非预包装食品中沙门氏菌快速检测方法的建立

Establishment of Rapid Detection Method for Salmonella in Non-prepackaged Foods

当前沙门氏菌传统的国家标准检测方法不能满足食品安全监管高效性的需求,尤其是针对非预包装食品这种较易腐败变质的食品。研究针对非预包装食品利用聚合酶链式反应(polymerase chain reaction,PCR)分子层析方法得出一个快速、简单、准确的沙门氏菌检测方法。通过对60批普通基质食品(面食制品、乳制品、肉制品)和40批复杂基质食品(凉菜类、生鲜蛋类)模拟不同程度被沙门氏菌污染的实验,采用国标法和PCR分子层析法对比检测。结果显示:普通基质食品PCR分子层析法检测灵敏度更高,可达1 CFU/mL^10 CFU/mL的阳性菌添加浓度;复杂基质食品尤其对生鲜蛋类食品,两种检测方法的灵敏度均较低。故对沙门氏菌的增菌液进行了优化,在沙门氏菌的增菌液缓冲蛋白胨水(buffered peptone water,BPW)中添加一定比例六水氯化镁,即利用提高增菌液渗透压和降低pH值抑制其他肠道菌群的生长的原理,使其检测灵敏度提高,由104 CFU/mL的阳性菌添加浓度降低至102 CFU/mL的添加浓度,灵敏度提高了100倍;经特异性试验验证,PCR分子层析法具有较强的抗干扰能力,样本体系中存在其他干扰菌株的脱氧核糖核酸(deoxyribonucleic acid,DNA)亦不会影响沙门氏菌的检测。PCR分子层析法检测时间只需22 h,对比需要6 d检测时间的传统国标方法的检测效率提高6倍。

Due to the needs of high efficiency of food safety supervision and management could not be met by the traditional national standard detection methods of Salmonella,especially for non-prepackaged foods which are more prone to spoilage.A rapid,accurate,simple and efficient Salmonella polymerase chain reaction(PCR)molecular chromatography detection method was established for non-prepackaged foods.The national standard method and PCR molecular chromatograph are used to detect the different degrees of Salmonella contamination in 60 batches of common substrate foods(pasta products,dairy products,and meat products)and 40 batches of complex substrate foods(cold dishes and fresh eggs).The results showed that the detection sensitivity of PCR molecular chromatography was higher for common substrate foods.And the minimal detectable concentration of positive bacteria could reach 1 CFU/mL-10 CFU/mL.The complex substrate such as fresh food and eggs were less sensitive to both detection methods.Therefore,MgCl·6H2O was added to optimize the enrichment microbial solution buffered peptone water(BPW)of Salmonella to improve the sensitivity by using the principle of increasing the osmotic pressure of enrichment solution and reducing the pH value to inhibit the growth of other interfering intestinal flora.The minimal detectable concentration of positive bacteria 104 CFU/mL was reduced to 102 CFU/mL and the detection sensitivity was improved by 100 times.In addition,verified by the specific experiment that PCR molecular chromatography had strong anti-interference ability.The detection of Salmonella could not be affected by other interfering strains deoxyribonucleic acid(DNA)existing in the system.The detection time of was only 22 h for the PCR molecular chromatography,which was greatly shorter than that(6 d)of the national standard method.The detection efficiency was increased by 6 times.