Ihira Masaru, Yoshikawa Tetsushi, Enomoto Yoshihiko, Akimoto Shiho, Ohashi Masahiro, Suga Sadao, Nishimura Naoko, Ozaki Takao, Nishiyama Yukihiro, Notomi Tsugunori, Ohta Yoshinori, Asano Yoshizo
Department of Medical Information Technology, Fujita Health University College, Konan, Aichi, Japan.
J Clin Microbiol. 2004 Jan;42(1):140-5. doi: 10.1128/JCM.42.1.140-145.2004.
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 HHV-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 HHV-6 LAMP protocol in not only the acute phase but also the convalescent phase. HHV-6 DNA was detected by modified HHV-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 HHV-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.
一种名为环介导等温扩增（LAMP）的新型核酸扩增方法，可在等温条件下高效、快速且特异性地扩增DNA，可能是快速检测感染因子的一种有价值的工具。LAMP最初是针对人类疱疹病毒6型（HHV-6）研发的，本研究对其可靠性进行了评估。虽然在HHV-6 B型和HHV-6 A型DNA中检测到了LAMP产物，但在HHV-7型和人巨细胞病毒DNA中未检测到。原始的HHV-6 LAMP方案的灵敏度为50拷贝/管。为提高该方法的灵敏度，通过增加引物浓度对HHV-6 LAMP进行了改进。改进后，灵敏度提高到了每管25拷贝。在这些初步验证研究之后，对13名发热患者进行了病毒分离、血清学分析和HHV-6 LAMP检测以筛查HHV-6。在8例原发性HHV-6感染患者中的3例中，不仅在急性期而且在恢复期，通过原始的HHV-6 LAMP方案在全血中检测到了HHV-6 DNA。通过改进的HHV-6 LAMP在急性期采集的所有8份血浆样本中均检测到了HHV-6 DNA；然而，在恢复期采集的血浆样本中未检测到HHV-6 DNA。虽然在既往有HHV-6感染的患者的全血样本的急性期和恢复期均检测到了HHV-6 DNA，但在不含有潜伏病毒DNA的血浆样本中未检测到。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。
