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- Title
融合域点突变的埃博拉病毒GP蛋白重组新城疫病毒的 构建与免疫评价.
- Authors
张海琳; 王金良; 王 翀; 温志远; 刘任强; 步志高; 葛金英
- Abstract
Zaire Ebola virus (EBOV) can cause severe hemorrhagic fever in humans with a case fatality rate of up to 90% and poses a significant threat to public health. Envelope protein (GP) is the main immunogenic protein of EBOV. Our previous study showed that the incorporation of EBOV GP protein into NDV particles significantly altered the entry of NDV into cells. In order to develop a safer Ebola vaccine, the GP of EBOV was mutated into GP(I532R), and a recombinant NDV expressing the mutant Zaire Ebola virus glycoprotein rLa- EBOVGP(I532R) was rescued via reverse genetics. The recombinant virus rLa- EBOVGP(I532R) was continuously passaged in embryonated chicken eggs and its genetic stability during in vitro propagation was evaluated. RT- PCR results showed that EBOV GP(I532R) was correctly inserted into the NDV, which indicated that rLa-EBOVGP(I532R) was rescued successfully. The recombinant virus was passaged 20 times in embryonated chicken eggs, and the EBOV GP(I532R) in the virus from each passage was examined by RT-PCR and sequencing. The result showed that the GP(I532R) gene could maintain its genetic stability throughout the propagation in embryonated chicken eggs. rLa, rLa-EBOVGP, and rLa-EBOVGP(I532R) were purified by sucrose gradient centrifugation followed by western blot detection. The results showed that GP(532R) could be expressed correctly. The localization of the GP(I532R) at the cell membrane of BHK- 21 cells infected with the recombinant virus was confirmed by confocal assay. Transmission electron microscopy revealed that the rLa-EBOVGP(I532R) had the same morphological characteristics as those of rLa and observed the expression of GP (I532R) on the surface of the chimeric virus. The growth kinetic curve of rLaEBOVGP(I532R) in embryonated chicken eggs was similar to that of parent strain. Mice immunization assay demonstrated that intranasal and intramuscular immunization with rLa-EBOVGP(I532R) could induce NVD specific neutralizing antibody and EBOV GP specific neutralizing antibody significantly in mice. The above results indicate that rLa-EBOVGP(I532R) has the potential to be a vaccine candidate against EBOV infection. This study also lays a foundation for further study of the biological function of GP (I532R).
- Publication
Chinese Journal of Preventive Veterinary Medicine / Zhongguo Yufang Shouyi Xuebao, 2021, Vol 43, Issue 2, p171
- ISSN
1008-0589
- Publication type
Article
- DOI
10.3969/j.issn.1008-0589.202005005