Full-genome next-generation sequencing of hepatitis C virus to assess the accuracy of genotyping by the commercial assay LiPA and the prevalence of resistance-associated substitutions in a Belgian cohort

Background: Although most currently used regimens for Hepatitis C virus (HCV) infections can be initiated without prior knowledge of genotype and subtype, genotyping is still useful to identify patients who might benefit from a personalized treatment due to resistance to direct-acting antivirals (DAA).Objectives: To assess the utility of full-genome next-generation sequencing (FG-NGS) for HCV genotyping. Study design: 138 HCV plasma samples previously genotyped by VERSANT HCV Genotype Assay (LiPA) were subjected to FG-NGS and phylogenetically genotyped Genome Detective. Consensuses were analysed by HCV-GLUE for resistance-associated substitutions (RASs) and their impact on treatment response was investigated.Results: 102/138 (73.9%) samples were sequenced to a genome coverage and depth of >90% of the HCV open reading frame covered by >100 reads/site. Concordant genotype and subtype results were assigned in 97.1% and 79.4% of samples, respectively. FG-NGS resolved the subtype of 13.7% samples that had ambiguous calls by LiPA and identified one dual infection and one recombinant strain. At least one RAS was found for the HCV genes NS3, NS5A, and NS5B in 2.91%, 36.98% and 27.3% samples, respectively. Irrespective of the observed RAS, all patients responded well to DAA treatment, except for HCV1b-infected patients treated with Zepatier (33.3% failure rate (5/15)).Conclusion: While LiPA and FG-NGS showed overall good concordance, FG-NGS improved specificity for sub-types, recombinant and mixed infections. FG-NGS enabled the detection of RAS, but its predictive value for treatment outcome in DAA-naive patients remains uncertain. With additional refinements, FG-NGS may be the way forward for HCV genotyping.

Automated summary

The utility of full-genome next-generation sequencing (FG-NGS) for HCV genotyping was assessed in this study. FG-NGS showed improved specificity for subtypes, identified mixed infections and recombinant strains, and detected resistance-associated substitutions (RASs). However, the predictive value of FG-NGS for treatment outcome in DAA-naive patients remains uncertain. With further refinements, FG-NGS may be the future of HCV genotyping.