建立基于多重聚合酶链式反应(polymerase chain reaction, PCR)和表面增强拉曼光谱(surface-enhanced Raman scattering, SERS)的沙门氏菌、大肠杆菌和金黄色葡萄球菌检测模型,优化检测方案,提高食源性病原菌检测效率。以病原菌hilA、uidA和clfA的基因序列设计特异性引物,优化多重PCR反应的扩增条件,检测多重PCR的特异性和灵敏性。通过SERS技术检测3种细菌并确定检测限,对3种病原菌进行主成分分析,然后对2种方法的检测限进行比较。结果显示,建立的多重PCR和SERS技术均可特异性地检测出沙门氏菌、大肠杆菌和金黄色葡萄球菌,三重PCR反应的最低检测限为104 CFU/mL,最低检出DNA量为50 pg/μL,而SERS检测沙门氏菌最低检出量为103 CFU/mL,大肠杆菌和金黄色葡萄球菌的最低检出量为104 CFU/mL。PCA分析的前2个主成分的累积贡献率达93.8%。与多重PCR相比,研究建立的SERS方法提高了沙门氏菌的检测限,敏感度更高,并且缩短了检测时间,对食品中食源性病原菌的监测起到指导作用。
The detection of common foodborne pathogens is an important prerequisite to ensure the safety of animal food. Consequently, developing effective methods for detecting common foodborne pathogens is necessary. In this study, surface-enhanced Raman scattering (SERS) method was applied to detect three common foodborne pathogens (Salmonella spp., Escherichia coli and Staphylococcus aureus) and compared with multiplex polymerase chain reaction (PCR) method. Specific primers were designed for multiplex PCR based on gene hilA, uidA and clfA, and followed by the amplification condition optimized and the specificity and sensitivity of the multiplex PCR system evaluated. Besides, the detection limits of the two methods were compared. The results showed that both the multiplex PCR and SERS methods could specifically detect the three foodborne pathogens. For the multiplex PCR, the minimum detection limit for microbes was 104 CFU/mL while for DNA was 50 pg per reaction. For SERS method, the detection limits for Salmonella sp., E. coli and S. aureus were 103 CFU/mL, 104 CFU/mL and 104 CFU/mL, respectively. In addition, the cumulative contribution rate of the first two principal components in the SERS method reached 93.8% according to the principal component analysis (PCA). In summary, the SERS method applied in this study revealed higher sensitivity for Salmonella sp., and shorter detection time than multiplex PCR. Therefore, the SERS method exhibited a good prospect in detecting foodborne pathogens.
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