[ERROR coverm::bam_generator] The STDERR for the minimap2 part

[WoCer2019]

Hi, I used the nice tool CoverM for calculating the coverage of MAG, but get an error. Please give me some advice, thanks!

Code: coverm genome --coupled Reads/BPA_1.fastq Reads/BPA_2.fastq Reads/DBP_1.fastq Reads/DBP_2.fastq Reads/DEP_1.fastq Reads/DEP_2.fastq Reads/DOP_1.fastq Reads/DOP_2.fastq Reads/GLU_1.fastq Reads/GLU_2.fastq Reads/NP_1.fastq Reads/NP_2.fastq Reads/S29.raw_1.fastq Reads/S29.raw_2.fastq Reads/S30.raw_1.fastq Reads/S30.raw_2.fastqReads/S31.raw_1.fastq Reads/S31.raw_2.fastq Reads/S32.raw_1.fastq Reads/S32.raw_2.fastq Reads/S33.raw_1.fastq Reads/S33.raw_2.fastq Reads/S34.raw_1.fastq Reads/S34.raw_2.fastq Reads/S35.raw_1.fastq Reads/S35.raw_2.fastq Reads/Seed_1.fastq Reads/Seed_2.fastq Reads/TC_1.fastq Reads/TC_2.fastq -d dereplicated/output/dereplicated_genomes -x fa -m rpkm -o MAG.abundance.csv -t 80

log file: nohup.out.txt

[wwood]

Hi,

At the bottom of the log it suggests a pair of files have different numbers of reads to each other, when it is expecting the same number in each. Do those files, and other pairs, appear truncated or corrupt at all?

-------------- Ben Woodcroft Group leader, Centre for Microbiome Research, QUT

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From: WoCer2019 @.> Sent: Tuesday, December 27, 2022 3:25:54 PM To: wwood/CoverM @.> Cc: Subscribed @.***> Subject: [wwood/CoverM] [ERROR coverm::bam_generator] The STDERR for the minimap2 part (Issue #148)

Hi, I used the nice tool CoverM for calculating the coverage of MAG, but get an error. Please give me some advice, thanks!

Code: coverm genome --coupled Reads/BPA_1.fastq Reads/BPA_2.fastq Reads/DBP_1.fastq Reads/DBP_2.fastq Reads/DEP_1.fastq Reads/DEP_2.fastq Reads/DOP_1.fastq Reads/DOP_2.fastq Reads/GLU_1.fastq Reads/GLU_2.fastq Reads/NP_1.fastq Reads/NP_2.fastq Reads/S29.raw_1.fastq Reads/S29.raw_2.fastq Reads/S30.raw_1.fastq Reads/S30.raw_2.fastqReads/S31.raw_1.fastq Reads/S31.raw_2.fastq Reads/S32.raw_1.fastq Reads/S32.raw_2.fastq Reads/S33.raw_1.fastq Reads/S33.raw_2.fastq Reads/S34.raw_1.fastq Reads/S34.raw_2.fastq Reads/S35.raw_1.fastq Reads/S35.raw_2.fastq Reads/Seed_1.fastq Reads/Seed_2.fastq Reads/TC_1.fastq Reads/TC_2.fastq -d dereplicated/output/dereplicated_genomes -x fa -m rpkm -o MAG.abundance.csv -t 80

log file: nohup.out.txthttps://github.com/wwood/CoverM/files/10306075/nohup.out.txt

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[WoCer2019]

Thanks for your reply! I used the same reads file for calculating the coverage of contig, it was successful. So, if the reads file has a problem, the result of calculating the coverage of contig will not be created. I feel it very strange. Why can calculate the coverage of contig and cannot calculate MAG?

Code (calculate contig): coverm contig --coupled Reads/BPA_1.fastq Reads/BPA_2.fastq Reads/DBP_1.fastq Reads/DBP_2.fastq Reads/DEP_1.fastq Reads/DEP_2.fastq Reads/DOP_1.fastq Reads/DOP_2.fastq Reads/GLU_1.fastq Reads/GLU_2.fastq Reads/NP_1.fastq Reads/NP_2.fastq Reads/S29.raw_1.fastq Reads/S29.raw_2.fastq Reads/S30.raw_1.fastq Reads/S30.raw_2.fastqReads/S31.raw_1.fastq Reads/S31.raw_2.fastq Reads/S32.raw_1.fastq Reads/S32.raw_2.fastq Reads/S33.raw_1.fastq Reads/S33.raw_2.fastq Reads/S34.raw_1.fastq Reads/S34.raw_2.fastq Reads/S35.raw_1.fastq Reads/S35.raw_2.fastq Reads/Seed_1.fastq Reads/Seed_2.fastq Reads/TC_1.fastq Reads/TC_2.fastq -r all.contig.fa --min-read-percent-identity 95 --min-read-aligned-percent 75 -m rpkm -o contig.abundance.csv -t 80

[wwood]

Hmm, good question. My feeling is that it might be an issue where contig mode should be failing, but isn't. Hard for me to tell until I get to a real computer, but in the meantime can you check the number of reads in each file please?

-------------- Ben Woodcroft Group leader, Centre for Microbiome Research, QUT

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From: WoCer2019 @.> Sent: Tuesday, December 27, 2022 6:33:47 PM To: wwood/CoverM @.> Cc: Ben J Woodcroft @.>; Comment @.> Subject: Re: [wwood/CoverM] [ERROR coverm::bam_generator] The STDERR for the minimap2 part (Issue #148)

Thanks for your reply! I used the same reads file for calculating the coverage of contig, it was successful. So, if the reads file has a problem, the result of calculating the coverage of contig will not be created. I feel it very strange. Why can calculate the coverage of contig and cannot calculate MAG?

Code (calculate contig): coverm contig --coupled Reads/BPA_1.fastq Reads/BPA_2.fastq Reads/DBP_1.fastq Reads/DBP_2.fastq Reads/DEP_1.fastq Reads/DEP_2.fastq Reads/DOP_1.fastq Reads/DOP_2.fastq Reads/GLU_1.fastq Reads/GLU_2.fastq Reads/NP_1.fastq Reads/NP_2.fastq Reads/S29.raw_1.fastq Reads/S29.raw_2.fastq Reads/S30.raw_1.fastq Reads/S30.raw_2.fastqReads/S31.raw_1.fastq Reads/S31.raw_2.fastq Reads/S32.raw_1.fastq Reads/S32.raw_2.fastq Reads/S33.raw_1.fastq Reads/S33.raw_2.fastq Reads/S34.raw_1.fastq Reads/S34.raw_2.fastq Reads/S35.raw_1.fastq Reads/S35.raw_2.fastq Reads/Seed_1.fastq Reads/Seed_2.fastq Reads/TC_1.fastq Reads/TC_2.fastq -r all.contig.fa --min-read-percent-identity 95 --min-read-aligned-percent 75 -m rpkm -o contig.abundance.csv -t 80

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[WoCer2019]

Ok. I check the number of reads. Meanwhile, I wish to get the subsequent reply too. Thanks!

[nvpatin]

Hi, was this issue ever resolved? I'm getting the same error trying to map interleaved reads to a contigs fasta. According to the log file, the BAM files are successfully generated and cached in the provided directory, but then the mapping calculations fail with the below error. Also, when I look in the bam cache directory, it only has two bam files (out of >60 expected).

[2023-04-28T20:38:44Z ERROR coverm::bam_generator] Failed to correctly find or parse BAM file at "/tmp/coverm_fifo.pWJk6n4hDNHa/foo.pipe": unable to open SAM/BAM/CRAM file at /tmp/coverm_fifo.pWJk6n4hDNHa/foo.pipe

And here is my full command:

coverm genome \ --reference eCruises_2018-2021_MAGs_HQ_dRepconcat.fa \ -s "" \ -m relative_abundance \ --interleaved *fq.gz \ --min-read-aligned-percent 0.75 \ --min-read-percent-identity 0.95 \ --min-covered-fraction 0 \ -x fa -t 10 -o Flyer2018_MAG_relabund.tsv \ --bam-file-cache-directory Flyer2018_MAG_bams &> log-relabund.txt

[wwood]

Hi @nvpatin that seems like a completely different issue. Maybe ran out of RAM?

Can you please check and if not can you post another issue with more details ie the whole output please?

For this issue need to know read numbers in each file, seems like corrupted input read files.

[nvpatin]

Thanks for the quick response. I think it's because I ran out of temporary storage space during the mapping process. I removed the --bam-file-cache-directory parameter and it worked fine, but the bam files were not saved so I guess I'll have to generate them anew if I want to run coverage calculations.

If I'm running this job in a loop on 50+ metagenomes, is there any way to specify that files in /tmp are deleted after each alignment and/or mapping job? I'm happy to open a new issue if you prefer but maybe it's an easy (or impossible) fix.

[wwood]

Hi,

I can't remember actually, what files are kept that you would like to see deleted?

Maybe the shortest path to a solution for you is to specify TMPDIR - see the FAQ. ben