Determination of the hepatitis C virus subtype: comparison of sequencing and reverse hybridization assays.
BACKGROUND
Hepatitis C virus (HCV) genotyping and accurate subtyping is becoming increasingly relevant to epidemiological studies, clinical management, pathogenicity, and vaccine development.


METHODS
The TRUGENE HCV 5'NC Genotyping Kit, the new VERSANT HCV Genotype 2.0 Assay (LiPA), and a new laboratory-developed HCV NS5b sequencing assay designed for automated sequencing of the HCV NS5b region were used. Clinical samples and a molecular diagnostics HCV genotyping proficiency program panel were used to determine accuracy and differentiate performance characteristics of the three methods.


RESULTS
All amplified samples from among the members of a HCV genotyping proficiency program panel that contained a single HCV genotype were subtyped correctly using all three HCV genotyping assays. With the TRUGENE HCV 5'NC Genotyping Kit, the HCV subtype was determined in 357 of 441 of routine clinical samples. When the 84 samples with only genotype results were retested with the VERSANT HCV Genotype 2.0 Assay (LiPA), 61 could be further subtyped accurately. With the new laboratory-developed HCV NS5b sequencing assay, all 84 could be subtyped accurately.


CONCLUSIONS
The two new methods show advantages over the routinely used TRUGENE HCV 5'NC Genotyping Kit in terms of genotyping and subtyping accuracy by utilizing part of the HCV core region and NS5b region, respectively.
Stelzl
Evelyn
E
Molecular Diagnostics Laboratory, Institute of Hygiene, Medical University of Graz, Graz, Austria.
van der Meer
Carola
C
Gouw
Remko
R
Beld
Marcel
M
Grahovac
Maja
M
Marth
Egon
E
Kessler
Harald H
HH
D005838:Genotype; D016174:Hepacivirus / Q000302:isolation & purification*; D009693:Nucleic Acid Hybridization / Q000379:methods*; D011933:Reagent Kits, Diagnostic / Q000592:standards; D017422:Sequence Analysis, DNA / Q000379:methods*; D017361:Viral Nonstructural Proteins / Q000235:genetics; D012324:RNA-Dependent RNA Polymerase
D011933:Reagent Kits, Diagnostic; D017361:Viral Nonstructural Proteins; C087036:NS-5 protein, hepatitis C virus; D012324:RNA-Dependent RNA Polymerase