Plugin error from phylogeny，

Technical Support
1

Hi! I got a error when I was running the qiime2 command on my lab's server:

nohup qiime phylogeny align-to-tree-mafft-fasttree \
  --i-sequences rep-seqs.qza \
  --o-alignment aligned-rep-seqs.qza \
  --o-masked-alignment masked-aligned-rep-seqs.qza \
  --o-tree unrooted-tree.qza \
  --o-rooted-tree rooted-tree.qza

Here are the error:

Plugin error from phylogeny:

  Command '['mafft', '--preservecase', '--inputorder', '--thread', '1', '/tmp/qiime2-archive-jxdayz10/8e7f3709-fdd2-4c07-9b70-0d6060c5143b/data/dna-sequences.fasta']' returned non-zero exit status 1.

Debug info has been saved to /tmp/qiime2-q2cli-err-yk52dmc3.log

And the .log file is:

inputfile = orig
321112 x 424 - 220 d
nthread = 1
stacksize: 8192 kb->62717 kb
generating a scoring matrix for nucleotide (dist=200) ... done
Gap Penalty = -1.53, +0.00, +0.00

Making a distance matrix ..

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/bmp/exp/yuyue/miniconda2/envs/qiime2-2020.2/bin/mafft: line 2440: 55172 Killed                  "$prefix/disttbfast" -q $npickup -E $cycledisttbfast -V "-"$gopdist -s $unalignlevel $legacygapopt $mergearg -W $tuplesize $termgapopt $outnum $addarg $add2ndhalfarg -C $numthreadstb $memopt $weightopt $treeinopt $treeoutopt $distoutopt $seqtype $model -f "-"$gop -Q $spfactor -h $aof $param_fft $algopt $treealg $scoreoutarg < infile > pre 2>> "$progressfile"

Running external command line application. This may print messages to stdout and/or stderr.
The command being run is below. This command cannot be manually re-run as it will depend on temporary files that no longer exist.

Command: mafft --preservecase --inputorder --thread 1 /bmp/exp/yuyue/tmp/qiime2-archive-d6cqbuxx/8e7f3709-fdd2-4c07-9b70-0d6060c5143b/data/dna-sequences.fasta

Traceback (most recent call last):
  File "/bmp/exp/yuyue/miniconda2/envs/qiime2-2020.2/lib/python3.6/site-packages/q2cli/commands.py", line 328, in __call__
    results = action(**arguments)
  File "</bmp/exp/yuyue/miniconda2/envs/qiime2-2020.2/lib/python3.6/site-packages/decorator.py:decorator-gen-226>", line 2, in align_to_tree_mafft_fasttree
  File "/bmp/exp/yuyue/miniconda2/envs/qiime2-2020.2/lib/python3.6/site-packages/qiime2/sdk/action.py", line 245, in bound_callable
    output_types, provenance)
  File "/bmp/exp/yuyue/miniconda2/envs/qiime2-2020.2/lib/python3.6/site-packages/qiime2/sdk/action.py", line 484, in _callable_executor_
    outputs = self._callable(scope.ctx, **view_args)
  File "/bmp/exp/yuyue/miniconda2/envs/qiime2-2020.2/lib/python3.6/site-packages/q2_phylogeny/_align_to_tree_mafft_fasttree.py", line 20, in align_to_tree_mafft_fasttree
    parttree=parttree)
  File "</bmp/exp/yuyue/miniconda2/envs/qiime2-2020.2/lib/python3.6/site-packages/decorator.py:decorator-gen-477>", line 2, in mafft
  File "/bmp/exp/yuyue/miniconda2/envs/qiime2-2020.2/lib/python3.6/site-packages/qiime2/sdk/action.py", line 245, in bound_callable
    output_types, provenance)
  File "/bmp/exp/yuyue/miniconda2/envs/qiime2-2020.2/lib/python3.6/site-packages/qiime2/sdk/action.py", line 390, in _callable_executor_
    output_views = self._callable(**view_args)
  File "/bmp/exp/yuyue/miniconda2/envs/qiime2-2020.2/lib/python3.6/site-packages/q2_alignment/_mafft.py", line 85, in mafft
    run_command(cmd, aligned_fp)
  File "/bmp/exp/yuyue/miniconda2/envs/qiime2-2020.2/lib/python3.6/site-packages/q2_alignment/_mafft.py", line 27, in run_command
    subprocess.run(cmd, stdout=output_f, check=True)
  File "/bmp/exp/yuyue/miniconda2/envs/qiime2-2020.2/lib/python3.6/subprocess.py", line 418, in run
    output=stdout, stderr=stderr)
subprocess.CalledProcessError: Command '['mafft', '--preservecase', '--inputorder', '--thread', '1', '/bmp/exp/yuyue/tmp/qiime2-archive-d6cqbuxx/8e7f3709-fdd2-4c07-9b70-0d6060c5143b/data/dna-sequences.fasta']' returned non-zero exit status 1.

I'm confused about how to deal with this problem, is the only one way to solve this problem by change a server or personnel computer?

BY THE WAY, I had change the ASV names which were create by DADA2, and the command is following:

less dna-sequences.fasta |paste - -|sed '1i ASVID,seq' > rep.fa
**and another command run by R:**
library(pacman)
pacman::p_load(tidyverse,magrittr,stringr)
rep <- "rep.fa" %>%
read.delim(check.names = FALSE, row.names = 1) %>%
set_rownames(paste0(">ASV", seq_len(nrow(.))))
write.table (rep,file ="rep.xls", sep ="\t", row.names = T,quote = F)
less rep.xls|sed '1d'|sed 's/"//g'|\
sed 's/\r//g'|tr "\t" "\n" > rep-seqs.fasta

WERE THOSE COMMANDS INFLUENCE THE RESULT?
THANKS FOR YOUR PATIENCE!!!!! :smiling_face_with_three_hearts:

2

Hello @Xieguangkun,

I'm not sure what caused this error. :thinking:

How much memory does your computer have and how big is your rep-seqs.qza file?

Colin :bird:

P.S. I reformatted your code so it was easier to read. Check out how to make code blocks!

```bash
nohup qiime phylogeny align-to-tree-mafft-fasttree \
  --i-sequences rep-seqs.qza \
  ...
  ...
3

My server memory have 512G, and the rep-seqs.qza is 19.46MB.THANKS FOR YOUR ANSWER!

4

That final code block appears to be from this page, and it should not cause issues with mafft. 微生物多样性qiime2分析流程(2）qiime2和dada2处理16s序列 - 哔哩哔哩

How long was this qiime2 plugin running? Was it running on a SLURM cluster or some sort of supercomputer, or running directly on the command line?

I ask because of this line mafft: line 2440: 55172 Killed
When a unix process is 'killed' it just means it was canceled by the system, and I'm trying to figure how why the system would cancel this job.

5

It appears you have about 321,112 ASVs (that is a lot!) with an alignment length of 424 positions. Generally, that should not be an issue for mafft. Perhaps you can privately share the sequence QZA file?

Also, does pipeline run with the normal ASV names? If you are able to get the pipeline run, then that'd imply that the modifications you applied generate invalid headers for mafft.

-Mike

1 Like
6

rep-seqs.qza
THANKS A LOT!

And I changed the ASV names before the command
qiime phylogeny align-to-tree-mafft-fasttree.

Running this command on my lab's Rack-mounted Server.

7

Are you still having issues running this plugin?

If you don't change the ASV names, does that fix the issue?

8

I tried to run this plugin without changed the ASV names, it showed the same question.

10

I use this plugin with other data, it finish the work. BUT when I used this plugin with the rep-seqs.qza which I upload, the error was happend again.

11

Are you using a SLURM queue or torque queue to run this command on your server? Can you post the full job submission script, as that could give us clues!

12

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未命名16466619631163×845 36.6 KB

13

That looks like three accounts (wangyue, wangrui..., guohao) are running directly on the server.

Sometimes processes / jobs can be killed because they use too much memory. Next time you run the plugin, try opening up top like you did here and see if your Qiime command using a lot of the %MEM (memory) column, or if something else is using up all the memory on the server.

14

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