Submitted to: Journal of Virological Methods
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 4/23/2009
Publication Date: 10/1/2009
Citation: McClenahan, S.D., Bok, K., Neill, J.D., Smith, A.W., Rhodes, C.R., Sosnovtsev, S.V., Green, K.Y., Romero, C.H. 2009. A Capsid Gene-Based Real-Time RT-PCR Assay for the Detection of Marine Vesiviruses in the Caliciviridae. Journal of Virological Methods. 161(1):12-18. Interpretive Summary: Emerging or newly discovered viruses are becoming more of a threat to both human and animal health. Some of these viruses belong to the calicivirus family of viruses. There is a group of caliciviruses that can cause disease in many different species of marine mammals, called the San Miguel sea lion viruses (SMSV). These viruses are closely related to the exotic caliciviruses that cause vesicular exanthema of swine. There is evidence that many different species of marine mammals may serve as reservoirs for these viruses to enter into domestic livestock herds. In response to this, a novel real-time PCR test was developed that was able to detect very low levels of these viruses. This test could reliably detect less than 100 copies of viral genomic RNA. In addition, it successfully detected 12 different SMSV strains. It was shown to be specific for SMSV by testing RNA from feline caliciviruses and members of the Norovirus group of caliciviruses. Testing of these latter viruses resulted in no specific amplification products. This real-time RT-PCR assay may find use as a diagnostic tool for the detection of currently circulating and previously described SMSV strains in clinical samples that include vesicular fluids, and oral and rectal swabs. It can also be employed in the screening of clinical samples that may contain unknown SMSV strains as well. This test will be especially valuable when large numbers of clinical samples are screened as in the case of surveillance for these viruses.
Technical Abstract: A real-time reverse transcription polymerase chain reaction (rtRT-PCR) assay was developed for the identification of marine vesiviruses. The primers were designed to target a 176-nucleotide fragment within a highly conserved region of the San Miguel sea lion viruses (SMSVs) capsid gene. The assay detected viral RNA from nine marine vesivirus serotypes described previously, including two serotypes (SMSV-8 and -12) not identified with presently available molecular assays, a highly related bovine vesivirus strain (Bos-1), a mink vesivirus strain (MCV), and two novel genotypes isolated recently from Steller sea lions (SSL V810 and V1415). The real-time assay did not amplify sequences from the corresponding genomic regions of feline calicivirus (also in the genus Vesivirus) and representative members of the genus Norovirus. The rtRT-PCR assay described below may prove useful as a diagnostic tool for the detection of currently circulating, emerging and previously described marine vesiviruses in clinical samples, especially when large numbers are screened in surveillance studies of these restricted viruses.