title: Sequencing assays for failed genotyping with the versant hepatitis C virus genotype assay (LiPA), version 2.0.
abstract: For optimal antiviral therapy, the hepatitis C virus (HCV) genotype needs to be determined, as it remains a strong predictor of sustained viral response. In this study, we assessed the number of HCV genotyping results that could not be determined using the commercially available line probe assay (LiPA) (Versant hepatitis C virus genotype 2.0 assay) in a large international panel of samples from 9,874 HCV-positive patients. In-house sequencing assays targeting the 5' untranslated region (UTR), core region, NS3 region, and NS5B region of the HCV genome and phylogenetic analyses were used to resolve these LiPA failures. Among all cases, the genotypes of 51 samples (0.52%) could not be determined with the LiPA. These undetermined results were observed more frequently among samples from non-European regions (mainly the Arabian Peninsula). The use of sequencing assays coupled with phylogenetic analysis provided reliable genotype results for 86% of the LiPA failures, which exhibited higher rates of genotypes 4, 5, and 6 than did LiPA-resolved genotypes. As expected, the 5' UTR was not sufficiently variable for clear discrimination between genotypes 1 and 6, but it also resulted in errors in classification of some genotype 3 and 4 cases using well-known Web-based BLAST programs. This study demonstrates the low frequency of genotyping failures with the Versant hepatitis C virus genotype 2.0 assay (LiPA) and also underlines the need for a complex combination of sequences and phylogenetic analyses in order to genotype these particular HCV strains correctly.
authors.affiliation: Laboratoire de Virologie, Département des Agents Infectieux, Pôle Biologie, Centre Hospitalier Universitaire Grenoble, Grenoble, France. slarrat@chu-grenoble.fr
authors.forename: Sylvie
authors.initials: S
authors.lastname: Larrat
authors.forename: Jean-Dominique
authors.initials: JD
authors.lastname: Poveda
authors.forename: Camille
authors.initials: C
authors.lastname: Coudret
authors.forename: Katia
authors.initials: K
authors.lastname: Fusillier
authors.forename: Nelly
authors.initials: N
authors.lastname: Magnat
authors.forename: Anne
authors.initials: A
authors.lastname: Signori-Schmuck
authors.forename: Vincent
authors.initials: V
authors.lastname: Thibault
authors.forename: Patrice
authors.initials: P
authors.lastname: Morand
mesh_terms: D000328:Adult; D000368:Aged; D001483:Base Sequence; D005188:False Negative Reactions; D005260:Female; D014644:Genetic Variation; D005838:Genotype; D016174:Hepacivirus / Q000145:classification* / Q000235:genetics*; D006526:Hepatitis C / Q000175:diagnosis / Q000821:virology; D006801:Humans; D008297:Male; D008875:Middle Aged; D025202:Molecular Diagnostic Techniques / Q000379:methods; D008969:Molecular Sequence Data; D009154:Mutation; D012367:RNA, Viral / Q000235:genetics*; D017422:Sequence Analysis, DNA; D014773:Virology / Q000379:methods
chemical_list: D012367:RNA, Viral