HCV genomes are considerable heterogeneities in nucleotide and amino acid sequences among individual isolates. The primary structure of the putative core and NS3 protein regions are relatively conserved among HCV isolates, while those of envelope proteins (E1 and E2) and NS4 are variable. On the basis of nucleotide sequence homology of parts of HCV genomes, several research groups have reported the possible existence of multiple subtypes of HCV isolates. From comparative sequence studies on HCV cDNA clones corresponding to NS3 and NS4 regions followed by diagnostic studies using these clones, have shown that there are at least two groups of HCV, group I and group II. The peptide produced in E. coli, carrying group I and II cDNA clones (A.A. positions 1676-1736 of NS4 region) are recognized by circulating antibodies specific to group I and II HCV. These group specific antigens of NS4 region are highly reliable in detecting (over 90%) circulating antibodies against either groups of HCV. In this study, most of the HCV isolates used were shown to be classified into group I or II HCV based on their amino acid differences and phylogenic analysis within the putative 5'UTR, core and NS3-4 regions. Biological significance of these two groups of HCV is suggested by the observation that the group I HCV was resistance to the IFN therapy (10-20% showing a good response), on the other hand the group II HCV showed a good response (70-90%).