Real-time loop-mediated isothermal amplification of propidium monoazide for visualization of viable Bacillus cereus in food

IF 3 3区农林科学 Q2 FOOD SCIENCE & TECHNOLOGY

European Food Research and Technology Pub Date : 2024-08-16 DOI:10.1007/s00217-024-04592-y

Xiaoting Song, Zuwei Wang, Zhaoxin Lu, Xiaomei Bie

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引用次数: 0

Abstract

Bacillus cereus is a common contaminating bacteria and conditioned pathogens in food. Immunological methods or molecular biology assays that are currently available cannot distinguish between live and dead bacterial cells, which may overestimate the number of bacteria and lead to false-positive results. To solve this problem, we introduced propidium monoazide (PMA) and calcein in this study and established a real-time visual loop-mediated isothermal amplification (LAMP) detection method to detect viable B. cereus. In addition, the results can be detected visually with calcein dye or quantitatively by monitoring the amplification curve of the fluorescence signal. In this study, the results showed that the visual PMA-LAMP method had high specificity, strong anti-interference ability, a sensitivity of 1.16 × 10² CFU/mL, and that the whole reaction could be completed within 80 min. This method not only has high sensitivity, but also has a shorter reaction time and is faster. In the detection of artificially contaminated food samples, the lowest detection limit reached 6 CFU/mL after 3 or 4 h of enrichment. Therefore, the visual PMA-LAMP technology can provide an efficient and rapid method for the detection of live B. cereus bacteria.

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利用单氮化丙啶的环介导实时等温扩增技术检测食品中的蜡样芽孢杆菌存活率

蜡样芽孢杆菌是食品中常见的污染菌和条件致病菌。目前可用的免疫学方法或分子生物学检测方法无法区分活的和死的细菌细胞，可能会高估细菌数量，导致假阳性结果。为了解决这个问题，我们在本研究中引入了单氮化丙啶（PMA）和钙蓝蛋白，并建立了一种实时可视环介导等温扩增（LAMP）检测方法来检测存活的蜡样芽孢杆菌。此外，还可通过钙黄绿素染料直观检测结果，或通过监测荧光信号的放大曲线定量检测结果。研究结果表明，可视 PMA-LAMP 法特异性高，抗干扰能力强，灵敏度为 1.16 × 102 CFU/mL，整个反应可在 80 分钟内完成。该方法不仅灵敏度高，而且反应时间短、速度快。在检测人工污染的食品样品时，经过 3 或 4 小时的富集，最低检测限达到 6 CFU/mL。因此，可视化 PMA-LAMP 技术可为检测活的蜡样芽孢杆菌提供一种高效、快速的方法。

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来源期刊

European Food Research and Technology 工程技术-食品科技

CiteScore

6.60

自引率

3.00%

发文量

232

审稿时长

2.0 months

期刊介绍： The journal European Food Research and Technology publishes state-of-the-art research papers and review articles on fundamental and applied food research. The journal''s mission is the fast publication of high quality papers on front-line research, newest techniques and on developing trends in the following sections: -chemistry and biochemistry- technology and molecular biotechnology- nutritional chemistry and toxicology- analytical and sensory methodologies- food physics. Out of the scope of the journal are: - contributions which are not of international interest or do not have a substantial impact on food sciences, - submissions which comprise merely data collections, based on the use of routine analytical or bacteriological methods, - contributions reporting biological or functional effects without profound chemical and/or physical structure characterization of the compound(s) under research.