Hepatitis B virus DNA polymerase gene polymorphism based prediction of  genotypes in chronic HBV patients from Western India.

Yashwant G Chavan, Sharad R Pawar, Minal Wani, Amol D Raut, Rabindra N Misra
1. Dr. D Y Patil Biotechnology and Bioinformatics Institute, Tathwade, Pune 411033. Dr. D Y Patil Vidyapeeth,
Sant Tukaram Nagar, Pimpri, Pune 411018. Maharashtra, India.
2. Medical Genetics, geneOmbio Technologies Private Limited, Baner, Pune 411045. Maharashtra, India.
3. Department of Microbiology, Dr. D Y Patil Medical College, Dr. D Y Patil Vidyapeeth, Sant Tukaram Nagar,
Pimpri, Pune 411018. Maharashtra, India.

Abstract
Background: Hepatitis B Virus (HBV) infection is one of the major causes of liver cirrhosis, hepatocellular carcinoma and  deaths due to the acute or chronic consequences worldwide. HBV is distributed into various genotypes based on nucleic acid  sequence variation.

Objectives: To develop a method of HBV genotyping and drug resistance interpretation using partial sequencing of polymerase  gene.

Methods: This study was performed on 98 HBV infected patients’ serum samples from Western India. A nested PCR protocol  was designed for amplification of pol gene from HBV genome and Sanger’s sequencing of the gene fragment. Sequences were  aligned with HBV reference sequences for phylogenetic analysis and for characterization of genetic diversity. Drug resistance  mutations were screened using HBVSeq program from Stanford University.

Results: Distribution of HBV genotypes showed predominance of genotype D, circulating in 76 (77.55%) patients (p < 0.05).  Genotypes A and C were less prevalent and were identified in 4 (4.08%) and 18 (18.37%) patients, respectively. Anti-retroviral  drug resistance mutations were not detected in any patient.

Conclusion: A method for determination of HBV genotypes using pol gene sequencing which simultaneously detects major  drug resistance mutations has been established. HBV genetic diversity may play an important role in treatment decision.

Keywords: Hepatitis B virus, nested PCR, genotype, sub-genotypes, YMDD mutations