MetaPhlAn 4 + HUMAnN 4 compatibility

You’re absolutely right, @Faeze_Darbaniyan. Your issue does appear to be database‑related. However, I’m not sure if it’s related to MetaPhlAn + HUMAnN 4 compatibility, which I assume was the original focus of this thread based on its title.

I wouldn’t expect humann to require Internet access during normal execution unless it needs to download missing files. Since you already downloaded all databases, that shouldn’t be happening.

After re‑reading your previous comment, I’d like to focus on this specific error:

It seems that you do not have Internet access.
ERROR: Cannot find a local database. Please run MetaPhlAn using option “-x <database_name>”

and the command you are using:

$ humann -r -i ../examples/demo.fastq.gz -o ../SAMPLE/ --threads 16 --protein-database uniref --nucleotide-database chocophlan --metaphlan-options “-t rel_ab_w_read_stats --bowtie2db vOct22 --index mpa_vOct22_CHOCOPhlAnSGB_202403”

From metaphlan --help, the --bowtie2db argument is expected to point to the folder containing the MetaPhlAn database:

$ metaphlan --help
usage: metaphlan --input_type {fastq,fasta,bowtie2out,sam} [--force] [--bowtie2db METAPHLAN_BOWTIE2_DB] [-x INDEX] [--bt2_ps BowTie2 presets] [--bowtie2_exe BOWTIE2_EXE] [--bowtie2_build BOWTIE2_BUILD]
                 [--bowtie2out FILE_NAME] [--min_mapq_val MIN_MAPQ_VAL] [--no_map] [--tmp_dir] [--tax_lev TAXONOMIC_LEVEL] [--min_cu_len] [--min_alignment_len] [--add_viruses] [--ignore_eukaryotes]
                 [--ignore_bacteria] [--ignore_archaea] [--ignore_ksgbs] [--ignore_usgbs] [--stat_q] [--perc_nonzero] [--ignore_markers IGNORE_MARKERS] [--avoid_disqm] [--stat] [-t ANALYSIS TYPE]
                 [--nreads NUMBER_OF_READS] [--pres_th PRESENCE_THRESHOLD] [--clade] [--min_ab] [--profile_vsc] [--vsc_out VSC_OUT] [--vsc_breadth VSC_BREADTH] [-o output file] [--sample_id_key name]
                 [--use_group_representative] [--sample_id value] [-s sam_output_file] [--legacy-output] [--CAMI_format_output] [--unclassified_estimation] [--mpa3] [--biom biom_output] [--mdelim mdelim]
                 [--nproc N] [--subsampling SUBSAMPLING] [--subsampling_output SUBSAMPLING_OUTPUT] [--subsampling_paired SUBSAMPLING_PAIRED] [-1 FORWARD_READS] [-2 REVERSE_READS] [--mapping_subsampling]
                 [--subsampling_seed SUBSAMPLING_SEED] [--install] [--offline] [--force_download] [--read_min_len READ_MIN_LEN] [-v] [-h]
                 [INPUT_FILE] [OUTPUT_FILE]

[...]

Mapping arguments:
[...]
  --bowtie2db METAPHLAN_BOWTIE2_DB
                        Folder containing the MetaPhlAn database. You can specify the location by exporting the DEFAULT_DB_FOLDER variable in the shell.

[...]

Given this, could you double‑check the actual content of your vOct22 directory?
It should contain the Bowtie2 index files and associated metadata for the selected version.

For reference, here is what my directory looks like:

$ ls -1 /home/vllorens/Fran/pipeline_benchmarking/pipeline_benchmarking/data/databases/metaphlan/vOct22_CHOCOPhlAnSGB_202403
mpa_vOct22_CHOCOPhlAnSGB_202403.1.bt2l
mpa_vOct22_CHOCOPhlAnSGB_202403.2.bt2l
mpa_vOct22_CHOCOPhlAnSGB_202403.3.bt2l
mpa_vOct22_CHOCOPhlAnSGB_202403.4.bt2l
mpa_vOct22_CHOCOPhlAnSGB_202403.pkl
mpa_vOct22_CHOCOPhlAnSGB_202403.rev.1.bt2l
mpa_vOct22_CHOCOPhlAnSGB_202403.rev.2.bt2l
mpa_vOct22_CHOCOPhlAnSGB_202403_VINFO.csv
mpa_vOct22_CHOCOPhlAnSGB_202403_VSG.fna

If some files are missing, misnamed, or if the path passed to --bowtie2db doesn’t actually point to this directory, MetaPhlAn will believe the database is absent and may try to download it, triggering that “no Internet access” message.

Let me know what your directory contains, and we can troubleshoot from there.

I really appreciate your fast help! Apparently I was missing some files as you mentioned. It solved my problem with running the demo example.

Just wondering if you are able to run $ humann_test --run-functional-tests-end-to-end as well.

Really grateful for all your kind help!

Faezeh

Certainly! I just ran the humann_test --run-functional-tests-end-to-end command to see if I could run end-to-end tests.

I encountered some errors during my first attempt, beginning with:

ERROR: The MetaPhlAn taxonomic profile provided does not contain the database version vOct22_CHOCOPhlAnSGB_202403 in any of its header lines.
ERROR

After doing some research, I returned to MetaPhlAn’s mapping arguments:

$ metaphlan --help
usage: metaphlan --input_type {fastq,fasta,bowtie2out,sam} [--force] [--bowtie2db METAPHLAN_BOWTIE2_DB] [-x INDEX] [--bt2_ps BowTie2 presets] [--bowtie2_exe BOWTIE2_EXE] [--bowtie2_build BOWTIE2_BUILD]
                 [--bowtie2out FILE_NAME] [--min_mapq_val MIN_MAPQ_VAL] [--no_map] [--tmp_dir] [--tax_lev TAXONOMIC_LEVEL] [--min_cu_len] [--min_alignment_len] [--add_viruses] [--ignore_eukaryotes]
                 [--ignore_bacteria] [--ignore_archaea] [--ignore_ksgbs] [--ignore_usgbs] [--stat_q] [--perc_nonzero] [--ignore_markers IGNORE_MARKERS] [--avoid_disqm] [--stat] [-t ANALYSIS TYPE]
                 [--nreads NUMBER_OF_READS] [--pres_th PRESENCE_THRESHOLD] [--clade] [--min_ab] [--profile_vsc] [--vsc_out VSC_OUT] [--vsc_breadth VSC_BREADTH] [-o output file] [--sample_id_key name]
                 [--use_group_representative] [--sample_id value] [-s sam_output_file] [--legacy-output] [--CAMI_format_output] [--unclassified_estimation] [--mpa3] [--biom biom_output] [--mdelim mdelim]
                 [--nproc N] [--subsampling SUBSAMPLING] [--subsampling_output SUBSAMPLING_OUTPUT] [--subsampling_paired SUBSAMPLING_PAIRED] [-1 FORWARD_READS] [-2 REVERSE_READS] [--mapping_subsampling]
                 [--subsampling_seed SUBSAMPLING_SEED] [--install] [--offline] [--force_download] [--read_min_len READ_MIN_LEN] [-v] [-h]
                 [INPUT_FILE] [OUTPUT_FILE]

[...]

Mapping arguments:
[...]
  --bowtie2db METAPHLAN_BOWTIE2_DB
                        Folder containing the MetaPhlAn database. You can specify the location by exporting the DEFAULT_DB_FOLDER variable in the shell.[default *****/.pixi/envs/default/lib/python3.12/site-packages/metaphlan/metaphlan_databases]

I defined the DEFAULT_DB_FOLDER environment variable to point to that data/databases/metaphlan/vOct22_CHOCOPhlAnSGB_202403/ location (I use its absolute path instead of the relative path shown here).

However, that didn’t fix the errors:

$ DEFAULT_DB_FOLDER=<ABSOLUTE_PATH>/data/databases/metaphlan/vOct22_CHOCOPhlAnSGB_202403/ ; humann_test --run-functional-tests-end-to-end


Please note running functional end to end tests requires all dependencies of HUMAnN.

test_calculate_percent_identity (basic_tests_nucleotide_search.TestBasicHumannNucleotideSearchFunctions.test_calculate_percent_identity)
Test the calculate percent identity function ... ok

[... output omitted for brevity ...]

test_humann_fasta (functional_tests_humann.TestFunctionalHumannEndtoEnd.test_humann_fasta)
Test the standard humann flow on a fasta input file ...
Testing command:
humann --input *****/envs/default/lib/python3.12/site-packages/humann/tests/data/demo.fasta --output /tmp/humann_test_gdwdsxho_fasta --nucleotide-database *****/envs/default/lib/python3.12/site-packages/humann/tests/../data/chocophlan_DEMO --protein-database *****/envs/default/lib/python3.12/site-packages/humann/tests/../data/uniref_DEMO
Output files will be written to: /tmp/humann_test_gdwdsxho_fasta

Running metaphlan ........



ERROR: The MetaPhlAn taxonomic profile provided does not contain the database version vOct22_CHOCOPhlAnSGB_202403 in any of its header lines.
ERROR

[... output omitted for brevity ...]

test_humann_fasta_bypass_translated_search (functional_tests_humann.TestFunctionalHumannEndtoEnd.test_humann_fasta_bypass_translated_search)
Test the standard humann flow on a fasta input file ...
Testing command:
humann --input *****/envs/default/lib/python3.12/site-packages/humann/tests/data/demo.fasta --output /tmp/humann_test_l4uz9dj1_fasta_bypass_translated_search --bypass-translated-search --nucleotide-database *****/envs/default/lib/python3.12/site-packages/humann/tests/../data/chocophlan_DEMO --protein-database *****/envs/default/lib/python3.12/site-packages/humann/tests/../data/uniref_DEMO
Output files will be written to: /tmp/humann_test_l4uz9dj1_fasta_bypass_translated_search

Running metaphlan ........



ERROR: The MetaPhlAn taxonomic profile provided does not contain the database version vOct22_CHOCOPhlAnSGB_202403 in any of its header lines.
ERROR
test_humann_fastq (functional_tests_humann.TestFunctionalHumannEndtoEnd.test_humann_fastq)
Test the standard humann flow on a fastq input file ...
Testing command:
humann --input *****/envs/default/lib/python3.12/site-packages/humann/tests/data/demo.fastq --output /tmp/humann_test_9pjawlwt_fastq --nucleotide-database *****/envs/default/lib/python3.12/site-packages/humann/tests/../data/chocophlan_DEMO --protein-database *****/envs/default/lib/python3.12/site-packages/humann/tests/../data/uniref_DEMO
Output files will be written to: /tmp/humann_test_9pjawlwt_fastq

Running metaphlan ........



ERROR: The MetaPhlAn taxonomic profile provided does not contain the database version vOct22_CHOCOPhlAnSGB_202403 in any of its header lines.
ERROR

[... output omitted for brevity ...]

test_humann_fastq_bypass_translated_search (functional_tests_humann.TestFunctionalHumannEndtoEnd.test_humann_fastq_bypass_translated_search)
Test the standard humann flow on a fastq input file ...
Testing command:
humann --input *****/envs/default/lib/python3.12/site-packages/humann/tests/data/demo.fastq --output /tmp/humann_test_t121b3t3_fastq_bypass_translated_search --bypass-translated-search --nucleotide-database *****/envs/default/lib/python3.12/site-packages/humann/tests/../data/chocophlan_DEMO --protein-database *****/envs/default/lib/python3.12/site-packages/humann/tests/../data/uniref_DEMO
Output files will be written to: /tmp/humann_test_t121b3t3_fastq_bypass_translated_search

Running metaphlan ........



ERROR: The MetaPhlAn taxonomic profile provided does not contain the database version vOct22_CHOCOPhlAnSGB_202403 in any of its header lines.
ERROR
test_humann_fastq_custom_taxonomic_profile (functional_tests_humann.TestFunctionalHumannEndtoEnd.test_humann_fastq_custom_taxonomic_profile)
Test the standard humann flow on a fastq input file ...
Testing command:
humann --input *****/envs/default/lib/python3.12/site-packages/humann/tests/data/demo.fastq --output /tmp/humann_test_cciaavqf_fastq_custom_taxonomic_profile --taxonomic-profile *****/envs/default/lib/python3.12/site-packages/humann/tests/data/demo_metaphlan_bugs_list.tsv --nucleotide-database *****/envs/default/lib/python3.12/site-packages/humann/tests/../data/chocophlan_DEMO --protein-database *****/envs/default/lib/python3.12/site-packages/humann/tests/../data/uniref_DEMO
Output files will be written to: /tmp/humann_test_cciaavqf_fastq_custom_taxonomic_profile


ERROR: The MetaPhlAn taxonomic profile provided does not contain the database version vOct22_CHOCOPhlAnSGB_202403 in any of its header lines.
ERROR

[... output omitted for brevity ...]

test_humann_fastq_biom_output (functional_tests_biom_humann.TestFunctionalHumannEndtoEndBiom.test_humann_fastq_biom_output)
Test the standard humann flow on a fastq input file ...
Testing command:
humann --input *****/envs/default/lib/python3.12/site-packages/humann/tests/data/demo.fastq --output /tmp/humann_test_6zf3z75c_fastq --output-format biom --nucleotide-database *****/envs/default/lib/python3.12/site-packages/humann/tests/../data/chocophlan_DEMO --protein-database *****/envs/default/lib/python3.12/site-packages/humann/tests/../data/uniref_DEMO
Output files will be written to: /tmp/humann_test_6zf3z75c_fastq

Running metaphlan ........



ERROR: The MetaPhlAn taxonomic profile provided does not contain the database version vOct22_CHOCOPhlAnSGB_202403 in any of its header lines.
ERROR

[... output omitted for brevity ...]

======================================================================
ERROR: test_humann_fasta (functional_tests_humann.TestFunctionalHumannEndtoEnd.test_humann_fasta)
Test the standard humann flow on a fasta input file
----------------------------------------------------------------------
Traceback (most recent call last):
  File "*****/envs/default/lib/python3.12/site-packages/humann/tests/utils.py", line 59, in run_command
    subprocess.check_call(command)
  File "*****/envs/default/lib/python3.12/subprocess.py", line 413, in check_call
    raise CalledProcessError(retcode, cmd)
subprocess.CalledProcessError: Command '['humann', '--input', '*****/envs/default/lib/python3.12/site-packages/humann/tests/data/demo.fasta', '--output', '/tmp/humann_test_gdwdsxho_fasta', '--nucleotide-database', '*****/envs/default/lib/python3.12/site-packages/humann/tests/../data/chocophlan_DEMO', '--protein-database', '*****/envs/default/lib/python3.12/site-packages/humann/tests/../data/uniref_DEMO']' returned non-zero exit status 1.

During handling of the above exception, another exception occurred:

Traceback (most recent call last):
  File "*****/envs/default/lib/python3.12/site-packages/humann/tests/functional_tests_humann.py", line 45, in test_humann_fasta
    utils.run_humann(command)
  File "*****/envs/default/lib/python3.12/site-packages/humann/tests/utils.py", line 50, in run_humann
    run_command(command)
  File "*****/envs/default/lib/python3.12/site-packages/humann/tests/utils.py", line 61, in run_command
    raise EnvironmentError("Warning: Unable to execute command in test.\n"+" ".join(command))
OSError: Warning: Unable to execute command in test.
humann --input *****/envs/default/lib/python3.12/site-packages/humann/tests/data/demo.fasta --output /tmp/humann_test_gdwdsxho_fasta --nucleotide-database *****/envs/default/lib/python3.12/site-packages/humann/tests/../data/chocophlan_DEMO --protein-database *****/envs/default/lib/python3.12/site-packages/humann/tests/../data/uniref_DEMO

======================================================================
ERROR: test_humann_fasta_bypass_translated_search (functional_tests_humann.TestFunctionalHumannEndtoEnd.test_humann_fasta_bypass_translated_search)
Test the standard humann flow on a fasta input file
----------------------------------------------------------------------
Traceback (most recent call last):
  File "*****/envs/default/lib/python3.12/site-packages/humann/tests/utils.py", line 59, in run_command
    subprocess.check_call(command)
  File "*****/envs/default/lib/python3.12/subprocess.py", line 413, in check_call
    raise CalledProcessError(retcode, cmd)
subprocess.CalledProcessError: Command '['humann', '--input', '*****/envs/default/lib/python3.12/site-packages/humann/tests/data/demo.fasta', '--output', '/tmp/humann_test_l4uz9dj1_fasta_bypass_translated_search', '--bypass-translated-search', '--nucleotide-database', '*****/envs/default/lib/python3.12/site-packages/humann/tests/../data/chocophlan_DEMO', '--protein-database', '*****/envs/default/lib/python3.12/site-packages/humann/tests/../data/uniref_DEMO']' returned non-zero exit status 1.

During handling of the above exception, another exception occurred:

Traceback (most recent call last):
  File "*****/envs/default/lib/python3.12/site-packages/humann/tests/functional_tests_humann.py", line 182, in test_humann_fasta_bypass_translated_search
    utils.run_humann(command)
  File "*****/envs/default/lib/python3.12/site-packages/humann/tests/utils.py", line 50, in run_humann
    run_command(command)
  File "*****/envs/default/lib/python3.12/site-packages/humann/tests/utils.py", line 61, in run_command
    raise EnvironmentError("Warning: Unable to execute command in test.\n"+" ".join(command))
OSError: Warning: Unable to execute command in test.
humann --input *****/envs/default/lib/python3.12/site-packages/humann/tests/data/demo.fasta --output /tmp/humann_test_l4uz9dj1_fasta_bypass_translated_search --bypass-translated-search --nucleotide-database *****/envs/default/lib/python3.12/site-packages/humann/tests/../data/chocophlan_DEMO --protein-database *****/envs/default/lib/python3.12/site-packages/humann/tests/../data/uniref_DEMO

======================================================================
ERROR: test_humann_fastq (functional_tests_humann.TestFunctionalHumannEndtoEnd.test_humann_fastq)
Test the standard humann flow on a fastq input file
----------------------------------------------------------------------
Traceback (most recent call last):
  File "*****/envs/default/lib/python3.12/site-packages/humann/tests/utils.py", line 59, in run_command
    subprocess.check_call(command)
  File "*****/envs/default/lib/python3.12/subprocess.py", line 413, in check_call
    raise CalledProcessError(retcode, cmd)
subprocess.CalledProcessError: Command '['humann', '--input', '*****/envs/default/lib/python3.12/site-packages/humann/tests/data/demo.fastq', '--output', '/tmp/humann_test_9pjawlwt_fastq', '--nucleotide-database', '*****/envs/default/lib/python3.12/site-packages/humann/tests/../data/chocophlan_DEMO', '--protein-database', '*****/envs/default/lib/python3.12/site-packages/humann/tests/../data/uniref_DEMO']' returned non-zero exit status 1.

During handling of the above exception, another exception occurred:

Traceback (most recent call last):
  File "*****/envs/default/lib/python3.12/site-packages/humann/tests/functional_tests_humann.py", line 26, in test_humann_fastq
    utils.run_humann(command)
  File "*****/envs/default/lib/python3.12/site-packages/humann/tests/utils.py", line 50, in run_humann
    run_command(command)
  File "*****/envs/default/lib/python3.12/site-packages/humann/tests/utils.py", line 61, in run_command
    raise EnvironmentError("Warning: Unable to execute command in test.\n"+" ".join(command))
OSError: Warning: Unable to execute command in test.
humann --input *****/envs/default/lib/python3.12/site-packages/humann/tests/data/demo.fastq --output /tmp/humann_test_9pjawlwt_fastq --nucleotide-database *****/envs/default/lib/python3.12/site-packages/humann/tests/../data/chocophlan_DEMO --protein-database *****/envs/default/lib/python3.12/site-packages/humann/tests/../data/uniref_DEMO

======================================================================
ERROR: test_humann_fastq_bypass_translated_search (functional_tests_humann.TestFunctionalHumannEndtoEnd.test_humann_fastq_bypass_translated_search)
Test the standard humann flow on a fastq input file
----------------------------------------------------------------------
Traceback (most recent call last):
  File "*****/envs/default/lib/python3.12/site-packages/humann/tests/utils.py", line 59, in run_command
    subprocess.check_call(command)
  File "*****/envs/default/lib/python3.12/subprocess.py", line 413, in check_call
    raise CalledProcessError(retcode, cmd)
subprocess.CalledProcessError: Command '['humann', '--input', '*****/envs/default/lib/python3.12/site-packages/humann/tests/data/demo.fastq', '--output', '/tmp/humann_test_t121b3t3_fastq_bypass_translated_search', '--bypass-translated-search', '--nucleotide-database', '*****/envs/default/lib/python3.12/site-packages/humann/tests/../data/chocophlan_DEMO', '--protein-database', '*****/envs/default/lib/python3.12/site-packages/humann/tests/../data/uniref_DEMO']' returned non-zero exit status 1.

During handling of the above exception, another exception occurred:

Traceback (most recent call last):
  File "*****/envs/default/lib/python3.12/site-packages/humann/tests/functional_tests_humann.py", line 162, in test_humann_fastq_bypass_translated_search
    utils.run_humann(command)
  File "*****/envs/default/lib/python3.12/site-packages/humann/tests/utils.py", line 50, in run_humann
    run_command(command)
  File "*****/envs/default/lib/python3.12/site-packages/humann/tests/utils.py", line 61, in run_command
    raise EnvironmentError("Warning: Unable to execute command in test.\n"+" ".join(command))
OSError: Warning: Unable to execute command in test.
humann --input *****/envs/default/lib/python3.12/site-packages/humann/tests/data/demo.fastq --output /tmp/humann_test_t121b3t3_fastq_bypass_translated_search --bypass-translated-search --nucleotide-database *****/envs/default/lib/python3.12/site-packages/humann/tests/../data/chocophlan_DEMO --protein-database *****/envs/default/lib/python3.12/site-packages/humann/tests/../data/uniref_DEMO

======================================================================
ERROR: test_humann_fastq_custom_taxonomic_profile (functional_tests_humann.TestFunctionalHumannEndtoEnd.test_humann_fastq_custom_taxonomic_profile)
Test the standard humann flow on a fastq input file
----------------------------------------------------------------------
Traceback (most recent call last):
  File "*****/envs/default/lib/python3.12/site-packages/humann/tests/utils.py", line 59, in run_command
    subprocess.check_call(command)
  File "*****/envs/default/lib/python3.12/subprocess.py", line 413, in check_call
    raise CalledProcessError(retcode, cmd)
subprocess.CalledProcessError: Command '['humann', '--input', '*****/envs/default/lib/python3.12/site-packages/humann/tests/data/demo.fastq', '--output', '/tmp/humann_test_cciaavqf_fastq_custom_taxonomic_profile', '--taxonomic-profile', '*****/envs/default/lib/python3.12/site-packages/humann/tests/data/demo_metaphlan_bugs_list.tsv', '--nucleotide-database', '*****/envs/default/lib/python3.12/site-packages/humann/tests/../data/chocophlan_DEMO', '--protein-database', '*****/envs/default/lib/python3.12/site-packages/humann/tests/../data/uniref_DEMO']' returned non-zero exit status 1.

During handling of the above exception, another exception occurred:

Traceback (most recent call last):
  File "*****/envs/default/lib/python3.12/site-packages/humann/tests/functional_tests_humann.py", line 243, in test_humann_fastq_custom_taxonomic_profile
    utils.run_humann(command)
  File "*****/envs/default/lib/python3.12/site-packages/humann/tests/utils.py", line 50, in run_humann
    run_command(command)
  File "*****/envs/default/lib/python3.12/site-packages/humann/tests/utils.py", line 61, in run_command
    raise EnvironmentError("Warning: Unable to execute command in test.\n"+" ".join(command))
OSError: Warning: Unable to execute command in test.
humann --input *****/envs/default/lib/python3.12/site-packages/humann/tests/data/demo.fastq --output /tmp/humann_test_cciaavqf_fastq_custom_taxonomic_profile --taxonomic-profile *****/envs/default/lib/python3.12/site-packages/humann/tests/data/demo_metaphlan_bugs_list.tsv --nucleotide-database *****/envs/default/lib/python3.12/site-packages/humann/tests/../data/chocophlan_DEMO --protein-database *****/envs/default/lib/python3.12/site-packages/humann/tests/../data/uniref_DEMO

======================================================================
ERROR: test_humann_fastq_biom_output (functional_tests_biom_humann.TestFunctionalHumannEndtoEndBiom.test_humann_fastq_biom_output)
Test the standard humann flow on a fastq input file
----------------------------------------------------------------------
Traceback (most recent call last):
  File "*****/envs/default/lib/python3.12/site-packages/humann/tests/utils.py", line 59, in run_command
    subprocess.check_call(command)
  File "*****/envs/default/lib/python3.12/subprocess.py", line 413, in check_call
    raise CalledProcessError(retcode, cmd)
subprocess.CalledProcessError: Command '['humann', '--input', '*****/envs/default/lib/python3.12/site-packages/humann/tests/data/demo.fastq', '--output', '/tmp/humann_test_6zf3z75c_fastq', '--output-format', 'biom', '--nucleotide-database', '*****/envs/default/lib/python3.12/site-packages/humann/tests/../data/chocophlan_DEMO', '--protein-database', '*****/envs/default/lib/python3.12/site-packages/humann/tests/../data/uniref_DEMO']' returned non-zero exit status 1.

During handling of the above exception, another exception occurred:

Traceback (most recent call last):
  File "*****/envs/default/lib/python3.12/site-packages/humann/tests/functional_tests_biom_humann.py", line 28, in test_humann_fastq_biom_output
    utils.run_humann(command)
  File "*****/envs/default/lib/python3.12/site-packages/humann/tests/utils.py", line 50, in run_humann
    run_command(command)
  File "*****/envs/default/lib/python3.12/site-packages/humann/tests/utils.py", line 61, in run_command
    raise EnvironmentError("Warning: Unable to execute command in test.\n"+" ".join(command))
OSError: Warning: Unable to execute command in test.
humann --input *****/envs/default/lib/python3.12/site-packages/humann/tests/data/demo.fastq --output /tmp/humann_test_6zf3z75c_fastq --output-format biom --nucleotide-database *****/envs/default/lib/python3.12/site-packages/humann/tests/../data/chocophlan_DEMO --protein-database *****/envs/default/lib/python3.12/site-packages/humann/tests/../data/uniref_DEMO

----------------------------------------------------------------------
Ran 189 tests in 2413.340s

FAILED (errors=6)

Note that I have been successfully running HUMAnN 4 for the last several months.

I’m glad you managed to solve all the issues, @Faeze_Darbaniyan. Happy profiling! :slight_smile:

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