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Specificity and Geographic Range of Detection of a Multiplex Real-Time ReverseTranscriptase PCR Assay for Vesicular Stomatitis Virus

Jessica Snyder1, George R. Smoliga2, and Luis L. Rodriguez2

1 Texas A&M University, College of Veterinary Medicine, College Station, TX, 77845

2ARS, USDA, Plum Island Animal Disease Center, Greenport, NY, 11944

Background: Vesicular Stomatitis (VS) is an important disease of horses and livestock caused by Vesicular Stomatitis Virus (VSV), an arbovirus in the genus Vesiculovirus of the family Rhabdoviridae.  Because VS is clinically indistinguishable from Foot-and-Mouth Disease (FMD), an economically devastating viral disease of cloven-hoofed animals, it is important to rule-out FMD during outbreaks of vesicular disease.  Therefore, much work has been devoted to the development of a rapid, portable multiplex real-time reverse transcriptase (rRT) PCR assay capable of detecting VSV strains of the two common serotypes New Jersey (NJ) and Indiana 1 (IN1) from various geographical regions. 

Objective: In this study specificity and geographic range of detection of the assay were determined.

Procedures: RNA was extracted from 145 previously obtained Bovine and Equine epithelium and tissue culture supernatant samples from 8 North American, Central American and northern South American countries and from VSV subtypes, near neighbors and look-alike viruses, including all 7 FMDV serotypes.  rRT-PCR was performed on the extracted RNA using a Smart Cycler II (Cepheid).  The TaqMan probe target area of any sample not detected by the rRT-PCR assay was sequenced.

Results: The rRT-PCR assay was not positive for any of the VSV subtypes, near neighbors or look-alike viruses tested (100% specificity).  Virus was detected by the rRT-PCR assay in samples from all 8 countries tested.  Six samples, all serotype NJ and all from El Salvador, were not detected and sequence data from these was used to design a new NJ probe (testing of new probe in progress).

Conclusions: The rRT-PCR assay for VSV is highly specific for the VSV serotypes NJ and IN1.  The assay has a wide geographic range of detection that is expected to be expanded by the NJ probe designed in this study.