禽冠状病毒纤突蛋白在病毒跨种感染与致病进程中的分子作用机制研究

The cross-species transmission of viruses such as SARS-CoV caused disease outbreaks. the phenomenon of cross-species infection of animal pathogens is increasingly arousing people's vigilance. The in-depth exploration of its mechanism has attracted the attention of researchers. Recent studies have found that avian coronaviruses from different hosts can replicate in different kinds of birds including poultry. Therefore, these viruses maybe play a very important role in the evolution of IBV, and the reasonable methods must be taken to block such cross-species. But, the the basic theory about the transmission and infection still need to be clarified. As we all know, the binding and fusion of viruses to host cells is the first step in the determination of viral tissue tropism, host specificity, and pathogenicity. This project serve the knowledge as an entry point and based on the recent finding, bird coronavirus isolates isolated from common pheasant caused significant differences in infection between SPF chickens and common pheasants. It is presumed that the cross-species infection and pathogenic mechanism of the virus strains may be related to the binding and recognition of the virus to different target animal cell receptors, and may also be related to the fusion process of virus into the target animal cell membranes. So, this project is based on the previous discovery, using the avian coronavirus spike protein as the main goal to systematically study its mechanism of action in the cross-species infection and pathogenicity. The research results will be useful in provide new ideas about trying to study the genetic evolution of avian coronavirus pathogens and disease prevention and control.

随着SARS-CoV等跨种感染并造成重大疫情，动物病原跨种感染现象越来越引起人们的警惕，对其机制的深入探索得到研究者的重视。近年来的研究发现来源不同的禽类冠状病毒能够在包括家禽的不同禽体内复制，因而该类病毒在IBV的演化进程中扮演着非常重要的角色，要想采取合理措施来阻断这类跨种感染，尚需明确其中的机制，但目前仍缺乏基础理论支持。众所周知，病毒与宿主细胞的结合与融合是病毒组织嗜性、宿主特异性以及致病性的第一步决定因素，本项目以此为切入点，并基于近期分离获得雉鸡源禽冠状病毒对SPF鸡及雉鸡感染特点存在的显著差异，推测毒株跨种感染和致病机制可能与病毒株对不同靶动物细胞受体的结合和识别有关，也可能与病毒进入靶动物细胞膜的融合过程有关，本项目拟基于前期发现，以禽冠状病毒纤突蛋白为主要目标通过系统研究其在病毒跨种间感染与致病进程中的作用机制，试图为该类病原遗传演化研究及基于该机制的免疫防控提供新思路。

随着近年来冠状病毒感染并造成重大疫情，病原跨种感染现象越来越引起人们的警惕，对其机制的深入探索得到重视。此前研究发现来源不同的禽类冠状病毒能够在包括家禽在内的不同禽体内复制，推测这些病毒在IBV的演化进程中扮演着非常重要的角色，采取合理措施阻断这类跨种感染尤为必要，尚需明确其中的机制，但目前仍缺乏基础理论支持。本项目以基于前期分离获得雉鸡源禽冠状病毒对SPF鸡及雉鸡感染特点存在显著差异，推测毒株跨种感染和致病机制可能与病毒株对不同靶动物细胞受体的结合和识别有关，也可能与病毒进入靶动物细胞膜的融合过程有关，本项目拟基于前期发现，建立了雉鸡源和鸡源感染特性不同的禽冠状病毒跨种感染模型，在此基础上通过不同宿主来源毒株在体内跨种交互感染试验验证以及毒株跨种适应性传代感染，验证毒株跨宿主感染后毒株的复制和组织嗜性等特点，结合不同源病毒在跨种适应中纤突蛋白受体结合域及其他主要蛋白氨基酸位点变化，探究了鸡源和雉鸡源冠状病毒跨种感染特性的关键分子变化特征。结果表明，与不同株IBV表现出不同的特性相似，I0623/17和I0710/17与M41在发病机制、趋向性和受体特异性上存在差异。我们发现PhCoV I0623/17和M41在S1亚基最小RBD的关键氨基酸上存在明显差异，相比之下，在两种禽冠状病毒中，S蛋白中影响病毒与唾液酸糖苷结合的5个特异性潜在N-糖基化位点是保守的，雉鸡体内PhCoV I0623/17的受体可能对IBV M41和H120株的亲和性较低，这解释了IBV M41和H120株在雉鸡体内复制能力较差的原因，同样地，鸡体内IBV M41和H120株的受体可能对PhCoV I0623/17的亲和性较低，也解释了IBV M41和H120株在雉鸡体内复制能力较差的原因。我们以表达不同毒株受体结合域在体外验证了对靶细胞结合活性，进一步证实S1亚基这种可变性很可能决定了禽冠状病毒的宿主和组织嗜性。