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Journal of Veterinary Diagnostic Investigation Vol. 20 Issue 3, 325-328
Copyright © 2008 by the American Association of Veterinary Laboratory Diagnosticians
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Brief Communication

Real-time fluorogenic reverse transcription polymerase chain reaction assay for detection of African horse sickness virus

Montserrat Agüero, Concepción Gómez-Tejedor, María Ángeles Cubillo, Consuelo Rubio, Esther Romero and Angel Jiménez-Clavero1

Correspondence: 1Corresponding Author: Miguel Angel Jiménez-Clavero, CISA-INIA, Ctra Algete-El Casar, s/n, 28130, Valdeolmos, Madrid, Spain, e-mail: majimenez{at}inia.es

African horse sickness is an arthropod-borne disease of the equine included in the World Organization for Animal Health (OIE) list with important economic consequences for horse trade. The disease is caused by African horse sickness virus (AHSV; family Reoviridae, genus Orbivirus), which is transmitted by Culicoides midges. It is endemic in sub-Saharan Africa, spreading occasionally outside this area where the occurrence of Culicoides vectors allows virus transmission. Currently, only conventional (gel-based) reverse transcription polymerase chain reaction (RT-PCR) protocols are available for its detection; however, these methods are cumbersome and difficult to apply when large numbers of samples are to be tested, as in the case of epizootics. To overcome this problem, a real-time RT-PCR method has been developed, based on a 5'-Taq nuclease-3'-minor groove binder-DNA probe (TaqMan MGB) for detection of a wide range of AHSV serotypes and strains designed to the highly conserved region of the VP7 gene (segment 7). The method was able to detect all prototype strains from the 9 known serotypes of the virus, with a high analytical sensitivity; no cross-reactions were observed with other orbiviruses or with other viruses affecting horses. The diagnostic sensitivity was assessed using a panel of AHSV-positive tissue samples from an epizootic that occurred in Spain between 1987 and 1990. This method, which can be performed in 96-well format, is suitable for large-scale surveillance of AHSV in areas where it can potentially spread.

Key Words: African horse sickness virus • fluorogenic • real-time reverse transcription polymerase chain reaction






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