lrtk ALIGN fails in bam_stats step

[Leelaa30]

I'm using LRTK to align stLFR reads to a reference genome using the following command:

lrtk ALIGN -FQ1 /scratch/project/adna/Papaya/Murdoch_sequencing/Papaya_stlfr/PAPHI001.R1.fq.gz -FQ2 /scratch/project/adna/Papaya/Murdoch_sequencing/Papaya_stlfr/PAPHI001.R2.fq.gz -R /scratch/project/adna/Papaya/Murdoch_sequencing/papaya_ref_grade/PAPHI003/hap2/PAPHI003_hap2.fasta -O /scratch/project/adna/leela/FB_SNP_calling_10_10_2025/aligned_reads/PAPHI001/PAPHI001.rg.bam -RG "@RG\tID:PAPHI001.E150016243\tSM:PAPHI001\tLB:PAPHI001.E150016243\tPU:E150016243\tPL:MGI\tPM:DNBseq_T7\tCN:MU_SABC" -T 10 -P stLFR 

However, the alignemnt fails with this error:

The output directory is: /scratch/project/adna/leela/FB_SNP_calling_10_10_2025/aligned_reads/PAPHI001
@RG\tID:PAPHI001.E150016243\tSM:PAPHI001\tLB:PAPHI001.E150016243\tPU:E150016243\tPL:MGI\tPM:DNBseq_T7\tCN:MU_SABC
[2026-03-04 15:32:12.339734] sort_barcoded_FQ1 starts
[2026-03-04 15:32:45.481001] sort_barcoded_FQ1 ends
[2026-03-04 15:32:45.481069] sort_unbarcoded_FQ1 starts
[2026-03-04 15:33:42.542408] sort_unbarcoded_FQ1 ends
[2026-03-04 15:33:42.542474] sort_barcoded_FQ2 starts
[2026-03-04 15:34:15.231935] sort_barcoded_FQ2 ends
[2026-03-04 15:34:15.232026] sort_unbarcoded_FQ2 starts
[2026-03-04 15:35:12.082798] sort_unbarcoded_FQ2 ends
[2026-03-04 15:35:12.086547] EMA_alignment starts
BWA initialization...
[M::bwa_idx_load_from_disk] read 0 ALT contigs
Processing reads...
[2026-03-04 15:35:12.554866] EMA_alignment ends
[2026-03-04 15:35:12.554966] BWA_alignment starts
[M::bwa_idx_load_from_disk] read 0 ALT contigs
[main] Version: 0.7.18-r1243-dirty
[main] CMD: bwa mem -1 -R @RG\tID:PAPHI001.E150016243\tSM:PAPHI001\tLB:PAPHI001.E150016243\tPU:E150016243\tPL:MGI\tPM:DNBseq_T7\tCN:MU_SABC -t 8 /scratch/project/adna/Papaya/Murdoch_sequencing/papaya_ref_gr
ade/PAPHI003/hap2/PAPHI003_hap2.fasta /scratch/project/adna/leela/FB_SNP_calling_10_10_2025/aligned_reads/PAPHI001/tmp.unbarcoded.1.fq /scratch/project/adna/leela/FB_SNP_calling_10_10_2025/aligned_reads/PAP
HI001/tmp.unbarcoded.2.fq
[main] Real time: 0.098 sec; CPU: 0.099 sec
[2026-03-04 15:35:12.659517] BWA_alignment ends
[2026-03-04 15:35:12.659581] merge_bam starts
[2026-03-04 15:35:12.667025] merge_bam ends
[2026-03-04 15:35:12.667086] sort_bam starts
[2026-03-04 15:35:12.675469] sort_bam ends
[2026-03-04 15:35:12.675539] Mark_bam starts
[2026-03-04 15:35:15.320830] Mark_bam ends
[2026-03-04 15:35:15.320900] index_bam starts
[2026-03-04 15:35:15.328093] index_bam ends
[2026-03-04 15:35:15.328277] bam_stat starts
Illegal division by zero at /home/leelamanoharan/miniforge3/envs/genomics/lib/python3.9/site-packages/script/AlignStat_WGS.pl line 84.
[2026-03-04 15:35:15.366629] bam_stat fails: perl /home/leelamanoharan/miniforge3/envs/genomics/lib/python3.9/site-packages/script/AlignStat_WGS.pl -in /scratch/project/adna/leela/FB_SNP_calling_10_10_2025/
aligned_reads/PAPHI001/PAPHI001.rg.bam -sam PAPHI001.rg.bam -samtools /home/leelamanoharan/miniforge3/envs/genomics/bin/samtools -outdir /scratch/project/adna/leela/FB_SNP_calling_10_10_2025/aligned_reads/P
APHI001 -move /scratch/project/adna/leela/FB_SNP_calling_10_10_2025/aligned_reads/PAPHI001

Also, note that the documentation didn't say anything about indexing the genome with bwa, so I had to figure it out myself.

I would appreciate any help or suggestions.

Thanks, Leela