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Bacterial Infections

Role for FimH in Extraintestinal Pathogenic Escherichia coli Invasion and Translocation through the Intestinal Epithelium

Nina M. Poole, Sabrina I. Green, Anubama Rajan, Luz E. Vela, Xi-Lei Zeng, Mary K. Estes, Anthony W. Maresso
Andreas J. Bäumler, Editor
Nina M. Poole
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Sabrina I. Green
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Anubama Rajan
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Luz E. Vela
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Xi-Lei Zeng
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Mary K. Estes
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Anthony W. Maresso
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Andreas J. Bäumler
University of California, Davis
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DOI: 10.1128/IAI.00581-17
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ABSTRACT

The translocation of bacteria across the intestinal epithelium of immunocompromised patients can lead to bacteremia and life-threatening sepsis. Extraintestinal pathogenic Escherichia coli (ExPEC), so named because this pathotype infects tissues distal to the intestinal tract, is a frequent cause of such infections, is often multidrug resistant, and chronically colonizes a sizable portion of the healthy population. Although several virulence factors and their roles in pathogenesis are well described for ExPEC strains that cause urinary tract infections and meningitis, they have not been linked to translocation through intestinal barriers, a fundamentally distant yet important clinical phenomenon. Using untransformed ex situ human intestinal enteroids and transformed Caco-2 cells, we report that ExPEC strain CP9 binds to and invades the intestinal epithelium. ExPEC harboring a deletion of the gene encoding the mannose-binding type 1 pilus tip protein FimH demonstrated reduced binding and invasion compared to strains lacking known E. coli virulence factors. Furthermore, in a murine model of chemotherapy-induced translocation, ExPEC lacking fimH colonized at levels comparable to that of the wild type but demonstrated a statistically significant reduction in translocation to the kidneys, spleen, and lungs. Collectively, this study indicates that FimH is important for ExPEC translocation, suggesting that the type 1 pilus is a therapeutic target for the prevention of this process. Our study also highlights the use of human intestinal enteroids in the study of enteric diseases.

INTRODUCTION

The gastrointestinal tract (GIT) not only functions in food digestion and excretion but also serves as an immunological barrier against microorganisms. However, under certain conditions, the bacteria residing in the GIT breach this barrier and migrate to extraintestinal sites, which can lead to gut-derived bacteremia and sepsis. The process by which bacteria and their products migrate from the GIT to the bloodstream and other organs is called “bacterial translocation,” which was first described as a clinical phenomenon by Wolochow and colleagues in the 1960s (1) and was investigated experimentally by Berg and coworkers in the 1980s (2–10). The latter set of studies defined several risk factors that contribute to translocation, including disruption of gut microflora by antibiotics (4, 6), damage or increased permeability of the intestinal epithelial barrier (4, 9, 11, 12), and the health status of the host (4, 5, 7, 8, 10–12). In addition, several studies have established an association between the intestinal microflora and systemic infections in immunocompromised individuals (13–18). Extraintestinal pathogenic E. coli (ExPEC), the general name prescribed to E. coli strains that cause infections distal to the intestine, is the most frequently isolated Gram-negative organism in such cases (14, 15, 17). ExPECs include E. coli associated with urinary tract infections (UTIs) (uropathogenic E. coli [UPEC]), neonatal meningitis (neonatal meningitis-associated E. coli [NMEC]), and septicemia (sepsis-causing E. coli [SEPEC]) (19–21). ExPEC strains are steadily acquiring resistance to commonly used antibiotics (22–25), and pandemic strains (sequence type 131) have been described previously (26).

ExPEC organisms are often indistinguishable from commensal organisms, colonize a sizable fraction of the healthy population (27), exhibit large genomic diversity, and carry multiple virulence factors in various combinations (21, 28, 29). Although several ExPEC virulence factors have been identified (21, 28, 29), they have not been directly linked to translocation from the intestine. This is problematic because knowledge of the bacterial factors that facilitate translocation would constitute attractive targets for the development of antitranslocative medicines. This study aims to fill this void by identifying the ExPEC factors involved in the early stages of translocation. Our findings establish a role for the mannose-binding type 1 pilus tip protein FimH in ExPEC invasion and translocation.

RESULTS

ExPEC invades the intestinal epithelium.Gut-derived bacteremia and sepsis originate from the movement of enteric bacteria from the intestinal lumen to the bloodstream, whereby a systemic infection and/or excessive immune response facilitates a life-threatening condition (13, 14). Passive translocation results when there is a physical break in the GI wall and intestinal contents, including bacteria, leach into the lymphatics and circulation (30). In contrast, active translocation occurs when the inherent properties of the bacteria allow for their passage through a GI barrier in the absence of evidence of pathology or trauma to the epithelial wall (31). We hypothesized that ExPEC would pass through an intact intestinal epithelium. To test this hypothesis, we adapted the use of a confluent and polarized Caco-2 monolayer, a system sometimes used to measure the invasion of Listeria (32, 33) and Salmonella (34), to measure that of ExPEC. This system allows the investigator to measure three fundamental steps in the process of translocation: adherence, invasion, and the actual movement through an intact intestinal barrier.

We first tested if ExPEC will adhere to Caco-2 cells. ExPEC strain CP9 and E. coli laboratory strain MG 1655 (K-12) were incubated with Caco-2 cells for 1 h, the monolayer was washed, and bacterial cells were enumerated by plating on Luria-Bertani agar (LB/agar). As demonstrated in Fig. 1A, both strains of E. coli adhered to Caco-2 cells. When gentamicin was added for 2 h to kill extracellular bacteria and the monolayer was assessed for the presence of E. coli (invasion), there was an average of 10 times more ExPEC than K-12 organisms (multiplicities of infection [MOIs], 10 and 50; P ≤ 0.05) (Fig. 1B). When mock infections in which the only omission from the experiment was the presence of the Caco-2 cells were performed, neither ExPEC nor K-12 survived.

FIG 1
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FIG 1

ExPEC adheres to and invades Caco-2 cell monolayers. (A) ExPEC strain CP9 and E. coli laboratory strain K-12 were seeded at MOIs of 10 and 50, and bacterial adherence was measured as described in Materials and Methods. (B) Bacterial invasion was measured by gentamicin protection assay at MOIs of 10 and 50 (see Materials and Methods). Data are means ± SEM; n = 4 to 6, assayed in triplicate; *, P ≤ 0.05.

ExPEC invasion is dependent on the type 1 pilus adhesin FimH.We reasoned that virulence factors that are important for urinary tract infection may be a starting point for identifying the bacterial factors important for the invasion and translocation through the intestinal epithelium. Therefore, we assessed if iroN, cnf-1, or fimH is necessary for ExPEC's invasion and translocation through intestinal epithelial cells. The iroN gene encodes the salmochelin siderophore receptor, which is important for iron uptake in iron-limited environments (35, 36), cnf-1 encodes a potent cytotoxin that induces the rearrangement of the actin cytoskeleton (37), and fimH encodes a type I pilus tip adhesin protein (38). In particular, FimH and allelic variants of this protein are important for the invasion of the urinary epithelium, as well as the formation of intracellular bacterial communities (IBCs), a persistent intracellular infection that forms a reservoir of UPEC that is a source of recurring infections in the urinary tract (38–42).

We used recombineering and transduction to generate deletions of iroN, cnf-1, and fimH in strain CP9, and these strains were evaluated for invasion and translocation using the Caco-2 cell system described above. Wild-type, ΔiroN, and Δcnf-1 strains all adhered (see Fig. S1A in the supplemental material) and invaded (Fig. 2A) the Caco-2 cells with similar efficiency, independent of the MOI used. The fimH mutant strain's ability to invade and adhere was reduced at all MOIs, but this reduction was 3-fold and 6-fold at the higher MOIs, respectively (Fig. 2A, P ≤ 0.05; Fig. S1A). To further evaluate the role of the mannose-sensitive adhesin FimH on ExPEC adhesion and invasion, Caco-2 cells were infected with ExPEC CP9 or E. coli K-12 at an MOI of 50 in the presence of methyl α-d-mannopyranoside (mannoside), or methyl α-d-glucopyranoside (glucoside). The mannoside reduced the percentage of ExPEC associated with the Caco-2 cells compared to the untreated control (P ≤ 0.05) (Fig. S1C; ExPEC+mannoside), whereas the total number of intracellular bacteria was reduced almost 5-fold, indicating that the invasive phenotype is indeed partially due to the loss of FimH (P ≤ 0.05) (Fig. 2B, ExPEC+mannoside).

FIG 2
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FIG 2

ExPEC invasion is partially dependent on the type 1 mannose-sensitive pilus adhesin FimH. (A) Caco-2 cells were infected with ExPEC wild-type (Wt) ΔfimH, ΔiroN, or Δcnf-1 strain, and invasion was evaluated at MOIs of 10 and 50 (see Materials and Methods). (B) Caco-2 cells were infected with ExPEC strain CP9 or E. coli K-12 at an MOI of 50 in the presence of mannoside or glucoside, and invasion was measured (see Materials and Methods). Data are means ± SEM, n = 3 to 6, assayed in triplicate; *, P ≤ 0.05.

Cultured transformed cancer cell lines, similar to the Caco-2 cells used in this study, are often the default infection model system for the study of enteric pathogens. To provide a more physiologic model system to further test the hypothesis that ExPEC adheres to and invades the intestinal epithelium, a process dependent on FimH, we developed the use of organotypic human intestinal enteroids to measure ExPEC invasion. Human enteroids are untransformed, composed of all the major cell types found in villus intestinal epithelium (enterocytes and Paneth, goblet, enteroendocrine, and stem cells) (43), display physiological responses upon infection (44–46), and can be grown in two dimensions (2-D) or 3-D. Here we used monolayers generated from multilobular 3-D human jejunal intestinal enteroids (HJIEMs) to assess the invasive capacity of ExPEC (see Materials and Methods). Wild-type, ΔfimH, or fimH-complemented ExPEC strains were applied (MOI, 100) to HJIEMs, and adhesion and intracellular invasion were measured. Whereas all ExPEC strains adhered to the HJIEMs (Fig. 3A), the strain lacking fimH was reduced by approximately 1 log in its ability to invade, a phenotype that was partially complemented with expression of FimH from a plasmid (Fig. 3B, ΔfimH) (P ≤ 0.05). Consistent with a role for FimH in this process, the invasion of enteroids was also inhibited by the addition of mannose (Fig. 3C, ExPEC with mannose) (P ≤ 0.05). These results mirror very closely those observed for wild-type ExPEC and the interaction with Caco-2 intestinal epithelial cells.

FIG 3
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FIG 3

FimH mediates ExPEC invasion of human jejunal intestinal enteroid monolayers (HJIEMs). HJIEMs were infected at an MOI of 100 with ExPEC wild-type (Wt) or ΔfimH strain or pUC19:ΔfimH ExPEC strain CP9, and adherence (A) and invasion (B) were measured (see Materials and Methods). (C) The effects of mannose on wild-type (Wt) ExPEC and ΔfimH mutant strains were evaluated as described in Materials and Methods. Data are means ± SEM; n = 4 to 8, assayed in triplicate; *, P ≤ 0.05.

ExPEC translocation through a polarized intestinal monolayer is mediated by FimH.The finding that ExPEC could invade intestinal epithelial cells suggests that it may also be able to translocate through this barrier. We first tested if a 6-hour incubation of ExPEC with Caco-2 cells leads to a loss of intestinal cell barrier integrity as measured by a potential reduction in the transepithelial electrical resistance (TER). Polarized Caco-2 monolayers normally have resistances above 200 (Ω cm2) when grown on Transwell inserts (47, 48). At an MOI of 10, the TER of Caco-2 cells remained unchanged throughout the experiment for both ExPEC and E. coli K-12 and did not differ from cells that were not infected (Fig. 4A). However, at an MOI of 50 and starting about 3 h into the infection, the TER of the K-12 cells slightly increased with time, whereas that of the ExPEC-infected cells was reduced, a finding not observed for the uninfected control (Fig. 4B, ExPEC and K-12) (P ≤ 0.05). These results suggest that ExPEC strain CP9 does not disrupt the integrity of the epithelium at lower infection doses, such as an MOI of 10. It also indicates that we can test if ExPEC is capable of translocating through these cells in the absence of the physical disruption of the monolayer.

FIG 4
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FIG 4

ExPEC translocation through a polarized intestinal monolayer is mediated by FimH. The transepithelial electrical resistance (TER) was measured at an MOI of 10 (A) or 50 (B) for Caco-2 cells infected with ExPEC strain CP9 or E. coli K-12 (see Materials and Methods). (C) Caco-2 cells were infected with strain ExPEC CP9 or E. coli K-12 at an MOI of 10, and levels of translocation were measured 6 h postinfection as described in Materials and Methods. (D) Translocation levels for wild-type (Wt) and ΔfimH, ΔiroN, and Δcnf-1 mutant strains at an MOI of 10 were evaluated after 6 h. Data are means ± SEM; n = 3 to 6, assayed in triplicate; *, P ≤ 0.05; ***, P ≤ 0.001.

To test this hypothesis, Caco-2 cells were grown on a Transwell insert seeded for 21 days until polarized (as measured by TER). ExPEC or E. coli K-12 (MOI, 10) was added to the apical surface (top chamber) of the monolayer for a 6-h incubation, and passage of bacteria to the basolateral side (bottom chamber) was measured by plating the medium on LB/agar. As demonstrated in Fig. 4C, some level of basal translocation was observed for E. coli K-12; however, approximately 2 and 3 orders of magnitude more ExPEC isolates passed from the apical to the basolateral side at the later time points, respectively (Fig. 4C, 4 h, P ≤ 0.05; 6 h, P ≤ 0.001). Collectively, these findings indicate that ExPEC can translocate through a polarized epithelial layer of intestinal origin grown on Transwell inserts. The finding that ExPEC, a leading cause of human bacteremia and gut-derived sepsis, can both invade and pass through an intestinal epithelium is a novel observation for this pathogen. Having identified FimH as a determinant of ExPEC's invasion, we next assessed the ExPEC mutant strains' ability to translocate through Caco-2 cells seeded on a Transwell system with apical and basolateral compartments. The mutant strains lacking iroN and cnf-1 migrated across the Caco-2 cell monolayer as well as the wild-type strains after 6 h; however, ExPEC lacking fimH did translocate at approximately 1 order of magnitude less than the wild type (P ≤ 0.05) (Fig. 4D, 4 and 6 h ΔfimH).

FimH is important for ExPEC translocation in vivo.Berg and colleagues defined translocation as the movement of bacteria from the intestinal lumen to extraintestinal sites, including the mesenteric lymph nodes and major organ systems (2–4). The Caco-2 and enteroid cell culture models indicate that ExPEC can invade and pass through an intact intestinal epithelium in a process partially dependent on FimH. To further test this hypothesis, we utilized a murine model of chemotherapy-induced ExPEC translocation that we developed (49) to evaluate if the loss of fimH affected ExPEC translocation during a developing infection. Mice were subjected to gavage with the wild-type, ΔfimH mutant, or fimH-complemented strain, colonization was established, and chemotherapy was administered as described previously (49). As shown in Fig. 5A, the levels of each mutant ExPEC strain shed into the feces from days 1 to 7 were comparable to wild-type levels, with a characteristic drop in the levels of colonized ExPEC after day 1 as previously observed (49). At 8 days, the mice were sacrificed and necropsied, and tissues were examined for the levels of ExPEC. Both the small and large intestines had a mean level of ExPEC ΔfimH that was slightly lower than the wild-type and the complemented strain levels, but this was not significant (Fig. 5B and C). When assessed for translocation, the fimH mutant displayed on average ∼1-log reduction in the mean CFU per gram of tissue in the kidneys, spleen, and lungs (kidneys, ΔfimH strain, P ≤ 0.05; spleen, ΔfimH strain, P ≤ 0.05; lungs, ΔfimH strain, P ≤ 0.05) (Fig. 5D to F). This phenotype was restored with the fimH-complemented strain (kidneys, ΔfimH strain, P ≤ 0.05; spleen, ΔfimH strain, P ≤ 0.01; lungs, ΔfimH strain, P ≤ 0.01) (Fig. 5D to F). Although there was a definite decrease in the levels of the fimH mutant strain in the liver and these levels were restored with the complemented strain, these trends were not significant (Fig. 5G). To determine if the higher numbers of wild-type isolates found in the kidneys, spleen, and lungs were due to some tropism for these tissues, we performed an intraperitoneal infection with the wild-type or fimH mutant strain into animals and assessed the levels of ExPEC in tissues. There was no difference in the levels of ExPEC in each organ between the wild-type and mutant strains (Fig. 5H).

FIG 5
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FIG 5

FimH is important for ExPEC translocation of an immunocompromised host. (A to G) BALB/c mice were subjected to oral gavage with wild-type (Wt), fimH mutant, or complemented pUC19:ΔfimH ExPEC strains at 109 CFU/100 μl. Mice were given cyclophosphamide at a total dose of 450 mg/kg (three 150-mg/kg doses on days 1, 3, and 5) by intraperitoneal injection. Intestinal colonization was measured by fecal homogenates (days 1, 3, 5, and 7) (A) or intestinal homogenates (8 days postinfection immediately after euthanasia) (B, C) plated on LB/agar plus 10 μg/ml CM. (D to G) Translocation out of the GI tract was determined by the presence of bacteria in organs as described in Materials and Methods. Data are means ± SEM; n = 14 to 16 for each group. *, P ≤ 0.05; **, P ≤ 0.01. (H) For an intraperitoneal infection, BALB/c mice were given 107 CFU/100 μl of wild-type (Wt) ExPEC or fimH mutant strains. Colonization was measured by organ homogenates plated on LB/agar plus 10 μg/ml CM (see Materials and Methods). Data are means ± SEM; n = 8 for each group.

DISCUSSION

Bacterial translocation occurs in healthy individuals and is very important for the differentiation and maturation of the host's immune system (50). However, in those with weakened immune systems, either because of severe illness or medical treatments that render the patient immunocompromised, or because gut barrier integrity has been disrupted, the potential for translocation of resident enteric bacteria into the blood is high (13–18). Sepsis is responsible for approximately 1.6 million hospital stays in the United States per year (11). Although not all cases of sepsis originate from gut enterics, the most frequently isolated Gram-negative bacterium from the blood of such patients is E. coli (17, 51, 52), generally classified as ExPEC. Particularly concerning is the notion that humans, pets, or livestock may be asymptomatic carriers of these bacteria and thus constitute a hidden reservoir for high-risk individuals, with evidence of transmission between family members (53, 54). There are major knowledge gaps in our understanding of the bacterial factors that promote transmission, colonization, and the translocation of these bacteria. The results presented in this study advance this field by demonstrating that (i) ExPEC adheres to and invades a polarized Caco-2 intestinal monolayer as well as monolayers derived from human intestinal enteroids, (ii) this process is dependent on FimH, (iii) ExPEC translocates through polarized intestinal epithelial cells seeded onto a Transwell insert, (iv) translocation is also FimH dependent, and (v) FimH mediates translocation of ExPEC from the intestine to the peripheral organs upon immunosuppression in a murine host.

ExPEC, like most members of the Enterobacteriaceae family, produces type 1 pili, which aid in the attachment to mucosal epithelial surfaces. These pili are encoded by the fim operon, which consists of nine genes that are transcriptionally controlled by phase variation of an invertible promoter element (reviewed in reference 55). FimH, the tip protein, is well known to facilitate adhesion to the urinary epithelium and be a prominent player in UPEC pathogenicity (56–62). Adhesion via FimH is largely driven by the direct binding to mannose moieties on receptors expressed on the surface epithelium. The mannose-binding pocket of FimH does not vary among sequenced UPEC strains (63, 64), but residues outside this pocket can influence binding affinity and are evolving under positive selection in clinical UPEC strains (64). FimH is also required for UPEC to enter the bladder epithelium and persist in the form of IBCs (38–42). Because of its role in UPEC virulence, several therapeutic strategies have targeted FimH, including vaccines (65, 66), mannosides as competitive inhibitors of FimH binding (67–69), and compounds that inhibit FimH assembly (70), some of which can treat chronic cystitis caused by circulating UPEC strains in animal models (71). Our finding that FimH is also important for the early stages of translocation now implicates this pilus protein in the onset of gut-derived bacteremia. Since the GIT is the likely initial reservoir for these pathogens, future work should give strong consideration to the testing and use of these therapeutic measures in the GIT microenvironment as a means to prevent translocation.

It is well established that the genetic heterogeneity of E. coli as a species allows this pathogen to infect a variety of different human tissues, resulting in vastly different clinical outcomes or conditions. The combination of distinct virulence factors and antibiotic resistance elements, driven together primarily by horizontal transfer mechanisms, generates unique E. coli strains that may be not only more virulent but also selected for upon antibiotic treatment. The extensive pangenome of E. coli, which may contain as many genes as the human genome itself, offers a plethora of gene combinations that provide for an astounding level of adaptability (28). In addition to this versatility, there is now a growing body of evidence that virulence factors also harbor intergene allelic heterogeneity that may confer special pathoadaptive features to E. coli, a finding that has been observed with FimH and may have direct implications on the topic presented in this report (72, 73). For example, in addition to the fim operon capable of being acquired by horizontal transfer, this region of the genome undergoes de novo mutation at a higher rate than that of housekeeping genes, of which some changes seemingly enhance the binding of FimH to mannosylated receptors (74). Reports suggest that upwards of 30% of fimH genes harbor a different allele, a level of genetic diversity that is the basis for organizing strains into distinct sequence types (75, 76). Different FimH variants display different binding affinities to monomannose moieties (77), and these differences are factors in determining the environmental niche, intestinal or urinary, that E. coli may preferentially occupy, possibly by enhancing colonization, especially in the urinary tract (72). More-recent work suggests that some fimH alleles are undergoing positive selection that leads to enhanced fitness (64), with at least some of these alleles altering pilus tip conformation without directly acting on mannose contact residues (78). Not well understood at this point is whether different fimH alleles, which confer an altered functionality on the pilus tip protein, also influence the colonization or translocation of ExPEC through the gastrointestinal epithelium. Given the preponderance of different alleles in circulating ExPEC strains, the segmentation of certain alleles associated with highly virulent and resistant superbugs (74), and the already important role for them in uropathogenesis, understanding the effect of these alleles in gut-derived sepsis would appear to be a pressing need. It has been recently shown, using ExPEC strain UTI89 in a murine model, that type 1 and F-17-like pili enhance intestinal colonization (79). M4284, a high-affinity biphenyl mannoside that targets FimH, when used to treat a UTI also lowered intestinal colonization (79). In this study, colonization of the intestines by ExPEC CP9 lacking FimH was slightly lower on average than that by the wild type, but the greatest attribute hampered was its ability to translocate. Both studies highlight the gut as a reservoir for ExPEC infection and FimH as a therapeutic target to combat these infections.

The two issues left unresolved in this work are the determination of the mechanism by which ExPEC enters and translocates through the intestinal epithelium and the other virulence factors that induce this process. Understanding this mechanism and identifying the additional key determinants are of the utmost importance, since knowledge of this process can be harnessed to design smart drugs that prevent it. The data presented here suggest that ExPEC can enter intestinal epithelial cells, a process that is in part but not exclusively FimH dependent. One may put forth three general models that may explain how enteric commensals or pathobionts translocate from the GI lumen to the blood in immunocompromised patients, none of which are mutually exclusive. The first involves overt destruction of the intestinal epithelium and complete loss of barrier integrity. In this scenario, a reduced mucin layer and/or compromised epithelial junctions provide a clear path for commensals to reach the basolateral compartment. Barrier loss can be induced by medical treatments or trauma; for example, some cancer chemotherapies lead to increased permeability in the intestinal epithelium (80). Barrier loss may also be bacterially induced, and there is growing evidence that E. coli makes enzymes that “chip away” at host barriers. For example, ExPEC produces a protein termed surface and secreted lipoprotein E (SslE), which degrades mucin and is very important for translocation in a murine chemotherapy-induced model of gut-derived sepsis (49). Many clinical strains of E. coli also make α-hemolysin, a secreted lipase, which is reported to generate intestinal “focal leaks,” small holes in an otherwise-intact epithelium through which bacteria can paracellularly traverse enterocytes (81). In the two culture model systems (Caco-2 and HJIEMs) reported here, there is evidence of epithelial cell destruction (as measured by a drop in TER), but only at high MOIs and later time points postinfection. At least for ExPEC strain CP9, adherence, invasion, and translocation occur before a drop in TER. Nevertheless, the finding that SslE and hemolysin may aid in translocation, along with a role for FimH as defined in this report, implies that at least some of the translocation is a specific process mediated by distinct virulence factors.

A second potential way enteric bacteria may access the underlying lamina propria is if they are taken up and brought there by host immune cells such as dendritic cells (82) or enter through specialized enterocytes within the follicle-associated epithelium of the Peyer's patches (M cells) (83). Since glycoprotein 2 (GP2) can mediate the binding of FimH-containing bacteria and the subsequent transcytosis from the apical to basolateral compartment (83), our finding that FimH is important for translocation is consistent with this observation. Shigella species, which are closely related to E. coli, are thought to enter M cells to access the underlying lamina propria (84). Finally, a third potential mechanism is that ExPEC squeezes between cells to access the basolateral compartment. In this route, referred to as paracellular invasion and commonly associated with Campylobacter jejuni, it is thought that the pathogen disrupts tight and adherens junctions to facilitate transmigration (85).

The discussion of the mechanism of translocation must consider the proposed receptor or receptors needed to adhere to the intestinal epithelium. In the case of UPEC and the urothelium, FimH binds to uroplakins, mainly UP1a, found on the apical surface of the umbrella cells lining the lumen (86–88). The FimH-UP1a-dependent invasion of urothelial cells involves lipid rafts and its components caveolin-1 (89) and Rac1 (90). Alternatively, UPEC can also invade the urothelium in a zipper-like fashion caused by FimH interacting with α3β1 integrin, which activates a downstream signaling cascade involving focal adhesion, Src, and phosphoinositide 3-kinases, Rho-family GTPases, α-actinin, vinculin, and clathrin (38, 91, 92). This actin-dependent entry into the host cell can also be modulated by microtubules (93). After the FimH-induced internalization, UPEC can be found in exocytic vesicles (94) or in the cytoplasm, where it rapidly replicates to form IBCs supporting the establishment of a persistent infection (38–42). It appears that uroplakins are highly specific to urothelium; therefore, it is more than plausible that FimH-mediated invasion of intestinal epithelial cells involves integrin receptors; and we have preliminary data to support this hypothesis (unpublished data). There still remain several unanswered questions that warrant further investigation; however, our work has begun to unveil how ExPEC virulence determinants contribute to gut-derived sepsis.

MATERIALS AND METHODS

Bacterial strains.In this study, we used ExPEC strain CP9 (serotype O4:K54:H5) (95), a blood isolate from a bacteremic patient (96, 97), generously provided by James Johnson (University of Minnesota). This strain was selected because its virulence in mouse models is well characterized, some virulence factors for survival have been identified (97–100), and Green et al. (49) recently showed that it can translocate out of the mouse intestinal tract.

CP9 and K-12 MG1655 (provided by Christophe Hermann, Baylor College of Medicine) were modified through the addition of a chloramphenicol (CM) resistance and green fluorescence protein (GFP) cassette by classical P1 phage transduction. The same procedure was used to generate a knockout of fimH (ΔfimH) in ExPEC CP9 (referred to as the ΔfimH mutant). The complementing strain (pUC19:ΔfimH) was generated by amplifying the fimH gene using isolated wild-type CP9 chromosomal DNA using primers FimH1-FWD (5′-GTATACGGATCCATGAAACGAGTTATTACCC-3′), which included a BamHI site, and FimH2-REV (5′-GTATACGAATTCTTATTGATAAACAAAACGTCAC-3′), which included an EcoRI site. The product was cloned into pUC19 (New England BioLabs, Ipswich, MA, USA) and transformed into chemicompetent E. coli DH5-α cells (New England BioLabs, Ipswich, MA, USA) selected for on Luria-Bertani agar (LB/agar) with 100 μg/ml ampicillin (AMP; EMD Millipore, Darmstadt, Germany) coated with X-Gal (5-bromo-4-chloro-3-indolyl-β-d-galactopyranoside) and IPTG (isopropyl-β-d-1-thiogalactopyranoside; Sigma-Aldrich, St. Louis, MO, USA) for blue-white colony screening. The white colonies were screened using M13 forward and reverse primers for pUC19. The vector carrying fimH was isolated and then used to transform electrocompetent ΔfimH cells, and FimH was expressed by IPTG induction.

CP9 ΔiroN and Δcnf-1 mutants were generated by recombineering utilizing the λ red system. The wild-type strain was transformed with AMP-resistant pKD46, which harbors the λ red recombinase enzymes, and selected for accordingly. PCR products were made using primers that corresponded to upstream and downstream regions flanking the gene of interest and FLP recombination target (FRT) sites using pBA169CM:FRT as the template DNA (IroN1-FWD, 5′-CTCCAGCCTTCGGAAGCCAGAAGAATGGCTTTAATACGATCATATGAATATCCTCCTTAG-3′); IroN2-REV, 5′-CCGCTTTTACGCGGGCAGTGCGCCTGAAACACTACGATCAGTGTAGGCTGGAGCTGCTTC-3′; CNF-11-FWD, 5′-CATATGAATATCCTCCTTAGCCCGAAATACTGTATTTCTCAGCCATCAGTACAGCACTTTCATATGAATATCCTCCTTAG-3′; and (CNF-12-REV, 5′-GTGTAGGCTGGAGCTGCTTCCAGATCGCAGCACTGGAGCGTAAAGCCAGTGATGACGAGGGTGTAGGCTGGAGCTGCTTC-3′). These products were purified, digested, and transformed into electrocompetent cells grown in the presence of l-arabinose to induce recombinase enzymes. Transformed colonies were selected for on LB/agar with 10 μg/ml CM (EMD Millipore, Darmstadt, Germany), isolated, and screened for the recombineered product by PCR using amplifying primers complementary to regions upstream and downstream of the CM:FRT insertion site: IroN3-FWD (5′-ACAAACGCAATAACATCGCG-3′), IroN4-REV (5′-CCACCCTCACTATACTGAAA-3′), CNF-13-FWD (5′-GAAATACTGTATTTCTCAGC-3′), and CNF-14-REV (5′-CTCCAGTGCTGCGATCTGG-3′). After mutants were confirmed, pDK46 was removed from each strain by incubating streak plates overnight at 42°C. Colonies were sequentially tested for AMP sensitivity after pDK46 removal. For experiments, strains were grown from a single isolate in Luria broth at 37°C with aeration and diluted to the appropriate concentration.

Cell and tissue culture.Caco-2 colonic cells (clone C2BBe1; ATCC CRL-2102) were maintained in 75-cm2 flasks in Dulbecco's modified Eagle's medium (DMEM; Mediatech, Inc., a Corning subsidiary, Manassas, VA, USA) supplemented with 10% fetal bovine serum (FBS; Atlanta Biologicals, Flowery Branch, GA, USA) and a 1% mixture containing 100 U/ml penicillin–100 μg/ml streptomycin (Thermo Fisher Scientific, Boston, MA, USA). Cells were subcultured weekly using Ca2+/Mg2+-free Hanks balanced salt solution (Mediatech, Inc., a Corning subsidiary, Manassas, VA, USA) and 0.25% trypsin–0.2% EDTA (Mediatech, Inc., a Corning subsidiary, Manassas, VA, USA). Then, cells were seeded at a density of 5 × 104 cells/ml at 37°C in the presence of 5% CO2 in a humidified incubator.

Human jejunum intestinal enteroids (HJIE) were derived from jejunal intestinal crypts extracted from tissues obtained from patients undergoing bariatric surgery biopsies using the protocol of Sato et al. (43) and as previously described (44). Briefly, minced intestinal tissue was prepared and placed in a 6-well plate with approximately 3 ml of complete chelating solution (CCS; consisting of 100 ml incomplete chelating solution [500 ml Milli-Q H2O, 2.49 g Na2HPO4 · 2H2O, 2.7 g KH2PO4, 14 g NaCl, 0.3 g KCl, 37.5 g sucrose, and 25 g d-sorbitol], 400 ml Milli-Q H2O, and 40 mg DL-dithiothreitol) with 120 μl of 0.5 M EDTA, and the plates were orbitally shaken (∼250 rpm) at 4°C for 30 min. Two milliliters of FBS was added and mixed by pipetting, and the tissue was allowed to settle. Crypts were isolated as described previously (44) and resuspended in 2 ml of complete medium without growth factors (CMGF−), which consisted of advanced Dulbecco's modified Eagle medium/F-12 (Gibco, Carlsbad, CA, USA) supplemented with 1% 100 U/ml penicillin–100 μg/ml streptomycin, 1% 1 M HEPES (Thermo Fisher Scientific, Boston, MA, USA), and 1% 100× GlutaMAX (Thermo Fisher Scientific, Boston, MA, USA), followed by a final spin at 200 × g for 5 min at 4°C. The supernatant was removed, and the pellet containing the crypts was resuspended in growth factor and phenol red-free BD Matrigel basement membrane matrix (BD Biosciences, San Jose, CA, USA) on ice to avoid solidification (30 μl). Once plated, the Matrigel-enteroid culture was allowed to solidify at 37°C for 10 to 15 min, and the cultures were overlaid with 500 μl of complete media with growth factors (CMGF+, described below). All CMGF reagents were purchased from Thermo Fisher Scientific, Boston, MA, USA, or R&D Systems, Inc., Minneapolis, MN, USA. Complete medium with growth factors (CMGF+) consisted of CMGF− with 100 ng/ml of epidermal growth factor (EGF), 20% R-spondin-conditioned medium (Trevigen cell line HA-R-Spondin-Fc293T), 10% Noggin conditioned medium produced according to the methods described by Heijmans et al. (101), 50% Wnt3 conditioned medium (ATCC L-Wnt3 cell line CRL-2647), 1× N2 supplement, 1× B27 supplement, 1 mM n-acetylcysteine, 10 nM gastrin, 10 mM nicotinamide, 500 nM A83 (transforming growth factor beta 1 [TGF-β1] type-I receptor inhibitor), and 10 μM SB202190 (P38 inhibitor). The protocol for generating HJIE monolayers (HJIEMs) was modified from VanDussen et al. (102) and established by Estes's laboratory (103). Briefly, a 2.5% solution of Matrigel in cold phosphate-buffered saline (PBS) was overlaid into the wells of a 96-well plate. Four-day-old undifferentiated HJIEs were mixed with 0.05% trypsin–EDTA to promote the disruption of the enteroid into individual cells, the mixture was filtered through a 40-μm cell strainer to remove large aggregated cell structures, and the cell suspension in CMGF+ with 10 μM Y-27632 (ROCK inhibitor; Sigma-Aldrich, St. Louis, MO, USA) was added to the matrigel-coated wells to promote the formation of an enteroid monolayer. After 1 day, the medium was changed to differentiating medium (CMGF+ without R-spondin, Wnt3, nicotinamide, and SB202190) to promote differentiation and formation of the intact and polarized monolayer.

Bacterial adhesion and invasion assays.For adhesion (modified from reference 32) and invasion (modified from reference 33) assays, Caco-2 cells were seeded in 24-well plates at a density of 5 × 104 cells/well. Cells were grown until visible dome formation, which indicates differentiation based on the unidirectional movement of water and ions (104). Monolayers were washed once with PBS to remove remnants of antibiotic from culture medium, and then DMEM with 10% FBS with or without 1% d-mannose, methyl α-d-mannopyranoside (mannoside), or methyl α-d-glucopyranoside (glucoside) (Sigma-Aldrich, St. Louis, MO, USA) was added. Diluted overnight bacterial cultures were washed once with PBS, resuspended in DMEM plus 10% FBS, and added to Caco-2 monolayers at a multiplicity of infection (MOI) of 1, 10, or 50, followed by incubation for 1 h at 37°C in the presence of 5% CO2 in a humidified incubator. To quantify adherent bacteria, monolayers were washed 2 times with PBS and lysed with cold PBS containing 0.1% Triton X-100, and bacteria were enumerated by plating on LB/agar plus 10 μg/ml CM. For invasion, monolayers were washed as previously described and subsequently incubated in DMEM plus 10% FBS containing 50 μg/ml gentamicin (Santa Cruz Biotechnology, Inc., Dallas, TX, USA) for an additional 2 h to kill extracellular bacteria. Caco-2 monolayers were washed and lysed, and intracellular bacteria were enumerated as mentioned above. The HJIEMs also were used to determine bacterial internalization. The HJIEMs were seeded as noted above in the protocol and infected with an MOI of 100.

Transepithelial bacterial translocation.Bacterial translocation (modified from reference 34) was determined by seeding Caco-2 cells at 1 × 105 cells/Transwell insert (filter pore size, 3.0 μm in diameter; BD Biosciences, San Jose, CA, USA) in 24-well plates and allowing differentiation for 21 days to form a tight polarized monolayer. To check the integrity of the monolayer, transepithelial electrical resistance (TER) was measured (Voltmeter; Millipore, Billerica, MA, USA), and the monolayers with a TER of at least 200 Ω cm2 were used (47, 48). The wells and inserts were washed once with PBS, and then DMEM with 10% FBS was added to the top and bottom chambers. Diluted overnight bacterial cultures were washed once with PBS, resuspended in DMEM–10% FBS, and added apically to the Caco-2 monolayers at an MOI of 10 or 50. To determine the number of translocated bacteria after 1, 2, 4, and 6 h, medium from the bottom chamber was removed, and the bacteria were enumerated by plating on LB/agar with 10 μg/ml CM. During the infection, the effect of bacteria on monolayer integrity was monitored by TER measurements at times 0, 3, and 6 h.

Murine model of translocation.Our lab has previously established a murine model of chemotherapy-induced ExPEC translocation (49), which we used here to evaluate the role of FimH in this process. Briefly, 8-week-old male and female BALB/c mice were infected with 1 × 109 CFU ExPEC CP9 wild-type, ΔfimH, and pUC19:ΔfimH strains. Mice infected with pUC19:ΔfimH were also given water containing 10 mM IPTG (changed on the 4th day postinfection) to induce and maintain FimH expression (105). The day of the experiment (day 0), the cultures were centrifuged at 15,000 rpm, washed twice, and suspended in PBS. Each mouse (n = 14 to 16 per group) was subjected to gavage with a sterile (20-gauge, 38-mm) flexible needle that contained 1 × 109 CFU/100 μl. Cyclophosphamide (Baxter Healthcare Corporation, Deerfield, IL, USA) was dissolved in sterile water at a concentration of 10 mg/ml, and the mice were given a total dose of 450 mg/kg of body weight (3 × 150-mg/kg doses on days 1, 3, and 5) by intraperitoneal injection. Colonization was measured by plating fecal homogenates obtained on days 1, 3, 5, and 7. On day 8, mice were euthanized with CO2 (2 liters/min, 5 min), and organs (lungs, spleen, kidneys, liver, small intestine, and large intestine) were weighed, homogenized, and serially diluted. For the intraperitoneal infection, each mouse was given 107 CFU/100 μl of wild-type or fimH mutant strains, and the extent of dissemination was assessed 16 h postinfection as described above. All experiments were done with approval by Baylor College of Medicine’s Institutional Animal Care and Use Committee.

Statistical analysis.Adhesion, invasion, and translocation experiments data are means ± standard errors of the means (SEM) of assays (n = 3 to 8) performed in triplicate and over several passages of cells. For the chemotherapy-induced murine model of translocation, each group (3 groups) had 14 to 16 animals, and data are means ± SEM. The intraperitoneal murine model of infection had 8 animals for both groups, and data are means ± SEM. Statistical significance was determined by the Kruskal-Wallis and two-tailed unpaired t tests; differences between groups were assessed by Dunn's and Mann-Whitney test employing GraphPad Prism version 7.03 Windows (GraphPad Software, San Diego, CA; www.graphpad.com ). Differences in means were considered significant at P levels of ≤0.05.

ACKNOWLEDGMENTS

We thank James R. Johnson from University of Minnesota Medical School for donating the ExPEC strain CP9 and Christophe Herman from Baylor College of Medicine for donating the K-12 strain MG1655. We also thank Mary Girard for her assistance in developing mutant strains using recombineering and P1 phage transduction. Our sincere gratitude to Joseph Hyser, Sarah Blutt, and Noah Shroyer from Baylor College of Medicine for their help in generating monolayers from human enteroids.

FOOTNOTES

    • Received 10 August 2017.
    • Accepted 11 August 2017.
    • Accepted manuscript posted online 14 August 2017.
  • Supplemental material for this article may be found at https://doi.org/10.1128/IAI.00581-17 .

  • Copyright © 2017 American Society for Microbiology.

All Rights Reserved .

REFERENCES

  1. 1.↵
    1. Wolochow H,
    2. Hildebrand GJ,
    3. Lamanna C
    . 1966. Translocation of microorganisms across the intestinal wall of the rat: effect of microbial size and concentration. J Infect Dis116:523–528. doi:10.1093/infdis/116.4.523.
    OpenUrlCrossRefPubMedWeb of Science
  2. 2.↵
    1. Berg RD,
    2. Garlington AW
    . 1979. Translocation of certain indigenous bacteria from the gastrointestinal tract to the mesenteric lymph nodes and other organs in a gnotobiotic mouse model. Infect Immun23:403–411.
    OpenUrlAbstract/FREE Full Text
  3. 3.↵
    1. Steffen EK,
    2. Berg RD,
    3. Deitch EA
    . 1988. Comparison of translocation rates of various indigenous bacteria from the gastrointestinal tract to the mesenteric lymph node. J Infect Dis157:1032–1038. doi:10.1093/infdis/157.5.1032.
    OpenUrlCrossRefPubMedWeb of Science
  4. 4.↵
    1. Berg RD
    . 1995. Bacterial translocation from the gastrointestinal tract. Trends Microbiol3:149–154. doi:10.1016/S0966-842X(00)88906-4.
    OpenUrlCrossRefPubMed
  5. 5.↵
    1. Berg RD
    . 1992. The scientific basis of the probiotic concept, vol 3. Chapman and Hall, London, United Kingdom.
  6. 6.↵
    1. Berg RD
    . 1981. Promotion of the translocation of enteric bacteria from the gastrointestinal tracts of mice by oral treatment with penicillin, clindamycin, or metronidazole. Infect Immun33:854–861.
    OpenUrlAbstract/FREE Full Text
  7. 7.↵
    1. Owens WE,
    2. Berg RD
    . 1980. Bacterial translocation from the gastrointestinal tract of athymic (nu/nu) mice. Infect Immun27:461–467.
    OpenUrlAbstract/FREE Full Text
  8. 8.↵
    1. Gautreaux MD,
    2. Deitch EA,
    3. Berg RD
    . 1994. T lymphocytes in host defense against bacterial translocation from the gastrointestinal tract. Infect Immun62:2874–2884.
    OpenUrlAbstract/FREE Full Text
  9. 9.↵
    1. Parks DA,
    2. Bulkley GB,
    3. Granger DN,
    4. Hamilton SR,
    5. McCord JM
    . 1982. Ischemic injury in the cat small intestine: role of superoxide radicals. Gastroenterology82:9–15.
    OpenUrlPubMedWeb of Science
  10. 10.↵
    1. Berg R
    . 1983. Bacterial translocation from the gastrointestinal tracts of mice receiving immunosuppressive chemotherapeutic agents. Curr Microbiol8:285–292. doi:10.1007/BF01577729.
    OpenUrlCrossRefWeb of Science
  11. 11.↵
    1. Morehouse JL,
    2. Specian RD,
    3. Stewart JJ,
    4. Berg RD
    . 1986. Translocation of indigenous bacteria from the gastrointestinal tract of mice after oral ricinoleic acid treatment. Gastroenterology91:673–682. doi:10.1016/0016-5085(86)90638-4.
    OpenUrlCrossRefPubMed
  12. 12.↵
    1. Deitch EA,
    2. Bridges W,
    3. Baker J,
    4. Ma JW,
    5. Ma L,
    6. Grisham MB,
    7. Granger DN,
    8. Specian RD,
    9. Berg R
    . 1988. Hemorrhagic shock-induced bacterial translocation is reduced by xanthine oxidase inhibition or inactivation. Surgery104:191–198.
    OpenUrlPubMedWeb of Science
  13. 13.↵
    1. MacFie J
    . 2004. Current status of bacterial translocation as a cause of surgical sepsis. Br Med Bull71:1–11. doi:10.1093/bmb/ldh029.
    OpenUrlCrossRefPubMedWeb of Science
  14. 14.↵
    1. MacFie J,
    2. O'Boyle CJ,
    3. Mitchell CJ,
    4. Buckley P,
    5. Johnstone D,
    6. Sudworth D
    . 1999. Gut origin of sepsis: a prospective study investigating associations between bacterial translocation, gastric microflora, and septic morbidity. Gut45:223–228. doi:10.1136/gut.45.2.223.
    OpenUrlAbstract/FREE Full Text
  15. 15.↵
    1. O'Boyle CJ,
    2. MacFie J,
    3. Mitchell CJ,
    4. Johnstone D,
    5. Sagar P,
    6. Sedman P
    . 1998. Microbiology of bacterial translocation in humans. Gut42:29–35. doi:10.1136/gut.42.1.29.
    OpenUrlAbstract/FREE Full Text
  16. 16.↵
    1. Huang E
    . 2000. Internal medicine: handbook for clinicians, resident survival guide. Scrub Hill Press, Arlington, VA.
  17. 17.↵
    1. Wisplinghoff H,
    2. Bischoff T,
    3. Tallent SM,
    4. Seifert H,
    5. Wenzel RP,
    6. Edmond MB
    . 2004. Nosocomial bloodstream infections in US hospitals: analysis of 24,179 cases from a prospective nationwide surveillance study. Clin Infect Dis39:309–317. doi:10.1086/421946.
    OpenUrlCrossRefPubMedWeb of Science
  18. 18.↵
    1. Samet A,
    2. Sledzinska A,
    3. Krawczyk B,
    4. Hellmann A,
    5. Nowicki S,
    6. Kur J,
    7. Nowicki B
    . 2013. Leukemia and risk of recurrent Escherichia coli bacteremia: genotyping implicates E. coli translocation from the colon to the bloodstream. Eur J Clin Microbiol Infect Dis32:1393–1400. doi:10.1007/s10096-013-1886-9.
    OpenUrlCrossRefPubMed
  19. 19.↵
    1. Russo TA,
    2. Johnson JR
    . 2000. A proposal for an inclusive designation for extraintestinal pathogenic Escherichia coli: ExPEC. J Infect Dis181:1753–1754. doi:10.1086/315418.
    OpenUrlCrossRefPubMedWeb of Science
  20. 20.↵
    1. Kaper JB,
    2. Natoro JP,
    3. Mobley HL
    . 2004. Pathogenic Escherichia coli. Nat Rev Microbiol2:123–140. doi:10.1038/nrmicro818.
    OpenUrlCrossRefPubMedWeb of Science
  21. 21.↵
    1. Johnson JR,
    2. Russo TA
    . 2005. Molecular epidemiology of extraintestinal pathogenic (uropathogenic) Escherichia coli. Int J Med Microbiol295:383–404. doi:10.1016/j.ijmm.2005.07.005.
    OpenUrlCrossRefPubMedWeb of Science
  22. 22.↵
    1. Lautenbach E,
    2. Patel JB,
    3. Bilker WB,
    4. Edelstein PH,
    5. Fishman NO
    . 2001. Extended-spectrum beta-lactamase-producing Escherichia coli and Klebsiella pneumoniae: risk factors for infection and impact of resistance on outcomes. Clinl Infect Dis32:1162–1171. doi:10.1086/319757.
    OpenUrlCrossRefPubMedWeb of Science
  23. 23.↵
    1. Tumbarello M,
    2. Sanguinetti M,
    3. Montuori E,
    4. Trecarichi EM,
    5. Posteraro B,
    6. Fiori B,
    7. Citton R,
    8. D'Inzeo T,
    9. Fadda G,
    10. Cauda R,
    11. Spanu T
    . 2007. Predictors of mortality in patients with bloodstream infections caused by extended-spectrum-beta-lactamase-producing Enterobacteriaceae: importance of inadequate initial antimicrobial treatment. Antimicrob Agents Chemother51:1987–1994. doi:10.1128/AAC.01509-06.
    OpenUrlAbstract/FREE Full Text
  24. 24.↵
    1. Pitout JD,
    2. Laupland KB
    . 2008. Extended-spectrum beta-lactamase-producing Enterobacteriaceae: an emerging public-health concern. Lancet Infect Dis8:159–166. doi:10.1016/S1473-3099(08)70041-0.
    OpenUrlCrossRefPubMedWeb of Science
  25. 25.↵
    1. Foxman B
    . 2010. The epidemiology of urinary tract infection. Nat Rev Urol7:653–660. doi:10.1038/nrurol.2010.190.
    OpenUrlCrossRefPubMed
  26. 26.↵
    1. Banerjee R,
    2. Johnson JR
    . 2013. A new clone sweeps clean: the enigmatic emergence of Escherichia coli sequence type 131. Antimicrob Agents Chemother58:4997–5004. doi:10.1128/AAC.02824-14.
    OpenUrlCrossRef
  27. 27.↵
    1. Köhler CD,
    2. Dobrindt U
    . 2011. What defines extraintestinal pathogenic Escherichia coli?Int J Med Microbiol301:642–627. doi:10.1016/j.ijmm.2011.09.006.
    OpenUrlCrossRefPubMed
  28. 28.↵
    1. Mokady D,
    2. Gophna U,
    3. Ron EZ
    . 2005. Virulence factors of septicemic Escherichia coli strains. Int J Med Microbiol295:455–462. doi:10.1016/j.ijmm.2005.07.007.
    OpenUrlCrossRefPubMed
  29. 29.↵
    1. Pitout JD
    . 2012. Extraintestinal pathogenic Escherichia coli: a combination of virulence with antibiotic resistance. Front Microbiol3:1–7. doi:10.3389/fmicb.2012.00009.
    OpenUrlCrossRefPubMed
  30. 30.↵
    1. Deitch EA
    . 2002. Bacterial translocation or lymphatic drainage of toxic products from the gut: what is important in human beings?Surgery13:241–244.
    OpenUrl
  31. 31.↵
    1. Gautreaux MD,
    2. Deitch EA,
    3. Berg RD
    . 1994. Bacterial translocation from the gastrointestinal tract to various segments of the mesenteric lymph mode complex. Infect Immun62:2132–2134.
    OpenUrlAbstract/FREE Full Text
  32. 32.↵
    1. Kim KP,
    2. Jagadeesan B,
    3. Burkholder KM,
    4. Jaradat ZW,
    5. Wampler JL,
    6. Lathrop AA,
    7. Morgan MT,
    8. Bhunia AK
    . 2006. Adhesion characteristics of Listeria adhesion protein (LAP)-expressing Escherichia coli to Caco-2 cells and of recombinant LAP to eukaryotic receptor Hsp60 as examined in a surface plasmon resonance sensor. FEMS Microbiol Lett256:324–332. doi:10.1111/j.1574-6968.2006.00140.x.
    OpenUrlCrossRefPubMed
  33. 33.↵
    1. Pizarro-Cerdá J,
    2. Lecuit M,
    3. Cossart P
    . 2002. Measuring and analysing invasion of mammalian cells by bacterial pathogens: the Listeria monocytogenes system. Methods Microbiol31:161–177. doi:10.1016/S0580-9517(02)31009-2.
    OpenUrlCrossRef
  34. 34.↵
    1. Kortman GM,
    2. Bolei A,
    3. Swinkels DW,
    4. Tjalsma H
    . 2012. Iron availability increases the pathogenic potential of Salmonella typhimurium and other enteric pathogens at the intestinal epithelial interface. PLoS One7:e29968. doi:10.1371/journal.pone.0029968.
    OpenUrlCrossRefPubMed
  35. 35.↵
    1. Russo TA,
    2. McFadden CD,
    3. Carlino-MacDonald UB,
    4. Beanan JM,
    5. Barnard TJ,
    6. Johnson JR
    . 2002. IroN functions as a siderophore receptor and is a urovirulence factor in an extraintestinal pathogenic isolate of Escherichia coli. Infect Immun70:7156–7160. doi:10.1128/IAI.70.12.7156-7160.2002.
    OpenUrlAbstract/FREE Full Text
  36. 36.↵
    1. Feldmann F,
    2. Sorsa LJ,
    3. Hildinger K,
    4. Schubert S
    . 2007. The salmochelin siderophore receptor IroN contributes to invasion of urothelial cells by extraintestinal pathogenic Escherichia coli in vitro. Infect Immun75:3183–3187. doi:10.1128/IAI.00656-06.
    OpenUrlAbstract/FREE Full Text
  37. 37.↵
    1. Rippere-Lampe KE,
    2. O'Brien AD,
    3. Conran R,
    4. Lockman HA
    . 2001. Mutation of the gene encoding cytotoxic necrotizing factor type 1 (cnf(1)) attenuates the virulence of uropathogenic Escherichia coli. Infect Immun69:3954–3964. doi:10.1128/IAI.69.6.3954-3964.2001.
    OpenUrlAbstract/FREE Full Text
  38. 38.↵
    1. Martinez JJ,
    2. Mulvey MA,
    3. Schilling JD,
    4. Pinker JS,
    5. Hultgren SJ
    . 2000. Type-1 pilus-mediated invasion of bladder epithelial cells. EMBO J19:2803–2812. doi:10.1093/emboj/19.12.2803.
    OpenUrlAbstract/FREE Full Text
  39. 39.↵
    1. Mulvey MA,
    2. Schilling JD,
    3. Hultgren SJ
    . 2001. Establishment of a persistent Escherichia coli reservoir during the acute phase of a bladder infection. Infect Immun69:4572–4579. doi:10.1128/IAI.69.7.4572-4579.2001.
    OpenUrlAbstract/FREE Full Text
  40. 40.↵
    1. Anderson GG,
    2. Palermo JJ,
    3. Schilling JD,
    4. Roth R,
    5. Heuser J,
    6. Hultgren SJ
    . 2003. Intracellular bacterial biofilm-like pods in urinary tract infections. Science301:105–107. doi:10.1126/science.1084550.
    OpenUrlAbstract/FREE Full Text
  41. 41.↵
    1. Wright KJ,
    2. Seed P,
    3. Hultgren SJ
    . 2007. Development of intracellular bacterial communities of uropathogenic Escherichia coli depends on type 1 pili. Cell Microbiol9:2230–2241. doi:10.1111/j.1462-5822.2007.00952.x.
    OpenUrlCrossRefPubMedWeb of Science
  42. 42.↵
    1. Berry RE,
    2. Klumpp DJ,
    3. Schaeffer AJ
    . 2009. Urothelial cultures support intracellular bacterial community formation by uropathogenic Escherichia coli. Infect Immun77:2762–2772. doi:10.1128/IAI.00323-09.
    OpenUrlAbstract/FREE Full Text
  43. 43.↵
    1. Sato T,
    2. Stange DE,
    3. Ferrante M,
    4. Vries RG,
    5. Van Es JH,
    6. Van den Brink S,
    7. Van Houdt WJ,
    8. Pronk A,
    9. Van Gorp J,
    10. Siersema PD,
    11. Clevers H
    . 2011. Long-term expansion of epithelial organoids from human colon, adenoma, adenocarcinoma, and Barrett's epithelium. Gastroenterology141:1762–1772. doi:10.1053/j.gastro.2011.07.050.
    OpenUrlCrossRefPubMedWeb of Science
  44. 44.↵
    1. Saxena K,
    2. Blutt SE,
    3. Ettayebi K,
    4. Zeng XL,
    5. Broughman JR,
    6. Crawford SE,
    7. Karandikar UC,
    8. Sastri NP,
    9. Conner ME,
    10. Opekun AR,
    11. Graham DY,
    12. Qureshi W,
    13. Sherman V,
    14. Foulke-Abel J,
    15. In J,
    16. Kovbasnjuk O,
    17. Zachos NC,
    18. Donowitz M,
    19. Estes MK
    . 2015. Human intestinal enteroids: a new model to study human rotavirus infection, host restriction, and pathophysiology. J Virol90:43–56. doi:10.1128/JVI.01930-15.
    OpenUrlAbstract/FREE Full Text
  45. 45.↵
    1. Foulke-Abel J,
    2. In J,
    3. Kovbasnjuk O,
    4. Zachos NC,
    5. Ettayebi K,
    6. Blutt SE,
    7. Hyser JM,
    8. Zeng XL,
    9. Crawford SE,
    10. Broughman JR,
    11. Estes MK,
    12. Donowitz M
    . 2014. Human enteroids as an ex-vivo model of host–pathogen interactions in the gastrointestinal tract. Exp Biol Med239:124–1134. doi:10.1177/1535370214529398.
    OpenUrlCrossRefPubMed
  46. 46.↵
    1. In J,
    2. Foulke-Abel J,
    3. Zachos NC,
    4. Hansen AM,
    5. Kaper JB,
    6. Bernstein HD,
    7. Halushka M,
    8. Blutt S,
    9. Estes MK,
    10. Donowitz M,
    11. Kovbasnjuk O
    . 2016. Enterohemorrhagic Escherichia coli reduce mucus and intermicovillar bridges in human stem cell-derived colonoids. Cell Mol Gastroenterol Hepatol2:48–62. doi:10.1016/j.jcmgh.2015.10.001.
    OpenUrlCrossRefPubMed
  47. 47.↵
    1. Burkholder KM,
    2. Bhunia AK
    . 2010. Listeria monocytogenes uses Listeria adhesion protein (LAP) to promote bacterial transepithelial translocation and induces expression of LAP receptor Hsp60. Infect Immun78:5062–5063. doi:10.1128/IAI.00516-10.
    OpenUrlAbstract/FREE Full Text
  48. 48.↵
    1. Cruz N,
    2. Qi L,
    3. Alvarez X,
    4. Berg RD,
    5. Deitch EA
    . 1994. The Caco-2 cell monolayer system as an in vitro model for studying bacterial-enterocyte interactions and bacterial translocation. J Burn Care Rehabil15:207–212. doi:10.1097/00004630-199405000-00002.
    OpenUrlCrossRefPubMed
  49. 49.↵
    1. Green SI,
    2. Ajami NJ,
    3. Ma L,
    4. Poole NM,
    5. Price RE,
    6. Petrosino JF,
    7. Maresso AW
    . 2015. Murine model of chemotherapy-induced extraintestinal pathogenic Escherichia coli translocation. Infect Immun83:3243–3256. doi:10.1128/IAI.00684-15.
    OpenUrlAbstract/FREE Full Text
  50. 50.↵
    1. Cerf-Bensussan N,
    2. Gaboriau-Routhiau V
    . 2010. The immune system and the gut microbiota: friends or foes?Nat Rev Immunol10:735–744. doi:10.1038/nri2850.
    OpenUrlCrossRefPubMedWeb of Science
  51. 51.↵
    1. Gudiol C,
    2. Bodro M,
    3. Simonetti A,
    4. Tubau F,
    5. González-Barca E,
    6. Cisnal M,
    7. Domingo-Domenech E,
    8. Jiménez L,
    9. Carratalá J
    . 2013. Changing aetiology, clinical features, antimicrobial resistance, and outcomes of bloodstream infection in neutropenic cancer patients. Clin Microbiol Infect19:474–479. doi:10.1111/j.1469-0691.2012.03879.x.
    OpenUrlCrossRefPubMed
  52. 52.↵
    1. Velasco E,
    2. Byington R,
    3. Martins CAS,
    4. Schirmer M,
    5. Dias LMC,
    6. Gonçalves VMSC
    . 2006. Comparative study of clinical characteristics of neutropenic and non-neutropenic adult cancer patients with bloodstream infections. Eur J Clin Microbiol Infect Dis25:1–7. doi:10.1007/s10096-005-0077-8.
    OpenUrlCrossRefPubMed
  53. 53.↵
    1. Madigan T,
    2. Johnson JR,
    3. Clabots C,
    4. Johnston BD,
    5. Porter SB,
    6. Slater BS,
    7. Banerjee R
    . 2015. Extensive household outbreak of urinary tract infection and intestinal colonization due to extended-spectrum β-lactamase-producing Escherichia coli sequence type 131. Clin Infect Dis61:e5–e12. doi:10.1093/cid/civ273.
    OpenUrlCrossRefPubMed
  54. 54.↵
    1. Spellberg B,
    2. Doi Y
    . 2015. The rise of fluoroquinolone-resistant Escherichia coli in the community: scarier than we thought. J Infect Dis212:1853–1855. doi:10.1093/infdis/jiv279.
    OpenUrlCrossRefPubMed
  55. 55.↵
    1. Schwan WR
    . 2011. Regulation of fim genes in uropathogenic Escherichia coli. World J Clin Infect Dis1:17–25. doi:10.5495/wjcid.v1.i1.17.
    OpenUrlCrossRefPubMed
  56. 56.↵
    1. Ofek I,
    2. Mosek A,
    3. Sharon N
    . 1981. Mannose-specific adherence of Escherichia coli freshly excreted in the urine of patients with urinary tract infections, and of isolates subcultured from the infected urine. Infect Immun34:708–711.
    OpenUrlAbstract/FREE Full Text
  57. 57.↵
    1. Keith BR,
    2. Maurer L,
    3. Spears PA,
    4. Orndorff PE
    . 1986. Receptor binding function of type 1 pili effects bladder colonization by a clinical isolate of Escherichia coli. Infect Immun53:693–696.
    OpenUrlAbstract/FREE Full Text
  58. 58.↵
    1. Pere A,
    2. Nowicki B,
    3. Saxén H,
    4. Siitonen A,
    5. Korhonen TK
    . 1987. Expression of P, type-1, and type-1C fimbriae of Escherichia coli in the urine of patients with acute urinary tract infection. J Infect Dis156:567–574. doi:10.1093/infdis/156.4.567.
    OpenUrlCrossRefPubMedWeb of Science
  59. 59.↵
    1. Mobley HL,
    2. Chippendale GR,
    3. Tenney JH,
    4. Hull RA,
    5. Warren JW
    . 1987. Expression of type 1 fimbriae may be required for persistence of Escherichia coli in the catheterized urinary tract. J Clin Microbiol25:2253–2257.
    OpenUrlAbstract/FREE Full Text
  60. 60.↵
    1. Kisielius PV,
    2. Schwan WR,
    3. Amundsen SK,
    4. Duncan JL,
    5. Schaeffer AJ
    . 1989. In vivo expression and variation of Escherichia coli type 1 and P pili in the urine of adults with acute urinary tract infections. Infect Immun57:1656–1662.
    OpenUrlAbstract/FREE Full Text
  61. 61.↵
    1. Lim JK,
    2. Gunther NW,
    3. Zhao H,
    4. Johnson DE,
    5. Keay SK,
    6. Mobley HL
    . 1998. In vivo phase variation of Escherichia coli type 1fimbrial genes in women with urinary tract infection. Infect Immun66:3303–3310.
    OpenUrlAbstract/FREE Full Text
  62. 62.↵
    1. Snyder JA,
    2. Lloyd AL,
    3. Lockatell CV,
    4. Johnson DE,
    5. Mobley HL
    . 2006. Role of phase variation of type 1 fimbriae in a uropathogenic Escherichia coli cystitis isolate during urinary tract infection. Infect Immun74:1387–1393. doi:10.1128/IAI.74.2.1387-1393.2006.
    OpenUrlAbstract/FREE Full Text
  63. 63.↵
    1. Hung CS,
    2. Bouckaert J,
    3. Hung D,
    4. Pinkner J,
    5. Widberg C,
    6. DeFusco A,
    7. Auguste CG,
    8. Strouse R,
    9. Langermann S,
    10. Waksman G,
    11. Hultgren SJ
    . 2002. Structural basis of tropism of Escherichia coli to the bladder during urinary tract infection. Mol Microbiol44:903–915. doi:10.1046/j.1365-2958.2002.02915.x.
    OpenUrlCrossRefPubMedWeb of Science
  64. 64.↵
    1. Chen SL,
    2. Hung CS,
    3. Pinkner JS,
    4. Walker JN,
    5. Cusumano CK,
    6. Li Z,
    7. Bouckaert J,
    8. Gordon JI,
    9. Hultgren SJ
    . 2009. Positive selection identifies an in vivo role for FimH during urinary tract infection in addition to mannose binding. Proc Natl Acad Sci U S A106:22439–22444. doi:10.1073/pnas.0902179106.
    OpenUrlAbstract/FREE Full Text
  65. 65.↵
    1. Meiland RS,
    2. Geerlings E,
    3. Langermann S,
    4. Brouwer EC,
    5. Coenjaerts FE,
    6. Hoepelman AI
    . 2004. Fimch antiserum inhibits the adherence of Escherichia coli to cells collected by voided urine specimens of diabetic women. J Urol171:1589–1593. doi:10.1097/01.ju.0000118402.01034.fb.
    OpenUrlCrossRefPubMed
  66. 66.↵
    1. Connell I,
    2. Agace W,
    3. Klemm P,
    4. Schembri M,
    5. Marild S,
    6. Svanborg C
    . 1996. Type 1 fimbrial expression enhances Escherichia coli virulence for the urinary tract. Proc Natl Acad Sci U S A93:9827–9832. doi:10.1073/pnas.93.18.9827.
    OpenUrlAbstract/FREE Full Text
  67. 67.↵
    1. Bouckaert J,
    2. Berglund J,
    3. Schembri M,
    4. De Genst E,
    5. Cools L,
    6. Wuhrer M,
    7. Hung CS,
    8. Pinkner J,
    9. Slättegård R,
    10. Zavialov A,
    11. Choudhury D,
    12. Langermann S,
    13. Hultgren SJ,
    14. Wyns L,
    15. Klemm P,
    16. Oscarson S,
    17. Knight SD,
    18. De Greve H
    . 2005. Receptor binding studies disclose a novel class of high-affinity inhibitors of the Escherichia coli FimH adhesin. Mol Microbiol55:441–455. doi:10.1111/j.1365-2958.2004.04415.x.
    OpenUrlCrossRefPubMed
  68. 68.↵
    1. Scharenberg M,
    2. Schwardt O,
    3. Rabbani S,
    4. Ernst B
    . 2012. Target selectivity of FimH antagonists. J Med Chem55:9810–9816. doi:10.1021/jm3010338.
    OpenUrlCrossRefPubMed
  69. 69.↵
    1. Jarvis C,
    2. Han Z,
    3. Kalas V,
    4. Klein R,
    5. Pinkner JS,
    6. Ford B,
    7. Binkley J,
    8. Cusumano CK,
    9. Cusumano Z,
    10. Mydock-McGrane L,
    11. Hultgren SJ,
    12. Janetka JW
    . 2016. Antivirulence isoquinolone mannosides: optimization of the biaryl aglycone for FimH lectin binding affinity and efficacy in the treatment of chronic UTI. Chem Med Chem11:367–373. doi:10.1002/cmdc.201600006.
    OpenUrlCrossRef
  70. 70.↵
    1. Lo AW,
    2. Van de Water K,
    3. Gane PJ,
    4. Chan AW,
    5. Steadman D,
    6. Stevens K,
    7. Selwood DL,
    8. Waksman G,
    9. Remaut H
    . 2014. Suppression of type 1 pilus assembly in uropathogenic Escherichia coli by chemical inhibition of subunit polymerization. J Antimicrob Chemother69:1017–1026. doi:10.1093/jac/dkt467.
    OpenUrlCrossRefPubMedWeb of Science
  71. 71.↵
    1. Totsika M,
    2. Kostakioti M,
    3. Hannan TJ,
    4. Upton M,
    5. Beatson SA,
    6. Janetka JW,
    7. Hultgren SJ,
    8. Schembri MA
    . 2013. A FimH inhibitor prevents acute bladder infection and treats chronic cystitis caused by multidrug-resistant uropathogenic Escherichia coli ST131. J Infect Dis208:921–928. doi:10.1093/infdis/jit245.
    OpenUrlCrossRefPubMed
  72. 72.↵
    1. Sokurenko EV,
    2. Chesnokova V,
    3. Dykhuizen DE,
    4. Ofek I,
    5. Wu XR,
    6. Krogfelt KA,
    7. Struve C,
    8. Schembri MA,
    9. Hasty DL
    . 1998. Pathogenic adaptation of Escherichia coli by natural variation of the FimH adhesin. Proc Natl Acad Sci U S A95:8922–8926. doi:10.1073/pnas.95.15.8922.
    OpenUrlAbstract/FREE Full Text
  73. 73.↵
    1. Weissman SJ,
    2. Beskhlebnaya V,
    3. Chesnokova V,
    4. Chattopadhyay S,
    5. Stamm WE,
    6. Hooton TM,
    7. Sokurenko EV
    . 2007. Differential stability and trade-off effects of pathoadaptive mutations in the Escherichia coli FimH adhesin. Infect Immun75:3548–3555. doi:10.1128/IAI.01963-06.
    OpenUrlAbstract/FREE Full Text
  74. 74.↵
    1. Weissman SJ,
    2. Chattopadhyay S,
    3. Aprikian P,
    4. Obata-Yasuoka M,
    5. Yarova-Yarovaya Y,
    6. Stapleton A,
    7. Ba-Thein W,
    8. Dykhuizen D,
    9. Johnson JR,
    10. Sokurenko EV
    . 2006. Clonal analysis reveals high rate of structural mutations in fimbrial adhesins of extraintestinal pathogenic Escherichia coli. Mol Microbiol59:975–988. doi:10.1111/j.1365-2958.2005.04985.x.
    OpenUrlCrossRefPubMedWeb of Science
  75. 75.↵
    1. Abdallah KS,
    2. Cao Y,
    3. Wei DJ
    . 2011. Epidemiologic investigation of extra-intestinal pathogenic E. coli (ExPEC) based on PCR phylogenetic group and fimH single nucleotide polymorphisms (SNPs) in China. Int J Mol Epidemiol Genet2:339–353.
    OpenUrlPubMed
  76. 76.↵
    1. Weissman SJ,
    2. Johnson JR,
    3. Tchesnokova V,
    4. Billig M,
    5. Dykhuizen D,
    6. Riddell K,
    7. Rogers P,
    8. Qin X,
    9. Butler-Wu S,
    10. Cookson BT,
    11. Fang FC,
    12. Scholes D,
    13. Chattopadhyay S,
    14. Sokurenko E
    . 2012. High-resolution two-locus clonal typing of extraintestinal pathogenic Escherichia coli. Appl Environ Microbiol78:1353–1360. doi:10.1128/AEM.06663-11.
    OpenUrlAbstract/FREE Full Text
  77. 77.↵
    1. Sokurenko EV,
    2. Chesnokova V,
    3. Doyle RJ,
    4. Hasty DL
    . 1997. Differential binding to mannosides and uroepithelial cells. J Biol Chem272:17880–17886. doi:10.1074/jbc.272.28.17880.
    OpenUrlAbstract/FREE Full Text
  78. 78.↵
    1. Schwartz DJ,
    2. Kalas V,
    3. Pinkner JS,
    4. Chen SL,
    5. Spaulding CN,
    6. Dodson KW,
    7. Hultgren SJ
    . 2013. Positively selected FimH residues enhance virulence during urinary tract infection by altering FimH conformation. Proc Natl Acad Sci U S A110:15530–15537. doi:10.1073/pnas.1315203110.
    OpenUrlAbstract/FREE Full Text
  79. 79.↵
    1. Spaulding CN,
    2. Klein RD,
    3. Ruer S,
    4. Kau AL,
    5. Schreiber IV HL,
    6. Cusumano ZT,
    7. Dodson KW,
    8. Pinkner JS,
    9. Fremont DH,
    10. Janetka JW,
    11. Remaut H,
    12. Gordon JI,
    13. Hultgren SJ
    . 2017. Selective depletion of uropathogenic E. coli from the gut by a FimH antagonist. Nature546:528–532. doi:10.1038/nature22972.
    OpenUrlCrossRef
  80. 80.↵
    1. Bellm LA,
    2. Epstein JB,
    3. Rose-Ped A,
    4. Martin P,
    5. Fuchs HJ
    . 2000. Patient reports of complications of bone marrow transplantation. Support Care Cancer8:33–39.
    OpenUrlPubMedWeb of Science
  81. 81.↵
    1. Troeger H,
    2. Richter JF,
    3. Beutin L,
    4. Günzel D,
    5. Dobrindt U,
    6. Epple HJ,
    7. Gitter AH,
    8. Zeitz M,
    9. Fromm M,
    10. Schulzke JD
    . 2007. Escherichia coli a-haemolysin induces focal leaks in colonic epithelium: a novel mechanism of bacterial translocation. Cell Microbiol9:2530–2540. doi:10.1111/j.1462-5822.2007.00978.x.
    OpenUrlCrossRefPubMed
  82. 82.↵
    1. Rescigno M,
    2. Urbano M,
    3. Valzasina B,
    4. Francolini M,
    5. Rotta G,
    6. Bonasio R,
    7. Granucci F,
    8. Kraehenbuhl JP,
    9. Ricciardi-Castagnoli P
    . 2001. Dendritic cells express tight junction proteins and penetrate gut epithelial monolayers to sample bacteria. Nat Immunol2:361–367. doi:10.1038/86373.
    OpenUrlCrossRefPubMedWeb of Science
  83. 83.↵
    1. Hase K,
    2. Kawano K,
    3. Nochi T,
    4. Pontes GS,
    5. Fukuda S,
    6. Ebisawa M,
    7. Kadokura K,
    8. Tobe T,
    9. Fujimura Y,
    10. Kawano S,
    11. Yabashi A,
    12. Waguri S,
    13. Nakato G,
    14. Kimura S,
    15. Murakami T,
    16. Iimura M,
    17. Hamura K,
    18. Fukuoka S,
    19. Lowe AW,
    20. Itoh K,
    21. Kiyono H,
    22. Ohno H
    . 2009. Uptake through glycoprotein 2 of FimH(+) bacteria by M cells initiates mucosal immune response. Nature462:226–230. doi:10.1038/nature08529.
    OpenUrlCrossRefPubMedWeb of Science
  84. 84.↵
    1. Phalipon A,
    2. Sansonetti PJ
    . 2003. Shigellosis: innate mechanisms of inflammatory destruction of the intestinal epithelium, adaptive immune response, and vaccine development. Crit Rev Immunol23:371–401. doi:10.1615/CritRevImmunol.v23.i56.20.
    OpenUrlCrossRefPubMedWeb of Science
  85. 85.↵
    1. Backert S,
    2. Boehm M,
    3. Wessler S,
    4. Tegtmeyer N
    . 2013. Transmigration route of Campylobacter jejuni across polarized intestinal epithelial cells: paracellular, transcellular or both?J Cell Commun Signaling11:72. doi:10.1186/1478-811X-11-72.
    OpenUrlCrossRefPubMed
  86. 86.↵
    1. Wu XR,
    2. Sun TT,
    3. Medina JJ
    . 1996. In vitro binding of type 1-fimbriated Escherichia coli to uroplakins Ia and Ib: relation to urinary tract infections. Proc Natl Acad Sci U S A93:9630–9635. doi:10.1073/pnas.93.18.9630.
    OpenUrlAbstract/FREE Full Text
  87. 87.↵
    1. Xie B,
    2. Zhou G,
    3. Chan SY,
    4. Shapiro E,
    5. Kong XP,
    6. Wu XR,
    7. Sun TT,
    8. Costello CE
    . 2006. Distinct glycan structures of uroplakins Ia and Ib: structural basis for the selective binding of FimH adhesin to uroplakin Ia. J Biol Chem281:14644–14653. doi:10.1074/jbc.M600877200.
    OpenUrlAbstract/FREE Full Text
  88. 88.↵
    1. Zhou G,
    2. Mo WJ,
    3. Sebbel P,
    4. Min G,
    5. Neubert TA,
    6. Glockshuber R,
    7. Wu XR,
    8. Sun TT,
    9. Kong XP
    . 2001. Uroplakin Ia is the urothelial receptor for uropathogenic Escherichia coli: evidence from in vitro FimH binding. J Cell Sci114:4095–4103.
    OpenUrlAbstract/FREE Full Text
  89. 89.↵
    1. Duncan MJ,
    2. Li G,
    3. Shin JS,
    4. Carson JL,
    5. Abraham SN
    . 2004. Bacterial penetration of bladder epithelium through lipid rafts. J Biol Chem279:18944–18951. doi:10.1074/jbc.M400769200.
    OpenUrlAbstract/FREE Full Text
  90. 90.↵
    1. Song J,
    2. Bishop BL,
    3. Li G,
    4. Duncan MJ,
    5. Abraham SN
    . 2007. TLR4-initiated and cAMP-mediated abrogation of bacterial invasion of the bladder. Cell Host Microbe1:287–298. doi:10.1016/j.chom.2007.05.007.
    OpenUrlCrossRefPubMedWeb of Science
  91. 91.↵
    1. Eto DS,
    2. Jones TA,
    3. Sundsbak JL,
    4. Mulvey MA
    . 2007. Integrin-mediated host cell invasion by type 1-piliated uropathogenic Escherichia coli. PLoS Pathog3:e100. doi:10.1371/journal.ppat.0030100.
    OpenUrlCrossRefPubMed
  92. 92.↵
    1. Martinez JJ,
    2. Hultgren SJ
    . 2002. Requirement of Rho-family GTPases in the invasion of type 1-piliated uropathogenic Escherichia coli. Cell Microbiol4:19–28. doi:10.1046/j.1462-5822.2002.00166.x.
    OpenUrlCrossRefPubMedWeb of Science
  93. 93.↵
    1. Dhakal BK,
    2. Mulvey MA
    . 2009. Uropathogenic Escherichia coli invades host cells via an HDAC6-modulated microtubule-dependent pathway. J Biol Chem284:446–454. doi:10.1074/jbc.M805010200.
    OpenUrlAbstract/FREE Full Text
  94. 94.↵
    1. Bishop BL,
    2. Duncan MJ,
    3. Song J,
    4. Li G,
    5. Zaas D,
    6. Abraham SN
    . 2007. Cyclic AMP-regulated exocytosis of Escherichia coli from infected bladder epithelial cells. Nat Med13:625–630. doi:10.1038/nm1572.
    OpenUrlCrossRefPubMedWeb of Science
  95. 95.↵
    1. Johnson JR,
    2. Russo TA,
    3. Scheutz F,
    4. Brown JJ,
    5. Zhang L,
    6. Palin K,
    7. Rode C,
    8. Bloch C,
    9. Marrs CF,
    10. Foxman B
    . 1997. Discovery of disseminated J96-like strains of uropathogenic Escherichia coli O4:H5 containing genes for both PapG (J96) (class I) and PrsG (J96) (class III) Gal (alpha1-4) Gal-binding adhesins. J Infect Dis175:983–988. doi:10.1086/514006.
    OpenUrlCrossRefPubMedWeb of Science
  96. 96.↵
    1. Russo TA,
    2. Guenther JE,
    3. Wenderoth S,
    4. Frank MM
    . 1993. Generation of isogenic K54 capsule638 deficient Escherichia coli strains through TnphoA-mediated gene disruption. Mol Microbiol9:357–364. doi:10.1111/j.1365-2958.1993.tb01696.x.
    OpenUrlCrossRefPubMedWeb of Science
  97. 97.↵
    1. Russo TA,
    2. Singh G
    . 1993. An extraintestinal, pathogenic isolate of Escherichia coli (O4/K54/H5) can produce a group 1 capsule which is divergently regulated from its constitutively produced group 2, K54 capsular polysaccharide. J Bacteriol175:7617–7623. doi:10.1128/jb.175.23.7617-7623.1993.
    OpenUrlAbstract/FREE Full Text
  98. 98.↵
    1. Johnson JR,
    2. Porter SB,
    3. Zhanel G,
    4. Kuskowski MA,
    5. Denamur E
    . 2012. Virulence of Escherichia coli clinical isolates in a murine sepsis model in relation to sequence type ST131 status, fluoroquinolone resistance, and virulence genotype. Infect Immun80:1554–1562. doi:10.1128/IAI.06388-11.
    OpenUrlAbstract/FREE Full Text
  99. 99.↵
    1. Russo TA,
    2. Wang Z,
    3. Davidson BA,
    4. Genagon SA,
    5. Beanan JM,
    6. Olson R,
    7. Holm BA,
    8. Knight PR III,
    9. Chess PR,
    10. Notter RH
    . 2007. Surfactant dysfunction and lung injury due to the E. coli virulence factor hemolysin in a rat pneumonia model. Am J Physiol292:L632–L643.
    OpenUrl
  100. 100.↵
    1. Johnson JR,
    2. Clermont O,
    3. Menard M,
    4. Kuskowski MA,
    5. Picard B,
    6. Denamur E
    . 2006. Experimental mouse lethality of Escherichia coli isolates, in relation to accessory traits, phylogenetic group, and ecological source. J Infect Dis194:1141–1150. doi:10.1086/507305.
    OpenUrlCrossRefPubMedWeb of Science
  101. 101.↵
    1. Heijmans J,
    2. van Lidth de Jeude JF,
    3. Koo BK,
    4. Rosekrans SL,
    5. Wielenga MC,
    6. Van de Wetering M,
    7. Ferrante M,
    8. Lee AS,
    9. Onderwater JJ,
    10. Paton JC,
    11. Paton AW,
    12. Mommaas AM,
    13. Kodach LL,
    14. Hardwick JC,
    15. Hommes DW,
    16. Clevers H,
    17. Muncan V,
    18. Van den Brink GR
    . 2013. ER stress causes rapid loss of intestinal epithelial stemness through activation of the unfolded protein response. Cell Rep3:1128–1139. doi:10.1016/j.celrep.2013.02.031.
    OpenUrlCrossRefPubMedWeb of Science
  102. 102.↵
    1. VanDussen KL,
    2. Marinshaw JM,
    3. Shaikh N,
    4. Miyoshi H,
    5. Moon C,
    6. Tarr PI,
    7. Ciorba MA,
    8. Stappenbeck TS
    . 2015. Development of an enhanced human gastrointestinal epithelial culture system to facilitate patient-based assays. Gut64:911–920. doi:10.1136/gutjnl-2013-306651.
    OpenUrlAbstract/FREE Full Text
  103. 103.↵
    1. Ettayebi K,
    2. Crawford SE,
    3. Murakami K,
    4. Broughman JR,
    5. Karandikar U,
    6. Tenge VR,
    7. Neill FH,
    8. Blutt SE,
    9. Zeng XL,
    10. Qu L,
    11. Kou B,
    12. Opekun AR,
    13. Burrin D,
    14. Graham DY,
    15. Ramani S,
    16. Atmar RL,
    17. Estes MK
    . 2016. Replication of human noroviruses in stem cell-derived human enteroids. Science353:1387–1393. doi:10.1126/science.aaf5211.
    OpenUrlAbstract/FREE Full Text
  104. 104.↵
    1. Ferraretto A,
    2. Gravaghi C,
    3. Donetti E,
    4. Cosentino S,
    5. Donida BM,
    6. Bedoni M,
    7. Lombardi G,
    8. Fiorilli A,
    9. Tettamanti G
    . 2007. New methodological approach to induce a differentiation phenotype in Caco-2 cells prior to post-confluence stage. Anticancer Res27:3919–3926.
    OpenUrlAbstract/FREE Full Text
  105. 105.↵
    1. Cronin CA,
    2. Gluba W,
    3. Scrable H
    . 2001. The lac operator-repressor system is functional in the mouse. Genes Dev15:1506–1517. doi:10.1101/gad.892001.
    OpenUrlAbstract/FREE Full Text
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Role for FimH in Extraintestinal Pathogenic Escherichia coli Invasion and Translocation through the Intestinal Epithelium
Nina M. Poole, Sabrina I. Green, Anubama Rajan, Luz E. Vela, Xi-Lei Zeng, Mary K. Estes, Anthony W. Maresso
Infection and Immunity Oct 2017, 85 (11) e00581-17; DOI: 10.1128/IAI.00581-17

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Role for FimH in Extraintestinal Pathogenic Escherichia coli Invasion and Translocation through the Intestinal Epithelium
Nina M. Poole, Sabrina I. Green, Anubama Rajan, Luz E. Vela, Xi-Lei Zeng, Mary K. Estes, Anthony W. Maresso
Infection and Immunity Oct 2017, 85 (11) e00581-17; DOI: 10.1128/IAI.00581-17
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    • ABSTRACT
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KEYWORDS

Adhesins, Escherichia coli
Bacterial Translocation
epithelial cells
Escherichia coli Infections
Extraintestinal Pathogenic Escherichia coli
Fimbriae Proteins
Fimbriae, Bacterial
ExPEC
FimH
intestinal epithelium
invasion
translocation

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