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Amphibians are the most imperilled class of animals on the planet. A major contributor to this has been the amphibian disease chytridiomycosis, caused by the chytrid fungus Batrachochytrium dendrobatidis (Bd), which spread around the world during the 20th century because of increasing global trade. Following epizootic waves across multiple continents which contributed to the extinction of around 100 species, several species appear to have recovered after often precipitous declines. However, few of these species have been studied extensively to investigate their current response to chytridiomycosis. The purpose of this project was to understand the effects of Bd infection on mortality and to quantify infection dynamics across multiple life stages of a recovering species in the field. I conducted extensive surveys of all life stages of Fleay’s barred frog (Mixophyes fleayi) across three sites. I applied rigorous, bespoke statistical models to assess the effect of Bd infection on mortality. Upon the recognition of imperfect pathogen detection in the Bd sampling protocols, I developed a novel extension to multistate models to propagate this uncertainty into the ecological inference. Chapter 3 reports on a four-year mark-recapture study on adults across three sites, where I show that Bd infection intensity, and not infection status, is an important predictor of mortality, but that populations remain stable despite infections. Chapter 4 details the development of a novel multievent model to account for imperfect pathogen detection, where I show that infection dynamics parameters are severely biased in traditional models. In Chapter 5, I report the findings of a four-month mark-recapture study on juveniles—the first of its kind—where I found that this life stage of M. fleayi is not more vulnerable to chytridiomycosis than adults, in contrast to results reported in previous literature. In Chapter 6, I assessed the seasonal patterns of Bd infections in tadpoles at two independent streams and investigated the effect of infection on mouthpart abnormalities. Finally, the general discussion provides an integrated assessment of the response of M. fleayi to Bd, where I suggest continued persistence is likely and that other recovered species can potentially follow similar trajectories. This project represents a significant contribution to the literature of post-epizootic species recovery. Additionally, the statistical methods presented herein are considered an important methodological advance for improved inference on infection prevalence and dynamics in a wide range of disease systems.

Chytridiomycosis is affecting hundreds of amphibian species worldwide, but while in tropical areas, adult individuals have been the focus of most investigations, the exact role played by infection intensity of breeding adults is not well understood in temperate areas. We conducted mark-recapture-capture surveys during spiny common toad breeding seasons from 2006 to 2018 at the site of the first recorded outbreak of chytridiomycosis in Europe, the Peñalara Massif (Sierra de Guadarrama National Park, central Spain), and collected infection samples and several variables related to the reproductive effort of male individuals. We used general linear mixed models to evaluate the contribution of study variables on the infection loads of adult male toads exhibited at their capturing date. We also analysed the differences on several male characteristics between the pond with the largest breeding population against the rest of the ponds. We found that the duration of time spent in the waterbody and the condition of the host predicted infection loads. Animals of good physical condition, that spent longer in water, have higher infection levels than individuals with the opposite set of traits. The pond supporting the largest breeding population housed smaller male toads and in poorer condition. Our results are consistent with a shift in reproductive strategy in response to infection and potentially a strategy of tolerance, rather than resistance to infection. These findings have applications for disease mitigation and theoretical implications related to the trade-offs made and the evolution of traits in response to the disease., Open Access funding provided thanks to the CRUE-CSIC agreement with Springer Nature. This study was funded by Organismo Autónomo Parques Nacionales of Spain (Project Ref. 2399/2017, PI: JBo). BT was supported by ‘Doctorados Industriales de la Comunidad de Madrid’ (Ref. IND2020/AMB-17438).

In 2020 and 2021 we detected in Catalonia (NE Spain) six new episodes of mortality or presence of dying specimens in introduced Mediterranean painted frogs (Discoglossus pictus). All affected individuals were positive to the amphibian chytrid fungus Batrachochytrium dendrobatidis (Bd). Histological examinations showed intense involvement of the superficial layers of the epidermis and qPCR analyses yielded infection loads greater than 100 000 genomic equivalents of zoospores. These results suggest that this introduced species could behave as a Bd ‘supershedder’, and recommend to control its expansion due to its important role in the dynamic of chytridiomycosis.

Amphibian declines are a threat to global biodiversity and part of Earth’s sixth mass extinction. Amphibian extinctions and extirpations are caused by a variety of factors, including the Chytridiomycosis panzootic and its causative agents: the batrachochytridsBatrachochytrium salamandrivorans(Bsal)andBatrachochytrium dendrobatidis(Bd).Until recently, the genomic basis for the batrachochytrids’ evolution and their virulence was largely unknown. To investigate their virulence and evolution, high-quality genome assemblies were needed. Prior to 2022, only a highly fragmented short-read assembly ofBsal’s genome assembly had been generated and made available, largely owing to its repeat-richness. In 2022, a new assembly based on deep nanopore long-read sequencing provided a much-needed improvement in both contiguity and completeness. The new assembly revealed thatBsalhas undergone a repeat-driven genome expansion and has a highly compartmentalized genome architecture. Here, the road map to assembling and polishing a highly repeat-rich chytrid fungus genome using both long and short reads is described, detailing the methodology for each step, and explaining the rationales underpinning those steps. Considerations for tailoring this methodology to other similar genome assemblies are illustrated. Additionally, we describe how a high-quality assembly can be analysed to provide insight into the genomic architecture and the underlying evolutionary processes.&nbsp

Based on examination of most of the existing museum specimens of the rare bufonid frog Incilius tacanensis, we present a redescription and new diagnosis for this species. The species is limited to small region of the Pacific chain of volcanoes in southeastern Chiapas, Mexico, and adjacent areas of Guatemala. The species has not been observed in the wild since 1984 and may have been reduced or eliminated by regional epidemics of chytridiomycosis.

Mycoviruses may influence the pathogenicity of disease-causing fungi. Although mycoviruses have been found in some chytrid fungi, limited testing has not detected them in Batrachochytrium dendrobatidis (Bd), the cause of the devastating amphibian disease, chytridiomycosis. Here we conducted a survey for mycovirus presence in 38 Bd isolates from Australia (n = 31), Brazil (n = 5) and South Korea (n = 2) with a combination of modern high-throughput sequencing and conventional dsRNA cellulose chromatography. Mycoviruses were not detected in any isolates. This result was unexpected, given the long evolutionary history of Bd, as well as the high prevalence of mycoviruses in related fungal species. Given our widespread sampling in Australia and the limited number of Bd introductions, we suggest that mycoviruses are uncommon or absent from Australian Bd. Testing more isolates from regions where Bd originated, as well as regions with high diversity or low fungal virulence may identify mycoviruses that could aid in disease control.

Chytridiomycosis, the disease caused by the fungal pathogen Batrachochytrium dendrobatidis (Bd), has been unambiguously implicated in the decline of amphibian populations worldwide. However, the impact of this devastating infectious disease can be difficult to gauge without empirical data on the population-level effects of Bd. Often, assessments of the amphibian chytridiomycosis panzootic are based primarily on expert opinions; as a result, declines in tropical areas are promptly attributed to Bd while its impact on temperate species not suffering from adult mass mortalities is frequently overlooked. Here, we investigated the survival probability in an amphibian species from a temperate area that until now has not been considered to be severely impacted by the disease. Specifically, we related individual survival to Bd infection status using long-term capture-mark-recapture data of male spiny common toads (Bufo spinosus) in Sierra de Guadarrama National Park in central Spain. Even though the study population has demonstrated potential for adaptation to Bd and die-offs of adult individuals have not been recorded, our results clearly indicated that the probability of survival was lower for Bd-positive individuals. Moreover, the probability of becoming Bd-positive was higher than the probability of clearance, driving the population to a slow but certain decline. These results are consistent with other indicators of a negative population trend and suggest that the impact of Bd on temperate species of less concern may be greater than previously thought.

Anthropogenic habitat disturbances can dramatically alter ecological community interactions, including host-pathogen dynamics. Recent work has highlighted the potential for habitat disturbances to alter host-associated microbial communities, but the associations between anthropogenic disturbance, host microbiomes, and pathogens are unresolved. Amphibian skin microbial communities are particularly responsive to factors like temperature, physiochemistry, pathogen infection, and environmental microbial reservoirs. Through a field survey on wild populations of Acris crepitans (Hylidae) and Lithobates catesbeianus (Ranidae), we assessed effects of habitat disturbance on environmental bacterial resevoirs, Batrachochytrium dendrobatidis (Bd) infection, and skin microbiome composition. We found higher measures of microbiome dispersion (a measure of community stability) in A. crepitans from more disturbed ponds, supporting the hypothesis that disturbance increases stochasticity in biological communties. We also found that habitat disturbance limited microbiome similarity between locations for both species, suggesting less bacterial exchange in more disturbed areas. Higher disturbance was associated with lower Bd prevalence for A. crepitans, which could signify suboptimal microclimates for Bd in disturbed habitats. In this system we use microbiome dispersion as a metric of population health. Combined, our findings show that reduced microbiome stability stemming from habitat disturbance could compromise population health, even in the absence of pathogenic infection.

We report the first cases of mortality in anurans associated with the presence of the fungus Batrachochytrium dendrobatidis at four different localities from Catalonia (NE Iberian Peninsula, Spain). All cases were confirmed by both molecular techniques and histology. The infected individuals were two Mediterranean painted frogs (Discoglossus pictus) from Girona province found in 2018 and 2020, one Iberian waterfrog (Pelophylax perezi) from Tarragona province found in 2018, and one European common frog (Rana temporaria) from Barcelona province found in 2019. This is the first time that mortality associated with this pathogen has been confirmed in D. pictus and P. perezi. The role of the fungus as an agent possibly leading to death, in association with other external environmental factors, is discussed. These findings could suggest a recent increase of the incidence of this disease in the region. Some of these cases are particularly worrying because of their occurrence close to sites where some endemic amphibian species with extremely reduced distributions inhabit.

While the amphibian disease chytridiomycosis is causing ongoing population declines and biodiversity losses around the globe, efficient mitigation strategies are lacking. The free-living zoospores of the causative agents of this disease, the chytrid pathogens Batrachochytrium dendrobatidis (Bd) and Batrachochytrium salamandrivorans (Bsal), are a potential food source for filter-feeding micropredators as part of the aquatic food web. While consumption of zoospores can lower environmental pathogen loads, alternative food sources may interfere with pathogen removal rates. We compared the ability of three filter-feeding zooplankton species, the cladoceran Daphnia magna, the rotifer Brachionus calyciflorus and the ostracod Heterocypris incongruens, to remove Bd zoospores in water and investigated the effect of alternative food sources, the green algae Pseudokirchneriella subcapitata and Chlorella vulgaris, on zoospore ingestion by D. magna. Daphnia magna was the only micropredator candidate that effectively removed Bd zoospores from its environment, with an average removal rate of 1,012 +/- 542 GE ind.(-1) hr(-1) within our test system. High concentrations (1 x 10(5) cells/ml) of large and easily ingestible P. subcapitata reduced pathogen removal rates, whereas the small and less edible C. vulgaris did not interfere with pathogen removal. Synthesis and applications. We showed that Daphnia spp., which are keystone species in all sorts of aquatic habitats worldwide, are promising target agents for biologically mitigating chytridiomycosis infections and how natural food sources may interfere with this strategy. We also suggest potential management actions for biological disease mitigation, aiming to optimize environmental conditions for these target filter feeders, thereby reducing pathogen densities and eventually infection pressure in amphibian hosts. Examples of such management actions include, but are not limited to, removal of planktivorous fish, habitat restoration, nutrient control or agrochemical regulation in the vicinity of amphibian breeding ponds. Further studies, including field trials, are needed to confirm the effects of pathogen consumption on infection dynamics in natural situations and investigate the impact of intervention actions.

The skin microbiome in amphibians has gained a lot of attention as some of its members play a protective role against pathogens such as the fungus Batrachochytrium dendrobatidis (Bd). The composition of skin bacterial communities has been suggested as one of the factors explaining differences in susceptibility to Bd among amphibian species and populations. The boreal toad Anaxyrus boreas is known to be susceptible to Bd, and severe population declines in its southeastern range have been documented. However, throughout A. boreas distribution, populations present differences in susceptibility to Bd infections which may be associated with differences in skin microbial diversity. This study compared the skin bacterial diversity and Bd infection levels of A. boreas in one desert population and one pine forest population from Baja California, Mexico. We found that desert and pine forest toad populations exhibit differences in skin bacterial community structure but show similar Bd infection levels. Using a predictive method, we found that the abundance of bacteria with potential Bd-inhibitory properties differed between uninfected and infected individuals but not between populations. Our data suggest that several bacteria in the skin community may be offering protection from Bd infections in these A. boreas populations. This study provides foundational evidence for future studies seeking to understand the skin-microbial variation among boreal toads’ populations and its relation with Bd susceptibility.

More than a third of the world’s amphibian species are listed as Threatened or Extinct, with a recent assessment identifying 45 Australian frogs (18.4% of the currently recognised species) as ‘Threatened’ based on IUCN criteria. We applied structured expert elicitation to 26 frogs assessed as Critically Endangered and Endangered to estimate their probability of extinction by 2040. We also investigated whether participant experience (measured as a self-assigned categorical score, i.e. ‘expert’ or ‘non-expert’) influenced the estimates. Collation and analysis of participant opinion indicated that eight species are at high risk (>50% chance) of becoming extinct by 2040, with the disease chytridiomycosis identified as the primary threat. A further five species are at moderate–high risk (30–50% chance), primarily due to climate change. Fourteen of the 26 frog species are endemic to Queensland, with many species restricted to small geographic ranges that are susceptible to stochastic events (e.g. a severe heatwave or a large bushfire). Experts were more likely to rate extinction probability higher for poorly known species (those with

The ability of an organism to tolerate seasonal temperature changes, such as extremely cold temperatures during the winter, can be influenced by their pathogens. We tested how exposure to a virulent fungal pathogen, Batrachochytrium dendrobatidis (Bd), affected the critical thermal minimum (CTmin) of two frog species, Hyla versicolor (gray treefrog) and Lithobates palustris (pickerel frog). The CTmin is the minimum thermal performance point of an organism, which we estimated via righting response trials. For both frog species, we compared the righting response of Bd‐exposed and Bd‐unexposed individuals in either a constant (15ºC) environment or with decreasing temperatures (−1°C/2.5 min) starting from 15°C. The CTmin for both species was higher for Bd‐exposed frogs than unexposed frogs, and the CTmin of H. versicolor was higher than L. palustris. We also found that Bd‐exposed frogs of both species righted themselves significantly fewer times in both decreasing and constant temperature trials. Our findings show that pathogen exposure can reduce cold tolerance and limit the thermal performance range of hosts, which may lead to increased overwintering mortality., We tested how exposure to a virulent fungal pathogen, Batrachochytrium dendrobatidis, affected the critical thermal minimum of two frog species. In both frog species, exposure to the fungal pathogen increased the critical thermal minimum. Our findings show that pathogen exposure can reduce cold tolerance and limit the thermal performance range of hosts.

Chytridiomycosis, an emergent infectious disease caused by the fungusBatrachochytrium dendrobatidis(Bd), is considered one of the drivers of the current amphibian biodiversity loss. To inform endangered species conservation efforts, it is essential to improve our knowledge about the abiotic and biotic factors that influenceBdinfection dynamics in the wild. Here, we analyzed variation ofBdinfection in the redbelly toadMelanophryniscus montevidensis, a threatened bufonid from Uruguay. We tested the influence of temperature, precipitation, season, and host population size onBdprevalence and intensity. Additionally, considering the sub-lethal effectsofBd, we tested if these variables, potentially through their effect onBd, also explain the variation in host body condition. We determined a highBdprevalence of 41% (100/241), and that population size influenced bothBdprevalence and infection intensity. We identified an effect of precipitation and season onBdinfection intensity and an effect of season on toad body condition. In addition, we found a negative effect of infection intensity on body condition; moreover, while some toads cleared the infection, their body condition did not improve, suggesting a long-term cost. This is the first report on host population size as an important factor inBdinfection dynamics in a threatened anuran species, and seasonal demographic changes appear to play an important role in the dynamics. Finally, we highlight the need for monitoringBdin this and other endangered amphibian populations, especially those within the genusMelanophryniscus, which includes several Endangered and Data Deficient species in South America.

Crawshaw, L., Buchanan. T., Shirose, L., Palahnuk, A., Cai, H.Y., Bennett, A.M., Jardine, C.M., Davy, C.M. We conducted surveillance for two chytrid fungi (Batrachochytrium dendrobatidis and B. salamandrivorans) in south-central Canada, in an area where previous niche modelling suggests low habitat suitability for B. dendrobatidis. Sampling of 1041 amphibians across Ontario and Akimiski Island (Nunavut) revealed that B. dendrobatidis is ubiquitous in our study area, but did not detect B. salamandrivorans. Niche models informed by these data show high habitat suitability for B. dendrobatidis, and temperature models suggest high habitat suitability for B. salamandrivorans if it is introduced, emphasizing the need for enhanced coordinated, trans-boundary regulation of trade and surveillance of potential pathways of transmission.

Bu çalışmada, Türkiye'de yaşamakta olan amfibi türleri üzerinde IUCN (İnternational Union for Conservation of Nature) kriterleri doğrultusunda Nesli Tükenmekte Olan Türler kategorisinde Red Listte endemik olan Tavass Kurbağasının( Rana tavasensis ) azaldığı bilşinmekte, bireyleri üzerinde Real time PCR tekniği kullanılarak ölümcül hastalıklarla ilişkili olduğu bilinen amfibi patojenleri Batrachochytrium dendrobatidis ve poikilotherm hayvanlara rahatlıkla bulaşabilmekte olan Ranavirus [ Frog virus (FV-3) ] genom equivalentlerine ait primerler yardımıyla varlık veya yokluğu üzerinde inceleme yapılmıştır. Araştırma sonucu olarak amfibi türleri üzerinde yok oluşlardan sorumlu olarak kabul edilen iki önemli patojen; Batrachochytrium dendrobatidis (Bd) fungusu ve Ranavirus (Irıdoviridae)'ün ülkemiz de endemik tür, Tavas Kurbağası ( Rana tavasensis ) bireylerinde tespit edilmiştir. Ülkemizin biyoçeşitliliğin de önemli yeri olan kuyruksuz türlerin nesillerinin azaldığını ve tükenme tehlikesinde oldukları ortaya konulmuştır. In this study, the endemic derivatives Frog (Rana panasisis), which is in the Red List of the IUCN (National Union for Conservation of Nature) on the things that are living in Turkey and it is under construction, on what has actually been done - over time. related amphibian tracers Batrachochytrium Batrachochytrium (Frog virus) [Frog virus [Frog virus] images showing absence on or on primers of their equivalents on the genome. Two main pathogens considered responsible for extinction on amphibians as a research process; Batrachochytrium dendrobatidis (Bd) fungus and endemic species of Ranavirus (Iridoviridae), Pan-Frog (Rana Tavasensis) were detected. It reveals itself to be unable to meet a high level of biodiversity. Bu tez Çalışması; Afyon Kocatepe Üniversitesi Bilimsel Araştırma Projeleri Koordinasyon Birimi (BAPK) Tarafından Desteklenmiştir. Proje No: “19.SAĞ.BİL.16”

Recent evidence indicates that infectious diseases are one of the main causes of amphibian decline worldwide. Chytridiomycosis is an emergent disease caused by the waterborne fungus Batrachochytrium dendrobatidis (Bd). The infectious stage of this pathogen comprises motile zoospores that encyst in amphibian host to form a zoosporangium, from which new zoospores are subsequently released into the water. Therefore, the aquatic environment facilitates Bd transmission, suggesting that tadpoles could be more prone to infection than adults. However, unlike adults, infection during the amphibian larval stage is rarely lethal. This leads to a high prevalence of Bd in tadpoles, consequently acting as reservoirs for this pathogen. With this study, we aim to understand the effects of intrinsic and extrinsic factors on the prevalence of Bd in an assemblage of stream tadpoles in the Atlantic Forest, south-eastern Brazil. For Bd detection, we inspected individuals’ mouthparts searching for a specific pattern of depigmentation that has been widely associated with infection by this pathogen in tadpoles. Specimens presenting depigmentation compatible with the described patterns within the literature were classified as Bd-positive. We found 38.7% (209 out of 540) Bd-positive tadpoles. Slow-developing tadpoles presented a higher Bd prevalence compared to fast-developing tadpoles. Also, tadpoles inhabiting streams with lower temperatures and slower water flow presented high Bd prevalence regardless of their developmental duration. Our results suggest that slow-developing tadpoles can potentially act as effective Bd reservoir, and that cooler, slow-water streams could favour the pathogen maintenance in the environment.

Considering that emerging infectious diseases are one of the major drivers of global amphibian decline, controlling the spread of infections are evenmore challenging. Amphibian skin disease chytridiomycosis, which is caused by two species of fungi belonging to the Batrachochytrium genus, has been detected in at least 700 amphibian species causingmass mortalities in all continents where amphibian soccur. Most Alytes species, including the Beticmid wife toad (A. dickhilleni), are highly susceptible to B. dendrobatidis(Bd) with lethal consequences. The presence of Bd infection in A. dickhilleni was confirmed ten years ago in just three localities across the entire distribution range of this threatened species. Here we report the extraordinary Bd expansion through the entire distribution range of A. dickhilleniand analyse if former infected populations acted as the source of transmission events to current infected populations. Currently,Bd infection is broadly distributed across the entire distribution range of the species and the increase of infection prevalence reached 30–50% during a decade. The populations where the infection was detected a decade ago could be identified as likely sources of infection for somelocations where the pathogen is now present. The introduction of infected amphibian hosts into previously naïve A. dickhilleni breeding sites, and otheranthropogenic processes, are seeming to be the most plausible way of Bd range expansion, motivating mass mortalities, population declines and extirpation events of this threatened amphibian species., This work was funded by the Consejería de Medio Ambiente of Junta de Andalucía (Contract Number NET378406/1).

The skin microbiome is an important part of amphibian immune defenses and protects against pathogens such as the chytrid fungus Batrachochytrium dendrobatidis (Bd), which causes the skin disease chytridiomycosis. Alteration of the microbiome by anthropogenic factors, like pesticides, can impact this protective trait, disrupting its functionality. Chlorothalonil is a widely used fungicide that has been recognized as having an impact on amphibians, but so far, no studies have investigated its effects on amphibian microbial communities. In the present study, we used the amphibian Lithobates vibicarius from the montane forest of Costa Rica, which now appears to persist despite ongoing Bd-exposure, as an experimental model organism. We used 16S rRNA amplicon sequencing to investigate the effect of chlorothalonil on tadpoles’ skin microbiome. We found that exposure to chlorothalonil changes bacterial community composition, with more significant changes at a higher concentration. We also found that a larger number of bacteria were reduced on tadpoles’ skin when exposed to the higher concentration of chlorothalonil. We detected four presumed Bd-inhibitory bacteria being suppressed on tadpoles exposed to the fungicide. Our results suggest that exposure to a widely used fungicide could be impacting host-associated bacterial communities, potentially disrupting an amphibian protective trait against pathogens.

Environmental variation along elevational gradients shapes conditions for pathogen development, which influences disease outcomes. Chytridiomycosis is a non-vectored disease caused by the aquatic fungus Batrachochytrium dendrobatidis (Bd) and is responsible for massive declines of amphibian populations all over the world. Several biotic and abiotic factors are known to influence Bd infection dynamics in amphibians, including temperature and host species richness. Here, we quantified Bd prevalence and load along an elevational gradient in the Caparaó National Park (CNP), Brazil, and tested for associations of Bd infections with elevation, temperature, and species richness. We hypothesized that Bd infections would increase as local species richness decreased with elevation. We detected Bd along the entire elevational gradient and found a negative association between infection load and elevation. We did not detect significant associations between infection prevalence and elevation. Our findings are consistent with other wide elevational gradient studies, but are contrary to 2 other studies performed in the Atlantic Forest. We did not find the minimum elevational range that should be sampled to detect the influence of elevation on Bd variation. Our study represents the widest elevational gradient that has been sampled in Brazil and contributes to a better understanding of Bd distribution and dynamics in natural systems.

Chytridiomycosis is an emerging infectious disease affecting amphibians globally and it is caused by the fungal pathogen Batrachochytrium dendrobatidis (Bd). Chytridiomycosis has caused dramatic declines and even extinctions in wild amphibian populations in Europe, Australia, Central and North America. Spanning over two and a half decades, extensive research has led to discovery of epizootic and enzootic lineages of this pathogen. However, the Bd-amphibian system had garnered less attention in Asia until recently when an ancestral Bd lineage was identified in the Korean peninsula. Amphibians co-exist with the pathogen in Asia, only sub-lethal effects have been documented on hosts. Such regions are 'coldspots' of infection and are an important resource to understand the dynamics between the enzootic pathogen-Bd and its obligate host-amphibians. Insights into the biology of infection have provided new knowledge on the multi-faceted interaction of Bd in a hyperdiverse Asian amphibian community. We present the findings and highlight the knowledge gap that exists, and propose the ways to bridge them. We emphasize that chytridiomycosis in Asia is an important wildlife disease and it needs focussed research, as it is a dynamic front of pathogen diversity and virulence.

Chytridiomycosis, a leading cause for the global decline in the number of amphibians, is caused by the fungal pathogen Batrachochytrium dendrobatidis. In this study, global distribution data of B. dendrobatidis were collected from January 2009 to May 2019. Space-time scan statistics and the maximum entropy (MaxEnt) model were used to analyze the epidemic trends and aggregation of the pathogen, and predict B. dendrobatidis distribution through its relationships with climate factors, wind speed, and solar radiation. The results of space-time scan statistics show seven clusters of data for the distribution of B. dendrobatidis. The time was mainly concentrated in 2009, 2013, 2015, and 2016, and the regions were primarily concentrated in southeastern Canada, southwestern France, Nigeria, Cameroon, eastern Brazil, southeastern Brazil, central Madagascar, and central and eastern Australia. MaxEnt showed that annual precipitation had the largest contribution percentage in the model, and annual mean temperature highly influenced the distribution of B. dendrobatidis. The global high-risk areas of B. dendrobatidis distribution were mainly observed in western Canada, southern Brazil, Chile, the United Kingdom, Japan, the Republic of Korea, eastern South Africa, eastern Madagascar, southeastern Australia, and southern China.

Chytridiomycosis is a catastrophic disease currently decimating worldwide amphibian populations, caused by the panzootic chytrid fungus Batrachochytrium dendrobatidis. Massive species decline to extinction catalyzes radical changes in ecosystems globally, including the largest continuous rainforest ecosystem on Earth, the Amazon rainforest. Innovative research that aims to propose feasible mechanisms of mitigation and the origins of the disease is vital, including studies addressing climatic effects on the expansion of chytridiomycosis. Thus, this publication aims to provide a comprehensive review of: i) the current technologies used for B. dendrobatidis detection and monitoring, and ii) the known Neotropical amphibian's skin microbiota with anti-fungal properties against B. dendrobatidis. Several immunologic and DNA-based methods are discussed to understand the emerging fungal pathogens and their effects on the biosphere, which can help to mitigate the devastating ecological impacts of mass amphibian morbidity. The establishment of rapid and highly accurate B. dendrobatidis detection techniques and methods for monitoring amphibian's cutaneous microbiome is crucial in the fight against chytridiomycosis.

Wildlife disease is a major cause of global biodiversity loss. Amongst the most devastating is the disease chytridiomycosis, caused by the amphibian chytrid fungusBatrachochytrium dendrobatidis(Bd). This disease has contributed to declines and extinctions in hundreds of amphibian species, but not all species are affected equally. Some amphibian hosts are capable of carrying high levels ofBdinfection without population declines, acting as reservoir species for the pathogen and driving population declines in sympatric species. In Australia, several species have been proposed as reservoir species; however, our understanding ofBdis derived from studies that are highly geographically and taxonomically biased, and our ability to extrapolate from these systems is unknown. We examined the prevalence and intensity ofBdinfection in 3 frog species in a previously unstudied host-pathogen system in temperate eastern Australia: the Blue Mountains tree frogLitoria citropa, a poorly-known species predicted to be susceptible toBdinfection; and the common eastern frogletCrinia signiferaand the stony creek frogL. lesueuri, which have both been identified as reservoir species in other regions. We found thatL. citropaandL. lesueuriwere infected withBdat a high prevalence and often high intensity, while the reverse was true forC. signifera. All species were detected at moderate abundance and there was no evidence of morbidity and mortality. Our findings do not supportC. signiferaandL. lesueuribeing reservoir species in this system, highlighting the importance of region-specific studies to inform conservation management.

Animals challenged with disease may select specific habitat conditions that help prevent or reduce infection. Whereas preinfection avoidance of habitats with a high risk of disease exposure has been documented in both captive and free-ranging animals, evidence of switching habitats after infection to support the clearing of the infection is limited to laboratory experiments. The extent to which wild animals proximately modify habitat choices in response to infection status thus remains unclear. We investigated preinfection behavioral avoidance and postinfection habitat switching using wild, radio-tracked boreal toads (Anaxyrus boreas boreas) in a population challenged with Batrachochytrium dendrobatidis (Bd), a pathogenic fungus responsible for a catastrophic panzootic affecting hundreds of amphibian species worldwide. Boreal toads did not preemptively avoid microhabitats with conditions conducive to Bd growth. Infected individuals, however, selected warmer, more open habitats, which were associated with elevated body temperature and the subsequent clearing of infection. Our results suggest that disease can comprise an important selective pressure on animal habitat and space use. Habitat selection models, therefore, may be greatly improved by including variables that quantify infection risk and/or the infection status of individuals through time.

Fungal diseases have not been taken seriously in public health agendas as well as research priorities, despite of globally causing an estimated two million deaths every year, and the emergence of many troublesome fungal pathogens like Candida auris, azole resistant Aspergillus fumigatus, terbinafine and azole resistant dermatophytes, and zoonotic sporotrichosis in humans. Fungi are also responsible for huge losses of agricultural products and stored crops as well as recent massive and unexpected mortality in animals caused by white-nose syndrome in the bats and Chytridiomycosis in amphibians. This review aims to underscore the need for collaborative, multisectoral, and trans-disciplinary approach to include the One Health approach as an essential component of surveillance, prevention, and control of globally emerging fungal diseases. Rigorous evidence based surveillance of the environment as well as strengthening rapid and quality diagnosis of fungal diseases can save millions of lives and reduce significant morbidity.

The amphibian chytrid fungal pathogen (Batrachochytrium dendrobatidis, henceforth Bd) has had a devastating impact on biodiversity, causing the decline or extinction of over 500 amphibian species. Yet, our understanding of Bd transmission pathways remains incomplete, in particular for host species with weak aquatic associations, and between reservoir and susceptible host species. We examined Bd transmission from a potential reservoir host to a potentially susceptible critically endangered host; directly assessing the capacity of the former to transmit Bd, and the susceptibility of the latter as a Bd host. Using cohousing versus sequential use of the same enclosure by the two species, we distinguished the effects of direct versus indirect (environmental) transmission. Our study provides clear evidence that both direct and indirect terrestrial transmission from a reservoir to susceptible host species results in fatal chytridiomycosis. Transmission mode had no effect on overall morbidity or disease progression in the susceptible species. Our results demonstrate that reservoir and susceptible hosts do not need to be in the same place at the same time, or within an aquatic environment for transmission to occur. Our demonstration of indirect terrestrial transmission from a reservoir to susceptible host identifies mechanisms by which Bd may drive ongoing declines in populations where the pathogen is now endemic. Identifying these transmission pathways is important for understanding long-term extinction vulnerability of remnant populations of declining species challenged by disease.

The chytrid fungusBatrachochytrium dendrobatidis(Bd) infects numerous amphibian species worldwide and is suggested to drive population declines and extinction events. We report a study ofBdinfection at the northernmost distribution of the European yellow-bellied toadBombina variegata. A total of 577 individuals from ponds in 16 study sites were sampled for DNA andBdthroughout the breeding season. Microsatellite genotyping revealed 3 genetic clusters for the hostB. variegatawith an overall low genetic diversity. One of the clusters displayed a low microsatellite heterozygosity, a high inbreeding coefficient as well as highBdinfection prevalence and intensities. Multi-model estimates identified site, time of sampling, and heterozygosity to be important predictors of an individual’sBdinfection status, and identified a strong effect of site on individualBdinfection intensity. The study site effects are suggestive of localized infection peaks, and the increase of individualBdinfection probabilities towards the end of the sampling period suggests cumulative infection during the breeding season. This study highlights the need for regular monitoring ofBdinfection variables at multiple localities and times to gain insights intoBddynamics. Due to the detected relationship between individualBdinfection status and heterozygosity, conservation measures should focus on the maintenance of high genetic diversity and connectivity within and among amphibian populations.

Chytridiomycosis, caused by the fungus Batrachochytrium dendrobatidis (Bd), is a devastating infectious disease of amphibians. Retrospective studies using museum vouchers and genetic samples supported the hypothesis that Bd colonized Mexico from North America and then continued to spread into Central and South America, where it led to dramatic losses in tropical amphibian biodiversity (the epizootic wave hypothesis). While these studies suggest that Bd has been in Mexico since the 1970s, information regarding the historical and contemporary occurrence of different pathogen genetic lineages across the country is limited. In the current study, we investigated the historical and contemporary patterns of Bd in Mexico. We combined the swabbing of historical museum vouchers and sampling of wild amphibians with a custom Bd genotyping assay to assess the presence, prevalence, and genetic diversity of Bd over time in Mexico. We found Bd-positive museum specimens from the late 1800s, far earlier than previous records and well before recent amphibian declines. With Bd genotypes from samples collected between 1975-2019, we observed a contemporary dominance of the global panzootic lineage in Mexico and report four genetic subpopulations and potential for admixture among these populations. The observed genetic variation did not have a clear geographic signature or provide clear support for the epizootic wave hypothesis. These results provide a framework for testing new questions regarding Bd invasions and their temporal relationship to observed amphibian declines in the Americas.

Batrachochytrium salamandrivorans (Bsal), a pathogenic fungus causing the fatal disease chytridiomycosis in amphibians, was likely introduced to Europe through the trade in pet salamanders from Asia and then escaped into wild populations. Among European countries, Spain has a large number of private breeders and keepers of pet salamanders, and cases of Bsal in wild and captive populations already have been confirmed there. However, surveillance for the pathogen in Spanish collections of amphibians is sparse. Therefore, assisted by private owners and breeders, we surveyed 10 amphibian collections and analysed a total of 317 samples for presence of Bsal. All of our analyses yielded negative results. However, this apparent lack of Bsal cases in captivity should not encourage relaxation of vigilance, quarantine efforts or good practices. Because amphibian collections represent highly dynamic environments (animals are coming in and out), the pathogen could easily be introduced into a collection by new individuals. Any case of Bsal infection in captive animals could lead to its further spread to wild populations of susceptible species, potentially decimating them, and thus should be prevented.

Over the past half century, the chytridiomycosis panzootic has led to the decline of over 500 amphibian species with 90 attributed extinctions. Chytridiomycosis of amphibians is caused by two fungal species Batrachochytrium dendrobatidis (Bd) and Batrachochytrium salamandrivorans (Bsal). The genetic mechanisms underlying host-specificity and pathology in the Batrachochytrium genus remain elusive and their evolution and origins of virulence are largely unknown. Using deep nanopore sequencing, we found that Bsal is extremely repeat-rich with high numbers of long terminal repeats, long interspersed nuclear elements and transposable elements. This repeat-driven genome expansion in Bsal has resulted in a tripling of its length compared with Bd. Key pathogenicity genes including M36 metalloproteases have expanded compared with Bd, and are enriched for flanking transposable elements, suggesting its genome expansion is connected to selective evolutionary processes. Both batrachochytrids have evidence of a two speed genome architecture, including an enrichment of functional categories in compartments of repeat richness or sparsity. Furthermore, among Bd lineages, M36 metalloproteases with signatures of positive selection and, both in Bsal and Bd, genes upregulated during infection in vivo are enriched in repeat-rich and gene-sparse compartment of the genome. This is the first evidence for a two-speed genome in an animal pathogen, shedding new light on the role of repetitive sequences on the evolution of fungal pathogens driving global declines and extinctions of amphibians., {"references":["Scheele, B. C.et al., Science 363, 1459-1463 (2019)","Carey, C.et al., EcoHealth 3, 5-21 (2006)","Stegen, G.et al., Nature 544, 353-356 (2017)","Martel, A.et al., Proc. Natl. Acad. Sci. 110, 15325-15329 (2013)","Farrer, R. A. , Trends Microbiol. 27, 892-893 (2019)","Sabino-Pinto, J. et al. , AMRE 36, 411-416 (2015)","Wang, Y. et al. , New Phytol. 220, 922-935 (2018)","Zhang, S.-J. et al., Genomics Proteomics Bioinformatics 18, 321-332 (2020)","Wos, G. et al., Mob. DNA 12, 7 (2021)","Klein, S. J. & O'Neill, R. J. , Chromosome Res. 26, 5-23 (2018)","Oliver, K. R. & Greene, W. K. , BioEssays 31, 703-714 (2009)"]}

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.

While epizootics increasingly affect wildlife, it remains poorly understood how the environment shapes most host-pathogen systems. Here, we employ a three-step framework to study microclimate influence on ectotherm host thermal behaviour, focusing on amphibian chytridiomycosis in fire salamanders (Salamandra salamandra) infected with the fungal pathogen Batrachochytrium salamandrivorans (Bsal). Laboratory trials reveal that innate variation in thermal preference, rather than behavioural fever, can inhibit infection and facilitate salamander recovery under humidity-saturated conditions. Yet, a 3-year field study and a mesocosm experiment close to the invasive Bsal range show that microclimate constraints suppress host thermal behaviour favourable to disease control. A final mechanistic model, that estimates range-wide, year-round host body temperature relative to microclimate, suggests that these constraints are rule rather than exception. Our results demonstrate how innate host defences against epizootics may remain constrained in the wild, which predisposes to range-wide disease outbreaks and population declines.

Understanding host immune function and ecoimmunology is increasingly important at a time when emerging infectious diseases (EIDs) threaten wildlife. One EID that has emerged and spread widely in recent years is chytridiomycosis, caused by the fungal pathogen Batrachochytrium dendrobatidis (Bd), which is implicated unprecedented amphibian declines around the world. The impacts of Bd have been severe for many amphibian species, but some populations have exhibited signs of persistence, and even recovery, in some regions. Many mechanisms may underpin this pattern and amphibian immune responses are likely one key component. Although we have made great strides in understanding amphibian immunity, the complement system remains poorly understood. The complement system is a nonspecific, innate immune defense that is known to enhance other immune responses. Complement activation can occur by three different biochemical pathways and result in protective mechanisms, such as inflammation, opsonization, and pathogen lysis, thereby providing protection to the host. We currently lack an understanding of complement pathway activation for chytridiomycosis, but several studies have suggested that it may be a key part of an early and robust immune response that confers host resistance. Here, we review the available research on the complement system in general as well as amphibian complement responses to Bd infection. Additionally, we propose future research directions that will increase our understanding of the amphibian complement system and other immune responses to Bd. Finally, we suggest how a deeper understanding of amphibian immunity could enhance the conservation and management of amphibian species that are threatened by chytridiomycosis., The fungal pathogen Batrachochytrium dendrobatidis (Bd), is implicated in worldwide amphibian declines. The complement system is a nonspecific, innate immune defense that is known to enhance other immune responses. Here, we address what is known about the complement system in amphibians and how it may respond to infection with Bd., Highlights The complement system is an immune component present in vertebrates, but little is known about its role in amphibian chytridiomycosis.We review recent research, current gaps in knowledge, and we suggest future research to fill these gaps.

Wildlife diseases are contributing to the current Earth’s sixth mass extinction; one disease, chytridiomycosis, has caused mass amphibian die-offs. While global spread of a hypervirulent lineage of the fungus Batrachochytrium dendrobatidis (BdGPL) causes unprecedented loss of vertebrate diversity by decimating amphibian populations, its impact on amphibian communities is highly variable across regions. Here, we combine field data with in vitro and in vivo trials that demonstrate the presence of a markedly diverse variety of low virulence isolates of BdGPL in northern European amphibian communities. Pre-exposure to some of these low virulence isolates protects against disease following subsequent exposure to highly virulent BdGPL in midwife toads (Alytes obstetricans) and alters infection dynamics of its sister species B. salamandrivorans in newts (Triturus marmoratus), but not in salamanders (Salamandra salamandra). The key role of pathogen virulence in the complex host-pathogen-environment interaction supports efforts to limit pathogen pollution in a globalized world., The pathogen Batrachochytrium dendrobatidis (BD) associated with widespread amphibian declines is present in Europe but has not consistently caused disease-induced declines in that region. Here, the authors suggest that an endemic strain of BD with low virulence may protect the hosts upon co-infection with more virulent strains.

Environmental DNA (eDNA) detection is a valuable conservation tool that can be used to identify and monitor imperiled or invasive species and wildlife pathogens. Batrachochytrium pathogens are of global conservation concern because they are a leading cause of amphibian decline. While eDNA techniques have been used to detect Batrachochytrium DNA in the environment, a systematic comparison of extraction methods across environmental samples is lacking. In this study, we first compared eDNA extraction methods and found that a soil extraction kit (Qiagen PowerSoil) was the most effective for detecting Batrachochytrium dendrobatidis in water samples. The PowerSoil extraction had a minimum detection level of 100 zoospores and had a two- to four-fold higher detection probability than other commonly used extraction methods (e.g., QIAamp extraction, DNeasy+Qiashredder extraction method, respectively). Next, we used this extraction method on field-collected water and sediment samples and were able to detect pathogen DNA in both. While field-collected water filters were equivalent to amphibian skin swab samples in detecting the presence of pathogen DNA, the seasonal patterns in pathogen quantity were different between skin swabs and water samples. Detection rate was lowest in sediment samples. We also found that detection probability increases with the volume of water filtered. Our results indicate that water filter eDNA samples can be accurate in detecting pathogen presence at the habitat scale but their utility for quantifying pathogen loads in the environment appears limited. We suggest that eDNA techniques be used for early warning detection to guide animal sampling efforts.

The disease chytridiomycosis caused by the fungus Bd has devastated amphibian populations worldwide. Functional genomic contributions to host susceptibility remain enigmatic and vary between species and populations. We conducted experimental Bd infections in Rana yavapaiensis, a species with intraspecific variation in chytridiomycosis susceptibility, to assess the skin and spleen transcriptomic response to infection over time. We predicted that increased immune gene expression would be associated with a positive disease outcome, but we instead found that surviving frogs had significantly reduced immune gene expression compared to susceptible frogs and to uninfected controls. MHC class IIβ gene expression was also significantly higher in susceptible frogs compared to surviving frogs. Furthermore, susceptible frogs expressed a significantly larger number of distinct class IIβ alleles, demonstrating a negative correlation between class IIβ expression, functional diversity, and survival. Expression of the MHC class IIβ locus previously associated with Bd disease outcomes was a significant predictor of Bd infection intensity at early infection stages but not at late infection stages, suggesting initial MHC-linked immune processes are important for ultimate disease outcomes. We infer through disease association and phylogenetic analysis that certain MHC variants are linked to the immune expression that was negatively associated with survival, and we hypothesize that frogs that did not express these alleles could better survive infections. Our study finds that MHC expression at early and late infection stages predicts Bd infection intensity, and suggests that generating a sustained immune response against Bd may be counterproductive for surviving chytridiomycosis in this partially susceptible species.

Microorganisms are increasingly recognized as ecosystem-relevant components because they affect the population dynamics of hosts. Functioning at the interface of the host and pathogen, skin and gut microbiomes are vital components of immunity. Recent work reveals a strong influence of biotic and abiotic environmental factors (including the environmental microbiome) on disease dynamics, yet the importance of the host-host microbiome-pathogen-environment interaction has been poorly reflected in theory. We use amphibians and the disease chytridiomycosis caused by the fungal pathogen Batrachochytrium dendrobatidis to show how interactions between host, host microbiome, pathogen, and the environment all affect disease outcome. Our review provides new perspectives that improve our understanding of disease dynamics and ecology by incorporating environmental factors and microbiomes into disease theory.

Light gradients are an inherent feature in aquatic ecosystems and play a key role in shaping the biology of phytoplankton. Parasitism by chytrid fungi is gaining increasing attention as a major control agent of phytoplankton due to its previously overlooked ubiquity, and profound ecological and evolutionary consequences. Despite this interest, if and how light conditions modulate phytoplankton chytridiomycosis remains poorly studied. We investigated life-history traits of a chytrid parasite, Rhizophydium megarrhizum, under different light intensities and spectral compositions when infecting two closely related planktonic cyanobacteria with different light-harvesting strategies: Planktothrix rubescens and P. agardhii. In general, parasite transmission was highest under light conditions (both intensity and quality) that maximized growth rates for uninfected cyanobacteria. Chytrid encystment on hosts was significantly affected by light intensity and host strain identity. This likely resulted from higher irradiances stimulating the increased discharge of photosynthetic by-products, which drive parasite chemotaxis, and from strain-specific differences at the cell-surface. Comparisons of parasite transmission and host growth rates under different light conditions suggest the potential for epidemic development at higher irradiances, whereas host and parasite could coexist without epidemic outbreaks at lower light levels. These results illustrate the close relationship between parasite transmission and host fitness, which is ultimately modulated by the external environment.

Emerging infectious diseases can cause dramatic declines in wildlife populations. Sometimes, these declines are followed by recovery, but many populations do not recover. Studying differential recovery patterns may yield important information for managing disease-afflicted populations and facilitating population recoveries. In the late 1980s, a chytridiomycosis outbreak caused multiple frog species in Australia’s Wet Tropics to decline. Populations of some species (e.g., Litoria nannotis) subsequently recovered, while others (e.g., Litoria dayi) did not. We examined the population genetics and current infection status of L. dayi, to test several hypotheses regarding the failure of its populations to recover: (1) a lack of individual dispersal abilities has prevented recolonization of previously occupied locations, (2) a loss of genetic variation has resulted in limited adaptive potential, and (3) L. dayi is currently adapting to chytridiomycosis. We found moderate-to-high levels of gene flow and diversity (Fst range

Emerging infectious diseases are responsible for declines in wildlife populations around the globe. Mass mortality events associated with emerging infectious diseases are often associated with high number of infected individuals (prevalence) and high pathogen loads within individuals (intensity). At the landscape scale, spatial and temporal variation in environmental conditions can alter the relationship between these infection parameters and blur the overall picture of disease dynamics. Quantitative estimates of how infection parameters covary with environmental heterogeneity at the landscape scale are scarce. If we are to identify wild populations at risk of disease epidemics, we must elucidate the factors that shape, and potentially decouple, the link between pathogen prevalence and intensity of infection over complex ecological landscapes. Using a network of 41 populations of the amphibian host Rana pipiens in Ontario, Canada, we present the spatial and temporal heterogeneity in pathogen prevalence and intensity of infection of the chytrid fungus Batrachochytrium dendrobatidis (Bd), across a 3-year period. We then quantify how covariation between both infection parameters measured during late summer is modified by previously experienced spatiotemporal environmental heterogeneity across 14 repeat sampled populations. Late summer Bd infection parameters are governed, at least in part, by different environmental factors operating during separate host life-history events. Our results provide evidence for a relationship between Bd prevalence and thermal regimes prior to host breeding at the site level, and a relationship between intensity of infection and aquatic conditions (precipitation, hydroshed size and river density) throughout host breeding period at the site level. This demonstrates that microclimatic variation within temporal windows can drive divergent patterns of pathogen dynamics within and across years, by effecting changes in host behaviour which interfere with the pathogen's ability to infect and re-infect hosts. A clearer understanding of the role that spatiotemporal heterogeneity has upon infection parameters will provide valuable insights into host-pathogen epidemiology, as well as more fundamental aspects of the ecology and evolution of interspecific interactions.

Background Emerging infectious diseases (EIDs) are contributing to species die-offs worldwide. We can better understand EIDs by using ecological approaches to study pathogen biology. For example, pathogens are exposed to variable temperatures across daily, seasonal, and annual scales. Exposure to temperature fluctuations may reduce pathogen growth and reproduction, which could affect pathogen virulence, transmission, and environmental persistence with implications for disease. We examined the effect of a variable thermal environment on reproductive life history traits of the fungal pathogen Batrachochytrium dendrobatidis (Bd). Bd causes chytridiomycosis, an emerging infectious disease of amphibians. As a pathogen of ectothermic hosts, Bd can be exposed to large temperature fluctuations in nature. To determine the effect of fluctuating temperatures on Bd growth and reproduction, we collected temperature data from breeding pools of the Yosemite toad (Anaxyrus canorus), a federally threatened species that is susceptible to chytridiomycosis. We cultured Bd under a daily fluctuating temperature regime that simulated Yosemite toad breeding pool temperatures and measured Bd growth, reproduction, fecundity, and viability. Results We observed decreased Bd growth and reproduction in a diurnally fluctuating thermal environment as compared to cultures grown at constant temperatures within the optimal Bd thermal range. We also found that Bd exhibits temperature-induced trade-offs under constant low and constant high temperature conditions. Conclusions Our results provide novel insights on variable responses of Bd to dynamic thermal conditions and highlight the importance of incorporating realistic temperature fluctuations into investigations of pathogen ecology and EIDs.

The mechanisms by which invasive species negatively affect native species include competition, predation, and the introduction of novel pathogens. Moreover, if an invasive species is a competent disease reservoir, it may facilitate the long-term maintenance and spread of pathogens in ecological assemblages and drive the extinction of less tolerant or less resistant species. Disease-driven loss of biodiversity is exemplified by the amphibian–chytrid fungus system. The disease chytridiomycosis is caused by the aquatic chytrid fungus Batrachochytrium dendrobatidis (Bd) in anurans and is associated with worldwide amphibian population declines and extinctions. For amphibian species that metamorphose and leave infected aquatic habitats, the mechanisms by which Bd persists over winter in these habitats remains a critical open question. A leading hypothesis is that American bullfrogs (Rana catesbeiana), a worldwide invasive species, are tolerant to Bd and serve as a reservoir host for Bd during winter months and subsequently infect native species that return to breed in spring. Using outdoor mesocosms, we experimentally examined if two strains of Bd could overwinter in aquatic systems, in the presence or absence of bullfrog tadpoles, and if overwintered Bd could be transmitted to tadpoles of two spring-breeding species: Pacific treefrogs (Pseudacris regilla) and Cascades frogs (Rana cascadae). We found that only 4 of 448 total animals (one bullfrog and three spring breeders) tested positive for Bd after overwintering. Moreover, two of the three infected spring breeders emerged from tanks that contained overwintered Bd but in the absence of infected bullfrogs. This suggests that Bd can persist over winter without bullfrogs as a reservoir host. We found no effect of Bd strain on bullfrog survival after overwintering. For Pacific treefrogs, Bd exposure did not significantly affect mass at or time to metamorphosis while exposure to bullfrogs reduced survival. For Cascades frogs, we found an interactive effect of Bd strain and bullfrog presence on time to metamorphosis, but no main or interactive effects on their survival or mass at metamorphosis. In short, bullfrog tadpoles rarely retained and transmitted Bd infection in our experiment and we found limited evidence that Bd successfully overwinters in the absence of bullfrog tadpoles and infects spring-breeding amphibians.

The emerging fungal pathogen Batrachochytrium salamandrivorans (Bsal) is a major threat to amphibian species worldwide with potential to infect many species if it invades salamander biodiversity hotspots in the Americas. Bsal can cause the disease chytridiomycosis, and it is important to assess the risk of Bsal-induced chytridiomycosis to species in North America. We evaluated the susceptibility to Bsal of the common and widespread spotted salamander, Ambystoma maculatum, across life history stages and monitored the effect of Bsal exposure on growth rate and response of the stress hormone, corticosterone. We conclude that spotted salamanders appear resistant to Bsal because they showed no indication of disease or infection, and experienced minor effects on growth upon exposure. While we focused on a single population for this study, results were consistent across conditions of exposure including high or repeated doses of Bsal, life-stage at exposure, environmental conditions including two temperatures and two substrates, and promoting pathogen infectivity by conditioning Bsal cultures with thyroid hormone. Exposure to high levels of Bsal elicited an acute but not chronic increase in corticosterone in spotted salamanders, and reduced growth. We hypothesize that the early acute increase in corticosterone facilitated mounting an immune response to the pathogen, perhaps through immunoredistribution to the skin, but further study is needed to determine immune responses to Bsal. These results will contribute to development of appropriate Bsal management plans to conserve species at risk of emerging disease.