Process `NFCORE_CHIPSEQ:CHIPSEQ:FASTQ_ALIGN_BWA:BAM_SORT_STATS_SAMTOOLS:SAMTOOLS_SORT (SRR5627136_REP2_T1)` terminated with an error exit status (1)

[NicoleChant]

Description of the bug

Bug Description

Hello, I am encountering a bug during SAMTOOLS SORT when I run nf-core/chipseq pipeline from master branch on Red Hat Linux.

The pipeline runs perfectly up to the point of SAMTOOLS-SORT.

All programs are installed properly. The alignment step (using bwa mem) finishes properly without any errors.

In the next step, the pipeline is not able to locate the alignment files of the previous step.

All programs required are properly installed (bwa mem, samtools, etc.)

Any help would be DEEPLY appreciated.

Command used and terminal output

I use a custom environment and I initialize the pipeline as follows:

nextflow run nf-core/chipseq --input chip_seq_input_new_with_second.csv --outdir chip_seq_experiment2 --fasta <my_fasta> --save_reference --read_length 50 --gtf  <my_gtf>

Bug Error Messages from the stacktrace

The pipeline was proceeding as expected up to SAMTOOLS SORT operation:

[f3/eaad2d] NFC…0_genomic_replaced.gtf.gz) | 1 of 1 ✔
[b2/86dc81] NFC…v2.0_genomic_replaced.gtf) | 1 of 1 ✔
[f5/c0cff6] NFC…TCHROMSIZES (chm13v2.0.fa) | 1 of 1 ✔
[ef/c6092f] NFC…GIONS (chm13v2.0.fa.sizes) | 1 of 1 ✔
[07/4c9520] NFC…E:BWA_INDEX (chm13v2.0.fa) | 1 of 1 ✔
[b7/30bf98] NFC…input_new_with_second.csv) | 1 of 1 ✔
[b4/5e88cc] NFC…ASTQC (SRR5627150_REP2_T1) | 12 of 12 ✔
[2b/be1a95] NFC…ALORE (SRR5627151_REP1_T1) | 12 of 12 ✔
[9f/74b0cf] NFC…A_MEM (SRR5627151_REP1_T1) | 12 of 12 ✔
[48/9acab1] NFC…_SORT (SRR5627135_REP1_T1) | 4 of 12, failed: 4
[-        ] NFC…TS_SAMTOOLS:SAMTOOLS_INDEX -
[-        ] NFC…TS_SAMTOOLS:SAMTOOLS_STATS -
[-        ] NFC…SAMTOOLS:SAMTOOLS_FLAGSTAT -
[-        ] NFC…SAMTOOLS:SAMTOOLS_IDXSTATS -
[-        ] NFC…IPSEQ:PICARD_MERGESAMFILES -
[-        ] NFC…CARD:PICARD_MARKDUPLICATES -
[-        ] NFC…ATES_PICARD:SAMTOOLS_INDEX -
[-        ] NFC…TS_SAMTOOLS:SAMTOOLS_STATS -
[-        ] NFC…SAMTOOLS:SAMTOOLS_FLAGSTAT -

This is the error I encountered:

Caused by:
  Process `NFCORE_CHIPSEQ:CHIPSEQ:FASTQ_ALIGN_BWA:BAM_SORT_STATS_SAMTOOLS:SAMTOOLS_SORT (SRR5627136_REP2_T1)` terminated with an error exit status (1)

Command executed:

  samtools cat \
      --threads 6 \
      SRR5627136_REP2_T1.Lb.bam \
  | \
  samtools sort \
       \
      -T SRR5627136_REP2_T1.Lb.sorted \
      --threads 6 \
      --reference chm13v2.0.fa \
      -o SRR5627136_REP2_T1.Lb.sorted.bam \
      -
  
  cat <<-END_VERSIONS > versions.yml
  "NFCORE_CHIPSEQ:CHIPSEQ:FASTQ_ALIGN_BWA:BAM_SORT_STATS_SAMTOOLS:SAMTOOLS_SORT":
      samtools: $(echo $(samtools --version 2>&1) | sed 's/^.*samtools //; s/Using.*$//')
  END_VERSIONS

Command exit status:
  1

Command output:
  (empty)

Command error:
  Usage: samtools cat [options] <in1.bam>  [... <inN.bam>]
         samtools cat [options] <in1.cram> [... <inN.cram>]

Relevant files

This was my input file:

sample,fastq_1,fastq_2,replicate,antibody,control,control_replicate
SRR5627135,SRR5627135_1.fastq.gz,SRR5627135_2.fastq.gz,1,H3K4me3,SRR5627137,1
SRR5627135,SRR5627135_1.fastq.gz,SRR5627135_2.fastq.gz,2,H3K4me3,SRR5627142,1
SRR5627136,SRR5627136_1.fastq.gz,SRR5627136_2.fastq.gz,1,H3K4me3,SRR5627137,1
SRR5627136,SRR5627136_1.fastq.gz,SRR5627136_2.fastq.gz,2,H3K4me3,SRR5627142,1
SRR5627148,SRR5627148_1.fastq.gz,SRR5627148_2.fastq.gz,1,H3K36me3,SRR5627137,1
SRR5627148,SRR5627148_1.fastq.gz,SRR5627148_2.fastq.gz,2,H3K36me3,SRR5627142,1
SRR5627150,SRR5627150_1.fastq.gz,SRR5627150_2.fastq.gz,1,H3K4me3,SRR5627137,1
SRR5627150,SRR5627150_1.fastq.gz,SRR5627150_2.fastq.gz,2,H3K4me3,SRR5627142,1
SRR5627151,SRR5627151_1.fastq.gz,SRR5627151_2.fastq.gz,1,H3K4me3,SRR5627137,1
SRR5627151,SRR5627151_1.fastq.gz,SRR5627151_2.fastq.gz,2,H3K4me3,SRR5627142,1
SRR5627137,SRR5627137_1.fastq.gz,SRR5627137_2.fastq.gz,1,,,
SRR5627142,SRR5627142_1.fastq.gz,SRR5627142_2.fastq.gz,1,,,

System information

samtools 1.21
Operating System: Red Hat Enterprise Linux 8.10 (Ootpa)
Architecture: x86-64

Thank you,
Nikol

[user-mb]

the samtools used for the pipeline is version 1.20
it should idealy be loaded by the pipeline, however on my system to local installed version is used (which is 1.22.1 at the point of wrinting).
option handles --threads and -@ do not exist for samtools cat, see: samtools/samtools#2162

i resolved this pipeline bug by downgrading samtools to version 1.20

hope this helps anyone that encounters this issue.