Metaphlan4 discordance between relative_abundance and estimated_number_of_reads

Microbial community profiling MetaPhlAn
1

I am using metaphlan4 in a conda environment. When I look at the profile results, there are nonzero relative_abundance value while the estimated_number_of_reads_from_the_clade is 0 or -. Can you please explain how is this possible? I might have missed something. You can look at the unclassified clade_name or t__SGB7962 for example. There are a few others here with that issue.

The profile output for one of my samples is:
#mpa_vJan21_CHOCOPhlAnSGB_202103
#/bulk/IMCbinf_bulk/hramay/projects/Arrieta/PC1000/antibiotic_babies/analysis/metaphlan4/.snakemake/conda/87013b619fc67c1a9da33c332c64e376/bin/metaphlan -t rel_ab_w_read_stats --unclassified_estimation output_merged_sam/merged_data/S00CI-0093_S54.fastq --add_viruses --input_type fastq --bowtie2db /bulk/IMCshared_bulk/shared/dbs/metaphlan4 --bowtie2out output_merged_sam/metaphlan/S00CI-0093_S54_bowtie2.bz2 --nproc 8 -o output_merged_sam/metaphlan/S00CI-0093_S54_profile.txt
#SampleID Metaphlan_Analysis
#estimated_reads_mapped_to_known_clades:428080
#clade_name clade_taxid relative_abundance coverage estimated_number_of_reads_from_the_clade
UNCLASSIFIED -1 1.80294 - 0
k__Bacteria 2 98.19706 0.01638 428080
k__Bacteria|p__Firmicutes 2|1239 74.6264 0.01244 269612
k__Bacteria|p__Actinobacteria 2|201174 11.07805 0.00185 31318
k__Bacteria|p__Proteobacteria 2|1224 6.34208 0.00106 37898
k__Bacteria|p__Bacteroidetes 2|976 6.15053 0.00103 35742
k__Bacteria|p__Firmicutes|c__Bacilli 2|1239|91061 61.71549 0.01029 186348
k__Bacteria|p__Firmicutes|c__Clostridia 2|1239|186801 12.91091 0.00215 44748
k__Bacteria|p__Actinobacteria|c__Actinobacteria 2|201174|1760 11.07805 0.00185 26844
k__Bacteria|p__Proteobacteria|c__Gammaproteobacteria 2|1224|1236 6.34208 0.00106 32484
k__Bacteria|p__Bacteroidetes|c__Bacteroidia 2|976|200643 6.15053 0.00103 30636
k__Bacteria|p__Firmicutes|c__Bacilli|o__Lactobacillales 2|1239|91061|186826 61.71549 0.01029 155290
k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales 2|1239|186801|186802 12.91091 0.00215 37290
k__Bacteria|p__Actinobacteria|c__Actinobacteria|o__Bifidobacteriales 2|201174|1760|85004 11.07805 0.00185 22370
k__Bacteria|p__Bacteroidetes|c__Bacteroidia|o__Bacteroidales 2|976|200643|171549 6.15053 0.00103 25530
k__Bacteria|p__Proteobacteria|c__Gammaproteobacteria|o__Enterobacterales 2|1224|1236|91347 6.13684 0.00102 25930
k__Bacteria|p__Proteobacteria|c__Gammaproteobacteria|o__Pseudomonadales 2|1224|1236|72274 0.20524 3e-05 1140
k__Bacteria|p__Firmicutes|c__Bacilli|o__Lactobacillales|f__Enterococcaceae 2|1239|91061|186826|81852 61.71549 0.01029 124232
k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Lachnospiraceae 2|1239|186801|186802|186803 12.91091 0.00215 29832
k__Bacteria|p__Actinobacteria|c__Actinobacteria|o__Bifidobacteriales|f__Bifidobacteriaceae 2|201174|1760|85004|31953 11.07805 0.00185 17896
k__Bacteria|p__Proteobacteria|c__Gammaproteobacteria|o__Enterobacterales|f__Enterobacteriaceae 2|1224|1236|91347|543 6.13684 0.00102 20744
k__Bacteria|p__Bacteroidetes|c__Bacteroidia|o__Bacteroidales|f__Bacteroidaceae 2|976|200643|171549|815 5.0969 0.00085 16532
k__Bacteria|p__Bacteroidetes|c__Bacteroidia|o__Bacteroidales|f__Bacteroidales_unclassified 2|976|200643|171549| 1.05363 0.00018 3892
k__Bacteria|p__Proteobacteria|c__Gammaproteobacteria|o__Pseudomonadales|f__Pseudomonadaceae 2|1224|1236|72274|135621 0.20524 3e-05 912
k__Bacteria|p__Firmicutes|c__Bacilli|o__Lactobacillales|f__Enterococcaceae|g__Enterococcus 2|1239|91061|186826|81852|1350 61.71549 0.01029 93174
k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Lachnospiraceae|g__Mediterraneibacter 2|1239|186801|186802|186803|2316020 12.91091 0.00215 22374
k__Bacteria|p__Actinobacteria|c__Actinobacteria|o__Bifidobacteriales|f__Bifidobacteriaceae|g__Bifidobacterium 2|201174|1760|85004|31953|1678 11.07805 0.00185 13422
k__Bacteria|p__Proteobacteria|c__Gammaproteobacteria|o__Enterobacterales|f__Enterobacteriaceae|g__Escherichia 2|1224|1236|91347|543|561 6.13684 0.00102 15558
k__Bacteria|p__Bacteroidetes|c__Bacteroidia|o__Bacteroidales|f__Bacteroidaceae|g__Bacteroides 2|976|200643|171549|815|816 5.0969 0.00085 12399
k__Bacteria|p__Bacteroidetes|c__Bacteroidia|o__Bacteroidales|f__Bacteroidales_unclassified|g__Phocaeicola 2|976|200643|171549||909656 1.05363 0.00018 2919
k__Bacteria|p__Proteobacteria|c__Gammaproteobacteria|o__Pseudomonadales|f__Pseudomonadaceae|g__Pseudomonas 2|1224|1236|72274|135621|286 0.20524 3e-05 684
k__Bacteria|p__Firmicutes|c__Bacilli|o__Lactobacillales|f__Enterococcaceae|g__Enterococcus|s__Enterococcus_faecalis 2|1239|91061|186826|81852|1350|1351 61.71549 0.01029 62116
k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Lachnospiraceae|g__Mediterraneibacter|s__Ruminococcus_gnavus 2|1239|186801|186802|186803|2316020|33038 12.91091 0.00215 14916
k__Bacteria|p__Actinobacteria|c__Actinobacteria|o__Bifidobacteriales|f__Bifidobacteriaceae|g__Bifidobacterium|s__Bifidobacterium_longum 2|201174|1760|85004|31953|1678|216816 11.07805 0.00185 8948
k__Bacteria|p__Proteobacteria|c__Gammaproteobacteria|o__Enterobacterales|f__Enterobacteriaceae|g__Escherichia|s__Escherichia_coli 2|1224|1236|91347|543|561|562 6.13684 0.00102 10372
k__Bacteria|p__Bacteroidetes|c__Bacteroidia|o__Bacteroidales|f__Bacteroidaceae|g__Bacteroides|s__Bacteroides_uniformis 2|976|200643|171549|815|816|820 5.0969 0.00085 8266
k__Bacteria|p__Bacteroidetes|c__Bacteroidia|o__Bacteroidales|f__Bacteroidales_unclassified|g__Phocaeicola|s__Phocaeicola_dorei 2|976|200643|171549||909656|357276 1.05363 0.00018 1946
k__Bacteria|p__Proteobacteria|c__Gammaproteobacteria|o__Pseudomonadales|f__Pseudomonadaceae|g__Pseudomonas|s__Pseudomonas_aeruginosa 2|1224|1236|72274|135621|286|287 0.20524 3e-05 456
k__Bacteria|p__Firmicutes|c__Bacilli|o__Lactobacillales|f__Enterococcaceae|g__Enterococcus|s__Enterococcus_faecalis|t__SGB7962 2|1239|91061|186826|81852|1350|1351| 61.71549 - -
k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Lachnospiraceae|g__Mediterraneibacter|s__Ruminococcus_gnavus|t__SGB4584 2|1239|186801|186802|186803|2316020|33038| 12.91091 - -
k__Bacteria|p__Actinobacteria|c__Actinobacteria|o__Bifidobacteriales|f__Bifidobacteriaceae|g__Bifidobacterium|s__Bifidobacterium_longum|t__SGB17248 2|201174|1760|85004|31953|1678|216816| 11.07805 - -
k__Bacteria|p__Proteobacteria|c__Gammaproteobacteria|o__Enterobacterales|f__Enterobacteriaceae|g__Escherichia|s__Escherichia_coli|t__SGB10068 2|1224|1236|91347|543|561|562| 6.13684 - -
k__Bacteria|p__Bacteroidetes|c__Bacteroidia|o__Bacteroidales|f__Bacteroidaceae|g__Bacteroides|s__Bacteroides_uniformis|t__SGB1836_group 2|976|200643|171549|815|816|820| 5.0969 - -
k__Bacteria|p__Bacteroidetes|c__Bacteroidia|o__Bacteroidales|f__Bacteroidales_unclassified|g__Phocaeicola|s__Phocaeicola_dorei|t__SGB1815 2|976|200643|171549||909656|357276| 1.05363 - -
k__Bacteria|p__Proteobacteria|c__Gammaproteobacteria|o__Pseudomonadales|f__Pseudomonadaceae|g__Pseudomonas|s__Pseudomonas_aeruginosa|t__SGB12320 2|1224|1236|72274|135621|286|287| 0.20524 - -

2

Hi,
Does anyone has input on this issue?
Thanks!
Hena

3

Hi @Hena
sorry for the late reply. It seems that the “-t rel_ab_w_read_stats” is not working at the SGB (t__) level of the profiles. Thanking for reporting this issue, I will start working on a fix asap and will let you know when it is out there.

4

Thanks @aitor.blancomiguez !
Hena

5

@aitor.blancomiguez just wondering if you had a chance to fix the problem. I am also concerned about the unclassified results. There are 0s in estimated clade values while there are relative abundance values for the same sample. Can you please explain why is that? You can see for the example file that I have placed above UNCLASSIFIED -1 1.80294 - 0 . Just trying to understand how is this possible? I am seeing this in multiple samples.
Thanks!

6

Hi @Hena
We were working in the fix and we are planning to push the new version at the beginning of next week.
For the Unclassified estimation, we also providing a fix in the next version.

7

Hi @Hena
The new 4.0.3 version is now available in conda and should solve the issues