Add mutect2 force calling using '--alleles' flag

[jchchiu]

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Regarding issue #1883 I have:

• Added two params (mutect2_force_call, mutect2_force_call_tbi) to supply a path to a vcf.gz file and associated index file for force calling.
• Added Channel to collect .vcf.gz; added Channel to collect .vcf.gz tbi (if not found will create a tbi file); added params to main workflow pipeline.
• Added params to pipeline initialisation (raise an error if mutect2_force_call file format not in .vcf.gz format.
• Added TABIX creation for mutect2_force_call if user does not supply it.
• Added params to the [somatic/tumor_only]_[ALL/mutect2] subworkflows.
• Added params to GATK4 mutect2 module; created new command in script for force calling (mfc_command) and added to Mutect2 execution.
• Temporarily edited some tests configs in tools to test for the force call parameter (also tests if TABIX creation works if associated index file not supplied).

Some questions:

• Regarding tests:
  • For the test data I'm just thinking of creating a VCF with a few random alleles to be used to force call. Is there any other data you prefer?
  • I created test_force_call.vcf.gz which works for both tumor only and somatic mode:

##fileformat=VCFv4.2
##contig=<ID=chr21,length=46709983>
#CHROM	POS	ID	REF	ALT	QUAL	FILTER	INFO
chr21	46664275	.	A	C	.	.	.
chr21	46664280	.	A	C	.	.	.
chr21	46664285	.	A	G	.	.	.
chr21	46664290	.	T	A	.	.	.

• In the Mutect2 docs there is another boolean --genotype-filtered-alleles (or maybe it's now --force-call-filtered-alleles) flag that can be used in conjunction with --alleles. Is it worth it to also implement this?

[jchchiu]

Also, while I was editing ./subworkflows/local/samplesheet_to_channel/main.nf, I had a small question about bcftools annotate here.

// Fails when bcftools annotate is used but no files are supplied
if (tools && tools.split(',').contains('bcfann') && !(bcftools_annotations && bcftools_annotations_tbi && bcftools_header_lines)) {
    error("Please specify --bcftools_annotations, --bcftools_annotations_tbi, and --bcftools_header_lines, when using BCFTools annotations")
}

• In the ./main.nf NFCORE_SAREK workflow here, the PREPARE_GENOME workflow is called for bcftools_annotation to create a bcftools_annotation_tbi file if it doesn’t exist.
• However, in the main workflow here the PIPELINE_INITIALISATION workflow (and thus, ./subworkflows/local/samplesheet_to_channel/main.nf) is called before this to create a samplesheet to be passed into the NFCORE_SAREK workflow.
• Thus, if a user has to has to specify a bcftools_annotations_tbi file in the first place anyway for the pipeline to run, this makes the TABIX call for bcftools_annotation redundant.

Just a nitpick, but should either the !bcftools_annotations_tbi requirement, or the code for the TABIX creation of bcftools_annotation be removed?

If this is of interest and doesn't disrupt too much, I can open another PR with one of these updates.

[FriederikeHanssen]

This looks good already. I added some comments on where changes should be handeled instead.

What we are missing are updates to the output documentation of Mutect2.

[FriederikeHanssen]

we should proabbly have one test that checks things work with a supplied tbi file

[jchchiu]

Following remove validation error; handled by schema, I have:

• Removed validation error for samplesheets (handles in schema) and removed temporary tests
• Updated the gatk4/mutect module using nf-core and any associated subworkflows
• Updated output documentation and README
• Added a new test config specific to mutect2 to test the parameter, but unsure if this is the best way to add a test for it (maybe change the test to tumoronly?). As of Add config for readgroups to fq2bam #1939, I've been running with nextflow run . -profile test,tools_somatic_mutect2,mutect,docker --tools mutect2 --outdir results -step variant_calling

Sorry I also messed up the commit history, should have merged instead of rebased. The previous comments are up to:
temp: added param to config tests; testing TABIX creation

[maxulysse]

Can't we add the params we need to the tests?

[jchchiu]

Hey, I've tried two ways to do this, could you have a look at which one you prefer?

jchchiu#2: Is what makes most sense to me by adding test scenarios and updating snap; is this what you had in mind?

jchchiu#3: Just updates the somatic and tumor_only configs to include the parameter, but I believe this changes all mutect2 tests to always test with the force_call parameter which is not very good.

[maxulysse]

2nd is definitvely best for me

[nf-core-bot]

Warning

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Please update your pipeline to the latest version.

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[jchchiu]

@nf-core-bot fix linting

[jchchiu]

Heya, I've seen that 'Prepare genome refactor #1979' has been merged so I've updated my PR.

• The linting error is from the updated mutect2 module; haven't updated the module because of the extra ', path(gzi)' in the updated module that I'm guessing should be updated in a different PR

• I think everything should be good, just wondering whether there are any issues with the new snapshots for mutect2 using newer nf-test and nextflow versions? ( "nf-test": "0.9.3", "nextflow": "25.10.0" vs "nf-test": "0.9.2", "nextflow": "25.04.6")

• Also want to confirm that there is no need for a mutect2_force_call input in the SAMPLESHEET_TO_CHANNEL script; if so I'll also do another quick PR after to update Add mutect2 force calling using '--alleles' flag #1952 (comment) (also removing the bcftools_annotations_tbi // Path: bcftools annotations tbi input since it'll no longer be used in the script)