Nested PCR Amplification and Sequencing of a Reptile Reovirus Associated with Disease in Mojave Rattlesnakes (Crotalus scutulatus)
Abstract
Reoviruses have been shown to cause fatal pneumonia and subacute tracheitis in reptiles.1 Diagnosis is challenging, as there are no inclusions, and histologic lesions resemble paramyxoviral disease. Serologic diagnosis has been limited to virus neutralization,2 which is labor intensive and may not cross-react between strains. As reptile reoviruses do not hemagglutinate, hemagglutination inhibition is not an option. No published sequence for reptile reoviruses has been available. RNA-dependent RNA polymerase sequences from mammalian orthoreoviruses and piscine aquareoviruses were aligned. Degenerate primers were designed targeting conserved regions. These primers were used in a nested PCR to amplify sequences from a reovirus isolate associated with an outbreak of disease in Mojave rattlesnakes (Crotalus scutulatus). Nucleotide sequencing of the PCR products showed that the reoviral sequences from these snakes were novel. Comparative sequence analysis shows that these viruses are probable members of the genus Orthoreovirus. These primers may be of use for obtaining initial sequence data from novel reoviruses. Sequence data will enable design of diagnostic PCR testing for specific viruses.
Literature Cited
1. Lamirande, E.W., D.K. Nichols, J.W. Owens, J.M. Gaskin, and E.R. Jacobson. 1999. Isolation and experimental transmission of a reovirus pathogenic in ratsnakes (Elaphe species). Virus Res. 63:135–41.
2. Marschang, R.E., S. Donahoe, R. Manvell, and J Lemos-Espinal. 2002. Paramyxovirus and reovirus infections in wild-caught Mexican lizards (Xenosaurus and Abronia spp.). J Zoo Wildl Med. 33:317–21.