SEQUENCE-ANALYSIS OF HEPATITIS-C VIRUS VARIANTS PRODUCING DISCREPANT RESULTS WITH 2 DIFFERENT GENOTYPING ASSAYS

Methods for identifying the genotype of hepatitis C virus (HCV) in cli nical specimens are frequently based upon the direct characterisation of viral RNA sequences by polymerase chain reaction (PCR) amplificatio n, or by serologically based methods, in which the infecting genotype is inferred from the pattern of antibody reactivity to type-specific p eptides or recombinant proteins used as antigens in an Enzyme Linked I mmunosorbent Assay (ELISA). Although genotyping by direct, PCR-based m ethods show generally highly concordant results with the genotype infe rred from serological typing assays (>95% agreement), there exist a sm all number of samples that produce discrepant results. To investigate the underlying reasons for the discrepancies, we obtained eleven sampl es from haemophiliacs and four samples from patients with chronic hepa titis C that produced discordant results between a PCR based assay (In noLipa I and Il)and a serotyping assay (Murex HCO2). Nucleotide sequen ces in the 5'noncoding region (5'NCR), core, and NS4 region were used to identify the genotype of the circulating virus and to identify amin o acid changes in NS4 that might alter antigenicity. In 14 samples, se quence analysis of all three regions was concordant with the results o f the InnoLipa assay. There were few if any amino acid substitutions i n NS4 that might have accounted for the discrepant serotyping results, which were found predominantly in samples from individuals with a his tory of multiple exposure to HCV. It remains unclear whether the detec tion of antibody in such discrepant samples corresponds to previous ex pression of a different genotype than detected by PCR, or whether the virus population in plasma is more restricted in genotype diversity th an the population in the liver or at other sites of viral replication. (C) 1997 Wiley-Liss, Inc.