Dear Qiime2 Experts:

This is a question related to the interpretation of the output of the plugin “qiime diversity adonis”, that I am starting to use. We have rRNA 16S V4 data from a simple experimental design. In a soil translocation experiment we moved soils from location L, H and M with a factorial design (so, samples from for ewample L were moved to L, H and M, and so on…). Then there is two factors 1) “Home” (the origin of soils), and 2) “Away” (the destination of the soils). Then the soils were left for a while to check what happened to the microbial communities.

I have run the analysis in qiime2-2019.4 with no troubles and produced the unw-Unifrac distance matrix. After exploring with PCOAs it seems clear that the “home” factor is very determinant (=samples tend to cluster by the origin), and the “away” needs further testing. In order to check this I run the following two comands:

qiime diversity adonis

–i-distance-matrix unweighted_unifrac_distance_matrix.qza

–m-metadata-file …/metadata.tsv

–p-formula Away:Home

–p-permutations 999

–o-visualization beta-significance/ADONIS-only_interaction–un-unifrac-sign.qzv

qiime diversity adonis

–i-distance-matrix unweighted_unifrac_distance_matrix.qza

–m-metadata-file …/metadata.tsv

–p-formula Away*Home

–p-permutations 999

–o-visualization beta-significance/ADONIS-2factors–un-unifrac-sign.qzv

I attach the two qzv and the metadata ADONIS-2factors–un-unifrac-sign.qzv (264.7 KB) ADONIS-only_interaction–un-unifrac-sign.qzv (264.5 KB) metadata.txt (846 Bytes) . Note that the comparison is not even because some samples we gone during the experiment but all home/away combinations have >3 samples.

I have problems to interpret the results. If the formula used for the model is Away:Home I get the result of the interaction among the two factors, that is clearly significant (p=0.001). When using the formula Away*Home, I get the results of each factor (I understand, treated independently) AND their interaction, that with this formula is clarly NOT significant (p=0.418). The df of the interaction also changed.

So: 1) something wrong?, 2) which is the correct way to test the interaction, 3) I used the order Away(: ,*)Home, instead Home(: ,*)Away because we measure what happend in the soils after being translocated, and want to link that to the origin to detect possible effects; is this correct?

Thanks a lot!