CHANGELOG.md

Changelog

All notable changes to this project will be documented in this file.

The format is based on Keep a Changelog, and this project adheres to Semantic Versioning.

[v2.2.0]

Added

• Output {{sample}}.stats.json file describing some key metrics for the analysis.
• Summary of gene coverage if a 4-column BED is provided.
• Automated sex determination using relative coverage of chrX and chrY.
• Retry strategy added to snp:aggregate_pileup_variants to prevent out of memory error.

Changed

• --GVCF --phased will produce a phased GVCF.
• Changed default phasing algorithm to whatshap, with the possibility to change the phasing to longphase with --use_longphase true.
  • The intermediate phasing is still performed using longphase.
• Setting --snp --sv --phased will emit individually phased SNPs and SVs.
• Phased bedMethyl files now follow the pattern {{ alias }}.wf_mods.{{ haplotype }}.bedmethyl.gz.
• --sex parameter uses XX and XY rather than "female" and "male".
• Update modkit to v0.2.6.
• Improved modkit runtime by increasing default threads and increasing the default interval size.
• Improved modkit runtime by increasing the default interval size and running modkit on individual contigs.
• modkit is now run only on chromosomes 1-22, X, Y and MT, unless --include_all_ctgs is provided.
• Increased minimum CPU requirement for the workflow to 16.
• Filtering of SVs using a BED file now includes sites only partially overlapping the specified regions.
• basecaller_cfg will be inferred from the basecall_model DS key of input read groups, if available
  • Providing --basecaller_cfg will not be required if basecall_model is present in the DS tag of the read groups of the input BAM
  • --basecaller_cfg will be ignored if a basecall_model is found in the input BAM
• Reconciled workflow with wf-template v5.1.2
• Update to Clair3 v1.0.8.
• Update to longphase v1.7.1.

Fixed

• Update schema to allow selection of multiple BAM files per sample in EPI2ME.
• Spectre CNV report not handling cases when no CNVs detected.
• Lines denoting normal maximum and pathogenic minimum thresholds now correctly displayed on STR repeat content plots.
• Workflow will not emit sample.cram if sample.haplotagged.cram has been created by the workflow to save storage.
• Emitting nonsense input.1 file

Removed

• Single-step joint phasing of SV and SNP.
• --output_separate_phased as the workflow emits only individually phased VCFs.
• A copy of the reference and the generated reference cache is no longer output by the workflow.
  • The workflow encourages use of readily available standard reference sequences, so re-emitting the input reference as a workflow output is unnecessarily consuming disk space.

[v2.1.0]

Changed

• ClinVar version in SnpEff container updated to version 20240307.
• Convert to BAM only when --cnv --use_qdnaseq is selected.
• Update to Clair3 v1.0.6.
• Update Spectre to fix an error when parsing Clair3 VCFs with multiple AFs.
• Support for an input folder of multiple BAM files per sample with --bam (instead of only allowing a single BAM per sample).
• refine_with_sv to be run by chromosome in order to reduce memory footprint.

Fixed

• Force minimap2 to clean up memory more aggressively. Empirically this reduces peak-memory use over the course of execution.
• Handling of input VCF files with --vcf_fn.
• --phased --sv --snp generates a truncated VCF file when # appears in the VCF INFO field
• Some reporting scripts using too much memory.

Removed

• CRAM as supported input format.
• old_ref parameter as providing the reference of an existing CRAM is no longer needed.

[v2.0.0]

Changed

• CNV calling with --cnv is now performed using Spectre, which is optimised for long reads.
  • Legacy CNV calling using QDNAseq may still be carried out with --cnv --use_qdnaseq.
  • The bin size parameter has been renamed from --bin_size to --qdnaseq_bin_size.
• Skip CNV CRAM to BAM conversion if downsampling is required, to avoid creating an unnecessary intermediate file.
• The output of --depth_intervals now has .bedgraph.gz extension.
• SV workflow outputs SVs in the autosomes, sex chromosomes and MT; use --include_all_ctgs to output calls on all the sequences.

Added

• Output definitions for coverage files.
• N50 and mean coverage added to alignment report.

Fixed

• EPI2ME Desktop incorrectly allowed selection of directory for tr_bed.
• failedQCReport failing to generate a report.

[v1.11.0]

Changed

• Add an additional whatshap haplotag process after the final VCF phasing.
• Updates to the phasing subworkflow significantly impact the runtime and storage requirement for the workflow, as detailed here.
• Several performance improvements which should noticeably reduce the running time of the workflow

Fixed

• Updated the version of Straglr, which addresses the following:
  • Repeats can now be called in RFC1
  • Start position of called STRs is 1-based rather than 0-based
  • VCF headers now match those in the FORMAT field
• Generate allChromosomes.bed using samtools faidx index instead of pyfaidx, to avoid a KeyError
• Inconsistent file ownership of bundled Clair3 model files which could lead to subuid errors in some environments

[v1.10.1]

Fixed

• Bug report form.

[v1.10.0]

Added

• Clair3 4.3.0 models.

Changed

• --phase_vcf, --joint_phasing and --phase_mod are now deprecated for --phased; see the README for more details.
• --use_longphase_intermediate is now deprecated, and --use_longphase false will use whatshap throughout consistently
• Running --phase --snp --use_longphase false will now phase indels too
• --basecalling_cfg currently provides the configuration to Clair3.
• The clair3: prefix to Clair3 specific models is no longer required.

Fixed

• SNP workflow ignoring the mitochondrial genome.
• CNV report generation fails if there is no consensus on the copy number of a chromosome
  • Undetermined category has been added to the Chromosome Copy Summary to account for these cases
• readStats reports metrics on the downsampled BAM when --downsample_coverage is requested.
• Spurious warning for missing MM/ML tags when a BAM fails the coverage threshold

Removed

• wf-basecalling subworkflow
  • fast5_dir input and other basecalling related options have been removed from the workflow parameters
  • Users should run the standalone wf-basecalling workflow and provide the output to wf-human-variation
• Mapula statistics with --mapula

[v1.9.2]

Fixed

• --joint_phasing generating single-chromosome VCF files.

[v1.9.1]

Changed

• ClinVar annotation of SVs has been temporarily removed due to not being correctly incorporated. SnpEff annotations are still produced as part of the final SV VCF.
• New documentation

Removed

• --annotation_threads parameter, as the SnpEff process does not support multithreading.

Fixed

• Truncated SV VCF header generated from vcfsort.
• sed crashing with I/O error in some instances.
• Missing flagstats file in output directory.

[v1.9.0]

Added

• STR workflow report now includes additional plots which display repeat units and interruptions in each supporting read
• CNV workflow now outputs an indexed VCF file to the output directory

Changed

• Legend symbols in STR genotpying plot
• Unambiguous naming of bedMethyl files generated with --mod
  • Unphased outputs will have the pattern [sample_name].wf_mods.bedmethyl.gz
  • Phased outputs will have the pattern [sample_name]_[1|2|ungrouped].wf_mods.bedmethyl.gz

Fixed

• Report step failing if bcftools stats file has only some sub-sections
• Clair3 ignoring the bed file
• merge_haplotagged_contigs incorrectly generating intermediate CRAM when input is BAM
• STR content generation failing due to forward slash in disease name in variant_catalog_hg38.json
• Report name for the read alignment statistics now follows the pattern [sample_name].wf-human-alignment-report.html

[v1.8.3]

Fixed

• configure-jbrowse breaking on unescaped spaces

[v1.8.2]

Added

• The SNP workflow will filter the final VCF to only return calls in the regions specified with --bed
  • Avoid clair3 calling variants in regions that flank those specified in the BED
• Add a process to sanitise BED files

Fixed

• SNP subworkflow was ignoring BED file and analysing all regions
• Report SV crashing when generating the size plots with only large indels
• Downsampling not working when targeting regions with --bed
• --phase_mod not emitting the haplotagged bam files
• Report crashing when loading a clinvar-annotated VCF file with multiple GENEINFO/CLNVC entries

Removed

• Default local executor CPU and RAM limits

[v1.8.1]

Fixed

• SV size barchart causing extremely large reports.
  • The plot has been replaced with a distribution plot fixed to +/- 5KBp. The summary table reports the INS and DEL min/max values.
• Workflow could not be launched from EPI2ME desktop app due to incorrectly quoted fields in schema

[v1.8.0]

Changed

• replaced --methyl with --mod, and --phase_methyl with --phase_mod
• replaced modbam2bed with modkit (v0.1.12)
• --phase_mod will generate three bed files, one for each haplotype and one for the reads that are not tagged.

Added

• Add locus for LRP12 to BED file of STR repeats (GRCh38)

[v1.7.2]

Changed

• When --bed and --bam_min_coverage are specified, the workflow will process the regions passing the coverage filters
• whatshap v2.0 in base workflow

Fixed

• Patch whatshap stats crashing when no heterozygote sites are found in a contig
• Patch makeReport crashing when loading empty ClinVar VCFs

[v1.7.1]

Changed

• Increased minimum memory required for the workflow to 16 GB of RAM to reduce alignment failures

Fixed

• sv.filterBam missing output file when creating BAM CSI

[v1.7.0]

Changed

• VCFs generated by the --sv option are now automatically annotated with SnpEff, incorporating ClinVar annotations
  • This can be switched off with --skip_annotation
• —-skip_annotation disables attempt to determine human genome version, enabling --snp, --sv and --phase_methyl to be called on genomes which aren’t hg19 or hg38
• Updated example command displayed when running --help
• The ClinVar table in --snp and --sv reports is now sorted according to clinical significance, and includes HGVS cDNA and protein descriptions
• Workflow options for disabling steps have been updated for consistency:
  • --skip_annotation is now --annotation false
  • --skip_refine_snp_with_sv is now --refine_snp_with_sv false
• --phase_methyl also calls modifications using all reads to account for unphased regions
• Input options and Output options have been combined in the Main options category
• Updated Clair3 to v1.0.4
• SV workflow does not filter the SV calls by size and type
• Report for bam files failing the depth threshold is now consistent with the report of bam passing the hard threshold
• Perform Sniffles SV phasing with --phase_sv
• --phase_vcf now run Sniffles SV in phasing mode

Fixed

• 'mosdepth_downsampled' is defined more than once warning
• Workflow crashing when providing a reference with spaces/brackets in file name
• Workflow not emitting GVCF even when requested
• GVCF sample name not matching sample_name
• --cnv subworkflow sometimes reporting incorrect genetic sex, due to the way segment copy numbers were aggregated across chromosomes

Added

• Add downsampling of large bam files
• --joint_phasing allows an additional joint SNP and SV phasing
• --joint_phasing and --phase_vcf now emit a phased block GTF file to facilitate visualization

Removed

• --sv_types, all SV types are returned without filtering
• --max_sv_length, all SVs are returned regardless of maximum size

[v1.6.1]

Changed

• GPU tasks are limited to run in serial by default to avoid memory errors
  • Users in cluster and cloud environments where GPU devices are scheduled must use -profile discrete_gpus to parallelise GPU work
  • A warning will be printed if the workflow detects it is running non-local execution but the discrete_gpus profile is not enabled
  • Additional guidance on GPU support is provided in our Quickstart

Fixed

• ModuleNotFoundError on callCNV step when transforming some VCFs
• Malformed VCF created by annotation step if Java emits a warning

[v1.6.0]

Changed

• VCFs generated by the --snp option are now automatically annotated with SnpEff, incorporating ClinVar annotations
  • This can be switched off with --skip_annotation
• Bumped minimum required Nextflow version to 22.10.8
• Updated wf-basecalling subworkflow to 0.7.1 (Dorado 0.3.0)
• Enum choices are enumerated in the --help output
• Enum choices are enumerated as part of the error message when a user has selected an invalid choice
• Made it easier to see which basecaller configurations can be used for basecalling and which configurations are presented to allow small variant calling of existing datasets
  • Basecaller configurations prefixed with clair3: cannot be used for basecalling

Added

• v4.2.0 basecalling models, which must be used for sequencing runs performed at new 5 kHz sampling rate
• v4.1.0 basecalling models replace v4.0.0 models and must be used for sequencing runs performed at 4 kHz sampling rate
• Clair3 260bps models

Fixed

• Workflow get_filter_calls_command crashes with stranded interval BED
• CRAM inputs are now converted internally to BAM to avoid CNV subworkflow crash
• Basecalls and alignments are emitted in BAM to support CNV subworkflow if selected
• Workflow incorrectly prompting for an --old_ref when providing unaligned CRAM

[v1.5.2]

Added

• Configuration for running demo data in AWS

Changed

• Reports for wf-human-sv and wf-human-snp

Fixed

• Coverage plot not working when a bed region file is provided
• Workflow crashing with --bam_min_coverage 0
• Subworkflows that require hg19/GRCh37 now correctly accept references where chromosome sequence names do not have the 'chr' prefix

[v1.5.1]

Changed

• Depth of sequencing plots moved to alignment report

Fixed

• Corrected bin size unit displayed on CNV report

[v1.5.0]

Added

• Workflow outputs alignment statistics report when alignment has been performed

Changed

• Updated Clair3 to v1.0.1 to bump WhatsHap dependency to v1.7
• Bumped base container to use samtools 1.17 to prevent user reported segfault during minimap2_ubam process

[v1.4.0]

Added

• --depth_intervals will output a bedGraph file with entries for each genomic interval featuring homogeneous depth
• --phase_methyl will output haplotype-level methylation calls, using the HP tags added by whatshap in the wf-human-snp workflow
• --sv_benchmark will benchmark SV calls with Truvari
  • The workflow uses the 'NIST_SVs_Integration_v0.6' truth set and benchmarking should be carried out on HG002 data.

Changed

• Added coverage barrier to wf-human-variation
• When the SNP and SV subworkflows are both selected, the workflow will use the results of the SV subworkflow to refine the SNP calls
  • This new behaviour can be disabled with --skip_refine_snp_with_sv
• Updated to Oxford Nanopore Technologies PLC. Public License
• Workflow requires target regions to have 20x mean coverage to permit analysis

Fixed

• report_sv.py now handles empty VCF files
• report_str.py now handles mono-allelic STR
• WhatsHap processes do not block for long time with CRAM inputs due to missing REF_PATH
• filterCalls failure when input BED has more than three columns
• get_genome assumed STR subworkflow was always enabled, preventing CNV analysis with hg19
• "genome build (...) is not compatible with this workflow" was incorrectly thrown when the workflow stopped before calling get_genome

[v1.3.0]

Added

• --str enables STR genotyping using Straglr (compatible with genome build 38 only)

Changed

• Minor performance improvement when checking for empty VCF in aggregate pileup steps

[v1.2.0]

Added

• --cnv enables CNV calling using QDNAseq

[v1.1.0]

Changed

• Updated Dorado container image to use Dorado v0.1.1
  • Latest models are now v4.0.0
  • Workflow prints a more helpful error when Dorado fails due to unknown model name
• Updated wf-human-snp container image to load new Clair3 models for v4 basecalling
• Default basecaller_cfg set to dna_r10.4.1_e8.2_400bps_sup@v4.0.0

Added

• --basecaller_args may be used to provide custom arguments to the basecalling process

[v1.0.1]

Changed

• Default basecaller_cfg set to dna_r10.4.1_e8.2_400bps_sup@v3.5.2
• Updated description in manifest

[v1.0.0]

Added

• nextflow run epi2me-labs/wf-human-variation --version will now print the workflow version number and exit

Changed

• --modbam2bed_args can be used to further configure the wf-methylation modbam2bed process
• modbam2bed outputs are now prefixed with <sample_name>.methyl
• --basecall_cfg is now required by the SNP calling subworkflow to automatically pick a suitable Clair3 model
  • Users no longer have to provide --model for the SNP calling subworkflow
• Tidied workflow parameter schema
  • Some advanced options that are primarily used for benchmarking are now hidden but can be listed with --help --show_hidden_params
• wf-basecalling subworkflow now separates reads into pass and fail CRAMs based on mean qscore
  • The workflow will index and align both the pass and fail reads and provide a CRAM for each in the output directory
  • Only pass reads are used for downstream variant calling
• Updated wf-human-variation-snp container to use Sniffles v2.0.7

Removed

• -profile conda is no longer supported, users should use -profile standard (Docker) or -profile singularity instead
• --report_name is no longer required and reports will be prefixed with --sample_name instead

Fixed

• Workflow will exit with "No files match pattern" if no suitable files are found to basecall
  • Ensure to set --dorado_ext to fast5 or pod5 as appropriate

[v0.4.1]

Fixed

• JBrowse2 configuration failed to load AlignmentTrack for CRAM output

[v0.4.0]

Added

• Workflow will now output a JBrowse2 jbrowse.json configuration
• Workflow reports will be visible in the Nextflow Tower reports tab

Changed

• wf-basecalling 0.0.1 has been integrated to wf-human-variation
  • Basecalling is now conducted with Dorado
  • Basecalling options have changed, users are advised to check the basecalling options in --help for guidance
• GPU accelerated processes can now have their process directives generically modified in downstream user profiles using withLabel:gpu
• mapula will no longer run by default due to performance issues on large data sets and must be enabled with --mapula
  • This step will be deprecated and replaced with an equivalent in a future release

Fixed

• uBAM input no longer requires an index
• CRAM is written to the output directory in all cases where alignment was completed

[v0.3.1]

Fixed

• check_for_alignment did not support uBAM
• Set PYTHONNOUSERSITE to reduce risk of environment bleed

[v0.3.0]

Added

• Experimental guppy subworkflow
  • We do not provide a Docker container for Guppy at this time, users will need to override process.withLabel:wf_guppy.container

Changed

• --ubam option removed, users can now pass unaligned or aligned BAM (or CRAM) to --bam
  • If the input BAM is aligned and the provided --ref does not match the SQ lines (exact match to name and size) the file will be realigned to --ref
  • If the input CRAM is aligned and the provided --ref does not match the SQ lines (exact match to name and size) the file will be realigned to --ref, but will also require the old reference to be provided with --old_ref to avoid reference lookups
  • If the input is not aligned, the file will be aligned to --ref

Fixed

• Chunks without variants will no longer terminate the workflow at the create_candidates step

[v0.2.3]

Added

• mapula used to generate basic alignment QC statistics in both CSV and JSON

Changed

• Updated Clair3 to 0.1.12 (build 6) which bundles Longphase 1.3 to enable CRAM support and small improvement to accuracy
• mosdepth artifacts are now written to the output directory
  • Additionally outputs read counts at 1,10,20,30X coverage thresholds for each region in the input BAM (or each sequence of the reference if no BED is provided)
• Simplified wf-human-sv Docker image and conda definition

Fixed

• Outdated conda environment definitions
• Docker based profiles no longer requires an internet connection to fetch the cram_cache cache building script

[v0.2.2]

Fixed

• "No such property" when using the minimap2_ubam alignment step
• Slow performance on minimap2_ubam step when providing CRAM as --ubam
• Slow performance on snp:readStats process

Removed

• "Missing reference index" warning was unnecessary

[v0.2.0]

Added

• An experimental methylation subworkflow has been integrated, using modbam2bed to aggregate modified base counts (input BAM should have MM and ML tags), enable this with --methyl

Changed

• Workflow experimentally supports CRAM as input, uBAM input uses CRAM for intermediate files
• Reference FAI is now created if it does not exist, rather than raising an error
• --sniffles_args may be used to provide custom arguments to the sniffles2 process
• Output files are uniformly prefixed with --sample_name
• Output alignment from --ubam is now CRAM formatted

Fixed

• Existence of Clair3 model directory is checked before starting workflow
• --GVCF and --include_all_ctgs are correctly typed as booleans
  • --GVCF now outputs GVCF files to the output directory as intended
  • --include_all_ctgs no longer needs to be set as --include_all_ctgs y

[v0.1.1]

Added

• --ubam_bam2fq_threads and --ubam_sort_threads allow finer control over the resourcing of the alignment step
  • The number of CPU required for minimap2_ubam is sum(ubam_bam2fq_threads, ubam_map_threads, ubam_sort_threads)

Changed

• --ubam_threads is now --ubam_map_threads and only sets the threads for the mapping specifically
• --ref is now required by the schema, preventing obscure errors when it is not provided
• Print helpful warning if neither --snp or --sv have been specified
• Fastqingress metadata map
• Disable dag creation by default to suppress graphviz warning

Fixed

• Tandem repeat BED is now correctly set by --tr_bed
• Update fastcat dependency to 0.4.11 to catch cases where input FASTQ cannot be read
• Sanitize fastq intermittent null object error

Note

• Switched to "new flavour" CI, meaning that containers are released from workflows independently

[v0.1.0]

Added

• Ported wf-human-snp (v0.3.2) to modules/wf-human-snp
• Ported wf-human-sv (v0.1.0) to modules/wf-human-sv

[v0.0.0]

• Initialised wf-human-variation from wf-template #195cab5