TY - JOUR
T1 - Direct detection of lamivudine-resistant hepatitis B virus mutants by a multiplex PCR using dual-priming oligonucleotide primers
AU - Kim, Jong Kee
AU - Lee, Hyeon Ji
AU - Lee, Young Jo
AU - Chun, Jong Yoon
AU - Lee, In Kyoung
AU - Lim, Young Suk
AU - Suh, Dong Jin
AU - Ko, Sun Young
AU - Kim, Myeong Hee
AU - Oh, Heung Bum
N1 - Funding Information:
This research was supported, in part, by the Intelligent Microsystem Center (IMC; http://www.microsystem.re.kr ), which is performing one of the 21st century Frontier R&D Projects sponsored by the Korea Ministry Of Commerce, Industry and Energy, and by a grant (2007-219) from the Asan Institute for Life Sciences, Seoul, Korea.
Copyright:
Copyright 2008 Elsevier B.V., All rights reserved.
PY - 2008/4
Y1 - 2008/4
N2 - Mutations in the YMDD motif of the hepatitis B virus (HBV) polymerase gene increase lamivudine resistance of HBV, highlighting the clinical importance of accurate and sensitive detection of HBV mutants. Using dual-priming oligonucleotide primer technology, an assay that can detect mutations at codons 180 (L528M) and 204 (YVDD, YIDD, and YSDD) by a single-step multiplex PCR was developed. This Seeplex Lami-DR assay was sufficiently sensitive to detect 103 HBV/ml and was able to detect minor mutants comprising as little as 2% of the viral population. Mutants were detected in 57 of 65 serum samples (88%) from patients with chronic hepatitis B who had been treated with lamivudine (median, 32 months; range, 1-83 months). The agreement with direct sequencing was only 38.5% (25/65). Discrepancies between these methods resulted from detection of additional mutants by the Seeplex Lami-DR assay, as confirmed by a novel verification analysis. This assay is not only highly accurate and sensitive, but is also simple and cost-effective, requiring no expensive probes, laborious sequencing procedures, or digestion with restriction enzymes. Accordingly, the Seeplex HBV Lami-DR assay should be considered as a first-line, cost-effective tool for detecting viral mutations in patients with chronic hepatitis B receiving lamivudine therapy.
AB - Mutations in the YMDD motif of the hepatitis B virus (HBV) polymerase gene increase lamivudine resistance of HBV, highlighting the clinical importance of accurate and sensitive detection of HBV mutants. Using dual-priming oligonucleotide primer technology, an assay that can detect mutations at codons 180 (L528M) and 204 (YVDD, YIDD, and YSDD) by a single-step multiplex PCR was developed. This Seeplex Lami-DR assay was sufficiently sensitive to detect 103 HBV/ml and was able to detect minor mutants comprising as little as 2% of the viral population. Mutants were detected in 57 of 65 serum samples (88%) from patients with chronic hepatitis B who had been treated with lamivudine (median, 32 months; range, 1-83 months). The agreement with direct sequencing was only 38.5% (25/65). Discrepancies between these methods resulted from detection of additional mutants by the Seeplex Lami-DR assay, as confirmed by a novel verification analysis. This assay is not only highly accurate and sensitive, but is also simple and cost-effective, requiring no expensive probes, laborious sequencing procedures, or digestion with restriction enzymes. Accordingly, the Seeplex HBV Lami-DR assay should be considered as a first-line, cost-effective tool for detecting viral mutations in patients with chronic hepatitis B receiving lamivudine therapy.
KW - Chronic hepatitis B
KW - HBV
KW - Lamivudine resistance
KW - Seeplex Lami-DR assay
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U2 - 10.1016/j.jviromet.2008.01.003
DO - 10.1016/j.jviromet.2008.01.003
M3 - Article
C2 - 18291537
AN - SCOPUS:40849101514
VL - 149
SP - 76
EP - 84
JO - Journal of Virological Methods
JF - Journal of Virological Methods
SN - 0166-0934
IS - 1
ER -