Prevalence and genetic diversity of Batrachochytrium dendrobatidis in Central African island and continental amphibian communities

The fungal pathogen Batrachochytrium dendrobatidis (Bd) infects hundreds of amphibian species and is implicated in global amphibian declines. Bd is comprised of several lineages that differ in pathogenicity, thus, identifying which Bd strains are present in a given amphibian community is essential for understanding host–pathogen dynamics. The presence of Bd has been confirmed in Central Africa, yet vast expanses of this region have not yet been surveyed for Bd prevalence, and the genetic diversity of Bd is largely unknown in this part of the world. Using retrospective surveys of museum specimens and contemporary field surveys, we estimated the prevalence of Bd in Central African island and continental amphibian assemblages, and genotyped strains of Bd present in each community. Our sampling of museum specimens included just a few individuals collected in the Gulf of Guinea archipelago prior to 1998, yet one of these individuals was Bd‐positive indicating that the pathogen has been on Bioko Island since 1966. We detected Bd across all subsequent sample years in our study and found modest support for a relationship between host life history and Bd prevalence, a positive relationship between prevalence and host community species richness, and no significant relationship between elevation and prevalence. The Global Panzootic Lineage (BdGPL) was present in all the island and continental amphibian communities we surveyed. Our results are consistent with a long‐term and widespread distribution of Bd in amphibian communities of Gabon and the Gulf of Guinea archipelago.     

The thermal mismatch hypothesis explains host susceptibility to an emerging infectious disease

Parasites typically have broader thermal limits than hosts, so large performance gaps between pathogens and their cold‐ and warm‐adapted hosts should occur at relatively warm and cold temperatures, respectively. We tested this thermal mismatch hypothesis by quantifying the temperature‐dependent susceptibility of cold‐ and warm‐adapted amphibian species to the fungal pathogen Batrachochytrium dendrobatidis (Bd) using laboratory experiments and field prevalence estimates from 15 410 individuals in 598 populations. In both the laboratory and field, we found that the greatest susceptibility of cold‐ and warm‐adapted hosts occurred at relatively warm and cool temperatures, respectively, providing support for the thermal mismatch hypothesis. Our results suggest that as climate change shifts hosts away from their optimal temperatures, the probability of increased host susceptibility to infectious disease might increase, but the effect will depend on the host species and the direction of the climate shift. Our findings help explain the tremendous variation in species responses to Bd across climates and spatial, temporal and species‐level variation in disease outbreaks associated with extreme weather events that are becoming more common with climate change.     

Host‐specific thermal profiles affect fitness of a widespread pathogen

Host behavior can interact with environmental context to influence outcomes of pathogen exposure and the impact of disease on species and populations. Determining whether the thermal behaviors of individual species influence susceptibility to disease can help enhance our ability to explain and predict how and when disease outbreaks are likely to occur. The widespread disease chytridiomycosis (caused by the fungal pathogen Batrachochytrium dendrobatidis, Bd) often has species‐specific impacts on amphibian communities; some host species are asymptomatic, whereas others experience mass mortalities and population extirpation. We determined whether the average natural thermal regimes experienced by sympatric frog species in nature, in and of themselves, can account for differences in vulnerability to disease. We did this by growing Bd under temperatures mimicking those experienced by frogs in the wild. At low and high elevations, the rainforest frogs Litoria nannotis, L. rheocola, and L. serrata maintained mean thermal regimes within the optimal range for pathogen growth (15–25°C). Thermal regimes for L. serrata, which has recovered from Bd‐related declines, resulted in slower pathogen growth than the cooler and less variable thermal regimes for the other two species, which have experienced more long‐lasting declines. For L. rheocola and L. serrata, pathogen growth was faster in thermal regimes corresponding to high elevations than in those corresponding to low elevations, where temperatures were warmer. For L. nannotis, which prefers moist and thermally stable microenvironments, pathogen growth was fastest for low‐elevation thermal regimes. All of the thermal regimes we tested resulted in pathogen growth rates equivalent to, or significantly faster than, rates expected from constant‐temperature experiments. The effects of host body temperature on Bd can explain many of the broad ecological patterns of population declines in our focal species, via direct effects on pathogen fitness. Understanding the functional response of pathogens to conditions experienced by the host is important for determining the ecological drivers of disease outbreaks.     

Design‐ and model‐based recommendations for detecting and quantifying an amphibian pathogen in environmental samples

Accurate pathogen detection is essential for developing management strategies to address emerging infectious diseases, an increasingly prominent threat to wildlife. Sampling for free‐living pathogens outside of their hosts has benefits for inference and study efficiency, but is still uncommon. We used a laboratory experiment to evaluate the influences of pathogen concentration, water type, and qPCR inhibitors on the detection and quantification of Batrachochytrium dendrobatidis (Bd) using water filtration. We compared results pre‐ and post‐inhibitor removal, and assessed inferential differences when single versus multiple samples were collected across space or time. We found that qPCR inhibition influenced both Bd detection and quantification in natural water samples, resulting in biased inferences about Bd occurrence and abundance. Biases in occurrence could be mitigated by collecting multiple samples in space or time, but biases in Bd quantification were persistent. Differences in Bd concentration resulted in variation in detection probability, indicating that occupancy modeling could be used to explore factors influencing heterogeneity in Bd abundance among samples, sites, or over time. Our work will influence the design of studies involving amphibian disease dynamics and studies utilizing environmental DNA (eDNA) to understand species distributions.     

Resistance,tolerance and environmental transmission dynamics determine host extinction risk in a load‐dependent amphibian disease

While disease‐induced extinction is generally considered rare, a number of recently emerging infectious diseases with load‐dependent pathology have led to extinction in wildlife populations. Transmission is a critical factor affecting disease‐induced extinction, but the relative importance of transmission compared to load‐dependent host resistance and tolerance is currently unknown. Using a combination of models and experiments on an amphibian species suffering extirpations from the fungal pathogen Batrachochytrium dendrobatidis (Bd), we show that while transmission from an environmental Bd reservoir increased the ability of Bd to invade an amphibian population and the extinction risk of that population, Bd‐induced extinction dynamics were far more sensitive to host resistance and tolerance than to Bd transmission. We demonstrate that this is a general result for load‐dependent pathogens, where non‐linear resistance and tolerance functions can interact such that small changes in these functions lead to drastic changes in extinction dynamics.     

Incapacitating effects of fungal coinfection in a novel pathogen system

As globalization lowers geographic barriers to movement, coinfection with novel and enzootic pathogens is increasingly likely. Novel and enzootic pathogens can interact synergistically or antagonistically, leading to increased or decreased disease severity. Here we examine host immune responses to coinfection with two closely related fungal pathogens: Batrachochytrium dendrobatidis (Bd) and Batrachochytrium salamandrivorans (Bsal). Both pathogens have had detrimental effects on amphibian populations, with Bd now largely enzootic, while Bsal is currently spreading and causing epizootics. Recent experimental work revealed that newts coinfected with Bd and Bsal had significantly higher mortality than those infected with either pathogen alone. Here we characterize host immunogenomic responses to chytrid coinfection relative to single infection. Across several classes of immune genes including pattern recognition receptors, cytokines, and MHC, coinfected host gene expression was weakly upregulated or comparable to that seen in single Bd infection, but significantly decreased when compared to Bsal infection. Combined with strong complement pathway downregulation and keratin upregulation, these results indicate that coinfection with Bd and Bsal compromises immune responses active against Bsal alone. As Bsal continues to invade naïve habitats where Bd is enzootic, coinfection will be increasingly common. If other Bd‐susceptible species in the region have similar responses, interactions between the two pathogens could cause severe population and community‐level declines.     

Precipitation Constrains Amphibian Chytrid Fungus Infection Rates in a Terrestrial Frog Assemblage in Jamaica,West Indies

We model Batrachochytrium dendrobatidis (Bd) infection rates in Jamaican frogs—one of the most threatened amphibian fauna in the world. The majority of species we surveyed were terrestrial direct‐developing frogs or frogs that breed in tank bromeliads, rather than those that use permanent water bodies to breed. Thus, we were able to investigate the climatic correlates of Bd infection in a frog assemblage that does not rely on permanent water bodies. We sampled frogs for Bd across all of the major habitat types on the island, used machine learning algorithms to identify climatic variables that are correlated with infection rates, and extrapolated infection rates across the island. We compared the effectiveness of the machine learning algorithms for species distribution modeling in the context of our study, and found that infection rate rose quickly with precipitation in the driest month. Infection rates also increased with mean temperature in the warmest quarter until 22 °C, and remained relatively level thereafter. Both of these results are in accordance with previous studies of the physiology of Bd. Based on our environmental results, we suggest that frogs occupying high‐precipitation habitats with cool rainy‐season temperatures, though zcurrently experiencing low frequencies of infection, may experience an increase in infection rates as global warming increases temperatures in their habitat.     

After the epizootic: Host–pathogen dynamics in montane tropical amphibian communities with high prevalence of chytridiomycosis

The amphibian fungal disease chytridiomycosis, caused by Batrachochytrium dendrobatidis (Bd), poses a great threat to global amphibian biodiversity. In Peruvian cloud forests of the Kosñipata Valley of Manu National Park where chytrid infection is highly prevalent, we have monitored species-rich amphibian communities since 1996. An epizootic of chytridiomycosis is thought to have caused the disappearance of 35% of species richness in the early 2000s. We investigated the post-epizootic Bd prevalence and infection intensity within the remnant amphibian community from 2008 to 2015, and modeled Bd dynamics as a function of species, season, reproductive mode, life stage, and elevation. Prevalence was higher in 2012–2015 than in 2008–2009, but overall prevalence has remained fairly constant (~50%) post-epizootic. We also found that while prevalence decreased with elevation during the wet season, it generally increased with elevation during the dry season, potentially due to seasonal changes in temperature and precipitation. In aquatic habitats, Bd is likely maintained through a single, stream-breeding, putative reservoir species (which survived epizootics, in contrast to other aquatic-breeding species). The now-dominant terrestrial-breeding species allow Bd to persist and spread in terrestrial habitats, possibly through individual dispersal into naïve areas. We conclude that Bd prevalence in the Kosñipata Valley has stabilized over time, suggesting that Bd is now enzootic. Long-term monitoring of host infection is important because temporal changes in prevalence and infection intensity can cause changes in host species richness and abundance, which in turn may alter the trajectory of host–pathogen dynamics.     

Mitigating amphibian chytridiomycosis with bioaugmentation: characteristics of effective probiotics and strategies for their selection and use

Probiotic therapy through bioaugmentation is a feasible disease mitigation strategy based on growing evidence that microbes contribute to host defences of plants and animals. Amphibians are currently threatened by the rapid global spread of the pathogen, Batrachochytrium dendrobatidis (Bd), which causes the disease chytridiomycosis. Bioaugmentation of locally occurring protective bacteria on amphibians has mitigated this disease effectively in laboratory trials and one recent field trial. Areas still naïve to Bd provide an opportunity for conservationists to proactively implement probiotic strategies to prevent further amphibian declines. In areas where Bd is endemic, bioaugmentation can facilitate repatriation of susceptible amphibians currently maintained in assurance colonies. Here, we synthesise the current research in amphibian microbial ecology and bioaugmentation to identify characteristics of effective probiotics in relation to their interactions with Bd, their host, other resident microbes and the environment. To target at‐risk species and amphibian communities, we develop sampling strategies and filtering protocols that result in probiotics that inhibit Bd under ecologically relevant conditions and persist on susceptible amphibians. This filtering tool can be used proactively to guide amphibian disease mitigation and can be extended to other taxa threatened by emerging infectious diseases.     

Long‐term endemism of two highly divergent lineages of the amphibian‐killing fungus in the Atlantic Forest of Brazil

The recent global spread of the amphibian‐killing fungus [Batrachochytrium dendrobatidis (Bd)] has been closely tied to anthropogenic activities; however, regional patterns of spread are not completely understood. Using historical samples, we can test whether Bd was a spreading or endemic pathogen in a region within a particular time frame, because those two disease states provide different predictions for the regional demographic dynamics and population genetics of Bd. Testing historical patterns of pathogen prevalence and population genetics under these predictions is key to understanding the evolution and origin of Bd. Focusing on the Atlantic Forest (AF) of Brazil, we used qPCR assays to determine the presence or absence of Bd on 2799 preserved postmetamorphic anurans collected between 1894 and 2010 and used semi‐nested PCRs to determine the frequency of rRNA ITS1 haplotypes from 52 samples. Our earliest date of detection was 1894. A mean prevalence of 23.9% over time and spatiotemporal patterns of Bd clusters indicate that Bd has been enzootic in the Brazilian AF with no evidence of regional spread within the last 116 years. ITS1 haplotypes confirm the long‐term presence of two divergent strains of Bd (BdGPL and Bd‐Brazil) and three spatiotemporally broad genetic demes within BdGPL, indicating that Bd was not introduced into southeast Brazil by the bullfrog trade. Our data show that the evolutionary history and pathogen dynamics of Bd in Brazil is better explained by the endemic pathogen hypothesis.     

Amphibian‐killing chytrid in Brazil comprises both locally endemic and globally expanding populations

Chytridiomycosis, caused by the fungus Batrachochytrium dendrobatidis (Bd), is the emerging infectious disease implicated in recent population declines and extinctions of amphibian species worldwide. Bd strains from regions of disease‐associated amphibian decline to date have all belonged to a single, hypervirulent clonal genotype (Bd‐GPL). However, earlier studies in the Atlantic Forest of southeastern Brazil detected a novel, putatively enzootic lineage (Bd‐Brazil), and indicated hybridization between Bd‐GPL and Bd‐Brazil. Here, we characterize the spatial distribution and population history of these sympatric lineages in the Brazilian Atlantic Forest. To investigate the genetic structure of Bd in this region, we collected and genotyped Bd strains along a 2400‐km transect of the Atlantic Forest. Bd‐Brazil genotypes were restricted to a narrow geographic range in the southern Atlantic Forest, while Bd‐GPL strains were widespread and largely geographically unstructured. Bd population genetics in this region support the hypothesis that the recently discovered Brazilian lineage is enzootic in the Atlantic Forest of Brazil and that Bd‐GPL is a more recently expanded invasive. We collected additional hybrid isolates that demonstrate the recurrence of hybridization between panzootic and enzootic lineages, thereby confirming the existence of a hybrid zone in the Serra da Graciosa mountain range of Paraná State. Our field observations suggest that Bd‐GPL may be more infective towards native Brazilian amphibians, and potentially more effective at dispersing across a fragmented landscape. We also provide further evidence of pathogen translocations mediated by the Brazilian ranaculture industry with implications for regulations and policies on global amphibian trade.     

Widespread historical presence of Batrachochytrium dendrobatidis in African pipid frogs

Aim Amphibian chytridiomycosis, an emerging infectious disease caused by the chytrid fungus Batrachochytrium dendrobatidis (Bd), is associated with global amphibian population declines and species extinctions. Current evidence indicates that the pathogen has recently spread globally from an enzootic focus, with Xenopus spp. (family Pipidae) in South Africa having been identified as a likely source. The aim of this study was to investigate further the likelihood of African Xenopus spp. as the original source of Bd. Location We examined 665 museum specimens of 20 species of African and South American pipid frogs collected between 1844 and 1994 and held in the collection of the Natural History Museum, London. Methods Skin brushings taken from adult amphibians and brushings from the mouthparts, lips and developing hind limbs of larval pipid frogs were examined for the presence of Bd using real‐time PCR. Results We found six cases of Bd infection in three Xenopus spp. (from Africa), but none of the South American pipids was positive, although only 45 South American frogs were available for examination. The earliest case of Bd infection was in a specimen of Xenopus fraseri collected from Cameroon in 1933. A consistently low prevalence of infection over time indicates that a historical equilibrium existed between Xenopus spp. and Bd infection in Africa. Main conclusions Our results suggest that Bd infection was present in Xenopus spp. across sub‐Saharan Africa by the 1930s, providing additional support for the ‘out of Africa’ hypothesis. If this hypothesis is correct, it strengthens the argument for stringent control of human‐assisted movements of amphibians and other wildlife world‐wide to minimize the likelihood of pathogen introduction and disease emergence that can threaten species globally. Our findings help inform species selection for conservation in the face of the current Bd pandemic and also guide future research directions for selecting Bd isolates for sequencing and virulence testing.     

Impacts of thermal mismatches on chytrid fungus Batrachochytrium dendrobatidis prevalence are moderated by life stage,body size,elevation and latitude

Global climate change is increasing the frequency of unpredictable weather conditions; however, it remains unclear how species‐level and geographic factors, including body size and latitude, moderate impacts of unusually warm or cool temperatures on disease. Because larger and lower‐latitude hosts generally have slower acclimation times than smaller and higher‐latitude hosts, we hypothesised that their disease susceptibility increases under ‘thermal mismatches’ or differences between baseline climate and the temperature during surveying for disease. Here, we examined how thermal mismatches interact with body size, life stage, habitat, latitude, elevation, phylogeny and International Union for Conservation of Nature (IUCN) conservation status to predict infection prevalence of the chytrid fungus Batrachochytrium dendrobatidis (Bd) in a global analysis of 32 291 amphibian hosts. As hypothesised, we found that the susceptibility of larger hosts and hosts from lower latitudes to Bd was influenced by thermal mismatches. Furthermore, hosts of conservation concern were more susceptible than others following thermal mismatches, suggesting that thermal mismatches might have contributed to recent amphibian declines.     

Confronting inconsistencies in the amphibian‐chytridiomycosis system: implications for disease management

Chytridiomycosis, caused by the pathogenic fungus Batrachochytrium dendrobatidis (Bd), is one of the largest threats to wildlife and is putatively linked to the extirpation of numerous amphibians. Despite over a decade of research on Bd, conflicting results from a number of studies make it difficult to forecast where future epizootics will occur and how to manage this pathogen effectively. Here, we emphasize how resolving these conflicts will advance Bd management and amphibian conservation efforts. We synthesize current knowledge on whether Bd is novel or endemic, whether amphibians exhibit acquired resistance to Bd, the importance of host resistance versus tolerance to Bd, and how biotic (e.g. species richness) and abiotic factors (e.g. climate change) affect Bd abundance. Advances in our knowledge of amphibian–chytrid interactions might inform the management of fungal pathogens in general, which are becoming more common and problematic globally.     

Current and predicted distribution of the pathogenic fungus Batrachochytrium dendrobatidis in Colombia,a hotspot of amphibian biodiversity

Global amphibian declines have been attributed to several factors including the chytrid fungal pathogen, Batrachochytrium dendrobatidis (Bd), that infects hosts’ skin and causes death by inhibiting immune response and impairing osmoregulatory function. Here, we integrate extensive new field data with previously published locality records of Bd in Colombia, a megadiverse and environmentally heterogeneous country in northwestern South America, to determine the relative importance of environmental variables and reproductive mode for predicting the risk of Bd infection in amphibians. We surveyed 81 localities across Colombia and sampled 2876 individual amphibians belonging to 14 taxonomic families. Through a combination of end‐point PCR and real‐time PCR analyses, Bd was detected in 338 individuals (12%) representing 43 localities (53%) distributed from sea level to 3200 m. We found that annual mean temperature and variables related with seasonality in precipitation and temperature appeared to define the most suitable areas for the establishment of the pathogen. In addition, prevalence of infection appeared to be higher in species with a terrestrial reproductive mode. Our study provides the first large‐scale study of the current and potential distribution of Bd in the biodiversity hotspot centered on Colombia. We hope the newly provided information on the extent of the distribution of the pathogen and the potential areas where Bd may impact the amphibian fauna will inform decision making by environmental authorities and future conservation action.     

Effects of amphibian phylogeny,climate and human impact on the occurrence of the amphibian‐killing chytrid fungus

Chytridiomycosis, due to the fungus Batrachochytrium dendrobatidis (Bd), has been associated with the alarming decline and extinction crisis of amphibians worldwide. Because conservation programs are implemented locally, it is essential to understand how the complex interactions among host species, climate and human activities contribute to Bd occurrence at regional scales. Using weighted phylogenetic regressions and model selection, we investigated geographic patterns of Bd occurrence along a latitudinal gradient of 1500 km within a biodiversity hot spot in Chile (1845 individuals sampled from 253 sites and representing 24 species), and its association with climatic, socio‐demographic and economic variables. Analyses show that Bd prevalence decreases with latitude although it has increased by almost 10% between 2008 and 2013, possibly reflecting an ongoing spread of Bd following the introduction of Xenopus laevis. Occurrence of Bd was higher in regions with high gross domestic product (particularly near developed centers) and with a high variability in rainfall regimes, whereas models including other bioclimatic or geographic variables, including temperature, exhibited substantially lower fit and virtually no support based on Akaike weights. In addition, Bd prevalence exhibited a strong phylogenetic signal, with five species having high numbers of infected individuals and higher prevalence than the average of 13.3% across all species. Taken together, our results highlight that Bd in Chile might still be spreading south, facilitated by a subset of species that seem to play an important epidemiological role maintaining this pathogen in the communities, in combination with climatic and human factors affecting the availability and quality of amphibian breeding sites. This information may be employed to design conservation strategies and mitigate the impacts of Bd in the biodiversity hot spot of southern Chile, and similar studies may prove useful to disentangle the role of different factors contributing to the emergence and spread of this catastrophic disease.     

Mapping the Global Emergence of Batrachochytrium dendrobatidis,the Amphibian Chytrid Fungus

The rapid worldwide emergence of the amphibian pathogen Batrachochytrium dendrobatidis (Bd) is having a profound negative impact on biodiversity. However, global research efforts are fragmented and an overarching synthesis of global infection data is lacking. Here, we provide results from a community tool for the compilation of worldwide Bd presence and report on the analyses of data collated over a four-year period. Using this online database, we analysed: 1) spatial and taxonomic patterns of infection, including amphibian families that appear over- and under-infected; 2) relationships between Bd occurrence and declining amphibian species, including associations among Bd occurrence, species richness, and enigmatic population declines; and 3) patterns of environmental correlates with Bd, including climate metrics for all species combined and three families (Hylidae, Bufonidae, Ranidae) separately, at both a global scale and regional (U.S.A.) scale. These associations provide new insights for downscaled hypothesis testing. The pathogen has been detected in 52 of 82 countries in which sampling was reported, and it has been detected in 516 of 1240 (42%) amphibian species. We show that detected Bd infections are related to amphibian biodiversity and locations experiencing rapid enigmatic declines, supporting the hypothesis that greater complexity of amphibian communities increases the likelihood of emergence of infection and transmission of Bd. Using a global model including all sampled species, the odds of Bd detection decreased with increasing temperature range at a site. Further consideration of temperature range, rather than maximum or minimum temperatures, may provide new insights into Bd-host ecology. Whereas caution is necessary when interpreting such a broad global dataset, the use of our pathogen database is helping to inform studies of the epidemiology of Bd, as well as enabling regional, national, and international prioritization of conservation efforts. We provide recommendations for adaptive management to enhance the database utility and relevance.     

Species distribution models predict the geographic expansion of an enzootic amphibian pathogen

Globally, numerous amphibian species have declined due to the introduction of the chytrid fungus Batrachochytrium dendrobatidis (Bd). However, the understanding of the spatiotemporal dynamics remains incomplete. Therefore, estimating the current geographic distribution of Bd is urgently needed, especially in countries like Costa Rica, where susceptible species are still recovering from Bd‐driven declines. We conducted model tuning and spatial analysis to compare the habitat suitability for epizootic and enzootic Bd in Costa Rica and to identify data‐deficient regions, opportunistic sampling, and Bd hotspots. Our dataset combined two methods of detection (histology and PCR methods) for a total of 451 Bd‐positive records from 34 localities. We found that the distribution of enzootic Bd in Costa Rica increased 60% since previous estimates in the early 2000s and extended to highlands and dry lowlands that were considered unsuitable for Bd. We also found that Bd is common across protected lands (80%) and within the herpetological provinces containing the highest amphibian richness and endemism in Costa Rica. Opportunistic sampling of Bd has focused on sites where epizootics occurred with the strongest intensity, leading to deficient or absent sampling across the Talamanca Range, the Nicoya Peninsula, and the northern lowlands. Our results showed that PCR increased the power of Bd detection in lowlands and favored the identification of Bd hotspots across the Caribbean side of Costa Rica. Our results add to the understanding of disease spread during enzootics and can be used to identify new hotspots for disease to mitigate future outbreaks of this pathogen. Abstract in Spanish is available with online material