Position: Home > Articles > Detection of Listeria monocytogenes in Poultry and Livestock Meat by Propidium Monoazide-Hydroxynaphthol Blue-Loop-Mediated Isothermal Amplification Method
MEAT RESEARCH
2020
(5)
48-52
叠氮溴化丙锭-羟基萘酚蓝-环介导等温扩增法检测畜禽肉中单核细胞李斯特菌
作 者：
戴小芳;陈琼;董华夏;向春燕;虞伟明;郦娟
单 位：
武汉食品化妆品检验所
关键词：
单核细胞李斯特菌;叠氮溴化丙锭;羟基萘酚蓝;环介导等温扩增;畜禽肉
摘 要：
将叠氮溴化丙锭(propidium monoazide,PMA)、羟基萘酚蓝(hydroxynaphthol blue,HNB)与环介导等温扩增(loop-mediated isothermal amplification,LAMP)技术相结合,建立一种可视化PMA-HNB-LAMP方法快速检测畜禽肉中单核细胞李斯特菌(Listeria monocytogenes)。结果表明,当PMA终质量浓度为5.0μg/mL、孵育和曝光时间均为15 min时,可有效抑制10~5 CFU/m L的单核细胞李斯特菌死菌DNA的聚合酶链式反应扩增,而对活菌DNA的扩增不具有抑制作用。LAMP反应体系中HNB终浓度为210μmol/L时,检测显色结果肉眼可见。该方法的纯菌液灵敏度为2.4×10~2 CFU/m L,人工污染肉制品的检出限为2.3×10~4 CFU/m L。分别采用国标法、PMA-HNB-LAMP法和HNB-LAMP法检测20份市售畜禽肉样,3种方法的单核细胞李斯特菌检出率分别为5%、10%和20%。PMA-HNB-LAMP法在一定程度上可解决LAMP法的假阳性问题,并且具有操作简单、可视性强的优点。
译 名：
Detection of Listeria monocytogenes in Poultry and Livestock Meat by Propidium Monoazide-Hydroxynaphthol Blue-Loop-Mediated Isothermal Amplification Method
作 者：
DAI Xiaofang;CHEN Qiong;DONG Huaxia;XIANG Chunyan;YU Weiming;LI Juan;Wuhan Institute for Food and Cosmetic Control;
关键词：
Listeria monocytogenes;;propidium monoazide;;hydroxynaphthol blue;;loop-mediated isothermal ampli?cation;;poultry and livestock meat
摘 要：
Using propidium monoazide(PMA) treatment and hydroxynaphthol blue(HNB) staining combined with loopmediated isothermal amplification, a rapid and visual detection method(PMA-HNB-LAMP) for Listeria monocytogenes was established in this study. The optimization of experimental parameters showed that a final PMA concentration of 5.0 μg/m L, incubation time of 15 min and exposure time of 15 min could restrain DNA amplification of 10~5 CFU/m L dead cells but not of live cells. A final HNB concentration of 210 μmol/L could give results visible to the naked eyes. The sensitivity of the PMAHNB-LAMP method was 2.4 × 10~2 CFU/m L for pure bacterial culture. The detection limit for artificially contaminated meat was 2.3 × 10~4 CFU/m L. Using the national standard method, PMA-HNB-LAMP and HNB-LAMP, the detection rates of L. monocytogenes in 20 meat products available on the market were 5%, 10% and 20%, respectively. PMA-HNB-LAMP is superior to traditional LAMP in avoiding false-positive results to a certain extent, and it has the advantages of simple operation and strong visibility.
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