可视化环介导恒温扩增法检测蔬菜产地环境灌溉水源中沙门氏菌

Determination of Salmonella in irrigated water of vegetable producing environment by visual loop-mediated isothermal amplification method

中文关键词:  环介导等温扩增  灌溉水源  沙门氏菌

英文关键词:loop-mediated isothermal amplification  irrigated water  Salmonella

中文摘要:

      目的 建立可视化环介导恒温扩增体系(loop-mediated isothermal amplification, LAMP)检测灌溉水源中沙门氏菌的分析方法。方法 以沙门氏菌侵袭蛋白A (invA)基因序列设计特异性LAMP检测引物, 优化LAMP反应体系和反应条件后, 通过特异性实验、灵敏度实验对LAMP检测方法进行测试, 并与国标检测方法对比检测人工污染的灌溉水源样品, 验证该方法的准确性。结果 灵敏度实验结果显示本LAMP检测方法最低检出限为7 CFU/25 μL, 特异性实验结果显示本LAMP检测方法具有良好的特异性, 干扰菌株对本LAMP检测方法无影响。该方法在与国家标准检测方法对比, 检测人工污染的灌溉水源样品时具有相同的准确性, 检测灵敏度为1×102 CFU/mL。结论 本研究建立了快速、准确、灵敏的恒温扩增体系, 可以应用于灌溉水源中沙门氏菌的快速检测。

英文摘要:

      Objective To establish a method for the detection of Salmonella in irrigated water by visual loop mediated isothermal amplification (LAMP). Methods One group of specific LAMP primers were designed based on the sequences of invasion protein A (invA) gene of Salmonella. After the LAMP reaction system and reaction conditions were optimized, the LAMP method for Salmonella detection was tested by specificity experiment and sensitivity test and verified by comparing with the national standard detection method through Salmonella detection in artificially contaminated irrigation water. Results The results of sensitivity test showed that the minimum limit of detection of the LAMP detection method was 7 CFU/25 μL, the results of specificity test showed that the LAMP method had good specificity, and the genomic nucleic acid of interference strains had no effect on the LAMP method. Compared with the national standard method, the LAMP method had the same accuracy for Salmonella detection in artificially contaminated irrigation water, and the detection sensitivity for Salmonella was 1×102 CFU/mL. Conclusion This study establishes a rapid, reliable and sensitive constant temperature amplification system, which can be applied to the rapid detection of Salmonella in irrigated water.