Quantitative multiplex assay for simultaneous detection and identification of Indiana and New Jersey serotypes of vesicular stomatitis virus

Thomas Bruun Rasmussen, Åse Uttenthal, Jovita Fernandez, Torben Storgaard

    Research output: Contribution to journalJournal articleResearchpeer-review

    Abstract

    In order to establish a rapid and reliable system for the detection of vesicular stomatitis virus (VSV), we developed a quantitative reverse transcription-PCR assay for the detection, quantification, and differentiation of the major serotypes, VSV Indiana and VSV New Jersey, using a closed-tube multiplex format. The detection system is based on the recently invented primer-probe energy transfer (PriProET) system. A region of the gene encoding the RNA-dependent RNA polymerase was amplified by using VSV-specific primers in the presence of two serotype-specific fluorescent probes. By incorporating nucleotide analogues in the primers, both serotypes were amplified with similar efficiencies. The generation of specific amplicons resulted in fluorescent signals for either of the two serotypes, and the specificities of the reactions were confirmed from the melting temperature profiles of the fluorescent probes. The limits of detection ware found to be less than 10 50% tissue culture infective doses/ml for both serotypes. The diagnostic value of the new method was tested with clinical materials from experimentally infected pigs, and it is concluded that the method is a powerful tool for the rapid identification of VSV.
    Original languageEnglish
    JournalJournal of Clinical Microbiology
    Volume43
    Issue number1
    Pages (from-to)356-362
    ISSN0095-1137
    DOIs
    Publication statusPublished - 2005

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