An improved reverse dot hybridization for simple and rapid detection of adefovir dipivoxil-resistant hepatitis B virus.
An improved reverse dot hybridization for simple and rapid detection of adefovir dipivoxil-resistant hepatitis B virus.
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DOI：
10.4238/2012.january.9.6
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发表时间：
2012-01
期刊：
影响因子：
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通讯作者：
Y. Hu;W. Zhang;S. Xie;Y. Zhao;J. Hu;X. Cai;G. Lai;A. Huang
中科院分区：
文献类型：
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作者：
Y. Hu;W. Zhang;S. Xie;Y. Zhao;J. Hu;X. Cai;G. Lai;A. Huang
在长期使用阿德福韦酯治疗慢性B型肝炎病毒（HBV）感染的过程中，早期发现阿德福韦酯耐药突变株具有重要的临床意义。我们开发了一种改良的反向斑点杂交试验，用于简单快速地检测慢性B型肝炎患者阿德福韦酯耐药相关的rtA 181 V/T和rtN 236 T突变。针对这种耐药特征的基因型B、C和D设计探针;用这种改进的反向斑点杂交法分析了总共70个临床样本。通过与测序数据的比较验证了其有效性。通过亚克隆测序证实了不一致的结果。该反向斑点杂交试验的灵敏度足以检测10（3）拷贝/mL;当阿德福韦酯耐药突变株占混合病毒群体的5%以上时，也可检出阿德福韦酯耐药突变株。该反向斑点杂交阵列正确鉴定了阿德福韦酯耐药突变体，与直接测序数据具有较高的一致性（98.5%）。HBV基因型与阿德福韦酯耐药突变株的发生之间没有明确的关系。这种反向斑点杂交法被证明是简单和快速检测与阿德福韦酯耐药相关的rtA181 V/T和rtN236 T突变。
Early detection of adefovir dipivoxil-resistant mutants during long-term treatment of chronic hepatitis B virus (HBV) infection with this drug is of great clinical importance. We developed an improved reverse dot hybridization test for simple and rapid detection of the rtA181V/T and rtN236T mutations associated with adefovir dipivoxil resistance in chronic hepatitis B patients. Probes were designed for genotypes B, C, and D of this resistance characteristic; a total of 70 clinical samples were analyzed with this improved reverse dot hybridization assay. Its usefulness was validated by comparing with sequencing data. Discordant results were confirmed by subclone sequencing. This reverse dot hybridization assay was sufficiently sensitive to detect 10(3) copies/mL; it also detected adefovir dipivoxil-resistant mutant strains when they comprised more than 5% of a mixed virus population. This reverse dot hybridization array correctly identified adefovir dipivoxil-resistant mutants; it had high concordance (98.5%) with direct sequencing data. There was no clear relationship between the HBV genotype and the development of adefovir dipivoxil-resistant mutants. This reverse dot hybridization assay proved to be simple and rapid for detection of rtA181V/T and rtN236T mutations associated with resistance to adefovir dipivoxil.