Towards a better resolution of hepatitis C virus variants: CLIP sequencing of an HCV core fragment and automated assignment of genotypes and subtypes.
Commercially available assays for typing of hepatitis C virus (HCV) isolates satisfy the current clinical needs. They are, however, limited in their ability to identify the multitude of existing HCV subtypes correctly. Therefore, these kits should only be used cautiously in epidemiological studies and will also not meet future clinical demands which might arise, e.g., from the application of HCV subtype-specific antiviral compounds. In an attempt to overcome the drawbacks of commercial typing procedures based on the analysis of the 5' untranslated region (5' UTR), an approach was developed which relies on CLIP sequencing of an HCV core fragment with automated assignments of types and subtypes via an originally created "core-specific" sequence database. The performance characteristics of the new technique were evaluated in comparison to the Trugene 5' NC Genotyping Kit. The core-based sequencing method could regularly detect HCV isolates of genotypes 1-6 with an analytical sensitivity of 5000 IU/ml. The accuracy of typing results obtained by the Trugene test was 97% (genotypes) and 81% (subtypes). The core-linked approach classified all HCV strains correctly on the level of genotypes and led to an adequate subtype assignment in 96% of all cases. This analytical performance characteristics recorded for the newly devised typing technique was superior to those reported for all commercially available assays, including a most recently released new generation of the line probe assay. Consequently, CLIP sequencing of an HCV core fragment with subsequent automated assignment of types and subtypes can be confidently used in clinical laboratory practice to answer current and also future questions in the context of HCV typing.
Institute of Virology, National Reference Centre for Hepatitis C, Essen University Hospital, University of Duisburg-Essen, Essen, Germany. stefan.ross@uni-due.de <stefan.ross@uni-due.de>
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Ross
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Viazov
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Wolters
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Roggendorf
D020121:5' Untranslated Regions / Q000235:genetics; D000293:Adolescent; D000328:Adult; D000368:Aged; D001331:Automation; D002648:Child; D030561:Databases, Nucleic Acid; D005260:Female; D005838:Genotype; D016174:Hepacivirus / Q000145:classification* / Q000235:genetics* / Q000302:isolation & purification; D006526:Hepatitis C / Q000821:virology*; D006801:Humans; D008297:Male; D008875:Middle Aged; D010802:Phylogeny; D011110:Polymorphism, Genetic; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA / Q000379:methods*; D014758:Viral Core Proteins / Q000235:genetics*
D020121:5' Untranslated Regions; D014758:Viral Core Proteins; C069130:nucleocapsid protein, Hepatitis C virus