狂犬病病毒Vero ERA株G蛋白333位点精氨酸突变株的构建

Rescue of Vero Cell Adapted Rabies Virus ERA Vaccine Strain with R333E Mutation in Glycoprotein

狂犬病病毒糖蛋白(G)在病毒的致病性中起着主要作用,其第333位点精氨酸(Arg)是决定病毒神经细胞侵嗜性的重要分子基础之一。为研制更加安全有效的狂犬病减毒活疫苗,本试验采用负链RNA病毒反向遗传学方法,在体外将狂犬病病毒Vero细胞适应株ERA糖蛋白G333位点精氨酸突变为谷氨酸,构建减毒株ERAGΔ,减弱了其潜在的毒力返强的危险。救获的ERAGΔ突变株在Vero细胞上表现出与ERA株亲本病毒相似的生长动力学特性。本研究成功构建并拯救出了狂犬病病毒ERA减毒疫苗株,为研制开发狂犬病病毒无毒疫苗提供了候选株。

The glycoprotein （G） of rabies virus （RV） is known to play a predominant role in the pathogenesis of rabies. And G333 （Arg） is one of molecular basis that decided neurocyte tropism in rabies virus. For developing new vaccine of rabies virus which is safer and has good immunogenicity, based on reverse genetics, the antigen site R333 of Glycoprotein, in the Vero cell adapted ERA vaccine strain, was mutated from R to E. And rescued the attenuated vaccine strain ERAG^Δ successfully. TCID50 was between 10 ^-4 and 10^-5 , in vitro growth the virus of ERAG^Δ was reproduce after 24 hours, and the highest titer was in five days, then fall-off. The result shows the same typical grows kinetics with parents ERA in Vero cells. It was a substantial technique for the next research on attenuated vaccine. In this experiment, the attenuated vaccine strain ERAGa was rescued successfully, and provided a good candidate for vaccine. It has great value for next research and practical application.