Direct detection of Escherichia coli O-157 and its major virulence factor genes in animal faeces at slaughter using multiplex polymerase chain reaction (PCR)

Abstract

Although more than 100 serotypes of shigatoxigenic Escherichia coli (STEC) have been implicated in cases of human diseases, E. coli O-157 is the most common serogroup connected with sporadic cases and large outbreaks of diseases in many countries. Rapid and sensitive identification of this dangerous pathogen is important for patient management and for prompt epidemiological investigations. PCR has become a very rapid and reliable tool for the molecular diagnosis of E. coli O-157. PCR assays are usually aimed at detecting the shiga toxins, the intimin protein and enterohaemolysin. In the present study, a mPCR-based protocol is described as that which uses one primer set to detect the gene responsible for the production of the O-antigen synthesis (rfb O-157) and four primer set to detect the major virulence factor genes including the Shigatoxin type 1 and 2 (stx(1) and stx(2)), intimin (eaeA) and enterohemolysin (EHEC hlyA) directly from 190 samples of animal faeces at the time of slaughter after overnight incubation of stool specimens in BPW. In this research, we use one primer set for detection of the gene responsible for the production of the O-antigen synthesis (rfb O-157) and four primer set for detection of the Shigatoxin type 1 and 2 (stx(1) and stx(2)), intimin (eaeA) and enterohemolysin (EHEC hlyA) producing genes directly from 190 samples of animal faeces at the time of slaughter after overnight incubation of stool specimens in BPW. This study has established the presence of rather high prevalence of E. coli O-157-positive animals at abattoirs (These consisted of 4.2% of cattle and 2.1% of sheep), providing an increased risk of transmission of E. coli O-157 to the food chain and contamination of human.