多重实时荧光定量PCR快速检测婴幼儿奶粉中沙门氏菌、克罗诺杆菌和金黄色葡萄球菌

Simultaneous detection of Salmonella spp., Staphylococcus aureus and Cronobacter in infant milk powder by multiplex quantitative real-time PCR

中文关键词:  多重实时荧光PCR  奶粉  金黄色葡萄球菌  沙门氏菌  克罗诺杆菌

英文关键词:multiplex quantitative real-time PCR  milk powder  Salmonella spp.  Staphylococcus aureus  Cronobacter

中文摘要:

      目的 建立多重实时荧光定量PCR(multiplex quantitative real-time PCR, multiplex qPCR)快速检测奶粉中金黄色葡萄球菌、沙门氏菌和克罗诺杆菌3种常见致病菌的方法。方法 筛选目标菌株的特异性引物与探针, 优化反应体系, 建立稳定的多重qPCR反应体系。通过阳性菌株加标的方式验证体系的特异性, 并确定人工污染奶粉的检出限。结果 各对引物探针对目标菌株均能扩增, 多重实时荧光PCR未发现交叉反应, 对17株非目标菌进行检测均未检出, 人工污染奶粉中克罗诺杆菌和沙门氏菌的检出限均为103 CFU/mL, 金黄色葡萄球菌的检出限为104 CFU/mL。结论 本研究方法可实现婴幼儿奶粉样品中3种致病菌qPCR高效率检测。

英文摘要:

      Objective To establish a method for simultaneous detection of Salmonella spp., Staphylococcus aureus and Cronobacter in milk powder by multiplex quantitative real-time PCR (multiplex qPCR). Methods The specific primers and probes were selected at first. After optimization of reaction conditions, a multiplex qPCR system was established, and the specificity and limits of detection were further verified. Results All target strains were detected. In contrast, none of the 17 non-target strains were detected. The limits of detection of multiplex qPCR in milk power were 103 CFU/mL for Salmonella spp. and Cronobacter, and 104 CFU/mL for Staphylococcus aureus. Conclusion The established multiplex PCR method is accurate, fast and effective, which meets the satisfaction of detection of Salmonella spp., Staphylococcus aureus and Cronobacter in infant milk powder.