Multiplex polymerase chain reaction typing scheme based on Escherichia coli O157:H7 Sakai prophage (Sp)-associated genes

Abstract

Objectives: Escherichia coli strains of the O157 serogroup include significant foodborne pathogens: enterohemorrhagic E. coli (EHEC) and enteropathogenic E. coli, which are responsible for a considerable number of hospitalizations and deaths worldwide each year. There is a constant need for rapid, reliable, and easy-to-use methods for their identification, typing, and phylogenetic classification. In this study, we proposed a new multiplex polymerase chain reaction (PCR)-based typing system for pathogenic E. coli, focusing on the O157 serogroup.

Methods: We designed primers targeting 12 lambdoid prophage regions carried by the prototypic polylysogenic strain of EHEC, the O157:H7 Sakai strain. The reactions were tested in vitro as well as in silico with the PubMLST database.

Results: The PCR assays can be grouped into four multiplex reactions, and their results can be given as a four-digit code. In vitro and in silico testing showed that these Sakai prophage regions are prevalent not only in E. coli O157 strains but also in Shiga toxigenic E. coli non-O157 strains and the method provides appropriate resolution.

Conclusions: The proposed method could be a valuable tool in epidemiologic tracing and preliminary phylogenetic grouping of this diverse group of pathogens.

Keywords: EHEC; O157; multiplex PCR; pathogenic Escherichia coli; prophage; typing.