No paired-end reads were detected in paired-end read library

[chengmingbo]

Description of the bug

Dear authors,
Thanks very much for developing this amazing chip-seq pipeline. I'm not sure if it's a bug, but to my understanding, the samplesheets.csv I compiled would automatically being detected as single-ended. However, the featureCounts was added to -p parameter that cause an error.

chipseq nextflow version

Nextflow 24.10.5 is available - Please consider updating your version to it
N E X T F L O W  ~  version 23.10.1
Launching `https://github.com/nf-core/chipseq` [compassionate_golick] DSL2 - revision: 76e2382b6d [master]
nf-core/chipseq v2.1.0-g76e2382

the samplesheets

sample,fastq_1,fastq_2,replicate,antibody,control,control_replicate
Bach2_TGFB_plus1,ds213_S1_R1_001.fastq.gz,,1,Bach2,Input_TGFB_plus1,1
Bach2_TGFB_plus2,ds215_S3_R1_001.fastq.gz,,1,Bach2,Input_TGFB_plus2,1
Bach2_TGFB_plus_old2,ds217_S5_R1_001.fastq.gz,,1,Bach2,Input_TGFB_plus_old2,1
Bach2_TGFB_minus1,ds219_S7_R1_001.fastq.gz,,1,Bach2,Input_TGFB_minus1,1
Bach2_TGFB_minus2,ds221_S9_R1_001.fastq.gz,,1,Bach2,Input_TGFB_minus2,1
Bach2_TGFB_plus25_10,ds222_2_S10_R1_001.fastq.gz,,1,Bach2,Input_TGFB_plus25_10,1
Input_TGFB_plus1,ds214_2_S2_R1_001.fastq.gz,,1,,,
Input_TGFB_plus2,ds216_S4_R1_001.fastq.gz,,1,,,
Input_TGFB_plus_old2,ds218_S6_R1_001.fastq.gz,,1,,,
Input_TGFB_minus1,ds220_2_S8_R1_001.fastq.gz,,1,,,
Input_TGFB_minus2,ds222_2_S10_R1_001.fastq.gz,,1,,,
Input_TGFB_plus25_10,ds226_2_S12_R1_001.fastq.gz,,1,,,

Command used and terminal output

`nextflow run nf-core/chipseq --input ./samplesheet.csv --outdir ./results --genome GRCh38 --macs_gsize 2913022398 --igenomes_base $hg38  --skip_preseq -resume`


### Error log

`
ERROR ~ Error executing process > 'NFCORE_CHIPSEQ:CHIPSEQ:BED_CONSENSUS_QUANTIFY_QC_BEDTOOLS_FEATURECOUNTS_DESEQ2:SUBREAD_FEATURECOUNTS (Bach2)'

Caused by:
  Process `NFCORE_CHIPSEQ:CHIPSEQ:BED_CONSENSUS_QUANTIFY_QC_BEDTOOLS_FEATURECOUNTS_DESEQ2:SUBREAD_FEATURECOUNTS (Bach2)` terminated with an error exit status (255)

Command executed:

  featureCounts \
      -F SAF -O --fracOverlap 0.2 \
      -p \
      -T 6 \
      -a Bach2.consensus_peaks.saf \
      -s 0 \
      -o Bach2.consensus_peaks.featureCounts.txt \
      Bach2_TGFB_plus25_10_REP1.mLb.clN.sorted.bam Bach2_TGFB_minus2_REP1.mLb.clN.sorted.bam Bach2_TGFB_plus2_REP1.mLb.clN.sorted.bam Bach2_TGFB_plus_old2_REP1.mLb.clN.sorted.bam Bach2_TGFB_plus1_REP1.mLb.clN.sorted.bam Bach2_TGFB_minus1_REP1.mLb.clN.sorted.bam

  cat <<-END_VERSIONS > versions.yml
  "NFCORE_CHIPSEQ:CHIPSEQ:BED_CONSENSUS_QUANTIFY_QC_BEDTOOLS_FEATURECOUNTS_DESEQ2:SUBREAD_FEATURECOUNTS":
      subread: $( echo $(featureCounts -v 2>&1) | sed -e "s/featureCounts v//g")
  END_VERSIONS
Command exit status:
  255

Command output:
  (empty)

Command error:

          ==========     _____ _    _ ____  _____  ______          _____
          =====         / ____| |  | |  _ \|  __ \|  ____|   /\   |  __ \
            =====      | (___ | |  | | |_) | |__) | |__     /  \  | |  | |
              ====      \___ \| |  | |  _ <|  _  /|  __|   / /\ \ | |  | |
                ====    ____) | |__| | |_) | | \ \| |____ / ____ \| |__| |
          ==========   |_____/ \____/|____/|_|  \_\______/_/    \_\_____/
  	  v2.0.2

  //========================== featureCounts setting ===========================\\
  ||                                                                            ||
  ||             Input files : 6 BAM files                                      ||
  ||                                                                            ||
  ||                           Bach2_TGFB_plus25_10_REP1.mLb.clN.sorted.bam     ||
  ||                           Bach2_TGFB_minus2_REP1.mLb.clN.sorted.bam        ||
  ||                           Bach2_TGFB_plus2_REP1.mLb.clN.sorted.bam         ||
  ||                           Bach2_TGFB_plus_old2_REP1.mLb.clN.sorted.bam     ||
  ||                           Bach2_TGFB_plus1_REP1.mLb.clN.sorted.bam         ||
  ||                           Bach2_TGFB_minus1_REP1.mLb.clN.sorted.bam        ||
  ||                                                                            ||
  ||             Output file : Bach2.consensus_peaks.featureCounts.txt          ||
  ||                 Summary : Bach2.consensus_peaks.featureCounts.txt.summary  ||
  ||              Paired-end : yes                                              ||
  ||        Count read pairs : no                                               ||
  ||              Annotation : Bach2.consensus_peaks.saf (SAF)                  ||
  ||      Dir for temp files : ./                                               ||
  ||                                                                            ||
  ||                 Threads : 6                                                ||
  ||                   Level : meta-feature level                               ||
  ||      Multimapping reads : not counted                                      ||
  || Multi-overlapping reads : counted                                          ||
  ||   Min overlapping bases : 1                                                ||
  ||   Min overlapping frac. : 20.0% to reads                                   ||
  ||                                                                            ||
  \\============================================================================//

  //================================= Running ==================================\\
  ||                                                                            ||
  || Load annotation file Bach2.consensus_peaks.saf ...                         ||
  ||    Features : 15216                                                        ||
  ||    Meta-features : 15216                                                   ||
  ||    Chromosomes/contigs : 46                                                ||
  ||                                                                            ||
  || Process BAM file Bach2_TGFB_plus25_10_REP1.mLb.clN.sorted.bam...           ||
  ERROR: No paired-end reads were detected in paired-end read library : Bach2_TGFB_plus25_10_REP1.mLb.clN.sorted.bam

Relevant files

nextflow_2025-03-04_11:52:06.883245.txt

System information

Nextflow version 23.10.1
Hardware HPC
executor slurm
OS: Rocky Linux release 8.10
Version of nf-core/chipseq v2.1.0-g76e2382

[JoseEspinosa]

Thanks for reporting, that is indeed strange as we use the meta information obtained from the samplesheet to set the parameter, see here. I will try to see whether there is an issue when reading your samplesheet although it LGTM...

[JoseEspinosa]

This has been fixed in #453 and is now on dev, it would be nice if you could give it a try @chengmingbo