RESUMEN
A rapid, duplex, high-resolution melting interleukin-28B gene (IL28B) genotyping assay, targeting both rs12979860 and rs8099917 polymorphisms, was developed and validated using 30 DNA samples from healthy volunteers. A linkage study on 300 healthy Singaporeans showed variable haplotypes. When the assay was applied to plasma DNA from 50 hepatitis C virus genotype-1 (HCV-1)-infected patients, five compound heterozygous types were detected.
Asunto(s)
Técnicas de Laboratorio Clínico/métodos , Hepatitis C/tratamiento farmacológico , Interleucinas/genética , Técnicas de Diagnóstico Molecular/métodos , Polimorfismo de Nucleótido Simple , Genotipo , Haplotipos , Hepacivirus/clasificación , Hepacivirus/genética , Hepacivirus/aislamiento & purificación , Hepatitis C/virología , Humanos , Interferones , Desnaturalización de Ácido Nucleico , Singapur , Temperatura de TransiciónRESUMEN
BACKGROUND: Correct identification of infecting hepatitis C virus (HCV) genotype is helpful for targeted antiviral therapy. OBJECTIVES: Here, we compared the HCV genotyping performance of the cobas HCV GT assay against the Versant HCV Genotype 2.0 (LiPA) assay, using 97 archived serum samples. STUDY DESIGN: In the event of discrepant or indeterminate results produced by either assay, the core and NS5B regions were sequenced. RESULTS: Of the 97 samples tested by the cobas, 25 (26%) were deemed indeterminate. Sequencing analyses confirmed 21 (84%) of the 25 samples as genotype 6 viruses with either subtype 6m, 6n, 6v, 6xa, or unknown subtype. Of the 97 samples tested by the LiPA, thirteen (13%) were deemed indeterminate. Seven (7%) were assigned with genotype 1, with unavailable/inconclusive results from the core region of the LiPA. Notably, the 7 samples were later found to be either genotype 3 or 6 by sequencing analyses. Moreover, 1 sample by the LiPA was assigned as genotypes 4 (cobas: indeterminate) but were later found to be genotype 3 by sequencing analyses, highlighting its limitation in assigning the correct genotype. CONCLUSIONS: The cobas showed similar or slightly higher accuracy (100%; 95% CI 94-100%) compared to the LiPA (99%; 95% CI 92-100%). Twenty-six percent of the 97 samples tested by the cobas had indeterminate results, mainly due to its limitation in identifying genotype 6 other than subtypes 6a and 6b. This presents a significant assay limitation in Southeast Asia, where genotype 6 infection is highly prevalent.