<tool id="hydra" name="Hydra pipeline" version="0.1.0">
    <description>Identifies drug resistance within an NGS dataset</description>
    <requirements>
          <requirement type="package" version="0.2.2">quasitools</requirement>
    </requirements>
    <command detect_errors="exit_code"><![CDATA[

        quasitools hydra '$forward' 

        #if $reverse:
            '$reverse' 
        #end if

        #if $mutation_db:
            -m '$mutation_db' 
        #end if

        #if $reporting_threshold:
            -rt '$reporting_threshold' 
        #end if

        #if $consensus_pct:
            -cp '$consensus_pct' 
        #end if

        #if $length_cutoff:
            -lc '$length_cutoff' 
        #end if

        #if $score_cutoff:
            -sc '$score_cutoff' 
        #end if

        #if $error_rate:
            -e '$error_rate' 
        #end if

        #if $min_qual:
            -mq '$min_qual' 
        #end if

        #if $min_depth:
            -md '$min_depth' 
        #end if

        #if $min_ac:
            -ma '$min_ac' 
        #end if

        #if $min_freq:
            -mf '$min_freq' 
        #end if

        #if $generate_consensus:
            --generate_consensus 
        #end if

        #if $filter_ns:
            --ns 
        #end if

        -o output

    ]]></command>
    <inputs>
        <param name="forward" type="data" format="fastq" optional="false" label="Forward read" />
        <param name="reverse" type="data" format="fastq" optional="true" label="Reverse read" help="Not required."/>
        <param name="mutation_db" type="data" format="tsv" optional="true" label="Mutation DB" help="Defaults to HIV mutation database." />
        <param name="reporting_threshold" type="integer" optional="true" min="1" max="100" value="1" label="Reporting threshold. Defaults to 1." help="Minimum mutation frequency to report." />
        <param name="consensus_pct" type="integer" optional="true" min="1" max="20" value="20" label="Reporting threshold" help="Minimum mutation frequency to report. Defaults to 20." />
        <param name="length_cutoff" type="integer" optional="true" min="0" max="1000" label="Length cutoff" value="100" help="Reads which fall short of the specified length will be filtered out. Defaults to 100." />
        <param name="score_cutoff" type="integer" optional="true" min="0" max="40" label="Score cutoff" value="30" help="Reads whose average quality score is less than the specified score will be filtered out. Defaults to 30." />
        <param name="error_rate" type="float" optional="true" min="0" max="1" label="Error rate" value="0.0021" help="Estimated sequencing error rate. Defaults to 0.0021."/>
        <param name="min_qual" type="integer" optional="true" min="1" max="100" label="Minimum quality" value="30" help="Minimum required quality for variant to be considered later on in the pipeline. Defaults to 30." />
        <param name="min_depth" type="integer" optional="true" min="0" max="5000" label="Minimum depth" value="100" help="Minimum required depth for variant to be considered later on in the pipeline. Defaults to 100." />
        <param name="min_ac" type="integer" optional="true" min="0" max="5000" label="Minimum allele count" value="5" help="Minimum required allele count for variant to be considered later on in the pipeline. Defaults to 5." />
        <param name="min_freq" type="float" optional="true" min="0" max="1" label="Minimum frequency" value="0.01" help="Minimum required frequency for variant to be considered later on in the pipeline. Defaults to 0.01." />
        <param name="generate_consensus" type="boolean" truevalue="--generate_consensus" falsevalue="" checked="False" label="Generate consensus" />
        <param name="filter_ns" type="boolean" truevalue="--ns" falsevalue="" checked="False" label="Filter out n's" />
    </inputs>
    <outputs>
        <data format="bam" label="HyDRA: alignment bam output" name="output_bam" from_work_dir="output/align.bam" />
        <data format="bam" label="HyDRA: bam.bai output" name="output_bam_bai" from_work_dir="output/align.bam.bai" />
        <data format="csv" label="HyDRA: coverage output" name="output_coverage" from_work_dir="output/coverage_file.csv" />
        <data format="csv" label="HyDRA: drug resistance output" name="output_dr" from_work_dir="output/dr_report.csv" />
        <data format="fastq" label="HyDRA: filtered reads output" name="output_filtered" from_work_dir="output/filtered.fastq" />
        <data format="vcf" label="HyDRA: variants output" name="output_hydra" from_work_dir="output/hydra.vcf" />
        <data format="vcf" label="HyDRA: aa mutations output" name="output_aa_mt" from_work_dir="output/mutation_report.hmcf" />
        <data format="txt" label="HyDRA: stats output" name="output_stats" from_work_dir="output/stats.txt" />
    </outputs>
    <tests>
        <test>
            <param name="forward" value="forward.fastq" />
            <output name="output_coverage">
                <assert_contents>
                    <has_text text="frame: 0" />
                    <has_text text="1,0" />
                    <has_text text="948,0" />
                </assert_contents>
            </output>
            <output name="output_dr">
                <assert_contents>
                    <has_text text="Chromosome,Gene,Category,Surveillance,Wildtype,Position,Mutation,Mutation Frequency,Coverage" />
                    <has_text text="hxb2_pol,RT,NNRTI,Yes,K,101,P,14.23,1574" />
                    <has_text text="hxb2_pol,RT,NNRTI,Yes,K,103,N,5.49,1912" />
                    <has_text text="hxb2_pol,RT,NNRTI,Yes,Y,181,C,24.07,4557" />
                    <has_text text="hxb2_pol,RT,NNRTI,Yes,Y,181,I,18.04,4557" />
                    <has_text text="hxb2_pol,RT,NNRTI,Yes,Y,181,V,20.08,4557" />
                    <has_text text="hxb2_pol,RT,NNRTI,Yes,Y,188,C,2.81,3454" />
                    <has_text text="hxb2_pol,RT,NNRTI,Yes,G,190,A,5.20,3233" />
                    <has_text text="hxb2_pol,RT,NNRTI,Yes,G,190,S,6.68,3233" />
                </assert_contents>
            </output>
            <output name="output_hydra">
                <assert_contents>
                    <has_text_matching expression="#CHROM\tPOS\tID\tREF\tALT\tQUAL\tFILTER\tINFO"/>
                    <has_text_matching expression="hxb2_pol\s576\s.\sa\sg\s100\sPASS\sDP=805;AC=245;AF=0.3043" />
                    <has_text_matching expression="hxb2_pol\s958\s.\sc\sa\s100\sPASS\sDP=2503;AC=28;AF=0.0112" />
                </assert_contents>
            </output>
            <output name="output_aa_mt">
                <assert_contents>
                    <has_text_matching expression="#CHROM\sGENE\sTYPE\sWILDTYPE\sPOS\sMUTANT\sFILTER\sMUTANT_FREQ\sCOVERAGE\sINFO"/>
                    <has_text_matching expression="hxb2_pol\sRT\smutation\sK\s101\sP\sPASS\s0.1423\s1574\sWC=aaa;MC=CCa;MCF=0.1423;CAT=NNRTI;SRVL=Yes" />
                    <has_text_matching expression="hxb2_pol\sRT\smutation\sH\s221\sN\sPASS\s0.0113\s2475\sWC=cat;MC=Aat;MCF=0.0113;CAT=.;SRVL=." />
                </assert_contents>
            </output>
            <output name="output_stats">
                <assert_contents>
                    <has_text text="Input Size: 25000"/>
                    <has_text text="Number of reads filtered due to length: 15074"/>
                    <has_text text="Number of reads filtered due to average quality score: 501"/>
                    <has_text text="Number of reads filtered due to presence of Ns: 0"/>
                    <has_text text="Number of reads filtered due to excess coverage: 0"/>
                    <has_text text="Number of reads filtered due to poor mapping: 12"/>
                    <has_text text="Percentage of reads filtered: 62.35"/>
                </assert_contents>
            </output>
        </test>
    </tests>
    <help><![CDATA[

HyDRA - HIV Drug Resistance Analyzer
====================================

The HyDRA pipeline provides a pipeline for identifying drug resistance within a Next Generation Sequencing dataset. The pipeline takes as input the raw reads produced by a Next Generation Sequencer and produces a report detailing found drug resistance per sample.

Authors
-------

The HyDRA pipeline was developed by Eric Enns and David Peddle.

Stages
------

The HyDRA pipleine proceeds through the following stages:

1. Quality Control/Filtering
2. Reference mapping using bowtie2.
3. Variant Calling and filtering using a Poisson distribution.
4. AA Mutation Calling and filtering.
5. Drug Resistance report generation.

Details
-------

The following is an example for running the pipeline, using our included test dataset:
    * Output directory name: "/tmp/hydra_out"
    * Forward reads: "reads_w_K103N.fastq"

### Output ###

The detailed output directory tree looks as follows:

    /tmp/hydra_out/
        * align.bam
        * align.bam.bai
        * coverage_file.csv
        * dr_report.csv
        * filtered.fastq
        * hydra.vcf
        * mutation_report.hmcf
        * stats.txt

The description of each of these directories/files are as follows:

* __run.conf__: The configuration used when this output was produced.
* __reads_w_K103N/__: The results directory for the input file reads_w_K103N.fastq
    * __align.bam__: The alignment file in bam format.
    * __align.bam.bai__: The index to the alignment file.
    * __coverage_file.csv__: A file with one entry per line with the AA position and the coverage at the position.
    * __dr_report.csv__: A report detailing the drug resistant mutations found, above the reporting threshold (default: 1%).
    * __filtered.fastq__: The reads remaining after the filtering stage.
    * __hydra.vcf__: The variants found by the pipeline.
    * __mutation_report.hmcf__: The AA mutations found by the pipeline.
    * __stats.txt__: A log file detailing size after filtering and major stages.

The __dr_report.csv__ file lists all found drug resistant mutations (mutations included in the mutation database) which have frequency greater than the reporting threshold. An example of this file is given below.

Example: __dr_report.csv__

    Gene,Category,Surveillance,Wildtype,Position,Mutation,Mutation Frequency,Coverage
    RT,NNRTI,Yes,K,103,N,9.03,155

The __mutation_report.hmcf__ files is our custom VCF like file which details all of the AA mutations found by the pipeline. An example if this file is given below.

Example: __mutation_report.hmcf__

    ##fileformat=HMCFv1
    ##fileDate=20150008
    ##source=HyDRA
    ##reference=/home/ericenns/hydra/var/hxb2_pol.fas
    ##INFO=<ID=MC,Number=.,Description="String">
    ##INFO=<ID=MCF,Number=.,Description="String">
    ##INFO=<ID=WC,Number=.,Description="String">
    ##FILTER=<ID=mf0.01,Description="Mutant freq below 0.01">
    #GENE   CATEGORY        SURVEILLANCE    TYPE    WILDTYPE        POS     MUTANT  FILTER  MUTANT_FREQ     COVERAGE INFO
    RT      NNRTI   Yes     mutation        K       103     N       PASS    0.0903  155     WC=aaa;MC=aaC;MCF=0.0903

    ]]></help>
    <citations>
    </citations>
</tool>