遗传密码调节禽流感病毒对新宿主的适应机制研究

Avian influenza A viruses could cross the barrier and infect human beings. The mechanism of interspecies transmission for avian influenza A virus is a hot and important issue for scientific community, including two aspects whose are virus invasion mechanism and virus effective replication mechanism. Currently most of the researches focus on the viral invasion, phylogenesis and pathogenicity, its mechanism is basically clear. Although virus effective replication in host is a key step for interspecies transmission, yet the mechanism of is still unclear. In present program, based on our previous research results that H7N9 virus get internal genes from H9N2 and enhance the replication ability and the pathogenicity for mammalian host, we will select the three subtypes avian influenza virus (H9N2、H5N1and H3N2) that can cross the barrier and important hosts (chicken, pig and human) as subjects in order to explore change regulation of codon usage bias for genes of virus and host by bioinformatics strategy. Furthermore we will identify characterization of the key codons by analyzing the codon usage bias of virus and host and reconstruct the recombinant virus strains using point mutation technique and reverse genetic technique. Finally, we will clarify the Shinjuku survival and host adaptability mechanism of the virus by verifying the rationality of the adaptability mechanism through cell model and animal model. The research results of the project have important significance for a comprehensive understanding of interspecies transmission of avian influenza virus, which lays the scientific foundation for the design of vaccine and evolutionary trend prediction of novel avian influenza virus in the future.

禽流感病毒跨种间传播机制是当前科学界重要科学问题，主要包括病毒侵入机制和病毒有效复制机制两个方面。当前大部分研究主要集中在病毒侵入机制方面，对病毒有效复制的遗传机制研究不足，病毒对新宿主适应性机制尚不清楚。本项目在前期基础上（H9N2为H7N9提供了内部基因增强了H7N9病毒的复制能力和对人的致病性），对H9N2、H5N1和H3N2等3个突破种间屏障的禽流感病毒亚型和鸡、猪和人类等3个重要宿主的遗传密码子的偏好性分析，揭示不同宿主与相应的亚型病毒基因的密码子偏好性规律，找出宿主特异性的病毒基因中关键的密码子，利用点突变技术和反向遗传技术构建相应的基因重组病毒株，进而利用细胞模型和动物模型进行验证，阐明病毒在新宿主内生存和病毒对宿主的适应性机制。揭示这一机制对于全面了解禽流感病毒跨种间传播具有重要意义，为禽流感病毒疫苗的设计以及病毒未来的进化趋势的预测奠定科学基础。

禽流感病毒跨种间传播机制是当前科学界重要科学问题，主要包括病毒侵入机制和病毒有 效复制机制两个方面。当前大部分研究主要集中在病毒侵入机制方面，对病毒有效复制的遗传 机制研究不足，病毒对新宿主适应性机制尚不清楚。本项目在前期基础上，对H9N2、H5N1和H3N2等3个突破种间 屏障的禽流感病毒亚型和鸡、猪和人类等3个重要宿主的遗传密码子的偏好性分析，发现尽管流感病毒的GC含量均低于AU含量，偏向使用以A或U结尾的密码子；但人源毒株与鸡源毒株的相对聚集，鸭、水鸟、野鸟源毒株聚集，人、鸡源与鸭、水鸟、野鸟源存在分歧。.利用点突变技术和反向遗传技术构建相应的基因重组病毒株，进而利用细胞模型和动物模型进行验证，发现K91R和K198R能够显著降低病毒株在哺乳动物细胞内的聚合酶活性和干扰细胞定位；而K470R显著增强毒株在哺乳细胞中的复制能力和在小鼠中的致病力。在小鼠等哺乳动物中具有更高复制能力和致病能力的毒株在禽类中的复制能力和致病能力存在差异。.本研究结果表明，在分析流感病毒对宿主的可能威胁时，除了要评估毒株是否携带有利于其在该宿主体内复制或致病的关键突变位点，还应考虑密码子使用偏好带来的复制和致病能力的差异，为流感病毒跨种间传播和进化趋势预测提供新的研究思路。