Clinical evaluation of a newly developed automated massively parallel sequencing assay for hepatitis C virus genotyping and detection of resistance-association variants. Comparison with a line probe assay

•The HCV genotype has been shown to compromise therapy. Genotype1 is important to select optimal drug regimens.•This comparative evaluation between the two methods demonstrates an efficient method for accurate HCV genotyping.•Sanger sequencing is a reference method that assists accurate HCV genotyping.•The Q80K associates with reduced response to NS3 inhibitor, and Y93H with reduced response to NS5A inhibitor found frequently in Thai patients.

Hepatitis C virus (HCV) infection is a leading cause of chronic liver disease, cirrhosis and hepatocellular carcinoma. Recently, HCV was classified into 6 major genotypes (GTs) and 67 subtypes (STs). Efficient genotyping has become an essential tool for prognosis and indicating suitable treatment, prior to starting therapy in all HCV-infected individuals. The widely used genotyping assays have limitation with regard to genotype accuracy. This study was a comparative evaluation of exact HCV genotyping in a newly developed automated-massively parallel sequencing (MPS) system, versus the established Line probe assay 2.0 (LiPA), substantiated by Sanger sequencing, using 120 previously identified-HCV RNA positive specimens. LiPA gave identical genotypes in the majority of samples tested with MPS. However, as much as 25% of LiPA did not identify subtypes, whereas MPS did, and 0.83% of results were incompatible. Interestingly, only MPS could identify mixed infections in the remaining cases (1.67%). In addition, MPS could detect Resistance-Associated Variants (RAVs) simultaneously in GT1 in 56.82% of the specimens, which were known to affect drug resistance in the HCV NS3/NS4A and NS5A genomic regions. MPS can thus be deemed beneficial for guiding decisions on HCV therapy and saving costs in the long term when compared to traditional methods.