Rapid Communication
Copyright ©2006 Baishideng Publishing Group Co., Limited. All rights reserved.
World J Gastroenterol. Feb 28, 2006; 12(8): 1308-1311
Published online Feb 28, 2006. doi: 10.3748/wjg.v12.i8.1308
Detection of YMDD mutants using universal template real-time PCR
Rong-Sheng Wang, Hua Zhang, Yu-Fen Zhu, Bei Han, Zhi-Jun Yang
Rong-Sheng Wang, Hua Zhang, Yu-Fen Zhu, Bei Han, Jiangsu Oil Field Hospital, Shaobo, Jiangdu 225261, Jiangsu Province, China
Zhi-Jun Yang, Fosun Diagnositcs, Shanghai 200233, China
Author contributions: All authors contributed equally to the work.
Correspondence to: Zhi-Jun Yang, Fosun Diagnositics, 1289 Yishan Road, Shanghai 200233, China. yangzj@fosun.com.cn
Telephone: +86-21-64952059 Fax: +86-21-64955476
Received: August 10, 2005
Revised: October 15, 2005
Accepted: October 26, 2005
Published online: February 28, 2006

AIM: To establish a rapid and accurate method for the detection of lamivudine-resistant mutations in hepatitis B virus and monitor of lamivudine resistance during lamivudine treatment in patients with chronic hepatitis B virus infection.

METHODS: We established a real-time PCR method using a universal template and TaqMan probe to detect YMDD mutants. Variants of YVDD and YIDD were tested by individual reactions (reaction V and reaction I) and total hepatitis B viruses were detected in another reaction for control (reaction C). Results were determined by ΔCt < 3.5 (ΔCt = Ct of reaction V or I - Ct of reaction C). Clones of the HBV polymerase gene containing different YMDD mutations were tested. Serum samples from 163 lamivudine-treated patients with chronic hepatitis B virus infection were detected using this method and the results were confirmed by DNA sequencing.

RESULTS: As many as 1 000 copies per milliliter of wide-type plasmid were detected and nonspecific priming was excluded. In the 163 samples from patients treated with lamivudine, lamivudine-resistant mutations were detected in 51 samples.

CONCLUSION: This universal real-time PCR is a rapid and accurate method for quantification of YMDD mutants of HBV virus in lamivudine-treated patients and can be used to monitor lamivudine-resistant mutations before and during lamivudine therapy.

Keywords: HBV, YMDD, Mutation, UT-PCR