免疫磁珠富集-实时荧光PCR检测生畜肉中大肠埃希菌O157∶H7的实验研究

Immune magnetic beads enrichment-real-time fluorescent PCR detection of meat in e coli O157∶H7 experimental research

目的建立免疫磁珠富集（immunomagnetic separation,IMS）-实时荧光PCR（q PCR）技术快速检测肠出血性大肠埃希菌O157∶H7的方法。方法根据肠出血性大肠埃希菌O157∶H7抗原特异性基因rfb EO157设计引物和Taq Man探针,建立q PCR检测体系;对样品中的O157∶H7大肠埃希菌用出血性大肠O157抗体标记免疫磁珠进行特异性捕获,再利用q PCR技术对磁珠吸附的菌株进行检测。结果采用IMS-q PCR方法检测,当25 g样本中菌量为10^0cfu,样品被1∶2稀释时,检测结果呈阳性,检测全过程需时2-3 h。结论 IMS-q PCR检测O157∶H7大肠埃希菌的方法具有灵敏度高、特异性强、快速等优点,可用于大肠埃希菌O157∶H7食物中毒的快速诊断和食品微生物检测,为食源性疾病的分子流行病学调查提供新的检测手段。

Objective Immune magnetic beads enrichment - real - time fluorescent PCR technology rapid detection of enterohem- orrhagie e coli O157： H7. Methods According to enterohemorrhagie e coli O157：H7 antigen specific gene rfbEO157 design primers and TaqMan probe, establishing qPCR detection system; For e coli O157：H7 in his sample, with hemorrhagic mark immune mag- netic beads of e coli O157 antibody specificity to capture, recycle qPCR technology of magnetic beads adsorption tested strains. Results The IMS -qPCR method detection, when 25 g sample for 10^0 cfu, bacteria content in the samples was 1：2 dilu- tion, positive test results, test process takes about 2 - 3 h. Conclusion The IMS - qPCR detection method of e coli O157：H7 has the advantages of high sensitivity, strong specificity, fast, and can be used for e coli O157：H7 rapid diagnosis of food poisoning and food microbe testing, to investigate the molecular epidemiology of foodborne disease provide new detection means.