Dynamic Interactions of a Conserved Enterotoxigenic Escherichia coli Adhesin with Intestinal Mucins Govern Epithelium Engagement and Toxin Delivery

Abstract

e At present, there is no vaccine for enterotoxigenic Escherichia coli (ETEC), an important cause of diarrheal illness. Nevertheless, recent microbial pathogenesis studies have identified a number of molecules produced by ETEC that contribute to its virulence and are novel antigenic targets to complement canonical vaccine approaches. EtpA is a secreted two-partner adhesin that is conserved within the ETEC pathovar. EtpA interacts with the tips of ETEC flagella to promote bacterial adhesion, toxin delivery, and intestinal colonization by forming molecular bridges between the bacteria and the epithelial surface. However, the nature of EtpA interactions with the intestinal epithelium remains poorly defined. Here, we demonstrate that EtpA interacts with glycans presented by transmembrane and secreted intestinal mucins at epithelial surfaces to facilitate pathogen-host interactions that culminate in toxin delivery. Moreover, we found that a major effector molecule of ETEC, the heat-labile enterotoxin (LT), may enhance these interactions by stimulating the production of the gel-forming mucin MUC2. Our studies suggest, however, that EtpA participates in complex and dynamic interactions between ETEC and the gastrointestinal mucosae in which host glycoproteins promote bacterial attachment while simultaneously limiting the epithelial engagement required for effective toxin delivery. Collectively, these data provide additional insight into the intricate nature of ETEC interactions with the intestinal epithelium that have potential implications for rational approaches to vaccine design. Enterotoxigenic Escherichia coli (ETEC) is a major cause of infectious diarrhea in the developing world (1), where this organism causes millions of infections and hundreds of thousands of deaths, particularly in young children (2). This organism contributes substantially to the large global burden of diarrheal illness that has been associated with a number of important but poorly understood sequelae, including "environmental enteropathy" and compromised growth and cognitive development (3).ETEC was discovered more than 40 years ago in cases of diarrheal illness clinically indistinguishable from cholera (4) and is defined by the production of plasmid-encoded heat-labile enterotoxin (LT) and/or heat-stable enterotoxin (ST). LT and ST must successfully engage cognate receptors (GM-1 ganglioside and guanylyl cyclase C, respectively) on the surface of epithelial cells to activate the production of cyclic nucleotides. The respective increases in intracellular cyclic AMP (cAMP) and cGMP trigger signaling events culminating in altered salt and water absorption in the small intestine accompanied by voluminous watery diarrhea (5).Because of their substantial impact on global health, these pathogens are a principal target for vaccine development. Unfortunately, there is currently no vaccine that affords significant broad-based protection against ETEC (6). Despite decades of research, the current understanding of the pathogenesis of this complex and hig...

Interplay of a secreted protein with type IVb pilus for efficient enterotoxigenic Escherichia coli colonization

“…Notably, the wild-type CFA/III strain is nonflagellated and nonmotile, and some ETEC strains that possess Longus, one of the most prevalent CFs among ETEC clinical isolates that has substantial structural homology with CFA/III, are also nonflagellated ( 49 , 50 ). Our functional model of CofJ indicated it acts as a molecular bridge, in striking resemblance with EtpA; therefore, it is tempting to speculate that the interplay between the secreted protein and relatively long (5–15 μm or more) proteinaceous fibers, such as flagellum and T4bP ( 48 ), is a common strategy for initial attachment of ETEC, avoiding the mucosal barrier. Recent reports demonstrate that EtpA-targeted vaccination is effective ( 51 ), and our results showed that Fab fragments of an antibody against CofJ significantly inhibited adhesion of a CFA/III-expressing E. coli strain ( Figs.…”

“…Before intimate association with the target-cell surface, possibly via CFs of relatively short (1–5 μm) CU pili ( 44 , 45 ), another specific mechanism is apparently required for initial attachment while avoiding the mucosal barrier consisting of an inner layer of ∼15–30 μm ( 46 ). One such mechanism was recently proposed for the non-CF ETEC virulence factor EtpA, which is secreted and interacts as a molecular bridge with target cells and the tip of flagellum, a common bacterial appendage with typical length of 10–15 μm ( 47 , 48 ). Notably, the wild-type CFA/III strain is nonflagellated and nonmotile, and some ETEC strains that possess Longus, one of the most prevalent CFs among ETEC clinical isolates that has substantial structural homology with CFA/III, are also nonflagellated ( 49 , 50 ).…”

Virulence and transcriptome profile of multidrug-resistant Escherichia coli from chicken

“…The transcriptional analysis of the virulence genes in E. coli were evaluated. Adhesin and fimH are fimbrial tip adhesion molecules in E. coli which facilitate adhesion, internalization, and biofilm formation 62 , 63 . In the present study, many fimbrial genes that encode the type-1 fimbrial proteins fimA, fimC , fimH , and fimI were involved in infection 11 , 62 .…”

Interrogation of a live-attenuated enterotoxigenic Escherichia coli vaccine highlights features unique to wild-type infection

“…The genomes of the three vaccine strains and parents also revealed that the etpBAC locus, which encodes the two-partner secretion system responsible for production and export of the EtpA adhesin, 18 was present in two of the three parental strains but missing from the vaccine altogether, likely the result of engineering the vaccine strains to remove the plasmid-encoded toxins. EtpA is a high molecular weight glycoprotein secreted by ETEC that appears to facilitate bacterial adhesion by serving as a molecular bridge between flagella 37 and the enterocyte surface where it binds to N-acetylgalactosamine (GalNAc) residues 38 particularly when they are presented as the terminal glycan on human A blood group antigen. Interestingly, human challenge studies with H10407 demonstrate that individuals with A blood group are significantly more likely to experience severe diarrhea when challenged with this EtpA-producing strain.…”