High frequency of complex mutational patterns in lamivudine resistant hepatitis B virus isolates

The evolution of hepatitis-B virus (HBV) drug resistance is characterized by the emergence of resistance conferring mutations and compensatory mutations. Therefore HBV drug-resistant isolates often harbor multiple mutations in the reverse transcriptase (RT) and corresponding mutations in the hepatitis B surface antigen (HBsAg). In this study mutational patterns of 60 HBV isolates harboring drug resistance mutations rtM204V or rtM204I were retrospectively analyzed. Both mutations, especially mutation rtM204V, were most often accompanied by compensatory mutations rtV173L and rtL180M but also by mutations conferring entecavir (n = 5) or adefovir resistance (n = 4). In addition, 22 (36.7%) drug resistant HBV isolates carried mutations related to immune escape in the HBsAg. In seven cases premature stop codons in HBsAg were detected resulting in the expression of truncated HBsAg. Clonal analysis of these seven quasispecies even disclosed the presence of HBV isolates carrying further stop codons and in one case the occurrence of resistance mutation rtA181T without the stop codon mutation sW172*. Interestingly, only one HBV clone carried the resistance mutations rtM204V and rtA181T. HBV drug resistant isolates frequently harbored HBsAg mutations associated with immune escape or disease progression pointing to a complex interaction of both proteins. HBV genotypic resistance tests based on population sequencing methods seemed to be inappropriate for determining the clinical relevance of stop codons in the HBsAg.