Amphibian skin microbiomes exhibited high microeukaryote diversity and were dominated by Proteobacteria
In this study, we examined amphibian skin microbiome structure and diversity with respect to geography and host genetics. In analyzing both bacterial and microeukaryote sequences, we recovered microbial associations with geographic and host genetic factors, as well as unexpected patterns of microbial co-occurrence across domains. The diversity of microeukaryotes we recovered is higher than that previously reported from other wild frogs: we recovered 845 OTUs in our study compared with e.g. , 255 OTUs on Rana cascadae (16). In contrast, the level of bacterial diversity we recovered is lower than has been previously reported: in our study we recovered 303 bacterial OTUs compared with ~600 OTUs on Rana italica in Federici et al. 2015). However, our recovery of bacteria from 11 phyla is within the range of taxonomic diversity previously recovered from amphibian skin, with for example 10-18 bacterial phyla reported from three species (55). Our analysis showed that total microeukaryotic and bacterial diversity were positively correlated across all samples, which is a novel finding to our knowledge. It seems unlikely that this pattern is an artifact of sequencing: different MiSeq runs (and different research facilities) were used to sequence microeukaryotes and bacteria.
Proteobacteria were the most dominant bacterial phylum on T. taophora skin across all study populations, in terms of both OTUs and reads (Fig. 3). This is similar to findings from bacterial microbiome studies of other tropical post-metamorphic anurans (56–59). One hypothesis for the dominance of Proteobacteria on frog skin is that many members of the Proteobacteria produce anti-Bd metabolites (60,61). The presence of a high number of Proteobacteria on T. taophora skin could hypothetically contribute to the low apparent susceptibility to Bd previously observed in this species (17). It is important to note however that the present study is correlative; without experimental manipulations it is difficult to pinpoint which factors (e.g. , the physiology of the skin, mucosal biochemistry, host-microbial evolutionary processes, or interactions with the saline coastal environment) are responsible for the overwhelming dominance of Proteobacteria on anuran skin.
Although bacteria were less diverse than microeukaryotes in our samples, bacteria could nevertheless dominate the skin microbiome according to microbial biomass, which we did not quantify in our study. Sequence reads are sometimes used as a proxy for relative abundance, but this has been shown to be an unreliable measure due to known sequencing biases among microbial taxa (62). It is possible that taxa representing fewer OTUs (i.e. , bacteria) represent a higher proportion of microbial biomass, and this should be considered in interpretations of our results. Future research to address the relationship between microbial diversity and abundance could utilize high-throughput sequencing alongside quantitative analyses, for example quantitative PCR.