Abstract
The nearest taxon index (NTI) and beta nearest taxon index (βNTI) are popular metrics used to infer community assembly in a variety of ecosystems. However, the dependence these metrics have on the underlying phylogenetic tree is often overlooked and can potentially lead to erroneous conclusions. In this paper, we investigated how misclassified sequences in the phylogenetic tree and reduction of the metacommunity richness affect estimates of the assembly processes. We estimated assembly processes in two datasets investigating fish-microbiota interactions. We found that non-bacterial misclassified sequences can drastically change the interpretation of assembly processes. NTI consistently increased after misclassified sequences were removed, which changed the ecological interpretation for 82.8% of the communities in one of the datasets. Similarly, βNTI was strongly affected by misclassified sequences in the phylogenetic tree, changing the ecological interpretation for 44.7% (Dataset 1) and 6.3% (Dataset 2) of the community comparisons. Lower metacommunity richness generally resulted in decreased NTI and increased βNTI, but the effect sizes were small and generally did not result in a change in ecological interpretation. Based on these findings, we recommend: (1) constructing and visually inspecting a phylogenetic tree for 16 S rRNA gene amplicon datasets to identify and remove misclassified sequences, and (2) evaluating the robustness of community assembly conclusions by performing analyses with varying metacommunity richness and clearly specifying the metacommunity used for the null model approach.