Help with error 512

[mradz19]

Phyloseq worked with the test data. But loading in my sample (approx 50 million reads) phyloseq fails with error 512:

[17:29:56] running subcommand: /mnt/nfs/home/.conda/envs/phyloflash/lib/phyloFlash/barrnap-HGV/bin/barrnap_HGV --evalue 1e-100 --reject 0.6 --kingdom bac --gene ssu --threads 22 p100.spades/scaffolds.fasta >p100.scaffolds.bac.gff 2>p100.barrnap.out [17:29:56] FATAL: Tool execution failed!. Error was 'No such file or directory' and return code '512' Aborting. [17:29:56] Saving log to file phyloFlash_log_on_error

I checked the bin directory and barnap_HGV is definitely present.

[kbseah]

Hello, Could you please attach the file p100.barrnap.out here, as well as the phyloFlash command line that you used? What type of sequencing library do you have? (Illumina, paired or single, sample type) Thanks, Brandon

[mradz19]

HI kbeash, This is the command I used:

phyloFlash.pl -lib 152_test -CPUs 22 -read1 p152.fastq.gz -interleaved

This is the content of that file.

[19:14:07] This is barrnap_HGV 0.7 [19:14:07] Written by Torsten Seemann torsten.seemann@gmail.com [19:14:07] Obtained from https://github.com/Victorian-Bioinformatics-Consortium/barrnap [19:14:07] Detected operating system: linux [19:14:07] Using HMMER binary: /mnt/nfs/home/30041036/.conda/envs/phyloflash/lib/phyloFlash/barrnap-HGV/bin/../binaries/linux/nhmmer [19:14:07] Will use 22 threads [19:14:07] Setting evalue cutoff to 1e-100 [19:14:07] Will tag genes < 0.8 of expected length. [19:14:07] Will reject genes < 0.6 of expected length. [19:14:07] Using database: /mnt/nfs/home/.conda/envs/phyloflash/lib/phyloFlash/barrnap-HGV/bin/../db/ssu/bac.hmm [19:14:07] Usage: barrnap_HGV

The reads are paired end illumina shotgun reads isolated from infected human tissue, they have been merged with bbmerge and set as interleaved.

[kbseah]

Hello,

Hm it looks like the log snippet that you originally posted was from a run named p100 while the command in the second post is for a run called 152_test. Do both runs give an identical error? Could you please post the entire phyloFlash log file here, and not just a snippet? You can use the -keeptmp option of phyloFlash to retain all the temporary and log files for troubleshooting.

Thanks again, Brandon

[HRGV]

Hi Michael Would you mind trying to input your raw reads as read1 and read2 instead of the reads merged with bbmerge? We have never tested pre-phyloFLash read merging and this could lead to fails in the read processing and ultimately an almost useless assembly Cheers Harald

[kbseah]

@HRGV i think we can close this issue now