登革3型病毒义乌流行株免疫原性研究

目的对2009年浙江省义乌市暴发流行的登革3型病毒进行系统进化分析并初步研究其E蛋白的免疫原性。方法 RT-PCR扩增义乌分离株全基因组,利用MEGA 4分别构建E及NS1区域核苷酸及氨基酸系统发生树,进行系统进化分析。构建重组质粒JESS-ywD3prM/E397JEV-pcDNA5,IFA及Western Blot检测E蛋白在293T细胞中的表达及分泌情况。将纯化后的分泌性E蛋白免疫BALB/c小鼠,通过ELISA、中和试验等方法对体液免疫进行测定。结果登革3型病毒义乌分离株与广州GZ1D3株及印度GWL-25株核苷酸及氨基酸序列同源性均达到99%以上;其分泌性E蛋白可以刺激小鼠产生登革特异性IgG抗体及针对同型病毒的中和抗体。结论义乌分离株属于登革3型病毒亚型Ⅲ,其分泌性E蛋白具有一定的免疫原性。

Biological characteristics of dengue virus type 3 isolated from Yiwu,China

To study the biological characteristics of dengue virus type 3 isolated from Yiwu,China,the genome was amplified by RT-PCR,and phylogenetic trees were constructed by MEGA 4 based on E and NS1 nucleotide and amino acid.E protein expression plasmid named JESS-ywD3prM/E397JEV-pcDNA5 was constructed and transfected into 293T cells,the supernatants containing secreted E protein were purified by sucrose density gradient centrifugation and inoculated into BALB/c mice,the humoral immunity was evaluated by ELISA and neutralization analysis.Phylogenetic tree analysis showed that the similarity with Guangzhou strain GZ1D3 and India strain GWL-25 were reached above 99%.Secreted E protein could induce dengue specific IgG antibodies and neutralizing antibodies against homotypic dengue virus.