Dynamic study on genomic characterization of hepatitis virus B isolated from two multi-drug refractory patients

OJEDA DIEGO, CASSINO LUCILA, MORETTI FRANCO, BOLCIC FEDERICO, BENETTI SILVINA, TANNO HUGO, QUARLERI JORGE

Paphos, Cyprus

Workshop; 9th European Workshop on HIV & Hepatitis; 2011

Background: Antiviral drug-resistant hepatitis B virus (HBV) mutants under a variety of treatment protocols are complex and only partly understood. Prolonged treatment of chronic hepatitis B (CHB) with nucleoside analogues (NAs) almost invariably engenders viral resistance, and sequential NAs monotherapy can promote multi-drug resistance. This study is a dynamic analysis from the serial sera of two patients (ARG-V and ARG-N) during the antiviral treatment aimed to investigate the molecular characteristics and the mutation profile of multi-drug resistant hepatitis B virus (HBV). Material & Methods: Two patients were followed-up during 14 (ARG-V) and 4 (ARG-N) years. HBV viral load by real time PCR (COBAS TaqMan Roche Molecular Systems, dynamic range 30 IU/ml to 110,000,000 IU/ml), HBeAg/Anti-HBe serological profile determined by electrochemiluminescence (ELECSYS 2010, Roche Diagnostic) and amino transferase and gamma-glutamil transpeptidase levels were measured during the follow-up. The partial surface/polymerase genomic sequences from consecutive HBV isolates from two multi-drug refractory patients were amplified and sequenced. The analysis was performed using an ABI3100 instrument (Applied Biosystems), and the sequences introduced in this work as well as those obtained from the GenBank database were aligned with ClustalX v1.83 and edited with Bioedit v7.0.9.0. HBV genotyping was carried out by phylogenetic inference by using maximum likelihood method.  Results: The phylogenetic analysis showed that isolates from patient ARG-V and ARG-N were classified as subgenotype A2 and F1, respectively. Conclusions: In conclusion, the complex HBV mutation profile detected in multi-drug refractory patients highlights the problems associated with the ongoing selection of mutations, including further compensatory mutations as well as potential cross-resistance mutations.