鼠伤寒沙门氏菌多重PCR检测方法的研究

Studies on Detection of Salmonella typhimurium in Food by Multiplex Polymerase Chain Methods

分别根据沙门氏菌16S rRNA、质粒毒力基因spvC、致病基因invB、fimA序列设计4对引物,对沙门氏菌株及非沙门氏株菌基因组DNA进行多重PCR检测。结果该方法能检测出6.3×102个cfu/ml纯培养的沙门氏菌,人工染菌食品模拟检测结果显示,熟鸡肉初始含菌量为17cfu/g、全脂奶粉为11cfu/g、生牛肉为13.6cfu/g,经过8h增菌,PCR检测为阳性。该体系能鉴定产生多种毒力因子的沙门氏菌,特异性强、敏感性高,为检测和鉴定沙门氏菌株提供了一个新方法。

Four sets of primers were designed according to 16S rRNA,spvC,invB,fimA genes of Salmonella typhimurium,which were used to amplify genomic DNA of Salmonella and non Salmonellas by multiplex polymerase chain reaction（m-PCR）.Specificity results showed that only Salmonella strains had specifically amplified the target fragments,but none others.Sensitivity of m-PCR assay for pure cell cultures of Salmonella typhimurium was 6.3×102cfu/ml.However the detection limits of this method for artificially contaminated cooked chicken,milk powder,and beef were 17cfu/g,11cfu/g,13.6cfu/g following 8h of enrichment,respectively.This multiplex PCR assay successfully identifed main virulence genes of Salmonella typhimurium and may be applied to rapid and sensitive detection of Salmonella typhimurium in food when combined with an enrichement step.