Genotyping of hepatitis C virus isolates by a modified polymerase chain reaction assay using type specific primers: Epidemiological applications

Abstract

A polymerase chain reaction (PCR)‐based assay using primers against the hepatitis C core gene has been described [Okamoto et al. (1992a): Journal of General Virology 73:673–679]. Within the two major HCV genotypes 1 and 2, the Okamoto system identifies two subtypes each (1a,1 b and 2a, 2b, respectively). Typing is achieved by a primary PCR with consensus primers followed by a nested PCR with type specific primers. The original assay was modified by addition of a parallel second PCR identifying the recently described major genotype 3. The assay also identifies in duplicate subtype 1 b (type II by Okamoto), suggested to respond poorly to interferon. Reaction conditions were reviewed and melting temperatures of all typing primers equalised to increase stringency. The modified system functioned well and typing results were supported by partial core sequencing. The following distribution of genotypes was found in 53 hepatitis C virus (HCV) infected Swedish blood donors: genotype 1a (57%), 3 (19%), 1b (13%), and 2b (11%). In six recipients of HCV infected blood identified in a retrospective study, the recipient HCV genotype was identical to donor HCV genotype. Furthermore, in HCV positive couples identical genotype was observed when only one partner had an external risk factor; whereas genotypes were often diverse if both sex partners had parenteral risk factors. Finally, a cluster of hepatitis C cases in a haemodialysis unit was evaluated retrospectively. Eight patients had genotype 1b, two had mixed 1a and 1b, and one had type 1a. The modified HCV genotyping assay was of value in examining different epidemiological situations and can be expanded presumably to include future genotypes.