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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.

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.

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.

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.

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.

Tracking and understanding variation in pathogens such as Batrachochytrium dendrobatidis (Bd), the agent of amphibian chytridiomycosis which has caused population declines globally, is a priority for many land managers. However, relatively little sampling of amphibian communities has occurred at high latitudes. We used skin swabs collected during 2005-2017 from boreal toads Anaxyrus boreas (n = 248), in southeast Alaska (USA; primarily in and near Klondike Gold Rush National Historical Park [KLGO]) and northwest British Columbia (Canada) to determine how Bd prevalence varied across life stages, habitat characteristics, local species richness, and time. Across all years, Bd prevalence peaked in June and was >3 times greater for adult toads (37.5%) vs. juveniles and metamorphs (11.2%). Bd prevalence for toads in the KLGO area, where other amphibian species are rare or absent, was highest from river habitats (55.0%), followed by human-modified upland wetlands (32.3%) and natural upland wetlands (12.7%)-the same rank-order these habitats are used for toad breeding. None of the 12 Columbia spotted frogs Rana luteiventris or 2 wood frogs R. sylvatica from the study area tested Bd-positive, although all were from an area of low host density where Bd has not been detected. Prevalence of Bd on toads in the KLGO area decreased during 2005-2015. This trend from a largely single-species system may be encouraging or concerning, depending on how Bd is affecting vital rates, and emphasizes the need to understand effects of pathogens before translating disease prevalence into management actions.

Infection caused by the fungus Batrachochytrium dendrobatidis (Bd) produces chytridiomycosis, a disease considered one of the main causes of amphibian population declines in the world. In Brazil, Bd has been recorded in several regions, but mainly in the Atlantic Forest biome. This study aimed to investigate the occurrence of Bd in amphibian species in Bahia State to test the hypothesis that Bd is widespread in other Brazilian biomes. Using histological analysis, we evaluated the skin of 190 anurans of 85 species preserved in herpetological collections. Based on these analyses, the distribution of Bd was extended approximately 400 km to the west, 150 km to the north and 105 km to the east in the state of Bahia. Of the 190 specimens analyzed, Bd infection was diagnosed in 16 individuals, from 14 species, with the earliest record from a specimen collected in 1996 in the Caatinga biome. We identified Bd in 13 adult specimens, including 2 individuals showing suggestive signs of the disease (loss of skin pigmentation). In tadpoles, we recorded fungal structures in the oral region and on the epidermis adjacent to the rows of teeth. The results of this study corroborate the prediction that Bd is widespread in the Atlantic Forest biome, and suggest that it is widespread in the other biomes of the state (Cerrado and Caatinga, at least since 1996). Conservation efforts should involve long-term studies aimed at providing information on the dynamics of the infection, its relationship with its host and its effect on amphibian populations.

The pathogenic chytrid fungi Batrachochytrium dendrobatidis (Bd) and B. salamandrivorans (Bsal) cause infections that have become primary drivers of amphibian biodiversity loss. While globally widespread, the distribution margins of Bd and Bsal have not been determined, and the presence of these pathogens has probably gone unnoticed in many areas, especially in northern Eurasia. To better understand the presence and distribution of both pathogens in the northern temperate and boreal forest biomes, 243 individuals were sampled from 8 native amphibian species across Estonia. Additionally, 68 amphibians were sampled from captive collections in Estonia and Latvia. Pathogen infection was assessed using metabarcoding of the ITS2 marker. No positive matches for Bsal infection were found. Bd was detected in 13 specimens, 3 of which were sampled at the Riga Zoo (with a prevalence of 5.2%) and 10 in natural environments in Estonia (3.3%). The infected wild individuals belonged to 6 amphibian species and were detected throughout the mainland of Estonia, but not on islands. Prevalence of infection with Bd ranged between 3.1 and 12.5% among native species. In addition, we found molecular evidence for a potentially new sister species to Bd in nature. Although outbreaks of chytridiomycosis have never been observed in Estonia, it cannot be excluded that the dynamics of local amphibian populations are affected by Bd infections. Therefore, further work, including capture-mark-recapture studies and long-term monitoring, are required to clarify the impact of Bd on amphibians in Northern Europe.

Complex interactions among hosts, pathogens, and the environment affect the vulnerability of amphibians to the emergence of infectious diseases such as chytridiomycosis, caused by Batrachochytrium dendrobatidis (Bd). Boana curupi is a forest-dwelling amphibian endemic to the southern Atlantic Forest of South America, a severely fragmented region. Here, we evaluated whether abiotic factors (including air and water temperature, relative air humidity, and landscape) are correlated with chytrid infection intensity and prevalence in B. curupi. We found individuals infected with Bd in all populations sampled. Prevalence ranged from 25-86%, and the infection burden ranged from 1 to over 130000 zoospore genomic equivalents (g.e.) (mean ± SD: 4913 ± 18081 g.e.). The infection load differed among populations and was influenced by forest cover at scales of 100, 500, and 1000 m, with the highest infection rates recorded in areas with a higher proportion of forest cover. Our results suggest that the fungus is widely distributed in the populations of B. curupi in southern Brazil. Population and disease monitoring are necessary to better understand the relationships between host, pathogen, and environment, especially when, as in the case of B. curupi, threatened species are involved.

Amphibians are globally threatened by emerging infectious diseases, and ranaviruses are among the most concerning pathogens to threaten species in the wild. We sampled for ranaviruses in wild amphibians at 8 sites in Costa Rica, spanning broad climatic zones and taxonomic associations. Seven of these sites are inhabited by highly threatened amphibian species that persist at low global population sizes after population declines due to amphibian chytridiomycosis. One of the surveyed sites is occupied by an introduced amphibian species, which is relatively rare in Central America but may be an important pathway for long-distance transport of ranaviruses. We detected ranavirus using quantitative polymerase chain reaction in 16.3% of the 243 individuals and among 5 of our 8 sites, but not at the site with the introduced species. Infection prevalence varied among species and sites, but not with mean annual temperature or mean annual precipitation. Infection intensity did not vary with species, site, temperature, or precipitation. Our results show that ranavirus infection is spatially widespread in Costa Rica, affecting a broad range of host species, and occurs across climatic zones—though we encountered no mortality or morbidity in our sampled species. Ranaviruses are known to cause intermittent mass mortality in amphibian populations, and the threatened species sampled here are likely vulnerable to population impacts from emerging ranaviruses. Therefore, we believe the potential impacts of ranaviruses on amphibian populations in tropical regions have likely been underestimated, and that they should be viewed as a potential major stressor to threatened amphibians in tropical regions.

Human-induced changes of the environment, including landscape alteration and habitat loss, may affect wildlife disease dynamics and have important ramifications for wildlife conservation. Amphibians are among the vertebrate taxa most threatened by anthropogenic habitat change. The emerging fungal pathogen Batrachochytrium dendrobatidis (Bd) has caused extinctions and population declines in hundreds of anuran species globally. We studied how the urban landscape is associated with the prevalence of Bd infections by sampling 655 anurans of 3 species (mainly the common toad Bufo bufo) in 42 ponds surrounded by different amounts of urban habitat (defined as towns, cities or villages). We also examined the association between Bd infections and a potential reservoir host species (the moor frog Rana arvalis). We found that 38% of the sites were positive for Bd with an infection prevalence of 4.4%. The extent of urban landscape was negatively correlated with Bd infection prevalence. However, the positive association of Bd with the presence of the possible reservoir species was substantially stronger than the urban effects. The body condition index of B. bufo was negatively associated with Bd infection. This Bd effect was stronger than the negative effect of urban landscape on body condition. Our results suggest that urban environments in Sweden have a negative impact on Bd infections, while the presence of the reservoir species has a positive impact on Bd prevalence. Our study also highlights the potential importance of Bd infection on host fitness, especially in rural landscapes.

Amphibian diversity has declined due to the infectious disease chytridiomycosis, caused by the fungal pathogen Batrachochytrium dendrobatidis (Bd). Coexistence between amphibian hosts and this pathogen in some locations is attributed to the presence of the cutaneous bacterium Janthinobacterium lividum (Jliv). This microbe inhibits the growth of Bd on the host, reduces morbidity, and improves survival. Andean water frogs in the genus Telmatobius seem to be particularly vulnerable to the disease yet populations of T. intermedius and T. marmoratus persist in southern and central Peru. We investigated the presence of Jliv on these 2 frog species and assessed the relationship of Jliv presence with prevalence and intensity of Bd infection. By sampling 125 frogs from 7 streams (3323-3950 m a.s.l.) and 27 from a city market, we found spatial variation in the mutualism among populations (range 0-40% proportion of Jliv-positives). Overall, 57% of frogs were infected with Bd, 12.5% of frogs hosted both Jliv and Bd, while 7.2% hosted just Jliv. We found that the probability of an individual being infected with Bd was independent of the presence of Jliv; however, we did detect a protective effect of Jliv with respect to intensity of infection. The extent of Jliv distribution in the high Andes stands in stark contrast to the rarity of Jliv on frogs in lower elevation cloud forest biomes.

In Australia, the cane toad Rhinella marina and chytrid fungus Batrachochytrium dendrobatidis (Bd) are examples of invasive species that have had dramatic impacts on native fauna. However, little is known about the interaction between Bd and cane toads. We aimed to explore the interaction of these 2 species in 3 parts. First, we collated data from the literature on Bd infection in wild cane toads. Second, we tested the susceptibility of recently metamorphosed cane toads to Bd infection. Finally, we modelled the distribution of the 2 species in Australia to identify where they overlap and, therefore, might interact. Through our data collation, we found that adult cane toads are infrequently infected and do not carry high infection burdens; however, our infection experiment showed that metamorphs are highly susceptible to infection and disease, but resistance appears to increase with increasing toad size. Niche modelling revealed overlapping distributions and the potential for cane toads to be affected by chytridiomycosis in the wild. While Bd can cause mortality in small juveniles in the laboratory, warm microhabitats used by wild toads likely prevent infection, and furthermore, high mortality of juveniles is unlikely to affect the adult populations because they are highly fecund. However, to demonstrate the impact of Bd on wild cane toad populations, targeted field studies are required to assess (1) the overall impact of chytridiomycosis on recruitment especially in cooler areas more favourable for Bd and (2) whether cane toad juveniles can amplify Bd exposure of native amphibian species in these areas.

The emerging infectious disease chytridiomycosis, caused by the fungus Batrachochytrium dendrobatidis, is a major driver pushing many amphibian species to the brink of extinction. Substantial efforts to develop effective protocols that use antifungal drugs have had notable success. Here, we used the antifungal agents itraconazole and thiophanate-methyl, singly and in combination, in an attempt to treat common midwife toad Alytes obstetricans larvae naturally infected with the globalized hypervirulent lineage of B. dendrobatidis. Despite the successful use of itraconazole in a closely related species (A. muletensis), our results show that these antifungal treatments are not always effective and that full clearance of animals cannot be assumed following treatment., This research was conducted under permits of the Consejería de Medio Ambiente of Aragón, Castilla y León and Madrid, and supported by the Fundación BBVA. T.W.J.G. was supported by NERC Standard Grant NE/N009967/1.

Disease monitoring is an essential step in translocation projects, specifically in amphibians where emerging pathogens such as the chytrid fungus Batrachochytrium dendrobatidis (Bd) are linked to population declines. The eastern hellbender Cryptobranchus alleganiensis is a large, fully aquatic salamander experiencing precipitous range-wide population declines; however, the role Bd plays in these declines is unclear. To augment declining hellbender populations and determine effects of translocation on Bd prevalence, we conducted a translocation study of wild adult hellbenders from 2 source streams with abundant hellbender populations to 2 streams with declining populations in east Tennessee, USA. In 2018, we implanted radio transmitters into 30 hellbenders and sampled them periodically for Bd until 17 of the 30 hellbenders were translocated in 2019. We attempted to recapture translocated hellbenders approximately every 45 d for 3 mo to determine Bd prevalence post-release. We used qPCR to detect Bd and quantify zoospore loads on positive samples. Hellbenders had a pre-translocation Bd prevalence of 50% (15 of 30), which decreased to 10% (1 of 10) post-translocation. The average zoospore load for positive samples was 73.63 ± 30.82, and no hellbenders showed signs of chytridiomycosis throughout the study. Although we detected no significant effect of translocation on Bd prevalence, we observed a reduction in Bd prevalence post-release. Our results indicate that translocation did not lead to an increase in pathogen prevalence in translocated wild adult hellbenders, suggesting that chytrid did not impact the success of short-term translocations of eastern hellbenders in the Blue Ridge ecoregion.

Amphibians have been facing a pandemic caused by the deadly fungus Batrachochytrium dendrobatidis (Bd). Although studies have elucidated cutaneous and homeostatic disturbances, it is still unknown if the hepatic function can be affected or if hepatic effects differ among host species. Thus, we evaluated the effects of an experimental Bd infection on the liver (histopathology and the hepatosomatic index) of 2 anuran species (Xenopus laevis and Physalaemus albonotatus) with different susceptibilities to Bd infection and compared them to uninfected controls. Bd infection increased the melanomacrophage cell area and induced leukocyte infiltration in both species. The effects were more pronounced in the sensitive species, P. albonotatus, which showed severe reduction in glycogen stores and liver atrophy, due to energetic imbalance. Hepatocytes of P. albonotatus also showed ballooning degeneration (vacuolization), which could lead to cell death and liver failure. Our results provide evidence that although the sensitive species showed more severe effects, the tolerant species also had hepatic responses to the infection. These findings indicate that hepatic function can play an important role in detoxification and in immune responses to chytridiomycosis, and that it may be used as a new biomarker of health status in chytrid infections.

Batrachochytrium dendrobatidis (Bd) has been detected in wild hellbender Cryptobranchus alleganiensis populations, with rare instances of chytridiomycosis and Bd-induced mortality. To date, Bd surveillance in hellbender populations has been disproportionately focused on adult age classes. A lingering question is whether Bd might contribute to hellbender declines through disproportionate negative effects on immature age classes. The objective of this study was to quantify Bd prevalence and load in immature hellbenders in western North Carolina, USA. We conducted field surveys during 2018 and 2019 and collected 88 skin swabs from 84 hellbenders spanning 3 age classes. Bd was detected on 11% of individuals, including 8 larvae and 1 juvenile. We did not detect symptoms of chytridiomycosis or a decline in body condition in Bd-positive hellbenders. Load varied from approximately 1-153 zoospore equivalents for the 9 Bd-positive hellbenders and was not associated with size class of the individual. While hellbenders appeared to be abundant in each survey reach, more work is needed to determine whether Bd may increase the vulnerability of immature hellbenders to anthropogenic stressors.

Discovered in 2013, Batrachochytrium salamandrivorans (Bsal) is an emerging amphibian pathogen that causes ulcerative skin lesions and multifocal erosion. A closely related pathogen, Batrachochytrium dendrobatidis (Bd), has devastated amphibian populations worldwide, suggesting that Bsal poses a significant threat to global salamander biodiversity. To expedite research into this emerging threat, we seek to standardize protocols across the field so that results of laboratory studies are reproducible and comparable. We have collated data and experience from multiple labs to standardize culturing practices of Bsal. Here we outline common culture practices including a media for optimal Bsal growth, standard culture protocols, and a method for isolating Bsal from infected tissue.

Chytridiomycosis and ranavirosis are 2 emerging infectious diseases that have caused significant global amphibian decline. Although both have received much scrutiny, little is known about interactions between the 2 causative agents Batrachochytrium dendrobatidis (Bd) and ranavirus (Rv) at the individual host and population levels. We present the first longitudinal assessment of Bd, Rv, and co-infections of a temperate amphibian assemblage in North America. From 2012 to 2016, we assessed the temporal oscillations of Bd, Rv and co-infection dynamics in a sample of 729 animals representing 13 species. Bd, Rv, and co-infected amphibians were detected during all 5 yr. Bd, Rv, and co-infection prevalence all varied annually, with the lowest instances of each at 2.1% (2013), 7.9% (2016), and 0.6% (2016), respectively. The highest Bd, Rv, and co-infection prevalence were recorded in 2012 (26.8%), 2016 (38.3%), and 2015 (10.3%), respectively. There was no association between Bd or Rv infection prevalence and co-infection, either when assessing the entire amphibian assemblage as a whole (odds ratio 1.32, 95% CI: 0.83-2.1, p = 0.29) or within species for amphibians that were more numerically represented (n > 40, p > 0.05). This suggests neither Bd nor Rv facilitate host co-infections within the sampled host assemblage. Instead, the basis for co-infections is the spatiotemporal distribution of both pathogens. Despite lack of interplay between Bd and Rv in this population, our study highlights the importance of considering numerous pathogens that may be present within amphibian habitats in order to properly anticipate interactions that may have direct bearing on disease outcomes.

Author(s): Peralta-Garcia, A; Adams, AJ; Briggs, CJ; Galina-Tessaro, P; Valdez-Villavicencio, JH; Hollingsworth, BD; Shaffer, HB; Fisher, RN | Abstract: Chytridiomycosis is caused by the fungal pathogen Batrachochytrium dendrobatidis (Bd) and is regarded as one of the most significant threats to global amphibian populations. In Mexico, Bd was first reported in 2003 and has now been documented in 13 states. We visited 33 localities and swabbed 199 wild-caught anurans from 7 species (5 native, 2 exotic) across the Mediterranean region of the state of Baja California. Using quantitative PCR, Bd was detected in 94 individuals (47.2% of samples) at 25 of the 33 survey localities for 5 native and 1 exotic frog species. The exotic Xenopus laevis was the only species that tested completely negative for Bd. We found that remoteness, distance to agricultural land, and elevation were the best positive predictors of Bd presence. These are the first Bd-positive results for the state of Baja California, and its presence should be regarded as an additional conservation threat to the region’s native frog species.

Amphibians suffer from a number of factors that make them the most threatened group of vertebrates. One threat is the fungal disease chytridiomycosis caused by the emerging pathogen Batrachochytrium dendrobatidis (Bd), which has rapidly spread and caused the loss of massive amphibian biodiversity worldwide. Recently, Bd was associated with a few amphibian population declines and extinctions in some areas of the Brazilian Atlantic Forest. However, the mechanisms underlying such declines are not fully understood. Therefore, it is essential to improve our knowledge of abiotic factors that can possibly influence Bd prevalence and chytridiomycosis disease severity. Herein we tested the hypothesis that water availability (such as in perennial streams, where Bd is frequently present in larvae) and rainfall would increase the prevalence of Bd. To test this, we sampled frogs from 6 transects with different numbers of perennial waterbodies, and we report that the more water available in the area, the higher the probability of Bd infection on anurans. Seasonality also influenced both the Bd prevalence in the area and the intensity of infection in infected frogs. However, Bd prevalence was higher during the rainy months whereas the infection burden was lower. We suggest that Bd is likely spread during the summer, when most anuran species gather near the water for spawning and when rainfall overfills ephemeral wetlands. On the other hand, during the drier months, a higher infection burden may be explained by increased disease susceptibility.