Chytridiomycosis causes high amphibian mortality prior to the completion of metamorphosis.

Amphibian populations are undergoing extensive declines globally. The fungal disease chytridiomycosis, caused by the pathogenic fungus Batrachochytrium dendrobatidis (Bd), is a primary contributor to these declines. The amphibian metamorphic stages (Gosner stages 42-46) are particularly vulnerable to a range of stressors, including Bd. Despite this, studies that explicitly examine host response to chytridiomycosis throughout the metamorphic stages are lacking. We aimed to determine how Bd exposure during the larval stages impacts metamorphic development and infection progression in the endangered Fleay's barred frog (Mixophyes fleayi). We exposed M. fleayi to Bd during pro-metamorphosis (Gosner stages 35-38) and monitored infection dynamics throughout metamorphosis. We took weekly morphological measurements (weight, total body length, snout-vent-length and Gosner stage) and quantified Bd load using qPCR. While we observed minimal impact of Bd infection on animal growth and development, Bd load varied throughout ontogeny, with an infection load plateau during the tadpole stages (Gosner stages 35-41) and temporary infection clearance at Gosner stage 42. Bd load increased exponentially between Gosner stages 42 and 45, with most exposed animals becoming moribund at Gosner stage 45, prior to the completion of metamorphosis. There was variability in infection outcome of exposed individuals, with a subgroup of animals (n = 5/29) apparently clearing their infection while the majority (n = 21/29) became moribund with high infection burdens. This study demonstrates the role that metamorphic restructuring plays in shaping Bd infection dynamics and raises the concern that substantial Bd-associated mortality could be overlooked in the field due to the often cryptic nature of these latter metamorphic stages. We recommend future studies that directly examine the host immune response to Bd infection throughout metamorphosis, incorporating histological and molecular methods to elucidate the mechanisms responsible for the observed trends.

Recovered frog populations coexist with endemic Batrachochytrium dendrobatidis despite load-dependent mortality.

Novel infectious diseases, particularly those caused by fungal pathogens, pose considerable risks to global biodiversity. The amphibian chytrid fungus (Batrachochytrium dendrobatidis, Bd) has demonstrated the scale of the threat, having caused the greatest recorded loss of vertebrate biodiversity attributable to a pathogen. Despite catastrophic declines on several continents, many affected species have experienced population recoveries after epidemics. However, the potential ongoing threat of endemic Bd in these recovered or recovering populations is still poorly understood. We investigated the threat of endemic Bd to frog populations that recovered after initial precipitous declines, focusing on the endangered rainforest frog Mixophyes fleayi. We conducted extensive field surveys over 4 years at three independent sites in eastern Australia. First, we compared Bd infection prevalence and infection intensities within frog communities to reveal species-specific infection patterns. Then, we analyzed mark-recapture data of M. fleayi to estimate the impact of Bd infection intensity on apparent mortality rates and Bd infection dynamics. We found that M. fleayi had lower infection intensities than sympatric frogs across the three sites, and cleared infections at higher rates than they gained infections throughout the study period. By incorporating time-varying individual infection intensities, we show that healthy M. fleayi populations persist despite increased apparent mortality associated with infrequent high Bd loads. Infection dynamics were influenced by environmental conditions, with Bd prevalence, infection intensity, and rates of gaining infection associated with lower temperatures and increased rainfall. However, mortality remained constant year-round despite these fluctuations in Bd infections, suggesting major mortality events did not occur over the study period. Together, our results demonstrate that while Bd is still a potential threat to recovered populations of M. fleayi, high rates of clearing infections and generally low average infection loads likely minimize mortality caused by Bd. Our results are consistent with pathogen resistance contributing to the coexistence of M. fleayi with endemic Bd. We emphasize the importance of incorporating infection intensity into disease models rather than infection status alone. Similar population and infection dynamics likely exist within other recovered amphibian-Bd systems around the globe, promising longer-term persistence in the face of endemic chytridiomycosis.

Limited impact of chytridiomycosis on juvenile frogs in a recovered species.

The amphibian chytrid fungus Batrachochytrium dendrobatidis (Bd) has caused catastrophic frog declines on several continents, but disease outcome is mediated by a number of factors. Host life stage is an important consideration and many studies have highlighted the vulnerability of recently metamorphosed or juvenile frogs compared to adults. The majority of these studies have taken place in a laboratory setting, and there is a general paucity of longitudinal field studies investigating the influence of life stage on disease outcome. In this study, we assessed the effect of endemic Bd on juvenile Mixophyes fleayi (Fleay's barred frog) in subtropical eastern Australian rainforest. Using photographic mark-recapture, we made 386 captures of 116 individuals and investigated the effect of Bd infection intensity on the apparent mortality rates of frogs using a multievent model correcting for infection state misclassification. We found that neither Bd infection status nor infection intensity predicted mortality in juvenile frogs, counter to the expectation that early life stages are more vulnerable to disease, despite average high infection prevalence (0.35, 95% HDPI [0.14, 0.52]). Additionally, we found that observed infection prevalence and intensity were somewhat lower for juveniles than adults. Our results indicate that in this Bd-recovered species, the realized impacts of chytridiomycosis on juveniles were apparently low, likely resulting in high recruitment contributing to population stability. We highlight the importance of investigating factors relating to disease outcome in a field setting and make recommendations for future studies.

Mechanisms underlying host persistence following amphibian disease emergence determine appropriate management strategies.

Emerging infectious diseases have caused many species declines, changes in communities and even extinctions. There are also many species that persist following devastating declines due to disease. The broad mechanisms that enable host persistence following declines include evolution of resistance or tolerance, changes in immunity and behaviour, compensatory recruitment, pathogen attenuation, environmental refugia, density-dependent transmission and changes in community composition. Here we examine the case of chytridiomycosis, the most important wildlife disease of the past century. We review the full breadth of mechanisms allowing host persistence, and synthesise research on host, pathogen, environmental and community factors driving persistence following chytridiomycosis-related declines and overview the current evidence and the information required to support each mechanism. We found that for most species the mechanisms facilitating persistence have not been identified. We illustrate how the mechanisms that drive long-term host population dynamics determine the most effective conservation management strategies. Therefore, understanding mechanisms of host persistence is important because many species continue to be threatened by disease, some of which will require intervention. The conceptual framework we describe is broadly applicable to other novel disease systems.

Susceptibility of frogs to chytridiomycosis correlates with increased levels of immunomodulatory serotonin in the skin.

Chytridiomycosis, caused by the fungus Batrachochytrium dendrobatidis (Bd), is a skin disease responsible for the global decline of amphibians. Frog species and populations can vary in susceptibility, but this phenomenon remains poorly understood. Here, we investigated serotonin in the skin of infected and uninfected frogs. In more susceptible frog populations, skin serotonin rose with increasing infection intensity, but decreased in later stages of the disease. The more resistant population maintained a basal level of skin serotonin. Serotonin inhibited both Bd sporangial growth and Jurkat lymphocyte proliferation in vitro. However, serotonin accumulates in skin granular glands, and this compartmentalisation may prevent inhibition of Bd growth in vivo. We suggest that skin serotonin increases in susceptible frogs due to pathogen excretion of precursor tryptophan, but that resistant frogs are able to control the levels of serotonin. Overall, the immunosuppressive effects of serotonin may contribute to the susceptibility of frogs to chytridiomycosis.

Evolution of resistance to chytridiomycosis is associated with a robust early immune response.

Potentiating the evolution of immunity is a promising strategy for addressing biodiversity diseases. Assisted selection for infection resistance may enable the recovery and persistence of amphibians threatened by chytridiomycosis, a devastating fungal skin disease threatening hundreds of species globally. However, knowledge of the mechanisms involved in the natural evolution of immunity to chytridiomycosis is limited. Understanding the mechanisms of such resistance may help speed-assisted selection. Using a transcriptomics approach, we examined gene expression responses of endangered alpine tree frogs (Litoria verreauxii alpina) to subclinical infection, comparing two long-exposed populations with a naïve population. We performed a blinded, randomized and controlled exposure experiment, collecting skin, liver and spleen tissues at 4, 8 and 14 days postexposure from 51 wild-caught captively reared infection-naïve adult frogs for transcriptome assembly and differential gene expression analyses. We analysed our results in conjunction with infection intensity data, and the results of a large clinical survival experiment run concurrently with individuals from the same clutches. Here, we show that frogs from an evolutionarily long-exposed and phenotypically more resistant population of the highly susceptible alpine tree frog demonstrate a more robust innate and adaptive immune response at the critical early subclinical stage of infection when compared with two more susceptible populations. These results are consistent with the occurrence of evolution of resistance against chytridiomycosis, help to explain underlying resistance mechanisms, and provide genes of potential interest and sequence data for future research. We recommend further investigation of cell-mediated immunity pathways, the role of interferons and mechanisms of lymphocyte suppression.

Endemicity of chytridiomycosis features pathogen overdispersion.

Pathogens can be critical drivers of the abundance and distribution of wild animal populations. The presence of an overdispersed pathogen load distribution between hosts (where few hosts harbour heavy parasite burdens and light infections are common) can have an important stabilizing effect on host-pathogen dynamics where infection intensity determines pathogenicity. This may potentially lead to endemicity of an introduced pathogen rather than extirpation of the host and/or pathogen. Overdispersed pathogen load distributions have rarely been considered in wild animal populations as an important component of the infection dynamics of microparasites such as bacteria, viruses, protozoa and fungi. Here we examined the abundance, distribution and transmission of the model fungal pathogen Batrachochytrium dendrobatidis (Bd, cause of amphibian chytridiomycosis) between wild-caught Litoria rheocola (common mist frogs) to investigate the effects of an overdispersed pathogen load distribution on the host population in the wild. We quantified host survival, infection incidence and recovery probabilities relative to infectious burden, and compared the results of models where pathogen overdispersion either was or was not considered an important feature of host-pathogen dynamics. We found the distribution of Bd load between hosts to be highly overdispersed. We found that host survival was related to infection burden and that accounting for pathogen overdispersion allowed us to better understand infection dynamics and their implications for disease control. In addition, we found that the pattern of host infections and recoveries varied markedly with season whereby (i) infections established more in winter, consistent with temperature-dependent effects on fungal growth, and (ii) recoveries (loss of infection) occurred frequently in the field throughout the year but were less likely in winter. Our results suggest that pathogen overdispersion is an important feature of endemic chytridiomycosis and that intensity of infection determines disease impact. These findings have important implications for our understanding of chytridiomycosis dynamics and the application of management strategies for disease mitigation. We recommend quantifying individual infectious burdens rather than infection state where possible in microparasitic diseases.

Interventions for reducing extinction risk in chytridiomycosis-threatened amphibians.

Wildlife diseases pose an increasing threat to biodiversity and are a major management challenge. A striking example of this threat is the emergence of chytridiomycosis. Despite diagnosis of chytridiomycosis as an important driver of global amphibian declines 15 years ago, researchers have yet to devise effective large-scale management responses other than biosecurity measures to mitigate disease spread and the establishment of disease-free captive assurance colonies prior to or during disease outbreaks. We examined the development of management actions that can be implemented after an epidemic in surviving populations. We developed a conceptual framework with clear interventions to guide experimental management and applied research so that further extinctions of amphibian species threatened by chytridiomycosis might be prevented. Within our framework, there are 2 management approaches: reducing Batrachochytrium dendrobatidis (the fungus that causes chytridiomycosis) in the environment or on amphibians and increasing the capacity of populations to persist despite increased mortality from disease. The latter approach emphasizes that mitigation does not necessarily need to focus on reducing disease-associated mortality. We propose promising management actions that can be implemented and tested based on current knowledge and that include habitat manipulation, antifungal treatments, animal translocation, bioaugmentation, head starting, and selection for resistance. Case studies where these strategies are being implemented will demonstrate their potential to save critically endangered species.

Metabolomic response of striped marsh frog (Limnodynastes peronii) tadpoles exposed to the fire retardant Phos-Chek LC95W.

Climate change and other factors have contributed to an increased frequency and intensity of global wildfires in recent years. Ammonium-based fire retardants are widely used to suppress or delay the spread of fire and have generally been regarded as presenting a low risk of acute toxicity to fauna. However, studies have raised concerns about their potential to cause indirect or sub-lethal effects, and toxicity information regarding the potential for such impacts in aquatic species is limited. To address these knowledge gaps, we used an untargeted metabolomics approach to evaluate the sub-lethal physiological and metabolic responses of striped marsh frog (Limnodynastes peronii) tadpoles exposed to a concentration gradient of the ammonium polyphosphate (APP)-based fire retardant Phos-Chek LC95W (PC). Acute exposure (96 h) to PC significantly altered the relative abundance of 14 metabolites in whole tadpoles. The overall metabolic response pattern was consistent with effects reported for ammonia toxicity and also suggestive of energy dysregulation and osmotic stress associated with alterations to physicochemical water quality parameters in the PC treatments. Results suggest that run-off or accidental application of this formulation into waterways can have significant sub-lethal consequences on the biochemical profiles (i.e., the metabolome) of aquatic organisms and may be a concern for frog species that breed and develop in small, often ephemeral, waterbodies. Our study highlights the benefits of integrating untargeted metabolomics with other ecological and toxicological endpoints to provide a more holistic characterisation of the sub-lethal impacts associated with bushfire-fighting chemicals and with environmental contaminants more broadly.

Local resource availability drives habitat use by a threatened avian granivore in savanna woodlands.

Conserving threatened species relies on an understanding of their habitat requirements. This is especially relevant for granivorous birds, whose habitat use and movement patterns are intricately linked to the spatial and temporal availability of resources such as food and water. In this study, we investigated the habitat use, home range and daily activity patterns of the Endangered Southern Black-throated Finch (SBTF; Poephila cincta cincta) within a 75,000 ha savanna woodland study area in northeastern Australia. This semi-arid region is one of the key remaining strongholds for the species and is characterised by substantially different habitat and climatic conditions than areas where previous research on this species has been undertaken. We radio tracked 142 SBTF using both manual tracking and an array of 27 automated radio towers, which revealed a strong preference for eucalypt-dominated grassy woodland communities. The preference for these habitats also increased with decreasing distance to permanent water. While SBTF occupied large home ranges, individual SBTF were largely sedentary during the radio tracking period (21.8, range = 0.83-120 days), with few landscape-scale movements of more than 4.5 km. Daily foraging activity of SBTF exhibited bimodal peaks in the early morning and late afternoon, while other activities were greatest from the late morning to the early afternoon. Compared to other estrildid finches, our research suggests that SBTF track resources at a local scale across a large home range. We postulate that in times of resource scarcity SBTF may use dietary diversification, instead of landscape or regional-scale nomadic movements, to meet their resource needs. The species' movement patterns underscore the importance of local scale habitat management to facilitate resource availability throughout the year. Furthermore, our research helps target monitoring designs for granivorous birds that focus on the species' diurnal activity patterns.

Chytridiomycosis and seasonal mortality of tropical stream-associated frogs 15 years after introduction of Batrachochytrium dendrobatidis.

Assessing the effects of diseases on wildlife populations can be difficult in the absence of observed mortalities, but it is crucial for threat assessment and conservation. We performed an intensive capture-mark-recapture study across seasons and years to investigate the effect of chytridiomycosis on demographics in 2 populations of the threatened common mist frog (Litoria rheocola) in the lowland wet tropics of Queensland, Australia. Infection prevalence was the best predictor for apparent survival probability in adult males and varied widely with season (0-65%). Infection prevalence was highest in winter months when monthly survival probabilities were low (approximately 70%). Populations at both sites exhibited very low annual survival probabilities (12-15%) but high recruitment (71-91%), which resulted in population growth rates that fluctuated seasonally. Our results suggest that even in the absence of observed mortalities and continued declines, and despite host-pathogen co-existence for multiple host generations over almost 2 decades, chytridiomycosis continues to have substantial seasonally fluctuating population-level effects on amphibian survival, which necessitates increased recruitment for population persistence. Similarly infected populations may thus be under continued threat from chytridiomycosis which may render them vulnerable to other threatening processes, particularly those affecting recruitment success.

Amphibian infection tolerance to chytridiomycosis.

Animal defences against infection involve two distinct but complementary mechanisms: tolerance and resistance. Tolerance measures the animal's ability to limit detrimental effects from a given infection, whereas resistance is the ability to limit the intensity of that infection. Tolerance is a valuable defence for highly prevalent, persistent or endemic infections where mitigation strategies based on traditional resistance mechanisms are less effective or evolutionarily stable. Selective breeding of amphibians for enhanced tolerance to Batrachochytrium spp. has been suggested as a strategy for mitigating the impacts of the fungal disease, chytridiomycosis. Here, we define infection tolerance and resistance in the context of chytridiomycosis, present evidence for variation in tolerance to chytridiomycosis, and explore epidemiological, ecological and evolutionary implications of tolerance to chytridiomycosis. We found that exposure risk and environmental moderation of infection burdens are major confounders of resistance and tolerance, chytridiomycosis is primarily characterized by variation in constitutive rather than adaptive resistance, tolerance is epidemiologically important in driving pathogen spread and maintenance, heterogeneity of tolerance leads to ecological trade-offs, and natural selection for resistance and tolerance is likely to be dilute. Improving our understanding of infection tolerance broadens our capacity for mitigating the ongoing impacts of emerging infectious diseases such as chytridiomycosis. This article is part of the theme issue 'Amphibian immunity: stress, disease and ecoimmunology'.

Effects of two firefighting chemical formulations, Phos-Chek LC95W and BlazeTamer380, on striped marsh frog (Limodynastes peronii) tadpole survival, growth, development and behaviour.

Global wildfire events are projected to become more frequent and severe due to the continual threat of climate change, resulting in increasing demand for effective fire mitigation methods. Firefighting chemicals (FFCs), including retardants, foams and water enhancers, are often used to prevent the spread of wildfires. However, the impact of FFCs on wildlife and ecosystems is poorly understood. We investigated the effects of two common FFC formulations, Phos-Chek LC95W and BlazeTamer380, on tadpole survival, growth, development and swimming behaviour. Tadpoles of the striped marsh frog (Limnodynastes peronii) were exposed to two concentrations of either Phos-Chek (0.25 and 1 g/L) or BlazeTamer (0.05 and 0.2 g/L) for 16 days. The highest concentration of Phos-Chek was lethal to tadpoles, with mortalities gradually increasing over time and only 8% of animals surviving to day 16. Both FFCs influenced the growth and development of tadpoles, though effects were more severe in tadpoles exposed to the Phos-Chek formulation. Phos-Chek was found to completely stop tadpole growth and development over the 16-day exposure, whereas BlazeTamer significantly delayed growth and development in comparison to controls. Nevertheless, treatments had no apparent effect on tadpole movement patterns and swimming activity. Greater toxicity caused by the Phos-Chek treatment likely relates to the increased ammonia and altered water quality parameters. Runoff or accidental application of commonly used FFCs into small waterways may therefore have important ramifications for aquatic biota.

Do immune system changes at metamorphosis predict vulnerability to chytridiomycosis? An update.

Amphibians are among the vertebrate groups suffering great losses of biodiversity due to a variety of causes including diseases, such as chytridiomycosis (caused by the fungal pathogens Batrachochytrium dendrobatidis and B. salamandrivorans). The amphibian metamorphic period has been identified as being particularly vulnerable to chytridiomycosis, with dramatic physiological and immunological reorganisation likely contributing to this vulnerability. Here, we overview the processes behind these changes at metamorphosis and then perform a systematic literature review to capture the breadth of empirical research performed over the last two decades on the metamorphic immune response. We found that few studies focused specifically on the immune response during the peri-metamorphic stages of amphibian development and fewer still on the implications of their findings with respect to chytridiomycosis. We recommend future studies consider components of the immune system that are currently under-represented in the literature on amphibian metamorphosis, particularly pathogen recognition pathways. Although logistically challenging, we suggest varying the timing of exposure to Bd across metamorphosis to examine the relative importance of pathogen evasion, suppression or dysregulation of the immune system. We also suggest elucidating the underlying mechanisms of the increased susceptibility to chytridiomycosis at metamorphosis and the associated implications for population persistence. For species that overlap a distribution where Bd/Bsal are now endemic, we recommend a greater focus on management strategies that consider the important peri-metamorphic period.

Declining amphibians might be evolving increased reproductive effort in the face of devastating disease.

The devastating infectious disease chytridiomycosis has caused declines of amphibians across the globe, yet some populations are persisting and even recovering. One understudied effect of wildlife disease is changes in reproductive effort. Here, we aimed to understand if the disease has plastic effects on reproduction and if reproductive effort could evolve with disease endemism. We compared the effects of experimental pathogen exposure (trait plasticity) and population-level disease history (evolution in trait baseline) on reproductive effort using gametogenesis as a proxy in the declining and endangered frog Litoria verreauxii alpina. We found that unexposed males from disease-endemic populations had higher reproductive effort, which is consistent with an evolutionary response to chytridiomycosis. We also found evidence of trait plasticity, where males and females were affected differently by infection: pathogen exposed males had higher reproductive effort (larger testes), whereas females had reduced reproductive effort (smaller and fewer developed eggs) regardless of the population of origin. Infectious diseases can cause plastic changes in the reproductive effort at an individual level, and population-level disease exposure can result in changes to baseline reproductive effort; therefore, individual- and population-level effects of disease should be considered when designing management and conservation programs for threatened and declining species.

A SLC4-like anion exchanger from renal tubules of the mosquito (Aedes aegypti): evidence for a novel role of stellate cells in diuretic fluid secretion.

Transepithelial fluid secretion across the renal (Malpighian) tubule epithelium of the mosquito (Aedes aegypti) is energized by the vacuolar-type (V-type) H(+)-ATPase and not the Na(+)-K(+)-ATPase. Located at the apical membrane of principal cells, the V-type H(+)-ATPase translocates protons from the cytoplasm to the tubule lumen. Secreted protons are likely to derive from metabolic H(2)CO(3), which raises questions about the handling of HCO(3)(-) by principal cells. Accordingly, we tested the hypothesis that a Cl/HCO(3) anion exchanger (AE) related to the solute-linked carrier 4 (SLC4) superfamily mediates the extrusion of HCO(3)(-) across the basal membrane of principal cells. We began by cloning from Aedes Malpighian tubules a full-length cDNA encoding an SLC4-like AE, termed AeAE. When expressed heterologously in Xenopus oocytes, AeAE is both N- and O-glycosylated and mediates Na(+)-independent intracellular pH changes that are sensitive to extracellular Cl(-) concentration and to DIDS. In Aedes Malpighian tubules, AeAE is expressed as two distinct forms: one is O-glycosylated, and the other is N-glycosylated. Significantly, AeAE immunoreactivity localizes to the basal regions of stellate cells but not principal cells. Concentrations of DIDS that inhibit AeAE activity in Xenopus oocytes have no effects on the unstimulated rates of fluid secretion mediated by Malpighian tubules as measured by the Ramsay assay. However, in Malpighian tubules stimulated with kinin or calcitonin-like diuretic peptides, DIDS reduces the diuretic rates of fluid secretion to basal levels. In conclusion, Aedes Malpighian tubules express AeAE in the basal region of stellate cells, where this transporter may participate in producing diuretic rates of transepithelial fluid secretion.

Immunological Aspects of Chytridiomycosis.

Amphibians are currently the most threatened vertebrate class, with the disease chytridiomycosis being a major contributor to their global declines. Chytridiomycosis is a frequently fatal skin disease caused by the fungal pathogens Batrachochytrium dendrobatidis (Bd) and Batrachochytrium salamandrivorans (Bsal). The severity and extent of the impact of the infection caused by these pathogens across modern Amphibia are unprecedented in the history of vertebrate infectious diseases. The immune system of amphibians is thought to be largely similar to that of other jawed vertebrates, such as mammals. However, amphibian hosts are both ectothermic and water-dependent, which are characteristics favouring fungal proliferation. Although amphibians possess robust constitutive host defences, Bd/Bsal replicate within host cells once these defences have been breached. Intracellular fungal localisation may contribute to evasion of the induced innate immune response. Increasing evidence suggests that once the innate defences are surpassed, fungal virulence factors suppress the targeted adaptive immune responses whilst promoting an ineffectual inflammatory cascade, resulting in immunopathology and systemic metabolic disruption. Thus, although infections are contained within the integument, crucial homeostatic processes become compromised, leading to mortality. In this paper, we present an integrated synthesis of amphibian post-metamorphic immunological responses and the corresponding outcomes of infection with Bd, focusing on recent developments within the field and highlighting future directions.

Extreme Competence: Keystone Hosts of Infections.

Individual hosts differ extensively in their competence for parasites, but traditional research has discounted this variation, partly because modeling such heterogeneity is difficult. This discounting has diminished as tools have improved and recognition has grown that some hosts, the extremely competent, can have exceptional impacts on disease dynamics. Most prominent among these hosts are the superspreaders, but other forms of extreme competence (EC) exist and others await discovery; each with potentially strong but distinct implications for disease emergence and spread. Here, we propose a framework for the study and discovery of EC, suitable for different host-parasite systems, which we hope enhances our understanding of how parasites circulate and evolve in host communities.

Chytridiomycosis causes catastrophic organism-wide metabolic dysregulation including profound failure of cellular energy pathways.

Chytridiomycosis is among several recently emerged fungal diseases of wildlife that have caused decline or extinction of naïve populations. Despite recent advances in understanding pathogenesis, host response to infection remains poorly understood. Here we modelled a total of 162 metabolites across skin and liver tissues of 61 frogs from four populations (three long-exposed and one naïve to the fungus) of the Australian alpine tree frog (Litoria verreauxii alpina) throughout a longitudinal exposure experiment involving both infected and negative control individuals. We found that chytridiomycosis dramatically altered the organism-wide metabolism of clinically diseased frogs. Chytridiomycosis caused catastrophic failure of normal homeostatic mechanisms (interruption of biosynthetic and degradation metabolic pathways), and pronounced dysregulation of cellular energy metabolism. Key intermediates of the tricarboxylic acid cycle were markedly depleted, including in particular α-ketoglutarate and glutamate that together constitute a key nutrient pathway for immune processes. This study was the first to apply a non-targeted metabolomics approach to a fungal wildlife disease and specifically to dissect the host-pathogen interface of Bd-infected frogs. The patterns of metabolite accumulation we have identified reveal whole-body metabolic dysfunction induced by a fungal skin infection, and these findings have broad relevance for other fungal diseases.