Problem with Phylogenetic tree construction

Technical Support
1

Hello

I want to construct phylogenetic tree for bacterial sequence. I used a reference-based fragment insertion approach.

This is the command I ran

qiime fragment-insertion sepp --i-representative-sequences si_cs.qza --p-threads 40 --p-alignment-subset-size 1000 --p-placement-subset-size 5000 --o-tree Phylogeny[Rooted] --o-placements Placements

This was the error I got

p_error
p_error1831×378 213 KB

Removing /tmp/tmp.B5onA1CVkA/sepp-tmp-Wrjwf
Traceback (most recent call last):
File "/home/ruthra/miniconda2/envs/qiime2-2019.1/lib/python3.6/site-packages/q2cli/commands.py", line 274, in call
results = action(**arguments)
File "</home/ruthra/miniconda2/envs/qiime2-2019.1/lib/python3.6/site-packages/decorator.py:decorator-gen-290>", line 2, in sepp
File "/home/ruthra/miniconda2/envs/qiime2-2019.1/lib/python3.6/site-packages/qiime2/sdk/action.py", line 231, in bound_callable
output_types, provenance)
File "/home/ruthra/miniconda2/envs/qiime2-2019.1/lib/python3.6/site-packages/qiime2/sdk/action.py", line 365, in callable_executor
output_views = self._callable(**view_args)
File "/home/ruthra/miniconda2/envs/qiime2-2019.1/lib/python3.6/site-packages/q2_fragment_insertion/_insertion.py", line 179, in sepp
reference_alignment, reference_phylogeny, debug)
File "/home/ruthra/miniconda2/envs/qiime2-2019.1/lib/python3.6/site-packages/q2_fragment_insertion/_insertion.py", line 137, in _run
subprocess.run(cmd, check=True, cwd=cwd)
File "/home/ruthra/miniconda2/envs/qiime2-2019.1/lib/python3.6/subprocess.py", line 418, in run
output=stdout, stderr=stderr)
subprocess.CalledProcessError: Command '['run-sepp.sh', '/tmp/qiime2-archive-nmlejvgb/5addfdcf-d8e1-4d49-9f46-991dd9e237b8/data/dna-sequences.fasta', 'q2-fragment-insertion', '-x', '40', '-A', '1000', '-P', '5000']' returned non-zero exit status 1.

Plugin error from fragment-insertion:

Command '['run-sepp.sh', '/tmp/qiime2-archive-nmlejvgb/5addfdcf-d8e1-4d49-9f46-991dd9e237b8/data/dna-sequences.fasta', 'q2-fragment-insertion', '-x', '40', '-A', '1000', '-P', '5000']' returned non-zero exit status 1.

See above for debug info.

Could you please help me how to solve this error?

2

Hi @Asha1,

This might take a bit to troubleshoot, so please be patient with us! Could you please run the command,

locale

and share the result here?

Best,
Justine

3

Hi Ma’am,

Thank you so much for your prompt reply ma’am.

(qiime2-2019.1) ruthra@bioinformatics-PowerEdge-T630:~/miniconda2/envs/qiime2-2019.1/v3-v4_region_result/sp/mo/moraxellaca.taxaplot$ locale
LANG=C.UTF-8
LANGUAGE=
LC_CTYPE=“C.UTF-8”
LC_NUMERIC=“C.UTF-8”
LC_TIME=“C.UTF-8”
LC_COLLATE=“C.UTF-8”
LC_MONETARY=“C.UTF-8”
LC_MESSAGES=“C.UTF-8”
LC_PAPER=“C.UTF-8”
LC_NAME=“C.UTF-8”
LC_ADDRESS=“C.UTF-8”
LC_TELEPHONE=“C.UTF-8”
LC_MEASUREMENT=“C.UTF-8”
LC_IDENTIFICATION=“C.UTF-8”
LC_ALL=

Looking forward to your reply.

4

Hi @Asha1,

This is an easy fix, then!

Please run

export LC_ALL="en_US.utf-8"
export LANG="en_US.utf-8"
locale

You should see everything set to en_US.utf-8.

If you want to solve the problem forever, on a mac you can add this line to your ~/.bash_profile and on a linux server your ~/.bashrc, which is kind of a master file which tells the system how to configure itself self.

Best,
Justine

5

Thank you so much ma’am for your reply., I will check once and get back to you.

6

I ran the commanline as per your suggestion. But, I am getting the same error again.
(qiime2-2019.1) ruthra@bioinformatics-PowerEdge-T630:~/miniconda2/envs/qiime2-2019.1/v3-v4_region_result/sp/mo/moraxellaca.taxaplot$ export LC_ALL=“en_US.utf-8”
(qiime2-2019.1) ruthra@bioinformatics-PowerEdge-T630:~/miniconda2/envs/qiime2-2019.1/v3-v4_region_result/sp/mo/moraxellaca.taxaplot$ export LANG=“en_US.utf-8”
(qiime2-2019.1) ruthra@bioinformatics-PowerEdge-T630:~/miniconda2/envs/qiime2-2019.1/v3-v4_region_result/sp/mo/moraxellaca.taxaplot$ locale
LANG=en_US.utf-8
LANGUAGE=
LC_CTYPE=“en_US.utf-8”
LC_NUMERIC=“en_US.utf-8”
LC_TIME=“en_US.utf-8”
LC_COLLATE=“en_US.utf-8”
LC_MONETARY=“en_US.utf-8”
LC_MESSAGES=“en_US.utf-8”
LC_PAPER=“en_US.utf-8”
LC_NAME=“en_US.utf-8”
LC_ADDRESS=“en_US.utf-8”
LC_TELEPHONE=“en_US.utf-8”
LC_MEASUREMENT=“en_US.utf-8”
LC_IDENTIFICATION=“en_US.utf-8”
LC_ALL=en_US.utf-8
(qiime2-2019.1) ruthra@bioinformatics-PowerEdge-T630:~/miniconda2/envs/qiime2-2019.1/v3-v4_region_result/sp/mo/moraxellaca.taxaplot$ qiime fragment-insertion sepp --i-representative-sequences si_cs.qza --p-threads 40 --p-alignmen
t-subset-size 1000 --p-placement-subset-size 5000 --o-tree Phylogeny[Rooted] --o-placements Placements
Plugin error from fragment-insertion:

Command ‘[‘run-sepp.sh’, ‘/tmp/qiime2-archive-q49w7763/d5ce9526-e2a9-4b53-8b72-a37a2c11da39/data/dna-sequences.fasta’, ‘q2-fragment-insertion’, ‘-x’, ‘40’, ‘-A’, ‘1000’, ‘-P’, ‘5000’]’ returned non-zero exit status 1.

Debug info has been saved to /tmp/qiime2-q2cli-err-cw12znbe.log
What to do now Ma’am?

7

Can you please run the command with the --debug flag and post the results?

Best,
Justine

8

Hi @Asha1,

Based on your other post, is this ITS or 16s? The current fragment insertion doesn't support ITS.

Best,
Justine