Adhesion of Escherichia coli under flow conditions reveals potential novel effects of FimH mutations

T Feenstra,R Brandes,T Stockner,A Peschel,S C Satchell,R Kain,A J Rees,M S Thøgersen,E Wieser,F M Aarestrup,M J Ball

doi:10.1007/s10096-016-2820-8

Abstract

FimH-mediated adhesion of Escherichia coli to bladder epithelium is a prerequisite for urinary tract infections. FimH is also essential for blood-borne bacterial dissemination, but the mechanisms are poorly understood. The purpose of this study was to assess the influence of different FimH mutations on bacterial adhesion using a novel adhesion assay, which models the physiological flow conditions bacteria are exposed to. We introduced 12 different point mutations in the mannose binding pocket of FimH in an E. coli strain expressing type 1 fimbriae only (MSC95-FimH). We compared the bacterial adhesion of each mutant across several commonly used adhesion assays, including agglutination of yeast, adhesion to mono- and tri-mannosylated substrates, and static adhesion to bladder epithelial and endothelial cells. We performed a comparison of these assays to a novel method that we developed to study bacterial adhesion to mammalian cells under flow conditions. We showed that E. coli MSC95-FimH adheres more efficiently to microvascular endothelium than to bladder epithelium, and that only endothelium supports adhesion at physiological shear stress. The results confirmed that mannose binding pocket mutations abrogated adhesion. We demonstrated that FimH residues E50 and T53 are crucial for adhesion under flow conditions. The coating of endothelial cells on biochips and modelling of physiological flow conditions enabled us to identify FimH residues crucial for adhesion. These results provide novel insights into screening methods to determine the effect of FimH mutants and potentially FimH antagonists.

Highlights

Infection with Escherichia coli is the most frequent cause of septicaemia in humans and commonly originates from the urinary tract [1]
We showed that E. coli MSC95-FimH adheres more efficiently to microvascular endothelium than to bladder epithelium, and that only endothelium supports adhesion at physiological shear stress
There is a lack of data on FimH-dependent bacterial adhesion to microvascular endothelium that is thought to underlie blood-borne dissemination of E. coli [4]. We addressed this issue by generating and validating a panel of multiply disabled E. coli strains that uniquely express type 1 fimbriae and normal or mutated FimH [14], and systematically analysing the ability of the mutant strains to adhere to microvascular endothelium and bladder epithelium, under both static conditions and physiological shear stress

Summary

Introduction

Infection with Escherichia coli is the most frequent cause of septicaemia in humans and commonly originates from the urinary tract [1]. Uropathogenic E. coli (UPEC) adhere to bladder epithelium in a process mediated by type 1 fimbriae via FimH engaging uroplakin 1a on urothelium, leading to urinary tract infection [2, 3]. FimH promotes invasion and is critical for blood-borne dissemination to other tissues [4]. In neonatal meningitis, FimH is essential for the localisation of UPEC to brain microvascular endothelium and invasion of the meninges [5, 6]. This establishes the pathogenic importance of FimH-mediated adhesion beyond the urinary tract. FimH is located at the tip of type 1 fimbriae expressed by Gram-negative pathogens, including E. coli, Salmonella

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