Add filter for variants not supported by both strands

.github/workflows/juno_mapping_conda.yaml

@@ -25,6 +25,7 @@ jobs:
       - name: Install Conda environment with Micromamba
         uses: mamba-org/setup-micromamba@v1
         with:
+          generate-run-shell: false # see https://github.com/mamba-org/setup-micromamba/issues/130
           cache-downloads: true
           environment-file: envs/juno_mapping.yaml
       - name: Cache minikraken

.github/workflows/juno_mapping_singularity.yaml

@@ -25,6 +25,7 @@ jobs:
       - name: Install Conda environment with Micromamba
         uses: mamba-org/setup-micromamba@v1
         with:
+          generate-run-shell: false # see https://github.com/mamba-org/setup-micromamba/issues/130
           cache-downloads: true
           environment-file: envs/juno_mapping.yaml
       - name: Cache minikraken

envs/juno_mapping.yaml

@@ -8,9 +8,9 @@ dependencies:
   - git=2.40.*
   - mamba=1.3.*
   - pandas=1.5.*
-  - snakemake=7.18.*
+  - snakemake=7.32.0
   - pip=23.*
   - python=3.11.*
   - pyvcf
   - pip:
-    - "--editable=git+https://github.com/RIVM-bioinformatics/juno-library.git@v2.0.0#egg=juno_library"
+    - "--editable=git+https://github.com/RIVM-bioinformatics/juno-library.git@v2.1.2#egg=juno_library"

juno_mapping.py

@@ -7,15 +7,17 @@
 Date: 25-05-2023   
 """
 
-from pathlib import Path
-import pathlib
-import yaml
 import argparse
+import pathlib
 import sys
 from dataclasses import dataclass, field
+from pathlib import Path
+from typing import Any, Callable, Optional, Union
+
+import yaml
 from juno_library import Pipeline
-from typing import Optional, Callable, Union, Any
-from version import __package_name__, __version__, __description__
+
+from version import __description__, __package_name__, __version__
 
 
 def main() -> None:
@@ -150,6 +152,13 @@ def _add_args_to_parser(self) -> None:
             default=0.8,
             help="Minimum allele frequency to filter variants on.",
         )
+        self.add_argument(
+            "--min-reads-per-strand",
+            type=check_number_within_range(minimum=0, maximum=9999),
+            metavar="INT",
+            default=1,
+            help="Minimum number of reads per strand to call a variant.",
+        )
 
     def _parse_args(self) -> argparse.Namespace:
         args = super()._parse_args()
@@ -168,6 +177,7 @@ def _parse_args(self) -> argparse.Namespace:
         self.species: str = args.species
         self.minimum_depth_variant: int = args.minimum_depth_variant
         self.minimum_allele_frequency: float = args.minimum_allele_frequency
+        self.min_reads_per_strand: int = args.min_reads_per_strand
 
         return args
 
@@ -249,6 +259,7 @@ def setup(self) -> None:
             "species": str(self.species),
             "minimum_depth": int(self.minimum_depth_variant),
             "minimum_allele_frequency": float(self.minimum_allele_frequency),
+            "min_reads_per_strand": int(self.min_reads_per_strand),
         }
 
 

workflow/rules/variant_filtering.smk

@@ -1,59 +1,62 @@
-rule filter_af:
+rule FilterMutectCalls:
     input:
-        vcf=OUT + "/variants_raw/FMC/{sample}.vcf",
+        vcf=OUT + "/variants_raw/raw/{sample}.vcf",
         stats=OUT + "/variants_raw/raw/{sample}.vcf.stats",
         ref=OUT + "/reference/reference.fasta",
     output:
-        vcf=OUT + "/variants_raw/FMC_af/{sample}.vcf",
+        vcf=OUT + "/variants_raw/FMC/{sample}.vcf",
     message:
-        "Marking minority variants for {wildcards.sample}"
+        "Marking low confidence variants for {wildcards.sample}, based on FilterMutectCalls microbial mode"
     container:
-        "docker://staphb/bcftools:1.16"
+        "docker://broadinstitute/gatk:4.3.0.0"
     conda:
-        "../envs/bcftools.yaml"
+        "../envs/gatk_picard.yaml"
     params:
-        min_af=config["minimum_allele_frequency"],
+        min_reads_per_strand=config["min_reads_per_strand"],
     log:
-        OUT + "/log/filter_af/{sample}.log",
+        OUT + "/log/FilterMutectCalls/{sample}.log",
     threads: config["threads"]["filter_variants"]
     resources:
         mem_gb=config["mem_gb"]["filter_variants"],
     shell:
         """
-bcftools filter \
---exclude \"FORMAT/AF < {params.min_af}\" \
---soft-filter "min_af_{params.min_af}" \
-{input.vcf} \
-1>{output.vcf} \
-2>{log}
+gatk FilterMutectCalls -V {input.vcf} \
+-R {input.ref} \
+-O {output.vcf} \
+--stats {input.stats} \
+--min-reads-per-strand {params.min_reads_per_strand} \
+--microbial-mode 2>&1>{log}
         """
 
 
-rule FilterMutectCalls:
+rule filter_af:
     input:
-        vcf=OUT + "/variants_raw/raw/{sample}.vcf",
+        vcf=OUT + "/variants_raw/FMC/{sample}.vcf",
         stats=OUT + "/variants_raw/raw/{sample}.vcf.stats",
         ref=OUT + "/reference/reference.fasta",
     output:
-        vcf=OUT + "/variants_raw/FMC/{sample}.vcf",
+        vcf=OUT + "/variants_raw/FMC_af/{sample}.vcf",
     message:
-        "Marking low confidence variants for {wildcards.sample}, based on FilterMutectCalls microbial mode"
+        "Marking minority variants for {wildcards.sample}"
     container:
-        "docker://broadinstitute/gatk:4.3.0.0"
+        "docker://staphb/bcftools:1.16"
     conda:
-        "../envs/gatk_picard.yaml"
+        "../envs/bcftools.yaml"
+    params:
+        min_af=config["minimum_allele_frequency"],
     log:
-        OUT + "/log/FilterMutectCalls/{sample}.log",
+        OUT + "/log/filter_af/{sample}.log",
     threads: config["threads"]["filter_variants"]
     resources:
         mem_gb=config["mem_gb"]["filter_variants"],
     shell:
         """
-gatk FilterMutectCalls -V {input.vcf} \
--R {input.ref} \
--O {output.vcf} \
---stats {input.stats} \
---microbial-mode 2>&1>{log}
+bcftools filter \
+--exclude \"FORMAT/AF < {params.min_af}\" \
+--soft-filter "min_af_{params.min_af}" \
+{input.vcf} \
+1>{output.vcf} \
+2>{log}
         """