title: Development and validation of one-step SYBR green real-time RT-PCR for the rapid detection of newly emerged duck Tembusu virus.
abstract: Duck Tembusu virus (DTMUV) is a single-stranded positive-sense RNA virus that causes disease to emerge in duck flocks and results in huge economic losses to the duck industry. However, no vaccines and control measures are available in China to date. Development of reliable and fast detection methods is necessary to prevent and control this disease. Therefore, a one-step SYBR Green real-time reverse transcription polymerase chain reaction (RT-PCR) method is established here for DTMUV detection. The results show that the method can specifically detect DTMUV without cross-reactions with selected avian pathogens. The sensitivity of the assay was 1000 times greater than that of a conventional RT-PCR and able to test as few as 20 copies from RNA standard samples. The coefficients of variations of inter- and intra-assay values ranged from 0.09% to 0.36% and 0.1% to 0.23%, respectively. Testing 168 field samples and 96 experimentally infected samples by conventional RT-PCR and the one-step SYBR Green real-time RT-PCR, the positive rates were 35.1% and 73.8% from field samples and 30.2% and 64.6% from infected samples. The one-step SYBR Green real-time RT-PCR developed in this study was shown to be a sensitive, specific, high-throughput, cost-effective, and simple diagnostic tool for the rapid detection and epidemiological surveillance of the emerging DTMUV infection.
authors.affiliation: State Key Laboratory of Veterinary Etiological Biology, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, 1 Xujiaping, Chengguan District, Lanzhou 730046, PR China.
authors.forename: Zongliang
authors.initials: Z
authors.lastname: Liu
authors.forename: Yuguang
authors.initials: Y
authors.lastname: Fu
authors.forename: Yanhong
authors.initials: Y
authors.lastname: Ji
authors.forename: Jianzhong
authors.initials: J
authors.lastname: Wei
authors.forename: Xuepeng
authors.initials: X
authors.lastname: Cai
authors.forename: Qiyun
authors.initials: Q
authors.lastname: Zhu
mesh_terms: D000818:Animals; D052160:Benzothiazoles; D002681:China; D003959:Diamines; D004372:Ducks; D005416:Flavivirus / Q000302:isolation & purification*; D018177:Flavivirus Infections / Q000175:diagnosis / Q000662:veterinary* / Q000821:virology; D009930:Organic Chemicals; D011201:Poultry Diseases / Q000175:diagnosis* / Q000821:virology; D011804:Quinolines; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity
chemical_list: D052160:Benzothiazoles; D003959:Diamines; D009930:Organic Chemicals; D011804:Quinolines; C098022:SYBR Green I