Objective To investigate the presentation of a neutralization epitope-containing peptide antigen of hepatitis E virus （HEV） on chimeric virus-like particles （VLPs） of hepatitis B surface antigen （HBsAg）. MethodsThe gene fragment corresponding to amino acids （aa） 551-607 （HEnAg） of HEV capsid protein

which contains the only neutralization epitope identified to date

was fused via a synthetic glycine linker in frame with the gene of HBsAg. The resulted fusion gene was then integrated through transformation into the genome of Pichiapastorisunder the control of a methanol-induced alcohol oxidase 1 （AOX1） promoter and expressed intracellularly. The expression products in the soluble cell extracts were characterized by Western blot

ELISA

CsCl density gradient analysis

and electron microscopic visualiza-tion. Results The novel fusion protein incorporating HBsAg and the neutralization epitope-containing HEnAg was expressed successfully in Pichiapastoriswith an expected molecular weight of approximately 32 kD. It was found to possess the ability to assemble into chimeric HBV/HEV VLPs with immunological

physical and morphological characteristics akin to HBsAg particles. Not only did the chimeric VLPs show high activity levels in a HBsAg particle-specific ELISA but they were also strongly immunoreactive with hepatitis E （HE） positive human serum in a HEV specific ELISA

indicating that HEnAg peptide fragments were exposed on VLP surfaces and would be expected to be readily accessible by cells and molecules of the immune system. Similarity between chimeric VLPs to highly immunogenic HBsAg particles may confer good immuno-genicity on surface-displayed HEnAg. Conclusion The chimeric HBV/HEV VLPs produced in this study may have potential to be a recombinant HBV/HEV bivalent vaccine candidate.