Identification and prevalence of in vivo-induced genes in enterohaemorrhagic Escherichia coli

Marion Gardette,Simon Le Hello,Patricia Mariani-Kurkdjian,Laetitia Fabre,François Gravey,Annie Garrivier,Estelle Loukiadis,Grégory Jubelin

doi:10.1080/21505594.2019.1582976

Abstract

ABSTRACTEnterohaemorrhagic Escherichia coli (EHEC) are food-borne pathogens responsible for bloody diarrhoea and renal failure in humans. While Shiga toxin (Stx) is the cardinal virulence factor of EHEC, its production by E. coli is not sufficient to cause disease and many Shiga-toxin producing E. coli (STEC) strains have never been implicated in human infection. So far, the pathophysiology of EHEC infection is not fully understood and more knowledge is needed to characterize the “auxiliary” factors that enable a STEC strain to cause disease in humans. In this study, we applied a recombinase-based in vivo expression technology (RIVET) to the EHEC reference strain EDL933 in order to identify genes specifically induced during the infectious process, using mouse as an infection model. We identified 31 in vivo-induced (ivi) genes having functions related to metabolism, stress adaptive response and bacterial virulence or fitness. Eight of the 31 ivi genes were found to be heterogeneously distributed in EHEC strains circulating in France these last years. In addition, they are more prevalent in strains from the TOP seven priority serotypes and particularly strains carrying significant virulence determinants such as Stx2 and intimin adhesin. This work sheds further light on bacterial determinants over-expressed in vivo during infection that may contribute to the potential of STEC strains to cause disease in humans.

Highlights

Shiga toxin-producing Escherichia coli (STEC) are recognized as an important cause of food-borne disease in humans and cause large outbreaks worldwide
To evaluate the efficiency of the recombinase-based in vivo expression technology (RIVET) method described by Osorio et al [18] in enterohaemorrhagic E. coli (EHEC), we placed the reporter gene tnpR under the control of the promoter region of chuA, for which expression is well known to be induced in iron starvation conditions [28]
This result was expected since the RES1 cassette is excised by TnpR with a lower efficiency in comparison with the original RES cassette [18]. Both strains harbouring the PchuAtnpR135 construct had a low resolution that did not exceed 15%. These results indicate that the RIVET strategy based on RES cassette – tnpR combination can be readily used in EHEC and that the use of different alleles of tnpR and RES cassettes may allow the identification of promoter regions with a broad range of promoter strength

Summary

Introduction

Shiga toxin-producing Escherichia coli (STEC) are recognized as an important cause of food-borne disease in humans and cause large outbreaks worldwide. The two cardinal virulence factors in EHEC are type III secretion system (T3SS) and Shiga toxins (Stx). Tir is crucial since it acts as a receptor for the adhesin intimin (encoded by gene eae), upon its integration into the host cell plasma membrane [2]. Thirty distinct intimin subtypes with variation in their C-terminal part (involved in Tir binding) have been described so far [3] and this represents a standard tool for typing EHEC strains as well as for epidemiological studies. The other cardinal virulence factor in EHEC is Stx, which is produced in the gut lumen and crosses the epithelial barrier to reach the bloodstream and the target organs including brain and kidneys. Stx2producing E. coli strains, especially subtypes Stx2a and Stx2d, are more often associated with the most severe symptoms of the disease than Stx1-producing strains [7,8]

Methods

Results

Conclusion