Low-level pathogen infection and geographic location correlate with the skin microbiomes of Columbia spotted frogs (Rana luteiventris) in a montane landscape

The skin microbiome of amphibians can influence host susceptibility towards the fungal pathogen Batrachochytrium dendrobatidis (Bd), while simultaneously having the potential to be altered by Bd. Severe Bd infections are known to alter the amphibian skin microbiome; however, little is known about microbiome interactions in amphibians with low infection intensity. In addition to disease dynamics, environmental factors may influence the microbiome. To test for patterns in bacterial diversity based on pathogen infection and environmental factors, 399 Columbia spotted frogs (Rana luteiventris) were sampled throughout northern Idaho and northeastern Washington across two years. Bd prevalence and intensity were measured in 376 frogs, revealing a prevalence of 69%, but generally low infection intensity (Mean = 127 Bd zoospore equivalents among infected frogs). Skin bacterial communities were characterized in 92 frogs using 16S rRNA gene amplicon sequencing. Our results indicated correlations of decreasing Shannon diversity and evenness as infection intensity increased. Latitude was correlated with bacterial richness and Faith's Phylogenetic Diversity measures, indicating increased diversity in northern locations. Beta diversity (UniFrac) analyses revealed that skin microbiomes were distinct between infected and uninfected frogs, and infection intensity had a significant effect on microbiome composition. Site explained the majority of microbiome variation (weighted UniFrac: 57.5%), suggesting a combination of local habitat conditions explain variation, as only small proportions of variation could be explained by year, month, temperature, elevation, and latitude individually. Bacterial genera with potential for Bd-inhibitory properties were found with differential relative abundance in infected and uninfected frogs, with higher Stenotrophomonas and lower Pseudomonas relative abundance observed in infected frogs. Further study may indicate if Bd inhibition by members of the skin microbiome is an influence behind the low infection intensities observed and whether low Bd infection intensities are capable of altering skin microbiome composition.