Inconsistent pathway stratification


Could anyone please help me with how pathways are stratified? I noticed that for the pathways did not have any species assigned, in the pathabundance.tsv, it could either be only having the main entry, such as:

S1_pathabundance.tsv:PWY-5677: succinate fermentation to butanoate 114.7999908573

or, in some cases, it could have the main entry and the sub-entry “unclassified”, e.g.,

S2_pathabundance.tsv:ARGORNPROST-PWY 66.8159830865
S2_pathabundance.tsv:ARGORNPROST-PWY|unclassified 48.8755175661

What may have caused the difference? This is Humann 3.0.0.


If you only see a total for a pathway (i.e. no stratifications), it means that the pathway was satisfied at the community level but not by any single detected species. For example, maybe the pathway requires reactions A + B + C + D and species 1 contributes A + B and species 2 contributes C + D.

In the second example you’re seeing stratification to unclassified, which means that all of the required reactions were detected by translated search, potentially coming from different species. The total is larger than the unclassified stratification because there are additional incomplete pathways being satisfied at the community level.

Returning to my example above, if we also saw one copy of A + B + C + D in the unclassified mapping, then there would be two complete copies at the community (total) level: the one from unclassified and another one from the combination of Species 1 and Species 2.