Real-world performance of the NeuMoDx™ HCV Quant Test for quantification of hepatitis C virus (HCV)-RNA.

Quantification of hepatitis C virus (HCV)-RNA in serum or plasma samples is an essential parameter in HCV diagnostics. Here, the NeuMoDx™Molecular System (Qiagen) was tested for the most common HCV genotypes and compared to the cobas c6800 system (Roche). HCV-RNA from 131 plasma/serum samples from chronically infected patients was determined in parallel on the NeuMoDx and c6800 systems. Linearity was analysed using the four most common HCV genotypes (1-4) in our cohort. The coefficient of variation (CV) within (intra-assay) and between (inter-assay) runs was calculated based on HCV-RNA concentration. Quantitative HCV-RNA results were highly correlated on both test systems (R2 = 0.7947; y = 0.94 x + 0.37). On average, the NeuMoDx and c6800 HCV RNA levels showed a mean difference of only 0.05 log10 IU/mL but with a broad distribution (±1.2 2 x SD). The NeuMoDx demonstrated very good linearity across all HCV genotypes tested at concentrations between 1.7 and 6.2 log10 IU/mL (R2 range: 0.9257-0.9991) with the highest mean coefficient of determination for genotype 1 (R2 = 0.9909). The mean intra- and inter-assay CV for both serum and plasma samples was <5 %. The NeuMoDx HCV-RNA Assay demonstrates high subtype-independent comparability, linearity, and reproducibility for the quantification of HCV-RNA in serum and plasma samples from chronically infected patients.

A genotype independent, full-genome reverse-transcription protocol for HCV genotyping and resistance testing.

BACKGROUND: HCV treatment options and cure rates have tremendously increased in the last decade. Although a pan-genotype HCV treatment has recently been approved, most DAA therapies are still genotype specific. Resistance-associated variants (RAVs) can limit the efficacy of DAA therapy and are associated with increased risk for therapy failure. With the approval of DAA regimens that recommend resistance testing prior to therapy, correct assessment of the genotype and testing for viruses with RAVs is clinically relevant. However, genotyping and resistance testing is generally done in costly and laborious separate reactions. OBJECTIVE: The aim of the study was to establish a genotype-independent full-genome reverse transcription protocol to generate a template for both genotyping and resistance testing and to implement it into our routine diagnostic setup. STUDY DESIGN: The complete HCV genome was reverse transcribed with a pan-genotype primer binding at the 3'end of the viral RNA. This cDNA served as template for transcription of the genotyping amplicon in the core region as well as for the resistance testing of NS3, NS5A, and NS5B. RESULTS: With the established RT-protocol the HCV core region was successfully amplified and genotyped from 124 out of 125 (99.2%) HCV-positive samples. The amplification efficiency of RAV containing regions in NS3, NS5A, NS5B was 96.2%, 96.6% and 94.4%, respectively. CONCLUSIONS: We developed a method for HCV full-genome cDNA synthesis and implemented it into a routine diagnostic setup. This cDNA can be used as template for genotyping amplicons covering the core or NS5B region as well as for resistance testing amplicons in NS3, NS5A and NS5B.