登革3型病毒义乌流行株免疫原性研究(英文)

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

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。

This study performed phylogenic analysis on a dengue strain isolated from an outbreak of dengue fever in 2009 at Yiwu City of Zhejiang Province, China, and further to analyze the immunogenicity of E protein of this viral isolate. Firstly, the viral genome was amplified by RT-PCR and phylogenetic trees were constructed by MEGA 4 based on both nucleotide and amino acid sequences of E and NS1 proteins. The phylogenetic analysis showed that the similarity of Yiwu strain with the Guangzhou GZID3 strain and the India GWL-25 strain was over 990/00. Secondly, the expression plasmid of E protein was con- structed and transfected into 293T cells. The secreted E protein were then purified by sucrose density gradient centrifugation and used to inoculate BALB/c mice. The humoral immunity was evaluated by ELISA and neutralizing antibody analysis. Re- sults showed that the E protein of Yiwu strain could induce dengue specific IgG antibodies and neutralizing antibodies. There- fore, the study found that the Yiwu strain was classified into the subtype III of dengue virus type 3 （DENV-3）, and the E pro tein of this strain had strong immunogenicity