From f280fa93f882f57637949840f9c957e5fa4465cf Mon Sep 17 00:00:00 2001 From: daniel Date: Fri, 3 Mar 2023 13:08:38 +0100 Subject: [PATCH 001/158] update to 2.6.0dev --- nextflow.config | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/nextflow.config b/nextflow.config index 5f4ae444..28efd87b 100644 --- a/nextflow.config +++ b/nextflow.config @@ -260,7 +260,7 @@ manifest { description = """Amplicon sequencing analysis workflow using DADA2 and QIIME2""" mainScript = 'main.nf' nextflowVersion = '!>=22.10.1' - version = '2.5.0' + version = '2.6.0dev' doi = '10.3389/fmicb.2020.550420' } From b012f29b211f13f7a65444a40bc568006dc82a67 Mon Sep 17 00:00:00 2001 From: daniel Date: Fri, 3 Mar 2023 13:08:56 +0100 Subject: [PATCH 002/158] update CHANGELOG --- CHANGELOG.md | 12 ++++++++++++ 1 file changed, 12 insertions(+) diff --git a/CHANGELOG.md b/CHANGELOG.md index 0df376d7..a071b375 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -3,6 +3,18 @@ The format is based on [Keep a Changelog](https://keepachangelog.com/en/1.0.0/) and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0.html). +## nf-core/ampliseq version 2.6.0dev + +### `Added` + +### `Changed` + +### `Fixed` + +### `Dependencies` + +### `Removed` + ## nf-core/ampliseq version 2.5.0 - 2023-03-02 ### `Added` From 7741fee6f20f0fa8155b52e97971f613d476c4f3 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Tue, 7 Mar 2023 13:16:47 +0100 Subject: [PATCH 003/158] Import vsearch/sintax module --- modules/nf-core/vsearch/sintax/main.nf | 38 ++++++++++++++++++++++++ modules/nf-core/vsearch/sintax/meta.yml | 39 +++++++++++++++++++++++++ workflows/ampliseq.nf | 1 + 3 files changed, 78 insertions(+) create mode 100644 modules/nf-core/vsearch/sintax/main.nf create mode 100644 modules/nf-core/vsearch/sintax/meta.yml diff --git a/modules/nf-core/vsearch/sintax/main.nf b/modules/nf-core/vsearch/sintax/main.nf new file mode 100644 index 00000000..273ca698 --- /dev/null +++ b/modules/nf-core/vsearch/sintax/main.nf @@ -0,0 +1,38 @@ +process VSEARCH_SINTAX { + tag "${meta.id}" + label 'process_low' + + conda "bioconda::vsearch=2.21.1" + container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? + 'https://depot.galaxyproject.org/singularity/vsearch:2.21.1--h95f258a_0': + 'quay.io/biocontainers/vsearch:2.21.1--h95f258a_0' }" + + input: + tuple val(meta), path(queryfasta) + path db + + output: + tuple val(meta), path('*.tsv') , optional: true, emit: tsv + path "versions.yml" , emit: versions + + when: + task.ext.when == null || task.ext.when + + script: + def args = task.ext.args ?: '' + def prefix = task.ext.prefix ?: "${meta.id}" + + """ + vsearch \\ + --sintax $queryfasta \\ + --db $db \\ + --threads $task.cpus \\ + $args \\ + --tabbedout ${prefix}.tsv + + cat <<-END_VERSIONS > versions.yml + "${task.process}": + vsearch: \$(vsearch --version 2>&1 | head -n 1 | sed 's/vsearch //g' | sed 's/,.*//g' | sed 's/^v//' | sed 's/_.*//') + END_VERSIONS + """ +} diff --git a/modules/nf-core/vsearch/sintax/meta.yml b/modules/nf-core/vsearch/sintax/meta.yml new file mode 100644 index 00000000..61019499 --- /dev/null +++ b/modules/nf-core/vsearch/sintax/meta.yml @@ -0,0 +1,39 @@ +name: "vsearch_sintax" +description: Taxonomic classification using the sintax algorithm. +keywords: + - vsearch + - sintax +tools: + - "vsearch": + description: "VSEARCH is a versatile open-source tool for microbiome analysis, including chimera detection, clustering, dereplication and rereplication, extraction, FASTA/FASTQ/SFF file processing, masking, orienting, pair-wise alignment, restriction site cutting, searching, shuffling, sorting, subsampling, and taxonomic classification of amplicon sequences for metagenomics, genomics, and population genetics. (USEARCH alternative)" + homepage: "https://github.com/torognes/vsearch" + + tool_dev_url: "https://github.com/torognes/vsearch" + doi: "10.7717/peerj.2584" + licence: "['GPL v3-or-later OR BSD-2-clause']" + +input: + - meta: + type: map + description: Groovy Map containing query file information e.g. [ id:'test' ] + - queryfasta: + type: file + description: Query sequences in FASTA or FASTQ format + pattern: "*.{fasta,fa,fna,faa,fastq,fq}" + - db: + type: file + description: Reference database file in FASTA or UDB format + pattern: "*" + +output: + - tsv: + type: file + description: Results written to tab-delimited file + pattern: "*.{tsv}" + - versions: + type: file + description: File containing software versions + pattern: "versions.yml" + +authors: + - "@jtangrot" diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 864d0802..87e52f0e 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -182,6 +182,7 @@ include { CUTADAPT as CUTADAPT_TAXONOMY } from '../modules/nf-core/cutadapt/ include { FASTQC } from '../modules/nf-core/fastqc/main' include { MULTIQC } from '../modules/nf-core/multiqc/main' include { CUSTOM_DUMPSOFTWAREVERSIONS } from '../modules/nf-core/custom/dumpsoftwareversions/main' +include { VSEARCH_SINTAX } from '../modules/nf-core/vsearch/sintax/main' include { VSEARCH_USEARCHGLOBAL } from '../modules/nf-core/vsearch/usearchglobal/main' From 4ff980e7a55aaafcaa0fa77c728c996ef9da3a07 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Tue, 7 Mar 2023 13:19:25 +0100 Subject: [PATCH 004/158] Add sintax databases to config --- conf/ref_databases.config | 49 +++++++++++++++++++++++++++++++++++++++ 1 file changed, 49 insertions(+) diff --git a/conf/ref_databases.config b/conf/ref_databases.config index a8b55ba2..48dd93f0 100644 --- a/conf/ref_databases.config +++ b/conf/ref_databases.config @@ -258,4 +258,53 @@ params { fmtscript = "taxref_reformat_qiime_greengenes85.sh" } } + //Sintax taxonomic reference databases + sintax_ref_databases { + 'coidb' { + title = "COIDB - CO1 Taxonomy Database - Release 221216" + file = [ "https://figshare.scilifelab.se/ndownloader/files/38787072", "https://figshare.scilifelab.se/ndownloader/files/38787069" ] + citation = "Sundh J, Manoharan L, Iwaszkiewicz-Eggebrecht E, Miraldo A, Andersson A, Ronquist F. COI reference sequences from BOLD DB. doi: https://doi.org/10.17044/scilifelab.20514192.v2" + fmtscript = "taxref_reformat_sintax_coidb.sh" + dbversion = "COIDB 221216 (https://doi.org/10.17044/scilifelab.20514192.v2)" + } + 'coidb=221216' { + title = "COIDB - CO1 Taxonomy Database - Release 221216" + file = [ "https://figshare.scilifelab.se/ndownloader/files/38787072", "https://figshare.scilifelab.se/ndownloader/files/38787069" ] + citation = "Sundh J, Manoharan L, Iwaszkiewicz-Eggebrecht E, Miraldo A, Andersson A, Ronquist F. COI reference sequences from BOLD DB. doi: https://doi.org/10.17044/scilifelab.20514192.v2" + fmtscript = "taxref_reformat_sintax_coidb.sh" + dbversion = "COIDB 221216 (https://doi.org/10.17044/scilifelab.20514192.v2)" + } + 'unite-fungi' { + title = "UNITE USEARCH/UTAX release for Fungi - Version 8.3" + file = [ "https://files.plutof.ut.ee/public/orig/82/CB/82CB44BBAAA7D3AEAC297B5689BDA2963E8D0666E01FE0B54096147AFAF85263.gz" ] + citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2021): UNITE USEARCH/UTAX release for Fungi. UNITE Community. 10.15156/BIO/1280276" + fmtscript = "taxref_reformat_sintax_unite.sh" + dbversion = "UNITE-fungi v8.3 (https://dx.doi.org/10.15156/BIO/1280276)" + shfile = [ "https://scilifelab.figshare.com/ndownloader/files/34497977", "https://scilifelab.figshare.com/ndownloader/files/34497980"] + } + 'unite-fungi=8.3' { + title = "UNITE USEARCH/UTAX release for Fungi - Version 8.3" + file = [ "https://files.plutof.ut.ee/public/orig/82/CB/82CB44BBAAA7D3AEAC297B5689BDA2963E8D0666E01FE0B54096147AFAF85263.gz" ] + citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2021): UNITE USEARCH/UTAX release for Fungi. UNITE Community. 10.15156/BIO/1280276" + fmtscript = "taxref_reformat_sintax_unite.sh" + dbversion = "UNITE-fungi v8.3 (https://doi.org/10.15156/BIO/1280049)" + shfile = [ "https://scilifelab.figshare.com/ndownloader/files/34497977", "https://scilifelab.figshare.com/ndownloader/files/34497980"] + } + 'unite-alleuk' { + title = "UNITE USEARCH/UTAX release for eukaryotes - Version 8.3" + file = [ "https://files.plutof.ut.ee/public/orig/B9/35/B9351C91550A52713CB66DB7A1CEF35765310EBB23B6667AC93E714E9A9D020B.gz"] + citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2021): UNITE USEARCH/UTAX release for eukaryotes. UNITE Community. 10.15156/BIO/1280317" + fmtscript = "taxref_reformat_sintax_unite.sh" + dbversion = "UNITE-alleuk v8.3 (https://doi.org/10.15156/BIO/1280127)" + shfile = [ "https://scilifelab.figshare.com/ndownloader/files/34994575", "https://scilifelab.figshare.com/ndownloader/files/34994578"] + } + 'unite-alleuk=8.3' { + title = "UNITE USEARCH/UTAX release for eukaryotes - Version 8.3" + file = [ "https://files.plutof.ut.ee/public/orig/B9/35/B9351C91550A52713CB66DB7A1CEF35765310EBB23B6667AC93E714E9A9D020B.gz"] + citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2021): UNITE USEARCH/UTAX release for eukaryotes. UNITE Community. 10.15156/BIO/1280317" + fmtscript = "taxref_reformat_sintax_unite.sh" + dbversion = "UNITE-alleuk v8.3 (https://doi.org/10.15156/BIO/1280127)" + shfile = [ "https://scilifelab.figshare.com/ndownloader/files/34994575", "https://scilifelab.figshare.com/ndownloader/files/34994578"] + } + } } From dfbff728b88c3ff383f92761916ffb2cec573934 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Wed, 8 Mar 2023 11:35:58 +0100 Subject: [PATCH 005/158] Include settings for vsearch sintax --- conf/modules.config | 19 +++++++++++++++++++ modules.json | 5 +++++ nextflow.config | 2 ++ nextflow_schema.json | 4 ++++ 4 files changed, 30 insertions(+) diff --git a/conf/modules.config b/conf/modules.config index d0bf626a..ff10456e 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -381,6 +381,25 @@ process { ] } + withName: FORMAT_TAXONOMY_SINTAX { + publishDir = [ + path: { "${params.outdir}/sintax" }, + mode: params.publish_dir_mode, + pattern: "{ref_taxonomy_sintax.txt}" + ] + } + + withName: VSEARCH_SINTAX { + ext.args = "--randseed ${params.seed}" + publishDir = [ + [ + path: { "${params.outdir}/sintax" }, + mode: params.publish_dir_mode, + pattern: "*.tsv" + ] + ] + } + withName: VSEARCH_USEARCHGLOBAL { ext.args = '--top_hits_only --output_no_hits --maxaccepts 50 --query_cov 0.9' publishDir = [ diff --git a/modules.json b/modules.json index c3b153ee..382df979 100644 --- a/modules.json +++ b/modules.json @@ -25,6 +25,11 @@ "git_sha": "ee80d14721e76e2e079103b8dcd5d57129e584ba", "installed_by": ["modules"] }, + "vsearch/sintax": { + "branch": "master", + "git_sha": "0f8a77ff00e65eaeebc509b8156eaa983192474b", + "installed_by": ["modules"] + }, "vsearch/usearchglobal": { "branch": "master", "git_sha": "c8e35eb2055c099720a75538d1b8adb3fb5a464c", diff --git a/nextflow.config b/nextflow.config index 5f4ae444..51cc3689 100644 --- a/nextflow.config +++ b/nextflow.config @@ -46,6 +46,7 @@ params { its_partial = 0 picrust = false sbdiexport = false + sintax = false addsh = false dada_tax_agglom_min = 2 dada_tax_agglom_max = 6 @@ -83,6 +84,7 @@ params { dada_ref_tax_custom = null dada_ref_tax_custom_sp = null cut_dada_ref_taxonomy = false + sintax_ref_taxonomy = null qiime_ref_taxonomy = null // MultiQC options diff --git a/nextflow_schema.json b/nextflow_schema.json index 4a343309..f0e1d63a 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -279,6 +279,10 @@ "description": "Path to QIIME2 trained classifier file (typically *-classifier.qza)", "help_text": "If you have trained a compatible classifier before, from sources such as SILVA (https://www.arb-silva.de/), Greengenes (http://greengenes.secondgenome.com/downloads) or RDP (https://rdp.cme.msu.edu/). \n\nFor example:\n\n```bash\n--classifier \"FW_primer-RV_primer-classifier.qza\"\n```\n\nPlease note the following requirements:\n\n1. The path must be enclosed in quotes\n2. The classifier is a Naive Bayes classifier produced by `qiime feature-classifier fit-classifier-naive-bayes` (e.g. by this pipeline)\n3. The primer pair for the amplicon PCR and the computing of the classifier are exactly the same (or full-length, potentially lower performance)\n4. The classifier has to be trained by the same version of scikit-learn as this version of the pipeline uses" }, + "sintax": { + "type": "boolean", + "description": "If VSEARCH sintax should be used for taxonomic assingment of ASVs." + }, "addsh": { "type": "boolean", "description": "If ASVs should be assigned to UNITE species hypotheses (SHs). Only relevant for ITS data." From 8b0c952f8e870e91edccfb839f164fc76e217999 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Wed, 8 Mar 2023 11:36:58 +0100 Subject: [PATCH 006/158] Add sintax option to workflow --- bin/taxref_reformat_sintax_unite.sh | 7 +++++ modules/local/format_taxonomy_sintax.nf | 35 +++++++++++++++++++++++++ workflows/ampliseq.nf | 27 ++++++++++++++++--- 3 files changed, 66 insertions(+), 3 deletions(-) create mode 100755 bin/taxref_reformat_sintax_unite.sh create mode 100644 modules/local/format_taxonomy_sintax.nf diff --git a/bin/taxref_reformat_sintax_unite.sh b/bin/taxref_reformat_sintax_unite.sh new file mode 100755 index 00000000..39005428 --- /dev/null +++ b/bin/taxref_reformat_sintax_unite.sh @@ -0,0 +1,7 @@ +#!/bin/sh + +# Handles the UNITE USEARCH/UTAX database. + +# There's a preformatted USEARCH/UTAX file that is suitable for sintax -- this is just renamed +mv *.gz sintaxdb.fa.gz + diff --git a/modules/local/format_taxonomy_sintax.nf b/modules/local/format_taxonomy_sintax.nf new file mode 100644 index 00000000..825d4ecf --- /dev/null +++ b/modules/local/format_taxonomy_sintax.nf @@ -0,0 +1,35 @@ +process FORMAT_TAXONOMY_SINTAX { + label 'process_low' + + conda "conda-forge::sed=4.7" + container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? + 'https://containers.biocontainers.pro/s3/SingImgsRepo/biocontainers/v1.2.0_cv1/biocontainers_v1.2.0_cv1.img' : + 'biocontainers/biocontainers:v1.2.0_cv1' }" + + input: + path(database) + + output: + path( "sintaxdb.fa.gz" ), emit: db + path( "ref_taxonomy_sintax.txt") , emit: ref_tax_info + path "versions.yml" , emit: versions + + when: + task.ext.when == null || task.ext.when + + script: + """ + ${params.sintax_ref_databases[params.sintax_ref_taxonomy]["fmtscript"]} + + #Giving out information + echo -e "--sintax_ref_taxonomy: ${params.sintax_ref_taxonomy}\n" >ref_taxonomy_sintax.txt + echo -e "Title: ${params.sintax_ref_databases[params.sintax_ref_taxonomy]["title"]}\n" >>ref_taxonomy_sintax.txt + echo -e "Citation: ${params.sintax_ref_databases[params.sintax_ref_taxonomy]["citation"]}\n" >>ref_taxonomy_sintax.txt + echo "All entries: ${params.sintax_ref_databases[params.sintax_ref_taxonomy]}" >>ref_taxonomy_sintax.txt + + cat <<-END_VERSIONS > versions.yml + "${task.process}": + bash: \$(bash --version | sed -n 1p | sed 's/GNU bash, version //g') + END_VERSIONS + """ +} diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 87e52f0e..0d17cb19 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -63,6 +63,10 @@ if (params.qiime_ref_taxonomy && !params.skip_taxonomy && !params.classifier) { ch_qiime_ref_taxonomy = Channel.fromList(params.qiime_ref_databases[params.qiime_ref_taxonomy]["file"]).map { file(it) } } else { ch_qiime_ref_taxonomy = Channel.empty() } +if (params.sintax_ref_taxonomy && !params.skip_taxonomy) { + ch_sintax_ref_taxonomy = Channel.fromList(params.sintax_ref_databases[params.sintax_ref_taxonomy]["file"]).map { file(it) } +} else { ch_sintax_ref_taxonomy = Channel.empty() } + // Set non-params Variables @@ -134,6 +138,7 @@ include { MERGE_STATS as MERGE_STATS_FILTERSSU } from '../modules/local/merge_st include { MERGE_STATS as MERGE_STATS_FILTERLENASV } from '../modules/local/merge_stats' include { FORMAT_FASTAINPUT } from '../modules/local/format_fastainput' include { FORMAT_TAXONOMY } from '../modules/local/format_taxonomy' +include { FORMAT_TAXONOMY_SINTAX } from '../modules/local/format_taxonomy_sintax' include { ITSX_CUTASV } from '../modules/local/itsx_cutasv' include { MERGE_STATS as MERGE_STATS_STD} from '../modules/local/merge_stats' include { DADA2_TAXONOMY } from '../modules/local/dada2_taxonomy' @@ -410,9 +415,9 @@ workflow AMPLISEQ { ASSIGNSH( DADA2_TAXONOMY.out.tsv, ch_shinfo.collect(), VSEARCH_USEARCHGLOBAL.out.txt, 'ASV_tax_SH.tsv') ch_versions = ch_versions.mix(ASSIGNSH.out.versions.ifEmpty(null)) ch_dada2_tax = ASSIGNSH.out.tsv - } else { - ch_dada2_tax = DADA2_TAXONOMY.out.tsv - } + } else { + ch_dada2_tax = DADA2_TAXONOMY.out.tsv + } } //Cut out ITS region if long ITS reads } else { @@ -472,6 +477,22 @@ workflow AMPLISEQ { } } + // Sintax + if (params.sintax) { + FORMAT_TAXONOMY_SINTAX ( ch_sintax_ref_taxonomy.collect() ) + ch_sintaxdb = FORMAT_TAXONOMY_SINTAX.out.db + ch_fasta + .map { + fasta -> + def meta = [:] + meta.id = "ASV_tax_sintax" + [ meta, fasta ] } + .set { ch_fasta_sintax } + VSEARCH_SINTAX( ch_fasta_sintax, ch_sintaxdb ) + ch_versions = ch_versions.mix(VSEARCH_SINTAX.out.versions) + } + + //QIIME2 if ( run_qiime2 ) { if (params.qiime_ref_taxonomy && !params.classifier) { From 957cc55c6cdc757ce3d9fbaa59fcd9bc7b0bda49 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Wed, 8 Mar 2023 13:10:19 +0100 Subject: [PATCH 007/158] add sintax_ref_taxonomy to schema --- nextflow.config | 2 +- nextflow_schema.json | 16 ++++++++++++++++ 2 files changed, 17 insertions(+), 1 deletion(-) diff --git a/nextflow.config b/nextflow.config index 51cc3689..24708ea4 100644 --- a/nextflow.config +++ b/nextflow.config @@ -107,7 +107,7 @@ params { version = false validate_params = true show_hidden_params = false - schema_ignore_params = 'dada_ref_databases,qiime_ref_databases,igenomes_base' + schema_ignore_params = 'dada_ref_databases,qiime_ref_databases,sintax_ref_databases,igenomes_base' // Config options diff --git a/nextflow_schema.json b/nextflow_schema.json index f0e1d63a..a2a4c43c 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -279,6 +279,22 @@ "description": "Path to QIIME2 trained classifier file (typically *-classifier.qza)", "help_text": "If you have trained a compatible classifier before, from sources such as SILVA (https://www.arb-silva.de/), Greengenes (http://greengenes.secondgenome.com/downloads) or RDP (https://rdp.cme.msu.edu/). \n\nFor example:\n\n```bash\n--classifier \"FW_primer-RV_primer-classifier.qza\"\n```\n\nPlease note the following requirements:\n\n1. The path must be enclosed in quotes\n2. The classifier is a Naive Bayes classifier produced by `qiime feature-classifier fit-classifier-naive-bayes` (e.g. by this pipeline)\n3. The primer pair for the amplicon PCR and the computing of the classifier are exactly the same (or full-length, potentially lower performance)\n4. The classifier has to be trained by the same version of scikit-learn as this version of the pipeline uses" }, + "sintax_ref_taxonomy": { + "type": "string", + "help_text": "Choose any of the supported databases, and optionally also specify the version. Database and version are separated by an equal sign (`=`, e.g. `coidb=221216`) . This will download the desired database and if needed format it to produce a file that is compatible with USEARCH sintax.\n\nThe following databases are supported:\n- UNITE - eukaryotic nuclear ribosomal ITS region - ITS\n- COIDB - eukaryotic Cytochrome Oxidase I (COI) from The Barcode of Life Data System (BOLD) - COI\n\nGenerally, using `coidb`, `unite-fungi`, or `unite-alleuk` will select the most recent supported version. For details on what values are valid, please either use an invalid value such as `x` (causing the pipeline to send an error message with a list of all valid values) or see `conf/ref_databases.config`.", + "description": "Name of supported database, and optionally also version number", + "default": "null", + "enum": [ + "coidb", + "coidb=221216", + "unite-fungi=8.3", + "unite-fungi=8.2", + "unite-fungi", + "unite-alleuk=8.3", + "unite-alleuk=8.2", + "unite-alleuk" + ] + }, "sintax": { "type": "boolean", "description": "If VSEARCH sintax should be used for taxonomic assingment of ASVs." From 9ada47a9101efb874bf576716b054441280b024a Mon Sep 17 00:00:00 2001 From: jtangrot Date: Wed, 8 Mar 2023 13:38:58 +0100 Subject: [PATCH 008/158] Set 1 cpu for reproducibility --- conf/modules.config | 1 + 1 file changed, 1 insertion(+) diff --git a/conf/modules.config b/conf/modules.config index ff10456e..b6711efb 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -391,6 +391,7 @@ process { withName: VSEARCH_SINTAX { ext.args = "--randseed ${params.seed}" + cpus = 1 publishDir = [ [ path: { "${params.outdir}/sintax" }, From 167e166c80917bf3abb32f95bd99832b43f7091d Mon Sep 17 00:00:00 2001 From: jtangrot Date: Wed, 8 Mar 2023 15:25:14 +0100 Subject: [PATCH 009/158] Add coidb and unite 8.2 --- bin/taxref_reformat_sintax.sh | 8 +++++++ bin/taxref_reformat_sintax_unite.sh | 7 ------ conf/ref_databases.config | 36 ++++++++++++++++++----------- 3 files changed, 31 insertions(+), 20 deletions(-) create mode 100755 bin/taxref_reformat_sintax.sh delete mode 100755 bin/taxref_reformat_sintax_unite.sh diff --git a/bin/taxref_reformat_sintax.sh b/bin/taxref_reformat_sintax.sh new file mode 100755 index 00000000..b076d0af --- /dev/null +++ b/bin/taxref_reformat_sintax.sh @@ -0,0 +1,8 @@ +#!/bin/sh + +# Handles preformatted database files suitable for sintax + +# Just rename the preformatted file +# Assumes only one (gzipped) file +mv * sintaxdb.fa.gz + diff --git a/bin/taxref_reformat_sintax_unite.sh b/bin/taxref_reformat_sintax_unite.sh deleted file mode 100755 index 39005428..00000000 --- a/bin/taxref_reformat_sintax_unite.sh +++ /dev/null @@ -1,7 +0,0 @@ -#!/bin/sh - -# Handles the UNITE USEARCH/UTAX database. - -# There's a preformatted USEARCH/UTAX file that is suitable for sintax -- this is just renamed -mv *.gz sintaxdb.fa.gz - diff --git a/conf/ref_databases.config b/conf/ref_databases.config index 48dd93f0..cebc4462 100644 --- a/conf/ref_databases.config +++ b/conf/ref_databases.config @@ -262,49 +262,59 @@ params { sintax_ref_databases { 'coidb' { title = "COIDB - CO1 Taxonomy Database - Release 221216" - file = [ "https://figshare.scilifelab.se/ndownloader/files/38787072", "https://figshare.scilifelab.se/ndownloader/files/38787069" ] + file = [ "https://figshare.scilifelab.se/ndownloader/files/38787078" ] citation = "Sundh J, Manoharan L, Iwaszkiewicz-Eggebrecht E, Miraldo A, Andersson A, Ronquist F. COI reference sequences from BOLD DB. doi: https://doi.org/10.17044/scilifelab.20514192.v2" - fmtscript = "taxref_reformat_sintax_coidb.sh" + fmtscript = "taxref_reformat_sintax.sh" dbversion = "COIDB 221216 (https://doi.org/10.17044/scilifelab.20514192.v2)" } 'coidb=221216' { title = "COIDB - CO1 Taxonomy Database - Release 221216" - file = [ "https://figshare.scilifelab.se/ndownloader/files/38787072", "https://figshare.scilifelab.se/ndownloader/files/38787069" ] + file = [ "https://figshare.scilifelab.se/ndownloader/files/38787078" ] citation = "Sundh J, Manoharan L, Iwaszkiewicz-Eggebrecht E, Miraldo A, Andersson A, Ronquist F. COI reference sequences from BOLD DB. doi: https://doi.org/10.17044/scilifelab.20514192.v2" - fmtscript = "taxref_reformat_sintax_coidb.sh" + fmtscript = "taxref_reformat_sintax.sh" dbversion = "COIDB 221216 (https://doi.org/10.17044/scilifelab.20514192.v2)" } 'unite-fungi' { title = "UNITE USEARCH/UTAX release for Fungi - Version 8.3" file = [ "https://files.plutof.ut.ee/public/orig/82/CB/82CB44BBAAA7D3AEAC297B5689BDA2963E8D0666E01FE0B54096147AFAF85263.gz" ] citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2021): UNITE USEARCH/UTAX release for Fungi. UNITE Community. 10.15156/BIO/1280276" - fmtscript = "taxref_reformat_sintax_unite.sh" + fmtscript = "taxref_reformat_sintax.sh" dbversion = "UNITE-fungi v8.3 (https://dx.doi.org/10.15156/BIO/1280276)" - shfile = [ "https://scilifelab.figshare.com/ndownloader/files/34497977", "https://scilifelab.figshare.com/ndownloader/files/34497980"] } 'unite-fungi=8.3' { title = "UNITE USEARCH/UTAX release for Fungi - Version 8.3" file = [ "https://files.plutof.ut.ee/public/orig/82/CB/82CB44BBAAA7D3AEAC297B5689BDA2963E8D0666E01FE0B54096147AFAF85263.gz" ] citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2021): UNITE USEARCH/UTAX release for Fungi. UNITE Community. 10.15156/BIO/1280276" - fmtscript = "taxref_reformat_sintax_unite.sh" - dbversion = "UNITE-fungi v8.3 (https://doi.org/10.15156/BIO/1280049)" - shfile = [ "https://scilifelab.figshare.com/ndownloader/files/34497977", "https://scilifelab.figshare.com/ndownloader/files/34497980"] + fmtscript = "taxref_reformat_sintax.sh" + dbversion = "UNITE-fungi v8.3 (https://dx.doi.org/10.15156/BIO/1280276)" + } + 'unite-fungi=8.2' { + title = "UNITE USEARCH/UTAX release for Fungi - Version 8.2" + file = [ "https://files.plutof.ut.ee/public/orig/E8/83/E883EB19E3EA7B64C1F652521301239831FAFE0BFF015C9E2B4786DC0976C0FC.gz" ] + citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2020): UNITE USEARCH/UTAX release for Fungi. UNITE Community. 10.15156/BIO/786375" + fmtscript = "taxref_reformat_sintax.sh" + dbversion = "UNITE-fungi v8.2 (https://doi.org/10.15156/BIO/786375)" } 'unite-alleuk' { title = "UNITE USEARCH/UTAX release for eukaryotes - Version 8.3" file = [ "https://files.plutof.ut.ee/public/orig/B9/35/B9351C91550A52713CB66DB7A1CEF35765310EBB23B6667AC93E714E9A9D020B.gz"] citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2021): UNITE USEARCH/UTAX release for eukaryotes. UNITE Community. 10.15156/BIO/1280317" - fmtscript = "taxref_reformat_sintax_unite.sh" + fmtscript = "taxref_reformat_sintax.sh" dbversion = "UNITE-alleuk v8.3 (https://doi.org/10.15156/BIO/1280127)" - shfile = [ "https://scilifelab.figshare.com/ndownloader/files/34994575", "https://scilifelab.figshare.com/ndownloader/files/34994578"] } 'unite-alleuk=8.3' { title = "UNITE USEARCH/UTAX release for eukaryotes - Version 8.3" file = [ "https://files.plutof.ut.ee/public/orig/B9/35/B9351C91550A52713CB66DB7A1CEF35765310EBB23B6667AC93E714E9A9D020B.gz"] citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2021): UNITE USEARCH/UTAX release for eukaryotes. UNITE Community. 10.15156/BIO/1280317" - fmtscript = "taxref_reformat_sintax_unite.sh" + fmtscript = "taxref_reformat_sintax.sh" dbversion = "UNITE-alleuk v8.3 (https://doi.org/10.15156/BIO/1280127)" - shfile = [ "https://scilifelab.figshare.com/ndownloader/files/34994575", "https://scilifelab.figshare.com/ndownloader/files/34994578"] + } + 'unite-alleuk=8.2' { + title = "UNITE USEARCH/UTAX release for eukaryotes - Version 8.2" + file = [ "https://files.plutof.ut.ee/public/orig/7B/B5/7BB51166C988E448392B213A72D4604E2ABB494E20E19E4392F7819FEBFCD036.gz"] + citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2020): UNITE USEARCH/UTAX release for eukaryotes. UNITE Community. 10.15156/BIO/786376" + fmtscript = "taxref_reformat_sintax.sh" + dbversion = "UNITE-alleuk v8.2 (https://dx.doi.org/10.15156/BIO/786376)" } } } From fcb922a7ab7a10ebc3ab9bc49f25a6f88f3a9238 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Fri, 10 Mar 2023 11:13:19 +0100 Subject: [PATCH 010/158] Make sbdiexport handle empty barrnap file --- bin/sbdiexportreannotate.R | 5 +++++ workflows/ampliseq.nf | 2 +- 2 files changed, 6 insertions(+), 1 deletion(-) diff --git a/bin/sbdiexportreannotate.R b/bin/sbdiexportreannotate.R index 0413e30b..761dda15 100755 --- a/bin/sbdiexportreannotate.R +++ b/bin/sbdiexportreannotate.R @@ -25,6 +25,11 @@ taxonomy <- read.delim(taxfile, sep = '\t', stringsAsFactors = FALSE) # Read the predictions table if provided, otherwise create one if ( ! is.na(predfile) ) { predictions <- read.delim(predfile, sep = '\t', stringsAsFactors = FALSE) + if ( nrow(predictions) < 1 ) { + colnames <- names(predictions) + predictions <- data.frame(ASV_ID = taxonomy$ASV_ID) + predictions[,colnames[colnames != "ASV_ID"]] <- NA + } } else { predictions <- data.frame(ASV_ID = taxonomy$ASV_ID) } diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 864d0802..666e3985 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -617,7 +617,7 @@ workflow AMPLISEQ { if ( params.sbdiexport ) { SBDIEXPORT ( ch_dada2_asv, ch_dada2_tax, ch_metadata ) ch_versions = ch_versions.mix(SBDIEXPORT.out.versions.first()) - SBDIEXPORTREANNOTATE ( ch_dada2_tax, ch_barrnapsummary ) + SBDIEXPORTREANNOTATE ( ch_dada2_tax, ch_barrnapsummary.ifEmpty([]) ) } CUSTOM_DUMPSOFTWAREVERSIONS ( From 95b101f30bb9ff5467fe11c5e3d614b830459bdb Mon Sep 17 00:00:00 2001 From: jtangrot Date: Mon, 13 Mar 2023 09:01:18 +0100 Subject: [PATCH 011/158] Add warning if barrnap gives no hits --- modules/local/barrnapsummary.nf | 7 +++++++ workflows/ampliseq.nf | 2 ++ 2 files changed, 9 insertions(+) diff --git a/modules/local/barrnapsummary.nf b/modules/local/barrnapsummary.nf index 13a38385..7d6ecbb8 100644 --- a/modules/local/barrnapsummary.nf +++ b/modules/local/barrnapsummary.nf @@ -11,6 +11,7 @@ process BARRNAPSUMMARY { output: path "summary.tsv" , emit: summary + path "*warning.txt" , emit: warning when: task.ext.when == null || task.ext.when @@ -21,6 +22,12 @@ process BARRNAPSUMMARY { """ summarize_barrnap.py $predictions + if [[ \$(wc -l < summary.tsv ) -le 1 ]]; then + touch WARNING_no_rRNA_found_warning.txt + else + touch no_warning.txt + fi + cat <<-END_VERSIONS > versions.yml "${task.process}": python: \$( python --version ) diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 666e3985..2e7ed7af 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -315,6 +315,7 @@ workflow AMPLISEQ { if (!params.skip_barrnap && params.filter_ssu) { BARRNAP ( ch_unfiltered_fasta ) BARRNAPSUMMARY ( BARRNAP.out.gff.collect() ) + BARRNAPSUMMARY.out.warning.subscribe { if ( it.baseName.toString().startsWith("WARNING") ) log.warn "Barrnap could not identify any rRNA in the ASV sequences, which seem to be non-ribosomal. We recommended to use the --skip_barrnap option for these sequences." } ch_barrnapsummary = BARRNAPSUMMARY.out.summary ch_versions = ch_versions.mix(BARRNAP.out.versions.ifEmpty(null)) FILTER_SSU ( DADA2_MERGE.out.fasta, DADA2_MERGE.out.asv, BARRNAPSUMMARY.out.summary ) @@ -325,6 +326,7 @@ workflow AMPLISEQ { } else if (!params.skip_barrnap && !params.filter_ssu) { BARRNAP ( ch_unfiltered_fasta ) BARRNAPSUMMARY ( BARRNAP.out.gff.collect() ) + BARRNAPSUMMARY.out.warning.subscribe { if ( it.baseName.toString().startsWith("WARNING") ) log.warn "Barrnap could not identify any rRNA in the ASV sequences, which seem to be non-ribosomal. We recommended to use the --skip_barrnap option for these sequences." } ch_barrnapsummary = BARRNAPSUMMARY.out.summary ch_versions = ch_versions.mix(BARRNAP.out.versions.ifEmpty(null)) ch_dada2_fasta = ch_unfiltered_fasta From da588e20dabb93ac88e070d37c162a3508684570 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Mon, 13 Mar 2023 09:31:55 +0100 Subject: [PATCH 012/158] Error if filter_ssu and no barrnap hits --- workflows/ampliseq.nf | 6 +++++- 1 file changed, 5 insertions(+), 1 deletion(-) diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 2e7ed7af..981f6590 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -315,7 +315,11 @@ workflow AMPLISEQ { if (!params.skip_barrnap && params.filter_ssu) { BARRNAP ( ch_unfiltered_fasta ) BARRNAPSUMMARY ( BARRNAP.out.gff.collect() ) - BARRNAPSUMMARY.out.warning.subscribe { if ( it.baseName.toString().startsWith("WARNING") ) log.warn "Barrnap could not identify any rRNA in the ASV sequences, which seem to be non-ribosomal. We recommended to use the --skip_barrnap option for these sequences." } + BARRNAPSUMMARY.out.warning.subscribe { + if ( it.baseName.toString().startsWith("WARNING") ) + log.error "Barrnap could not identify any rRNA in the ASV sequences! This will result in all sequences being removed with SSU filtering." + System.exit(1) + } ch_barrnapsummary = BARRNAPSUMMARY.out.summary ch_versions = ch_versions.mix(BARRNAP.out.versions.ifEmpty(null)) FILTER_SSU ( DADA2_MERGE.out.fasta, DADA2_MERGE.out.asv, BARRNAPSUMMARY.out.summary ) From c17ce2a1ba069f060dcb8b1a676c02173dbbc3ba Mon Sep 17 00:00:00 2001 From: jtangrot Date: Mon, 13 Mar 2023 13:12:54 +0100 Subject: [PATCH 013/158] Update changelog --- CHANGELOG.md | 2 ++ 1 file changed, 2 insertions(+) diff --git a/CHANGELOG.md b/CHANGELOG.md index a071b375..cfc05c29 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -11,6 +11,8 @@ and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0 ### `Fixed` +- [#551](https://github.com/nf-core/ampliseq/pull/551) - Handle empty barrnap results files + ### `Dependencies` ### `Removed` From 7a32b87aaffd5e5055e1a059b9519f1bf12d6e90 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Mon, 13 Mar 2023 13:29:56 +0100 Subject: [PATCH 014/158] Sort barrnap columns in summary --- bin/summarize_barrnap.py | 4 ++-- workflows/ampliseq.nf | 3 ++- 2 files changed, 4 insertions(+), 3 deletions(-) diff --git a/bin/summarize_barrnap.py b/bin/summarize_barrnap.py index 97cb31bb..c7690bbe 100755 --- a/bin/summarize_barrnap.py +++ b/bin/summarize_barrnap.py @@ -32,8 +32,8 @@ # Write results fh = open("summary.tsv", mode="w") -orglist = list(orgs) -header = list(orgs) +orglist = sorted(orgs) +header = sorted(orgs) header.insert(0, "ASV_ID") header.append("eval_method") fh.write("\t".join(header) + "\n") diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 981f6590..4994f4a3 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -316,9 +316,10 @@ workflow AMPLISEQ { BARRNAP ( ch_unfiltered_fasta ) BARRNAPSUMMARY ( BARRNAP.out.gff.collect() ) BARRNAPSUMMARY.out.warning.subscribe { - if ( it.baseName.toString().startsWith("WARNING") ) + if ( it.baseName.toString().startsWith("WARNING") ) { log.error "Barrnap could not identify any rRNA in the ASV sequences! This will result in all sequences being removed with SSU filtering." System.exit(1) + } } ch_barrnapsummary = BARRNAPSUMMARY.out.summary ch_versions = ch_versions.mix(BARRNAP.out.versions.ifEmpty(null)) From 36d1b55e40231d5ccf7d632d51ed749b28757abb Mon Sep 17 00:00:00 2001 From: jtangrot Date: Tue, 14 Mar 2023 08:38:26 +0100 Subject: [PATCH 015/158] Change wording in warning --- workflows/ampliseq.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 4994f4a3..6523d174 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -331,7 +331,7 @@ workflow AMPLISEQ { } else if (!params.skip_barrnap && !params.filter_ssu) { BARRNAP ( ch_unfiltered_fasta ) BARRNAPSUMMARY ( BARRNAP.out.gff.collect() ) - BARRNAPSUMMARY.out.warning.subscribe { if ( it.baseName.toString().startsWith("WARNING") ) log.warn "Barrnap could not identify any rRNA in the ASV sequences, which seem to be non-ribosomal. We recommended to use the --skip_barrnap option for these sequences." } + BARRNAPSUMMARY.out.warning.subscribe { if ( it.baseName.toString().startsWith("WARNING") ) log.warn "Barrnap could not identify any rRNA in the ASV sequences. We recommended to use the --skip_barrnap option for these sequences." } ch_barrnapsummary = BARRNAPSUMMARY.out.summary ch_versions = ch_versions.mix(BARRNAP.out.versions.ifEmpty(null)) ch_dada2_fasta = ch_unfiltered_fasta From 66f7157db272591cfca81ac67b694e8aa0feb1d5 Mon Sep 17 00:00:00 2001 From: daniel Date: Wed, 15 Mar 2023 14:45:06 +0100 Subject: [PATCH 016/158] fix hash and apostrophe in taxonomy strings for QIIME2 --- CHANGELOG.md | 1 + bin/parse_dada2_taxonomy.r | 2 +- 2 files changed, 2 insertions(+), 1 deletion(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index cfc05c29..18903dc6 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -12,6 +12,7 @@ and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0 ### `Fixed` - [#551](https://github.com/nf-core/ampliseq/pull/551) - Handle empty barrnap results files +- [#552](https://github.com/nf-core/ampliseq/pull/552) - Accept taxonomy strings that contain `#`,`'` ### `Dependencies` diff --git a/bin/parse_dada2_taxonomy.r b/bin/parse_dada2_taxonomy.r index 54568472..7ebd1ec2 100755 --- a/bin/parse_dada2_taxonomy.r +++ b/bin/parse_dada2_taxonomy.r @@ -11,7 +11,7 @@ tax_file <- args[1] OUT="tax.tsv" # read required files -tax = read.table(tax_file, header = TRUE, sep = "\t", stringsAsFactors = FALSE) +tax = read.table(tax_file, header = TRUE, sep = "\t", stringsAsFactors = FALSE, comment.char = '', quote = '') # Join columns 2:ncol(.) - 1, the taxonomy ranks (sequence is the last) r <- colnames(tax)[!colnames(tax) %in% c('ASV_ID', 'sequence')] From 4db5f777696301287742e06528af35ca2a57c1bb Mon Sep 17 00:00:00 2001 From: daniel Date: Wed, 15 Mar 2023 14:54:45 +0100 Subject: [PATCH 017/158] update CHANGELOG --- CHANGELOG.md | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index 18903dc6..a371e745 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -11,8 +11,8 @@ and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0 ### `Fixed` -- [#551](https://github.com/nf-core/ampliseq/pull/551) - Handle empty barrnap results files -- [#552](https://github.com/nf-core/ampliseq/pull/552) - Accept taxonomy strings that contain `#`,`'` +- [#553](https://github.com/nf-core/ampliseq/pull/553) - Handle empty barrnap results files +- [#554](https://github.com/nf-core/ampliseq/pull/554) - Accept taxonomy strings that contain `#`,`'` ### `Dependencies` From 0d3cc2c4a306398b5a0157e6fa11c2b2c8a1d368 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Fri, 24 Mar 2023 14:53:19 +0100 Subject: [PATCH 018/158] Use only --sintax_ref_taxonomy and remove --sintax parameter --- modules.json | 26 +++++++++++++++++++------- nextflow.config | 1 - nextflow_schema.json | 7 +------ workflows/ampliseq.nf | 27 +++++++++++++-------------- 4 files changed, 33 insertions(+), 28 deletions(-) diff --git a/modules.json b/modules.json index 382df979..31c011f5 100644 --- a/modules.json +++ b/modules.json @@ -8,35 +8,47 @@ "custom/dumpsoftwareversions": { "branch": "master", "git_sha": "c8e35eb2055c099720a75538d1b8adb3fb5a464c", - "installed_by": ["modules"] + "installed_by": [ + "modules" + ] }, "cutadapt": { "branch": "master", "git_sha": "c8e35eb2055c099720a75538d1b8adb3fb5a464c", - "installed_by": ["modules"] + "installed_by": [ + "modules" + ] }, "fastqc": { "branch": "master", "git_sha": "c8e35eb2055c099720a75538d1b8adb3fb5a464c", - "installed_by": ["modules"] + "installed_by": [ + "modules" + ] }, "multiqc": { "branch": "master", "git_sha": "ee80d14721e76e2e079103b8dcd5d57129e584ba", - "installed_by": ["modules"] + "installed_by": [ + "modules" + ] }, "vsearch/sintax": { "branch": "master", "git_sha": "0f8a77ff00e65eaeebc509b8156eaa983192474b", - "installed_by": ["modules"] + "installed_by": [ + "modules" + ] }, "vsearch/usearchglobal": { "branch": "master", "git_sha": "c8e35eb2055c099720a75538d1b8adb3fb5a464c", - "installed_by": ["modules"] + "installed_by": [ + "modules" + ] } } } } } -} +} \ No newline at end of file diff --git a/nextflow.config b/nextflow.config index 24708ea4..1e54da4f 100644 --- a/nextflow.config +++ b/nextflow.config @@ -46,7 +46,6 @@ params { its_partial = 0 picrust = false sbdiexport = false - sintax = false addsh = false dada_tax_agglom_min = 2 dada_tax_agglom_max = 6 diff --git a/nextflow_schema.json b/nextflow_schema.json index a2a4c43c..eeab4d28 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -281,9 +281,8 @@ }, "sintax_ref_taxonomy": { "type": "string", - "help_text": "Choose any of the supported databases, and optionally also specify the version. Database and version are separated by an equal sign (`=`, e.g. `coidb=221216`) . This will download the desired database and if needed format it to produce a file that is compatible with USEARCH sintax.\n\nThe following databases are supported:\n- UNITE - eukaryotic nuclear ribosomal ITS region - ITS\n- COIDB - eukaryotic Cytochrome Oxidase I (COI) from The Barcode of Life Data System (BOLD) - COI\n\nGenerally, using `coidb`, `unite-fungi`, or `unite-alleuk` will select the most recent supported version. For details on what values are valid, please either use an invalid value such as `x` (causing the pipeline to send an error message with a list of all valid values) or see `conf/ref_databases.config`.", + "help_text": "Choose any of the supported databases, and optionally also specify the version. Database and version are separated by an equal sign (`=`, e.g. `coidb=221216`) . This will download the desired database and initiate taxonomic classification with USEARCH sintax and the chosen database, which if needed is formatted it to produce a file that is compatible with USEARCH sintax.\n\nThe following databases are supported:\n- UNITE - eukaryotic nuclear ribosomal ITS region - ITS\n- COIDB - eukaryotic Cytochrome Oxidase I (COI) from The Barcode of Life Data System (BOLD) - COI\n\nGenerally, using `coidb`, `unite-fungi`, or `unite-alleuk` will select the most recent supported version. For details on what values are valid, please either use an invalid value such as `x` (causing the pipeline to send an error message with a list of all valid values) or see `conf/ref_databases.config`.", "description": "Name of supported database, and optionally also version number", - "default": "null", "enum": [ "coidb", "coidb=221216", @@ -295,10 +294,6 @@ "unite-alleuk" ] }, - "sintax": { - "type": "boolean", - "description": "If VSEARCH sintax should be used for taxonomic assingment of ASVs." - }, "addsh": { "type": "boolean", "description": "If ASVs should be assigned to UNITE species hypotheses (SHs). Only relevant for ITS data." diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 0d17cb19..04a6f970 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -478,20 +478,19 @@ workflow AMPLISEQ { } // Sintax - if (params.sintax) { - FORMAT_TAXONOMY_SINTAX ( ch_sintax_ref_taxonomy.collect() ) - ch_sintaxdb = FORMAT_TAXONOMY_SINTAX.out.db - ch_fasta - .map { - fasta -> - def meta = [:] - meta.id = "ASV_tax_sintax" - [ meta, fasta ] } - .set { ch_fasta_sintax } - VSEARCH_SINTAX( ch_fasta_sintax, ch_sintaxdb ) - ch_versions = ch_versions.mix(VSEARCH_SINTAX.out.versions) - } - + // This will only run if --sintax_ref_taxonomy is defined, i.e. the channel + // ch_sintax_ref_taxonomy is not empty + FORMAT_TAXONOMY_SINTAX ( ch_sintax_ref_taxonomy.collect() ) + ch_sintaxdb = FORMAT_TAXONOMY_SINTAX.out.db + ch_fasta + .map { + fasta -> + def meta = [:] + meta.id = "ASV_tax_sintax" + [ meta, fasta ] } + .set { ch_fasta_sintax } + VSEARCH_SINTAX( ch_fasta_sintax, ch_sintaxdb ) + ch_versions = ch_versions.mix(VSEARCH_SINTAX.out.versions) //QIIME2 if ( run_qiime2 ) { From 33e5e57cd91f55f745526836e9d962db78c1a7a0 Mon Sep 17 00:00:00 2001 From: daniel Date: Mon, 27 Mar 2023 15:33:13 +0200 Subject: [PATCH 019/158] add ref tax abbreviation to DADA2 output tables --- CHANGELOG.md | 2 ++ conf/modules.config | 2 +- docs/output.md | 16 ++++++++-------- modules/local/format_taxonomy.nf | 11 ++++++----- workflows/ampliseq.nf | 23 ++++++++++++----------- 5 files changed, 29 insertions(+), 25 deletions(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index a371e745..6d4e1ef4 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -9,6 +9,8 @@ and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0 ### `Changed` +- [#563](https://github.com/nf-core/ampliseq/pull/563) - Renamed DADA2 taxonomic classification files to include the chosen reference taxonomy abbreviation. + ### `Fixed` - [#553](https://github.com/nf-core/ampliseq/pull/553) - Handle empty barrnap results files diff --git a/conf/modules.config b/conf/modules.config index d0bf626a..26776202 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -335,7 +335,7 @@ process { publishDir = [ path: { "${params.outdir}/dada2" }, mode: params.publish_dir_mode, - pattern: "{ref_taxonomy.txt,ASV_tax.tsv,ASV_tax_species.tsv}" + pattern: "{ref_taxonomy*.txt,ASV_tax*.tsv,ASV_tax_species*.tsv}" ] } diff --git a/docs/output.md b/docs/output.md index 42d4eeec..8119757d 100644 --- a/docs/output.md +++ b/docs/output.md @@ -198,9 +198,9 @@ Files when _not_ using ITSx (default): Output files - `dada2/` - - `ASV_tax.tsv`: Taxonomic classification for each ASV sequence. - - `ASV_tax_species.tsv`: Exact species classification for each ASV sequence. - - `ref_taxonomy.txt`: Information about the used reference taxonomy, such as title, version, citation. + - `ASV_tax.*.tsv`: Taxonomic classification for each ASV sequence. + - `ASV_tax_species.*.tsv`: Exact species classification for each ASV sequence. + - `ref_taxonomy.*.txt`: Information about the used reference taxonomy, such as title, version, citation. @@ -210,11 +210,11 @@ Files when using ITSx: Output files - `dada2/` - - `ASV_ITS_tax.tsv`: Taxonomic classification with ITS region of each ASV sequence. - - `ASV_ITS_tax_species.tsv`: Exact species classification with ITS region of each ASV sequence. - - `ASV_tax.tsv`: Taxonomic classification of each ASV sequence, based on the ITS region. - - `ASV_tax_species.tsv`: Exact species classification of each ASV sequence, based on the ITS region. - - `ref_taxonomy.txt`: Information about the used reference taxonomy, such as title, version, citation. + - `ASV_ITS_tax.*.tsv`: Taxonomic classification with ITS region of each ASV sequence. + - `ASV_ITS_tax_species.*.tsv`: Exact species classification with ITS region of each ASV sequence. + - `ASV_tax.*.tsv`: Taxonomic classification of each ASV sequence, based on the ITS region. + - `ASV_tax_species.*.tsv`: Exact species classification of each ASV sequence, based on the ITS region. + - `ref_taxonomy.*.txt`: Information about the used reference taxonomy, such as title, version, citation. diff --git a/modules/local/format_taxonomy.nf b/modules/local/format_taxonomy.nf index 8e085a47..52935ddf 100644 --- a/modules/local/format_taxonomy.nf +++ b/modules/local/format_taxonomy.nf @@ -8,11 +8,12 @@ process FORMAT_TAXONOMY { input: path(database) + val(suffix) output: path( "*assignTaxonomy.fna*" ), emit: assigntax path( "*addSpecies.fna*") , emit: addspecies - path( "ref_taxonomy.txt") , emit: ref_tax_info + path( "ref_taxonomy.*.txt") , emit: ref_tax_info path "versions.yml" , emit: versions when: @@ -23,10 +24,10 @@ process FORMAT_TAXONOMY { ${params.dada_ref_databases[params.dada_ref_taxonomy]["fmtscript"]} #Giving out information - echo -e "--dada_ref_taxonomy: ${params.dada_ref_taxonomy}\n" >ref_taxonomy.txt - echo -e "Title: ${params.dada_ref_databases[params.dada_ref_taxonomy]["title"]}\n" >>ref_taxonomy.txt - echo -e "Citation: ${params.dada_ref_databases[params.dada_ref_taxonomy]["citation"]}\n" >>ref_taxonomy.txt - echo "All entries: ${params.dada_ref_databases[params.dada_ref_taxonomy]}" >>ref_taxonomy.txt + echo -e "--dada_ref_taxonomy: ${params.dada_ref_taxonomy}\n" >ref_taxonomy.${suffix}.txt + echo -e "Title: ${params.dada_ref_databases[params.dada_ref_taxonomy]["title"]}\n" >>ref_taxonomy.${suffix}.txt + echo -e "Citation: ${params.dada_ref_databases[params.dada_ref_taxonomy]["citation"]}\n" >>ref_taxonomy.${suffix}.txt + echo "All entries: ${params.dada_ref_databases[params.dada_ref_taxonomy]}" >>ref_taxonomy.${suffix}.txt cat <<-END_VERSIONS > versions.yml "${task.process}": diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 6523d174..e748d2dd 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -54,6 +54,7 @@ if (params.dada_ref_tax_custom) { } else if (params.dada_ref_taxonomy && !params.skip_taxonomy) { //standard ref taxonomy input from params.dada_ref_taxonomy & conf/ref_databases.config ch_dada_ref_taxonomy = Channel.fromList(params.dada_ref_databases[params.dada_ref_taxonomy]["file"]).map { file(it) } + val_dada_ref_taxonomy = params.dada_ref_taxonomy.replace('=','_').replace('.','_') if (params.addsh) { ch_shinfo = Channel.fromList(params.dada_ref_databases[params.dada_ref_taxonomy]["shfile"]).map { file(it) } } @@ -363,7 +364,7 @@ workflow AMPLISEQ { if (!params.skip_taxonomy) { if (!params.dada_ref_tax_custom) { //standard ref taxonomy input from conf/ref_databases.config - FORMAT_TAXONOMY ( ch_dada_ref_taxonomy.collect() ) + FORMAT_TAXONOMY ( ch_dada_ref_taxonomy.collect(), val_dada_ref_taxonomy ) ch_assigntax = FORMAT_TAXONOMY.out.assigntax ch_addspecies = FORMAT_TAXONOMY.out.addspecies } @@ -382,10 +383,10 @@ workflow AMPLISEQ { .set { ch_assigntax } } if (params.cut_its == "none") { - DADA2_TAXONOMY ( ch_fasta, ch_assigntax, 'ASV_tax.tsv', taxlevels ) + DADA2_TAXONOMY ( ch_fasta, ch_assigntax, "ASV_tax.${val_dada_ref_taxonomy}.tsv", taxlevels ) ch_versions = ch_versions.mix(DADA2_TAXONOMY.out.versions) if (!params.skip_dada_addspecies) { - DADA2_ADDSPECIES ( DADA2_TAXONOMY.out.rds, ch_addspecies, 'ASV_tax_species.tsv', taxlevels ) + DADA2_ADDSPECIES ( DADA2_TAXONOMY.out.rds, ch_addspecies, "ASV_tax_species.${val_dada_ref_taxonomy}.tsv", taxlevels ) if ( params.addsh ) { ch_fasta .map { @@ -396,7 +397,7 @@ workflow AMPLISEQ { .set { ch_fasta_map } VSEARCH_USEARCHGLOBAL( ch_fasta_map, ch_assigntax, vsearch_cutoff, 'blast6out', "" ) ch_versions = ch_versions.mix(VSEARCH_USEARCHGLOBAL.out.versions.ifEmpty(null)) - ASSIGNSH( DADA2_ADDSPECIES.out.tsv, ch_shinfo.collect(), VSEARCH_USEARCHGLOBAL.out.txt, 'ASV_tax_species_SH.tsv') + ASSIGNSH( DADA2_ADDSPECIES.out.tsv, ch_shinfo.collect(), VSEARCH_USEARCHGLOBAL.out.txt, "ASV_tax_species_SH.${val_dada_ref_taxonomy}.tsv") ch_versions = ch_versions.mix(ASSIGNSH.out.versions.ifEmpty(null)) ch_dada2_tax = ASSIGNSH.out.tsv } else { @@ -413,7 +414,7 @@ workflow AMPLISEQ { .set { ch_fasta_map } VSEARCH_USEARCHGLOBAL( ch_fasta_map, ch_assigntax, vsearch_cutoff, 'blast6out', "" ) ch_versions = ch_versions.mix(VSEARCH_USEARCHGLOBAL.out.versions.ifEmpty(null)) - ASSIGNSH( DADA2_TAXONOMY.out.tsv, ch_shinfo.collect(), VSEARCH_USEARCHGLOBAL.out.txt, 'ASV_tax_SH.tsv') + ASSIGNSH( DADA2_TAXONOMY.out.tsv, ch_shinfo.collect(), VSEARCH_USEARCHGLOBAL.out.txt, "ASV_tax_SH.${val_dada_ref_taxonomy}.tsv") ch_versions = ch_versions.mix(ASSIGNSH.out.versions.ifEmpty(null)) ch_dada2_tax = ASSIGNSH.out.tsv } else { @@ -434,13 +435,13 @@ workflow AMPLISEQ { ITSX_CUTASV ( ch_fasta, outfile ) ch_versions = ch_versions.mix(ITSX_CUTASV.out.versions.ifEmpty(null)) ch_cut_fasta = ITSX_CUTASV.out.fasta - DADA2_TAXONOMY ( ch_cut_fasta, ch_assigntax, 'ASV_ITS_tax.tsv', taxlevels ) + DADA2_TAXONOMY ( ch_cut_fasta, ch_assigntax, "ASV_ITS_tax.${val_dada_ref_taxonomy}.tsv", taxlevels ) ch_versions = ch_versions.mix(DADA2_TAXONOMY.out.versions) FORMAT_TAXRESULTS_STD ( DADA2_TAXONOMY.out.tsv, ch_fasta, 'ASV_tax.tsv' ) ch_versions = ch_versions.mix( FORMAT_TAXRESULTS_STD.out.versions.ifEmpty(null) ) if (!params.skip_dada_addspecies) { - DADA2_ADDSPECIES ( DADA2_TAXONOMY.out.rds, ch_addspecies, 'ASV_ITS_tax_species.tsv', taxlevels ) - FORMAT_TAXRESULTS_ADDSP ( DADA2_ADDSPECIES.out.tsv, ch_fasta, 'ASV_tax_species.tsv' ) + DADA2_ADDSPECIES ( DADA2_TAXONOMY.out.rds, ch_addspecies, "ASV_ITS_tax_species.${val_dada_ref_taxonomy}.tsv", taxlevels ) + FORMAT_TAXRESULTS_ADDSP ( DADA2_ADDSPECIES.out.tsv, ch_fasta, "ASV_tax_species.${val_dada_ref_taxonomy}.tsv" ) if ( params.addsh ) { ch_cut_fasta .map { @@ -451,13 +452,13 @@ workflow AMPLISEQ { .set { ch_cut_fasta } VSEARCH_USEARCHGLOBAL( ch_cut_fasta, ch_assigntax, vsearch_cutoff, 'blast6out', "" ) ch_versions = ch_versions.mix(VSEARCH_USEARCHGLOBAL.out.versions.ifEmpty(null)) - ASSIGNSH( FORMAT_TAXRESULTS_ADDSP.out.tsv, ch_shinfo.collect(), VSEARCH_USEARCHGLOBAL.out.txt, 'ASV_tax_species_SH.tsv') + ASSIGNSH( FORMAT_TAXRESULTS_ADDSP.out.tsv, ch_shinfo.collect(), VSEARCH_USEARCHGLOBAL.out.txt, "ASV_tax_species_SH.${val_dada_ref_taxonomy}.tsv") ch_versions = ch_versions.mix(ASSIGNSH.out.versions.ifEmpty(null)) ch_dada2_tax = ASSIGNSH.out.tsv } else { ch_dada2_tax = FORMAT_TAXRESULTS_ADDSP.out.tsv } - } else { + } else { if ( params.addsh ) { ch_cut_fasta .map { @@ -468,7 +469,7 @@ workflow AMPLISEQ { .set { ch_cut_fasta } VSEARCH_USEARCHGLOBAL( ch_cut_fasta, ch_assigntax, vsearch_cutoff, 'blast6out', "" ) ch_versions = ch_versions.mix(VSEARCH_USEARCHGLOBAL.out.versions.ifEmpty(null)) - ASSIGNSH( FORMAT_TAXRESULTS_STD.out.tsv, ch_shinfo.collect(), VSEARCH_USEARCHGLOBAL.out.txt, 'ASV_tax_SH.tsv') + ASSIGNSH( FORMAT_TAXRESULTS_STD.out.tsv, ch_shinfo.collect(), VSEARCH_USEARCHGLOBAL.out.txt, "ASV_tax_SH.${val_dada_ref_taxonomy}.tsv") ch_versions = ch_versions.mix(ASSIGNSH.out.versions.ifEmpty(null)) ch_dada2_tax = ASSIGNSH.out.tsv } else { From 941f098729ddef6bb0d8a2be25163e422b70aac2 Mon Sep 17 00:00:00 2001 From: daniel Date: Mon, 27 Mar 2023 15:49:10 +0200 Subject: [PATCH 020/158] fix when dada_ref_taxonomy is false --- workflows/ampliseq.nf | 5 ++++- 1 file changed, 4 insertions(+), 1 deletion(-) diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index e748d2dd..05483f09 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -58,7 +58,10 @@ if (params.dada_ref_tax_custom) { if (params.addsh) { ch_shinfo = Channel.fromList(params.dada_ref_databases[params.dada_ref_taxonomy]["shfile"]).map { file(it) } } -} else { ch_dada_ref_taxonomy = Channel.empty() } +} else { + ch_dada_ref_taxonomy = Channel.empty() + val_dada_ref_taxonomy = "none" +} if (params.qiime_ref_taxonomy && !params.skip_taxonomy && !params.classifier) { ch_qiime_ref_taxonomy = Channel.fromList(params.qiime_ref_databases[params.qiime_ref_taxonomy]["file"]).map { file(it) } From fa84fb4d908eb3cb00b6b3ad037abe203155d402 Mon Sep 17 00:00:00 2001 From: Daniel Straub <42973691+d4straub@users.noreply.github.com> Date: Mon, 27 Mar 2023 15:51:44 +0200 Subject: [PATCH 021/158] Update modules/local/format_taxonomy.nf Co-authored-by: Daniel Lundin --- modules/local/format_taxonomy.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/modules/local/format_taxonomy.nf b/modules/local/format_taxonomy.nf index 52935ddf..ded8a104 100644 --- a/modules/local/format_taxonomy.nf +++ b/modules/local/format_taxonomy.nf @@ -13,7 +13,7 @@ process FORMAT_TAXONOMY { output: path( "*assignTaxonomy.fna*" ), emit: assigntax path( "*addSpecies.fna*") , emit: addspecies - path( "ref_taxonomy.*.txt") , emit: ref_tax_info + path( "ref_taxonomy.*.txt") , emit: ref_tax_info path "versions.yml" , emit: versions when: From 03402db2903b5ec6f16e7730eff1ae2ed34a88ea Mon Sep 17 00:00:00 2001 From: daniel Date: Mon, 27 Mar 2023 16:01:55 +0200 Subject: [PATCH 022/158] fix with custom ref tax database --- workflows/ampliseq.nf | 1 + 1 file changed, 1 insertion(+) diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 05483f09..f4c8902d 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -51,6 +51,7 @@ if (params.dada_ref_tax_custom) { ch_addspecies = Channel.fromPath("${params.dada_ref_tax_custom_sp}", checkIfExists: true) } ch_dada_ref_taxonomy = Channel.empty() + val_dada_ref_taxonomy = "user" } else if (params.dada_ref_taxonomy && !params.skip_taxonomy) { //standard ref taxonomy input from params.dada_ref_taxonomy & conf/ref_databases.config ch_dada_ref_taxonomy = Channel.fromList(params.dada_ref_databases[params.dada_ref_taxonomy]["file"]).map { file(it) } From 02b44e689e0bf51ac89421e2d6e409411479dade Mon Sep 17 00:00:00 2001 From: daniel Date: Tue, 28 Mar 2023 14:44:27 +0200 Subject: [PATCH 023/158] add phylogenetic placement --- .github/workflows/ci.yml | 1 + CHANGELOG.md | 2 + CITATIONS.md | 20 +++ README.md | 3 +- conf/modules.config | 155 ++++++++++++++++++ conf/test_pplace.config | 46 ++++++ docs/output.md | 19 +++ lib/WorkflowAmpliseq.groovy | 11 ++ modules.json | 59 +++++++ modules/nf-core/epang/place/main.nf | 54 ++++++ modules/nf-core/epang/place/meta.yml | 66 ++++++++ modules/nf-core/epang/split/main.nf | 37 +++++ modules/nf-core/epang/split/meta.yml | 50 ++++++ modules/nf-core/gappa/examineassign/main.nf | 42 +++++ modules/nf-core/gappa/examineassign/meta.yml | 55 +++++++ modules/nf-core/gappa/examinegraft/main.nf | 37 +++++ modules/nf-core/gappa/examinegraft/meta.yml | 41 +++++ modules/nf-core/gappa/examineheattree/main.nf | 45 +++++ .../nf-core/gappa/examineheattree/meta.yml | 64 ++++++++ modules/nf-core/hmmer/eslalimask/main.nf | 56 +++++++ modules/nf-core/hmmer/eslalimask/meta.yml | 105 ++++++++++++ modules/nf-core/hmmer/eslreformat/main.nf | 38 +++++ modules/nf-core/hmmer/eslreformat/meta.yml | 41 +++++ modules/nf-core/hmmer/hmmalign/main.nf | 35 ++++ modules/nf-core/hmmer/hmmalign/meta.yml | 46 ++++++ modules/nf-core/hmmer/hmmbuild/main.nf | 44 +++++ modules/nf-core/hmmer/hmmbuild/meta.yml | 44 +++++ modules/nf-core/mafft/main.nf | 38 +++++ modules/nf-core/mafft/meta.yml | 46 ++++++ nextflow.config | 7 + nextflow_schema.json | 31 ++++ .../nf-core/fasta_newick_epang_gappa/main.nf | 154 +++++++++++++++++ .../nf-core/fasta_newick_epang_gappa/meta.yml | 97 +++++++++++ workflows/ampliseq.nf | 25 +++ 34 files changed, 1613 insertions(+), 1 deletion(-) create mode 100644 conf/test_pplace.config create mode 100644 modules/nf-core/epang/place/main.nf create mode 100644 modules/nf-core/epang/place/meta.yml create mode 100644 modules/nf-core/epang/split/main.nf create mode 100644 modules/nf-core/epang/split/meta.yml create mode 100644 modules/nf-core/gappa/examineassign/main.nf create mode 100644 modules/nf-core/gappa/examineassign/meta.yml create mode 100644 modules/nf-core/gappa/examinegraft/main.nf create mode 100644 modules/nf-core/gappa/examinegraft/meta.yml create mode 100644 modules/nf-core/gappa/examineheattree/main.nf create mode 100644 modules/nf-core/gappa/examineheattree/meta.yml create mode 100644 modules/nf-core/hmmer/eslalimask/main.nf create mode 100644 modules/nf-core/hmmer/eslalimask/meta.yml create mode 100644 modules/nf-core/hmmer/eslreformat/main.nf create mode 100644 modules/nf-core/hmmer/eslreformat/meta.yml create mode 100644 modules/nf-core/hmmer/hmmalign/main.nf create mode 100644 modules/nf-core/hmmer/hmmalign/meta.yml create mode 100644 modules/nf-core/hmmer/hmmbuild/main.nf create mode 100644 modules/nf-core/hmmer/hmmbuild/meta.yml create mode 100644 modules/nf-core/mafft/main.nf create mode 100644 modules/nf-core/mafft/meta.yml create mode 100644 subworkflows/nf-core/fasta_newick_epang_gappa/main.nf create mode 100644 subworkflows/nf-core/fasta_newick_epang_gappa/meta.yml diff --git a/.github/workflows/ci.yml b/.github/workflows/ci.yml index b00c99b6..8c235eda 100644 --- a/.github/workflows/ci.yml +++ b/.github/workflows/ci.yml @@ -57,6 +57,7 @@ jobs: test_fasta, test_reftaxcustom, test_novaseq, + test_pplace, ] steps: - name: Check out pipeline code diff --git a/CHANGELOG.md b/CHANGELOG.md index a371e745..b51e2747 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -7,6 +7,8 @@ and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0 ### `Added` +- [#564](https://github.com/nf-core/ampliseq/pull/564) - Added phylogenetic placement + ### `Changed` ### `Fixed` diff --git a/CITATIONS.md b/CITATIONS.md index 97604283..832eee3c 100644 --- a/CITATIONS.md +++ b/CITATIONS.md @@ -67,6 +67,26 @@ > Sundh J, Manoharan L, Iwaszkiewicz-Eggebrecht E, Miraldo A, Andersson A, Ronquist F. COI reference sequences from BOLD DB. doi: https://doi.org/10.17044/scilifelab.20514192.v2. +### Phylogenetic placement + +- nf-core/phyloplace (https://github.com/nf-core/phyloplace, https://nf-co.re/phyloplace) was originally written by Daniel Lundin. + +- [HMMER](https://pubmed.ncbi.nlm.nih.gov/22039361/) + + > Eddy, Sean R. “Accelerated Profile HMM Searches.” PLoS Comput Biol 7, no. 10 (October 20, 2011): e1002195. https://doi.org/10.1371/journal.pcbi.1002195. + +- [MAFFT](https://pubmed.ncbi.nlm.nih.gov/12136088/) + + > Katoh, Kazutaka, Kazuharu Misawa, Kei‐ichi Kuma, and Takashi Miyata. “MAFFT: A Novel Method for Rapid Multiple Sequence Alignment Based on Fast Fourier Transform.” Nucleic Acids Research 30, no. 14 (July 15, 2002): 3059–66. https://doi.org/10.1093/nar/gkf436. + +- [EPA-NG](https://pubmed.ncbi.nlm.nih.gov/30165689/) + + > Barbera, Pierre, Alexey M Kozlov, Lucas Czech, Benoit Morel, Diego Darriba, Tomáš Flouri, and Alexandros Stamatakis. “EPA-Ng: Massively Parallel Evolutionary Placement of Genetic Sequences.” Systematic Biology 68, no. 2 (March 1, 2019): 365–69. https://doi.org/10.1093/sysbio/syy054. + +- [Gappa](https://pubmed.ncbi.nlm.nih.gov/32016344/) + + > Czech, Lucas, Pierre Barbera, and Alexandros Stamatakis. “Genesis and Gappa: Processing, Analyzing and Visualizing Phylogenetic (Placement) Data.” Bioinformatics 36, no. 10 (May 1, 2020): 3263–65. https://doi.org/10.1093/bioinformatics/btaa070. + ### Downstream analysis - [QIIME2](https://pubmed.ncbi.nlm.nih.gov/31341288/) diff --git a/README.md b/README.md index 60f56f01..778643c1 100644 --- a/README.md +++ b/README.md @@ -19,7 +19,7 @@ ## Introduction -**nfcore/ampliseq** is a bioinformatics analysis pipeline used for amplicon sequencing, supporting denoising of any amplicon and, currently, taxonomic assignment of 16S, ITS, CO1 and 18S amplicons. Supported is paired-end Illumina or single-end Illumina, PacBio and IonTorrent data. Default is the analysis of 16S rRNA gene amplicons sequenced paired-end with Illumina. +**nfcore/ampliseq** is a bioinformatics analysis pipeline used for amplicon sequencing, supporting denoising of any amplicon and, currently, taxonomic assignment of 16S, ITS, CO1 and 18S amplicons. Phylogenetic placement is also possible. Supported is paired-end Illumina or single-end Illumina, PacBio and IonTorrent data. Default is the analysis of 16S rRNA gene amplicons sequenced paired-end with Illumina.

nf-core/ampliseq workflow overview @@ -37,6 +37,7 @@ By default, the pipeline currently performs the following: - Trimming of reads ([Cutadapt](https://journal.embnet.org/index.php/embnetjournal/article/view/200)) - Infer Amplicon Sequence Variants (ASVs) ([DADA2](https://doi.org/10.1038/nmeth.3869)) - Predict whether ASVs are ribosomal RNA sequences ([Barrnap](https://github.com/tseemann/barrnap)) +- Phylogenetic placement ([EPA-NG](https://github.com/Pbdas/epa-ng)) - Taxonomical classification using DADA2 or [QIIME2](https://www.nature.com/articles/s41587-019-0209-9) - Excludes unwanted taxa, produces absolute and relative feature/taxa count tables and plots, plots alpha rarefaction curves, computes alpha and beta diversity indices and plots thereof ([QIIME2](https://www.nature.com/articles/s41587-019-0209-9)) - Calls differentially abundant taxa ([ANCOM](https://www.ncbi.nlm.nih.gov/pubmed/26028277)) diff --git a/conf/modules.config b/conf/modules.config index d0bf626a..bc4b2265 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -418,6 +418,161 @@ process { ] } + withName: HMMER_HMMBUILD { + ext.prefix = { "${meta.id}.ref" } + publishDir = [ + path: { "${params.outdir}/pplace/${task.process.tokenize(':')[-1].tokenize('_')[0].toLowerCase()}" }, + mode: params.publish_dir_mode, + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + ] + } + + withName: HMMER_UNALIGNREF { + ext.prefix = { "${meta.id}.ref.unaligned" } + ext.args = "--gapsym=- afa" + ext.postprocessing = '| sed "/^>/!s/-//g"' + publishDir = [ + path: { "${params.outdir}/pplace/${task.process.tokenize(':')[-1].tokenize('_')[0].toLowerCase()}" }, + mode: params.publish_dir_mode, + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + ] + } + + withName: HMMER_HMMALIGNREF { + ext.prefix = { "${meta.id}.ref.hmmalign" } + publishDir = [ + path: { "${params.outdir}/pplace/${task.process.tokenize(':')[-1].tokenize('_')[0].toLowerCase()}" }, + mode: params.publish_dir_mode, + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + ] + } + + withName: HMMER_HMMALIGNQUERY { + ext.prefix = { "${meta.id}.query.hmmalign" } + publishDir = [ + path: { "${params.outdir}/pplace/${task.process.tokenize(':')[-1].tokenize('_')[0].toLowerCase()}" }, + mode: params.publish_dir_mode, + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + ] + } + + withName: 'HMMER_MASK.*' { + ext.args = '--rf-is-mask' + publishDir = [ + path: { "${params.outdir}/pplace/${task.process.tokenize(':')[-1].tokenize('_')[0].toLowerCase()}" }, + mode: params.publish_dir_mode, + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + ] + } + + withName: 'HMMER_MASKQUERY.*' { + ext.prefix = { "${meta.id}.query.hmmalign" } + publishDir = [ + path: { "${params.outdir}/pplace/${task.process.tokenize(':')[-1].tokenize('_')[0].toLowerCase()}" }, + mode: params.publish_dir_mode, + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + ] + } + + withName: 'HMMER_MASKREF.*' { + ext.prefix = { "${meta.id}.ref.hmmalign" } + publishDir = [ + path: { "${params.outdir}/pplace/${task.process.tokenize(':')[-1].tokenize('_')[0].toLowerCase()}" }, + mode: params.publish_dir_mode, + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + ] + } + + withName: 'HMMER_AFAFORMATQUERY.*' { + ext.prefix = { "${meta.id}.query.hmmalign.masked" } + ext.args = 'afa' + publishDir = [ + path: { "${params.outdir}/pplace/${task.process.tokenize(':')[-1].tokenize('_')[0].toLowerCase()}" }, + mode: params.publish_dir_mode, + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + ] + } + + withName: 'HMMER_AFAFORMATREF.*' { + ext.prefix = { "${meta.id}.ref.hmmalign.masked" } + ext.args = 'afa' + publishDir = [ + path: { "${params.outdir}/pplace/${task.process.tokenize(':')[-1].tokenize('_')[0].toLowerCase()}" }, + mode: params.publish_dir_mode, + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + ] + } + + withName: 'MAFFT' { + ext.args = '--keeplength' + publishDir = [ + path: { "${params.outdir}/pplace/${task.process.tokenize(':')[-1].tokenize('_')[0].toLowerCase()}" }, + mode: params.publish_dir_mode, + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + ] + } + + withName: 'EPANG_PLACE' { + ext.args = { "--model ${meta.model}" } + publishDir = [ + path: { "${params.outdir}/pplace/${task.process.tokenize(':')[-1].tokenize('_')[0].toLowerCase()}" }, + mode: params.publish_dir_mode, + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + ] + } + + withName: 'GAPPA_GRAFT' { + ext.prefix = { "${meta.id}.graft" } + //test_pplace.graft.test_pplace.epa_result.newick + publishDir = [ + [ + path: { "${params.outdir}/pplace" }, + mode: params.publish_dir_mode, + pattern: "*.newick" + ], + [ + path: { "${params.outdir}/pplace/${task.process.tokenize(':')[-1].tokenize('_')[0].toLowerCase()}" }, + mode: params.publish_dir_mode, + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + ] + ] + } + + withName: 'GAPPA_ASSIGN' { + ext.prefix = { "${meta.id}.taxonomy" } + ext.args = "--per-query-results --krona --sativa" + ext.when = { taxonomy } + publishDir = [ + [ + path: { "${params.outdir}/pplace" }, + mode: params.publish_dir_mode, + pattern: "*.taxonomy.per_query.tsv" + ], + [ + path: { "${params.outdir}/pplace/${task.process.tokenize(':')[-1].tokenize('_')[0].toLowerCase()}" }, + mode: params.publish_dir_mode, + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + ] + ] + } + + withName: 'GAPPA_HEATTREE' { + ext.prefix = { "${meta.id}.heattree" } + ext.args = "--write-nexus-tree --write-phyloxml-tree --write-svg-tree" + publishDir = [ + [ + path: { "${params.outdir}/pplace" }, + mode: params.publish_dir_mode, + pattern: "*.tree.svg" + ], + [ + path: { "${params.outdir}/pplace/${task.process.tokenize(':')[-1].tokenize('_')[0].toLowerCase()}" }, + mode: params.publish_dir_mode, + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + ] + ] + } + withName: 'QIIME2_INASV|QIIME2_INSEQ|QIIME2_INTAX' { publishDir = [ path: { "${params.outdir}/qiime2/input" }, diff --git a/conf/test_pplace.config b/conf/test_pplace.config new file mode 100644 index 00000000..ade2a9e3 --- /dev/null +++ b/conf/test_pplace.config @@ -0,0 +1,46 @@ +/* +~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ + Nextflow config file for running minimal tests +~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ + Defines input files and everything required to run a fast and simple pipeline test. + + Use as follows: + nextflow run nf-core/ampliseq -profile test_pplace, --outdir + +---------------------------------------------------------------------------------------- +*/ + +params { + config_profile_name = 'Test profile' + config_profile_description = 'Minimal test dataset to check pipeline function' + + // Limit resources so that this can run on GitHub Actions + max_cpus = 2 + max_memory = '6.GB' + max_time = '6.h' + + // Input data + FW_primer = "GTGYCAGCMGCCGCGGTAA" + RV_primer = "GGACTACNVGGGTWTCTAAT" + input = "https://raw.githubusercontent.com/nf-core/test-datasets/ampliseq/samplesheets/Samplesheet.tsv" + metadata = "https://raw.githubusercontent.com/nf-core/test-datasets/ampliseq/samplesheets/Metadata.tsv" + dada_ref_taxonomy = false + qiime_ref_taxonomy = "greengenes85" + filter_ssu = "bac" + + //this is to remove low abundance ASVs to reduce runtime of downstream processes + min_samples = 2 + min_frequency = 10 + + //pplace + pplace_tree = "https://github.com/nf-core/test-datasets/raw/phyloplace/testdata/cyanos_16s.newick" + pplace_aln = "https://github.com/nf-core/test-datasets/raw/phyloplace/testdata/cyanos_16s.alnfna" + pplace_model = "GTR+F+I+I+R3" + pplace_taxonomy = "https://github.com/nf-core/test-datasets/raw/phyloplace/testdata/cyanos_16s.taxonomy.tsv" + pplace_name = "test_pplace" + + + //Skip some steps to reduce runtime + skip_alpha_rarefaction = true + skip_fastqc = true +} diff --git a/docs/output.md b/docs/output.md index 42d4eeec..da0b99c5 100644 --- a/docs/output.md +++ b/docs/output.md @@ -22,6 +22,7 @@ The pipeline is built using [Nextflow](https://www.nextflow.io/) and processes d - [ITSx](#itsx) - Optionally, the ITS region can be extracted - [Taxonomic classification with DADA2](#taxonomic-classification-with-dada2) - Taxonomic classification of (filtered) ASVs - [assignSH](#assignsh) - Optionally, a UNITE species hypothesis (SH) can be added to the taxonomy +- [Phlogenetic placement and taxonomic classification](#phylogenetic-placement-and-taxonomic-classification) - Placing ASVs into a phyloenetic tree - [QIIME2](#qiime2) - Secondary analysis - [Taxonomic classification](#taxonomic-classification) - Taxonomical classification of ASVs - [Abundance tables](#abundance-tables) - Exported abundance tables @@ -231,6 +232,24 @@ Optionally, a UNITE species hypothesis (SH) can be added to the taxonomy. In sho +### Phlogenetic placement and taxonomic classification + +Phylogenetic placement allows to graft sequences onto a phylogenetic reference tree and retrieve taxonomic annotations. The reference tree can be made of whole genomes (or other evolutionary information) that can yield more relevant information than a de-novo calculated phylogenetic tree from short amplicon sequences. On providing required reference files, ASV sequences are aligned to the reference alignment with either [HMMER](http://hmmer.org/) (default) or [MAFFT](https://mafft.cbrc.jp/alignment/software/), phylogenetic placement of query sequences is performed with [EPA-NG](https://github.com/Pbdas/epa-ng), and finally a number of summary operations are performed with [Gappa](https://github.com/lczech/gappa) after placement. This uses code (subworkflow) from [nf-core/phyloplace](https://nf-co.re/phyloplace) and its [subworkflow](https://github.com/nf-core/modules/tree/master/subworkflows/nf-core/fasta_newick_epang_gappa), therefore its detailed documentation also applies here. + +

+Output files + +- `pplace/` + - `*.graft.*.epa_result.newick`: Full phylogeny with query sequences grafted on to the reference phylogeny, in newick format. + - `*.taxonomy.per_query.tsv`: Tab separated file with taxonomy information per query from classification by `gappa examine examinassign` + - `*.heattree.tree.svg`: Heattree in SVG format from calling `gappa examine heattree`, see [Gappa documentation](https://github.com/Pbdas/epa-ng/blob/master/README.md) for details. + - `pplace/hmmer/`: Contains intermediatary files if HMMER is used + - `pplace/mafft/`: Contains intermediatary files if MAFFT is used + - `pplace/epang/`: Output files from EPA-NG. + - `pplace/gappa/`: Gappa output described in the [Gappa documentation](https://github.com/Pbdas/epa-ng/blob/master/README.md). + +
+ ### QIIME2 **Quantitative Insights Into Microbial Ecology 2** ([QIIME2](https://qiime2.org/)) is a next-generation microbiome bioinformatics platform and the successor of the widely used [QIIME1](https://www.nature.com/articles/nmeth.f.303). diff --git a/lib/WorkflowAmpliseq.groovy b/lib/WorkflowAmpliseq.groovy index 0162d5a4..0ee13f30 100755 --- a/lib/WorkflowAmpliseq.groovy +++ b/lib/WorkflowAmpliseq.groovy @@ -63,6 +63,17 @@ class WorkflowAmpliseq { System.exit(1) } + if (params.pplace_tree) { + if (!params.pplace_aln) { + log.error "Missing parameter: Phylogenetic placement requires in addition to `--pplace_tree` also `--pplace_aln`." + System.exit(1) + } + if (!params.pplace_model) { + log.error "Missing parameter: Phylogenetic placement requires in addition to `--pplace_tree` also `--pplace_model`." + System.exit(1) + } + } + if (params.dada_assign_taxlevels && params.sbdiexport) { log.error "Incompatible parameters: `--sbdiexport` expects specific taxonomics ranks (default) and therefore excludes modifying those using `--dada_assign_taxlevels`." System.exit(1) diff --git a/modules.json b/modules.json index c3b153ee..1f08e52c 100644 --- a/modules.json +++ b/modules.json @@ -15,11 +15,61 @@ "git_sha": "c8e35eb2055c099720a75538d1b8adb3fb5a464c", "installed_by": ["modules"] }, + "epang/place": { + "branch": "master", + "git_sha": "0f8a77ff00e65eaeebc509b8156eaa983192474b", + "installed_by": ["fasta_newick_epang_gappa"] + }, + "epang/split": { + "branch": "master", + "git_sha": "0f8a77ff00e65eaeebc509b8156eaa983192474b", + "installed_by": ["fasta_newick_epang_gappa"] + }, "fastqc": { "branch": "master", "git_sha": "c8e35eb2055c099720a75538d1b8adb3fb5a464c", "installed_by": ["modules"] }, + "gappa/examineassign": { + "branch": "master", + "git_sha": "0f8a77ff00e65eaeebc509b8156eaa983192474b", + "installed_by": ["fasta_newick_epang_gappa"] + }, + "gappa/examinegraft": { + "branch": "master", + "git_sha": "0f8a77ff00e65eaeebc509b8156eaa983192474b", + "installed_by": ["fasta_newick_epang_gappa"] + }, + "gappa/examineheattree": { + "branch": "master", + "git_sha": "0f8a77ff00e65eaeebc509b8156eaa983192474b", + "installed_by": ["fasta_newick_epang_gappa"] + }, + "hmmer/eslalimask": { + "branch": "master", + "git_sha": "0f8a77ff00e65eaeebc509b8156eaa983192474b", + "installed_by": ["fasta_newick_epang_gappa"] + }, + "hmmer/eslreformat": { + "branch": "master", + "git_sha": "0f8a77ff00e65eaeebc509b8156eaa983192474b", + "installed_by": ["fasta_newick_epang_gappa"] + }, + "hmmer/hmmalign": { + "branch": "master", + "git_sha": "0f8a77ff00e65eaeebc509b8156eaa983192474b", + "installed_by": ["fasta_newick_epang_gappa"] + }, + "hmmer/hmmbuild": { + "branch": "master", + "git_sha": "c8e35eb2055c099720a75538d1b8adb3fb5a464c", + "installed_by": ["fasta_newick_epang_gappa"] + }, + "mafft": { + "branch": "master", + "git_sha": "b265b4ff6a35b133b963b4eaddfca0ffb3395236", + "installed_by": ["fasta_newick_epang_gappa"] + }, "multiqc": { "branch": "master", "git_sha": "ee80d14721e76e2e079103b8dcd5d57129e584ba", @@ -31,6 +81,15 @@ "installed_by": ["modules"] } } + }, + "subworkflows": { + "nf-core": { + "fasta_newick_epang_gappa": { + "branch": "master", + "git_sha": "6ad90f5583fb375c60a913a24ed1c79339efc019", + "installed_by": ["subworkflows"] + } + } } } } diff --git a/modules/nf-core/epang/place/main.nf b/modules/nf-core/epang/place/main.nf new file mode 100644 index 00000000..35525d31 --- /dev/null +++ b/modules/nf-core/epang/place/main.nf @@ -0,0 +1,54 @@ +process EPANG_PLACE { + tag "$meta.id" + label 'process_high' + + conda "bioconda::epa-ng=0.3.8" + container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? + 'https://depot.galaxyproject.org/singularity/epa-ng:0.3.8--h9a82719_1': + 'quay.io/biocontainers/epa-ng:0.3.8--h9a82719_1' }" + + input: + tuple val(meta), path(queryaln), path(referencealn), path(referencetree) + path bfastfile + path binaryfile + + output: + tuple val(meta), path("./.") , emit: epang , optional: true + tuple val(meta), path("*.epa_result.jplace.gz"), emit: jplace , optional: true + path "*.epa_info.log" , emit: log + path "versions.yml" , emit: versions + + when: + task.ext.when == null || task.ext.when + + script: + def args = task.ext.args ?: '' + def prefix = task.ext.prefix ?: "${meta.id}" + def queryarg = queryaln ? "--query $queryaln" : "" + def refalnarg = referencealn ? "--ref-msa $referencealn" : "" + def reftreearg = referencetree ? "--tree $referencetree" : "" + def bfastarg = bfastfile ? "--bfast $bfastfile" : "" + def binaryarg = binaryfile ? "--binary $binaryfile" : "" + if ( binaryfile && ( referencealn || referencetree ) ) error "[EPANG] Cannot supply both binary and reference MSA or reference tree. Check input" + """ + epa-ng \\ + $args \\ + --threads $task.cpus \\ + $queryarg \\ + $refalnarg \\ + $reftreearg \\ + $bfastarg \\ + $binaryarg + + if [ -e epa_result.jplace ]; then + gzip epa_result.jplace + cp epa_result.jplace.gz ${prefix}.epa_result.jplace.gz + fi + [ -e epa_info.log ] && cp epa_info.log ${prefix}.epa_info.log + + cat <<-END_VERSIONS > versions.yml + "${task.process}": + epang: \$(echo \$(epa-ng --version 2>&1) | sed 's/^EPA-ng v//') + END_VERSIONS + """ +} diff --git a/modules/nf-core/epang/place/meta.yml b/modules/nf-core/epang/place/meta.yml new file mode 100644 index 00000000..483f4bad --- /dev/null +++ b/modules/nf-core/epang/place/meta.yml @@ -0,0 +1,66 @@ +name: "epang_place" +description: phylogenetic placement of query sequences in a reference tree +keywords: + - phylogeny + - phylogenetic placement + - sequences +tools: + - "epang": + description: "Massively parallel phylogenetic placement of genetic sequences" + homepage: "https://github.com/Pbdas/epa-ng" + documentation: "https://github.com/Pbdas/epa-ng/wiki/Full-Stack-Example" + tool_dev_url: "https://github.com/Pbdas/epa-ng" + doi: "10.1093/sysbio/syy054" + licence: "['GNU Affero General Public License v3.0']" + +input: + - meta: + type: map + description: | + Groovy Map containing sample information + e.g. [ id:'test' ] + - queryaln: + type: file + description: aligned query sequences in any supported format including phylip and fasta, may be gzipped + pattern: "*" + - referencealn: + type: file + description: reference alignment in any supported format including phylip and fasta, may be gzipped + pattern: "*" + - referencetree: + type: file + description: newick file containing the reference tree in which query sequences will be placed + pattern: "*" + - bfastfile: + type: file + description: file argument to the --bfast parameter + pattern: "*" + - binaryfile: + type: file + description: file argument to the --binary parameter + pattern: "*" + +output: + - meta: + type: map + description: | + Groovy Map containing sample information + e.g. [ id:'test' ] + - epang: + type: directory + description: directory in which EPA-NG was run + - jplace: + type: file + description: gzipped file with placement information + pattern: "*.jplace.gz" + - log: + type: file + description: log file from placement + pattern: "*.log" + - versions: + type: file + description: File containing software versions + pattern: "versions.yml" + +authors: + - "@erikrikarddaniel" diff --git a/modules/nf-core/epang/split/main.nf b/modules/nf-core/epang/split/main.nf new file mode 100644 index 00000000..973dd6dd --- /dev/null +++ b/modules/nf-core/epang/split/main.nf @@ -0,0 +1,37 @@ +process EPANG_SPLIT { + tag "$meta.id" + label 'process_single' + + conda "bioconda::epa-ng=0.3.8" + container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? + 'https://depot.galaxyproject.org/singularity/epa-ng:0.3.8--h9a82719_1': + 'quay.io/biocontainers/epa-ng:0.3.8--h9a82719_1' }" + + input: + tuple val(meta), path(refaln), path(fullaln) + + output: + tuple val(meta), path("*query.fasta.gz") , emit: query + tuple val(meta), path("*reference.fasta.gz"), emit: reference + path "versions.yml" , emit: versions + + when: + task.ext.when == null || task.ext.when + + script: + def args = task.ext.args ?: '' + def prefix = task.ext.prefix ?: "${meta.id}" + """ + epa-ng \\ + $args \\ + --split $refaln $fullaln + + gzip -c query.fasta > ${prefix}.query.fasta.gz; rm query.fasta + gzip -c reference.fasta > ${prefix}.reference.fasta.gz; rm reference.fasta + + cat <<-END_VERSIONS > versions.yml + "${task.process}": + epang: \$(echo \$(epa-ng --version 2>&1) | sed 's/^EPA-ng v//') + END_VERSIONS + """ +} diff --git a/modules/nf-core/epang/split/meta.yml b/modules/nf-core/epang/split/meta.yml new file mode 100644 index 00000000..b5d915a4 --- /dev/null +++ b/modules/nf-core/epang/split/meta.yml @@ -0,0 +1,50 @@ +name: "epang_split" +description: splits an alignment into reference and query parts +keywords: + - phylogeny + - phylogenetic placement + - sequences +tools: + - "epang": + description: "Massively parallel phylogenetic placement of genetic sequences" + homepage: "https://github.com/Pbdas/epa-ng" + documentation: "https://github.com/Pbdas/epa-ng/wiki/Full-Stack-Example" + tool_dev_url: "https://github.com/Pbdas/epa-ng" + doi: "10.1093/sysbio/syy054" + licence: "['GNU Affero General Public License v3.0']" + +input: + # Only when we have meta + - meta: + type: map + description: | + Groovy Map containing sample information + e.g. [ id:'test', single_end:false ] + - refaln: + type: file + description: reference alignment in any supported format including phylip and fasta, may be gzipped + pattern: "*.{faa,fna,fa,fasta,fa,phy,aln,alnfaa,alnfna,alnfa,mfa,faa.gz,fna.gz,fa.gz,fasta.gz,fa.gz,phy.gz,aln.gz,alnfaa.gz,alnfna.gz,alnfa.gz,mfa.gz}" + - fullaln: + type: file + description: full alignment in any supported format to split into reference and query alignments + pattern: "*.{faa,fna,fa,fasta,fa,phy,aln,alnfaa,alnfna,alnfa,mfa,faa.gz,fna.gz,fa.gz,fasta.gz,fa.gz,phy.gz,aln.gz,alnfaa.gz,alnfna.gz,alnfa.gz,mfa.gz}" + +output: + - meta: + type: map + description: | + Groovy Map containing sample information + e.g. [ id:'test', single_end:false ] + - query: + type: file + description: query sequence alignment in gzipped fasta format + - reference: + type: file + description: reference sequence alignment in gzipped fasta format + - versions: + type: file + description: File containing software versions + pattern: "versions.yml" + +authors: + - "@erikrikarddaniel" diff --git a/modules/nf-core/gappa/examineassign/main.nf b/modules/nf-core/gappa/examineassign/main.nf new file mode 100644 index 00000000..1c48d953 --- /dev/null +++ b/modules/nf-core/gappa/examineassign/main.nf @@ -0,0 +1,42 @@ +process GAPPA_EXAMINEASSIGN { + tag "$meta.id" + label 'process_medium' + + conda "bioconda::gappa=0.8.0" + container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? + 'https://depot.galaxyproject.org/singularity/gappa:0.8.0--h9a82719_0': + 'quay.io/biocontainers/gappa:0.8.0--h9a82719_0' }" + + input: + tuple val(meta), path(jplace), path(taxonomy) + + output: + tuple val(meta), path("./.") , emit: examineassign + tuple val(meta), path("*profile.tsv") , emit: profile + tuple val(meta), path("*labelled_tree.newick"), emit: labelled_tree + tuple val(meta), path("*per_query.tsv") , emit: per_query, optional: true + tuple val(meta), path("*krona.profile") , emit: krona , optional: true + tuple val(meta), path("*sativa.tsv") , emit: sativa , optional: true + path "versions.yml" , emit: versions + + when: + task.ext.when == null || task.ext.when + + script: + def args = task.ext.args ?: '' + def prefix = task.ext.prefix ?: "${meta.id}" + """ + gappa \\ + examine assign \\ + $args \\ + --threads $task.cpus \\ + --jplace-path $jplace \\ + --taxon-file $taxonomy \\ + --file-prefix ${prefix}. + + cat <<-END_VERSIONS > versions.yml + "${task.process}": + gappa: \$(echo \$(gappa --version 2>&1 | sed 's/v//' )) + END_VERSIONS + """ +} diff --git a/modules/nf-core/gappa/examineassign/meta.yml b/modules/nf-core/gappa/examineassign/meta.yml new file mode 100644 index 00000000..501c272b --- /dev/null +++ b/modules/nf-core/gappa/examineassign/meta.yml @@ -0,0 +1,55 @@ +name: "gappa_examineassign" +description: assigns taxonomy to query sequences in phylogenetic placement output +keywords: + - phylogeny + - phylogenetic placement + - classification + - taxonomy +tools: + - "gappa": + description: "Genesis Applications for Phylogenetic Placement Analysis" + homepage: "https://github.com/lczech/gappa" + documentation: "https://github.com/lczech/gappa/wiki" + tool_dev_url: "https://github.com/lczech/gappa" + doi: "10.1093/bioinformatics/btaa070" + licence: "['GPL v3']" + +input: + - meta: + type: map + description: | + Groovy Map containing sample information + e.g. [ id:'test' ] + - jplace: + type: file + description: jplace file output from phylogenetic placement, e.g. EPA-NG, gzipped or not + pattern: "*.{jplace,jplace.gz}" + +output: + - meta: + type: map + description: | + Groovy Map containing sample information + e.g. [ id:'test' ] + - examineassign: + type: directory + description: Execution directory + - profile: + type: file + description: profile tsv file + pattern: "*profile.tsv" + - labelled_tree: + type: file + description: labelled tree in newick format + pattern: "*labelled_tree.newick" + - per_query: + type: file + description: per query taxonomy assignments in tsv format + pattern: "*per_query.tsv" + - versions: + type: file + description: File containing software versions + pattern: "versions.yml" + +authors: + - "@erikrikarddaniel" diff --git a/modules/nf-core/gappa/examinegraft/main.nf b/modules/nf-core/gappa/examinegraft/main.nf new file mode 100644 index 00000000..b1121f3a --- /dev/null +++ b/modules/nf-core/gappa/examinegraft/main.nf @@ -0,0 +1,37 @@ +process GAPPA_EXAMINEGRAFT { + tag "$meta.id" + label 'process_low' + + conda "bioconda::gappa=0.8.0" + container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? + 'https://depot.galaxyproject.org/singularity/gappa:0.8.0--h9a82719_0': + 'quay.io/biocontainers/gappa:0.8.0--h9a82719_0' }" + + input: + tuple val(meta), path(jplace) + + output: + tuple val(meta), path("*.newick"), emit: newick + path "versions.yml" , emit: versions + + when: + task.ext.when == null || task.ext.when + + script: + def args = task.ext.args ?: '' + def prefix = task.ext.prefix ?: "${meta.id}" + """ + gappa \\ + examine \\ + graft \\ + $args \\ + --threads $task.cpus \\ + --file-prefix ${prefix}. \\ + --jplace-path $jplace + + cat <<-END_VERSIONS > versions.yml + "${task.process}": + gappa: \$(echo \$(gappa --version 2>&1 | sed 's/v//' )) + END_VERSIONS + """ +} diff --git a/modules/nf-core/gappa/examinegraft/meta.yml b/modules/nf-core/gappa/examinegraft/meta.yml new file mode 100644 index 00000000..0e813c85 --- /dev/null +++ b/modules/nf-core/gappa/examinegraft/meta.yml @@ -0,0 +1,41 @@ +name: "gappa_examinegraft" +description: grafts query sequences from phylogenetic placement on the reference tree +keywords: + - sort +tools: + - "gappa": + description: "Genesis Applications for Phylogenetic Placement Analysis" + homepage: "https://github.com/lczech/gappa" + documentation: "https://github.com/lczech/gappa/wiki" + tool_dev_url: "https://github.com/lczech/gappa" + doi: "10.1093/bioinformatics/btaa070" + licence: "['GPL v3']" + +input: + - meta: + type: map + description: | + Groovy Map containing sample information + e.g. [ id:'test' ] + - jplace: + type: file + description: jplace file output from phylogenetic placement, e.g. EPA-NG, gzipped or not + pattern: "*.{jplace,jplace.gz}" + +output: + - meta: + type: map + description: | + Groovy Map containing sample information + e.g. [ id:'test' ] + - versions: + type: file + description: File containing software versions + pattern: "versions.yml" + - newick: + type: file + description: phylogenetic tree file in newick format + pattern: "*.newick" + +authors: + - "@erikrikarddaniel" diff --git a/modules/nf-core/gappa/examineheattree/main.nf b/modules/nf-core/gappa/examineheattree/main.nf new file mode 100644 index 00000000..6898992f --- /dev/null +++ b/modules/nf-core/gappa/examineheattree/main.nf @@ -0,0 +1,45 @@ +process GAPPA_EXAMINEHEATTREE { + tag "$meta.id" + label 'process_low' + + conda "bioconda::gappa=0.8.0" + container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? + 'https://depot.galaxyproject.org/singularity/gappa:0.8.0--h9a82719_0': + 'quay.io/biocontainers/gappa:0.8.0--h9a82719_0' }" + + input: + tuple val(meta), path(jplace) + + output: + tuple val(meta), path("*.newick") , emit: newick , optional: true + tuple val(meta), path("*.nexus") , emit: nexus , optional: true + tuple val(meta), path("*.phyloxml") , emit: phyloxml, optional: true + tuple val(meta), path("*.svg") , emit: svg , optional: true + tuple val(meta), path("*.colours.txt"), emit: colours + tuple val(meta), path("*.log") , emit: log + path "versions.yml" , emit: versions + + when: + task.ext.when == null || task.ext.when + + script: + def args = task.ext.args ?: '' + def prefix = task.ext.prefix ?: "${meta.id}" + """ + gappa \\ + examine \\ + heat-tree \\ + --threads $task.cpus \\ + --file-prefix ${prefix}. \\ + --jplace-path $jplace \\ + $args \\ + --log-file ${prefix}.log + + grep '^ *At' ${prefix}.log > ${prefix}.colours.txt + + cat <<-END_VERSIONS > versions.yml + "${task.process}": + gappa: \$(echo \$(gappa --version 2>&1 | sed 's/v//' )) + END_VERSIONS + """ +} diff --git a/modules/nf-core/gappa/examineheattree/meta.yml b/modules/nf-core/gappa/examineheattree/meta.yml new file mode 100644 index 00000000..badf9719 --- /dev/null +++ b/modules/nf-core/gappa/examineheattree/meta.yml @@ -0,0 +1,64 @@ +name: "gappa_examineheattree" +description: colours a phylogeny with placement densities +keywords: + - phylogeny + - phylogenetic placement + - heattree + - visualisation +tools: + - "gappa": + description: "Genesis Applications for Phylogenetic Placement Analysis" + homepage: "https://github.com/lczech/gappa" + documentation: "https://github.com/lczech/gappa/wiki" + tool_dev_url: "https://github.com/lczech/gappa" + doi: "10.1093/bioinformatics/btaa070" + licence: "['GPL v3']" + +input: + - meta: + type: map + description: | + Groovy Map containing sample information + e.g. [ id:'test' ] + - jplace: + type: file + description: jplace file output from phylogenetic placement, e.g. EPA-NG, gzipped or not + pattern: "*.{jplace,jplace.gz}" + +output: + - meta: + type: map + description: | + Groovy Map containing sample information + e.g. [ id:'test' ] + - versions: + type: file + description: File containing software versions + pattern: "versions.yml" + - newick: + type: file + description: phylogenetic tree file in newick format + pattern: "*.newick" + - nexus: + type: file + description: coloured phylogenetic tree file in nexus format + pattern: "*.nexus" + - phyloxml: + type: file + description: coloured phylogenetic tree file in phyloxml format + pattern: "*.phyloxml" + - svg: + type: file + description: coloured phylogenetic tree file in svg format + pattern: "*.svg" + - colours: + type: file + description: colours used in plot + pattern: "*.colours.txt" + - log: + type: file + description: log file from the run + pattern: "*.log" + +authors: + - "@erikrikarddaniel" diff --git a/modules/nf-core/hmmer/eslalimask/main.nf b/modules/nf-core/hmmer/eslalimask/main.nf new file mode 100644 index 00000000..37776430 --- /dev/null +++ b/modules/nf-core/hmmer/eslalimask/main.nf @@ -0,0 +1,56 @@ +process HMMER_ESLALIMASK { + tag "$meta.id" + label 'process_single' + + conda "bioconda::hmmer=3.3.2" + container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? + 'https://depot.galaxyproject.org/singularity/hmmer:3.3.2--h1b792b2_1': + 'quay.io/biocontainers/hmmer:3.3.2--h1b792b2_1' }" + + input: + tuple val(meta), path(unmaskedaln), val(fmask_rf), val(fmask_all), val(gmask_rf), val(gmask_all), val(pmask_rf), val(pmask_all) + path maskfile + + output: + tuple val(meta), path("*.masked.sthlm.gz"), emit: maskedaln + path "*.fmask-rf.gz" , emit: fmask_rf , optional: true + path "*.fmask-all.gz" , emit: fmask_all, optional: true + path "*.gmask-rf.gz" , emit: gmask_rf , optional: true + path "*.gmask-all.gz" , emit: gmask_all, optional: true + path "*.pmask-rf.gz" , emit: pmask_rf , optional: true + path "*.pmask-all.gz" , emit: pmask_all, optional: true + path "versions.yml" , emit: versions + + when: + task.ext.when == null || task.ext.when + + script: + def args = task.ext.args ?: '' + def prefix = task.ext.prefix ?: "${meta.id}" + def fmask_rfarg = fmask_rf ? "--fmask-rf ${prefix}.fmask-rf" : "" + def fmask_allarg = fmask_all ? "--fmask-all ${prefix}.fmask-all" : "" + def gmask_rfarg = gmask_rf ? "--gmask-rf ${prefix}.gmask-rf" : "" + def gmask_allarg = gmask_all ? "--gmask-all ${prefix}.gmask-all" : "" + def pmask_rfarg = pmask_rf ? "--pmask-rf ${prefix}.pmask-rf" : "" + def pmask_allarg = pmask_all ? "--pmask-all ${prefix}.pmask-all" : "" + """ + esl-alimask \\ + $args \\ + $fmask_rfarg \\ + $fmask_allarg \\ + $gmask_rfarg \\ + $gmask_allarg \\ + $pmask_rfarg \\ + $pmask_allarg \\ + -o ${prefix}.masked.sthlm \\ + $unmaskedaln \\ + $maskfile + + gzip ${prefix}.*mask* + + cat <<-END_VERSIONS > versions.yml + "${task.process}": + hmmer/easel: \$(esl-reformat -h | grep -o '^# Easel [0-9.]*' | sed 's/^# Easel *//') + END_VERSIONS + """ +} diff --git a/modules/nf-core/hmmer/eslalimask/meta.yml b/modules/nf-core/hmmer/eslalimask/meta.yml new file mode 100644 index 00000000..238c7393 --- /dev/null +++ b/modules/nf-core/hmmer/eslalimask/meta.yml @@ -0,0 +1,105 @@ +name: "hmmer_eslalimask" +description: Mask multiple sequence alignments +keywords: + - hmmer + - alignment + - mask +tools: + - "hmmer": + description: "Biosequence analysis using profile hidden Markov models" + homepage: http://hmmer.org/ + documentation: http://hmmer.org/documentation.html + + doi: "10.1371/journal.pcbi.1002195" + licence: ["BSD-3-Clause"] + +input: + - meta: + type: map + description: | + Groovy Map containing sample information + e.g. [ id:'test' ] + + - unmaskedaln: + type: file + description: multiple sequence alignment, Stockholm or other formats + pattern: "*" + + - fmask_rf: + type: val + description: Flag to output optional file with final mask of non-gap RF len + + - fmask_all: + type: val + description: Flag to output optional file with final mask of full aln len + + - gmask_rf: + type: val + description: Flag to output optional file gap-based 0/1 mask of non-gap RF len + + - gmask_all: + type: val + description: Flag to output optional file gap-based 0/1 mask of full aln len + + - pmask_rf: + type: val + description: Flag to output optional file with PP-based 0/1 mask of non-gap RF len + + - pmask_all: + type: val + description: Flag to output optional file with PP-based 0/1 mask of full aln len + + - maskfile: + type: file + description: mask file, see program documentation + pattern: "*" + +output: + - meta: + type: map + description: | + Groovy Map containing sample information + e.g. [ id:'test', single_end:false ] + + - versions: + type: file + description: File containing software versions + pattern: "versions.yml" + + - maskedaln: + type: file + description: Masked alignment in gzipped Stockholm format + pattern: "*.sthlm.gz" + + - fmask_rf: + type: file + description: File with final mask of non-gap RF len + pattern: "*.fmask-rf.gz" + + - fmask_all: + type: file + description: File with final mask of full aln len + pattern: "*.fmask-all.gz" + + - gmask_rf: + type: file + description: File with gap-based 0/1 mask of non-gap RF len + pattern: "*.gmask-rf.gz" + + - gmask_all: + type: file + description: File with gap-based 0/1 mask of full aln len + pattern: "*.gmask-all.gz" + + - pmask_rf: + type: file + description: File with PP-based 0/1 mask of non-gap RF len + pattern: "*.pmask-rf.gz" + + - pmask_all: + type: file + description: File with PP-based 0/1 mask of full aln len + pattern: "*.pmask-all.gz" + +authors: + - "@erikrikarddaniel" diff --git a/modules/nf-core/hmmer/eslreformat/main.nf b/modules/nf-core/hmmer/eslreformat/main.nf new file mode 100644 index 00000000..6eaae099 --- /dev/null +++ b/modules/nf-core/hmmer/eslreformat/main.nf @@ -0,0 +1,38 @@ +process HMMER_ESLREFORMAT { + tag "$meta.id" + label 'process_single' + + conda "bioconda::hmmer=3.3.2" + container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? + 'https://depot.galaxyproject.org/singularity/hmmer:3.3.2--h1b792b2_1': + 'quay.io/biocontainers/hmmer:3.3.2--h1b792b2_1' }" + + input: + tuple val(meta), path(seqfile) + + output: + tuple val(meta), path("*.*.gz"), emit: seqreformated + path "versions.yml" , emit: versions + + when: + task.ext.when == null || task.ext.when + + script: + def args = task.ext.args ?: '' + def prefix = task.ext.prefix ?: "${meta.id}" + def suffix = args ? args.trim().tokenize(" ")[-1] : "sequences" + // Use for any postprocessing of the sequence file, e.g. removal of gap characters + def postproc = task.ext.postprocessing ?: "" + """ + esl-reformat \\ + $args \\ + $seqfile \\ + $postproc \\ + | gzip -c > ${prefix}.${suffix}.gz + + cat <<-END_VERSIONS > versions.yml + "${task.process}": + hmmer/easel: \$(esl-reformat -h | grep -o '^# Easel [0-9.]*' | sed 's/^# Easel *//') + END_VERSIONS + """ +} diff --git a/modules/nf-core/hmmer/eslreformat/meta.yml b/modules/nf-core/hmmer/eslreformat/meta.yml new file mode 100644 index 00000000..05ff04a9 --- /dev/null +++ b/modules/nf-core/hmmer/eslreformat/meta.yml @@ -0,0 +1,41 @@ +name: "hmmer_eslreformat" +description: reformats sequence files, see HMMER documentation for details. The module requires that the format is specified in ext.args in a config file, and that this comes last. See the tools help for possible values. +keywords: + - sort +tools: + - "hmmer": + description: "Biosequence analysis using profile hidden Markov models" + homepage: http://hmmer.org/ + documentation: http://hmmer.org/documentation.html + + doi: "10.1371/journal.pcbi.1002195" + licence: ["BSD-3-Clause"] + +input: + - meta: + type: map + description: | + Groovy Map containing sample information + e.g. [ id:'test' ] + - seqfile: + type: file + description: Sequences, aligned or not, in any supported format + pattern: "*" + +output: + - meta: + type: map + description: | + Groovy Map containing sample information + e.g. [ id:'test', single_end:false ] + - versions: + type: file + description: File containing software versions + pattern: "versions.yml" + - seqreformated: + type: file + description: Reformated sequence file + pattern: "*.*.gz" + +authors: + - "@erikrikarddaniel" diff --git a/modules/nf-core/hmmer/hmmalign/main.nf b/modules/nf-core/hmmer/hmmalign/main.nf new file mode 100644 index 00000000..63f2bbd1 --- /dev/null +++ b/modules/nf-core/hmmer/hmmalign/main.nf @@ -0,0 +1,35 @@ +process HMMER_HMMALIGN { + tag "$meta.id" + label 'process_single' + + conda "bioconda::hmmer=3.3.2" + container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? + 'https://depot.galaxyproject.org/singularity/hmmer:3.3.2--h1b792b2_1' : + 'quay.io/biocontainers/hmmer:3.3.2--h1b792b2_1' }" + + input: + tuple val(meta), path(fasta) + path hmm + + output: + tuple val(meta), path("*.sthlm.gz"), emit: sthlm + path "versions.yml" , emit: versions + + when: + task.ext.when == null || task.ext.when + + script: + def args = task.ext.args ?: '' + def prefix = task.ext.prefix ?: "${meta.id}" + """ + hmmalign \\ + $args \\ + $hmm \\ + $fasta | gzip -c > ${prefix}.sthlm.gz + + cat <<-END_VERSIONS > versions.yml + "${task.process}": + hmmer: \$(hmmalign -h | grep -o '^# HMMER [0-9.]*' | sed 's/^# HMMER *//') + END_VERSIONS + """ +} diff --git a/modules/nf-core/hmmer/hmmalign/meta.yml b/modules/nf-core/hmmer/hmmalign/meta.yml new file mode 100644 index 00000000..df1bd37e --- /dev/null +++ b/modules/nf-core/hmmer/hmmalign/meta.yml @@ -0,0 +1,46 @@ +name: hmmer_hmmalign +description: hmmalign from the HMMER suite aligns a number of sequences to an HMM profile +keywords: + - alignment +tools: + - hmmer: + description: Biosequence analysis using profile hidden Markov models + homepage: http://hmmer.org/ + documentation: http://hmmer.org/documentation.html + + doi: "10.1371/journal.pcbi.1002195" + licence: ["BSD-3-Clause"] + +input: + - meta: + type: map + description: | + Groovy Map containing sample information + e.g. [ id:'test' ] + - fasta: + type: file + description: Amino acid or nucleotide gzipped compressed fasta file + pattern: "*.{fna.gz,faa.gz,fasta.gz,fa.gz}" + - hmm: + type: file + description: A gzipped HMM file + pattern: "*.hmm.gz" + +output: + - meta: + type: map + description: | + Groovy Map containing sample information + e.g. [ id:'test', single_end:false ] + - versions: + type: file + description: File containing software versions + pattern: "versions.yml" + - sthlm: + type: file + description: Multiple alignment in gzipped Stockholm format + pattern: "*.sthlm.gz" + +authors: + - "@erikrikarddaniel" + - "@jfy133" diff --git a/modules/nf-core/hmmer/hmmbuild/main.nf b/modules/nf-core/hmmer/hmmbuild/main.nf new file mode 100644 index 00000000..91924b99 --- /dev/null +++ b/modules/nf-core/hmmer/hmmbuild/main.nf @@ -0,0 +1,44 @@ +process HMMER_HMMBUILD { + tag "$meta.id" + label 'process_low' + + conda "bioconda::hmmer=3.3.2" + container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? + 'https://depot.galaxyproject.org/singularity/hmmer:3.3.2--h87f3376_2': + 'quay.io/biocontainers/hmmer:3.3.2--h1b792b2_1' }" + + input: + tuple val(meta), path(alignment) + path mxfile + + output: + tuple val(meta), path("*.hmm.gz"), emit: hmm + path "*.hmmbuild.txt", emit: hmmbuildout + path "versions.yml", emit: versions + + when: + task.ext.when == null || task.ext.when + + script: + def args = task.ext.args ?: '' + def prefix = task.ext.prefix ?: "${meta.id}" + def mxfileopt = mxfile ? "--mxfile ${mxfile}" : "" + + """ + hmmbuild \\ + $args \\ + --cpu $task.cpus \\ + -n ${prefix} \\ + -o ${prefix}.hmmbuild.txt \\ + ${mxfileopt} \\ + ${prefix}.hmm \\ + $alignment + + gzip ${prefix}.hmm + + cat <<-END_VERSIONS > versions.yml + "${task.process}": + hmmer: \$(echo \$(hmmbuild -h | grep HMMER | sed 's/# HMMER //' | sed 's/ .*//' 2>&1)) + END_VERSIONS + """ +} diff --git a/modules/nf-core/hmmer/hmmbuild/meta.yml b/modules/nf-core/hmmer/hmmbuild/meta.yml new file mode 100644 index 00000000..77af69cf --- /dev/null +++ b/modules/nf-core/hmmer/hmmbuild/meta.yml @@ -0,0 +1,44 @@ +name: "hmmer_hmmbuild" +description: create an hmm profile from a multiple sequence alignment +keywords: + - search + - hidden Markov model + - HMM + - hmmer + - hmmsearch +tools: + - "hmmer": + description: "Biosequence analysis using profile hidden Markov models" + homepage: "http://hmmer.org" + documentation: "http://hmmer.org/documentation.html" + tool_dev_url: "https://github.com/EddyRivasLab/hmmer" + doi: "10.1371/journal.pcbi.1002195" + licence: "['BSD']" + +input: + - meta: + type: map + description: | + Groovy Map containing sample information + e.g. [ id:'test', single_end:false ] + - alignment: + type: file + description: multiple sequence alignment in fasta, clustal, stockholm or phylip format + pattern: "*" + - mxfile: + type: file + description: read substitution score matrix, for use when building profiles from single sequences (--singlemx option) + pattern: "*" + +output: + - versions: + type: file + description: File containing software versions + pattern: "versions.yml" + - hmm: + type: file + description: Gzipped HMM file + pattern: "*.{hmm.gz}" + +authors: + - "@erikrikarddaniel" diff --git a/modules/nf-core/mafft/main.nf b/modules/nf-core/mafft/main.nf new file mode 100644 index 00000000..d2b02b78 --- /dev/null +++ b/modules/nf-core/mafft/main.nf @@ -0,0 +1,38 @@ +process MAFFT { + tag "$meta.id" + label 'process_high' + + conda "bioconda::mafft=7.508" + container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? + 'https://depot.galaxyproject.org/singularity/mafft:7.508--hec16e2b_0': + 'quay.io/biocontainers/mafft:7.508--hec16e2b_0' }" + + input: + tuple val(meta), path(fasta) + path addsequences + + output: + tuple val(meta), path("*.fas"), emit: fas + path "versions.yml" , emit: versions + + when: + task.ext.when == null || task.ext.when + + script: + def args = task.ext.args ?: '' + def prefix = task.ext.prefix ?: "${meta.id}" + def add = addsequences ? "--add $addsequences" : '' + """ + mafft \\ + --thread ${task.cpus} \\ + ${args} \\ + ${add} \\ + ${fasta} \\ + > ${prefix}.fas + + cat <<-END_VERSIONS > versions.yml + "${task.process}": + mafft: \$(mafft --version 2>&1 | sed 's/^v//' | sed 's/ (.*)//') + END_VERSIONS + """ +} diff --git a/modules/nf-core/mafft/meta.yml b/modules/nf-core/mafft/meta.yml new file mode 100644 index 00000000..a366a4b8 --- /dev/null +++ b/modules/nf-core/mafft/meta.yml @@ -0,0 +1,46 @@ +name: mafft +description: Multiple sequence alignment using MAFFT +keywords: + - msa + - multiple sequence alignment +tools: + - mafft: + description: Multiple alignment program for amino acid or nucleotide sequences based on fast Fourier transform + homepage: https://mafft.cbrc.jp/alignment/software/ + documentation: https://mafft.cbrc.jp/alignment/software/manual/manual.html + tool_dev_url: https://mafft.cbrc.jp/alignment/software/source.html + doi: "10.1093/nar/gkf436" + licence: ["BSD"] + +input: + - meta: + type: map + description: | + Groovy Map containing sample information + e.g. [ id:'test', single_end:false ] + - fasta: + type: file + description: FASTA file containing the sequences to align + pattern: "*.{fa,fasta}" + - addsequences: + type: file + description: FASTA file containing sequences to align to the sequences in `fasta` + pattern: "*.{fa,fasta}" + +output: + - meta: + type: map + description: | + Groovy Map containing sample information + e.g. [ id:'test', single_end:false ] + - versions: + type: file + description: File containing software versions + pattern: "versions.yml" + - fas: + type: file + description: Aligned sequences in FASTA format + pattern: "*.{fas}" + +authors: + - "@MillironX" diff --git a/nextflow.config b/nextflow.config index 28efd87b..ff2e4733 100644 --- a/nextflow.config +++ b/nextflow.config @@ -58,6 +58,12 @@ params { filter_ssu = null min_len_asv = null max_len_asv = null + pplace_tree = null + pplace_aln = null + pplace_model = null + pplace_alnmethod = 'hmmer' + pplace_taxonomy = null + pplace_name = null diversity_rarefaction_depth = 500 ancom_sample_min_count = 1 @@ -218,6 +224,7 @@ profiles { test_full { includeConfig 'conf/test_full.config' } test_reftaxcustom { includeConfig 'conf/test_reftaxcustom.config' } test_novaseq { includeConfig 'conf/test_novaseq.config' } + test_pplace { includeConfig 'conf/test_pplace.config' } } diff --git a/nextflow_schema.json b/nextflow_schema.json index 4a343309..937cfb51 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -260,6 +260,37 @@ "help_text": "Expected amplified sequences are extracted from the DADA2 reference taxonomy using the primer sequences, that might improve classification. This is not applied to species classification (assignSpecies) but only for lower taxonomic levels (assignTaxonomy).", "description": "If the expected amplified sequences are extracted from the DADA2 reference taxonomy database" }, + "pplace_tree": { + "type": "string", + "description": "Newick file with reference phylogenetic tree. Requires also `--pplace_aln` and `--pplace_model`." + }, + "pplace_aln": { + "type": "string", + "description": "File with reference sequences. Requires also `--pplace_tree` and `--pplace_model`." + }, + "pplace_model": { + "type": "string", + "description": "Phylogenetic model to use in placement, e.g. 'LG+F' or 'GTR+I+F'. Requires also `--pplace_tree` and `--pplace_aln`." + }, + "pplace_alnmethod": { + "type": "string", + "description": "Method used for alignment, \"hmmer\" or \"mafft\"", + "default": "hmmer", + "enum": [ + "hmmer", + "mafft" + ] + }, + "pplace_taxonomy": { + "type": "string", + "help_text": "Headerless, tab-separated, first column with tree leaves, second column with taxonomy ranks separated by semicolon `;`.", + "description": "Tab-separated file with taxonomy assignments of reference sequences." + }, + "pplace_name": { + "type": "string", + "description": "A name for the run", + "hidden": true + }, "qiime_ref_taxonomy": { "type": "string", "help_text": "Choose any of the supported databases, and optionally also specify the version. Database and version are separated by an equal sign (`=`, e.g. `silva=138`) . This will download the desired database and initiate taxonomic classification with QIIME2 and the chosen database.\n\nIf both, `--dada_ref_taxonomy` and `--qiime_ref_taxonomy` are used, DADA2 classification will be used for downstream analysis.\n\nThe following databases are supported:\n- SILVA ribosomal RNA gene database project - 16S rRNA\n- UNITE - eukaryotic nuclear ribosomal ITS region - ITS\n- Greengenes (only testing!)\n\nGenerally, using `silva`, `unite-fungi`, or `unite-alleuk` will select the most recent supported version. For testing purposes, the tiny database `greengenes85` (dereplicated at 85% sequence similarity) is available. For details on what values are valid, please either use an invalid value such as `x` (causing the pipeline to send an error message with all valid values) or see `conf/ref_databases.config`.", diff --git a/subworkflows/nf-core/fasta_newick_epang_gappa/main.nf b/subworkflows/nf-core/fasta_newick_epang_gappa/main.nf new file mode 100644 index 00000000..15cdb20c --- /dev/null +++ b/subworkflows/nf-core/fasta_newick_epang_gappa/main.nf @@ -0,0 +1,154 @@ +include { HMMER_HMMBUILD } from '../../../modules/nf-core/hmmer/hmmbuild/main' +include { HMMER_HMMALIGN as HMMER_HMMALIGNREF } from '../../../modules/nf-core/hmmer/hmmalign/main' +include { HMMER_HMMALIGN as HMMER_HMMALIGNQUERY } from '../../../modules/nf-core/hmmer/hmmalign/main' +include { HMMER_ESLALIMASK as HMMER_MASKREF } from '../../../modules/nf-core/hmmer/eslalimask/main' +include { HMMER_ESLALIMASK as HMMER_MASKQUERY } from '../../../modules/nf-core/hmmer/eslalimask/main' +include { HMMER_ESLREFORMAT as HMMER_UNALIGNREF } from '../../../modules/nf-core/hmmer/eslreformat/main' +include { HMMER_ESLREFORMAT as HMMER_AFAFORMATREF } from '../../../modules/nf-core/hmmer/eslreformat/main' +include { HMMER_ESLREFORMAT as HMMER_AFAFORMATQUERY } from '../../../modules/nf-core/hmmer/eslreformat/main' +include { MAFFT } from '../../../modules/nf-core/mafft/main' +include { EPANG_PLACE } from '../../../modules/nf-core/epang/place/main' +include { EPANG_SPLIT } from '../../../modules/nf-core/epang/split/main' +include { GAPPA_EXAMINEGRAFT as GAPPA_GRAFT } from '../../../modules/nf-core/gappa/examinegraft/main' +include { GAPPA_EXAMINEASSIGN as GAPPA_ASSIGN } from '../../../modules/nf-core/gappa/examineassign/main' +include { GAPPA_EXAMINEHEATTREE as GAPPA_HEATTREE } from '../../../modules/nf-core/gappa/examineheattree/main' + +workflow FASTA_NEWICK_EPANG_GAPPA { + + take: + ch_pp_data // channel: [ meta: val(meta), data: [ alignmethod: val(alignmethod), queryseqfile: file(queryseqfile), refseqfile: file(refseqfile), refphylogeny: file(refphylogeny), hmmfile: file(hmmfile), model: val(model) ] ] + + main: + ch_versions = Channel.empty() + + // Divide the input channel into two: One for hmmer and one for mafft alignment + ch_hmmer_data = ch_pp_data.filter { it.data.alignmethod == 'hmmer' } + ch_mafft_data = ch_pp_data.filter { it.data.alignmethod == 'mafft' } + + // 1.a.1 HMMER alignment: For entries that do not specify an hmm file, build one to use for alignment + HMMER_HMMBUILD ( + ch_hmmer_data + .filter { ! it.data.hmmfile } + .map { [ it.meta, it.data.refseqfile ] }, + [] + ) + // 1.a.2 This handles mixed input where some samples have hmmfile set, while others don't (sample sheet input) + ch_hmm = Channel.empty() + .mix(HMMER_HMMBUILD.out.hmm.map { [ it[0], it[1] ] }) + .mix( + ch_hmmer_data + .filter { it.data.hmmfile } + .map { [ it.meta, it.data.hmmfile ] } + ) + + ch_versions = ch_versions.mix(HMMER_HMMBUILD.out.versions.first()) + + // 1.b For entries that do not specify an hmm file, "unalign" the reference sequences before they can be aligned to the hmm. + HMMER_UNALIGNREF ( + ch_hmmer_data + .filter { ! it.data.hmmfile } + .map { [ it.meta, it.data.refseqfile ] } + ) + ch_hmmer_unaligned = Channel.empty() + .mix(HMMER_UNALIGNREF.out.seqreformated.map { [ it[0], it[1] ] }) + .mix( + ch_hmmer_data + .filter { it.data.hmmfile } + .map { [ it.meta, it.data.refseqfile ] } + ) + + ch_versions = ch_versions.mix(HMMER_UNALIGNREF.out.versions) + + // 1.c Align the reference and query sequences to the profile + ch_hmmer_alignref = ch_hmm + .mix(ch_hmmer_unaligned) + .groupTuple(size: 2, sort: { a, b -> a =~ /\.hmm/ ? 1 : -1 }) + + HMMER_HMMALIGNREF ( + ch_hmmer_alignref.map { [ it[0], it[1][0] ] }, + ch_hmmer_alignref.map { it[1][1] } + ) + ch_versions = ch_versions.mix(HMMER_HMMALIGNREF.out.versions) + + ch_hmmer_alignquery = Channel.empty() + .mix(ch_hmmer_data.map { [ it.meta, it.data.queryseqfile ] }) + .mix(ch_hmm) + .groupTuple(size: 2, sort: { a, b -> a =~ /\.hmm/ ? 1 : -1 }) + + HMMER_HMMALIGNQUERY ( + ch_hmmer_alignquery.map { [ it[0], it[1][0] ] }, + ch_hmmer_alignquery.map { it[1][1] } + ) + ch_versions = ch_versions.mix(HMMER_HMMALIGNQUERY.out.versions) + + // 1.d Mask the alignments (Add '--rf-is-mask' ext.args in config for the process.) + HMMER_MASKREF ( HMMER_HMMALIGNREF.out.sthlm.map { [ it[0], it[1], [], [], [], [], [], [] ] }, [] ) + ch_versions = ch_versions.mix(HMMER_MASKREF.out.versions) + + HMMER_MASKQUERY ( HMMER_HMMALIGNQUERY.out.sthlm.map { [ it[0], it[1], [], [], [], [], [], [] ] }, [] ) + ch_versions = ch_versions.mix(HMMER_MASKQUERY.out.versions) + + // 1.e Reformat alignments to "afa" (aligned fasta) + HMMER_AFAFORMATREF ( HMMER_MASKREF.out.maskedaln ) + ch_versions = ch_versions.mix(HMMER_AFAFORMATREF.out.versions) + + HMMER_AFAFORMATQUERY ( HMMER_MASKQUERY.out.maskedaln ) + ch_versions = ch_versions.mix(HMMER_AFAFORMATQUERY.out.versions) + + // 2.a MAFFT profile alignment of query sequences to reference alignment + MAFFT ( + ch_mafft_data.map { [ it.meta, it.data.refseqfile ] }, + ch_mafft_data.map { [ it.data.queryseqfile ] } + ) + ch_versions = ch_versions.mix(MAFFT.out.versions) + + // 2.b Split the profile alignment into reference and query parts + EPANG_SPLIT ( + ch_mafft_data.map { [ it.meta, it.data.refseqfile ] } + .join(MAFFT.out.fas) + ) + ch_versions = ch_versions.mix(EPANG_SPLIT.out.versions) + + // 3. Do the placement + ch_epang_query = ch_pp_data.map { [ it.meta, it.data.model, it.data.refphylogeny ] } + .join ( HMMER_AFAFORMATQUERY.out.seqreformated ) + .join ( HMMER_AFAFORMATREF.out.seqreformated ) + .mix( + ch_pp_data.map { [ it.meta, it.data.model, it.data.refphylogeny ] } + .join(EPANG_SPLIT.out.query.map { [ it[0], it[1] ] } ) + .join(EPANG_SPLIT.out.reference.map { [ it[0], it[1] ] } ) + ) + .map { [ [ id:it[0].id, model:it[1] ], it[3], it[4], it[2] ] } + + EPANG_PLACE ( + ch_epang_query, + [], [] + ) + ch_versions = ch_versions.mix(EPANG_PLACE.out.versions) + + // 7. Calculate a tree with the placed sequences + GAPPA_GRAFT ( EPANG_PLACE.out.jplace ) + ch_versions = ch_versions.mix(GAPPA_GRAFT.out.versions) + + // 8. Classify + GAPPA_ASSIGN ( + EPANG_PLACE.out.jplace + .map { [ [ id:it[0].id ], it[1] ] } + .join( ch_pp_data.map { [ it.meta, it.data.taxonomy ] } ) + ) + ch_versions = ch_versions.mix(GAPPA_ASSIGN.out.versions) + + // 9. Heat tree output + GAPPA_HEATTREE ( EPANG_PLACE.out.jplace ) + ch_versions = ch_versions.mix(GAPPA_HEATTREE.out.versions) + + emit: + epang = EPANG_PLACE.out.epang + jplace = EPANG_PLACE.out.jplace + grafted_phylogeny = GAPPA_GRAFT.out.newick + taxonomy_profile = GAPPA_ASSIGN.out.profile + taxonomy_per_query = GAPPA_ASSIGN.out.per_query + heattree = GAPPA_HEATTREE.out.svg + versions = ch_versions // channel: [ versions.yml ] +} + diff --git a/subworkflows/nf-core/fasta_newick_epang_gappa/meta.yml b/subworkflows/nf-core/fasta_newick_epang_gappa/meta.yml new file mode 100644 index 00000000..a6dbe999 --- /dev/null +++ b/subworkflows/nf-core/fasta_newick_epang_gappa/meta.yml @@ -0,0 +1,97 @@ +name: "fasta_newick_epang_gappa" +description: Run phylogenetic placement with a number of query sequences plus a reference alignment and phylogeny. Used in nf-core/phyloplace. +keywords: + - phylogenetic placement + - phylogenetics + - alignment + - fasta + - newick +modules: + - hmmer/hmmbuild + - hmmer/hmmalign + - hmmer/eslalimask + - hmmer/eslreformat + - mafft + - epang/place + - epang/split + - gappa/examinegraft + - gappa/examineassign + - gappa/examineheattree +input: + - ch_pp_data: + type: map + description: | + Structure: [ + meta: val(meta), + data: [ + alignmethod: 'hmmer', + queryseqfile: path("*.faa"), + refseqfile: path("*.alnfaa"), + refphylogeny: path("*.newick"), + model: "LG", + taxonomy: path("*.tsv") + ] + ] + - meta: + type: map + description: | + Groovy Map containing sample information + e.g. [ id:'test' ] + - queryseqfile: + type: file + description: Fasta file with query sequences + pattern: "*.{faa,fna,fa,fasta,fa,faa.gz,fna.gz,fa.gz,fasta.gz,fa.gz}" + - refseqfile: + type: file + description: File with reference sequences; aligned unless an hmmfile is provided + pattern: "*.{faa,fna,fa,fasta,fa,phy,aln,alnfaa,alnfna,alnfa,mfa,faa.gz,fna.gz,fa.gz,fasta.gz,fa.gz,phy.gz,aln.gz,alnfaa.gz,alnfna.gz,alnfa.gz,mfa.gz}" + - refphylogeny: + type: file + description: Newick file with reference phylogenetic tree + pattern: "*.{newick,tree}" + - hmmfile: + type: file + description: HMM file to use for alignment; implies that refseqfile is not aligned. Optional. + pattern: "*.{hmm,HMM,hmm.gz,HMM.gz}" + - model: + type: string + description: Phylogenetic model to use in placement, e.g. 'LG+F' or 'GTR+I+F' + - alignmethod: + type: string + description: Method used for alignment, 'hmmer' or 'mafft' +output: + - meta: + type: map + description: | + Groovy Map containing sample information + e.g. [ id:'test' ] + - epang: + type: directory + description: All output from EPA-NG + pattern: "*" + - jplace: + type: file + description: jplace file from EPA-NG + pattern: "*.jplace" + - grafted_phylogeny: + type: file + description: Newick file with query sequences placed in reference tree + pattern: "*.newick" + - taxonomy_profile: + type: file + description: Tab separated file with taxonomy information from classification + pattern: "*.tsv" + - taxonomy_per_query: + type: file + description: Tab separated file with taxonomy information per query from classification + pattern: "*.tsv" + - heattree: + type: file + description: Heattree in SVG format + pattern: "*.svg" + - versions: + type: file + description: File containing software versions + pattern: "versions.yml" +authors: + - "@erikrikarddaniel" diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 6523d174..c17760a8 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -183,6 +183,7 @@ include { FASTQC } from '../modules/nf-core/fastqc/ma include { MULTIQC } from '../modules/nf-core/multiqc/main' include { CUSTOM_DUMPSOFTWAREVERSIONS } from '../modules/nf-core/custom/dumpsoftwareversions/main' include { VSEARCH_USEARCHGLOBAL } from '../modules/nf-core/vsearch/usearchglobal/main' +include { FASTA_NEWICK_EPANG_GAPPA } from '../subworkflows/nf-core/fasta_newick_epang_gappa/main' /* @@ -478,6 +479,30 @@ workflow AMPLISEQ { } } + //Phylo placement + if ( params.pplace_tree ) { + ch_pp_data = ch_fasta.map { it -> + [ meta: [ id: params.pplace_name ?: 'user_tree' ], + data: [ + alignmethod: params.pplace_alnmethod ?: 'hmmer', + queryseqfile: it, + refseqfile: file( params.pplace_aln, checkIfExists: true ), + hmmfile: [], + refphylogeny: file( params.pplace_tree, checkIfExists: true ), + model: params.pplace_model, + taxonomy: params.pplace_taxonomy ? file( params.pplace_taxonomy, checkIfExists: true ) : [] + ] ] + } + FASTA_NEWICK_EPANG_GAPPA ( ch_pp_data ) + ch_versions = ch_versions.mix( FASTA_NEWICK_EPANG_GAPPA.out.versions ) + //TODO: if params.pplace_taxonomy is given, use taxonomy over DADA2 taxonomy for downstream analysis --> FASTA_NEWICK_EPANG_GAPPA.out.taxonomy_per_query --> test_pplace.taxonomy.per_query.tsv + //Use for each ASV the annotation with lowest LWR (likelihood-weight-ration) -> conversion script + //if tie -> use annotation with less entries (i.e. conservative choice) + //if same number of entries -> remove last entry + //if those truncated annotations arent identical, remove one more entry; repeat until identical + //TODO: use newick tree in downstream analysis --> FASTA_NEWICK_EPANG_GAPPA.out.grafted_phylogeny --> test_pplace.graft.test_pplace.epa_result.newick + } + //QIIME2 if ( run_qiime2 ) { if (params.qiime_ref_taxonomy && !params.classifier) { From 9e525e84c6a8e68480fe7c539ed4a35112c0c994 Mon Sep 17 00:00:00 2001 From: Daniel Straub <42973691+d4straub@users.noreply.github.com> Date: Tue, 28 Mar 2023 15:20:34 +0200 Subject: [PATCH 024/158] Update docs/output.md Co-authored-by: Daniel Lundin --- docs/output.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/docs/output.md b/docs/output.md index da0b99c5..bcb0c6ee 100644 --- a/docs/output.md +++ b/docs/output.md @@ -234,7 +234,7 @@ Optionally, a UNITE species hypothesis (SH) can be added to the taxonomy. In sho ### Phlogenetic placement and taxonomic classification -Phylogenetic placement allows to graft sequences onto a phylogenetic reference tree and retrieve taxonomic annotations. The reference tree can be made of whole genomes (or other evolutionary information) that can yield more relevant information than a de-novo calculated phylogenetic tree from short amplicon sequences. On providing required reference files, ASV sequences are aligned to the reference alignment with either [HMMER](http://hmmer.org/) (default) or [MAFFT](https://mafft.cbrc.jp/alignment/software/), phylogenetic placement of query sequences is performed with [EPA-NG](https://github.com/Pbdas/epa-ng), and finally a number of summary operations are performed with [Gappa](https://github.com/lczech/gappa) after placement. This uses code (subworkflow) from [nf-core/phyloplace](https://nf-co.re/phyloplace) and its [subworkflow](https://github.com/nf-core/modules/tree/master/subworkflows/nf-core/fasta_newick_epang_gappa), therefore its detailed documentation also applies here. +Phylogenetic placement grafts sequences onto a phylogenetic reference tree and optionally outputs taxonomic annotations. The reference tree is ideally made from full-length high-quality sequences containing better evolutionary signal than short amplicons. It is hence superior to estimating de-novo phylogenetic trees from short amplicon sequences. On providing required reference files, ASV sequences are aligned to the reference alignment with either [HMMER](http://hmmer.org/) (default) or [MAFFT](https://mafft.cbrc.jp/alignment/software/). Subsequently, phylogenetic placement of query sequences is performed with [EPA-NG](https://github.com/Pbdas/epa-ng), and finally a number of summary operations are performed with [Gappa](https://github.com/lczech/gappa). This uses code from [nf-core/phyloplace](https://nf-co.re/phyloplace) in the form of its main [subworkflow](https://github.com/nf-core/modules/tree/master/subworkflows/nf-core/fasta_newick_epang_gappa), therefore its detailed documentation also applies here.
Output files From 2a5d302aef1191b99cfcafb0b20868ede18f292f Mon Sep 17 00:00:00 2001 From: Daniel Straub <42973691+d4straub@users.noreply.github.com> Date: Tue, 28 Mar 2023 15:20:42 +0200 Subject: [PATCH 025/158] Update nextflow.config Co-authored-by: Daniel Lundin --- nextflow.config | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/nextflow.config b/nextflow.config index ff2e4733..38c5571b 100644 --- a/nextflow.config +++ b/nextflow.config @@ -224,7 +224,7 @@ profiles { test_full { includeConfig 'conf/test_full.config' } test_reftaxcustom { includeConfig 'conf/test_reftaxcustom.config' } test_novaseq { includeConfig 'conf/test_novaseq.config' } - test_pplace { includeConfig 'conf/test_pplace.config' } + test_pplace { includeConfig 'conf/test_pplace.config' } } From 821c766a8f51e5d9c614b49dada5cce13b91073a Mon Sep 17 00:00:00 2001 From: Daniel Straub <42973691+d4straub@users.noreply.github.com> Date: Tue, 28 Mar 2023 15:20:50 +0200 Subject: [PATCH 026/158] Update workflows/ampliseq.nf Co-authored-by: Daniel Lundin --- workflows/ampliseq.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index c17760a8..c89d2cff 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -479,7 +479,7 @@ workflow AMPLISEQ { } } - //Phylo placement + // Phylo placement if ( params.pplace_tree ) { ch_pp_data = ch_fasta.map { it -> [ meta: [ id: params.pplace_name ?: 'user_tree' ], From 5aad68bd4a5010cbe35bd09a4b69d03419d7ddcf Mon Sep 17 00:00:00 2001 From: daniel Date: Tue, 28 Mar 2023 16:00:42 +0200 Subject: [PATCH 027/158] fix prettier --- nextflow_schema.json | 5 +---- 1 file changed, 1 insertion(+), 4 deletions(-) diff --git a/nextflow_schema.json b/nextflow_schema.json index 937cfb51..e334f391 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -276,10 +276,7 @@ "type": "string", "description": "Method used for alignment, \"hmmer\" or \"mafft\"", "default": "hmmer", - "enum": [ - "hmmer", - "mafft" - ] + "enum": ["hmmer", "mafft"] }, "pplace_taxonomy": { "type": "string", From 68ada6aa81273ce758be7afd699d3dd11ed140ed Mon Sep 17 00:00:00 2001 From: dariader Date: Wed, 29 Mar 2023 15:38:17 +0300 Subject: [PATCH 028/158] added description to --exclude_taxa param --- nextflow_schema.json | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/nextflow_schema.json b/nextflow_schema.json index e334f391..6213d6ca 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -367,7 +367,7 @@ "type": "string", "default": "mitochondria,chloroplast", "description": "Comma separated list of unwanted taxa, to skip taxa filtering use \"none\"", - "help_text": "Depending on the primers used, PCR might amplify unwanted or off-target DNA. By default sequences originating from mitochondria or chloroplasts are removed. The taxa specified are excluded from further analysis.\nFor example to exclude any taxa that contain mitochondria, chloroplast, or archaea:\n\n```bash\n--exclude_taxa \"mitochondria,chloroplast,archaea\"\n```\n\nIf you prefer not filtering the data, specify:\n\n```bash\n--exclude_taxa \"none\"\n```\n\nPlease note the following requirements:\n\n1. Comma separated list enclosed in quotes\n2. May not contain whitespace characters\n3. Features that contain one or several of these terms in their taxonomical classification are excluded from further analysis\n4. The taxonomy level is not taken into consideration" + "help_text": "Depending on the primers used, PCR might amplify unwanted or off-target DNA. By default sequences originating from mitochondria or chloroplasts are removed. The taxa specified are excluded from further analysis.\nFor example to exclude any taxa that contain mitochondria, chloroplast, or archaea:\n\n```bash\n--exclude_taxa \"mitochondria,chloroplast,archaea\"\n```\n\nIf you prefer not filtering the data, specify:\n\n```bash\n--exclude_taxa \"none\"\n```\n\nPlease note the following requirements:\n\n1. Comma separated list enclosed in quotes\n2. May not contain whitespace characters\n3. Features that contain one or several of these terms in their taxonomical classification are excluded from further analysis\n4. The taxonomy level is not taken into consideration\n5. Taxa names should be as in Taxonomic database (Default: Silva138), example: 'Bacteria', 'Armatimonadia', 'unidentified', 'p__'\n6. Taxon names are case-insensitive and partial match is possible." }, "min_frequency": { "type": "integer", From f9c86b85f808b3018c8794db620827d5d90e9232 Mon Sep 17 00:00:00 2001 From: daniel Date: Wed, 29 Mar 2023 16:22:33 +0200 Subject: [PATCH 029/158] use-phylogenetic-placement-output-in-qiime --- CHANGELOG.md | 2 +- conf/modules.config | 10 ++- conf/test_pplace.config | 10 ++- docs/output.md | 1 + modules/local/format_pplacetax.nf | 99 ++++++++++++++++++++++++++ modules/local/qiime2_intree.nf | 34 +++++++++ subworkflows/local/qiime2_diversity.nf | 10 ++- workflows/ampliseq.nf | 27 ++++--- 8 files changed, 176 insertions(+), 17 deletions(-) create mode 100644 modules/local/format_pplacetax.nf create mode 100644 modules/local/qiime2_intree.nf diff --git a/CHANGELOG.md b/CHANGELOG.md index 35123649..bef0daca 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -7,7 +7,7 @@ and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0 ### `Added` -- [#564](https://github.com/nf-core/ampliseq/pull/564) - Added phylogenetic placement +- [#564](https://github.com/nf-core/ampliseq/pull/564),[#565](https://github.com/nf-core/ampliseq/pull/565) - Added phylogenetic placement ### `Changed` diff --git a/conf/modules.config b/conf/modules.config index 6372a1fe..498351fa 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -573,7 +573,7 @@ process { ] } - withName: 'QIIME2_INASV|QIIME2_INSEQ|QIIME2_INTAX' { + withName: 'QIIME2_INASV|QIIME2_INSEQ|QIIME2_INTAX|QIIME2_INTREE' { publishDir = [ path: { "${params.outdir}/qiime2/input" }, mode: params.publish_dir_mode, @@ -581,6 +581,14 @@ process { ] } + withName: FORMAT_PPLACETAX { + publishDir = [ + path: { "${params.outdir}/pplace" }, + mode: params.publish_dir_mode, + pattern: "*.per_query_unique.tsv" + ] + } + withName: 'QIIME2_INASV_BPAVG' { publishDir = [ path: { "${params.outdir}/qiime2/input" }, diff --git a/conf/test_pplace.config b/conf/test_pplace.config index ade2a9e3..e4f3e737 100644 --- a/conf/test_pplace.config +++ b/conf/test_pplace.config @@ -28,19 +28,23 @@ params { qiime_ref_taxonomy = "greengenes85" filter_ssu = "bac" - //this is to remove low abundance ASVs to reduce runtime of downstream processes + // this is to remove low abundance ASVs to reduce runtime of downstream processes min_samples = 2 min_frequency = 10 - //pplace + // pplace pplace_tree = "https://github.com/nf-core/test-datasets/raw/phyloplace/testdata/cyanos_16s.newick" pplace_aln = "https://github.com/nf-core/test-datasets/raw/phyloplace/testdata/cyanos_16s.alnfna" pplace_model = "GTR+F+I+I+R3" pplace_taxonomy = "https://github.com/nf-core/test-datasets/raw/phyloplace/testdata/cyanos_16s.taxonomy.tsv" pplace_name = "test_pplace" + // Adjust taxonomic levels + dada_tax_agglom_max = 1 + dada_tax_agglom_max = 3 - //Skip some steps to reduce runtime + // Skip some steps to reduce runtime skip_alpha_rarefaction = true skip_fastqc = true + skip_ancom = true } diff --git a/docs/output.md b/docs/output.md index 0ef749a5..7de89e50 100644 --- a/docs/output.md +++ b/docs/output.md @@ -242,6 +242,7 @@ Phylogenetic placement grafts sequences onto a phylogenetic reference tree and o - `pplace/` - `*.graft.*.epa_result.newick`: Full phylogeny with query sequences grafted on to the reference phylogeny, in newick format. - `*.taxonomy.per_query.tsv`: Tab separated file with taxonomy information per query from classification by `gappa examine examinassign` + - `*.per_query_unique.tsv`: Tab separated file with taxonomy information as aboe, but one row per query, by filtering for lowest LWR (likelihood weight ratio) - `*.heattree.tree.svg`: Heattree in SVG format from calling `gappa examine heattree`, see [Gappa documentation](https://github.com/Pbdas/epa-ng/blob/master/README.md) for details. - `pplace/hmmer/`: Contains intermediatary files if HMMER is used - `pplace/mafft/`: Contains intermediatary files if MAFFT is used diff --git a/modules/local/format_pplacetax.nf b/modules/local/format_pplacetax.nf new file mode 100644 index 00000000..ac35cda3 --- /dev/null +++ b/modules/local/format_pplacetax.nf @@ -0,0 +1,99 @@ +process FORMAT_PPLACETAX { + tag "${tax.baseName}" + label 'process_high' + + conda "bioconda::bioconductor-dada2=1.22.0 conda-forge::r-digest=0.6.30" + container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? + 'https://depot.galaxyproject.org/singularity/bioconductor-dada2:1.22.0--r41h399db7b_0' : + 'quay.io/biocontainers/bioconductor-dada2:1.22.0--r41h399db7b_0' }" + + input: + tuple val(meta), path(tax) + + output: + path("*.per_query_unique.tsv"), emit: unique + path("*.taxonomy.tsv") , emit: tsv + path "versions.yml" , emit: versions + + when: + task.ext.when == null || task.ext.when + + script: + """ + #!/usr/bin/env Rscript + + tax = read.table("$tax", header = TRUE, sep = "\t", stringsAsFactors = FALSE, comment.char = '', quote = '') + + df <- data.frame( + name=character(), + LWR=character(), + fract=character(), + aLWR=character(), + afract=character(), + taxopath=character(), + stringsAsFactors=FALSE) + + for (asvid in unique(tax\$name)) { + # subset per ASV ID + temp = subset(tax, tax\$name == asvid) + maxLWR = max(temp\$LWR) + temp = subset(temp, temp\$LWR == maxLWR) + + if ( nrow(temp) == 1 ) { + # STEP 1: if there is just 1 maximum, choose this row + print( paste ( asvid,"was added in STEP 1" ) ) + temp = temp[which.max(temp\$LWR),] + df <- rbind(df, temp) + } else { + # STEP2: be conservative, choose the taxonomy with less entries + # count number of entries in "taxopath", simplified: number of semicolons + temp\$taxlevels = nchar( temp\$taxopath ) - nchar(gsub(';', '', temp\$taxopath )) + 1 + minTaxlevels = min(temp\$taxlevels) + temp = subset(temp, temp\$taxlevels == minTaxlevels) + temp\$taxlevels = NULL + if ( nrow(temp) == 1 ) { + df <- rbind(df, temp) + print( paste ( asvid,"was added in STEP 2" ) ) + } else { + # STEP 3: if number of entries is same, remove last entry + # then, check if reduced entries are identical > if yes, choose any row, if no, repeat + # at that step the taxonomies have same length + print( paste ( asvid,"enters STEP 3" ) ) + list_taxonpath <- str_split( temp\$taxopath, ";") + df_taxonpath <- as.data.frame(do.call(rbind, list_taxonpath)) + for (i in ncol(df_taxonpath):0) { + # choose first column and change taxon to reduced overlap + print(paste("i:",i)) + print(df_taxonpath[,1:i]) + print(paste("len:",length(unique(df_taxonpath[,i])))) + if ( length(unique(df_taxonpath[,i])) == 1 ) { + if (i>1) { + temp\$taxopath <- apply(df_taxonpath[1,1:i], 1, paste, collapse=";")[[1]] + } else if (i==1) { + temp\$taxopath <- df_taxonpath[1,1] + } + df <- rbind(df, temp[1,]) + print( paste ( asvid,"was added with",temp\$taxopath[[1]],"in STEP 3a" ) ) + break + } else if (i==0) { + # if all fails, i.e. there is no consensus on any taxonomic level, use NA + temp\$taxopath <- "NA" + df <- rbind(df, temp[1,]) + print( paste ( asvid,"was added with",temp\$taxopath[[1]],"in STEP 3b" ) ) + } + } + } + } + } + + # output the cleaned, unified, unique taxonomic classification per ASV + write.table(df, file = "${meta.id}.taxonomy.per_query_unique.tsv", row.names = FALSE, col.names = TRUE, quote = FALSE, na = '', sep = "\\t") + + # output ASV taxonomy table for QIIME2 + df_clean <- subset(df, select = c("name","taxopath")) + colnames(df_clean) <- c("ASV_ID","taxonomy") + write.table(df_clean, file = "${meta.id}.taxonomy.tsv", row.names = FALSE, col.names = TRUE, quote = FALSE, na = '', sep = "\\t") + + writeLines(c("\\"${task.process}\\":", paste0(" R: ", paste0(R.Version()[c("major","minor")], collapse = ".")),paste0(" dada2: ", packageVersion("dada2")) ), "versions.yml") + """ +} diff --git a/modules/local/qiime2_intree.nf b/modules/local/qiime2_intree.nf new file mode 100644 index 00000000..f9f35b97 --- /dev/null +++ b/modules/local/qiime2_intree.nf @@ -0,0 +1,34 @@ +process QIIME2_INTREE { + tag "${meta.id}:${meta.model}" + label 'process_low' + + container "quay.io/qiime2/core:2022.11" + + // Exit if running this module with -profile conda / -profile mamba + if (workflow.profile.tokenize(',').intersect(['conda', 'mamba']).size() >= 1) { + exit 1, "QIIME2 does not support Conda. Please use Docker / Singularity / Podman instead." + } + + input: + tuple val(meta), path(tree) + + output: + path("tree.qza") , emit: qza + path "versions.yml", emit: versions + + when: + task.ext.when == null || task.ext.when + + script: + """ + qiime tools import \\ + --type 'Phylogeny[Rooted]' \\ + --input-path $tree \\ + --output-path tree.qza + + cat <<-END_VERSIONS > versions.yml + "${task.process}": + qiime2: \$( qiime --version | sed '1!d;s/.* //' ) + END_VERSIONS + """ +} diff --git a/subworkflows/local/qiime2_diversity.nf b/subworkflows/local/qiime2_diversity.nf index d5ce511d..b3d7f64b 100644 --- a/subworkflows/local/qiime2_diversity.nf +++ b/subworkflows/local/qiime2_diversity.nf @@ -15,6 +15,7 @@ workflow QIIME2_DIVERSITY { ch_metadata ch_asv ch_seq + ch_tree ch_stats //QIIME2_FILTERTAXA.out.tsv ch_metacolumn_pairwise //METADATA_PAIRWISE.out ch_metacolumn_all //METADATA_ALL.out @@ -24,17 +25,20 @@ workflow QIIME2_DIVERSITY { main: //Phylogenetic tree for beta & alpha diversities - QIIME2_TREE ( ch_seq ) + if (!ch_tree) { + QIIME2_TREE ( ch_seq ) + ch_tree = QIIME2_TREE.out.qza + } //Alpha-rarefaction if (!skip_alpha_rarefaction) { - QIIME2_ALPHARAREFACTION ( ch_metadata, ch_asv, QIIME2_TREE.out.qza, ch_stats ) + QIIME2_ALPHARAREFACTION ( ch_metadata, ch_asv, ch_tree, ch_stats ) } //Calculate diversity indices if (!skip_diversity_indices) { - QIIME2_DIVERSITY_CORE ( ch_metadata, ch_asv, QIIME2_TREE.out.qza, ch_stats, diversity_rarefaction_depth ) + QIIME2_DIVERSITY_CORE ( ch_metadata, ch_asv, ch_tree, ch_stats, diversity_rarefaction_depth ) //Print warning if rarefaction depth is <10000 QIIME2_DIVERSITY_CORE.out.depth.subscribe { if ( it.baseName.toString().startsWith("WARNING") ) log.warn it.baseName.toString().replace("WARNING ","QIIME2_DIVERSITY_CORE: ") } diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 32bcc635..13d643da 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -149,6 +149,8 @@ include { FORMAT_TAXRESULTS as FORMAT_TAXRESULTS_ADDSP } from '../modules/local/ include { QIIME2_INSEQ } from '../modules/local/qiime2_inseq' include { QIIME2_FILTERTAXA } from '../modules/local/qiime2_filtertaxa' include { QIIME2_INASV } from '../modules/local/qiime2_inasv' +include { QIIME2_INTREE } from '../modules/local/qiime2_intree' +include { FORMAT_PPLACETAX } from '../modules/local/format_pplacetax' include { FILTER_STATS } from '../modules/local/filter_stats' include { MERGE_STATS as MERGE_STATS_FILTERTAXA } from '../modules/local/merge_stats' include { QIIME2_BARPLOT } from '../modules/local/qiime2_barplot' @@ -500,12 +502,8 @@ workflow AMPLISEQ { } FASTA_NEWICK_EPANG_GAPPA ( ch_pp_data ) ch_versions = ch_versions.mix( FASTA_NEWICK_EPANG_GAPPA.out.versions ) - //TODO: if params.pplace_taxonomy is given, use taxonomy over DADA2 taxonomy for downstream analysis --> FASTA_NEWICK_EPANG_GAPPA.out.taxonomy_per_query --> test_pplace.taxonomy.per_query.tsv - //Use for each ASV the annotation with lowest LWR (likelihood-weight-ration) -> conversion script - //if tie -> use annotation with less entries (i.e. conservative choice) - //if same number of entries -> remove last entry - //if those truncated annotations arent identical, remove one more entry; repeat until identical - //TODO: use newick tree in downstream analysis --> FASTA_NEWICK_EPANG_GAPPA.out.grafted_phylogeny --> test_pplace.graft.test_pplace.epa_result.newick + + ch_pplace_tax = FORMAT_PPLACETAX ( FASTA_NEWICK_EPANG_GAPPA.out.taxonomy_per_query ).tsv } //QIIME2 @@ -528,16 +526,26 @@ workflow AMPLISEQ { // SUBWORKFLOW / MODULES : Downstream analysis with QIIME2 // if ( run_qiime2 ) { - //Import ASV abundance table and sequences into QIIME2 + // Import ASV abundance table and sequences into QIIME2 QIIME2_INASV ( ch_dada2_asv ) QIIME2_INSEQ ( ch_fasta ) - //Import taxonomic classification into QIIME2, if available + // Import phylogenetic tree into QIIME2 + if ( params.pplace_tree ) { + ch_tree = QIIME2_INTREE ( FASTA_NEWICK_EPANG_GAPPA.out.grafted_phylogeny ).qza + } else { ch_tree = Channel.empty() } + + // Import taxonomic classification into QIIME2, if available if ( params.skip_taxonomy ) { log.info "Skip taxonomy classification" ch_tax = Channel.empty() tax_agglom_min = 1 tax_agglom_max = 2 + } else if ( params.pplace_tree && params.pplace_taxonomy) { + log.info "Use EPA-NG / GAPPA taxonomy classification" + ch_tax = QIIME2_INTAX ( ch_pplace_tax ).qza + tax_agglom_min = params.dada_tax_agglom_min + tax_agglom_max = params.dada_tax_agglom_max } else if ( params.dada_ref_taxonomy ) { log.info "Use DADA2 taxonomy classification" ch_tax = QIIME2_INTAX ( ch_dada2_tax ).qza @@ -555,7 +563,7 @@ workflow AMPLISEQ { tax_agglom_max = 2 } - //Filtering ASVs by taxonomy & prevalence & counts + // Filtering ASVs by taxonomy & prevalence & counts if (params.exclude_taxa != "none" || params.min_frequency != 1 || params.min_samples != 1) { QIIME2_FILTERTAXA ( QIIME2_INASV.out.qza, @@ -614,6 +622,7 @@ workflow AMPLISEQ { ch_metadata, ch_asv, ch_seq, + ch_tree, ch_tsv, ch_metacolumn_pairwise, ch_metacolumn_all, From 1e0784102c42dbbc3702a726e5ef5147345f94e6 Mon Sep 17 00:00:00 2001 From: daniel Date: Wed, 29 Mar 2023 16:32:19 +0200 Subject: [PATCH 030/158] fix typo --- conf/test_pplace.config | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/conf/test_pplace.config b/conf/test_pplace.config index e4f3e737..24c81e83 100644 --- a/conf/test_pplace.config +++ b/conf/test_pplace.config @@ -40,7 +40,7 @@ params { pplace_name = "test_pplace" // Adjust taxonomic levels - dada_tax_agglom_max = 1 + dada_tax_agglom_min = 1 dada_tax_agglom_max = 3 // Skip some steps to reduce runtime From 54e349acaf8c1a4ee81cd4a9accd6e0278daddaf Mon Sep 17 00:00:00 2001 From: daniel Date: Wed, 29 Mar 2023 16:33:47 +0200 Subject: [PATCH 031/158] update CHANGELOG --- CHANGELOG.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index bef0daca..a6f59673 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -7,7 +7,7 @@ and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0 ### `Added` -- [#564](https://github.com/nf-core/ampliseq/pull/564),[#565](https://github.com/nf-core/ampliseq/pull/565) - Added phylogenetic placement +- [#564](https://github.com/nf-core/ampliseq/pull/564),[#567](https://github.com/nf-core/ampliseq/pull/567) - Added phylogenetic placement ### `Changed` From 28c4e1de2d4dba33d1fa4c6526bc7bbdc56466e6 Mon Sep 17 00:00:00 2001 From: daniel Date: Thu, 30 Mar 2023 15:20:03 +0200 Subject: [PATCH 032/158] remove some print() --- modules/local/format_pplacetax.nf | 3 --- 1 file changed, 3 deletions(-) diff --git a/modules/local/format_pplacetax.nf b/modules/local/format_pplacetax.nf index ac35cda3..09376354 100644 --- a/modules/local/format_pplacetax.nf +++ b/modules/local/format_pplacetax.nf @@ -63,9 +63,6 @@ process FORMAT_PPLACETAX { df_taxonpath <- as.data.frame(do.call(rbind, list_taxonpath)) for (i in ncol(df_taxonpath):0) { # choose first column and change taxon to reduced overlap - print(paste("i:",i)) - print(df_taxonpath[,1:i]) - print(paste("len:",length(unique(df_taxonpath[,i])))) if ( length(unique(df_taxonpath[,i])) == 1 ) { if (i>1) { temp\$taxopath <- apply(df_taxonpath[1,1:i], 1, paste, collapse=";")[[1]] From 317b059d1d6ad4bfdc1837edd2562b0da1188f21 Mon Sep 17 00:00:00 2001 From: daniel Date: Thu, 30 Mar 2023 15:28:32 +0200 Subject: [PATCH 033/158] update documentation --- docs/output.md | 4 ++-- nextflow_schema.json | 2 +- 2 files changed, 3 insertions(+), 3 deletions(-) diff --git a/docs/output.md b/docs/output.md index 7de89e50..f2dd0587 100644 --- a/docs/output.md +++ b/docs/output.md @@ -271,7 +271,7 @@ Intermediate data imported to QIIME2 is saved as QIIME2 fragments, that can be c #### Taxonomic classification -Taxonomic classification with QIIME2 is typically similar to DADA2 classifications. However, both options are available. When taxonomic classification with DADA2 and QIIME2 is performed, DADA2 classification takes precedence over QIIME2 classifications for all downstream analysis. +Taxonomic classification with QIIME2 is typically similar to DADA2 classifications. However, both options are available. When taxonomic classification with DADA2 and QIIME2 is performed, DADA2 classification takes precedence over QIIME2 classifications for all downstream analysis. Taxonomic classification by phylogenetic placement superseeds DADA2 and QIIME2 classification.
Output files @@ -285,7 +285,7 @@ Taxonomic classification with QIIME2 is typically similar to DADA2 classificatio #### Abundance tables -The abundance tables are the final data for further downstream analysis and visualisations. The tables are based on the computed ASVs and taxonomic classification (DADA2 classification takes precedence over QIIME2 classifications), but after removal of unwanted taxa. Unwanted taxa are often off-targets generated in PCR with primers that are not perfectly specific for the target DNA (can be specified by `--exclude_taxa`), by default mitrochondria and chloroplast sequences are removed because these are frequent unwanted non-bacteria PCR products. +The abundance tables are the final data for further downstream analysis and visualisations. The tables are based on the computed ASVs and taxonomic classification (in the following priotity: phylogenetic placement [EPA-NG, Gappa], DADA2, QIIME2), but after removal of unwanted taxa. Unwanted taxa are often off-targets generated in PCR with primers that are not perfectly specific for the target DNA (can be specified by `--exclude_taxa`), by default mitrochondria and chloroplast sequences are removed because these are frequent unwanted non-bacteria PCR products. All following analysis is based on these filtered tables. diff --git a/nextflow_schema.json b/nextflow_schema.json index e334f391..a64f1fb6 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -280,7 +280,7 @@ }, "pplace_taxonomy": { "type": "string", - "help_text": "Headerless, tab-separated, first column with tree leaves, second column with taxonomy ranks separated by semicolon `;`.", + "help_text": "Headerless, tab-separated, first column with tree leaves, second column with taxonomy ranks separated by semicolon `;`. The results take precedence over DADA2 and QIIME2 classifications.", "description": "Tab-separated file with taxonomy assignments of reference sequences." }, "pplace_name": { From c831737a0125b232a9b7132345f1aa6490f8f63a Mon Sep 17 00:00:00 2001 From: daniel Date: Thu, 30 Mar 2023 15:55:59 +0200 Subject: [PATCH 034/158] simplifiying dada_tax_agglom_* --- CHANGELOG.md | 1 + conf/test.config | 4 ++-- conf/test_pplace.config | 4 ++-- lib/WorkflowAmpliseq.groovy | 9 ++------- nextflow.config | 6 ++---- nextflow_schema.json | 22 ++++------------------ workflows/ampliseq.nf | 12 ++++++------ 7 files changed, 19 insertions(+), 39 deletions(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index a6f59673..dc2b371b 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -12,6 +12,7 @@ and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0 ### `Changed` - [#563](https://github.com/nf-core/ampliseq/pull/563) - Renamed DADA2 taxonomic classification files to include the chosen reference taxonomy abbreviation. +- [#567](https://github.com/nf-core/ampliseq/pull/567) - Renamed `--dada_tax_agglom_min` and `--qiime_tax_agglom_min` to `--tax_agglom_min` and `--dada_tax_agglom_max` and `--qiime_tax_agglom_max` to `--tax_agglom_max` ### `Fixed` diff --git a/conf/test.config b/conf/test.config index 1da1bc4d..77deea96 100644 --- a/conf/test.config +++ b/conf/test.config @@ -38,8 +38,8 @@ params { metadata_category_barplot = "treatment1,badpairwise10" //restrict ANCOM analysis to higher taxonomic levels - dada_tax_agglom_max = 4 - qiime_tax_agglom_max = 4 + tax_agglom_max = 4 + tax_agglom_max = 4 sbdiexport = true diff --git a/conf/test_pplace.config b/conf/test_pplace.config index 24c81e83..b6eaff1d 100644 --- a/conf/test_pplace.config +++ b/conf/test_pplace.config @@ -40,8 +40,8 @@ params { pplace_name = "test_pplace" // Adjust taxonomic levels - dada_tax_agglom_min = 1 - dada_tax_agglom_max = 3 + tax_agglom_min = 1 + tax_agglom_max = 3 // Skip some steps to reduce runtime skip_alpha_rarefaction = true diff --git a/lib/WorkflowAmpliseq.groovy b/lib/WorkflowAmpliseq.groovy index 0ee13f30..a1b0ffa5 100755 --- a/lib/WorkflowAmpliseq.groovy +++ b/lib/WorkflowAmpliseq.groovy @@ -43,13 +43,8 @@ class WorkflowAmpliseq { System.exit(1) } - if (params.dada_tax_agglom_min > params.dada_tax_agglom_max) { - log.error "Incompatible parameters: `--dada_tax_agglom_min` may not be greater than `--dada_tax_agglom_max`." - System.exit(1) - } - - if (params.qiime_tax_agglom_min > params.qiime_tax_agglom_max) { - log.error "Incompatible parameters: `--qiime_tax_agglom_min` may not be greater than `--qiime_tax_agglom_max`." + if (params.tax_agglom_min > params.tax_agglom_max) { + log.error "Incompatible parameters: `--tax_agglom_min` may not be greater than `--tax_agglom_max`." System.exit(1) } diff --git a/nextflow.config b/nextflow.config index 38c5571b..a8afbf08 100644 --- a/nextflow.config +++ b/nextflow.config @@ -47,10 +47,8 @@ params { picrust = false sbdiexport = false addsh = false - dada_tax_agglom_min = 2 - dada_tax_agglom_max = 6 - qiime_tax_agglom_min = 2 - qiime_tax_agglom_max = 6 + tax_agglom_min = 2 + tax_agglom_max = 6 ignore_failed_trimming = false ignore_empty_input_files = false qiime_adonis_formula = null diff --git a/nextflow_schema.json b/nextflow_schema.json index a64f1fb6..ab9851f3 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -426,33 +426,19 @@ "description": "Minimum sample counts to retain a sample for ANCOM analysis. Any sample below that threshold will be removed.", "fa_icon": "fas fa-greater-than-equal" }, - "dada_tax_agglom_min": { + "tax_agglom_min": { "type": "integer", "default": 2, - "description": "Minimum taxonomy agglomeration level for DADA2 classification", + "description": "Minimum taxonomy agglomeration level for taxonomic classifications", "fa_icon": "fas fa-greater-than-equal", "help_text": "Depends on the reference taxonomy database used." }, - "dada_tax_agglom_max": { + "tax_agglom_max": { "type": "integer", "default": 6, - "description": "Maximum taxonomy agglomeration level for DADA2 classification", + "description": "Maximum taxonomy agglomeration level for taxonomic classifications", "fa_icon": "fas fa-greater-than-equal", "help_text": "Depends on the reference taxonomy database used. Most default databases have genus level at 6." - }, - "qiime_tax_agglom_min": { - "type": "integer", - "default": 2, - "description": "Minimum taxonomy agglomeration level for QIIME2 classification", - "fa_icon": "fas fa-greater-than-equal", - "help_text": "Depends on the reference taxonomy database used." - }, - "qiime_tax_agglom_max": { - "type": "integer", - "default": 6, - "description": "Maximum taxonomy agglomeration level for QIIME2 classification", - "fa_icon": "fas fa-greater-than-equal", - "help_text": "Depends on the reference taxonomy database used. Default databases should have genus level at 6." } } }, diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 13d643da..2561fb23 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -544,18 +544,18 @@ workflow AMPLISEQ { } else if ( params.pplace_tree && params.pplace_taxonomy) { log.info "Use EPA-NG / GAPPA taxonomy classification" ch_tax = QIIME2_INTAX ( ch_pplace_tax ).qza - tax_agglom_min = params.dada_tax_agglom_min - tax_agglom_max = params.dada_tax_agglom_max + tax_agglom_min = params.tax_agglom_min + tax_agglom_max = params.tax_agglom_max } else if ( params.dada_ref_taxonomy ) { log.info "Use DADA2 taxonomy classification" ch_tax = QIIME2_INTAX ( ch_dada2_tax ).qza - tax_agglom_min = params.dada_tax_agglom_min - tax_agglom_max = params.dada_tax_agglom_max + tax_agglom_min = params.tax_agglom_min + tax_agglom_max = params.tax_agglom_max } else if ( params.qiime_ref_taxonomy || params.classifier ) { log.info "Use QIIME2 taxonomy classification" ch_tax = QIIME2_TAXONOMY.out.qza - tax_agglom_min = params.qiime_tax_agglom_min - tax_agglom_max = params.qiime_tax_agglom_max + tax_agglom_min = params.tax_agglom_min + tax_agglom_max = params.tax_agglom_max } else { log.info "Use no taxonomy classification" ch_tax = Channel.empty() From a40a6028571d0b81b3a9673c9039258f0a51a4c7 Mon Sep 17 00:00:00 2001 From: daniel Date: Thu, 30 Mar 2023 16:18:21 +0200 Subject: [PATCH 035/158] export relative abundance table with phylogenetic placement taxonomy --- subworkflows/local/qiime2_export.nf | 5 +++++ workflows/ampliseq.nf | 4 ++-- 2 files changed, 7 insertions(+), 2 deletions(-) diff --git a/subworkflows/local/qiime2_export.nf b/subworkflows/local/qiime2_export.nf index d7f57e0f..71b24f43 100644 --- a/subworkflows/local/qiime2_export.nf +++ b/subworkflows/local/qiime2_export.nf @@ -7,6 +7,7 @@ include { QIIME2_EXPORT_RELASV } from '../../modules/local/qiim include { QIIME2_EXPORT_RELTAX } from '../../modules/local/qiime2_export_reltax' include { COMBINE_TABLE as COMBINE_TABLE_QIIME2 } from '../../modules/local/combine_table' include { COMBINE_TABLE as COMBINE_TABLE_DADA2 } from '../../modules/local/combine_table' +include { COMBINE_TABLE as COMBINE_TABLE_PPLACE } from '../../modules/local/combine_table' workflow QIIME2_EXPORT { take: @@ -15,6 +16,7 @@ workflow QIIME2_EXPORT { ch_tax ch_QIIME2_tax_tsv ch_DADA2_tax_tsv + ch_pplace_tax_tsv tax_agglom_min tax_agglom_max @@ -34,6 +36,9 @@ workflow QIIME2_EXPORT { //combine_table.r (optional), similar to DADA2_table.tsv but with additionally taxonomy merged COMBINE_TABLE_DADA2 ( QIIME2_EXPORT_RELASV.out.tsv, QIIME2_EXPORT_ABSOLUTE.out.fasta, ch_DADA2_tax_tsv, 'rel-table-ASV_with-DADA2-tax.tsv' ) + //combine_table.r (optional), similar to DADA2_table.tsv but with additionally taxonomy merged + COMBINE_TABLE_PPLACE ( QIIME2_EXPORT_RELASV.out.tsv, QIIME2_EXPORT_ABSOLUTE.out.fasta, ch_pplace_tax_tsv, 'rel-table-ASV_with-PPLACE-tax.tsv' ) + emit: abs_fasta = QIIME2_EXPORT_ABSOLUTE.out.fasta abs_tsv = QIIME2_EXPORT_ABSOLUTE.out.tsv diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 2561fb23..7bbb8429 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -504,7 +504,7 @@ workflow AMPLISEQ { ch_versions = ch_versions.mix( FASTA_NEWICK_EPANG_GAPPA.out.versions ) ch_pplace_tax = FORMAT_PPLACETAX ( FASTA_NEWICK_EPANG_GAPPA.out.taxonomy_per_query ).tsv - } + } else { ch_pplace_tax = Channel.empty() } //QIIME2 if ( run_qiime2 ) { @@ -586,7 +586,7 @@ workflow AMPLISEQ { } //Export various ASV tables if (!params.skip_abundance_tables) { - QIIME2_EXPORT ( ch_asv, ch_seq, ch_tax, QIIME2_TAXONOMY.out.tsv, ch_dada2_tax, tax_agglom_min, tax_agglom_max ) + QIIME2_EXPORT ( ch_asv, ch_seq, ch_tax, QIIME2_TAXONOMY.out.tsv, ch_dada2_tax, ch_pplace_tax, tax_agglom_min, tax_agglom_max ) } if (!params.skip_barplot) { From cdb9abb7697e30c1b0c1c1f1df4bbdeea605489c Mon Sep 17 00:00:00 2001 From: daniel Date: Thu, 30 Mar 2023 16:29:32 +0200 Subject: [PATCH 036/158] update docs --- docs/output.md | 1 + 1 file changed, 1 insertion(+) diff --git a/docs/output.md b/docs/output.md index f2dd0587..28e5abc6 100644 --- a/docs/output.md +++ b/docs/output.md @@ -318,6 +318,7 @@ Absolute abundance tables produced by the previous steps contain count data, but - `rel-table-ASV.tsv`: Tab-separated relative abundance table for all ASVs. - `rel-table-ASV_with-DADA2-tax.tsv`: Tab-separated table for all ASVs with DADA2 taxonomic classification, sequence and relative abundance. - `rel-table-ASV_with-QIIME2-tax.tsv`: Tab-separated table for all ASVs with QIIME2 taxonomic classification, sequence and relative abundance. + - `rel-table-ASV_with-PPLACE-tax.tsv`: Tab-separated table for all ASVs with EPA-NG - Gappa taxonomic classification, sequence and relative abundance.
From 46e2a8fc504f97e8110413d1c6dba8b6e305cf8f Mon Sep 17 00:00:00 2001 From: jtangrot Date: Fri, 31 Mar 2023 09:38:11 +0200 Subject: [PATCH 037/158] Convert sintax output and enable input to qiime --- bin/convert_sintax_output.py | 70 +++++++++++++++++++++++ conf/modules.config | 10 +++- modules.json | 26 +++------ modules/local/format_taxresults_sintax.nf | 31 ++++++++++ nextflow.config | 2 + nextflow_schema.json | 14 +++++ workflows/ampliseq.nf | 39 ++++++++----- 7 files changed, 158 insertions(+), 34 deletions(-) create mode 100755 bin/convert_sintax_output.py create mode 100644 modules/local/format_taxresults_sintax.nf diff --git a/bin/convert_sintax_output.py b/bin/convert_sintax_output.py new file mode 100755 index 00000000..39a4100e --- /dev/null +++ b/bin/convert_sintax_output.py @@ -0,0 +1,70 @@ +#!/usr/bin/env python3 +##### Program description ####### +# +# Title: Convert sintax output to DADA2-like tsv format +# +# Author(s): Lokeshwaran Manoharan, Jeanette Tångrot +# +# Description: +# +# List of subroutines: +# +# Usage: convert_sintax_output.py -i -o -t +################################## + +import re +import argparse + +usage = """This program takes the output of SINTAX and creates taxonomy file in a format suitable for the ampliseq pipeline""" + +parser = argparse.ArgumentParser(description=usage) + +parser.add_argument( + "-i", "--infile", dest="infile", metavar="INFILE", type=argparse.FileType("r"), help="SINTAX output", required=True +) + +parser.add_argument( + "-o", + "--outfile", + dest="outfile", + metavar="OUTFILE", + type=argparse.FileType("w"), + help="Final Taxonomy in tsv format", +) + +parser.add_argument( + "-t", + "--taxlevels", + dest="taxlevels", + metavar="TAXLEVELS", + help="Taxonomy levels to use, comma separated list", + default="", +) + +args = parser.parse_args() + + +def complete_list(some_list, target_len): + return some_list[:target_len] + ["NA"] * (target_len - len(some_list)) + + +header = ["ASV"] + args.taxlevels.split(",") +num_taxa = len(header) - 1 +print("\t".join(header), file=args.outfile) + +for line in args.infile: + line = line.rstrip("\n") + tmp_list = line.split("\t") + if tmp_list[3] != "": + annot = re.sub(".\:", "", tmp_list[3]) + tmp_list1 = annot.split(",") + if len(tmp_list1) != num_taxa: + tmp_list1 = complete_list(tmp_list1, num_taxa) + print(tmp_list[0], "\t".join(tmp_list1), sep="\t", file=args.outfile) + else: + print(tmp_list[0], "\t".join(tmp_list1), sep="\t", file=args.outfile) + else: + print(tmp_list[0], "\t".join(["NA"] * num_taxa), sep="\t", file=args.outfile) + +args.outfile.close() +args.infile.close() diff --git a/conf/modules.config b/conf/modules.config index b6711efb..0042e160 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -390,7 +390,7 @@ process { } withName: VSEARCH_SINTAX { - ext.args = "--randseed ${params.seed}" + ext.args = "--sintax_cutoff 0.8 --randseed ${params.seed}" cpus = 1 publishDir = [ [ @@ -401,6 +401,14 @@ process { ] } + withName: FORMAT_TAXRESULTS_SINTAX { + publishDir = [ + path: { "${params.outdir}/sintax" }, + mode: params.publish_dir_mode, + pattern: "*.tsv" + ] + } + withName: VSEARCH_USEARCHGLOBAL { ext.args = '--top_hits_only --output_no_hits --maxaccepts 50 --query_cov 0.9' publishDir = [ diff --git a/modules.json b/modules.json index 31c011f5..382df979 100644 --- a/modules.json +++ b/modules.json @@ -8,47 +8,35 @@ "custom/dumpsoftwareversions": { "branch": "master", "git_sha": "c8e35eb2055c099720a75538d1b8adb3fb5a464c", - "installed_by": [ - "modules" - ] + "installed_by": ["modules"] }, "cutadapt": { "branch": "master", "git_sha": "c8e35eb2055c099720a75538d1b8adb3fb5a464c", - "installed_by": [ - "modules" - ] + "installed_by": ["modules"] }, "fastqc": { "branch": "master", "git_sha": "c8e35eb2055c099720a75538d1b8adb3fb5a464c", - "installed_by": [ - "modules" - ] + "installed_by": ["modules"] }, "multiqc": { "branch": "master", "git_sha": "ee80d14721e76e2e079103b8dcd5d57129e584ba", - "installed_by": [ - "modules" - ] + "installed_by": ["modules"] }, "vsearch/sintax": { "branch": "master", "git_sha": "0f8a77ff00e65eaeebc509b8156eaa983192474b", - "installed_by": [ - "modules" - ] + "installed_by": ["modules"] }, "vsearch/usearchglobal": { "branch": "master", "git_sha": "c8e35eb2055c099720a75538d1b8adb3fb5a464c", - "installed_by": [ - "modules" - ] + "installed_by": ["modules"] } } } } } -} \ No newline at end of file +} diff --git a/modules/local/format_taxresults_sintax.nf b/modules/local/format_taxresults_sintax.nf new file mode 100644 index 00000000..8a27153f --- /dev/null +++ b/modules/local/format_taxresults_sintax.nf @@ -0,0 +1,31 @@ +process FORMAT_TAXRESULTS_SINTAX { + label 'process_low' + + conda "conda-forge::python=3.9" + container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? + 'https://depot.galaxyproject.org/singularity/python:3.9' : + 'quay.io/biocontainers/python:3.9' }" + + input: + tuple val(meta), path(taxtable) + val(outfile) + val(taxlevels_input) + + output: + path(outfile) , emit: tsv + path "versions.yml", emit: versions + + when: + task.ext.when == null || task.ext.when + + script: + def taxlevels = taxlevels_input ? taxlevels_input : "Kingdom,Phylum,Class,Order,Family,Genus,Species" + """ + convert_sintax_output.py -i $taxtable -o $outfile -t $taxlevels + + cat <<-END_VERSIONS > versions.yml + "${task.process}": + python: \$(python --version 2>&1 | sed 's/Python //g') + END_VERSIONS + """ +} diff --git a/nextflow.config b/nextflow.config index 1e54da4f..a54cbd59 100644 --- a/nextflow.config +++ b/nextflow.config @@ -49,6 +49,8 @@ params { addsh = false dada_tax_agglom_min = 2 dada_tax_agglom_max = 6 + sintax_tax_agglom_min = 2 + sintax_tax_agglom_max = 6 qiime_tax_agglom_min = 2 qiime_tax_agglom_max = 6 ignore_failed_trimming = false diff --git a/nextflow_schema.json b/nextflow_schema.json index eeab4d28..c2205662 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -427,6 +427,20 @@ "fa_icon": "fas fa-greater-than-equal", "help_text": "Depends on the reference taxonomy database used. Most default databases have genus level at 6." }, + "sintax_tax_agglom_min": { + "type": "integer", + "default": 2, + "description": "Minimum taxonomy agglomeration level for SINTAX classification", + "fa_icon": "fas fa-greater-than-equal", + "help_text": "Depends on the reference taxonomy database used." + }, + "sintax_tax_agglom_max": { + "type": "integer", + "default": 6, + "description": "Maximum taxonomy agglomeration level for SINTAX classification", + "fa_icon": "fas fa-greater-than-equal", + "help_text": "Depends on the reference taxonomy database used. Most databases have genus level at 6." + }, "qiime_tax_agglom_min": { "type": "integer", "default": 2, diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 04a6f970..c2cb27ef 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -146,6 +146,7 @@ include { DADA2_ADDSPECIES } from '../modules/local/dada2_addspecie include { ASSIGNSH } from '../modules/local/assignsh' include { FORMAT_TAXRESULTS as FORMAT_TAXRESULTS_STD } from '../modules/local/format_taxresults' include { FORMAT_TAXRESULTS as FORMAT_TAXRESULTS_ADDSP } from '../modules/local/format_taxresults' +include { FORMAT_TAXRESULTS_SINTAX } from '../modules/local/format_taxresults_sintax' include { QIIME2_INSEQ } from '../modules/local/qiime2_inseq' include { QIIME2_FILTERTAXA } from '../modules/local/qiime2_filtertaxa' include { QIIME2_INASV } from '../modules/local/qiime2_inasv' @@ -354,7 +355,7 @@ workflow AMPLISEQ { } // - // SUBWORKFLOW / MODULES : Taxonomic classification with DADA2 and/or QIIME2 + // SUBWORKFLOW / MODULES : Taxonomic classification with DADA2, SINTAX and/or QIIME2 // ch_fasta = ch_dada2_fasta @@ -478,19 +479,24 @@ workflow AMPLISEQ { } // Sintax - // This will only run if --sintax_ref_taxonomy is defined, i.e. the channel - // ch_sintax_ref_taxonomy is not empty - FORMAT_TAXONOMY_SINTAX ( ch_sintax_ref_taxonomy.collect() ) - ch_sintaxdb = FORMAT_TAXONOMY_SINTAX.out.db - ch_fasta - .map { - fasta -> - def meta = [:] - meta.id = "ASV_tax_sintax" - [ meta, fasta ] } - .set { ch_fasta_sintax } - VSEARCH_SINTAX( ch_fasta_sintax, ch_sintaxdb ) - ch_versions = ch_versions.mix(VSEARCH_SINTAX.out.versions) + // This will only run if --sintax_ref_taxonomy is defined, + // i.e. if the channel ch_sintax_ref_taxonomy is not empty + if (!params.skip_taxonomy) { + FORMAT_TAXONOMY_SINTAX ( ch_sintax_ref_taxonomy.collect() ) + ch_sintaxdb = FORMAT_TAXONOMY_SINTAX.out.db + ch_fasta + .map { + fasta -> + def meta = [:] + meta.id = "ASV_tax_sintax.raw" + [ meta, fasta ] } + .set { ch_fasta_sintax } + VSEARCH_SINTAX( ch_fasta_sintax, ch_sintaxdb ) + ch_versions = ch_versions.mix(VSEARCH_SINTAX.out.versions) + FORMAT_TAXRESULTS_SINTAX( VSEARCH_SINTAX.out.tsv, 'ASV_tax_sintax.tsv', taxlevels ) + //FORMAT_TAXRESULTS_SINTAX( VSEARCH_SINTAX.out.tsv, 'ASV_tax_sintax.tsv', 'Kingdom,Genus' ) + ch_sintax_tax = FORMAT_TAXRESULTS_SINTAX.out.tsv + } //QIIME2 if ( run_qiime2 ) { @@ -522,6 +528,11 @@ workflow AMPLISEQ { ch_tax = Channel.empty() tax_agglom_min = 1 tax_agglom_max = 2 + } else if ( params.sintax_ref_taxonomy ) { + log.info "Use SINTAX taxonomy classification" + ch_tax = QIIME2_INTAX ( ch_sintax_tax ).qza + tax_agglom_min = params.sintax_tax_agglom_min + tax_agglom_max = params.sintax_tax_agglom_max } else if ( params.dada_ref_taxonomy ) { log.info "Use DADA2 taxonomy classification" ch_tax = QIIME2_INTAX ( ch_dada2_tax ).qza From 11ec174e1ce91a944ad78a0cc1796d68f83edfa8 Mon Sep 17 00:00:00 2001 From: lokeshbio Date: Fri, 31 Mar 2023 09:55:19 +0200 Subject: [PATCH 038/158] filter coi test 1 --- bin/filt_InsBiom_asv.py | 107 ++++++++++++++++++++++++++++++++ bin/filt_coi_stats.r | 23 +++++++ conf/modules.config | 8 +++ modules/local/filter_coi_asv.nf | 36 +++++++++++ workflows/ampliseq.nf | 12 ++++ 5 files changed, 186 insertions(+) create mode 100755 bin/filt_InsBiom_asv.py create mode 100644 bin/filt_coi_stats.r create mode 100644 modules/local/filter_coi_asv.nf diff --git a/bin/filt_InsBiom_asv.py b/bin/filt_InsBiom_asv.py new file mode 100755 index 00000000..5c32e887 --- /dev/null +++ b/bin/filt_InsBiom_asv.py @@ -0,0 +1,107 @@ +#!/usr/bin/env python3 +##### Program description ####### +# +# Title: Subsetting Insect Biome ASVs +# +# Author(s): Lokeshwaran Manoharan +# +# +# +# Description: The ASVs that are supposed to be of particular length (418 ± nx3 and between 403 and 418), possibly does not contain any stop codon in the right reading frame. +# +# List of subroutines: +# +# +# +# Overall procedure: using functions and dictionaries in python +# +# Usage: filt_InsBiom_asv.py +# +################################## + +import re +import sys +import copy +import argparse + +usage = '''This program takes the output from the AmpliSeq dada2 pipeline and filteres the COI ASVs and only keep the ASVs of particular length (418 ± nx3 and between 403 and 418) and does not contain any stop codon in the right reading frame''' + +parser = argparse.ArgumentParser(description=usage) + +parser.add_argument( + '-f', '--fasta', + dest='fasta', + type=argparse.FileType('r'), + help='Fasta file of the ASVs from the AmpliSeq pipeline', + required=True + ) + +parser.add_argument( + '-t', '--count-table', + dest='count', + type=argparse.FileType('r'), + help='Count table file of the ASVs from the AmpliSeq pipeline', + required=True + ) + +parser.add_argument( + '-p', '--out-prefix', + dest='prefix', + type=str, + help='Prefix of the output files with filtered ASVs and the corresponding count table. The output files will be in *_filtered.* format', + required=True + ) + + +args = parser.parse_args() + + +p1 = re.compile('\t') +p2 = re.compile('>') + +def check_asv(seq): + sub_seq = seq[1:] + stop_list = ['TAA', 'TAG'] + sub_list = [] + for x in range(0, len(sub_seq), 3): + sub_list.append(sub_seq[x:x+3]) + if not 402 <= len(sub_seq) <= 417: + return False + elif not len(sub_seq)%3 == 0: + return False + elif any(x in sub_list for x in stop_list): + return False + else: + return True + +Out_Seq = open(args.prefix +'_filtered.fna', 'w') +Out_list = open(args.prefix+'_filtered.list', 'w') +Out_table = open(args.prefix+'_filtered.table.tsv', 'w') + +count_dict = {} + +count = 0 +for line in args.count: + line = line.rstrip('\n') + if count == 0: + print(line, file = Out_table) + count += 1 + else: + tmp_list = re.split(p1, line) + count_dict[tmp_list[0]] = line + +for line in args.fasta: + line = line.rstrip('\n') + if re.match(p2, line) is not None: + bin_head = re.sub(p2, '', line) + elif check_asv(line): + print('>', bin_head,'\n', line, file = Out_Seq, sep='') + print(bin_head, file = Out_list) + print(count_dict[bin_head], file = Out_table) + +args.count.close() +args.fasta.close() +Out_Seq.close() +Out_list.close() +Out_table.close() + diff --git a/bin/filt_coi_stats.r b/bin/filt_coi_stats.r new file mode 100644 index 00000000..df876ce5 --- /dev/null +++ b/bin/filt_coi_stats.r @@ -0,0 +1,23 @@ +#!/usr/bin/env Rscript + +args = commandArgs(trailingOnly=TRUE) + +if(length(args) < 2){ + stop("Usage: filt_coi_stats.r ") +} + +dada2_stats <- args[1] +coi_filt <- args[2] + + <- read.table(dada2_stats, header = T, check.names = F, row.names = 1) + +filt_table <- read.table(coi_filt, header = T, check.names = F, row.names = 1) + +filt_counts <- as.data.frame(colSums(filt_table)) +names(filt_counts) <- "coi_filtered" + +new_stats <- merge(old_stats, filt_counts, by = 'row.names') +new_stats$sample <- row.names(new_stats) + +write.table(new_stats, file = "stats.filt.tsv", row.names=FALSE, sep="\t", col.names = TRUE, quote = FALSE, na = '') + diff --git a/conf/modules.config b/conf/modules.config index d0bf626a..853cdf3f 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -309,6 +309,14 @@ process { ] } + withName: FILTER_COI_ASV { + publishDir = [ + path: { "${params.outdir}/COI_filtered" }, + mode: params.publish_dir_mode, + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + ] + } + withName: 'MERGE_STATS_STD|MERGE_STATS_FILTERSSU|MERGE_STATS_FILTERTAXA' { publishDir = [ path: { "${params.outdir}" }, diff --git a/modules/local/filter_coi_asv.nf b/modules/local/filter_coi_asv.nf new file mode 100644 index 00000000..a4d418c8 --- /dev/null +++ b/modules/local/filter_coi_asv.nf @@ -0,0 +1,36 @@ +process FILTER_COI_ASV { + tag "${fasta}" + label 'process_low' + + conda (params.enable_conda ? { exit 1 "QIIME2 has no conda package" } : null) + container "quay.io/qiime2/core:2022.8" + + input: + path(fasta) + path(asv) + path(dada2stats) + + output: + path( "ASV_filtered.table.tsv" ) , emit: asv + path( "ASV_filtered.fna" ) , emit: fasta + path( "ASV_filtered.list" ) , emit: list + path( "stats.filt.tsv" ), emit: stas + path "versions.yml" , emit: versions + + when: + task.ext.when == null || task.ext.when + + script: + """ + filt_InsBiom_asv.py -f ${fasta} -t ${asv} -p ASV + filt_coi_stats.r ${dada2stats} ASV_filtered.table.tsv + + cat <<-END_VERSIONS > versions.yml + "${task.process}": + python: \$(python --version 2>&1 | sed 's/Python //g') + R: \$(R --version | sed -n 1p | sed 's/R version //g' | sed 's/\\s.*\$//') + END_VERSIONS + + """ + +} \ No newline at end of file diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 864d0802..17616d1d 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -132,6 +132,7 @@ include { FILTER_SSU } from '../modules/local/filter_ssu' include { FILTER_LEN_ASV } from '../modules/local/filter_len_asv' include { MERGE_STATS as MERGE_STATS_FILTERSSU } from '../modules/local/merge_stats' include { MERGE_STATS as MERGE_STATS_FILTERLENASV } from '../modules/local/merge_stats' +include { FILTER_COI_ASV } from '../modules/local/filter_coi_asv' include { FORMAT_FASTAINPUT } from '../modules/local/format_fastainput' include { FORMAT_TAXONOMY } from '../modules/local/format_taxonomy' include { ITSX_CUTASV } from '../modules/local/itsx_cutasv' @@ -347,6 +348,17 @@ workflow AMPLISEQ { ch_dada2_asv = FILTER_LEN_ASV.out.asv } + // + // Modules : COI ASV Filtering + // + if (params.filter_coi_asv) { + FILTER_COI_ASV ( ch_dada2_fasta, ch_dada2_asv, ch_stats ) + ch_versions = ch_versions.mix(FILTER_COI_ASV.out.versions.ifEmpty(null)) + ch_stats = FILTER_COI_ASV.out.stats + ch_dada2_fasta = FILTER_COI_ASV.out.fasta + ch_dada2_asv = FILTER_COI_ASV.out.asv + } + // // SUBWORKFLOW / MODULES : Taxonomic classification with DADA2 and/or QIIME2 // From 9b87c66939d8e13e34a70ee3493abeb5f381ed8d Mon Sep 17 00:00:00 2001 From: daniel Date: Fri, 31 Mar 2023 11:26:08 +0200 Subject: [PATCH 039/158] update nf-core/phyloplace citation --- CITATIONS.md | 4 +++- 1 file changed, 3 insertions(+), 1 deletion(-) diff --git a/CITATIONS.md b/CITATIONS.md index 832eee3c..997b12b2 100644 --- a/CITATIONS.md +++ b/CITATIONS.md @@ -69,7 +69,9 @@ ### Phylogenetic placement -- nf-core/phyloplace (https://github.com/nf-core/phyloplace, https://nf-co.re/phyloplace) was originally written by Daniel Lundin. +- [nf-core/phyloplace](https://nf-co.re/phyloplace) + + > Daniel Lundin. (2023). nf-core/phyloplace: First release (1.0.0). Zenodo. https://doi.org/10.5281/zenodo.7643948 - [HMMER](https://pubmed.ncbi.nlm.nih.gov/22039361/) From bf5c4f221d80d9a68c5c5ac3d70f641c4f9f9ec2 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Fri, 31 Mar 2023 13:05:05 +0200 Subject: [PATCH 040/158] Enable cutits for sintax --- bin/convert_sintax_output.py | 55 ++++++++++++++++++++--- modules/local/format_taxresults_sintax.nf | 3 +- workflows/ampliseq.nf | 52 +++++++++++++++------ 3 files changed, 88 insertions(+), 22 deletions(-) diff --git a/bin/convert_sintax_output.py b/bin/convert_sintax_output.py index 39a4100e..c7c0d2d1 100755 --- a/bin/convert_sintax_output.py +++ b/bin/convert_sintax_output.py @@ -9,7 +9,7 @@ # # List of subroutines: # -# Usage: convert_sintax_output.py -i -o -t +# Usage: convert_sintax_output.py -i -f -o -t ################################## import re @@ -20,7 +20,23 @@ parser = argparse.ArgumentParser(description=usage) parser.add_argument( - "-i", "--infile", dest="infile", metavar="INFILE", type=argparse.FileType("r"), help="SINTAX output", required=True + "-i", + "--infile", + dest="infile", + metavar="INFILE", + type=argparse.FileType("r"), + help="SINTAX output", + required=True, +) + +parser.add_argument( + "-f", + "--fasta", + dest="fastafile", + metavar="FASTAFILE", + type=argparse.FileType("r"), + help="FASTA file with sequences", + required=True, ) parser.add_argument( @@ -30,6 +46,7 @@ metavar="OUTFILE", type=argparse.FileType("w"), help="Final Taxonomy in tsv format", + default="sintax_taxonomy.tsv", ) parser.add_argument( @@ -48,23 +65,47 @@ def complete_list(some_list, target_len): return some_list[:target_len] + ["NA"] * (target_len - len(some_list)) -header = ["ASV"] + args.taxlevels.split(",") -num_taxa = len(header) - 1 +# Read fasta file and store as dictionary +# If multiple sequences have the same name only the first will be stored +seqs = dict() +seq = "" +name = "" +for line in args.fastafile: + if line.startswith(">"): + if seq != "" and name != "": + if name not in seqs: + seqs[name] = seq + seq = "" + name = line.lstrip(">").rstrip() + else: + seq = seq + line.rstrip("\n") +if seq != "" and name != "" and name not in seqs: + seqs[name] = seq + +# Print header to outfile +if args.taxlevels != "": + header = ["ASV"] + args.taxlevels.split(",") + ["sequence"] +else: + header = ["ASV", "sequence"] +num_taxa = len(header) - 2 print("\t".join(header), file=args.outfile) +# Read sintax file, parse results, and write taxonomies together with sequence to outfile for line in args.infile: line = line.rstrip("\n") tmp_list = line.split("\t") + sequence = seqs[tmp_list[0]] if tmp_list[0] in seqs else "" if tmp_list[3] != "": annot = re.sub(".\:", "", tmp_list[3]) tmp_list1 = annot.split(",") if len(tmp_list1) != num_taxa: tmp_list1 = complete_list(tmp_list1, num_taxa) - print(tmp_list[0], "\t".join(tmp_list1), sep="\t", file=args.outfile) + print(tmp_list[0], "\t".join(tmp_list1), sequence, sep="\t", file=args.outfile) else: - print(tmp_list[0], "\t".join(tmp_list1), sep="\t", file=args.outfile) + print(tmp_list[0], "\t".join(tmp_list1), sequence, sep="\t", file=args.outfile) else: - print(tmp_list[0], "\t".join(["NA"] * num_taxa), sep="\t", file=args.outfile) + print(tmp_list[0], "\t".join(["NA"] * num_taxa), sequence, sep="\t", file=args.outfile) + args.outfile.close() args.infile.close() diff --git a/modules/local/format_taxresults_sintax.nf b/modules/local/format_taxresults_sintax.nf index 8a27153f..9af239f6 100644 --- a/modules/local/format_taxresults_sintax.nf +++ b/modules/local/format_taxresults_sintax.nf @@ -8,6 +8,7 @@ process FORMAT_TAXRESULTS_SINTAX { input: tuple val(meta), path(taxtable) + path(fastafile) val(outfile) val(taxlevels_input) @@ -21,7 +22,7 @@ process FORMAT_TAXRESULTS_SINTAX { script: def taxlevels = taxlevels_input ? taxlevels_input : "Kingdom,Phylum,Class,Order,Family,Genus,Species" """ - convert_sintax_output.py -i $taxtable -o $outfile -t $taxlevels + convert_sintax_output.py -i $taxtable -f $fastafile -o $outfile -t $taxlevels cat <<-END_VERSIONS > versions.yml "${task.process}": diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index c2cb27ef..081e5627 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -134,12 +134,13 @@ include { BARRNAP } from '../modules/local/barrnap' include { BARRNAPSUMMARY } from '../modules/local/barrnapsummary' include { FILTER_SSU } from '../modules/local/filter_ssu' include { FILTER_LEN_ASV } from '../modules/local/filter_len_asv' -include { MERGE_STATS as MERGE_STATS_FILTERSSU } from '../modules/local/merge_stats' +include { MERGE_STATS as MERGE_STATS_FILTERSSU } from '../modules/local/merge_stats' include { MERGE_STATS as MERGE_STATS_FILTERLENASV } from '../modules/local/merge_stats' include { FORMAT_FASTAINPUT } from '../modules/local/format_fastainput' include { FORMAT_TAXONOMY } from '../modules/local/format_taxonomy' include { FORMAT_TAXONOMY_SINTAX } from '../modules/local/format_taxonomy_sintax' -include { ITSX_CUTASV } from '../modules/local/itsx_cutasv' +include { ITSX_CUTASV as ITSX_CUTASV_DADA2 } from '../modules/local/itsx_cutasv' +include { ITSX_CUTASV as ITSX_CUTASV_SINTAX } from '../modules/local/itsx_cutasv' include { MERGE_STATS as MERGE_STATS_STD} from '../modules/local/merge_stats' include { DADA2_TAXONOMY } from '../modules/local/dada2_taxonomy' include { DADA2_ADDSPECIES } from '../modules/local/dada2_addspecies' @@ -431,9 +432,9 @@ workflow AMPLISEQ { else if (params.cut_its == "its2") { outfile = params.its_partial ? "ASV_ITS_seqs.ITS2.full_and_partial.fasta" : "ASV_ITS_seqs.ITS2.fasta" } - ITSX_CUTASV ( ch_fasta, outfile ) - ch_versions = ch_versions.mix(ITSX_CUTASV.out.versions.ifEmpty(null)) - ch_cut_fasta = ITSX_CUTASV.out.fasta + ITSX_CUTASV_DADA2 ( ch_fasta, outfile ) + ch_versions = ch_versions.mix(ITSX_CUTASV_DADA2.out.versions.ifEmpty(null)) + ch_cut_fasta = ITSX_CUTASV_DADA2.out.fasta DADA2_TAXONOMY ( ch_cut_fasta, ch_assigntax, 'ASV_ITS_tax.tsv', taxlevels ) ch_versions = ch_versions.mix(DADA2_TAXONOMY.out.versions) FORMAT_TAXRESULTS_STD ( DADA2_TAXONOMY.out.tsv, ch_fasta, 'ASV_tax.tsv' ) @@ -484,17 +485,40 @@ workflow AMPLISEQ { if (!params.skip_taxonomy) { FORMAT_TAXONOMY_SINTAX ( ch_sintax_ref_taxonomy.collect() ) ch_sintaxdb = FORMAT_TAXONOMY_SINTAX.out.db - ch_fasta - .map { - fasta -> - def meta = [:] - meta.id = "ASV_tax_sintax.raw" - [ meta, fasta ] } - .set { ch_fasta_sintax } + if (params.cut_its == "none") { + ASV_tax_name = "ASV_tax_sintax" + ch_fasta + .map { + fasta -> + def meta = [:] + meta.id = ASV_tax_name + ".raw" + [ meta, fasta ] } + .set { ch_fasta_sintax } + } else { + ASV_tax_name = "ASV_ITS_tax_sintax" + if (params.cut_its == "full") { + outfile = params.its_partial ? "ASV_ITS_seqs.full_and_partial.fasta" : "ASV_ITS_seqs.full.fasta" + } + else if (params.cut_its == "its1") { + outfile = params.its_partial ? "ASV_ITS_seqs.ITS1.full_and_partial.fasta" : "ASV_ITS_seqs.ITS1.fasta" + } + else if (params.cut_its == "its2") { + outfile = params.its_partial ? "ASV_ITS_seqs.ITS2.full_and_partial.fasta" : "ASV_ITS_seqs.ITS2.fasta" + } + ITSX_CUTASV_SINTAX ( ch_fasta, outfile ) + ch_versions = ch_versions.mix(ITSX_CUTASV_SINTAX.out.versions.ifEmpty(null)) + ch_cut_fasta = ITSX_CUTASV_SINTAX.out.fasta + ch_cut_fasta + .map { + fasta -> + def meta = [:] + meta.id = ASV_tax_name + ".raw" + [ meta, fasta ] } + .set { ch_fasta_sintax } + } VSEARCH_SINTAX( ch_fasta_sintax, ch_sintaxdb ) ch_versions = ch_versions.mix(VSEARCH_SINTAX.out.versions) - FORMAT_TAXRESULTS_SINTAX( VSEARCH_SINTAX.out.tsv, 'ASV_tax_sintax.tsv', taxlevels ) - //FORMAT_TAXRESULTS_SINTAX( VSEARCH_SINTAX.out.tsv, 'ASV_tax_sintax.tsv', 'Kingdom,Genus' ) + FORMAT_TAXRESULTS_SINTAX( VSEARCH_SINTAX.out.tsv, ch_fasta, ASV_tax_name + '.tsv', taxlevels ) ch_sintax_tax = FORMAT_TAXRESULTS_SINTAX.out.tsv } From c5e813ffe4abcb3a0811be3a26c9c1d3b17e8b4e Mon Sep 17 00:00:00 2001 From: jtangrot Date: Fri, 31 Mar 2023 13:26:12 +0200 Subject: [PATCH 041/158] Fix typo --- workflows/ampliseq.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 1f04aac0..b12aaba3 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -70,7 +70,7 @@ if (params.qiime_ref_taxonomy && !params.skip_taxonomy && !params.classifier) { if (params.sintax_ref_taxonomy && !params.skip_taxonomy) { ch_sintax_ref_taxonomy = Channel.fromList(params.sintax_ref_databases[params.sintax_ref_taxonomy]["file"]).map { file(it) } - val_sintax_ref_taxonomy = params.sintax_ref_taxonomy.replace('=','_').replace('.','_')" + val_sintax_ref_taxonomy = params.sintax_ref_taxonomy.replace('=','_').replace('.','_') } else { ch_sintax_ref_taxonomy = Channel.empty() val_sintax_ref_taxonomy = "none" From f5c855c50e64f2b6798d884f4dabca6e1e2264be Mon Sep 17 00:00:00 2001 From: jtangrot Date: Fri, 31 Mar 2023 13:51:42 +0200 Subject: [PATCH 042/158] Fix quotation marks --- workflows/ampliseq.nf | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index b12aaba3..6b4b2800 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -452,7 +452,7 @@ workflow AMPLISEQ { ITSX_CUTASV_DADA2 ( ch_fasta, outfile ) ch_versions = ch_versions.mix(ITSX_CUTASV_DADA2.out.versions.ifEmpty(null)) ch_cut_fasta = ITSX_CUTASV_DADA2.out.fasta - DADA2_TAXONOMY ( ch_cut_fasta, ch_assigntax, 'ASV_ITS_tax.${val_dada_ref_taxonomy}.tsv', taxlevels ) + DADA2_TAXONOMY ( ch_cut_fasta, ch_assigntax, "ASV_ITS_tax.${val_dada_ref_taxonomy}.tsv", taxlevels ) ch_versions = ch_versions.mix(DADA2_TAXONOMY.out.versions) FORMAT_TAXRESULTS_STD ( DADA2_TAXONOMY.out.tsv, ch_fasta, 'ASV_tax.tsv' ) ch_versions = ch_versions.mix( FORMAT_TAXRESULTS_STD.out.versions.ifEmpty(null) ) @@ -497,7 +497,7 @@ workflow AMPLISEQ { } // Sintax - // This will only run if --sintax_ref_taxonomy is defined, + // This will only run if --sintax_ref_taxonomy is defined, // i.e. if the channel ch_sintax_ref_taxonomy is not empty if (!params.skip_taxonomy) { FORMAT_TAXONOMY_SINTAX ( ch_sintax_ref_taxonomy.collect() ) From 5da307d10987701ddaa1a1e673e5142be76be1ae Mon Sep 17 00:00:00 2001 From: lokeshbio Date: Fri, 31 Mar 2023 17:48:29 +0200 Subject: [PATCH 043/158] tested coi filtering --- bin/filt_coi_stats.r | 2 +- modules/local/filter_coi_asv.nf | 2 +- workflows/ampliseq.nf | 4 +++- 3 files changed, 5 insertions(+), 3 deletions(-) mode change 100644 => 100755 bin/filt_coi_stats.r diff --git a/bin/filt_coi_stats.r b/bin/filt_coi_stats.r old mode 100644 new mode 100755 index df876ce5..3c4ef204 --- a/bin/filt_coi_stats.r +++ b/bin/filt_coi_stats.r @@ -9,7 +9,7 @@ if(length(args) < 2){ dada2_stats <- args[1] coi_filt <- args[2] - <- read.table(dada2_stats, header = T, check.names = F, row.names = 1) +old_stats <- read.table(dada2_stats, header = T, check.names = F, row.names = 1) filt_table <- read.table(coi_filt, header = T, check.names = F, row.names = 1) diff --git a/modules/local/filter_coi_asv.nf b/modules/local/filter_coi_asv.nf index a4d418c8..cb98c1a1 100644 --- a/modules/local/filter_coi_asv.nf +++ b/modules/local/filter_coi_asv.nf @@ -14,7 +14,7 @@ process FILTER_COI_ASV { path( "ASV_filtered.table.tsv" ) , emit: asv path( "ASV_filtered.fna" ) , emit: fasta path( "ASV_filtered.list" ) , emit: list - path( "stats.filt.tsv" ), emit: stas + path( "stats.filt.tsv" ), emit: stats path "versions.yml" , emit: versions when: diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 17616d1d..fe162e30 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -132,7 +132,7 @@ include { FILTER_SSU } from '../modules/local/filter_ssu' include { FILTER_LEN_ASV } from '../modules/local/filter_len_asv' include { MERGE_STATS as MERGE_STATS_FILTERSSU } from '../modules/local/merge_stats' include { MERGE_STATS as MERGE_STATS_FILTERLENASV } from '../modules/local/merge_stats' -include { FILTER_COI_ASV } from '../modules/local/filter_coi_asv' +include { FILTER_COI_ASV } from '../modules/local/filter_coi_asv' include { FORMAT_FASTAINPUT } from '../modules/local/format_fastainput' include { FORMAT_TAXONOMY } from '../modules/local/format_taxonomy' include { ITSX_CUTASV } from '../modules/local/itsx_cutasv' @@ -303,6 +303,8 @@ workflow AMPLISEQ { ch_stats = DADA2_MERGE.out.dada2stats } + + // // Modules : Filter rRNA // From 3fbdaf8df1db437243c230f1e1846d35946a9063 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Mon, 3 Apr 2023 14:27:05 +0200 Subject: [PATCH 044/158] Add test profile for sintax --- conf/test_sintax.config | 51 +++++++++++++++++++++++++++++++++++++++++ nextflow.config | 1 + 2 files changed, 52 insertions(+) create mode 100644 conf/test_sintax.config diff --git a/conf/test_sintax.config b/conf/test_sintax.config new file mode 100644 index 00000000..f1d705ec --- /dev/null +++ b/conf/test_sintax.config @@ -0,0 +1,51 @@ +/* +~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ + Nextflow config file for running minimal tests +~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ + Defines input files and everything required to run a fast and simple pipeline test. + + Use as follows: + nextflow run nf-core/ampliseq -profile test_sintax, --outdir + +---------------------------------------------------------------------------------------- +*/ + +params { + config_profile_name = 'Test sintax profile' + config_profile_description = 'Minimal test dataset to check pipeline function when using sintax for taxonomy assignment' + + // Limit resources so that this can run on GitHub Actions + max_cpus = 2 + max_memory = '6.GB' + max_time = '6.h' + + // Input data + FW_primer = "CTTGGTCATTTAGAGGAAGTAA" + RV_primer = "TCCTGAGGGAAACTTCG" + input = "https://raw.githubusercontent.com/nf-core/test-datasets/ampliseq/samplesheets/Samplesheet_pacbio_ITS.tsv" + metadata = "https://raw.githubusercontent.com/nf-core/test-datasets/ampliseq/samplesheets/Metadata_pacbio_ITS.tsv" + pacbio = true + max_ee = 12 + cut_its = "its2" + + dada_ref_taxonomy = "unite-fungi" + sintax_ref_taxonomy = "unite-fungi" + + //this is to remove low abundance ASVs to reduce runtime of downstream processes + min_samples = 2 + min_frequency = 10 + + //produce average barplots + metadata_category_barplot = "treatment1,badpairwise10" + + //restrict ANCOM analysis to higher taxonomic levels + dada_tax_agglom_max = 4 + qiime_tax_agglom_max = 4 + sintax_tax_agglom_max = 4 + + sbdiexport = true + + qiime_adonis_formula = "treatment1,mix8" + + diversity_rarefaction_depth = 500 +} diff --git a/nextflow.config b/nextflow.config index 221785ea..663c54e9 100644 --- a/nextflow.config +++ b/nextflow.config @@ -228,6 +228,7 @@ profiles { test_reftaxcustom { includeConfig 'conf/test_reftaxcustom.config' } test_novaseq { includeConfig 'conf/test_novaseq.config' } test_pplace { includeConfig 'conf/test_pplace.config' } + test_sintax { includeConfig 'conf/test_sintax.config' } } From fbee8c35eb9480401be076e5a2e77a99b9c34f5d Mon Sep 17 00:00:00 2001 From: lokeshbio Date: Mon, 3 Apr 2023 15:55:54 +0200 Subject: [PATCH 045/158] output.md is fixed --- docs/output.md | 27 +++++++++++++++++++++++++++ 1 file changed, 27 insertions(+) diff --git a/docs/output.md b/docs/output.md index 42d4eeec..918c351d 100644 --- a/docs/output.md +++ b/docs/output.md @@ -20,6 +20,7 @@ The pipeline is built using [Nextflow](https://www.nextflow.io/) and processes d - [Barrnap](#barrnap) - Predict ribosomal RNA sequences and optional filtering - [Length filter](#length-filter) - Optionally, ASV can be filtered by length thresholds - [ITSx](#itsx) - Optionally, the ITS region can be extracted + - [COI](#coi) - Optionally the COI ASVs can be filtered. - [Taxonomic classification with DADA2](#taxonomic-classification-with-dada2) - Taxonomic classification of (filtered) ASVs - [assignSH](#assignsh) - Optionally, a UNITE species hypothesis (SH) can be added to the taxonomy - [QIIME2](#qiime2) - Secondary analysis @@ -186,6 +187,32 @@ Optionally, the ITS region can be extracted from each ASV sequence using ITSx, a
+#### COI + +Optionally, the COI ASVs can be filtered based on their length and also their codons. The filtering is based on length of ASVs (418 ± nx3 and between 403 and 418), possibly does not contain any stop codon in the right reading frame. This applied to the samples that use any of the combination of following primers: + + forward: + - "CCHGAYATRGCHTTYCCHCG" + - "ACCHGAYATRGCHTTYCCHCG" + - "GACCHGAYATRGCHTTYCCHCG" + - "TGACCHGAYATRGCHTTYCCHCG" + reverse: + - "CDGGRTGNCCRAARAAYCA" + - "TCDGGRTGNCCRAARAAYCA" + - "ATCDGGRTGNCCRAARAAYCA" + - "GATCDGGRTGNCCRAARAAYCA" + +
+Output files + +- `COI_filtered/` + - `ASV_filtered.fna`: Fasta file of ASV sequences that passes the filter thresholds explained above. + - `ASV_filtered.table.tsv`: The count table of ASVs that successfully passed through the filter thresholds. + - `ASV_filtered.list`: List of ASV IDs that pass through the filter thresholds. + - `stats.filt.tsv`: The statistics from this filtering step is appeneded to the `DADA2_stats.tsv` + +
+ ### Taxonomic classification with DADA2 DADA2 taxonomically classifies the ASVs using a choice of supplied databases (specified with `--dada_ref_taxonomy`). The taxonomic classification will be done based on filtered ASV sequences (see above). From ba1e9ebb4be039dd45dcff9ef3fb5b8277ca7a05 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Wed, 5 Apr 2023 08:50:49 +0200 Subject: [PATCH 046/158] Fix typo --- bin/convert_sintax_output.py | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/bin/convert_sintax_output.py b/bin/convert_sintax_output.py index c7c0d2d1..8c3fa693 100755 --- a/bin/convert_sintax_output.py +++ b/bin/convert_sintax_output.py @@ -84,9 +84,9 @@ def complete_list(some_list, target_len): # Print header to outfile if args.taxlevels != "": - header = ["ASV"] + args.taxlevels.split(",") + ["sequence"] + header = ["ASV_ID"] + args.taxlevels.split(",") + ["sequence"] else: - header = ["ASV", "sequence"] + header = ["ASV_ID", "sequence"] num_taxa = len(header) - 2 print("\t".join(header), file=args.outfile) From fd05b12917120965a5a83b571f39e36d32e8aceb Mon Sep 17 00:00:00 2001 From: jtangrot Date: Wed, 5 Apr 2023 09:06:02 +0200 Subject: [PATCH 047/158] Fix test config --- conf/test_sintax.config | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/conf/test_sintax.config b/conf/test_sintax.config index f1d705ec..d93d0434 100644 --- a/conf/test_sintax.config +++ b/conf/test_sintax.config @@ -36,7 +36,7 @@ params { min_frequency = 10 //produce average barplots - metadata_category_barplot = "treatment1,badpairwise10" + metadata_category_barplot = "var2,var3" //restrict ANCOM analysis to higher taxonomic levels dada_tax_agglom_max = 4 From 16bed62b0d095b8b25441d2bd98325eaf569e065 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Wed, 5 Apr 2023 09:36:07 +0200 Subject: [PATCH 048/158] Fix test config --- conf/test_sintax.config | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/conf/test_sintax.config b/conf/test_sintax.config index d93d0434..e80066fc 100644 --- a/conf/test_sintax.config +++ b/conf/test_sintax.config @@ -45,7 +45,7 @@ params { sbdiexport = true - qiime_adonis_formula = "treatment1,mix8" + qiime_adonis_formula = "var2" diversity_rarefaction_depth = 500 } From c66a36a6ced2c03b6985301415d743a133edd2c3 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Wed, 5 Apr 2023 10:19:05 +0200 Subject: [PATCH 049/158] Remove unused file --- bin/create_unite_taxfile.py | 51 ------------------------------------- 1 file changed, 51 deletions(-) delete mode 100755 bin/create_unite_taxfile.py diff --git a/bin/create_unite_taxfile.py b/bin/create_unite_taxfile.py deleted file mode 100755 index ba2c85e9..00000000 --- a/bin/create_unite_taxfile.py +++ /dev/null @@ -1,51 +0,0 @@ -#!/usr/bin/env python3 -# -# Convert fasta file from UNITE into -# (i) fasta file with only sequence names and sequences -# (ii) Taxonomy file with 7 levels D_0__ to D_6__ -# -# By Jeanette Tångrot 2020-09-02 - -# --- Import libraries, do initializations ---# -import sys -from Bio import SeqIO - -usage = """create_unite_taxfile.py - : Input. Fasta file from UNITE containing taxonomies in description line - : Output. Name of fasta file. - : Output. Name of text file with taxonomies. -""" - -# --- Check and read arguments ---# -if len(sys.argv) != 4: - exit("Usage: " + usage) - -fasta_in = sys.argv[1] -fasta_out = sys.argv[2] -tax_out = sys.argv[3] - -# --- Read sequence file and create new records ---# -replace_dict = { - ";p__": ";D_1__", - ";c__": ";D_2__", - ";o__": ";D_3__", - ";f__": ";D_4__", - ";g__": ";D_5__", - ";s__": ";D_6__", -} - -fh_fasta = open(fasta_out, mode="w") -fh_tax = open(tax_out, mode="w") - -for entry in SeqIO.parse(fasta_in, "fasta"): - (name, tax) = entry.id.split("|k__") - tax = "D_0__" + tax - tax = tax.replace("unidentified", "") - for n1, n2 in replace_dict.items(): - tax = tax.replace(n1, n2) - tax = tax.replace("|SH", "_SH") - fh_fasta.write(">" + name + "\n" + str(entry.seq).upper() + "\n") - fh_tax.write(name + "\t" + tax + "\n") - -fh_fasta.close() -fh_tax.close() From 1133ea6df6e6afe4e526369b9aa1c8a43e37d25c Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Jeanette=20T=C3=A5ngrot?= Date: Wed, 5 Apr 2023 10:38:40 +0200 Subject: [PATCH 050/158] Make header in overall_summary consistent The input/DADA2_input column is named differently when using single end data compared to paired end, I suggest to use "DADA2_input" in both cases. I cannot find any conflicts in the ampliseq code. --- modules/local/dada2_stats.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/modules/local/dada2_stats.nf b/modules/local/dada2_stats.nf index 05ac3737..81e0f4da 100644 --- a/modules/local/dada2_stats.nf +++ b/modules/local/dada2_stats.nf @@ -85,7 +85,7 @@ process DADA2_STATS { } else { track <- cbind(filter_and_trim, sapply(dadaFs, getN), rowSums(seqtab.nochim)) } - colnames(track) <- c("input", "filtered", "denoised", "nonchim") + colnames(track) <- c("DADA2_input", "filtered", "denoised", "nonchim") track <- cbind(sample = sub(pattern = "(.*?)\\\\..*\$", replacement = "\\\\1", rownames(track)), track) write.table( track, file = "${meta.run}.stats.tsv", sep = "\\t", row.names = FALSE, quote = FALSE, na = '') From e33f83201fc64d0f2db1620a372c0ba37ef975e7 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Jeanette=20T=C3=A5ngrot?= Date: Wed, 5 Apr 2023 11:13:48 +0200 Subject: [PATCH 051/158] Update CHANGELOG.md --- CHANGELOG.md | 1 + 1 file changed, 1 insertion(+) diff --git a/CHANGELOG.md b/CHANGELOG.md index 35123649..1ccace81 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -17,6 +17,7 @@ and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0 - [#553](https://github.com/nf-core/ampliseq/pull/553) - Handle empty barrnap results files - [#554](https://github.com/nf-core/ampliseq/pull/554) - Accept taxonomy strings that contain `#`,`'` +- [#569](https://github.com/nf-core/ampliseq/pull/569) - Make header of overall_summary.tsv consistent between input data types ### `Dependencies` From 32532f8c71e0cecb2a1e32e858c191228e416e9a Mon Sep 17 00:00:00 2001 From: daniel Date: Wed, 5 Apr 2023 11:35:35 +0200 Subject: [PATCH 052/158] fix test.config --- conf/test.config | 1 - 1 file changed, 1 deletion(-) diff --git a/conf/test.config b/conf/test.config index 77deea96..e46c93a8 100644 --- a/conf/test.config +++ b/conf/test.config @@ -39,7 +39,6 @@ params { //restrict ANCOM analysis to higher taxonomic levels tax_agglom_max = 4 - tax_agglom_max = 4 sbdiexport = true From c0b6f844e3d1ac68bcbb4f422435e334404afcaf Mon Sep 17 00:00:00 2001 From: Daniel Straub <42973691+d4straub@users.noreply.github.com> Date: Wed, 5 Apr 2023 11:36:31 +0200 Subject: [PATCH 053/158] Fix typo Co-authored-by: Till E. <64961761+tillenglert@users.noreply.github.com> --- docs/output.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/docs/output.md b/docs/output.md index 28e5abc6..155e13f8 100644 --- a/docs/output.md +++ b/docs/output.md @@ -242,7 +242,7 @@ Phylogenetic placement grafts sequences onto a phylogenetic reference tree and o - `pplace/` - `*.graft.*.epa_result.newick`: Full phylogeny with query sequences grafted on to the reference phylogeny, in newick format. - `*.taxonomy.per_query.tsv`: Tab separated file with taxonomy information per query from classification by `gappa examine examinassign` - - `*.per_query_unique.tsv`: Tab separated file with taxonomy information as aboe, but one row per query, by filtering for lowest LWR (likelihood weight ratio) + - `*.per_query_unique.tsv`: Tab separated file with taxonomy information as above, but one row per query, by filtering for lowest LWR (likelihood weight ratio) - `*.heattree.tree.svg`: Heattree in SVG format from calling `gappa examine heattree`, see [Gappa documentation](https://github.com/Pbdas/epa-ng/blob/master/README.md) for details. - `pplace/hmmer/`: Contains intermediatary files if HMMER is used - `pplace/mafft/`: Contains intermediatary files if MAFFT is used From 5ce1c7e3386cfda7676e479765d4bbfd95c71f8d Mon Sep 17 00:00:00 2001 From: daniel Date: Wed, 5 Apr 2023 12:31:23 +0200 Subject: [PATCH 054/158] simplyfy tax_agglom_mi/ax --- workflows/ampliseq.nf | 10 ++++------ 1 file changed, 4 insertions(+), 6 deletions(-) diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 7bbb8429..c52aa646 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -91,6 +91,10 @@ if ( !is_fasta_input && (!params.FW_primer || !params.RV_primer) && !params.skip System.exit(1) } +// save params to values to be able to overwrite it +tax_agglom_min = params.tax_agglom_min +tax_agglom_max = params.tax_agglom_max + //use custom taxlevels from --dada_assign_taxlevels or database specific taxlevels if specified in conf/ref_databases.config if ( params.dada_ref_taxonomy ) { taxlevels = params.dada_assign_taxlevels ? "${params.dada_assign_taxlevels}" : @@ -544,18 +548,12 @@ workflow AMPLISEQ { } else if ( params.pplace_tree && params.pplace_taxonomy) { log.info "Use EPA-NG / GAPPA taxonomy classification" ch_tax = QIIME2_INTAX ( ch_pplace_tax ).qza - tax_agglom_min = params.tax_agglom_min - tax_agglom_max = params.tax_agglom_max } else if ( params.dada_ref_taxonomy ) { log.info "Use DADA2 taxonomy classification" ch_tax = QIIME2_INTAX ( ch_dada2_tax ).qza - tax_agglom_min = params.tax_agglom_min - tax_agglom_max = params.tax_agglom_max } else if ( params.qiime_ref_taxonomy || params.classifier ) { log.info "Use QIIME2 taxonomy classification" ch_tax = QIIME2_TAXONOMY.out.qza - tax_agglom_min = params.tax_agglom_min - tax_agglom_max = params.tax_agglom_max } else { log.info "Use no taxonomy classification" ch_tax = Channel.empty() From 572a423f3799f4cddf61a11d7afc02176dd70aec Mon Sep 17 00:00:00 2001 From: lokeshbio Date: Thu, 6 Apr 2023 10:58:56 +0200 Subject: [PATCH 055/158] added info to config and schema --- docs/output.md | 2 ++ nextflow.config | 1 + nextflow_schema.json | 5 +++++ 3 files changed, 8 insertions(+) diff --git a/docs/output.md b/docs/output.md index 918c351d..319991cf 100644 --- a/docs/output.md +++ b/docs/output.md @@ -202,6 +202,8 @@ Optionally, the COI ASVs can be filtered based on their length and also their co - "ATCDGGRTGNCCRAARAAYCA" - "GATCDGGRTGNCCRAARAAYCA" +This particular COI filtering can be set by using the option `filter_coi_asv`. +
Output files diff --git a/nextflow.config b/nextflow.config index 5f4ae444..bb643e4e 100644 --- a/nextflow.config +++ b/nextflow.config @@ -58,6 +58,7 @@ params { filter_ssu = null min_len_asv = null max_len_asv = null + filter_coi_asv = false diversity_rarefaction_depth = 500 ancom_sample_min_count = 1 diff --git a/nextflow_schema.json b/nextflow_schema.json index 4a343309..304e9012 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -335,6 +335,11 @@ "description": "Maximum ASV length", "help_text": "Remove ASV that are above the maximum length threshold (default: filter is disabled, otherwise 1000000). Lowering the threshold might reduce false positive ASVs (e.g. PCR off-targets)." }, + "filter_coi_asv": { + "type": "boolean", + "description": "Filter COI ASVs", + "help_text": "This setting is to be used only when any of the combination of following primers have been used \n\nforward:\n - \"CCHGAYATRGCHTTYCCHCG\"\n - \"ACCHGAYATRGCHTTYCCHCG\"\n - \"GACCHGAYATRGCHTTYCCHCG\"\n - \"TGACCHGAYATRGCHTTYCCHCG\"\n\nreverse:\n - \"CDGGRTGNCCRAARAAYCA\"\n - \"TCDGGRTGNCCRAARAAYCA\"\n - \"ATCDGGRTGNCCRAARAAYCA\"\n - \"GATCDGGRTGNCCRAARAAYCA\"\nASVs that do not follow the length category: 418 ± nx3 and between 403 and 418 and the ones that contain stop codon are filtered." + }, "exclude_taxa": { "type": "string", "default": "mitochondria,chloroplast", From c690195266c1fdb4f3788889d1247512ad8898db Mon Sep 17 00:00:00 2001 From: jtangrot Date: Thu, 6 Apr 2023 11:24:39 +0200 Subject: [PATCH 056/158] Update help text --- nextflow_schema.json | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/nextflow_schema.json b/nextflow_schema.json index 7f5f0bd8..33a62b8f 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -16,7 +16,7 @@ "mimetype": "text/tsv", "fa_icon": "fas fa-dna", "description": "Either a tab-separated sample sheet, a fasta file, or a folder containing zipped FastQ files", - "help_text": "Points to the main pipeline input, one of the following:\n- folder containing compressed fastq files\n- sample sheet ending with `.tsv` that points towards compressed fastq files\n- fasta file ending with `.fasta`, `.fna` or `.fa` that will be taxonomically classified\n\nRelated parameters are:\n- `--pacbio` and `--iontorrent` if the sequencing data is PacBio data or IonTorrent data (default expected: paired-end Illumina data)\n- `--single_end` if the sequencing data is single-ended Illumina data (default expected: paired-end Illumina data)\n- `--multiple_sequencing_runs` (folder input only) if the sequencing data originates from multiple sequencing runs\n- `--extension` (folder input only) if the sequencing file names do not follow the default (`\"/*_R{1,2}_001.fastq.gz\"`)\n- `--dada_ref_taxonomy` and `--qiime_ref_taxonomy` to choose an appropriate reference taxonomy for the type of amplicon (16S/18S/ITS/CO1) (default: 16S rRNA sequence database)\n\n##### Folder containing zipped FastQ files\n\nFor example:\n\n```bash\n--input 'path/to/data'\n```\n\nExample for input data organization from one sequencing run with two samples, paired-end data:\n\n```bash\ndata\n \u251c\u2500sample1_1_L001_R1_001.fastq.gz\n \u251c\u2500sample1_1_L001_R2_001.fastq.gz\n \u251c\u2500sample2_1_L001_R1_001.fastq.gz\n \u2514\u2500sample2_1_L001_R2_001.fastq.gz\n```\n\nPlease note the following requirements:\n\n1. The path must be enclosed in quotes\n2. The folder must contain gzip compressed demultiplexed fastq files. If the file names do not follow the default (`\"/*_R{1,2}_001.fastq.gz\"`), please check `--extension`.\n3. Sample identifiers are extracted from file names, i.e. the string before the first underscore `_`, these must be unique\n4. If your data is scattered, produce a sample sheet\n5. All sequencing data should originate from one sequencing run, because processing relies on run-specific error models that are unreliable when data from several sequencing runs are mixed. Sequencing data originating from multiple sequencing runs requires additionally the parameter `--multiple_sequencing_runs` and a specific folder structure.\n\n##### Sample sheet\n\nThe sample sheet file is an alternative way to provide input reads, it must be a tab-separated file ending with `.tsv` that must have two to four columns with the following headers: \n- `sampleID` (required): Unique sample identifiers, any unique string (may not contain dots `.`, must not start with a number)\n- `forwardReads` (required): Paths to (forward) reads zipped FastQ files\n- `reverseReads` (optional): Paths to reverse reads zipped FastQ files, required if the data is paired-end\n- `run` (optional): If the data was produced by multiple sequencing runs, any string\n\nFor example:\n\n```bash\n--input 'path/to/samplesheet.tsv'\n```\n\n##### Fasta file\n\nWhen pointing at a file ending with `.fasta`, `.fna` or `.fa`, the containing sequences will be taxonomically classified. All other pipeline steps will be skipped.\n\nThe sequence header line may contain a description, that will be kept as part of the sequence name. However, tabs will be changed into spaces.\n\nThe fasta file input option can be used to taxonomically classify previously produced ASV/OTU sequences.\n\nFor example:\n\n```bash\n--input 'path/to/amplicon_sequences.fasta'\n```" + "help_text": "Points to the main pipeline input, one of the following:\n- folder containing compressed fastq files\n- sample sheet ending with `.tsv` that points towards compressed fastq files\n- fasta file ending with `.fasta`, `.fna` or `.fa` that will be taxonomically classified\n\nRelated parameters are:\n- `--pacbio` and `--iontorrent` if the sequencing data is PacBio data or IonTorrent data (default expected: paired-end Illumina data)\n- `--single_end` if the sequencing data is single-ended Illumina data (default expected: paired-end Illumina data)\n- `--multiple_sequencing_runs` (folder input only) if the sequencing data originates from multiple sequencing runs\n- `--extension` (folder input only) if the sequencing file names do not follow the default (`\"/*_R{1,2}_001.fastq.gz\"`)\n- `--dada_ref_taxonomy`, `--qiime_ref_taxonomy`, and/or `--sintax_ref_taxonomy` to choose an appropriate reference taxonomy for the type of amplicon (16S/18S/ITS/CO1). If the `--sintax_ref_taxonomy` is given, taxonomic assignment is performed using the USEARCH sintax method in addition to DADA2 assignTaxonomy (default: DADA2 assignTaxonomy and 16S rRNA sequence database)\n\n##### Folder containing zipped FastQ files\n\nFor example:\n\n```bash\n--input 'path/to/data'\n```\n\nExample for input data organization from one sequencing run with two samples, paired-end data:\n\n```bash\ndata\n \u251c\u2500sample1_1_L001_R1_001.fastq.gz\n \u251c\u2500sample1_1_L001_R2_001.fastq.gz\n \u251c\u2500sample2_1_L001_R1_001.fastq.gz\n \u2514\u2500sample2_1_L001_R2_001.fastq.gz\n```\n\nPlease note the following requirements:\n\n1. The path must be enclosed in quotes\n2. The folder must contain gzip compressed demultiplexed fastq files. If the file names do not follow the default (`\"/*_R{1,2}_001.fastq.gz\"`), please check `--extension`.\n3. Sample identifiers are extracted from file names, i.e. the string before the first underscore `_`, these must be unique\n4. If your data is scattered, produce a sample sheet\n5. All sequencing data should originate from one sequencing run, because processing relies on run-specific error models that are unreliable when data from several sequencing runs are mixed. Sequencing data originating from multiple sequencing runs requires additionally the parameter `--multiple_sequencing_runs` and a specific folder structure.\n\n##### Sample sheet\n\nThe sample sheet file is an alternative way to provide input reads, it must be a tab-separated file ending with `.tsv` that must have two to four columns with the following headers: \n- `sampleID` (required): Unique sample identifiers, any unique string (may not contain dots `.`, must not start with a number)\n- `forwardReads` (required): Paths to (forward) reads zipped FastQ files\n- `reverseReads` (optional): Paths to reverse reads zipped FastQ files, required if the data is paired-end\n- `run` (optional): If the data was produced by multiple sequencing runs, any string\n\nFor example:\n\n```bash\n--input 'path/to/samplesheet.tsv'\n```\n\n##### Fasta file\n\nWhen pointing at a file ending with `.fasta`, `.fna` or `.fa`, the containing sequences will be taxonomically classified. All other pipeline steps will be skipped.\n\nThe sequence header line may contain a description, that will be kept as part of the sequence name. However, tabs will be changed into spaces.\n\nThe fasta file input option can be used to taxonomically classify previously produced ASV/OTU sequences.\n\nFor example:\n\n```bash\n--input 'path/to/amplicon_sequences.fasta'\n```" }, "FW_primer": { "type": "string", @@ -309,7 +309,7 @@ }, "sintax_ref_taxonomy": { "type": "string", - "help_text": "Choose any of the supported databases, and optionally also specify the version. Database and version are separated by an equal sign (`=`, e.g. `coidb=221216`) . This will download the desired database and initiate taxonomic classification with USEARCH sintax and the chosen database, which if needed is formatted it to produce a file that is compatible with USEARCH sintax.\n\nThe following databases are supported:\n- UNITE - eukaryotic nuclear ribosomal ITS region - ITS\n- COIDB - eukaryotic Cytochrome Oxidase I (COI) from The Barcode of Life Data System (BOLD) - COI\n\nGenerally, using `coidb`, `unite-fungi`, or `unite-alleuk` will select the most recent supported version. For details on what values are valid, please either use an invalid value such as `x` (causing the pipeline to send an error message with a list of all valid values) or see `conf/ref_databases.config`.", + "help_text": "Choose any of the supported databases, and optionally also specify the version. Database and version are separated by an equal sign (`=`, e.g. `coidb=221216`) . This will download the desired database and initiate taxonomic classification with USEARCH sintax and the chosen database, which if needed is formatted to produce a file that is compatible with USEARCH sintax.\n\nThe following databases are supported:\n- COIDB - eukaryotic Cytochrome Oxidase I (COI) from The Barcode of Life Data System (BOLD) - COI\n- UNITE - eukaryotic nuclear ribosomal ITS region - ITS\n\nGenerally, using `coidb`, `unite-fungi`, or `unite-alleuk` will select the most recent supported version. For details on what values are valid, please either use an invalid value such as `x` (causing the pipeline to send an error message with a list of all valid values) or see `conf/ref_databases.config`.", "description": "Name of supported database, and optionally also version number", "enum": [ "coidb", From 8764715b44a93db7b30d81374862778db11337e6 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Wed, 19 Apr 2023 11:31:49 +0200 Subject: [PATCH 057/158] Use sintax results in sbdiexport --- bin/convert_sintax_output.py | 48 ++++++++++++++++++----- bin/sbdiexport.R | 7 ++-- bin/sbdiexportreannotate.R | 14 +++---- conf/ref_databases.config | 2 + modules/local/format_taxresults_sintax.nf | 2 +- modules/local/sbdiexportreannotate.nf | 9 ++++- workflows/ampliseq.nf | 16 ++++++-- 7 files changed, 71 insertions(+), 27 deletions(-) diff --git a/bin/convert_sintax_output.py b/bin/convert_sintax_output.py index 8c3fa693..44557245 100755 --- a/bin/convert_sintax_output.py +++ b/bin/convert_sintax_output.py @@ -9,7 +9,7 @@ # # List of subroutines: # -# Usage: convert_sintax_output.py -i -f -o -t +# Usage: convert_sintax_output.py -i -f -o -t -d ################################## import re @@ -58,6 +58,15 @@ default="", ) +parser.add_argument( + "-d", + "--dbversion", + dest="db", + metavar="DBVERSION", + help="Taxonomy database name", + default="", +) + args = parser.parse_args() @@ -65,6 +74,11 @@ def complete_list(some_list, target_len): return some_list[:target_len] + ["NA"] * (target_len - len(some_list)) +# Find out if taxonomies contain SH groups +dbtype = "default" +if "UNITE" in args.db: + dbtype = "UNITE" + # Read fasta file and store as dictionary # If multiple sequences have the same name only the first will be stored seqs = dict() @@ -84,10 +98,16 @@ def complete_list(some_list, target_len): # Print header to outfile if args.taxlevels != "": - header = ["ASV_ID"] + args.taxlevels.split(",") + ["sequence"] + if dbtype == "UNITE": + header = ["ASV_ID"] + args.taxlevels.split(",") + ["SH", "confidence", "sequence"] + else: + header = ["ASV_ID"] + args.taxlevels.split(",") + ["confidence", "sequence"] else: - header = ["ASV_ID", "sequence"] -num_taxa = len(header) - 2 + header = ["ASV_ID", "confidence", "sequence"] +num_taxa = len(header) - 3 +num_cols = num_taxa +if dbtype == "UNITE": + num_taxa = num_taxa - 1 print("\t".join(header), file=args.outfile) # Read sintax file, parse results, and write taxonomies together with sequence to outfile @@ -97,14 +117,22 @@ def complete_list(some_list, target_len): sequence = seqs[tmp_list[0]] if tmp_list[0] in seqs else "" if tmp_list[3] != "": annot = re.sub(".\:", "", tmp_list[3]) - tmp_list1 = annot.split(",") - if len(tmp_list1) != num_taxa: - tmp_list1 = complete_list(tmp_list1, num_taxa) - print(tmp_list[0], "\t".join(tmp_list1), sequence, sep="\t", file=args.outfile) + annot_list = annot.split(",") + raw_annot = re.sub(".\:", "", tmp_list[1]) + raw_annot_list = raw_annot.split(",") + confidence = raw_annot_list[len(annot_list) - 1].split("(")[1] + confidence = confidence[:-1] + if len(annot_list) != num_taxa: + annot_list = complete_list(annot_list, num_cols) + print(tmp_list[0], "\t".join(annot_list), confidence, sequence, sep="\t", file=args.outfile) else: - print(tmp_list[0], "\t".join(tmp_list1), sequence, sep="\t", file=args.outfile) + if dbtype == "UNITE": + (species, sh) = annot_list[num_taxa - 1].rsplit("_", 1) + annot_list[num_taxa - 1] = species + annot_list.append(sh) + print(tmp_list[0], "\t".join(annot_list), confidence, sequence, sep="\t", file=args.outfile) else: - print(tmp_list[0], "\t".join(["NA"] * num_taxa), sequence, sep="\t", file=args.outfile) + print(tmp_list[0], "\t".join(["NA"] * num_cols), "", sequence, sep="\t", file=args.outfile) args.outfile.close() diff --git a/bin/sbdiexport.R b/bin/sbdiexport.R index 715871ac..e0a3575d 100755 --- a/bin/sbdiexport.R +++ b/bin/sbdiexport.R @@ -52,7 +52,7 @@ taxonomy <- read.delim(taxtable, sep = '\t', stringsAsFactors = FALSE) %>% mutate(Genus = if("Genus" %in% colnames(.)) Genus else '') %>% mutate(Species = if("Species" %in% colnames(.)) Species else '') %>% mutate(Species_exact = if("Species_exact" %in% colnames(.)) Species_exact else '') %>% - mutate(SH = if("SH" %in% colnames(.)) SH else '') %>% + mutate(otu = if("SH" %in% colnames(.)) SH else if ("BOLD_bin" %in% colnames(.)) BOLD_bin else '') %>% relocate(Domain, .after = sequence) %>% relocate(Kingdom, .after = Domain) %>% relocate(Phylum, .after = Kingdom) %>% @@ -62,7 +62,7 @@ taxonomy <- read.delim(taxtable, sep = '\t', stringsAsFactors = FALSE) %>% relocate(Genus, .after = Family) %>% relocate(Species, .after = Genus) %>% relocate(Species_exact, .after = Species) %>% - relocate(SH, .after = Species_exact) + relocate(otu, .after = Species_exact) # Read the metadata table if provided, otherwise create one @@ -144,7 +144,6 @@ asvtax <- asvs %>% mutate(across(domain:species, ~str_replace_all(.,' ','_'))) %>% rename( specificEpithet = species, - otu = SH, asv_id_alias = ASV_ID, DNA_sequence = sequence ) %>% @@ -161,5 +160,5 @@ asvtax <- asvs %>% ) %>% relocate(otu, .after = infraspecificEpithet) %>% relocate(associatedSequences, .before = domain) %>% - select(-confidence, -domain, -Species_exact) %>% + select_if(!names(.) %in% c('confidence','domain', 'Species_exact', 'SH', 'BOLD_bin')) %>% write_tsv("asv-table.tsv", na = '') diff --git a/bin/sbdiexportreannotate.R b/bin/sbdiexportreannotate.R index 761dda15..ccbc958a 100755 --- a/bin/sbdiexportreannotate.R +++ b/bin/sbdiexportreannotate.R @@ -16,8 +16,9 @@ suppressPackageStartupMessages(library(tidyverse)) args <- commandArgs(trailingOnly=TRUE) dbversion <- args[1] taxfile <- args[2] -wfversion <- args[3] -predfile <- args[4] +taxmethod <- args[3] +wfversion <- args[4] +predfile <- args[5] # Read taxonomy table taxonomy <- read.delim(taxfile, sep = '\t', stringsAsFactors = FALSE) @@ -54,7 +55,7 @@ taxtable <- taxonomy %>% mutate(Genus = if("Genus" %in% colnames(.)) Genus else '') %>% mutate(Species = if("Species" %in% colnames(.)) Species else '') %>% mutate(Species_exact = if("Species_exact" %in% colnames(.)) Species_exact else '') %>% - mutate(SH = if("SH" %in% colnames(.)) SH else '') %>% + mutate(otu = if("SH" %in% colnames(.)) SH else if ("BOLD_bin" %in% colnames(.)) BOLD_bin else '') %>% relocate(Domain, .after = sequence) %>% relocate(Kingdom, .after = Domain) %>% relocate(Phylum, .after = Kingdom) %>% @@ -64,13 +65,12 @@ taxtable <- taxonomy %>% relocate(Genus, .after = Family) %>% relocate(Species, .after = Genus) %>% relocate(Species_exact, .after = Species) %>% - relocate(SH, .after = Species_exact) %>% + relocate(otu, .after = Species_exact) %>% rename_with(tolower, Domain:Species_exact) %>% rename( asv_id_alias = ASV_ID, asv_sequence = sequence, specificEpithet = species, - otu = SH, annotation_confidence = confidence ) %>% mutate(across(.fns = ~str_replace_all(.,' ','_'))) %>% @@ -108,6 +108,7 @@ taxtable <- taxonomy %>% date_identified = as.character(lubridate::today()), reference_db = dbversion, annotation_algorithm = case_when( + (taxmethod == 'sintax') ~ paste('Ampliseq',wfversion,'(https://nf-co.re/ampliseq) VSEARCH:sintax', sep=' '), (!(is.na(otu) | otu == '')) ~ paste('Ampliseq',wfversion,'(https://nf-co.re/ampliseq) addsh', sep=' '), (!(is.na(species_exact) | species_exact == '')) ~ paste('Ampliseq',wfversion,'(https://nf-co.re/ampliseq) DADA2:assignTaxonomy:addSpecies', sep=' '), TRUE ~ paste('Ampliseq',wfversion,'(https://nf-co.re/ampliseq) DADA2:assignTaxonomy', sep=' ') @@ -121,6 +122,5 @@ taxtable <- taxonomy %>% relocate(infraspecificEpithet, .after = specificEpithet) %>% relocate(annotation_confidence, .after = otu) %>% relocate(date_identified:taxon_remarks, .after = annotation_confidence) %>% - select(-domain) %>% - select(-species_exact) %>% + select_if(!names(.) %in% c('domain', 'species_exact', 'SH', 'BOLD_bin')) %>% write_tsv("annotation.tsv", na = '') diff --git a/conf/ref_databases.config b/conf/ref_databases.config index cebc4462..9ffe2300 100644 --- a/conf/ref_databases.config +++ b/conf/ref_databases.config @@ -266,6 +266,7 @@ params { citation = "Sundh J, Manoharan L, Iwaszkiewicz-Eggebrecht E, Miraldo A, Andersson A, Ronquist F. COI reference sequences from BOLD DB. doi: https://doi.org/10.17044/scilifelab.20514192.v2" fmtscript = "taxref_reformat_sintax.sh" dbversion = "COIDB 221216 (https://doi.org/10.17044/scilifelab.20514192.v2)" + taxlevels = "Kingdom,Phylum,Class,Order,Family,Genus,Species,BOLD_bin" } 'coidb=221216' { title = "COIDB - CO1 Taxonomy Database - Release 221216" @@ -273,6 +274,7 @@ params { citation = "Sundh J, Manoharan L, Iwaszkiewicz-Eggebrecht E, Miraldo A, Andersson A, Ronquist F. COI reference sequences from BOLD DB. doi: https://doi.org/10.17044/scilifelab.20514192.v2" fmtscript = "taxref_reformat_sintax.sh" dbversion = "COIDB 221216 (https://doi.org/10.17044/scilifelab.20514192.v2)" + taxlevels = "Kingdom,Phylum,Class,Order,Family,Genus,Species,BOLD_bin" } 'unite-fungi' { title = "UNITE USEARCH/UTAX release for Fungi - Version 8.3" diff --git a/modules/local/format_taxresults_sintax.nf b/modules/local/format_taxresults_sintax.nf index 9af239f6..2496a7b3 100644 --- a/modules/local/format_taxresults_sintax.nf +++ b/modules/local/format_taxresults_sintax.nf @@ -22,7 +22,7 @@ process FORMAT_TAXRESULTS_SINTAX { script: def taxlevels = taxlevels_input ? taxlevels_input : "Kingdom,Phylum,Class,Order,Family,Genus,Species" """ - convert_sintax_output.py -i $taxtable -f $fastafile -o $outfile -t $taxlevels + convert_sintax_output.py -i $taxtable -f $fastafile -o $outfile -t $taxlevels -d \"${params.sintax_ref_databases[params.sintax_ref_taxonomy]["dbversion"]}\" cat <<-END_VERSIONS > versions.yml "${task.process}": diff --git a/modules/local/sbdiexportreannotate.nf b/modules/local/sbdiexportreannotate.nf index 8036fafc..7f1377ad 100644 --- a/modules/local/sbdiexportreannotate.nf +++ b/modules/local/sbdiexportreannotate.nf @@ -9,6 +9,7 @@ process SBDIEXPORTREANNOTATE { input: path taxonomytable + val taxonomymethod path predictions output: @@ -20,13 +21,19 @@ process SBDIEXPORTREANNOTATE { script: """ + if [[ $taxonomymethod == "sintax" ]] ; then + db_version="${params.sintax_ref_databases[params.sintax_ref_taxonomy]["dbversion"]}" + else + db_version="${params.dada_ref_databases[params.dada_ref_taxonomy]["dbversion"]}" + fi + if [[ $workflow.manifest.version == *dev ]]; then ampliseq_version="v$workflow.manifest.version, revision: ${workflow.scriptId.substring(0,10)}" else ampliseq_version="v$workflow.manifest.version" fi - sbdiexportreannotate.R \"${params.dada_ref_databases[params.dada_ref_taxonomy]["dbversion"]}\" $taxonomytable \"\$ampliseq_version\" $predictions + sbdiexportreannotate.R \"\$db_version\" $taxonomytable $taxonomymethod \"\$ampliseq_version\" $predictions cat <<-END_VERSIONS > versions.yml "${task.process}": diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 6b4b2800..beff2331 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -104,11 +104,14 @@ if ( params.dada_ref_taxonomy ) { taxlevels = params.dada_assign_taxlevels ? "${params.dada_assign_taxlevels}" : params.dada_ref_databases[params.dada_ref_taxonomy]["taxlevels"] ?: "" } else { taxlevels = params.dada_assign_taxlevels ? "${params.dada_assign_taxlevels}" : "" } +if ( params.sintax_ref_taxonomy ) { + sintax_taxlevels = params.sintax_ref_databases[params.sintax_ref_taxonomy]["taxlevels"] ?: "" +} //make sure that taxlevels adheres to requirements when mixed with addSpecies if ( params.dada_ref_taxonomy && !params.skip_dada_addspecies && !params.skip_taxonomy && taxlevels ) { if ( !taxlevels.endsWith(",Genus,Species") && !taxlevels.endsWith(",Genus") ) { - log.error "Incompatible settings: To use exact species annotations, taxonomic levels must end with `,Genus,Species` or `,Genus,Species` but are currently `${taxlevels}`. Taxonomic levels can be set with `--dada_assign_taxlevels`. Skip exact species annotations with `--skip_dada_addspecies`.\n" + log.error "Incompatible settings: To use exact species annotations, taxonomic levels must end with `,Genus,Species` or `,Genus` but are currently `${taxlevels}`. Taxonomic levels can be set with `--dada_assign_taxlevels`. Skip exact species annotations with `--skip_dada_addspecies`.\n" System.exit(1) } } @@ -535,7 +538,7 @@ workflow AMPLISEQ { } VSEARCH_SINTAX( ch_fasta_sintax, ch_sintaxdb ) ch_versions = ch_versions.mix(VSEARCH_SINTAX.out.versions) - FORMAT_TAXRESULTS_SINTAX( VSEARCH_SINTAX.out.tsv, ch_fasta, ASV_tax_name + '.tsv', taxlevels ) + FORMAT_TAXRESULTS_SINTAX( VSEARCH_SINTAX.out.tsv, ch_fasta, ASV_tax_name + '.tsv', sintax_taxlevels ) ch_sintax_tax = FORMAT_TAXRESULTS_SINTAX.out.tsv } @@ -712,9 +715,14 @@ workflow AMPLISEQ { // MODULE: Export data in SBDI's (Swedish biodiversity infrastructure) format // if ( params.sbdiexport ) { - SBDIEXPORT ( ch_dada2_asv, ch_dada2_tax, ch_metadata ) + if ( params.sintax_ref_taxonomy ) { + SBDIEXPORT ( ch_dada2_asv, ch_sintax_tax, ch_metadata ) + SBDIEXPORTREANNOTATE ( ch_sintax_tax, "sintax", ch_barrnapsummary.ifEmpty([]) ) + } else { + SBDIEXPORT ( ch_dada2_asv, ch_dada2_tax, ch_metadata ) + SBDIEXPORTREANNOTATE ( ch_dada2_tax, "dada2", ch_barrnapsummary.ifEmpty([]) ) + } ch_versions = ch_versions.mix(SBDIEXPORT.out.versions.first()) - SBDIEXPORTREANNOTATE ( ch_dada2_tax, ch_barrnapsummary.ifEmpty([]) ) } CUSTOM_DUMPSOFTWAREVERSIONS ( From 2d7482d8df47110a08df8552e00c3e9723643895 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Wed, 19 Apr 2023 12:44:11 +0200 Subject: [PATCH 058/158] Add parameter skip_dada_taxonomy --- conf/test_sintax.config | 18 +++++++++--------- nextflow.config | 1 + nextflow_schema.json | 4 ++++ workflows/ampliseq.nf | 12 ++++++------ 4 files changed, 20 insertions(+), 15 deletions(-) diff --git a/conf/test_sintax.config b/conf/test_sintax.config index e80066fc..b07b09b2 100644 --- a/conf/test_sintax.config +++ b/conf/test_sintax.config @@ -20,17 +20,17 @@ params { max_time = '6.h' // Input data - FW_primer = "CTTGGTCATTTAGAGGAAGTAA" - RV_primer = "TCCTGAGGGAAACTTCG" - input = "https://raw.githubusercontent.com/nf-core/test-datasets/ampliseq/samplesheets/Samplesheet_pacbio_ITS.tsv" - metadata = "https://raw.githubusercontent.com/nf-core/test-datasets/ampliseq/samplesheets/Metadata_pacbio_ITS.tsv" - pacbio = true - max_ee = 12 - cut_its = "its2" - - dada_ref_taxonomy = "unite-fungi" + input = "https://raw.githubusercontent.com/nf-core/test-datasets/ampliseq/samplesheets/Samplesheet_novaseq.tsv" + illumina_novaseq = true + metadata = "https://raw.githubusercontent.com/nf-core/test-datasets/ampliseq/samplesheets/Metadata_novaseq.tsv" + sintax_ref_taxonomy = "unite-fungi" + // Skip steps + skip_cutadapt = true + skip_barrnap = true + skip_dada_taxonomy = true + //this is to remove low abundance ASVs to reduce runtime of downstream processes min_samples = 2 min_frequency = 10 diff --git a/nextflow.config b/nextflow.config index 663c54e9..530e21b8 100644 --- a/nextflow.config +++ b/nextflow.config @@ -79,6 +79,7 @@ params { skip_abundance_tables = false skip_barplot = false skip_taxonomy = false + skip_dada_taxonomy = false skip_dada_addspecies = false skip_alpha_rarefaction = false skip_diversity_indices = false diff --git a/nextflow_schema.json b/nextflow_schema.json index 33a62b8f..29263275 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -515,6 +515,10 @@ "type": "boolean", "description": "Skip taxonomic classification. Incompatible with `--sbdiexport`" }, + "skip_dada_taxonomy": { + "type": "boolean", + "description": "Skip taxonomic classification with DADA2" + }, "skip_dada_addspecies": { "type": "boolean", "description": "Skip species level when using DADA2 for taxonomic classification. This reduces the required memory dramatically under certain conditions. Incompatible with `--sbdiexport`" diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index beff2331..20ed31bb 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -52,7 +52,7 @@ if (params.dada_ref_tax_custom) { } ch_dada_ref_taxonomy = Channel.empty() val_dada_ref_taxonomy = "user" -} else if (params.dada_ref_taxonomy && !params.skip_taxonomy) { +} else if (params.dada_ref_taxonomy && !params.skip_dada_taxonomy && !params.skip_taxonomy) { //standard ref taxonomy input from params.dada_ref_taxonomy & conf/ref_databases.config ch_dada_ref_taxonomy = Channel.fromList(params.dada_ref_databases[params.dada_ref_taxonomy]["file"]).map { file(it) } val_dada_ref_taxonomy = params.dada_ref_taxonomy.replace('=','_').replace('.','_') @@ -64,7 +64,7 @@ if (params.dada_ref_tax_custom) { val_dada_ref_taxonomy = "none" } -if (params.qiime_ref_taxonomy && !params.skip_taxonomy && !params.classifier) { +if (params.qiime_ref_taxonomy && !params.skip_dada_taxonomy && !params.skip_taxonomy && !params.classifier) { ch_qiime_ref_taxonomy = Channel.fromList(params.qiime_ref_databases[params.qiime_ref_taxonomy]["file"]).map { file(it) } } else { ch_qiime_ref_taxonomy = Channel.empty() } @@ -109,7 +109,7 @@ if ( params.sintax_ref_taxonomy ) { } //make sure that taxlevels adheres to requirements when mixed with addSpecies -if ( params.dada_ref_taxonomy && !params.skip_dada_addspecies && !params.skip_taxonomy && taxlevels ) { +if ( params.dada_ref_taxonomy && !params.skip_dada_addspecies && !params.skip_dada_taxonomy && !params.skip_taxonomy && taxlevels ) { if ( !taxlevels.endsWith(",Genus,Species") && !taxlevels.endsWith(",Genus") ) { log.error "Incompatible settings: To use exact species annotations, taxonomic levels must end with `,Genus,Species` or `,Genus` but are currently `${taxlevels}`. Taxonomic levels can be set with `--dada_assign_taxlevels`. Skip exact species annotations with `--skip_dada_addspecies`.\n" System.exit(1) @@ -117,7 +117,7 @@ if ( params.dada_ref_taxonomy && !params.skip_dada_addspecies && !params.skip_ta } //only run QIIME2 when taxonomy is actually calculated and all required data is available -if ( !(workflow.profile.tokenize(',').intersect(['conda', 'mamba']).size() >= 1) && !params.skip_taxonomy && !params.skip_qiime ) { +if ( !(workflow.profile.tokenize(',').intersect(['conda', 'mamba']).size() >= 1) && !params.skip_taxonomy && !params.skip_qiime && (!params.skip_dada_taxonomy || params.sintax_ref_taxonomy) ) { run_qiime2 = true } else { run_qiime2 = false @@ -381,7 +381,7 @@ workflow AMPLISEQ { ch_fasta = ch_dada2_fasta //DADA2 - if (!params.skip_taxonomy) { + if (!params.skip_taxonomy && !params.skip_dada_taxonomy) { if (!params.dada_ref_tax_custom) { //standard ref taxonomy input from conf/ref_databases.config FORMAT_TAXONOMY ( ch_dada_ref_taxonomy.collect(), val_dada_ref_taxonomy ) @@ -703,7 +703,7 @@ workflow AMPLISEQ { // MODULE: Predict functional potential of a bacterial community from marker genes with Picrust2 // if ( params.picrust ) { - if ( run_qiime2 && !params.skip_abundance_tables && ( params.dada_ref_taxonomy || params.qiime_ref_taxonomy || params.classifier ) && !params.skip_taxonomy ) { + if ( run_qiime2 && !params.skip_abundance_tables && ( params.dada_ref_taxonomy || params.qiime_ref_taxonomy || params.classifier || params.sintax_ref_taxonomy ) && !params.skip_taxonomy ) { PICRUST ( QIIME2_EXPORT.out.abs_fasta, QIIME2_EXPORT.out.abs_tsv, "QIIME2", "This Picrust2 analysis is based on filtered reads from QIIME2" ) } else { PICRUST ( ch_fasta, ch_dada2_asv, "DADA2", "This Picrust2 analysis is based on unfiltered reads from DADA2" ) From 5ae1f6b6508d49adb629f33b91f4c60f1144fade Mon Sep 17 00:00:00 2001 From: jtangrot Date: Wed, 19 Apr 2023 13:09:24 +0200 Subject: [PATCH 059/158] Fix input to qiime when no dada2 taxonomy --- subworkflows/local/qiime2_export.nf | 5 +++++ workflows/ampliseq.nf | 6 +++++- 2 files changed, 10 insertions(+), 1 deletion(-) diff --git a/subworkflows/local/qiime2_export.nf b/subworkflows/local/qiime2_export.nf index d7f57e0f..d67513e3 100644 --- a/subworkflows/local/qiime2_export.nf +++ b/subworkflows/local/qiime2_export.nf @@ -7,6 +7,7 @@ include { QIIME2_EXPORT_RELASV } from '../../modules/local/qiim include { QIIME2_EXPORT_RELTAX } from '../../modules/local/qiime2_export_reltax' include { COMBINE_TABLE as COMBINE_TABLE_QIIME2 } from '../../modules/local/combine_table' include { COMBINE_TABLE as COMBINE_TABLE_DADA2 } from '../../modules/local/combine_table' +include { COMBINE_TABLE as COMBINE_TABLE_SINTAX } from '../../modules/local/combine_table' workflow QIIME2_EXPORT { take: @@ -15,6 +16,7 @@ workflow QIIME2_EXPORT { ch_tax ch_QIIME2_tax_tsv ch_DADA2_tax_tsv + ch_SINTAX_tax_tsv tax_agglom_min tax_agglom_max @@ -34,6 +36,9 @@ workflow QIIME2_EXPORT { //combine_table.r (optional), similar to DADA2_table.tsv but with additionally taxonomy merged COMBINE_TABLE_DADA2 ( QIIME2_EXPORT_RELASV.out.tsv, QIIME2_EXPORT_ABSOLUTE.out.fasta, ch_DADA2_tax_tsv, 'rel-table-ASV_with-DADA2-tax.tsv' ) + //combine_table.r (optional), similar to DADA2_table.tsv but with additionally taxonomy merged + COMBINE_TABLE_SINTAX ( QIIME2_EXPORT_RELASV.out.tsv, QIIME2_EXPORT_ABSOLUTE.out.fasta, ch_SINTAX_tax_tsv, 'rel-table-ASV_with-DADA2-tax.tsv' ) + emit: abs_fasta = QIIME2_EXPORT_ABSOLUTE.out.fasta abs_tsv = QIIME2_EXPORT_ABSOLUTE.out.tsv diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 20ed31bb..de5824cc 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -497,6 +497,8 @@ workflow AMPLISEQ { } } } + } else { + ch_dada2_tax = Channel.empty() } // Sintax @@ -540,6 +542,8 @@ workflow AMPLISEQ { ch_versions = ch_versions.mix(VSEARCH_SINTAX.out.versions) FORMAT_TAXRESULTS_SINTAX( VSEARCH_SINTAX.out.tsv, ch_fasta, ASV_tax_name + '.tsv', sintax_taxlevels ) ch_sintax_tax = FORMAT_TAXRESULTS_SINTAX.out.tsv + } else { + ch_sintax_tax = Channel.empty() } // Phylo placement @@ -641,7 +645,7 @@ workflow AMPLISEQ { } //Export various ASV tables if (!params.skip_abundance_tables) { - QIIME2_EXPORT ( ch_asv, ch_seq, ch_tax, QIIME2_TAXONOMY.out.tsv, ch_dada2_tax, tax_agglom_min, tax_agglom_max ) + QIIME2_EXPORT ( ch_asv, ch_seq, ch_tax, QIIME2_TAXONOMY.out.tsv, ch_dada2_tax, ch_sintax_tax, tax_agglom_min, tax_agglom_max ) } if (!params.skip_barplot) { From 18036e514bfb71e48efb7cf52296370d15f5a9a4 Mon Sep 17 00:00:00 2001 From: daniel Date: Wed, 26 Apr 2023 15:35:11 +0200 Subject: [PATCH 060/158] fix single end empty file handling --- CHANGELOG.md | 1 + subworkflows/local/parse_input.nf | 4 ++-- 2 files changed, 3 insertions(+), 2 deletions(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index f6dcda98..1b17dbbb 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -19,6 +19,7 @@ and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0 - [#553](https://github.com/nf-core/ampliseq/pull/553) - Handle empty barrnap results files - [#554](https://github.com/nf-core/ampliseq/pull/554) - Accept taxonomy strings that contain `#`,`'` - [#569](https://github.com/nf-core/ampliseq/pull/569) - Make header of overall_summary.tsv consistent between input data types +- [#573](https://github.com/nf-core/ampliseq/pull/573) - Avoid parser error for single-end data when an empty read file is detected ### `Dependencies` diff --git a/subworkflows/local/parse_input.nf b/subworkflows/local/parse_input.nf index 3bed5705..6be99555 100644 --- a/subworkflows/local/parse_input.nf +++ b/subworkflows/local/parse_input.nf @@ -143,8 +143,8 @@ workflow PARSE_INPUT { //Filter empty files ch_reads .branch { - failed: it[0].single_end ? it[1][0].size() < 1.KB : it[1][0].size() < 1.KB || it[1][1].size() < 1.KB - passed: it[0].single_end ? it[1][0].size() >= 1.KB : it[1][0].size() >= 1.KB && it[1][1].size() >= 1.KB + failed: it[0].single_end ? it[1].size() < 1.KB : it[1][0].size() < 1.KB || it[1][1].size() < 1.KB + passed: it[0].single_end ? it[1].size() >= 1.KB : it[1][0].size() >= 1.KB && it[1][1].size() >= 1.KB } .set { ch_reads_result } ch_reads_result.passed.set { ch_reads_passed } From 778cc59dd8ed5c0ee97f2aec42ddb9b2c358f05b Mon Sep 17 00:00:00 2001 From: daniel Date: Thu, 27 Apr 2023 15:27:33 +0200 Subject: [PATCH 061/158] unify single end input channel from folder and samplesheet input --- subworkflows/local/parse_input.nf | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/subworkflows/local/parse_input.nf b/subworkflows/local/parse_input.nf index 6be99555..11cc5f3b 100644 --- a/subworkflows/local/parse_input.nf +++ b/subworkflows/local/parse_input.nf @@ -27,7 +27,7 @@ def parse_samplesheet(LinkedHashMap row, single_end) { exit 1, "ERROR: Please check input samplesheet -> Forward read FastQ file does not exist!\n${row.forwardReads}" } if (meta.single_end) { - array = [ meta, [ file(row.forwardReads) ] ] + array = [ meta, file(row.forwardReads) ] } else { if (!file(row.reverseReads).exists()) { exit 1, "ERROR: Please check input samplesheet -> Reverse read FastQ file does not exist!\n${row.reverseReads}" @@ -141,7 +141,7 @@ workflow PARSE_INPUT { .subscribe { if ( params.metadata && "$it"[0].isNumber() ) exit 1, "Please review data input, sampleIDs may not start with a number, but \"$it\" does. The pipeline unintentionally modifies such strings and the metadata will not match any more." } //Filter empty files - ch_reads + ch_reads.dump(tag:'parse_input.nf: ch_reads') .branch { failed: it[0].single_end ? it[1].size() < 1.KB : it[1][0].size() < 1.KB || it[1][1].size() < 1.KB passed: it[0].single_end ? it[1].size() >= 1.KB : it[1][0].size() >= 1.KB && it[1][1].size() >= 1.KB From 9eb271782ddbf2780540c20148a005cc48dfbdd0 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Fri, 28 Apr 2023 13:12:47 +0200 Subject: [PATCH 062/158] Revert test config to pacbio data --- conf/test_sintax.config | 17 +++++++++-------- 1 file changed, 9 insertions(+), 8 deletions(-) diff --git a/conf/test_sintax.config b/conf/test_sintax.config index b07b09b2..a1b506e8 100644 --- a/conf/test_sintax.config +++ b/conf/test_sintax.config @@ -20,15 +20,16 @@ params { max_time = '6.h' // Input data - input = "https://raw.githubusercontent.com/nf-core/test-datasets/ampliseq/samplesheets/Samplesheet_novaseq.tsv" - illumina_novaseq = true - metadata = "https://raw.githubusercontent.com/nf-core/test-datasets/ampliseq/samplesheets/Metadata_novaseq.tsv" + FW_primer = "CTTGGTCATTTAGAGGAAGTAA" + RV_primer = "TCCTGAGGGAAACTTCG" + input = "https://raw.githubusercontent.com/nf-core/test-datasets/ampliseq/samplesheets/Samplesheet_pacbio_ITS.tsv" + metadata = "https://raw.githubusercontent.com/nf-core/test-datasets/ampliseq/samplesheets/Metadata_pacbio_ITS.tsv" + pacbio = true + max_ee = 12 + cut_its = "its2" sintax_ref_taxonomy = "unite-fungi" - // Skip steps - skip_cutadapt = true - skip_barrnap = true skip_dada_taxonomy = true //this is to remove low abundance ASVs to reduce runtime of downstream processes @@ -36,7 +37,7 @@ params { min_frequency = 10 //produce average barplots - metadata_category_barplot = "var2,var3" + metadata_category_barplot = "treatment1,badpairwise10" //restrict ANCOM analysis to higher taxonomic levels dada_tax_agglom_max = 4 @@ -45,7 +46,7 @@ params { sbdiexport = true - qiime_adonis_formula = "var2" + qiime_adonis_formula = "treatment1,mix8" diversity_rarefaction_depth = 500 } From a50404bb60f55480cdf0ed9c7d6b97196b0db030 Mon Sep 17 00:00:00 2001 From: lokeshbio Date: Fri, 28 Apr 2023 13:21:47 +0200 Subject: [PATCH 063/158] filtering codons feature added --- bin/filt_codon_stats.r | 22 ++++++ bin/filt_codons.py | 128 +++++++++++++++++++++++++++++++++ conf/modules.config | 4 +- docs/output.md | 30 +++----- modules/local/filter_codons.nf | 54 ++++++++++++++ nextflow.config | 3 +- workflows/ampliseq.nf | 16 ++--- 7 files changed, 227 insertions(+), 30 deletions(-) create mode 100755 bin/filt_codon_stats.r create mode 100755 bin/filt_codons.py create mode 100644 modules/local/filter_codons.nf diff --git a/bin/filt_codon_stats.r b/bin/filt_codon_stats.r new file mode 100755 index 00000000..497338de --- /dev/null +++ b/bin/filt_codon_stats.r @@ -0,0 +1,22 @@ +#!/usr/bin/env Rscript + +args = commandArgs(trailingOnly=TRUE) + +if(length(args) < 2){ + stop("Usage: filt_codons_stats.r ") +} + +dada2_stats <- args[1] +coi_filt <- args[2] + +old_stats <- read.table(dada2_stats, header = T, check.names = F, row.names = 1) + +filt_table <- read.table(coi_filt, header = T, check.names = F, row.names = 1) + +filt_counts <- as.data.frame(colSums(filt_table)) +names(filt_counts) <- "codon_filtered" + +new_stats <- merge(old_stats, filt_counts, by = 'row.names') + +write.table(new_stats, file = "codon.filtered.stats.tsv", row.names=FALSE, sep="\t", col.names = TRUE, quote = FALSE, na = '') + diff --git a/bin/filt_codons.py b/bin/filt_codons.py new file mode 100755 index 00000000..ffe53018 --- /dev/null +++ b/bin/filt_codons.py @@ -0,0 +1,128 @@ +#!/usr/bin/env python3 +##### Program description ####### +# +# Title: Subsetting Insect Biome ASVs +# +# Author(s): Lokeshwaran Manoharan +# +# +# +# Description: The ASVs that are supposed to be of particular length (418 ± nx3 and between 403 and 418), possibly does not contain any stop codon in the right reading frame. +# +# List of subroutines: +# +# +# +# Overall procedure: using functions and dictionaries in python +# +# Usage: filt_InsBiom_asv.py +# +################################## + +import re +import sys +import copy +import argparse + +usage = '''This program filters ASVs that contains any stop codon or if the length from the position is not a multiple of 3 in the right reading frame.''' + +parser = argparse.ArgumentParser(description=usage) + +parser.add_argument( + '-f', '--fasta', + dest='fasta', + type=argparse.FileType('r'), + help='Fasta file of the ASVs from the AmpliSeq pipeline', + required=True + ) + +parser.add_argument( + '-t', '--count-table', + dest='count', + type=argparse.FileType('r'), + help='Count table file of the ASVs from the AmpliSeq pipeline', + required=True + ) + +parser.add_argument( + '-s', '--start-position', + dest='begin', + type=int, + help='Specific position of the ASV where to start checking for codons. By default it starts from position 1', + required=False, + default=0 + ) + +parser.add_argument( + '-e', '--end-position', + dest='end', + type=int, + help='Specific position of the ASV where to stop checking for codons. By default it checks until the end of the sequence', + required=False + ) + +parser.add_argument( + '-p', '--out-prefix', + dest='prefix', + type=str, + help='Prefix of the output files with filtered ASVs and the corresponding count table. The output files will be in *_filtered.* format', + required=True + ) + + +args = parser.parse_args() + +begin = args.begin + +p1 = re.compile('\t') +p2 = re.compile('>') + +def check_asv(seq, start, stop): + sub_seq = seq[start-1:stop] + stop_list = ['TAA', 'TAG'] + sub_list = [] + for x in range(0, len(sub_seq), 3): + sub_list.append(sub_seq[x:x+3]) + if not len(sub_seq)%3 == 0: + return False + elif any(x in sub_list for x in stop_list): + return False + else: + return True + +Out_Seq = open(args.prefix +'_filtered.fna', 'w') +Out_list = open(args.prefix+'_filtered.list', 'w') +Out_table = open(args.prefix+'_filtered.table.tsv', 'w') + +count_dict = {} + +count = 0 +for line in args.count: + line = line.rstrip('\n') + if count == 0: + print(line, file = Out_table) + count += 1 + else: + tmp_list = re.split(p1, line) + count_dict[tmp_list[0]] = line + +for line in args.fasta: + line = line.rstrip('\n') + if re.match(p2, line) is not None: + bin_head = re.sub(p2, '', line) + else: + if args.end is not None: + end = args.end + else: + end = len(line) + if check_asv(line, begin, end): + print('>', bin_head,'\n', line, file = Out_Seq, sep='') + print(bin_head, file = Out_list) + print(count_dict[bin_head], file = Out_table) + +args.count.close() +args.fasta.close() +Out_Seq.close() +Out_list.close() +Out_table.close() + diff --git a/conf/modules.config b/conf/modules.config index 853cdf3f..66aa5d3e 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -309,9 +309,9 @@ process { ] } - withName: FILTER_COI_ASV { + withName: FILTER_CODONS { publishDir = [ - path: { "${params.outdir}/COI_filtered" }, + path: { "${params.outdir}/Codons_filtered" }, mode: params.publish_dir_mode, saveAs: { filename -> filename.equals('versions.yml') ? null : filename } ] diff --git a/docs/output.md b/docs/output.md index 319991cf..3f1c2e46 100644 --- a/docs/output.md +++ b/docs/output.md @@ -20,7 +20,7 @@ The pipeline is built using [Nextflow](https://www.nextflow.io/) and processes d - [Barrnap](#barrnap) - Predict ribosomal RNA sequences and optional filtering - [Length filter](#length-filter) - Optionally, ASV can be filtered by length thresholds - [ITSx](#itsx) - Optionally, the ITS region can be extracted - - [COI](#coi) - Optionally the COI ASVs can be filtered. + - [Codons](#codons) - Optionally the ASVs can be filtered by presence of stop codons. - [Taxonomic classification with DADA2](#taxonomic-classification-with-dada2) - Taxonomic classification of (filtered) ASVs - [assignSH](#assignsh) - Optionally, a UNITE species hypothesis (SH) can be added to the taxonomy - [QIIME2](#qiime2) - Secondary analysis @@ -187,31 +187,23 @@ Optionally, the ITS region can be extracted from each ASV sequence using ITSx, a
-#### COI +#### Codons -Optionally, the COI ASVs can be filtered based on their length and also their codons. The filtering is based on length of ASVs (418 ± nx3 and between 403 and 418), possibly does not contain any stop codon in the right reading frame. This applied to the samples that use any of the combination of following primers: +Optionally, the ASVs can be filtered if they contain stop codons (`"TAA,TAG"`) in the specified open reading frame of the ASV. The filtering step also filters ASVs in the specifed checking range, if they are not multiple of 3. - forward: - - "CCHGAYATRGCHTTYCCHCG" - - "ACCHGAYATRGCHTTYCCHCG" - - "GACCHGAYATRGCHTTYCCHCG" - - "TGACCHGAYATRGCHTTYCCHCG" - reverse: - - "CDGGRTGNCCRAARAAYCA" - - "TCDGGRTGNCCRAARAAYCA" - - "ATCDGGRTGNCCRAARAAYCA" - - "GATCDGGRTGNCCRAARAAYCA" +This filtering can be done by the setting the `--orf_start` option to starting position on the ASV where you would like to start checking of codon usage! For example, setting this parameter to `1`, would start checking the ASV for codons from the beginning of the ASV or if it is set to `22`, the check starts the open reading frame from position 22 on each ASV. By default, the filtering applies from the `--orf_start` to the end of the ASV. But, this can be changed by setting the `--orf_end` to specify the end of the open reading frame where you want to look for stop codons + +> **NB:** just make sure that when you set `--orf_end`, the length between `--orf_start` and `--orf_end` is a multiple of 3, otherwise all ASVs would be filtered. -This particular COI filtering can be set by using the option `filter_coi_asv`.
Output files -- `COI_filtered/` - - `ASV_filtered.fna`: Fasta file of ASV sequences that passes the filter thresholds explained above. - - `ASV_filtered.table.tsv`: The count table of ASVs that successfully passed through the filter thresholds. - - `ASV_filtered.list`: List of ASV IDs that pass through the filter thresholds. - - `stats.filt.tsv`: The statistics from this filtering step is appeneded to the `DADA2_stats.tsv` +- `Codons_filtered/` + - `ASV_codon_filtered.fna`: Fasta file of ASV sequences that passes the filter thresholds explained above. + - `ASV_codon_filtered.table.tsv`: The count table of ASVs that successfully passed through the filter thresholds. + - `ASV_codon_filtered.list`: List of ASV IDs that pass through the filter thresholds. + - `codon.filtered.stats.tsv`: Tracking read numbers through filtering, for each sample.
diff --git a/modules/local/filter_codons.nf b/modules/local/filter_codons.nf new file mode 100644 index 00000000..10aa1b99 --- /dev/null +++ b/modules/local/filter_codons.nf @@ -0,0 +1,54 @@ +process FILTER_CODONS { + tag "${fasta}" + label 'process_low' + + conda (params.enable_conda ? { exit 1 "QIIME2 has no conda package" } : null) + container "quay.io/qiime2/core:2022.8" + + input: + path(fasta) + path(asv) + path(dada2stats) + + output: + path( "ASV_codon_filtered.table.tsv" ) , emit: asv + path( "ASV_codon_filtered.fna" ) , emit: fasta + path( "ASV_codon_filtered.list" ) , emit: list + path( "codon.filtered.stats.tsv" ), emit: stats + path "versions.yml" , emit: versions + + when: + task.ext.when == null || task.ext.when + + script: + if (!params.orf_end) { + def orf_start = params.orf_start ?: '1' + + """ + filt_codons.py -f ${fasta} -t ${asv} -s ${orf_start} -p ASV_codon + filt_codon_stats.r ${dada2stats} ASV_codon_filtered.table.tsv + + cat <<-END_VERSIONS > versions.yml + "${task.process}": + python: \$(python --version 2>&1 | sed 's/Python //g') + R: \$(R --version | sed -n 1p | sed 's/R version //g' | sed 's/\\s.*\$//') + END_VERSIONS + """ + + } else { + def orf_start = params.orf_start ?: '1' + def orf_end = params.orf_end ?: '1000000' + + """ + filt_codons.py -f ${fasta} -t ${asv} -s ${orf_start} -e ${orf_end} -p ASV_codon + filt_codon_stats.r ${dada2stats} ASV_codon_filtered.table.tsv + + cat <<-END_VERSIONS > versions.yml + "${task.process}": + python: \$(python --version 2>&1 | sed 's/Python //g') + R: \$(R --version | sed -n 1p | sed 's/R version //g' | sed 's/\\s.*\$//') + END_VERSIONS + """ + } + +} \ No newline at end of file diff --git a/nextflow.config b/nextflow.config index bb643e4e..3b408d99 100644 --- a/nextflow.config +++ b/nextflow.config @@ -58,7 +58,8 @@ params { filter_ssu = null min_len_asv = null max_len_asv = null - filter_coi_asv = false + orf_start = null + orf_end = null diversity_rarefaction_depth = 500 ancom_sample_min_count = 1 diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index fe162e30..74d35b50 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -132,7 +132,7 @@ include { FILTER_SSU } from '../modules/local/filter_ssu' include { FILTER_LEN_ASV } from '../modules/local/filter_len_asv' include { MERGE_STATS as MERGE_STATS_FILTERSSU } from '../modules/local/merge_stats' include { MERGE_STATS as MERGE_STATS_FILTERLENASV } from '../modules/local/merge_stats' -include { FILTER_COI_ASV } from '../modules/local/filter_coi_asv' +include { FILTER_CODONS } from '../modules/local/filter_codons' include { FORMAT_FASTAINPUT } from '../modules/local/format_fastainput' include { FORMAT_TAXONOMY } from '../modules/local/format_taxonomy' include { ITSX_CUTASV } from '../modules/local/itsx_cutasv' @@ -351,14 +351,14 @@ workflow AMPLISEQ { } // - // Modules : COI ASV Filtering + // Modules : Filtering based on codons in an open reading frame // - if (params.filter_coi_asv) { - FILTER_COI_ASV ( ch_dada2_fasta, ch_dada2_asv, ch_stats ) - ch_versions = ch_versions.mix(FILTER_COI_ASV.out.versions.ifEmpty(null)) - ch_stats = FILTER_COI_ASV.out.stats - ch_dada2_fasta = FILTER_COI_ASV.out.fasta - ch_dada2_asv = FILTER_COI_ASV.out.asv + if (params.orf_start || params.orf_end) { + FILTER_CODONS ( ch_dada2_fasta, ch_dada2_asv, ch_stats ) + ch_versions = ch_versions.mix(FILTER_CODONS.out.versions.ifEmpty(null)) + ch_stats = FILTER_CODONS.out.stats + ch_dada2_fasta = FILTER_CODONS.out.fasta + ch_dada2_asv = FILTER_CODONS.out.asv } // From 602590b514d0c37f7776c79973d97c51fe60a377 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Fri, 28 Apr 2023 13:27:05 +0200 Subject: [PATCH 064/158] Only run ITSx once --- workflows/ampliseq.nf | 199 +++++++++++++++++------------------------- 1 file changed, 82 insertions(+), 117 deletions(-) diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index de5824cc..4b2af77f 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -151,8 +151,7 @@ include { MERGE_STATS as MERGE_STATS_FILTERLENASV } from '../modules/local/merge include { FORMAT_FASTAINPUT } from '../modules/local/format_fastainput' include { FORMAT_TAXONOMY } from '../modules/local/format_taxonomy' include { FORMAT_TAXONOMY_SINTAX } from '../modules/local/format_taxonomy_sintax' -include { ITSX_CUTASV as ITSX_CUTASV_DADA2 } from '../modules/local/itsx_cutasv' -include { ITSX_CUTASV as ITSX_CUTASV_SINTAX } from '../modules/local/itsx_cutasv' +include { ITSX_CUTASV } from '../modules/local/itsx_cutasv' include { MERGE_STATS as MERGE_STATS_STD} from '../modules/local/merge_stats' include { DADA2_TAXONOMY } from '../modules/local/dada2_taxonomy' include { DADA2_ADDSPECIES } from '../modules/local/dada2_addspecies' @@ -375,10 +374,30 @@ workflow AMPLISEQ { ch_dada2_asv = FILTER_LEN_ASV.out.asv } + // + // Modules : ITSx - cut out ITS region if long ITS reads + // + ch_full_fasta = ch_dada2_fasta + if (params.cut_its == "none") { + ch_fasta = ch_dada2_fasta + } else { + if (params.cut_its == "full") { + outfile = params.its_partial ? "ASV_ITS_seqs.full_and_partial.fasta" : "ASV_ITS_seqs.full.fasta" + } + else if (params.cut_its == "its1") { + outfile = params.its_partial ? "ASV_ITS_seqs.ITS1.full_and_partial.fasta" : "ASV_ITS_seqs.ITS1.fasta" + } + else if (params.cut_its == "its2") { + outfile = params.its_partial ? "ASV_ITS_seqs.ITS2.full_and_partial.fasta" : "ASV_ITS_seqs.ITS2.fasta" + } + ITSX_CUTASV ( ch_full_fasta, outfile ) + ch_versions = ch_versions.mix(ITSX_CUTASV.out.versions.ifEmpty(null)) + ch_fasta = ITSX_CUTASV.out.fasta + } + // // SUBWORKFLOW / MODULES : Taxonomic classification with DADA2, SINTAX and/or QIIME2 // - ch_fasta = ch_dada2_fasta //DADA2 if (!params.skip_taxonomy && !params.skip_dada_taxonomy) { @@ -403,98 +422,61 @@ workflow AMPLISEQ { .set { ch_assigntax } } if (params.cut_its == "none") { - DADA2_TAXONOMY ( ch_fasta, ch_assigntax, "ASV_tax.${val_dada_ref_taxonomy}.tsv", taxlevels ) - ch_versions = ch_versions.mix(DADA2_TAXONOMY.out.versions) - if (!params.skip_dada_addspecies) { - DADA2_ADDSPECIES ( DADA2_TAXONOMY.out.rds, ch_addspecies, "ASV_tax_species.${val_dada_ref_taxonomy}.tsv", taxlevels ) - if ( params.addsh ) { - ch_fasta - .map { - fasta -> - def meta = [:] - meta.id = "ASV.vsearch" - [ meta, fasta ] } - .set { ch_fasta_map } - VSEARCH_USEARCHGLOBAL( ch_fasta_map, ch_assigntax, vsearch_cutoff, 'blast6out', "" ) - ch_versions = ch_versions.mix(VSEARCH_USEARCHGLOBAL.out.versions.ifEmpty(null)) - ASSIGNSH( DADA2_ADDSPECIES.out.tsv, ch_shinfo.collect(), VSEARCH_USEARCHGLOBAL.out.txt, "ASV_tax_species_SH.${val_dada_ref_taxonomy}.tsv") - ch_versions = ch_versions.mix(ASSIGNSH.out.versions.ifEmpty(null)) - ch_dada2_tax = ASSIGNSH.out.tsv - } else { - ch_dada2_tax = DADA2_ADDSPECIES.out.tsv - } - } else { - if ( params.addsh ) { - ch_fasta - .map { - fasta -> - def meta = [:] - meta.id = "ASV.vsearch" - [ meta, fasta ] } - .set { ch_fasta_map } - VSEARCH_USEARCHGLOBAL( ch_fasta_map, ch_assigntax, vsearch_cutoff, 'blast6out', "" ) - ch_versions = ch_versions.mix(VSEARCH_USEARCHGLOBAL.out.versions.ifEmpty(null)) - ASSIGNSH( DADA2_TAXONOMY.out.tsv, ch_shinfo.collect(), VSEARCH_USEARCHGLOBAL.out.txt, "ASV_tax_SH.${val_dada_ref_taxonomy}.tsv") - ch_versions = ch_versions.mix(ASSIGNSH.out.versions.ifEmpty(null)) - ch_dada2_tax = ASSIGNSH.out.tsv - } else { - ch_dada2_tax = DADA2_TAXONOMY.out.tsv - } - } - //Cut out ITS region if long ITS reads + ASV_tax_name = "ASV_tax" } else { - if (params.cut_its == "full") { - outfile = params.its_partial ? "ASV_ITS_seqs.full_and_partial.fasta" : "ASV_ITS_seqs.full.fasta" + ASV_tax_name = "ASV_ITS_tax" + } + DADA2_TAXONOMY ( ch_fasta, ch_assigntax, ASV_tax_name + ".${val_dada_ref_taxonomy}.tsv", taxlevels ) + ch_versions = ch_versions.mix(DADA2_TAXONOMY.out.versions) + if (params.cut_its != "none") { + FORMAT_TAXRESULTS_STD ( DADA2_TAXONOMY.out.tsv, ch_full_fasta, "ASV_tax.${val_dada_ref_taxonomy}.tsv" ) + ch_versions = ch_versions.mix( FORMAT_TAXRESULTS_STD.out.versions.ifEmpty(null) ) + } + if (!params.skip_dada_addspecies) { + DADA2_ADDSPECIES ( DADA2_TAXONOMY.out.rds, ch_addspecies, ASV_tax_name + "_species.${val_dada_ref_taxonomy}.tsv", taxlevels ) + if (params.cut_its == "none") { + ch_dada2_addsp = DADA2_ADDSPECIES.out.tsv + } else { + FORMAT_TAXRESULTS_ADDSP ( DADA2_ADDSPECIES.out.tsv, ch_fasta, "ASV_tax_species.${val_dada_ref_taxonomy}.tsv" ) + ch_dada2_addsp = FORMAT_TAXRESULTS_ADDSP.out.tsv } - else if (params.cut_its == "its1") { - outfile = params.its_partial ? "ASV_ITS_seqs.ITS1.full_and_partial.fasta" : "ASV_ITS_seqs.ITS1.fasta" + if ( params.addsh ) { + ch_fasta + .map { + fasta -> + def meta = [:] + meta.id = ASV_tax_name + ".vsearch" + [ meta, fasta ] } + .set { ch_fasta_map } + VSEARCH_USEARCHGLOBAL( ch_fasta_map, ch_assigntax, vsearch_cutoff, 'blast6out', "" ) + ch_versions = ch_versions.mix(VSEARCH_USEARCHGLOBAL.out.versions.ifEmpty(null)) + ASSIGNSH( ch_dada2_addsp, ch_shinfo.collect(), VSEARCH_USEARCHGLOBAL.out.txt, "ASV_tax_species_SH.${val_dada_ref_taxonomy}.tsv") + ch_versions = ch_versions.mix(ASSIGNSH.out.versions.ifEmpty(null)) + ch_dada2_tax = ASSIGNSH.out.tsv + } else { + ch_dada2_tax = ch_dada2_addsp } - else if (params.cut_its == "its2") { - outfile = params.its_partial ? "ASV_ITS_seqs.ITS2.full_and_partial.fasta" : "ASV_ITS_seqs.ITS2.fasta" + } else { + if (params.cut_its == "none") { + ch_dada2_tax1 = DADA2_TAXONOMY.out.tsv + } else { + ch_dada2_tax1 = FORMAT_TAXRESULTS_STD.out.tsv } - ITSX_CUTASV_DADA2 ( ch_fasta, outfile ) - ch_versions = ch_versions.mix(ITSX_CUTASV_DADA2.out.versions.ifEmpty(null)) - ch_cut_fasta = ITSX_CUTASV_DADA2.out.fasta - DADA2_TAXONOMY ( ch_cut_fasta, ch_assigntax, "ASV_ITS_tax.${val_dada_ref_taxonomy}.tsv", taxlevels ) - ch_versions = ch_versions.mix(DADA2_TAXONOMY.out.versions) - FORMAT_TAXRESULTS_STD ( DADA2_TAXONOMY.out.tsv, ch_fasta, 'ASV_tax.tsv' ) - ch_versions = ch_versions.mix( FORMAT_TAXRESULTS_STD.out.versions.ifEmpty(null) ) - if (!params.skip_dada_addspecies) { - DADA2_ADDSPECIES ( DADA2_TAXONOMY.out.rds, ch_addspecies, "ASV_ITS_tax_species.${val_dada_ref_taxonomy}.tsv", taxlevels ) - FORMAT_TAXRESULTS_ADDSP ( DADA2_ADDSPECIES.out.tsv, ch_fasta, "ASV_tax_species.${val_dada_ref_taxonomy}.tsv" ) - if ( params.addsh ) { - ch_cut_fasta - .map { - fasta -> - def meta = [:] - meta.id = "ASV_cut.vsearch" - [ meta, fasta ] } - .set { ch_cut_fasta } - VSEARCH_USEARCHGLOBAL( ch_cut_fasta, ch_assigntax, vsearch_cutoff, 'blast6out', "" ) - ch_versions = ch_versions.mix(VSEARCH_USEARCHGLOBAL.out.versions.ifEmpty(null)) - ASSIGNSH( FORMAT_TAXRESULTS_ADDSP.out.tsv, ch_shinfo.collect(), VSEARCH_USEARCHGLOBAL.out.txt, "ASV_tax_species_SH.${val_dada_ref_taxonomy}.tsv") - ch_versions = ch_versions.mix(ASSIGNSH.out.versions.ifEmpty(null)) - ch_dada2_tax = ASSIGNSH.out.tsv - } else { - ch_dada2_tax = FORMAT_TAXRESULTS_ADDSP.out.tsv - } + if ( params.addsh ) { + ch_fasta + .map { + fasta -> + def meta = [:] + meta.id = ASV_tax_name + ".vsearch" + [ meta, fasta ] } + .set { ch_fasta_map } + VSEARCH_USEARCHGLOBAL( ch_fasta_map, ch_assigntax, vsearch_cutoff, 'blast6out', "" ) + ch_versions = ch_versions.mix(VSEARCH_USEARCHGLOBAL.out.versions.ifEmpty(null)) + ASSIGNSH( ch_dada2_tax1.out.tsv, ch_shinfo.collect(), VSEARCH_USEARCHGLOBAL.out.txt, "ASV_tax_SH.${val_dada_ref_taxonomy}.tsv") + ch_versions = ch_versions.mix(ASSIGNSH.out.versions.ifEmpty(null)) + ch_dada2_tax = ASSIGNSH.out.tsv } else { - if ( params.addsh ) { - ch_cut_fasta - .map { - fasta -> - def meta = [:] - meta.id = "ASV_cut.vsearch" - [ meta, fasta ] } - .set { ch_cut_fasta } - VSEARCH_USEARCHGLOBAL( ch_cut_fasta, ch_assigntax, vsearch_cutoff, 'blast6out', "" ) - ch_versions = ch_versions.mix(VSEARCH_USEARCHGLOBAL.out.versions.ifEmpty(null)) - ASSIGNSH( FORMAT_TAXRESULTS_STD.out.tsv, ch_shinfo.collect(), VSEARCH_USEARCHGLOBAL.out.txt, "ASV_tax_SH.${val_dada_ref_taxonomy}.tsv") - ch_versions = ch_versions.mix(ASSIGNSH.out.versions.ifEmpty(null)) - ch_dada2_tax = ASSIGNSH.out.tsv - } else { - ch_dada2_tax = FORMAT_TAXRESULTS_STD.out.tsv - } + ch_dada2_tax = ch_dada2_tax1 } } } else { @@ -509,38 +491,21 @@ workflow AMPLISEQ { ch_sintaxdb = FORMAT_TAXONOMY_SINTAX.out.db if (params.cut_its == "none") { ASV_tax_name = "ASV_tax_sintax.${val_sintax_ref_taxonomy}" - ch_fasta - .map { - fasta -> - def meta = [:] - meta.id = ASV_tax_name + ".raw" - [ meta, fasta ] } - .set { ch_fasta_sintax } + ASV_tax_name2 = "ASV_tax_sintax.${val_sintax_ref_taxonomy}" } else { ASV_tax_name = "ASV_ITS_tax_sintax.${val_sintax_ref_taxonomy}" - if (params.cut_its == "full") { - outfile = params.its_partial ? "ASV_ITS_seqs.full_and_partial.fasta" : "ASV_ITS_seqs.full.fasta" - } - else if (params.cut_its == "its1") { - outfile = params.its_partial ? "ASV_ITS_seqs.ITS1.full_and_partial.fasta" : "ASV_ITS_seqs.ITS1.fasta" - } - else if (params.cut_its == "its2") { - outfile = params.its_partial ? "ASV_ITS_seqs.ITS2.full_and_partial.fasta" : "ASV_ITS_seqs.ITS2.fasta" - } - ITSX_CUTASV_SINTAX ( ch_fasta, outfile ) - ch_versions = ch_versions.mix(ITSX_CUTASV_SINTAX.out.versions.ifEmpty(null)) - ch_cut_fasta = ITSX_CUTASV_SINTAX.out.fasta - ch_cut_fasta - .map { - fasta -> - def meta = [:] - meta.id = ASV_tax_name + ".raw" - [ meta, fasta ] } - .set { ch_fasta_sintax } + ASV_tax_name2 = "ASV_tax_sintax.${val_sintax_ref_taxonomy}" } + ch_fasta + .map { + fasta -> + def meta = [:] + meta.id = ASV_tax_name + ".raw" + [ meta, fasta ] } + .set { ch_fasta_sintax } VSEARCH_SINTAX( ch_fasta_sintax, ch_sintaxdb ) ch_versions = ch_versions.mix(VSEARCH_SINTAX.out.versions) - FORMAT_TAXRESULTS_SINTAX( VSEARCH_SINTAX.out.tsv, ch_fasta, ASV_tax_name + '.tsv', sintax_taxlevels ) + FORMAT_TAXRESULTS_SINTAX( VSEARCH_SINTAX.out.tsv, ch_full_fasta, ASV_tax_name2 + '.tsv', sintax_taxlevels ) ch_sintax_tax = FORMAT_TAXRESULTS_SINTAX.out.tsv } else { ch_sintax_tax = Channel.empty() From 4d7fa386488d6a9ff4b05120f475533dfd739ce9 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Fri, 28 Apr 2023 15:31:59 +0200 Subject: [PATCH 065/158] Add docs and citation --- .github/workflows/ci.yml | 3 ++- CITATIONS.md | 6 +++++- README.md | 2 +- docs/output.md | 40 +++++++++++++++++++++++++++++++++++----- 4 files changed, 43 insertions(+), 8 deletions(-) diff --git a/.github/workflows/ci.yml b/.github/workflows/ci.yml index 8c235eda..140f892b 100644 --- a/.github/workflows/ci.yml +++ b/.github/workflows/ci.yml @@ -58,7 +58,8 @@ jobs: test_reftaxcustom, test_novaseq, test_pplace, - ] + test_sintax, + ] steps: - name: Check out pipeline code uses: actions/checkout@v2 diff --git a/CITATIONS.md b/CITATIONS.md index 997b12b2..777446bd 100644 --- a/CITATIONS.md +++ b/CITATIONS.md @@ -133,7 +133,11 @@ - VSEARCH option usearch_global implements the [USEARCH](https://doi.org/10.1093/bioinformatics/btq461) algorithm - > R.C. Edgar. Search and clustering orders of magnitude faster than BLAST. Bioinformatics. 2010 26(19) 2460-2461 + > Edgar RC. Search and clustering orders of magnitude faster than BLAST. Bioinformatics. 2010 26(19) 2460-2461 + +- VSEARCH option sintax implements the [SINTAX](https://doi.org/10.1101/074161) algorithm + + > Edgar RC. (2016) SINTAX: a simple non-Bayesian taxonomy classifier for 16S and ITS sequences, BioRxiv, 074161. Preprint. ### Summarizing software diff --git a/README.md b/README.md index 778643c1..bcf0722e 100644 --- a/README.md +++ b/README.md @@ -38,7 +38,7 @@ By default, the pipeline currently performs the following: - Infer Amplicon Sequence Variants (ASVs) ([DADA2](https://doi.org/10.1038/nmeth.3869)) - Predict whether ASVs are ribosomal RNA sequences ([Barrnap](https://github.com/tseemann/barrnap)) - Phylogenetic placement ([EPA-NG](https://github.com/Pbdas/epa-ng)) -- Taxonomical classification using DADA2 or [QIIME2](https://www.nature.com/articles/s41587-019-0209-9) +- Taxonomical classification using DADA2, [SINTAX](https://doi.org/10.1101/074161) or [QIIME2](https://www.nature.com/articles/s41587-019-0209-9) - Excludes unwanted taxa, produces absolute and relative feature/taxa count tables and plots, plots alpha rarefaction curves, computes alpha and beta diversity indices and plots thereof ([QIIME2](https://www.nature.com/articles/s41587-019-0209-9)) - Calls differentially abundant taxa ([ANCOM](https://www.ncbi.nlm.nih.gov/pubmed/26028277)) - Overall pipeline run summaries ([MultiQC](https://multiqc.info/)) diff --git a/docs/output.md b/docs/output.md index 155e13f8..b80549ad 100644 --- a/docs/output.md +++ b/docs/output.md @@ -187,11 +187,13 @@ Optionally, the ITS region can be extracted from each ASV sequence using ITSx, a -### Taxonomic classification with DADA2 +### Taxonomic classification -DADA2 taxonomically classifies the ASVs using a choice of supplied databases (specified with `--dada_ref_taxonomy`). The taxonomic classification will be done based on filtered ASV sequences (see above). +DADA2 and/or SINTAX can be used to taxonomically classify the ASVs using a choice of supplied databases (specified with `--dada_ref_taxonomy` and/or `sintax_ref_taxonomy`). By default, DADA2 is used for the classification. The taxonomic classification will be done based on filtered ASV sequences (see above). -Depending on the reference taxonomy database, sequences can be classified down to species rank. Species classification is reported in columns "Species" using DADA2's assignTaxonomy function or "Species_exact" using DADA2's addSpecies function, the latter only assigns exact sequence matches. Generally, species assignment without exact matches are much less trustworthy than those with exact matches. With short amplicons, e.g. 16S rRNA gene V4 region, the non-exact species annotation is not recommended to be trusted. The longer the ASVs are, the more acceptable is the non-exact species classification, e.g. PacBio (nearly) full length 16S rRNA gene sequences are thought to be trustworthy. +#### DADA2 + +DADA2 is the default method for taxonomy classification. Depending on the reference taxonomy database, sequences can be classified down to species rank. Species classification is reported in columns "Species" using DADA2's assignTaxonomy function or "Species_exact" using DADA2's addSpecies function, the latter only assigns exact sequence matches. Generally, species assignment without exact matches are much less trustworthy than those with exact matches. With short amplicons, e.g. 16S rRNA gene V4 region, the non-exact species annotation is not recommended to be trusted. The longer the ASVs are, the more acceptable is the non-exact species classification, e.g. PacBio (nearly) full length 16S rRNA gene sequences are thought to be trustworthy. Files when _not_ using ITSx (default): @@ -221,7 +223,7 @@ Files when using ITSx: #### assignSH -Optionally, a UNITE species hypothesis (SH) can be added to the taxonomy. In short, the sequences are matched to the selected UNITE database with VSEARCH, using the sequence identity cutoff 98.5%. If a good enough and unique match is found, the sequence is assigned the SH of the matching sequence, and the taxonomy assignment for the sequence is changed to that of the SH. If no match is found, the taxonomy generated by assignTaxonomy and/or assignSpecies is kept. +Optionally, a UNITE species hypothesis (SH) can be added to the DADA2 taxonomy. In short, the sequences are matched to the selected UNITE database with VSEARCH, using the sequence identity cutoff 98.5%. If a good enough and unique match is found, the sequence is assigned the SH of the matching sequence, and the taxonomy assignment for the sequence is changed to that of the SH. If no match is found, the taxonomy generated by assignTaxonomy and/or assignSpecies is kept.
Output files @@ -232,7 +234,35 @@ Optionally, a UNITE species hypothesis (SH) can be added to the taxonomy. In sho
-### Phlogenetic placement and taxonomic classification +#### SINTAX + +Alternatively, ASVs can be taxonomically classified using the SINTAX algorithm as implemented in VSEARCH. Depending on the reference taxonomy database (specified with `sintax_ref_taxonomy`), sequences can be classified down to species rank. + +Files when _not_ using ITSx (default): + +
+Output files + +- `sintax/` + - `ASV_tax_sintax.*.raw.tsv`: Raw output from SINTAX. + - `ASV_tax_sintax.*.tsv`: Taxonomic classification for each ASV sequence in a format similar to DADA2 output. + - `ref_taxonomy.*.txt`: Information about the used reference taxonomy, such as title, version, citation. + +
+ +Files when using ITSx: + +
+Output files + +- `sintax/` + - `ASV_ITS_tax_sintax.*.raw.tsv`: Raw output from SINTAX, when using cut sequences as input. + - `ASV_tax_sintax.*.tsv`: Taxonomic classification for each ASV sequence, based on the chosen ITS region, in a format similar to DADA2 output. + - `ref_taxonomy.*.txt`: Information about the used reference taxonomy, such as title, version, citation. + +
+ +### Phylogenetic placement and taxonomic classification Phylogenetic placement grafts sequences onto a phylogenetic reference tree and optionally outputs taxonomic annotations. The reference tree is ideally made from full-length high-quality sequences containing better evolutionary signal than short amplicons. It is hence superior to estimating de-novo phylogenetic trees from short amplicon sequences. On providing required reference files, ASV sequences are aligned to the reference alignment with either [HMMER](http://hmmer.org/) (default) or [MAFFT](https://mafft.cbrc.jp/alignment/software/). Subsequently, phylogenetic placement of query sequences is performed with [EPA-NG](https://github.com/Pbdas/epa-ng), and finally a number of summary operations are performed with [Gappa](https://github.com/lczech/gappa). This uses code from [nf-core/phyloplace](https://nf-co.re/phyloplace) in the form of its main [subworkflow](https://github.com/nf-core/modules/tree/master/subworkflows/nf-core/fasta_newick_epang_gappa), therefore its detailed documentation also applies here. From 25605bd418890f0b02bf6bad69ac1b6f5292c307 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Fri, 28 Apr 2023 15:37:50 +0200 Subject: [PATCH 066/158] Prettier --- .github/workflows/ci.yml | 2 +- CITATIONS.md | 4 ++-- docs/output.md | 2 +- 3 files changed, 4 insertions(+), 4 deletions(-) diff --git a/.github/workflows/ci.yml b/.github/workflows/ci.yml index 140f892b..07d7ab5b 100644 --- a/.github/workflows/ci.yml +++ b/.github/workflows/ci.yml @@ -59,7 +59,7 @@ jobs: test_novaseq, test_pplace, test_sintax, - ] + ] steps: - name: Check out pipeline code uses: actions/checkout@v2 diff --git a/CITATIONS.md b/CITATIONS.md index 777446bd..e488e7bd 100644 --- a/CITATIONS.md +++ b/CITATIONS.md @@ -134,10 +134,10 @@ - VSEARCH option usearch_global implements the [USEARCH](https://doi.org/10.1093/bioinformatics/btq461) algorithm > Edgar RC. Search and clustering orders of magnitude faster than BLAST. Bioinformatics. 2010 26(19) 2460-2461 - + - VSEARCH option sintax implements the [SINTAX](https://doi.org/10.1101/074161) algorithm - > Edgar RC. (2016) SINTAX: a simple non-Bayesian taxonomy classifier for 16S and ITS sequences, BioRxiv, 074161. Preprint. + > Edgar RC. (2016) SINTAX: a simple non-Bayesian taxonomy classifier for 16S and ITS sequences, BioRxiv, 074161. Preprint. ### Summarizing software diff --git a/docs/output.md b/docs/output.md index b80549ad..77f8bbd2 100644 --- a/docs/output.md +++ b/docs/output.md @@ -237,7 +237,7 @@ Optionally, a UNITE species hypothesis (SH) can be added to the DADA2 taxonomy. #### SINTAX Alternatively, ASVs can be taxonomically classified using the SINTAX algorithm as implemented in VSEARCH. Depending on the reference taxonomy database (specified with `sintax_ref_taxonomy`), sequences can be classified down to species rank. - + Files when _not_ using ITSx (default):
From 02503677ade3818552f6433b349500f501a6e00f Mon Sep 17 00:00:00 2001 From: nf-core-bot Date: Fri, 28 Apr 2023 14:17:36 +0000 Subject: [PATCH 067/158] Template update for nf-core/tools version 2.8 --- .editorconfig | 2 +- .github/ISSUE_TEMPLATE/bug_report.yml | 2 +- .github/PULL_REQUEST_TEMPLATE.md | 3 +- .github/workflows/awsfulltest.yml | 2 +- .github/workflows/awstest.yml | 2 +- .github/workflows/branch.yml | 2 +- .github/workflows/clean-up.yml | 24 ++++ .github/workflows/linting.yml | 2 +- .pre-commit-config.yaml | 5 + CHANGELOG.md | 2 +- README.md | 74 ++++++---- bin/check_samplesheet.py | 3 - conf/base.config | 2 +- conf/igenomes.config | 8 ++ conf/test_full.config | 2 + docs/usage.md | 130 +++++------------- lib/NfcoreSchema.groovy | 4 +- lib/WorkflowAmpliseq.groovy | 12 +- lib/WorkflowMain.groovy | 13 +- main.nf | 1 - modules.json | 4 +- modules/local/samplesheet_check.nf | 2 +- .../custom/dumpsoftwareversions/main.nf | 6 +- .../custom/dumpsoftwareversions/meta.yml | 2 + modules/nf-core/multiqc/main.nf | 6 +- modules/nf-core/multiqc/meta.yml | 3 +- nextflow.config | 31 ++++- tower.yml | 5 + 28 files changed, 195 insertions(+), 159 deletions(-) create mode 100644 .github/workflows/clean-up.yml create mode 100644 .pre-commit-config.yaml create mode 100644 tower.yml diff --git a/.editorconfig b/.editorconfig index b78de6e6..b6b31907 100644 --- a/.editorconfig +++ b/.editorconfig @@ -8,7 +8,7 @@ trim_trailing_whitespace = true indent_size = 4 indent_style = space -[*.{md,yml,yaml,html,css,scss,js,cff}] +[*.{md,yml,yaml,html,css,scss,js}] indent_size = 2 # These files are edited and tested upstream in nf-core/modules diff --git a/.github/ISSUE_TEMPLATE/bug_report.yml b/.github/ISSUE_TEMPLATE/bug_report.yml index b783e190..79bc7c1a 100644 --- a/.github/ISSUE_TEMPLATE/bug_report.yml +++ b/.github/ISSUE_TEMPLATE/bug_report.yml @@ -45,6 +45,6 @@ body: * Nextflow version _(eg. 22.10.1)_ * Hardware _(eg. HPC, Desktop, Cloud)_ * Executor _(eg. slurm, local, awsbatch)_ - * Container engine: _(e.g. Docker, Singularity, Conda, Podman, Shifter or Charliecloud)_ + * Container engine: _(e.g. Docker, Singularity, Conda, Podman, Shifter, Charliecloud, or Apptainer)_ * OS _(eg. CentOS Linux, macOS, Linux Mint)_ * Version of nf-core/ampliseq _(eg. 1.1, 1.5, 1.8.2)_ diff --git a/.github/PULL_REQUEST_TEMPLATE.md b/.github/PULL_REQUEST_TEMPLATE.md index ced09960..636e0714 100644 --- a/.github/PULL_REQUEST_TEMPLATE.md +++ b/.github/PULL_REQUEST_TEMPLATE.md @@ -15,7 +15,8 @@ Learn more about contributing: [CONTRIBUTING.md](https://github.com/nf-core/ampl - [ ] This comment contains a description of changes (with reason). - [ ] If you've fixed a bug or added code that should be tested, add tests! -- [ ] If you've added a new tool - have you followed the pipeline conventions in the [contribution docs](https://github.com/nf-core/ampliseq/tree/master/.github/CONTRIBUTING.md)- [ ] If necessary, also make a PR on the nf-core/ampliseq _branch_ on the [nf-core/test-datasets](https://github.com/nf-core/test-datasets) repository. +- [ ] If you've added a new tool - have you followed the pipeline conventions in the [contribution docs](https://github.com/nf-core/ampliseq/tree/master/.github/CONTRIBUTING.md) +- [ ] If necessary, also make a PR on the nf-core/ampliseq _branch_ on the [nf-core/test-datasets](https://github.com/nf-core/test-datasets) repository. - [ ] Make sure your code lints (`nf-core lint`). - [ ] Ensure the test suite passes (`nextflow run . -profile test,docker --outdir `). - [ ] Usage Documentation in `docs/usage.md` is updated. diff --git a/.github/workflows/awsfulltest.yml b/.github/workflows/awsfulltest.yml index f0efcb80..d66d066a 100644 --- a/.github/workflows/awsfulltest.yml +++ b/.github/workflows/awsfulltest.yml @@ -14,7 +14,7 @@ jobs: runs-on: ubuntu-latest steps: - name: Launch workflow via tower - uses: nf-core/tower-action@v3 + uses: seqeralabs/action-tower-launch@v1 # TODO nf-core: You can customise AWS full pipeline tests as required # Add full size test data (but still relatively small datasets for few samples) # on the `test_full.config` test runs with only one set of parameters diff --git a/.github/workflows/awstest.yml b/.github/workflows/awstest.yml index b6f5626a..d42ecdfd 100644 --- a/.github/workflows/awstest.yml +++ b/.github/workflows/awstest.yml @@ -12,7 +12,7 @@ jobs: steps: # Launch workflow using Tower CLI tool action - name: Launch workflow via tower - uses: nf-core/tower-action@v3 + uses: seqeralabs/action-tower-launch@v1 with: workspace_id: ${{ secrets.TOWER_WORKSPACE_ID }} access_token: ${{ secrets.TOWER_ACCESS_TOKEN }} diff --git a/.github/workflows/branch.yml b/.github/workflows/branch.yml index ffb5a222..2a9f5e0a 100644 --- a/.github/workflows/branch.yml +++ b/.github/workflows/branch.yml @@ -13,7 +13,7 @@ jobs: - name: Check PRs if: github.repository == 'nf-core/ampliseq' run: | - { [[ ${{github.event.pull_request.head.repo.full_name }} == nf-core/ampliseq ]] && [[ $GITHUB_HEAD_REF = "dev" ]]; } || [[ $GITHUB_HEAD_REF == "patch" ]] + { [[ ${{github.event.pull_request.head.repo.full_name }} == nf-core/ampliseq ]] && [[ $GITHUB_HEAD_REF == "dev" ]]; } || [[ $GITHUB_HEAD_REF == "patch" ]] # If the above check failed, post a comment on the PR explaining the failure # NOTE - this doesn't currently work if the PR is coming from a fork, due to limitations in GitHub actions secrets diff --git a/.github/workflows/clean-up.yml b/.github/workflows/clean-up.yml new file mode 100644 index 00000000..694e90ec --- /dev/null +++ b/.github/workflows/clean-up.yml @@ -0,0 +1,24 @@ +name: "Close user-tagged issues and PRs" +on: + schedule: + - cron: "0 0 * * 0" # Once a week + +jobs: + clean-up: + runs-on: ubuntu-latest + permissions: + issues: write + pull-requests: write + steps: + - uses: actions/stale@v7 + with: + stale-issue-message: "This issue has been tagged as awaiting-changes or awaiting-feedback by an nf-core contributor. Remove stale label or add a comment otherwise this issue will be closed in 20 days." + stale-pr-message: "This PR has been tagged as awaiting-changes or awaiting-feedback by an nf-core contributor. Remove stale label or add a comment if it is still useful." + close-issue-message: "This issue was closed because it has been tagged as awaiting-changes or awaiting-feedback by an nf-core contributor and then staled for 20 days with no activity." + days-before-stale: 30 + days-before-close: 20 + days-before-pr-close: -1 + any-of-labels: "awaiting-changes,awaiting-feedback" + exempt-issue-labels: "WIP" + exempt-pr-labels: "WIP" + repo-token: "${{ secrets.GITHUB_TOKEN }}" diff --git a/.github/workflows/linting.yml b/.github/workflows/linting.yml index 858d622e..888cb4bc 100644 --- a/.github/workflows/linting.yml +++ b/.github/workflows/linting.yml @@ -78,7 +78,7 @@ jobs: - uses: actions/setup-python@v4 with: - python-version: "3.7" + python-version: "3.8" architecture: "x64" - name: Install dependencies diff --git a/.pre-commit-config.yaml b/.pre-commit-config.yaml new file mode 100644 index 00000000..0c31cdb9 --- /dev/null +++ b/.pre-commit-config.yaml @@ -0,0 +1,5 @@ +repos: + - repo: https://github.com/pre-commit/mirrors-prettier + rev: "v2.7.1" + hooks: + - id: prettier diff --git a/CHANGELOG.md b/CHANGELOG.md index ec3c5bec..fed2891a 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -3,7 +3,7 @@ The format is based on [Keep a Changelog](https://keepachangelog.com/en/1.0.0/) and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0.html). -## v2.5.0dev - [date] +## v2.6.0dev - [date] Initial release of nf-core/ampliseq, created with the [nf-core](https://nf-co.re/) template. diff --git a/README.md b/README.md index 4ba6de4c..a891d317 100644 --- a/README.md +++ b/README.md @@ -8,57 +8,71 @@ [![run with singularity](https://img.shields.io/badge/run%20with-singularity-1d355c.svg?labelColor=000000)](https://sylabs.io/docs/) [![Launch on Nextflow Tower](https://img.shields.io/badge/Launch%20%F0%9F%9A%80-Nextflow%20Tower-%234256e7)](https://tower.nf/launch?pipeline=https://github.com/nf-core/ampliseq) -[![Get help on Slack](http://img.shields.io/badge/slack-nf--core%20%23ampliseq-4A154B?labelColor=000000&logo=slack)](https://nfcore.slack.com/channels/ampliseq)[![Follow on Twitter](http://img.shields.io/badge/twitter-%40nf__core-1DA1F2?labelColor=000000&logo=twitter)](https://twitter.com/nf_core)[![Watch on YouTube](http://img.shields.io/badge/youtube-nf--core-FF0000?labelColor=000000&logo=youtube)](https://www.youtube.com/c/nf-core) +[![Get help on Slack](http://img.shields.io/badge/slack-nf--core%20%23ampliseq-4A154B?labelColor=000000&logo=slack)](https://nfcore.slack.com/channels/ampliseq)[![Follow on Twitter](http://img.shields.io/badge/twitter-%40nf__core-1DA1F2?labelColor=000000&logo=twitter)](https://twitter.com/nf_core)[![Follow on Mastodon](https://img.shields.io/badge/mastodon-nf__core-6364ff?labelColor=FFFFFF&logo=mastodon)](https://mstdn.science/@nf_core)[![Watch on YouTube](http://img.shields.io/badge/youtube-nf--core-FF0000?labelColor=000000&logo=youtube)](https://www.youtube.com/c/nf-core) ## Introduction - +**nf-core/ampliseq** is a bioinformatics pipeline that ... -**nf-core/ampliseq** is a bioinformatics best-practice analysis pipeline for Amplicon sequencing analysis workflow using DADA2 and QIIME2. - -The pipeline is built using [Nextflow](https://www.nextflow.io), a workflow tool to run tasks across multiple compute infrastructures in a very portable manner. It uses Docker/Singularity containers making installation trivial and results highly reproducible. The [Nextflow DSL2](https://www.nextflow.io/docs/latest/dsl2.html) implementation of this pipeline uses one container per process which makes it much easier to maintain and update software dependencies. Where possible, these processes have been submitted to and installed from [nf-core/modules](https://github.com/nf-core/modules) in order to make them available to all nf-core pipelines, and to everyone within the Nextflow community! - - - -On release, automated continuous integration tests run the pipeline on a full-sized dataset on the AWS cloud infrastructure. This ensures that the pipeline runs on AWS, has sensible resource allocation defaults set to run on real-world datasets, and permits the persistent storage of results to benchmark between pipeline releases and other analysis sources.The results obtained from the full-sized test can be viewed on the [nf-core website](https://nf-co.re/ampliseq/results). - -## Pipeline summary + + 1. Read QC ([`FastQC`](https://www.bioinformatics.babraham.ac.uk/projects/fastqc/)) 2. Present QC for raw reads ([`MultiQC`](http://multiqc.info/)) -## Quick Start +## Usage + +> **Note** +> If you are new to Nextflow and nf-core, please refer to [this page](https://nf-co.re/docs/usage/installation) on how +> to set-up Nextflow. Make sure to [test your setup](https://nf-co.re/docs/usage/introduction#how-to-run-a-pipeline) +> with `-profile test` before running the workflow on actual data. + + - Note that some form of configuration will be needed so that Nextflow knows how to fetch the required software. This is usually done in the form of a config profile (`YOURPROFILE` in the example command above). You can chain multiple config profiles in a comma-separated string. +Now, you can run the pipeline using: - > - The pipeline comes with config profiles called `docker`, `singularity`, `podman`, `shifter`, `charliecloud` and `conda` which instruct the pipeline to use the named tool for software management. For example, `-profile test,docker`. - > - Please check [nf-core/configs](https://github.com/nf-core/configs#documentation) to see if a custom config file to run nf-core pipelines already exists for your Institute. If so, you can simply use `-profile ` in your command. This will enable either `docker` or `singularity` and set the appropriate execution settings for your local compute environment. - > - If you are using `singularity`, please use the [`nf-core download`](https://nf-co.re/tools/#downloading-pipelines-for-offline-use) command to download images first, before running the pipeline. Setting the [`NXF_SINGULARITY_CACHEDIR` or `singularity.cacheDir`](https://www.nextflow.io/docs/latest/singularity.html?#singularity-docker-hub) Nextflow options enables you to store and re-use the images from a central location for future pipeline runs. - > - If you are using `conda`, it is highly recommended to use the [`NXF_CONDA_CACHEDIR` or `conda.cacheDir`](https://www.nextflow.io/docs/latest/conda.html) settings to store the environments in a central location for future pipeline runs. + -4. Start running your own analysis! +```bash +nextflow run nf-core/ampliseq \ + -profile \ + --input samplesheet.csv \ + --outdir +``` - +> **Warning:** +> Please provide pipeline parameters via the CLI or Nextflow `-params-file` option. Custom config files including those +> provided by the `-c` Nextflow option can be used to provide any configuration _**except for parameters**_; +> see [docs](https://nf-co.re/usage/configuration#custom-configuration-files). - ```bash - nextflow run nf-core/ampliseq --input samplesheet.csv --outdir --genome GRCh37 -profile - ``` +For more details, please refer to the [usage documentation](https://nf-co.re/ampliseq/usage) and the [parameter documentation](https://nf-co.re/ampliseq/parameters). -## Documentation +## Pipeline output -The nf-core/ampliseq pipeline comes with documentation about the pipeline [usage](https://nf-co.re/ampliseq/usage), [parameters](https://nf-co.re/ampliseq/parameters) and [output](https://nf-co.re/ampliseq/output). +To see the the results of a test run with a full size dataset refer to the [results](https://nf-co.re/ampliseq/results) tab on the nf-core website pipeline page. +For more details about the output files and reports, please refer to the +[output documentation](https://nf-co.re/ampliseq/output). ## Credits diff --git a/bin/check_samplesheet.py b/bin/check_samplesheet.py index 11b15572..4a758fe0 100755 --- a/bin/check_samplesheet.py +++ b/bin/check_samplesheet.py @@ -158,9 +158,6 @@ def sniff_format(handle): peek = read_head(handle) handle.seek(0) sniffer = csv.Sniffer() - if not sniffer.has_header(peek): - logger.critical("The given sample sheet does not appear to contain a header.") - sys.exit(1) dialect = sniffer.sniff(peek) return dialect diff --git a/conf/base.config b/conf/base.config index 26449840..478c1657 100644 --- a/conf/base.config +++ b/conf/base.config @@ -15,7 +15,7 @@ process { memory = { check_max( 6.GB * task.attempt, 'memory' ) } time = { check_max( 4.h * task.attempt, 'time' ) } - errorStrategy = { task.exitStatus in [143,137,104,134,139] ? 'retry' : 'finish' } + errorStrategy = { task.exitStatus in ((130..145) + 104) ? 'retry' : 'finish' } maxRetries = 1 maxErrors = '-1' diff --git a/conf/igenomes.config b/conf/igenomes.config index 7a1b3ac6..3f114377 100644 --- a/conf/igenomes.config +++ b/conf/igenomes.config @@ -36,6 +36,14 @@ params { macs_gsize = "2.7e9" blacklist = "${projectDir}/assets/blacklists/hg38-blacklist.bed" } + 'CHM13' { + fasta = "${params.igenomes_base}/Homo_sapiens/UCSC/CHM13/Sequence/WholeGenomeFasta/genome.fa" + bwa = "${params.igenomes_base}/Homo_sapiens/UCSC/CHM13/Sequence/BWAIndex/" + bwamem2 = "${params.igenomes_base}/Homo_sapiens/UCSC/CHM13/Sequence/BWAmem2Index/" + gtf = "${params.igenomes_base}/Homo_sapiens/NCBI/CHM13/Annotation/Genes/genes.gtf" + gff = "ftp://ftp.ncbi.nlm.nih.gov/genomes/all/GCF/009/914/755/GCF_009914755.1_T2T-CHM13v2.0/GCF_009914755.1_T2T-CHM13v2.0_genomic.gff.gz" + mito_name = "chrM" + } 'GRCm38' { fasta = "${params.igenomes_base}/Mus_musculus/Ensembl/GRCm38/Sequence/WholeGenomeFasta/genome.fa" bwa = "${params.igenomes_base}/Mus_musculus/Ensembl/GRCm38/Sequence/BWAIndex/version0.6.0/" diff --git a/conf/test_full.config b/conf/test_full.config index 80fecde4..9bd5e2e9 100644 --- a/conf/test_full.config +++ b/conf/test_full.config @@ -10,6 +10,8 @@ ---------------------------------------------------------------------------------------- */ +cleanup = true + params { config_profile_name = 'Full test profile' config_profile_description = 'Full test dataset to check pipeline function' diff --git a/docs/usage.md b/docs/usage.md index 9e859ddb..fab27df7 100644 --- a/docs/usage.md +++ b/docs/usage.md @@ -71,6 +71,29 @@ work # Directory containing the nextflow working files # Other nextflow hidden files, eg. history of pipeline runs and old logs. ``` +If you wish to repeatedly use the same parameters for multiple runs, rather than specifying each flag in the command, you can specify these in a params file. + +Pipeline settings can be provided in a `yaml` or `json` file via `-params-file `. + +> ⚠️ Do not use `-c ` to specify parameters as this will result in errors. Custom config files specified with `-c` must only be used for [tuning process resource specifications](https://nf-co.re/docs/usage/configuration#tuning-workflow-resources), other infrastructural tweaks (such as output directories), or module arguments (args). +> The above pipeline run specified with a params file in yaml format: + +```bash +nextflow run nf-core/ampliseq -profile docker -params-file params.yaml +``` + +with `params.yaml` containing: + +```yaml +input: './samplesheet.csv' +outdir: './results/' +genome: 'GRCh37' +input: 'data' +<...> +``` + +You can also generate such `YAML`/`JSON` files via [nf-core/launch](https://nf-co.re/launch). + ### Updating the pipeline When you run the above command, Nextflow automatically pulls the pipeline code from GitHub and stores it as a cached version. When running the pipeline after this, it will always use the cached version if available - even if the pipeline has been updated since. To make sure that you're running the latest version of the pipeline, make sure that you regularly update the cached version of the pipeline: @@ -87,6 +110,10 @@ First, go to the [nf-core/ampliseq releases page](https://github.com/nf-core/amp This version number will be logged in reports when you run the pipeline, so that you'll know what you used when you look back in the future. For example, at the bottom of the MultiQC reports. +To further assist in reproducbility, you can use share and re-use [parameter files](#running-the-pipeline) to repeat pipeline runs with the same settings without having to write out a command with every single parameter. + +> 💡 If you wish to share such profile (such as upload as supplementary material for academic publications), make sure to NOT include cluster specific paths to files, nor institutional specific profiles. + ## Core Nextflow arguments > **NB:** These options are part of Nextflow and use a _single_ hyphen (pipeline parameters use a double-hyphen). @@ -95,7 +122,7 @@ This version number will be logged in reports when you run the pipeline, so that Use this parameter to choose a configuration profile. Profiles can give configuration presets for different compute environments. -Several generic profiles are bundled with the pipeline which instruct the pipeline to use software packaged using different methods (Docker, Singularity, Podman, Shifter, Charliecloud, Conda) - see below. +Several generic profiles are bundled with the pipeline which instruct the pipeline to use software packaged using different methods (Docker, Singularity, Podman, Shifter, Charliecloud, Apptainer, Conda) - see below. > We highly recommend the use of Docker or Singularity containers for full pipeline reproducibility, however when this is not possible, Conda is also supported. @@ -119,8 +146,10 @@ If `-profile` is not specified, the pipeline will run locally and expect all sof - A generic configuration profile to be used with [Shifter](https://nersc.gitlab.io/development/shifter/how-to-use/) - `charliecloud` - A generic configuration profile to be used with [Charliecloud](https://hpc.github.io/charliecloud/) +- `apptainer` + - A generic configuration profile to be used with [Apptainer](https://apptainer.org/) - `conda` - - A generic configuration profile to be used with [Conda](https://conda.io/docs/). Please only use Conda as a last resort i.e. when it's not possible to run the pipeline with Docker, Singularity, Podman, Shifter or Charliecloud. + - A generic configuration profile to be used with [Conda](https://conda.io/docs/). Please only use Conda as a last resort i.e. when it's not possible to run the pipeline with Docker, Singularity, Podman, Shifter, Charliecloud, or Apptainer. ### `-resume` @@ -138,102 +167,19 @@ Specify the path to a specific config file (this is a core Nextflow command). Se Whilst the default requirements set within the pipeline will hopefully work for most people and with most input data, you may find that you want to customise the compute resources that the pipeline requests. Each step in the pipeline has a default set of requirements for number of CPUs, memory and time. For most of the steps in the pipeline, if the job exits with any of the error codes specified [here](https://github.com/nf-core/rnaseq/blob/4c27ef5610c87db00c3c5a3eed10b1d161abf575/conf/base.config#L18) it will automatically be resubmitted with higher requests (2 x original, then 3 x original). If it still fails after the third attempt then the pipeline execution is stopped. -For example, if the nf-core/rnaseq pipeline is failing after multiple re-submissions of the `STAR_ALIGN` process due to an exit code of `137` this would indicate that there is an out of memory issue: - -```console -[62/149eb0] NOTE: Process `NFCORE_RNASEQ:RNASEQ:ALIGN_STAR:STAR_ALIGN (WT_REP1)` terminated with an error exit status (137) -- Execution is retried (1) -Error executing process > 'NFCORE_RNASEQ:RNASEQ:ALIGN_STAR:STAR_ALIGN (WT_REP1)' - -Caused by: - Process `NFCORE_RNASEQ:RNASEQ:ALIGN_STAR:STAR_ALIGN (WT_REP1)` terminated with an error exit status (137) - -Command executed: - STAR \ - --genomeDir star \ - --readFilesIn WT_REP1_trimmed.fq.gz \ - --runThreadN 2 \ - --outFileNamePrefix WT_REP1. \ - - -Command exit status: - 137 - -Command output: - (empty) - -Command error: - .command.sh: line 9: 30 Killed STAR --genomeDir star --readFilesIn WT_REP1_trimmed.fq.gz --runThreadN 2 --outFileNamePrefix WT_REP1. -Work dir: - /home/pipelinetest/work/9d/172ca5881234073e8d76f2a19c88fb - -Tip: you can replicate the issue by changing to the process work dir and entering the command `bash .command.run` -``` - -#### For beginners - -A first step to bypass this error, you could try to increase the amount of CPUs, memory, and time for the whole pipeline. Therefor you can try to increase the resource for the parameters `--max_cpus`, `--max_memory`, and `--max_time`. Based on the error above, you have to increase the amount of memory. Therefore you can go to the [parameter documentation of rnaseq](https://nf-co.re/rnaseq/3.9/parameters) and scroll down to the `show hidden parameter` button to get the default value for `--max_memory`. In this case 128GB, you than can try to run your pipeline again with `--max_memory 200GB -resume` to skip all process, that were already calculated. If you can not increase the resource of the complete pipeline, you can try to adapt the resource for a single process as mentioned below. - -#### Advanced option on process level - -To bypass this error you would need to find exactly which resources are set by the `STAR_ALIGN` process. The quickest way is to search for `process STAR_ALIGN` in the [nf-core/rnaseq Github repo](https://github.com/nf-core/rnaseq/search?q=process+STAR_ALIGN). -We have standardised the structure of Nextflow DSL2 pipelines such that all module files will be present in the `modules/` directory and so, based on the search results, the file we want is `modules/nf-core/star/align/main.nf`. -If you click on the link to that file you will notice that there is a `label` directive at the top of the module that is set to [`label process_high`](https://github.com/nf-core/rnaseq/blob/4c27ef5610c87db00c3c5a3eed10b1d161abf575/modules/nf-core/software/star/align/main.nf#L9). -The [Nextflow `label`](https://www.nextflow.io/docs/latest/process.html#label) directive allows us to organise workflow processes in separate groups which can be referenced in a configuration file to select and configure subset of processes having similar computing requirements. -The default values for the `process_high` label are set in the pipeline's [`base.config`](https://github.com/nf-core/rnaseq/blob/4c27ef5610c87db00c3c5a3eed10b1d161abf575/conf/base.config#L33-L37) which in this case is defined as 72GB. -Providing you haven't set any other standard nf-core parameters to **cap** the [maximum resources](https://nf-co.re/usage/configuration#max-resources) used by the pipeline then we can try and bypass the `STAR_ALIGN` process failure by creating a custom config file that sets at least 72GB of memory, in this case increased to 100GB. -The custom config below can then be provided to the pipeline via the [`-c`](#-c) parameter as highlighted in previous sections. - -```nextflow -process { - withName: 'NFCORE_RNASEQ:RNASEQ:ALIGN_STAR:STAR_ALIGN' { - memory = 100.GB - } -} -``` - -> **NB:** We specify the full process name i.e. `NFCORE_RNASEQ:RNASEQ:ALIGN_STAR:STAR_ALIGN` in the config file because this takes priority over the short name (`STAR_ALIGN`) and allows existing configuration using the full process name to be correctly overridden. -> -> If you get a warning suggesting that the process selector isn't recognised check that the process name has been specified correctly. - -### Updating containers (advanced users) - -The [Nextflow DSL2](https://www.nextflow.io/docs/latest/dsl2.html) implementation of this pipeline uses one container per process which makes it much easier to maintain and update software dependencies. If for some reason you need to use a different version of a particular tool with the pipeline then you just need to identify the `process` name and override the Nextflow `container` definition for that process using the `withName` declaration. For example, in the [nf-core/viralrecon](https://nf-co.re/viralrecon) pipeline a tool called [Pangolin](https://github.com/cov-lineages/pangolin) has been used during the COVID-19 pandemic to assign lineages to SARS-CoV-2 genome sequenced samples. Given that the lineage assignments change quite frequently it doesn't make sense to re-release the nf-core/viralrecon everytime a new version of Pangolin has been released. However, you can override the default container used by the pipeline by creating a custom config file and passing it as a command-line argument via `-c custom.config`. - -1. Check the default version used by the pipeline in the module file for [Pangolin](https://github.com/nf-core/viralrecon/blob/a85d5969f9025409e3618d6c280ef15ce417df65/modules/nf-core/software/pangolin/main.nf#L14-L19) -2. Find the latest version of the Biocontainer available on [Quay.io](https://quay.io/repository/biocontainers/pangolin?tag=latest&tab=tags) -3. Create the custom config accordingly: - - - For Docker: +To change the resource requests, please see the [max resources](https://nf-co.re/docs/usage/configuration#max-resources) and [tuning workflow resources](https://nf-co.re/docs/usage/configuration#tuning-workflow-resources) section of the nf-core website. - ```nextflow - process { - withName: PANGOLIN { - container = 'quay.io/biocontainers/pangolin:3.0.5--pyhdfd78af_0' - } - } - ``` +### Custom Containers - - For Singularity: +In some cases you may wish to change which container or conda environment a step of the pipeline uses for a particular tool. By default nf-core pipelines use containers and software from the [biocontainers](https://biocontainers.pro/) or [bioconda](https://bioconda.github.io/) projects. However in some cases the pipeline specified version maybe out of date. - ```nextflow - process { - withName: PANGOLIN { - container = 'https://depot.galaxyproject.org/singularity/pangolin:3.0.5--pyhdfd78af_0' - } - } - ``` +To use a different container from the default container or conda environment specified in a pipeline, please see the [updating tool versions](https://nf-co.re/docs/usage/configuration#updating-tool-versions) section of the nf-core website. - - For Conda: +### Custom Tool Arguments - ```nextflow - process { - withName: PANGOLIN { - conda = 'bioconda::pangolin=3.0.5' - } - } - ``` +A pipeline might not always support every possible argument or option of a particular tool used in pipeline. Fortunately, nf-core pipelines provide some freedom to users to insert additional parameters that the pipeline does not include by default. -> **NB:** If you wish to periodically update individual tool-specific results (e.g. Pangolin) generated by the pipeline then you must ensure to keep the `work/` directory otherwise the `-resume` ability of the pipeline will be compromised and it will restart from scratch. +To learn how to provide additional arguments to a particular tool of the pipeline, please see the [customising tool arguments](https://nf-co.re/docs/usage/configuration#customising-tool-arguments) section of the nf-core website. ### nf-core/configs diff --git a/lib/NfcoreSchema.groovy b/lib/NfcoreSchema.groovy index 33cd4f6e..9b34804d 100755 --- a/lib/NfcoreSchema.groovy +++ b/lib/NfcoreSchema.groovy @@ -2,6 +2,7 @@ // This file holds several functions used to perform JSON parameter validation, help and summary rendering for the nf-core pipeline template. // +import nextflow.Nextflow import org.everit.json.schema.Schema import org.everit.json.schema.loader.SchemaLoader import org.everit.json.schema.ValidationException @@ -83,6 +84,7 @@ class NfcoreSchema { 'stub-run', 'test', 'w', + 'with-apptainer', 'with-charliecloud', 'with-conda', 'with-dag', @@ -177,7 +179,7 @@ class NfcoreSchema { } if (has_error) { - System.exit(1) + Nextflow.error('Exiting!') } } diff --git a/lib/WorkflowAmpliseq.groovy b/lib/WorkflowAmpliseq.groovy index 02a96f23..a576b038 100755 --- a/lib/WorkflowAmpliseq.groovy +++ b/lib/WorkflowAmpliseq.groovy @@ -2,6 +2,7 @@ // This file holds several functions specific to the workflow/ampliseq.nf in the nf-core/ampliseq pipeline // +import nextflow.Nextflow import groovy.text.SimpleTemplateEngine class WorkflowAmpliseq { @@ -14,8 +15,7 @@ class WorkflowAmpliseq { if (!params.fasta) { - log.error "Genome fasta file not specified with e.g. '--fasta genome.fa' or via a detectable config file." - System.exit(1) + Nextflow.error "Genome fasta file not specified with e.g. '--fasta genome.fa' or via a detectable config file." } } @@ -61,17 +61,19 @@ class WorkflowAmpliseq { def description_html = engine.createTemplate(methods_text).make(meta) return description_html - }// + } + + // // Exit pipeline if incorrect --genome key provided // private static void genomeExistsError(params, log) { if (params.genomes && params.genome && !params.genomes.containsKey(params.genome)) { - log.error "~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~\n" + + def error_string = "~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~\n" + " Genome '${params.genome}' not found in any config files provided to the pipeline.\n" + " Currently, the available genome keys are:\n" + " ${params.genomes.keySet().join(", ")}\n" + "~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~" - System.exit(1) + Nextflow.error(error_string) } } } diff --git a/lib/WorkflowMain.groovy b/lib/WorkflowMain.groovy index a8d64eec..88093eb2 100755 --- a/lib/WorkflowMain.groovy +++ b/lib/WorkflowMain.groovy @@ -2,6 +2,8 @@ // This file holds several functions specific to the main.nf workflow in the nf-core/ampliseq pipeline // +import nextflow.Nextflow + class WorkflowMain { // @@ -21,7 +23,7 @@ class WorkflowMain { // // Generate help string // - public static String help(workflow, params, log) { + public static String help(workflow, params) { def command = "nextflow run ${workflow.manifest.name} --input samplesheet.csv --genome GRCh37 -profile docker" def help_string = '' help_string += NfcoreTemplate.logo(workflow, params.monochrome_logs) @@ -34,7 +36,7 @@ class WorkflowMain { // // Generate parameter summary log string // - public static String paramsSummaryLog(workflow, params, log) { + public static String paramsSummaryLog(workflow, params) { def summary_log = '' summary_log += NfcoreTemplate.logo(workflow, params.monochrome_logs) summary_log += NfcoreSchema.paramsSummaryLog(workflow, params) @@ -49,7 +51,7 @@ class WorkflowMain { public static void initialise(workflow, params, log) { // Print help to screen if required if (params.help) { - log.info help(workflow, params, log) + log.info help(workflow, params) System.exit(0) } @@ -61,7 +63,7 @@ class WorkflowMain { } // Print parameter summary log to screen - log.info paramsSummaryLog(workflow, params, log) + log.info paramsSummaryLog(workflow, params) // Validate workflow parameters via the JSON schema if (params.validate_params) { @@ -81,8 +83,7 @@ class WorkflowMain { // Check input has been provided if (!params.input) { - log.error "Please provide an input samplesheet to the pipeline e.g. '--input samplesheet.csv'" - System.exit(1) + Nextflow.error("Please provide an input samplesheet to the pipeline e.g. '--input samplesheet.csv'") } } // diff --git a/main.nf b/main.nf index 98ce2481..ab10ddb1 100644 --- a/main.nf +++ b/main.nf @@ -4,7 +4,6 @@ nf-core/ampliseq ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ Github : https://github.com/nf-core/ampliseq - Website: https://nf-co.re/ampliseq Slack : https://nfcore.slack.com/channels/ampliseq ---------------------------------------------------------------------------------------- diff --git a/modules.json b/modules.json index 41975cad..e48dd38b 100644 --- a/modules.json +++ b/modules.json @@ -7,7 +7,7 @@ "nf-core": { "custom/dumpsoftwareversions": { "branch": "master", - "git_sha": "c8e35eb2055c099720a75538d1b8adb3fb5a464c", + "git_sha": "76cc4938c1f6ea5c7d83fed1eeffc146787f9543", "installed_by": ["modules"] }, "fastqc": { @@ -17,7 +17,7 @@ }, "multiqc": { "branch": "master", - "git_sha": "c8e35eb2055c099720a75538d1b8adb3fb5a464c", + "git_sha": "f2d63bd5b68925f98f572eed70993d205cc694b7", "installed_by": ["modules"] } } diff --git a/modules/local/samplesheet_check.nf b/modules/local/samplesheet_check.nf index d34407ff..c2633ba8 100644 --- a/modules/local/samplesheet_check.nf +++ b/modules/local/samplesheet_check.nf @@ -5,7 +5,7 @@ process SAMPLESHEET_CHECK { conda "conda-forge::python=3.8.3" container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? 'https://depot.galaxyproject.org/singularity/python:3.8.3' : - 'quay.io/biocontainers/python:3.8.3' }" + 'biocontainers/python:3.8.3' }" input: path samplesheet diff --git a/modules/nf-core/custom/dumpsoftwareversions/main.nf b/modules/nf-core/custom/dumpsoftwareversions/main.nf index 3df21765..800a6099 100644 --- a/modules/nf-core/custom/dumpsoftwareversions/main.nf +++ b/modules/nf-core/custom/dumpsoftwareversions/main.nf @@ -2,10 +2,10 @@ process CUSTOM_DUMPSOFTWAREVERSIONS { label 'process_single' // Requires `pyyaml` which does not have a dedicated container but is in the MultiQC container - conda "bioconda::multiqc=1.13" + conda "bioconda::multiqc=1.14" container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? - 'https://depot.galaxyproject.org/singularity/multiqc:1.13--pyhdfd78af_0' : - 'quay.io/biocontainers/multiqc:1.13--pyhdfd78af_0' }" + 'https://depot.galaxyproject.org/singularity/multiqc:1.14--pyhdfd78af_0' : + 'quay.io/biocontainers/multiqc:1.14--pyhdfd78af_0' }" input: path versions diff --git a/modules/nf-core/custom/dumpsoftwareversions/meta.yml b/modules/nf-core/custom/dumpsoftwareversions/meta.yml index 60b546a0..c32657de 100644 --- a/modules/nf-core/custom/dumpsoftwareversions/meta.yml +++ b/modules/nf-core/custom/dumpsoftwareversions/meta.yml @@ -1,7 +1,9 @@ +# yaml-language-server: $schema=https://raw.githubusercontent.com/nf-core/modules/master/modules/yaml-schema.json name: custom_dumpsoftwareversions description: Custom module used to dump software versions within the nf-core pipeline template keywords: - custom + - dump - version tools: - custom: diff --git a/modules/nf-core/multiqc/main.nf b/modules/nf-core/multiqc/main.nf index 68f66bea..4b604749 100644 --- a/modules/nf-core/multiqc/main.nf +++ b/modules/nf-core/multiqc/main.nf @@ -1,10 +1,10 @@ process MULTIQC { label 'process_single' - conda "bioconda::multiqc=1.13" + conda "bioconda::multiqc=1.14" container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? - 'https://depot.galaxyproject.org/singularity/multiqc:1.13--pyhdfd78af_0' : - 'quay.io/biocontainers/multiqc:1.13--pyhdfd78af_0' }" + 'https://depot.galaxyproject.org/singularity/multiqc:1.14--pyhdfd78af_0' : + 'quay.io/biocontainers/multiqc:1.14--pyhdfd78af_0' }" input: path multiqc_files, stageAs: "?/*" diff --git a/modules/nf-core/multiqc/meta.yml b/modules/nf-core/multiqc/meta.yml index ebc29b27..f93b5ee5 100644 --- a/modules/nf-core/multiqc/meta.yml +++ b/modules/nf-core/multiqc/meta.yml @@ -1,3 +1,4 @@ +# yaml-language-server: $schema=https://raw.githubusercontent.com/nf-core/modules/master/modules/yaml-schema.json name: MultiQC description: Aggregate results from bioinformatics analyses across many samples into a single report keywords: @@ -37,7 +38,7 @@ output: description: MultiQC report file pattern: "multiqc_report.html" - data: - type: dir + type: directory description: MultiQC data dir pattern: "multiqc_data" - plots: diff --git a/nextflow.config b/nextflow.config index 9a159e4b..9c823f7d 100644 --- a/nextflow.config +++ b/nextflow.config @@ -78,7 +78,11 @@ try { profiles { - debug { process.beforeScript = 'echo $HOSTNAME' } + debug { + dumpHashes = true + process.beforeScript = 'echo $HOSTNAME' + cleanup = false + } conda { conda.enabled = true docker.enabled = false @@ -86,6 +90,7 @@ profiles { podman.enabled = false shifter.enabled = false charliecloud.enabled = false + apptainer.enabled = false } mamba { conda.enabled = true @@ -95,14 +100,18 @@ profiles { podman.enabled = false shifter.enabled = false charliecloud.enabled = false + apptainer.enabled = false } docker { docker.enabled = true + docker.registry = 'quay.io' docker.userEmulation = true + conda.enabled = false singularity.enabled = false podman.enabled = false shifter.enabled = false charliecloud.enabled = false + apptainer.enabled = false } arm { docker.runOptions = '-u $(id -u):$(id -g) --platform=linux/amd64' @@ -110,31 +119,49 @@ profiles { singularity { singularity.enabled = true singularity.autoMounts = true + conda.enabled = false docker.enabled = false podman.enabled = false shifter.enabled = false charliecloud.enabled = false + apptainer.enabled = false } podman { podman.enabled = true + podman.registry = 'quay.io' + conda.enabled = false docker.enabled = false singularity.enabled = false shifter.enabled = false charliecloud.enabled = false + apptainer.enabled = false } shifter { shifter.enabled = true + conda.enabled = false docker.enabled = false singularity.enabled = false podman.enabled = false charliecloud.enabled = false + apptainer.enabled = false } charliecloud { charliecloud.enabled = true + conda.enabled = false docker.enabled = false singularity.enabled = false podman.enabled = false shifter.enabled = false + apptainer.enabled = false + } + apptainer { + apptainer.enabled = true + conda.enabled = false + docker.enabled = false + singularity.enabled = false + podman.enabled = false + shifter.enabled = false + charliecloud.enabled = false } gitpod { executor.name = 'local' @@ -193,7 +220,7 @@ manifest { description = """Amplicon sequencing analysis workflow using DADA2 and QIIME2""" mainScript = 'main.nf' nextflowVersion = '!>=22.10.1' - version = '2.5.0dev' + version = '2.6.0dev' doi = '' } diff --git a/tower.yml b/tower.yml new file mode 100644 index 00000000..787aedfe --- /dev/null +++ b/tower.yml @@ -0,0 +1,5 @@ +reports: + multiqc_report.html: + display: "MultiQC HTML report" + samplesheet.csv: + display: "Auto-created samplesheet with collated metadata and FASTQ paths" From 39429882b0d833441b5db63f2318791745010731 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Wed, 3 May 2023 08:41:09 +0200 Subject: [PATCH 068/158] Revert test config to pacbio data, fix qiime test --- conf/test_sintax.config | 5 ++--- 1 file changed, 2 insertions(+), 3 deletions(-) diff --git a/conf/test_sintax.config b/conf/test_sintax.config index d220f434..0021d195 100644 --- a/conf/test_sintax.config +++ b/conf/test_sintax.config @@ -29,7 +29,6 @@ params { cut_its = "its2" sintax_ref_taxonomy = "unite-fungi" - skip_dada_taxonomy = true //this is to remove low abundance ASVs to reduce runtime of downstream processes @@ -37,14 +36,14 @@ params { min_frequency = 10 //produce average barplots - metadata_category_barplot = "treatment1,badpairwise10" + metadata_category_barplot = "var2,var3" //restrict ANCOM analysis to higher taxonomic levels tax_agglom_max = 4 sbdiexport = true - qiime_adonis_formula = "treatment1,mix8" + qiime_adonis_formula = "var2" diversity_rarefaction_depth = 500 } From e1916e3b8782ee2571a45f8258dffc4b306b9477 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Wed, 3 May 2023 09:27:49 +0200 Subject: [PATCH 069/158] Update CHANGELOG --- CHANGELOG.md | 1 + 1 file changed, 1 insertion(+) diff --git a/CHANGELOG.md b/CHANGELOG.md index 1b17dbbb..0ff77334 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -8,6 +8,7 @@ and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0 ### `Added` - [#564](https://github.com/nf-core/ampliseq/pull/564),[#567](https://github.com/nf-core/ampliseq/pull/567) - Added phylogenetic placement +- [#577](https://github.com/nf-core/ampliseq/pull/577),[#577](https://github.com/nf-core/ampliseq/pull/567) - Added SINTAX for taxonomic classification ### `Changed` From e1d0f0ec80bf143907a3f3b7843b10197dfa8753 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Wed, 3 May 2023 11:30:33 +0200 Subject: [PATCH 070/158] Fix spaces --- conf/modules.config | 4 ++-- conf/ref_databases.config | 2 +- 2 files changed, 3 insertions(+), 3 deletions(-) diff --git a/conf/modules.config b/conf/modules.config index 10707a68..d23d4c80 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -391,13 +391,13 @@ process { withName: VSEARCH_SINTAX { ext.args = "--sintax_cutoff 0.8 --randseed ${params.seed}" - cpus = 1 + cpus = 1 publishDir = [ [ path: { "${params.outdir}/sintax" }, mode: params.publish_dir_mode, pattern: "*.tsv" - ] + ] ] } diff --git a/conf/ref_databases.config b/conf/ref_databases.config index 9ffe2300..cd408bf7 100644 --- a/conf/ref_databases.config +++ b/conf/ref_databases.config @@ -292,7 +292,7 @@ params { } 'unite-fungi=8.2' { title = "UNITE USEARCH/UTAX release for Fungi - Version 8.2" - file = [ "https://files.plutof.ut.ee/public/orig/E8/83/E883EB19E3EA7B64C1F652521301239831FAFE0BFF015C9E2B4786DC0976C0FC.gz" ] + file = [ "https://files.plutof.ut.ee/public/orig/E8/83/E883EB19E3EA7B64C1F652521301239831FAFE0BFF015C9E2B4786DC0976C0FC.gz" ] citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2020): UNITE USEARCH/UTAX release for Fungi. UNITE Community. 10.15156/BIO/786375" fmtscript = "taxref_reformat_sintax.sh" dbversion = "UNITE-fungi v8.2 (https://doi.org/10.15156/BIO/786375)" From 139d5b5979dd064c544057d3fa6614be7f931b3d Mon Sep 17 00:00:00 2001 From: daniel Date: Wed, 3 May 2023 17:03:30 +0200 Subject: [PATCH 071/158] specify docker.io for ubuntu containers --- modules/local/format_fastainput.nf | 2 +- modules/local/rename_raw_data_files.nf | 2 +- 2 files changed, 2 insertions(+), 2 deletions(-) diff --git a/modules/local/format_fastainput.nf b/modules/local/format_fastainput.nf index b97b3184..92cb587b 100644 --- a/modules/local/format_fastainput.nf +++ b/modules/local/format_fastainput.nf @@ -4,7 +4,7 @@ process FORMAT_FASTAINPUT { conda "conda-forge::sed=4.7" container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? 'https://depot.galaxyproject.org/singularity/ubuntu:20.04' : - 'ubuntu:20.04' }" + 'docker.io/ubuntu:20.04' }" input: path(fastain) diff --git a/modules/local/rename_raw_data_files.nf b/modules/local/rename_raw_data_files.nf index 3f277c7a..8f8016bb 100644 --- a/modules/local/rename_raw_data_files.nf +++ b/modules/local/rename_raw_data_files.nf @@ -5,7 +5,7 @@ process RENAME_RAW_DATA_FILES { conda "conda-forge::sed=4.7" container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? 'https://depot.galaxyproject.org/singularity/ubuntu:20.04' : - 'ubuntu:20.04' }" + 'docker.io/ubuntu:20.04' }" input: tuple val(meta), path(reads) From d140af1f8e754cb33253e56a058039d7a9255a8c Mon Sep 17 00:00:00 2001 From: daniel Date: Wed, 3 May 2023 17:10:41 +0200 Subject: [PATCH 072/158] fix prettier --- docs/usage.md | 10 +++++----- 1 file changed, 5 insertions(+), 5 deletions(-) diff --git a/docs/usage.md b/docs/usage.md index 4bab9909..54be4b91 100644 --- a/docs/usage.md +++ b/docs/usage.md @@ -81,11 +81,11 @@ nextflow run nf-core/ampliseq -profile docker -params-file params.yaml with `params.yaml` containing: ```yaml -input: 'data' -FW_primer: 'GTGYCAGCMGCCGCGGTAA' -RV_primer: 'GGACTACNVGGGTWTCTAAT' -metadata: 'data/Metadata.tsv' -outdir: './results' +input: "data" +FW_primer: "GTGYCAGCMGCCGCGGTAA" +RV_primer: "GGACTACNVGGGTWTCTAAT" +metadata: "data/Metadata.tsv" +outdir: "./results" ``` You can also generate such `YAML`/`JSON` files via [nf-core/launch](https://nf-co.re/launch). From d4f1f1a7231c6ef5e8f9faf1c281061576a6fca8 Mon Sep 17 00:00:00 2001 From: daniel Date: Thu, 4 May 2023 09:02:00 +0200 Subject: [PATCH 073/158] add docker.io/ to biocontainers/biocontainers:v1.2.0_cv1 --- modules/local/format_taxonomy.nf | 2 +- modules/local/format_taxonomy_qiime.nf | 2 +- 2 files changed, 2 insertions(+), 2 deletions(-) diff --git a/modules/local/format_taxonomy.nf b/modules/local/format_taxonomy.nf index ded8a104..8981eaa3 100644 --- a/modules/local/format_taxonomy.nf +++ b/modules/local/format_taxonomy.nf @@ -4,7 +4,7 @@ process FORMAT_TAXONOMY { conda "conda-forge::sed=4.7" container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? 'https://containers.biocontainers.pro/s3/SingImgsRepo/biocontainers/v1.2.0_cv1/biocontainers_v1.2.0_cv1.img' : - 'biocontainers/biocontainers:v1.2.0_cv1' }" + 'docker.io/biocontainers/biocontainers:v1.2.0_cv1' }" input: path(database) diff --git a/modules/local/format_taxonomy_qiime.nf b/modules/local/format_taxonomy_qiime.nf index 8ed2ba1a..6d652745 100644 --- a/modules/local/format_taxonomy_qiime.nf +++ b/modules/local/format_taxonomy_qiime.nf @@ -5,7 +5,7 @@ process FORMAT_TAXONOMY_QIIME { conda "conda-forge::sed=4.7" container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? 'https://containers.biocontainers.pro/s3/SingImgsRepo/biocontainers/v1.2.0_cv1/biocontainers_v1.2.0_cv1.img' : - 'biocontainers/biocontainers:v1.2.0_cv1' }" + 'docker.io/biocontainers/biocontainers:v1.2.0_cv1' }" input: path(database) From d03babef8fd7623bc68fc9db833cd4cb289ae07e Mon Sep 17 00:00:00 2001 From: daniel Date: Thu, 4 May 2023 13:36:56 +0200 Subject: [PATCH 074/158] replace System.exit(1) with error() --- lib/WorkflowAmpliseq.groovy | 61 +++++++++---------------- lib/WorkflowMain.groovy | 8 ++-- subworkflows/local/cutadapt_workflow.nf | 3 +- subworkflows/local/parse_input.nf | 3 +- workflows/ampliseq.nf | 9 ++-- 5 files changed, 30 insertions(+), 54 deletions(-) diff --git a/lib/WorkflowAmpliseq.groovy b/lib/WorkflowAmpliseq.groovy index da8a1add..7aafa8d3 100755 --- a/lib/WorkflowAmpliseq.groovy +++ b/lib/WorkflowAmpliseq.groovy @@ -14,101 +14,83 @@ class WorkflowAmpliseq { if ( params.pacbio || params.iontorrent || params.single_end ) { if (params.trunclenr) { log.warn "Unused parameter: `--trunclenr` is ignored because the data is single end." } } else if (params.trunclenf && !params.trunclenr) { - log.error "Invalid command: `--trunclenf` is set, but `--trunclenr` is not. Either both parameters `--trunclenf` and `--trunclenr` must be set or none." - System.exit(1) + Nextflow.error("Invalid command: `--trunclenf` is set, but `--trunclenr` is not. Either both parameters `--trunclenf` and `--trunclenr` must be set or none.") } else if (!params.trunclenf && params.trunclenr) { - log.error "Invalid command: `--trunclenr` is set, but `--trunclenf` is not. Either both parameters `--trunclenf` and `--trunclenr` must be set or none." - System.exit(1) + Nextflow.error("Invalid command: `--trunclenr` is set, but `--trunclenf` is not. Either both parameters `--trunclenf` and `--trunclenr` must be set or none.") } if (!["pooled", "independent", "pseudo"].contains(params.sample_inference)) { - log.error "Please set `--sample_inference` to one of the following:\n" + + def error_string = "Please set `--sample_inference` to one of the following:\n" + "\t-\"independent\" (lowest sensitivity and lowest resources),\n" + "\t-\"pseudo\" (balance between required resources and sensitivity),\n" + "\t-\"pooled\" (highest sensitivity and resources)." - System.exit(1) + Nextflow.error(error_string) } if (params.double_primer && params.retain_untrimmed) { - log.error "Incompatible parameters `--double_primer` and `--retain_untrimmed` cannot be set at the same time." - System.exit(1) + Nextflow.error("Incompatible parameters `--double_primer` and `--retain_untrimmed` cannot be set at the same time.") } if ( params.min_len_asv && params.max_len_asv && (params.min_len_asv > params.max_len_asv) ) { - log.error "Incompatible parameters: `--min_len_asv` may not be greater than `--max_len_asv`." - System.exit(1) + Nextflow.error("Incompatible parameters: `--min_len_asv` may not be greater than `--max_len_asv`.") } if ( params.skip_dada_quality && (params.trunclenf == null || params.trunclenr == null) ) { - log.error "Incompatible parameters: `--skip_dada_quality` may not be used without setting `--trunclenf` and `--trunclenr`." - System.exit(1) + Nextflow.error("Incompatible parameters: `--skip_dada_quality` may not be used without setting `--trunclenf` and `--trunclenr`.") } if (params.tax_agglom_min > params.tax_agglom_max) { - log.error "Incompatible parameters: `--tax_agglom_min` may not be greater than `--tax_agglom_max`." - System.exit(1) + Nextflow.error("Incompatible parameters: `--tax_agglom_min` may not be greater than `--tax_agglom_max`.") } if (!params.dada_ref_tax_custom && params.dada_ref_tax_custom_sp) { - log.error "Incompatible parameters: `--dada_ref_tax_custom_sp` requires `--dada_ref_tax_custom`." - System.exit(1) + Nextflow.error("Incompatible parameters: `--dada_ref_tax_custom_sp` requires `--dada_ref_tax_custom`.") } if (params.dada_ref_tax_custom && !params.dada_ref_tax_custom_sp && !params.skip_dada_addspecies) { - log.error "Incompatible parameters: Either `--skip_dada_addspecies` or `--dada_ref_tax_custom_sp` is additionally required to `--dada_ref_tax_custom`." - System.exit(1) + Nextflow.error("Incompatible parameters: Either `--skip_dada_addspecies` or `--dada_ref_tax_custom_sp` is additionally required to `--dada_ref_tax_custom`.") } if (params.pplace_tree) { if (!params.pplace_aln) { - log.error "Missing parameter: Phylogenetic placement requires in addition to `--pplace_tree` also `--pplace_aln`." - System.exit(1) + Nextflow.error("Missing parameter: Phylogenetic placement requires in addition to `--pplace_tree` also `--pplace_aln`.") } if (!params.pplace_model) { - log.error "Missing parameter: Phylogenetic placement requires in addition to `--pplace_tree` also `--pplace_model`." - System.exit(1) + Nextflow.error("Missing parameter: Phylogenetic placement requires in addition to `--pplace_tree` also `--pplace_model`.") } } if (params.dada_assign_taxlevels && params.sbdiexport) { - log.error "Incompatible parameters: `--sbdiexport` expects specific taxonomics ranks (default) and therefore excludes modifying those using `--dada_assign_taxlevels`." - System.exit(1) + Nextflow.error("Incompatible parameters: `--sbdiexport` expects specific taxonomics ranks (default) and therefore excludes modifying those using `--dada_assign_taxlevels`.") } if (params.skip_dada_addspecies && params.sbdiexport) { - log.error "Incompatible parameters: `--sbdiexport` expects species annotation and therefore excludes `--skip_dada_addspecies`." - System.exit(1) + Nextflow.error("Incompatible parameters: `--sbdiexport` expects species annotation and therefore excludes `--skip_dada_addspecies`.") } if (params.skip_taxonomy && params.sbdiexport) { - log.error "Incompatible parameters: `--sbdiexport` expects taxa annotation and therefore excludes `--skip_taxonomy`." - System.exit(1) + Nextflow.error("Incompatible parameters: `--sbdiexport` expects taxa annotation and therefore excludes `--skip_taxonomy`.") } if ( (!params.FW_primer || !params.RV_primer) && params.qiime_ref_taxonomy && !params.skip_qiime && !params.skip_taxonomy ) { - log.error "Incompatible parameters: `--FW_primer` and `--RV_primer` are required for cutting the QIIME2 reference database to the amplicon sequences. Please specify primers or do not use `--qiime_ref_taxonomy`." - System.exit(1) + Nextflow.error("Incompatible parameters: `--FW_primer` and `--RV_primer` are required for cutting the QIIME2 reference database to the amplicon sequences. Please specify primers or do not use `--qiime_ref_taxonomy`.") } if ( (!params.FW_primer || !params.RV_primer) && params.cut_dada_ref_taxonomy && !params.skip_taxonomy ) { - log.error "Incompatible parameters: `--FW_primer` and `--RV_primer` are required for cutting the DADA2 reference database to the amplicon sequences. Please specify primers or do not use `--cut_dada_ref_taxonomy`." - System.exit(1) + Nextflow.error("Incompatible parameters: `--FW_primer` and `--RV_primer` are required for cutting the DADA2 reference database to the amplicon sequences. Please specify primers or do not use `--cut_dada_ref_taxonomy`.") } if (params.qiime_ref_taxonomy && params.classifier) { - log.error "Incompatible parameters: `--qiime_ref_taxonomy` will produce a classifier but `--classifier` points to a precomputed classifier, therefore, only use one of those." - System.exit(1) + Nextflow.error("Incompatible parameters: `--qiime_ref_taxonomy` will produce a classifier but `--classifier` points to a precomputed classifier, therefore, only use one of those.") } if (params.filter_ssu && params.skip_barrnap) { - log.error "Incompatible parameters: `--filter_ssu` cannot be used with `--skip_barrnap` because filtering for SSU's depends on barrnap." - System.exit(1) + Nextflow.error("Incompatible parameters: `--filter_ssu` cannot be used with `--skip_barrnap` because filtering for SSU's depends on barrnap.") } String[] sbdi_compatible_databases = ["coidb","coidb=221216","gtdb","gtdb=R07-RS207","gtdb=R06-RS202","gtdb=R05-RS95","midori2-co1","midori2-co1=gb250","pr2","pr2=4.14.0","pr2=4.13.0","rdp","rdp=18","sbdi-gtdb","sbdi-gtdb=R07-RS207-1","silva","silva=138","silva=132","unite-fungi","unite-fungi=8.3","unite-fungi=8.2","unite-alleuk","unite-alleuk=8.3","unite-alleuk=8.2"] if ( params.sbdiexport && !Arrays.stream(sbdi_compatible_databases).anyMatch(entry -> params.dada_ref_taxonomy.toString().equals(entry)) ) { - log.error "Incompatible parameters: `--sbdiexport` does not work with the chosen database of `--dada_ref_taxonomy`, because the expected taxonomic levels do not match." - System.exit(1) + Nextflow.error("Incompatible parameters: `--sbdiexport` does not work with the chosen database of `--dada_ref_taxonomy`, because the expected taxonomic levels do not match.") } if (params.addsh && !params.dada_ref_databases[params.dada_ref_taxonomy]["shfile"]) { @@ -118,8 +100,7 @@ class WorkflowAmpliseq { validDBs += " " + db } } - log.error "UNITE species hypothesis information is not available for the selected reference database, please use the option `--dada_ref_taxonomy` to select an appropriate database. Currently, the option `--addsh` can only be used together with the following UNITE reference databases:\n" + validDBs + "." - System.exit(1) + Nextflow.error("UNITE species hypothesis information is not available for the selected reference database, please use the option `--dada_ref_taxonomy` to select an appropriate database. Currently, the option `--addsh` can only be used together with the following UNITE reference databases:\n" + validDBs + ".") } if (params.addsh && params.cut_its == "none") { diff --git a/lib/WorkflowMain.groovy b/lib/WorkflowMain.groovy index 1d88d052..3cd7c5fd 100755 --- a/lib/WorkflowMain.groovy +++ b/lib/WorkflowMain.groovy @@ -96,12 +96,12 @@ class WorkflowMain { // private static void dadareftaxonomyExistsError(params, log) { if (params.dada_ref_databases && params.dada_ref_taxonomy && !params.dada_ref_databases.containsKey(params.dada_ref_taxonomy)) { - log.error "=============================================================================\n" + + def error_string = "=============================================================================\n" + " DADA2 reference database '${params.dada_ref_taxonomy}' not found in any config files provided to the pipeline.\n" + " Currently, the available reference taxonomy keys for `--dada_ref_taxonomy` are:\n" + " ${params.dada_ref_databases.keySet().join(", ")}\n" + "===================================================================================" - System.exit(1) + Nextflow.error(error_string) } } // @@ -109,12 +109,12 @@ class WorkflowMain { // private static void qiimereftaxonomyExistsError(params, log) { if (params.qiime_ref_databases && params.qiime_ref_taxonomy && !params.qiime_ref_databases.containsKey(params.qiime_ref_taxonomy)) { - log.error "=============================================================================\n" + + def error_string = "=============================================================================\n" + " QIIME2 reference database '${params.qiime_ref_taxonomy}' not found in any config files provided to the pipeline.\n" + " Currently, the available reference taxonomy keys for `--qiime_ref_taxonomy` are:\n" + " ${params.qiime_ref_databases.keySet().join(", ")}\n" + "===================================================================================" - System.exit(1) + Nextflow.error(error_string) } } } diff --git a/subworkflows/local/cutadapt_workflow.nf b/subworkflows/local/cutadapt_workflow.nf index 0d68b097..6f280b9d 100644 --- a/subworkflows/local/cutadapt_workflow.nf +++ b/subworkflows/local/cutadapt_workflow.nf @@ -63,8 +63,7 @@ workflow CUTADAPT_WORKFLOW { if (params.ignore_failed_trimming) { log.warn "The following samples had too small file size (<1KB) after trimming with cutadapt:\n$samples\nIgnoring failed samples and continue!\n" } else { - log.error "The following samples had too small file size (<1KB) after trimming with cutadapt:\n$samples\nPlease check whether the correct primer sequences for trimming were supplied. Ignore that samples using `--ignore_failed_trimming`." - System.exit(1) + error("The following samples had too small file size (<1KB) after trimming with cutadapt:\n$samples\nPlease check whether the correct primer sequences for trimming were supplied. Ignore that samples using `--ignore_failed_trimming`.") } } diff --git a/subworkflows/local/parse_input.nf b/subworkflows/local/parse_input.nf index 11cc5f3b..640820a3 100644 --- a/subworkflows/local/parse_input.nf +++ b/subworkflows/local/parse_input.nf @@ -156,8 +156,7 @@ workflow PARSE_INPUT { if (params.ignore_empty_input_files) { log.warn "At least one input file for the following sample(s) was too small (<1KB):\n$samples\nIgnoring failed samples and continue!\n" } else { - log.error "At least one input file for the following sample(s) was too small (<1KB):\n$samples\nEither remove those samples or ignore that samples using `--ignore_empty_input_files`." - System.exit(1) + error("At least one input file for the following sample(s) was too small (<1KB):\n$samples\nEither remove those samples or ignore that samples using `--ignore_empty_input_files`.") } } } diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index c52aa646..63d16aae 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -87,8 +87,7 @@ if ( !single_end && !params.illumina_pe_its && (params.trunclenf == null || para } else { find_truncation_values = false } if ( !is_fasta_input && (!params.FW_primer || !params.RV_primer) && !params.skip_cutadapt ) { - log.error "Incompatible parameters: `--FW_primer` and `--RV_primer` are required for primer trimming. If primer trimming is not needed, use `--skip_cutadapt`." - System.exit(1) + error("Incompatible parameters: `--FW_primer` and `--RV_primer` are required for primer trimming. If primer trimming is not needed, use `--skip_cutadapt`.") } // save params to values to be able to overwrite it @@ -104,8 +103,7 @@ if ( params.dada_ref_taxonomy ) { //make sure that taxlevels adheres to requirements when mixed with addSpecies if ( params.dada_ref_taxonomy && !params.skip_dada_addspecies && !params.skip_taxonomy && taxlevels ) { if ( !taxlevels.endsWith(",Genus,Species") && !taxlevels.endsWith(",Genus") ) { - log.error "Incompatible settings: To use exact species annotations, taxonomic levels must end with `,Genus,Species` or `,Genus,Species` but are currently `${taxlevels}`. Taxonomic levels can be set with `--dada_assign_taxlevels`. Skip exact species annotations with `--skip_dada_addspecies`.\n" - System.exit(1) + error("Incompatible settings: To use exact species annotations, taxonomic levels must end with `,Genus,Species` or `,Genus,Species` but are currently `${taxlevels}`. Taxonomic levels can be set with `--dada_assign_taxlevels`. Skip exact species annotations with `--skip_dada_addspecies`.\n") } } @@ -329,8 +327,7 @@ workflow AMPLISEQ { BARRNAPSUMMARY ( BARRNAP.out.gff.collect() ) BARRNAPSUMMARY.out.warning.subscribe { if ( it.baseName.toString().startsWith("WARNING") ) { - log.error "Barrnap could not identify any rRNA in the ASV sequences! This will result in all sequences being removed with SSU filtering." - System.exit(1) + error("Barrnap could not identify any rRNA in the ASV sequences! This will result in all sequences being removed with SSU filtering.") } } ch_barrnapsummary = BARRNAPSUMMARY.out.summary From ec794cd308095348434427c12d90332611d367f2 Mon Sep 17 00:00:00 2001 From: daniel Date: Thu, 4 May 2023 13:42:38 +0200 Subject: [PATCH 075/158] replace exit 1 with error() --- subworkflows/local/parse_input.nf | 24 ++++++++++++------------ workflows/ampliseq.nf | 2 +- 2 files changed, 13 insertions(+), 13 deletions(-) diff --git a/subworkflows/local/parse_input.nf b/subworkflows/local/parse_input.nf index 640820a3..16f8ad64 100644 --- a/subworkflows/local/parse_input.nf +++ b/subworkflows/local/parse_input.nf @@ -6,15 +6,15 @@ def parse_samplesheet(LinkedHashMap row, single_end) { //Check if samplesheet contains column sampleID & forwardReads if (row.sampleID == null || row.forwardReads == null) { - exit 1, "ERROR: Please check input samplesheet -> Column 'sampleID' and 'forwardReads' are required but not detected." + error("ERROR: Please check input samplesheet -> Column 'sampleID' and 'forwardReads' are required but not detected.") } //Check if samplesheet contains a column for reverse reads if (row.reverseReads == null && !single_end) { - exit 1, "ERROR: Please check input samplesheet -> Column 'reverseReads' is missing. In case you do have only single ended reads, please specify '--single_end', '--pacbio', or '--iontorrent'." + error("ERROR: Please check input samplesheet -> Column 'reverseReads' is missing. In case you do have only single ended reads, please specify '--single_end', '--pacbio', or '--iontorrent'.") } //Check if samplesheet contains a column run and empty fields if (row.run != null && row.run == "") { - exit 1, "ERROR: Please check input samplesheet -> Column 'run' contains an empty field. Either remove column 'run' or fill each field with a value." + error("ERROR: Please check input samplesheet -> Column 'run' contains an empty field. Either remove column 'run' or fill each field with a value.") } //read meta info def meta = [:] @@ -24,13 +24,13 @@ def parse_samplesheet(LinkedHashMap row, single_end) { //read data info def array = [] if (!file(row.forwardReads).exists()) { - exit 1, "ERROR: Please check input samplesheet -> Forward read FastQ file does not exist!\n${row.forwardReads}" + error("ERROR: Please check input samplesheet -> Forward read FastQ file does not exist!\n${row.forwardReads}") } if (meta.single_end) { array = [ meta, file(row.forwardReads) ] } else { if (!file(row.reverseReads).exists()) { - exit 1, "ERROR: Please check input samplesheet -> Reverse read FastQ file does not exist!\n${row.reverseReads}" + error("ERROR: Please check input samplesheet -> Reverse read FastQ file does not exist!\n${row.reverseReads}") } array = [ meta, [ file(row.forwardReads), file(row.reverseReads) ] ] } @@ -60,7 +60,7 @@ workflow PARSE_INPUT { // extracts read files from TSV and distribute into channels Channel .fromPath(input) - .ifEmpty {exit 1, log.info "Cannot find path file ${tsvFile}"} + .ifEmpty { error("Cannot find path file ${tsvFile}") } .splitCsv(header:true, sep:'\t') .map { parse_samplesheet(it, single_end) } .set { ch_reads } @@ -79,7 +79,7 @@ workflow PARSE_INPUT { //Get files - single end Channel .fromPath( input + folders + extension ) - .ifEmpty { exit 1, "${error_message}" } + .ifEmpty { error("${error_message}") } .map { read -> def meta = [:] meta.id = read.baseName.toString().indexOf("_") != -1 ? read.baseName.toString().take(read.baseName.toString().indexOf("_")) : read.baseName @@ -91,7 +91,7 @@ workflow PARSE_INPUT { //Get files - paired end Channel .fromFilePairs( input + folders + extension, size: 2 ) - .ifEmpty { exit 1, "${error_message}" } + .ifEmpty { error("${error_message}") } .map { name, reads -> def meta = [:] meta.id = name.toString().indexOf("_") != -1 ? name.toString().take(name.toString().indexOf("_")) : name @@ -115,7 +115,7 @@ workflow PARSE_INPUT { //Stop if folder count is 1 and multiple_sequencing_runs ch_folders .count() - .subscribe { if ( it == 1 ) exit 1, "Found only one folder with read data but \"--multiple_sequencing_runs\" was specified. Please review data input." } + .subscribe { if ( it == 1 ) error("Found only one folder with read data but \"--multiple_sequencing_runs\" was specified. Please review data input.") } } } @@ -126,19 +126,19 @@ workflow PARSE_INPUT { .subscribe { if( it.size() != it.unique().size() ) { ids = it.take(10); - exit 1, "Please review data input, sample IDs are not unique! First IDs are $ids" + error("Please review data input, sample IDs are not unique! First IDs are $ids") } } //Check that no dots "." are in sampleID ch_reads .map { meta, reads -> meta.id } - .subscribe { if ( "$it".contains(".") ) exit 1, "Please review data input, sampleIDs may not contain dots, but \"$it\" does." } + .subscribe { if ( "$it".contains(".") ) error("Please review data input, sampleIDs may not contain dots, but \"$it\" does.") } //Check that sampleIDs do not start with a number when using metadata (sampleID gets X prepended by R and metadata wont match any more!) ch_reads .map { meta, reads -> meta.id } - .subscribe { if ( params.metadata && "$it"[0].isNumber() ) exit 1, "Please review data input, sampleIDs may not start with a number, but \"$it\" does. The pipeline unintentionally modifies such strings and the metadata will not match any more." } + .subscribe { if ( params.metadata && "$it"[0].isNumber() ) error("Please review data input, sampleIDs may not start with a number, but \"$it\" does. The pipeline unintentionally modifies such strings and the metadata will not match any more.") } //Filter empty files ch_reads.dump(tag:'parse_input.nf: ch_reads') diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 63d16aae..d86a87cd 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -15,7 +15,7 @@ def checkPathParamList = [ params.multiqc_config, params.metadata, params.classi for (param in checkPathParamList) { if (param) { file(param, checkIfExists: true) } } // Check mandatory parameters -if (params.input) { ch_input = file(params.input) } else { exit 1, 'Input samplesheet not specified!' } +if (params.input) { ch_input = file(params.input) } else { error('Input samplesheet not specified!') } /* ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ From 05db2b81e5ef0a2551075fced4133d13232d8d3e Mon Sep 17 00:00:00 2001 From: daniel Date: Thu, 4 May 2023 14:03:30 +0200 Subject: [PATCH 076/158] update changelog --- CHANGELOG.md | 1 + 1 file changed, 1 insertion(+) diff --git a/CHANGELOG.md b/CHANGELOG.md index 1b17dbbb..ba3b57c6 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -20,6 +20,7 @@ and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0 - [#554](https://github.com/nf-core/ampliseq/pull/554) - Accept taxonomy strings that contain `#`,`'` - [#569](https://github.com/nf-core/ampliseq/pull/569) - Make header of overall_summary.tsv consistent between input data types - [#573](https://github.com/nf-core/ampliseq/pull/573) - Avoid parser error for single-end data when an empty read file is detected +- [#578](https://github.com/nf-core/ampliseq/pull/578) - Template update for nf-core/tools version 2.8, including changing `System.exit(1)` to `Nextflow.error()` ### `Dependencies` From 0f7b7fa21f149a9b55c63b8db6a5e1c4ec789fa3 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Thu, 4 May 2023 15:00:53 +0200 Subject: [PATCH 077/158] Fix bug when sintax_ref_taxonomy is not set --- modules/local/sbdiexportreannotate.nf | 9 ++------- workflows/ampliseq.nf | 8 ++++++-- 2 files changed, 8 insertions(+), 9 deletions(-) diff --git a/modules/local/sbdiexportreannotate.nf b/modules/local/sbdiexportreannotate.nf index 7f1377ad..50125217 100644 --- a/modules/local/sbdiexportreannotate.nf +++ b/modules/local/sbdiexportreannotate.nf @@ -10,6 +10,7 @@ process SBDIEXPORTREANNOTATE { input: path taxonomytable val taxonomymethod + val dbversion path predictions output: @@ -21,19 +22,13 @@ process SBDIEXPORTREANNOTATE { script: """ - if [[ $taxonomymethod == "sintax" ]] ; then - db_version="${params.sintax_ref_databases[params.sintax_ref_taxonomy]["dbversion"]}" - else - db_version="${params.dada_ref_databases[params.dada_ref_taxonomy]["dbversion"]}" - fi - if [[ $workflow.manifest.version == *dev ]]; then ampliseq_version="v$workflow.manifest.version, revision: ${workflow.scriptId.substring(0,10)}" else ampliseq_version="v$workflow.manifest.version" fi - sbdiexportreannotate.R \"\$db_version\" $taxonomytable $taxonomymethod \"\$ampliseq_version\" $predictions + sbdiexportreannotate.R \"$dbversion\" $taxonomytable $taxonomymethod \"\$ampliseq_version\" $predictions cat <<-END_VERSIONS > versions.yml "${task.process}": diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 4c02c41e..97289c2a 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -110,6 +110,8 @@ if ( params.dada_ref_taxonomy ) { } else { taxlevels = params.dada_assign_taxlevels ? "${params.dada_assign_taxlevels}" : "" } if ( params.sintax_ref_taxonomy ) { sintax_taxlevels = params.sintax_ref_databases[params.sintax_ref_taxonomy]["taxlevels"] ?: "" +} else { + sintax_taxlevels = "" } //make sure that taxlevels adheres to requirements when mixed with addSpecies @@ -691,10 +693,12 @@ workflow AMPLISEQ { if ( params.sbdiexport ) { if ( params.sintax_ref_taxonomy ) { SBDIEXPORT ( ch_dada2_asv, ch_sintax_tax, ch_metadata ) - SBDIEXPORTREANNOTATE ( ch_sintax_tax, "sintax", ch_barrnapsummary.ifEmpty([]) ) + db_version = params.sintax_ref_databases[params.sintax_ref_taxonomy]["dbversion"] + SBDIEXPORTREANNOTATE ( ch_sintax_tax, "sintax", db_version, ch_barrnapsummary.ifEmpty([]) ) } else { SBDIEXPORT ( ch_dada2_asv, ch_dada2_tax, ch_metadata ) - SBDIEXPORTREANNOTATE ( ch_dada2_tax, "dada2", ch_barrnapsummary.ifEmpty([]) ) + db_version = params.dada_ref_databases[params.dada_ref_taxonomy]["dbversion"] + SBDIEXPORTREANNOTATE ( ch_dada2_tax, "dada2", db_version, ch_barrnapsummary.ifEmpty([]) ) } ch_versions = ch_versions.mix(SBDIEXPORT.out.versions.first()) } From 8956649375c64473759032ce8657755140f0d4d2 Mon Sep 17 00:00:00 2001 From: Daniel Straub <42973691+d4straub@users.noreply.github.com> Date: Fri, 5 May 2023 09:01:56 +0200 Subject: [PATCH 078/158] Add docker.io to modules/local/format_taxonomy_sintax.nf --- modules/local/format_taxonomy_sintax.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/modules/local/format_taxonomy_sintax.nf b/modules/local/format_taxonomy_sintax.nf index 825d4ecf..7ccfa745 100644 --- a/modules/local/format_taxonomy_sintax.nf +++ b/modules/local/format_taxonomy_sintax.nf @@ -4,7 +4,7 @@ process FORMAT_TAXONOMY_SINTAX { conda "conda-forge::sed=4.7" container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? 'https://containers.biocontainers.pro/s3/SingImgsRepo/biocontainers/v1.2.0_cv1/biocontainers_v1.2.0_cv1.img' : - 'biocontainers/biocontainers:v1.2.0_cv1' }" + 'docker.io/biocontainers/biocontainers:v1.2.0_cv1' }" input: path(database) From d155d529a097b5d4666e2ddf775936c187840aea Mon Sep 17 00:00:00 2001 From: jtangrot Date: Fri, 5 May 2023 10:47:56 +0200 Subject: [PATCH 079/158] Move sintax taxonomy to subworkflow --- subworkflows/local/sintax_taxonomy.nf | 52 +++++++++++++++++++++++++++ workflows/ampliseq.nf | 34 ++++++------------ 2 files changed, 62 insertions(+), 24 deletions(-) create mode 100644 subworkflows/local/sintax_taxonomy.nf diff --git a/subworkflows/local/sintax_taxonomy.nf b/subworkflows/local/sintax_taxonomy.nf new file mode 100644 index 00000000..1c4d1b62 --- /dev/null +++ b/subworkflows/local/sintax_taxonomy.nf @@ -0,0 +1,52 @@ +/* + * Taxonomic classification with SINTAX + */ + +include { VSEARCH_SINTAX } from '../../modules/nf-core/vsearch/sintax/main' + +include { FORMAT_TAXONOMY_SINTAX } from '../../modules/local/format_taxonomy_sintax' +include { FORMAT_TAXRESULTS_SINTAX } from '../../modules/local/format_taxresults_sintax' + +workflow SINTAX_TAXONOMY { + take: + ch_sintax_ref_taxonomy + val_sintax_ref_taxonomy + ch_fasta + ch_full_fasta + sintax_taxlevels + + main: + ch_versions_sintax_taxonomy = Channel.empty() + + //format taxonomy file + FORMAT_TAXONOMY_SINTAX ( ch_sintax_ref_taxonomy ) + ch_sintaxdb = FORMAT_TAXONOMY_SINTAX.out.db + + //set file prefix + if (params.cut_its == "none") { + ASV_tax_name = "ASV_tax_sintax.${val_sintax_ref_taxonomy}" + ASV_tax_name2 = "ASV_tax_sintax.${val_sintax_ref_taxonomy}" + } else { + ASV_tax_name = "ASV_ITS_tax_sintax.${val_sintax_ref_taxonomy}" + ASV_tax_name2 = "ASV_tax_sintax.${val_sintax_ref_taxonomy}" + } + + //search taxonomy database with SINTAX + ch_fasta + .map { + fasta -> + def meta = [:] + meta.id = ASV_tax_name + ".raw" + [ meta, fasta ] } + .set { ch_fasta_sintax } + VSEARCH_SINTAX( ch_fasta_sintax, ch_sintaxdb ) + ch_versions_sintax_taxonomy = ch_versions_sintax_taxonomy.mix(VSEARCH_SINTAX.out.versions) + + //convert SINTAX output to DADA2 like taxonomy table + FORMAT_TAXRESULTS_SINTAX( VSEARCH_SINTAX.out.tsv, ch_full_fasta, ASV_tax_name2 + '.tsv', sintax_taxlevels ) + ch_sintax_tax = FORMAT_TAXRESULTS_SINTAX.out.tsv + + emit: + tax = ch_sintax_tax + versions = ch_versions_sintax_taxonomy +} diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 291780fd..36f8ab80 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -154,7 +154,6 @@ include { MERGE_STATS as MERGE_STATS_FILTERSSU } from '../modules/local/merge include { MERGE_STATS as MERGE_STATS_FILTERLENASV } from '../modules/local/merge_stats' include { FORMAT_FASTAINPUT } from '../modules/local/format_fastainput' include { FORMAT_TAXONOMY } from '../modules/local/format_taxonomy' -include { FORMAT_TAXONOMY_SINTAX } from '../modules/local/format_taxonomy_sintax' include { ITSX_CUTASV } from '../modules/local/itsx_cutasv' include { MERGE_STATS as MERGE_STATS_STD} from '../modules/local/merge_stats' include { DADA2_TAXONOMY } from '../modules/local/dada2_taxonomy' @@ -162,7 +161,6 @@ include { DADA2_ADDSPECIES } from '../modules/local/dada2_addspecie include { ASSIGNSH } from '../modules/local/assignsh' include { FORMAT_TAXRESULTS as FORMAT_TAXRESULTS_STD } from '../modules/local/format_taxresults' include { FORMAT_TAXRESULTS as FORMAT_TAXRESULTS_ADDSP } from '../modules/local/format_taxresults' -include { FORMAT_TAXRESULTS_SINTAX } from '../modules/local/format_taxresults_sintax' include { QIIME2_INSEQ } from '../modules/local/qiime2_inseq' include { QIIME2_FILTERTAXA } from '../modules/local/qiime2_filtertaxa' include { QIIME2_INASV } from '../modules/local/qiime2_inasv' @@ -187,6 +185,7 @@ include { DADA2_PREPROCESSING } from '../subworkflows/local/dada2_prep include { QIIME2_PREPTAX } from '../subworkflows/local/qiime2_preptax' include { QIIME2_TAXONOMY } from '../subworkflows/local/qiime2_taxonomy' include { CUTADAPT_WORKFLOW } from '../subworkflows/local/cutadapt_workflow' +include { SINTAX_TAXONOMY } from '../subworkflows/local/sintax_taxonomy' include { QIIME2_EXPORT } from '../subworkflows/local/qiime2_export' include { QIIME2_BARPLOTAVG } from '../subworkflows/local/qiime2_barplotavg' include { QIIME2_DIVERSITY } from '../subworkflows/local/qiime2_diversity' @@ -206,7 +205,6 @@ include { CUTADAPT as CUTADAPT_TAXONOMY } from '../modules/nf-core/cutadapt/ include { FASTQC } from '../modules/nf-core/fastqc/main' include { MULTIQC } from '../modules/nf-core/multiqc/main' include { CUSTOM_DUMPSOFTWAREVERSIONS } from '../modules/nf-core/custom/dumpsoftwareversions/main' -include { VSEARCH_SINTAX } from '../modules/nf-core/vsearch/sintax/main' include { VSEARCH_USEARCHGLOBAL } from '../modules/nf-core/vsearch/usearchglobal/main' include { FASTA_NEWICK_EPANG_GAPPA } from '../subworkflows/nf-core/fasta_newick_epang_gappa/main' @@ -488,30 +486,17 @@ workflow AMPLISEQ { ch_dada2_tax = Channel.empty() } - // Sintax + // SINTAX // This will only run if --sintax_ref_taxonomy is defined, // i.e. if the channel ch_sintax_ref_taxonomy is not empty if (!params.skip_taxonomy) { - FORMAT_TAXONOMY_SINTAX ( ch_sintax_ref_taxonomy.collect() ) - ch_sintaxdb = FORMAT_TAXONOMY_SINTAX.out.db - if (params.cut_its == "none") { - ASV_tax_name = "ASV_tax_sintax.${val_sintax_ref_taxonomy}" - ASV_tax_name2 = "ASV_tax_sintax.${val_sintax_ref_taxonomy}" - } else { - ASV_tax_name = "ASV_ITS_tax_sintax.${val_sintax_ref_taxonomy}" - ASV_tax_name2 = "ASV_tax_sintax.${val_sintax_ref_taxonomy}" - } - ch_fasta - .map { - fasta -> - def meta = [:] - meta.id = ASV_tax_name + ".raw" - [ meta, fasta ] } - .set { ch_fasta_sintax } - VSEARCH_SINTAX( ch_fasta_sintax, ch_sintaxdb ) - ch_versions = ch_versions.mix(VSEARCH_SINTAX.out.versions) - FORMAT_TAXRESULTS_SINTAX( VSEARCH_SINTAX.out.tsv, ch_full_fasta, ASV_tax_name2 + '.tsv', sintax_taxlevels ) - ch_sintax_tax = FORMAT_TAXRESULTS_SINTAX.out.tsv + SINTAX_TAXONOMY ( + ch_sintax_ref_taxonomy.collect(), + val_sintax_ref_taxonomy, + ch_fasta, + ch_full_fasta, + sintax_taxlevels + ).tax.set { ch_sintax_tax } } else { ch_sintax_tax = Channel.empty() } @@ -552,6 +537,7 @@ workflow AMPLISEQ { ) ch_versions = ch_versions.mix( QIIME2_TAXONOMY.out.versions.ifEmpty(null) ) //usually a .first() is here, dont know why this leads here to a warning } + // // SUBWORKFLOW / MODULES : Downstream analysis with QIIME2 // From afa4f9ad832d3cbdd5672e7b00e1ab9e8c94d6b1 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Fri, 5 May 2023 11:29:24 +0200 Subject: [PATCH 080/158] Move pplace taxonomy to subworkflow --- subworkflows/local/pplace_taxonomy.nf | 36 +++++++++++++++++++++++++++ workflows/ampliseq.nf | 26 ++++++------------- 2 files changed, 43 insertions(+), 19 deletions(-) create mode 100644 subworkflows/local/pplace_taxonomy.nf diff --git a/subworkflows/local/pplace_taxonomy.nf b/subworkflows/local/pplace_taxonomy.nf new file mode 100644 index 00000000..f8ecdd31 --- /dev/null +++ b/subworkflows/local/pplace_taxonomy.nf @@ -0,0 +1,36 @@ +/* + * Phylogenetic placement and taxonomic classification + */ + +include { FASTA_NEWICK_EPANG_GAPPA } from '../../subworkflows/nf-core/fasta_newick_epang_gappa/main' + +include { FORMAT_PPLACETAX } from '../../modules/local/format_pplacetax' + +workflow PPLACE_TAXONOMY { + take: + ch_fasta + + main: + ch_versions_pplace_taxonomy = Channel.empty() + + ch_pp_data = ch_fasta.map { it -> + [ meta: [ id: params.pplace_name ?: 'user_tree' ], + data: [ + alignmethod: params.pplace_alnmethod ?: 'hmmer', + queryseqfile: it, + refseqfile: file( params.pplace_aln, checkIfExists: true ), + hmmfile: [], + refphylogeny: file( params.pplace_tree, checkIfExists: true ), + model: params.pplace_model, + taxonomy: params.pplace_taxonomy ? file( params.pplace_taxonomy, checkIfExists: true ) : [] + ] ] + } + FASTA_NEWICK_EPANG_GAPPA ( ch_pp_data ) + ch_versions_pplace_taxonomy = ch_versions_pplace_taxonomy.mix( FASTA_NEWICK_EPANG_GAPPA.out.versions ) + + ch_pplace_tax = FORMAT_PPLACETAX ( FASTA_NEWICK_EPANG_GAPPA.out.taxonomy_per_query ).tsv + + emit: + tax = ch_pplace_tax + versions = ch_versions_pplace_taxonomy +} diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 36f8ab80..fa13d9de 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -165,7 +165,6 @@ include { QIIME2_INSEQ } from '../modules/local/qiime2_inseq' include { QIIME2_FILTERTAXA } from '../modules/local/qiime2_filtertaxa' include { QIIME2_INASV } from '../modules/local/qiime2_inasv' include { QIIME2_INTREE } from '../modules/local/qiime2_intree' -include { FORMAT_PPLACETAX } from '../modules/local/format_pplacetax' include { FILTER_STATS } from '../modules/local/filter_stats' include { MERGE_STATS as MERGE_STATS_FILTERTAXA } from '../modules/local/merge_stats' include { QIIME2_BARPLOT } from '../modules/local/qiime2_barplot' @@ -186,6 +185,7 @@ include { QIIME2_PREPTAX } from '../subworkflows/local/qiime2_pre include { QIIME2_TAXONOMY } from '../subworkflows/local/qiime2_taxonomy' include { CUTADAPT_WORKFLOW } from '../subworkflows/local/cutadapt_workflow' include { SINTAX_TAXONOMY } from '../subworkflows/local/sintax_taxonomy' +include { PPLACE_TAXONOMY } from '../subworkflows/local/pplace_taxonomy' include { QIIME2_EXPORT } from '../subworkflows/local/qiime2_export' include { QIIME2_BARPLOTAVG } from '../subworkflows/local/qiime2_barplotavg' include { QIIME2_DIVERSITY } from '../subworkflows/local/qiime2_diversity' @@ -206,7 +206,6 @@ include { FASTQC } from '../modules/nf-core/fastqc/ma include { MULTIQC } from '../modules/nf-core/multiqc/main' include { CUSTOM_DUMPSOFTWAREVERSIONS } from '../modules/nf-core/custom/dumpsoftwareversions/main' include { VSEARCH_USEARCHGLOBAL } from '../modules/nf-core/vsearch/usearchglobal/main' -include { FASTA_NEWICK_EPANG_GAPPA } from '../subworkflows/nf-core/fasta_newick_epang_gappa/main' /* @@ -497,29 +496,18 @@ workflow AMPLISEQ { ch_full_fasta, sintax_taxlevels ).tax.set { ch_sintax_tax } + ch_versions = ch_versions.mix(SINTAX_TAXONOMY.out.versions) } else { ch_sintax_tax = Channel.empty() } // Phylo placement if ( params.pplace_tree ) { - ch_pp_data = ch_fasta.map { it -> - [ meta: [ id: params.pplace_name ?: 'user_tree' ], - data: [ - alignmethod: params.pplace_alnmethod ?: 'hmmer', - queryseqfile: it, - refseqfile: file( params.pplace_aln, checkIfExists: true ), - hmmfile: [], - refphylogeny: file( params.pplace_tree, checkIfExists: true ), - model: params.pplace_model, - taxonomy: params.pplace_taxonomy ? file( params.pplace_taxonomy, checkIfExists: true ) : [] - ] ] - } - FASTA_NEWICK_EPANG_GAPPA ( ch_pp_data ) - ch_versions = ch_versions.mix( FASTA_NEWICK_EPANG_GAPPA.out.versions ) - - ch_pplace_tax = FORMAT_PPLACETAX ( FASTA_NEWICK_EPANG_GAPPA.out.taxonomy_per_query ).tsv - } else { ch_pplace_tax = Channel.empty() } + ch_pplace_tax = PPLACE_TAXONOMY ( ch_fasta ).tax + ch_versions = ch_versions.mix(PPLACE_TAXONOMY.out.versions) + } else { + ch_pplace_tax = Channel.empty() + } //QIIME2 if ( run_qiime2 ) { From 6a6310ecce5dd1743a51a62e5c54df6ee2293956 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Fri, 5 May 2023 15:03:56 +0200 Subject: [PATCH 081/158] Move dada2 taxonomy to subworkflow --- subworkflows/local/dada2_taxonomy_wf.nf | 118 ++++++++++++++++++ ...lace_taxonomy.nf => pplace_taxonomy_wf.nf} | 2 +- ...ntax_taxonomy.nf => sintax_taxonomy_wf.nf} | 2 +- workflows/ampliseq.nf | 108 +++------------- 4 files changed, 136 insertions(+), 94 deletions(-) create mode 100644 subworkflows/local/dada2_taxonomy_wf.nf rename subworkflows/local/{pplace_taxonomy.nf => pplace_taxonomy_wf.nf} (97%) rename subworkflows/local/{sintax_taxonomy.nf => sintax_taxonomy_wf.nf} (98%) diff --git a/subworkflows/local/dada2_taxonomy_wf.nf b/subworkflows/local/dada2_taxonomy_wf.nf new file mode 100644 index 00000000..c26761b1 --- /dev/null +++ b/subworkflows/local/dada2_taxonomy_wf.nf @@ -0,0 +1,118 @@ +/* + * Taxonomic classification with DADA2 + */ + +include { CUTADAPT as CUTADAPT_TAXONOMY } from '../../modules/nf-core/cutadapt/main' +include { VSEARCH_USEARCHGLOBAL } from '../../modules/nf-core/vsearch/usearchglobal/main' + +include { DADA2_TAXONOMY } from '../../modules/local/dada2_taxonomy' +include { DADA2_ADDSPECIES } from '../../modules/local/dada2_addspecies' +include { FORMAT_TAXRESULTS as FORMAT_TAXRESULTS_STD } from '../../modules/local/format_taxresults' +include { FORMAT_TAXRESULTS as FORMAT_TAXRESULTS_ADDSP } from '../../modules/local/format_taxresults' +include { ASSIGNSH } from '../../modules/local/assignsh' + + +workflow DADA2_TAXONOMY_WF { + take: + ch_assigntax + ch_addspecies + val_dada_ref_taxonomy + ch_fasta + ch_full_fasta + taxlevels + + main: + ch_versions_dada_taxonomy = Channel.empty() + + // Set cutoff to use for SH assignment and path to SH taxonomy file + if ( params.addsh ) { + vsearch_cutoff = 0.985 + ch_shinfo = Channel.fromList(params.dada_ref_databases[params.dada_ref_taxonomy]["shfile"]).map { file(it) } + } + + //cut taxonomy to expected amplicon + if (params.cut_dada_ref_taxonomy) { + ch_assigntax + .map { + db -> + def meta = [:] + meta.single_end = true + meta.id = "assignTaxonomy" + [ meta, db ] } + .set { ch_assigntax } + CUTADAPT_TAXONOMY ( ch_assigntax ).reads + .map { meta, db -> db } + .set { ch_assigntax } + } + + //set file name prefix + if (params.cut_its == "none") { + ASV_tax_name = "ASV_tax" + } else { + ASV_tax_name = "ASV_ITS_tax" + } + + //DADA2 assignTaxonomy + DADA2_TAXONOMY ( ch_fasta, ch_assigntax, ASV_tax_name + ".${val_dada_ref_taxonomy}.tsv", taxlevels ) + ch_versions_dada_taxonomy = ch_versions_dada_taxonomy.mix(DADA2_TAXONOMY.out.versions) + if (params.cut_its != "none") { + FORMAT_TAXRESULTS_STD ( DADA2_TAXONOMY.out.tsv, ch_full_fasta, "ASV_tax.${val_dada_ref_taxonomy}.tsv" ) + ch_versions_dada_taxonomy = ch_versions_dada_taxonomy.mix( FORMAT_TAXRESULTS_STD.out.versions.ifEmpty(null) ) + } + + //DADA2 addSpecies + if (!params.skip_dada_addspecies) { + DADA2_ADDSPECIES ( DADA2_TAXONOMY.out.rds, ch_addspecies, ASV_tax_name + "_species.${val_dada_ref_taxonomy}.tsv", taxlevels ) + if (params.cut_its == "none") { + ch_dada2_addsp = DADA2_ADDSPECIES.out.tsv + } else { + FORMAT_TAXRESULTS_ADDSP ( DADA2_ADDSPECIES.out.tsv, ch_full_fasta, "ASV_tax_species.${val_dada_ref_taxonomy}.tsv" ) + ch_dada2_addsp = FORMAT_TAXRESULTS_ADDSP.out.tsv + } + //if addsh set: add SH assignments + if ( params.addsh ) { + ch_fasta + .map { + fasta -> + def meta = [:] + meta.id = ASV_tax_name + ".vsearch" + [ meta, fasta ] } + .set { ch_fasta_map } + VSEARCH_USEARCHGLOBAL( ch_fasta_map, ch_assigntax, vsearch_cutoff, 'blast6out', "" ) + ch_versions_dada_taxonomy = ch_versions_dada_taxonomy.mix(VSEARCH_USEARCHGLOBAL.out.versions.ifEmpty(null)) + ASSIGNSH( ch_dada2_addsp, ch_shinfo.collect(), VSEARCH_USEARCHGLOBAL.out.txt, "ASV_tax_species_SH.${val_dada_ref_taxonomy}.tsv") + ch_versions_dada_taxonomy = ch_versions_dada_taxonomy.mix(ASSIGNSH.out.versions.ifEmpty(null)) + ch_dada2_tax = ASSIGNSH.out.tsv + } else { + ch_dada2_tax = ch_dada2_addsp + } + //no DADA2 addSpecies, use results from assignTaxonomy: + } else { + if (params.cut_its == "none") { + ch_dada2_tax1 = DADA2_TAXONOMY.out.tsv + } else { + ch_dada2_tax1 = FORMAT_TAXRESULTS_STD.out.tsv + } + //if addsh set: add SH assignments + if ( params.addsh ) { + ch_fasta + .map { + fasta -> + def meta = [:] + meta.id = ASV_tax_name + ".vsearch" + [ meta, fasta ] } + .set { ch_fasta_map } + VSEARCH_USEARCHGLOBAL( ch_fasta_map, ch_assigntax, vsearch_cutoff, 'blast6out', "" ) + ch_versions_dada_taxonomy = ch_versions_dada_taxonomy.mix(VSEARCH_USEARCHGLOBAL.out.versions.ifEmpty(null)) + ASSIGNSH( ch_dada2_tax1.out.tsv, ch_shinfo.collect(), VSEARCH_USEARCHGLOBAL.out.txt, "ASV_tax_SH.${val_dada_ref_taxonomy}.tsv") + ch_versions_dada_taxonomy = ch_versions_dada_taxonomy.mix(ASSIGNSH.out.versions.ifEmpty(null)) + ch_dada2_tax = ASSIGNSH.out.tsv + } else { + ch_dada2_tax = ch_dada2_tax1 + } + } + + emit: + tax = ch_dada2_tax + versions = ch_versions_dada_taxonomy +} diff --git a/subworkflows/local/pplace_taxonomy.nf b/subworkflows/local/pplace_taxonomy_wf.nf similarity index 97% rename from subworkflows/local/pplace_taxonomy.nf rename to subworkflows/local/pplace_taxonomy_wf.nf index f8ecdd31..8a5f24de 100644 --- a/subworkflows/local/pplace_taxonomy.nf +++ b/subworkflows/local/pplace_taxonomy_wf.nf @@ -6,7 +6,7 @@ include { FASTA_NEWICK_EPANG_GAPPA } from '../../subworkflows/nf-core/fasta include { FORMAT_PPLACETAX } from '../../modules/local/format_pplacetax' -workflow PPLACE_TAXONOMY { +workflow PPLACE_TAXONOMY_WF { take: ch_fasta diff --git a/subworkflows/local/sintax_taxonomy.nf b/subworkflows/local/sintax_taxonomy_wf.nf similarity index 98% rename from subworkflows/local/sintax_taxonomy.nf rename to subworkflows/local/sintax_taxonomy_wf.nf index 1c4d1b62..99bbf971 100644 --- a/subworkflows/local/sintax_taxonomy.nf +++ b/subworkflows/local/sintax_taxonomy_wf.nf @@ -7,7 +7,7 @@ include { VSEARCH_SINTAX } from '../../modules/nf-core/vsearch/si include { FORMAT_TAXONOMY_SINTAX } from '../../modules/local/format_taxonomy_sintax' include { FORMAT_TAXRESULTS_SINTAX } from '../../modules/local/format_taxresults_sintax' -workflow SINTAX_TAXONOMY { +workflow SINTAX_TAXONOMY_WF { take: ch_sintax_ref_taxonomy val_sintax_ref_taxonomy diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index fa13d9de..268e9190 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -56,9 +56,6 @@ if (params.dada_ref_tax_custom) { //standard ref taxonomy input from params.dada_ref_taxonomy & conf/ref_databases.config ch_dada_ref_taxonomy = Channel.fromList(params.dada_ref_databases[params.dada_ref_taxonomy]["file"]).map { file(it) } val_dada_ref_taxonomy = params.dada_ref_taxonomy.replace('=','_').replace('.','_') - if (params.addsh) { - ch_shinfo = Channel.fromList(params.dada_ref_databases[params.dada_ref_taxonomy]["shfile"]).map { file(it) } - } } else { ch_dada_ref_taxonomy = Channel.empty() val_dada_ref_taxonomy = "none" @@ -128,10 +125,6 @@ if ( !(workflow.profile.tokenize(',').intersect(['conda', 'mamba']).size() >= 1) if ( workflow.profile.tokenize(',').intersect(['conda', 'mamba']).size() >= 1 ) { log.warn "Conda or mamba is enabled, any steps involving QIIME2 are not available. Use a container engine instead of conda to enable all software." } } -// Set cutoff to use for SH assignment -if ( params.addsh ) { - vsearch_cutoff = 0.985 -} /* ======================================================================================== @@ -156,11 +149,6 @@ include { FORMAT_FASTAINPUT } from '../modules/local/format_fastainp include { FORMAT_TAXONOMY } from '../modules/local/format_taxonomy' include { ITSX_CUTASV } from '../modules/local/itsx_cutasv' include { MERGE_STATS as MERGE_STATS_STD} from '../modules/local/merge_stats' -include { DADA2_TAXONOMY } from '../modules/local/dada2_taxonomy' -include { DADA2_ADDSPECIES } from '../modules/local/dada2_addspecies' -include { ASSIGNSH } from '../modules/local/assignsh' -include { FORMAT_TAXRESULTS as FORMAT_TAXRESULTS_STD } from '../modules/local/format_taxresults' -include { FORMAT_TAXRESULTS as FORMAT_TAXRESULTS_ADDSP } from '../modules/local/format_taxresults' include { QIIME2_INSEQ } from '../modules/local/qiime2_inseq' include { QIIME2_FILTERTAXA } from '../modules/local/qiime2_filtertaxa' include { QIIME2_INASV } from '../modules/local/qiime2_inasv' @@ -184,8 +172,9 @@ include { DADA2_PREPROCESSING } from '../subworkflows/local/dada2_prep include { QIIME2_PREPTAX } from '../subworkflows/local/qiime2_preptax' include { QIIME2_TAXONOMY } from '../subworkflows/local/qiime2_taxonomy' include { CUTADAPT_WORKFLOW } from '../subworkflows/local/cutadapt_workflow' -include { SINTAX_TAXONOMY } from '../subworkflows/local/sintax_taxonomy' -include { PPLACE_TAXONOMY } from '../subworkflows/local/pplace_taxonomy' +include { DADA2_TAXONOMY_WF } from '../subworkflows/local/dada2_taxonomy_wf' +include { SINTAX_TAXONOMY_WF } from '../subworkflows/local/sintax_taxonomy_wf' +include { PPLACE_TAXONOMY_WF } from '../subworkflows/local/pplace_taxonomy_wf' include { QIIME2_EXPORT } from '../subworkflows/local/qiime2_export' include { QIIME2_BARPLOTAVG } from '../subworkflows/local/qiime2_barplotavg' include { QIIME2_DIVERSITY } from '../subworkflows/local/qiime2_diversity' @@ -201,11 +190,9 @@ include { QIIME2_ANCOM } from '../subworkflows/local/qiime2_anc // MODULE: Installed directly from nf-core/modules // -include { CUTADAPT as CUTADAPT_TAXONOMY } from '../modules/nf-core/cutadapt/main' include { FASTQC } from '../modules/nf-core/fastqc/main' include { MULTIQC } from '../modules/nf-core/multiqc/main' include { CUSTOM_DUMPSOFTWAREVERSIONS } from '../modules/nf-core/custom/dumpsoftwareversions/main' -include { VSEARCH_USEARCHGLOBAL } from '../modules/nf-core/vsearch/usearchglobal/main' /* @@ -409,78 +396,15 @@ workflow AMPLISEQ { ch_assigntax = FORMAT_TAXONOMY.out.assigntax ch_addspecies = FORMAT_TAXONOMY.out.addspecies } - //Cut taxonomy to expected amplicon - if (params.cut_dada_ref_taxonomy) { - ch_assigntax - .map { - db -> - def meta = [:] - meta.single_end = true - meta.id = "assignTaxonomy" - [ meta, db ] } - .set { ch_assigntax } - CUTADAPT_TAXONOMY ( ch_assigntax ).reads - .map { meta, db -> db } - .set { ch_assigntax } - } - if (params.cut_its == "none") { - ASV_tax_name = "ASV_tax" - } else { - ASV_tax_name = "ASV_ITS_tax" - } - DADA2_TAXONOMY ( ch_fasta, ch_assigntax, ASV_tax_name + ".${val_dada_ref_taxonomy}.tsv", taxlevels ) - ch_versions = ch_versions.mix(DADA2_TAXONOMY.out.versions) - if (params.cut_its != "none") { - FORMAT_TAXRESULTS_STD ( DADA2_TAXONOMY.out.tsv, ch_full_fasta, "ASV_tax.${val_dada_ref_taxonomy}.tsv" ) - ch_versions = ch_versions.mix( FORMAT_TAXRESULTS_STD.out.versions.ifEmpty(null) ) - } - if (!params.skip_dada_addspecies) { - DADA2_ADDSPECIES ( DADA2_TAXONOMY.out.rds, ch_addspecies, ASV_tax_name + "_species.${val_dada_ref_taxonomy}.tsv", taxlevels ) - if (params.cut_its == "none") { - ch_dada2_addsp = DADA2_ADDSPECIES.out.tsv - } else { - FORMAT_TAXRESULTS_ADDSP ( DADA2_ADDSPECIES.out.tsv, ch_fasta, "ASV_tax_species.${val_dada_ref_taxonomy}.tsv" ) - ch_dada2_addsp = FORMAT_TAXRESULTS_ADDSP.out.tsv - } - if ( params.addsh ) { - ch_fasta - .map { - fasta -> - def meta = [:] - meta.id = ASV_tax_name + ".vsearch" - [ meta, fasta ] } - .set { ch_fasta_map } - VSEARCH_USEARCHGLOBAL( ch_fasta_map, ch_assigntax, vsearch_cutoff, 'blast6out', "" ) - ch_versions = ch_versions.mix(VSEARCH_USEARCHGLOBAL.out.versions.ifEmpty(null)) - ASSIGNSH( ch_dada2_addsp, ch_shinfo.collect(), VSEARCH_USEARCHGLOBAL.out.txt, "ASV_tax_species_SH.${val_dada_ref_taxonomy}.tsv") - ch_versions = ch_versions.mix(ASSIGNSH.out.versions.ifEmpty(null)) - ch_dada2_tax = ASSIGNSH.out.tsv - } else { - ch_dada2_tax = ch_dada2_addsp - } - } else { - if (params.cut_its == "none") { - ch_dada2_tax1 = DADA2_TAXONOMY.out.tsv - } else { - ch_dada2_tax1 = FORMAT_TAXRESULTS_STD.out.tsv - } - if ( params.addsh ) { - ch_fasta - .map { - fasta -> - def meta = [:] - meta.id = ASV_tax_name + ".vsearch" - [ meta, fasta ] } - .set { ch_fasta_map } - VSEARCH_USEARCHGLOBAL( ch_fasta_map, ch_assigntax, vsearch_cutoff, 'blast6out', "" ) - ch_versions = ch_versions.mix(VSEARCH_USEARCHGLOBAL.out.versions.ifEmpty(null)) - ASSIGNSH( ch_dada2_tax1.out.tsv, ch_shinfo.collect(), VSEARCH_USEARCHGLOBAL.out.txt, "ASV_tax_SH.${val_dada_ref_taxonomy}.tsv") - ch_versions = ch_versions.mix(ASSIGNSH.out.versions.ifEmpty(null)) - ch_dada2_tax = ASSIGNSH.out.tsv - } else { - ch_dada2_tax = ch_dada2_tax1 - } - } + DADA2_TAXONOMY_WF ( + ch_assigntax, + ch_addspecies, + val_dada_ref_taxonomy, + ch_fasta, + ch_full_fasta, + taxlevels + ).tax.set { ch_dada2_tax } + ch_versions = ch_versions.mix(DADA2_TAXONOMY_WF.out.versions) } else { ch_dada2_tax = Channel.empty() } @@ -489,22 +413,22 @@ workflow AMPLISEQ { // This will only run if --sintax_ref_taxonomy is defined, // i.e. if the channel ch_sintax_ref_taxonomy is not empty if (!params.skip_taxonomy) { - SINTAX_TAXONOMY ( + SINTAX_TAXONOMY_WF ( ch_sintax_ref_taxonomy.collect(), val_sintax_ref_taxonomy, ch_fasta, ch_full_fasta, sintax_taxlevels ).tax.set { ch_sintax_tax } - ch_versions = ch_versions.mix(SINTAX_TAXONOMY.out.versions) + ch_versions = ch_versions.mix(SINTAX_TAXONOMY_WF.out.versions) } else { ch_sintax_tax = Channel.empty() } // Phylo placement if ( params.pplace_tree ) { - ch_pplace_tax = PPLACE_TAXONOMY ( ch_fasta ).tax - ch_versions = ch_versions.mix(PPLACE_TAXONOMY.out.versions) + ch_pplace_tax = PPLACE_TAXONOMY_WF ( ch_fasta ).tax + ch_versions = ch_versions.mix(PPLACE_TAXONOMY_WF.out.versions) } else { ch_pplace_tax = Channel.empty() } From b34a18704f86630819a379c6bff5032f2cf41045 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Mon, 8 May 2023 09:31:37 +0200 Subject: [PATCH 082/158] Fix docs --- docs/output.md | 44 ++++++++++++++++++++++++++------------------ 1 file changed, 26 insertions(+), 18 deletions(-) diff --git a/docs/output.md b/docs/output.md index 77f8bbd2..7b4aa5b0 100644 --- a/docs/output.md +++ b/docs/output.md @@ -1,3 +1,9 @@ +--- +output: + pdf_document: default + html_document: default +--- + # nf-core/ampliseq: Output ## Introduction @@ -20,11 +26,13 @@ The pipeline is built using [Nextflow](https://www.nextflow.io/) and processes d - [Barrnap](#barrnap) - Predict ribosomal RNA sequences and optional filtering - [Length filter](#length-filter) - Optionally, ASV can be filtered by length thresholds - [ITSx](#itsx) - Optionally, the ITS region can be extracted -- [Taxonomic classification with DADA2](#taxonomic-classification-with-dada2) - Taxonomic classification of (filtered) ASVs - - [assignSH](#assignsh) - Optionally, a UNITE species hypothesis (SH) can be added to the taxonomy +- [Taxonomic classification](#taxonomic-classification) - Taxonomic classification of (filtered) ASVs + - [DADA2](#dada2) - Taxonomic classification with DADA2 + - [assignSH](#assignsh) - Optionally, a UNITE species hypothesis (SH) can be added to the DADA2 taxonomy + - [SINTAX](#sintax) - Taxonomic classification with SINTAX + - [Taxonomic classification with QIIME2](#taxonomic-classification-with-qiime2) - Taxonomic classification with QIIME2 - [Phlogenetic placement and taxonomic classification](#phylogenetic-placement-and-taxonomic-classification) - Placing ASVs into a phyloenetic tree - [QIIME2](#qiime2) - Secondary analysis - - [Taxonomic classification](#taxonomic-classification) - Taxonomical classification of ASVs - [Abundance tables](#abundance-tables) - Exported abundance tables - [Relative abundance tables](#relative-abundance-tables) - Exported relative abundance tables - [Barplot](#barplot) - Interactive barplot @@ -189,7 +197,7 @@ Optionally, the ITS region can be extracted from each ASV sequence using ITSx, a ### Taxonomic classification -DADA2 and/or SINTAX can be used to taxonomically classify the ASVs using a choice of supplied databases (specified with `--dada_ref_taxonomy` and/or `sintax_ref_taxonomy`). By default, DADA2 is used for the classification. The taxonomic classification will be done based on filtered ASV sequences (see above). +DADA2 and/or SINTAX can be used to taxonomically classify the ASVs using a choice of supplied databases (specified with `--dada_ref_taxonomy` and/or `--sintax_ref_taxonomy`). By default, DADA2 is used for the classification. The taxonomic classification will be done based on filtered ASV sequences (see above). #### DADA2 @@ -262,6 +270,20 @@ Files when using ITSx:
+#### Taxonomic classification with QIIME2 + +Taxonomic classification with QIIME2 is typically similar to DADA2 classifications. However, both options are available. When taxonomic classification with DADA2 and QIIME2 is performed, DADA2 classification takes precedence over QIIME2 classifications for all downstream analysis. Taxonomic classification by SINTAX or phylogenetic placement superseeds DADA2 and QIIME2 classification. + +
+Output files + +- `qiime2/taxonomy/` + - `taxonomy.tsv`: Tab-separated table with taxonomic classification for each ASV + - `*-classifier.qza`: QIIME2 artefact of the trained classifier. Can be supplied to other pipeline runs with `--classifier` + - `ref_taxonomy.txt`: Information about the used reference taxonomy, such as title, version, citation. + +
+ ### Phylogenetic placement and taxonomic classification Phylogenetic placement grafts sequences onto a phylogenetic reference tree and optionally outputs taxonomic annotations. The reference tree is ideally made from full-length high-quality sequences containing better evolutionary signal than short amplicons. It is hence superior to estimating de-novo phylogenetic trees from short amplicon sequences. On providing required reference files, ASV sequences are aligned to the reference alignment with either [HMMER](http://hmmer.org/) (default) or [MAFFT](https://mafft.cbrc.jp/alignment/software/). Subsequently, phylogenetic placement of query sequences is performed with [EPA-NG](https://github.com/Pbdas/epa-ng), and finally a number of summary operations are performed with [Gappa](https://github.com/lczech/gappa). This uses code from [nf-core/phyloplace](https://nf-co.re/phyloplace) in the form of its main [subworkflow](https://github.com/nf-core/modules/tree/master/subworkflows/nf-core/fasta_newick_epang_gappa), therefore its detailed documentation also applies here. @@ -299,20 +321,6 @@ Intermediate data imported to QIIME2 is saved as QIIME2 fragments, that can be c -#### Taxonomic classification - -Taxonomic classification with QIIME2 is typically similar to DADA2 classifications. However, both options are available. When taxonomic classification with DADA2 and QIIME2 is performed, DADA2 classification takes precedence over QIIME2 classifications for all downstream analysis. Taxonomic classification by phylogenetic placement superseeds DADA2 and QIIME2 classification. - -
-Output files - -- `qiime2/taxonomy/` - - `taxonomy.tsv`: Tab-separated table with taxonomic classification for each ASV - - `*-classifier.qza`: QIIME2 artefact of the trained classifier. Can be supplied to other pipeline runs with `--classifier` - - `ref_taxonomy.txt`: Information about the used reference taxonomy, such as title, version, citation. - -
- #### Abundance tables The abundance tables are the final data for further downstream analysis and visualisations. The tables are based on the computed ASVs and taxonomic classification (in the following priotity: phylogenetic placement [EPA-NG, Gappa], DADA2, QIIME2), but after removal of unwanted taxa. Unwanted taxa are often off-targets generated in PCR with primers that are not perfectly specific for the target DNA (can be specified by `--exclude_taxa`), by default mitrochondria and chloroplast sequences are removed because these are frequent unwanted non-bacteria PCR products. From 528516e4d63cc13db3c5a5ab716063caae0db92b Mon Sep 17 00:00:00 2001 From: jtangrot Date: Mon, 8 May 2023 13:09:34 +0200 Subject: [PATCH 083/158] Fix bug in sintax results conversion --- bin/convert_sintax_output.py | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/bin/convert_sintax_output.py b/bin/convert_sintax_output.py index 44557245..1cbd8073 100755 --- a/bin/convert_sintax_output.py +++ b/bin/convert_sintax_output.py @@ -116,9 +116,9 @@ def complete_list(some_list, target_len): tmp_list = line.split("\t") sequence = seqs[tmp_list[0]] if tmp_list[0] in seqs else "" if tmp_list[3] != "": - annot = re.sub(".\:", "", tmp_list[3]) + annot = re.sub("[dkpcofgs]\:", "", tmp_list[3]) annot_list = annot.split(",") - raw_annot = re.sub(".\:", "", tmp_list[1]) + raw_annot = re.sub("[dkpcofgs]\:", "", tmp_list[1]) raw_annot_list = raw_annot.split(",") confidence = raw_annot_list[len(annot_list) - 1].split("(")[1] confidence = confidence[:-1] From 14229f3bf4f8ac2ed9a208582e332889e193e03d Mon Sep 17 00:00:00 2001 From: jtangrot Date: Tue, 9 May 2023 13:33:11 +0200 Subject: [PATCH 084/158] Tidy up dada2_taxonomy workflow --- subworkflows/local/dada2_taxonomy_wf.nf | 62 +++++++++++-------------- 1 file changed, 27 insertions(+), 35 deletions(-) diff --git a/subworkflows/local/dada2_taxonomy_wf.nf b/subworkflows/local/dada2_taxonomy_wf.nf index c26761b1..e7caf74a 100644 --- a/subworkflows/local/dada2_taxonomy_wf.nf +++ b/subworkflows/local/dada2_taxonomy_wf.nf @@ -64,27 +64,10 @@ workflow DADA2_TAXONOMY_WF { if (!params.skip_dada_addspecies) { DADA2_ADDSPECIES ( DADA2_TAXONOMY.out.rds, ch_addspecies, ASV_tax_name + "_species.${val_dada_ref_taxonomy}.tsv", taxlevels ) if (params.cut_its == "none") { - ch_dada2_addsp = DADA2_ADDSPECIES.out.tsv + ch_dada2_tax1 = DADA2_ADDSPECIES.out.tsv } else { FORMAT_TAXRESULTS_ADDSP ( DADA2_ADDSPECIES.out.tsv, ch_full_fasta, "ASV_tax_species.${val_dada_ref_taxonomy}.tsv" ) - ch_dada2_addsp = FORMAT_TAXRESULTS_ADDSP.out.tsv - } - //if addsh set: add SH assignments - if ( params.addsh ) { - ch_fasta - .map { - fasta -> - def meta = [:] - meta.id = ASV_tax_name + ".vsearch" - [ meta, fasta ] } - .set { ch_fasta_map } - VSEARCH_USEARCHGLOBAL( ch_fasta_map, ch_assigntax, vsearch_cutoff, 'blast6out', "" ) - ch_versions_dada_taxonomy = ch_versions_dada_taxonomy.mix(VSEARCH_USEARCHGLOBAL.out.versions.ifEmpty(null)) - ASSIGNSH( ch_dada2_addsp, ch_shinfo.collect(), VSEARCH_USEARCHGLOBAL.out.txt, "ASV_tax_species_SH.${val_dada_ref_taxonomy}.tsv") - ch_versions_dada_taxonomy = ch_versions_dada_taxonomy.mix(ASSIGNSH.out.versions.ifEmpty(null)) - ch_dada2_tax = ASSIGNSH.out.tsv - } else { - ch_dada2_tax = ch_dada2_addsp + ch_dada2_tax1 = FORMAT_TAXRESULTS_ADDSP.out.tsv } //no DADA2 addSpecies, use results from assignTaxonomy: } else { @@ -93,25 +76,34 @@ workflow DADA2_TAXONOMY_WF { } else { ch_dada2_tax1 = FORMAT_TAXRESULTS_STD.out.tsv } - //if addsh set: add SH assignments - if ( params.addsh ) { - ch_fasta - .map { - fasta -> - def meta = [:] - meta.id = ASV_tax_name + ".vsearch" - [ meta, fasta ] } - .set { ch_fasta_map } - VSEARCH_USEARCHGLOBAL( ch_fasta_map, ch_assigntax, vsearch_cutoff, 'blast6out', "" ) - ch_versions_dada_taxonomy = ch_versions_dada_taxonomy.mix(VSEARCH_USEARCHGLOBAL.out.versions.ifEmpty(null)) - ASSIGNSH( ch_dada2_tax1.out.tsv, ch_shinfo.collect(), VSEARCH_USEARCHGLOBAL.out.txt, "ASV_tax_SH.${val_dada_ref_taxonomy}.tsv") - ch_versions_dada_taxonomy = ch_versions_dada_taxonomy.mix(ASSIGNSH.out.versions.ifEmpty(null)) - ch_dada2_tax = ASSIGNSH.out.tsv - } else { - ch_dada2_tax = ch_dada2_tax1 + } + + //if addsh set: add SH assignments + if ( params.addsh ) { + //set file name prefix for SH assignments + if (!params.skip_dada_addspecies) { + ASV_SH_name = "ASV_tax_species_SH" + } else { + ASV_SH_name = "ASV_tax_SH" } + //find SHs + ch_fasta + .map { + fasta -> + def meta = [:] + meta.id = ASV_tax_name + ".vsearch" + [ meta, fasta ] } + .set { ch_fasta_map } + VSEARCH_USEARCHGLOBAL( ch_fasta_map, ch_assigntax, vsearch_cutoff, 'blast6out', "" ) + ch_versions_dada_taxonomy = ch_versions_dada_taxonomy.mix(VSEARCH_USEARCHGLOBAL.out.versions.ifEmpty(null)) + ASSIGNSH( ch_dada2_tax1, ch_shinfo.collect(), VSEARCH_USEARCHGLOBAL.out.txt, ASV_SH_name + ".${val_dada_ref_taxonomy}.tsv") + ch_versions_dada_taxonomy = ch_versions_dada_taxonomy.mix(ASSIGNSH.out.versions.ifEmpty(null)) + ch_dada2_tax = ASSIGNSH.out.tsv + } else { + ch_dada2_tax = ch_dada2_tax1 } + emit: tax = ch_dada2_tax versions = ch_versions_dada_taxonomy From 8a67f42c69243e3e2e539e3a8469b7fa89caca33 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Tue, 9 May 2023 13:59:48 +0200 Subject: [PATCH 085/158] Remove subworkflow for pplace --- subworkflows/local/dada2_taxonomy_wf.nf | 4 +-- subworkflows/local/pplace_taxonomy_wf.nf | 36 ------------------------ workflows/ampliseq.nf | 21 ++++++++++++-- 3 files changed, 20 insertions(+), 41 deletions(-) delete mode 100644 subworkflows/local/pplace_taxonomy_wf.nf diff --git a/subworkflows/local/dada2_taxonomy_wf.nf b/subworkflows/local/dada2_taxonomy_wf.nf index e7caf74a..8a1f11e6 100644 --- a/subworkflows/local/dada2_taxonomy_wf.nf +++ b/subworkflows/local/dada2_taxonomy_wf.nf @@ -29,7 +29,7 @@ workflow DADA2_TAXONOMY_WF { vsearch_cutoff = 0.985 ch_shinfo = Channel.fromList(params.dada_ref_databases[params.dada_ref_taxonomy]["shfile"]).map { file(it) } } - + //cut taxonomy to expected amplicon if (params.cut_dada_ref_taxonomy) { ch_assigntax @@ -51,7 +51,7 @@ workflow DADA2_TAXONOMY_WF { } else { ASV_tax_name = "ASV_ITS_tax" } - + //DADA2 assignTaxonomy DADA2_TAXONOMY ( ch_fasta, ch_assigntax, ASV_tax_name + ".${val_dada_ref_taxonomy}.tsv", taxlevels ) ch_versions_dada_taxonomy = ch_versions_dada_taxonomy.mix(DADA2_TAXONOMY.out.versions) diff --git a/subworkflows/local/pplace_taxonomy_wf.nf b/subworkflows/local/pplace_taxonomy_wf.nf deleted file mode 100644 index 8a5f24de..00000000 --- a/subworkflows/local/pplace_taxonomy_wf.nf +++ /dev/null @@ -1,36 +0,0 @@ -/* - * Phylogenetic placement and taxonomic classification - */ - -include { FASTA_NEWICK_EPANG_GAPPA } from '../../subworkflows/nf-core/fasta_newick_epang_gappa/main' - -include { FORMAT_PPLACETAX } from '../../modules/local/format_pplacetax' - -workflow PPLACE_TAXONOMY_WF { - take: - ch_fasta - - main: - ch_versions_pplace_taxonomy = Channel.empty() - - ch_pp_data = ch_fasta.map { it -> - [ meta: [ id: params.pplace_name ?: 'user_tree' ], - data: [ - alignmethod: params.pplace_alnmethod ?: 'hmmer', - queryseqfile: it, - refseqfile: file( params.pplace_aln, checkIfExists: true ), - hmmfile: [], - refphylogeny: file( params.pplace_tree, checkIfExists: true ), - model: params.pplace_model, - taxonomy: params.pplace_taxonomy ? file( params.pplace_taxonomy, checkIfExists: true ) : [] - ] ] - } - FASTA_NEWICK_EPANG_GAPPA ( ch_pp_data ) - ch_versions_pplace_taxonomy = ch_versions_pplace_taxonomy.mix( FASTA_NEWICK_EPANG_GAPPA.out.versions ) - - ch_pplace_tax = FORMAT_PPLACETAX ( FASTA_NEWICK_EPANG_GAPPA.out.taxonomy_per_query ).tsv - - emit: - tax = ch_pplace_tax - versions = ch_versions_pplace_taxonomy -} diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 268e9190..d64ccac5 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -153,6 +153,7 @@ include { QIIME2_INSEQ } from '../modules/local/qiime2_inseq' include { QIIME2_FILTERTAXA } from '../modules/local/qiime2_filtertaxa' include { QIIME2_INASV } from '../modules/local/qiime2_inasv' include { QIIME2_INTREE } from '../modules/local/qiime2_intree' +include { FORMAT_PPLACETAX } from '../modules/local/format_pplacetax' include { FILTER_STATS } from '../modules/local/filter_stats' include { MERGE_STATS as MERGE_STATS_FILTERTAXA } from '../modules/local/merge_stats' include { QIIME2_BARPLOT } from '../modules/local/qiime2_barplot' @@ -174,7 +175,6 @@ include { QIIME2_TAXONOMY } from '../subworkflows/local/qiime2_tax include { CUTADAPT_WORKFLOW } from '../subworkflows/local/cutadapt_workflow' include { DADA2_TAXONOMY_WF } from '../subworkflows/local/dada2_taxonomy_wf' include { SINTAX_TAXONOMY_WF } from '../subworkflows/local/sintax_taxonomy_wf' -include { PPLACE_TAXONOMY_WF } from '../subworkflows/local/pplace_taxonomy_wf' include { QIIME2_EXPORT } from '../subworkflows/local/qiime2_export' include { QIIME2_BARPLOTAVG } from '../subworkflows/local/qiime2_barplotavg' include { QIIME2_DIVERSITY } from '../subworkflows/local/qiime2_diversity' @@ -193,6 +193,7 @@ include { QIIME2_ANCOM } from '../subworkflows/local/qiime2_anc include { FASTQC } from '../modules/nf-core/fastqc/main' include { MULTIQC } from '../modules/nf-core/multiqc/main' include { CUSTOM_DUMPSOFTWAREVERSIONS } from '../modules/nf-core/custom/dumpsoftwareversions/main' +include { FASTA_NEWICK_EPANG_GAPPA } from '../subworkflows/nf-core/fasta_newick_epang_gappa/main' /* @@ -427,8 +428,22 @@ workflow AMPLISEQ { // Phylo placement if ( params.pplace_tree ) { - ch_pplace_tax = PPLACE_TAXONOMY_WF ( ch_fasta ).tax - ch_versions = ch_versions.mix(PPLACE_TAXONOMY_WF.out.versions) + ch_pp_data = ch_fasta.map { it -> + [ meta: [ id: params.pplace_name ?: 'user_tree' ], + data: [ + alignmethod: params.pplace_alnmethod ?: 'hmmer', + queryseqfile: it, + refseqfile: file( params.pplace_aln, checkIfExists: true ), + hmmfile: [], + refphylogeny: file( params.pplace_tree, checkIfExists: true ), + model: params.pplace_model, + taxonomy: params.pplace_taxonomy ? file( params.pplace_taxonomy, checkIfExists: true ) : [] + ] ] + } + FASTA_NEWICK_EPANG_GAPPA ( ch_pp_data ) + ch_versions = ch_versions.mix( FASTA_NEWICK_EPANG_GAPPA.out.versions ) + + ch_pplace_tax = FORMAT_PPLACETAX ( FASTA_NEWICK_EPANG_GAPPA.out.taxonomy_per_query ).tsv } else { ch_pplace_tax = Channel.empty() } From ddfebe21d544f08a29735b00f7a78ae3bb11dda5 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Wed, 10 May 2023 13:27:49 +0200 Subject: [PATCH 086/158] Change NA to blank in sintax table --- bin/convert_sintax_output.py | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/bin/convert_sintax_output.py b/bin/convert_sintax_output.py index 1cbd8073..9a5ca22c 100755 --- a/bin/convert_sintax_output.py +++ b/bin/convert_sintax_output.py @@ -71,7 +71,7 @@ def complete_list(some_list, target_len): - return some_list[:target_len] + ["NA"] * (target_len - len(some_list)) + return some_list[:target_len] + [""] * (target_len - len(some_list)) # Find out if taxonomies contain SH groups @@ -132,7 +132,7 @@ def complete_list(some_list, target_len): annot_list.append(sh) print(tmp_list[0], "\t".join(annot_list), confidence, sequence, sep="\t", file=args.outfile) else: - print(tmp_list[0], "\t".join(["NA"] * num_cols), "", sequence, sep="\t", file=args.outfile) + print(tmp_list[0], "\t".join([""] * num_cols), "", sequence, sep="\t", file=args.outfile) args.outfile.close() From 0135398f88b07dc8b8c0ab269e9d7ba440b1997b Mon Sep 17 00:00:00 2001 From: jtangrot Date: Wed, 10 May 2023 14:01:20 +0200 Subject: [PATCH 087/158] Add check if sintax should run --- subworkflows/local/dada2_taxonomy_wf.nf | 1 - workflows/ampliseq.nf | 2 +- 2 files changed, 1 insertion(+), 2 deletions(-) diff --git a/subworkflows/local/dada2_taxonomy_wf.nf b/subworkflows/local/dada2_taxonomy_wf.nf index 8a1f11e6..c5259e6c 100644 --- a/subworkflows/local/dada2_taxonomy_wf.nf +++ b/subworkflows/local/dada2_taxonomy_wf.nf @@ -103,7 +103,6 @@ workflow DADA2_TAXONOMY_WF { ch_dada2_tax = ch_dada2_tax1 } - emit: tax = ch_dada2_tax versions = ch_versions_dada_taxonomy diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index d64ccac5..37526333 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -413,7 +413,7 @@ workflow AMPLISEQ { // SINTAX // This will only run if --sintax_ref_taxonomy is defined, // i.e. if the channel ch_sintax_ref_taxonomy is not empty - if (!params.skip_taxonomy) { + if (!params.skip_taxonomy && params.sintax_ref_taxonomy) { SINTAX_TAXONOMY_WF ( ch_sintax_ref_taxonomy.collect(), val_sintax_ref_taxonomy, From 9b2688d6a91036ea1f9abecc90e70750474ebf3a Mon Sep 17 00:00:00 2001 From: jtangrot Date: Wed, 10 May 2023 14:02:16 +0200 Subject: [PATCH 088/158] Remove comment --- workflows/ampliseq.nf | 2 -- 1 file changed, 2 deletions(-) diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 37526333..ca1db1e1 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -411,8 +411,6 @@ workflow AMPLISEQ { } // SINTAX - // This will only run if --sintax_ref_taxonomy is defined, - // i.e. if the channel ch_sintax_ref_taxonomy is not empty if (!params.skip_taxonomy && params.sintax_ref_taxonomy) { SINTAX_TAXONOMY_WF ( ch_sintax_ref_taxonomy.collect(), From 1efaad998306641d8d1f48b659714772dc02a0f2 Mon Sep 17 00:00:00 2001 From: daniel Date: Wed, 10 May 2023 15:27:05 +0200 Subject: [PATCH 089/158] fix diversity analysis --- CHANGELOG.md | 2 +- workflows/ampliseq.nf | 2 +- 2 files changed, 2 insertions(+), 2 deletions(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index ba3b57c6..d8603fbf 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -7,7 +7,7 @@ and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0 ### `Added` -- [#564](https://github.com/nf-core/ampliseq/pull/564),[#567](https://github.com/nf-core/ampliseq/pull/567) - Added phylogenetic placement +- [#564](https://github.com/nf-core/ampliseq/pull/564),[#567](https://github.com/nf-core/ampliseq/pull/567),[#582](https://github.com/nf-core/ampliseq/pull/582) - Added phylogenetic placement ### `Changed` diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index d86a87cd..40dd835b 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -534,7 +534,7 @@ workflow AMPLISEQ { // Import phylogenetic tree into QIIME2 if ( params.pplace_tree ) { ch_tree = QIIME2_INTREE ( FASTA_NEWICK_EPANG_GAPPA.out.grafted_phylogeny ).qza - } else { ch_tree = Channel.empty() } + } else { ch_tree = [] } // Import taxonomic classification into QIIME2, if available if ( params.skip_taxonomy ) { From a92193b3c794aa03213a9bcbd08abe8b1dc88cc4 Mon Sep 17 00:00:00 2001 From: lokeshbio Date: Fri, 12 May 2023 16:15:09 +0200 Subject: [PATCH 090/158] codons updated chnages based on comments --- bin/filt_InsBiom_asv.py | 107 -------------------------------- bin/filt_codons.py | 31 ++++++--- bin/filt_coi_stats.r | 23 ------- conf/modules.config | 4 ++ docs/output.md | 2 +- modules/local/filter_codons.nf | 32 +++------- modules/local/filter_coi_asv.nf | 36 ----------- nextflow.config | 2 + nextflow_schema.json | 24 +++++-- workflows/ampliseq.nf | 4 +- 10 files changed, 60 insertions(+), 205 deletions(-) delete mode 100755 bin/filt_InsBiom_asv.py delete mode 100755 bin/filt_coi_stats.r delete mode 100644 modules/local/filter_coi_asv.nf diff --git a/bin/filt_InsBiom_asv.py b/bin/filt_InsBiom_asv.py deleted file mode 100755 index 5c32e887..00000000 --- a/bin/filt_InsBiom_asv.py +++ /dev/null @@ -1,107 +0,0 @@ -#!/usr/bin/env python3 -##### Program description ####### -# -# Title: Subsetting Insect Biome ASVs -# -# Author(s): Lokeshwaran Manoharan -# -# -# -# Description: The ASVs that are supposed to be of particular length (418 ± nx3 and between 403 and 418), possibly does not contain any stop codon in the right reading frame. -# -# List of subroutines: -# -# -# -# Overall procedure: using functions and dictionaries in python -# -# Usage: filt_InsBiom_asv.py -# -################################## - -import re -import sys -import copy -import argparse - -usage = '''This program takes the output from the AmpliSeq dada2 pipeline and filteres the COI ASVs and only keep the ASVs of particular length (418 ± nx3 and between 403 and 418) and does not contain any stop codon in the right reading frame''' - -parser = argparse.ArgumentParser(description=usage) - -parser.add_argument( - '-f', '--fasta', - dest='fasta', - type=argparse.FileType('r'), - help='Fasta file of the ASVs from the AmpliSeq pipeline', - required=True - ) - -parser.add_argument( - '-t', '--count-table', - dest='count', - type=argparse.FileType('r'), - help='Count table file of the ASVs from the AmpliSeq pipeline', - required=True - ) - -parser.add_argument( - '-p', '--out-prefix', - dest='prefix', - type=str, - help='Prefix of the output files with filtered ASVs and the corresponding count table. The output files will be in *_filtered.* format', - required=True - ) - - -args = parser.parse_args() - - -p1 = re.compile('\t') -p2 = re.compile('>') - -def check_asv(seq): - sub_seq = seq[1:] - stop_list = ['TAA', 'TAG'] - sub_list = [] - for x in range(0, len(sub_seq), 3): - sub_list.append(sub_seq[x:x+3]) - if not 402 <= len(sub_seq) <= 417: - return False - elif not len(sub_seq)%3 == 0: - return False - elif any(x in sub_list for x in stop_list): - return False - else: - return True - -Out_Seq = open(args.prefix +'_filtered.fna', 'w') -Out_list = open(args.prefix+'_filtered.list', 'w') -Out_table = open(args.prefix+'_filtered.table.tsv', 'w') - -count_dict = {} - -count = 0 -for line in args.count: - line = line.rstrip('\n') - if count == 0: - print(line, file = Out_table) - count += 1 - else: - tmp_list = re.split(p1, line) - count_dict[tmp_list[0]] = line - -for line in args.fasta: - line = line.rstrip('\n') - if re.match(p2, line) is not None: - bin_head = re.sub(p2, '', line) - elif check_asv(line): - print('>', bin_head,'\n', line, file = Out_Seq, sep='') - print(bin_head, file = Out_list) - print(count_dict[bin_head], file = Out_table) - -args.count.close() -args.fasta.close() -Out_Seq.close() -Out_list.close() -Out_table.close() - diff --git a/bin/filt_codons.py b/bin/filt_codons.py index ffe53018..127e8473 100755 --- a/bin/filt_codons.py +++ b/bin/filt_codons.py @@ -1,13 +1,13 @@ #!/usr/bin/env python3 ##### Program description ####### # -# Title: Subsetting Insect Biome ASVs +# Title: Filtering ASVs with stop codons # # Author(s): Lokeshwaran Manoharan # # # -# Description: The ASVs that are supposed to be of particular length (418 ± nx3 and between 403 and 418), possibly does not contain any stop codon in the right reading frame. +# Description: The ASVs are filtered if they contain stop codons in the reading frame specified. # # List of subroutines: # @@ -15,7 +15,7 @@ # # Overall procedure: using functions and dictionaries in python # -# Usage: filt_InsBiom_asv.py +# Usage: filt_codons.py [-h] -f -t [-s BEGIN] [-e END] -p # ################################## @@ -61,6 +61,15 @@ required=False ) +parser.add_argument( + '-x', '--stop-codons', + dest='stopcodon', + type=str, + help='Specific stop codons to look for. Specify multiple codons with in a comma separated list like: "TAA,TAG". By default TAA and TAG are being looked for.', + required=False, + default="TAA,TAG" + ) + parser.add_argument( '-p', '--out-prefix', dest='prefix', @@ -72,14 +81,20 @@ args = parser.parse_args() -begin = args.begin +if args.begin is not None: + begin = args.begin +else: + begin = 1 -p1 = re.compile('\t') -p2 = re.compile('>') +if args.stopcodon is not None: + stopcodon = args.stopcodon +else: + stopcodon = 'TAA,TAG' + +stop_list = [item.strip() for item in stopcodon.split(',')] def check_asv(seq, start, stop): sub_seq = seq[start-1:stop] - stop_list = ['TAA', 'TAG'] sub_list = [] for x in range(0, len(sub_seq), 3): sub_list.append(sub_seq[x:x+3]) @@ -95,6 +110,8 @@ def check_asv(seq, start, stop): Out_table = open(args.prefix+'_filtered.table.tsv', 'w') count_dict = {} +p1 = re.compile('\t') +p2 = re.compile('>') count = 0 for line in args.count: diff --git a/bin/filt_coi_stats.r b/bin/filt_coi_stats.r deleted file mode 100755 index 3c4ef204..00000000 --- a/bin/filt_coi_stats.r +++ /dev/null @@ -1,23 +0,0 @@ -#!/usr/bin/env Rscript - -args = commandArgs(trailingOnly=TRUE) - -if(length(args) < 2){ - stop("Usage: filt_coi_stats.r ") -} - -dada2_stats <- args[1] -coi_filt <- args[2] - -old_stats <- read.table(dada2_stats, header = T, check.names = F, row.names = 1) - -filt_table <- read.table(coi_filt, header = T, check.names = F, row.names = 1) - -filt_counts <- as.data.frame(colSums(filt_table)) -names(filt_counts) <- "coi_filtered" - -new_stats <- merge(old_stats, filt_counts, by = 'row.names') -new_stats$sample <- row.names(new_stats) - -write.table(new_stats, file = "stats.filt.tsv", row.names=FALSE, sep="\t", col.names = TRUE, quote = FALSE, na = '') - diff --git a/conf/modules.config b/conf/modules.config index 66aa5d3e..cd78aa14 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -310,6 +310,10 @@ process { } withName: FILTER_CODONS { + ext.args = [params.orf_start ? "-s ${params.orf_start}" : '', + params.orf_end ? "-e ${params.orf_end}" : '', + params.stop_codons ? "-x ${params.stop_codons}" : '' + ].join(' ') publishDir = [ path: { "${params.outdir}/Codons_filtered" }, mode: params.publish_dir_mode, diff --git a/docs/output.md b/docs/output.md index 3f1c2e46..34bf9f08 100644 --- a/docs/output.md +++ b/docs/output.md @@ -191,7 +191,7 @@ Optionally, the ITS region can be extracted from each ASV sequence using ITSx, a Optionally, the ASVs can be filtered if they contain stop codons (`"TAA,TAG"`) in the specified open reading frame of the ASV. The filtering step also filters ASVs in the specifed checking range, if they are not multiple of 3. -This filtering can be done by the setting the `--orf_start` option to starting position on the ASV where you would like to start checking of codon usage! For example, setting this parameter to `1`, would start checking the ASV for codons from the beginning of the ASV or if it is set to `22`, the check starts the open reading frame from position 22 on each ASV. By default, the filtering applies from the `--orf_start` to the end of the ASV. But, this can be changed by setting the `--orf_end` to specify the end of the open reading frame where you want to look for stop codons +This filtering can be done by the setting the `--filter_codons`. By default, the codons are calculated and checked from the beginning to the end of the ASV sequence and the filter also checks if the length is of multiple of 3. By default the stop codons that are being screend for is `TAA` and `TAG`. These settings can be changed by `--orf_start` option to change the starting position of the codon on the ASV where you would like to start checking of codon usage! For example, setting this parameter to `1`, would start checking the ASV for codons from the beginning of the ASV or if it is set to `22`, the check starts the open reading frame from position 22 on each ASV. By default, the filtering applies from the `--orf_start` to the end of the ASV. But, this can be changed by setting the `--orf_end` to specify the end of the open reading frame where you want to look for stop codons. To look for different stop codons than default, `--stop_codons` can be used by specifying other single/multiple stop codons as a comma-separated list with this parameter like: `--stop_codons "TAA,TAG,TGA"` > **NB:** just make sure that when you set `--orf_end`, the length between `--orf_start` and `--orf_end` is a multiple of 3, otherwise all ASVs would be filtered. diff --git a/modules/local/filter_codons.nf b/modules/local/filter_codons.nf index 10aa1b99..d1e91532 100644 --- a/modules/local/filter_codons.nf +++ b/modules/local/filter_codons.nf @@ -11,36 +11,19 @@ process FILTER_CODONS { path(dada2stats) output: - path( "ASV_codon_filtered.table.tsv" ) , emit: asv - path( "ASV_codon_filtered.fna" ) , emit: fasta - path( "ASV_codon_filtered.list" ) , emit: list - path( "codon.filtered.stats.tsv" ), emit: stats - path "versions.yml" , emit: versions + path( "ASV_codon_filtered.table.tsv" ) , emit: asv + path( "ASV_codon_filtered.fna" ) , emit: fasta + path( "ASV_codon_filtered.list" ) , emit: list + path( "codon.filtered.stats.tsv" ) , emit: stats + path( "versions.yml" ) , emit: versions when: task.ext.when == null || task.ext.when script: - if (!params.orf_end) { - def orf_start = params.orf_start ?: '1' - - """ - filt_codons.py -f ${fasta} -t ${asv} -s ${orf_start} -p ASV_codon - filt_codon_stats.r ${dada2stats} ASV_codon_filtered.table.tsv - - cat <<-END_VERSIONS > versions.yml - "${task.process}": - python: \$(python --version 2>&1 | sed 's/Python //g') - R: \$(R --version | sed -n 1p | sed 's/R version //g' | sed 's/\\s.*\$//') - END_VERSIONS - """ - - } else { - def orf_start = params.orf_start ?: '1' - def orf_end = params.orf_end ?: '1000000' - + def args = task.ext.args ?: '' """ - filt_codons.py -f ${fasta} -t ${asv} -s ${orf_start} -e ${orf_end} -p ASV_codon + filt_codons.py -f ${fasta} -t ${asv} -p ASV_codon ${args} filt_codon_stats.r ${dada2stats} ASV_codon_filtered.table.tsv cat <<-END_VERSIONS > versions.yml @@ -49,6 +32,5 @@ process FILTER_CODONS { R: \$(R --version | sed -n 1p | sed 's/R version //g' | sed 's/\\s.*\$//') END_VERSIONS """ - } } \ No newline at end of file diff --git a/modules/local/filter_coi_asv.nf b/modules/local/filter_coi_asv.nf deleted file mode 100644 index cb98c1a1..00000000 --- a/modules/local/filter_coi_asv.nf +++ /dev/null @@ -1,36 +0,0 @@ -process FILTER_COI_ASV { - tag "${fasta}" - label 'process_low' - - conda (params.enable_conda ? { exit 1 "QIIME2 has no conda package" } : null) - container "quay.io/qiime2/core:2022.8" - - input: - path(fasta) - path(asv) - path(dada2stats) - - output: - path( "ASV_filtered.table.tsv" ) , emit: asv - path( "ASV_filtered.fna" ) , emit: fasta - path( "ASV_filtered.list" ) , emit: list - path( "stats.filt.tsv" ), emit: stats - path "versions.yml" , emit: versions - - when: - task.ext.when == null || task.ext.when - - script: - """ - filt_InsBiom_asv.py -f ${fasta} -t ${asv} -p ASV - filt_coi_stats.r ${dada2stats} ASV_filtered.table.tsv - - cat <<-END_VERSIONS > versions.yml - "${task.process}": - python: \$(python --version 2>&1 | sed 's/Python //g') - R: \$(R --version | sed -n 1p | sed 's/R version //g' | sed 's/\\s.*\$//') - END_VERSIONS - - """ - -} \ No newline at end of file diff --git a/nextflow.config b/nextflow.config index 3b408d99..7216381c 100644 --- a/nextflow.config +++ b/nextflow.config @@ -58,8 +58,10 @@ params { filter_ssu = null min_len_asv = null max_len_asv = null + filter_codons = null orf_start = null orf_end = null + stop_codons = null diversity_rarefaction_depth = 500 ancom_sample_min_count = 1 diff --git a/nextflow_schema.json b/nextflow_schema.json index 304e9012..ae7c6528 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -335,10 +335,26 @@ "description": "Maximum ASV length", "help_text": "Remove ASV that are above the maximum length threshold (default: filter is disabled, otherwise 1000000). Lowering the threshold might reduce false positive ASVs (e.g. PCR off-targets)." }, - "filter_coi_asv": { - "type": "boolean", - "description": "Filter COI ASVs", - "help_text": "This setting is to be used only when any of the combination of following primers have been used \n\nforward:\n - \"CCHGAYATRGCHTTYCCHCG\"\n - \"ACCHGAYATRGCHTTYCCHCG\"\n - \"GACCHGAYATRGCHTTYCCHCG\"\n - \"TGACCHGAYATRGCHTTYCCHCG\"\n\nreverse:\n - \"CDGGRTGNCCRAARAAYCA\"\n - \"TCDGGRTGNCCRAARAAYCA\"\n - \"ATCDGGRTGNCCRAARAAYCA\"\n - \"GATCDGGRTGNCCRAARAAYCA\"\nASVs that do not follow the length category: 418 ± nx3 and between 403 and 418 and the ones that contain stop codon are filtered." + "filter_codons": { + "type": "boolean", + "description": "Filter ASVs based on codon usage", + "help_text": "This setting is to be used when the ASVs need to be filtered if they contain STOP codons in their sequence and they are of length in the multiple of 3.", + "properties": { + "orf_start": { + "type": "integer", + "description": "Starting position to check for the stop codons in the ASVs", + "help_text": "By default, when the `--filter_codons` is triggered, the codons start from the beginning of the ASV sequences. If you would like to change this setting, you can specify from which position of the ASV does the codon triplets be calculated and checked. \n\nPlease note that the length from this position to the end of the ASV or until the `--orf_end` should be a multiple of 3 to make sure the filtering works properly." + }, + "orf_end": { + "type": "integer", + "description": "Ending position to check for the stop codons in the ASVs", + "help_text": "By default, when the `--filter_codons` is triggered, the codons are checked until the end of the ASV sequences. If you would like to change this setting, you can specify until which position of the ASV sequences does the codon triplets be checked.\n\nPlease note that the length of the ASV from the beginning or from the `--orf_start` until this position should be a multiple of 3 to make sure the filtering works properly." + },"stop_codons": { + "type": "string", + "description": "Specific stop codons to check for in the ASV", + "help_text": "By default, when the `--filter_codons` is triggered, the codons `TAA,TAG` are checked in the ASVs that are to be filtered. If there are other single/multiple stop codons that needs to be checked, the codons can be specified as a comma-separated list with this parameter like: `--stop_codons \"TAA,TAG,TGA\"`" + } + } }, "exclude_taxa": { "type": "string", diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 74d35b50..95df1f55 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -132,7 +132,7 @@ include { FILTER_SSU } from '../modules/local/filter_ssu' include { FILTER_LEN_ASV } from '../modules/local/filter_len_asv' include { MERGE_STATS as MERGE_STATS_FILTERSSU } from '../modules/local/merge_stats' include { MERGE_STATS as MERGE_STATS_FILTERLENASV } from '../modules/local/merge_stats' -include { FILTER_CODONS } from '../modules/local/filter_codons' +include { FILTER_CODONS } from '../modules/local/filter_codons' include { FORMAT_FASTAINPUT } from '../modules/local/format_fastainput' include { FORMAT_TAXONOMY } from '../modules/local/format_taxonomy' include { ITSX_CUTASV } from '../modules/local/itsx_cutasv' @@ -353,7 +353,7 @@ workflow AMPLISEQ { // // Modules : Filtering based on codons in an open reading frame // - if (params.orf_start || params.orf_end) { + if (params.filter_codons || params.orf_start || params.orf_end || params.stop_codons) { FILTER_CODONS ( ch_dada2_fasta, ch_dada2_asv, ch_stats ) ch_versions = ch_versions.mix(FILTER_CODONS.out.versions.ifEmpty(null)) ch_stats = FILTER_CODONS.out.stats From 316470060dc2951cbee44f43ddd677abe8be2314 Mon Sep 17 00:00:00 2001 From: lokeshbio Date: Mon, 15 May 2023 12:43:41 +0200 Subject: [PATCH 091/158] tried correcting run errors --- docs/output.md | 2 +- modules/local/filter_codons.nf | 3 +-- nextflow_schema.json | 32 +++++++++++++++----------------- 3 files changed, 17 insertions(+), 20 deletions(-) diff --git a/docs/output.md b/docs/output.md index 34bf9f08..68010100 100644 --- a/docs/output.md +++ b/docs/output.md @@ -193,7 +193,7 @@ Optionally, the ASVs can be filtered if they contain stop codons (`"TAA,TAG"`) i This filtering can be done by the setting the `--filter_codons`. By default, the codons are calculated and checked from the beginning to the end of the ASV sequence and the filter also checks if the length is of multiple of 3. By default the stop codons that are being screend for is `TAA` and `TAG`. These settings can be changed by `--orf_start` option to change the starting position of the codon on the ASV where you would like to start checking of codon usage! For example, setting this parameter to `1`, would start checking the ASV for codons from the beginning of the ASV or if it is set to `22`, the check starts the open reading frame from position 22 on each ASV. By default, the filtering applies from the `--orf_start` to the end of the ASV. But, this can be changed by setting the `--orf_end` to specify the end of the open reading frame where you want to look for stop codons. To look for different stop codons than default, `--stop_codons` can be used by specifying other single/multiple stop codons as a comma-separated list with this parameter like: `--stop_codons "TAA,TAG,TGA"` -> **NB:** just make sure that when you set `--orf_end`, the length between `--orf_start` and `--orf_end` is a multiple of 3, otherwise all ASVs would be filtered. +> **NB:** just make sure that when you set `--orf_start` and/or `--orf_end`, the length between `--orf_start` and `--orf_end` is a multiple of 3, otherwise all ASVs would be filtered.
diff --git a/modules/local/filter_codons.nf b/modules/local/filter_codons.nf index d1e91532..186c18cb 100644 --- a/modules/local/filter_codons.nf +++ b/modules/local/filter_codons.nf @@ -32,5 +32,4 @@ process FILTER_CODONS { R: \$(R --version | sed -n 1p | sed 's/R version //g' | sed 's/\\s.*\$//') END_VERSIONS """ - -} \ No newline at end of file +} diff --git a/nextflow_schema.json b/nextflow_schema.json index ae7c6528..779c1d9a 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -338,23 +338,21 @@ "filter_codons": { "type": "boolean", "description": "Filter ASVs based on codon usage", - "help_text": "This setting is to be used when the ASVs need to be filtered if they contain STOP codons in their sequence and they are of length in the multiple of 3.", - "properties": { - "orf_start": { - "type": "integer", - "description": "Starting position to check for the stop codons in the ASVs", - "help_text": "By default, when the `--filter_codons` is triggered, the codons start from the beginning of the ASV sequences. If you would like to change this setting, you can specify from which position of the ASV does the codon triplets be calculated and checked. \n\nPlease note that the length from this position to the end of the ASV or until the `--orf_end` should be a multiple of 3 to make sure the filtering works properly." - }, - "orf_end": { - "type": "integer", - "description": "Ending position to check for the stop codons in the ASVs", - "help_text": "By default, when the `--filter_codons` is triggered, the codons are checked until the end of the ASV sequences. If you would like to change this setting, you can specify until which position of the ASV sequences does the codon triplets be checked.\n\nPlease note that the length of the ASV from the beginning or from the `--orf_start` until this position should be a multiple of 3 to make sure the filtering works properly." - },"stop_codons": { - "type": "string", - "description": "Specific stop codons to check for in the ASV", - "help_text": "By default, when the `--filter_codons` is triggered, the codons `TAA,TAG` are checked in the ASVs that are to be filtered. If there are other single/multiple stop codons that needs to be checked, the codons can be specified as a comma-separated list with this parameter like: `--stop_codons \"TAA,TAG,TGA\"`" - } - } + "help_text": "This setting is to be used when the ASVs need to be filtered if they contain STOP codons in their sequence and they are of length in the multiple of 3." + }, + "orf_start": { + "type": "integer", + "description": "Starting position to check for the stop codons in the ASVs", + "help_text": "By default, when the `--filter_codons` is triggered, the codons start from the beginning of the ASV sequences. If you would like to change this setting, you can specify from which position of the ASV does the codon triplets be calculated and checked. \n\nPlease note that the length from this position to the end of the ASV or until the `--orf_end` should be a multiple of 3 to make sure the filtering works properly." + }, + "orf_end": { + "type": "integer", + "description": "Ending position to check for the stop codons in the ASVs", + "help_text": "By default, when the `--filter_codons` is triggered, the codons are checked until the end of the ASV sequences. If you would like to change this setting, you can specify until which position of the ASV sequences does the codon triplets be checked.\n\nPlease note that the length of the ASV from the beginning or from the `--orf_start` until this position should be a multiple of 3 to make sure the filtering works properly." + },"stop_codons": { + "type": "string", + "description": "Specific stop codons to check for in the ASV", + "help_text": "By default, when the `--filter_codons` is triggered, the codons `TAA,TAG` are checked in the ASVs that are to be filtered. If there are other single/multiple stop codons that needs to be checked, the codons can be specified as a comma-separated list with this parameter like: `--stop_codons \"TAA,TAG,TGA\"`" }, "exclude_taxa": { "type": "string", From 6c2070e7514b5fbd2aa559cbbd23754c49ba5462 Mon Sep 17 00:00:00 2001 From: lokeshbio Date: Mon, 15 May 2023 13:33:10 +0200 Subject: [PATCH 092/158] updated defaults in config and json --- modules/local/filter_codons.nf | 21 ++++++++++----------- nextflow.config | 4 ++-- nextflow_schema.json | 2 ++ workflows/ampliseq.nf | 2 +- 4 files changed, 15 insertions(+), 14 deletions(-) diff --git a/modules/local/filter_codons.nf b/modules/local/filter_codons.nf index 186c18cb..a0f19f5d 100644 --- a/modules/local/filter_codons.nf +++ b/modules/local/filter_codons.nf @@ -21,15 +21,14 @@ process FILTER_CODONS { task.ext.when == null || task.ext.when script: - def args = task.ext.args ?: '' - """ - filt_codons.py -f ${fasta} -t ${asv} -p ASV_codon ${args} - filt_codon_stats.r ${dada2stats} ASV_codon_filtered.table.tsv - - cat <<-END_VERSIONS > versions.yml - "${task.process}": - python: \$(python --version 2>&1 | sed 's/Python //g') - R: \$(R --version | sed -n 1p | sed 's/R version //g' | sed 's/\\s.*\$//') - END_VERSIONS - """ + def args = task.ext.args ?: '' + """ + filt_codons.py -f ${fasta} -t ${asv} -p ASV_codon ${args} + filt_codon_stats.r ${dada2stats} ASV_codon_filtered.table.tsv + cat <<-END_VERSIONS > versions.yml + "${task.process}": + python: \$(python --version 2>&1 | sed 's/Python //g') + R: \$(R --version | sed -n 1p | sed 's/R version //g' | sed 's/\\s.*\$//') + END_VERSIONS + """ } diff --git a/nextflow.config b/nextflow.config index 7216381c..8ce82b37 100644 --- a/nextflow.config +++ b/nextflow.config @@ -59,9 +59,9 @@ params { min_len_asv = null max_len_asv = null filter_codons = null - orf_start = null + orf_start = 1 orf_end = null - stop_codons = null + stop_codons = "TAA,TAG" diversity_rarefaction_depth = 500 ancom_sample_min_count = 1 diff --git a/nextflow_schema.json b/nextflow_schema.json index 779c1d9a..0274619a 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -342,6 +342,7 @@ }, "orf_start": { "type": "integer", + "default": 1, "description": "Starting position to check for the stop codons in the ASVs", "help_text": "By default, when the `--filter_codons` is triggered, the codons start from the beginning of the ASV sequences. If you would like to change this setting, you can specify from which position of the ASV does the codon triplets be calculated and checked. \n\nPlease note that the length from this position to the end of the ASV or until the `--orf_end` should be a multiple of 3 to make sure the filtering works properly." }, @@ -351,6 +352,7 @@ "help_text": "By default, when the `--filter_codons` is triggered, the codons are checked until the end of the ASV sequences. If you would like to change this setting, you can specify until which position of the ASV sequences does the codon triplets be checked.\n\nPlease note that the length of the ASV from the beginning or from the `--orf_start` until this position should be a multiple of 3 to make sure the filtering works properly." },"stop_codons": { "type": "string", + "default": "TAA,TAG", "description": "Specific stop codons to check for in the ASV", "help_text": "By default, when the `--filter_codons` is triggered, the codons `TAA,TAG` are checked in the ASVs that are to be filtered. If there are other single/multiple stop codons that needs to be checked, the codons can be specified as a comma-separated list with this parameter like: `--stop_codons \"TAA,TAG,TGA\"`" }, diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 95df1f55..35e6abf4 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -353,7 +353,7 @@ workflow AMPLISEQ { // // Modules : Filtering based on codons in an open reading frame // - if (params.filter_codons || params.orf_start || params.orf_end || params.stop_codons) { + if (params.filter_codons ) { FILTER_CODONS ( ch_dada2_fasta, ch_dada2_asv, ch_stats ) ch_versions = ch_versions.mix(FILTER_CODONS.out.versions.ifEmpty(null)) ch_stats = FILTER_CODONS.out.stats From 9b7aac32dc873471c940d1fc51ce86697d740161 Mon Sep 17 00:00:00 2001 From: lokeshbio Date: Mon, 15 May 2023 13:49:43 +0200 Subject: [PATCH 093/158] upadted error message for orf_end multiple of 3 --- workflows/ampliseq.nf | 5 +++++ 1 file changed, 5 insertions(+) diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 35e6abf4..97181fd9 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -100,6 +100,11 @@ if ( params.dada_ref_taxonomy && !params.skip_dada_addspecies && !params.skip_ta } } +// Error message for incompatible combination of --orf_start and --orf_end +if ( params.orf_end && ( ( ( params.orf_end + 1 ) - params.orf_start ) % 3 != 0 ) ) { + Nextflow.error("Incompatible parameters: The difference of `--orf_end` and `--orf_start` must be a multiple of 3.") +} + //only run QIIME2 when taxonomy is actually calculated and all required data is available if ( !(workflow.profile.tokenize(',').intersect(['conda', 'mamba']).size() >= 1) && !params.skip_taxonomy && !params.skip_qiime ) { run_qiime2 = true From 83ce01b9edea6d4c0f930acd323660e0ce6037a5 Mon Sep 17 00:00:00 2001 From: lokeshbio Date: Tue, 16 May 2023 10:55:36 +0200 Subject: [PATCH 094/158] ran prettier and black --- bin/filt_codons.py | 175 ++++++++++++++++++++++--------------------- conf/modules.config | 2 +- core.24117 | Bin 0 -> 10240 bytes core.725 | Bin 0 -> 10240 bytes docs/output.md | 3 +- nextflow_schema.json | 3 +- 6 files changed, 95 insertions(+), 88 deletions(-) create mode 100644 core.24117 create mode 100644 core.725 diff --git a/bin/filt_codons.py b/bin/filt_codons.py index 127e8473..09754b57 100755 --- a/bin/filt_codons.py +++ b/bin/filt_codons.py @@ -1,15 +1,15 @@ #!/usr/bin/env python3 ##### Program description ####### # -# Title: Filtering ASVs with stop codons +# Title: Filtering ASVs with stop codons # # Author(s): Lokeshwaran Manoharan # # # -# Description: The ASVs are filtered if they contain stop codons in the reading frame specified. -# -# List of subroutines: +# Description: The ASVs are filtered if they contain stop codons in the reading frame specified. +# +# List of subroutines: # # # @@ -24,122 +24,129 @@ import copy import argparse -usage = '''This program filters ASVs that contains any stop codon or if the length from the position is not a multiple of 3 in the right reading frame.''' +usage = """This program filters ASVs that contains any stop codon or if the length from the position is not a multiple of 3 in the right reading frame.""" parser = argparse.ArgumentParser(description=usage) parser.add_argument( - '-f', '--fasta', - dest='fasta', - type=argparse.FileType('r'), - help='Fasta file of the ASVs from the AmpliSeq pipeline', - required=True - ) + "-f", + "--fasta", + dest="fasta", + type=argparse.FileType("r"), + help="Fasta file of the ASVs from the AmpliSeq pipeline", + required=True, +) parser.add_argument( - '-t', '--count-table', - dest='count', - type=argparse.FileType('r'), - help='Count table file of the ASVs from the AmpliSeq pipeline', - required=True - ) + "-t", + "--count-table", + dest="count", + type=argparse.FileType("r"), + help="Count table file of the ASVs from the AmpliSeq pipeline", + required=True, +) parser.add_argument( - '-s', '--start-position', - dest='begin', - type=int, - help='Specific position of the ASV where to start checking for codons. By default it starts from position 1', - required=False, - default=0 - ) + "-s", + "--start-position", + dest="begin", + type=int, + help="Specific position of the ASV where to start checking for codons. By default it starts from position 1", + required=False, + default=0, +) parser.add_argument( - '-e', '--end-position', - dest='end', - type=int, - help='Specific position of the ASV where to stop checking for codons. By default it checks until the end of the sequence', - required=False - ) + "-e", + "--end-position", + dest="end", + type=int, + help="Specific position of the ASV where to stop checking for codons. By default it checks until the end of the sequence", + required=False, +) parser.add_argument( - '-x', '--stop-codons', - dest='stopcodon', - type=str, - help='Specific stop codons to look for. Specify multiple codons with in a comma separated list like: "TAA,TAG". By default TAA and TAG are being looked for.', - required=False, - default="TAA,TAG" - ) + "-x", + "--stop-codons", + dest="stopcodon", + type=str, + help='Specific stop codons to look for. Specify multiple codons with in a comma separated list like: "TAA,TAG". By default TAA and TAG are being looked for.', + required=False, + default="TAA,TAG", +) parser.add_argument( - '-p', '--out-prefix', - dest='prefix', - type=str, - help='Prefix of the output files with filtered ASVs and the corresponding count table. The output files will be in *_filtered.* format', - required=True - ) + "-p", + "--out-prefix", + dest="prefix", + type=str, + help="Prefix of the output files with filtered ASVs and the corresponding count table. The output files will be in *_filtered.* format", + required=True, +) args = parser.parse_args() if args.begin is not None: - begin = args.begin + begin = args.begin else: - begin = 1 + begin = 1 if args.stopcodon is not None: - stopcodon = args.stopcodon + stopcodon = args.stopcodon else: - stopcodon = 'TAA,TAG' + stopcodon = "TAA,TAG" + +stop_list = [item.strip() for item in stopcodon.split(",")] -stop_list = [item.strip() for item in stopcodon.split(',')] def check_asv(seq, start, stop): - sub_seq = seq[start-1:stop] - sub_list = [] - for x in range(0, len(sub_seq), 3): - sub_list.append(sub_seq[x:x+3]) - if not len(sub_seq)%3 == 0: - return False - elif any(x in sub_list for x in stop_list): - return False - else: - return True - -Out_Seq = open(args.prefix +'_filtered.fna', 'w') -Out_list = open(args.prefix+'_filtered.list', 'w') -Out_table = open(args.prefix+'_filtered.table.tsv', 'w') + sub_seq = seq[start - 1 : stop] + sub_list = [] + for x in range(0, len(sub_seq), 3): + sub_list.append(sub_seq[x : x + 3]) + if not len(sub_seq) % 3 == 0: + return False + elif any(x in sub_list for x in stop_list): + return False + else: + return True + + +Out_Seq = open(args.prefix + "_filtered.fna", "w") +Out_list = open(args.prefix + "_filtered.list", "w") +Out_table = open(args.prefix + "_filtered.table.tsv", "w") count_dict = {} -p1 = re.compile('\t') -p2 = re.compile('>') +p1 = re.compile("\t") +p2 = re.compile(">") count = 0 for line in args.count: - line = line.rstrip('\n') - if count == 0: - print(line, file = Out_table) - count += 1 - else: - tmp_list = re.split(p1, line) - count_dict[tmp_list[0]] = line + line = line.rstrip("\n") + if count == 0: + print(line, file=Out_table) + count += 1 + else: + tmp_list = re.split(p1, line) + count_dict[tmp_list[0]] = line for line in args.fasta: - line = line.rstrip('\n') - if re.match(p2, line) is not None: - bin_head = re.sub(p2, '', line) - else: - if args.end is not None: - end = args.end - else: - end = len(line) - if check_asv(line, begin, end): - print('>', bin_head,'\n', line, file = Out_Seq, sep='') - print(bin_head, file = Out_list) - print(count_dict[bin_head], file = Out_table) + line = line.rstrip("\n") + if re.match(p2, line) is not None: + bin_head = re.sub(p2, "", line) + else: + if args.end is not None: + end = args.end + else: + end = len(line) + if check_asv(line, begin, end): + print(">", bin_head, "\n", line, file=Out_Seq, sep="") + print(bin_head, file=Out_list) + print(count_dict[bin_head], file=Out_table) args.count.close() args.fasta.close() Out_Seq.close() Out_list.close() Out_table.close() - diff --git a/conf/modules.config b/conf/modules.config index cd78aa14..176fcf7f 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -313,7 +313,7 @@ process { ext.args = [params.orf_start ? "-s ${params.orf_start}" : '', params.orf_end ? "-e ${params.orf_end}" : '', params.stop_codons ? "-x ${params.stop_codons}" : '' - ].join(' ') + ].join(' ') publishDir = [ path: { "${params.outdir}/Codons_filtered" }, mode: params.publish_dir_mode, diff --git a/core.24117 b/core.24117 new file mode 100644 index 0000000000000000000000000000000000000000..bb9cdc4dd0abef990fb0c7b4b512ad256289f061 GIT binary patch literal 10240 zcmb8!e@v8h90%~jKo18A8yGlfN6CT(PUca^yhB)EqM(OKlYo+vB?bj~=ur|-ARthX zf|5|M@l(V=U_l-QC7W#A294Wb!KNE-&<=+UO$%G2=lvDh+Mav!&-1sK~9 zg&VUvtPLo{Bje~I-MQ*qkzTy~@8dkNa?TyeQS*i;W@RDgJdt*BQc}(vX=54ZeBk^l zIp+&E@h#{4;rjA6VkI5qo|izRYgJB^3qo40dzx~=NS76IE(Gr8;wGw3hxD8lFG1Xv zdtSnkQ$>nbsQ+2bMPEz7HkBg+7OsZLS>YtZ?P#KALk4aEBH+*950`6Q5|(hTQqJ!DVdb zTnF4z;EPz<*4KsfvAvw@g6v=L)Ct=A1iR>siiu3a8dnb9%$s7jw=BuJeyrs?Qg$@y&yj^M~u3=UgD1 z>H+71;L>7Wqvi~TQ-8|25V*tJ;;23y+(RAb!r>;)aV`=rpevr5GYZadB7t%-aMRU^ zl#7MiewTA`aC1&cR9`%t>Tk{^!5O{ZqWY5ImNp!rT#7iq*OMui3g=Pu-%DopcoZCQ z5n?4QT|Y~QvktyP^<~0oCXZ4s3vR+8m2%mlul+dXa^RH8_b8VSH@owF$`!&LyUV!} zxVbMrp!&+;+=?HICmRPyUe*eaI=27)SUBhnv;2yTY&Sr zkWaaLaN73ID0d&u&9#7XkKnA)g_L^?SCPm$yLD1c{Yz)4J_oqgS4$}82p6&GEajZx zOi7$`g)>~`oI9Mtvy7V46K=YNbKY>Zlj2IRR4ey;osXFFx6i3Lec`Otm6Y>`Gj@MT zxj?wVp!1Xqf?NEMbHQ--bDRr-)23BXbL!x3?5n0+IGn!V0_7s%0#03|TohcbQw`-} z;BJL~MY&kGn=zLt7YAoLS4+8gxals=CBfwla4s3{;{4atoGEa|&<4t-!YOMTDVGkn z^ni1jaI^JIR9_aHVY!)d*>Lrvt(41w^J{6NT)sHJEte@*2se?#xe~ae>FrcsIb5;f z3gs%{^v&WwP!gB>T&aR9=;mAvoJ;9dYR)>ih<7_F*8peDy#UZ8~edTISZUY^CRV^;P(8=xf!@4fBp17eg6PBzUM9g literal 0 HcmV?d00001 diff --git a/core.725 b/core.725 new file mode 100644 index 0000000000000000000000000000000000000000..4770ba1b03284a9e99dd0e5412e85d2aa84c49c2 GIT binary patch literal 10240 zcmbW7JFn$56oq|9!=obw1t0{LK)f0Z;wdpGj06QB5eXp%naplmik>B;FnOY_FBEcj7rYbBeot6MtT7FZ*QfIhrqCeD$RZXJ>}5i{^QA z?Kfje-iT^xo-q$z2p2Bm`j4MQ6Qbe$SE65{;iBP}k-?JDtT4QMC`A|_X5VxelB+)W z^0!CAJ^6`w#$vUIyZJDofu8r<9QlZo9$RpY;wbGg$Z@mcsAJch^w&YJ_$c*By>iWo zPjXd;qi*C=Rw7sL=Mp`yIHeQspX@`=JQZir7i;8G^hR)<;;84qN1Wt2fg2Tv>1JAU zqDQX(czYlG5zc;+a`dU{!JXtX!>8NN-69rn-pl=*=oR29#aZ;l8s#aR1LqZ2-5o&I zh?6{J#&}-nX)%16L^ya)w}}C+B71Zu@HU?*i&Zoa9nMuino|e-${N z;cz>C-%1NLZnTP%o991fU|tp4OkI-hNK{Yru6G&ckrzQ?9h&3YYrC^lX3R zOqFt2pE!o8us?`y0TyI`w((M_#%8BWIhGJJ#Q5!@6!iC+B4X zS1K;7W{i77;#=iO&j0*%@;)-RgG7yT33^Y2`~FJGu|L>}^<*DraINC#v&TT|kt=^m z^~i1LiBEF1;^0*6lcWEIgA={a;17UEbM!)C zX9;dr9H!GB>#~33%)h7QLatEUv0N(nwEH>9r3U9RoXh0m!PSamF7=udJ#s^avq!i| zamR9LxPOYH4lCAK_OAn1{Fm+@^)@&;FFo^A9NID)c}gzi>I}D`mvD{Z=u_3@xir9M zyPp%i5nPwy`ta>s{C<#JCUDi+H|gKk2&TR>4B&U!2Xg&g+uV5T^||K0lydL0ru~tl zZp4LK&@16n%)_Zqiw0SBe>FI(xa;Uio*tZ29OSs)tcP@^KXT;_a+Tt)<5SLA z1E2MNPJFiDe1_Y|Q*!CRwTfdd8=UBo>oc6{bIFt3pg1)@-UrF0htKguZU8qa&fq!C z3jM|B@N!6CdMrJk^tq9Xu`E=lB#qo1HYs5*O?zOP+ z{8V~g=(B=u#K}1;!Ie*^9Htv{NaFkzaiX_y&unwGPDm9^ y(}0^Z+=ia`Bv<-Wk3Lmh@@%13DULd<;1_NJ*C}o}C_Z!=apH5D!}V_3^!Y!h=$O?2 literal 0 HcmV?d00001 diff --git a/docs/output.md b/docs/output.md index 68010100..3196fb67 100644 --- a/docs/output.md +++ b/docs/output.md @@ -193,8 +193,7 @@ Optionally, the ASVs can be filtered if they contain stop codons (`"TAA,TAG"`) i This filtering can be done by the setting the `--filter_codons`. By default, the codons are calculated and checked from the beginning to the end of the ASV sequence and the filter also checks if the length is of multiple of 3. By default the stop codons that are being screend for is `TAA` and `TAG`. These settings can be changed by `--orf_start` option to change the starting position of the codon on the ASV where you would like to start checking of codon usage! For example, setting this parameter to `1`, would start checking the ASV for codons from the beginning of the ASV or if it is set to `22`, the check starts the open reading frame from position 22 on each ASV. By default, the filtering applies from the `--orf_start` to the end of the ASV. But, this can be changed by setting the `--orf_end` to specify the end of the open reading frame where you want to look for stop codons. To look for different stop codons than default, `--stop_codons` can be used by specifying other single/multiple stop codons as a comma-separated list with this parameter like: `--stop_codons "TAA,TAG,TGA"` -> **NB:** just make sure that when you set `--orf_start` and/or `--orf_end`, the length between `--orf_start` and `--orf_end` is a multiple of 3, otherwise all ASVs would be filtered. - +> **NB:** just make sure that when you set `--orf_start` and/or `--orf_end`, the length between `--orf_start` and `--orf_end` is a multiple of 3, otherwise all ASVs would be filtered.
Output files diff --git a/nextflow_schema.json b/nextflow_schema.json index 0274619a..093a672b 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -350,7 +350,8 @@ "type": "integer", "description": "Ending position to check for the stop codons in the ASVs", "help_text": "By default, when the `--filter_codons` is triggered, the codons are checked until the end of the ASV sequences. If you would like to change this setting, you can specify until which position of the ASV sequences does the codon triplets be checked.\n\nPlease note that the length of the ASV from the beginning or from the `--orf_start` until this position should be a multiple of 3 to make sure the filtering works properly." - },"stop_codons": { + }, + "stop_codons": { "type": "string", "default": "TAA,TAG", "description": "Specific stop codons to check for in the ASV", From ac0255780db0870509370e79432fdaa073f93fd5 Mon Sep 17 00:00:00 2001 From: lokeshbio Date: Tue, 16 May 2023 10:56:00 +0200 Subject: [PATCH 095/158] ran prettier and black --- core.24117 | Bin 10240 -> 0 bytes core.725 | Bin 10240 -> 0 bytes 2 files changed, 0 insertions(+), 0 deletions(-) delete mode 100644 core.24117 delete mode 100644 core.725 diff --git a/core.24117 b/core.24117 deleted file mode 100644 index bb9cdc4dd0abef990fb0c7b4b512ad256289f061..0000000000000000000000000000000000000000 GIT binary patch literal 0 HcmV?d00001 literal 10240 zcmb8!e@v8h90%~jKo18A8yGlfN6CT(PUca^yhB)EqM(OKlYo+vB?bj~=ur|-ARthX zf|5|M@l(V=U_l-QC7W#A294Wb!KNE-&<=+UO$%G2=lvDh+Mav!&-1sK~9 zg&VUvtPLo{Bje~I-MQ*qkzTy~@8dkNa?TyeQS*i;W@RDgJdt*BQc}(vX=54ZeBk^l zIp+&E@h#{4;rjA6VkI5qo|izRYgJB^3qo40dzx~=NS76IE(Gr8;wGw3hxD8lFG1Xv zdtSnkQ$>nbsQ+2bMPEz7HkBg+7OsZLS>YtZ?P#KALk4aEBH+*950`6Q5|(hTQqJ!DVdb zTnF4z;EPz<*4KsfvAvw@g6v=L)Ct=A1iR>siiu3a8dnb9%$s7jw=BuJeyrs?Qg$@y&yj^M~u3=UgD1 z>H+71;L>7Wqvi~TQ-8|25V*tJ;;23y+(RAb!r>;)aV`=rpevr5GYZadB7t%-aMRU^ zl#7MiewTA`aC1&cR9`%t>Tk{^!5O{ZqWY5ImNp!rT#7iq*OMui3g=Pu-%DopcoZCQ z5n?4QT|Y~QvktyP^<~0oCXZ4s3vR+8m2%mlul+dXa^RH8_b8VSH@owF$`!&LyUV!} zxVbMrp!&+;+=?HICmRPyUe*eaI=27)SUBhnv;2yTY&Sr zkWaaLaN73ID0d&u&9#7XkKnA)g_L^?SCPm$yLD1c{Yz)4J_oqgS4$}82p6&GEajZx zOi7$`g)>~`oI9Mtvy7V46K=YNbKY>Zlj2IRR4ey;osXFFx6i3Lec`Otm6Y>`Gj@MT zxj?wVp!1Xqf?NEMbHQ--bDRr-)23BXbL!x3?5n0+IGn!V0_7s%0#03|TohcbQw`-} z;BJL~MY&kGn=zLt7YAoLS4+8gxals=CBfwla4s3{;{4atoGEa|&<4t-!YOMTDVGkn z^ni1jaI^JIR9_aHVY!)d*>Lrvt(41w^J{6NT)sHJEte@*2se?#xe~ae>FrcsIb5;f z3gs%{^v&WwP!gB>T&aR9=;mAvoJ;9dYR)>ih<7_F*8peDy#UZ8~edTISZUY^CRV^;P(8=xf!@4fBp17eg6PBzUM9g diff --git a/core.725 b/core.725 deleted file mode 100644 index 4770ba1b03284a9e99dd0e5412e85d2aa84c49c2..0000000000000000000000000000000000000000 GIT binary patch literal 0 HcmV?d00001 literal 10240 zcmbW7JFn$56oq|9!=obw1t0{LK)f0Z;wdpGj06QB5eXp%naplmik>B;FnOY_FBEcj7rYbBeot6MtT7FZ*QfIhrqCeD$RZXJ>}5i{^QA z?Kfje-iT^xo-q$z2p2Bm`j4MQ6Qbe$SE65{;iBP}k-?JDtT4QMC`A|_X5VxelB+)W z^0!CAJ^6`w#$vUIyZJDofu8r<9QlZo9$RpY;wbGg$Z@mcsAJch^w&YJ_$c*By>iWo zPjXd;qi*C=Rw7sL=Mp`yIHeQspX@`=JQZir7i;8G^hR)<;;84qN1Wt2fg2Tv>1JAU zqDQX(czYlG5zc;+a`dU{!JXtX!>8NN-69rn-pl=*=oR29#aZ;l8s#aR1LqZ2-5o&I zh?6{J#&}-nX)%16L^ya)w}}C+B71Zu@HU?*i&Zoa9nMuino|e-${N z;cz>C-%1NLZnTP%o991fU|tp4OkI-hNK{Yru6G&ckrzQ?9h&3YYrC^lX3R zOqFt2pE!o8us?`y0TyI`w((M_#%8BWIhGJJ#Q5!@6!iC+B4X zS1K;7W{i77;#=iO&j0*%@;)-RgG7yT33^Y2`~FJGu|L>}^<*DraINC#v&TT|kt=^m z^~i1LiBEF1;^0*6lcWEIgA={a;17UEbM!)C zX9;dr9H!GB>#~33%)h7QLatEUv0N(nwEH>9r3U9RoXh0m!PSamF7=udJ#s^avq!i| zamR9LxPOYH4lCAK_OAn1{Fm+@^)@&;FFo^A9NID)c}gzi>I}D`mvD{Z=u_3@xir9M zyPp%i5nPwy`ta>s{C<#JCUDi+H|gKk2&TR>4B&U!2Xg&g+uV5T^||K0lydL0ru~tl zZp4LK&@16n%)_Zqiw0SBe>FI(xa;Uio*tZ29OSs)tcP@^KXT;_a+Tt)<5SLA z1E2MNPJFiDe1_Y|Q*!CRwTfdd8=UBo>oc6{bIFt3pg1)@-UrF0htKguZU8qa&fq!C z3jM|B@N!6CdMrJk^tq9Xu`E=lB#qo1HYs5*O?zOP+ z{8V~g=(B=u#K}1;!Ie*^9Htv{NaFkzaiX_y&unwGPDm9^ y(}0^Z+=ia`Bv<-Wk3Lmh@@%13DULd<;1_NJ*C}o}C_Z!=apH5D!}V_3^!Y!h=$O?2 From 36033df541fd345013149d4ea9376738663f4c79 Mon Sep 17 00:00:00 2001 From: lokeshbio Date: Tue, 16 May 2023 11:02:18 +0200 Subject: [PATCH 096/158] moved orf check from wf to groovy --- lib/WorkflowAmpliseq.groovy | 5 +++++ workflows/ampliseq.nf | 5 ----- 2 files changed, 5 insertions(+), 5 deletions(-) diff --git a/lib/WorkflowAmpliseq.groovy b/lib/WorkflowAmpliseq.groovy index 0162d5a4..5e4e4220 100755 --- a/lib/WorkflowAmpliseq.groovy +++ b/lib/WorkflowAmpliseq.groovy @@ -118,6 +118,11 @@ class WorkflowAmpliseq { if (params.addsh && params.cut_its == "none") { log.warn "Adding UNITE species hypothesis (SH) assignments is only feasible for ITS sequences. Please use option `--cut_its` to find ITS regions in the ASV sequences, unless the given sequences are already cut to the ITS region.\n" } + + // Error message for incompatible combination of --orf_start and --orf_end + if ( params.orf_end && ( ( ( params.orf_end + 1 ) - params.orf_start ) % 3 != 0 ) ) { + Nextflow.error("Incompatible parameters: The difference of `--orf_end` and `--orf_start` must be a multiple of 3.") + } } // diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index 97181fd9..35e6abf4 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -100,11 +100,6 @@ if ( params.dada_ref_taxonomy && !params.skip_dada_addspecies && !params.skip_ta } } -// Error message for incompatible combination of --orf_start and --orf_end -if ( params.orf_end && ( ( ( params.orf_end + 1 ) - params.orf_start ) % 3 != 0 ) ) { - Nextflow.error("Incompatible parameters: The difference of `--orf_end` and `--orf_start` must be a multiple of 3.") -} - //only run QIIME2 when taxonomy is actually calculated and all required data is available if ( !(workflow.profile.tokenize(',').intersect(['conda', 'mamba']).size() >= 1) && !params.skip_taxonomy && !params.skip_qiime ) { run_qiime2 = true From c98831a642c2d9a986b4b6e3f49533652326f484 Mon Sep 17 00:00:00 2001 From: lokeshbio Date: Tue, 16 May 2023 15:01:52 +0200 Subject: [PATCH 097/158] WorkflowAmpliseq.groovy error and changelog --- CHANGELOG.md | 24 ++++++++++++++++++++++++ lib/WorkflowAmpliseq.groovy | 2 +- 2 files changed, 25 insertions(+), 1 deletion(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index 0df376d7..524f9a90 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -3,6 +3,30 @@ The format is based on [Keep a Changelog](https://keepachangelog.com/en/1.0.0/) and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0.html). +## nf-core/ampliseq version 2.6.0dev + +### `Added` + +- [#564](https://github.com/nf-core/ampliseq/pull/564),[#567](https://github.com/nf-core/ampliseq/pull/567),[#582](https://github.com/nf-core/ampliseq/pull/582) - Added phylogenetic placement +- [#577](https://github.com/nf-core/ampliseq/pull/577),[#577](https://github.com/nf-core/ampliseq/pull/567) - Added SINTAX for taxonomic classification +- [#575](https://github.com/nf-core/ampliseq/pull/575) - Added filtering step for stop codons for ASVs that are of coding regions! + +### `Changed` + +- [#563](https://github.com/nf-core/ampliseq/pull/563) - Renamed DADA2 taxonomic classification files to include the chosen reference taxonomy abbreviation. +- [#567](https://github.com/nf-core/ampliseq/pull/567) - Renamed `--dada_tax_agglom_min` and `--qiime_tax_agglom_min` to `--tax_agglom_min` and `--dada_tax_agglom_max` and `--qiime_tax_agglom_max` to `--tax_agglom_max` + +### `Fixed` + +- [#553](https://github.com/nf-core/ampliseq/pull/553) - Handle empty barrnap results files +- [#554](https://github.com/nf-core/ampliseq/pull/554) - Accept taxonomy strings that contain `#`,`'` +- [#569](https://github.com/nf-core/ampliseq/pull/569) - Make header of overall_summary.tsv consistent between input data types +- [#573](https://github.com/nf-core/ampliseq/pull/573) - Avoid parser error for single-end data when an empty read file is detected +- [#578](https://github.com/nf-core/ampliseq/pull/578) - Template update for nf-core/tools version 2.8, including changing `System.exit(1)` to `Nextflow.error()` + +### `Dependencies` + +### `Removed` ## nf-core/ampliseq version 2.5.0 - 2023-03-02 ### `Added` diff --git a/lib/WorkflowAmpliseq.groovy b/lib/WorkflowAmpliseq.groovy index 5e4e4220..1d907bb2 100755 --- a/lib/WorkflowAmpliseq.groovy +++ b/lib/WorkflowAmpliseq.groovy @@ -119,7 +119,7 @@ class WorkflowAmpliseq { log.warn "Adding UNITE species hypothesis (SH) assignments is only feasible for ITS sequences. Please use option `--cut_its` to find ITS regions in the ASV sequences, unless the given sequences are already cut to the ITS region.\n" } - // Error message for incompatible combination of --orf_start and --orf_end + // Error message for incompatible combination of --orf_start and --orf_end if ( params.orf_end && ( ( ( params.orf_end + 1 ) - params.orf_start ) % 3 != 0 ) ) { Nextflow.error("Incompatible parameters: The difference of `--orf_end` and `--orf_start` must be a multiple of 3.") } From de49d4004c70f0e31b6b49796206d2bb60dcac72 Mon Sep 17 00:00:00 2001 From: daniel Date: Fri, 19 May 2023 16:23:15 +0200 Subject: [PATCH 098/158] activate codon filter and use pandas instead of QIIME2 environment --- bin/filt_codon_stats.py | 27 +++++++++++++++++++++++++++ bin/filt_codon_stats.r | 22 ---------------------- conf/modules.config | 4 ++-- conf/test_novaseq.config | 7 ++++++- docs/output.md | 2 +- modules/local/filter_codons.nf | 15 +++++++++------ workflows/ampliseq.nf | 4 +++- 7 files changed, 48 insertions(+), 33 deletions(-) create mode 100755 bin/filt_codon_stats.py delete mode 100755 bin/filt_codon_stats.r diff --git a/bin/filt_codon_stats.py b/bin/filt_codon_stats.py new file mode 100755 index 00000000..69728203 --- /dev/null +++ b/bin/filt_codon_stats.py @@ -0,0 +1,27 @@ +#!/usr/bin/env python3 + +import pandas as pd +import sys + +# argument check +if len(sys.argv) != 2: + exit("Usage: count_table_max_reads.py ") + +# read tsv and skip first two rows +data = pd.read_csv(sys.argv[1], sep="\t", skiprows=None) # count table + +# drop feature ids +df = data.drop(data.columns[0], axis=1) + +# sum each column +sums = df.sum(axis=0) + +# add column with sample names at beginning +out = sums.rename_axis("sample").reset_index() + +# rename columns +out = out.rename(columns={0: "encoding"}) + +# write file +out.to_csv("codon.filtered.stats.tsv", sep="\t", index=False) + diff --git a/bin/filt_codon_stats.r b/bin/filt_codon_stats.r deleted file mode 100755 index 497338de..00000000 --- a/bin/filt_codon_stats.r +++ /dev/null @@ -1,22 +0,0 @@ -#!/usr/bin/env Rscript - -args = commandArgs(trailingOnly=TRUE) - -if(length(args) < 2){ - stop("Usage: filt_codons_stats.r ") -} - -dada2_stats <- args[1] -coi_filt <- args[2] - -old_stats <- read.table(dada2_stats, header = T, check.names = F, row.names = 1) - -filt_table <- read.table(coi_filt, header = T, check.names = F, row.names = 1) - -filt_counts <- as.data.frame(colSums(filt_table)) -names(filt_counts) <- "codon_filtered" - -new_stats <- merge(old_stats, filt_counts, by = 'row.names') - -write.table(new_stats, file = "codon.filtered.stats.tsv", row.names=FALSE, sep="\t", col.names = TRUE, quote = FALSE, na = '') - diff --git a/conf/modules.config b/conf/modules.config index debda5f1..95d8569a 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -315,13 +315,13 @@ process { params.stop_codons ? "-x ${params.stop_codons}" : '' ].join(' ') publishDir = [ - path: { "${params.outdir}/Codons_filtered" }, + path: { "${params.outdir}/codon_filter" }, mode: params.publish_dir_mode, saveAs: { filename -> filename.equals('versions.yml') ? null : filename } ] } - withName: 'MERGE_STATS_STD|MERGE_STATS_FILTERSSU|MERGE_STATS_FILTERTAXA' { + withName: 'MERGE_STATS_STD|MERGE_STATS_FILTERSSU|MERGE_STATS_FILTERTAXA|MERGE_STATS_CODONS' { publishDir = [ path: { "${params.outdir}" }, mode: params.publish_dir_mode, diff --git a/conf/test_novaseq.config b/conf/test_novaseq.config index a8fa1263..b82e4d27 100644 --- a/conf/test_novaseq.config +++ b/conf/test_novaseq.config @@ -12,7 +12,7 @@ params { config_profile_name = 'Test profile Novaseq data' - config_profile_description = 'Minimal test dataset to check pipeline function with Novaseq data' + config_profile_description = 'Minimal test dataset to check pipeline function with Novaseq data and codon filtering' // Limit resources so that this can run on GitHub Actions max_cpus = 2 @@ -23,6 +23,11 @@ params { input = "https://raw.githubusercontent.com/nf-core/test-datasets/ampliseq/samplesheets/Samplesheet_novaseq.tsv" illumina_novaseq = true + // Codon filter, only for the first 2 codons (because otherwise all is filtered) + filter_codons = true + orf_start = 1 + orf_end = 6 + // Skip steps skip_cutadapt = true skip_barrnap = true diff --git a/docs/output.md b/docs/output.md index 18f1c693..e8d8f896 100644 --- a/docs/output.md +++ b/docs/output.md @@ -207,7 +207,7 @@ This filtering can be done by the setting the `--filter_codons`. By default, the
Output files -- `Codons_filtered/` +- `codons_filter/` - `ASV_codon_filtered.fna`: Fasta file of ASV sequences that passes the filter thresholds explained above. - `ASV_codon_filtered.table.tsv`: The count table of ASVs that successfully passed through the filter thresholds. - `ASV_codon_filtered.list`: List of ASV IDs that pass through the filter thresholds. diff --git a/modules/local/filter_codons.nf b/modules/local/filter_codons.nf index a0f19f5d..a3b51d2d 100644 --- a/modules/local/filter_codons.nf +++ b/modules/local/filter_codons.nf @@ -2,8 +2,10 @@ process FILTER_CODONS { tag "${fasta}" label 'process_low' - conda (params.enable_conda ? { exit 1 "QIIME2 has no conda package" } : null) - container "quay.io/qiime2/core:2022.8" + conda "conda-forge::pandas=1.1.5" + container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? + 'https://depot.galaxyproject.org/singularity/pandas:1.1.5': + 'quay.io/biocontainers/pandas:1.1.5' }" input: path(fasta) @@ -24,11 +26,12 @@ process FILTER_CODONS { def args = task.ext.args ?: '' """ filt_codons.py -f ${fasta} -t ${asv} -p ASV_codon ${args} - filt_codon_stats.r ${dada2stats} ASV_codon_filtered.table.tsv + filt_codon_stats.py ASV_codon_filtered.table.tsv + cat <<-END_VERSIONS > versions.yml - "${task.process}": + "${task.process}": python: \$(python --version 2>&1 | sed 's/Python //g') - R: \$(R --version | sed -n 1p | sed 's/R version //g' | sed 's/\\s.*\$//') - END_VERSIONS + pandas: \$(python -c "import pkg_resources; print(pkg_resources.get_distribution('pandas').version)") + END_VERSIONS """ } diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index ff185574..2dd78b3c 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -145,6 +145,7 @@ include { FILTER_SSU } from '../modules/local/filter_ssu' include { FILTER_LEN_ASV } from '../modules/local/filter_len_asv' include { MERGE_STATS as MERGE_STATS_FILTERSSU } from '../modules/local/merge_stats' include { MERGE_STATS as MERGE_STATS_FILTERLENASV } from '../modules/local/merge_stats' +include { MERGE_STATS as MERGE_STATS_CODONS } from '../modules/local/merge_stats' include { FILTER_CODONS } from '../modules/local/filter_codons' include { FORMAT_FASTAINPUT } from '../modules/local/format_fastainput' include { FORMAT_TAXONOMY } from '../modules/local/format_taxonomy' @@ -373,7 +374,8 @@ workflow AMPLISEQ { if (params.filter_codons ) { FILTER_CODONS ( ch_dada2_fasta, ch_dada2_asv, ch_stats ) ch_versions = ch_versions.mix(FILTER_CODONS.out.versions.ifEmpty(null)) - ch_stats = FILTER_CODONS.out.stats + MERGE_STATS_CODONS( ch_stats, FILTER_CODONS.out.stats ) + ch_stats = MERGE_STATS_CODONS.out.tsv ch_dada2_fasta = FILTER_CODONS.out.fasta ch_dada2_asv = FILTER_CODONS.out.asv } From f1638408c7edd89e64feead98bd349ff38b33ed2 Mon Sep 17 00:00:00 2001 From: daniel Date: Fri, 19 May 2023 16:30:36 +0200 Subject: [PATCH 099/158] update CHANGELOG and contributors --- CHANGELOG.md | 2 +- README.md | 1 + 2 files changed, 2 insertions(+), 1 deletion(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index 287fe43e..c8735fac 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -9,7 +9,7 @@ and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0 - [#564](https://github.com/nf-core/ampliseq/pull/564),[#567](https://github.com/nf-core/ampliseq/pull/567),[#582](https://github.com/nf-core/ampliseq/pull/582) - Added phylogenetic placement - [#577](https://github.com/nf-core/ampliseq/pull/577),[#577](https://github.com/nf-core/ampliseq/pull/567) - Added SINTAX for taxonomic classification -- [#575](https://github.com/nf-core/ampliseq/pull/575) - Added filtering step for stop codons for ASVs that are of coding regions! +- [#575](https://github.com/nf-core/ampliseq/pull/575), [#586](https://github.com/nf-core/ampliseq/pull/586) - Added filtering step for stop codons for ASVs that are of coding regions. ### `Changed` diff --git a/README.md b/README.md index 0befb267..1fb0499a 100644 --- a/README.md +++ b/README.md @@ -89,6 +89,7 @@ We thank the following people for their extensive assistance in the development - [Diego Brambilla](https://github.com/DiegoBrambilla) - [Emelie Nilsson](https://github.com/emnilsson) - [Jeanette Tångrot](https://github.com/jtangrot) +- [Lokeshwaran Manoharan](https://github.com/lokeshbio) - [Marissa Dubbelaar](https://github.com/marissaDubbelaar) - [Sabrina Krakau](https://github.com/skrakau) From 3991be8f7260b3063fa7bddf3fbc1833f1b612b3 Mon Sep 17 00:00:00 2001 From: daniel Date: Fri, 19 May 2023 16:39:58 +0200 Subject: [PATCH 100/158] update output docs --- docs/output.md | 8 +++----- 1 file changed, 3 insertions(+), 5 deletions(-) diff --git a/docs/output.md b/docs/output.md index e8d8f896..624b9fc0 100644 --- a/docs/output.md +++ b/docs/output.md @@ -198,16 +198,14 @@ Optionally, the ITS region can be extracted from each ASV sequence using ITSx, a #### Codons -Optionally, the ASVs can be filtered if they contain stop codons (`"TAA,TAG"`) in the specified open reading frame of the ASV. The filtering step also filters ASVs in the specifed checking range, if they are not multiple of 3. +Optionally, the ASVs can be filtered against the presence of stop codons in the specified open reading frame of the ASV. The filtering step can also filter out ASVs that are not multiple of 3 in length. -This filtering can be done by the setting the `--filter_codons`. By default, the codons are calculated and checked from the beginning to the end of the ASV sequence and the filter also checks if the length is of multiple of 3. By default the stop codons that are being screend for is `TAA` and `TAG`. These settings can be changed by `--orf_start` option to change the starting position of the codon on the ASV where you would like to start checking of codon usage! For example, setting this parameter to `1`, would start checking the ASV for codons from the beginning of the ASV or if it is set to `22`, the check starts the open reading frame from position 22 on each ASV. By default, the filtering applies from the `--orf_start` to the end of the ASV. But, this can be changed by setting the `--orf_end` to specify the end of the open reading frame where you want to look for stop codons. To look for different stop codons than default, `--stop_codons` can be used by specifying other single/multiple stop codons as a comma-separated list with this parameter like: `--stop_codons "TAA,TAG,TGA"` - -> **NB:** just make sure that when you set `--orf_start` and/or `--orf_end`, the length between `--orf_start` and `--orf_end` is a multiple of 3, otherwise all ASVs would be filtered. +Codon filtering can be activated by `--filter_codons`. By default, the codons are calculated and checked from the beginning (`--orf_start 1`) to the end (`--orf_end` not set) of the ASV sequence and the filter also checks if the length is a multiple of 3. By default, the filter screens for the presence of the stop codons `TAA` and `TAG`. To define different stop codons than default, use `--stop_codons`(comma-separated list, e.g. `--stop_codons "TAA,TAG,TGA"`).
Output files -- `codons_filter/` +- `codon_filter/` - `ASV_codon_filtered.fna`: Fasta file of ASV sequences that passes the filter thresholds explained above. - `ASV_codon_filtered.table.tsv`: The count table of ASVs that successfully passed through the filter thresholds. - `ASV_codon_filtered.list`: List of ASV IDs that pass through the filter thresholds. From 0e99b78c300291e20c23378b238e980dc7f58e62 Mon Sep 17 00:00:00 2001 From: daniel Date: Fri, 19 May 2023 16:50:46 +0200 Subject: [PATCH 101/158] update parameter description --- nextflow_schema.json | 14 +++++++------- 1 file changed, 7 insertions(+), 7 deletions(-) diff --git a/nextflow_schema.json b/nextflow_schema.json index d7e138de..250797a4 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -381,24 +381,24 @@ "filter_codons": { "type": "boolean", "description": "Filter ASVs based on codon usage", - "help_text": "This setting is to be used when the ASVs need to be filtered if they contain STOP codons in their sequence and they are of length in the multiple of 3." + "help_text": "ASVs will be filtered to contain no stop codon in their coding sequence and that their length is a multiple of 3." }, "orf_start": { "type": "integer", "default": 1, - "description": "Starting position to check for the stop codons in the ASVs", - "help_text": "By default, when the `--filter_codons` is triggered, the codons start from the beginning of the ASV sequences. If you would like to change this setting, you can specify from which position of the ASV does the codon triplets be calculated and checked. \n\nPlease note that the length from this position to the end of the ASV or until the `--orf_end` should be a multiple of 3 to make sure the filtering works properly." + "description": "Starting position of codon tripletts", + "help_text": "By default, when `--filter_codons` is set, the codons start from the first position of the ASV sequences. The start of the codons can be changed to any position." }, "orf_end": { "type": "integer", - "description": "Ending position to check for the stop codons in the ASVs", - "help_text": "By default, when the `--filter_codons` is triggered, the codons are checked until the end of the ASV sequences. If you would like to change this setting, you can specify until which position of the ASV sequences does the codon triplets be checked.\n\nPlease note that the length of the ASV from the beginning or from the `--orf_start` until this position should be a multiple of 3 to make sure the filtering works properly." + "description": "Ending position of codon tripletts", + "help_text": "By default, when `--filter_codons` is set, the codons are checked until the end of the ASV sequences. If you would like to change this setting, you can specify until which position of the ASV sequences does the codon triplets be checked.\n\nPlease note that the length of the ASV from the beginning or from the `--orf_start` until this position must be a multiple of 3." }, "stop_codons": { "type": "string", "default": "TAA,TAG", - "description": "Specific stop codons to check for in the ASV", - "help_text": "By default, when the `--filter_codons` is triggered, the codons `TAA,TAG` are checked in the ASVs that are to be filtered. If there are other single/multiple stop codons that needs to be checked, the codons can be specified as a comma-separated list with this parameter like: `--stop_codons \"TAA,TAG,TGA\"`" + "description": "Define stop codons", + "help_text": "By default, when `--filter_codons` is set, the codons `TAA,TAG` are set as stop codons. Here you can specify any comma-separated list of codons to be used as stop codons, e.g. `--stop_codons \"TAA,TAG,TGA\"`" }, "exclude_taxa": { "type": "string", From 011bdf984c97c47828ff10b14a4678b5e398da3b Mon Sep 17 00:00:00 2001 From: daniel Date: Fri, 19 May 2023 17:05:03 +0200 Subject: [PATCH 102/158] fix black formatting --- bin/filt_codon_stats.py | 1 - 1 file changed, 1 deletion(-) diff --git a/bin/filt_codon_stats.py b/bin/filt_codon_stats.py index 69728203..2efd2712 100755 --- a/bin/filt_codon_stats.py +++ b/bin/filt_codon_stats.py @@ -24,4 +24,3 @@ # write file out.to_csv("codon.filtered.stats.tsv", sep="\t", index=False) - From 72bdd43d377d0c7f57ff0cbe79b26d9bb00613e6 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Wed, 24 May 2023 12:44:34 +0200 Subject: [PATCH 103/158] Add paths to UNITE 9.0 to ref_databases --- conf/ref_databases.config | 36 ++++++++++++++++++++++++++---------- 1 file changed, 26 insertions(+), 10 deletions(-) diff --git a/conf/ref_databases.config b/conf/ref_databases.config index a8b55ba2..037fa5f4 100644 --- a/conf/ref_databases.config +++ b/conf/ref_databases.config @@ -161,12 +161,20 @@ params { dbversion = "SILVA v132 (https://zenodo.org/record/1172783)" } 'unite-fungi' { - title = "UNITE general FASTA release for Fungi - Version 8.3" - file = [ "https://files.plutof.ut.ee/public/orig/7B/23/7B235835FAF5C85D7B01E40FEF17F687914CB81A182554C5BD95E3168328E604.tgz" ] - citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2021): UNITE general FASTA release for Fungi. UNITE Community. 10.15156/BIO/1280049" + title = "UNITE general FASTA release for Fungi - Version 9.0" + file = [ "https://files.plutof.ut.ee/public/orig/1E/25/1E25CA4CC30A31C2E2B8CB2C89824C83D080A7F5A62E6263A0E95B37C6628067.tgz" ] + citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2022): UNITE general FASTA release for Fungi. Version 16.10.2022. UNITE Community. https://doi.org/10.15156/BIO/2483911" fmtscript = "taxref_reformat_unite.sh" - dbversion = "UNITE-fungi v8.3 (https://doi.org/10.15156/BIO/1280049)" - shfile = [ "https://scilifelab.figshare.com/ndownloader/files/34497977", "https://scilifelab.figshare.com/ndownloader/files/34497980"] + dbversion = "UNITE-fungi v9.0 (https://doi.org/10.15156/BIO/2483911)" + shfile = [ "https://figshare.scilifelab.se/ndownloader/files/40788767", "https://figshare.scilifelab.se/ndownloader/files/40788770"] + } + 'unite-fungi=9.0' { + title = "UNITE general FASTA release for Fungi - Version 9.0" + file = [ "https://files.plutof.ut.ee/public/orig/1E/25/1E25CA4CC30A31C2E2B8CB2C89824C83D080A7F5A62E6263A0E95B37C6628067.tgz" ] + citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2022): UNITE general FASTA release for Fungi. Version 16.10.2022. UNITE Community. https://doi.org/10.15156/BIO/2483911" + fmtscript = "taxref_reformat_unite.sh" + dbversion = "UNITE-fungi v9.0 (https://doi.org/10.15156/BIO/2483911)" + shfile = [ "https://figshare.scilifelab.se/ndownloader/files/40788767", "https://figshare.scilifelab.se/ndownloader/files/40788770"] } 'unite-fungi=8.3' { title = "UNITE general FASTA release for Fungi - Version 8.3" @@ -185,12 +193,20 @@ params { shfile = [ "https://scilifelab.figshare.com/ndownloader/files/34497971", "https://scilifelab.figshare.com/ndownloader/files/34497974"] } 'unite-alleuk' { - title = "UNITE general FASTA release for eukaryotes - Version 8.3" - file = [ "https://files.plutof.ut.ee/public/orig/E5/F5/E5F5E426DEC78BA2F7EC530621DDBD3F10564A09CBC2A5C4D3B3CBE7E37C5E1A.tgz" ] - citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2021): UNITE general FASTA release for eukaryotes. UNITE Community. 10.15156/BIO/1280127" + title = "UNITE general FASTA release for eukaryotes - Version 9.0" + file = [ "https://files.plutof.ut.ee/public/orig/F2/62/F262D942DEB8CAB3AEB9F313F67B04050E364B72E6707F99755DDCB271C45A48.tgz" ] + citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2022): UNITE general FASTA release for eukaryotes. Version 16.10.2022. UNITE Community. https://doi.org/10.15156/BIO/2483913" fmtscript = "taxref_reformat_unite.sh" - dbversion = "UNITE-alleuk v8.3 (https://doi.org/10.15156/BIO/1280127)" - shfile = [ "https://scilifelab.figshare.com/ndownloader/files/34994575", "https://scilifelab.figshare.com/ndownloader/files/34994578"] + dbversion = "UNITE-alleuk v9.0 (https://doi.org/10.15156/BIO/2483913)" + shfile = [ "https://figshare.scilifelab.se/ndownloader/files/40788773", "https://figshare.scilifelab.se/ndownloader/files/40788776"] + } + 'unite-alleuk=9.0' { + title = "UNITE general FASTA release for eukaryotes - Version 9.0" + file = [ "https://files.plutof.ut.ee/public/orig/F2/62/F262D942DEB8CAB3AEB9F313F67B04050E364B72E6707F99755DDCB271C45A48.tgz" ] + citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2022): UNITE general FASTA release for eukaryotes. Version 16.10.2022. UNITE Community. https://doi.org/10.15156/BIO/2483913" + fmtscript = "taxref_reformat_unite.sh" + dbversion = "UNITE-alleuk v9.0 (https://doi.org/10.15156/BIO/2483913)" + shfile = [ "https://figshare.scilifelab.se/ndownloader/files/40788773", "https://figshare.scilifelab.se/ndownloader/files/40788776"] } 'unite-alleuk=8.3' { title = "UNITE general FASTA release for eukaryotes - Version 8.3" From b7f8938ca914eb870d6c20a31f1b0df579928811 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Thu, 25 May 2023 13:29:32 +0200 Subject: [PATCH 104/158] Fix reformatting script for new UNITE --- bin/taxref_reformat_unite.sh | 3 ++- 1 file changed, 2 insertions(+), 1 deletion(-) diff --git a/bin/taxref_reformat_unite.sh b/bin/taxref_reformat_unite.sh index 1aa4547f..0778dc47 100755 --- a/bin/taxref_reformat_unite.sh +++ b/bin/taxref_reformat_unite.sh @@ -5,10 +5,11 @@ # Untar the Unite file tar xzf *gz +mv */*[[:digit:]].fasta . # Remove leading "k__" and the like, remove ranks classified as "unknown", # and replace space with underscore to create assignTaxonomy.fna -cat */*[[:digit:]].fasta | sed '/^>/s/;k__.*//' | sed '/^>/s/[a-z]__unidentified//g' | sed '/^>/s/[a-z]__//g' | sed '/^>/s/ /_/g' > assignTaxonomy.fna +cat *[[:digit:]].fasta | sed '/^>/s/;k__.*//' | sed '/^>/s/[a-z]__unidentified//g' | sed '/^>/s/[a-z]__//g' | sed '/^>/s/ /_/g' > assignTaxonomy.fna # Reformat to addSpecies format sed 's/>\([^|]\+\)|\([^|]\+|[^|]\+\)|.*/>\2 \1/' assignTaxonomy.fna | sed '/^>/s/_/ /g' > addSpecies.fna From 9fa6c2d522d41d02afa2b099e24e633a4fb6786b Mon Sep 17 00:00:00 2001 From: jtangrot Date: Thu, 25 May 2023 14:11:55 +0200 Subject: [PATCH 105/158] Add new unite to schema --- lib/WorkflowAmpliseq.groovy | 2 +- nextflow_schema.json | 2 ++ 2 files changed, 3 insertions(+), 1 deletion(-) diff --git a/lib/WorkflowAmpliseq.groovy b/lib/WorkflowAmpliseq.groovy index 30c0ff2b..4a8190d6 100755 --- a/lib/WorkflowAmpliseq.groovy +++ b/lib/WorkflowAmpliseq.groovy @@ -88,7 +88,7 @@ class WorkflowAmpliseq { Nextflow.error("Incompatible parameters: `--filter_ssu` cannot be used with `--skip_barrnap` because filtering for SSU's depends on barrnap.") } - String[] sbdi_compatible_databases = ["coidb","coidb=221216","gtdb","gtdb=R07-RS207","gtdb=R06-RS202","gtdb=R05-RS95","midori2-co1","midori2-co1=gb250","pr2","pr2=4.14.0","pr2=4.13.0","rdp","rdp=18","sbdi-gtdb","sbdi-gtdb=R07-RS207-1","silva","silva=138","silva=132","unite-fungi","unite-fungi=8.3","unite-fungi=8.2","unite-alleuk","unite-alleuk=8.3","unite-alleuk=8.2"] + String[] sbdi_compatible_databases = ["coidb","coidb=221216","gtdb","gtdb=R07-RS207","gtdb=R06-RS202","gtdb=R05-RS95","midori2-co1","midori2-co1=gb250","pr2","pr2=4.14.0","pr2=4.13.0","rdp","rdp=18","sbdi-gtdb","sbdi-gtdb=R07-RS207-1","silva","silva=138","silva=132","unite-fungi","unite-fungi=9.0","unite-fungi=8.3","unite-fungi=8.2","unite-alleuk","unite-alleuk=9.0","unite-alleuk=8.3","unite-alleuk=8.2"] if ( params.sbdiexport && !Arrays.stream(sbdi_compatible_databases).anyMatch(entry -> params.dada_ref_taxonomy.toString().equals(entry)) ) { Nextflow.error("Incompatible parameters: `--sbdiexport` does not work with the chosen database of `--dada_ref_taxonomy`, because the expected taxonomic levels do not match.") } diff --git a/nextflow_schema.json b/nextflow_schema.json index d7e138de..d14f9dd1 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -232,9 +232,11 @@ "silva=132", "silva=138", "silva", + "unite-fungi=9.0", "unite-fungi=8.3", "unite-fungi=8.2", "unite-fungi", + "unite-alleuk=9.0", "unite-alleuk=8.3", "unite-alleuk=8.2", "unite-alleuk" From 5fadb81e3745069c86ba38bbbd44c2702a9ef754 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Thu, 25 May 2023 14:16:05 +0200 Subject: [PATCH 106/158] Add link to SH file generation --- bin/add_sh_to_taxonomy.py | 1 + docs/output.md | 2 +- 2 files changed, 2 insertions(+), 1 deletion(-) diff --git a/bin/add_sh_to_taxonomy.py b/bin/add_sh_to_taxonomy.py index 629329a3..77b7ec2c 100755 --- a/bin/add_sh_to_taxonomy.py +++ b/bin/add_sh_to_taxonomy.py @@ -2,6 +2,7 @@ # @author Jeanette Tångrot # Adds UNITE species hypothesis (SH) information to ASV table based on vsearch usearch_global results in blast6 format. +# The look-up files required (, ), see below, can be generated as described in https://github.com/biodiversitydata-se/unite-shinfo # # Usage: add_sh_to_taxonomy.py # Input: diff --git a/docs/output.md b/docs/output.md index 18f1c693..a86f5b74 100644 --- a/docs/output.md +++ b/docs/output.md @@ -251,7 +251,7 @@ Files when using ITSx: #### assignSH -Optionally, a UNITE species hypothesis (SH) can be added to the DADA2 taxonomy. In short, the sequences are matched to the selected UNITE database with VSEARCH, using the sequence identity cutoff 98.5%. If a good enough and unique match is found, the sequence is assigned the SH of the matching sequence, and the taxonomy assignment for the sequence is changed to that of the SH. If no match is found, the taxonomy generated by assignTaxonomy and/or assignSpecies is kept. +Optionally, a UNITE species hypothesis (SH) can be added to the DADA2 taxonomy. In short, the sequences are matched to the selected UNITE database with VSEARCH, using the sequence identity cutoff 98.5%. If a good enough and unique match is found, the sequence is assigned the SH of the matching sequence, and the taxonomy assignment for the sequence is changed to that of the SH. The look-up tables for SH taxonomies are generated as descibed at https://github.com/biodiversitydata-se/unite-shinfo. If no match is found, the taxonomy generated by assignTaxonomy and/or assignSpecies is kept.
Output files From 0d933d30bad0db753d3ae60b143571dbdeabc92b Mon Sep 17 00:00:00 2001 From: jtangrot Date: Fri, 26 May 2023 08:36:44 +0200 Subject: [PATCH 107/158] Add sintax dbs for UNITE9.0 --- conf/ref_databases.config | 30 ++++++++++++++++++++++-------- nextflow_schema.json | 2 ++ 2 files changed, 24 insertions(+), 8 deletions(-) diff --git a/conf/ref_databases.config b/conf/ref_databases.config index 97bc2b74..1156005c 100644 --- a/conf/ref_databases.config +++ b/conf/ref_databases.config @@ -293,11 +293,18 @@ params { taxlevels = "Kingdom,Phylum,Class,Order,Family,Genus,Species,BOLD_bin" } 'unite-fungi' { - title = "UNITE USEARCH/UTAX release for Fungi - Version 8.3" - file = [ "https://files.plutof.ut.ee/public/orig/82/CB/82CB44BBAAA7D3AEAC297B5689BDA2963E8D0666E01FE0B54096147AFAF85263.gz" ] - citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2021): UNITE USEARCH/UTAX release for Fungi. UNITE Community. 10.15156/BIO/1280276" + title = "UNITE USEARCH/UTAX release for Fungi - Version 9.0" + file = [ "https://files.plutof.ut.ee/public/orig/19/1B/191B0D889A6B7B05DF4C103B118ABB3E0CF8EDBEA5B3E3FAB3EAFE3B72D7F3C8.gz" ] + citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2022): UNITE USEARCH/UTAX release for Fungi. Version 16.10.2022. UNITE Community. https://doi.org/10.15156/BIO/2483923" fmtscript = "taxref_reformat_sintax.sh" - dbversion = "UNITE-fungi v8.3 (https://dx.doi.org/10.15156/BIO/1280276)" + dbversion = "UNITE-fungi v9.0 (https://doi.org/10.15156/BIO/2483923)" + } + 'unite-fungi=9.0' { + title = "UNITE USEARCH/UTAX release for Fungi - Version 9.0" + file = [ "https://files.plutof.ut.ee/public/orig/19/1B/191B0D889A6B7B05DF4C103B118ABB3E0CF8EDBEA5B3E3FAB3EAFE3B72D7F3C8.gz" ] + citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2022): UNITE USEARCH/UTAX release for Fungi. Version 16.10.2022. UNITE Community. https://doi.org/10.15156/BIO/2483923" + fmtscript = "taxref_reformat_sintax.sh" + dbversion = "UNITE-fungi v9.0 (https://doi.org/10.15156/BIO/2483923)" } 'unite-fungi=8.3' { title = "UNITE USEARCH/UTAX release for Fungi - Version 8.3" @@ -314,11 +321,18 @@ params { dbversion = "UNITE-fungi v8.2 (https://doi.org/10.15156/BIO/786375)" } 'unite-alleuk' { - title = "UNITE USEARCH/UTAX release for eukaryotes - Version 8.3" - file = [ "https://files.plutof.ut.ee/public/orig/B9/35/B9351C91550A52713CB66DB7A1CEF35765310EBB23B6667AC93E714E9A9D020B.gz"] - citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2021): UNITE USEARCH/UTAX release for eukaryotes. UNITE Community. 10.15156/BIO/1280317" + title = "UNITE USEARCH/UTAX release for eukaryotes - Version 9.0" + file = [ "https://files.plutof.ut.ee/public/orig/AB/8C/AB8C119FC82CF5AFAFCB93CA4FFFF2B42A03CF1275DE23F60B887392E8FDEA21.gz"] + citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2022): UNITE USEARCH/UTAX release for eukaryotes. Version 16.10.2022. UNITE Community. https://doi.org/10.15156/BIO/2483924" fmtscript = "taxref_reformat_sintax.sh" - dbversion = "UNITE-alleuk v8.3 (https://doi.org/10.15156/BIO/1280127)" + dbversion = "UNITE-alleuk v9.0 (https://doi.org/10.15156/BIO/2483924)" + } + 'unite-alleuk=9.0' { + title = "UNITE USEARCH/UTAX release for eukaryotes - Version 9.0" + file = [ "https://files.plutof.ut.ee/public/orig/AB/8C/AB8C119FC82CF5AFAFCB93CA4FFFF2B42A03CF1275DE23F60B887392E8FDEA21.gz"] + citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2022): UNITE USEARCH/UTAX release for eukaryotes. Version 16.10.2022. UNITE Community. https://doi.org/10.15156/BIO/2483924" + fmtscript = "taxref_reformat_sintax.sh" + dbversion = "UNITE-alleuk v9.0 (https://doi.org/10.15156/BIO/2483924)" } 'unite-alleuk=8.3' { title = "UNITE USEARCH/UTAX release for eukaryotes - Version 8.3" diff --git a/nextflow_schema.json b/nextflow_schema.json index d14f9dd1..c070aee9 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -316,9 +316,11 @@ "enum": [ "coidb", "coidb=221216", + "unite-fungi=9.0", "unite-fungi=8.3", "unite-fungi=8.2", "unite-fungi", + "unite-alleuk=9.0", "unite-alleuk=8.3", "unite-alleuk=8.2", "unite-alleuk" From 5765e3c8a9b72ac2f9b7c60cac62b3177f156876 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Fri, 26 May 2023 09:07:26 +0200 Subject: [PATCH 108/158] Add qiime dbs for UNITE 8.3 & 9.0 --- bin/taxref_reformat_qiime_unite.sh | 7 ++--- conf/ref_databases.config | 42 +++++++++++++++++++++++------- 2 files changed, 37 insertions(+), 12 deletions(-) diff --git a/bin/taxref_reformat_qiime_unite.sh b/bin/taxref_reformat_qiime_unite.sh index 6fd80ac9..f863ec65 100755 --- a/bin/taxref_reformat_qiime_unite.sh +++ b/bin/taxref_reformat_qiime_unite.sh @@ -1,8 +1,9 @@ #!/bin/sh # Untar the Unite file -tar xzf *.gz +tar xzf *gz +mv */*_dynamic_*.fasta . # Select and rename dynamic files -cat */*_dynamic_*.fasta > unite.fna -cat */*_dynamic_*.txt > unite.tax +cat *_dynamic_*.fasta > unite.fna +cat *_dynamic_*.txt > unite.tax diff --git a/conf/ref_databases.config b/conf/ref_databases.config index 1156005c..06b4777c 100644 --- a/conf/ref_databases.config +++ b/conf/ref_databases.config @@ -243,25 +243,49 @@ params { fmtscript = "taxref_reformat_qiime_silva138.sh" } //UNITE for QIIME2, see https://unite.ut.ee/repository.php + 'unite-fungi' { + title = "UNITE QIIME release for Fungi - Version 9.0" + file = [ "https://files.plutof.ut.ee/public/orig/59/12/591225E8985EFC44B595C79AF5F467421B4D9A95093A0811B13CB4CC13A6DA46.tgz" ] + citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2022): UNITE QIIME release for Fungi. Version 16.10.2022. UNITE Community. https://doi.org/10.15156/BIO/2483915" + fmtscript = "taxref_reformat_qiime_unite.sh" + } + 'unite-fungi=9.0' { + title = "UNITE QIIME release for Fungi - Version 9.0" + file = [ "https://files.plutof.ut.ee/public/orig/59/12/591225E8985EFC44B595C79AF5F467421B4D9A95093A0811B13CB4CC13A6DA46.tgz" ] + citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2022): UNITE QIIME release for Fungi. Version 16.10.2022. UNITE Community. https://doi.org/10.15156/BIO/2483915" + fmtscript = "taxref_reformat_qiime_unite.sh" + } + 'unite-fungi=8.3' { + title = "UNITE QIIME release for Fungi - Version 8.3" + file = [ "https://files.plutof.ut.ee/public/orig/C5/54/C5547B97AAA979E45F79DC4C8C4B12113389343D7588716B5AD330F8BDB300C9.tgz" ] + citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2021): UNITE QIIME release for Fungi. Version 10.05.2021. UNITE Community. https://doi.org/10.15156/BIO/1264708" + fmtscript = "taxref_reformat_qiime_unite.sh" + } 'unite-fungi=8.2' { title = "UNITE QIIME release for Fungi - Version 8.2" file = [ "https://files.plutof.ut.ee/public/orig/98/AE/98AE96C6593FC9C52D1C46B96C2D9064291F4DBA625EF189FEC1CCAFCF4A1691.gz" ] citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2020): UNITE QIIME release for Fungi. Version 04.02.2020. UNITE Community. https://doi.org/10.15156/BIO/786385" fmtscript = "taxref_reformat_qiime_unite.sh" } - 'unite-fungi' { - title = "UNITE QIIME release for Fungi - Version 8.2" - file = [ "https://files.plutof.ut.ee/public/orig/98/AE/98AE96C6593FC9C52D1C46B96C2D9064291F4DBA625EF189FEC1CCAFCF4A1691.gz" ] - citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2020): UNITE QIIME release for Fungi. Version 04.02.2020. UNITE Community. https://doi.org/10.15156/BIO/786385" + 'unite-alleuk' { + title = "UNITE QIIME release for eukaryotes - Version 9.0" + file = [ "https://files.plutof.ut.ee/public/orig/8F/FC/8FFCC8A730E50FEEF8CFFEEFEF02A22FBCF7E02B7FD31C6649754834D2CB0E6F.tgz" ] + citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2022): UNITE QIIME release for eukaryotes. Version 16.10.2022. UNITE Community. https://doi.org/10.15156/BIO/2483917" fmtscript = "taxref_reformat_qiime_unite.sh" } - 'unite-alleuk=8.2' { - title = "UNITE QIIME release for eukaryotes - Version 8.2" - file = [ "https://files.plutof.ut.ee/public/orig/6E/0E/6E0EDD5592003B47C70A1B384C3C784AA32B726AC861CD7E2BD22AEB0278675E.gz" ] - citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2020): UNITE QIIME release for eukaryotes. Version 04.02.2020. UNITE Community. https://doi.org/10.15156/BIO/786386" + 'unite-alleuk=9.0' { + title = "UNITE QIIME release for eukaryotes - Version 9.0" + file = [ "https://files.plutof.ut.ee/public/orig/8F/FC/8FFCC8A730E50FEEF8CFFEEFEF02A22FBCF7E02B7FD31C6649754834D2CB0E6F.tgz" ] + citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2022): UNITE QIIME release for eukaryotes. Version 16.10.2022. UNITE Community. https://doi.org/10.15156/BIO/2483917" fmtscript = "taxref_reformat_qiime_unite.sh" } - 'unite-alleuk' { + 'unite-alleuk=8.3' { + title = "UNITE QIIME release for eukaryotes - Version 8.3" + file = [ "https://files.plutof.ut.ee/public/orig/48/29/4829D91F763E20F0F4376A60AA53FC9FBE6029A7D1BDC1B45347DD64EDE5D560.tgz" ] + citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2021): UNITE QIIME release for eukaryotes. Version 10.05.2021. UNITE Community. https://doi.org/10.15156/BIO/1264819" + fmtscript = "taxref_reformat_qiime_unite.sh" + } + 'unite-alleuk=8.2' { title = "UNITE QIIME release for eukaryotes - Version 8.2" file = [ "https://files.plutof.ut.ee/public/orig/6E/0E/6E0EDD5592003B47C70A1B384C3C784AA32B726AC861CD7E2BD22AEB0278675E.gz" ] citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2020): UNITE QIIME release for eukaryotes. Version 04.02.2020. UNITE Community. https://doi.org/10.15156/BIO/786386" From 25944432bfa912b536af87998047b2d2aa087a07 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Fri, 26 May 2023 09:09:13 +0200 Subject: [PATCH 109/158] Add qiime dbs for UNITE 8.3 & 9.0 --- nextflow_schema.json | 4 ++++ 1 file changed, 4 insertions(+) diff --git a/nextflow_schema.json b/nextflow_schema.json index c070aee9..a0bd2591 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -297,8 +297,12 @@ "enum": [ "silva=138", "silva", + "unite-fungi=9.0", + "unite-fungi=8.3", "unite-fungi=8.2", "unite-fungi", + "unite-alleuk=9.0", + "unite-alleuk=8.3", "unite-alleuk=8.2", "unite-alleuk", "greengenes85" From f944c6c6e2001cfd818c8bc69f9fd7ea62eb0cbd Mon Sep 17 00:00:00 2001 From: jtangrot Date: Fri, 26 May 2023 09:29:30 +0200 Subject: [PATCH 110/158] Make qiime run even if skip_dada_taxonomy is set --- workflows/ampliseq.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index ff185574..b4c56143 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -61,7 +61,7 @@ if (params.dada_ref_tax_custom) { val_dada_ref_taxonomy = "none" } -if (params.qiime_ref_taxonomy && !params.skip_dada_taxonomy && !params.skip_taxonomy && !params.classifier) { +if (params.qiime_ref_taxonomy && !params.skip_taxonomy && !params.classifier) { ch_qiime_ref_taxonomy = Channel.fromList(params.qiime_ref_databases[params.qiime_ref_taxonomy]["file"]).map { file(it) } } else { ch_qiime_ref_taxonomy = Channel.empty() } From 54048cb84479b326dae37e31da20494739732f45 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Fri, 26 May 2023 10:11:31 +0200 Subject: [PATCH 111/158] Make qiime run even if skip_dada_taxonomy is set --- workflows/ampliseq.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index b4c56143..ca7949fc 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -118,7 +118,7 @@ if ( params.dada_ref_taxonomy && !params.skip_dada_addspecies && !params.skip_da } //only run QIIME2 when taxonomy is actually calculated and all required data is available -if ( !(workflow.profile.tokenize(',').intersect(['conda', 'mamba']).size() >= 1) && !params.skip_taxonomy && !params.skip_qiime && (!params.skip_dada_taxonomy || params.sintax_ref_taxonomy) ) { +if ( !(workflow.profile.tokenize(',').intersect(['conda', 'mamba']).size() >= 1) && !params.skip_taxonomy && !params.skip_qiime && (!params.skip_dada_taxonomy || params.sintax_ref_taxonomy || params.qiime_ref_taxonomy) ) { run_qiime2 = true } else { run_qiime2 = false From d3bdb056d759115bebced56c2f4f97f843562d7c Mon Sep 17 00:00:00 2001 From: jtangrot Date: Fri, 26 May 2023 10:18:07 +0200 Subject: [PATCH 112/158] Make qiime run even if skip_dada_taxonomy is set --- workflows/ampliseq.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/workflows/ampliseq.nf b/workflows/ampliseq.nf index ca7949fc..9190c6be 100644 --- a/workflows/ampliseq.nf +++ b/workflows/ampliseq.nf @@ -502,7 +502,7 @@ workflow AMPLISEQ { } else if ( params.pplace_tree && params.pplace_taxonomy) { log.info "Use EPA-NG / GAPPA taxonomy classification" ch_tax = QIIME2_INTAX ( ch_pplace_tax ).qza - } else if ( params.dada_ref_taxonomy ) { + } else if ( params.dada_ref_taxonomy && !params.skip_dada_taxonomy ) { log.info "Use DADA2 taxonomy classification" ch_tax = QIIME2_INTAX ( ch_dada2_tax ).qza } else if ( params.qiime_ref_taxonomy || params.classifier ) { From 25822d6c70a616ffb67a4e396463c392d7eb175a Mon Sep 17 00:00:00 2001 From: jtangrot Date: Fri, 26 May 2023 10:33:15 +0200 Subject: [PATCH 113/158] Fix bug in qiime reformatting --- bin/taxref_reformat_qiime_unite.sh | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/bin/taxref_reformat_qiime_unite.sh b/bin/taxref_reformat_qiime_unite.sh index f863ec65..07862048 100755 --- a/bin/taxref_reformat_qiime_unite.sh +++ b/bin/taxref_reformat_qiime_unite.sh @@ -2,7 +2,7 @@ # Untar the Unite file tar xzf *gz -mv */*_dynamic_*.fasta . +mv */*_dynamic_* . # Select and rename dynamic files cat *_dynamic_*.fasta > unite.fna From 009ff8ea349f50294c837b18c730c680aefde623 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Fri, 26 May 2023 13:27:29 +0200 Subject: [PATCH 114/158] Fix bug in qiime reformatting --- bin/taxref_reformat_qiime_unite.sh | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/bin/taxref_reformat_qiime_unite.sh b/bin/taxref_reformat_qiime_unite.sh index 07862048..5c140978 100755 --- a/bin/taxref_reformat_qiime_unite.sh +++ b/bin/taxref_reformat_qiime_unite.sh @@ -5,5 +5,5 @@ tar xzf *gz mv */*_dynamic_* . # Select and rename dynamic files -cat *_dynamic_*.fasta > unite.fna -cat *_dynamic_*.txt > unite.tax +cat *_dynamic_*[[:digit:]].fasta > unite.fna +cat *_dynamic_*[[:digit:]].txt > unite.tax From b8b4ddda6bea05828cc274aab7f10c5cc2a58e07 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Fri, 9 Jun 2023 13:41:20 +0200 Subject: [PATCH 115/158] Remove unite-fungi=9.0 from qiime reference databases --- conf/ref_databases.config | 12 +++--------- nextflow_schema.json | 1 - 2 files changed, 3 insertions(+), 10 deletions(-) diff --git a/conf/ref_databases.config b/conf/ref_databases.config index 06b4777c..1fb8c666 100644 --- a/conf/ref_databases.config +++ b/conf/ref_databases.config @@ -244,15 +244,9 @@ params { } //UNITE for QIIME2, see https://unite.ut.ee/repository.php 'unite-fungi' { - title = "UNITE QIIME release for Fungi - Version 9.0" - file = [ "https://files.plutof.ut.ee/public/orig/59/12/591225E8985EFC44B595C79AF5F467421B4D9A95093A0811B13CB4CC13A6DA46.tgz" ] - citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2022): UNITE QIIME release for Fungi. Version 16.10.2022. UNITE Community. https://doi.org/10.15156/BIO/2483915" - fmtscript = "taxref_reformat_qiime_unite.sh" - } - 'unite-fungi=9.0' { - title = "UNITE QIIME release for Fungi - Version 9.0" - file = [ "https://files.plutof.ut.ee/public/orig/59/12/591225E8985EFC44B595C79AF5F467421B4D9A95093A0811B13CB4CC13A6DA46.tgz" ] - citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2022): UNITE QIIME release for Fungi. Version 16.10.2022. UNITE Community. https://doi.org/10.15156/BIO/2483915" + title = "UNITE QIIME release for Fungi - Version 8.3" + file = [ "https://files.plutof.ut.ee/public/orig/C5/54/C5547B97AAA979E45F79DC4C8C4B12113389343D7588716B5AD330F8BDB300C9.tgz" ] + citation = "Abarenkov, Kessy; Zirk, Allan; Piirmann, Timo; Pöhönen, Raivo; Ivanov, Filipp; Nilsson, R. Henrik; Kõljalg, Urmas (2021): UNITE QIIME release for Fungi. Version 10.05.2021. UNITE Community. https://doi.org/10.15156/BIO/1264708" fmtscript = "taxref_reformat_qiime_unite.sh" } 'unite-fungi=8.3' { diff --git a/nextflow_schema.json b/nextflow_schema.json index a0bd2591..a87a2f14 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -297,7 +297,6 @@ "enum": [ "silva=138", "silva", - "unite-fungi=9.0", "unite-fungi=8.3", "unite-fungi=8.2", "unite-fungi", From f7f17a3f5144d827c7dac089642e49bea8d1e758 Mon Sep 17 00:00:00 2001 From: jtangrot Date: Fri, 9 Jun 2023 14:00:45 +0200 Subject: [PATCH 116/158] prettier --- docs/output.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/docs/output.md b/docs/output.md index a86f5b74..90d37ded 100644 --- a/docs/output.md +++ b/docs/output.md @@ -251,7 +251,7 @@ Files when using ITSx: #### assignSH -Optionally, a UNITE species hypothesis (SH) can be added to the DADA2 taxonomy. In short, the sequences are matched to the selected UNITE database with VSEARCH, using the sequence identity cutoff 98.5%. If a good enough and unique match is found, the sequence is assigned the SH of the matching sequence, and the taxonomy assignment for the sequence is changed to that of the SH. The look-up tables for SH taxonomies are generated as descibed at https://github.com/biodiversitydata-se/unite-shinfo. If no match is found, the taxonomy generated by assignTaxonomy and/or assignSpecies is kept. +Optionally, a UNITE species hypothesis (SH) can be added to the DADA2 taxonomy. In short, the sequences are matched to the selected UNITE database with VSEARCH, using the sequence identity cutoff 98.5%. If a good enough and unique match is found, the sequence is assigned the SH of the matching sequence, and the taxonomy assignment for the sequence is changed to that of the SH. The look-up tables for SH taxonomies are generated as descibed at https://github.com/biodiversitydata-se/unite-shinfo. If no match is found, the taxonomy generated by assignTaxonomy and/or assignSpecies is kept.
Output files From 36087193fe024eb94dc4c14ad1705d1c7ea80b4d Mon Sep 17 00:00:00 2001 From: jtangrot Date: Fri, 9 Jun 2023 14:12:42 +0200 Subject: [PATCH 117/158] Update CHANGELOG --- CHANGELOG.md | 1 + 1 file changed, 1 insertion(+) diff --git a/CHANGELOG.md b/CHANGELOG.md index 287fe43e..b9cd7bbe 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -7,6 +7,7 @@ and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0 ### `Added` +- [#591](https://github.com/nf-core/ampliseq/pull/591) - New version of the Unite taxonomy databases: 9.0 - [#564](https://github.com/nf-core/ampliseq/pull/564),[#567](https://github.com/nf-core/ampliseq/pull/567),[#582](https://github.com/nf-core/ampliseq/pull/582) - Added phylogenetic placement - [#577](https://github.com/nf-core/ampliseq/pull/577),[#577](https://github.com/nf-core/ampliseq/pull/567) - Added SINTAX for taxonomic classification - [#575](https://github.com/nf-core/ampliseq/pull/575) - Added filtering step for stop codons for ASVs that are of coding regions! From 590f8ce592f9ce046787e9bdba96b33c69c54ce6 Mon Sep 17 00:00:00 2001 From: daniel Date: Mon, 12 Jun 2023 16:28:07 +0200 Subject: [PATCH 118/158] fix metadata documentation --- CHANGELOG.md | 1 + docs/usage.md | 2 +- nextflow_schema.json | 2 +- 3 files changed, 3 insertions(+), 2 deletions(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index 287fe43e..acf34238 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -23,6 +23,7 @@ and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0 - [#569](https://github.com/nf-core/ampliseq/pull/569) - Make header of overall_summary.tsv consistent between input data types - [#573](https://github.com/nf-core/ampliseq/pull/573) - Avoid parser error for single-end data when an empty read file is detected - [#578](https://github.com/nf-core/ampliseq/pull/578) - Template update for nf-core/tools version 2.8, including changing `System.exit(1)` to `Nextflow.error()` +- [#594](https://github.com/nf-core/ampliseq/pull/594) - Update metadata documentation ### `Dependencies` diff --git a/docs/usage.md b/docs/usage.md index 54be4b91..73930402 100644 --- a/docs/usage.md +++ b/docs/usage.md @@ -222,7 +222,7 @@ Please note the following requirements: - The path must be enclosed in quotes - The metadata file has to follow the QIIME2 specifications (https://docs.qiime2.org/2021.2/tutorials/metadata/) -The metadata file must be tab-separated with a header line. The first column in the tab-separated metadata file is the sample identifier column (required header: ID) and defines the sample or feature IDs associated with the dataset. In addition to the sample identifier column, metadata files are required to have additional metadata columns. +The metadata file must be tab-separated with a header line. The first column in the tab-separated metadata file is the sample identifier column (required header: ID) and defines the sample or feature IDs associated with the dataset. In addition to the sample identifier column, the metadata file is required to have at least one column with multiple different non-numeric values but not all unique. Sample identifiers should be 36 characters long or less, and also contain only ASCII alphanumeric characters (i.e. in the range of [a-z], [A-Z], or [0-9]), or the dash (-) character. For downstream analysis, by default all numeric columns, blanks or NA are removed, and only columns with multiple different values but not all unique are selected. diff --git a/nextflow_schema.json b/nextflow_schema.json index d7e138de..257c718d 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -33,7 +33,7 @@ "metadata": { "type": "string", "description": "Path to metadata sheet, when missing most downstream analysis are skipped (barplots, PCoA plots, ...).", - "help_text": "This is optional, but for performing downstream analysis such as barplots, diversity indices or differential abundance testing, a metadata file is essential.\n\nRelated parameter:\n- `--metadata_category` (optional) to choose columns that are used for testing significance\n\nFor example:\n\n```bash\n--metadata \"path/to/metadata.tsv\"\n```\n\nPlease note the following requirements:\n\n1. The path must be enclosed in quotes\n2. The metadata file has to follow the QIIME2 specifications (https://docs.qiime2.org/2021.2/tutorials/metadata/)\n\nThe first column in the tab-separated metadata file is the sample identifier column (required header: `ID`) and defines the sample or feature IDs associated with your study. Metadata files are not required to have additional metadata columns, so a file containing only an ID column is a valid QIIME 2 metadata file. Additional columns defining metadata associated with each sample or feature ID are optional.\n**NB**: without additional columns there might be no groupings for the downstream analyses.\n\nSample identifiers should be 36 characters long or less, and also contain only ASCII alphanumeric characters (i.e. in the range of [a-z], [A-Z], or [0-9]), or the dash (-) character. For downstream analysis, by default all numeric columns, blanks or NA are removed, and only columns with multiple different values but not all unique are selected.\n\nThe columns which are to be assessed can be specified by `--metadata_category`. If `--metadata_category` isn't specified than all columns that fit the specification are automatically chosen.", + "help_text": "This is optional, but for performing downstream analysis such as barplots, diversity indices or differential abundance testing, a metadata file is essential.\n\nRelated parameter:\n- `--metadata_category` (optional) to choose columns that are used for testing significance\n\nFor example:\n\n```bash\n--metadata \"path/to/metadata.tsv\"\n```\n\nPlease note the following requirements:\n\n1. The path must be enclosed in quotes\n2. The metadata file has to follow the QIIME2 specifications (https://docs.qiime2.org/2021.2/tutorials/metadata/)\n\nThe first column in the tab-separated metadata file is the sample identifier column (required header: `ID`) and defines the sample or feature IDs associated with your study. In addition to the sample identifier column, the metadata file is required to have at least one column with multiple different non-numeric values but not all unique.\n**NB**: without additional columns there might be no groupings for the downstream analyses.\n\nSample identifiers should be 36 characters long or less, and also contain only ASCII alphanumeric characters (i.e. in the range of [a-z], [A-Z], or [0-9]), or the dash (-) character. For downstream analysis, by default all numeric columns, blanks or NA are removed, and only columns with multiple different values but not all unique are selected.\n\nThe columns which are to be assessed can be specified by `--metadata_category`. If `--metadata_category` isn't specified than all columns that fit the specification are automatically chosen.", "fa_icon": "fas fa-file-csv" }, "outdir": { From 2c38d61c56f9c93378d2905ce129c65b4f9a17d6 Mon Sep 17 00:00:00 2001 From: daniel Date: Mon, 12 Jun 2023 16:53:17 +0200 Subject: [PATCH 119/158] restructure reporting of mindepth in QIIME2_DIVERSITY_CORE --- CHANGELOG.md | 1 + modules/local/qiime2_diversity_core.nf | 15 +++++++++++---- 2 files changed, 12 insertions(+), 4 deletions(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index 287fe43e..dd5eba7b 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -23,6 +23,7 @@ and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0 - [#569](https://github.com/nf-core/ampliseq/pull/569) - Make header of overall_summary.tsv consistent between input data types - [#573](https://github.com/nf-core/ampliseq/pull/573) - Avoid parser error for single-end data when an empty read file is detected - [#578](https://github.com/nf-core/ampliseq/pull/578) - Template update for nf-core/tools version 2.8, including changing `System.exit(1)` to `Nextflow.error()` +- [#595](https://github.com/nf-core/ampliseq/pull/595) - Closing gaps in rarefaction depth for diversity calculations (`mindepth` in QIIME2_DIVERSITY_CORE) ### `Dependencies` diff --git a/modules/local/qiime2_diversity_core.nf b/modules/local/qiime2_diversity_core.nf index ebfbff2f..99fe9280 100644 --- a/modules/local/qiime2_diversity_core.nf +++ b/modules/local/qiime2_diversity_core.nf @@ -31,10 +31,17 @@ process QIIME2_DIVERSITY_CORE { mindepth=\$(count_table_minmax_reads.py $stats minimum 2>&1) if [ \"\$mindepth\" -lt \"$mindepth\" ]; then mindepth=$mindepth; fi - if [ \"\$mindepth\" -gt \"10000\" ]; then echo \$mindepth >\"Use the sampling depth of \$mindepth for rarefaction.txt\" ; fi - if [ \"\$mindepth\" -lt \"10000\" -a \"\$mindepth\" -gt \"5000\" ]; then echo \$mindepth >\"WARNING The sampling depth of \$mindepth is quite small for rarefaction.txt\" ; fi - if [ \"\$mindepth\" -lt \"5000\" -a \"\$mindepth\" -gt \"1000\" ]; then echo \$mindepth >\"WARNING The sampling depth of \$mindepth is very small for rarefaction.txt\" ; fi - if [ \"\$mindepth\" -lt \"1000\" ]; then echo \$mindepth >\"WARNING The sampling depth of \$mindepth seems too small for rarefaction.txt\" ; fi + + # report the rarefaction depth and return warning, if needed + if [ \"\$mindepth\" -lt \"1000\" ]; then + echo \$mindepth >\"WARNING The sampling depth of \$mindepth seems too small for rarefaction.txt\" + elif [ \"\$mindepth\" -lt \"5000\" ]; then + echo \$mindepth >\"WARNING The sampling depth of \$mindepth is very small for rarefaction.txt\" + elif [ \"\$mindepth\" -lt \"10000\" ]; then + echo \$mindepth >\"WARNING The sampling depth of \$mindepth is quite small for rarefaction.txt\" + else + echo \$mindepth >\"Use the sampling depth of \$mindepth for rarefaction.txt\" + fi qiime diversity core-metrics-phylogenetic \\ --m-metadata-file ${metadata} \\ From 070620463289bd1cd1e8f00404e63477c3336ae4 Mon Sep 17 00:00:00 2001 From: Daniel Straub <42973691+d4straub@users.noreply.github.com> Date: Tue, 13 Jun 2023 09:00:07 +0200 Subject: [PATCH 120/158] Update bin/filt_codon_stats.py --- bin/filt_codon_stats.py | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/bin/filt_codon_stats.py b/bin/filt_codon_stats.py index 2efd2712..5c9484df 100755 --- a/bin/filt_codon_stats.py +++ b/bin/filt_codon_stats.py @@ -7,7 +7,7 @@ if len(sys.argv) != 2: exit("Usage: count_table_max_reads.py ") -# read tsv and skip first two rows +# read tsv data = pd.read_csv(sys.argv[1], sep="\t", skiprows=None) # count table # drop feature ids From 10b2b995d7b0e5a2c3ed6f53e307459fd85b6090 Mon Sep 17 00:00:00 2001 From: Daniel Straub <42973691+d4straub@users.noreply.github.com> Date: Tue, 13 Jun 2023 09:02:08 +0200 Subject: [PATCH 121/158] Update nextflow_schema.json Co-authored-by: Till E. <64961761+tillenglert@users.noreply.github.com> --- nextflow_schema.json | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/nextflow_schema.json b/nextflow_schema.json index 250797a4..68f43542 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -392,7 +392,7 @@ "orf_end": { "type": "integer", "description": "Ending position of codon tripletts", - "help_text": "By default, when `--filter_codons` is set, the codons are checked until the end of the ASV sequences. If you would like to change this setting, you can specify until which position of the ASV sequences does the codon triplets be checked.\n\nPlease note that the length of the ASV from the beginning or from the `--orf_start` until this position must be a multiple of 3." + "help_text": "By default, when `--filter_codons` is set, the codons are checked until the end of the ASV sequences. If you would like to change this setting, you can specify until which position of the ASV sequences the codon triplets are checked.\n\nPlease note that the length of the ASV from the beginning or from the `--orf_start` until this position must be a multiple of 3." }, "stop_codons": { "type": "string", From 08892b15520ed5052b84d75c2622b82c37a6018a Mon Sep 17 00:00:00 2001 From: jtangrot Date: Tue, 13 Jun 2023 10:50:22 +0200 Subject: [PATCH 122/158] Specify unite version in test configs --- conf/test_iontorrent.config | 2 +- conf/test_pacbio_its.config | 2 +- 2 files changed, 2 insertions(+), 2 deletions(-) diff --git a/conf/test_iontorrent.config b/conf/test_iontorrent.config index 06f88af6..c4092fdd 100644 --- a/conf/test_iontorrent.config +++ b/conf/test_iontorrent.config @@ -22,7 +22,7 @@ params { // Input data FW_primer = "GTGARTCATCGARTCTTTG" RV_primer = "TCCTCSSCTTATTGATATGC" - dada_ref_taxonomy = "unite-fungi" + dada_ref_taxonomy = "unite-fungi=8.3" input = "https://raw.githubusercontent.com/nf-core/test-datasets/ampliseq/samplesheets/Samplesheet_it_SE_ITS.tsv" iontorrent = true max_ee = 5 diff --git a/conf/test_pacbio_its.config b/conf/test_pacbio_its.config index e4e4e2d2..f64706ea 100644 --- a/conf/test_pacbio_its.config +++ b/conf/test_pacbio_its.config @@ -22,7 +22,7 @@ params { // Input data FW_primer = "CTTGGTCATTTAGAGGAAGTAA" RV_primer = "TCCTGAGGGAAACTTCG" - dada_ref_taxonomy = "unite-fungi" + dada_ref_taxonomy = "unite-fungi=8.3" input = "https://raw.githubusercontent.com/nf-core/test-datasets/ampliseq/samplesheets/Samplesheet_pacbio_ITS.tsv" metadata = "https://raw.githubusercontent.com/nf-core/test-datasets/ampliseq/samplesheets/Metadata_pacbio_ITS.tsv" pacbio = true From e25a96a047b81cb0d0531c85e20c1d6d09bf0827 Mon Sep 17 00:00:00 2001 From: daniel Date: Thu, 15 Jun 2023 11:21:43 +0200 Subject: [PATCH 123/158] add pr2 v5.0.0 --- CHANGELOG.md | 1 + conf/ref_databases.config | 16 ++++++++++++---- nextflow_schema.json | 1 + 3 files changed, 14 insertions(+), 4 deletions(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index 48a95085..6e7f4604 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -8,6 +8,7 @@ and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0 ### `Added` - [#591](https://github.com/nf-core/ampliseq/pull/591) - New version of the Unite taxonomy databases: 9.0 +- [#596](https://github.com/nf-core/ampliseq/pull/596) - New version of the PR2 taxonomy database: 5.0.0, only available with DADA2 (`--dada_ref_taxonomy`) - [#564](https://github.com/nf-core/ampliseq/pull/564),[#567](https://github.com/nf-core/ampliseq/pull/567),[#582](https://github.com/nf-core/ampliseq/pull/582) - Added phylogenetic placement - [#577](https://github.com/nf-core/ampliseq/pull/577),[#577](https://github.com/nf-core/ampliseq/pull/567) - Added SINTAX for taxonomic classification - [#575](https://github.com/nf-core/ampliseq/pull/575), [#586](https://github.com/nf-core/ampliseq/pull/586) - Added filtering step for stop codons for ASVs that are of coding regions. diff --git a/conf/ref_databases.config b/conf/ref_databases.config index 1fb8c666..7a2417e5 100644 --- a/conf/ref_databases.config +++ b/conf/ref_databases.config @@ -70,12 +70,20 @@ params { dbversion = "GTDB R05-RS95 (https://data.ace.uq.edu.au/public/gtdb/data/releases/release95/95.0/)" } 'pr2' { - title = "PR2 - Protist Reference Ribosomal Database - Version 4.14.0" - file = [ "https://github.com/pr2database/pr2database/releases/download/v4.14.0/pr2_version_4.14.0_SSU_dada2.fasta.gz", "https://github.com/pr2database/pr2database/releases/download/v4.14.0/pr2_version_4.14.0_SSU_UTAX.fasta.gz" ] + title = "PR2 - Protist Reference Ribosomal Database - Version 5.0.0" + file = [ "https://github.com/pr2database/pr2database/releases/download/v5.0.0/pr2_version_5.0.0_SSU_dada2.fasta.gz", "https://github.com/pr2database/pr2database/releases/download/v5.0.0/pr2_version_5.0.0_SSU_UTAX.fasta.gz" ] citation = "Guillou L, Bachar D, Audic S, Bass D, Berney C, Bittner L, Boutte C, Burgaud G, de Vargas C, Decelle J, Del Campo J, Dolan JR, Dunthorn M, Edvardsen B, Holzmann M, Kooistra WH, Lara E, Le Bescot N, Logares R, Mahé F, Massana R, Montresor M, Morard R, Not F, Pawlowski J, Probert I, Sauvadet AL, Siano R, Stoeck T, Vaulot D, Zimmermann P, Christen R. The Protist Ribosomal Reference database (PR2): a catalog of unicellular eukaryote small sub-unit rRNA sequences with curated taxonomy. Nucleic Acids Res. 2013 Jan;41(Database issue):D597-604. doi: 10.1093/nar/gks1160. Epub 2012 Nov 27. PMID: 23193267; PMCID: PMC3531120." fmtscript = "taxref_reformat_pr2.sh" - dbversion = "PR2 v4.14.0 (https://github.com/pr2database/pr2database/releases/tag/v4.14.0)" - taxlevels = "Domain,Kingdom,Phylum,Class,Order,Family,Genus,Species" + dbversion = "PR2 v5.0.0 (https://github.com/pr2database/pr2database/releases/tag/v5.0.0)" + taxlevels = "Domain,Clade,Kingdom,Phylum,Class,Order,Family,Genus,Species" + } + 'pr2=5.0.0' { + title = "PR2 - Protist Reference Ribosomal Database - Version 5.0.0" + file = [ "https://github.com/pr2database/pr2database/releases/download/v5.0.0/pr2_version_5.0.0_SSU_dada2.fasta.gz", "https://github.com/pr2database/pr2database/releases/download/v5.0.0/pr2_version_5.0.0_SSU_UTAX.fasta.gz" ] + citation = "Guillou L, Bachar D, Audic S, Bass D, Berney C, Bittner L, Boutte C, Burgaud G, de Vargas C, Decelle J, Del Campo J, Dolan JR, Dunthorn M, Edvardsen B, Holzmann M, Kooistra WH, Lara E, Le Bescot N, Logares R, Mahé F, Massana R, Montresor M, Morard R, Not F, Pawlowski J, Probert I, Sauvadet AL, Siano R, Stoeck T, Vaulot D, Zimmermann P, Christen R. The Protist Ribosomal Reference database (PR2): a catalog of unicellular eukaryote small sub-unit rRNA sequences with curated taxonomy. Nucleic Acids Res. 2013 Jan;41(Database issue):D597-604. doi: 10.1093/nar/gks1160. Epub 2012 Nov 27. PMID: 23193267; PMCID: PMC3531120." + fmtscript = "taxref_reformat_pr2.sh" + dbversion = "PR2 v5.0.0 (https://github.com/pr2database/pr2database/releases/tag/v5.0.0)" + taxlevels = "Domain,SuperClade,Clade,Phylum,Class,Order,Family,Genus,Species" } 'pr2=4.14.0' { title = "PR2 - Protist Reference Ribosomal Database - Version 4.14.0" diff --git a/nextflow_schema.json b/nextflow_schema.json index 9d1b42f3..089e2d5f 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -220,6 +220,7 @@ "gtdb", "coidb", "coidb=221216", + "pr2=5.0.0", "pr2=4.14.0", "pr2=4.13.0", "pr2", From b095ebf94781093d0eb29a8b9386edd551c84d97 Mon Sep 17 00:00:00 2001 From: daniel Date: Thu, 15 Jun 2023 15:11:52 +0200 Subject: [PATCH 124/158] change from file size to read counts as sample read threshold --- CHANGELOG.md | 1 + nextflow.config | 1 + nextflow_schema.json | 13 +++++++++++-- subworkflows/local/cutadapt_workflow.nf | 8 ++++---- subworkflows/local/parse_input.nf | 8 ++++---- 5 files changed, 21 insertions(+), 10 deletions(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index 48a95085..21925306 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -11,6 +11,7 @@ and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0 - [#564](https://github.com/nf-core/ampliseq/pull/564),[#567](https://github.com/nf-core/ampliseq/pull/567),[#582](https://github.com/nf-core/ampliseq/pull/582) - Added phylogenetic placement - [#577](https://github.com/nf-core/ampliseq/pull/577),[#577](https://github.com/nf-core/ampliseq/pull/567) - Added SINTAX for taxonomic classification - [#575](https://github.com/nf-core/ampliseq/pull/575), [#586](https://github.com/nf-core/ampliseq/pull/586) - Added filtering step for stop codons for ASVs that are of coding regions. +- [#597](https://github.com/nf-core/ampliseq/pull/597) - Samples with less reads than specified with `--min_read_counts` (default: 1) stop the pipeline, previously the threshold was 1KB in size. ### `Changed` diff --git a/nextflow.config b/nextflow.config index ff7e3125..693b1a26 100644 --- a/nextflow.config +++ b/nextflow.config @@ -49,6 +49,7 @@ params { addsh = false tax_agglom_min = 2 tax_agglom_max = 6 + min_read_counts = 1 ignore_failed_trimming = false ignore_empty_input_files = false qiime_adonis_formula = null diff --git a/nextflow_schema.json b/nextflow_schema.json index 9d1b42f3..24b4d942 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -85,9 +85,17 @@ "description": "Naming of sequencing files", "help_text": "Indicates the naming of sequencing files (default: `\"/*_R{1,2}_001.fastq.gz\"`).\n\nPlease note:\n\n1. The prepended slash (`/`) is required\n2. The star (`*`) is the required wildcard for sample names\n3. The curly brackets (`{}`) enclose the orientation for paired end reads, separated by a comma (`,`).\n4. The pattern must be enclosed in quotes\n\nFor example for one sample (name: `1`) with forward (file: `1_a.fastq.gz`) and reverse (file: `1_b.fastq.gz`) reads in folder `data`:\n\n```bash\n--input \"data\" --extension \"/*_{a,b}.fastq.gz\"\n```" }, + "min_read_counts": { + "type": "integer", + "default": 1, + "description": "Set read count threshold for failed input samples.", + "help_text": "Samples with less reads than this threshold at input or after trimming stops the pipeline. Using `--ignore_empty_input_files` or `--ignore_failed_trimming ignores` samples with read numbers below the threshold and lets the pipeline continue with less samples.", + "fa_icon": "fas fa-greater-than-equal" + }, "ignore_empty_input_files": { "type": "boolean", - "description": "Ignore input files considered too small (<1KB) for individual samples and continue the pipeline without those samples." + "description": "Ignore input files with too less reads.", + "help_text": "Ignore input files with less reads than specified by `--min_read_counts` and continue the pipeline without those samples." } } }, @@ -121,7 +129,8 @@ }, "ignore_failed_trimming": { "type": "boolean", - "description": "Ignore files considered too small (<1KB) after trimming and continue the pipeline without those samples." + "description": "Ignore files with too less reads after trimming.", + "help_text": "Ignore files with less reads than specified by `--min_read_counts` after trimming and continue the pipeline without those samples." } } }, diff --git a/subworkflows/local/cutadapt_workflow.nf b/subworkflows/local/cutadapt_workflow.nf index 6f280b9d..1ce12a1d 100644 --- a/subworkflows/local/cutadapt_workflow.nf +++ b/subworkflows/local/cutadapt_workflow.nf @@ -50,8 +50,8 @@ workflow CUTADAPT_WORKFLOW { //Filter empty files ch_trimmed_reads .branch { - failed: it[0].single_end ? it[1].size() < 1.KB : it[1][0].size() < 1.KB || it[1][1].size() < 1.KB - passed: it[0].single_end ? it[1].size() >= 1.KB : it[1][0].size() >= 1.KB && it[1][1].size() >= 1.KB + failed: it[0].single_end ? it[1].countFastq() < params.min_read_counts : it[1][0].countFastq() < params.min_read_counts || it[1][1].countFastq() < params.min_read_counts + passed: true } .set { ch_trimmed_reads_result } ch_trimmed_reads_result.passed.set { ch_trimmed_reads_passed } @@ -61,9 +61,9 @@ workflow CUTADAPT_WORKFLOW { .subscribe { samples = it.join("\n") if (params.ignore_failed_trimming) { - log.warn "The following samples had too small file size (<1KB) after trimming with cutadapt:\n$samples\nIgnoring failed samples and continue!\n" + log.warn "The following samples had too less reads (<$params.min_read_counts) after trimming with cutadapt:\n$samples\nIgnoring failed samples and continue!\n" } else { - error("The following samples had too small file size (<1KB) after trimming with cutadapt:\n$samples\nPlease check whether the correct primer sequences for trimming were supplied. Ignore that samples using `--ignore_failed_trimming`.") + error("The following samples had too too less reads (<$params.min_read_counts) after trimming with cutadapt:\n$samples\nPlease check whether the correct primer sequences for trimming were supplied. Ignore that samples using `--ignore_failed_trimming` or adjust the threshold with `--min_read_counts`.") } } diff --git a/subworkflows/local/parse_input.nf b/subworkflows/local/parse_input.nf index 16f8ad64..5871d845 100644 --- a/subworkflows/local/parse_input.nf +++ b/subworkflows/local/parse_input.nf @@ -143,8 +143,8 @@ workflow PARSE_INPUT { //Filter empty files ch_reads.dump(tag:'parse_input.nf: ch_reads') .branch { - failed: it[0].single_end ? it[1].size() < 1.KB : it[1][0].size() < 1.KB || it[1][1].size() < 1.KB - passed: it[0].single_end ? it[1].size() >= 1.KB : it[1][0].size() >= 1.KB && it[1][1].size() >= 1.KB + failed: it[0].single_end ? it[1].countFastq() < params.min_read_counts : it[1][0].countFastq() < params.min_read_counts || it[1][1].countFastq() < params.min_read_counts + passed: true } .set { ch_reads_result } ch_reads_result.passed.set { ch_reads_passed } @@ -154,9 +154,9 @@ workflow PARSE_INPUT { .subscribe { samples = it.join("\n") if (params.ignore_empty_input_files) { - log.warn "At least one input file for the following sample(s) was too small (<1KB):\n$samples\nIgnoring failed samples and continue!\n" + log.warn "At least one input file for the following sample(s) had too less reads (<$params.min_read_counts):\n$samples\nThe threshold can be adjusted with `--min_read_counts`. Ignoring failed samples and continue!\n" } else { - error("At least one input file for the following sample(s) was too small (<1KB):\n$samples\nEither remove those samples or ignore that samples using `--ignore_empty_input_files`.") + error("At least one input file for the following sample(s) had too less reads (<$params.min_read_counts):\n$samples\nEither remove those samples, adjust the threshold with `--min_read_counts`, or ignore that samples using `--ignore_empty_input_files`.") } } } From ecc79dd4442a681761306fe4524de59ad4ba4e6c Mon Sep 17 00:00:00 2001 From: daniel Date: Thu, 15 Jun 2023 15:20:06 +0200 Subject: [PATCH 125/158] fix typo --- nextflow_schema.json | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/nextflow_schema.json b/nextflow_schema.json index 24b4d942..d3b50e55 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -88,8 +88,8 @@ "min_read_counts": { "type": "integer", "default": 1, - "description": "Set read count threshold for failed input samples.", - "help_text": "Samples with less reads than this threshold at input or after trimming stops the pipeline. Using `--ignore_empty_input_files` or `--ignore_failed_trimming ignores` samples with read numbers below the threshold and lets the pipeline continue with less samples.", + "description": "Set read count threshold for failed samples.", + "help_text": "Samples with less reads than this threshold at input or after trimming stop the pipeline. Using `--ignore_empty_input_files` or `--ignore_failed_trimming ignores` samples with read numbers below the threshold and lets the pipeline continue with less samples.", "fa_icon": "fas fa-greater-than-equal" }, "ignore_empty_input_files": { From 988eeb27511249b30e91741d513165b6ea9b569d Mon Sep 17 00:00:00 2001 From: daniel Date: Thu, 15 Jun 2023 16:37:18 +0200 Subject: [PATCH 126/158] update workflow figure --- CHANGELOG.md | 3 +- docs/images/ampliseq_workflow.png | Bin 709452 -> 745412 bytes docs/images/ampliseq_workflow.svg | 401 ++++++++++++++++++++---------- 3 files changed, 266 insertions(+), 138 deletions(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index 48a95085..99856307 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -9,13 +9,14 @@ and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0 - [#591](https://github.com/nf-core/ampliseq/pull/591) - New version of the Unite taxonomy databases: 9.0 - [#564](https://github.com/nf-core/ampliseq/pull/564),[#567](https://github.com/nf-core/ampliseq/pull/567),[#582](https://github.com/nf-core/ampliseq/pull/582) - Added phylogenetic placement -- [#577](https://github.com/nf-core/ampliseq/pull/577),[#577](https://github.com/nf-core/ampliseq/pull/567) - Added SINTAX for taxonomic classification +- [#577](https://github.com/nf-core/ampliseq/pull/577) - Added SINTAX for taxonomic classification - [#575](https://github.com/nf-core/ampliseq/pull/575), [#586](https://github.com/nf-core/ampliseq/pull/586) - Added filtering step for stop codons for ASVs that are of coding regions. ### `Changed` - [#563](https://github.com/nf-core/ampliseq/pull/563) - Renamed DADA2 taxonomic classification files to include the chosen reference taxonomy abbreviation. - [#567](https://github.com/nf-core/ampliseq/pull/567) - Renamed `--dada_tax_agglom_min` and `--qiime_tax_agglom_min` to `--tax_agglom_min` and `--dada_tax_agglom_max` and `--qiime_tax_agglom_max` to `--tax_agglom_max` +- [#598](https://github.com/nf-core/ampliseq/pull/598) - Updated Workflow figure with SINTAX and phylogenetic placement ### `Fixed` diff --git a/docs/images/ampliseq_workflow.png b/docs/images/ampliseq_workflow.png index eca455acd75a1de761dc5ba0fb75dc4467c6dd0c..0dee77202c6b189ff4bf027aaa24158f483c4978 100644 GIT binary patch literal 745412 zcmZ^L2V9PA|Mz9zMs`UQveM8{qK&k*OOlF08%^!ZG-*lE(9mAAOT#Foq@5o{HHWyPu1(XFFUC{&UXqKXvChMg43s{DUe z;}s>((qa6u((JUP@;~^o`$zW?{=N35#04`7h1Hb&x9m~bIZM2_)BLQOxzdem=2n+Y zuTre6td8oN7@Fx^zIpZN4b$t-2Lm(+Wy-)9gzGYwkDd@tm{i{sUM4nCpp2Ibq)>=)BSHM zYC=z~2oJ}NS5{VzhzfB~=j7xR6JKWFS5P4GcBE*R>CKxQd}M++F^|39B&=5zAxW(s zj?W@*W=<`d8tvk_J^X!L!jB(6UK%&0tLRZl_Ev_AKd5g=H-0W4Ab?}O7v8g&pPgEl zF!JqsNDSUft*-O?R|&O4NT&yHtNgl@`(FNC+&Z< z_U@|n%&B4$5|6Hx2dcc}A!ic!>8R_5R+~Py@@JgyyJ_)~NWzv~yW-X`+!oozYZ5=~ z%#MRfbh-3@esEFGy!ok@##<-`USnhCAB&1q^{B+fi`R91 zDlbpC>3Qf}pz;alh;?00BHq1ww=U_{pW(}Mv!fN%>g~O?anxUK_146!b3CW0sL1?c 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a/docs/images/ampliseq_workflow.svg b/docs/images/ampliseq_workflow.svg index ae56ffbe..d360c4ca 100644 --- a/docs/images/ampliseq_workflow.svg +++ b/docs/images/ampliseq_workflow.svg @@ -148,7 +148,7 @@ id="linearGradient5670-9-95" xlink:href="#linearGradient5668" inkscape:collect="always" - gradientTransform="matrix(0.26458333,0,0,0.28227706,103.97164,88.655711)" /> + gradientTransform="matrix(0.26458333,0,0,0.2796764,103.97164,89.114258)" /> + gradientTransform="matrix(-0.26458333,0,0,-0.10679224,158.30358,174.09233)" /> + - Infer AmpliconSequence VariantsInfer sequences(ASVs) @@ -810,21 +805,21 @@ inkscape:export-xdpi="305.81" inkscape:connector-curvature="0" id="path5662-9-2" - d="m 123.78452,110.91469 7e-5,-28.328499 0.22614,1.865045 c 2.01924,16.652964 3.07454,20.735714 6.67589,25.827654 l 0.92328,1.30539 -0.88175,1.47985 c -1.65672,2.78045 -2.41296,4.35193 -3.26592,6.78668 -1.10577,3.15639 -2.04838,7.75931 -3.03195,14.80541 -0.21285,1.52479 -0.44524,3.18064 -0.51642,3.67967 -0.11127,0.78008 -0.1294,-3.06489 -0.12934,-27.4212 z" + d="m 123.78452,111.16816 7e-5,-28.067503 0.22614,1.847862 c 2.01924,16.499541 3.07454,20.544671 6.67589,25.589701 l 0.92328,1.29336 -0.88175,1.46622 c -1.65672,2.75483 -2.41296,4.31183 -3.26592,6.72415 -1.10577,3.12731 -2.04838,7.68782 -3.03195,14.66901 -0.21285,1.51074 -0.44524,3.15133 -0.51642,3.64577 -0.11127,0.77289 -0.1294,-3.03666 -0.12934,-27.16857 z" style="opacity:1;fill:url(#linearGradient5670-9-95);fill-opacity:1;stroke:#000000;stroke-width:0.05;stroke-linecap:square;stroke-linejoin:bevel;stroke-miterlimit:4;stroke-dasharray:none;stroke-dashoffset:0;stroke-opacity:1;paint-order:normal" inkscape:export-filename="C:\Users\fagernaes\Pictures\eager2\eager2_workflow_review.png" /> + rx="3.8618414" /> DADA2 @@ -890,21 +885,21 @@ inkscape:export-ydpi="305.81" inkscape:export-xdpi="305.81" id="text4732-0-6-7-2" - y="118.36993" + y="116.78242" x="96.411736" - style="font-style:normal;font-variant:normal;font-weight:bold;font-stretch:normal;font-size:3.52778px;line-height:1;font-family:'Maven Pro';-inkscape-font-specification:'Maven Pro Bold';text-align:center;letter-spacing:0px;word-spacing:0px;text-anchor:middle;fill:#000000;fill-opacity:1;stroke:none;stroke-width:0.264583" + style="font-style:normal;font-variant:normal;font-weight:bold;font-stretch:normal;font-size:3.52778006px;line-height:1;font-family:'Maven Pro';-inkscape-font-specification:'Maven Pro Bold';text-align:center;letter-spacing:0px;word-spacing:0px;text-anchor:middle;fill:#000000;fill-opacity:1;stroke:none;stroke-width:0.26458299" xml:space="preserve" inkscape:export-filename="C:\Users\fagernaes\Pictures\eager2\eager2_workflow_review.png">QIIME2 + transform="translate(-4.9145208,-3.1750001)"> assignTaxonomy + style="font-size:3.17499995px;stroke-width:0.26458299">assignTaxonomy + transform="translate(-4.9145208,-4.7625002)"> addSpecies + style="font-size:3.17499995px;stroke-width:0.26458299">addSpecies + transform="translate(0,-9.5250005)"> extract-reads + style="font-size:3.17499995px;stroke-width:0.26458299">extract-reads + transform="translate(0,-10.583334)"> classify-sklearn + style="font-size:3.17499995px;stroke-width:0.26458299">classify-sklearn + transform="translate(0,-10.054167)"> fit-classifier-naive-bayes + style="font-size:2.82222009px;stroke-width:0.26458299">fit-classifier-naive-bayes + rx="1.8965623" /> Reference taxonomy + transform="translate(55.409533,-6.0830591)"> + transform="translate(88.479685,-11.661582)"> + transform="translate(82.758013,-21.976031)"> + transform="translate(55.700763,-26.377237)"> + transform="translate(93.656771,-27.554416)"> + transform="translate(-34.925004,-24.08092)"> + transform="translate(-34.925004,-1.8596019)"> + transform="translate(-34.925004,-27.792687)"> removeBimeraDenovo + style="font-size:3.17499995px;stroke-width:0.26458299">removeBimeraDenovo + transform="translate(0,-12.700002)"> Barrnap + style="font-size:3.17499995px;stroke-width:0.26458299">Barrnap + transform="translate(0,-7.1437543)"> length filtering + style="font-size:3.17499995px;stroke-width:0.26458299">length filtering + transform="translate(-34.925004,-26.987511)"> mergeSequenceTables + style="font-size:3.17499995px;stroke-width:0.26458299">mergeSequenceTables + transform="translate(-34.925004,-20.578741)"> ITSx + style="font-size:3.17499995px;stroke-width:0.26458299">ITSx Extract ITS region @@ -2218,13 +2213,13 @@ inkscape:export-ydpi="305.81" inkscape:export-xdpi="305.81" id="text4732-3-3-2" - y="134.49713" + y="129.20554" x="41.395306" - style="font-style:normal;font-variant:normal;font-weight:bold;font-stretch:normal;font-size:3.52778px;line-height:0.25;font-family:'Maven Pro';-inkscape-font-specification:'Maven Pro Bold';letter-spacing:0px;word-spacing:0px;fill:#000000;fill-opacity:1;stroke:none;stroke-width:0.264583" + style="font-style:normal;font-variant:normal;font-weight:bold;font-stretch:normal;font-size:3.52778006px;line-height:0.25;font-family:'Maven Pro';-inkscape-font-specification:'Maven Pro Bold';letter-spacing:0px;word-spacing:0px;fill:#000000;fill-opacity:1;stroke:none;stroke-width:0.26458299" xml:space="preserve" inkscape:export-filename="C:\Users\fagernaes\Pictures\eager2\eager2_workflow_review.png">Filter ASVs @@ -2281,30 +2276,9 @@ style="font-style:normal;font-variant:normal;font-weight:normal;font-stretch:normal;font-size:3.175px;font-family:Arial;-inkscape-font-specification:Arial;stroke-width:0.264583px" x="40.422691" y="161.44469">mandatory - DADA2 only - - + transform="translate(66.443358,-27.554416)"> CO1 + + + coding region + + SINTAX + + + VSEARCH + + Phylogeneticplacement + + + HMMER / MAFFT + + + + Gappa + + + + EPA-NG + From dde0637f03f5d6d41d0dfb8cfbaa96738b3c5163 Mon Sep 17 00:00:00 2001 From: daniel Date: Mon, 19 Jun 2023 12:18:52 +0200 Subject: [PATCH 127/158] 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