detection of YMDD mutation of hepatitis B virus by allele-specific PCR and a cross-contamination proofed device

ABSTRACT

Lamivudine [LAM] is commonly used to treat Hepatitis B virus [HBV] infection, but its use frequently induces drug resistance. Therefore, rapid and correct detection of drug-resistant HBV is important for effective treatment of HBV infection. In this study, we aimed to develop a novel, simple, and user-friendly method for the detection of LAM resistant HBV. Samples were collected from 60 HBV infected patients for the analysis by allele-specific polymerase chain reaction [AS-PCR], nucleic acid detection strip [NADS] and a cross-contamination proofed device. HBV YMDD mutations were detected by AS-PCR, restriction fragment length polymorphism [RFLP] and DNA sequencing. A 91.7% concordance between all three methods was obtained. Compared to sequencing and RFLP, AS-PCR detected more samples with mutant variants and was more sensitive. This novel method had a detection limit of approximately 10[3]copies/ml and detected a variant of only 5% of total HBV population. In conclusion, we develop a new assay which could be useful for the detection of HBV LAM resistance, especially in resource-poor settings