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DetermineGermlineContigPloidy

Determines the baseline contig ploidy for germline samples given counts data

Category Copy Number Variant Discovery


Overview

Determines the integer ploidy state of all contigs for germline samples given counts data. These should be either HDF5 or TSV count files generated by CollectReadCounts.

Introduction

Germline karyotyping is a frequently performed task in bioinformatics pipelines, e.g. for sex determination and aneuploidy identification. This tool uses counts data for germline karyotyping.

Performing germline karyotyping using counts data requires calibrating ("modeling") the technical coverage bias and variance for each contig. The Bayesian model and the associated inference scheme implemented in DetermineGermlineContigPloidy includes provisions for inferring and explaining away much of the technical variation. Furthermore, karyotyping confidence is automatically adjusted for individual samples and contigs.

Running DetermineGermlineContigPloidy is the first computational step in the GATK germline CNV calling pipeline. It provides a baseline ("default") copy-number state for each contig/sample with respect to which the probability of alternative states is allocated.

Python environment setup

The computation done by this tool, aside from input data parsing and validation, is performed outside of the Java Virtual Machine and using the gCNV computational python module, namely gcnvkernel. It is crucial that the user has properly set up a python conda environment with gcnvkernel and its dependencies installed. If the user intends to run DetermineGermlineContigPloidy using one of the official GATK Docker images, the python environment is already set up. Otherwise, the environment must be created and activated as described in the main GATK README.md file.

Tool run modes

This tool has two operation modes as described below:

COHORT mode:
If a ploidy model parameter path is not provided via the model argument, the tool will run in the COHORT mode. In this mode, ploidy model parameters (e.g. coverage bias and variance for each contig) are inferred, along with baseline contig ploidy states of each sample. It is possible to run the tool over a subset of all intervals present in the input count files, which can be specified by -L; this can be used to pass a filtered interval list produced by FilterIntervals to mask intervals from modeling. The specified intervals must be present in all of the input count files.

A TSV file specifying prior probabilities for each integer ploidy state and for each contig is required in this mode and must be specified via the contig-ploidy-priors argument. The following shows an example of such a table:


CONTIG_NAME PLOIDY_PRIOR_0 PLOIDY_PRIOR_1 PLOIDY_PRIOR_2 PLOIDY_PRIOR_3
1 0.01 0.01 0.97 0.01
2 0.01 0.01 0.97 0.01
X 0.01 0.49 0.49 0.01
Y 0.50 0.50 0.00 0.00

Note that the contig names appearing under CONTIG_NAME column must match contig names in the input counts files, and all contigs appearing in the input counts files must have a corresponding entry in the priors table. The order of contigs is immaterial in the priors table. The highest ploidy state is determined by the prior table (3 in the above example). A ploidy state can be strictly forbidden by setting its prior probability to 0. For example, the X contig in the above example can only assume 0 and 1 ploidy states.

The tool output in the COHORT mode will contain two subdirectories, one ending with "-model" and the other ending with "-calls". The model subdirectory contains the inferred parameters of the ploidy model, which may be used later on for karyotyping one or more similarly-sequenced samples (see below). The calls subdirectory contains one subdirectory for each sample, listing various sample-specific quantities such as the global read-depth, average ploidy, per-contig baseline ploidies, and per-contig coverage variance estimates.

CASE mode:
If a path containing previously inferred ploidy model parameters is provided via the model argument, then the tool will run in the CASE mode. In this mode, the parameters of the ploidy model are loaded from the provided directory and only sample-specific quantities are inferred. The modeled intervals are then specified by a file contained in the model directory, all interval-related arguments are ignored in this mode, and all model intervals must be present in all of the input count files. The tool output in the CASE mode is only the "-calls" subdirectory and is organized similarly to the COHORT mode.

In the CASE mode, the contig ploidy prior table is taken directly from the provided model parameters path and must be not provided again.

Important Remarks

Choice of hyperparameters:

The quality of ploidy model parametrization and the sensitivity/precision of germline karyotyping are sensitive to the choice of model hyperparameters, including standard deviation of mean contig coverage bias (set using the mean-bias-standard-deviation argument), mapping error rate (set using the mapping-error-rate argument), and the typical scale of contig- and sample-specific unexplained variance (set using the global-psi-scale and sample-psi-scale arguments, respectively). It is crucial to note that these hyperparameters are not universal and must be tuned for each sequencing protocol and properly set at runtime.

Mosaicism and fractional ploidies:

The model underlying this tool assumes integer ploidy states (in contrast to fractional/variable ploidy states). Therefore, it is to be used strictly on germline samples and for the purpose of sex determination, autosomal aneuploidy detection, or as a part of the GATK germline CNV calling pipeline. The presence of large somatic events and mosaicism (e.g., sex chromosome loss and somatic trisomy) will naturally lead to unreliable results. We strongly recommended inspecting genotyping qualities (GQ) from the tool output and considering to drop low-GQ contigs in downstream analyses. Finally, given the Bayesian status of this tool, we suggest including as many high-quality germline samples as possible for ploidy model parametrizaton in the COHORT mode. This will downplay the role of questionable samples and will yield a more reliable estimation of genuine sequencing biases.

Coverage-based germline karyotyping:

Accurate germline karyotyping requires incorporating SNP allele-fraction data and counts data in a unified probabilistic model and is beyond the scope of the present tool. The current implementation only uses counts data for karyotyping and while being fast, it may not provide the most reliable results.

Usage examples

COHORT mode:

 gatk DetermineGermlineContigPloidy \
   --input normal_1.counts.hdf5 \
   --input normal_2.counts.hdf5 \
   ... \
   --contig-ploidy-priors a_valid_ploidy_priors_table.tsv
   --output output_dir \
   --output-prefix normal_cohort
 

COHORT mode (with optional interval filtering):

 gatk DetermineGermlineContigPloidy \
   -L intervals.interval_list \
   -XL blacklist_intervals.interval_list \
   --input normal_1.counts.hdf5 \
   --input normal_2.counts.hdf5 \
   ... \
   --contig-ploidy-priors a_valid_ploidy_priors_table.tsv
   --output output_dir \
   --output-prefix normal_cohort
 

CASE mode:

 gatk DetermineGermlineContigPloidy \
   --model a_valid_ploidy_model_dir
   --input normal_1.counts.hdf5 \
   --input normal_2.counts.hdf5 \
   ... \
   --output output_dir \
   --output-prefix normal_case
 

DetermineGermlineContigPloidy specific arguments

This table summarizes the command-line arguments that are specific to this tool. For more details on each argument, see the list further down below the table or click on an argument name to jump directly to that entry in the list.

Argument name(s) Default value Summary
Required Arguments
--input
 -I
[] Input read-count files containing integer read counts in genomic intervals for all samples. Intervals must be identical and in the same order for all samples. If only a single sample is specified, an input ploidy-model directory must also be specified.
--output
 -O
null Output directory for sample contig-ploidy calls and the contig-ploidy model parameters for future use.
--output-prefix
null Prefix for output filenames.
Optional Tool Arguments
--adamax-beta-1
0.9 Adamax optimizer first moment estimation forgetting factor.
--adamax-beta-2
0.999 Adamax optimizer second moment estimation forgetting factor.
--arguments_file
[] read one or more arguments files and add them to the command line
--caller-external-admixing-rate
0.75 Admixing ratio of new and old called posteriors (between 0 and 1; larger values implies using more of the new posterior and less of the old posterior) after convergence.
--caller-internal-admixing-rate
0.75 Admixing ratio of new and old called posteriors (between 0 and 1; larger values implies using more of the new posterior and less of the old posterior) for internal convergence loops.
--caller-update-convergence-threshold
0.001 Maximum tolerated calling update size for convergence.
--contig-ploidy-priors
null Input file specifying contig-ploidy priors. If only a single sample is specified, this input should not be provided. If multiple samples are specified, this input is required.
--convergence-snr-averaging-window
5000 Averaging window for calculating training signal-to-noise ratio (SNR) for convergence checking.
--convergence-snr-countdown-window
10 The number of ADVI iterations during which the SNR is required to stay below the set threshold for convergence.
--convergence-snr-trigger-threshold
0.1 The SNR threshold to be reached before triggering the convergence countdown.
--disable-annealing
false (advanced) Disable annealing.
--disable-caller
false (advanced) Disable caller.
--disable-sampler
false (advanced) Disable sampler.
--gcs-max-retries
 -gcs-retries
20 If the GCS bucket channel errors out, how many times it will attempt to re-initiate the connection
--gcs-project-for-requester-pays
"" Project to bill when accessing "requester pays" buckets. If unset, these buckets cannot be accessed.
--global-psi-scale
0.001 Prior scale of contig coverage unexplained variance. If a single sample is provided, this input will be ignored.
--help
 -h
false display the help message
--initial-temperature
2.0 Initial temperature (for DA-ADVI).
--interval-merging-rule
 -imr
ALL Interval merging rule for abutting intervals
--intervals
 -L
[] One or more genomic intervals over which to operate
--learning-rate
0.05 Adamax optimizer learning rate.
--log-emission-samples-per-round
2000 Log emission samples drawn per round of sampling.
--log-emission-sampling-median-rel-error
5.0E-4 Maximum tolerated median relative error in log emission sampling.
--log-emission-sampling-rounds
100 Log emission maximum sampling rounds.
--mapping-error-rate
0.01 Typical mapping error rate.
--max-advi-iter-first-epoch
1000 Maximum ADVI iterations in the first epoch.
--max-advi-iter-subsequent-epochs
1000 Maximum ADVI iterations in subsequent epochs.
--max-calling-iters
1 Maximum number of internal self-consistency iterations within each calling step.
--max-training-epochs
100 Maximum number of training epochs.
--mean-bias-standard-deviation
0.01 Prior standard deviation of the contig-level mean coverage bias. If a single sample is provided, this input will be ignored.
--min-training-epochs
20 Minimum number of training epochs.
--model
null Input ploidy-model directory. If only a single sample is specified, this input is required. If multiple samples are specified, this input should not be provided.
--num-thermal-advi-iters
5000 Number of thermal ADVI iterations (for DA-ADVI).
--sample-psi-scale
1.0E-4 Prior scale of the sample-specific correction to the coverage unexplained variance.
--version
false display the version number for this tool
Optional Common Arguments
--exclude-intervals
 -XL
[] One or more genomic intervals to exclude from processing
--gatk-config-file
null A configuration file to use with the GATK.
--interval-exclusion-padding
 -ixp
0 Amount of padding (in bp) to add to each interval you are excluding.
--interval-padding
 -ip
0 Amount of padding (in bp) to add to each interval you are including.
--interval-set-rule
 -isr
UNION Set merging approach to use for combining interval inputs
--QUIET
false Whether to suppress job-summary info on System.err.
--tmp-dir
null Temp directory to use.
--use-jdk-deflater
 -jdk-deflater
false Whether to use the JdkDeflater (as opposed to IntelDeflater)
--use-jdk-inflater
 -jdk-inflater
false Whether to use the JdkInflater (as opposed to IntelInflater)
--verbosity
INFO Control verbosity of logging.
Advanced Arguments
--showHidden
false display hidden arguments

Argument details

Arguments in this list are specific to this tool. Keep in mind that other arguments are available that are shared with other tools (e.g. command-line GATK arguments); see Inherited arguments above.


--adamax-beta-1 / NA

Adamax optimizer first moment estimation forgetting factor.

double  0.9  [ [ 0  1 ] ]


--adamax-beta-2 / NA

Adamax optimizer second moment estimation forgetting factor.

double  0.999  [ [ 0  1 ] ]


--arguments_file / NA

read one or more arguments files and add them to the command line

List[File]  []


--caller-external-admixing-rate / NA

Admixing ratio of new and old called posteriors (between 0 and 1; larger values implies using more of the new posterior and less of the old posterior) after convergence.

double  0.75  [ [ 0  ∞ ] ]


--caller-internal-admixing-rate / NA

Admixing ratio of new and old called posteriors (between 0 and 1; larger values implies using more of the new posterior and less of the old posterior) for internal convergence loops.

double  0.75  [ [ 0  ∞ ] ]


--caller-update-convergence-threshold / NA

Maximum tolerated calling update size for convergence.

double  0.001  [ [ 0  ∞ ] ]


--contig-ploidy-priors / NA

Input file specifying contig-ploidy priors. If only a single sample is specified, this input should not be provided. If multiple samples are specified, this input is required.

File  null


--convergence-snr-averaging-window / NA

Averaging window for calculating training signal-to-noise ratio (SNR) for convergence checking.

int  5000  [ [ 0  ∞ ] ]


--convergence-snr-countdown-window / NA

The number of ADVI iterations during which the SNR is required to stay below the set threshold for convergence.

int  10  [ [ 0  ∞ ] ]


--convergence-snr-trigger-threshold / NA

The SNR threshold to be reached before triggering the convergence countdown.

double  0.1  [ [ 0  ∞ ] ]


--disable-annealing / NA

(advanced) Disable annealing.

boolean  false


--disable-caller / NA

(advanced) Disable caller.

boolean  false


--disable-sampler / NA

(advanced) Disable sampler.

boolean  false


--exclude-intervals / -XL

One or more genomic intervals to exclude from processing
Use this argument to exclude certain parts of the genome from the analysis (like -L, but the opposite). This argument can be specified multiple times. You can use samtools-style intervals either explicitly on the command line (e.g. -XL 1 or -XL 1:100-200) or by loading in a file containing a list of intervals (e.g. -XL myFile.intervals).

List[String]  []


--gatk-config-file / NA

A configuration file to use with the GATK.

String  null


--gcs-max-retries / -gcs-retries

If the GCS bucket channel errors out, how many times it will attempt to re-initiate the connection

int  20  [ [ -∞  ∞ ] ]


--gcs-project-for-requester-pays / NA

Project to bill when accessing "requester pays" buckets. If unset, these buckets cannot be accessed.

String  ""


--global-psi-scale / NA

Prior scale of contig coverage unexplained variance. If a single sample is provided, this input will be ignored.

double  0.001  [ [ 0  ∞ ] ]


--help / -h

display the help message

boolean  false


--initial-temperature / NA

Initial temperature (for DA-ADVI).

double  2.0  [ [ 0  ∞ ] ]


--input / -I

Input read-count files containing integer read counts in genomic intervals for all samples. Intervals must be identical and in the same order for all samples. If only a single sample is specified, an input ploidy-model directory must also be specified.

R List[File]  []


--interval-exclusion-padding / -ixp

Amount of padding (in bp) to add to each interval you are excluding.
Use this to add padding to the intervals specified using -XL. For example, '-XL 1:100' with a padding value of 20 would turn into '-XL 1:80-120'. This is typically used to add padding around targets when analyzing exomes.

int  0  [ [ -∞  ∞ ] ]


--interval-merging-rule / -imr

Interval merging rule for abutting intervals
By default, the program merges abutting intervals (i.e. intervals that are directly side-by-side but do not actually overlap) into a single continuous interval. However you can change this behavior if you want them to be treated as separate intervals instead.

The --interval-merging-rule argument is an enumerated type (IntervalMergingRule), which can have one of the following values:

ALL
OVERLAPPING_ONLY

IntervalMergingRule  ALL


--interval-padding / -ip

Amount of padding (in bp) to add to each interval you are including.
Use this to add padding to the intervals specified using -L. For example, '-L 1:100' with a padding value of 20 would turn into '-L 1:80-120'. This is typically used to add padding around targets when analyzing exomes.

int  0  [ [ -∞  ∞ ] ]


--interval-set-rule / -isr

Set merging approach to use for combining interval inputs
By default, the program will take the UNION of all intervals specified using -L and/or -XL. However, you can change this setting for -L, for example if you want to take the INTERSECTION of the sets instead. E.g. to perform the analysis only on chromosome 1 exomes, you could specify -L exomes.intervals -L 1 --interval-set-rule INTERSECTION. However, it is not possible to modify the merging approach for intervals passed using -XL (they will always be merged using UNION). Note that if you specify both -L and -XL, the -XL interval set will be subtracted from the -L interval set.

The --interval-set-rule argument is an enumerated type (IntervalSetRule), which can have one of the following values:

UNION
Take the union of all intervals
INTERSECTION
Take the intersection of intervals (the subset that overlaps all intervals specified)

IntervalSetRule  UNION


--intervals / -L

One or more genomic intervals over which to operate

List[String]  []


--learning-rate / NA

Adamax optimizer learning rate.

double  0.05  [ [ 0  ∞ ] ]


--log-emission-samples-per-round / NA

Log emission samples drawn per round of sampling.

int  2000  [ [ 0  ∞ ] ]


--log-emission-sampling-median-rel-error / NA

Maximum tolerated median relative error in log emission sampling.

double  5.0E-4  [ [ 0  ∞ ] ]


--log-emission-sampling-rounds / NA

Log emission maximum sampling rounds.

int  100  [ [ 0  ∞ ] ]


--mapping-error-rate / NA

Typical mapping error rate.

double  0.01  [ [ 0  ∞ ] ]


--max-advi-iter-first-epoch / NA

Maximum ADVI iterations in the first epoch.

int  1000  [ [ 0  ∞ ] ]


--max-advi-iter-subsequent-epochs / NA

Maximum ADVI iterations in subsequent epochs.

int  1000  [ [ 0  ∞ ] ]


--max-calling-iters / NA

Maximum number of internal self-consistency iterations within each calling step.

int  1  [ [ 0  ∞ ] ]


--max-training-epochs / NA

Maximum number of training epochs.

int  100  [ [ 0  ∞ ] ]


--mean-bias-standard-deviation / NA

Prior standard deviation of the contig-level mean coverage bias. If a single sample is provided, this input will be ignored.

double  0.01  [ [ 0  ∞ ] ]


--min-training-epochs / NA

Minimum number of training epochs.

int  20  [ [ 0  ∞ ] ]


--model / NA

Input ploidy-model directory. If only a single sample is specified, this input is required. If multiple samples are specified, this input should not be provided.

String  null


--num-thermal-advi-iters / NA

Number of thermal ADVI iterations (for DA-ADVI).

int  5000  [ [ 0  ∞ ] ]


--output / -O

Output directory for sample contig-ploidy calls and the contig-ploidy model parameters for future use.

R String  null


--output-prefix / NA

Prefix for output filenames.

R String  null


--QUIET / NA

Whether to suppress job-summary info on System.err.

Boolean  false


--sample-psi-scale / NA

Prior scale of the sample-specific correction to the coverage unexplained variance.

double  1.0E-4  [ [ 0  ∞ ] ]


--showHidden / -showHidden

display hidden arguments

boolean  false


--tmp-dir / NA

Temp directory to use.

String  null


--use-jdk-deflater / -jdk-deflater

Whether to use the JdkDeflater (as opposed to IntelDeflater)

boolean  false


--use-jdk-inflater / -jdk-inflater

Whether to use the JdkInflater (as opposed to IntelInflater)

boolean  false


--verbosity / -verbosity

Control verbosity of logging.

The --verbosity argument is an enumerated type (LogLevel), which can have one of the following values:

ERROR
WARNING
INFO
DEBUG

LogLevel  INFO


--version / NA

display the version number for this tool

boolean  false


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GATK version 4.1.0.0 built at Wed, 30 Jan 2019 10:21:04 +0530.