Real Time PCR versus Conventional Methods For Detection of Viable But NonCulturable E. Coli O157 Isolated From Food of Animal Origin

Abstract

Escherichia coliO157:H7 consider an important zoonotic food borne pathogen that causebloody diarrhea, hemorrhagic colitis (HC), hemolytic-uremic syndrome (HUS), andthrombotic thrombocytopenic purpura (TTP) in human. E. coli O157:H7 use a viable butnon culturable (VBNC) state as a survival strategy which regarded as a threat to foodsecurity and the health of society. In this dormant stage, pathogens can evadeidentification by standard techniques. So, PCR methods are used for identifying allviable (culturable and non-culturable). This study was performed on 500 samples (300raw meat and 200 raw milk samples) collected from different supermarkets alloverMenoufia, Egypt. Samples were subjected to conventional culture method then positiveculture samples were confirmed by biochemical and serological identification whilenegative-culture sample subjected to conventional PCR and PMA in combination withSYBR green real-time PCR for VBNC E. coli O157:H7 detection. The obtained datashowed that the incidence of E. coli O157:H7 by the conventional method was 3% whilethe incidence of VBNC E. coli O157:H7 by PMA with SYBR green real-time PCR togetherwas 10% for ten tested samples and 0.21% of 475 negative-culture samples. Thisstudy revealed that conventional and immunoassay methods were unable to identifyVBNC case, also conventional PCR is unable to distinguish between dead and livebacterial cells. However, employing a real-time PCR technique alongside PMA has thenecessary efficiency and sensitivity for identifying all viable (culturable and nonculturable) E. coli O157:H7.