Diarrhea due to Escherichia coli in the rabbit: a novel mechanism.

A strain of Escherichia coli O15 (RDEC-1) isolated from several rabbits with diarrhea was examined to determine (1) whether the strain could produce diarrhea when administered by the orogastric route to other rabbits and (2) whether this strain was invasive or enterotoxigenic. Strain RDEC-1 produced diarrhea in 48 of 62 rabbits when given by the orogastric route in doses that ranged from 1.5 X 10(2) to 4 X 10(10) bacteria. The organism did not give a positive result in the 18-hr ileal loop or Serény tests and did not invade HeLa cells. Culture supernatants of strain RDEC-1 did not give positive results in the 6- or 18-hr rabbit ileal loop, suckling mouse, Y-1 adrenal cell, or Chinese hamster ovary cell assays. Fluorescent antibody staining of sections of intestine prepared in a cryostat revealed great numbers of E. coli strain RDEC-1 that adhered to the epithelial surface. It is evident that E. coli can produce diarrhea without being able to invade the mucosa or synthesize enterotoxin. Strains of E. coli similar to RDEC-1 may account for some of the E. coli-associated diarrhea that occurs in humans.

Production of diarrhea in the rabbit by a mutant of Escherichia coli (RDEC-1) that does not express adherence (AF/R1) pili.

Escherichia coli (RDEC-1) adheres to Peyer's patch and absorptive epithelium in the rabbit in the closely adhering manner characteristic of enteropathogenic and enterohemorrhagic E. coli. An adherence pilus (AF/R1) is important for adherence to Peyer's patch M cells in vivo and ileal brush borders in vitro. A nonpiliated mutant (42-2-37-8) of the RDEC-1 strain colonized the gut lumen less readily than the parent strain. The mutant adhered infrequently to Peyer's patch lymphoid follicle epithelium (22% vs. 84%, P less than .0001). Ileal close adherence was less frequent at 3 d, but by 9 d after inoculation had increased, approaching that of the parent RDEC-1 strain. However, adherence was focal, and fewer bacteria were present at each adherence site compared with the parent RDEC-1 strain. The result was a lower frequency of diarrhea (34% vs. 65%, P = .003) and mortality (9.4% vs. 27%, P = .035) with the 42-2-37-8 strain. Loss of AF/R1 pili compromised the ability of the RDEC-1 strain to adhere to Peyer's patch and absorptive epithelium and to produce diarrhea.

A rabbit model of diarrhea due to invasive Escherichia coli.

A nonpiliated strain of invasive Escherichia coli of human origin (HInvEC) was given to rabbits (weight, 0.7-1.1 kg) in doses ranging from 1.5 X 10(8) to 2.5 X 10(10) bacteria. E. coli strain HInvEC colonized the ileum, cecum, and colon in large numbers for one to three days and produced diarrhea in 91 ((58%) of the 156 rabbits. A dose of 2.5 X 10(10) bacteria reliably and repeatedly elicited diarrheal disease in 80% of the animals. The acute pathohistology as determined by light microscopy, transmission electron microscopy, and immune fluorescent microscopy was manifest in the distal ileum, cecum, and colon. Prominent findings included mucosal ulcers, bacterial invasion of the lamina propria, and a polymorphonuclear infiltrate in the lamina propria. Diarrhea due to strain HInvEC was followed by a rise in the titer of serum antibody to O antigen of E. coli.

Characterization of the Escherichia coli AF/R1 pilus operon: novel genes necessary for transcriptional regulation and for pilus-mediated adherence.

We isolated the genetic determinant of AF/R1 pilus production in attaching/effacing Escherichia coli RDEC-1 and identified seven genes required for pilus expression and function. DNA sequence analysis of the structural subunit gene afrA corrected an error in the published sequence and extended homology with the F18 pilus subunit of pig edema E. coli strains. AfrB and AfrC, encoded downstream from AfrA, were required for pilus expression. AfrB was related to the usher protein PefC of Salmonella typhimurium plasmid-encoded fimbriae, and AfrC was related to PefD, a chaperone protein. AfrD and AfrE, encoded downstream from AfrC, were not necessary for the expression of AF/R1 pili but were required for ileal adherence as assayed by ileal brush border aggregation. Thus, the adhesive subunit of the AF/R1 pilus is distinct from the structural subunit, as is the case for Pap pili and type 1 pili. AfrD was related to FedE of the F18 fimbrial operon of the E. coli strain that causes edema disease in pigs. AfrE was a novel protein. AfrR and AfrS are encoded upstream from AfrA, in the opposite orientation. AfrR is related to the AraC family of transcriptional regulators, and AfrR and AfrS interact to function in a novel mode of transcriptional activation of afrA. AF/R1 pili mediate the adherence to Peyer's patch M cells, ileal mucosa, and colonic mucosa in a rabbit model of diarrhea caused by enteropathogenic E. coli. Our observations will facilitate the further study of the phenomena of M-cell adherence.