Host-associated microbiomes play important roles in host health and pathogen defense. In amphibians, the skin-associated microbiota can contribute to innate immunity with potential implications for disease management. Few studies have examined season-long temporal variation in the amphibian skin-associated microbiome, and the interactions between bacteria and fungi on amphibian skin remain poorly understood. We characterize season-long temporal variation in the skin-associated microbiome of the western tiger salamander ( Ambystoma mavortium ) for both bacteria and fungi between sites and across salamander life stages. Two hundred seven skin-associated microbiome samples were collected from salamanders at two Rocky Mountain lakes throughout the summer and fall of 2018, and 127 additional microbiome samples were collected from lake water and lake substrate. We used 16S rRNA and ITS amplicon sequencing with Bayesian Dirichlet-multinomial regression to estimate the relative abundances of bacterial and fungal taxa, test for differential abundance, examine microbial selection, and derive alpha diversity. We predicted the ability of bacterial communities to inhibit the amphibian chytrid fungus Batrachochytrium dendrobatidis ( Bd ), a cutaneous fungal pathogen, using stochastic character mapping and a database of Bd -inhibitory bacterial isolates. For both bacteria and fungi, we observed variation in community composition through time, between sites, and with salamander age and life stage. We further found that temporal trends in community composition were specific to each combination of salamander age, life stage, and lake. We found salamander skin to be selective for microbes, with many taxa disproportionately represented relative to the environment. Salamander skin appeared to select for predicted Bd -inhibitory bacteria, and we found a negative relationship between the relative abundances of predicted Bd -inhibitory bacteria and Bd . We hope these findings will assist in the conservation of amphibian species threatened by chytridiomycosis and other emerging diseases.
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Longitudinal patterns in the skin microbiome of wild, individually marked frogs from the Sierra Nevada, California
The amphibian skin microbiome has been the focus of numerous studies because of the protective effects that some bacteria provide against the pathogenic fungus Batrachochytrium dendrobatidis, which has caused a global panzootic among amphibians. However, the mechanisms driving community structure and function in the amphibian skin microbiome are still poorly understood, and longitudinal analyses of the skin microbiome have not yet been conducted in wild populations. In this study, we investigate longitudinal patterns in the skin microbiome of 19 individually marked adult frogs from two wild populations of the endangered Sierra Nevada yellow-legged frog (Rana sierrae), sampled over the course of 2 years. We found that individuals with low bacterial diversity (dominated by order Burkhorderiales) had significantly more stable bacterial communities than those with higher diversity. Amplicon sequence variants (ASVs) with high relative abundance were significantly less transient than those with low relative abundance, and ASVs with intermediate-level relative abundances experienced the greatest volatility over time. Based on these results, we suggest that efforts to develop probiotic treatments to combat B. dendrobatidis should focus on bacteria that are found at high relative abundances in some members of a population, as these strains are more likely to persist and remain stable in the long term.
more » « less- Award ID(s):
- 1633948
- NSF-PAR ID:
- 10384517
- Publisher / Repository:
- Oxford University Press
- Date Published:
- Journal Name:
- ISME Communications
- Volume:
- 1
- Issue:
- 1
- ISSN:
- 2730-6151
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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Abstract Chytridiomycosis, caused by the pathogen Batrachochytrium dendrobatidis (Bd), has led to population declines and extinctions of frog species around the world. While it is known that symbiotic skin bacteria can play a protective role against pathogens, it is not known how these defensive bacteria are integrated into the bacterial community on amphibian skin. In this study, we used 16S rRNA gene amplicon sequencing, culturing and Bd inhibition bioassays to characterize the communities of skin bacteria on three Neotropical frog species that persist in a Bd-infected area in Panama and determined the abundance and integration of anti-Bd bacteria into the community. We found that the two treefrog species had a similar bacterial community structure, which differed from the more diverse community found on the terrestrial frog. Co-occurrence networks also revealed differences between frog species such that the treefrogs had a significantly higher number of culturable Bd-inhibitory OTUs with high centrality scores compared with the terrestrial frog. We found that culture-dependent OTUs captured between 21 and 39% of the total relative abundance revealed in culture-independent communities. Our results suggest different ecological strategies occurring within skin antifungal communities on host species that have not succumbed to Bd infections in the wild.
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Abstract Accurately predicting the impacts of climate change on wildlife health requires a deeper understanding of seasonal rhythms in host–pathogen interactions. The amphibian pathogen,
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Rana [Lithobates ]sphenocephala ) in Tennessee, United States, were surveyed for host immunity, microbiome and pathogen dynamics. Frogs were swabbed for pathogen load and skin bacterial diversity and stimulated to release stored antimicrobial peptides (AMPs). Secretions were analysed to estimate total hydrophobic peptide concentrations, presence of known AMPs and effectiveness ofBd growth inhibition in vitro. The diversity and proportion of bacterial reads with a 99% match to sequences of isolates known to inhibitBd growth in vitro were used as an estimate of predicted anti‐Bd function of the skin microbiome.Batrachochytrium dendrobatidis dynamics followed the expected seasonal fluctuations—peaks in cooler months—which coincided with when host mucosal defences were most potent againstBd . Specifically, the concentration and expression of stored AMPs cycled synchronously withBd dynamics. Although microbiome changes followed more linear trends over time, the proportion of bacteria that can function to inhibitBd growth was greatest when risk ofBd infection was highest.We interpret the increase in peptide storage in the fall and the shift to a more anti‐
Bd microbiome over winter as a preparatory response for subsequent infection risk during the colder periods when AMP synthesis and bacterial growth is slow and pathogen pressure from this cool‐adapted fungus is high. Given that a decrease in stored AMP concentrations as temperatures warm in spring likely means greater secretion rates, the subsequent decrease in prevalence suggests seasonality ofBd in this host may be in part regulated by annual immune rhythms, and dominated by the effects of temperature. -
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ABSTRACT Host-associated microbial communities can influence physiological processes of macroorganisms, including contributing to infectious disease resistance. For instance, some bacteria that live on amphibian skin produce antifungal compounds that inhibit two lethal fungal pathogens, Batrachochytrium dendrobatidis (Bd) and Batrachochytrium salamandrivorans (Bsal). Therefore, differences in microbiome composition among host species or populations within a species can contribute to variation in susceptibility to Bd/Bsal. This study applies 16S rRNA sequencing to characterize the skin bacterial microbiomes of three widespread terrestrial salamander genera native to the western United States. Using a metacommunity structure analysis, we identified dispersal barriers for these influential bacteria between salamander families and localities. We also analysed the effects of habitat characteristics such as percent natural cover and temperature seasonality on the microbiome. We found that certain environmental variables may influence the skin microbial communities of some salamander genera more strongly than others. Each salamander family had a somewhat distinct community of putative anti-Bd skin bacteria, suggesting that salamanders may select for a functional assembly of cutaneous symbionts that could differ in its ability to protect these amphibians from disease. Our observations raise the need to consider host identity and environmental heterogeneity during the selection of probiotics to treat wildlife diseases.
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