Critical role of T cell-dependent serum antibody, but not the gut-associated lymphoid tissue, for surviving acute mucosal infection with Citrobacter rodentium, an attaching and effacing pathogen. (1/177)

Citrobacter rodentium uses virulence factors similar to the enteropathogenic Escherichia coli to produce attaching and effacing lesions in the distal colon of mice. We used this infection model to determine components of adaptive immunity needed to survive infection. During acute infection, wild-type mice develop breaks across infected epithelial surfaces but resolve infection. Surprisingly, mice markedly deficient in mucosal lymphocyte populations from beta(7) integrin deficiency resolve infection, as do CD8alpha-/- or TCR-delta-/- mice. In contrast, CD4-/- or TCR-beta-/- mice develop polymicrobial sepsis and end-organ damage, and succumb during acute infection, despite epithelial damage similar to wild-type mice. B cell-deficient (MuMT-/-) or B and T cell-deficient (recombinase-activating gene 2-/-) mice develop severe pathology in colon and internal organs, and deteriorate rapidly during acute infection. Surviving mice develop robust Citrobacter-specific serum IgM responses during acute infection, whereas mice that succumb do not. However, CD4-/- mice receiving serum Igs from infected wild-type mice survive and clear the infection. Our data show that survival of apparently self-limited and luminal mucosal infections requires a systemic T cell-dependent Ab response against bacteria that enter through damaged mucosa. These findings have implications for understanding host defense against mucosal infections, including the pathogenesis of these diseases in immunocompromised populations.  (+info)

Dissecting virulence: systematic and functional analyses of a pathogenicity island. (2/177)

Bacterial pathogenicity islands (PAI) often encode both effector molecules responsible for disease and secretion systems that deliver these effectors to host cells. Human enterohemorrhagic Escherichia coli (EHEC), enteropathogenic E. coli, and the mouse pathogen Citrobacter rodentium (CR) possess the locus of enterocyte effacement (LEE) PAI. We systematically mutagenized all 41 CR LEE genes and functionally characterized these mutants in vitro and in a murine infection model. We identified 33 virulence factors, including two virulence regulators and a hierarchical switch for type III secretion. In addition, 7 potential type III effectors encoded outside the LEE were identified by using a proteomics approach. These non-LEE effectors are encoded by three uncharacterized PAIs in EHEC O157, suggesting that these PAIs act cooperatively with the LEE in pathogenesis. Our findings provide significant insights into bacterial virulence mechanisms and disease.  (+info)

Identification of a novel Citrobacter rodentium type III secreted protein, EspI, and roles of this and other secreted proteins in infection. (3/177)

Citrobacter rodentium is a member of a group of pathogens that colonize the lumen of the host gastrointestinal tract via attaching and effacing (A/E) lesion formation. C. rodentium, which causes transmissible colonic hyperplasia in mice, is used as an in vivo model system for the clinically significant A/E pathogens enterohemorrhagic and enteropathogenic Escherichia coli. These bacteria all contain a pathogenicity island called the locus of enterocyte effacement (LEE), which encodes a type III secretion system that is designed to deliver effector proteins into eukaryotic host cells. These effectors are involved in the subversion of host eukaryotic cell functions to the benefit of the bacterium. In this study we used mutant strains to determine the effects of the C. rodentium LEE-encoded effectors EspF, EspG, EspH, and Map on virulence in the mouse model. In addition, we identified a novel secreted protein, EspI encoded outside the LEE, whose secretion is also dependent on a functional type III secretion system. Mutant strains with each of the effectors investigated were found to be outcompeted by wild-type bacteria in mixed-infection experiments in vivo, although the effects of EspF and EspH were only subtle. In single-infection experiments, we found that EspF, EspG, and EspH are not required for efficient colonization of the mouse colon or for the production of hyperplasia. In contrast, strains producing EspI and Map had significant colonization defects and resulted in dramatically reduced levels of hyperplasia, and they exhibited very different growth dynamics in mice than the wild-type strain exhibited.  (+info)

Clearance of Citrobacter rodentium requires B cells but not secretory immunoglobulin A (IgA) or IgM antibodies. (4/177)

Citrobacter rodentium, a murine model pathogen for human enteropathogenic Escherichia coli, predominantly colonizes the lumen and mucosal surface of the colon and cecum and causes crypt hyperplasia and mucosal inflammation. Mice infected with C. rodentium develop a secretory immunoglobulin A (IgA) response, but the role of B cells or secretory antibodies in host defense is unknown. To address this question, we conducted oral C. rodentium infections in mice lacking B cells, IgA, secreted IgM, polymeric Ig receptor (pIgR), or J chain. Normal mice showed peak bacterial numbers in colon and feces at 1 week and bacterial eradication after 3 to 4 weeks. B-cell-deficient mice were equally susceptible initially but could not control infection subsequently. Tissue responses showed marked differences, as infection of normal mice was accompanied by transient crypt hyperplasia and mucosal inflammation in the colon and cecum at 2 but not 6 weeks, whereas B-cell-deficient mice had few mucosal changes at 2 weeks but severe epithelial hyperplasia with ulcerations and mucosal inflammation at 6 weeks. The functions of B cells were not mediated by secretory antibodies, since mice lacking IgA or secreted IgM or proteins required for their transport into the lumen, pIgR or J chain, cleared C. rodentium normally. Nonetheless, systemic administration of immune sera reduced bacterial numbers significantly in normal and pIgR-deficient mice, and depletion of IgG abrogated this effect. These results indicate that host defense against C. rodentium depends on B cells and IgG antibodies but does not require production or transepithelial transport of IgA or secreted IgM.  (+info)

Protective role of arginase in a mouse model of colitis. (5/177)

Arginase is the endogenous inhibitor of inducible NO synthase (iNOS), because both enzymes use the same substrate, l-arginine (Arg). Importantly, arginase synthesizes ornithine, which is metabolized by the enzyme ornithine decarboxylase (ODC) to produce polyamines. We investigated the role of these enzymes in the Citrobacter rodentium model of colitis. Arginase I, iNOS, and ODC were induced in the colon during the infection, while arginase II was not up-regulated. l-Arg supplementation of wild-type mice or iNOS deletion significantly improved colitis, and l-Arg treatment of iNOS(-/-) mice led to an additive improvement. There was a significant induction of IFN-gamma, IL-1, and TNF-alpha mRNA expression in colitis tissues that was markedly attenuated with l-Arg treatment or iNOS deletion. Treatment with the arginase inhibitor S-(2-boronoethyl)-l-cysteine worsened colitis in both wild-type and iNOS(-/-) mice. Polyamine levels were increased in colitis tissues, and were further increased by l-Arg. In addition, in vivo inhibition of ODC with alpha-difluoromethylornithine also exacerbated the colitis. Taken together, these data indicate that arginase is protective in C. rodentium colitis by enhancing the generation of polyamines in addition to competitive inhibition of iNOS. Modulation of the balance of iNOS and arginase, and of the arginase-ODC metabolic pathway may represent a new strategy for regulating intestinal inflammation.  (+info)

Macroscopic, microscopic and biochemical characterisation of spontaneous colitis in a transgenic mouse, deficient in the multiple drug resistance 1a gene. (6/177)

1 A novel animal model of spontaneous colonic inflammation, the multiple drug-resistant (mdr1) a(-/-) mouse, was identified by Panwala and colleagues in 1998. The aim of our study was to further characterise this model, specifically by measuring cytokines that have been implicated in inflammatory bowel disease (IBD) (IL-8 and IFN-gamma) in the colon/rectum of mdr1a(-/-) mice, and by determining the sensitivity of these, together with the macroscopic, microscopic and disease signs of colitis, to dexamethasone (0.05, 0.3 and 2 mg kg(-1) subcutaneously (s.c.) daily for 7 days). 2 All mdr1a(-/-) mice had microscopic evidence of inflammation in the caecum and colon/rectum, while control mice with the same genetic background did not. Significant increases in colon/rectum and caecal weights and also in cytokine levels (both IFN-gamma and IL-8) in homogenised colon/rectum were observed in mdr1a(-/-) mice compared to mdr1a(+/+) mice. 3 Dexamethasone reduced the increases in tissue weights and also microscopic grading of colitis severity, but had no effect on IFN-gamma or IL-8. 4 This study supports the similarity of the gastrointestinal inflammation present in mdr1a(-/-) mice to that of human IBD, in particular Crohn's disease. This has been demonstrated at the macroscopic and microscopic levels, and was supported further by elevations in colonic levels of IFN-gamma and IL-8 and the disease signs observed. The incidence of colitis was much higher than previously reported, with all mice having microscopic evidence of colitis. The limited variance between animals in the parameters measured suggests that this model is reproducible.  (+info)

Impaired immunity to intestinal bacterial infection in stromelysin-1 (matrix metalloproteinase-3)-deficient mice. (7/177)

Infection of mice with the intestinal bacterial pathogen Citrobacter rodentium results in colonic mucosal hyperplasia and a local Th1 inflammatory response similar to that seen in mouse models of inflammatory bowel disease. Matrix metalloproteinases (MMPs) have been shown to mediate matrix remodeling and cell migration during tissue injury and repair in the intestine. We have previously shown enhanced pathology in infected TNFRp55-/-, IL-12p40-/-, and IFN-gamma-/- mice, and here we show that this is associated with an increase in stromelysin-1 (MMP3) transcripts in colonic tissues. We have therefore investigated the role of MMP3 in colonic mucosal hyperplasia and the local Th1 responses using MMP3-/- mice. In MMP3-/- mice, similar mucosal thickening was observed after infection as in wild-type (WT) mice. Colonic tissues from MMP3-/- mice showed a compensatory increase in the expression of other MMP transcripts, such as MMP7 and MMP12. However, MMP3-/- mice showed delayed clearance of bacteria and delayed appearance of CD4+ T lymphocytes into intestinal lamina propria. CSFE-labeled mesenteric lymph node CD4+ T lymphocytes from infected WT mice migrated in fewer numbers into the mesenteric lymph nodes and colon of MMP3-/- mice than into those of WT mice. These studies show that mucosal remodeling can occur in the absence of MMP3, but that MMP3 plays a role in the migration of CD4+ T lymphocytes to the intestinal mucosa.  (+info)

EspJ is a prophage-carried type III effector protein of attaching and effacing pathogens that modulates infection dynamics. (8/177)

Enterohemorrhagic Escherichia coli, enteropathogenic E. coli, and Citrobacter rodentium are highly adapted enteropathogens that successfully colonize their host's gastrointestinal tract via the formation of attaching and effacing (A/E) lesions. These pathogens utilize a type III secretion system (TTSS) apparatus, encoded by the locus of enterocyte effacement, to translocate bacterial effector proteins into epithelial cells. Here, we report the identification of EspJ (E. coli-secreted protein J), a translocated TTSS effector that is carried on the 5' end of the cryptic prophage CP-933U. Infection of epithelial cells in culture revealed that EspJ is not required for A/E lesion activity in vivo and ex vivo. However, in vivo studies performed with mice demonstrated that EspJ possesses properties that influence the dynamics of clearance of the pathogen from the host's intestinal tract, suggesting a role in host survival and pathogen transmission.  (+info)

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