Genotyping and resistant mutants analysis of P gene in HBV fragment sequencing

Abstract

Objective To establish a method for efficient, accurate genotyping and nucleoside drug-resistant mutation analysis for hepatitis B virus (HBV). Methods The 48 HBV serum samples were collected from the Third People′s Hospital of Taiyuan from July to August 2011，and HBV DNA were extracted using the commercial kit.The HBV whole genome and P gene were amplified and sequenced.Each HBV sample was genotyped by both constructing phylogenetic trees and genotyping software analysis.The results from two strategies were compared for every sample. Results A total of 48 HBV full genome sequences were identified into 12 B and 36 C genotype′s by both constructing phylogenetic trees and genotyping software analysis, which was exactly the same as the analysis using P gene fragment sequencing.Seven forms of nucleoside drug-resistant mutation were found in the P gene for all the samples, with the ratio of 27.1%（13/48）, in which all the mutation forms were associated with lamivudine or adefovir, and no other nucleotide drugs-related resistance mutations existed.In addition, there were 11 B and 35 C genotype and 2 B/C hybrid type with the analysis using Real-time PCR genotyping for the 48 samples. Conclusion P gene sequencing can be used as a new clinical method for efficient, accurate HBV genotyping and resistant mutation analysis，which provides guidance for hepatitis B treatment. （Chin J Lab Med,2012,35:726-729） Key words: Hepatitis B virus; Genotype; Sequence analysis，DNA; Lamivudine; Drug resistance，viral