Chemical disinfection as a simple and reliable method to control the amphibian chytrid fungus at breeding points of endangered amphibians.

Chytridiomycosis caused by the fungal pathogen Batrachochytrium dendrobatidis (Bd) is pushing amphibians towards extinction. Whilst mitigation methods were suggested a decade ago, we lack field trials testing their efficacy. We used the agrochemical fungicide, tebuconazole, to treat Bd infected breeding waterbodies of an endangered species that is highly susceptible to the fungus. Just two applications of tebuconazole led to a significant reduction in infection loads in the vast majority of sites, and at six sites the disinfection remained one/two-years post-application. Tebuconazole values drastically decreased in the waterbodies within a week after application, with no significant effects on their hydrochemical and hydrobiological characteristics. Although the use of chemicals in natural populations is undesirable, the growing existential threat to amphibians all over the world indicates that effective interventions in selected populations of endangered species are urgently needed.

Gut microbiota of the critically endangered Saiga antelope across two wild populations in a year without mass mortality.

The Saiga are migratory antelopes inhabiting the grasslands of Eurasia. Over the last century, Saiga have been pushed to the brink of extinction by mass mortality events and intense poaching. Yet, despite the high profile of the Saiga as an animal of conservation concern, little is known of its biology. In particular, the gut microbiota of Saiga has not been studied, despite its potential importance in health. Here, we characterise the gut microbiota of Saiga from two geographically distinct populations in Kazakhstan and compare it with that of other antelope species. We identified a consistent gut microbial diversity and composition among individuals and across two Saiga populations during a year without die-offs, with over 85% of bacterial genera being common to both populations despite vast geographic separation. We further show that the Saiga gut microbiota resembled that of five other antelopes. The putative causative agent of Saiga mass die-offs, Pasteurella multocida, was not detected in the Saiga microbiota. Our findings provide the first description of the Saiga gut microbiota, generating a baseline for future work investigating the microbiota's role in health and mass die-offs, and supporting the conservation of this critically endangered species.

Turning the tide on sex and the microbiota in aquatic animals.

Sex-based differences in animal microbiota are increasingly recognized as of biological importance. While most animal biomass is found in aquatic ecosystems and many water-dwelling species are of high economic and ecological value, biological sex is rarely included as an explanatory variable in studies of the aquatic animal microbiota. In this opinion piece, we argue for greater consideration of host sex in studying the microbiota of aquatic animals, emphasizing the many advancements that this information could provide in the life sciences, from the evolution of sex to aquaculture.

Experimental temperatures shape host microbiome diversity and composition.

Global climate change has led to more extreme thermal events. Plants and animals harbour diverse microbial communities, which may be vital for their physiological performance and help them survive stressful climatic conditions. The extent to which microbiome communities change in response to warming or cooling may be important for predicting host performance under global change. Using a meta-analysis of 1377 microbiomes from 43 terrestrial and aquatic species, we found a decrease in the amplicon sequence variant-level microbiome phylogenetic diversity and alteration of microbiome composition under both experimental warming and cooling. Microbiome beta dispersion was not affected by temperature changes. We showed that the host habitat and experimental factors affected microbiome diversity and composition more than host biological traits. In particular, aquatic organisms-especially in marine habitats-experienced a greater depletion in microbiome diversity under cold conditions, compared to terrestrial hosts. Exposure involving a sudden long and static temperature shift was associated with microbiome diversity loss, but this reduction was attenuated by prior-experimental lab acclimation or when a ramped regime (i.e., warming) was used. Microbial differential abundance and co-occurrence network analyses revealed several potential indicator bacterial classes for hosts in heated environments and on different biome levels. Overall, our findings improve our understanding on the impact of global temperature changes on animal and plant microbiome structures across a diverse range of habitats. The next step is to link these changes to measures of host fitness, as well as microbial community functions, to determine whether microbiomes can buffer some species against a more thermally variable and extreme world.

Rapid evolution of a novel protective symbiont into keystone taxon in Caenorhabditis elegans microbiota.

Protective microbes have a major role in shaping host-pathogen interactions, but their relative importance in the structure of the host microbiota remains unclear. Here, we used a network approach to characterize the impact of a novel, experimentally evolved 'protective microbial symbiont' (Enterococcus faecalis) on the structure and predicted function of the natural microbiota of the model organism Caenorhabditis elegans. We used microbial network analysis to identify keystone taxa and describe the hierarchical placement of protective and non-protective symbionts in the microbiota. We found that early colonization with symbionts produce statistically significant changes in the structure of the community. Notably, only the protective E. faecalis became a keystone taxon in the nematode microbiota. Non-protective lineages of the same bacterial species remained comparatively unimportant to the community. Prediction of functional profiles in bacterial communities using PICRUSt2 showed that the presence of highly protective E. faecalis decreased the abundance of ergothioneine (EGT) biosynthesis pathway involved in the synthesis of the antioxidant molecule EGT, a potential public good. These data show that in addition to direct antagonism with virulent pathogens, keystone protective symbionts are linked to modified bacterial community structure and possible reductions in public goods, potentially driving decreased antioxidant defense. We suggest that this response could suppress infection via wholesale microbial community changes to further benefit the host. These findings extend the concept of protective symbionts beyond bodyguards to ecosystem engineers.

Microbiome function predicts amphibian chytridiomycosis disease dynamics.

BACKGROUND: The fungal pathogen Batrachochytrium dendrobatidis (Bd) threatens amphibian biodiversity and ecosystem stability worldwide. Amphibian skin microbial community structure has been linked to the clinical outcome of Bd infections, yet its overall functional importance is poorly understood. METHODS: Microbiome taxonomic and functional profiles were assessed using high-throughput bacterial 16S rRNA and fungal ITS2 gene sequencing, bacterial shotgun metagenomics and skin mucosal metabolomics. We sampled 56 wild midwife toads (Alytes obstetricans) from montane populations exhibiting Bd epizootic or enzootic disease dynamics. In addition, to assess whether disease-specific microbiome profiles were linked to microbe-mediated protection or Bd-induced perturbation, we performed a laboratory Bd challenge experiment whereby 40 young adult A. obstetricans were exposed to Bd or a control sham infection. We measured temporal changes in the microbiome as well as functional profiles of Bd-exposed and control animals at peak infection. RESULTS: Microbiome community structure and function differed in wild populations based on infection history and in experimental control versus Bd-exposed animals. Bd exposure in the laboratory resulted in dynamic changes in microbiome community structure and functional differences, with infection clearance in all but one infected animal. Sphingobacterium, Stenotrophomonas and an unclassified Commamonadaceae were associated with wild epizootic dynamics and also had reduced abundance in laboratory Bd-exposed animals that cleared infection, indicating a negative association with Bd resistance. This was further supported by microbe-metabolite integration which identified functionally relevant taxa driving disease outcome, of which Sphingobacterium and Bd were most influential in wild epizootic dynamics. The strong correlation between microbial taxonomic community composition and skin metabolome in the laboratory and field is inconsistent with microbial functional redundancy, indicating that differences in microbial taxonomy drive functional variation. Shotgun metagenomic analyses support these findings, with similar disease-associated patterns in beta diversity. Analysis of differentially abundant bacterial genes and pathways indicated that bacterial environmental sensing and Bd resource competition are likely to be important in driving infection outcomes. CONCLUSIONS: Bd infection drives altered microbiome taxonomic and functional profiles across laboratory and field environments. Our application of multi-omics analyses in experimental and field settings robustly predicts Bd disease dynamics and identifies novel candidate biomarkers of infection. Video Abstract.

A globally ubiquitous symbiont can drive experimental host evolution.

Organisms harbour myriad microbes which can be parasitic or protective against harm. The costs and benefits resulting from these symbiotic relationships can be context-dependent, but the evolutionary consequences to hosts of these transitions remain unclear. Here, we mapped the Leucobacter genus across 13,715 microbiome samples (163 studies) to reveal a global distribution as a free-living microbe or a symbiont of animals and plants. We showed that across geographically distant locations (South Africa, France, Cape Verde), Leucobacter isolates vary substantially in their virulence to an associated animal host, Caenorhabditis nematodes. We further found that multiple Leucobacter sequence variants co-occur in wild Caenorhabditis spp. which combined with natural variation in virulence provides real-world potential for Leucobacter community composition to influence host fitness. We examined this by competing C. elegans genotypes that differed in susceptibility to different Leucobacter species in an evolution experiment. One Leucobacter species was found to be host-protective against another, virulent parasitic species. We tested the impact of host genetic background and Leucobacter community composition on patterns of host-based defence evolution. We found host genotypes conferring defence against the parasitic species were maintained during infection. However, when hosts were protected during coinfection, host-based defences were nearly lost from the population. Overall, our results provide insight into the role of community context in shaping host evolution during symbioses.

Host microbiota can facilitate pathogen infection.

Animals live in symbiosis with numerous microbe species. While some can protect hosts from infection and benefit host health, components of the microbiota or changes to the microbial landscape have the potential to facilitate infections and worsen disease severity. Pathogens and pathobionts can exploit microbiota metabolites, or can take advantage of a depletion in host defences and changing conditions within a host, to cause opportunistic infection. The microbiota might also favour a more virulent evolutionary trajectory for invading pathogens. In this review, we consider the ways in which a host microbiota contributes to infectious disease throughout the host's life and potentially across evolutionary time. We further discuss the implications of these negative outcomes for microbiota manipulation and engineering in disease management.

Effects of multiple stressors on northern leopard frogs in agricultural wetlands.

Natural and anthropogenic stressors, including parasites and pesticides, may induce oxidative stress in animals. Measuring oxidative stress responses in sentinel species that are particularly responsive to environmental perturbations not only provides insight into host physiology but is also a useful readout of ecosystem health. Newly metamorphosed northern leopard frogs (Lithobates pipiens), a sentinel species, were collected from agricultural and non-agricultural wetlands exposed to varying concentrations of the herbicide atrazine. Significant effects of certain parasites' abundance and their interaction with atrazine exposure on frog oxidative stress were identified. Specifically, increased protein levels were detected in frogs infected with echinostome metacercariae. In addition, the nematode Oswaldocruzia sp. was significantly associated with increased thiol concentration and catalase activity. Significant parasite × atrazine interactions were observed for atrazine exposure and the abundance of Oswaldocruzia sp. on thiol, as thiol concentrations increased with parasite abundance at low atrazine localities and decreased in high atrazine wetlands. In addition, a significant interaction between the abundances of Oswaldocruzia sp. and gorgoderid trematodes on thiol concentrations was observed. These findings demonstrate that studies of oxidative stress on animals in natural ecosystems should account for the confounding effects of parasitism, particularly for amphibians in agricultural landscapes.

Captivity and Infection by the Fungal Pathogen Batrachochytrium salamandrivorans Perturb the Amphibian Skin Microbiome.

The emerging fungal pathogen, Batrachochytrium salamandrivorans (Bsal) is responsible for the catastrophic decline of European salamanders and poses a threat to amphibians globally. The amphibian skin microbiome can influence disease outcome for several host-pathogen systems, yet little is known of its role in Bsal infection. In addition, many experimental in-vivo amphibian disease studies to date have relied on specimens that have been kept in captivity for long periods without considering the influence of environment on the microbiome and how this may impact the host response to pathogen exposure. We characterized the impact of captivity and exposure to Bsal on the skin bacterial and fungal communities of two co-occurring European newt species, the smooth newt, Lissotriton vulgaris and the great-crested newt, Triturus cristatus. We show that captivity led to significant losses in bacterial and fungal diversity of amphibian skin, which may be indicative of a decline in microbe-mediated protection. We further demonstrate that in both L. vulgaris and T. cristatus, Bsal infection was associated with changes in the composition of skin bacterial communities with possible negative consequences to host health. Our findings advance current understanding of the role of host-associated microbiota in Bsal infection and highlight important considerations for ex-situ amphibian conservation programmes.

Diagnosing Emerging Fungal Threats: A One Health Perspective.

Emerging fungal pathogens are a growing threat to global health, ecosystems, food security, and the world economy. Over the last century, environmental change and globalized transport, twinned with the increasing application of antifungal chemical drugs have led to increases in outbreaks of fungal diseases with sometimes catastrophic effects. In order to tackle contemporary epidemics and predemic threats, there is a pressing need for a unified approach in identification and monitoring of fungal pathogens. In this paper, we discuss current high throughput technologies, as well as new platforms capable of combining diverse data types to inform practical epidemiological strategies with a focus on emerging fungal pathogens of wildlife.

Recent Asian origin of chytrid fungi causing global amphibian declines.

Globalized infectious diseases are causing species declines worldwide, but their source often remains elusive. We used whole-genome sequencing to solve the spatiotemporal origins of the most devastating panzootic to date, caused by the fungus Batrachochytrium dendrobatidis, a proximate driver of global amphibian declines. We traced the source of B. dendrobatidis to the Korean peninsula, where one lineage, BdASIA-1, exhibits the genetic hallmarks of an ancestral population that seeded the panzootic. We date the emergence of this pathogen to the early 20th century, coinciding with the global expansion of commercial trade in amphibians, and we show that intercontinental transmission is ongoing. Our findings point to East Asia as a geographic hotspot for B. dendrobatidis biodiversity and the original source of these lineages that now parasitize amphibians worldwide.

Amphibian chytridiomycosis outbreak dynamics are linked with host skin bacterial community structure.

Host-associated microbes are vital for combatting infections and maintaining health. In amphibians, certain skin-associated bacteria inhibit the fungal pathogen Batrachochytrium dendrobatidis (Bd), yet our understanding of host microbial ecology and its role in disease outbreaks is limited. We sampled skin-associated bacteria and Bd from Pyrenean midwife toad populations exhibiting enzootic or epizootic disease dynamics. We demonstrate that bacterial communities differ between life stages with few shared taxa, indicative of restructuring at metamorphosis. We detected a significant effect of infection history on metamorph skin microbiota, with reduced bacterial diversity in epizootic populations and differences in community structure and predicted function. Genome sequencing of Bd isolates supports a single introduction to the Pyrenees and reveals no association between pathogen genetics and epidemiological trends. Our findings provide an ecologically relevant insight into the microbial ecology of amphibian skin and highlight the relative importance of host microbiota and pathogen genetics in predicting disease outcome.