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Infection and Immunity, September 2004, p. 5402-5411, Vol. 72, No. 9
0019-9567/04/$08.00+0     DOI: 10.1128/IAI.72.9.5402-5411.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Mutation of toxB and a Truncated Version of the efa-1 Gene in Escherichia coli O157:H7 Influences the Expression and Secretion of Locus of Enterocyte Effacement-Encoded Proteins but not Intestinal Colonization in Calves or Sheep

Mark P. Stevens,1* Andrew J. Roe,2 Isabella Vlisidou,1 Pauline M. van Diemen,1 Roberto M. La Ragione,3 Angus Best,3 Martin J. Woodward,3 David L. Gally,2 and Timothy S. Wallis1

Division of Microbiology, Institute for Animal Health, Compton Laboratory, Berkshire,1 Zoonotic and Animal Pathogens Laboratory, Department of Medical Microbiology, University of Edinburgh, Edinburgh,2 Department of Food and Environmental Safety, Veterinary Laboratories Agency, New Haw, Addlestone, Surrey, United Kingdom3

Received 13 February 2004/ Returned for modification 13 May 2004/ Accepted 17 May 2004

Enterohemorrhagic Escherichia coli (EHEC) strains comprise a broad group of bacteria, some of which cause attaching and effacing (AE) lesions and enteritis in humans and animals. Non-O157:H7 EHEC strains contain the gene efa-1 (referred to in previous publications as efa1), which influences adherence to cultured epithelial cells. An almost identical gene in enteropathogenic E. coli (lifA) mediates the inhibition of lymphocyte proliferation and proinflammatory cytokine synthesis. We have shown previously that significantly lower numbers of EHEC O5 and O111 efa-1 mutants are shed in feces following experimental infection in calves and that these mutants exhibit reduced adherence to intestinal epithelia compared with isogenic wild-type strains. E. coli O157:H7 strains lack efa-1 but encode a homolog on the pO157 plasmid (toxB/l7095) and contain a truncated version of the efa-1 gene (efa-1'/z4332 in O island 122 of the EDL933 chromosome). Here we report that E. coli O157:H7 toxB and efa-1' single and double mutants exhibit reduced adherence to cultured epithelial cells and show reduced expression and secretion of proteins encoded by the locus of enterocyte effacement (LEE), which plays a key role in the host-cell interactions of EHEC. The activity of LEE1, LEE4, and LEE5 promoters was not significantly altered in E. coli O157:H7 strains harboring toxB or efa-1' mutations, indicating that the effect on the expression of LEE-encoded secreted proteins occurs at a posttranscriptional level. Despite affecting type III secretion, mutation of toxB and efa-1' did not significantly affect the course of fecal shedding of E. coli O157:H7 following experimental inoculation of 10- to 14-day-old calves or 6-week-old sheep. Mutation of tir caused a significant reduction in fecal shedding of E. coli O157:H7 in calves, indicating that the formation of AE lesions is important for colonization of the bovine intestine.

* Corresponding author. Mailing address: Division of Microbiology, Institute for Animal Health, Compton Laboratory, Berkshire RG20 7NN, United Kingdom. Phone: 1635 578411. Fax: 1635 577243. E-mail: mark-p.stevens{at}bbsrc.ac.uk.

Editor: A. D. O'Brien

Infection and Immunity, September 2004, p. 5402-5411, Vol. 72, No. 9
0019-9567/04/$08.00+0     DOI: 10.1128/IAI.72.9.5402-5411.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



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