华东地区首例人感染2.3.4.4b分支H5N1禽流感病毒毒株分子特征分析

Molecular characteristics of the first human case infected with the avian influenza virus H5N1 clade 2.3.4.4b in eastern China

目的分析华东地区首例感染人的2.3.4.4b H5N1禽流感病毒分子进化基因特性。方法通过实时RT-PCR、病毒培养和序列分析,对患者痰标本和支气管肺泡灌洗液进行病毒培养和鉴定。利用Mega 6.1等软件构建系统发育树以探索病毒的进化和起源。结果从1例患严重呼吸道症状患者的支气管肺泡灌洗液标本中分离到1株甲型流感病毒,属于甲型H5N1禽流感2.3.4.4b分支。对该毒株进行了全基因组测序分析,8个基因组片段与2021年以来在欧亚大陆特别是中国南方流行的H5N1分离株具有极高的核苷酸序列相似性(99.1%~99.7%)。该病毒HA蛋白的226~228位氨基酸未发生突变,保留了禽类受体α2-3半乳糖苷唾液酸(SAα2-3Gal)结合特性,为高致病性禽流感H5N1病毒;其NA和M2蛋白中未发现与抗病毒药物(包括奥司他韦、扎那米韦和金刚烷)耐药性相关的突变;JS210的PB2蛋白的E627K和D701N位点以及NS1蛋白的D92E位点均未发生突变,表明该毒株对人体的适应力有限。结论分支2.3.4.4b H5N1禽流感病毒可以溢出族群感染人类并引起严重的疾病,应持续加强对禽流感病毒的监测。

Objective To investigate the molecular genetic characteristics of the first human case infected with the avian influenza virus H5N1 clade 2.3.4.4b in eastern China.Methods Patient′s sputum and bronchoalveolar lavage fluid(BALF)specimens were collected for virus identification using real-time reverse-transcription PCR(RT-PCR)assay,virus culture and sequencing.The virus evolution and origin were investigated using phylogenetic analysiswith the software Mega 6.1.Results An influenza A virus isolate was obtained from the BALF specimens of a case with severe respiratory syndromes,which belonged to influenza virus H5N1 clade 2.3.4.4b.Whole-genome sequencing of the virus isolate showed that the 8 genome fragment shared high nucleotide sequence homology(99.1%to 99.7%)with H5N1 isolates that had been prevalent in Eurasia since 2021,notably in southern China.No mutation was identified in 226 to 228 amino acids in the HA protein of the virus,and the binding ability of avian receptor alpha-2,3 galactoside sialic acid(SAα2,3Gal)was reserved,which was characterized as highly pathogenic avian influenza H5N1 virus.No mutationassociated with resistance to oseltamivir,zanamivir or adamantane was detected in NA or M2 protein,and no mutations were identified in E627K or D701N sites of JS210 or PB2 protein,or D92E site of NS1 protein,indicating limited adaptability of this virus isolate to humans.Conclusions The avian influenza virus H5N1 clade 2.3.4.4b may spillover avian to humans and cause severe diseases.Persistent intensified surveillance of avian influenza viruses is required.