蚊媒体内登革病毒的快速分型检测

The Rapid Detection and Typing of Dengue Virus in Mosguito

目的建立一种用于蚊媒体内登革病毒(DEN)快速检测和分型的实验方法。方法利用标记地高辛(Dig)的登革病毒通用引物进行RT-PCR扩增蚊媒体内的病毒RNA,然后以反向斑点杂交法检测病毒扩增片段并进行分型。结果RT-PCR可扩增出预期的DEN基因片段,杂交检测和分型结果特异,易于判断,可测出3pg的病毒RNA。结论该方法灵敏、特异、稳定性好,可用于登革病毒分型及混合感染的检测,4h左右即可获得结果,为早期快速诊断蚊媒体内的登革病毒提供了一种可靠方法。

Objective To establish a rapid and stable method for detection and typing of dengue virus(DEN) in mosquito,the major vector of DEN. Methods A pair of universal primes labeled with Dig was designed according to the highly reserved domain of DEN type 1 4. It was used in the RT PCR to amplify DEN RNA extracted from DEN infected mosquitos. Then the product was hybridized with the universal and type specific probes which have been inoculated on nylon membrane. Results The prospective fragment of DEN sequence can be amplified. The effects of detecting and typing by hybridization are specific and objective. As low as 3pg DEN RNA can be detected. And it can also identify concurrent infection. Conclusion This assay maybe helpful to develop a surveillance system to provide an early warning of DF epidemics and to furnish information for epidemiology and the effective vector control measures.