COMPARATIVE-EVALUATION OF 4 GENOTYPING METHODS FOR HEPATITIS-C VIRUS

Four most widely accepted genotyping methods for hepatitis C virus (HC V) were applied to 40 HCV RNA isolates obtained from Slovenian patient s in order to determine the concordance and applicability of various g enotyping systems. The four methods are: (i) amplification of the core region with genotype-specific primers; (ii) nested polymerase chain r eaction (PCR) in the core region followed by hybridization to HCV type -specific probes; (iii) reverse hybridization with the line probe assa y Inno LiPA (Innogenetics, Gent, Belgium) using type-specific probes f or the 5' non-coding region (NCR); and (iv) restriction fragment lengt h polymorphism analysis of DNA amplified from the 5' NCR. Additionally , in isolates with discordant results nucleotide sequence analysis of a part of the NS-5 region was performed. Both genotyping methods based on the analysis of the 5' NCR were found more sensitive than those me thods based on the analysis of the HCV core region. None of the four g enotyping methods correctly classified all Slovenian HCV RNA isolates. PCR with genotype-specific primers was identified as entirely unsuita ble for genotyping of Slovenian HCV RNA isolates. The remaining genoty ping methods could clearly differentiate between HCV genotypes, but we re not entirely reliable for HCV subtyping. The specificity of genotyp ing methods, which are based on the 5' NCR or the core region, was occ asionally hampered, due to a lack or excess of sequence variation in t heir respective target regions.