Importance of Heat-Labile Enterotoxin in Colonization of the Adult Mouse Small Intestine by Human EnterotoxigenicEscherichia coliStrains

Abstract

Enterotoxigenic Escherichia coli (ETEC) infections are a significant cause of diarrheal disease and infant mortality in developing countries. Studies of ETEC pathogenesis relevant to vaccine development have been greatly hampered by the lack of a suitable small-animal model of infection with human ETEC strains. Here, we demonstrate that adult immunocompetent outbred mice can be effectively colonized with the prototypical human ETEC H10407 strain (colonization factor antigen I; heat-labile and heat-stable enterotoxin positive) and that production of heat-labile holotoxin provides a significant advantage in colonization of the small intestine in this model.Enterotoxigenic Escherichia coli (ETEC) infections are a significant cause of diarrheal disease worldwide. Infections caused by this heterogeneous group of pathogens remain major causes of diarrheal morbidity and infant mortality in developing countries (11,33,61), are perennially associated with disease in travelers (4, 7, 43) and in soldiers deployed to developing countries (8, 32), and have recently been associated with large outbreaks in developed countries, including the United States (1, 13).In the presently accepted paradigm for ETEC pathogenesis, fimbrial (or fibrillar) colonization factors (CFs) (10,22,27) mediate colonization of the small intestine, where organisms elaborate heat-labile enterotoxin (LT) and/or heat-stable enterotoxin (ST) (21, 68). Vaccine development has largely focused on the CFs; however, development of a broadly protective ETEC vaccine has been hampered due to the considerable heterogeneity of the known CFs (6,52,58,62).While a number of other surface antigens of ETEC have been described (20,25,51), their utility as potential vaccine candidates has not been effectively explored, largely due to the lack of a suitable animal model for testing. Animal models that have been used in previous studies of ETEC include infant (3, 16-18, 28, 47) and adult (9) mice, rats (34, 35), and rabbits (19,46,51). All of these models have inherent difficulties in utilization or require anesthesia and/or significant surgical manipulation. Some of the models have not been thoroughly evaluated.We sought to develop a murine model of small-intestinal colonization with human ETEC isolates using adult immunocompetent mice. Here, we report our initial use of this model using the prototypical human ETEC H10407 strain. Furthermore, similar to recent studies of porcine ETEC infection of gnotobiotic piglets (2), we demonstrate that elaboration of the heat-labile toxin provides a distinct advantage to the organism in establishing early colonization of the small-intestinal mucosa.

Heat-Labile Enterotoxin PromotesEscherichia coliAdherence to Intestinal Epithelial Cells

“…1A and 2A). These data parallel prior observations that the eltA mutant was markedly defective (ϳ10-fold) in intestinal colonization relative to wt ETEC (1). Coadministration of LT with ETEC inocula was sufficient to increase adherence of ETEC lacking eltAB to near-wt levels ( Fig.…”

“…Recent in vivo observations suggesting a potential role for bacterial enterotoxins in initial stages of host colonization (1,5) prompted us to quantify, in vitro, potential contributions of LT in modulating early stages of pathogen-host interactions. We utilized an IPEC-J2 cell culture system to quantify the bacterial adherence levels of porcine ETEC isolates differing in their expression of enterotoxins.…”

“…Recent in vivo experiments with both human and porcine ETEC isolates are consistent with this hypothesis. Preliminary characterization of a mouse model of ETEC infection indicated that production of LT provides an advantage in colonization of the murine small intestine (1). A porcine ETEC strain deficient in LT expression colonized piglets less efficiently than either the parent strain or a strain in which the mutation was complemented (5).…”

Concomitant EnterotoxigenicEscherichia coliInfection Induces Increased Immune Responses toVibrio choleraeO1 Antigens in Patients with Cholera in Bangladesh

“…In addition, LT was shown to increase colonization by ETEC in both mouse and pig experimental models (1,2). Adherence may be enhanced by the ADP-ribosylating activity of LT and by the levels of cyclic AMP (cAMP) that can affect host cell physiology (4), leading to an enhanced adherence of V. cholerae to the gut (20,29).…”

Decreased Expression of Colonic Slc26a3 and Carbonic Anhydrase IV as a Cause of Fatal Infectious Diarrhea in Mice

“…Mice are generally considered poor models for studying watery diarrhea caused by pathogenic agents that infect people. Thus, diarrheagenic Escherichia coli, Vibrio cholerae, Campylobacter jejuni, Shigella spp., and Salmonella spp., as well as rotavirus, either fail to efficiently infect mice or do not cause appreciable diarrhea except in infant mice or older mice that have been manipulated (germfree, antibiotic pretreated, iron loaded, or given gastric acid-reducing agents) to increase susceptibility (1,5,16,26,27,36,39,43,45,47,56). Thus, elucidating the pathogenesis of acute diarrheal illness is hampered by the lack of a suitable small-animal model.…”