多重PCR方法快速检测3种消化道病原菌

Development of a multiplex PCR for simultaneous detection of three conditioned pathogen bacteria of alimentary tract diseases

为建立一种检测和鉴别诊断动物奇异变形杆菌、沙门氏菌和产气荚膜梭菌感染的方法,本研究针对奇异变形杆菌脲酶调节子基因(ureR)、沙门氏菌侵袭蛋白A基因(invA)及产气荚膜梭菌α毒素基因(α-toxin)序列设计引物,建立一种同时检测3种细菌的多重PCR方法,并对扩增体系和条件进行了优化,建立的多重PCR检测方法特异性良好,对3种目的菌最低检出限均在105cfu/mL以上。初步应用该方法检测3种细菌单独人工感染小鼠的病变组织和人工模拟细菌混合感染的临床样品,结果证明该方法可以用于临床样本检测。

Proteus mirabilis, Salmonella and Clostfidium perfringens are major conditioned pathogens causing diarrhea in animals. To simultaneous detection of three bacteria, the multiplex PCR method was established with three pairs of primers designed according to the corresponding sequences of the ureR, invA and a-toxin, respectively. The method had high specificity and sensitivity with a detection limit of 10s cfu/mL. Furthermore, the detections of tissue samples ~om artificially infected mice or artificially contaminated tissue samples proved that the multiplex PCR was specific and sensitive. Our data demonstrated the detection method was applicable for the differential diagnosis of P.mirabilis, Salmonella and C.perfringens infections.