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ARS Home » Midwest Area » Ames, Iowa » National Animal Disease Center » Virus and Prion Research » Research » Publications at this Location » Publication #218285
Title: Development of realtime PCR for rapid detection and differentiation of field pseudorabies and vaccine viruses

Author
item MA, WENJUN - IOWA STATE UNIVERSITY
item Lager, Kelly
item Richt, Juergen
item Stoffregen, William
item ZHOU, FANGHONG - IOWA STATE UNIVERSITY
item JANKE, BRUCE - IOWA STATE UNIVERSITY
item YOON, KYOUNG-JIN - IOWA STATE UNIVERSITY

Submitted to: Journal of Veterinary Diagnostic Investigation
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/1/2008
Publication Date: 7/1/2008
Citation: Ma, W., Lager, K.M., Richt, J.,A. Stoffregen, W.C., Zhou, F., Janke, B.H., Yoon, K.J.. 2008. Development of real-time polymerase chain reaction assays for rapid detection and differentiation of wild-type pseudorabies and gene-deleted vaccine viruses. Journal of Veterinary Diagnostic Investigation. 20(4):440-447.

Interpretive Summary: The successful eradication of pseudorabies in U.S. domestic swine was accomplished through the use of modified-live virus vaccines, and an accompanying blood test that could differentiate vaccinated swine from swine that have been infected with pseudorabies virus (PRV), the virus that causes pseudorabies. Some feral swine in the U.S. are infected with pseudorabies virus, and they represent a potential reservoir of this virus for infection of domestic swine and other native wildlife. A critical need for the current PRV surveillance program is the rapid detection of PRV. For this reason, a multiplex real-time PCR test was developed and evaluated for its capability in the detection and differentiation of field and vaccine strains of PRV. Archived samples from previous PRV field cases were analyzed with this test as well as samples that had recently been collected from feral swine. This test was shown to be able to distinguish between vaccine and field strains of PRV, and it could identify virus in feral swine samples. These observations demonstrated the potential application of this multiplex real-time PCR for rapid and specific detection of PRV in domestic and feral swine.

Technical Abstract: The successful eradication of pseudorabies in U.S. domestic swine was accomplished through the use of gE-deleted modified live virus vaccines and an accompanying gE differential ELISA. Pseudorabies virus (PrV) has been established in feral swine in the United States, and they represent a potential reservoir of this virus for infection of domestic swine and other native wildlife. A critical need for the current PrV surveillance program in the United States is the rapid detection of PrV. For this reason, a multiplex real-time PCR using TaqMan chemistry was developed and evaluated for its capability in the detection and differentiation of field and vaccine strains of PrV. Based on sequence information available in GeneBank and sequences of all PrV marker vaccines which were commercially available, PCR primers and probes were designed for gB and gE genes for general detection and differentiation, respectively. The realtime PCR assay could detect all PrV from diagnostic submissions and vaccine strains in a differential manner. The analytical sensitivity of the assay was approximately 0.1 PFU per reaction. The assay was specific for PrV as no positive results were obtained from testing common swine viral pathogens and other herpesviruses. These observations demonstrated the potential application of the developed multiplex real-time PCR for rapid and specific detection of PRV in domestic and feral swine, as well as non-porcine species that can be infected with PRV and serve as carriers.