From 47fb693992ffe452d87897e1c19e6d9a7ed00c04 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Fri, 19 Feb 2021 15:04:33 -0800 Subject: [PATCH 001/223] Add mod --- vAMPirus.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 4542133..8c7bbb9 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -2946,7 +2946,7 @@ if (params.Analyze) { if (params.ncASV) { - + // mopd this process Report_ASV { label 'norm_cpus' From e9eb02c55af18668aa020cd97df340471d2e6136 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Wed, 24 Feb 2021 09:48:46 -0800 Subject: [PATCH 002/223] Restructure processes. Use cache from DataCheck in Analyze mode --- vAMPirus.nf | 4478 ++++++++++++++++++++++++--------------------------- 1 file changed, 2130 insertions(+), 2348 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 8c7bbb9..a849587 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -471,7 +471,7 @@ if (params.readsTest) { .into{ reads_ch; reads_qc_ch } } -if (params.Analyze) { +if (params.DataCheck || params.Analyze) { println("\n\tRunning vAMPirus Analyze pipeline - This might take a while, check out Nextflow tower (tower.nf) to remotely monitor the run.\n") @@ -546,7 +546,7 @@ if (params.Analyze) { tuple sample_id, file(reads) from reads_qc_ch output: - tuple sample_id, file("*_fastqc.{zip,html}") into fastqc_results_OAS + tuple sample_id, file("*_fastqc.{zip,html}") into fastqc_results script: """ @@ -572,7 +572,7 @@ if (params.Analyze) { output: tuple sample_id, file("*.fastp.{json,html}") into fastp_results tuple sample_id, file("*.filter.fq") into reads_fastp_ch - file("*.csv") into fastp_csv + file("*.csv") into ( fastp_csv_in1, fastp_csv_in2 ) script: """ @@ -613,7 +613,7 @@ if (params.Analyze) { """ bbduk.sh in1=${reads[0]} out=${sample_id}_bb_R1.fastq.gz ftl=${params.defaultFwdTrim} t=${task.cpus} bbduk.sh in=${reads[1]} out=${sample_id}_bb_R2.fastq.gz ftl=${params.defaultRevTrim} t=${task.cpus} - repair.sh in1=${sample_id}_bb_R1.fastq.gz in2=${sample_id}_bb_R2.fastq.gz out1=${sample_id}_bbduk_R1.fastq.gz out2=${sample_id}_bbduk_R2.fastq.gz outs=sing.fq repair + repair.sh in1=${sample_id}_bb_R1.fastq.gz in2=${sample_id}_bb_R2.fastq.gz out1=${sample_id}_bbduk_R1.fastq.gz out2=${sample_id}_bbduk_R2.fastq.gz outs=sing.fq repair """ } else if ( params.GlobTrim && !params.GlobTrim == "" ) { """ @@ -621,7 +621,7 @@ if (params.Analyze) { RTRIM=\$( echo ${GlobTrim} | cut -f 2 -d "," ) bbduk.sh in=${reads[0]} out=${sample_id}_bb_R1.fastq.gz ftl=\${FTRIM} t=${task.cpus} bbduk.sh in=${reads[1]} out=${sample_id}_bb_R2.fastq.gz ftl=\${RTRIM} t=${task.cpus} - repair.sh in1=${sample_id}_bb_R1.fastq.gz in2=${sample_id}_bb_R2.fastq.gz out1=${sample_id}_bbduk_R1.fastq.gz out2=${sample_id}_bbduk_R2.fastq.gz outs=sing.fq repair + repair.sh in1=${sample_id}_bb_R1.fastq.gz in2=${sample_id}_bb_R2.fastq.gz out1=${sample_id}_bbduk_R1.fastq.gz out2=${sample_id}_bbduk_R2.fastq.gz outs=sing.fq repair """ } else if ( params.multi && params.primers ) { """ @@ -652,7 +652,7 @@ if (params.Analyze) { tuple sample_id, file(reads) from readsforqc2 output: - tuple sample_id, file("*_fastqc.{zip,html}") into fastqc2_results_OAS + tuple sample_id, file("*_fastqc.{zip,html}") into fastqc2_results script: """ @@ -726,6 +726,7 @@ if (params.Analyze) { } + /* BEfore examples process Length_Filtering { //changed label 'norm_cpus' @@ -752,6 +753,73 @@ if (params.Analyze) { bbduk.sh in=${params.projtag}_merged_preFilt_clean.fastq out=${params.projtag}_merged_clean_Lengthfiltered.fastq minlength=${params.maxLen} maxlength=${params.maxLen} t=${task.cpus} bbduk.sh in=${params.projtag}_merged_clean_Lengthfiltered.fastq bhist=${params.projtag}_all_merged_postFilt_baseFrequency_hist.txt qhist=${params.projtag}_all_merged_postFilt_qualityScore_hist.txt gchist=${params.projtag}_all_merged_postFilt_gcContent_hist.txt aqhist=${params.projtag}_all_merged_postFilt_averageQuaulity_hist.txt lhist=${params.projtag}_all_merged_postFilt_length_hist.txt gcbins=auto """ + }*/ + + process Length_Filtering { + + label 'norm_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging/LengthFiltering", mode: "copy", overwrite: true, pattern: "*_merged_preFilt*.fasta" + publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging", mode: "copy", overwrite: true, pattern: "*Lengthfiltered.fastq" + publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging/Histograms/pre_length_filtering", mode: "copy", overwrite: true, pattern: "*preFilt_*st.txt" + publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging/Histograms/post_length_filtering", mode: "copy", overwrite: true, pattern: "*postFilt_*st.txt" + + input: + file(reads) from collect_samples_ch + + output: + file("*_merged_preFilt_clean.fastq") into ( mergeforprotcounts, mergeforpcASVaacounts ) + + file("*_merged_preFilt_clean.fasta") into ( nuclCounts_mergedreads_ch, pcASV_mergedreads_ch ) + file("*_merged_clean_Lengthfiltered.fastq") into reads_vsearch2_ch + + file("*preFilt_preClean_baseFrequency_hist.csv") into prefilt_basefreq + file("*preFilt_preClean_qualityScore_hist.csv") into prefilt_qualityscore + file("*preFilt_preClean_gcContent_hist.csv") into prefilt_gccontent + file("*preFilt_preClean_averageQuality_hist.csv") into prefilt_averagequality + file("*preFilt_preClean_length_hist.csv") into prefilt_length + + file("*postFilt_baseFrequency_hist.csv") into postFilt_basefreq + file("*postFilt_qualityScore_hist.csv") into postFilt_qualityscore + file("*postFilt_gcContent_hist.csv") into postFilt_gccontent + file("*postFilt_averageQuaulity_hist.csv") into postFilt_averagequality + file("*postFilt_length_hist.csv") into postFilt_length + + file("reads_per_sample_preFilt_preClean.csv") into reads_per_sample_preFilt + file("read_per_sample_postFilt_postClean.csv") into reads_per_sample_postFilt + + script: + """ + #bbduk.sh in=${reads} bhist=${params.projtag}_all_merged_preFilt_preClean_baseFrequency_hist.txt qhist=${params.projtag}_all_merged_preFilt_preClean_qualityScore_hist.txt gchist=${params.projtag}_all_merged_preFilt_preClean_gcContent_hist.txt aqhist=${params.projtag}_all_merged_preFilt_preClean_averageQuality_hist.txt lhist=${params.projtag}_all_merged_preFilt__preClean_length_hist.txt gcbins=auto + #fastp -i ${reads} -o ${params.projtag}_merged_preFilt_clean.fastq -b ${params.maxLen} -l ${params.minLen} --thread ${task.cpus} -n 1 + #reformat.sh in=${params.projtag}_merged_preFilt_clean.fastq out=${params.projtag}_merged_preFilt_clean.fasta t=${task.cpus} + #bbduk.sh in=${params.projtag}_merged_preFilt_clean.fastq out=${params.projtag}_merged_clean_Lengthfiltered.fastq minlength=${params.maxLen} maxlength=${params.maxLen} t=${task.cpus} + #bbduk.sh in=${params.projtag}_merged_clean_Lengthfiltered.fastq bhist=${params.projtag}_all_merged_postFilt_baseFrequency_hist.txt qhist=${params.projtag}_all_merged_postFilt_qualityScore_hist.txt gchist=${params.projtag}_all_merged_postFilt_gcContent_hist.txt aqhist=${params.projtag}_all_merged_postFilt_averageQuaulity_hist.txt lhist=${params.projtag}_all_merged_postFilt_length_hist.txt gcbins=auto + + # from DC + bbduk.sh in=${reads} bhist=${params.projtag}_all_merged_preFilt_preClean_baseFrequency_hist.txt qhist=${params.projtag}_all_merged_preFilt_preClean_qualityScore_hist.txt gchist=${params.projtag}_all_merged_preFilt_preClean_gcContent_hist.txt aqhist=${params.projtag}_all_merged_preFilt_preClean_averageQuality_hist.txt lhist=${params.projtag}_all_merged_preFilt_preClean_length_hist.txt gcbins=auto + for x in *preFilt*hist.txt;do + pre=\$(echo \$x | awk -F ".txt" '{print \$1}') + cat \$x | tr "\t" "," > \${pre}.csv + rm \$x + done + reformat.sh in=${reads} out=${params.projtag}_preFilt_preclean.fasta t=${task.cpus} + echo "sample,reads" >> reads_per_sample_preFilt_preClean.csv + grep ">" ${params.projtag}_preFilt_preclean.fasta | awk -F ">" '{print \$2}' | awk -F "." '{print \$1}' | sort --parallel=${task.cpus} | uniq -c | sort -brg --parallel=${task.cpus} | awk '{print \$2","\$1}' >> reads_per_sample_preFilt_preClean.csv + rm ${params.projtag}_preFilt_preclean.fasta + fastp -i ${reads} -o ${params.projtag}_merged_preFilt_clean.fastq -b ${params.maxLen} -l ${params.minLen} --thread ${task.cpus} -n 1 + reformat.sh in=${params.projtag}_merged_preFilt_clean.fastq out=${params.projtag}_merged_preFilt_clean.fasta t=${task.cpus} + bbduk.sh in=${params.projtag}_merged_preFilt_clean.fastq out=${params.projtag}_merged_clean_Lengthfiltered.fastq minlength=${params.maxLen} maxlength=${params.maxLen} t=${task.cpus} + bbduk.sh in=${params.projtag}_merged_clean_Lengthfiltered.fastq bhist=${params.projtag}_all_merged_postFilt_baseFrequency_hist.txt qhist=${params.projtag}_all_merged_postFilt_qualityScore_hist.txt gchist=${params.projtag}_all_merged_postFilt_gcContent_hist.txt aqhist=${params.projtag}_all_merged_postFilt_averageQuaulity_hist.txt lhist=${params.projtag}_all_merged_postFilt_length_hist.txt gcbins=auto + for x in *postFilt*hist.txt;do + pre=\$(echo \$x | awk -F ".txt" '{print \$1}') + cat \$x | tr "\t" "," > \${pre}.csv + rm \$x + done + reformat.sh in=${params.projtag}_merged_clean_Lengthfiltered.fastq out=${params.projtag}_merged_clean_Lengthfiltered.fasta t=${task.cpus} + echo "sample,reads" >> read_per_sample_postFilt_postClean.csv + grep ">" ${params.projtag}_merged_clean_Lengthfiltered.fasta | awk -F ">" '{print \$2}' | awk -F "." '{print \$1}' | sort --parallel=${task.cpus} | uniq -c | sort -brg --parallel=${task.cpus} | awk '{print \$2","\$1}' >> read_per_sample_postFilt_postClean.csv + """ } process Extract_Uniques { @@ -810,351 +878,330 @@ if (params.Analyze) { // UNTIL HERE DEFAULT - if (params.ncASV) { + if (params.DataCheck) { + + process NucleotideBased_ASV_clustering_DC { + + label 'norm_cpus' + + publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Nucleotide", mode: "copy", overwrite: true, pattern: '*{.csv}' + + input: + file(fasta) from reads_vsearch5_ch + + output: + file("number_per_percentage_nucl.csv") into number_per_percent_nucl_plot + + script: + if (params.datacheckntIDlist) { + """ + for id in `echo ${params.datacheckntIDlist} | tr "," "\\n"`;do + vsearch --cluster_fast ${fasta} --centroids ${params.projtag}_ncASV\${id}.fasta --threads ${task.cpus} --relabel OTU --id \${id} + done + for x in *ncASV*.fasta;do + id=\$( echo \$x | awk -F "_ncASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') + numb=\$( grep -c ">" \$x ) + echo "\${id},\${numb}" >> number_per_percentage_nucl.csv + done + yo=\$(grep -c ">" ${fasta}) + echo "1.0,\${yo}" >> number_per_percentage_nucl.csv + """ + } + } + + if (params.sing) { + + process Translating_For_ProteinClustering_DC { + + label 'low_cpus' + + publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Aminoacid/translation", mode: "copy", overwrite: true - process NucleotideBased_ASV_clustering { + input: + file(fasta) from nucl2aa + + output: + file("*ASVprotforclust.fasta") into clustering_aa + file("*_translation_report") into reportaa_VR + file("*_ASV_all.fasta") into asvfastaforaaclust + + script: + """ + conda init && source activate virtualribosome + + ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_ASVprotforclust.fasta --report ${params.projtag}_translation_report + cp ${fasta} ${params.projtag}_ASV_all.fasta + + """ + + } + + } else { + + process Translation_For_ProteinBased_Clustering_DC { label 'norm_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/ncASV", mode: "copy", overwrite: true, pattern: '*ncASV*.fasta' + conda 'python=2.7' + + publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Aminoacid/translation", mode: "copy", overwrite: true input: - file(fasta) from reads_vsearch5_ch + file(fasta) from nucl2aa output: - tuple file("*_ncASV*.fasta"), file("*ASV.fasta") into ( nuclFastas_forDiamond_ch, nuclFastas_forCounts_ch, nuclFastas_forMatrix_ch) - tuple file("*_ncASV*.fasta"), file("*ASV.fasta") into nuclFastas_forphylogeny + file("*ASVprotforclust.fasta") into clustering_aa + file("*_translation_report") into reportaa_VR + file("*_ASV_all.fasta") into asvfastaforaaclust script: - if (params.clusterNuclIDlist) { """ - cp ${fasta} ./${params.projtag}_ASV.fasta - for id in `echo ${params.clusterNuclIDlist} | tr "," "\\n"`;do - vsearch --cluster_fast ${params.projtag}_ASV.fasta --centroids ${params.projtag}_ncASV\${id}.fasta --threads ${task.cpus} --relabel ncASV --id \${id} - done - """ - } else if (params.clusterNuclID) { + ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_ASVprotforclust.fasta --report ${params.projtag}_translation_report + cp ${fasta} ${params.projtag}_ASV_all.fasta """ - cp ${fasta} ./${params.projtag}_ASV.fasta - id=${params.clusterNuclID} - vsearch --cluster_fast ${params.projtag}_ASV.fasta --centroids ${params.projtag}_ncASV\${id}.fasta --threads ${task.cpus} --relabel ncASV --id \${id} - """ - } - } - } else { - reads_vsearch5_ch - .into{ nuclFastas_forDiamond_ch; nuclFastas_forCounts_ch; nuclFastas_forphylogeny; nuclFastas_forMatrix_ch } - } - - if (!params.skipTaxonomy) { - - if (params.ncASV) { + } + } - process Nucleotide_Taxonomy_Inference { + process Protein_clustering_DC { - label 'high_cpus' + label 'norm_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy", mode: "copy", overwrite: true, pattern: '*_ASV*.{fasta,csv,tsv}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Taxonomy", mode: "copy", overwrite: true, pattern: '*ncASV*.{fasta,csv,tsv}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*_ASV*dmd.out' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*ncASV*dmd.out' + publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Aminoacid", mode: "copy", overwrite: true, pattern: '*{.csv}' input: - tuple file(notus), file(asvs) from nuclFastas_forDiamond_ch + file(fasta) from clustering_aa + file(asvs) from asvfastaforaaclust output: - file("*.fasta") into tax_labeled_fasta - tuple file("*_phyloseqObject.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond - file("*ncASV*summary_for_plot.csv") into taxplot1a - file("*_ASV*_summary_for_plot.csv") into taxplot1 + file("number_per_percentage_prot.csv") into number_per_percent_prot_plot script: + // add awk script to count seqs """ + set +e cp ${params.vampdir}/bin/rename_seq.py . - virdb=${params.dbdir}/${params.dbname} - grep ">" \${virdb} > headers.list - headers="headers.list" - for filename in ${notus};do - name=\$(ls \${filename} | awk -F ".fasta" '{print \$1}') - diamond blastx -q \${filename} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 - echo "Preparing lists to generate summary .csv's" - echo "[Best hit accession number]" > access.list - echo "[e-value]" > evalue.list - echo "[Bitscore]" > bit.list - echo "[Percent ID (aa)]" > pid.list - echo "[Organism ID]" > "\$name"_virus.list - echo "[Gene]" > "\$name"_genes.list - grep ">" \${filename} | awk -F ">" '{print \$2}' > seqids.lst - echo "extracting genes and names" - touch new_"\$name"_asvnames.txt - j=1 - if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then - echo "[ncASV#]" > otu.list - echo "[ncASV sequence length]" > length.list - for s in \$(cat seqids.lst);do - echo "Checking for \$s hit in diamond output" - if [[ ${params.refseq} == "T" ]];then - echo "RefSeq headers specified" - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">ncASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list - virus="NO" - gene="HIT" - echo ">ncASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - fi - else - echo "Using RVDB headers." - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">ncASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list - virus="NO" - gene="HIT" - echo ">ncASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - fi - fi - echo "Done with \$s" - done + for id in `echo ${params.datacheckaaIDlist} | tr "," "\\n"`;do + if [ \${id} == ".55" ];then + word=3 + elif [ \${id} == ".65" ];then + word=4 + else + word=5 fi - echo "Now editing "\$name" fasta headers" - ###### rename_seq.py - ./rename_seq.py \${filename} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta - awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta - echo "[Sequence header]" > newnames.list - cat new_"\$name"_asvnames.txt >> newnames.list - touch sequence.list - echo " " > sequence.list - grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list - rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - for x in *phyloseqObject.csv;do - echo "\$x" - lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) - tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; - sed 's/ /_/g' tmpcol.list > tmp2col.list; - cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; - rm tmpcol.list tmp2col.list + awk 'BEGIN{RS=">";ORS=""}length(\$2)>="${params.minAA}"{print ">"\$0}' ${fasta} > ${params.projtag}_filtered_proteins.fasta + cd-hit -i ${params.projtag}_filtered_proteins.fasta -n \${word} -c \${id} -o ${params.projtag}_pcASV\${id}.fasta + sed 's/>Cluster />Cluster_/g' ${params.projtag}_pcASV\${id}.fasta.clstr >${params.projtag}_pcASV\${id}.clstr + grep ">Cluster_" ${params.projtag}_pcASV\${id}.clstr >temporaryclusters.list + y=\$(grep -c ">Cluster_" ${params.projtag}_pcASV\${id}.clstr) + echo ">Cluster_"\${y}"" >> ${params.projtag}_pcASV\${id}.clstr + t=1 + b=1 + for x in \$(cat temporaryclusters.list);do + echo "Extracting \$x" + name="\$( echo \$x | awk -F ">" '{print \$2}')" + clust="pcASV"\${t}"" + echo "\${name}" + awk '/^>'\${name}'\$/,/^>Cluster_'\${b}'\$/' ${params.projtag}_pcASV\${id}.clstr > "\${name}"_"\${clust}"_tmp.list + t=\$(( \${t}+1 )) + b=\$(( \${b}+1 )) done - rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; - rm *asvnames.txt - rm "\$name"_virus.list - rm "\$name"_genes.list - rm newnames.list - rm access.list - echo "Taxonomy inferred for: \${filename} " - done - for filename in ${asvs};do - name=\$(ls \${filename} | awk -F ".fasta" '{print \$1}') - diamond blastx -q \${filename} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 - echo "Preparing lists to generate summary .csv's" - echo "[Best hit accession number]" > access.list - echo "[e-value]" > evalue.list - echo "[Bitscore]" > bit.list - echo "[Percent ID (aa)]" > pid.list - echo "[Organism ID]" > "\$name"_virus.list - echo "[Gene]" > "\$name"_genes.list - grep ">" \${filename} | awk -F ">" '{print \$2}' > seqids.lst - echo "extracting genes and names" - touch new_"\$name"_asvnames.txt - j=1 - if [ `echo \${filename} | grep -c "ASV"` -eq 1 ];then - for s in \$(cat seqids.lst);do - echo "[ASV#]" > otu.list - echo "[ASV sequence length]" > length.list - echo "Checking for \$s hit in diamond output" - if [[ ${params.refseq} == "T" ]];then - echo "RefSeq headers specified" - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list - virus="NO" - gene="HIT" - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - fi - else - echo "Using RVDB headers." - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list - virus="NO" - gene="HIT" - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - fi - fi - echo "Done with \$s" + ls *_tmp.list + u=1 + for x in *_tmp.list;do + name="\$(echo \$x | awk -F "_p" '{print \$1}')" + echo "\${name}" + cluster="\$(echo \$x | awk -F "_" '{print \$3}')" + echo "\${cluster}" + grep "ASV" \$x | awk -F ", " '{print \$2}' | awk -F "_" '{print \$1}' | awk -F ">" '{print \$2}' > \${name}_\${cluster}_seqs_tmps.list + seqtk subseq ${asvs} \${name}_\${cluster}_seqs_tmps.list > \${name}_\${cluster}_nucleotide_sequences.fasta + vsearch --cluster_fast \${name}_\${cluster}_nucleotide_sequences.fasta --id 0.2 --centroids \${name}_\${cluster}_centroids.fasta + grep ">" \${name}_\${cluster}_centroids.fasta >> \${name}_\${cluster}_tmp_centroids.list + for y in \$( cat \${name}_\${cluster}_tmp_centroids.list );do + echo ">\${cluster}_type"\$u"" >> \${name}_\${cluster}_tmp_centroid.newheaders + u=\$(( \${u}+1 )) done - fi - echo "Now editing "\$name" fasta headers" - ###### rename_seq.py - ./rename_seq.py \${filename} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta - awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta - echo "[Sequence header]" > newnames.list - cat new_"\$name"_asvnames.txt >> newnames.list - touch sequence.list - echo " " > sequence.list - grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list - rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - for x in *phyloseqObject.csv;do - echo "\$x" - lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) - tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; - sed 's/ /_/g' tmpcol.list > tmp2col.list; - cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; - rm tmpcol.list tmp2col.list + u=1 + ./rename_seq.py \${name}_\${cluster}_centroids.fasta \${name}_\${cluster}_tmp_centroid.newheaders \${cluster}_types_labeled.fasta + done + cat *_types_labeled.fasta >> ${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta + grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$3}' | awk -F "." '{print \$1}' >tmphead.list + grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$2}' | awk -F "," '{print \$1}' >tmplen.list + paste -d"," temporaryclusters.list tmphead.list >tmp.info.csv + grep ">" ${params.projtag}_pcASV\${id}.fasta >lala.list + j=1 + for x in \$(cat lala.list);do + echo ">${params.projtag}_pcASV\${j}" >>${params.projtag}_aminoheaders.list + echo "\${x},>${params.projtag}_pcASV\${j}" >>tmpaminotype.info.csv + j=\$(( \${j}+1 )) + done + rm lala.list + awk -F "," '{print \$2}' tmp.info.csv >>tmporder.list + for x in \$(cat tmporder.list);do + grep -w "\$x" tmpaminotype.info.csv | awk -F "," '{print \$2}' >>tmpder.list done - rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; - rm *asvnames.txt - rm "\$name"_virus.list - rm "\$name"_genes.list - rm newnames.list - rm access.list - echo "Taxonomy inferred for: \${filename} " + paste -d "," temporaryclusters.list tmplen.list tmphead.list tmpder.list >${params.projtag}_pcASVCluster\${id}_summary.csv + ./rename_seq.py ${params.projtag}_pcASV\${id}.fasta ${params.projtag}_aminoheaders.list ${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta + stats.sh in=${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_aminoacid_clustered.gc gcformat=4 overwrite=true + stats.sh in=${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_nucleotide_clustered.gc gcformat=4 overwrite=true + awk 'BEGIN{RS=">";ORS=""}length(\$2)<"${params.minAA}"{print ">"\$0}' ${fasta} >${params.projtag}_pcASV\${id}_problematic_translations.fasta + if [ `wc -l ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk '{print \$1}'` -gt 1 ];then + grep ">" ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk -F ">" '{print \$2}' > problem_tmp.list + seqtk subseq ${asvs} problem_tmp.list > ${params.projtag}_pcASV\${id}_problematic_nucleotides.fasta + else + rm ${params.projtag}_pcASV\${id}_problematic_translations.fasta + fi + rm *.list + rm Cluster* + rm *types* + rm *tmp* + rm ${params.projtag}_pcASV\${id}.fast* + done + for x in *aminoacid*noTaxonomy.fasta;do + id=\$( echo \$x | awk -F "_noTax" '{print \$1}' | awk -F "pcASV" '{print \$2}') + numb=\$( grep -c ">" \$x) + echo "\${id},\${numb}" >> number_per_percentage_protz.csv + done + yesirr=\$( wc -l number_per_percentage_protz.csv | awk '{print \$1}') + tail -\$(( \${yesirr}-1 )) number_per_percentage_protz.csv > number_per_percentage_prot.csv + head -1 number_per_percentage_protz.csv >> number_per_percentage_prot.csv + rm number_per_percentage_protz.csv + """ + } + + process combine_csv_DC { + + input: + file(csv) from fastp_csv_in1 + .collect() + + output: + file("final_reads_stats.csv") into fastp_csv_dc + + script: + """ + cat ${csv} >all_reads_stats.csv + head -n1 all_reads_stats.csv >tmp.names.csv + cat all_reads_stats.csv | grep -v ""Sample,Total_"" >tmp.reads.stats.csv + cat tmp.names.csv tmp.reads.stats.csv >final_reads_stats.csv + rm tmp.names.csv tmp.reads.stats.csv + """ + + } + + process Report_DataCheck { + + label 'norm_cpus' + + publishDir "${params.workingdir}/${params.outdir}/DataCheck/Report", mode: "copy", overwrite: true, pattern: '*.{html}' + + input: + file(fastpcsv) from fastp_csv_dc + file(reads_per_sample_preFilt) from reads_per_sample_preFilt + file(read_per_sample_postFilt) from reads_per_sample_postFilt + file(preFilt_baseFrequency) from prefilt_basefreq + file(postFilt_baseFrequency) from postFilt_basefreq + file(preFilt_qualityScore) from prefilt_qualityscore + file(postFilt_qualityScore) from postFilt_qualityscore + file(preFilt_gcContent) from prefilt_gccontent + file(postFilt_gcContent) from postFilt_gccontent + file(preFilt_averageQuality) from prefilt_averagequality + file(postFilt_averageQuaulity) from postFilt_averagequality + file(preFilt_length) from prefilt_length + file(postFilt_length) from postFilt_length + file(number_per_percentage_nucl) from number_per_percent_nucl_plot + file(number_per_percentage_prot) from number_per_percent_prot_plot + + output: + file("*.html") into datacheckreport + + script: + """ + cp ${params.vampdir}/bin/vAMPirus_DC_Report.Rmd . + cp ${params.vampdir}/example_data/conf/vamplogo.png . + Rscript -e "rmarkdown::render('vAMPirus_DC_Report.Rmd',output_file='${params.projtag}_DataCheck_Report.html')" ${params.projtag} \ + ${fastpcsv} \ + ${reads_per_sample_preFilt} \ + ${read_per_sample_postFilt} \ + ${preFilt_baseFrequency} \ + ${postFilt_baseFrequency} \ + ${preFilt_qualityScore} \ + ${postFilt_qualityScore} \ + ${preFilt_averageQuality} \ + ${postFilt_averageQuaulity} \ + ${preFilt_length} \ + ${postFilt_length} \ + ${number_per_percentage_nucl} \ + ${number_per_percentage_prot} + """ + } + + } else if (params.Analyze) { + + if (params.ncASV) { + + process NucleotideBased_ASV_clustering { + + label 'norm_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/ncASV", mode: "copy", overwrite: true, pattern: '*ncASV*.fasta' + + input: + file(fasta) from reads_vsearch5_ch + + output: + tuple file("*_ncASV*.fasta"), file("*ASV.fasta") into ( nuclFastas_forDiamond_ch, nuclFastas_forCounts_ch, nuclFastas_forMatrix_ch) + tuple file("*_ncASV*.fasta"), file("*ASV.fasta") into nuclFastas_forphylogeny + + script: + if (params.clusterNuclIDlist) { + """ + cp ${fasta} ./${params.projtag}_ASV.fasta + for id in `echo ${params.clusterNuclIDlist} | tr "," "\\n"`;do + vsearch --cluster_fast ${params.projtag}_ASV.fasta --centroids ${params.projtag}_ncASV\${id}.fasta --threads ${task.cpus} --relabel ncASV --id \${id} done - rm headers.list + """ + } else if (params.clusterNuclID) { + """ + cp ${fasta} ./${params.projtag}_ASV.fasta + id=${params.clusterNuclID} + vsearch --cluster_fast ${params.projtag}_ASV.fasta --centroids ${params.projtag}_ncASV\${id}.fasta --threads ${task.cpus} --relabel ncASV --id \${id} """ } + } + } else { + reads_vsearch5_ch + .into{ nuclFastas_forDiamond_ch; nuclFastas_forCounts_ch; nuclFastas_forphylogeny; nuclFastas_forMatrix_ch } + } - } else { + if (!params.skipTaxonomy) { - process ASV_Taxonomy_Inference { + if (params.ncASV) { + + process Nucleotide_Taxonomy_Inference { label 'high_cpus' publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy", mode: "copy", overwrite: true, pattern: '*_ASV*.{fasta,csv,tsv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Taxonomy", mode: "copy", overwrite: true, pattern: '*ncASV*.{fasta,csv,tsv}' publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*_ASV*dmd.out' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*ncASV*dmd.out' input: - file(reads) from nuclFastas_forDiamond_ch + tuple file(notus), file(asvs) from nuclFastas_forDiamond_ch output: file("*.fasta") into tax_labeled_fasta tuple file("*_phyloseqObject.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond + file("*ncASV*summary_for_plot.csv") into taxplot1a file("*_ASV*_summary_for_plot.csv") into taxplot1 + script: """ cp ${params.vampdir}/bin/rename_seq.py . virdb=${params.dbdir}/${params.dbname} grep ">" \${virdb} > headers.list headers="headers.list" - for filename in ${reads};do + for filename in ${notus};do name=\$(ls \${filename} | awk -F ".fasta" '{print \$1}') diamond blastx -q \${filename} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 echo "Preparing lists to generate summary .csv's" @@ -1169,8 +1216,8 @@ if (params.Analyze) { touch new_"\$name"_asvnames.txt j=1 if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then - echo "[ASV#]" > otu.list - echo "[ASV sequence length]" > length.list + echo "[ncASV#]" > otu.list + echo "[ncASV sequence length]" > length.list for s in \$(cat seqids.lst);do echo "Checking for \$s hit in diamond output" if [[ ${params.refseq} == "T" ]];then @@ -1191,7 +1238,7 @@ if (params.Analyze) { echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + echo ">ncASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt j=\$((\$j+1)) echo "\$s done." else @@ -1207,7 +1254,7 @@ if (params.Analyze) { echo "NO_HIT" >> length.list virus="NO" gene="HIT" - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + echo ">ncASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt j=\$((\$j+1)) echo "\$s done." fi @@ -1229,7 +1276,7 @@ if (params.Analyze) { echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + echo ">ncASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt j=\$((\$j+1)) echo "\$s done." else @@ -1245,25 +1292,68 @@ if (params.Analyze) { echo "NO_HIT" >> length.list virus="NO" gene="HIT" - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + echo ">ncASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt j=\$((\$j+1)) echo "\$s done." fi fi echo "Done with \$s" - done - else - for s in \$(cat seqids.lst);do - echo "[ASV#]" > otu.list - echo "[ASV sequence length]" > length.list - echo "Checking for \$s hit in diamond output" - if [[ ${params.refseq} == "T" ]];then - echo "RefSeq headers specified" - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') - echo "\$s" >> otu.list + done + fi + echo "Now editing "\$name" fasta headers" + ###### rename_seq.py + ./rename_seq.py \${filename} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta + echo "[Sequence header]" > newnames.list + cat new_"\$name"_asvnames.txt >> newnames.list + touch sequence.list + echo " " > sequence.list + grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list + rm "\$name"_tmpssasv.fasta + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + for x in *phyloseqObject.csv;do + echo "\$x" + lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) + tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; + sed 's/ /_/g' tmpcol.list > tmp2col.list; + cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; + rm tmpcol.list tmp2col.list + done + rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; + rm *asvnames.txt + rm "\$name"_virus.list + rm "\$name"_genes.list + rm newnames.list + rm access.list + echo "Taxonomy inferred for: \${filename} " + done + for filename in ${asvs};do + name=\$(ls \${filename} | awk -F ".fasta" '{print \$1}') + diamond blastx -q \${filename} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + echo "Preparing lists to generate summary .csv's" + echo "[Best hit accession number]" > access.list + echo "[e-value]" > evalue.list + echo "[Bitscore]" > bit.list + echo "[Percent ID (aa)]" > pid.list + echo "[Organism ID]" > "\$name"_virus.list + echo "[Gene]" > "\$name"_genes.list + grep ">" \${filename} | awk -F ">" '{print \$2}' > seqids.lst + echo "extracting genes and names" + touch new_"\$name"_asvnames.txt + j=1 + if [ `echo \${filename} | grep -c "ASV"` -eq 1 ];then + for s in \$(cat seqids.lst);do + echo "[ASV#]" > otu.list + echo "[ASV sequence length]" > length.list + echo "Checking for \$s hit in diamond output" + if [[ ${params.refseq} == "T" ]];then + echo "RefSeq headers specified" + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') + echo "\$s" >> otu.list echo "\$acc" >> access.list line="\$(grep -w "\$s" "\$name"_dmd.out)" echo "\$line" | awk '{print \$10}' >> evalue.list @@ -1334,7 +1424,7 @@ if (params.Analyze) { echo "\$s done." fi fi - echo "Done with \$s" + echo "Done with \$s" done fi echo "Now editing "\$name" fasta headers" @@ -1367,811 +1457,1121 @@ if (params.Analyze) { done rm headers.list """ - } - } - } - - if (params.ncASV) { - - process Generate_Counts_Tables_Nucleotide { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Counts", mode: "copy", overwrite: true, pattern: '*_ASV*.{biome,csv}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Counts", mode: "copy", overwrite: true, pattern: '*ncASV*.{biome,csv}' - - input: - tuple file(notus), file(asvs) from nuclFastas_forCounts_ch - file(merged) from nuclCounts_mergedreads_ch - - output: - tuple file("*_counts.csv"), file("*_counts.biome") into counts_vsearch - file("*ncASV*counts.csv") into notu_counts_plots - file("*_ASV*counts.csv") into asv_counts_plots + } - script: - """ - for filename in ${notus};do - if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then - ident=\$( echo \${filename} | awk -F "ncASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') - name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') - vsearch --usearch_global ${merged} --db \${filename} --id \${ident} --threads ${task.cpus} --otutabout \${name}_counts.txt --biomout \${name}_counts.biome - cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv - sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv - rm \${name}_count.csv - fi - done - if [ `echo ${asvs} | grep -c "ASV"` -eq 1 ];then - name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - vsearch --usearch_global ${merged} --db ${asvs} --id ${params.asvcountID} --threads ${task.cpus} --otutabout "\$name"_counts.txt --biomout "\$name"_counts.biome - cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv - sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv - rm \${name}_count.csv - fi - """ - } - } else { - process Generate_ASV_Counts_Tables { + } else { - label 'norm_cpus' + process ASV_Taxonomy_Inference { - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Counts", mode: "copy", overwrite: true, pattern: '*ASV*.{biome,csv}' + label 'high_cpus' - input: - file(asvs) from nuclFastas_forCounts_ch - file(merged) from nuclCounts_mergedreads_ch + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy", mode: "copy", overwrite: true, pattern: '*_ASV*.{fasta,csv,tsv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*_ASV*dmd.out' - output: - tuple file("*_counts.csv"), file("*_counts.biome") into counts_vsearch - file("*_ASV*counts.csv") into asv_counts_plots + input: + file(reads) from nuclFastas_forDiamond_ch - script: - """ - if [ `echo ${asvs} | grep -c "ASV"` -eq 1 ];then - name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - vsearch --usearch_global ${merged} --db ${asvs} --id ${params.asvcountID} --threads ${task.cpus} --otutabout "\$name"_counts.txt --biomout "\$name"_counts.biome - cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv - sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv - rm \${name}_count.csv - fi - """ + output: + file("*.fasta") into tax_labeled_fasta + tuple file("*_phyloseqObject.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond + file("*_ASV*_summary_for_plot.csv") into taxplot1 + script: + """ + cp ${params.vampdir}/bin/rename_seq.py . + virdb=${params.dbdir}/${params.dbname} + grep ">" \${virdb} > headers.list + headers="headers.list" + for filename in ${reads};do + name=\$(ls \${filename} | awk -F ".fasta" '{print \$1}') + diamond blastx -q \${filename} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + echo "Preparing lists to generate summary .csv's" + echo "[Best hit accession number]" > access.list + echo "[e-value]" > evalue.list + echo "[Bitscore]" > bit.list + echo "[Percent ID (aa)]" > pid.list + echo "[Organism ID]" > "\$name"_virus.list + echo "[Gene]" > "\$name"_genes.list + grep ">" \${filename} | awk -F ">" '{print \$2}' > seqids.lst + echo "extracting genes and names" + touch new_"\$name"_asvnames.txt + j=1 + if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then + echo "[ASV#]" > otu.list + echo "[ASV sequence length]" > length.list + for s in \$(cat seqids.lst);do + echo "Checking for \$s hit in diamond output" + if [[ ${params.refseq} == "T" ]];then + echo "RefSeq headers specified" + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list + virus="NO" + gene="HIT" + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + fi + else + echo "Using RVDB headers." + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list + virus="NO" + gene="HIT" + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + fi + fi + echo "Done with \$s" + done + else + for s in \$(cat seqids.lst);do + echo "[ASV#]" > otu.list + echo "[ASV sequence length]" > length.list + echo "Checking for \$s hit in diamond output" + if [[ ${params.refseq} == "T" ]];then + echo "RefSeq headers specified" + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list + virus="NO" + gene="HIT" + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + fi + else + echo "Using RVDB headers." + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list + virus="NO" + gene="HIT" + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + fi + fi + echo "Done with \$s" + done + fi + echo "Now editing "\$name" fasta headers" + ###### rename_seq.py + ./rename_seq.py \${filename} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta + echo "[Sequence header]" > newnames.list + cat new_"\$name"_asvnames.txt >> newnames.list + touch sequence.list + echo " " > sequence.list + grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list + rm "\$name"_tmpssasv.fasta + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + for x in *phyloseqObject.csv;do + echo "\$x" + lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) + tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; + sed 's/ /_/g' tmpcol.list > tmp2col.list; + cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; + rm tmpcol.list tmp2col.list + done + rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; + rm *asvnames.txt + rm "\$name"_virus.list + rm "\$name"_genes.list + rm newnames.list + rm access.list + echo "Taxonomy inferred for: \${filename} " + done + rm headers.list + """ + } } } - if (params.ncASV) { + if (params.ncASV) { - process Generate_Nucleotide_Matrix { + process Generate_Counts_Tables_Nucleotide { - label 'low_cpus' + label 'norm_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Matrix", mode: "copy", overwrite: true, pattern: '*_ASV*PercentID.matrix' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Matrix", mode: "copy", overwrite: true, pattern: '*ncASV*PercentID.matrix' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Counts", mode: "copy", overwrite: true, pattern: '*_ASV*.{biome,csv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Counts", mode: "copy", overwrite: true, pattern: '*ncASV*.{biome,csv}' - input: - tuple file(notus), file(asvs) from nuclFastas_forMatrix_ch + input: + tuple file(notus), file(asvs) from nuclFastas_forCounts_ch + file(merged) from nuclCounts_mergedreads_ch - output: - file("*.matrix") into clustmatrices - file("*ncASV*PercentID.matrix") into notu_heatmap - file("*_ASV*PercentID.matrix") into asv_heatmap + output: + tuple file("*_counts.csv"), file("*_counts.biome") into counts_vsearch + file("*ncASV*counts.csv") into notu_counts_plots + file("*_ASV*counts.csv") into asv_counts_plots - script: - // remove if statement later (no fin) - """ - for filename in ${notus};do - if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then - ident=\$( echo \${filename} | awk -F "ncASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') - name=\$( echo \${filename}| awk -F ".fasta" '{print \$1}') - clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} - clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} - for x in *q.matrix;do - pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') - ya=\$(wc -l \$x | awk '{print \$1}') - echo "\$((\$ya-1))" - tail -"\$((\$ya-1))" \$x > \${pre}z.matrix - rm \$x - cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix - rm \${pre}z.matrix + script: + """ + for filename in ${notus};do + if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then + ident=\$( echo \${filename} | awk -F "ncASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') + name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') + vsearch --usearch_global ${merged} --db \${filename} --id \${ident} --threads ${task.cpus} --otutabout \${name}_counts.txt --biomout \${name}_counts.biome + cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv + sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv + rm \${name}_count.csv + fi done - fi - done - if [ `echo ${asvs} | grep -c "_ASV"` -eq 1 ];then - name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - clustalo -i ${asvs} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} - clustalo -i ${asvs} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} - for x in *q.matrix;do - pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') - ya=\$(wc -l \$x | awk '{print \$1}') - echo "\$((\$ya-1))" - tail -"\$((\$ya-1))" \$x > \${pre}z.matrix - rm \$x - cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix - rm \${pre}z.matrix - done - fi - """ - } - } else { - - process Generate_ASV_Matrix { - - label 'low_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Matrix", mode: "copy", overwrite: true, pattern: '*ASV*PercentID.matrix' - - input: - file(reads) from nuclFastas_forMatrix_ch - - output: - file("*.matrix") into clustmatrices - file("*_ASV*PercentID.matrix") into asv_heatmap - - script: - // remove if statement later (no fin) - """ - for filename in ${reads};do - if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then - ident=\$( echo \${filename} | awk -F "ncASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') - name=\$( echo \${filename}| awk -F ".fasta" '{print \$1}') - clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} - clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} - for x in *q.matrix;do - pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') - ya=\$(wc -l \$x | awk '{print \$1}') - echo "\$((\$ya-1))" - tail -"\$((\$ya-1))" \$x > \${pre}z.matrix - rm \$x - cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix - rm \${pre}z.matrix - done - else - name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') - clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} - clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} - for x in *q.matrix;do - pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') - ya=\$(wc -l \$x | awk '{print \$1}') - echo "\$((\$ya-1))" - tail -"\$((\$ya-1))" \$x > \${pre}z.matrix - rm \$x - cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix - rm \${pre}z.matrix - done + if [ `echo ${asvs} | grep -c "ASV"` -eq 1 ];then + name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') + vsearch --usearch_global ${merged} --db ${asvs} --id ${params.asvcountID} --threads ${task.cpus} --otutabout "\$name"_counts.txt --biomout "\$name"_counts.biome + cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv + sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv + rm \${name}_count.csv fi - done - """ + """ } - } - - if (!params.skipPhylogeny) { // need to edit paths - - if (params.ncASV) { - - process Nucleotide_Phylogeny { + } else { + process Generate_ASV_Counts_Tables { label 'norm_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*ncASV*aln.*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*ncASV*mt*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*ncASV*iq*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*_ASV*aln.*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*_ASV*mt*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*_ASV*iq*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Counts", mode: "copy", overwrite: true, pattern: '*ASV*.{biome,csv}' input: - tuple file(notus), file(asvs) from nuclFastas_forphylogeny + file(asvs) from nuclFastas_forCounts_ch + file(merged) from nuclCounts_mergedreads_ch output: - tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into align_results - file("*iq.treefile") into nucl_phyl_plot + tuple file("*_counts.csv"), file("*_counts.biome") into counts_vsearch + file("*_ASV*counts.csv") into asv_counts_plots script: """ - for filename in ${notus};do - pre=\$(echo \${filename} | awk -F ".fasta" '{print \$1}' ) - mafft --thread ${task.cpus} --maxiterate 15000 --auto \${filename} >\${pre}_ALN.fasta - trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html - # Nucleotide_ModelTest - modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint - # Nucleotide_Phylogeny - if [ "${params.iqCustomnt}" != "" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -t \${pre}_mt.tree -T auto ${params.iqCustomnt} - elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} - elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni - elif [ "${params.nonparametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} - elif [ "${params.parametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni - else - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni - fi - done - for filename in ${asvs};do - pre=\$(echo \${filename} | awk -F ".fasta" '{print \$1}' ) - mafft --thread ${task.cpus} --maxiterate 15000 --auto \${filename} >\${pre}_ALN.fasta - trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html - # Nucleotide_ModelTest - modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint - # Nucleotide_Phylogeny - if [ "${params.iqCustomnt}" != "" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomnt} - elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} - elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni - elif [ "${params.nonparametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} - elif [ "${params.parametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - else - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - fi - done + if [ `echo ${asvs} | grep -c "ASV"` -eq 1 ];then + name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') + vsearch --usearch_global ${merged} --db ${asvs} --id ${params.asvcountID} --threads ${task.cpus} --otutabout "\$name"_counts.txt --biomout "\$name"_counts.biome + cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv + sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv + rm \${name}_count.csv + fi """ + } } - } else { - process ASV_Phylogeny { + if (params.ncASV) { - label 'norm_cpus' + process Generate_Nucleotide_Matrix { + + label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*ASV*aln.*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*ASV*mt*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*ASV*iq*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Matrix", mode: "copy", overwrite: true, pattern: '*_ASV*PercentID.matrix' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Matrix", mode: "copy", overwrite: true, pattern: '*ncASV*PercentID.matrix' input: - file(asvs) from nuclFastas_forphylogeny + tuple file(notus), file(asvs) from nuclFastas_forMatrix_ch output: - tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into align_results - file("*iq.treefile") into nucl_phyl_plot + file("*.matrix") into clustmatrices + file("*ncASV*PercentID.matrix") into notu_heatmap + file("*_ASV*PercentID.matrix") into asv_heatmap script: + // remove if statement later (no fin) """ - for filename in ${asvs};do - pre=\$(echo \${filename} | awk -F ".fasta" '{print \$1}' ) - mafft --thread ${task.cpus} --maxiterate 15000 --auto \${filename} >\${pre}_ALN.fasta - trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html - # Nucleotide_ModelTest - modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint - # Nucleotide_Phylogeny - if [ "${params.iqCustomnt}" != "" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomnt} - elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} - elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni - elif [ "${params.nonparametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} - elif [ "${params.parametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - else - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + for filename in ${notus};do + if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then + ident=\$( echo \${filename} | awk -F "ncASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') + name=\$( echo \${filename}| awk -F ".fasta" '{print \$1}') + clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} + clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} + for x in *q.matrix;do + pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') + ya=\$(wc -l \$x | awk '{print \$1}') + echo "\$((\$ya-1))" + tail -"\$((\$ya-1))" \$x > \${pre}z.matrix + rm \$x + cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix + rm \${pre}z.matrix + done fi done + if [ `echo ${asvs} | grep -c "_ASV"` -eq 1 ];then + name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') + clustalo -i ${asvs} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} + clustalo -i ${asvs} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} + for x in *q.matrix;do + pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') + ya=\$(wc -l \$x | awk '{print \$1}') + echo "\$((\$ya-1))" + tail -"\$((\$ya-1))" \$x > \${pre}z.matrix + rm \$x + cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix + rm \${pre}z.matrix + done + fi """ - } - } - } + } + } else { - if (!params.skipAminoTyping) { + process Generate_ASV_Matrix { - if (params.sing) { + label 'low_cpus' - process Translating_For_Aminotypes { + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Matrix", mode: "copy", overwrite: true, pattern: '*ASV*PercentID.matrix' - label 'low_cpus' + input: + file(reads) from nuclFastas_forMatrix_ch - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes/Translation", mode: "copy", overwrite: true + output: + file("*.matrix") into clustmatrices + file("*_ASV*PercentID.matrix") into asv_heatmap - input: - file(fasta) from asvsforAminotyping + script: + // remove if statement later (no fin) + """ + for filename in ${reads};do + if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then + ident=\$( echo \${filename} | awk -F "ncASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') + name=\$( echo \${filename}| awk -F ".fasta" '{print \$1}') + clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} + clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} + for x in *q.matrix;do + pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') + ya=\$(wc -l \$x | awk '{print \$1}') + echo "\$((\$ya-1))" + tail -"\$((\$ya-1))" \$x > \${pre}z.matrix + rm \$x + cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix + rm \${pre}z.matrix + done + else + name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') + clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} + clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} + for x in *q.matrix;do + pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') + ya=\$(wc -l \$x | awk '{print \$1}') + echo "\$((\$ya-1))" + tail -"\$((\$ya-1))" \$x > \${pre}z.matrix + rm \$x + cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix + rm \${pre}z.matrix + done + fi + done + """ + } + } - output: - file("${params.projtag}_all_translations.fasta") into amintypegen - file("${params.projtag}_translation_report") into proteinstage_vap_report + if (!params.skipPhylogeny) { // need to edit paths + + if (params.ncASV) { + + process Nucleotide_Phylogeny { + + label 'norm_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*ncASV*aln.*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*ncASV*mt*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*ncASV*iq*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*_ASV*aln.*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*_ASV*mt*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*_ASV*iq*' + + input: + tuple file(notus), file(asvs) from nuclFastas_forphylogeny + + output: + tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into align_results + file("*iq.treefile") into nucl_phyl_plot + + script: + """ + for filename in ${notus};do + pre=\$(echo \${filename} | awk -F ".fasta" '{print \$1}' ) + mafft --thread ${task.cpus} --maxiterate 15000 --auto \${filename} >\${pre}_ALN.fasta + trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html + # Nucleotide_ModelTest + modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint + # Nucleotide_Phylogeny + if [ "${params.iqCustomnt}" != "" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -t \${pre}_mt.tree -T auto ${params.iqCustomnt} + elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} + elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni + elif [ "${params.nonparametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} + elif [ "${params.parametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni + else + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni + fi + done + for filename in ${asvs};do + pre=\$(echo \${filename} | awk -F ".fasta" '{print \$1}' ) + mafft --thread ${task.cpus} --maxiterate 15000 --auto \${filename} >\${pre}_ALN.fasta + trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html + # Nucleotide_ModelTest + modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint + # Nucleotide_Phylogeny + if [ "${params.iqCustomnt}" != "" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomnt} + elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} + elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni + elif [ "${params.nonparametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} + elif [ "${params.parametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + else + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + fi + done + """ + } + } else { + + process ASV_Phylogeny { + + label 'norm_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*ASV*aln.*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*ASV*mt*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*ASV*iq*' + + input: + file(asvs) from nuclFastas_forphylogeny + + output: + tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into align_results + file("*iq.treefile") into nucl_phyl_plot + + script: + """ + for filename in ${asvs};do + pre=\$(echo \${filename} | awk -F ".fasta" '{print \$1}' ) + mafft --thread ${task.cpus} --maxiterate 15000 --auto \${filename} >\${pre}_ALN.fasta + trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html + # Nucleotide_ModelTest + modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint + # Nucleotide_Phylogeny + if [ "${params.iqCustomnt}" != "" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomnt} + elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} + elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni + elif [ "${params.nonparametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} + elif [ "${params.parametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + else + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + fi + done + """ + } + } + } - script: - """ - conda init && source activate virtualribosome + if (!params.skipAminoTyping) { - ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_all_translations.fasta --report ${params.projtag}_translation_report - """ + if (params.sing) { - } + process Translating_For_Aminotypes { - } else { + label 'low_cpus' - process Translate_For_AminoTyping { + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes/Translation", mode: "copy", overwrite: true - label 'low_cpus' + input: + file(fasta) from asvsforAminotyping - conda 'python=2.7' + output: + file("${params.projtag}_all_translations.fasta") into amintypegen + file("${params.projtag}_translation_report") into proteinstage_vap_report - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes/Translation", mode: "copy", overwrite: true + script: + """ + conda init && source activate virtualribosome - input: - file(fasta) from asvsforAminotyping + ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_all_translations.fasta --report ${params.projtag}_translation_report + """ - output: - file("${params.projtag}_all_translations.fasta") into amintypegen - file("${params.projtag}_translation_report") into proteinstage_vap_report + } - script: - """ - ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_all_translations.fasta --report ${params.projtag}_translation_report - """ - } + } else { - } + process Translate_For_AminoTyping { - process Generate_AminoTypes { + label 'low_cpus' - label 'norm_cpus' + conda 'python=2.7' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{clstr,csv,gc}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes/Problematic", mode: "copy", overwrite: true, pattern: '*problematic*.{fasta}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes", mode: "copy", overwrite: true, pattern: '*AminoTypes_noTaxonomy.{fasta}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes/Translation", mode: "copy", overwrite: true - input: - file(prot) from amintypegen - file(asvs) from asvaminocheck + input: + file(fasta) from asvsforAminotyping - output: - tuple file("*.fasta"), file("${params.projtag}_AminoTypes.clstr"), file("${params.projtag}_AminoType_summary_map.csv"), file("${params.projtag}_clustered.gc") into ( supplementalfiles ) - file("${params.projtag}_AminoTypes_noTaxonomy.fasta") into ( aminotypesCounts, aminotypesMafft, aminotypesClustal, aminotypesBlast, aminotypesEmboss ) + output: + file("${params.projtag}_all_translations.fasta") into amintypegen + file("${params.projtag}_translation_report") into proteinstage_vap_report - script: - """ - set +e - cp ${params.vampdir}/bin/rename_seq.py . - awk 'BEGIN{RS=">";ORS=""}length(\$2)>="${params.minAA}"{print ">"\$0}' ${prot} >${params.projtag}_filtered_translations.fasta - awk 'BEGIN{RS=">";ORS=""}length(\$2)<"${params.minAA}"{print ">"\$0}' ${prot} >${params.projtag}_problematic_translations.fasta - if [ `wc -l ${params.projtag}_problematic_translations.fasta | awk '{print \$1}'` -gt 1 ];then - grep ">" ${params.projtag}_problematic_translations.fasta | awk -F ">" '{print \$2}' > problem_tmp.list - seqtk subseq ${asvs} problem_tmp.list > ${params.projtag}_problematic_nucleotides.fasta - else - rm ${params.projtag}_problematic_translations.fasta - fi - cd-hit -i ${params.projtag}_filtered_translations.fasta -c 1.0 -o ${params.projtag}_unlabeled_types.fasta - sed 's/>Cluster />Cluster_/g' ${params.projtag}_unlabeled_types.fasta.clstr >${params.projtag}_AminoTypes.clstr - grep ">Cluster_" ${params.projtag}_AminoTypes.clstr >tmpclusters.list - grep -w "*" ${params.projtag}_AminoTypes.clstr | awk '{print \$3}' | awk -F "." '{print \$1}' >tmphead.list - grep -w "*" ${params.projtag}_AminoTypes.clstr | awk '{print \$2}' | awk -F "," '{print \$1}' >tmplen.list - paste -d"," tmpclusters.list tmphead.list >tmp.info.csv - grep ">" ${params.projtag}_unlabeled_types.fasta >lala.list - j=1 - for x in \$(cat lala.list);do - echo ">${params.projtag}_AminoType\${j}" >>${params.projtag}_aminoheaders.list - echo "\${x},>${params.projtag}_AminoType\${j}" >>tmpaminotype.info.csv - j=\$(( \${j}+1 )) - done - rm lala.list - awk -F "," '{print \$2}' tmp.info.csv >>tmporder.list - for x in \$(cat tmporder.list);do - grep -w "\$x" tmpaminotype.info.csv | awk -F "," '{print \$2}' >>tmpder.list - done - paste -d "," tmpclusters.list tmplen.list tmphead.list tmpder.list >${params.projtag}_AminoType_summary_map.csv - rm tmp* - ./rename_seq.py ${params.projtag}_unlabeled_types.fasta ${params.projtag}_aminoheaders.list ${params.projtag}_AminoTypes_noTaxonomy.fasta - stats.sh in=${params.projtag}_AminoTypes_noTaxonomy.fasta gc=${params.projtag}_clustered.gc gcformat=4 - """ - } + script: + """ + ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_all_translations.fasta --report ${params.projtag}_translation_report + """ + } + } - process Generate_AminoType_Matrix { + process Generate_AminoTypes { - label 'low_cpus' + label 'norm_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Matrix", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{clstr,csv,gc}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes/Problematic", mode: "copy", overwrite: true, pattern: '*problematic*.{fasta}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes", mode: "copy", overwrite: true, pattern: '*AminoTypes_noTaxonomy.{fasta}' - input: - file(prot) from aminotypesClustal + input: + file(prot) from amintypegen + file(asvs) from asvaminocheck - output: - file("*.matrix") into proclustmatrices - file("*PercentID.matrix") into aminotype_heatmap + output: + tuple file("*.fasta"), file("${params.projtag}_AminoTypes.clstr"), file("${params.projtag}_AminoType_summary_map.csv"), file("${params.projtag}_clustered.gc") into ( supplementalfiles ) + file("${params.projtag}_AminoTypes_noTaxonomy.fasta") into ( aminotypesCounts, aminotypesMafft, aminotypesClustal, aminotypesBlast, aminotypesEmboss ) - script: - """ - name=\$( echo ${prot} | awk -F ".fasta" '{print \$1}') - clustalo -i ${prot} --distmat-out=\${name}_PairwiseDistanceq.matrix --full --force --threads=${task.cpus} - clustalo -i ${prot} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} - for x in *q.matrix;do - pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') - ya=\$(wc -l \$x | awk '{print \$1}') - echo "\$((\$ya-1))" - tail -"\$(( \$ya-1))" \$x > \${pre}z.matrix - rm \$x - cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix - rm \${pre}z.matrix - done - """ - } + script: + """ + set +e + cp ${params.vampdir}/bin/rename_seq.py . + awk 'BEGIN{RS=">";ORS=""}length(\$2)>="${params.minAA}"{print ">"\$0}' ${prot} >${params.projtag}_filtered_translations.fasta + awk 'BEGIN{RS=">";ORS=""}length(\$2)<"${params.minAA}"{print ">"\$0}' ${prot} >${params.projtag}_problematic_translations.fasta + if [ `wc -l ${params.projtag}_problematic_translations.fasta | awk '{print \$1}'` -gt 1 ];then + grep ">" ${params.projtag}_problematic_translations.fasta | awk -F ">" '{print \$2}' > problem_tmp.list + seqtk subseq ${asvs} problem_tmp.list > ${params.projtag}_problematic_nucleotides.fasta + else + rm ${params.projtag}_problematic_translations.fasta + fi + cd-hit -i ${params.projtag}_filtered_translations.fasta -c 1.0 -o ${params.projtag}_unlabeled_types.fasta + sed 's/>Cluster />Cluster_/g' ${params.projtag}_unlabeled_types.fasta.clstr >${params.projtag}_AminoTypes.clstr + grep ">Cluster_" ${params.projtag}_AminoTypes.clstr >tmpclusters.list + grep -w "*" ${params.projtag}_AminoTypes.clstr | awk '{print \$3}' | awk -F "." '{print \$1}' >tmphead.list + grep -w "*" ${params.projtag}_AminoTypes.clstr | awk '{print \$2}' | awk -F "," '{print \$1}' >tmplen.list + paste -d"," tmpclusters.list tmphead.list >tmp.info.csv + grep ">" ${params.projtag}_unlabeled_types.fasta >lala.list + j=1 + for x in \$(cat lala.list);do + echo ">${params.projtag}_AminoType\${j}" >>${params.projtag}_aminoheaders.list + echo "\${x},>${params.projtag}_AminoType\${j}" >>tmpaminotype.info.csv + j=\$(( \${j}+1 )) + done + rm lala.list + awk -F "," '{print \$2}' tmp.info.csv >>tmporder.list + for x in \$(cat tmporder.list);do + grep -w "\$x" tmpaminotype.info.csv | awk -F "," '{print \$2}' >>tmpder.list + done + paste -d "," tmpclusters.list tmplen.list tmphead.list tmpder.list >${params.projtag}_AminoType_summary_map.csv + rm tmp* + ./rename_seq.py ${params.projtag}_unlabeled_types.fasta ${params.projtag}_aminoheaders.list ${params.projtag}_AminoTypes_noTaxonomy.fasta + stats.sh in=${params.projtag}_AminoTypes_noTaxonomy.fasta gc=${params.projtag}_clustered.gc gcformat=4 + """ + } - if (!params.skipEMBOSS) { + process Generate_AminoType_Matrix { - process AminoType_EMBOSS_Analyses { + label 'low_cpus' - label 'low_cpus' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Matrix", mode: "copy", overwrite: true - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/2dStructure", mode: "copy", overwrite: true, pattern: '*.{garnier}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/HydrophobicMoment", mode: "copy", overwrite: true, pattern: '*HydrophobicMoments.{svg}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/IsoelectricPoint", mode: "copy", overwrite: true, pattern: '*IsoelectricPoint.{iep,svg}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/ProteinProperties", mode: "copy", overwrite: true, pattern: '*.{pepstats,pepinfo}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/ProteinProperties/Plots", mode: "copy", overwrite: true, pattern: '*PropertiesPlot.{svg}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/2dStructure/Plots", mode: "copy", overwrite: true, pattern: '*Helical*.{svg}' + input: + file(prot) from aminotypesClustal - input: - file(prot) from aminotypesEmboss + output: + file("*.matrix") into proclustmatrices + file("*PercentID.matrix") into aminotype_heatmap - output: - tuple file("*.garnier"), file("*HydrophobicMoments.svg"), file("*IsoelectricPoint*"), file("*.pepstats"), file("*PropertiesPlot*"), file("*Helical*") into amino_emboss + script: + """ + name=\$( echo ${prot} | awk -F ".fasta" '{print \$1}') + clustalo -i ${prot} --distmat-out=\${name}_PairwiseDistanceq.matrix --full --force --threads=${task.cpus} + clustalo -i ${prot} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} + for x in *q.matrix;do + pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') + ya=\$(wc -l \$x | awk '{print \$1}') + echo "\$((\$ya-1))" + tail -"\$(( \$ya-1))" \$x > \${pre}z.matrix + rm \$x + cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix + rm \${pre}z.matrix + done + """ + } - script: - """ - name=\$( echo ${prot} | awk -F ".fasta" '{print \$1}') - garnier -sequence ${prot} -outfile \${name}_2dStructures.garnier - hmoment -seqall ${prot} -graph svg -plot - mv hmoment.svg ./"\${name}"_HydrophobicMoments.svg - iep -sequence ${prot} -graph svg -plot -outfile "\${name}"_IsoelectricPoint.iep - mv iep.svg ./"\${name}"_IsoelectricPoint.svg - pepstats -sequence ${prot} -outfile \${name}_ProteinProperties.pepstats - grep ">" ${prot} | awk -F ">" '{print \$2}' > tmpsequence.list - for x in \$(cat tmpsequence.list);do - echo \$x > tmp1.list - seqtk subseq ${prot} tmp1.list > tmp2.fasta - len=\$(tail -1 tmp2.fasta | awk '{print length}') - pepinfo -sequence tmp2.fasta -graph svg -outfile "\$x"_PropertiesPlot.pepinfo - mv pepinfo.svg ./"\$x"_PropertiesPlot.svg - cat "\$x"_PropertiesPlot.pepinfo >> "\${name}"_PropertiesPlot.pepinfo - rm "\$x"_PropertiesPlot.pepinfo - pepnet -sask -sequence tmp2.fasta -graph svg -sbegin1 1 -send1 \$len - mv pepnet.svg ./"\$x"_HelicalNet.svg - pepwheel -sequence tmp2.fasta -graph svg -sbegin1 1 -send1 \$len - mv pepwheel.svg ./"\$x"_HelicalWheel.svg - rm tmp1.list tmp2.fasta - done - rm tmpsequence.list - """ + if (!params.skipEMBOSS) { + + process AminoType_EMBOSS_Analyses { + + label 'low_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/2dStructure", mode: "copy", overwrite: true, pattern: '*.{garnier}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/HydrophobicMoment", mode: "copy", overwrite: true, pattern: '*HydrophobicMoments.{svg}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/IsoelectricPoint", mode: "copy", overwrite: true, pattern: '*IsoelectricPoint.{iep,svg}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/ProteinProperties", mode: "copy", overwrite: true, pattern: '*.{pepstats,pepinfo}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/ProteinProperties/Plots", mode: "copy", overwrite: true, pattern: '*PropertiesPlot.{svg}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/2dStructure/Plots", mode: "copy", overwrite: true, pattern: '*Helical*.{svg}' + + input: + file(prot) from aminotypesEmboss + + output: + tuple file("*.garnier"), file("*HydrophobicMoments.svg"), file("*IsoelectricPoint*"), file("*.pepstats"), file("*PropertiesPlot*"), file("*Helical*") into amino_emboss + + script: + """ + name=\$( echo ${prot} | awk -F ".fasta" '{print \$1}') + garnier -sequence ${prot} -outfile \${name}_2dStructures.garnier + hmoment -seqall ${prot} -graph svg -plot + mv hmoment.svg ./"\${name}"_HydrophobicMoments.svg + iep -sequence ${prot} -graph svg -plot -outfile "\${name}"_IsoelectricPoint.iep + mv iep.svg ./"\${name}"_IsoelectricPoint.svg + pepstats -sequence ${prot} -outfile \${name}_ProteinProperties.pepstats + grep ">" ${prot} | awk -F ">" '{print \$2}' > tmpsequence.list + for x in \$(cat tmpsequence.list);do + echo \$x > tmp1.list + seqtk subseq ${prot} tmp1.list > tmp2.fasta + len=\$(tail -1 tmp2.fasta | awk '{print length}') + pepinfo -sequence tmp2.fasta -graph svg -outfile "\$x"_PropertiesPlot.pepinfo + mv pepinfo.svg ./"\$x"_PropertiesPlot.svg + cat "\$x"_PropertiesPlot.pepinfo >> "\${name}"_PropertiesPlot.pepinfo + rm "\$x"_PropertiesPlot.pepinfo + pepnet -sask -sequence tmp2.fasta -graph svg -sbegin1 1 -send1 \$len + mv pepnet.svg ./"\$x"_HelicalNet.svg + pepwheel -sequence tmp2.fasta -graph svg -sbegin1 1 -send1 \$len + mv pepwheel.svg ./"\$x"_HelicalWheel.svg + rm tmp1.list tmp2.fasta + done + rm tmpsequence.list + """ + } } - } - if (!params.skipTaxonomy) { + if (!params.skipTaxonomy) { - process AminoType_Taxonomy_Inference { + process AminoType_Taxonomy_Inference { - label 'high_cpus' + label 'high_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{csv,tsv}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Taxonomy", mode: "copy", overwrite: true, pattern: '*TaxonomyLabels.fasta' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{csv,tsv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Taxonomy", mode: "copy", overwrite: true, pattern: '*TaxonomyLabels.fasta' - input: - file(reads) from aminotypesBlast + input: + file(reads) from aminotypesBlast - output: - tuple file("*_phyloseqObject.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AA_diamond - file("*_summary_for_plot.csv") into taxplot2 - file("*TaxonomyLabels.fasta") into tax_labeled_fasta2 + output: + tuple file("*_phyloseqObject.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AA_diamond + file("*_summary_for_plot.csv") into taxplot2 + file("*TaxonomyLabels.fasta") into tax_labeled_fasta2 - script: - """ - cp ${params.vampdir}/bin/rename_seq.py . - virdb=${params.dbdir}/${params.dbname} - grep ">" \${virdb} >> headers.list - headers="headers.list" - name=\$(ls ${reads} | awk -F "_noTaxonomy" '{print \$1}') - diamond blastp -q ${reads} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 - echo "Preparing lists to generate summary .csv's" - echo "[Best hit accession number]" >access.list - echo "[pcASV sequence length]" >length.list - echo "[e-value]" >evalue.list - echo "[Bitscore]" >bit.list - echo "[Percent ID (aa)]" >pid.list - echo "[AminoType#]" >otu.list - echo "[Virus ID]" >"\$name"_virus.list - echo "[Gene]" >"\$name"_genes.list - grep ">" ${reads} | awk -F ">" '{print \$2}' > seqids.lst - echo "extracting genes and names" - touch new_"\$name"_asvnames.txt - j=1 - for s in \$(cat seqids.lst);do - echo "Checking for \$s hit in diamond output" - if [[ ${params.refseq} == "T" ]];then - echo "RefSeq headers specified" - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >>evalue.list - echo "\$line" | awk '{print \$11}' >>bit.list - echo "\$line" | awk '{print \$12}' >>pid.list - echo "\$line" | awk '{print \$2}' >>length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">AminoType\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >>access.list - echo "NO_HIT" >>"\$name"_genes.list - echo "NO_HIT" >>"\$name"_virus.list - echo "NO_HIT" >>evalue.list - echo "NO_HIT" >>bit.list - echo "NO_HIT" >>pid.list - echo "NO_HIT" >>length.list - virus="NO" - gene="HIT" - echo ">AminoType\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - fi - else - echo "Using RVDB headers." - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') - echo "\$s" >>otu.list - echo "\$acc" >>access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >>evalue.list - echo "\$line" | awk '{print \$11}' >>bit.list - echo "\$line" | awk '{print \$12}' >>pid.list - echo "\$line" | awk '{print \$2}' >>length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') - echo "\$gene" | sed 's/_/ /g' >>"\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') - echo "\$virus" | sed 's/_/ /g' >>"\$name"_virus.list - echo ">AminoType\${j}_"\$virus"_"\$gene"" >>new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >>otu.list - echo "NO_HIT" >>access.list - echo "NO_HIT" >>"\$name"_genes.list - echo "NO_HIT" >>"\$name"_virus.list - echo "NO_HIT" >>evalue.list - echo "NO_HIT" >>bit.list - echo "NO_HIT" >>pid.list - echo "NO_HIT" >>length.list - virus="NO" - gene="HIT" - echo ">AminoType\${j}_"\$virus"_"\$gene"" >>new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - fi - fi - echo "Done with \$s" - done - echo "Now editing "\$name" fasta headers" - ###### rename_seq.py - ./rename_seq.py ${reads} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta - awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta > "\$name"_tmpssasv.fasta - echo "[Sequence header]" > newnames.list - cat new_"\$name"_asvnames.txt >> newnames.list - touch sequence.list - echo " " > sequence.list - grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list - rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - for x in *phyloseqObject.csv;do - echo "\$x" - lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) - tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; - sed 's/ /_/g' tmpcol.list > tmp2col.list; - cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; - rm tmpcol.list tmp2col.list - done - rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst headers.list otu.list ; - rm *asvnames.txt - rm "\$name"_virus.list - rm "\$name"_genes.list - rm newnames.list - rm access.list - echo "Taxonomy inferred for: ${reads} " - """ - } - } + script: + """ + cp ${params.vampdir}/bin/rename_seq.py . + virdb=${params.dbdir}/${params.dbname} + grep ">" \${virdb} >> headers.list + headers="headers.list" + name=\$(ls ${reads} | awk -F "_noTaxonomy" '{print \$1}') + diamond blastp -q ${reads} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + echo "Preparing lists to generate summary .csv's" + echo "[Best hit accession number]" >access.list + echo "[pcASV sequence length]" >length.list + echo "[e-value]" >evalue.list + echo "[Bitscore]" >bit.list + echo "[Percent ID (aa)]" >pid.list + echo "[AminoType#]" >otu.list + echo "[Virus ID]" >"\$name"_virus.list + echo "[Gene]" >"\$name"_genes.list + grep ">" ${reads} | awk -F ">" '{print \$2}' > seqids.lst + echo "extracting genes and names" + touch new_"\$name"_asvnames.txt + j=1 + for s in \$(cat seqids.lst);do + echo "Checking for \$s hit in diamond output" + if [[ ${params.refseq} == "T" ]];then + echo "RefSeq headers specified" + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >>evalue.list + echo "\$line" | awk '{print \$11}' >>bit.list + echo "\$line" | awk '{print \$12}' >>pid.list + echo "\$line" | awk '{print \$2}' >>length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">AminoType\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >>access.list + echo "NO_HIT" >>"\$name"_genes.list + echo "NO_HIT" >>"\$name"_virus.list + echo "NO_HIT" >>evalue.list + echo "NO_HIT" >>bit.list + echo "NO_HIT" >>pid.list + echo "NO_HIT" >>length.list + virus="NO" + gene="HIT" + echo ">AminoType\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + fi + else + echo "Using RVDB headers." + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') + echo "\$s" >>otu.list + echo "\$acc" >>access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >>evalue.list + echo "\$line" | awk '{print \$11}' >>bit.list + echo "\$line" | awk '{print \$12}' >>pid.list + echo "\$line" | awk '{print \$2}' >>length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') + echo "\$gene" | sed 's/_/ /g' >>"\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') + echo "\$virus" | sed 's/_/ /g' >>"\$name"_virus.list + echo ">AminoType\${j}_"\$virus"_"\$gene"" >>new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >>otu.list + echo "NO_HIT" >>access.list + echo "NO_HIT" >>"\$name"_genes.list + echo "NO_HIT" >>"\$name"_virus.list + echo "NO_HIT" >>evalue.list + echo "NO_HIT" >>bit.list + echo "NO_HIT" >>pid.list + echo "NO_HIT" >>length.list + virus="NO" + gene="HIT" + echo ">AminoType\${j}_"\$virus"_"\$gene"" >>new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + fi + fi + echo "Done with \$s" + done + echo "Now editing "\$name" fasta headers" + ###### rename_seq.py + ./rename_seq.py ${reads} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta > "\$name"_tmpssasv.fasta + echo "[Sequence header]" > newnames.list + cat new_"\$name"_asvnames.txt >> newnames.list + touch sequence.list + echo " " > sequence.list + grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list + rm "\$name"_tmpssasv.fasta + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + for x in *phyloseqObject.csv;do + echo "\$x" + lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) + tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; + sed 's/ /_/g' tmpcol.list > tmp2col.list; + cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; + rm tmpcol.list tmp2col.list + done + rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst headers.list otu.list ; + rm *asvnames.txt + rm "\$name"_virus.list + rm "\$name"_genes.list + rm newnames.list + rm access.list + echo "Taxonomy inferred for: ${reads} " + """ + } + } - if (!params.skipPhylogeny) { + if (!params.skipPhylogeny) { - process AminoType_Phylogeny { + process AminoType_Phylogeny { - label 'norm_cpus' + label 'norm_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*aln.*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Phylogeny/Modeltest", mode: "copy", overwrite: true, pattern: '*mt*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*iq*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*aln.*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Phylogeny/Modeltest", mode: "copy", overwrite: true, pattern: '*mt*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*iq*' - input: - file(prot) from aminotypesMafft + input: + file(prot) from aminotypesMafft - output: - tuple file("*_aln.fasta"), file("*_aln.html"), file("*.log"), file("*iq*"), file("*mt*") into alignprot_results - file("*iq.treefile") into amino_rax_plot + output: + tuple file("*_aln.fasta"), file("*_aln.html"), file("*.log"), file("*iq*"), file("*mt*") into alignprot_results + file("*iq.treefile") into amino_rax_plot - script: - """ - # Protein_Alignment - pre=\$(echo ${prot} | awk -F ".fasta" '{print \$1}' ) - mafft --thread ${task.cpus} --maxiterate 15000 --auto ${prot} >\${pre}_ALN.fasta - trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html + script: + """ + # Protein_Alignment + pre=\$(echo ${prot} | awk -F ".fasta" '{print \$1}' ) + mafft --thread ${task.cpus} --maxiterate 15000 --auto ${prot} >\${pre}_ALN.fasta + trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html - # Protein_ModelTest - modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d aa -s 203 --disable-checkpoint + # Protein_ModelTest + modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d aa -s 203 --disable-checkpoint - # Protein_Phylogeny - if [ "${params.iqCustomaa}" != "" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomaa} + # Protein_Phylogeny + if [ "${params.iqCustomaa}" != "" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomaa} - elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} + elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} - elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni + elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni - elif [ "${params.nonparametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} + elif [ "${params.nonparametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} - elif [ "${params.parametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + elif [ "${params.parametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - else - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - fi - """ - } - } + else + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + fi + """ + } + } - process Generate_AminoTypes_Counts_Table { + process Generate_AminoTypes_Counts_Table { - label 'high_cpus' + label 'high_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Counts", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Counts", mode: "copy", overwrite: true - input: - file(fasta) from aminotypesCounts - file(merged) from mergeforprotcounts - file(samplist) from samplelist + input: + file(fasta) from aminotypesCounts + file(merged) from mergeforprotcounts + file(samplist) from samplelist - output: - tuple file("*_protcounts.csv"), file("*dmd.out") into counts_summary - file("*_protcounts.csv") into aminocounts_plot + output: + tuple file("*_protcounts.csv"), file("*dmd.out") into counts_summary + file("*_protcounts.csv") into aminocounts_plot - script: - """ - set +e - diamond makedb --in ${fasta} --db ${fasta} - diamond blastx -q ${merged} -d ${fasta} -p ${task.cpus} --min-score ${params.ProtCountsBit} --id ${params.ProtCountID} -l ${params.ProtCountsLength} --more-sensitive -o ${params.projtag}_protCounts_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 --max-hsps 1 - echo "OTU_ID" >tmp.col1.txt - echo "Generating sample id list" - grep ">" ${fasta} | awk -F ">" '{print \$2}' | sort | uniq > otuid.list - cat otuid.list >> tmp.col1.txt - echo "Beginning them counts tho my g" - for y in \$( cat ${samplist} );do - echo "Starting with \$y now ..." - grep "\$y" ${params.projtag}_protCounts_dmd.out > tmp."\$y".out - echo "Isolated hits" - echo "Created uniq subject id list" - echo "\$y" > "\$y"_col.txt - echo "Starting my counts" - for z in \$(cat otuid.list);do - echo "Counting \$z hits" - echo "grep -wc "\$z" >> "\$y"_col.txt" - grep -wc "\$z" tmp."\$y".out >> "\$y"_col.txt - echo "\$z counted" - done - done - paste -d "," tmp.col1.txt *col.txt > ${params.projtag}_protcounts.csv - rm tmp* - rm *col.txt - """ - } - } + script: + """ + set +e + diamond makedb --in ${fasta} --db ${fasta} + diamond blastx -q ${merged} -d ${fasta} -p ${task.cpus} --min-score ${params.ProtCountsBit} --id ${params.ProtCountID} -l ${params.ProtCountsLength} --more-sensitive -o ${params.projtag}_protCounts_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 --max-hsps 1 + echo "OTU_ID" >tmp.col1.txt + echo "Generating sample id list" + grep ">" ${fasta} | awk -F ">" '{print \$2}' | sort | uniq > otuid.list + cat otuid.list >> tmp.col1.txt + echo "Beginning them counts tho my g" + for y in \$( cat ${samplist} );do + echo "Starting with \$y now ..." + grep "\$y" ${params.projtag}_protCounts_dmd.out > tmp."\$y".out + echo "Isolated hits" + echo "Created uniq subject id list" + echo "\$y" > "\$y"_col.txt + echo "Starting my counts" + for z in \$(cat otuid.list);do + echo "Counting \$z hits" + echo "grep -wc "\$z" >> "\$y"_col.txt" + grep -wc "\$z" tmp."\$y".out >> "\$y"_col.txt + echo "\$z counted" + done + done + paste -d "," tmp.col1.txt *col.txt > ${params.projtag}_protcounts.csv + rm tmp* + rm *col.txt + """ + } + } - if (params.pcASV) { // ASV_nucl -> ASV_aa -> clusteraa by %id with ch-hit -> extract representative nucl sequences to generate new OTU file + if (params.pcASV) { // ASV_nucl -> ASV_aa -> clusteraa by %id with ch-hit -> extract representative nucl sequences to generate new OTU file - if (params.sing) { + if (params.sing) { - process Translating_For_pcASV_Generation { + process Translating_For_pcASV_Generation { - label 'low_cpus' + label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV/Translation", mode: "copy", overwrite: true, pattern: '*_ASV_translations*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV/Translation", mode: "copy", overwrite: true, pattern: '*_ASV_translations*' - input: - file(fasta) from nucl2aa + input: + file(fasta) from nucl2aa - output: - file("*ASV*translations.fasta") into clustering_aa - file("*_ASV_translations_report") into reportaa_VR - file("*_ASV_nucleotide.fasta") into asvfastaforaaclust + output: + file("*ASV*translations.fasta") into clustering_aa + file("*_ASV_translations_report") into reportaa_VR + file("*_ASV_nucleotide.fasta") into asvfastaforaaclust - script: - """ - conda init && source activate virtualribosome + script: + """ + conda init && source activate virtualribosome - ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_ASV_translations.fasta --report ${params.projtag}_ASV_translations_report - cp ${fasta} ${params.projtag}_ASV_nucleotide.fasta - """ + ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_ASV_translations.fasta --report ${params.projtag}_ASV_translations_report + cp ${fasta} ${params.projtag}_ASV_nucleotide.fasta + """ - } + } - } else { + } else { - process Translation_For_pcASV_Generation { + process Translation_For_pcASV_Generation { - label 'low_cpus' + label 'low_cpus' - conda 'python=2.7' + conda 'python=2.7' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV/Translation", mode: "copy", overwrite: true, pattern: '*_ASV_translations*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV/Translation", mode: "copy", overwrite: true, pattern: '*_ASV_translations*' - input: - file(fasta) from nucl2aa + input: + file(fasta) from nucl2aa - output: - file("*ASV*translations.fasta") into clustering_aa - file("*_ASV_translations_report") into reportaa_VR - file("*_ASV_nucleotide.fasta") into asvfastaforaaclust + output: + file("*ASV*translations.fasta") into clustering_aa + file("*_ASV_translations_report") into reportaa_VR + file("*_ASV_nucleotide.fasta") into asvfastaforaaclust - script: - """ - ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_ASV_translations.fasta --report ${params.projtag}_ASV_translations_report - cp ${fasta} ${params.projtag}_ASV_nucleotide.fasta - """ - } - } + script: + """ + ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_ASV_translations.fasta --report ${params.projtag}_ASV_translations_report + cp ${fasta} ${params.projtag}_ASV_nucleotide.fasta + """ + } + } - process Generate_pcASVs { + process Generate_pcASVs { - label 'norm_cpus' + label 'norm_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV", mode: "copy", overwrite: true, pattern: '*pcASV*.{fasta}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{clstr,csv,gc}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV/Problematic", mode: "copy", overwrite: true, pattern: '*problem*.{fasta}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV", mode: "copy", overwrite: true, pattern: '*pcASV*.{fasta}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{clstr,csv,gc}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV/Problematic", mode: "copy", overwrite: true, pattern: '*problem*.{fasta}' - input: - file(fasta) from clustering_aa - file(asvs) from asvfastaforaaclust + input: + file(fasta) from clustering_aa + file(asvs) from asvfastaforaaclust - output: - file("${params.projtag}_nucleotide_pcASV*.fasta") into ( pcASV_ntDiamond_ch, pcASV_nt_counts_ch, pcASV_ntmatrix_ch, pcASV_ntmafft_ch ) - file("*_aminoacid_pcASV*_noTaxonomy.fasta") into ( pcASV_aaMatrix_ch, pcASV_aaDiamond_ch, pcASV_aaMafft_ch, pcASV_aaCounts_ch, pcASVEMBOSS ) - tuple file("*.fasta"), file("*.clstr"), file("*.csv"), file("*.gc") into ( pcASVsupplementalfiles ) + output: + file("${params.projtag}_nucleotide_pcASV*.fasta") into ( pcASV_ntDiamond_ch, pcASV_nt_counts_ch, pcASV_ntmatrix_ch, pcASV_ntmafft_ch ) + file("*_aminoacid_pcASV*_noTaxonomy.fasta") into ( pcASV_aaMatrix_ch, pcASV_aaDiamond_ch, pcASV_aaMafft_ch, pcASV_aaCounts_ch, pcASVEMBOSS ) + tuple file("*.fasta"), file("*.clstr"), file("*.csv"), file("*.gc") into ( pcASVsupplementalfiles ) - script: - // add awk script to count seqs - if (params.clusterAAIDlist) { - """ - set +e - cp ${params.vampdir}/bin/rename_seq.py . - for id in `echo ${params.clusterAAIDlist} | tr "," "\\n"`;do + script: + // add awk script to count seqs + if (params.clusterAAIDlist) { + """ + set +e + cp ${params.vampdir}/bin/rename_seq.py . + for id in `echo ${params.clusterAAIDlist} | tr "," "\\n"`;do + awk 'BEGIN{RS=">";ORS=""}length(\$2)>="${params.minAA}"{print ">"\$0}' ${fasta} > ${params.projtag}_filtered_proteins.fasta + cd-hit -i ${params.projtag}_filtered_proteins.fasta -c \${id} -o ${params.projtag}_pcASV\${id}.fasta + sed 's/>Cluster />Cluster_/g' ${params.projtag}_pcASV\${id}.fasta.clstr >${params.projtag}_pcASV\${id}.clstr + grep ">Cluster_" ${params.projtag}_pcASV\${id}.clstr >temporaryclusters.list + y=\$(grep -c ">Cluster_" ${params.projtag}_pcASV\${id}.clstr) + echo ">Cluster_"\${y}"" >> ${params.projtag}_pcASV\${id}.clstr + t=1 + b=1 + for x in \$(cat temporaryclusters.list);do + echo "Extracting \$x" + name="\$( echo \$x | awk -F ">" '{print \$2}')" + clust="pcASV"\${t}"" + echo "\${name}" + awk '/^>'\${name}'\$/,/^>Cluster_'\${b}'\$/' ${params.projtag}_pcASV\${id}.clstr > "\${name}"_"\${clust}"_tmp.list + t=\$(( \${t}+1 )) + b=\$(( \${b}+1 )) + done + ls *_tmp.list + u=1 + for x in *_tmp.list;do + name="\$(echo \$x | awk -F "_p" '{print \$1}')" + echo "\${name}" + cluster="\$(echo \$x | awk -F "_" '{print \$3}')" + echo "\${cluster}" + grep "ASV" \$x | awk -F ", " '{print \$2}' | awk -F "_" '{print \$1}' | awk -F ">" '{print \$2}' > \${name}_\${cluster}_seqs_tmps.list + seqtk subseq ${asvs} \${name}_\${cluster}_seqs_tmps.list > \${name}_\${cluster}_nucleotide_sequences.fasta + vsearch --cluster_fast \${name}_\${cluster}_nucleotide_sequences.fasta --id 0.2 --centroids \${name}_\${cluster}_centroids.fasta + grep ">" \${name}_\${cluster}_centroids.fasta >> \${name}_\${cluster}_tmp_centroids.list + for y in \$( cat \${name}_\${cluster}_tmp_centroids.list );do + echo ">\${cluster}_type"\$u"" >> \${name}_\${cluster}_tmp_centroid.newheaders + u=\$(( \${u}+1 )) + done + u=1 + ./rename_seq.py \${name}_\${cluster}_centroids.fasta \${name}_\${cluster}_tmp_centroid.newheaders \${cluster}_types_labeled.fasta + done + cat *_types_labeled.fasta >> ${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta + grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$3}' | awk -F "." '{print \$1}' >tmphead.list + grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$2}' | awk -F "," '{print \$1}' >tmplen.list + paste -d"," temporaryclusters.list tmphead.list >tmp.info.csv + grep ">" ${params.projtag}_pcASV\${id}.fasta >lala.list + j=1 + for x in \$(cat lala.list);do + echo ">${params.projtag}_pcASV\${j}" >>${params.projtag}_aminoheaders.list + echo "\${x},>${params.projtag}_pcASV\${j}" >>tmpaminotype.info.csv + j=\$(( \${j}+1 )) + done + rm lala.list + awk -F "," '{print \$2}' tmp.info.csv >>tmporder.list + for x in \$(cat tmporder.list);do + grep -w "\$x" tmpaminotype.info.csv | awk -F "," '{print \$2}' >>tmpder.list + done + paste -d "," temporaryclusters.list tmplen.list tmphead.list tmpder.list >${params.projtag}_pcASVCluster\${id}_summary.csv + ./rename_seq.py ${params.projtag}_pcASV\${id}.fasta ${params.projtag}_aminoheaders.list ${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta + stats.sh in=${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_aminoacid_clustered.gc gcformat=4 overwrite=true + stats.sh in=${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_nucleotide_clustered.gc gcformat=4 overwrite=true + awk 'BEGIN{RS=">";ORS=""}length(\$2)<"${params.minAA}"{print ">"\$0}' ${fasta} >${params.projtag}_pcASV\${id}_problematic_translations.fasta + if [ `wc -l ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk '{print \$1}'` -gt 1 ];then + grep ">" ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk -F ">" '{print \$2}' > problem_tmp.list + seqtk subseq ${asvs} > ${params.projtag}_pcASV\${id}_problematic_nucleotides.fasta + else + rm ${params.projtag}_pcASV\${id}_problematic_translations.fasta + fi + rm *.list + rm Cluster* + rm *types* + rm *tmp* + rm ${params.projtag}_pcASV\${id}.fast* + done + """ + } else if (params.clusterAAID) { + """ + set +e + cp ${params.vampdir}/bin/rename_seq.py . + id=${params.clusterAAID} awk 'BEGIN{RS=">";ORS=""}length(\$2)>="${params.minAA}"{print ">"\$0}' ${fasta} > ${params.projtag}_filtered_proteins.fasta - cd-hit -i ${params.projtag}_filtered_proteins.fasta -c \${id} -o ${params.projtag}_pcASV\${id}.fasta + cd-hit -i ${params.projtag}_filtered_proteins.fasta -c ${params.clusterAAID} -o ${params.projtag}_pcASV\${id}.fasta sed 's/>Cluster />Cluster_/g' ${params.projtag}_pcASV\${id}.fasta.clstr >${params.projtag}_pcASV\${id}.clstr grep ">Cluster_" ${params.projtag}_pcASV\${id}.clstr >temporaryclusters.list y=\$(grep -c ">Cluster_" ${params.projtag}_pcASV\${id}.clstr) @@ -2187,7 +2587,8 @@ if (params.Analyze) { t=\$(( \${t}+1 )) b=\$(( \${b}+1 )) done - ls *_tmp.list + + ls *_tmp.list u=1 for x in *_tmp.list;do name="\$(echo \$x | awk -F "_p" '{print \$1}')" @@ -2223,12 +2624,12 @@ if (params.Analyze) { done paste -d "," temporaryclusters.list tmplen.list tmphead.list tmpder.list >${params.projtag}_pcASVCluster\${id}_summary.csv ./rename_seq.py ${params.projtag}_pcASV\${id}.fasta ${params.projtag}_aminoheaders.list ${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta - stats.sh in=${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_aminoacid_clustered.gc gcformat=4 overwrite=true - stats.sh in=${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_nucleotide_clustered.gc gcformat=4 overwrite=true + stats.sh in=${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_aminoacid_clustered.gc gcformat=4 + stats.sh in=${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_nucleotide_clustered.gc gcformat=4 awk 'BEGIN{RS=">";ORS=""}length(\$2)<"${params.minAA}"{print ">"\$0}' ${fasta} >${params.projtag}_pcASV\${id}_problematic_translations.fasta if [ `wc -l ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk '{print \$1}'` -gt 1 ];then grep ">" ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk -F ">" '{print \$2}' > problem_tmp.list - seqtk subseq ${asvs} > ${params.projtag}_pcASV\${id}_problematic_nucleotides.fasta + seqtk subseq ${asvs} problem_tmp.list > ${params.projtag}_pcASV\${id}_problematic_nucleotides.fasta else rm ${params.projtag}_pcASV\${id}_problematic_translations.fasta fi @@ -2237,833 +2638,651 @@ if (params.Analyze) { rm *types* rm *tmp* rm ${params.projtag}_pcASV\${id}.fast* - done - """ - } else if (params.clusterAAID) { - """ - set +e - cp ${params.vampdir}/bin/rename_seq.py . - id=${params.clusterAAID} - awk 'BEGIN{RS=">";ORS=""}length(\$2)>="${params.minAA}"{print ">"\$0}' ${fasta} > ${params.projtag}_filtered_proteins.fasta - cd-hit -i ${params.projtag}_filtered_proteins.fasta -c ${params.clusterAAID} -o ${params.projtag}_pcASV\${id}.fasta - sed 's/>Cluster />Cluster_/g' ${params.projtag}_pcASV\${id}.fasta.clstr >${params.projtag}_pcASV\${id}.clstr - grep ">Cluster_" ${params.projtag}_pcASV\${id}.clstr >temporaryclusters.list - y=\$(grep -c ">Cluster_" ${params.projtag}_pcASV\${id}.clstr) - echo ">Cluster_"\${y}"" >> ${params.projtag}_pcASV\${id}.clstr - t=1 - b=1 - for x in \$(cat temporaryclusters.list);do - echo "Extracting \$x" - name="\$( echo \$x | awk -F ">" '{print \$2}')" - clust="pcASV"\${t}"" - echo "\${name}" - awk '/^>'\${name}'\$/,/^>Cluster_'\${b}'\$/' ${params.projtag}_pcASV\${id}.clstr > "\${name}"_"\${clust}"_tmp.list - t=\$(( \${t}+1 )) - b=\$(( \${b}+1 )) - done - - ls *_tmp.list - u=1 - for x in *_tmp.list;do - name="\$(echo \$x | awk -F "_p" '{print \$1}')" - echo "\${name}" - cluster="\$(echo \$x | awk -F "_" '{print \$3}')" - echo "\${cluster}" - grep "ASV" \$x | awk -F ", " '{print \$2}' | awk -F "_" '{print \$1}' | awk -F ">" '{print \$2}' > \${name}_\${cluster}_seqs_tmps.list - seqtk subseq ${asvs} \${name}_\${cluster}_seqs_tmps.list > \${name}_\${cluster}_nucleotide_sequences.fasta - vsearch --cluster_fast \${name}_\${cluster}_nucleotide_sequences.fasta --id 0.2 --centroids \${name}_\${cluster}_centroids.fasta - grep ">" \${name}_\${cluster}_centroids.fasta >> \${name}_\${cluster}_tmp_centroids.list - for y in \$( cat \${name}_\${cluster}_tmp_centroids.list );do - echo ">\${cluster}_type"\$u"" >> \${name}_\${cluster}_tmp_centroid.newheaders - u=\$(( \${u}+1 )) - done - u=1 - ./rename_seq.py \${name}_\${cluster}_centroids.fasta \${name}_\${cluster}_tmp_centroid.newheaders \${cluster}_types_labeled.fasta - done - cat *_types_labeled.fasta >> ${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta - grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$3}' | awk -F "." '{print \$1}' >tmphead.list - grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$2}' | awk -F "," '{print \$1}' >tmplen.list - paste -d"," temporaryclusters.list tmphead.list >tmp.info.csv - grep ">" ${params.projtag}_pcASV\${id}.fasta >lala.list - j=1 - for x in \$(cat lala.list);do - echo ">${params.projtag}_pcASV\${j}" >>${params.projtag}_aminoheaders.list - echo "\${x},>${params.projtag}_pcASV\${j}" >>tmpaminotype.info.csv - j=\$(( \${j}+1 )) - done - rm lala.list - awk -F "," '{print \$2}' tmp.info.csv >>tmporder.list - for x in \$(cat tmporder.list);do - grep -w "\$x" tmpaminotype.info.csv | awk -F "," '{print \$2}' >>tmpder.list - done - paste -d "," temporaryclusters.list tmplen.list tmphead.list tmpder.list >${params.projtag}_pcASVCluster\${id}_summary.csv - ./rename_seq.py ${params.projtag}_pcASV\${id}.fasta ${params.projtag}_aminoheaders.list ${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta - stats.sh in=${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_aminoacid_clustered.gc gcformat=4 - stats.sh in=${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_nucleotide_clustered.gc gcformat=4 - awk 'BEGIN{RS=">";ORS=""}length(\$2)<"${params.minAA}"{print ">"\$0}' ${fasta} >${params.projtag}_pcASV\${id}_problematic_translations.fasta - if [ `wc -l ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk '{print \$1}'` -gt 1 ];then - grep ">" ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk -F ">" '{print \$2}' > problem_tmp.list - seqtk subseq ${asvs} problem_tmp.list > ${params.projtag}_pcASV\${id}_problematic_nucleotides.fasta - else - rm ${params.projtag}_pcASV\${id}_problematic_translations.fasta - fi - rm *.list - rm Cluster* - rm *types* - rm *tmp* - rm ${params.projtag}_pcASV\${id}.fast* - """ - } - } + """ + } + } - if (!params.skipTaxonomy) { + if (!params.skipTaxonomy) { - process pcASV_Nucleotide_Taxonomy_Inference { + process pcASV_Nucleotide_Taxonomy_Inference { - label 'high_cpus' + label 'high_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{csv,tsv}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*dmd.{out}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{fasta}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{csv,tsv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*dmd.{out}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{fasta}' - input: - file(reads) from pcASV_ntDiamond_ch + input: + file(reads) from pcASV_ntDiamond_ch - output: - file("*.fasta") into ( pcASV_labeled ) - tuple file("*_phyloseqObject.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AAdiamond - file("*_summary_for_plot.csv") into taxplot3 + output: + file("*.fasta") into ( pcASV_labeled ) + tuple file("*_phyloseqObject.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AAdiamond + file("*_summary_for_plot.csv") into taxplot3 - script: - """ - set +e - cp ${params.vampdir}/bin/rename_seq.py . - virdb=${params.dbdir}/${params.dbname} - grep ">" \${virdb} >> headers.list - headers="headers.list" - for filename in ${reads};do - name=\$(ls \${filename} | awk -F "_noTax" '{print \$1}') - diamond blastx -q \${filename} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 - echo "Preparing lists to generate summary .csv's" - echo "[Best hit accession number]" >access.list - echo "[pcASV sequence length]" >length.list - echo "[e-value]" >evalue.list - echo "[Bitscore]" >bit.list - echo "[Percent ID (aa)]" >pid.list - echo "[pcASV#]" >otu.list - echo "[Virus ID]" >"\$name"_virus.list - echo "[Gene]" >"\$name"_genes.list - grep ">" \${filename} | awk -F ">" '{print \$2}' > seqids.lst - echo "extracting genes and names" - touch new_"\$name"_headers.txt - j=1 - for s in \$(cat seqids.lst);do - echo "Checking for \$s hit in diamond output" - if [[ ${params.refseq} == "T" ]];then - echo "RefSeq headers specified" - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >>evalue.list - echo "\$line" | awk '{print \$11}' >>bit.list - echo "\$line" | awk '{print \$12}' >>pid.list - echo "\$line" | awk '{print \$2}' >>length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">pcASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_headers.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >>access.list - echo "NO_HIT" >>"\$name"_genes.list - echo "NO_HIT" >>"\$name"_virus.list - echo "NO_HIT" >>evalue.list - echo "NO_HIT" >>bit.list - echo "NO_HIT" >>pid.list - echo "NO_HIT" >>length.list - virus="NO" - gene="HIT" - echo ">pcASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_headers.txt - j=\$((\$j+1)) - echo "\$s done." - fi - else - echo "Using RVDB headers." - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') - echo "\$s" >>otu.list - echo "\$acc" >>access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >>evalue.list - echo "\$line" | awk '{print \$11}' >>bit.list - echo "\$line" | awk '{print \$12}' >>pid.list - echo "\$line" | awk '{print \$2}' >>length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >>"\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && - echo "\$virus" | sed 's/_/ /g' >>"\$name"_virus.list - echo ">pcASV\${j}_"\$virus"_"\$gene"" >>new_"\$name"_headers.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >>otu.list - echo "NO_HIT" >>access.list - echo "NO_HIT" >>"\$name"_genes.list - echo "NO_HIT" >>"\$name"_virus.list - echo "NO_HIT" >>evalue.list - echo "NO_HIT" >>bit.list - echo "NO_HIT" >>pid.list - echo "NO_HIT" >>length.list - virus="NO" - gene="HIT" - echo ">pcASV\${j}_"\$virus"_"\$gene"" >>new_"\$name"_headers.txt - j=\$((\$j+1)) - echo "\$s done." - fi - fi - echo "Done with \$s" - done - echo "Now editing "\$name" fasta headers" - ###### rename_seq.py - ./rename_seq.py \${filename} new_"\$name"_headers.txt "\$name"_TaxonomyLabels.fasta - awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta > "\$name"_tmpssasv.fasta - echo "[Sequence header]" > newnames.list - cat new_"\$name"_headers.txt >> newnames.list - touch sequence.list - echo " " > sequence.list - grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list - rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - for x in *phyloseqObject.csv;do - echo "\$x" - lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) - tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; - sed 's/ /_/g' tmpcol.list > tmp2col.list; - cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; - rm tmpcol.list tmp2col.list - done - rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; - rm "\$name"_virus.list - rm "\$name"_genes.list - rm newnames.list - rm access.list - echo "Taxonomy inferred for: \${filename} " - done - rm *headers.list - """ - } - } + script: + """ + set +e + cp ${params.vampdir}/bin/rename_seq.py . + virdb=${params.dbdir}/${params.dbname} + grep ">" \${virdb} >> headers.list + headers="headers.list" + for filename in ${reads};do + name=\$(ls \${filename} | awk -F "_noTax" '{print \$1}') + diamond blastx -q \${filename} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + echo "Preparing lists to generate summary .csv's" + echo "[Best hit accession number]" >access.list + echo "[pcASV sequence length]" >length.list + echo "[e-value]" >evalue.list + echo "[Bitscore]" >bit.list + echo "[Percent ID (aa)]" >pid.list + echo "[pcASV#]" >otu.list + echo "[Virus ID]" >"\$name"_virus.list + echo "[Gene]" >"\$name"_genes.list + grep ">" \${filename} | awk -F ">" '{print \$2}' > seqids.lst + echo "extracting genes and names" + touch new_"\$name"_headers.txt + j=1 + for s in \$(cat seqids.lst);do + echo "Checking for \$s hit in diamond output" + if [[ ${params.refseq} == "T" ]];then + echo "RefSeq headers specified" + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >>evalue.list + echo "\$line" | awk '{print \$11}' >>bit.list + echo "\$line" | awk '{print \$12}' >>pid.list + echo "\$line" | awk '{print \$2}' >>length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">pcASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_headers.txt + j=\$((\$j+1)) + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >>access.list + echo "NO_HIT" >>"\$name"_genes.list + echo "NO_HIT" >>"\$name"_virus.list + echo "NO_HIT" >>evalue.list + echo "NO_HIT" >>bit.list + echo "NO_HIT" >>pid.list + echo "NO_HIT" >>length.list + virus="NO" + gene="HIT" + echo ">pcASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_headers.txt + j=\$((\$j+1)) + echo "\$s done." + fi + else + echo "Using RVDB headers." + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') + echo "\$s" >>otu.list + echo "\$acc" >>access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >>evalue.list + echo "\$line" | awk '{print \$11}' >>bit.list + echo "\$line" | awk '{print \$12}' >>pid.list + echo "\$line" | awk '{print \$2}' >>length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >>"\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && + echo "\$virus" | sed 's/_/ /g' >>"\$name"_virus.list + echo ">pcASV\${j}_"\$virus"_"\$gene"" >>new_"\$name"_headers.txt + j=\$((\$j+1)) + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >>otu.list + echo "NO_HIT" >>access.list + echo "NO_HIT" >>"\$name"_genes.list + echo "NO_HIT" >>"\$name"_virus.list + echo "NO_HIT" >>evalue.list + echo "NO_HIT" >>bit.list + echo "NO_HIT" >>pid.list + echo "NO_HIT" >>length.list + virus="NO" + gene="HIT" + echo ">pcASV\${j}_"\$virus"_"\$gene"" >>new_"\$name"_headers.txt + j=\$((\$j+1)) + echo "\$s done." + fi + fi + echo "Done with \$s" + done + echo "Now editing "\$name" fasta headers" + ###### rename_seq.py + ./rename_seq.py \${filename} new_"\$name"_headers.txt "\$name"_TaxonomyLabels.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta > "\$name"_tmpssasv.fasta + echo "[Sequence header]" > newnames.list + cat new_"\$name"_headers.txt >> newnames.list + touch sequence.list + echo " " > sequence.list + grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list + rm "\$name"_tmpssasv.fasta + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + for x in *phyloseqObject.csv;do + echo "\$x" + lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) + tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; + sed 's/ /_/g' tmpcol.list > tmp2col.list; + cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; + rm tmpcol.list tmp2col.list + done + rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; + rm "\$name"_virus.list + rm "\$name"_genes.list + rm newnames.list + rm access.list + echo "Taxonomy inferred for: \${filename} " + done + rm *headers.list + """ + } + } - process Generate_Nucleotide_pcASV_Counts { + process Generate_Nucleotide_pcASV_Counts { - label 'norm_cpus' + label 'norm_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Counts", mode: "copy", overwrite: true, pattern: '*.{biome,csv,txt}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Counts", mode: "copy", overwrite: true, pattern: '*.{biome,csv,txt}' - input: - file(potus) from pcASV_nt_counts_ch - file(merged) from pcASV_mergedreads_ch + input: + file(potus) from pcASV_nt_counts_ch + file(merged) from pcASV_mergedreads_ch - output: - tuple file("*_counts.txt"), file("*_counts.biome") into pcASVcounts_vsearch - file("*.csv") into potu_Ncounts_for_report + output: + tuple file("*_counts.txt"), file("*_counts.biome") into pcASVcounts_vsearch + file("*.csv") into potu_Ncounts_for_report - script: - """ - for filename in ${potus};do - ident=\$( echo \${filename} | awk -F "pcASV" '{print \$2}' | awk -F "_noTaxonomy.fasta" '{print \$1}') - name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') - vsearch --usearch_global ${merged} --db \${filename} --id \${ident} --threads ${task.cpus} --otutabout \${name}_counts.txt --biomout \${name}_counts.biome - cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv - sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv - rm \${name}_count.csv - done - """ - } + script: + """ + for filename in ${potus};do + ident=\$( echo \${filename} | awk -F "pcASV" '{print \$2}' | awk -F "_noTaxonomy.fasta" '{print \$1}') + name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') + vsearch --usearch_global ${merged} --db \${filename} --id \${ident} --threads ${task.cpus} --otutabout \${name}_counts.txt --biomout \${name}_counts.biome + cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv + sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv + rm \${name}_count.csv + done + """ + } - process Generate_pcASV_Nucleotide_Matrix { + process Generate_pcASV_Nucleotide_Matrix { - label 'low_cpus' + label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Matrix", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Matrix", mode: "copy", overwrite: true - input: - file(potus) from pcASV_ntmatrix_ch + input: + file(potus) from pcASV_ntmatrix_ch - output: - file("*.matrix") into pcASVclustmatrices - file("*PercentID.matrix") into potu_nucl_heatmap + output: + file("*.matrix") into pcASVclustmatrices + file("*PercentID.matrix") into potu_nucl_heatmap - script: - """ - for filename in ${potus};do - ident=\$( echo \${filename} | awk -F "pcASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') - name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') - clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistanceq.matrix --full --force --threads=${task.cpus} - clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} - for x in *q.matrix;do - pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') - ya=\$(wc -l \$x | awk '{print \$1}') - echo "\$((\$ya-1))" - tail -"\$((\$ya-1))" \$x > \${pre}z.matrix - rm \$x - cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix - rm \${pre}z.matrix - done - done - """ - } + script: + """ + for filename in ${potus};do + ident=\$( echo \${filename} | awk -F "pcASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') + name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') + clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistanceq.matrix --full --force --threads=${task.cpus} + clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} + for x in *q.matrix;do + pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') + ya=\$(wc -l \$x | awk '{print \$1}') + echo "\$((\$ya-1))" + tail -"\$((\$ya-1))" \$x > \${pre}z.matrix + rm \$x + cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix + rm \${pre}z.matrix + done + done + """ + } - if (!params.skipPhylogeny) { + if (!params.skipPhylogeny) { - process pcASV_Nucleotide_Phylogeny { + process pcASV_Nucleotide_Phylogeny { - label 'norm_cpus' + label 'norm_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*aln.*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*mt*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*iq*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*aln.*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*mt*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*iq*' - input: - file(reads) from pcASV_ntmafft_ch + input: + file(reads) from pcASV_ntmafft_ch - output: - tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into pcASV_nucleotide_phylogeny_results - file("*iq.treefile") into potu_Ntree_plot + output: + tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into pcASV_nucleotide_phylogeny_results + file("*iq.treefile") into potu_Ntree_plot - script: - """ - for filename in ${reads};do - pre=\$( echo \${filename} | awk -F "_noTax" '{print \$1}' ) - mafft --maxiterate 5000 --auto \${filename} >\${pre}_ALN.fasta - trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html + script: + """ + for filename in ${reads};do + pre=\$( echo \${filename} | awk -F "_noTax" '{print \$1}' ) + mafft --maxiterate 5000 --auto \${filename} >\${pre}_ALN.fasta + trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html - # pcASV_Nucleotide_ModelTest - modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint + # pcASV_Nucleotide_ModelTest + modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint - # pcASV_Nucleotide_Phylogeny - if [ "${params.iqCustomnt}" != "" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -t \${pre}_mt.tree -T auto ${params.iqCustomnt} + # pcASV_Nucleotide_Phylogeny + if [ "${params.iqCustomnt}" != "" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -t \${pre}_mt.tree -T auto ${params.iqCustomnt} - elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} + elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} - elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni + elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni - elif [ "${params.nonparametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} + elif [ "${params.nonparametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} - elif [ "${params.parametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni + elif [ "${params.parametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni - else - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni - fi - done - """ - } - } + else + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni + fi + done + """ + } + } - process pcASV_AminoAcid_Matrix { + process pcASV_AminoAcid_Matrix { - label 'low_cpus' + label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Matrix", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Matrix", mode: "copy", overwrite: true - input: - file(prot) from pcASV_aaMatrix_ch + input: + file(prot) from pcASV_aaMatrix_ch - output: - file("*.matrix") into pcASVaaMatrix - file("*PercentID.matrix") into potu_aa_heatmap + output: + file("*.matrix") into pcASVaaMatrix + file("*PercentID.matrix") into potu_aa_heatmap - script: - """ - for filename in ${prot};do - name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') - clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistanceq.matrix --full --force --threads=${task.cpus} - clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} - for x in *q.matrix;do - pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') - ya=\$(wc -l \$x | awk '{print \$1}') - echo "\$((\$ya-1))" - tail -"\$((\$ya-1))" \$x > \${pre}z.matrix - rm \$x - cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix - rm \${pre}z.matrix - done - done - """ - } + script: + """ + for filename in ${prot};do + name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') + clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistanceq.matrix --full --force --threads=${task.cpus} + clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} + for x in *q.matrix;do + pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') + ya=\$(wc -l \$x | awk '{print \$1}') + echo "\$((\$ya-1))" + tail -"\$((\$ya-1))" \$x > \${pre}z.matrix + rm \$x + cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix + rm \${pre}z.matrix + done + done + """ + } - if (!params.skipEMBOSS) { + if (!params.skipEMBOSS) { - process pcASV_EMBOSS_Analyses { + process pcASV_EMBOSS_Analyses { - label 'low_cpus' + label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/2dStructure", mode: "copy", overwrite: true, pattern: '*.{garnier}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/HydrophobicMoment", mode: "copy", overwrite: true, pattern: '*HydrophobicMoments.{svg}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/IsoelectricPoint", mode: "copy", overwrite: true, pattern: '*IsoelectricPoint.{iep,svg}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/ProteinProperties", mode: "copy", overwrite: true, pattern: '*.{pepstats,pepinfo}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/ProteinProperties/Plots", mode: "copy", overwrite: true, pattern: '*PropertiesPlot.{svg}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/2dStructure/Plots", mode: "copy", overwrite: true, pattern: '*Helical*.{svg}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/2dStructure", mode: "copy", overwrite: true, pattern: '*.{garnier}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/HydrophobicMoment", mode: "copy", overwrite: true, pattern: '*HydrophobicMoments.{svg}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/IsoelectricPoint", mode: "copy", overwrite: true, pattern: '*IsoelectricPoint.{iep,svg}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/ProteinProperties", mode: "copy", overwrite: true, pattern: '*.{pepstats,pepinfo}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/ProteinProperties/Plots", mode: "copy", overwrite: true, pattern: '*PropertiesPlot.{svg}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/2dStructure/Plots", mode: "copy", overwrite: true, pattern: '*Helical*.{svg}' - input: - file(prot) from pcASVEMBOSS + input: + file(prot) from pcASVEMBOSS - output: - tuple file("*.garnier"), file("*HydrophobicMoments.svg"), file("*IsoelectricPoint*"), file("*.pepstats"), file("*PropertiesPlot*"), file("*Helical*") into pcASV_emboss + output: + tuple file("*.garnier"), file("*HydrophobicMoments.svg"), file("*IsoelectricPoint*"), file("*.pepstats"), file("*PropertiesPlot*"), file("*Helical*") into pcASV_emboss - script: - """ - for filename in ${prot};do - name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') - garnier -sequence \${filename} -outfile \${name}_2dStructures.garnier - hmoment -seqall \${filename} -graph svg -plot - mv hmoment.svg ./"\${name}"_HydrophobicMoments.svg - iep -sequence \${filename} -graph svg -plot -outfile "\${name}"_IsoelectricPoint.iep - mv iep.svg ./"\${name}"_IsoelectricPoint.svg - pepstats -sequence \${filename} -outfile \${name}_ProteinProperties.pepstats - grep ">" \${filename} | awk -F ">" '{print \$2}' > tmpsequence.list - for x in \$(cat tmpsequence.list);do - echo \$x > tmp1.list - seqtk subseq \${filename} tmp1.list > tmp2.fasta - len=\$(tail -1 tmp2.fasta | awk '{print length}') - pepinfo -sequence tmp2.fasta -graph svg -outfile "\$x"_PropertiesPlot.pepinfo - mv pepinfo.svg ./"\$x"_PropertiesPlot.svg - cat "\$x"_PropertiesPlot.pepinfo >> "\${name}"_PropertiesPlot.pepinfo - rm "\$x"_PropertiesPlot.pepinfo - pepnet -sask -sequence tmp2.fasta -graph svg -sbegin1 1 -send1 \$len - mv pepnet.svg ./"\$x"_HelicalNet.svg - pepwheel -sequence tmp2.fasta -graph svg -sbegin1 1 -send1 \$len - mv pepwheel.svg ./"\$x"_HelicalWheel.svg - rm tmp1.list tmp2.fasta - done - rm tmpsequence.list - done - """ + script: + """ + for filename in ${prot};do + name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') + garnier -sequence \${filename} -outfile \${name}_2dStructures.garnier + hmoment -seqall \${filename} -graph svg -plot + mv hmoment.svg ./"\${name}"_HydrophobicMoments.svg + iep -sequence \${filename} -graph svg -plot -outfile "\${name}"_IsoelectricPoint.iep + mv iep.svg ./"\${name}"_IsoelectricPoint.svg + pepstats -sequence \${filename} -outfile \${name}_ProteinProperties.pepstats + grep ">" \${filename} | awk -F ">" '{print \$2}' > tmpsequence.list + for x in \$(cat tmpsequence.list);do + echo \$x > tmp1.list + seqtk subseq \${filename} tmp1.list > tmp2.fasta + len=\$(tail -1 tmp2.fasta | awk '{print length}') + pepinfo -sequence tmp2.fasta -graph svg -outfile "\$x"_PropertiesPlot.pepinfo + mv pepinfo.svg ./"\$x"_PropertiesPlot.svg + cat "\$x"_PropertiesPlot.pepinfo >> "\${name}"_PropertiesPlot.pepinfo + rm "\$x"_PropertiesPlot.pepinfo + pepnet -sask -sequence tmp2.fasta -graph svg -sbegin1 1 -send1 \$len + mv pepnet.svg ./"\$x"_HelicalNet.svg + pepwheel -sequence tmp2.fasta -graph svg -sbegin1 1 -send1 \$len + mv pepwheel.svg ./"\$x"_HelicalWheel.svg + rm tmp1.list tmp2.fasta + done + rm tmpsequence.list + done + """ + } } - } - if (!params.skipTaxonomy) { - - process pcASV_AminoAcid_Taxonomy_Inference { + if (!params.skipTaxonomy) { - label 'high_cpus' + process pcASV_AminoAcid_Taxonomy_Inference { - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{csv,tsv}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*dmd.{out}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{fasta}' + label 'high_cpus' - input: - file(reads) from pcASV_aaDiamond_ch + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{csv,tsv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*dmd.{out}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{fasta}' - output: - file("*.fasta") into ( pcASV_labeledAA ) - tuple file("*phyloseqObject.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_potuaadiamond - file("*_summary_for_plot.csv") into taxplot4 + input: + file(reads) from pcASV_aaDiamond_ch - script: - """ - cp ${params.vampdir}/bin/rename_seq.py . - virdb=${params.dbdir}/${params.dbname} - grep ">" \${virdb} >> headers.list - headers="headers.list" - for filename in ${reads};do - name=\$(ls \${filename} | awk -F ".fasta" '{print \$1}') - diamond blastp -q \${filename} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 - echo "Preparing lists to generate summary .csv's" - echo "[Best hit accession number]" >access.list - echo "[pcASV sequence length]" >length.list - echo "[e-value]" >evalue.list - echo "[Bitscore]" >bit.list - echo "[Percent ID (aa)]" >pid.list - echo "[pcASVaa#]" >otu.list - echo "[Virus ID]" >"\$name"_virus.list - echo "[Gene]" >"\$name"_genes.list - grep ">" \${filename} | awk -F ">" '{print \$2}' > seqids.lst - echo "extracting genes and names" - touch new_"\$name"_headers.txt - j=1 - for s in \$(cat seqids.lst);do - echo "Checking for \$s hit in diamond output" - if [[ ${params.refseq} == "T" ]];then - echo "RefSeq headers specified" - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >>evalue.list - echo "\$line" | awk '{print \$11}' >>bit.list - echo "\$line" | awk '{print \$12}' >>pid.list - echo "\$line" | awk '{print \$2}' >>length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">pcASVaa\${j}_"\$virus"_"\$gene"" >> new_"\$name"_headers.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >>access.list - echo "NO_HIT" >>"\$name"_genes.list - echo "NO_HIT" >>"\$name"_virus.list - echo "NO_HIT" >>evalue.list - echo "NO_HIT" >>bit.list - echo "NO_HIT" >>pid.list - echo "NO_HIT" >>length.list - virus="NO" - gene="HIT" - echo ">pcASVaa\${j}_"\$virus"_"\$gene"" >> new_"\$name"_headers.txt - j=\$((\$j+1)) - echo "\$s done." - fi - else - echo "Using RVDB headers." - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') - echo "\$s" >>otu.list - echo "\$acc" >>access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >>evalue.list - echo "\$line" | awk '{print \$11}' >>bit.list - echo "\$line" | awk '{print \$12}' >>pid.list - echo "\$line" | awk '{print \$2}' >>length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >>"\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && - echo "\$virus" | sed 's/_/ /g' >>"\$name"_virus.list - echo ">pcASVaa\${j}_"\$virus"_"\$gene"" >>new_"\$name"_headers.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >>otu.list - echo "NO_HIT" >>access.list - echo "NO_HIT" >>"\$name"_genes.list - echo "NO_HIT" >>"\$name"_virus.list - echo "NO_HIT" >>evalue.list - echo "NO_HIT" >>bit.list - echo "NO_HIT" >>pid.list - echo "NO_HIT" >>length.list - virus="NO" - gene="HIT" - echo ">pcASVaa\${j}_\${virus}_\${gene}" >>new_"\$name"_headers.txt - j=\$((\$j+1)) - echo "\$s done." - fi - fi - echo "Done with \$s" - done - echo "Now editing "\$name" fasta headers" - ###### rename_seq.py - ./rename_seq.py \${filename} new_"\$name"_headers.txt "\$name"_wTax.fasta - awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_wTax.fasta > "\$name"_tmpssasv.fasta - echo "[Sequence header]" > newnames.list - cat new_"\$name"_headers.txt >> newnames.list - touch sequence.list - awk 'BEGIN{RS=">";ORS=""}{print \$2"\\n"}' \${name}_tmpssasv.fasta >>sequence.list - rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_summary_phyloseqObject.csv - paste -d"\\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - for x in *phyloseqObject.csv;do - echo "\$x" - lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) - tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; - sed 's/ /_/g' tmpcol.list > tmp2col.list; - cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; - rm tmpcol.list tmp2col.list - done - rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; - rm "\$name"_virus.list - rm "\$name"_genes.list - rm newnames.list - rm access.list - echo "Taxonomy inferred for: \${filename} " - done - rm *headers.list - """ - } - } + output: + file("*.fasta") into ( pcASV_labeledAA ) + tuple file("*phyloseqObject.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_potuaadiamond + file("*_summary_for_plot.csv") into taxplot4 - if (!params.skipPhylogeny) { + script: + """ + cp ${params.vampdir}/bin/rename_seq.py . + virdb=${params.dbdir}/${params.dbname} + grep ">" \${virdb} >> headers.list + headers="headers.list" + for filename in ${reads};do + name=\$(ls \${filename} | awk -F ".fasta" '{print \$1}') + diamond blastp -q \${filename} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + echo "Preparing lists to generate summary .csv's" + echo "[Best hit accession number]" >access.list + echo "[pcASV sequence length]" >length.list + echo "[e-value]" >evalue.list + echo "[Bitscore]" >bit.list + echo "[Percent ID (aa)]" >pid.list + echo "[pcASVaa#]" >otu.list + echo "[Virus ID]" >"\$name"_virus.list + echo "[Gene]" >"\$name"_genes.list + grep ">" \${filename} | awk -F ">" '{print \$2}' > seqids.lst + echo "extracting genes and names" + touch new_"\$name"_headers.txt + j=1 + for s in \$(cat seqids.lst);do + echo "Checking for \$s hit in diamond output" + if [[ ${params.refseq} == "T" ]];then + echo "RefSeq headers specified" + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >>evalue.list + echo "\$line" | awk '{print \$11}' >>bit.list + echo "\$line" | awk '{print \$12}' >>pid.list + echo "\$line" | awk '{print \$2}' >>length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">pcASVaa\${j}_"\$virus"_"\$gene"" >> new_"\$name"_headers.txt + j=\$((\$j+1)) + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >>access.list + echo "NO_HIT" >>"\$name"_genes.list + echo "NO_HIT" >>"\$name"_virus.list + echo "NO_HIT" >>evalue.list + echo "NO_HIT" >>bit.list + echo "NO_HIT" >>pid.list + echo "NO_HIT" >>length.list + virus="NO" + gene="HIT" + echo ">pcASVaa\${j}_"\$virus"_"\$gene"" >> new_"\$name"_headers.txt + j=\$((\$j+1)) + echo "\$s done." + fi + else + echo "Using RVDB headers." + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') + echo "\$s" >>otu.list + echo "\$acc" >>access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >>evalue.list + echo "\$line" | awk '{print \$11}' >>bit.list + echo "\$line" | awk '{print \$12}' >>pid.list + echo "\$line" | awk '{print \$2}' >>length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >>"\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && + echo "\$virus" | sed 's/_/ /g' >>"\$name"_virus.list + echo ">pcASVaa\${j}_"\$virus"_"\$gene"" >>new_"\$name"_headers.txt + j=\$((\$j+1)) + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >>otu.list + echo "NO_HIT" >>access.list + echo "NO_HIT" >>"\$name"_genes.list + echo "NO_HIT" >>"\$name"_virus.list + echo "NO_HIT" >>evalue.list + echo "NO_HIT" >>bit.list + echo "NO_HIT" >>pid.list + echo "NO_HIT" >>length.list + virus="NO" + gene="HIT" + echo ">pcASVaa\${j}_\${virus}_\${gene}" >>new_"\$name"_headers.txt + j=\$((\$j+1)) + echo "\$s done." + fi + fi + echo "Done with \$s" + done + echo "Now editing "\$name" fasta headers" + ###### rename_seq.py + ./rename_seq.py \${filename} new_"\$name"_headers.txt "\$name"_wTax.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_wTax.fasta > "\$name"_tmpssasv.fasta + echo "[Sequence header]" > newnames.list + cat new_"\$name"_headers.txt >> newnames.list + touch sequence.list + awk 'BEGIN{RS=">";ORS=""}{print \$2"\\n"}' \${name}_tmpssasv.fasta >>sequence.list + rm "\$name"_tmpssasv.fasta + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_summary_phyloseqObject.csv + paste -d"\\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + for x in *phyloseqObject.csv;do + echo "\$x" + lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) + tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; + sed 's/ /_/g' tmpcol.list > tmp2col.list; + cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; + rm tmpcol.list tmp2col.list + done + rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; + rm "\$name"_virus.list + rm "\$name"_genes.list + rm newnames.list + rm access.list + echo "Taxonomy inferred for: \${filename} " + done + rm *headers.list + """ + } + } - process pcASV_Protein_Phylogeny { + if (!params.skipPhylogeny) { - label 'norm_cpus' + process pcASV_Protein_Phylogeny { - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*aln.*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Phylogeny/Modeltest", mode: "copy", overwrite: true, pattern: '*mt*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*iq*' + label 'norm_cpus' - input: - file(prot) from pcASV_aaMafft_ch + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*aln.*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Phylogeny/Modeltest", mode: "copy", overwrite: true, pattern: '*mt*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*iq*' - output: - tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into pcASV_protein_phylogeny_results - file("*iq.treefile") into potu_Atree_plot + input: + file(prot) from pcASV_aaMafft_ch - script: - """ - for filename in ${prot};do - pre=\$( echo \${filename} | awk -F ".fasta" '{print \$1}' ) - mafft --maxiterate 5000 --auto \${filename} >\${pre}_ALN.fasta - trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html + output: + tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into pcASV_protein_phylogeny_results + file("*iq.treefile") into potu_Atree_plot - # pcASV_Protein_ModelTest - modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d aa -s 203 --disable-checkpoint + script: + """ + for filename in ${prot};do + pre=\$( echo \${filename} | awk -F ".fasta" '{print \$1}' ) + mafft --maxiterate 5000 --auto \${filename} >\${pre}_ALN.fasta + trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html - # pcASV_Protein_Phylogeny - if [ "${params.iqCustomaa}" != "" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomaa} + # pcASV_Protein_ModelTest + modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d aa -s 203 --disable-checkpoint - elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} + # pcASV_Protein_Phylogeny + if [ "${params.iqCustomaa}" != "" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomaa} - elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni + elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} - elif [ "${params.nonparametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} + elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni - elif [ "${params.parametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + elif [ "${params.nonparametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} - else - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - fi - done - """ - } - } + elif [ "${params.parametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - process Generate_pcASV_Protein_Counts { + else + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + fi + done + """ + } + } - label 'high_cpus' + process Generate_pcASV_Protein_Counts { - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Counts", mode: "copy", overwrite: true + label 'high_cpus' - input: - file(fasta) from pcASV_aaCounts_ch - file(merged) from mergeforpcASVaacounts - file(samplist) from samplistpotu + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Counts", mode: "copy", overwrite: true - output: - tuple file("*_counts.csv"), file("*dmd.out") into potuaacounts_summary - file("*counts.csv") into potu_Acounts + input: + file(fasta) from pcASV_aaCounts_ch + file(merged) from mergeforpcASVaacounts + file(samplist) from samplistpotu - script: - """ - set +e - for filename in ${fasta};do - potu="\$( echo \${filename} | awk -F "_" '{print \$3}')" - diamond makedb --in \${filename} --db \${filename} - diamond blastx -q ${merged} -d \${filename} -p ${task.cpus} --min-score ${params.ProtCountsBit} --id ${params.ProtCountID} -l ${params.ProtCountsLength} --more-sensitive -o ${params.projtag}_\${potu}_Counts_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 --max-hsps 1 - echo "OTU_ID" >tmp.col1.txt - echo "Generating sample id list" - grep ">" \${filename} | awk -F ">" '{print \$2}' | sort | uniq > otuid.list - cat otuid.list >> tmp.col1.txt - echo "Beginning them counts tho my g" - for y in \$( cat ${samplist} );do - echo "Starting with \$y now ..." - grep "\$y" ${params.projtag}_\${potu}_Counts_dmd.out > tmp."\$y".out - echo "Isolated hits" - echo "Created uniq subject id list" - echo "\$y" > "\$y"_col.txt - echo "Starting my counts" - for z in \$(cat otuid.list);do - echo "Counting \$z hits" - echo "grep -wc "\$z" >> "\$y"_col.txt" - grep -wc "\$z" tmp."\$y".out >> "\$y"_col.txt - echo "\$z counted" - done - done - paste -d "," tmp.col1.txt *col.txt > ${params.projtag}_\${potu}_counts.csv - rm tmp* - rm *col.txt - done - """ - } - } + output: + tuple file("*_counts.csv"), file("*dmd.out") into potuaacounts_summary + file("*counts.csv") into potu_Acounts - if (!params.skipReport) { + script: + """ + set +e + for filename in ${fasta};do + potu="\$( echo \${filename} | awk -F "_" '{print \$3}')" + diamond makedb --in \${filename} --db \${filename} + diamond blastx -q ${merged} -d \${filename} -p ${task.cpus} --min-score ${params.ProtCountsBit} --id ${params.ProtCountID} -l ${params.ProtCountsLength} --more-sensitive -o ${params.projtag}_\${potu}_Counts_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 --max-hsps 1 + echo "OTU_ID" >tmp.col1.txt + echo "Generating sample id list" + grep ">" \${filename} | awk -F ">" '{print \$2}' | sort | uniq > otuid.list + cat otuid.list >> tmp.col1.txt + echo "Beginning them counts tho my g" + for y in \$( cat ${samplist} );do + echo "Starting with \$y now ..." + grep "\$y" ${params.projtag}_\${potu}_Counts_dmd.out > tmp."\$y".out + echo "Isolated hits" + echo "Created uniq subject id list" + echo "\$y" > "\$y"_col.txt + echo "Starting my counts" + for z in \$(cat otuid.list);do + echo "Counting \$z hits" + echo "grep -wc "\$z" >> "\$y"_col.txt" + grep -wc "\$z" tmp."\$y".out >> "\$y"_col.txt + echo "\$z counted" + done + done + paste -d "," tmp.col1.txt *col.txt > ${params.projtag}_\${potu}_counts.csv + rm tmp* + rm *col.txt + done + """ + } + } - if (!params.skipAdapterRemoval) { + if (!params.skipReport) { - process combine_csv { + if (!params.skipAdapterRemoval) { - input: - file(csv) from fastp_csv - .collect() + process combine_csv { - output: - file("final_reads_stats.csv") into ( fastp_csv1, fastp_csv2, fastp_csv3, fastp_csv4, fastp_csv5 ) + input: + file(csv) from fastp_csv_in2 + .collect() - script: - """ - cat ${csv} >all_reads_stats.csv - head -n1 all_reads_stats.csv >tmp.names.csv - cat all_reads_stats.csv | grep -v ""Sample,Total_"" >tmp.reads.stats.csv - cat tmp.names.csv tmp.reads.stats.csv >final_reads_stats.csv - rm tmp.names.csv tmp.reads.stats.csv - """ + output: + file("final_reads_stats.csv") into ( fastp_csv1, fastp_csv2, fastp_csv3, fastp_csv4, fastp_csv5 ) - } - } + script: + """ + cat ${csv} >all_reads_stats.csv + head -n1 all_reads_stats.csv >tmp.names.csv + cat all_reads_stats.csv | grep -v ""Sample,Total_"" >tmp.reads.stats.csv + cat tmp.names.csv tmp.reads.stats.csv >final_reads_stats.csv + rm tmp.names.csv tmp.reads.stats.csv + """ - if (params.ncASV) { + } + } - // mopd this - process Report_ASV { + if (params.ncASV) { - label 'norm_cpus' + // mopd this + process Report_ASV { - publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport", mode: "copy", overwrite: true + label 'norm_cpus' - input: - file(counts) from asv_counts_plots - file(taxonomy) from taxplot1 - file(matrix) from asv_heatmap - file(readsstats) from fastp_csv1 - - output: - file("*.html") into report_summaryA - - script: - """ - name=\$( echo ${taxonomy} | awk -F "_summary_for_plot.csv" '{print \$1}') - cp ${params.vampdir}/bin/vAMPirus_ReportA.Rmd . - cp ${params.vampdir}/example_data/conf/vamplogo.png . - Rscript -e "rmarkdown::render('vAMPirus_ReportA.Rmd',output_file='\${name}_ASV_Report.html')" \${name} \ - ${readsstats} \ - ${counts} \ - ${params.metadata} \ - ${params.minimumCounts} \ - ${matrix} \ - ${taxonomy} \ - ${params.trymax} \ - ${params.stats} - """ - } - - process Report_ncASV { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport/ncASV", mode: "copy", overwrite: true - - input: - file(counts) from notu_counts_plots - file(taxonomy) from taxplot1a - file(matrix) from notu_heatmap - file(phylogeny) from nucl_phyl_plot - file(readsstats) from fastp_csv2 - - output: - file("*.html") into report_summaryB - - script: - """ - cp ${params.vampdir}/bin/vAMPirus_ReportB.Rmd . - cp ${params.vampdir}/example_data/conf/vamplogo.png . - for x in *_summary_for_plot.csv;do - name=\$( echo \${x} | awk -F "_summary_for_plot.csv" '{print \$1}') - id=\$( echo \${x} | awk -F "_summary_for_plot.csv" '{print \$1}' | cut -f 2 -d "." ) - Rscript -e "rmarkdown::render('vAMPirus_ReportB.Rmd',output_file='\${name}_ncASV\${id}_Report.html')" \${name} \ - ${readsstats} \ - \$( echo ${counts} | tr " " "\\n" | grep "\${id}" ) \ - ${params.metadata} \ - ${params.minimumCounts} \ - \$( echo ${matrix} | tr " " "\\n" | grep "\${id}" ) \ - \$( echo ${taxonomy} | tr " " "\\n" | grep "\${id}" ) \ - \$( echo ${phylogeny} | tr " " "\\n" | grep "\${id}" ) \ - ${params.trymax} \ - ${params.stats} - done - """ - } - - if (!params.skipAminoTyping) { - - process Report_AminoTypes { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport", mode: "copy", overwrite: true - - input: - file(counts) from aminocounts_plot - file(taxonomy) from taxplot2 - file(matrix) from aminotype_heatmap - file(phylogeny) from amino_rax_plot - file(readsstats) from fastp_csv5 - - output: - file("*.html") into report_summaryE - - script: - """ - name=\$( echo ${taxonomy} | awk -F "_summary_for_plot.csv" '{print \$1}') - cp ${params.vampdir}/bin/vAMPirus_ReportB.Rmd . - cp ${params.vampdir}/example_data/conf/vamplogo.png . - Rscript -e "rmarkdown::render('vAMPirus_ReportB.Rmd',output_file='\${name}_AminoType_Report.html')" \${name} \ - ${readsstats} \ - ${counts} \ - ${params.metadata} \ - ${params.minimumCounts} ${matrix} \ - ${taxonomy} \ - ${phylogeny} \ - ${params.trymax} \ - ${params.stats} - """ - } - } - } else { - - process Report_ASVs { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport", mode: "copy", overwrite: true input: file(counts) from asv_counts_plots file(taxonomy) from taxplot1 file(matrix) from asv_heatmap - file(phylogeny) from nucl_phyl_plot file(readsstats) from fastp_csv1 output: @@ -3072,24 +3291,60 @@ if (params.Analyze) { script: """ name=\$( echo ${taxonomy} | awk -F "_summary_for_plot.csv" '{print \$1}') - cp ${params.vampdir}/bin/vAMPirus_ReportB.Rmd . + cp ${params.vampdir}/bin/vAMPirus_ReportA.Rmd . cp ${params.vampdir}/example_data/conf/vamplogo.png . - Rscript -e "rmarkdown::render('vAMPirus_ReportB.Rmd',output_file='\${name}_ASV_Report.html')" \${name} \ + Rscript -e "rmarkdown::render('vAMPirus_ReportA.Rmd',output_file='\${name}_ASV_Report.html')" \${name} \ ${readsstats} \ ${counts} \ ${params.metadata} \ ${params.minimumCounts} \ - ${matrix} \ + ${matrix} \ ${taxonomy} \ - ${phylogeny} \ ${params.trymax} \ ${params.stats} """ - } + } + + process Report_ncASV { + + label 'norm_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport/ncASV", mode: "copy", overwrite: true + + input: + file(counts) from notu_counts_plots + file(taxonomy) from taxplot1a + file(matrix) from notu_heatmap + file(phylogeny) from nucl_phyl_plot + file(readsstats) from fastp_csv2 + + output: + file("*.html") into report_summaryB + + script: + """ + cp ${params.vampdir}/bin/vAMPirus_ReportB.Rmd . + cp ${params.vampdir}/example_data/conf/vamplogo.png . + for x in *_summary_for_plot.csv;do + name=\$( echo \${x} | awk -F "_summary_for_plot.csv" '{print \$1}') + id=\$( echo \${x} | awk -F "_summary_for_plot.csv" '{print \$1}' | cut -f 2 -d "." ) + Rscript -e "rmarkdown::render('vAMPirus_ReportB.Rmd',output_file='\${name}_ncASV\${id}_Report.html')" \${name} \ + ${readsstats} \ + \$( echo ${counts} | tr " " "\\n" | grep "\${id}" ) \ + ${params.metadata} \ + ${params.minimumCounts} \ + \$( echo ${matrix} | tr " " "\\n" | grep "\${id}" ) \ + \$( echo ${taxonomy} | tr " " "\\n" | grep "\${id}" ) \ + \$( echo ${phylogeny} | tr " " "\\n" | grep "\${id}" ) \ + ${params.trymax} \ + ${params.stats} + done + """ + } if (!params.skipAminoTyping) { - process Report_AminoType { + process Report_AminoTypes { label 'norm_cpus' @@ -3120,632 +3375,159 @@ if (params.Analyze) { ${params.trymax} \ ${params.stats} """ + } } - } - } - - if (params.pcASV) { - - process Report_pcASV_AminoAcid { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport/pcASV/Aminoacid", mode: "copy", overwrite: true - - input: - file(counts) from potu_Acounts - file(taxonomy) from taxplot4 - file(matrix) from potu_aa_heatmap - file(phylogeny) from potu_Atree_plot - file(readsstats) from fastp_csv3 - - output: - file("*.html") into report_summaryC - - script: - """ - cp ${params.vampdir}/bin/vAMPirus_ReportB.Rmd . - cp ${params.vampdir}/example_data/conf/vamplogo.png . - for x in *_summary_for_plot.csv;do - name=\$( echo \${x} | awk -F "_noTaxonomy_summary_for_plot.csv" '{print \$1}') - id=\$( echo \${x} | awk -F "_noTaxonomy_summary_for_plot.csv" '{print \$1}' | cut -f 2 -d "." ) - Rscript -e "rmarkdown::render('vAMPirus_ReportB.Rmd',output_file='\${name}_pcASVaa\${id}_Report.html')" \${name} \ - ${readsstats} \ - \$( echo ${counts} | tr " " "\\n" | grep "\${id}" ) \ - ${params.metadata} \ - ${params.minimumCounts} \ - \$( echo ${matrix} | tr " " "\\n" | grep "\${id}" ) \ - \$( echo ${taxonomy} | tr " " "\\n" | grep "\${id}" ) \ - \$( echo ${phylogeny} | tr " " "\\n" | grep "\${id}" ) \ - ${params.trymax} \ - ${params.stats} - done - """ - } - - process Report_pcASV_Nucleotide { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport/pcASV/Nucleotide", mode: "copy", overwrite: true - - input: - file(counts) from potu_Ncounts_for_report - file(taxonomy) from taxplot3 - file(matrix) from potu_nucl_heatmap - file(phylogeny) from potu_Ntree_plot - file(readsstats) from fastp_csv4 - - output: - file("*.html") into report_summaryD - - script: - """ - cp ${params.vampdir}/bin/vAMPirus_ReportB.Rmd . - cp ${params.vampdir}/example_data/conf/vamplogo.png . - for x in *_summary_for_plot.csv;do - name=\$( echo \${x} | awk -F "_summary_for_plot.csv" '{print \$1}') - id=\$( echo \${x} | awk -F "_summary_for_plot.csv" '{print \$1}' | cut -f 2 -d "." ) - Rscript -e "rmarkdown::render('vAMPirus_ReportB.Rmd',output_file='\${name}_pcASVnt\${id}_Report.html')" \${name} \ - ${readsstats} \ - \$( echo ${counts} | tr " " "\\n" | grep "\${id}" ) \ - ${params.metadata} \ - ${params.minimumCounts} \ - \$( echo ${matrix} | tr " " "\\n" | grep "\${id}" ) \ - \$( echo ${taxonomy} | tr " " "\\n" | grep "\${id}" ) \ - \$( echo ${phylogeny} | tr " " "\\n" | grep "\${id}" ) \ - ${params.trymax} \ - ${params.stats} - done - """ - } - } - } - -} else if (params.DataCheck) { - - println("\n\tRunning vAMPirus DataCheck \n") - - process QualityCheck_1DC { - - label 'low_cpus' - - tag "${sample_id}" - - publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/FastQC/PreClean", mode: "copy", overwrite: true - - input: - tuple sample_id, file(reads) from reads_qc_ch - - output: - tuple sample_id, file("*_fastqc.{zip,html}") into fastqc_results_OAS - - script: - """ - fastqc --quiet --threads ${task.cpus} ${reads} - """ - } + } else { - process Adapter_Removal_DC { + process Report_ASVs { + + label 'norm_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport", mode: "copy", overwrite: true + + input: + file(counts) from asv_counts_plots + file(taxonomy) from taxplot1 + file(matrix) from asv_heatmap + file(phylogeny) from nucl_phyl_plot + file(readsstats) from fastp_csv1 + + output: + file("*.html") into report_summaryA + + script: + """ + name=\$( echo ${taxonomy} | awk -F "_summary_for_plot.csv" '{print \$1}') + cp ${params.vampdir}/bin/vAMPirus_ReportB.Rmd . + cp ${params.vampdir}/example_data/conf/vamplogo.png . + Rscript -e "rmarkdown::render('vAMPirus_ReportB.Rmd',output_file='\${name}_ASV_Report.html')" \${name} \ + ${readsstats} \ + ${counts} \ + ${params.metadata} \ + ${params.minimumCounts} \ + ${matrix} \ + ${taxonomy} \ + ${phylogeny} \ + ${params.trymax} \ + ${params.stats} + """ + } + + if (!params.skipAminoTyping) { + + process Report_AminoType { + + label 'norm_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport", mode: "copy", overwrite: true + + input: + file(counts) from aminocounts_plot + file(taxonomy) from taxplot2 + file(matrix) from aminotype_heatmap + file(phylogeny) from amino_rax_plot + file(readsstats) from fastp_csv5 + + output: + file("*.html") into report_summaryE + + script: + """ + name=\$( echo ${taxonomy} | awk -F "_summary_for_plot.csv" '{print \$1}') + cp ${params.vampdir}/bin/vAMPirus_ReportB.Rmd . + cp ${params.vampdir}/example_data/conf/vamplogo.png . + Rscript -e "rmarkdown::render('vAMPirus_ReportB.Rmd',output_file='\${name}_AminoType_Report.html')" \${name} \ + ${readsstats} \ + ${counts} \ + ${params.metadata} \ + ${params.minimumCounts} ${matrix} \ + ${taxonomy} \ + ${phylogeny} \ + ${params.trymax} \ + ${params.stats} + """ + } + } + } - label 'norm_cpus' + if (params.pcASV) { - tag "${sample_id}" + process Report_pcASV_AminoAcid { - publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/AdapterRemoval", mode: "copy", overwrite: true, pattern: "*.filter.fq" - publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/AdapterRemoval/fastpOut", mode: "copy", overwrite: true, pattern: "*.fastp.{json,html}" - - input: - tuple sample_id, file(reads) from reads_ch + label 'norm_cpus' - output: - tuple sample_id, file("*.fastp.{json,html}") into fastp_results - tuple sample_id, file("*.filter.fq") into reads_fastp_ch - file("*.csv") into fastp_csv + publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport/pcASV/Aminoacid", mode: "copy", overwrite: true - script: - """ - echo ${sample_id} - - fastp -i ${reads[0]} -I ${reads[1]} -o left-${sample_id}.filter.fq -O right-${sample_id}.filter.fq --detect_adapter_for_pe \ - --average_qual 25 -c --overrepresentation_analysis --html ${sample_id}.fastp.html --json ${sample_id}.fastp.json --thread ${task.cpus} \ - --report_title ${sample_id} + input: + file(counts) from potu_Acounts + file(taxonomy) from taxplot4 + file(matrix) from potu_aa_heatmap + file(phylogeny) from potu_Atree_plot + file(readsstats) from fastp_csv3 - bash get_readstats.sh ${sample_id}.fastp.json - """ - } + output: + file("*.html") into report_summaryC - process Primer_Removal_DC { + script: + """ + cp ${params.vampdir}/bin/vAMPirus_ReportB.Rmd . + cp ${params.vampdir}/example_data/conf/vamplogo.png . + for x in *_summary_for_plot.csv;do + name=\$( echo \${x} | awk -F "_noTaxonomy_summary_for_plot.csv" '{print \$1}') + id=\$( echo \${x} | awk -F "_noTaxonomy_summary_for_plot.csv" '{print \$1}' | cut -f 2 -d "." ) + Rscript -e "rmarkdown::render('vAMPirus_ReportB.Rmd',output_file='\${name}_pcASVaa\${id}_Report.html')" \${name} \ + ${readsstats} \ + \$( echo ${counts} | tr " " "\\n" | grep "\${id}" ) \ + ${params.metadata} \ + ${params.minimumCounts} \ + \$( echo ${matrix} | tr " " "\\n" | grep "\${id}" ) \ + \$( echo ${taxonomy} | tr " " "\\n" | grep "\${id}" ) \ + \$( echo ${phylogeny} | tr " " "\\n" | grep "\${id}" ) \ + ${params.trymax} \ + ${params.stats} + done + """ + } - label 'norm_cpus' + process Report_pcASV_Nucleotide { - tag "${sample_id}" + label 'norm_cpus' - publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/PrimerRemoval", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport/pcASV/Nucleotide", mode: "copy", overwrite: true - input: - tuple sample_id, file(reads) from reads_fastp_ch + input: + file(counts) from potu_Ncounts_for_report + file(taxonomy) from taxplot3 + file(matrix) from potu_nucl_heatmap + file(phylogeny) from potu_Ntree_plot + file(readsstats) from fastp_csv4 - output: - tuple sample_id, file("*bbduk*.fastq.gz") into ( reads_bbduk_ch, readsforqc2 ) + output: + file("*.html") into report_summaryD - script: - // add cocktail primer removal, easy, set a list, line by line seperated by , in a for loop - if ( params.fwd == "" && params.rev == "" && !params.multi) { - """ - bbduk.sh in1=${reads[0]} out=${sample_id}_bb_R1.fastq.gz ftl=${params.defaultFwdTrim} t=${task.cpus} - bbduk.sh in=${reads[1]} out=${sample_id}_bb_R2.fastq.gz ftl=${params.defaultRevTrim} t=${task.cpus} - repair.sh in1=${sample_id}_bb_R1.fastq.gz in2=${sample_id}_bb_R2.fastq.gz out1=${sample_id}_bbduk_R1.fastq.gz out2=${sample_id}_bbduk_R2.fastq.gz minlength=${params.minilen} outs=sing.fq repair - """ - } else if ( params.GlobTrim && !params.GlobTrim == "" ) { - """ - FTRIM=\$( echo ${GlobTrim} | cut -f 1 -d "," ) - RTRIM=\$( echo ${GlobTrim} | cut -f 2 -d "," ) - bbduk.sh in=${reads[0]} out=${sample_id}_bb_R1.fastq.gz ftl=\${FTRIM} t=${task.cpus} - bbduk.sh in=${reads[1]} out=${sample_id}_bb_R2.fastq.gz ftl=\${RTRIM} t=${task.cpus} - repair.sh in1=${sample_id}_bb_R1.fastq.gz in2=${sample_id}_bb_R2.fastq.gz out1=${sample_id}_bbduk_R1.fastq.gz out2=${sample_id}_bbduk_R2.fastq.gz minlength=${params.minilen} outs=sing.fq repair - """ - } else if ( params.multi && params.primers ) { - """ - bbduk.sh in=${reads[0]} in2=${reads[1]} out=${sample_id}_bbduk_R1.fastq.gz out2=${sample_id}_bbduk_R2.fastq.gz ref=${params.primers} copyundefined=t t=${task.cpus} restrictleft=${params.primerLength} k=${params.maxkmer} ordered=t mink=${params.minkmer} ktrim=l ecco=t rcomp=t minlength=${params.minilen} tbo tpe - """ - } else { - """ - bbduk.sh in=${reads[0]} in2=${reads[1]} out=${sample_id}_bbduk_R1.fastq.gz out2=${sample_id}_bbduk_R2.fastq.gz literal=${params.fwd},${params.rev} copyundefined=t t=${task.cpus} restrictleft=${params.primerLength} k=${params.maxkmer} ordered=t mink=${params.minkmer} ktrim=l ecco=t rcomp=t minlength=${params.minilen} tbo tpe - """ + script: + """ + cp ${params.vampdir}/bin/vAMPirus_ReportB.Rmd . + cp ${params.vampdir}/example_data/conf/vamplogo.png . + for x in *_summary_for_plot.csv;do + name=\$( echo \${x} | awk -F "_summary_for_plot.csv" '{print \$1}') + id=\$( echo \${x} | awk -F "_summary_for_plot.csv" '{print \$1}' | cut -f 2 -d "." ) + Rscript -e "rmarkdown::render('vAMPirus_ReportB.Rmd',output_file='\${name}_pcASVnt\${id}_Report.html')" \${name} \ + ${readsstats} \ + \$( echo ${counts} | tr " " "\\n" | grep "\${id}" ) \ + ${params.metadata} \ + ${params.minimumCounts} \ + \$( echo ${matrix} | tr " " "\\n" | grep "\${id}" ) \ + \$( echo ${taxonomy} | tr " " "\\n" | grep "\${id}" ) \ + \$( echo ${phylogeny} | tr " " "\\n" | grep "\${id}" ) \ + ${params.trymax} \ + ${params.stats} + done + """ + } + } } - } - - process QualityCheck_2_DC { - - label 'low_cpus' - - tag "${sample_id}" - - publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/FastQC/PostClean", mode: "copy", overwrite: true - - input: - tuple sample_id, file(reads) from readsforqc2 - - output: - tuple sample_id, file("*_fastqc.{zip,html}") into fastqc2_results_OAS - - script: - """ - fastqc --quiet --threads ${task.cpus} ${reads} - """ } - - process Read_Merging_DC { - - label 'norm_cpus' - - tag "${sample_id}" - - publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/ReadMerging/Individual", mode: "copy", overwrite: true, pattern: "*mergedclean.fastq" - publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/ReadMerging/Individual/notmerged", mode: "copy", overwrite: true, pattern: "*notmerged*.fastq" - - input: - tuple sample_id, file(reads) from reads_bbduk_ch - - output: - file("*_mergedclean.fastq") into reads_vsearch1_ch - file("*.name") into names - file("*notmerged*.fastq") into notmerged - - script: - """ - vsearch --fastq_mergepairs ${reads[0]} --reverse ${reads[1]} --threads ${task.cpus} --fastqout ${sample_id}_mergedclean.fastq --fastqout_notmerged_fwd ${sample_id}_notmerged_fwd.fastq --fastqout_notmerged_rev ${sample_id}_notmerged_rev.fastq --fastq_maxee ${params.maxEE} --relabel ${sample_id}. - echo ${sample_id} > ${sample_id}.name - """ - - } - - process Compile_Reads_DC { - - label 'low_cpus' - - publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/ReadMerging/LengthFiltering", mode: "copy", overwrite: true - - input: - file(reads) from reads_vsearch1_ch - .collect() - - output: - file("*_all_merged_preFilt_preClean.fastq") into collect_samples_ch - - script: - """ - cat ${reads} >>${params.projtag}_all_merged_preFilt_preClean.fastq - """ - } - - process Compile_Names_DC { - - label 'low_cpus' - - publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/ReadMerging", mode: "copy", overwrite: true - - input: - file(names) from names - .collect() - - output: - file("*sample_ids.list") into ( samplelist, samplistpotu ) - - script: - """ - cat ${names} >>${params.projtag}_sample_ids.list - """ - } - - process Length_Filtering_DC { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/ReadMerging/LengthFiltering", mode: "copy", overwrite: true, pattern: "*_merged_preFilt*.fasta" - publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/ReadMerging", mode: "copy", overwrite: true, pattern: "*Lengthfiltered.fastq" - publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/ReadMerging/Histograms/pre_length_filtering", mode: "copy", overwrite: true, pattern: "*preFilt_*st.txt" - publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/ReadMerging/Histograms/post_length_filtering", mode: "copy", overwrite: true, pattern: "*postFilt_*st.txt" - - input: - file(reads) from collect_samples_ch - - output: - file("*_merged_preFilt_clean.fasta") into ( nuclCounts_mergedreads_ch, pcASV_mergedreads_ch ) - file("*_merged_clean_Lengthfiltered.fastq") into reads_vsearch2_ch - - file("*preFilt_preClean_baseFrequency_hist.csv") into prefilt_basefreq - file("*preFilt_preClean_qualityScore_hist.csv") into prefilt_qualityscore - file("*preFilt_preClean_gcContent_hist.csv") into prefilt_gccontent - file("*preFilt_preClean_averageQuality_hist.csv") into prefilt_averagequality - file("*preFilt_preClean_length_hist.csv") into prefilt_length - - file("*postFilt_baseFrequency_hist.csv") into postFilt_basefreq - file("*postFilt_qualityScore_hist.csv") into postFilt_qualityscore - file("*postFilt_gcContent_hist.csv") into postFilt_gccontent - file("*postFilt_averageQuaulity_hist.csv") into postFilt_averagequality - file("*postFilt_length_hist.csv") into postFilt_length - - file("reads_per_sample_preFilt_preClean.csv") into reads_per_sample_preFilt - file("read_per_sample_postFilt_postClean.csv") into reads_per_sample_postFilt - script: - """ - bbduk.sh in=${reads} bhist=${params.projtag}_all_merged_preFilt_preClean_baseFrequency_hist.txt qhist=${params.projtag}_all_merged_preFilt_preClean_qualityScore_hist.txt gchist=${params.projtag}_all_merged_preFilt_preClean_gcContent_hist.txt aqhist=${params.projtag}_all_merged_preFilt_preClean_averageQuality_hist.txt lhist=${params.projtag}_all_merged_preFilt_preClean_length_hist.txt gcbins=auto - for x in *preFilt*hist.txt;do - pre=\$(echo \$x | awk -F ".txt" '{print \$1}') - cat \$x | tr "\t" "," > \${pre}.csv - rm \$x - done - reformat.sh in=${reads} out=${params.projtag}_preFilt_preclean.fasta t=${task.cpus} - echo "sample,reads" >> reads_per_sample_preFilt_preClean.csv - grep ">" ${params.projtag}_preFilt_preclean.fasta | awk -F ">" '{print \$2}' | awk -F "." '{print \$1}' | sort --parallel=${task.cpus} | uniq -c | sort -brg --parallel=${task.cpus} | awk '{print \$2","\$1}' >> reads_per_sample_preFilt_preClean.csv - rm ${params.projtag}_preFilt_preclean.fasta - fastp -i ${reads} -o ${params.projtag}_merged_preFilt_clean.fastq -b ${params.maxLen} -l ${params.minLen} --thread ${task.cpus} -n 1 - reformat.sh in=${params.projtag}_merged_preFilt_clean.fastq out=${params.projtag}_merged_preFilt_clean.fasta t=${task.cpus} - bbduk.sh in=${params.projtag}_merged_preFilt_clean.fastq out=${params.projtag}_merged_clean_Lengthfiltered.fastq minlength=${params.maxLen} maxlength=${params.maxLen} t=${task.cpus} - bbduk.sh in=${params.projtag}_merged_clean_Lengthfiltered.fastq bhist=${params.projtag}_all_merged_postFilt_baseFrequency_hist.txt qhist=${params.projtag}_all_merged_postFilt_qualityScore_hist.txt gchist=${params.projtag}_all_merged_postFilt_gcContent_hist.txt aqhist=${params.projtag}_all_merged_postFilt_averageQuaulity_hist.txt lhist=${params.projtag}_all_merged_postFilt_length_hist.txt gcbins=auto - for x in *postFilt*hist.txt;do - pre=\$(echo \$x | awk -F ".txt" '{print \$1}') - cat \$x | tr "\t" "," > \${pre}.csv - rm \$x - done - reformat.sh in=${params.projtag}_merged_clean_Lengthfiltered.fastq out=${params.projtag}_merged_clean_Lengthfiltered.fasta t=${task.cpus} - echo "sample,reads" >> read_per_sample_postFilt_postClean.csv - grep ">" ${params.projtag}_merged_clean_Lengthfiltered.fasta | awk -F ">" '{print \$2}' | awk -F "." '{print \$1}' | sort --parallel=${task.cpus} | uniq -c | sort -brg --parallel=${task.cpus} | awk '{print \$2","\$1}' >> read_per_sample_postFilt_postClean.csv - """ - } - - process Extract_Uniques_DC { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/ReadMerging/Uniques", mode: "copy", overwrite: true - - input: - file(reads) from reads_vsearch2_ch - - output: - file("*unique_sequences.fasta") into reads_vsearch3_ch - - script: - """ - vsearch --derep_fulllength ${reads} --sizeout --relabel_keep --output ${params.projtag}_unique_sequences.fasta - """ - } - - process Identify_ASVs_DC { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/ASVs/ChimeraCheck", mode: "copy", overwrite: true - - input: - file(reads) from reads_vsearch3_ch - - output: - file("*notChecked.fasta") into reads_vsearch4_ch - - script: - """ - vsearch --cluster_unoise ${reads} --unoise_alpha ${params.alpha} --relabel ASV --centroids ${params.projtag}_notChecked.fasta --minsize ${params.minSize} - """ - } - - process Chimera_Check_DC { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/ASVs", mode: "copy", overwrite: true - - input: - file(fasta) from reads_vsearch4_ch - - output: - file("*ASVs.fasta") into ( reads_vsearch5_ch, nucl2aa, asvsforAminotyping, asvfastaforcounts, asvaminocheck ) - - script: - """ - vsearch --uchime3_denovo ${fasta} --relabel ASV --nonchimeras ${params.projtag}_ASVs.fasta - """ - } - - process NucleotideBased_ASV_clustering_DC { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Nucleotide", mode: "copy", overwrite: true, pattern: '*{.csv}' - - input: - file(fasta) from reads_vsearch5_ch - - output: - file("number_per_percentage_nucl.csv") into number_per_percent_nucl_plot - - script: - if (params.datacheckntIDlist) { - """ - for id in `echo ${params.datacheckntIDlist} | tr "," "\\n"`;do - vsearch --cluster_fast ${fasta} --centroids ${params.projtag}_ncASV\${id}.fasta --threads ${task.cpus} --relabel OTU --id \${id} - done - for x in *ncASV*.fasta;do - id=\$( echo \$x | awk -F "_ncASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') - numb=\$( grep -c ">" \$x ) - echo "\${id},\${numb}" >> number_per_percentage_nucl.csv - done - yo=\$(grep -c ">" ${fasta}) - echo "1.0,\${yo}" >> number_per_percentage_nucl.csv - """ - } - } - - - if (params.sing) { - - process Translating_For_ProteinClustering_DC { - - label 'low_cpus' - - publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Aminoacid/translation", mode: "copy", overwrite: true - - input: - file(fasta) from nucl2aa - - output: - file("*ASVprotforclust.fasta") into clustering_aa - file("*_translation_report") into reportaa_VR - file("*_ASV_all.fasta") into asvfastaforaaclust - - script: - """ - conda init && source activate virtualribosome - - ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_ASVprotforclust.fasta --report ${params.projtag}_translation_report - cp ${fasta} ${params.projtag}_ASV_all.fasta - - """ - - } - - } else { - - process Translation_For_ProteinBased_Clustering_DC { - - label 'norm_cpus' - - conda 'python=2.7' - - publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Aminoacid/translation", mode: "copy", overwrite: true - - input: - file(fasta) from nucl2aa - - output: - file("*ASVprotforclust.fasta") into clustering_aa - file("*_translation_report") into reportaa_VR - file("*_ASV_all.fasta") into asvfastaforaaclust - - script: - """ - ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_ASVprotforclust.fasta --report ${params.projtag}_translation_report - cp ${fasta} ${params.projtag}_ASV_all.fasta - """ - } - } - - process Protein_clustering_DC { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Aminoacid", mode: "copy", overwrite: true, pattern: '*{.csv}' - - input: - file(fasta) from clustering_aa - file(asvs) from asvfastaforaaclust - - output: - file("number_per_percentage_prot.csv") into number_per_percent_prot_plot - - script: - // add awk script to count seqs - """ - set +e - cp ${params.vampdir}/bin/rename_seq.py . - for id in `echo ${params.datacheckaaIDlist} | tr "," "\\n"`;do - if [ \${id} == ".55" ];then - word=3 - elif [ \${id} == ".65" ];then - word=4 - else - word=5 - fi - awk 'BEGIN{RS=">";ORS=""}length(\$2)>="${params.minAA}"{print ">"\$0}' ${fasta} > ${params.projtag}_filtered_proteins.fasta - cd-hit -i ${params.projtag}_filtered_proteins.fasta -n \${word} -c \${id} -o ${params.projtag}_pcASV\${id}.fasta - sed 's/>Cluster />Cluster_/g' ${params.projtag}_pcASV\${id}.fasta.clstr >${params.projtag}_pcASV\${id}.clstr - grep ">Cluster_" ${params.projtag}_pcASV\${id}.clstr >temporaryclusters.list - y=\$(grep -c ">Cluster_" ${params.projtag}_pcASV\${id}.clstr) - echo ">Cluster_"\${y}"" >> ${params.projtag}_pcASV\${id}.clstr - t=1 - b=1 - for x in \$(cat temporaryclusters.list);do - echo "Extracting \$x" - name="\$( echo \$x | awk -F ">" '{print \$2}')" - clust="pcASV"\${t}"" - echo "\${name}" - awk '/^>'\${name}'\$/,/^>Cluster_'\${b}'\$/' ${params.projtag}_pcASV\${id}.clstr > "\${name}"_"\${clust}"_tmp.list - t=\$(( \${t}+1 )) - b=\$(( \${b}+1 )) - done - ls *_tmp.list - u=1 - for x in *_tmp.list;do - name="\$(echo \$x | awk -F "_p" '{print \$1}')" - echo "\${name}" - cluster="\$(echo \$x | awk -F "_" '{print \$3}')" - echo "\${cluster}" - grep "ASV" \$x | awk -F ", " '{print \$2}' | awk -F "_" '{print \$1}' | awk -F ">" '{print \$2}' > \${name}_\${cluster}_seqs_tmps.list - seqtk subseq ${asvs} \${name}_\${cluster}_seqs_tmps.list > \${name}_\${cluster}_nucleotide_sequences.fasta - vsearch --cluster_fast \${name}_\${cluster}_nucleotide_sequences.fasta --id 0.2 --centroids \${name}_\${cluster}_centroids.fasta - grep ">" \${name}_\${cluster}_centroids.fasta >> \${name}_\${cluster}_tmp_centroids.list - for y in \$( cat \${name}_\${cluster}_tmp_centroids.list );do - echo ">\${cluster}_type"\$u"" >> \${name}_\${cluster}_tmp_centroid.newheaders - u=\$(( \${u}+1 )) - done - u=1 - ./rename_seq.py \${name}_\${cluster}_centroids.fasta \${name}_\${cluster}_tmp_centroid.newheaders \${cluster}_types_labeled.fasta - done - cat *_types_labeled.fasta >> ${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta - grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$3}' | awk -F "." '{print \$1}' >tmphead.list - grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$2}' | awk -F "," '{print \$1}' >tmplen.list - paste -d"," temporaryclusters.list tmphead.list >tmp.info.csv - grep ">" ${params.projtag}_pcASV\${id}.fasta >lala.list - j=1 - for x in \$(cat lala.list);do - echo ">${params.projtag}_pcASV\${j}" >>${params.projtag}_aminoheaders.list - echo "\${x},>${params.projtag}_pcASV\${j}" >>tmpaminotype.info.csv - j=\$(( \${j}+1 )) - done - rm lala.list - awk -F "," '{print \$2}' tmp.info.csv >>tmporder.list - for x in \$(cat tmporder.list);do - grep -w "\$x" tmpaminotype.info.csv | awk -F "," '{print \$2}' >>tmpder.list - done - paste -d "," temporaryclusters.list tmplen.list tmphead.list tmpder.list >${params.projtag}_pcASVCluster\${id}_summary.csv - ./rename_seq.py ${params.projtag}_pcASV\${id}.fasta ${params.projtag}_aminoheaders.list ${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta - stats.sh in=${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_aminoacid_clustered.gc gcformat=4 overwrite=true - stats.sh in=${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_nucleotide_clustered.gc gcformat=4 overwrite=true - awk 'BEGIN{RS=">";ORS=""}length(\$2)<"${params.minAA}"{print ">"\$0}' ${fasta} >${params.projtag}_pcASV\${id}_problematic_translations.fasta - if [ `wc -l ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk '{print \$1}'` -gt 1 ];then - grep ">" ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk -F ">" '{print \$2}' > problem_tmp.list - seqtk subseq ${asvs} problem_tmp.list > ${params.projtag}_pcASV\${id}_problematic_nucleotides.fasta - else - rm ${params.projtag}_pcASV\${id}_problematic_translations.fasta - fi - rm *.list - rm Cluster* - rm *types* - rm *tmp* - rm ${params.projtag}_pcASV\${id}.fast* - done - for x in *aminoacid*noTaxonomy.fasta;do - id=\$( echo \$x | awk -F "_noTax" '{print \$1}' | awk -F "pcASV" '{print \$2}') - numb=\$( grep -c ">" \$x) - echo "\${id},\${numb}" >> number_per_percentage_protz.csv - done - yesirr=\$( wc -l number_per_percentage_protz.csv | awk '{print \$1}') - tail -\$(( \${yesirr}-1 )) number_per_percentage_protz.csv > number_per_percentage_prot.csv - head -1 number_per_percentage_protz.csv >> number_per_percentage_prot.csv - rm number_per_percentage_protz.csv - """ - } - - process combine_csv_DC { - - input: - file(csv) from fastp_csv - .collect() - - output: - file("final_reads_stats.csv") into fastp_csv1 - - script: - """ - cat ${csv} >all_reads_stats.csv - head -n1 all_reads_stats.csv >tmp.names.csv - cat all_reads_stats.csv | grep -v ""Sample,Total_"" >tmp.reads.stats.csv - cat tmp.names.csv tmp.reads.stats.csv >final_reads_stats.csv - rm tmp.names.csv tmp.reads.stats.csv - """ - - } - - process Report_DataCheck { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/DataCheck/Report", mode: "copy", overwrite: true, pattern: '*.{html}' - - input: - file(fastpcsv) from fastp_csv1 - file(reads_per_sample_preFilt) from reads_per_sample_preFilt - file(read_per_sample_postFilt) from reads_per_sample_postFilt - file(preFilt_baseFrequency) from prefilt_basefreq - file(postFilt_baseFrequency) from postFilt_basefreq - file(preFilt_qualityScore) from prefilt_qualityscore - file(postFilt_qualityScore) from postFilt_qualityscore - file(preFilt_gcContent) from prefilt_gccontent - file(postFilt_gcContent) from postFilt_gccontent - file(preFilt_averageQuality) from prefilt_averagequality - file(postFilt_averageQuaulity) from postFilt_averagequality - file(preFilt_length) from prefilt_length - file(postFilt_length) from postFilt_length - file(number_per_percentage_nucl) from number_per_percent_nucl_plot - file(number_per_percentage_prot) from number_per_percent_prot_plot - - output: - file("*.html") into datacheckreport - - script: - """ - cp ${params.vampdir}/bin/vAMPirus_DC_Report.Rmd . - cp ${params.vampdir}/example_data/conf/vamplogo.png . - Rscript -e "rmarkdown::render('vAMPirus_DC_Report.Rmd',output_file='${params.projtag}_DataCheck_Report.html')" ${params.projtag} \ - ${fastpcsv} \ - ${reads_per_sample_preFilt} \ - ${read_per_sample_postFilt} \ - ${preFilt_baseFrequency} \ - ${postFilt_baseFrequency} \ - ${preFilt_qualityScore} \ - ${postFilt_qualityScore} \ - ${preFilt_averageQuality} \ - ${postFilt_averageQuaulity} \ - ${preFilt_length} \ - ${postFilt_length} \ - ${number_per_percentage_nucl} \ - ${number_per_percentage_prot} - """ - } - } else { println("\n\t\033[0;31mMandatory argument not specified. For more info use `nextflow run vAMPirus.nf --help`\n\033[0m") exit 0 From 3ceef117d2a8b78d78cdb618fbf10be32b36fae7 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Thu, 25 Feb 2021 09:53:54 -0800 Subject: [PATCH 003/223] Add conda parameter --- vampirus.config | 4 +++- 1 file changed, 3 insertions(+), 1 deletion(-) diff --git a/vampirus.config b/vampirus.config index dfa3e09..edbfee7 100644 --- a/vampirus.config +++ b/vampirus.config @@ -121,6 +121,8 @@ params { // Maximum number of iteration performed by metaMDS trymax="900" + // Conda env PATH (added automatically by startup script) + condaDir="CONDADIR" /* // ------------------------------------ STOP ------------------------------------ // @@ -253,7 +255,7 @@ profiles { params.condaActivate = true // cache for condaEnv created individually conda.cacheDir = "${params.localCacheDir}/condaEnv/" - process.conda = "CONDADIR" + process.conda = "${params.condaDir}" } docker { docker.enabled = true From 2b540897cf7b6d999740626d5247da689cf0b46c Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Thu, 25 Feb 2021 09:54:56 -0800 Subject: [PATCH 004/223] Change publishDir name --- vAMPirus.nf | 36 ++++++++++++++++++------------------ 1 file changed, 18 insertions(+), 18 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index a849587..309cf96 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -540,7 +540,7 @@ if (params.DataCheck || params.Analyze) { tag "${sample_id}" - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/FastQC/PreClean", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/FastQC/PreClean", mode: "copy", overwrite: true input: tuple sample_id, file(reads) from reads_qc_ch @@ -563,8 +563,8 @@ if (params.DataCheck || params.Analyze) { tag "${sample_id}" - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/AdapterRemoval", mode: "copy", overwrite: true, pattern: "*.filter.fq" - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/AdapterRemoval/fastpOut", mode: "copy", overwrite: true, pattern: "*.fastp.{json,html}" + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/AdapterRemoval", mode: "copy", overwrite: true, pattern: "*.filter.fq" + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/AdapterRemoval/fastpOut", mode: "copy", overwrite: true, pattern: "*.fastp.{json,html}" input: tuple sample_id, file(reads) from reads_ch @@ -599,7 +599,7 @@ if (params.DataCheck || params.Analyze) { tag "${sample_id}" - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/PrimerRemoval", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/PrimerRemoval", mode: "copy", overwrite: true input: tuple sample_id, file(reads) from reads_fastp_ch @@ -646,7 +646,7 @@ if (params.DataCheck || params.Analyze) { tag "${sample_id}" - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/FastQC/PostClean", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/FastQC/PostClean", mode: "copy", overwrite: true input: tuple sample_id, file(reads) from readsforqc2 @@ -668,8 +668,8 @@ if (params.DataCheck || params.Analyze) { tag "${sample_id}" - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging/Individual", mode: "copy", overwrite: true, pattern: "*mergedclean.fastq" - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging/Individual/notmerged", mode: "copy", overwrite: true, pattern: "*notmerged*.fastq" + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/Individual", mode: "copy", overwrite: true, pattern: "*mergedclean.fastq" + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/Individual/notmerged", mode: "copy", overwrite: true, pattern: "*notmerged*.fastq" input: tuple sample_id, file(reads) from reads_bbduk_ch @@ -691,7 +691,7 @@ if (params.DataCheck || params.Analyze) { label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging/LengthFiltering", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/LengthFiltering", mode: "copy", overwrite: true input: file(reads) from reads_vsearch1_ch @@ -710,7 +710,7 @@ if (params.DataCheck || params.Analyze) { label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging", mode: "copy", overwrite: true input: file(names) from names @@ -731,10 +731,10 @@ if (params.DataCheck || params.Analyze) { label 'norm_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging/LengthFiltering", mode: "copy", overwrite: true, pattern: "*_merged_preFilt*.fasta" - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging", mode: "copy", overwrite: true, pattern: "*Lengthfiltered.fastq" - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging/Histograms/pre_length_filtering", mode: "copy", overwrite: true, pattern: "*preFilt_*st.txt" - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging/Histograms/post_length_filtering", mode: "copy", overwrite: true, pattern: "*postFilt_*st.txt" + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/LengthFiltering", mode: "copy", overwrite: true, pattern: "*_merged_preFilt*.fasta" + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging", mode: "copy", overwrite: true, pattern: "*Lengthfiltered.fastq" + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/Histograms/pre_length_filtering", mode: "copy", overwrite: true, pattern: "*preFilt_*st.txt" + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/Histograms/post_length_filtering", mode: "copy", overwrite: true, pattern: "*postFilt_*st.txt" input: file(reads) from collect_samples_ch @@ -759,10 +759,10 @@ if (params.DataCheck || params.Analyze) { label 'norm_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging/LengthFiltering", mode: "copy", overwrite: true, pattern: "*_merged_preFilt*.fasta" - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging", mode: "copy", overwrite: true, pattern: "*Lengthfiltered.fastq" - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging/Histograms/pre_length_filtering", mode: "copy", overwrite: true, pattern: "*preFilt_*st.txt" - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging/Histograms/post_length_filtering", mode: "copy", overwrite: true, pattern: "*postFilt_*st.txt" + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/LengthFiltering", mode: "copy", overwrite: true, pattern: "*_merged_preFilt*.fasta" + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging", mode: "copy", overwrite: true, pattern: "*Lengthfiltered.fastq" + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/Histograms/pre_length_filtering", mode: "copy", overwrite: true, pattern: "*preFilt_*st.txt" + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/Histograms/post_length_filtering", mode: "copy", overwrite: true, pattern: "*postFilt_*st.txt" input: file(reads) from collect_samples_ch @@ -826,7 +826,7 @@ if (params.DataCheck || params.Analyze) { label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging/Uniques", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/Uniques", mode: "copy", overwrite: true input: file(reads) from reads_vsearch2_ch From 0c408231ee82fda8b2e82b1c0a2d76bcd72fbf6f Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Thu, 25 Feb 2021 12:18:02 -0800 Subject: [PATCH 005/223] Fix multiple env issue --- vampirus_startup.sh | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vampirus_startup.sh b/vampirus_startup.sh index 5d09829..5f5089a 100644 --- a/vampirus_startup.sh +++ b/vampirus_startup.sh @@ -170,7 +170,7 @@ else echo "Alright, lets check your system for Conda..." conda_c echo "Editing path to conda directory in vampirus.config" - environment="$(conda env list | sed 's/*//g' | grep "vAMPirus" | head -1 | awk '{print $2}')" + environment="$(conda info -e | awk '$1 == "vAMPirus" {print $2}')" sed "s|CONDADIR|${environment}|g" "$mypwd"/vampirus.config > tmp1.config cat tmp1.config > "$mypwd"/vampirus.config rm tmp1.config From 5635f39c16339f509da9e276e9e03163b3643a0b Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Sat, 27 Feb 2021 07:49:07 -0800 Subject: [PATCH 006/223] Add skip merging parameter --- vampirus.config | 2 ++ 1 file changed, 2 insertions(+) diff --git a/vampirus.config b/vampirus.config index edbfee7..c778930 100644 --- a/vampirus.config +++ b/vampirus.config @@ -163,6 +163,8 @@ params { skipEMBOSS = false // Skip Reports skipReport = false + // Skip Merging steps -> will also skip all read Processing + skipMerging = false // Data check parameters datacheckntIDlist=".55,.65,.75,.80,.81,.82,.83,.84,.85,.86,.87,.88,.89,.90,.91,.92,.93,.94,.95,.96,.97,.98,.99" datacheckaaIDlist=".55,.65,.75,.80,.81,.82,.83,.84,.85,.86,.87,.88,.89,.90,.91,.92,.93,.94,.95,.96,.97,.98,.99,1.0" From 22ed88298f50c2538e917e6576a2017955b442d4 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Sat, 27 Feb 2021 07:54:25 -0800 Subject: [PATCH 007/223] Add skipMerging. Change DataChek report --- vAMPirus.nf | 172 +++++++++++++++++++++++++++------------------------- 1 file changed, 90 insertions(+), 82 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 309cf96..b6096c6 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -463,12 +463,12 @@ if (params.readsTest) { Channel .fromFilePairs(params.readsTest) .ifEmpty{ exit 1, "params.readTest was empty - no input files supplied" } - .into{ reads_ch; reads_qc_ch } + .into{ reads_ch; reads_qc_ch; reads_processing } } else { println("\n\tEverything ready for launch.\n") Channel .fromFilePairs("${params.reads}", checkIfExists: true) - .into{ reads_ch; reads_qc_ch } + .into{ reads_ch; reads_qc_ch; reads_processing } } if (params.DataCheck || params.Analyze) { @@ -530,7 +530,7 @@ if (params.DataCheck || params.Analyze) { } } - if (!params.skipReadProcessing) { + if (!params.skipReadProcessing || !params.skipMerging ) { if (!params.skipFastQC) { @@ -589,6 +589,7 @@ if (params.DataCheck || params.Analyze) { reads_ch .set{ reads_fastp_ch } fastp_results = Channel.empty() + } if (!params.skipPrimerRemoval) { @@ -613,7 +614,7 @@ if (params.DataCheck || params.Analyze) { """ bbduk.sh in1=${reads[0]} out=${sample_id}_bb_R1.fastq.gz ftl=${params.defaultFwdTrim} t=${task.cpus} bbduk.sh in=${reads[1]} out=${sample_id}_bb_R2.fastq.gz ftl=${params.defaultRevTrim} t=${task.cpus} - repair.sh in1=${sample_id}_bb_R1.fastq.gz in2=${sample_id}_bb_R2.fastq.gz out1=${sample_id}_bbduk_R1.fastq.gz out2=${sample_id}_bbduk_R2.fastq.gz outs=sing.fq repair + repair.sh in1=${sample_id}_bb_R1.fastq.gz in2=${sample_id}_bb_R2.fastq.gz out1=${sample_id}_bbduk_R1.fastq.gz out2=${sample_id}_bbduk_R2.fastq.gz outs=sing.fq repair """ } else if ( params.GlobTrim && !params.GlobTrim == "" ) { """ @@ -621,7 +622,7 @@ if (params.DataCheck || params.Analyze) { RTRIM=\$( echo ${GlobTrim} | cut -f 2 -d "," ) bbduk.sh in=${reads[0]} out=${sample_id}_bb_R1.fastq.gz ftl=\${FTRIM} t=${task.cpus} bbduk.sh in=${reads[1]} out=${sample_id}_bb_R2.fastq.gz ftl=\${RTRIM} t=${task.cpus} - repair.sh in1=${sample_id}_bb_R1.fastq.gz in2=${sample_id}_bb_R2.fastq.gz out1=${sample_id}_bbduk_R1.fastq.gz out2=${sample_id}_bbduk_R2.fastq.gz outs=sing.fq repair + repair.sh in1=${sample_id}_bb_R1.fastq.gz in2=${sample_id}_bb_R2.fastq.gz out1=${sample_id}_bbduk_R1.fastq.gz out2=${sample_id}_bbduk_R2.fastq.gz outs=sing.fq repair """ } else if ( params.multi && params.primers ) { """ @@ -636,9 +637,10 @@ if (params.DataCheck || params.Analyze) { } else { reads_fastp_ch .set{ reads_bbduk_ch } + } - if (!params.skipFastQC) { + if (!params.skipFastQC && !params.skipPrimerRemoval) { process QualityCheck_2 { @@ -660,33 +662,45 @@ if (params.DataCheck || params.Analyze) { """ } } + } else { + reads_ch + .set{ reads_bbduk_ch } + } - process Read_Merging { + if (!params.skipMerging) { - label 'norm_cpus' + process Read_Merging { - tag "${sample_id}" + label 'norm_cpus' - publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/Individual", mode: "copy", overwrite: true, pattern: "*mergedclean.fastq" - publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/Individual/notmerged", mode: "copy", overwrite: true, pattern: "*notmerged*.fastq" + tag "${sample_id}" - input: - tuple sample_id, file(reads) from reads_bbduk_ch + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/Individual", mode: "copy", overwrite: true, pattern: "*mergedclean.fastq" + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/Individual/notmerged", mode: "copy", overwrite: true, pattern: "*notmerged*.fastq" - output: - file("*_mergedclean.fastq") into reads_vsearch1_ch - file("*.name") into names - file("*notmerged*.fastq") into notmerged + input: + tuple sample_id, file(reads) from reads_bbduk_ch - script: - """ - vsearch --fastq_mergepairs ${reads[0]} --reverse ${reads[1]} --threads ${task.cpus} --fastqout ${sample_id}_mergedclean.fastq --fastqout_notmerged_fwd ${sample_id}_notmerged_fwd.fastq --fastqout_notmerged_rev ${sample_id}_notmerged_rev.fastq --fastq_maxee ${params.maxEE} --relabel ${sample_id}. - echo ${sample_id} > ${sample_id}.name - """ + output: + file("*_mergedclean.fastq") into reads_vsearch1_ch + file("*.name") into names + file("*notmerged*.fastq") into notmerged + + script: + """ + vsearch --fastq_mergepairs ${reads[0]} --reverse ${reads[1]} --threads ${task.cpus} --fastqout ${sample_id}_mergedclean.fastq --fastqout_notmerged_fwd ${sample_id}_notmerged_fwd.fastq --fastqout_notmerged_rev ${sample_id}_notmerged_rev.fastq --fastq_maxee ${params.maxEE} --relabel ${sample_id}. + echo ${sample_id} > ${sample_id}.name + """ + } + + } else { + reads_bbduk_ch + .set{ reads_vsearch1_ch } } + process Compile_Reads { label 'low_cpus' @@ -904,14 +918,14 @@ if (params.DataCheck || params.Analyze) { echo "\${id},\${numb}" >> number_per_percentage_nucl.csv done yo=\$(grep -c ">" ${fasta}) - echo "1.0,\${yo}" >> number_per_percentage_nucl.csv + echo "1.0,\${yo}" >> number_per_percentage_nucl.csv """ } } if (params.sing) { - process Translating_For_ProteinClustering_DC { + process Translating_For_ProteinClustering_DC { label 'low_cpus' @@ -931,34 +945,33 @@ if (params.DataCheck || params.Analyze) { ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_ASVprotforclust.fasta --report ${params.projtag}_translation_report cp ${fasta} ${params.projtag}_ASV_all.fasta - """ - } + } } else { process Translation_For_ProteinBased_Clustering_DC { - label 'norm_cpus' + label 'norm_cpus' - conda 'python=2.7' + conda 'python=2.7' - publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Aminoacid/translation", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Aminoacid/translation", mode: "copy", overwrite: true - input: - file(fasta) from nucl2aa + input: + file(fasta) from nucl2aa - output: - file("*ASVprotforclust.fasta") into clustering_aa - file("*_translation_report") into reportaa_VR - file("*_ASV_all.fasta") into asvfastaforaaclust + output: + file("*ASVprotforclust.fasta") into clustering_aa + file("*_translation_report") into reportaa_VR + file("*_ASV_all.fasta") into asvfastaforaaclust - script: - """ - ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_ASVprotforclust.fasta --report ${params.projtag}_translation_report - cp ${fasta} ${params.projtag}_ASV_all.fasta - """ + script: + """ + ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_ASVprotforclust.fasta --report ${params.projtag}_translation_report + cp ${fasta} ${params.projtag}_ASV_all.fasta + """ } } @@ -1068,26 +1081,44 @@ if (params.DataCheck || params.Analyze) { """ } - process combine_csv_DC { + if (!params.skipReadProcessing || !params.skipMerging ) { - input: - file(csv) from fastp_csv_in1 - .collect() + process combine_csv_DC { - output: - file("final_reads_stats.csv") into fastp_csv_dc + input: + file(csv) from fastp_csv_in1 + .collect() - script: - """ - cat ${csv} >all_reads_stats.csv - head -n1 all_reads_stats.csv >tmp.names.csv - cat all_reads_stats.csv | grep -v ""Sample,Total_"" >tmp.reads.stats.csv - cat tmp.names.csv tmp.reads.stats.csv >final_reads_stats.csv - rm tmp.names.csv tmp.reads.stats.csv - """ + output: + file("final_reads_stats.csv") into fastp_csv_dc + script: + """ + cat ${csv} >all_reads_stats.csv + head -n1 all_reads_stats.csv >tmp.names.csv + cat all_reads_stats.csv | grep -v ""Sample,Total_"" >tmp.reads.stats.csv + cat tmp.names.csv tmp.reads.stats.csv >final_reads_stats.csv + rm tmp.names.csv tmp.reads.stats.csv + """ + + } + } else { + + process skip_combine_csv_DC { + output: + file("filter_reads.txt") into fastp_csv_dc + + script: + """ + echo "Read processing steps skipped." >filter_reads.txt + """ + } } + report_dc_in = Channel.create() + fastp_csv_dc.mix( reads_per_sample_preFilt, reads_per_sample_postFilt, prefilt_basefreq, postFilt_basefreq, prefilt_qualityscore, postFilt_qualityscore, prefilt_gccontent, postFilt_gccontent, prefilt_averagequality, postFilt_averagequality, prefilt_length, postFilt_length, number_per_percent_nucl_plot, number_per_percent_prot_plot + ).into(report_dc_in) + process Report_DataCheck { label 'norm_cpus' @@ -1095,21 +1126,8 @@ if (params.DataCheck || params.Analyze) { publishDir "${params.workingdir}/${params.outdir}/DataCheck/Report", mode: "copy", overwrite: true, pattern: '*.{html}' input: - file(fastpcsv) from fastp_csv_dc - file(reads_per_sample_preFilt) from reads_per_sample_preFilt - file(read_per_sample_postFilt) from reads_per_sample_postFilt - file(preFilt_baseFrequency) from prefilt_basefreq - file(postFilt_baseFrequency) from postFilt_basefreq - file(preFilt_qualityScore) from prefilt_qualityscore - file(postFilt_qualityScore) from postFilt_qualityscore - file(preFilt_gcContent) from prefilt_gccontent - file(postFilt_gcContent) from postFilt_gccontent - file(preFilt_averageQuality) from prefilt_averagequality - file(postFilt_averageQuaulity) from postFilt_averagequality - file(preFilt_length) from prefilt_length - file(postFilt_length) from postFilt_length - file(number_per_percentage_nucl) from number_per_percent_nucl_plot - file(number_per_percentage_prot) from number_per_percent_prot_plot + file(files) from report_dc_in + .collect() output: file("*.html") into datacheckreport @@ -1119,19 +1137,9 @@ if (params.DataCheck || params.Analyze) { cp ${params.vampdir}/bin/vAMPirus_DC_Report.Rmd . cp ${params.vampdir}/example_data/conf/vamplogo.png . Rscript -e "rmarkdown::render('vAMPirus_DC_Report.Rmd',output_file='${params.projtag}_DataCheck_Report.html')" ${params.projtag} \ - ${fastpcsv} \ - ${reads_per_sample_preFilt} \ - ${read_per_sample_postFilt} \ - ${preFilt_baseFrequency} \ - ${postFilt_baseFrequency} \ - ${preFilt_qualityScore} \ - ${postFilt_qualityScore} \ - ${preFilt_averageQuality} \ - ${postFilt_averageQuaulity} \ - ${preFilt_length} \ - ${postFilt_length} \ - ${number_per_percentage_nucl} \ - ${number_per_percentage_prot} + ${params.skipReadProcessing} \ + ${params.skipMerging} \ + ${params.skipAdapterRemoval} """ } From 58109ad21bf2380d0774a63c9384a03f6ae57122 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Sat, 27 Feb 2021 07:58:39 -0800 Subject: [PATCH 008/223] Add parameters check. --- bin/vAMPirus_DC_Report.Rmd | 134 ++++++++++++++++++++++--------------- 1 file changed, 81 insertions(+), 53 deletions(-) diff --git a/bin/vAMPirus_DC_Report.Rmd b/bin/vAMPirus_DC_Report.Rmd index 3320070..a0b4a1a 100644 --- a/bin/vAMPirus_DC_Report.Rmd +++ b/bin/vAMPirus_DC_Report.Rmd @@ -4,19 +4,10 @@ date: "Generated on: `r Sys.time()`" output: html_document params: interactive: TRUE - fastpcsv: !r commandArgs(trailingOnly=T)[2] - reads_per_sample_preFilt: !r commandArgs(trailingOnly=T)[3] - read_per_sample_postFilt: !r commandArgs(trailingOnly=T)[4] - preFilt_baseFrequency: !r commandArgs(trailingOnly=T)[5] - postFilt_baseFrequency: !r commandArgs(trailingOnly=T)[6] - preFilt_qualityScore: !r commandArgs(trailingOnly=T)[7] - postFilt_qualityScore: !r commandArgs(trailingOnly=T)[8] - preFilt_averageQuality: !r commandArgs(trailingOnly=T)[9] - postFilt_averageQuaulity: !r commandArgs(trailingOnly=T)[10] - preFilt_length: !r commandArgs(trailingOnly=T)[11] - postFilt_length: !r commandArgs(trailingOnly=T)[12] - number_per_percentage_nucl: !r commandArgs(trailingOnly=T)[13] - number_per_percentage_prot: !r commandArgs(trailingOnly=T)[14] + projtag: !r commandArgs(trailingOnly=T)[1] + skipReadProcessing: !r commandArgs(trailingOnly=T)[2] + skipMerging: !r commandArgs(trailingOnly=T)[3] + skipAdapterRemoval: !r commandArgs(trailingOnly=T)[4] --- + +```{r setup, include=FALSE} +knitr::opts_chunk$set(echo = TRUE, + message = FALSE, + warning = FALSE, + out.width="100%") +``` + +```{r pathways, echo=FALSE} +knitr::include_graphics("vamplogo.png") +``` + +```{r load_libraries, include=FALSE} +library(vegan) +library(tidyverse) +library(scales) +library(cowplot) +library(dplyr) +library(ggtree) +library(plotly) +library(knitr) +library(kableExtra) +library(rmarkdown) +library(processx) +``` + +```{r colors, include=FALSE} +mycol=c('#088da5','#73cdc8','#ff6f61','#7cb8df','#88b04b','#00a199','#6B5B95','#92A8D1','#b0e0e6','#ff7f50','#088d9b','#E15D44','#e19336') +``` +
+------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------ +NOTE: Most plots are interactive and you can use the legend to specify samples/treatment of interest. You can also download an .svg version of each figure within this report. +------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------ +
+

  Pre- and Post-Adapter Removal Read Stats

+
+```{r readstats, echo=FALSE} +if (params$skipReadProcessing == "true" || params$skipMerging == "true" ) { + writeLines("\n--------------------------------------------------------------\n") + cat(readLines(list.files(pattern="filter_reads.txt")) , sep = '\n') + writeLines("\n--------------------------------------------------------------\n") +} else { + if (params$skipAdapterRemoval == "false") { + reads_stats=read.csv("final_reads_stats.csv") + paged_table(reads_stats,options = list(rows.print = 20)) + } else { + writeLines("\n--------------------------------------------------------------\n") + cat(readLines(list.files(pattern="filter_reads.txt")) , sep = '\n') + writeLines("\n--------------------------------------------------------------\n") + } +} +``` +
+
+ + +### Total number of reads before and after adapter removal + +```{r readstats_plot, echo=FALSE} +# Plot of reads before and after +if (params$skipReadProcessing == "true" || params$skipMerging == "true" ) { + writeLines("\n--------------------------------------------------------------\n") + cat(readLines(list.files(pattern="filter_reads.txt")) , sep = '\n') + writeLines("\n--------------------------------------------------------------\n") +} else { + if (params$skipAdapterRemoval == "false") { + ptotal <- plot_ly(type="box",marker=list(colors=mycol)) + ptotal <- ptotal %>% add_boxplot(y=reads_stats$Total_before, name="Reads before filtering") + ptotal <- ptotal %>% add_boxplot(y=reads_stats$Total_after, name="Reads after filtering") + #ptotal <- ptotal %>% layout(title=list(text="Number of reads before and after filtering")) + ptotal <- ptotal %>% layout(legend = list(x=10,y=.5)) + ptotal <- ptotal %>% config(toImageButtonOptions=list(format='svg',filename='TotReads_b4_af_adaptrem', height= 500, width= 800, scale= 1)) + ptotal + } else { + # file fore here + writeLines("\n--------------------------------------------------------------\n") + cat(readLines(list.files(pattern="filter_reads.txt")) , sep = '\n') + writeLines("\n--------------------------------------------------------------\n") + } +} +``` +
+ +### Forward (R1) and reverse (R2) read length before and after adapter removal + +```{r readstats_plot2, echo=FALSE} +# Plot of R1 and R2 before and after +if (params$skipReadProcessing == "true" || params$skipMerging == "true" ) { + writeLines("\n--------------------------------------------------------------\n") + cat(readLines(list.files(pattern="filter_reads.txt")) , sep = '\n') + writeLines("\n--------------------------------------------------------------\n") +} else { + if (params$skipAdapterRemoval == "false") { + pr <- plot_ly(y=reads_stats$R1_before_length, type="box", name="R1 length before") + pr <- pr %>% add_boxplot(y=reads_stats$R1_after_length, name="R1 length after") + pr <- pr %>% add_boxplot(y=reads_stats$R2_before_length, name="R2 length before") + pr <- pr %>% add_boxplot(y=reads_stats$R2_after_length, name="R2 length after") + #pr <- pr %>% layout(title = "R1 and R2 Length") + pr <- pr %>% layout(legend = list(x=10,y=.5)) + pr <- pr %>% config(toImageButtonOptions=list(format='svg',filename='readlen_b4_af_adaptrem', height= 500, width= 800, scale= 1)) + pr + } else { + # file fore here + writeLines("\n--------------------------------------------------------------\n") + cat(readLines(list.files(pattern="filter_reads.txt")) , sep = '\n') + writeLines("\n--------------------------------------------------------------\n") + } +} +``` +
+
+
+```{r load_datasets_bash, include=FALSE} +cat *_counts.csv >counts.csv +cat *_summary_for_plot.csv >sum.csv +cat *_PercentID.matrix >matrix.txt +cat *.treefile >tree.txt +``` +```{r load_datasets, include=FALSE} +sample_name=read.csv("counts.csv") +sample_metadata=params$metadata +data<- read.csv(sample_name, check.names=FALSE) +data2 <-as.data.frame(t(data)) +data2$sample <- row.names(data2) +colnames(data2)<- as.matrix(data2[1,]) +as.data.frame(data2) +data2 <- data2[-1,] + +#X.OTU.ID for X.Sequence. +data2 <- data2 %>% + rename(sample=OTU_ID) +data2dim <- dim(data2) + +##Loading metadata +samples <- read.csv(sample_metadata) + +##Combining data and metadata +data3 <- merge(data2, samples, by="sample") + +dim_data3 <- dim(data3) +dim_samples <- dim(samples) +cols <- dim_data3[2]-dim_samples[2]+1 +first <-colnames(data3)[2] +last <- colnames(data3)[cols] +data3[,2:cols] <- lapply(data3[,2:cols], as.character) +data3[,2:cols] <- lapply(data3[,2:cols], as.numeric) + +#Calculate total reads per sample +data4 <- data3%>% + mutate(sum=select(.,2:cols)%>% + apply(1, sum, na.rm=TRUE)) + +``` + + +

  Number of Reads Per Sample

+ +```{r plot, echo=FALSE} +# sample and count +con <- plot_ly(data4, x = ~sum, y = ~sample, name = "Sample", type = 'scatter', + mode = "markers", marker = list(color = "#088da5"), hovertemplate = paste('Sample: %{y}','
Total reads: %{x}','')) +con <- con %>% layout(xaxis = list(title = "Total reads"),yaxis = list(title = "Sample")) +con <- con %>% config(toImageButtonOptions=list(format='svg',filename='Counts_per_sample', height= 500, width= 800, scale= 1)) +con +``` +
+
+
+```{r filter_data, include=FALSE} +##Filter samples with low reads +nfil=params$minimumCounts +data5 <- data4 %>% + filter(sum>nfil) + #can cause errors +data5dim <-dim(data5) +minreads<-min(data5$sum) +``` +
+
+
+
+
+ +

  Rarefaction

+```{r rarefaction, echo=FALSE, cache=FALSE} +##Rarefaction curves +rarefaction <- rarecurve(data5[,2:cols]) + +##rarefied dataset +raredata <- as.data.frame(rrarefy(data5[,2:cols], sample=minreads)) +``` +
+
+
+
+
+ + +

  Diversity Analyses Plots

+ +
+
+ +### Shannon diversty + +
+```{r diversity_analysis, echo=FALSE} +metadata <- data5[,(cols+1):data5dim[2]] +metadata$sample <- data5$sample +index <-diversity(raredata, index= "shannon") +shannondata5 <- as.data.frame(index) +shannondata5$sample<- data5$sample +shannondata5_2 <- merge(shannondata5, metadata, by="sample") + +#shannonplot <- ggplot(shannondata5_2, aes(x=treatment,color=treatment,y=index))+ +# geom_boxplot()+ +# geom_point()+ +# theme_classic()+ +# labs(y="Index",x="Treatment")+ +# theme(axis.text=element_text(size=12))+ +# theme(legend.position = "none") + +#sh<-ggplotly(shannonplot) +#sh <- sh %>% layout(title = list(text="Shannon diversty",y=.99)) +#sh + +sh <- plot_ly(shannondata5_2, x=~treatment, y=~index, color=~treatment, colors=mycol, type="box", boxpoints = "all", pointpos = 0, jitter = 0.5) +#sh <- sh %>% layout(title = list(text="Shannon diversty",y=.99)) +sh <- sh %>% layout(legend = list(x=10,y=.5), yaxis=list(title = "Index"), xaxis=list(title = "Treatment")) +sh <- sh %>% config(toImageButtonOptions=list(format='svg',filename='ShannonDiv', height= 500, width= 800, scale= 1)) +sh + +if (params$stats == "true" ) { + shannonaov <- aov(index ~ treatment, data= shannondata5_2) + st <- shapiro.test(resid(shannonaov)) + bt <- bartlett.test(index ~ treatment, data= shannondata5_2) + + if (st$p.value > .05 && bt$p.value > .05) { + print("Shapiro Test of normality - data is normal p-value > 0.05") + print(shapiro.test(resid(shannonaov))) + writeLines("\n--------------------------------------------------------------\n") + print("Bartlett Test variance homogeneity - variance is homogeneous p-value > 0.05") + print(bartlett.test(index ~ treatment, data= shannondata5_2)) + writeLines("\n--------------------------------------------------------------\n") + print("ANOVA Results") + print(summary(shannonaov)) + writeLines("\n--------------------------------------------------------------\n") + #Tukey Honest Significant Differences (pairwise comparison) - significant p <.05 + print("Tukey HSD - Pairwise comparison - significant differences indicated by p-value < 0.05") + print(TukeyHSD(shannonaov)) + writeLines("\n--------------------------------------------------------------\n") + } else { + print("Shapiro Test of normality - data is normal if p-value > 0.05") + print(shapiro.test(resid(shannonaov))) + writeLines("\n--------------------------------------------------------------\n") + print("Bartlett Test variance homogeneity - variance is homogeneous if p-value > 0.05") + print(bartlett.test(index ~ treatment, data= shannondata5_2)) + writeLines("\n--------------------------------------------------------------\n") + print("Data either not normal or variance not homogenous") + print("Kruskal-Wallis Test - test significant if p <.05") + #Kruskal-Wallis test - significant p <.05 + mykt <- kruskal.test(index ~ treatment, data= shannondata5_2) + print(mykt) + writeLines("\n--------------------------------------------------------------\n") + if (mykt$p.value < .05) { + #Pairwise comparison + print("Wilcox.test - pairwise comparison") + print(pairwise.wilcox.test(shannondata5_2$index, shannondata5_2$treatment, p.adjust.method = "BH")) + } else { + print("Data not significant. Skipping pairwise comparison") + } + writeLines("\n--------------------------------------------------------------\n") + print("ANOVA - one or more of the assumptions not met, take with a grain of salt.") + print(summary(shannonaov)) + writeLines("\n--------------------------------------------------------------\n") + } +} else { + print("Stats skipped. To toggle on use \"--stats run\" in vAMPirus launch command") +} +``` +
+
+
+
+ +### Simpson diversty + +
+```{r diversity_analysis2, echo=FALSE} +index <- diversity(raredata, index= "simpson") +simpsondata5 <- as.data.frame(index) +simpsondata5$sample<- data5$sample +simpsondata5_2 <- merge(simpsondata5, metadata, by="sample") + +#simpsonplot <- ggplot(simpsondata5_2, aes(x=treatment,color=treatment,y=index))+ +# geom_boxplot()+ +# geom_point()+ +# theme_classic()+ +# labs(y="Index",x="Treatment")+ +# theme(axis.text=element_text(size=12))+ +# theme(legend.position = "none") + +#s<-ggplotly(simpsonplot) +#s <- s %>% layout(title = list(text="Simpson diversty",y=.99)) +#s + +s <- plot_ly(simpsondata5_2, x=~treatment, y=~index, color=~treatment, colors=mycol, type="box", boxpoints = "all", pointpos = 0, jitter = 0.5) +#s <- s %>% layout(title = list(text="Simpson diversty",y=.99)) +s <- s %>% layout(legend = list(x=10,y=.5), yaxis=list(title = "Index"), xaxis=list(title = "Treatment")) +s <- s %>% config(toImageButtonOptions=list(format='svg',filename='SimpsonDiv', height= 500, width= 800, scale= 1)) +s + +if (params$stats == "true" ) { + simpsonaov <- aov(index ~ treatment, data= simpsondata5_2) + st <- shapiro.test(resid(simpsonaov)) + bt <- bartlett.test(index ~ treatment, data= simpsondata5_2) + + if (st$p.value > .05 && bt$p.value > .05) { + print("Shapiro Test of normality - data is normal p-value > 0.05") + print(shapiro.test(resid(simpsonaov))) + writeLines("\n--------------------------------------------------------------\n") + print("Bartlett Test variance homogeneity - variance is homogeneous p-value > 0.05") + print(bartlett.test(index ~ treatment, data= simpsondata5_2)) + writeLines("\n--------------------------------------------------------------\n") + print("ANOVA Results") + print(summary(simpsonaov)) + writeLines("\n--------------------------------------------------------------\n") + #Tukey Honest Significant Differences (pairwise comparison) - significant p <.05 + print("Tukey HSD - Pairwise comparison - significant differences indicated by p-value < 0.05") + print(TukeyHSD(simpsonaov)) + writeLines("\n--------------------------------------------------------------\n") + } else { + print("Shapiro Test of normality - data is normal if p-value > 0.05") + print(shapiro.test(resid(simpsonaov))) + writeLines("\n--------------------------------------------------------------\n") + print("Bartlett Test variance homogeneity - variance is homogeneous if p-value > 0.05") + print(bartlett.test(index ~ treatment, data= simpsondata5_2)) + writeLines("\n--------------------------------------------------------------\n") + print("Data either not normal or variance not homogenous") + print("Kruskal-Wallis Test - test significant if p <.05") + #Kruskal-Wallis test - significant p <.05 + mykt <- kruskal.test(index ~ treatment, data= simpsondata5_2) + print(mykt) + writeLines("\n--------------------------------------------------------------\n") + if (mykt$p.value < .05) { + #Pairwise comparison + print("Wilcox.test - pairwise comparison") + print(pairwise.wilcox.test(simpsondata5_2$index, simpsondata5_2$treatment, p.adjust.method = "BH")) + } else { + print("Data not significant. Skipping pairwise comparison") + } + writeLines("\n--------------------------------------------------------------\n") + print("ANOVA - one or more of the assumptions not met, take with a grain of salt.") + print(summary(simpsonaov)) + writeLines("\n--------------------------------------------------------------\n") + } +} else { + print("Stats skipped. To toggle on use \"--stats run\" in vAMPirus launch command") +} +``` +
+
+
+
+ +### Species Richness + +
+```{r diversity_analysis3, echo=FALSE} +mind5<-min(data5$sum) +index <- rarefy(data5[,2:cols], sample=mind5) +rarerichnessdata5 <- as.data.frame(index) +rarerichnessdata5$sample <-data5$sample +richdata5_2 <- merge(rarerichnessdata5, metadata, by="sample") + +#richnessplot <- ggplot(richdata5_2, aes(x=treatment,color=treatment,y=index))+ +# geom_boxplot()+ +# geom_point()+ +# theme_classic()+ +# labs(y="Richness",x="Treatment")+ +# theme(axis.text=element_text(size=12))+ +# theme(legend.position = "none") + +ri <- plot_ly(richdata5_2, x=~treatment, y=~index, color=~treatment, colors=mycol, type="box", boxpoints = "all", pointpos = 0, jitter = 0.5) +#ri <- ri %>% layout(title = list(text="ASV Richness",y=.99)) +ri <- ri %>% layout(legend = list(x=10,y=.5), yaxis=list(title = "Index"), xaxis=list(title = "Treatment")) +ri <- ri %>% config(toImageButtonOptions=list(format='svg',filename='SpeciesRich', height= 500, width= 800, scale= 1)) +ri + +if (params$stats == "true" ) { + richaov <- aov(index ~ treatment, data= richdata5_2) + st <- shapiro.test(resid(richaov)) + bt <- bartlett.test(index ~ treatment, data= richdata5_2) + + if (st$p.value > .05 && bt$p.value > .05) { + print("Shapiro Test of normality - data is normal p-value > 0.05") + print(shapiro.test(resid(richaov))) + writeLines("\n--------------------------------------------------------------\n") + print("Bartlett Test variance homogeneity - variance is homogeneous p-value > 0.05") + print(bartlett.test(index ~ treatment, data= richdata5_2)) + writeLines("\n--------------------------------------------------------------\n") + print("ANOVA Results") + print(summary(richaov)) + #Tukey Honest Significant Differences (pairwise comparison) - significant p <.05 + writeLines("\n--------------------------------------------------------------\n") + print("Tukey HSD - Pairwise comparison - significant differences indicated by p-value < 0.05") + print(TukeyHSD(richaov)) + writeLines("\n--------------------------------------------------------------\n") + } else { + print("Shapiro Test of normality - data is normal if p-value > 0.05") + print(shapiro.test(resid(richaov))) + writeLines("\n--------------------------------------------------------------\n") + print("Bartlett Test variance homogeneity - variance is homogeneous if p-value > 0.05") + print(bartlett.test(index ~ treatment, data= richdata5_2)) + writeLines("\n--------------------------------------------------------------\n") + print("Data either not normal or variance not homogenous") + print("Kruskal-Wallis Test - test significant if p <.05") + #Kruskal-Wallis test - significant p <.05 + mykt <- kruskal.test(index ~ treatment, data= richdata5_2) + print(mykt) + writeLines("\n--------------------------------------------------------------\n") + if (mykt$p.value < .05) { + #Pairwise comparison + print("Wilcox.test - pairwise comparison") + print(pairwise.wilcox.test(richdata5_2$index, richdata5_2$treatment, p.adjust.method = "BH")) + } else { + print("Data not significant. Skipping pairwise comparison") + } + writeLines("\n--------------------------------------------------------------\n") + print("ANOVA - one or more of the assumptions not met, take with a grain of salt.") + print(summary(richaov)) + writeLines("\n--------------------------------------------------------------\n") + } +} else { + print("Stats skipped. To toggle on use \"--stats run\" in vAMPirus launch command") +} +``` +
+
+
+
+
+ +

  Distance To Centroid

+
+```{r distance, echo=FALSE} +##Distance +intermediate <- raredata +bray.distance <- vegdist(sqrt(intermediate), method="bray") + +##Dispersion +disper <- betadisper(bray.distance, group = metadata$treatment, type="centroid") +df <- data.frame(Distance_to_centroid=disper$distances,Group=disper$group) +df$sample <- data5$sample +df2 <- merge(df, metadata, by="sample") + +#p<- ggplot(data=df2,aes(x=treatment,y=Distance_to_centroid,colour=treatment))+ +# geom_boxplot(outlier.alpha = 0)+ +# theme_classic()+ +# geom_point(position=position_dodge(width=0.75))+ +# labs(y="Distance",x="Treatment")+ +# theme(axis.text=element_text(size=12))+ +# theme(legend.position = "none") + +#cen <- ggplotly(p) +#cen <- cen %>% layout(title = list(text="Distance to centroid",y=.99)) +#cen + +cen <- plot_ly(df2, x=~treatment, y=~Distance_to_centroid, color=~treatment, colors=mycol, type="box", boxpoints = "all", pointpos = 0, jitter = 0.5) +#cen <- cen %>% layout(title = list(text="Distance to centroid",y=.99)) +cen <- cen %>% layout(legend = list(x=10,y=.5), yaxis=list(title = "Distance"), xaxis=list(title = "Treatment")) +cen <- cen %>% config(toImageButtonOptions=list(format='svg',filename='Dispersion', height= 500, width= 800, scale= 1)) +cen + +if (params$stats == "true" ) { + adn <- adonis(bray.distance~data5$treatment) + adn +} else { + print("Stats skipped. To toggle on use \"--stats run\" in vAMPirus launch command") +} +``` +
+
+
+
+
+ +

  NMDS Plots

+ +
+ +### 2D NMDS + +
+```{r nmds2d, echo=FALSE} +##NMDS +datax <- decostand(raredata,method="total") #method 'total' normalizes data to sum up to 1 --data5[,2:cols] + +MDS <- metaMDS(sqrt(datax), + distance = "bray",autotransform = FALSE, + k = 2, + maxit = 999, + trymax = params$try, + wascores = TRUE) + +if (MDS$converged == "TRUE") { + +data.scores <- as.data.frame(scores(MDS)) +data.scores$sample <- data5$sample +data.scores.2 <- merge(data.scores, metadata, by="sample") + +p <- ggplot(data.scores.2, aes(x=NMDS1, y=NMDS2,color=treatment))+ + geom_point(size=2)+ + theme_classic()+ + theme(legend.title = element_blank()) + +#fff <- ggplotly(p) +#fff <- fff %>% layout(legend=list(y=.5)) +#fff + +fff <-plot_ly(data.scores.2, x=~NMDS1, y=~NMDS2, color=~treatment, colors=mycol, text = ~paste("Sample: ", sample)) +fff <- fff %>% layout(legend=list(y=.5)) +fff <- fff %>% config(toImageButtonOptions=list(format='svg',filename='2Dnmds', height= 500, width= 800, scale= 1)) +fff + +} else { + print("No Convergence") +} +``` + +
+
+
+
+ +### 3D NMDS + +
+ +```{r nmds3d, echo=FALSE} +MDS3 <- metaMDS(sqrt(datax), + distance = "bray",autotransform = FALSE, + k = 3, + maxit = 999, + trymax = params$try, + wascores = TRUE) + +if (MDS3$converged == "TRUE") { + +data.scores3 <- as.data.frame(scores(MDS3)) +data.scores3$sample <- data5$sample +data.scores.3 <- merge(data.scores3, metadata, by="sample") +p3d <- plot_ly(data.scores.3, x=~NMDS1, y=~NMDS2, z=~NMDS3, text=~paste("Sample: ", sample), + color=~treatment, colors=mycol, + mode = 'markers', symbol = ~treatment, symbols = c('square','circle'), + marker = list(opacity = .8,line=list(color = 'darkblue',width = 1)) + ) +p3d <- p3d %>% layout(legend=list(y=.5)) +p3d <- p3d %>% config(toImageButtonOptions=list(format='svg',filename='3Dnmds', height= 500, width= 800, scale= 1)) +p3d + +} else { + print("No Convergence") +} +``` +
+
+
+
+
+ +

  OTU Abundance Per Sample

+ +```{r long, echo=FALSE} +dataz <- decostand(data5[,2:cols],method="total") #method 'total' normalizes data to sum up to 1 +dataz$sample <- data5$sample +datay <- merge(dataz, metadata, by="sample") +datalong <- datay %>% + tidyr::gather(first:last, key=hit, value=reads) + +##Barplot +## add better colors +#spec_bar <- ggplot(datalong, aes(x=forcats::fct_reorder(timepoint, as.numeric(as.character(timepoint))), +# y=reads, fill=hit))+ #Consider sqrt-transforming data +# geom_bar(aes(), stat="identity", position="fill")+ +# #coord_polar("y", start=0)+ +# theme_classic()+ +# facet_wrap(colony~treatment, nrow=2)+ +# #coord_flip()+ +# labs(x="Timepoint") #+ theme(legend.position = "none") +#spec_bar <- ggplot(datalong, aes(x=sample,y=reads,fill=hit))+geom_bar(aes(), stat="identity", position="fill")+ +# theme(axis.text.x=element_text(angle=90)) +#ggplotly(spec_bar) +#,'#00b300','#00b3b3','#0059b3','#6600ff','#b800e6','#ff3333','#ff8000','#ffff00','#bf8040','#42bcf5','#b428f5','#2e8d7e','#664e7e','#a4c700','#1aa3ff' + +ddd <- plot_ly(datalong, x=~sample, y=~reads, color=~hit, colors=mycol) +ddd <- ddd %>% layout(type='bar',barmode = 'stack') +ddd <- ddd %>% layout(legend = list(x=10,y=.5), xaxis=list(title = "Sample"), yaxis=list(title = "Relative abundance")) +ddd <- ddd %>% config(toImageButtonOptions=list(format='svg',filename='Relative_abundance', height= 500, width= 800, scale= 1)) +ddd +``` +
+
+
+
+
+ +

  OTU Abundance Per Treatment

+ +```{r asv_barplot, echo=FALSE} +datalong <- datalong %>% + filter(reads>0) + +#asv_bar <- ggplot(datalong, aes(x=reorder(hit,reads), y=reads, fill=treatment))+ +# geom_bar(stat="identity")+ +# scale_fill_manual(values=c('#088da5','#e19336'))+ #colors not working +# coord_flip()+ +# theme_classic()+ +# theme(axis.title.y = element_blank())+ +# theme(legend.title = element_blank()) + +asp2 <- plot_ly(datalong, y=~hit, x=~reads, color=~treatment, colors=mycol,text = ~paste("Sample: ", sample), opacity=.9) +asp2 <- asp2 %>% layout(type='bar', barmode = 'group') +asp2 <- asp2 %>% layout(yaxis = list(title = '', categoryorder = "total ascending"), legend = list(x=10,y=.5)) +asp2 <- asp2 %>% config(toImageButtonOptions=list(format='svg',filename='Most_abundant_hits_per_sample', height= 500, width= 800, scale= 1)) +asp2 +``` +
+
+
+
+
+ +

  Pairwise Percent-ID Heatmap

+ +
+
+```{r heatmap, echo=FALSE} +simmatrix<- read.csv("matrix.txt", header=FALSE) +rownames(simmatrix) <- simmatrix[,1] +simmatrix <- simmatrix[,-1] +colnames(simmatrix) <-rownames(simmatrix) +cols <- dim(simmatrix)[2] +simmatrix$AA <- rownames(simmatrix) +rval=nrow(simmatrix) +simmatrix2 <- simmatrix %>% + gather(1:rval, key=sequence, value=similarity) +x=reorder(simmatrix2$AA,simmatrix2$similarity) +y=reorder(simmatrix2$sequence,simmatrix2$similarity) +similaritymatrix <- ggplot(simmatrix2, aes(x=x, y=y,fill=similarity))+ + geom_raster()+ + scale_fill_distiller(palette="Spectral")+ + theme(axis.text.x = element_text(angle = 90))+ + theme(axis.title.x=element_blank())+ + theme(axis.title.y=element_blank()) + +heat <- ggplotly(similaritymatrix) +heat <- heat %>% config(toImageButtonOptions=list(format='svg',filename='heatmap', height= 500, width= 800, scale= 1)) +heat +``` +
+
+
+
+
+ + +

  Taxonomy Result Visualization

+ +
+
+ +```{r taxonomy, echo=FALSE} +if (params$skipTaxonomy == "true") + # file fore here + writeLines("\n--------------------------------------------------------------\n") + cat(readLines(list.files(pattern="filter_reads.txt")) , sep = '\n') + writeLines("\n--------------------------------------------------------------\n") +} else { + tax=read.csv("sum.csv",header=F) + tp <- plot_ly(tax, labels = ~V1, values = ~V2) + tp <- tp %>% add_pie(marker=list(colors=mycol, line=list(color='#000000', width=.5)), hole = 0.6) + tp <- tp %>% layout(title = "Taxonomy distribution", showlegend = F, + xaxis = list(showgrid = FALSE, zeroline = FALSE, showticklabels = FALSE), + yaxis = list(showgrid = FALSE, zeroline = FALSE, showticklabels = FALSE) + ) + tp <- tp %>% config(toImageButtonOptions=list(format='svg',filename='TaxDonut', height= 500, width= 800, scale= 1)) + tp +} + +``` + +
+
+
+
+
+ +

  Phylogenetic Tree

+ +
+
+ +```{r tree, echo=FALSE} +if (params$skipPhylogeny == "true") + # file fore here + writeLines("\n--------------------------------------------------------------\n") + cat(readLines(list.files(pattern="filter_reads.txt")) , sep = '\n') + writeLines("\n--------------------------------------------------------------\n") +} else { + #tree=read.newick("vAMPrun_otu.55.raxml.support") + tree=read.tree("tree.txt") + p1 <- ggtree(tree) + id <- tree$tip.label + dat <- tibble::tibble(id = id) + metat <- p1$data %>% dplyr::inner_join(dat, c('label' = 'id')) + p2 <- p1 + geom_point(data = metat, aes(x = x, y = y, label = id)) + ggplotly(p2, tooltip = "label") +} +``` +This tree is a maximum likelihood tree made with IQTREE2 and the parameters you specified in the vampirus.config file. Also, this is an interactive tree, you can zoom in and hover on nodes to know the sequence ID. For a better visualization of this tree, you can find the *.treefile with bootstrap support values within the results directory and visualize using programs like FigTree or ITOL. +
+
+
+
+
+
+
+
+
From 518e21f0fa9d05a79ca17a3ac7168d4abe4dba1f Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Tue, 2 Mar 2021 13:26:40 -0800 Subject: [PATCH 015/223] Change filenames. Create single report to handle all data. --- vAMPirus.nf | 396 ++++++++++------------------------------------------ 1 file changed, 76 insertions(+), 320 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 4d140a3..100f78a 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -2115,7 +2115,7 @@ if (params.DataCheck || params.Analyze) { script: """ - name=\$( echo ${prot} | awk -F ".fasta" '{print \$1}') + name=\$( echo ${prot} | awk -F "_noTax" '{print \$1}') clustalo -i ${prot} --distmat-out=\${name}_PairwiseDistanceq.matrix --full --force --threads=${task.cpus} clustalo -i ${prot} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} for x in *q.matrix;do @@ -2348,7 +2348,7 @@ if (params.DataCheck || params.Analyze) { script: """ # Protein_Alignment - pre=\$(echo ${prot} | awk -F ".fasta" '{print \$1}' ) + pre=\$(echo ${prot} | awk -F "_noTax" '{print \$1}' ) mafft --thread ${task.cpus} --maxiterate 15000 --auto ${prot} >\${pre}_ALN.fasta trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html @@ -2782,14 +2782,14 @@ if (params.DataCheck || params.Analyze) { echo " " > sequence.list grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqLikeObject.csv paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv for x in *phyloseqObject.csv;do echo "\$x" lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; sed 's/ /_/g' tmpcol.list > tmp2col.list; - cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; + cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_noTaxonomy_summary_for_plot.csv; rm tmpcol.list tmp2col.list done rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; @@ -2889,28 +2889,28 @@ if (params.DataCheck || params.Analyze) { trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html # pcASV_Nucleotide_ModelTest - modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint + modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_noTaxonomy_mt -d nt -s 203 --disable-checkpoint # pcASV_Nucleotide_Phylogeny if [ "${params.iqCustomnt}" != "" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -t \${pre}_mt.tree -T auto ${params.iqCustomnt} + iqtree -s \${pre}_aln.fasta --prefix \${pre}_noTaxonomy_iq --redo -T auto ${params.iqCustomnt} elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} + iqtree -s \${pre}_aln.fasta --prefix \${pre}_noTaxonomy_iq -m \${mod} --redo-nt auto -b ${params.boots} elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni + iqtree -s \${pre}_aln.fasta --prefix \${pre}_noTaxonomy_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni elif [ "${params.nonparametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} + iqtree -s \${pre}_aln.fasta --prefix \${pre}_noTaxonomy_iq -m MFP --redo -nt auto -b ${params.boots} elif [ "${params.parametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni + iqtree -s \${pre}_aln.fasta --prefix \${pre}_noTaxonomy_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni else - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni + iqtree -s \${pre}_aln.fasta --prefix \${pre}_noTaxonomy_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni fi done """ @@ -3245,7 +3245,7 @@ if (params.DataCheck || params.Analyze) { echo "\$z counted" done done - paste -d "," tmp.col1.txt *col.txt > ${params.projtag}_\${potu}_counts.csv + paste -d "," tmp.col1.txt *col.txt > ${params.projtag}_aminoacid_\${potu}_noTaxonomy_counts.csv rm tmp* rm *col.txt done @@ -3264,7 +3264,7 @@ if (params.DataCheck || params.Analyze) { .collect() output: - file("final_reads_stats.csv") into ( fastp_csv1, fastp_csv2, fastp_csv3, fastp_csv4, fastp_csv5 ) + file("final_reads_stats.csv") into fastp_csv_in script: """ @@ -3276,364 +3276,120 @@ if (params.DataCheck || params.Analyze) { """ } + } else { + + process skip_combine_csv { + + output: + file("filter_reads.txt") into fastp_csv_in + + script: + """ + echo "Read processing steps skipped." >filter_reads.txt + """ + + } + } //NEW REPORT !!!!!!!!!!!!!!!!! + /*Report_ASV + asv_counts_plots -> ${params.projtag}_ASV_counts.csv + taxplot1 -> ${params.projtag}_ASV_summary_for_plot.csv + asv_heatmap -> ${params.projtag}_ASV_PercentID.matrix + nucl_phyl_plot -> ${params.projtag}_ASV_iq.treefile + */ + report_asv = Channel.create() + asv_counts_plots.mix(taxplot1, asv_heatmap, nucl_phyl_plot).flatten().buffer(size:4).toList().into(report_asv) if (params.ncASV) { - - notu_counts_plots.mix(taxplot1a,notu_heatmap,nucl_phyl_plot,fastp_csv2) - + report_ncasv = Channel.empty() + notu_counts_plots.mix(taxplot1a, notu_heatmap, nucl_phyl_plot).flatten().toSortedList().flatten().buffer(size:4).toList().into(report_ncasv) /* notu_counts_plots -> ${params.projtag}_ncASV${id}_counts.csv taxplot1a -> ${params.projtag}_ncASV${id}_summary_for_plot.csv notu_heatmap -> ${params.projtag}_ncASV${id}_PercentID.matrix nucl_phyl_plot -> ${params.projtag}_ncASV${id}_iq.treefile - fastp_csv2 -> final_reads_stats.csv */ - + } else { + report_ncasv = Channel.empty() } if (params.pcASV) { - - potu_Acounts.mix(taxplot4,potu_aa_heatmap,potu_Atree_plot,fastp_csv3) - + report_pcasv_aa = Channel.create() + potu_Acounts.mix(taxplot4, potu_aa_heatmap, potu_Atree_plot).flatten().toSortedList().flatten().buffer(size:4).toList().into(report_pcasv_aa) /*Report_pcASV_AminoAcid - potu_Acounts -> ${params.projtag}_pcASV${id}_counts.csv + potu_Acounts -> ${params.projtag}_pcASV${id}_noTaxonomy_counts.csv taxplot4 -> ${params.projtag}_aminoacid_pcASV${id}_noTaxonomy_summary_for_plot.csv potu_aa_heatmap -> ${params.projtag}_aminoacid_pcASV${id}_noTaxonomy_PercentID.matrix potu_Atree_plot -> ${params.projtag}_aminoacid_pcASV${id}_noTaxonomy_iq.treefile - fastp_csv3 -> final_reads_stats.csv */ - - potu_Ncounts_for_report.mix(taxplot3,potu_nucl_heatmap,potu_Ntree_plot,fastp_csv4) - + report_pcasv_nucl = Channel.create() + potu_Ncounts_for_report.mix(taxplot3, potu_nucl_heatmap, potu_Ntree_plot).flatten().toSortedList().flatten().buffer(size:4).toList().into(report_pcasv_nucl) /*Report_pcASV_Nucleotide potu_Ncounts_for_report -> ${params.projtag}_nucleotide_pcASV${id}_noTaxonomy_counts.csv - taxplot3 -> ${params.projtag}_nucleotide_pcASV${id}_summary_for_plot.csv + taxplot3 -> ${params.projtag}_nucleotide_pcASV${id}_noTaxonomy_summary_for_plot.csv potu_nucl_heatmap -> ${params.projtag}_nucleotide_pcASV${id}_noTaxonomy_PercentID.matrix - potu_Ntree_plot -> ${params.projtag}_nucleotide_pcASV${id}_iq.treefile - fastp_csv4 -> final_reads_stats.csv + potu_Ntree_plot -> ${params.projtag}_nucleotide_pcASV${id}_noTaxonomy_iq.treefile */ - + } else { + report_pcasv_aa = Channel.empty() + report_pcasv_nucl = Channel.empty() } if (!params.skipAminoTyping) { - - aminocounts_plot.mix(taxplot2,aminotype_heatmap,amino_rax_plot,fastp_csv5) - + report_aminotypes = Channel.create() + aminocounts_plot.mix(taxplot2, aminotype_heatmap, amino_rax_plot).flatten().buffer(size:4).toList().into(report_aminotypes) /* Report_AminoTypes aminocounts_plot -> ${params.projtag}_AminoType_counts.csv taxplot2 -> ${params.projtag}_AminoTypes_summary_for_plot.csv - aminotype_heatmap -> ${params.projtag}_AminoTypes_noTaxonomy_PercentID.matrix - amino_rax_plot -> ${params.projtag}_AminoTypes_noTaxonomy_iq.treefile - fastp_csv5 -> final_reads_stats.csv + aminotype_heatmap -> ${params.projtag}_AminoTypes_PercentID.matrix + amino_rax_plot -> ${params.projtag}_AminoTypes_iq.treefile */ + } else { + report_aminotypes = Channel.empty() } - /*Report_ASV - asv_counts_plots -> ${params.projtag}_ASV_counts.csv - taxplot1 -> ${params.projtag}_ASV_summary_for_plot.csv - asv_heatmap -> ${params.projtag}_ASV_PercentID.matrix - nucl_phyl_plot -> ${params.projtag}_ASV_iq.treefile - fastp_csv1 -> final_reads_stats.csv - */ - asv_counts_plots.mix(taxplot1,asv_heatmap,nucl_phyl_plot,fastp_csv1) - - // what? - //process_combine_csv - //fastp_csv_in2 -> ${sample_id}_reads_stats.csv + report_asv.mix(report_ncasv, report_pcasv_aa, report_pcasv_nucl,report_aminotypes).buffer(size:1).into(report_all_ch) process Report { label 'norm_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReports", mode: "copy", overwrite: true input: + file(csv) from fastp_csv_in file(files) from report_all_ch .collect() output: file("*.html") into report_all_out - """ - name=\$( ls -1 | grep -w "_summary_for_plot.csv" | awk -F "_summary_for_plot.csv" '{print \$1}') - cp ${params.vampdir}/bin/vAMPirus_Report.Rmd . - cp ${params.vampdir}/example_data/conf/vamplogo.png . - Rscript -e "rmarkdown::render('vAMPirus_ReportA.Rmd',output_file='\${name}_Report.html')" \${name} \ - ${params.skipReadProcessing} \ - ${params.skipMerging} \ - ${params.skipAdapterRemoval} \ - ${params.skipTaxonomy} \ - ${params.skipPhylogeny} \ - ${params.trymax} \ - ${params.stats} \ - ${params.metadata} \ - ${params.minimumCounts} - """ + script: + """ + name=\$( ls -1 | grep -w "_summary_for_plot.csv" | awk -F "_summary_for_plot.csv" '{print \$1}') + cp ${params.vampdir}/bin/vAMPirus_Report.Rmd . + cp ${params.vampdir}/example_data/conf/vamplogo.png . + Rscript -e "rmarkdown::render('vAMPirus_Report.Rmd',output_file='\${name}_Report.html')" \${name} \ + ${params.skipReadProcessing} \ + ${params.skipMerging} \ + ${params.skipAdapterRemoval} \ + ${params.skipTaxonomy} \ + ${params.skipPhylogeny} \ + ${params.trymax} \ + ${params.stats} \ + ${params.metadata} \ + ${params.minimumCounts} + """ } - if (params.ncASV) { - - process Report_ASV { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport", mode: "copy", overwrite: true - - input: - file(counts) from asv_counts_plots - file(taxonomy) from taxplot1 - file(matrix) from asv_heatmap - file(readsstats) from fastp_csv1 - - output: - file("*.html") into report_summaryA - - script: - """ - name=\$( echo ${taxonomy} | awk -F "_summary_for_plot.csv" '{print \$1}') - cp ${params.vampdir}/bin/vAMPirus_ReportA.Rmd . - cp ${params.vampdir}/example_data/conf/vamplogo.png . - Rscript -e "rmarkdown::render('vAMPirus_ReportA.Rmd',output_file='\${name}_Report.html')" \${name} \ - ${readsstats} \ - ${counts} \ - ${params.metadata} \ - ${params.minimumCounts} \ - ${matrix} \ - ${taxonomy} \ - ${params.trymax} \ - ${params.stats} - - """ - } - - process Report_ncASV { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport/ncASV", mode: "copy", overwrite: true - - input: - file(counts) from notu_counts_plots - file(taxonomy) from taxplot1a - file(matrix) from notu_heatmap - file(phylogeny) from nucl_phyl_plot - file(readsstats) from fastp_csv2 - - output: - file("*.html") into report_summaryB - - script: - """ - cp ${params.vampdir}/bin/vAMPirus_ReportB.Rmd . - cp ${params.vampdir}/example_data/conf/vamplogo.png . - for x in *_summary_for_plot.csv;do - name=\$( echo \${x} | awk -F "_summary_for_plot.csv" '{print \$1}') - id=\$( echo \${x} | awk -F "_summary_for_plot.csv" '{print \$1}' | cut -f 2 -d "." ) - Rscript -e "rmarkdown::render('vAMPirus_ReportB.Rmd',output_file='\${name}_Report.html')" \${name} \ - ${readsstats} \ - \$( echo ${counts} | tr " " "\\n" | grep "\${id}" ) \ - ${params.metadata} \ - ${params.minimumCounts} \ - \$( echo ${matrix} | tr " " "\\n" | grep "\${id}" ) \ - \$( echo ${taxonomy} | tr " " "\\n" | grep "\${id}" ) \ - \$( echo ${phylogeny} | tr " " "\\n" | grep "\${id}" ) \ - ${params.trymax} \ - ${params.stats} - done - """ - } - - if (!params.skipAminoTyping) { - - process Report_AminoTypes { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport", mode: "copy", overwrite: true - - input: - file(counts) from aminocounts_plot - file(taxonomy) from taxplot2 - file(matrix) from aminotype_heatmap - file(phylogeny) from amino_rax_plot - file(readsstats) from fastp_csv5 - - output: - file("*.html") into report_summaryE - - script: - """ - name=\$( echo ${taxonomy} | awk -F "_summary_for_plot.csv" '{print \$1}') - cp ${params.vampdir}/bin/vAMPirus_ReportB.Rmd . - cp ${params.vampdir}/example_data/conf/vamplogo.png . - Rscript -e "rmarkdown::render('vAMPirus_ReportB.Rmd',output_file='\${name}_AminoType_Report.html')" \${name} \ - ${readsstats} \ - ${counts} \ - ${params.metadata} \ - ${params.minimumCounts} ${matrix} \ - ${taxonomy} \ - ${phylogeny} \ - ${params.trymax} \ - ${params.stats} - """ - } - } - } else { - - process Report_ASVs { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport", mode: "copy", overwrite: true - - input: - file(counts) from asv_counts_plots - file(taxonomy) from taxplot1 - file(matrix) from asv_heatmap - file(phylogeny) from nucl_phyl_plot - file(readsstats) from fastp_csv1 - - output: - file("*.html") into report_summaryA - - script: - """ - name=\$( echo ${taxonomy} | awk -F "_summary_for_plot.csv" '{print \$1}') - cp ${params.vampdir}/bin/vAMPirus_ReportB.Rmd . - cp ${params.vampdir}/example_data/conf/vamplogo.png . - Rscript -e "rmarkdown::render('vAMPirus_ReportB.Rmd',output_file='\${name}_ASV_Report.html')" \${name} \ - ${readsstats} \ - ${counts} \ - ${params.metadata} \ - ${params.minimumCounts} \ - ${matrix} \ - ${taxonomy} \ - ${phylogeny} \ - ${params.trymax} \ - ${params.stats} - """ - } - - if (!params.skipAminoTyping) { - - process Report_AminoType { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport", mode: "copy", overwrite: true - - input: - file(counts) from aminocounts_plot - file(taxonomy) from taxplot2 - file(matrix) from aminotype_heatmap - file(phylogeny) from amino_rax_plot - file(readsstats) from fastp_csv5 - - output: - file("*.html") into report_summaryE - - script: - """ - name=\$( echo ${taxonomy} | awk -F "_summary_for_plot.csv" '{print \$1}') - cp ${params.vampdir}/bin/vAMPirus_ReportB.Rmd . - cp ${params.vampdir}/example_data/conf/vamplogo.png . - Rscript -e "rmarkdown::render('vAMPirus_ReportB.Rmd',output_file='\${name}_AminoType_Report.html')" \${name} \ - ${readsstats} \ - ${counts} \ - ${params.metadata} \ - ${params.minimumCounts} ${matrix} \ - ${taxonomy} \ - ${phylogeny} \ - ${params.trymax} \ - ${params.stats} - """ - } - } - } - - if (params.pcASV) { - - process Report_pcASV_AminoAcid { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport/pcASV/Aminoacid", mode: "copy", overwrite: true - - input: - file(counts) from potu_Acounts - file(taxonomy) from taxplot4 - file(matrix) from potu_aa_heatmap - file(phylogeny) from potu_Atree_plot - file(readsstats) from fastp_csv3 - - output: - file("*.html") into report_summaryC - - script: - """ - cp ${params.vampdir}/bin/vAMPirus_ReportB.Rmd . - cp ${params.vampdir}/example_data/conf/vamplogo.png . - for x in *_summary_for_plot.csv;do - name=\$( echo \${x} | awk -F "_noTaxonomy_summary_for_plot.csv" '{print \$1}') - id=\$( echo \${x} | awk -F "_noTaxonomy_summary_for_plot.csv" '{print \$1}' | cut -f 2 -d "." ) - Rscript -e "rmarkdown::render('vAMPirus_ReportB.Rmd',output_file='\${name}_pcASVaa\${id}_Report.html')" \${name} \ - ${readsstats} \ - \$( echo ${counts} | tr " " "\\n" | grep "\${id}" ) \ - ${params.metadata} \ - ${params.minimumCounts} \ - \$( echo ${matrix} | tr " " "\\n" | grep "\${id}" ) \ - \$( echo ${taxonomy} | tr " " "\\n" | grep "\${id}" ) \ - \$( echo ${phylogeny} | tr " " "\\n" | grep "\${id}" ) \ - ${params.trymax} \ - ${params.stats} - done - """ - } - - process Report_pcASV_Nucleotide { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport/pcASV/Nucleotide", mode: "copy", overwrite: true - - input: - file(counts) from potu_Ncounts_for_report - file(taxonomy) from taxplot3 - file(matrix) from potu_nucl_heatmap - file(phylogeny) from potu_Ntree_plot - file(readsstats) from fastp_csv4 + } - output: - file("*.html") into report_summaryD + } - script: - """ - cp ${params.vampdir}/bin/vAMPirus_ReportB.Rmd . - cp ${params.vampdir}/example_data/conf/vamplogo.png . - for x in *_summary_for_plot.csv;do - name=\$( echo \${x} | awk -F "_summary_for_plot.csv" '{print \$1}') - id=\$( echo \${x} | awk -F "_summary_for_plot.csv" '{print \$1}' | cut -f 2 -d "." ) - Rscript -e "rmarkdown::render('vAMPirus_ReportB.Rmd',output_file='\${name}_pcASVnt\${id}_Report.html')" \${name} \ - ${readsstats} \ - \$( echo ${counts} | tr " " "\\n" | grep "\${id}" ) \ - ${params.metadata} \ - ${params.minimumCounts} \ - \$( echo ${matrix} | tr " " "\\n" | grep "\${id}" ) \ - \$( echo ${taxonomy} | tr " " "\\n" | grep "\${id}" ) \ - \$( echo ${phylogeny} | tr " " "\\n" | grep "\${id}" ) \ - ${params.trymax} \ - ${params.stats} - done - """ - } - } - } - } } else { println("\n\t\033[0;31mMandatory argument not specified. For more info use `nextflow run vAMPirus.nf --help`\n\033[0m") exit 0 From f167d4165bfc942a26da32bb21cdfbe96bd8f84e Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Tue, 2 Mar 2021 15:15:14 -0800 Subject: [PATCH 016/223] Fix missing ). Fix name --- bin/vAMPirus_Report.Rmd | 10 +++++----- 1 file changed, 5 insertions(+), 5 deletions(-) diff --git a/bin/vAMPirus_Report.Rmd b/bin/vAMPirus_Report.Rmd index 51603aa..300f310 100644 --- a/bin/vAMPirus_Report.Rmd +++ b/bin/vAMPirus_Report.Rmd @@ -134,14 +134,14 @@ if (params$skipReadProcessing == "true" || params$skipMerging == "true" ) {


-```{r load_datasets_bash, include=FALSE} +```{bash load_datasets_bash, include=FALSE} cat *_counts.csv >counts.csv cat *_summary_for_plot.csv >sum.csv cat *_PercentID.matrix >matrix.txt cat *.treefile >tree.txt ``` ```{r load_datasets, include=FALSE} -sample_name=read.csv("counts.csv") +sample_name="counts.csv" sample_metadata=params$metadata data<- read.csv(sample_name, check.names=FALSE) data2 <-as.data.frame(t(data)) @@ -156,7 +156,7 @@ data2 <- data2 %>% data2dim <- dim(data2) ##Loading metadata -samples <- read.csv(sample_metadata) +samples <- read.csv(sample_metadata, header = TRUE) ##Combining data and metadata data3 <- merge(data2, samples, by="sample") @@ -695,7 +695,7 @@ heat
```{r taxonomy, echo=FALSE} -if (params$skipTaxonomy == "true") +if (params$skipTaxonomy == "true") { # file fore here writeLines("\n--------------------------------------------------------------\n") cat(readLines(list.files(pattern="filter_reads.txt")) , sep = '\n') @@ -726,7 +726,7 @@ if (params$skipTaxonomy == "true")
```{r tree, echo=FALSE} -if (params$skipPhylogeny == "true") +if (params$skipPhylogeny == "true") { # file fore here writeLines("\n--------------------------------------------------------------\n") cat(readLines(list.files(pattern="filter_reads.txt")) , sep = '\n') From 5cafaf1c0474db6bb8c185145851fa2b710acb0f Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Tue, 2 Mar 2021 15:15:48 -0800 Subject: [PATCH 017/223] WIP - Divide processes ASV and ncASV. Use each in processes --- vAMPirus.nf | 177 +++++++++++++++++++++++++--------------------------- 1 file changed, 86 insertions(+), 91 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 100f78a..8810a6b 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -471,6 +471,21 @@ if (params.readsTest) { .into{ reads_ch; reads_qc_ch; reads_processing } } +// HERE fix if stament and list names. Also is not an integer +if (params.clusterNuclIDlist == "") { + msize=params.clusterNuclIDlist + def slist=msize.split(',').collect{it as int} + def nnuc=slist.size() +} else { + def nnuc=1 +} + +if (params.clusterAAIDlist == "") { + msize2=params.clusterAAIDlist + def slist2=msize2.split(',').collect{it as int} + def naa=slist2.size() +} + if (params.DataCheck || params.Analyze) { println("\n\tRunning vAMPirus Analyze pipeline - This might take a while, check out Nextflow tower (tower.nf) to remotely monitor the run.\n") @@ -1154,19 +1169,21 @@ if (params.DataCheck || params.Analyze) { publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/ncASV", mode: "copy", overwrite: true, pattern: '*ncASV*.fasta' input: + each x from 1..nnuc file(fasta) from reads_vsearch5_ch output: + //HERE - sepearte avs and ncasv out and proceeses. matrix, couts and phylogeny tuple file("*_ncASV*.fasta"), file("*ASV.fasta") into ( nuclFastas_forDiamond_ch, nuclFastas_forCounts_ch, nuclFastas_forMatrix_ch) - tuple file("*_ncASV*.fasta"), file("*ASV.fasta") into nuclFastas_forphylogeny + file("*_ncASV*.fasta") into nuclFastas_forphylogeny_ncasv + file("*ASV.fasta") into nuclFastas_forphylogeny_asv script: if (params.clusterNuclIDlist) { + nid=slist1.get(x-1) """ cp ${fasta} ./${params.projtag}_ASV.fasta - for id in `echo ${params.clusterNuclIDlist} | tr "," "\\n"`;do - vsearch --cluster_fast ${params.projtag}_ASV.fasta --centroids ${params.projtag}_ncASV\${id}.fasta --threads ${task.cpus} --relabel ncASV --id \${id} - done + vsearch --cluster_fast ${params.projtag}_ASV.fasta --centroids ${params.projtag}_ncASV\${id}.fasta --threads ${task.cpus} --relabel ncASV --id ${nid} """ } else if (params.clusterNuclID) { """ @@ -1721,7 +1738,7 @@ if (params.DataCheck || params.Analyze) { output: tuple file("*_counts.csv"), file("*_counts.biome") into counts_vsearch - file("*ncASV*counts.csv") into notu_counts_plots + tuple val("${id}"), file("*ncASV*counts.csv") into notu_counts_plots file("*_ASV*counts.csv") into asv_counts_plots script: @@ -1880,25 +1897,68 @@ if (params.DataCheck || params.Analyze) { if (!params.skipPhylogeny) { // need to edit paths + nuclFastas_forphylogeny_asv = Channel.create() + nuclFastas_forphylogeny_ncasv = Channel.create() + nuclFastas_forphylogeny.into(nuclFastas_forphylogeny_asv, nuclFastas_forphylogeny_ncasv) + + process ASV_Phylogeny { + + label 'norm_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*ASV*aln.*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*ASV*mt*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*ASV*iq*' + + input: + file(asvs) from nuclFastas_forphylogeny_asv + + output: + tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into align_results + file("*iq.treefile") into nucl_phyl_plot + + script: + """ + pre=\$(echo \${filename} | awk -F ".fasta" '{print \$1}' ) + mafft --thread ${task.cpus} --maxiterate 15000 --auto \${filename} >\${pre}_ALN.fasta + trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html + # Nucleotide_ModelTest + modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint + # Nucleotide_Phylogeny + if [ "${params.iqCustomnt}" != "" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomnt} + elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} + elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni + elif [ "${params.nonparametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} + elif [ "${params.parametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + else + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + fi + done + """ + } + if (params.ncASV) { - process Nucleotide_Phylogeny { + process ncASV_Phylogeny { label 'norm_cpus' publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*ncASV*aln.*' publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*ncASV*mt*' publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*ncASV*iq*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*_ASV*aln.*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*_ASV*mt*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*_ASV*iq*' input: - tuple file(notus), file(asvs) from nuclFastas_forphylogeny + tuple file(notus), file(asvs) from nuclFastas_forphylogeny_ncasv output: - tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into align_results - file("*iq.treefile") into nucl_phyl_plot + tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into align_results_ncasv + file("*iq.treefile") into nucl_phyl_plot_ncasv script: """ @@ -1925,77 +1985,12 @@ if (params.DataCheck || params.Analyze) { iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni fi done - for filename in ${asvs};do - pre=\$(echo \${filename} | awk -F ".fasta" '{print \$1}' ) - mafft --thread ${task.cpus} --maxiterate 15000 --auto \${filename} >\${pre}_ALN.fasta - trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html - # Nucleotide_ModelTest - modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint - # Nucleotide_Phylogeny - if [ "${params.iqCustomnt}" != "" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomnt} - elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} - elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni - elif [ "${params.nonparametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} - elif [ "${params.parametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - else - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - fi - done """ } - } else { - - process ASV_Phylogeny { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*ASV*aln.*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*ASV*mt*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*ASV*iq*' - - input: - file(asvs) from nuclFastas_forphylogeny - - output: - tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into align_results - file("*iq.treefile") into nucl_phyl_plot - - script: - """ - for filename in ${asvs};do - pre=\$(echo \${filename} | awk -F ".fasta" '{print \$1}' ) - mafft --thread ${task.cpus} --maxiterate 15000 --auto \${filename} >\${pre}_ALN.fasta - trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html - # Nucleotide_ModelTest - modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint - # Nucleotide_Phylogeny - if [ "${params.iqCustomnt}" != "" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomnt} - elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} - elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni - elif [ "${params.nonparametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} - elif [ "${params.parametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - else - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - fi - done - """ - } - } - } + + } + + } if (!params.skipAminoTyping) { @@ -2782,7 +2777,7 @@ if (params.DataCheck || params.Analyze) { echo " " > sequence.list grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqLikeObject.csv + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv for x in *phyloseqObject.csv;do echo "\$x" @@ -3301,11 +3296,11 @@ if (params.DataCheck || params.Analyze) { nucl_phyl_plot -> ${params.projtag}_ASV_iq.treefile */ report_asv = Channel.create() - asv_counts_plots.mix(taxplot1, asv_heatmap, nucl_phyl_plot).flatten().buffer(size:4).toList().into(report_asv) + asv_counts_plots.mix(taxplot1, asv_heatmap, nucl_phyl_plot).flatten().buffer(size:4).into(report_asv) if (params.ncASV) { - report_ncasv = Channel.empty() - notu_counts_plots.mix(taxplot1a, notu_heatmap, nucl_phyl_plot).flatten().toSortedList().flatten().buffer(size:4).toList().into(report_ncasv) + report_ncasv = Channel.create() + notu_counts_plots.mix(taxplot1a, notu_heatmap, nucl_phyl_plot_ncasv).flatten().toSortedList().flatten().buffer(size:4).into(report_ncasv) /* notu_counts_plots -> ${params.projtag}_ncASV${id}_counts.csv taxplot1a -> ${params.projtag}_ncASV${id}_summary_for_plot.csv @@ -3318,7 +3313,7 @@ if (params.DataCheck || params.Analyze) { if (params.pcASV) { report_pcasv_aa = Channel.create() - potu_Acounts.mix(taxplot4, potu_aa_heatmap, potu_Atree_plot).flatten().toSortedList().flatten().buffer(size:4).toList().into(report_pcasv_aa) + potu_Acounts.mix(taxplot4, potu_aa_heatmap, potu_Atree_plot).flatten().toSortedList().flatten().buffer(size:4).into(report_pcasv_aa) /*Report_pcASV_AminoAcid potu_Acounts -> ${params.projtag}_pcASV${id}_noTaxonomy_counts.csv taxplot4 -> ${params.projtag}_aminoacid_pcASV${id}_noTaxonomy_summary_for_plot.csv @@ -3326,7 +3321,7 @@ if (params.DataCheck || params.Analyze) { potu_Atree_plot -> ${params.projtag}_aminoacid_pcASV${id}_noTaxonomy_iq.treefile */ report_pcasv_nucl = Channel.create() - potu_Ncounts_for_report.mix(taxplot3, potu_nucl_heatmap, potu_Ntree_plot).flatten().toSortedList().flatten().buffer(size:4).toList().into(report_pcasv_nucl) + potu_Ncounts_for_report.mix(taxplot3, potu_nucl_heatmap, potu_Ntree_plot).flatten().toSortedList().flatten().buffer(size:4).into(report_pcasv_nucl) /*Report_pcASV_Nucleotide potu_Ncounts_for_report -> ${params.projtag}_nucleotide_pcASV${id}_noTaxonomy_counts.csv taxplot3 -> ${params.projtag}_nucleotide_pcASV${id}_noTaxonomy_summary_for_plot.csv @@ -3340,7 +3335,7 @@ if (params.DataCheck || params.Analyze) { if (!params.skipAminoTyping) { report_aminotypes = Channel.create() - aminocounts_plot.mix(taxplot2, aminotype_heatmap, amino_rax_plot).flatten().buffer(size:4).toList().into(report_aminotypes) + aminocounts_plot.mix(taxplot2, aminotype_heatmap, amino_rax_plot).flatten().buffer(size:4).into(report_aminotypes) /* Report_AminoTypes aminocounts_plot -> ${params.projtag}_AminoType_counts.csv @@ -3352,7 +3347,8 @@ if (params.DataCheck || params.Analyze) { report_aminotypes = Channel.empty() } - report_asv.mix(report_ncasv, report_pcasv_aa, report_pcasv_nucl,report_aminotypes).buffer(size:1).into(report_all_ch) + report_all_ch = Channel.create() + report_asv.mix(report_ncasv, report_pcasv_aa, report_pcasv_nucl, report_aminotypes).buffer(size:1).flatten().toList().view().into(report_all_ch) process Report { @@ -3363,14 +3359,13 @@ if (params.DataCheck || params.Analyze) { input: file(csv) from fastp_csv_in file(files) from report_all_ch - .collect() output: file("*.html") into report_all_out script: """ - name=\$( ls -1 | grep -w "_summary_for_plot.csv" | awk -F "_summary_for_plot.csv" '{print \$1}') + name=\$( ls *summary_for_plot.csv | awk -F "_summary_for_plot.csv" '{print \$1}') cp ${params.vampdir}/bin/vAMPirus_Report.Rmd . cp ${params.vampdir}/example_data/conf/vamplogo.png . Rscript -e "rmarkdown::render('vAMPirus_Report.Rmd',output_file='\${name}_Report.html')" \${name} \ From 76be22a1a3e2658d6267827ce40cc2e59cdf23c0 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Wed, 10 Mar 2021 12:42:10 -0800 Subject: [PATCH 018/223] Change python 2to3 --- bin/virtualribosomev2/dna2pep.py | 457 +++++++++++++------------ bin/virtualribosomev2/mod_translate.py | 108 +++--- 2 files changed, 283 insertions(+), 282 deletions(-) diff --git a/bin/virtualribosomev2/dna2pep.py b/bin/virtualribosomev2/dna2pep.py index 6f69015..9d4e9f9 100644 --- a/bin/virtualribosomev2/dna2pep.py +++ b/bin/virtualribosomev2/dna2pep.py @@ -1,4 +1,4 @@ -#!/usr/bin/env python2 +#!/usr/bin/env python3 # Copyright 2006 Rasmus Wernersson, Technical University of Denmark # @@ -29,60 +29,60 @@ SYNOPSIS dna2pep [options] [input files] [-f outfile] - + DESCRIPTION TRANSLATION: The translation engine of dna2pep has full support for handling degenerate nucleotides (IUPAC definition, e.g. W = A or T, S = G or C). - All translation table defined by the NCBI taxonomy group is included, + All translation table defined by the NCBI taxonomy group is included, and a number of options determining the behaviour of STOP and START codons is avialable. - - INTRON and EXONS: dna2pep natively understands TAB files containing + + INTRON and EXONS: dna2pep natively understands TAB files containing Intron/Exon annotation (gb2tab / FeatureExtract). When translating files containing Intron/Exon structure, dna2pep will annotate the underlying gene-structure in the annotation of the translated sequence. - + Input files can be in FASTA (no Intron/Exon annotation) RAW (single sequence with no header - all non-letters are discarded) or TAB (incluing annotation) FORMAT. The output format will by default be FASTA - for files without annotation and TAB for files including annotation. + for files without annotation and TAB for files including annotation. The file format is autodetected by investigating the first line of the input. If no input files are specified, dna2pep will read from STDIN. - + OPTIONS -F, --outfile - Optional - specify an output file. If no output file is + Optional - specify an output file. If no output file is specified the output will go to STDOUT. - + -O, --outformat - Specify output format (see also the --fasta, --tab, + Specify output format (see also the --fasta, --tab, --report options below): - + FASTA: Fasta format (plain DNA, no sequence annotation) - + TAB: Tab format. Each line contains the following four fields, separated by tabs: name, seq, ann, comment - + See gb2tab (FeatureExtract) for details. - - REPORT: A nice visualization of the results. - - AUTO: [Default] Generate a both a report and sequence output - (use the same format as the one detected from the for + + REPORT: A nice visualization of the results. + + AUTO: [Default] Generate a both a report and sequence output + (use the same format as the one detected from the for the input files). --fasta filename Write output sequences in FASTA format to the specified file. Use '-' to indicate STDOUT. - + --tab filename Write output sequences in TAB format to the specified file. Use '-' to indicate STDOUT. - + --report filename Write report to the specified file. Use '-' to indicate STDOUT. @@ -90,121 +90,121 @@ -m, --matrix tablename/file Use alternative translation matrix instead of the build-in Standard Genetic Code for translation. - - If "tablename" is 1-6,9-16 or 21-23 one of the alternative - translation tables defined by the NCBI taxonomy group will be + + If "tablename" is 1-6,9-16 or 21-23 one of the alternative + translation tables defined by the NCBI taxonomy group will be used. - + Briefly, the following tables are defined: ----------------------------------------- - 1: The Standard Code - 2: The Vertebrate Mitochondrial Code - 3: The Yeast Mitochondrial Code - 4: The Mold, Protozoan, and Coelenterate Mitochondrial Code - and the Mycoplasma/Spiroplasma Code - 5: The Invertebrate Mitochondrial Code - 6: The Ciliate, Dasycladacean and Hexamita Nuclear Code - 9: The Echinoderm and Flatworm Mitochondrial Code - 10: The Euplotid Nuclear Code - 11: The Bacterial and Plant Plastid Code - 12: The Alternative Yeast Nuclear Code - 13: The Ascidian Mitochondrial Code - 14: The Alternative Flatworm Mitochondrial Code - 15: Blepharisma Nuclear Code - 16: Chlorophycean Mitochondrial Code - 21: Trematode Mitochondrial Code - 22: Scenedesmus obliquus mitochondrial Code - 23: Thraustochytrium Mitochondrial Code - + 1: The Standard Code + 2: The Vertebrate Mitochondrial Code + 3: The Yeast Mitochondrial Code + 4: The Mold, Protozoan, and Coelenterate Mitochondrial Code + and the Mycoplasma/Spiroplasma Code + 5: The Invertebrate Mitochondrial Code + 6: The Ciliate, Dasycladacean and Hexamita Nuclear Code + 9: The Echinoderm and Flatworm Mitochondrial Code + 10: The Euplotid Nuclear Code + 11: The Bacterial and Plant Plastid Code + 12: The Alternative Yeast Nuclear Code + 13: The Ascidian Mitochondrial Code + 14: The Alternative Flatworm Mitochondrial Code + 15: Blepharisma Nuclear Code + 16: Chlorophycean Mitochondrial Code + 21: Trematode Mitochondrial Code + 22: Scenedesmus obliquus mitochondrial Code + 23: Thraustochytrium Mitochondrial Code + See http://www.ncbi.nlm.nih.gov/Taxonomy [Genetic Codes] for a detailed description. Please notice that the table of start codons is also used (see the --allinternal option below for details). - + If a filename is supplied the translation table is read from - file instead. - + file instead. + The file should contain one line per codon in the format: - + codonaa-single letter code - - All 64 codons must be included. Stop codons is specified + + All 64 codons must be included. Stop codons is specified by "*". T and U is interchangeable. Blank lines and lines starting with "#" are ignored. - + See the "gcMitVertebrate.mtx" file in the dna2pep source distribution for a well documented example. -r x, --readingframe=x Specify the reading frame. For input files in TAB format this options is ignored, and the reading frame is build from the - annotated Intron/Exon structure. - + annotated Intron/Exon structure. + 1: Reading frame 1 (e.g. ATGxxxxxx). DEFAULT. 2: Reading frame 2 (e.g. xATGxxxxx). 3: Reading frame 3 (e.g. xxATGxxxx). - + -1: Reading frame 1 on the minus strand. -2: Reading frame 2 on the minus strand. -3: Reading frame 3 on the minus strand. - - all: Try all reading frames. + + all: Try all reading frames. This option also implies the -x option. - + plus: All positive reading frames. This option also implies the -x option. - + minus: All negative reading frames. This option also implies the -x option. - + -o mode, --orf mode - Report longest ORF in the reading frame(s) specified with the + Report longest ORF in the reading frame(s) specified with the -r option. - - Mode governs which criterias are used to allow the opening of - an ORF. "Strict start codons" => codons _always_ coding for - methione (e.g. ATG in the standard code), "Minor start codons" - => codon only coding for methionine at the start positon - (e.g. TTG in the standard genetic code). - + + Mode governs which criterias are used to allow the opening of + an ORF. "Strict start codons" => codons _always_ coding for + methione (e.g. ATG in the standard code), "Minor start codons" + => codon only coding for methionine at the start positon + (e.g. TTG in the standard genetic code). + Mode can be: ------------ strict: Open an ORF at "strict start codons" only. any: Open an ORF at any start codon. - none: Do not use start codons - look for the longest + none: Do not use start codons - look for the longest fragment before a STOP codon. - - The DNA fragment usedfor encoding the ORF will be added to the + + The DNA fragment usedfor encoding the ORF will be added to the comment field (TAB format only). - + -a, --allinternal By default the very first codon in each sequences is assumed to be the initial codon on the transcript. This means certain - non-methionine codons actually codes for metionine at this + non-methionine codons actually codes for metionine at this position. For example "TTG" in the standard genetic code (see above). - - Selecting this option treats all codons as internal codons. - + + Selecting this option treats all codons as internal codons. + -x, --readthroughstop Allow the translation to continue after a stop codon is reached. The stop codon will be marked as "*". - + -p, --plain, --ignoreannotation Ignore annotation for TAB files. If this options is selected TAB files will be treated in same way as FASTA files. - -c, --comment + -c, --comment Preserve the comment field in TAB files. Normally the comment field is silently dropped, since it makes no sense for FASTA files. - + -C, --processcomment Works as the -c option described above, except a bit of intelligent parsing is done on the comment field: If a "/spliced_product" sub-field is found (from TAB files create by FeatureExtract / gb2tab) only the part of the comment field before the DNA specific information - is kept in the comment field. + is kept in the comment field. -e, --exonstructure Default for TAB files. Annotate the underlying exons structure @@ -212,19 +212,19 @@ are fully or partially encoded within the first exon get the annotation character "1", positions in the secon exon get the character "2" etc. - + The hex-decimal system is used, which means up to 15 exons can be uniquely annotated, before the numbering wraps around to "0". - + -i, --intronphase Annotate where an intron interrupted the DNA sequences, and how the intron did cut the readingframe. - + 0 : phase-0 intron (inbetween the previous and current position). 1 : phase-1 intron. 2 : phase-2 intron. - - + + AUTHOR Rasmus Wernersson, raz@cbs.dtu.dk Feb-Mar 2006 @@ -234,11 +234,11 @@ WEB PAGE http://www.cbs.dtu.dk/services/VirtualRibosome/ - + REFERENCE Rasmus Wernersson Virtual Ribosome - Comprehensive DNA translation tool. - Submitted to Nucleic Acids Research, 2006 + Submitted to Nucleic Acids Research, 2006 """ import sys, re, mod_translate,string @@ -271,41 +271,41 @@ def makePretty(title,vals,labels,max_len): val = vals[j] lab = labels[j] pos = min(max_len,i+pwidth) - if lab: + if lab: spos = "%d" % pos else: spos = "" - + s = "%-2s %s %s\n" % (lab,val[i:pos],spos) l.append(s) l.append("\n") - return l + return l def explodePep(s): l = list(s) return " "+" ".join(l)+" " - + def revCom(dna): l = list(dna) l.reverse() rev_dna = "".join(l) return rev_dna.translate(transTable) - + def dnaComplement(dna): return dna.translate(transTable) - + def revStr(s): l = list(s) l.reverse() return "".join(l) - + def isDNAValid(dna): for c in dna: if not c in allValid: return False - + return True - + def combineToTab(name, seql): seq = "".join(seql) ann= "."*len(seq) @@ -320,19 +320,19 @@ def readRaw(lines): sl = [] for c in line.strip().upper(): if c.isalpha(): sl.append(c) - + seql.append("".join(sl)) - + l.append(combineToTab("Seq1",seql)) - return l - + return l + # Read FASTA format def readFasta(lines): l = [] - + name = "" seql = [] - + for line in lines: line = line.strip() if line.startswith(">"): @@ -342,104 +342,105 @@ def readFasta(lines): seql = [] else: seql.append(line) - + if name: l.append(combineToTab(name,seql)) - + return l -# read TAB format +# read TAB format def readTab(lines): l = [] - + for line in lines: line = line.strip() tokens = line.split("\t") if len(tokens) < 2: continue - + name = tokens[0] seq = tokens[1] - + if len(tokens) > 2: ann = tokens[2] else: ann = "."*len(seq) - + if len(tokens) > 3: com = tokens[3] else: com = "" - + l.append( (name,seq,ann,com) ) - + return l - + def readInput(lines): if not lines: return ([], True) - - line = lines[0] + + line = lines[0] tokens = line.split("\t") - + if line.startswith(">") and len(tokens) < 3: l = readFasta(lines) isFasta = True - + elif len(tokens) == 1: l = readRaw(lines) isFasta = True - + else: l = readTab(lines) isFasta = False - + return (l, isFasta) - + def writeFasta(seqs, outstream): for (name, seq, ann, com) in seqs: - print >> outstream, ">"+name + print(">"+name, file=outstream) for i in range(0,len(seq),60): - print >> outstream, seq[i:i+60] - + print(seq[i:i+60], file=outstream) + def writeTab(seqs, outstream): for tokens in seqs: - print >> outstream, "\t".join(tokens) + print("\t".join(tokens), file=outstream) def openForWriteOrDie(outfile): try: outstream = file(outfile,"w") - except IOError, (strerror): - print >> sys.stderr, "ERROR - cannot write to the specified file %s [%s]" % (outfile,strerror) + except IOError as xxx_todo_changeme: + (strerror) = xxx_todo_changeme + print("ERROR - cannot write to the specified file %s [%s]" % (outfile,strerror), file=sys.stderr) sys.exit(-1) - + return outstream def parseOpts(): # Quick hack to overrule the -h and --help feature # build into the optpase module if "-h" in sys.argv or "--help" in sys.argv: - print __doc__ + print(__doc__) sys.exit(0) parser = OptionParser() - + # File handling parser.add_option("-F","--outfile", type="string", dest="outfile", default="") parser.add_option("-O","--outformat", type="string", dest="outformat", default="AUTO") parser.add_option("--tab", type="string", dest="tabfile", default="") parser.add_option("--fasta", type="string", dest="fastafile", default="") parser.add_option("--report", type="string", dest="reportfile", default="") - + # Matrix parser.add_option("-m","--matrix", type="string", dest="matrix", default="1") - + # Reading frame parser.add_option("-r","--readingframe", type="string", dest="readingframe", default="1") - - # ORF finding + + # ORF finding parser.add_option("-o","--orf", type="string", dest="orf", default="") - + # Stop and Start codons parser.add_option("-a","--allinternal", action="store_true", dest="allinternal", default = False) parser.add_option("-x","--readthroughstop", action="store_true", dest="readthroughstop", default = False) @@ -455,38 +456,38 @@ def parseOpts(): # Debug parser.add_option("-d","--debug", action="store_true", dest="debug", default=False) - + (opt, args) = parser.parse_args() - + # Check reading frame if not opt.readingframe in ["1", "2", "3", "-1", "-2", "-3", "all", "plus","minus"]: sys.stderr.write("Invalid reading frame [%s]\n" % opt.readingframe) sys.exit(-1) - + if opt.readingframe in ["all","plus","minus"]: opt.readthroughstop = True - + # Chech ORF mode if not opt.orf in ["","strict","any","none"]: - print >> sys.stderr, "Invalid ORF mode [%s]\n" % opt.orf + print("Invalid ORF mode [%s]\n" % opt.orf, file=sys.stderr) sys.exit(-1) - + # Chech output format opt.outformat = opt.outformat.upper() if not opt.outformat in ["AUTO","TAB","FASTA","REPORT"]: - print >> sys.stderr("Invalid output format [%s]\n" % opt.outformat) + print(file=sys.stderr("Invalid output format [%s]\n" % opt.outformat)) sys.exit(-1) - + # Check mutually exclusive options if opt.intronrf: opt.exonann=False - + return (opt, args) if __name__ == "__main__": reports = [] pepseqs = [] opt, args = parseOpts() - + # Initialize translation matrix mtx = mod_translate.parseMatrixFile(opt.matrix) if not mtx: @@ -508,12 +509,12 @@ def parseOpts(): except: sys.stderr.write("ERROR: Cannot read from file '%s'\n" % fn) sys.exit(-1) - + try: (seq_list, isFasta) = readInput(lines) - except Exception, msg: - print >> sys.stderr, "ERROR parsing input files. Please verify the format (FASTA, RAW or TAB)" - print >> sys.stderr, "[%s]" % (str(msg)) + except Exception as msg: + print("ERROR parsing input files. Please verify the format (FASTA, RAW or TAB)", file=sys.stderr) + print("[%s]" % (str(msg)), file=sys.stderr) sys.exit(-1) # Ignore annotation? @@ -525,36 +526,36 @@ def parseOpts(): for (name, dna, ann, com) in seq_list: seq_proc = ann_proc = exnum_proc = "" - + # Test if the DNA sequence is valid if not isDNAValid(dna): sys.stderr.write("Non IUPAC characters is detected in sequence '%s' - skipping this entry\n" %name) #sys.stderr.write("seq: %s\n" % seq) continue - + # Files without intron/exon annotation ------------------------------------------- if isFasta: # reports.append([ORF_ANNOTATION]) - + d_collect = {} if opt.readingframe in ["all"]: rf_list = ["1","2","3","-1","-2","-3"] echo_rf = True - + elif opt.readingframe == "plus": rf_list = ["1","2","3"] echo_rf = True - + elif opt.readingframe == "minus": rf_list = ["-1","-2","-3"] echo_rf = True - + else: rf_list = [opt.readingframe] echo_rf = False - + for rf in rf_list: - + # Find current reading frame if rf == "1": qseq = dna @@ -574,19 +575,19 @@ def parseOpts(): # Do the actual translation pep = mod_translate.translate(qseq,mtx,not opt.allinternal,opt.readthroughstop) pa = mod_translate.annotate(qseq,mtx) - + # The annotation string may be longer that the peptide, if the -x more is not used - pa = pa[:len(pep)] + pa = pa[:len(pep)] # Store translated sequence if echo_rf: cname = name+"_rframe"+rf else: cname = name - + data = ( cname,pep,pa,qseq ) d_collect[rf] = data - + # Do ORF finding? if opt.orf: #Find longest ORF @@ -594,7 +595,7 @@ def parseOpts(): bestspan = (0,0) bestdata = None bestrf = "" - for key in d_collect.keys(): + for key in list(d_collect.keys()): data = d_collect[key] seq = data[1] ann = data[2] @@ -617,52 +618,52 @@ def parseOpts(): j = j_strict else: j = min(j_strict,j_any) - + if j == -1: break - + #Step 2 - find stop m = ann.find("*",j) if m == -1: m = len(ann) - - #print j,m - + + #print j,m + if (m - j) > bestlen: bestlen = m - j bestspan = (j, m) bestdata = data bestrf = key - + #print rf, bestlen #print seq[j:m+1] #print ann[j:m+1] - + j = m # Format the best hit if not bestdata: - bestdata = d_collect.values()[0] + bestdata = list(d_collect.values())[0] bestspan = (0,0) - bestrf = d_collect.keys()[0] - + bestrf = list(d_collect.keys())[0] + msg = "NO ORF FOUND (given the criteria '%s') for sequence '%s'\n\n" % (opt.orf,name) - reports.append([msg]) - + reports.append([msg]) + # clist = list(bestdata[1].lower()) name = bestdata[0] pep = bestdata[1] ann = bestdata[2] dna_work = bestdata[3] - + bpos, epos = bestspan orf_dna = dna_work[bpos*3:epos*3] orf = mod_translate.translate(orf_dna,mtx,True,False) new_pep = " "*bpos + orf + " "*(len(pep)-epos) - + name += "_ORF" - + d_collect = {} d_collect[bestrf] = (name,new_pep,ann,orf_dna) #print d_collect - + # Processing and Pretty printing... if opt.readingframe in ["all","plus","minus"] and (not opt.orf): if opt.readingframe == "all": @@ -677,17 +678,17 @@ def parseOpts(): dna_plus = dna dna_minus = revCom(dna) - + #dnapl = list(dna_plus) #dnaml = list(dna_minus) # dnapann = [" "]*len(dna_plus) # dnamann = [" "]*len(dna_minus) dnapann = ["."]*len(dna_plus) dnamann = ["."]*len(dna_minus) - + vals = [] labels = [] - + if doPlus: pep1 = d_collect["1"][1] ann1 = d_collect["1"][2] @@ -720,8 +721,8 @@ def parseOpts(): #dna_plus = "".join(dnapl) vals += [pep3e,pep2e,pep1e,dna_plus,"".join(dnapann)] labels += ["","","","5'",""] - - if doMinus: + + if doMinus: pepm1 = d_collect["-1"][1] annm1 = d_collect["-1"][2] pepm1e = explodePep(pepm1)+" "+" " @@ -741,7 +742,7 @@ def parseOpts(): peps = [pepm1,pepm2,pepm3] maxlen = max(len(pepm1),len(pepm2)) maxlen = max(maxlen,len(pepm3)) - + for i in range(0,maxlen): for j in range(0,3): ann = anns[j] @@ -766,37 +767,37 @@ def parseOpts(): # vals = [pep3,pep2,pep1,dna_plus,revStr(dna_minus),revStr(pepm1),revStr(pepm2),revStr(pepm3)] # labels = ["","","","5'","3'","","",""] title = "%s - reading frame(s): %s" % (name,opt.readingframe) - + l = makePretty(title,vals,labels,len(dna_plus)) - + ###print "".join(l) reports.append(l) - + else: - rf_list = d_collect.keys() + rf_list = list(d_collect.keys()) rf = rf_list[0] - + vals = [] labels = [] title = "%s\nReading frame: %s" % (name,rf) - + #rf_int = abs(int(rf)) - 1 pep = d_collect[rf][1] pepx = explodePep(pep) - + vals = [] labels = [] - + vals.append(pepx) labels.append("") - + pa = d_collect[rf][2] pax = ["."]*len(pa)*3 if rf.startswith("-"): dna_minus = revCom(dna) #print len(dna_minus) dnamann = ["."] * len(dna_minus) - if rf == "-1": + if rf == "-1": pepm = d_collect["-1"][1] annm = d_collect["-1"][2] pepme = explodePep(pepm)+" "+" " @@ -811,9 +812,9 @@ def parseOpts(): annm = d_collect["-3"][2] pepme = " "+explodePep(pepm)+" "+" " j = 2 - + pepme = pepme[:len(dna_minus)] - + for i in range(0,len(pepm)): ac = annm[i] if ac in ["M","m","*"]: @@ -852,42 +853,42 @@ def parseOpts(): if ac in ["M","m","*"]: dnapos = (i*3) + j if ac == "M": c = ">" - elif ac == "m": c = ")" + elif ac == "m": c = ")" else: c = "*" for k in range(dnapos,dnapos+3): dnapann[k] = c - + ann = "".join(dnapann) vals = [pepx,dna_plus,ann] - labels = ["","5'",""] - - + labels = ["","5'",""] + + l = makePretty(title,vals,labels,len(dna)) reports.append(l) ###print "".join(l) - - - for rf in rf_list: + + + for rf in rf_list: name,seq,ann,com = d_collect[rf] new_com = "" - + # Seq may contain leding and trailing spaces if we are in ORF finde mode if opt.orf: new_seq = [] new_ann = [] for i in range(0,len(seq)): - if seq[i] <> " ": + if seq[i] != " ": new_seq.append(seq[i]) new_ann.append(ann[i]) - + seq = "".join(new_seq) ann = "".join(new_ann) - + new_com = '/orf_mode="%s"; /dna="%s";' % (opt.orf,com) - + pepseqs.append( (name,seq,ann,new_com) ) # pepseqs.append( (cname,pep,pa,com.strip()) ) - + # Files with intron/exon annotation --------------------------------------------- else: exon_count = 1 @@ -895,17 +896,17 @@ def parseOpts(): start, end = mo.span() seq_proc += dna[start:end] ann_proc += ann[start:end] - + exnum_chr = "%x" % exon_count exnum_proc += exnum_chr * (end-start) exon_count = (exon_count + 1) % 0x10 - + # DEBUG if opt.debug: - print seq_proc - print ann_proc - print rf_proc - print exnum_proc + print(seq_proc) + print(ann_proc) + print(rf_proc) + print(exnum_proc) # Process comments if opt.keepcomment or opt.processcomment: @@ -928,37 +929,37 @@ def parseOpts(): rf_proc = "012" * (len(ann_proc) / 3) rf_proc += "012"[:len(ann_proc) % 3] - for i in range(1,len(pa)*3): # Skip first position + for i in range(1,len(pa)*3): # Skip first position if ann_proc[i] == "(": pa[i/3] = rf_proc[i] # Store translated sequence prot_ann = "".join(pa) pepseqs.append( (name,pep,prot_ann,com.strip()) ) - + # Pretty printing for the report title = "%s - " % (name) if opt.exonann: title += "translation and annotation of the exonic structure" - + elif opt.intronrf: title += "translation and annotation of the position and phase of the introns" - + vals = [pep,prot_ann] labels = ["pep:","ann:"] l = makePretty(title,vals,labels,len(pep)) reports.append(l) - + # Output the results ------------------------------------------------------- # Step 1) Combined results. if opt.outfile: outstrem = openForWriteOrDie(opt.outfile) else: outstream = sys.stdout - + if opt.outformat in ["REPORT","AUTO"]: for l in reports: outstream.writelines(l) - print >> outstream, "//" - + print("//", file=outstream) + if "TAB" == opt.outformat: outFasta = False elif "FASTA" == opt.outformat: @@ -970,24 +971,24 @@ def parseOpts(): writeFasta(pepseqs,outstream) else: writeTab(pepseqs,outstream) - - #outstream.close() + + #outstream.close() # Step 2) Write specifik sub-result if requested if opt.reportfile: if opt.reportfile == "-": outstream = sys.stdout else: outstream = openForWriteOrDie(opt.reportfile) - + for l in reports: outstream.writelines(l) - + if opt.fastafile: if opt.fastafile == "-": outstream = sys.stdout else: outstream = openForWriteOrDie(opt.fastafile) - + writeFasta(pepseqs,outstream) - + if opt.tabfile: if opt.tabfile == "-": outstream = sys.stdout else: outstream = openForWriteOrDie(opt.tabfile) - + writeTab(pepseqs,outstream) diff --git a/bin/virtualribosomev2/mod_translate.py b/bin/virtualribosomev2/mod_translate.py index c9017f9..a555603 100644 --- a/bin/virtualribosomev2/mod_translate.py +++ b/bin/virtualribosomev2/mod_translate.py @@ -1,4 +1,4 @@ -#!/usr/local/python/bin/python +#!/usr/bin/env python3 # Copyright 2002,2003,2004,2005 Rasmus Wernersson, Technical University of Denmark # @@ -35,7 +35,7 @@ def __init__(self): self.description = "" self.d_all = {} self.d_first = {} - + def toString(self): return "Description: %s\nd_all: %s\nd_first: %s" % (self.description,str(self.d_all),str(self.d_first)) @@ -96,7 +96,7 @@ def toString(self): iupac["N"] = "ACGT" #aNy -alphaDNA = "ACGTRYMKWSBDHVN" +alphaDNA = "ACGTRYMKWSBDHVN" alphaDNAStrict = "ACGT" alphaPep ="*ACDEFGHIKLMNPQRSTVWY" @@ -112,19 +112,19 @@ def parseNcbiTable(lines): for line in lines.split("\n"): line = line.strip() #print "!!!"+line - + if line.startswith("name ") and (desc == ""): dRec.description += line.split('"')[1]+" " - + elif line.startswith("id "): tab_id = line.split()[1] - + elif line.startswith("ncbieaa"): aa_all = line.split('"')[1] - + elif line.startswith("sncbieaa"): aa_first = line.split('"')[1] - + c = 0 for b1 in "TCAG": for b2 in "TCAG": @@ -134,7 +134,7 @@ def parseNcbiTable(lines): aaf = aa_first[c] if aaf == "-": aaf = aa_all[c] dRec.d_first[codon] = aaf - + c += 1 result[tab_id] = dRec dRec = TransTableRec() @@ -145,51 +145,51 @@ def parseMatrixLines(iterator): result = {} for line in iterator: line = line.strip() - - if not line: + + if not line: continue # Ignore blank lines - if line.startswith("#"): + if line.startswith("#"): continue # Ignore comment lines - + tokens = line.split() try: codon, aa = tokens - + # Skip invalid entries - if len(codon) <> 3: + if len(codon) != 3: badCodon = 1 else: codon = codon.upper().replace("U","T") - for c in codon: + for c in codon: if not c in alphaDNAStrict: badCodon = 1 badCodon = 0 - + if badCodon: raise "Bad codon: %s [%s]" % (codon,line) - - - if len(aa) <> 1: #or (not aa in alphaPep): + + + if len(aa) != 1: #or (not aa in alphaPep): raise "Bad aa: %s [%s]" % (aa,line) - + result[codon] = aa - - except Exception, e: + + except Exception as e: if DEBUG: sys.stderr.write("Matrix Error - %s\n" % e) - + if len(d) != 64 and DEBUG: sys.stderr.write("Matrix Error - size of matrix differs from 64 [%i]\n" % len(d)) - + return result - + def parseMatrixFile(filename): - if d_ncbi_table.has_key(filename): + if filename in d_ncbi_table: dRec = d_ncbi_table[filename] return dRec - + dRec = TransTableRec() - dRec.d_all = parseMatrixLines(open(filename,"r").xreadlines()) + dRec.d_all = parseMatrixLines(open(filename,"r")) dRec.d_first = dRec.d_all dRec.description = "Custom translation table '%s'" % filename return dRec @@ -208,29 +208,29 @@ def trim_old(seq): # Assumption: Degenerate codons are rare - speed is not an issue def decode(codon,dRec,isFirst): if len(codon) != 3: return [] - + #Use the relevant translation table if isFirst: d_gc = dRec.d_first else: d_gc = dRec.d_all - + #Check for the simple case: this is a standard non-degenerate codon - if d_gc.has_key(codon): + if codon in d_gc: return [ d_gc[codon]] - + #The codon is to some degree degenerate - start the whole recursive scheme result = [] - + for i in range(0,3): p = iupac[codon[i]] if (len(p) > 1): for c in p: result += (decode(codon[0:i]+c+codon[i+1:3],dRec,isFirst)) return result - + if len(p) == 0: return [] # Unknown/illegal char #return [d[codon]] - + def condense(lst): result = [] for e in lst: @@ -239,7 +239,7 @@ def condense(lst): def translate(seq,transRec): return translate(seq,transRec,True,True) - + def translate(seq,transRec,firstIsStartCodon,readThroughStopCodon): debug = False if not transRec: @@ -248,14 +248,14 @@ def translate(seq,transRec,firstIsStartCodon,readThroughStopCodon): result = [] seq = trim(seq) if firstIsStartCodon: isFirst = True - else: isFirst = False + else: isFirst = False for i in range(0,len(seq),3): aa = condense(decode(seq[i:i+3],transRec,isFirst)) - if debug: print seq[i:i+3], aa + if debug: print(seq[i:i+3], aa) if aa: - if aa[0] == "*" and not readThroughStopCodon: + if aa[0] == "*" and not readThroughStopCodon: break - + if len(aa) == 1: result.append(aa[0]) else: s = string.join(aa,"") @@ -264,12 +264,12 @@ def translate(seq,transRec,firstIsStartCodon,readThroughStopCodon): else : result.append("X") #Any #print seq[i:i+3] isFirst = False - - pepseq = "".join(result) + + pepseq = "".join(result) if debug: - print pepseq + print(pepseq) return pepseq - + # Annotate all possible start and stop codons def annotate(seq,transRec): debug = False @@ -279,18 +279,18 @@ def annotate(seq,transRec): for i in range(0,len(seq),3): codon = seq[i:i+3] aa = condense(decode(codon,transRec,True)) - if debug: print seq[i:i+3], aa + if debug: print(seq[i:i+3], aa) if aa: - if "*" in aa: + if "*" in aa: result.append("*") elif ["M"] == aa: aa_int = condense(decode(codon,transRec,False)) if aa_int == ["M"]: result.append("M") else: result.append("m") - else: - result.append(".") - - return "".join(result) + else: + result.append(".") + + return "".join(result) #def translate(seq): # translate(seq,None) @@ -298,10 +298,10 @@ def annotate(seq,transRec): try: import ncbi_genetic_codes d_ncbi_table = parseNcbiTable(ncbi_genetic_codes.ncbi_gc_table) - + except: pass - + if __name__ == "__main__": for line in sys.stdin.readlines(): - print translate(line,None,True,False) + print(translate(line,None,True,False)) From 33f5f3b0e95d772c65923eb10445b54694938ffb Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Wed, 10 Mar 2021 14:10:55 -0800 Subject: [PATCH 019/223] Change file to open. Fix tabs --- bin/virtualribosomev2/dna2pep.py | 22 +++++++++++----------- 1 file changed, 11 insertions(+), 11 deletions(-) diff --git a/bin/virtualribosomev2/dna2pep.py b/bin/virtualribosomev2/dna2pep.py index 9d4e9f9..1ab8eff 100644 --- a/bin/virtualribosomev2/dna2pep.py +++ b/bin/virtualribosomev2/dna2pep.py @@ -248,7 +248,7 @@ complDNA = "TAACGRYWSMKVHDBN" allValid = validDNA+validDNA.lower() -transTable = string.maketrans(validDNA+validDNA.lower(),complDNA+complDNA.lower()) +transTable = str.maketrans(validDNA+validDNA.lower(),complDNA+complDNA.lower()) pwidth = 90 @@ -407,7 +407,7 @@ def writeTab(seqs, outstream): def openForWriteOrDie(outfile): try: - outstream = file(outfile,"w") + outstream = open(outfile,"w") except IOError as xxx_todo_changeme: (strerror) = xxx_todo_changeme @@ -417,11 +417,11 @@ def openForWriteOrDie(outfile): return outstream def parseOpts(): - # Quick hack to overrule the -h and --help feature - # build into the optpase module - if "-h" in sys.argv or "--help" in sys.argv: - print(__doc__) - sys.exit(0) + # Quick hack to overrule the -h and --help feature + # build into the optpase module + if "-h" in sys.argv or "--help" in sys.argv: + print(__doc__) + sys.exit(0) parser = OptionParser() @@ -454,10 +454,10 @@ def parseOpts(): parser.add_option("-c","--comment", action = "store_true", dest = "keepcomment", default=False) parser.add_option("-C","--processcomment", action = "store_true", dest = "processcomment", default=False) - # Debug - parser.add_option("-d","--debug", action="store_true", dest="debug", default=False) + # Debug + parser.add_option("-d","--debug", action="store_true", dest="debug", default=False) - (opt, args) = parser.parse_args() + (opt, args) = parser.parse_args() # Check reading frame if not opt.readingframe in ["1", "2", "3", "-1", "-2", "-3", "all", "plus","minus"]: @@ -505,7 +505,7 @@ def parseOpts(): lines = [] for fn in args: try: - lines += (file(fn,"r").readlines()) + lines += (open(fn,"r").readlines()) except: sys.stderr.write("ERROR: Cannot read from file '%s'\n" % fn) sys.exit(-1) From e1d01d1864484325c37f8477d3c24a41f86404b8 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Wed, 10 Mar 2021 14:13:55 -0800 Subject: [PATCH 020/223] Change permissions --- bin/virtualribosomev2/dna2pep.py | 0 bin/virtualribosomev2/mod_translate.py | 0 2 files changed, 0 insertions(+), 0 deletions(-) mode change 100644 => 100755 bin/virtualribosomev2/dna2pep.py mode change 100644 => 100755 bin/virtualribosomev2/mod_translate.py diff --git a/bin/virtualribosomev2/dna2pep.py b/bin/virtualribosomev2/dna2pep.py old mode 100644 new mode 100755 diff --git a/bin/virtualribosomev2/mod_translate.py b/bin/virtualribosomev2/mod_translate.py old mode 100644 new mode 100755 From 412b1d4a6f714b7589b45cf83482b85edddd936f Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Wed, 10 Mar 2021 14:24:31 -0800 Subject: [PATCH 021/223] Remove py2 processes(translation) --- vAMPirus.nf | 453 +++++++++++++++++++++------------------------------- 1 file changed, 181 insertions(+), 272 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 8810a6b..3870224 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -938,225 +938,194 @@ if (params.DataCheck || params.Analyze) { } } - if (params.sing) { + process Translation_For_ProteinBased_Clustering_DC { - process Translating_For_ProteinClustering_DC { + label 'norm_cpus' - label 'low_cpus' + publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Aminoacid/translation", mode: "copy", overwrite: true - publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Aminoacid/translation", mode: "copy", overwrite: true + input: + file(fasta) from nucl2aa - input: - file(fasta) from nucl2aa + output: + file("*ASVprotforclust.fasta") into clustering_aa + file("*_translation_report") into reportaa_VR + file("*_ASV_all.fasta") into asvfastaforaaclust - output: - file("*ASVprotforclust.fasta") into clustering_aa - file("*_translation_report") into reportaa_VR - file("*_ASV_all.fasta") into asvfastaforaaclust + script: + """ + ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_ASVprotforclust.fasta --report ${params.projtag}_translation_report + cp ${fasta} ${params.projtag}_ASV_all.fasta + """ + } - script: - """ - conda init && source activate virtualribosome + process Protein_clustering_DC { - ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_ASVprotforclust.fasta --report ${params.projtag}_translation_report - cp ${fasta} ${params.projtag}_ASV_all.fasta - """ + label 'norm_cpus' - } + publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Aminoacid", mode: "copy", overwrite: true, pattern: '*{.csv}' - } else { + input: + file(fasta) from clustering_aa + file(asvs) from asvfastaforaaclust - process Translation_For_ProteinBased_Clustering_DC { + output: + file("number_per_percentage_prot.csv") into number_per_percent_prot_plot - label 'norm_cpus' + script: + // add awk script to count seqs + """ + set +e + cp ${params.vampdir}/bin/rename_seq.py . + for id in `echo ${params.datacheckaaIDlist} | tr "," "\\n"`;do + if [ \${id} == ".55" ];then + word=3 + elif [ \${id} == ".65" ];then + word=4 + else + word=5 + fi + awk 'BEGIN{RS=">";ORS=""}length(\$2)>="${params.minAA}"{print ">"\$0}' ${fasta} > ${params.projtag}_filtered_proteins.fasta + cd-hit -i ${params.projtag}_filtered_proteins.fasta -n \${word} -c \${id} -o ${params.projtag}_pcASV\${id}.fasta + sed 's/>Cluster />Cluster_/g' ${params.projtag}_pcASV\${id}.fasta.clstr >${params.projtag}_pcASV\${id}.clstr + grep ">Cluster_" ${params.projtag}_pcASV\${id}.clstr >temporaryclusters.list + y=\$(grep -c ">Cluster_" ${params.projtag}_pcASV\${id}.clstr) + echo ">Cluster_"\${y}"" >> ${params.projtag}_pcASV\${id}.clstr + t=1 + b=1 + for x in \$(cat temporaryclusters.list);do + echo "Extracting \$x" + name="\$( echo \$x | awk -F ">" '{print \$2}')" + clust="pcASV"\${t}"" + echo "\${name}" + awk '/^>'\${name}'\$/,/^>Cluster_'\${b}'\$/' ${params.projtag}_pcASV\${id}.clstr > "\${name}"_"\${clust}"_tmp.list + t=\$(( \${t}+1 )) + b=\$(( \${b}+1 )) + done + ls *_tmp.list + u=1 + for x in *_tmp.list;do + name="\$(echo \$x | awk -F "_p" '{print \$1}')" + echo "\${name}" + cluster="\$(echo \$x | awk -F "_" '{print \$3}')" + echo "\${cluster}" + grep "ASV" \$x | awk -F ", " '{print \$2}' | awk -F "_" '{print \$1}' | awk -F ">" '{print \$2}' > \${name}_\${cluster}_seqs_tmps.list + seqtk subseq ${asvs} \${name}_\${cluster}_seqs_tmps.list > \${name}_\${cluster}_nucleotide_sequences.fasta + vsearch --cluster_fast \${name}_\${cluster}_nucleotide_sequences.fasta --id 0.2 --centroids \${name}_\${cluster}_centroids.fasta + grep ">" \${name}_\${cluster}_centroids.fasta >> \${name}_\${cluster}_tmp_centroids.list + for y in \$( cat \${name}_\${cluster}_tmp_centroids.list );do + echo ">\${cluster}_type"\$u"" >> \${name}_\${cluster}_tmp_centroid.newheaders + u=\$(( \${u}+1 )) + done + u=1 + ./rename_seq.py \${name}_\${cluster}_centroids.fasta \${name}_\${cluster}_tmp_centroid.newheaders \${cluster}_types_labeled.fasta + done + cat *_types_labeled.fasta >> ${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta + grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$3}' | awk -F "." '{print \$1}' >tmphead.list + grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$2}' | awk -F "," '{print \$1}' >tmplen.list + paste -d"," temporaryclusters.list tmphead.list >tmp.info.csv + grep ">" ${params.projtag}_pcASV\${id}.fasta >lala.list + j=1 + for x in \$(cat lala.list);do + echo ">${params.projtag}_pcASV\${j}" >>${params.projtag}_aminoheaders.list + echo "\${x},>${params.projtag}_pcASV\${j}" >>tmpaminotype.info.csv + j=\$(( \${j}+1 )) + done + rm lala.list + awk -F "," '{print \$2}' tmp.info.csv >>tmporder.list + for x in \$(cat tmporder.list);do + grep -w "\$x" tmpaminotype.info.csv | awk -F "," '{print \$2}' >>tmpder.list + done + paste -d "," temporaryclusters.list tmplen.list tmphead.list tmpder.list >${params.projtag}_pcASVCluster\${id}_summary.csv + ./rename_seq.py ${params.projtag}_pcASV\${id}.fasta ${params.projtag}_aminoheaders.list ${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta + stats.sh in=${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_aminoacid_clustered.gc gcformat=4 overwrite=true + stats.sh in=${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_nucleotide_clustered.gc gcformat=4 overwrite=true + awk 'BEGIN{RS=">";ORS=""}length(\$2)<"${params.minAA}"{print ">"\$0}' ${fasta} >${params.projtag}_pcASV\${id}_problematic_translations.fasta + if [ `wc -l ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk '{print \$1}'` -gt 1 ];then + grep ">" ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk -F ">" '{print \$2}' > problem_tmp.list + seqtk subseq ${asvs} problem_tmp.list > ${params.projtag}_pcASV\${id}_problematic_nucleotides.fasta + else + rm ${params.projtag}_pcASV\${id}_problematic_translations.fasta + fi + rm *.list + rm Cluster* + rm *types* + rm *tmp* + rm ${params.projtag}_pcASV\${id}.fast* + done + for x in *aminoacid*noTaxonomy.fasta;do + id=\$( echo \$x | awk -F "_noTax" '{print \$1}' | awk -F "pcASV" '{print \$2}') + numb=\$( grep -c ">" \$x) + echo "\${id},\${numb}" >> number_per_percentage_protz.csv + done + yesirr=\$( wc -l number_per_percentage_protz.csv | awk '{print \$1}') + tail -\$(( \${yesirr}-1 )) number_per_percentage_protz.csv > number_per_percentage_prot.csv + head -1 number_per_percentage_protz.csv >> number_per_percentage_prot.csv + rm number_per_percentage_protz.csv + """ + } - conda 'python=2.7' + if (!params.skipReadProcessing || !params.skipMerging ) { - publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Aminoacid/translation", mode: "copy", overwrite: true + process combine_csv_DC { - input: - file(fasta) from nucl2aa + input: + file(csv) from fastp_csv_in1 + .collect() output: - file("*ASVprotforclust.fasta") into clustering_aa - file("*_translation_report") into reportaa_VR - file("*_ASV_all.fasta") into asvfastaforaaclust + file("final_reads_stats.csv") into fastp_csv_dc script: """ - ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_ASVprotforclust.fasta --report ${params.projtag}_translation_report - cp ${fasta} ${params.projtag}_ASV_all.fasta + cat ${csv} >all_reads_stats.csv + head -n1 all_reads_stats.csv >tmp.names.csv + cat all_reads_stats.csv | grep -v ""Sample,Total_"" >tmp.reads.stats.csv + cat tmp.names.csv tmp.reads.stats.csv >final_reads_stats.csv + rm tmp.names.csv tmp.reads.stats.csv """ - } - } - process Protein_clustering_DC { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Aminoacid", mode: "copy", overwrite: true, pattern: '*{.csv}' - - input: - file(fasta) from clustering_aa - file(asvs) from asvfastaforaaclust + } + } else { + process skip_combine_csv_DC { output: - file("number_per_percentage_prot.csv") into number_per_percent_prot_plot + file("filter_reads.txt") into fastp_csv_dc script: - // add awk script to count seqs """ - set +e - cp ${params.vampdir}/bin/rename_seq.py . - for id in `echo ${params.datacheckaaIDlist} | tr "," "\\n"`;do - if [ \${id} == ".55" ];then - word=3 - elif [ \${id} == ".65" ];then - word=4 - else - word=5 - fi - awk 'BEGIN{RS=">";ORS=""}length(\$2)>="${params.minAA}"{print ">"\$0}' ${fasta} > ${params.projtag}_filtered_proteins.fasta - cd-hit -i ${params.projtag}_filtered_proteins.fasta -n \${word} -c \${id} -o ${params.projtag}_pcASV\${id}.fasta - sed 's/>Cluster />Cluster_/g' ${params.projtag}_pcASV\${id}.fasta.clstr >${params.projtag}_pcASV\${id}.clstr - grep ">Cluster_" ${params.projtag}_pcASV\${id}.clstr >temporaryclusters.list - y=\$(grep -c ">Cluster_" ${params.projtag}_pcASV\${id}.clstr) - echo ">Cluster_"\${y}"" >> ${params.projtag}_pcASV\${id}.clstr - t=1 - b=1 - for x in \$(cat temporaryclusters.list);do - echo "Extracting \$x" - name="\$( echo \$x | awk -F ">" '{print \$2}')" - clust="pcASV"\${t}"" - echo "\${name}" - awk '/^>'\${name}'\$/,/^>Cluster_'\${b}'\$/' ${params.projtag}_pcASV\${id}.clstr > "\${name}"_"\${clust}"_tmp.list - t=\$(( \${t}+1 )) - b=\$(( \${b}+1 )) - done - ls *_tmp.list - u=1 - for x in *_tmp.list;do - name="\$(echo \$x | awk -F "_p" '{print \$1}')" - echo "\${name}" - cluster="\$(echo \$x | awk -F "_" '{print \$3}')" - echo "\${cluster}" - grep "ASV" \$x | awk -F ", " '{print \$2}' | awk -F "_" '{print \$1}' | awk -F ">" '{print \$2}' > \${name}_\${cluster}_seqs_tmps.list - seqtk subseq ${asvs} \${name}_\${cluster}_seqs_tmps.list > \${name}_\${cluster}_nucleotide_sequences.fasta - vsearch --cluster_fast \${name}_\${cluster}_nucleotide_sequences.fasta --id 0.2 --centroids \${name}_\${cluster}_centroids.fasta - grep ">" \${name}_\${cluster}_centroids.fasta >> \${name}_\${cluster}_tmp_centroids.list - for y in \$( cat \${name}_\${cluster}_tmp_centroids.list );do - echo ">\${cluster}_type"\$u"" >> \${name}_\${cluster}_tmp_centroid.newheaders - u=\$(( \${u}+1 )) - done - u=1 - ./rename_seq.py \${name}_\${cluster}_centroids.fasta \${name}_\${cluster}_tmp_centroid.newheaders \${cluster}_types_labeled.fasta - done - cat *_types_labeled.fasta >> ${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta - grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$3}' | awk -F "." '{print \$1}' >tmphead.list - grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$2}' | awk -F "," '{print \$1}' >tmplen.list - paste -d"," temporaryclusters.list tmphead.list >tmp.info.csv - grep ">" ${params.projtag}_pcASV\${id}.fasta >lala.list - j=1 - for x in \$(cat lala.list);do - echo ">${params.projtag}_pcASV\${j}" >>${params.projtag}_aminoheaders.list - echo "\${x},>${params.projtag}_pcASV\${j}" >>tmpaminotype.info.csv - j=\$(( \${j}+1 )) - done - rm lala.list - awk -F "," '{print \$2}' tmp.info.csv >>tmporder.list - for x in \$(cat tmporder.list);do - grep -w "\$x" tmpaminotype.info.csv | awk -F "," '{print \$2}' >>tmpder.list - done - paste -d "," temporaryclusters.list tmplen.list tmphead.list tmpder.list >${params.projtag}_pcASVCluster\${id}_summary.csv - ./rename_seq.py ${params.projtag}_pcASV\${id}.fasta ${params.projtag}_aminoheaders.list ${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta - stats.sh in=${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_aminoacid_clustered.gc gcformat=4 overwrite=true - stats.sh in=${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_nucleotide_clustered.gc gcformat=4 overwrite=true - awk 'BEGIN{RS=">";ORS=""}length(\$2)<"${params.minAA}"{print ">"\$0}' ${fasta} >${params.projtag}_pcASV\${id}_problematic_translations.fasta - if [ `wc -l ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk '{print \$1}'` -gt 1 ];then - grep ">" ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk -F ">" '{print \$2}' > problem_tmp.list - seqtk subseq ${asvs} problem_tmp.list > ${params.projtag}_pcASV\${id}_problematic_nucleotides.fasta - else - rm ${params.projtag}_pcASV\${id}_problematic_translations.fasta - fi - rm *.list - rm Cluster* - rm *types* - rm *tmp* - rm ${params.projtag}_pcASV\${id}.fast* - done - for x in *aminoacid*noTaxonomy.fasta;do - id=\$( echo \$x | awk -F "_noTax" '{print \$1}' | awk -F "pcASV" '{print \$2}') - numb=\$( grep -c ">" \$x) - echo "\${id},\${numb}" >> number_per_percentage_protz.csv - done - yesirr=\$( wc -l number_per_percentage_protz.csv | awk '{print \$1}') - tail -\$(( \${yesirr}-1 )) number_per_percentage_protz.csv > number_per_percentage_prot.csv - head -1 number_per_percentage_protz.csv >> number_per_percentage_prot.csv - rm number_per_percentage_protz.csv + echo "Read processing steps skipped." >filter_reads.txt """ } + } - if (!params.skipReadProcessing || !params.skipMerging ) { - - process combine_csv_DC { - - input: - file(csv) from fastp_csv_in1 - .collect() - - output: - file("final_reads_stats.csv") into fastp_csv_dc - - script: - """ - cat ${csv} >all_reads_stats.csv - head -n1 all_reads_stats.csv >tmp.names.csv - cat all_reads_stats.csv | grep -v ""Sample,Total_"" >tmp.reads.stats.csv - cat tmp.names.csv tmp.reads.stats.csv >final_reads_stats.csv - rm tmp.names.csv tmp.reads.stats.csv - """ - - } - } else { - - process skip_combine_csv_DC { - output: - file("filter_reads.txt") into fastp_csv_dc - - script: - """ - echo "Read processing steps skipped." >filter_reads.txt - """ - } - } - - report_dc_in = Channel.create() - fastp_csv_dc.mix( reads_per_sample_preFilt, reads_per_sample_postFilt, prefilt_basefreq, postFilt_basefreq, prefilt_qualityscore, postFilt_qualityscore, prefilt_gccontent, postFilt_gccontent, prefilt_averagequality, postFilt_averagequality, prefilt_length, postFilt_length, number_per_percent_nucl_plot, number_per_percent_prot_plot - ).into(report_dc_in) + report_dc_in = Channel.create() + fastp_csv_dc.mix( reads_per_sample_preFilt, reads_per_sample_postFilt, prefilt_basefreq, postFilt_basefreq, prefilt_qualityscore, postFilt_qualityscore, prefilt_gccontent, postFilt_gccontent, prefilt_averagequality, postFilt_averagequality, prefilt_length, postFilt_length, number_per_percent_nucl_plot, number_per_percent_prot_plot + ).into(report_dc_in) - process Report_DataCheck { + process Report_DataCheck { - label 'norm_cpus' + label 'norm_cpus' - publishDir "${params.workingdir}/${params.outdir}/DataCheck/Report", mode: "copy", overwrite: true, pattern: '*.{html}' + publishDir "${params.workingdir}/${params.outdir}/DataCheck/Report", mode: "copy", overwrite: true, pattern: '*.{html}' - input: - file(files) from report_dc_in - .collect() + input: + file(files) from report_dc_in + .collect() - output: - file("*.html") into datacheckreport + output: + file("*.html") into datacheckreport - script: - """ - cp ${params.vampdir}/bin/vAMPirus_DC_Report.Rmd . - cp ${params.vampdir}/example_data/conf/vamplogo.png . - Rscript -e "rmarkdown::render('vAMPirus_DC_Report.Rmd',output_file='${params.projtag}_DataCheck_Report.html')" ${params.projtag} \ - ${params.skipReadProcessing} \ - ${params.skipMerging} \ - ${params.skipAdapterRemoval} - """ - } + script: + """ + cp ${params.vampdir}/bin/vAMPirus_DC_Report.Rmd . + cp ${params.vampdir}/example_data/conf/vamplogo.png . + Rscript -e "rmarkdown::render('vAMPirus_DC_Report.Rmd',output_file='${params.projtag}_DataCheck_Report.html')" ${params.projtag} \ + ${params.skipReadProcessing} \ + ${params.skipMerging} \ + ${params.skipAdapterRemoval} + """ + } } else if (params.Analyze) { @@ -1994,53 +1963,24 @@ if (params.DataCheck || params.Analyze) { if (!params.skipAminoTyping) { - if (params.sing) { - - process Translating_For_Aminotypes { - - label 'low_cpus' + process Translate_For_AminoTyping { - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes/Translation", mode: "copy", overwrite: true - - input: - file(fasta) from asvsforAminotyping - - output: - file("${params.projtag}_all_translations.fasta") into amintypegen - file("${params.projtag}_translation_report") into proteinstage_vap_report - - script: - """ - conda init && source activate virtualribosome - - ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_all_translations.fasta --report ${params.projtag}_translation_report - """ - - } - - } else { - - process Translate_For_AminoTyping { - - label 'low_cpus' - - conda 'python=2.7' + label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes/Translation", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes/Translation", mode: "copy", overwrite: true - input: - file(fasta) from asvsforAminotyping + input: + file(fasta) from asvsforAminotyping - output: - file("${params.projtag}_all_translations.fasta") into amintypegen - file("${params.projtag}_translation_report") into proteinstage_vap_report + output: + file("${params.projtag}_all_translations.fasta") into amintypegen + file("${params.projtag}_translation_report") into proteinstage_vap_report - script: - """ - ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_all_translations.fasta --report ${params.projtag}_translation_report - """ - } - } + script: + """ + ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_all_translations.fasta --report ${params.projtag}_translation_report + """ + } process Generate_AminoTypes { @@ -2423,57 +2363,26 @@ if (params.DataCheck || params.Analyze) { if (params.pcASV) { // ASV_nucl -> ASV_aa -> clusteraa by %id with ch-hit -> extract representative nucl sequences to generate new OTU file - if (params.sing) { - - process Translating_For_pcASV_Generation { + process Translation_For_pcASV_Generation { - label 'low_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV/Translation", mode: "copy", overwrite: true, pattern: '*_ASV_translations*' - - input: - file(fasta) from nucl2aa - - output: - file("*ASV*translations.fasta") into clustering_aa - file("*_ASV_translations_report") into reportaa_VR - file("*_ASV_nucleotide.fasta") into asvfastaforaaclust - - script: - """ - conda init && source activate virtualribosome - - ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_ASV_translations.fasta --report ${params.projtag}_ASV_translations_report - cp ${fasta} ${params.projtag}_ASV_nucleotide.fasta - """ - - } - - } else { - - process Translation_For_pcASV_Generation { - - label 'low_cpus' - - conda 'python=2.7' + label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV/Translation", mode: "copy", overwrite: true, pattern: '*_ASV_translations*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV/Translation", mode: "copy", overwrite: true, pattern: '*_ASV_translations*' - input: - file(fasta) from nucl2aa + input: + file(fasta) from nucl2aa - output: - file("*ASV*translations.fasta") into clustering_aa - file("*_ASV_translations_report") into reportaa_VR - file("*_ASV_nucleotide.fasta") into asvfastaforaaclust + output: + file("*ASV*translations.fasta") into clustering_aa + file("*_ASV_translations_report") into reportaa_VR + file("*_ASV_nucleotide.fasta") into asvfastaforaaclust - script: - """ - ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_ASV_translations.fasta --report ${params.projtag}_ASV_translations_report - cp ${fasta} ${params.projtag}_ASV_nucleotide.fasta - """ - } - } + script: + """ + ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_ASV_translations.fasta --report ${params.projtag}_ASV_translations_report + cp ${fasta} ${params.projtag}_ASV_nucleotide.fasta + """ + } process Generate_pcASVs { From b97a41ad6f4034ce333d8411c88ffac7d3a41203 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Wed, 10 Mar 2021 16:03:34 -0800 Subject: [PATCH 022/223] Refactor code. Divide asv and ncasv processes --- vAMPirus.nf | 1077 +++++++++++++++++++++------------------------------ 1 file changed, 452 insertions(+), 625 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 3870224..38124f7 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1131,6 +1131,9 @@ if (params.DataCheck || params.Analyze) { if (params.ncASV) { + reads_vsearch5_ch + .into{ asv_file_for_ncasvs; nuclFastas_forDiamond_asv_ch; nuclFastas_forCounts_asv_ch; nuclFastas_forphylogeny_asv; nuclFastas_forMatrix_asv_ch } + process NucleotideBased_ASV_clustering { label 'norm_cpus' @@ -1139,55 +1142,40 @@ if (params.DataCheck || params.Analyze) { input: each x from 1..nnuc - file(fasta) from reads_vsearch5_ch + file(fasta) from asv_file_for_ncasvs output: - //HERE - sepearte avs and ncasv out and proceeses. matrix, couts and phylogeny - tuple file("*_ncASV*.fasta"), file("*ASV.fasta") into ( nuclFastas_forDiamond_ch, nuclFastas_forCounts_ch, nuclFastas_forMatrix_ch) - file("*_ncASV*.fasta") into nuclFastas_forphylogeny_ncasv - file("*ASV.fasta") into nuclFastas_forphylogeny_asv + file("*_ncASV*.fasta") into ( nuclFastas_forphylogeny_ncasv, nuclFastas_forDiamond_ncasv_ch, nuclFastas_forCounts_ncasv_ch, nuclFastas_forMatrix_ncasv_ch ) script: if (params.clusterNuclIDlist) { nid=slist1.get(x-1) """ - cp ${fasta} ./${params.projtag}_ASV.fasta - vsearch --cluster_fast ${params.projtag}_ASV.fasta --centroids ${params.projtag}_ncASV\${id}.fasta --threads ${task.cpus} --relabel ncASV --id ${nid} + vsearch --cluster_fast ${fasta} --centroids ${params.projtag}_ncASV\${id}.fasta --threads ${task.cpus} --relabel ncASV --id ${nid} """ } else if (params.clusterNuclID) { """ - cp ${fasta} ./${params.projtag}_ASV.fasta - id=${params.clusterNuclID} - vsearch --cluster_fast ${params.projtag}_ASV.fasta --centroids ${params.projtag}_ncASV\${id}.fasta --threads ${task.cpus} --relabel ncASV --id \${id} + vsearch --cluster_fast ${fasta} --centroids ${params.projtag}_ncASV\${id}.fasta --threads ${task.cpus} --relabel ncASV --id ${params.clusterNuclID} """ } } - } else { - reads_vsearch5_ch - .into{ nuclFastas_forDiamond_ch; nuclFastas_forCounts_ch; nuclFastas_forphylogeny; nuclFastas_forMatrix_ch } - } - - if (!params.skipTaxonomy) { - if (params.ncASV) { + if (!params.skipTaxonomy) { process Nucleotide_Taxonomy_Inference { label 'high_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy", mode: "copy", overwrite: true, pattern: '*_ASV*.{fasta,csv,tsv}' publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Taxonomy", mode: "copy", overwrite: true, pattern: '*ncASV*.{fasta,csv,tsv}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*_ASV*dmd.out' publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*ncASV*dmd.out' input: - tuple file(notus), file(asvs) from nuclFastas_forDiamond_ch + file(asvs) from nuclFastas_forDiamond_ncasv_ch output: - file("*.fasta") into tax_labeled_fasta - tuple file("*_phyloseqObject.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond - file("*ncASV*summary_for_plot.csv") into taxplot1a - file("*_ASV*_summary_for_plot.csv") into taxplot1 + file("*.fasta") into tax_labeled_fasta1 + tuple file("*_phyloseqObject.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond_ncasv + file("*ncASV*summary_for_plot.csv") into taxplot_ncasv script: """ @@ -1195,133 +1183,6 @@ if (params.DataCheck || params.Analyze) { virdb=${params.dbdir}/${params.dbname} grep ">" \${virdb} > headers.list headers="headers.list" - for filename in ${notus};do - name=\$(ls \${filename} | awk -F ".fasta" '{print \$1}') - diamond blastx -q \${filename} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 - echo "Preparing lists to generate summary .csv's" - echo "[Best hit accession number]" > access.list - echo "[e-value]" > evalue.list - echo "[Bitscore]" > bit.list - echo "[Percent ID (aa)]" > pid.list - echo "[Organism ID]" > "\$name"_virus.list - echo "[Gene]" > "\$name"_genes.list - grep ">" \${filename} | awk -F ">" '{print \$2}' > seqids.lst - echo "extracting genes and names" - touch new_"\$name"_asvnames.txt - j=1 - if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then - echo "[ncASV#]" > otu.list - echo "[ncASV sequence length]" > length.list - for s in \$(cat seqids.lst);do - echo "Checking for \$s hit in diamond output" - if [[ ${params.refseq} == "T" ]];then - echo "RefSeq headers specified" - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">ncASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list - virus="NO" - gene="HIT" - echo ">ncASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - fi - else - echo "Using RVDB headers." - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">ncASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list - virus="NO" - gene="HIT" - echo ">ncASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - fi - fi - echo "Done with \$s" - done - fi - echo "Now editing "\$name" fasta headers" - ###### rename_seq.py - ./rename_seq.py \${filename} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta - awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta - echo "[Sequence header]" > newnames.list - cat new_"\$name"_asvnames.txt >> newnames.list - touch sequence.list - echo " " > sequence.list - grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list - rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - for x in *phyloseqObject.csv;do - echo "\$x" - lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) - tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; - sed 's/ /_/g' tmpcol.list > tmp2col.list; - cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; - rm tmpcol.list tmp2col.list - done - rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; - rm *asvnames.txt - rm "\$name"_virus.list - rm "\$name"_genes.list - rm newnames.list - rm access.list - echo "Taxonomy inferred for: \${filename} " - done for filename in ${asvs};do name=\$(ls \${filename} | awk -F ".fasta" '{print \$1}') diamond blastx -q \${filename} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 @@ -1452,513 +1313,479 @@ if (params.DataCheck || params.Analyze) { rm headers.list """ } + } - } else { + process Generate_ncASV_Counts_Table { - process ASV_Taxonomy_Inference { + label 'norm_cpus' - label 'high_cpus' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Counts", mode: "copy", overwrite: true, pattern: '*_ASV*.{biome,csv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Counts", mode: "copy", overwrite: true, pattern: '*ncASV*.{biome,csv}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy", mode: "copy", overwrite: true, pattern: '*_ASV*.{fasta,csv,tsv}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*_ASV*dmd.out' + input: + file(notus) from nuclFastas_forCounts_ncasv_ch + file(merged) from nuclCounts_mergedreads_ch - input: - file(reads) from nuclFastas_forDiamond_ch + output: + tuple file("*_counts.csv"), file("*_counts.biome") into counts_vsearch + tuple val("${id}"), file("*ncASV*counts.csv") into notu_counts_plots + file("*_ASV*counts.csv") into asv_counts_plots - output: - file("*.fasta") into tax_labeled_fasta - tuple file("*_phyloseqObject.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond - file("*_ASV*_summary_for_plot.csv") into taxplot1 - script: - """ - cp ${params.vampdir}/bin/rename_seq.py . - virdb=${params.dbdir}/${params.dbname} - grep ">" \${virdb} > headers.list - headers="headers.list" - for filename in ${reads};do - name=\$(ls \${filename} | awk -F ".fasta" '{print \$1}') - diamond blastx -q \${filename} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 - echo "Preparing lists to generate summary .csv's" - echo "[Best hit accession number]" > access.list - echo "[e-value]" > evalue.list - echo "[Bitscore]" > bit.list - echo "[Percent ID (aa)]" > pid.list - echo "[Organism ID]" > "\$name"_virus.list - echo "[Gene]" > "\$name"_genes.list - grep ">" \${filename} | awk -F ">" '{print \$2}' > seqids.lst - echo "extracting genes and names" - touch new_"\$name"_asvnames.txt - j=1 - if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then - echo "[ASV#]" > otu.list - echo "[ASV sequence length]" > length.list - for s in \$(cat seqids.lst);do - echo "Checking for \$s hit in diamond output" - if [[ ${params.refseq} == "T" ]];then - echo "RefSeq headers specified" - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list - virus="NO" - gene="HIT" - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - fi - else - echo "Using RVDB headers." - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list - virus="NO" - gene="HIT" - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - fi - fi - echo "Done with \$s" - done - else - for s in \$(cat seqids.lst);do - echo "[ASV#]" > otu.list - echo "[ASV sequence length]" > length.list - echo "Checking for \$s hit in diamond output" - if [[ ${params.refseq} == "T" ]];then - echo "RefSeq headers specified" - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list - virus="NO" - gene="HIT" - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - fi - else - echo "Using RVDB headers." - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list - virus="NO" - gene="HIT" - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - fi - fi - echo "Done with \$s" - done - fi - echo "Now editing "\$name" fasta headers" - ###### rename_seq.py - ./rename_seq.py \${filename} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta - awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta - echo "[Sequence header]" > newnames.list - cat new_"\$name"_asvnames.txt >> newnames.list - touch sequence.list - echo " " > sequence.list - grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list - rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - for x in *phyloseqObject.csv;do - echo "\$x" - lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) - tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; - sed 's/ /_/g' tmpcol.list > tmp2col.list; - cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; - rm tmpcol.list tmp2col.list - done - rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; - rm *asvnames.txt - rm "\$name"_virus.list - rm "\$name"_genes.list - rm newnames.list - rm access.list - echo "Taxonomy inferred for: \${filename} " - done - rm headers.list - """ - } - } + script: + """ + for filename in ${notus};do + if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then + ident=\$( echo \${filename} | awk -F "ncASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') + name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') + vsearch --usearch_global ${merged} --db \${filename} --id \${ident} --threads ${task.cpus} --otutabout \${name}_counts.txt --biomout \${name}_counts.biome + cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv + sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv + rm \${name}_count.csv + fi + done + """ } - if (params.ncASV) { + process Generate_ncASV_Matrices { - process Generate_Counts_Tables_Nucleotide { + label 'low_cpus' - label 'norm_cpus' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Matrices", mode: "copy", overwrite: true, pattern: '*ncASV*PercentID.matrix' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Counts", mode: "copy", overwrite: true, pattern: '*_ASV*.{biome,csv}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Counts", mode: "copy", overwrite: true, pattern: '*ncASV*.{biome,csv}' + input: - input: - tuple file(notus), file(asvs) from nuclFastas_forCounts_ch - file(merged) from nuclCounts_mergedreads_ch + file(notus) from nuclFastas_forMatrix_ncasv_ch - output: - tuple file("*_counts.csv"), file("*_counts.biome") into counts_vsearch - tuple val("${id}"), file("*ncASV*counts.csv") into notu_counts_plots - file("*_ASV*counts.csv") into asv_counts_plots + output: + file("*.matrix") into clustmatrices_ncasv + file("*ncASV*PercentID.matrix") into notu_heatmap - script: - """ - for filename in ${notus};do - if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then - ident=\$( echo \${filename} | awk -F "ncASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') - name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') - vsearch --usearch_global ${merged} --db \${filename} --id \${ident} --threads ${task.cpus} --otutabout \${name}_counts.txt --biomout \${name}_counts.biome - cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv - sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv - rm \${name}_count.csv - fi - done - if [ `echo ${asvs} | grep -c "ASV"` -eq 1 ];then - name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - vsearch --usearch_global ${merged} --db ${asvs} --id ${params.asvcountID} --threads ${task.cpus} --otutabout "\$name"_counts.txt --biomout "\$name"_counts.biome - cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv - sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv - rm \${name}_count.csv + script: + // remove if statement later (no fin) + """ + for filename in ${notus};do + if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then + ident=\$( echo \${filename} | awk -F "ncASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') + name=\$( echo \${filename}| awk -F ".fasta" '{print \$1}') + clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} + clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} + for x in *q.matrix;do + pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') + ya=\$(wc -l \$x | awk '{print \$1}') + echo "\$((\$ya-1))" + tail -"\$((\$ya-1))" \$x > \${pre}z.matrix + rm \$x + cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix + rm \${pre}z.matrix + done fi - """ + done + """ } - } else { - process Generate_ASV_Counts_Tables { - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Counts", mode: "copy", overwrite: true, pattern: '*ASV*.{biome,csv}' - - input: - file(asvs) from nuclFastas_forCounts_ch - file(merged) from nuclCounts_mergedreads_ch + if (!params.skipPhylogeny) { - output: - tuple file("*_counts.csv"), file("*_counts.biome") into counts_vsearch - file("*_ASV*counts.csv") into asv_counts_plots + process ncASV_Phylogeny { + + label 'norm_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*ncASV*aln.*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*ncASV*mt*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*ncASV*iq*' + + input: + file(asvs) from nuclFastas_forphylogeny_ncasv + + output: + tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into align_results_ncasv + file("*iq.treefile") into nucl_phyl_plot_ncasv + + script: + """ + pre=\$(echo ${asvs} | awk -F ".fasta" '{print \$1}' ) + mafft --thread ${task.cpus} --maxiterate 15000 --auto ${asvs} >\${pre}_ALN.fasta + trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html + # Nucleotide_ModelTest + modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint + # Nucleotide_Phylogeny + if [ "${params.iqCustomnt}" != "" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -t \${pre}_mt.tree -T auto ${params.iqCustomnt} + elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} + elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni + elif [ "${params.nonparametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} + elif [ "${params.parametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni + else + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni + fi + """ + } - script: - """ - if [ `echo ${asvs} | grep -c "ASV"` -eq 1 ];then - name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - vsearch --usearch_global ${merged} --db ${asvs} --id ${params.asvcountID} --threads ${task.cpus} --otutabout "\$name"_counts.txt --biomout "\$name"_counts.biome - cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv - sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv - rm \${name}_count.csv - fi - """ - } } - if (params.ncASV) { + } else { + reads_vsearch5_ch + .into{ nuclFastas_forDiamond_ch; nuclFastas_forCounts_ch; nuclFastas_forphylogeny_asv; nuclFastas_forMatrix_ch } + } - process Generate_Nucleotide_Matrix { + process ASV_Taxonomy_Inference { - label 'low_cpus' + label 'high_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Matrix", mode: "copy", overwrite: true, pattern: '*_ASV*PercentID.matrix' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Matrix", mode: "copy", overwrite: true, pattern: '*ncASV*PercentID.matrix' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy", mode: "copy", overwrite: true, pattern: '*_ASV*.{fasta,csv,tsv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*_ASV*dmd.out' - input: - tuple file(notus), file(asvs) from nuclFastas_forMatrix_ch + input: + file(reads) from nuclFastas_forDiamond_asv_ch - output: - file("*.matrix") into clustmatrices - file("*ncASV*PercentID.matrix") into notu_heatmap - file("*_ASV*PercentID.matrix") into asv_heatmap + output: + file("*.fasta") into tax_labeled_fasta2 + tuple file("*_phyloseqObject.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond_asv + file("*_ASV*_summary_for_plot.csv") into taxplot_asv - script: - // remove if statement later (no fin) - """ - for filename in ${notus};do - if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then - ident=\$( echo \${filename} | awk -F "ncASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') - name=\$( echo \${filename}| awk -F ".fasta" '{print \$1}') - clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} - clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} - for x in *q.matrix;do - pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') - ya=\$(wc -l \$x | awk '{print \$1}') - echo "\$((\$ya-1))" - tail -"\$((\$ya-1))" \$x > \${pre}z.matrix - rm \$x - cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix - rm \${pre}z.matrix - done + script: + """ + cp ${params.vampdir}/bin/rename_seq.py . + virdb=${params.dbdir}/${params.dbname} + grep ">" \${virdb} > headers.list + headers="headers.list" + for filename in ${reads};do + name=\$(ls \${filename} | awk -F ".fasta" '{print \$1}') + diamond blastx -q \${filename} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + echo "Preparing lists to generate summary .csv's" + echo "[Best hit accession number]" > access.list + echo "[e-value]" > evalue.list + echo "[Bitscore]" > bit.list + echo "[Percent ID (aa)]" > pid.list + echo "[Organism ID]" > "\$name"_virus.list + echo "[Gene]" > "\$name"_genes.list + grep ">" \${filename} | awk -F ">" '{print \$2}' > seqids.lst + echo "extracting genes and names" + touch new_"\$name"_asvnames.txt + j=1 + if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then + echo "[ASV#]" > otu.list + echo "[ASV sequence length]" > length.list + for s in \$(cat seqids.lst);do + echo "Checking for \$s hit in diamond output" + if [[ ${params.refseq} == "T" ]];then + echo "RefSeq headers specified" + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list + virus="NO" + gene="HIT" + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + fi + else + echo "Using RVDB headers." + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list + virus="NO" + gene="HIT" + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." fi + fi + echo "Done with \$s" done - if [ `echo ${asvs} | grep -c "_ASV"` -eq 1 ];then - name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - clustalo -i ${asvs} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} - clustalo -i ${asvs} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} - for x in *q.matrix;do - pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') - ya=\$(wc -l \$x | awk '{print \$1}') - echo "\$((\$ya-1))" - tail -"\$((\$ya-1))" \$x > \${pre}z.matrix - rm \$x - cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix - rm \${pre}z.matrix - done + else + for s in \$(cat seqids.lst);do + echo "[ASV#]" > otu.list + echo "[ASV sequence length]" > length.list + echo "Checking for \$s hit in diamond output" + if [[ ${params.refseq} == "T" ]];then + echo "RefSeq headers specified" + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list + virus="NO" + gene="HIT" + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + fi + else + echo "Using RVDB headers." + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list + virus="NO" + gene="HIT" + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + fi fi - """ - } - } else { + echo "Done with \$s" + done + fi + echo "Now editing "\$name" fasta headers" + ###### rename_seq.py + ./rename_seq.py \${filename} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta + echo "[Sequence header]" > newnames.list + cat new_"\$name"_asvnames.txt >> newnames.list + touch sequence.list + echo " " > sequence.list + grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list + rm "\$name"_tmpssasv.fasta + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + for x in *phyloseqObject.csv;do + echo "\$x" + lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) + tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; + sed 's/ /_/g' tmpcol.list > tmp2col.list; + cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; + rm tmpcol.list tmp2col.list + done + rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; + rm *asvnames.txt + rm "\$name"_virus.list + rm "\$name"_genes.list + rm newnames.list + rm access.list + echo "Taxonomy inferred for: \${filename} " + done + rm headers.list + """ + } - process Generate_ASV_Matrix { - - label 'low_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Matrix", mode: "copy", overwrite: true, pattern: '*ASV*PercentID.matrix' - - input: - file(reads) from nuclFastas_forMatrix_ch - - output: - file("*.matrix") into clustmatrices - file("*_ASV*PercentID.matrix") into asv_heatmap - - script: - // remove if statement later (no fin) - """ - for filename in ${reads};do - if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then - ident=\$( echo \${filename} | awk -F "ncASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') - name=\$( echo \${filename}| awk -F ".fasta" '{print \$1}') - clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} - clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} - for x in *q.matrix;do - pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') - ya=\$(wc -l \$x | awk '{print \$1}') - echo "\$((\$ya-1))" - tail -"\$((\$ya-1))" \$x > \${pre}z.matrix - rm \$x - cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix - rm \${pre}z.matrix - done - else - name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') - clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} - clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} - for x in *q.matrix;do - pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') - ya=\$(wc -l \$x | awk '{print \$1}') - echo "\$((\$ya-1))" - tail -"\$((\$ya-1))" \$x > \${pre}z.matrix - rm \$x - cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix - rm \${pre}z.matrix - done - fi - done - """ - } - } + process Generate_ASV_Counts_Tables { - if (!params.skipPhylogeny) { // need to edit paths - - nuclFastas_forphylogeny_asv = Channel.create() - nuclFastas_forphylogeny_ncasv = Channel.create() - nuclFastas_forphylogeny.into(nuclFastas_forphylogeny_asv, nuclFastas_forphylogeny_ncasv) - - process ASV_Phylogeny { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*ASV*aln.*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*ASV*mt*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*ASV*iq*' - - input: - file(asvs) from nuclFastas_forphylogeny_asv - - output: - tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into align_results - file("*iq.treefile") into nucl_phyl_plot - - script: - """ - pre=\$(echo \${filename} | awk -F ".fasta" '{print \$1}' ) - mafft --thread ${task.cpus} --maxiterate 15000 --auto \${filename} >\${pre}_ALN.fasta - trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html - # Nucleotide_ModelTest - modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint - # Nucleotide_Phylogeny - if [ "${params.iqCustomnt}" != "" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomnt} - elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} - elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni - elif [ "${params.nonparametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} - elif [ "${params.parametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - else - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - fi - done - """ - } - - if (params.ncASV) { - - process ncASV_Phylogeny { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*ncASV*aln.*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*ncASV*mt*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*ncASV*iq*' - - input: - tuple file(notus), file(asvs) from nuclFastas_forphylogeny_ncasv - - output: - tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into align_results_ncasv - file("*iq.treefile") into nucl_phyl_plot_ncasv - - script: - """ - for filename in ${notus};do - pre=\$(echo \${filename} | awk -F ".fasta" '{print \$1}' ) - mafft --thread ${task.cpus} --maxiterate 15000 --auto \${filename} >\${pre}_ALN.fasta - trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html - # Nucleotide_ModelTest - modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint - # Nucleotide_Phylogeny - if [ "${params.iqCustomnt}" != "" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -t \${pre}_mt.tree -T auto ${params.iqCustomnt} - elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} - elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni - elif [ "${params.nonparametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} - elif [ "${params.parametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni - else - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni - fi - done - """ - } + label 'norm_cpus' - } + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Counts", mode: "copy", overwrite: true, pattern: '*ASV*.{biome,csv}' + + input: + file(asvs) from nuclFastas_forCounts_asv_ch + file(merged) from nuclCounts_mergedreads_ch + + output: + tuple file("*_counts.csv"), file("*_counts.biome") into counts_vsearch_asv + file("*_ASV*counts.csv") into asv_counts_plots_asv + + script: + """ + if [ `echo ${asvs} | grep -c "ASV"` -eq 1 ];then + name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') + vsearch --usearch_global ${merged} --db ${asvs} --id ${params.asvcountID} --threads ${task.cpus} --otutabout "\$name"_counts.txt --biomout "\$name"_counts.biome + cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv + sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv + rm \${name}_count.csv + fi + """ + } + + process Generate_ASV_Matrices { + + label 'low_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Matrix", mode: "copy", overwrite: true, pattern: '*ASV*PercentID.matrix' + input: + file(reads) from nuclFastas_forMatrix_asv_ch + + output: + file("*.matrix") into clustmatrices_asv + file("*_ASV*PercentID.matrix") into asv_heatmap_asv + + script: + // remove if statement later (no fin) + """ + for filename in ${reads};do + if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then + ident=\$( echo \${filename} | awk -F "ncASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') + name=\$( echo \${filename}| awk -F ".fasta" '{print \$1}') + clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} + clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} + for x in *q.matrix;do + pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') + ya=\$(wc -l \$x | awk '{print \$1}') + echo "\$((\$ya-1))" + tail -"\$((\$ya-1))" \$x > \${pre}z.matrix + rm \$x + cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix + rm \${pre}z.matrix + done + else + name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') + clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} + clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} + for x in *q.matrix;do + pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') + ya=\$(wc -l \$x | awk '{print \$1}') + echo "\$((\$ya-1))" + tail -"\$((\$ya-1))" \$x > \${pre}z.matrix + rm \$x + cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix + rm \${pre}z.matrix + done + fi + done + """ + } + + if (!params.skipPhylogeny) { // need to edit paths + + process ASV_Phylogeny { + + label 'norm_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*ASV*aln.*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*ASV*mt*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*ASV*iq*' + + input: + file(asvs) from nuclFastas_forphylogeny_asv + + output: + tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into align_results_asv + file("*iq.treefile") into nucl_phyl_plot_asv + + script: + """ + pre=\$(echo ${asvs} | awk -F ".fasta" '{print \$1}' ) + mafft --thread ${task.cpus} --maxiterate 15000 --auto ${asvs} >\${pre}_ALN.fasta + trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html + # Nucleotide_ModelTest + modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint + # Nucleotide_Phylogeny + if [ "${params.iqCustomnt}" != "" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomnt} + elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} + elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni + elif [ "${params.nonparametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} + elif [ "${params.parametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + else + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + fi + """ + } } if (!params.skipAminoTyping) { @@ -2035,7 +1862,7 @@ if (params.DataCheck || params.Analyze) { """ } - process Generate_AminoType_Matrix { + process Generate_AminoType_Matrices { label 'low_cpus' From 449669eebc98eb7bd35f94552d04b1b22f90dd56 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Wed, 10 Mar 2021 16:20:09 -0800 Subject: [PATCH 023/223] Remove . --- vampirus.config | 6 +++--- 1 file changed, 3 insertions(+), 3 deletions(-) diff --git a/vampirus.config b/vampirus.config index fbd73e3..aff8d25 100644 --- a/vampirus.config +++ b/vampirus.config @@ -58,11 +58,11 @@ params { // Minimum size or representation for sequence to be considered in ASV generation minSize="8" // Percent similarity to cluster nucleotide ASV sequences - clusterNuclID=".85" + clusterNuclID="85" // List of percent similarities to cluster nucleotide ASV sequences - must be separated by a comma (ex. ".95,.96") clusterNuclIDlist="" // Default percent similarity to cluster aminoacid sequences - clusterAAID=".97" + clusterAAID="97" // List of percent similarities to cluster aminoacid sequences - must be separated by ".95,.96" clusterAAIDlist="" // Minimum length of amino acid translation to be considered during protein clustered ASV (pcASV) generation. Recommended to put this at the expected aminoacid sequence length based on your maximum read length (e.g. if maxLen="420", then minAA should be 420/3 or 140) @@ -71,7 +71,7 @@ params { // Counts table generation parameters // Percent similarity to use for ASV/cASV counts table generation with vsearch - asvcountID=".97" + asvcountID="97" // Parameters for protein counts table generation // Minimum Bitscore for counts ProtCountsBit="50" From 1e858a7be875e8013b401d908be39e59cae1e6b3 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Wed, 10 Mar 2021 16:20:19 -0800 Subject: [PATCH 024/223] Add percentage --- vAMPirus.nf | 24 ++++++++++++------------ 1 file changed, 12 insertions(+), 12 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 38124f7..f3793e7 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -472,7 +472,7 @@ if (params.readsTest) { } // HERE fix if stament and list names. Also is not an integer -if (params.clusterNuclIDlist == "") { +if (!params.clusterNuclIDlist == "") { msize=params.clusterNuclIDlist def slist=msize.split(',').collect{it as int} def nnuc=slist.size() @@ -1151,11 +1151,11 @@ if (params.DataCheck || params.Analyze) { if (params.clusterNuclIDlist) { nid=slist1.get(x-1) """ - vsearch --cluster_fast ${fasta} --centroids ${params.projtag}_ncASV\${id}.fasta --threads ${task.cpus} --relabel ncASV --id ${nid} + vsearch --cluster_fast ${fasta} --centroids ${params.projtag}_ncASV\${id}.fasta --threads ${task.cpus} --relabel ncASV --id .${nid} """ } else if (params.clusterNuclID) { """ - vsearch --cluster_fast ${fasta} --centroids ${params.projtag}_ncASV\${id}.fasta --threads ${task.cpus} --relabel ncASV --id ${params.clusterNuclID} + vsearch --cluster_fast ${fasta} --centroids ${params.projtag}_ncASV\${id}.fasta --threads ${task.cpus} --relabel ncASV --id .${params.clusterNuclID} """ } } @@ -1173,7 +1173,7 @@ if (params.DataCheck || params.Analyze) { file(asvs) from nuclFastas_forDiamond_ncasv_ch output: - file("*.fasta") into tax_labeled_fasta1 + file("*.fasta") into tax_labeled_fasta_ncasv tuple file("*_phyloseqObject.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond_ncasv file("*ncASV*summary_for_plot.csv") into taxplot_ncasv @@ -1327,9 +1327,9 @@ if (params.DataCheck || params.Analyze) { file(merged) from nuclCounts_mergedreads_ch output: - tuple file("*_counts.csv"), file("*_counts.biome") into counts_vsearch + tuple file("*_counts.csv"), file("*_counts.biome") into counts_vsearch_ncasv tuple val("${id}"), file("*ncASV*counts.csv") into notu_counts_plots - file("*_ASV*counts.csv") into asv_counts_plots + file("*_ASV*counts.csv") into ncasv_counts_plots script: """ @@ -1430,7 +1430,7 @@ if (params.DataCheck || params.Analyze) { } else { reads_vsearch5_ch - .into{ nuclFastas_forDiamond_ch; nuclFastas_forCounts_ch; nuclFastas_forphylogeny_asv; nuclFastas_forMatrix_ch } + .into{ nuclFastas_forDiamond_asv_ch; nuclFastas_forCounts_asv_ch; nuclFastas_forphylogeny_asv; nuclFastas_forMatrix_asv_ch } } process ASV_Taxonomy_Inference { @@ -1444,7 +1444,7 @@ if (params.DataCheck || params.Analyze) { file(reads) from nuclFastas_forDiamond_asv_ch output: - file("*.fasta") into tax_labeled_fasta2 + file("*.fasta") into tax_labeled_fasta_asv tuple file("*_phyloseqObject.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond_asv file("*_ASV*_summary_for_plot.csv") into taxplot_asv @@ -1687,7 +1687,7 @@ if (params.DataCheck || params.Analyze) { """ if [ `echo ${asvs} | grep -c "ASV"` -eq 1 ];then name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - vsearch --usearch_global ${merged} --db ${asvs} --id ${params.asvcountID} --threads ${task.cpus} --otutabout "\$name"_counts.txt --biomout "\$name"_counts.biome + vsearch --usearch_global ${merged} --db ${asvs} --id .${params.asvcountID} --threads ${task.cpus} --otutabout "\$name"_counts.txt --biomout "\$name"_counts.biome cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv rm \${name}_count.csv @@ -1699,7 +1699,7 @@ if (params.DataCheck || params.Analyze) { label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Matrix", mode: "copy", overwrite: true, pattern: '*ASV*PercentID.matrix' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Matrices", mode: "copy", overwrite: true, pattern: '*ASV*PercentID.matrix' input: file(reads) from nuclFastas_forMatrix_asv_ch @@ -1866,7 +1866,7 @@ if (params.DataCheck || params.Analyze) { label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Matrix", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Matrices", mode: "copy", overwrite: true input: file(prot) from aminotypesClustal @@ -1967,7 +1967,7 @@ if (params.DataCheck || params.Analyze) { diamond blastp -q ${reads} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" >access.list - echo "[pcASV sequence length]" >length.list + echo "[AminoType sequence length]" >length.list echo "[e-value]" >evalue.list echo "[Bitscore]" >bit.list echo "[Percent ID (aa)]" >pid.list From 24f645b3478800e783f1273f2fb2bfdaf9ef22a9 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Wed, 10 Mar 2021 17:24:22 -0800 Subject: [PATCH 025/223] Remove . --- example_data/conf/test.config | 6 +++--- 1 file changed, 3 insertions(+), 3 deletions(-) diff --git a/example_data/conf/test.config b/example_data/conf/test.config index 774f905..d213a47 100644 --- a/example_data/conf/test.config +++ b/example_data/conf/test.config @@ -15,9 +15,9 @@ params { outdir="vAMPtestresults" workingdir="${projectDir}/example_data/" projtag="vAMPtest" - clusterNuclID=".85" - clusterAAID=".85" + clusterNuclID="85" + clusterAAID="85" dbname="test_db.fasta" dbdir="${projectDir}/example_data/conf/" - stats="run" + stats = true } From e380799a3d58691b5581f66533a252123d5f24d7 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Wed, 10 Mar 2021 17:33:26 -0800 Subject: [PATCH 026/223] Add name --- example_data/conf/test.config | 3 +-- 1 file changed, 1 insertion(+), 2 deletions(-) diff --git a/example_data/conf/test.config b/example_data/conf/test.config index d213a47..b68646b 100644 --- a/example_data/conf/test.config +++ b/example_data/conf/test.config @@ -3,8 +3,7 @@ Test Config File vAMPirus ======================================================================================== Virus Amplicon Analysis Pipeline - Author: Alex J.Veglia - Version: 1.0 (dev) + Author: Alex J.Veglia, Ramón E. Rivera Vicéns ---------------------------------------------------------------------------------------- */ From b6f1ddced4672b6df021cfe857d1e8d8af6568a4 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Wed, 10 Mar 2021 17:47:08 -0800 Subject: [PATCH 027/223] Update id list --- vampirus.config | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/vampirus.config b/vampirus.config index aff8d25..dae7c9d 100644 --- a/vampirus.config +++ b/vampirus.config @@ -59,11 +59,11 @@ params { minSize="8" // Percent similarity to cluster nucleotide ASV sequences clusterNuclID="85" - // List of percent similarities to cluster nucleotide ASV sequences - must be separated by a comma (ex. ".95,.96") + // List of percent similarities to cluster nucleotide ASV sequences - must be separated by a comma (ex. "95,96") clusterNuclIDlist="" // Default percent similarity to cluster aminoacid sequences clusterAAID="97" - // List of percent similarities to cluster aminoacid sequences - must be separated by ".95,.96" + // List of percent similarities to cluster aminoacid sequences - must be separated by "95,96" clusterAAIDlist="" // Minimum length of amino acid translation to be considered during protein clustered ASV (pcASV) generation. Recommended to put this at the expected aminoacid sequence length based on your maximum read length (e.g. if maxLen="420", then minAA should be 420/3 or 140) minAA="140" From 21507faf40b08618c44801100831f7622d2e8928 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Wed, 10 Mar 2021 17:50:13 -0800 Subject: [PATCH 028/223] WIP - Add if for nnuc --- vAMPirus.nf | 10 ++++++---- 1 file changed, 6 insertions(+), 4 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index f3793e7..e14f64c 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -472,15 +472,17 @@ if (params.readsTest) { } // HERE fix if stament and list names. Also is not an integer -if (!params.clusterNuclIDlist == "") { +if (params.clusterNuclIDlist == "") { + def nnuc=1 +} else { msize=params.clusterNuclIDlist def slist=msize.split(',').collect{it as int} def nnuc=slist.size() -} else { - def nnuc=1 } if (params.clusterAAIDlist == "") { + def naa=1 +} else { msize2=params.clusterAAIDlist def slist2=msize2.split(',').collect{it as int} def naa=slist2.size() @@ -1155,7 +1157,7 @@ if (params.DataCheck || params.Analyze) { """ } else if (params.clusterNuclID) { """ - vsearch --cluster_fast ${fasta} --centroids ${params.projtag}_ncASV\${id}.fasta --threads ${task.cpus} --relabel ncASV --id .${params.clusterNuclID} + vsearch --cluster_fast ${fasta} --centroids ${params.projtag}_ncASV${params.clusterNuclID}.fasta --threads ${task.cpus} --relabel ncASV --id .${params.clusterNuclID} """ } } From 22aecd74f4473562875cf66a921fe795edd4db04 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Wed, 10 Mar 2021 18:08:02 -0800 Subject: [PATCH 029/223] WIP - remove def --- vAMPirus.nf | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index e14f64c..cd996c7 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -473,7 +473,7 @@ if (params.readsTest) { // HERE fix if stament and list names. Also is not an integer if (params.clusterNuclIDlist == "") { - def nnuc=1 + nnuc=1 } else { msize=params.clusterNuclIDlist def slist=msize.split(',').collect{it as int} @@ -481,7 +481,7 @@ if (params.clusterNuclIDlist == "") { } if (params.clusterAAIDlist == "") { - def naa=1 + naa=1 } else { msize2=params.clusterAAIDlist def slist2=msize2.split(',').collect{it as int} From 0cceba5c05a59d4bf2c6e529e917b5b46a7d3b8c Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Wed, 10 Mar 2021 18:56:32 -0800 Subject: [PATCH 030/223] Add id tag. Fix list --- vAMPirus.nf | 23 +++++++++++++++-------- 1 file changed, 15 insertions(+), 8 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index cd996c7..e42f513 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -473,19 +473,23 @@ if (params.readsTest) { // HERE fix if stament and list names. Also is not an integer if (params.clusterNuclIDlist == "") { - nnuc=1 + a=params.clusterNuclID + slist=[a] + nnuc=slist.size() } else { msize=params.clusterNuclIDlist - def slist=msize.split(',').collect{it as int} - def nnuc=slist.size() + slist=msize.split(',').collect{it as int} + nnuc=slist.size() } if (params.clusterAAIDlist == "") { - naa=1 + b=params.clusterAAID + slist2=[b] + naa=slist2.size() } else { msize2=params.clusterAAIDlist def slist2=msize2.split(',').collect{it as int} - def naa=slist2.size() + naa=slist2.size() } if (params.DataCheck || params.Analyze) { @@ -1140,6 +1144,8 @@ if (params.DataCheck || params.Analyze) { label 'norm_cpus' + tag "${mtag}" + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/ncASV", mode: "copy", overwrite: true, pattern: '*ncASV*.fasta' input: @@ -1150,10 +1156,11 @@ if (params.DataCheck || params.Analyze) { file("*_ncASV*.fasta") into ( nuclFastas_forphylogeny_ncasv, nuclFastas_forDiamond_ncasv_ch, nuclFastas_forCounts_ncasv_ch, nuclFastas_forMatrix_ncasv_ch ) script: + nid=slist.get(x-1) + mtag="ID=" + slist.get(x-1) if (params.clusterNuclIDlist) { - nid=slist1.get(x-1) """ - vsearch --cluster_fast ${fasta} --centroids ${params.projtag}_ncASV\${id}.fasta --threads ${task.cpus} --relabel ncASV --id .${nid} + vsearch --cluster_fast ${fasta} --centroids ${params.projtag}_ncASV${nid}.fasta --threads ${task.cpus} --relabel ncASV --id .${nid} """ } else if (params.clusterNuclID) { """ @@ -1339,7 +1346,7 @@ if (params.DataCheck || params.Analyze) { if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then ident=\$( echo \${filename} | awk -F "ncASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') - vsearch --usearch_global ${merged} --db \${filename} --id \${ident} --threads ${task.cpus} --otutabout \${name}_counts.txt --biomout \${name}_counts.biome + vsearch --usearch_global ${merged} --db \${filename} --id .\${ident} --threads ${task.cpus} --otutabout \${name}_counts.txt --biomout \${name}_counts.biome cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv rm \${name}_count.csv From 129305a1b103cb504a6b87694c749b11367fa637 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Wed, 10 Mar 2021 19:40:42 -0800 Subject: [PATCH 031/223] Add id tag. Fix missing channels --- vAMPirus.nf | 353 ++++++++++++++++++++++++---------------------------- 1 file changed, 161 insertions(+), 192 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index e42f513..a44cb7d 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -761,35 +761,6 @@ if (params.DataCheck || params.Analyze) { } - /* BEfore examples - process Length_Filtering { //changed - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/LengthFiltering", mode: "copy", overwrite: true, pattern: "*_merged_preFilt*.fasta" - publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging", mode: "copy", overwrite: true, pattern: "*Lengthfiltered.fastq" - publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/Histograms/pre_length_filtering", mode: "copy", overwrite: true, pattern: "*preFilt_*st.txt" - publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/Histograms/post_length_filtering", mode: "copy", overwrite: true, pattern: "*postFilt_*st.txt" - - input: - file(reads) from collect_samples_ch - - output: - file("*_merged_preFilt_clean.fasta") into ( nuclCounts_mergedreads_ch, pcASV_mergedreads_ch ) - file("*_merged_clean_Lengthfiltered.fastq") into reads_vsearch2_ch - file("*_merged_preFilt_clean.fastq") into ( mergeforprotcounts, mergeforpcASVaacounts ) - file("**hist.txt") into histos - - script: - """ - bbduk.sh in=${reads} bhist=${params.projtag}_all_merged_preFilt_preClean_baseFrequency_hist.txt qhist=${params.projtag}_all_merged_preFilt_preClean_qualityScore_hist.txt gchist=${params.projtag}_all_merged_preFilt_preClean_gcContent_hist.txt aqhist=${params.projtag}_all_merged_preFilt_preClean_averageQuality_hist.txt lhist=${params.projtag}_all_merged_preFilt__preClean_length_hist.txt gcbins=auto - fastp -i ${reads} -o ${params.projtag}_merged_preFilt_clean.fastq -b ${params.maxLen} -l ${params.minLen} --thread ${task.cpus} -n 1 - reformat.sh in=${params.projtag}_merged_preFilt_clean.fastq out=${params.projtag}_merged_preFilt_clean.fasta t=${task.cpus} - bbduk.sh in=${params.projtag}_merged_preFilt_clean.fastq out=${params.projtag}_merged_clean_Lengthfiltered.fastq minlength=${params.maxLen} maxlength=${params.maxLen} t=${task.cpus} - bbduk.sh in=${params.projtag}_merged_clean_Lengthfiltered.fastq bhist=${params.projtag}_all_merged_postFilt_baseFrequency_hist.txt qhist=${params.projtag}_all_merged_postFilt_qualityScore_hist.txt gchist=${params.projtag}_all_merged_postFilt_gcContent_hist.txt aqhist=${params.projtag}_all_merged_postFilt_averageQuaulity_hist.txt lhist=${params.projtag}_all_merged_postFilt_length_hist.txt gcbins=auto - """ - }*/ - process Length_Filtering { label 'norm_cpus' @@ -805,7 +776,7 @@ if (params.DataCheck || params.Analyze) { output: file("*_merged_preFilt_clean.fastq") into ( mergeforprotcounts, mergeforpcASVaacounts ) - file("*_merged_preFilt_clean.fasta") into ( nuclCounts_mergedreads_ch, pcASV_mergedreads_ch ) + file("*_merged_preFilt_clean.fasta") into ( nuclCounts_mergedreads_asv_ch, nuclCounts_mergedreads_ncasv_ch, pcASV_mergedreads_ch ) file("*_merged_clean_Lengthfiltered.fastq") into reads_vsearch2_ch file("*preFilt_preClean_baseFrequency_hist.csv") into prefilt_basefreq @@ -1153,7 +1124,7 @@ if (params.DataCheck || params.Analyze) { file(fasta) from asv_file_for_ncasvs output: - file("*_ncASV*.fasta") into ( nuclFastas_forphylogeny_ncasv, nuclFastas_forDiamond_ncasv_ch, nuclFastas_forCounts_ncasv_ch, nuclFastas_forMatrix_ncasv_ch ) + tuple nid, file("*_ncASV*.fasta") into ( nuclFastas_forphylogeny_ncasv, nuclFastas_forDiamond_ncasv_ch, nuclFastas_forCounts_ncasv_ch, nuclFastas_forMatrix_ncasv_ch ) script: nid=slist.get(x-1) @@ -1170,16 +1141,18 @@ if (params.DataCheck || params.Analyze) { } if (!params.skipTaxonomy) { - + // here nid for output?? process Nucleotide_Taxonomy_Inference { label 'high_cpus' + tag "${mtag}" + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Taxonomy", mode: "copy", overwrite: true, pattern: '*ncASV*.{fasta,csv,tsv}' publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*ncASV*dmd.out' input: - file(asvs) from nuclFastas_forDiamond_ncasv_ch + tuple nid, file(asvs) from nuclFastas_forDiamond_ncasv_ch output: file("*.fasta") into tax_labeled_fasta_ncasv @@ -1187,138 +1160,137 @@ if (params.DataCheck || params.Analyze) { file("*ncASV*summary_for_plot.csv") into taxplot_ncasv script: + mtag="ID=" + nid """ cp ${params.vampdir}/bin/rename_seq.py . virdb=${params.dbdir}/${params.dbname} grep ">" \${virdb} > headers.list headers="headers.list" - for filename in ${asvs};do - name=\$(ls \${filename} | awk -F ".fasta" '{print \$1}') - diamond blastx -q \${filename} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 - echo "Preparing lists to generate summary .csv's" - echo "[Best hit accession number]" > access.list - echo "[e-value]" > evalue.list - echo "[Bitscore]" > bit.list - echo "[Percent ID (aa)]" > pid.list - echo "[Organism ID]" > "\$name"_virus.list - echo "[Gene]" > "\$name"_genes.list - grep ">" \${filename} | awk -F ">" '{print \$2}' > seqids.lst - echo "extracting genes and names" - touch new_"\$name"_asvnames.txt - j=1 - if [ `echo \${filename} | grep -c "ASV"` -eq 1 ];then - for s in \$(cat seqids.lst);do - echo "[ASV#]" > otu.list - echo "[ASV sequence length]" > length.list - echo "Checking for \$s hit in diamond output" - if [[ ${params.refseq} == "T" ]];then - echo "RefSeq headers specified" - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list - virus="NO" - gene="HIT" - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - fi + name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') + diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + echo "Preparing lists to generate summary .csv's" + echo "[Best hit accession number]" > access.list + echo "[e-value]" > evalue.list + echo "[Bitscore]" > bit.list + echo "[Percent ID (aa)]" > pid.list + echo "[Organism ID]" > "\$name"_virus.list + echo "[Gene]" > "\$name"_genes.list + grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst + echo "extracting genes and names" + touch new_"\$name"_asvnames.txt + j=1 + if [ `echo ${asvs} | grep -c "ASV"` -eq 1 ];then + for s in \$(cat seqids.lst);do + echo "[ASV#]" > otu.list + echo "[ASV sequence length]" > length.list + echo "Checking for \$s hit in diamond output" + if [[ ${params.refseq} == "T" ]];then + echo "RefSeq headers specified" + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." else - echo "Using RVDB headers." - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list - virus="NO" - gene="HIT" - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - fi + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list + virus="NO" + gene="HIT" + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." fi - echo "Done with \$s" - done - fi - echo "Now editing "\$name" fasta headers" - ###### rename_seq.py - ./rename_seq.py \${filename} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta - awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta - echo "[Sequence header]" > newnames.list - cat new_"\$name"_asvnames.txt >> newnames.list - touch sequence.list - echo " " > sequence.list - grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list - rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - for x in *phyloseqObject.csv;do - echo "\$x" - lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) - tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; - sed 's/ /_/g' tmpcol.list > tmp2col.list; - cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; - rm tmpcol.list tmp2col.list + else + echo "Using RVDB headers." + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list + virus="NO" + gene="HIT" + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + fi + fi + echo "Done with \$s" done - rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; - rm *asvnames.txt - rm "\$name"_virus.list - rm "\$name"_genes.list - rm newnames.list - rm access.list - echo "Taxonomy inferred for: \${filename} " + fi + echo "Now editing "\$name" fasta headers" + ###### rename_seq.py + ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta + echo "[Sequence header]" > newnames.list + cat new_"\$name"_asvnames.txt >> newnames.list + touch sequence.list + echo " " > sequence.list + grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list + rm "\$name"_tmpssasv.fasta + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + for x in *phyloseqObject.csv;do + echo "\$x" + lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) + tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; + sed 's/ /_/g' tmpcol.list > tmp2col.list; + cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; + rm tmpcol.list tmp2col.list done + rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; + rm *asvnames.txt + rm "\$name"_virus.list + rm "\$name"_genes.list + rm newnames.list + rm access.list + echo "Taxonomy inferred for: ${asvs}" rm headers.list """ } @@ -1328,66 +1300,60 @@ if (params.DataCheck || params.Analyze) { label 'norm_cpus' + tag "${mtag}" + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Counts", mode: "copy", overwrite: true, pattern: '*_ASV*.{biome,csv}' publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Counts", mode: "copy", overwrite: true, pattern: '*ncASV*.{biome,csv}' input: - file(notus) from nuclFastas_forCounts_ncasv_ch - file(merged) from nuclCounts_mergedreads_ch + tuple nid, file(notus) from nuclFastas_forCounts_ncasv_ch + file(merged) from nuclCounts_mergedreads_ncasv_ch output: tuple file("*_counts.csv"), file("*_counts.biome") into counts_vsearch_ncasv tuple val("${id}"), file("*ncASV*counts.csv") into notu_counts_plots - file("*_ASV*counts.csv") into ncasv_counts_plots + file("*ASV*counts.csv") into ncasv_counts_plots script: + mtag="ID=" + nid """ - for filename in ${notus};do - if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then - ident=\$( echo \${filename} | awk -F "ncASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') - name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') - vsearch --usearch_global ${merged} --db \${filename} --id .\${ident} --threads ${task.cpus} --otutabout \${name}_counts.txt --biomout \${name}_counts.biome - cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv - sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv - rm \${name}_count.csv - fi - done + name=\$( echo ${notus} | awk -F ".fasta" '{print \$1}') + vsearch --usearch_global ${merged} --db ${notus} --id .${nid} --threads ${task.cpus} --otutabout \${name}_counts.txt --biomout \${name}_counts.biome + cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv + sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv + rm \${name}_count.csv """ } process Generate_ncASV_Matrices { - + //here do I need the for loop label 'low_cpus' + tag "${mtag}" + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Matrices", mode: "copy", overwrite: true, pattern: '*ncASV*PercentID.matrix' input: - - file(notus) from nuclFastas_forMatrix_ncasv_ch + tuple nid, file(asvs) from nuclFastas_forMatrix_ncasv_ch output: file("*.matrix") into clustmatrices_ncasv file("*ncASV*PercentID.matrix") into notu_heatmap script: - // remove if statement later (no fin) + mtag="ID=" + nid """ - for filename in ${notus};do - if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then - ident=\$( echo \${filename} | awk -F "ncASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') - name=\$( echo \${filename}| awk -F ".fasta" '{print \$1}') - clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} - clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} - for x in *q.matrix;do - pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') - ya=\$(wc -l \$x | awk '{print \$1}') - echo "\$((\$ya-1))" - tail -"\$((\$ya-1))" \$x > \${pre}z.matrix - rm \$x - cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix - rm \${pre}z.matrix - done - fi + name=\$( echo ${asvs}| awk -F ".fasta" '{print \$1}') + clustalo -i ${asvs} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} + clustalo -i ${asvs} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} + for x in *q.matrix;do + pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') + ya=\$(wc -l \$x | awk '{print \$1}') + echo "\$((\$ya-1))" + tail -"\$((\$ya-1))" \$x > \${pre}z.matrix + rm \$x + cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix + rm \${pre}z.matrix done """ } @@ -1398,18 +1364,21 @@ if (params.DataCheck || params.Analyze) { label 'norm_cpus' + tag "${mtag}" + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*ncASV*aln.*' publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*ncASV*mt*' publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*ncASV*iq*' input: - file(asvs) from nuclFastas_forphylogeny_ncasv + tuple nid, file(asvs) from nuclFastas_forphylogeny_ncasv output: - tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into align_results_ncasv + tuple nid, file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into align_results_ncasv file("*iq.treefile") into nucl_phyl_plot_ncasv script: + mtag="ID=" + nid """ pre=\$(echo ${asvs} | awk -F ".fasta" '{print \$1}' ) mafft --thread ${task.cpus} --maxiterate 15000 --auto ${asvs} >\${pre}_ALN.fasta @@ -1686,7 +1655,7 @@ if (params.DataCheck || params.Analyze) { input: file(asvs) from nuclFastas_forCounts_asv_ch - file(merged) from nuclCounts_mergedreads_ch + file(merged) from nuclCounts_mergedreads_asv_ch output: tuple file("*_counts.csv"), file("*_counts.biome") into counts_vsearch_asv From 0ed4e8d73e306a3d97037f32160970747596a08b Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Wed, 10 Mar 2021 19:58:30 -0800 Subject: [PATCH 032/223] Fix Generate_ncASV_Matrices process --- vAMPirus.nf | 16 +++------------- 1 file changed, 3 insertions(+), 13 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index a44cb7d..8f3f63e 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -445,7 +445,6 @@ if (params.help) { exit 0 } -// This will be printed to the user in each run. here thy can check if the values the selected are fine log.info """\ ================================================================================================================================================ vAMPirus v${workflow.manifest.version} - Virus Amplicon Sequencing Analysis Pipeline @@ -471,7 +470,6 @@ if (params.readsTest) { .into{ reads_ch; reads_qc_ch; reads_processing } } -// HERE fix if stament and list names. Also is not an integer if (params.clusterNuclIDlist == "") { a=params.clusterNuclID slist=[a] @@ -1141,7 +1139,7 @@ if (params.DataCheck || params.Analyze) { } if (!params.skipTaxonomy) { - // here nid for output?? + process Nucleotide_Taxonomy_Inference { label 'high_cpus' @@ -1326,7 +1324,6 @@ if (params.DataCheck || params.Analyze) { } process Generate_ncASV_Matrices { - //here do I need the for loop label 'low_cpus' tag "${mtag}" @@ -1346,15 +1343,8 @@ if (params.DataCheck || params.Analyze) { name=\$( echo ${asvs}| awk -F ".fasta" '{print \$1}') clustalo -i ${asvs} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} clustalo -i ${asvs} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} - for x in *q.matrix;do - pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') - ya=\$(wc -l \$x | awk '{print \$1}') - echo "\$((\$ya-1))" - tail -"\$((\$ya-1))" \$x > \${pre}z.matrix - rm \$x - cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix - rm \${pre}z.matrix - done + cat \${name}_PercentIDq.matrix | tr " " "," | grep "," >\${name}_PercentIDq.matrix + rm \${name}_PercentIDq.matrix """ } From e9a23a2168363cfa0267994fd4d92daa359b9cd6 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Wed, 10 Mar 2021 20:08:23 -0800 Subject: [PATCH 033/223] Remove extra letter --- vAMPirus.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 8f3f63e..8fab14a 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1343,7 +1343,7 @@ if (params.DataCheck || params.Analyze) { name=\$( echo ${asvs}| awk -F ".fasta" '{print \$1}') clustalo -i ${asvs} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} clustalo -i ${asvs} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} - cat \${name}_PercentIDq.matrix | tr " " "," | grep "," >\${name}_PercentIDq.matrix + cat \${name}_PercentIDq.matrix | tr " " "," | grep "," >\${name}_PercentID.matrix rm \${name}_PercentIDq.matrix """ } From 242815a9c9d2eba7f5dadb5ff6774bafe8fb0219 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Wed, 10 Mar 2021 20:52:53 -0800 Subject: [PATCH 034/223] Add nid for pcASV processes. Refactor code --- vAMPirus.nf | 674 +++++++++++++++++++++++----------------------------- 1 file changed, 296 insertions(+), 378 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 8fab14a..01c9f59 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -486,7 +486,7 @@ if (params.clusterAAIDlist == "") { naa=slist2.size() } else { msize2=params.clusterAAIDlist - def slist2=msize2.split(',').collect{it as int} + slist2=msize2.split(',').collect{it as int} naa=slist2.size() } @@ -1125,17 +1125,11 @@ if (params.DataCheck || params.Analyze) { tuple nid, file("*_ncASV*.fasta") into ( nuclFastas_forphylogeny_ncasv, nuclFastas_forDiamond_ncasv_ch, nuclFastas_forCounts_ncasv_ch, nuclFastas_forMatrix_ncasv_ch ) script: - nid=slist.get(x-1) - mtag="ID=" + slist.get(x-1) - if (params.clusterNuclIDlist) { + nid=slist.get(x-1) + mtag="ID=" + slist.get(x-1) """ vsearch --cluster_fast ${fasta} --centroids ${params.projtag}_ncASV${nid}.fasta --threads ${task.cpus} --relabel ncASV --id .${nid} """ - } else if (params.clusterNuclID) { - """ - vsearch --cluster_fast ${fasta} --centroids ${params.projtag}_ncASV${params.clusterNuclID}.fasta --threads ${task.cpus} --relabel ncASV --id .${params.clusterNuclID} - """ - } } if (!params.skipTaxonomy) { @@ -2183,106 +2177,35 @@ if (params.DataCheck || params.Analyze) { label 'norm_cpus' + tag "${mtag}" + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV", mode: "copy", overwrite: true, pattern: '*pcASV*.{fasta}' publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{clstr,csv,gc}' publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV/Problematic", mode: "copy", overwrite: true, pattern: '*problem*.{fasta}' input: + each x from 1..naa file(fasta) from clustering_aa file(asvs) from asvfastaforaaclust output: - file("${params.projtag}_nucleotide_pcASV*.fasta") into ( pcASV_ntDiamond_ch, pcASV_nt_counts_ch, pcASV_ntmatrix_ch, pcASV_ntmafft_ch ) - file("*_aminoacid_pcASV*_noTaxonomy.fasta") into ( pcASV_aaMatrix_ch, pcASV_aaDiamond_ch, pcASV_aaMafft_ch, pcASV_aaCounts_ch, pcASVEMBOSS ) - tuple file("*.fasta"), file("*.clstr"), file("*.csv"), file("*.gc") into ( pcASVsupplementalfiles ) + tuple nid, file("${params.projtag}_nucleotide_pcASV*.fasta") into ( pcASV_ntDiamond_ch, pcASV_nt_counts_ch, pcASV_ntmatrix_ch, pcASV_ntmafft_ch ) + tuple nid, file("*_aminoacid_pcASV*_noTaxonomy.fasta") into ( pcASV_aaMatrix_ch, pcASV_aaDiamond_ch, pcASV_aaMafft_ch, pcASV_aaCounts_ch, pcASVEMBOSS ) + tuple nid, file("*.fasta"), file("*.clstr"), file("*.csv"), file("*.gc") into ( pcASVsupplementalfiles ) script: - // add awk script to count seqs - if (params.clusterAAIDlist) { - """ - set +e - cp ${params.vampdir}/bin/rename_seq.py . - for id in `echo ${params.clusterAAIDlist} | tr "," "\\n"`;do - awk 'BEGIN{RS=">";ORS=""}length(\$2)>="${params.minAA}"{print ">"\$0}' ${fasta} > ${params.projtag}_filtered_proteins.fasta - cd-hit -i ${params.projtag}_filtered_proteins.fasta -c \${id} -o ${params.projtag}_pcASV\${id}.fasta - sed 's/>Cluster />Cluster_/g' ${params.projtag}_pcASV\${id}.fasta.clstr >${params.projtag}_pcASV\${id}.clstr - grep ">Cluster_" ${params.projtag}_pcASV\${id}.clstr >temporaryclusters.list - y=\$(grep -c ">Cluster_" ${params.projtag}_pcASV\${id}.clstr) - echo ">Cluster_"\${y}"" >> ${params.projtag}_pcASV\${id}.clstr - t=1 - b=1 - for x in \$(cat temporaryclusters.list);do - echo "Extracting \$x" - name="\$( echo \$x | awk -F ">" '{print \$2}')" - clust="pcASV"\${t}"" - echo "\${name}" - awk '/^>'\${name}'\$/,/^>Cluster_'\${b}'\$/' ${params.projtag}_pcASV\${id}.clstr > "\${name}"_"\${clust}"_tmp.list - t=\$(( \${t}+1 )) - b=\$(( \${b}+1 )) - done - ls *_tmp.list - u=1 - for x in *_tmp.list;do - name="\$(echo \$x | awk -F "_p" '{print \$1}')" - echo "\${name}" - cluster="\$(echo \$x | awk -F "_" '{print \$3}')" - echo "\${cluster}" - grep "ASV" \$x | awk -F ", " '{print \$2}' | awk -F "_" '{print \$1}' | awk -F ">" '{print \$2}' > \${name}_\${cluster}_seqs_tmps.list - seqtk subseq ${asvs} \${name}_\${cluster}_seqs_tmps.list > \${name}_\${cluster}_nucleotide_sequences.fasta - vsearch --cluster_fast \${name}_\${cluster}_nucleotide_sequences.fasta --id 0.2 --centroids \${name}_\${cluster}_centroids.fasta - grep ">" \${name}_\${cluster}_centroids.fasta >> \${name}_\${cluster}_tmp_centroids.list - for y in \$( cat \${name}_\${cluster}_tmp_centroids.list );do - echo ">\${cluster}_type"\$u"" >> \${name}_\${cluster}_tmp_centroid.newheaders - u=\$(( \${u}+1 )) - done - u=1 - ./rename_seq.py \${name}_\${cluster}_centroids.fasta \${name}_\${cluster}_tmp_centroid.newheaders \${cluster}_types_labeled.fasta - done - cat *_types_labeled.fasta >> ${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta - grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$3}' | awk -F "." '{print \$1}' >tmphead.list - grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$2}' | awk -F "," '{print \$1}' >tmplen.list - paste -d"," temporaryclusters.list tmphead.list >tmp.info.csv - grep ">" ${params.projtag}_pcASV\${id}.fasta >lala.list - j=1 - for x in \$(cat lala.list);do - echo ">${params.projtag}_pcASV\${j}" >>${params.projtag}_aminoheaders.list - echo "\${x},>${params.projtag}_pcASV\${j}" >>tmpaminotype.info.csv - j=\$(( \${j}+1 )) - done - rm lala.list - awk -F "," '{print \$2}' tmp.info.csv >>tmporder.list - for x in \$(cat tmporder.list);do - grep -w "\$x" tmpaminotype.info.csv | awk -F "," '{print \$2}' >>tmpder.list - done - paste -d "," temporaryclusters.list tmplen.list tmphead.list tmpder.list >${params.projtag}_pcASVCluster\${id}_summary.csv - ./rename_seq.py ${params.projtag}_pcASV\${id}.fasta ${params.projtag}_aminoheaders.list ${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta - stats.sh in=${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_aminoacid_clustered.gc gcformat=4 overwrite=true - stats.sh in=${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_nucleotide_clustered.gc gcformat=4 overwrite=true - awk 'BEGIN{RS=">";ORS=""}length(\$2)<"${params.minAA}"{print ">"\$0}' ${fasta} >${params.projtag}_pcASV\${id}_problematic_translations.fasta - if [ `wc -l ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk '{print \$1}'` -gt 1 ];then - grep ">" ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk -F ">" '{print \$2}' > problem_tmp.list - seqtk subseq ${asvs} > ${params.projtag}_pcASV\${id}_problematic_nucleotides.fasta - else - rm ${params.projtag}_pcASV\${id}_problematic_translations.fasta - fi - rm *.list - rm Cluster* - rm *types* - rm *tmp* - rm ${params.projtag}_pcASV\${id}.fast* - done - """ - } else if (params.clusterAAID) { + // add awk script to count seqs + nid=slist2.get(x-1) + mtag="ID=" + slist2.get(x-1) """ set +e cp ${params.vampdir}/bin/rename_seq.py . - id=${params.clusterAAID} awk 'BEGIN{RS=">";ORS=""}length(\$2)>="${params.minAA}"{print ">"\$0}' ${fasta} > ${params.projtag}_filtered_proteins.fasta - cd-hit -i ${params.projtag}_filtered_proteins.fasta -c ${params.clusterAAID} -o ${params.projtag}_pcASV\${id}.fasta - sed 's/>Cluster />Cluster_/g' ${params.projtag}_pcASV\${id}.fasta.clstr >${params.projtag}_pcASV\${id}.clstr - grep ">Cluster_" ${params.projtag}_pcASV\${id}.clstr >temporaryclusters.list - y=\$(grep -c ">Cluster_" ${params.projtag}_pcASV\${id}.clstr) - echo ">Cluster_"\${y}"" >> ${params.projtag}_pcASV\${id}.clstr + cd-hit -i ${params.projtag}_filtered_proteins.fasta -c ${nid} -o ${params.projtag}_pcASV${nid}.fasta + sed 's/>Cluster />Cluster_/g' ${params.projtag}_pcASV${nid}.fasta.clstr >${params.projtag}_pcASV${nid}.clstr + grep ">Cluster_" ${params.projtag}_pcASV${nid}.clstr >temporaryclusters.list + y=\$(grep -c ">Cluster_" ${params.projtag}_pcASV${nid}.clstr) + echo ">Cluster_"\${y}"" >> ${params.projtag}_pcASV${nid}.clstr t=1 b=1 for x in \$(cat temporaryclusters.list);do @@ -2290,7 +2213,7 @@ if (params.DataCheck || params.Analyze) { name="\$( echo \$x | awk -F ">" '{print \$2}')" clust="pcASV"\${t}"" echo "\${name}" - awk '/^>'\${name}'\$/,/^>Cluster_'\${b}'\$/' ${params.projtag}_pcASV\${id}.clstr > "\${name}"_"\${clust}"_tmp.list + awk '/^>'\${name}'\$/,/^>Cluster_'\${b}'\$/' ${params.projtag}_pcASV${nid}.clstr > "\${name}"_"\${clust}"_tmp.list t=\$(( \${t}+1 )) b=\$(( \${b}+1 )) done @@ -2313,11 +2236,11 @@ if (params.DataCheck || params.Analyze) { u=1 ./rename_seq.py \${name}_\${cluster}_centroids.fasta \${name}_\${cluster}_tmp_centroid.newheaders \${cluster}_types_labeled.fasta done - cat *_types_labeled.fasta >> ${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta - grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$3}' | awk -F "." '{print \$1}' >tmphead.list - grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$2}' | awk -F "," '{print \$1}' >tmplen.list + cat *_types_labeled.fasta >> ${params.projtag}_nucleotide_pcASV${nid}_noTaxonomy.fasta + grep -w "*" ${params.projtag}_pcASV${nid}.clstr | awk '{print \$3}' | awk -F "." '{print \$1}' >tmphead.list + grep -w "*" ${params.projtag}_pcASV${nid}.clstr | awk '{print \$2}' | awk -F "," '{print \$1}' >tmplen.list paste -d"," temporaryclusters.list tmphead.list >tmp.info.csv - grep ">" ${params.projtag}_pcASV\${id}.fasta >lala.list + grep ">" ${params.projtag}_pcASV${nid}.fasta >lala.list j=1 for x in \$(cat lala.list);do echo ">${params.projtag}_pcASV\${j}" >>${params.projtag}_aminoheaders.list @@ -2329,22 +2252,22 @@ if (params.DataCheck || params.Analyze) { for x in \$(cat tmporder.list);do grep -w "\$x" tmpaminotype.info.csv | awk -F "," '{print \$2}' >>tmpder.list done - paste -d "," temporaryclusters.list tmplen.list tmphead.list tmpder.list >${params.projtag}_pcASVCluster\${id}_summary.csv - ./rename_seq.py ${params.projtag}_pcASV\${id}.fasta ${params.projtag}_aminoheaders.list ${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta - stats.sh in=${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_aminoacid_clustered.gc gcformat=4 - stats.sh in=${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_nucleotide_clustered.gc gcformat=4 - awk 'BEGIN{RS=">";ORS=""}length(\$2)<"${params.minAA}"{print ">"\$0}' ${fasta} >${params.projtag}_pcASV\${id}_problematic_translations.fasta - if [ `wc -l ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk '{print \$1}'` -gt 1 ];then - grep ">" ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk -F ">" '{print \$2}' > problem_tmp.list - seqtk subseq ${asvs} problem_tmp.list > ${params.projtag}_pcASV\${id}_problematic_nucleotides.fasta + paste -d "," temporaryclusters.list tmplen.list tmphead.list tmpder.list >${params.projtag}_pcASVCluster${nid}_summary.csv + ./rename_seq.py ${params.projtag}_pcASV${nid}.fasta ${params.projtag}_aminoheaders.list ${params.projtag}_aminoacid_pcASV${nid}_noTaxonomy.fasta + stats.sh in=${params.projtag}_aminoacid_pcASV${nid}_noTaxonomy.fasta gc=${params.projtag}_pcASV${nid}_aminoacid_clustered.gc gcformat=4 + stats.sh in=${params.projtag}_nucleotide_pcASV${nid}_noTaxonomy.fasta gc=${params.projtag}_pcASV${nid}_nucleotide_clustered.gc gcformat=4 + awk 'BEGIN{RS=">";ORS=""}length(\$2)<"${params.minAA}"{print ">"\$0}' ${fasta} >${params.projtag}_pcASV${nid}_problematic_translations.fasta + if [ `wc -l ${params.projtag}_pcASV${nid}_problematic_translations.fasta | awk '{print \$1}'` -gt 1 ];then + grep ">" ${params.projtag}_pcASV${nid}_problematic_translations.fasta | awk -F ">" '{print \$2}' > problem_tmp.list + seqtk subseq ${asvs} problem_tmp.list > ${params.projtag}_pcASV${nid}_problematic_nucleotides.fasta else - rm ${params.projtag}_pcASV\${id}_problematic_translations.fasta + rm ${params.projtag}_pcASV${nid}_problematic_translations.fasta fi rm *.list rm Cluster* rm *types* rm *tmp* - rm ${params.projtag}_pcASV\${id}.fast* + rm ${params.projtag}_pcASV${nid}.fast* """ } } @@ -2355,12 +2278,14 @@ if (params.DataCheck || params.Analyze) { label 'high_cpus' + tag "${mtag}" + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{csv,tsv}' publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*dmd.{out}' publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{fasta}' input: - file(reads) from pcASV_ntDiamond_ch + tuple nid, file(reads) from pcASV_ntDiamond_ch output: file("*.fasta") into ( pcASV_labeled ) @@ -2368,15 +2293,16 @@ if (params.DataCheck || params.Analyze) { file("*_summary_for_plot.csv") into taxplot3 script: + mtag="ID=" + slist2.get(x-1) """ set +e cp ${params.vampdir}/bin/rename_seq.py . virdb=${params.dbdir}/${params.dbname} grep ">" \${virdb} >> headers.list headers="headers.list" - for filename in ${reads};do - name=\$(ls \${filename} | awk -F "_noTax" '{print \$1}') - diamond blastx -q \${filename} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + + name=\$(ls ${reads} | awk -F "_noTax" '{print \$1}') + diamond blastx -q ${reads} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" >access.list echo "[pcASV sequence length]" >length.list @@ -2386,7 +2312,7 @@ if (params.DataCheck || params.Analyze) { echo "[pcASV#]" >otu.list echo "[Virus ID]" >"\$name"_virus.list echo "[Gene]" >"\$name"_genes.list - grep ">" \${filename} | awk -F ">" '{print \$2}' > seqids.lst + grep ">" ${reads} | awk -F ">" '{print \$2}' > seqids.lst echo "extracting genes and names" touch new_"\$name"_headers.txt j=1 @@ -2473,7 +2399,7 @@ if (params.DataCheck || params.Analyze) { done echo "Now editing "\$name" fasta headers" ###### rename_seq.py - ./rename_seq.py \${filename} new_"\$name"_headers.txt "\$name"_TaxonomyLabels.fasta + ./rename_seq.py ${reads} new_"\$name"_headers.txt "\$name"_TaxonomyLabels.fasta awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta > "\$name"_tmpssasv.fasta echo "[Sequence header]" > newnames.list cat new_"\$name"_headers.txt >> newnames.list @@ -2496,7 +2422,7 @@ if (params.DataCheck || params.Analyze) { rm "\$name"_genes.list rm newnames.list rm access.list - echo "Taxonomy inferred for: \${filename} " + echo "Taxonomy inferred for: ${reads} " done rm *headers.list """ @@ -2507,10 +2433,12 @@ if (params.DataCheck || params.Analyze) { label 'norm_cpus' + tag "${mtag}" + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Counts", mode: "copy", overwrite: true, pattern: '*.{biome,csv,txt}' input: - file(potus) from pcASV_nt_counts_ch + tuple nid, file(potus) from pcASV_nt_counts_ch file(merged) from pcASV_mergedreads_ch output: @@ -2518,15 +2446,13 @@ if (params.DataCheck || params.Analyze) { file("*.csv") into potu_Ncounts_for_report script: + mtag="ID=" + slist2.get(x-1) """ - for filename in ${potus};do - ident=\$( echo \${filename} | awk -F "pcASV" '{print \$2}' | awk -F "_noTaxonomy.fasta" '{print \$1}') - name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') - vsearch --usearch_global ${merged} --db \${filename} --id \${ident} --threads ${task.cpus} --otutabout \${name}_counts.txt --biomout \${name}_counts.biome - cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv - sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv - rm \${name}_count.csv - done + name=\$( echo ${potus} | awk -F ".fasta" '{print \$1}') + vsearch --usearch_global ${merged} --db ${potus} --id .${nid} --threads ${task.cpus} --otutabout \${name}_counts.txt --biomout \${name}_counts.biome + cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv + sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv + rm \${name}_count.csv """ } @@ -2534,32 +2460,26 @@ if (params.DataCheck || params.Analyze) { label 'low_cpus' + tag "${mtag}" + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Matrix", mode: "copy", overwrite: true input: - file(potus) from pcASV_ntmatrix_ch + tuple nid, file(potus) from pcASV_ntmatrix_ch output: file("*.matrix") into pcASVclustmatrices file("*PercentID.matrix") into potu_nucl_heatmap script: + //check --percent-id second clustalo + mtag="ID=" + slist2.get(x-1) """ - for filename in ${potus};do - ident=\$( echo \${filename} | awk -F "pcASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') - name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') - clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistanceq.matrix --full --force --threads=${task.cpus} - clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} - for x in *q.matrix;do - pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') - ya=\$(wc -l \$x | awk '{print \$1}') - echo "\$((\$ya-1))" - tail -"\$((\$ya-1))" \$x > \${pre}z.matrix - rm \$x - cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix - rm \${pre}z.matrix - done - done + name=\$( echo ${potus} | awk -F ".fasta" '{print \$1}') + clustalo -i ${potus} --distmat-out=\${name}_PairwiseDistanceq.matrix --full --force --threads=${task.cpus} + clustalo -i ${potus} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} + cat \${name}_PercentIDq.matrix | tr " " "," | grep "," >\${name}_PercentID.matrix + rm \${name}_PercentIDq.matrix """ } @@ -2569,49 +2489,50 @@ if (params.DataCheck || params.Analyze) { label 'norm_cpus' + tag "${mtag}" + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*aln.*' publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*mt*' publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*iq*' input: - file(reads) from pcASV_ntmafft_ch + tuple nid, file(reads) from pcASV_ntmafft_ch output: tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into pcASV_nucleotide_phylogeny_results file("*iq.treefile") into potu_Ntree_plot script: + mtag="ID=" + slist2.get(x-1) """ - for filename in ${reads};do - pre=\$( echo \${filename} | awk -F "_noTax" '{print \$1}' ) - mafft --maxiterate 5000 --auto \${filename} >\${pre}_ALN.fasta - trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html + pre=\$( echo ${reads} | awk -F "_noTax" '{print \$1}' ) + mafft --maxiterate 5000 --auto ${reads} >\${pre}_ALN.fasta + trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html - # pcASV_Nucleotide_ModelTest - modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_noTaxonomy_mt -d nt -s 203 --disable-checkpoint + # pcASV_Nucleotide_ModelTest + modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_noTaxonomy_mt -d nt -s 203 --disable-checkpoint - # pcASV_Nucleotide_Phylogeny - if [ "${params.iqCustomnt}" != "" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_noTaxonomy_iq --redo -T auto ${params.iqCustomnt} + # pcASV_Nucleotide_Phylogeny + if [ "${params.iqCustomnt}" != "" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_noTaxonomy_iq --redo -T auto ${params.iqCustomnt} - elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_noTaxonomy_iq -m \${mod} --redo-nt auto -b ${params.boots} + elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_noTaxonomy_iq -m \${mod} --redo-nt auto -b ${params.boots} - elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_noTaxonomy_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni + elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_noTaxonomy_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni - elif [ "${params.nonparametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_noTaxonomy_iq -m MFP --redo -nt auto -b ${params.boots} + elif [ "${params.nonparametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_noTaxonomy_iq -m MFP --redo -nt auto -b ${params.boots} - elif [ "${params.parametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_noTaxonomy_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + elif [ "${params.parametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_noTaxonomy_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - else - iqtree -s \${pre}_aln.fasta --prefix \${pre}_noTaxonomy_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - fi - done + else + iqtree -s \${pre}_aln.fasta --prefix \${pre}_noTaxonomy_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + fi """ } } @@ -2620,31 +2541,25 @@ if (params.DataCheck || params.Analyze) { label 'low_cpus' + tag "${mtag}" + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Matrix", mode: "copy", overwrite: true input: - file(prot) from pcASV_aaMatrix_ch + tuple nid, file(prot) from pcASV_aaMatrix_ch output: file("*.matrix") into pcASVaaMatrix file("*PercentID.matrix") into potu_aa_heatmap script: + mtag="ID=" + slist2.get(x-1) """ - for filename in ${prot};do - name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') - clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistanceq.matrix --full --force --threads=${task.cpus} - clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} - for x in *q.matrix;do - pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') - ya=\$(wc -l \$x | awk '{print \$1}') - echo "\$((\$ya-1))" - tail -"\$((\$ya-1))" \$x > \${pre}z.matrix - rm \$x - cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix - rm \${pre}z.matrix - done - done + name=\$( echo ${prot} | awk -F ".fasta" '{print \$1}') + clustalo -i ${prot} --distmat-out=\${name}_PairwiseDistanceq.matrix --full --force --threads=${task.cpus} + clustalo -i ${prot} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} + cat \${name}_PercentIDq.matrix | tr " " "," | grep "," >\${name}_PercentID.matrix + rm \${name}_PercentIDq.matrix """ } @@ -2654,6 +2569,8 @@ if (params.DataCheck || params.Analyze) { label 'low_cpus' + tag "${mtag}" + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/2dStructure", mode: "copy", overwrite: true, pattern: '*.{garnier}' publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/HydrophobicMoment", mode: "copy", overwrite: true, pattern: '*HydrophobicMoments.{svg}' publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/IsoelectricPoint", mode: "copy", overwrite: true, pattern: '*IsoelectricPoint.{iep,svg}' @@ -2662,38 +2579,38 @@ if (params.DataCheck || params.Analyze) { publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/2dStructure/Plots", mode: "copy", overwrite: true, pattern: '*Helical*.{svg}' input: - file(prot) from pcASVEMBOSS + tuple nid, file(prot) from pcASVEMBOSS output: tuple file("*.garnier"), file("*HydrophobicMoments.svg"), file("*IsoelectricPoint*"), file("*.pepstats"), file("*PropertiesPlot*"), file("*Helical*") into pcASV_emboss script: + // check do I need for loop + mtag="ID=" + slist2.get(x-1) """ - for filename in ${prot};do - name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') - garnier -sequence \${filename} -outfile \${name}_2dStructures.garnier - hmoment -seqall \${filename} -graph svg -plot - mv hmoment.svg ./"\${name}"_HydrophobicMoments.svg - iep -sequence \${filename} -graph svg -plot -outfile "\${name}"_IsoelectricPoint.iep - mv iep.svg ./"\${name}"_IsoelectricPoint.svg - pepstats -sequence \${filename} -outfile \${name}_ProteinProperties.pepstats - grep ">" \${filename} | awk -F ">" '{print \$2}' > tmpsequence.list - for x in \$(cat tmpsequence.list);do - echo \$x > tmp1.list - seqtk subseq \${filename} tmp1.list > tmp2.fasta - len=\$(tail -1 tmp2.fasta | awk '{print length}') - pepinfo -sequence tmp2.fasta -graph svg -outfile "\$x"_PropertiesPlot.pepinfo - mv pepinfo.svg ./"\$x"_PropertiesPlot.svg - cat "\$x"_PropertiesPlot.pepinfo >> "\${name}"_PropertiesPlot.pepinfo - rm "\$x"_PropertiesPlot.pepinfo - pepnet -sask -sequence tmp2.fasta -graph svg -sbegin1 1 -send1 \$len - mv pepnet.svg ./"\$x"_HelicalNet.svg - pepwheel -sequence tmp2.fasta -graph svg -sbegin1 1 -send1 \$len - mv pepwheel.svg ./"\$x"_HelicalWheel.svg - rm tmp1.list tmp2.fasta - done - rm tmpsequence.list + name=\$( echo ${prot} | awk -F ".fasta" '{print \$1}') + garnier -sequence ${prot} -outfile \${name}_2dStructures.garnier + hmoment -seqall ${prot} -graph svg -plot + mv hmoment.svg ./"\${name}"_HydrophobicMoments.svg + iep -sequence ${prot} -graph svg -plot -outfile "\${name}"_IsoelectricPoint.iep + mv iep.svg ./"\${name}"_IsoelectricPoint.svg + pepstats -sequence ${prot} -outfile \${name}_ProteinProperties.pepstats + grep ">" ${prot} | awk -F ">" '{print \$2}' > tmpsequence.list + for x in \$(cat tmpsequence.list);do + echo \$x > tmp1.list + seqtk subseq ${prot} tmp1.list > tmp2.fasta + len=\$(tail -1 tmp2.fasta | awk '{print length}') + pepinfo -sequence tmp2.fasta -graph svg -outfile "\$x"_PropertiesPlot.pepinfo + mv pepinfo.svg ./"\$x"_PropertiesPlot.svg + cat "\$x"_PropertiesPlot.pepinfo >> "\${name}"_PropertiesPlot.pepinfo + rm "\$x"_PropertiesPlot.pepinfo + pepnet -sask -sequence tmp2.fasta -graph svg -sbegin1 1 -send1 \$len + mv pepnet.svg ./"\$x"_HelicalNet.svg + pepwheel -sequence tmp2.fasta -graph svg -sbegin1 1 -send1 \$len + mv pepwheel.svg ./"\$x"_HelicalWheel.svg + rm tmp1.list tmp2.fasta done + rm tmpsequence.list """ } } @@ -2704,12 +2621,14 @@ if (params.DataCheck || params.Analyze) { label 'high_cpus' + tag "${mtag}" + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{csv,tsv}' publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*dmd.{out}' publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{fasta}' input: - file(reads) from pcASV_aaDiamond_ch + tuple nid, file(reads) from pcASV_aaDiamond_ch output: file("*.fasta") into ( pcASV_labeledAA ) @@ -2717,134 +2636,133 @@ if (params.DataCheck || params.Analyze) { file("*_summary_for_plot.csv") into taxplot4 script: + mtag="ID=" + slist2.get(x-1) """ cp ${params.vampdir}/bin/rename_seq.py . virdb=${params.dbdir}/${params.dbname} grep ">" \${virdb} >> headers.list headers="headers.list" - for filename in ${reads};do - name=\$(ls \${filename} | awk -F ".fasta" '{print \$1}') - diamond blastp -q \${filename} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 - echo "Preparing lists to generate summary .csv's" - echo "[Best hit accession number]" >access.list - echo "[pcASV sequence length]" >length.list - echo "[e-value]" >evalue.list - echo "[Bitscore]" >bit.list - echo "[Percent ID (aa)]" >pid.list - echo "[pcASVaa#]" >otu.list - echo "[Virus ID]" >"\$name"_virus.list - echo "[Gene]" >"\$name"_genes.list - grep ">" \${filename} | awk -F ">" '{print \$2}' > seqids.lst - echo "extracting genes and names" - touch new_"\$name"_headers.txt - j=1 - for s in \$(cat seqids.lst);do - echo "Checking for \$s hit in diamond output" - if [[ ${params.refseq} == "T" ]];then - echo "RefSeq headers specified" - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >>evalue.list - echo "\$line" | awk '{print \$11}' >>bit.list - echo "\$line" | awk '{print \$12}' >>pid.list - echo "\$line" | awk '{print \$2}' >>length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">pcASVaa\${j}_"\$virus"_"\$gene"" >> new_"\$name"_headers.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >>access.list - echo "NO_HIT" >>"\$name"_genes.list - echo "NO_HIT" >>"\$name"_virus.list - echo "NO_HIT" >>evalue.list - echo "NO_HIT" >>bit.list - echo "NO_HIT" >>pid.list - echo "NO_HIT" >>length.list - virus="NO" - gene="HIT" - echo ">pcASVaa\${j}_"\$virus"_"\$gene"" >> new_"\$name"_headers.txt - j=\$((\$j+1)) - echo "\$s done." - fi + name=\$(ls ${reads} | awk -F ".fasta" '{print \$1}') + diamond blastp -q ${reads} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + echo "Preparing lists to generate summary .csv's" + echo "[Best hit accession number]" >access.list + echo "[pcASV sequence length]" >length.list + echo "[e-value]" >evalue.list + echo "[Bitscore]" >bit.list + echo "[Percent ID (aa)]" >pid.list + echo "[pcASVaa#]" >otu.list + echo "[Virus ID]" >"\$name"_virus.list + echo "[Gene]" >"\$name"_genes.list + grep ">" ${reads} | awk -F ">" '{print \$2}' > seqids.lst + echo "extracting genes and names" + touch new_"\$name"_headers.txt + j=1 + for s in \$(cat seqids.lst);do + echo "Checking for \$s hit in diamond output" + if [[ ${params.refseq} == "T" ]];then + echo "RefSeq headers specified" + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >>evalue.list + echo "\$line" | awk '{print \$11}' >>bit.list + echo "\$line" | awk '{print \$12}' >>pid.list + echo "\$line" | awk '{print \$2}' >>length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">pcASVaa\${j}_"\$virus"_"\$gene"" >> new_"\$name"_headers.txt + j=\$((\$j+1)) + echo "\$s done." else - echo "Using RVDB headers." - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') - echo "\$s" >>otu.list - echo "\$acc" >>access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >>evalue.list - echo "\$line" | awk '{print \$11}' >>bit.list - echo "\$line" | awk '{print \$12}' >>pid.list - echo "\$line" | awk '{print \$2}' >>length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >>"\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && - echo "\$virus" | sed 's/_/ /g' >>"\$name"_virus.list - echo ">pcASVaa\${j}_"\$virus"_"\$gene"" >>new_"\$name"_headers.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >>otu.list - echo "NO_HIT" >>access.list - echo "NO_HIT" >>"\$name"_genes.list - echo "NO_HIT" >>"\$name"_virus.list - echo "NO_HIT" >>evalue.list - echo "NO_HIT" >>bit.list - echo "NO_HIT" >>pid.list - echo "NO_HIT" >>length.list - virus="NO" - gene="HIT" - echo ">pcASVaa\${j}_\${virus}_\${gene}" >>new_"\$name"_headers.txt - j=\$((\$j+1)) - echo "\$s done." - fi + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >>access.list + echo "NO_HIT" >>"\$name"_genes.list + echo "NO_HIT" >>"\$name"_virus.list + echo "NO_HIT" >>evalue.list + echo "NO_HIT" >>bit.list + echo "NO_HIT" >>pid.list + echo "NO_HIT" >>length.list + virus="NO" + gene="HIT" + echo ">pcASVaa\${j}_"\$virus"_"\$gene"" >> new_"\$name"_headers.txt + j=\$((\$j+1)) + echo "\$s done." + fi + else + echo "Using RVDB headers." + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') + echo "\$s" >>otu.list + echo "\$acc" >>access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >>evalue.list + echo "\$line" | awk '{print \$11}' >>bit.list + echo "\$line" | awk '{print \$12}' >>pid.list + echo "\$line" | awk '{print \$2}' >>length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >>"\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && + echo "\$virus" | sed 's/_/ /g' >>"\$name"_virus.list + echo ">pcASVaa\${j}_"\$virus"_"\$gene"" >>new_"\$name"_headers.txt + j=\$((\$j+1)) + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >>otu.list + echo "NO_HIT" >>access.list + echo "NO_HIT" >>"\$name"_genes.list + echo "NO_HIT" >>"\$name"_virus.list + echo "NO_HIT" >>evalue.list + echo "NO_HIT" >>bit.list + echo "NO_HIT" >>pid.list + echo "NO_HIT" >>length.list + virus="NO" + gene="HIT" + echo ">pcASVaa\${j}_\${virus}_\${gene}" >>new_"\$name"_headers.txt + j=\$((\$j+1)) + echo "\$s done." fi - echo "Done with \$s" - done - echo "Now editing "\$name" fasta headers" - ###### rename_seq.py - ./rename_seq.py \${filename} new_"\$name"_headers.txt "\$name"_wTax.fasta - awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_wTax.fasta > "\$name"_tmpssasv.fasta - echo "[Sequence header]" > newnames.list - cat new_"\$name"_headers.txt >> newnames.list - touch sequence.list - awk 'BEGIN{RS=">";ORS=""}{print \$2"\\n"}' \${name}_tmpssasv.fasta >>sequence.list - rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_summary_phyloseqObject.csv - paste -d"\\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - for x in *phyloseqObject.csv;do - echo "\$x" - lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) - tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; - sed 's/ /_/g' tmpcol.list > tmp2col.list; - cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; - rm tmpcol.list tmp2col.list - done - rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; - rm "\$name"_virus.list - rm "\$name"_genes.list - rm newnames.list - rm access.list - echo "Taxonomy inferred for: \${filename} " + fi + echo "Done with \$s" done + echo "Now editing "\$name" fasta headers" + ###### rename_seq.py + ./rename_seq.py ${reads} new_"\$name"_headers.txt "\$name"_wTax.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_wTax.fasta > "\$name"_tmpssasv.fasta + echo "[Sequence header]" > newnames.list + cat new_"\$name"_headers.txt >> newnames.list + touch sequence.list + awk 'BEGIN{RS=">";ORS=""}{print \$2"\\n"}' \${name}_tmpssasv.fasta >>sequence.list + rm "\$name"_tmpssasv.fasta + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_summary_phyloseqObject.csv + paste -d"\\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + for x in *phyloseqObject.csv;do + echo "\$x" + lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) + tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; + sed 's/ /_/g' tmpcol.list > tmp2col.list; + cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; + rm tmpcol.list tmp2col.list + done + rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; + rm "\$name"_virus.list + rm "\$name"_genes.list + rm newnames.list + rm access.list + echo "Taxonomy inferred for: ${reads} " rm *headers.list """ } @@ -2856,49 +2774,50 @@ if (params.DataCheck || params.Analyze) { label 'norm_cpus' + tag "${mtag}" + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*aln.*' publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Phylogeny/Modeltest", mode: "copy", overwrite: true, pattern: '*mt*' publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*iq*' input: - file(prot) from pcASV_aaMafft_ch + tuple nid, file(prot) from pcASV_aaMafft_ch output: tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into pcASV_protein_phylogeny_results file("*iq.treefile") into potu_Atree_plot script: + mtag="ID=" + slist2.get(x-1) """ - for filename in ${prot};do - pre=\$( echo \${filename} | awk -F ".fasta" '{print \$1}' ) - mafft --maxiterate 5000 --auto \${filename} >\${pre}_ALN.fasta - trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html + pre=\$( echo ${prot} | awk -F ".fasta" '{print \$1}' ) + mafft --maxiterate 5000 --auto ${prot} >\${pre}_ALN.fasta + trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html - # pcASV_Protein_ModelTest - modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d aa -s 203 --disable-checkpoint + # pcASV_Protein_ModelTest + modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d aa -s 203 --disable-checkpoint - # pcASV_Protein_Phylogeny - if [ "${params.iqCustomaa}" != "" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomaa} + # pcASV_Protein_Phylogeny + if [ "${params.iqCustomaa}" != "" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomaa} - elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} + elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} - elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni + elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni - elif [ "${params.nonparametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} + elif [ "${params.nonparametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} - elif [ "${params.parametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + elif [ "${params.parametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - else - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - fi - done + else + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + fi """ } } @@ -2910,7 +2829,7 @@ if (params.DataCheck || params.Analyze) { publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Counts", mode: "copy", overwrite: true input: - file(fasta) from pcASV_aaCounts_ch + tuple nid, file(fasta) from pcASV_aaCounts_ch file(merged) from mergeforpcASVaacounts file(samplist) from samplistpotu @@ -2919,35 +2838,34 @@ if (params.DataCheck || params.Analyze) { file("*counts.csv") into potu_Acounts script: + // check do I need for loop """ set +e - for filename in ${fasta};do - potu="\$( echo \${filename} | awk -F "_" '{print \$3}')" - diamond makedb --in \${filename} --db \${filename} - diamond blastx -q ${merged} -d \${filename} -p ${task.cpus} --min-score ${params.ProtCountsBit} --id ${params.ProtCountID} -l ${params.ProtCountsLength} --more-sensitive -o ${params.projtag}_\${potu}_Counts_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 --max-hsps 1 - echo "OTU_ID" >tmp.col1.txt - echo "Generating sample id list" - grep ">" \${filename} | awk -F ">" '{print \$2}' | sort | uniq > otuid.list - cat otuid.list >> tmp.col1.txt - echo "Beginning them counts tho my g" - for y in \$( cat ${samplist} );do - echo "Starting with \$y now ..." - grep "\$y" ${params.projtag}_\${potu}_Counts_dmd.out > tmp."\$y".out - echo "Isolated hits" - echo "Created uniq subject id list" - echo "\$y" > "\$y"_col.txt - echo "Starting my counts" - for z in \$(cat otuid.list);do - echo "Counting \$z hits" - echo "grep -wc "\$z" >> "\$y"_col.txt" - grep -wc "\$z" tmp."\$y".out >> "\$y"_col.txt - echo "\$z counted" - done - done - paste -d "," tmp.col1.txt *col.txt > ${params.projtag}_aminoacid_\${potu}_noTaxonomy_counts.csv - rm tmp* - rm *col.txt - done + potu="\$( echo ${fasta} | awk -F "_" '{print \$3}')" + diamond makedb --in ${fasta} --db ${fasta} + diamond blastx -q ${merged} -d ${fasta} -p ${task.cpus} --min-score ${params.ProtCountsBit} --id ${params.ProtCountID} -l ${params.ProtCountsLength} --more-sensitive -o ${params.projtag}_\${potu}_Counts_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 --max-hsps 1 + echo "OTU_ID" >tmp.col1.txt + echo "Generating sample id list" + grep ">" ${fasta} | awk -F ">" '{print \$2}' | sort | uniq > otuid.list + cat otuid.list >> tmp.col1.txt + echo "Beginning them counts tho my g" + for y in \$( cat ${samplist} );do + echo "Starting with \$y now ..." + grep "\$y" ${params.projtag}_\${potu}_Counts_dmd.out > tmp."\$y".out + echo "Isolated hits" + echo "Created uniq subject id list" + echo "\$y" > "\$y"_col.txt + echo "Starting my counts" + for z in \$(cat otuid.list);do + echo "Counting \$z hits" + echo "grep -wc "\$z" >> "\$y"_col.txt" + grep -wc "\$z" tmp."\$y".out >> "\$y"_col.txt + echo "\$z counted" + done + done + paste -d "," tmp.col1.txt *col.txt > ${params.projtag}_aminoacid_\${potu}_noTaxonomy_counts.csv + rm tmp* + rm *col.txt """ } } From bd90e4d66cc3f106c1a785db85363f6155571ed5 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Wed, 10 Mar 2021 21:01:51 -0800 Subject: [PATCH 035/223] Fix tag. Add . in ID parameter --- vAMPirus.nf | 23 ++++++++++++----------- 1 file changed, 12 insertions(+), 11 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 01c9f59..e53043a 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -2201,7 +2201,7 @@ if (params.DataCheck || params.Analyze) { set +e cp ${params.vampdir}/bin/rename_seq.py . awk 'BEGIN{RS=">";ORS=""}length(\$2)>="${params.minAA}"{print ">"\$0}' ${fasta} > ${params.projtag}_filtered_proteins.fasta - cd-hit -i ${params.projtag}_filtered_proteins.fasta -c ${nid} -o ${params.projtag}_pcASV${nid}.fasta + cd-hit -i ${params.projtag}_filtered_proteins.fasta -c .${nid} -o ${params.projtag}_pcASV${nid}.fasta sed 's/>Cluster />Cluster_/g' ${params.projtag}_pcASV${nid}.fasta.clstr >${params.projtag}_pcASV${nid}.clstr grep ">Cluster_" ${params.projtag}_pcASV${nid}.clstr >temporaryclusters.list y=\$(grep -c ">Cluster_" ${params.projtag}_pcASV${nid}.clstr) @@ -2293,7 +2293,7 @@ if (params.DataCheck || params.Analyze) { file("*_summary_for_plot.csv") into taxplot3 script: - mtag="ID=" + slist2.get(x-1) + mtag="ID=" + nid """ set +e cp ${params.vampdir}/bin/rename_seq.py . @@ -2446,7 +2446,7 @@ if (params.DataCheck || params.Analyze) { file("*.csv") into potu_Ncounts_for_report script: - mtag="ID=" + slist2.get(x-1) + mtag="ID=" + nid """ name=\$( echo ${potus} | awk -F ".fasta" '{print \$1}') vsearch --usearch_global ${merged} --db ${potus} --id .${nid} --threads ${task.cpus} --otutabout \${name}_counts.txt --biomout \${name}_counts.biome @@ -2473,7 +2473,7 @@ if (params.DataCheck || params.Analyze) { script: //check --percent-id second clustalo - mtag="ID=" + slist2.get(x-1) + mtag="ID=" + nid """ name=\$( echo ${potus} | awk -F ".fasta" '{print \$1}') clustalo -i ${potus} --distmat-out=\${name}_PairwiseDistanceq.matrix --full --force --threads=${task.cpus} @@ -2503,7 +2503,7 @@ if (params.DataCheck || params.Analyze) { file("*iq.treefile") into potu_Ntree_plot script: - mtag="ID=" + slist2.get(x-1) + mtag="ID=" + nid """ pre=\$( echo ${reads} | awk -F "_noTax" '{print \$1}' ) mafft --maxiterate 5000 --auto ${reads} >\${pre}_ALN.fasta @@ -2553,7 +2553,7 @@ if (params.DataCheck || params.Analyze) { file("*PercentID.matrix") into potu_aa_heatmap script: - mtag="ID=" + slist2.get(x-1) + mtag="ID=" + nid """ name=\$( echo ${prot} | awk -F ".fasta" '{print \$1}') clustalo -i ${prot} --distmat-out=\${name}_PairwiseDistanceq.matrix --full --force --threads=${task.cpus} @@ -2586,7 +2586,7 @@ if (params.DataCheck || params.Analyze) { script: // check do I need for loop - mtag="ID=" + slist2.get(x-1) + mtag="ID=" + nid """ name=\$( echo ${prot} | awk -F ".fasta" '{print \$1}') garnier -sequence ${prot} -outfile \${name}_2dStructures.garnier @@ -2636,7 +2636,7 @@ if (params.DataCheck || params.Analyze) { file("*_summary_for_plot.csv") into taxplot4 script: - mtag="ID=" + slist2.get(x-1) + mtag="ID=" + nid """ cp ${params.vampdir}/bin/rename_seq.py . virdb=${params.dbdir}/${params.dbname} @@ -2788,7 +2788,7 @@ if (params.DataCheck || params.Analyze) { file("*iq.treefile") into potu_Atree_plot script: - mtag="ID=" + slist2.get(x-1) + mtag="ID=" + nid """ pre=\$( echo ${prot} | awk -F ".fasta" '{print \$1}' ) mafft --maxiterate 5000 --auto ${prot} >\${pre}_ALN.fasta @@ -2826,6 +2826,8 @@ if (params.DataCheck || params.Analyze) { label 'high_cpus' + tag "${mtag}" + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Counts", mode: "copy", overwrite: true input: @@ -2839,6 +2841,7 @@ if (params.DataCheck || params.Analyze) { script: // check do I need for loop + mtag="ID=" + nid """ set +e potu="\$( echo ${fasta} | awk -F "_" '{print \$3}')" @@ -3005,8 +3008,6 @@ if (params.DataCheck || params.Analyze) { } - } - } else { println("\n\t\033[0;31mMandatory argument not specified. For more info use `nextflow run vAMPirus.nf --help`\n\033[0m") exit 0 From 6dd02e8c50a74741392aeb7bfba5dc1cf41b502e Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Wed, 10 Mar 2021 21:04:37 -0800 Subject: [PATCH 036/223] Fix process pcASV_Nucleotide_Taxonomy_Inference --- vAMPirus.nf | 240 ++++++++++++++++++++++++++-------------------------- 1 file changed, 119 insertions(+), 121 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index e53043a..d86cfc7 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -2300,130 +2300,128 @@ if (params.DataCheck || params.Analyze) { virdb=${params.dbdir}/${params.dbname} grep ">" \${virdb} >> headers.list headers="headers.list" - - name=\$(ls ${reads} | awk -F "_noTax" '{print \$1}') - diamond blastx -q ${reads} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 - echo "Preparing lists to generate summary .csv's" - echo "[Best hit accession number]" >access.list - echo "[pcASV sequence length]" >length.list - echo "[e-value]" >evalue.list - echo "[Bitscore]" >bit.list - echo "[Percent ID (aa)]" >pid.list - echo "[pcASV#]" >otu.list - echo "[Virus ID]" >"\$name"_virus.list - echo "[Gene]" >"\$name"_genes.list - grep ">" ${reads} | awk -F ">" '{print \$2}' > seqids.lst - echo "extracting genes and names" - touch new_"\$name"_headers.txt - j=1 - for s in \$(cat seqids.lst);do - echo "Checking for \$s hit in diamond output" - if [[ ${params.refseq} == "T" ]];then - echo "RefSeq headers specified" - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >>evalue.list - echo "\$line" | awk '{print \$11}' >>bit.list - echo "\$line" | awk '{print \$12}' >>pid.list - echo "\$line" | awk '{print \$2}' >>length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">pcASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_headers.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >>access.list - echo "NO_HIT" >>"\$name"_genes.list - echo "NO_HIT" >>"\$name"_virus.list - echo "NO_HIT" >>evalue.list - echo "NO_HIT" >>bit.list - echo "NO_HIT" >>pid.list - echo "NO_HIT" >>length.list - virus="NO" - gene="HIT" - echo ">pcASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_headers.txt - j=\$((\$j+1)) - echo "\$s done." - fi + name=\$(ls ${reads} | awk -F "_noTax" '{print \$1}') + diamond blastx -q ${reads} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + echo "Preparing lists to generate summary .csv's" + echo "[Best hit accession number]" >access.list + echo "[pcASV sequence length]" >length.list + echo "[e-value]" >evalue.list + echo "[Bitscore]" >bit.list + echo "[Percent ID (aa)]" >pid.list + echo "[pcASV#]" >otu.list + echo "[Virus ID]" >"\$name"_virus.list + echo "[Gene]" >"\$name"_genes.list + grep ">" ${reads} | awk -F ">" '{print \$2}' > seqids.lst + echo "extracting genes and names" + touch new_"\$name"_headers.txt + j=1 + for s in \$(cat seqids.lst);do + echo "Checking for \$s hit in diamond output" + if [[ ${params.refseq} == "T" ]];then + echo "RefSeq headers specified" + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >>evalue.list + echo "\$line" | awk '{print \$11}' >>bit.list + echo "\$line" | awk '{print \$12}' >>pid.list + echo "\$line" | awk '{print \$2}' >>length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">pcASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_headers.txt + j=\$((\$j+1)) + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >>access.list + echo "NO_HIT" >>"\$name"_genes.list + echo "NO_HIT" >>"\$name"_virus.list + echo "NO_HIT" >>evalue.list + echo "NO_HIT" >>bit.list + echo "NO_HIT" >>pid.list + echo "NO_HIT" >>length.list + virus="NO" + gene="HIT" + echo ">pcASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_headers.txt + j=\$((\$j+1)) + echo "\$s done." + fi + else + echo "Using RVDB headers." + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') + echo "\$s" >>otu.list + echo "\$acc" >>access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >>evalue.list + echo "\$line" | awk '{print \$11}' >>bit.list + echo "\$line" | awk '{print \$12}' >>pid.list + echo "\$line" | awk '{print \$2}' >>length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >>"\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && + echo "\$virus" | sed 's/_/ /g' >>"\$name"_virus.list + echo ">pcASV\${j}_"\$virus"_"\$gene"" >>new_"\$name"_headers.txt + j=\$((\$j+1)) + echo "\$s done." else - echo "Using RVDB headers." - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') - echo "\$s" >>otu.list - echo "\$acc" >>access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >>evalue.list - echo "\$line" | awk '{print \$11}' >>bit.list - echo "\$line" | awk '{print \$12}' >>pid.list - echo "\$line" | awk '{print \$2}' >>length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >>"\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && - echo "\$virus" | sed 's/_/ /g' >>"\$name"_virus.list - echo ">pcASV\${j}_"\$virus"_"\$gene"" >>new_"\$name"_headers.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >>otu.list - echo "NO_HIT" >>access.list - echo "NO_HIT" >>"\$name"_genes.list - echo "NO_HIT" >>"\$name"_virus.list - echo "NO_HIT" >>evalue.list - echo "NO_HIT" >>bit.list - echo "NO_HIT" >>pid.list - echo "NO_HIT" >>length.list - virus="NO" - gene="HIT" - echo ">pcASV\${j}_"\$virus"_"\$gene"" >>new_"\$name"_headers.txt - j=\$((\$j+1)) - echo "\$s done." - fi + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >>otu.list + echo "NO_HIT" >>access.list + echo "NO_HIT" >>"\$name"_genes.list + echo "NO_HIT" >>"\$name"_virus.list + echo "NO_HIT" >>evalue.list + echo "NO_HIT" >>bit.list + echo "NO_HIT" >>pid.list + echo "NO_HIT" >>length.list + virus="NO" + gene="HIT" + echo ">pcASV\${j}_"\$virus"_"\$gene"" >>new_"\$name"_headers.txt + j=\$((\$j+1)) + echo "\$s done." fi - echo "Done with \$s" - done - echo "Now editing "\$name" fasta headers" - ###### rename_seq.py - ./rename_seq.py ${reads} new_"\$name"_headers.txt "\$name"_TaxonomyLabels.fasta - awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta > "\$name"_tmpssasv.fasta - echo "[Sequence header]" > newnames.list - cat new_"\$name"_headers.txt >> newnames.list - touch sequence.list - echo " " > sequence.list - grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list - rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - for x in *phyloseqObject.csv;do - echo "\$x" - lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) - tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; - sed 's/ /_/g' tmpcol.list > tmp2col.list; - cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_noTaxonomy_summary_for_plot.csv; - rm tmpcol.list tmp2col.list - done - rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; - rm "\$name"_virus.list - rm "\$name"_genes.list - rm newnames.list - rm access.list - echo "Taxonomy inferred for: ${reads} " + fi + echo "Done with \$s" + done + echo "Now editing "\$name" fasta headers" + ###### rename_seq.py + ./rename_seq.py ${reads} new_"\$name"_headers.txt "\$name"_TaxonomyLabels.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta > "\$name"_tmpssasv.fasta + echo "[Sequence header]" > newnames.list + cat new_"\$name"_headers.txt >> newnames.list + touch sequence.list + echo " " > sequence.list + grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list + rm "\$name"_tmpssasv.fasta + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + for x in *phyloseqObject.csv;do + echo "\$x" + lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) + tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; + sed 's/ /_/g' tmpcol.list > tmp2col.list; + cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_noTaxonomy_summary_for_plot.csv; + rm tmpcol.list tmp2col.list done + rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; + rm "\$name"_virus.list + rm "\$name"_genes.list + rm newnames.list + rm access.list + echo "Taxonomy inferred for: ${reads} " rm *headers.list """ } From 7efd1e2af1643c337fa9dad075878450f59fea04 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Fri, 12 Mar 2021 16:12:57 -0800 Subject: [PATCH 037/223] Add missing } --- vAMPirus.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index d86cfc7..ba65098 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -2270,7 +2270,6 @@ if (params.DataCheck || params.Analyze) { rm ${params.projtag}_pcASV${nid}.fast* """ } - } if (!params.skipTaxonomy) { @@ -3005,6 +3004,7 @@ if (params.DataCheck || params.Analyze) { } } + } } else { println("\n\t\033[0;31mMandatory argument not specified. For more info use `nextflow run vAMPirus.nf --help`\n\033[0m") From 284c6ecd2b888e9d82f11f653ba7b321f91ce530 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Fri, 12 Mar 2021 17:26:54 -0800 Subject: [PATCH 038/223] WIP. Add tuple. Fix report channels --- vAMPirus.nf | 54 ++++++++++++++++++++++++----------------------------- 1 file changed, 24 insertions(+), 30 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index ba65098..5625089 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1149,7 +1149,7 @@ if (params.DataCheck || params.Analyze) { output: file("*.fasta") into tax_labeled_fasta_ncasv tuple file("*_phyloseqObject.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond_ncasv - file("*ncASV*summary_for_plot.csv") into taxplot_ncasv + tuple nid, file("*ncASV*summary_for_plot.csv") into taxplot_ncasv script: mtag="ID=" + nid @@ -1303,8 +1303,7 @@ if (params.DataCheck || params.Analyze) { output: tuple file("*_counts.csv"), file("*_counts.biome") into counts_vsearch_ncasv - tuple val("${id}"), file("*ncASV*counts.csv") into notu_counts_plots - file("*ASV*counts.csv") into ncasv_counts_plots + tuple nid, file("*ncASV*counts.csv") into notu_counts_plots script: mtag="ID=" + nid @@ -1329,7 +1328,7 @@ if (params.DataCheck || params.Analyze) { output: file("*.matrix") into clustmatrices_ncasv - file("*ncASV*PercentID.matrix") into notu_heatmap + tuple nid, file("*ncASV*PercentID.matrix") into notu_heatmap script: mtag="ID=" + nid @@ -1359,7 +1358,7 @@ if (params.DataCheck || params.Analyze) { output: tuple nid, file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into align_results_ncasv - file("*iq.treefile") into nucl_phyl_plot_ncasv + tuple nid, file("*iq.treefile") into nucl_phyl_plot_ncasv script: mtag="ID=" + nid @@ -1643,17 +1642,15 @@ if (params.DataCheck || params.Analyze) { output: tuple file("*_counts.csv"), file("*_counts.biome") into counts_vsearch_asv - file("*_ASV*counts.csv") into asv_counts_plots_asv + file("*_ASV*counts.csv") into asv_counts_plots script: """ - if [ `echo ${asvs} | grep -c "ASV"` -eq 1 ];then - name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - vsearch --usearch_global ${merged} --db ${asvs} --id .${params.asvcountID} --threads ${task.cpus} --otutabout "\$name"_counts.txt --biomout "\$name"_counts.biome - cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv - sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv - rm \${name}_count.csv - fi + name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') + vsearch --usearch_global ${merged} --db ${asvs} --id .${params.asvcountID} --threads ${task.cpus} --otutabout "\$name"_counts.txt --biomout "\$name"_counts.biome + cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv + sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv + rm \${name}_count.csv """ } @@ -1668,7 +1665,7 @@ if (params.DataCheck || params.Analyze) { output: file("*.matrix") into clustmatrices_asv - file("*_ASV*PercentID.matrix") into asv_heatmap_asv + file("*_ASV*PercentID.matrix") into asv_heatmap script: // remove if statement later (no fin) @@ -2289,7 +2286,7 @@ if (params.DataCheck || params.Analyze) { output: file("*.fasta") into ( pcASV_labeled ) tuple file("*_phyloseqObject.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AAdiamond - file("*_summary_for_plot.csv") into taxplot3 + tuple nid, file("*_summary_for_plot.csv") into taxplot3 script: mtag="ID=" + nid @@ -2440,7 +2437,7 @@ if (params.DataCheck || params.Analyze) { output: tuple file("*_counts.txt"), file("*_counts.biome") into pcASVcounts_vsearch - file("*.csv") into potu_Ncounts_for_report + tuple nid, file("*.csv") into potu_Ncounts_for_report script: mtag="ID=" + nid @@ -2466,7 +2463,7 @@ if (params.DataCheck || params.Analyze) { output: file("*.matrix") into pcASVclustmatrices - file("*PercentID.matrix") into potu_nucl_heatmap + tuple nid, file("*PercentID.matrix") into potu_nucl_heatmap script: //check --percent-id second clustalo @@ -2497,7 +2494,7 @@ if (params.DataCheck || params.Analyze) { output: tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into pcASV_nucleotide_phylogeny_results - file("*iq.treefile") into potu_Ntree_plot + tuple nid, file("*iq.treefile") into potu_Ntree_plot script: mtag="ID=" + nid @@ -2547,7 +2544,7 @@ if (params.DataCheck || params.Analyze) { output: file("*.matrix") into pcASVaaMatrix - file("*PercentID.matrix") into potu_aa_heatmap + tuple nid, file("*PercentID.matrix") into potu_aa_heatmap script: mtag="ID=" + nid @@ -2630,7 +2627,7 @@ if (params.DataCheck || params.Analyze) { output: file("*.fasta") into ( pcASV_labeledAA ) tuple file("*phyloseqObject.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_potuaadiamond - file("*_summary_for_plot.csv") into taxplot4 + tuple nid, file("*_summary_for_plot.csv") into taxplot4 script: mtag="ID=" + nid @@ -2782,7 +2779,7 @@ if (params.DataCheck || params.Analyze) { output: tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into pcASV_protein_phylogeny_results - file("*iq.treefile") into potu_Atree_plot + tuple nid, file("*iq.treefile") into potu_Atree_plot script: mtag="ID=" + nid @@ -2834,7 +2831,7 @@ if (params.DataCheck || params.Analyze) { output: tuple file("*_counts.csv"), file("*dmd.out") into potuaacounts_summary - file("*counts.csv") into potu_Acounts + tuple nid, file("*counts.csv") into potu_Acounts script: // check do I need for loop @@ -2904,12 +2901,9 @@ if (params.DataCheck || params.Analyze) { """ echo "Read processing steps skipped." >filter_reads.txt """ - } - } - //NEW REPORT !!!!!!!!!!!!!!!!! /*Report_ASV asv_counts_plots -> ${params.projtag}_ASV_counts.csv @@ -2918,11 +2912,11 @@ if (params.DataCheck || params.Analyze) { nucl_phyl_plot -> ${params.projtag}_ASV_iq.treefile */ report_asv = Channel.create() - asv_counts_plots.mix(taxplot1, asv_heatmap, nucl_phyl_plot).flatten().buffer(size:4).into(report_asv) + asv_counts_plots.mix(taxplot_asv, asv_heatmap, nucl_phyl_plot_asv).flatten().buffer(size:4).into(report_asv) if (params.ncASV) { report_ncasv = Channel.create() - notu_counts_plots.mix(taxplot1a, notu_heatmap, nucl_phyl_plot_ncasv).flatten().toSortedList().flatten().buffer(size:4).into(report_ncasv) + notu_counts_plots.mix(taxplot_ncasv, notu_heatmap, nucl_phyl_plot_ncasv).groupTuple(by:0, size:4).into(report_ncasv) /* notu_counts_plots -> ${params.projtag}_ncASV${id}_counts.csv taxplot1a -> ${params.projtag}_ncASV${id}_summary_for_plot.csv @@ -2935,7 +2929,7 @@ if (params.DataCheck || params.Analyze) { if (params.pcASV) { report_pcasv_aa = Channel.create() - potu_Acounts.mix(taxplot4, potu_aa_heatmap, potu_Atree_plot).flatten().toSortedList().flatten().buffer(size:4).into(report_pcasv_aa) + potu_Acounts.mix(taxplot4, potu_aa_heatmap, potu_Atree_plot).groupTuple(by:0, size:4).into(report_pcasv_aa) /*Report_pcASV_AminoAcid potu_Acounts -> ${params.projtag}_pcASV${id}_noTaxonomy_counts.csv taxplot4 -> ${params.projtag}_aminoacid_pcASV${id}_noTaxonomy_summary_for_plot.csv @@ -2943,7 +2937,7 @@ if (params.DataCheck || params.Analyze) { potu_Atree_plot -> ${params.projtag}_aminoacid_pcASV${id}_noTaxonomy_iq.treefile */ report_pcasv_nucl = Channel.create() - potu_Ncounts_for_report.mix(taxplot3, potu_nucl_heatmap, potu_Ntree_plot).flatten().toSortedList().flatten().buffer(size:4).into(report_pcasv_nucl) + potu_Ncounts_for_report.mix(taxplot3, potu_nucl_heatmap, potu_Ntree_plot).groupTuple(by:0, size:4).into(report_pcasv_nucl) /*Report_pcASV_Nucleotide potu_Ncounts_for_report -> ${params.projtag}_nucleotide_pcASV${id}_noTaxonomy_counts.csv taxplot3 -> ${params.projtag}_nucleotide_pcASV${id}_noTaxonomy_summary_for_plot.csv @@ -2970,7 +2964,7 @@ if (params.DataCheck || params.Analyze) { } report_all_ch = Channel.create() - report_asv.mix(report_ncasv, report_pcasv_aa, report_pcasv_nucl, report_aminotypes).buffer(size:1).flatten().toList().view().into(report_all_ch) + report_asv.mix(report_ncasv, report_pcasv_aa, report_pcasv_nucl, report_aminotypes).view().into(report_all_ch) process Report { From 790095f428568f015c602a35527164d9c51fada0 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Fri, 12 Mar 2021 18:12:18 -0800 Subject: [PATCH 039/223] Fix issue with input files in report --- vAMPirus.nf | 1 + 1 file changed, 1 insertion(+) diff --git a/vAMPirus.nf b/vAMPirus.nf index 5625089..8688a19 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -2981,6 +2981,7 @@ if (params.DataCheck || params.Analyze) { script: """ + if [ -e input.3 ];then cat input.3 | sed -r 's|\\[(\\/\\w+.*)\\]|\\1|g' | tr -d " " | tr "," "\\n" >flist.txt ; for x in `cat flist.txt`;do cp \$x . ;done ;fi name=\$( ls *summary_for_plot.csv | awk -F "_summary_for_plot.csv" '{print \$1}') cp ${params.vampdir}/bin/vAMPirus_Report.Rmd . cp ${params.vampdir}/example_data/conf/vamplogo.png . From a23b1fedae76c9980860e519a60d437bd3a84ee7 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Sun, 14 Mar 2021 09:19:13 -0700 Subject: [PATCH 040/223] Fix channel. Erase sed line --- vAMPirus.nf | 3 +-- 1 file changed, 1 insertion(+), 2 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 8688a19..8cc6478 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -2964,7 +2964,7 @@ if (params.DataCheck || params.Analyze) { } report_all_ch = Channel.create() - report_asv.mix(report_ncasv, report_pcasv_aa, report_pcasv_nucl, report_aminotypes).view().into(report_all_ch) + report_asv.mix(report_ncasv, report_pcasv_aa, report_pcasv_nucl, report_aminotypes).map{it.flatten()}.into(report_all_ch) process Report { @@ -2981,7 +2981,6 @@ if (params.DataCheck || params.Analyze) { script: """ - if [ -e input.3 ];then cat input.3 | sed -r 's|\\[(\\/\\w+.*)\\]|\\1|g' | tr -d " " | tr "," "\\n" >flist.txt ; for x in `cat flist.txt`;do cp \$x . ;done ;fi name=\$( ls *summary_for_plot.csv | awk -F "_summary_for_plot.csv" '{print \$1}') cp ${params.vampdir}/bin/vAMPirus_Report.Rmd . cp ${params.vampdir}/example_data/conf/vamplogo.png . From 1c026f1cf534b5344c06ef630220e6c7b54383c1 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Sun, 14 Mar 2021 09:36:18 -0700 Subject: [PATCH 041/223] Fix onComplete print --- vAMPirus.nf | 33 ++++++++++++++++++++++----------- 1 file changed, 22 insertions(+), 11 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 8cc6478..8f345fe 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -3004,15 +3004,26 @@ if (params.DataCheck || params.Analyze) { println("\n\t\033[0;31mMandatory argument not specified. For more info use `nextflow run vAMPirus.nf --help`\n\033[0m") exit 0 } - -workflow.onComplete { - log.info ( workflow.success ? \ - "---------------------------------------------------------------------------------" \ - + "\n\033[0;32mDone! Open the following reports in your browser\033[0m" \ - + "\n\033[0;32mPipeline performance report: ${params.workingdir}/${params.outdir}/${params.tracedir}/vampirus_report.html\033[0m" \ - + "\n\033[0;32mvAMPirus --DataCheck interactive report: ${params.workingdir}/${params.outdir}/DataCheck/*.hmtl\033[0m" \ - + "\n\033[0;32mvAMPirus --Analyze interactive report: ${params.workingdir}/${params.outdir}/Analyze/*.hmtl\033[0m" \ - : \ - "---------------------------------------------------------------------------------" \ - + "\n\033[0;31mSomething went wrong. Check error message below and/or log files.\033[0m" ) +if (params.DataCheck) { + workflow.onComplete { + log.info ( workflow.success ? \ + "---------------------------------------------------------------------------------" \ + + "\n\033[0;32mDone! Open the following reports in your browser\033[0m" \ + + "\n\033[0;32mPipeline performance report: ${params.workingdir}/${params.outdir}/${params.tracedir}/vampirus_report.html\033[0m" \ + + "\n\033[0;32mvAMPirus --DataCheck interactive report: ${params.workingdir}/${params.outdir}/DataCheck/*.hmtl\033[0m" \ + : \ + "---------------------------------------------------------------------------------" \ + + "\n\033[0;31mSomething went wrong. Check error message below and/or log files.\033[0m" ) + } +} else if (params.Analyze) { + workflow.onComplete { + log.info ( workflow.success ? \ + "---------------------------------------------------------------------------------" \ + + "\n\033[0;32mDone! Open the following reports in your browser\033[0m" \ + + "\n\033[0;32mPipeline performance report: ${params.workingdir}/${params.outdir}/${params.tracedir}/vampirus_report.html\033[0m" \ + + "\n\033[0;32mvAMPirus --Analyze interactive report: ${params.workingdir}/${params.outdir}/Analyze/*.hmtl\033[0m" \ + : \ + "---------------------------------------------------------------------------------" \ + + "\n\033[0;31mSomething went wrong. Check error message below and/or log files.\033[0m" ) + } } From 05f6f61cd24b92829854cbb4a2ef372b82cd9542 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Tue, 23 Mar 2021 09:24:12 -0500 Subject: [PATCH 042/223] added oligtyping installation and databse annotation download and options --- vampirus.config | 8 +++++--- vampirus_startup.sh | 17 +++++++++++++---- 2 files changed, 18 insertions(+), 7 deletions(-) diff --git a/vampirus.config b/vampirus.config index dae7c9d..db310e0 100644 --- a/vampirus.config +++ b/vampirus.config @@ -25,7 +25,7 @@ params { // Merged read length filtering parameters - // Minimum merged read length - reads below the specified maximum read length will be used for counts only + // Minimum merged read length - reads with lengths greater than minLen and below the specified maximum read length will be used for counts only minLen="400" // Maximum merged read length - reads with length equal to the specified max read length will be used to generate uniques and ASVs (safe to set at expected amplicon size to start) maxLen="420" @@ -57,7 +57,7 @@ params { alpha="1" // Minimum size or representation for sequence to be considered in ASV generation minSize="8" - // Percent similarity to cluster nucleotide ASV sequences + // Percent similarity to cluster nucleotide ASV sequences (used when --ncASV is set) clusterNuclID="85" // List of percent similarities to cluster nucleotide ASV sequences - must be separated by a comma (ex. "95,96") clusterNuclIDlist="" @@ -65,7 +65,7 @@ params { clusterAAID="97" // List of percent similarities to cluster aminoacid sequences - must be separated by "95,96" clusterAAIDlist="" - // Minimum length of amino acid translation to be considered during protein clustered ASV (pcASV) generation. Recommended to put this at the expected aminoacid sequence length based on your maximum read length (e.g. if maxLen="420", then minAA should be 420/3 or 140) + // Minimum length of amino acid translation to be considered during protein clustered ASV (pcASV) generation. Recommended to put this at the expected amino acid sequence length based on your maximum read length (e.g. if maxLen="420", then minAA should be 420/3 so 140) minAA="140" // Counts table generation parameters @@ -86,6 +86,8 @@ params { dbname="DATABASENAME" // Path to Directory where database is being stored dbdir="DATABASEDIR" + // Path to hmm annotation .txt file - see manual for information on this. Leave as is if not planning on using. + dbanno="DATABASEANNOT" // Toggle use of RefSeq header format; default is Reverence Viral DataBase (RVDB) refseq="F" // Set minimum bitscore for best hit in taxonomy assignment diff --git a/vampirus_startup.sh b/vampirus_startup.sh index 5f5089a..e4a618e 100644 --- a/vampirus_startup.sh +++ b/vampirus_startup.sh @@ -18,6 +18,7 @@ vampirus_startup.sh -h [-d 1|2|3|4] [-s] [ -d 1|2|3|4 ] Set this option to create a database directiory within the current working directory and download the following databases for taxonomy assignment: 1 - Download the proteic version of the Reference Viral DataBase (See the paper for more information on this database: https://f1000research.com/articles/8-530) + NOTE: need to use RVDB for sequence classification 2 - Download only NCBIs Viral protein RefSeq database 3 - Download only the complete NCBI NR protein database 4 - Download all three databases @@ -124,6 +125,11 @@ conda_c() { fi } +echo "Downloading oligotyping program" +conda init && source activate vAMPirus +pip install oligotyping --yes +conda deactivate + nextflow_c() { source_c cd $mypwd @@ -186,15 +192,18 @@ if [[ $DATABASE -eq 1 ]] then mkdir "$mypwd"/Databases cd "$mypwd"/Databases dir="$(pwd)" - echo "Database installation: RVDB version 19.0 (latest as of 2020-06)" - curl -o U-RVDBv19.0-prot.fasta.bz2 https://rvdb-prot.pasteur.fr/files/U-RVDBv19.0-prot.fasta.bz2 - bunzip2 U-RVDBv19.0-prot.fasta.bz2 + echo "Database installation: RVDB version 21.0 (latest as of 2021-02)" + curl -o U-RVDBv21.0-prot.fasta.bz2 https://rvdb-prot.pasteur.fr/files/U-RVDBv21.0-prot.fasta.bz2 + bunzip2 U-RVDBv21.0-prot.fasta.bz2 + curl -o U-RVDBv21.0-prot-hmm-txt.zip https://rvdb-prot.pasteur.fr/files/U-RVDBv21.0-prot-hmm-txt.zip echo "Editing confiration file for you now..." sed 's/DATABASENAME/U-RVDBv19.0-prot.fasta/g' "$mypwd"/vampirus.config > tmp1.config sed "s|DATABASEDIR|${dir}|g" tmp1.config > tmp2.config + sed "s|DATABASEANNOT|${dir}/U-RVDBv21.0-prot-hmm-txt.zip|g" tmp2.config > tmp3.config rm tmp1.config - cat tmp2.config > "$mypwd"/vampirus.config rm tmp2.config + cat tmp3.config > "$mypwd"/vampirus.config + rm tmp3.config echo "Database downloaded and configuration file edited, you should confirm the path and database name was set correctly in the config file." elif [[ $DATABASE -eq 2 ]] then mkdir "$mypwd"/Databases From 141c6b085fd896c03c2ac913b789c2d595d20cfc Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Tue, 23 Mar 2021 09:50:11 -0500 Subject: [PATCH 043/223] fixed oligotyping install --- vampirus_startup.sh | 11 ++++++----- 1 file changed, 6 insertions(+), 5 deletions(-) diff --git a/vampirus_startup.sh b/vampirus_startup.sh index e4a618e..13c7e09 100644 --- a/vampirus_startup.sh +++ b/vampirus_startup.sh @@ -125,11 +125,6 @@ conda_c() { fi } -echo "Downloading oligotyping program" -conda init && source activate vAMPirus -pip install oligotyping --yes -conda deactivate - nextflow_c() { source_c cd $mypwd @@ -181,6 +176,12 @@ else cat tmp1.config > "$mypwd"/vampirus.config rm tmp1.config fi + +echo "Downloading oligotyping program" +conda init && source activate vAMPirus +pip install oligotyping +conda deactivate + echo "-------------------------------------------------------------------------------- Conda check/install done" echo "Now lets check the status of Nextflow on your system..." From 9698c229b12b10103fc97ea47e32193127815c0d Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Tue, 23 Mar 2021 14:48:40 -0500 Subject: [PATCH 044/223] Added blast to yml file and added Shannon entropy analysis to datacheck --- vAMPirus.nf | 164 +++++++++++++++++++++++++++++++++++++++++++++-- vampirus_env.yml | 1 + 2 files changed, 160 insertions(+), 5 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 8f345fe..e37dc75 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -826,7 +826,7 @@ if (params.DataCheck || params.Analyze) { """ } - process Extract_Uniques { + process Extracting_Uniques { label 'low_cpus' @@ -872,7 +872,7 @@ if (params.DataCheck || params.Analyze) { file(fasta) from reads_vsearch4_ch output: - file("*ASVs.fasta") into ( reads_vsearch5_ch, nucl2aa, asvsforAminotyping, asvfastaforcounts, asvaminocheck ) + file("*ASVs.fasta") into ( reads_vsearch5_ch, asv_med, nucl2aa, asvsforAminotyping, asvfastaforcounts, asvaminocheck ) script: """ @@ -946,7 +946,7 @@ if (params.DataCheck || params.Analyze) { output: file("number_per_percentage_prot.csv") into number_per_percent_prot_plot - + file("*pcASV100*") into amino_med script: // add awk script to count seqs """ @@ -1040,6 +1040,158 @@ if (params.DataCheck || params.Analyze) { """ } + process ASV_Shannons_Entropy_Analysis { + + label 'norm_cpus' + + publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Nucleotide/ShannonEntropy", mode: "copy", overwrite: true + + input: + file(asvs) from asv_med + + output: + + file("*_ASV_entropy_breakdown.csv") into asv_entro_csv + file("*ASV*") into entrop + + script: + """ + set +e + #alignment + mafft --thread ${task.cpus} --maxiterate 15000 --auto ${asvs} > ${params.projtag}_ASVs_mafftAlign.fasta + #trimming + trimal -in ${params.projtag}_ASVs_mafftAlign.fasta -out ${params.projtag}_ASVs_mafftAligned.fasta -keepheader -fasta -automated1 + rm ${params.projtag}_ASVs_mafftAlign.fasta + o-trim-uninformative-columns-from-alignment ${params.projtag}_ASVs_mafftAligned.fasta + mv ${params.projtag}_ASVs_mafftAligned.fasta-TRIMMED ./${params.projtag}_ASVs_Aligned_informativeonly.fasta + #entopy analysis + entropy-analysis ${params.projtag}_ASVs_Aligned_informativeonly.fasta + #summarize entropy peaks + awk '{print \$2}' testaligned.fasta-ENTROPY >> tmp_value.list + for x in \$(cat tmp_value.list) + do echo "\$x" + if [[ \$(echo ""\$x" > 0.0"|bc -l) -eq 1 ]]; + then echo dope >> above-0.0-.list + fi + if [[ \$(echo ""\$x" > 0.1"|bc -l) -eq 1 ]]; + then echo dope >> above-0.1-.list + fi + if [[ \$(echo ""\$x" > 0.2"|bc -l) -eq 1 ]]; + then echo dope >> above-0.2-.list + fi + if [[ \$(echo ""\$x" > 0.3"|bc -l) -eq 1 ]]; + then echo dope >> above-0.3-.list + fi + if [[ \$(echo ""\$x" > 0.4"|bc -l) -eq 1 ]]; + then echo dope >> above-0.4-.list + fi + if [[ \$(echo ""\$x" > 0.5"|bc -l) -eq 1 ]]; + then echo dope >> above-0.5-.list + fi + if [[ \$(echo ""\$x" > 0.6"|bc -l) -eq 1 ]]; + then echo dope >> above-0.6-.list + fi + if [[ \$(echo ""\$x" > 0.7"|bc -l) -eq 1 ]]; + then echo dope >> above-0.7-.list + fi + if [[ \$(echo ""\$x" > 0.8"|bc -l) -eq 1 ]]; + then echo dope >> above-0.8-.list + fi + if [[ \$(echo ""\$x" > 0.9"|bc -l) -eq 1 ]]; + then echo dope >> above-0.9-.list + fi + if [[ \$(echo ""\$x" > 1.0"|bc -l) -eq 1 ]]; + then echo dope >> above-1.0-.list + fi + if [[ \$(echo ""\$x" > 1.5"|bc -l) -eq 1 ]]; + then echo dope >> above-1.5-.list + fi + done + for z in above*.list; + do entrop=\$(echo \$z | awk -F "-" '{print \$2}') + echo ""\$entrop", "\$(wc -l \$z | awk '{print \$1}')"" >> ${params.projtag}_ASV_entropy_breakdown.csv + done + rm above* + """ + + } + + process AminoType_Shannons_Entropy_Analysis { + + label 'norm_cpus' + + publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Aminoacid/ShannonEntropy", mode: "copy", overwrite: true, pattern: '*{.csv}' + + input: + file(aminos) from amino_med + + output: + + file("*AminoType_entropy_breakdown.csv") into amino_entro_csv + file("*AminoTypes*") into aminos + + script: + """ + #alignment + mafft --thread ${task.cpus} --maxiterate 15000 --auto ${aminos} > ${params.projtag}_AminoTypes_mafftAlign.fasta + #trimming + trimal -in ${params.projtag}_AminoTypes_mafftAlign.fasta -out ${params.projtag}_AminoTypes_mafftAligned.fasta -keepheader -fasta -automated1 + rm ${params.projtag}_AminoTypes_mafftAlign.fasta + o-trim-uninformative-columns-from-alignment ${params.projtag}_AminoTypes_mafftAligned.fasta + mv ${params.projtag}_AminoTypes_mafftAligned.fasta-TRIMMED ./${params.projtag}_AminoTypes_Aligned_informativeonly.fasta + #entropy analysis + entropy-analysis ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta --amino-acid-sequences + #summarize entropy peaks + awk '{print \$2}' ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta-ENTROPY >> tmp_value.list + for x in \$(cat tmp_value.list) + do echo "\$x" + if [[ \$(echo ""\$x" > 0.0"|bc -l) -eq 1 ]]; + then echo dope >> above-0.0-.list + fi + if [[ \$(echo ""\$x" > 0.1"|bc -l) -eq 1 ]]; + then echo dope >> above-0.1-.list + fi + if [[ \$(echo ""\$x" > 0.2"|bc -l) -eq 1 ]]; + then echo dope >> above-0.2-.list + fi + if [[ \$(echo ""\$x" > 0.3"|bc -l) -eq 1 ]]; + then echo dope >> above-0.3-.list + fi + if [[ \$(echo ""\$x" > 0.4"|bc -l) -eq 1 ]]; + then echo dope >> above-0.4-.list + fi + if [[ \$(echo ""\$x" > 0.5"|bc -l) -eq 1 ]]; + then echo dope >> above-0.5-.list + fi + if [[ \$(echo ""\$x" > 0.6"|bc -l) -eq 1 ]]; + then echo dope >> above-0.6-.list + fi + if [[ \$(echo ""\$x" > 0.7"|bc -l) -eq 1 ]]; + then echo dope >> above-0.7-.list + fi + if [[ \$(echo ""\$x" > 0.8"|bc -l) -eq 1 ]]; + then echo dope >> above-0.8-.list + fi + if [[ \$(echo ""\$x" > 0.9"|bc -l) -eq 1 ]]; + then echo dope >> above-0.9-.list + fi + if [[ \$(echo ""\$x" > 1.0"|bc -l) -eq 1 ]]; + then echo dope >> above-1.0-.list + fi + if [[ \$(echo ""\$x" > 1.5"|bc -l) -eq 1 ]]; + then echo dope >> above-1.5-.list + fi + done + for z in above*.list; + do entrop=\$(echo \$z | awk -F "-" '{print \$2}') + echo ""\$entrop", "\$(wc -l \$z | awk '{print \$1}')"" >> ${params.projtag}_AminoType_entropy_breakdown.csv + done + rm above* + + """ + + } + if (!params.skipReadProcessing || !params.skipMerging ) { process combine_csv_DC { @@ -1366,11 +1518,13 @@ if (params.DataCheck || params.Analyze) { pre=\$(echo ${asvs} | awk -F ".fasta" '{print \$1}' ) mafft --thread ${task.cpus} --maxiterate 15000 --auto ${asvs} >\${pre}_ALN.fasta trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html + o-trim-uninformative-columns-from-alignment \${pre}_aln.fasta + mv \${pre}_aln.fasta-TRIMMED ./\${pre}_Aligned_informativeonly.fasta # Nucleotide_ModelTest - modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint + modeltest-ng -i \${pre}_Aligned_informativeonly.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint # Nucleotide_Phylogeny if [ "${params.iqCustomnt}" != "" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -t \${pre}_mt.tree -T auto ${params.iqCustomnt} + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq --redo -t \${pre}_mt.tree -T auto ${params.iqCustomnt} elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} diff --git a/vampirus_env.yml b/vampirus_env.yml index 95423f3..df49583 100644 --- a/vampirus_env.yml +++ b/vampirus_env.yml @@ -9,6 +9,7 @@ channels: - genomedk dependencies: - python=3.6 + - blast=2.11.0 - diamond=0.9.30 - fastqc=0.11.9 - fastp=0.20.1 From bcf09885f7afb34bbc872b8151b3dac0067e5c7d Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Tue, 23 Mar 2021 15:53:57 -0500 Subject: [PATCH 045/223] Added options for MED and processes for MED in Analyze loop --- vAMPirus.nf | 65 ++++++++++++++++++++++++++++++++++++++++++++++--- vampirus.config | 14 ++++++++++- 2 files changed, 74 insertions(+), 5 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index e37dc75..19750fd 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1128,6 +1128,7 @@ if (params.DataCheck || params.Analyze) { output: file("*AminoType_entropy_breakdown.csv") into amino_entro_csv + file ("*.png") into file("*AminoTypes*") into aminos script: @@ -1286,7 +1287,7 @@ if (params.DataCheck || params.Analyze) { if (!params.skipTaxonomy) { - process Nucleotide_Taxonomy_Inference { + process Nucleotide_Taxonomy_Inference { /////// editttt label 'high_cpus' @@ -1545,10 +1546,10 @@ if (params.DataCheck || params.Analyze) { } else { reads_vsearch5_ch - .into{ nuclFastas_forDiamond_asv_ch; nuclFastas_forCounts_asv_ch; nuclFastas_forphylogeny_asv; nuclFastas_forMatrix_asv_ch } + .into{ nuclFastas_forDiamond_asv_ch; nuclFastas_forCounts_asv_ch; nuclFastas_forphylogeny_asv; nuclFastas_forMatrix_asv_ch, asv_for_med } } - process ASV_Taxonomy_Inference { + process ASV_Taxonomy_Inference { /////// editttt label 'high_cpus' @@ -1901,6 +1902,40 @@ if (params.DataCheck || params.Analyze) { } } + if (params.asvMED) { + + process ASV_Minimum_Entropy_Decomposition { + + label 'low_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/ASV/MED", mode: "copy", overwrite: true + + input: + + file(asvs) from asv_for_med + + output: + + script: + """ + set +e + #alignment + mafft --thread ${task.cpus} --maxiterate 15000 --auto ${asvs} > ${params.projtag}_ASVs_mafftAlign.fasta + #trimming + trimal -in ${params.projtag}_ASVs_mafftAlign.fasta -out ${params.projtag}_ASVs_mafftAligned.fasta -keepheader -fasta -automated1 + rm ${params.projtag}_ASVs_mafftAlign.fasta + o-trim-uninformative-columns-from-alignment ${params.projtag}_ASVs_mafftAligned.fasta + mv ${params.projtag}_ASVs_mafftAligned.fasta-TRIMMED ./${params.projtag}_ASVs_Aligned_informativeonly.fasta + #entopy analysis + entropy-analysis ${params.projtag}_ASVs_Aligned_informativeonly.fasta + #Decomposition + oligotype ${params.projtag}_ASVs_Aligned_informativeonly.fasta ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY -o ${params.projtag}_asvMED_${params.asvC} -M 1 -c ${params.asvC} --skip-check-input --no-figures + + """ + } + + } + if (!params.skipAminoTyping) { process Translate_For_AminoTyping { @@ -1936,7 +1971,7 @@ if (params.DataCheck || params.Analyze) { output: tuple file("*.fasta"), file("${params.projtag}_AminoTypes.clstr"), file("${params.projtag}_AminoType_summary_map.csv"), file("${params.projtag}_clustered.gc") into ( supplementalfiles ) - file("${params.projtag}_AminoTypes_noTaxonomy.fasta") into ( aminotypesCounts, aminotypesMafft, aminotypesClustal, aminotypesBlast, aminotypesEmboss ) + file("${params.projtag}_AminoTypes_noTaxonomy.fasta") into ( aminotypesCounts, aminotypesMafft, aminotypesClustal, aminotypesBlast, aminotypesEmboss, aminos_for_med ) script: """ @@ -2301,6 +2336,28 @@ if (params.DataCheck || params.Analyze) { } } + if (params.aminoMED) { + + process AminoType_Minimum_Entropy_Decomposition { + + label 'low_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/ASV/MED", mode: "copy", overwrite: true + + input: + + file(aminos) from aminos_for_med + + output: + + script: + """ + + """ + } + + } + if (params.pcASV) { // ASV_nucl -> ASV_aa -> clusteraa by %id with ch-hit -> extract representative nucl sequences to generate new OTU file process Translation_For_pcASV_Generation { diff --git a/vampirus.config b/vampirus.config index db310e0..aef6beb 100644 --- a/vampirus.config +++ b/vampirus.config @@ -68,6 +68,13 @@ params { // Minimum length of amino acid translation to be considered during protein clustered ASV (pcASV) generation. Recommended to put this at the expected amino acid sequence length based on your maximum read length (e.g. if maxLen="420", then minAA should be 420/3 so 140) minAA="140" + // Minimum Entropy Decomposition (MED) parameters for clustering (https://merenlab.org/2012/05/11/oligotyping-pipeline-explained/) + + // If you plan to do MED on ASVs using the option "--asvMED" you can set here the number of positions for oligotyping to take into consideration + asvC="" + // If you plan to do MED on ASVs using the option "--aminoMED" you can set here the number of positions for oligotyping to take into consideration + aminoC="" + // Counts table generation parameters // Percent similarity to use for ASV/cASV counts table generation with vsearch @@ -141,11 +148,16 @@ params { // Manadotory arguments Analyze=false DataCheck=false -// Clustering options +// Non-Mandatory Clustering options // Cluster nucleotide sequences (ncASVs) ncASV = false // Cluster by aminoacid translations and generate protein-based OTUs (pcASVs) pcASV = false + // Generate virus types with MED of ASV sequences + asvMED = false + // Generate virus types with MED of ASV sequences + aminoMED = false + // Skip options // Skip all Read Processing steps skipReadProcessing=false From 38a05d797ded1dda08806e1df576b79cfe4f915e Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Tue, 23 Mar 2021 17:23:40 -0500 Subject: [PATCH 046/223] added lines for csv generation --- vAMPirus.nf | 14 +++++++------- 1 file changed, 7 insertions(+), 7 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 19750fd..31898f9 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1052,6 +1052,7 @@ if (params.DataCheck || params.Analyze) { output: file("*_ASV_entropy_breakdown.csv") into asv_entro_csv + file("") file("*ASV*") into entrop script: @@ -1067,7 +1068,7 @@ if (params.DataCheck || params.Analyze) { #entopy analysis entropy-analysis ${params.projtag}_ASVs_Aligned_informativeonly.fasta #summarize entropy peaks - awk '{print \$2}' testaligned.fasta-ENTROPY >> tmp_value.list + awk '{print \$2}' ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY >> tmp_value.list for x in \$(cat tmp_value.list) do echo "\$x" if [[ \$(echo ""\$x" > 0.0"|bc -l) -eq 1 ]]; @@ -1107,6 +1108,7 @@ if (params.DataCheck || params.Analyze) { then echo dope >> above-1.5-.list fi done + echo "Base_position, Shannon's_Entropy" >> ${params.projtag}_ASV_entropy_breakdown.csv for z in above*.list; do entrop=\$(echo \$z | awk -F "-" '{print \$2}') echo ""\$entrop", "\$(wc -l \$z | awk '{print \$1}')"" >> ${params.projtag}_ASV_entropy_breakdown.csv @@ -1126,13 +1128,12 @@ if (params.DataCheck || params.Analyze) { file(aminos) from amino_med output: - file("*AminoType_entropy_breakdown.csv") into amino_entro_csv file ("*.png") into file("*AminoTypes*") into aminos script: - """ + """ #alignment mafft --thread ${task.cpus} --maxiterate 15000 --auto ${aminos} > ${params.projtag}_AminoTypes_mafftAlign.fasta #trimming @@ -1183,14 +1184,13 @@ if (params.DataCheck || params.Analyze) { then echo dope >> above-1.5-.list fi done + echo "Base_position, Shannon's_Entropy" >> ${params.projtag}_AminoType_entropy_breakdown.csv for z in above*.list; do entrop=\$(echo \$z | awk -F "-" '{print \$2}') echo ""\$entrop", "\$(wc -l \$z | awk '{print \$1}')"" >> ${params.projtag}_AminoType_entropy_breakdown.csv done rm above* - - """ - + """ } if (!params.skipReadProcessing || !params.skipMerging ) { @@ -1930,7 +1930,7 @@ if (params.DataCheck || params.Analyze) { entropy-analysis ${params.projtag}_ASVs_Aligned_informativeonly.fasta #Decomposition oligotype ${params.projtag}_ASVs_Aligned_informativeonly.fasta ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY -o ${params.projtag}_asvMED_${params.asvC} -M 1 -c ${params.asvC} --skip-check-input --no-figures - + """ } From 960fccaf1d2a1c60756a31c91cb15d3736037cd9 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Tue, 23 Mar 2021 17:54:21 -0500 Subject: [PATCH 047/223] edits --- vAMPirus.nf | 14 ++++++++++++-- 1 file changed, 12 insertions(+), 2 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 31898f9..54bb6ed 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1108,12 +1108,18 @@ if (params.DataCheck || params.Analyze) { then echo dope >> above-1.5-.list fi done - echo "Base_position, Shannon's_Entropy" >> ${params.projtag}_ASV_entropy_breakdown.csv + echo "Entropy,Peaks_above" >> ${params.projtag}_ASV_entropy_breakdown.csv for z in above*.list; do entrop=\$(echo \$z | awk -F "-" '{print \$2}') echo ""\$entrop", "\$(wc -l \$z | awk '{print \$1}')"" >> ${params.projtag}_ASV_entropy_breakdown.csv done rm above* + + mv ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY ./tmp.fasta + cat "Base_position Shannons_Entropy" >> ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY + cat tmp.fasta >> ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY + rm tmp.fasta + """ } @@ -1184,12 +1190,16 @@ if (params.DataCheck || params.Analyze) { then echo dope >> above-1.5-.list fi done - echo "Base_position, Shannon's_Entropy" >> ${params.projtag}_AminoType_entropy_breakdown.csv + echo "Entropy,Peaks_above" >> ${params.projtag}_AminoType_entropy_breakdown.csv for z in above*.list; do entrop=\$(echo \$z | awk -F "-" '{print \$2}') echo ""\$entrop", "\$(wc -l \$z | awk '{print \$1}')"" >> ${params.projtag}_AminoType_entropy_breakdown.csv done rm above* + mv ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta-ENTROPY ./tmp.fasta + cat "Base_position Shannons_Entropy" >> ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta-ENTROPY + cat tmp.fasta >> ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta-ENTROPY + rm tmp.fasta """ } From 2368c440c6cb1ebac5f110aaae7227a7283037aa Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Tue, 23 Mar 2021 15:54:30 -0700 Subject: [PATCH 048/223] Add MED plots and tables --- bin/vAMPirus_DC_Report.Rmd | 66 ++++++++++++++++++++++++++++++++++++++ 1 file changed, 66 insertions(+) diff --git a/bin/vAMPirus_DC_Report.Rmd b/bin/vAMPirus_DC_Report.Rmd index 0914be4..7a6b8f1 100644 --- a/bin/vAMPirus_DC_Report.Rmd +++ b/bin/vAMPirus_DC_Report.Rmd @@ -398,6 +398,72 @@ pnp ``` NOTE: The "1" represents the number of AminoTypes which are unique amino acid sequences in your dataset
+ + +### ASVs MED + +
+ +```{r med_asv, echo=FALSE} +med_asv=read.delim(paste(params$projtag, sep="","_ASVs_Aligned_informativeonly.fasta-ENTROPY"), sep = "\t", header=TRUE) + +mplot=plot_ly(med_asv, x=~Base_position, y=~Shannons_Entropy,type="bar", name="Entropy", opacity = .8, line=list(color='#088da5'), + hovertemplate = paste('Base position: %{x}','
Entropy: %{y}','')) +mplot<- mplot %>% layout(xaxis=list(title="Base position")) +mplot<- mplot %>% layout(yaxis=list(title="Entropy")) +mplot<- mplot %>% config(toImageButtonOptions=list(format='svg',filename='ASV_MED', height= 500, width= 800, scale= 1), displaylogo = FALSE) +mplot +``` +
+ +### ASVs Peaks Range Information + +
+ +```{r med_asv_peak, echo=FALSE} +med_asv_csv=read.csv(paste(params$projtag, sep="", "_ASV_entropy_breakdown.csv"), header=TRUE) +paged_table(med_asv_csv, options = list(rows.print = 10)) +``` +
+ +### AminoTypes MED + +
+ +```{r med_amino, echo=FALSE} +med_amino=read.delim(paste(params$projtag, sep="","_AminoTypes_Aligned_informativeonly.fasta-ENTROPY"), sep = "\t", header=TRUE) + +mplot=plot_ly(med_amino, x=~Base_position, y=~Shannons_Entropy,type="bar", name="Entropy", opacity = .8, line=list(color='#088da5'), + hovertemplate = paste('Base position: %{x}','
Entropy: %{y}','')) +mplot<- mplot %>% layout(xaxis=list(title="Base position")) +mplot<- mplot %>% layout(yaxis=list(title="Entropy")) +mplot<- mplot %>% config(toImageButtonOptions=list(format='svg',filename='AminoTypes_MED', height= 500, width= 800, scale= 1), displaylogo = FALSE) +mplot +``` +
+ +### AminoTypes Peaks Range Information + +
+ +```{r med_amino_peak, echo=FALSE} +med_amino_csv=read.csv(paste(params$projtag, sep="", "_AminoType_entropy_breakdown.csv"), header=TRUE) +paged_table(med_amino_csv, options = list(rows.print = 10)) +``` +
+ +### ASVs vs AminoTypes + +
+ +```{r med_comparison, echo=FALSE} +mplotbox <- plot_ly(med_asv, y=~Shannons_Entropy, type="box", boxmean='sd', boxpoints="outlier", jitter=0.1, pointpos=0, name="ASVs") +mplotbox<- mplotbox %>% add_trace(med_amino, y=~Shannons_Entropy, type="box", boxmean='sd', boxpoints="outliers", jitter=0.1, pointpos=0, name="AminoTypes") +mplotbox<- mplotbox %>% layout(legend = list(y=.5)) +mplotbox<- mplotbox %>% config(toImageButtonOptions=list(format='svg',filename='ASV_vs_AminoTypes', height= 500, width= 800, scale= 1), displaylogo = FALSE) +mplotbox +``` +



From 78ff79f081b8f41d7fe8d19646a88493a3c29c90 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Tue, 23 Mar 2021 18:06:06 -0500 Subject: [PATCH 049/223] connected datacheck med channels --- vAMPirus.nf | 7 +++---- 1 file changed, 3 insertions(+), 4 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 54bb6ed..c231e5d 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1052,7 +1052,7 @@ if (params.DataCheck || params.Analyze) { output: file("*_ASV_entropy_breakdown.csv") into asv_entro_csv - file("") + file("Aligned_informativeonly.fasta-ENTROPY") into asv_entropy file("*ASV*") into entrop script: @@ -1114,7 +1114,6 @@ if (params.DataCheck || params.Analyze) { echo ""\$entrop", "\$(wc -l \$z | awk '{print \$1}')"" >> ${params.projtag}_ASV_entropy_breakdown.csv done rm above* - mv ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY ./tmp.fasta cat "Base_position Shannons_Entropy" >> ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY cat tmp.fasta >> ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY @@ -1135,7 +1134,7 @@ if (params.DataCheck || params.Analyze) { output: file("*AminoType_entropy_breakdown.csv") into amino_entro_csv - file ("*.png") into + file ("*Aligned_informativeonly.fasta-ENTROPY") into amino_entropy file("*AminoTypes*") into aminos script: @@ -1238,7 +1237,7 @@ if (params.DataCheck || params.Analyze) { } report_dc_in = Channel.create() - fastp_csv_dc.mix( reads_per_sample_preFilt, reads_per_sample_postFilt, prefilt_basefreq, postFilt_basefreq, prefilt_qualityscore, postFilt_qualityscore, prefilt_gccontent, postFilt_gccontent, prefilt_averagequality, postFilt_averagequality, prefilt_length, postFilt_length, number_per_percent_nucl_plot, number_per_percent_prot_plot + fastp_csv_dc.mix( reads_per_sample_preFilt, reads_per_sample_postFilt, prefilt_basefreq, postFilt_basefreq, prefilt_qualityscore, postFilt_qualityscore, prefilt_gccontent, postFilt_gccontent, prefilt_averagequality, postFilt_averagequality, prefilt_length, postFilt_length, number_per_percent_nucl_plot, number_per_percent_prot_plot, amino_entro_csv, amino_entropy, asv_entro_csv, asv_entropy ).into(report_dc_in) process Report_DataCheck { From 70dc57f78d4b0c181691d01e211d65af749c7826 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Tue, 23 Mar 2021 18:17:15 -0500 Subject: [PATCH 050/223] conda replaced --- vampirus_startup.sh | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/vampirus_startup.sh b/vampirus_startup.sh index 13c7e09..5df8e89 100644 --- a/vampirus_startup.sh +++ b/vampirus_startup.sh @@ -178,8 +178,8 @@ else fi echo "Downloading oligotyping program" -conda init && source activate vAMPirus -pip install oligotyping +conda init && conda activate vAMPirus +pip install oligotyping conda deactivate echo "-------------------------------------------------------------------------------- Conda check/install done" From c8127f8bf82ca0bdbfb1094928d75be294234f56 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Tue, 23 Mar 2021 18:19:22 -0500 Subject: [PATCH 051/223] changed semicolon --- vAMPirus.nf | 5 ++--- 1 file changed, 2 insertions(+), 3 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index c231e5d..8bef557 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1237,8 +1237,7 @@ if (params.DataCheck || params.Analyze) { } report_dc_in = Channel.create() - fastp_csv_dc.mix( reads_per_sample_preFilt, reads_per_sample_postFilt, prefilt_basefreq, postFilt_basefreq, prefilt_qualityscore, postFilt_qualityscore, prefilt_gccontent, postFilt_gccontent, prefilt_averagequality, postFilt_averagequality, prefilt_length, postFilt_length, number_per_percent_nucl_plot, number_per_percent_prot_plot, amino_entro_csv, amino_entropy, asv_entro_csv, asv_entropy - ).into(report_dc_in) + fastp_csv_dc.mix( reads_per_sample_preFilt, reads_per_sample_postFilt, prefilt_basefreq, postFilt_basefreq, prefilt_qualityscore, postFilt_qualityscore, prefilt_gccontent, postFilt_gccontent, prefilt_averagequality, postFilt_averagequality, prefilt_length, postFilt_length, number_per_percent_nucl_plot, number_per_percent_prot_plot, amino_entro_csv, amino_entropy, asv_entro_csv, asv_entropy).into(report_dc_in) process Report_DataCheck { @@ -1555,7 +1554,7 @@ if (params.DataCheck || params.Analyze) { } else { reads_vsearch5_ch - .into{ nuclFastas_forDiamond_asv_ch; nuclFastas_forCounts_asv_ch; nuclFastas_forphylogeny_asv; nuclFastas_forMatrix_asv_ch, asv_for_med } + .into{ nuclFastas_forDiamond_asv_ch; nuclFastas_forCounts_asv_ch; nuclFastas_forphylogeny_asv; nuclFastas_forMatrix_asv_ch; asv_for_med } } process ASV_Taxonomy_Inference { /////// editttt From 6ab2a17498899018bc2a25a157a0e24b38fbda5d Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Tue, 23 Mar 2021 18:26:13 -0500 Subject: [PATCH 052/223] editted output file nae --- vAMPirus.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 8bef557..4ba62d7 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -946,7 +946,7 @@ if (params.DataCheck || params.Analyze) { output: file("number_per_percentage_prot.csv") into number_per_percent_prot_plot - file("*pcASV100*") into amino_med + file("*pcASV1.0_noTaxonomy*") into amino_med script: // add awk script to count seqs """ From 4311757ccd0766ec09840550b19bd2eb69128eeb Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Tue, 23 Mar 2021 18:41:41 -0500 Subject: [PATCH 053/223] editted errors in datacheck --- vAMPirus.nf | 6 +++--- 1 file changed, 3 insertions(+), 3 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 4ba62d7..81ce02b 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -946,7 +946,7 @@ if (params.DataCheck || params.Analyze) { output: file("number_per_percentage_prot.csv") into number_per_percent_prot_plot - file("*pcASV1.0_noTaxonomy*") into amino_med + file("*aminoacid_pcASV1.0_noTaxonomy.fasta") into amino_med script: // add awk script to count seqs """ @@ -1115,7 +1115,7 @@ if (params.DataCheck || params.Analyze) { done rm above* mv ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY ./tmp.fasta - cat "Base_position Shannons_Entropy" >> ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY + echo "Base_position Shannons_Entropy" >> ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY cat tmp.fasta >> ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY rm tmp.fasta @@ -1196,7 +1196,7 @@ if (params.DataCheck || params.Analyze) { done rm above* mv ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta-ENTROPY ./tmp.fasta - cat "Base_position Shannons_Entropy" >> ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta-ENTROPY + echo "Base_position Shannons_Entropy" >> ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta-ENTROPY cat tmp.fasta >> ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta-ENTROPY rm tmp.fasta """ From 40e1807c086ae872f2c2646ff58912c4ddf7d069 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Tue, 23 Mar 2021 17:20:29 -0700 Subject: [PATCH 054/223] Fix figures --- bin/vAMPirus_DC_Report.Rmd | 14 +++++++------- 1 file changed, 7 insertions(+), 7 deletions(-) diff --git a/bin/vAMPirus_DC_Report.Rmd b/bin/vAMPirus_DC_Report.Rmd index 7a6b8f1..e7ab7cf 100644 --- a/bin/vAMPirus_DC_Report.Rmd +++ b/bin/vAMPirus_DC_Report.Rmd @@ -407,7 +407,7 @@ NOTE: The "1" represents the number of AminoTypes which are unique amino acid se ```{r med_asv, echo=FALSE} med_asv=read.delim(paste(params$projtag, sep="","_ASVs_Aligned_informativeonly.fasta-ENTROPY"), sep = "\t", header=TRUE) -mplot=plot_ly(med_asv, x=~Base_position, y=~Shannons_Entropy,type="bar", name="Entropy", opacity = .8, line=list(color='#088da5'), +mplot=plot_ly(med_asv, x=med_asv$Base_position, y=med_asv$Shannons_Entropy,type="bar", name="Entropy", opacity = .8, line=list(color='#088da5'), hovertemplate = paste('Base position: %{x}','
Entropy: %{y}','')) mplot<- mplot %>% layout(xaxis=list(title="Base position")) mplot<- mplot %>% layout(yaxis=list(title="Entropy")) @@ -416,7 +416,7 @@ mplot ```
-### ASVs Peaks Range Information +### ASVs Peaks Range Information
@@ -433,7 +433,7 @@ paged_table(med_asv_csv, options = list(rows.print = 10)) ```{r med_amino, echo=FALSE} med_amino=read.delim(paste(params$projtag, sep="","_AminoTypes_Aligned_informativeonly.fasta-ENTROPY"), sep = "\t", header=TRUE) -mplot=plot_ly(med_amino, x=~Base_position, y=~Shannons_Entropy,type="bar", name="Entropy", opacity = .8, line=list(color='#088da5'), +mplot=plot_ly(med_amino, x=med_amino$Base_position, y=med_amino$Shannons_Entropy,type="bar", name="Entropy", opacity = .8, line=list(color='#088da5'), hovertemplate = paste('Base position: %{x}','
Entropy: %{y}','')) mplot<- mplot %>% layout(xaxis=list(title="Base position")) mplot<- mplot %>% layout(yaxis=list(title="Entropy")) @@ -442,7 +442,7 @@ mplot ```
-### AminoTypes Peaks Range Information +### AminoTypes Peaks Range Information
@@ -452,13 +452,13 @@ paged_table(med_amino_csv, options = list(rows.print = 10)) ```
-### ASVs vs AminoTypes +### ASVs vs AminoTypes
```{r med_comparison, echo=FALSE} -mplotbox <- plot_ly(med_asv, y=~Shannons_Entropy, type="box", boxmean='sd', boxpoints="outlier", jitter=0.1, pointpos=0, name="ASVs") -mplotbox<- mplotbox %>% add_trace(med_amino, y=~Shannons_Entropy, type="box", boxmean='sd', boxpoints="outliers", jitter=0.1, pointpos=0, name="AminoTypes") +mplotbox <- plot_ly(med_asv, y=med_asv$Shannons_Entropy, type="box", boxmean='sd', boxpoints="outlier", jitter=0.1, pointpos=0, name="ASVs") +mplotbox<- mplotbox %>% add_trace(med_amino, y=med_amino$Shannons_Entropy, type="box", boxmean='sd', boxpoints="outliers", jitter=0.1, pointpos=0, name="AminoTypes") mplotbox<- mplotbox %>% layout(legend = list(y=.5)) mplotbox<- mplotbox %>% config(toImageButtonOptions=list(format='svg',filename='ASV_vs_AminoTypes', height= 500, width= 800, scale= 1), displaylogo = FALSE) mplotbox From 91c744cb3865cb4086856705ad96869a2e041675 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Tue, 23 Mar 2021 17:24:08 -0700 Subject: [PATCH 055/223] Add missing * --- vAMPirus.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 81ce02b..3a817ce 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1052,7 +1052,7 @@ if (params.DataCheck || params.Analyze) { output: file("*_ASV_entropy_breakdown.csv") into asv_entro_csv - file("Aligned_informativeonly.fasta-ENTROPY") into asv_entropy + file("*Aligned_informativeonly.fasta-ENTROPY") into asv_entropy file("*ASV*") into entrop script: From cc283b1849c3b15e443f69f50de541a94026bcd5 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Tue, 23 Mar 2021 17:28:27 -0700 Subject: [PATCH 056/223] Fix name --- vAMPirus.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 3a817ce..24df4cd 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -3229,7 +3229,7 @@ if (params.DataCheck) { "---------------------------------------------------------------------------------" \ + "\n\033[0;32mDone! Open the following reports in your browser\033[0m" \ + "\n\033[0;32mPipeline performance report: ${params.workingdir}/${params.outdir}/${params.tracedir}/vampirus_report.html\033[0m" \ - + "\n\033[0;32mvAMPirus --DataCheck interactive report: ${params.workingdir}/${params.outdir}/DataCheck/*.hmtl\033[0m" \ + + "\n\033[0;32mvAMPirus --DataCheck interactive report: ${params.workingdir}/${params.outdir}/DataCheck/Report/*.hmtl\033[0m" \ : \ "---------------------------------------------------------------------------------" \ + "\n\033[0;31mSomething went wrong. Check error message below and/or log files.\033[0m" ) From d66bba98511f98d379dae9f18003c4d1aea17512 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Tue, 23 Mar 2021 17:37:36 -0700 Subject: [PATCH 057/223] Edit space --- bin/vAMPirus_DC_Report.Rmd | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/bin/vAMPirus_DC_Report.Rmd b/bin/vAMPirus_DC_Report.Rmd index e7ab7cf..6ee6df3 100644 --- a/bin/vAMPirus_DC_Report.Rmd +++ b/bin/vAMPirus_DC_Report.Rmd @@ -407,7 +407,7 @@ NOTE: The "1" represents the number of AminoTypes which are unique amino acid se ```{r med_asv, echo=FALSE} med_asv=read.delim(paste(params$projtag, sep="","_ASVs_Aligned_informativeonly.fasta-ENTROPY"), sep = "\t", header=TRUE) -mplot=plot_ly(med_asv, x=med_asv$Base_position, y=med_asv$Shannons_Entropy,type="bar", name="Entropy", opacity = .8, line=list(color='#088da5'), +mplot=plot_ly(med_asv, x=med_asv$Base_position, y=med_asv$Shannons_Entropy, type="bar", name="Entropy", opacity = .8, line=list(color='#088da5'), hovertemplate = paste('Base position: %{x}','
Entropy: %{y}','')) mplot<- mplot %>% layout(xaxis=list(title="Base position")) mplot<- mplot %>% layout(yaxis=list(title="Entropy")) @@ -433,7 +433,7 @@ paged_table(med_asv_csv, options = list(rows.print = 10)) ```{r med_amino, echo=FALSE} med_amino=read.delim(paste(params$projtag, sep="","_AminoTypes_Aligned_informativeonly.fasta-ENTROPY"), sep = "\t", header=TRUE) -mplot=plot_ly(med_amino, x=med_amino$Base_position, y=med_amino$Shannons_Entropy,type="bar", name="Entropy", opacity = .8, line=list(color='#088da5'), +mplot=plot_ly(med_amino, x=med_amino$Base_position, y=med_amino$Shannons_Entropy, type="bar", name="Entropy", opacity = .8, line=list(color='#088da5'), hovertemplate = paste('Base position: %{x}','
Entropy: %{y}','')) mplot<- mplot %>% layout(xaxis=list(title="Base position")) mplot<- mplot %>% layout(yaxis=list(title="Entropy")) From a1608c32e354e09954e9a10f820fe27da94a6c39 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Tue, 23 Mar 2021 17:39:04 -0700 Subject: [PATCH 058/223] Add files for figure --- bin/vAMPirus_DC_Report.Rmd | 3 +++ 1 file changed, 3 insertions(+) diff --git a/bin/vAMPirus_DC_Report.Rmd b/bin/vAMPirus_DC_Report.Rmd index 6ee6df3..0578dce 100644 --- a/bin/vAMPirus_DC_Report.Rmd +++ b/bin/vAMPirus_DC_Report.Rmd @@ -457,6 +457,9 @@ paged_table(med_amino_csv, options = list(rows.print = 10))
```{r med_comparison, echo=FALSE} +med_asv=read.delim(paste(params$projtag, sep="","_ASVs_Aligned_informativeonly.fasta-ENTROPY"), sep = "\t", header=TRUE) +med_amino=read.delim(paste(params$projtag, sep="","_AminoTypes_Aligned_informativeonly.fasta-ENTROPY"), sep = "\t", header=TRUE) + mplotbox <- plot_ly(med_asv, y=med_asv$Shannons_Entropy, type="box", boxmean='sd', boxpoints="outlier", jitter=0.1, pointpos=0, name="ASVs") mplotbox<- mplotbox %>% add_trace(med_amino, y=med_amino$Shannons_Entropy, type="box", boxmean='sd', boxpoints="outliers", jitter=0.1, pointpos=0, name="AminoTypes") mplotbox<- mplotbox %>% layout(legend = list(y=.5)) From 5dc272e746a47d74bd5f91e573662781f5cea691 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Wed, 24 Mar 2021 08:28:09 -0500 Subject: [PATCH 059/223] fixed outputname --- vAMPirus.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 81ce02b..3a817ce 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1052,7 +1052,7 @@ if (params.DataCheck || params.Analyze) { output: file("*_ASV_entropy_breakdown.csv") into asv_entro_csv - file("Aligned_informativeonly.fasta-ENTROPY") into asv_entropy + file("*Aligned_informativeonly.fasta-ENTROPY") into asv_entropy file("*ASV*") into entrop script: From 8dfbd30b592c8bd1babb57a645d6c2adbb1b6215 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Fri, 26 Mar 2021 17:03:18 -0500 Subject: [PATCH 060/223] Added MED to Analyze --- vAMPirus.nf | 194 ++++++++++++++++++++++++++++++++++++++++++++++++---- 1 file changed, 182 insertions(+), 12 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 24df4cd..7e339ee 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -720,7 +720,7 @@ if (params.DataCheck || params.Analyze) { } - process Compile_Reads { + process Filtering_Prep1 { label 'low_cpus' @@ -739,7 +739,7 @@ if (params.DataCheck || params.Analyze) { """ } - process Compile_Names { + process Filtering_Prep2 { label 'low_cpus' @@ -1805,7 +1805,7 @@ if (params.DataCheck || params.Analyze) { output: tuple file("*_counts.csv"), file("*_counts.biome") into counts_vsearch_asv - file("*_ASV*counts.csv") into asv_counts_plots + file("*_ASV*counts.csv") into (asv_counts_plots, asvcount_med) script: """ @@ -1881,7 +1881,7 @@ if (params.DataCheck || params.Analyze) { output: tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into align_results_asv - file("*iq.treefile") into nucl_phyl_plot_asv + file("*iq.treefile") into (nucl_phyl_plot_asv, asvphy_med) script: """ @@ -1916,17 +1916,18 @@ if (params.DataCheck || params.Analyze) { label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/ASV/MED", mode: "copy", overwrite: true - + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/ASV/${params.projtag}_asvMED_${params.asvC}", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/ASV/${params.projtag}_asvMED_${params.asvC}/OLIGO-REPRESENTATIVES", mode: "copy", overwrite: true input: file(asvs) from asv_for_med output: + file("*_ASV_Grouping.csv") into asvgroupscsv + file("${params.projtag}_ASV_group_reps_aligned.fasta") into groupreps script: """ - set +e #alignment mafft --thread ${task.cpus} --maxiterate 15000 --auto ${asvs} > ${params.projtag}_ASVs_mafftAlign.fasta #trimming @@ -1937,11 +1938,98 @@ if (params.DataCheck || params.Analyze) { #entopy analysis entropy-analysis ${params.projtag}_ASVs_Aligned_informativeonly.fasta #Decomposition - oligotype ${params.projtag}_ASVs_Aligned_informativeonly.fasta ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY -o ${params.projtag}_asvMED_${params.asvC} -M 1 -c ${params.asvC} --skip-check-input --no-figures + oligotype ${params.projtag}_ASVs_Aligned_informativeonly.fasta ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY -o ${params.projtag}_asvMED_${params.asvC} -M 1 -c ${params.asvC} -N ${task.cpus} --skip-check-input --no-figures --skip-gen-html + #generatemaps + cd ./${params.projtag}_asvMED_${params.asvC}/OLIGO-REPRESENTATIVES/ + echo "ASV,Group,IDPattern" + j=1 + for x in *_unique; + do gid=$(echo \$x | awk -F "_" '{print \$1}') + uni=$(echo \$x | awk -F ""\${gid}"_" '{print \$2}' | awk -F "_uni" '{print \$1}') + grep ">" "\$gid"_"\$uni" | awk -F ">" '{print \$2}' > asv.list + seqtk subseq ${asvs} asv.list > Group"\${j}"_sequences.fasta + for z in $( cat asv.list) + do echo ""\$z",Group"\$gid","\$uni"" >> ${params.projtag}_ASV_Grouping.csv + done + rm asv.list + echo "Group\${j}" >> ${params.projtag}_group_reps_aligned.fasta + echo "\$uni" > group.list + seqtk subseq ../OLIGO-REPRESENTATIVES.fasta group.list > group.fasta + tail -1 group.fasta >> ${params.projtag}_group_reps_aligned.fasta + mv "\$gid"_"\$uni" ./Group"\$j"_"\$uni"_aligned.fasta + mv "\$gid"_"\$uni"_unique ./Group"\$j"_"\$uni"_unqiues_aligned.fasta + rm "\$gid"*.cPickle + j=\$((\$j+1)) + done + mv ${params.projtag}_ASV_Grouping.csv ../../ + mv ${params.projtag}_ASV_group_reps_aligned.fasta ../../ + cd .. """ } + process ASV_MED_Reps_phylogeny { + + label 'low_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/MED/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*ASV*mt*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/MED/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*ASV*iq*' + + input: + file(reps) from groupreps + + output: + tuple file("*_Aminotype_Group_Reps") into align_results_asvmed + file("*iq.treefile") into asv_group_rep_tree + + script: + """ + # Protein_ModelTest + modeltest-ng -i ${reps} -p ${task.cpus} -o ${params.protag}_ASV_Group_Reps_mt -d aa -s 203 --disable-checkpoint + + # Protein_Phylogeny + if [ "${params.iqCustomaa}" != "" ];then + iqtree -s ${reps} --prefix ${params.protag}_ASV_Group_Reps_iq --redo -T auto ${params.iqCustomaa} + + elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then + mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') + iqtree -s ${reps} --prefix ${params.protag}_ASV_Group_Reps_iq -m \${mod} --redo -nt auto -b ${params.boots} + + elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then + mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') + iqtree -s ${reps} --prefix ${params.protag}_ASV_Group_Reps_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni + + elif [ "${params.nonparametric}" != "false" ];then + iqtree -s ${reps} --prefix ${params.protag}_ASV_Group_Reps_iq -m MFP --redo -nt auto -b ${params.boots} + + elif [ "${params.parametric}" != "false" ];then + iqtree -s ${reps} --prefix ${params.protag}_ASV_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + + else + iqtree -s ${reps} --prefix ${params.protag}_ASV_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + fi + """ + } + + process Adding_ASV_MED_Info { + + label 'low_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/MED/Phylogeny", mode: "copy", overwrite: true + + input: + file(counts) from asvcount_med + file(tree) from asvphy_med + file(map) from asvgroupscsv + output: + + + script: + """ + + + """ + } } if (!params.skipAminoTyping) { @@ -1978,8 +2066,9 @@ if (params.DataCheck || params.Analyze) { file(asvs) from asvaminocheck output: - tuple file("*.fasta"), file("${params.projtag}_AminoTypes.clstr"), file("${params.projtag}_AminoType_summary_map.csv"), file("${params.projtag}_clustered.gc") into ( supplementalfiles ) + tuple file("*.fasta"), file("${params.projtag}_AminoTypes.clstr"), file("${params.projtag}_clustered.gc") into ( supplementalfiles ) file("${params.projtag}_AminoTypes_noTaxonomy.fasta") into ( aminotypesCounts, aminotypesMafft, aminotypesClustal, aminotypesBlast, aminotypesEmboss, aminos_for_med ) + file("${params.projtag}_AminoType_summary_map.csv") into aminomapmed script: """ @@ -2171,7 +2260,7 @@ if (params.DataCheck || params.Analyze) { virus="NO" gene="HIT" echo ">AminoType\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) + j=\$((\$\$1)) echo "\$s done." fi else @@ -2350,17 +2439,98 @@ if (params.DataCheck || params.Analyze) { label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/ASV/MED", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes/MED", mode: "copy", overwrite: true input: - file(aminos) from aminos_for_med output: + file("*_AminoType_Grouping.csv") into atygroupscsv + file("${params.projtag}_AminoType_group_reps_aligned.fasta") into atygroupreps script: """ + #alignment + mafft --thread ${task.cpus} --maxiterate 15000 --auto ${aminos} > ${params.projtag}_AminoTypes_mafftAlign.fasta + #trimming + trimal -in ${params.projtag}_AmminoTypes_mafftAlign.fasta -out ${params.projtag}_AminoTypes_mafftAligned.fasta -keepheader -fasta -automated1 + rm ${params.projtag}_AminoTypes_mafftAlign.fasta + o-trim-uninformative-columns-from-alignment ${params.projtag}_AminoTypes_mafftAligned.fasta + mv ${params.projtag}_AminoTypes_mafftAligned.fasta-TRIMMED ./${params.projtag}_AminoTypes_Aligned_informativeonly.fasta + #entopy analysis + entropy-analysis ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta + #Decomposition + oligotype ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta-ENTROPY -o ${params.projtag}_asvMED_${params.asvC} -M 1 -c ${params.asvC} -N ${task.cpus} --skip-check-input --no-figures --skip-gen-html + #generatemaps + cd ./${params.projtag}_asvMED_${params.asvC}/OLIGO-REPRESENTATIVES/ + echo "AminoType,Group,IDPattern" + j=1 + for x in *_unique; + do gid=$(echo \$x | awk -F "_" '{print \$1}') + uni=$(echo \$x | awk -F ""\${gid}"_" '{print \$2}' | awk -F "_uni" '{print \$1}') + grep ">" "\$gid"_"\$uni" | awk -F ">" '{print \$2}' > asv.list + seqtk subseq ${aminos} asv.list > Group"\${j}"_sequences.fasta + for z in $( cat asv.list) + do echo ""\$z",Group"\$gid","\$uni"" >> ${params.projtag}_AminoType_Grouping.csv + + done + rm asv.list + echo "Group\${j}" >> ${params.projtag}_group_reps_aligned.fasta + echo "\$uni" > group.list + seqtk subseq ../OLIGO-REPRESENTATIVES.fasta group.list > group.fasta + tail -1 group.fasta >> ${params.projtag}_group_reps_aligned.fasta + mv "\$gid"_"\$uni" ./Group"\$j"_"\$uni"_aligned.fasta + mv "\$gid"_"\$uni"_unique ./Group"\$j"_"\$uni"_unqiues_aligned.fasta + rm "\$gid"*.cPickle + j=\$((\$j+1)) + done + mv ${params.projtag}_AminoType_Grouping.csv ../../ + mv ${params.projtag}_AminoType_group_reps_aligned.fasta ../../ + cd .. + + """ + } + + process AminoType_MED_Reps_phylogeny { + + label 'low_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/MED/Phylogeny/Modeltest", mode: "copy", overwrite: true, pattern: '*mt*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/MED/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*iq*' + + input: + file(reps) from atygroupreps + + output: + tuple file("*_Aminotype_Group_Reps") into align_results_aminmed + file("*iq.treefile") into amino_group_rep_tree + + script: + """ + # Protein_ModelTest + modeltest-ng -i ${reps} -p ${task.cpus} -o ${params.protag}_AminoType_Group_Reps_mt -d aa -s 203 --disable-checkpoint + + # Protein_Phylogeny + if [ "${params.iqCustomaa}" != "" ];then + iqtree -s ${reps} --prefix ${params.protag}_AminoType_Group_Reps_iq --redo -T auto ${params.iqCustomaa} + + elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then + mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') + iqtree -s ${reps} --prefix ${params.protag}_AminoType_Group_Reps_iq -m \${mod} --redo -nt auto -b ${params.boots} + + elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then + mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') + iqtree -s ${reps} --prefix ${params.protag}_AminoType_Group_Reps_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni + + elif [ "${params.nonparametric}" != "false" ];then + iqtree -s ${reps} --prefix ${params.protag}_AminoType_Group_Reps_iq -m MFP --redo -nt auto -b ${params.boots} + + elif [ "${params.parametric}" != "false" ];then + iqtree -s ${reps} --prefix ${params.protag}_AminoType_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + else + iqtree -s ${reps} --prefix ${params.protag}_AminoType_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + fi """ } From 72bdaa85d2c62e218941cf7fe4465b74f37d4207 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Fri, 26 Mar 2021 17:47:39 -0500 Subject: [PATCH 061/223] Added Scripts for MED info incorportation --- bin/vAMPirus_ReportA.Rmd | 705 ------------------------------------- bin/vAMPirus_ReportB.Rmd | 725 --------------------------------------- vAMPirus.nf | 101 ++++-- 3 files changed, 71 insertions(+), 1460 deletions(-) delete mode 100644 bin/vAMPirus_ReportA.Rmd delete mode 100644 bin/vAMPirus_ReportB.Rmd diff --git a/bin/vAMPirus_ReportA.Rmd b/bin/vAMPirus_ReportA.Rmd deleted file mode 100644 index ad6fbf1..0000000 --- a/bin/vAMPirus_ReportA.Rmd +++ /dev/null @@ -1,705 +0,0 @@ ---- -title: "vAMPirus Analyze Report `r commandArgs(trailingOnly=T)[1]`" -date: "Generated on: `r Sys.time()`" -output: html_document -params: - interactive: TRUE - reads: !r commandArgs(trailingOnly=T)[2] # reads - counts: !r commandArgs(trailingOnly=T)[3] # csv - metadata: !r commandArgs(trailingOnly=T)[4] # metadata - filter: !r commandArgs(trailingOnly=T)[5] # filter min counts - heatmap: !r commandArgs(trailingOnly=T)[6] # heatmap - tax: !r commandArgs(trailingOnly=T)[7] # tax - try: !r commandArgs(trailingOnly=T)[8] # trymax - stats: !r commandArgs(trailingOnly=T)[9] # stats ---- - - -```{r setup, include=FALSE} -knitr::opts_chunk$set(echo = TRUE, - message = FALSE, - warning = FALSE, - out.width="100%") -``` - -```{r pathways, echo=FALSE} -knitr::include_graphics("vamplogo.png") -``` - -```{r load_libraries, include=FALSE} - -library(BiocManager) -library(vegan) -#library(rstatix) -library(tidyverse) -library(scales) -library(cowplot) -library(dplyr) -library(ggtree) -library(plotly) -#library(BiocParallel) -library(knitr) -library(kableExtra) #install.packages("kableExtra") -library(rmarkdown) -library(processx) #install.packages("processx") -#register(MulticoreParam(4)) -``` - -```{r colors, include=FALSE} -mycol=c('#088da5','#73cdc8','#ff6f61','#7cb8df','#88b04b','#00a199','#6B5B95','#92A8D1','#b0e0e6','#ff7f50','#088d9b','#E15D44','#e19336') -``` -
------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- -NOTE: Most plots are interactive and you can use the legend to specify samples/treatment of interest. You can also download an .svg version of each figure within this report. ------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- - -
-

  Pre- and Post-Adapter Removal Read Stats

-
-```{r readstats, echo=FALSE} -reads_stats=read.csv(params$reads) -#reads_stats=read.csv("PVID_final_reads_stats.csv") -paged_table(reads_stats,options = list(rows.print = 20)) - -#test plotly table -fig <- plot_ly( - type = 'table', - header = list(values=names(reads_stats), - align = c('left'), - line = list(width = 1, color = 'black'), - fill = list(color = 'rgb(45, 100, 230)')), - cells = list(values=unname(reads_stats), - line = list(width = 1, color = 'black'), - fill = list(color = 'white'), - align = c('center'), - font = list(color = '#506784', size = 10)) - ) - -#fig -``` -
-
- - -### Total number of reads before and after adapter removal - -```{r readstats_plot, echo=FALSE} -# Plot of reads before and after -ptotal <- plot_ly(typle="box",marker=list(colors=mycol)) -ptotal <- ptotal %>% add_boxplot(y=reads_stats$Total_before, name="Reads before filtering") -ptotal <- ptotal %>% add_boxplot(y=reads_stats$Total_after, name="Reads after filtering") -#ptotal <- ptotal %>% layout(title=list(text="Number of reads before and after filtering")) -ptotal <- ptotal %>% layout(legend = list(x=10,y=.5)) -ptotal <- ptotal %>% config(toImageButtonOptions=list(format='svg',filename='TotReads_b4_af_adaptrem', height= 500, width= 800, scale= 1)) -ptotal -``` -
- -### Forward (R1) and reverse (R2) read length before and after adapter removal - -```{r readstats_plot2, echo=FALSE} -# Plot of R1 and R2 before and after -pr <- plot_ly(y=reads_stats$R1_before_length, type="box", name="R1 length before") -pr <- pr %>% add_boxplot(y=reads_stats$R1_after_length, name="R1 length after") -pr <- pr %>% add_boxplot(y=reads_stats$R2_before_length, name="R2 length before") -pr <- pr %>% add_boxplot(y=reads_stats$R2_after_length, name="R2 length after") -#pr <- pr %>% layout(title = "R1 and R2 Length") -pr <- pr %>% layout(legend = list(x=10,y=.5)) -pr <- pr %>% config(toImageButtonOptions=list(format='svg',filename='readlen_b4_af_adaptrem', height= 500, width= 800, scale= 1)) -pr -``` -
-
-
-```{r load_datasets, include=FALSE} -sample_name=params$counts -sample_metadata=params$metadata -#sample_name="vAMPset_nOTU.90_counts.csv" -#sample_metadata="meta.csv" -data<- read.csv(sample_name, check.names=FALSE) -data2 <-as.data.frame(t(data)) -data2$sample <- row.names(data2) -colnames(data2)<- as.matrix(data2[1,]) -as.data.frame(data2) -data2 <- data2[-1,] - -#X.OTU.ID for X.Sequence. -data2 <- data2 %>% - rename(sample=OTU_ID) -data2dim <- dim(data2) - -##Loading metadata -samples <- read.csv(sample_metadata, header=TRUE) - -##Combining data and metadata -data3 <- merge(data2, samples, by="sample") - -dim_data3 <- dim(data3) -dim_samples <- dim(samples) -cols <- dim_data3[2]-dim_samples[2]+1 -first <-colnames(data3)[2] -last <- colnames(data3)[cols] -data3[,2:cols] <- lapply(data3[,2:cols], as.character) -data3[,2:cols] <- lapply(data3[,2:cols], as.numeric) - -#Calculate total reads per sample -data4 <- data3%>% - mutate(sum=select(.,2:cols)%>% - apply(1, sum, na.rm=TRUE)) - -``` - - -

  Number of Reads Per Sample

- -```{r plot, echo=FALSE} -# sample and count -con <- plot_ly(data4, x = ~sum, y = ~sample, name = "Sample", type = 'scatter', - mode = "markers", marker = list(color = "#088da5"), hovertemplate = paste('Sample: %{y}','
Total reads: %{x}','')) -con <- con %>% layout(xaxis = list(title = "Total reads"), yaxis = list(title = "Sample")) -con <- con %>% config(toImageButtonOptions=list(format='svg',filename='Counts_per_sample', height= 500, width= 800, scale= 1)) -con -``` -
-
-
-```{r filter_data, include=FALSE} -##Filter samples with low reads -nfil=params$filter -#nfil=1000 -data5 <- data4 %>% - filter(sum>nfil) - #can cause errors -data5dim <-dim(data5) -minreads<-min(data5$sum) -``` -
-
-
-
-
- -

  Rarefaction

-```{r rarefaction, echo=FALSE, cache=FALSE} -##Rarefaction curves -rarefaction <- rarecurve(data5[,2:cols]) - -##rarefied dataset -raredata <- as.data.frame(rrarefy(data5[,2:cols], sample=minreads)) -``` -
-
-
-
-
- - -

  Diversity Analyses Plots

- -
-
- -### Shannon diversity - -
-```{r diversity_analysis, echo=FALSE} -metadata <- data5[,(cols+1):data5dim[2]] -metadata$sample <- data5$sample -index <-diversity(raredata, index= "shannon") -shannondata5 <- as.data.frame(index) -shannondata5$sample<- data5$sample -shannondata5_2 <- merge(shannondata5, metadata, by="sample") - -#shannonplot <- ggplot(shannondata5_2, aes(x=treatment,color=treatment,y=index))+ -# geom_boxplot()+ -# geom_point()+ -# theme_classic()+ -# labs(y="Index",x="Treatment")+ -# theme(axis.text=element_text(size=12))+ -# theme(legend.position = "none") - -#sh<-ggplotly(shannonplot) -#sh <- sh %>% layout(title = list(text="Shannon diversty",y=.99)) -#sh - -sh <- plot_ly(shannondata5_2, x=~treatment, y=~index, color=~treatment, colors=mycol, type="box", boxpoints = "all", pointpos = 0, jitter = 0.5) -#sh <- sh %>% layout(title = list(text="Shannon diversty",y=.99)) -sh <- sh %>% layout(legend = list(x=10,y=.5), yaxis=list(title = "Index"), xaxis=list(title = "Treatment")) -sh <- sh %>% config(toImageButtonOptions=list(format='svg',filename='ShannonDiv', height= 500, width= 800, scale= 1)) -sh - -if (params$stats == "true" ) { - shannonaov <- aov(index ~ treatment, data= shannondata5_2) - st <- shapiro.test(resid(shannonaov)) - bt <- bartlett.test(index ~ treatment, data= shannondata5_2) - - if (st$p.value > .05 && bt$p.value > .05) { - print("Shapiro Test of normality - data is normal p-value > 0.05") - print(shapiro.test(resid(shannonaov))) - writeLines("\n--------------------------------------------------------------\n") - print("Bartlett Test variance homogeneity - variance is homogeneous p-value > 0.05") - print(bartlett.test(index ~ treatment, data= shannondata5_2)) - writeLines("\n--------------------------------------------------------------\n") - print("ANOVA Results") - print(summary(shannonaov)) - writeLines("\n--------------------------------------------------------------\n") - #Tukey Honest Significant Differences (pairwise comparison) - significant p <.05 - print("Tukey HSD - Pairwise comparison - significant differences indicated by p-value < 0.05") - print(TukeyHSD(shannonaov)) - writeLines("\n--------------------------------------------------------------\n") - } else { - print("Shapiro Test of normality - data is normal if p-value > 0.05") - print(shapiro.test(resid(shannonaov))) - writeLines("\n--------------------------------------------------------------\n") - print("Bartlett Test variance homogeneity - variance is homogeneous if p-value > 0.05") - print(bartlett.test(index ~ treatment, data= shannondata5_2)) - writeLines("\n--------------------------------------------------------------\n") - print("Data either not normal or variance not homogenous") - print("Kruskal-Wallis Test - test significant if p <.05") - #Kruskal-Wallis test - significant p <.05 - mykt <- kruskal.test(index ~ treatment, data= shannondata5_2) - print(mykt) - writeLines("\n--------------------------------------------------------------\n") - if (mykt$p.value < .05) { - #Pairwise comparison - print("Wilcox.test - pairwise comparison") - print(pairwise.wilcox.test(shannondata5_2$index, shannondata5_2$treatment, p.adjust.method = "BH")) - } else { - print("Data not significant. Skipping pairwise comparison") - } - writeLines("\n--------------------------------------------------------------\n") - print("ANOVA - one or more of the assumptions not met, take with a grain of salt.") - print(summary(shannonaov)) - writeLines("\n--------------------------------------------------------------\n") - } -} else { - print("Stats skipped. To toggle on use \"--stats run\" in vAMPirus launch command") -} -``` -
-
-
-
- -### Simpson diversty - -
-```{r diversity_analysis2, echo=FALSE} -index <- diversity(raredata, index= "simpson") -simpsondata5 <- as.data.frame(index) -simpsondata5$sample<- data5$sample -simpsondata5_2 <- merge(simpsondata5, metadata, by="sample") - -#simpsonplot <- ggplot(simpsondata5_2, aes(x=treatment,color=treatment,y=index))+ -# geom_boxplot()+ -# geom_point()+ -# theme_classic()+ -# labs(y="Index",x="Treatment")+ -# theme(axis.text=element_text(size=12))+ -# theme(legend.position = "none") - -#s<-ggplotly(simpsonplot) -#s <- s %>% layout(title = list(text="Simpson diversty",y=.99)) -#s - -s <- plot_ly(simpsondata5_2, x=~treatment, y=~index, color=~treatment, colors=mycol, type="box", boxpoints = "all", pointpos = 0, jitter = 0.5) -#s <- s %>% layout(title = list(text="Simpson diversty",y=.99)) -s <- s %>% layout(legend = list(x=10,y=.5), yaxis=list(title = "Index"), xaxis=list(title = "Treatment")) -s <- s %>% config(toImageButtonOptions=list(format='svg',filename='SimpsonDiv', height= 500, width= 800, scale= 1)) -s - -if (params$stats == "true" ) { - simpsonaov <- aov(index ~ treatment, data= simpsondata5_2) - st <- shapiro.test(resid(simpsonaov)) - bt <- bartlett.test(index ~ treatment, data= simpsondata5_2) - - if (st$p.value > .05 && bt$p.value > .05) { - print("Shapiro Test of normality - data is normal p-value > 0.05") - print(shapiro.test(resid(simpsonaov))) - writeLines("\n--------------------------------------------------------------\n") - print("Bartlett Test variance homogeneity - variance is homogeneous p-value > 0.05") - print(bartlett.test(index ~ treatment, data= simpsondata5_2)) - writeLines("\n--------------------------------------------------------------\n") - print("ANOVA Results") - print(summary(simpsonaov)) - writeLines("\n--------------------------------------------------------------\n") - #Tukey Honest Significant Differences (pairwise comparison) - significant p <.05 - print("Tukey HSD - Pairwise comparison - significant differences indicated by p-value < 0.05") - print(TukeyHSD(simpsonaov)) - writeLines("\n--------------------------------------------------------------\n") - } else { - print("Shapiro Test of normality - data is normal if p-value > 0.05") - print(shapiro.test(resid(simpsonaov))) - writeLines("\n--------------------------------------------------------------\n") - print("Bartlett Test variance homogeneity - variance is homogeneous if p-value > 0.05") - print(bartlett.test(index ~ treatment, data= simpsondata5_2)) - writeLines("\n--------------------------------------------------------------\n") - print("Data either not normal or variance not homogenous") - print("Kruskal-Wallis Test - test significant if p <.05") - #Kruskal-Wallis test - significant p <.05 - mykt <- kruskal.test(index ~ treatment, data= simpsondata5_2) - print(mykt) - writeLines("\n--------------------------------------------------------------\n") - if (mykt$p.value < .05) { - #Pairwise comparison - print("Wilcox.test - pairwise comparison") - print(pairwise.wilcox.test(simpsondata5_2$index, simpsondata5_2$treatment, p.adjust.method = "BH")) - } else { - print("Data not significant. Skipping pairwise comparison") - } - writeLines("\n--------------------------------------------------------------\n") - print("ANOVA - one or more of the assumptions not met, take with a grain of salt.") - print(summary(simpsonaov)) - writeLines("\n--------------------------------------------------------------\n") - } -} else { - print("Stats skipped. To toggle on use \"--stats run\" in vAMPirus launch command") -} -``` -
-
-
-
- -### Species Richness - -
-```{r diversity_analysis3, echo=FALSE} -mind5<-min(data5$sum) -index <- rarefy(data5[,2:cols], sample=mind5) -rarerichnessdata5 <- as.data.frame(index) -rarerichnessdata5$sample <-data5$sample -richdata5_2 <- merge(rarerichnessdata5, metadata, by="sample") - -#richnessplot <- ggplot(richdata5_2, aes(x=treatment,color=treatment,y=index))+ -# geom_boxplot()+ -# geom_point()+ -# theme_classic()+ -# labs(y="Richness",x="Treatment")+ -# theme(axis.text=element_text(size=12))+ -# theme(legend.position = "none") - -ri <- plot_ly(richdata5_2, x=~treatment, y=~index, color=~treatment, colors=mycol, type="box", boxpoints = "all", pointpos = 0, jitter = 0.5) -#ri <- ri %>% layout(title = list(text="ASV Richness",y=.99)) -ri <- ri %>% layout(legend = list(x=10,y=.5), yaxis=list(title = "Index"), xaxis=list(title = "Treatment")) -ri <- ri %>% config(toImageButtonOptions=list(format='svg',filename='SpeciesRich', height= 500, width= 800, scale= 1)) -ri - -if (params$stats == "true" ) { - richaov <- aov(index ~ treatment, data= richdata5_2) - st <- shapiro.test(resid(richaov)) - bt <- bartlett.test(index ~ treatment, data= richdata5_2) - - if (st$p.value > .05 && bt$p.value > .05) { - print("Shapiro Test of normality - data is normal p-value > 0.05") - print(shapiro.test(resid(richaov))) - writeLines("\n--------------------------------------------------------------\n") - print("Bartlett Test variance homogeneity - variance is homogeneous p-value > 0.05") - print(bartlett.test(index ~ treatment, data= richdata5_2)) - writeLines("\n--------------------------------------------------------------\n") - print("ANOVA Results") - print(summary(richaov)) - writeLines("\n--------------------------------------------------------------\n") - #Tukey Honest Significant Differences (pairwise comparison) - significant p <.05 - print("Tukey HSD - Pairwise comparison - significant differences indicated by p-value < 0.05") - print(TukeyHSD(richaov)) - writeLines("\n--------------------------------------------------------------\n") - } else { - print("Shapiro Test of normality - data is normal if p-value > 0.05") - print(shapiro.test(resid(richaov))) - writeLines("\n--------------------------------------------------------------\n") - print("Bartlett Test variance homogeneity - variance is homogeneous if p-value > 0.05") - print(bartlett.test(index ~ treatment, data= richdata5_2)) - writeLines("\n--------------------------------------------------------------\n") - print("Data either not normal or variance not homogenous") - print("Kruskal-Wallis Test - test significant if p <.05") - #Kruskal-Wallis test - significant p <.05 - mykt <- kruskal.test(index ~ treatment, data= richdata5_2) - print(mykt) - writeLines("\n--------------------------------------------------------------\n") - if (mykt$p.value < .05) { - #Pairwise comparison - print("Wilcox.test - pairwise comparison") - print(pairwise.wilcox.test(richdata5_2$index, richdata5_2$treatment, p.adjust.method = "BH")) - } else { - print("Data not significant. Skipping pairwise comparison") - } - writeLines("\n--------------------------------------------------------------\n") - print("ANOVA - one or more of the assumptions not met, take with a grain of salt.") - print(summary(richaov)) - writeLines("\n--------------------------------------------------------------\n") - } -} else { - print("Stats skipped. To toggle on use \"--stats run\" in vAMPirus launch command") -} -``` -
-
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-
-
- -

  Distance To Centroid

-
-```{r distance, echo=FALSE} -##Distance -intermediate <- raredata -bray.distance <- vegdist(intermediate, method="bray",autotransform=TRUE) - -##Dispersion -disper <- betadisper(bray.distance, group = metadata$treatment, type="centroid") -#anova(disper) -df <- data.frame(Distance_to_centroid=disper$distances,Group=disper$group) -df$sample <- data5$sample -df2 <- merge(df, metadata, by="sample") - -#p<- ggplot(data=df2,aes(x=treatment,y=Distance_to_centroid,colour=treatment))+ -# geom_boxplot(outlier.alpha = 0)+ -# theme_classic()+ -# geom_point(position=position_dodge(width=0.75))+ -# labs(y="Distance",x="Treatment")+ -# theme(axis.text=element_text(size=12))+ -# theme(legend.position = "none") - -#cen <- ggplotly(p) -#cen <- cen %>% layout(title = list(text="Distance to centroid",y=.99)) -#cen - -cen <- plot_ly(df2, x=~treatment, y=~Distance_to_centroid, color=~treatment, colors=mycol, type="box", boxpoints = "all", pointpos = 0, jitter = 0.5) -#cen <- cen %>% layout(title = list(text="Distance to centroid",y=.99)) -cen <- cen %>% layout(legend = list(x=10,y=.5), yaxis=list(title = "Distance"), xaxis=list(title = "Treatment")) -cen <- cen %>% config(toImageButtonOptions=list(format='svg',filename='Dispersion', height= 500, width= 800, scale= 1)) -cen - -if (params$stats == "true" ) { - adn <- adonis(bray.distance~data5$treatment) - adn -} else { - print("Stats skipped. To toggle on use \"--stats run\" in vAMPirus launch command") -} -``` -
-
-
-
-
- -

  NMDS Plots

- -
- -### 2D NMDS - -
-```{r nmds2d, echo=FALSE} -##NMDS -datax <- decostand(raredata,method="total") #method 'total' normalizes data to sum up to 1 --data5[,2:cols] - -MDS <- metaMDS(sqrt(datax), - distance = "bray",autotransform = FALSE, - k = 2, - maxit = 999, - trymax = params$try, - wascores = TRUE) - -if (MDS$converged == "TRUE") { - -data.scores <- as.data.frame(scores(MDS)) -data.scores$sample <- data5$sample -data.scores.2 <- merge(data.scores, metadata, by="sample") - -p <- ggplot(data.scores.2, aes(x=NMDS1, y=NMDS2,color=treatment))+ - geom_point(size=2)+ - theme_classic()+ - theme(legend.title = element_blank()) - - -#fff <- ggplotly(p) -#fff <- fff %>% layout(legend=list(y=.5)) -#fff - -fff <-plot_ly(data.scores.2, x=~NMDS1, y=~NMDS2, color=~treatment, colors=mycol, text = ~paste("Sample: ", sample)) -fff <- fff %>% layout(legend=list(y=.5)) -fff <- fff %>% config(toImageButtonOptions=list(format='svg',filename='2Dnmds', height= 500, width= 800, scale= 1)) -fff - -} else { - print("No Convergence") -} -``` - -
-
-
-
- -### 3D NMDS - -
- -```{r nmds3d, echo=FALSE} -MDS3 <- metaMDS(sqrt(datax), - distance = "bray",autotransform = FALSE, - k = 3, - maxit = 999, - trymax = params$try, - wascores = TRUE) - -if (MDS3$converged == "TRUE") { - -data.scores3 <- as.data.frame(scores(MDS3)) -data.scores3$sample <- data5$sample -data.scores.3 <- merge(data.scores3, metadata, by="sample") -p3d <- plot_ly(data.scores.3, x= ~ NMDS1, y = ~ NMDS2, z = ~ NMDS3, text = ~paste("Sample: ", sample), - color = ~treatment, colors = mycol, - mode = 'markers', symbol = ~treatment, symbols = c('square','circle'), - marker = list(opacity = .8,line=list(color = 'darkblue',width = 1)) - ) -p3d <- p3d %>% layout(legend=list(y=.5)) -p3d <- p3d %>% config(toImageButtonOptions=list(format='svg',filename='3Dnmds', height= 500, width= 800, scale= 1)) -p3d - -} else { - print("No Convergence") -} -``` -
-
-
-
-
- -

  OTU Abundance Per Sample

- -```{r long, echo=FALSE} -dataz <- decostand(data5[,2:cols],method="total") #method 'total' normalizes data to sum up to 1 -dataz$sample <- data5$sample -datay <- merge(dataz, metadata, by="sample") -datalong <- datay %>% - tidyr::gather(first:last, key=hit, value=reads) - -##Barplot -## add better colors -#spec_bar <- ggplot(datalong, aes(x=forcats::fct_reorder(timepoint, as.numeric(as.character(timepoint))), -# y=reads, fill=hit))+ #Consider sqrt-transforming data -# geom_bar(aes(), stat="identity", position="fill")+ -# #coord_polar("y", start=0)+ -# theme_classic()+ -# facet_wrap(colony~treatment, nrow=2)+ -# #coord_flip()+ -# labs(x="Timepoint") #+ theme(legend.position = "none") -#spec_bar <- ggplot(datalong, aes(x=sample,y=reads,fill=hit))+geom_bar(aes(), stat="identity", position="fill")+ -# theme(axis.text.x=element_text(angle=90)) -#ggplotly(spec_bar) -#,'#00b300','#00b3b3','#0059b3','#6600ff','#b800e6','#ff3333','#ff8000','#ffff00','#bf8040','#42bcf5','#b428f5','#2e8d7e','#664e7e','#a4c700','#1aa3ff' - -ddd <- plot_ly(datalong, x=~sample, y=~reads, color=~hit, colors=mycol) -ddd <- ddd %>% layout(type='bar', barmode = 'stack') -ddd <- ddd %>% layout(legend = list(x=10,y=.5), xaxis=list(title = "Sample"), yaxis=list(title = "Relative abundance")) -ddd <- ddd %>% config(toImageButtonOptions=list(format='svg',filename='Relative_abundance', height= 500, width= 800, scale= 1)) -ddd -``` -
-
-
-
-
- -

  OTU Abundance Per Treatment

- -```{r asv_barplot, echo=FALSE} -datalong <- datalong %>% - filter(reads>0) - -#asv_bar <- ggplot(datalong, aes(x=reorder(hit,reads), y=reads, fill=treatment))+ -# geom_bar(stat="identity")+ -# scale_fill_manual(values=c('#088da5','#e19336'))+ #colors not working -# coord_flip()+ -# theme_classic()+ -# theme(axis.title.y = element_blank())+ -# theme(legend.title = element_blank()) - -asp2 <- plot_ly(datalong, y=~hit, x=~reads, color=~treatment, colors=mycol,text = ~paste("Sample: ", sample), opacity=.9) -asp2 <- asp2 %>% layout(type='bar', barmode = 'group') -asp2 <- asp2 %>% layout(yaxis = list(title = '', categoryorder = "total ascending"), legend = list(x=10,y=.5)) -asp2 <- asp2 %>% config(toImageButtonOptions=list(format='svg',filename='Most_abundant_hits_per_sample', height= 500, width= 800, scale= 1)) -asp2 -``` -
-
-
-
-
- -

  Pairwise Percent-ID Heatmap

- -
-
-```{r heatmap, echo=FALSE} -heatdata=params$heatmap -#heatdata="a.csv" -#heatdata="PVID_vAMPtest1_otu.85_PercentID.matrix" -simmatrix<- read.csv(heatdata, header=FALSE) -rownames(simmatrix) <- simmatrix[,1] -simmatrix <- simmatrix[,-1] -colnames(simmatrix) <-rownames(simmatrix) -cols <- dim(simmatrix)[2] -simmatrix$AA <- rownames(simmatrix) -rval=nrow(simmatrix) -simmatrix2 <- simmatrix %>% - gather(1:rval, key=sequence, value=similarity) -x=reorder(simmatrix2$AA,simmatrix2$similarity) -y=reorder(simmatrix2$sequence,simmatrix2$similarity) -similaritymatrix <- ggplot(simmatrix2, aes(x=x, y=y,fill=similarity))+ - geom_raster()+ - scale_fill_distiller(palette="Spectral")+ - theme(axis.text.x = element_text(angle = 90))+ - theme(axis.title.x=element_blank())+ - theme(axis.title.y=element_blank()) - - -heat <- ggplotly(similaritymatrix) -heat <- heat %>% config(toImageButtonOptions=list(format='svg',filename='heatmap', height= 500, width= 800, scale= 1)) -heat -``` -
-
-
-
-
- - -

  Taxonomy Result Visualization

- -
-
- -```{r taxonomy, echo=FALSE} -tax=read.csv(params$tax,header=F) -#tax=read.csv("tax.csv",header=F) -#tax=read.csv("PVID_vAMPtest1_otu.85_summary_for_plot.csv", header=F) -tp <- plot_ly(tax, labels = ~V1, values = ~V2) -tp <- tp %>% add_pie(marker=list(colors=mycol, line=list(color='#000000', width=.5)), hole = 0.6) -tp <- tp %>% layout(title = "Taxonomy distribution", showlegend = F, - xaxis = list(showgrid = FALSE, zeroline = FALSE, showticklabels = FALSE), - yaxis = list(showgrid = FALSE, zeroline = FALSE, showticklabels = FALSE) - ) -tp <- tp %>% config(toImageButtonOptions=list(format='svg',filename='TaxDonut', height= 500, width= 800, scale= 1)) -tp -``` -
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-
diff --git a/bin/vAMPirus_ReportB.Rmd b/bin/vAMPirus_ReportB.Rmd deleted file mode 100644 index 29b0385..0000000 --- a/bin/vAMPirus_ReportB.Rmd +++ /dev/null @@ -1,725 +0,0 @@ ---- -title: "vAMPirus Analyze Report `r commandArgs(trailingOnly=T)[1]`" -date: "Generated on: `r Sys.time()`" -output: html_document -params: - interactive: TRUE - reads: !r commandArgs(trailingOnly=T)[2] # reads - counts: !r commandArgs(trailingOnly=T)[3] # csv - metadata: !r commandArgs(trailingOnly=T)[4] # metadata - filter: !r commandArgs(trailingOnly=T)[5] # filter min counts - heatmap: !r commandArgs(trailingOnly=T)[6] # heatmap - tax: !r commandArgs(trailingOnly=T)[7] # tax - tree: !r commandArgs(trailingOnly=T)[8] # tree - try: !r commandArgs(trailingOnly=T)[9] # trymax - stats: !r commandArgs(trailingOnly=T)[10] # stats - ---- - - -```{r setup, include=FALSE} -knitr::opts_chunk$set(echo = TRUE, - message = FALSE, - warning = FALSE, - out.width="100%") -``` - -```{r pathways, echo=FALSE} -knitr::include_graphics("vamplogo.png") -``` - -```{r load_libraries, include=FALSE} - -library(vegan) -#library(rstatix) -library(tidyverse) -library(scales) -library(cowplot) -library(dplyr) -library(ggtree) -library(plotly) -#library(BiocParallel) -library(knitr) -library(kableExtra) #install.packages("kableExtra") -library(rmarkdown) -library(processx) #install.packages("processx") -#register(MulticoreParam(4)) -``` - -```{r colors, include=FALSE} -mycol=c('#088da5','#73cdc8','#ff6f61','#7cb8df','#88b04b','#00a199','#6B5B95','#92A8D1','#b0e0e6','#ff7f50','#088d9b','#E15D44','#e19336') -``` -
------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- -NOTE: Most plots are interactive and you can use the legend to specify samples/treatment of interest. You can also download an .svg version of each figure within this report. ------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- -
-

  Pre- and Post-Adapter Removal Read Stats

-
-```{r readstats, echo=FALSE} -reads_stats=read.csv(params$reads) -#reads_stats=read.csv("PVID_final_reads_stats.csv") -paged_table(reads_stats,options = list(rows.print = 20)) - -#test plotly table -fig <- plot_ly( - type = 'table', - header = list(values=names(reads_stats), - align = c('left'), - line = list(width = 1, color = 'black'), - fill = list(color = 'rgb(45, 100, 230)')), - cells = list(values=unname(reads_stats), - line = list(width = 1, color = 'black'), - fill = list(color = 'white'), - align = c('center'), - font = list(color = '#506784', size = 10)) - ) - -#fig -``` -
-
- - -### Total number of reads before and after adapter removal - -```{r readstats_plot, echo=FALSE} -# Plot of reads before and after -ptotal <- plot_ly(type="box",marker=list(colors=mycol)) -ptotal <- ptotal %>% add_boxplot(y=reads_stats$Total_before, name="Reads before filtering") -ptotal <- ptotal %>% add_boxplot(y=reads_stats$Total_after, name="Reads after filtering") -#ptotal <- ptotal %>% layout(title=list(text="Number of reads before and after filtering")) -ptotal <- ptotal %>% layout(legend = list(x=10,y=.5)) -ptotal <- ptotal %>% config(toImageButtonOptions=list(format='svg',filename='TotReads_b4_af_adaptrem', height= 500, width= 800, scale= 1)) -ptotal -``` -
- -### Forward (R1) and reverse (R2) read length before and after adapter removal - -```{r readstats_plot2, echo=FALSE} -# Plot of R1 and R2 before and after -pr <- plot_ly(y=reads_stats$R1_before_length, type="box", name="R1 length before") -pr <- pr %>% add_boxplot(y=reads_stats$R1_after_length, name="R1 length after") -pr <- pr %>% add_boxplot(y=reads_stats$R2_before_length, name="R2 length before") -pr <- pr %>% add_boxplot(y=reads_stats$R2_after_length, name="R2 length after") -#pr <- pr %>% layout(title = "R1 and R2 Length") -pr <- pr %>% layout(legend = list(x=10,y=.5)) -pr <- pr %>% config(toImageButtonOptions=list(format='svg',filename='readlen_b4_af_adaptrem', height= 500, width= 800, scale= 1)) -pr -``` -
-
-
-```{r load_datasets, include=FALSE} -sample_name=params$counts -sample_metadata=params$metadata -#sample_name="PVID_vAMPtest1_otu.85_counts.csv" -#sample_metadata="PVID_fiscesTestmeta.csv" -#sample_name="vAMPrun_otu.55_counts.csv" -#sample_metadata="pvid_samples.csv" -data<- read.csv(sample_name, check.names=FALSE) -data2 <-as.data.frame(t(data)) -data2$sample <- row.names(data2) -colnames(data2)<- as.matrix(data2[1,]) -as.data.frame(data2) -data2 <- data2[-1,] - -#X.OTU.ID for X.Sequence. -data2 <- data2 %>% - rename(sample=OTU_ID) -data2dim <- dim(data2) - -##Loading metadata -samples <- read.csv(sample_metadata) - -##Combining data and metadata -data3 <- merge(data2, samples, by="sample") - -dim_data3 <- dim(data3) -dim_samples <- dim(samples) -cols <- dim_data3[2]-dim_samples[2]+1 -first <-colnames(data3)[2] -last <- colnames(data3)[cols] -data3[,2:cols] <- lapply(data3[,2:cols], as.character) -data3[,2:cols] <- lapply(data3[,2:cols], as.numeric) - -#Calculate total reads per sample -data4 <- data3%>% - mutate(sum=select(.,2:cols)%>% - apply(1, sum, na.rm=TRUE)) - -``` - - -

  Number of Reads Per Sample

- -```{r plot, echo=FALSE} -# sample and count -con <- plot_ly(data4, x = ~sum, y = ~sample, name = "Sample", type = 'scatter', - mode = "markers", marker = list(color = "#088da5"), hovertemplate = paste('Sample: %{y}','
Total reads: %{x}','')) -con <- con %>% layout(xaxis = list(title = "Total reads"),yaxis = list(title = "Sample")) -con <- con %>% config(toImageButtonOptions=list(format='svg',filename='Counts_per_sample', height= 500, width= 800, scale= 1)) -con -``` -
-
-
-```{r filter_data, include=FALSE} -##Filter samples with low reads -nfil=params$filter -data5 <- data4 %>% - filter(sum>nfil) - #can cause errors -data5dim <-dim(data5) -minreads<-min(data5$sum) -``` -
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-
-
-
- -

  Rarefaction

-```{r rarefaction, echo=FALSE, cache=FALSE} -##Rarefaction curves -rarefaction <- rarecurve(data5[,2:cols]) - -##rarefied dataset -raredata <- as.data.frame(rrarefy(data5[,2:cols], sample=minreads)) -``` -
-
-
-
-
- - -

  Diversity Analyses Plots

- -
-
- -### Shannon diversty - -
-```{r diversity_analysis, echo=FALSE} -metadata <- data5[,(cols+1):data5dim[2]] -metadata$sample <- data5$sample -index <-diversity(raredata, index= "shannon") -shannondata5 <- as.data.frame(index) -shannondata5$sample<- data5$sample -shannondata5_2 <- merge(shannondata5, metadata, by="sample") - -#shannonplot <- ggplot(shannondata5_2, aes(x=treatment,color=treatment,y=index))+ -# geom_boxplot()+ -# geom_point()+ -# theme_classic()+ -# labs(y="Index",x="Treatment")+ -# theme(axis.text=element_text(size=12))+ -# theme(legend.position = "none") - -#sh<-ggplotly(shannonplot) -#sh <- sh %>% layout(title = list(text="Shannon diversty",y=.99)) -#sh - -sh <- plot_ly(shannondata5_2, x=~treatment, y=~index, color=~treatment, colors=mycol, type="box", boxpoints = "all", pointpos = 0, jitter = 0.5) -#sh <- sh %>% layout(title = list(text="Shannon diversty",y=.99)) -sh <- sh %>% layout(legend = list(x=10,y=.5), yaxis=list(title = "Index"), xaxis=list(title = "Treatment")) -sh <- sh %>% config(toImageButtonOptions=list(format='svg',filename='ShannonDiv', height= 500, width= 800, scale= 1)) -sh - -if (params$stats == "true" ) { - shannonaov <- aov(index ~ treatment, data= shannondata5_2) - st <- shapiro.test(resid(shannonaov)) - bt <- bartlett.test(index ~ treatment, data= shannondata5_2) - - if (st$p.value > .05 && bt$p.value > .05) { - print("Shapiro Test of normality - data is normal p-value > 0.05") - print(shapiro.test(resid(shannonaov))) - writeLines("\n--------------------------------------------------------------\n") - print("Bartlett Test variance homogeneity - variance is homogeneous p-value > 0.05") - print(bartlett.test(index ~ treatment, data= shannondata5_2)) - writeLines("\n--------------------------------------------------------------\n") - print("ANOVA Results") - print(summary(shannonaov)) - writeLines("\n--------------------------------------------------------------\n") - #Tukey Honest Significant Differences (pairwise comparison) - significant p <.05 - print("Tukey HSD - Pairwise comparison - significant differences indicated by p-value < 0.05") - print(TukeyHSD(shannonaov)) - writeLines("\n--------------------------------------------------------------\n") - } else { - print("Shapiro Test of normality - data is normal if p-value > 0.05") - print(shapiro.test(resid(shannonaov))) - writeLines("\n--------------------------------------------------------------\n") - print("Bartlett Test variance homogeneity - variance is homogeneous if p-value > 0.05") - print(bartlett.test(index ~ treatment, data= shannondata5_2)) - writeLines("\n--------------------------------------------------------------\n") - print("Data either not normal or variance not homogenous") - print("Kruskal-Wallis Test - test significant if p <.05") - #Kruskal-Wallis test - significant p <.05 - mykt <- kruskal.test(index ~ treatment, data= shannondata5_2) - print(mykt) - writeLines("\n--------------------------------------------------------------\n") - if (mykt$p.value < .05) { - #Pairwise comparison - print("Wilcox.test - pairwise comparison") - print(pairwise.wilcox.test(shannondata5_2$index, shannondata5_2$treatment, p.adjust.method = "BH")) - } else { - print("Data not significant. Skipping pairwise comparison") - } - writeLines("\n--------------------------------------------------------------\n") - print("ANOVA - one or more of the assumptions not met, take with a grain of salt.") - print(summary(shannonaov)) - writeLines("\n--------------------------------------------------------------\n") - } -} else { - print("Stats skipped. To toggle on use \"--stats run\" in vAMPirus launch command") -} -``` -
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-
-
- -### Simpson diversty - -
-```{r diversity_analysis2, echo=FALSE} -index <- diversity(raredata, index= "simpson") -simpsondata5 <- as.data.frame(index) -simpsondata5$sample<- data5$sample -simpsondata5_2 <- merge(simpsondata5, metadata, by="sample") - -#simpsonplot <- ggplot(simpsondata5_2, aes(x=treatment,color=treatment,y=index))+ -# geom_boxplot()+ -# geom_point()+ -# theme_classic()+ -# labs(y="Index",x="Treatment")+ -# theme(axis.text=element_text(size=12))+ -# theme(legend.position = "none") - -#s<-ggplotly(simpsonplot) -#s <- s %>% layout(title = list(text="Simpson diversty",y=.99)) -#s - -s <- plot_ly(simpsondata5_2, x=~treatment, y=~index, color=~treatment, colors=mycol, type="box", boxpoints = "all", pointpos = 0, jitter = 0.5) -#s <- s %>% layout(title = list(text="Simpson diversty",y=.99)) -s <- s %>% layout(legend = list(x=10,y=.5), yaxis=list(title = "Index"), xaxis=list(title = "Treatment")) -s <- s %>% config(toImageButtonOptions=list(format='svg',filename='SimpsonDiv', height= 500, width= 800, scale= 1)) -s - -if (params$stats == "true" ) { - simpsonaov <- aov(index ~ treatment, data= simpsondata5_2) - st <- shapiro.test(resid(simpsonaov)) - bt <- bartlett.test(index ~ treatment, data= simpsondata5_2) - - if (st$p.value > .05 && bt$p.value > .05) { - print("Shapiro Test of normality - data is normal p-value > 0.05") - print(shapiro.test(resid(simpsonaov))) - writeLines("\n--------------------------------------------------------------\n") - print("Bartlett Test variance homogeneity - variance is homogeneous p-value > 0.05") - print(bartlett.test(index ~ treatment, data= simpsondata5_2)) - writeLines("\n--------------------------------------------------------------\n") - print("ANOVA Results") - print(summary(simpsonaov)) - writeLines("\n--------------------------------------------------------------\n") - #Tukey Honest Significant Differences (pairwise comparison) - significant p <.05 - print("Tukey HSD - Pairwise comparison - significant differences indicated by p-value < 0.05") - print(TukeyHSD(simpsonaov)) - writeLines("\n--------------------------------------------------------------\n") - } else { - print("Shapiro Test of normality - data is normal if p-value > 0.05") - print(shapiro.test(resid(simpsonaov))) - writeLines("\n--------------------------------------------------------------\n") - print("Bartlett Test variance homogeneity - variance is homogeneous if p-value > 0.05") - print(bartlett.test(index ~ treatment, data= simpsondata5_2)) - writeLines("\n--------------------------------------------------------------\n") - print("Data either not normal or variance not homogenous") - print("Kruskal-Wallis Test - test significant if p <.05") - #Kruskal-Wallis test - significant p <.05 - mykt <- kruskal.test(index ~ treatment, data= simpsondata5_2) - print(mykt) - writeLines("\n--------------------------------------------------------------\n") - if (mykt$p.value < .05) { - #Pairwise comparison - print("Wilcox.test - pairwise comparison") - print(pairwise.wilcox.test(simpsondata5_2$index, simpsondata5_2$treatment, p.adjust.method = "BH")) - } else { - print("Data not significant. Skipping pairwise comparison") - } - writeLines("\n--------------------------------------------------------------\n") - print("ANOVA - one or more of the assumptions not met, take with a grain of salt.") - print(summary(simpsonaov)) - writeLines("\n--------------------------------------------------------------\n") - } -} else { - print("Stats skipped. To toggle on use \"--stats run\" in vAMPirus launch command") -} -``` -
-
-
-
- -### Species Richness - -
-```{r diversity_analysis3, echo=FALSE} -mind5<-min(data5$sum) -index <- rarefy(data5[,2:cols], sample=mind5) -rarerichnessdata5 <- as.data.frame(index) -rarerichnessdata5$sample <-data5$sample -richdata5_2 <- merge(rarerichnessdata5, metadata, by="sample") - -#richnessplot <- ggplot(richdata5_2, aes(x=treatment,color=treatment,y=index))+ -# geom_boxplot()+ -# geom_point()+ -# theme_classic()+ -# labs(y="Richness",x="Treatment")+ -# theme(axis.text=element_text(size=12))+ -# theme(legend.position = "none") - -ri <- plot_ly(richdata5_2, x=~treatment, y=~index, color=~treatment, colors=mycol, type="box", boxpoints = "all", pointpos = 0, jitter = 0.5) -#ri <- ri %>% layout(title = list(text="ASV Richness",y=.99)) -ri <- ri %>% layout(legend = list(x=10,y=.5), yaxis=list(title = "Index"), xaxis=list(title = "Treatment")) -ri <- ri %>% config(toImageButtonOptions=list(format='svg',filename='SpeciesRich', height= 500, width= 800, scale= 1)) -ri - -if (params$stats == "true" ) { - richaov <- aov(index ~ treatment, data= richdata5_2) - st <- shapiro.test(resid(richaov)) - bt <- bartlett.test(index ~ treatment, data= richdata5_2) - - if (st$p.value > .05 && bt$p.value > .05) { - print("Shapiro Test of normality - data is normal p-value > 0.05") - print(shapiro.test(resid(richaov))) - writeLines("\n--------------------------------------------------------------\n") - print("Bartlett Test variance homogeneity - variance is homogeneous p-value > 0.05") - print(bartlett.test(index ~ treatment, data= richdata5_2)) - writeLines("\n--------------------------------------------------------------\n") - print("ANOVA Results") - print(summary(richaov)) - #Tukey Honest Significant Differences (pairwise comparison) - significant p <.05 - writeLines("\n--------------------------------------------------------------\n") - print("Tukey HSD - Pairwise comparison - significant differences indicated by p-value < 0.05") - print(TukeyHSD(richaov)) - writeLines("\n--------------------------------------------------------------\n") - } else { - print("Shapiro Test of normality - data is normal if p-value > 0.05") - print(shapiro.test(resid(richaov))) - writeLines("\n--------------------------------------------------------------\n") - print("Bartlett Test variance homogeneity - variance is homogeneous if p-value > 0.05") - print(bartlett.test(index ~ treatment, data= richdata5_2)) - writeLines("\n--------------------------------------------------------------\n") - print("Data either not normal or variance not homogenous") - print("Kruskal-Wallis Test - test significant if p <.05") - #Kruskal-Wallis test - significant p <.05 - mykt <- kruskal.test(index ~ treatment, data= richdata5_2) - print(mykt) - writeLines("\n--------------------------------------------------------------\n") - if (mykt$p.value < .05) { - #Pairwise comparison - print("Wilcox.test - pairwise comparison") - print(pairwise.wilcox.test(richdata5_2$index, richdata5_2$treatment, p.adjust.method = "BH")) - } else { - print("Data not significant. Skipping pairwise comparison") - } - writeLines("\n--------------------------------------------------------------\n") - print("ANOVA - one or more of the assumptions not met, take with a grain of salt.") - print(summary(richaov)) - writeLines("\n--------------------------------------------------------------\n") - } -} else { - print("Stats skipped. To toggle on use \"--stats run\" in vAMPirus launch command") -} -``` -
-
-
-
-
- -

  Distance To Centroid

-
-```{r distance, echo=FALSE} -##Distance -intermediate <- raredata -bray.distance <- vegdist(sqrt(intermediate), method="bray") - -##Dispersion -disper <- betadisper(bray.distance, group = metadata$treatment, type="centroid") -df <- data.frame(Distance_to_centroid=disper$distances,Group=disper$group) -df$sample <- data5$sample -df2 <- merge(df, metadata, by="sample") - -#p<- ggplot(data=df2,aes(x=treatment,y=Distance_to_centroid,colour=treatment))+ -# geom_boxplot(outlier.alpha = 0)+ -# theme_classic()+ -# geom_point(position=position_dodge(width=0.75))+ -# labs(y="Distance",x="Treatment")+ -# theme(axis.text=element_text(size=12))+ -# theme(legend.position = "none") - -#cen <- ggplotly(p) -#cen <- cen %>% layout(title = list(text="Distance to centroid",y=.99)) -#cen - -cen <- plot_ly(df2, x=~treatment, y=~Distance_to_centroid, color=~treatment, colors=mycol, type="box", boxpoints = "all", pointpos = 0, jitter = 0.5) -#cen <- cen %>% layout(title = list(text="Distance to centroid",y=.99)) -cen <- cen %>% layout(legend = list(x=10,y=.5), yaxis=list(title = "Distance"), xaxis=list(title = "Treatment")) -cen <- cen %>% config(toImageButtonOptions=list(format='svg',filename='Dispersion', height= 500, width= 800, scale= 1)) -cen - -if (params$stats == "true" ) { - adn <- adonis(bray.distance~data5$treatment) - adn -} else { - print("Stats skipped. To toggle on use \"--stats run\" in vAMPirus launch command") -} -``` -
-
-
-
-
- -

  NMDS Plots

- -
- -### 2D NMDS - -
-```{r nmds2d, echo=FALSE} -##NMDS -datax <- decostand(raredata,method="total") #method 'total' normalizes data to sum up to 1 --data5[,2:cols] - -MDS <- metaMDS(sqrt(datax), - distance = "bray",autotransform = FALSE, - k = 2, - maxit = 999, - trymax = params$try, - wascores = TRUE) - -if (MDS$converged == "TRUE") { - -data.scores <- as.data.frame(scores(MDS)) -data.scores$sample <- data5$sample -data.scores.2 <- merge(data.scores, metadata, by="sample") - -p <- ggplot(data.scores.2, aes(x=NMDS1, y=NMDS2,color=treatment))+ - geom_point(size=2)+ - theme_classic()+ - theme(legend.title = element_blank()) - -#fff <- ggplotly(p) -#fff <- fff %>% layout(legend=list(y=.5)) -#fff - -fff <-plot_ly(data.scores.2, x=~NMDS1, y=~NMDS2, color=~treatment, colors=mycol, text = ~paste("Sample: ", sample)) -fff <- fff %>% layout(legend=list(y=.5)) -fff <- fff %>% config(toImageButtonOptions=list(format='svg',filename='2Dnmds', height= 500, width= 800, scale= 1)) -fff - -} else { - print("No Convergence") -} -``` - -
-
-
-
- -### 3D NMDS - -
- -```{r nmds3d, echo=FALSE} -MDS3 <- metaMDS(sqrt(datax), - distance = "bray",autotransform = FALSE, - k = 3, - maxit = 999, - trymax = params$try, - wascores = TRUE) - -if (MDS3$converged == "TRUE") { - -data.scores3 <- as.data.frame(scores(MDS3)) -data.scores3$sample <- data5$sample -data.scores.3 <- merge(data.scores3, metadata, by="sample") -p3d <- plot_ly(data.scores.3, x=~NMDS1, y=~NMDS2, z=~NMDS3, text=~paste("Sample: ", sample), - color=~treatment, colors=mycol, - mode = 'markers', symbol = ~treatment, symbols = c('square','circle'), - marker = list(opacity = .8,line=list(color = 'darkblue',width = 1)) - ) -p3d <- p3d %>% layout(legend=list(y=.5)) -p3d <- p3d %>% config(toImageButtonOptions=list(format='svg',filename='3Dnmds', height= 500, width= 800, scale= 1)) -p3d - -} else { - print("No Convergence") -} -``` -
-
-
-
-
- -

  OTU Abundance Per Sample

- -```{r long, echo=FALSE} -dataz <- decostand(data5[,2:cols],method="total") #method 'total' normalizes data to sum up to 1 -dataz$sample <- data5$sample -datay <- merge(dataz, metadata, by="sample") -datalong <- datay %>% - tidyr::gather(first:last, key=hit, value=reads) - -##Barplot -## add better colors -#spec_bar <- ggplot(datalong, aes(x=forcats::fct_reorder(timepoint, as.numeric(as.character(timepoint))), -# y=reads, fill=hit))+ #Consider sqrt-transforming data -# geom_bar(aes(), stat="identity", position="fill")+ -# #coord_polar("y", start=0)+ -# theme_classic()+ -# facet_wrap(colony~treatment, nrow=2)+ -# #coord_flip()+ -# labs(x="Timepoint") #+ theme(legend.position = "none") -#spec_bar <- ggplot(datalong, aes(x=sample,y=reads,fill=hit))+geom_bar(aes(), stat="identity", position="fill")+ -# theme(axis.text.x=element_text(angle=90)) -#ggplotly(spec_bar) -#,'#00b300','#00b3b3','#0059b3','#6600ff','#b800e6','#ff3333','#ff8000','#ffff00','#bf8040','#42bcf5','#b428f5','#2e8d7e','#664e7e','#a4c700','#1aa3ff' - -ddd <- plot_ly(datalong, x=~sample, y=~reads, color=~hit, colors=mycol) -ddd <- ddd %>% layout(type='bar',barmode = 'stack') -ddd <- ddd %>% layout(legend = list(x=10,y=.5), xaxis=list(title = "Sample"), yaxis=list(title = "Relative abundance")) -ddd <- ddd %>% config(toImageButtonOptions=list(format='svg',filename='Relative_abundance', height= 500, width= 800, scale= 1)) -ddd -``` -
-
-
-
-
- -

  OTU Abundance Per Treatment

- -```{r asv_barplot, echo=FALSE} -datalong <- datalong %>% - filter(reads>0) - -#asv_bar <- ggplot(datalong, aes(x=reorder(hit,reads), y=reads, fill=treatment))+ -# geom_bar(stat="identity")+ -# scale_fill_manual(values=c('#088da5','#e19336'))+ #colors not working -# coord_flip()+ -# theme_classic()+ -# theme(axis.title.y = element_blank())+ -# theme(legend.title = element_blank()) - -asp2 <- plot_ly(datalong, y=~hit, x=~reads, color=~treatment, colors=mycol,text = ~paste("Sample: ", sample), opacity=.9) -asp2 <- asp2 %>% layout(type='bar', barmode = 'group') -asp2 <- asp2 %>% layout(yaxis = list(title = '', categoryorder = "total ascending"), legend = list(x=10,y=.5)) -asp2 <- asp2 %>% config(toImageButtonOptions=list(format='svg',filename='Most_abundant_hits_per_sample', height= 500, width= 800, scale= 1)) -asp2 -``` -
-
-
-
-
- -

  Pairwise Percent-ID Heatmap

- -
-
-```{r heatmap, echo=FALSE} -heatdata=params$heatmap -#heatdata="a.csv" -#heatdata="PVID_vAMPtest1_otu.85_PercentID.matrix" -simmatrix<- read.csv(heatdata, header=FALSE) -rownames(simmatrix) <- simmatrix[,1] -simmatrix <- simmatrix[,-1] -colnames(simmatrix) <-rownames(simmatrix) -cols <- dim(simmatrix)[2] -simmatrix$AA <- rownames(simmatrix) -rval=nrow(simmatrix) -simmatrix2 <- simmatrix %>% - gather(1:rval, key=sequence, value=similarity) -x=reorder(simmatrix2$AA,simmatrix2$similarity) -y=reorder(simmatrix2$sequence,simmatrix2$similarity) -similaritymatrix <- ggplot(simmatrix2, aes(x=x, y=y,fill=similarity))+ - geom_raster()+ - scale_fill_distiller(palette="Spectral")+ - theme(axis.text.x = element_text(angle = 90))+ - theme(axis.title.x=element_blank())+ - theme(axis.title.y=element_blank()) - -heat <- ggplotly(similaritymatrix) -heat <- heat %>% config(toImageButtonOptions=list(format='svg',filename='heatmap', height= 500, width= 800, scale= 1)) -heat -``` -
-
-
-
-
- - -

  Taxonomy Result Visualization

- -
-
- -```{r taxonomy, echo=FALSE} -tax=read.csv(params$tax,header=F) -#tax=read.csv("tax.csv",header=F) -#tax=read.csv("PVID_vAMPtest1_otu.85_summary_for_plot.csv", header=F) -tp <- plot_ly(tax, labels = ~V1, values = ~V2) -tp <- tp %>% add_pie(marker=list(colors=mycol, line=list(color='#000000', width=.5)), hole = 0.6) -tp <- tp %>% layout(title = "Taxonomy distribution", showlegend = F, - xaxis = list(showgrid = FALSE, zeroline = FALSE, showticklabels = FALSE), - yaxis = list(showgrid = FALSE, zeroline = FALSE, showticklabels = FALSE) - ) -tp <- tp %>% config(toImageButtonOptions=list(format='svg',filename='TaxDonut', height= 500, width= 800, scale= 1)) -tp -``` - -
-
-
-
-
- -

  Phylogenetic Tree

- -
-
- -```{r tree, echo=FALSE} -#tree=read.newick("vAMPrun_otu.55.raxml.support") -tree=read.tree(params$tree) -p1 <- ggtree(tree) -id <- tree$tip.label -dat <- tibble::tibble(id = id) -metat <- p1$data %>% dplyr::inner_join(dat, c('label' = 'id')) -p2 <- p1 + geom_point(data = metat, aes(x = x, y = y, label = id)) -ggplotly(p2, tooltip = "label") -``` -This tree is a maximum likelihood tree made with IQTREE2 and the parameters you specified in the vampirus.config file. Also, this is an interactive tree, you can zoom in and hover on nodes to know the sequence ID. For a better visualization of this tree, you can find the *.treefile with bootstrap support values within the results directory and visualize using programs like FigTree or ITOL. -
-
-
-
-
-
-
-
-
diff --git a/vAMPirus.nf b/vAMPirus.nf index 7e339ee..2f6035e 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1916,8 +1916,8 @@ if (params.DataCheck || params.Analyze) { label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/ASV/${params.projtag}_asvMED_${params.asvC}", mode: "copy", overwrite: true - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/ASV/${params.projtag}_asvMED_${params.asvC}/OLIGO-REPRESENTATIVES", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/ASV/MED", mode: "copy", overwrite: true + input: file(asvs) from asv_for_med @@ -1925,9 +1925,10 @@ if (params.DataCheck || params.Analyze) { output: file("*_ASV_Grouping.csv") into asvgroupscsv file("${params.projtag}_ASV_group_reps_aligned.fasta") into groupreps + file("${params.projtag}_asvMED_${params.asvC}") script: - """ + """ #alignment mafft --thread ${task.cpus} --maxiterate 15000 --auto ${asvs} > ${params.projtag}_ASVs_mafftAlign.fasta #trimming @@ -1941,7 +1942,7 @@ if (params.DataCheck || params.Analyze) { oligotype ${params.projtag}_ASVs_Aligned_informativeonly.fasta ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY -o ${params.projtag}_asvMED_${params.asvC} -M 1 -c ${params.asvC} -N ${task.cpus} --skip-check-input --no-figures --skip-gen-html #generatemaps cd ./${params.projtag}_asvMED_${params.asvC}/OLIGO-REPRESENTATIVES/ - echo "ASV,Group,IDPattern" + echo "ASV,GroupID,IDPattern" j=1 for x in *_unique; do gid=$(echo \$x | awk -F "_" '{print \$1}') @@ -1965,7 +1966,7 @@ if (params.DataCheck || params.Analyze) { mv ${params.projtag}_ASV_Grouping.csv ../../ mv ${params.projtag}_ASV_group_reps_aligned.fasta ../../ cd .. - """ + """ } process ASV_MED_Reps_phylogeny { @@ -1983,32 +1984,32 @@ if (params.DataCheck || params.Analyze) { file("*iq.treefile") into asv_group_rep_tree script: - """ - # Protein_ModelTest - modeltest-ng -i ${reps} -p ${task.cpus} -o ${params.protag}_ASV_Group_Reps_mt -d aa -s 203 --disable-checkpoint + """ + # Protein_ModelTest + modeltest-ng -i ${reps} -p ${task.cpus} -o ${params.protag}_ASV_Group_Reps_mt -d aa -s 203 --disable-checkpoint - # Protein_Phylogeny - if [ "${params.iqCustomaa}" != "" ];then - iqtree -s ${reps} --prefix ${params.protag}_ASV_Group_Reps_iq --redo -T auto ${params.iqCustomaa} + # Protein_Phylogeny + if [ "${params.iqCustomaa}" != "" ];then + iqtree -s ${reps} --prefix ${params.protag}_ASV_Group_Reps_iq --redo -T auto ${params.iqCustomaa} - elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') - iqtree -s ${reps} --prefix ${params.protag}_ASV_Group_Reps_iq -m \${mod} --redo -nt auto -b ${params.boots} + elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then + mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') + iqtree -s ${reps} --prefix ${params.protag}_ASV_Group_Reps_iq -m \${mod} --redo -nt auto -b ${params.boots} - elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') - iqtree -s ${reps} --prefix ${params.protag}_ASV_Group_Reps_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni + elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then + mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') + iqtree -s ${reps} --prefix ${params.protag}_ASV_Group_Reps_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni - elif [ "${params.nonparametric}" != "false" ];then - iqtree -s ${reps} --prefix ${params.protag}_ASV_Group_Reps_iq -m MFP --redo -nt auto -b ${params.boots} + elif [ "${params.nonparametric}" != "false" ];then + iqtree -s ${reps} --prefix ${params.protag}_ASV_Group_Reps_iq -m MFP --redo -nt auto -b ${params.boots} - elif [ "${params.parametric}" != "false" ];then - iqtree -s ${reps} --prefix ${params.protag}_ASV_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + elif [ "${params.parametric}" != "false" ];then + iqtree -s ${reps} --prefix ${params.protag}_ASV_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - else - iqtree -s ${reps} --prefix ${params.protag}_ASV_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - fi - """ + else + iqtree -s ${reps} --prefix ${params.protag}_ASV_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + fi + """ } process Adding_ASV_MED_Info { @@ -2021,14 +2022,20 @@ if (params.DataCheck || params.Analyze) { file(counts) from asvcount_med file(tree) from asvphy_med file(map) from asvgroupscsv - output: + output: + file("${params.projtag}_ASV_Grouping_counts.csv") into asvgroupcounts script: - """ - - - """ + """ + awk -F "," '{print \$1}' ${counts} | sed '1d' > asv.list + echo "GroupID" >> group.list + for x in $(cat asv.list); + do group=$(grep -w \$x ${map} | awk -F "," '{print \$2}') + echo "\$group" >> group.list + done + paste -d',' group.list ${counts} > ${params.projtag}_ASV_Grouping_counts.csv + """ } } @@ -2534,6 +2541,40 @@ if (params.DataCheck || params.Analyze) { """ } + process Adding_AminoType_MED_Info { + + label 'low_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/MED/Phylogeny", mode: "copy", overwrite: true + + input: + file(counts) from asvcount_med + file(tree) from asvphy_med + file(map) from asvgroupscsv + + output: + file("${params.projtag}_AminoType_Grouping_counts.csv") into asvgroupcounts + + script: + #map + ASV1,GroupA,TAGCGAT + + #count + #ASvs, Sample1,sample2 + asv1,1,2 + + + + """ + awk -F "," '{print \$1}' ${counts} | sed '1d' > amino.list + echo "GroupID" >> group.list + for x in $(cat amino.list); + do group=$(grep -w \$x ${map} | awk -F "," '{print \$2}') + echo "\$group" >> group.list + done + paste -d',' group.list ${counts} > ${params.projtag}_AminoType_Grouping_counts.csv + """ + } } if (params.pcASV) { // ASV_nucl -> ASV_aa -> clusteraa by %id with ch-hit -> extract representative nucl sequences to generate new OTU file From 152b3fd0d35b97c73106fbaae5285e349debf42f Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Fri, 26 Mar 2021 17:50:26 -0500 Subject: [PATCH 062/223] fixed --- vAMPirus.nf | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 2f6035e..d89887b 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1945,8 +1945,8 @@ if (params.DataCheck || params.Analyze) { echo "ASV,GroupID,IDPattern" j=1 for x in *_unique; - do gid=$(echo \$x | awk -F "_" '{print \$1}') - uni=$(echo \$x | awk -F ""\${gid}"_" '{print \$2}' | awk -F "_uni" '{print \$1}') + do gid=\$(echo \$x | awk -F "_" '{print \$1}') + uni=\$(echo \$x | awk -F ""\${gid}"_" '{print \$2}' | awk -F "_uni" '{print \$1}') grep ">" "\$gid"_"\$uni" | awk -F ">" '{print \$2}' > asv.list seqtk subseq ${asvs} asv.list > Group"\${j}"_sequences.fasta for z in $( cat asv.list) From f738e93fe1fbac00c2cb0e6186f3708f9f4d2560 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Fri, 26 Mar 2021 17:51:37 -0500 Subject: [PATCH 063/223] edits --- vAMPirus.nf | 6 +++--- 1 file changed, 3 insertions(+), 3 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index d89887b..42bbeac 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1949,7 +1949,7 @@ if (params.DataCheck || params.Analyze) { uni=\$(echo \$x | awk -F ""\${gid}"_" '{print \$2}' | awk -F "_uni" '{print \$1}') grep ">" "\$gid"_"\$uni" | awk -F ">" '{print \$2}' > asv.list seqtk subseq ${asvs} asv.list > Group"\${j}"_sequences.fasta - for z in $( cat asv.list) + for z in \$( cat asv.list) do echo ""\$z",Group"\$gid","\$uni"" >> ${params.projtag}_ASV_Grouping.csv done @@ -2030,8 +2030,8 @@ if (params.DataCheck || params.Analyze) { """ awk -F "," '{print \$1}' ${counts} | sed '1d' > asv.list echo "GroupID" >> group.list - for x in $(cat asv.list); - do group=$(grep -w \$x ${map} | awk -F "," '{print \$2}') + for x in \$(cat asv.list); + do group=\$(grep -w \$x ${map} | awk -F "," '{print \$2}') echo "\$group" >> group.list done paste -d',' group.list ${counts} > ${params.projtag}_ASV_Grouping_counts.csv From ec6ea3c95a60a3cdb5c9407c23d0e772cc37c3df Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Fri, 26 Mar 2021 17:55:12 -0500 Subject: [PATCH 064/223] pushing --- vAMPirus.nf | 27 +++++++++------------------ 1 file changed, 9 insertions(+), 18 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 42bbeac..5a78ed1 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -2357,7 +2357,7 @@ if (params.DataCheck || params.Analyze) { output: tuple file("*_aln.fasta"), file("*_aln.html"), file("*.log"), file("*iq*"), file("*mt*") into alignprot_results - file("*iq.treefile") into amino_rax_plot + file("*iq.treefile") into (amino_rax_plot, amino_repphy) script: """ @@ -2407,7 +2407,7 @@ if (params.DataCheck || params.Analyze) { output: tuple file("*_AminoType_counts.csv"), file("*dmd.out") into counts_summary - file("*_AminoType_counts.csv") into aminocounts_plot + file("*_AminoType_counts.csv") into (aminocounts_plot, aminocountmed) script: """ @@ -2473,8 +2473,8 @@ if (params.DataCheck || params.Analyze) { echo "AminoType,Group,IDPattern" j=1 for x in *_unique; - do gid=$(echo \$x | awk -F "_" '{print \$1}') - uni=$(echo \$x | awk -F ""\${gid}"_" '{print \$2}' | awk -F "_uni" '{print \$1}') + do gid=\$(echo \$x | awk -F "_" '{print \$1}') + uni=\$(echo \$x | awk -F ""\${gid}"_" '{print \$2}' | awk -F "_uni" '{print \$1}') grep ">" "\$gid"_"\$uni" | awk -F ">" '{print \$2}' > asv.list seqtk subseq ${aminos} asv.list > Group"\${j}"_sequences.fasta for z in $( cat asv.list) @@ -2548,28 +2548,19 @@ if (params.DataCheck || params.Analyze) { publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/MED/Phylogeny", mode: "copy", overwrite: true input: - file(counts) from asvcount_med - file(tree) from asvphy_med - file(map) from asvgroupscsv + file(counts) from aminocountmed + file(tree) from amino_repphy + file(map) from atygroupscsv output: file("${params.projtag}_AminoType_Grouping_counts.csv") into asvgroupcounts script: - #map - ASV1,GroupA,TAGCGAT - - #count - #ASvs, Sample1,sample2 - asv1,1,2 - - - """ awk -F "," '{print \$1}' ${counts} | sed '1d' > amino.list echo "GroupID" >> group.list - for x in $(cat amino.list); - do group=$(grep -w \$x ${map} | awk -F "," '{print \$2}') + for x in \$(cat amino.list); + do group=\$(grep -w \$x ${map} | awk -F "," '{print \$2}') echo "\$group" >> group.list done paste -d',' group.list ${counts} > ${params.projtag}_AminoType_Grouping_counts.csv From dda3e248439e8ec625a00b8c1e10f71375e7b402 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Fri, 26 Mar 2021 17:56:42 -0500 Subject: [PATCH 065/223] edit --- vAMPirus.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 5a78ed1..0579b4a 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -2477,7 +2477,7 @@ if (params.DataCheck || params.Analyze) { uni=\$(echo \$x | awk -F ""\${gid}"_" '{print \$2}' | awk -F "_uni" '{print \$1}') grep ">" "\$gid"_"\$uni" | awk -F ">" '{print \$2}' > asv.list seqtk subseq ${aminos} asv.list > Group"\${j}"_sequences.fasta - for z in $( cat asv.list) + for z in \$( cat asv.list) do echo ""\$z",Group"\$gid","\$uni"" >> ${params.projtag}_AminoType_Grouping.csv done From 7147feb820a14efdd9aa05be132ad2d7f7d6159b Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Fri, 26 Mar 2021 18:01:13 -0500 Subject: [PATCH 066/223] try --- vAMPirus.nf | 6 +++--- 1 file changed, 3 insertions(+), 3 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 0579b4a..25f254f 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -2016,7 +2016,7 @@ if (params.DataCheck || params.Analyze) { label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/MED/Phylogeny", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/MED/", mode: "copy", overwrite: true input: file(counts) from asvcount_med @@ -2545,7 +2545,7 @@ if (params.DataCheck || params.Analyze) { label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/MED/Phylogeny", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/MED/", mode: "copy", overwrite: true input: file(counts) from aminocountmed @@ -2553,7 +2553,7 @@ if (params.DataCheck || params.Analyze) { file(map) from atygroupscsv output: - file("${params.projtag}_AminoType_Grouping_counts.csv") into asvgroupcounts + file("${params.projtag}_AminoType_Grouping_counts.csv") into amino_groupcounts script: """ From c2c91cec716a2314dd3137592ec76dfaf1343d5f Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Fri, 26 Mar 2021 18:03:17 -0500 Subject: [PATCH 067/223] yaya --- vAMPirus.nf | 46 +++++++++++++++++++++++----------------------- 1 file changed, 23 insertions(+), 23 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 25f254f..04b7a09 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1980,7 +1980,7 @@ if (params.DataCheck || params.Analyze) { file(reps) from groupreps output: - tuple file("*_Aminotype_Group_Reps") into align_results_asvmed + file("*_Aminotype_Group_Reps") into align_results_asvmed file("*iq.treefile") into asv_group_rep_tree script: @@ -2509,36 +2509,36 @@ if (params.DataCheck || params.Analyze) { file(reps) from atygroupreps output: - tuple file("*_Aminotype_Group_Reps") into align_results_aminmed - file("*iq.treefile") into amino_group_rep_tree + file("*_Aminotype_Group_Reps") into align_results_aminmed + file("*iq.treefile") into amino_group_rep_tree script: - """ - # Protein_ModelTest - modeltest-ng -i ${reps} -p ${task.cpus} -o ${params.protag}_AminoType_Group_Reps_mt -d aa -s 203 --disable-checkpoint + """ + # Protein_ModelTest + modeltest-ng -i ${reps} -p ${task.cpus} -o ${params.protag}_AminoType_Group_Reps_mt -d aa -s 203 --disable-checkpoint - # Protein_Phylogeny - if [ "${params.iqCustomaa}" != "" ];then - iqtree -s ${reps} --prefix ${params.protag}_AminoType_Group_Reps_iq --redo -T auto ${params.iqCustomaa} + # Protein_Phylogeny + if [ "${params.iqCustomaa}" != "" ];then + iqtree -s ${reps} --prefix ${params.protag}_AminoType_Group_Reps_iq --redo -T auto ${params.iqCustomaa} - elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') - iqtree -s ${reps} --prefix ${params.protag}_AminoType_Group_Reps_iq -m \${mod} --redo -nt auto -b ${params.boots} + elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then + mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') + iqtree -s ${reps} --prefix ${params.protag}_AminoType_Group_Reps_iq -m \${mod} --redo -nt auto -b ${params.boots} - elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') - iqtree -s ${reps} --prefix ${params.protag}_AminoType_Group_Reps_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni + elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then + mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') + iqtree -s ${reps} --prefix ${params.protag}_AminoType_Group_Reps_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni - elif [ "${params.nonparametric}" != "false" ];then - iqtree -s ${reps} --prefix ${params.protag}_AminoType_Group_Reps_iq -m MFP --redo -nt auto -b ${params.boots} + elif [ "${params.nonparametric}" != "false" ];then + iqtree -s ${reps} --prefix ${params.protag}_AminoType_Group_Reps_iq -m MFP --redo -nt auto -b ${params.boots} - elif [ "${params.parametric}" != "false" ];then - iqtree -s ${reps} --prefix ${params.protag}_AminoType_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + elif [ "${params.parametric}" != "false" ];then + iqtree -s ${reps} --prefix ${params.protag}_AminoType_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - else - iqtree -s ${reps} --prefix ${params.protag}_AminoType_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - fi - """ + else + iqtree -s ${reps} --prefix ${params.protag}_AminoType_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + fi + """ } process Adding_AminoType_MED_Info { From 041d4777f1b31055eb35c4f0daeec79cf6982df4 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Fri, 26 Mar 2021 18:06:11 -0500 Subject: [PATCH 068/223] yayay --- vAMPirus.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 04b7a09..e4d96b6 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -2460,7 +2460,7 @@ if (params.DataCheck || params.Analyze) { #alignment mafft --thread ${task.cpus} --maxiterate 15000 --auto ${aminos} > ${params.projtag}_AminoTypes_mafftAlign.fasta #trimming - trimal -in ${params.projtag}_AmminoTypes_mafftAlign.fasta -out ${params.projtag}_AminoTypes_mafftAligned.fasta -keepheader -fasta -automated1 + trimal -in ${params.projtag}_AminoTypes_mafftAlign.fasta -out ${params.projtag}_AminoTypes_mafftAligned.fasta -keepheader -fasta -automated1 rm ${params.projtag}_AminoTypes_mafftAlign.fasta o-trim-uninformative-columns-from-alignment ${params.projtag}_AminoTypes_mafftAligned.fasta mv ${params.projtag}_AminoTypes_mafftAligned.fasta-TRIMMED ./${params.projtag}_AminoTypes_Aligned_informativeonly.fasta From fbd47899a7221a45852feda7d68419cebf45e089 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Fri, 26 Mar 2021 18:12:18 -0500 Subject: [PATCH 069/223] fixed seqtk --- vAMPirus.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index e4d96b6..244ed75 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -2476,7 +2476,7 @@ if (params.DataCheck || params.Analyze) { do gid=\$(echo \$x | awk -F "_" '{print \$1}') uni=\$(echo \$x | awk -F ""\${gid}"_" '{print \$2}' | awk -F "_uni" '{print \$1}') grep ">" "\$gid"_"\$uni" | awk -F ">" '{print \$2}' > asv.list - seqtk subseq ${aminos} asv.list > Group"\${j}"_sequences.fasta + seqtk subseq ../../${aminos} asv.list > Group"\${j}"_sequences.fasta for z in \$( cat asv.list) do echo ""\$z",Group"\$gid","\$uni"" >> ${params.projtag}_AminoType_Grouping.csv From c735ed1fb33dc6fb15493acab955239f86352d39 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Fri, 26 Mar 2021 18:16:24 -0500 Subject: [PATCH 070/223] aminoadded --- vAMPirus.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 244ed75..3867f72 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -2482,7 +2482,7 @@ if (params.DataCheck || params.Analyze) { done rm asv.list - echo "Group\${j}" >> ${params.projtag}_group_reps_aligned.fasta + echo "Group\${j}" >> ${params.projtag}_AminoType_group_reps_aligned.fasta echo "\$uni" > group.list seqtk subseq ../OLIGO-REPRESENTATIVES.fasta group.list > group.fasta tail -1 group.fasta >> ${params.projtag}_group_reps_aligned.fasta From dcccd0c77660201d38b4ddc66cbc9887fdaa1e68 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Fri, 26 Mar 2021 18:22:59 -0500 Subject: [PATCH 071/223] fix --- vAMPirus.nf | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 3867f72..fd550d8 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -2482,10 +2482,10 @@ if (params.DataCheck || params.Analyze) { done rm asv.list - echo "Group\${j}" >> ${params.projtag}_AminoType_group_reps_aligned.fasta + echo ">Group\${j}" >> ${params.projtag}_AminoType_group_reps_aligned.fasta echo "\$uni" > group.list seqtk subseq ../OLIGO-REPRESENTATIVES.fasta group.list > group.fasta - tail -1 group.fasta >> ${params.projtag}_group_reps_aligned.fasta + tail -1 group.fasta >> ${params.projtag}_AminoType_group_reps_aligned.fasta mv "\$gid"_"\$uni" ./Group"\$j"_"\$uni"_aligned.fasta mv "\$gid"_"\$uni"_unique ./Group"\$j"_"\$uni"_unqiues_aligned.fasta rm "\$gid"*.cPickle From bf891d0ef48948aff5f63a72ed69a568b4b3e909 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Fri, 26 Mar 2021 18:26:02 -0500 Subject: [PATCH 072/223] fixed asv med --- vAMPirus.nf | 6 +++--- 1 file changed, 3 insertions(+), 3 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index fd550d8..a03986b 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1948,16 +1948,16 @@ if (params.DataCheck || params.Analyze) { do gid=\$(echo \$x | awk -F "_" '{print \$1}') uni=\$(echo \$x | awk -F ""\${gid}"_" '{print \$2}' | awk -F "_uni" '{print \$1}') grep ">" "\$gid"_"\$uni" | awk -F ">" '{print \$2}' > asv.list - seqtk subseq ${asvs} asv.list > Group"\${j}"_sequences.fasta + seqtk subseq ../../${asvs} asv.list > Group"\${j}"_sequences.fasta for z in \$( cat asv.list) do echo ""\$z",Group"\$gid","\$uni"" >> ${params.projtag}_ASV_Grouping.csv done rm asv.list - echo "Group\${j}" >> ${params.projtag}_group_reps_aligned.fasta + echo "Group\${j}" >> ${params.projtag}_ASV_group_reps_aligned.fasta echo "\$uni" > group.list seqtk subseq ../OLIGO-REPRESENTATIVES.fasta group.list > group.fasta - tail -1 group.fasta >> ${params.projtag}_group_reps_aligned.fasta + tail -1 group.fasta >> ${params.projtag}_ASV_group_reps_aligned.fasta mv "\$gid"_"\$uni" ./Group"\$j"_"\$uni"_aligned.fasta mv "\$gid"_"\$uni"_unique ./Group"\$j"_"\$uni"_unqiues_aligned.fasta rm "\$gid"*.cPickle From 48e3e67c6639bba0d98635438f494880787bdda4 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Fri, 26 Mar 2021 18:30:43 -0500 Subject: [PATCH 073/223] fix --- vAMPirus.nf | 32 ++++++++++++++++---------------- 1 file changed, 16 insertions(+), 16 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index a03986b..e4d244c 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1980,34 +1980,34 @@ if (params.DataCheck || params.Analyze) { file(reps) from groupreps output: - file("*_Aminotype_Group_Reps") into align_results_asvmed + file("*_Aminotype_Group_Reps*") into align_results_asvmed file("*iq.treefile") into asv_group_rep_tree script: """ # Protein_ModelTest - modeltest-ng -i ${reps} -p ${task.cpus} -o ${params.protag}_ASV_Group_Reps_mt -d aa -s 203 --disable-checkpoint + modeltest-ng -i ${reps} -p ${task.cpus} -o ${params.projtag}_ASV_Group_Reps_mt -d aa -s 203 --disable-checkpoint # Protein_Phylogeny if [ "${params.iqCustomaa}" != "" ];then - iqtree -s ${reps} --prefix ${params.protag}_ASV_Group_Reps_iq --redo -T auto ${params.iqCustomaa} + iqtree -s ${reps} --prefix ${params.projtag}_ASV_Group_Reps_iq --redo -T auto ${params.iqCustomaa} elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') - iqtree -s ${reps} --prefix ${params.protag}_ASV_Group_Reps_iq -m \${mod} --redo -nt auto -b ${params.boots} + iqtree -s ${reps} --prefix ${params.projtag}_ASV_Group_Reps_iq -m \${mod} --redo -nt auto -b ${params.boots} elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') - iqtree -s ${reps} --prefix ${params.protag}_ASV_Group_Reps_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni + iqtree -s ${reps} --prefix ${params.projtag}_ASV_Group_Reps_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni elif [ "${params.nonparametric}" != "false" ];then - iqtree -s ${reps} --prefix ${params.protag}_ASV_Group_Reps_iq -m MFP --redo -nt auto -b ${params.boots} + iqtree -s ${reps} --prefix ${params.projtag}_ASV_Group_Reps_iq -m MFP --redo -nt auto -b ${params.boots} elif [ "${params.parametric}" != "false" ];then - iqtree -s ${reps} --prefix ${params.protag}_ASV_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + iqtree -s ${reps} --prefix ${params.projtag}_ASV_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni else - iqtree -s ${reps} --prefix ${params.protag}_ASV_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + iqtree -s ${reps} --prefix ${params.projtag}_ASV_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni fi """ } @@ -2509,34 +2509,34 @@ if (params.DataCheck || params.Analyze) { file(reps) from atygroupreps output: - file("*_Aminotype_Group_Reps") into align_results_aminmed + file("*_Aminotype_Group_Reps*") into align_results_aminmed file("*iq.treefile") into amino_group_rep_tree script: """ # Protein_ModelTest - modeltest-ng -i ${reps} -p ${task.cpus} -o ${params.protag}_AminoType_Group_Reps_mt -d aa -s 203 --disable-checkpoint + modeltest-ng -i ${reps} -p ${task.cpus} -o ${params.projtag}_AminoType_Group_Reps_mt -d aa -s 203 --disable-checkpoint # Protein_Phylogeny if [ "${params.iqCustomaa}" != "" ];then - iqtree -s ${reps} --prefix ${params.protag}_AminoType_Group_Reps_iq --redo -T auto ${params.iqCustomaa} + iqtree -s ${reps} --prefix ${params.projtag}_AminoType_Group_Reps_iq --redo -T auto ${params.iqCustomaa} elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') - iqtree -s ${reps} --prefix ${params.protag}_AminoType_Group_Reps_iq -m \${mod} --redo -nt auto -b ${params.boots} + iqtree -s ${reps} --prefix ${params.projtag}_AminoType_Group_Reps_iq -m \${mod} --redo -nt auto -b ${params.boots} elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') - iqtree -s ${reps} --prefix ${params.protag}_AminoType_Group_Reps_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni + iqtree -s ${reps} --prefix ${params.projtag}_AminoType_Group_Reps_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni elif [ "${params.nonparametric}" != "false" ];then - iqtree -s ${reps} --prefix ${params.protag}_AminoType_Group_Reps_iq -m MFP --redo -nt auto -b ${params.boots} + iqtree -s ${reps} --prefix ${params.projtag}_AminoType_Group_Reps_iq -m MFP --redo -nt auto -b ${params.boots} elif [ "${params.parametric}" != "false" ];then - iqtree -s ${reps} --prefix ${params.protag}_AminoType_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + iqtree -s ${reps} --prefix ${params.projtag}_AminoType_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni else - iqtree -s ${reps} --prefix ${params.protag}_AminoType_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + iqtree -s ${reps} --prefix ${params.projtag}_AminoType_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni fi """ } From 94aa5cebd5db16562629d4af16c138001377a01d Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Fri, 26 Mar 2021 18:32:11 -0500 Subject: [PATCH 074/223] fit --- vAMPirus.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index e4d244c..4a13427 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1954,7 +1954,7 @@ if (params.DataCheck || params.Analyze) { done rm asv.list - echo "Group\${j}" >> ${params.projtag}_ASV_group_reps_aligned.fasta + echo ">Group\${j}" >> ${params.projtag}_ASV_group_reps_aligned.fasta echo "\$uni" > group.list seqtk subseq ../OLIGO-REPRESENTATIVES.fasta group.list > group.fasta tail -1 group.fasta >> ${params.projtag}_ASV_group_reps_aligned.fasta From 01982f93d3faf5458dd3f53a62a3e1f51c4333ac Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Fri, 26 Mar 2021 18:35:55 -0500 Subject: [PATCH 075/223] push --- vAMPirus.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 4a13427..03e5935 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -2509,7 +2509,7 @@ if (params.DataCheck || params.Analyze) { file(reps) from atygroupreps output: - file("*_Aminotype_Group_Reps*") into align_results_aminmed + file("*_AminoType_Group_Reps*") into align_results_aminmed file("*iq.treefile") into amino_group_rep_tree script: From 21c667827f9fb7d3a36823b8970cc786471efa6d Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Fri, 26 Mar 2021 18:38:07 -0500 Subject: [PATCH 076/223] fixed out --- vAMPirus.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 03e5935..8429faa 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1980,7 +1980,7 @@ if (params.DataCheck || params.Analyze) { file(reps) from groupreps output: - file("*_Aminotype_Group_Reps*") into align_results_asvmed + file("*_ASV_Group_Reps*") into align_results_asvmed file("*iq.treefile") into asv_group_rep_tree script: From 237f1cda83eaa4fc550b3c8ae4e36db1cbb5598c Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Sun, 28 Mar 2021 19:02:48 -0500 Subject: [PATCH 077/223] revamped taxonomy processes and added classification --- vAMPirus.nf | 1431 ++++++++++++++++++++++++++----------------- vampirus_startup.sh | 16 +- 2 files changed, 870 insertions(+), 577 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 8429faa..d0956ea 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1295,7 +1295,9 @@ if (params.DataCheck || params.Analyze) { if (!params.skipTaxonomy) { - process Nucleotide_Taxonomy_Inference { /////// editttt + if (params.headers == "RefSeq") { + + process ncASV_Taxonomy_Inference { /////// editttt label 'high_cpus' @@ -1309,7 +1311,7 @@ if (params.DataCheck || params.Analyze) { output: file("*.fasta") into tax_labeled_fasta_ncasv - tuple file("*_phyloseqObject.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond_ncasv + tuple file("*_phyloformat.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond_ncasv tuple nid, file("*ncASV*summary_for_plot.csv") into taxplot_ncasv script: @@ -1328,18 +1330,14 @@ if (params.DataCheck || params.Analyze) { echo "[Percent ID (aa)]" > pid.list echo "[Organism ID]" > "\$name"_virus.list echo "[Gene]" > "\$name"_genes.list + echo "[ncASV#]" > otu.list + echo "[Sequence length]" > length.list grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst echo "extracting genes and names" touch new_"\$name"_asvnames.txt - j=1 - if [ `echo ${asvs} | grep -c "ASV"` -eq 1 ];then - for s in \$(cat seqids.lst);do - echo "[ASV#]" > otu.list - echo "[ASV sequence length]" > length.list - echo "Checking for \$s hit in diamond output" - if [[ ${params.refseq} == "T" ]];then - echo "RefSeq headers specified" - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + for s in \$(cat seqids.lst);do + echo "Checking for \$s hit in diamond output" + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then echo "Yep, there was a hit for \$s" echo "Extracting the information now:" acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') @@ -1355,48 +1353,9 @@ if (params.DataCheck || params.Analyze) { echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list - virus="NO" - gene="HIT" - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) + echo ">"\${s}"_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt echo "\$s done." - fi - else - echo "Using RVDB headers." - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - else + else echo "Ugh, there was no hit for \$s .." echo "We still love \$s though and we will add it to the final fasta file" echo "\$s" >> otu.list @@ -1409,14 +1368,10 @@ if (params.DataCheck || params.Analyze) { echo "NO_HIT" >> length.list virus="NO" gene="HIT" - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) + echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt echo "\$s done." - fi - fi - echo "Done with \$s" - done - fi + fi + done echo "Now editing "\$name" fasta headers" ###### rename_seq.py ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta @@ -1427,9 +1382,9 @@ if (params.DataCheck || params.Analyze) { echo " " > sequence.list grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - for x in *phyloseqObject.csv;do + for x in *phyloformat.csv;do echo "\$x" lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; @@ -1437,17 +1392,134 @@ if (params.DataCheck || params.Analyze) { cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; rm tmpcol.list tmp2col.list done - rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; - rm *asvnames.txt - rm "\$name"_virus.list - rm "\$name"_genes.list - rm newnames.list - rm access.list - echo "Taxonomy inferred for: ${asvs}" - rm headers.list + rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list """ - } + } + } else if (params.headers == "RVDB") { + + process ncASV_Taxonomy_Inference { /////// editttt + + label 'high_cpus' + + tag "${mtag}" + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Taxonomy", mode: "copy", overwrite: true, pattern: '*ncASV*.{fasta,csv,tsv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*ncASV*dmd.out' + + input: + tuple nid, file(asvs) from nuclFastas_forDiamond_ncasv_ch + + output: + file("*.fasta") into tax_labeled_fasta_ncasv + tuple file("*_phyloformat.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond_ncasv + tuple nid, file("*ncASV*summary_for_plot.csv") into taxplot_ncasv + tuple nid, file("*_quick_Taxbreakdown.csv") into tax_table_ncasv + + script: + mtag="ID=" + nid + """ + cp ${params.vampdir}/bin/rename_seq.py . + virdb=${params.dbdir}/${params.dbname} + grep ">" \${virdb} > headers.list + headers="headers.list" + name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') + diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + echo "Preparing lists to generate summary .csv's" + echo "[Best hit accession number]" > access.list + echo "[e-value]" > evalue.list + echo "[Bitscore]" > bit.list + echo "[Percent ID (aa)]" > pid.list + echo "[Organism ID]" > "\$name"_virus.list + echo "[Gene]" > "\$name"_genes.list + echo "[ncASV#]" > otu.list + echo "[Sequence length]" > length.list + grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst + if [[ ${params.lsa} == "T" ]] + then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list + echo "[Classification]" > classfication.list + else echo "skipped" >> \${name}_quick_Taxbreakdown.csv + fi + echo "extracting genes and names" + touch new_"\$name"_asvnames.txt + for s in \$(cat seqids.lst);do + echo "Using RVDB headers." + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + if [[ ${params.lca} == "T" ]] + then group=$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}' + lcla=$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') + echo "\$lcla" >> classfication.list + fi + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list + virus="NO" + gene="HIT" + echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + if [[ ${params.lca} == "T" ]] + then echo "N/A" >> classfication.list + fi + echo "\$s done." + fi + echo "Done with \$s" + done + echo "Now editing "\$name" fasta headers" + ###### rename_seq.py + ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta + echo "[Sequence header]" > newnames.list + cat new_"\$name"_asvnames.txt >> newnames.list + touch sequence.list + echo " " > sequence.list + grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list + rm "\$name"_tmpssasv.fasta + if [[ ${params.lca} == "T" ]] + then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list classfication.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + else paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + fi + if [[ ${params.lca} == "T" ]] + then paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv + fi + for x in *phyloformat.csv;do + echo "\$x" + lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) + tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; + sed 's/ /_/g' tmpcol.list > tmp2col.list; + cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; + rm tmpcol.list tmp2col.list + done + rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list + """ + } } + } + process Generate_ncASV_Counts_Table { @@ -1535,17 +1607,17 @@ if (params.DataCheck || params.Analyze) { if [ "${params.iqCustomnt}" != "" ];then iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq --redo -t \${pre}_mt.tree -T auto ${params.iqCustomnt} elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} + mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni + mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni elif [ "${params.nonparametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} elif [ "${params.parametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni else - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni fi """ } @@ -1557,240 +1629,225 @@ if (params.DataCheck || params.Analyze) { .into{ nuclFastas_forDiamond_asv_ch; nuclFastas_forCounts_asv_ch; nuclFastas_forphylogeny_asv; nuclFastas_forMatrix_asv_ch; asv_for_med } } - process ASV_Taxonomy_Inference { /////// editttt + if (!params.skipTaxonomy) { - label 'high_cpus' + if (params.headers == "RefSeq") { - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy", mode: "copy", overwrite: true, pattern: '*_ASV*.{fasta,csv,tsv}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*_ASV*dmd.out' + process ASV_Taxonomy_Inference { /////// editttt - input: - file(reads) from nuclFastas_forDiamond_asv_ch + label 'high_cpus' - output: - file("*.fasta") into tax_labeled_fasta_asv - tuple file("*_phyloseqObject.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond_asv - file("*_ASV*_summary_for_plot.csv") into taxplot_asv + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy", mode: "copy", overwrite: true, pattern: '*_ASV*.{fasta,csv,tsv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*_ASV*dmd.out' - script: - """ - cp ${params.vampdir}/bin/rename_seq.py . - virdb=${params.dbdir}/${params.dbname} - grep ">" \${virdb} > headers.list - headers="headers.list" - for filename in ${reads};do - name=\$(ls \${filename} | awk -F ".fasta" '{print \$1}') - diamond blastx -q \${filename} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 - echo "Preparing lists to generate summary .csv's" - echo "[Best hit accession number]" > access.list - echo "[e-value]" > evalue.list - echo "[Bitscore]" > bit.list - echo "[Percent ID (aa)]" > pid.list - echo "[Organism ID]" > "\$name"_virus.list - echo "[Gene]" > "\$name"_genes.list - grep ">" \${filename} | awk -F ">" '{print \$2}' > seqids.lst - echo "extracting genes and names" - touch new_"\$name"_asvnames.txt - j=1 - if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then + input: + file(asvs) from nuclFastas_forDiamond_asv_ch + + output: + file("*.fasta") into tax_labeled_fasta_asv + tuple file("*_phyloseqObject.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond_asv + file("*_ASV*_summary_for_plot.csv") into taxplot_asv + + script: + """ + cp ${params.vampdir}/bin/rename_seq.py . + virdb=${params.dbdir}/${params.dbname} + grep ">" \${virdb} > headers.list + headers="headers.list" + name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') + diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + echo "Preparing lists to generate summary .csv's" + echo "[Best hit accession number]" > access.list + echo "[e-value]" > evalue.list + echo "[Bitscore]" > bit.list + echo "[Percent ID (aa)]" > pid.list + echo "[Organism ID]" > "\$name"_virus.list + echo "[Gene]" > "\$name"_genes.list echo "[ASV#]" > otu.list - echo "[ASV sequence length]" > length.list - for s in \$(cat seqids.lst);do - echo "Checking for \$s hit in diamond output" - if [[ ${params.refseq} == "T" ]];then - echo "RefSeq headers specified" - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list - virus="NO" - gene="HIT" - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - fi - else - echo "Using RVDB headers." - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list - virus="NO" - gene="HIT" - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - fi - fi - echo "Done with \$s" - done - else + echo "[Sequence length]" > length.list + grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst + echo "extracting genes and names" + touch new_"\$name"_asvnames.txt for s in \$(cat seqids.lst);do - echo "[ASV#]" > otu.list - echo "[ASV sequence length]" > length.list echo "Checking for \$s hit in diamond output" - if [[ ${params.refseq} == "T" ]];then - echo "RefSeq headers specified" - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list - virus="NO" - gene="HIT" - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - fi + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">"\${s}"_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + echo "\$s done." else - echo "Using RVDB headers." - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list - virus="NO" - gene="HIT" - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - fi - fi - echo "Done with \$s" + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list + virus="NO" + gene="HIT" + echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + echo "\$s done." + fi done - fi - echo "Now editing "\$name" fasta headers" - ###### rename_seq.py - ./rename_seq.py \${filename} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta - awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta - echo "[Sequence header]" > newnames.list - cat new_"\$name"_asvnames.txt >> newnames.list - touch sequence.list - echo " " > sequence.list - grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list - rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - for x in *phyloseqObject.csv;do - echo "\$x" - lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) - tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; - sed 's/ /_/g' tmpcol.list > tmp2col.list; - cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; - rm tmpcol.list tmp2col.list - done - rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; - rm *asvnames.txt - rm "\$name"_virus.list - rm "\$name"_genes.list - rm newnames.list - rm access.list - echo "Taxonomy inferred for: \${filename} " - done - rm headers.list - """ + echo "Now editing "\$name" fasta headers" + ###### rename_seq.py + ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta + echo "[Sequence header]" > newnames.list + cat new_"\$name"_asvnames.txt >> newnames.list + touch sequence.list + echo " " > sequence.list + grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list + rm "\$name"_tmpssasv.fasta + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + for x in *phyloformat.csv;do + echo "\$x" + lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) + tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; + sed 's/ /_/g' tmpcol.list > tmp2col.list; + cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; + rm tmpcol.list tmp2col.list + done + rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list + """ + } + } else if (params.headers == "RVDB") { + + process ASV_Taxonomy_Inference { /////// editttt + + label 'high_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy", mode: "copy", overwrite: true, pattern: '*_ASV*.{fasta,csv,tsv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*_ASV*dmd.out' + + input: + file(asvs) from nuclFastas_forDiamond_asv_ch + + output: + file("*.fasta") into tax_labeled_fasta_asv + tuple file("*_phyloseqObject.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond_asv + file("*_ASV*_summary_for_plot.csv") into taxplot_asv + file("*_quick_Taxbreakdown.csv") into tax_table_asv + + script: + """ + cp ${params.vampdir}/bin/rename_seq.py . + virdb=${params.dbdir}/${params.dbname} + grep ">" \${virdb} > headers.list + headers="headers.list" + name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') + diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + echo "Preparing lists to generate summary .csv's" + echo "[Best hit accession number]" > access.list + echo "[e-value]" > evalue.list + echo "[Bitscore]" > bit.list + echo "[Percent ID (aa)]" > pid.list + echo "[Organism ID]" > "\$name"_virus.list + echo "[Gene]" > "\$name"_genes.list + echo "[ASV#]" > otu.list + echo "[Sequence length]" > length.list + grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst + if [[ ${params.lsa} == "T" ]] + then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list + echo "[Classification]" > classfication.list + else echo "skipped" >> \${name}_quick_Taxbreakdown.csv + fi + echo "extracting genes and names" + touch new_"\$name"_asvnames.txt + for s in \$(cat seqids.lst);do + echo "Using RVDB headers." + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + if [[ ${params.lca} == "T" ]] + then group=$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}' + lcla=$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') + echo "\$lcla" >> classfication.list + fi + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list + virus="NO" + gene="HIT" + echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + if [[ ${params.lca} == "T" ]] + then echo "N/A" >> classfication.list + fi + echo "\$s done." + fi + echo "Done with \$s" + done + echo "Now editing "\$name" fasta headers" + ###### rename_seq.py + ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta + echo "[Sequence header]" > newnames.list + cat new_"\$name"_asvnames.txt >> newnames.list + touch sequence.list + echo " " > sequence.list + grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list + rm "\$name"_tmpssasv.fasta + if [[ ${params.lca} == "T" ]] + then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list classfication.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + else paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + fi + if [[ ${params.lca} == "T" ]] + then paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv + fi + for x in *phyloformat.csv;do + echo "\$x" + lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) + tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; + sed 's/ /_/g' tmpcol.list > tmp2col.list; + cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; + rm tmpcol.list tmp2col.list + done + rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list + """ + } + } } process Generate_ASV_Counts_Tables { @@ -1888,23 +1945,25 @@ if (params.DataCheck || params.Analyze) { pre=\$(echo ${asvs} | awk -F ".fasta" '{print \$1}' ) mafft --thread ${task.cpus} --maxiterate 15000 --auto ${asvs} >\${pre}_ALN.fasta trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html + o-trim-uninformative-columns-from-alignment \${pre}_aln.fasta + mv \${pre}_aln.fasta-TRIMMED ./\${pre}_Aligned_informativeonly.fasta # Nucleotide_ModelTest - modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint + modeltest-ng -i \${pre}_Aligned_informativeonly.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint # Nucleotide_Phylogeny if [ "${params.iqCustomnt}" != "" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomnt} + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomnt} elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} + mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni + mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni elif [ "${params.nonparametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} elif [ "${params.parametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni else - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni fi """ } @@ -1916,7 +1975,7 @@ if (params.DataCheck || params.Analyze) { label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/ASV/MED", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/ASVs/MED", mode: "copy", overwrite: true input: @@ -2194,6 +2253,8 @@ if (params.DataCheck || params.Analyze) { if (!params.skipTaxonomy) { + if (params.headers == "RefSeq") { + process AminoType_Taxonomy_Inference { label 'high_cpus' @@ -2202,7 +2263,7 @@ if (params.DataCheck || params.Analyze) { publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Taxonomy", mode: "copy", overwrite: true, pattern: '*TaxonomyLabels.fasta' input: - file(reads) from aminotypesBlast + file(asvs) from aminotypesBlast output: tuple file("*_phyloseqObject.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AA_diamond @@ -2213,117 +2274,72 @@ if (params.DataCheck || params.Analyze) { """ cp ${params.vampdir}/bin/rename_seq.py . virdb=${params.dbdir}/${params.dbname} - grep ">" \${virdb} >> headers.list + grep ">" \${virdb} > headers.list headers="headers.list" - name=\$(ls ${reads} | awk -F "_noTaxonomy" '{print \$1}') - diamond blastp -q ${reads} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') + diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 echo "Preparing lists to generate summary .csv's" - echo "[Best hit accession number]" >access.list - echo "[AminoType sequence length]" >length.list - echo "[e-value]" >evalue.list - echo "[Bitscore]" >bit.list - echo "[Percent ID (aa)]" >pid.list - echo "[AminoType#]" >otu.list - echo "[Virus ID]" >"\$name"_virus.list - echo "[Gene]" >"\$name"_genes.list - grep ">" ${reads} | awk -F ">" '{print \$2}' > seqids.lst + echo "[Best hit accession number]" > access.list + echo "[e-value]" > evalue.list + echo "[Bitscore]" > bit.list + echo "[Percent ID (aa)]" > pid.list + echo "[Organism ID]" > "\$name"_virus.list + echo "[Gene]" > "\$name"_genes.list + echo "[AminoType#]" > otu.list + echo "[Sequence length]" > length.list + grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst echo "extracting genes and names" touch new_"\$name"_asvnames.txt - j=1 for s in \$(cat seqids.lst);do echo "Checking for \$s hit in diamond output" - if [[ ${params.refseq} == "T" ]];then - echo "RefSeq headers specified" - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then echo "Yep, there was a hit for \$s" echo "Extracting the information now:" acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') echo "\$s" >> otu.list echo "\$acc" >> access.list line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >>evalue.list - echo "\$line" | awk '{print \$11}' >>bit.list - echo "\$line" | awk '{print \$12}' >>pid.list - echo "\$line" | awk '{print \$2}' >>length.list + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') + gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">AminoType\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) + echo ">"\${s}"_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt echo "\$s done." - else + else echo "Ugh, there was no hit for \$s .." echo "We still love \$s though and we will add it to the final fasta file" echo "\$s" >> otu.list - echo "NO_HIT" >>access.list - echo "NO_HIT" >>"\$name"_genes.list - echo "NO_HIT" >>"\$name"_virus.list - echo "NO_HIT" >>evalue.list - echo "NO_HIT" >>bit.list - echo "NO_HIT" >>pid.list - echo "NO_HIT" >>length.list - virus="NO" - gene="HIT" - echo ">AminoType\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$\$1)) - echo "\$s done." - fi - else - echo "Using RVDB headers." - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') - echo "\$s" >>otu.list - echo "\$acc" >>access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >>evalue.list - echo "\$line" | awk '{print \$11}' >>bit.list - echo "\$line" | awk '{print \$12}' >>pid.list - echo "\$line" | awk '{print \$2}' >>length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') - echo "\$gene" | sed 's/_/ /g' >>"\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') - echo "\$virus" | sed 's/_/ /g' >>"\$name"_virus.list - echo ">AminoType\${j}_"\$virus"_"\$gene"" >>new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >>otu.list - echo "NO_HIT" >>access.list - echo "NO_HIT" >>"\$name"_genes.list - echo "NO_HIT" >>"\$name"_virus.list - echo "NO_HIT" >>evalue.list - echo "NO_HIT" >>bit.list - echo "NO_HIT" >>pid.list - echo "NO_HIT" >>length.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list virus="NO" gene="HIT" - echo ">AminoType\${j}_"\$virus"_"\$gene"" >>new_"\$name"_asvnames.txt - j=\$((\$j+1)) + echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt echo "\$s done." - fi - fi - echo "Done with \$s" + fi done echo "Now editing "\$name" fasta headers" ###### rename_seq.py - ./rename_seq.py ${reads} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta - awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta > "\$name"_tmpssasv.fasta + ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta echo "[Sequence header]" > newnames.list cat new_"\$name"_asvnames.txt >> newnames.list touch sequence.list echo " " > sequence.list grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - for x in *phyloseqObject.csv;do + for x in *phyloformat.csv;do echo "\$x" lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; @@ -2331,16 +2347,130 @@ if (params.DataCheck || params.Analyze) { cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; rm tmpcol.list tmp2col.list done - rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst headers.list otu.list ; - rm *asvnames.txt - rm "\$name"_virus.list - rm "\$name"_genes.list - rm newnames.list - rm access.list - echo "Taxonomy inferred for: ${reads} " + rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list """ + } + } else if (params.headers == "RVDB") { + + process AminoType_Taxonomy_Inference { + + label 'high_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{csv,tsv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Taxonomy", mode: "copy", overwrite: true, pattern: '*TaxonomyLabels.fasta' + + input: + file(asvs) from aminotypesBlast + + output: + tuple file("*_phyloseqObject.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AA_diamond + file("*_summary_for_plot.csv") into taxplot2 + file("*TaxonomyLabels.fasta") into tax_labeled_fasta2 + file("*_quick_Taxbreakdown.csv") into tax_table_amino + + script: + """ + cp ${params.vampdir}/bin/rename_seq.py . + virdb=${params.dbdir}/${params.dbname} + grep ">" \${virdb} > headers.list + headers="headers.list" + name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') + diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + echo "Preparing lists to generate summary .csv's" + echo "[Best hit accession number]" > access.list + echo "[e-value]" > evalue.list + echo "[Bitscore]" > bit.list + echo "[Percent ID (aa)]" > pid.list + echo "[Organism ID]" > "\$name"_virus.list + echo "[Gene]" > "\$name"_genes.list + echo "[AminoType#]" > otu.list + echo "[Sequence length]" > length.list + grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst + if [[ ${params.lsa} == "T" ]] + then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list + echo "[Classification]" > classfication.list + else echo "skipped" >> \${name}_quick_Taxbreakdown.csv + fi + echo "extracting genes and names" + touch new_"\$name"_asvnames.txt + for s in \$(cat seqids.lst);do + echo "Using RVDB headers." + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + if [[ ${params.lca} == "T" ]] + then group=$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}' + lcla=$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') + echo "\$lcla" >> classfication.list + fi + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list + virus="NO" + gene="HIT" + echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + if [[ ${params.lca} == "T" ]] + then echo "N/A" >> classfication.list + fi + echo "\$s done." + fi + echo "Done with \$s" + done + echo "Now editing "\$name" fasta headers" + ###### rename_seq.py + ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta + echo "[Sequence header]" > newnames.list + cat new_"\$name"_asvnames.txt >> newnames.list + touch sequence.list + echo " " > sequence.list + grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list + rm "\$name"_tmpssasv.fasta + if [[ ${params.lca} == "T" ]] + then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list classfication.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + else paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + fi + if [[ ${params.lca} == "T" ]] + then paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv + fi + for x in *phyloformat.csv;do + echo "\$x" + lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) + tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; + sed 's/ /_/g' tmpcol.list > tmp2col.list; + cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; + rm tmpcol.list tmp2col.list + done + rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list + """ } } + } if (!params.skipPhylogeny) { @@ -2691,6 +2821,8 @@ if (params.DataCheck || params.Analyze) { if (!params.skipTaxonomy) { + if (params.headers == "RefSeq") { + process pcASV_Nucleotide_Taxonomy_Inference { label 'high_cpus' @@ -2702,7 +2834,7 @@ if (params.DataCheck || params.Analyze) { publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{fasta}' input: - tuple nid, file(reads) from pcASV_ntDiamond_ch + tuple nid, file(asvs) from pcASV_ntDiamond_ch output: file("*.fasta") into ( pcASV_labeled ) @@ -2715,134 +2847,209 @@ if (params.DataCheck || params.Analyze) { set +e cp ${params.vampdir}/bin/rename_seq.py . virdb=${params.dbdir}/${params.dbname} - grep ">" \${virdb} >> headers.list + grep ">" \${virdb} > headers.list headers="headers.list" - name=\$(ls ${reads} | awk -F "_noTax" '{print \$1}') - diamond blastx -q ${reads} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') + diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 echo "Preparing lists to generate summary .csv's" - echo "[Best hit accession number]" >access.list - echo "[pcASV sequence length]" >length.list - echo "[e-value]" >evalue.list - echo "[Bitscore]" >bit.list - echo "[Percent ID (aa)]" >pid.list - echo "[pcASV#]" >otu.list - echo "[Virus ID]" >"\$name"_virus.list - echo "[Gene]" >"\$name"_genes.list - grep ">" ${reads} | awk -F ">" '{print \$2}' > seqids.lst + echo "[Best hit accession number]" > access.list + echo "[e-value]" > evalue.list + echo "[Bitscore]" > bit.list + echo "[Percent ID (aa)]" > pid.list + echo "[Organism ID]" > "\$name"_virus.list + echo "[Gene]" > "\$name"_genes.list + echo "[pcASV#]" > otu.list + echo "[Sequence length]" > length.list + grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst echo "extracting genes and names" - touch new_"\$name"_headers.txt - j=1 + touch new_"\$name"_asvnames.txt for s in \$(cat seqids.lst);do echo "Checking for \$s hit in diamond output" - if [[ ${params.refseq} == "T" ]];then - echo "RefSeq headers specified" - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then echo "Yep, there was a hit for \$s" echo "Extracting the information now:" acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') echo "\$s" >> otu.list echo "\$acc" >> access.list line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >>evalue.list - echo "\$line" | awk '{print \$11}' >>bit.list - echo "\$line" | awk '{print \$12}' >>pid.list - echo "\$line" | awk '{print \$2}' >>length.list + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list echo "Extracting virus and gene ID for \$s now" gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">pcASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_headers.txt - j=\$((\$j+1)) + echo ">"\${s}"_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt echo "\$s done." - else + else echo "Ugh, there was no hit for \$s .." echo "We still love \$s though and we will add it to the final fasta file" echo "\$s" >> otu.list - echo "NO_HIT" >>access.list - echo "NO_HIT" >>"\$name"_genes.list - echo "NO_HIT" >>"\$name"_virus.list - echo "NO_HIT" >>evalue.list - echo "NO_HIT" >>bit.list - echo "NO_HIT" >>pid.list - echo "NO_HIT" >>length.list - virus="NO" - gene="HIT" - echo ">pcASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_headers.txt - j=\$((\$j+1)) - echo "\$s done." - fi - else - echo "Using RVDB headers." - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') - echo "\$s" >>otu.list - echo "\$acc" >>access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >>evalue.list - echo "\$line" | awk '{print \$11}' >>bit.list - echo "\$line" | awk '{print \$12}' >>pid.list - echo "\$line" | awk '{print \$2}' >>length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >>"\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && - echo "\$virus" | sed 's/_/ /g' >>"\$name"_virus.list - echo ">pcASV\${j}_"\$virus"_"\$gene"" >>new_"\$name"_headers.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >>otu.list - echo "NO_HIT" >>access.list - echo "NO_HIT" >>"\$name"_genes.list - echo "NO_HIT" >>"\$name"_virus.list - echo "NO_HIT" >>evalue.list - echo "NO_HIT" >>bit.list - echo "NO_HIT" >>pid.list - echo "NO_HIT" >>length.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list virus="NO" gene="HIT" - echo ">pcASV\${j}_"\$virus"_"\$gene"" >>new_"\$name"_headers.txt - j=\$((\$j+1)) + echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt echo "\$s done." - fi - fi - echo "Done with \$s" + fi done echo "Now editing "\$name" fasta headers" ###### rename_seq.py - ./rename_seq.py ${reads} new_"\$name"_headers.txt "\$name"_TaxonomyLabels.fasta - awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta > "\$name"_tmpssasv.fasta + ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta echo "[Sequence header]" > newnames.list - cat new_"\$name"_headers.txt >> newnames.list + cat new_"\$name"_asvnames.txt >> newnames.list touch sequence.list echo " " > sequence.list grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - for x in *phyloseqObject.csv;do + for x in *phyloformat.csv;do echo "\$x" lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; sed 's/ /_/g' tmpcol.list > tmp2col.list; - cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_noTaxonomy_summary_for_plot.csv; + cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; rm tmpcol.list tmp2col.list done - rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; - rm "\$name"_virus.list - rm "\$name"_genes.list - rm newnames.list - rm access.list - echo "Taxonomy inferred for: ${reads} " - rm *headers.list + rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list """ + } + } else if (params.headers == "RVDB") { + + process pcASV_Nucleotide_Taxonomy_Inference { + + label 'high_cpus' + + tag "${mtag}" + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{csv,tsv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*dmd.{out}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{fasta}' + + input: + tuple nid, file(asvs) from pcASV_ntDiamond_ch + + output: + file("*.fasta") into ( pcASV_labeled ) + tuple file("*_phyloseqObject.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AAdiamond + tuple nid, file("*_summary_for_plot.csv") into taxplot3 + file("*_quick_Taxbreakdown.csv") into tax_table_pcasvnt + + script: + mtag="ID=" + nid + """ + set +e + cp ${params.vampdir}/bin/rename_seq.py . + virdb=${params.dbdir}/${params.dbname} + grep ">" \${virdb} > headers.list + headers="headers.list" + name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') + diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + echo "Preparing lists to generate summary .csv's" + echo "[Best hit accession number]" > access.list + echo "[e-value]" > evalue.list + echo "[Bitscore]" > bit.list + echo "[Percent ID (aa)]" > pid.list + echo "[Organism ID]" > "\$name"_virus.list + echo "[Gene]" > "\$name"_genes.list + echo "[pcASV#]" > otu.list + echo "[Sequence length]" > length.list + grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst + if [[ ${params.lsa} == "T" ]] + then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list + echo "[Classification]" > classfication.list + else echo "skipped" >> \${name}_quick_Taxbreakdown.csv + fi + echo "extracting genes and names" + touch new_"\$name"_asvnames.txt + for s in \$(cat seqids.lst);do + echo "Using RVDB headers." + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + if [[ ${params.lca} == "T" ]] + then group=$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}' + lcla=$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') + echo "\$lcla" >> classfication.list + fi + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list + virus="NO" + gene="HIT" + echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + if [[ ${params.lca} == "T" ]] + then echo "N/A" >> classfication.list + fi + echo "\$s done." + fi + echo "Done with \$s" + done + echo "Now editing "\$name" fasta headers" + ###### rename_seq.py + ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta + echo "[Sequence header]" > newnames.list + cat new_"\$name"_asvnames.txt >> newnames.list + touch sequence.list + echo " " > sequence.list + grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list + rm "\$name"_tmpssasv.fasta + if [[ ${params.lca} == "T" ]] + then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list classfication.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + else paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + fi + if [[ ${params.lca} == "T" ]] + then paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv + fi + for x in *phyloformat.csv;do + echo "\$x" + lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) + tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; + sed 's/ /_/g' tmpcol.list > tmp2col.list; + cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; + rm tmpcol.list tmp2col.list + done + rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list + """ } } + } + process Generate_Nucleotide_pcASV_Counts { @@ -3032,6 +3239,8 @@ if (params.DataCheck || params.Analyze) { if (!params.skipTaxonomy) { + if (params.headers == "RefSeq") { + process pcASV_AminoAcid_Taxonomy_Inference { label 'high_cpus' @@ -3043,7 +3252,7 @@ if (params.DataCheck || params.Analyze) { publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{fasta}' input: - tuple nid, file(reads) from pcASV_aaDiamond_ch + tuple nid, file(asvs) from pcASV_aaDiamond_ch output: file("*.fasta") into ( pcASV_labeledAA ) @@ -3055,116 +3264,72 @@ if (params.DataCheck || params.Analyze) { """ cp ${params.vampdir}/bin/rename_seq.py . virdb=${params.dbdir}/${params.dbname} - grep ">" \${virdb} >> headers.list + grep ">" \${virdb} > headers.list headers="headers.list" - name=\$(ls ${reads} | awk -F ".fasta" '{print \$1}') - diamond blastp -q ${reads} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') + diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 echo "Preparing lists to generate summary .csv's" - echo "[Best hit accession number]" >access.list - echo "[pcASV sequence length]" >length.list - echo "[e-value]" >evalue.list - echo "[Bitscore]" >bit.list - echo "[Percent ID (aa)]" >pid.list - echo "[pcASVaa#]" >otu.list - echo "[Virus ID]" >"\$name"_virus.list - echo "[Gene]" >"\$name"_genes.list - grep ">" ${reads} | awk -F ">" '{print \$2}' > seqids.lst + echo "[Best hit accession number]" > access.list + echo "[e-value]" > evalue.list + echo "[Bitscore]" > bit.list + echo "[Percent ID (aa)]" > pid.list + echo "[Organism ID]" > "\$name"_virus.list + echo "[Gene]" > "\$name"_genes.list + echo "[pcASV#]" > otu.list + echo "[Sequence length]" > length.list + grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst echo "extracting genes and names" - touch new_"\$name"_headers.txt - j=1 + touch new_"\$name"_asvnames.txt for s in \$(cat seqids.lst);do echo "Checking for \$s hit in diamond output" - if [[ ${params.refseq} == "T" ]];then - echo "RefSeq headers specified" - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then echo "Yep, there was a hit for \$s" echo "Extracting the information now:" acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') echo "\$s" >> otu.list echo "\$acc" >> access.list line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >>evalue.list - echo "\$line" | awk '{print \$11}' >>bit.list - echo "\$line" | awk '{print \$12}' >>pid.list - echo "\$line" | awk '{print \$2}' >>length.list + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list echo "Extracting virus and gene ID for \$s now" gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">pcASVaa\${j}_"\$virus"_"\$gene"" >> new_"\$name"_headers.txt - j=\$((\$j+1)) + echo ">"\${s}"_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt echo "\$s done." - else + else echo "Ugh, there was no hit for \$s .." echo "We still love \$s though and we will add it to the final fasta file" echo "\$s" >> otu.list - echo "NO_HIT" >>access.list - echo "NO_HIT" >>"\$name"_genes.list - echo "NO_HIT" >>"\$name"_virus.list - echo "NO_HIT" >>evalue.list - echo "NO_HIT" >>bit.list - echo "NO_HIT" >>pid.list - echo "NO_HIT" >>length.list - virus="NO" - gene="HIT" - echo ">pcASVaa\${j}_"\$virus"_"\$gene"" >> new_"\$name"_headers.txt - j=\$((\$j+1)) - echo "\$s done." - fi - else - echo "Using RVDB headers." - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') - echo "\$s" >>otu.list - echo "\$acc" >>access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >>evalue.list - echo "\$line" | awk '{print \$11}' >>bit.list - echo "\$line" | awk '{print \$12}' >>pid.list - echo "\$line" | awk '{print \$2}' >>length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >>"\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && - echo "\$virus" | sed 's/_/ /g' >>"\$name"_virus.list - echo ">pcASVaa\${j}_"\$virus"_"\$gene"" >>new_"\$name"_headers.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >>otu.list - echo "NO_HIT" >>access.list - echo "NO_HIT" >>"\$name"_genes.list - echo "NO_HIT" >>"\$name"_virus.list - echo "NO_HIT" >>evalue.list - echo "NO_HIT" >>bit.list - echo "NO_HIT" >>pid.list - echo "NO_HIT" >>length.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list virus="NO" gene="HIT" - echo ">pcASVaa\${j}_\${virus}_\${gene}" >>new_"\$name"_headers.txt - j=\$((\$j+1)) + echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt echo "\$s done." - fi - fi - echo "Done with \$s" + fi done echo "Now editing "\$name" fasta headers" ###### rename_seq.py - ./rename_seq.py ${reads} new_"\$name"_headers.txt "\$name"_wTax.fasta - awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_wTax.fasta > "\$name"_tmpssasv.fasta + ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta echo "[Sequence header]" > newnames.list - cat new_"\$name"_headers.txt >> newnames.list + cat new_"\$name"_asvnames.txt >> newnames.list touch sequence.list - awk 'BEGIN{RS=">";ORS=""}{print \$2"\\n"}' \${name}_tmpssasv.fasta >>sequence.list + echo " " > sequence.list + grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_summary_phyloseqObject.csv - paste -d"\\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - for x in *phyloseqObject.csv;do + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + for x in *phyloformat.csv;do echo "\$x" lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; @@ -3172,16 +3337,134 @@ if (params.DataCheck || params.Analyze) { cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; rm tmpcol.list tmp2col.list done - rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; - rm "\$name"_virus.list - rm "\$name"_genes.list - rm newnames.list - rm access.list - echo "Taxonomy inferred for: ${reads} " - rm *headers.list + rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list """ } + } else if (params.headers == "RVDB") { + + process pcASV_AminoAcid_Taxonomy_Inference { + + label 'high_cpus' + + tag "${mtag}" + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{csv,tsv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*dmd.{out}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{fasta}' + + input: + tuple nid, file(asvs) from pcASV_aaDiamond_ch + + output: + file("*.fasta") into ( pcASV_labeledAA ) + tuple file("*phyloseqObject.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_potuaadiamond + tuple nid, file("*_summary_for_plot.csv") into taxplot4 + file("*_quick_Taxbreakdown.csv") into tax_table_pcasvaa + + script: + mtag="ID=" + nid + """ + cp ${params.vampdir}/bin/rename_seq.py . + virdb=${params.dbdir}/${params.dbname} + grep ">" \${virdb} > headers.list + headers="headers.list" + name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') + diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + echo "Preparing lists to generate summary .csv's" + echo "[Best hit accession number]" > access.list + echo "[e-value]" > evalue.list + echo "[Bitscore]" > bit.list + echo "[Percent ID (aa)]" > pid.list + echo "[Organism ID]" > "\$name"_virus.list + echo "[Gene]" > "\$name"_genes.list + echo "[pcASV#]" > otu.list + echo "[Sequence length]" > length.list + grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst + if [[ ${params.lsa} == "T" ]] + then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list + echo "[Classification]" > classfication.list + else echo "skipped" >> \${name}_quick_Taxbreakdown.csv + fi + echo "extracting genes and names" + touch new_"\$name"_asvnames.txt + for s in \$(cat seqids.lst);do + echo "Using RVDB headers." + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + if [[ ${params.lca} == "T" ]] + then group=$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}' + lcla=$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') + echo "\$lcla" >> classfication.list + fi + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list + virus="NO" + gene="HIT" + echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + if [[ ${params.lca} == "T" ]] + then echo "N/A" >> classfication.list + fi + echo "\$s done." + fi + echo "Done with \$s" + done + echo "Now editing "\$name" fasta headers" + ###### rename_seq.py + ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta + echo "[Sequence header]" > newnames.list + cat new_"\$name"_asvnames.txt >> newnames.list + touch sequence.list + echo " " > sequence.list + grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list + rm "\$name"_tmpssasv.fasta + if [[ ${params.lca} == "T" ]] + then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list classfication.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + else paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + fi + if [[ ${params.lca} == "T" ]] + then paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv + fi + for x in *phyloformat.csv;do + echo "\$x" + lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) + tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; + sed 's/ /_/g' tmpcol.list > tmp2col.list; + cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; + rm tmpcol.list tmp2col.list + done + rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list + """ } + } + } if (!params.skipPhylogeny) { diff --git a/vampirus_startup.sh b/vampirus_startup.sh index 5df8e89..02edcf1 100644 --- a/vampirus_startup.sh +++ b/vampirus_startup.sh @@ -197,10 +197,12 @@ then mkdir "$mypwd"/Databases curl -o U-RVDBv21.0-prot.fasta.bz2 https://rvdb-prot.pasteur.fr/files/U-RVDBv21.0-prot.fasta.bz2 bunzip2 U-RVDBv21.0-prot.fasta.bz2 curl -o U-RVDBv21.0-prot-hmm-txt.zip https://rvdb-prot.pasteur.fr/files/U-RVDBv21.0-prot-hmm-txt.zip + unzip U-RVDBv21.0-prot-hmm-txt.zip + mv annot ./RVDBannot echo "Editing confiration file for you now..." - sed 's/DATABASENAME/U-RVDBv19.0-prot.fasta/g' "$mypwd"/vampirus.config > tmp1.config + sed 's/DATABASENAME/U-RVDBv21.0-prot.fasta/g' "$mypwd"/vampirus.config > tmp1.config sed "s|DATABASEDIR|${dir}|g" tmp1.config > tmp2.config - sed "s|DATABASEANNOT|${dir}/U-RVDBv21.0-prot-hmm-txt.zip|g" tmp2.config > tmp3.config + sed "s|DATABASEANNOT|${dir}/RVDBannot|g" tmp2.config > tmp3.config rm tmp1.config rm tmp2.config cat tmp3.config > "$mypwd"/vampirus.config @@ -212,9 +214,17 @@ then mkdir "$mypwd"/Databases dir="$(pwd)" echo "Database installation: Viral RefSeq database version 2.0 (latest as of 2020-07)" curl -o viral.2.protein.faa.gz https://ftp.ncbi.nlm.nih.gov/refseq/release/viral/viral.2.protein.faa.gz + curl -o viral.1.protein.faa.gz https://ftp.ncbi.nlm.nih.gov/refseq/release/viral/viral.1.protein.faa.gz + curl -o viral.3.protein.faa.gz https://ftp.ncbi.nlm.nih.gov/refseq/release/viral/viral.3.protein.faa.gz + gunzip viral.1.protein.faa.gz + cat viral.1.protein.faa >> complete_virus_refseq_prot.fasta gunzip viral.2.protein.faa.gz + cat viral.2.protein.faa >> complete_virus_refseq_prot.fasta + gunzip viral.3.protein.faa.gz + cat viral.3.protein.faa >> complete_virus_refseq_prot.fasta + rm viral.*.protein.faa echo "Editing confiration file for you now..." - sed 's/DATABASENAME/viral.2.protein.faa/g' "$mypwd"/vampirus.config > tmp1.config + sed 's/DATABASENAME/complete_virus_refseq_prot.fasta/g' "$mypwd"/vampirus.config > tmp1.config sed "s|DATABASEDIR|${dir}|g" tmp1.config > tmp2.config rm tmp1.config cat tmp2.config > "$mypwd"/vampirus.config From 167a530eaab6a10ad6c2b938a1a96a42b1f9099c Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Sun, 28 Mar 2021 19:12:52 -0500 Subject: [PATCH 078/223] updated with annotation info download --- vampirus_startup.sh | 4 +++- 1 file changed, 3 insertions(+), 1 deletion(-) diff --git a/vampirus_startup.sh b/vampirus_startup.sh index 02edcf1..447a90c 100644 --- a/vampirus_startup.sh +++ b/vampirus_startup.sh @@ -202,7 +202,7 @@ then mkdir "$mypwd"/Databases echo "Editing confiration file for you now..." sed 's/DATABASENAME/U-RVDBv21.0-prot.fasta/g' "$mypwd"/vampirus.config > tmp1.config sed "s|DATABASEDIR|${dir}|g" tmp1.config > tmp2.config - sed "s|DATABASEANNOT|${dir}/RVDBannot|g" tmp2.config > tmp3.config + sed "s|DATABASEANNOT|${dir}/RVDBannot|g" tmp2.config | sed "s|LCAT|T|g" > tmp3.config rm tmp1.config rm tmp2.config cat tmp3.config > "$mypwd"/vampirus.config @@ -251,6 +251,8 @@ then mkdir "$mypwd"/Databases echo "Database installation: We want 'em all! Might take a little while....'" curl -o NCBI_nr_proteindb.faa.gz https://ftp.ncbi.nlm.nih.gov/blast/db/FASTA/nr.gz curl -o viral.2.protein.faa.gz https://ftp.ncbi.nlm.nih.gov/refseq/release/viral/viral.2.protein.faa.gz + curl -o viral.1.protein.faa.gz https://ftp.ncbi.nlm.nih.gov/refseq/release/viral/viral.1.protein.faa.gz + curl -o viral.3.protein.faa.gz https://ftp.ncbi.nlm.nih.gov/refseq/release/viral/viral.3.protein.faa.gz curl -o U-RVDBv19.0-prot.fasta.bz2 https://rvdb-prot.pasteur.fr/files/U-RVDBv19.0-prot.fasta.bz2 sed "s|DATABASEDIR|${dir}|g" "$mypwd"/vampirus.config > tmp1.config cat tmp1.config > "$mypwd"/vampirus.config From 120cbdb5a27dac1fbe305b64de11a3dc1ad2d2d5 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Sun, 28 Mar 2021 19:22:12 -0500 Subject: [PATCH 079/223] added the removal of uniformative gaps in phylogeny processes --- vAMPirus.nf | 63 ++++++++++++++++++++++++++----------------------- vampirus.config | 8 ++++--- 2 files changed, 38 insertions(+), 33 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index d0956ea..60a0d01 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -2495,30 +2495,31 @@ if (params.DataCheck || params.Analyze) { pre=\$(echo ${prot} | awk -F "_noTax" '{print \$1}' ) mafft --thread ${task.cpus} --maxiterate 15000 --auto ${prot} >\${pre}_ALN.fasta trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html - + o-trim-uninformative-columns-from-alignment \${pre}_aln.fasta + mv \${pre}_aln.fasta-TRIMMED ./\${pre}_Aligned_informativeonly.fasta # Protein_ModelTest - modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d aa -s 203 --disable-checkpoint + modeltest-ng -i \${pre}_Aligned_informativeonly.fasta -p ${task.cpus} -o \${pre}_mt -d aa -s 203 --disable-checkpoint # Protein_Phylogeny if [ "${params.iqCustomaa}" != "" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomaa} + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomaa} elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} + mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni + mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni elif [ "${params.nonparametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} + iqtree -s \${pre}_Aligned_informativeonly.fasta.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} elif [ "${params.parametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + iqtree -s\${pre}_Aligned_informativeonly.fasta.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni else - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + iqtree -s \${pre}_Aligned_informativeonly.fasta.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni fi """ } @@ -3130,30 +3131,31 @@ if (params.DataCheck || params.Analyze) { pre=\$( echo ${reads} | awk -F "_noTax" '{print \$1}' ) mafft --maxiterate 5000 --auto ${reads} >\${pre}_ALN.fasta trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html - + o-trim-uninformative-columns-from-alignment \${pre}_aln.fasta + mv \${pre}_aln.fasta-TRIMMED ./\${pre}_Aligned_informativeonly.fasta # pcASV_Nucleotide_ModelTest - modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_noTaxonomy_mt -d nt -s 203 --disable-checkpoint + modeltest-ng -i \${pre}_Aligned_informativeonly.fasta -p ${task.cpus} -o \${pre}_noTaxonomy_mt -d nt -s 203 --disable-checkpoint # pcASV_Nucleotide_Phylogeny if [ "${params.iqCustomnt}" != "" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_noTaxonomy_iq --redo -T auto ${params.iqCustomnt} + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_noTaxonomy_iq --redo -T auto ${params.iqCustomnt} elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_noTaxonomy_iq -m \${mod} --redo-nt auto -b ${params.boots} + mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_noTaxonomy_iq -m \${mod} --redo-nt auto -b ${params.boots} elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_noTaxonomy_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni + mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_noTaxonomy_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni elif [ "${params.nonparametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_noTaxonomy_iq -m MFP --redo -nt auto -b ${params.boots} + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_noTaxonomy_iq -m MFP --redo -nt auto -b ${params.boots} elif [ "${params.parametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_noTaxonomy_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_noTaxonomy_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni else - iqtree -s \${pre}_aln.fasta --prefix \${pre}_noTaxonomy_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_noTaxonomy_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni fi """ } @@ -3491,30 +3493,31 @@ if (params.DataCheck || params.Analyze) { pre=\$( echo ${prot} | awk -F ".fasta" '{print \$1}' ) mafft --maxiterate 5000 --auto ${prot} >\${pre}_ALN.fasta trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html - + o-trim-uninformative-columns-from-alignment \${pre}_aln.fasta + mv \${pre}_aln.fasta-TRIMMED ./\${pre}_Aligned_informativeonly.fasta # pcASV_Protein_ModelTest - modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d aa -s 203 --disable-checkpoint + modeltest-ng -i \${pre}_Aligned_informativeonly.fasta -p ${task.cpus} -o \${pre}_mt -d aa -s 203 --disable-checkpoint # pcASV_Protein_Phylogeny if [ "${params.iqCustomaa}" != "" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomaa} + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomaa} elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} + mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni + mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni elif [ "${params.nonparametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} elif [ "${params.parametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni else - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni fi """ } diff --git a/vampirus.config b/vampirus.config index aef6beb..3fdc778 100644 --- a/vampirus.config +++ b/vampirus.config @@ -93,10 +93,12 @@ params { dbname="DATABASENAME" // Path to Directory where database is being stored dbdir="DATABASEDIR" - // Path to hmm annotation .txt file - see manual for information on this. Leave as is if not planning on using. + // Set taxonomy database header format -> headers= "RefSeq" to toggle use of RefSeq header format; set to "RVDB" to signal the use of Reverence Viral DataBase (RVDB) headers + headers="HEAD" + // Path to directory hmm annotation .txt file - see manual for information on this. Leave as is if not planning on using. dbanno="DATABASEANNOT" - // Toggle use of RefSeq header format; default is Reverence Viral DataBase (RVDB) - refseq="F" + // Set lca="T" if you would like to add taxonomic classification to taxonomy results - example: "ASV1, Viruses::Duplodnaviria::Heunggongvirae::Peploviricota::Herviviricetes::Herpesvirales::Herpesviridae::Gammaherpesvirinae::Macavirus" + lca="LCAT" // Set minimum bitscore for best hit in taxonomy assignment bitscore="50" // Set minimum percent amino acid similarity for best hit to be counted in taxonomy assignment From 4699ae8b02aee5017a573e186de3012800267134 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Sun, 28 Mar 2021 19:45:59 -0500 Subject: [PATCH 080/223] updated options in help --- vAMPirus.nf | 225 ++++++++++++++++++++++++++------------------ vampirus.config | 4 +- vampirus_startup.sh | 14 +-- 3 files changed, 145 insertions(+), 98 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 60a0d01..cde16b1 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -57,6 +57,12 @@ def helpMessage() { --pcASV Set this option to have vAMPirus cluster nucleotide and translated ASVs into protein-based operational taxonomic units (pcASVs) - See options below to define a single percent similarity or a list + --Minimum Entropy Decomposition arguments-- + + --asvMED Set this option to perform Minimum Entropy Decomposition on ASV sequences, see manual for more information. You will need to set a value for --asvC to perform this analysis + + --amino_med Set this option to perform Minimum Entropy Decomposition on AminoType sequences, see manual for more information. You will need to set a value for --aminoC to perform this analysis + --Skip options-- --skipReadProcessing Set this option to skip all read processing steps in the pipeline @@ -65,7 +71,9 @@ def helpMessage() { --skipAdapterRemoval Set this option to skip adapter removal in the pipeline - --skipPrimerRemoval Set this option to skup Skip primer removal process + --skipPrimerRemoval Set this option to skip primer removal process + + --skipMerging Set this option to skip read merging --skipAminoTyping Set this option to skip AminoTyping processes @@ -73,6 +81,10 @@ def helpMessage() { --skipPhylogeny Set this option to skip phylogeny processes + --skipEMBOSS Set this option to skip EMBOSS processes + + --skipReport Set this option to skip html report generation + **NOTE** Most opitons below can be set using the configuration file (vampirus.config) to avoid a lengthy launch command. --Project/analysis information-- @@ -140,6 +152,11 @@ def helpMessage() { --minAA With --pcASV set, use this option to set the expected or minimum amino acid sequence length of open reading frames within your amplicon sequences + --Minimum Entropy Decomposition-- + + --asvC Number of high entropy positions to use for ASV MED analysis and generate "Groups" + + --aminoC Number of high entropy positions to use for AminoType MED analysis and generate "Groups" --Counts table generation-- @@ -158,7 +175,11 @@ def helpMessage() { --dbdir Path to Directory where database is being stored - --refseq Set "--refseq T" to toggle use of RefSeq header format; default is "F" to use Reverence Viral DataBase (RVDB) header + --headers Set taxonomy database header format -> headers= "RefSeq" to toggle use of RefSeq header format; set to "RVDB" to signal the use of Reverence Viral DataBase (RVDB) headers + + --dbanno Path to directory hmm annotation .txt file - see manual for information on this. Leave as is if not planning on using. + + --lca Set --lca T if you would like to add taxonomic classification to taxonomy results - example: "ASV1, Viruses::Duplodnaviria::Heunggongvirae::Peploviricota::Herviviricetes::Herpesvirales::Herpesviridae::Gammaherpesvirinae::Macavirus" --bitscore Set minimum bitscore to allow for best hit in taxonomy assignment @@ -205,186 +226,208 @@ def fullHelpMessage() { THIS IS A LONGER HELP WITH USAGE EXAMPLES vAMPirus v${workflow.manifest.version} ============================================================================================================================================================================================== - Steps: - 1- Before launching the vAMPirus.nf, be sure to run the vampirus_startup.sh script to install dependencies and/or databases + Steps: + 1- Before launching the vAMPirus.nf, be sure to run the vampirus_startup.sh script to install dependencies and/or databases + + 2- Test the vAMPirus installation with the provided test dataset (if you have ran the start up script, you can see STARTUP_HELP.txt for test commands and other examples) + + 3. Edit parameters in vampirus.config file - 2- Test the vAMPirus installation with the provided test dataset (if you have ran the start up script, you can see STARTUP_HELP.txt for test commands and other examples) + 4. Launch the DataCheck pipeline to get summary information about your dataset - 3. Edit parameters in vampirus.config file + 5. Change any parameters in vampirus.config file that might aid your analysis (e.g. clustering ID, maximum merged read length) - 4. Launch the DataCheck pipeline to get summary information about your dataset + 6. Launch the Analyze pipeline to perform a comprehensive analysis with your dataset - 5. Change any parameters in vampirus.config file that might aid your analysis (e.g. clustering ID, maximum merged read length) + 7. Explore results directories and produced final reports - 6. Launch the Analyze pipeline to perform a comprehensive analysis with your dataset - 7. Explore results directories and produced final reports + Usage: + nextflow run vAMPirus.nf -c vampirus.config -profile [conda|singularity] --[Analyze|DataCheck] [--ncASV] [--pcASV] - Usage: - nextflow run vAMPirus.nf -c vampirus.config -profile [conda|singularity] --[Analyze|DataCheck] [--ncASV] [--pcASV] + --Help options-- + --help Print help information - --Help options-- + --fullHelp Print even more help information - --help Print help information - --fullHelp Print even more help information + --Mandatory arguments (choose one)-- + --Analyze Run absolutely everything - --Mandatory arguments (choose one)-- + --DataCheck Assess how data performs with during processing and clustering - --Analyze Run absolutely everything - --DataCheck Assess how data performs with during processing and clustering + --ASV clustering arguments-- + --ncASV Set this option to have vAMPirus cluster nucleotide amplicon sequence variants (ASVs) into nucleotide-based operational taxonomic units (ncASVs) - See options below to define a single percent similarity or a list - --ASV clustering arguments-- + --pcASV Set this option to have vAMPirus cluster nucleotide and translated ASVs into protein-based operational taxonomic units (pcASVs) - See options below to define a single percent similarity or a list - --ncASV Set this option to have vAMPirus cluster nucleotide amplicon sequence variants (ASVs) into nucleotide-based operational taxonomic units (ncASVs) - See options below to define a single percent similarity or a list - --pcASV Set this option to have vAMPirus cluster nucleotide and translated ASVs into protein-based operational taxonomic units (pcASVs) - See options below to define a single percent similarity or a list + --Minimum Entropy Decomposition arguments-- + --asvMED Set this option to perform Minimum Entropy Decomposition on ASV sequences, see manual for more information. You will need to set a value for --asvC to perform this analysis - --Skip options-- + --amino_med Set this option to perform Minimum Entropy Decomposition on AminoType sequences, see manual for more information. You will need to set a value for --aminoC to perform this analysis - --skipReadProcessing Set this option to skip all read processing steps in the pipeline + --Skip options-- - --skipFastQC Set this option to skiip FastQC steps in the pipeline + --skipReadProcessing Set this option to skip all read processing steps in the pipeline - --skipAdapterRemoval Set this option to skip adapter removal in the pipeline + --skipFastQC Set this option to skiip FastQC steps in the pipeline - --skipPrimerRemoval Set this option to skup Skip primer removal process + --skipAdapterRemoval Set this option to skip adapter removal in the pipeline - --skipAminoTyping Set this option to skip AminoTyping processes + --skipPrimerRemoval Set this option to skip primer removal process - --skipTaxonomy Set this option to skip taxonomy assignment processes + --skipMerging Set this option to skip read merging - --skipPhylogeny Set this option to skip phylogeny processes + --skipAminoTyping Set this option to skip AminoTyping processes - **NOTE** Most opitons below can be set using the configuration file (vampirus.config) to avoid a lengthy launch command. + --skipTaxonomy Set this option to skip taxonomy assignment processes - --Project/analysis information-- + --skipPhylogeny Set this option to skip phylogeny processes - --projtag Set project name to be used as a prefix for output files + --skipEMBOSS Set this option to skip EMBOSS processes - --metadata Set path to metadata spreadsheet file to be used for report generation (must be defined if generating report) + --skipReport Set this option to skip html report generation - --reads Path to directory containing read libraries, must have *R{1,2}* in the library names - --workingdir Path to working directory where Nextflow will put all Nextflow and vAMPirus generated output files + **NOTE** Most opitons below can be set using the configuration file (vampirus.config) to avoid a lengthy launch command. - --outdir Name of results directory containing all output from the chosen pipeline (will be made within the working directory) + --Project/analysis information-- + --projtag Set project name to be used as a prefix for output files - --Merged read length filtering-- + --metadata Set path to metadata spreadsheet file to be used for report generation (must be defined if generating report) - --minLen Minimum merged read length - reads below the specified maximum read length will be used for counts only + --reads Path to directory containing read libraries, must have *R{1,2}* in the library names - --maxLen Maximum merged read length - reads with length equal to the specified max read length will be used to identifying unique sequences and subsequent Amplicon Sequence Variant (ASV) analysis + --workingdir Path to working directory where Nextflow will put all Nextflow and vAMPirus generated output files - --maxEE Use this option to set the maximum expected error rate for vsearch merging. Default is 1. + --outdir Name of results directory containing all output from the chosen pipeline (will be made within the working directory) - --Primer removal-- + --Merged read length filtering-- - General primer removal parameters + --minLen Minimum merged read length - reads below the specified maximum read length will be used for counts only - --primerLength Use this option to set the max primer length to restrict bbduk.sh primer trimming to the first x number of bases + --maxLen Maximum merged read length - reads with length equal to the specified max read length will be used to identifying unique sequences and subsequent Amplicon Sequence Variant (ASV) analysis - --maxkmer Maximum kmer length for bbduk.sh to use for primer detection and removal (must be shorter than your primer length; default = 13) + --maxEE Use this option to set the maximum expected error rate for vsearch merging. Default is 1. - --minkmer Minimum kmer length for primer removal (default = 3) - --minilen Minimum read length after adapter and primer removal (default = 200) + --Primer removal-- - Single primer set removal- + General primer removal parameters - --GlobTrim Set this option to perform global trimming to reads to remove primer sequences. Example usage "--GlobTrim #basesfromforward,#basesfromreverse" + --primerLength Use this option to set the max primer length to restrict bbduk.sh primer trimming to the first x number of bases - --fwd Forward primer sequence for reads to be detected and removed from reads (must specify reverse sequence if providing forward) + --maxkmer Maximum kmer length for bbduk.sh to use for primer detection and removal (must be shorter than your primer length; default = 13) - --rev Reverse primer sequence for reads to be detected and removed from reads (must specify forward sequence if providing reverse) + --minkmer Minimum kmer length for primer removal (default = 3) - Multiple primer set removal- + --minilen Minimum read length after adapter and primer removal (default = 200) - --multi Use this option to signal multiple primer sequence removal within the specified pipeline + Single primer set removal- - --primers Use this option to set the path to a fasta file with all of the primer sequences to be detected and removed from reads + --GlobTrim Set this option to perform global trimming to reads to remove primer sequences. Example usage "--GlobTrim #basesfromforward,#basesfromreverse" + --fwd Forward primer sequence for reads to be detected and removed from reads (must specify reverse sequence if providing forward) - --Amplicon Sequence Variant (ASV) genration and clustering-- + --rev Reverse primer sequence for reads to be detected and removed from reads (must specify forward sequence if providing reverse) - --alpha Alpha value for denoising - the higher the alpha the higher the chance of false positives in ASV generation (1 or 2) + Multiple primer set removal- - --minSize Minimum size or representation for sequence to be considered in ASV generation + --multi Use this option to signal multiple primer sequence removal within the specified pipeline - --clusterNuclID With --ncASV set, use this option to set a single percent similarity to cluster nucleotide ASV sequences into ncASVs by [ Example: --clusterNuclID .97 ] + --primers Use this option to set the path to a fasta file with all of the primer sequences to be detected and removed from reads - --clusterNuclIDlist With --ncASV set, use this option to perform nucleotide-based clustering of ASVs with a comma separated list of percent similarities [ Example: --clusterNuclIDlist .95,.96,.97,.98 ] - --clusterAAID With --pcASV set, use this option to set a single percent similarity for protein-based ASV clustering to generate pcASVs[ Example: --clusterAAID .97 ] + --Amplicon Sequence Variant (ASV) genration and clustering-- - --clusterAAIDlist With --pcASV set, use this option to perform protein-based ASV clustering to generate pcASVs with a comma separated list of percent similarities [ Example: --clusterAAIDlist .95,.96,.97,.98 ] + --alpha Alpha value for denoising - the higher the alpha the higher the chance of false positives in ASV generation (1 or 2) - --minAA With --pcASV set, use this option to set the expected or minimum amino acid sequence length of open reading frames within your amplicon sequences + --minSize Minimum size or representation for sequence to be considered in ASV generation + --clusterNuclID With --ncASV set, use this option to set a single percent similarity to cluster nucleotide ASV sequences into ncASVs by [ Example: --clusterNuclID .97 ] - --Counts table generation-- + --clusterNuclIDlist With --ncASV set, use this option to perform nucleotide clustering with a comma separated list of percent similarities [ Example: --clusterNuclIDlist .95,.96,.97,.98 ] - --asvcountID Similarity ID to use for ASV counts + --clusterAAID With --pcASV set, use this option to set a single percent similarity for protein-based ASV clustering to generation pcASVs [ Example: --clusterAAID .97 ] - --ProtCountID Minimum amino acid sequence similarity for hit to count + --clusterAAIDlist With --pcASV set, use this option to perform protein-based ASV clustering to generate pcASVs with a comma separated list of percent similarities [ Example: --clusterAAIDlist .95,.96,.97,.98 ] - --ProtCountsLength Minimum alignment length for hit to count + --minAA With --pcASV set, use this option to set the expected or minimum amino acid sequence length of open reading frames within your amplicon sequences - --ProtCountsBit Minimum bitscore for hit to be counted + --Minimum Entropy Decomposition-- + --asvC Number of high entropy positions to use for ASV MED analysis and generate "Groups" - --Taxonomy inference parameters-- + --aminoC Number of high entropy positions to use for AminoType MED analysis and generate "Groups" - --dbname Specify name of database to use for analysis + --Counts table generation-- - --dbdir Path to Directory where database is being stored + --asvcountID Similarity ID to use for ASV counts - --refseq Set "--refseq T" to toggle use of RefSeq header format; default is "F" to use Reverence Viral DataBase (RVDB) header + --ProtCountID Minimum amino acid sequence similarity for hit to count - --bitscore Set minimum bitscore to allow for best hit in taxonomy assignment + --ProtCountsLength Minimum alignment length for hit to count - --minID Set minimum percent amino acid similarity for best hit to be counted in taxonomy assignment + --ProtCountsBit Minimum bitscore for hit to be counted - --minaln Set minimum amino acid alignment length for best hit to be counted in taxonomy assignment + --Taxonomy inference parameters-- - --Phylogeny analysis parameters-- + --dbname Specify name of database to use for analysis - Setting customs options for IQ-TREE (Example: "-option1 A -option2 B -option3 C -option4 D") - might be easier to set in the vampirus.config file at lines 108/109 + --dbdir Path to Directory where database is being stored - --iqCustomnt Use option to set custom options to use in all IQTREE analyses with nuceoltide sequences + --headers Set taxonomy database header format -> headers= "RefSeq" to toggle use of RefSeq header format; set to "RVDB" to signal the use of Reverence Viral DataBase (RVDB) headers - --iqCustomaa Use option to set custom options to use in all IQTREE analyses with amino acid sequences + --dbanno Path to directory hmm annotation .txt file - see manual for information on this. Leave as is if not planning on using. - These options below you can set at the command, for example, to set to use model from ModelTest-NG with parametric bootstrapping --ModelTnt --ModelTaa --parametric + --lca Set --lca T if you would like to add taxonomic classification to taxonomy results - example: "ASV1, Viruses::Duplodnaviria::Heunggongvirae::Peploviricota::Herviviricetes::Herpesvirales::Herpesviridae::Gammaherpesvirinae::Macavirus" - --ModelTnt=false Signal for IQ-TREE to use model determined by ModelTest-NG for all IQTREE analyses with nuceoltide sequences (Default is IQ-TREE will do automatic model testing with ModelFinder Plus) + --bitscore Set minimum bitscore to allow for best hit in taxonomy assignment - --ModelTaa=false Signal for IQ-TREE to use model determined by ModelTest-NG for all IQTREE analyses with amino acid sequences + --minID Set minimum percent amino acid similarity for best hit to be counted in taxonomy assignment - --parametric Set to use parametric bootstrapping in IQTREE analyses + --minaln Set minimum amino acid alignment length for best hit to be counted in taxonomy assignment - --nonparametric Set to use parametric bootstrapping in IQTREE analyses - --boots Number of bootstraps (recommended 1000 for parametric and 100 for non-parametric) + --Phylogeny analysis parameters-- + Setting customs options for IQ-TREE (Example: "-option1 A -option2 B -option3 C -option4 D") - might be easier to set in the vampirus.config file at lines 108/109 - --Statistics options-- + --iqCustomnt Use option to set custom options to use in all IQTREE analyses with nuceoltide sequences - --stats Set "--stats run" to signal statstical tests to be performed and included in the final report + --iqCustomaa Use option to set custom options to use in all IQTREE analyses with amino acid sequences - --minimumCounts Minimum number of hit counts for a sample to have to be included in the downstream statistical analyses and report generation + These options below you can set at the command, for example, to set to use model from ModelTest-NG with parametric bootstrapping --ModelTnt --ModelTaa --parametric - --trymax Maximum number of iterations performed by metaMDS + --ModelTnt=false Signal for IQ-TREE to use model determined by ModelTest-NG for all IQTREE analyses with nuceoltide sequences (Default is IQ-TREE will do automatic model testing with ModelFinder Plus) + + --ModelTaa=false Signal for IQ-TREE to use model determined by ModelTest-NG for all IQTREE analyses with amino acid sequences + + --parametric Set to use parametric bootstrapping in IQTREE analyses + + --nonparametric Set to use parametric bootstrapping in IQTREE analyses + + --boots Number of bootstraps (recommended 1000 for parametric and 100 for non-parametric) + + + --Statistics options-- + + --stats Set "--stats run" to signal statstical tests to be performed and included in the final report + + --minimumCounts Minimum number of hit counts for a sample to have to be included in the downstream statistical analyses and report generation + + --trymax Maximum number of iterations performed by metaMDS ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||| @@ -414,7 +457,7 @@ def fullHelpMessage() { Example 4. Launching the vAMPirus Analyze pipeline with singularity with ASV and AminoType generation with all accesory analyses (taxonomy assignment, EMBOSS, IQTREE, statistics) - nextflow run vAMPirus.nf -c vampirus.config -profile singularity --Analyze --stats run + nextflow run vAMPirus.nf -c vampirus.config -profile singularity --Analyze --stats Example 5. Launching the vAMPirus Analyze pipeline with conda to perform multiple primer removal and protein-based clustering of ASVs, but skip most of the extra analyses @@ -422,7 +465,11 @@ def fullHelpMessage() { Example 6. Launching vAMPirus Analyze pipeline with conda to produce only ASV-related results - nextflow run vAMPirus.nf -c vampirus.config -profile conda --Analyze --skipAminoTyping --stats run + nextflow run vAMPirus.nf -c vampirus.config -profile conda --Analyze --skipAminoTyping --stats + + Example 7. Launching vAMPirus Analyze pipeline with conda to perform ASV analyses with Minimum Entropy Decomposition to form "Groups" + + nextflow run vAMPirus.nf -c vampirus.config -profile conda --Analyze --skipAminoTyping --stats --asvMED --asvC 24 Resuming analyses => diff --git a/vampirus.config b/vampirus.config index 3fdc778..05db904 100644 --- a/vampirus.config +++ b/vampirus.config @@ -125,8 +125,8 @@ params { boots="1000" // Stats options - // Tell vAMPirus to perform statistical analyses by setting "stats="run"" below or in the launch command by adding "--stats run" to it - stats=false + // Tell vAMPirus to perform statistical analyses by setting "stats = true" below or in the launch command by adding "--stats" to it + stats = false // Minimum number of hit counts for a sample to have to be included in the downstream statistical analyses and report generation minimumCounts="1000" // Maximum number of iteration performed by metaMDS diff --git a/vampirus_startup.sh b/vampirus_startup.sh index 447a90c..7b1f980 100644 --- a/vampirus_startup.sh +++ b/vampirus_startup.sh @@ -202,7 +202,7 @@ then mkdir "$mypwd"/Databases echo "Editing confiration file for you now..." sed 's/DATABASENAME/U-RVDBv21.0-prot.fasta/g' "$mypwd"/vampirus.config > tmp1.config sed "s|DATABASEDIR|${dir}|g" tmp1.config > tmp2.config - sed "s|DATABASEANNOT|${dir}/RVDBannot|g" tmp2.config | sed "s|LCAT|T|g" > tmp3.config + sed "s|DATABASEANNOT|${dir}/RVDBannot|g" tmp2.config | sed "s|LCAT|T|g" | sed 's/HEAD/RVDB/g' > tmp3.config rm tmp1.config rm tmp2.config cat tmp3.config > "$mypwd"/vampirus.config @@ -225,7 +225,7 @@ then mkdir "$mypwd"/Databases rm viral.*.protein.faa echo "Editing confiration file for you now..." sed 's/DATABASENAME/complete_virus_refseq_prot.fasta/g' "$mypwd"/vampirus.config > tmp1.config - sed "s|DATABASEDIR|${dir}|g" tmp1.config > tmp2.config + sed "s|DATABASEDIR|${dir}|g" tmp1.config | sed 's/HEAD/RefSeq/g' > tmp2.config rm tmp1.config cat tmp2.config > "$mypwd"/vampirus.config rm tmp2.config @@ -239,7 +239,7 @@ then mkdir "$mypwd"/Databases gunzip NCBI_nr_proteindb.faa.gz echo "Editing confiration file for you now..." sed 's/DATABASENAME/NCBI_nr_proteindb.faa/g' "$mypwd"/vampirus.config > tmp1.config - sed "s|DATABASEDIR|${dir}|g" tmp1.config > tmp2.config + sed "s|DATABASEDIR|${dir}|g" tmp1.config | sed 's/HEAD/RefSeq/g' > tmp2.config rm tmp1.config cat tmp2.config > "$mypwd"/vampirus.config rm tmp2.config @@ -284,11 +284,11 @@ then echo ""$mypwd"/nextflow run "$mypwd"/vAMPirus.nf -c "$mypwd"/vampirus.config -profile singularity,test --DataCheck" >> STARTUP_HELP.txt echo " " >> STARTUP_HELP.txt echo "Next, test the analysis pipeline:" >> STARTUP_HELP.txt - echo ""$mypwd"/nextflow run "$mypwd"/vAMPirus.nf -c "$mypwd"/vampirus.config -profile conda,test --Analyze --ncASV --pcASV --stats run" >> STARTUP_HELP.txt + echo ""$mypwd"/nextflow run "$mypwd"/vAMPirus.nf -c "$mypwd"/vampirus.config -profile conda,test --Analyze --ncASV --pcASV --stats" >> STARTUP_HELP.txt echo " " >> STARTUP_HELP.txt echo "Or if you plan to run vAMPirus using Singularity, use this test command:" >> STARTUP_HELP.txt echo " " >> STARTUP_HELP.txt - echo ""$mypwd"/nextflow run "$mypwd"/vAMPirus.nf -c "$mypwd"/vampirus.config -profile singularity,test --Analyze --ncASV --pcASV --stats run" >> STARTUP_HELP.txt + echo ""$mypwd"/nextflow run "$mypwd"/vAMPirus.nf -c "$mypwd"/vampirus.config -profile singularity,test --Analyze --ncASV --pcASV --stats" >> STARTUP_HELP.txt echo "--------------------------------------------------------------------------------------------------------------------------------" >> STARTUP_HELP.txt echo " " >> STARTUP_HELP.txt echo "Ok, if everything went well (green text was spit out by Nextflow), now you can move on to the fun. First, you should review the help docs and the vampirus.config in the vAMPirus directory." >> STARTUP_HELP.txt @@ -308,11 +308,11 @@ then echo "-------------------------------------------------------------------------------------------------------------------------------- RUNNING Analyze PIPELINE WITH YOUR DATA" >> STARTUP_HELP.txt echo "Then you can run the analysis using the -with-conda Nextflow option, here is a launch command to run the complete analysis and statistical tests:" >> STARTUP_HELP.txt echo " " >> STARTUP_HELP.txt - echo ""$mypwd"/nextflow run "$mypwd"/vAMPirus.nf -c "$mypwd"/vampirus.config -with-conda "$environment" --Analyze --ncASV --pcASV --stats run" >> STARTUP_HELP.txt + echo ""$mypwd"/nextflow run "$mypwd"/vAMPirus.nf -c "$mypwd"/vampirus.config -with-conda "$environment" --Analyze --ncASV --pcASV --stats" >> STARTUP_HELP.txt echo " " >> STARTUP_HELP.txt echo "OR same command using -profile option of Nextflow ..." >> STARTUP_HELP.txt echo " " >> STARTUP_HELP.txt - echo ""$mypwd"/nextflow run "$mypwd"/vAMPirus.nf -c "$mypwd"/vampirus.config -profile [conda|singularity] --Analyze --ncASV --pcASV --stats run" >> STARTUP_HELP.txt + echo ""$mypwd"/nextflow run "$mypwd"/vAMPirus.nf -c "$mypwd"/vampirus.config -profile [conda|singularity] --Analyze --ncASV --pcASV --stats" >> STARTUP_HELP.txt echo " " >> STARTUP_HELP.txt echo "--------------------------------------------------------------------------------------------------------------------------------" >> STARTUP_HELP.txt fi From 463553cbe53e1fbd390dd6bc5bf2651fd575c38c Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Mon, 29 Mar 2021 14:19:10 -0700 Subject: [PATCH 081/223] Fix typo --- bin/vAMPirus_Report.Rmd | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/bin/vAMPirus_Report.Rmd b/bin/vAMPirus_Report.Rmd index 300f310..af10a49 100644 --- a/bin/vAMPirus_Report.Rmd +++ b/bin/vAMPirus_Report.Rmd @@ -225,7 +225,7 @@ raredata <- as.data.frame(rrarefy(data5[,2:cols], sample=minreads))

-### Shannon diversty +### Shannon diversity
```{r diversity_analysis, echo=FALSE} @@ -307,7 +307,7 @@ if (params$stats == "true" ) {

-### Simpson diversty +### Simpson diversity
```{r diversity_analysis2, echo=FALSE} From 9034ec7b016dd1c46009b39eaef9edcf0907f4f5 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Tue, 30 Mar 2021 13:41:18 -0500 Subject: [PATCH 082/223] update --- vAMPirus.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index cde16b1..f5e3aca 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -33,7 +33,7 @@ def helpMessage() { Usage: - nextflow run vAMPirus.nf -c vampirus.config -profile [conda|singularity] --[Analyze|DataCheck] [--ncASV] [--pcASV] + nextflow run vAMPirus.nf -c vampirus.config -profile [conda|singularity] --[Analyze|DataCheck] [--ncASV] [--pcASV] [--asvMED] [--aminoMED] [--stats] --Help options-- From 5643f065b9ac9d5a7425563b1673ad3af33cb973 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Tue, 30 Mar 2021 11:52:45 -0700 Subject: [PATCH 083/223] Add pip install command --- vampirus_env.yml | 2 ++ 1 file changed, 2 insertions(+) diff --git a/vampirus_env.yml b/vampirus_env.yml index df49583..7ecc2be 100644 --- a/vampirus_env.yml +++ b/vampirus_env.yml @@ -37,3 +37,5 @@ dependencies: - bioconductor-biocparallel=1.24.0 - pigz=2.4 - r-biocmanager=1.30.10 + - pip: + - oligotyping From 41c29e60855d498e7f85a585da8ea3550ae1e63b Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Tue, 30 Mar 2021 11:53:07 -0700 Subject: [PATCH 084/223] Remove pip install code --- vampirus_startup.sh | 5 ----- 1 file changed, 5 deletions(-) diff --git a/vampirus_startup.sh b/vampirus_startup.sh index 7b1f980..0c3f1f3 100644 --- a/vampirus_startup.sh +++ b/vampirus_startup.sh @@ -177,11 +177,6 @@ else rm tmp1.config fi -echo "Downloading oligotyping program" -conda init && conda activate vAMPirus -pip install oligotyping -conda deactivate - echo "-------------------------------------------------------------------------------- Conda check/install done" echo "Now lets check the status of Nextflow on your system..." From d16c3341dba9d470bf61e4b21f419a75374d5c5e Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Tue, 30 Mar 2021 12:06:01 -0700 Subject: [PATCH 085/223] Remove extra code --- vAMPirus.nf | 20 +++++--------------- 1 file changed, 5 insertions(+), 15 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index f5e3aca..0487021 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1547,12 +1547,10 @@ if (params.DataCheck || params.Analyze) { if [[ ${params.lca} == "T" ]] then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list classfication.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv else paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv fi - if [[ ${params.lca} == "T" ]] - then paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv - fi for x in *phyloformat.csv;do echo "\$x" lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) @@ -1877,12 +1875,10 @@ if (params.DataCheck || params.Analyze) { if [[ ${params.lca} == "T" ]] then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list classfication.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv else paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv fi - if [[ ${params.lca} == "T" ]] - then paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv - fi for x in *phyloformat.csv;do echo "\$x" lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) @@ -2499,12 +2495,10 @@ if (params.DataCheck || params.Analyze) { if [[ ${params.lca} == "T" ]] then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list classfication.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv else paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv fi - if [[ ${params.lca} == "T" ]] - then paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv - fi for x in *phyloformat.csv;do echo "\$x" lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) @@ -3078,12 +3072,10 @@ if (params.DataCheck || params.Analyze) { if [[ ${params.lca} == "T" ]] then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list classfication.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv else paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv fi - if [[ ${params.lca} == "T" ]] - then paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv - fi for x in *phyloformat.csv;do echo "\$x" lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) @@ -3495,12 +3487,10 @@ if (params.DataCheck || params.Analyze) { if [[ ${params.lca} == "T" ]] then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list classfication.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv else paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv fi - if [[ ${params.lca} == "T" ]] - then paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv - fi for x in *phyloformat.csv;do echo "\$x" lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) From 336159e4cac45af0ffd8dfa031e3023bfc10a01a Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Tue, 30 Mar 2021 12:09:13 -0700 Subject: [PATCH 086/223] Add missing "\" --- vAMPirus.nf | 20 ++++++++++---------- 1 file changed, 10 insertions(+), 10 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 0487021..228b5c3 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1508,8 +1508,8 @@ if (params.DataCheck || params.Analyze) { echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt if [[ ${params.lca} == "T" ]] - then group=$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}' - lcla=$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') + then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}' + lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') echo "\$lcla" >> classfication.list fi echo "\$s done." @@ -1836,8 +1836,8 @@ if (params.DataCheck || params.Analyze) { echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt if [[ ${params.lca} == "T" ]] - then group=$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}' - lcla=$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') + then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}' + lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') echo "\$lcla" >> classfication.list fi echo "\$s done." @@ -2456,8 +2456,8 @@ if (params.DataCheck || params.Analyze) { echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt if [[ ${params.lca} == "T" ]] - then group=$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}' - lcla=$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') + then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}' + lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') echo "\$lcla" >> classfication.list fi echo "\$s done." @@ -3033,8 +3033,8 @@ if (params.DataCheck || params.Analyze) { echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt if [[ ${params.lca} == "T" ]] - then group=$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}' - lcla=$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') + then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}' + lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') echo "\$lcla" >> classfication.list fi echo "\$s done." @@ -3448,8 +3448,8 @@ if (params.DataCheck || params.Analyze) { echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt if [[ ${params.lca} == "T" ]] - then group=$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}' - lcla=$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') + then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}' + lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') echo "\$lcla" >> classfication.list fi echo "\$s done." From 596f4d32de8c61b0f53209eca8f410b80c39ff4c Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Tue, 30 Mar 2021 12:13:19 -0700 Subject: [PATCH 087/223] Add process names --- vAMPirus.nf | 20 ++++++++++---------- 1 file changed, 10 insertions(+), 10 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 228b5c3..4434f5c 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1344,7 +1344,7 @@ if (params.DataCheck || params.Analyze) { if (params.headers == "RefSeq") { - process ncASV_Taxonomy_Inference { /////// editttt + process ncASV_Taxonomy_Inference_RefSeq { /////// editttt label 'high_cpus' @@ -1444,7 +1444,7 @@ if (params.DataCheck || params.Analyze) { } } else if (params.headers == "RVDB") { - process ncASV_Taxonomy_Inference { /////// editttt + process ncASV_Taxonomy_Inference_RVDB { /////// editttt label 'high_cpus' @@ -1678,7 +1678,7 @@ if (params.DataCheck || params.Analyze) { if (params.headers == "RefSeq") { - process ASV_Taxonomy_Inference { /////// editttt + process ASV_Taxonomy_Inference_RefSeq { /////// editttt label 'high_cpus' @@ -1775,7 +1775,7 @@ if (params.DataCheck || params.Analyze) { } } else if (params.headers == "RVDB") { - process ASV_Taxonomy_Inference { /////// editttt + process ASV_Taxonomy_Inference_RVDB { /////// editttt label 'high_cpus' @@ -2298,7 +2298,7 @@ if (params.DataCheck || params.Analyze) { if (params.headers == "RefSeq") { - process AminoType_Taxonomy_Inference { + process AminoType_Taxonomy_Inference_RefSeq { label 'high_cpus' @@ -2395,7 +2395,7 @@ if (params.DataCheck || params.Analyze) { } } else if (params.headers == "RVDB") { - process AminoType_Taxonomy_Inference { + process AminoType_Taxonomy_Inference_RVDB { label 'high_cpus' @@ -2865,7 +2865,7 @@ if (params.DataCheck || params.Analyze) { if (params.headers == "RefSeq") { - process pcASV_Nucleotide_Taxonomy_Inference { + process pcASV_Nucleotide_Taxonomy_Inference_RefSeq { label 'high_cpus' @@ -2967,7 +2967,7 @@ if (params.DataCheck || params.Analyze) { } } else if (params.headers == "RVDB") { - process pcASV_Nucleotide_Taxonomy_Inference { + process pcASV_Nucleotide_Taxonomy_Inference_RVDB { label 'high_cpus' @@ -3282,7 +3282,7 @@ if (params.DataCheck || params.Analyze) { if (params.headers == "RefSeq") { - process pcASV_AminoAcid_Taxonomy_Inference { + process pcASV_AminoAcid_Taxonomy_Inference_RefSeq { label 'high_cpus' @@ -3383,7 +3383,7 @@ if (params.DataCheck || params.Analyze) { } } else if (params.headers == "RVDB") { - process pcASV_AminoAcid_Taxonomy_Inference { + process pcASV_AminoAcid_Taxonomy_Inference_RVDB { label 'high_cpus' From e8224307ed2f71e26a090d0188523e1c791b3ce8 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Tue, 30 Mar 2021 12:21:49 -0700 Subject: [PATCH 088/223] Add " --- vAMPirus.nf | 32 ++++++++++++++++---------------- 1 file changed, 16 insertions(+), 16 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 4434f5c..16c0f37 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1507,7 +1507,7 @@ if (params.DataCheck || params.Analyze) { virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - if [[ ${params.lca} == "T" ]] + if [[ "${params.lca}" == "T" ]] then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}' lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') echo "\$lcla" >> classfication.list @@ -1527,7 +1527,7 @@ if (params.DataCheck || params.Analyze) { virus="NO" gene="HIT" echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - if [[ ${params.lca} == "T" ]] + if [[ "${params.lca}" == "T" ]] then echo "N/A" >> classfication.list fi echo "\$s done." @@ -1544,7 +1544,7 @@ if (params.DataCheck || params.Analyze) { echo " " > sequence.list grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list rm "\$name"_tmpssasv.fasta - if [[ ${params.lca} == "T" ]] + if [[ "${params.lca}" == "T" ]] then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list classfication.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv @@ -1835,7 +1835,7 @@ if (params.DataCheck || params.Analyze) { virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - if [[ ${params.lca} == "T" ]] + if [[ "${params.lca}" == "T" ]] then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}' lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') echo "\$lcla" >> classfication.list @@ -1855,7 +1855,7 @@ if (params.DataCheck || params.Analyze) { virus="NO" gene="HIT" echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - if [[ ${params.lca} == "T" ]] + if [[ "${params.lca}" == "T" ]] then echo "N/A" >> classfication.list fi echo "\$s done." @@ -1872,7 +1872,7 @@ if (params.DataCheck || params.Analyze) { echo " " > sequence.list grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list rm "\$name"_tmpssasv.fasta - if [[ ${params.lca} == "T" ]] + if [[ "${params.lca}" == "T" ]] then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list classfication.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv @@ -2418,7 +2418,7 @@ if (params.DataCheck || params.Analyze) { grep ">" \${virdb} > headers.list headers="headers.list" name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" > access.list echo "[e-value]" > evalue.list @@ -2455,7 +2455,7 @@ if (params.DataCheck || params.Analyze) { virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - if [[ ${params.lca} == "T" ]] + if [[ "${params.lca}" == "T" ]] then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}' lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') echo "\$lcla" >> classfication.list @@ -2475,7 +2475,7 @@ if (params.DataCheck || params.Analyze) { virus="NO" gene="HIT" echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - if [[ ${params.lca} == "T" ]] + if [[ "${params.lca}" == "T" ]] then echo "N/A" >> classfication.list fi echo "\$s done." @@ -2492,7 +2492,7 @@ if (params.DataCheck || params.Analyze) { echo " " > sequence.list grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list rm "\$name"_tmpssasv.fasta - if [[ ${params.lca} == "T" ]] + if [[ "${params.lca}" == "T" ]] then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list classfication.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv @@ -3032,7 +3032,7 @@ if (params.DataCheck || params.Analyze) { virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - if [[ ${params.lca} == "T" ]] + if [[ "${params.lca}" == "T" ]] then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}' lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') echo "\$lcla" >> classfication.list @@ -3052,7 +3052,7 @@ if (params.DataCheck || params.Analyze) { virus="NO" gene="HIT" echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - if [[ ${params.lca} == "T" ]] + if [[ "${params.lca}" == "T" ]] then echo "N/A" >> classfication.list fi echo "\$s done." @@ -3069,7 +3069,7 @@ if (params.DataCheck || params.Analyze) { echo " " > sequence.list grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list rm "\$name"_tmpssasv.fasta - if [[ ${params.lca} == "T" ]] + if [[ "${params.lca}" == "T" ]] then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list classfication.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv @@ -3447,7 +3447,7 @@ if (params.DataCheck || params.Analyze) { virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - if [[ ${params.lca} == "T" ]] + if [[ "${params.lca}" == "T" ]] then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}' lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') echo "\$lcla" >> classfication.list @@ -3467,7 +3467,7 @@ if (params.DataCheck || params.Analyze) { virus="NO" gene="HIT" echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - if [[ ${params.lca} == "T" ]] + if [[ "${params.lca}" == "T" ]] then echo "N/A" >> classfication.list fi echo "\$s done." @@ -3484,7 +3484,7 @@ if (params.DataCheck || params.Analyze) { echo " " > sequence.list grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list rm "\$name"_tmpssasv.fasta - if [[ ${params.lca} == "T" ]] + if [[ "${params.lca}" == "T" ]] then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list classfication.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv From d13c8fbbdf4240fa155d16b3f4554063827e26bd Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Tue, 30 Mar 2021 12:24:40 -0700 Subject: [PATCH 089/223] Add missing ) --- vAMPirus.nf | 10 +++++----- 1 file changed, 5 insertions(+), 5 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 16c0f37..87dadb1 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1508,7 +1508,7 @@ if (params.DataCheck || params.Analyze) { echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt if [[ "${params.lca}" == "T" ]] - then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}' + then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}') lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') echo "\$lcla" >> classfication.list fi @@ -1836,7 +1836,7 @@ if (params.DataCheck || params.Analyze) { echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt if [[ "${params.lca}" == "T" ]] - then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}' + then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}') lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') echo "\$lcla" >> classfication.list fi @@ -2456,7 +2456,7 @@ if (params.DataCheck || params.Analyze) { echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt if [[ "${params.lca}" == "T" ]] - then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}' + then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}') lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') echo "\$lcla" >> classfication.list fi @@ -3033,7 +3033,7 @@ if (params.DataCheck || params.Analyze) { echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt if [[ "${params.lca}" == "T" ]] - then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}' + then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}') lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') echo "\$lcla" >> classfication.list fi @@ -3448,7 +3448,7 @@ if (params.DataCheck || params.Analyze) { echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt if [[ "${params.lca}" == "T" ]] - then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}' + then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}') lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') echo "\$lcla" >> classfication.list fi From f43d786aa5328a272bd9e69ba5529cfb769fd979 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Tue, 30 Mar 2021 12:32:19 -0700 Subject: [PATCH 090/223] Fix output filename --- vAMPirus.nf | 16 ++++++++-------- 1 file changed, 8 insertions(+), 8 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 87dadb1..a0fd3b8 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1690,7 +1690,7 @@ if (params.DataCheck || params.Analyze) { output: file("*.fasta") into tax_labeled_fasta_asv - tuple file("*_phyloseqObject.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond_asv + tuple file("*_phyloformat.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond_asv file("*_ASV*_summary_for_plot.csv") into taxplot_asv script: @@ -1787,7 +1787,7 @@ if (params.DataCheck || params.Analyze) { output: file("*.fasta") into tax_labeled_fasta_asv - tuple file("*_phyloseqObject.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond_asv + tuple file("*_phyloformat.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond_asv file("*_ASV*_summary_for_plot.csv") into taxplot_asv file("*_quick_Taxbreakdown.csv") into tax_table_asv @@ -2309,7 +2309,7 @@ if (params.DataCheck || params.Analyze) { file(asvs) from aminotypesBlast output: - tuple file("*_phyloseqObject.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AA_diamond + tuple file("*_phyloformat.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AA_diamond file("*_summary_for_plot.csv") into taxplot2 file("*TaxonomyLabels.fasta") into tax_labeled_fasta2 @@ -2406,7 +2406,7 @@ if (params.DataCheck || params.Analyze) { file(asvs) from aminotypesBlast output: - tuple file("*_phyloseqObject.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AA_diamond + tuple file("*_phyloformat.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AA_diamond file("*_summary_for_plot.csv") into taxplot2 file("*TaxonomyLabels.fasta") into tax_labeled_fasta2 file("*_quick_Taxbreakdown.csv") into tax_table_amino @@ -2880,7 +2880,7 @@ if (params.DataCheck || params.Analyze) { output: file("*.fasta") into ( pcASV_labeled ) - tuple file("*_phyloseqObject.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AAdiamond + tuple file("*_phyloformat.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AAdiamond tuple nid, file("*_summary_for_plot.csv") into taxplot3 script: @@ -2982,7 +2982,7 @@ if (params.DataCheck || params.Analyze) { output: file("*.fasta") into ( pcASV_labeled ) - tuple file("*_phyloseqObject.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AAdiamond + tuple file("*_phyloformat.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AAdiamond tuple nid, file("*_summary_for_plot.csv") into taxplot3 file("*_quick_Taxbreakdown.csv") into tax_table_pcasvnt @@ -3297,7 +3297,7 @@ if (params.DataCheck || params.Analyze) { output: file("*.fasta") into ( pcASV_labeledAA ) - tuple file("*phyloseqObject.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_potuaadiamond + tuple file("*phyloformat.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_potuaadiamond tuple nid, file("*_summary_for_plot.csv") into taxplot4 script: @@ -3398,7 +3398,7 @@ if (params.DataCheck || params.Analyze) { output: file("*.fasta") into ( pcASV_labeledAA ) - tuple file("*phyloseqObject.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_potuaadiamond + tuple file("*phyloformat.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_potuaadiamond tuple nid, file("*_summary_for_plot.csv") into taxplot4 file("*_quick_Taxbreakdown.csv") into tax_table_pcasvaa From 8f632573453269991f85c6e0fc06fa820a1b6d96 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Tue, 30 Mar 2021 12:35:45 -0700 Subject: [PATCH 091/223] Fix process error. --- vAMPirus.nf | 8 ++++---- 1 file changed, 4 insertions(+), 4 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index a0fd3b8..e55fe8c 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -2550,17 +2550,17 @@ if (params.DataCheck || params.Analyze) { iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.fasta.log | head -1 | awk '{print \$6}') + mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.log | head -1 | awk '{print \$6}') iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni elif [ "${params.nonparametric}" != "false" ];then - iqtree -s \${pre}_Aligned_informativeonly.fasta.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} elif [ "${params.parametric}" != "false" ];then - iqtree -s\${pre}_Aligned_informativeonly.fasta.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + iqtree -s\${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni else - iqtree -s \${pre}_Aligned_informativeonly.fasta.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni fi """ } From 467e0aa398202969d25bf54372a19cefe61ba432 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Tue, 30 Mar 2021 12:42:16 -0700 Subject: [PATCH 092/223] Fix file issue --- vAMPirus.nf | 6 +++++- 1 file changed, 5 insertions(+), 1 deletion(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index e55fe8c..c8e3dab 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1485,6 +1485,7 @@ if (params.DataCheck || params.Analyze) { then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list echo "[Classification]" > classfication.list else echo "skipped" >> \${name}_quick_Taxbreakdown.csv + echo "[Classification]" > classfication.list fi echo "extracting genes and names" touch new_"\$name"_asvnames.txt @@ -1813,6 +1814,7 @@ if (params.DataCheck || params.Analyze) { then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list echo "[Classification]" > classfication.list else echo "skipped" >> \${name}_quick_Taxbreakdown.csv + echo "[Classification]" > classfication.list fi echo "extracting genes and names" touch new_"\$name"_asvnames.txt @@ -2122,7 +2124,6 @@ if (params.DataCheck || params.Analyze) { input: file(counts) from asvcount_med - file(tree) from asvphy_med file(map) from asvgroupscsv output: @@ -2433,6 +2434,7 @@ if (params.DataCheck || params.Analyze) { then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list echo "[Classification]" > classfication.list else echo "skipped" >> \${name}_quick_Taxbreakdown.csv + echo "[Classification]" > classfication.list fi echo "extracting genes and names" touch new_"\$name"_asvnames.txt @@ -3010,6 +3012,7 @@ if (params.DataCheck || params.Analyze) { then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list echo "[Classification]" > classfication.list else echo "skipped" >> \${name}_quick_Taxbreakdown.csv + echo "[Classification]" > classfication.list fi echo "extracting genes and names" touch new_"\$name"_asvnames.txt @@ -3425,6 +3428,7 @@ if (params.DataCheck || params.Analyze) { then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list echo "[Classification]" > classfication.list else echo "skipped" >> \${name}_quick_Taxbreakdown.csv + echo "[Classification]" > classfication.list fi echo "extracting genes and names" touch new_"\$name"_asvnames.txt From 54594979d1a5885aa721f7dc3a4e95452c04c1b3 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Tue, 30 Mar 2021 12:49:46 -0700 Subject: [PATCH 093/223] Fix group labeling --- vAMPirus.nf | 16 ++++++++-------- 1 file changed, 8 insertions(+), 8 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index c8e3dab..1999c60 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -2123,11 +2123,11 @@ if (params.DataCheck || params.Analyze) { publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/MED/", mode: "copy", overwrite: true input: - file(counts) from asvcount_med - file(map) from asvgroupscsv + file(counts) from asvcount_med + file(map) from asvgroupscsv output: - file("${params.projtag}_ASV_Grouping_counts.csv") into asvgroupcounts + file("${params.projtag}_ASV_Grouping_counts.csv") into asvgroupcounts script: """ @@ -2652,7 +2652,7 @@ if (params.DataCheck || params.Analyze) { grep ">" "\$gid"_"\$uni" | awk -F ">" '{print \$2}' > asv.list seqtk subseq ../../${aminos} asv.list > Group"\${j}"_sequences.fasta for z in \$( cat asv.list) - do echo ""\$z",Group"\$gid","\$uni"" >> ${params.projtag}_AminoType_Grouping.csv + do echo ""\$z",Group"\$j","\$uni"" >> ${params.projtag}_AminoType_Grouping.csv done rm asv.list @@ -2722,12 +2722,12 @@ if (params.DataCheck || params.Analyze) { publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/MED/", mode: "copy", overwrite: true input: - file(counts) from aminocountmed - file(tree) from amino_repphy - file(map) from atygroupscsv + file(counts) from aminocountmed + file(tree) from amino_repphy + file(map) from atygroupscsv output: - file("${params.projtag}_AminoType_Grouping_counts.csv") into amino_groupcounts + file("${params.projtag}_AminoType_Grouping_counts.csv") into amino_groupcounts script: """ From 2f011fb0e489a3cd8d3b7079a04b1eaa5f72f822 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Tue, 30 Mar 2021 12:50:36 -0700 Subject: [PATCH 094/223] Fix typo --- vAMPirus.nf | 12 ++++++------ 1 file changed, 6 insertions(+), 6 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 1999c60..8df4db3 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1481,7 +1481,7 @@ if (params.DataCheck || params.Analyze) { echo "[ncASV#]" > otu.list echo "[Sequence length]" > length.list grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst - if [[ ${params.lsa} == "T" ]] + if [[ ${params.lca} == "T" ]] then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list echo "[Classification]" > classfication.list else echo "skipped" >> \${name}_quick_Taxbreakdown.csv @@ -1810,7 +1810,7 @@ if (params.DataCheck || params.Analyze) { echo "[ASV#]" > otu.list echo "[Sequence length]" > length.list grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst - if [[ ${params.lsa} == "T" ]] + if [[ ${params.lca} == "T" ]] then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list echo "[Classification]" > classfication.list else echo "skipped" >> \${name}_quick_Taxbreakdown.csv @@ -2054,7 +2054,7 @@ if (params.DataCheck || params.Analyze) { grep ">" "\$gid"_"\$uni" | awk -F ">" '{print \$2}' > asv.list seqtk subseq ../../${asvs} asv.list > Group"\${j}"_sequences.fasta for z in \$( cat asv.list) - do echo ""\$z",Group"\$gid","\$uni"" >> ${params.projtag}_ASV_Grouping.csv + do echo ""\$z",Group"\$j","\$uni"" >> ${params.projtag}_ASV_Grouping.csv done rm asv.list @@ -2430,7 +2430,7 @@ if (params.DataCheck || params.Analyze) { echo "[AminoType#]" > otu.list echo "[Sequence length]" > length.list grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst - if [[ ${params.lsa} == "T" ]] + if [[ ${params.lca} == "T" ]] then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list echo "[Classification]" > classfication.list else echo "skipped" >> \${name}_quick_Taxbreakdown.csv @@ -3008,7 +3008,7 @@ if (params.DataCheck || params.Analyze) { echo "[pcASV#]" > otu.list echo "[Sequence length]" > length.list grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst - if [[ ${params.lsa} == "T" ]] + if [[ ${params.lca} == "T" ]] then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list echo "[Classification]" > classfication.list else echo "skipped" >> \${name}_quick_Taxbreakdown.csv @@ -3424,7 +3424,7 @@ if (params.DataCheck || params.Analyze) { echo "[pcASV#]" > otu.list echo "[Sequence length]" > length.list grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst - if [[ ${params.lsa} == "T" ]] + if [[ ${params.lca} == "T" ]] then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list echo "[Classification]" > classfication.list else echo "skipped" >> \${name}_quick_Taxbreakdown.csv From 00ff0eb2b3564780bc0732f5d83cf2f366356fef Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Tue, 30 Mar 2021 13:13:08 -0700 Subject: [PATCH 095/223] WIP - Fix report channels --- vAMPirus.nf | 24 ++++++++++++++++++------ 1 file changed, 18 insertions(+), 6 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 8df4db3..4887746 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -3658,18 +3658,24 @@ if (params.DataCheck || params.Analyze) { taxplot1 -> ${params.projtag}_ASV_summary_for_plot.csv asv_heatmap -> ${params.projtag}_ASV_PercentID.matrix nucl_phyl_plot -> ${params.projtag}_ASV_iq.treefile + file("*_ASV_Grouping.csv") into asvgroupscsv + "${params.projtag}_ASV_Grouping_counts.csv") into asvgroupcounts + *_quick_Taxbreakdown.csv") into tax_table_asv + \\${params.projtag}_ASV_Group_Reps_iq.treefile */ + report_asv = Channel.create() - asv_counts_plots.mix(taxplot_asv, asv_heatmap, nucl_phyl_plot_asv).flatten().buffer(size:4).into(report_asv) + asv_counts_plots.mix(taxplot_asv, asv_heatmap, nucl_phyl_plot_asv, asvgroups, asvgroupcounts, asv_group_rep_tree, tax_table_asv).flatten().buffer(size:8).dump(tag:'asv').into(report_asv) if (params.ncASV) { report_ncasv = Channel.create() - notu_counts_plots.mix(taxplot_ncasv, notu_heatmap, nucl_phyl_plot_ncasv).groupTuple(by:0, size:4).into(report_ncasv) + notu_counts_plots.mix(taxplot_ncasv, notu_heatmap, nucl_phyl_plot_ncasv, tax_table_ncasv).groupTuple(by:0, size:5).dump(tag:'ncasv').into(report_ncasv) /* notu_counts_plots -> ${params.projtag}_ncASV${id}_counts.csv taxplot1a -> ${params.projtag}_ncASV${id}_summary_for_plot.csv notu_heatmap -> ${params.projtag}_ncASV${id}_PercentID.matrix nucl_phyl_plot -> ${params.projtag}_ncASV${id}_iq.treefile + ${params.projtag}_ncASV${id}_quick_Taxbreakdown.csv */ } else { report_ncasv = Channel.empty() @@ -3677,20 +3683,22 @@ if (params.DataCheck || params.Analyze) { if (params.pcASV) { report_pcasv_aa = Channel.create() - potu_Acounts.mix(taxplot4, potu_aa_heatmap, potu_Atree_plot).groupTuple(by:0, size:4).into(report_pcasv_aa) + potu_Acounts.mix(taxplot4, potu_aa_heatmap, potu_Atree_plot, tax_table_pcasvaa).groupTuple(by:0, size:5).dump(tag:'pcasv1').into(report_pcasv_aa) /*Report_pcASV_AminoAcid potu_Acounts -> ${params.projtag}_pcASV${id}_noTaxonomy_counts.csv taxplot4 -> ${params.projtag}_aminoacid_pcASV${id}_noTaxonomy_summary_for_plot.csv potu_aa_heatmap -> ${params.projtag}_aminoacid_pcASV${id}_noTaxonomy_PercentID.matrix potu_Atree_plot -> ${params.projtag}_aminoacid_pcASV${id}_noTaxonomy_iq.treefile + tax_table_pcasvaa -> ${params.projtag}_aminoacid_pcASV${id}_quick_Taxbreakdown.csv */ report_pcasv_nucl = Channel.create() - potu_Ncounts_for_report.mix(taxplot3, potu_nucl_heatmap, potu_Ntree_plot).groupTuple(by:0, size:4).into(report_pcasv_nucl) + potu_Ncounts_for_report.mix(taxplot3, potu_nucl_heatmap, potu_Ntree_plot, tax_table_pcasvnt).groupTuple(by:0, size:5).dump(tag:'pcasv2').into(report_pcasv_nucl) /*Report_pcASV_Nucleotide potu_Ncounts_for_report -> ${params.projtag}_nucleotide_pcASV${id}_noTaxonomy_counts.csv taxplot3 -> ${params.projtag}_nucleotide_pcASV${id}_noTaxonomy_summary_for_plot.csv potu_nucl_heatmap -> ${params.projtag}_nucleotide_pcASV${id}_noTaxonomy_PercentID.matrix potu_Ntree_plot -> ${params.projtag}_nucleotide_pcASV${id}_noTaxonomy_iq.treefile + tax_table_pcasvnt -> ${params.projtag}_nucleotide_pcASV${id}_quick_Taxbreakdown.csv */ } else { report_pcasv_aa = Channel.empty() @@ -3699,20 +3707,24 @@ if (params.DataCheck || params.Analyze) { if (!params.skipAminoTyping) { report_aminotypes = Channel.create() - aminocounts_plot.mix(taxplot2, aminotype_heatmap, amino_rax_plot).flatten().buffer(size:4).into(report_aminotypes) + aminocounts_plot.mix(taxplot2, aminotype_heatmap, amino_rax_plot, atygroupscsv, amino_group_rep_tree, amino_groupcounts, tax_table_amino).flatten().buffer(size:8).dump(tag:'amino').into(report_aminotypes) /* Report_AminoTypes aminocounts_plot -> ${params.projtag}_AminoType_counts.csv taxplot2 -> ${params.projtag}_AminoTypes_summary_for_plot.csv aminotype_heatmap -> ${params.projtag}_AminoTypes_PercentID.matrix amino_rax_plot -> ${params.projtag}_AminoTypes_iq.treefile + atygroupscsv -> *_AminoType_Grouping.csv + amino_group_rep_tree -> ${params.projtag}_AminoType_Group_Reps_iq.treefile + params.projtag}_AminoType_Grouping_counts.csv") into amino_groupcounts + *_quick_Taxbreakdown.csv") into tax_table_amino */ } else { report_aminotypes = Channel.empty() } report_all_ch = Channel.create() - report_asv.mix(report_ncasv, report_pcasv_aa, report_pcasv_nucl, report_aminotypes).map{it.flatten()}.into(report_all_ch) + report_asv.mix(report_ncasv, report_pcasv_aa, report_pcasv_nucl, report_aminotypes).map{it.flatten()}.dump(tag:'report').into(report_all_ch) process Report { From 4e339ec8483ead07d2c3a1fd361f103d6d5780b7 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Tue, 30 Mar 2021 13:15:36 -0700 Subject: [PATCH 096/223] WIP - Fix missing name --- vAMPirus.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 4887746..3baa0f3 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -3665,7 +3665,7 @@ if (params.DataCheck || params.Analyze) { */ report_asv = Channel.create() - asv_counts_plots.mix(taxplot_asv, asv_heatmap, nucl_phyl_plot_asv, asvgroups, asvgroupcounts, asv_group_rep_tree, tax_table_asv).flatten().buffer(size:8).dump(tag:'asv').into(report_asv) + asv_counts_plots.mix(taxplot_asv, asv_heatmap, nucl_phyl_plot_asv, asvgroupscsv, asvgroupcounts, asv_group_rep_tree, tax_table_asv).flatten().buffer(size:8).dump(tag:'asv').into(report_asv) if (params.ncASV) { report_ncasv = Channel.create() From 991e5ce1482c8ff485eae044ac10b75ee5d67cd9 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Tue, 30 Mar 2021 13:25:05 -0700 Subject: [PATCH 097/223] WIP - Fix missing channel --- vAMPirus.nf | 3 +-- 1 file changed, 1 insertion(+), 2 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 3baa0f3..79fc583 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1315,7 +1315,7 @@ if (params.DataCheck || params.Analyze) { if (params.ncASV) { reads_vsearch5_ch - .into{ asv_file_for_ncasvs; nuclFastas_forDiamond_asv_ch; nuclFastas_forCounts_asv_ch; nuclFastas_forphylogeny_asv; nuclFastas_forMatrix_asv_ch } + .into{ asv_file_for_ncasvs; nuclFastas_forDiamond_asv_ch; nuclFastas_forCounts_asv_ch; nuclFastas_forphylogeny_asv; nuclFastas_forMatrix_asv_ch; asv_for_med } process NucleotideBased_ASV_clustering { @@ -2023,7 +2023,6 @@ if (params.DataCheck || params.Analyze) { publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/ASVs/MED", mode: "copy", overwrite: true input: - file(asvs) from asv_for_med output: From 421581f00c189056c0555767c566a39b7cd6c5bf Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Tue, 30 Mar 2021 13:27:23 -0700 Subject: [PATCH 098/223] WIP - Fix diamond databse --- vAMPirus.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 79fc583..12171a4 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -3412,7 +3412,7 @@ if (params.DataCheck || params.Analyze) { grep ">" \${virdb} > headers.list headers="headers.list" name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" > access.list echo "[e-value]" > evalue.list From 956660165fac86c66c50475b4bab131433dd466e Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Tue, 30 Mar 2021 13:46:26 -0700 Subject: [PATCH 099/223] Fix space --- vAMPirus.nf | 3 +-- 1 file changed, 1 insertion(+), 2 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 12171a4..d03a85f 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1297,7 +1297,7 @@ if (params.DataCheck || params.Analyze) { .collect() output: - file("*.html") into datacheckreport + file("*.html") into datacheckreport script: """ @@ -3635,7 +3635,6 @@ if (params.DataCheck || params.Analyze) { cat tmp.names.csv tmp.reads.stats.csv >final_reads_stats.csv rm tmp.names.csv tmp.reads.stats.csv """ - } } else { From afc606397955925c9df9f4a77e95306ed0aa965d Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Wed, 31 Mar 2021 11:03:06 -0700 Subject: [PATCH 100/223] Add info of change to pyhton3 --- bin/virtualribosomev2/dna2pep.py | 2 ++ bin/virtualribosomev2/mod_translate.py | 2 ++ 2 files changed, 4 insertions(+) diff --git a/bin/virtualribosomev2/dna2pep.py b/bin/virtualribosomev2/dna2pep.py index 1ab8eff..73196d6 100755 --- a/bin/virtualribosomev2/dna2pep.py +++ b/bin/virtualribosomev2/dna2pep.py @@ -1,5 +1,7 @@ #!/usr/bin/env python3 +# Script was modified form original to use python3 + # Copyright 2006 Rasmus Wernersson, Technical University of Denmark # # This file is part of VirtualRibosome. diff --git a/bin/virtualribosomev2/mod_translate.py b/bin/virtualribosomev2/mod_translate.py index a555603..af9af91 100755 --- a/bin/virtualribosomev2/mod_translate.py +++ b/bin/virtualribosomev2/mod_translate.py @@ -1,5 +1,7 @@ #!/usr/bin/env python3 +# Script was modified form original to use python3 + # Copyright 2002,2003,2004,2005 Rasmus Wernersson, Technical University of Denmark # # This file is part of VirtualRibosome. From 4e06fe9fdcf92a42db0dc70237b1deca779f86af Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Wed, 31 Mar 2021 12:30:53 -0700 Subject: [PATCH 101/223] Fix tuple in output channels --- vAMPirus.nf | 9 +++++++-- 1 file changed, 7 insertions(+), 2 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index d03a85f..17a6b72 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1360,6 +1360,7 @@ if (params.DataCheck || params.Analyze) { file("*.fasta") into tax_labeled_fasta_ncasv tuple file("*_phyloformat.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond_ncasv tuple nid, file("*ncASV*summary_for_plot.csv") into taxplot_ncasv + tuple nid, file("*_quick_Taxbreakdown.csv") into tax_table_ncasv script: mtag="ID=" + nid @@ -1693,6 +1694,7 @@ if (params.DataCheck || params.Analyze) { file("*.fasta") into tax_labeled_fasta_asv tuple file("*_phyloformat.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond_asv file("*_ASV*_summary_for_plot.csv") into taxplot_asv + file("*_quick_Taxbreakdown.csv") into tax_table_asv script: """ @@ -2312,6 +2314,7 @@ if (params.DataCheck || params.Analyze) { tuple file("*_phyloformat.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AA_diamond file("*_summary_for_plot.csv") into taxplot2 file("*TaxonomyLabels.fasta") into tax_labeled_fasta2 + file("*_quick_Taxbreakdown.csv") into tax_table_amino script: """ @@ -2883,6 +2886,7 @@ if (params.DataCheck || params.Analyze) { file("*.fasta") into ( pcASV_labeled ) tuple file("*_phyloformat.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AAdiamond tuple nid, file("*_summary_for_plot.csv") into taxplot3 + tuple nid, file("*_quick_Taxbreakdown.csv") into tax_table_pcasvnt script: mtag="ID=" + nid @@ -2985,7 +2989,7 @@ if (params.DataCheck || params.Analyze) { file("*.fasta") into ( pcASV_labeled ) tuple file("*_phyloformat.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AAdiamond tuple nid, file("*_summary_for_plot.csv") into taxplot3 - file("*_quick_Taxbreakdown.csv") into tax_table_pcasvnt + tuple nid, file("*_quick_Taxbreakdown.csv") into tax_table_pcasvnt script: mtag="ID=" + nid @@ -3301,6 +3305,7 @@ if (params.DataCheck || params.Analyze) { file("*.fasta") into ( pcASV_labeledAA ) tuple file("*phyloformat.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_potuaadiamond tuple nid, file("*_summary_for_plot.csv") into taxplot4 + tuple nid, file("*_quick_Taxbreakdown.csv") into tax_table_pcasvaa script: mtag="ID=" + nid @@ -3402,7 +3407,7 @@ if (params.DataCheck || params.Analyze) { file("*.fasta") into ( pcASV_labeledAA ) tuple file("*phyloformat.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_potuaadiamond tuple nid, file("*_summary_for_plot.csv") into taxplot4 - file("*_quick_Taxbreakdown.csv") into tax_table_pcasvaa + tuple nid, file("*_quick_Taxbreakdown.csv") into tax_table_pcasvaa script: mtag="ID=" + nid From bdb3dcc3f397abf54f30a6ed5453fe272e69b2c3 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ram=C3=B3n=20Rivera-Vic=C3=A9ns?= Date: Wed, 31 Mar 2021 14:01:05 -0700 Subject: [PATCH 102/223] Fix paste issue --- vAMPirus.nf | 12 ++++++------ 1 file changed, 6 insertions(+), 6 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 17a6b72..a16c0bf 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -2128,7 +2128,7 @@ if (params.DataCheck || params.Analyze) { file(map) from asvgroupscsv output: - file("${params.projtag}_ASV_Grouping_counts.csv") into asvgroupcounts + file("${params.projtag}_ASV_Groupingcounts.csv") into asvgroupcounts script: """ @@ -2138,7 +2138,7 @@ if (params.DataCheck || params.Analyze) { do group=\$(grep -w \$x ${map} | awk -F "," '{print \$2}') echo "\$group" >> group.list done - paste -d',' group.list ${counts} > ${params.projtag}_ASV_Grouping_counts.csv + paste -d',' group.list ${counts} > ${params.projtag}_ASV_Groupingcounts.csv """ } } @@ -2729,7 +2729,7 @@ if (params.DataCheck || params.Analyze) { file(map) from atygroupscsv output: - file("${params.projtag}_AminoType_Grouping_counts.csv") into amino_groupcounts + file("${params.projtag}_AminoType_Groupingcounts.csv") into amino_groupcounts script: """ @@ -2739,7 +2739,7 @@ if (params.DataCheck || params.Analyze) { do group=\$(grep -w \$x ${map} | awk -F "," '{print \$2}') echo "\$group" >> group.list done - paste -d',' group.list ${counts} > ${params.projtag}_AminoType_Grouping_counts.csv + paste -d',' group.list ${counts} > ${params.projtag}_AminoType_Groupingcounts.csv """ } } @@ -3662,7 +3662,7 @@ if (params.DataCheck || params.Analyze) { asv_heatmap -> ${params.projtag}_ASV_PercentID.matrix nucl_phyl_plot -> ${params.projtag}_ASV_iq.treefile file("*_ASV_Grouping.csv") into asvgroupscsv - "${params.projtag}_ASV_Grouping_counts.csv") into asvgroupcounts + "${params.projtag}_ASV_Groupingcounts.csv") into asvgroupcounts *_quick_Taxbreakdown.csv") into tax_table_asv \\${params.projtag}_ASV_Group_Reps_iq.treefile */ @@ -3719,7 +3719,7 @@ if (params.DataCheck || params.Analyze) { amino_rax_plot -> ${params.projtag}_AminoTypes_iq.treefile atygroupscsv -> *_AminoType_Grouping.csv amino_group_rep_tree -> ${params.projtag}_AminoType_Group_Reps_iq.treefile - params.projtag}_AminoType_Grouping_counts.csv") into amino_groupcounts + params.projtag}_AminoType_Groupingcounts.csv") into amino_groupcounts *_quick_Taxbreakdown.csv") into tax_table_amino */ } else { From f3318f42322533273730a1696e2467be6492a520 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Tue, 6 Apr 2021 08:55:48 -0500 Subject: [PATCH 103/223] Added max n, merge staggere and max differences options in merging command --- vAMPirus.nf | 2 +- vampirus.config | 8 ++++++-- 2 files changed, 7 insertions(+), 3 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index a16c0bf..5534e34 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -755,7 +755,7 @@ if (params.DataCheck || params.Analyze) { script: """ - vsearch --fastq_mergepairs ${reads[0]} --reverse ${reads[1]} --threads ${task.cpus} --fastqout ${sample_id}_mergedclean.fastq --fastqout_notmerged_fwd ${sample_id}_notmerged_fwd.fastq --fastqout_notmerged_rev ${sample_id}_notmerged_rev.fastq --fastq_maxee ${params.maxEE} --relabel ${sample_id}. + vsearch --fastq_mergepairs ${reads[0]} --reverse ${reads[1]} --threads ${task.cpus} --fastqout ${sample_id}_mergedclean.fastq --fastqout_notmerged_fwd ${sample_id}_notmerged_fwd.fastq --fastqout_notmerged_rev ${sample_id}_notmerged_rev.fastq --fastq_maxdiffs ${params.diffs} --fastq_maxns ${params.maxn} --fastq_allowmergestagger --fastq_maxee ${params.maxEE} --relabel ${sample_id}. echo ${sample_id} > ${sample_id}.name """ diff --git a/vampirus.config b/vampirus.config index 05db904..a987467 100644 --- a/vampirus.config +++ b/vampirus.config @@ -29,8 +29,12 @@ params { minLen="400" // Maximum merged read length - reads with length equal to the specified max read length will be used to generate uniques and ASVs (safe to set at expected amplicon size to start) maxLen="420" - // Maximum expected error for vsearch merge command - maxEE="1" + // Maximum expected error for vsearch merge command - vsearch discard sequences with more than the specified number of expected errors + maxEE="3" + // Maximum number of non-matching nucleotides allowed in overlap region + diffs="20" + // Maximum number of "N"'s in a sequence - if above the specified value, sequence will be discarded + maxn="20" // Primer Removal parameters From df70d51b531f7fd7782b9a496a9f8cbbd76be274 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Tue, 6 Apr 2021 13:18:18 -0500 Subject: [PATCH 104/223] fixed aminoC --- vAMPirus.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 5534e34..0b04ac4 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -2643,7 +2643,7 @@ if (params.DataCheck || params.Analyze) { #entopy analysis entropy-analysis ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta #Decomposition - oligotype ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta-ENTROPY -o ${params.projtag}_asvMED_${params.asvC} -M 1 -c ${params.asvC} -N ${task.cpus} --skip-check-input --no-figures --skip-gen-html + oligotype ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta-ENTROPY -o ${params.projtag}_asvMED_${params.asvC} -M 1 -c ${params.aminoC} -N ${task.cpus} --skip-check-input --no-figures --skip-gen-html #generatemaps cd ./${params.projtag}_asvMED_${params.asvC}/OLIGO-REPRESENTATIVES/ echo "AminoType,Group,IDPattern" From b992dd402ef59a07f5511759333af868d56619b5 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Tue, 6 Apr 2021 13:20:33 -0500 Subject: [PATCH 105/223] fixed aminomed proc --- vAMPirus.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 0b04ac4..6f08f3f 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -2643,7 +2643,7 @@ if (params.DataCheck || params.Analyze) { #entopy analysis entropy-analysis ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta #Decomposition - oligotype ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta-ENTROPY -o ${params.projtag}_asvMED_${params.asvC} -M 1 -c ${params.aminoC} -N ${task.cpus} --skip-check-input --no-figures --skip-gen-html + oligotype ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta-ENTROPY -o ${params.projtag}_AminoTypeMED_${params.aminoC} -M 1 -c ${params.aminoC} -N ${task.cpus} --skip-check-input --no-figures --skip-gen-html #generatemaps cd ./${params.projtag}_asvMED_${params.asvC}/OLIGO-REPRESENTATIVES/ echo "AminoType,Group,IDPattern" From bf9c3ccdc81df2b299af9bfd04026d29f2dd08c8 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Tue, 6 Apr 2021 13:25:03 -0500 Subject: [PATCH 106/223] fixed again --- vAMPirus.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 6f08f3f..3af44b5 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -2645,7 +2645,7 @@ if (params.DataCheck || params.Analyze) { #Decomposition oligotype ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta-ENTROPY -o ${params.projtag}_AminoTypeMED_${params.aminoC} -M 1 -c ${params.aminoC} -N ${task.cpus} --skip-check-input --no-figures --skip-gen-html #generatemaps - cd ./${params.projtag}_asvMED_${params.asvC}/OLIGO-REPRESENTATIVES/ + cd ./${params.projtag}_AminoTypeMED_${params.aminoC}/OLIGO-REPRESENTATIVES/ echo "AminoType,Group,IDPattern" j=1 for x in *_unique; From 3af23fa3e716ba34be2abbe1249a8af4edcdc33f Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Thu, 8 Apr 2021 15:16:12 -0500 Subject: [PATCH 107/223] bbtools updated version --- vampirus_env.yml | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vampirus_env.yml b/vampirus_env.yml index 7ecc2be..fa80f48 100644 --- a/vampirus_env.yml +++ b/vampirus_env.yml @@ -19,7 +19,7 @@ dependencies: - mafft=7.464 - vsearch=2.14.2 - biopython=1.76 - - bbmap=38.79 + - bbmap=38.90 - trimal=1.4.1 - cd-hit=4.8.1 - emboss=6.5.7.0 From d300c0f91c4c78a2f59f749f77de23815bcc9b75 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Tue, 13 Apr 2021 10:37:52 -0500 Subject: [PATCH 108/223] edits --- README.md | 7 +- vAMPirus.nf | 5450 +++++++++++++++++++++++------------------------ vampirus.config | 2 +- 3 files changed, 2716 insertions(+), 2743 deletions(-) diff --git a/README.md b/README.md index c623462..aae8925 100644 --- a/README.md +++ b/README.md @@ -197,11 +197,11 @@ OR ### Analyze test => - `/path/to/nextflow run /path/to/vAMPirus.nf -c /path/to/vampirus.config -profile conda,test --Analyze --ncASV --pcASV --stats run` + `/path/to/nextflow run /path/to/vAMPirus.nf -c /path/to/vampirus.config -profile conda,test --Analyze --ncASV --pcASV --stats` OR - `nextflow run vAMPirus.nf -c vampirus.config -profile singularity,test --Analyze --ncASV --pcASV --stats run` + `nextflow run vAMPirus.nf -c vampirus.config -profile singularity,test --Analyze --ncASV --pcASV --stats` # Running vAMPirus @@ -209,6 +209,7 @@ OR If you done the setup and confirmed installation success with the test commands, you are good to get going with your own data. Before getting started edit the configuration file with the parameters and other options you plan to use. Here are some example vAMPirus launch commands: + ### DataCheck pipeline => Example 1. Launching the vAMPirus DataCheck pipeline using conda @@ -245,7 +246,7 @@ If an analysis is interrupted, you can use Nextflows "-resume" option that will For example if the analysis launched with the command from Example 6 above was interrupted, all you would need to do is add the "-resume" to the end of the command like so: - `nextflow run vAMPirus.nf -c vampirus.config -profile conda --Analyze --skipAminoTyping --stats run -resume` + `nextflow run vAMPirus.nf -c vampirus.config -profile conda --Analyze --skipAminoTyping --stats -resume` # Who to cite: diff --git a/vAMPirus.nf b/vAMPirus.nf index 3af44b5..4542133 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -33,7 +33,7 @@ def helpMessage() { Usage: - nextflow run vAMPirus.nf -c vampirus.config -profile [conda|singularity] --[Analyze|DataCheck] [--ncASV] [--pcASV] [--asvMED] [--aminoMED] [--stats] + nextflow run vAMPirus.nf -c vampirus.config -profile [conda|singularity] --[Analyze|DataCheck] [--ncASV] [--pcASV] --Help options-- @@ -57,12 +57,6 @@ def helpMessage() { --pcASV Set this option to have vAMPirus cluster nucleotide and translated ASVs into protein-based operational taxonomic units (pcASVs) - See options below to define a single percent similarity or a list - --Minimum Entropy Decomposition arguments-- - - --asvMED Set this option to perform Minimum Entropy Decomposition on ASV sequences, see manual for more information. You will need to set a value for --asvC to perform this analysis - - --amino_med Set this option to perform Minimum Entropy Decomposition on AminoType sequences, see manual for more information. You will need to set a value for --aminoC to perform this analysis - --Skip options-- --skipReadProcessing Set this option to skip all read processing steps in the pipeline @@ -71,9 +65,7 @@ def helpMessage() { --skipAdapterRemoval Set this option to skip adapter removal in the pipeline - --skipPrimerRemoval Set this option to skip primer removal process - - --skipMerging Set this option to skip read merging + --skipPrimerRemoval Set this option to skup Skip primer removal process --skipAminoTyping Set this option to skip AminoTyping processes @@ -81,10 +73,6 @@ def helpMessage() { --skipPhylogeny Set this option to skip phylogeny processes - --skipEMBOSS Set this option to skip EMBOSS processes - - --skipReport Set this option to skip html report generation - **NOTE** Most opitons below can be set using the configuration file (vampirus.config) to avoid a lengthy launch command. --Project/analysis information-- @@ -152,11 +140,6 @@ def helpMessage() { --minAA With --pcASV set, use this option to set the expected or minimum amino acid sequence length of open reading frames within your amplicon sequences - --Minimum Entropy Decomposition-- - - --asvC Number of high entropy positions to use for ASV MED analysis and generate "Groups" - - --aminoC Number of high entropy positions to use for AminoType MED analysis and generate "Groups" --Counts table generation-- @@ -175,11 +158,7 @@ def helpMessage() { --dbdir Path to Directory where database is being stored - --headers Set taxonomy database header format -> headers= "RefSeq" to toggle use of RefSeq header format; set to "RVDB" to signal the use of Reverence Viral DataBase (RVDB) headers - - --dbanno Path to directory hmm annotation .txt file - see manual for information on this. Leave as is if not planning on using. - - --lca Set --lca T if you would like to add taxonomic classification to taxonomy results - example: "ASV1, Viruses::Duplodnaviria::Heunggongvirae::Peploviricota::Herviviricetes::Herpesvirales::Herpesviridae::Gammaherpesvirinae::Macavirus" + --refseq Set "--refseq T" to toggle use of RefSeq header format; default is "F" to use Reverence Viral DataBase (RVDB) header --bitscore Set minimum bitscore to allow for best hit in taxonomy assignment @@ -226,208 +205,186 @@ def fullHelpMessage() { THIS IS A LONGER HELP WITH USAGE EXAMPLES vAMPirus v${workflow.manifest.version} ============================================================================================================================================================================================== - Steps: - 1- Before launching the vAMPirus.nf, be sure to run the vampirus_startup.sh script to install dependencies and/or databases - - 2- Test the vAMPirus installation with the provided test dataset (if you have ran the start up script, you can see STARTUP_HELP.txt for test commands and other examples) - - 3. Edit parameters in vampirus.config file - - 4. Launch the DataCheck pipeline to get summary information about your dataset - - 5. Change any parameters in vampirus.config file that might aid your analysis (e.g. clustering ID, maximum merged read length) - - 6. Launch the Analyze pipeline to perform a comprehensive analysis with your dataset - - 7. Explore results directories and produced final reports + Steps: + 1- Before launching the vAMPirus.nf, be sure to run the vampirus_startup.sh script to install dependencies and/or databases + 2- Test the vAMPirus installation with the provided test dataset (if you have ran the start up script, you can see STARTUP_HELP.txt for test commands and other examples) - Usage: + 3. Edit parameters in vampirus.config file - nextflow run vAMPirus.nf -c vampirus.config -profile [conda|singularity] --[Analyze|DataCheck] [--ncASV] [--pcASV] + 4. Launch the DataCheck pipeline to get summary information about your dataset + 5. Change any parameters in vampirus.config file that might aid your analysis (e.g. clustering ID, maximum merged read length) - --Help options-- + 6. Launch the Analyze pipeline to perform a comprehensive analysis with your dataset - --help Print help information + 7. Explore results directories and produced final reports - --fullHelp Print even more help information + Usage: - --Mandatory arguments (choose one)-- + nextflow run vAMPirus.nf -c vampirus.config -profile [conda|singularity] --[Analyze|DataCheck] [--ncASV] [--pcASV] - --Analyze Run absolutely everything - --DataCheck Assess how data performs with during processing and clustering + --Help options-- + --help Print help information - --ASV clustering arguments-- + --fullHelp Print even more help information - --ncASV Set this option to have vAMPirus cluster nucleotide amplicon sequence variants (ASVs) into nucleotide-based operational taxonomic units (ncASVs) - See options below to define a single percent similarity or a list - --pcASV Set this option to have vAMPirus cluster nucleotide and translated ASVs into protein-based operational taxonomic units (pcASVs) - See options below to define a single percent similarity or a list + --Mandatory arguments (choose one)-- + --Analyze Run absolutely everything - --Minimum Entropy Decomposition arguments-- + --DataCheck Assess how data performs with during processing and clustering - --asvMED Set this option to perform Minimum Entropy Decomposition on ASV sequences, see manual for more information. You will need to set a value for --asvC to perform this analysis - --amino_med Set this option to perform Minimum Entropy Decomposition on AminoType sequences, see manual for more information. You will need to set a value for --aminoC to perform this analysis + --ASV clustering arguments-- - --Skip options-- + --ncASV Set this option to have vAMPirus cluster nucleotide amplicon sequence variants (ASVs) into nucleotide-based operational taxonomic units (ncASVs) - See options below to define a single percent similarity or a list - --skipReadProcessing Set this option to skip all read processing steps in the pipeline + --pcASV Set this option to have vAMPirus cluster nucleotide and translated ASVs into protein-based operational taxonomic units (pcASVs) - See options below to define a single percent similarity or a list - --skipFastQC Set this option to skiip FastQC steps in the pipeline - --skipAdapterRemoval Set this option to skip adapter removal in the pipeline + --Skip options-- - --skipPrimerRemoval Set this option to skip primer removal process + --skipReadProcessing Set this option to skip all read processing steps in the pipeline - --skipMerging Set this option to skip read merging + --skipFastQC Set this option to skiip FastQC steps in the pipeline - --skipAminoTyping Set this option to skip AminoTyping processes + --skipAdapterRemoval Set this option to skip adapter removal in the pipeline - --skipTaxonomy Set this option to skip taxonomy assignment processes + --skipPrimerRemoval Set this option to skup Skip primer removal process - --skipPhylogeny Set this option to skip phylogeny processes + --skipAminoTyping Set this option to skip AminoTyping processes - --skipEMBOSS Set this option to skip EMBOSS processes + --skipTaxonomy Set this option to skip taxonomy assignment processes - --skipReport Set this option to skip html report generation + --skipPhylogeny Set this option to skip phylogeny processes + **NOTE** Most opitons below can be set using the configuration file (vampirus.config) to avoid a lengthy launch command. - **NOTE** Most opitons below can be set using the configuration file (vampirus.config) to avoid a lengthy launch command. + --Project/analysis information-- - --Project/analysis information-- + --projtag Set project name to be used as a prefix for output files - --projtag Set project name to be used as a prefix for output files + --metadata Set path to metadata spreadsheet file to be used for report generation (must be defined if generating report) - --metadata Set path to metadata spreadsheet file to be used for report generation (must be defined if generating report) + --reads Path to directory containing read libraries, must have *R{1,2}* in the library names - --reads Path to directory containing read libraries, must have *R{1,2}* in the library names + --workingdir Path to working directory where Nextflow will put all Nextflow and vAMPirus generated output files - --workingdir Path to working directory where Nextflow will put all Nextflow and vAMPirus generated output files + --outdir Name of results directory containing all output from the chosen pipeline (will be made within the working directory) - --outdir Name of results directory containing all output from the chosen pipeline (will be made within the working directory) + --Merged read length filtering-- - --Merged read length filtering-- + --minLen Minimum merged read length - reads below the specified maximum read length will be used for counts only - --minLen Minimum merged read length - reads below the specified maximum read length will be used for counts only + --maxLen Maximum merged read length - reads with length equal to the specified max read length will be used to identifying unique sequences and subsequent Amplicon Sequence Variant (ASV) analysis - --maxLen Maximum merged read length - reads with length equal to the specified max read length will be used to identifying unique sequences and subsequent Amplicon Sequence Variant (ASV) analysis + --maxEE Use this option to set the maximum expected error rate for vsearch merging. Default is 1. - --maxEE Use this option to set the maximum expected error rate for vsearch merging. Default is 1. + --Primer removal-- - --Primer removal-- + General primer removal parameters - General primer removal parameters + --primerLength Use this option to set the max primer length to restrict bbduk.sh primer trimming to the first x number of bases - --primerLength Use this option to set the max primer length to restrict bbduk.sh primer trimming to the first x number of bases + --maxkmer Maximum kmer length for bbduk.sh to use for primer detection and removal (must be shorter than your primer length; default = 13) - --maxkmer Maximum kmer length for bbduk.sh to use for primer detection and removal (must be shorter than your primer length; default = 13) + --minkmer Minimum kmer length for primer removal (default = 3) - --minkmer Minimum kmer length for primer removal (default = 3) + --minilen Minimum read length after adapter and primer removal (default = 200) - --minilen Minimum read length after adapter and primer removal (default = 200) + Single primer set removal- - Single primer set removal- + --GlobTrim Set this option to perform global trimming to reads to remove primer sequences. Example usage "--GlobTrim #basesfromforward,#basesfromreverse" - --GlobTrim Set this option to perform global trimming to reads to remove primer sequences. Example usage "--GlobTrim #basesfromforward,#basesfromreverse" + --fwd Forward primer sequence for reads to be detected and removed from reads (must specify reverse sequence if providing forward) - --fwd Forward primer sequence for reads to be detected and removed from reads (must specify reverse sequence if providing forward) + --rev Reverse primer sequence for reads to be detected and removed from reads (must specify forward sequence if providing reverse) - --rev Reverse primer sequence for reads to be detected and removed from reads (must specify forward sequence if providing reverse) + Multiple primer set removal- - Multiple primer set removal- + --multi Use this option to signal multiple primer sequence removal within the specified pipeline - --multi Use this option to signal multiple primer sequence removal within the specified pipeline + --primers Use this option to set the path to a fasta file with all of the primer sequences to be detected and removed from reads - --primers Use this option to set the path to a fasta file with all of the primer sequences to be detected and removed from reads + --Amplicon Sequence Variant (ASV) genration and clustering-- - --Amplicon Sequence Variant (ASV) genration and clustering-- + --alpha Alpha value for denoising - the higher the alpha the higher the chance of false positives in ASV generation (1 or 2) - --alpha Alpha value for denoising - the higher the alpha the higher the chance of false positives in ASV generation (1 or 2) + --minSize Minimum size or representation for sequence to be considered in ASV generation - --minSize Minimum size or representation for sequence to be considered in ASV generation + --clusterNuclID With --ncASV set, use this option to set a single percent similarity to cluster nucleotide ASV sequences into ncASVs by [ Example: --clusterNuclID .97 ] - --clusterNuclID With --ncASV set, use this option to set a single percent similarity to cluster nucleotide ASV sequences into ncASVs by [ Example: --clusterNuclID .97 ] + --clusterNuclIDlist With --ncASV set, use this option to perform nucleotide-based clustering of ASVs with a comma separated list of percent similarities [ Example: --clusterNuclIDlist .95,.96,.97,.98 ] - --clusterNuclIDlist With --ncASV set, use this option to perform nucleotide clustering with a comma separated list of percent similarities [ Example: --clusterNuclIDlist .95,.96,.97,.98 ] + --clusterAAID With --pcASV set, use this option to set a single percent similarity for protein-based ASV clustering to generate pcASVs[ Example: --clusterAAID .97 ] - --clusterAAID With --pcASV set, use this option to set a single percent similarity for protein-based ASV clustering to generation pcASVs [ Example: --clusterAAID .97 ] + --clusterAAIDlist With --pcASV set, use this option to perform protein-based ASV clustering to generate pcASVs with a comma separated list of percent similarities [ Example: --clusterAAIDlist .95,.96,.97,.98 ] - --clusterAAIDlist With --pcASV set, use this option to perform protein-based ASV clustering to generate pcASVs with a comma separated list of percent similarities [ Example: --clusterAAIDlist .95,.96,.97,.98 ] + --minAA With --pcASV set, use this option to set the expected or minimum amino acid sequence length of open reading frames within your amplicon sequences - --minAA With --pcASV set, use this option to set the expected or minimum amino acid sequence length of open reading frames within your amplicon sequences - --Minimum Entropy Decomposition-- + --Counts table generation-- - --asvC Number of high entropy positions to use for ASV MED analysis and generate "Groups" + --asvcountID Similarity ID to use for ASV counts - --aminoC Number of high entropy positions to use for AminoType MED analysis and generate "Groups" + --ProtCountID Minimum amino acid sequence similarity for hit to count - --Counts table generation-- + --ProtCountsLength Minimum alignment length for hit to count - --asvcountID Similarity ID to use for ASV counts + --ProtCountsBit Minimum bitscore for hit to be counted - --ProtCountID Minimum amino acid sequence similarity for hit to count - --ProtCountsLength Minimum alignment length for hit to count + --Taxonomy inference parameters-- - --ProtCountsBit Minimum bitscore for hit to be counted + --dbname Specify name of database to use for analysis + --dbdir Path to Directory where database is being stored - --Taxonomy inference parameters-- + --refseq Set "--refseq T" to toggle use of RefSeq header format; default is "F" to use Reverence Viral DataBase (RVDB) header - --dbname Specify name of database to use for analysis + --bitscore Set minimum bitscore to allow for best hit in taxonomy assignment - --dbdir Path to Directory where database is being stored + --minID Set minimum percent amino acid similarity for best hit to be counted in taxonomy assignment - --headers Set taxonomy database header format -> headers= "RefSeq" to toggle use of RefSeq header format; set to "RVDB" to signal the use of Reverence Viral DataBase (RVDB) headers + --minaln Set minimum amino acid alignment length for best hit to be counted in taxonomy assignment - --dbanno Path to directory hmm annotation .txt file - see manual for information on this. Leave as is if not planning on using. - --lca Set --lca T if you would like to add taxonomic classification to taxonomy results - example: "ASV1, Viruses::Duplodnaviria::Heunggongvirae::Peploviricota::Herviviricetes::Herpesvirales::Herpesviridae::Gammaherpesvirinae::Macavirus" + --Phylogeny analysis parameters-- - --bitscore Set minimum bitscore to allow for best hit in taxonomy assignment + Setting customs options for IQ-TREE (Example: "-option1 A -option2 B -option3 C -option4 D") - might be easier to set in the vampirus.config file at lines 108/109 - --minID Set minimum percent amino acid similarity for best hit to be counted in taxonomy assignment + --iqCustomnt Use option to set custom options to use in all IQTREE analyses with nuceoltide sequences - --minaln Set minimum amino acid alignment length for best hit to be counted in taxonomy assignment + --iqCustomaa Use option to set custom options to use in all IQTREE analyses with amino acid sequences + These options below you can set at the command, for example, to set to use model from ModelTest-NG with parametric bootstrapping --ModelTnt --ModelTaa --parametric - --Phylogeny analysis parameters-- + --ModelTnt=false Signal for IQ-TREE to use model determined by ModelTest-NG for all IQTREE analyses with nuceoltide sequences (Default is IQ-TREE will do automatic model testing with ModelFinder Plus) - Setting customs options for IQ-TREE (Example: "-option1 A -option2 B -option3 C -option4 D") - might be easier to set in the vampirus.config file at lines 108/109 + --ModelTaa=false Signal for IQ-TREE to use model determined by ModelTest-NG for all IQTREE analyses with amino acid sequences - --iqCustomnt Use option to set custom options to use in all IQTREE analyses with nuceoltide sequences + --parametric Set to use parametric bootstrapping in IQTREE analyses - --iqCustomaa Use option to set custom options to use in all IQTREE analyses with amino acid sequences + --nonparametric Set to use parametric bootstrapping in IQTREE analyses - These options below you can set at the command, for example, to set to use model from ModelTest-NG with parametric bootstrapping --ModelTnt --ModelTaa --parametric + --boots Number of bootstraps (recommended 1000 for parametric and 100 for non-parametric) - --ModelTnt=false Signal for IQ-TREE to use model determined by ModelTest-NG for all IQTREE analyses with nuceoltide sequences (Default is IQ-TREE will do automatic model testing with ModelFinder Plus) - --ModelTaa=false Signal for IQ-TREE to use model determined by ModelTest-NG for all IQTREE analyses with amino acid sequences + --Statistics options-- - --parametric Set to use parametric bootstrapping in IQTREE analyses + --stats Set "--stats run" to signal statstical tests to be performed and included in the final report - --nonparametric Set to use parametric bootstrapping in IQTREE analyses + --minimumCounts Minimum number of hit counts for a sample to have to be included in the downstream statistical analyses and report generation - --boots Number of bootstraps (recommended 1000 for parametric and 100 for non-parametric) - - - --Statistics options-- - - --stats Set "--stats run" to signal statstical tests to be performed and included in the final report - - --minimumCounts Minimum number of hit counts for a sample to have to be included in the downstream statistical analyses and report generation - - --trymax Maximum number of iterations performed by metaMDS + --trymax Maximum number of iterations performed by metaMDS ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||| @@ -457,7 +414,7 @@ def fullHelpMessage() { Example 4. Launching the vAMPirus Analyze pipeline with singularity with ASV and AminoType generation with all accesory analyses (taxonomy assignment, EMBOSS, IQTREE, statistics) - nextflow run vAMPirus.nf -c vampirus.config -profile singularity --Analyze --stats + nextflow run vAMPirus.nf -c vampirus.config -profile singularity --Analyze --stats run Example 5. Launching the vAMPirus Analyze pipeline with conda to perform multiple primer removal and protein-based clustering of ASVs, but skip most of the extra analyses @@ -465,11 +422,7 @@ def fullHelpMessage() { Example 6. Launching vAMPirus Analyze pipeline with conda to produce only ASV-related results - nextflow run vAMPirus.nf -c vampirus.config -profile conda --Analyze --skipAminoTyping --stats - - Example 7. Launching vAMPirus Analyze pipeline with conda to perform ASV analyses with Minimum Entropy Decomposition to form "Groups" - - nextflow run vAMPirus.nf -c vampirus.config -profile conda --Analyze --skipAminoTyping --stats --asvMED --asvC 24 + nextflow run vAMPirus.nf -c vampirus.config -profile conda --Analyze --skipAminoTyping --stats run Resuming analyses => @@ -492,6 +445,7 @@ if (params.help) { exit 0 } +// This will be printed to the user in each run. here thy can check if the values the selected are fine log.info """\ ================================================================================================================================================ vAMPirus v${workflow.manifest.version} - Virus Amplicon Sequencing Analysis Pipeline @@ -509,35 +463,15 @@ if (params.readsTest) { Channel .fromFilePairs(params.readsTest) .ifEmpty{ exit 1, "params.readTest was empty - no input files supplied" } - .into{ reads_ch; reads_qc_ch; reads_processing } + .into{ reads_ch; reads_qc_ch } } else { println("\n\tEverything ready for launch.\n") Channel .fromFilePairs("${params.reads}", checkIfExists: true) - .into{ reads_ch; reads_qc_ch; reads_processing } -} - -if (params.clusterNuclIDlist == "") { - a=params.clusterNuclID - slist=[a] - nnuc=slist.size() -} else { - msize=params.clusterNuclIDlist - slist=msize.split(',').collect{it as int} - nnuc=slist.size() -} - -if (params.clusterAAIDlist == "") { - b=params.clusterAAID - slist2=[b] - naa=slist2.size() -} else { - msize2=params.clusterAAIDlist - slist2=msize2.split(',').collect{it as int} - naa=slist2.size() + .into{ reads_ch; reads_qc_ch } } -if (params.DataCheck || params.Analyze) { +if (params.Analyze) { println("\n\tRunning vAMPirus Analyze pipeline - This might take a while, check out Nextflow tower (tower.nf) to remotely monitor the run.\n") @@ -596,7 +530,7 @@ if (params.DataCheck || params.Analyze) { } } - if (!params.skipReadProcessing || !params.skipMerging ) { + if (!params.skipReadProcessing) { if (!params.skipFastQC) { @@ -606,13 +540,13 @@ if (params.DataCheck || params.Analyze) { tag "${sample_id}" - publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/FastQC/PreClean", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/FastQC/PreClean", mode: "copy", overwrite: true input: tuple sample_id, file(reads) from reads_qc_ch output: - tuple sample_id, file("*_fastqc.{zip,html}") into fastqc_results + tuple sample_id, file("*_fastqc.{zip,html}") into fastqc_results_OAS script: """ @@ -629,8 +563,8 @@ if (params.DataCheck || params.Analyze) { tag "${sample_id}" - publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/AdapterRemoval", mode: "copy", overwrite: true, pattern: "*.filter.fq" - publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/AdapterRemoval/fastpOut", mode: "copy", overwrite: true, pattern: "*.fastp.{json,html}" + publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/AdapterRemoval", mode: "copy", overwrite: true, pattern: "*.filter.fq" + publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/AdapterRemoval/fastpOut", mode: "copy", overwrite: true, pattern: "*.fastp.{json,html}" input: tuple sample_id, file(reads) from reads_ch @@ -638,7 +572,7 @@ if (params.DataCheck || params.Analyze) { output: tuple sample_id, file("*.fastp.{json,html}") into fastp_results tuple sample_id, file("*.filter.fq") into reads_fastp_ch - file("*.csv") into ( fastp_csv_in1, fastp_csv_in2 ) + file("*.csv") into fastp_csv script: """ @@ -655,7 +589,6 @@ if (params.DataCheck || params.Analyze) { reads_ch .set{ reads_fastp_ch } fastp_results = Channel.empty() - } if (!params.skipPrimerRemoval) { @@ -666,7 +599,7 @@ if (params.DataCheck || params.Analyze) { tag "${sample_id}" - publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/PrimerRemoval", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/PrimerRemoval", mode: "copy", overwrite: true input: tuple sample_id, file(reads) from reads_fastp_ch @@ -703,10 +636,9 @@ if (params.DataCheck || params.Analyze) { } else { reads_fastp_ch .set{ reads_bbduk_ch } - } - if (!params.skipFastQC && !params.skipPrimerRemoval) { + if (!params.skipFastQC) { process QualityCheck_2 { @@ -714,13 +646,13 @@ if (params.DataCheck || params.Analyze) { tag "${sample_id}" - publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/FastQC/PostClean", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/FastQC/PostClean", mode: "copy", overwrite: true input: tuple sample_id, file(reads) from readsforqc2 output: - tuple sample_id, file("*_fastqc.{zip,html}") into fastqc2_results + tuple sample_id, file("*_fastqc.{zip,html}") into fastqc2_results_OAS script: """ @@ -728,50 +660,38 @@ if (params.DataCheck || params.Analyze) { """ } } - } else { - reads_ch - .set{ reads_bbduk_ch } - } - if (!params.skipMerging) { - - process Read_Merging { - - label 'norm_cpus' + process Read_Merging { - tag "${sample_id}" + label 'norm_cpus' - publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/Individual", mode: "copy", overwrite: true, pattern: "*mergedclean.fastq" - publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/Individual/notmerged", mode: "copy", overwrite: true, pattern: "*notmerged*.fastq" + tag "${sample_id}" - input: - tuple sample_id, file(reads) from reads_bbduk_ch + publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging/Individual", mode: "copy", overwrite: true, pattern: "*mergedclean.fastq" + publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging/Individual/notmerged", mode: "copy", overwrite: true, pattern: "*notmerged*.fastq" - output: - file("*_mergedclean.fastq") into reads_vsearch1_ch - file("*.name") into names - file("*notmerged*.fastq") into notmerged + input: + tuple sample_id, file(reads) from reads_bbduk_ch - script: - """ - vsearch --fastq_mergepairs ${reads[0]} --reverse ${reads[1]} --threads ${task.cpus} --fastqout ${sample_id}_mergedclean.fastq --fastqout_notmerged_fwd ${sample_id}_notmerged_fwd.fastq --fastqout_notmerged_rev ${sample_id}_notmerged_rev.fastq --fastq_maxdiffs ${params.diffs} --fastq_maxns ${params.maxn} --fastq_allowmergestagger --fastq_maxee ${params.maxEE} --relabel ${sample_id}. - echo ${sample_id} > ${sample_id}.name - """ + output: + file("*_mergedclean.fastq") into reads_vsearch1_ch + file("*.name") into names + file("*notmerged*.fastq") into notmerged - } + script: + """ + vsearch --fastq_mergepairs ${reads[0]} --reverse ${reads[1]} --threads ${task.cpus} --fastqout ${sample_id}_mergedclean.fastq --fastqout_notmerged_fwd ${sample_id}_notmerged_fwd.fastq --fastqout_notmerged_rev ${sample_id}_notmerged_rev.fastq --fastq_maxee ${params.maxEE} --relabel ${sample_id}. + echo ${sample_id} > ${sample_id}.name + """ - } else { - reads_bbduk_ch - .set{ reads_vsearch1_ch } } - - process Filtering_Prep1 { + process Compile_Reads { label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/LengthFiltering", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging/LengthFiltering", mode: "copy", overwrite: true input: file(reads) from reads_vsearch1_ch @@ -786,11 +706,11 @@ if (params.DataCheck || params.Analyze) { """ } - process Filtering_Prep2 { + process Compile_Names { label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging", mode: "copy", overwrite: true input: file(names) from names @@ -806,78 +726,39 @@ if (params.DataCheck || params.Analyze) { } - process Length_Filtering { + process Length_Filtering { //changed label 'norm_cpus' - publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/LengthFiltering", mode: "copy", overwrite: true, pattern: "*_merged_preFilt*.fasta" - publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging", mode: "copy", overwrite: true, pattern: "*Lengthfiltered.fastq" - publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/Histograms/pre_length_filtering", mode: "copy", overwrite: true, pattern: "*preFilt_*st.txt" - publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/Histograms/post_length_filtering", mode: "copy", overwrite: true, pattern: "*postFilt_*st.txt" + publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging/LengthFiltering", mode: "copy", overwrite: true, pattern: "*_merged_preFilt*.fasta" + publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging", mode: "copy", overwrite: true, pattern: "*Lengthfiltered.fastq" + publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging/Histograms/pre_length_filtering", mode: "copy", overwrite: true, pattern: "*preFilt_*st.txt" + publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging/Histograms/post_length_filtering", mode: "copy", overwrite: true, pattern: "*postFilt_*st.txt" input: file(reads) from collect_samples_ch output: - file("*_merged_preFilt_clean.fastq") into ( mergeforprotcounts, mergeforpcASVaacounts ) - - file("*_merged_preFilt_clean.fasta") into ( nuclCounts_mergedreads_asv_ch, nuclCounts_mergedreads_ncasv_ch, pcASV_mergedreads_ch ) + file("*_merged_preFilt_clean.fasta") into ( nuclCounts_mergedreads_ch, pcASV_mergedreads_ch ) file("*_merged_clean_Lengthfiltered.fastq") into reads_vsearch2_ch - - file("*preFilt_preClean_baseFrequency_hist.csv") into prefilt_basefreq - file("*preFilt_preClean_qualityScore_hist.csv") into prefilt_qualityscore - file("*preFilt_preClean_gcContent_hist.csv") into prefilt_gccontent - file("*preFilt_preClean_averageQuality_hist.csv") into prefilt_averagequality - file("*preFilt_preClean_length_hist.csv") into prefilt_length - - file("*postFilt_baseFrequency_hist.csv") into postFilt_basefreq - file("*postFilt_qualityScore_hist.csv") into postFilt_qualityscore - file("*postFilt_gcContent_hist.csv") into postFilt_gccontent - file("*postFilt_averageQuaulity_hist.csv") into postFilt_averagequality - file("*postFilt_length_hist.csv") into postFilt_length - - file("reads_per_sample_preFilt_preClean.csv") into reads_per_sample_preFilt - file("read_per_sample_postFilt_postClean.csv") into reads_per_sample_postFilt + file("*_merged_preFilt_clean.fastq") into ( mergeforprotcounts, mergeforpcASVaacounts ) + file("**hist.txt") into histos script: """ - #bbduk.sh in=${reads} bhist=${params.projtag}_all_merged_preFilt_preClean_baseFrequency_hist.txt qhist=${params.projtag}_all_merged_preFilt_preClean_qualityScore_hist.txt gchist=${params.projtag}_all_merged_preFilt_preClean_gcContent_hist.txt aqhist=${params.projtag}_all_merged_preFilt_preClean_averageQuality_hist.txt lhist=${params.projtag}_all_merged_preFilt__preClean_length_hist.txt gcbins=auto - #fastp -i ${reads} -o ${params.projtag}_merged_preFilt_clean.fastq -b ${params.maxLen} -l ${params.minLen} --thread ${task.cpus} -n 1 - #reformat.sh in=${params.projtag}_merged_preFilt_clean.fastq out=${params.projtag}_merged_preFilt_clean.fasta t=${task.cpus} - #bbduk.sh in=${params.projtag}_merged_preFilt_clean.fastq out=${params.projtag}_merged_clean_Lengthfiltered.fastq minlength=${params.maxLen} maxlength=${params.maxLen} t=${task.cpus} - #bbduk.sh in=${params.projtag}_merged_clean_Lengthfiltered.fastq bhist=${params.projtag}_all_merged_postFilt_baseFrequency_hist.txt qhist=${params.projtag}_all_merged_postFilt_qualityScore_hist.txt gchist=${params.projtag}_all_merged_postFilt_gcContent_hist.txt aqhist=${params.projtag}_all_merged_postFilt_averageQuaulity_hist.txt lhist=${params.projtag}_all_merged_postFilt_length_hist.txt gcbins=auto - - # from DC - bbduk.sh in=${reads} bhist=${params.projtag}_all_merged_preFilt_preClean_baseFrequency_hist.txt qhist=${params.projtag}_all_merged_preFilt_preClean_qualityScore_hist.txt gchist=${params.projtag}_all_merged_preFilt_preClean_gcContent_hist.txt aqhist=${params.projtag}_all_merged_preFilt_preClean_averageQuality_hist.txt lhist=${params.projtag}_all_merged_preFilt_preClean_length_hist.txt gcbins=auto - for x in *preFilt*hist.txt;do - pre=\$(echo \$x | awk -F ".txt" '{print \$1}') - cat \$x | tr "\t" "," > \${pre}.csv - rm \$x - done - reformat.sh in=${reads} out=${params.projtag}_preFilt_preclean.fasta t=${task.cpus} - echo "sample,reads" >> reads_per_sample_preFilt_preClean.csv - grep ">" ${params.projtag}_preFilt_preclean.fasta | awk -F ">" '{print \$2}' | awk -F "." '{print \$1}' | sort --parallel=${task.cpus} | uniq -c | sort -brg --parallel=${task.cpus} | awk '{print \$2","\$1}' >> reads_per_sample_preFilt_preClean.csv - rm ${params.projtag}_preFilt_preclean.fasta + bbduk.sh in=${reads} bhist=${params.projtag}_all_merged_preFilt_preClean_baseFrequency_hist.txt qhist=${params.projtag}_all_merged_preFilt_preClean_qualityScore_hist.txt gchist=${params.projtag}_all_merged_preFilt_preClean_gcContent_hist.txt aqhist=${params.projtag}_all_merged_preFilt_preClean_averageQuality_hist.txt lhist=${params.projtag}_all_merged_preFilt__preClean_length_hist.txt gcbins=auto fastp -i ${reads} -o ${params.projtag}_merged_preFilt_clean.fastq -b ${params.maxLen} -l ${params.minLen} --thread ${task.cpus} -n 1 reformat.sh in=${params.projtag}_merged_preFilt_clean.fastq out=${params.projtag}_merged_preFilt_clean.fasta t=${task.cpus} bbduk.sh in=${params.projtag}_merged_preFilt_clean.fastq out=${params.projtag}_merged_clean_Lengthfiltered.fastq minlength=${params.maxLen} maxlength=${params.maxLen} t=${task.cpus} bbduk.sh in=${params.projtag}_merged_clean_Lengthfiltered.fastq bhist=${params.projtag}_all_merged_postFilt_baseFrequency_hist.txt qhist=${params.projtag}_all_merged_postFilt_qualityScore_hist.txt gchist=${params.projtag}_all_merged_postFilt_gcContent_hist.txt aqhist=${params.projtag}_all_merged_postFilt_averageQuaulity_hist.txt lhist=${params.projtag}_all_merged_postFilt_length_hist.txt gcbins=auto - for x in *postFilt*hist.txt;do - pre=\$(echo \$x | awk -F ".txt" '{print \$1}') - cat \$x | tr "\t" "," > \${pre}.csv - rm \$x - done - reformat.sh in=${params.projtag}_merged_clean_Lengthfiltered.fastq out=${params.projtag}_merged_clean_Lengthfiltered.fasta t=${task.cpus} - echo "sample,reads" >> read_per_sample_postFilt_postClean.csv - grep ">" ${params.projtag}_merged_clean_Lengthfiltered.fasta | awk -F ">" '{print \$2}' | awk -F "." '{print \$1}' | sort --parallel=${task.cpus} | uniq -c | sort -brg --parallel=${task.cpus} | awk '{print \$2","\$1}' >> read_per_sample_postFilt_postClean.csv - """ + """ } - process Extracting_Uniques { + process Extract_Uniques { label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/Uniques", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging/Uniques", mode: "copy", overwrite: true input: file(reads) from reads_vsearch2_ch @@ -919,7 +800,7 @@ if (params.DataCheck || params.Analyze) { file(fasta) from reads_vsearch4_ch output: - file("*ASVs.fasta") into ( reads_vsearch5_ch, asv_med, nucl2aa, asvsforAminotyping, asvfastaforcounts, asvaminocheck ) + file("*ASVs.fasta") into ( reads_vsearch5_ch, nucl2aa, asvsforAminotyping, asvfastaforcounts, asvaminocheck ) script: """ @@ -929,448 +810,73 @@ if (params.DataCheck || params.Analyze) { // UNTIL HERE DEFAULT - if (params.DataCheck) { - - process NucleotideBased_ASV_clustering_DC { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Nucleotide", mode: "copy", overwrite: true, pattern: '*{.csv}' - - input: - file(fasta) from reads_vsearch5_ch - - output: - file("number_per_percentage_nucl.csv") into number_per_percent_nucl_plot - - script: - if (params.datacheckntIDlist) { - """ - for id in `echo ${params.datacheckntIDlist} | tr "," "\\n"`;do - vsearch --cluster_fast ${fasta} --centroids ${params.projtag}_ncASV\${id}.fasta --threads ${task.cpus} --relabel OTU --id \${id} - done - for x in *ncASV*.fasta;do - id=\$( echo \$x | awk -F "_ncASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') - numb=\$( grep -c ">" \$x ) - echo "\${id},\${numb}" >> number_per_percentage_nucl.csv - done - yo=\$(grep -c ">" ${fasta}) - echo "1.0,\${yo}" >> number_per_percentage_nucl.csv - """ - } - } - - process Translation_For_ProteinBased_Clustering_DC { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Aminoacid/translation", mode: "copy", overwrite: true - - input: - file(fasta) from nucl2aa - - output: - file("*ASVprotforclust.fasta") into clustering_aa - file("*_translation_report") into reportaa_VR - file("*_ASV_all.fasta") into asvfastaforaaclust - - script: - """ - ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_ASVprotforclust.fasta --report ${params.projtag}_translation_report - cp ${fasta} ${params.projtag}_ASV_all.fasta - """ - } + if (params.ncASV) { - process Protein_clustering_DC { + process NucleotideBased_ASV_clustering { label 'norm_cpus' - publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Aminoacid", mode: "copy", overwrite: true, pattern: '*{.csv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/ncASV", mode: "copy", overwrite: true, pattern: '*ncASV*.fasta' input: - file(fasta) from clustering_aa - file(asvs) from asvfastaforaaclust + file(fasta) from reads_vsearch5_ch output: - file("number_per_percentage_prot.csv") into number_per_percent_prot_plot - file("*aminoacid_pcASV1.0_noTaxonomy.fasta") into amino_med + tuple file("*_ncASV*.fasta"), file("*ASV.fasta") into ( nuclFastas_forDiamond_ch, nuclFastas_forCounts_ch, nuclFastas_forMatrix_ch) + tuple file("*_ncASV*.fasta"), file("*ASV.fasta") into nuclFastas_forphylogeny + script: - // add awk script to count seqs + if (params.clusterNuclIDlist) { """ - set +e - cp ${params.vampdir}/bin/rename_seq.py . - for id in `echo ${params.datacheckaaIDlist} | tr "," "\\n"`;do - if [ \${id} == ".55" ];then - word=3 - elif [ \${id} == ".65" ];then - word=4 - else - word=5 - fi - awk 'BEGIN{RS=">";ORS=""}length(\$2)>="${params.minAA}"{print ">"\$0}' ${fasta} > ${params.projtag}_filtered_proteins.fasta - cd-hit -i ${params.projtag}_filtered_proteins.fasta -n \${word} -c \${id} -o ${params.projtag}_pcASV\${id}.fasta - sed 's/>Cluster />Cluster_/g' ${params.projtag}_pcASV\${id}.fasta.clstr >${params.projtag}_pcASV\${id}.clstr - grep ">Cluster_" ${params.projtag}_pcASV\${id}.clstr >temporaryclusters.list - y=\$(grep -c ">Cluster_" ${params.projtag}_pcASV\${id}.clstr) - echo ">Cluster_"\${y}"" >> ${params.projtag}_pcASV\${id}.clstr - t=1 - b=1 - for x in \$(cat temporaryclusters.list);do - echo "Extracting \$x" - name="\$( echo \$x | awk -F ">" '{print \$2}')" - clust="pcASV"\${t}"" - echo "\${name}" - awk '/^>'\${name}'\$/,/^>Cluster_'\${b}'\$/' ${params.projtag}_pcASV\${id}.clstr > "\${name}"_"\${clust}"_tmp.list - t=\$(( \${t}+1 )) - b=\$(( \${b}+1 )) - done - ls *_tmp.list - u=1 - for x in *_tmp.list;do - name="\$(echo \$x | awk -F "_p" '{print \$1}')" - echo "\${name}" - cluster="\$(echo \$x | awk -F "_" '{print \$3}')" - echo "\${cluster}" - grep "ASV" \$x | awk -F ", " '{print \$2}' | awk -F "_" '{print \$1}' | awk -F ">" '{print \$2}' > \${name}_\${cluster}_seqs_tmps.list - seqtk subseq ${asvs} \${name}_\${cluster}_seqs_tmps.list > \${name}_\${cluster}_nucleotide_sequences.fasta - vsearch --cluster_fast \${name}_\${cluster}_nucleotide_sequences.fasta --id 0.2 --centroids \${name}_\${cluster}_centroids.fasta - grep ">" \${name}_\${cluster}_centroids.fasta >> \${name}_\${cluster}_tmp_centroids.list - for y in \$( cat \${name}_\${cluster}_tmp_centroids.list );do - echo ">\${cluster}_type"\$u"" >> \${name}_\${cluster}_tmp_centroid.newheaders - u=\$(( \${u}+1 )) - done - u=1 - ./rename_seq.py \${name}_\${cluster}_centroids.fasta \${name}_\${cluster}_tmp_centroid.newheaders \${cluster}_types_labeled.fasta - done - cat *_types_labeled.fasta >> ${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta - grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$3}' | awk -F "." '{print \$1}' >tmphead.list - grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$2}' | awk -F "," '{print \$1}' >tmplen.list - paste -d"," temporaryclusters.list tmphead.list >tmp.info.csv - grep ">" ${params.projtag}_pcASV\${id}.fasta >lala.list - j=1 - for x in \$(cat lala.list);do - echo ">${params.projtag}_pcASV\${j}" >>${params.projtag}_aminoheaders.list - echo "\${x},>${params.projtag}_pcASV\${j}" >>tmpaminotype.info.csv - j=\$(( \${j}+1 )) - done - rm lala.list - awk -F "," '{print \$2}' tmp.info.csv >>tmporder.list - for x in \$(cat tmporder.list);do - grep -w "\$x" tmpaminotype.info.csv | awk -F "," '{print \$2}' >>tmpder.list - done - paste -d "," temporaryclusters.list tmplen.list tmphead.list tmpder.list >${params.projtag}_pcASVCluster\${id}_summary.csv - ./rename_seq.py ${params.projtag}_pcASV\${id}.fasta ${params.projtag}_aminoheaders.list ${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta - stats.sh in=${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_aminoacid_clustered.gc gcformat=4 overwrite=true - stats.sh in=${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_nucleotide_clustered.gc gcformat=4 overwrite=true - awk 'BEGIN{RS=">";ORS=""}length(\$2)<"${params.minAA}"{print ">"\$0}' ${fasta} >${params.projtag}_pcASV\${id}_problematic_translations.fasta - if [ `wc -l ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk '{print \$1}'` -gt 1 ];then - grep ">" ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk -F ">" '{print \$2}' > problem_tmp.list - seqtk subseq ${asvs} problem_tmp.list > ${params.projtag}_pcASV\${id}_problematic_nucleotides.fasta - else - rm ${params.projtag}_pcASV\${id}_problematic_translations.fasta - fi - rm *.list - rm Cluster* - rm *types* - rm *tmp* - rm ${params.projtag}_pcASV\${id}.fast* + cp ${fasta} ./${params.projtag}_ASV.fasta + for id in `echo ${params.clusterNuclIDlist} | tr "," "\\n"`;do + vsearch --cluster_fast ${params.projtag}_ASV.fasta --centroids ${params.projtag}_ncASV\${id}.fasta --threads ${task.cpus} --relabel ncASV --id \${id} done - for x in *aminoacid*noTaxonomy.fasta;do - id=\$( echo \$x | awk -F "_noTax" '{print \$1}' | awk -F "pcASV" '{print \$2}') - numb=\$( grep -c ">" \$x) - echo "\${id},\${numb}" >> number_per_percentage_protz.csv - done - yesirr=\$( wc -l number_per_percentage_protz.csv | awk '{print \$1}') - tail -\$(( \${yesirr}-1 )) number_per_percentage_protz.csv > number_per_percentage_prot.csv - head -1 number_per_percentage_protz.csv >> number_per_percentage_prot.csv - rm number_per_percentage_protz.csv """ - } - - process ASV_Shannons_Entropy_Analysis { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Nucleotide/ShannonEntropy", mode: "copy", overwrite: true - - input: - file(asvs) from asv_med - - output: - - file("*_ASV_entropy_breakdown.csv") into asv_entro_csv - file("*Aligned_informativeonly.fasta-ENTROPY") into asv_entropy - file("*ASV*") into entrop - - script: - """ - set +e - #alignment - mafft --thread ${task.cpus} --maxiterate 15000 --auto ${asvs} > ${params.projtag}_ASVs_mafftAlign.fasta - #trimming - trimal -in ${params.projtag}_ASVs_mafftAlign.fasta -out ${params.projtag}_ASVs_mafftAligned.fasta -keepheader -fasta -automated1 - rm ${params.projtag}_ASVs_mafftAlign.fasta - o-trim-uninformative-columns-from-alignment ${params.projtag}_ASVs_mafftAligned.fasta - mv ${params.projtag}_ASVs_mafftAligned.fasta-TRIMMED ./${params.projtag}_ASVs_Aligned_informativeonly.fasta - #entopy analysis - entropy-analysis ${params.projtag}_ASVs_Aligned_informativeonly.fasta - #summarize entropy peaks - awk '{print \$2}' ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY >> tmp_value.list - for x in \$(cat tmp_value.list) - do echo "\$x" - if [[ \$(echo ""\$x" > 0.0"|bc -l) -eq 1 ]]; - then echo dope >> above-0.0-.list - fi - if [[ \$(echo ""\$x" > 0.1"|bc -l) -eq 1 ]]; - then echo dope >> above-0.1-.list - fi - if [[ \$(echo ""\$x" > 0.2"|bc -l) -eq 1 ]]; - then echo dope >> above-0.2-.list - fi - if [[ \$(echo ""\$x" > 0.3"|bc -l) -eq 1 ]]; - then echo dope >> above-0.3-.list - fi - if [[ \$(echo ""\$x" > 0.4"|bc -l) -eq 1 ]]; - then echo dope >> above-0.4-.list - fi - if [[ \$(echo ""\$x" > 0.5"|bc -l) -eq 1 ]]; - then echo dope >> above-0.5-.list - fi - if [[ \$(echo ""\$x" > 0.6"|bc -l) -eq 1 ]]; - then echo dope >> above-0.6-.list - fi - if [[ \$(echo ""\$x" > 0.7"|bc -l) -eq 1 ]]; - then echo dope >> above-0.7-.list - fi - if [[ \$(echo ""\$x" > 0.8"|bc -l) -eq 1 ]]; - then echo dope >> above-0.8-.list - fi - if [[ \$(echo ""\$x" > 0.9"|bc -l) -eq 1 ]]; - then echo dope >> above-0.9-.list - fi - if [[ \$(echo ""\$x" > 1.0"|bc -l) -eq 1 ]]; - then echo dope >> above-1.0-.list - fi - if [[ \$(echo ""\$x" > 1.5"|bc -l) -eq 1 ]]; - then echo dope >> above-1.5-.list - fi - done - echo "Entropy,Peaks_above" >> ${params.projtag}_ASV_entropy_breakdown.csv - for z in above*.list; - do entrop=\$(echo \$z | awk -F "-" '{print \$2}') - echo ""\$entrop", "\$(wc -l \$z | awk '{print \$1}')"" >> ${params.projtag}_ASV_entropy_breakdown.csv - done - rm above* - mv ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY ./tmp.fasta - echo "Base_position Shannons_Entropy" >> ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY - cat tmp.fasta >> ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY - rm tmp.fasta - - """ - - } - - process AminoType_Shannons_Entropy_Analysis { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Aminoacid/ShannonEntropy", mode: "copy", overwrite: true, pattern: '*{.csv}' - - input: - file(aminos) from amino_med - - output: - file("*AminoType_entropy_breakdown.csv") into amino_entro_csv - file ("*Aligned_informativeonly.fasta-ENTROPY") into amino_entropy - file("*AminoTypes*") into aminos - - script: - """ - #alignment - mafft --thread ${task.cpus} --maxiterate 15000 --auto ${aminos} > ${params.projtag}_AminoTypes_mafftAlign.fasta - #trimming - trimal -in ${params.projtag}_AminoTypes_mafftAlign.fasta -out ${params.projtag}_AminoTypes_mafftAligned.fasta -keepheader -fasta -automated1 - rm ${params.projtag}_AminoTypes_mafftAlign.fasta - o-trim-uninformative-columns-from-alignment ${params.projtag}_AminoTypes_mafftAligned.fasta - mv ${params.projtag}_AminoTypes_mafftAligned.fasta-TRIMMED ./${params.projtag}_AminoTypes_Aligned_informativeonly.fasta - #entropy analysis - entropy-analysis ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta --amino-acid-sequences - #summarize entropy peaks - awk '{print \$2}' ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta-ENTROPY >> tmp_value.list - for x in \$(cat tmp_value.list) - do echo "\$x" - if [[ \$(echo ""\$x" > 0.0"|bc -l) -eq 1 ]]; - then echo dope >> above-0.0-.list - fi - if [[ \$(echo ""\$x" > 0.1"|bc -l) -eq 1 ]]; - then echo dope >> above-0.1-.list - fi - if [[ \$(echo ""\$x" > 0.2"|bc -l) -eq 1 ]]; - then echo dope >> above-0.2-.list - fi - if [[ \$(echo ""\$x" > 0.3"|bc -l) -eq 1 ]]; - then echo dope >> above-0.3-.list - fi - if [[ \$(echo ""\$x" > 0.4"|bc -l) -eq 1 ]]; - then echo dope >> above-0.4-.list - fi - if [[ \$(echo ""\$x" > 0.5"|bc -l) -eq 1 ]]; - then echo dope >> above-0.5-.list - fi - if [[ \$(echo ""\$x" > 0.6"|bc -l) -eq 1 ]]; - then echo dope >> above-0.6-.list - fi - if [[ \$(echo ""\$x" > 0.7"|bc -l) -eq 1 ]]; - then echo dope >> above-0.7-.list - fi - if [[ \$(echo ""\$x" > 0.8"|bc -l) -eq 1 ]]; - then echo dope >> above-0.8-.list - fi - if [[ \$(echo ""\$x" > 0.9"|bc -l) -eq 1 ]]; - then echo dope >> above-0.9-.list - fi - if [[ \$(echo ""\$x" > 1.0"|bc -l) -eq 1 ]]; - then echo dope >> above-1.0-.list - fi - if [[ \$(echo ""\$x" > 1.5"|bc -l) -eq 1 ]]; - then echo dope >> above-1.5-.list - fi - done - echo "Entropy,Peaks_above" >> ${params.projtag}_AminoType_entropy_breakdown.csv - for z in above*.list; - do entrop=\$(echo \$z | awk -F "-" '{print \$2}') - echo ""\$entrop", "\$(wc -l \$z | awk '{print \$1}')"" >> ${params.projtag}_AminoType_entropy_breakdown.csv - done - rm above* - mv ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta-ENTROPY ./tmp.fasta - echo "Base_position Shannons_Entropy" >> ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta-ENTROPY - cat tmp.fasta >> ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta-ENTROPY - rm tmp.fasta - """ - } - - if (!params.skipReadProcessing || !params.skipMerging ) { - - process combine_csv_DC { - - input: - file(csv) from fastp_csv_in1 - .collect() - - output: - file("final_reads_stats.csv") into fastp_csv_dc - - script: - """ - cat ${csv} >all_reads_stats.csv - head -n1 all_reads_stats.csv >tmp.names.csv - cat all_reads_stats.csv | grep -v ""Sample,Total_"" >tmp.reads.stats.csv - cat tmp.names.csv tmp.reads.stats.csv >final_reads_stats.csv - rm tmp.names.csv tmp.reads.stats.csv - """ - - } - } else { - - process skip_combine_csv_DC { - output: - file("filter_reads.txt") into fastp_csv_dc - - script: - """ - echo "Read processing steps skipped." >filter_reads.txt - """ - } - } - - report_dc_in = Channel.create() - fastp_csv_dc.mix( reads_per_sample_preFilt, reads_per_sample_postFilt, prefilt_basefreq, postFilt_basefreq, prefilt_qualityscore, postFilt_qualityscore, prefilt_gccontent, postFilt_gccontent, prefilt_averagequality, postFilt_averagequality, prefilt_length, postFilt_length, number_per_percent_nucl_plot, number_per_percent_prot_plot, amino_entro_csv, amino_entropy, asv_entro_csv, asv_entropy).into(report_dc_in) - - process Report_DataCheck { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/DataCheck/Report", mode: "copy", overwrite: true, pattern: '*.{html}' - - input: - file(files) from report_dc_in - .collect() - - output: - file("*.html") into datacheckreport - - script: + } else if (params.clusterNuclID) { """ - cp ${params.vampdir}/bin/vAMPirus_DC_Report.Rmd . - cp ${params.vampdir}/example_data/conf/vamplogo.png . - Rscript -e "rmarkdown::render('vAMPirus_DC_Report.Rmd',output_file='${params.projtag}_DataCheck_Report.html')" ${params.projtag} \ - ${params.skipReadProcessing} \ - ${params.skipMerging} \ - ${params.skipAdapterRemoval} + cp ${fasta} ./${params.projtag}_ASV.fasta + id=${params.clusterNuclID} + vsearch --cluster_fast ${params.projtag}_ASV.fasta --centroids ${params.projtag}_ncASV\${id}.fasta --threads ${task.cpus} --relabel ncASV --id \${id} """ + } } + } else { + reads_vsearch5_ch + .into{ nuclFastas_forDiamond_ch; nuclFastas_forCounts_ch; nuclFastas_forphylogeny; nuclFastas_forMatrix_ch } + } - } else if (params.Analyze) { + if (!params.skipTaxonomy) { if (params.ncASV) { - reads_vsearch5_ch - .into{ asv_file_for_ncasvs; nuclFastas_forDiamond_asv_ch; nuclFastas_forCounts_asv_ch; nuclFastas_forphylogeny_asv; nuclFastas_forMatrix_asv_ch; asv_for_med } - - process NucleotideBased_ASV_clustering { - - label 'norm_cpus' + process Nucleotide_Taxonomy_Inference { - tag "${mtag}" + label 'high_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/ncASV", mode: "copy", overwrite: true, pattern: '*ncASV*.fasta' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy", mode: "copy", overwrite: true, pattern: '*_ASV*.{fasta,csv,tsv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Taxonomy", mode: "copy", overwrite: true, pattern: '*ncASV*.{fasta,csv,tsv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*_ASV*dmd.out' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*ncASV*dmd.out' input: - each x from 1..nnuc - file(fasta) from asv_file_for_ncasvs + tuple file(notus), file(asvs) from nuclFastas_forDiamond_ch output: - tuple nid, file("*_ncASV*.fasta") into ( nuclFastas_forphylogeny_ncasv, nuclFastas_forDiamond_ncasv_ch, nuclFastas_forCounts_ncasv_ch, nuclFastas_forMatrix_ncasv_ch ) + file("*.fasta") into tax_labeled_fasta + tuple file("*_phyloseqObject.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond + file("*ncASV*summary_for_plot.csv") into taxplot1a + file("*_ASV*_summary_for_plot.csv") into taxplot1 script: - nid=slist.get(x-1) - mtag="ID=" + slist.get(x-1) - """ - vsearch --cluster_fast ${fasta} --centroids ${params.projtag}_ncASV${nid}.fasta --threads ${task.cpus} --relabel ncASV --id .${nid} """ - } - - if (!params.skipTaxonomy) { - - if (params.headers == "RefSeq") { - - process ncASV_Taxonomy_Inference_RefSeq { /////// editttt - - label 'high_cpus' - - tag "${mtag}" - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Taxonomy", mode: "copy", overwrite: true, pattern: '*ncASV*.{fasta,csv,tsv}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*ncASV*dmd.out' - - input: - tuple nid, file(asvs) from nuclFastas_forDiamond_ncasv_ch - - output: - file("*.fasta") into tax_labeled_fasta_ncasv - tuple file("*_phyloformat.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond_ncasv - tuple nid, file("*ncASV*summary_for_plot.csv") into taxplot_ncasv - tuple nid, file("*_quick_Taxbreakdown.csv") into tax_table_ncasv - - script: - mtag="ID=" + nid - """ - cp ${params.vampdir}/bin/rename_seq.py . - virdb=${params.dbdir}/${params.dbname} - grep ">" \${virdb} > headers.list - headers="headers.list" - name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + cp ${params.vampdir}/bin/rename_seq.py . + virdb=${params.dbdir}/${params.dbname} + grep ">" \${virdb} > headers.list + headers="headers.list" + for filename in ${notus};do + name=\$(ls \${filename} | awk -F ".fasta" '{print \$1}') + diamond blastx -q \${filename} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" > access.list echo "[e-value]" > evalue.list @@ -1378,14 +884,18 @@ if (params.DataCheck || params.Analyze) { echo "[Percent ID (aa)]" > pid.list echo "[Organism ID]" > "\$name"_virus.list echo "[Gene]" > "\$name"_genes.list - echo "[ncASV#]" > otu.list - echo "[Sequence length]" > length.list - grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst + grep ">" \${filename} | awk -F ">" '{print \$2}' > seqids.lst echo "extracting genes and names" touch new_"\$name"_asvnames.txt - for s in \$(cat seqids.lst);do - echo "Checking for \$s hit in diamond output" - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + j=1 + if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then + echo "[ncASV#]" > otu.list + echo "[ncASV sequence length]" > length.list + for s in \$(cat seqids.lst);do + echo "Checking for \$s hit in diamond output" + if [[ ${params.refseq} == "T" ]];then + echo "RefSeq headers specified" + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then echo "Yep, there was a hit for \$s" echo "Extracting the information now:" acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') @@ -1401,9 +911,10 @@ if (params.DataCheck || params.Analyze) { echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">"\${s}"_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + echo ">ncASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) echo "\$s done." - else + else echo "Ugh, there was no hit for \$s .." echo "We still love \$s though and we will add it to the final fasta file" echo "\$s" >> otu.list @@ -1416,356 +927,1530 @@ if (params.DataCheck || params.Analyze) { echo "NO_HIT" >> length.list virus="NO" gene="HIT" - echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + echo ">ncASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) echo "\$s done." - fi - done - echo "Now editing "\$name" fasta headers" - ###### rename_seq.py - ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta - awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta - echo "[Sequence header]" > newnames.list - cat new_"\$name"_asvnames.txt >> newnames.list - touch sequence.list - echo " " > sequence.list - grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list - rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - for x in *phyloformat.csv;do - echo "\$x" - lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) - tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; - sed 's/ /_/g' tmpcol.list > tmp2col.list; - cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; + fi + else + echo "Using RVDB headers." + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">ncASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list + virus="NO" + gene="HIT" + echo ">ncASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + fi + fi + echo "Done with \$s" + done + fi + echo "Now editing "\$name" fasta headers" + ###### rename_seq.py + ./rename_seq.py \${filename} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta + echo "[Sequence header]" > newnames.list + cat new_"\$name"_asvnames.txt >> newnames.list + touch sequence.list + echo " " > sequence.list + grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list + rm "\$name"_tmpssasv.fasta + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + for x in *phyloseqObject.csv;do + echo "\$x" + lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) + tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; + sed 's/ /_/g' tmpcol.list > tmp2col.list; + cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; rm tmpcol.list tmp2col.list done - rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list + rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; + rm *asvnames.txt + rm "\$name"_virus.list + rm "\$name"_genes.list + rm newnames.list + rm access.list + echo "Taxonomy inferred for: \${filename} " + done + for filename in ${asvs};do + name=\$(ls \${filename} | awk -F ".fasta" '{print \$1}') + diamond blastx -q \${filename} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + echo "Preparing lists to generate summary .csv's" + echo "[Best hit accession number]" > access.list + echo "[e-value]" > evalue.list + echo "[Bitscore]" > bit.list + echo "[Percent ID (aa)]" > pid.list + echo "[Organism ID]" > "\$name"_virus.list + echo "[Gene]" > "\$name"_genes.list + grep ">" \${filename} | awk -F ">" '{print \$2}' > seqids.lst + echo "extracting genes and names" + touch new_"\$name"_asvnames.txt + j=1 + if [ `echo \${filename} | grep -c "ASV"` -eq 1 ];then + for s in \$(cat seqids.lst);do + echo "[ASV#]" > otu.list + echo "[ASV sequence length]" > length.list + echo "Checking for \$s hit in diamond output" + if [[ ${params.refseq} == "T" ]];then + echo "RefSeq headers specified" + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list + virus="NO" + gene="HIT" + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + fi + else + echo "Using RVDB headers." + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list + virus="NO" + gene="HIT" + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + fi + fi + echo "Done with \$s" + done + fi + echo "Now editing "\$name" fasta headers" + ###### rename_seq.py + ./rename_seq.py \${filename} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta + echo "[Sequence header]" > newnames.list + cat new_"\$name"_asvnames.txt >> newnames.list + touch sequence.list + echo " " > sequence.list + grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list + rm "\$name"_tmpssasv.fasta + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + for x in *phyloseqObject.csv;do + echo "\$x" + lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) + tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; + sed 's/ /_/g' tmpcol.list > tmp2col.list; + cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; + rm tmpcol.list tmp2col.list + done + rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; + rm *asvnames.txt + rm "\$name"_virus.list + rm "\$name"_genes.list + rm newnames.list + rm access.list + echo "Taxonomy inferred for: \${filename} " + done + rm headers.list + """ + } + + } else { + + process ASV_Taxonomy_Inference { + + label 'high_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy", mode: "copy", overwrite: true, pattern: '*_ASV*.{fasta,csv,tsv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*_ASV*dmd.out' + + input: + file(reads) from nuclFastas_forDiamond_ch + + output: + file("*.fasta") into tax_labeled_fasta + tuple file("*_phyloseqObject.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond + file("*_ASV*_summary_for_plot.csv") into taxplot1 + script: + """ + cp ${params.vampdir}/bin/rename_seq.py . + virdb=${params.dbdir}/${params.dbname} + grep ">" \${virdb} > headers.list + headers="headers.list" + for filename in ${reads};do + name=\$(ls \${filename} | awk -F ".fasta" '{print \$1}') + diamond blastx -q \${filename} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + echo "Preparing lists to generate summary .csv's" + echo "[Best hit accession number]" > access.list + echo "[e-value]" > evalue.list + echo "[Bitscore]" > bit.list + echo "[Percent ID (aa)]" > pid.list + echo "[Organism ID]" > "\$name"_virus.list + echo "[Gene]" > "\$name"_genes.list + grep ">" \${filename} | awk -F ">" '{print \$2}' > seqids.lst + echo "extracting genes and names" + touch new_"\$name"_asvnames.txt + j=1 + if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then + echo "[ASV#]" > otu.list + echo "[ASV sequence length]" > length.list + for s in \$(cat seqids.lst);do + echo "Checking for \$s hit in diamond output" + if [[ ${params.refseq} == "T" ]];then + echo "RefSeq headers specified" + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list + virus="NO" + gene="HIT" + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + fi + else + echo "Using RVDB headers." + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list + virus="NO" + gene="HIT" + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + fi + fi + echo "Done with \$s" + done + else + for s in \$(cat seqids.lst);do + echo "[ASV#]" > otu.list + echo "[ASV sequence length]" > length.list + echo "Checking for \$s hit in diamond output" + if [[ ${params.refseq} == "T" ]];then + echo "RefSeq headers specified" + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list + virus="NO" + gene="HIT" + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + fi + else + echo "Using RVDB headers." + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list + virus="NO" + gene="HIT" + echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + fi + fi + echo "Done with \$s" + done + fi + echo "Now editing "\$name" fasta headers" + ###### rename_seq.py + ./rename_seq.py \${filename} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta + echo "[Sequence header]" > newnames.list + cat new_"\$name"_asvnames.txt >> newnames.list + touch sequence.list + echo " " > sequence.list + grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list + rm "\$name"_tmpssasv.fasta + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + for x in *phyloseqObject.csv;do + echo "\$x" + lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) + tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; + sed 's/ /_/g' tmpcol.list > tmp2col.list; + cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; + rm tmpcol.list tmp2col.list + done + rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; + rm *asvnames.txt + rm "\$name"_virus.list + rm "\$name"_genes.list + rm newnames.list + rm access.list + echo "Taxonomy inferred for: \${filename} " + done + rm headers.list + """ + } + } + } + + if (params.ncASV) { + + process Generate_Counts_Tables_Nucleotide { + + label 'norm_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Counts", mode: "copy", overwrite: true, pattern: '*_ASV*.{biome,csv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Counts", mode: "copy", overwrite: true, pattern: '*ncASV*.{biome,csv}' + + input: + tuple file(notus), file(asvs) from nuclFastas_forCounts_ch + file(merged) from nuclCounts_mergedreads_ch + + output: + tuple file("*_counts.csv"), file("*_counts.biome") into counts_vsearch + file("*ncASV*counts.csv") into notu_counts_plots + file("*_ASV*counts.csv") into asv_counts_plots + + script: + """ + for filename in ${notus};do + if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then + ident=\$( echo \${filename} | awk -F "ncASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') + name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') + vsearch --usearch_global ${merged} --db \${filename} --id \${ident} --threads ${task.cpus} --otutabout \${name}_counts.txt --biomout \${name}_counts.biome + cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv + sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv + rm \${name}_count.csv + fi + done + if [ `echo ${asvs} | grep -c "ASV"` -eq 1 ];then + name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') + vsearch --usearch_global ${merged} --db ${asvs} --id ${params.asvcountID} --threads ${task.cpus} --otutabout "\$name"_counts.txt --biomout "\$name"_counts.biome + cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv + sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv + rm \${name}_count.csv + fi + """ + } + } else { + process Generate_ASV_Counts_Tables { + + label 'norm_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Counts", mode: "copy", overwrite: true, pattern: '*ASV*.{biome,csv}' + + input: + file(asvs) from nuclFastas_forCounts_ch + file(merged) from nuclCounts_mergedreads_ch + + output: + tuple file("*_counts.csv"), file("*_counts.biome") into counts_vsearch + file("*_ASV*counts.csv") into asv_counts_plots + + script: + """ + if [ `echo ${asvs} | grep -c "ASV"` -eq 1 ];then + name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') + vsearch --usearch_global ${merged} --db ${asvs} --id ${params.asvcountID} --threads ${task.cpus} --otutabout "\$name"_counts.txt --biomout "\$name"_counts.biome + cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv + sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv + rm \${name}_count.csv + fi + """ + } + } + + if (params.ncASV) { + + process Generate_Nucleotide_Matrix { + + label 'low_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Matrix", mode: "copy", overwrite: true, pattern: '*_ASV*PercentID.matrix' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Matrix", mode: "copy", overwrite: true, pattern: '*ncASV*PercentID.matrix' + + input: + tuple file(notus), file(asvs) from nuclFastas_forMatrix_ch + + output: + file("*.matrix") into clustmatrices + file("*ncASV*PercentID.matrix") into notu_heatmap + file("*_ASV*PercentID.matrix") into asv_heatmap + + script: + // remove if statement later (no fin) + """ + for filename in ${notus};do + if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then + ident=\$( echo \${filename} | awk -F "ncASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') + name=\$( echo \${filename}| awk -F ".fasta" '{print \$1}') + clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} + clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} + for x in *q.matrix;do + pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') + ya=\$(wc -l \$x | awk '{print \$1}') + echo "\$((\$ya-1))" + tail -"\$((\$ya-1))" \$x > \${pre}z.matrix + rm \$x + cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix + rm \${pre}z.matrix + done + fi + done + if [ `echo ${asvs} | grep -c "_ASV"` -eq 1 ];then + name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') + clustalo -i ${asvs} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} + clustalo -i ${asvs} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} + for x in *q.matrix;do + pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') + ya=\$(wc -l \$x | awk '{print \$1}') + echo "\$((\$ya-1))" + tail -"\$((\$ya-1))" \$x > \${pre}z.matrix + rm \$x + cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix + rm \${pre}z.matrix + done + fi + """ + } + } else { + + process Generate_ASV_Matrix { + + label 'low_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Matrix", mode: "copy", overwrite: true, pattern: '*ASV*PercentID.matrix' + + input: + file(reads) from nuclFastas_forMatrix_ch + + output: + file("*.matrix") into clustmatrices + file("*_ASV*PercentID.matrix") into asv_heatmap + + script: + // remove if statement later (no fin) + """ + for filename in ${reads};do + if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then + ident=\$( echo \${filename} | awk -F "ncASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') + name=\$( echo \${filename}| awk -F ".fasta" '{print \$1}') + clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} + clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} + for x in *q.matrix;do + pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') + ya=\$(wc -l \$x | awk '{print \$1}') + echo "\$((\$ya-1))" + tail -"\$((\$ya-1))" \$x > \${pre}z.matrix + rm \$x + cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix + rm \${pre}z.matrix + done + else + name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') + clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} + clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} + for x in *q.matrix;do + pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') + ya=\$(wc -l \$x | awk '{print \$1}') + echo "\$((\$ya-1))" + tail -"\$((\$ya-1))" \$x > \${pre}z.matrix + rm \$x + cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix + rm \${pre}z.matrix + done + fi + done + """ + } + } + + if (!params.skipPhylogeny) { // need to edit paths + + if (params.ncASV) { + + process Nucleotide_Phylogeny { + + label 'norm_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*ncASV*aln.*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*ncASV*mt*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*ncASV*iq*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*_ASV*aln.*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*_ASV*mt*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*_ASV*iq*' + + input: + tuple file(notus), file(asvs) from nuclFastas_forphylogeny + + output: + tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into align_results + file("*iq.treefile") into nucl_phyl_plot + + script: + """ + for filename in ${notus};do + pre=\$(echo \${filename} | awk -F ".fasta" '{print \$1}' ) + mafft --thread ${task.cpus} --maxiterate 15000 --auto \${filename} >\${pre}_ALN.fasta + trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html + # Nucleotide_ModelTest + modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint + # Nucleotide_Phylogeny + if [ "${params.iqCustomnt}" != "" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -t \${pre}_mt.tree -T auto ${params.iqCustomnt} + elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} + elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni + elif [ "${params.nonparametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} + elif [ "${params.parametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni + else + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni + fi + done + for filename in ${asvs};do + pre=\$(echo \${filename} | awk -F ".fasta" '{print \$1}' ) + mafft --thread ${task.cpus} --maxiterate 15000 --auto \${filename} >\${pre}_ALN.fasta + trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html + # Nucleotide_ModelTest + modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint + # Nucleotide_Phylogeny + if [ "${params.iqCustomnt}" != "" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomnt} + elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} + elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni + elif [ "${params.nonparametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} + elif [ "${params.parametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + else + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + fi + done """ - } - } else if (params.headers == "RVDB") { + } + } else { + + process ASV_Phylogeny { + + label 'norm_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*ASV*aln.*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*ASV*mt*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*ASV*iq*' - process ncASV_Taxonomy_Inference_RVDB { /////// editttt + input: + file(asvs) from nuclFastas_forphylogeny + + output: + tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into align_results + file("*iq.treefile") into nucl_phyl_plot + + script: + """ + for filename in ${asvs};do + pre=\$(echo \${filename} | awk -F ".fasta" '{print \$1}' ) + mafft --thread ${task.cpus} --maxiterate 15000 --auto \${filename} >\${pre}_ALN.fasta + trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html + # Nucleotide_ModelTest + modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint + # Nucleotide_Phylogeny + if [ "${params.iqCustomnt}" != "" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomnt} + elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} + elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni + elif [ "${params.nonparametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} + elif [ "${params.parametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + else + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + fi + done + """ + } + } + } - label 'high_cpus' + if (!params.skipAminoTyping) { - tag "${mtag}" + if (params.sing) { - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Taxonomy", mode: "copy", overwrite: true, pattern: '*ncASV*.{fasta,csv,tsv}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*ncASV*dmd.out' + process Translating_For_Aminotypes { + + label 'low_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes/Translation", mode: "copy", overwrite: true input: - tuple nid, file(asvs) from nuclFastas_forDiamond_ncasv_ch + file(fasta) from asvsforAminotyping output: - file("*.fasta") into tax_labeled_fasta_ncasv - tuple file("*_phyloformat.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond_ncasv - tuple nid, file("*ncASV*summary_for_plot.csv") into taxplot_ncasv - tuple nid, file("*_quick_Taxbreakdown.csv") into tax_table_ncasv + file("${params.projtag}_all_translations.fasta") into amintypegen + file("${params.projtag}_translation_report") into proteinstage_vap_report script: - mtag="ID=" + nid """ - cp ${params.vampdir}/bin/rename_seq.py . - virdb=${params.dbdir}/${params.dbname} - grep ">" \${virdb} > headers.list - headers="headers.list" - name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 - echo "Preparing lists to generate summary .csv's" - echo "[Best hit accession number]" > access.list - echo "[e-value]" > evalue.list - echo "[Bitscore]" > bit.list - echo "[Percent ID (aa)]" > pid.list - echo "[Organism ID]" > "\$name"_virus.list - echo "[Gene]" > "\$name"_genes.list - echo "[ncASV#]" > otu.list - echo "[Sequence length]" > length.list - grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst - if [[ ${params.lca} == "T" ]] - then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list - echo "[Classification]" > classfication.list - else echo "skipped" >> \${name}_quick_Taxbreakdown.csv - echo "[Classification]" > classfication.list - fi - echo "extracting genes and names" - touch new_"\$name"_asvnames.txt - for s in \$(cat seqids.lst);do - echo "Using RVDB headers." - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - if [[ "${params.lca}" == "T" ]] - then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}') - lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') - echo "\$lcla" >> classfication.list - fi - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list - virus="NO" - gene="HIT" - echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - if [[ "${params.lca}" == "T" ]] - then echo "N/A" >> classfication.list - fi - echo "\$s done." - fi - echo "Done with \$s" - done - echo "Now editing "\$name" fasta headers" - ###### rename_seq.py - ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta - awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta - echo "[Sequence header]" > newnames.list - cat new_"\$name"_asvnames.txt >> newnames.list - touch sequence.list - echo " " > sequence.list - grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list - rm "\$name"_tmpssasv.fasta - if [[ "${params.lca}" == "T" ]] - then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list classfication.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv - else paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - fi - for x in *phyloformat.csv;do - echo "\$x" - lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) - tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; - sed 's/ /_/g' tmpcol.list > tmp2col.list; - cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; - rm tmpcol.list tmp2col.list - done - rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list + conda init && source activate virtualribosome + + ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_all_translations.fasta --report ${params.projtag}_translation_report """ + + } + + } else { + + process Translate_For_AminoTyping { + + label 'low_cpus' + + conda 'python=2.7' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes/Translation", mode: "copy", overwrite: true + + input: + file(fasta) from asvsforAminotyping + + output: + file("${params.projtag}_all_translations.fasta") into amintypegen + file("${params.projtag}_translation_report") into proteinstage_vap_report + + script: + """ + ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_all_translations.fasta --report ${params.projtag}_translation_report + """ + } + + } + + process Generate_AminoTypes { + + label 'norm_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{clstr,csv,gc}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes/Problematic", mode: "copy", overwrite: true, pattern: '*problematic*.{fasta}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes", mode: "copy", overwrite: true, pattern: '*AminoTypes_noTaxonomy.{fasta}' + + input: + file(prot) from amintypegen + file(asvs) from asvaminocheck + + output: + tuple file("*.fasta"), file("${params.projtag}_AminoTypes.clstr"), file("${params.projtag}_AminoType_summary_map.csv"), file("${params.projtag}_clustered.gc") into ( supplementalfiles ) + file("${params.projtag}_AminoTypes_noTaxonomy.fasta") into ( aminotypesCounts, aminotypesMafft, aminotypesClustal, aminotypesBlast, aminotypesEmboss ) + + script: + """ + set +e + cp ${params.vampdir}/bin/rename_seq.py . + awk 'BEGIN{RS=">";ORS=""}length(\$2)>="${params.minAA}"{print ">"\$0}' ${prot} >${params.projtag}_filtered_translations.fasta + awk 'BEGIN{RS=">";ORS=""}length(\$2)<"${params.minAA}"{print ">"\$0}' ${prot} >${params.projtag}_problematic_translations.fasta + if [ `wc -l ${params.projtag}_problematic_translations.fasta | awk '{print \$1}'` -gt 1 ];then + grep ">" ${params.projtag}_problematic_translations.fasta | awk -F ">" '{print \$2}' > problem_tmp.list + seqtk subseq ${asvs} problem_tmp.list > ${params.projtag}_problematic_nucleotides.fasta + else + rm ${params.projtag}_problematic_translations.fasta + fi + cd-hit -i ${params.projtag}_filtered_translations.fasta -c 1.0 -o ${params.projtag}_unlabeled_types.fasta + sed 's/>Cluster />Cluster_/g' ${params.projtag}_unlabeled_types.fasta.clstr >${params.projtag}_AminoTypes.clstr + grep ">Cluster_" ${params.projtag}_AminoTypes.clstr >tmpclusters.list + grep -w "*" ${params.projtag}_AminoTypes.clstr | awk '{print \$3}' | awk -F "." '{print \$1}' >tmphead.list + grep -w "*" ${params.projtag}_AminoTypes.clstr | awk '{print \$2}' | awk -F "," '{print \$1}' >tmplen.list + paste -d"," tmpclusters.list tmphead.list >tmp.info.csv + grep ">" ${params.projtag}_unlabeled_types.fasta >lala.list + j=1 + for x in \$(cat lala.list);do + echo ">${params.projtag}_AminoType\${j}" >>${params.projtag}_aminoheaders.list + echo "\${x},>${params.projtag}_AminoType\${j}" >>tmpaminotype.info.csv + j=\$(( \${j}+1 )) + done + rm lala.list + awk -F "," '{print \$2}' tmp.info.csv >>tmporder.list + for x in \$(cat tmporder.list);do + grep -w "\$x" tmpaminotype.info.csv | awk -F "," '{print \$2}' >>tmpder.list + done + paste -d "," tmpclusters.list tmplen.list tmphead.list tmpder.list >${params.projtag}_AminoType_summary_map.csv + rm tmp* + ./rename_seq.py ${params.projtag}_unlabeled_types.fasta ${params.projtag}_aminoheaders.list ${params.projtag}_AminoTypes_noTaxonomy.fasta + stats.sh in=${params.projtag}_AminoTypes_noTaxonomy.fasta gc=${params.projtag}_clustered.gc gcformat=4 + """ + } + + process Generate_AminoType_Matrix { + + label 'low_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Matrix", mode: "copy", overwrite: true + + input: + file(prot) from aminotypesClustal + + output: + file("*.matrix") into proclustmatrices + file("*PercentID.matrix") into aminotype_heatmap + + script: + """ + name=\$( echo ${prot} | awk -F ".fasta" '{print \$1}') + clustalo -i ${prot} --distmat-out=\${name}_PairwiseDistanceq.matrix --full --force --threads=${task.cpus} + clustalo -i ${prot} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} + for x in *q.matrix;do + pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') + ya=\$(wc -l \$x | awk '{print \$1}') + echo "\$((\$ya-1))" + tail -"\$(( \$ya-1))" \$x > \${pre}z.matrix + rm \$x + cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix + rm \${pre}z.matrix + done + """ + } + + if (!params.skipEMBOSS) { + + process AminoType_EMBOSS_Analyses { + + label 'low_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/2dStructure", mode: "copy", overwrite: true, pattern: '*.{garnier}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/HydrophobicMoment", mode: "copy", overwrite: true, pattern: '*HydrophobicMoments.{svg}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/IsoelectricPoint", mode: "copy", overwrite: true, pattern: '*IsoelectricPoint.{iep,svg}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/ProteinProperties", mode: "copy", overwrite: true, pattern: '*.{pepstats,pepinfo}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/ProteinProperties/Plots", mode: "copy", overwrite: true, pattern: '*PropertiesPlot.{svg}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/2dStructure/Plots", mode: "copy", overwrite: true, pattern: '*Helical*.{svg}' + + input: + file(prot) from aminotypesEmboss + + output: + tuple file("*.garnier"), file("*HydrophobicMoments.svg"), file("*IsoelectricPoint*"), file("*.pepstats"), file("*PropertiesPlot*"), file("*Helical*") into amino_emboss + + script: + """ + name=\$( echo ${prot} | awk -F ".fasta" '{print \$1}') + garnier -sequence ${prot} -outfile \${name}_2dStructures.garnier + hmoment -seqall ${prot} -graph svg -plot + mv hmoment.svg ./"\${name}"_HydrophobicMoments.svg + iep -sequence ${prot} -graph svg -plot -outfile "\${name}"_IsoelectricPoint.iep + mv iep.svg ./"\${name}"_IsoelectricPoint.svg + pepstats -sequence ${prot} -outfile \${name}_ProteinProperties.pepstats + grep ">" ${prot} | awk -F ">" '{print \$2}' > tmpsequence.list + for x in \$(cat tmpsequence.list);do + echo \$x > tmp1.list + seqtk subseq ${prot} tmp1.list > tmp2.fasta + len=\$(tail -1 tmp2.fasta | awk '{print length}') + pepinfo -sequence tmp2.fasta -graph svg -outfile "\$x"_PropertiesPlot.pepinfo + mv pepinfo.svg ./"\$x"_PropertiesPlot.svg + cat "\$x"_PropertiesPlot.pepinfo >> "\${name}"_PropertiesPlot.pepinfo + rm "\$x"_PropertiesPlot.pepinfo + pepnet -sask -sequence tmp2.fasta -graph svg -sbegin1 1 -send1 \$len + mv pepnet.svg ./"\$x"_HelicalNet.svg + pepwheel -sequence tmp2.fasta -graph svg -sbegin1 1 -send1 \$len + mv pepwheel.svg ./"\$x"_HelicalWheel.svg + rm tmp1.list tmp2.fasta + done + rm tmpsequence.list + """ } + } + + if (!params.skipTaxonomy) { + + process AminoType_Taxonomy_Inference { + + label 'high_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{csv,tsv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Taxonomy", mode: "copy", overwrite: true, pattern: '*TaxonomyLabels.fasta' + + input: + file(reads) from aminotypesBlast + + output: + tuple file("*_phyloseqObject.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AA_diamond + file("*_summary_for_plot.csv") into taxplot2 + file("*TaxonomyLabels.fasta") into tax_labeled_fasta2 + + script: + """ + cp ${params.vampdir}/bin/rename_seq.py . + virdb=${params.dbdir}/${params.dbname} + grep ">" \${virdb} >> headers.list + headers="headers.list" + name=\$(ls ${reads} | awk -F "_noTaxonomy" '{print \$1}') + diamond blastp -q ${reads} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + echo "Preparing lists to generate summary .csv's" + echo "[Best hit accession number]" >access.list + echo "[pcASV sequence length]" >length.list + echo "[e-value]" >evalue.list + echo "[Bitscore]" >bit.list + echo "[Percent ID (aa)]" >pid.list + echo "[AminoType#]" >otu.list + echo "[Virus ID]" >"\$name"_virus.list + echo "[Gene]" >"\$name"_genes.list + grep ">" ${reads} | awk -F ">" '{print \$2}' > seqids.lst + echo "extracting genes and names" + touch new_"\$name"_asvnames.txt + j=1 + for s in \$(cat seqids.lst);do + echo "Checking for \$s hit in diamond output" + if [[ ${params.refseq} == "T" ]];then + echo "RefSeq headers specified" + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >>evalue.list + echo "\$line" | awk '{print \$11}' >>bit.list + echo "\$line" | awk '{print \$12}' >>pid.list + echo "\$line" | awk '{print \$2}' >>length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">AminoType\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >>access.list + echo "NO_HIT" >>"\$name"_genes.list + echo "NO_HIT" >>"\$name"_virus.list + echo "NO_HIT" >>evalue.list + echo "NO_HIT" >>bit.list + echo "NO_HIT" >>pid.list + echo "NO_HIT" >>length.list + virus="NO" + gene="HIT" + echo ">AminoType\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + fi + else + echo "Using RVDB headers." + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') + echo "\$s" >>otu.list + echo "\$acc" >>access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >>evalue.list + echo "\$line" | awk '{print \$11}' >>bit.list + echo "\$line" | awk '{print \$12}' >>pid.list + echo "\$line" | awk '{print \$2}' >>length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') + echo "\$gene" | sed 's/_/ /g' >>"\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') + echo "\$virus" | sed 's/_/ /g' >>"\$name"_virus.list + echo ">AminoType\${j}_"\$virus"_"\$gene"" >>new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >>otu.list + echo "NO_HIT" >>access.list + echo "NO_HIT" >>"\$name"_genes.list + echo "NO_HIT" >>"\$name"_virus.list + echo "NO_HIT" >>evalue.list + echo "NO_HIT" >>bit.list + echo "NO_HIT" >>pid.list + echo "NO_HIT" >>length.list + virus="NO" + gene="HIT" + echo ">AminoType\${j}_"\$virus"_"\$gene"" >>new_"\$name"_asvnames.txt + j=\$((\$j+1)) + echo "\$s done." + fi + fi + echo "Done with \$s" + done + echo "Now editing "\$name" fasta headers" + ###### rename_seq.py + ./rename_seq.py ${reads} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta > "\$name"_tmpssasv.fasta + echo "[Sequence header]" > newnames.list + cat new_"\$name"_asvnames.txt >> newnames.list + touch sequence.list + echo " " > sequence.list + grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list + rm "\$name"_tmpssasv.fasta + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + for x in *phyloseqObject.csv;do + echo "\$x" + lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) + tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; + sed 's/ /_/g' tmpcol.list > tmp2col.list; + cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; + rm tmpcol.list tmp2col.list + done + rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst headers.list otu.list ; + rm *asvnames.txt + rm "\$name"_virus.list + rm "\$name"_genes.list + rm newnames.list + rm access.list + echo "Taxonomy inferred for: ${reads} " + """ } - } + } + if (!params.skipPhylogeny) { - process Generate_ncASV_Counts_Table { + process AminoType_Phylogeny { label 'norm_cpus' - tag "${mtag}" - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Counts", mode: "copy", overwrite: true, pattern: '*_ASV*.{biome,csv}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Counts", mode: "copy", overwrite: true, pattern: '*ncASV*.{biome,csv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*aln.*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Phylogeny/Modeltest", mode: "copy", overwrite: true, pattern: '*mt*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*iq*' input: - tuple nid, file(notus) from nuclFastas_forCounts_ncasv_ch - file(merged) from nuclCounts_mergedreads_ncasv_ch + file(prot) from aminotypesMafft output: - tuple file("*_counts.csv"), file("*_counts.biome") into counts_vsearch_ncasv - tuple nid, file("*ncASV*counts.csv") into notu_counts_plots + tuple file("*_aln.fasta"), file("*_aln.html"), file("*.log"), file("*iq*"), file("*mt*") into alignprot_results + file("*iq.treefile") into amino_rax_plot + + script: + """ + # Protein_Alignment + pre=\$(echo ${prot} | awk -F ".fasta" '{print \$1}' ) + mafft --thread ${task.cpus} --maxiterate 15000 --auto ${prot} >\${pre}_ALN.fasta + trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html + + # Protein_ModelTest + modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d aa -s 203 --disable-checkpoint + + # Protein_Phylogeny + if [ "${params.iqCustomaa}" != "" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomaa} + + elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} + + elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni + + elif [ "${params.nonparametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} + + elif [ "${params.parametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + + else + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + fi + """ + } + } + + process Generate_AminoTypes_Counts_Table { + + label 'high_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Counts", mode: "copy", overwrite: true + + input: + file(fasta) from aminotypesCounts + file(merged) from mergeforprotcounts + file(samplist) from samplelist + + output: + tuple file("*_protcounts.csv"), file("*dmd.out") into counts_summary + file("*_protcounts.csv") into aminocounts_plot + + script: + """ + set +e + diamond makedb --in ${fasta} --db ${fasta} + diamond blastx -q ${merged} -d ${fasta} -p ${task.cpus} --min-score ${params.ProtCountsBit} --id ${params.ProtCountID} -l ${params.ProtCountsLength} --more-sensitive -o ${params.projtag}_protCounts_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 --max-hsps 1 + echo "OTU_ID" >tmp.col1.txt + echo "Generating sample id list" + grep ">" ${fasta} | awk -F ">" '{print \$2}' | sort | uniq > otuid.list + cat otuid.list >> tmp.col1.txt + echo "Beginning them counts tho my g" + for y in \$( cat ${samplist} );do + echo "Starting with \$y now ..." + grep "\$y" ${params.projtag}_protCounts_dmd.out > tmp."\$y".out + echo "Isolated hits" + echo "Created uniq subject id list" + echo "\$y" > "\$y"_col.txt + echo "Starting my counts" + for z in \$(cat otuid.list);do + echo "Counting \$z hits" + echo "grep -wc "\$z" >> "\$y"_col.txt" + grep -wc "\$z" tmp."\$y".out >> "\$y"_col.txt + echo "\$z counted" + done + done + paste -d "," tmp.col1.txt *col.txt > ${params.projtag}_protcounts.csv + rm tmp* + rm *col.txt + """ + } + } + + if (params.pcASV) { // ASV_nucl -> ASV_aa -> clusteraa by %id with ch-hit -> extract representative nucl sequences to generate new OTU file + + if (params.sing) { + + process Translating_For_pcASV_Generation { + + label 'low_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV/Translation", mode: "copy", overwrite: true, pattern: '*_ASV_translations*' + + input: + file(fasta) from nucl2aa + + output: + file("*ASV*translations.fasta") into clustering_aa + file("*_ASV_translations_report") into reportaa_VR + file("*_ASV_nucleotide.fasta") into asvfastaforaaclust + + script: + """ + conda init && source activate virtualribosome + + ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_ASV_translations.fasta --report ${params.projtag}_ASV_translations_report + cp ${fasta} ${params.projtag}_ASV_nucleotide.fasta + """ + + } + + } else { + + process Translation_For_pcASV_Generation { + + label 'low_cpus' + + conda 'python=2.7' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV/Translation", mode: "copy", overwrite: true, pattern: '*_ASV_translations*' + + input: + file(fasta) from nucl2aa + + output: + file("*ASV*translations.fasta") into clustering_aa + file("*_ASV_translations_report") into reportaa_VR + file("*_ASV_nucleotide.fasta") into asvfastaforaaclust + + script: + """ + ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_ASV_translations.fasta --report ${params.projtag}_ASV_translations_report + cp ${fasta} ${params.projtag}_ASV_nucleotide.fasta + """ + } + } + + process Generate_pcASVs { + + label 'norm_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV", mode: "copy", overwrite: true, pattern: '*pcASV*.{fasta}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{clstr,csv,gc}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV/Problematic", mode: "copy", overwrite: true, pattern: '*problem*.{fasta}' + + input: + file(fasta) from clustering_aa + file(asvs) from asvfastaforaaclust + + output: + file("${params.projtag}_nucleotide_pcASV*.fasta") into ( pcASV_ntDiamond_ch, pcASV_nt_counts_ch, pcASV_ntmatrix_ch, pcASV_ntmafft_ch ) + file("*_aminoacid_pcASV*_noTaxonomy.fasta") into ( pcASV_aaMatrix_ch, pcASV_aaDiamond_ch, pcASV_aaMafft_ch, pcASV_aaCounts_ch, pcASVEMBOSS ) + tuple file("*.fasta"), file("*.clstr"), file("*.csv"), file("*.gc") into ( pcASVsupplementalfiles ) + + script: + // add awk script to count seqs + if (params.clusterAAIDlist) { + """ + set +e + cp ${params.vampdir}/bin/rename_seq.py . + for id in `echo ${params.clusterAAIDlist} | tr "," "\\n"`;do + awk 'BEGIN{RS=">";ORS=""}length(\$2)>="${params.minAA}"{print ">"\$0}' ${fasta} > ${params.projtag}_filtered_proteins.fasta + cd-hit -i ${params.projtag}_filtered_proteins.fasta -c \${id} -o ${params.projtag}_pcASV\${id}.fasta + sed 's/>Cluster />Cluster_/g' ${params.projtag}_pcASV\${id}.fasta.clstr >${params.projtag}_pcASV\${id}.clstr + grep ">Cluster_" ${params.projtag}_pcASV\${id}.clstr >temporaryclusters.list + y=\$(grep -c ">Cluster_" ${params.projtag}_pcASV\${id}.clstr) + echo ">Cluster_"\${y}"" >> ${params.projtag}_pcASV\${id}.clstr + t=1 + b=1 + for x in \$(cat temporaryclusters.list);do + echo "Extracting \$x" + name="\$( echo \$x | awk -F ">" '{print \$2}')" + clust="pcASV"\${t}"" + echo "\${name}" + awk '/^>'\${name}'\$/,/^>Cluster_'\${b}'\$/' ${params.projtag}_pcASV\${id}.clstr > "\${name}"_"\${clust}"_tmp.list + t=\$(( \${t}+1 )) + b=\$(( \${b}+1 )) + done + ls *_tmp.list + u=1 + for x in *_tmp.list;do + name="\$(echo \$x | awk -F "_p" '{print \$1}')" + echo "\${name}" + cluster="\$(echo \$x | awk -F "_" '{print \$3}')" + echo "\${cluster}" + grep "ASV" \$x | awk -F ", " '{print \$2}' | awk -F "_" '{print \$1}' | awk -F ">" '{print \$2}' > \${name}_\${cluster}_seqs_tmps.list + seqtk subseq ${asvs} \${name}_\${cluster}_seqs_tmps.list > \${name}_\${cluster}_nucleotide_sequences.fasta + vsearch --cluster_fast \${name}_\${cluster}_nucleotide_sequences.fasta --id 0.2 --centroids \${name}_\${cluster}_centroids.fasta + grep ">" \${name}_\${cluster}_centroids.fasta >> \${name}_\${cluster}_tmp_centroids.list + for y in \$( cat \${name}_\${cluster}_tmp_centroids.list );do + echo ">\${cluster}_type"\$u"" >> \${name}_\${cluster}_tmp_centroid.newheaders + u=\$(( \${u}+1 )) + done + u=1 + ./rename_seq.py \${name}_\${cluster}_centroids.fasta \${name}_\${cluster}_tmp_centroid.newheaders \${cluster}_types_labeled.fasta + done + cat *_types_labeled.fasta >> ${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta + grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$3}' | awk -F "." '{print \$1}' >tmphead.list + grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$2}' | awk -F "," '{print \$1}' >tmplen.list + paste -d"," temporaryclusters.list tmphead.list >tmp.info.csv + grep ">" ${params.projtag}_pcASV\${id}.fasta >lala.list + j=1 + for x in \$(cat lala.list);do + echo ">${params.projtag}_pcASV\${j}" >>${params.projtag}_aminoheaders.list + echo "\${x},>${params.projtag}_pcASV\${j}" >>tmpaminotype.info.csv + j=\$(( \${j}+1 )) + done + rm lala.list + awk -F "," '{print \$2}' tmp.info.csv >>tmporder.list + for x in \$(cat tmporder.list);do + grep -w "\$x" tmpaminotype.info.csv | awk -F "," '{print \$2}' >>tmpder.list + done + paste -d "," temporaryclusters.list tmplen.list tmphead.list tmpder.list >${params.projtag}_pcASVCluster\${id}_summary.csv + ./rename_seq.py ${params.projtag}_pcASV\${id}.fasta ${params.projtag}_aminoheaders.list ${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta + stats.sh in=${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_aminoacid_clustered.gc gcformat=4 overwrite=true + stats.sh in=${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_nucleotide_clustered.gc gcformat=4 overwrite=true + awk 'BEGIN{RS=">";ORS=""}length(\$2)<"${params.minAA}"{print ">"\$0}' ${fasta} >${params.projtag}_pcASV\${id}_problematic_translations.fasta + if [ `wc -l ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk '{print \$1}'` -gt 1 ];then + grep ">" ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk -F ">" '{print \$2}' > problem_tmp.list + seqtk subseq ${asvs} > ${params.projtag}_pcASV\${id}_problematic_nucleotides.fasta + else + rm ${params.projtag}_pcASV\${id}_problematic_translations.fasta + fi + rm *.list + rm Cluster* + rm *types* + rm *tmp* + rm ${params.projtag}_pcASV\${id}.fast* + done + """ + } else if (params.clusterAAID) { + """ + set +e + cp ${params.vampdir}/bin/rename_seq.py . + id=${params.clusterAAID} + awk 'BEGIN{RS=">";ORS=""}length(\$2)>="${params.minAA}"{print ">"\$0}' ${fasta} > ${params.projtag}_filtered_proteins.fasta + cd-hit -i ${params.projtag}_filtered_proteins.fasta -c ${params.clusterAAID} -o ${params.projtag}_pcASV\${id}.fasta + sed 's/>Cluster />Cluster_/g' ${params.projtag}_pcASV\${id}.fasta.clstr >${params.projtag}_pcASV\${id}.clstr + grep ">Cluster_" ${params.projtag}_pcASV\${id}.clstr >temporaryclusters.list + y=\$(grep -c ">Cluster_" ${params.projtag}_pcASV\${id}.clstr) + echo ">Cluster_"\${y}"" >> ${params.projtag}_pcASV\${id}.clstr + t=1 + b=1 + for x in \$(cat temporaryclusters.list);do + echo "Extracting \$x" + name="\$( echo \$x | awk -F ">" '{print \$2}')" + clust="pcASV"\${t}"" + echo "\${name}" + awk '/^>'\${name}'\$/,/^>Cluster_'\${b}'\$/' ${params.projtag}_pcASV\${id}.clstr > "\${name}"_"\${clust}"_tmp.list + t=\$(( \${t}+1 )) + b=\$(( \${b}+1 )) + done - script: - mtag="ID=" + nid - """ - name=\$( echo ${notus} | awk -F ".fasta" '{print \$1}') - vsearch --usearch_global ${merged} --db ${notus} --id .${nid} --threads ${task.cpus} --otutabout \${name}_counts.txt --biomout \${name}_counts.biome - cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv - sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv - rm \${name}_count.csv - """ + ls *_tmp.list + u=1 + for x in *_tmp.list;do + name="\$(echo \$x | awk -F "_p" '{print \$1}')" + echo "\${name}" + cluster="\$(echo \$x | awk -F "_" '{print \$3}')" + echo "\${cluster}" + grep "ASV" \$x | awk -F ", " '{print \$2}' | awk -F "_" '{print \$1}' | awk -F ">" '{print \$2}' > \${name}_\${cluster}_seqs_tmps.list + seqtk subseq ${asvs} \${name}_\${cluster}_seqs_tmps.list > \${name}_\${cluster}_nucleotide_sequences.fasta + vsearch --cluster_fast \${name}_\${cluster}_nucleotide_sequences.fasta --id 0.2 --centroids \${name}_\${cluster}_centroids.fasta + grep ">" \${name}_\${cluster}_centroids.fasta >> \${name}_\${cluster}_tmp_centroids.list + for y in \$( cat \${name}_\${cluster}_tmp_centroids.list );do + echo ">\${cluster}_type"\$u"" >> \${name}_\${cluster}_tmp_centroid.newheaders + u=\$(( \${u}+1 )) + done + u=1 + ./rename_seq.py \${name}_\${cluster}_centroids.fasta \${name}_\${cluster}_tmp_centroid.newheaders \${cluster}_types_labeled.fasta + done + cat *_types_labeled.fasta >> ${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta + grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$3}' | awk -F "." '{print \$1}' >tmphead.list + grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$2}' | awk -F "," '{print \$1}' >tmplen.list + paste -d"," temporaryclusters.list tmphead.list >tmp.info.csv + grep ">" ${params.projtag}_pcASV\${id}.fasta >lala.list + j=1 + for x in \$(cat lala.list);do + echo ">${params.projtag}_pcASV\${j}" >>${params.projtag}_aminoheaders.list + echo "\${x},>${params.projtag}_pcASV\${j}" >>tmpaminotype.info.csv + j=\$(( \${j}+1 )) + done + rm lala.list + awk -F "," '{print \$2}' tmp.info.csv >>tmporder.list + for x in \$(cat tmporder.list);do + grep -w "\$x" tmpaminotype.info.csv | awk -F "," '{print \$2}' >>tmpder.list + done + paste -d "," temporaryclusters.list tmplen.list tmphead.list tmpder.list >${params.projtag}_pcASVCluster\${id}_summary.csv + ./rename_seq.py ${params.projtag}_pcASV\${id}.fasta ${params.projtag}_aminoheaders.list ${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta + stats.sh in=${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_aminoacid_clustered.gc gcformat=4 + stats.sh in=${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_nucleotide_clustered.gc gcformat=4 + awk 'BEGIN{RS=">";ORS=""}length(\$2)<"${params.minAA}"{print ">"\$0}' ${fasta} >${params.projtag}_pcASV\${id}_problematic_translations.fasta + if [ `wc -l ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk '{print \$1}'` -gt 1 ];then + grep ">" ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk -F ">" '{print \$2}' > problem_tmp.list + seqtk subseq ${asvs} problem_tmp.list > ${params.projtag}_pcASV\${id}_problematic_nucleotides.fasta + else + rm ${params.projtag}_pcASV\${id}_problematic_translations.fasta + fi + rm *.list + rm Cluster* + rm *types* + rm *tmp* + rm ${params.projtag}_pcASV\${id}.fast* + """ } + } - process Generate_ncASV_Matrices { - label 'low_cpus' + if (!params.skipTaxonomy) { - tag "${mtag}" + process pcASV_Nucleotide_Taxonomy_Inference { - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Matrices", mode: "copy", overwrite: true, pattern: '*ncASV*PercentID.matrix' + label 'high_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{csv,tsv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*dmd.{out}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{fasta}' input: - tuple nid, file(asvs) from nuclFastas_forMatrix_ncasv_ch + file(reads) from pcASV_ntDiamond_ch output: - file("*.matrix") into clustmatrices_ncasv - tuple nid, file("*ncASV*PercentID.matrix") into notu_heatmap + file("*.fasta") into ( pcASV_labeled ) + tuple file("*_phyloseqObject.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AAdiamond + file("*_summary_for_plot.csv") into taxplot3 script: - mtag="ID=" + nid - """ - name=\$( echo ${asvs}| awk -F ".fasta" '{print \$1}') - clustalo -i ${asvs} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} - clustalo -i ${asvs} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} - cat \${name}_PercentIDq.matrix | tr " " "," | grep "," >\${name}_PercentID.matrix - rm \${name}_PercentIDq.matrix """ - } - - if (!params.skipPhylogeny) { - - process ncASV_Phylogeny { - - label 'norm_cpus' - - tag "${mtag}" - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*ncASV*aln.*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*ncASV*mt*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*ncASV*iq*' - - input: - tuple nid, file(asvs) from nuclFastas_forphylogeny_ncasv - - output: - tuple nid, file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into align_results_ncasv - tuple nid, file("*iq.treefile") into nucl_phyl_plot_ncasv - - script: - mtag="ID=" + nid - """ - pre=\$(echo ${asvs} | awk -F ".fasta" '{print \$1}' ) - mafft --thread ${task.cpus} --maxiterate 15000 --auto ${asvs} >\${pre}_ALN.fasta - trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html - o-trim-uninformative-columns-from-alignment \${pre}_aln.fasta - mv \${pre}_aln.fasta-TRIMMED ./\${pre}_Aligned_informativeonly.fasta - # Nucleotide_ModelTest - modeltest-ng -i \${pre}_Aligned_informativeonly.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint - # Nucleotide_Phylogeny - if [ "${params.iqCustomnt}" != "" ];then - iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq --redo -t \${pre}_mt.tree -T auto ${params.iqCustomnt} - elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} - elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni - elif [ "${params.nonparametric}" != "false" ];then - iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} - elif [ "${params.parametric}" != "false" ];then - iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni - else - iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni - fi - """ - } - - } - - } else { - reads_vsearch5_ch - .into{ nuclFastas_forDiamond_asv_ch; nuclFastas_forCounts_asv_ch; nuclFastas_forphylogeny_asv; nuclFastas_forMatrix_asv_ch; asv_for_med } - } - - if (!params.skipTaxonomy) { - - if (params.headers == "RefSeq") { - - process ASV_Taxonomy_Inference_RefSeq { /////// editttt - - label 'high_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy", mode: "copy", overwrite: true, pattern: '*_ASV*.{fasta,csv,tsv}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*_ASV*dmd.out' - - input: - file(asvs) from nuclFastas_forDiamond_asv_ch - - output: - file("*.fasta") into tax_labeled_fasta_asv - tuple file("*_phyloformat.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond_asv - file("*_ASV*_summary_for_plot.csv") into taxplot_asv - file("*_quick_Taxbreakdown.csv") into tax_table_asv - - script: - """ - cp ${params.vampdir}/bin/rename_seq.py . - virdb=${params.dbdir}/${params.dbname} - grep ">" \${virdb} > headers.list - headers="headers.list" - name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + set +e + cp ${params.vampdir}/bin/rename_seq.py . + virdb=${params.dbdir}/${params.dbname} + grep ">" \${virdb} >> headers.list + headers="headers.list" + for filename in ${reads};do + name=\$(ls \${filename} | awk -F "_noTax" '{print \$1}') + diamond blastx -q \${filename} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 echo "Preparing lists to generate summary .csv's" - echo "[Best hit accession number]" > access.list - echo "[e-value]" > evalue.list - echo "[Bitscore]" > bit.list - echo "[Percent ID (aa)]" > pid.list - echo "[Organism ID]" > "\$name"_virus.list - echo "[Gene]" > "\$name"_genes.list - echo "[ASV#]" > otu.list - echo "[Sequence length]" > length.list - grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst + echo "[Best hit accession number]" >access.list + echo "[pcASV sequence length]" >length.list + echo "[e-value]" >evalue.list + echo "[Bitscore]" >bit.list + echo "[Percent ID (aa)]" >pid.list + echo "[pcASV#]" >otu.list + echo "[Virus ID]" >"\$name"_virus.list + echo "[Gene]" >"\$name"_genes.list + grep ">" \${filename} | awk -F ">" '{print \$2}' > seqids.lst echo "extracting genes and names" - touch new_"\$name"_asvnames.txt + touch new_"\$name"_headers.txt + j=1 for s in \$(cat seqids.lst);do echo "Checking for \$s hit in diamond output" - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + if [[ ${params.refseq} == "T" ]];then + echo "RefSeq headers specified" + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then echo "Yep, there was a hit for \$s" echo "Extracting the information now:" acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') echo "\$s" >> otu.list echo "\$acc" >> access.list line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list + echo "\$line" | awk '{print \$10}' >>evalue.list + echo "\$line" | awk '{print \$11}' >>bit.list + echo "\$line" | awk '{print \$12}' >>pid.list + echo "\$line" | awk '{print \$2}' >>length.list echo "Extracting virus and gene ID for \$s now" gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">"\${s}"_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + echo ">pcASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_headers.txt + j=\$((\$j+1)) echo "\$s done." - else + else echo "Ugh, there was no hit for \$s .." echo "We still love \$s though and we will add it to the final fasta file" echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list + echo "NO_HIT" >>access.list + echo "NO_HIT" >>"\$name"_genes.list + echo "NO_HIT" >>"\$name"_virus.list + echo "NO_HIT" >>evalue.list + echo "NO_HIT" >>bit.list + echo "NO_HIT" >>pid.list + echo "NO_HIT" >>length.list + virus="NO" + gene="HIT" + echo ">pcASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_headers.txt + j=\$((\$j+1)) + echo "\$s done." + fi + else + echo "Using RVDB headers." + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') + echo "\$s" >>otu.list + echo "\$acc" >>access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >>evalue.list + echo "\$line" | awk '{print \$11}' >>bit.list + echo "\$line" | awk '{print \$12}' >>pid.list + echo "\$line" | awk '{print \$2}' >>length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >>"\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && + echo "\$virus" | sed 's/_/ /g' >>"\$name"_virus.list + echo ">pcASV\${j}_"\$virus"_"\$gene"" >>new_"\$name"_headers.txt + j=\$((\$j+1)) + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >>otu.list + echo "NO_HIT" >>access.list + echo "NO_HIT" >>"\$name"_genes.list + echo "NO_HIT" >>"\$name"_virus.list + echo "NO_HIT" >>evalue.list + echo "NO_HIT" >>bit.list + echo "NO_HIT" >>pid.list + echo "NO_HIT" >>length.list virus="NO" gene="HIT" - echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + echo ">pcASV\${j}_"\$virus"_"\$gene"" >>new_"\$name"_headers.txt + j=\$((\$j+1)) echo "\$s done." - fi + fi + fi + echo "Done with \$s" done echo "Now editing "\$name" fasta headers" ###### rename_seq.py - ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta - awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta + ./rename_seq.py \${filename} new_"\$name"_headers.txt "\$name"_TaxonomyLabels.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta > "\$name"_tmpssasv.fasta echo "[Sequence header]" > newnames.list - cat new_"\$name"_asvnames.txt >> newnames.list + cat new_"\$name"_headers.txt >> newnames.list touch sequence.list echo " " > sequence.list grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - for x in *phyloformat.csv;do + for x in *phyloseqObject.csv;do echo "\$x" lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; @@ -1773,2020 +2458,1307 @@ if (params.DataCheck || params.Analyze) { cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; rm tmpcol.list tmp2col.list done - rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list - """ - } - } else if (params.headers == "RVDB") { - - process ASV_Taxonomy_Inference_RVDB { /////// editttt - - label 'high_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy", mode: "copy", overwrite: true, pattern: '*_ASV*.{fasta,csv,tsv}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*_ASV*dmd.out' - - input: - file(asvs) from nuclFastas_forDiamond_asv_ch - - output: - file("*.fasta") into tax_labeled_fasta_asv - tuple file("*_phyloformat.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond_asv - file("*_ASV*_summary_for_plot.csv") into taxplot_asv - file("*_quick_Taxbreakdown.csv") into tax_table_asv - - script: - """ - cp ${params.vampdir}/bin/rename_seq.py . - virdb=${params.dbdir}/${params.dbname} - grep ">" \${virdb} > headers.list - headers="headers.list" - name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 - echo "Preparing lists to generate summary .csv's" - echo "[Best hit accession number]" > access.list - echo "[e-value]" > evalue.list - echo "[Bitscore]" > bit.list - echo "[Percent ID (aa)]" > pid.list - echo "[Organism ID]" > "\$name"_virus.list - echo "[Gene]" > "\$name"_genes.list - echo "[ASV#]" > otu.list - echo "[Sequence length]" > length.list - grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst - if [[ ${params.lca} == "T" ]] - then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list - echo "[Classification]" > classfication.list - else echo "skipped" >> \${name}_quick_Taxbreakdown.csv - echo "[Classification]" > classfication.list - fi - echo "extracting genes and names" - touch new_"\$name"_asvnames.txt - for s in \$(cat seqids.lst);do - echo "Using RVDB headers." - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - if [[ "${params.lca}" == "T" ]] - then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}') - lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') - echo "\$lcla" >> classfication.list - fi - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list - virus="NO" - gene="HIT" - echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - if [[ "${params.lca}" == "T" ]] - then echo "N/A" >> classfication.list - fi - echo "\$s done." - fi - echo "Done with \$s" - done - echo "Now editing "\$name" fasta headers" - ###### rename_seq.py - ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta - awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta - echo "[Sequence header]" > newnames.list - cat new_"\$name"_asvnames.txt >> newnames.list - touch sequence.list - echo " " > sequence.list - grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list - rm "\$name"_tmpssasv.fasta - if [[ "${params.lca}" == "T" ]] - then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list classfication.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv - else paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - fi - for x in *phyloformat.csv;do - echo "\$x" - lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) - tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; - sed 's/ /_/g' tmpcol.list > tmp2col.list; - cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; - rm tmpcol.list tmp2col.list - done - rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list - """ - } + rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; + rm "\$name"_virus.list + rm "\$name"_genes.list + rm newnames.list + rm access.list + echo "Taxonomy inferred for: \${filename} " + done + rm *headers.list + """ } } - process Generate_ASV_Counts_Tables { + process Generate_Nucleotide_pcASV_Counts { label 'norm_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Counts", mode: "copy", overwrite: true, pattern: '*ASV*.{biome,csv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Counts", mode: "copy", overwrite: true, pattern: '*.{biome,csv,txt}' input: - file(asvs) from nuclFastas_forCounts_asv_ch - file(merged) from nuclCounts_mergedreads_asv_ch + file(potus) from pcASV_nt_counts_ch + file(merged) from pcASV_mergedreads_ch output: - tuple file("*_counts.csv"), file("*_counts.biome") into counts_vsearch_asv - file("*_ASV*counts.csv") into (asv_counts_plots, asvcount_med) + tuple file("*_counts.txt"), file("*_counts.biome") into pcASVcounts_vsearch + file("*.csv") into potu_Ncounts_for_report script: """ - name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - vsearch --usearch_global ${merged} --db ${asvs} --id .${params.asvcountID} --threads ${task.cpus} --otutabout "\$name"_counts.txt --biomout "\$name"_counts.biome - cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv - sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv - rm \${name}_count.csv + for filename in ${potus};do + ident=\$( echo \${filename} | awk -F "pcASV" '{print \$2}' | awk -F "_noTaxonomy.fasta" '{print \$1}') + name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') + vsearch --usearch_global ${merged} --db \${filename} --id \${ident} --threads ${task.cpus} --otutabout \${name}_counts.txt --biomout \${name}_counts.biome + cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv + sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv + rm \${name}_count.csv + done """ } - process Generate_ASV_Matrices { + process Generate_pcASV_Nucleotide_Matrix { label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Matrices", mode: "copy", overwrite: true, pattern: '*ASV*PercentID.matrix' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Matrix", mode: "copy", overwrite: true input: - file(reads) from nuclFastas_forMatrix_asv_ch + file(potus) from pcASV_ntmatrix_ch output: - file("*.matrix") into clustmatrices_asv - file("*_ASV*PercentID.matrix") into asv_heatmap - - script: - // remove if statement later (no fin) - """ - for filename in ${reads};do - if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then - ident=\$( echo \${filename} | awk -F "ncASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') - name=\$( echo \${filename}| awk -F ".fasta" '{print \$1}') - clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} - clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} - for x in *q.matrix;do - pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') - ya=\$(wc -l \$x | awk '{print \$1}') - echo "\$((\$ya-1))" - tail -"\$((\$ya-1))" \$x > \${pre}z.matrix - rm \$x - cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix - rm \${pre}z.matrix - done - else - name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') - clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} - clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} - for x in *q.matrix;do - pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') - ya=\$(wc -l \$x | awk '{print \$1}') - echo "\$((\$ya-1))" - tail -"\$((\$ya-1))" \$x > \${pre}z.matrix - rm \$x - cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix - rm \${pre}z.matrix - done - fi - done - """ - } - - if (!params.skipPhylogeny) { // need to edit paths - - process ASV_Phylogeny { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*ASV*aln.*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*ASV*mt*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*ASV*iq*' - - input: - file(asvs) from nuclFastas_forphylogeny_asv - - output: - tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into align_results_asv - file("*iq.treefile") into (nucl_phyl_plot_asv, asvphy_med) - - script: - """ - pre=\$(echo ${asvs} | awk -F ".fasta" '{print \$1}' ) - mafft --thread ${task.cpus} --maxiterate 15000 --auto ${asvs} >\${pre}_ALN.fasta - trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html - o-trim-uninformative-columns-from-alignment \${pre}_aln.fasta - mv \${pre}_aln.fasta-TRIMMED ./\${pre}_Aligned_informativeonly.fasta - # Nucleotide_ModelTest - modeltest-ng -i \${pre}_Aligned_informativeonly.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint - # Nucleotide_Phylogeny - if [ "${params.iqCustomnt}" != "" ];then - iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomnt} - elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} - elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni - elif [ "${params.nonparametric}" != "false" ];then - iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} - elif [ "${params.parametric}" != "false" ];then - iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - else - iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - fi - """ - } - } - - if (params.asvMED) { - - process ASV_Minimum_Entropy_Decomposition { - - label 'low_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/ASVs/MED", mode: "copy", overwrite: true - - input: - file(asvs) from asv_for_med - - output: - file("*_ASV_Grouping.csv") into asvgroupscsv - file("${params.projtag}_ASV_group_reps_aligned.fasta") into groupreps - file("${params.projtag}_asvMED_${params.asvC}") - - script: - """ - #alignment - mafft --thread ${task.cpus} --maxiterate 15000 --auto ${asvs} > ${params.projtag}_ASVs_mafftAlign.fasta - #trimming - trimal -in ${params.projtag}_ASVs_mafftAlign.fasta -out ${params.projtag}_ASVs_mafftAligned.fasta -keepheader -fasta -automated1 - rm ${params.projtag}_ASVs_mafftAlign.fasta - o-trim-uninformative-columns-from-alignment ${params.projtag}_ASVs_mafftAligned.fasta - mv ${params.projtag}_ASVs_mafftAligned.fasta-TRIMMED ./${params.projtag}_ASVs_Aligned_informativeonly.fasta - #entopy analysis - entropy-analysis ${params.projtag}_ASVs_Aligned_informativeonly.fasta - #Decomposition - oligotype ${params.projtag}_ASVs_Aligned_informativeonly.fasta ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY -o ${params.projtag}_asvMED_${params.asvC} -M 1 -c ${params.asvC} -N ${task.cpus} --skip-check-input --no-figures --skip-gen-html - #generatemaps - cd ./${params.projtag}_asvMED_${params.asvC}/OLIGO-REPRESENTATIVES/ - echo "ASV,GroupID,IDPattern" - j=1 - for x in *_unique; - do gid=\$(echo \$x | awk -F "_" '{print \$1}') - uni=\$(echo \$x | awk -F ""\${gid}"_" '{print \$2}' | awk -F "_uni" '{print \$1}') - grep ">" "\$gid"_"\$uni" | awk -F ">" '{print \$2}' > asv.list - seqtk subseq ../../${asvs} asv.list > Group"\${j}"_sequences.fasta - for z in \$( cat asv.list) - do echo ""\$z",Group"\$j","\$uni"" >> ${params.projtag}_ASV_Grouping.csv - - done - rm asv.list - echo ">Group\${j}" >> ${params.projtag}_ASV_group_reps_aligned.fasta - echo "\$uni" > group.list - seqtk subseq ../OLIGO-REPRESENTATIVES.fasta group.list > group.fasta - tail -1 group.fasta >> ${params.projtag}_ASV_group_reps_aligned.fasta - mv "\$gid"_"\$uni" ./Group"\$j"_"\$uni"_aligned.fasta - mv "\$gid"_"\$uni"_unique ./Group"\$j"_"\$uni"_unqiues_aligned.fasta - rm "\$gid"*.cPickle - j=\$((\$j+1)) - done - mv ${params.projtag}_ASV_Grouping.csv ../../ - mv ${params.projtag}_ASV_group_reps_aligned.fasta ../../ - cd .. - """ - } - - process ASV_MED_Reps_phylogeny { - - label 'low_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/MED/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*ASV*mt*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/MED/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*ASV*iq*' - - input: - file(reps) from groupreps - - output: - file("*_ASV_Group_Reps*") into align_results_asvmed - file("*iq.treefile") into asv_group_rep_tree - - script: - """ - # Protein_ModelTest - modeltest-ng -i ${reps} -p ${task.cpus} -o ${params.projtag}_ASV_Group_Reps_mt -d aa -s 203 --disable-checkpoint + file("*.matrix") into pcASVclustmatrices + file("*PercentID.matrix") into potu_nucl_heatmap - # Protein_Phylogeny - if [ "${params.iqCustomaa}" != "" ];then - iqtree -s ${reps} --prefix ${params.projtag}_ASV_Group_Reps_iq --redo -T auto ${params.iqCustomaa} - - elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') - iqtree -s ${reps} --prefix ${params.projtag}_ASV_Group_Reps_iq -m \${mod} --redo -nt auto -b ${params.boots} - - elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') - iqtree -s ${reps} --prefix ${params.projtag}_ASV_Group_Reps_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni - - elif [ "${params.nonparametric}" != "false" ];then - iqtree -s ${reps} --prefix ${params.projtag}_ASV_Group_Reps_iq -m MFP --redo -nt auto -b ${params.boots} - - elif [ "${params.parametric}" != "false" ];then - iqtree -s ${reps} --prefix ${params.projtag}_ASV_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - - else - iqtree -s ${reps} --prefix ${params.projtag}_ASV_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - fi - """ - } - - process Adding_ASV_MED_Info { - - label 'low_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/MED/", mode: "copy", overwrite: true - - input: - file(counts) from asvcount_med - file(map) from asvgroupscsv - - output: - file("${params.projtag}_ASV_Groupingcounts.csv") into asvgroupcounts - - script: - """ - awk -F "," '{print \$1}' ${counts} | sed '1d' > asv.list - echo "GroupID" >> group.list - for x in \$(cat asv.list); - do group=\$(grep -w \$x ${map} | awk -F "," '{print \$2}') - echo "\$group" >> group.list - done - paste -d',' group.list ${counts} > ${params.projtag}_ASV_Groupingcounts.csv - """ - } - } - - if (!params.skipAminoTyping) { - - process Translate_For_AminoTyping { - - label 'low_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes/Translation", mode: "copy", overwrite: true - - input: - file(fasta) from asvsforAminotyping - - output: - file("${params.projtag}_all_translations.fasta") into amintypegen - file("${params.projtag}_translation_report") into proteinstage_vap_report - - script: - """ - ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_all_translations.fasta --report ${params.projtag}_translation_report - """ - } - - process Generate_AminoTypes { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{clstr,csv,gc}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes/Problematic", mode: "copy", overwrite: true, pattern: '*problematic*.{fasta}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes", mode: "copy", overwrite: true, pattern: '*AminoTypes_noTaxonomy.{fasta}' - - input: - file(prot) from amintypegen - file(asvs) from asvaminocheck - - output: - tuple file("*.fasta"), file("${params.projtag}_AminoTypes.clstr"), file("${params.projtag}_clustered.gc") into ( supplementalfiles ) - file("${params.projtag}_AminoTypes_noTaxonomy.fasta") into ( aminotypesCounts, aminotypesMafft, aminotypesClustal, aminotypesBlast, aminotypesEmboss, aminos_for_med ) - file("${params.projtag}_AminoType_summary_map.csv") into aminomapmed - - script: - """ - set +e - cp ${params.vampdir}/bin/rename_seq.py . - awk 'BEGIN{RS=">";ORS=""}length(\$2)>="${params.minAA}"{print ">"\$0}' ${prot} >${params.projtag}_filtered_translations.fasta - awk 'BEGIN{RS=">";ORS=""}length(\$2)<"${params.minAA}"{print ">"\$0}' ${prot} >${params.projtag}_problematic_translations.fasta - if [ `wc -l ${params.projtag}_problematic_translations.fasta | awk '{print \$1}'` -gt 1 ];then - grep ">" ${params.projtag}_problematic_translations.fasta | awk -F ">" '{print \$2}' > problem_tmp.list - seqtk subseq ${asvs} problem_tmp.list > ${params.projtag}_problematic_nucleotides.fasta - else - rm ${params.projtag}_problematic_translations.fasta - fi - cd-hit -i ${params.projtag}_filtered_translations.fasta -c 1.0 -o ${params.projtag}_unlabeled_types.fasta - sed 's/>Cluster />Cluster_/g' ${params.projtag}_unlabeled_types.fasta.clstr >${params.projtag}_AminoTypes.clstr - grep ">Cluster_" ${params.projtag}_AminoTypes.clstr >tmpclusters.list - grep -w "*" ${params.projtag}_AminoTypes.clstr | awk '{print \$3}' | awk -F "." '{print \$1}' >tmphead.list - grep -w "*" ${params.projtag}_AminoTypes.clstr | awk '{print \$2}' | awk -F "," '{print \$1}' >tmplen.list - paste -d"," tmpclusters.list tmphead.list >tmp.info.csv - grep ">" ${params.projtag}_unlabeled_types.fasta >lala.list - j=1 - for x in \$(cat lala.list);do - echo ">${params.projtag}_AminoType\${j}" >>${params.projtag}_aminoheaders.list - echo "\${x},>${params.projtag}_AminoType\${j}" >>tmpaminotype.info.csv - j=\$(( \${j}+1 )) - done - rm lala.list - awk -F "," '{print \$2}' tmp.info.csv >>tmporder.list - for x in \$(cat tmporder.list);do - grep -w "\$x" tmpaminotype.info.csv | awk -F "," '{print \$2}' >>tmpder.list - done - paste -d "," tmpclusters.list tmplen.list tmphead.list tmpder.list >${params.projtag}_AminoType_summary_map.csv - rm tmp* - ./rename_seq.py ${params.projtag}_unlabeled_types.fasta ${params.projtag}_aminoheaders.list ${params.projtag}_AminoTypes_noTaxonomy.fasta - stats.sh in=${params.projtag}_AminoTypes_noTaxonomy.fasta gc=${params.projtag}_clustered.gc gcformat=4 - """ - } + script: + """ + for filename in ${potus};do + ident=\$( echo \${filename} | awk -F "pcASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') + name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') + clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistanceq.matrix --full --force --threads=${task.cpus} + clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} + for x in *q.matrix;do + pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') + ya=\$(wc -l \$x | awk '{print \$1}') + echo "\$((\$ya-1))" + tail -"\$((\$ya-1))" \$x > \${pre}z.matrix + rm \$x + cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix + rm \${pre}z.matrix + done + done + """ + } - process Generate_AminoType_Matrices { + if (!params.skipPhylogeny) { - label 'low_cpus' + process pcASV_Nucleotide_Phylogeny { - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Matrices", mode: "copy", overwrite: true + label 'norm_cpus' - input: - file(prot) from aminotypesClustal + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*aln.*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*mt*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*iq*' - output: - file("*.matrix") into proclustmatrices - file("*PercentID.matrix") into aminotype_heatmap + input: + file(reads) from pcASV_ntmafft_ch - script: - """ - name=\$( echo ${prot} | awk -F "_noTax" '{print \$1}') - clustalo -i ${prot} --distmat-out=\${name}_PairwiseDistanceq.matrix --full --force --threads=${task.cpus} - clustalo -i ${prot} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} - for x in *q.matrix;do - pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') - ya=\$(wc -l \$x | awk '{print \$1}') - echo "\$((\$ya-1))" - tail -"\$(( \$ya-1))" \$x > \${pre}z.matrix - rm \$x - cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix - rm \${pre}z.matrix - done - """ - } + output: + tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into pcASV_nucleotide_phylogeny_results + file("*iq.treefile") into potu_Ntree_plot - if (!params.skipEMBOSS) { + script: + """ + for filename in ${reads};do + pre=\$( echo \${filename} | awk -F "_noTax" '{print \$1}' ) + mafft --maxiterate 5000 --auto \${filename} >\${pre}_ALN.fasta + trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html - process AminoType_EMBOSS_Analyses { + # pcASV_Nucleotide_ModelTest + modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint - label 'low_cpus' + # pcASV_Nucleotide_Phylogeny + if [ "${params.iqCustomnt}" != "" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -t \${pre}_mt.tree -T auto ${params.iqCustomnt} - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/2dStructure", mode: "copy", overwrite: true, pattern: '*.{garnier}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/HydrophobicMoment", mode: "copy", overwrite: true, pattern: '*HydrophobicMoments.{svg}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/IsoelectricPoint", mode: "copy", overwrite: true, pattern: '*IsoelectricPoint.{iep,svg}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/ProteinProperties", mode: "copy", overwrite: true, pattern: '*.{pepstats,pepinfo}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/ProteinProperties/Plots", mode: "copy", overwrite: true, pattern: '*PropertiesPlot.{svg}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/2dStructure/Plots", mode: "copy", overwrite: true, pattern: '*Helical*.{svg}' + elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} - input: - file(prot) from aminotypesEmboss + elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni - output: - tuple file("*.garnier"), file("*HydrophobicMoments.svg"), file("*IsoelectricPoint*"), file("*.pepstats"), file("*PropertiesPlot*"), file("*Helical*") into amino_emboss + elif [ "${params.nonparametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} - script: - """ - name=\$( echo ${prot} | awk -F ".fasta" '{print \$1}') - garnier -sequence ${prot} -outfile \${name}_2dStructures.garnier - hmoment -seqall ${prot} -graph svg -plot - mv hmoment.svg ./"\${name}"_HydrophobicMoments.svg - iep -sequence ${prot} -graph svg -plot -outfile "\${name}"_IsoelectricPoint.iep - mv iep.svg ./"\${name}"_IsoelectricPoint.svg - pepstats -sequence ${prot} -outfile \${name}_ProteinProperties.pepstats - grep ">" ${prot} | awk -F ">" '{print \$2}' > tmpsequence.list - for x in \$(cat tmpsequence.list);do - echo \$x > tmp1.list - seqtk subseq ${prot} tmp1.list > tmp2.fasta - len=\$(tail -1 tmp2.fasta | awk '{print length}') - pepinfo -sequence tmp2.fasta -graph svg -outfile "\$x"_PropertiesPlot.pepinfo - mv pepinfo.svg ./"\$x"_PropertiesPlot.svg - cat "\$x"_PropertiesPlot.pepinfo >> "\${name}"_PropertiesPlot.pepinfo - rm "\$x"_PropertiesPlot.pepinfo - pepnet -sask -sequence tmp2.fasta -graph svg -sbegin1 1 -send1 \$len - mv pepnet.svg ./"\$x"_HelicalNet.svg - pepwheel -sequence tmp2.fasta -graph svg -sbegin1 1 -send1 \$len - mv pepwheel.svg ./"\$x"_HelicalWheel.svg - rm tmp1.list tmp2.fasta - done - rm tmpsequence.list - """ - } - } + elif [ "${params.parametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni - if (!params.skipTaxonomy) { + else + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni + fi + done + """ + } + } - if (params.headers == "RefSeq") { + process pcASV_AminoAcid_Matrix { - process AminoType_Taxonomy_Inference_RefSeq { + label 'low_cpus' - label 'high_cpus' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Matrix", mode: "copy", overwrite: true - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{csv,tsv}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Taxonomy", mode: "copy", overwrite: true, pattern: '*TaxonomyLabels.fasta' + input: + file(prot) from pcASV_aaMatrix_ch - input: - file(asvs) from aminotypesBlast + output: + file("*.matrix") into pcASVaaMatrix + file("*PercentID.matrix") into potu_aa_heatmap - output: - tuple file("*_phyloformat.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AA_diamond - file("*_summary_for_plot.csv") into taxplot2 - file("*TaxonomyLabels.fasta") into tax_labeled_fasta2 - file("*_quick_Taxbreakdown.csv") into tax_table_amino + script: + """ + for filename in ${prot};do + name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') + clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistanceq.matrix --full --force --threads=${task.cpus} + clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} + for x in *q.matrix;do + pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') + ya=\$(wc -l \$x | awk '{print \$1}') + echo "\$((\$ya-1))" + tail -"\$((\$ya-1))" \$x > \${pre}z.matrix + rm \$x + cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix + rm \${pre}z.matrix + done + done + """ + } - script: - """ - cp ${params.vampdir}/bin/rename_seq.py . - virdb=${params.dbdir}/${params.dbname} - grep ">" \${virdb} > headers.list - headers="headers.list" - name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 - echo "Preparing lists to generate summary .csv's" - echo "[Best hit accession number]" > access.list - echo "[e-value]" > evalue.list - echo "[Bitscore]" > bit.list - echo "[Percent ID (aa)]" > pid.list - echo "[Organism ID]" > "\$name"_virus.list - echo "[Gene]" > "\$name"_genes.list - echo "[AminoType#]" > otu.list - echo "[Sequence length]" > length.list - grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst - echo "extracting genes and names" - touch new_"\$name"_asvnames.txt - for s in \$(cat seqids.lst);do - echo "Checking for \$s hit in diamond output" - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">"\${s}"_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list - virus="NO" - gene="HIT" - echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - echo "\$s done." - fi - done - echo "Now editing "\$name" fasta headers" - ###### rename_seq.py - ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta - awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta - echo "[Sequence header]" > newnames.list - cat new_"\$name"_asvnames.txt >> newnames.list - touch sequence.list - echo " " > sequence.list - grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list - rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - for x in *phyloformat.csv;do - echo "\$x" - lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) - tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; - sed 's/ /_/g' tmpcol.list > tmp2col.list; - cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; - rm tmpcol.list tmp2col.list - done - rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list - """ - } - } else if (params.headers == "RVDB") { - - process AminoType_Taxonomy_Inference_RVDB { - - label 'high_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{csv,tsv}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Taxonomy", mode: "copy", overwrite: true, pattern: '*TaxonomyLabels.fasta' - - input: - file(asvs) from aminotypesBlast - - output: - tuple file("*_phyloformat.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AA_diamond - file("*_summary_for_plot.csv") into taxplot2 - file("*TaxonomyLabels.fasta") into tax_labeled_fasta2 - file("*_quick_Taxbreakdown.csv") into tax_table_amino - - script: - """ - cp ${params.vampdir}/bin/rename_seq.py . - virdb=${params.dbdir}/${params.dbname} - grep ">" \${virdb} > headers.list - headers="headers.list" - name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 - echo "Preparing lists to generate summary .csv's" - echo "[Best hit accession number]" > access.list - echo "[e-value]" > evalue.list - echo "[Bitscore]" > bit.list - echo "[Percent ID (aa)]" > pid.list - echo "[Organism ID]" > "\$name"_virus.list - echo "[Gene]" > "\$name"_genes.list - echo "[AminoType#]" > otu.list - echo "[Sequence length]" > length.list - grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst - if [[ ${params.lca} == "T" ]] - then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list - echo "[Classification]" > classfication.list - else echo "skipped" >> \${name}_quick_Taxbreakdown.csv - echo "[Classification]" > classfication.list - fi - echo "extracting genes and names" - touch new_"\$name"_asvnames.txt - for s in \$(cat seqids.lst);do - echo "Using RVDB headers." - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - if [[ "${params.lca}" == "T" ]] - then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}') - lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') - echo "\$lcla" >> classfication.list - fi - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list - virus="NO" - gene="HIT" - echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - if [[ "${params.lca}" == "T" ]] - then echo "N/A" >> classfication.list - fi - echo "\$s done." - fi - echo "Done with \$s" - done - echo "Now editing "\$name" fasta headers" - ###### rename_seq.py - ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta - awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta - echo "[Sequence header]" > newnames.list - cat new_"\$name"_asvnames.txt >> newnames.list - touch sequence.list - echo " " > sequence.list - grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list - rm "\$name"_tmpssasv.fasta - if [[ "${params.lca}" == "T" ]] - then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list classfication.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv - else paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - fi - for x in *phyloformat.csv;do - echo "\$x" - lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) - tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; - sed 's/ /_/g' tmpcol.list > tmp2col.list; - cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; - rm tmpcol.list tmp2col.list - done - rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list - """ - } - } - } + if (!params.skipEMBOSS) { - if (!params.skipPhylogeny) { + process pcASV_EMBOSS_Analyses { - process AminoType_Phylogeny { + label 'low_cpus' - label 'norm_cpus' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/2dStructure", mode: "copy", overwrite: true, pattern: '*.{garnier}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/HydrophobicMoment", mode: "copy", overwrite: true, pattern: '*HydrophobicMoments.{svg}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/IsoelectricPoint", mode: "copy", overwrite: true, pattern: '*IsoelectricPoint.{iep,svg}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/ProteinProperties", mode: "copy", overwrite: true, pattern: '*.{pepstats,pepinfo}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/ProteinProperties/Plots", mode: "copy", overwrite: true, pattern: '*PropertiesPlot.{svg}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/2dStructure/Plots", mode: "copy", overwrite: true, pattern: '*Helical*.{svg}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*aln.*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Phylogeny/Modeltest", mode: "copy", overwrite: true, pattern: '*mt*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*iq*' + input: + file(prot) from pcASVEMBOSS - input: - file(prot) from aminotypesMafft + output: + tuple file("*.garnier"), file("*HydrophobicMoments.svg"), file("*IsoelectricPoint*"), file("*.pepstats"), file("*PropertiesPlot*"), file("*Helical*") into pcASV_emboss - output: - tuple file("*_aln.fasta"), file("*_aln.html"), file("*.log"), file("*iq*"), file("*mt*") into alignprot_results - file("*iq.treefile") into (amino_rax_plot, amino_repphy) + script: + """ + for filename in ${prot};do + name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') + garnier -sequence \${filename} -outfile \${name}_2dStructures.garnier + hmoment -seqall \${filename} -graph svg -plot + mv hmoment.svg ./"\${name}"_HydrophobicMoments.svg + iep -sequence \${filename} -graph svg -plot -outfile "\${name}"_IsoelectricPoint.iep + mv iep.svg ./"\${name}"_IsoelectricPoint.svg + pepstats -sequence \${filename} -outfile \${name}_ProteinProperties.pepstats + grep ">" \${filename} | awk -F ">" '{print \$2}' > tmpsequence.list + for x in \$(cat tmpsequence.list);do + echo \$x > tmp1.list + seqtk subseq \${filename} tmp1.list > tmp2.fasta + len=\$(tail -1 tmp2.fasta | awk '{print length}') + pepinfo -sequence tmp2.fasta -graph svg -outfile "\$x"_PropertiesPlot.pepinfo + mv pepinfo.svg ./"\$x"_PropertiesPlot.svg + cat "\$x"_PropertiesPlot.pepinfo >> "\${name}"_PropertiesPlot.pepinfo + rm "\$x"_PropertiesPlot.pepinfo + pepnet -sask -sequence tmp2.fasta -graph svg -sbegin1 1 -send1 \$len + mv pepnet.svg ./"\$x"_HelicalNet.svg + pepwheel -sequence tmp2.fasta -graph svg -sbegin1 1 -send1 \$len + mv pepwheel.svg ./"\$x"_HelicalWheel.svg + rm tmp1.list tmp2.fasta + done + rm tmpsequence.list + done + """ + } + } - script: - """ - # Protein_Alignment - pre=\$(echo ${prot} | awk -F "_noTax" '{print \$1}' ) - mafft --thread ${task.cpus} --maxiterate 15000 --auto ${prot} >\${pre}_ALN.fasta - trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html - o-trim-uninformative-columns-from-alignment \${pre}_aln.fasta - mv \${pre}_aln.fasta-TRIMMED ./\${pre}_Aligned_informativeonly.fasta - # Protein_ModelTest - modeltest-ng -i \${pre}_Aligned_informativeonly.fasta -p ${task.cpus} -o \${pre}_mt -d aa -s 203 --disable-checkpoint + if (!params.skipTaxonomy) { - # Protein_Phylogeny - if [ "${params.iqCustomaa}" != "" ];then - iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomaa} + process pcASV_AminoAcid_Taxonomy_Inference { - elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} + label 'high_cpus' - elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{csv,tsv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*dmd.{out}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{fasta}' - elif [ "${params.nonparametric}" != "false" ];then - iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} + input: + file(reads) from pcASV_aaDiamond_ch - elif [ "${params.parametric}" != "false" ];then - iqtree -s\${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + output: + file("*.fasta") into ( pcASV_labeledAA ) + tuple file("*phyloseqObject.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_potuaadiamond + file("*_summary_for_plot.csv") into taxplot4 - else - iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - fi - """ - } + script: + """ + cp ${params.vampdir}/bin/rename_seq.py . + virdb=${params.dbdir}/${params.dbname} + grep ">" \${virdb} >> headers.list + headers="headers.list" + for filename in ${reads};do + name=\$(ls \${filename} | awk -F ".fasta" '{print \$1}') + diamond blastp -q \${filename} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + echo "Preparing lists to generate summary .csv's" + echo "[Best hit accession number]" >access.list + echo "[pcASV sequence length]" >length.list + echo "[e-value]" >evalue.list + echo "[Bitscore]" >bit.list + echo "[Percent ID (aa)]" >pid.list + echo "[pcASVaa#]" >otu.list + echo "[Virus ID]" >"\$name"_virus.list + echo "[Gene]" >"\$name"_genes.list + grep ">" \${filename} | awk -F ">" '{print \$2}' > seqids.lst + echo "extracting genes and names" + touch new_"\$name"_headers.txt + j=1 + for s in \$(cat seqids.lst);do + echo "Checking for \$s hit in diamond output" + if [[ ${params.refseq} == "T" ]];then + echo "RefSeq headers specified" + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >>evalue.list + echo "\$line" | awk '{print \$11}' >>bit.list + echo "\$line" | awk '{print \$12}' >>pid.list + echo "\$line" | awk '{print \$2}' >>length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">pcASVaa\${j}_"\$virus"_"\$gene"" >> new_"\$name"_headers.txt + j=\$((\$j+1)) + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >>access.list + echo "NO_HIT" >>"\$name"_genes.list + echo "NO_HIT" >>"\$name"_virus.list + echo "NO_HIT" >>evalue.list + echo "NO_HIT" >>bit.list + echo "NO_HIT" >>pid.list + echo "NO_HIT" >>length.list + virus="NO" + gene="HIT" + echo ">pcASVaa\${j}_"\$virus"_"\$gene"" >> new_"\$name"_headers.txt + j=\$((\$j+1)) + echo "\$s done." + fi + else + echo "Using RVDB headers." + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') + echo "\$s" >>otu.list + echo "\$acc" >>access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >>evalue.list + echo "\$line" | awk '{print \$11}' >>bit.list + echo "\$line" | awk '{print \$12}' >>pid.list + echo "\$line" | awk '{print \$2}' >>length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >>"\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && + echo "\$virus" | sed 's/_/ /g' >>"\$name"_virus.list + echo ">pcASVaa\${j}_"\$virus"_"\$gene"" >>new_"\$name"_headers.txt + j=\$((\$j+1)) + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >>otu.list + echo "NO_HIT" >>access.list + echo "NO_HIT" >>"\$name"_genes.list + echo "NO_HIT" >>"\$name"_virus.list + echo "NO_HIT" >>evalue.list + echo "NO_HIT" >>bit.list + echo "NO_HIT" >>pid.list + echo "NO_HIT" >>length.list + virus="NO" + gene="HIT" + echo ">pcASVaa\${j}_\${virus}_\${gene}" >>new_"\$name"_headers.txt + j=\$((\$j+1)) + echo "\$s done." + fi + fi + echo "Done with \$s" + done + echo "Now editing "\$name" fasta headers" + ###### rename_seq.py + ./rename_seq.py \${filename} new_"\$name"_headers.txt "\$name"_wTax.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_wTax.fasta > "\$name"_tmpssasv.fasta + echo "[Sequence header]" > newnames.list + cat new_"\$name"_headers.txt >> newnames.list + touch sequence.list + awk 'BEGIN{RS=">";ORS=""}{print \$2"\\n"}' \${name}_tmpssasv.fasta >>sequence.list + rm "\$name"_tmpssasv.fasta + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_summary_phyloseqObject.csv + paste -d"\\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + for x in *phyloseqObject.csv;do + echo "\$x" + lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) + tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; + sed 's/ /_/g' tmpcol.list > tmp2col.list; + cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; + rm tmpcol.list tmp2col.list + done + rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; + rm "\$name"_virus.list + rm "\$name"_genes.list + rm newnames.list + rm access.list + echo "Taxonomy inferred for: \${filename} " + done + rm *headers.list + """ } + } - process Generate_AminoTypes_Counts_Table { + if (!params.skipPhylogeny) { - label 'high_cpus' + process pcASV_Protein_Phylogeny { - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Counts", mode: "copy", overwrite: true + label 'norm_cpus' - input: - file(fasta) from aminotypesCounts - file(merged) from mergeforprotcounts - file(samplist) from samplelist + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*aln.*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Phylogeny/Modeltest", mode: "copy", overwrite: true, pattern: '*mt*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*iq*' - output: - tuple file("*_AminoType_counts.csv"), file("*dmd.out") into counts_summary - file("*_AminoType_counts.csv") into (aminocounts_plot, aminocountmed) + input: + file(prot) from pcASV_aaMafft_ch - script: - """ - set +e - diamond makedb --in ${fasta} --db ${fasta} - diamond blastx -q ${merged} -d ${fasta} -p ${task.cpus} --min-score ${params.ProtCountsBit} --id ${params.ProtCountID} -l ${params.ProtCountsLength} --more-sensitive -o ${params.projtag}_protCounts_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 --max-hsps 1 - echo "OTU_ID" >tmp.col1.txt - echo "Generating sample id list" - grep ">" ${fasta} | awk -F ">" '{print \$2}' | sort | uniq > otuid.list - cat otuid.list >> tmp.col1.txt - echo "Beginning them counts tho my g" - for y in \$( cat ${samplist} );do - echo "Starting with \$y now ..." - grep "\$y" ${params.projtag}_protCounts_dmd.out > tmp."\$y".out - echo "Isolated hits" - echo "Created uniq subject id list" - echo "\$y" > "\$y"_col.txt - echo "Starting my counts" - for z in \$(cat otuid.list);do - echo "Counting \$z hits" - echo "grep -wc "\$z" >> "\$y"_col.txt" - grep -wc "\$z" tmp."\$y".out >> "\$y"_col.txt - echo "\$z counted" - done - done - paste -d "," tmp.col1.txt *col.txt > ${params.projtag}_AminoType_counts.csv - rm tmp* - rm *col.txt - """ - } - } + output: + tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into pcASV_protein_phylogeny_results + file("*iq.treefile") into potu_Atree_plot - if (params.aminoMED) { + script: + """ + for filename in ${prot};do + pre=\$( echo \${filename} | awk -F ".fasta" '{print \$1}' ) + mafft --maxiterate 5000 --auto \${filename} >\${pre}_ALN.fasta + trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html - process AminoType_Minimum_Entropy_Decomposition { + # pcASV_Protein_ModelTest + modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d aa -s 203 --disable-checkpoint - label 'low_cpus' + # pcASV_Protein_Phylogeny + if [ "${params.iqCustomaa}" != "" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomaa} - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes/MED", mode: "copy", overwrite: true + elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} - input: - file(aminos) from aminos_for_med + elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni - output: - file("*_AminoType_Grouping.csv") into atygroupscsv - file("${params.projtag}_AminoType_group_reps_aligned.fasta") into atygroupreps + elif [ "${params.nonparametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} - script: - """ - #alignment - mafft --thread ${task.cpus} --maxiterate 15000 --auto ${aminos} > ${params.projtag}_AminoTypes_mafftAlign.fasta - #trimming - trimal -in ${params.projtag}_AminoTypes_mafftAlign.fasta -out ${params.projtag}_AminoTypes_mafftAligned.fasta -keepheader -fasta -automated1 - rm ${params.projtag}_AminoTypes_mafftAlign.fasta - o-trim-uninformative-columns-from-alignment ${params.projtag}_AminoTypes_mafftAligned.fasta - mv ${params.projtag}_AminoTypes_mafftAligned.fasta-TRIMMED ./${params.projtag}_AminoTypes_Aligned_informativeonly.fasta - #entopy analysis - entropy-analysis ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta - #Decomposition - oligotype ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta-ENTROPY -o ${params.projtag}_AminoTypeMED_${params.aminoC} -M 1 -c ${params.aminoC} -N ${task.cpus} --skip-check-input --no-figures --skip-gen-html - #generatemaps - cd ./${params.projtag}_AminoTypeMED_${params.aminoC}/OLIGO-REPRESENTATIVES/ - echo "AminoType,Group,IDPattern" - j=1 - for x in *_unique; - do gid=\$(echo \$x | awk -F "_" '{print \$1}') - uni=\$(echo \$x | awk -F ""\${gid}"_" '{print \$2}' | awk -F "_uni" '{print \$1}') - grep ">" "\$gid"_"\$uni" | awk -F ">" '{print \$2}' > asv.list - seqtk subseq ../../${aminos} asv.list > Group"\${j}"_sequences.fasta - for z in \$( cat asv.list) - do echo ""\$z",Group"\$j","\$uni"" >> ${params.projtag}_AminoType_Grouping.csv - - done - rm asv.list - echo ">Group\${j}" >> ${params.projtag}_AminoType_group_reps_aligned.fasta - echo "\$uni" > group.list - seqtk subseq ../OLIGO-REPRESENTATIVES.fasta group.list > group.fasta - tail -1 group.fasta >> ${params.projtag}_AminoType_group_reps_aligned.fasta - mv "\$gid"_"\$uni" ./Group"\$j"_"\$uni"_aligned.fasta - mv "\$gid"_"\$uni"_unique ./Group"\$j"_"\$uni"_unqiues_aligned.fasta - rm "\$gid"*.cPickle - j=\$((\$j+1)) - done - mv ${params.projtag}_AminoType_Grouping.csv ../../ - mv ${params.projtag}_AminoType_group_reps_aligned.fasta ../../ - cd .. + elif [ "${params.parametric}" != "false" ];then + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - """ - } + else + iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + fi + done + """ + } + } - process AminoType_MED_Reps_phylogeny { + process Generate_pcASV_Protein_Counts { - label 'low_cpus' + label 'high_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/MED/Phylogeny/Modeltest", mode: "copy", overwrite: true, pattern: '*mt*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/MED/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*iq*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Counts", mode: "copy", overwrite: true - input: - file(reps) from atygroupreps + input: + file(fasta) from pcASV_aaCounts_ch + file(merged) from mergeforpcASVaacounts + file(samplist) from samplistpotu - output: - file("*_AminoType_Group_Reps*") into align_results_aminmed - file("*iq.treefile") into amino_group_rep_tree + output: + tuple file("*_counts.csv"), file("*dmd.out") into potuaacounts_summary + file("*counts.csv") into potu_Acounts - script: - """ - # Protein_ModelTest - modeltest-ng -i ${reps} -p ${task.cpus} -o ${params.projtag}_AminoType_Group_Reps_mt -d aa -s 203 --disable-checkpoint + script: + """ + set +e + for filename in ${fasta};do + potu="\$( echo \${filename} | awk -F "_" '{print \$3}')" + diamond makedb --in \${filename} --db \${filename} + diamond blastx -q ${merged} -d \${filename} -p ${task.cpus} --min-score ${params.ProtCountsBit} --id ${params.ProtCountID} -l ${params.ProtCountsLength} --more-sensitive -o ${params.projtag}_\${potu}_Counts_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 --max-hsps 1 + echo "OTU_ID" >tmp.col1.txt + echo "Generating sample id list" + grep ">" \${filename} | awk -F ">" '{print \$2}' | sort | uniq > otuid.list + cat otuid.list >> tmp.col1.txt + echo "Beginning them counts tho my g" + for y in \$( cat ${samplist} );do + echo "Starting with \$y now ..." + grep "\$y" ${params.projtag}_\${potu}_Counts_dmd.out > tmp."\$y".out + echo "Isolated hits" + echo "Created uniq subject id list" + echo "\$y" > "\$y"_col.txt + echo "Starting my counts" + for z in \$(cat otuid.list);do + echo "Counting \$z hits" + echo "grep -wc "\$z" >> "\$y"_col.txt" + grep -wc "\$z" tmp."\$y".out >> "\$y"_col.txt + echo "\$z counted" + done + done + paste -d "," tmp.col1.txt *col.txt > ${params.projtag}_\${potu}_counts.csv + rm tmp* + rm *col.txt + done + """ + } + } - # Protein_Phylogeny - if [ "${params.iqCustomaa}" != "" ];then - iqtree -s ${reps} --prefix ${params.projtag}_AminoType_Group_Reps_iq --redo -T auto ${params.iqCustomaa} + if (!params.skipReport) { - elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') - iqtree -s ${reps} --prefix ${params.projtag}_AminoType_Group_Reps_iq -m \${mod} --redo -nt auto -b ${params.boots} + if (!params.skipAdapterRemoval) { - elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') - iqtree -s ${reps} --prefix ${params.projtag}_AminoType_Group_Reps_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni + process combine_csv { - elif [ "${params.nonparametric}" != "false" ];then - iqtree -s ${reps} --prefix ${params.projtag}_AminoType_Group_Reps_iq -m MFP --redo -nt auto -b ${params.boots} + input: + file(csv) from fastp_csv + .collect() - elif [ "${params.parametric}" != "false" ];then - iqtree -s ${reps} --prefix ${params.projtag}_AminoType_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + output: + file("final_reads_stats.csv") into ( fastp_csv1, fastp_csv2, fastp_csv3, fastp_csv4, fastp_csv5 ) - else - iqtree -s ${reps} --prefix ${params.projtag}_AminoType_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - fi - """ - } + script: + """ + cat ${csv} >all_reads_stats.csv + head -n1 all_reads_stats.csv >tmp.names.csv + cat all_reads_stats.csv | grep -v ""Sample,Total_"" >tmp.reads.stats.csv + cat tmp.names.csv tmp.reads.stats.csv >final_reads_stats.csv + rm tmp.names.csv tmp.reads.stats.csv + """ - process Adding_AminoType_MED_Info { + } + } - label 'low_cpus' + if (params.ncASV) { - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/MED/", mode: "copy", overwrite: true - input: - file(counts) from aminocountmed - file(tree) from amino_repphy - file(map) from atygroupscsv + process Report_ASV { - output: - file("${params.projtag}_AminoType_Groupingcounts.csv") into amino_groupcounts + label 'norm_cpus' - script: - """ - awk -F "," '{print \$1}' ${counts} | sed '1d' > amino.list - echo "GroupID" >> group.list - for x in \$(cat amino.list); - do group=\$(grep -w \$x ${map} | awk -F "," '{print \$2}') - echo "\$group" >> group.list - done - paste -d',' group.list ${counts} > ${params.projtag}_AminoType_Groupingcounts.csv - """ - } - } + publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport", mode: "copy", overwrite: true + + input: + file(counts) from asv_counts_plots + file(taxonomy) from taxplot1 + file(matrix) from asv_heatmap + file(readsstats) from fastp_csv1 + + output: + file("*.html") into report_summaryA - if (params.pcASV) { // ASV_nucl -> ASV_aa -> clusteraa by %id with ch-hit -> extract representative nucl sequences to generate new OTU file + script: + """ + name=\$( echo ${taxonomy} | awk -F "_summary_for_plot.csv" '{print \$1}') + cp ${params.vampdir}/bin/vAMPirus_ReportA.Rmd . + cp ${params.vampdir}/example_data/conf/vamplogo.png . + Rscript -e "rmarkdown::render('vAMPirus_ReportA.Rmd',output_file='\${name}_ASV_Report.html')" \${name} \ + ${readsstats} \ + ${counts} \ + ${params.metadata} \ + ${params.minimumCounts} \ + ${matrix} \ + ${taxonomy} \ + ${params.trymax} \ + ${params.stats} + """ + } - process Translation_For_pcASV_Generation { + process Report_ncASV { - label 'low_cpus' + label 'norm_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV/Translation", mode: "copy", overwrite: true, pattern: '*_ASV_translations*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport/ncASV", mode: "copy", overwrite: true - input: - file(fasta) from nucl2aa + input: + file(counts) from notu_counts_plots + file(taxonomy) from taxplot1a + file(matrix) from notu_heatmap + file(phylogeny) from nucl_phyl_plot + file(readsstats) from fastp_csv2 - output: - file("*ASV*translations.fasta") into clustering_aa - file("*_ASV_translations_report") into reportaa_VR - file("*_ASV_nucleotide.fasta") into asvfastaforaaclust + output: + file("*.html") into report_summaryB - script: - """ - ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_ASV_translations.fasta --report ${params.projtag}_ASV_translations_report - cp ${fasta} ${params.projtag}_ASV_nucleotide.fasta - """ + script: + """ + cp ${params.vampdir}/bin/vAMPirus_ReportB.Rmd . + cp ${params.vampdir}/example_data/conf/vamplogo.png . + for x in *_summary_for_plot.csv;do + name=\$( echo \${x} | awk -F "_summary_for_plot.csv" '{print \$1}') + id=\$( echo \${x} | awk -F "_summary_for_plot.csv" '{print \$1}' | cut -f 2 -d "." ) + Rscript -e "rmarkdown::render('vAMPirus_ReportB.Rmd',output_file='\${name}_ncASV\${id}_Report.html')" \${name} \ + ${readsstats} \ + \$( echo ${counts} | tr " " "\\n" | grep "\${id}" ) \ + ${params.metadata} \ + ${params.minimumCounts} \ + \$( echo ${matrix} | tr " " "\\n" | grep "\${id}" ) \ + \$( echo ${taxonomy} | tr " " "\\n" | grep "\${id}" ) \ + \$( echo ${phylogeny} | tr " " "\\n" | grep "\${id}" ) \ + ${params.trymax} \ + ${params.stats} + done + """ } - process Generate_pcASVs { + if (!params.skipAminoTyping) { + + process Report_AminoTypes { label 'norm_cpus' - tag "${mtag}" - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV", mode: "copy", overwrite: true, pattern: '*pcASV*.{fasta}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{clstr,csv,gc}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV/Problematic", mode: "copy", overwrite: true, pattern: '*problem*.{fasta}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport", mode: "copy", overwrite: true input: - each x from 1..naa - file(fasta) from clustering_aa - file(asvs) from asvfastaforaaclust + file(counts) from aminocounts_plot + file(taxonomy) from taxplot2 + file(matrix) from aminotype_heatmap + file(phylogeny) from amino_rax_plot + file(readsstats) from fastp_csv5 output: - tuple nid, file("${params.projtag}_nucleotide_pcASV*.fasta") into ( pcASV_ntDiamond_ch, pcASV_nt_counts_ch, pcASV_ntmatrix_ch, pcASV_ntmafft_ch ) - tuple nid, file("*_aminoacid_pcASV*_noTaxonomy.fasta") into ( pcASV_aaMatrix_ch, pcASV_aaDiamond_ch, pcASV_aaMafft_ch, pcASV_aaCounts_ch, pcASVEMBOSS ) - tuple nid, file("*.fasta"), file("*.clstr"), file("*.csv"), file("*.gc") into ( pcASVsupplementalfiles ) + file("*.html") into report_summaryE script: - // add awk script to count seqs - nid=slist2.get(x-1) - mtag="ID=" + slist2.get(x-1) """ - set +e - cp ${params.vampdir}/bin/rename_seq.py . - awk 'BEGIN{RS=">";ORS=""}length(\$2)>="${params.minAA}"{print ">"\$0}' ${fasta} > ${params.projtag}_filtered_proteins.fasta - cd-hit -i ${params.projtag}_filtered_proteins.fasta -c .${nid} -o ${params.projtag}_pcASV${nid}.fasta - sed 's/>Cluster />Cluster_/g' ${params.projtag}_pcASV${nid}.fasta.clstr >${params.projtag}_pcASV${nid}.clstr - grep ">Cluster_" ${params.projtag}_pcASV${nid}.clstr >temporaryclusters.list - y=\$(grep -c ">Cluster_" ${params.projtag}_pcASV${nid}.clstr) - echo ">Cluster_"\${y}"" >> ${params.projtag}_pcASV${nid}.clstr - t=1 - b=1 - for x in \$(cat temporaryclusters.list);do - echo "Extracting \$x" - name="\$( echo \$x | awk -F ">" '{print \$2}')" - clust="pcASV"\${t}"" - echo "\${name}" - awk '/^>'\${name}'\$/,/^>Cluster_'\${b}'\$/' ${params.projtag}_pcASV${nid}.clstr > "\${name}"_"\${clust}"_tmp.list - t=\$(( \${t}+1 )) - b=\$(( \${b}+1 )) - done - - ls *_tmp.list - u=1 - for x in *_tmp.list;do - name="\$(echo \$x | awk -F "_p" '{print \$1}')" - echo "\${name}" - cluster="\$(echo \$x | awk -F "_" '{print \$3}')" - echo "\${cluster}" - grep "ASV" \$x | awk -F ", " '{print \$2}' | awk -F "_" '{print \$1}' | awk -F ">" '{print \$2}' > \${name}_\${cluster}_seqs_tmps.list - seqtk subseq ${asvs} \${name}_\${cluster}_seqs_tmps.list > \${name}_\${cluster}_nucleotide_sequences.fasta - vsearch --cluster_fast \${name}_\${cluster}_nucleotide_sequences.fasta --id 0.2 --centroids \${name}_\${cluster}_centroids.fasta - grep ">" \${name}_\${cluster}_centroids.fasta >> \${name}_\${cluster}_tmp_centroids.list - for y in \$( cat \${name}_\${cluster}_tmp_centroids.list );do - echo ">\${cluster}_type"\$u"" >> \${name}_\${cluster}_tmp_centroid.newheaders - u=\$(( \${u}+1 )) - done - u=1 - ./rename_seq.py \${name}_\${cluster}_centroids.fasta \${name}_\${cluster}_tmp_centroid.newheaders \${cluster}_types_labeled.fasta - done - cat *_types_labeled.fasta >> ${params.projtag}_nucleotide_pcASV${nid}_noTaxonomy.fasta - grep -w "*" ${params.projtag}_pcASV${nid}.clstr | awk '{print \$3}' | awk -F "." '{print \$1}' >tmphead.list - grep -w "*" ${params.projtag}_pcASV${nid}.clstr | awk '{print \$2}' | awk -F "," '{print \$1}' >tmplen.list - paste -d"," temporaryclusters.list tmphead.list >tmp.info.csv - grep ">" ${params.projtag}_pcASV${nid}.fasta >lala.list - j=1 - for x in \$(cat lala.list);do - echo ">${params.projtag}_pcASV\${j}" >>${params.projtag}_aminoheaders.list - echo "\${x},>${params.projtag}_pcASV\${j}" >>tmpaminotype.info.csv - j=\$(( \${j}+1 )) - done - rm lala.list - awk -F "," '{print \$2}' tmp.info.csv >>tmporder.list - for x in \$(cat tmporder.list);do - grep -w "\$x" tmpaminotype.info.csv | awk -F "," '{print \$2}' >>tmpder.list - done - paste -d "," temporaryclusters.list tmplen.list tmphead.list tmpder.list >${params.projtag}_pcASVCluster${nid}_summary.csv - ./rename_seq.py ${params.projtag}_pcASV${nid}.fasta ${params.projtag}_aminoheaders.list ${params.projtag}_aminoacid_pcASV${nid}_noTaxonomy.fasta - stats.sh in=${params.projtag}_aminoacid_pcASV${nid}_noTaxonomy.fasta gc=${params.projtag}_pcASV${nid}_aminoacid_clustered.gc gcformat=4 - stats.sh in=${params.projtag}_nucleotide_pcASV${nid}_noTaxonomy.fasta gc=${params.projtag}_pcASV${nid}_nucleotide_clustered.gc gcformat=4 - awk 'BEGIN{RS=">";ORS=""}length(\$2)<"${params.minAA}"{print ">"\$0}' ${fasta} >${params.projtag}_pcASV${nid}_problematic_translations.fasta - if [ `wc -l ${params.projtag}_pcASV${nid}_problematic_translations.fasta | awk '{print \$1}'` -gt 1 ];then - grep ">" ${params.projtag}_pcASV${nid}_problematic_translations.fasta | awk -F ">" '{print \$2}' > problem_tmp.list - seqtk subseq ${asvs} problem_tmp.list > ${params.projtag}_pcASV${nid}_problematic_nucleotides.fasta - else - rm ${params.projtag}_pcASV${nid}_problematic_translations.fasta - fi - rm *.list - rm Cluster* - rm *types* - rm *tmp* - rm ${params.projtag}_pcASV${nid}.fast* + name=\$( echo ${taxonomy} | awk -F "_summary_for_plot.csv" '{print \$1}') + cp ${params.vampdir}/bin/vAMPirus_ReportB.Rmd . + cp ${params.vampdir}/example_data/conf/vamplogo.png . + Rscript -e "rmarkdown::render('vAMPirus_ReportB.Rmd',output_file='\${name}_AminoType_Report.html')" \${name} \ + ${readsstats} \ + ${counts} \ + ${params.metadata} \ + ${params.minimumCounts} ${matrix} \ + ${taxonomy} \ + ${phylogeny} \ + ${params.trymax} \ + ${params.stats} """ } + } + } else { - if (!params.skipTaxonomy) { - - if (params.headers == "RefSeq") { - - process pcASV_Nucleotide_Taxonomy_Inference_RefSeq { - - label 'high_cpus' + process Report_ASVs { - tag "${mtag}" + label 'norm_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{csv,tsv}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*dmd.{out}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{fasta}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport", mode: "copy", overwrite: true input: - tuple nid, file(asvs) from pcASV_ntDiamond_ch + file(counts) from asv_counts_plots + file(taxonomy) from taxplot1 + file(matrix) from asv_heatmap + file(phylogeny) from nucl_phyl_plot + file(readsstats) from fastp_csv1 output: - file("*.fasta") into ( pcASV_labeled ) - tuple file("*_phyloformat.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AAdiamond - tuple nid, file("*_summary_for_plot.csv") into taxplot3 - tuple nid, file("*_quick_Taxbreakdown.csv") into tax_table_pcasvnt + file("*.html") into report_summaryA script: - mtag="ID=" + nid """ - set +e - cp ${params.vampdir}/bin/rename_seq.py . - virdb=${params.dbdir}/${params.dbname} - grep ">" \${virdb} > headers.list - headers="headers.list" - name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 - echo "Preparing lists to generate summary .csv's" - echo "[Best hit accession number]" > access.list - echo "[e-value]" > evalue.list - echo "[Bitscore]" > bit.list - echo "[Percent ID (aa)]" > pid.list - echo "[Organism ID]" > "\$name"_virus.list - echo "[Gene]" > "\$name"_genes.list - echo "[pcASV#]" > otu.list - echo "[Sequence length]" > length.list - grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst - echo "extracting genes and names" - touch new_"\$name"_asvnames.txt - for s in \$(cat seqids.lst);do - echo "Checking for \$s hit in diamond output" - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">"\${s}"_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list - virus="NO" - gene="HIT" - echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - echo "\$s done." - fi - done - echo "Now editing "\$name" fasta headers" - ###### rename_seq.py - ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta - awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta - echo "[Sequence header]" > newnames.list - cat new_"\$name"_asvnames.txt >> newnames.list - touch sequence.list - echo " " > sequence.list - grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list - rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - for x in *phyloformat.csv;do - echo "\$x" - lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) - tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; - sed 's/ /_/g' tmpcol.list > tmp2col.list; - cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; - rm tmpcol.list tmp2col.list - done - rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list + name=\$( echo ${taxonomy} | awk -F "_summary_for_plot.csv" '{print \$1}') + cp ${params.vampdir}/bin/vAMPirus_ReportB.Rmd . + cp ${params.vampdir}/example_data/conf/vamplogo.png . + Rscript -e "rmarkdown::render('vAMPirus_ReportB.Rmd',output_file='\${name}_ASV_Report.html')" \${name} \ + ${readsstats} \ + ${counts} \ + ${params.metadata} \ + ${params.minimumCounts} \ + ${matrix} \ + ${taxonomy} \ + ${phylogeny} \ + ${params.trymax} \ + ${params.stats} """ + } + + if (!params.skipAminoTyping) { + + process Report_AminoType { + + label 'norm_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport", mode: "copy", overwrite: true + + input: + file(counts) from aminocounts_plot + file(taxonomy) from taxplot2 + file(matrix) from aminotype_heatmap + file(phylogeny) from amino_rax_plot + file(readsstats) from fastp_csv5 + + output: + file("*.html") into report_summaryE + + script: + """ + name=\$( echo ${taxonomy} | awk -F "_summary_for_plot.csv" '{print \$1}') + cp ${params.vampdir}/bin/vAMPirus_ReportB.Rmd . + cp ${params.vampdir}/example_data/conf/vamplogo.png . + Rscript -e "rmarkdown::render('vAMPirus_ReportB.Rmd',output_file='\${name}_AminoType_Report.html')" \${name} \ + ${readsstats} \ + ${counts} \ + ${params.metadata} \ + ${params.minimumCounts} ${matrix} \ + ${taxonomy} \ + ${phylogeny} \ + ${params.trymax} \ + ${params.stats} + """ } - } else if (params.headers == "RVDB") { - - process pcASV_Nucleotide_Taxonomy_Inference_RVDB { - - label 'high_cpus' - - tag "${mtag}" - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{csv,tsv}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*dmd.{out}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{fasta}' - - input: - tuple nid, file(asvs) from pcASV_ntDiamond_ch - - output: - file("*.fasta") into ( pcASV_labeled ) - tuple file("*_phyloformat.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AAdiamond - tuple nid, file("*_summary_for_plot.csv") into taxplot3 - tuple nid, file("*_quick_Taxbreakdown.csv") into tax_table_pcasvnt - - script: - mtag="ID=" + nid - """ - set +e - cp ${params.vampdir}/bin/rename_seq.py . - virdb=${params.dbdir}/${params.dbname} - grep ">" \${virdb} > headers.list - headers="headers.list" - name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 - echo "Preparing lists to generate summary .csv's" - echo "[Best hit accession number]" > access.list - echo "[e-value]" > evalue.list - echo "[Bitscore]" > bit.list - echo "[Percent ID (aa)]" > pid.list - echo "[Organism ID]" > "\$name"_virus.list - echo "[Gene]" > "\$name"_genes.list - echo "[pcASV#]" > otu.list - echo "[Sequence length]" > length.list - grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst - if [[ ${params.lca} == "T" ]] - then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list - echo "[Classification]" > classfication.list - else echo "skipped" >> \${name}_quick_Taxbreakdown.csv - echo "[Classification]" > classfication.list - fi - echo "extracting genes and names" - touch new_"\$name"_asvnames.txt - for s in \$(cat seqids.lst);do - echo "Using RVDB headers." - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - if [[ "${params.lca}" == "T" ]] - then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}') - lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') - echo "\$lcla" >> classfication.list - fi - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list - virus="NO" - gene="HIT" - echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - if [[ "${params.lca}" == "T" ]] - then echo "N/A" >> classfication.list - fi - echo "\$s done." - fi - echo "Done with \$s" - done - echo "Now editing "\$name" fasta headers" - ###### rename_seq.py - ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta - awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta - echo "[Sequence header]" > newnames.list - cat new_"\$name"_asvnames.txt >> newnames.list - touch sequence.list - echo " " > sequence.list - grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list - rm "\$name"_tmpssasv.fasta - if [[ "${params.lca}" == "T" ]] - then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list classfication.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv - else paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - fi - for x in *phyloformat.csv;do - echo "\$x" - lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) - tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; - sed 's/ /_/g' tmpcol.list > tmp2col.list; - cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; - rm tmpcol.list tmp2col.list - done - rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list - """ } } - } + if (params.pcASV) { - process Generate_Nucleotide_pcASV_Counts { + process Report_pcASV_AminoAcid { label 'norm_cpus' - tag "${mtag}" - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Counts", mode: "copy", overwrite: true, pattern: '*.{biome,csv,txt}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport/pcASV/Aminoacid", mode: "copy", overwrite: true input: - tuple nid, file(potus) from pcASV_nt_counts_ch - file(merged) from pcASV_mergedreads_ch + file(counts) from potu_Acounts + file(taxonomy) from taxplot4 + file(matrix) from potu_aa_heatmap + file(phylogeny) from potu_Atree_plot + file(readsstats) from fastp_csv3 output: - tuple file("*_counts.txt"), file("*_counts.biome") into pcASVcounts_vsearch - tuple nid, file("*.csv") into potu_Ncounts_for_report + file("*.html") into report_summaryC script: - mtag="ID=" + nid """ - name=\$( echo ${potus} | awk -F ".fasta" '{print \$1}') - vsearch --usearch_global ${merged} --db ${potus} --id .${nid} --threads ${task.cpus} --otutabout \${name}_counts.txt --biomout \${name}_counts.biome - cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv - sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv - rm \${name}_count.csv + cp ${params.vampdir}/bin/vAMPirus_ReportB.Rmd . + cp ${params.vampdir}/example_data/conf/vamplogo.png . + for x in *_summary_for_plot.csv;do + name=\$( echo \${x} | awk -F "_noTaxonomy_summary_for_plot.csv" '{print \$1}') + id=\$( echo \${x} | awk -F "_noTaxonomy_summary_for_plot.csv" '{print \$1}' | cut -f 2 -d "." ) + Rscript -e "rmarkdown::render('vAMPirus_ReportB.Rmd',output_file='\${name}_pcASVaa\${id}_Report.html')" \${name} \ + ${readsstats} \ + \$( echo ${counts} | tr " " "\\n" | grep "\${id}" ) \ + ${params.metadata} \ + ${params.minimumCounts} \ + \$( echo ${matrix} | tr " " "\\n" | grep "\${id}" ) \ + \$( echo ${taxonomy} | tr " " "\\n" | grep "\${id}" ) \ + \$( echo ${phylogeny} | tr " " "\\n" | grep "\${id}" ) \ + ${params.trymax} \ + ${params.stats} + done """ - } - - process Generate_pcASV_Nucleotide_Matrix { + } - label 'low_cpus' + process Report_pcASV_Nucleotide { - tag "${mtag}" + label 'norm_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Matrix", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport/pcASV/Nucleotide", mode: "copy", overwrite: true input: - tuple nid, file(potus) from pcASV_ntmatrix_ch + file(counts) from potu_Ncounts_for_report + file(taxonomy) from taxplot3 + file(matrix) from potu_nucl_heatmap + file(phylogeny) from potu_Ntree_plot + file(readsstats) from fastp_csv4 output: - file("*.matrix") into pcASVclustmatrices - tuple nid, file("*PercentID.matrix") into potu_nucl_heatmap + file("*.html") into report_summaryD script: - //check --percent-id second clustalo - mtag="ID=" + nid """ - name=\$( echo ${potus} | awk -F ".fasta" '{print \$1}') - clustalo -i ${potus} --distmat-out=\${name}_PairwiseDistanceq.matrix --full --force --threads=${task.cpus} - clustalo -i ${potus} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} - cat \${name}_PercentIDq.matrix | tr " " "," | grep "," >\${name}_PercentID.matrix - rm \${name}_PercentIDq.matrix + cp ${params.vampdir}/bin/vAMPirus_ReportB.Rmd . + cp ${params.vampdir}/example_data/conf/vamplogo.png . + for x in *_summary_for_plot.csv;do + name=\$( echo \${x} | awk -F "_summary_for_plot.csv" '{print \$1}') + id=\$( echo \${x} | awk -F "_summary_for_plot.csv" '{print \$1}' | cut -f 2 -d "." ) + Rscript -e "rmarkdown::render('vAMPirus_ReportB.Rmd',output_file='\${name}_pcASVnt\${id}_Report.html')" \${name} \ + ${readsstats} \ + \$( echo ${counts} | tr " " "\\n" | grep "\${id}" ) \ + ${params.metadata} \ + ${params.minimumCounts} \ + \$( echo ${matrix} | tr " " "\\n" | grep "\${id}" ) \ + \$( echo ${taxonomy} | tr " " "\\n" | grep "\${id}" ) \ + \$( echo ${phylogeny} | tr " " "\\n" | grep "\${id}" ) \ + ${params.trymax} \ + ${params.stats} + done """ } + } + } - if (!params.skipPhylogeny) { +} else if (params.DataCheck) { - process pcASV_Nucleotide_Phylogeny { + println("\n\tRunning vAMPirus DataCheck \n") - label 'norm_cpus' + process QualityCheck_1DC { - tag "${mtag}" + label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*aln.*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*mt*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*iq*' + tag "${sample_id}" - input: - tuple nid, file(reads) from pcASV_ntmafft_ch + publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/FastQC/PreClean", mode: "copy", overwrite: true - output: - tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into pcASV_nucleotide_phylogeny_results - tuple nid, file("*iq.treefile") into potu_Ntree_plot + input: + tuple sample_id, file(reads) from reads_qc_ch - script: - mtag="ID=" + nid - """ - pre=\$( echo ${reads} | awk -F "_noTax" '{print \$1}' ) - mafft --maxiterate 5000 --auto ${reads} >\${pre}_ALN.fasta - trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html - o-trim-uninformative-columns-from-alignment \${pre}_aln.fasta - mv \${pre}_aln.fasta-TRIMMED ./\${pre}_Aligned_informativeonly.fasta - # pcASV_Nucleotide_ModelTest - modeltest-ng -i \${pre}_Aligned_informativeonly.fasta -p ${task.cpus} -o \${pre}_noTaxonomy_mt -d nt -s 203 --disable-checkpoint + output: + tuple sample_id, file("*_fastqc.{zip,html}") into fastqc_results_OAS + + script: + """ + fastqc --quiet --threads ${task.cpus} ${reads} + """ + } + + process Adapter_Removal_DC { + + label 'norm_cpus' + + tag "${sample_id}" + + publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/AdapterRemoval", mode: "copy", overwrite: true, pattern: "*.filter.fq" + publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/AdapterRemoval/fastpOut", mode: "copy", overwrite: true, pattern: "*.fastp.{json,html}" + + input: + tuple sample_id, file(reads) from reads_ch + + output: + tuple sample_id, file("*.fastp.{json,html}") into fastp_results + tuple sample_id, file("*.filter.fq") into reads_fastp_ch + file("*.csv") into fastp_csv + + script: + """ + echo ${sample_id} + + fastp -i ${reads[0]} -I ${reads[1]} -o left-${sample_id}.filter.fq -O right-${sample_id}.filter.fq --detect_adapter_for_pe \ + --average_qual 25 -c --overrepresentation_analysis --html ${sample_id}.fastp.html --json ${sample_id}.fastp.json --thread ${task.cpus} \ + --report_title ${sample_id} + + bash get_readstats.sh ${sample_id}.fastp.json + """ + } + + process Primer_Removal_DC { + + label 'norm_cpus' + + tag "${sample_id}" + + publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/PrimerRemoval", mode: "copy", overwrite: true + + input: + tuple sample_id, file(reads) from reads_fastp_ch + + output: + tuple sample_id, file("*bbduk*.fastq.gz") into ( reads_bbduk_ch, readsforqc2 ) + + script: + // add cocktail primer removal, easy, set a list, line by line seperated by , in a for loop + if ( params.fwd == "" && params.rev == "" && !params.multi) { + """ + bbduk.sh in1=${reads[0]} out=${sample_id}_bb_R1.fastq.gz ftl=${params.defaultFwdTrim} t=${task.cpus} + bbduk.sh in=${reads[1]} out=${sample_id}_bb_R2.fastq.gz ftl=${params.defaultRevTrim} t=${task.cpus} + repair.sh in1=${sample_id}_bb_R1.fastq.gz in2=${sample_id}_bb_R2.fastq.gz out1=${sample_id}_bbduk_R1.fastq.gz out2=${sample_id}_bbduk_R2.fastq.gz minlength=${params.minilen} outs=sing.fq repair + """ + } else if ( params.GlobTrim && !params.GlobTrim == "" ) { + """ + FTRIM=\$( echo ${GlobTrim} | cut -f 1 -d "," ) + RTRIM=\$( echo ${GlobTrim} | cut -f 2 -d "," ) + bbduk.sh in=${reads[0]} out=${sample_id}_bb_R1.fastq.gz ftl=\${FTRIM} t=${task.cpus} + bbduk.sh in=${reads[1]} out=${sample_id}_bb_R2.fastq.gz ftl=\${RTRIM} t=${task.cpus} + repair.sh in1=${sample_id}_bb_R1.fastq.gz in2=${sample_id}_bb_R2.fastq.gz out1=${sample_id}_bbduk_R1.fastq.gz out2=${sample_id}_bbduk_R2.fastq.gz minlength=${params.minilen} outs=sing.fq repair + """ + } else if ( params.multi && params.primers ) { + """ + bbduk.sh in=${reads[0]} in2=${reads[1]} out=${sample_id}_bbduk_R1.fastq.gz out2=${sample_id}_bbduk_R2.fastq.gz ref=${params.primers} copyundefined=t t=${task.cpus} restrictleft=${params.primerLength} k=${params.maxkmer} ordered=t mink=${params.minkmer} ktrim=l ecco=t rcomp=t minlength=${params.minilen} tbo tpe + """ + } else { + """ + bbduk.sh in=${reads[0]} in2=${reads[1]} out=${sample_id}_bbduk_R1.fastq.gz out2=${sample_id}_bbduk_R2.fastq.gz literal=${params.fwd},${params.rev} copyundefined=t t=${task.cpus} restrictleft=${params.primerLength} k=${params.maxkmer} ordered=t mink=${params.minkmer} ktrim=l ecco=t rcomp=t minlength=${params.minilen} tbo tpe + """ + } + } + + process QualityCheck_2_DC { + + label 'low_cpus' + + tag "${sample_id}" + + publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/FastQC/PostClean", mode: "copy", overwrite: true + + input: + tuple sample_id, file(reads) from readsforqc2 + + output: + tuple sample_id, file("*_fastqc.{zip,html}") into fastqc2_results_OAS + + script: + """ + fastqc --quiet --threads ${task.cpus} ${reads} + """ + } + + process Read_Merging_DC { + + label 'norm_cpus' + + tag "${sample_id}" + + publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/ReadMerging/Individual", mode: "copy", overwrite: true, pattern: "*mergedclean.fastq" + publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/ReadMerging/Individual/notmerged", mode: "copy", overwrite: true, pattern: "*notmerged*.fastq" + + input: + tuple sample_id, file(reads) from reads_bbduk_ch + + output: + file("*_mergedclean.fastq") into reads_vsearch1_ch + file("*.name") into names + file("*notmerged*.fastq") into notmerged + + script: + """ + vsearch --fastq_mergepairs ${reads[0]} --reverse ${reads[1]} --threads ${task.cpus} --fastqout ${sample_id}_mergedclean.fastq --fastqout_notmerged_fwd ${sample_id}_notmerged_fwd.fastq --fastqout_notmerged_rev ${sample_id}_notmerged_rev.fastq --fastq_maxee ${params.maxEE} --relabel ${sample_id}. + echo ${sample_id} > ${sample_id}.name + """ + + } + + process Compile_Reads_DC { + + label 'low_cpus' + + publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/ReadMerging/LengthFiltering", mode: "copy", overwrite: true + + input: + file(reads) from reads_vsearch1_ch + .collect() + + output: + file("*_all_merged_preFilt_preClean.fastq") into collect_samples_ch + + script: + """ + cat ${reads} >>${params.projtag}_all_merged_preFilt_preClean.fastq + """ + } - # pcASV_Nucleotide_Phylogeny - if [ "${params.iqCustomnt}" != "" ];then - iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_noTaxonomy_iq --redo -T auto ${params.iqCustomnt} + process Compile_Names_DC { - elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_noTaxonomy_iq -m \${mod} --redo-nt auto -b ${params.boots} + label 'low_cpus' - elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_noTaxonomy_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni + publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/ReadMerging", mode: "copy", overwrite: true - elif [ "${params.nonparametric}" != "false" ];then - iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_noTaxonomy_iq -m MFP --redo -nt auto -b ${params.boots} + input: + file(names) from names + .collect() - elif [ "${params.parametric}" != "false" ];then - iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_noTaxonomy_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + output: + file("*sample_ids.list") into ( samplelist, samplistpotu ) - else - iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_noTaxonomy_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - fi - """ - } - } + script: + """ + cat ${names} >>${params.projtag}_sample_ids.list + """ + } - process pcASV_AminoAcid_Matrix { + process Length_Filtering_DC { - label 'low_cpus' + label 'norm_cpus' - tag "${mtag}" + publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/ReadMerging/LengthFiltering", mode: "copy", overwrite: true, pattern: "*_merged_preFilt*.fasta" + publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/ReadMerging", mode: "copy", overwrite: true, pattern: "*Lengthfiltered.fastq" + publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/ReadMerging/Histograms/pre_length_filtering", mode: "copy", overwrite: true, pattern: "*preFilt_*st.txt" + publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/ReadMerging/Histograms/post_length_filtering", mode: "copy", overwrite: true, pattern: "*postFilt_*st.txt" - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Matrix", mode: "copy", overwrite: true + input: + file(reads) from collect_samples_ch - input: - tuple nid, file(prot) from pcASV_aaMatrix_ch + output: + file("*_merged_preFilt_clean.fasta") into ( nuclCounts_mergedreads_ch, pcASV_mergedreads_ch ) + file("*_merged_clean_Lengthfiltered.fastq") into reads_vsearch2_ch + + file("*preFilt_preClean_baseFrequency_hist.csv") into prefilt_basefreq + file("*preFilt_preClean_qualityScore_hist.csv") into prefilt_qualityscore + file("*preFilt_preClean_gcContent_hist.csv") into prefilt_gccontent + file("*preFilt_preClean_averageQuality_hist.csv") into prefilt_averagequality + file("*preFilt_preClean_length_hist.csv") into prefilt_length + + file("*postFilt_baseFrequency_hist.csv") into postFilt_basefreq + file("*postFilt_qualityScore_hist.csv") into postFilt_qualityscore + file("*postFilt_gcContent_hist.csv") into postFilt_gccontent + file("*postFilt_averageQuaulity_hist.csv") into postFilt_averagequality + file("*postFilt_length_hist.csv") into postFilt_length + + file("reads_per_sample_preFilt_preClean.csv") into reads_per_sample_preFilt + file("read_per_sample_postFilt_postClean.csv") into reads_per_sample_postFilt + script: + """ + bbduk.sh in=${reads} bhist=${params.projtag}_all_merged_preFilt_preClean_baseFrequency_hist.txt qhist=${params.projtag}_all_merged_preFilt_preClean_qualityScore_hist.txt gchist=${params.projtag}_all_merged_preFilt_preClean_gcContent_hist.txt aqhist=${params.projtag}_all_merged_preFilt_preClean_averageQuality_hist.txt lhist=${params.projtag}_all_merged_preFilt_preClean_length_hist.txt gcbins=auto + for x in *preFilt*hist.txt;do + pre=\$(echo \$x | awk -F ".txt" '{print \$1}') + cat \$x | tr "\t" "," > \${pre}.csv + rm \$x + done + reformat.sh in=${reads} out=${params.projtag}_preFilt_preclean.fasta t=${task.cpus} + echo "sample,reads" >> reads_per_sample_preFilt_preClean.csv + grep ">" ${params.projtag}_preFilt_preclean.fasta | awk -F ">" '{print \$2}' | awk -F "." '{print \$1}' | sort --parallel=${task.cpus} | uniq -c | sort -brg --parallel=${task.cpus} | awk '{print \$2","\$1}' >> reads_per_sample_preFilt_preClean.csv + rm ${params.projtag}_preFilt_preclean.fasta + fastp -i ${reads} -o ${params.projtag}_merged_preFilt_clean.fastq -b ${params.maxLen} -l ${params.minLen} --thread ${task.cpus} -n 1 + reformat.sh in=${params.projtag}_merged_preFilt_clean.fastq out=${params.projtag}_merged_preFilt_clean.fasta t=${task.cpus} + bbduk.sh in=${params.projtag}_merged_preFilt_clean.fastq out=${params.projtag}_merged_clean_Lengthfiltered.fastq minlength=${params.maxLen} maxlength=${params.maxLen} t=${task.cpus} + bbduk.sh in=${params.projtag}_merged_clean_Lengthfiltered.fastq bhist=${params.projtag}_all_merged_postFilt_baseFrequency_hist.txt qhist=${params.projtag}_all_merged_postFilt_qualityScore_hist.txt gchist=${params.projtag}_all_merged_postFilt_gcContent_hist.txt aqhist=${params.projtag}_all_merged_postFilt_averageQuaulity_hist.txt lhist=${params.projtag}_all_merged_postFilt_length_hist.txt gcbins=auto + for x in *postFilt*hist.txt;do + pre=\$(echo \$x | awk -F ".txt" '{print \$1}') + cat \$x | tr "\t" "," > \${pre}.csv + rm \$x + done + reformat.sh in=${params.projtag}_merged_clean_Lengthfiltered.fastq out=${params.projtag}_merged_clean_Lengthfiltered.fasta t=${task.cpus} + echo "sample,reads" >> read_per_sample_postFilt_postClean.csv + grep ">" ${params.projtag}_merged_clean_Lengthfiltered.fasta | awk -F ">" '{print \$2}' | awk -F "." '{print \$1}' | sort --parallel=${task.cpus} | uniq -c | sort -brg --parallel=${task.cpus} | awk '{print \$2","\$1}' >> read_per_sample_postFilt_postClean.csv + """ + } - output: - file("*.matrix") into pcASVaaMatrix - tuple nid, file("*PercentID.matrix") into potu_aa_heatmap + process Extract_Uniques_DC { - script: - mtag="ID=" + nid - """ - name=\$( echo ${prot} | awk -F ".fasta" '{print \$1}') - clustalo -i ${prot} --distmat-out=\${name}_PairwiseDistanceq.matrix --full --force --threads=${task.cpus} - clustalo -i ${prot} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} - cat \${name}_PercentIDq.matrix | tr " " "," | grep "," >\${name}_PercentID.matrix - rm \${name}_PercentIDq.matrix - """ - } + label 'norm_cpus' - if (!params.skipEMBOSS) { + publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/ReadMerging/Uniques", mode: "copy", overwrite: true - process pcASV_EMBOSS_Analyses { + input: + file(reads) from reads_vsearch2_ch - label 'low_cpus' + output: + file("*unique_sequences.fasta") into reads_vsearch3_ch - tag "${mtag}" + script: + """ + vsearch --derep_fulllength ${reads} --sizeout --relabel_keep --output ${params.projtag}_unique_sequences.fasta + """ + } - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/2dStructure", mode: "copy", overwrite: true, pattern: '*.{garnier}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/HydrophobicMoment", mode: "copy", overwrite: true, pattern: '*HydrophobicMoments.{svg}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/IsoelectricPoint", mode: "copy", overwrite: true, pattern: '*IsoelectricPoint.{iep,svg}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/ProteinProperties", mode: "copy", overwrite: true, pattern: '*.{pepstats,pepinfo}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/ProteinProperties/Plots", mode: "copy", overwrite: true, pattern: '*PropertiesPlot.{svg}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/2dStructure/Plots", mode: "copy", overwrite: true, pattern: '*Helical*.{svg}' + process Identify_ASVs_DC { - input: - tuple nid, file(prot) from pcASVEMBOSS + label 'norm_cpus' - output: - tuple file("*.garnier"), file("*HydrophobicMoments.svg"), file("*IsoelectricPoint*"), file("*.pepstats"), file("*PropertiesPlot*"), file("*Helical*") into pcASV_emboss + publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/ASVs/ChimeraCheck", mode: "copy", overwrite: true - script: - // check do I need for loop - mtag="ID=" + nid - """ - name=\$( echo ${prot} | awk -F ".fasta" '{print \$1}') - garnier -sequence ${prot} -outfile \${name}_2dStructures.garnier - hmoment -seqall ${prot} -graph svg -plot - mv hmoment.svg ./"\${name}"_HydrophobicMoments.svg - iep -sequence ${prot} -graph svg -plot -outfile "\${name}"_IsoelectricPoint.iep - mv iep.svg ./"\${name}"_IsoelectricPoint.svg - pepstats -sequence ${prot} -outfile \${name}_ProteinProperties.pepstats - grep ">" ${prot} | awk -F ">" '{print \$2}' > tmpsequence.list - for x in \$(cat tmpsequence.list);do - echo \$x > tmp1.list - seqtk subseq ${prot} tmp1.list > tmp2.fasta - len=\$(tail -1 tmp2.fasta | awk '{print length}') - pepinfo -sequence tmp2.fasta -graph svg -outfile "\$x"_PropertiesPlot.pepinfo - mv pepinfo.svg ./"\$x"_PropertiesPlot.svg - cat "\$x"_PropertiesPlot.pepinfo >> "\${name}"_PropertiesPlot.pepinfo - rm "\$x"_PropertiesPlot.pepinfo - pepnet -sask -sequence tmp2.fasta -graph svg -sbegin1 1 -send1 \$len - mv pepnet.svg ./"\$x"_HelicalNet.svg - pepwheel -sequence tmp2.fasta -graph svg -sbegin1 1 -send1 \$len - mv pepwheel.svg ./"\$x"_HelicalWheel.svg - rm tmp1.list tmp2.fasta - done - rm tmpsequence.list - """ - } - } + input: + file(reads) from reads_vsearch3_ch - if (!params.skipTaxonomy) { + output: + file("*notChecked.fasta") into reads_vsearch4_ch - if (params.headers == "RefSeq") { + script: + """ + vsearch --cluster_unoise ${reads} --unoise_alpha ${params.alpha} --relabel ASV --centroids ${params.projtag}_notChecked.fasta --minsize ${params.minSize} + """ + } - process pcASV_AminoAcid_Taxonomy_Inference_RefSeq { + process Chimera_Check_DC { - label 'high_cpus' + label 'norm_cpus' - tag "${mtag}" + publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/ASVs", mode: "copy", overwrite: true - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{csv,tsv}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*dmd.{out}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{fasta}' + input: + file(fasta) from reads_vsearch4_ch - input: - tuple nid, file(asvs) from pcASV_aaDiamond_ch + output: + file("*ASVs.fasta") into ( reads_vsearch5_ch, nucl2aa, asvsforAminotyping, asvfastaforcounts, asvaminocheck ) - output: - file("*.fasta") into ( pcASV_labeledAA ) - tuple file("*phyloformat.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_potuaadiamond - tuple nid, file("*_summary_for_plot.csv") into taxplot4 - tuple nid, file("*_quick_Taxbreakdown.csv") into tax_table_pcasvaa + script: + """ + vsearch --uchime3_denovo ${fasta} --relabel ASV --nonchimeras ${params.projtag}_ASVs.fasta + """ + } - script: - mtag="ID=" + nid - """ - cp ${params.vampdir}/bin/rename_seq.py . - virdb=${params.dbdir}/${params.dbname} - grep ">" \${virdb} > headers.list - headers="headers.list" - name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 - echo "Preparing lists to generate summary .csv's" - echo "[Best hit accession number]" > access.list - echo "[e-value]" > evalue.list - echo "[Bitscore]" > bit.list - echo "[Percent ID (aa)]" > pid.list - echo "[Organism ID]" > "\$name"_virus.list - echo "[Gene]" > "\$name"_genes.list - echo "[pcASV#]" > otu.list - echo "[Sequence length]" > length.list - grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst - echo "extracting genes and names" - touch new_"\$name"_asvnames.txt - for s in \$(cat seqids.lst);do - echo "Checking for \$s hit in diamond output" - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">"\${s}"_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list - virus="NO" - gene="HIT" - echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - echo "\$s done." - fi - done - echo "Now editing "\$name" fasta headers" - ###### rename_seq.py - ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta - awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta - echo "[Sequence header]" > newnames.list - cat new_"\$name"_asvnames.txt >> newnames.list - touch sequence.list - echo " " > sequence.list - grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list - rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - for x in *phyloformat.csv;do - echo "\$x" - lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) - tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; - sed 's/ /_/g' tmpcol.list > tmp2col.list; - cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; - rm tmpcol.list tmp2col.list - done - rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list - """ - } - } else if (params.headers == "RVDB") { - - process pcASV_AminoAcid_Taxonomy_Inference_RVDB { - - label 'high_cpus' - - tag "${mtag}" - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{csv,tsv}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*dmd.{out}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{fasta}' - - input: - tuple nid, file(asvs) from pcASV_aaDiamond_ch - - output: - file("*.fasta") into ( pcASV_labeledAA ) - tuple file("*phyloformat.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_potuaadiamond - tuple nid, file("*_summary_for_plot.csv") into taxplot4 - tuple nid, file("*_quick_Taxbreakdown.csv") into tax_table_pcasvaa - - script: - mtag="ID=" + nid - """ - cp ${params.vampdir}/bin/rename_seq.py . - virdb=${params.dbdir}/${params.dbname} - grep ">" \${virdb} > headers.list - headers="headers.list" - name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 - echo "Preparing lists to generate summary .csv's" - echo "[Best hit accession number]" > access.list - echo "[e-value]" > evalue.list - echo "[Bitscore]" > bit.list - echo "[Percent ID (aa)]" > pid.list - echo "[Organism ID]" > "\$name"_virus.list - echo "[Gene]" > "\$name"_genes.list - echo "[pcASV#]" > otu.list - echo "[Sequence length]" > length.list - grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst - if [[ ${params.lca} == "T" ]] - then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list - echo "[Classification]" > classfication.list - else echo "skipped" >> \${name}_quick_Taxbreakdown.csv - echo "[Classification]" > classfication.list - fi - echo "extracting genes and names" - touch new_"\$name"_asvnames.txt - for s in \$(cat seqids.lst);do - echo "Using RVDB headers." - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - if [[ "${params.lca}" == "T" ]] - then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}') - lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') - echo "\$lcla" >> classfication.list - fi - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list - virus="NO" - gene="HIT" - echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - if [[ "${params.lca}" == "T" ]] - then echo "N/A" >> classfication.list - fi - echo "\$s done." - fi - echo "Done with \$s" - done - echo "Now editing "\$name" fasta headers" - ###### rename_seq.py - ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta - awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta - echo "[Sequence header]" > newnames.list - cat new_"\$name"_asvnames.txt >> newnames.list - touch sequence.list - echo " " > sequence.list - grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list - rm "\$name"_tmpssasv.fasta - if [[ "${params.lca}" == "T" ]] - then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list classfication.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv - else paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - fi - for x in *phyloformat.csv;do - echo "\$x" - lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) - tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; - sed 's/ /_/g' tmpcol.list > tmp2col.list; - cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; - rm tmpcol.list tmp2col.list - done - rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list - """ - } - } - } + process NucleotideBased_ASV_clustering_DC { - if (!params.skipPhylogeny) { + label 'norm_cpus' - process pcASV_Protein_Phylogeny { + publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Nucleotide", mode: "copy", overwrite: true, pattern: '*{.csv}' - label 'norm_cpus' + input: + file(fasta) from reads_vsearch5_ch - tag "${mtag}" + output: + file("number_per_percentage_nucl.csv") into number_per_percent_nucl_plot - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*aln.*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Phylogeny/Modeltest", mode: "copy", overwrite: true, pattern: '*mt*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*iq*' + script: + if (params.datacheckntIDlist) { + """ + for id in `echo ${params.datacheckntIDlist} | tr "," "\\n"`;do + vsearch --cluster_fast ${fasta} --centroids ${params.projtag}_ncASV\${id}.fasta --threads ${task.cpus} --relabel OTU --id \${id} + done + for x in *ncASV*.fasta;do + id=\$( echo \$x | awk -F "_ncASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') + numb=\$( grep -c ">" \$x ) + echo "\${id},\${numb}" >> number_per_percentage_nucl.csv + done + yo=\$(grep -c ">" ${fasta}) + echo "1.0,\${yo}" >> number_per_percentage_nucl.csv + """ + } + } - input: - tuple nid, file(prot) from pcASV_aaMafft_ch - output: - tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into pcASV_protein_phylogeny_results - tuple nid, file("*iq.treefile") into potu_Atree_plot + if (params.sing) { - script: - mtag="ID=" + nid - """ - pre=\$( echo ${prot} | awk -F ".fasta" '{print \$1}' ) - mafft --maxiterate 5000 --auto ${prot} >\${pre}_ALN.fasta - trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html - o-trim-uninformative-columns-from-alignment \${pre}_aln.fasta - mv \${pre}_aln.fasta-TRIMMED ./\${pre}_Aligned_informativeonly.fasta - # pcASV_Protein_ModelTest - modeltest-ng -i \${pre}_Aligned_informativeonly.fasta -p ${task.cpus} -o \${pre}_mt -d aa -s 203 --disable-checkpoint + process Translating_For_ProteinClustering_DC { - # pcASV_Protein_Phylogeny - if [ "${params.iqCustomaa}" != "" ];then - iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomaa} + label 'low_cpus' - elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} + publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Aminoacid/translation", mode: "copy", overwrite: true - elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni + input: + file(fasta) from nucl2aa - elif [ "${params.nonparametric}" != "false" ];then - iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} + output: + file("*ASVprotforclust.fasta") into clustering_aa + file("*_translation_report") into reportaa_VR + file("*_ASV_all.fasta") into asvfastaforaaclust - elif [ "${params.parametric}" != "false" ];then - iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + script: + """ + conda init && source activate virtualribosome - else - iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - fi - """ - } - } + ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_ASVprotforclust.fasta --report ${params.projtag}_translation_report + cp ${fasta} ${params.projtag}_ASV_all.fasta - process Generate_pcASV_Protein_Counts { + """ - label 'high_cpus' + } - tag "${mtag}" + } else { - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Counts", mode: "copy", overwrite: true + process Translation_For_ProteinBased_Clustering_DC { - input: - tuple nid, file(fasta) from pcASV_aaCounts_ch - file(merged) from mergeforpcASVaacounts - file(samplist) from samplistpotu + label 'norm_cpus' - output: - tuple file("*_counts.csv"), file("*dmd.out") into potuaacounts_summary - tuple nid, file("*counts.csv") into potu_Acounts + conda 'python=2.7' - script: - // check do I need for loop - mtag="ID=" + nid - """ - set +e - potu="\$( echo ${fasta} | awk -F "_" '{print \$3}')" - diamond makedb --in ${fasta} --db ${fasta} - diamond blastx -q ${merged} -d ${fasta} -p ${task.cpus} --min-score ${params.ProtCountsBit} --id ${params.ProtCountID} -l ${params.ProtCountsLength} --more-sensitive -o ${params.projtag}_\${potu}_Counts_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 --max-hsps 1 - echo "OTU_ID" >tmp.col1.txt - echo "Generating sample id list" - grep ">" ${fasta} | awk -F ">" '{print \$2}' | sort | uniq > otuid.list - cat otuid.list >> tmp.col1.txt - echo "Beginning them counts tho my g" - for y in \$( cat ${samplist} );do - echo "Starting with \$y now ..." - grep "\$y" ${params.projtag}_\${potu}_Counts_dmd.out > tmp."\$y".out - echo "Isolated hits" - echo "Created uniq subject id list" - echo "\$y" > "\$y"_col.txt - echo "Starting my counts" - for z in \$(cat otuid.list);do - echo "Counting \$z hits" - echo "grep -wc "\$z" >> "\$y"_col.txt" - grep -wc "\$z" tmp."\$y".out >> "\$y"_col.txt - echo "\$z counted" - done - done - paste -d "," tmp.col1.txt *col.txt > ${params.projtag}_aminoacid_\${potu}_noTaxonomy_counts.csv - rm tmp* - rm *col.txt - """ - } - } + publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Aminoacid/translation", mode: "copy", overwrite: true - if (!params.skipReport) { + input: + file(fasta) from nucl2aa - if (!params.skipAdapterRemoval || !params.skipReadProcessing || !params.skipMerging) { + output: + file("*ASVprotforclust.fasta") into clustering_aa + file("*_translation_report") into reportaa_VR + file("*_ASV_all.fasta") into asvfastaforaaclust - process combine_csv { + script: + """ + ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_ASVprotforclust.fasta --report ${params.projtag}_translation_report + cp ${fasta} ${params.projtag}_ASV_all.fasta + """ + } + } - input: - file(csv) from fastp_csv_in2 - .collect() + process Protein_clustering_DC { - output: - file("final_reads_stats.csv") into fastp_csv_in + label 'norm_cpus' - script: - """ - cat ${csv} >all_reads_stats.csv - head -n1 all_reads_stats.csv >tmp.names.csv - cat all_reads_stats.csv | grep -v ""Sample,Total_"" >tmp.reads.stats.csv - cat tmp.names.csv tmp.reads.stats.csv >final_reads_stats.csv - rm tmp.names.csv tmp.reads.stats.csv - """ - } - } else { + publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Aminoacid", mode: "copy", overwrite: true, pattern: '*{.csv}' - process skip_combine_csv { + input: + file(fasta) from clustering_aa + file(asvs) from asvfastaforaaclust - output: - file("filter_reads.txt") into fastp_csv_in + output: + file("number_per_percentage_prot.csv") into number_per_percent_prot_plot - script: - """ - echo "Read processing steps skipped." >filter_reads.txt - """ - } - } + script: + // add awk script to count seqs + """ + set +e + cp ${params.vampdir}/bin/rename_seq.py . + for id in `echo ${params.datacheckaaIDlist} | tr "," "\\n"`;do + if [ \${id} == ".55" ];then + word=3 + elif [ \${id} == ".65" ];then + word=4 + else + word=5 + fi + awk 'BEGIN{RS=">";ORS=""}length(\$2)>="${params.minAA}"{print ">"\$0}' ${fasta} > ${params.projtag}_filtered_proteins.fasta + cd-hit -i ${params.projtag}_filtered_proteins.fasta -n \${word} -c \${id} -o ${params.projtag}_pcASV\${id}.fasta + sed 's/>Cluster />Cluster_/g' ${params.projtag}_pcASV\${id}.fasta.clstr >${params.projtag}_pcASV\${id}.clstr + grep ">Cluster_" ${params.projtag}_pcASV\${id}.clstr >temporaryclusters.list + y=\$(grep -c ">Cluster_" ${params.projtag}_pcASV\${id}.clstr) + echo ">Cluster_"\${y}"" >> ${params.projtag}_pcASV\${id}.clstr + t=1 + b=1 + for x in \$(cat temporaryclusters.list);do + echo "Extracting \$x" + name="\$( echo \$x | awk -F ">" '{print \$2}')" + clust="pcASV"\${t}"" + echo "\${name}" + awk '/^>'\${name}'\$/,/^>Cluster_'\${b}'\$/' ${params.projtag}_pcASV\${id}.clstr > "\${name}"_"\${clust}"_tmp.list + t=\$(( \${t}+1 )) + b=\$(( \${b}+1 )) + done + ls *_tmp.list + u=1 + for x in *_tmp.list;do + name="\$(echo \$x | awk -F "_p" '{print \$1}')" + echo "\${name}" + cluster="\$(echo \$x | awk -F "_" '{print \$3}')" + echo "\${cluster}" + grep "ASV" \$x | awk -F ", " '{print \$2}' | awk -F "_" '{print \$1}' | awk -F ">" '{print \$2}' > \${name}_\${cluster}_seqs_tmps.list + seqtk subseq ${asvs} \${name}_\${cluster}_seqs_tmps.list > \${name}_\${cluster}_nucleotide_sequences.fasta + vsearch --cluster_fast \${name}_\${cluster}_nucleotide_sequences.fasta --id 0.2 --centroids \${name}_\${cluster}_centroids.fasta + grep ">" \${name}_\${cluster}_centroids.fasta >> \${name}_\${cluster}_tmp_centroids.list + for y in \$( cat \${name}_\${cluster}_tmp_centroids.list );do + echo ">\${cluster}_type"\$u"" >> \${name}_\${cluster}_tmp_centroid.newheaders + u=\$(( \${u}+1 )) + done + u=1 + ./rename_seq.py \${name}_\${cluster}_centroids.fasta \${name}_\${cluster}_tmp_centroid.newheaders \${cluster}_types_labeled.fasta + done + cat *_types_labeled.fasta >> ${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta + grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$3}' | awk -F "." '{print \$1}' >tmphead.list + grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$2}' | awk -F "," '{print \$1}' >tmplen.list + paste -d"," temporaryclusters.list tmphead.list >tmp.info.csv + grep ">" ${params.projtag}_pcASV\${id}.fasta >lala.list + j=1 + for x in \$(cat lala.list);do + echo ">${params.projtag}_pcASV\${j}" >>${params.projtag}_aminoheaders.list + echo "\${x},>${params.projtag}_pcASV\${j}" >>tmpaminotype.info.csv + j=\$(( \${j}+1 )) + done + rm lala.list + awk -F "," '{print \$2}' tmp.info.csv >>tmporder.list + for x in \$(cat tmporder.list);do + grep -w "\$x" tmpaminotype.info.csv | awk -F "," '{print \$2}' >>tmpder.list + done + paste -d "," temporaryclusters.list tmplen.list tmphead.list tmpder.list >${params.projtag}_pcASVCluster\${id}_summary.csv + ./rename_seq.py ${params.projtag}_pcASV\${id}.fasta ${params.projtag}_aminoheaders.list ${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta + stats.sh in=${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_aminoacid_clustered.gc gcformat=4 overwrite=true + stats.sh in=${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_nucleotide_clustered.gc gcformat=4 overwrite=true + awk 'BEGIN{RS=">";ORS=""}length(\$2)<"${params.minAA}"{print ">"\$0}' ${fasta} >${params.projtag}_pcASV\${id}_problematic_translations.fasta + if [ `wc -l ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk '{print \$1}'` -gt 1 ];then + grep ">" ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk -F ">" '{print \$2}' > problem_tmp.list + seqtk subseq ${asvs} problem_tmp.list > ${params.projtag}_pcASV\${id}_problematic_nucleotides.fasta + else + rm ${params.projtag}_pcASV\${id}_problematic_translations.fasta + fi + rm *.list + rm Cluster* + rm *types* + rm *tmp* + rm ${params.projtag}_pcASV\${id}.fast* + done + for x in *aminoacid*noTaxonomy.fasta;do + id=\$( echo \$x | awk -F "_noTax" '{print \$1}' | awk -F "pcASV" '{print \$2}') + numb=\$( grep -c ">" \$x) + echo "\${id},\${numb}" >> number_per_percentage_protz.csv + done + yesirr=\$( wc -l number_per_percentage_protz.csv | awk '{print \$1}') + tail -\$(( \${yesirr}-1 )) number_per_percentage_protz.csv > number_per_percentage_prot.csv + head -1 number_per_percentage_protz.csv >> number_per_percentage_prot.csv + rm number_per_percentage_protz.csv + """ + } - //NEW REPORT !!!!!!!!!!!!!!!!! - /*Report_ASV - asv_counts_plots -> ${params.projtag}_ASV_counts.csv - taxplot1 -> ${params.projtag}_ASV_summary_for_plot.csv - asv_heatmap -> ${params.projtag}_ASV_PercentID.matrix - nucl_phyl_plot -> ${params.projtag}_ASV_iq.treefile - file("*_ASV_Grouping.csv") into asvgroupscsv - "${params.projtag}_ASV_Groupingcounts.csv") into asvgroupcounts - *_quick_Taxbreakdown.csv") into tax_table_asv - \\${params.projtag}_ASV_Group_Reps_iq.treefile - */ - - report_asv = Channel.create() - asv_counts_plots.mix(taxplot_asv, asv_heatmap, nucl_phyl_plot_asv, asvgroupscsv, asvgroupcounts, asv_group_rep_tree, tax_table_asv).flatten().buffer(size:8).dump(tag:'asv').into(report_asv) - - if (params.ncASV) { - report_ncasv = Channel.create() - notu_counts_plots.mix(taxplot_ncasv, notu_heatmap, nucl_phyl_plot_ncasv, tax_table_ncasv).groupTuple(by:0, size:5).dump(tag:'ncasv').into(report_ncasv) - /* - notu_counts_plots -> ${params.projtag}_ncASV${id}_counts.csv - taxplot1a -> ${params.projtag}_ncASV${id}_summary_for_plot.csv - notu_heatmap -> ${params.projtag}_ncASV${id}_PercentID.matrix - nucl_phyl_plot -> ${params.projtag}_ncASV${id}_iq.treefile - ${params.projtag}_ncASV${id}_quick_Taxbreakdown.csv - */ - } else { - report_ncasv = Channel.empty() - } + process combine_csv_DC { - if (params.pcASV) { - report_pcasv_aa = Channel.create() - potu_Acounts.mix(taxplot4, potu_aa_heatmap, potu_Atree_plot, tax_table_pcasvaa).groupTuple(by:0, size:5).dump(tag:'pcasv1').into(report_pcasv_aa) - /*Report_pcASV_AminoAcid - potu_Acounts -> ${params.projtag}_pcASV${id}_noTaxonomy_counts.csv - taxplot4 -> ${params.projtag}_aminoacid_pcASV${id}_noTaxonomy_summary_for_plot.csv - potu_aa_heatmap -> ${params.projtag}_aminoacid_pcASV${id}_noTaxonomy_PercentID.matrix - potu_Atree_plot -> ${params.projtag}_aminoacid_pcASV${id}_noTaxonomy_iq.treefile - tax_table_pcasvaa -> ${params.projtag}_aminoacid_pcASV${id}_quick_Taxbreakdown.csv - */ - report_pcasv_nucl = Channel.create() - potu_Ncounts_for_report.mix(taxplot3, potu_nucl_heatmap, potu_Ntree_plot, tax_table_pcasvnt).groupTuple(by:0, size:5).dump(tag:'pcasv2').into(report_pcasv_nucl) - /*Report_pcASV_Nucleotide - potu_Ncounts_for_report -> ${params.projtag}_nucleotide_pcASV${id}_noTaxonomy_counts.csv - taxplot3 -> ${params.projtag}_nucleotide_pcASV${id}_noTaxonomy_summary_for_plot.csv - potu_nucl_heatmap -> ${params.projtag}_nucleotide_pcASV${id}_noTaxonomy_PercentID.matrix - potu_Ntree_plot -> ${params.projtag}_nucleotide_pcASV${id}_noTaxonomy_iq.treefile - tax_table_pcasvnt -> ${params.projtag}_nucleotide_pcASV${id}_quick_Taxbreakdown.csv - */ - } else { - report_pcasv_aa = Channel.empty() - report_pcasv_nucl = Channel.empty() - } + input: + file(csv) from fastp_csv + .collect() - if (!params.skipAminoTyping) { - report_aminotypes = Channel.create() - aminocounts_plot.mix(taxplot2, aminotype_heatmap, amino_rax_plot, atygroupscsv, amino_group_rep_tree, amino_groupcounts, tax_table_amino).flatten().buffer(size:8).dump(tag:'amino').into(report_aminotypes) - /* - Report_AminoTypes - aminocounts_plot -> ${params.projtag}_AminoType_counts.csv - taxplot2 -> ${params.projtag}_AminoTypes_summary_for_plot.csv - aminotype_heatmap -> ${params.projtag}_AminoTypes_PercentID.matrix - amino_rax_plot -> ${params.projtag}_AminoTypes_iq.treefile - atygroupscsv -> *_AminoType_Grouping.csv - amino_group_rep_tree -> ${params.projtag}_AminoType_Group_Reps_iq.treefile - params.projtag}_AminoType_Groupingcounts.csv") into amino_groupcounts - *_quick_Taxbreakdown.csv") into tax_table_amino - */ - } else { - report_aminotypes = Channel.empty() - } + output: + file("final_reads_stats.csv") into fastp_csv1 - report_all_ch = Channel.create() - report_asv.mix(report_ncasv, report_pcasv_aa, report_pcasv_nucl, report_aminotypes).map{it.flatten()}.dump(tag:'report').into(report_all_ch) + script: + """ + cat ${csv} >all_reads_stats.csv + head -n1 all_reads_stats.csv >tmp.names.csv + cat all_reads_stats.csv | grep -v ""Sample,Total_"" >tmp.reads.stats.csv + cat tmp.names.csv tmp.reads.stats.csv >final_reads_stats.csv + rm tmp.names.csv tmp.reads.stats.csv + """ - process Report { + } - label 'norm_cpus' + process Report_DataCheck { - publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReports", mode: "copy", overwrite: true + label 'norm_cpus' - input: - file(csv) from fastp_csv_in - file(files) from report_all_ch + publishDir "${params.workingdir}/${params.outdir}/DataCheck/Report", mode: "copy", overwrite: true, pattern: '*.{html}' - output: - file("*.html") into report_all_out + input: + file(fastpcsv) from fastp_csv1 + file(reads_per_sample_preFilt) from reads_per_sample_preFilt + file(read_per_sample_postFilt) from reads_per_sample_postFilt + file(preFilt_baseFrequency) from prefilt_basefreq + file(postFilt_baseFrequency) from postFilt_basefreq + file(preFilt_qualityScore) from prefilt_qualityscore + file(postFilt_qualityScore) from postFilt_qualityscore + file(preFilt_gcContent) from prefilt_gccontent + file(postFilt_gcContent) from postFilt_gccontent + file(preFilt_averageQuality) from prefilt_averagequality + file(postFilt_averageQuaulity) from postFilt_averagequality + file(preFilt_length) from prefilt_length + file(postFilt_length) from postFilt_length + file(number_per_percentage_nucl) from number_per_percent_nucl_plot + file(number_per_percentage_prot) from number_per_percent_prot_plot - script: - """ - name=\$( ls *summary_for_plot.csv | awk -F "_summary_for_plot.csv" '{print \$1}') - cp ${params.vampdir}/bin/vAMPirus_Report.Rmd . - cp ${params.vampdir}/example_data/conf/vamplogo.png . - Rscript -e "rmarkdown::render('vAMPirus_Report.Rmd',output_file='\${name}_Report.html')" \${name} \ - ${params.skipReadProcessing} \ - ${params.skipMerging} \ - ${params.skipAdapterRemoval} \ - ${params.skipTaxonomy} \ - ${params.skipPhylogeny} \ - ${params.trymax} \ - ${params.stats} \ - ${params.metadata} \ - ${params.minimumCounts} - """ - } + output: + file("*.html") into datacheckreport - } - } + script: + """ + cp ${params.vampdir}/bin/vAMPirus_DC_Report.Rmd . + cp ${params.vampdir}/example_data/conf/vamplogo.png . + Rscript -e "rmarkdown::render('vAMPirus_DC_Report.Rmd',output_file='${params.projtag}_DataCheck_Report.html')" ${params.projtag} \ + ${fastpcsv} \ + ${reads_per_sample_preFilt} \ + ${read_per_sample_postFilt} \ + ${preFilt_baseFrequency} \ + ${postFilt_baseFrequency} \ + ${preFilt_qualityScore} \ + ${postFilt_qualityScore} \ + ${preFilt_averageQuality} \ + ${postFilt_averageQuaulity} \ + ${preFilt_length} \ + ${postFilt_length} \ + ${number_per_percentage_nucl} \ + ${number_per_percentage_prot} + """ + } } else { println("\n\t\033[0;31mMandatory argument not specified. For more info use `nextflow run vAMPirus.nf --help`\n\033[0m") exit 0 } -if (params.DataCheck) { - workflow.onComplete { - log.info ( workflow.success ? \ - "---------------------------------------------------------------------------------" \ - + "\n\033[0;32mDone! Open the following reports in your browser\033[0m" \ - + "\n\033[0;32mPipeline performance report: ${params.workingdir}/${params.outdir}/${params.tracedir}/vampirus_report.html\033[0m" \ - + "\n\033[0;32mvAMPirus --DataCheck interactive report: ${params.workingdir}/${params.outdir}/DataCheck/Report/*.hmtl\033[0m" \ - : \ - "---------------------------------------------------------------------------------" \ - + "\n\033[0;31mSomething went wrong. Check error message below and/or log files.\033[0m" ) - } -} else if (params.Analyze) { - workflow.onComplete { - log.info ( workflow.success ? \ - "---------------------------------------------------------------------------------" \ - + "\n\033[0;32mDone! Open the following reports in your browser\033[0m" \ - + "\n\033[0;32mPipeline performance report: ${params.workingdir}/${params.outdir}/${params.tracedir}/vampirus_report.html\033[0m" \ - + "\n\033[0;32mvAMPirus --Analyze interactive report: ${params.workingdir}/${params.outdir}/Analyze/*.hmtl\033[0m" \ - : \ - "---------------------------------------------------------------------------------" \ - + "\n\033[0;31mSomething went wrong. Check error message below and/or log files.\033[0m" ) - } + +workflow.onComplete { + log.info ( workflow.success ? \ + "---------------------------------------------------------------------------------" \ + + "\n\033[0;32mDone! Open the following reports in your browser\033[0m" \ + + "\n\033[0;32mPipeline performance report: ${params.workingdir}/${params.outdir}/${params.tracedir}/vampirus_report.html\033[0m" \ + + "\n\033[0;32mvAMPirus --DataCheck interactive report: ${params.workingdir}/${params.outdir}/DataCheck/*.hmtl\033[0m" \ + + "\n\033[0;32mvAMPirus --Analyze interactive report: ${params.workingdir}/${params.outdir}/Analyze/*.hmtl\033[0m" \ + : \ + "---------------------------------------------------------------------------------" \ + + "\n\033[0;31mSomething went wrong. Check error message below and/or log files.\033[0m" ) } diff --git a/vampirus.config b/vampirus.config index a987467..c1610aa 100644 --- a/vampirus.config +++ b/vampirus.config @@ -106,7 +106,7 @@ params { // Set minimum bitscore for best hit in taxonomy assignment bitscore="50" // Set minimum percent amino acid similarity for best hit to be counted in taxonomy assignment - minID="80" + minID="40" // Set minimum amino acid alignment length for best hit to be counted in taxonomy assignment minaln="30" From 48a923dd498779fcfb45b126dbaeb7832190aeb8 Mon Sep 17 00:00:00 2001 From: Ramon Rivera Date: Mon, 3 May 2021 15:37:24 -0700 Subject: [PATCH 109/223] Update vAMPirus.nf --- vAMPirus.nf | 5402 ++++++++++++++++++++++++++------------------------- 1 file changed, 2715 insertions(+), 2687 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 4542133..3af44b5 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -33,7 +33,7 @@ def helpMessage() { Usage: - nextflow run vAMPirus.nf -c vampirus.config -profile [conda|singularity] --[Analyze|DataCheck] [--ncASV] [--pcASV] + nextflow run vAMPirus.nf -c vampirus.config -profile [conda|singularity] --[Analyze|DataCheck] [--ncASV] [--pcASV] [--asvMED] [--aminoMED] [--stats] --Help options-- @@ -57,6 +57,12 @@ def helpMessage() { --pcASV Set this option to have vAMPirus cluster nucleotide and translated ASVs into protein-based operational taxonomic units (pcASVs) - See options below to define a single percent similarity or a list + --Minimum Entropy Decomposition arguments-- + + --asvMED Set this option to perform Minimum Entropy Decomposition on ASV sequences, see manual for more information. You will need to set a value for --asvC to perform this analysis + + --amino_med Set this option to perform Minimum Entropy Decomposition on AminoType sequences, see manual for more information. You will need to set a value for --aminoC to perform this analysis + --Skip options-- --skipReadProcessing Set this option to skip all read processing steps in the pipeline @@ -65,7 +71,9 @@ def helpMessage() { --skipAdapterRemoval Set this option to skip adapter removal in the pipeline - --skipPrimerRemoval Set this option to skup Skip primer removal process + --skipPrimerRemoval Set this option to skip primer removal process + + --skipMerging Set this option to skip read merging --skipAminoTyping Set this option to skip AminoTyping processes @@ -73,6 +81,10 @@ def helpMessage() { --skipPhylogeny Set this option to skip phylogeny processes + --skipEMBOSS Set this option to skip EMBOSS processes + + --skipReport Set this option to skip html report generation + **NOTE** Most opitons below can be set using the configuration file (vampirus.config) to avoid a lengthy launch command. --Project/analysis information-- @@ -140,6 +152,11 @@ def helpMessage() { --minAA With --pcASV set, use this option to set the expected or minimum amino acid sequence length of open reading frames within your amplicon sequences + --Minimum Entropy Decomposition-- + + --asvC Number of high entropy positions to use for ASV MED analysis and generate "Groups" + + --aminoC Number of high entropy positions to use for AminoType MED analysis and generate "Groups" --Counts table generation-- @@ -158,7 +175,11 @@ def helpMessage() { --dbdir Path to Directory where database is being stored - --refseq Set "--refseq T" to toggle use of RefSeq header format; default is "F" to use Reverence Viral DataBase (RVDB) header + --headers Set taxonomy database header format -> headers= "RefSeq" to toggle use of RefSeq header format; set to "RVDB" to signal the use of Reverence Viral DataBase (RVDB) headers + + --dbanno Path to directory hmm annotation .txt file - see manual for information on this. Leave as is if not planning on using. + + --lca Set --lca T if you would like to add taxonomic classification to taxonomy results - example: "ASV1, Viruses::Duplodnaviria::Heunggongvirae::Peploviricota::Herviviricetes::Herpesvirales::Herpesviridae::Gammaherpesvirinae::Macavirus" --bitscore Set minimum bitscore to allow for best hit in taxonomy assignment @@ -205,186 +226,208 @@ def fullHelpMessage() { THIS IS A LONGER HELP WITH USAGE EXAMPLES vAMPirus v${workflow.manifest.version} ============================================================================================================================================================================================== - Steps: - 1- Before launching the vAMPirus.nf, be sure to run the vampirus_startup.sh script to install dependencies and/or databases + Steps: + 1- Before launching the vAMPirus.nf, be sure to run the vampirus_startup.sh script to install dependencies and/or databases + + 2- Test the vAMPirus installation with the provided test dataset (if you have ran the start up script, you can see STARTUP_HELP.txt for test commands and other examples) + + 3. Edit parameters in vampirus.config file + + 4. Launch the DataCheck pipeline to get summary information about your dataset + + 5. Change any parameters in vampirus.config file that might aid your analysis (e.g. clustering ID, maximum merged read length) + + 6. Launch the Analyze pipeline to perform a comprehensive analysis with your dataset + + 7. Explore results directories and produced final reports - 2- Test the vAMPirus installation with the provided test dataset (if you have ran the start up script, you can see STARTUP_HELP.txt for test commands and other examples) - 3. Edit parameters in vampirus.config file + Usage: - 4. Launch the DataCheck pipeline to get summary information about your dataset + nextflow run vAMPirus.nf -c vampirus.config -profile [conda|singularity] --[Analyze|DataCheck] [--ncASV] [--pcASV] - 5. Change any parameters in vampirus.config file that might aid your analysis (e.g. clustering ID, maximum merged read length) - 6. Launch the Analyze pipeline to perform a comprehensive analysis with your dataset + --Help options-- - 7. Explore results directories and produced final reports + --help Print help information + --fullHelp Print even more help information - Usage: - nextflow run vAMPirus.nf -c vampirus.config -profile [conda|singularity] --[Analyze|DataCheck] [--ncASV] [--pcASV] + --Mandatory arguments (choose one)-- + --Analyze Run absolutely everything - --Help options-- + --DataCheck Assess how data performs with during processing and clustering - --help Print help information - --fullHelp Print even more help information + --ASV clustering arguments-- + --ncASV Set this option to have vAMPirus cluster nucleotide amplicon sequence variants (ASVs) into nucleotide-based operational taxonomic units (ncASVs) - See options below to define a single percent similarity or a list - --Mandatory arguments (choose one)-- + --pcASV Set this option to have vAMPirus cluster nucleotide and translated ASVs into protein-based operational taxonomic units (pcASVs) - See options below to define a single percent similarity or a list - --Analyze Run absolutely everything - --DataCheck Assess how data performs with during processing and clustering + --Minimum Entropy Decomposition arguments-- + --asvMED Set this option to perform Minimum Entropy Decomposition on ASV sequences, see manual for more information. You will need to set a value for --asvC to perform this analysis - --ASV clustering arguments-- + --amino_med Set this option to perform Minimum Entropy Decomposition on AminoType sequences, see manual for more information. You will need to set a value for --aminoC to perform this analysis - --ncASV Set this option to have vAMPirus cluster nucleotide amplicon sequence variants (ASVs) into nucleotide-based operational taxonomic units (ncASVs) - See options below to define a single percent similarity or a list + --Skip options-- - --pcASV Set this option to have vAMPirus cluster nucleotide and translated ASVs into protein-based operational taxonomic units (pcASVs) - See options below to define a single percent similarity or a list + --skipReadProcessing Set this option to skip all read processing steps in the pipeline + --skipFastQC Set this option to skiip FastQC steps in the pipeline - --Skip options-- + --skipAdapterRemoval Set this option to skip adapter removal in the pipeline - --skipReadProcessing Set this option to skip all read processing steps in the pipeline + --skipPrimerRemoval Set this option to skip primer removal process - --skipFastQC Set this option to skiip FastQC steps in the pipeline + --skipMerging Set this option to skip read merging - --skipAdapterRemoval Set this option to skip adapter removal in the pipeline + --skipAminoTyping Set this option to skip AminoTyping processes - --skipPrimerRemoval Set this option to skup Skip primer removal process + --skipTaxonomy Set this option to skip taxonomy assignment processes - --skipAminoTyping Set this option to skip AminoTyping processes + --skipPhylogeny Set this option to skip phylogeny processes - --skipTaxonomy Set this option to skip taxonomy assignment processes + --skipEMBOSS Set this option to skip EMBOSS processes - --skipPhylogeny Set this option to skip phylogeny processes + --skipReport Set this option to skip html report generation - **NOTE** Most opitons below can be set using the configuration file (vampirus.config) to avoid a lengthy launch command. - --Project/analysis information-- + **NOTE** Most opitons below can be set using the configuration file (vampirus.config) to avoid a lengthy launch command. - --projtag Set project name to be used as a prefix for output files + --Project/analysis information-- - --metadata Set path to metadata spreadsheet file to be used for report generation (must be defined if generating report) + --projtag Set project name to be used as a prefix for output files - --reads Path to directory containing read libraries, must have *R{1,2}* in the library names + --metadata Set path to metadata spreadsheet file to be used for report generation (must be defined if generating report) - --workingdir Path to working directory where Nextflow will put all Nextflow and vAMPirus generated output files + --reads Path to directory containing read libraries, must have *R{1,2}* in the library names - --outdir Name of results directory containing all output from the chosen pipeline (will be made within the working directory) + --workingdir Path to working directory where Nextflow will put all Nextflow and vAMPirus generated output files + --outdir Name of results directory containing all output from the chosen pipeline (will be made within the working directory) - --Merged read length filtering-- - --minLen Minimum merged read length - reads below the specified maximum read length will be used for counts only + --Merged read length filtering-- - --maxLen Maximum merged read length - reads with length equal to the specified max read length will be used to identifying unique sequences and subsequent Amplicon Sequence Variant (ASV) analysis + --minLen Minimum merged read length - reads below the specified maximum read length will be used for counts only - --maxEE Use this option to set the maximum expected error rate for vsearch merging. Default is 1. + --maxLen Maximum merged read length - reads with length equal to the specified max read length will be used to identifying unique sequences and subsequent Amplicon Sequence Variant (ASV) analysis + --maxEE Use this option to set the maximum expected error rate for vsearch merging. Default is 1. - --Primer removal-- - General primer removal parameters + --Primer removal-- - --primerLength Use this option to set the max primer length to restrict bbduk.sh primer trimming to the first x number of bases + General primer removal parameters - --maxkmer Maximum kmer length for bbduk.sh to use for primer detection and removal (must be shorter than your primer length; default = 13) + --primerLength Use this option to set the max primer length to restrict bbduk.sh primer trimming to the first x number of bases - --minkmer Minimum kmer length for primer removal (default = 3) + --maxkmer Maximum kmer length for bbduk.sh to use for primer detection and removal (must be shorter than your primer length; default = 13) - --minilen Minimum read length after adapter and primer removal (default = 200) + --minkmer Minimum kmer length for primer removal (default = 3) - Single primer set removal- + --minilen Minimum read length after adapter and primer removal (default = 200) - --GlobTrim Set this option to perform global trimming to reads to remove primer sequences. Example usage "--GlobTrim #basesfromforward,#basesfromreverse" + Single primer set removal- - --fwd Forward primer sequence for reads to be detected and removed from reads (must specify reverse sequence if providing forward) + --GlobTrim Set this option to perform global trimming to reads to remove primer sequences. Example usage "--GlobTrim #basesfromforward,#basesfromreverse" - --rev Reverse primer sequence for reads to be detected and removed from reads (must specify forward sequence if providing reverse) + --fwd Forward primer sequence for reads to be detected and removed from reads (must specify reverse sequence if providing forward) - Multiple primer set removal- + --rev Reverse primer sequence for reads to be detected and removed from reads (must specify forward sequence if providing reverse) - --multi Use this option to signal multiple primer sequence removal within the specified pipeline + Multiple primer set removal- - --primers Use this option to set the path to a fasta file with all of the primer sequences to be detected and removed from reads + --multi Use this option to signal multiple primer sequence removal within the specified pipeline + --primers Use this option to set the path to a fasta file with all of the primer sequences to be detected and removed from reads - --Amplicon Sequence Variant (ASV) genration and clustering-- - --alpha Alpha value for denoising - the higher the alpha the higher the chance of false positives in ASV generation (1 or 2) + --Amplicon Sequence Variant (ASV) genration and clustering-- - --minSize Minimum size or representation for sequence to be considered in ASV generation + --alpha Alpha value for denoising - the higher the alpha the higher the chance of false positives in ASV generation (1 or 2) - --clusterNuclID With --ncASV set, use this option to set a single percent similarity to cluster nucleotide ASV sequences into ncASVs by [ Example: --clusterNuclID .97 ] + --minSize Minimum size or representation for sequence to be considered in ASV generation - --clusterNuclIDlist With --ncASV set, use this option to perform nucleotide-based clustering of ASVs with a comma separated list of percent similarities [ Example: --clusterNuclIDlist .95,.96,.97,.98 ] + --clusterNuclID With --ncASV set, use this option to set a single percent similarity to cluster nucleotide ASV sequences into ncASVs by [ Example: --clusterNuclID .97 ] - --clusterAAID With --pcASV set, use this option to set a single percent similarity for protein-based ASV clustering to generate pcASVs[ Example: --clusterAAID .97 ] + --clusterNuclIDlist With --ncASV set, use this option to perform nucleotide clustering with a comma separated list of percent similarities [ Example: --clusterNuclIDlist .95,.96,.97,.98 ] - --clusterAAIDlist With --pcASV set, use this option to perform protein-based ASV clustering to generate pcASVs with a comma separated list of percent similarities [ Example: --clusterAAIDlist .95,.96,.97,.98 ] + --clusterAAID With --pcASV set, use this option to set a single percent similarity for protein-based ASV clustering to generation pcASVs [ Example: --clusterAAID .97 ] - --minAA With --pcASV set, use this option to set the expected or minimum amino acid sequence length of open reading frames within your amplicon sequences + --clusterAAIDlist With --pcASV set, use this option to perform protein-based ASV clustering to generate pcASVs with a comma separated list of percent similarities [ Example: --clusterAAIDlist .95,.96,.97,.98 ] + --minAA With --pcASV set, use this option to set the expected or minimum amino acid sequence length of open reading frames within your amplicon sequences - --Counts table generation-- + --Minimum Entropy Decomposition-- - --asvcountID Similarity ID to use for ASV counts + --asvC Number of high entropy positions to use for ASV MED analysis and generate "Groups" - --ProtCountID Minimum amino acid sequence similarity for hit to count + --aminoC Number of high entropy positions to use for AminoType MED analysis and generate "Groups" - --ProtCountsLength Minimum alignment length for hit to count + --Counts table generation-- - --ProtCountsBit Minimum bitscore for hit to be counted + --asvcountID Similarity ID to use for ASV counts + --ProtCountID Minimum amino acid sequence similarity for hit to count - --Taxonomy inference parameters-- + --ProtCountsLength Minimum alignment length for hit to count - --dbname Specify name of database to use for analysis + --ProtCountsBit Minimum bitscore for hit to be counted - --dbdir Path to Directory where database is being stored - --refseq Set "--refseq T" to toggle use of RefSeq header format; default is "F" to use Reverence Viral DataBase (RVDB) header + --Taxonomy inference parameters-- - --bitscore Set minimum bitscore to allow for best hit in taxonomy assignment + --dbname Specify name of database to use for analysis - --minID Set minimum percent amino acid similarity for best hit to be counted in taxonomy assignment + --dbdir Path to Directory where database is being stored - --minaln Set minimum amino acid alignment length for best hit to be counted in taxonomy assignment + --headers Set taxonomy database header format -> headers= "RefSeq" to toggle use of RefSeq header format; set to "RVDB" to signal the use of Reverence Viral DataBase (RVDB) headers + --dbanno Path to directory hmm annotation .txt file - see manual for information on this. Leave as is if not planning on using. - --Phylogeny analysis parameters-- + --lca Set --lca T if you would like to add taxonomic classification to taxonomy results - example: "ASV1, Viruses::Duplodnaviria::Heunggongvirae::Peploviricota::Herviviricetes::Herpesvirales::Herpesviridae::Gammaherpesvirinae::Macavirus" - Setting customs options for IQ-TREE (Example: "-option1 A -option2 B -option3 C -option4 D") - might be easier to set in the vampirus.config file at lines 108/109 + --bitscore Set minimum bitscore to allow for best hit in taxonomy assignment - --iqCustomnt Use option to set custom options to use in all IQTREE analyses with nuceoltide sequences + --minID Set minimum percent amino acid similarity for best hit to be counted in taxonomy assignment - --iqCustomaa Use option to set custom options to use in all IQTREE analyses with amino acid sequences + --minaln Set minimum amino acid alignment length for best hit to be counted in taxonomy assignment - These options below you can set at the command, for example, to set to use model from ModelTest-NG with parametric bootstrapping --ModelTnt --ModelTaa --parametric - --ModelTnt=false Signal for IQ-TREE to use model determined by ModelTest-NG for all IQTREE analyses with nuceoltide sequences (Default is IQ-TREE will do automatic model testing with ModelFinder Plus) + --Phylogeny analysis parameters-- - --ModelTaa=false Signal for IQ-TREE to use model determined by ModelTest-NG for all IQTREE analyses with amino acid sequences + Setting customs options for IQ-TREE (Example: "-option1 A -option2 B -option3 C -option4 D") - might be easier to set in the vampirus.config file at lines 108/109 - --parametric Set to use parametric bootstrapping in IQTREE analyses + --iqCustomnt Use option to set custom options to use in all IQTREE analyses with nuceoltide sequences - --nonparametric Set to use parametric bootstrapping in IQTREE analyses + --iqCustomaa Use option to set custom options to use in all IQTREE analyses with amino acid sequences - --boots Number of bootstraps (recommended 1000 for parametric and 100 for non-parametric) + These options below you can set at the command, for example, to set to use model from ModelTest-NG with parametric bootstrapping --ModelTnt --ModelTaa --parametric + --ModelTnt=false Signal for IQ-TREE to use model determined by ModelTest-NG for all IQTREE analyses with nuceoltide sequences (Default is IQ-TREE will do automatic model testing with ModelFinder Plus) - --Statistics options-- + --ModelTaa=false Signal for IQ-TREE to use model determined by ModelTest-NG for all IQTREE analyses with amino acid sequences - --stats Set "--stats run" to signal statstical tests to be performed and included in the final report + --parametric Set to use parametric bootstrapping in IQTREE analyses - --minimumCounts Minimum number of hit counts for a sample to have to be included in the downstream statistical analyses and report generation + --nonparametric Set to use parametric bootstrapping in IQTREE analyses - --trymax Maximum number of iterations performed by metaMDS + --boots Number of bootstraps (recommended 1000 for parametric and 100 for non-parametric) + + + --Statistics options-- + + --stats Set "--stats run" to signal statstical tests to be performed and included in the final report + + --minimumCounts Minimum number of hit counts for a sample to have to be included in the downstream statistical analyses and report generation + + --trymax Maximum number of iterations performed by metaMDS ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||| @@ -414,7 +457,7 @@ def fullHelpMessage() { Example 4. Launching the vAMPirus Analyze pipeline with singularity with ASV and AminoType generation with all accesory analyses (taxonomy assignment, EMBOSS, IQTREE, statistics) - nextflow run vAMPirus.nf -c vampirus.config -profile singularity --Analyze --stats run + nextflow run vAMPirus.nf -c vampirus.config -profile singularity --Analyze --stats Example 5. Launching the vAMPirus Analyze pipeline with conda to perform multiple primer removal and protein-based clustering of ASVs, but skip most of the extra analyses @@ -422,7 +465,11 @@ def fullHelpMessage() { Example 6. Launching vAMPirus Analyze pipeline with conda to produce only ASV-related results - nextflow run vAMPirus.nf -c vampirus.config -profile conda --Analyze --skipAminoTyping --stats run + nextflow run vAMPirus.nf -c vampirus.config -profile conda --Analyze --skipAminoTyping --stats + + Example 7. Launching vAMPirus Analyze pipeline with conda to perform ASV analyses with Minimum Entropy Decomposition to form "Groups" + + nextflow run vAMPirus.nf -c vampirus.config -profile conda --Analyze --skipAminoTyping --stats --asvMED --asvC 24 Resuming analyses => @@ -445,7 +492,6 @@ if (params.help) { exit 0 } -// This will be printed to the user in each run. here thy can check if the values the selected are fine log.info """\ ================================================================================================================================================ vAMPirus v${workflow.manifest.version} - Virus Amplicon Sequencing Analysis Pipeline @@ -463,15 +509,35 @@ if (params.readsTest) { Channel .fromFilePairs(params.readsTest) .ifEmpty{ exit 1, "params.readTest was empty - no input files supplied" } - .into{ reads_ch; reads_qc_ch } + .into{ reads_ch; reads_qc_ch; reads_processing } } else { println("\n\tEverything ready for launch.\n") Channel .fromFilePairs("${params.reads}", checkIfExists: true) - .into{ reads_ch; reads_qc_ch } + .into{ reads_ch; reads_qc_ch; reads_processing } +} + +if (params.clusterNuclIDlist == "") { + a=params.clusterNuclID + slist=[a] + nnuc=slist.size() +} else { + msize=params.clusterNuclIDlist + slist=msize.split(',').collect{it as int} + nnuc=slist.size() +} + +if (params.clusterAAIDlist == "") { + b=params.clusterAAID + slist2=[b] + naa=slist2.size() +} else { + msize2=params.clusterAAIDlist + slist2=msize2.split(',').collect{it as int} + naa=slist2.size() } -if (params.Analyze) { +if (params.DataCheck || params.Analyze) { println("\n\tRunning vAMPirus Analyze pipeline - This might take a while, check out Nextflow tower (tower.nf) to remotely monitor the run.\n") @@ -530,7 +596,7 @@ if (params.Analyze) { } } - if (!params.skipReadProcessing) { + if (!params.skipReadProcessing || !params.skipMerging ) { if (!params.skipFastQC) { @@ -540,13 +606,13 @@ if (params.Analyze) { tag "${sample_id}" - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/FastQC/PreClean", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/FastQC/PreClean", mode: "copy", overwrite: true input: tuple sample_id, file(reads) from reads_qc_ch output: - tuple sample_id, file("*_fastqc.{zip,html}") into fastqc_results_OAS + tuple sample_id, file("*_fastqc.{zip,html}") into fastqc_results script: """ @@ -563,8 +629,8 @@ if (params.Analyze) { tag "${sample_id}" - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/AdapterRemoval", mode: "copy", overwrite: true, pattern: "*.filter.fq" - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/AdapterRemoval/fastpOut", mode: "copy", overwrite: true, pattern: "*.fastp.{json,html}" + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/AdapterRemoval", mode: "copy", overwrite: true, pattern: "*.filter.fq" + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/AdapterRemoval/fastpOut", mode: "copy", overwrite: true, pattern: "*.fastp.{json,html}" input: tuple sample_id, file(reads) from reads_ch @@ -572,7 +638,7 @@ if (params.Analyze) { output: tuple sample_id, file("*.fastp.{json,html}") into fastp_results tuple sample_id, file("*.filter.fq") into reads_fastp_ch - file("*.csv") into fastp_csv + file("*.csv") into ( fastp_csv_in1, fastp_csv_in2 ) script: """ @@ -589,6 +655,7 @@ if (params.Analyze) { reads_ch .set{ reads_fastp_ch } fastp_results = Channel.empty() + } if (!params.skipPrimerRemoval) { @@ -599,7 +666,7 @@ if (params.Analyze) { tag "${sample_id}" - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/PrimerRemoval", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/PrimerRemoval", mode: "copy", overwrite: true input: tuple sample_id, file(reads) from reads_fastp_ch @@ -636,9 +703,10 @@ if (params.Analyze) { } else { reads_fastp_ch .set{ reads_bbduk_ch } + } - if (!params.skipFastQC) { + if (!params.skipFastQC && !params.skipPrimerRemoval) { process QualityCheck_2 { @@ -646,13 +714,13 @@ if (params.Analyze) { tag "${sample_id}" - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/FastQC/PostClean", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/FastQC/PostClean", mode: "copy", overwrite: true input: tuple sample_id, file(reads) from readsforqc2 output: - tuple sample_id, file("*_fastqc.{zip,html}") into fastqc2_results_OAS + tuple sample_id, file("*_fastqc.{zip,html}") into fastqc2_results script: """ @@ -660,38 +728,50 @@ if (params.Analyze) { """ } } + } else { + reads_ch + .set{ reads_bbduk_ch } + } - process Read_Merging { + if (!params.skipMerging) { - label 'norm_cpus' + process Read_Merging { - tag "${sample_id}" + label 'norm_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging/Individual", mode: "copy", overwrite: true, pattern: "*mergedclean.fastq" - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging/Individual/notmerged", mode: "copy", overwrite: true, pattern: "*notmerged*.fastq" + tag "${sample_id}" - input: - tuple sample_id, file(reads) from reads_bbduk_ch + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/Individual", mode: "copy", overwrite: true, pattern: "*mergedclean.fastq" + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/Individual/notmerged", mode: "copy", overwrite: true, pattern: "*notmerged*.fastq" - output: - file("*_mergedclean.fastq") into reads_vsearch1_ch - file("*.name") into names - file("*notmerged*.fastq") into notmerged + input: + tuple sample_id, file(reads) from reads_bbduk_ch - script: - """ - vsearch --fastq_mergepairs ${reads[0]} --reverse ${reads[1]} --threads ${task.cpus} --fastqout ${sample_id}_mergedclean.fastq --fastqout_notmerged_fwd ${sample_id}_notmerged_fwd.fastq --fastqout_notmerged_rev ${sample_id}_notmerged_rev.fastq --fastq_maxee ${params.maxEE} --relabel ${sample_id}. - echo ${sample_id} > ${sample_id}.name - """ + output: + file("*_mergedclean.fastq") into reads_vsearch1_ch + file("*.name") into names + file("*notmerged*.fastq") into notmerged + + script: + """ + vsearch --fastq_mergepairs ${reads[0]} --reverse ${reads[1]} --threads ${task.cpus} --fastqout ${sample_id}_mergedclean.fastq --fastqout_notmerged_fwd ${sample_id}_notmerged_fwd.fastq --fastqout_notmerged_rev ${sample_id}_notmerged_rev.fastq --fastq_maxdiffs ${params.diffs} --fastq_maxns ${params.maxn} --fastq_allowmergestagger --fastq_maxee ${params.maxEE} --relabel ${sample_id}. + echo ${sample_id} > ${sample_id}.name + """ + + } + } else { + reads_bbduk_ch + .set{ reads_vsearch1_ch } } - process Compile_Reads { + + process Filtering_Prep1 { label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging/LengthFiltering", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/LengthFiltering", mode: "copy", overwrite: true input: file(reads) from reads_vsearch1_ch @@ -706,11 +786,11 @@ if (params.Analyze) { """ } - process Compile_Names { + process Filtering_Prep2 { label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging", mode: "copy", overwrite: true input: file(names) from names @@ -726,39 +806,78 @@ if (params.Analyze) { } - process Length_Filtering { //changed + process Length_Filtering { label 'norm_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging/LengthFiltering", mode: "copy", overwrite: true, pattern: "*_merged_preFilt*.fasta" - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging", mode: "copy", overwrite: true, pattern: "*Lengthfiltered.fastq" - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging/Histograms/pre_length_filtering", mode: "copy", overwrite: true, pattern: "*preFilt_*st.txt" - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging/Histograms/post_length_filtering", mode: "copy", overwrite: true, pattern: "*postFilt_*st.txt" + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/LengthFiltering", mode: "copy", overwrite: true, pattern: "*_merged_preFilt*.fasta" + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging", mode: "copy", overwrite: true, pattern: "*Lengthfiltered.fastq" + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/Histograms/pre_length_filtering", mode: "copy", overwrite: true, pattern: "*preFilt_*st.txt" + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/Histograms/post_length_filtering", mode: "copy", overwrite: true, pattern: "*postFilt_*st.txt" input: file(reads) from collect_samples_ch output: - file("*_merged_preFilt_clean.fasta") into ( nuclCounts_mergedreads_ch, pcASV_mergedreads_ch ) - file("*_merged_clean_Lengthfiltered.fastq") into reads_vsearch2_ch file("*_merged_preFilt_clean.fastq") into ( mergeforprotcounts, mergeforpcASVaacounts ) - file("**hist.txt") into histos + + file("*_merged_preFilt_clean.fasta") into ( nuclCounts_mergedreads_asv_ch, nuclCounts_mergedreads_ncasv_ch, pcASV_mergedreads_ch ) + file("*_merged_clean_Lengthfiltered.fastq") into reads_vsearch2_ch + + file("*preFilt_preClean_baseFrequency_hist.csv") into prefilt_basefreq + file("*preFilt_preClean_qualityScore_hist.csv") into prefilt_qualityscore + file("*preFilt_preClean_gcContent_hist.csv") into prefilt_gccontent + file("*preFilt_preClean_averageQuality_hist.csv") into prefilt_averagequality + file("*preFilt_preClean_length_hist.csv") into prefilt_length + + file("*postFilt_baseFrequency_hist.csv") into postFilt_basefreq + file("*postFilt_qualityScore_hist.csv") into postFilt_qualityscore + file("*postFilt_gcContent_hist.csv") into postFilt_gccontent + file("*postFilt_averageQuaulity_hist.csv") into postFilt_averagequality + file("*postFilt_length_hist.csv") into postFilt_length + + file("reads_per_sample_preFilt_preClean.csv") into reads_per_sample_preFilt + file("read_per_sample_postFilt_postClean.csv") into reads_per_sample_postFilt script: """ - bbduk.sh in=${reads} bhist=${params.projtag}_all_merged_preFilt_preClean_baseFrequency_hist.txt qhist=${params.projtag}_all_merged_preFilt_preClean_qualityScore_hist.txt gchist=${params.projtag}_all_merged_preFilt_preClean_gcContent_hist.txt aqhist=${params.projtag}_all_merged_preFilt_preClean_averageQuality_hist.txt lhist=${params.projtag}_all_merged_preFilt__preClean_length_hist.txt gcbins=auto + #bbduk.sh in=${reads} bhist=${params.projtag}_all_merged_preFilt_preClean_baseFrequency_hist.txt qhist=${params.projtag}_all_merged_preFilt_preClean_qualityScore_hist.txt gchist=${params.projtag}_all_merged_preFilt_preClean_gcContent_hist.txt aqhist=${params.projtag}_all_merged_preFilt_preClean_averageQuality_hist.txt lhist=${params.projtag}_all_merged_preFilt__preClean_length_hist.txt gcbins=auto + #fastp -i ${reads} -o ${params.projtag}_merged_preFilt_clean.fastq -b ${params.maxLen} -l ${params.minLen} --thread ${task.cpus} -n 1 + #reformat.sh in=${params.projtag}_merged_preFilt_clean.fastq out=${params.projtag}_merged_preFilt_clean.fasta t=${task.cpus} + #bbduk.sh in=${params.projtag}_merged_preFilt_clean.fastq out=${params.projtag}_merged_clean_Lengthfiltered.fastq minlength=${params.maxLen} maxlength=${params.maxLen} t=${task.cpus} + #bbduk.sh in=${params.projtag}_merged_clean_Lengthfiltered.fastq bhist=${params.projtag}_all_merged_postFilt_baseFrequency_hist.txt qhist=${params.projtag}_all_merged_postFilt_qualityScore_hist.txt gchist=${params.projtag}_all_merged_postFilt_gcContent_hist.txt aqhist=${params.projtag}_all_merged_postFilt_averageQuaulity_hist.txt lhist=${params.projtag}_all_merged_postFilt_length_hist.txt gcbins=auto + + # from DC + bbduk.sh in=${reads} bhist=${params.projtag}_all_merged_preFilt_preClean_baseFrequency_hist.txt qhist=${params.projtag}_all_merged_preFilt_preClean_qualityScore_hist.txt gchist=${params.projtag}_all_merged_preFilt_preClean_gcContent_hist.txt aqhist=${params.projtag}_all_merged_preFilt_preClean_averageQuality_hist.txt lhist=${params.projtag}_all_merged_preFilt_preClean_length_hist.txt gcbins=auto + for x in *preFilt*hist.txt;do + pre=\$(echo \$x | awk -F ".txt" '{print \$1}') + cat \$x | tr "\t" "," > \${pre}.csv + rm \$x + done + reformat.sh in=${reads} out=${params.projtag}_preFilt_preclean.fasta t=${task.cpus} + echo "sample,reads" >> reads_per_sample_preFilt_preClean.csv + grep ">" ${params.projtag}_preFilt_preclean.fasta | awk -F ">" '{print \$2}' | awk -F "." '{print \$1}' | sort --parallel=${task.cpus} | uniq -c | sort -brg --parallel=${task.cpus} | awk '{print \$2","\$1}' >> reads_per_sample_preFilt_preClean.csv + rm ${params.projtag}_preFilt_preclean.fasta fastp -i ${reads} -o ${params.projtag}_merged_preFilt_clean.fastq -b ${params.maxLen} -l ${params.minLen} --thread ${task.cpus} -n 1 reformat.sh in=${params.projtag}_merged_preFilt_clean.fastq out=${params.projtag}_merged_preFilt_clean.fasta t=${task.cpus} bbduk.sh in=${params.projtag}_merged_preFilt_clean.fastq out=${params.projtag}_merged_clean_Lengthfiltered.fastq minlength=${params.maxLen} maxlength=${params.maxLen} t=${task.cpus} bbduk.sh in=${params.projtag}_merged_clean_Lengthfiltered.fastq bhist=${params.projtag}_all_merged_postFilt_baseFrequency_hist.txt qhist=${params.projtag}_all_merged_postFilt_qualityScore_hist.txt gchist=${params.projtag}_all_merged_postFilt_gcContent_hist.txt aqhist=${params.projtag}_all_merged_postFilt_averageQuaulity_hist.txt lhist=${params.projtag}_all_merged_postFilt_length_hist.txt gcbins=auto - """ + for x in *postFilt*hist.txt;do + pre=\$(echo \$x | awk -F ".txt" '{print \$1}') + cat \$x | tr "\t" "," > \${pre}.csv + rm \$x + done + reformat.sh in=${params.projtag}_merged_clean_Lengthfiltered.fastq out=${params.projtag}_merged_clean_Lengthfiltered.fasta t=${task.cpus} + echo "sample,reads" >> read_per_sample_postFilt_postClean.csv + grep ">" ${params.projtag}_merged_clean_Lengthfiltered.fasta | awk -F ">" '{print \$2}' | awk -F "." '{print \$1}' | sort --parallel=${task.cpus} | uniq -c | sort -brg --parallel=${task.cpus} | awk '{print \$2","\$1}' >> read_per_sample_postFilt_postClean.csv + """ } - process Extract_Uniques { + process Extracting_Uniques { label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/ReadProcessing/ReadMerging/Uniques", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ReadMerging/Uniques", mode: "copy", overwrite: true input: file(reads) from reads_vsearch2_ch @@ -800,7 +919,7 @@ if (params.Analyze) { file(fasta) from reads_vsearch4_ch output: - file("*ASVs.fasta") into ( reads_vsearch5_ch, nucl2aa, asvsforAminotyping, asvfastaforcounts, asvaminocheck ) + file("*ASVs.fasta") into ( reads_vsearch5_ch, asv_med, nucl2aa, asvsforAminotyping, asvfastaforcounts, asvaminocheck ) script: """ @@ -810,73 +929,448 @@ if (params.Analyze) { // UNTIL HERE DEFAULT - if (params.ncASV) { + if (params.DataCheck) { + + process NucleotideBased_ASV_clustering_DC { + + label 'norm_cpus' + + publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Nucleotide", mode: "copy", overwrite: true, pattern: '*{.csv}' + + input: + file(fasta) from reads_vsearch5_ch + + output: + file("number_per_percentage_nucl.csv") into number_per_percent_nucl_plot + + script: + if (params.datacheckntIDlist) { + """ + for id in `echo ${params.datacheckntIDlist} | tr "," "\\n"`;do + vsearch --cluster_fast ${fasta} --centroids ${params.projtag}_ncASV\${id}.fasta --threads ${task.cpus} --relabel OTU --id \${id} + done + for x in *ncASV*.fasta;do + id=\$( echo \$x | awk -F "_ncASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') + numb=\$( grep -c ">" \$x ) + echo "\${id},\${numb}" >> number_per_percentage_nucl.csv + done + yo=\$(grep -c ">" ${fasta}) + echo "1.0,\${yo}" >> number_per_percentage_nucl.csv + """ + } + } + + process Translation_For_ProteinBased_Clustering_DC { + + label 'norm_cpus' + + publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Aminoacid/translation", mode: "copy", overwrite: true + + input: + file(fasta) from nucl2aa + + output: + file("*ASVprotforclust.fasta") into clustering_aa + file("*_translation_report") into reportaa_VR + file("*_ASV_all.fasta") into asvfastaforaaclust + + script: + """ + ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_ASVprotforclust.fasta --report ${params.projtag}_translation_report + cp ${fasta} ${params.projtag}_ASV_all.fasta + """ + } - process NucleotideBased_ASV_clustering { + process Protein_clustering_DC { label 'norm_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/ncASV", mode: "copy", overwrite: true, pattern: '*ncASV*.fasta' + publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Aminoacid", mode: "copy", overwrite: true, pattern: '*{.csv}' input: - file(fasta) from reads_vsearch5_ch + file(fasta) from clustering_aa + file(asvs) from asvfastaforaaclust output: - tuple file("*_ncASV*.fasta"), file("*ASV.fasta") into ( nuclFastas_forDiamond_ch, nuclFastas_forCounts_ch, nuclFastas_forMatrix_ch) - tuple file("*_ncASV*.fasta"), file("*ASV.fasta") into nuclFastas_forphylogeny - + file("number_per_percentage_prot.csv") into number_per_percent_prot_plot + file("*aminoacid_pcASV1.0_noTaxonomy.fasta") into amino_med script: - if (params.clusterNuclIDlist) { + // add awk script to count seqs """ - cp ${fasta} ./${params.projtag}_ASV.fasta - for id in `echo ${params.clusterNuclIDlist} | tr "," "\\n"`;do - vsearch --cluster_fast ${params.projtag}_ASV.fasta --centroids ${params.projtag}_ncASV\${id}.fasta --threads ${task.cpus} --relabel ncASV --id \${id} + set +e + cp ${params.vampdir}/bin/rename_seq.py . + for id in `echo ${params.datacheckaaIDlist} | tr "," "\\n"`;do + if [ \${id} == ".55" ];then + word=3 + elif [ \${id} == ".65" ];then + word=4 + else + word=5 + fi + awk 'BEGIN{RS=">";ORS=""}length(\$2)>="${params.minAA}"{print ">"\$0}' ${fasta} > ${params.projtag}_filtered_proteins.fasta + cd-hit -i ${params.projtag}_filtered_proteins.fasta -n \${word} -c \${id} -o ${params.projtag}_pcASV\${id}.fasta + sed 's/>Cluster />Cluster_/g' ${params.projtag}_pcASV\${id}.fasta.clstr >${params.projtag}_pcASV\${id}.clstr + grep ">Cluster_" ${params.projtag}_pcASV\${id}.clstr >temporaryclusters.list + y=\$(grep -c ">Cluster_" ${params.projtag}_pcASV\${id}.clstr) + echo ">Cluster_"\${y}"" >> ${params.projtag}_pcASV\${id}.clstr + t=1 + b=1 + for x in \$(cat temporaryclusters.list);do + echo "Extracting \$x" + name="\$( echo \$x | awk -F ">" '{print \$2}')" + clust="pcASV"\${t}"" + echo "\${name}" + awk '/^>'\${name}'\$/,/^>Cluster_'\${b}'\$/' ${params.projtag}_pcASV\${id}.clstr > "\${name}"_"\${clust}"_tmp.list + t=\$(( \${t}+1 )) + b=\$(( \${b}+1 )) + done + ls *_tmp.list + u=1 + for x in *_tmp.list;do + name="\$(echo \$x | awk -F "_p" '{print \$1}')" + echo "\${name}" + cluster="\$(echo \$x | awk -F "_" '{print \$3}')" + echo "\${cluster}" + grep "ASV" \$x | awk -F ", " '{print \$2}' | awk -F "_" '{print \$1}' | awk -F ">" '{print \$2}' > \${name}_\${cluster}_seqs_tmps.list + seqtk subseq ${asvs} \${name}_\${cluster}_seqs_tmps.list > \${name}_\${cluster}_nucleotide_sequences.fasta + vsearch --cluster_fast \${name}_\${cluster}_nucleotide_sequences.fasta --id 0.2 --centroids \${name}_\${cluster}_centroids.fasta + grep ">" \${name}_\${cluster}_centroids.fasta >> \${name}_\${cluster}_tmp_centroids.list + for y in \$( cat \${name}_\${cluster}_tmp_centroids.list );do + echo ">\${cluster}_type"\$u"" >> \${name}_\${cluster}_tmp_centroid.newheaders + u=\$(( \${u}+1 )) + done + u=1 + ./rename_seq.py \${name}_\${cluster}_centroids.fasta \${name}_\${cluster}_tmp_centroid.newheaders \${cluster}_types_labeled.fasta + done + cat *_types_labeled.fasta >> ${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta + grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$3}' | awk -F "." '{print \$1}' >tmphead.list + grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$2}' | awk -F "," '{print \$1}' >tmplen.list + paste -d"," temporaryclusters.list tmphead.list >tmp.info.csv + grep ">" ${params.projtag}_pcASV\${id}.fasta >lala.list + j=1 + for x in \$(cat lala.list);do + echo ">${params.projtag}_pcASV\${j}" >>${params.projtag}_aminoheaders.list + echo "\${x},>${params.projtag}_pcASV\${j}" >>tmpaminotype.info.csv + j=\$(( \${j}+1 )) + done + rm lala.list + awk -F "," '{print \$2}' tmp.info.csv >>tmporder.list + for x in \$(cat tmporder.list);do + grep -w "\$x" tmpaminotype.info.csv | awk -F "," '{print \$2}' >>tmpder.list + done + paste -d "," temporaryclusters.list tmplen.list tmphead.list tmpder.list >${params.projtag}_pcASVCluster\${id}_summary.csv + ./rename_seq.py ${params.projtag}_pcASV\${id}.fasta ${params.projtag}_aminoheaders.list ${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta + stats.sh in=${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_aminoacid_clustered.gc gcformat=4 overwrite=true + stats.sh in=${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_nucleotide_clustered.gc gcformat=4 overwrite=true + awk 'BEGIN{RS=">";ORS=""}length(\$2)<"${params.minAA}"{print ">"\$0}' ${fasta} >${params.projtag}_pcASV\${id}_problematic_translations.fasta + if [ `wc -l ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk '{print \$1}'` -gt 1 ];then + grep ">" ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk -F ">" '{print \$2}' > problem_tmp.list + seqtk subseq ${asvs} problem_tmp.list > ${params.projtag}_pcASV\${id}_problematic_nucleotides.fasta + else + rm ${params.projtag}_pcASV\${id}_problematic_translations.fasta + fi + rm *.list + rm Cluster* + rm *types* + rm *tmp* + rm ${params.projtag}_pcASV\${id}.fast* done + for x in *aminoacid*noTaxonomy.fasta;do + id=\$( echo \$x | awk -F "_noTax" '{print \$1}' | awk -F "pcASV" '{print \$2}') + numb=\$( grep -c ">" \$x) + echo "\${id},\${numb}" >> number_per_percentage_protz.csv + done + yesirr=\$( wc -l number_per_percentage_protz.csv | awk '{print \$1}') + tail -\$(( \${yesirr}-1 )) number_per_percentage_protz.csv > number_per_percentage_prot.csv + head -1 number_per_percentage_protz.csv >> number_per_percentage_prot.csv + rm number_per_percentage_protz.csv """ - } else if (params.clusterNuclID) { + } + + process ASV_Shannons_Entropy_Analysis { + + label 'norm_cpus' + + publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Nucleotide/ShannonEntropy", mode: "copy", overwrite: true + + input: + file(asvs) from asv_med + + output: + + file("*_ASV_entropy_breakdown.csv") into asv_entro_csv + file("*Aligned_informativeonly.fasta-ENTROPY") into asv_entropy + file("*ASV*") into entrop + + script: + """ + set +e + #alignment + mafft --thread ${task.cpus} --maxiterate 15000 --auto ${asvs} > ${params.projtag}_ASVs_mafftAlign.fasta + #trimming + trimal -in ${params.projtag}_ASVs_mafftAlign.fasta -out ${params.projtag}_ASVs_mafftAligned.fasta -keepheader -fasta -automated1 + rm ${params.projtag}_ASVs_mafftAlign.fasta + o-trim-uninformative-columns-from-alignment ${params.projtag}_ASVs_mafftAligned.fasta + mv ${params.projtag}_ASVs_mafftAligned.fasta-TRIMMED ./${params.projtag}_ASVs_Aligned_informativeonly.fasta + #entopy analysis + entropy-analysis ${params.projtag}_ASVs_Aligned_informativeonly.fasta + #summarize entropy peaks + awk '{print \$2}' ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY >> tmp_value.list + for x in \$(cat tmp_value.list) + do echo "\$x" + if [[ \$(echo ""\$x" > 0.0"|bc -l) -eq 1 ]]; + then echo dope >> above-0.0-.list + fi + if [[ \$(echo ""\$x" > 0.1"|bc -l) -eq 1 ]]; + then echo dope >> above-0.1-.list + fi + if [[ \$(echo ""\$x" > 0.2"|bc -l) -eq 1 ]]; + then echo dope >> above-0.2-.list + fi + if [[ \$(echo ""\$x" > 0.3"|bc -l) -eq 1 ]]; + then echo dope >> above-0.3-.list + fi + if [[ \$(echo ""\$x" > 0.4"|bc -l) -eq 1 ]]; + then echo dope >> above-0.4-.list + fi + if [[ \$(echo ""\$x" > 0.5"|bc -l) -eq 1 ]]; + then echo dope >> above-0.5-.list + fi + if [[ \$(echo ""\$x" > 0.6"|bc -l) -eq 1 ]]; + then echo dope >> above-0.6-.list + fi + if [[ \$(echo ""\$x" > 0.7"|bc -l) -eq 1 ]]; + then echo dope >> above-0.7-.list + fi + if [[ \$(echo ""\$x" > 0.8"|bc -l) -eq 1 ]]; + then echo dope >> above-0.8-.list + fi + if [[ \$(echo ""\$x" > 0.9"|bc -l) -eq 1 ]]; + then echo dope >> above-0.9-.list + fi + if [[ \$(echo ""\$x" > 1.0"|bc -l) -eq 1 ]]; + then echo dope >> above-1.0-.list + fi + if [[ \$(echo ""\$x" > 1.5"|bc -l) -eq 1 ]]; + then echo dope >> above-1.5-.list + fi + done + echo "Entropy,Peaks_above" >> ${params.projtag}_ASV_entropy_breakdown.csv + for z in above*.list; + do entrop=\$(echo \$z | awk -F "-" '{print \$2}') + echo ""\$entrop", "\$(wc -l \$z | awk '{print \$1}')"" >> ${params.projtag}_ASV_entropy_breakdown.csv + done + rm above* + mv ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY ./tmp.fasta + echo "Base_position Shannons_Entropy" >> ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY + cat tmp.fasta >> ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY + rm tmp.fasta + + """ + + } + + process AminoType_Shannons_Entropy_Analysis { + + label 'norm_cpus' + + publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Aminoacid/ShannonEntropy", mode: "copy", overwrite: true, pattern: '*{.csv}' + + input: + file(aminos) from amino_med + + output: + file("*AminoType_entropy_breakdown.csv") into amino_entro_csv + file ("*Aligned_informativeonly.fasta-ENTROPY") into amino_entropy + file("*AminoTypes*") into aminos + + script: + """ + #alignment + mafft --thread ${task.cpus} --maxiterate 15000 --auto ${aminos} > ${params.projtag}_AminoTypes_mafftAlign.fasta + #trimming + trimal -in ${params.projtag}_AminoTypes_mafftAlign.fasta -out ${params.projtag}_AminoTypes_mafftAligned.fasta -keepheader -fasta -automated1 + rm ${params.projtag}_AminoTypes_mafftAlign.fasta + o-trim-uninformative-columns-from-alignment ${params.projtag}_AminoTypes_mafftAligned.fasta + mv ${params.projtag}_AminoTypes_mafftAligned.fasta-TRIMMED ./${params.projtag}_AminoTypes_Aligned_informativeonly.fasta + #entropy analysis + entropy-analysis ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta --amino-acid-sequences + #summarize entropy peaks + awk '{print \$2}' ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta-ENTROPY >> tmp_value.list + for x in \$(cat tmp_value.list) + do echo "\$x" + if [[ \$(echo ""\$x" > 0.0"|bc -l) -eq 1 ]]; + then echo dope >> above-0.0-.list + fi + if [[ \$(echo ""\$x" > 0.1"|bc -l) -eq 1 ]]; + then echo dope >> above-0.1-.list + fi + if [[ \$(echo ""\$x" > 0.2"|bc -l) -eq 1 ]]; + then echo dope >> above-0.2-.list + fi + if [[ \$(echo ""\$x" > 0.3"|bc -l) -eq 1 ]]; + then echo dope >> above-0.3-.list + fi + if [[ \$(echo ""\$x" > 0.4"|bc -l) -eq 1 ]]; + then echo dope >> above-0.4-.list + fi + if [[ \$(echo ""\$x" > 0.5"|bc -l) -eq 1 ]]; + then echo dope >> above-0.5-.list + fi + if [[ \$(echo ""\$x" > 0.6"|bc -l) -eq 1 ]]; + then echo dope >> above-0.6-.list + fi + if [[ \$(echo ""\$x" > 0.7"|bc -l) -eq 1 ]]; + then echo dope >> above-0.7-.list + fi + if [[ \$(echo ""\$x" > 0.8"|bc -l) -eq 1 ]]; + then echo dope >> above-0.8-.list + fi + if [[ \$(echo ""\$x" > 0.9"|bc -l) -eq 1 ]]; + then echo dope >> above-0.9-.list + fi + if [[ \$(echo ""\$x" > 1.0"|bc -l) -eq 1 ]]; + then echo dope >> above-1.0-.list + fi + if [[ \$(echo ""\$x" > 1.5"|bc -l) -eq 1 ]]; + then echo dope >> above-1.5-.list + fi + done + echo "Entropy,Peaks_above" >> ${params.projtag}_AminoType_entropy_breakdown.csv + for z in above*.list; + do entrop=\$(echo \$z | awk -F "-" '{print \$2}') + echo ""\$entrop", "\$(wc -l \$z | awk '{print \$1}')"" >> ${params.projtag}_AminoType_entropy_breakdown.csv + done + rm above* + mv ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta-ENTROPY ./tmp.fasta + echo "Base_position Shannons_Entropy" >> ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta-ENTROPY + cat tmp.fasta >> ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta-ENTROPY + rm tmp.fasta + """ + } + + if (!params.skipReadProcessing || !params.skipMerging ) { + + process combine_csv_DC { + + input: + file(csv) from fastp_csv_in1 + .collect() + + output: + file("final_reads_stats.csv") into fastp_csv_dc + + script: + """ + cat ${csv} >all_reads_stats.csv + head -n1 all_reads_stats.csv >tmp.names.csv + cat all_reads_stats.csv | grep -v ""Sample,Total_"" >tmp.reads.stats.csv + cat tmp.names.csv tmp.reads.stats.csv >final_reads_stats.csv + rm tmp.names.csv tmp.reads.stats.csv + """ + + } + } else { + + process skip_combine_csv_DC { + output: + file("filter_reads.txt") into fastp_csv_dc + + script: + """ + echo "Read processing steps skipped." >filter_reads.txt + """ + } + } + + report_dc_in = Channel.create() + fastp_csv_dc.mix( reads_per_sample_preFilt, reads_per_sample_postFilt, prefilt_basefreq, postFilt_basefreq, prefilt_qualityscore, postFilt_qualityscore, prefilt_gccontent, postFilt_gccontent, prefilt_averagequality, postFilt_averagequality, prefilt_length, postFilt_length, number_per_percent_nucl_plot, number_per_percent_prot_plot, amino_entro_csv, amino_entropy, asv_entro_csv, asv_entropy).into(report_dc_in) + + process Report_DataCheck { + + label 'norm_cpus' + + publishDir "${params.workingdir}/${params.outdir}/DataCheck/Report", mode: "copy", overwrite: true, pattern: '*.{html}' + + input: + file(files) from report_dc_in + .collect() + + output: + file("*.html") into datacheckreport + + script: """ - cp ${fasta} ./${params.projtag}_ASV.fasta - id=${params.clusterNuclID} - vsearch --cluster_fast ${params.projtag}_ASV.fasta --centroids ${params.projtag}_ncASV\${id}.fasta --threads ${task.cpus} --relabel ncASV --id \${id} + cp ${params.vampdir}/bin/vAMPirus_DC_Report.Rmd . + cp ${params.vampdir}/example_data/conf/vamplogo.png . + Rscript -e "rmarkdown::render('vAMPirus_DC_Report.Rmd',output_file='${params.projtag}_DataCheck_Report.html')" ${params.projtag} \ + ${params.skipReadProcessing} \ + ${params.skipMerging} \ + ${params.skipAdapterRemoval} """ - } } - } else { - reads_vsearch5_ch - .into{ nuclFastas_forDiamond_ch; nuclFastas_forCounts_ch; nuclFastas_forphylogeny; nuclFastas_forMatrix_ch } - } - if (!params.skipTaxonomy) { + } else if (params.Analyze) { if (params.ncASV) { - process Nucleotide_Taxonomy_Inference { + reads_vsearch5_ch + .into{ asv_file_for_ncasvs; nuclFastas_forDiamond_asv_ch; nuclFastas_forCounts_asv_ch; nuclFastas_forphylogeny_asv; nuclFastas_forMatrix_asv_ch; asv_for_med } + + process NucleotideBased_ASV_clustering { + + label 'norm_cpus' - label 'high_cpus' + tag "${mtag}" - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy", mode: "copy", overwrite: true, pattern: '*_ASV*.{fasta,csv,tsv}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Taxonomy", mode: "copy", overwrite: true, pattern: '*ncASV*.{fasta,csv,tsv}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*_ASV*dmd.out' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*ncASV*dmd.out' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/ncASV", mode: "copy", overwrite: true, pattern: '*ncASV*.fasta' input: - tuple file(notus), file(asvs) from nuclFastas_forDiamond_ch + each x from 1..nnuc + file(fasta) from asv_file_for_ncasvs output: - file("*.fasta") into tax_labeled_fasta - tuple file("*_phyloseqObject.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond - file("*ncASV*summary_for_plot.csv") into taxplot1a - file("*_ASV*_summary_for_plot.csv") into taxplot1 + tuple nid, file("*_ncASV*.fasta") into ( nuclFastas_forphylogeny_ncasv, nuclFastas_forDiamond_ncasv_ch, nuclFastas_forCounts_ncasv_ch, nuclFastas_forMatrix_ncasv_ch ) script: + nid=slist.get(x-1) + mtag="ID=" + slist.get(x-1) + """ + vsearch --cluster_fast ${fasta} --centroids ${params.projtag}_ncASV${nid}.fasta --threads ${task.cpus} --relabel ncASV --id .${nid} """ - cp ${params.vampdir}/bin/rename_seq.py . - virdb=${params.dbdir}/${params.dbname} - grep ">" \${virdb} > headers.list - headers="headers.list" - for filename in ${notus};do - name=\$(ls \${filename} | awk -F ".fasta" '{print \$1}') - diamond blastx -q \${filename} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + } + + if (!params.skipTaxonomy) { + + if (params.headers == "RefSeq") { + + process ncASV_Taxonomy_Inference_RefSeq { /////// editttt + + label 'high_cpus' + + tag "${mtag}" + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Taxonomy", mode: "copy", overwrite: true, pattern: '*ncASV*.{fasta,csv,tsv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*ncASV*dmd.out' + + input: + tuple nid, file(asvs) from nuclFastas_forDiamond_ncasv_ch + + output: + file("*.fasta") into tax_labeled_fasta_ncasv + tuple file("*_phyloformat.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond_ncasv + tuple nid, file("*ncASV*summary_for_plot.csv") into taxplot_ncasv + tuple nid, file("*_quick_Taxbreakdown.csv") into tax_table_ncasv + + script: + mtag="ID=" + nid + """ + cp ${params.vampdir}/bin/rename_seq.py . + virdb=${params.dbdir}/${params.dbname} + grep ">" \${virdb} > headers.list + headers="headers.list" + name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') + diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" > access.list echo "[e-value]" > evalue.list @@ -884,18 +1378,14 @@ if (params.Analyze) { echo "[Percent ID (aa)]" > pid.list echo "[Organism ID]" > "\$name"_virus.list echo "[Gene]" > "\$name"_genes.list - grep ">" \${filename} | awk -F ">" '{print \$2}' > seqids.lst + echo "[ncASV#]" > otu.list + echo "[Sequence length]" > length.list + grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst echo "extracting genes and names" touch new_"\$name"_asvnames.txt - j=1 - if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then - echo "[ncASV#]" > otu.list - echo "[ncASV sequence length]" > length.list - for s in \$(cat seqids.lst);do - echo "Checking for \$s hit in diamond output" - if [[ ${params.refseq} == "T" ]];then - echo "RefSeq headers specified" - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + for s in \$(cat seqids.lst);do + echo "Checking for \$s hit in diamond output" + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then echo "Yep, there was a hit for \$s" echo "Extracting the information now:" acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') @@ -911,10 +1401,9 @@ if (params.Analyze) { echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">ncASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) + echo ">"\${s}"_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt echo "\$s done." - else + else echo "Ugh, there was no hit for \$s .." echo "We still love \$s though and we will add it to the final fasta file" echo "\$s" >> otu.list @@ -927,55 +1416,13 @@ if (params.Analyze) { echo "NO_HIT" >> length.list virus="NO" gene="HIT" - echo ">ncASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) + echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt echo "\$s done." - fi - else - echo "Using RVDB headers." - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">ncASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list - virus="NO" - gene="HIT" - echo ">ncASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - fi - fi - echo "Done with \$s" - done - fi + fi + done echo "Now editing "\$name" fasta headers" ###### rename_seq.py - ./rename_seq.py \${filename} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta + ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta echo "[Sequence header]" > newnames.list cat new_"\$name"_asvnames.txt >> newnames.list @@ -983,9 +1430,9 @@ if (params.Analyze) { echo " " > sequence.list grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - for x in *phyloseqObject.csv;do + for x in *phyloformat.csv;do echo "\$x" lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; @@ -993,17 +1440,270 @@ if (params.Analyze) { cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; rm tmpcol.list tmp2col.list done - rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; - rm *asvnames.txt - rm "\$name"_virus.list - rm "\$name"_genes.list - rm newnames.list - rm access.list - echo "Taxonomy inferred for: \${filename} " - done - for filename in ${asvs};do - name=\$(ls \${filename} | awk -F ".fasta" '{print \$1}') - diamond blastx -q \${filename} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list + """ + } + } else if (params.headers == "RVDB") { + + process ncASV_Taxonomy_Inference_RVDB { /////// editttt + + label 'high_cpus' + + tag "${mtag}" + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Taxonomy", mode: "copy", overwrite: true, pattern: '*ncASV*.{fasta,csv,tsv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*ncASV*dmd.out' + + input: + tuple nid, file(asvs) from nuclFastas_forDiamond_ncasv_ch + + output: + file("*.fasta") into tax_labeled_fasta_ncasv + tuple file("*_phyloformat.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond_ncasv + tuple nid, file("*ncASV*summary_for_plot.csv") into taxplot_ncasv + tuple nid, file("*_quick_Taxbreakdown.csv") into tax_table_ncasv + + script: + mtag="ID=" + nid + """ + cp ${params.vampdir}/bin/rename_seq.py . + virdb=${params.dbdir}/${params.dbname} + grep ">" \${virdb} > headers.list + headers="headers.list" + name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') + diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + echo "Preparing lists to generate summary .csv's" + echo "[Best hit accession number]" > access.list + echo "[e-value]" > evalue.list + echo "[Bitscore]" > bit.list + echo "[Percent ID (aa)]" > pid.list + echo "[Organism ID]" > "\$name"_virus.list + echo "[Gene]" > "\$name"_genes.list + echo "[ncASV#]" > otu.list + echo "[Sequence length]" > length.list + grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst + if [[ ${params.lca} == "T" ]] + then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list + echo "[Classification]" > classfication.list + else echo "skipped" >> \${name}_quick_Taxbreakdown.csv + echo "[Classification]" > classfication.list + fi + echo "extracting genes and names" + touch new_"\$name"_asvnames.txt + for s in \$(cat seqids.lst);do + echo "Using RVDB headers." + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + if [[ "${params.lca}" == "T" ]] + then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}') + lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') + echo "\$lcla" >> classfication.list + fi + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list + virus="NO" + gene="HIT" + echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + if [[ "${params.lca}" == "T" ]] + then echo "N/A" >> classfication.list + fi + echo "\$s done." + fi + echo "Done with \$s" + done + echo "Now editing "\$name" fasta headers" + ###### rename_seq.py + ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta + echo "[Sequence header]" > newnames.list + cat new_"\$name"_asvnames.txt >> newnames.list + touch sequence.list + echo " " > sequence.list + grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list + rm "\$name"_tmpssasv.fasta + if [[ "${params.lca}" == "T" ]] + then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list classfication.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv + else paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + fi + for x in *phyloformat.csv;do + echo "\$x" + lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) + tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; + sed 's/ /_/g' tmpcol.list > tmp2col.list; + cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; + rm tmpcol.list tmp2col.list + done + rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list + """ + } + } + } + + + process Generate_ncASV_Counts_Table { + + label 'norm_cpus' + + tag "${mtag}" + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Counts", mode: "copy", overwrite: true, pattern: '*_ASV*.{biome,csv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Counts", mode: "copy", overwrite: true, pattern: '*ncASV*.{biome,csv}' + + input: + tuple nid, file(notus) from nuclFastas_forCounts_ncasv_ch + file(merged) from nuclCounts_mergedreads_ncasv_ch + + output: + tuple file("*_counts.csv"), file("*_counts.biome") into counts_vsearch_ncasv + tuple nid, file("*ncASV*counts.csv") into notu_counts_plots + + script: + mtag="ID=" + nid + """ + name=\$( echo ${notus} | awk -F ".fasta" '{print \$1}') + vsearch --usearch_global ${merged} --db ${notus} --id .${nid} --threads ${task.cpus} --otutabout \${name}_counts.txt --biomout \${name}_counts.biome + cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv + sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv + rm \${name}_count.csv + """ + } + + process Generate_ncASV_Matrices { + label 'low_cpus' + + tag "${mtag}" + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Matrices", mode: "copy", overwrite: true, pattern: '*ncASV*PercentID.matrix' + + input: + tuple nid, file(asvs) from nuclFastas_forMatrix_ncasv_ch + + output: + file("*.matrix") into clustmatrices_ncasv + tuple nid, file("*ncASV*PercentID.matrix") into notu_heatmap + + script: + mtag="ID=" + nid + """ + name=\$( echo ${asvs}| awk -F ".fasta" '{print \$1}') + clustalo -i ${asvs} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} + clustalo -i ${asvs} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} + cat \${name}_PercentIDq.matrix | tr " " "," | grep "," >\${name}_PercentID.matrix + rm \${name}_PercentIDq.matrix + """ + } + + if (!params.skipPhylogeny) { + + process ncASV_Phylogeny { + + label 'norm_cpus' + + tag "${mtag}" + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*ncASV*aln.*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*ncASV*mt*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*ncASV*iq*' + + input: + tuple nid, file(asvs) from nuclFastas_forphylogeny_ncasv + + output: + tuple nid, file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into align_results_ncasv + tuple nid, file("*iq.treefile") into nucl_phyl_plot_ncasv + + script: + mtag="ID=" + nid + """ + pre=\$(echo ${asvs} | awk -F ".fasta" '{print \$1}' ) + mafft --thread ${task.cpus} --maxiterate 15000 --auto ${asvs} >\${pre}_ALN.fasta + trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html + o-trim-uninformative-columns-from-alignment \${pre}_aln.fasta + mv \${pre}_aln.fasta-TRIMMED ./\${pre}_Aligned_informativeonly.fasta + # Nucleotide_ModelTest + modeltest-ng -i \${pre}_Aligned_informativeonly.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint + # Nucleotide_Phylogeny + if [ "${params.iqCustomnt}" != "" ];then + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq --redo -t \${pre}_mt.tree -T auto ${params.iqCustomnt} + elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} + elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni + elif [ "${params.nonparametric}" != "false" ];then + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} + elif [ "${params.parametric}" != "false" ];then + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni + else + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni + fi + """ + } + + } + + } else { + reads_vsearch5_ch + .into{ nuclFastas_forDiamond_asv_ch; nuclFastas_forCounts_asv_ch; nuclFastas_forphylogeny_asv; nuclFastas_forMatrix_asv_ch; asv_for_med } + } + + if (!params.skipTaxonomy) { + + if (params.headers == "RefSeq") { + + process ASV_Taxonomy_Inference_RefSeq { /////// editttt + + label 'high_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy", mode: "copy", overwrite: true, pattern: '*_ASV*.{fasta,csv,tsv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*_ASV*dmd.out' + + input: + file(asvs) from nuclFastas_forDiamond_asv_ch + + output: + file("*.fasta") into tax_labeled_fasta_asv + tuple file("*_phyloformat.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond_asv + file("*_ASV*_summary_for_plot.csv") into taxplot_asv + file("*_quick_Taxbreakdown.csv") into tax_table_asv + + script: + """ + cp ${params.vampdir}/bin/rename_seq.py . + virdb=${params.dbdir}/${params.dbname} + grep ">" \${virdb} > headers.list + headers="headers.list" + name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') + diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" > access.list echo "[e-value]" > evalue.list @@ -1011,18 +1711,14 @@ if (params.Analyze) { echo "[Percent ID (aa)]" > pid.list echo "[Organism ID]" > "\$name"_virus.list echo "[Gene]" > "\$name"_genes.list - grep ">" \${filename} | awk -F ">" '{print \$2}' > seqids.lst + echo "[ASV#]" > otu.list + echo "[Sequence length]" > length.list + grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst echo "extracting genes and names" touch new_"\$name"_asvnames.txt - j=1 - if [ `echo \${filename} | grep -c "ASV"` -eq 1 ];then - for s in \$(cat seqids.lst);do - echo "[ASV#]" > otu.list - echo "[ASV sequence length]" > length.list - echo "Checking for \$s hit in diamond output" - if [[ ${params.refseq} == "T" ]];then - echo "RefSeq headers specified" - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + for s in \$(cat seqids.lst);do + echo "Checking for \$s hit in diamond output" + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then echo "Yep, there was a hit for \$s" echo "Extracting the information now:" acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') @@ -1038,48 +1734,9 @@ if (params.Analyze) { echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list - virus="NO" - gene="HIT" - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - fi - else - echo "Using RVDB headers." - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) + echo ">"\${s}"_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt echo "\$s done." - else + else echo "Ugh, there was no hit for \$s .." echo "We still love \$s though and we will add it to the final fasta file" echo "\$s" >> otu.list @@ -1092,17 +1749,13 @@ if (params.Analyze) { echo "NO_HIT" >> length.list virus="NO" gene="HIT" - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) + echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt echo "\$s done." - fi - fi - echo "Done with \$s" - done - fi + fi + done echo "Now editing "\$name" fasta headers" ###### rename_seq.py - ./rename_seq.py \${filename} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta + ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta echo "[Sequence header]" > newnames.list cat new_"\$name"_asvnames.txt >> newnames.list @@ -1110,9 +1763,9 @@ if (params.Analyze) { echo " " > sequence.list grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - for x in *phyloseqObject.csv;do + for x in *phyloformat.csv;do echo "\$x" lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; @@ -1120,146 +1773,572 @@ if (params.Analyze) { cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; rm tmpcol.list tmp2col.list done - rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; - rm *asvnames.txt - rm "\$name"_virus.list - rm "\$name"_genes.list - rm newnames.list - rm access.list - echo "Taxonomy inferred for: \${filename} " + rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list + """ + } + } else if (params.headers == "RVDB") { + + process ASV_Taxonomy_Inference_RVDB { /////// editttt + + label 'high_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy", mode: "copy", overwrite: true, pattern: '*_ASV*.{fasta,csv,tsv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*_ASV*dmd.out' + + input: + file(asvs) from nuclFastas_forDiamond_asv_ch + + output: + file("*.fasta") into tax_labeled_fasta_asv + tuple file("*_phyloformat.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond_asv + file("*_ASV*_summary_for_plot.csv") into taxplot_asv + file("*_quick_Taxbreakdown.csv") into tax_table_asv + + script: + """ + cp ${params.vampdir}/bin/rename_seq.py . + virdb=${params.dbdir}/${params.dbname} + grep ">" \${virdb} > headers.list + headers="headers.list" + name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') + diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + echo "Preparing lists to generate summary .csv's" + echo "[Best hit accession number]" > access.list + echo "[e-value]" > evalue.list + echo "[Bitscore]" > bit.list + echo "[Percent ID (aa)]" > pid.list + echo "[Organism ID]" > "\$name"_virus.list + echo "[Gene]" > "\$name"_genes.list + echo "[ASV#]" > otu.list + echo "[Sequence length]" > length.list + grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst + if [[ ${params.lca} == "T" ]] + then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list + echo "[Classification]" > classfication.list + else echo "skipped" >> \${name}_quick_Taxbreakdown.csv + echo "[Classification]" > classfication.list + fi + echo "extracting genes and names" + touch new_"\$name"_asvnames.txt + for s in \$(cat seqids.lst);do + echo "Using RVDB headers." + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + if [[ "${params.lca}" == "T" ]] + then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}') + lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') + echo "\$lcla" >> classfication.list + fi + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list + virus="NO" + gene="HIT" + echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + if [[ "${params.lca}" == "T" ]] + then echo "N/A" >> classfication.list + fi + echo "\$s done." + fi + echo "Done with \$s" + done + echo "Now editing "\$name" fasta headers" + ###### rename_seq.py + ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta + echo "[Sequence header]" > newnames.list + cat new_"\$name"_asvnames.txt >> newnames.list + touch sequence.list + echo " " > sequence.list + grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list + rm "\$name"_tmpssasv.fasta + if [[ "${params.lca}" == "T" ]] + then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list classfication.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv + else paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + fi + for x in *phyloformat.csv;do + echo "\$x" + lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) + tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; + sed 's/ /_/g' tmpcol.list > tmp2col.list; + cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; + rm tmpcol.list tmp2col.list + done + rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list + """ + } + } + } + + process Generate_ASV_Counts_Tables { + + label 'norm_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Counts", mode: "copy", overwrite: true, pattern: '*ASV*.{biome,csv}' + + input: + file(asvs) from nuclFastas_forCounts_asv_ch + file(merged) from nuclCounts_mergedreads_asv_ch + + output: + tuple file("*_counts.csv"), file("*_counts.biome") into counts_vsearch_asv + file("*_ASV*counts.csv") into (asv_counts_plots, asvcount_med) + + script: + """ + name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') + vsearch --usearch_global ${merged} --db ${asvs} --id .${params.asvcountID} --threads ${task.cpus} --otutabout "\$name"_counts.txt --biomout "\$name"_counts.biome + cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv + sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv + rm \${name}_count.csv + """ + } + + process Generate_ASV_Matrices { + + label 'low_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Matrices", mode: "copy", overwrite: true, pattern: '*ASV*PercentID.matrix' + + input: + file(reads) from nuclFastas_forMatrix_asv_ch + + output: + file("*.matrix") into clustmatrices_asv + file("*_ASV*PercentID.matrix") into asv_heatmap + + script: + // remove if statement later (no fin) + """ + for filename in ${reads};do + if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then + ident=\$( echo \${filename} | awk -F "ncASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') + name=\$( echo \${filename}| awk -F ".fasta" '{print \$1}') + clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} + clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} + for x in *q.matrix;do + pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') + ya=\$(wc -l \$x | awk '{print \$1}') + echo "\$((\$ya-1))" + tail -"\$((\$ya-1))" \$x > \${pre}z.matrix + rm \$x + cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix + rm \${pre}z.matrix + done + else + name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') + clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} + clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} + for x in *q.matrix;do + pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') + ya=\$(wc -l \$x | awk '{print \$1}') + echo "\$((\$ya-1))" + tail -"\$((\$ya-1))" \$x > \${pre}z.matrix + rm \$x + cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix + rm \${pre}z.matrix + done + fi + done + """ + } + + if (!params.skipPhylogeny) { // need to edit paths + + process ASV_Phylogeny { + + label 'norm_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*ASV*aln.*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*ASV*mt*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*ASV*iq*' + + input: + file(asvs) from nuclFastas_forphylogeny_asv + + output: + tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into align_results_asv + file("*iq.treefile") into (nucl_phyl_plot_asv, asvphy_med) + + script: + """ + pre=\$(echo ${asvs} | awk -F ".fasta" '{print \$1}' ) + mafft --thread ${task.cpus} --maxiterate 15000 --auto ${asvs} >\${pre}_ALN.fasta + trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html + o-trim-uninformative-columns-from-alignment \${pre}_aln.fasta + mv \${pre}_aln.fasta-TRIMMED ./\${pre}_Aligned_informativeonly.fasta + # Nucleotide_ModelTest + modeltest-ng -i \${pre}_Aligned_informativeonly.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint + # Nucleotide_Phylogeny + if [ "${params.iqCustomnt}" != "" ];then + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomnt} + elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} + elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni + elif [ "${params.nonparametric}" != "false" ];then + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} + elif [ "${params.parametric}" != "false" ];then + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + else + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + fi + """ + } + } + + if (params.asvMED) { + + process ASV_Minimum_Entropy_Decomposition { + + label 'low_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/ASVs/MED", mode: "copy", overwrite: true + + input: + file(asvs) from asv_for_med + + output: + file("*_ASV_Grouping.csv") into asvgroupscsv + file("${params.projtag}_ASV_group_reps_aligned.fasta") into groupreps + file("${params.projtag}_asvMED_${params.asvC}") + + script: + """ + #alignment + mafft --thread ${task.cpus} --maxiterate 15000 --auto ${asvs} > ${params.projtag}_ASVs_mafftAlign.fasta + #trimming + trimal -in ${params.projtag}_ASVs_mafftAlign.fasta -out ${params.projtag}_ASVs_mafftAligned.fasta -keepheader -fasta -automated1 + rm ${params.projtag}_ASVs_mafftAlign.fasta + o-trim-uninformative-columns-from-alignment ${params.projtag}_ASVs_mafftAligned.fasta + mv ${params.projtag}_ASVs_mafftAligned.fasta-TRIMMED ./${params.projtag}_ASVs_Aligned_informativeonly.fasta + #entopy analysis + entropy-analysis ${params.projtag}_ASVs_Aligned_informativeonly.fasta + #Decomposition + oligotype ${params.projtag}_ASVs_Aligned_informativeonly.fasta ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY -o ${params.projtag}_asvMED_${params.asvC} -M 1 -c ${params.asvC} -N ${task.cpus} --skip-check-input --no-figures --skip-gen-html + #generatemaps + cd ./${params.projtag}_asvMED_${params.asvC}/OLIGO-REPRESENTATIVES/ + echo "ASV,GroupID,IDPattern" + j=1 + for x in *_unique; + do gid=\$(echo \$x | awk -F "_" '{print \$1}') + uni=\$(echo \$x | awk -F ""\${gid}"_" '{print \$2}' | awk -F "_uni" '{print \$1}') + grep ">" "\$gid"_"\$uni" | awk -F ">" '{print \$2}' > asv.list + seqtk subseq ../../${asvs} asv.list > Group"\${j}"_sequences.fasta + for z in \$( cat asv.list) + do echo ""\$z",Group"\$j","\$uni"" >> ${params.projtag}_ASV_Grouping.csv + + done + rm asv.list + echo ">Group\${j}" >> ${params.projtag}_ASV_group_reps_aligned.fasta + echo "\$uni" > group.list + seqtk subseq ../OLIGO-REPRESENTATIVES.fasta group.list > group.fasta + tail -1 group.fasta >> ${params.projtag}_ASV_group_reps_aligned.fasta + mv "\$gid"_"\$uni" ./Group"\$j"_"\$uni"_aligned.fasta + mv "\$gid"_"\$uni"_unique ./Group"\$j"_"\$uni"_unqiues_aligned.fasta + rm "\$gid"*.cPickle + j=\$((\$j+1)) done - rm headers.list + mv ${params.projtag}_ASV_Grouping.csv ../../ + mv ${params.projtag}_ASV_group_reps_aligned.fasta ../../ + cd .. """ - } + } - } else { + process ASV_MED_Reps_phylogeny { - process ASV_Taxonomy_Inference { + label 'low_cpus' - label 'high_cpus' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/MED/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*ASV*mt*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/MED/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*ASV*iq*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy", mode: "copy", overwrite: true, pattern: '*_ASV*.{fasta,csv,tsv}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*_ASV*dmd.out' + input: + file(reps) from groupreps - input: - file(reads) from nuclFastas_forDiamond_ch + output: + file("*_ASV_Group_Reps*") into align_results_asvmed + file("*iq.treefile") into asv_group_rep_tree - output: - file("*.fasta") into tax_labeled_fasta - tuple file("*_phyloseqObject.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_diamond - file("*_ASV*_summary_for_plot.csv") into taxplot1 - script: - """ - cp ${params.vampdir}/bin/rename_seq.py . - virdb=${params.dbdir}/${params.dbname} - grep ">" \${virdb} > headers.list - headers="headers.list" - for filename in ${reads};do - name=\$(ls \${filename} | awk -F ".fasta" '{print \$1}') - diamond blastx -q \${filename} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 - echo "Preparing lists to generate summary .csv's" - echo "[Best hit accession number]" > access.list - echo "[e-value]" > evalue.list - echo "[Bitscore]" > bit.list - echo "[Percent ID (aa)]" > pid.list - echo "[Organism ID]" > "\$name"_virus.list + script: + """ + # Protein_ModelTest + modeltest-ng -i ${reps} -p ${task.cpus} -o ${params.projtag}_ASV_Group_Reps_mt -d aa -s 203 --disable-checkpoint + + # Protein_Phylogeny + if [ "${params.iqCustomaa}" != "" ];then + iqtree -s ${reps} --prefix ${params.projtag}_ASV_Group_Reps_iq --redo -T auto ${params.iqCustomaa} + + elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then + mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') + iqtree -s ${reps} --prefix ${params.projtag}_ASV_Group_Reps_iq -m \${mod} --redo -nt auto -b ${params.boots} + + elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then + mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') + iqtree -s ${reps} --prefix ${params.projtag}_ASV_Group_Reps_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni + + elif [ "${params.nonparametric}" != "false" ];then + iqtree -s ${reps} --prefix ${params.projtag}_ASV_Group_Reps_iq -m MFP --redo -nt auto -b ${params.boots} + + elif [ "${params.parametric}" != "false" ];then + iqtree -s ${reps} --prefix ${params.projtag}_ASV_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + + else + iqtree -s ${reps} --prefix ${params.projtag}_ASV_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + fi + """ + } + + process Adding_ASV_MED_Info { + + label 'low_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/MED/", mode: "copy", overwrite: true + + input: + file(counts) from asvcount_med + file(map) from asvgroupscsv + + output: + file("${params.projtag}_ASV_Groupingcounts.csv") into asvgroupcounts + + script: + """ + awk -F "," '{print \$1}' ${counts} | sed '1d' > asv.list + echo "GroupID" >> group.list + for x in \$(cat asv.list); + do group=\$(grep -w \$x ${map} | awk -F "," '{print \$2}') + echo "\$group" >> group.list + done + paste -d',' group.list ${counts} > ${params.projtag}_ASV_Groupingcounts.csv + """ + } + } + + if (!params.skipAminoTyping) { + + process Translate_For_AminoTyping { + + label 'low_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes/Translation", mode: "copy", overwrite: true + + input: + file(fasta) from asvsforAminotyping + + output: + file("${params.projtag}_all_translations.fasta") into amintypegen + file("${params.projtag}_translation_report") into proteinstage_vap_report + + script: + """ + ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_all_translations.fasta --report ${params.projtag}_translation_report + """ + } + + process Generate_AminoTypes { + + label 'norm_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{clstr,csv,gc}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes/Problematic", mode: "copy", overwrite: true, pattern: '*problematic*.{fasta}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes", mode: "copy", overwrite: true, pattern: '*AminoTypes_noTaxonomy.{fasta}' + + input: + file(prot) from amintypegen + file(asvs) from asvaminocheck + + output: + tuple file("*.fasta"), file("${params.projtag}_AminoTypes.clstr"), file("${params.projtag}_clustered.gc") into ( supplementalfiles ) + file("${params.projtag}_AminoTypes_noTaxonomy.fasta") into ( aminotypesCounts, aminotypesMafft, aminotypesClustal, aminotypesBlast, aminotypesEmboss, aminos_for_med ) + file("${params.projtag}_AminoType_summary_map.csv") into aminomapmed + + script: + """ + set +e + cp ${params.vampdir}/bin/rename_seq.py . + awk 'BEGIN{RS=">";ORS=""}length(\$2)>="${params.minAA}"{print ">"\$0}' ${prot} >${params.projtag}_filtered_translations.fasta + awk 'BEGIN{RS=">";ORS=""}length(\$2)<"${params.minAA}"{print ">"\$0}' ${prot} >${params.projtag}_problematic_translations.fasta + if [ `wc -l ${params.projtag}_problematic_translations.fasta | awk '{print \$1}'` -gt 1 ];then + grep ">" ${params.projtag}_problematic_translations.fasta | awk -F ">" '{print \$2}' > problem_tmp.list + seqtk subseq ${asvs} problem_tmp.list > ${params.projtag}_problematic_nucleotides.fasta + else + rm ${params.projtag}_problematic_translations.fasta + fi + cd-hit -i ${params.projtag}_filtered_translations.fasta -c 1.0 -o ${params.projtag}_unlabeled_types.fasta + sed 's/>Cluster />Cluster_/g' ${params.projtag}_unlabeled_types.fasta.clstr >${params.projtag}_AminoTypes.clstr + grep ">Cluster_" ${params.projtag}_AminoTypes.clstr >tmpclusters.list + grep -w "*" ${params.projtag}_AminoTypes.clstr | awk '{print \$3}' | awk -F "." '{print \$1}' >tmphead.list + grep -w "*" ${params.projtag}_AminoTypes.clstr | awk '{print \$2}' | awk -F "," '{print \$1}' >tmplen.list + paste -d"," tmpclusters.list tmphead.list >tmp.info.csv + grep ">" ${params.projtag}_unlabeled_types.fasta >lala.list + j=1 + for x in \$(cat lala.list);do + echo ">${params.projtag}_AminoType\${j}" >>${params.projtag}_aminoheaders.list + echo "\${x},>${params.projtag}_AminoType\${j}" >>tmpaminotype.info.csv + j=\$(( \${j}+1 )) + done + rm lala.list + awk -F "," '{print \$2}' tmp.info.csv >>tmporder.list + for x in \$(cat tmporder.list);do + grep -w "\$x" tmpaminotype.info.csv | awk -F "," '{print \$2}' >>tmpder.list + done + paste -d "," tmpclusters.list tmplen.list tmphead.list tmpder.list >${params.projtag}_AminoType_summary_map.csv + rm tmp* + ./rename_seq.py ${params.projtag}_unlabeled_types.fasta ${params.projtag}_aminoheaders.list ${params.projtag}_AminoTypes_noTaxonomy.fasta + stats.sh in=${params.projtag}_AminoTypes_noTaxonomy.fasta gc=${params.projtag}_clustered.gc gcformat=4 + """ + } + + process Generate_AminoType_Matrices { + + label 'low_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Matrices", mode: "copy", overwrite: true + + input: + file(prot) from aminotypesClustal + + output: + file("*.matrix") into proclustmatrices + file("*PercentID.matrix") into aminotype_heatmap + + script: + """ + name=\$( echo ${prot} | awk -F "_noTax" '{print \$1}') + clustalo -i ${prot} --distmat-out=\${name}_PairwiseDistanceq.matrix --full --force --threads=${task.cpus} + clustalo -i ${prot} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} + for x in *q.matrix;do + pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') + ya=\$(wc -l \$x | awk '{print \$1}') + echo "\$((\$ya-1))" + tail -"\$(( \$ya-1))" \$x > \${pre}z.matrix + rm \$x + cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix + rm \${pre}z.matrix + done + """ + } + + if (!params.skipEMBOSS) { + + process AminoType_EMBOSS_Analyses { + + label 'low_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/2dStructure", mode: "copy", overwrite: true, pattern: '*.{garnier}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/HydrophobicMoment", mode: "copy", overwrite: true, pattern: '*HydrophobicMoments.{svg}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/IsoelectricPoint", mode: "copy", overwrite: true, pattern: '*IsoelectricPoint.{iep,svg}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/ProteinProperties", mode: "copy", overwrite: true, pattern: '*.{pepstats,pepinfo}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/ProteinProperties/Plots", mode: "copy", overwrite: true, pattern: '*PropertiesPlot.{svg}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/2dStructure/Plots", mode: "copy", overwrite: true, pattern: '*Helical*.{svg}' + + input: + file(prot) from aminotypesEmboss + + output: + tuple file("*.garnier"), file("*HydrophobicMoments.svg"), file("*IsoelectricPoint*"), file("*.pepstats"), file("*PropertiesPlot*"), file("*Helical*") into amino_emboss + + script: + """ + name=\$( echo ${prot} | awk -F ".fasta" '{print \$1}') + garnier -sequence ${prot} -outfile \${name}_2dStructures.garnier + hmoment -seqall ${prot} -graph svg -plot + mv hmoment.svg ./"\${name}"_HydrophobicMoments.svg + iep -sequence ${prot} -graph svg -plot -outfile "\${name}"_IsoelectricPoint.iep + mv iep.svg ./"\${name}"_IsoelectricPoint.svg + pepstats -sequence ${prot} -outfile \${name}_ProteinProperties.pepstats + grep ">" ${prot} | awk -F ">" '{print \$2}' > tmpsequence.list + for x in \$(cat tmpsequence.list);do + echo \$x > tmp1.list + seqtk subseq ${prot} tmp1.list > tmp2.fasta + len=\$(tail -1 tmp2.fasta | awk '{print length}') + pepinfo -sequence tmp2.fasta -graph svg -outfile "\$x"_PropertiesPlot.pepinfo + mv pepinfo.svg ./"\$x"_PropertiesPlot.svg + cat "\$x"_PropertiesPlot.pepinfo >> "\${name}"_PropertiesPlot.pepinfo + rm "\$x"_PropertiesPlot.pepinfo + pepnet -sask -sequence tmp2.fasta -graph svg -sbegin1 1 -send1 \$len + mv pepnet.svg ./"\$x"_HelicalNet.svg + pepwheel -sequence tmp2.fasta -graph svg -sbegin1 1 -send1 \$len + mv pepwheel.svg ./"\$x"_HelicalWheel.svg + rm tmp1.list tmp2.fasta + done + rm tmpsequence.list + """ + } + } + + if (!params.skipTaxonomy) { + + if (params.headers == "RefSeq") { + + process AminoType_Taxonomy_Inference_RefSeq { + + label 'high_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{csv,tsv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Taxonomy", mode: "copy", overwrite: true, pattern: '*TaxonomyLabels.fasta' + + input: + file(asvs) from aminotypesBlast + + output: + tuple file("*_phyloformat.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AA_diamond + file("*_summary_for_plot.csv") into taxplot2 + file("*TaxonomyLabels.fasta") into tax_labeled_fasta2 + file("*_quick_Taxbreakdown.csv") into tax_table_amino + + script: + """ + cp ${params.vampdir}/bin/rename_seq.py . + virdb=${params.dbdir}/${params.dbname} + grep ">" \${virdb} > headers.list + headers="headers.list" + name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') + diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + echo "Preparing lists to generate summary .csv's" + echo "[Best hit accession number]" > access.list + echo "[e-value]" > evalue.list + echo "[Bitscore]" > bit.list + echo "[Percent ID (aa)]" > pid.list + echo "[Organism ID]" > "\$name"_virus.list echo "[Gene]" > "\$name"_genes.list - grep ">" \${filename} | awk -F ">" '{print \$2}' > seqids.lst + echo "[AminoType#]" > otu.list + echo "[Sequence length]" > length.list + grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst echo "extracting genes and names" touch new_"\$name"_asvnames.txt - j=1 - if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then - echo "[ASV#]" > otu.list - echo "[ASV sequence length]" > length.list - for s in \$(cat seqids.lst);do - echo "Checking for \$s hit in diamond output" - if [[ ${params.refseq} == "T" ]];then - echo "RefSeq headers specified" - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list - virus="NO" - gene="HIT" - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - fi - else - echo "Using RVDB headers." - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list - virus="NO" - gene="HIT" - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - fi - fi - echo "Done with \$s" - done - else - for s in \$(cat seqids.lst);do - echo "[ASV#]" > otu.list - echo "[ASV sequence length]" > length.list - echo "Checking for \$s hit in diamond output" - if [[ ${params.refseq} == "T" ]];then - echo "RefSeq headers specified" - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + for s in \$(cat seqids.lst);do + echo "Checking for \$s hit in diamond output" + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then echo "Yep, there was a hit for \$s" echo "Extracting the information now:" acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') @@ -1275,48 +2354,9 @@ if (params.Analyze) { echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >> access.list - echo "NO_HIT" >> "\$name"_genes.list - echo "NO_HIT" >> "\$name"_virus.list - echo "NO_HIT" >> evalue.list - echo "NO_HIT" >> bit.list - echo "NO_HIT" >> pid.list - echo "NO_HIT" >> length.list - virus="NO" - gene="HIT" - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - fi - else - echo "Using RVDB headers." - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >> evalue.list - echo "\$line" | awk '{print \$11}' >> bit.list - echo "\$line" | awk '{print \$12}' >> pid.list - echo "\$line" | awk '{print \$2}' >> length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) + echo ">"\${s}"_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt echo "\$s done." - else + else echo "Ugh, there was no hit for \$s .." echo "We still love \$s though and we will add it to the final fasta file" echo "\$s" >> otu.list @@ -1329,17 +2369,13 @@ if (params.Analyze) { echo "NO_HIT" >> length.list virus="NO" gene="HIT" - echo ">ASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) + echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt echo "\$s done." - fi - fi - echo "Done with \$s" - done - fi + fi + done echo "Now editing "\$name" fasta headers" ###### rename_seq.py - ./rename_seq.py \${filename} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta + ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta echo "[Sequence header]" > newnames.list cat new_"\$name"_asvnames.txt >> newnames.list @@ -1347,9 +2383,9 @@ if (params.Analyze) { echo " " > sequence.list grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - for x in *phyloseqObject.csv;do + for x in *phyloformat.csv;do echo "\$x" lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; @@ -1357,2408 +2393,1400 @@ if (params.Analyze) { cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; rm tmpcol.list tmp2col.list done - rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; - rm *asvnames.txt - rm "\$name"_virus.list - rm "\$name"_genes.list - rm newnames.list - rm access.list - echo "Taxonomy inferred for: \${filename} " - done - rm headers.list - """ + rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list + """ + } + } else if (params.headers == "RVDB") { + + process AminoType_Taxonomy_Inference_RVDB { + + label 'high_cpus' + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{csv,tsv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Taxonomy", mode: "copy", overwrite: true, pattern: '*TaxonomyLabels.fasta' + + input: + file(asvs) from aminotypesBlast + + output: + tuple file("*_phyloformat.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AA_diamond + file("*_summary_for_plot.csv") into taxplot2 + file("*TaxonomyLabels.fasta") into tax_labeled_fasta2 + file("*_quick_Taxbreakdown.csv") into tax_table_amino + + script: + """ + cp ${params.vampdir}/bin/rename_seq.py . + virdb=${params.dbdir}/${params.dbname} + grep ">" \${virdb} > headers.list + headers="headers.list" + name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') + diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + echo "Preparing lists to generate summary .csv's" + echo "[Best hit accession number]" > access.list + echo "[e-value]" > evalue.list + echo "[Bitscore]" > bit.list + echo "[Percent ID (aa)]" > pid.list + echo "[Organism ID]" > "\$name"_virus.list + echo "[Gene]" > "\$name"_genes.list + echo "[AminoType#]" > otu.list + echo "[Sequence length]" > length.list + grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst + if [[ ${params.lca} == "T" ]] + then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list + echo "[Classification]" > classfication.list + else echo "skipped" >> \${name}_quick_Taxbreakdown.csv + echo "[Classification]" > classfication.list + fi + echo "extracting genes and names" + touch new_"\$name"_asvnames.txt + for s in \$(cat seqids.lst);do + echo "Using RVDB headers." + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + if [[ "${params.lca}" == "T" ]] + then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}') + lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') + echo "\$lcla" >> classfication.list + fi + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list + virus="NO" + gene="HIT" + echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + if [[ "${params.lca}" == "T" ]] + then echo "N/A" >> classfication.list + fi + echo "\$s done." + fi + echo "Done with \$s" + done + echo "Now editing "\$name" fasta headers" + ###### rename_seq.py + ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta + echo "[Sequence header]" > newnames.list + cat new_"\$name"_asvnames.txt >> newnames.list + touch sequence.list + echo " " > sequence.list + grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list + rm "\$name"_tmpssasv.fasta + if [[ "${params.lca}" == "T" ]] + then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list classfication.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv + else paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + fi + for x in *phyloformat.csv;do + echo "\$x" + lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) + tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; + sed 's/ /_/g' tmpcol.list > tmp2col.list; + cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; + rm tmpcol.list tmp2col.list + done + rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list + """ + } } - } - } + } - if (params.ncASV) { + if (!params.skipPhylogeny) { - process Generate_Counts_Tables_Nucleotide { + process AminoType_Phylogeny { - label 'norm_cpus' + label 'norm_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Counts", mode: "copy", overwrite: true, pattern: '*_ASV*.{biome,csv}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Counts", mode: "copy", overwrite: true, pattern: '*ncASV*.{biome,csv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*aln.*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Phylogeny/Modeltest", mode: "copy", overwrite: true, pattern: '*mt*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*iq*' - input: - tuple file(notus), file(asvs) from nuclFastas_forCounts_ch - file(merged) from nuclCounts_mergedreads_ch + input: + file(prot) from aminotypesMafft - output: - tuple file("*_counts.csv"), file("*_counts.biome") into counts_vsearch - file("*ncASV*counts.csv") into notu_counts_plots - file("*_ASV*counts.csv") into asv_counts_plots + output: + tuple file("*_aln.fasta"), file("*_aln.html"), file("*.log"), file("*iq*"), file("*mt*") into alignprot_results + file("*iq.treefile") into (amino_rax_plot, amino_repphy) - script: - """ - for filename in ${notus};do - if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then - ident=\$( echo \${filename} | awk -F "ncASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') - name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') - vsearch --usearch_global ${merged} --db \${filename} --id \${ident} --threads ${task.cpus} --otutabout \${name}_counts.txt --biomout \${name}_counts.biome - cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv - sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv - rm \${name}_count.csv - fi - done - if [ `echo ${asvs} | grep -c "ASV"` -eq 1 ];then - name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - vsearch --usearch_global ${merged} --db ${asvs} --id ${params.asvcountID} --threads ${task.cpus} --otutabout "\$name"_counts.txt --biomout "\$name"_counts.biome - cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv - sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv - rm \${name}_count.csv - fi - """ - } - } else { - process Generate_ASV_Counts_Tables { + script: + """ + # Protein_Alignment + pre=\$(echo ${prot} | awk -F "_noTax" '{print \$1}' ) + mafft --thread ${task.cpus} --maxiterate 15000 --auto ${prot} >\${pre}_ALN.fasta + trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html + o-trim-uninformative-columns-from-alignment \${pre}_aln.fasta + mv \${pre}_aln.fasta-TRIMMED ./\${pre}_Aligned_informativeonly.fasta + # Protein_ModelTest + modeltest-ng -i \${pre}_Aligned_informativeonly.fasta -p ${task.cpus} -o \${pre}_mt -d aa -s 203 --disable-checkpoint - label 'norm_cpus' + # Protein_Phylogeny + if [ "${params.iqCustomaa}" != "" ];then + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomaa} - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Counts", mode: "copy", overwrite: true, pattern: '*ASV*.{biome,csv}' + elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} - input: - file(asvs) from nuclFastas_forCounts_ch - file(merged) from nuclCounts_mergedreads_ch + elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni - output: - tuple file("*_counts.csv"), file("*_counts.biome") into counts_vsearch - file("*_ASV*counts.csv") into asv_counts_plots + elif [ "${params.nonparametric}" != "false" ];then + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} - script: - """ - if [ `echo ${asvs} | grep -c "ASV"` -eq 1 ];then - name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - vsearch --usearch_global ${merged} --db ${asvs} --id ${params.asvcountID} --threads ${task.cpus} --otutabout "\$name"_counts.txt --biomout "\$name"_counts.biome - cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv - sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv - rm \${name}_count.csv - fi - """ + elif [ "${params.parametric}" != "false" ];then + iqtree -s\${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + + else + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + fi + """ + } } - } - if (params.ncASV) { + process Generate_AminoTypes_Counts_Table { - process Generate_Nucleotide_Matrix { + label 'high_cpus' - label 'low_cpus' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Counts", mode: "copy", overwrite: true - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Matrix", mode: "copy", overwrite: true, pattern: '*_ASV*PercentID.matrix' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Matrix", mode: "copy", overwrite: true, pattern: '*ncASV*PercentID.matrix' - - input: - tuple file(notus), file(asvs) from nuclFastas_forMatrix_ch - - output: - file("*.matrix") into clustmatrices - file("*ncASV*PercentID.matrix") into notu_heatmap - file("*_ASV*PercentID.matrix") into asv_heatmap - - script: - // remove if statement later (no fin) - """ - for filename in ${notus};do - if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then - ident=\$( echo \${filename} | awk -F "ncASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') - name=\$( echo \${filename}| awk -F ".fasta" '{print \$1}') - clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} - clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} - for x in *q.matrix;do - pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') - ya=\$(wc -l \$x | awk '{print \$1}') - echo "\$((\$ya-1))" - tail -"\$((\$ya-1))" \$x > \${pre}z.matrix - rm \$x - cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix - rm \${pre}z.matrix - done - fi - done - if [ `echo ${asvs} | grep -c "_ASV"` -eq 1 ];then - name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - clustalo -i ${asvs} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} - clustalo -i ${asvs} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} - for x in *q.matrix;do - pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') - ya=\$(wc -l \$x | awk '{print \$1}') - echo "\$((\$ya-1))" - tail -"\$((\$ya-1))" \$x > \${pre}z.matrix - rm \$x - cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix - rm \${pre}z.matrix - done - fi - """ - } - } else { - - process Generate_ASV_Matrix { - - label 'low_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Matrix", mode: "copy", overwrite: true, pattern: '*ASV*PercentID.matrix' - - input: - file(reads) from nuclFastas_forMatrix_ch - - output: - file("*.matrix") into clustmatrices - file("*_ASV*PercentID.matrix") into asv_heatmap - - script: - // remove if statement later (no fin) - """ - for filename in ${reads};do - if [ `echo \${filename} | grep -c "ncASV"` -eq 1 ];then - ident=\$( echo \${filename} | awk -F "ncASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') - name=\$( echo \${filename}| awk -F ".fasta" '{print \$1}') - clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} - clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} - for x in *q.matrix;do - pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') - ya=\$(wc -l \$x | awk '{print \$1}') - echo "\$((\$ya-1))" - tail -"\$((\$ya-1))" \$x > \${pre}z.matrix - rm \$x - cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix - rm \${pre}z.matrix - done - else - name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') - clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistance.matrix --full --force --threads=${task.cpus} - clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} - for x in *q.matrix;do - pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') - ya=\$(wc -l \$x | awk '{print \$1}') - echo "\$((\$ya-1))" - tail -"\$((\$ya-1))" \$x > \${pre}z.matrix - rm \$x - cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix - rm \${pre}z.matrix - done - fi - done - """ - } - } - - if (!params.skipPhylogeny) { // need to edit paths - - if (params.ncASV) { - - process Nucleotide_Phylogeny { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*ncASV*aln.*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*ncASV*mt*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ncASV/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*ncASV*iq*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*_ASV*aln.*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*_ASV*mt*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*_ASV*iq*' - - input: - tuple file(notus), file(asvs) from nuclFastas_forphylogeny + input: + file(fasta) from aminotypesCounts + file(merged) from mergeforprotcounts + file(samplist) from samplelist output: - tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into align_results - file("*iq.treefile") into nucl_phyl_plot + tuple file("*_AminoType_counts.csv"), file("*dmd.out") into counts_summary + file("*_AminoType_counts.csv") into (aminocounts_plot, aminocountmed) script: """ - for filename in ${notus};do - pre=\$(echo \${filename} | awk -F ".fasta" '{print \$1}' ) - mafft --thread ${task.cpus} --maxiterate 15000 --auto \${filename} >\${pre}_ALN.fasta - trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html - # Nucleotide_ModelTest - modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint - # Nucleotide_Phylogeny - if [ "${params.iqCustomnt}" != "" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -t \${pre}_mt.tree -T auto ${params.iqCustomnt} - elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} - elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni - elif [ "${params.nonparametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} - elif [ "${params.parametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni - else - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni - fi - done - for filename in ${asvs};do - pre=\$(echo \${filename} | awk -F ".fasta" '{print \$1}' ) - mafft --thread ${task.cpus} --maxiterate 15000 --auto \${filename} >\${pre}_ALN.fasta - trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html - # Nucleotide_ModelTest - modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint - # Nucleotide_Phylogeny - if [ "${params.iqCustomnt}" != "" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomnt} - elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} - elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni - elif [ "${params.nonparametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} - elif [ "${params.parametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - else - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - fi - done - """ + set +e + diamond makedb --in ${fasta} --db ${fasta} + diamond blastx -q ${merged} -d ${fasta} -p ${task.cpus} --min-score ${params.ProtCountsBit} --id ${params.ProtCountID} -l ${params.ProtCountsLength} --more-sensitive -o ${params.projtag}_protCounts_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 --max-hsps 1 + echo "OTU_ID" >tmp.col1.txt + echo "Generating sample id list" + grep ">" ${fasta} | awk -F ">" '{print \$2}' | sort | uniq > otuid.list + cat otuid.list >> tmp.col1.txt + echo "Beginning them counts tho my g" + for y in \$( cat ${samplist} );do + echo "Starting with \$y now ..." + grep "\$y" ${params.projtag}_protCounts_dmd.out > tmp."\$y".out + echo "Isolated hits" + echo "Created uniq subject id list" + echo "\$y" > "\$y"_col.txt + echo "Starting my counts" + for z in \$(cat otuid.list);do + echo "Counting \$z hits" + echo "grep -wc "\$z" >> "\$y"_col.txt" + grep -wc "\$z" tmp."\$y".out >> "\$y"_col.txt + echo "\$z counted" + done + done + paste -d "," tmp.col1.txt *col.txt > ${params.projtag}_AminoType_counts.csv + rm tmp* + rm *col.txt + """ } - } else { - - process ASV_Phylogeny { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*ASV*aln.*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*ASV*mt*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*ASV*iq*' - - input: - file(asvs) from nuclFastas_forphylogeny - - output: - tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into align_results - file("*iq.treefile") into nucl_phyl_plot - - script: - """ - for filename in ${asvs};do - pre=\$(echo \${filename} | awk -F ".fasta" '{print \$1}' ) - mafft --thread ${task.cpus} --maxiterate 15000 --auto \${filename} >\${pre}_ALN.fasta - trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html - # Nucleotide_ModelTest - modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint - # Nucleotide_Phylogeny - if [ "${params.iqCustomnt}" != "" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomnt} - elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} - elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni - elif [ "${params.nonparametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} - elif [ "${params.parametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - else - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - fi - done - """ - } } - } - - if (!params.skipAminoTyping) { - if (params.sing) { + if (params.aminoMED) { - process Translating_For_Aminotypes { + process AminoType_Minimum_Entropy_Decomposition { label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes/Translation", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes/MED", mode: "copy", overwrite: true input: - file(fasta) from asvsforAminotyping + file(aminos) from aminos_for_med output: - file("${params.projtag}_all_translations.fasta") into amintypegen - file("${params.projtag}_translation_report") into proteinstage_vap_report + file("*_AminoType_Grouping.csv") into atygroupscsv + file("${params.projtag}_AminoType_group_reps_aligned.fasta") into atygroupreps script: - """ - conda init && source activate virtualribosome - - ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_all_translations.fasta --report ${params.projtag}_translation_report - """ - - } - - } else { - - process Translate_For_AminoTyping { - - label 'low_cpus' - - conda 'python=2.7' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes/Translation", mode: "copy", overwrite: true - - input: - file(fasta) from asvsforAminotyping - - output: - file("${params.projtag}_all_translations.fasta") into amintypegen - file("${params.projtag}_translation_report") into proteinstage_vap_report - - script: - """ - ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_all_translations.fasta --report ${params.projtag}_translation_report """ - } - - } - - process Generate_AminoTypes { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{clstr,csv,gc}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes/Problematic", mode: "copy", overwrite: true, pattern: '*problematic*.{fasta}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes", mode: "copy", overwrite: true, pattern: '*AminoTypes_noTaxonomy.{fasta}' - - input: - file(prot) from amintypegen - file(asvs) from asvaminocheck - - output: - tuple file("*.fasta"), file("${params.projtag}_AminoTypes.clstr"), file("${params.projtag}_AminoType_summary_map.csv"), file("${params.projtag}_clustered.gc") into ( supplementalfiles ) - file("${params.projtag}_AminoTypes_noTaxonomy.fasta") into ( aminotypesCounts, aminotypesMafft, aminotypesClustal, aminotypesBlast, aminotypesEmboss ) - - script: - """ - set +e - cp ${params.vampdir}/bin/rename_seq.py . - awk 'BEGIN{RS=">";ORS=""}length(\$2)>="${params.minAA}"{print ">"\$0}' ${prot} >${params.projtag}_filtered_translations.fasta - awk 'BEGIN{RS=">";ORS=""}length(\$2)<"${params.minAA}"{print ">"\$0}' ${prot} >${params.projtag}_problematic_translations.fasta - if [ `wc -l ${params.projtag}_problematic_translations.fasta | awk '{print \$1}'` -gt 1 ];then - grep ">" ${params.projtag}_problematic_translations.fasta | awk -F ">" '{print \$2}' > problem_tmp.list - seqtk subseq ${asvs} problem_tmp.list > ${params.projtag}_problematic_nucleotides.fasta - else - rm ${params.projtag}_problematic_translations.fasta - fi - cd-hit -i ${params.projtag}_filtered_translations.fasta -c 1.0 -o ${params.projtag}_unlabeled_types.fasta - sed 's/>Cluster />Cluster_/g' ${params.projtag}_unlabeled_types.fasta.clstr >${params.projtag}_AminoTypes.clstr - grep ">Cluster_" ${params.projtag}_AminoTypes.clstr >tmpclusters.list - grep -w "*" ${params.projtag}_AminoTypes.clstr | awk '{print \$3}' | awk -F "." '{print \$1}' >tmphead.list - grep -w "*" ${params.projtag}_AminoTypes.clstr | awk '{print \$2}' | awk -F "," '{print \$1}' >tmplen.list - paste -d"," tmpclusters.list tmphead.list >tmp.info.csv - grep ">" ${params.projtag}_unlabeled_types.fasta >lala.list - j=1 - for x in \$(cat lala.list);do - echo ">${params.projtag}_AminoType\${j}" >>${params.projtag}_aminoheaders.list - echo "\${x},>${params.projtag}_AminoType\${j}" >>tmpaminotype.info.csv - j=\$(( \${j}+1 )) - done - rm lala.list - awk -F "," '{print \$2}' tmp.info.csv >>tmporder.list - for x in \$(cat tmporder.list);do - grep -w "\$x" tmpaminotype.info.csv | awk -F "," '{print \$2}' >>tmpder.list - done - paste -d "," tmpclusters.list tmplen.list tmphead.list tmpder.list >${params.projtag}_AminoType_summary_map.csv - rm tmp* - ./rename_seq.py ${params.projtag}_unlabeled_types.fasta ${params.projtag}_aminoheaders.list ${params.projtag}_AminoTypes_noTaxonomy.fasta - stats.sh in=${params.projtag}_AminoTypes_noTaxonomy.fasta gc=${params.projtag}_clustered.gc gcformat=4 - """ - } - - process Generate_AminoType_Matrix { - - label 'low_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Matrix", mode: "copy", overwrite: true - - input: - file(prot) from aminotypesClustal - - output: - file("*.matrix") into proclustmatrices - file("*PercentID.matrix") into aminotype_heatmap - - script: - """ - name=\$( echo ${prot} | awk -F ".fasta" '{print \$1}') - clustalo -i ${prot} --distmat-out=\${name}_PairwiseDistanceq.matrix --full --force --threads=${task.cpus} - clustalo -i ${prot} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} - for x in *q.matrix;do - pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') - ya=\$(wc -l \$x | awk '{print \$1}') - echo "\$((\$ya-1))" - tail -"\$(( \$ya-1))" \$x > \${pre}z.matrix - rm \$x - cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix - rm \${pre}z.matrix - done - """ - } - - if (!params.skipEMBOSS) { - - process AminoType_EMBOSS_Analyses { - - label 'low_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/2dStructure", mode: "copy", overwrite: true, pattern: '*.{garnier}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/HydrophobicMoment", mode: "copy", overwrite: true, pattern: '*HydrophobicMoments.{svg}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/IsoelectricPoint", mode: "copy", overwrite: true, pattern: '*IsoelectricPoint.{iep,svg}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/ProteinProperties", mode: "copy", overwrite: true, pattern: '*.{pepstats,pepinfo}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/ProteinProperties/Plots", mode: "copy", overwrite: true, pattern: '*PropertiesPlot.{svg}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/EMBOSS/2dStructure/Plots", mode: "copy", overwrite: true, pattern: '*Helical*.{svg}' - - input: - file(prot) from aminotypesEmboss - - output: - tuple file("*.garnier"), file("*HydrophobicMoments.svg"), file("*IsoelectricPoint*"), file("*.pepstats"), file("*PropertiesPlot*"), file("*Helical*") into amino_emboss - - script: - """ - name=\$( echo ${prot} | awk -F ".fasta" '{print \$1}') - garnier -sequence ${prot} -outfile \${name}_2dStructures.garnier - hmoment -seqall ${prot} -graph svg -plot - mv hmoment.svg ./"\${name}"_HydrophobicMoments.svg - iep -sequence ${prot} -graph svg -plot -outfile "\${name}"_IsoelectricPoint.iep - mv iep.svg ./"\${name}"_IsoelectricPoint.svg - pepstats -sequence ${prot} -outfile \${name}_ProteinProperties.pepstats - grep ">" ${prot} | awk -F ">" '{print \$2}' > tmpsequence.list - for x in \$(cat tmpsequence.list);do - echo \$x > tmp1.list - seqtk subseq ${prot} tmp1.list > tmp2.fasta - len=\$(tail -1 tmp2.fasta | awk '{print length}') - pepinfo -sequence tmp2.fasta -graph svg -outfile "\$x"_PropertiesPlot.pepinfo - mv pepinfo.svg ./"\$x"_PropertiesPlot.svg - cat "\$x"_PropertiesPlot.pepinfo >> "\${name}"_PropertiesPlot.pepinfo - rm "\$x"_PropertiesPlot.pepinfo - pepnet -sask -sequence tmp2.fasta -graph svg -sbegin1 1 -send1 \$len - mv pepnet.svg ./"\$x"_HelicalNet.svg - pepwheel -sequence tmp2.fasta -graph svg -sbegin1 1 -send1 \$len - mv pepwheel.svg ./"\$x"_HelicalWheel.svg - rm tmp1.list tmp2.fasta - done - rm tmpsequence.list - """ - } - } - - if (!params.skipTaxonomy) { - - process AminoType_Taxonomy_Inference { - - label 'high_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{csv,tsv}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Taxonomy", mode: "copy", overwrite: true, pattern: '*TaxonomyLabels.fasta' - - input: - file(reads) from aminotypesBlast - - output: - tuple file("*_phyloseqObject.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AA_diamond - file("*_summary_for_plot.csv") into taxplot2 - file("*TaxonomyLabels.fasta") into tax_labeled_fasta2 - - script: - """ - cp ${params.vampdir}/bin/rename_seq.py . - virdb=${params.dbdir}/${params.dbname} - grep ">" \${virdb} >> headers.list - headers="headers.list" - name=\$(ls ${reads} | awk -F "_noTaxonomy" '{print \$1}') - diamond blastp -q ${reads} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 - echo "Preparing lists to generate summary .csv's" - echo "[Best hit accession number]" >access.list - echo "[pcASV sequence length]" >length.list - echo "[e-value]" >evalue.list - echo "[Bitscore]" >bit.list - echo "[Percent ID (aa)]" >pid.list - echo "[AminoType#]" >otu.list - echo "[Virus ID]" >"\$name"_virus.list - echo "[Gene]" >"\$name"_genes.list - grep ">" ${reads} | awk -F ">" '{print \$2}' > seqids.lst - echo "extracting genes and names" - touch new_"\$name"_asvnames.txt - j=1 - for s in \$(cat seqids.lst);do - echo "Checking for \$s hit in diamond output" - if [[ ${params.refseq} == "T" ]];then - echo "RefSeq headers specified" - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >>evalue.list - echo "\$line" | awk '{print \$11}' >>bit.list - echo "\$line" | awk '{print \$12}' >>pid.list - echo "\$line" | awk '{print \$2}' >>length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">AminoType\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >>access.list - echo "NO_HIT" >>"\$name"_genes.list - echo "NO_HIT" >>"\$name"_virus.list - echo "NO_HIT" >>evalue.list - echo "NO_HIT" >>bit.list - echo "NO_HIT" >>pid.list - echo "NO_HIT" >>length.list - virus="NO" - gene="HIT" - echo ">AminoType\${j}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - fi - else - echo "Using RVDB headers." - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') - echo "\$s" >>otu.list - echo "\$acc" >>access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >>evalue.list - echo "\$line" | awk '{print \$11}' >>bit.list - echo "\$line" | awk '{print \$12}' >>pid.list - echo "\$line" | awk '{print \$2}' >>length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') - echo "\$gene" | sed 's/_/ /g' >>"\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') - echo "\$virus" | sed 's/_/ /g' >>"\$name"_virus.list - echo ">AminoType\${j}_"\$virus"_"\$gene"" >>new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >>otu.list - echo "NO_HIT" >>access.list - echo "NO_HIT" >>"\$name"_genes.list - echo "NO_HIT" >>"\$name"_virus.list - echo "NO_HIT" >>evalue.list - echo "NO_HIT" >>bit.list - echo "NO_HIT" >>pid.list - echo "NO_HIT" >>length.list - virus="NO" - gene="HIT" - echo ">AminoType\${j}_"\$virus"_"\$gene"" >>new_"\$name"_asvnames.txt - j=\$((\$j+1)) - echo "\$s done." - fi - fi - echo "Done with \$s" - done - echo "Now editing "\$name" fasta headers" - ###### rename_seq.py - ./rename_seq.py ${reads} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta - awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta > "\$name"_tmpssasv.fasta - echo "[Sequence header]" > newnames.list - cat new_"\$name"_asvnames.txt >> newnames.list - touch sequence.list - echo " " > sequence.list - grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list - rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - for x in *phyloseqObject.csv;do - echo "\$x" - lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) - tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; - sed 's/ /_/g' tmpcol.list > tmp2col.list; - cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; - rm tmpcol.list tmp2col.list - done - rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst headers.list otu.list ; - rm *asvnames.txt - rm "\$name"_virus.list - rm "\$name"_genes.list - rm newnames.list - rm access.list - echo "Taxonomy inferred for: ${reads} " - """ - } - } - - if (!params.skipPhylogeny) { - - process AminoType_Phylogeny { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*aln.*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Phylogeny/Modeltest", mode: "copy", overwrite: true, pattern: '*mt*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*iq*' - - input: - file(prot) from aminotypesMafft - - output: - tuple file("*_aln.fasta"), file("*_aln.html"), file("*.log"), file("*iq*"), file("*mt*") into alignprot_results - file("*iq.treefile") into amino_rax_plot - - script: - """ - # Protein_Alignment - pre=\$(echo ${prot} | awk -F ".fasta" '{print \$1}' ) - mafft --thread ${task.cpus} --maxiterate 15000 --auto ${prot} >\${pre}_ALN.fasta - trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html - - # Protein_ModelTest - modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d aa -s 203 --disable-checkpoint - - # Protein_Phylogeny - if [ "${params.iqCustomaa}" != "" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomaa} - - elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} - - elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni - - elif [ "${params.nonparametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} - - elif [ "${params.parametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - - else - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - fi - """ - } - } - - process Generate_AminoTypes_Counts_Table { - - label 'high_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/Counts", mode: "copy", overwrite: true - - input: - file(fasta) from aminotypesCounts - file(merged) from mergeforprotcounts - file(samplist) from samplelist + #alignment + mafft --thread ${task.cpus} --maxiterate 15000 --auto ${aminos} > ${params.projtag}_AminoTypes_mafftAlign.fasta + #trimming + trimal -in ${params.projtag}_AminoTypes_mafftAlign.fasta -out ${params.projtag}_AminoTypes_mafftAligned.fasta -keepheader -fasta -automated1 + rm ${params.projtag}_AminoTypes_mafftAlign.fasta + o-trim-uninformative-columns-from-alignment ${params.projtag}_AminoTypes_mafftAligned.fasta + mv ${params.projtag}_AminoTypes_mafftAligned.fasta-TRIMMED ./${params.projtag}_AminoTypes_Aligned_informativeonly.fasta + #entopy analysis + entropy-analysis ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta + #Decomposition + oligotype ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta-ENTROPY -o ${params.projtag}_AminoTypeMED_${params.aminoC} -M 1 -c ${params.aminoC} -N ${task.cpus} --skip-check-input --no-figures --skip-gen-html + #generatemaps + cd ./${params.projtag}_AminoTypeMED_${params.aminoC}/OLIGO-REPRESENTATIVES/ + echo "AminoType,Group,IDPattern" + j=1 + for x in *_unique; + do gid=\$(echo \$x | awk -F "_" '{print \$1}') + uni=\$(echo \$x | awk -F ""\${gid}"_" '{print \$2}' | awk -F "_uni" '{print \$1}') + grep ">" "\$gid"_"\$uni" | awk -F ">" '{print \$2}' > asv.list + seqtk subseq ../../${aminos} asv.list > Group"\${j}"_sequences.fasta + for z in \$( cat asv.list) + do echo ""\$z",Group"\$j","\$uni"" >> ${params.projtag}_AminoType_Grouping.csv + + done + rm asv.list + echo ">Group\${j}" >> ${params.projtag}_AminoType_group_reps_aligned.fasta + echo "\$uni" > group.list + seqtk subseq ../OLIGO-REPRESENTATIVES.fasta group.list > group.fasta + tail -1 group.fasta >> ${params.projtag}_AminoType_group_reps_aligned.fasta + mv "\$gid"_"\$uni" ./Group"\$j"_"\$uni"_aligned.fasta + mv "\$gid"_"\$uni"_unique ./Group"\$j"_"\$uni"_unqiues_aligned.fasta + rm "\$gid"*.cPickle + j=\$((\$j+1)) + done + mv ${params.projtag}_AminoType_Grouping.csv ../../ + mv ${params.projtag}_AminoType_group_reps_aligned.fasta ../../ + cd .. - output: - tuple file("*_protcounts.csv"), file("*dmd.out") into counts_summary - file("*_protcounts.csv") into aminocounts_plot + """ + } - script: - """ - set +e - diamond makedb --in ${fasta} --db ${fasta} - diamond blastx -q ${merged} -d ${fasta} -p ${task.cpus} --min-score ${params.ProtCountsBit} --id ${params.ProtCountID} -l ${params.ProtCountsLength} --more-sensitive -o ${params.projtag}_protCounts_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 --max-hsps 1 - echo "OTU_ID" >tmp.col1.txt - echo "Generating sample id list" - grep ">" ${fasta} | awk -F ">" '{print \$2}' | sort | uniq > otuid.list - cat otuid.list >> tmp.col1.txt - echo "Beginning them counts tho my g" - for y in \$( cat ${samplist} );do - echo "Starting with \$y now ..." - grep "\$y" ${params.projtag}_protCounts_dmd.out > tmp."\$y".out - echo "Isolated hits" - echo "Created uniq subject id list" - echo "\$y" > "\$y"_col.txt - echo "Starting my counts" - for z in \$(cat otuid.list);do - echo "Counting \$z hits" - echo "grep -wc "\$z" >> "\$y"_col.txt" - grep -wc "\$z" tmp."\$y".out >> "\$y"_col.txt - echo "\$z counted" - done - done - paste -d "," tmp.col1.txt *col.txt > ${params.projtag}_protcounts.csv - rm tmp* - rm *col.txt - """ - } - } + process AminoType_MED_Reps_phylogeny { - if (params.pcASV) { // ASV_nucl -> ASV_aa -> clusteraa by %id with ch-hit -> extract representative nucl sequences to generate new OTU file + label 'low_cpus' - if (params.sing) { + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/MED/Phylogeny/Modeltest", mode: "copy", overwrite: true, pattern: '*mt*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/MED/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*iq*' - process Translating_For_pcASV_Generation { + input: + file(reps) from atygroupreps - label 'low_cpus' + output: + file("*_AminoType_Group_Reps*") into align_results_aminmed + file("*iq.treefile") into amino_group_rep_tree - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV/Translation", mode: "copy", overwrite: true, pattern: '*_ASV_translations*' + script: + """ + # Protein_ModelTest + modeltest-ng -i ${reps} -p ${task.cpus} -o ${params.projtag}_AminoType_Group_Reps_mt -d aa -s 203 --disable-checkpoint - input: - file(fasta) from nucl2aa + # Protein_Phylogeny + if [ "${params.iqCustomaa}" != "" ];then + iqtree -s ${reps} --prefix ${params.projtag}_AminoType_Group_Reps_iq --redo -T auto ${params.iqCustomaa} - output: - file("*ASV*translations.fasta") into clustering_aa - file("*_ASV_translations_report") into reportaa_VR - file("*_ASV_nucleotide.fasta") into asvfastaforaaclust + elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then + mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') + iqtree -s ${reps} --prefix ${params.projtag}_AminoType_Group_Reps_iq -m \${mod} --redo -nt auto -b ${params.boots} - script: - """ - conda init && source activate virtualribosome + elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then + mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') + iqtree -s ${reps} --prefix ${params.projtag}_AminoType_Group_Reps_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni - ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_ASV_translations.fasta --report ${params.projtag}_ASV_translations_report - cp ${fasta} ${params.projtag}_ASV_nucleotide.fasta - """ + elif [ "${params.nonparametric}" != "false" ];then + iqtree -s ${reps} --prefix ${params.projtag}_AminoType_Group_Reps_iq -m MFP --redo -nt auto -b ${params.boots} - } + elif [ "${params.parametric}" != "false" ];then + iqtree -s ${reps} --prefix ${params.projtag}_AminoType_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - } else { + else + iqtree -s ${reps} --prefix ${params.projtag}_AminoType_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + fi + """ + } - process Translation_For_pcASV_Generation { + process Adding_AminoType_MED_Info { label 'low_cpus' - conda 'python=2.7' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV/Translation", mode: "copy", overwrite: true, pattern: '*_ASV_translations*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/MED/", mode: "copy", overwrite: true input: - file(fasta) from nucl2aa + file(counts) from aminocountmed + file(tree) from amino_repphy + file(map) from atygroupscsv output: - file("*ASV*translations.fasta") into clustering_aa - file("*_ASV_translations_report") into reportaa_VR - file("*_ASV_nucleotide.fasta") into asvfastaforaaclust + file("${params.projtag}_AminoType_Groupingcounts.csv") into amino_groupcounts script: """ - ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_ASV_translations.fasta --report ${params.projtag}_ASV_translations_report - cp ${fasta} ${params.projtag}_ASV_nucleotide.fasta + awk -F "," '{print \$1}' ${counts} | sed '1d' > amino.list + echo "GroupID" >> group.list + for x in \$(cat amino.list); + do group=\$(grep -w \$x ${map} | awk -F "," '{print \$2}') + echo "\$group" >> group.list + done + paste -d',' group.list ${counts} > ${params.projtag}_AminoType_Groupingcounts.csv """ - } - } - - process Generate_pcASVs { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV", mode: "copy", overwrite: true, pattern: '*pcASV*.{fasta}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{clstr,csv,gc}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV/Problematic", mode: "copy", overwrite: true, pattern: '*problem*.{fasta}' - - input: - file(fasta) from clustering_aa - file(asvs) from asvfastaforaaclust - - output: - file("${params.projtag}_nucleotide_pcASV*.fasta") into ( pcASV_ntDiamond_ch, pcASV_nt_counts_ch, pcASV_ntmatrix_ch, pcASV_ntmafft_ch ) - file("*_aminoacid_pcASV*_noTaxonomy.fasta") into ( pcASV_aaMatrix_ch, pcASV_aaDiamond_ch, pcASV_aaMafft_ch, pcASV_aaCounts_ch, pcASVEMBOSS ) - tuple file("*.fasta"), file("*.clstr"), file("*.csv"), file("*.gc") into ( pcASVsupplementalfiles ) - - script: - // add awk script to count seqs - if (params.clusterAAIDlist) { - """ - set +e - cp ${params.vampdir}/bin/rename_seq.py . - for id in `echo ${params.clusterAAIDlist} | tr "," "\\n"`;do - awk 'BEGIN{RS=">";ORS=""}length(\$2)>="${params.minAA}"{print ">"\$0}' ${fasta} > ${params.projtag}_filtered_proteins.fasta - cd-hit -i ${params.projtag}_filtered_proteins.fasta -c \${id} -o ${params.projtag}_pcASV\${id}.fasta - sed 's/>Cluster />Cluster_/g' ${params.projtag}_pcASV\${id}.fasta.clstr >${params.projtag}_pcASV\${id}.clstr - grep ">Cluster_" ${params.projtag}_pcASV\${id}.clstr >temporaryclusters.list - y=\$(grep -c ">Cluster_" ${params.projtag}_pcASV\${id}.clstr) - echo ">Cluster_"\${y}"" >> ${params.projtag}_pcASV\${id}.clstr - t=1 - b=1 - for x in \$(cat temporaryclusters.list);do - echo "Extracting \$x" - name="\$( echo \$x | awk -F ">" '{print \$2}')" - clust="pcASV"\${t}"" - echo "\${name}" - awk '/^>'\${name}'\$/,/^>Cluster_'\${b}'\$/' ${params.projtag}_pcASV\${id}.clstr > "\${name}"_"\${clust}"_tmp.list - t=\$(( \${t}+1 )) - b=\$(( \${b}+1 )) - done - ls *_tmp.list - u=1 - for x in *_tmp.list;do - name="\$(echo \$x | awk -F "_p" '{print \$1}')" - echo "\${name}" - cluster="\$(echo \$x | awk -F "_" '{print \$3}')" - echo "\${cluster}" - grep "ASV" \$x | awk -F ", " '{print \$2}' | awk -F "_" '{print \$1}' | awk -F ">" '{print \$2}' > \${name}_\${cluster}_seqs_tmps.list - seqtk subseq ${asvs} \${name}_\${cluster}_seqs_tmps.list > \${name}_\${cluster}_nucleotide_sequences.fasta - vsearch --cluster_fast \${name}_\${cluster}_nucleotide_sequences.fasta --id 0.2 --centroids \${name}_\${cluster}_centroids.fasta - grep ">" \${name}_\${cluster}_centroids.fasta >> \${name}_\${cluster}_tmp_centroids.list - for y in \$( cat \${name}_\${cluster}_tmp_centroids.list );do - echo ">\${cluster}_type"\$u"" >> \${name}_\${cluster}_tmp_centroid.newheaders - u=\$(( \${u}+1 )) - done - u=1 - ./rename_seq.py \${name}_\${cluster}_centroids.fasta \${name}_\${cluster}_tmp_centroid.newheaders \${cluster}_types_labeled.fasta - done - cat *_types_labeled.fasta >> ${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta - grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$3}' | awk -F "." '{print \$1}' >tmphead.list - grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$2}' | awk -F "," '{print \$1}' >tmplen.list - paste -d"," temporaryclusters.list tmphead.list >tmp.info.csv - grep ">" ${params.projtag}_pcASV\${id}.fasta >lala.list - j=1 - for x in \$(cat lala.list);do - echo ">${params.projtag}_pcASV\${j}" >>${params.projtag}_aminoheaders.list - echo "\${x},>${params.projtag}_pcASV\${j}" >>tmpaminotype.info.csv - j=\$(( \${j}+1 )) - done - rm lala.list - awk -F "," '{print \$2}' tmp.info.csv >>tmporder.list - for x in \$(cat tmporder.list);do - grep -w "\$x" tmpaminotype.info.csv | awk -F "," '{print \$2}' >>tmpder.list - done - paste -d "," temporaryclusters.list tmplen.list tmphead.list tmpder.list >${params.projtag}_pcASVCluster\${id}_summary.csv - ./rename_seq.py ${params.projtag}_pcASV\${id}.fasta ${params.projtag}_aminoheaders.list ${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta - stats.sh in=${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_aminoacid_clustered.gc gcformat=4 overwrite=true - stats.sh in=${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_nucleotide_clustered.gc gcformat=4 overwrite=true - awk 'BEGIN{RS=">";ORS=""}length(\$2)<"${params.minAA}"{print ">"\$0}' ${fasta} >${params.projtag}_pcASV\${id}_problematic_translations.fasta - if [ `wc -l ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk '{print \$1}'` -gt 1 ];then - grep ">" ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk -F ">" '{print \$2}' > problem_tmp.list - seqtk subseq ${asvs} > ${params.projtag}_pcASV\${id}_problematic_nucleotides.fasta - else - rm ${params.projtag}_pcASV\${id}_problematic_translations.fasta - fi - rm *.list - rm Cluster* - rm *types* - rm *tmp* - rm ${params.projtag}_pcASV\${id}.fast* - done - """ - } else if (params.clusterAAID) { - """ - set +e - cp ${params.vampdir}/bin/rename_seq.py . - id=${params.clusterAAID} - awk 'BEGIN{RS=">";ORS=""}length(\$2)>="${params.minAA}"{print ">"\$0}' ${fasta} > ${params.projtag}_filtered_proteins.fasta - cd-hit -i ${params.projtag}_filtered_proteins.fasta -c ${params.clusterAAID} -o ${params.projtag}_pcASV\${id}.fasta - sed 's/>Cluster />Cluster_/g' ${params.projtag}_pcASV\${id}.fasta.clstr >${params.projtag}_pcASV\${id}.clstr - grep ">Cluster_" ${params.projtag}_pcASV\${id}.clstr >temporaryclusters.list - y=\$(grep -c ">Cluster_" ${params.projtag}_pcASV\${id}.clstr) - echo ">Cluster_"\${y}"" >> ${params.projtag}_pcASV\${id}.clstr - t=1 - b=1 - for x in \$(cat temporaryclusters.list);do - echo "Extracting \$x" - name="\$( echo \$x | awk -F ">" '{print \$2}')" - clust="pcASV"\${t}"" - echo "\${name}" - awk '/^>'\${name}'\$/,/^>Cluster_'\${b}'\$/' ${params.projtag}_pcASV\${id}.clstr > "\${name}"_"\${clust}"_tmp.list - t=\$(( \${t}+1 )) - b=\$(( \${b}+1 )) - done - - ls *_tmp.list - u=1 - for x in *_tmp.list;do - name="\$(echo \$x | awk -F "_p" '{print \$1}')" - echo "\${name}" - cluster="\$(echo \$x | awk -F "_" '{print \$3}')" - echo "\${cluster}" - grep "ASV" \$x | awk -F ", " '{print \$2}' | awk -F "_" '{print \$1}' | awk -F ">" '{print \$2}' > \${name}_\${cluster}_seqs_tmps.list - seqtk subseq ${asvs} \${name}_\${cluster}_seqs_tmps.list > \${name}_\${cluster}_nucleotide_sequences.fasta - vsearch --cluster_fast \${name}_\${cluster}_nucleotide_sequences.fasta --id 0.2 --centroids \${name}_\${cluster}_centroids.fasta - grep ">" \${name}_\${cluster}_centroids.fasta >> \${name}_\${cluster}_tmp_centroids.list - for y in \$( cat \${name}_\${cluster}_tmp_centroids.list );do - echo ">\${cluster}_type"\$u"" >> \${name}_\${cluster}_tmp_centroid.newheaders - u=\$(( \${u}+1 )) - done - u=1 - ./rename_seq.py \${name}_\${cluster}_centroids.fasta \${name}_\${cluster}_tmp_centroid.newheaders \${cluster}_types_labeled.fasta - done - cat *_types_labeled.fasta >> ${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta - grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$3}' | awk -F "." '{print \$1}' >tmphead.list - grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$2}' | awk -F "," '{print \$1}' >tmplen.list - paste -d"," temporaryclusters.list tmphead.list >tmp.info.csv - grep ">" ${params.projtag}_pcASV\${id}.fasta >lala.list - j=1 - for x in \$(cat lala.list);do - echo ">${params.projtag}_pcASV\${j}" >>${params.projtag}_aminoheaders.list - echo "\${x},>${params.projtag}_pcASV\${j}" >>tmpaminotype.info.csv - j=\$(( \${j}+1 )) - done - rm lala.list - awk -F "," '{print \$2}' tmp.info.csv >>tmporder.list - for x in \$(cat tmporder.list);do - grep -w "\$x" tmpaminotype.info.csv | awk -F "," '{print \$2}' >>tmpder.list - done - paste -d "," temporaryclusters.list tmplen.list tmphead.list tmpder.list >${params.projtag}_pcASVCluster\${id}_summary.csv - ./rename_seq.py ${params.projtag}_pcASV\${id}.fasta ${params.projtag}_aminoheaders.list ${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta - stats.sh in=${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_aminoacid_clustered.gc gcformat=4 - stats.sh in=${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_nucleotide_clustered.gc gcformat=4 - awk 'BEGIN{RS=">";ORS=""}length(\$2)<"${params.minAA}"{print ">"\$0}' ${fasta} >${params.projtag}_pcASV\${id}_problematic_translations.fasta - if [ `wc -l ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk '{print \$1}'` -gt 1 ];then - grep ">" ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk -F ">" '{print \$2}' > problem_tmp.list - seqtk subseq ${asvs} problem_tmp.list > ${params.projtag}_pcASV\${id}_problematic_nucleotides.fasta - else - rm ${params.projtag}_pcASV\${id}_problematic_translations.fasta - fi - rm *.list - rm Cluster* - rm *types* - rm *tmp* - rm ${params.projtag}_pcASV\${id}.fast* - """ - } - } - - if (!params.skipTaxonomy) { - - process pcASV_Nucleotide_Taxonomy_Inference { - - label 'high_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{csv,tsv}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*dmd.{out}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{fasta}' - - input: - file(reads) from pcASV_ntDiamond_ch - - output: - file("*.fasta") into ( pcASV_labeled ) - tuple file("*_phyloseqObject.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AAdiamond - file("*_summary_for_plot.csv") into taxplot3 - - script: - """ - set +e - cp ${params.vampdir}/bin/rename_seq.py . - virdb=${params.dbdir}/${params.dbname} - grep ">" \${virdb} >> headers.list - headers="headers.list" - for filename in ${reads};do - name=\$(ls \${filename} | awk -F "_noTax" '{print \$1}') - diamond blastx -q \${filename} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 - echo "Preparing lists to generate summary .csv's" - echo "[Best hit accession number]" >access.list - echo "[pcASV sequence length]" >length.list - echo "[e-value]" >evalue.list - echo "[Bitscore]" >bit.list - echo "[Percent ID (aa)]" >pid.list - echo "[pcASV#]" >otu.list - echo "[Virus ID]" >"\$name"_virus.list - echo "[Gene]" >"\$name"_genes.list - grep ">" \${filename} | awk -F ">" '{print \$2}' > seqids.lst - echo "extracting genes and names" - touch new_"\$name"_headers.txt - j=1 - for s in \$(cat seqids.lst);do - echo "Checking for \$s hit in diamond output" - if [[ ${params.refseq} == "T" ]];then - echo "RefSeq headers specified" - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >>evalue.list - echo "\$line" | awk '{print \$11}' >>bit.list - echo "\$line" | awk '{print \$12}' >>pid.list - echo "\$line" | awk '{print \$2}' >>length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">pcASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_headers.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >>access.list - echo "NO_HIT" >>"\$name"_genes.list - echo "NO_HIT" >>"\$name"_virus.list - echo "NO_HIT" >>evalue.list - echo "NO_HIT" >>bit.list - echo "NO_HIT" >>pid.list - echo "NO_HIT" >>length.list - virus="NO" - gene="HIT" - echo ">pcASV\${j}_"\$virus"_"\$gene"" >> new_"\$name"_headers.txt - j=\$((\$j+1)) - echo "\$s done." - fi - else - echo "Using RVDB headers." - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') - echo "\$s" >>otu.list - echo "\$acc" >>access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >>evalue.list - echo "\$line" | awk '{print \$11}' >>bit.list - echo "\$line" | awk '{print \$12}' >>pid.list - echo "\$line" | awk '{print \$2}' >>length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >>"\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && - echo "\$virus" | sed 's/_/ /g' >>"\$name"_virus.list - echo ">pcASV\${j}_"\$virus"_"\$gene"" >>new_"\$name"_headers.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >>otu.list - echo "NO_HIT" >>access.list - echo "NO_HIT" >>"\$name"_genes.list - echo "NO_HIT" >>"\$name"_virus.list - echo "NO_HIT" >>evalue.list - echo "NO_HIT" >>bit.list - echo "NO_HIT" >>pid.list - echo "NO_HIT" >>length.list - virus="NO" - gene="HIT" - echo ">pcASV\${j}_"\$virus"_"\$gene"" >>new_"\$name"_headers.txt - j=\$((\$j+1)) - echo "\$s done." - fi - fi - echo "Done with \$s" - done - echo "Now editing "\$name" fasta headers" - ###### rename_seq.py - ./rename_seq.py \${filename} new_"\$name"_headers.txt "\$name"_TaxonomyLabels.fasta - awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta > "\$name"_tmpssasv.fasta - echo "[Sequence header]" > newnames.list - cat new_"\$name"_headers.txt >> newnames.list - touch sequence.list - echo " " > sequence.list - grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list - rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloseqObject.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - for x in *phyloseqObject.csv;do - echo "\$x" - lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) - tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; - sed 's/ /_/g' tmpcol.list > tmp2col.list; - cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; - rm tmpcol.list tmp2col.list - done - rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; - rm "\$name"_virus.list - rm "\$name"_genes.list - rm newnames.list - rm access.list - echo "Taxonomy inferred for: \${filename} " - done - rm *headers.list - """ - } - } - - process Generate_Nucleotide_pcASV_Counts { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Counts", mode: "copy", overwrite: true, pattern: '*.{biome,csv,txt}' - - input: - file(potus) from pcASV_nt_counts_ch - file(merged) from pcASV_mergedreads_ch - - output: - tuple file("*_counts.txt"), file("*_counts.biome") into pcASVcounts_vsearch - file("*.csv") into potu_Ncounts_for_report - - script: - """ - for filename in ${potus};do - ident=\$( echo \${filename} | awk -F "pcASV" '{print \$2}' | awk -F "_noTaxonomy.fasta" '{print \$1}') - name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') - vsearch --usearch_global ${merged} --db \${filename} --id \${ident} --threads ${task.cpus} --otutabout \${name}_counts.txt --biomout \${name}_counts.biome - cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv - sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv - rm \${name}_count.csv - done - """ - } - - process Generate_pcASV_Nucleotide_Matrix { - - label 'low_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Matrix", mode: "copy", overwrite: true - - input: - file(potus) from pcASV_ntmatrix_ch - - output: - file("*.matrix") into pcASVclustmatrices - file("*PercentID.matrix") into potu_nucl_heatmap - - script: - """ - for filename in ${potus};do - ident=\$( echo \${filename} | awk -F "pcASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') - name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') - clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistanceq.matrix --full --force --threads=${task.cpus} - clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} - for x in *q.matrix;do - pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') - ya=\$(wc -l \$x | awk '{print \$1}') - echo "\$((\$ya-1))" - tail -"\$((\$ya-1))" \$x > \${pre}z.matrix - rm \$x - cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix - rm \${pre}z.matrix - done - done - """ - } - - if (!params.skipPhylogeny) { - - process pcASV_Nucleotide_Phylogeny { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*aln.*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*mt*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*iq*' - - input: - file(reads) from pcASV_ntmafft_ch - - output: - tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into pcASV_nucleotide_phylogeny_results - file("*iq.treefile") into potu_Ntree_plot - - script: - """ - for filename in ${reads};do - pre=\$( echo \${filename} | awk -F "_noTax" '{print \$1}' ) - mafft --maxiterate 5000 --auto \${filename} >\${pre}_ALN.fasta - trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html - - # pcASV_Nucleotide_ModelTest - modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d nt -s 203 --disable-checkpoint - - # pcASV_Nucleotide_Phylogeny - if [ "${params.iqCustomnt}" != "" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -t \${pre}_mt.tree -T auto ${params.iqCustomnt} - - elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} - - elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni - - elif [ "${params.nonparametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -b ${params.boots} - - elif [ "${params.parametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni - - else - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -t \${pre}_mt.tree -nt auto -bb ${params.boots} -bnni - fi - done - """ - } - } - - process pcASV_AminoAcid_Matrix { - - label 'low_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Matrix", mode: "copy", overwrite: true - - input: - file(prot) from pcASV_aaMatrix_ch - - output: - file("*.matrix") into pcASVaaMatrix - file("*PercentID.matrix") into potu_aa_heatmap - - script: - """ - for filename in ${prot};do - name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') - clustalo -i \${filename} --distmat-out=\${name}_PairwiseDistanceq.matrix --full --force --threads=${task.cpus} - clustalo -i \${filename} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} - for x in *q.matrix;do - pre=\$(echo "\$x" | awk -F "q.matrix" '{print \$1}') - ya=\$(wc -l \$x | awk '{print \$1}') - echo "\$((\$ya-1))" - tail -"\$((\$ya-1))" \$x > \${pre}z.matrix - rm \$x - cat \${pre}z.matrix | sed 's/ /,/g' | sed -E 's/(,*),/,/g' >\${pre}.matrix - rm \${pre}z.matrix - done - done - """ - } - - if (!params.skipEMBOSS) { - - process pcASV_EMBOSS_Analyses { - - label 'low_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/2dStructure", mode: "copy", overwrite: true, pattern: '*.{garnier}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/HydrophobicMoment", mode: "copy", overwrite: true, pattern: '*HydrophobicMoments.{svg}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/IsoelectricPoint", mode: "copy", overwrite: true, pattern: '*IsoelectricPoint.{iep,svg}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/ProteinProperties", mode: "copy", overwrite: true, pattern: '*.{pepstats,pepinfo}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/ProteinProperties/Plots", mode: "copy", overwrite: true, pattern: '*PropertiesPlot.{svg}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/2dStructure/Plots", mode: "copy", overwrite: true, pattern: '*Helical*.{svg}' - - input: - file(prot) from pcASVEMBOSS - - output: - tuple file("*.garnier"), file("*HydrophobicMoments.svg"), file("*IsoelectricPoint*"), file("*.pepstats"), file("*PropertiesPlot*"), file("*Helical*") into pcASV_emboss - - script: - """ - for filename in ${prot};do - name=\$( echo \${filename} | awk -F ".fasta" '{print \$1}') - garnier -sequence \${filename} -outfile \${name}_2dStructures.garnier - hmoment -seqall \${filename} -graph svg -plot - mv hmoment.svg ./"\${name}"_HydrophobicMoments.svg - iep -sequence \${filename} -graph svg -plot -outfile "\${name}"_IsoelectricPoint.iep - mv iep.svg ./"\${name}"_IsoelectricPoint.svg - pepstats -sequence \${filename} -outfile \${name}_ProteinProperties.pepstats - grep ">" \${filename} | awk -F ">" '{print \$2}' > tmpsequence.list - for x in \$(cat tmpsequence.list);do - echo \$x > tmp1.list - seqtk subseq \${filename} tmp1.list > tmp2.fasta - len=\$(tail -1 tmp2.fasta | awk '{print length}') - pepinfo -sequence tmp2.fasta -graph svg -outfile "\$x"_PropertiesPlot.pepinfo - mv pepinfo.svg ./"\$x"_PropertiesPlot.svg - cat "\$x"_PropertiesPlot.pepinfo >> "\${name}"_PropertiesPlot.pepinfo - rm "\$x"_PropertiesPlot.pepinfo - pepnet -sask -sequence tmp2.fasta -graph svg -sbegin1 1 -send1 \$len - mv pepnet.svg ./"\$x"_HelicalNet.svg - pepwheel -sequence tmp2.fasta -graph svg -sbegin1 1 -send1 \$len - mv pepwheel.svg ./"\$x"_HelicalWheel.svg - rm tmp1.list tmp2.fasta - done - rm tmpsequence.list - done - """ - } - } - - if (!params.skipTaxonomy) { - - process pcASV_AminoAcid_Taxonomy_Inference { - - label 'high_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{csv,tsv}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*dmd.{out}' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{fasta}' - - input: - file(reads) from pcASV_aaDiamond_ch - - output: - file("*.fasta") into ( pcASV_labeledAA ) - tuple file("*phyloseqObject.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_potuaadiamond - file("*_summary_for_plot.csv") into taxplot4 - - script: - """ - cp ${params.vampdir}/bin/rename_seq.py . - virdb=${params.dbdir}/${params.dbname} - grep ">" \${virdb} >> headers.list - headers="headers.list" - for filename in ${reads};do - name=\$(ls \${filename} | awk -F ".fasta" '{print \$1}') - diamond blastp -q \${filename} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 - echo "Preparing lists to generate summary .csv's" - echo "[Best hit accession number]" >access.list - echo "[pcASV sequence length]" >length.list - echo "[e-value]" >evalue.list - echo "[Bitscore]" >bit.list - echo "[Percent ID (aa)]" >pid.list - echo "[pcASVaa#]" >otu.list - echo "[Virus ID]" >"\$name"_virus.list - echo "[Gene]" >"\$name"_genes.list - grep ">" \${filename} | awk -F ">" '{print \$2}' > seqids.lst - echo "extracting genes and names" - touch new_"\$name"_headers.txt - j=1 - for s in \$(cat seqids.lst);do - echo "Checking for \$s hit in diamond output" - if [[ ${params.refseq} == "T" ]];then - echo "RefSeq headers specified" - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') - echo "\$s" >> otu.list - echo "\$acc" >> access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >>evalue.list - echo "\$line" | awk '{print \$11}' >>bit.list - echo "\$line" | awk '{print \$12}' >>pid.list - echo "\$line" | awk '{print \$2}' >>length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') - echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list - echo ">pcASVaa\${j}_"\$virus"_"\$gene"" >> new_"\$name"_headers.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >> otu.list - echo "NO_HIT" >>access.list - echo "NO_HIT" >>"\$name"_genes.list - echo "NO_HIT" >>"\$name"_virus.list - echo "NO_HIT" >>evalue.list - echo "NO_HIT" >>bit.list - echo "NO_HIT" >>pid.list - echo "NO_HIT" >>length.list - virus="NO" - gene="HIT" - echo ">pcASVaa\${j}_"\$virus"_"\$gene"" >> new_"\$name"_headers.txt - j=\$((\$j+1)) - echo "\$s done." - fi - else - echo "Using RVDB headers." - if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then - echo "Yep, there was a hit for \$s" - echo "Extracting the information now:" - acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') - echo "\$s" >>otu.list - echo "\$acc" >>access.list - line="\$(grep -w "\$s" "\$name"_dmd.out)" - echo "\$line" | awk '{print \$10}' >>evalue.list - echo "\$line" | awk '{print \$11}' >>bit.list - echo "\$line" | awk '{print \$12}' >>pid.list - echo "\$line" | awk '{print \$2}' >>length.list - echo "Extracting virus and gene ID for \$s now" - gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && - echo "\$gene" | sed 's/_/ /g' >>"\$name"_genes.list - virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && - echo "\$virus" | sed 's/_/ /g' >>"\$name"_virus.list - echo ">pcASVaa\${j}_"\$virus"_"\$gene"" >>new_"\$name"_headers.txt - j=\$((\$j+1)) - echo "\$s done." - else - echo "Ugh, there was no hit for \$s .." - echo "We still love \$s though and we will add it to the final fasta file" - echo "\$s" >>otu.list - echo "NO_HIT" >>access.list - echo "NO_HIT" >>"\$name"_genes.list - echo "NO_HIT" >>"\$name"_virus.list - echo "NO_HIT" >>evalue.list - echo "NO_HIT" >>bit.list - echo "NO_HIT" >>pid.list - echo "NO_HIT" >>length.list - virus="NO" - gene="HIT" - echo ">pcASVaa\${j}_\${virus}_\${gene}" >>new_"\$name"_headers.txt - j=\$((\$j+1)) - echo "\$s done." - fi - fi - echo "Done with \$s" - done - echo "Now editing "\$name" fasta headers" - ###### rename_seq.py - ./rename_seq.py \${filename} new_"\$name"_headers.txt "\$name"_wTax.fasta - awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_wTax.fasta > "\$name"_tmpssasv.fasta - echo "[Sequence header]" > newnames.list - cat new_"\$name"_headers.txt >> newnames.list - touch sequence.list - awk 'BEGIN{RS=">";ORS=""}{print \$2"\\n"}' \${name}_tmpssasv.fasta >>sequence.list - rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_summary_phyloseqObject.csv - paste -d"\\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - for x in *phyloseqObject.csv;do - echo "\$x" - lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) - tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; - sed 's/ /_/g' tmpcol.list > tmp2col.list; - cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; - rm tmpcol.list tmp2col.list - done - rm evalue.list ; rm sequence.list ; rm bit.list ; rm pid.list ; rm length.list seqids.lst otu.list ; - rm "\$name"_virus.list - rm "\$name"_genes.list - rm newnames.list - rm access.list - echo "Taxonomy inferred for: \${filename} " - done - rm *headers.list - """ - } - } - - if (!params.skipPhylogeny) { - - process pcASV_Protein_Phylogeny { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*aln.*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Phylogeny/Modeltest", mode: "copy", overwrite: true, pattern: '*mt*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*iq*' - - input: - file(prot) from pcASV_aaMafft_ch - - output: - tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into pcASV_protein_phylogeny_results - file("*iq.treefile") into potu_Atree_plot - - script: - """ - for filename in ${prot};do - pre=\$( echo \${filename} | awk -F ".fasta" '{print \$1}' ) - mafft --maxiterate 5000 --auto \${filename} >\${pre}_ALN.fasta - trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html - - # pcASV_Protein_ModelTest - modeltest-ng -i \${pre}_aln.fasta -p ${task.cpus} -o \${pre}_mt -d aa -s 203 --disable-checkpoint - - # pcASV_Protein_Phylogeny - if [ "${params.iqCustomaa}" != "" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomaa} - - elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} - - elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 \${pre}_aln.fasta.log | head -1 | awk '{print \$6}') - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni - - elif [ "${params.nonparametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} - - elif [ "${params.parametric}" != "false" ];then - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - - else - iqtree -s \${pre}_aln.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - fi - done - """ - } - } - - process Generate_pcASV_Protein_Counts { - - label 'high_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Counts", mode: "copy", overwrite: true - - input: - file(fasta) from pcASV_aaCounts_ch - file(merged) from mergeforpcASVaacounts - file(samplist) from samplistpotu - - output: - tuple file("*_counts.csv"), file("*dmd.out") into potuaacounts_summary - file("*counts.csv") into potu_Acounts - - script: - """ - set +e - for filename in ${fasta};do - potu="\$( echo \${filename} | awk -F "_" '{print \$3}')" - diamond makedb --in \${filename} --db \${filename} - diamond blastx -q ${merged} -d \${filename} -p ${task.cpus} --min-score ${params.ProtCountsBit} --id ${params.ProtCountID} -l ${params.ProtCountsLength} --more-sensitive -o ${params.projtag}_\${potu}_Counts_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 --max-hsps 1 - echo "OTU_ID" >tmp.col1.txt - echo "Generating sample id list" - grep ">" \${filename} | awk -F ">" '{print \$2}' | sort | uniq > otuid.list - cat otuid.list >> tmp.col1.txt - echo "Beginning them counts tho my g" - for y in \$( cat ${samplist} );do - echo "Starting with \$y now ..." - grep "\$y" ${params.projtag}_\${potu}_Counts_dmd.out > tmp."\$y".out - echo "Isolated hits" - echo "Created uniq subject id list" - echo "\$y" > "\$y"_col.txt - echo "Starting my counts" - for z in \$(cat otuid.list);do - echo "Counting \$z hits" - echo "grep -wc "\$z" >> "\$y"_col.txt" - grep -wc "\$z" tmp."\$y".out >> "\$y"_col.txt - echo "\$z counted" - done - done - paste -d "," tmp.col1.txt *col.txt > ${params.projtag}_\${potu}_counts.csv - rm tmp* - rm *col.txt - done - """ - } - } - - if (!params.skipReport) { - - if (!params.skipAdapterRemoval) { - - process combine_csv { - - input: - file(csv) from fastp_csv - .collect() - - output: - file("final_reads_stats.csv") into ( fastp_csv1, fastp_csv2, fastp_csv3, fastp_csv4, fastp_csv5 ) - - script: - """ - cat ${csv} >all_reads_stats.csv - head -n1 all_reads_stats.csv >tmp.names.csv - cat all_reads_stats.csv | grep -v ""Sample,Total_"" >tmp.reads.stats.csv - cat tmp.names.csv tmp.reads.stats.csv >final_reads_stats.csv - rm tmp.names.csv tmp.reads.stats.csv - """ - - } - } - - if (params.ncASV) { - - - process Report_ASV { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport", mode: "copy", overwrite: true - - input: - file(counts) from asv_counts_plots - file(taxonomy) from taxplot1 - file(matrix) from asv_heatmap - file(readsstats) from fastp_csv1 - - output: - file("*.html") into report_summaryA + } + } - script: - """ - name=\$( echo ${taxonomy} | awk -F "_summary_for_plot.csv" '{print \$1}') - cp ${params.vampdir}/bin/vAMPirus_ReportA.Rmd . - cp ${params.vampdir}/example_data/conf/vamplogo.png . - Rscript -e "rmarkdown::render('vAMPirus_ReportA.Rmd',output_file='\${name}_ASV_Report.html')" \${name} \ - ${readsstats} \ - ${counts} \ - ${params.metadata} \ - ${params.minimumCounts} \ - ${matrix} \ - ${taxonomy} \ - ${params.trymax} \ - ${params.stats} - """ - } + if (params.pcASV) { // ASV_nucl -> ASV_aa -> clusteraa by %id with ch-hit -> extract representative nucl sequences to generate new OTU file - process Report_ncASV { + process Translation_For_pcASV_Generation { - label 'norm_cpus' + label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport/ncASV", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV/Translation", mode: "copy", overwrite: true, pattern: '*_ASV_translations*' - input: - file(counts) from notu_counts_plots - file(taxonomy) from taxplot1a - file(matrix) from notu_heatmap - file(phylogeny) from nucl_phyl_plot - file(readsstats) from fastp_csv2 + input: + file(fasta) from nucl2aa - output: - file("*.html") into report_summaryB + output: + file("*ASV*translations.fasta") into clustering_aa + file("*_ASV_translations_report") into reportaa_VR + file("*_ASV_nucleotide.fasta") into asvfastaforaaclust - script: - """ - cp ${params.vampdir}/bin/vAMPirus_ReportB.Rmd . - cp ${params.vampdir}/example_data/conf/vamplogo.png . - for x in *_summary_for_plot.csv;do - name=\$( echo \${x} | awk -F "_summary_for_plot.csv" '{print \$1}') - id=\$( echo \${x} | awk -F "_summary_for_plot.csv" '{print \$1}' | cut -f 2 -d "." ) - Rscript -e "rmarkdown::render('vAMPirus_ReportB.Rmd',output_file='\${name}_ncASV\${id}_Report.html')" \${name} \ - ${readsstats} \ - \$( echo ${counts} | tr " " "\\n" | grep "\${id}" ) \ - ${params.metadata} \ - ${params.minimumCounts} \ - \$( echo ${matrix} | tr " " "\\n" | grep "\${id}" ) \ - \$( echo ${taxonomy} | tr " " "\\n" | grep "\${id}" ) \ - \$( echo ${phylogeny} | tr " " "\\n" | grep "\${id}" ) \ - ${params.trymax} \ - ${params.stats} - done - """ + script: + """ + ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_ASV_translations.fasta --report ${params.projtag}_ASV_translations_report + cp ${fasta} ${params.projtag}_ASV_nucleotide.fasta + """ } - if (!params.skipAminoTyping) { - - process Report_AminoTypes { + process Generate_pcASVs { label 'norm_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport", mode: "copy", overwrite: true + tag "${mtag}" + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV", mode: "copy", overwrite: true, pattern: '*pcASV*.{fasta}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{clstr,csv,gc}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/pcASV/Problematic", mode: "copy", overwrite: true, pattern: '*problem*.{fasta}' input: - file(counts) from aminocounts_plot - file(taxonomy) from taxplot2 - file(matrix) from aminotype_heatmap - file(phylogeny) from amino_rax_plot - file(readsstats) from fastp_csv5 + each x from 1..naa + file(fasta) from clustering_aa + file(asvs) from asvfastaforaaclust output: - file("*.html") into report_summaryE + tuple nid, file("${params.projtag}_nucleotide_pcASV*.fasta") into ( pcASV_ntDiamond_ch, pcASV_nt_counts_ch, pcASV_ntmatrix_ch, pcASV_ntmafft_ch ) + tuple nid, file("*_aminoacid_pcASV*_noTaxonomy.fasta") into ( pcASV_aaMatrix_ch, pcASV_aaDiamond_ch, pcASV_aaMafft_ch, pcASV_aaCounts_ch, pcASVEMBOSS ) + tuple nid, file("*.fasta"), file("*.clstr"), file("*.csv"), file("*.gc") into ( pcASVsupplementalfiles ) script: + // add awk script to count seqs + nid=slist2.get(x-1) + mtag="ID=" + slist2.get(x-1) """ - name=\$( echo ${taxonomy} | awk -F "_summary_for_plot.csv" '{print \$1}') - cp ${params.vampdir}/bin/vAMPirus_ReportB.Rmd . - cp ${params.vampdir}/example_data/conf/vamplogo.png . - Rscript -e "rmarkdown::render('vAMPirus_ReportB.Rmd',output_file='\${name}_AminoType_Report.html')" \${name} \ - ${readsstats} \ - ${counts} \ - ${params.metadata} \ - ${params.minimumCounts} ${matrix} \ - ${taxonomy} \ - ${phylogeny} \ - ${params.trymax} \ - ${params.stats} + set +e + cp ${params.vampdir}/bin/rename_seq.py . + awk 'BEGIN{RS=">";ORS=""}length(\$2)>="${params.minAA}"{print ">"\$0}' ${fasta} > ${params.projtag}_filtered_proteins.fasta + cd-hit -i ${params.projtag}_filtered_proteins.fasta -c .${nid} -o ${params.projtag}_pcASV${nid}.fasta + sed 's/>Cluster />Cluster_/g' ${params.projtag}_pcASV${nid}.fasta.clstr >${params.projtag}_pcASV${nid}.clstr + grep ">Cluster_" ${params.projtag}_pcASV${nid}.clstr >temporaryclusters.list + y=\$(grep -c ">Cluster_" ${params.projtag}_pcASV${nid}.clstr) + echo ">Cluster_"\${y}"" >> ${params.projtag}_pcASV${nid}.clstr + t=1 + b=1 + for x in \$(cat temporaryclusters.list);do + echo "Extracting \$x" + name="\$( echo \$x | awk -F ">" '{print \$2}')" + clust="pcASV"\${t}"" + echo "\${name}" + awk '/^>'\${name}'\$/,/^>Cluster_'\${b}'\$/' ${params.projtag}_pcASV${nid}.clstr > "\${name}"_"\${clust}"_tmp.list + t=\$(( \${t}+1 )) + b=\$(( \${b}+1 )) + done + + ls *_tmp.list + u=1 + for x in *_tmp.list;do + name="\$(echo \$x | awk -F "_p" '{print \$1}')" + echo "\${name}" + cluster="\$(echo \$x | awk -F "_" '{print \$3}')" + echo "\${cluster}" + grep "ASV" \$x | awk -F ", " '{print \$2}' | awk -F "_" '{print \$1}' | awk -F ">" '{print \$2}' > \${name}_\${cluster}_seqs_tmps.list + seqtk subseq ${asvs} \${name}_\${cluster}_seqs_tmps.list > \${name}_\${cluster}_nucleotide_sequences.fasta + vsearch --cluster_fast \${name}_\${cluster}_nucleotide_sequences.fasta --id 0.2 --centroids \${name}_\${cluster}_centroids.fasta + grep ">" \${name}_\${cluster}_centroids.fasta >> \${name}_\${cluster}_tmp_centroids.list + for y in \$( cat \${name}_\${cluster}_tmp_centroids.list );do + echo ">\${cluster}_type"\$u"" >> \${name}_\${cluster}_tmp_centroid.newheaders + u=\$(( \${u}+1 )) + done + u=1 + ./rename_seq.py \${name}_\${cluster}_centroids.fasta \${name}_\${cluster}_tmp_centroid.newheaders \${cluster}_types_labeled.fasta + done + cat *_types_labeled.fasta >> ${params.projtag}_nucleotide_pcASV${nid}_noTaxonomy.fasta + grep -w "*" ${params.projtag}_pcASV${nid}.clstr | awk '{print \$3}' | awk -F "." '{print \$1}' >tmphead.list + grep -w "*" ${params.projtag}_pcASV${nid}.clstr | awk '{print \$2}' | awk -F "," '{print \$1}' >tmplen.list + paste -d"," temporaryclusters.list tmphead.list >tmp.info.csv + grep ">" ${params.projtag}_pcASV${nid}.fasta >lala.list + j=1 + for x in \$(cat lala.list);do + echo ">${params.projtag}_pcASV\${j}" >>${params.projtag}_aminoheaders.list + echo "\${x},>${params.projtag}_pcASV\${j}" >>tmpaminotype.info.csv + j=\$(( \${j}+1 )) + done + rm lala.list + awk -F "," '{print \$2}' tmp.info.csv >>tmporder.list + for x in \$(cat tmporder.list);do + grep -w "\$x" tmpaminotype.info.csv | awk -F "," '{print \$2}' >>tmpder.list + done + paste -d "," temporaryclusters.list tmplen.list tmphead.list tmpder.list >${params.projtag}_pcASVCluster${nid}_summary.csv + ./rename_seq.py ${params.projtag}_pcASV${nid}.fasta ${params.projtag}_aminoheaders.list ${params.projtag}_aminoacid_pcASV${nid}_noTaxonomy.fasta + stats.sh in=${params.projtag}_aminoacid_pcASV${nid}_noTaxonomy.fasta gc=${params.projtag}_pcASV${nid}_aminoacid_clustered.gc gcformat=4 + stats.sh in=${params.projtag}_nucleotide_pcASV${nid}_noTaxonomy.fasta gc=${params.projtag}_pcASV${nid}_nucleotide_clustered.gc gcformat=4 + awk 'BEGIN{RS=">";ORS=""}length(\$2)<"${params.minAA}"{print ">"\$0}' ${fasta} >${params.projtag}_pcASV${nid}_problematic_translations.fasta + if [ `wc -l ${params.projtag}_pcASV${nid}_problematic_translations.fasta | awk '{print \$1}'` -gt 1 ];then + grep ">" ${params.projtag}_pcASV${nid}_problematic_translations.fasta | awk -F ">" '{print \$2}' > problem_tmp.list + seqtk subseq ${asvs} problem_tmp.list > ${params.projtag}_pcASV${nid}_problematic_nucleotides.fasta + else + rm ${params.projtag}_pcASV${nid}_problematic_translations.fasta + fi + rm *.list + rm Cluster* + rm *types* + rm *tmp* + rm ${params.projtag}_pcASV${nid}.fast* """ } - } - } else { - process Report_ASVs { + if (!params.skipTaxonomy) { - label 'norm_cpus' + if (params.headers == "RefSeq") { + + process pcASV_Nucleotide_Taxonomy_Inference_RefSeq { - publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport", mode: "copy", overwrite: true + label 'high_cpus' + + tag "${mtag}" + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{csv,tsv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*dmd.{out}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{fasta}' input: - file(counts) from asv_counts_plots - file(taxonomy) from taxplot1 - file(matrix) from asv_heatmap - file(phylogeny) from nucl_phyl_plot - file(readsstats) from fastp_csv1 + tuple nid, file(asvs) from pcASV_ntDiamond_ch output: - file("*.html") into report_summaryA + file("*.fasta") into ( pcASV_labeled ) + tuple file("*_phyloformat.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AAdiamond + tuple nid, file("*_summary_for_plot.csv") into taxplot3 + tuple nid, file("*_quick_Taxbreakdown.csv") into tax_table_pcasvnt script: + mtag="ID=" + nid """ - name=\$( echo ${taxonomy} | awk -F "_summary_for_plot.csv" '{print \$1}') - cp ${params.vampdir}/bin/vAMPirus_ReportB.Rmd . - cp ${params.vampdir}/example_data/conf/vamplogo.png . - Rscript -e "rmarkdown::render('vAMPirus_ReportB.Rmd',output_file='\${name}_ASV_Report.html')" \${name} \ - ${readsstats} \ - ${counts} \ - ${params.metadata} \ - ${params.minimumCounts} \ - ${matrix} \ - ${taxonomy} \ - ${phylogeny} \ - ${params.trymax} \ - ${params.stats} + set +e + cp ${params.vampdir}/bin/rename_seq.py . + virdb=${params.dbdir}/${params.dbname} + grep ">" \${virdb} > headers.list + headers="headers.list" + name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') + diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + echo "Preparing lists to generate summary .csv's" + echo "[Best hit accession number]" > access.list + echo "[e-value]" > evalue.list + echo "[Bitscore]" > bit.list + echo "[Percent ID (aa)]" > pid.list + echo "[Organism ID]" > "\$name"_virus.list + echo "[Gene]" > "\$name"_genes.list + echo "[pcASV#]" > otu.list + echo "[Sequence length]" > length.list + grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst + echo "extracting genes and names" + touch new_"\$name"_asvnames.txt + for s in \$(cat seqids.lst);do + echo "Checking for \$s hit in diamond output" + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">"\${s}"_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list + virus="NO" + gene="HIT" + echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + echo "\$s done." + fi + done + echo "Now editing "\$name" fasta headers" + ###### rename_seq.py + ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta + echo "[Sequence header]" > newnames.list + cat new_"\$name"_asvnames.txt >> newnames.list + touch sequence.list + echo " " > sequence.list + grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list + rm "\$name"_tmpssasv.fasta + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + for x in *phyloformat.csv;do + echo "\$x" + lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) + tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; + sed 's/ /_/g' tmpcol.list > tmp2col.list; + cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; + rm tmpcol.list tmp2col.list + done + rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list """ - } - - if (!params.skipAminoTyping) { - - process Report_AminoType { - - label 'norm_cpus' - - publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport", mode: "copy", overwrite: true - - input: - file(counts) from aminocounts_plot - file(taxonomy) from taxplot2 - file(matrix) from aminotype_heatmap - file(phylogeny) from amino_rax_plot - file(readsstats) from fastp_csv5 - - output: - file("*.html") into report_summaryE - - script: - """ - name=\$( echo ${taxonomy} | awk -F "_summary_for_plot.csv" '{print \$1}') - cp ${params.vampdir}/bin/vAMPirus_ReportB.Rmd . - cp ${params.vampdir}/example_data/conf/vamplogo.png . - Rscript -e "rmarkdown::render('vAMPirus_ReportB.Rmd',output_file='\${name}_AminoType_Report.html')" \${name} \ - ${readsstats} \ - ${counts} \ - ${params.metadata} \ - ${params.minimumCounts} ${matrix} \ - ${taxonomy} \ - ${phylogeny} \ - ${params.trymax} \ - ${params.stats} - """ } + } else if (params.headers == "RVDB") { + + process pcASV_Nucleotide_Taxonomy_Inference_RVDB { + + label 'high_cpus' + + tag "${mtag}" + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{csv,tsv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*dmd.{out}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{fasta}' + + input: + tuple nid, file(asvs) from pcASV_ntDiamond_ch + + output: + file("*.fasta") into ( pcASV_labeled ) + tuple file("*_phyloformat.csv"), file("*_summaryTable.tsv"), file("*dmd.out") into summary_AAdiamond + tuple nid, file("*_summary_for_plot.csv") into taxplot3 + tuple nid, file("*_quick_Taxbreakdown.csv") into tax_table_pcasvnt + + script: + mtag="ID=" + nid + """ + set +e + cp ${params.vampdir}/bin/rename_seq.py . + virdb=${params.dbdir}/${params.dbname} + grep ">" \${virdb} > headers.list + headers="headers.list" + name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') + diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + echo "Preparing lists to generate summary .csv's" + echo "[Best hit accession number]" > access.list + echo "[e-value]" > evalue.list + echo "[Bitscore]" > bit.list + echo "[Percent ID (aa)]" > pid.list + echo "[Organism ID]" > "\$name"_virus.list + echo "[Gene]" > "\$name"_genes.list + echo "[pcASV#]" > otu.list + echo "[Sequence length]" > length.list + grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst + if [[ ${params.lca} == "T" ]] + then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list + echo "[Classification]" > classfication.list + else echo "skipped" >> \${name}_quick_Taxbreakdown.csv + echo "[Classification]" > classfication.list + fi + echo "extracting genes and names" + touch new_"\$name"_asvnames.txt + for s in \$(cat seqids.lst);do + echo "Using RVDB headers." + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + if [[ "${params.lca}" == "T" ]] + then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}') + lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') + echo "\$lcla" >> classfication.list + fi + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list + virus="NO" + gene="HIT" + echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + if [[ "${params.lca}" == "T" ]] + then echo "N/A" >> classfication.list + fi + echo "\$s done." + fi + echo "Done with \$s" + done + echo "Now editing "\$name" fasta headers" + ###### rename_seq.py + ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta + echo "[Sequence header]" > newnames.list + cat new_"\$name"_asvnames.txt >> newnames.list + touch sequence.list + echo " " > sequence.list + grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list + rm "\$name"_tmpssasv.fasta + if [[ "${params.lca}" == "T" ]] + then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list classfication.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv + else paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + fi + for x in *phyloformat.csv;do + echo "\$x" + lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) + tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; + sed 's/ /_/g' tmpcol.list > tmp2col.list; + cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; + rm tmpcol.list tmp2col.list + done + rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list + """ } } + } - if (params.pcASV) { - process Report_pcASV_AminoAcid { + process Generate_Nucleotide_pcASV_Counts { label 'norm_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport/pcASV/Aminoacid", mode: "copy", overwrite: true + tag "${mtag}" + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Counts", mode: "copy", overwrite: true, pattern: '*.{biome,csv,txt}' input: - file(counts) from potu_Acounts - file(taxonomy) from taxplot4 - file(matrix) from potu_aa_heatmap - file(phylogeny) from potu_Atree_plot - file(readsstats) from fastp_csv3 + tuple nid, file(potus) from pcASV_nt_counts_ch + file(merged) from pcASV_mergedreads_ch output: - file("*.html") into report_summaryC + tuple file("*_counts.txt"), file("*_counts.biome") into pcASVcounts_vsearch + tuple nid, file("*.csv") into potu_Ncounts_for_report script: + mtag="ID=" + nid """ - cp ${params.vampdir}/bin/vAMPirus_ReportB.Rmd . - cp ${params.vampdir}/example_data/conf/vamplogo.png . - for x in *_summary_for_plot.csv;do - name=\$( echo \${x} | awk -F "_noTaxonomy_summary_for_plot.csv" '{print \$1}') - id=\$( echo \${x} | awk -F "_noTaxonomy_summary_for_plot.csv" '{print \$1}' | cut -f 2 -d "." ) - Rscript -e "rmarkdown::render('vAMPirus_ReportB.Rmd',output_file='\${name}_pcASVaa\${id}_Report.html')" \${name} \ - ${readsstats} \ - \$( echo ${counts} | tr " " "\\n" | grep "\${id}" ) \ - ${params.metadata} \ - ${params.minimumCounts} \ - \$( echo ${matrix} | tr " " "\\n" | grep "\${id}" ) \ - \$( echo ${taxonomy} | tr " " "\\n" | grep "\${id}" ) \ - \$( echo ${phylogeny} | tr " " "\\n" | grep "\${id}" ) \ - ${params.trymax} \ - ${params.stats} - done + name=\$( echo ${potus} | awk -F ".fasta" '{print \$1}') + vsearch --usearch_global ${merged} --db ${potus} --id .${nid} --threads ${task.cpus} --otutabout \${name}_counts.txt --biomout \${name}_counts.biome + cat \${name}_counts.txt | tr "\t" "," >\${name}_count.csv + sed 's/#OTU ID/OTU_ID/g' \${name}_count.csv >\${name}_counts.csv + rm \${name}_count.csv """ - } + } - process Report_pcASV_Nucleotide { + process Generate_pcASV_Nucleotide_Matrix { - label 'norm_cpus' + label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReport/pcASV/Nucleotide", mode: "copy", overwrite: true + tag "${mtag}" + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Matrix", mode: "copy", overwrite: true input: - file(counts) from potu_Ncounts_for_report - file(taxonomy) from taxplot3 - file(matrix) from potu_nucl_heatmap - file(phylogeny) from potu_Ntree_plot - file(readsstats) from fastp_csv4 + tuple nid, file(potus) from pcASV_ntmatrix_ch output: - file("*.html") into report_summaryD + file("*.matrix") into pcASVclustmatrices + tuple nid, file("*PercentID.matrix") into potu_nucl_heatmap script: + //check --percent-id second clustalo + mtag="ID=" + nid """ - cp ${params.vampdir}/bin/vAMPirus_ReportB.Rmd . - cp ${params.vampdir}/example_data/conf/vamplogo.png . - for x in *_summary_for_plot.csv;do - name=\$( echo \${x} | awk -F "_summary_for_plot.csv" '{print \$1}') - id=\$( echo \${x} | awk -F "_summary_for_plot.csv" '{print \$1}' | cut -f 2 -d "." ) - Rscript -e "rmarkdown::render('vAMPirus_ReportB.Rmd',output_file='\${name}_pcASVnt\${id}_Report.html')" \${name} \ - ${readsstats} \ - \$( echo ${counts} | tr " " "\\n" | grep "\${id}" ) \ - ${params.metadata} \ - ${params.minimumCounts} \ - \$( echo ${matrix} | tr " " "\\n" | grep "\${id}" ) \ - \$( echo ${taxonomy} | tr " " "\\n" | grep "\${id}" ) \ - \$( echo ${phylogeny} | tr " " "\\n" | grep "\${id}" ) \ - ${params.trymax} \ - ${params.stats} - done + name=\$( echo ${potus} | awk -F ".fasta" '{print \$1}') + clustalo -i ${potus} --distmat-out=\${name}_PairwiseDistanceq.matrix --full --force --threads=${task.cpus} + clustalo -i ${potus} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} + cat \${name}_PercentIDq.matrix | tr " " "," | grep "," >\${name}_PercentID.matrix + rm \${name}_PercentIDq.matrix """ } - } - } - -} else if (params.DataCheck) { - - println("\n\tRunning vAMPirus DataCheck \n") - - process QualityCheck_1DC { - - label 'low_cpus' - - tag "${sample_id}" - - publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/FastQC/PreClean", mode: "copy", overwrite: true - - input: - tuple sample_id, file(reads) from reads_qc_ch - - output: - tuple sample_id, file("*_fastqc.{zip,html}") into fastqc_results_OAS - - script: - """ - fastqc --quiet --threads ${task.cpus} ${reads} - """ - } - - process Adapter_Removal_DC { - - label 'norm_cpus' - - tag "${sample_id}" - - publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/AdapterRemoval", mode: "copy", overwrite: true, pattern: "*.filter.fq" - publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/AdapterRemoval/fastpOut", mode: "copy", overwrite: true, pattern: "*.fastp.{json,html}" - - input: - tuple sample_id, file(reads) from reads_ch - - output: - tuple sample_id, file("*.fastp.{json,html}") into fastp_results - tuple sample_id, file("*.filter.fq") into reads_fastp_ch - file("*.csv") into fastp_csv - - script: - """ - echo ${sample_id} - - fastp -i ${reads[0]} -I ${reads[1]} -o left-${sample_id}.filter.fq -O right-${sample_id}.filter.fq --detect_adapter_for_pe \ - --average_qual 25 -c --overrepresentation_analysis --html ${sample_id}.fastp.html --json ${sample_id}.fastp.json --thread ${task.cpus} \ - --report_title ${sample_id} - - bash get_readstats.sh ${sample_id}.fastp.json - """ - } - - process Primer_Removal_DC { - label 'norm_cpus' - - tag "${sample_id}" - - publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/PrimerRemoval", mode: "copy", overwrite: true - - input: - tuple sample_id, file(reads) from reads_fastp_ch - - output: - tuple sample_id, file("*bbduk*.fastq.gz") into ( reads_bbduk_ch, readsforqc2 ) - - script: - // add cocktail primer removal, easy, set a list, line by line seperated by , in a for loop - if ( params.fwd == "" && params.rev == "" && !params.multi) { - """ - bbduk.sh in1=${reads[0]} out=${sample_id}_bb_R1.fastq.gz ftl=${params.defaultFwdTrim} t=${task.cpus} - bbduk.sh in=${reads[1]} out=${sample_id}_bb_R2.fastq.gz ftl=${params.defaultRevTrim} t=${task.cpus} - repair.sh in1=${sample_id}_bb_R1.fastq.gz in2=${sample_id}_bb_R2.fastq.gz out1=${sample_id}_bbduk_R1.fastq.gz out2=${sample_id}_bbduk_R2.fastq.gz minlength=${params.minilen} outs=sing.fq repair - """ - } else if ( params.GlobTrim && !params.GlobTrim == "" ) { - """ - FTRIM=\$( echo ${GlobTrim} | cut -f 1 -d "," ) - RTRIM=\$( echo ${GlobTrim} | cut -f 2 -d "," ) - bbduk.sh in=${reads[0]} out=${sample_id}_bb_R1.fastq.gz ftl=\${FTRIM} t=${task.cpus} - bbduk.sh in=${reads[1]} out=${sample_id}_bb_R2.fastq.gz ftl=\${RTRIM} t=${task.cpus} - repair.sh in1=${sample_id}_bb_R1.fastq.gz in2=${sample_id}_bb_R2.fastq.gz out1=${sample_id}_bbduk_R1.fastq.gz out2=${sample_id}_bbduk_R2.fastq.gz minlength=${params.minilen} outs=sing.fq repair - """ - } else if ( params.multi && params.primers ) { - """ - bbduk.sh in=${reads[0]} in2=${reads[1]} out=${sample_id}_bbduk_R1.fastq.gz out2=${sample_id}_bbduk_R2.fastq.gz ref=${params.primers} copyundefined=t t=${task.cpus} restrictleft=${params.primerLength} k=${params.maxkmer} ordered=t mink=${params.minkmer} ktrim=l ecco=t rcomp=t minlength=${params.minilen} tbo tpe - """ - } else { - """ - bbduk.sh in=${reads[0]} in2=${reads[1]} out=${sample_id}_bbduk_R1.fastq.gz out2=${sample_id}_bbduk_R2.fastq.gz literal=${params.fwd},${params.rev} copyundefined=t t=${task.cpus} restrictleft=${params.primerLength} k=${params.maxkmer} ordered=t mink=${params.minkmer} ktrim=l ecco=t rcomp=t minlength=${params.minilen} tbo tpe - """ - } - } - - process QualityCheck_2_DC { - - label 'low_cpus' - - tag "${sample_id}" - - publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/FastQC/PostClean", mode: "copy", overwrite: true - - input: - tuple sample_id, file(reads) from readsforqc2 - - output: - tuple sample_id, file("*_fastqc.{zip,html}") into fastqc2_results_OAS - - script: - """ - fastqc --quiet --threads ${task.cpus} ${reads} - """ - } - - process Read_Merging_DC { - - label 'norm_cpus' - - tag "${sample_id}" - - publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/ReadMerging/Individual", mode: "copy", overwrite: true, pattern: "*mergedclean.fastq" - publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/ReadMerging/Individual/notmerged", mode: "copy", overwrite: true, pattern: "*notmerged*.fastq" - - input: - tuple sample_id, file(reads) from reads_bbduk_ch + if (!params.skipPhylogeny) { - output: - file("*_mergedclean.fastq") into reads_vsearch1_ch - file("*.name") into names - file("*notmerged*.fastq") into notmerged - - script: - """ - vsearch --fastq_mergepairs ${reads[0]} --reverse ${reads[1]} --threads ${task.cpus} --fastqout ${sample_id}_mergedclean.fastq --fastqout_notmerged_fwd ${sample_id}_notmerged_fwd.fastq --fastqout_notmerged_rev ${sample_id}_notmerged_rev.fastq --fastq_maxee ${params.maxEE} --relabel ${sample_id}. - echo ${sample_id} > ${sample_id}.name - """ - - } + process pcASV_Nucleotide_Phylogeny { - process Compile_Reads_DC { + label 'norm_cpus' - label 'low_cpus' + tag "${mtag}" - publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/ReadMerging/LengthFiltering", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*aln.*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*mt*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*iq*' - input: - file(reads) from reads_vsearch1_ch - .collect() + input: + tuple nid, file(reads) from pcASV_ntmafft_ch - output: - file("*_all_merged_preFilt_preClean.fastq") into collect_samples_ch + output: + tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into pcASV_nucleotide_phylogeny_results + tuple nid, file("*iq.treefile") into potu_Ntree_plot - script: - """ - cat ${reads} >>${params.projtag}_all_merged_preFilt_preClean.fastq - """ - } + script: + mtag="ID=" + nid + """ + pre=\$( echo ${reads} | awk -F "_noTax" '{print \$1}' ) + mafft --maxiterate 5000 --auto ${reads} >\${pre}_ALN.fasta + trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html + o-trim-uninformative-columns-from-alignment \${pre}_aln.fasta + mv \${pre}_aln.fasta-TRIMMED ./\${pre}_Aligned_informativeonly.fasta + # pcASV_Nucleotide_ModelTest + modeltest-ng -i \${pre}_Aligned_informativeonly.fasta -p ${task.cpus} -o \${pre}_noTaxonomy_mt -d nt -s 203 --disable-checkpoint - process Compile_Names_DC { + # pcASV_Nucleotide_Phylogeny + if [ "${params.iqCustomnt}" != "" ];then + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_noTaxonomy_iq --redo -T auto ${params.iqCustomnt} - label 'low_cpus' + elif [[ "${params.ModelTnt}" != "false" && "${params.nonparametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_noTaxonomy_iq -m \${mod} --redo-nt auto -b ${params.boots} - publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/ReadMerging", mode: "copy", overwrite: true + elif [[ "${params.ModelTnt}" != "false" && "${params.parametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_noTaxonomy_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni - input: - file(names) from names - .collect() + elif [ "${params.nonparametric}" != "false" ];then + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_noTaxonomy_iq -m MFP --redo -nt auto -b ${params.boots} - output: - file("*sample_ids.list") into ( samplelist, samplistpotu ) + elif [ "${params.parametric}" != "false" ];then + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_noTaxonomy_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - script: - """ - cat ${names} >>${params.projtag}_sample_ids.list - """ - } + else + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_noTaxonomy_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + fi + """ + } + } - process Length_Filtering_DC { + process pcASV_AminoAcid_Matrix { - label 'norm_cpus' + label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/ReadMerging/LengthFiltering", mode: "copy", overwrite: true, pattern: "*_merged_preFilt*.fasta" - publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/ReadMerging", mode: "copy", overwrite: true, pattern: "*Lengthfiltered.fastq" - publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/ReadMerging/Histograms/pre_length_filtering", mode: "copy", overwrite: true, pattern: "*preFilt_*st.txt" - publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/ReadMerging/Histograms/post_length_filtering", mode: "copy", overwrite: true, pattern: "*postFilt_*st.txt" + tag "${mtag}" - input: - file(reads) from collect_samples_ch + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Matrix", mode: "copy", overwrite: true - output: - file("*_merged_preFilt_clean.fasta") into ( nuclCounts_mergedreads_ch, pcASV_mergedreads_ch ) - file("*_merged_clean_Lengthfiltered.fastq") into reads_vsearch2_ch - - file("*preFilt_preClean_baseFrequency_hist.csv") into prefilt_basefreq - file("*preFilt_preClean_qualityScore_hist.csv") into prefilt_qualityscore - file("*preFilt_preClean_gcContent_hist.csv") into prefilt_gccontent - file("*preFilt_preClean_averageQuality_hist.csv") into prefilt_averagequality - file("*preFilt_preClean_length_hist.csv") into prefilt_length - - file("*postFilt_baseFrequency_hist.csv") into postFilt_basefreq - file("*postFilt_qualityScore_hist.csv") into postFilt_qualityscore - file("*postFilt_gcContent_hist.csv") into postFilt_gccontent - file("*postFilt_averageQuaulity_hist.csv") into postFilt_averagequality - file("*postFilt_length_hist.csv") into postFilt_length - - file("reads_per_sample_preFilt_preClean.csv") into reads_per_sample_preFilt - file("read_per_sample_postFilt_postClean.csv") into reads_per_sample_postFilt - script: - """ - bbduk.sh in=${reads} bhist=${params.projtag}_all_merged_preFilt_preClean_baseFrequency_hist.txt qhist=${params.projtag}_all_merged_preFilt_preClean_qualityScore_hist.txt gchist=${params.projtag}_all_merged_preFilt_preClean_gcContent_hist.txt aqhist=${params.projtag}_all_merged_preFilt_preClean_averageQuality_hist.txt lhist=${params.projtag}_all_merged_preFilt_preClean_length_hist.txt gcbins=auto - for x in *preFilt*hist.txt;do - pre=\$(echo \$x | awk -F ".txt" '{print \$1}') - cat \$x | tr "\t" "," > \${pre}.csv - rm \$x - done - reformat.sh in=${reads} out=${params.projtag}_preFilt_preclean.fasta t=${task.cpus} - echo "sample,reads" >> reads_per_sample_preFilt_preClean.csv - grep ">" ${params.projtag}_preFilt_preclean.fasta | awk -F ">" '{print \$2}' | awk -F "." '{print \$1}' | sort --parallel=${task.cpus} | uniq -c | sort -brg --parallel=${task.cpus} | awk '{print \$2","\$1}' >> reads_per_sample_preFilt_preClean.csv - rm ${params.projtag}_preFilt_preclean.fasta - fastp -i ${reads} -o ${params.projtag}_merged_preFilt_clean.fastq -b ${params.maxLen} -l ${params.minLen} --thread ${task.cpus} -n 1 - reformat.sh in=${params.projtag}_merged_preFilt_clean.fastq out=${params.projtag}_merged_preFilt_clean.fasta t=${task.cpus} - bbduk.sh in=${params.projtag}_merged_preFilt_clean.fastq out=${params.projtag}_merged_clean_Lengthfiltered.fastq minlength=${params.maxLen} maxlength=${params.maxLen} t=${task.cpus} - bbduk.sh in=${params.projtag}_merged_clean_Lengthfiltered.fastq bhist=${params.projtag}_all_merged_postFilt_baseFrequency_hist.txt qhist=${params.projtag}_all_merged_postFilt_qualityScore_hist.txt gchist=${params.projtag}_all_merged_postFilt_gcContent_hist.txt aqhist=${params.projtag}_all_merged_postFilt_averageQuaulity_hist.txt lhist=${params.projtag}_all_merged_postFilt_length_hist.txt gcbins=auto - for x in *postFilt*hist.txt;do - pre=\$(echo \$x | awk -F ".txt" '{print \$1}') - cat \$x | tr "\t" "," > \${pre}.csv - rm \$x - done - reformat.sh in=${params.projtag}_merged_clean_Lengthfiltered.fastq out=${params.projtag}_merged_clean_Lengthfiltered.fasta t=${task.cpus} - echo "sample,reads" >> read_per_sample_postFilt_postClean.csv - grep ">" ${params.projtag}_merged_clean_Lengthfiltered.fasta | awk -F ">" '{print \$2}' | awk -F "." '{print \$1}' | sort --parallel=${task.cpus} | uniq -c | sort -brg --parallel=${task.cpus} | awk '{print \$2","\$1}' >> read_per_sample_postFilt_postClean.csv - """ - } + input: + tuple nid, file(prot) from pcASV_aaMatrix_ch - process Extract_Uniques_DC { + output: + file("*.matrix") into pcASVaaMatrix + tuple nid, file("*PercentID.matrix") into potu_aa_heatmap - label 'norm_cpus' + script: + mtag="ID=" + nid + """ + name=\$( echo ${prot} | awk -F ".fasta" '{print \$1}') + clustalo -i ${prot} --distmat-out=\${name}_PairwiseDistanceq.matrix --full --force --threads=${task.cpus} + clustalo -i ${prot} --distmat-out=\${name}_PercentIDq.matrix --percent-id --full --force --threads=${task.cpus} + cat \${name}_PercentIDq.matrix | tr " " "," | grep "," >\${name}_PercentID.matrix + rm \${name}_PercentIDq.matrix + """ + } - publishDir "${params.workingdir}/${params.outdir}/DataCheck/ReadProcessing/ReadMerging/Uniques", mode: "copy", overwrite: true + if (!params.skipEMBOSS) { - input: - file(reads) from reads_vsearch2_ch + process pcASV_EMBOSS_Analyses { - output: - file("*unique_sequences.fasta") into reads_vsearch3_ch + label 'low_cpus' - script: - """ - vsearch --derep_fulllength ${reads} --sizeout --relabel_keep --output ${params.projtag}_unique_sequences.fasta - """ - } + tag "${mtag}" - process Identify_ASVs_DC { + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/2dStructure", mode: "copy", overwrite: true, pattern: '*.{garnier}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/HydrophobicMoment", mode: "copy", overwrite: true, pattern: '*HydrophobicMoments.{svg}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/IsoelectricPoint", mode: "copy", overwrite: true, pattern: '*IsoelectricPoint.{iep,svg}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/ProteinProperties", mode: "copy", overwrite: true, pattern: '*.{pepstats,pepinfo}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/ProteinProperties/Plots", mode: "copy", overwrite: true, pattern: '*PropertiesPlot.{svg}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/EMBOSS/2dStructure/Plots", mode: "copy", overwrite: true, pattern: '*Helical*.{svg}' - label 'norm_cpus' + input: + tuple nid, file(prot) from pcASVEMBOSS - publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/ASVs/ChimeraCheck", mode: "copy", overwrite: true + output: + tuple file("*.garnier"), file("*HydrophobicMoments.svg"), file("*IsoelectricPoint*"), file("*.pepstats"), file("*PropertiesPlot*"), file("*Helical*") into pcASV_emboss - input: - file(reads) from reads_vsearch3_ch + script: + // check do I need for loop + mtag="ID=" + nid + """ + name=\$( echo ${prot} | awk -F ".fasta" '{print \$1}') + garnier -sequence ${prot} -outfile \${name}_2dStructures.garnier + hmoment -seqall ${prot} -graph svg -plot + mv hmoment.svg ./"\${name}"_HydrophobicMoments.svg + iep -sequence ${prot} -graph svg -plot -outfile "\${name}"_IsoelectricPoint.iep + mv iep.svg ./"\${name}"_IsoelectricPoint.svg + pepstats -sequence ${prot} -outfile \${name}_ProteinProperties.pepstats + grep ">" ${prot} | awk -F ">" '{print \$2}' > tmpsequence.list + for x in \$(cat tmpsequence.list);do + echo \$x > tmp1.list + seqtk subseq ${prot} tmp1.list > tmp2.fasta + len=\$(tail -1 tmp2.fasta | awk '{print length}') + pepinfo -sequence tmp2.fasta -graph svg -outfile "\$x"_PropertiesPlot.pepinfo + mv pepinfo.svg ./"\$x"_PropertiesPlot.svg + cat "\$x"_PropertiesPlot.pepinfo >> "\${name}"_PropertiesPlot.pepinfo + rm "\$x"_PropertiesPlot.pepinfo + pepnet -sask -sequence tmp2.fasta -graph svg -sbegin1 1 -send1 \$len + mv pepnet.svg ./"\$x"_HelicalNet.svg + pepwheel -sequence tmp2.fasta -graph svg -sbegin1 1 -send1 \$len + mv pepwheel.svg ./"\$x"_HelicalWheel.svg + rm tmp1.list tmp2.fasta + done + rm tmpsequence.list + """ + } + } - output: - file("*notChecked.fasta") into reads_vsearch4_ch + if (!params.skipTaxonomy) { - script: - """ - vsearch --cluster_unoise ${reads} --unoise_alpha ${params.alpha} --relabel ASV --centroids ${params.projtag}_notChecked.fasta --minsize ${params.minSize} - """ - } + if (params.headers == "RefSeq") { - process Chimera_Check_DC { + process pcASV_AminoAcid_Taxonomy_Inference_RefSeq { - label 'norm_cpus' + label 'high_cpus' - publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/ASVs", mode: "copy", overwrite: true + tag "${mtag}" - input: - file(fasta) from reads_vsearch4_ch + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{csv,tsv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*dmd.{out}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{fasta}' - output: - file("*ASVs.fasta") into ( reads_vsearch5_ch, nucl2aa, asvsforAminotyping, asvfastaforcounts, asvaminocheck ) + input: + tuple nid, file(asvs) from pcASV_aaDiamond_ch - script: - """ - vsearch --uchime3_denovo ${fasta} --relabel ASV --nonchimeras ${params.projtag}_ASVs.fasta - """ - } + output: + file("*.fasta") into ( pcASV_labeledAA ) + tuple file("*phyloformat.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_potuaadiamond + tuple nid, file("*_summary_for_plot.csv") into taxplot4 + tuple nid, file("*_quick_Taxbreakdown.csv") into tax_table_pcasvaa - process NucleotideBased_ASV_clustering_DC { + script: + mtag="ID=" + nid + """ + cp ${params.vampdir}/bin/rename_seq.py . + virdb=${params.dbdir}/${params.dbname} + grep ">" \${virdb} > headers.list + headers="headers.list" + name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') + diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + echo "Preparing lists to generate summary .csv's" + echo "[Best hit accession number]" > access.list + echo "[e-value]" > evalue.list + echo "[Bitscore]" > bit.list + echo "[Percent ID (aa)]" > pid.list + echo "[Organism ID]" > "\$name"_virus.list + echo "[Gene]" > "\$name"_genes.list + echo "[pcASV#]" > otu.list + echo "[Sequence length]" > length.list + grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst + echo "extracting genes and names" + touch new_"\$name"_asvnames.txt + for s in \$(cat seqids.lst);do + echo "Checking for \$s hit in diamond output" + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "." '{ print \$2 }' | awk -F "[" '{ print \$1 }' | awk -F " " print substr(\$0, index(\$0,\$2)) | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">"\${s}"_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list + virus="NO" + gene="HIT" + echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + echo "\$s done." + fi + done + echo "Now editing "\$name" fasta headers" + ###### rename_seq.py + ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta + echo "[Sequence header]" > newnames.list + cat new_"\$name"_asvnames.txt >> newnames.list + touch sequence.list + echo " " > sequence.list + grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list + rm "\$name"_tmpssasv.fasta + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + for x in *phyloformat.csv;do + echo "\$x" + lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) + tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; + sed 's/ /_/g' tmpcol.list > tmp2col.list; + cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; + rm tmpcol.list tmp2col.list + done + rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list + """ + } + } else if (params.headers == "RVDB") { + + process pcASV_AminoAcid_Taxonomy_Inference_RVDB { + + label 'high_cpus' + + tag "${mtag}" + + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy/SummaryFiles", mode: "copy", overwrite: true, pattern: '*.{csv,tsv}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy/DiamondOutput", mode: "copy", overwrite: true, pattern: '*dmd.{out}' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Taxonomy", mode: "copy", overwrite: true, pattern: '*.{fasta}' + + input: + tuple nid, file(asvs) from pcASV_aaDiamond_ch + + output: + file("*.fasta") into ( pcASV_labeledAA ) + tuple file("*phyloformat.csv"), file("*summaryTable.tsv"), file("*dmd.out") into summary_potuaadiamond + tuple nid, file("*_summary_for_plot.csv") into taxplot4 + tuple nid, file("*_quick_Taxbreakdown.csv") into tax_table_pcasvaa + + script: + mtag="ID=" + nid + """ + cp ${params.vampdir}/bin/rename_seq.py . + virdb=${params.dbdir}/${params.dbname} + grep ">" \${virdb} > headers.list + headers="headers.list" + name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') + diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + echo "Preparing lists to generate summary .csv's" + echo "[Best hit accession number]" > access.list + echo "[e-value]" > evalue.list + echo "[Bitscore]" > bit.list + echo "[Percent ID (aa)]" > pid.list + echo "[Organism ID]" > "\$name"_virus.list + echo "[Gene]" > "\$name"_genes.list + echo "[pcASV#]" > otu.list + echo "[Sequence length]" > length.list + grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst + if [[ ${params.lca} == "T" ]] + then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list + echo "[Classification]" > classfication.list + else echo "skipped" >> \${name}_quick_Taxbreakdown.csv + echo "[Classification]" > classfication.list + fi + echo "extracting genes and names" + touch new_"\$name"_asvnames.txt + for s in \$(cat seqids.lst);do + echo "Using RVDB headers." + if [[ "\$(grep -wc "\$s" "\$name"_dmd.out)" -eq 1 ]];then + echo "Yep, there was a hit for \$s" + echo "Extracting the information now:" + acc=\$(grep -w "\$s" "\$name"_dmd.out | awk '{print \$3}' | awk -F "|" '{print \$3}') + echo "\$s" >> otu.list + echo "\$acc" >> access.list + line="\$(grep -w "\$s" "\$name"_dmd.out)" + echo "\$line" | awk '{print \$10}' >> evalue.list + echo "\$line" | awk '{print \$11}' >> bit.list + echo "\$line" | awk '{print \$12}' >> pid.list + echo "\$line" | awk '{print \$2}' >> length.list + echo "Extracting virus and gene ID for \$s now" + gene=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$1 }' | sed 's/ /_/g') && + echo "\$gene" | sed 's/_/ /g' >> "\$name"_genes.list + virus=\$(grep -w "\$acc" "\$headers" | awk -F "|" '{ print \$6 }' | awk -F "[" '{ print \$2 }' | awk -F "]" '{print \$1}' | sed 's/ /_/g') && + echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list + echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + if [[ "${params.lca}" == "T" ]] + then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}') + lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') + echo "\$lcla" >> classfication.list + fi + echo "\$s done." + else + echo "Ugh, there was no hit for \$s .." + echo "We still love \$s though and we will add it to the final fasta file" + echo "\$s" >> otu.list + echo "NO_HIT" >> access.list + echo "NO_HIT" >> "\$name"_genes.list + echo "NO_HIT" >> "\$name"_virus.list + echo "NO_HIT" >> evalue.list + echo "NO_HIT" >> bit.list + echo "NO_HIT" >> pid.list + echo "NO_HIT" >> length.list + virus="NO" + gene="HIT" + echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + if [[ "${params.lca}" == "T" ]] + then echo "N/A" >> classfication.list + fi + echo "\$s done." + fi + echo "Done with \$s" + done + echo "Now editing "\$name" fasta headers" + ###### rename_seq.py + ./rename_seq.py ${asvs} new_"\$name"_asvnames.txt "\$name"_TaxonomyLabels.fasta + awk 'BEGIN {RS=">";FS="\\n";OFS=""} NR>1 {print ">"\$1; \$1=""; print}' "\$name"_TaxonomyLabels.fasta >"\$name"_tmpssasv.fasta + echo "[Sequence header]" > newnames.list + cat new_"\$name"_asvnames.txt >> newnames.list + touch sequence.list + echo " " > sequence.list + grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list + rm "\$name"_tmpssasv.fasta + if [[ "${params.lca}" == "T" ]] + then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list classfication.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv + else paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + fi + for x in *phyloformat.csv;do + echo "\$x" + lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) + tail -"\$lin" \$x | awk -F "," '{print \$2}' > tmpcol.list; + sed 's/ /_/g' tmpcol.list > tmp2col.list; + cat tmp2col.list | sort | uniq -c | sort -nr | awk '{print \$2","\$1}' > \${name}_summary_for_plot.csv; + rm tmpcol.list tmp2col.list + done + rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list + """ + } + } + } - label 'norm_cpus' + if (!params.skipPhylogeny) { - publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Nucleotide", mode: "copy", overwrite: true, pattern: '*{.csv}' + process pcASV_Protein_Phylogeny { - input: - file(fasta) from reads_vsearch5_ch + label 'norm_cpus' - output: - file("number_per_percentage_nucl.csv") into number_per_percent_nucl_plot + tag "${mtag}" - script: - if (params.datacheckntIDlist) { - """ - for id in `echo ${params.datacheckntIDlist} | tr "," "\\n"`;do - vsearch --cluster_fast ${fasta} --centroids ${params.projtag}_ncASV\${id}.fasta --threads ${task.cpus} --relabel OTU --id \${id} - done - for x in *ncASV*.fasta;do - id=\$( echo \$x | awk -F "_ncASV" '{print \$2}' | awk -F ".fasta" '{print \$1}') - numb=\$( grep -c ">" \$x ) - echo "\${id},\${numb}" >> number_per_percentage_nucl.csv - done - yo=\$(grep -c ">" ${fasta}) - echo "1.0,\${yo}" >> number_per_percentage_nucl.csv - """ - } - } + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Phylogeny/Alignment", mode: "copy", overwrite: true, pattern: '*aln.*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Phylogeny/Modeltest", mode: "copy", overwrite: true, pattern: '*mt*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*iq*' + input: + tuple nid, file(prot) from pcASV_aaMafft_ch - if (params.sing) { + output: + tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into pcASV_protein_phylogeny_results + tuple nid, file("*iq.treefile") into potu_Atree_plot - process Translating_For_ProteinClustering_DC { + script: + mtag="ID=" + nid + """ + pre=\$( echo ${prot} | awk -F ".fasta" '{print \$1}' ) + mafft --maxiterate 5000 --auto ${prot} >\${pre}_ALN.fasta + trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html + o-trim-uninformative-columns-from-alignment \${pre}_aln.fasta + mv \${pre}_aln.fasta-TRIMMED ./\${pre}_Aligned_informativeonly.fasta + # pcASV_Protein_ModelTest + modeltest-ng -i \${pre}_Aligned_informativeonly.fasta -p ${task.cpus} -o \${pre}_mt -d aa -s 203 --disable-checkpoint - label 'low_cpus' + # pcASV_Protein_Phylogeny + if [ "${params.iqCustomaa}" != "" ];then + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq --redo -T auto ${params.iqCustomaa} - publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Aminoacid/translation", mode: "copy", overwrite: true + elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -b ${params.boots} - input: - file(fasta) from nucl2aa + elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then + mod=\$(tail -12 \${pre}_Aligned_informativeonly.fasta.log | head -1 | awk '{print \$6}') + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni - output: - file("*ASVprotforclust.fasta") into clustering_aa - file("*_translation_report") into reportaa_VR - file("*_ASV_all.fasta") into asvfastaforaaclust + elif [ "${params.nonparametric}" != "false" ];then + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} - script: - """ - conda init && source activate virtualribosome + elif [ "${params.parametric}" != "false" ];then + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_ASVprotforclust.fasta --report ${params.projtag}_translation_report - cp ${fasta} ${params.projtag}_ASV_all.fasta + else + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + fi + """ + } + } - """ + process Generate_pcASV_Protein_Counts { - } + label 'high_cpus' - } else { + tag "${mtag}" - process Translation_For_ProteinBased_Clustering_DC { + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Aminoacid/Counts", mode: "copy", overwrite: true - label 'norm_cpus' + input: + tuple nid, file(fasta) from pcASV_aaCounts_ch + file(merged) from mergeforpcASVaacounts + file(samplist) from samplistpotu - conda 'python=2.7' + output: + tuple file("*_counts.csv"), file("*dmd.out") into potuaacounts_summary + tuple nid, file("*counts.csv") into potu_Acounts - publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Aminoacid/translation", mode: "copy", overwrite: true + script: + // check do I need for loop + mtag="ID=" + nid + """ + set +e + potu="\$( echo ${fasta} | awk -F "_" '{print \$3}')" + diamond makedb --in ${fasta} --db ${fasta} + diamond blastx -q ${merged} -d ${fasta} -p ${task.cpus} --min-score ${params.ProtCountsBit} --id ${params.ProtCountID} -l ${params.ProtCountsLength} --more-sensitive -o ${params.projtag}_\${potu}_Counts_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 --max-hsps 1 + echo "OTU_ID" >tmp.col1.txt + echo "Generating sample id list" + grep ">" ${fasta} | awk -F ">" '{print \$2}' | sort | uniq > otuid.list + cat otuid.list >> tmp.col1.txt + echo "Beginning them counts tho my g" + for y in \$( cat ${samplist} );do + echo "Starting with \$y now ..." + grep "\$y" ${params.projtag}_\${potu}_Counts_dmd.out > tmp."\$y".out + echo "Isolated hits" + echo "Created uniq subject id list" + echo "\$y" > "\$y"_col.txt + echo "Starting my counts" + for z in \$(cat otuid.list);do + echo "Counting \$z hits" + echo "grep -wc "\$z" >> "\$y"_col.txt" + grep -wc "\$z" tmp."\$y".out >> "\$y"_col.txt + echo "\$z counted" + done + done + paste -d "," tmp.col1.txt *col.txt > ${params.projtag}_aminoacid_\${potu}_noTaxonomy_counts.csv + rm tmp* + rm *col.txt + """ + } + } - input: - file(fasta) from nucl2aa + if (!params.skipReport) { - output: - file("*ASVprotforclust.fasta") into clustering_aa - file("*_translation_report") into reportaa_VR - file("*_ASV_all.fasta") into asvfastaforaaclust + if (!params.skipAdapterRemoval || !params.skipReadProcessing || !params.skipMerging) { - script: - """ - ${tools}/virtualribosomev2/dna2pep.py ${fasta} -r all -x -o none --fasta ${params.projtag}_ASVprotforclust.fasta --report ${params.projtag}_translation_report - cp ${fasta} ${params.projtag}_ASV_all.fasta - """ - } - } + process combine_csv { - process Protein_clustering_DC { + input: + file(csv) from fastp_csv_in2 + .collect() - label 'norm_cpus' + output: + file("final_reads_stats.csv") into fastp_csv_in - publishDir "${params.workingdir}/${params.outdir}/DataCheck/Clustering/Aminoacid", mode: "copy", overwrite: true, pattern: '*{.csv}' + script: + """ + cat ${csv} >all_reads_stats.csv + head -n1 all_reads_stats.csv >tmp.names.csv + cat all_reads_stats.csv | grep -v ""Sample,Total_"" >tmp.reads.stats.csv + cat tmp.names.csv tmp.reads.stats.csv >final_reads_stats.csv + rm tmp.names.csv tmp.reads.stats.csv + """ + } + } else { - input: - file(fasta) from clustering_aa - file(asvs) from asvfastaforaaclust + process skip_combine_csv { - output: - file("number_per_percentage_prot.csv") into number_per_percent_prot_plot + output: + file("filter_reads.txt") into fastp_csv_in - script: - // add awk script to count seqs - """ - set +e - cp ${params.vampdir}/bin/rename_seq.py . - for id in `echo ${params.datacheckaaIDlist} | tr "," "\\n"`;do - if [ \${id} == ".55" ];then - word=3 - elif [ \${id} == ".65" ];then - word=4 - else - word=5 - fi - awk 'BEGIN{RS=">";ORS=""}length(\$2)>="${params.minAA}"{print ">"\$0}' ${fasta} > ${params.projtag}_filtered_proteins.fasta - cd-hit -i ${params.projtag}_filtered_proteins.fasta -n \${word} -c \${id} -o ${params.projtag}_pcASV\${id}.fasta - sed 's/>Cluster />Cluster_/g' ${params.projtag}_pcASV\${id}.fasta.clstr >${params.projtag}_pcASV\${id}.clstr - grep ">Cluster_" ${params.projtag}_pcASV\${id}.clstr >temporaryclusters.list - y=\$(grep -c ">Cluster_" ${params.projtag}_pcASV\${id}.clstr) - echo ">Cluster_"\${y}"" >> ${params.projtag}_pcASV\${id}.clstr - t=1 - b=1 - for x in \$(cat temporaryclusters.list);do - echo "Extracting \$x" - name="\$( echo \$x | awk -F ">" '{print \$2}')" - clust="pcASV"\${t}"" - echo "\${name}" - awk '/^>'\${name}'\$/,/^>Cluster_'\${b}'\$/' ${params.projtag}_pcASV\${id}.clstr > "\${name}"_"\${clust}"_tmp.list - t=\$(( \${t}+1 )) - b=\$(( \${b}+1 )) - done - ls *_tmp.list - u=1 - for x in *_tmp.list;do - name="\$(echo \$x | awk -F "_p" '{print \$1}')" - echo "\${name}" - cluster="\$(echo \$x | awk -F "_" '{print \$3}')" - echo "\${cluster}" - grep "ASV" \$x | awk -F ", " '{print \$2}' | awk -F "_" '{print \$1}' | awk -F ">" '{print \$2}' > \${name}_\${cluster}_seqs_tmps.list - seqtk subseq ${asvs} \${name}_\${cluster}_seqs_tmps.list > \${name}_\${cluster}_nucleotide_sequences.fasta - vsearch --cluster_fast \${name}_\${cluster}_nucleotide_sequences.fasta --id 0.2 --centroids \${name}_\${cluster}_centroids.fasta - grep ">" \${name}_\${cluster}_centroids.fasta >> \${name}_\${cluster}_tmp_centroids.list - for y in \$( cat \${name}_\${cluster}_tmp_centroids.list );do - echo ">\${cluster}_type"\$u"" >> \${name}_\${cluster}_tmp_centroid.newheaders - u=\$(( \${u}+1 )) - done - u=1 - ./rename_seq.py \${name}_\${cluster}_centroids.fasta \${name}_\${cluster}_tmp_centroid.newheaders \${cluster}_types_labeled.fasta - done - cat *_types_labeled.fasta >> ${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta - grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$3}' | awk -F "." '{print \$1}' >tmphead.list - grep -w "*" ${params.projtag}_pcASV\${id}.clstr | awk '{print \$2}' | awk -F "," '{print \$1}' >tmplen.list - paste -d"," temporaryclusters.list tmphead.list >tmp.info.csv - grep ">" ${params.projtag}_pcASV\${id}.fasta >lala.list - j=1 - for x in \$(cat lala.list);do - echo ">${params.projtag}_pcASV\${j}" >>${params.projtag}_aminoheaders.list - echo "\${x},>${params.projtag}_pcASV\${j}" >>tmpaminotype.info.csv - j=\$(( \${j}+1 )) - done - rm lala.list - awk -F "," '{print \$2}' tmp.info.csv >>tmporder.list - for x in \$(cat tmporder.list);do - grep -w "\$x" tmpaminotype.info.csv | awk -F "," '{print \$2}' >>tmpder.list - done - paste -d "," temporaryclusters.list tmplen.list tmphead.list tmpder.list >${params.projtag}_pcASVCluster\${id}_summary.csv - ./rename_seq.py ${params.projtag}_pcASV\${id}.fasta ${params.projtag}_aminoheaders.list ${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta - stats.sh in=${params.projtag}_aminoacid_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_aminoacid_clustered.gc gcformat=4 overwrite=true - stats.sh in=${params.projtag}_nucleotide_pcASV\${id}_noTaxonomy.fasta gc=${params.projtag}_pcASV\${id}_nucleotide_clustered.gc gcformat=4 overwrite=true - awk 'BEGIN{RS=">";ORS=""}length(\$2)<"${params.minAA}"{print ">"\$0}' ${fasta} >${params.projtag}_pcASV\${id}_problematic_translations.fasta - if [ `wc -l ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk '{print \$1}'` -gt 1 ];then - grep ">" ${params.projtag}_pcASV\${id}_problematic_translations.fasta | awk -F ">" '{print \$2}' > problem_tmp.list - seqtk subseq ${asvs} problem_tmp.list > ${params.projtag}_pcASV\${id}_problematic_nucleotides.fasta - else - rm ${params.projtag}_pcASV\${id}_problematic_translations.fasta - fi - rm *.list - rm Cluster* - rm *types* - rm *tmp* - rm ${params.projtag}_pcASV\${id}.fast* - done - for x in *aminoacid*noTaxonomy.fasta;do - id=\$( echo \$x | awk -F "_noTax" '{print \$1}' | awk -F "pcASV" '{print \$2}') - numb=\$( grep -c ">" \$x) - echo "\${id},\${numb}" >> number_per_percentage_protz.csv - done - yesirr=\$( wc -l number_per_percentage_protz.csv | awk '{print \$1}') - tail -\$(( \${yesirr}-1 )) number_per_percentage_protz.csv > number_per_percentage_prot.csv - head -1 number_per_percentage_protz.csv >> number_per_percentage_prot.csv - rm number_per_percentage_protz.csv - """ - } + script: + """ + echo "Read processing steps skipped." >filter_reads.txt + """ + } + } - process combine_csv_DC { + //NEW REPORT !!!!!!!!!!!!!!!!! + /*Report_ASV + asv_counts_plots -> ${params.projtag}_ASV_counts.csv + taxplot1 -> ${params.projtag}_ASV_summary_for_plot.csv + asv_heatmap -> ${params.projtag}_ASV_PercentID.matrix + nucl_phyl_plot -> ${params.projtag}_ASV_iq.treefile + file("*_ASV_Grouping.csv") into asvgroupscsv + "${params.projtag}_ASV_Groupingcounts.csv") into asvgroupcounts + *_quick_Taxbreakdown.csv") into tax_table_asv + \\${params.projtag}_ASV_Group_Reps_iq.treefile + */ + + report_asv = Channel.create() + asv_counts_plots.mix(taxplot_asv, asv_heatmap, nucl_phyl_plot_asv, asvgroupscsv, asvgroupcounts, asv_group_rep_tree, tax_table_asv).flatten().buffer(size:8).dump(tag:'asv').into(report_asv) + + if (params.ncASV) { + report_ncasv = Channel.create() + notu_counts_plots.mix(taxplot_ncasv, notu_heatmap, nucl_phyl_plot_ncasv, tax_table_ncasv).groupTuple(by:0, size:5).dump(tag:'ncasv').into(report_ncasv) + /* + notu_counts_plots -> ${params.projtag}_ncASV${id}_counts.csv + taxplot1a -> ${params.projtag}_ncASV${id}_summary_for_plot.csv + notu_heatmap -> ${params.projtag}_ncASV${id}_PercentID.matrix + nucl_phyl_plot -> ${params.projtag}_ncASV${id}_iq.treefile + ${params.projtag}_ncASV${id}_quick_Taxbreakdown.csv + */ + } else { + report_ncasv = Channel.empty() + } - input: - file(csv) from fastp_csv - .collect() + if (params.pcASV) { + report_pcasv_aa = Channel.create() + potu_Acounts.mix(taxplot4, potu_aa_heatmap, potu_Atree_plot, tax_table_pcasvaa).groupTuple(by:0, size:5).dump(tag:'pcasv1').into(report_pcasv_aa) + /*Report_pcASV_AminoAcid + potu_Acounts -> ${params.projtag}_pcASV${id}_noTaxonomy_counts.csv + taxplot4 -> ${params.projtag}_aminoacid_pcASV${id}_noTaxonomy_summary_for_plot.csv + potu_aa_heatmap -> ${params.projtag}_aminoacid_pcASV${id}_noTaxonomy_PercentID.matrix + potu_Atree_plot -> ${params.projtag}_aminoacid_pcASV${id}_noTaxonomy_iq.treefile + tax_table_pcasvaa -> ${params.projtag}_aminoacid_pcASV${id}_quick_Taxbreakdown.csv + */ + report_pcasv_nucl = Channel.create() + potu_Ncounts_for_report.mix(taxplot3, potu_nucl_heatmap, potu_Ntree_plot, tax_table_pcasvnt).groupTuple(by:0, size:5).dump(tag:'pcasv2').into(report_pcasv_nucl) + /*Report_pcASV_Nucleotide + potu_Ncounts_for_report -> ${params.projtag}_nucleotide_pcASV${id}_noTaxonomy_counts.csv + taxplot3 -> ${params.projtag}_nucleotide_pcASV${id}_noTaxonomy_summary_for_plot.csv + potu_nucl_heatmap -> ${params.projtag}_nucleotide_pcASV${id}_noTaxonomy_PercentID.matrix + potu_Ntree_plot -> ${params.projtag}_nucleotide_pcASV${id}_noTaxonomy_iq.treefile + tax_table_pcasvnt -> ${params.projtag}_nucleotide_pcASV${id}_quick_Taxbreakdown.csv + */ + } else { + report_pcasv_aa = Channel.empty() + report_pcasv_nucl = Channel.empty() + } - output: - file("final_reads_stats.csv") into fastp_csv1 + if (!params.skipAminoTyping) { + report_aminotypes = Channel.create() + aminocounts_plot.mix(taxplot2, aminotype_heatmap, amino_rax_plot, atygroupscsv, amino_group_rep_tree, amino_groupcounts, tax_table_amino).flatten().buffer(size:8).dump(tag:'amino').into(report_aminotypes) + /* + Report_AminoTypes + aminocounts_plot -> ${params.projtag}_AminoType_counts.csv + taxplot2 -> ${params.projtag}_AminoTypes_summary_for_plot.csv + aminotype_heatmap -> ${params.projtag}_AminoTypes_PercentID.matrix + amino_rax_plot -> ${params.projtag}_AminoTypes_iq.treefile + atygroupscsv -> *_AminoType_Grouping.csv + amino_group_rep_tree -> ${params.projtag}_AminoType_Group_Reps_iq.treefile + params.projtag}_AminoType_Groupingcounts.csv") into amino_groupcounts + *_quick_Taxbreakdown.csv") into tax_table_amino + */ + } else { + report_aminotypes = Channel.empty() + } - script: - """ - cat ${csv} >all_reads_stats.csv - head -n1 all_reads_stats.csv >tmp.names.csv - cat all_reads_stats.csv | grep -v ""Sample,Total_"" >tmp.reads.stats.csv - cat tmp.names.csv tmp.reads.stats.csv >final_reads_stats.csv - rm tmp.names.csv tmp.reads.stats.csv - """ + report_all_ch = Channel.create() + report_asv.mix(report_ncasv, report_pcasv_aa, report_pcasv_nucl, report_aminotypes).map{it.flatten()}.dump(tag:'report').into(report_all_ch) - } + process Report { - process Report_DataCheck { + label 'norm_cpus' - label 'norm_cpus' + publishDir "${params.workingdir}/${params.outdir}/Analyze/FinalReports", mode: "copy", overwrite: true - publishDir "${params.workingdir}/${params.outdir}/DataCheck/Report", mode: "copy", overwrite: true, pattern: '*.{html}' + input: + file(csv) from fastp_csv_in + file(files) from report_all_ch - input: - file(fastpcsv) from fastp_csv1 - file(reads_per_sample_preFilt) from reads_per_sample_preFilt - file(read_per_sample_postFilt) from reads_per_sample_postFilt - file(preFilt_baseFrequency) from prefilt_basefreq - file(postFilt_baseFrequency) from postFilt_basefreq - file(preFilt_qualityScore) from prefilt_qualityscore - file(postFilt_qualityScore) from postFilt_qualityscore - file(preFilt_gcContent) from prefilt_gccontent - file(postFilt_gcContent) from postFilt_gccontent - file(preFilt_averageQuality) from prefilt_averagequality - file(postFilt_averageQuaulity) from postFilt_averagequality - file(preFilt_length) from prefilt_length - file(postFilt_length) from postFilt_length - file(number_per_percentage_nucl) from number_per_percent_nucl_plot - file(number_per_percentage_prot) from number_per_percent_prot_plot + output: + file("*.html") into report_all_out - output: - file("*.html") into datacheckreport + script: + """ + name=\$( ls *summary_for_plot.csv | awk -F "_summary_for_plot.csv" '{print \$1}') + cp ${params.vampdir}/bin/vAMPirus_Report.Rmd . + cp ${params.vampdir}/example_data/conf/vamplogo.png . + Rscript -e "rmarkdown::render('vAMPirus_Report.Rmd',output_file='\${name}_Report.html')" \${name} \ + ${params.skipReadProcessing} \ + ${params.skipMerging} \ + ${params.skipAdapterRemoval} \ + ${params.skipTaxonomy} \ + ${params.skipPhylogeny} \ + ${params.trymax} \ + ${params.stats} \ + ${params.metadata} \ + ${params.minimumCounts} + """ + } - script: - """ - cp ${params.vampdir}/bin/vAMPirus_DC_Report.Rmd . - cp ${params.vampdir}/example_data/conf/vamplogo.png . - Rscript -e "rmarkdown::render('vAMPirus_DC_Report.Rmd',output_file='${params.projtag}_DataCheck_Report.html')" ${params.projtag} \ - ${fastpcsv} \ - ${reads_per_sample_preFilt} \ - ${read_per_sample_postFilt} \ - ${preFilt_baseFrequency} \ - ${postFilt_baseFrequency} \ - ${preFilt_qualityScore} \ - ${postFilt_qualityScore} \ - ${preFilt_averageQuality} \ - ${postFilt_averageQuaulity} \ - ${preFilt_length} \ - ${postFilt_length} \ - ${number_per_percentage_nucl} \ - ${number_per_percentage_prot} - """ - } + } + } } else { println("\n\t\033[0;31mMandatory argument not specified. For more info use `nextflow run vAMPirus.nf --help`\n\033[0m") exit 0 } - -workflow.onComplete { - log.info ( workflow.success ? \ - "---------------------------------------------------------------------------------" \ - + "\n\033[0;32mDone! Open the following reports in your browser\033[0m" \ - + "\n\033[0;32mPipeline performance report: ${params.workingdir}/${params.outdir}/${params.tracedir}/vampirus_report.html\033[0m" \ - + "\n\033[0;32mvAMPirus --DataCheck interactive report: ${params.workingdir}/${params.outdir}/DataCheck/*.hmtl\033[0m" \ - + "\n\033[0;32mvAMPirus --Analyze interactive report: ${params.workingdir}/${params.outdir}/Analyze/*.hmtl\033[0m" \ - : \ - "---------------------------------------------------------------------------------" \ - + "\n\033[0;31mSomething went wrong. Check error message below and/or log files.\033[0m" ) +if (params.DataCheck) { + workflow.onComplete { + log.info ( workflow.success ? \ + "---------------------------------------------------------------------------------" \ + + "\n\033[0;32mDone! Open the following reports in your browser\033[0m" \ + + "\n\033[0;32mPipeline performance report: ${params.workingdir}/${params.outdir}/${params.tracedir}/vampirus_report.html\033[0m" \ + + "\n\033[0;32mvAMPirus --DataCheck interactive report: ${params.workingdir}/${params.outdir}/DataCheck/Report/*.hmtl\033[0m" \ + : \ + "---------------------------------------------------------------------------------" \ + + "\n\033[0;31mSomething went wrong. Check error message below and/or log files.\033[0m" ) + } +} else if (params.Analyze) { + workflow.onComplete { + log.info ( workflow.success ? \ + "---------------------------------------------------------------------------------" \ + + "\n\033[0;32mDone! Open the following reports in your browser\033[0m" \ + + "\n\033[0;32mPipeline performance report: ${params.workingdir}/${params.outdir}/${params.tracedir}/vampirus_report.html\033[0m" \ + + "\n\033[0;32mvAMPirus --Analyze interactive report: ${params.workingdir}/${params.outdir}/Analyze/*.hmtl\033[0m" \ + : \ + "---------------------------------------------------------------------------------" \ + + "\n\033[0;31mSomething went wrong. Check error message below and/or log files.\033[0m" ) + } } From f5c083dc8815589ae28e300f8f56aa6b34dccfd6 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Mon, 3 May 2021 17:40:19 -0500 Subject: [PATCH 110/223] Corrected aminoMED in help --- vAMPirus.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 3af44b5..d82a494 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -61,7 +61,7 @@ def helpMessage() { --asvMED Set this option to perform Minimum Entropy Decomposition on ASV sequences, see manual for more information. You will need to set a value for --asvC to perform this analysis - --amino_med Set this option to perform Minimum Entropy Decomposition on AminoType sequences, see manual for more information. You will need to set a value for --aminoC to perform this analysis + --aminoMED Set this option to perform Minimum Entropy Decomposition on AminoType sequences, see manual for more information. You will need to set a value for --aminoC to perform this analysis --Skip options-- From 06da69920764c548653bc1fb283ff06ab2cf5b4d Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Wed, 5 May 2021 16:05:04 -0500 Subject: [PATCH 111/223] Added skip phylogeny to MED phylogeny steps --- vAMPirus.nf | 225 +++++++++++++++++++++++++++------------------------- 1 file changed, 115 insertions(+), 110 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index d82a494..9aa37cb 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -2074,49 +2074,52 @@ if (params.DataCheck || params.Analyze) { """ } - process ASV_MED_Reps_phylogeny { + if (!params.skipPhylogeny) { - label 'low_cpus' + process ASV_MED_Reps_phylogeny { - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/MED/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*ASV*mt*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/MED/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*ASV*iq*' + label 'low_cpus' - input: - file(reps) from groupreps + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/MED/Phylogeny/ModelTest", mode: "copy", overwrite: true, pattern: '*ASV*mt*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/ASVs/MED/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*ASV*iq*' - output: - file("*_ASV_Group_Reps*") into align_results_asvmed - file("*iq.treefile") into asv_group_rep_tree + input: + file(reps) from groupreps - script: - """ - # Protein_ModelTest - modeltest-ng -i ${reps} -p ${task.cpus} -o ${params.projtag}_ASV_Group_Reps_mt -d aa -s 203 --disable-checkpoint + output: + file("*_ASV_Group_Reps*") into align_results_asvmed + file("*iq.treefile") into asv_group_rep_tree + + script: + """ + # Protein_ModelTest + modeltest-ng -i ${reps} -p ${task.cpus} -o ${params.projtag}_ASV_Group_Reps_mt -d aa -s 203 --disable-checkpoint - # Protein_Phylogeny - if [ "${params.iqCustomaa}" != "" ];then - iqtree -s ${reps} --prefix ${params.projtag}_ASV_Group_Reps_iq --redo -T auto ${params.iqCustomaa} + # Protein_Phylogeny + if [ "${params.iqCustomaa}" != "" ];then + iqtree -s ${reps} --prefix ${params.projtag}_ASV_Group_Reps_iq --redo -T auto ${params.iqCustomaa} - elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') - iqtree -s ${reps} --prefix ${params.projtag}_ASV_Group_Reps_iq -m \${mod} --redo -nt auto -b ${params.boots} + elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then + mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') + iqtree -s ${reps} --prefix ${params.projtag}_ASV_Group_Reps_iq -m \${mod} --redo -nt auto -b ${params.boots} - elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') - iqtree -s ${reps} --prefix ${params.projtag}_ASV_Group_Reps_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni + elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then + mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') + iqtree -s ${reps} --prefix ${params.projtag}_ASV_Group_Reps_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni - elif [ "${params.nonparametric}" != "false" ];then - iqtree -s ${reps} --prefix ${params.projtag}_ASV_Group_Reps_iq -m MFP --redo -nt auto -b ${params.boots} + elif [ "${params.nonparametric}" != "false" ];then + iqtree -s ${reps} --prefix ${params.projtag}_ASV_Group_Reps_iq -m MFP --redo -nt auto -b ${params.boots} - elif [ "${params.parametric}" != "false" ];then - iqtree -s ${reps} --prefix ${params.projtag}_ASV_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + elif [ "${params.parametric}" != "false" ];then + iqtree -s ${reps} --prefix ${params.projtag}_ASV_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - else - iqtree -s ${reps} --prefix ${params.projtag}_ASV_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - fi - """ + else + iqtree -s ${reps} --prefix ${params.projtag}_ASV_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + fi + """ + } } - + process Adding_ASV_MED_Info { label 'low_cpus' @@ -2616,107 +2619,109 @@ if (params.DataCheck || params.Analyze) { } } - if (params.aminoMED) { + if (params.aminoMED) { - process AminoType_Minimum_Entropy_Decomposition { + process AminoType_Minimum_Entropy_Decomposition { - label 'low_cpus' + label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes/MED", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/Analyze/Clustering/AminoTypes/MED", mode: "copy", overwrite: true - input: - file(aminos) from aminos_for_med + input: + file(aminos) from aminos_for_med - output: - file("*_AminoType_Grouping.csv") into atygroupscsv - file("${params.projtag}_AminoType_group_reps_aligned.fasta") into atygroupreps + output: + file("*_AminoType_Grouping.csv") into atygroupscsv + file("${params.projtag}_AminoType_group_reps_aligned.fasta") into atygroupreps - script: - """ - #alignment - mafft --thread ${task.cpus} --maxiterate 15000 --auto ${aminos} > ${params.projtag}_AminoTypes_mafftAlign.fasta - #trimming - trimal -in ${params.projtag}_AminoTypes_mafftAlign.fasta -out ${params.projtag}_AminoTypes_mafftAligned.fasta -keepheader -fasta -automated1 - rm ${params.projtag}_AminoTypes_mafftAlign.fasta - o-trim-uninformative-columns-from-alignment ${params.projtag}_AminoTypes_mafftAligned.fasta - mv ${params.projtag}_AminoTypes_mafftAligned.fasta-TRIMMED ./${params.projtag}_AminoTypes_Aligned_informativeonly.fasta - #entopy analysis - entropy-analysis ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta - #Decomposition - oligotype ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta-ENTROPY -o ${params.projtag}_AminoTypeMED_${params.aminoC} -M 1 -c ${params.aminoC} -N ${task.cpus} --skip-check-input --no-figures --skip-gen-html - #generatemaps - cd ./${params.projtag}_AminoTypeMED_${params.aminoC}/OLIGO-REPRESENTATIVES/ - echo "AminoType,Group,IDPattern" - j=1 - for x in *_unique; - do gid=\$(echo \$x | awk -F "_" '{print \$1}') - uni=\$(echo \$x | awk -F ""\${gid}"_" '{print \$2}' | awk -F "_uni" '{print \$1}') - grep ">" "\$gid"_"\$uni" | awk -F ">" '{print \$2}' > asv.list - seqtk subseq ../../${aminos} asv.list > Group"\${j}"_sequences.fasta - for z in \$( cat asv.list) - do echo ""\$z",Group"\$j","\$uni"" >> ${params.projtag}_AminoType_Grouping.csv + script: + """ + #alignment + mafft --thread ${task.cpus} --maxiterate 15000 --auto ${aminos} > ${params.projtag}_AminoTypes_mafftAlign.fasta + #trimming + trimal -in ${params.projtag}_AminoTypes_mafftAlign.fasta -out ${params.projtag}_AminoTypes_mafftAligned.fasta -keepheader -fasta -automated1 + rm ${params.projtag}_AminoTypes_mafftAlign.fasta + o-trim-uninformative-columns-from-alignment ${params.projtag}_AminoTypes_mafftAligned.fasta + mv ${params.projtag}_AminoTypes_mafftAligned.fasta-TRIMMED ./${params.projtag}_AminoTypes_Aligned_informativeonly.fasta + #entopy analysis + entropy-analysis ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta + #Decomposition + oligotype ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta-ENTROPY -o ${params.projtag}_AminoTypeMED_${params.aminoC} -M 1 -c ${params.aminoC} -N ${task.cpus} --skip-check-input --no-figures --skip-gen-html + #generatemaps + cd ./${params.projtag}_AminoTypeMED_${params.aminoC}/OLIGO-REPRESENTATIVES/ + echo "AminoType,Group,IDPattern" + j=1 + for x in *_unique; + do gid=\$(echo \$x | awk -F "_" '{print \$1}') + uni=\$(echo \$x | awk -F ""\${gid}"_" '{print \$2}' | awk -F "_uni" '{print \$1}') + grep ">" "\$gid"_"\$uni" | awk -F ">" '{print \$2}' > asv.list + seqtk subseq ../../${aminos} asv.list > Group"\${j}"_sequences.fasta + for z in \$( cat asv.list) + do echo ""\$z",Group"\$j","\$uni"" >> ${params.projtag}_AminoType_Grouping.csv - done - rm asv.list - echo ">Group\${j}" >> ${params.projtag}_AminoType_group_reps_aligned.fasta - echo "\$uni" > group.list - seqtk subseq ../OLIGO-REPRESENTATIVES.fasta group.list > group.fasta - tail -1 group.fasta >> ${params.projtag}_AminoType_group_reps_aligned.fasta - mv "\$gid"_"\$uni" ./Group"\$j"_"\$uni"_aligned.fasta - mv "\$gid"_"\$uni"_unique ./Group"\$j"_"\$uni"_unqiues_aligned.fasta - rm "\$gid"*.cPickle - j=\$((\$j+1)) - done - mv ${params.projtag}_AminoType_Grouping.csv ../../ - mv ${params.projtag}_AminoType_group_reps_aligned.fasta ../../ - cd .. + done + rm asv.list + echo ">Group\${j}" >> ${params.projtag}_AminoType_group_reps_aligned.fasta + echo "\$uni" > group.list + seqtk subseq ../OLIGO-REPRESENTATIVES.fasta group.list > group.fasta + tail -1 group.fasta >> ${params.projtag}_AminoType_group_reps_aligned.fasta + mv "\$gid"_"\$uni" ./Group"\$j"_"\$uni"_aligned.fasta + mv "\$gid"_"\$uni"_unique ./Group"\$j"_"\$uni"_unqiues_aligned.fasta + rm "\$gid"*.cPickle + j=\$((\$j+1)) + done + mv ${params.projtag}_AminoType_Grouping.csv ../../ + mv ${params.projtag}_AminoType_group_reps_aligned.fasta ../../ + cd .. - """ + """ + } } + if (!params.skipPhylogeny) { - process AminoType_MED_Reps_phylogeny { + process AminoType_MED_Reps_phylogeny { - label 'low_cpus' + label 'low_cpus' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/MED/Phylogeny/Modeltest", mode: "copy", overwrite: true, pattern: '*mt*' - publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/MED/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*iq*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/MED/Phylogeny/Modeltest", mode: "copy", overwrite: true, pattern: '*mt*' + publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/AminoTypes/MED/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*iq*' - input: - file(reps) from atygroupreps + input: + file(reps) from atygroupreps - output: - file("*_AminoType_Group_Reps*") into align_results_aminmed - file("*iq.treefile") into amino_group_rep_tree + output: + file("*_AminoType_Group_Reps*") into align_results_aminmed + file("*iq.treefile") into amino_group_rep_tree - script: - """ - # Protein_ModelTest - modeltest-ng -i ${reps} -p ${task.cpus} -o ${params.projtag}_AminoType_Group_Reps_mt -d aa -s 203 --disable-checkpoint + script: + """ + # Protein_ModelTest + modeltest-ng -i ${reps} -p ${task.cpus} -o ${params.projtag}_AminoType_Group_Reps_mt -d aa -s 203 --disable-checkpoint - # Protein_Phylogeny - if [ "${params.iqCustomaa}" != "" ];then - iqtree -s ${reps} --prefix ${params.projtag}_AminoType_Group_Reps_iq --redo -T auto ${params.iqCustomaa} + # Protein_Phylogeny + if [ "${params.iqCustomaa}" != "" ];then + iqtree -s ${reps} --prefix ${params.projtag}_AminoType_Group_Reps_iq --redo -T auto ${params.iqCustomaa} - elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then - mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') - iqtree -s ${reps} --prefix ${params.projtag}_AminoType_Group_Reps_iq -m \${mod} --redo -nt auto -b ${params.boots} + elif [[ "${params.ModelTaa}" != "false" && "${params.nonparametric}" != "false" ]];then + mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') + iqtree -s ${reps} --prefix ${params.projtag}_AminoType_Group_Reps_iq -m \${mod} --redo -nt auto -b ${params.boots} - elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then - mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') - iqtree -s ${reps} --prefix ${params.projtag}_AminoType_Group_Reps_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni + elif [[ "${params.ModelTaa}" != "false" && "${params.parametric}" != "false" ]];then + mod=\$(tail -12 ${reps}.log | head -1 | awk '{print \$6}') + iqtree -s ${reps} --prefix ${params.projtag}_AminoType_Group_Reps_iq -m \${mod} --redo -nt auto -bb ${params.boots} -bnni - elif [ "${params.nonparametric}" != "false" ];then - iqtree -s ${reps} --prefix ${params.projtag}_AminoType_Group_Reps_iq -m MFP --redo -nt auto -b ${params.boots} + elif [ "${params.nonparametric}" != "false" ];then + iqtree -s ${reps} --prefix ${params.projtag}_AminoType_Group_Reps_iq -m MFP --redo -nt auto -b ${params.boots} - elif [ "${params.parametric}" != "false" ];then - iqtree -s ${reps} --prefix ${params.projtag}_AminoType_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + elif [ "${params.parametric}" != "false" ];then + iqtree -s ${reps} --prefix ${params.projtag}_AminoType_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - else - iqtree -s ${reps} --prefix ${params.projtag}_AminoType_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni - fi - """ + else + iqtree -s ${reps} --prefix ${params.projtag}_AminoType_Group_Reps_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + fi + """ + } } - process Adding_AminoType_MED_Info { label 'low_cpus' From 221411faf6531d6285309b073c9197da5ed36897 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Sun, 27 Jun 2021 11:46:52 -0500 Subject: [PATCH 112/223] updated diamond version in yaml and removed line in nf --- vAMPirus.nf | 1 - vampirus_env.yml | 2 +- 2 files changed, 1 insertion(+), 2 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 9aa37cb..6c6b967 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -2119,7 +2119,6 @@ if (params.DataCheck || params.Analyze) { """ } } - process Adding_ASV_MED_Info { label 'low_cpus' diff --git a/vampirus_env.yml b/vampirus_env.yml index fa80f48..92ee9d5 100644 --- a/vampirus_env.yml +++ b/vampirus_env.yml @@ -10,7 +10,7 @@ channels: dependencies: - python=3.6 - blast=2.11.0 - - diamond=0.9.30 + - diamond=2.0.9 - fastqc=0.11.9 - fastp=0.20.1 - clustalo=1.2.4 From 42b17e2aa63c71dc07b6404df0006e83d9bf9e3d Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Wed, 28 Jul 2021 15:49:14 -0500 Subject: [PATCH 113/223] replaced mafft with muscle v5 --- bin/muscle5.0.1278_linux64 | Bin 0 -> 3805336 bytes vAMPirus.nf | 68 ++++++++++++++++++++++--------------- vampirus_env.yml | 1 - vampirus_startup.sh | 2 +- 4 files changed, 42 insertions(+), 29 deletions(-) create mode 100644 bin/muscle5.0.1278_linux64 diff --git a/bin/muscle5.0.1278_linux64 b/bin/muscle5.0.1278_linux64 new file mode 100644 index 0000000000000000000000000000000000000000..53f3e8f38063b2b65ce5da492440d85ab47220d9 GIT binary patch literal 3805336 zcmeFa33L=i_x?Wt2Ev+vEMbc=1Pq86KoKNhf*=##fMFF8V~B`|hyjV928bpsLkNad zMa77Sh=|B0Vt~jN+z=5JMFNJch6I9ufDp*2$x 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zBfq0R{hz)vf0sYLXDQH7cmJHKZy OIgJk*E@=F?@qYoSx%p%O literal 0 HcmV?d00001 diff --git a/vAMPirus.nf b/vAMPirus.nf index 6c6b967..8b6a27b 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1,4 +1,4 @@ -#!/usr/bin/env nextflow +muscle#!/usr/bin/env nextflow /* ======================================================================================== @@ -1106,11 +1106,12 @@ if (params.DataCheck || params.Analyze) { """ set +e #alignment - mafft --thread ${task.cpus} --maxiterate 15000 --auto ${asvs} > ${params.projtag}_ASVs_mafftAlign.fasta + ${tools}/muscle5.0.1278_linux64 -in ${asvs} -out ${params.projtag}_ASVs_muscleAlign.fasta -thread ${task.cpus} -quiet + #mafft --thread ${task.cpus} --maxiterate 15000 --auto ${asvs} > ${params.projtag}_ASVs_muscleAlign.fasta #trimming - trimal -in ${params.projtag}_ASVs_mafftAlign.fasta -out ${params.projtag}_ASVs_mafftAligned.fasta -keepheader -fasta -automated1 - rm ${params.projtag}_ASVs_mafftAlign.fasta - o-trim-uninformative-columns-from-alignment ${params.projtag}_ASVs_mafftAligned.fasta + trimal -in ${params.projtag}_ASVs_mafftAlign.fasta -out ${params.projtag}_ASVs_muscleAligned.fasta -keepheader -fasta -automated1 + rm ${params.projtag}_ASVs_muscleAlign.fasta + o-trim-uninformative-columns-from-alignment ${params.projtag}_ASVs_muscleAligned.fasta mv ${params.projtag}_ASVs_mafftAligned.fasta-TRIMMED ./${params.projtag}_ASVs_Aligned_informativeonly.fasta #entopy analysis entropy-analysis ${params.projtag}_ASVs_Aligned_informativeonly.fasta @@ -1187,12 +1188,14 @@ if (params.DataCheck || params.Analyze) { script: """ #alignment - mafft --thread ${task.cpus} --maxiterate 15000 --auto ${aminos} > ${params.projtag}_AminoTypes_mafftAlign.fasta + if [[ $(grep -c ">" ${aminos}) -gt 499 ]]; then algo="super5"; else algo="mpc"; fi + ${tools}/muscle5.0.1278_linux64 -"\${algo}" ${aminos} -out ${params.projtag}_AminoTypes_muscleAlign.fasta -thread ${task.cpus} -quiet + #mafft --thread ${task.cpus} --maxiterate 15000 --auto ${aminos} > ${params.projtag}_AminoTypes_muscleAlign.fasta #trimming - trimal -in ${params.projtag}_AminoTypes_mafftAlign.fasta -out ${params.projtag}_AminoTypes_mafftAligned.fasta -keepheader -fasta -automated1 - rm ${params.projtag}_AminoTypes_mafftAlign.fasta - o-trim-uninformative-columns-from-alignment ${params.projtag}_AminoTypes_mafftAligned.fasta - mv ${params.projtag}_AminoTypes_mafftAligned.fasta-TRIMMED ./${params.projtag}_AminoTypes_Aligned_informativeonly.fasta + trimal -in ${params.projtag}_AminoTypes_muscleAlign.fasta -out ${params.projtag}_AminoTypes_muscleAligned.fasta -keepheader -fasta -automated1 + rm ${params.projtag}_AminoTypes_muscleAlign.fasta + o-trim-uninformative-columns-from-alignment ${params.projtag}_AminoTypes_muscleAligned.fasta + mv ${params.projtag}_AminoTypes_muscleAligned.fasta-TRIMMED ./${params.projtag}_AminoTypes_Aligned_informativeonly.fasta #entropy analysis entropy-analysis ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta --amino-acid-sequences #summarize entropy peaks @@ -1644,7 +1647,8 @@ if (params.DataCheck || params.Analyze) { mtag="ID=" + nid """ pre=\$(echo ${asvs} | awk -F ".fasta" '{print \$1}' ) - mafft --thread ${task.cpus} --maxiterate 15000 --auto ${asvs} >\${pre}_ALN.fasta + ${tools}/muscle5.0.1278_linux64 -in ${asvs} -out \${pre}_ALN.fasta -thread ${task.cpus} -quiet + #mafft --thread ${task.cpus} --maxiterate 15000 --auto ${asvs} >\${pre}_ALN.fasta trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html o-trim-uninformative-columns-from-alignment \${pre}_aln.fasta mv \${pre}_aln.fasta-TRIMMED ./\${pre}_Aligned_informativeonly.fasta @@ -1990,7 +1994,8 @@ if (params.DataCheck || params.Analyze) { script: """ pre=\$(echo ${asvs} | awk -F ".fasta" '{print \$1}' ) - mafft --thread ${task.cpus} --maxiterate 15000 --auto ${asvs} >\${pre}_ALN.fasta + ${tools}/muscle5.0.1278_linux64 -in ${asvs} -out \${pre}_ALN.fasta -thread ${task.cpus} -quiet + #mafft --thread ${task.cpus} --maxiterate 15000 --auto ${asvs} >\${pre}_ALN.fasta trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html o-trim-uninformative-columns-from-alignment \${pre}_aln.fasta mv \${pre}_aln.fasta-TRIMMED ./\${pre}_Aligned_informativeonly.fasta @@ -2035,12 +2040,13 @@ if (params.DataCheck || params.Analyze) { script: """ #alignment - mafft --thread ${task.cpus} --maxiterate 15000 --auto ${asvs} > ${params.projtag}_ASVs_mafftAlign.fasta + ${tools}/muscle5.0.1278_linux64 -in ${asvs} -out -thread ${task.cpus} -quiet + #mafft --thread ${task.cpus} --maxiterate 15000 --auto ${asvs} > ${params.projtag}_ASVs_muscleAlign.fasta #trimming - trimal -in ${params.projtag}_ASVs_mafftAlign.fasta -out ${params.projtag}_ASVs_mafftAligned.fasta -keepheader -fasta -automated1 - rm ${params.projtag}_ASVs_mafftAlign.fasta - o-trim-uninformative-columns-from-alignment ${params.projtag}_ASVs_mafftAligned.fasta - mv ${params.projtag}_ASVs_mafftAligned.fasta-TRIMMED ./${params.projtag}_ASVs_Aligned_informativeonly.fasta + trimal -in ${params.projtag}_ASVs_muscleAlign.fasta -out ${params.projtag}_ASVs_muscleAligned.fasta -keepheader -fasta -automated1 + rm ${params.projtag}_ASVs_muscleAlign.fasta + o-trim-uninformative-columns-from-alignment ${params.projtag}_ASVs_muscleAligned.fasta + mv ${params.projtag}_ASVs_muscleAligned.fasta-TRIMMED ./${params.projtag}_ASVs_Aligned_informativeonly.fasta #entopy analysis entropy-analysis ${params.projtag}_ASVs_Aligned_informativeonly.fasta #Decomposition @@ -2540,7 +2546,9 @@ if (params.DataCheck || params.Analyze) { """ # Protein_Alignment pre=\$(echo ${prot} | awk -F "_noTax" '{print \$1}' ) - mafft --thread ${task.cpus} --maxiterate 15000 --auto ${prot} >\${pre}_ALN.fasta + if [[ $(grep -c ">" ${prot}) -gt 499 ]]; then algo="super5"; else algo="mpc"; fi + ${tools}/muscle5.0.1278_linux64 -"\${algo}" ${prot} -out \${pre}_ALN.fasta -thread ${task.cpus} -quiet + #mafft --thread ${task.cpus} --maxiterate 15000 --auto ${prot} >\${pre}_ALN.fasta trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html o-trim-uninformative-columns-from-alignment \${pre}_aln.fasta mv \${pre}_aln.fasta-TRIMMED ./\${pre}_Aligned_informativeonly.fasta @@ -2636,12 +2644,14 @@ if (params.DataCheck || params.Analyze) { script: """ #alignment - mafft --thread ${task.cpus} --maxiterate 15000 --auto ${aminos} > ${params.projtag}_AminoTypes_mafftAlign.fasta + if [[ $(grep -c ">" ${aminos}) -gt 499 ]]; then algo="super5"; else algo="mpc"; fi + ${tools}/muscle5.0.1278_linux64 -"\${algo}" ${aminos} -out ${params.projtag}_AminoTypes_muscleAlign.fasta -thread ${task.cpus} -quiet + #mafft --thread ${task.cpus} --maxiterate 15000 --auto ${aminos} > ${params.projtag}_AminoTypes_mafftAlign.fasta #trimming - trimal -in ${params.projtag}_AminoTypes_mafftAlign.fasta -out ${params.projtag}_AminoTypes_mafftAligned.fasta -keepheader -fasta -automated1 - rm ${params.projtag}_AminoTypes_mafftAlign.fasta - o-trim-uninformative-columns-from-alignment ${params.projtag}_AminoTypes_mafftAligned.fasta - mv ${params.projtag}_AminoTypes_mafftAligned.fasta-TRIMMED ./${params.projtag}_AminoTypes_Aligned_informativeonly.fasta + trimal -in ${params.projtag}_AminoTypes_muscleAlign.fasta -out ${params.projtag}_AminoTypes_muscleAligned.fasta -keepheader -fasta -automated1 + rm ${params.projtag}_AminoTypes_muscleAlign.fasta + o-trim-uninformative-columns-from-alignment ${params.projtag}_AminoTypes_muscleAligned.fasta + mv ${params.projtag}_AminoTypes_muscleAligned.fasta-TRIMMED ./${params.projtag}_AminoTypes_Aligned_informativeonly.fasta #entopy analysis entropy-analysis ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta #Decomposition @@ -3168,7 +3178,7 @@ if (params.DataCheck || params.Analyze) { publishDir "${params.workingdir}/${params.outdir}/Analyze/Analyses/pcASV/Nucleotide/Phylogeny/IQ-TREE", mode: "copy", overwrite: true, pattern: '*iq*' input: - tuple nid, file(reads) from pcASV_ntmafft_ch + tuple nid, file(prots) from pcASV_ntmafft_ch output: tuple file("*_aln.fasta"), file("*_aln.html"), file("*.tree"), file("*.log"), file("*iq*"), file("*mt*") into pcASV_nucleotide_phylogeny_results @@ -3177,8 +3187,10 @@ if (params.DataCheck || params.Analyze) { script: mtag="ID=" + nid """ - pre=\$( echo ${reads} | awk -F "_noTax" '{print \$1}' ) - mafft --maxiterate 5000 --auto ${reads} >\${pre}_ALN.fasta + pre=\$( echo ${prots} | awk -F "_noTax" '{print \$1}' ) + if [[ $(grep -c ">" ${prots}) -gt 499 ]]; then algo="super5"; else algo="mpc"; fi + ${tools}/muscle5.0.1278_linux64 -"\${algo}" ${prots} -out \${pre}_ALN.fasta -thread ${task.cpus} -quiet + #mafft --maxiterate 5000 --auto ${reads} >\${pre}_ALN.fasta trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html o-trim-uninformative-columns-from-alignment \${pre}_aln.fasta mv \${pre}_aln.fasta-TRIMMED ./\${pre}_Aligned_informativeonly.fasta @@ -3540,7 +3552,9 @@ if (params.DataCheck || params.Analyze) { mtag="ID=" + nid """ pre=\$( echo ${prot} | awk -F ".fasta" '{print \$1}' ) - mafft --maxiterate 5000 --auto ${prot} >\${pre}_ALN.fasta + if [[ $(grep -c ">" ${prots}) -gt 499 ]]; then algo="super5"; else algo="mpc"; fi + ${tools}/muscle5.0.1278_linux64 -"\${algo}" ${prots} -out \${pre}_ALN.fasta -thread ${task.cpus} -quiet + #mafft --maxiterate 5000 --auto ${prot} >\${pre}_ALN.fasta trimal -in \${pre}_ALN.fasta -out \${pre}_aln.fasta -keepheader -fasta -automated1 -htmlout \${pre}_aln.html o-trim-uninformative-columns-from-alignment \${pre}_aln.fasta mv \${pre}_aln.fasta-TRIMMED ./\${pre}_Aligned_informativeonly.fasta diff --git a/vampirus_env.yml b/vampirus_env.yml index 92ee9d5..9edfd7b 100644 --- a/vampirus_env.yml +++ b/vampirus_env.yml @@ -16,7 +16,6 @@ dependencies: - clustalo=1.2.4 - iqtree=2.0.3 - modeltest-ng=0.1.6 - - mafft=7.464 - vsearch=2.14.2 - biopython=1.76 - bbmap=38.90 diff --git a/vampirus_startup.sh b/vampirus_startup.sh index 0c3f1f3..67fc100 100644 --- a/vampirus_startup.sh +++ b/vampirus_startup.sh @@ -262,7 +262,7 @@ echo "-------------------------------------------------------------------------- cd "$mypwd" echo "Ok, everything downloaded. To test installation, check out the STARTUP_HELP.txt file within "$mypwd" for instructions for testing the installation and running vAMPirus with your own data." chmod a+x "$mypwd"/bin/virtualribosomev2/* - +chmod a+x "$mypwd"/bin/muscle5.0.1278_linux64 if [[ $(ls "$mypwd"| grep -wc "STARTUP_HELP.txt") -eq 0 ]] then touch STARTUP_HELP.txt From a03978b3f7280575b20675479cb05912940b7276 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Wed, 28 Jul 2021 15:50:15 -0500 Subject: [PATCH 114/223] added sensitivty argument to diamond command --- vAMPirus.nf | 24 ++++++++++++------------ vampirus.config | 3 ++- 2 files changed, 14 insertions(+), 13 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 8b6a27b..1e1c086 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1373,7 +1373,7 @@ if (params.DataCheck || params.Analyze) { grep ">" \${virdb} > headers.list headers="headers.list" name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" > access.list echo "[e-value]" > evalue.list @@ -1474,7 +1474,7 @@ if (params.DataCheck || params.Analyze) { grep ">" \${virdb} > headers.list headers="headers.list" name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" > access.list echo "[e-value]" > evalue.list @@ -1707,7 +1707,7 @@ if (params.DataCheck || params.Analyze) { grep ">" \${virdb} > headers.list headers="headers.list" name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" > access.list echo "[e-value]" > evalue.list @@ -1805,7 +1805,7 @@ if (params.DataCheck || params.Analyze) { grep ">" \${virdb} > headers.list headers="headers.list" name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" > access.list echo "[e-value]" > evalue.list @@ -2331,7 +2331,7 @@ if (params.DataCheck || params.Analyze) { grep ">" \${virdb} > headers.list headers="headers.list" name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" > access.list echo "[e-value]" > evalue.list @@ -2429,7 +2429,7 @@ if (params.DataCheck || params.Analyze) { grep ">" \${virdb} > headers.list headers="headers.list" name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" > access.list echo "[e-value]" > evalue.list @@ -2599,7 +2599,7 @@ if (params.DataCheck || params.Analyze) { """ set +e diamond makedb --in ${fasta} --db ${fasta} - diamond blastx -q ${merged} -d ${fasta} -p ${task.cpus} --min-score ${params.ProtCountsBit} --id ${params.ProtCountID} -l ${params.ProtCountsLength} --more-sensitive -o ${params.projtag}_protCounts_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 --max-hsps 1 + diamond blastx -q ${merged} -d ${fasta} -p ${task.cpus} --min-score ${params.ProtCountsBit} --id ${params.ProtCountID} -l ${params.ProtCountsLength} --${sensitivity} -o ${params.projtag}_protCounts_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 --max-hsps 1 echo "OTU_ID" >tmp.col1.txt echo "Generating sample id list" grep ">" ${fasta} | awk -F ">" '{print \$2}' | sort | uniq > otuid.list @@ -2911,7 +2911,7 @@ if (params.DataCheck || params.Analyze) { grep ">" \${virdb} > headers.list headers="headers.list" name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" > access.list echo "[e-value]" > evalue.list @@ -3014,7 +3014,7 @@ if (params.DataCheck || params.Analyze) { grep ">" \${virdb} > headers.list headers="headers.list" name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" > access.list echo "[e-value]" > evalue.list @@ -3331,7 +3331,7 @@ if (params.DataCheck || params.Analyze) { grep ">" \${virdb} > headers.list headers="headers.list" name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" > access.list echo "[e-value]" > evalue.list @@ -3433,7 +3433,7 @@ if (params.DataCheck || params.Analyze) { grep ">" \${virdb} > headers.list headers="headers.list" name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --more-sensitive -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" > access.list echo "[e-value]" > evalue.list @@ -3610,7 +3610,7 @@ if (params.DataCheck || params.Analyze) { set +e potu="\$( echo ${fasta} | awk -F "_" '{print \$3}')" diamond makedb --in ${fasta} --db ${fasta} - diamond blastx -q ${merged} -d ${fasta} -p ${task.cpus} --min-score ${params.ProtCountsBit} --id ${params.ProtCountID} -l ${params.ProtCountsLength} --more-sensitive -o ${params.projtag}_\${potu}_Counts_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 --max-hsps 1 + diamond blastx -q ${merged} -d ${fasta} -p ${task.cpus} --min-score ${params.ProtCountsBit} --id ${params.ProtCountID} -l ${params.ProtCountsLength} --${sensitivity} -o ${params.projtag}_\${potu}_Counts_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 --max-hsps 1 echo "OTU_ID" >tmp.col1.txt echo "Generating sample id list" grep ">" ${fasta} | awk -F ">" '{print \$2}' | sort | uniq > otuid.list diff --git a/vampirus.config b/vampirus.config index c1610aa..831f90f 100644 --- a/vampirus.config +++ b/vampirus.config @@ -109,7 +109,8 @@ params { minID="40" // Set minimum amino acid alignment length for best hit to be counted in taxonomy assignment minaln="30" - + // Set sensitivity parameters for DIAMOND aligner (read more here: https://github.com/bbuchfink/diamond/wiki; default = ultra-sensitive) + sensitivity="ultra-sensitive" // Phylogeny analysis parameters From f8bed36f9dc76cf5e5e23fb88f322aca31bf8d49 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Wed, 28 Jul 2021 15:50:49 -0500 Subject: [PATCH 115/223] removed mafft and updated diamond version to be downloaded --- vampirus_env.yml | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vampirus_env.yml b/vampirus_env.yml index 9edfd7b..cb10503 100644 --- a/vampirus_env.yml +++ b/vampirus_env.yml @@ -10,7 +10,7 @@ channels: dependencies: - python=3.6 - blast=2.11.0 - - diamond=2.0.9 + - diamond=2.0.11 - fastqc=0.11.9 - fastp=0.20.1 - clustalo=1.2.4 From d9ee754911a607d52ccd473309647c43a858facd Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Fri, 30 Jul 2021 11:14:22 -0500 Subject: [PATCH 116/223] added ncbi tax map files to nf and startup --- vAMPirus.nf | 5 ++++- vampirus.config | 4 +++- vampirus_env.yml | 1 + vampirus_startup.sh | 21 ++++++++++++++++++--- 4 files changed, 26 insertions(+), 5 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 1e1c086..f5cfab9 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -566,7 +566,10 @@ if (params.DataCheck || params.Analyze) { cp ${params.vampdir}/Databases/${params.dbname}* ${params.dbdir}/ if [ ! -e ${params.dbdir}/${params.dbname}.dmnd ];then echo "It needs to be built upp, doing it now" - diamond makedb --in ${params.dbdir}/${params.dbname} -d ${params.dbdir}/${params.dbname} + if [[ ${ncbitax} == "true" ]] + then diamond makedb --in ${params.dbdir}/${params.dbname} -d ${params.dbdir}/${params.dbname} --taxonmap ${params.dbdir}/NCBItaxonomy/prot.accession2taxid.FULL --taxonnodes ${params.dbdir}/NCBItaxonomy/nodes.dmp --taxonnames ${params.dbdir}/NCBItaxonomy/names.dmp + else diamond makedb --in ${params.dbdir}/${params.dbname} -d ${params.dbdir}/${params.dbname} + fi export virdb=${params.dbdir}/${params.dbname} else echo "Database looks to be present and built." diff --git a/vampirus.config b/vampirus.config index 831f90f..eaecc9b 100644 --- a/vampirus.config +++ b/vampirus.config @@ -101,7 +101,9 @@ params { headers="HEAD" // Path to directory hmm annotation .txt file - see manual for information on this. Leave as is if not planning on using. dbanno="DATABASEANNOT" - // Set lca="T" if you would like to add taxonomic classification to taxonomy results - example: "ASV1, Viruses::Duplodnaviria::Heunggongvirae::Peploviricota::Herviviricetes::Herpesvirales::Herpesviridae::Gammaherpesvirinae::Macavirus" + // DIAMOND taxonomy inference using NCBI taxmap files (can be downloaded using the startup script using the option -t); set to "true" for this to run + ncbitax="false" + // Set lca="T" if you would like to add taxonomic classification to taxonomy results using annotation information provided by - example: "ASV1, Viruses::Duplodnaviria::Heunggongvirae::Peploviricota::Herviviricetes::Herpesvirales::Herpesviridae::Gammaherpesvirinae::Macavirus" lca="LCAT" // Set minimum bitscore for best hit in taxonomy assignment bitscore="50" diff --git a/vampirus_env.yml b/vampirus_env.yml index cb10503..0957c96 100644 --- a/vampirus_env.yml +++ b/vampirus_env.yml @@ -10,6 +10,7 @@ channels: dependencies: - python=3.6 - blast=2.11.0 + - blast-legacy=2.2.26 - diamond=2.0.11 - fastqc=0.11.9 - fastp=0.20.1 diff --git a/vampirus_startup.sh b/vampirus_startup.sh index 67fc100..3fda920 100644 --- a/vampirus_startup.sh +++ b/vampirus_startup.sh @@ -25,15 +25,18 @@ vampirus_startup.sh -h [-d 1|2|3|4] [-s] [ -s ] Set this option to skip conda installation and environment set up (you can use if you plan to run with Singularity and the vAMPirus Docker container) + [ -t ] Set this option to download NCBI taxonomy files needed for DIAMOND to assign taxonomic classification to sequences + " } -while getopts "hsd:" OPTION; do +while getopts "hstd:" OPTION; do case $OPTION in h) usage; exit;; d) DATABASE=${OPTARG};; s) CONDA="no";; + t) TAX="yes";; esac done shift $((OPTIND-1)) # required, to "eat" the options that have been processed @@ -189,8 +192,8 @@ then mkdir "$mypwd"/Databases cd "$mypwd"/Databases dir="$(pwd)" echo "Database installation: RVDB version 21.0 (latest as of 2021-02)" - curl -o U-RVDBv21.0-prot.fasta.bz2 https://rvdb-prot.pasteur.fr/files/U-RVDBv21.0-prot.fasta.bz2 - bunzip2 U-RVDBv21.0-prot.fasta.bz2 + curl -o U-RVDBv21.0-prot.fasta.xz https://rvdb-prot.pasteur.fr/files/U-RVDBv21.0-prot.fasta.xz + xz -d U-RVDBv21.0-prot.fasta.xz curl -o U-RVDBv21.0-prot-hmm-txt.zip https://rvdb-prot.pasteur.fr/files/U-RVDBv21.0-prot-hmm-txt.zip unzip U-RVDBv21.0-prot-hmm-txt.zip mv annot ./RVDBannot @@ -257,12 +260,24 @@ elif [[ $DATABASE != "" ]] then echo "Error: Database download signaled but not given a value between 1-4" exit 1 fi + +if [[ "$TAX" == "yes" ]] +then mkdir "$mypwd"/Databases/NCBItaxonomy + cd "$mypwd"/Databases/NCBItaxonomy + curl -o prot.accession2taxid.FULL.gz ftp://ftp.ncbi.nlm.nih.gov/pub/taxonomy/accession2taxid/prot.accession2taxid.FULL.gz + gunzip prot.accession2taxid.FULL.gz + curl -o taxdmp.zip ftp://ftp.ncbi.nlm.nih.gov/pub/taxonomy/taxdmp.zip + unzip taxdmp.zip +fi + echo "-------------------------------------------------------------------------------- Database loop done" cd "$mypwd" echo "Ok, everything downloaded. To test installation, check out the STARTUP_HELP.txt file within "$mypwd" for instructions for testing the installation and running vAMPirus with your own data." chmod a+x "$mypwd"/bin/virtualribosomev2/* chmod a+x "$mypwd"/bin/muscle5.0.1278_linux64 + + if [[ $(ls "$mypwd"| grep -wc "STARTUP_HELP.txt") -eq 0 ]] then touch STARTUP_HELP.txt From f13a14f3e53186c62f440bf57d1e7fa044fe7061 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Mon, 2 Aug 2021 16:31:27 -0500 Subject: [PATCH 117/223] Added ncbi taconomy based assigment and LCA from RVDB to all taxonomy processes --- vAMPirus.nf | 392 ++++++++++++++++++++++++++++++++++++-------- vampirus.config | 46 +++--- vampirus_startup.sh | 17 +- 3 files changed, 358 insertions(+), 97 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index f5cfab9..3f73bb5 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -566,7 +566,7 @@ if (params.DataCheck || params.Analyze) { cp ${params.vampdir}/Databases/${params.dbname}* ${params.dbdir}/ if [ ! -e ${params.dbdir}/${params.dbname}.dmnd ];then echo "It needs to be built upp, doing it now" - if [[ ${ncbitax} == "true" ]] + if [[ ${ncbitax} == "true" && ${dbtype} == "NCBI" ]] then diamond makedb --in ${params.dbdir}/${params.dbname} -d ${params.dbdir}/${params.dbname} --taxonmap ${params.dbdir}/NCBItaxonomy/prot.accession2taxid.FULL --taxonnodes ${params.dbdir}/NCBItaxonomy/nodes.dmp --taxonnames ${params.dbdir}/NCBItaxonomy/names.dmp else diamond makedb --in ${params.dbdir}/${params.dbname} -d ${params.dbdir}/${params.dbname} fi @@ -1348,7 +1348,7 @@ if (params.DataCheck || params.Analyze) { if (!params.skipTaxonomy) { - if (params.headers == "RefSeq") { + if (params.dbtype == "NCBI") { process ncASV_Taxonomy_Inference_RefSeq { /////// editttt @@ -1376,7 +1376,10 @@ if (params.DataCheck || params.Analyze) { grep ">" \${virdb} > headers.list headers="headers.list" name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + if [[ ${ncbitax} == "true" ]] + then diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop staxids sskingdoms skingdoms sphylums --max-target-seqs 1 --max-hsps 1 + else diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + fi echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" > access.list echo "[e-value]" > evalue.list @@ -1387,6 +1390,15 @@ if (params.DataCheck || params.Analyze) { echo "[ncASV#]" > otu.list echo "[Sequence length]" > length.list grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst + if [[ ${params.lca} == "T" ]] + then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list + echo "[Taxonomic classification from RVDB annotations]" > lca_classification.list + else + echo "[Taxonomic classification from RVDB annotations]" > lca_classification.list + fi + if [[ ${ncbitax} == "true" ]] + then echo "[NCBI Taxonomy ID],[Taxonomic classification from NCBI]" > ncbi_classification.list + fi echo "extracting genes and names" touch new_"\$name"_asvnames.txt for s in \$(cat seqids.lst);do @@ -1408,6 +1420,17 @@ if (params.DataCheck || params.Analyze) { virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list echo ">"\${s}"_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + if [[ "${params.lca}" == "T" ]] + then if [[ \$(grep -w "\$acc" ${params.dbanno}/*.txt | wc -l) -eq 1 ]] + then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}') + lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') + echo "\$lcla" >> lca_classification.list + else echo "Viruses::unclassified" >> lca_classification.list + fi + fi + if [[ ${ncbitax} == "true" ]] + then echo "\$line" | awk -F "\t" '{print \$14","\$16"::"\$18"::"\$17}' >> ncbi_classification.list + fi echo "\$s done." else echo "Ugh, there was no hit for \$s .." @@ -1423,6 +1446,12 @@ if (params.DataCheck || params.Analyze) { virus="NO" gene="HIT" echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + if [[ "${params.lca}" == "T" ]] + then echo "N/A" >> lca_classification.list + fi + if [[ "${ncbitax}" == "true" ]] + then echo "N/A" >> ncbi_classification.list + fi echo "\$s done." fi done @@ -1436,8 +1465,26 @@ if (params.DataCheck || params.Analyze) { echo " " > sequence.list grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + if [[ "${params.lca}" == "T" && "${ncbitax}" == "true" ]] + then + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list lca_classification.list ncbi_classification.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list lca_classification.list ncbi_classification.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list lca_classification.list ncbi_classification.list >> \${name}_quick_Taxbreakdown.csv + elif [[ "${params.lca}" == "T" && "${ncbitax}" != "true" ]] + then + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list lca_classification.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list lca_classification.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list lca_classification.list >> \${name}_quick_Taxbreakdown.csv + elif [[ "${ncbitax}" == "true" && "${params.lca}" != "T"]] + then + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list ncbi_classification.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list ncbi_classification.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list ncbi_classification.list >> \${name}_quick_Taxbreakdown.csv + else + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + echo "skipped" >> \${name}_quick_Taxbreakdown.csv + fi for x in *phyloformat.csv;do echo "\$x" lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) @@ -1449,7 +1496,7 @@ if (params.DataCheck || params.Analyze) { rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list """ } - } else if (params.headers == "RVDB") { + } else if (params.dbtype== "RVDB") { process ncASV_Taxonomy_Inference_RVDB { /////// editttt @@ -1490,9 +1537,9 @@ if (params.DataCheck || params.Analyze) { grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst if [[ ${params.lca} == "T" ]] then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list - echo "[Classification]" > classfication.list + echo "[Taxonomic classification from RVDB annotations]" > lca_classification.list else echo "skipped" >> \${name}_quick_Taxbreakdown.csv - echo "[Classification]" > classfication.list + echo "[Taxonomic classification from RVDB annotations]" > lca_classification.list fi echo "extracting genes and names" touch new_"\$name"_asvnames.txt @@ -1516,9 +1563,12 @@ if (params.DataCheck || params.Analyze) { echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt if [[ "${params.lca}" == "T" ]] - then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}') - lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') - echo "\$lcla" >> classfication.list + then if [[ \$(grep -w "\$acc" ${params.dbanno}/*.txt | wc -l) -eq 1 ]] + then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}') + lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') + echo "\$lcla" >> lca_classification.list + else echo "Viruses::unclassified" >> lca_classification.list + fi fi echo "\$s done." else @@ -1536,7 +1586,7 @@ if (params.DataCheck || params.Analyze) { gene="HIT" echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt if [[ "${params.lca}" == "T" ]] - then echo "N/A" >> classfication.list + then echo "N/A" >> lca_classification.list fi echo "\$s done." fi @@ -1553,9 +1603,9 @@ if (params.DataCheck || params.Analyze) { grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list rm "\$name"_tmpssasv.fasta if [[ "${params.lca}" == "T" ]] - then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list classfication.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv + then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list lca_classification.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list lca_classification.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list lca_classification.list >> \${name}_quick_Taxbreakdown.csv else paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv fi @@ -1685,7 +1735,7 @@ if (params.DataCheck || params.Analyze) { if (!params.skipTaxonomy) { - if (params.headers == "RefSeq") { + if (params.dbtype == "NCBI") { process ASV_Taxonomy_Inference_RefSeq { /////// editttt @@ -1710,7 +1760,10 @@ if (params.DataCheck || params.Analyze) { grep ">" \${virdb} > headers.list headers="headers.list" name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + if [[ ${ncbitax} == "true" ]] + then diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop staxids sskingdoms skingdoms sphylums --max-target-seqs 1 --max-hsps 1 + else diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + fi echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" > access.list echo "[e-value]" > evalue.list @@ -1721,6 +1774,15 @@ if (params.DataCheck || params.Analyze) { echo "[ASV#]" > otu.list echo "[Sequence length]" > length.list grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst + if [[ ${params.lca} == "T" ]] + then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list + echo "[Taxonomic classification from RVDB annotations]" > lca_classification.list + else + echo "[Taxonomic classification from RVDB annotations]" > lca_classification.list + fi + if [[ ${ncbitax} == "true" ]] + then echo "[NCBI Taxonomy ID],[Taxonomic classification from NCBI]" > ncbi_classification.list + fi echo "extracting genes and names" touch new_"\$name"_asvnames.txt for s in \$(cat seqids.lst);do @@ -1742,6 +1804,17 @@ if (params.DataCheck || params.Analyze) { virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list echo ">"\${s}"_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + if [[ "${params.lca}" == "T" ]] + then if [[ \$(grep -w "\$acc" ${params.dbanno}/*.txt | wc -l) -eq 1 ]] + then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}') + lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') + echo "\$lcla" >> lca_classification.list + else echo "Viruses::unclassified" >> lca_classification.list + fi + fi + if [[ ${ncbitax} == "true" ]] + then echo "\$line" | awk -F "\t" '{print \$14","\$16"::"\$18"::"\$17}' >> ncbi_classification.list + fi echo "\$s done." else echo "Ugh, there was no hit for \$s .." @@ -1757,6 +1830,12 @@ if (params.DataCheck || params.Analyze) { virus="NO" gene="HIT" echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + if [[ "${params.lca}" == "T" ]] + then echo "N/A" >> lca_classification.list + fi + if [[ "${ncbitax}" == "true" ]] + then echo "N/A" >> ncbi_classification.list + fi echo "\$s done." fi done @@ -1770,8 +1849,26 @@ if (params.DataCheck || params.Analyze) { echo " " > sequence.list grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + if [[ "${params.lca}" == "T" && "${ncbitax}" == "true" ]] + then + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list lca_classification.list ncbi_classification.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list lca_classification.list ncbi_classification.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list lca_classification.list ncbi_classification.list >> \${name}_quick_Taxbreakdown.csv + elif [[ "${params.lca}" == "T" && "${ncbitax}" != "true" ]] + then + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list lca_classification.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list lca_classification.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list lca_classification.list >> \${name}_quick_Taxbreakdown.csv + elif [[ "${ncbitax}" == "true" && "${params.lca}" != "T"]] + then + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list ncbi_classification.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list ncbi_classification.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list ncbi_classification.list >> \${name}_quick_Taxbreakdown.csv + else + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + echo "skipped" >> \${name}_quick_Taxbreakdown.csv + fi for x in *phyloformat.csv;do echo "\$x" lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) @@ -1783,7 +1880,7 @@ if (params.DataCheck || params.Analyze) { rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list """ } - } else if (params.headers == "RVDB") { + } else if (params.dbtype== "RVDB") { process ASV_Taxonomy_Inference_RVDB { /////// editttt @@ -1821,9 +1918,9 @@ if (params.DataCheck || params.Analyze) { grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst if [[ ${params.lca} == "T" ]] then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list - echo "[Classification]" > classfication.list + echo "[Taxonomic classification from RVDB annotations]" > lca_classification.list else echo "skipped" >> \${name}_quick_Taxbreakdown.csv - echo "[Classification]" > classfication.list + echo "[Taxonomic classification from RVDB annotations]" > lca_classification.list fi echo "extracting genes and names" touch new_"\$name"_asvnames.txt @@ -1847,9 +1944,12 @@ if (params.DataCheck || params.Analyze) { echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt if [[ "${params.lca}" == "T" ]] - then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}') - lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') - echo "\$lcla" >> classfication.list + then if [[ \$(grep -w "\$acc" ${params.dbanno}/*.txt | wc -l) -eq 1 ]] + then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}') + lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') + echo "\$lcla" >> lca_classification.list + else echo "Viruses::unclassified" >> lca_classification.list + fi fi echo "\$s done." else @@ -1867,7 +1967,7 @@ if (params.DataCheck || params.Analyze) { gene="HIT" echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt if [[ "${params.lca}" == "T" ]] - then echo "N/A" >> classfication.list + then echo "N/A" >> lca_classification.list fi echo "\$s done." fi @@ -1884,9 +1984,9 @@ if (params.DataCheck || params.Analyze) { grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list rm "\$name"_tmpssasv.fasta if [[ "${params.lca}" == "T" ]] - then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list classfication.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv + then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list lca_classification.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list lca_classification.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list lca_classification.list >> \${name}_quick_Taxbreakdown.csv else paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv fi @@ -2309,7 +2409,7 @@ if (params.DataCheck || params.Analyze) { if (!params.skipTaxonomy) { - if (params.headers == "RefSeq") { + if (params.dbtype == "NCBI") { process AminoType_Taxonomy_Inference_RefSeq { @@ -2334,7 +2434,10 @@ if (params.DataCheck || params.Analyze) { grep ">" \${virdb} > headers.list headers="headers.list" name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + if [[ ${ncbitax} == "true" ]] + then diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop staxids sskingdoms skingdoms sphylums --max-target-seqs 1 --max-hsps 1 + else diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + fi echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" > access.list echo "[e-value]" > evalue.list @@ -2345,6 +2448,15 @@ if (params.DataCheck || params.Analyze) { echo "[AminoType#]" > otu.list echo "[Sequence length]" > length.list grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst + if [[ ${params.lca} == "T" ]] + then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list + echo "[Taxonomic classification from RVDB annotations]" > lca_classification.list + else + echo "[Taxonomic classification from RVDB annotations]" > lca_classification.list + fi + if [[ ${ncbitax} == "true" ]] + then echo "[NCBI Taxonomy ID],[Taxonomic classification from NCBI]" > ncbi_classification.list + fi echo "extracting genes and names" touch new_"\$name"_asvnames.txt for s in \$(cat seqids.lst);do @@ -2366,6 +2478,17 @@ if (params.DataCheck || params.Analyze) { virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list echo ">"\${s}"_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + if [[ "${params.lca}" == "T" ]] + then if [[ \$(grep -w "\$acc" ${params.dbanno}/*.txt | wc -l) -eq 1 ]] + then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}') + lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') + echo "\$lcla" >> lca_classification.list + else echo "Viruses::unclassified" >> lca_classification.list + fi + fi + if [[ ${ncbitax} == "true" ]] + then echo "\$line" | awk -F "\t" '{print \$14","\$16"::"\$18"::"\$17}' >> ncbi_classification.list + fi echo "\$s done." else echo "Ugh, there was no hit for \$s .." @@ -2381,6 +2504,12 @@ if (params.DataCheck || params.Analyze) { virus="NO" gene="HIT" echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + if [[ "${params.lca}" == "T" ]] + then echo "N/A" >> lca_classification.list + fi + if [[ "${ncbitax}" == "true" ]] + then echo "N/A" >> ncbi_classification.list + fi echo "\$s done." fi done @@ -2394,8 +2523,26 @@ if (params.DataCheck || params.Analyze) { echo " " > sequence.list grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + if [[ "${params.lca}" == "T" && "${ncbitax}" == "true" ]] + then + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list lca_classification.list ncbi_classification.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list lca_classification.list ncbi_classification.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list lca_classification.list ncbi_classification.list >> \${name}_quick_Taxbreakdown.csv + elif [[ "${params.lca}" == "T" && "${ncbitax}" != "true" ]] + then + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list lca_classification.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list lca_classification.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list lca_classification.list >> \${name}_quick_Taxbreakdown.csv + elif [[ "${ncbitax}" == "true" && "${params.lca}" != "T"]] + then + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list ncbi_classification.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list ncbi_classification.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list ncbi_classification.list >> \${name}_quick_Taxbreakdown.csv + else + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + echo "skipped" >> \${name}_quick_Taxbreakdown.csv + fi for x in *phyloformat.csv;do echo "\$x" lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) @@ -2407,7 +2554,7 @@ if (params.DataCheck || params.Analyze) { rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list """ } - } else if (params.headers == "RVDB") { + } else if (params.dbtype== "RVDB") { process AminoType_Taxonomy_Inference_RVDB { @@ -2445,9 +2592,9 @@ if (params.DataCheck || params.Analyze) { grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst if [[ ${params.lca} == "T" ]] then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list - echo "[Classification]" > classfication.list + echo "[Taxonomic classification from RVDB annotations]" > lca_classification.list else echo "skipped" >> \${name}_quick_Taxbreakdown.csv - echo "[Classification]" > classfication.list + echo "[Taxonomic classification from RVDB annotations]" > lca_classification.list fi echo "extracting genes and names" touch new_"\$name"_asvnames.txt @@ -2471,9 +2618,12 @@ if (params.DataCheck || params.Analyze) { echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt if [[ "${params.lca}" == "T" ]] - then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}') - lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') - echo "\$lcla" >> classfication.list + then if [[ \$(grep -w "\$acc" ${params.dbanno}/*.txt | wc -l) -eq 1 ]] + then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}') + lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') + echo "\$lcla" >> lca_classification.list + else echo "Viruses::unclassified" >> lca_classification.list + fi fi echo "\$s done." else @@ -2491,7 +2641,7 @@ if (params.DataCheck || params.Analyze) { gene="HIT" echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt if [[ "${params.lca}" == "T" ]] - then echo "N/A" >> classfication.list + then echo "N/A" >> lca_classification.list fi echo "\$s done." fi @@ -2508,9 +2658,9 @@ if (params.DataCheck || params.Analyze) { grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list rm "\$name"_tmpssasv.fasta if [[ "${params.lca}" == "T" ]] - then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list classfication.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv + then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list lca_classification.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list lca_classification.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list lca_classification.list >> \${name}_quick_Taxbreakdown.csv else paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv fi @@ -2884,7 +3034,7 @@ if (params.DataCheck || params.Analyze) { if (!params.skipTaxonomy) { - if (params.headers == "RefSeq") { + if (params.dbtype == "NCBI") { process pcASV_Nucleotide_Taxonomy_Inference_RefSeq { @@ -2914,7 +3064,10 @@ if (params.DataCheck || params.Analyze) { grep ">" \${virdb} > headers.list headers="headers.list" name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + if [[ ${ncbitax} == "true" ]] + then diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop staxids sskingdoms skingdoms sphylums --max-target-seqs 1 --max-hsps 1 + else diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + fi echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" > access.list echo "[e-value]" > evalue.list @@ -2925,6 +3078,15 @@ if (params.DataCheck || params.Analyze) { echo "[pcASV#]" > otu.list echo "[Sequence length]" > length.list grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst + if [[ ${params.lca} == "T" ]] + then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list + echo "[Taxonomic classification from RVDB annotations]" > lca_classification.list + else + echo "[Taxonomic classification from RVDB annotations]" > lca_classification.list + fi + if [[ ${ncbitax} == "true" ]] + then echo "[NCBI Taxonomy ID],[Taxonomic classification from NCBI]" > ncbi_classification.list + fi echo "extracting genes and names" touch new_"\$name"_asvnames.txt for s in \$(cat seqids.lst);do @@ -2946,6 +3108,17 @@ if (params.DataCheck || params.Analyze) { virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list echo ">"\${s}"_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + if [[ "${params.lca}" == "T" ]] + then if [[ \$(grep -w "\$acc" ${params.dbanno}/*.txt | wc -l) -eq 1 ]] + then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}') + lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') + echo "\$lcla" >> lca_classification.list + else echo "Viruses::unclassified" >> lca_classification.list + fi + fi + if [[ ${ncbitax} == "true" ]] + then echo "\$line" | awk -F "\t" '{print \$14","\$16"::"\$18"::"\$17}' >> ncbi_classification.list + fi echo "\$s done." else echo "Ugh, there was no hit for \$s .." @@ -2961,6 +3134,12 @@ if (params.DataCheck || params.Analyze) { virus="NO" gene="HIT" echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + if [[ "${params.lca}" == "T" ]] + then echo "N/A" >> lca_classification.list + fi + if [[ "${ncbitax}" == "true" ]] + then echo "N/A" >> ncbi_classification.list + fi echo "\$s done." fi done @@ -2974,8 +3153,26 @@ if (params.DataCheck || params.Analyze) { echo " " > sequence.list grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + if [[ "${params.lca}" == "T" && "${ncbitax}" == "true" ]] + then + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list lca_classification.list ncbi_classification.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list lca_classification.list ncbi_classification.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list lca_classification.list ncbi_classification.list >> \${name}_quick_Taxbreakdown.csv + elif [[ "${params.lca}" == "T" && "${ncbitax}" != "true" ]] + then + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list lca_classification.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list lca_classification.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list lca_classification.list >> \${name}_quick_Taxbreakdown.csv + elif [[ "${ncbitax}" == "true" && "${params.lca}" != "T"]] + then + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list ncbi_classification.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list ncbi_classification.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list ncbi_classification.list >> \${name}_quick_Taxbreakdown.csv + else + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + echo "skipped" >> \${name}_quick_Taxbreakdown.csv + fi for x in *phyloformat.csv;do echo "\$x" lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) @@ -2987,7 +3184,7 @@ if (params.DataCheck || params.Analyze) { rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list """ } - } else if (params.headers == "RVDB") { + } else if (params.dbtype== "RVDB") { process pcASV_Nucleotide_Taxonomy_Inference_RVDB { @@ -3030,9 +3227,9 @@ if (params.DataCheck || params.Analyze) { grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst if [[ ${params.lca} == "T" ]] then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list - echo "[Classification]" > classfication.list + echo "[Taxonomic classification from RVDB annotations]" > lca_classification.list else echo "skipped" >> \${name}_quick_Taxbreakdown.csv - echo "[Classification]" > classfication.list + echo "[Taxonomic classification from RVDB annotations]" > lca_classification.list fi echo "extracting genes and names" touch new_"\$name"_asvnames.txt @@ -3056,9 +3253,12 @@ if (params.DataCheck || params.Analyze) { echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt if [[ "${params.lca}" == "T" ]] - then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}') - lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') - echo "\$lcla" >> classfication.list + then if [[ \$(grep -w "\$acc" ${params.dbanno}/*.txt | wc -l) -eq 1 ]] + then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}') + lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') + echo "\$lcla" >> lca_classification.list + else echo "Viruses::unclassified" >> lca_classification.list + fi fi echo "\$s done." else @@ -3076,7 +3276,7 @@ if (params.DataCheck || params.Analyze) { gene="HIT" echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt if [[ "${params.lca}" == "T" ]] - then echo "N/A" >> classfication.list + then echo "N/A" >> lca_classification.list fi echo "\$s done." fi @@ -3093,9 +3293,9 @@ if (params.DataCheck || params.Analyze) { grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list rm "\$name"_tmpssasv.fasta if [[ "${params.lca}" == "T" ]] - then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list classfication.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv + then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list lca_classification.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list lca_classification.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list lca_classification.list >> \${name}_quick_Taxbreakdown.csv else paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv fi @@ -3305,7 +3505,7 @@ if (params.DataCheck || params.Analyze) { if (!params.skipTaxonomy) { - if (params.headers == "RefSeq") { + if (params.dbtype == "NCBI") { process pcASV_AminoAcid_Taxonomy_Inference_RefSeq { @@ -3334,7 +3534,10 @@ if (params.DataCheck || params.Analyze) { grep ">" \${virdb} > headers.list headers="headers.list" name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + if [[ ${ncbitax} == "true" ]] + then diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop staxids sskingdoms skingdoms sphylums --max-target-seqs 1 --max-hsps 1 + else diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + fi echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" > access.list echo "[e-value]" > evalue.list @@ -3345,6 +3548,15 @@ if (params.DataCheck || params.Analyze) { echo "[pcASV#]" > otu.list echo "[Sequence length]" > length.list grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst + if [[ ${params.lca} == "T" ]] + then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list + echo "[Taxonomic classification from RVDB annotations]" > lca_classification.list + else echo "skipped" >> \${name}_quick_Taxbreakdown.csv + echo "[Taxonomic classification from RVDB annotations]" > lca_classification.list + fi + if [[ ${ncbitax} == "true" ]] + then echo "[NCBI Taxonomy ID],[Taxonomic classification from NCBI]" > ncbi_classification.list + fi echo "extracting genes and names" touch new_"\$name"_asvnames.txt for s in \$(cat seqids.lst);do @@ -3366,6 +3578,17 @@ if (params.DataCheck || params.Analyze) { virus=\$(grep -w "\$acc" "\$headers" | awk -F "[" '{ print \$2 }' | awk -F "]" '{ print \$1 }'| sed 's/ /_/g') echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list echo ">"\${s}"_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + if [[ "${params.lca}" == "T" ]] + then if [[ \$(grep -w "\$acc" ${params.dbanno}/*.txt | wc -l) -eq 1 ]] + then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}') + lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') + echo "\$lcla" >> lca_classification.list + else echo "Viruses::unclassified" >> lca_classification.list + fi + fi + if [[ ${ncbitax} == "true" ]] + then echo "\$line" | awk -F "\t" '{print \$14","\$16"::"\$18"::"\$17}' >> ncbi_classification.list + fi echo "\$s done." else echo "Ugh, there was no hit for \$s .." @@ -3381,6 +3604,12 @@ if (params.DataCheck || params.Analyze) { virus="NO" gene="HIT" echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt + if [[ "${params.lca}" == "T" ]] + then echo "N/A" >> lca_classification.list + fi + if [[ "${ncbitax}" == "true" ]] + then echo "N/A" >> ncbi_classification.list + fi echo "\$s done." fi done @@ -3394,8 +3623,26 @@ if (params.DataCheck || params.Analyze) { echo " " > sequence.list grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list rm "\$name"_tmpssasv.fasta - paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + if [[ "${params.lca}" == "T" && "${ncbitax}" == "true" ]] + then + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list lca_classification.list ncbi_classification.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list lca_classification.list ncbi_classification.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list lca_classification.list ncbi_classification.list >> \${name}_quick_Taxbreakdown.csv + elif [[ "${params.lca}" == "T" && "${ncbitax}" != "true" ]] + then + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list lca_classification.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list lca_classification.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list lca_classification.list >> \${name}_quick_Taxbreakdown.csv + elif [[ "${ncbitax}" == "true" && "${params.lca}" != "T"]] + then + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list ncbi_classification.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list ncbi_classification.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list ncbi_classification.list >> \${name}_quick_Taxbreakdown.csv + else + paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + echo "skipped" >> \${name}_quick_Taxbreakdown.csv + fi for x in *phyloformat.csv;do echo "\$x" lin=\$(( \$(wc -l \$x | awk '{print \$1}')-1)) @@ -3407,7 +3654,7 @@ if (params.DataCheck || params.Analyze) { rm evalue.list sequence.list bit.list pid.list length.list seqids.lst otu.list *asvnames.txt "\$name"_virus.list "\$name"_genes.list newnames.list access.list headers.list """ } - } else if (params.headers == "RVDB") { + } else if (params.dbtype== "RVDB") { process pcASV_AminoAcid_Taxonomy_Inference_RVDB { @@ -3449,9 +3696,9 @@ if (params.DataCheck || params.Analyze) { grep ">" ${asvs} | awk -F ">" '{print \$2}' > seqids.lst if [[ ${params.lca} == "T" ]] then grep -w "LCA" ${params.dbanno}/*.txt > lcainfo.list - echo "[Classification]" > classfication.list + echo "[Taxonomic classification from RVDB annotations]" > lca_classification.list else echo "skipped" >> \${name}_quick_Taxbreakdown.csv - echo "[Classification]" > classfication.list + echo "[Taxonomic classification from RVDB annotations]" > lca_classification.list fi echo "extracting genes and names" touch new_"\$name"_asvnames.txt @@ -3475,9 +3722,12 @@ if (params.DataCheck || params.Analyze) { echo "\$virus" | sed 's/_/ /g' >> "\$name"_virus.list echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt if [[ "${params.lca}" == "T" ]] - then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}') - lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') - echo "\$lcla" >> classfication.list + then if [[ \$(grep -w "\$acc" ${params.dbanno}/*.txt | wc -l) -eq 1 ]] + then group=\$(grep -w "\$acc" ${params.dbanno}/*.txt | awk -F ":" '{print \$1}') + lcla=\$(grep -w "\$group" lcainfo.list | awk -F "\t" '{print \$2}') + echo "\$lcla" >> lca_classification.list + else echo "Viruses::unclassified" >> lca_classification.list + fi fi echo "\$s done." else @@ -3495,7 +3745,7 @@ if (params.DataCheck || params.Analyze) { gene="HIT" echo ">\${s}_"\$virus"_"\$gene"" >> new_"\$name"_asvnames.txt if [[ "${params.lca}" == "T" ]] - then echo "N/A" >> classfication.list + then echo "N/A" >> lca_classification.list fi echo "\$s done." fi @@ -3512,9 +3762,9 @@ if (params.DataCheck || params.Analyze) { grep -v ">" "\$name"_tmpssasv.fasta >> sequence.list rm "\$name"_tmpssasv.fasta if [[ "${params.lca}" == "T" ]] - then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list classfication.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv - paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv - paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list classfication.list >> \${name}_quick_Taxbreakdown.csv + then paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list lca_classification.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv + paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list lca_classification.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv + paste -d"," otu.list access.list "\$name"_virus.list "\$name"_genes.list lca_classification.list >> \${name}_quick_Taxbreakdown.csv else paste -d "," sequence.list "\$name"_virus.list "\$name"_genes.list otu.list newnames.list length.list bit.list evalue.list pid.list access.list >> "\$name"_phyloformat.csv paste -d"\t" otu.list access.list "\$name"_virus.list "\$name"_genes.list sequence.list length.list bit.list evalue.list pid.list >> "\$name"_summaryTable.tsv fi diff --git a/vampirus.config b/vampirus.config index eaecc9b..de3df9d 100644 --- a/vampirus.config +++ b/vampirus.config @@ -93,26 +93,32 @@ params { // Taxonomy inference parameters - // Specify name of database to use for analysis - dbname="DATABASENAME" - // Path to Directory where database is being stored - dbdir="DATABASEDIR" - // Set taxonomy database header format -> headers= "RefSeq" to toggle use of RefSeq header format; set to "RVDB" to signal the use of Reverence Viral DataBase (RVDB) headers - headers="HEAD" - // Path to directory hmm annotation .txt file - see manual for information on this. Leave as is if not planning on using. - dbanno="DATABASEANNOT" - // DIAMOND taxonomy inference using NCBI taxmap files (can be downloaded using the startup script using the option -t); set to "true" for this to run - ncbitax="false" - // Set lca="T" if you would like to add taxonomic classification to taxonomy results using annotation information provided by - example: "ASV1, Viruses::Duplodnaviria::Heunggongvirae::Peploviricota::Herviviricetes::Herpesvirales::Herpesviridae::Gammaherpesvirinae::Macavirus" - lca="LCAT" - // Set minimum bitscore for best hit in taxonomy assignment - bitscore="50" - // Set minimum percent amino acid similarity for best hit to be counted in taxonomy assignment - minID="40" - // Set minimum amino acid alignment length for best hit to be counted in taxonomy assignment - minaln="30" - // Set sensitivity parameters for DIAMOND aligner (read more here: https://github.com/bbuchfink/diamond/wiki; default = ultra-sensitive) - sensitivity="ultra-sensitive" + //Parameters for diamond command + // Set minimum bitscore for best hit in taxonomy assignment + bitscore="50" + // Set minimum percent amino acid similarity for best hit to be counted in taxonomy assignment + minID="40" + // Set minimum amino acid alignment length for best hit to be counted in taxonomy assignment + minaln="30" + // Set sensitivity parameters for DIAMOND aligner (read more here: https://github.com/bbuchfink/diamond/wiki; default = ultra-sensitive) + sensitivity="ultra-sensitive" + + // Database information + // Specify name of database to use for analysis + dbname="DATABASENAME" + // Path to Directory where database is being stored - vAMPirus will look here to make sure the database with the name provided above is present and built + dbdir="DATABASEDIR" + // Set database type (NCBI or RVDB). Lets vAMPirus know which sequence header format is being used and must be set to NCBI when using RefSeq or Non-Redundant databases. -> dbtype="NCBI" to toggle use of RefSeq header format; set to "RVDB" to signal the use of Reverence Viral DataBase (RVDB) headers (see manual) + dbtype="TYPE" + + // Classification settings - if planning on inferring LCA from RVDB annotation files OR using NCBI taxonomy files, confirm options below are accurate. + // Path to directory RVDB hmm annotation .txt file - see manual for information on this. Leave as is if not planning on using. + dbanno="DATABASEANNOT" + // Set lca="T" if you would like to add "Least Common Ancestor" classifications to taxonomy results using information provided by RVDB annotation files (works when using NCBI or RVDB databases) - example: "ASV1, Viruses::Duplodnaviria::Heunggongvirae::Peploviricota::Herviviricetes::Herpesvirales::Herpesviridae::Gammaherpesvirinae::Macavirus" + lca="LCAT" + // DIAMOND taxonomy inference using NCBI taxmap files (can be downloaded using the startup script using the option -t); set to "true" for this to run (ONLY WORKS WITH dbtype="NCBI") + ncbitax="false" + // Phylogeny analysis parameters diff --git a/vampirus_startup.sh b/vampirus_startup.sh index 3fda920..7730296 100644 --- a/vampirus_startup.sh +++ b/vampirus_startup.sh @@ -18,14 +18,13 @@ vampirus_startup.sh -h [-d 1|2|3|4] [-s] [ -d 1|2|3|4 ] Set this option to create a database directiory within the current working directory and download the following databases for taxonomy assignment: 1 - Download the proteic version of the Reference Viral DataBase (See the paper for more information on this database: https://f1000research.com/articles/8-530) - NOTE: need to use RVDB for sequence classification 2 - Download only NCBIs Viral protein RefSeq database 3 - Download only the complete NCBI NR protein database 4 - Download all three databases [ -s ] Set this option to skip conda installation and environment set up (you can use if you plan to run with Singularity and the vAMPirus Docker container) - [ -t ] Set this option to download NCBI taxonomy files needed for DIAMOND to assign taxonomic classification to sequences + [ -t ] Set this option to download NCBI taxonomy files needed for DIAMOND to assign taxonomic classification to sequences (works with NCBI type databases only, see manual for more information) " @@ -196,11 +195,11 @@ then mkdir "$mypwd"/Databases xz -d U-RVDBv21.0-prot.fasta.xz curl -o U-RVDBv21.0-prot-hmm-txt.zip https://rvdb-prot.pasteur.fr/files/U-RVDBv21.0-prot-hmm-txt.zip unzip U-RVDBv21.0-prot-hmm-txt.zip - mv annot ./RVDBannot + mv annot ./RVDBannot/ echo "Editing confiration file for you now..." sed 's/DATABASENAME/U-RVDBv21.0-prot.fasta/g' "$mypwd"/vampirus.config > tmp1.config sed "s|DATABASEDIR|${dir}|g" tmp1.config > tmp2.config - sed "s|DATABASEANNOT|${dir}/RVDBannot|g" tmp2.config | sed "s|LCAT|T|g" | sed 's/HEAD/RVDB/g' > tmp3.config + sed "s|DATABASEANNOT|${dir}/RVDBannot|g" tmp2.config | sed "s|LCAT|T|g" | sed 's/TYPE/RVDB/g' > tmp3.config rm tmp1.config rm tmp2.config cat tmp3.config > "$mypwd"/vampirus.config @@ -221,9 +220,12 @@ then mkdir "$mypwd"/Databases gunzip viral.3.protein.faa.gz cat viral.3.protein.faa >> complete_virus_refseq_prot.fasta rm viral.*.protein.faa + curl -o U-RVDBv21.0-prot-hmm-txt.zip https://rvdb-prot.pasteur.fr/files/U-RVDBv21.0-prot-hmm-txt.zip + unzip U-RVDBv21.0-prot-hmm-txt.zip + mv annot ./RVDBannot echo "Editing confiration file for you now..." sed 's/DATABASENAME/complete_virus_refseq_prot.fasta/g' "$mypwd"/vampirus.config > tmp1.config - sed "s|DATABASEDIR|${dir}|g" tmp1.config | sed 's/HEAD/RefSeq/g' > tmp2.config + sed "s|DATABASEDIR|${dir}|g" tmp1.config | sed "s|DATABASEANNOT|${dir}/RVDBannot|g" | sed 's/TYPE/NCBI/g' > tmp2.config rm tmp1.config cat tmp2.config > "$mypwd"/vampirus.config rm tmp2.config @@ -235,9 +237,12 @@ then mkdir "$mypwd"/Databases echo "Database installation: NCBI NR protein database (should be the most up to date at time of running this script)" curl -o NCBI_nr_proteindb.faa.gz https://ftp.ncbi.nlm.nih.gov/blast/db/FASTA/nr.gz gunzip NCBI_nr_proteindb.faa.gz + curl -o U-RVDBv21.0-prot-hmm-txt.zip https://rvdb-prot.pasteur.fr/files/U-RVDBv21.0-prot-hmm-txt.zip + unzip U-RVDBv21.0-prot-hmm-txt.zip + mv annot ./RVDBannot echo "Editing confiration file for you now..." sed 's/DATABASENAME/NCBI_nr_proteindb.faa/g' "$mypwd"/vampirus.config > tmp1.config - sed "s|DATABASEDIR|${dir}|g" tmp1.config | sed 's/HEAD/RefSeq/g' > tmp2.config + sed "s|DATABASEDIR|${dir}|g" tmp1.config | sed "s|DATABASEANNOT|${dir}/RVDBannot|g" | sed 's/TYPE/NCBI/g' > tmp2.config rm tmp1.config cat tmp2.config > "$mypwd"/vampirus.config rm tmp2.config From c8acefa4a3007588efd2122e4a52e9a503ca0e2a Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Tue, 3 Aug 2021 08:32:24 -0500 Subject: [PATCH 118/223] added by position MED analysis --- vAMPirus.nf | 30 +++++++++++++++++++++--------- vampirus.config | 4 ++-- 2 files changed, 23 insertions(+), 11 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 3f73bb5..ee40f57 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -175,7 +175,7 @@ def helpMessage() { --dbdir Path to Directory where database is being stored - --headers Set taxonomy database header format -> headers= "RefSeq" to toggle use of RefSeq header format; set to "RVDB" to signal the use of Reverence Viral DataBase (RVDB) headers + --headers Set taxonomy database header format -> headers= "NCBI" to toggle use of NCBI header format; set to "RVDB" to signal the use of Reverence Viral DataBase (RVDB) headers --dbanno Path to directory hmm annotation .txt file - see manual for information on this. Leave as is if not planning on using. @@ -387,7 +387,7 @@ def fullHelpMessage() { --dbdir Path to Directory where database is being stored - --headers Set taxonomy database header format -> headers= "RefSeq" to toggle use of RefSeq header format; set to "RVDB" to signal the use of Reverence Viral DataBase (RVDB) headers + --headers Set taxonomy database header format -> headers= "NCBI" to toggle use of NCBI header format; set to "RVDB" to signal the use of Reverence Viral DataBase (RVDB) headers --dbanno Path to directory hmm annotation .txt file - see manual for information on this. Leave as is if not planning on using. @@ -1350,7 +1350,7 @@ if (params.DataCheck || params.Analyze) { if (params.dbtype == "NCBI") { - process ncASV_Taxonomy_Inference_RefSeq { /////// editttt + process ncASV_Taxonomy_Inference_NCBI { /////// editttt label 'high_cpus' @@ -1737,7 +1737,7 @@ if (params.DataCheck || params.Analyze) { if (params.dbtype == "NCBI") { - process ASV_Taxonomy_Inference_RefSeq { /////// editttt + process ASV_Taxonomy_Inference_NCBI { /////// editttt label 'high_cpus' @@ -2153,7 +2153,13 @@ if (params.DataCheck || params.Analyze) { #entopy analysis entropy-analysis ${params.projtag}_ASVs_Aligned_informativeonly.fasta #Decomposition - oligotype ${params.projtag}_ASVs_Aligned_informativeonly.fasta ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY -o ${params.projtag}_asvMED_${params.asvC} -M 1 -c ${params.asvC} -N ${task.cpus} --skip-check-input --no-figures --skip-gen-html + if [[ \$(echo ${params.asvC} | grep -c ",") -eq 1 ]] + then + tag=$(echo ${params.asvC} | sed 's/,/_/g') + oligotype ${params.projtag}_ASVs_Aligned_informativeonly.fasta ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY -o ${params.projtag}_asvMED_"\$tag -M 1 -C ${params.asvC} -N ${task.cpus} --skip-check-input --no-figures --skip-gen-html + else + oligotype ${params.projtag}_ASVs_Aligned_informativeonly.fasta ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY -o ${params.projtag}_asvMED_${params.asvC} -M 1 -c ${params.asvC} -N ${task.cpus} --skip-check-input --no-figures --skip-gen-html + fi #generatemaps cd ./${params.projtag}_asvMED_${params.asvC}/OLIGO-REPRESENTATIVES/ echo "ASV,GroupID,IDPattern" @@ -2411,7 +2417,7 @@ if (params.DataCheck || params.Analyze) { if (params.dbtype == "NCBI") { - process AminoType_Taxonomy_Inference_RefSeq { + process AminoType_Taxonomy_Inference_NCBI { label 'high_cpus' @@ -2808,7 +2814,13 @@ if (params.DataCheck || params.Analyze) { #entopy analysis entropy-analysis ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta #Decomposition - oligotype ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta-ENTROPY -o ${params.projtag}_AminoTypeMED_${params.aminoC} -M 1 -c ${params.aminoC} -N ${task.cpus} --skip-check-input --no-figures --skip-gen-html + if [[ \$(echo ${params.aminoC} | grep -c ",") -eq 1 ]] + then + tag=$(echo ${params.aminoC} | sed 's/,/_/g') + oligotype ${params.projtag}_ASVs_Aligned_informativeonly.fasta ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY -o ${params.projtag}_asvMED_"\$tag -M 1 -C ${params.aminoC} -N ${task.cpus} --skip-check-input --no-figures --skip-gen-html + else + oligotype ${params.projtag}_ASVs_Aligned_informativeonly.fasta ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY -o ${params.projtag}_asvMED_${params.aminoC} -M 1 -c ${params.aminoC} -N ${task.cpus} --skip-check-input --no-figures --skip-gen-html + fi #generatemaps cd ./${params.projtag}_AminoTypeMED_${params.aminoC}/OLIGO-REPRESENTATIVES/ echo "AminoType,Group,IDPattern" @@ -3036,7 +3048,7 @@ if (params.DataCheck || params.Analyze) { if (params.dbtype == "NCBI") { - process pcASV_Nucleotide_Taxonomy_Inference_RefSeq { + process pcASV_Nucleotide_Taxonomy_Inference_NCBI { label 'high_cpus' @@ -3507,7 +3519,7 @@ if (params.DataCheck || params.Analyze) { if (params.dbtype == "NCBI") { - process pcASV_AminoAcid_Taxonomy_Inference_RefSeq { + process pcASV_AminoAcid_Taxonomy_Inference_NCBI { label 'high_cpus' diff --git a/vampirus.config b/vampirus.config index de3df9d..4e99635 100644 --- a/vampirus.config +++ b/vampirus.config @@ -74,7 +74,7 @@ params { // Minimum Entropy Decomposition (MED) parameters for clustering (https://merenlab.org/2012/05/11/oligotyping-pipeline-explained/) - // If you plan to do MED on ASVs using the option "--asvMED" you can set here the number of positions for oligotyping to take into consideration + // If you plan to do MED on ASVs using the option "--asvMED" you can set here the number of entopy peak positions or a comma seperated list of biologically meaningful positons (e.g. 35,122,21) for oligotyping to take into consideration asvC="" // If you plan to do MED on ASVs using the option "--aminoMED" you can set here the number of positions for oligotyping to take into consideration aminoC="" @@ -112,7 +112,7 @@ params { dbtype="TYPE" // Classification settings - if planning on inferring LCA from RVDB annotation files OR using NCBI taxonomy files, confirm options below are accurate. - // Path to directory RVDB hmm annotation .txt file - see manual for information on this. Leave as is if not planning on using. + // Path to directory RVDB hmm annotation .txt file - see manual for information on this. Leave as is if not planning on using RVDB LCA. dbanno="DATABASEANNOT" // Set lca="T" if you would like to add "Least Common Ancestor" classifications to taxonomy results using information provided by RVDB annotation files (works when using NCBI or RVDB databases) - example: "ASV1, Viruses::Duplodnaviria::Heunggongvirae::Peploviricota::Herviviricetes::Herpesvirales::Herpesviridae::Gammaherpesvirinae::Macavirus" lca="LCAT" From bc6efd549fbdd74aa395b7d6ac2308799542b5fd Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Tue, 3 Aug 2021 11:16:39 -0500 Subject: [PATCH 119/223] update --- docs/HelpDocumentation.md | 15 +++++++++++++-- vampirus_env.yml | 1 - vampirus_startup.sh | 1 + 3 files changed, 14 insertions(+), 3 deletions(-) diff --git a/docs/HelpDocumentation.md b/docs/HelpDocumentation.md index bcaa271..ab03987 100644 --- a/docs/HelpDocumentation.md +++ b/docs/HelpDocumentation.md @@ -44,13 +44,22 @@ The vAMPirus program contains two different pipelines: If you have a feature request or any feedback/questions, feel free to email vAMPirusHelp@gmail.com or you can open an Issue on GitHub. +## Changes in version 2.0.0 + +1. Added Minimum Entropy Decomposition analysis using the oligotyping program produced by the Meren Lab. This allows for sequence clustering based on sequence positions of interest (biologically meaningful) or top positions with the highest Shannon's Entropy (read more here: https://merenlab.org/software/oligotyping/ ; and below) + +2. Added more useful taxonomic classification of sequences leveraging the RVDB annotation database and/or NCBI taxonomy files (read more below) + +3. Replaced the used of MAFFT with muscle5 (Edgar 2021) for more accurate virus gene alignments (see https://www.biorxiv.org/content/10.1101/2021.06.20.449169v1.full) + + ## Who to cite If you do use vAMPirus for your analyses, please cite the following -> 1. vAMPirus - Veglia, A.J., Rivera Vicens, R., Grupstra, C., Howe-Kerr, L., and Correa A.M.S. (2020) vAMPirus: An automated virus amplicon sequencing analysis pipeline. Zenodo. *DOI:* -2. Diamond - Buchfink B, Xie C, Huson DH. (2015) Fast and sensitive protein alignment using DIAMOND. Nat Methods. 12(1):59-60. doi:10.1038/nmeth.3176 +2. DIAMOND - Buchfink B, Xie C, Huson DH. (2015) Fast and sensitive protein alignment using DIAMOND. Nat Methods. 12(1):59-60. doi:10.1038/nmeth.3176 3. FastQC - Andrews, S. (2010). FastQC: A Quality Control Tool for High Throughput Sequence Data [Online]. Available online at: http://www.bioinformatics.babraham.ac.uk/projects/fastqc/ @@ -62,7 +71,7 @@ If you do use vAMPirus for your analyses, please cite the following -> 7. ModelTest-NG - Darriba, D., Posada, D., Kozlov, A. M., Stamatakis, A., Morel, B., & Flouri, T. (2020). ModelTest-NG: a new and scalable tool for the selection of DNA and protein evolutionary models. Molecular biology and evolution, 37(1), 291-294. -8. MAFFT - Katoh, K., & Standley, D. M. (2013). MAFFT multiple sequence alignment software version 7: improvements in performance and usability. Molecular biology and evolution, 30(4), 772-780. +8. muscle v5 - R.C. Edgar (2021) "MUSCLE v5 enables improved estimates of phylogenetic tree confidence by ensemble bootstrapping" https://www.biorxiv.org/content/10.1101/2021.06.20.449169v1.full.pdf 9. vsearch - Rognes, T., Flouri, T., Nichols, B., Quince, C., & Mahé, F. (2016). VSEARCH: a versatile open source tool for metagenomics. PeerJ, 4, e2584. @@ -78,6 +87,8 @@ If you do use vAMPirus for your analyses, please cite the following -> 15. UNOISE algorithm - R.C. Edgar (2016). UNOISE2: improved error-correction for Illumina 16S and ITS amplicon sequencing, https://doi.org/10.1101/081257 +16. Oligotyping - A. Murat Eren, Gary G. Borisy, Susan M. Huse, Jessica L. Mark Welch (2014). Oligotyping analysis of the human oral microbiome. Proceedings of the National Academy of Sciences Jul 2014, 111 (28) E2875-E2884; DOI: 10.1073/pnas.1409644111 + # Getting started with vAMPirus diff --git a/vampirus_env.yml b/vampirus_env.yml index 0957c96..cb10503 100644 --- a/vampirus_env.yml +++ b/vampirus_env.yml @@ -10,7 +10,6 @@ channels: dependencies: - python=3.6 - blast=2.11.0 - - blast-legacy=2.2.26 - diamond=2.0.11 - fastqc=0.11.9 - fastp=0.20.1 diff --git a/vampirus_startup.sh b/vampirus_startup.sh index 7730296..28b00f5 100644 --- a/vampirus_startup.sh +++ b/vampirus_startup.sh @@ -270,6 +270,7 @@ if [[ "$TAX" == "yes" ]] then mkdir "$mypwd"/Databases/NCBItaxonomy cd "$mypwd"/Databases/NCBItaxonomy curl -o prot.accession2taxid.FULL.gz ftp://ftp.ncbi.nlm.nih.gov/pub/taxonomy/accession2taxid/prot.accession2taxid.FULL.gz + echo "Gunzipping accession2tax map, might take a moment.." gunzip prot.accession2taxid.FULL.gz curl -o taxdmp.zip ftp://ftp.ncbi.nlm.nih.gov/pub/taxonomy/taxdmp.zip unzip taxdmp.zip From 7500d1b8003b996d2ddc7f109b2339ee8b46a653 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Tue, 3 Aug 2021 11:44:28 -0500 Subject: [PATCH 120/223] fixed --- vAMPirus.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index ee40f57..8bec68a 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1,4 +1,4 @@ -muscle#!/usr/bin/env nextflow +#!/usr/bin/env nextflow /* ======================================================================================== From 1154871171b21ad55489e2ad942e9040968b0319 Mon Sep 17 00:00:00 2001 From: Ramon Rivera Date: Tue, 3 Aug 2021 18:58:16 +0200 Subject: [PATCH 121/223] Update vAMPirus_Report.Rmd --- bin/vAMPirus_Report.Rmd | 675 +++++++++++++++++++++++++++++++++++++++- 1 file changed, 667 insertions(+), 8 deletions(-) diff --git a/bin/vAMPirus_Report.Rmd b/bin/vAMPirus_Report.Rmd index af10a49..9a86013 100644 --- a/bin/vAMPirus_Report.Rmd +++ b/bin/vAMPirus_Report.Rmd @@ -14,6 +14,9 @@ params: stats: !r commandArgs(trailingOnly=T)[8] metadata: !r commandArgs(trailingOnly=T)[9] minimumCounts: !r commandArgs(trailingOnly=T)[10] + asvMED: !r commandArgs(trailingOnly=T)[11] + aminoMED: !r commandArgs(trailingOnly=T)[12] + type: !r commandArgs(trailingOnly=T)[13] --- @@ -71,7 +71,7 @@ if (params$skipReadProcessing == "true" || params$skipMerging == "true" ) { reads_stats=read.csv("final_reads_stats.csv") #paged_table(reads_stats,options = list(rows.print = 20)) knitr::kable(reads_stats, digits = 2, align = 'c', booktabs = TRUE, caption = "Table 1: Read summary stats") %>% - kable_styling(font_size = 12, full_width = F)%>% + kable_styling(font_size = 12, full_width = F)%>% scroll_box(width = "100%", height = "100%") } else { writeLines("\n--------------------------------------------------------------\n") @@ -764,7 +764,7 @@ if (params$asvMED == "true" && params$type == "ASV"){ #paged_table(med_group,options = list(rows.print = 20)) med_group=read.csv("asv_medtable.csv", header = T) knitr::kable(med_group, digits = 2, align = 'c', booktabs = TRUE, caption = "Table: ASV MED Table") %>% - kable_styling(font_size = 12, full_width = F)%>% + kable_styling(font_size = 12, full_width = F)%>% scroll_box(width = "100%", height = "100%") } @@ -774,7 +774,7 @@ if (params$aminoMED == "true" && params$type == "AminoType"){ #paged_table(med_group,options = list(rows.print = 20)) med_group=read.csv("amino_medtable.csv", header = T) knitr::kable(med_group, digits = 2, align = 'c', booktabs = TRUE, caption = "Table: Aminotype MED Table") %>% - kable_styling(font_size = 12, full_width = F) %>% + kable_styling(font_size = 12, full_width = F) %>% scroll_box(width = "100%", height = "100%") } ``` From 575505f6b647d96b3026e27bc3ecbda2dbfcd203 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Fri, 10 Sep 2021 10:39:48 -0500 Subject: [PATCH 172/223] edits --- docs/HelpDocumentation.md | 2 +- example_data/conf/test.config | 1 + 2 files changed, 2 insertions(+), 1 deletion(-) diff --git a/docs/HelpDocumentation.md b/docs/HelpDocumentation.md index 036dc66..3e91cfe 100644 --- a/docs/HelpDocumentation.md +++ b/docs/HelpDocumentation.md @@ -1103,7 +1103,7 @@ Example launch command: In vAMPirus v2, we added the ability for the user to use the oligotyping program employing the Minimum Entropy Decomposition (MED) algorithm developed by Eren et al. 2015 (read more about MED here - https://www.nature.com/articles/ismej2014195#citeas) to cluster ASV or AminoType sequences. -The MED algorithm provides an alternative way of clustering marker gene sequences using "information theory-guided decomposition" - "By employing Shannon entropy, MED uses only the information-rich nucleotide positions across reads and iteratively partitions large datasets while omitting stochastic variation." -Ereb et al. 2015 +The MED algorithm provides an alternative way of clustering marker gene sequences using "information theory-guided decomposition" - "By employing Shannon entropy, MED uses only the information-rich nucleotide positions across reads and iteratively partitions large datasets while omitting stochastic variation." -Eren et al. 2015 When you run the DataCheck pipeline with your dataset, the report will include a figure and table that breakdown the Shannon Entropy analysis results for both ASVs and AminoTypes. The figure visualizes entropy values per sequence position revealing positions or regions of high entropy. The table beneath the figure breaks down the number of positions with entropy values above "0.x". Generally, you would want to decompose sequences using all high extropy positions (>0.1). Although, if you know the positions on your sequence that have the potential to contain biologically or ecologically meaningful mutations, you can specify decomposition based on these positions. diff --git a/example_data/conf/test.config b/example_data/conf/test.config index b68646b..c75ce75 100644 --- a/example_data/conf/test.config +++ b/example_data/conf/test.config @@ -18,5 +18,6 @@ params { clusterAAID="85" dbname="test_db.fasta" dbdir="${projectDir}/example_data/conf/" + dbtype="RVDB" stats = true } From d3dd6bf2acf9a7d2ffa436a6213905c4581f9ac1 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Fri, 10 Sep 2021 11:51:48 -0500 Subject: [PATCH 173/223] added info in help docs --- docs/HelpDocumentation.md | 60 +- .../ex_reports/example_Analyze_Report.html | 882 ------------------ .../ex_reports/example_DataCheck_Report.html | 391 -------- example_data/conf/test.config | 2 + 4 files changed, 33 insertions(+), 1302 deletions(-) delete mode 100644 example_data/conf/ex_reports/example_Analyze_Report.html delete mode 100644 example_data/conf/ex_reports/example_DataCheck_Report.html diff --git a/docs/HelpDocumentation.md b/docs/HelpDocumentation.md index 3e91cfe..35fcf4a 100644 --- a/docs/HelpDocumentation.md +++ b/docs/HelpDocumentation.md @@ -537,7 +537,7 @@ Analyze test => OR - nextflow run vAMPirus.nf -c vampirus.config -profile singularity,test --Analyze --ncASV --pcASV --stats + nextflow run vAMPirus.nf -c /path/to/vampirus.config -profile singularity,test --Analyze --ncASV --pcASV --asvMED --aminoMED --stats ### Resuming test analyses if you ran into an error @@ -1105,7 +1105,7 @@ In vAMPirus v2, we added the ability for the user to use the oligotyping program The MED algorithm provides an alternative way of clustering marker gene sequences using "information theory-guided decomposition" - "By employing Shannon entropy, MED uses only the information-rich nucleotide positions across reads and iteratively partitions large datasets while omitting stochastic variation." -Eren et al. 2015 -When you run the DataCheck pipeline with your dataset, the report will include a figure and table that breakdown the Shannon Entropy analysis results for both ASVs and AminoTypes. The figure visualizes entropy values per sequence position revealing positions or regions of high entropy. The table beneath the figure breaks down the number of positions with entropy values above "0.x". Generally, you would want to decompose sequences using all high extropy positions (>0.1). Although, if you know the positions on your sequence that have the potential to contain biologically or ecologically meaningful mutations, you can specify decomposition based on these positions. +When you run the DataCheck pipeline with your dataset, the report will include a figure and table that breakdown the Shannon Entropy analysis results for both ASVs and AminoTypes. The figure visualizes entropy values per sequence position revealing positions or regions of high entropy. The table beneath the figure breaks down the number of positions with entropy values above "0.x". Although, if you know the positions on your sequence that have the potential to contain biologically or ecologically meaningful mutations, you can specify decomposition based on these positions. If you decide to use MED, vAMPirus will run all the same analyses that would be done with the ASV or AminoType sequences (e.g. diversity analyses, statistics) and be appended results to the ASV or AminoType report. The ASV or AminoType sequence nodes on the phylogenetic tree will also be colored based on which MED group they were assigned to. @@ -1188,7 +1188,7 @@ By default IQTREE will determine the best model to use with ModelFinder Plus. ### Bootstrapping -IQ-TREE is capable of performing parametric or non-parametric bootstrapping. You can specify which one using "--parametric" or "--nonparametric" and to set how many boostraps to perform, you would use "--boots #ofbootstraps" or edit lime 114 in the vampirus.config file. +IQ-TREE is capable of performing parametric or non-parametric bootstrapping. You can specify which one using "--parametric" or "--nonparametric" and to set how many bootstraps to perform, you would use "--boots #ofbootstraps" or edit lime 114 in the vampirus.config file. Here is an example for creating a tree using the model determined by ModelTest-NG, non-parametric bootstrapping and 500 bootstraps: @@ -1236,7 +1236,7 @@ AminoType IQTREE command -> ## Taxonomy Inference -vAMPirus uses DIAMOND blastx/blastp and the provided protein database to infer taxonomy of amplicons to ASVs/cASVs/AminoTypes. There are summary files generated, one in the formated to process with phyloseq and the other as a .tsv with information in a different arrangement. Results are also visualized as a donut graph in the final reports. +vAMPirus uses DIAMOND blastx/blastp and the provided protein database to infer taxonomy of amplicons to ASVs/cASVs/AminoTypes. There are summary files generated, one in the format compatable with phyloseq and the other as a .tsv with information in a different arrangement. Results are also visualized as a donut graph in the final reports. First, lets take a look at the taxonomy section of the configuration file: @@ -1328,17 +1328,17 @@ NOTE=> Be sure that there is more than 1 sample in each treatment category or th There are several files created throughout the vAMPirus pipeline that are stored and organized in directories within the specified results/output directory (ex. ${working_directory}/results; lime 27 in the configuration file). We will go through the structure of the output directory and where to find which files here: -## Pipeline performance information - ${working_directory}/results/PipelinePerformance/ +## Pipeline performance information - ${working_directory}/${outdir}/PipelinePerformance/ Nextflow produces a couple files that breakdown how and what parts of the vAMPirus pipeline was ran. The first file is a report that contains information on how the pipeline performed along with other pipeline performance-related information like how much memory or CPUs were used during certain processes. You can use this information to alter how many resources you would request for a given task in the pipeline. For example, the counts table generation process may take a long time with the current amount of resources requested, you can see this in the report then edit the resources requested at lines 144-183 in the vAMPirus configuration file. The second file produced by Nextflow is just the visualization of the workflow and analyses ran as a flowchart. -## Output of "--DataCheck" - ${working_directory}/results/DataCheck +## Output of "--DataCheck" - ${working_directory}/${outdir}/DataCheck The DataCheck performed by vAMPirus includes "ReadProcessing", "Clustering", and "Report" generation. Here again is the launch command to run the DataCheck mode: `nextflow run vAMPirus.nf -c vampirus.config -profile [conda|singularity] --DataCheck` -### ReadProcessing - ${working_directory}/results/DataCheck/ReadProcessing +### ReadProcessing - ${working_directory}/${outdir}/ReadProcessing Within the ReadProcessing directory you will find all files related to each step of read processing: @@ -1358,7 +1358,7 @@ Similar to the adapter removal directory, here you have the clean read libraries There is a little bit more going on in this directory compared to the others. The first major file to pay attention to here is the file \*\_merged_clean_Lengthfiltered.fastq. This is the "final" merged read file that contains all merged reads from each samples and is used to identify unique sequences and then ASVs. "Pre-filtered" and "pre-cleaned" combined merged read files can be found in "./LengthFiltering". If you would like to review or use the separate merged read files per sample, these fastq files are found in the "./Individual" directory. Finally, a fasta file with unique sequences are found in the "./Uniques" directory and the "./Histograms" directory is full of several different sequence property (length, per base quality, etc.) histogram files which can be visualized manually and reviewed in the DataCheck report. -### Clustering - ${working_directory}/results/DataCheck/Clustering +### Clustering - ${working_directory}/${outdir}/DataCheck/Clustering As the name would suggest, the files within this directory are related to the clustering process of "--DataCheck". There isn't too much in here, but here is the breakdown anyway: @@ -1368,13 +1368,13 @@ In this directory, there is the fasta file with the generated ASV sequences and 2. Nucleotide - ${working_directory}/results/DataCheck/Clustering/Nucleotide -This directory stores a .csv file that shows the number of clusters or ncASVs per clustering percentage. The file can be visualized manually or can be reviewed in the DataCheck report. +This directory stores a .csv file that shows the number of clusters or ncASVs per clustering percentage. The file can be visualized manually or can be reviewed in the DataCheck report. In this directory you will also find Shannon Entropy analysis results files. 3. Aminoacid - ${working_directory}/results/DataCheck/Clustering/Aminoacid -Similar to Nucleotide, the Aminoacid directory contained the .csv that shows the number of clusters or pcASVs per clustering percentage. The file can be visualized manually or can be reviewed in the DataCheck report. +Similar to Nucleotide, the Aminoacid directory contained the .csv that shows the number of clusters or pcASVs per clustering percentage. The file can be visualized manually or can be reviewed in the DataCheck report. In this directory you will also find Shannon Entropy analysis results files. -### Report - ${working_directory}/results/DataCheck/Report +### Report - ${working_directory}/${outdir}/DataCheck/Report In this directory, you will find a .html DataCheck report that can be opened in any browser. The report contains the following information and it meant to allow the user to tailor their vAMPirus pipeline run to their data (i.e. maximum read length, clustering percentage, etc.): @@ -1393,15 +1393,15 @@ In this section of the report, vAMPirus is showing the number of nucleotide- and NOTE: Most, if not all, plots in vAMPirus reports are interactive meaning you can select and zoom on certain parts of the plot or you can use the legend to remove certain samples. -## Output of "--Analyze" - ${working_directory}/results/Analyze +## Output of "--Analyze" - ${working_directory}/${outdir}/Analyze Depending on which optional arguments you add to your analyses (e.g. --pcASV, --ncASV, skip options), you will have different files produced, here we will go through the output of the full analysis stemming from this launch command: nextflow run vAMPirus.nf -c vampirus.config -profile [conda|singularity] --Analyze --ncASV --pcASV --stats -### ReadProcessing - ${working_directory}/results/Analyze/ReadProcessing +### ReadProcessing - ${working_directory}/${outdir}/ReadProcessing -Very similar to the "ReadProcessing" directory created in DataCheck, you will find the following: +Contains the same files produced during DataCheck run (analyses will not be redone) 1. FastQC - ${working_directory}/results/Analyze/ReadProcessing/FastQC @@ -1419,33 +1419,33 @@ Similar to the adapter removal directory, here you have the clean read libraries There is a little bit more going on in this directory compared to the others. The first major file to pay attention to here is the file \*\_merged_clean_Lengthfiltered.fastq. This is the "final" merged read file that contains all merged reads from each samples and is used to identify unique sequences and then ASVs. "Pre-filtered" and "pre-cleaned" combined merged read files can be found in "./LengthFiltering". If you would like to review or use the separate merged read files per sample, these fastq files are found in the "./Individual" directory. Finally, a fasta file with unique sequences are found in the "./Uniques" directory and the "./Histograms" directory is full of several different sequence property (length, per base quality, etc.) histogram files which can be visualized manually and reviewed in the DataCheck report if ran before Analyze. -### Clustering - ${working_directory}/results/Analyze/Clustering +### Clustering - ${working_directory}/${outdir}/Analyze/Clustering The clustering directory will contain all files produced for whichever clustering technique you specified (with the launch command above, all are specified): -1. ASVs - ${working_directory}/results/Analyze/Clustering/ASVsIn this directory, there is the fasta file with the generated ASV sequences and there is another directory "./ChimeraCheck" where the pre-chimera filered ASV fasta sits. +1. ASVs -- ${working_directory}/results/Analyze/Clustering/ASVs -- In this directory, there is the fasta file with the generated ASV sequences and there is another directory "./ChimeraCheck" where the pre-chimera filered ASV fasta sits. In this directory, if --asvMED was set to run, you will be a MED/ directory containing all output files from oligotyping analyses. -2. AminoTypes - ${working_directory}/results/Analyze/Clustering/AminoTypesThe AminoTypes directory has a few different subdirectories, in the main directory, however, is the fasta file with the AminoTypes used in all subsequent analyses. The first subdirectory is called "Translation" which includes the raw ASV translation file along with a report spit out by VirtualRibosome. The next subdirectory is "Problematic", where any translations that were below the given "--minAA" length will be reported, if none were deemed "problematic" then the directory will be empty. All problematic amino acid sequence AND their corresponding ASVs are stored in fasta files for you to review. The final subdirectory is "SummaryFiles" where you can find a "map" of sorts to track which ASVs contributed to which AminoTypes and a .gc file containing information on length of translated sequences. +2. AminoTypes -- ${working_directory}/results/Analyze/Clustering/AminoTypes -- The AminoTypes directory has a few different subdirectories, in the main directory, however, is the fasta file with the AminoTypes used in all subsequent analyses. The first subdirectory is called "Translation" which includes the raw ASV translation file along with a report spit out by VirtualRibosome. The next subdirectory is "Problematic", where any translations that were below the given "--minAA" length will be reported, if none were deemed "problematic" then the directory will be empty. All problematic amino acid sequence AND their corresponding ASVs are stored in fasta files for you to review. The final subdirectory is "SummaryFiles" where you can find a "map" of sorts to track which ASVs contributed to which AminoTypes and a .gc file containing information on length of translated sequences. In this directory, if --aminoMED was set to run, you will be a MED/ directory containing all output files from oligotyping analyses. -3. ncASV - ${working_directory}/results/Analyze/Clustering/ncASVIn this directory, you will find the fasta files corresponding to the clustering percentage(s) you specified for the run. +3. ncASV -- ${working_directory}/results/Analyze/Clustering/ncASV -- In this directory, you will find the fasta files corresponding to the clustering percentage(s) you specified for the run. -4. pcASV - ${working_directory}/results/Analyze/Clustering/pcASVLooking in this directory, you probably notice some similar subdirectories. The pcASV directory also contains the Summary, Problematic, and Translation subsirectories we saw in the AminoType directory. The other important files in this directory is the nucleotide and amino acid versions of the pcASVs generated for whichever clustering percentage(s) specified.An important note for when creating pcASVs is that the subsequent analyses (phylogenies, taxonomy assignment, etc.) are run on both nucloetide and amino acid pcASV fastas. To create these files, vAMPirus translates the ASVs, checks for problematic sequences, then clusters the translated sequences by the given percentage(s). After clustering, vAMPirus will go pcASV by pcASV extracting the nucleotide sequences of the ASVs that clustered within a given pcASV. The extracted nucleotide sequences are then used to generate a consensus nucleotide sequence(s) per pcASV. +4. pcASV -- ${working_directory}/results/Analyze/Clustering/pcASV -- Looking in this directory, you probably notice some similar subdirectories. The pcASV directory also contains the Summary, Problematic, and Translation subsirectories we saw in the AminoType directory. The other important files in this directory is the nucleotide and amino acid versions of the pcASVs generated for whichever clustering percentage(s) specified.An important note for when creating pcASVs is that the subsequent analyses (phylogenies, taxonomy assignment, etc.) are run on both nucloetide and amino acid pcASV fastas. To create these files, vAMPirus translates the ASVs, checks for problematic sequences, then clusters the translated sequences by the given percentage(s). After clustering, vAMPirus will go pcASV by pcASV extracting the nucleotide sequences of the ASVs that clustered within a given pcASV. The extracted nucleotide sequences are then used to generate a consensus nucleotide sequence(s) per pcASV. -### Analyses - ${working_directory}/results/Analyze/Analyses +### Analyses - ${working_directory}/${outdir}/Analyze/Analyses For each clustering technique (i.e. ASVs, AminoTypes, ncASVs and pcASVs) performed in a given run, resulting taxonomic unit fastas will go through the following analyses (unless skip options are used): -1. Counts - ${working_directory}/results/Analyze/Analyses/${clustertechnique}/CountsThe Counts directory is where you can find the counts tables as .csv files (and .biome as well for nucleotide counts tables). +1. Counts -- ${working_directory}/results/Analyze/Analyses/${clustertechnique}/Counts -- The Counts directory is where you can find the counts tables as .csv files (and .biome as well for nucleotide counts tables). -2. Phylogeny - ${working_directory}/results/Analyze/Analyses/${clustertechnique}/PhylogenyUnless told otherwise, vAMPirus will produce phylogenetic trees for all taxonomic unit fastas using IQ-TREE. The options for this analysis was discussed in a previous section of the docs. In the phylogeny output directory, you will find three subdirectories: (i) ./Alignment - contains trimmed MAFFT alignment used for tree, (ii) ./ModelTest - contains output files from substitution model prediction with ModelTest-NG, and (iii) ./IQ-TREE - where you can find all output files from IQ-TREE with the file of (usual) interest is the ".treefile". +2. Phylogeny -- ${working_directory}/results/Analyze/Analyses/${clustertechnique}/Phylogeny -- Unless told otherwise, vAMPirus will produce phylogenetic trees for all taxonomic unit fastas using IQ-TREE. The options for this analysis was discussed in a previous section of the docs. In the phylogeny output directory, you will find three subdirectories: (i) ./Alignment - contains trimmed MAFFT alignment used for tree, (ii) ./ModelTest - contains output files from substitution model prediction with ModelTest-NG, and (iii) ./IQ-TREE - where you can find all output files from IQ-TREE with the file of (usual) interest is the ".treefile". -3. Taxonomy - ${working_directory}/results/Analyze/Analyses/${clustertechnique}/TaxonomyvAMPirus uses DIAMOND blastp/x and the supplied PROTEIN database for taxonomy assignment of sequences. In the Taxonomy directory, you will find (i) a subdirectory called "DIAMONDOutput" which contains the original output file produced by DIAMOND, (ii) a fasta file that has taxonomy assignments within the sequence headers, and (iii) three different summary files (one being a phyloseq object with taxonomic information, a tab-separated summary file for review by the user and a summary table looking at abundance of specific hits). +3. Taxonomy -- ${working_directory}/results/Analyze/Analyses/${clustertechnique}/Taxonomy -- vAMPirus uses DIAMOND blastp/x and the supplied PROTEIN database for taxonomy assignment of sequences. In the Taxonomy directory, you will find (i) a subdirectory called "DIAMONDOutput" which contains the original output file produced by DIAMOND, (ii) a fasta file that has taxonomy assignments within the sequence headers, and (iii) three different summary files (one being a phyloseq object with taxonomic information, a tab-separated summary file for review by the user and a summary table looking at abundance of specific hits). -4. Matrix - ${working_directory}/results/Analyze/Analyses/${clustertechnique}/MatrixThe Matric directory is where you can find all Percent Identity matrices for produced ASV/cASV/AmintoType fastas. +4. Matrix -- ${working_directory}/results/Analyze/Analyses/${clustertechnique}/Matrix -- The Matric directory is where you can find all Percent Identity matrices for produced ASV/cASV/AmintoType fastas. -5. EMBOSS - ${working_directory}/results/Analyze/Analyses/${clustertechnique}/EMBOSSSeveral different protein physiochemical properties for all amino acid sequences are assessed using EMBOSS scripts (http://emboss.sourceforge.net/apps/release/6.6/emboss/apps/groups.html). There are four different subdirectories within EMBOSS, these include (i) ./ProteinProperties - contains files and plots regarding multiple different physiochemical properties (http://emboss.sourceforge.net/apps/release/6.6/emboss/apps/pepstats.html), (ii) ./IsoelectricPoint - contains a text file and a .svg image with plots showing the isoelectric point of protein (http://emboss.sourceforge.net/apps/release/6.6/emboss/apps/iep.html), (iii) ./HydrophobicMoment - information related to hydrophobic moments of amino acid sequences (http://emboss.sourceforge.net/apps/release/6.6/emboss/apps/hmoment.html), and (iv) ./2dStructure - information about 2D structure of proteins (http://emboss.sourceforge.net/apps/release/6.6/emboss/apps/protein_2d_structure_group.html). +5. EMBOSS -- ${working_directory}/results/Analyze/Analyses/${clustertechnique}/EMBOSS -- Several different protein physiochemical properties for all amino acid sequences are assessed using EMBOSS scripts (http://emboss.sourceforge.net/apps/release/6.6/emboss/apps/groups.html). There are four different subdirectories within EMBOSS, these include (i) ./ProteinProperties - contains files and plots regarding multiple different physiochemical properties (http://emboss.sourceforge.net/apps/release/6.6/emboss/apps/pepstats.html), (ii) ./IsoelectricPoint - contains a text file and a .svg image with plots showing the isoelectric point of protein (http://emboss.sourceforge.net/apps/release/6.6/emboss/apps/iep.html), (iii) ./HydrophobicMoment - information related to hydrophobic moments of amino acid sequences (http://emboss.sourceforge.net/apps/release/6.6/emboss/apps/hmoment.html), and (iv) ./2dStructure - information about 2D structure of proteins (http://emboss.sourceforge.net/apps/release/6.6/emboss/apps/protein_2d_structure_group.html). -### FinalReport - ${working_directory}/results/Analyze/FinalReport +### FinalReport - ${working_directory}/${outdir}/Analyze/FinalReports vAMPirus produces final reports for all taxonomic unit fastas produced in the run. These reports contain the following information: @@ -1461,7 +1461,7 @@ This is a plot that looks at number of reads per sample, similar to what is seen 4. Diversity analyses box Plots -The plots in order are (i) Shannon Diversity, (ii) Simpson Diversity, (iii) Species Richness. +The plots in order are (i) Shannon Diversity, (ii) Simpson Diversity, (iii) Richness. Stats tests included with "--stats": @@ -1478,9 +1478,9 @@ Stats tests included with "--stats": 6. NMDS plots (2D and 3D) -7. Relative ASV/cASV abundance per sample bar chart +7. Relative sequence abundance per sample bar chart -8. Absolute ASV/cASV abundance per treatment bar chart +8. Absolute sequence abundance per treatment bar chart 9. Pairwise percent ID heatmap @@ -1488,6 +1488,8 @@ Stats tests included with "--stats": 11. Visualized phylogenetic tree +12. Post-Minimum Entropy Decomposition Analyses (combination of above) + # All of the options diff --git a/example_data/conf/ex_reports/example_Analyze_Report.html b/example_data/conf/ex_reports/example_Analyze_Report.html deleted file mode 100644 index bc65412..0000000 --- a/example_data/conf/ex_reports/example_Analyze_Report.html +++ /dev/null @@ -1,882 +0,0 @@ - - - - - - - - - - - - - -vAMPirus Analyze Report vAMPtest_ASVs - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - -

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    NOTE: Most plots are interactive and you can use the legend to specify samples/treatment of interest. You can also download an .svg version of each figure within this report.

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    ## [1] "Shapiro Test of normality - data is normal p-value > 0.05"
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-##  Shapiro-Wilk normality test
-##
-## data:  resid(shannonaov)
-## W = 0.9291, p-value = 0.5732
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-##
-## --------------------------------------------------------------
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-## [1] "Bartlett Test variance homogeneity - variance is homogeneous p-value > 0.05"
-##
-##  Bartlett test of homogeneity of variances
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-## data:  index by treatment
-## Bartlett's K-squared = 1.0859, df = 1, p-value = 0.2974
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-## --------------------------------------------------------------
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-## [1] "ANOVA Results"
-##             Df Sum Sq Mean Sq F value Pr(>F)
-## treatment    1 0.0527  0.0527    0.15  0.718
-## Residuals    4 1.4061  0.3515
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-## --------------------------------------------------------------
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-## [1] "Tukey HSD - Pairwise comparison - significant differences indicated by p-value < 0.05"
-##   Tukey multiple comparisons of means
-##     95% family-wise confidence level
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-## Fit: aov(formula = index ~ treatment, data = shannondata5_2)
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-##                     diff       lwr     upr     p adj
-## Group2-Group1 -0.1874213 -1.531512 1.15667 0.7183604
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    ## [1] "Shapiro Test of normality - data is normal p-value > 0.05"
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-##  Shapiro-Wilk normality test
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-## data:  resid(simpsonaov)
-## W = 0.88738, p-value = 0.3047
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-## [1] "Bartlett Test variance homogeneity - variance is homogeneous p-value > 0.05"
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-##  Bartlett test of homogeneity of variances
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-## data:  index by treatment
-## Bartlett's K-squared = 0.74903, df = 1, p-value = 0.3868
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-## [1] "ANOVA Results"
-##             Df Sum Sq Mean Sq F value Pr(>F)
-## treatment    1 0.0052 0.00519   0.059   0.82
-## Residuals    4 0.3506 0.08764
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-## [1] "Tukey HSD - Pairwise comparison - significant differences indicated by p-value < 0.05"
-##   Tukey multiple comparisons of means
-##     95% family-wise confidence level
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-## Fit: aov(formula = index ~ treatment, data = simpsondata5_2)
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-## Group2-Group1 -0.05883652 -0.7299494 0.6122763 0.8196598
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-## data:  resid(richaov)
-## W = 0.93289, p-value = 0.6026
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-## [1] "Bartlett Test variance homogeneity - variance is homogeneous p-value > 0.05"
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-## Bartlett's K-squared = 3.2991, df = 1, p-value = 0.06932
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-##             Df Sum Sq Mean Sq F value Pr(>F)
-## treatment    1  7.288   7.288   1.561   0.28
-## Residuals    4 18.670   4.667
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-## [1] "Tukey HSD - Pairwise comparison - significant differences indicated by p-value < 0.05"
-##   Tukey multiple comparisons of means
-##     95% family-wise confidence level
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-## Fit: aov(formula = index ~ treatment, data = richdata5_2)
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-## Group2-Group1 2.204189 -2.693449 7.101827 0.2795884
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-## adonis(formula = bray.distance ~ data5$treatment)
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-## Number of permutations: 719
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-##                 Df SumsOfSqs MeanSqs F.Model      R2 Pr(>F)
-## data5$treatment  1   1.01309 1.01309  4.6942 0.53992    0.1
-## Residuals        4   0.86326 0.21582         0.46008
-## Total            5   1.87635                 1.00000
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    -  NMDS Plots -

    -
    -


    -
    -
    -

    2D NMDS

    -


    -
    ## Run 0 stress 4.35166e-05
-## Run 1 stress 9.206128e-05
-## ... Procrustes: rmse 0.2192141  max resid 0.4447335
-## Run 2 stress 0
-## ... New best solution
-## ... Procrustes: rmse 0.1475603  max resid 0.2719593
-## Run 3 stress 0.1967694
-## Run 4 stress 0
-## ... Procrustes: rmse 0.1503594  max resid 0.2805032
-## Run 5 stress 0
-## ... Procrustes: rmse 0.1890022  max resid 0.2367295
-## Run 6 stress 0
-## ... Procrustes: rmse 0.1525354  max resid 0.2259206
-## Run 7 stress 0
-## ... Procrustes: rmse 0.1446005  max resid 0.2074821
-## Run 8 stress 0
-## ... Procrustes: rmse 0.132074  max resid 0.2500861
-## Run 9 stress 0
-## ... Procrustes: rmse 0.1888654  max resid 0.3103119
-## Run 10 stress 0
-## ... Procrustes: rmse 0.1321098  max resid 0.2335853
-## Run 11 stress 0
-## ... Procrustes: rmse 0.1086904  max resid 0.198957
-## Run 12 stress 0
-## ... Procrustes: rmse 0.184796  max resid 0.2895882
-## Run 13 stress 0
-## ... Procrustes: rmse 0.1985743  max resid 0.2828424
-## Run 14 stress 0
-## ... Procrustes: rmse 0.1839276  max resid 0.339448
-## Run 15 stress 0
-## ... Procrustes: rmse 0.1569956  max resid 0.2738898
-## Run 16 stress 0
-## ... Procrustes: rmse 0.08147595  max resid 0.1293814
-## Run 17 stress 0
-## ... Procrustes: rmse 0.1231843  max resid 0.1929613
-## Run 18 stress 0
-## ... Procrustes: rmse 0.2154224  max resid 0.3725087
-## Run 19 stress 0
-## ... Procrustes: rmse 0.09658586  max resid 0.169947
-## Run 20 stress 0
-## ... Procrustes: rmse 0.1123304  max resid 0.1810664
-## Run 21 stress 0
-## ... Procrustes: rmse 0.07114148  max resid 0.08562701
-## Run 22 stress 0
-## ... Procrustes: rmse 0.1207696  max resid 0.2088403
-## Run 23 stress 0
-## ... Procrustes: rmse 0.1338395  max resid 0.2011499
-## Run 24 stress 0
-## ... Procrustes: rmse 0.2011198  max resid 0.2879716
-## Run 25 stress 7.960692e-05
-## ... Procrustes: rmse 0.1835879  max resid 0.3361377
-## Run 26 stress 7.465245e-05
-## ... Procrustes: rmse 0.1516833  max resid 0.21161
-## Run 27 stress 4.134366e-05
-## ... Procrustes: rmse 0.158652  max resid 0.1955455
-## Run 28 stress 9.401493e-05
-## ... Procrustes: rmse 0.1705531  max resid 0.3146807
-## Run 29 stress 0
-## ... Procrustes: rmse 0.140281  max resid 0.1899751
-## Run 30 stress 0
-## ... Procrustes: rmse 0.1569951  max resid 0.237474
-## Run 31 stress 0
-## ... Procrustes: rmse 0.21383  max resid 0.3404873
-## Run 32 stress 0
-## ... Procrustes: rmse 0.1831665  max resid 0.2903409
-## Run 33 stress 0
-## ... Procrustes: rmse 0.1862874  max resid 0.2430944
-## Run 34 stress 0
-## ... Procrustes: rmse 0.1398732  max resid 0.194408
-## Run 35 stress 0
-## ... Procrustes: rmse 0.08385465  max resid 0.1208572
-## Run 36 stress 0
-## ... Procrustes: rmse 0.1531769  max resid 0.2654934
-## Run 37 stress 0.1420473
-## Run 38 stress 0
-## ... Procrustes: rmse 0.168821  max resid 0.2760271
-## Run 39 stress 0
-## ... Procrustes: rmse 0.2398039  max resid 0.4101748
-## Run 40 stress 4.958545e-05
-## ... Procrustes: rmse 0.1690092  max resid 0.224612
-## Run 41 stress 0
-## ... Procrustes: rmse 0.1763536  max resid 0.2979007
-## Run 42 stress 8.50985e-05
-## ... Procrustes: rmse 0.2037257  max resid 0.3652396
-## Run 43 stress 0
-## ... Procrustes: rmse 0.1373896  max resid 0.1978843
-## Run 44 stress 0
-## ... Procrustes: rmse 0.1721457  max resid 0.2384637
-## Run 45 stress 8.338233e-06
-## ... Procrustes: rmse 0.1350539  max resid 0.1826055
-## Run 46 stress 8.733644e-05
-## ... Procrustes: rmse 0.1864883  max resid 0.2843063
-## Run 47 stress 0
-## ... Procrustes: rmse 0.1287125  max resid 0.1800035
-## Run 48 stress 1.294414e-05
-## ... Procrustes: rmse 0.06488975  max resid 0.09540012
-## Run 49 stress 0
-## ... Procrustes: rmse 0.1123794  max resid 0.2193415
-## Run 50 stress 0
-## ... Procrustes: rmse 0.0873931  max resid 0.1388494
-## Run 51 stress 0
-## ... Procrustes: rmse 0.1836172  max resid 0.2892836
-## Run 52 stress 0
-## ... Procrustes: rmse 0.1349277  max resid 0.2496178
-## Run 53 stress 0
-## ... Procrustes: rmse 0.1597566  max resid 0.3010393
-## Run 54 stress 0
-## ... Procrustes: rmse 0.1578135  max resid 0.2739341
-## Run 55 stress 0.1420473
-## Run 56 stress 9.580941e-05
-## ... Procrustes: rmse 0.2385671  max resid 0.3099702
-## Run 57 stress 0
-## ... Procrustes: rmse 0.1424363  max resid 0.203378
-## Run 58 stress 0
-## ... Procrustes: rmse 0.1498625  max resid 0.2312135
-## Run 59 stress 0
-## ... Procrustes: rmse 0.08465851  max resid 0.1346232
-## Run 60 stress 0
-## ... Procrustes: rmse 0.1430299  max resid 0.2610707
-## Run 61 stress 0
-## ... Procrustes: rmse 0.1369128  max resid 0.1816959
-## Run 62 stress 0
-## ... Procrustes: rmse 0.08119163  max resid 0.1444967
-## Run 63 stress 0
-## ... Procrustes: rmse 0.1710694  max resid 0.240634
-## Run 64 stress 9.84601e-05
-## ... Procrustes: rmse 0.1878078  max resid 0.2714905
-## Run 65 stress 0
-## ... Procrustes: rmse 0.05651144  max resid 0.09672819
-## Run 66 stress 8.62831e-05
-## ... Procrustes: rmse 0.0727498  max resid 0.09722458
-## Run 67 stress 0
-## ... Procrustes: rmse 0.1958305  max resid 0.2910121
-## Run 68 stress 2.701363e-05
-## ... Procrustes: rmse 0.2225825  max resid 0.3507306
-## Run 69 stress 7.042554e-05
-## ... Procrustes: rmse 0.173566  max resid 0.2626667
-## Run 70 stress 2.745518e-05
-## ... Procrustes: rmse 0.08284383  max resid 0.124135
-## Run 71 stress 0
-## ... Procrustes: rmse 0.1840454  max resid 0.3503948
-## Run 72 stress 0
-## ... Procrustes: rmse 0.1540941  max resid 0.2302387
-## Run 73 stress 0
-## ... Procrustes: rmse 0.07616243  max resid 0.1113843
-## Run 74 stress 0
-## ... Procrustes: rmse 0.1348955  max resid 0.2398141
-## Run 75 stress 0
-## ... Procrustes: rmse 0.1593818  max resid 0.2624199
-## Run 76 stress 0
-## ... Procrustes: rmse 0.1658679  max resid 0.2368498
-## Run 77 stress 0
-## ... Procrustes: rmse 0.1083634  max resid 0.1355875
-## Run 78 stress 0
-## ... Procrustes: rmse 0.173316  max resid 0.2188972
-## Run 79 stress 2.412692e-05
-## ... Procrustes: rmse 0.1513869  max resid 0.2559945
-## Run 80 stress 0
-## ... Procrustes: rmse 0.1286293  max resid 0.1708846
-## Run 81 stress 8.249084e-05
-## ... Procrustes: rmse 0.1600543  max resid 0.2790534
-## Run 82 stress 0
-## ... Procrustes: rmse 0.06425533  max resid 0.09782641
-## Run 83 stress 0
-## ... Procrustes: rmse 0.1859754  max resid 0.3306764
-## Run 84 stress 1.218079e-05
-## ... Procrustes: rmse 0.1579732  max resid 0.2679705
-## Run 85 stress 0
-## ... Procrustes: rmse 0.1373995  max resid 0.1704452
-## Run 86 stress 0
-## ... Procrustes: rmse 0.1135352  max resid 0.1456319
-## Run 87 stress 3.345478e-06
-## ... Procrustes: rmse 0.1136627  max resid 0.1526098
-## Run 88 stress 7.30298e-05
-## ... Procrustes: rmse 0.09087387  max resid 0.1523526
-## Run 89 stress 0
-## ... Procrustes: rmse 0.1294085  max resid 0.20968
-## Run 90 stress 0
-## ... Procrustes: rmse 0.2133609  max resid 0.307295
-## Run 91 stress 0
-## ... Procrustes: rmse 0.208796  max resid 0.3331994
-## *** No convergence -- monoMDS stopping criteria:
-##     88: stress < smin
-##      1: stress ratio > sratmax
-##      2: scale factor of the gradient < sfgrmin
    -
    ## [1] "No Convergence"
    -





    -
    -
    -

    3D NMDS

    -


    -
    ## Run 0 stress 9.179259e-05
-## Run 1 stress 0
-## ... New best solution
-## ... Procrustes: rmse 0.1912644  max resid 0.3064663
-## Run 2 stress 0
-## ... Procrustes: rmse 0.1515697  max resid 0.1940969
-## Run 3 stress 8.011522e-05
-## ... Procrustes: rmse 0.2408658  max resid 0.3233412
-## Run 4 stress 0
-## ... Procrustes: rmse 0.1502567  max resid 0.2373834
-## Run 5 stress 0
-## ... Procrustes: rmse 0.1824273  max resid 0.2309358
-## Run 6 stress 0
-## ... Procrustes: rmse 0.2044446  max resid 0.3064849
-## Run 7 stress 0
-## ... Procrustes: rmse 0.1360824  max resid 0.1868316
-## Run 8 stress 0
-## ... Procrustes: rmse 0.224697  max resid 0.3889081
-## Run 9 stress 1.601269e-06
-## ... Procrustes: rmse 0.2017323  max resid 0.253622
-## Run 10 stress 0
-## ... Procrustes: rmse 0.2332277  max resid 0.2830118
-## Run 11 stress 0
-## ... Procrustes: rmse 0.2094351  max resid 0.3393129
-## Run 12 stress 0
-## ... Procrustes: rmse 0.1924744  max resid 0.2476726
-## Run 13 stress 0
-## ... Procrustes: rmse 0.1723438  max resid 0.2779048
-## Run 14 stress 0
-## ... Procrustes: rmse 0.231938  max resid 0.3082758
-## Run 15 stress 0
-## ... Procrustes: rmse 0.1591339  max resid 0.2941593
-## Run 16 stress 1.844663e-05
-## ... Procrustes: rmse 0.1367305  max resid 0.1893536
-## Run 17 stress 0
-## ... Procrustes: rmse 0.2432104  max resid 0.3742779
-## Run 18 stress 0
-## ... Procrustes: rmse 0.2021536  max resid 0.3190184
-## Run 19 stress 8.261138e-05
-## ... Procrustes: rmse 0.2661264  max resid 0.3395278
-## Run 20 stress 0
-## ... Procrustes: rmse 0.2050021  max resid 0.3470228
-## Run 21 stress 0
-## ... Procrustes: rmse 0.2349675  max resid 0.3899205
-## Run 22 stress 0
-## ... Procrustes: rmse 0.1743172  max resid 0.3327414
-## Run 23 stress 0
-## ... Procrustes: rmse 0.1805854  max resid 0.2484891
-## Run 24 stress 0
-## ... Procrustes: rmse 0.2214565  max resid 0.3735158
-## Run 25 stress 6.602248e-05
-## ... Procrustes: rmse 0.2633094  max resid 0.3328158
-## Run 26 stress 0
-## ... Procrustes: rmse 0.1989565  max resid 0.3402747
-## Run 27 stress 0
-## ... Procrustes: rmse 0.2434265  max resid 0.2853429
-## Run 28 stress 0
-## ... Procrustes: rmse 0.1437204  max resid 0.2008141
-## Run 29 stress 0
-## ... Procrustes: rmse 0.2207409  max resid 0.2922027
-## Run 30 stress 0
-## ... Procrustes: rmse 0.140996  max resid 0.2320052
-## Run 31 stress 0
-## ... Procrustes: rmse 0.1889331  max resid 0.2900138
-## Run 32 stress 0
-## ... Procrustes: rmse 0.1834047  max resid 0.2422162
-## Run 33 stress 0
-## ... Procrustes: rmse 0.2447367  max resid 0.3580808
-## Run 34 stress 0
-## ... Procrustes: rmse 0.2359379  max resid 0.3172238
-## Run 35 stress 0
-## ... Procrustes: rmse 0.2428633  max resid 0.3027342
-## Run 36 stress 0
-## ... Procrustes: rmse 0.2011346  max resid 0.344021
-## Run 37 stress 0
-## ... Procrustes: rmse 0.2398276  max resid 0.4186627
-## Run 38 stress 6.012963e-05
-## ... Procrustes: rmse 0.2290966  max resid 0.3002383
-## Run 39 stress 0
-## ... Procrustes: rmse 0.2681661  max resid 0.4893852
-## Run 40 stress 2.102041e-05
-## ... Procrustes: rmse 0.1696101  max resid 0.2438495
-## Run 41 stress 0
-## ... Procrustes: rmse 0.2461656  max resid 0.3874782
-## Run 42 stress 8.813298e-05
-## ... Procrustes: rmse 0.2458929  max resid 0.3572947
-## Run 43 stress 1.301719e-06
-## ... Procrustes: rmse 0.196147  max resid 0.322213
-## Run 44 stress 0
-## ... Procrustes: rmse 0.1521486  max resid 0.2327491
-## Run 45 stress 7.430631e-05
-## ... Procrustes: rmse 0.2806763  max resid 0.3795962
-## Run 46 stress 0
-## ... Procrustes: rmse 0.1268192  max resid 0.1828082
-## Run 47 stress 0
-## ... Procrustes: rmse 0.1656652  max resid 0.2345423
-## Run 48 stress 0
-## ... Procrustes: rmse 0.2071554  max resid 0.3748716
-## Run 49 stress 8.899945e-05
-## ... Procrustes: rmse 0.2411995  max resid 0.3239697
-## Run 50 stress 8.49387e-05
-## ... Procrustes: rmse 0.2235364  max resid 0.2957555
-## Run 51 stress 8.946254e-05
-## ... Procrustes: rmse 0.2411916  max resid 0.3239556
-## Run 52 stress 0
-## ... Procrustes: rmse 0.1024931  max resid 0.1772733
-## Run 53 stress 0
-## ... Procrustes: rmse 0.1805318  max resid 0.2266111
-## Run 54 stress 0
-## ... Procrustes: rmse 0.222073  max resid 0.3309312
-## Run 55 stress 0
-## ... Procrustes: rmse 0.1984136  max resid 0.2676206
-## Run 56 stress 4.061045e-05
-## ... Procrustes: rmse 0.1650274  max resid 0.2850801
-## Run 57 stress 0
-## ... Procrustes: rmse 0.2278851  max resid 0.350922
-## Run 58 stress 0
-## ... Procrustes: rmse 0.1401086  max resid 0.185101
-## Run 59 stress 8.737338e-05
-## ... Procrustes: rmse 0.2411935  max resid 0.3239583
-## Run 60 stress 0
-## ... Procrustes: rmse 0.2191345  max resid 0.3940798
-## Run 61 stress 2.198444e-05
-## ... Procrustes: rmse 0.2350082  max resid 0.3891647
-## Run 62 stress 0
-## ... Procrustes: rmse 0.2048775  max resid 0.265205
-## Run 63 stress 0
-## ... Procrustes: rmse 0.2213181  max resid 0.3503163
-## Run 64 stress 0
-## ... Procrustes: rmse 0.1501829  max resid 0.2294057
-## Run 65 stress 0
-## ... Procrustes: rmse 0.1411666  max resid 0.2546569
-## Run 66 stress 0
-## ... Procrustes: rmse 0.1464234  max resid 0.2116882
-## Run 67 stress 0
-## ... Procrustes: rmse 0.1652729  max resid 0.2477694
-## Run 68 stress 9.870175e-05
-## ... Procrustes: rmse 0.2505968  max resid 0.3328449
-## Run 69 stress 0.07976986
-## Run 70 stress 0
-## ... Procrustes: rmse 0.194331  max resid 0.2432624
-## Run 71 stress 6.941015e-05
-## ... Procrustes: rmse 0.2130009  max resid 0.3004891
-## Run 72 stress 7.615578e-05
-## ... Procrustes: rmse 0.259831  max resid 0.34112
-## Run 73 stress 0
-## ... Procrustes: rmse 0.1647591  max resid 0.2388743
-## Run 74 stress 0
-## ... Procrustes: rmse 0.1644595  max resid 0.2847747
-## Run 75 stress 0
-## ... Procrustes: rmse 0.1906808  max resid 0.3100185
-## Run 76 stress 0
-## ... Procrustes: rmse 0.146477  max resid 0.2315368
-## Run 77 stress 7.361458e-05
-## ... Procrustes: rmse 0.2474006  max resid 0.3599023
-## Run 78 stress 0
-## ... Procrustes: rmse 0.1476474  max resid 0.2302322
-## Run 79 stress 0
-## ... Procrustes: rmse 0.1500008  max resid 0.1972427
-## Run 80 stress 0
-## ... Procrustes: rmse 0.2317812  max resid 0.3787597
-## Run 81 stress 0
-## ... Procrustes: rmse 0.2462261  max resid 0.3285394
-## Run 82 stress 0
-## ... Procrustes: rmse 0.1343457  max resid 0.1992015
-## Run 83 stress 0
-## ... Procrustes: rmse 0.1171339  max resid 0.2019657
-## Run 84 stress 0
-## ... Procrustes: rmse 0.1754114  max resid 0.255796
-## Run 85 stress 0
-## ... Procrustes: rmse 0.17915  max resid 0.2562237
-## Run 86 stress 0
-## ... Procrustes: rmse 0.2158695  max resid 0.2893261
-## Run 87 stress 0
-## ... Procrustes: rmse 0.1897326  max resid 0.3018936
-## Run 88 stress 8.264709e-05
-## ... Procrustes: rmse 0.2704232  max resid 0.3402092
-## Run 89 stress 0
-## ... Procrustes: rmse 0.1770887  max resid 0.2768628
-## Run 90 stress 7.124833e-05
-## ... Procrustes: rmse 0.2274275  max resid 0.3037865
-## Run 91 stress 0
-## ... Procrustes: rmse 0.2291196  max resid 0.3758455
-## *** No convergence -- monoMDS stopping criteria:
-##     90: stress < smin
-##      1: stress ratio > sratmax
    -
    ## [1] "No Convergence"
    -






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    -  OTU Abundance Per Sample -

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    -  OTU Abundance Per Treatment -

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    -

    -  Pairwise Percent-ID Heatmap -

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    -  Taxonomy Result Visualization -

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    -

    -  Phylogenetic Tree -

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    -

    - This tree is a maximum likelihood tree made with IQTREE2 and the parameters you specified in the vampirus.config file. Also, this is an interactive tree, you can zoom in and hover on nodes to know the sequence ID. For a better visualization of this tree, you can find the *.treefile with bootstrap support values within the results directory and visualize using programs like FigTree or ITOL.








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    - - - - - - - - - - - - - - - diff --git a/example_data/conf/ex_reports/example_DataCheck_Report.html b/example_data/conf/ex_reports/example_DataCheck_Report.html deleted file mode 100644 index c903911..0000000 --- a/example_data/conf/ex_reports/example_DataCheck_Report.html +++ /dev/null @@ -1,391 +0,0 @@ - - - - - - - - - - - - - -vAMPirus DataCheck Report: vAMPtest - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - -
    - - - - - - - - -

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    —————————————————————————————————————————————————————————————————————— NOTE: Plots are interactive and you can use the legend to specify samples/treatment of interest. You can also download an .svg version of each figure within this report. ——————————————————————————————————————————————————————————————————————
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    -  Pre- and Post-Adapter Removal Read Stats -

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    Total number of reads before and after adapter removal

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    Forward (R1) and reverse (R2) read length before and after adapter removal

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    -  Post-Merging Read Stats -

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    Pre-filtering reads per sample

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    Post-filtering reads per sample

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    Pre-filtering base frequency per position on reads

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    Post-filtering base frequency per position on reads

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    Pre-filtering mean quality score per position on reads

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    Post-filtering mean quality score per position on reads

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    Pre-filtering read GC-content

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    -
    ## Mean,45.279
-## Median,45.309
-## Mode,45.110
-## STDev,1.617
-## GC,Count
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    Post-filtering read GC-content

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    -
    ## Mean,45.656
-## Median,45.476
-## Mode,44.762
-## STDev,1.219
-## GC,Count
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    Number of reads per quality score (pre-filtering)

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    Number of reads per quality score (post-filtering)

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    Number of reads per length (pre-filtering)

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    Number of reads per length (post-filtering)

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    -  Clustering Statistics -

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    Number of ncASVs per clustering percentage – “1” represents number of ASVs

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    - NOTE: The “1” on the x-axis represents number of ASVs identified by vsearch

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    Number of pcASVs per clustering percentage

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    - NOTE: The “1” represents the number of AminoTypes which are unique amino acid sequences in your dataset








    -
    - - - - -
    - - - - - - - - - - - - - - - diff --git a/example_data/conf/test.config b/example_data/conf/test.config index c75ce75..7149b5d 100644 --- a/example_data/conf/test.config +++ b/example_data/conf/test.config @@ -19,5 +19,7 @@ params { dbname="test_db.fasta" dbdir="${projectDir}/example_data/conf/" dbtype="RVDB" + aminoC=5 + asvC=5 stats = true } From c969d278601fe20b50b02a9ddd2ce339c76fd785 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Tue, 21 Sep 2021 16:10:23 -0500 Subject: [PATCH 174/223] added filtering processes and added the use of evalue in taxonomy step --- docs/HelpDocumentation.md | 16 ++-- vAMPirus.nf | 182 ++++++++++++++++++++++++++++++++++---- vampirus.config | 37 ++++++-- 3 files changed, 205 insertions(+), 30 deletions(-) diff --git a/docs/HelpDocumentation.md b/docs/HelpDocumentation.md index 35fcf4a..689d892 100644 --- a/docs/HelpDocumentation.md +++ b/docs/HelpDocumentation.md @@ -412,7 +412,7 @@ General execution: vampirus_startup.sh -h [-d 1|2|3|4] [-s] [-t] - Command lime options: + Command line options: [ -h ] Print help information @@ -449,7 +449,7 @@ RVDB format -> ">acc|GENBANK|AYD68780.1|GENBANK|MH171300|structural polyprotein NCBI NR/RefSeq format -> ">KJX92028.1 hypothetical protein TI39_contig5958g00003 [Zymoseptoria brevis]" -To set/inform vAMPirus of which header format for the reference database is being used, you can edit the vampirus.config file at lime 112 "dbtype="NCBI"" for NCBI header format or "dbtype="RVDB"" for RVDB format. +To set/inform vAMPirus of which header format for the reference database is being used, you can edit the vampirus.config file at line 122 "dbtype="NCBI"" for NCBI header format or "dbtype="RVDB"" for RVDB format. An example of custom headers in RVDB format if you plan to use a custom database: @@ -636,7 +636,7 @@ Nextflow deployment of vAMPirus relies on the use of the configuration file (vam Furthermore, the configuration file contains analysis-specific parameters AND resource-specific Nextflow launching parameters. A benefit of Nextflow integration, is that you can run the vAMPirus workflow on a large HPC just as easily as you could on your local machine. -If you look at lime 151 and greater in the vampirus.config file, you will see resource-specific parameters that you can alter before any run. Nexflow is capable of submitting jobs automatically using slurm and PBS, check out the Nextflow docs to learn more (https://www.nextflow.io/docs/latest/executor.html)! +If you look at line 233 and greater in the vampirus.config file, you will see resource-specific parameters that you can alter before any run. Nexflow is capable of submitting jobs automatically using slurm and PBS, check out the Nextflow docs to learn more (https://www.nextflow.io/docs/latest/executor.html)! ### Setting parameter values @@ -672,15 +672,15 @@ Instead of editing the configuration file directly, you could set parameters wit nextflow run vAMPirus.nf -c vampirus.config -profile [conda|singularity] --Analyze --ncASV --clusterNuclID .95 -Here we use the "--Analyze" option that tells vAMPirus that we are ready to analyze some data. Then the "--ncASV" argument with the "--clisterNuclID .95" tells vAMPirus we would like to cluster our ASVs based on 95% nucleotide similarity. The default ID value is stored at lime 51 in the vampirus.config file (currently 85%), but as soon as you specify and provide a value in the command, the value within the config file is ignored. +Here we use the "--Analyze" option that tells vAMPirus that we are ready to analyze some data. Then the "--ncASV" argument with the "--clisterNuclID .95" tells vAMPirus we would like to cluster our ASVs based on 95% nucleotide similarity. The default ID value is stored at line 66 in the vampirus.config file (currently 85%), but as soon as you specify and provide a value in the command, the value within the config file is ignored. NOTE: Nextflow also has options in the launch command. To tell them apart, Nextflow options uses a single dash (e.g. -with-conda or -profile) while vAMPirus options are always with a double dash (e.g. --Analyze) -### Setting computing resource parameters - Edit in lines 151-171 in vampirus.config +### Setting computing resource parameters - Edit in lines 241-261 in vampirus.config Each process within the vAMPirus workflow is tagged with either "low_cpus", "norm_cpus", or "high_cpus" (see below) which let's Nextflow know the amount of cpus and memory required for each process, which will then be used for when Nextflow submits a given job or task. Nexflow actively assesses the amount of available resources on your machine and will submit tasks only when the proper amount of resources can be requested. -From lime 203-217 in the vAMPirus.config file is where you can edit these values for whichever machine you plan to run the workflow on. +From line 241-261 in the vAMPirus.config file is where you can edit these values for whichever machine you plan to run the workflow on. process { withLabel: low_cpus { @@ -763,7 +763,7 @@ This launch command will run all aspects of the vAMPirus workflow on your data a ### Sequencing reads -Input can be raw or processed compressed or non-compressed fastq files with names containing "\_R1" or "\_R2". You can specify the directory containing your reads in lime 25 of the vampirus.config file. +Input can be raw or processed compressed or non-compressed fastq files with names containing "\_R1" or "\_R2". You can specify the directory containing your reads in line 20 of the vampirus.config file. NOTE: Sample names are extracted from read library names by using the string to the left of the "\_R" in the filename automatically. @@ -1326,7 +1326,7 @@ NOTE=> Be sure that there is more than 1 sample in each treatment category or th # vAMPirus output -There are several files created throughout the vAMPirus pipeline that are stored and organized in directories within the specified results/output directory (ex. ${working_directory}/results; lime 27 in the configuration file). We will go through the structure of the output directory and where to find which files here: +There are several files created throughout the vAMPirus pipeline that are stored and organized in directories within the specified results/output directory (ex. ${working_directory}/results; line 24 in the configuration file). We will go through the structure of the output directory and where to find which files here: ## Pipeline performance information - ${working_directory}/${outdir}/PipelinePerformance/ diff --git a/vAMPirus.nf b/vAMPirus.nf index b65d710..4170216 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -756,7 +756,7 @@ if (params.DataCheck || params.Analyze) { script: """ - vsearch --fastq_mergepairs ${reads[0]} --reverse ${reads[1]} --threads ${task.cpus} --fastqout ${sample_id}_mergedclean.fastq --fastqout_notmerged_fwd ${sample_id}_notmerged_fwd.fastq --fastqout_notmerged_rev ${sample_id}_notmerged_rev.fastq --fastq_maxdiffs ${params.diffs} --fastq_maxns ${params.maxn} --fastq_allowmergestagger --fastq_maxee ${params.maxEE} --relabel ${sample_id}. + vsearch --fastq_mergepairs ${reads[0]} --reverse ${reads[1]} --threads ${task.cpus} --fastqout ${sample_id}_mergedclean.fastq --fastqout_notmerged_fwd ${sample_id}_notmerged_fwd.fastq --fastqout_notmerged_rev ${sample_id}_notmerged_rev.fastq --fastq_maxdiffs ${params.diffs} --fastq_maxns ${params.maxn} --fastq_allowmergestagger --fastq_maxee ${params.maxEE} --fastq_minovlen ${params.minoverlap} --relabel ${sample_id}. echo ${sample_id} > ${sample_id}.name """ @@ -914,7 +914,8 @@ if (params.DataCheck || params.Analyze) { file(fasta) from reads_vsearch4_ch output: - file("*ASVs.fasta") into ( reads_vsearch5_ch, asv_med, nucl2aa, asvsforAminotyping, asvfastaforcounts, asvaminocheck ) + file("*ASVs.fasta") into asvforfilt + script: """ @@ -923,6 +924,93 @@ if (params.DataCheck || params.Analyze) { } // UNTIL HERE DEFAULT + if (params.filter) { + + process Filtering { + + label 'norm_cpus' + + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/FilterCont", mode: "copy", overwrite: true, pattern: '*.fasta' + + input: + file(asv) from asvforfilt + + output: + file("*ASV.fasta") into ( reads_vsearch5_ch, asv_med, nucl2aa, asvsforAminotyping, asvfastaforcounts, asvaminocheck ) + + script: + """ + cp ${params.vampdir}/bin/rename_seq.py . + + #create and rename filter database + grep ">" ${params.filtDB} | awk -F ">" '{print \$2}' >> filt.head + j=1 + for y in \$( cat filt.head );do + echo ">Filt"\$j"" >> filt.headers + j=\$(( \${j}+1 )) + done + ./rename_seq.py ${params.filtDB} filt.headers filterdatabaserenamed.fasta + cat filterdatabase.fasta >> combodatabase.fasta + paste -d',' filt.head filt.headers > filtername_map.csv + rm filterdatabaserenamed.fasta + + #create and rename keep database + grep ">" ${params.keepDB} | awk -F ">" '{print \$2}' >> keep.head + d=1 + for y in \$( cat keep.head );do + echo ">keep"\$d"" >> keep.headers + d=\$(( \${d}+1 )) + done + ./rename_seq.py ${params.keepDB} keep.headers keepdatabaserenamed.fasta + cat keepdatabaserenamed.fasta >> combodatabase.fasta + paste -d',' keep.head keep.headers > keepername_map.csv + rm filterdatabaserenamed.fasta + #index database + diamond makedb --in combodatabase.fasta --db combodatabase.fasta + #run diamond_db + diamond blastx -q ${asv} -d combodatabase.fasta -p ${task.cpus} --id ${params.filtminID} -l ${params.filtminaln} -e ${params.filtevalue} --${params.filtsensitivity} -o ${params.projtag}_diamondfilter.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident --max-target-seqs 1 --max-hsps 1 + #get asvs + grep ">" ${asv} | awk -F ">" '{print \$2}' > asv.list + for x in $(cat asv.list) + do #check for a hit + if [[ \$(grep -c "\$x" ${params.projtag}_diamondfilter.out) -eq 1 ]] + then #check if hit is to filter + hit=\$(grep "\$x" ${params.projtag}_diamondfilter.out | awk '{print \$3}') + if [[ \$(grep -c "\$hit" filt.headers) -eq 1 ]] + then echo "\$x,\$hit >> filtered_asvs_summary.csv + elif [[ \$(grep -c "\$hit" keep.headers) -eq 1 ]] + then echo "\$x" >> kep.list + fi + else echo \$x >> nohit.list + fi + done + if [[ ${params.keepnohit} == "true" ]] + then echo nohit.list >> kep.list + cat kep.list | sort > keep.list + seqtk subseq ${asv} keep.list > kept.fasta + u=1 + for y in \$( cat keep.list );do + echo ">ASV"\$u"" >> asvrename.list + u=\$(( \${u}+1 )) + done + ./rename_seq.py ${asv} asvrename.list ${params.projtag}_ASV.fasta + else + cat kep.list | sort > keep.list + seqtk subseq ${asv} keep.list > kept.fasta + u=1 + for y in \$( cat keep.list );do + echo ">ASV"\$u"" >> asvrename.list + u=\$(( \${u}+1 )) + done + ./rename_seq.py ${asv} asvrename.list ${params.projtag}_ASV.fasta + fi + """ + } + + } else { + asvforfilt + .into{ reads_vsearch5_ch, asv_med, nucl2aa, asvsforAminotyping, asvfastaforcounts, asvaminocheck } + } if (params.DataCheck) { @@ -1367,12 +1455,18 @@ if (params.DataCheck || params.Analyze) { """ cp ${params.vampdir}/bin/rename_seq.py . virdb=${params.dbdir}/${params.dbname} + if [[ ${params.measurement} == "bitscore"]] + then measure="--min-score ${params.bitscore}" + elif [[ ${params.measurement} == "evalue"]] + then measure="-e ${params.evalue}" + else measure="--min-score ${params.bitscore}" + fi grep ">" \${virdb} > headers.list headers="headers.list" name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') if [[ ${params.ncbitax} == "true" ]] - then diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${params.sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop staxids sskingdoms skingdoms sphylums --max-target-seqs 1 --max-hsps 1 - else diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${params.sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + then diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} \${measure} --${params.sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop staxids sskingdoms skingdoms sphylums --max-target-seqs 1 --max-hsps 1 + else diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} \${measure} --${params.sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 fi echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" > access.list @@ -1515,10 +1609,16 @@ if (params.DataCheck || params.Analyze) { """ cp ${params.vampdir}/bin/rename_seq.py . virdb=${params.dbdir}/${params.dbname} + if [[ ${params.measurement} == "bitscore"]] + then measure="--min-score ${params.bitscore}" + elif [[ ${params.measurement} == "evalue"]] + then measure="-e ${params.evalue}" + else measure="--min-score ${params.bitscore}" + fi grep ">" \${virdb} > headers.list headers="headers.list" name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${params.sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} \${measure} --${params.sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" > access.list echo "[e-value]" > evalue.list @@ -1750,12 +1850,18 @@ if (params.DataCheck || params.Analyze) { """ cp ${params.vampdir}/bin/rename_seq.py . virdb=${params.dbdir}/${params.dbname} + if [[ ${params.measurement} == "bitscore"]] + then measure="--min-score ${params.bitscore}" + elif [[ ${params.measurement} == "evalue"]] + then measure="-e ${params.evalue}" + else measure="--min-score ${params.bitscore}" + fi grep ">" \${virdb} > headers.list headers="headers.list" name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') if [[ ${params.ncbitax} == "true" ]] - then diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${params.sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop staxids sskingdoms skingdoms sphylums --max-target-seqs 1 --max-hsps 1 - else diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${params.sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + then diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} \${measure} --${params.sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop staxids sskingdoms skingdoms sphylums --max-target-seqs 1 --max-hsps 1 + else diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} \${measure} --${params.sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 fi echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" > access.list @@ -1896,9 +2002,15 @@ if (params.DataCheck || params.Analyze) { cp ${params.vampdir}/bin/rename_seq.py . virdb=${params.dbdir}/${params.dbname} grep ">" \${virdb} > headers.list + if [[ ${params.measurement} == "bitscore"]] + then measure="--min-score ${params.bitscore}" + elif [[ ${params.measurement} == "evalue"]] + then measure="-e ${params.evalue}" + else measure="--min-score ${params.bitscore}" + fi headers="headers.list" name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${params.sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} \${measure} --${params.sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" > access.list echo "[e-value]" > evalue.list @@ -2431,12 +2543,18 @@ if (params.DataCheck || params.Analyze) { """ cp ${params.vampdir}/bin/rename_seq.py . virdb=${params.dbdir}/${params.dbname} + if [[ ${params.measurement} == "bitscore"]] + then measure="--min-score ${params.bitscore}" + elif [[ ${params.measurement} == "evalue"]] + then measure="-e ${params.evalue}" + else measure="--min-score ${params.bitscore}" + fi grep ">" \${virdb} > headers.list headers="headers.list" name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') if [[ ${params.ncbitax} == "true" ]] - then diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${params.sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop staxids sskingdoms skingdoms sphylums --max-target-seqs 1 --max-hsps 1 - else diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${params.sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + then diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} \${measure} --${params.sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop staxids sskingdoms skingdoms sphylums --max-target-seqs 1 --max-hsps 1 + else diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} \${measure} --${params.sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 fi echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" > access.list @@ -2576,10 +2694,16 @@ if (params.DataCheck || params.Analyze) { """ cp ${params.vampdir}/bin/rename_seq.py . virdb=${params.dbdir}/${params.dbname} + if [[ ${params.measurement} == "bitscore"]] + then measure="--min-score ${params.bitscore}" + elif [[ ${params.measurement} == "evalue"]] + then measure="-e ${params.evalue}" + else measure="--min-score ${params.bitscore}" + fi grep ">" \${virdb} > headers.list headers="headers.list" name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${params.sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} \${measure} --${params.sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" > access.list echo "[e-value]" > evalue.list @@ -3070,12 +3194,18 @@ if (params.DataCheck || params.Analyze) { set +e cp ${params.vampdir}/bin/rename_seq.py . virdb=${params.dbdir}/${params.dbname} + if [[ ${params.measurement} == "bitscore"]] + then measure="--min-score ${params.bitscore}" + elif [[ ${params.measurement} == "evalue"]] + then measure="-e ${params.evalue}" + else measure="--min-score ${params.bitscore}" + fi grep ">" \${virdb} > headers.list headers="headers.list" name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') if [[ ${params.ncbitax} == "true" ]] - then diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${params.sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop staxids sskingdoms skingdoms sphylums --max-target-seqs 1 --max-hsps 1 - else diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${params.sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + then diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} \${measure} --${params.sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop staxids sskingdoms skingdoms sphylums --max-target-seqs 1 --max-hsps 1 + else diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} \${measure} --${params.sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 fi echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" > access.list @@ -3220,10 +3350,16 @@ if (params.DataCheck || params.Analyze) { set +e cp ${params.vampdir}/bin/rename_seq.py . virdb=${params.dbdir}/${params.dbname} + if [[ ${params.measurement} == "bitscore"]] + then measure="--min-score ${params.bitscore}" + elif [[ ${params.measurement} == "evalue"]] + then measure="-e ${params.evalue}" + else measure="--min-score ${params.bitscore}" + fi grep ">" \${virdb} > headers.list headers="headers.list" name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${params.sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + diamond blastx -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} \${measure} --${params.sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" > access.list echo "[e-value]" > evalue.list @@ -3539,12 +3675,18 @@ if (params.DataCheck || params.Analyze) { """ cp ${params.vampdir}/bin/rename_seq.py . virdb=${params.dbdir}/${params.dbname} + if [[ ${params.measurement} == "bitscore"]] + then measure="--min-score ${params.bitscore}" + elif [[ ${params.measurement} == "evalue"]] + then measure="-e ${params.evalue}" + else measure="--min-score ${params.bitscore}" + fi grep ">" \${virdb} > headers.list headers="headers.list" name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') if [[ ${params.ncbitax} == "true" ]] - then diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${params.sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop staxids sskingdoms skingdoms sphylums --max-target-seqs 1 --max-hsps 1 - else diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${params.sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + then diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} \${measure} --${params.sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop staxids sskingdoms skingdoms sphylums --max-target-seqs 1 --max-hsps 1 + else diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} \${measure} --${params.sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 fi echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" > access.list @@ -3688,10 +3830,16 @@ if (params.DataCheck || params.Analyze) { """ cp ${params.vampdir}/bin/rename_seq.py . virdb=${params.dbdir}/${params.dbname} + if [[ ${params.measurement} == "bitscore"]] + then measure="--min-score ${params.bitscore}" + elif [[ ${params.measurement} == "evalue"]] + then measure="-e ${params.evalue}" + else measure="--min-score ${params.bitscore}" + fi grep ">" \${virdb} > headers.list headers="headers.list" name=\$( echo ${asvs} | awk -F ".fasta" '{print \$1}') - diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} --min-score ${params.bitscore} --${params.sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 + diamond blastp -q ${asvs} -d \${virdb} -p ${task.cpus} --id ${params.minID} -l ${params.minaln} \${measure} --${params.sensitivity} -o "\$name"_dmd.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident btop --max-target-seqs 1 --max-hsps 1 echo "Preparing lists to generate summary .csv's" echo "[Best hit accession number]" > access.list echo "[e-value]" > evalue.list diff --git a/vampirus.config b/vampirus.config index a892240..8c34321 100644 --- a/vampirus.config +++ b/vampirus.config @@ -55,6 +55,8 @@ params { diffs="20" // Maximum number of "N"'s in a sequence - if above the specified value, sequence will be discarded maxn="20" + // Minimum overlap for sequence merging + minoverlap="10" // ASV generation and clustering parameters @@ -73,6 +75,27 @@ params { // Minimum length of amino acid translation to be considered during protein clustered ASV (pcASV) generation. Recommended to put this at the expected amino acid sequence length based on your maximum read length (e.g. if maxLen="420", then minAA should be 420/3 so 140) minAA="140" + // Filtering parameters - You can set the filtering to run with the command --filter + + // Path to database + filtDB="" + // Path to database of + keepDB="" + // Keep any sequences without hits - for yes, set keepnohit to ="true" + keepnohit="true" + + //Parameters for diamond command for filtering + + // Set minimum percent amino acid similarity for best hit to be counted in taxonomy assignment + filtminID="80" + // Set minimum amino acid alignment length for best hit to be counted in taxonomy assignment + filtminaln="30" + // Set sensitivity parameters for DIAMOND aligner (read more here: https://github.com/bbuchfink/diamond/wiki; default = ultra-sensitive) + filtsensitivity="ultra-sensitive" + // Set the max e-value for best hit to be recorded + filtevalue="0.001" + + // Minimum Entropy Decomposition (MED) parameters for clustering (https://merenlab.org/2012/05/11/oligotyping-pipeline-explained/) // If you plan to do MED on ASVs using the option "--asvMED" you can set here the number of entopy peak positions or a comma seperated list of biologically meaningful positons (e.g. 35,122,21) for oligotyping to take into consideration. If you want to use a single specific position, make "asvSingle="true"". @@ -97,8 +120,12 @@ params { // Taxonomy inference parameters //Parameters for diamond command - // Set minimum bitscore for best hit in taxonomy assignment - bitscore="50" + // Set which measurement to use for a minimum threshold in taxonomy inference - must be either "evalue" or "bitscore" + measurement="bitscore" + // Set maximum e-value for hits to be counted + evalue="0.001" + // Set minimum bitscore for best hit in taxonomy assignment (default = 30) + bitscore="30" // Set minimum percent amino acid similarity for best hit to be counted in taxonomy assignment minID="40" // Set minimum amino acid alignment length for best hit to be counted in taxonomy assignment @@ -122,7 +149,6 @@ params { // DIAMOND taxonomy inference using NCBI taxmap files (can be downloaded using the startup script using the option -t); set to "true" for this to run (ONLY WORKS WITH dbtype="NCBI") ncbitax="false" - // Phylogeny analysis parameters // Customs options for IQ-TREE (Example: "-option1 A -option2 B -option3 C -option4 D") @@ -166,7 +192,7 @@ params { // Manadotory arguments Analyze=false DataCheck=false -// Non-Mandatory Clustering options +// Non-Mandatory options // Cluster nucleotide sequences (ncASVs) ncASV = false // Cluster by aminoacid translations and generate protein-based OTUs (pcASVs) @@ -175,7 +201,8 @@ params { asvMED = false // Generate virus types with MED of ASV sequences aminoMED = false - + // Filtr ASVs + filter = false // Skip options // Skip all Read Processing steps skipReadProcessing=false From fd354888b68bcec6b93371003ae65b6c75dc76d7 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Wed, 22 Sep 2021 09:39:08 -0500 Subject: [PATCH 175/223] revised y axis label for clustering curves --- bin/vAMPirus_DC_Report.Rmd | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/bin/vAMPirus_DC_Report.Rmd b/bin/vAMPirus_DC_Report.Rmd index 5cb0384..327b044 100644 --- a/bin/vAMPirus_DC_Report.Rmd +++ b/bin/vAMPirus_DC_Report.Rmd @@ -374,7 +374,7 @@ nn=read.csv("number_per_percentage_nucl.csv", header=TRUE) #nn=read.csv("number_per_percentage_nucl.csv", header=T) nnp <- plot_ly(nn, x=nn[,1], y=nn[,2], type="scatter", mode = 'lines+markers', marker=list(color='#088da5', line=list(color = 'black', width = .1)), hovertemplate = paste('ID%: %{x}','
    Number of ncASVs: %{y}','')) -nnp <- nnp %>% layout(yaxis = list(title = "Number of nOTU")) +nnp <- nnp %>% layout(yaxis = list(title = "Number of ncASVs")) nnp <- nnp %>% layout(xaxis = list(title = "Clustering ID %")) nnp <- nnp %>% config(toImageButtonOptions=list(format='svg',filename='NuclClustresults', height= 500, width= 800, scale= 1)) nnp @@ -391,7 +391,7 @@ pn=read.csv("number_per_percentage_prot.csv", header=TRUE) #pn=read.csv("number_per_percentage_prot.csv", header=T) pnp <- plot_ly(pn, x=pn[,1], y=pn[,2], type="scatter", mode = 'lines+markers', marker=list(color='#088da5', line=list(color = 'black', width = .1)), hovertemplate = paste('ID%: %{x}','
    Number of pcASVs: %{y}','')) -pnp <- pnp %>% layout(yaxis = list(title = "Number of pOTU")) +pnp <- pnp %>% layout(yaxis = list(title = "Number of pcASVs")) pnp <- pnp %>% layout(xaxis = list(title = "Clustering ID %")) pnp <- pnp %>% config(toImageButtonOptions=list(format='svg',filename='Protclustresults', height= 500, width= 800, scale= 1)) pnp From af04eb504ffb27f36d66c00b45f1c0036c8fac00 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Wed, 22 Sep 2021 09:40:22 -0500 Subject: [PATCH 176/223] added med options to test config --- example_data/conf/test.config | 2 ++ 1 file changed, 2 insertions(+) diff --git a/example_data/conf/test.config b/example_data/conf/test.config index 7149b5d..cbc63e1 100644 --- a/example_data/conf/test.config +++ b/example_data/conf/test.config @@ -22,4 +22,6 @@ params { aminoC=5 asvC=5 stats = true + aminoMED = true + asvMED = true } From f4e787147843375e0ef78082c679831b1f244c2d Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Wed, 22 Sep 2021 10:32:40 -0500 Subject: [PATCH 177/223] Added ASV filtering options and help info --- docs/HelpDocumentation.md | 36 ++++++++++++++++++++++++++++++++++-- vampirus.config | 6 +++--- 2 files changed, 37 insertions(+), 5 deletions(-) diff --git a/docs/HelpDocumentation.md b/docs/HelpDocumentation.md index 689d892..5c085c8 100644 --- a/docs/HelpDocumentation.md +++ b/docs/HelpDocumentation.md @@ -319,7 +319,7 @@ After running the above you should now have Nextflow installed. Now, build the S then test the Analyze pipeline with: - ./nextflow run vAMPirus.nf -c vampirus.config -profile singularity,test --Analyze --ncASV --pcASV --stats + ./nextflow run vAMPirus.nf -c vampirus.config -profile singularity,test --Analyze --ncASV --pcASV --asvMED --aminoMED --stats Please check out http://sourabhbajaj.com/mac-setup/Vagrant/README.html and https://www.vagrantup.com/docs/providers/virtualbox for understanding how to use Vagrant commands like "halt", "suspend" or "reload" @@ -1021,7 +1021,7 @@ This is where the DataCheck report is very useful, you can review the report and The goal of vAMPirus was to make it easy for the user to analyze their data is many different ways to potentially reveal patterns that would have been missed if limited to one method/pipeline. -A major and sometimes difficult step in analyzing virus amplicon sequence data is deciding the method to use for identifying or defining different viral "species" in the data. To aid this process, vAMPirus has the DataCheck mode discussed above and has several different options for sequence clustering/analysis for the user to decide between. +A major, and sometimes difficult, step in analyzing virus amplicon sequence data is deciding the method to use for identifying or defining different viral "species" in the data. To aid this process, vAMPirus has the DataCheck mode discussed above and has several different options for sequence clustering/analysis for the user to decide between. vAMPirus relies on vsearch using the UNOISE3 algorithm to generate Amplicon Sequencing Variants (ASVs) from dereplicated amplicon reads. ASVs are always generated by default and there are two parameters that the user can specify either in the launch command or by editing the configuration file at lines 45-49: @@ -1037,6 +1037,38 @@ Launch command to produce only ASV-related analyses: nextflow run vAMPirus.nf -c vampirus.config -profile [conda|singularity] --Analyze --stats --skipAminoTyping +### ASV filtering + +New to version 2 you can now filter ASVs to remove sequences that belong to taxonomic groups that are not of interest for a given run. + +A great example of when this feature is useful is Prodinger et al. 2020 (https://www.mdpi.com/2076-2607/8/4/506). In this study they looked to amplify and analyze Mimiviridae polB sequences, however, polB is also found in cellular genomes like bacteria. In this case, Prodinger et al. looked to avoid including any bacterial polB in their final results and thus used a filtering step to remove microbial sequences. The ASV filtering feature can be used to do exactly this type of filtering where you provide paths to a "filter database" containing sequences belonging to non-target groups (e.g. microbial polB) and a "keep database" containing sequences belonging to the target group (e.g. Mimiviridae polB). Any ASVs that match non-target sequences will then be filtered from the ASV file prior to running the DataCheck or Analyze pipeline. + +Here are the options stored within the configuration file: + + // ASV filtering parameters - You can set the filtering to run with the command --filter + + // Path to database containing sequences that if ASVs match to, are then removed prior to any analyses + filtDB="" + // Path to database containing sequences that if ASVs match to, are kept for final ASV file to be used in subsequent analyses + keepDB="" + // Keep any sequences without hits - for yes, set keepnohit to ="true" + keepnohit="true" + + //Parameters for diamond command for filtering + + // Set minimum percent amino acid similarity for best hit to be counted in taxonomy assignment + filtminID="80" + // Set minimum amino acid alignment length for best hit to be counted in taxonomy assignment + filtminaln="30" + // Set sensitivity parameters for DIAMOND aligner (read more here: https://github.com/bbuchfink/diamond/wiki; default = ultra-sensitive) + filtsensitivity="ultra-sensitive" + // Set the max e-value for best hit to be recorded + filtevalue="0.001" + + + +### Clustering options + Now, onto clustering ASVs into clustered ASVs (cASVs). vAMPirus is able to use two different techniques for generating cASVs for the user: diff --git a/vampirus.config b/vampirus.config index 8c34321..0e0deae 100644 --- a/vampirus.config +++ b/vampirus.config @@ -75,11 +75,11 @@ params { // Minimum length of amino acid translation to be considered during protein clustered ASV (pcASV) generation. Recommended to put this at the expected amino acid sequence length based on your maximum read length (e.g. if maxLen="420", then minAA should be 420/3 so 140) minAA="140" - // Filtering parameters - You can set the filtering to run with the command --filter + // ASV filtering parameters - You can set the filtering to run with the command --filter - // Path to database + // Path to database containing sequences that if ASVs match, are then removed prior to any analyses filtDB="" - // Path to database of + // Path to database containing sequences that if ASVs match to, are kept for final ASV file to be used in susequent analyses keepDB="" // Keep any sequences without hits - for yes, set keepnohit to ="true" keepnohit="true" From 589672859ab1cfb7da34d855a40a5d2e8854c875 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Wed, 22 Sep 2021 10:48:58 -0500 Subject: [PATCH 178/223] removed MED boxplot --- bin/vAMPirus_DC_Report.Rmd | 15 --------------- 1 file changed, 15 deletions(-) diff --git a/bin/vAMPirus_DC_Report.Rmd b/bin/vAMPirus_DC_Report.Rmd index 327b044..50d6ec1 100644 --- a/bin/vAMPirus_DC_Report.Rmd +++ b/bin/vAMPirus_DC_Report.Rmd @@ -451,21 +451,6 @@ med_amino_csv=read.csv(paste(params$projtag, sep="", "_AminoType_entropy_breakdo paged_table(med_amino_csv, options = list(rows.print = 10)) ```
    - -### ASVs vs AminoTypes - -
    - -```{r med_comparison, echo=FALSE} -med_asv=read.delim(paste(params$projtag, sep="","_ASVs_Aligned_informativeonly.fasta-ENTROPY"), sep = ",", header=TRUE) -med_amino=read.delim(paste(params$projtag, sep="","_AminoTypes_Aligned_informativeonly.fasta-ENTROPY"), sep = ",", header=TRUE) - -mplotbox <- plot_ly(med_asv, y=med_asv$Shannons_Entropy, type="box", boxmean='sd', boxpoints="outlier", jitter=0.1, pointpos=0, name="ASVs") -mplotbox<- mplotbox %>% add_trace(med_amino, y=med_amino$Shannons_Entropy, type="box", boxmean='sd', boxpoints="outliers", jitter=0.1, pointpos=0, name="AminoTypes") -mplotbox<- mplotbox %>% layout(legend = list(y=.5)) -mplotbox<- mplotbox %>% config(toImageButtonOptions=list(format='svg',filename='ASV_vs_AminoTypes', height= 500, width= 800, scale= 1), displaylogo = FALSE) -mplotbox -```


    From d512df36e734f8e2d888853743c71e819b77004b Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Wed, 22 Sep 2021 13:45:29 -0500 Subject: [PATCH 179/223] added more values for med table in datacheck --- vAMPirus.nf | 53 +++++++++++++++++++++++++++++++++++++++++++++++++++++ 1 file changed, 53 insertions(+) diff --git a/vAMPirus.nf b/vAMPirus.nf index 4170216..3a92691 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1234,9 +1234,36 @@ if (params.DataCheck || params.Analyze) { if [[ \$(echo ""\$x" > 1.0"|bc -l) -eq 1 ]]; then echo dope >> above-1.0-.list fi + if [[ \$(echo ""\$x" > 1.1"|bc -l) -eq 1 ]]; + then echo dope >> above-1.1-.list + fi + if [[ \$(echo ""\$x" > 1.2"|bc -l) -eq 1 ]]; + then echo dope >> above-1.2-.list + fi + if [[ \$(echo ""\$x" > 1.3"|bc -l) -eq 1 ]]; + then echo dope >> above-1.3-.list + fi + if [[ \$(echo ""\$x" > 1.4"|bc -l) -eq 1 ]]; + then echo dope >> above-1.4-.list + fi if [[ \$(echo ""\$x" > 1.5"|bc -l) -eq 1 ]]; then echo dope >> above-1.5-.list fi + if [[ \$(echo ""\$x" > 1.6"|bc -l) -eq 1 ]]; + then echo dope >> above-1.6-.list + fi + if [[ \$(echo ""\$x" > 1.7"|bc -l) -eq 1 ]]; + then echo dope >> above-1.7-.list + fi + if [[ \$(echo ""\$x" > 1.8"|bc -l) -eq 1 ]]; + then echo dope >> above-1.8-.list + fi + if [[ \$(echo ""\$x" > 1.9"|bc -l) -eq 1 ]]; + then echo dope >> above-1.9-.list + fi + if [[ \$(echo ""\$x" > 2.0"|bc -l) -eq 1 ]]; + then echo dope >> above-2.0-.list + fi done echo "Entropy,Peaks_above" >> ${params.projtag}_ASV_entropy_breakdown.csv for z in above*.list; @@ -1318,9 +1345,35 @@ if (params.DataCheck || params.Analyze) { if [[ \$(echo ""\$x" > 1.0"|bc -l) -eq 1 ]]; then echo dope >> above-1.0-.list fi + if [[ \$(echo ""\$x" > 1.1"|bc -l) -eq 1 ]]; + then echo dope >> above-1.1-.list + fi + if [[ \$(echo ""\$x" > 1.2"|bc -l) -eq 1 ]]; + then echo dope >> above-1.2-.list + fi + if [[ \$(echo ""\$x" > 1.3"|bc -l) -eq 1 ]]; + then echo dope >> above-1.3-.list + fi + if [[ \$(echo ""\$x" > 1.4"|bc -l) -eq 1 ]]; + then echo dope >> above-1.4-.list if [[ \$(echo ""\$x" > 1.5"|bc -l) -eq 1 ]]; then echo dope >> above-1.5-.list fi + if [[ \$(echo ""\$x" > 1.6"|bc -l) -eq 1 ]]; + then echo dope >> above-1.6-.list + fi + if [[ \$(echo ""\$x" > 1.7"|bc -l) -eq 1 ]]; + then echo dope >> above-1.7-.list + fi + if [[ \$(echo ""\$x" > 1.8"|bc -l) -eq 1 ]]; + then echo dope >> above-1.8-.list + fi + if [[ \$(echo ""\$x" > 1.9"|bc -l) -eq 1 ]]; + then echo dope >> above-1.9-.list + fi + if [[ \$(echo ""\$x" > 2.0"|bc -l) -eq 1 ]]; + then echo dope >> above-2.0-.list + fi done echo "Entropy,Peaks_above" >> ${params.projtag}_AminoType_entropy_breakdown.csv for z in above*.list; From 86d636279e7c66705f0839ed8db9d8d3a33819b1 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Wed, 22 Sep 2021 14:43:54 -0500 Subject: [PATCH 180/223] fixed for loop in filtering --- vAMPirus.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 3a92691..59cde39 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -971,7 +971,7 @@ if (params.DataCheck || params.Analyze) { diamond blastx -q ${asv} -d combodatabase.fasta -p ${task.cpus} --id ${params.filtminID} -l ${params.filtminaln} -e ${params.filtevalue} --${params.filtsensitivity} -o ${params.projtag}_diamondfilter.out -f 6 qseqid qlen sseqid qstart qend qseq sseq length qframe evalue bitscore pident --max-target-seqs 1 --max-hsps 1 #get asvs grep ">" ${asv} | awk -F ">" '{print \$2}' > asv.list - for x in $(cat asv.list) + for x in \$(cat asv.list) do #check for a hit if [[ \$(grep -c "\$x" ${params.projtag}_diamondfilter.out) -eq 1 ]] then #check if hit is to filter From 2bd9ab2bc3e2949d39f1f267dbd0f6f0b2b41b64 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Wed, 22 Sep 2021 14:52:59 -0500 Subject: [PATCH 181/223] fixed if statement for fitering --- vAMPirus.nf | 4 ++-- vampirus.config | 2 +- 2 files changed, 3 insertions(+), 3 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 59cde39..899cef1 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -930,7 +930,7 @@ if (params.DataCheck || params.Analyze) { label 'norm_cpus' - publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/FilterCont", mode: "copy", overwrite: true, pattern: '*.fasta' + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/FilterCont", mode: "copy", overwrite: true input: file(asv) from asvforfilt @@ -1009,7 +1009,7 @@ if (params.DataCheck || params.Analyze) { } else { asvforfilt - .into{ reads_vsearch5_ch, asv_med, nucl2aa, asvsforAminotyping, asvfastaforcounts, asvaminocheck } + .into{ reads_vsearch5_ch; asv_med; nucl2aa; asvsforAminotyping; asvfastaforcounts; asvaminocheck } } if (params.DataCheck) { diff --git a/vampirus.config b/vampirus.config index 0e0deae..80876ea 100644 --- a/vampirus.config +++ b/vampirus.config @@ -201,7 +201,7 @@ params { asvMED = false // Generate virus types with MED of ASV sequences aminoMED = false - // Filtr ASVs + // Filter ASVs filter = false // Skip options // Skip all Read Processing steps From 6dd92a7341ec0a2e244122ba12f073ff0e710c55 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Wed, 22 Sep 2021 15:06:08 -0500 Subject: [PATCH 182/223] fixed aminomed datacheck --- vAMPirus.nf | 84 ++++++++++++++++++++++++++--------------------------- 1 file changed, 42 insertions(+), 42 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 899cef1..66c520a 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1201,67 +1201,67 @@ if (params.DataCheck || params.Analyze) { awk '{print \$2}' ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY >> tmp_value.list for x in \$(cat tmp_value.list) do echo "\$x" - if [[ \$(echo ""\$x" > 0.0"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 0.0"|bc -l) -eq 1 ]]; then echo dope >> above-0.0-.list fi - if [[ \$(echo ""\$x" > 0.1"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 0.1"|bc -l) -eq 1 ]]; then echo dope >> above-0.1-.list fi - if [[ \$(echo ""\$x" > 0.2"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 0.2"|bc -l) -eq 1 ]]; then echo dope >> above-0.2-.list fi - if [[ \$(echo ""\$x" > 0.3"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 0.3"|bc -l) -eq 1 ]]; then echo dope >> above-0.3-.list fi - if [[ \$(echo ""\$x" > 0.4"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 0.4"|bc -l) -eq 1 ]]; then echo dope >> above-0.4-.list fi - if [[ \$(echo ""\$x" > 0.5"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 0.5"|bc -l) -eq 1 ]]; then echo dope >> above-0.5-.list fi - if [[ \$(echo ""\$x" > 0.6"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 0.6"|bc -l) -eq 1 ]]; then echo dope >> above-0.6-.list fi - if [[ \$(echo ""\$x" > 0.7"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 0.7"|bc -l) -eq 1 ]]; then echo dope >> above-0.7-.list fi - if [[ \$(echo ""\$x" > 0.8"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 0.8"|bc -l) -eq 1 ]]; then echo dope >> above-0.8-.list fi - if [[ \$(echo ""\$x" > 0.9"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 0.9"|bc -l) -eq 1 ]]; then echo dope >> above-0.9-.list fi - if [[ \$(echo ""\$x" > 1.0"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 1.0"|bc -l) -eq 1 ]]; then echo dope >> above-1.0-.list fi - if [[ \$(echo ""\$x" > 1.1"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 1.1"|bc -l) -eq 1 ]]; then echo dope >> above-1.1-.list fi - if [[ \$(echo ""\$x" > 1.2"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 1.2"|bc -l) -eq 1 ]]; then echo dope >> above-1.2-.list fi - if [[ \$(echo ""\$x" > 1.3"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 1.3"|bc -l) -eq 1 ]]; then echo dope >> above-1.3-.list fi - if [[ \$(echo ""\$x" > 1.4"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 1.4"|bc -l) -eq 1 ]]; then echo dope >> above-1.4-.list fi - if [[ \$(echo ""\$x" > 1.5"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 1.5"|bc -l) -eq 1 ]]; then echo dope >> above-1.5-.list fi - if [[ \$(echo ""\$x" > 1.6"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 1.6"|bc -l) -eq 1 ]]; then echo dope >> above-1.6-.list fi - if [[ \$(echo ""\$x" > 1.7"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 1.7"|bc -l) -eq 1 ]]; then echo dope >> above-1.7-.list fi - if [[ \$(echo ""\$x" > 1.8"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 1.8"|bc -l) -eq 1 ]]; then echo dope >> above-1.8-.list fi - if [[ \$(echo ""\$x" > 1.9"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 1.9"|bc -l) -eq 1 ]]; then echo dope >> above-1.9-.list fi - if [[ \$(echo ""\$x" > 2.0"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 2.0"|bc -l) -eq 1 ]]; then echo dope >> above-2.0-.list fi done @@ -1312,66 +1312,66 @@ if (params.DataCheck || params.Analyze) { awk '{print \$2}' ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta-ENTROPY >> tmp_value.list for x in \$(cat tmp_value.list) do echo "\$x" - if [[ \$(echo ""\$x" > 0.0"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 0.0"|bc -l) -eq 1 ]]; then echo dope >> above-0.0-.list fi - if [[ \$(echo ""\$x" > 0.1"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 0.1"|bc -l) -eq 1 ]]; then echo dope >> above-0.1-.list fi - if [[ \$(echo ""\$x" > 0.2"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 0.2"|bc -l) -eq 1 ]]; then echo dope >> above-0.2-.list fi - if [[ \$(echo ""\$x" > 0.3"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 0.3"|bc -l) -eq 1 ]]; then echo dope >> above-0.3-.list fi - if [[ \$(echo ""\$x" > 0.4"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 0.4"|bc -l) -eq 1 ]]; then echo dope >> above-0.4-.list fi - if [[ \$(echo ""\$x" > 0.5"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 0.5"|bc -l) -eq 1 ]]; then echo dope >> above-0.5-.list fi - if [[ \$(echo ""\$x" > 0.6"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 0.6"|bc -l) -eq 1 ]]; then echo dope >> above-0.6-.list fi - if [[ \$(echo ""\$x" > 0.7"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 0.7"|bc -l) -eq 1 ]]; then echo dope >> above-0.7-.list fi - if [[ \$(echo ""\$x" > 0.8"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 0.8"|bc -l) -eq 1 ]]; then echo dope >> above-0.8-.list fi - if [[ \$(echo ""\$x" > 0.9"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 0.9"|bc -l) -eq 1 ]]; then echo dope >> above-0.9-.list fi - if [[ \$(echo ""\$x" > 1.0"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 1.0"|bc -l) -eq 1 ]]; then echo dope >> above-1.0-.list fi - if [[ \$(echo ""\$x" > 1.1"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 1.1"|bc -l) -eq 1 ]]; then echo dope >> above-1.1-.list fi - if [[ \$(echo ""\$x" > 1.2"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 1.2"|bc -l) -eq 1 ]]; then echo dope >> above-1.2-.list fi - if [[ \$(echo ""\$x" > 1.3"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 1.3"|bc -l) -eq 1 ]]; then echo dope >> above-1.3-.list fi - if [[ \$(echo ""\$x" > 1.4"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 1.4"|bc -l) -eq 1 ]]; then echo dope >> above-1.4-.list - if [[ \$(echo ""\$x" > 1.5"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 1.5"|bc -l) -eq 1 ]]; then echo dope >> above-1.5-.list fi - if [[ \$(echo ""\$x" > 1.6"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 1.6"|bc -l) -eq 1 ]]; then echo dope >> above-1.6-.list fi - if [[ \$(echo ""\$x" > 1.7"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 1.7"|bc -l) -eq 1 ]]; then echo dope >> above-1.7-.list fi - if [[ \$(echo ""\$x" > 1.8"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 1.8"|bc -l) -eq 1 ]]; then echo dope >> above-1.8-.list fi - if [[ \$(echo ""\$x" > 1.9"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 1.9"|bc -l) -eq 1 ]]; then echo dope >> above-1.9-.list fi - if [[ \$(echo ""\$x" > 2.0"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x" > 2.0"|bc -l) -eq 1 ]]; then echo dope >> above-2.0-.list fi done From 07af4f395448ce745553cbcd31e75425f254e296 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Wed, 22 Sep 2021 15:15:15 -0500 Subject: [PATCH 183/223] test --- vAMPirus.nf | 84 ++++++++++++++++++++++++++--------------------------- 1 file changed, 42 insertions(+), 42 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 66c520a..899cef1 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1201,67 +1201,67 @@ if (params.DataCheck || params.Analyze) { awk '{print \$2}' ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY >> tmp_value.list for x in \$(cat tmp_value.list) do echo "\$x" - if [[ \$(echo "\$x" > 0.0"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 0.0"|bc -l) -eq 1 ]]; then echo dope >> above-0.0-.list fi - if [[ \$(echo "\$x" > 0.1"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 0.1"|bc -l) -eq 1 ]]; then echo dope >> above-0.1-.list fi - if [[ \$(echo "\$x" > 0.2"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 0.2"|bc -l) -eq 1 ]]; then echo dope >> above-0.2-.list fi - if [[ \$(echo "\$x" > 0.3"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 0.3"|bc -l) -eq 1 ]]; then echo dope >> above-0.3-.list fi - if [[ \$(echo "\$x" > 0.4"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 0.4"|bc -l) -eq 1 ]]; then echo dope >> above-0.4-.list fi - if [[ \$(echo "\$x" > 0.5"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 0.5"|bc -l) -eq 1 ]]; then echo dope >> above-0.5-.list fi - if [[ \$(echo "\$x" > 0.6"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 0.6"|bc -l) -eq 1 ]]; then echo dope >> above-0.6-.list fi - if [[ \$(echo "\$x" > 0.7"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 0.7"|bc -l) -eq 1 ]]; then echo dope >> above-0.7-.list fi - if [[ \$(echo "\$x" > 0.8"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 0.8"|bc -l) -eq 1 ]]; then echo dope >> above-0.8-.list fi - if [[ \$(echo "\$x" > 0.9"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 0.9"|bc -l) -eq 1 ]]; then echo dope >> above-0.9-.list fi - if [[ \$(echo "\$x" > 1.0"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 1.0"|bc -l) -eq 1 ]]; then echo dope >> above-1.0-.list fi - if [[ \$(echo "\$x" > 1.1"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 1.1"|bc -l) -eq 1 ]]; then echo dope >> above-1.1-.list fi - if [[ \$(echo "\$x" > 1.2"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 1.2"|bc -l) -eq 1 ]]; then echo dope >> above-1.2-.list fi - if [[ \$(echo "\$x" > 1.3"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 1.3"|bc -l) -eq 1 ]]; then echo dope >> above-1.3-.list fi - if [[ \$(echo "\$x" > 1.4"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 1.4"|bc -l) -eq 1 ]]; then echo dope >> above-1.4-.list fi - if [[ \$(echo "\$x" > 1.5"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 1.5"|bc -l) -eq 1 ]]; then echo dope >> above-1.5-.list fi - if [[ \$(echo "\$x" > 1.6"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 1.6"|bc -l) -eq 1 ]]; then echo dope >> above-1.6-.list fi - if [[ \$(echo "\$x" > 1.7"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 1.7"|bc -l) -eq 1 ]]; then echo dope >> above-1.7-.list fi - if [[ \$(echo "\$x" > 1.8"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 1.8"|bc -l) -eq 1 ]]; then echo dope >> above-1.8-.list fi - if [[ \$(echo "\$x" > 1.9"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 1.9"|bc -l) -eq 1 ]]; then echo dope >> above-1.9-.list fi - if [[ \$(echo "\$x" > 2.0"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 2.0"|bc -l) -eq 1 ]]; then echo dope >> above-2.0-.list fi done @@ -1312,66 +1312,66 @@ if (params.DataCheck || params.Analyze) { awk '{print \$2}' ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta-ENTROPY >> tmp_value.list for x in \$(cat tmp_value.list) do echo "\$x" - if [[ \$(echo "\$x" > 0.0"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 0.0"|bc -l) -eq 1 ]]; then echo dope >> above-0.0-.list fi - if [[ \$(echo "\$x" > 0.1"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 0.1"|bc -l) -eq 1 ]]; then echo dope >> above-0.1-.list fi - if [[ \$(echo "\$x" > 0.2"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 0.2"|bc -l) -eq 1 ]]; then echo dope >> above-0.2-.list fi - if [[ \$(echo "\$x" > 0.3"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 0.3"|bc -l) -eq 1 ]]; then echo dope >> above-0.3-.list fi - if [[ \$(echo "\$x" > 0.4"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 0.4"|bc -l) -eq 1 ]]; then echo dope >> above-0.4-.list fi - if [[ \$(echo "\$x" > 0.5"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 0.5"|bc -l) -eq 1 ]]; then echo dope >> above-0.5-.list fi - if [[ \$(echo "\$x" > 0.6"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 0.6"|bc -l) -eq 1 ]]; then echo dope >> above-0.6-.list fi - if [[ \$(echo "\$x" > 0.7"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 0.7"|bc -l) -eq 1 ]]; then echo dope >> above-0.7-.list fi - if [[ \$(echo "\$x" > 0.8"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 0.8"|bc -l) -eq 1 ]]; then echo dope >> above-0.8-.list fi - if [[ \$(echo "\$x" > 0.9"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 0.9"|bc -l) -eq 1 ]]; then echo dope >> above-0.9-.list fi - if [[ \$(echo "\$x" > 1.0"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 1.0"|bc -l) -eq 1 ]]; then echo dope >> above-1.0-.list fi - if [[ \$(echo "\$x" > 1.1"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 1.1"|bc -l) -eq 1 ]]; then echo dope >> above-1.1-.list fi - if [[ \$(echo "\$x" > 1.2"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 1.2"|bc -l) -eq 1 ]]; then echo dope >> above-1.2-.list fi - if [[ \$(echo "\$x" > 1.3"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 1.3"|bc -l) -eq 1 ]]; then echo dope >> above-1.3-.list fi - if [[ \$(echo "\$x" > 1.4"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 1.4"|bc -l) -eq 1 ]]; then echo dope >> above-1.4-.list - if [[ \$(echo "\$x" > 1.5"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 1.5"|bc -l) -eq 1 ]]; then echo dope >> above-1.5-.list fi - if [[ \$(echo "\$x" > 1.6"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 1.6"|bc -l) -eq 1 ]]; then echo dope >> above-1.6-.list fi - if [[ \$(echo "\$x" > 1.7"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 1.7"|bc -l) -eq 1 ]]; then echo dope >> above-1.7-.list fi - if [[ \$(echo "\$x" > 1.8"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 1.8"|bc -l) -eq 1 ]]; then echo dope >> above-1.8-.list fi - if [[ \$(echo "\$x" > 1.9"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 1.9"|bc -l) -eq 1 ]]; then echo dope >> above-1.9-.list fi - if [[ \$(echo "\$x" > 2.0"|bc -l) -eq 1 ]]; + if [[ \$(echo ""\$x" > 2.0"|bc -l) -eq 1 ]]; then echo dope >> above-2.0-.list fi done From 41c79337cce4696b933404a77fe27a56101eeb96 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Wed, 22 Sep 2021 15:21:03 -0500 Subject: [PATCH 184/223] fixe quotes --- vAMPirus.nf | 84 ++++++++++++++++++++++++++--------------------------- 1 file changed, 42 insertions(+), 42 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 899cef1..3a874c3 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1201,67 +1201,67 @@ if (params.DataCheck || params.Analyze) { awk '{print \$2}' ${params.projtag}_ASVs_Aligned_informativeonly.fasta-ENTROPY >> tmp_value.list for x in \$(cat tmp_value.list) do echo "\$x" - if [[ \$(echo ""\$x" > 0.0"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 0.0"|bc -l) -eq 1 ]]; then echo dope >> above-0.0-.list fi - if [[ \$(echo ""\$x" > 0.1"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 0.1"|bc -l) -eq 1 ]]; then echo dope >> above-0.1-.list fi - if [[ \$(echo ""\$x" > 0.2"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 0.2"|bc -l) -eq 1 ]]; then echo dope >> above-0.2-.list fi - if [[ \$(echo ""\$x" > 0.3"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 0.3"|bc -l) -eq 1 ]]; then echo dope >> above-0.3-.list fi - if [[ \$(echo ""\$x" > 0.4"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 0.4"|bc -l) -eq 1 ]]; then echo dope >> above-0.4-.list fi - if [[ \$(echo ""\$x" > 0.5"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 0.5"|bc -l) -eq 1 ]]; then echo dope >> above-0.5-.list fi - if [[ \$(echo ""\$x" > 0.6"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 0.6"|bc -l) -eq 1 ]]; then echo dope >> above-0.6-.list fi - if [[ \$(echo ""\$x" > 0.7"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 0.7"|bc -l) -eq 1 ]]; then echo dope >> above-0.7-.list fi - if [[ \$(echo ""\$x" > 0.8"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 0.8"|bc -l) -eq 1 ]]; then echo dope >> above-0.8-.list fi - if [[ \$(echo ""\$x" > 0.9"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 0.9"|bc -l) -eq 1 ]]; then echo dope >> above-0.9-.list fi - if [[ \$(echo ""\$x" > 1.0"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 1.0"|bc -l) -eq 1 ]]; then echo dope >> above-1.0-.list fi - if [[ \$(echo ""\$x" > 1.1"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 1.1"|bc -l) -eq 1 ]]; then echo dope >> above-1.1-.list fi - if [[ \$(echo ""\$x" > 1.2"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 1.2"|bc -l) -eq 1 ]]; then echo dope >> above-1.2-.list fi - if [[ \$(echo ""\$x" > 1.3"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 1.3"|bc -l) -eq 1 ]]; then echo dope >> above-1.3-.list fi - if [[ \$(echo ""\$x" > 1.4"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 1.4"|bc -l) -eq 1 ]]; then echo dope >> above-1.4-.list fi - if [[ \$(echo ""\$x" > 1.5"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 1.5"|bc -l) -eq 1 ]]; then echo dope >> above-1.5-.list fi - if [[ \$(echo ""\$x" > 1.6"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 1.6"|bc -l) -eq 1 ]]; then echo dope >> above-1.6-.list fi - if [[ \$(echo ""\$x" > 1.7"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 1.7"|bc -l) -eq 1 ]]; then echo dope >> above-1.7-.list fi - if [[ \$(echo ""\$x" > 1.8"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 1.8"|bc -l) -eq 1 ]]; then echo dope >> above-1.8-.list fi - if [[ \$(echo ""\$x" > 1.9"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 1.9"|bc -l) -eq 1 ]]; then echo dope >> above-1.9-.list fi - if [[ \$(echo ""\$x" > 2.0"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 2.0"|bc -l) -eq 1 ]]; then echo dope >> above-2.0-.list fi done @@ -1312,66 +1312,66 @@ if (params.DataCheck || params.Analyze) { awk '{print \$2}' ${params.projtag}_AminoTypes_Aligned_informativeonly.fasta-ENTROPY >> tmp_value.list for x in \$(cat tmp_value.list) do echo "\$x" - if [[ \$(echo ""\$x" > 0.0"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 0.0"|bc -l) -eq 1 ]]; then echo dope >> above-0.0-.list fi - if [[ \$(echo ""\$x" > 0.1"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 0.1"|bc -l) -eq 1 ]]; then echo dope >> above-0.1-.list fi - if [[ \$(echo ""\$x" > 0.2"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 0.2"|bc -l) -eq 1 ]]; then echo dope >> above-0.2-.list fi - if [[ \$(echo ""\$x" > 0.3"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 0.3"|bc -l) -eq 1 ]]; then echo dope >> above-0.3-.list fi - if [[ \$(echo ""\$x" > 0.4"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 0.4"|bc -l) -eq 1 ]]; then echo dope >> above-0.4-.list fi - if [[ \$(echo ""\$x" > 0.5"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 0.5"|bc -l) -eq 1 ]]; then echo dope >> above-0.5-.list fi - if [[ \$(echo ""\$x" > 0.6"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 0.6"|bc -l) -eq 1 ]]; then echo dope >> above-0.6-.list fi - if [[ \$(echo ""\$x" > 0.7"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 0.7"|bc -l) -eq 1 ]]; then echo dope >> above-0.7-.list fi - if [[ \$(echo ""\$x" > 0.8"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 0.8"|bc -l) -eq 1 ]]; then echo dope >> above-0.8-.list fi - if [[ \$(echo ""\$x" > 0.9"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 0.9"|bc -l) -eq 1 ]]; then echo dope >> above-0.9-.list fi - if [[ \$(echo ""\$x" > 1.0"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 1.0"|bc -l) -eq 1 ]]; then echo dope >> above-1.0-.list fi - if [[ \$(echo ""\$x" > 1.1"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 1.1"|bc -l) -eq 1 ]]; then echo dope >> above-1.1-.list fi - if [[ \$(echo ""\$x" > 1.2"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 1.2"|bc -l) -eq 1 ]]; then echo dope >> above-1.2-.list fi - if [[ \$(echo ""\$x" > 1.3"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 1.3"|bc -l) -eq 1 ]]; then echo dope >> above-1.3-.list fi - if [[ \$(echo ""\$x" > 1.4"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 1.4"|bc -l) -eq 1 ]]; then echo dope >> above-1.4-.list - if [[ \$(echo ""\$x" > 1.5"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 1.5"|bc -l) -eq 1 ]]; then echo dope >> above-1.5-.list fi - if [[ \$(echo ""\$x" > 1.6"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 1.6"|bc -l) -eq 1 ]]; then echo dope >> above-1.6-.list fi - if [[ \$(echo ""\$x" > 1.7"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 1.7"|bc -l) -eq 1 ]]; then echo dope >> above-1.7-.list fi - if [[ \$(echo ""\$x" > 1.8"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 1.8"|bc -l) -eq 1 ]]; then echo dope >> above-1.8-.list fi - if [[ \$(echo ""\$x" > 1.9"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 1.9"|bc -l) -eq 1 ]]; then echo dope >> above-1.9-.list fi - if [[ \$(echo ""\$x" > 2.0"|bc -l) -eq 1 ]]; + if [[ \$(echo "\$x > 2.0"|bc -l) -eq 1 ]]; then echo dope >> above-2.0-.list fi done From fbdcc4dd8925be3e84ffdcba0d7c771bd049d7f9 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Wed, 22 Sep 2021 15:26:01 -0500 Subject: [PATCH 185/223] fixed for loop --- vAMPirus.nf | 1 + 1 file changed, 1 insertion(+) diff --git a/vAMPirus.nf b/vAMPirus.nf index 3a874c3..4ac928a 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1356,6 +1356,7 @@ if (params.DataCheck || params.Analyze) { fi if [[ \$(echo "\$x > 1.4"|bc -l) -eq 1 ]]; then echo dope >> above-1.4-.list + fi if [[ \$(echo "\$x > 1.5"|bc -l) -eq 1 ]]; then echo dope >> above-1.5-.list fi From 5e54ca2634d83a52d8a295dc1112d80284f12040 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Wed, 22 Sep 2021 15:52:39 -0500 Subject: [PATCH 186/223] updated ggtree in yaml --- vampirus_env.yml | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vampirus_env.yml b/vampirus_env.yml index cb10503..f7222a3 100644 --- a/vampirus_env.yml +++ b/vampirus_env.yml @@ -32,7 +32,7 @@ dependencies: - r-tidyverse=1.3.0 - r-cowplot=1.0.0 - r-scales=1.1.1 - - bioconductor-ggtree=2.2.1 + - bioconductor-ggtree=3.0.1 - bioconductor-biocparallel=1.24.0 - pigz=2.4 - r-biocmanager=1.30.10 From 4eeaa440c5acc06ae218eacd486ed379530ff6c8 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Wed, 22 Sep 2021 16:42:59 -0500 Subject: [PATCH 187/223] ggtree info changes --- bin/vAMPirus_DC_Report.Rmd | 4 +++- vampirus_env.yml | 2 +- 2 files changed, 4 insertions(+), 2 deletions(-) diff --git a/bin/vAMPirus_DC_Report.Rmd b/bin/vAMPirus_DC_Report.Rmd index 50d6ec1..728e770 100644 --- a/bin/vAMPirus_DC_Report.Rmd +++ b/bin/vAMPirus_DC_Report.Rmd @@ -36,7 +36,6 @@ library(tidyverse) library(scales) library(cowplot) library(dplyr) -library(ggtree) library(plotly) #library(BiocParallel) library(knitr) @@ -460,3 +459,6 @@ paged_table(med_amino_csv, options = list(rows.print = 10))


    +
    +
    +
    diff --git a/vampirus_env.yml b/vampirus_env.yml index f7222a3..cb10503 100644 --- a/vampirus_env.yml +++ b/vampirus_env.yml @@ -32,7 +32,7 @@ dependencies: - r-tidyverse=1.3.0 - r-cowplot=1.0.0 - r-scales=1.1.1 - - bioconductor-ggtree=3.0.1 + - bioconductor-ggtree=2.2.1 - bioconductor-biocparallel=1.24.0 - pigz=2.4 - r-biocmanager=1.30.10 From 8121ca88b07d43d7d16773e3df5203b934904b40 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Wed, 22 Sep 2021 16:48:40 -0500 Subject: [PATCH 188/223] fixed measure if statement --- vAMPirus.nf | 40 ++++++++++++++++++++-------------------- 1 file changed, 20 insertions(+), 20 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 4ac928a..1991bac 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1509,9 +1509,9 @@ if (params.DataCheck || params.Analyze) { """ cp ${params.vampdir}/bin/rename_seq.py . virdb=${params.dbdir}/${params.dbname} - if [[ ${params.measurement} == "bitscore"]] + if [[ ${params.measurement} == "bitscore" ]] then measure="--min-score ${params.bitscore}" - elif [[ ${params.measurement} == "evalue"]] + elif [[ ${params.measurement} == "evalue" ]] then measure="-e ${params.evalue}" else measure="--min-score ${params.bitscore}" fi @@ -1663,9 +1663,9 @@ if (params.DataCheck || params.Analyze) { """ cp ${params.vampdir}/bin/rename_seq.py . virdb=${params.dbdir}/${params.dbname} - if [[ ${params.measurement} == "bitscore"]] + if [[ ${params.measurement} == "bitscore" ]] then measure="--min-score ${params.bitscore}" - elif [[ ${params.measurement} == "evalue"]] + elif [[ ${params.measurement} == "evalue" ]] then measure="-e ${params.evalue}" else measure="--min-score ${params.bitscore}" fi @@ -1904,9 +1904,9 @@ if (params.DataCheck || params.Analyze) { """ cp ${params.vampdir}/bin/rename_seq.py . virdb=${params.dbdir}/${params.dbname} - if [[ ${params.measurement} == "bitscore"]] + if [[ ${params.measurement} == "bitscore" ]] then measure="--min-score ${params.bitscore}" - elif [[ ${params.measurement} == "evalue"]] + elif [[ ${params.measurement} == "evalue" ]] then measure="-e ${params.evalue}" else measure="--min-score ${params.bitscore}" fi @@ -2056,9 +2056,9 @@ if (params.DataCheck || params.Analyze) { cp ${params.vampdir}/bin/rename_seq.py . virdb=${params.dbdir}/${params.dbname} grep ">" \${virdb} > headers.list - if [[ ${params.measurement} == "bitscore"]] + if [[ ${params.measurement} == "bitscore" ]] then measure="--min-score ${params.bitscore}" - elif [[ ${params.measurement} == "evalue"]] + elif [[ ${params.measurement} == "evalue" ]] then measure="-e ${params.evalue}" else measure="--min-score ${params.bitscore}" fi @@ -2597,9 +2597,9 @@ if (params.DataCheck || params.Analyze) { """ cp ${params.vampdir}/bin/rename_seq.py . virdb=${params.dbdir}/${params.dbname} - if [[ ${params.measurement} == "bitscore"]] + if [[ ${params.measurement} == "bitscore" ]] then measure="--min-score ${params.bitscore}" - elif [[ ${params.measurement} == "evalue"]] + elif [[ ${params.measurement} == "evalue" ]] then measure="-e ${params.evalue}" else measure="--min-score ${params.bitscore}" fi @@ -2748,9 +2748,9 @@ if (params.DataCheck || params.Analyze) { """ cp ${params.vampdir}/bin/rename_seq.py . virdb=${params.dbdir}/${params.dbname} - if [[ ${params.measurement} == "bitscore"]] + if [[ ${params.measurement} == "bitscore" ]] then measure="--min-score ${params.bitscore}" - elif [[ ${params.measurement} == "evalue"]] + elif [[ ${params.measurement} == "evalue" ]] then measure="-e ${params.evalue}" else measure="--min-score ${params.bitscore}" fi @@ -3248,9 +3248,9 @@ if (params.DataCheck || params.Analyze) { set +e cp ${params.vampdir}/bin/rename_seq.py . virdb=${params.dbdir}/${params.dbname} - if [[ ${params.measurement} == "bitscore"]] + if [[ ${params.measurement} == "bitscore" ]] then measure="--min-score ${params.bitscore}" - elif [[ ${params.measurement} == "evalue"]] + elif [[ ${params.measurement} == "evalue" ]] then measure="-e ${params.evalue}" else measure="--min-score ${params.bitscore}" fi @@ -3404,9 +3404,9 @@ if (params.DataCheck || params.Analyze) { set +e cp ${params.vampdir}/bin/rename_seq.py . virdb=${params.dbdir}/${params.dbname} - if [[ ${params.measurement} == "bitscore"]] + if [[ ${params.measurement} == "bitscore" ]] then measure="--min-score ${params.bitscore}" - elif [[ ${params.measurement} == "evalue"]] + elif [[ ${params.measurement} == "evalue" ]] then measure="-e ${params.evalue}" else measure="--min-score ${params.bitscore}" fi @@ -3729,9 +3729,9 @@ if (params.DataCheck || params.Analyze) { """ cp ${params.vampdir}/bin/rename_seq.py . virdb=${params.dbdir}/${params.dbname} - if [[ ${params.measurement} == "bitscore"]] + if [[ ${params.measurement} == "bitscore" ]] then measure="--min-score ${params.bitscore}" - elif [[ ${params.measurement} == "evalue"]] + elif [[ ${params.measurement} == "evalue" ]] then measure="-e ${params.evalue}" else measure="--min-score ${params.bitscore}" fi @@ -3884,9 +3884,9 @@ if (params.DataCheck || params.Analyze) { """ cp ${params.vampdir}/bin/rename_seq.py . virdb=${params.dbdir}/${params.dbname} - if [[ ${params.measurement} == "bitscore"]] + if [[ ${params.measurement} == "bitscore" ]] then measure="--min-score ${params.bitscore}" - elif [[ ${params.measurement} == "evalue"]] + elif [[ ${params.measurement} == "evalue" ]] then measure="-e ${params.evalue}" else measure="--min-score ${params.bitscore}" fi From 817797accdb7aa1cc337bfe30bd072723056adae Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Wed, 22 Sep 2021 18:22:57 -0500 Subject: [PATCH 189/223] fix to filter process --- vAMPirus.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 1991bac..f009151 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -950,7 +950,7 @@ if (params.DataCheck || params.Analyze) { j=\$(( \${j}+1 )) done ./rename_seq.py ${params.filtDB} filt.headers filterdatabaserenamed.fasta - cat filterdatabase.fasta >> combodatabase.fasta + cat filterdatabaserenamed.fasta >> combodatabase.fasta paste -d',' filt.head filt.headers > filtername_map.csv rm filterdatabaserenamed.fasta From 2851b00081139ee0b72ca28ba5e662ab297944a3 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Wed, 22 Sep 2021 19:03:34 -0500 Subject: [PATCH 190/223] fixed filter process --- vAMPirus.nf | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index f009151..066f20a 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -943,7 +943,7 @@ if (params.DataCheck || params.Analyze) { cp ${params.vampdir}/bin/rename_seq.py . #create and rename filter database - grep ">" ${params.filtDB} | awk -F ">" '{print \$2}' >> filt.head + grep ">" ${params.filtDB} | sed 's/ //g' | awk -F ">" '{print \$2}' >> filt.head j=1 for y in \$( cat filt.head );do echo ">Filt"\$j"" >> filt.headers @@ -955,7 +955,7 @@ if (params.DataCheck || params.Analyze) { rm filterdatabaserenamed.fasta #create and rename keep database - grep ">" ${params.keepDB} | awk -F ">" '{print \$2}' >> keep.head + grep ">" ${params.keepDB} | sed 's/ //g' | awk -F ">" '{print \$2}' >> keep.head d=1 for y in \$( cat keep.head );do echo ">keep"\$d"" >> keep.headers From 361c168cce4605956e2cd4fd7650456f4ad390ce Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Wed, 22 Sep 2021 19:09:30 -0500 Subject: [PATCH 191/223] removed rm command from filter proc --- vAMPirus.nf | 3 +-- 1 file changed, 1 insertion(+), 2 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 066f20a..4feb1b1 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -952,8 +952,7 @@ if (params.DataCheck || params.Analyze) { ./rename_seq.py ${params.filtDB} filt.headers filterdatabaserenamed.fasta cat filterdatabaserenamed.fasta >> combodatabase.fasta paste -d',' filt.head filt.headers > filtername_map.csv - rm filterdatabaserenamed.fasta - + #create and rename keep database grep ">" ${params.keepDB} | sed 's/ //g' | awk -F ">" '{print \$2}' >> keep.head d=1 From 9c2948eb0692a731ac8b14b64088fea9c3fe70fb Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Wed, 22 Sep 2021 19:16:06 -0500 Subject: [PATCH 192/223] fixed quotes in process --- vAMPirus.nf | 10 +++++----- 1 file changed, 5 insertions(+), 5 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 4feb1b1..f7d1938 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -952,7 +952,7 @@ if (params.DataCheck || params.Analyze) { ./rename_seq.py ${params.filtDB} filt.headers filterdatabaserenamed.fasta cat filterdatabaserenamed.fasta >> combodatabase.fasta paste -d',' filt.head filt.headers > filtername_map.csv - + #create and rename keep database grep ">" ${params.keepDB} | sed 's/ //g' | awk -F ">" '{print \$2}' >> keep.head d=1 @@ -984,12 +984,12 @@ if (params.DataCheck || params.Analyze) { fi done if [[ ${params.keepnohit} == "true" ]] - then echo nohit.list >> kep.list - cat kep.list | sort > keep.list + then cat nohit.list >> kep.list + cat kep.list | sort >> keep.list seqtk subseq ${asv} keep.list > kept.fasta u=1 for y in \$( cat keep.list );do - echo ">ASV"\$u"" >> asvrename.list + echo ">ASV\${u} >> asvrename.list u=\$(( \${u}+1 )) done ./rename_seq.py ${asv} asvrename.list ${params.projtag}_ASV.fasta @@ -998,7 +998,7 @@ if (params.DataCheck || params.Analyze) { seqtk subseq ${asv} keep.list > kept.fasta u=1 for y in \$( cat keep.list );do - echo ">ASV"\$u"" >> asvrename.list + echo ">ASV\${u}" >> asvrename.list u=\$(( \${u}+1 )) done ./rename_seq.py ${asv} asvrename.list ${params.projtag}_ASV.fasta From 4fb917ab324f1559ce9cc0d28b18d429f1e38432 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Wed, 22 Sep 2021 19:22:23 -0500 Subject: [PATCH 193/223] fixed quotes in filter process --- vAMPirus.nf | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index f7d1938..0f73199 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -976,7 +976,7 @@ if (params.DataCheck || params.Analyze) { then #check if hit is to filter hit=\$(grep "\$x" ${params.projtag}_diamondfilter.out | awk '{print \$3}') if [[ \$(grep -c "\$hit" filt.headers) -eq 1 ]] - then echo "\$x,\$hit >> filtered_asvs_summary.csv + then echo "\$x,\$hit" >> filtered_asvs_summary.csv elif [[ \$(grep -c "\$hit" keep.headers) -eq 1 ]] then echo "\$x" >> kep.list fi @@ -989,7 +989,7 @@ if (params.DataCheck || params.Analyze) { seqtk subseq ${asv} keep.list > kept.fasta u=1 for y in \$( cat keep.list );do - echo ">ASV\${u} >> asvrename.list + echo ">ASV\${u}" >> asvrename.list u=\$(( \${u}+1 )) done ./rename_seq.py ${asv} asvrename.list ${params.projtag}_ASV.fasta From b2a84033abbcccc0729c205656b145b7185936fb Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Wed, 22 Sep 2021 19:34:17 -0500 Subject: [PATCH 194/223] added extra to output from filter processes --- vAMPirus.nf | 6 ++++-- 1 file changed, 4 insertions(+), 2 deletions(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index 0f73199..f71d640 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -930,14 +930,15 @@ if (params.DataCheck || params.Analyze) { label 'norm_cpus' - publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/FilterCont", mode: "copy", overwrite: true + publishDir "${params.workingdir}/${params.outdir}/ReadProcessing/ASVFiltering", mode: "copy", overwrite: true input: file(asv) from asvforfilt output: file("*ASV.fasta") into ( reads_vsearch5_ch, asv_med, nucl2aa, asvsforAminotyping, asvfastaforcounts, asvaminocheck ) - + file("*.csv") into ( nothing ) + file("*diamondfilter.out") into ( noth) script: """ cp ${params.vampdir}/bin/rename_seq.py . @@ -1003,6 +1004,7 @@ if (params.DataCheck || params.Analyze) { done ./rename_seq.py ${asv} asvrename.list ${params.projtag}_ASV.fasta fi + paste -d',' keep.list asvrename.list > ASV_rename_map.csv """ } From a02ae106283bbfa3f5bdec56566dcab61be32393 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Wed, 22 Sep 2021 19:37:37 -0500 Subject: [PATCH 195/223] changed name of process --- vAMPirus.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index f71d640..e45dfc4 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -926,7 +926,7 @@ if (params.DataCheck || params.Analyze) { // UNTIL HERE DEFAULT if (params.filter) { - process Filtering { + process ASV_Filtering { label 'norm_cpus' From 11f91ba6b5a761cd2b424ce8b3c50ddcdcfda453 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Thu, 23 Sep 2021 09:29:34 -0500 Subject: [PATCH 196/223] fixed mscule command in aminoMED in datacheck --- vAMPirus.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index e45dfc4..cabfb98 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -1301,7 +1301,7 @@ if (params.DataCheck || params.Analyze) { """ #alignment if [[ \$(grep -c ">" ${aminos}) -gt 499 ]]; then algo="super5"; else algo="mpc"; fi - ${tools}/muscle5.0.1278_linux64 -"\${algo}" ${aminos} -out ${params.projtag}_AminoTypes_muscleAlign.fasta -threads ${task.cpus} -quiet + ${tools}/muscle5.0.1278_linux64 -\${algo} ${aminos} -out ${params.projtag}_AminoTypes_muscleAlign.fasta -threads ${task.cpus} -quiet #trimming trimal -in ${params.projtag}_AminoTypes_muscleAlign.fasta -out ${params.projtag}_AminoTypes_muscleAligned.fasta -keepheader -fasta -automated1 rm ${params.projtag}_AminoTypes_muscleAlign.fasta From 3277a65ec3fa8e88ec927760f114f23bc7938e3b Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Thu, 23 Sep 2021 22:13:29 -0500 Subject: [PATCH 197/223] fixed typo in aminotype tree --- vAMPirus.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/vAMPirus.nf b/vAMPirus.nf index cabfb98..3c6b0dd 100644 --- a/vAMPirus.nf +++ b/vAMPirus.nf @@ -2902,7 +2902,7 @@ if (params.DataCheck || params.Analyze) { iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -b ${params.boots} elif [ "${params.parametric}" != "false" ];then - iqtree -s\${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni + iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni else iqtree -s \${pre}_Aligned_informativeonly.fasta --prefix \${pre}_iq -m MFP --redo -nt auto -bb ${params.boots} -bnni From c1bd3fea13229a1a91c4d89881d2a3928f901ebc Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Mon, 27 Sep 2021 14:40:17 -0500 Subject: [PATCH 198/223] updated headers fo shannon entropy figures in datacheck report --- bin/vAMPirus_DC_Report.Rmd | 8 ++++---- 1 file changed, 4 insertions(+), 4 deletions(-) diff --git a/bin/vAMPirus_DC_Report.Rmd b/bin/vAMPirus_DC_Report.Rmd index 728e770..b55fa92 100644 --- a/bin/vAMPirus_DC_Report.Rmd +++ b/bin/vAMPirus_DC_Report.Rmd @@ -399,7 +399,7 @@ NOTE: The "1" represents the number of AminoTypes which are unique amino acid se
    -### ASVs MED +### ASV Shannon Entropy Analysis (https://merenlab.org/2012/05/11/oligotyping-pipeline-explained/)
    @@ -415,7 +415,7 @@ mplot ```
    -### ASVs Peaks Range Information +### ASV Shannon Entropy Peaks Breakdown
    @@ -425,7 +425,7 @@ paged_table(med_asv_csv, options = list(rows.print = 10)) ```
    -### AminoTypes MED +### AminoTypes Shannon Entropy Analysis (https://merenlab.org/2012/05/11/oligotyping-pipeline-explained/)
    @@ -441,7 +441,7 @@ mplot ```
    -### AminoTypes Peaks Range Information +### AminoTypes Shannon Entropy Peaks Breakdown
    From 7badd77b77675d19f20abdfcdedb7c5309b2ea75 Mon Sep 17 00:00:00 2001 From: Alex Veglia Date: Wed, 29 Sep 2021 16:40:23 -0500 Subject: [PATCH 199/223] added options for datacheck MED processes --- bin/vAMPirus_DC_Report.Rmd | 2 + docs/HelpDocumentation.md | 16 +- vAMPirus.nf | 632 +++++++++++++++++++------------------ vampirus.config | 10 +- 4 files changed, 346 insertions(+), 314 deletions(-) diff --git a/bin/vAMPirus_DC_Report.Rmd b/bin/vAMPirus_DC_Report.Rmd index b55fa92..0eb3ea8 100644 --- a/bin/vAMPirus_DC_Report.Rmd +++ b/bin/vAMPirus_DC_Report.Rmd @@ -8,6 +8,8 @@ params: skipReadProcessing: !r commandArgs(trailingOnly=T)[2] skipMerging: !r commandArgs(trailingOnly=T)[3] skipAdapterRemoval: !r commandArgs(trailingOnly=T)[4] + asvMED: !r commandArgs(trailingOnly=T)[5] + aminoMED: !r commandArgs(trailingOnly=T)[6] ---

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