Abstract
Venezuelan equine encephalitis virus (VEEV) is an Alphavirus from the family Togaviridae that causes epizootic outbreaks in equids and humans in Central and South America. So far, most studies use conventional reverse transcriptase PCR assays for the detection of the different VEEV subtypes. Here we describe the development of a TaqMan quantitative real-time reverse transcriptase PCR assay for the specific detection and quantitation of all VEEV subtypes which uses in parallel a universal equine encephalitis virus control RNA carrying target sequences of the three equine encephalitis viruses. The control RNA was used to generate standard curves for the calculation of copy numbers of viral genome of Eastern equine encephalitis virus (EEEV), Western equine encephalitis virus (WEEV), and VEEV. The new assay provides a reliable high-throughput method for the detection and quantitation of VEEV RNA in clinical and field samples and allows a rapid differentiation from potentially cocirculating EEEV and WEEV strains. The capability to detect all known VEEV variants was experimentally demonstrated and makes this assay suitable especially for the surveillance of VEEV.
Highlights
Western equine encephalitis virus (WEEV), Eastern equine encephalitis virus (EEEV), and Venezuelan equine encephalitis virus (VEEV) are arthropod-borne viruses of the genus Alphavirus of the virus family Togaviridae
They are transmitted by sanguivorous mosquitoes within bird (WEEV, EEEV, and epizootic VEEV) or rodent populations (VEEV, enzootic strains), respectively
In the here presented study, we are introducing a rapid, sensitive, and reliable one-step quantitative RT-PCR assay for VEEV as well as a synthetic RNA construct which can be used as calibrator for the quantification of alpha virus associated equine encephalitis viruses
Summary
Western equine encephalitis virus (WEEV), Eastern equine encephalitis virus (EEEV), and Venezuelan equine encephalitis virus (VEEV) are arthropod-borne (arbo) viruses of the genus Alphavirus of the virus family Togaviridae. To date these viruses are restricted to the Americas but due to worldwide travelling and trade they might be introduced to other parts of the world in the future. All three equine encephalitis viruses are classified as Category B agent by the Centers for Disease Control and Prevention, Atlanta (https:// emergency.cdc.gov/agent/agentlist-category.asp). They are transmitted by sanguivorous mosquitoes within bird (WEEV, EEEV, and epizootic VEEV (epizootic strains)) or rodent populations (VEEV, enzootic strains), respectively. For some epizootic VEEV strains a productive replication followed by successful intra- and interspecies transmission cycles was observed in horses and humans [2,3,4,5,6]
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