CRITICAL: Can not find file sample_616_metaphlan_bugs_list.tsv

Microbial community profiling HUMAnN
1

Hello,
I am running Humann3.9 together with Metaphlan4 and keep getting: " Can not find file sample_616_metaphlan_bugs_list.tsv".

When inspecting the log file, the first error is related to the version of the database:
humann.search.prescreen - ERROR: The MetaPhlAn taxonomic profile provided was not generated with the database version v3 or vJun23 . Please update your version of MetaPhlAn to at least v3.0 or if you are using MetaPhlAn v4 please use the database vJun23.

I then dowloaded vJun23 and specified in the parameters but still keep getting a similar error:
humann.utilities - DEBUG: b’WARNING: It seems that you do not have Internet access.\nERROR: Cannot find a local database. Please run MetaPhlAn using option “-x <database_name>”.\n You can download the MetaPhlAn database from \n Index of /biobakery4/metaphlan_databases \n \n’
04/17/2024 04:55:41 PM - humann.utilities - CRITICAL: Can not find file sample_616_metaphlan_bugs_list.tsv

The command I used and got this last error was the following:
humann -i sample_616.fastq -o /humann3-output --input-format fastq --nucleotide-database /metaphlan_database_v31 --metaphlan-options=“–stat_q 0.01 -t rel_ab_w_read_stats --tax_lev g --ignore_eukaryotes --ignore_archaea -x mpa_vJun23_CHOCOPhlAnSGB_202307 --bowtie2db /metaphlan_database_vJun23_202307” --protein-database /humann3-database-uniref --threads 5

Any ideas on how to solve this, please?
This is the content of the /metaphlan_database_vJun23_202307 database folder I specified in the --bowtie2db parameter.
image

Thank you,

Thais

2

It sounds like your MetaPhlAn install is not working properly. I would see if you can get MetaPhlAn working with the vJun23 marker database outside of HUMAnN first.

3

Thanks for the reply!

I tried your suggestion with the following command and got the error below:

metaphlan --input_type fastq --stat_q 0.01 -t rel_ab_w_read_stats --tax_lev g --ignore_eukaryotes --ignore_archaea -x mpa_vJun23_CHOCOPhlAnSGB_202307 --bowtie2db /home/metaphlan_database_vJun23_202307 --bowtie2out sample_616.bowtie2.bz2

WARNING: It seems that you do not have Internet access.
ERROR: Cannot find a local database. Please run MetaPhlAn using option “-x <database_name>”.
** You can download the MetaPhlAn database from**
** Index of /biobakery4/metaphlan_databases**

It seems something is wrong recognizing this version of the database. It tells me to include the “-x” option although I already included it.

It’s worth mentioning I am working on a multiuser HPC for the analysis above.

To troubleshoot, I additionally tried running a Metaphlan4 installation I have in my own user (independent of the Humann3.9 installation) with this vJun23 database, and the same error as above appears. But, when I use a different database version - like the mpa_vJan21_CHOCOPhlAnSGB_202103 - it runs perfectly.

So it’s definitely something connected to this vJun23 database. On the other hand, Metaphlan within Humann3.9 does not let me run with a different version.

Any tips how to troubleshoot, please?