Error: returned non-zero exit status 2

[GriffyGe]

Hi @fasincar
I am trying to run phylophlan for the first time. I would like to get phylogenetic tree for Xanthomonas (.fna). I am using following 2 commands:
phylophlan_write_config_file -d a -o griffy_config.cfg --db_aa diamond --map_dna blastn --msa muscle --tree1 fasttree --verbose 2>&1 | tee phylophlan_write_config_file.log
phylophlan -i input_genomes --force_nucleotides -d phylophlan -t a -f griffy_config.cfg --diversity low -o output_Xanthomonas --verbose 2>&1 | tee logs/phylophlan.log
And I got the following error massage:

Mapping "phylophlan" on 38 inputs (key: "map_dna")
Mapping "output_Xanthomonas/tmp/clean_dna/GCA_002019225.fna"
Removing "output_Xanthomonas/tmp/map_dna/GCA_002019225.b6o.bkp"

[e] Command '['/home/griffy/miniconda3/envs/Griffy_env/bin/blastn', '-outfmt', '6', '-evalue', '0.1', '-max_target_seqs', '1000000', '-perc_identity', '75', '-query', 'output_Xanthomonas/tmp/clean_dna/GCA_002019225.fna', '-db', 'phylophlan_databases/phylophlan/phylophlan.dmnd', '-out', 'output_Xanthomonas/tmp/map_dna/GCA_002019225.b6o.bkp']'
returned non-zero exit status 2.

[e] cannot execute command
    command_line: /home/griffy/miniconda3/envs/Griffy_env/bin/blastn -outfmt 6 -evalue 0.1 -max_target_seqs 1000000 -perc_identity 75 -query output_Xanthomonas/tmp/clean_dna/GCA_002019225.fna -db phylophlan_databases/phylophlan/phylophlan.dmnd -out output_Xanthomonas/tmp/map_dna/GCA_002019225.b6o.bkp
           stdin: None
          stdout: None...
[e] Command '['/home/griffy/miniconda3/envs/Griffy_env/bin/blastn', '-outfmt', '6', '-evalue', '0.1', '-max_target_seqs', '1000000', '-perc_identity', '75', '-query', 'output_Xanthomonas/tmp/clean_dna/GCA_002019225.fna', '-db', 'phylophlan_databases/phylophlan/phylophlan.dmnd', '-out', 'output_Xanthomonas/tmp/map_dna/GCA_002019225.b6o.bkp']' returned non-zero exit status 2.

[e] **error while mapping**
    {'program_name': '/home/griffy/miniconda3/envs/Griffy_env/bin/blastn', 'params': '-outfmt 6 -evalue 0.1 -max_target_seqs 1000000 -perc_identity 75', 'input': '-query', 'database': '-db', 'output': '-out', 'version': '-version', 'command_line': '#program_name# #params# #input# #database# #output#'}
    output_Xanthomonas/tmp/clean_dna/GCA_002019225.fna
    phylophlan_databases/phylophlan/phylophlan.dmnd
    output_Xanthomonas/tmp/map_dna
    GCA_002019225.b6o.bkp
    True

[e] Command '['/home/griffy/miniconda3/envs/Griffy_env/bin/blastn', '-outfmt', '6', '-evalue', '0.1', '-max_target_seqs', '1000000', '-perc_identity', '75', '-query', 'output_Xanthomonas/tmp/clean_dna/GCA_002019225.fna', '-db', 'phylophlan_databases/phylophlan/phylophlan.dmnd', '-out', 'output_Xanthomonas/tmp/map_dna/GCA_002019225.b6o.bkp']' returned non-zero exit status 2.

[e] gene_markers_identification crashed

How can I fix this problem?
I am looking forward to your reply!

[fasnicar]

Dear @GriffyGe, thanks for reporting this.

I think your problem is due to a wrong configuration figuessI gues the error is because blastn can't find a properly indexed database. With phylophlan_write_config_file you specified to index a database of proteins/amino acids with --db_aa diamond but to use blastn for mapping genomes/DNA (-map_dna blastn). Now, blastn can't do a translated search, so here instead you should use diamond, for instance.

Just FYI, if your inputs are all genomes, then you can also consider running the multiple sequence alignments and the phylogeny reconstruction steps on the DNA space instead on amino acids, and you can do so by adding the --force_nucleotides param to both the phylophlan_write_config_file and phylophlan commands.

Please, let me know if something is not clear.

Many thanks,
Francesco