多重荧光PCR检测婴幼儿食品中常见病原菌

Establishment of a multiple real-time PCR method for detection of pathogens in infant food

目的建立婴幼儿食品中3种常见病原菌的多重荧光PCR检测方法。方法根据阪崎杆菌16S-23SrDNA保守区、金黄色葡萄球菌耐热核酸酶(nuc)基因序列和蜡样芽胞杆菌cerA特异基因,设计合成引物和探针,优化反应条件。结果建立的多重荧光PCR方法只特异性地扩增目标病原菌;用国标法和多重荧光PCR方法同时对194份奶粉和米粉样品进行检测,结果完全一致。多重荧光PCR方法检测过程可在8h内完成。结论建立的多重荧光PCR法敏感性高、特异性强、快速、准确,可同时检测婴幼儿食品中的阪崎杆菌、金黄色葡萄球菌和蜡样芽胞杆菌,适合批量样本检测,有很好的应用前景。

Objective To establish a multiple real-time PCR method for detection of three kinds of pathogens in infant food.Methods Primers and probes were designed based on the conservative sequence of 16S-23SrDNA of Enterobacter sakazakii and nuc gene of Staphylococcus aureus and cerA gene of Bacillus cereus,and the reaction conditions of multiple PCR were optimized.Results Enterobacter sakazakii and Staphylococcus aureus and Bacillus cereus generated fluorescent signals only.The infant food samples were detected by real-time PCR and GB4789method.The results of Guobiao and PCR method for determining 194milk and rice flour samples were coincidental.The detection could be finished in 8hours.Conclusion The detection of real-time PCR method was of sensitivity and specificity,and can be applied to rapid diagnosis of Enterobacter sakazakii,Staphylococcus aureus and Bacillus cereus in infant food and can be applied to detection for bulk samples.