Analysis of virus adaptations in the novel HCV cell culture and infection system

我们用从一例重型肝炎患者中分离的JFH-1株建立了丙型肝炎病毒的培养体系。这个系统使我们能够详细分析丙型肝炎病毒的生命周期。在这个研究项目中，我们分析了丙型肝炎病毒如何在培养的细胞和生物体中适应。我们还试图确定哪些病毒成分参与了丙型肝炎病毒的适应。培养细胞中的丙型肝炎适应：我们将体外合成的丙型肝炎病毒RNA导入Huh7细胞，并收集具有感染性的丙型肝炎病毒颗粒。丙型肝炎病毒感染有时会导致斑块形成。收集形成丙型肝炎病毒的空斑，感染未成熟的HuH7细胞。从单个斑块中分离的病毒亚克隆显示出比亲本病毒更高的复制效率和细胞致病性。我们分析了该丙型肝炎病毒的序列，发现在NS5B编码区有几个氨基酸替换的核苷酸突变。这些突变可能与空斑形成能力和病毒基因组复制有关。

We have established HCV culture system using JFH-1 strain isolated from a fulminant hepatitis patient. This system enabled us to analyze HCV life cycles in detail. In this research project, we have analyzed how HCV adapted in cultured cells and organisms. We have also tried to determine which virus components involved in HCV adaptation.HCV adaptation in cultured cells: we transfected in vitro-synthesized HCV RNA into Huh7 cells and collected infectious HCV particles. HCV infection sometimes causes plaque formation. We collected plaque forming HCVs, and infected to naive Huh7 cells. Isolated virus subclones from individual plaque showed greater replication efficiency and cytopathogenicity than parental virus. We analyzed the sequence of this HCV and found several nucleotide mutations with amino acid substitution in NS5B coding region. These mutations may be involved in plaque forming ability and virus genome replication.