Development of real-time RT-PCR for the detection of low concentrations of Rift Valley fever virus

• Published in: Journal of virological methods 2014-01, Vol.195, p.92-99
• Main Authors: Maquart, Marianne, Temmam, Sarah, Héraud, Jean-Michel, Leparc-Goffart, Isabelle, Cêtre-Sossah, Catherine, Dellagi, Koussay, Cardinale, Eric, Pascalis, Hervé
• Format: Article
• Language: English
• Subjects: Animals; Bioengineering; Comoros; Diagnostic tool; Inter-epidemic period; Life Sciences; Madagascar; Microbiology and Parasitology; Polymerase Chain Reaction; Polymerase Chain Reaction - methods; Real-Time Polymerase Chain Reaction; Real-Time Polymerase Chain Reaction - methods; Reverse Transcriptase Polymerase Chain Reaction; Reverse Transcriptase Polymerase Chain Reaction - methods; Rift Valley Fever; Rift Valley Fever - diagnosis; Rift Valley Fever - veterinary; Rift Valley Fever - virology; Rift Valley fever virus; Rift Valley fever virus - isolation & purification; Ruminants; Santé publique et épidémiologie; Semi-nested RT-PCR; Sensitivity and Specificity; Veterinary Medicine; Veterinary Medicine - methods; Virology; Virology - methods
• ISSN: 0166-0934; 1879-0984
• DOI: 10.1016/j.jviromet.2013.10.001

•A highly sensitive real-time PCR assay for the detection of Rift Valley Fever virus.•Sensibility tested on 2756 serum from Madagascar and the Comoros archipelago.•Comparison with a RVF specific quantitative real time RT-PCR reference technique.•System specific and 7.7 times more sensitive than the reference technique.•Diagnostic tool for the detection of low viral loads in inter-epizootic periods. In recent years, Madagascar and the Comoros archipelago have been affected by epidemics of Rift Valley fever (RVF), however detection of Rift Valley fever virus (RVFV) in zebu, sheep and goats during the post epidemic periods was frequently unsuccessful. Thus, a highly sensitive real-time RT-PCR assay was developed for the detection of RVFV at low viral loads. A new RVF SYBR Green RT-PCR targeting the M segment was tested on serum from different RVF seronegative ruminant species collected from May 2010 to August 2011 in Madagascar and the Comoros archipelago and compared with a RVF specific quantitative real time RT-PCR technique, which is considered as the reference technique. The specificity was tested on a wide range of arboviruses or other viruses giving RVF similar clinical signs. A total of 38 out of 2756 serum samples tested positive with the new RT-PCR, whereas the reference technique only detected 5 out of the 2756. The described RT-PCR is an efficient diagnostic tool for the investigation of enzootic circulation of the RVF virus. It allows the detection of low viral RNA loads adapted for the investigations of reservoirs or specific epidemiological situations such as inter-epizootic periods.