q2-woltka ( returned non-zero exit status 1 )

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1

Hello!

I am attempting to use WOLTKA on my dataset. I have 32 SAM files I’ve retrieved from bowtie. I proceeded to use the following:

woltka classify -i 01_SAMPLES -o table.biom

qiime tools import --type 'FeatureTable[Frequency]' --input-path table.biom --output-path table.qza

qiime diversity core-metrics-phylogenetic --i-phylogeny tree.qza --i-table table.qza --p-sampling-depth 10000 --m-metadata-file mappingfile.txt --output-dir output

I used the tree.qza provided from WOL. I receive this error:

Plugin error from diversity:

Command '['faithpd', '-i', '/var/folders/tm/yf1fs3ys08bb7v_rfjyt8rz80000gr/T/qiime2/user/data/601fc3f6-dae5-4eb4-960c-27f853e599db/data/feature-table.biom', '-t', '/var/folders/tm/yf1fs3ys08bb7v_rfjyt8rz80000gr/T/qiime2/user/data/901fdadf-7b9b-4532-85f4-5386e1b4dd6f/data/tree.nwk', '-o', '/var/folders/tm/yf1fs3ys08bb7v_rfjyt8rz80000gr/T/qiime2/user/processes/4357-1758564090.049032@fa74451/tmp/q2-OutPath-vytdbchm']' returned non-zero exit status 1.

Debug info has been saved to /var/folders/tm/yf1fs3ys08bb7v_rfjyt8rz80000gr/T/qiime2-q2cli-err-aqfnghze.log

I’m not sure what this error may mean. In addition, I’ve tried to directly import my samples into Q2 using qiime tools import --type FeatureData\[SeqAlnMap\] --input-path alignment.sam --output-path alignment.qza, but I have 32 SAM files (sam/sample). How would I import all SAM files into one alignment.qza file? Thanks!

2

Hello @zahra , I am the main developer of Woltka and WoL. Let me try to address your question.

The error message most likely means the input alignment file and the tree file don’t match. It might be because you used a reference database other than WoL (and the right version) for Bowtie2 alignment. Can you let us know where did you get the database and the tree.qza file?

To import 32 SAM files (one file per sample) into a single alignment.qza file, you will need to “multiplex” them, then concatenate them into a single SAM file. Here is the relevant explanation in the Woltka documentation.

Typically, sequencing data generated at a sequencing facility is initially in multiplexed format, and they will demultiplex it before shipping to you. Now you will need to re-multiplex them back manually.

For example, a demultiplexed SAM file for sample X01 reads like:

S0R16	73	G000147875	2025907	255	150M	=	2025907	0	*	*
S0R17	99	G001792925	279403	255	150M	=	279501	248	*	*
S0R17	147	G001792925	279501	255	150M	=	279403	-248	*	*
S0R18	83	G001792925	377844	255	150M	=	377760	-234	*	*
S0R18	163	G001792925	377760	255	150M	=	377844	234	*	*
...

You will need to add a prefix X01_ before each line, such that query IDs become like X01_S0R16, then concatenate all SAM files into a single file. Woltka doesn’t have this feature, but it can be achieved with some Linux commands, like the following:

for file in /path/to/*.sam.gz; do
  id=$(basename ${file%.sam.gz})
  zcat $file | sed 's/^/'${id}'_/' | gzip >> multiplexed.sam.gz
done

Hope this helps!

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