error: INFO: processed 0 reads for BAM file

[cccnrc]

I am struggling having Whamg running for a WGS .bam file. I used GATKSV/Wham docker image and the following command:

whamg \
    -c ${chr_list} \
    -x 30 \
    -a ${reference_fasta} \
    -f ${bam_file}

Returns:

Invoking whamg:
whamg -c /var/lib/cwl/stg5910348b-6df0-43de-baff-568ee78530fc/hg38.primary_contig.csv -x 30 -a /var/lib/cwl/stg85dd1661-e28a-42b5-a410-ee90e45240cc/Homo_sapiens_assembly38.fasta -f /var/lib/cwl/stg80c5a3ef-bc35-40ce-9482-f42a56486e04/DAKIKI.sorted.markup.bam

INFO: WHAM will analyze seqid: /var/lib/cwl/stg5910348b-6df0-43de-baff-568ee78530fc/hg38.primary_contig.csv
INFO: OpenMP will roughly use 30 threads
INFO: fasta file: /var/lib/cwl/stg85dd1661-e28a-42b5-a410-ee90e45240cc/Homo_sapiens_assembly38.fasta
INFO: target bams:
/var/lib/cwl/stg80c5a3ef-bc35-40ce-9482-f42a56486e04/DAKIKI.sorted.markup.bam
INFO: gathering stats (may take some time) for bam: /var/lib/cwl/stg80c5a3ef-bc35-40ce-9482-f42a56486e04/DAKIKI.sorted.markup.bam
INFO: processed 0 reads for: /var/lib/cwl/stg80c5a3ef-bc35-40ce-9482-f42a56486e04/DAKIKI.sorted.markup.bam

and nothing else. I also tried to reconvert the .bam to .fastq and realign with BWA-MEM:

bedtools bamtofastq -i DAKIKI.final.sorted.bam -fq DAKIKI.final.sorted.fq
bwa mem -t 30 -R "@RG\tID:DAKIKI\tSM:DAKIKI" \
          /media/kong/enrico/fasta_X01_hg38/Homo_sapiens_assembly38.fasta \
          /media/kong/enrico/wgs_cnv/DAKIKI.final.sorted.fq \
               > /media/kong/enrico/wgs_cnv/DAKIKI.sam
samtools view -@ 30 -F 4 -S -b -h /media/kong/enrico/wgs_cnv/DAKIKI.sam > /media/kong/enrico/wgs_cnv/DAKIKI.bam
samtools sort -@ 30 -n  /media/kong/enrico/wgs_cnv/DAKIKI.bam -o  /media/kong/enrico/wgs_cnv/DAKIKI.sorted.bam
samtools fixmate -@ 30 -m /media/kong/enrico/wgs_cnv/DAKIKI.sorted.bam /media/kong/enrico/wgs_cnv/DAKIKI.fixmate.bam > /media/kong/enrico/wgs_cnv/DAKIKI_fixmate.out 
samtools sort -@ 30 /media/kong/enrico/wgs_cnv/DAKIKI.fixmate.bam -o /media/kong/enrico/wgs_cnv/DAKIKI.fixmate.sorted.bam
samtools markdup -@ 30 -r -s /media/kong/enrico/wgs_cnv/DAKIKI.fixmate.sorted.bam /media/kong/enrico/wgs_cnv/DAKIKI.sorted.markup.bam 
samtools index -@ 30 /media/kong/enrico/wgs_cnv/DAKIKI.sorted.markup.bam

but nothing works, keep getting the same output.

Thank you in advance for any help!

[zeeev]

Hi @cccnrc, the wham suite of tools was designed for paired-end reads? Looking over the commands, above, I'm guessing you're using single-end reads?

Best,

Zev

[cccnrc]

Thanks for your reply @zeeev
The .bam is actually sequenced with paired-end....any other idea?

[carolynzy]

I had the same issue using whamg. I'm processing paired-end samples. whamg used to be working on this sample, the only change I made was using bbduk.sh to trim low quality bases from fastq file and then went through the same processes. When I used sickle to trim the same file, whamg just worked fine.

I looked into the .err file produced, and the content is as follows:

INFO: target bams:
../../bam_trimed_sorted/1_trimebbduk_sorted_addRG.bam
INFO: fasta file: ../../GCF_000195955.2_ASM19595v2_genomic.fna
INFO: OpenMP will roughly use 3 threads
INFO: gathering stats (may take some time) for bam: ../../bam_trimed_sorted/1_trimebbduk_sorted_addRG.bam
INFO: processed 0 reads for: ../../bam_trimed_sorted/1_trimebbduk_sorted_addRG.bam
INFO: processed 0 reads for: ../../bam_trimed_sorted/1_trimebbduk_sorted_addRG.bam
INFO: processed 0 reads for: ../../bam_trimed_sorted/1_trimebbduk_sorted_addRG.bam
INFO: processed 0 reads for: ../../bam_trimed_sorted/1_trimebbduk_sorted_addRG.bam
INFO: processed 0 reads for: ../../bam_trimed_sorted/1_trimebbduk_sorted_addRG.bam
INFO: processed 0 reads for: ../../bam_trimed_sorted/1_trimebbduk_sorted_addRG.bam
INFO: processed 0 reads for: ../../bam_trimed_sorted/1_trimebbduk_sorted_addRG.bam
INFO: processed 0 reads for: ../../bam_trimed_sorted/1_trimebbduk_sorted_addRG.bam
INFO: processed 0 reads for: ../../bam_trimed_sorted/1_trimebbduk_sorted_addRG.bam
INFO: processed 0 reads for: ../../bam_trimed_sorted/1_trimebbduk_sorted_addRG.bam

and then the program will hault here for ever. Any idea what's going on?

[SMMusgrave]

I'm having the exact same issue as cccnrc, running whamg on BWA-aligned paired-end .bam and getting stuck at

INFO: processed 0 reads for: SM.markdup.bqsr.bam

and not proceeding. Any ideas on how to resolve this?

whamg command:

whamg \
    -c primary_contigs.list \
    -x 4 \
    -a  Homo_sapiens_assembly38.fasta \
    -f $ID.markdup.bqsr.bam

[zeeev]

Please try limiting it only to the standard chromosomes chr1-chr22,X,Y.

[SMMusgrave]

Thank you for the suggestion. It appears that there was an issue with proving the regions to include as a list file. This was resolved when I instead provided the list in a variable like example 1 in the README:
$ export INCLUDE=chr1,chr2,chr3,chr4,chr5,chr6,chr7,chr8,chr9,chr10,chr11,chr12,chr13,chr14,chr15,chr16,chr17,chr18,chr19,chr20,chr21,chr22,chrX,chrY
$ whamg -c $INCLUDE -x 4 -a Homo_sapiens_assembly38.fasta -f $ID.markdup.bqsr.bam