Ministry of Food and Drug Safety, 187, Osongsaengmyeong 2-ro, Cheongju, Chungbuk 28159, Republic of Korea.
Animal and Plant Quarantine Agency, 177, Hyeoksin 8-ro, Gimcheon, Gyeongbuk 39660, Republic of Korea.
J Food Prot. 2022 Nov 1;85(11):1531-1537. doi: 10.4315/JFP-22-117.
Listeria monocytogenes is a common foodborne pathogen affecting public health. Thus, detecting L. monocytogenes, even at low levels, in food matrices is essential. However, the current culture methods used for its detection and quantification are time consuming and difficult owing to background flora and interference by food matrices. DNA-based assays depend on DNA extraction and purification techniques. No optimal DNA extraction kit has been developed for analyzing L. monocytogenes in dairy products by real-time quantitative PCR (RT-qPCR). Therefore, in this study, we aimed to determine the efficiency of three DNA extraction kits for detecting L. monocytogenes in dairy products by RT-qPCR. We tested the efficiency of three commercial kits for DNA extraction from L. monocytogenes artificially inoculated in milk and dairy products. For the PrepSEQ rapid spin sample preparation kit and Exgene Cell SV mini, the limit of detection of was 100, 100, and 101 CFU/mL L. monocytogenes in milk, processed cheese, and infant formula, respectively, whereas that of the QIAamp DNA mini kit was 101, 103, and 102 CFU/mL, respectively. In addition, the Exgene Cell SV mini was better than the PrepSEQ rapid spin sample preparation kit for obtaining a standard curve for RT-qPCR of L. monocytogenes DNA in milk and dairy products, with a high correlation coefficient and amplification efficiency. The results of this study may be valuable for diagnostic laboratories and for developing an effective extraction method for processing food samples, such as dairy products, to subsequently detect and quantify L. monocytogenes by RT-qPCR.
单增李斯特菌是一种常见的食源性致病菌，影响公共健康。因此，检测食品基质中的单增李斯特菌（即使是低水平的）是至关重要的。然而，目前用于其检测和定量的培养方法由于背景菌群和食品基质的干扰而耗时且困难。DNA 检测方法依赖于 DNA 提取和纯化技术。目前还没有开发出用于乳制品中实时定量 PCR（RT-qPCR）分析单增李斯特菌的最佳 DNA 提取试剂盒。因此，在本研究中，我们旨在确定三种 DNA 提取试剂盒用于通过 RT-qPCR 检测乳制品中单增李斯特菌的效率。我们测试了三种商业试剂盒从人工接种于牛奶和乳制品中的单增李斯特菌中提取 DNA 的效率。对于 PrepSEQ 快速旋转样品制备试剂盒和 Exgene Cell SV mini，单增李斯特菌在牛奶、加工奶酪和婴儿配方奶粉中的检测限分别为 100、100 和 101 CFU/mL，而 QIAamp DNA mini 试剂盒的检测限分别为 101、103 和 102 CFU/mL。此外，对于从牛奶和乳制品中 RT-qPCR 检测单增李斯特菌 DNA 的标准曲线，Exgene Cell SV mini 比 PrepSEQ 快速旋转样品制备试剂盒获得更好，具有更高的相关系数和扩增效率。本研究的结果对于诊断实验室和开发有效的食品处理样品（如乳制品）提取方法可能具有重要价值，以便随后通过 RT-qPCR 检测和定量单增李斯特菌。
