食品中沙门氏菌PCR快速检测方法的建立

Development of polymerase chain methods for Salmonella detection in foods

目的寻求快速检测食品中沙门氏菌的技术方法支持。方法根据沙门氏菌编码侵袭蛋白E(invasion protean E,invE)的基因设计一对引物,对沙门氏菌株及非沙门氏株菌基因组DNA进行PCR扩增,建立了沙门氏菌特异、敏感和快速的PCR检测方法,并对食品中的沙门氏菌进行了检测。结果该方法能检测出3.0×102cfu/mL纯培养的沙门氏菌,4种人工染菌食品的模拟检测结果显示,当各食品中初始含菌量分别为1.5cfu/g(火腿肠)、2.4cfu/g(鲜猪肉)、1cfu/ml(包装鲜奶)和1cfu/g(鲜鸡蛋)时,分别经过6h、18h、12h和18h增菌,PCR检测为阳性,而阴性对照组均为阴性。结论方法具有耗时少,特异性强,敏感性高,操作简便,费用低的优点。

A pair of primers was designed according to the published nucleotide sequence of inv E gene of Salmonella typhimurium, which specifically amplified an inv E segment of 463bp. Polymerase chain reaction （PCR） assay for detection of nucleotide of Salmonella was established by optimizing the reaction condition and the concentrations of each component. Specificity results showed that all the 7 Salmonella bacterial strains had specifically amplified the purpose fragment, but 10 other non-Salmonella bacterial strains submitted negative reactions. Sensitivity of PCR assay for pure cell culture of Salmonella typhimurium was 3.0 10^2cfu per ml pure cell culture. However, the detection limits of this method for artificially contaminated ham, fresh meat, milk and eggs were 1.5cfu/g, 2.4cfu/g, 1cfu/ml and 1cfu/g following 6h, 18h, 12h and 18h of enrichment respectively, while there was no amplification from the negative control. Results showed that a easy, specific and sensitive PCR method was established. This method could detect Salmonella in a short period and be used for Salmonella detection in foods.