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README.md

venn

VCF-comp

VCF-comp is a Scala library for pairwise comparison of annotated VCF files. Uses Apache Spark, ADAM and adam-fx.

VCF-comp is intended for performing VCF analyses using the Scala programming language within a Spark-notebook environment.

VCF-comp is open source software, available on both Github and BitBucket.

VCF-comp artifacts are published to Bintray. Latest version: 0.4.0

--

CHANGE NOTES

  • 0.4.0 Added filtering functions:
    • filtering by gene name(s) with respect to .bed and .gtf files
    • filtering by contig
  • 0.3.0 Stable version

GETTING STARTED

Using the Docker image

TODO

(A Docker image containing Spark Notebook and example notebooks with VCF-comp analyses will be available in the near future)

Manual setup

We will assume you have a Spark-notebook instance available. If not, have a look at the launch instructions.

Remote artifact repository

First, we specify the remote Maven repository on Bintray from which the VCF-comp library artifact are available.

:remote-repo bintray-tmoerman % default % http://dl.bintray.com/tmoerman/maven % maven

Library dependencies

Next, we specify the library dependencies. We need both the VCF-comp library and the BDGenomics Adam library, but without Hadoop and Spark dependencies, because these are already provided automatically in the Spark Notebook environment.

:dp org.tmoerman % vcf-comp_2.10 % 0.4.0
:dp org.bdgenomics.adam % adam-core % 0.17.1
- org.apache.hadoop % hadoop-client %   _
- org.apache.spark  %     _         %   _
- org.scala-lang    %     _         %   _
- org.scoverage     %     _         %   _
- joda-time         %     _         %   _

Initialize the SparkContext

We are now ready to configure the running SparkContext in the notebook. VCF-comp uses Adam's rich domain classes, therefore we need to define Kryo as the Spark serializer and the Adam-FX Kryo registrator. Don't worry too much about this, just execute the following snippet in a Spark Notebook cell.

reset(lastChanges = _.set("spark.app.name", "My VCF analysis")
                     .set("spark.serializer", "org.apache.spark.serializer.KryoSerializer")
                     .set("spark.kryo.registrator", "org.tmoerman.adam.fx.serialization.AdamFxKryoRegistrator")
                     .set("spark.kryoserializer.buffer", "32m")
                     .set("spark.kryoserializer.buffer.max value", "128")
                     .set("spark.kryo.referenceTracking", "true"))

Import VCF-comp functionality

Almost there! The final setup step is to import the necessary VCF-comp classes and functions.

import scala.language.implicitConversions

import org.tmoerman.vcf.comp.VcfComparisonContext._
import org.tmoerman.vcf.comp.core.Model._
import org.tmoerman.vcf.comp.viz.SparkNoteBookDimpleGraphs._

import org.tmoerman.adam.fx.avro.AnnotatedGenotype

implicit def toDimpleChart(chart: DimpleChart) = chart match {
  case DimpleChart(data, js, s) => DiyChart(data, js, maxPoints = chart.maxPoints, sizes = s)
}

Test the setup

We can now test the setup of the library by executing the .help function on the SparkContext variable (sc or sparkContext).

sc.help

This method should return a list of methods we can invoke on the SparkContext:

res9: String =
- getMetaFields
- startQcComparison
- startSnpComparison

This is VCF-comp's so-called discoverable API in action. We discuss this concept in more detail later.

Well done! We are now ready to perform an actual pairwise VCF comparison analysis! Read on to find out how.

USAGE

Some proficiency in Scala is expected. Don't worry, the level of Scala programming required to effectively use this library is quite basic. If you have some experience with Java, Python, C++ or C#, most of the code in this tutorial will feel very familiar.

The motivation for adopting a programmatic Notebook approach in this tool is because this is essentially a data science tool. Data science tools ideally show exactly how an analysis has been performed, by making the individual steps explicit as code. Although the threshold to get started is higher than a point-and-click interface, this disadvantage is outweighed in the long run by the benefits of the reproducible nature of a notebook approach.

VCF-comp focuses on pairwise comparison of VCF files, so let's get two interesting files ready: tumor.vcf and normal.vcf. In this example, we read these files from HDFS:

val workDir = "hdfs://bla:54310/my-data/VCF/"

val tumor  = workDir + "tumor.vcf"
val normal = workDir + "normal.vcf"

We can now start two types of VCF comparison: a quality control (QC) comparison and a SNP comparison. Let's start with a QC comparison.

Starting a QC comparison

To start a QC comparison, we invoke the appropriate method on the SparkContext instance sc. Remember, as previously mentioned, we can always invoke the .help method on different objects in the analysis, to give us a list of methods available at that point. Let's do that one more time:

sc.help
res9: String =
- getMetaFields
- startQcComparison
- startSnpComparison

The method startQcComparison is the one we need, let's invoke it while setting some parameters:

val qcParams = new ComparisonParams(labels = ("TUMOR", "NORMAL")) // parameters

val qcComparison = sc.startQcComparison(tumor, normal, qcParams) // a Spark RDD
                     .cache()                                    // cache the RDD

The ComparisonParams object defines labels for our VCF files. The result of invoking startQcComparison is a Spark RDD, which we cache in memory because it is an intermediate result we will use in future computations. We store this RDD in a value named qcComparison.

Let's once again use the .help method to discover how we can proceed. This time we don't invoke it on the SparkContext sc, but on the qcComparison value.

qcComparison.help
res11: String =
- countByProjection
- countByTraversableProjection
- indelLengthDistribution
- snpCountByContig
- snpQualityDistribution
- snpReadDepthDistribution
- variantTypeCount

Looks like we can do some interesting analyses! Let's have a look at the number of SNPs we can find in the different contigs, so we choose snpCountByContig.

qcComparison.snpCountByContig

Oops, now we get some gobbledigook:

res16: Iterable[org.tmoerman.vcf.comp.core.Model.QcProjectionCount[String]] = List(QcProjectionCount(NORMAL,9,1361),
QcProjectionCount(NORMAL,1,3304), QcProjectionCount(TUMOR,20,844), QcProjectionCount(NORMAL,12,1564),
QcProjectionCount(TUMOR,19,2607), QcProjectionCount(TUMOR,21,476), QcProjectionCount(TUMOR,1,3376),
QcProjectionCount(TUMOR,13,547), QcProjectionCount(TUMOR,Y,12), QcProjectionCount(NORMAL,11,2300),
QcProjectionCount(TUMOR,22,733), QcProjectionCount(NORMAL,2,2197), QcProjectionCount(TUMOR,12,1520),
QcProjectionCount(NORMAL,7,1522), QcProjectionCount(NORMAL,16,1387), QcProjectionCount(NORMAL,18,54...

That doesn't look right, we'd prefer to see some graphical output. Let's consult the .help function once more, this time we invoke it on the result of the qcComparison.snpCountByContig calculation:

qcComparison.snpCountByContig.help
res17: String =
- groupedBarChart
- lineChart

Okay, a histogram is probably the most sensible chart for a SNP count by contig, so let's choose that one:

qcComparison.snpCountByContig
            .groupedBarChart(x_title = "SNP count by contig",
                             x_order = true)

We also specified some overriding attributes of the grouped bar chart. The result is an object that is automagically turned into a chart by the notebook.

chart

Nice!

This sequence of steps illustrates the primary usage pattern of the VCF-comp library. See section QC analyses for an overview of QC functionality.

Let's now apply this for a SNP comparison.

Starting a SNP comparison

Analogously, we launch a SNP comparison.

val snpParams = new ComparisonParams(labels = ("TUMOR", "NORMAL"))

val snpComparison = sc.startSnpComparison(tumor, normal, snpParams) // a Spark RDD
                      .cache()                                      // cache the RDD		

In the same spirit as the QC example, we create a Spark RDD instance by invoking the startSnpComparison function on the SparkContext sc. Analogously, we can inspect the list of available functions by invoking the .help function on the snpComparison value.

snpComparison.help
res17: String =
- alleleFrequencyDistribution
- allelesSwitchCount
- baseChangeCount
- baseChangePatternCount
- baseChangeTypeCount
- categoryCount
- clinvarRatio
- commonSnpRatio
- countByProjection
- countBySwitch
- countByTraversableProjection
- functionalAnnotationCount
- functionalImpactCount
- qualityDistribution
- readDepthDistribution
- synonymousRatio
- transcriptBiotypeCount
- viewOnly
- zygosityCount
- zygositySwitchCount

Let's choose the readDepthDistribution analysis, and inspect which visualizations are available. Note that not all visualization make sense for a "distribution" type analysis.

snpComparison.readDepthDistribution().help
res21: String =
- groupedBarChart
- lineChart
- lollipopPieChart
- percentageBarChart
- stackedBarChart
- table

In this case, a line chart makes most sense, so let's try that.

snpComparison.readDepthDistribution(step = 5)
             .lineChart(width = 700,
                        x_title = "read depth distribution")

RD

Sweet!

Notice that again, we have overridden some of the default arguments in both the readDepthDistribution as well as the lineChart methods. Check the documentation for more information about which arguments are available on the analysis methods.

This concludes two examples of how to launch a QC comparison and a SNP comparison. The next section deals with the core concept of the SNP comparison functionality: the 5 concordance categories.

CONCORDANCE CATEGORIES

The heart of VCF-comp is an algorithm that matches variants per position by concordance. By concordance, we mean a degree to which both files agree about the variants or their genotypes on that position, according to a configurable matching criterion. VCF-comp defines 5 categories:

name meaning
A-unique File A has a variant on this position, file B does not
B-unique File B has a variant on this position, file A does not
Concordant Both file A and B have the same variant on this position with respect
to the matching criterion. A concordant variant is counted once.
A-discordant File A and B have a variant on this position, but do not agree with
respect to the matching criterion. This variant is the one from file A.
B-discordant File A and B have a variant on this position, but do not agree with
respect to the matching criterion. This variant is the one from file B.

The default matching criterion is: matching genotype alleles in both files A and B.

In our opinion, this is the most sensible default behaviour. However, a researcher might want to define a different matching criterion. This is possible. The ComparisonParams class defines the default behaviour, but this is overridable, as illustrated in following example:

val snpParamsAA = new ComparisonParams(
  labels = ("TUMOR", "NORMAL"),
  matchFunction = (gt: AnnotatedGenotype) => gt.getGenotype.getVariant.getAlternateAllele) // overridden match function

val snpComparisonAA = sc.startSnpComparison(tumor, normal, snpParamsAA)
                        .cache()

The matchFunction is the function used in the matching algorithm to determine concordance. The matchFunction can return anything, or in Scala jargon: the function has the Any return type. If file A and B have a variant on a position, this function is applied on the corresponding genotypes and the results are compared for equality. If the results are equal we have a concordant variant, if they are not equal we have two discordant variants.

Here we have overridden it with a function that takes an AnnotatedGenotype and returns the alternate allele of the variant associated with the genotype. This is a less strict matching criterion than the default one, and could be useful for a particular analysis.

Note that overriding the matchFunction only affects the balance between concordant and discordant variants. The unique variants remain stable.

OVERVIEW OF ANALYSES

This section contains an overview of all available analyses. Additionally, the higher-order functions that act as building blocks for the available analyses and ad-hoc analyses are discussed. Using the building block functions requires a more advanced mastery of the Scala language.

TODO (Scala docs will be hosted on a dedicated site in the near future.)

QC

QC analyses

def variantTypeCount
def snpCountByContig
snpReadDepthDistribution(step: Int = DEFAULT_READ_DEPTH_STEP)
def snpQualityDistribution(step: Double = DEFAULT_QUALITY_STEP)
def indelLengthDistribution

QC analysis building blocks

def countByProjection[P: ClassTag](projection: VariantContext => P): Iterable[QcProjectionCount[P]]
def countByTraversableProjection[P: ClassTag](projection: VariantContext => Traversable[P]): Iterable[QcProjectionCount[P]]

SNP comparison

SNP comparison analyses

/**
  * @param occurrences vararg. Accepts one or more Strings from "unique", "concordant", "discordant".
  *
  * @return Returns the RDD, filtered on the specified occurrences.
  */
def viewOnly(occurrences: String*)
/**
  * @return Returns SNP count by concordance category.
  */
def categoryCount
/**
  * @return Returns SNP count by base change per concordance category.
  *
  *         Base change is a String: "ref->alt" where ref and alt are the variant alleles, e.g. "A->T", "G->C", etc...
  */
def baseChangeCount
/**
  * @return Returns SNP count by base change pattern per concordance category.
  *
  *         Base change pattern is a String: "ref:alt", analogous to base change, but without taking into account
  *         the order of ref to alt, e.g. "A:T", "C:G", etc...
  */
def baseChangePatternCount
/**
  * @return Returns SNP count by base change type per concordance category.
  *
  *         Base change types are "Ti" (Transition) and "Tv" (Transversion).
  */
def baseChangeTypeCount
/**
  * @return Returns SNP count by zygosity per concordance category.
  *
  *         Zygosity values are: "HOM_REF", "HET", "HOM_ALT", "NO_CALL".
  */
def zygosityCount
/**
  * @return Returns SNP count by functional impact per concordance category.
  *
  *         Functional impact is a scale value provided by SnpEff: "HIGH", "MODERATE", "LOW" and "MODIFIER".
  *
  *         Cfr. SnpEff http://snpeff.sourceforge.net/VCFannotationformat_v1.0.pdf
  */
def functionalImpactCount
/**
  * @return Returns SNP count by functional annotation per concordance category.
  *
  *         Functional annotation is a SnpEff annotation, including: "synonymous_variant", "missense_variant",
  *         "stop_gained", "start_lost".
  *
  *         Assumes annotation with SnpEff http://snpeff.sourceforge.net/VCFannotationformat_v1.0.pdf
  */
def functionalAnnotationCount
/**
  * @return Returns SNP count by transcript biotype per concordance category.
  *
  *         Transcript biotype is a SnpEff annotation: including: "protein_coding", "retained_intron"
  *         "nonsense_mediated_decay".
  *
  *         Assumes annotation with SnpEff http://snpeff.sourceforge.net/VCFannotationformat_v1.0.pdf
  */
def transcriptBiotypeCount
/**
  * @param label (optional). Maps the Boolean to a descriptive label.
  *
  * @return Returns the ratio of SNPs with a Clinvar annotation vs. SNPs without.
  *
  *         Assumes VCF annotation with SnpSift http://snpeff.sourceforge.net/SnpSift.html
  */
def clinvarRatio(label: Boolean => String)
/**
  * @param label (optional). Maps the Boolean to a descriptive label.
  *
  * @return Returns the ratio of SNPs with a DbSNP annotation vs. SNPs without.
  *
  *         Assumes VCF annotation with SnpSift http://snpeff.sourceforge.net/SnpSift.html
  */
def commonSnpRatio(label: Boolean => String)
/**
  * @param label (optional). Maps the boolean to a descriptive label.
  *
  * @return Returns the ratio of SNPs with "synonymous_variant" vs. "missense_variant" annotation. If a SNP has
  *         neither annotation, it is not taken into account.
  *
  *         Assumes annotation with SnpEff http://snpeff.sourceforge.net/VCFannotationformat_v1.0.pdf
  */
def synonymousRatio(label: Boolean => String)
/**
  * @param step (optional). Size of the step interval for binning the read depth values.
  *
  * @return Returns the distribution of SNPs by read depth.
  */
def readDepthDistribution(step: Int = DEFAULT_READ_DEPTH_STEP)
/**
  * @param step (optional). Size of the step interval for binning the quality values.
  *
  * @return Returns the distribution of SNPs by quality.
  */
def qualityDistribution(step: Double = DEFAULT_QUALITY_STEP)
/**
  * @param step (optional). Size of the step interval for binning the allele frequency values.
  *
  * @return Returns the distribution of SNPs by allele frequency.
  */
def alleleFrequencyDistribution(step: Double = DEFAULT_ALLELE_FREQUENCY_STEP)
def zygositySwitchCount
def allelesSwitchCount

SNP comparison building blocks

def countByProjection[P](projection: AnnotatedGenotype => P): Iterable[CategoryProjectionCount[P]]
def countByTraversableProjection[P](projection: AnnotatedGenotype => Traversable[P]): Iterable[CategoryProjectionCount[P]]
def countBySwitch[P](projection: AnnotatedGenotype => Option[P]): Iterable[CategoryProjectionCount[String]]

Visualizations

TODO

HOW IT WORKS

Pimp my library

VCF-comp makes use of a Scala idiom called "Pimp my library", through Scala's implicit conversions.

TODO

Dimple.js

TODO

http://snpeff.sourceforge.net/VCFannotationformat_v1.0.pdf

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