The Establishment And Application Of The Multiplex PCR To Detect Eight Different Serotype Of Enterohemorrhagic Escherichia Coli

Enterohemorrhagic Escherichia coli(EHEC)is transmitted to human mainly by ingestion of food consumption,can cause severe foodborne diseases,and infectious diseases epidemics and outbreaks caused have been reported in several countries.EHEC O157:H7 was classified as an new-emerging foodborne disease pathogen in 1997 by the world health organization(WHO).EHEC O157:H7 is the most important serogroup of EHEC associated with public health and food safety,EHEC of other serogroups eg O26,O45,O103,O104,O111,O121 and O145 are also common.Therefore,these 7 serogroups of non-O157 EHEC and EHEC O157 have been studied in this subject.The mainly researches and results are given as below.Primers were designed for 11 specific genes of O26,O45,O103,O104,O111,O121 and O145 serogroups of non-O157 EHEC and EHEC O157,all amplification products of single PCR of 11 pairs of primers have shown clear target bands and none of impurity bands have been observed.4 sets of multiplex PCR systems for detection of EHEC in food have been developed through optimizing the reaction systems and conditions and was verifying the specificity of the primers,and the sensitivity of the methods was evalued too.Simultaneous and rapid detection methods with high sensitivity and high specification of for 8 kinds of serogroups of EHEC were realiaed by this reserch,It is helpful in overcomeing the shortcomings of traditional methods in time-consuming,laborious,tedious steps features.multiple PCR systems established in this method can be selected according to the type of food matrix.A total of 339 food samples have been detected for 11 genes mentioned.The results showed every gene that the positive ratio of O45 serogroup gene and O145 serogroup gene were 11.5% and 9.1% respectively.Positive ratio of all 3 virulence genes detected in the same sample was 2.4%.Positive ratio of stx2 gene was 20.1%,and was higher than that of stx1 gene,4.4%.Positive ratio of both stx2 and eae detected in the same sample was 15.9%,and was higher than that of both stx2 and eae detected in the same sample,1.5%.Positive ratio of the seven non-O157 EHEC,10.3%,was found higher than that of EHEC O157,0.9%。The results showed that positive ratios of unprocessed raw pork and raw beef is the highest in the investigated sample types,and other processed products,the positive ratios of aquatic products,poultry products,edible vegetables which have been processed are lower.Therefore,positive ratios of EHEC was somewhat related to the type of food matrix and the levels of processing.