Rapid diagnosis of human herpesvirus 6 infection by a novel DNA amplification method, loop-mediated isothermal amplification
Rapid diagnosis of human herpesvirus 6 infection by a novel DNA amplification method, loop-mediated isothermal amplification
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DOI：
10.1128/jcm.42.1.140-145.2004
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发表时间：
2004-01-01
影响因子：
9.4
通讯作者：
Asano, Y
中科院分区：
文献类型：
--
作者：
Ihira, M;Yoshikawa, T;Asano, Y
一种新的核酸扩增方法，称为环介导等温扩增（LAMP），可在等温条件下以高特异性、高效和快速的方式扩增 DNA，可能是快速检测传染源的有价值的工具。 LAMP 是针对人类疱疹病毒 6 (HHV-6) 开发的，本研究评估了其可靠性。尽管在 HHV-6 B 和 HHV-6 A DNA 中检测到了 LAMP 产物，但在 HHV-7 和人巨细胞病毒 DNA 中未检测到 LAMP 产物。原始 HRV-6 LAMP 方案的灵敏度为 50 份/管。为了提高方法的灵敏度，通过增加引物浓度对HHV-6 LAMP进行了修饰。修改后，灵敏度增加至 25 个拷贝/管。在这些初步验证研究之后，通过病毒分离、血清学分析和 HHV-6 LAMP 对 13 名发烧患者进行了 HHV-6 检测。在 8 例原发性 HHV-6 感染患者中，有 3 例在急性期和恢复期均通过原始 HRV-6 LAMP 方案在全血中检测到了 HHV-6 DNA。在急性期收集的所有八份血浆样本中，通过改良的 HRV-6 LAMP 检测到 HHV-6 DNA；然而，在恢复期收集的血浆样本中未检测到 HHV-6 DNA。尽管在过去感染过 HRV-6 的患者的全血样本的急性期和恢复期均检测到了 HHV-6 DNA，但在不含潜伏病毒 DNA 的血浆样本中未检测到 HHV-6 DNA。因此，使用这种改进的 HHV-6 LAMP 方案检测血浆中的 HHV-6 DNA 适用于诊断活动性 HHV-6 感染。
A novel nucleic acid amplification method, termed loop-mediated isothermal amplification (LAMP), which amplifies DNA with high specificity, efficiency, and rapidity under isothermal conditions, may be a valuable tool for the rapid detection of infectious agents. LAMP was developed for human herpesvirus 6 (HHV-6), and its reliability was evaluated in this study. Although LAMP products were detected in HHV-6 B and HHV-6 A DNA, they were not detected in HHV-7 and human cytomegalovirus DNA. The sensitivity of the original HRV-6 LAMP protocol was 50 copies/tube. In order to increase the method's sensitivity, HHV-6 LAMP was modified by increasing the primer concentration. As a result of the modification, sensitivity increased to 25 copies/tube. After these initial validation studies, 13 patients with fever were tested for HHV-6 by viral isolation, serological analysis, and HHV-6 LAMP. In three of the eight patients with primary HHV-6 infection, HHV-6 DNA was detected in whole blood by the original HRV-6 LAMP protocol in not only the acute phase but also the convalescent phase. HHV-6 DNA was detected by modified HRV-6 LAMP in all eight plasma samples collected in the acute phase; however, no HHV-6 DNA was detected in plasma samples collected in the convalescent phase. Although HHV-6 DNA was detected in both the acute and convalescent phases of whole-blood samples in patients with past HRV-6 infection, it was not detected in plasma samples that did not contain latent viral DNA. Thus, detection of HHV-6 DNA in plasma by using this modified HHV-6 LAMP protocol is appropriate for diagnosis of active HHV-6 infection.