Rhesus monkey rhadinovirus (RRV): construction of a RRV-GFP recombinant virus and development of assays to assess viral replication

Rhesus monkey rhadinovirus (RRV) is a γ-2-herpesvirus that is closely related to Kaposi’s sarcoma-associated herpesvirus (KSHV/HHV-8). Lack of an efficient culture system to grow high titers of virus, and the lack of an in vivo animal model system, has hampered the study of KSHV replication and path...

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Bibliographic Details
Published in:Virology (New York, N.Y.) N.Y.), 2003-07, Vol.312 (1), p.122-134
Main Authors: DeWire, Scott M, Money, Eric S, Krall, Stuart P, Damania, Blossom
Format: Article
Language:English
Subjects:
Animals
Antiviral Agents - pharmacology
DNA, Recombinant - genetics
Genes, Viral
Genetic Engineering
Genome, Viral
Green Fluorescent Proteins
Luminescent Proteins - analysis
Luminescent Proteins - genetics
Macaca mulatta - virology
Plaque assay
Real-time PCR
Recombinant rhadinovirus
Rhadinovirus - drug effects
Rhadinovirus - genetics
Rhadinovirus - physiology
Rhesus monkey rhadinovirus
Viral Plaque Assay
Virus Replication - drug effects
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Summary:Rhesus monkey rhadinovirus (RRV) is a γ-2-herpesvirus that is closely related to Kaposi’s sarcoma-associated herpesvirus (KSHV/HHV-8). Lack of an efficient culture system to grow high titers of virus, and the lack of an in vivo animal model system, has hampered the study of KSHV replication and pathogenesis. RRV is capable of replicating to high titers on fibroblasts, thus facilitating the construction of recombinant rhadinoviruses. In addition, the ability to experimentally infect naïve rhesus macaques with RRV makes it an excellent model system to study γ-herpesvirus replication. Our study describes, for the first time, the construction of a GFP-expressing RRV recombinant virus using a traditional homologous recombination strategy. We have also developed two new methods for determining viral titers of RRV including a traditional viral plaque assay and a quantitative real-time PCR assay. We have compared the replication of wild-type RRV with that of the RRV-GFP recombinant virus in one-step growth curves. We have also measured the sensitivity of RRV to a small panel of antiviral drugs. The development of both the recombination strategy and the viral quantitation assays for RRV will lay the foundation for future studies to evaluate the contribution of individual genes to viral replication both in vitro and in vivo.
ISSN:0042-6822
1096-0341
DOI:10.1016/S0042-6822(03)00195-8