Assessments of intra- and inter-host diversity of hepatitis C virus using Next Generation Sequencing and Mass spectrometry

Recent advances in sequencing methods allow the analysis of an unprecedented number of viral variants from infected patients and present a novel opportunity for understanding viral evolution, drug resistance and immune escape. In the present paper, we compared three technologies for amplicon analysis: (i) Next Generation Sequencing; (ii) Clonal sequencing using End-point Limiting-dilution for isolation of individual sequence variants followed by Real-Time PCR and sequencing; and (iii) Mass spectrometry of base-specific cleavage reactions of a target sequence. Hypervariable region 1 of hepatitis C virus was analyzed using these three technologies to assess diversity and genetic relatedness of intra-host viral populations in specimens obtained from 38 patients. Estimates of population heterogeneity varied among technologies. However, all three technologies were equally accurate in identification of genetic relatedness among viral strains, supporting their application in molecular epidemiology for tracking viral variants and detecting transmission events.