Author
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MECHAM, JAMES |
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Submitted to: American Society for Virology Meeting
Publication Type: Abstract Only Publication Acceptance Date: 7/12/2004 Publication Date: 7/12/2004 Citation: Mecham, J.O. 2004. Detection and quantitation of bluetongue virus in insect cell culture [abstract]. American Society for Virology Meeting, p35-7, p. 270. Interpretive Summary: Bluetongue virus (BTV) infects sheep, cattle and other ruminants by small biting midges (Culicoides spp.). Disease caused by this virus is classified as a list A disease by the Office of International Epizootics (OIE), and causes considerable economic loss worldwide due to both direct losses and trade restrictions on the movement of livestock and livestock products. Virus isolation is required for diagnosis and typically involves the use of cell cultures derived from vertebrates. Although quite sensitive for virus isolation from infected mammals, these cells may have limited sensitivity for virus isolation from invertebrate insects. Cell cultures derived from the Culicoides vector have been developed in this laboratory and may be useful for BTV isolation and for better understanding viral replication in the insect vector. However, detection of BTV in these insect cells has required co-cultivation with susceptible vertebrate cells or staining techniques that are not easily quantitated. Techniques were developed for the direct detection and quantitation of BTV in infected Culicoides cell culture by enzyme-linked immunosorbent assay (ELISA) and by direct staining. These assays should facilitate the use of these insect cells for virus isolation and studies of BTV replication. Technical Abstract: Bluetongue virus (BTV) infects sheep, cattle and other ruminants and is transmitted by Culicoides spp. of biting midges. Virus is typically isolated and characterized by infection of susceptible vertebrate cells that undergo detectable and measurable cytopathic effects. Cell lines derived from C. sonorensis provide tools for better understanding BTV infection of the vector insect. These cell lines may also be more sensitive, compared to vertebrate cell culture, for isolation of virus from field collected insects. However, the use of these insect cell cultures for virus isolation and characterization is hampered because BTV does not produce significant cytopathic effects in these cells. Detection typically requires co-cultivation of the infected cells with susceptible vertebrate cell culture or staining methods that are not easily quantitated. This report describes the use of enzyme-linked immunosorbent assays and In situ fluorescent staining techniques to directly detect and quantitate BTV in infected Culicoides cell lines. The sensitivity and specificity of these assays should facilitate the use of these cell lines for virus isolation and studies of BTV replication. |
