Post Somatic Calling Filtering

The following options are available for post somatic calling filtering:

Option

Description

--vc-sq-call-threshold

Emits calls in the VCF. The default is 3.0 for tumor-normal and 0.1 for tumor-only. If the value for vc-sq-filter-threshold is lower than vc-sq-call-threshold, the filter threshold value is used instead of the call threshold value.

--vc-sq-filter-threshold

Marks emitted VCF calls as filtered. The default is 17.5 for tumor-normal and 3.0 for tumor-only.

--vc-enable-triallelic-filter

Enables the multiallelic filter. The default is true.

--vc-enable-non-primary-allelic-filter

Similar to the triallelic filter, but filters less aggressively. Keep the allele per multiallelic position with highest alt AD, and only filter the rest (Default=false). Cannot be enabled when the triallelic filter is also on.

--vc-enable-af-filter

Enables the allele frequency filter for nuclear chromosomes. The default value is false. When set to true, the VCF excludes variants with allele frequencies below the AF call threshold or variants with an allele frequency below the AF filter threshold and tagged with low AF filter tag. The default AF call threshold is 1% and the default AF filter threshold is 5%. To change the threshold values, use the following command line options: vc-af-call-threshold and vc-af-filter-threshold. Use vc-enable-af-filter-mito and the corresponding threshold options for mitochondrial allele frequency filtering.

--vc-enable-non-homref-normal-filter

Enables the non-homref normal filter. The default value is true. When set to true, the VCF filters out variants if the normal sample genotype is not a homozygous reference.

--vc-enable-vaf-ratio-filter

Adds one condition to be filtered out by the alt_allele_in_normal filter. The default value is false. When set to true, the VCF filters out variants if the normal sample AF is greater than 20% of tumor sample AF.

--vc-depth-filter-threshold

Filters all somatic variants (alt or homref) with a depth below this threshold. The default value is 0 (no filtering).

vc-homref-depth-filter-threshold

In gVCF mode, filters all somatic homref variants with a depth below this threshold. The default value is 3.

vc-depth-annotation-threshold

Filters all non-PASS somatic alt variants with a depth below this threshold. The default value is 0 (no filtering).

The following table describes the available somatic calling filters.

Somatic Mode

Filter ID

Description

Tumor-Only & Tumor-Normal

weak_evidence

Variant does not meet likelihood threshold. The likelihood ratio for SQ tumor-normal is < 17.5 or < 3.0 for SQ tumor-only.

Tumor-Only & Tumor-Normal

multiallelic

Site filtered if there are two or more ALT alleles at this location in the tumor.

Tumor-Only & Tumor-Normal

str_contraction

Suspected PCR error where the ALT allele is one repeat unit less than the reference. Enabled only when using --vc-enable-gatk-acceleration=true.

Tumor-Only & Tumor-Normal

base_quality

Median base quality of ALT reads at this locus is < 20.

Tumor-Only & Tumor-Normal

mapping_quality

Median mapping quality of ALT reads at this locus is < 20 (tumor-normal) or < 30 (tumor-only).

Tumor-Only & Tumor-Normal

fragment_length

Absolute difference between the median fragment length of alt reads and median fragment length of ref reads at a given locus > 10000.

Tumor-Only & Tumor-Normal

read_position

Median of distances between the start and end of read and a given locus < 5 (the variant is too close to edge of all the reads).

Tumor-Only & Tumor-Normal

low_af

Allele frequency is below the threshold specified with --vc-af-filter-threshold (default is 5%). Enabled only when using --vc-enable-af-filter=true.

Tumor-Only & Tumor-Normal

systematic_noise

If AQ score is < 10 (default) for tumor-normal or < 60 (default) for tumor-only, the site is filtered.

Tumor-Only & Tumor-Normal

low_frac_info_reads

The fraction of informative reads (denominator excludes filtered_out reads) is below the threshold. The default threshold value is 0.5.

Tumor-Only & Tumor-Normal

low_depth

The site was filtered because the number of reads is too low. The filter is off by default.

Tumor-Only & Tumor-Normal

low_tlen

The site was filtered because the fraction of low TLEN ALT supporting reads is above a threshold. The default threshold is 0.4. Reads with TLEN smaller than -2.25 (default) standard deviations from the mean are considered to be low TLEN. This filter is not applied for reads sampled from tight insert distributions ie, stddev / mean < 0.1 (default)

Somatic Mode

Filter ID

Description

Tumor-Normal

noisy_normal

More than three alleles are observed in the normal sample at allele frequency above 9.9%.

Tumor-Normal

alt_allele_in_normal

ALT allele frequency in the normal sample is above 0.2 plus the maximum contamination tolerance. For solid tumor mode, the value is 0. For liquid tumor mode, the default value is 0.15. See vc-enable-vaf-ratio-filter for optional conditions.

Tumor-Normal

filtered_reads

More than 90% of reads have been filtered out.

Tumor-Normal

no_reliable_supporting_read

No reliable supporting read was found in the tumor sample. A reliable supporting read is a read supporting the alt allele with mapping quality ≥ 40, fragment length ≤ 10,000, base call quality ≥ 25, and distance from start/end of read ≥ 5.

Tumor-Normal

too_few_supporting_reads

Variant is supported by < 3 reads in the tumor sample.

Tumor-Normal

non_homref_normal

Normal sample genotype is not a homozygous reference.

Tumor-Normal

germline_risk

Likelihood of allele being present in normal > 0.025. Enabled only when using --vc-enable-gatk-acceleration=true.

Tumor-Normal

artifact_in_normal

TLOD of the normal read set (Normal artifact LOD) is > 0.0. Not called normal artifact if allele fraction in normal is much smaller than allele fraction in tumor (normalAlleleFraction < (0.1 * tumorAlleleFraction). Enabled only when using --vc-enable-gatk-acceleration=true.

QUAL is not output in the somatic variant records. Instead, the confidence score is FORMAT/SQ.

##FORMAT=<ID=SQ,Number=1,Type=Float,Description="Somatic quality">

The field is specific to the sample. For the tumor samples, the field quantifies the evidence that a somatic variant is present at a given locus.

If a normal sample is also available, the corresponding FORMAT/SQ value quantifies the evidence that the normal sample is a homozygous reference at a given locus.

GQ is not output in the somatic variant records, because DRAGEN does not test for multiple diploid genotype candidates. Instead, an ALT allele is considered as a candidate somatic variant. If tumor SQ > vc-sq-call-threshold (default is 3), then the FORMAT/GT for the tumor sample is hard-coded to 0/1, and the FORMAT/AF yields an estimate on the somatic variant allele frequency, which ranges anywhere within [0,1].

• If tumor SQ < vc-sq-call-threshold, the variant is not emitted in the VCF.
• If tumor SQ > vc-sq-call-threshold but tumor SQ < vc-sq-filter-threshold, the variant is emitted in the VCF, but FILTER=weak_evidence.
• If tumor SQ > vc-sq-call-threshold and tumor SQ > vc-sq-filter-threshold, the variant is emitted in the VCF and FILTER=PASS (unless the variant is filtered by a different filter).
• The default vc-sq-filter-threshold is 17.5 for tumor-normal and 3.0 for tumor-only analysis.

The following is an example somatic T/N VCF record. Tumor SQ > vc-sq-call-threshold but tumor SQ < vc-sq-filter-threshold, so the FILTER is marked as weak_evidence.

2 593701 . G A . weak_evidence

DP=97;MQ=48.74;SQ=3.86;NLOD=9.83;FractionInformativeReads=1.000

GT:SQ:AF:F1R2:F2R1:DP:SB:MB 0/0:9.83:33,0:0.000:14,0:19,0:33 0/1:3.86:61,3:0.047:29,2:32,1:64:35,26,0,3:39,22,1,2

The clustered-events penalty is an exception to the above rule for emitting variants. By default, the clustered-events penalty replaces the clustered-events filter. Instead of applying a hard filter when too many events are clustered together, DRAGEN applies a penalty to the SQ scores of cophased clustered events. Clustered events with weak evidence are no longer called, but clustered events with strong evidence can still be called. This is equivalent to lowering the prior probability of observing clustered cophased variants. The penalty is applied after the decision to emit variants, so that penalized variants still appear in the VCF if their unpenalized score is high enough. Variants that are combined into an MNV via the --combine-phased-variants-distance option are treated as a single variant for the purposes of the penalty. To disable the clustered-events penalty, set --vc-clustered-event-penalty=0.