Role of capsule and O antigen in the virulence of uropathogenic Escherichia coli.

Sohinee Sarkar,Minh-Duy Phan,Mark A Schembri,Makrina Totsika,Glen C Ulett,Christiane Forestier

doi:10.1371/journal.pone.0094786

Sohinee Sarkar, Minh-Duy Phan + Show 4 more

Open Access

https://doi.org/10.1371/journal.pone.0094786

Copy DOI

Journal: PloS one	Publication Date: Apr 10, 2014
Citations: 183	License type: CC BY 4.0

Affiliation: University of Queensland, Griffith University

Abstract

Urinary tract infection (UTI) is one of the most common bacterial infections in humans, with uropathogenic Escherichia coli (UPEC) the leading causative organism. UPEC has a number of virulence factors that enable it to overcome host defenses within the urinary tract and establish infection. The O antigen and the capsular polysaccharide are two such factors that provide a survival advantage to UPEC. Here we describe the application of the rpsL counter selection system to construct capsule (kpsD) and O antigen (waaL) mutants and complemented derivatives of three reference UPEC strains: CFT073 (O6:K2:H1), RS218 (O18:K1:H7) and 1177 (O1:K1:H7). We observed that while the O1, O6 and O18 antigens were required for survival in human serum, the role of the capsule was less clear and linked to O antigen type. In contrast, both the K1 and K2 capsular antigens provided a survival advantage to UPEC in whole blood. In the mouse urinary tract, mutation of the O6 antigen significantly attenuated CFT073 bladder colonization. Overall, this study contrasts the role of capsule and O antigen in three common UPEC serotypes using defined mutant and complemented strains. The combined mutagenesis-complementation strategy can be applied to study other virulence factors with complex functions both in vitro and in vivo.

Highlights

Urinary tract infections (UTIs) are one of the most common types of bacterial infection in humans
Construction of capsule and O antigen insertion mutants A prerequisite for the rpsL-neo counter-selection system is that the bacterial strain should be streptomycin resistant (StrepR) and that this resistance is the direct effect of a mutation within the rpsL gene
Mutation of the waaL (O antigen) and kpsD genes in the pRedETcontaining strains was performed by insertional inactivation following homologous recombination between the bacterial chromosome and the rpsL-neo cassette flanked by 50 bp homology regions and subsequent growth on kanamycin LB-agar

Summary

Introduction

Urinary tract infections (UTIs) are one of the most common types of bacterial infection in humans. The capsule and the O antigen represent two cell-surface associated polysaccharides that contribute to UPEC virulence [12,13,14]. We employed a recently described counter-selection strategy based on streptomycin-kanamycin resistance (mediated by the rpsL and neo genes, respectively) for the mutation of single genes and the subsequent repair of these mutations to generate complemented strains [36]. This enabled us to generate a defined set of wild-type (WT), mutant and complemented strains that were used to test the contributions of O antigen and capsule to UPEC virulence. Our study is the first to demonstrate the use of the rpsL-neo counter-selection system in UPEC, and shows that the O antigen is the major polysaccharide responsible for UPEC serum resistance and survival in the mouse urinary tract

Results

Discussion

Materials and Methods