Merge pull request #41 from Pathogen-Genomics-Cymru/0.9.7-dev

0.9.7 dev

.github/workflows/build-push-quay.yml

@@ -2,8 +2,8 @@ name: build-push-quay
 on:
   push:
     branches:
-      - main
       - v0.9.6
+      - 0.9.7-dev
     paths:
       - '**/Dockerfile*'
 

README.md

@@ -67,8 +67,7 @@ Run [vcfmix](https://github.com/AlexOrlek/VCFMIX), yes or no. Set to no for synt
 Run [gnomonicus](https://github.com/oxfordmmm/gnomonicus), yes or no<br />
 * **amr_cat**<br />
 Path to AMR catalogue for gnomonicus<br />
-* **afanc_myco_db**<br />
-Path to the [afanc](https://github.com/ArthurVM/Afanc) database used for speciation. Obtain from  https://s3.climb.ac.uk/microbial-bioin-sp3/Mycobacteriaciae_DB_6.0.tar.gz
+* **afanc_myco_db**<br />Path to the [afanc](https://github.com/ArthurVM/Afanc) database used for speciation. Obtain from  https://s3.climb.ac.uk/microbial-bioin-sp3/Mycobacteriaciae_DB_7.0.tar.gz
 <br />
 
 For more information on the parameters run `nextflow run main.nf --help`

bin/reformat_afanc_json.py

@@ -17,15 +17,28 @@ def reformat_json(afanc_report):
         new_json[sample_id]["phylogenetics"]["species"] = {}
         species_dict = new_json[sample_id]["phylogenetics"]["species"]
 
-        for event in data["results"]["Detection_events"]:
-            if "closest_variant" in event:
-                event = event["closest_variant"]
+        cr = data["results"]["Detection_events"]["Clustering_results"]
+        vp = data["results"]["Detection_events"]["Variant_profile"]
+
+        for c_event in cr:
+            event = c_event
+            species = "_".join(c_event["name"].split(" ")[:2])
+            id  = species.replace(".", "")
+
+            ## capture closest variant info
+            if "closest_variant" in c_event:
+                event = c_event["closest_variant"]
+                id = event["name"].replace(" ", "_")
+
+            ## check to see if the top species hit exists within the variant profile
+            if species in vp and len(vp[species]) > 0:
+                ## if so, construct a new hit id
+                id  = species + "_" + sorted(list(vp[species].keys()))[-1]
 
-            id = event["name"].replace(" ", "_")
-            id = id.replace(".", "")
             species_dict[id] = { "percent_coverage" : event["reference_cov"]*100, "median_depth" : event["median_DOC"] }
 
     new_json["arguments"] = data["arguments"]
+    new_json["versions"] = data["versions"]
 
     with open(f"{sample_id}_afanc_report.json", "w") as fout:
         json.dump(new_json, fout, indent = 4)

docker/Dockerfile.preprocessing-0.9.6 → docker/Dockerfile.preprocessing-0.9.7

@@ -5,19 +5,21 @@ LABEL maintainer="[email protected]" \
 about.summary="container for the preprocessing workflow"
 
 ENV samtools_version=1.12 \
+bcftools_version=1.12 \
 htslib_version=1.12 \
 bedtools_version=2.29.2 \
 bowtie2_version=2.4.2 \
 fastp_version=0.20.1 \
 fastqc_version=0.11.9 \
 fqtools_version=2.3 \
 kraken2_version=2.1.1 \
-afanc_version=0.9.2 \
+afanc_version=0.10.2 \
 mykrobe_version=0.12.1 \
 bwa_version=0.7.17 \
-mash_version=2.3
+mash_version=2.3 \
+fastani_version=1.33
 
-ENV PACKAGES="procps curl git wget build-essential zlib1g-dev libncurses-dev libz-dev libbz2-dev liblzma-dev libcurl4-openssl-dev rsync unzip ncbi-blast+ pigz jq libtbb-dev openjdk-11-jre-headless autoconf r-base-core locales locales-all" \
+ENV PACKAGES="procps curl git wget build-essential zlib1g-dev libncurses-dev libz-dev libbz2-dev liblzma-dev libcurl4-openssl-dev libgsl-dev rsync unzip ncbi-blast+ pigz jq libtbb-dev openjdk-11-jre-headless autoconf r-base-core locales locales-all" \
 PYTHON="python3 python3-pip python3-dev" \
 PYTHON_PACKAGES="biopython"
 
@@ -36,7 +38,11 @@ RUN curl -fsSL https://github.com/samtools/samtools/archive/${samtools_version}.
 && curl -fsSL https://github.com/samtools/htslib/releases/download/${htslib_version}/htslib-${htslib_version}.tar.bz2 | tar -xj \
 && make -C samtools-${samtools_version} -j HTSDIR=../htslib-${htslib_version} \
 && make -C samtools-${samtools_version} -j HTSDIR=../htslib-${htslib_version} prefix=/usr/local install \
-&& rm -r samtools-${samtools_version}
+&& rm -r samtools-${samtools_version} \
+&& curl -fsSL https://github.com/samtools/bcftools/archive/refs/tags/${bcftools_version}.tar.gz | tar -xz \
+&& make -C bcftools-${bcftools_version} -j HTSDIR=../htslib-${htslib_version} \
+&& make -C bcftools-${bcftools_version} -j HTSDIR=../htslib-${htslib_version} prefix=/usr/local install \
+&& rm -r bcftools-${bcftools_version}
 
 RUN curl -fsSL https://github.com/alastair-droop/fqtools/archive/v${fqtools_version}.tar.gz | tar -xz \
 && mv htslib-${htslib_version} fqtools-${fqtools_version} \
@@ -90,7 +96,10 @@ RUN curl -fsSL https://github.com/ArthurVM/Afanc/archive/refs/tags/v${afanc_vers
 && cd .. \
 && curl -fsSL "https://github.com/marbl/Mash/releases/download/v${mash_version}/mash-Linux64-v${mash_version}.tar" | tar -x \
 && mv mash-Linux64-v${mash_version}/mash /usr/local/bin \
-&& rm -r mash-Linux*
+&& rm -r mash-Linux* \
+&& wget https://github.com/ParBLiSS/FastANI/releases/download/v${fastani_version}/fastANI-Linux64-v${fastani_version}.zip \
+&& unzip fastANI-Linux64-v${fastani_version}.zip \
+&& mv fastANI /usr/local/bin
 
 RUN sh -c "$(curl -fsSL ftp://ftp.ncbi.nlm.nih.gov/entrez/entrezdirect/install-edirect.sh)" \
 && mkdir -p /opt/edirect \

modules/preprocessingModules.nf

@@ -253,7 +253,7 @@ process fastQC {
 
 process kraken2 {
     /**
-    * @QCcheckpoint only pass to Afanc if Kraken's top family classification is Mycobacteriaceae
+    * @QCcheckpoint if Kraken's top family classification is NOT Mycobacteriaceae, sample will not proceed further than afanc
     */
 
     tag { sample_name }
@@ -301,7 +301,7 @@ process kraken2 {
 
     run_afanc=\$(jq '.afanc' ${kraken2_json})
 
-    if [ \$run_afanc == '\"true\"' ]; then printf "${sample_name}"; else echo '{"error":"Kraken's top family hit either wasn't Mycobacteriaceae, or there were < 100k Mycobacteriaceae reads"}' | jq '.' > ${error_log} && printf "no" && jq -s ".[0] * .[1]" ${software_json} ${error_log} > ${report_json}; fi
+    if [ \$run_afanc == '\"true\"' ]; then printf "${sample_name}"; else echo '{"error":"Kraken's top family hit either wasn't Mycobacteriaceae, or there were < 100k Mycobacteriaceae reads. Sample will not proceed further than afanc."}' | jq '.' > ${error_log} && printf "no" && jq -s ".[0] * .[1]" ${software_json} ${error_log} > ${report_json}; fi
     """
 
     stub:
@@ -334,26 +334,40 @@ process afanc {
     label 'medium_memory'
     label 'retryAfanc'
 
-    publishDir "${params.output_dir}/$sample_name/speciation_reports_for_reads_postFastP", mode: 'copy', pattern: '*.json'
+    publishDir "${params.output_dir}/$sample_name/speciation_reports_for_reads_postFastP", mode: 'copy', pattern: '*_afanc_report.json'
+    publishDir "${params.output_dir}/$sample_name", mode: 'copy', overwrite: 'true', pattern: '*{_err.json,_report.json}'
 
     input:
-    tuple val(sample_name), path(fq1), path(fq2), val(run_afanc), path(software_json)
+    tuple val(sample_name), path(fq1), path(fq2), val(run_afanc), path(software_json), path(kraken_report), path(kraken_json)
     path(afanc_myco_db)
 
-    when:
-    run_afanc =~ /${sample_name}/
-
     output:
-    tuple val(sample_name), path("${sample_name}_afanc_report.json"), stdout, emit: afanc_report
+    tuple val(sample_name), path("${sample_name}_afanc_report.json"), stdout, emit: afanc_json
+    path "${sample_name}_err.json", emit: afanc_log optional true
+    path "${sample_name}_report.json", emit: afanc_report optional true
     // tuple val(sample_name), path(fq1), path(fq2), stdout, emit: afanc_fqs
 
     script:
     afanc_report = "${sample_name}_afanc_report.json"
+    error_log = "${sample_name}_err.json"
+    report_json = "${sample_name}_report.json"
 
     """
-    afanc screen ${afanc_myco_db} ${fq1} ${fq2} -p 5.0 -n 1000 -o ${sample_name} -t ${task.cpus}
-    python3 ${baseDir}/bin/reformat_afanc_json.py ${sample_name}/${sample_name}.json
-    printf ${sample_name}
+    if [[ ${run_afanc} =~ /${sample_name}/ ]]
+    then
+	afanc screen ${afanc_myco_db} ${fq1} ${fq2} -p 5.0 -n 1000 -o ${sample_name} -t ${task.cpus} -v ${afanc_myco_db}/lineage_profiles/TB_variants.tsv
+	python3 ${baseDir}/bin/reformat_afanc_json.py ${sample_name}/${sample_name}.json
+	printf ${sample_name}
+    else
+	afanc screen ${afanc_myco_db} ${fq1} ${fq2} -p 2.0 -n 500 -o ${sample_name} -t ${task.cpus} -v ${afanc_myco_db}/lineage_profiles/TB_variants.tsv
+	python3 ${baseDir}/bin/reformat_afanc_json.py ${sample_name}/${sample_name}.json
+
+	python3 ${baseDir}/bin/identify_tophit_and_contaminants2.py ${afanc_report} ${kraken_json} ${baseDir}/resources/assembly_summary_refseq.txt ${params.species} ${params.unmix_myco} ${baseDir}/resources null
+
+	echo '{"error":"Kraken's top family hit either wasn't Mycobacteriaceae, or there were < 100k Mycobacteriaceae reads. Sample will not proceed further than afanc."}' | jq '.' > ${error_log} && printf "no" && jq -s ".[0] * .[1] * .[2]" ${software_json} ${error_log} ${sample_name}_species_in_sample.json > ${report_json}
+
+    fi
+
     """
 
     stub:
@@ -490,7 +504,7 @@ process identifyBacterialContaminants {
 
     if [ \$contam_to_remove == 'yes' ]; then cp ${sample_name}_species_in_sample.json ${sample_name}_species_in_sample_previous.json; fi
 
-    if [ \$contam_to_remove == 'yes' ]; then printf "NOW_DECONTAMINATE_${sample_name}"; elif [ \$contam_to_remove == 'no' ] && [ \$acceptable_species == 'yes' ]; then printf "NOW_ALIGN_TO_REF_${sample_name}" && mv $fq1 ${sample_name}_nocontam_1.fq.gz && mv $fq2 ${sample_name}_nocontam_2.fq.gz; elif [ \$contam_to_remove == 'no' ] && [ \$acceptable_species == 'no' ]; then jq -n --arg key "\$top_hit" '{"error": ("top hit " + \$key + " is not one of the 10 accepted mycobacteria")}' > ${error_log} && jq -s ".[0] * .[1] * .[2]" ${software_json} ${error_log} ${sample_name}_species_in_sample.json > ${report_json}; fi
+    if [ \$contam_to_remove == 'yes' ]; then printf "NOW_DECONTAMINATE_${sample_name}"; elif [ \$contam_to_remove == 'no' ] && [ \$acceptable_species == 'yes' ]; then printf "NOW_ALIGN_TO_REF_${sample_name}" && mv $fq1 ${sample_name}_nocontam_1.fq.gz && mv $fq2 ${sample_name}_nocontam_2.fq.gz; elif [ \$contam_to_remove == 'no' ] && [ \$acceptable_species == 'no' ]; then jq -n --arg key "\$top_hit" '{"error": ("top hit " + \$key + " does not have a reference genome. Sample will not proceed beyond preprocessing workflow.")}' > ${error_log} && jq -s ".[0] * .[1] * .[2]" ${software_json} ${error_log} ${sample_name}_species_in_sample.json > ${report_json}; fi
     """
 
     stub:
@@ -680,7 +694,7 @@ process reAfanc {
     afanc_report = "${sample_name}_afanc_report.json"
 
     """
-    afanc screen ${afanc_myco_db} ${fq1} ${fq2} -p 5.0 -n 1000 -o ${sample_name} -t ${task.cpus}
+    afanc screen ${afanc_myco_db} ${fq1} ${fq2} -p 5.0 -n 1000 -o ${sample_name} -t ${task.cpus} -v ${afanc_myco_db}/lineage_profiles/TB_variants.tsv
     python3 ${baseDir}/bin/reformat_afanc_json.py ${sample_name}/${sample_name}.json
     printf ${sample_name}
     """
@@ -759,7 +773,7 @@ process summarise {
 
     if [ \$contam_to_remove == 'yes' ]; then echo '{"error":"sample remains contaminated, even after attempting to resolve this"}' | jq '.' > ${error_log} && jq -s ".[0] * .[1] * .[2]" ${software_json} ${error_log} ${sample_name}_species_in_sample.json > ${report_json}; fi
 
-    if [ \$contam_to_remove == 'no' ] && [ \$acceptable_species == 'yes' ]; then printf "NOW_ALIGN_TO_REF_${sample_name}"; elif [ \$contam_to_remove == 'no' ] && [ \$acceptable_species == 'no' ]; then jq -n --arg key "\$top_hit" '{"error": ("top hit " + \$key + " is not one of the 10 accepted mycobacteria")}' > ${error_log} && jq -s ".[0] * .[1] * .[2]" ${software_json} ${error_log} ${sample_name}_species_in_sample.json > ${report_json}; fi
+    if [ \$contam_to_remove == 'no' ] && [ \$acceptable_species == 'yes' ]; then printf "NOW_ALIGN_TO_REF_${sample_name}"; elif [ \$contam_to_remove == 'no' ] && [ \$acceptable_species == 'no' ]; then jq -n --arg key "\$top_hit" '{"error": ("top hit " + \$key + " does not have a reference genome. Sample will not proceed beyond preprocessing workflow.")}' > ${error_log} && jq -s ".[0] * .[1] * .[2]" ${software_json} ${error_log} ${sample_name}_species_in_sample.json > ${report_json}; fi
     """
 
     stub:

nextflow.config

@@ -50,7 +50,7 @@ params {
   config_dir = "${baseDir}/nextflow.config"
 
   // path to the Afanc Myco database
-  afanc_myco_db = "${baseDir}/Mycobacteriaciae_DB_6.0/"
+  afanc_myco_db = "${baseDir}/Mycobacteriaciae_DB_7.0/"
 
 }
 
@@ -70,11 +70,11 @@ profiles {
       withLabel:high_memory { memory = '18GB' }
 
       withLabel:getversion {
-        container = "quay.io/pathogen-genomics-cymru/preprocessing:0.9.6"
+        container = "quay.io/pathogen-genomics-cymru/preprocessing:0.9.7"
       }
 
       withLabel:preprocessing {
-        container = "quay.io/pathogen-genomics-cymru/preprocessing:0.9.6"
+        container = "quay.io/pathogen-genomics-cymru/preprocessing:0.9.7"
       }
 
       withName:downloadContamGenomes {
@@ -84,17 +84,18 @@ profiles {
       }
 
       withLabel:retryAfanc {
+        shell = ['/bin/bash','-u']
         // Afanc sometimes fails curl in slurm, retry if so (error is masked as error status 1)
-        errorStrategy = {task.exitStatus == 1 ? 'retry' : 'terminate' }
+        errorStrategy = {task.exitStatus == 1 ? 'retry' : 'ignore' }
         maxRetries = 5
       }
 
       withLabel:clockwork {
-        container = "quay.io/pathogen-genomics-cymru/clockwork:0.9.6"
+        container = "quay.io/pathogen-genomics-cymru/clockwork:0.9.7"
       }
 
       withLabel:vcfpredict {
-        container = "quay.io/pathogen-genomics-cymru/vcfpredict:0.9.6"
+        container = "quay.io/pathogen-genomics-cymru/vcfpredict:0.9.7"
       }
 
     }
@@ -118,11 +119,11 @@ profiles {
       withLabel:high_memory { memory = '18GB' }
 
       withLabel:getversion {
-        container = "quay.io/pathogen-genomics-cymru/preprocessing:0.9.6"
+        container = "quay.io/pathogen-genomics-cymru/preprocessing:0.9.7"
       }
 
       withLabel:preprocessing {
-        container = "quay.io/pathogen-genomics-cymru/preprocessing:0.9.6"
+        container = "quay.io/pathogen-genomics-cymru/preprocessing:0.9.7"
       }
 
       withName:downloadContamGenomes {
@@ -132,16 +133,17 @@ profiles {
       }
 
       withLabel:retryAfanc {
-        errorStrategy = {task.exitStatus == 1 ? 'retry' : 'terminate' }
+	shell = ['/bin/bash','-u']
+        errorStrategy = {task.exitStatus == 1 ? 'retry' : 'ignore' }
         maxRetries = 5
       }
 
       withLabel:clockwork {
-        container = "quay.io/pathogen-genomics-cymru/clockwork:0.9.6"
+        container = "quay.io/pathogen-genomics-cymru/clockwork:0.9.7"
       }
 
       withLabel:vcfpredict {
-        container = "quay.io/pathogen-genomics-cymru/vcfpredict:0.9.6"
+        container = "quay.io/pathogen-genomics-cymru/vcfpredict:0.9.7"
       }
     }
   }
@@ -161,11 +163,11 @@ profiles {
       withLabel:high_memory { memory = '18GB' }
 
       withLabel:getversion {
-        container = "quay.io/pathogen-genomics-cymru/preprocessing:0.9.6"
+        container = "quay.io/pathogen-genomics-cymru/preprocessing:0.9.7"
       }
 
       withLabel:preprocessing {
-        container = "quay.io/pathogen-genomics-cymru/preprocessing:0.9.6"
+        container = "quay.io/pathogen-genomics-cymru/preprocessing:0.9.7"
       }
 
       withName:downloadContamGenomes {
@@ -175,16 +177,17 @@ profiles {
       }
 
       withLabel:retryAfanc {
-        errorStrategy = {task.exitStatus == 1 ? 'retry' : 'terminate' }
+	shell = ['/bin/bash','-u']
+        errorStrategy = {task.exitStatus == 1 ? 'retry' : 'ignore' }
         maxRetries = 5
       }
 
        withLabel:clockwork {
-        container = "quay.io/pathogen-genomics-cymru/clockwork:0.9.6"
+        container = "quay.io/pathogen-genomics-cymru/clockwork:0.9.7"
       }
 
       withLabel:vcfpredict {
-        container = "quay.io/pathogen-genomics-cymru/vcfpredict:0.9.6"
+        container = "quay.io/pathogen-genomics-cymru/vcfpredict:0.9.7"
       }
     }
   }

singularity/Singularity.preprocessing-0.9.6 → singularity/Singularity.preprocessing-0.9.7

@@ -9,19 +9,21 @@ about.summary="container for the preprocessing workflow"
 
 
 samtools_version=1.12
+bcftools_version=1.12
 htslib_version=1.12
 bedtools_version=2.29.2
 bowtie2_version=2.4.2
 fastp_version=0.20.1
 fastqc_version=0.11.9
 fqtools_version=2.3
 kraken2_version=2.1.1
-afanc_version=0.9.2
+afanc_version=0.10.2
 mykrobe_version=0.12.1
 bwa_version=0.7.17
 mash_version=2.3
+fastani_version=1.33
 
-PACKAGES="procps curl git wget build-essential zlib1g-dev libncurses-dev libz-dev libbz2-dev liblzma-dev libcurl4-openssl-dev rsync unzip ncbi-blast+ pigz jq libtbb-dev openjdk-11-jre-headless autoconf r-base-core locales locales-all"
+PACKAGES="procps curl git wget build-essential zlib1g-dev libncurses-dev libz-dev libbz2-dev liblzma-dev libcurl4-openssl-dev libgsl-dev rsync unzip ncbi-blast+ pigz jq libtbb-dev openjdk-11-jre-headless autoconf r-base-core locales locales-all"
 PYTHON="python3 python3-pip python3-dev"
 PYTHON_PACKAGES="biopython"
 
@@ -40,7 +42,11 @@ curl -fsSL https://github.com/samtools/samtools/archive/${samtools_version}.tar.
 && curl -fsSL https://github.com/samtools/htslib/releases/download/${htslib_version}/htslib-${htslib_version}.tar.bz2 | tar -xj \
 && make -C samtools-${samtools_version} -j HTSDIR=../htslib-${htslib_version} \
 && make -C samtools-${samtools_version} -j HTSDIR=../htslib-${htslib_version} prefix=/usr/local install \
-&& rm -r samtools-${samtools_version}
+&& rm -r samtools-${samtools_version} \
+&& curl -fsSL https://github.com/samtools/bcftools/archive/refs/tags/${bcftools_version}.tar.gz | tar -xz \
+&& make -C bcftools-${bcftools_version} -j HTSDIR=../htslib-${htslib_version} \
+&& make -C bcftools-${bcftools_version} -j HTSDIR=../htslib-${htslib_version} prefix=/usr/local install \
+&& rm -r bcftools-${bcftools_version}
 
 curl -fsSL https://github.com/alastair-droop/fqtools/archive/v${fqtools_version}.tar.gz | tar -xz \
 && mv htslib-${htslib_version} fqtools-${fqtools_version} \
@@ -94,7 +100,11 @@ curl -fsSL https://github.com/ArthurVM/Afanc/archive/refs/tags/v${afanc_version}
 && cd .. \
 && curl -fsSL "https://github.com/marbl/Mash/releases/download/v${mash_version}/mash-Linux64-v${mash_version}.tar" | tar -x \
 && mv mash-Linux64-v${mash_version}/mash /usr/local/bin \
-&& rm -r mash-Linux*
+&& rm -r mash-Linux* \
+&& wget https://github.com/ParBLiSS/FastANI/releases/download/v${fastani_version}/fastANI-Linux64-v${fastani_version}.zip \
+&& unzip fastANI-Linux64-v${fastani_version}.zip  \
+&& mv fastANI /usr/local/bin
+
 
 sh -c "$(curl -fsSL ftp://ftp.ncbi.nlm.nih.gov/entrez/entrezdirect/install-edirect.sh)" \
 && mkdir -p /opt/edirect \
@@ -124,18 +134,20 @@ LANGUAGE=en_US.UTF-8
 
 %environment
 export samtools_version=1.12
+export bcftools_version=1.12
 export htslib_version=1.12
 export bedtools_version=2.29.2
 export bowtie2_version=2.4.2
 export fastp_version=0.20.1
 export fastqc_version=0.11.9
 export fqtools_version=2.3
 export kraken2_version=2.1.1
-export afanc_version=0.9.2
+export afanc_version=0.10.2
 export mykrobe_version=0.12.1
 export bwa_version=0.7.17
 export mash_version=2.3
-export PACKAGES="procps curl git wget build-essential zlib1g-dev libncurses-dev libz-dev libbz2-dev liblzma-dev libcurl4-openssl-dev rsync unzip ncbi-blast+ pigz jq libtbb-dev openjdk-11-jre-headless autoconf r-base-core locales locales-all"
+export fastani_version=1.33
+export PACKAGES="procps curl git wget build-essential zlib1g-dev libncurses-dev libz-dev libbz2-dev liblzma-dev libcurl4-openssl-dev libgsl-dev rsync unzip ncbi-blast+ pigz jq libtbb-dev openjdk-11-jre-headless autoconf r-base-core locales locales-all"
 export PYTHON="python3 python3-pip python3-dev"
 export PYTHON_PACKAGES="biopython"
 export PATH=${PATH}:/usr/local/bin/mccortex/bin:/usr/local/bin/bwa-${bwa_version}:/opt/edirect