BACKGROUND: Steatosis is a common histological finding and a poor prognostic indicator in patients with hepatitis C virus (HCV) infection. In HCV genotype 3-infected patients, the etiology of steatosis appears to be closely correlated with unknown viral factors that increase intracellular lipid levels. We hypothesize that specific sequence polymorphisms in HCV genotype 3 core protein may be associated with hepatic intracellular lipid accumulation. METHODS: Using selected serum samples from 8 HCV genotype 3-infected patients with or without steatosis, we sequenced the HCV core gene to identify candidate polymorphisms associated with increased intracellular lipid levels. RESULTS: Two polymorphisms at positions 182 and 186 of the core protein correlated with the presence (P= .03) and absence (P= .005) of intrahepatic steatosis. Transfected liver cell lines expressing core protein with steatosis-associated polymorphisms had increased intracellular lipid levels compared with non-steatosis-associated core isolates, as measured by oil red O staining (P= .02). Site-specific mutagenesis performed at positions 182 and 186 in steatosis-associated core genes yielded proteins that had decreased intracellular lipid levels in transfected cells (P= .03). CONCLUSIONS: We have identified polymorphisms in HCV core protein genotype 3 that produce increased intracellular lipid levels and thus may play a significant role in lipid metabolism or trafficking, contributing to steatosis.