SFTSV N端糖蛋白重组腺病毒的鉴定和免疫研究

Identification and immunological study of recombinant adenovirus with Gn of severe fever with thrombocytopenia syndrome virus

目的获得表达发热伴血小板减少综合征病毒(SFTSV)N端糖蛋白(Gn)的重组人5型腺病毒(Ad5-Gn),鉴定其生物学特性及诱导产生中和抗体的能力。方法利用AdEasy腺病毒包装系统,构建pAD-Amp-Gn表达载体,转染HEK-293细胞,获得重组腺病毒Ad5-Gn。用Ad5-Gn感染细胞,利用免疫荧光染色和Western印迹鉴定SFTSV Gn的表达水平;将Ad5-Gn病毒注射BALB/c小鼠,观察小鼠行为学并检测特异性中和抗体水平,评价Ad5-Gn的安全性和免疫原性。结果免疫荧光染色和Western印迹结果表明,Ad5-Gn感染细胞后能检测到SFTSV Gn表达,且大小符合预期。Ad5-Gn不影响小鼠生长,免疫小鼠4周后产生的中和抗体平均效价为1∶341。结论获得了表达SFTSV Gn的Ad5-Gn候选疫苗,其毒力低且能诱导小鼠产生较高的中和抗体,为进一步研制SFTSV疫苗奠定了基础。

Objective To obtain a recombinant human adenovirus type 5 vector(Ad5)vaccine expressing the glycoprotein N(Gn)of severe fever with thrombocytopenia syndrome virus(SFTSV),and to identify its biological characteristics and production of neutralizing antibodies.Methods The AdEasy adenovirus packaging system was used to construct a pAD-Amp-Gn expression vector and transfect it into HEK-293 cells to obtain recombinant adenovirus Ad5-Gn.After infecting the cells with Ad5-Gn,the expression of SFTSV Gn was identified by immunofluorescence and Western blotting.BALB/c mice were injected with Ad5-Gn to observe the behavior and detect the level of specific neutralizing antibodies,and to evaluate the safety and immunogenicity of Ad5-Gn.Results Immunofluorescence and Western blotting showed that SFTSV Gn expression could be detected in Ad5-Gn infected cells,and the size was as big as expected.Ad5-Gn did not affect the growth of mice,and the average titer of neutralizing antibodies produced after 4 weeks of infection was 1∶341.Conclusion An Ad5-Gn vaccine candidate expressing SFTSV Gn has been obtained,which has low virulence and can induce high levels of neutralizing antibodies in mice.