置换HN基因对新城疫病毒LaSota株致病力的影响

Evaluation of Newcastle disease virus with derivated Hemaggluti-nin-Neuraminidase gene of mesogenic strain

摘要:

【目的】新城疫病毒的血凝素-神经氨酸酶（HN）和融合蛋白（F）在病毒装配、出芽、释放及侵入宿主细胞的过程中发挥关键作用，但HN对病毒致病力的影响程度尚不完全清楚。【方法】为探讨这一问题，本研究以中等毒力毒株Mukteswar的HN基因替换我国广泛应用的LaSota疫苗株HN基因，通过反向遗传操作技术拯救出嵌合病毒(rL-MuHN)。【结果】rL-MuHN红细胞吸附能力较亲本株rLaSota无显著升高，具有相似的细胞融合活性；嵌合病毒ICPI由rLaSota株的0.36降为0，MDT≥90，IVPI=0与rLaSota株相同，保持典型低致病力缓发型特点不变。进一步以Mukteswar株F基因替换rL-MuHN的F基因，拯救出F和HN双基因替换嵌合病毒rL-MuFHN，尽管该病毒的细胞融合能力显著提高，但其MDT、ICPI和IVPI分别为98 h，0.59和0，显示F和HN双基因替换仍未能使嵌合新城疫病毒rL-MuFHN的致病力达到中等毒力毒株Mukteswar（MDT、ICPI及IVPI分别为46 h、1.32和0.64）的水平。【结论】试验结果表明，F及HN囊膜蛋白基因之外的病毒基因组骨架背景对病毒的致病性同样具有重要的决定性意义，不同HN蛋白对嵌合病毒的致病能力的影响不同，与供体毒株毒力无关；以流行野毒株HN替代rLaSota疫苗株构建抗原针对性更强的弱毒疫苗株存在技术可行性。

关键词:重组新城疫病毒;F蛋白;HN蛋白;致病力

Abstract:

[Objective] The hemagglutinin-neuraminidase (HN) and fusion protein of Newcastle disease virus (NDV) plays a crucial role in the process of budding and infection. To understand the exact contribution of the HN gene to NDV pathogenecity, a reverse genetics system was developed using the lentogenic NDV LaSota strain . [Methods] the HN genes of an avirulent recombinant NDV strain (rLaSota) was replaced by an HN gene from Mukteswar mesogenic NDV strain by reverse genetics. Furthermore, the F gene of rL-MuHN was replaced with that of Mukteswar strain, resulting the double genes replaced chmeric virus, rL-MuFHN. [Results] Although the rescued chimeric virus (rL-MuHN) did not show significant increase in ability of hemadsorption, the intracerebral pathogenicity index test (ICPI=0) in chickens and mean death time for eggs (MDT≥90 h). rL-MuHN kept the low pathogenicity similar to its parent rLaSota strain. Compared to single gene replaced rL-MuHN, rL-MuFHN induced stronger cell fusion and showed a mild increase in ICPI (from 0 to 0.59) and no significant change in MDT (≥90 h). rL-MuFHN showed much lower pathogenicity than that of Mukteswar (ICPI.=1.32 and MDT=46,respectively). A HN gene exchange alone within the context of the NDV rLaSota backbone failed to increase virus virulence from unvelogenic to mesogenic pathotype. [Conclusion] These results indicated that the virulence of NDV is determined multigenically. The heterotypic HN and F pairs were not equally effective in virus pathogenicity. The HN gene derivated from mesogenic strain dose not alter the lentogenic property of NDV LaSota strain. NDV can be manipulated by gene replacement in the future for use as a vaccine candidate.

Key words:recombinant newcastle disease virus; F protein; HN protein; virulence