No species detected in RNA data


I’m trying to run metaphlan → humann (both version 3) with a mouse microbiome sample. I have RNA and DNA samples, and ran metaphlan (following kneaddata) on both, but only the DNA was able to detect any bacteria (all Unkown). I was wondering if this is likely because the RNA isn’t sequenced deep enough or if this is a typical problem associated with RNA given how metaphlan operates?

Also, since the DNA should have the same bacterial species / present, could the output from the DNA metaphlan run be used as a sort of reference for humann?

Thank you!

What do you mean by “DNA samples” and “RNA samples”? Is the DNA experiment whole genome sequencing or exome sequencing? Is the RNA experiment poly-A tail enrichment or total RNA extraction followed by ribosomal depletion? Some protocols are suited to sequencing bacteria but others are not. What tissue or fluid are you analysing?


These were stool samples from mice.

DNA samples: extracted genomic DNA
RNA samples: total RNA extraction followed by DNA and ribosomal depletion