The object of this proposal is to apply molecular techniques to study the biology and progression of Woodchuck Hepatitis Virus (WHV) infection. Of the three animal viruses that are related to human Hepatitis B Virus (HBV), WHV is the only one that is known to cause hepatocellular carcinoma as HBV does. This study will define the environment in which viral hepatocarcinogenesis takes place, and it will provide baseline data required to develop WHV as an experimental model for treatment of HBV infection. To make serological assays for WHV analogous to those used to study HBV in Man the surface, core and e antigens of WHV will be purified from appropriate tissues of infected animals and used to immunise rabbits. The antigens and antibodies so obtained will be used to set up enzyme immunoassays. WHV-specific RNA species present in infected liver will be mapped in detail by methods based on primer extension, and the transcripts coding for the surface and core antigens will be identified by in vitro translation. The ends of the virion DNA strands will be precisely located relative to the known nucleotide sequence of the genome. The progression of acute and chronic WHV infections will be studied by serological sampling and by analysis of nucleic acids obtained by repeated liver biopsy of infected animals. RNA transcripts will be analysed by """"""""Northern"""""""" blot hybridisation, and DNA intermediates of replication by Southern blot, while covalently closed circular and integrated viral DNA will be analysed likewise after separation by BND-cellulose chromatography. It is hoped that this method of study will reveal the molecular changes that underlie the shift during chronic infection from production of infectious virus to expression of surface antigen on its own, as well as providing information about the resolution of chronic infection.
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