title: [Cell culture system for hepatitis E virus].
abstract: Early studies reported propagation of hepatitis E virus (HEV) in primary hepatocytes or several established cell lines, but replication was inefficient. Recently, using inocula comprised of fecal suspensions with high loads of HEV, originally obtained from Japanese patients who contracted domestic infection of genotype 3 HEV (the JE03-1760F strain, 2.0 x 10(7) copies/ml) or genotype 4 HEV (the HE-JF5/15F strain, 1.3 x 10(7) copies/ml), we developed an efficient cell culture system for HEV in PLC/PRF/5 and A549 cells, which yielded the highest HEV load of 10(8) copies/ml in the culture supernatant, and we successfully propagated six or more generations in serial passages of culture supernatant. In addition, we constructed a full-length infectious cDNA clone (pJE03-1760F/wt) of the JE03-1760F strain, which can replicate efficiently in PLC/PRF/5 and A549 cells. Using a derivative ORF3-deficient (delta ORF3) mutant, we demonstrated that the ORF3 protein of HEV is responsible for virion egress from infected cells and is present on the surface of released HEV particles, which is associated with lipids. Various HEV strains in blood circulation were also propagated efficiently in PLC/PRF/5 and A549 cells. Our in vitro cell culture system can be used for propagation of a wide variety of HEV strains in feces and sera from various infected patients, allowing extended studies on viral replication specific to different HEV strains.
authors.affiliation: Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, 3311-1 Yakushiji, Shimotsuke-shi, Tochigi-ken 329-0498, Japan. hokamoto@jichi.ac.jp
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Okamoto
mesh_terms: D000818:Animals; D005838:Genotype; D016751:Hepatitis E / Q000821:virology; D016752:Hepatitis E virus / Q000235:genetics / Q000502:physiology*; D006801:Humans; D008113:Liver Neoplasms / Q000821:virology; D008175:Lung Neoplasms / Q000821:virology; D009154:Mutation; D012367:RNA, Viral; D014407:Tumor Cells, Cultured; D014764:Viral Proteins / Q000502:physiology; D014773:Virology / Q000379:methods*; D014779:Virus Replication
chemical_list: C400973:ORF3 protein, Hepatitis E virus; D012367:RNA, Viral; D014764:Viral Proteins