Rapid and Sensitive Detection of Listeria monocytogenes by Loop-Mediated Isothermal Amplification
Rapid and Sensitive Detection of Listeria monocytogenes by Loop-Mediated Isothermal Amplification
复制标题
DOI：
10.1007/s00284-011-0013-3
复制
发表时间：
2011-12-01
影响因子：
2.6
通讯作者：
Gao, Yu-Shi
中科院分区：
文献类型：
--
作者：
Tang, Meng-Jun;Zhou, Sheng;Gao, Yu-Shi
环介导等温扩增 (LAMP) 设计用于检测单核细胞增生李斯特氏菌，这是一种重要的食源性致病菌，可引起人类和动物疾病。 hlyA 基因的引物组由针对特定基因上的八个区域的六个引物组成。 LAMP 测定可以在 65 摄氏度水浴中在 40 分钟内完成。通过钙黄绿素和锰离子以及琼脂糖凝胶电泳使扩增产物可视化。用于检测纯培养物中单核细胞增生李斯特菌的 LAMP 测定的灵敏度为每个反应 2.0 CFU。 LAMP 检测的灵敏度比传统 PCR 检测高 100 倍。通过这种方法，对 60 个鸡肉样本进行了单核细胞增生李斯特菌的检测。与“金标准”培养生物技术相比，LAMP 的准确性为 100%，而 PCR 检测未能检测到两个阳性样本中的单核细胞增生李斯特菌。结果表明，LAMP检测可作为一种灵敏、快速、简单的单核细胞增生李斯特菌检测工具，有助于食品中单核细胞增生李斯特菌污染的监测。
Loop-mediated isothermal amplification (LAMP) was designed for detection of Listeria monocytogenes, which is an important food-borne kind of pathogenic bacteria causing human and animal disease. The primers set for the hlyA gene consist of six primers targeting eight regions on specific gene. The LAMP assay could be performed within 40 min at 65 degrees C in a water bath. Amplification products were visualized by calcein and manganous ion and agarose gel electrophoresis. Sensitivity of the LAMP assay for detection of L. monocytogenes in pure cultures was 2.0 CFU per reaction. The LAMP assay was 100-fold higher sensitive than that of the conventional PCR assay. Taking this way, 60 chicken samples were investigated for L. monocytogenes. The accuracy of LAMP was shown to be 100% when compared to the "gold standard" culture-biotechnical, while the PCR assay failed to detect L. monocytogenes in two of the positive samples. It is shown that LAMP assay can be used as a sensitive, rapid, and simple detection tool for the detection of L. monocytogenes and will facilitate the surveillance for contamination of L. monocytogenes in food.