Lee F. Skerratt

Spread of chytridiomycosis has caused the rapid global decline and extinction of frogs

The global emergence and spread of the pathogenic, virulent, and highly transmissible fungus Batrachochytrium dendrobatidis, resulting in the disease chytridiomycosis, has caused the decline or extinction of up to about 200 species of frogs. Key postulates for this theory have been completely or partially fulfilled. In the absence of supportive evidence for alternative theories despite decades of research, it is important for the scientific community and conservation agencies to recognize and manage the threat of chytridiomycosis to remaining species of frogs, especially those that are naive to the pathogen. The impact of chytridiomycosis on frogs is the most spectacular loss of vertebrate biodiversity due to disease in recorded history.

Pathogenesis of chytridiomycosis, a cause of catastrophic amphibian declines.

Croaking Frogs The global amphibian decline has been attributed, among other causes, to an amphibian skin disease chytridiomycosis caused by the fungus Batrachochytrium dendrobatidis. However, how this pathogen causes mortality has been unclear. Voyles et al. (p. 582) show that this superficial skin infection may lead to cardiac failure owing to changes caused by lowered ion transport through the skin and consequent electrolyte reduction in the blood. A fungal disease that is associated with frog mortality causes changes in electrolyte transport across the skin. The pathogen Batrachochytrium dendrobatidis (Bd), which causes the skin disease chytridiomycosis, is one of the few highly virulent fungi in vertebrates and has been implicated in worldwide amphibian declines. However, the mechanism by which Bd causes death has not been determined. We show that Bd infection is associated with pathophysiological changes that lead to mortality in green tree frogs (Litoria caerulea). In diseased individuals, electrolyte transport across the epidermis was inhibited by >50%, plasma sodium and potassium concentrations were respectively reduced by ~20% and ~50%, and asystolic cardiac arrest resulted in death. Because the skin is critical in maintaining amphibian homeostasis, disruption to cutaneous function may be the mechanism by which Bd produces morbidity and mortality across a wide range of phylogenetically distant amphibian taxa.

Ecological dynamics of emerging bat virus spillover

Viruses that originate in bats may be the most notorious emerging zoonoses that spill over from wildlife into domestic animals and humans. Understanding how these infections filter through ecological systems to cause disease in humans is of profound importance to public health. Transmission of viruses from bats to humans requires a hierarchy of enabling conditions that connect the distribution of reservoir hosts, viral infection within these hosts, and exposure and susceptibility of recipient hosts. For many emerging bat viruses, spillover also requires viral shedding from bats, and survival of the virus in the environment. Focusing on Hendra virus, but also addressing Nipah virus, Ebola virus, Marburg virus and coronaviruses, we delineate this cross-species spillover dynamic from the within-host processes that drive virus excretion to land-use changes that increase interaction among species. We describe how land-use changes may affect co-occurrence and contact between bats and recipient hosts. Two hypotheses may explain temporal and spatial pulses of virus shedding in bat populations: episodic shedding from persistently infected bats or transient epidemics that occur as virus is transmitted among bat populations. Management of livestock also may affect the probability of exposure and disease. Interventions to decrease the probability of virus spillover can be implemented at multiple levels from targeting the reservoir host to managing recipient host exposure and susceptibility.

Environmental refuge from disease-driven amphibian extinction.

Species that are tolerant of broad environmental gradients may be less vulnerable to epizootic outbreaks of disease. Chytridriomycosis, caused by the fungus Batrachochytrium dendrobatidis, has been linked to extirpations and extinctions of amphibian species in many regions. The pathogen thrives in cool, moist environments, and high amphibian mortality rates have commonly occurred during chytridiomycosis outbreaks in amphibian populations in high-elevation tropical rainforests. In Australia several high-elevation species, including the armored mist frog (Litoria lorica), which is designated as critically endangered by the International Union for the Conservation of Nature (IUCN), were believed to have gone extinct during chytridiomycosis outbreaks in the 1980s and early 1990s. Species with greater elevational ranges disappeared from higher elevations, but remained common in the lowlands. In June 2008, we surveyed a stream in a high-elevation dry sclerophyll forest and discovered a previously unknown population of L. lorica and a population of the waterfall frog (Litoria nannotis). We conducted 6 additional surveys in June 2008, September 2008, March 2009, and August 2009. Prevalences of B. dendrobatidis infection (number infected per total sampled) were consistently high in frogs (mean 82.5%, minimum 69%) of both species and in tadpoles (100%) during both winter (starting July) and summer (starting February). However, no individuals of either species showed clinical signs of disease, and they remained abundant (3.25 - 8.75 individuals of L. lorica and 6.5-12.5 individuals of L. nannotis found/person/100 m over 13 months). The high-elevation dry sclerophyll site had little canopy cover, low annual precipitation, and a more defined dry season than a nearby rainforest site, where L. nannotis was more negatively affected by chytridiomycosis. We hypothesize this lack of canopy cover allowed the rocks on which frogs perched to warm up, thereby slowing growth and reproduction of the pathogen on the hosts. In addition, we suggest surveys for apparently extinct or rare species should not be limited to core environments.

Emerging infectious diseases of wildlife: a critical perspective

We review the literature to distinguish reports of vertebrate wildlife disease emergence with sufficient evidence, enabling a robust assessment of emergence drivers. For potentially emerging agents that cannot be confirmed, sufficient data on prior absence (or a prior difference in disease dynamics) are frequently lacking. Improved surveillance, particularly for neglected host taxa, geographical regions and infectious agents, would enable more effective management should emergence occur. Exposure to domestic sources of infection and human-assisted exposure to wild sources were identified as the two main drivers of emergence across host taxa; the domestic source was primary for fish while the wild source was primary for other taxa. There was generally insufficient evidence for major roles of other hypothesized drivers of emergence.

The mitochondrial 12S gene is a suitable marker of populations of Sarcoptes scabiei from wombats, dogs and humans in Australia

Abstract. We sequenced part of the mitochondrial 12S ribosomal RNA gene of 23 specimens of Sarcoptes scabiei from eight wombats, one dog and three humans. Twelve of the 326 nucleotide positions varied among these mites and there were nine haplotypes (sequences) that differed by 1–8 nucleotides. Phylogenetic analyses indicated that these mites were from two lineages: (1) mites from wombats from Victoria, Australia, and mites from the humans and dog from the Northern Territory, Australia (haplotypes 1–4, 9); and (2) mites from the humans and dog from the Northern Territory (haplotypes 5–8). Mites from the three different hosts (wombats, a dog and humans) had not diverged phylogenetically; rather, these mites had similar 12S sequences. Thus, we conclude that these mites from wombats, humans and a dog are closely related, and that they diverged from a common ancestor relatively recently. This conclusion is consistent with the argument that people and/or their dogs introduced to Australia the S. scabiei mites that infect wombats in Australia . So, S. scabiei, which has been blamed for the extinction of populations of wombats in Australia, may be a parasitic mite that was introduced to Australia with people and/or their dogs. These data show that the mitochondrial 12S rRNA gene may be a suitable population marker of S. scabiei from wombats, dogs and humans in Australia.

Survey protocol for detecting chytridiomycosis in all Australian frog populations

Spread of the amphibian chytrid fungus Batrachochytrium dendrobatidis (Bd) has caused the decline and extinction of frogs, but the distribution of Bd is not completely known. This information is crucial to implementing appropriate quarantine strategies, preparing for outbreaks of chytridiomycosis due to introduction of Bd, and for directing conservation actions towards affected species. This survey protocol provides a simple and standard method for sampling all frog populations in Australia to maximise the chances of detecting Bd. In order to structure and prioritise the protocol, areas are divided by bioregion and frog species are allocated depending on the water bodies they utilize into 3 groups representing different levels of risk of exposure to Bd. Sixty individuals per population need to be tested to achieve 95% certainty of detecting 1 positive frog, based on the minimum apparent prevalence of > or =5% in infected Australian frog populations and using a quantitative real-time TaqMan PCR test. The appropriate season to sample varies among bioregions and will ideally incorporate temperatures favourable for chytridiomycosis (e.g. maximum air temperatures generally <27 degrees C). Opportunistic collection and testing of sick frogs and tadpoles with abnormal mouth-parts should also be done to increase the probability of detecting Bd. The survey priorities in order are (1) threatened species that may have been exposed to Bd, (2) bioregions surrounding infected bioregions/ecological groups, and (3) species of frogs of unknown infection status in infected bioregions. Within these priority groups, sampling should first target ecological groups and species likely to be exposed to Bd, such as those associated with permanent water, and areas within bioregions that have high risk for Bd as indicated by climatic modelling. This protocol can be adapted for use in other countries and a standard protocol will enable comparison among amphibian populations globally.

BSA reduces inhibition in a TaqMan® assay for the detection of Batrachochytrium dendrobatidis

A TaqMan assay for the causative agent of chytridiomycosis in amphibians (Batrachochytrium dendrobatidis) can be inhibited by phenolic compounds, including humic and tannic acids, resulting in false negatives. Bovine serum albumin (BSA) is known to reduce inhibition of PCR when samples are contaminated with these inhibitors. We assessed the effect of BSA in reducing inhibition of the TaqMan assay when analyzing skin swabs for B. dendrobatidis. We found that the addition of BSA to the TaqMan reaction reduced inhibition to insignificant levels. BSA did not appreciably affect the efficiency or analytical sensitivity of the TaqMan reaction in the analysis of standard DNA solutions free from environmental inhibitors. We recommend the addition of 400 ng microl(-1) of BSA to the standard TaqMan assay to reduce inhibition associated with sampling wild amphibians.

Susceptibility of amphibians to chytridiomycosis is associated with MHC class II conformation

The pathogenic chytrid fungus Batrachochytrium dendrobatidis (Bd) can cause precipitous population declines in its amphibian hosts. Responses of individuals to infection vary greatly with the capacity of their immune system to respond to the pathogen. We used a combination of comparative and experimental approaches to identify major histocompatibility complex class II (MHC-II) alleles encoding molecules that foster the survival of Bd-infected amphibians. We found that Bd-resistant amphibians across four continents share common amino acids in three binding pockets of the MHC-II antigen-binding groove. Moreover, strong signals of selection acting on these specific sites were evident among all species co-existing with the pathogen. In the laboratory, we experimentally inoculated Australian tree frogs with Bd to test how each binding pocket conformation influences disease resistance. Only the conformation of MHC-II pocket 9 of surviving subjects matched those of Bd-resistant species. This MHC-II conformation thus may determine amphibian resistance to Bd, although other MHC-II binding pockets also may contribute to resistance. Rescuing amphibian biodiversity will depend on our understanding of amphibian immune defence mechanisms against Bd. The identification of adaptive genetic markers for Bd resistance represents an important step forward towards that goal.

Chytridiomycosis and Seasonal Mortality of Tropical Stream‐Associated Frogs 15 Years after Introduction of Batrachochytrium dendrobatidis

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