# question about unclassified **URL:** \ **Category:** MetaPhlAn\ **Created:** [April 8, 2020, 6:40pm UTC](https://forum.biobakery.org/t/question-about-unclassified/329 "2020-04-08T18:40:46Z")\ **Posts on this page:** 20\ **Page:** 1
**Author:** ![marilyne](https://avatars.discourse-cdn.com/v4/letter/m/a88e4f/32.png) [@marilyne](https://forum.biobakery.org/u/marilyne)\ **Post date:** [April 8, 2020, 6:40pm UTC](https://forum.biobakery.org/t/question-about-unclassified/329/1 "2020-04-08T18:40:46Z")
Dear metaphlan team, In my profile result (see below the last line) I got t\_\_Enterobacter\_cloacae\_unclassified 93.68831 My first question is: what does it mean ‘t’ ? My second question is: what does it mean ‘unclassified’ ? Does it mean it is a new subspecies or a new strain? #SampleID Metaphlan2\_Analysis k\_\_Bacteria 100.0 k\_\_Bacteria|p\_\_Proteobacteria 100.0 k\_\_Bacteria|p\_\_Proteobacteria|c\_\_Gammaproteobacteria 100.0 k\_\_Bacteria|p\_\_Proteobacteria|c\_\_Gammaproteobacteria|o\_\_Enterobacteriales 100.0 k\_\_Bacteria|p\_\_Proteobacteria|c\_\_Gammaproteobacteria|o\_\_Enterobacteriales|f\_\_Enterobacteriaceae 100.0 k\_\_Bacteria|p\_\_Proteobacteria|c\_\_Gammaproteobacteria|o\_\_Enterobacteriales|f\_\_Enterobacteriaceae|g\_\_Enterobacter 93.68831 k\_\_Bacteria|p\_\_Proteobacteria|c\_\_Gammaproteobacteria|o\_\_Enterobacteriales|f\_\_Enterobacteriaceae|g\_\_Escherichia 6.31169 k\_\_Bacteria|p\_\_Proteobacteria|c\_\_Gammaproteobacteria|o\_\_Enterobacteriales|f\_\_Enterobacteriaceae|g\_\_Enterobacter|s\_\_Enterobacter\_cloacae 93.68831 k\_\_Bacteria|p\_\_Proteobacteria|c\_\_Gammaproteobacteria|o\_\_Enterobacteriales|f\_\_Enterobacteriaceae|g\_\_Escherichia|s\_\_Escherichia\_unclassified 6.31169 k\_\_Bacteria|p\_\_Proteobacteria|c\_\_Gammaproteobacteria|o\_\_Enterobacteriales|f\_\_Enterobacteriaceae|g\_\_Enterobacter|s\_\_Enterobacter\_cloacae|t\_\_Enterobacter\_cloacae\_unclassified93.68831 Best regards, By the way this group is really helpful! Marilyne ---
**Author:** ![fbeghini](https://yyz1.discourse-cdn.com/flex027/user_avatar/forum.biobakery.org/fbeghini/32/81_2.png) [@fbeghini](https://forum.biobakery.org/u/fbeghini)\ **Post date:** [April 14, 2020, 3:15pm UTC](https://forum.biobakery.org/t/question-about-unclassified/329/2 "2020-04-14T15:15:08Z")
Hi Marilyne, `t __` is the taxonomy rank which identifies the strain, in this case, `unclassified` means that MetaPhlAn was not able to identify the exact strain present in the community, otherwise, you’ll get something like `t__ GCA_XXXXXX`. It’s fine to filter all the `t__` entries and focus only on the species level assigments. Best, Francesco ---
**Author:** ![marilyne](https://avatars.discourse-cdn.com/v4/letter/m/a88e4f/32.png) [@marilyne](https://forum.biobakery.org/u/marilyne)\ **Post date:** [April 15, 2020, 5:03pm UTC](https://forum.biobakery.org/t/question-about-unclassified/329/3 "2020-04-15T17:03:02Z")
Hi Francesco, Thank you I would like to know if metaphlan includes also fungi ? Or it is only bacteria ? ---
**Author:** ![fbeghini](https://yyz1.discourse-cdn.com/flex027/user_avatar/forum.biobakery.org/fbeghini/32/81_2.png) [@fbeghini](https://forum.biobakery.org/u/fbeghini)\ **Post date:** [April 16, 2020, 11:49am UTC](https://forum.biobakery.org/t/question-about-unclassified/329/4 "2020-04-16T11:49:09Z")
Yes, MetaPhlAn2 includes markers for some fungi, you can check which species can be profiles by looking into the marker information [marker information file here](https://www.dropbox.com/s/nhhx7i7glwdahru/mpa_v20_m200_marker_info.txt.bz2?dl=1). You can also have a look at [MetaPhlAn 3.0](https://github.com/biobakery/MetaPhlAn/tree/3.0) which includes more fungal markers ---
**Author:** ![marilyne](https://avatars.discourse-cdn.com/v4/letter/m/a88e4f/32.png) [@marilyne](https://forum.biobakery.org/u/marilyne)\ **Post date:** [April 23, 2020, 7:56pm UTC](https://forum.biobakery.org/t/question-about-unclassified/329/5 "2020-04-23T19:56:29Z")
Hi Francesco Great thanks! ---
**Author:** ![marilyne](https://avatars.discourse-cdn.com/v4/letter/m/a88e4f/32.png) [@marilyne](https://forum.biobakery.org/u/marilyne)\ **Post date:** [June 2, 2020, 2:24pm UTC](https://forum.biobakery.org/t/question-about-unclassified/329/6 "2020-06-02T14:24:46Z")
Hi Francesco, I have difficulties to install metaphlan3 do you mind to provide me the anaconda command lines ? I tried conda install metaphlan=3.0=pyh5ca1d4c\_2 --no-channel-priority but got: Fetching package metadata …An unexpected error has occurred. ---
**Author:** ![marilyne](https://avatars.discourse-cdn.com/v4/letter/m/a88e4f/32.png) [@marilyne](https://forum.biobakery.org/u/marilyne)\ **Post date:** [June 2, 2020, 4:38pm UTC](https://forum.biobakery.org/t/question-about-unclassified/329/7 "2020-06-02T16:38:11Z")
finally it works but got Disk quota exceeded how can I change the directory of installation envs/ and pkgs/ ? ---
**Author:** ![fbeghini](https://yyz1.discourse-cdn.com/flex027/user_avatar/forum.biobakery.org/fbeghini/32/81_2.png) [@fbeghini](https://forum.biobakery.org/u/fbeghini)\ **Post date:** [June 4, 2020, 12:36pm UTC](https://forum.biobakery.org/t/question-about-unclassified/329/8 "2020-06-04T12:36:51Z")
> [@marilyne](#): > > conda install metaphlan=3.0=pyh5ca1d4c\_2 --no-channel-priority There’s a newer build pyh5ca1d4c\_4 > [@marilyne](#): > > how can I change the directory of installation envs/ and pkgs/ You should move the whole anaconda installation to another location, see [this](https://docs.anaconda.com/anaconda/user-guide/tasks/move-directory/) for more information ---
**Author:** ![marilyne](https://avatars.discourse-cdn.com/v4/letter/m/a88e4f/32.png) [@marilyne](https://forum.biobakery.org/u/marilyne)\ **Post date:** [July 22, 2020, 8:49pm UTC](https://forum.biobakery.org/t/question-about-unclassified/329/9 "2020-07-22T20:49:50Z")
Hi Francesco, I could change the directory of installation and install metaphlan3 but when I run it I get FileNotFoundError: [Errno 2] No such file or directory: ‘/dev/fd/63’ do you know how to solve this problem ? It would be great if you could help me. regards Marilyne ---
**Author:** ![marilyne](https://avatars.discourse-cdn.com/v4/letter/m/a88e4f/32.png) [@marilyne](https://forum.biobakery.org/u/marilyne)\ **Post date:** [July 22, 2020, 9:42pm UTC](https://forum.biobakery.org/t/question-about-unclassified/329/10 "2020-07-22T21:42:16Z")
metaphlan --input\_type fastq \<(zcat …/…/batch9\_data/$1\_R1\_001.cleaned.fastq.gz …/…/batch9\_data/$1\_R2\_001.cleaned.fastq.gz) --bowtie2db …/…/database\_metaphlan/ --nproc 12 --bowtie2out $1.bowtie2out.txt -o $1\_profile.txt this is the command I use ---
**Author:** ![marilyne](https://avatars.discourse-cdn.com/v4/letter/m/a88e4f/32.png) [@marilyne](https://forum.biobakery.org/u/marilyne)\ **Post date:** [July 22, 2020, 9:43pm UTC](https://forum.biobakery.org/t/question-about-unclassified/329/11 "2020-07-22T21:43:30Z")
I get: slurmstepd: error: task/cgroup: unable to add task[pid=26585] to memory cg ‘(null)’ Use of uninitialized value $bt2\_args[2] in join or string at /home/mdebieu/.conda/envs/metaphlan\_version3b/bin/bowtie2 line 423. Use of uninitialized value bt2\_args[3] in join or string at /home/mdebieu/.conda/envs/metaphlan\_version3b/bin/bowtie2 line 423. Use of uninitialized value _[2] in string eq at /home/mdebieu/.conda/envs/metaphlan\_version3b/bin/bowtie2 line 360. Use of uninitialized value $_[3] in string eq at /home/mdebieu/.conda/envs/metaphlan\_version3b/bin/bowtie2 line 360. Use of uninitialized value in exists at /home/mdebieu/.conda/envs/metaphlan\_version3b/bin/bowtie2 line 81. Use of uninitialized value in exists at /home/mdebieu/.conda/envs/metaphlan\_version3b/bin/bowtie2 line 81. Use of uninitialized value $bt2\_args[2] in join or string at /home/mdebieu/.conda/envs/metaphlan\_version3b/bin/bowtie2 line 459. Use of uninitialized value $bt2\_args[3] in join or string at /home/mdebieu/.conda/envs/metaphlan\_version3b/bin/bowtie2 line 459. Traceback (most recent call last): File “/home/mdebieu/.conda/envs/metaphlan\_version3b/bin/read\_fastx.py”, line 10, in sys.exit(main()) File “/home/mdebieu/.conda/envs/metaphlan\_version3b/lib/python3.7/site-packages/metaphlan/utils/read\_fastx.py”, line 155, in main nreads += read\_and\_write\_raw(f, opened=False, min\_len=min\_len) File “/home/mdebieu/.conda/envs/metaphlan\_version3b/lib/python3.7/site-packages/metaphlan/utils/read\_fastx.py”, line 118, in read\_and\_write\_raw with fopen(fd) as inf: File “/home/mdebieu/.conda/envs/metaphlan\_version3b/lib/python3.7/site-packages/metaphlan/utils/read\_fastx.py”, line 53, in fopen return open(fn) FileNotFoundError: [Errno 2] No such file or directory: ‘/dev/fd/63’ ---
**Author:** ![marilyne](https://avatars.discourse-cdn.com/v4/letter/m/a88e4f/32.png) [@marilyne](https://forum.biobakery.org/u/marilyne)\ **Post date:** [July 23, 2020, 4:01am UTC](https://forum.biobakery.org/t/question-about-unclassified/329/12 "2020-07-23T04:01:35Z")
Finally it worked with /metaphlan …/…/batch9\_data/JC-0355\_S647\_L002\_R1\_001.cleaned.fastq.gz,…/…/batch9\_data/JC-0355\_S647\_L002\_R2\_001.cleaned.fastq.gz --bowtie2out JC-0355\_S647\_L002.bowtie2.bz2 --bowtie2db …/…/database\_metaphlan/ --nproc 12 --input\_type fastq -o JC-0355\_S647\_L002\_profile.txt what is the last version of metaphlan3 and the last version of the databases ? how much memory should I request ? what does it mean ‘additional\_species’ is it all possible subspecies ? ---
**Author:** ![fbeghini](https://yyz1.discourse-cdn.com/flex027/user_avatar/forum.biobakery.org/fbeghini/32/81_2.png) [@fbeghini](https://forum.biobakery.org/u/fbeghini)\ **Post date:** [July 23, 2020, 8:46am UTC](https://forum.biobakery.org/t/question-about-unclassified/329/13 "2020-07-23T08:46:41Z")
> [@marilyne](#): > > what is the last version of metaphlan3 and the last version of the databases ? The latest version available is 3.0.1, you can check for new releases [here]([Package Recipe 'metaphlan' — Bioconda documentation](https://bioconda.github.io/recipes/metaphlan/README.html), the latest database is v30. > [@marilyne](#): > > how much memory should I request ? To answer this, you should manually check how much memory uses to profile one of the metagenomes, more or less it should not require more than 6GB. > [@marilyne](#): > > what does it mean ‘additional\_species’ is it all possible subspecies ? See this [Unexpected output (format) - #2 by fbeghini](https://forum.biobakery.org/t/unexpected-output-format/658/2) ---
**Author:** ![marilyne](https://avatars.discourse-cdn.com/v4/letter/m/a88e4f/32.png) [@marilyne](https://forum.biobakery.org/u/marilyne)\ **Post date:** [July 23, 2020, 11:39am UTC](https://forum.biobakery.org/t/question-about-unclassified/329/14 "2020-07-23T11:39:53Z")
Hi Francesco, Great! Thank you! ---
**Author:** ![marilyne](https://avatars.discourse-cdn.com/v4/letter/m/a88e4f/32.png) [@marilyne](https://forum.biobakery.org/u/marilyne)\ **Post date:** [September 26, 2020, 1:23am UTC](https://forum.biobakery.org/t/question-about-unclassified/329/15 "2020-09-26T01:23:52Z")
Hi Francesco, What is the difference between metaphlan2 and metaphlan3 ? Is the method still the same ? How many markers are used per species ? what is the average length of a marker ? Best regards, Marilyne ---
**Author:** ![fbeghini](https://yyz1.discourse-cdn.com/flex027/user_avatar/forum.biobakery.org/fbeghini/32/81_2.png) [@fbeghini](https://forum.biobakery.org/u/fbeghini)\ **Post date:** [September 28, 2020, 8:12am UTC](https://forum.biobakery.org/t/question-about-unclassified/329/16 "2020-09-28T08:12:20Z")
Yes, the method is practically identical, we introduced a couple of QC on the alignment quality and a new expanded database comprising 12k species (see [https://forum.biobakery.org/t/can-you-tell-us-about-the-db-updates/310](https://forum.biobakery.org/t/can-you-tell-us-about-the-db-updates/310)). We try to use maximum 150 markers per species, for complete marker statistics you can check the marker info file here [https://www.dropbox.com/sh/7qze7m7g9fe2xjg/AAAlyQITZuUCtBUJxpxhIroIa/mpa\_v30\_CHOCOPhlAn\_201901\_marker\_info.txt.bz2?dl=1](https://www.dropbox.com/sh/7qze7m7g9fe2xjg/AAAlyQITZuUCtBUJxpxhIroIa/mpa_v30_CHOCOPhlAn_201901_marker_info.txt.bz2?dl=1) ---
**Author:** ![marilyne](https://avatars.discourse-cdn.com/v4/letter/m/a88e4f/32.png) [@marilyne](https://forum.biobakery.org/u/marilyne)\ **Post date:** [December 3, 2020, 4:38pm UTC](https://forum.biobakery.org/t/question-about-unclassified/329/17 "2020-12-03T16:38:39Z")
Hi Francesco, I am planning to use phyloseq to analyse alpha and beta diversities with my mataphlan2 outputs. So, I need three inputs: a table, a tree and a fasta file. I know how to get the table I can use the script “merge\_metaphlan\_tables.py”. But how can I generate the tree and the fasta file? I would like to know if a script is available to extract the fasta sequences ? And I suppose I could generate a tree using those sequences ? Which R package would you recommend to create the tree? Best regards, Marilyne ---
**Author:** ![fbeghini](https://yyz1.discourse-cdn.com/flex027/user_avatar/forum.biobakery.org/fbeghini/32/81_2.png) [@fbeghini](https://forum.biobakery.org/u/fbeghini)\ **Post date:** [December 7, 2020, 3:19pm UTC](https://forum.biobakery.org/t/question-about-unclassified/329/18 "2020-12-07T15:19:11Z")
Hi Marilyne, you can find the Newick tree built with all the genomes included in MetaPhlAn in the GitHub repository ([https://github.com/biobakery/MetaPhlAn/blob/master/metaphlan/utils/mpa\_v30\_CHOCOPhlAn\_201901\_species\_tree.nwk](https://github.com/biobakery/MetaPhlAn/blob/master/metaphlan/utils/mpa_v30_CHOCOPhlAn_201901_species_tree.nwk)), I don’t recall what the fasta file in phyloseq is needed for, but if you need to perform alpha and beta diversity measures, the tree should be enough ---
**Author:** ![marilyne](https://avatars.discourse-cdn.com/v4/letter/m/a88e4f/32.png) [@marilyne](https://forum.biobakery.org/u/marilyne)\ **Post date:** [December 9, 2020, 10:32pm UTC](https://forum.biobakery.org/t/question-about-unclassified/329/19 "2020-12-09T22:32:33Z")
Hi Francesco, Thank you! I was able to create a phyloseq object. I have now a question about the otu table, I got it from the script “merge\_metaphlan\_tables.py”. But when I run analysis with phyloseq, I got this error: “function accepts only integers (counts)” I assume I need to have the number of reads in my otu\_table, so I was thinking about to multiply the relative abundance per the total number of reads but it seems that I do not have relative abundance in the otu table. I saw that when I calculate the sum for each sample, I get a number between 100 and 800. What does it mean ? Is it correlated with the number of reads ? Or Do I need an other script to convert it in read counts ? Best regards, Marilyne ---
**Author:** ![fbeghini](https://yyz1.discourse-cdn.com/flex027/user_avatar/forum.biobakery.org/fbeghini/32/81_2.png) [@fbeghini](https://forum.biobakery.org/u/fbeghini)\ **Post date:** [December 10, 2020, 8:12am UTC](https://forum.biobakery.org/t/question-about-unclassified/329/20 "2020-12-10T08:12:48Z")
For importing the MetaPhlAn profiles into phyloseq you can have a look at this R function [Import a table of MetaPhlAn taxonomic abundances into phyloseq (github.com)](https://gist.github.com/lwaldron/512d1925a8102e921f05c5b25de7ec94). There is the possibility to have the estimated read counts in the output profile when MetaPhlAn is run with the option `-t rel_ab_w_read_stats`