Pathogenesis of Bohle Iridovirus (Genus Ranavirus) in Experimentally Infected Juvenile Eastern Water Dragons ( Intellagama lesueurii lesueurii).

Juvenile eastern water dragons ( Intellagama lesueurii lesueurii) are highly susceptible to infection with Bohle iridovirus (BIV), a species of ranavirus first isolated from ornate burrowing frogs in Townsville, Australia. To investigate the progression of BIV infection in eastern water dragons, 11 captive-bred juveniles were orally inoculated with a dose of 104.33 TCID50 and euthanized at 3, 6, 8, 10, 12, and 14 days postinfection (dpi). Viral DNA was detected via polymerase chain reaction (PCR) in the liver, kidney, and cloacal swabs at 3 dpi. Mild lymphocytic infiltration was observed in the submucosa and mucosa of the tongue and liver at 3 dpi. Immunohistochemistry (IHC) first identified viral antigen in foci of splenic necrosis and in hepatocytes with intracytoplasmic inclusion or rare single-cell necrosis at 6 dpi. By 14 dpi, positive IHC labeling was found in association with lesions in multiple tissues. Selected tissues from an individual euthanized at 14 dpi were probed using in situ hybridization (ISH). The ISH labeling matched the location and pattern detected by IHC. The progression of BIV infection in eastern water dragons, based on lesion severity and virus detection, appears to start in the spleen, followed by the liver, then other organs such as the kidney, pancreas, oral mucosa, and skin. The early detection of ranaviral DNA in cloacal swabs and liver and kidney tissue samples suggests these to be a reliable source of diagnostic samples in the early stage of disease before the appearance of clinical signs, as well as throughout the infection.

Susceptibility of eastern water dragons Intellagama lesueurii lesueurii to Bohle iridovirus.

Ranaviruses infect and have been associated with mass mortality events in fish, amphibians and reptiles and are capable of interclass transmission. Eastern water dragons (EWDs), a semi-aquatic squamate, have an overlapping distribution with several species shown to be susceptible to Bohle iridovirus (BIV). However, this species has not been previously investigated, and no known mass mortalities have occurred in wild populations. Here we report the experimental infection of juvenile EWDs with BIV to investigate a water-dwelling lizards' susceptibility to a ranaviral strain present in northern Queensland, Australia. Lizards were exposed via oral inoculation, intramuscular injection, or cohabitation with orally infected lizards. All exposure methods were effective in establishing an infection as demonstrated by skin lesions and pathological changes in the internal organs. Necrosis, haemorrhage and inflammation were observed histologically in the pancreas, liver, spleen, kidney and submucosa of the gastrointestinal tract of BIV-exposed lizards. Variably sized basophilic intracytoplasmic inclusion bodies were observed in the liver of 6/14 BIV-exposed lizards. Virus was isolated from the liver and kidney of all BIV-infected lizards and confirmed with quantitative PCR (qPCR). The outcome of this study demonstrates that juvenile EWDs are susceptible to BIV, thereby adding Australian lizards to the broad host range of ranaviruses. Furthermore, this study provides additional evidence of BIV's ability to infect different classes of ecothermic vertebrates.

Serological survey of Australian native reptiles for exposure to ranavirus.

Ranaviruses have been isolated from many ectothermic vertebrates, and serological surveys of both amphibians and reptiles have shown the presence of ranaviral antibodies in a proportion of these populations. An enzyme-linked immunosorbent assay (ELISA) was developed to measure serum antibodies against ranavirus in Australian reptiles. The ELISA was validated with serum from challenge trials with Bohle iridovirus (BIV) in 6 reptilian species. A preliminary sero-survey of northern Queensland riparian reptile fauna (saw-shelled turtles Myuchelys latisternum, Krefft's river turtles Emydura macquarii krefftii, freshwater crocodiles Crocodylus johnstoni, as well as the snakes Boiga irregularis, Dendrelaphis punctulatus, Tropidonophis mairii, Morelia spilota, Liasis childreni and L. fuscus) revealed evidence of past exposure to Bohle iridoviral antigens in part of the population at several locations sampled. Furthermore, in Krefft's river turtles and freshwater crocodiles, a statistically significant trend was apparent for larger reptiles to be more likely to have BIV-reactive sera than smaller individuals. The use of adult tortoise populations as sentinels can assist in monitoring the presence of BIV in northern Australian freshwater streams, and thereby the potential dangers to native fauna from this agent.