人源H5N1亚型禽流感病毒株空斑特性和致病性研究

Research on the plaque forming features and pathogenicity of H5N1 avian influenza virus strains isolated from human

目的观察不同H5N1亚型禽流感病毒株空斑特性,对比研究不同空斑类型H5N1病毒致病性和复制能力差异。方法将不同稀释度的H5N1亚型禽流感病毒株分别接种于MDCK单层细胞,加盖营养琼脂,合适时间染色,观察不同H5N1亚型禽流感病毒株空斑形成特点,按照DullbeccoR的方法计算病毒PFU数;根据空斑大小进行挑选、纯化、筛选不同类型空斑病毒,测定这些病毒对Balb/c小鼠致病性差异和在MDCK细胞上复制差异。结果野生的人源H5N1禽流感病毒株在空斑形成上都存在不同程度的不均一性,空斑大小、形状差异明显:A/Vietnam/1194/2004(H5N1)以大圆形斑为主,夹杂少量小点状斑。A/Beijing/01/03(H5N1)大多数为中等大小空斑,并有少数针尖状斑。野生A/Vietnam/1194/2004经过分离纯化后,得到的两种空斑类型病毒在致病性和复制能力存在明显差异。结论野生的人源H5N1禽流感病毒株在空斑形成上都存在不同程度的不均一性,从野生病毒株中分离、纯化的大小斑病毒在致病性和复制能力上存在明显差异。

Objective To observe the plaque forming characteristics of different H5N1 avian influenza virus strains.And contrast the differences of pathogenicity and replication of various virus strains with different plaque forming characterisfics.Methods Different H5N1 avian influenza virus strains in serial dilutions were inoculated to MDCK cells. Then the cells were covered with nutritious agar, and dyed some day after being infected with viruses. The plaques of different H5N1 virus strains were observed and titrated by Dullbeeco R method. Then the pathogenicity to Balb/c mice and replication in MDCK cells of virus strains with different plaque forming characteristic were determined. Results It was heterogeneous in/plaque forming for wild H5N1 avian influenza virus strains isolated from human. Plaques of these virus strains were obviously different in size and shape. The plaques of A/Vietnam/1194/2004 （H5N1） were mainly large in size and round in shape, except for a few smaller plaques appearing punctiform in shape. While A/Beijing/01/03（H5N1） were dominated by plaques of moderate size, also mixed with a few punctiform plaques, Two kinds of virus strains with different plaque forming characteristic were got from A/Vietnam/1194/2004 （H5N1） after purification and isolation with plaque forming assay. There were obvious differences in pathogenicity and replication between those two virus strains. Conclusion It was heterogeneous in plaque forming for wild H5N1 avian influenza virus strains isolated from human. Plaques of these virus strains were obviously different in size and shape, and it was heterogeneous for virus strains purified from wild virus strain in pathogenicity and replication.