Kono Y, Kanno T, Shimizu M, Yamada S, Ohashi S, Nakamine M, Shirai J
Exotic Diseases Research Division, National Institute of Animal Health, Tokyo, Japan.
J Vet Med Sci. 1996 Oct;58(10):941-6. doi: 10.1292/jvms.58.10_941.
A nested polymerase chain reaction (PCR) was developed to detect porcine reproductive and respiratory syndrome (PRRS) virus. A common primer set for European and North American type isolates of PRRS virus was designed for reverse transcription PCR, and a specific primer set for each of the 2 type isolates was designed for nested PCR. The PCR that used a specific primer set detected the corresponding type of the virus at a level equivalent to 1 TCID50/100 microliters, but not the other type of isolates. Therefore, the method clearly differentiated the 2 types of virus from each other. The detection of PRRS virus by the nested PCR was as sensitive as virus isolation in cultures of porcine alveolar macrophages from infected pigs at the acute stages, and was more sensitive from pigs at the convalescent stages. The infecting virus type was determined by use of 2 specific primer sets even when virus isolation was negative in naturally infected pigs. It was concluded that the nested PCR is useful for diagnosis and typing of PRRS virus and studies of persistent infection by the virus.
开发了一种巢式聚合酶链反应（PCR）来检测猪繁殖与呼吸综合征（PRRS）病毒。针对PRRS病毒的欧洲型和北美型分离株设计了一组通用引物用于逆转录PCR，并为这两种类型的分离株各设计了一组特异性引物用于巢式PCR。使用特异性引物组的PCR能检测到相应类型的病毒，其灵敏度相当于1个半数组织培养感染剂量（TCID50）/100微升，但检测不到其他类型的分离株。因此，该方法能清晰地区分这两种类型的病毒。巢式PCR检测PRRS病毒的灵敏度与在急性期感染猪的猪肺泡巨噬细胞培养物中进行病毒分离的灵敏度相当，而在恢复期猪中检测更为灵敏。即使在自然感染猪中病毒分离呈阴性，也可通过使用两组特异性引物来确定感染病毒的类型。得出结论，巢式PCR对PRRS病毒的诊断、分型以及该病毒持续感染的研究很有用。
