Chromosomal instability in enterohaemorrhagic Escherichia coli O157:H7: impact on adherence, tellurite resistance and colony phenotype

Martina Bielaszewska,Rita Prager,Thomas Aldick,Wenlan Zhang,Phillip I Tarr,Helge Karch,Alexander Mellmann,Ulrich Dobrindt,Barbara Middendorf

doi:10.1111/j.1365-2958.2010.07499.x

Abstract

Tellurite (Tel) resistant enterohaemorrhagic Escherichia coli (EHEC) O157:H7 is a global pathogen. In strain EDL933 Tel resistance (TelR) is encoded by duplicate ter cluster in O islands (OI) 43 and 48, which also harbour iha, encoding the adhesin and siderophore receptor Iha. We identified five EHEC O157:H7 strains that differentiate into large (L) colonies and small (S) colonies with high and low Tel minimal inhibitory concentrations (MICs) respectively. S colonies (Tel-MICs ≤ 4 µg ml−1) sustained large internal deletions within the TelR OIs via homologous recombination between IS elements and lost ter and iha. Moreover, complete excision of the islands occurred by site-specific recombination between flanking direct repeats. Complete excision of OI 43 and OI 48 occurred in 1.81 × 10−3 and 1.97 × 10−4 cells in culture, respectively; internal deletion of OI 48 was more frequent (9.7 × 10−1 cells). Under iron limitation that promotes iha transcription, iha-negative derivatives adhered less well to human intestinal epithelial cells and grew slower than did their iha-positive counterparts. Experiments utilizing iha deletion and complementation mutants identified Iha as the major factor responsible for these phenotypic differences. Spontaneous deletions affecting TelR OIs contribute to EHEC O157 genome plasticity and might impair virulence and/or fitness.

Highlights

Enterohaemorrhagic Escherichia coli (EHEC) O157:H7 is the predominant Shiga toxin (Stx)-producing pathogen of humans (Karch et al, 2005; Tarr et al, 2005)
We identified five EHEC O157:H7 strains that differentiate into large (L) colonies and small (S) colonies with high and low Tel minimal inhibitory concentrations (MICs) respectively
S colonies (Tel-MICs Յ 4 mg ml-1) sustained large internal deletions within the Tel resistance (TelR) O islands (OI) via homologous recombination between IS elements and lost ter and iha

Summary

Introduction

Enterohaemorrhagic Escherichia coli (EHEC) O157:H7 is the predominant Shiga toxin (Stx)-producing pathogen of humans (Karch et al, 2005; Tarr et al, 2005). EHEC O157:H7 resists the highly toxic tellurium oxyanion, tellurite (TeO32-; Tel), and grows in concentrations of Tel that inhibit most other E. coli (Zadik et al, 1993; Taylor et al, 2002; Bielaszewska et al, 2005; Orth et al, 2007) This characteristic, together with its inability to ferment sorbitol, has been exploited in selective strategies to isolate EHEC O157:H7 from faeces, food and the environment using Tel-containing media, such as cefixime-Tel sorbitol MacConkey agar (CT-SMAC) (Onoue et al, 1999; Van Duynhoven et al, 2002). O157 Sakai outbreak strain RIMD 0509952 harbours only a single ter cluster-containing island (SpLE1) integrated in serX (Hayashi et al, 2001). SF, sorbitol fermentation; GUD, production of b-D-glucuronidase; EHEC-Hly, EHEC haemolysin production; CT-SMAC, cefixime-tellurite sorbitol MacConkey agar

Results

Discussion

Conclusion