Resource competition drives an invasion-replacement event among shrew species on an island.

Invasive mammals are responsible for the majority of native species extinctions on islands. While most of these extinction events will be due to novel interactions between species (e.g. exotic predators and naive prey), it is more unusual to find incidences where a newly invasive species causes the decline/extinction of a native species on an island when they normally coexist elsewhere in their overlapping mainland ranges. We investigated if resource competition between two insectivorous small mammals was playing a significant role in the rapid replacement of the native pygmy shrew Sorex minutus in the presence of the recently invading greater white-toothed shrew Crocidura russula on the island of Ireland. We used DNA metabarcoding of gut contents from >300 individuals of both species to determine each species' diet and measured the body size (weight and length) during different stages of the invasion in Ireland (before, during and after the species come into contact with one another) and on a French island where both species have long coexisted (acting as a natural 'control' site). Dietary composition, niche width and overlap and body size were compared in these different stages. The body size of the invasive C. russula and composition of its diet changes between when it first invades an area and after it becomes established. During the initial stages of the invasion, individual shrews are larger and consume larger sized invertebrate prey species. During later stages of the invasion, C. russula switches to consuming smaller prey taxa that are more essential for the native species. As a result, the level of interspecific dietary overlap increases from between 11% and 14% when they first come into contact with each other to between 39% and 46% after the invasion. Here we show that an invasive species can quickly alter its dietary niche in a new environment, ultimately causing the replacement of a native species. In addition, the invasive shrew could also be potentially exhausting local resources of larger invertebrate species. These subsequent changes in terrestrial invertebrate communities could have severe impacts further downstream on ecosystem functioning and services.

Some observations on meaningful and objective inference in radioecological field studies.

Anthropogenic releases of radiation are of ongoing importance for environmental protection, but the radiation doses at which natural systems begin to show effects are controversial. More certainty is required in this area to achieve optimal regulation for radioactive substances. We recently carried out a large survey (268 sampled animals and 20 sites) of the association between environmental radiation exposures and small mammal gut-associated microbiomes (fungal and bacterial) in the Chornobyl Exclusion zone (CEZ). Using individual measurements of total absorbed dose rates and a study design and analyses that accounted for spatial non-independence, we found no, or only limited, association. Watts et al. have criticised our study: for not filtering candidate non-resident components prior to our fungal microbiome analyses, for our qualified speculations on the relative merits of faecal and gut samples, and for the design of our study which they felt lacked sufficient replication. The advantage of filtering non-resident-fungal taxa is not clear and it would not have changed the null (spatially adjusted) association we found between radioactive dose and mycobiome composition because the most discriminatory fungal taxa with regard to dose were non-resident taxa. We maintain that it was legitimate for us to make qualified discussion comments on the differences in results between our faecal and gut microbiome analyses and on the relative merits of these sample types. Most importantly, the criticism of our study design by Watts et al. and the designs and analysis of their recent studies in the CEZ show a misunderstanding of the true nature of independent replication in field studies. Recognising the importance of spatial non-independence is essential in the design and analysis of radioecological field surveys.

Evidence for the genetic similarity rule at an expanding mangrove range limit.

PREMISE: Host-plant genetic variation can shape associated communities of organisms. These community-genetic effects include (1) genetically similar hosts harboring similar associated communities (i.e., the genetic similarity rule) and (2) host-plant heterozygosity increasing associated community diversity. Community-genetic effects are predicted to be less prominent in plant systems with limited genetic variation, such as those at distributional range limits. Yet, empirical evidence from such systems is limited. METHODS: We sampled a natural population of a mangrove foundation species (Avicennia germinans) at an expanding range limit in Florida, USA. We measured genetic variation within and among 40 host trees with 24 nuclear microsatellite loci and characterized their foliar endophytic fungal communities with internal transcribed spacer (ITS1) gene amplicon sequencing. We evaluated relationships among host-tree genetic variation, host-tree spatial location, and the associated fungal communities. RESULTS: Genetic diversity was low across all host trees (mean: 2.6 alleles per locus) and associated fungal communities were relatively homogeneous (five sequence variants represented 78% of all reads). We found (1) genetically similar host trees harbored similar fungal communities, with no detectable effect of interhost geographic distance. (2) Host-tree heterozygosity had no detectable effect, while host-tree absolute spatial location affected community alpha diversity. CONCLUSIONS: This research supports the genetic similarity rule within a range limit population and helps broaden the current scope of community genetics theory by demonstrating that community-genetic effects can occur even at expanding distributional limits where host-plant genetic variation may be limited. Our findings also provide the first documentation of community-genetic effects in a natural mangrove system.

Fungal microbiomes are determined by host phylogeny and exhibit widespread associations with the bacterial microbiome.

Interactions between hosts and their resident microbial communities are a fundamental component of fitness for both agents. Though recent research has highlighted the importance of interactions between animals and their bacterial communities, comparative evidence for fungi is lacking, especially in natural populations. Using data from 49 species, we present novel evidence of strong covariation between fungal and bacterial communities across the host phylogeny, indicative of recruitment by hosts for specific suites of microbes. Using co-occurrence networks, we demonstrate marked variation across host taxonomy in patterns of covariation between bacterial and fungal abundances. Host phylogeny drives differences in the overall richness of bacterial and fungal communities, but the effect of diet on richness was only evident in the mammalian gut microbiome. Sample type, tissue storage and DNA extraction method also affected bacterial and fungal community composition, and future studies would benefit from standardized approaches to sample processing. Collectively these data indicate fungal microbiomes may play a key role in host fitness and suggest an urgent need to study multiple agents of the animal microbiome to accurately determine the strength and ecological significance of host-microbe interactions.

Exposure to airborne bacteria depends upon vertical stratification and vegetation complexity.

Exposure to biodiverse aerobiomes supports human health, but it is unclear which ecological factors influence exposure. Few studies have investigated near-surface green space aerobiome dynamics, and no studies have reported aerobiome vertical stratification in different urban green spaces. We used columnar sampling and next generation sequencing of the bacterial 16S rRNA gene, combined with geospatial and network analyses to investigate urban green space aerobiome spatio-compositional dynamics. We show a strong effect of habitat on bacterial diversity and network complexity. We observed aerobiome vertical stratification and network complexity that was contingent on habitat type. Tree density, closer proximity, and canopy coverage associated with greater aerobiome alpha diversity. Grassland aerobiomes exhibited greater proportions of putative pathogens compared to scrub, and also stratified vertically. We provide novel insights into the urban ecosystem with potential importance for public health, whereby the possibility of differential aerobiome exposures appears to depend on habitat type and height in the airspace. This has important implications for managing urban landscapes for the regulation of aerobiome exposure.

Impacts of radiation exposure on the bacterial and fungal microbiome of small mammals in the Chernobyl Exclusion Zone.

Environmental impacts of the 1986 Chernobyl Nuclear Power Plant accident are much debated, but the effects of radiation on host microbiomes have received little attention to date. We present the first analysis of small mammal gut microbiomes from the Chernobyl Exclusion Zone in relation to total absorbed dose rate, including both caecum and faeces samples. We provide novel evidence that host species determines fungal community composition, and that associations between microbiome (both bacterial and fungal) communities and radiation exposure vary between host species. Using ambient versus total weighted absorbed dose rates in analyses produced different results, with the latter more robust for interpreting microbiome changes at the individual level. We found considerable variation between results for faecal and gut samples of bank voles, suggesting faecal samples are not an accurate indicator of gut composition. Associations between radiation exposure and microbiome composition of gut samples were not robust against geographical variation, although we identified families of bacteria (Lachnospiraceae and Muribaculaceae) and fungi (Steccherinaceae and Strophariaceae) in the guts of bank voles that may serve as biomarkers of radiation exposure. Further studies considering a range of small mammal species are needed to establish the robustness of these potential biomarkers.

Mixed-Cropping Between Field Pea Varieties Alters Root Bacterial and Fungal Communities.

Modern agricultural practices have vastly increased crop production but negatively affected soil health. As such, there is a call to develop sustainable, ecologically-viable approaches to food production. Mixed-cropping of plant varieties can increase yields, although impacts on plant-associated microbial communities are unclear, despite their critical role in plant health and broader ecosystem function. We investigated how mixed-cropping between two field pea (Pisum sativum L.) varieties (Winfreda and Ambassador) influenced root-associated microbial communities and yield. The two varieties supported significantly different fungal and bacterial communities when grown as mono-crops. Mixed-cropping caused changes in microbial communities but with differences between varieties. Root bacterial communities of Winfreda remained stable in response to mixed-cropping, whereas those of Ambassador became more similar to Winfreda. Conversely, root fungal communities of Ambassador remained stable under mixed-cropping, and those of Winfreda shifted towards the composition of Ambassador. Microbial co-occurrence networks of both varieties were stronger and larger under mixed-cropping, which may improve stability and resilience in agricultural soils. Both varieties produced slightly higher yields under mixed-cropping, although overall Ambassador plants produced higher yields than Winfreda plants. Our results suggest that variety diversification may increase yield and promote microbial interactions.

Host genetics and geography influence microbiome composition in the sponge Ircinia campana.

Marine sponges are hosts to large, diverse communities of microorganisms. These microbiomes are distinct among sponge species and from seawater bacterial communities, indicating a key role of host identity in shaping its resident microbial community. However, the factors governing intraspecific microbiome variability are underexplored and may shed light on the evolutionary and ecological relationships between host and microbiome. Here, we examined the influence of genetic variation and geographic location on the composition of the Ircinia campana microbiome. We developed new microsatellite markers to genotype I. campana from two locations in the Florida Keys, USA, and characterized their microbiomes using V4 16S rRNA amplicon sequencing. We show that microbial community composition and diversity is influenced by host genotype, with more genetically similar sponges hosting more similar microbial communities. We also found that although I. campana was not genetically differentiated between sites, microbiome composition differed by location. Our results demonstrate that both host genetics and geography influence the composition of the sponge microbiome. Host genotypic influence on microbiome composition may be due to stable vertical transmission of the microbial community from parent to offspring, making microbiomes more similar by descent. Alternatively, sponge genotypic variation may reflect variation in functional traits that influence the acquisition of environmental microbes. This study reveals drivers of microbiome variation within and among locations, and shows the importance of intraspecific variability in mediating eco-evolutionary dynamics of host-associated microbiomes.

Multi-individual microsatellite identification: A multiple genome approach to microsatellite design (MiMi).

Bespoke microsatellite marker panels are increasingly affordable and tractable to researchers and conservationists. The rate of microsatellite discovery is very high within a shotgun genomic data set, but extensive laboratory testing of markers is required for confirmation of amplification and polymorphism. By incorporating shotgun next-generation sequencing data sets from multiple individuals of the same species, we have developed a new method for the optimal design of microsatellite markers. This new tool allows us to increase the rate at which suitable candidate markers are selected by 58% in direct comparisons and facilitate an estimated 16% reduction in costs associated with producing a novel microsatellite panel. Our method enables the visualisation of each microsatellite locus in a multiple sequence alignment allowing several important quality checks to be made. Polymorphic loci can be identified and prioritised. Loci containing fragment-length-altering mutations in the flanking regions, which may invalidate assumptions regarding the model of evolution underlying variation at the microsatellite, can be avoided. Priming regions containing point mutations can be detected and avoided, helping to reduce sample-site-marker specificity arising from genetic isolation, and the likelihood of null alleles occurring. We demonstrate the utility of this new approach in two species: an echinoderm and a bird. Our method makes a valuable contribution towards minimising genotyping errors and reducing costs associated with developing a novel marker panel. The Python script to perform our method of multi-individual microsatellite identification (MiMi) is freely available from GitHub (https://github.com/graemefox/mimi).

Rare gut microbiota associated with breeding success, hormone metabolites and ovarian cycle phase in the critically endangered eastern black rhino.

BACKGROUND: Host microbiomes play a role in hormone production and subsequent fertility in humans, but this is less well understood in non-model organisms. This is of particular relevance to species in zoo-based conservation breeding programmes, as relationships between host microbiome composition and reproductive output may allow for the development of microbial augmentation strategies to improve success. Here, we characterise faecal bacterial communities of breeding and non-breeding eastern black rhino (Diceros bicornis michaeli) using 16S rRNA gene amplicon sequencing and quantify progestagen and glucocorticoid metabolite concentrations through enzyme immunoassays to identify such relationships. RESULTS: We identified significant differences in black rhino gut microbiome composition according to ID, institution, breeding success and ovarian cycle phase. In particular, the gut microbiome during pregnancy and post-parturition was significantly altered. Around a third of bacterial genera showed more than ± 10% correlation with either progestagen and/or glucocorticoid concentration, and in general, microbial genera correlated with both hormones in the same direction. Through a combination of analyses, we identified four genera (Aerococcaceae, Atopostipes, Carnobacteriaceae and Solobacterium) that were significantly associated with breeding success, pregnancy and/or post-parturition, and higher faecal progestagen metabolite concentrations. These genera had a lower-than-average relative abundance in the gut microbiome. CONCLUSION: Our results indicate that many members of the gut microbiome of black rhino are associated with hormone production and breeding success, and some members of the rare microbiota appear to be particularly important. Although the directionality of the relationship is unclear, the variation in gut microbiome communities represents a potential biomarker of reproductive health. We identified four genera that were associated with multiple indicators of reproductive output; these could be candidate probiotics to improve the breeding success of black rhino in zoo-based conservation breeding programmes. Further work is required to understand the efficacy and feasibility of this, either directly through microbial augmentation (e.g. probiotics) or indirectly via dietary manipulation or prebiotics.

Designing Probiotic Therapies With Broad-Spectrum Activity Against a Wildlife Pathogen.

Host-associated microbes form an important component of immunity that protect against infection by pathogens. Treating wild individuals with these protective microbes, known as probiotics, can reduce rates of infection and disease in both wild and captive settings. However, the utility of probiotics for tackling wildlife disease requires that they offer consistent protection across the broad genomic variation of the pathogen that hosts can encounter in natural settings. Here we develop multi-isolate probiotic consortia with the aim of effecting broad-spectrum inhibition of growth of the lethal amphibian pathogen Batrachochytrium dendrobatidis (Bd) when tested against nine Bd isolates from two distinct lineages. Though we achieved strong growth inhibition between 70 and 100% for seven Bd isolates, two isolates appeared consistently resistant to inhibition, irrespective of probiotic strategy employed. We found no evidence that genomic relatedness of the chytrid predicted similarity of inhibition scores, nor that increasing the genetic diversity of the bacterial consortia could offer stronger inhibition of pathogen growth, even for the two resistant isolates. Our findings have important consequences for the application of probiotics to mitigate wildlife diseases in the face of extensive pathogen genomic variation.

Gut microbiome composition is associated with spatial structuring and social interactions in semi-feral Welsh Mountain ponies.

BACKGROUND: Microbiome composition is linked to host functional traits including metabolism and immune function. Drivers of microbiome composition are increasingly well-characterised; however, evidence of group-level microbiome convergence is limited and may represent a multi-level trait (i.e. across individuals and groups), whereby heritable phenotypes are influenced by social interactions. Here, we investigate the influence of spatial structuring and social interactions on the gut microbiome composition of Welsh mountain ponies. RESULTS: We show that semi-feral ponies exhibit variation in microbiome composition according to band (group) membership, in addition to considerable within-individual variation. Spatial structuring was also identified within bands, suggesting that despite communal living, social behaviours still influence microbiome composition. Indeed, we show that specific interactions (i.e. mother-offspring and stallion-mare) lead to more similar microbiomes, further supporting the notion that individuals influence the microbiome composition of one another and ultimately the group. Foals exhibited different microbiome composition to sub-adults and adults, most likely related to differences in diet. CONCLUSIONS: We provide novel evidence that microbiome composition is structured at multiple levels within populations of social mammals and thus may form a unit on which selection can act. High levels of within-individual variation in microbiome composition, combined with the potential for social interactions to influence microbiome composition, suggest the direction of microbiome selection may be influenced by the individual members present in the group. Although the functional implications of this require further research, these results lend support to the idea that multi-level selection can act on microbiomes.

Genetic variability and ontogeny predict microbiome structure in a disease-challenged montane amphibian.

Amphibian populations worldwide are at risk of extinction from infectious diseases, including chytridiomycosis caused by the fungal pathogen Batrachochytrium dendrobatidis (Bd). Amphibian cutaneous microbiomes interact with Bd and can confer protective benefits to the host. The composition of the microbiome itself is influenced by many environment- and host-related factors. However, little is known about the interacting effects of host population structure, genetic variation and developmental stage on microbiome composition and Bd prevalence across multiple sites. Here we explore these questions in Amietia hymenopus, a disease-affected frog in southern Africa. We use microsatellite genotyping and 16S amplicon sequencing to show that the microbiome associated with tadpole mouthparts is structured spatially, and is influenced by host genotype and developmental stage. We observed strong genetic structure in host populations based on rivers and geographic distances, but this did not correspond to spatial patterns in microbiome composition. These results indicate that demographic and host genetic factors affect microbiome composition within sites, but different factors are responsible for host population structure and microbiome structure at the between-site level. Our results help to elucidate complex within- and among- population drivers of microbiome structure in amphibian populations. That there is a genetic basis to microbiome composition in amphibians could help to inform amphibian conservation efforts against infectious diseases.

Probiotic consortia are not uniformly effective against different amphibian chytrid pathogen isolates.

Symbiotic bacterial communities can protect their hosts from infection by pathogens. Treatment of wild individuals with protective bacteria (probiotics) isolated from hosts can combat the spread of emerging infectious diseases. However, it is unclear whether candidate probiotic bacteria can offer consistent protection across multiple isolates of globally distributed pathogens. Here, we use the lethal amphibian fungal pathogen Batrachochytrium dendrobatidis to investigate whether probiotic richness (number of bacteria) or genetic distance among consortia members influences broad-scale in vitro inhibitory capabilities of probiotics across multiple isolates of the pathogen. We show that inhibition of multiple pathogen isolates by individual bacteria is rare, with no systematic pattern among bacterial genera in ability to inhibit multiple B. dendrobatidis isolates. Bacterial consortia can offer stronger protection against B. dendrobatidis compared to single strains, and this tended to be more pronounced for consortia containing multiple genera compared with those consisting of bacteria from a single genus (i.e., with lower genetic distance), but critically, this effect was not uniform across all B. dendrobatidis isolates. These novel insights have important implications for the effective design of bacterial probiotics to mitigate emerging infectious diseases.

Fifty important research questions in microbial ecology.

Microbial ecology provides insights into the ecological and evolutionary dynamics of microbial communities underpinning every ecosystem on Earth. Microbial communities can now be investigated in unprecedented detail, although there is still a wealth of open questions to be tackled. Here we identify 50 research questions of fundamental importance to the science or application of microbial ecology, with the intention of summarising the field and bringing focus to new research avenues. Questions are categorised into seven themes: host-microbiome interactions; health and infectious diseases; human health and food security; microbial ecology in a changing world; environmental processes; functional diversity; and evolutionary processes. Many questions recognise that microbes provide an extraordinary array of functional diversity that can be harnessed to solve real-world problems. Our limited knowledge of spatial and temporal variation in microbial diversity and function is also reflected, as is the need to integrate micro- and macro-ecological concepts, and knowledge derived from studies with humans and other diverse organisms. Although not exhaustive, the questions presented are intended to stimulate discussion and provide focus for researchers, funders and policy makers, informing the future research agenda in microbial ecology.

Using "Omics" and Integrated Multi-Omics Approaches to Guide Probiotic Selection to Mitigate Chytridiomycosis and Other Emerging Infectious Diseases.

Emerging infectious diseases in wildlife are responsible for massive population declines. In amphibians, chytridiomycosis caused by Batrachochytrium dendrobatidis, Bd, has severely affected many amphibian populations and species around the world. One promising management strategy is probiotic bioaugmentation of antifungal bacteria on amphibian skin. In vivo experimental trials using bioaugmentation strategies have had mixed results, and therefore a more informed strategy is needed to select successful probiotic candidates. Metagenomic, transcriptomic, and metabolomic methods, colloquially called "omics," are approaches that can better inform probiotic selection and optimize selection protocols. The integration of multiple omic data using bioinformatic and statistical tools and in silico models that link bacterial community structure with bacterial defensive function can allow the identification of species involved in pathogen inhibition. We recommend using 16S rRNA gene amplicon sequencing and methods such as indicator species analysis, the Kolmogorov-Smirnov Measure, and co-occurrence networks to identify bacteria that are associated with pathogen resistance in field surveys and experimental trials. In addition to 16S amplicon sequencing, we recommend approaches that give insight into symbiont function such as shotgun metagenomics, metatranscriptomics, or metabolomics to maximize the probability of finding effective probiotic candidates, which can then be isolated in culture and tested in persistence and clinical trials. An effective mitigation strategy to ameliorate chytridiomycosis and other emerging infectious diseases is necessary; the advancement of omic methods and the integration of multiple omic data provide a promising avenue toward conservation of imperiled species.

Amphibian Symbiotic Bacteria Do Not Show a Universal Ability To Inhibit Growth of the Global Panzootic Lineage of Batrachochytrium dendrobatidis.

Microbiomes associated with multicellular organisms influence the disease susceptibility of hosts. The potential exists for such bacteria to protect wildlife from infectious diseases, particularly in the case of the globally distributed and highly virulent fungal pathogen Batrachochytrium dendrobatidis of the global panzootic lineage (B. dendrobatidis GPL), responsible for mass extinctions and population declines of amphibians. B. dendrobatidis GPL exhibits wide genotypic and virulence variation, and the ability of candidate probiotics to restrict growth across B. dendrobatidis isolates has not previously been considered. Here we show that only a small proportion of candidate probiotics exhibited broad-spectrum inhibition across B. dendrobatidis GPL isolates. Moreover, some bacterial genera showed significantly greater inhibition than others, but overall, genus and species were not particularly reliable predictors of inhibitory capabilities. These findings indicate that bacterial consortia are likely to offer a more stable and effective approach to probiotics, particularly if related bacteria are selected from genera with greater antimicrobial capabilities. Together these results highlight a complex interaction between pathogens and host-associated symbiotic bacteria that will require consideration in the development of bacterial probiotics for wildlife conservation. Future efforts to construct protective microbiomes should incorporate bacteria that exhibit broad-spectrum inhibition of B. dendrobatidis GPL isolates.

Tagging frogs with passive integrated transponders causes disruption of the cutaneous bacterial community and proliferation of opportunistic fungi.

Symbiotic bacterial communities play a key role in protecting amphibians from infectious diseases including chytridiomycosis, caused by the pathogenic fungus Batrachochytrium dendrobatidis. Events that lead to the disruption of the bacterial community may have implications for the susceptibility of amphibians to such diseases. Amphibians are often marked both in the wild and in captivity for a variety of reasons, and although existing literature indicates that marking techniques have few negative effects, the response of cutaneous microbial communities has not yet been investigated. Here we determine the effects of passive integrated transponder (PIT) tagging on culturable cutaneous microbial communities of captive Morelet's tree frogs (Agalychnis moreletii) and assess the isolated bacterial strains for anti-B. dendrobatidis activity in vitro. We find that PIT tagging causes a major disruption to the bacterial community associated with the skin of frogs (∼12-fold increase in abundance), as well as a concurrent proliferation in resident fungi (up to ∼200-fold increase). Handling also caused a disruption the bacterial community, although to a lesser extent than PIT tagging. However, the effects of both tagging and handling were temporary, and after 2 weeks, the bacterial communities were similar to their original compositions. We also identify two bacterial strains that inhibit B. dendrobatidis, one of which increased in abundance on PIT-tagged frogs at 1 day postmarking, while the other was unaffected. These results show that PIT tagging has previously unobserved consequences for cutaneous microbial communities of frogs and may be particularly relevant for studies that intend to use PIT tagging to identify individuals involved in trials to develop probiotic treatments.

Impact of plant cover on fitness and behavioural traits of captive red-eyed tree frogs (Agalychnis callidryas).

Despite the importance of ex situ conservation programmes as highlighted in the Amphibian Conservation Action Plan, there are few empirical studies that examine the influence of captive conditions on the fitness of amphibians, even for basic components of enclosure design such as cover provision. Maintaining the fitness of captive amphibian populations is essential to the success of ex situ conservation projects. Here we examined the impact of plant cover on measures of fitness and behaviour in captive red-eyed tree frogs (Agalychnis callidryas). We found significant effects of plant provision on body size, growth rates and cutaneous bacterial communities that together demonstrate a compelling fitness benefit from cover provision. We also demonstrate a strong behavioural preference for planted rather than non-planted areas. We also assessed the impact of plant provision on the abiotic environment in the enclosure as a potential driver of these behavioural and fitness effects. Together this data provides valuable information regarding enclosure design for a non-model amphibian species and has implications for amphibian populations maintained in captivity for conservation breeding programmes and research.

Ex situ diet influences the bacterial community associated with the skin of red-eyed tree frogs (Agalychnis callidryas).

Amphibians support symbiotic bacterial communities on their skin that protect against a range of infectious pathogens, including the amphibian chytrid fungus. The conditions under which amphibians are maintained in captivity (e.g. diet, substrate, enrichment) in ex situ conservation programmes may affect the composition of the bacterial community. In addition, ex situ amphibian populations may support different bacterial communities in comparison to in situ populations of the same species. This could have implications for the suitability of populations intended for reintroduction, as well as the success of probiotic bacterial inoculations intended to provide amphibians with a bacterial community that resists invasion by the chytrid fungus. We aimed to investigate the effect of a carotenoid-enriched diet on the culturable bacterial community associated with captive red-eyed tree frogs (Agalychnis callidryas) and make comparisons to bacteria isolated from a wild population from the Chiquibul Rainforest in Belize. We successfully showed carotenoid availability influences the overall community composition, species richness and abundance of the bacterial community associated with the skin of captive frogs, with A. callidryas fed a carotenoid-enriched diet supporting a greater species richness and abundance of bacteria than those fed a carotenoid-free diet. Our results suggest that availability of carotenoids in the diet of captive frogs is likely to be beneficial for the bacterial community associated with the skin. We also found wild A. callidryas hosted more than double the number of different bacterial species than captive frogs with very little commonality between species. This suggests frogs in captivity may support a reduced and diverged bacterial community in comparison to wild populations of the same species, which could have particular relevance for ex situ conservation projects.