Abstract
West Nile virus, Kunjin strain (WNVKUN) is endemic in Northern Australia, but rarely causes clinical disease in humans and horses. Recently, WNVKUN genomic material was detected in cutaneous lesions of farmed saltwater crocodiles (Crocodylus porosus), but live virus could not be isolated, begging the question of the pathogenesis of these lesions. Crocodile hatchlings were experimentally infected with either 105 (n = 10) or 104 (n = 11) TCID50-doses of WNVKUN and each group co-housed with six uninfected hatchlings in a mosquito-free facility. Seven hatchlings were mock-infected and housed separately. Each crocodile was rotationally examined and blood-sampled every third day over a 3-week period. Eleven animals, including three crocodiles developing typical skin lesions, were culled and sampled 21 days post-infection (dpi). The remaining hatchlings were blood-sampled fortnightly until experimental endpoint 87 dpi. All hatchlings remained free of overt clinical disease, apart from skin lesions, throughout the experiment. Viremia was detected by qRT-PCR in infected animals during 2–17 dpi and in-contact animals 11–21 dpi, indicating horizontal mosquito-independent transmission. Detection of viral genome in tank-water as well as oral and cloacal swabs, collected on multiple days, suggests that shedding into pen-water and subsequent mucosal infection is the most likely route. All inoculated animals and some in-contact animals developed virus-neutralizing antibodies detectable from 17 dpi. Virus-neutralizing antibody titers continued to increase in exposed animals until the experimental endpoint, suggestive of persisting viral antigen. However, no viral antigen was detected by immunohistochemistry in any tissue sample, including from skin and intestine. While this study confirmed that infection of saltwater crocodiles with WNVKUN was associated with the formation of skin lesions, we were unable to elucidate the pathogenesis of these lesions or the nidus of viral persistence. Our results nevertheless suggest that prevention of WNVKUN infection and induction of skin lesions in farmed crocodiles may require management of both mosquito-borne and water-borne viral transmission in addition to vaccination strategies.
Highlights
West Nile virus (WNV) is a mosquito-transmitted flavivirus that produces a potentially fatal disease in humans, horses, birds and alligators and has been associated with outbreaks of viral encephalitis in Africa, Europe, and the Americas [1]
Our results suggest that despite absence of clinical signs or pathological lesions in the gastrointestinal tract, WNVKUN is shed into the water and can be transmitted directly to other animals in close contact
We have presented data that confirm that WNV does cause characteristic skin lesions, so-called “pix”, in a subset of infected saltwater crocodiles within a few weeks of infection and that so-called “pix”, in a subset of infected saltwater crocodiles within a few weeks of infection and that at least some of the infected animals sustain a viremia at a level sufficient for transmission to biting at least some of the infected animals sustain a viremia at a level sufficient for transmission to biting mosquitoes [32]
Summary
West Nile virus (WNV) is a mosquito-transmitted flavivirus that produces a potentially fatal disease in humans, horses, birds and alligators and has been associated with outbreaks of viral encephalitis in Africa, Europe, and the Americas [1]. Until 2011, the relatively benign WNVKUN had only been associated with a few cases of non-fatal encephalitis in humans and a handful of equine cases, since it was first isolated in 1960 [5]. In 2011 an emerging strain of WNVKUN (referred to as NSW2011) caused a major outbreak of equine encephalitis in SE Australia [4,6]. WNVNY99 was found to be associated with neurological and gastrointestinal disease and high mortality in farmed alligators in Georgia, Louisiana and Florida. Skin lesions were noted in animals surviving the acute infection, generally appearing 4–5 weeks after the acute disease, but it was not until later that a direct association between the skin lesions, known as “pix” or “Lymphohistiocytic proliferative cutaneous lesions”
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