Mistyping of two Slovenian hepatitis C virus subtype 2c isolates as subtype 2b by two 5' noncoding region genotyping methods.

Abstract

During our recent comparison of the reliabilities of four different hepatitis C virus (HCV) genotyping methods (4, 5), two Slovenian HCV subtype 2c isolates which were misclassified as subtype 2b by the two most widely used genotyping methods, second-generation line probe assay (Inno LiPA; Innogenetics, Ghent, Belgium) (7) and restriction fragment length polymorphism (RFLP) (1) analysis, were identified. Namely, although these two 59 noncoding region (59NCR)based genotyping methods clearly identified both isolates as HCV subtype 2b, this subtype could not be confirmed by two core-based genotyping methods and by sequence analysis of the 270-bp fragment of the NS-5 region. Thus, a commercial core-based nested PCR coupled with DNA enzyme immunoassay (GEN-ETI-K-DEIA; Sorin Biomedica, Saluggia, Italy) (8) assigned genotype 2 and excluded subtypes 2a and 2b for both Slovenian isolates, since the genotype 2-specific probe included in the assay hybridized with PCR products and both subtype 2a- and 2b-specific probes did not. By the second core-based genotyping method, Okamoto’s genotype-specific PCR (3), HCV subtype could not be exactly determined in either isolate due to multiple PCR products obtained. Finally, sequence and phylogenetic analysis of the 270-bp fragment of the NS-5 regions, performed as described previously (6), clearly classified both Slovenian isolates as subtype 2c.