Phylogenetic distribution of symbiotic bacteria from Panamanian amphibians that inhibit growth of the lethal fungal pathogen Batrachochytrium dendrobatidis

Abstract

The introduction of next-generation sequencing has allowed for greater understanding of community composition of symbiotic microbial communities. However, determining the function of individual members of these microbial communities still largely relies on culture-based methods. Here, we present results on the phylogenetic distribution of a defensive functional trait of cultured symbiotic bacteria associated with amphibians. Amphibians are host to a diverse community of cutaneous bacteria and some of these bacteria protect their host from the lethal fungal pathogen Batrachochytrium dendrobatidis (Bd) by secreting antifungal metabolites. We cultured over 450 bacterial isolates from the skins of Panamanian amphibian species and tested their interactions with Bd using an in vitro challenge assay. For a subset of isolates, we also completed coculture experiments and found that culturing isolates with Bd had no effect on inhibitory properties of the bacteria, but it significantly decreased metabolite secretion. In challenge assays, approximately 75% of the bacterial isolates inhibited Bd to some extent and these inhibitory isolates were widely distributed among all bacterial phyla. Although there was no clear phylogenetic signal of inhibition, three genera, Stenotrophomonas, Aeromonas and Pseudomonas, had a high proportion of inhibitory isolates (100%, 77% and 73%, respectively). Overall, our results demonstrate that antifungal properties are phylogenetically widespread in symbiotic microbial communities of Panamanian amphibians and that some functional redundancy for fungal inhibition occurs in these communities. We hope that these findings contribute to the discovery and development of probiotics for amphibians that can mitigate the threat of chytridiomycosis.

Antifungal Bacteria on Woodland Salamander Skin Exhibit High Taxonomic Diversity and Geographic Variability

“…dendrobatidis bacterial species, chiefly Pseudomonas and Stenotrophomonas, and detection of bacterial sequences in those genera likely indicates that many of those species have anti-B. dendrobatidis traits (94). Two possible, nonexclusive mechanisms may explain the lack of phylogenetic conservatism of B. dendrobatidis inhibition: (i) genes involved in inhibition are widely distributed among bacterial phylogenies, and (ii) horizontal gene transfer of antifungal genes may occur among unrelated bacteria.…”

Composition of symbiotic bacteria predicts survival in Panamanian golden frogs infected with a lethal fungus

“…For example, commensal bacteria that reside in the human gut competitively exclude pathogenic bacteria by producing antibiotics, preventing attachment to epithelial cells, competing for resources and stimulating the host's immune system [21,55]. In this study, many indicator OTUs which were associated with frogs that cleared infection belong to bacterial families that inhibit Bd in vitro and have been isolated from non-susceptible Panamanian amphibians (Flavobacteriaceae, Comamonadaceae, Pseudomonadaceae; [26]). For example, a Comamonadaceae indicator OTU (figure 4) had a 98% sequence similarity (16S rRNA gene) to four cultured isolates that inhibit Bd growth [26].…”

“…In this study, many indicator OTUs which were associated with frogs that cleared infection belong to bacterial families that inhibit Bd in vitro and have been isolated from non-susceptible Panamanian amphibians (Flavobacteriaceae, Comamonadaceae, Pseudomonadaceae; [26]). For example, a Comamonadaceae indicator OTU (figure 4) had a 98% sequence similarity (16S rRNA gene) to four cultured isolates that inhibit Bd growth [26]. Bacteria belonging to these families have also been detected from skin swabs collected from a captive population of A. zeteki housed at the National Zoo and from a wild population of A. zeteki in Panama [33].…”

“…Probiotic candidates were chosen from 484 bacteria isolated from skin swabs of 11 Panamanian frog species (67 individuals) sampled from January to April 2010 [26]. We tested each isolate's ability to inhibit Bd with a spectrophotometric assay (similar to Bell et al [27]).…”

Dietary Carotenoid Supplementation Enhances the Cutaneous Bacterial Communities of the Critically Endangered Southern Corroboree Frog (Pseudophryne corroboree)

“…Amphibians are known to support a diverse array of microbial symbionts on their skin surface that help mediate pathogen colonisation [ 5,6 ]. In particular, Bd infectivity and virulence is inhibited by antifungal metabolites, such as violacein, which are produced by certain bacteria species [ 7 ].…”