Isolation and Identification of Dengue Virus I from Zhongshan City and Sequence Analysis of E/NS1 Gene

Objective To isolate and identify the epidemic strains of dengue virus in Zhongshan city in 2007 and 2012,and analyze their origins and phylogeny at the molecular level.Methods Dengue viruses from the sera of 10 patients with positive dengue virus specific IgM antibody in 2007 and of 4 patients with positive dengue virus nucleic acid in 2012 were isolated with C6/36 cells.The serotypes were identified by real time-PCR.E/NS1 genes were amplified by RT-PCR and sequenced.The homology of the epidemic strains in Zhongshan city with those from different regions and with the international standard strains were compared.Their phylogeny was analyzed.Results Six dengue virus strains isolated were all identified as dengue virus I,among which 2 strains were from 2007 and named 2007ZSDenguel-2 and 4 from 2012 and named 2012ZSDengue1-4.The sequence similarities of the 2012ZSDengue1-2 with the dengue virus I strains prevalent in Cambodia and Vietnam in recent years were higher than 99.0%,and the evolutionary distances were the shortest(0.011-0.012).The sequence similarities of the 2012ZSDengue3-4,2007ZSDenguel-2 with the dengue virus I strains prevalent in Zhuhai in 2007 and in Singapore,Japan,Thailand in recent years were higher than 99.0%,and the evolutionary distances were the shortest(0.007-0.024).The sequence similarities of the strains in Zhongshan city with the dengue virus I international standard strains,the prevalent strains in Africa and America were between 90.0%-94.0%,and the evolutionary distances were far(0.065-0.108).The sequence similarities with dengue virus Ⅱ,Ⅲ,Ⅳ international standard strains NGC,H87,H241 were about 68.0%,66.0% and 71.0%,and the evolutionary distances were the farthest(0.369-0.505).Conclusions The epidemic strains of dengue virus in Zhongshan city in 2007 and 2012 are dengue virus type I.It could be imported strains from Southeast Asia but have different origins.