Kuboki Noritaka, Inoue Noboru, Sakurai Tatsuya, Di Cello Francescopaolo, Grab Dennis J, Suzuki Hiroshi, Sugimoto Chihiro, Igarashi Ikuo
National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Obihiro, Hokkaido 080-8555, Japan.
J Clin Microbiol. 2003 Dec;41(12):5517-24. doi: 10.1128/JCM.41.12.5517-5524.2003.
While PCR is a method of choice for the detection of African trypanosomes in both humans and animals, the expense of this method negates its use as a diagnostic method for the detection of endemic trypanosomiasis in African countries. The loop-mediated isothermal amplification (LAMP) reaction is a method that amplifies DNA with high specificity, efficiency, and rapidity under isothermal conditions with only simple incubators. An added advantage of LAMP over PCR-based methods is that DNA amplification can be monitored spectrophotometrically and/or with the naked eye without the use of dyes. Here we report our conditions for a highly sensitive, specific, and easy diagnostic assay based on LAMP technology for the detection of parasites in the Trypanosoma brucei group (including T. brucei brucei, T. brucei gambiense, T. brucei rhodesiense, and T. evansi) and T. congolense. We show that the sensitivity of the LAMP-based method for detection of trypanosomes in vitro is up to 100 times higher than that of PCR-based methods. In vivo studies in mice infected with human-infective T. brucei gambiense further highlight the potential clinical importance of LAMP as a diagnostic tool for the identification of African trypanosomiasis.
虽然聚合酶链反应（PCR）是检测人和动物体内非洲锥虫的首选方法，但该方法的成本使其无法作为非洲国家地方性锥虫病的诊断方法。环介导等温扩增（LAMP）反应是一种在等温条件下仅使用简单培养箱就能以高特异性、高效率和快速性扩增DNA的方法。与基于PCR的方法相比，LAMP的一个额外优势是DNA扩增可以通过分光光度法和/或肉眼进行监测，无需使用染料。在此，我们报告了基于LAMP技术的一种高度灵敏、特异且简便的诊断检测方法的条件，用于检测布氏锥虫属（包括布氏布氏锥虫、布氏冈比亚锥虫、布氏罗德西亚锥虫和伊氏锥虫）和刚果锥虫中的寄生虫。我们表明，基于LAMP的体外检测锥虫方法的灵敏度比基于PCR的方法高多达100倍。对感染人感染性布氏冈比亚锥虫的小鼠进行的体内研究进一步凸显了LAMP作为非洲锥虫病诊断工具的潜在临床重要性。
