表达中东呼吸综合征冠状病毒S蛋白的重组痘苗病毒的构建及免疫效果初步评价

摘要

目的：以痘苗病毒天坛株为载体，构建表达中东呼吸综合征冠状病毒（MERS-CoV）S蛋白的重组病毒疫苗，并进行免疫效果评价。方法：通过PCR扩增获得去除跨膜区的S蛋白基因片段SQ，并构建重组痘苗病毒载体质粒pJSC11LacZ7.5SQ，将重组质粒与痘苗病毒同源重组并单斑纯化获得重组病毒毒株RVVMERS-SQ，重组病毒免疫小鼠后用酶联免疫吸附实验（ELISA）和假病毒微量中和实验检测其诱发的S蛋白体液免疫反应水平。结果：构建了表达MERS-CoV去跨膜区S蛋白的重组病毒RVVMERS-SQ，其免疫小鼠后诱发了强的S蛋白体液免疫反应，血清IgG抗体滴度为1∶3200，中和抗体滴度达到1∶1000。结论：重组痘苗病毒RVVMERS-SQ可在BALB/c小鼠体内诱发强的免疫反应，为MERS-CoV疫苗的研发提供了实验基础。%Objective: To construct a recombinant vaccinia virus expressing middle east respiratory syndrome coronavirus(MERS-CoV) S protein and evaluate its immunogenicity. Methods: SQ fragment was amplified by PCR in which the transmembrane region of S protein was deleted, and then inserted into a vaccinia virus transfer pla⁃simid. The recombinant vaccinia virus expressing MERS-CoV S protein was constructed through homologous recom⁃binant based on Tiantan vaccinia virus strains in chicken embryo fibroblast cells. BALB/c mice were immunized with the recombinant virus, its humoral immune responses against MERS-CoV S protein were detected by enzyme-linked immunoabsorbent assay(ELISA) and pseudovirus micro-neutralization assay. Results: The recombinant vac⁃cinia virus(RVVMERS-SQ) expressing SQ protein was constructed. Mice vaccinated with RVVMERS-SQ produced strong humoral immune response against MERS-CoV S; and could be elicited high level of serum antibodies neu⁃tralizing MERS-CoV pseudovirus. Conclusion: Recombinant vaccinia virus RVVMERS-SQ could induce a robust humoral immune response against MERS-CoV S in vaccinated mice, it may serve for further study of a vaccine against MERS-CoV.