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or hepatitis C virus (HCV) co-infection, is a significant cause of global health burden. About 130–170 million people are infected with HCV, representing 3 per cent of the world’s population. NS5B is responsible for the synthesis of negative-sense RNA and subsequently of positive-sense RNA that is incorporated into progeny virions. Importantly, the selective pressures that shape non-structural regions of the viral genome are distinct from those targeting structural genomic regions. For instance, highly conserved secondary RNA structures limit NS5B diversity. Immune-mediated selection pressures contribute to NS5B polymorphism, and HLA-restricted epitopes may overlap with sites of drug resistance. Immuneor drug-selected mutations in NS5B dramatically reduce viral replication in vivo, although compensatory mutations may develop. NS5B variability also impacts pathogenesis as a higher mutation rate is associated with elevated ALT levels, and NS5B enzymatic activity positively correlates with ALT levels. As additional non-structural gene inhibitors are developed, characterization of the factors that shape HCV diversity in vivo will be necessary to limit HCV replication and increase the effectiveness of new antiviral agents. The HIV Epidemiologic Research Study (HERS) was established in 1993 to prospectively define the biological, psychological, and social effects of HIV in US women. Serum samples were obtained from women who were HCV RNA positive but HIV negative, HIV/HCV co-infected with CD4<350, and HIV/HCV co-infected with CD4 350. HCV RNA was extracted using the QIAamp UltraSens Virus Kit and subjected to RT-PCR for the entire NS5B region ( 1,798 bases). Next-generation sequencing was used to evaluate intra-patient and interpatient NS5B diversity. NGS data were visualized in Integrated Genome Viewer. Consensus sequences were aligned in ClustalX 2.1 and BEAST v1.8.4 under an uncorrelated log-normal relaxed molecular clock, the general time-reversible model with nucleotide site heterogeneity estimated using a gamma distribution, and a chain length of 100,000,000 with sampling every 10,000th generation.

A40 Genotypic diversity of HCV in Kosovo with an emphasis on phylogenetic investigation of subtype 4D