Alex D. Hyatt

Emerging infectious diseases and amphibian population declines.

We review recent research on the pathology, ecology, and biogeography of two emerging infectious wildlife diseases, chytridiomycosis and ranaviral disease, in the context of host-parasite population biology. We examine the role of these diseases in the global decline of amphibian populations and propose hypotheses for the origins and impact of these panzootics. Finally, we discuss emerging infectious diseases as a global threat to wildlife populations.

Anthropogenic environmental change and the emergence of infectious diseases in wildlife

By using the criteria that define emerging infectious diseases (EIDs) of humans, we can identify a similar group of EIDs in wildlife. In the current review we highlight an important series of wildlife EIDs: amphibian chytridiomycosis; diseases of marine invertebrates and vertebrates and two recently-emerged viral zoonoses, Nipah virus disease and West Nile virus disease. These exemplify the varied etiology, pathogenesis, zoonotic potential and ecological impact of wildlife EIDs. Strikingly similar underlying factors drive disease emergence in both human and wildlife populations. These are predominantly ecological and almost entirely the product of human environmental change. The implications of wildlife EIDs are twofold: emerging wildlife diseases cause direct and indirect loss of biodiversity and add to the threat of zoonotic disease emergence. Since human environmental changes are largely responsible for their emergence, the threats wildlife EIDs pose to biodiversity and human health represent yet another consequence of anthropogenic influence on ecosystems. We identify key areas where existing expertise in ecology, conservation biology, wildlife biology, veterinary medicine and the impact of environmental change would augment programs to investigate emerging diseases of humans, and we comment on the need for greater medical and microbiological input into the study of wildlife diseases.

Origin of the Amphibian Chytrid Fungus

Histologic evidence indicates southern Africa as the origin of the amphibian chytrid fungus.

Comparative studies of piscine and amphibian iridoviruses

Summary. A total of 30 iridoviruses collected from Australia, South-East Asia, North America, South America and Europe were characterised. With the exception of the South-East Asian iridoviruses all viruses were found to belong to the genus Ranavirus. All viruses, except those originating from South-East Asia, cross-reacted with antisera against epizootic haematopoietic necrosis virus (EHNV). Viruses or virus-infected cells were examined using electron microscopy, SDS PAGE, restriction endonuclease (RE) digestion, DNA hybridisation, and DNA sequencing. Data from RE digestion of genomic DNA, and from the sequencing of PCR products indicated that the viruses generally grouped according to their geographic and taxonomic (i.e. amphibian or fish) origin. The one exception to this was the viruses from the United Kingdom that grouped with the North American ranaviruses. The differences between specified genomic regions were small. To assess the validity of the differences in sequence homology, similar studies were performed with different isolates from two viruses (EHNV and Guatopo virus (GV), collected from different animals at different locations and time). The sequence data showed complete homology for the isolates for any one virus over the 200 and 586 bp regions examined. Collectively, the data showed that the coding region for the major coat protein (MCP) is stable for any one species (e.g. EHNV).

CHARACTERISATION OF A NOVEL LYSSAVIRUS ISOLATED FROM PTEROPID BATS IN AUSTRALIA

A novel lyssavirus isolated from Pteropid bats in Australia (Australian Bat Lyssavirus, ABLV) has been characterised using gene sequence analyses, electron microscopy and a panel of monoclonal antibodies. Electron microscopic examination of Pteropid bat and mouse brain material as well as virus isolated from tissue culture medium, showed the presence of bullet-shaped rhabdovirus particles and structures characteristic of lyssavirus. Analysis using nucleocapsid (N) specific monoclonal antibodies, showed a strong relationship between this new lyssavirus and serotype 1 rabies. The nucleotide sequence of the prototype strain of ABLV was determined from the initiator methionine codon for the nucleocapsid protein (N protein) to the amino terminus of the polymerase gene (L protein), a distance of 5344 nucleotides. Comparisons of the deduced N, phosphoprotein (P), matrix protein (M), and glycoprotein (G) proteins showed that ABLV was more closely related to serotype 1 classic rabies viruses than to other members of the Lyssavirus genus. The percent relatedness of the ABLV proteins when compared to the cognate proteins of PV (Pasteur vaccine strain) rabies was 92, 75, 87 and 75% for the N, P, M and G proteins, respectively. Phylogenetic studies of N protein sequences showed clearly that ABLV is an unrecognised member of the Lyssavirus genus and represents a new genotype, genotype 7.

Pteropid bats are confirmed as the reservoir hosts of henipaviruses: a comprehensive experimental study of virus transmission.

Bats of the genus Pteropus have been identified as the reservoir hosts for the henipaviruses Hendra virus (HeV) and Nipah virus (NiV). The aim of these studies was to assess likely mechanisms for henipaviruses transmission from bats. In a series of experiments, Pteropus bats from Malaysia and Australia were inoculated with NiV and HeV, respectively, by natural routes of infection. Despite an intensive sampling strategy, no NiV was recovered from the Malaysian bats and HeV was reisolated from only one Australian bat; no disease was seen. These experiments suggest that opportunities for henipavirus transmission may be limited; therefore, the probability of a spillover event is low. For spillover to occur, a range of conditions and events must coincide. An alternate assessment framework is required if we are to fully understand how this reservoir host maintains and transmits not only these but all viruses with which it has been associated.

Amphibian chytridiomycosis in Japan: distribution, haplotypes and possible route of entry into Japan.

A serious disease of amphibians caused by the chytrid fungus Batrachochytrium dendrobatidis was first found in Japan in December 2006 in imported pet frogs. This was the first report of chytridiomycosis in Asia. To assess the risk of pandemic chytridiomycosis to Japanese frogs, we surveyed the distribution of the fungus among captive and wild frog populations. We established a nested PCR assay that uses two pairs of PCR primers to amplify the internal transcribed spacer (ITS) region of a ribosomal RNA cassette to detect mild fungal infections from as little as 0.001 pg (1 fg) of B. dendrobatidis DNA. We collected swab samples from 265 amphibians sold at pet shops, 294 bred at institutes and 2103 collected at field sites from northern to southwestern Japan. We detected infections in native and exotic species, both in captivity and in the field. Sequencing of PCR products revealed 26 haplotypes of the B. dendrobatidis ITS region. Phylogenetic analysis showed that three of these haplotypes were specific to the Japanese giant salamander (Andrias japonicus) and appeared to have established a commensal relationship with this native amphibian. Many other haplotypes were carried by alien amphibians. The highest genetic diversity of B. dendrobatidis was found in the American bullfrog (Rana catesbeiana). Some strains of B. dendrobatidis appeared to be endemic to Japanese native amphibians, but many alien strains are being introduced into Japan via imported amphibians. To improve chytridiomycosis risk management, we must consider the risk of B. dendrobatidis changing hosts as a result of anthropogenic disturbance of the host‐specific distribution of the fungus.

A previously unknown reovirus of bat origin is associated with an acute respiratory disease in humans

Respiratory infections constitute the most widespread human infectious disease, and a substantial proportion of them are caused by unknown etiological agents. Reoviruses (respiratory enteric orphan viruses) were first isolated from humans in the early 1950s and so named because they were not associated with any known disease. Here, we report a previously unknown reovirus (named “Melaka virus”) isolated from a 39-year-old male patient in Melaka, Malaysia, who was suffering from high fever and acute respiratory disease at the time of virus isolation. Two of his family members developed similar symptoms ≈1 week later and had serological evidence of infection with the same virus. Epidemiological tracing revealed that the family was exposed to a bat in the house ≈1 week before the onset of the fathers clinical symptoms. Genome sequence analysis indicated a close genetic relationship between Melaka virus and Pulau virus, a reovirus isolated in 1999 from fruit bats in Tioman Island, Malaysia. Screening of sera collected from human volunteers on the island revealed that 14 of 109 (13%) were positive for both Pulau and Melaka viruses. This is the first report of an orthoreovirus in association with acute human respiratory diseases. Melaka virus is serologically not related to the different types of mammalian reoviruses that were known to infect humans asymptomatically. These data indicate that bat-borne reoviruses can be transmitted to and cause clinical diseases in humans.

Agricultural intensification, priming for persistence and the emergence of Nipah virus: a lethal bat-borne zoonosis

Emerging zoonoses threaten global health, yet the processes by which they emerge are complex and poorly understood. Nipah virus (NiV) is an important threat owing to its broad host and geographical range, high case fatality, potential for human-to-human transmission and lack of effective prevention or therapies. Here, we investigate the origin of the first identified outbreak of NiV encephalitis in Malaysia and Singapore. We analyse data on livestock production from the index site (a commercial pig farm in Malaysia) prior to and during the outbreak, on Malaysian agricultural production, and from surveys of NiVs wildlife reservoir (flying foxes). Our analyses suggest that repeated introduction of NiV from wildlife changed infection dynamics in pigs. Initial viral introduction produced an explosive epizootic that drove itself to extinction but primed the population for enzootic persistence upon reintroduction of the virus. The resultant within-farm persistence permitted regional spread and increased the number of human infections. This study refutes an earlier hypothesis that anomalous El Niño Southern Oscillation-related climatic conditions drove emergence and suggests that priming for persistence drove the emergence of a novel zoonotic pathogen. Thus, we provide empirical evidence for a causative mechanism previously proposed as a precursor to widespread infection with H5N1 avian influenza and other emerging pathogens.

Mass mortality associated with a viral-induced vacuolating encephalopathy and retinopathy of larval and juvenile barramundi, Lates calcarifer Bloch

Abstract Light-microscopic examinations revealed a consistent correlation between a syndrome of mass mertality in hatchery-raised larval and juvenile barramundi and vacuolation in the brains and retinas of these fish. Large amounts of picorna-like viral particles were found associated with these lesions when brain and retinal tissue was examined by electron microscopy. Epizootological data suggested that vertical transmission of the infectious agent is not a common occurrence, and most transfer of infection is probably from previously-hatched juveniles to recentlyhatched larvae. These data include the observation that adequate isolation of older juvenile fish from recently-hatched larvae, coupled with between-hatch cleaning and disinfection of the premises, has apparently controlled mass mortalities in one large hatchery.