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目的分离鉴定中山市2007年及2012年流行的登革病毒,从分子水平分析其地域来源和系统进化情况。方法用C6/36细胞分离10份2007年登革病毒特异性IgM抗体阳性血清和4份2012年登革病毒核酸阳性血清中的病毒,采用Real time-PCR对毒株进行血清型鉴定,RT-PCR扩增毒株E/NS1基因并进行序列测定,分析其与不同区域流行株和国际标准株的同源性和系统进化情况。结果共分离鉴定到6株登革I型病毒,其中2株为2007年流行株,4株为2012年流行株,分别命名为2007ZSDengue1-2和2012ZSDengue1-4。2012ZSDengue1-2与柬埔寨、越南等东南亚国家近年流行的登革I型毒株相似性高(大于99.0%),进化距离最短(0.011~0.012)。2012ZSDengue3-4,2007ZSDengue1-2与广东省珠海市2007年流行和新加坡、日本、泰国等亚洲国家近年流行的登革I型毒株相似性高(大于99.0%),进化距离最短(0.007~0.024)。中山流行株与登革I型国际标准株、非洲和美洲流行株相似性在90.0%~94.0%之间,进化距离较远(0.065~0.108);与登革Ⅱ、Ⅲ、Ⅳ型国际标准株NGC、H87、H241相似性约为68.0%、66.0%、71.0%,进化距离最远(0.369~0.505)。结论中山市2007年及2012年流行株为登革I型病毒,可能属于东南亚国家输入性毒株,但输入源头有所不同。
Objective To isolate and identify Dengue virus which was endemic to Zhongshan City in 2007 and 2012 and analyze its geographical origin and phylogenetic status at the molecular level. Methods Ten strains of Dengue virus-specific IgM antibody-positive sera and four strains of Dengue virus-positive sera from 2012 were isolated from C6 / 36 cells. The serotypes were identified by Real time-PCR. RT- The E / NS1 gene was amplified by PCR and sequenced. The homology and phylogenetic analysis of the E / NS1 gene with different strains and international standard strains were analyzed. Results Six strains of Dengue I virus were isolated and identified. Two of them were epidemic strains in 2007 and four were epidemic strains in 2012, which were named 2007ZSDengue1-2 and 2012ZSDengue1-4.2012ZSDengue1-2 respectively. Southeast Asia such as Cambodia and Vietnam In recent years, the epidemic of Dengue I strain in China has a high similarity (more than 99.0%) and the shortest evolutionary distance (0.011 ~ 0.012). 2012ZSDengue3-4,2007ZSDengue1-2 has the highest similarity (more than 99.0%) with the dengue type I strains prevalent in Zhuhai in 2007 and the Asian countries such as Singapore, Japan and Thailand in recent years, and the shortest evolutionary distance (0.007 ~ 0.024) . The epidemic strains of Zhongshan and Dengue type I international standard strains, Africa and the Americas epidemic strains in the 90.0% ~ 94.0% similarity range, the evolutionary distance is far (0.065 ~ 0.108); and Dengue Ⅱ, Ⅲ, Ⅳ international standard strains The similarities of NGC, H87 and H241 were 68.0%, 66.0% and 71.0%, respectively, and the farsighted distance (0.369-0.505). Conclusion The epidemic strains of dengue type I in Zhongshan in 2007 and 2012 may belong to the imported strains in Southeast Asia, but the sources of their input are different.