HBV YMDD 돌연변이 검출을 위한 염기서열분석법과 RestrictionFragmentMassPolymorphism 법의 비교
The Comparison of Restriction Fragment Mass Polymorphism Method withSequecing Method for the Detection of HBV YMDD Mutants
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배경 : 만성B형 간염 치료제인 라미부딘을 장기 투여시 약제 내성 HBV 변이형의 출현이 나타났으며, 대부분은 YMDD motif에서 일어난다. 최근 국내에서 restriction fragment mass polymor-phism (RFMP)법을 이용한 YMDD 변이형 검출법이 소개되었다. 저자들은 라미부딘 치료를 받은 후 바이러스 breakthrough가 있었던 만성B형 간염 환자들을 대상으로 염기서열분석법과 RFMP법을 이용하여 두 검사법의 유용성을 비교하고자 하였다. 방법 : 라미부딘을 투여하면서 Hybrid capture II HBV DNA test (Digene Inc., Gaithersburg, MD, USA)에서 치료전 양성, 치료 후 음성으로 전환 후 다시 재양성 결과를 보인 18명의 환자를 대상으로 하였다. 각 시기의 검체를 염기서열분석법과 RFMP법으로 YMDD 변이형 유무를 관찰하였다. 결과 : 바이러스 breakthrough 상태에서 두 방법 모두에서 YMDD 변이형 13예(72.2%)가 검출되었으며, RFMP법에서는 2예를 혼합감염 상태로 판독하였다. 바이러스 breakthrough 이전에는 염기서열분석법으로 3예(16.7%), RFMP법으로 4예(22.2%)에서 변이형이 관찰되었는데, RFMP법에서 검출된 변이형 중 2예가 염기서열분석 결과가 달랐다. 54건의 검체에서 두 검사간 일치율은 92.6%이었다. 결론 : 염기서열분석법이나 RFMP법의 결과의 일치도가 높다고 사료되는 바 두 방법 모두 변이형 검출에 적절하다고 사료된다.
Abstract
Background : YMDD motif mutants of the hepatitis B virus (HBV) emerged in some chronic hepatitis B patients after prolonged lamivudine treatment. Recently a novel genotyping assay, the restriction fragment mass polymorphism (RFMP) method, was introduced for the detection of YMDD mutations. We compared the performance of the RFMP method with that of sequencing method in chronic hepatitis B patients who had suffered the HBV DNA breakthrough after lamivudine treatment. Methods : Enrolled in this study were 18 chronic hepatitis B patients who experienced the DNA breakthrough after a period during which HBV DNA was undetectable by Hybrid capture II HBV DNA test (Digene Inc., Gaithersburg, MD, USA). Both sequencing and RFMP methods were used to detect YMDD variants in three phases such as before treatment, before breakthrough and after breakthrough. Results : YMDD mutants were detected in 13 samples (72.2%) by both methods after DNA breakthrough. Among them were two samples with a mixed HBV population detected by RFMP. Before breakthrough, the mutants were detected in three samples (16.7%) by sequencing and four (22.2%) by RFMP, showing discrepant results for two samples. The concordance rate between both methods was 92.6%. Conclusions : Both sequencing and RFMP methods were highly concordant except in a few cases, so it is suggested that both methods are appropriate in detecting YMDD mutants. (Korean J Lab Med 2004; 24: 320-6)
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