Genotyping the hepatitis B virus with a fragment of the HBV DNA polymerase gene in Shenyang, China

Department of Neurology, Shengjing Hospital of China Medical University, Shenyang 110817, China.
Virology Journal (Impact Factor: 2.18). 06/2011; 8(1):315. DOI: 10.1186/1743-422X-8-315
Source: PubMed


The hepatitis B virus (HBV) has been classified into eight genotypes (A-H) based on intergenotypic divergence of at least 8% in the complete nucleotide sequence or more than 4% in the S gene. To facilitate the investigation of the relationship between the efficacy of drug treatment and the mutation with specific genotype of HBV, we have established a new genotyping strategy based on a fragment of the HBV DNA polymerase gene. Pairwise sequence and phylogenetic analyses were performed using CLUSTAL V (DNASTAR) on the eight (A-H) standard full-length nucleotide sequences of HBV DNA from GenBank (NCBI) and the corresponding semi-nested PCR products from the HBV DNA polymerase gene. The differences in the semi-nested PCR fragments of the polymerase genes among genotypes A through F were greater than 4%, which is consistent with the intergenotypic divergence of at least 4% in HBV DNA S gene sequences. Genotyping using the semi-nested PCR products of the DNA polymerase genes revealed that only genotypes B, C, and D were present in the 50 cases, from Shenyang, China, with a distribution of 11 cases (22%), 25 cases (50%), and 14 cases (28%) respectively. These results demonstrate that our new genotyping method utilizing a fragment of the HBV DNA polymerase gene is valid and can be employed as a general genotyping strategy in areas with prevalent HBV genotypes A through F. In Shenyang, China, genotypes C, B, and D were identified with this new genotyping method, and genotype C was demonstrated to be the dominant genotype.

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    • "HBV genotype classification is based on a nucleotide sequence divergence of 8%4, although in sequences of the same genotype, naturally occurring polymorphic sites are often observed. Therefore, consensus genotype reference sequences of HBV polymerase for genotypes B and C are essential for homology model construction. "
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    ABSTRACT: Aim: To investigate the molecular mechanisms underlying the influence of DNA polymerase from different genotypes of hepatitis B virus (HBV) on the binding affinity of adefovir (ADV). Methods: Computational approaches, including homology modeling, docking, MD simulation and MM/PBSA free energy analyses were used. Results: Sequence analyses revealed that residue 238 near the binding pocket was not only a polymorphic site but also a genotype-specific site (His238 in genotype B; Asn238 in genotype C). The calculated binding free-energy supported the hypothesis that the polymerase from HBV genotype C was more sensitive to ADV than that from genotype B. By using MD simulation trajectory analysis, binding free energy decomposition and alanine scanning, some energy variation in the residues around the binding pocket was observed. Both the alanine mutations at residues 236 and 238 led to an increase of the energy difference between genotypes C and B (ΔΔGC–B), suggesting that these residues contributed to the genotype-associated antiviral variability with regard to the interaction with ADV. Conclusion: The results support the hypothesis that the HBV genotype C polymerase is more sensitive to ADV than that from genotype B. Moreover, residue N236 and the polymorphic site 238 play important roles in contributing to the higher sensitivity of genotype C over B in the interaction with ADV.
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    • "Considering the importance of determining the HBV genotypes, several methods have been developed for genotyping of HBV strains such as restriction fragment length polymorphism, line probe assay, and quantitative real-time PCR [16] [25] [26] [27] [28] [29] [30]. Here we report the prevalent genotype found among HBV-infected Filipino patients using DNA sequencing. "
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    • "China is a vast country and different genotypes of HBV have been reported. In Shenyang, the largest city in the northeast of China, HBV/C was widely distributed (50%) in chronic patients or asymptomatic carriers along with HBV/B (22%) and HBV/D (28%) [36]. In Xian from the northwest region of China, 11 strains from blood donors were clustered into genotype B and C [30]. "
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