Institute of Life Sciences & Resources and Department of Food Science and Biotechnology, Kyung Hee University, Yongin, 17104, South Korea.
Interdisciplinary Program in Biological and Chemical Engineering, Seoul National University, Seoul, 08826, South Korea.
Food Microbiol. 2023 Aug;113:104265. doi: 10.1016/j.fm.2023.104265. Epub 2023 Mar 10.
Droplet digital polymerase chain reaction (ddPCR) is an emerging molecular detection assay that provides an absolute quantification of targets. Despite its emerging applications in the detection of food microorganisms, there are limited reports of its use for the monitoring of microorganisms utilized as starters in the dairy industry. This study investigated the applicability of ddPCR as a detection platform for Lacticaseibacillus casei, a probiotic found in fermented foods and exerts beneficial effects on human health. In addition, this study compared the performance of ddPCR with that of real-time PCR. The ddPCR targeting the haloacid dehalogenase-like hydrolase (LBCZ_1793) exhibited high specificity against 102 nontarget bacteria, including Lacticaseibacillus species that is very closely related to L. casei. The ddPCR exhibited high linearity and efficiency within the quantitation range (10-10 CFU/ml), with the limit of detection being 10 CFU/ml. The ddPCR also demonstrated a higher sensitivity than real-time PCR in detecting low bacterial concentration in spiked milk samples. Furthermore, it provided an accurate absolute quantification of the concentration of L. casei, without the need for standard calibration curves. This study demonstrated that ddPCR is a useful method for monitoring starter cultures in dairy fermentations and detecting L. casei in foods.
微滴式数字聚合酶链式反应（ddPCR）是一种新兴的分子检测方法，可提供目标的绝对定量。尽管它在食品微生物检测中的应用不断涌现，但在监测用于乳制品行业的起始微生物方面，其应用的报道有限。本研究探讨了 ddPCR 作为一种检测工具的适用性，该检测工具针对的是发酵食品中存在的益生菌乳杆菌属，它对人类健康有有益影响。此外，本研究比较了 ddPCR 与实时 PCR 的性能。针对卤酸脱卤酶样水解酶（LBCZ_1793）的 ddPCR 对 102 种非目标细菌表现出高度特异性，包括与乳杆菌属非常密切相关的乳杆菌属。ddPCR 在定量范围内（10-10 CFU/ml）表现出高线性和高效率，检测限为 10 CFU/ml。与实时 PCR 相比，ddPCR 在检测添加到牛奶样品中的低浓度细菌时具有更高的灵敏度。此外，它无需标准校准曲线即可准确绝对定量乳杆菌属的浓度。本研究表明，ddPCR 是一种用于监测乳制品发酵中起始培养物和检测食品中乳杆菌属的有用方法。
