Vancomycin-functionalized magnetic bead-qPCR platform for rapid differentiation of viable and dead Listeria monocytogenes in food samples

文献类型： 外文期刊

第一作者： Yuan, Weijie

作者： Yuan, Weijie;Wang, Yuzhu;Li, Yansong;Ren, Honglin;Zhang, Bo;Shi, Ruoran;Li, Chengwei;Hou, Jiaqi;Hu, Shaohui;Hu, Xueyu;Lu, Shiying;Hu, Pan;Yuan, Weijie;Wang, Yuzhu;Zhang, Runze;Li, Yansong;Ren, Honglin;Zhang, Bo;Shi, Ruoran;Li, Chengwei;Hou, Jiaqi;Hu, Shaohui;Hu, Xueyu;Lu, Shiying;Wang, Xiaoxu;Hu, Pan;Wang, Xiaoxu;Zhang, Runze

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关键词： Listeria monocytogenes; Vancomycin-functionalized magnetic beads; Real-time quantitative PCR (qPCR); Live/dead discrimination; Food safety

期刊名称：MICROCHEMICAL JOURNAL （ 影响因子：5.1； 五年影响因子：4.7 ）

ISSN： 0026-265X

年卷期： 2025 年 216 卷

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收录情况： SCI

摘要： The present study reports a dual-functional platform that couples vancomycin-functionalized magnetic beads (VMBs) with real-time quantitative polymerase chain reaction (qPCR) to achieve rapid, culture-free discrimination and quantification of live and dead Listeria monocytogenes in complex food matrices. The exploitation of vancomycin's high-affinity binding to the D-Ala-D-Ala motif of intact peptidoglycan enables the selective capture and removal of viable cells, thereby leaving only DNA from non-viable bacteria for SYBR Green qPCR analysis. Optimisation of bead coupling and adsorption kinetics, in conjunction with a pre-calibrated live/dead correction algorithm, yields an LOD of 102 CFU/mL over a 101-107 CFU/mL linear range and completes within 12 h-a process that is significantly faster than conventional culture methods. The validation of the method in beef, salad, cheese, and salmon matrices has yielded recoveries ranging from 96.4 to 99.2 %, with RSDs consistently below 3.6 %. Stability tests have further confirmed that dead-cell DNA remains amplifiable for a minimum of one week, and non-specific capture remains below 5 %. In contrast to isothermal or immunomagnetic approaches, the VMB-qPCR system offers a unique capability to provide unambiguous live/dead resolution without the use of photoactive dyes or extended enrichment, thereby providing a robust, highthroughput tool for food safety surveillance and clinical pathogen monitoring.

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