Rapid detection of peste des petits ruminants virus by a reverse transcription loop-mediated isothermal amplification assay.
Peste des petits ruminants virus (PPRV) is the causative agent of peste des petits ruminants (PPR), an economically important viral disease of small ruminants. In this report, a one-step, single-tube, reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed for the detection of PPRV. A set of six LAMP primers were designed based on the matrix gene sequence of PPRV to amplify the target RNA by incubation at 63°C for 60min with Bst DNA polymerase and reverse transcriptase. The amplified products could be observed by the naked eye. The specificity of the RT-LAMP assay was validated by amplifying eight strains of PPRV isolated in different geographical areas. No cross-reactivity with other related viruses, including rinderpest virus, canine distemper virus and measles virus, was detected. The sensitivity of the assay was similar to that of real-time reverse transcription polymerase chain reaction (RT-PCR) and 10-fold higher than that of conventional RT-PCR. Twenty clinical samples were evaluated by the RT-LAMP assay, and the results were consistent with those of real-time RT-PCR. As a simple, rapid and accurate detection method, this RT-LAMP assay has important potential applications in the clinical diagnosis of PPR and the surveillance of PPRV.
Li
Lin
L
China Animal Health and Epidemiology Center, Qingdao, China.
Bao
Jingyue
J
Wu
Xiaodong
X
Wang
Zhiliang
Z
Wang
Junwei
J
Gong
Mingxia
M
Liu
Chunju
C
Li
Jinming
J
D000818:Animals; D001483:Base Sequence; D003429:Cross Reactions; D004587:Electrophoresis, Agar Gel; D005456:Fluorescent Dyes; D015511:Goat Diseases / Q000175:diagnosis / Q000821:virology; D006041:Goats / Q000821:virology; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D029021:Peste-des-Petits-Ruminants / Q000175:diagnosis / Q000662:veterinary* / Q000821:virology; D018107:Peste-des-petits-ruminants virus / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*; D012367:RNA, Viral / Q000032:analysis* / Q000302:isolation & purification; D020133:Reverse Transcriptase Polymerase Chain Reaction; D048348:Reverse Transcription; D012680:Sensitivity and Specificity; D012756:Sheep / Q000821:virology; D012757:Sheep Diseases / Q000175:diagnosis / Q000821:virology; D013194:Staining and Labeling / Q000379:methods
D005456:Fluorescent Dyes; D012367:RNA, Viral