2017–2019年我国南方地区高致病性H5N6亚型禽流感病毒血凝素蛋白分子特征分析

Molecular characteristic of the hemagglutinin of highly pathogenic avian influenza A H5N6 viruses in live-poultry market, 2017–2019, Southern China

摘要:

【背景】自2014年以来，H5N6禽流感病毒在我国家禽和活禽市场持续进化，成为人类和动物健康的重大威胁。【目的】对2017–2019年中国南方地区93株高致病性H5N6禽流感病毒的HA基因进行分子进化分析。【方法】接种9–11日龄鸡胚分离核酸检测阳性的H5N6标本，运用下一代测序平台对病毒分离物进行全基因组测序，从NCBI和GISAID数据库下载参考序列，利用Blast、Mega 6.1及ClustalX等软件进行序列分析。【结果】2017–2019年，从189份江苏省H5亚型禽类/环境标本和1名H5N6患者咽拭子标本中共分离到43株病毒，完成了33株H5N6病毒的全基因组测序。下载网上同时期中国其他地区流行的H5N6毒株序列，对总计93株H5N6病毒的HA基因进行分子进化分析。93株H5N6病毒中有78株属于Clade 2.3.4.4h，9株病毒属于Clade 2.3.4.4e，4株H5N6病毒属于Clade 2.3.4.4b，1株属于Clade 2.3.4.4f，1株属于Clade 2.3.4.4g。所有93株病毒HA蛋白的裂解位点含有多个碱性氨基酸，表明它们都属于高致病性禽流感病毒。所有93株病毒HA蛋白的Q222和G224位氨基酸没有发生突变，保留了禽类受体α2-3半乳糖苷唾液酸(SAα2-3Gal)结合特性；158位点丧失糖基化，同时124位出现一个新的潜在糖基化位点。【结论】2017–2019年间中国南方地区H5N6病毒进化活跃，具有明显的基因多样性，需要加强对病毒分子进化的监测。

Abstract:

[Background] Since 2014, H5N6 avian influenza virus has continued to evolve in domestic poultry and live-poultry markets, becoming a major threat to human and animal health. [Objective] To analyze the molecular characteristic of hemagglutinin (HA) and neuraminidase (NA) proteins of highly pathogenic avian influenza H5N6 viruses from 19 human cases. [Methods] Whole-genome sequences of original specimen and virus isolates were obtained by next-generation sequencing technology. Pairwise sequence alignments and phylogenetic analysis were performed by Blast, Mega 6.1 and ClustalX softwares. [Results] In 2017–2019, 43 viruses were isolated from 189 H5 subtype poultry/environmental samples and 1 H5N6 patient throat swab samples, and 33 H5N6 viruses were sequenced. Based on the sequence of H5N6 virus collected on the internet, the HA gene of 93 H5N6 viruses was analyzed. Among 93 H5N6 strains, 78 belong to 2.3.4.4h, 9 belong to 2.3.4.4e, 4 belong to 2.3.4.4b, 1 belong to 2.3.4.4f, 1 belong to 2.3.4.4g. The cleavage sites of HA proteins of all 93 strains contain several basic amino acids, indicating that they are highly pathogenic avian influenza viruses. Q222 and G224 amino acids of HA protein of all 93 viruses did not mutate, which retained the binding characteristics of avian receptor α2-3 galactoside sialic acid (SAα2-3Gal); 158 sites lost glycosylation, and 124 sites showed a new potential glycosylation site. [Conclusion] From 2017 to 2019, the evolution of H5N6 virus infection in China is active, and there is obvious genetic diversity. It is necessary to strengthen the monitoring of the evolution of H5N6 viruses in China.