Ultra-Deep Sequencing Analysis Of Hepatitis B Virus Precore/Core Variants Among Prison-Inmates

FARRID BOADU 94 PAGES (16843 WORDS) Microbiology Thesis

ABSTRACT Infection with hepatitis B virus constitutes a major public health problem globally. The lack of proofreading of the reverse transcriptase results in considerable mutations culminating into genotypes, sub-genotypes and variants. HBV quasispecies in an infected host has been linked to disease progression and therapeutic outcome. Limited data exist on the quasispecies diversity in other clinically relevant regions particularly the precore/core ORF which also is a determinant factor in the progression and treatment of this infection. Therefore, the need to explore this phenomenon is needed especially in populations with frequent exposure to HBV to help understand the pathogenesis of co-infections, recombination and evolution of HBV. The aim of this study was to examine the diversity and mutations at subpopulations level of HBV across the precore/core genomic region. Initially, viral DNA was extracted from twenty-six (26) archived plasma samples of prison-inmates who participated in a nationwide HBV/HIV surveillance study. Out of twenty-six samples, five (5) precore/core ORF (designated as S1) was successfully amplified and next-generation sequencing done with the Illumina MiSeq platform. Clones from 3 inmates were analysed for co-infection and clinically relevant mutations. A common pattern of mutations that was observed across the precore region was W28*. Additionally, in the core ORF mutations such as I97F, P130T and R181P were observed. Phylogenetic analysis of the individual clones of HBV revealed the predominance of genotype E at the subpopulation level. However, almost half (5 / 13) of the clones from 4S1 clustered with genotype G reference sequences. Majority of the 5S1 clones clustered with reference sequences of genotype E with the exception of one which clustered with genotype A. Clinically relevant mutations may be found in most quasispecies and will have implications for disease progression and e-antigenemia. HBV clones observed may give insight into the evolution of A/E and G/E recombinants.