title: Development and evaluation of a real-time reverse transcription-loop-mediated isothermal amplification assay for rapid detection of Rift Valley fever virus in clinical specimens.
abstract: This paper reports on the development and validation of a real-time reverse transcription-loop-mediated isothermal amplification assay (RT-LAMP) targeting the genomic large RNA segment of Rift Valley fever virus (RVFV). The set of six designed RT-LAMP primers identified strains of RVFV isolated in geographically distinct areas over a period of 50 years; there was no cross-reactivity with other genetically related and unrelated arboviruses. When testing serial sera and plasma from sheep experimentally infected with wild-type RVFV, there was 100% agreement between results of the RT-LAMP, a TaqMan-based real-time PCR, and virus isolation. Similarly, the assay had very high levels of diagnostic sensitivity and specificity when testing various clinical specimens from humans and animals naturally infected with the virus during recent outbreaks of the disease in Africa. The detection of specific viral genome targets in positive clinical specimens was achieved in less than 30 min. As a highly accurate, rapid, and very simple nucleic acid detection format, the RT-LAMP has the potential to be used in less-well-equipped laboratories in Africa and as a portable device during RVF outbreaks in remote areas, and it can be a valuable tool for the differential diagnosis of viral hemorrhagic fevers.
authors.affiliation: Special Pathogens Unit, National Institute for Communicable Diseases of the National Health Laboratory Services, Private Bag X4, Sandringham 2131, Republic of South Africa.
authors.forename: C A
authors.initials: CA
authors.lastname: Le Roux
authors.forename: T
authors.initials: T
authors.lastname: Kubo
authors.forename: A A
authors.initials: AA
authors.lastname: Grobbelaar
authors.forename: P Jansen
authors.initials: PJ
authors.lastname: van Vuren
authors.forename: J
authors.initials: J
authors.lastname: Weyer
authors.forename: L H
authors.initials: LH
authors.lastname: Nel
authors.forename: R
authors.initials: R
authors.lastname: Swanepoel
authors.forename: K
authors.initials: K
authors.lastname: Morita
authors.forename: J T
authors.initials: JT
authors.lastname: Paweska
mesh_terms: D000349:Africa; D000818:Animals; D017931:DNA Primers / Q000235:genetics; D006801:Humans; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D012295:Rift Valley Fever / Q000175:diagnosis* / Q000662:veterinary*; D012296:Rift Valley fever virus / Q000235:genetics* / Q000302:isolation & purification*; D012680:Sensitivity and Specificity; D012756:Sheep; D012757:Sheep Diseases / Q000175:diagnosis*; D013997:Time Factors
chemical_list: D017931:DNA Primers