Identification and Characterization of Novel Compounds Blocking Shiga Toxin Expression in Escherichia coli O157:H7.

Alejandro Huerta-Uribe,Rudi Marquez,Stephen Fitzgerald,James P R Connolly,Zoe R Marjenberg,Gillian Douce,Michael Elofsson,David L Gally,Hanna Uvell,Nao Yamaguchi,Olwyn Byron,Andrew J Roe,Nuria Carpena

doi:10.3389/fmicb.2016.01930

Abstract

Infections caused by Shiga toxin (Stx)-producing E. coli strains constitute a health problem, as they are problematic to treat. Stx production is a key virulence factor associated with the pathogenicity of enterohaemorrhagic E. coli (EHEC) and can result in the development of haemolytic uremic syndrome in infected patients. The genes encoding Stx are located on temperate lysogenic phages integrated into the bacterial chromosome and expression of the toxin is generally coupled to phage induction through the SOS response. We aimed to find new compounds capable of blocking expression of Stx type 2 (Stx2) as this subtype of Stx is more strongly associated with human disease. High-throughput screening of a small-molecule library identified a lead compound that reduced Stx2 expression in a dose-dependent manner. We show that the optimized compound interferes with the SOS response by directly affecting the activity and oligomerization of RecA, thus limiting phage activation and Stx2 expression. Our work suggests that RecA is highly susceptible to inhibition and that targeting this protein is a viable approach to limiting production of Stx2 by EHEC. This type of approach has the potential to limit production and transfer of other phage induced and transduced determinants.

Highlights

Enterohemorrhagic E. coli (EHEC) are a group of Shiga toxin (Stx) producing pathogenic E. coli strains, which are associated with a broad spectrum of disease ranging from mild diarrhea to severe haemorrhagic colitis and haemolytic uremic syndrome (HUS) (Paton and Paton, 1998)
The development of haemorrhagic colitis and HUS is dependent on the production of Stx, a family of related toxins that are essential for disease
Stx produced by enterohaemorrhagic E. coli (EHEC) strains are largely differentiated into two types that share 55% sequence homology (Fraser et al, 2004): Stx1, which differs from Shigella dysenteriae Stx by a single amino acid, and Stx type 2 (Stx2), which is structurally similar to Stx1 but antigenically distinct

Summary

Introduction

Enterohemorrhagic E. coli (EHEC) are a group of Shiga toxin (Stx) producing pathogenic E. coli strains, which are associated with a broad spectrum of disease ranging from mild diarrhea to severe haemorrhagic colitis and haemolytic uremic syndrome (HUS) (Paton and Paton, 1998). The first recognition of an EHEC strain as a foodborne pathogen occurred in the US in 1982 during an investigation of customers from a fast-food restaurant chain who had bloody diarrhea and severe abdominal cramping with no fever (Riley et al, 1983). Stx and Stx can be further classified into several subtypes based on the sequence-based relatedness of the proteins. These comprise three Stx subtypes (1a, 1c, 1d) and seven Stx subtypes (2a, 2b, 2c, 2d, 2e, 2f, and 2g)

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