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Infection and Immunity, November 2000, p. 6472-6477, Vol. 68, No. 11
0019-9567/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Contribution of Plasmid-Encoded Fimbriae and Intimin to Capacity of Rabbit-Specific Enteropathogenic Escherichia coli To Attach to and Colonize Rabbit Intestine

Efrosinia O. Krejany, Travis H. Grant, Vicki Bennett-Wood, Louise M. Adams, and Roy M. Robins-Browne*

Department of Microbiology and Immunology, University of Melbourne, and Microbiological Research Unit, Murdoch Children's Research Institute, Parkville, Victoria 3052, Australia

Received 27 March 2000/Returned for modification 15 June 2000/Accepted 7 August 2000

Attachment to the intestinal mucosa is an essential step in the pathogenesis of diarrhea caused by enteropathogenic Escherichia coli (EPEC). Fimbriae and intimin, the outer membrane protein product of the chromosomal eae gene, contribute to this process, but their relative roles and the nature of their interaction are not known. The aim of this study was to determine the relative contribution of plasmid-encoded fimbriae, termed Ral, and intimin to the capacity of rabbit-specific EPEC (REPEC) to attach to the intestinal mucosa of rabbits. To achieve this, we constructed a series of mutants in REPEC strain 83/39 (O15:H-), in which the ralE and eae genes were insertionally inactivated. These strains were then inoculated into ligated loops of rabbit ileum, which were resected 18 h later and examined by light and electron microscopy. The results showed that intimin, but not Ral, is essential for the elicitation of attaching-effacing lesions by REPEC. Nevertheless, a Delta eae Ral-bearing mutant adhered to the intestinal epithelium to the same extent as its eae-positive parent and far more extensively than an eae+ Delta ral strain. To examine the contribution of Ral and intimin to colonization of rabbit intestine, we fed these strains to weanling rabbits, which were killed 4 days later, so that the number of bacteria in various regions of the intestine could be determined. The results indicated that strain 83/39 requires both Ral and intimin to colonize the intestine successfully and that a Delta eae Delta ralE double mutant was incapable of colonizing the intestine. Taken together, these findings indicate that Ral and intimin act independently as adhesion factors of REPEC strain 83/39 and that this strain carries no other significant colonization factor. When both Ral and intimin are present, they appear to act cooperatively, with Ral-mediated adhesion preceding that mediated by intimin.

* Corresponding author. Mailing address: Microbiological Research Unit, Royal Children's Hospital, Parkville, Victoria 3052, Australia. Phone: (61-3) 9345-5741. Fax: (61-3) 9345-5764. E-mail: rbrowne{at}cryptic.rch.unimelb.edu.au.

Infection and Immunity, November 2000, p. 6472-6477, Vol. 68, No. 11
0019-9567/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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