Scientific Publications by FDA Staff

Arch Virol 2004 Apr;149(4):759-72

Rescue of Hepatitis A virus from cDNA-transfected but not virion RNA-transfected mouse Ltk- cells.

Lu JH, Dveksler G, Kaplan GG

Kaplan GG, 8800 Rockville Pike,Bldg 29-NIH,Rm 225,HFM-325, Bethesda, MD 20892 USA US FDA, Ctr Biol Evaluat & Res, Hepatatis Lab, Bethesda, MD USA Uniformed Serv Univ Hlth Sci, Dept Pathol, Bethesda, MD 20814 USA

Hepatitis A virus (HAV) has stringent replication requirements and a restricted host-range. Mouse Ltk- cells do not support growth of HAV upon infection or transfection of virion RNA. However, low levels of HAV were rescued from Ltk- cells transiently transfected with its infectious cDNA. Ltk- stable transfectants that expressed HAV antigens and produced infectious HAV were selected and termed Ltk-pJH15 cells. After a few serial passages, HAV became undetectable in the Ltk-pJH15 cells. Multiple rounds of single cell cloning of HAV antigen positive Ltk-pJH15 cells resulted in the isolation of clone E8 that produced higher levels of HAV for at least 5 passages. HAV produced in E8 cells was similar to the parental virus as shown by infectivity assays. Luciferase assays using a bi-cistronic construct containing the HAV 5' noncoding region showed similar levels of HAV IRES-dependent translation in Ltk- and Ltk-pJH15 cells, which suggested that HAV IRES-dependent translation was not a limiting factor for HAV growth in these cells. The availability of the Ltk-pHJ15 cells will allow the identification of cellular factors required for HAV growth, which could lead to the development of a mouse model to study pathogenesis of HAV.