title: Development and evaluation of an automated hepatitis C virus NS5B sequence-based subtyping assay.
abstract: BACKGROUND
  Hepatitis C virus (HCV) genotyping and accurate subtype determination is becoming increasingly important to better understand viral evolution, the development of resistance to STAT-C, and possibly even for the treatment and management of chronic HCV-infected patients.
  
  
  METHODS
  A subtyping assay based on a 329-bp sequence of the NS5B region, together with an automated subtype interpretation tool was developed. Clinical samples of the six major genotypes were used to assess assay performance characteristics.
  
  
  RESULTS
  The NS5B BLAST-based subtyping assay showed clinical sensitivity for amplification of 89% (n=603 random samples). Assessment of analytical sensitivity of amplification for genotypes 1, 2, 3 and 4 revealed a suitable performance for high viral load samples and decreased only with low viral loads. The results were 100% and 99% accurate at the genotype and subtype level, respectively. A concordance of 97% on genotype level and 62% on subtype level was observed by comparison with subtype results from 5' non-coding-based assays with a panel of 276 isolates.
  
  
  CONCLUSIONS
  The HCV NS5B subtyping assay has been validated for research use. Based on its performance, it is the method of choice in cases where subtype rather than genotype information is needed.
authors.affiliation: Virco BVBA, Mechelen, Belgium. dkoletzk@its.jnj.com
authors.forename: Diana
authors.initials: D
authors.lastname: Koletzki
authors.forename: Stéphanie
authors.initials: S
authors.lastname: Dumont
authors.forename: Hans
authors.initials: H
authors.lastname: Vermeiren
authors.forename: Bart
authors.initials: B
authors.lastname: Fevery
authors.forename: Pieter
authors.initials: P
authors.lastname: De Smet
authors.forename: Lieven J
authors.initials: LJ
authors.lastname: Stuyver
mesh_terms: D001331:Automation; D005838:Genotype; D016174:Hepacivirus / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D006801:Humans; D010802:Phylogeny; D021122:Protein Subunits / Q000235:genetics*; D012367:RNA, Viral / Q000235:genetics; D011933:Reagent Kits, Diagnostic; D017423:Sequence Analysis, RNA; D019562:Viral Load; D017361:Viral Nonstructural Proteins / Q000235:genetics*; D012324:RNA-Dependent RNA Polymerase
chemical_list: D021122:Protein Subunits; D012367:RNA, Viral; D011933:Reagent Kits, Diagnostic; D017361:Viral Nonstructural Proteins; C087036:NS-5 protein, hepatitis C virus; D012324:RNA-Dependent RNA Polymerase