Development of a TaqMan®–Minor Groove Binding Protein Assay for the Detection and Quantification of Crimean-Congo Hemorrhagic Fever Virus

Abstract

Crimean-Congo hemorrhagic fever virus (CCHFV) is a tick-borne virus of the genus Nairovirus and the family Bunyaviridae. It is a negative-strand RNA virus comprised of small (S), medium (M), and large (L) genome segments. The S segment encodes for nucleocapsid protein, the M segment codes for envelope glycoproteins (Gn and Gc), and the L segment codes for the RNA-dependent RNA polymerase. Currently, there are a limited number of methods for rapidly diagnosing CCHFV infections. We developed a real-time, reverse transcription-polymerase chain reaction assay for the rapid detection of CCHFV by using the TaqMan((R))-minor groove binding protein probe technology. The primers and probes were designed to amplify and detect a region in the S segment of CCHFV that is conserved across multiple strains. The limit of detection of the assay was 10 genome copies of RNA. This primer and probe set was specific to 18 strains of CCHFV tested and did not cross-react with either a DNA panel of 78 organisms or a panel of 28 diverse RNA viruses. This will rapidly and specifically detect CCHFV, and it has been used to detect CCHFV infection in samples from humans, animals, and ticks.