关键词: 登革病毒, 通用引物, SiO₂RT-PCR, 酶切分型

Abstract: Dengue viruses RNA was extracted rapidly by using silica.Dengue virus consensusprimers of types 1-4 and reverse transcriptase-polymerase chain reaction (RT-PCR) were usedfor detecting dengue virus in 55 serum specimens collected from patients with suspected denguefever in acute phase in Guangdong Province in 1995,and compared with the method of virus isolation.The results showed that the dengue viruses RNA detecting positive rate by RT-PCR was 38.18%, the sensitivity was 77.7 %, the specificity was 100%, and the agreement rate was 89.09%.The dengue viruses positive rate by using virus isolation was 49.09%,without significant differencecompared with the one in RT-PCR (χ²=2.08,P>0.05).The amplified bands of 511bp were observed in amplified products of the reference viruses strains of types 1, 2, 3,and 4,and specimensrespectively,by consensus primers,but not observed in Japanese encephalitis virus.The amplifiedproducts of dengue viruses by RT-PCR were cleaved by restriction endonucleases of Hinf Ⅰ,HaeⅢ,Hinc Ⅱ and San 3a Ⅰ,respectively,on agarose gel electrophoresis for differentiation among fourtypes of dengue viruses.The results showed that amplified products of dengue viruses of types Ⅰ- 4 were cut by Hinf Ⅰ and Hae Ⅲ, the products of types land2 were cut by Sau 3a Ⅰ, and the products of types 1,2,and 3 were cleaved by Hinc Ⅱ.The dengue viruses of 4 types can be differentiated by the cleavage of two or more restriction endonucleases.The result of typing of fivespecimens was identical with the one of the reference dengue virus of type 1.The method of this paper takes only 5 to 6 hours to detect dengue viruses RNA,and two days to differentiate the fourtypes of dengue viruses.It is a rapid and simple method for detecting and typing dengue viruses.