Brief Article
Copyright ©2010 Baishideng. All rights reserved.
World J Gastroenterol. Mar 14, 2010; 16(10): 1267-1273
Published online Mar 14, 2010. doi: 10.3748/wjg.v16.i10.1267
Establishment of a new quantitative detection approach to adefovir-resistant HBV and its clinical application
Wei-Feng Zhao, You-Lin Shao, Liang-Yun Chen, Jin-Hua Wu, Yi-Ling Zhu, Jian-He Gan, Hui Xiong
Wei-Feng Zhao, You-Lin Shao, Liang-Yun Chen, Jin-Hua Wu, Yi-Ling Zhu, Jian-He Gan, Department of Infectious Diseases, First Affiliated Hospital of Soochow University, Suzhou 215006, Jiangsu Province, China
Hui Xiong, Shanghai Laboratory of Disease and Health Genomics, Chinese National Human Genome Center, Shanghai 201203, China
Author contributions: Zhao WF performed the quantitative RT-PCR, direct sequencing analysis and wrote the manuscript; Shao YL and Chen LY performed the T-A cloning, quantitative RT-PCR and direct sequencing analysis, and wrote the manuscript; Wu JH collected the serum samples and clinical data; Zhu YL collected the serum samples; Gan JH coordinated the study; Xiong H contributed to the overall interpretation of data and participated in writing the manuscript.
Supported by The fund from Health Project of Jiangsu Province, No. H200711 and the AIDS, Hepatitis B and Other Infectious Diseases Prevention Program, No. 2009ZX10004-712
Correspondence to: Hui Xiong, PhD, Shanghai Laboratory of Disease and Health Genomics, Chinese National Human Genome Center, Shanghai 201203, China. xionghui@chgc.sh.cn
Telephone: +86-21-58554463 Fax: +86-21-58557486
Received: March 4, 2009
Revised: March 27, 2009
Accepted: April 3, 2009
Published online: March 14, 2010
Abstract

AIM: To establish the more feasible and sensitive assessment approach to the detection of adefovir (ADV) resistance-associated hepatitis B virus (HBV) quasispecies.

METHODS: Based on the characteristics of rtA181V/T and rtN236T mutations, a new approach based on real-time fluorescent quantitative polymerase chain reaction (RT-PCR) was established for the detection of ADV-resistant HBV quasispecies, total HBV DNA, rtA181 and rtN236 mutations in blood samples from 32 chronic hepatitis B (CHB) patients with unsatisfactory curative effect on ADV and compared with routine HBV DNA sequencing.

RESULTS: Both the sensitivity and specificity of this new detection approach to ADV-resistant HBV quasispecies were 100%, which were much higher than those of direct HBV DNA sequencing. The approach was able to detect 0.1% of mutated strains in a total plasmid population. Among the 32 clinical patients, single rtA181 and rtN236T mutation and double rtA181T and rtN236T mutations were detected in 20 and 8, respectively, while ADV-resistant mutations in 6 (including, rtA181V/T mutation alone in 5 patients) and no associated mutations in 26.

CONCLUSION: This new approach is more feasible and efficient to detect ADV-resistant mutants of HBV and ADV-resistant mutations before and during ADV treatment with a specificity of 100% and a sensitivity of 100%.

Keywords: Chronic hepatitis B, Adefovir, Drug resistance, Quantitative detection, Real-time fluorescent quantitative polymerase chain reaction