Simultaneous detection of six major virulent genes of Escherichia coli O157:H7 from seafood by multiplex PCR and its validation through spiked study

Multiplex PCR for E. coli O157:H7

Abstract

Enterohaemorrhagic Escherichia coli (EHEC), mainly the serotype O157:H7 is a global concern in foodborne outbreaks,. The incidence of E. coli O157:H7 in seafood sold in India has been reported as 8.4%. The conventional method for the detection of E. coli O157:H7 from seafood is cumbersome and time consuming making this strain particularly challenging to detect. In this study a multiplex PCR was developed targeting six major virulence factors of the pathogen viz. shigatoxin 1 and 2, attachment and effacement factor, flagellar antigen, hemolysin gene and O157 antigen. The sensitivity of the multiplex PCR developed in the study was found to be 120 pg μl-1 of DNA and that of E. coli 0157:H7 spiked samples was 2 cfu ml-1. The multiplex PCR developed in this study showed 100% specificity in identifying E. coli O157:H7, effectively distinguishing it from non-target bacteria. This highly sensitive and specific multiplex PCR is the first of its kind to detect E. coli O157:H7 in seafood by targeting six major virulence genes.

Keywords: E. coli O157:H7, EHEC, Foodborne pathogen, Seafood, Sensitivity, Shigatoxin, Specificity