# Understanding Host-Adherent-Invasive Escherichia coli Interaction in Crohn's Disease: Opening Up New Therapeutic Strategies

Allison Agus, Sébastien Massier, Arlette Darfeuille-Michaud, Elisabeth

Billard, Nicolas Barnich

► To cite this version:

Allison Agus, Sébastien Massier, Arlette Darfeuille-Michaud, Elisabeth Billard, Nicolas Barnich. Understanding Host-Adherent-Invasive Escherichia coli Interaction in Crohn's Disease: Opening Up New Therapeutic Strategies. BioMed Research International, 2014, 2014, pp.1 - 16. 10.1155/2014/567929. [hal-01931428]

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<figure><img src="image_1.png" /></figure>
## Review Article

# Understanding Host-Adherent-Invasive *Escherichia coli* Interaction in Crohn's Disease: Opening Up New Therapeutic Strategies

Allison Agus,<sup>1,2</sup> Sébastien Massier,<sup>1,2</sup> Arlette Darfeuille-Michaud,<sup>1,2,3</sup> Elisabeth Billard,<sup>1,2,3</sup> and Nicolas Barnich<sup>1,2,3</sup>

Correspondence should be addressed to Nicolas Barnich; nicolas.barnich@udamail.fr

Received 29 July 2014; Revised 16 September 2014; Accepted 16 September 2014; Published 15 December 2014

Academic Editor: Alfredo Torres

Copyright © 2014 Allison Agus et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

A trillion of microorganisms colonize the mammalian intestine. Most of them have coevolved with the host in a symbiotic relationship and some of them have developed strategies to promote their replication in the presence of competing microbiota. Recent evidence suggests that perturbation of the microbial community favors the emergence of opportunistic pathogens, in particular adherent-invasive *Escherichia coli* (AIEC) that can increase incidence and severity of gut inflammation in the context of Crohn's disease (CD). This review will report the importance of AIEC as triggers of intestinal inflammation, focusing on their impact on epithelial barrier function and stimulation of mucosal inflammation. Beyond manipulation of immune response, restoration of gut microbiota as a new treatment option for CD patients will be discussed.

## 1. Introduction

A population of 10¹⁴ commensal microorganisms composes the human gut microbiota. Their genome (also named metagenome or microbiome) represents, in terms of gene number, 150-fold the human genome [1]. Microbiota influences physiology and metabolism within the body. In addition to influencing the metabolism of the host, microbiota could also be involved in various pathological mechanisms. Both development and activation of our mucosal immune system in GI tract depend on this complex consortium of microorganisms [2]. Recent evidence has pointed to the role of gut microbiota in various human diseases such as IBD, colon cancer, type 1 diabetes, insulin resistance, nonalcoholic fatty-liver disorders, asthma, and allergies. Thus, it is important to understand the involvement of microbiota in the etiology of such diseases by characterizing species that compose a “healthy” microbiota [3–8].

In inflammatory bowel diseases (IBD), including Crohn's disease (CD) and ulcerative colitis (UC), a dysfunction of the immune response to gut microbiota occurs in a context of host genetic predisposition. CD is a chronic and commonly disabling inflammatory disorder of the intestine whose prevalence and incidence increase in the developed countries [9]. IBD preferentially occurs in the colon and the distal ileum, intestinal portions harboring the largest concentration of microorganisms. Involvement of microbiota in IBD pathogenesis was supported by experiments performed in germ-free animal models since the presence of microbiota was required to trigger intestinal inflammation in various models (IL-10 and IL-12 knock-out mice, chemically DSS- and TNBS-induced colitis) [10, 11]. More recently, genetic evidence has shown associations between IBD and genes involved in antibacterial response, such as NOD2, autophagy-related genes, and the IL23R pathway involved in Th17 polarization. Several nonexclusive mechanisms could

<footnote n="1"><sup>1</sup>Clermont Université, M2iSH, UMR 1071 INSERM/Université d'Auvergne, CBRV, 28 place Henri Dunant, 63001 Clermont-Ferrand, France</footnote>

<footnote n="2"><sup>2</sup>Unité Sous Contrat 2018, Institut National de la Recherche Agronomique, 63001 Clermont-Ferrand, France</footnote>

<footnote n="3"><sup>3</sup>Institut Universitaire de Technologie, Génie Biologique, 63172 Aubière, France</footnote>
drive the pathogenic immunologic response to microbiota: (i) involvement of microbial pathogens that induce intestinal inflammation, such as traditional pathogens (*Mycobacterium avium* subspecies *paratuberculosis*) or functional alteration of commensal bacteria (adherent-invasive *Escherichia coli*, toxigenic *Bacteroides fragilis*, etc.), (ii) dysbiosis of commensal microbiota, with a depletion of protective bacterial species versus an enrichment of harmful species, (iii) host genetic inability to contain commensal microbiota due to defective intracellular bacterial killing and impaired intestinal barrier function, and (iv) defective host immunoregulation.

## 2. Importance of *Escherichia coli* as Triggers of Intestinal Inflammation in Crohn's Disease

An altered gut microbiota has long been suspected to play an important part in the pathogenesis of IBD. The evidence that enteric bacterial antigens continuously drive chronic, immune-mediated colitis and ileitis is provided by rodent models of spontaneous or induced intestinal inflammation [12].

2.1. *Dysbiosis*. A general dysbiosis of gut microbiota has been well established in IBD patients by both culture-dependent and culture-independent techniques [13, 14]. This altered composition of the commensal bacterial populations may result from a modulation of oxygen levels in inflamed gastrointestinal tract, leading to an overgrowth of bacteria having proinflammatory properties and/or to a decrease of beneficial commensal species [15]. Although a specific pattern of dysbiosis in IBD patients is difficult to establish, many studies have reported an increase in the abundance of Proteobacteria and Bacteroidetes and a decrease in Firmicutes [16]. In samples from multiple gastrointestinal locations in a large pediatric CD cohort collected prior to treatment in new-onset cases, an increased representation of Enterobacteriaceae, Veillonellaceae, Fusobacteriaceae, and Pasteurellaceae populations and a reciprocal decrease in Bacteroidales, Clostridiales, and Erysipelotrichales were strongly associated with disease status [17]. This study also indicated that, at the early stage of the disease, analysis of the rectal mucosal-associated microbiota could help to diagnose CD. More recently, analysis of fungal microbiota showed that its composition differs in inflamed and noninflamed area, suggesting that gut fungal exploration could be used to evaluate CD disease activity [18]. Now, intestinal microbiota should be investigated at the ecological level. A recent study reported that, on intestinal mucosal surface, bacterial community is organized into five highly conserved modules in human, two of them displaying distinct metabolic functionalities and being reciprocally associated with IBD. An integrative view of microbial ecology associated with IBD status of individual patients during disease was possible based on the analysis of microbial modules organization [19]. *Bacteroides fragilis*, a human symbiont, had anti-inflammatory effects, via expression of polysaccharide A (PSA) in *Helicobacter hepaticus*-induced colitis in mice [2, 4]. On resected ileal

Crohn's mucosa, decreased levels of *Faecalibacterium prausnitzii* population were associated with endoscopic postoperative recurrence [20]. *F. prausnitzii*, a beneficial bacteria, is known to induce an immunoregulatory cytokine secretion in peripheral blood mononuclear cells with high amounts of IL-10 and low amounts of IL-12 [20, 21]. In the fecal microbiota of UC patients, decreased levels of the butyrate-producing *Roseburia hominis* and *Faecalibacterium prausnitzii* were recently reported [22]. Distinct ratio of *F. prausnitzii* and *E. coli* has been reported in ileal and colonic CD, respectively, therefore allowing to consider this ratio as a promising biomarker for differential diagnosis and personalized treatment [23].

2.2. *Traditional Pathogens*. Molecular techniques have identified specific pathogenic agents playing a role in inflammation of IBD. Much research has shown a higher prevalence of *Mycobacterium avium paratuberculosis*, *Helicobacter* species, and *Campylobacter concisus* in IBD patients than in control subjects [24–26]. Other bacterial pathogens are also suspected of involvement in these diseases, such as *Fusobacterium*, *Klebsiella*, *Salmonella*, and *Yersinia* [27–31]. Despite recurrent indications that traditional pathogens could be involved in IBD, their role as causative agents of CD remains very uncertain. However, it is possible that only a subset of patients is concerned and investigation is needed especially in those with defects in intracellular killing of bacteria due to genetic polymorphisms (ATG16L1, IGRM, or NCF4).

2.3. *Adherent-Invasive E. coli*. Over the last 10–15 years, the microbe that has attracted the most attention, with respect to CD etiology, is *Escherichia coli* [32, 33]. Overgrowth of *E. coli* population in inflammatory bowel disease patients is currently unexplained but may be related to increased production of reactive nitrogen species allowing nitrate respiration, which confers *E. coli* a fitness advantage [34]. A specific pathogenic group of *E. coli*, called adherent-invasive *E. coli* (AIEC), has been extensively implicated in human CD and is currently one of the most exciting players in the pathogen story (Table 1) [23, 35–66]. AIEC bacteria strongly adhere to and invade intestinal epithelial cells (IEC) by a mechanism involving microtubule polymerisation and actin recruitment [67], inducing inflammatory cytokine secretion [68]. AIEC survive and replicate inside macrophages, induce an important secretion of TNF-α, and promote granuloma formation *in vitro* [69–71]. AIEC strains induce IL-1β *via* an NLRP3-dependent mechanism, but their elimination by macrophages is independent of NLRP3 [72]. Invasiveness of intracellular *E. coli* strains into the intestinal mucosa and IL-1β production may contribute to CD and UC pathogenesis. AIEC strains have been shown to be the cause of granulomatous colitis in Boxer dogs and to induce granulomas, similar to early epithelioid granulomas, *in vitro* [71, 73]. AIEC have type one pili and flagella that can bind to host adhesion receptor carcinoembryonic antigen-related cell adhesion molecule 6 (CEACAM6) [74]. CEACAM6 has been shown to be overexpressed in ileal CD tissue compared to healthy controls, to be increased after IFN-γ or TNF-α stimulation, and to be upregulated by AIEC themselves [74]. AIEC have
<table><caption>TABLE 1: Abnormal prevalence of *Escherichia coli* in Crohn's disease patients.</caption><thead><tr><th>Date</th><th>Country</th><th>Method</th><th>Sample</th><th>References</th></tr></thead><tbody><tr><td>1978</td><td>United Kingdom</td><td>Antibody</td><td>Blood</td><td>[35]</td></tr><tr><td>1978</td><td>United Kingdom</td><td>Culture</td><td>Ileal and colonic biopsies</td><td>[36]</td></tr><tr><td>1995</td><td>United States of America</td><td>Immunocytochemical</td><td>Intestines and mesenteric lymph node specimens</td><td>[37]</td></tr><tr><td>1997</td><td>The Netherlands</td><td>DNA probe</td><td>Rectal biopsies and feces</td><td>[38]</td></tr><tr><td>1998</td><td>France</td><td>Culture</td><td>Ileal biopsies</td><td>[39]</td></tr><tr><td>2001</td><td>France</td><td>Ribotyping</td><td>Ileal biopsies</td><td>[40]</td></tr><tr><td>2002</td><td>Japan</td><td>qPCR<sup>1</sup></td><td>Small and large intestine and ileocolitis biopsies</td><td>[41]</td></tr><tr><td>2004</td><td>Ireland</td><td>Nested PCR</td><td>Microdissected granulomas</td><td>[42]</td></tr><tr><td>2004</td><td>United Kingdom</td><td>Culture</td><td>Ileal, ileocolonic, and colonic biopsies</td><td>[43]</td></tr><tr><td>2004</td><td>France</td><td>Culture</td><td>Ileal biopsies</td><td>[44]</td></tr><tr><td>2005</td><td>United Kingdom</td><td>FISH<sup>2</sup></td><td>Rectal biopsies</td><td>[45]</td></tr><tr><td>2006</td><td>United States of America</td><td>OmpC Antibody</td><td>Serum</td><td>[46]</td></tr><tr><td>2007</td><td>Canada</td><td>Culture</td><td>Ileocolonic and colonic biopsies</td><td>[47]</td></tr><tr><td>2007</td><td>United Kingdom</td><td>qPCR<sup>1</sup></td><td>Ileal biopsies</td><td>[48]</td></tr><tr><td>2007</td><td>United States of America</td><td>Culture</td><td>Colonic biopsies</td><td>[49]</td></tr><tr><td>2009</td><td>France</td><td>qPCR<sup>1</sup></td><td>Ileal biopsies</td><td>[50]</td></tr><tr><td>2009</td><td>Sweden</td><td>qPCR<sup>1</sup></td><td>Ileal biopsies</td><td>[51]</td></tr><tr><td>2009</td><td>Denmark</td><td>Culture</td><td>Feces</td><td>[52]</td></tr><tr><td>2010</td><td>Germany</td><td>Cloning</td><td>Colonic biopsies</td><td>[53]</td></tr><tr><td>2010</td><td>Germany</td><td>qPCR<sup>1</sup></td><td>Feces</td><td>[54]</td></tr><tr><td>2010</td><td>Australia</td><td>Microarray</td><td>Feces</td><td>[55]</td></tr><tr><td>2011</td><td>France</td><td>qPCR<sup>1</sup></td><td>Feces</td><td>[56]</td></tr><tr><td>2011</td><td>Brazil</td><td>Culture</td><td>Rectal biopsies</td><td>[57]</td></tr><tr><td>2012</td><td>Brazil</td><td>Culture</td><td>Ileal, colonic, and rectal biopsies</td><td>[58]</td></tr><tr><td>2013</td><td>United States of America</td><td>Culture</td><td>Ileal biopsies</td><td>[59]</td></tr><tr><td>2013</td><td>China</td><td>qPCR<sup>1</sup></td><td>Feces</td><td>[60]</td></tr><tr><td>2013</td><td>United Kingdom</td><td>culture</td><td>Ileal, ileocolonic, and colonic biopsies</td><td>[61]</td></tr><tr><td>2014</td><td>Spain</td><td>qPCR<sup>1</sup></td><td>Ileal, ileocolonic, and colonic biopsies</td><td>[23]</td></tr><tr><td>2014</td><td>Australia</td><td>qPCR<sup>1</sup></td><td>Ileal biopsies</td><td>[62]</td></tr></tbody></table>

also been shown to possess long polar fimbriae and so can cross the mucosal barrier to access lymphoid cells [75]. AIEC LF82 bacteria isolated from an ileal CD patient exacerbate an inflammatory mucosal immune response involving upregulation of TLR5 (toll-like receptor) and IPAF flagellin receptors [76]. In CD patients, increased expression of CEACAM6 on the apical membrane of ileal enterocytes could promote the abnormal ileal mucosa colonization by AIEC bacteria, since CEACAM6 acts as a receptor for AIEC attachment to the intestinal mucosa [74]. In transgenic CEABAC10 mice expressing human CEACAMs to mimic the high expression of CEACAM6 reported in CD patients, the AIEC reference strain LF82 induced development of severe clinical symptoms of colitis in a type 1 pili dependent manner [77, 78]. Additionally, another AIEC strain NRG857c has been reported to colonize intestinal mucosa of conventional mice following streptomycin treatment, leading to chronic inflammation involving Th1 and Th17 responses and intestinal fibrosis [79]. The abnormal persistence of AIEC bacteria in this model could be related to their recently reported ability to actively

resist antimicrobial peptides secreted by intestinal cells [80]. Of note, gut microbiota composition and host mucosal homeostasis are altered in CEABAC10 mice submitted to Western diet, favouring AIEC bacteria colonization of gut mucosa [81]. This is in line with the multifactorial etiology of CD and emphasizes the role of diet in CD pathogenesis.

2.4. *Is AIEC an Instigator or a Propagator of Colitis?* At present, it is difficult to determine whether AIEC bacteria trigger intestinal inflammation, thus leading to the disease, or whether they colonize the gut mucosa as a consequence of preexisting inflammatory context in which case they could be an aggravating factor. AIEC colonized WT and TLR5KO mice only transiently but chronic colitis persisted in TLR5KO months later, suggesting that this microbe acted as an instigator, rather than a propagator, of colitis [82]. Inversely, inflammation leads to a shift from Gram+ to Gram-, a proliferation of mucosally invasive *E. coli* and a decrease in microbial diversity [83]. An answer concerning the origin of AIEC persistence may be proposed in the future

<footnote n="1">¹Quantitative polymerase chain reaction; ²fluorescent in situ hybridization.</footnote>
with the development of fecal sample banks collecting spec-
imens throughout life from patients who develop IBD and
control subjects. With the current knowledge, AIEC bacteria
are considered as an intestinal pathobiont able to promote
disease only in specific host genetic or environmental con-
texts (Figure 1). Pathobionts were thus termed to distinguish
them from acquired infectious agents [4, 84]. Host factors
expressed specifically during intestinal inflammation have
been shown to play major roles in facilitating infection with
enteric bacteria and especially with AIEC. Ileal lesions in CD
patients are colonized by pathogenic AIEC bacteria owing
to the increased expression of a specific bacterial attachment
molecule, CEACAM6, at the brush border of the ileal epithe-
lium [78]. AIEC are also able to adhere to chitinase 3-like-
1 receptor (CHI3L1) via the chitin-binding domain of ChiA
bacterial protein promoting the pathogenic effect of AIEC
in IBD [85]. AIEC outer membrane protein OmpA has been
shown to interact with the endoplasmic reticulum (ER) stress
response glycoprotein Gp96, which is also overexpressed at
the apical membrane of ileal epithelial cells in CD patients
[86]. Given that ER stress is commonly associated with
inflammation [87], AIEC may also take advantage of the
ER stress occurring in CD patients to increase adherence to
the intestinal epithelium. AIEC delay apoptosis in infected
macrophages, favoring their own persistence in CD patients,
by a mechanism involving increase of S-nitrosylation and
proteasomal degradation of caspase-3 [88]. In addition, AIEC
bacteria modulate the ubiquitin proteasome system turnover
in infected-intestinal epithelial cells by downregulating the
NF-κB regulator CYLD, leading to IκB-α degradation and
NF-κB activation. This property plays a key role in the
pathogenicity of AIEC since it favours intracellular repli-
cation of AIEC reference strain LF82 [89]. An abnormal
autophagy, an innate defense mechanism allowing clearance
of intracellular pathogens, could also favor AIEC persistence
in the gut of IBD patients, as supported by several recent stud-
ies. Indeed, altered expression of ATG16L1, IRGM, or NOD2
favoured intramacrophagic replication of AIEC and led to
enhanced secretion of IL-6 and TNF-α in response to AIEC
infection [90]. Conversely, the numbers of intramacrophagic
AIEC and proinflammatory cytokine release are strongly
decreased upon pharmacological induction of autophagy
[90]. Of note, autophagy is blocked at the autolysosomal step
following AIEC infection of neutrophil-like PLB-985 cells,
allowing intracellular survival of bacteria and increased IL-
8 secretion [91]. Moreover, microRNAs MIR106B and MIR93
decrease the expression of ATG16L1 and prevent autophagy-
mediated elimination of intracellular bacteria, a process
that seems to be impaired in colonic mucosa of patients
with active CD [92]. AIEC infection upregulated levels of
microRNA- (MIR-) 30C and MIR130A in T84 cells and in
mouse enterocytes, leading to reduced levels of ATG5 and
ATG16L1 and to inhibition of autophagy, increased numbers
of intracellular AIEC, and increased inflammatory response
[66].

During the latter 20th century, increased CD incidence
has been associated with consumption of polysaccharides
in Western diets. AIEC LF82 specific biofilm formation was
strongly favored in the presence of maltodextrin (MDX),

a starch-derived polysaccharide [93]. MDX also promoted
bacterial adhesion to human intestinal epithelial cells *via* a
mechanism involving type 1 pili. However, this was indepen-
dent of the expression of CEACAM6, indicating a distinct
mechanism of AIEC adhesion to enterocytes [93]. As MDX is
an ubiquitous dietary component, this suggests that Western
diets, enriched in specific polysaccharides, may contribute
to dysbiosis and lead to disease susceptibility. This is also
supported by alteration of microbiota composition reported
in CEABAC10 mice fed a Western diet [81].

The concept of pathobionts is supported by clinical data
which reveal that, in IBD patients with underlying genetic
mutations, inflammation may be driven by specific members
of the microbiota rather than by infectious pathogens [94].
The analysis of AIEC genome revealed the presence of specific
genes that could be involved in bacterial virulence, but
pathoadaptive mutations in many other genes or bacterial
DNA sequences could also participate in AIEC pathogenicity
in a susceptible host [95, 96]. For example, OmpA proteins
of LF82 bacteria interact with the host molecule Gp96 and
allow adhesion to intestinal epithelial cells [86]. Recently
acquired nonsynonymous substitutions are considered as a
typical signature of the pathoadaptive evolution of bacterial
pathogens and have notably been reported in FimH variants
expressed by AIEC strains, conferring them higher adhesion
ability [97]. FimH pathoadaptive mutations required for
AIEC gut colonization have thus been selected, leading to
the development of inflammation in a genetically susceptible
host [97]. Therefore, fimH SNPs analysis could be a marker
of virulence for IBD patients' *E. coli* strains and could be
used for diagnosis or epidemiological studies. Moreover, new
therapeutic strategies to impair AIEC adhesion to the gut
mucosa in the early stages of IBD could be considered. Of
interest, the protease meprin has been shown to degrade
type 1 pili of AIEC bacteria and prevent their binding
to mannosylated host-receptors [98]. Meprin expression is
decreased in CD patients and this decrease correlates with
the severity of inflammation, suggesting that the lack of
protective meprin could favor AIEC colonization of gut
mucosa [98]. AIEC, and perhaps other pathobionts, may
therefore instigate chronic inflammation in susceptible hosts
by altering the gut microbiota composition.

3. Crosstalk between Epithelial Barrier and Adherent-Invasive *E. coli*

The gastrointestinal epithelium forms a single-cell layer
between the blood circulation and the external environment
of the intestinal lumen [99]. Functions of intestinal barrier
are to control uptake across the mucosa and to protect
mucosa against intraluminal toxins, invading microorgan-
isms, and lumen gut antigens [100]. The intestinal epithelium
provides a physical barrier with interconnections between
intestinal epithelial cells. The cell-cell contact is mediated
by different protein complexes including adherens junctions,
tight junctions, and desmosomes to stabilize the mechanical
cohesion of the cells [101]. Also, epithelium acts as a chemical
barrier through the intestinal mucus layer and the synthesis
<figure><img src="image_2.png" /><figcaption>FIGURE 1: Strategies used by AIEC to trigger intestinal inflammation. (1) AIEC are able to strongly adhere to intestinal epithelial cells and colonize gut mucosa using type 1 pili that can bind to mannose residues of CEACAM6, which is overexpressed on the apical surface of ileal epithelial cells in patients with ileal CD. AIEC are also able to adhere to chitinase 3-like-1 receptor (CHI3L1) *via* the chitin-binding domain of ChiA bacterial protein. AIEC actively resist antimicrobial peptides secreted by Paneth cells. This mechanism involves two genes, *arlA*, which encodes a Mig-14 family protein implicated in defensin resistance, and *arlC*, an OmpT family outer membrane protease. (2) AIEC translocation through the epithelial barrier is increased following different mechanisms leading to exacerbation of intestinal inflammation. Modulation of tight junctions (TJs) by AIEC induces paracellular barrier permeability involving ZO-1 redistribution, increased expression of pore-forming claudin-2, and decreased expression of occludin. The endoplasmic reticulum (ER) stress response glycoprotein Gp96 is overexpressed at the apical membrane of ileal epithelial cells in CD patients and acts as a host-receptor for AIEC outer membrane vesicles (OMV) carrying OmpA protein promoting the invasion of the intestinal mucosa. AIEC bacteria interact with Peyer's patches and translocate across M cells *via* long polar fimbriae (LPF) expression to access lymphoid cells. (3) AIEC intramacrophagic replication is favored in the submucosal compartment of host cells. AIEC intramacrophagic survival could be due to host autophagy defects leading to increased bacterial replication and also enhancing inflammatory responses. AIEC can also induce Th17 and CD8+ cytotoxic responses.</figcaption></figure>

of antimicrobial peptides, an integral part of innate immunity. An efficient intestinal mucosal barrier is crucial for protection against the external environment. A barrier dysfunction has been characterized in patients suffering from IBD that leads to enhanced intestinal permeability [102]. Barrier disorders including defects in thickness or composition of the intestinal mucus layer, alterations of tight junctional complexes, and disturbances of the synthesis of antimicrobial peptides result in an inadequate protection of the epithelium against the adherence and invasion of luminal bacteria *via* specific receptors in the epithelium abnormally expressed in the context of intestinal inflammation [101]. Bacterial adhesion to and colonization of intestinal epithelial cells are considered as the crucial initializing steps in IBD pathogenesis before bacteria translocate and enter the submucosal compartment [85]. There is some evidence that, following invasion, AIEC bacteria can alter epithelial barrier function by displacing and redistributing ZO-1, a protein required for the formation of

apical tight junctions [103]. The decrease in barrier integrity could result in an increase in AIEC translocation across the epithelial barrier leading to an exacerbation of AIEC pathogenesis [49]. Gut barrier damage and inflammatory responses are crucial for the perturbation and aggravation of intestinal inflammation [104].

### 3.1. Failure of the Intestinal Mucus Layer and Defective Production of Antimicrobial Peptides.
The small intestine is lined with a thin mucus layer while the colon and stomach are covered by two layers of mucus [105]. The thinner inner layer (50–200 µm) common to the stomach, small intestine, and colon is densely packed and strongly linked to the intestinal epithelium, which ensures its protection. This layer provides a matrix for the retention of antimicrobial peptides, including α-defensins, secreted by Paneth cells, thereby establishing a barrier between microorganisms and mucosal intestinal tissue. This inner layer prevents direct contact of the epithelium
with the luminal microorganisms, whereas the outer layer, much thicker and difficult to dislodge, is colonized by a large number of commensal bacteria [105]. In IBD, deficiencies in mucus production and the secretion of antimicrobial peptides allow commensal bacteria to become opportunistic pathogens and contribute to chronic intestinal inflammation [106]. A reduced mucus layer has been reported to correlate with increased disease severity. Moreover, in IBD patients, the mucolytic species *Ruminococcus torques* and *Ruminococcus gnavus* have also been reported to be more prevalent and more abundant [107]. This disproportionate increase in mucolytic bacteria could explain the total increase in mucosa-associated bacteria in IBD since their ability to degrade human secretory mucin (MUC2) could promote the adhesion and invasion of opportunistic bacteria [107]. In CEABAC10 transgenic mice expressing human CEACAMs, Western diet led to a shift in microbiota composition comparable to what is observed in CD patients, with an increase in the mucin-degrading bacterium *Ruminococcus torques* and the *Bacteroides/Prevotella* group [81]. Western diet altered barrier function by decreasing *Mucin-2*, *Klf4*, and *Tff3* expression, mucus layer thickness, and goblet cell number in colonic mucosa. In these mice, AIEC bacteria have better ability to colonize the gut mucosa and to trigger intestinal inflammation due to alteration of barrier function and increased TNF-α secretion [81]. Moreover, some AIEC strains have been shown to resist antimicrobial peptides, which could promote their survival in the inner mucus layer [80]. A better understanding of the molecular mechanisms involved in increased intestinal permeability in CD patients with a Western diet, in the context of CD genetic susceptibility and in the presence of AIEC, should help the development of new drugs to target AIEC-induced disruption of intestinal barrier integrity.

**3.2. Disruption of Epithelial Barrier Integrity by AIEC in the Context of CD.** The intestinal epithelial cells are joined at their apical side by tight junctions (TJs), a space between the enterocytes that is finely regulated and is crucial in regulating intestinal permeability. Host-microbiota interactions promote a reorganization of TJs [108]. These interactions between pathogenic bacteria and epithelial tissues often disturb the intestinal TJs barrier and often lead to a number of pathophysiological disorders. The alteration of TJs protein content increases paracellular barrier permeability and contributes to intestinal inflammation. Recent studies showed a reduced number of tight junction strands and an increased number of strand breaks in CD patients [109]. Claudins are the major functional and structural components of TJs. Specifically, an increased expression of the pore-forming claudin-2 and a decreased expression of occludin were detectable [109, 110]. Moreover, in CEACAMs-expressing mice colonized with AIEC bacteria, an increase of intestinal permeability has been reported that could be related to the induction of claudin-2 expression consequently to AIEC/CEACAM6 interaction [110]. In a spontaneous model of IBD closely mimicking CD (SAMP1/YitFc mice), a dysregulation of the epithelial barrier function has also

been shown, involving aberrant expression of claudin-2 and occludin and resulting in ileitis with a worsening of histological scores [111, 112]. Intestinal barrier function in CD patients may be restored by defending against type 1 pili-mediated AIEC/CEACAM6 interaction.

**3.3. Peyer's Patches as Portals of Entry for AIEC Bacteria.** Peyer's patches (PPs) play a major role in mucosal immunity. CD pathogenesis results from an inadequate innate and/or adaptive immune response to the microflora supported by the relationship between PPs and CD lesions [113]. PPs have a defined role in the interaction between immune response and microbiota and consequently participate in intestinal epithelial disorders [113]. Their interplay with the diversity and the function of the gut microbiota is becoming an effective area of research [113]. A number of pathogenic microorganisms have evolved original strategies to pass through the apical epithelial barrier and penetrate into the intestinal epithelium. Many studies now indicate that several microorganisms, particularly invasive pathogens, use specialized M cells as the primary portal of entry into the host to cross the intestinal barrier and initiate the disease. For example, *Yersinia enterocolitica* and *Yersinia pseudotuberculosis* cross the intestinal epithelial barrier by adhering to M cells of the follicle-associated epithelium [114]. *Salmonella* Typhimurium also invade M cells and thus access to the PPs, although they have also been reported to be sampled by dendritic cells extending transcellular protrusions through M cells [115, 116]. PPs were suspected to be the site of initial inflammation and thus to play a major role in the early stages of CD disease [117]. A recent study showed that AIEC interact with PPs and translocate through M cells via long polar fimbriae (LPF); moreover, the prevalence of LPF-expressing AIEC strains was higher among CD patients than among control subjects [75]. Following translocation through the epithelium, AIEC bacteria could be internalized into immune cells, especially macrophages and dendritic cells, which are able to release inflammatory mediators such as TNF-α that can drive functional alterations of the mucosal barrier and lead to general mucosal permeability defects [100].

# 4. Handling of Microbiota-Derived Antigens in Health and Inflammatory Bowel Diseases

The intestinal mucosa contains high numbers of effector lymphocytes, including IgA-producing plasma cells and effector CD4+ T cells, among them IFN-γ-producing Th1 cells, IL-17-producing Th17 cells, and Foxp3-expressing regulatory T cells (Tregs), but also CD8+ T cells and intraepithelial lymphocytes (mainly γδ T cells) [118]. In addition, recent studies have revealed the importance of innate lymphoid cells that share functional characteristics with T cells [119].

**4.1. Microbial Containment by Mucosal Firewall.** The intestinal immune system needs to ensure simultaneously the immune tolerance of microbiota and host defense against microbial invasion whether by pathogens or by commensals taking advantage of occasional barrier weakness. Among
other mechanisms, this is achieved by the reciprocal regula-
tion of inflammatory and regulatory immune responses [120].
Inflammatory responses are kept under control by FoxP3+
Tregs originating either from the thymus or from local differ-
entiation of naive CD4+ T cells. Surprisingly, a recent report
suggested that intestinal Tregs are mainly of thymic origin
and not locally induced following exposure to commensal
or diet antigens as previously assumed [121]. Of note, innate
lymphoid cells have also been shown to play a major role
in the regulation of effector T cell responses to commensals
[122]. Thus, at steady-state, the “mucosal firewall,” comprising
mucus layer, epithelial barrier, IgA, and regulatory cells, con-
tains microbiota- and food-derived antigens and limits inap-
propriate immune responses [123]. Microbiota-derived anti-
gens do not, therefore, normally stimulate systemic immunity
[124]. Captured either directly or more probably transferred
from other mucosal phagocytes, these antigens are then
handled by CD103+ dendritic cells, known to preferentially
induce Treg differentiation and homing to the gut mucosa
through the production of retinoic acid [125]. Pathogens are
then recognized on the basis of their invasiveness and sensed
by pattern-recognition receptors (PRR) expressed either in
an intracellular way or in the basolateral compartment of
the epithelium [126]. Any disruption of the mucosal firewall,
even local and transient, caused, for instance, by acute infec-
tion, epithelial damage, or just by increased permeability,
could thus allow for microbial translocation and impair
discrimination between commensals and pathogens. In line
with this, it is known that acute infection can lead to loss
of tolerance to commensals and induction of microbiota-
specific T cells with inflammatory phenotype [127]. Immune
memory cells are probably generated following this kind
of event and could be reactivated, as suggested by the
detection of antibodies to microbiota in healthy human
serum [128]. How the mucosal firewall is restored following
such events and whether commensal-specific memory T
cells remain and take part in the characteristic alternation
of clinical relapse and remission in IBD still need to be
elucidated.

4.2. Reciprocal Shaping of Microbiota and Mucosal Immunity.
Commensal flora plays an essential part in the development
of gut-associated lymphoid structures, that is, PPs and
isolated lymphoid follicles, as shown by studies on germ-free
animals [129]. Microbiota has also been recognized as a major
regulator of mucosal immune system activation and tuning,
through direct interactions with epithelial or immune
cells or by producing immunomodulatory metabolites
[120].

The microbiota stimulates regulatory responses, mainly
through the production of active metabolites. *Clostridium*
clusters IV and XIVa, which are decreased in IBD patients'
flora, are known to induce colonic IL-10-producing cells
[130, 131]. Belonging to this group of microorganisms, *Fae-*
*calibacterium prausnitzii* secrete a still unidentified factor
exerting anti-inflammatory effects *in vitro* on human intesti-
nal epithelial cells and peripheral blood mononuclear cells
[20]. In addition, intragastric administration of *F. prausnitzii*

or their supernatant ameliorates TNBS colitis in mice [20].
An increase in Treg frequency was also reported recently
in mice fed probiotics such as *Lactobacillus reuteri* [132].
Bacterial fermentation products, namely, short chain fatty
acids (SCFA) such as butyrate and acetate produced by
Bacteroidetes phylum or Clostridia, suppress inflammation
and have a protective effect against colitis by favoring the
differentiation and function of colonic Tregs in a GPR43-
dependent manner [133–135]. Other bacterial products such
as polysaccharide A from nonenterotoxinogenic *Bacteroides
fragilis* also promote expansion of IL-10-producing FoxP3+
Tregs and ameliorate colitis in mice [136, 137].

Conversely, evidence of effector T cell stimulation by
microbiota has also been documented. Segmented filamen-
tous bacteria (SFB) closely adhere to PPs and potently
stimulate Th17 differentiation in mice, even though they
also induce other types of helper T cells [138, 139]. Other
microbial products such as bacterial DNA also stimulate
Th17 activation at steady-state [140]. These RORγT+, IL-17
secreting CD4+ T cells are often considered detrimental for
the host owing to their recurrent implication in inflammatory
and autoimmune pathogenesis. For instance, Th17 response
to enterotoxinogenic *Bacteroides fragilis* induces colitis and
tumor formation in Min mice and SFB colonization has also
been reported to aggravate autoimmune arthritis in mice
[141, 142]. However, severe DSS-induced colitis in RORγT
deficient mice is reversed by antibiotic treatment suggesting
a crucial role for Th17 cells in microbiota containment and
mucosal homeostasis at steady-state [143]. In addition, Th17
response clearly plays a protective role against fungal and
bacterial enteric pathogens such as *Citrobacter rodentium*
[138]. Thus, Th17 responses probably contribute to intestinal
homeostasis at steady-state and become pathogenic only in
particular contexts such as a host autoimmune susceptibility
or immune overstimulation due to massive bacterial translo-
cation. According to this view, the increased expression of
IL-17 and IL-22 reported in CD patients’ mucosa would not
necessarily mean a pathogenic role for these cytokines but
could rather be the signature of an abnormal stimulation of
Th17 antimicrobial immunity. The deregulated expansion of
Th17 population, combined with the reinforcement of their
proinflammatory role by IL-23, would make them become
harmful and participate in the development of inflammatory
diseases [144].

Thus, gut microbiota seems to play a dual role in
immune regulation. By stimulating the adaptive immune
system and promoting the generation of different T cell
subsets in the gut mucosa, normal intestinal flora con-
tributes to immune homeostasis while being detrimental
in autoimmune models such as arthritis and experimental
autoimmune encephalomyelitis [136, 141]. Of note, Treg cells
in gut mucosa are specific to microbiota-derived antigens
and any perturbation of microbiota composition strongly
influences Treg repertoire [121]. Likewise, antibody repertoire
has been shown to be progressively shaped by changes in
microbiota and to adjust to the latest microorganisms present
[145]. Reciprocally, the shaping of microbiota composition by
the immune system has been demonstrated in several mouse
models, among them NOD2-deficient mice, PGRP- (pep-
tidoglycan recognition protein-) deficient mice, and TRUC
mice, in which immune defects induce the establishment of a
colitogenic flora able to transfer colitis to healthy mice [146–
148].

4.3. *Antigenic Stimulation of Adaptive Immunity in Inflammatory Bowel Diseases.* IBD is generally considered as the result of an inappropriate response of the adaptive immune system to microbiota-derived antigens. This view is supported by the fact that most genetic polymorphisms associated with a higher risk of IBD affect the responsiveness of the mucosal immune system to microorganisms [149]. This excessive stimulation is thought to induce effector T cell responses: CD has long been considered as a Th1 disease with increased levels of TNF-α, IFN-γ, and IL-12 in patients' inflamed mucosa whereas increased concentrations of Th2 cytokines such as IL-5 and IL-13 are rather a feature of UC [150–152]. This notion was next challenged by the recognition of excessive Th17 infiltration and elevated concentrations of IL-23, IL-17, IL-22, and IL-21 in patients' inflamed mucosa. A gain-of-function mutation on IL23R gene has been reported to predispose patients to both CD and UC whereas IL-17 secretion by PBMC of patients correlates with disease severity in UC but not in CD, suggesting a major role of the IL-23 in the pathogenesis of IBD but possibly a different involvement of Th17 cells in the two disorders [153, 154]. Of note, IL-23 not only sustains Th17 response but also promotes IL-17 and IFN-γ secretion by innate lymphoid cells, leading to colitis in mice [155]. These innate lymphoid cells are abnormally represented in IBD subjects mucosa compared to healthy controls [156]. A functional plasticity between Th1 and Th17 lineage has also been proposed and is consistent with the identification in CD patients of pathogenic Th1/Th17 cells releasing both IL-17 and IFN-γ [157]. Thus, Th1 and Th2 responses are currently considered as the true immunopathogenic components driving inflammation, respectively, in CD and UC, whereas Th17 response could initiate the deregulation of these effector responses [152].

While the exact nature of antigens stimulating the immune system remains elusive, evidence for microbiota-induced activation of B cell and T cell immunity does not suggest the involvement of a unique, pathogenic antigen in disease etiology but rather a generalized loss of tolerance to microbiota in IBD subjects. Elevated levels of antibodies directed against microbial structures, among them antibodies to *Saccharomyces cerevisiae* (ASCA) and to *E. coli* membrane protein OmpC and flagellin, have been found in IBD patients and associated with aggressive forms of the disease [158]. These systemic antibodies have also been detected in patients' unaffected relatives, in whom they are predictive of IBD development [159]. Of note, although there is no data available concerning the possible correlation between AIEC colonization and OmpC antibodies in CD patients, both have been specifically associated with ileal involvement [44, 160]. However, even if their concentration is strongly increased in IBD, antibodies against gut-resident flora also occur in healthy subjects, probably following transient barrier

dysfunction [128]. Whether these responses are cross-reactive
antibodies against conserved bacterial antigens is currently
not known. Nevertheless the simple presence of microbiota-
specific systemic immunity clearly cannot account by itself
for the development of IBD.

Regarding T cell response and in line with what has been found for antibody response, the diversity of the TCR repertoire is affected in CD patients and oligoclonal expansions of CD4+ T cells persisting after surgery have been detected in inflamed as well as noninflamed mucosa [161]. No common TCR specificity has been identified among patients, suggesting there is no shared antigenic response at the origin of these abnormal T cell oligoclonal proliferations. Of note, chronic infection with adherent-invasive *E. coli* has been reported to induce Th17 and cytotoxic T cell responses in mice [79]. Altogether, this favors a decisive role of a pathogenic T cell response in the triggering of inflammation in gut mucosa and in postoperative recurrence.

Current knowledge is in line with a multiple-hit model
in which IBD triggering is due to neither host susceptibility
nor the environment nor the microbiota but due to the
concomitant occurrence of intestinal barrier dysfunction that
allows abnormal antigenic stimulation of immunity in a
susceptible host prone to mount uncontrolled inflammatory
responses.

**5. New Treatment Options for CD Patients:
Beyond Manipulation of Immune Response,
Targeting of AIEC and Restoration
of Gut Microbiota?**

Current CD treatments including immunosuppressive agents and synthetic anti-TNF-α or anti-integrin antibodies mainly focus on reducing the symptoms but are unable to treat the cause of the disease insofar as the origin of initial inflammation remains elusive. Despite recent advances in this field, the available drugs are not devoid of side effects and a large subset of CD patients do not respond or undergo loss of responsiveness in the course of their disease [162]. Beyond improvement of diarrhea and abdominal pain, new therapeutic goals now intend to limit mucosal damage and promote mucosal healing in order to achieve long-term deep remission. In the past decade, many promising advances have been made and extensively reviewed in the field of IBD [162–164].

5.1. *Immunomodulation Treatments.* Besides the major breakthrough of the discovery of TNF-α antagonists, the blockade of inflammatory cytokines or their receptors did not reach expectations, with the exception of some therapeutic antibodies targeting the p40 subunit of IL-12 and IL-23, IL-6 receptor, or IL-13, whose efficacy still has to be confirmed [164]. Likewise, systemic administration of anti-inflammatory cytokines such as IL-10 did not result in clinical improvement [165]. Numerous other possible therapeutic agents have been suggested for IBD treatment, including agonists of peroxisome proliferator-activated receptor gamma (PPARγ) modulators and elafin, an endogenous
regulator of protease activity, thereby underlining an urgent need for efficient and safe drug-delivery systems to the gut mucosa [166, 167]. Although this approach does not appear feasible with current regulations, oral administration of genetically modified food-grade bacteria has been considered for local expression of therapeutic molecules at the mucosal surface [168, 169].

New strategies are currently emerging to dampen or manipulate the abnormal immune response in IBD patients. The manipulation of the T cell costimulatory pathway by anti-CD28 antibody Abatacept did not work [170]. However, the humanized antibody to α4β7 integrin Vedolizumab, which prevents homing of immune cells to gut mucosa, has yielded encouraging results in CD [171]. Among innovative strategies in this field, the injection of activated regulatory T cells into CD patients has shown promising potential and could open up the way to personalized immunotherapy [172]. Of note, autologous hematopoietic stem cell transplantation has also been reported to induce durable remission in CD patients refractory to conventional therapies [173-175].

Therapeutic interventions targeting dysbiosis in general and/or AIEC colonization in particular are promising for changing the natural CD history. Today the manipulation of patient microbiota through antibiotherapy, fecal transplantation, nutritional interventions, or pre/probiotic administration could be used either alone or in combination with immunotherapy to induce remission in active disease or as a postoperative therapy to prevent relapse.

**5.2. Antibiotics.** The use of antibiotics is currently restricted to bacterial complications of CD because their efficacy has not been clearly established and because of their side effects [176]. However, antibiotics may induce remission in active CD, especially in patients with colonic involvement [177, 178]. Their efficacy in preventing postoperative recurrence has also been studied, with conflicting results [179, 180]. Antibiotics nevertheless deserve further evaluation in IBD, notably in particular cases such as CD patients with evidence of AIEC colonization. Additionally, in the future it may become possible to specifically target antibiotic activity against aggressive bacterial species, thus reducing side effects and allowing selective elimination of undesirable bacteria.

**5.3. Fecal Transplantation.** Fecal microbiota transplantation is another drastic way of modifying the patient's microbiota and has been successfully tested in recurrent *Clostridium difficile* infections [181, 182]. By restoring essential components of intestinal flora, it could reverse the inappropriate immune stimulation in CD and make intestinal ecosystem less suitable for AIEC intestinal colonization. The mode of delivery and the preparation of donor stools still have to be perfected, and the potential long-term consequences of fecal transplantation remain to be established. Its safety and efficacy in CD patients are currently under investigation, especially in those for whom standard treatments have failed [183]. Nevertheless, the efficacy of fecal transplantation has recently been attested to in CD patients in independent studies [184, 185].

**5.4. Probiotics, Prebiotics, and Postbiotics.** As a promising way of modulating microbiota composition, the administration of presumed anti-inflammatory probiotics has been tested in CD [186]. Their ability to induce remission has not been clearly demonstrated but they may be effective in maintaining remission in postoperative prophylaxis [187-189]. The use of a yeast probiotic has recently been reported to prevent colitis in mice and therefore could represent a new strategy to treat patients with ileal CD that are abnormally colonized by AIEC [190]. As a complementary strategy, nondigestible prebiotics could also be given to stimulate growth or metabolic activity of beneficial microbial species; this approach is currently under investigation in CD patients [191]. New experimental models consisting of polarized explants of healthy or IBD gut mucosa have recently been developed to assess *ex vivo* the effect of probiotics [192]. In these models, some probiotics worsened inflammation in IBD mucosal explants, probably because of increased permeability and higher bacterial translocation, which suggests that the administration of probiotics to IBD patients should be considered with great caution [192]. The safety of probiotic use in active CD has also been recently questioned by the report of a break of tolerance to commensal flora in acute inflammatory context [127]. As an alternative to probiotics, the use of postbiotics (soluble factors produced by probiotics and able to elicit immunomodulatory response) as therapeutic agents could be of interest in CD since they could be administered in a purified and well-characterized form to guarantee their safety [186]. There is thus a need to identify bacterial immunomodulatory factors like, for instance, lactocepin, a serine protease secreted by *Lactobacillus casei* that decreases inflammation in a murine colitis model through selective degradation of proinflammatory cytokines, and to deliver them to the intestinal mucosa in a safe form [193].

**5.5. Phage Therapy.** Finally, the abundance and diversity of bacteriophage communities in the human gut have recently been investigated and certain differences between the bacteriophage colonization of IBD patients and that of healthy controls strongly suggest a possible implication of bacteriophages in IBD [194]. New animal models have been developed to study the dynamics of phage/bacterial communities in the gut that open up a new area of research and therapeutic possibilities [195].

**6. Conclusion**

Despite original therapeutic options available, current CD treatments have important limitations with regard to safety, efficacy, and applicability and often cause severe side effects. Anti-TNF-α have been involved in fatal blood disorders, infections, and liver injury. In the near future, a better understanding of microbiota function in intestinal inflammation will provide new therapeutic opportunities to treat CD patients. There is an accumulation of evidence that CD probably occurs as a result of inappropriate triggering of the mucosal immune system in a host genetic and/or epigenetic susceptibility and under certain dietary or environmental
conditions. The potential multiplicity of etiologies suggests that CD patients need personalized therapeutic strategies: a subgroup of CD patients abnormally colonized by AIEC could benefit from specific treatment aiming at eradicating these bacteria. More generally, there is an urgent need to identify biomarkers that can reliably predict the responsiveness and efficacy of treatments. Microbial signatures could prove to be useful as such biomarkers for diagnosis, for monitoring disease activity, and for therapeutic orientation [196].

## Conflict of Interests

The authors declare that there is no conflict of interests regarding the publication of this paper.

## References

[1] J. Qin, R. Li, J. Raes et al., "A human gut microbial gene catalogue established by metagenomic sequencing," *Nature*, vol. 464, no. 7285, pp. 59-65, 2010.

[2] S. K. Mazmanian, H. L. Cui, A. O. Tzianabos, and D. L. Kasper, "An immunomodulatory molecule of symbiotic bacteria directs maturation of the host immune system," *Cell*, vol. 122, no. 1, pp. 107-118, 2005.

[3] J. M. Kinross, A. C. von Roon, E. Holmes, A. Darzi, and J. K. Nicholson, "The human gut microbiome: implications for future health care," *Current Gastroenterology Reports*, vol. 10, no. 4, pp. 396-403, 2008.

[4] S. K. Mazmanian, J. L. Round, and D. L. Kasper, "A microbial symbiosis factor prevents intestinal inflammatory disease," *Nature*, vol. 453, no. 7195, pp. 620-625, 2008.

[5] J. Penders, E. E. Stobberingh, P. A. V. D. Brandt, and C. Thijs, "The role of the intestinal microbiota in the development of atopic disorders," *Allergy: European Journal of Allergy and Clinical Immunology*, vol. 62, no. 11, pp. 1223-1236, 2007.

[6] L. Wen, R. E. Ley, P. Y. Volchkov et al., "Innate immunity and intestinal microbiota in the development of Type 1 diabetes," *Nature*, vol. 455, no. 7216, pp. 1109-1113, 2008.

[7] F. Bäckhed, R. E. Ley, J. L. Sonnenburg, D. A. Peterson, and J. I. Gordon, "Host-bacterial mutualism in the human intestine," *Science*, vol. 307, no. 5717, pp. 1915-1920, 2005.

[8] M.-E. Dumas, R. H. Barton, A. Toye et al., "Metabolic profiling reveals a contribution of gut microbiota to fatty liver phenotype in insulin-resistant mice," *Proceedings of the National Academy of Sciences of the United States of America*, vol. 103, no. 33, pp. 12511-12516, 2006.

[9] R. J. Xavier and D. K. Podolsky, "Unravelling the pathogenesis of inflammatory bowel disease," *Nature*, vol. 448, no. 7152, pp. 427-434, 2007.

[10] C. O. Elson, R. B. Sartor, G. S. Tennyson, and R. H. Riddell, "Experimental models of inflammatory bowel disease," *Gastroenterology*, vol. 109, no. 4, pp. 1344-1367, 1995.

[11] M. Llopis, M. Antolín, F. Guarner, A. Salas, and J.-R. Malagelada, "Mucosal colonisation with *Lactobacillus casei* mitigates barrier injury induced by exposure to trinitronbenzene sulphonic acid," *Gut*, vol. 54, no. 7, pp. 955-959, 2005.

[12] A. Kaser, S. Zeissig, and R. S. Blumberg, "Genes and environment: how will our concepts on the pathophysiology of IBD develop in the future?" *Digestive Diseases*, vol. 28, no. 3, pp. 395-405, 2010.

[13] D. N. Frank, A. L. St. Amand, R. A. Feldman, E. C. Boedeker, N. Harpaz, and N. R. Pace, "Molecular-phylogenetic characterization of microbial community imbalances in human inflammatory bowel diseases," *Proceedings of the National Academy of Sciences of the United States of America*, vol. 104, no. 34, pp. 13780-13785, 2007.

[14] N. Kaur, C.-C. Chen, J. Luther, and J. Y. Kao, "Intestinal dysbiosis in inflammatory bowel disease," *Gut Microbes*, vol. 2, no. 4, pp. 211-216, 2011.

[15] L. Rigottier-Gois, "Dysbiosis in inflammatory bowel diseases: the oxygen hypothesis," *ISME Journal*, vol. 7, no. 7, pp. 1256-1261, 2013.

[16] P. Seksik, H. Sokol, P. Lepage et al., "Review article: the role of bacteria in onset and perpetuation of inflammatory bowel disease," *Alimentary Pharmacology and Therapeutics*, vol. 24, supplement 3, pp. 11-18, 2006.

[17] D. Gevers, S. Kugathasan, L. A. Denson et al., "The treatment-naïve microbiome in new-onset Crohn's disease," *Cell Host & Microbe*, vol. 15, no. 3, pp. 382-392, 2014.

[18] Q. Li, C. Wang, C. Tang, Q. He, N. Li, and J. Li, "Dysbiosis of gut fungal microbiota is associated with mucosal inflammation in crohn's disease," *Journal of Clinical Gastroenterology*, vol. 48, no. 6, pp. 513-523, 2014.

[19] M. Tong, X. Li, L. W. Parfrey et al., "A modular organization of the human intestinal mucosal microbiota and its association with inflammatory bowel disease," *PLoS ONE*, vol. 8, no. 11, Article ID e80702, 2013.

[20] H. Sokol, B. Pigneur, L. Watterlot et al., "Faecalibacterium prausnitzii is an anti-inflammatory commensal bacterium identified by gut microbiota analysis of Crohn disease patients," *Proceedings of the National Academy of Sciences of the United States of America*, vol. 105, no. 43, pp. 16731-16736, 2008.

[21] T. Fujimoto, H. Imaeda, K. Takahashi et al., "Decreased abundance of Faecalibacterium prausnitzii in the gut microbiota of Crohn's disease," *Journal of Gastroenterology and Hepatology*, vol. 28, no. 4, pp. 613-619, 2013.

[22] K. Machiels, M. Joossens, J. Sabino et al., "A decrease of the butyrate-producing species *Roseburia hominis* and *Faecalibacterium prausnitzii* defines dysbiosis in patients with ulcerative colitis," *Gut*, vol. 63, no. 8, pp. 1275-1283, 2014.

[23] M. Lopez-Siles, M. Martinez-Medina, D. Busquets et al., "Mucosa-associated *Faecalibacterium prausnitzii* and *Escherichia coli* co-abundance can distinguish Irritable Bowel Syndrome and Inflammatory Bowel Disease phenotypes," *International Journal of Medical Microbiology*, vol. 304, no. 3-4, pp. 464-475, 2014.

[24] F. Autschbach, S. Eisold, U. Hinz et al., "High prevalence of *Mycobacterium avium* subspecies *paratuberculosis* IS900 DNA in gut tissues from individuals with Crohn's disease," *Gut*, vol. 54, no. 7, pp. 944-949, 2005.

[25] J. M. Thomson, R. Hansen, S. H. Berry et al., "Enterohepatic *Helicobacter* in ulcerative colitis: potential pathogenic entities?" *PLoS ONE*, vol. 6, no. 2, Article ID e17184, 2011.

[26] V. Mahendran, S. M. Riordan, M. C. Grimm et al., "Prevalence of *Campylobacter* species in adult Crohn's disease and the preferential colonization sites of *Campylobacter* species in the human intestine," *PLoS ONE*, vol. 6, no. 9, Article ID e25417, 2011.

[27] T. Ohkusa, I. Okayasu, T. Ogihara, K. Morita, M. Ogawa, and N. Sato, "Induction of experimental ulcerative colitis by *Fusobacterium varium* isolated from colonic mucosa of patients with ulcerative colitis," *Gut*, vol. 52, no. 1, pp. 79-83, 2003.
[28] I.-A. Lee and D.-H. Kim, "Klebsiella pneumoniae" increases the risk of inflammation and colitis in a murine model of intestinal bowel disease," *Scandinavian Journal of Gastroenterology*, vol. 46, no. 6, pp. 684-693, 2011.

[29] K. O. Gradel, H. L. Nielsen, H. C. Schønheyder, T. Ejlertsen, and B. Kristensen, "Increased short- and long-term risk of inflammatory bowel disease after *Salmonella* or *Campylobacter* gastroenteritis," *Gastroenterology*, vol. 137, no. 2, pp. 495-501, 2009.

[30] A. Saebo, E. Vik, O. J. Lange, and L. Matuszkiewicz, "Inflammatory bowel disease associated with *Yersinia enterocolitica* O:3 infection," *European Journal of Internal Medicine*, vol. 16, no. 3, pp. 176-182, 2005.

[31] S. B. Leu, S. C. Shulman, C. K. Steelman et al., "Pathogenic *yersinia* DNA in intestinal specimens of pediatric patients with Crohn's disease," *Fetal & Pediatric Pathology*, vol. 32, no. 5, pp. 367-370, 2013.

[32] J. M. Rhodes, "The role of *Escherichia coli* in inflammatory bowel disease," *Gut*, vol. 56, no. 5, pp. 610-612, 2007.

[33] N. Rolhion and A. Darfeuille-Michaud, "Adherent-invasive *Escherichia coli* in inflammatory bowel disease," *Inflammatory Bowel Diseases*, vol. 13, no. 10, pp. 1277-1283, 2007.

[34] S. E. Winter, M. G. Winter, M. N. Xavier et al., "Host-derived nitrate boosts growth of *E. coli* in the inflamed gut," *Science*, vol. 339, no. 6120, pp. 708-711, 2013.

[35] S. Tabaqchali, D. P. O'Donoghue, and K. A. Bettelheim, "*Escherichia coli* antibodies in patients with inflammatory bowel disease," *Gut*, vol. 19, no. 2, pp. 108-113, 1978.

[36] M. R. B. Keighley, Y. Arabi, F. Dimock, D. W. Burdon, R. N. Allan, and J. Alexander-Williams, "Influence of inflammatory bowel disease on intestinal microflora," *Gut*, vol. 19, no. 12, pp. 1099-1104, 1978.

[37] Y. Liu, H. J. Van Kruiningen, A. B. West, R. W. Cartun, A. Cortot, and J.-F. Colombel, "Immunocytochemical evidence of *Listeria*, *Escherichia coli*, and *Streptococcus* antigens in Crohn's disease," *Gastroenterology*, vol. 108, no. 5, pp. 1396-1404, 1995.

[38] C. Schultsz, M. Moussa, R. Van Ketel, G. N. J. Tytgat, and J. Dankert, "Frequency of pathogenic and enteroadherent *Escherichia coli* in patients with inflammatory bowel disease and controls," *Journal of Clinical Pathology*, vol. 50, no. 7, pp. 573-579, 1997.

[39] A. Darfeuille-Michaud, C. Neut, N. Barnich et al., "Presence of adherent *Escherichia coli* strains in ileal mucosa of patients with Crohn's disease," *Gastroenterology*, vol. 115, no. 6, pp. 1405-1413, 1998.

[40] E. Masseret, J. Boudeau, J. F. Colombel et al., "Genetically related *Escherichia coli* strains associated with Crohn's disease," *Gut*, vol. 48, no. 3, pp. 320-325, 2001.

[41] H. Fujita, Y. Eishi, I. Ishige et al., "Quantitative analysis of bacterial DNA from *Mycobacteria* spp., *Bacteroides vulgatus*, and *Escherichia coli* in tissue samples from patients with inflammatory bowel diseases," *Journal of Gastroenterology*, vol. 37, no. 7, pp. 509-516, 2002.

[42] P. Ryan, R. G. Kelly, G. Lee et al., "Bacterial DNA within granulomas of patients with Crohn's disease—detection by laser capture microdissection and PCR," *The American Journal of Gastroenterology*, vol. 99, no. 8, pp. 1539-1543, 2004.

[43] H. M. Martin, B. J. Campbell, C. A. Hart et al., "Enhanced *Escherichia coli* adherence and invasion in Crohn's disease and colon cancer," *Gastroenterology*, vol. 127, no. 1, pp. 80-93, 2004.

[44] A. Darfeuille-Michaud, J. Boudeau, P. Bulois et al., "High prevalence of adherent-invasive *Escherichia coli* associated with ileal mucosa in Crohn's disease," *Gastroenterology*, vol. 127, no. 2, pp. 412-421, 2004.

[45] M. Mylonaki, N. B. Rayment, D. S. Rampton, B. N. Hudspith, and J. Brostoff, "Molecular characterization of rectal mucosa-associated bacterial flora in inflammatory bowel disease," *Inflammatory Bowel Diseases*, vol. 11, no. 5, pp. 481-487, 2005.

[46] L. Mei, S. R. Targan, C. J. Landers et al., "Familial expression of anti-*Escherichia coli* outer membrane porin C in relatives of patients with Crohn's disease," *Gastroenterology*, vol. 130, no. 4, pp. 1078-1085, 2006.

[47] R. Kotlowski, C. N. Bernstein, S. Sepehri, and D. O. Krause, "High prevalence of *Escherichia coli* belonging to the B2+D phylogenetic group in inflammatory bowel disease," *Gut*, vol. 56, no. 5, pp. 669-675, 2007.

[48] M. Baumgart, B. Dogan, M. Rishniw et al., "Culture independent analysis of ileal mucosa reveals a selective increase in invasive *Escherichia coli* of novel phylogeny relative to depletion of Clostridiales in Crohn's disease involving the ileum," *The ISME Journal*, vol. 1, no. 5, pp. 403-418, 2007.

[49] M. Sasaki, S. V. Sitaraman, B. A. Babbin et al., "Invasive *Escherichia coli* are a feature of Crohn's disease," *Laboratory Investigation*, vol. 87, no. 10, pp. 1042-1054, 2007.

[50] M. Martinez-Medina, X. Aldeguer, M. Lopez-Siles et al., "Molecular diversity of *Escherichia coli* in the human gut: new ecological evidence supporting the role of adherent-invasive *E. coli* (AIEC) in Crohn's disease," *Inflammatory Bowel Diseases*, vol. 15, no. 6, pp. 872-882, 2009.

[51] B. Willing, J. Halfvarson, J. Dicksved et al., "Twin studies reveal specific imbalances in the mucosa-associated microbiota of patients with ileal Crohn's disease," *Inflammatory Bowel Diseases*, vol. 15, no. 5, pp. 653-660, 2009.

[52] A. M. Petersen, E. M. Nielsen, E. Litrup, J. Brynskov, H. Mirsepassi, and K. A. Krogfelt, "A phylogenetic group of *Escherichia coli* associated with active left-sided inflammatory bowel disease," *BMC Microbiology*, vol. 9, article 171, 2009.

[53] A. Rehman, P. Lepage, A. Nolte, S. Hellmig, S. Schreiber, and S. J. Ott, "Transcriptional activity of the dominant gut mucosal microbiota in chronic inflammatory bowel disease patients," *Journal of Medical Microbiology*, vol. 59, part 9, pp. 1114-1122, 2010.

[54] A. Schwiertz, M. Jacobi, J. S. Frick, M. Richter, K. Rusch, and H. Köhler, "Microbiota in pediatric inflammatory bowel disease," *Journal of Pediatrics*, vol. 157, no. 2, pp. 240.e1-244.e1, 2010.

[55] S. Kang, S. E. Denman, M. Morrison et al., "Dysbiosis of fecal microbiota in Crohn's disease patients as revealed by a custom phylogenetic microarray," *Inflammatory Bowel Diseases*, vol. 16, no. 12, pp. 2034-2042, 2010.

[56] S. Mondot, S. Kang, J. P. Furet et al., "Highlighting new phylogenetic specificities of Crohn's disease microbiota," *Inflammatory Bowel Diseases*, vol. 17, no. 1, pp. 185-192, 2011.

[57] C. M. Thomazini, D. A. G. Samegima, M. A. M. Rodrigues, C. R. Victoria, and J. Rodrigues, "High prevalence of aggregative adherent *Escherichia coli* strains in the mucosa-associated microbiota of patients with inflammatory bowel diseases," *International Journal of Medical Microbiology*, vol. 301, no. 6, pp. 475-479, 2011.

[58] H. L. de Souza, V. R. de Carvalho, F. G. Romeiro, L. Y. Sassaki, R. Keller, and J. Rodrigues, "Mucosa-associated but not luminal *Escherichia coli* is augmented in Crohn's disease and ulcerative colitis," *Gut Pathogens*, vol. 4, article 21, 2012.
[59] B. Dogan, E. Scherl, B. Bosworth et al., "Multidrug resistance is common in *Escherichia coli* associated with ileal Crohn's disease," *Inflammatory Bowel Diseases*, vol. 19, no. 1, pp. 141-150, 2013.

[60] S. Sha, B. Xu, X. Wang et al., "The biodiversity and composition of the dominant fecal microbiota in patients with inflammatory bowel disease," *Diagnostic Microbiology and Infectious Disease*, vol. 75, no. 3, pp. 245-251, 2013.

[61] T. R. Elliott, B. N. Hudspith, G. Wu et al., "Quantification and characterization of mucosa-associated and intracellular *Escherichia coli* in inflammatory bowel disease," *Inflammatory Bowel Diseases*, vol. 19, no. 11, pp. 2326-2338, 2013.

[62] C. L. O'Brien, P. Pavli, D. M. Gordon, and G. E. Allison, "Detection of bacterial DNA in lymph nodes of Crohn's disease patients using high throughput sequencing," *Gut*, vol. 63, no. 10, pp. 1596-1606, 2014.

[63] T. Raso, S. Crivellaro, M. G. Chirillo, P. Pais, E. Gaia, and D. Savoia, "Analysis of escherichia coli isolated from patients affected by Crohn's Disease," *Current Microbiology*, vol. 63, no. 2, pp. 131-137, 2011.

[64] A. Negroni, M. Costanzo, R. Vitali et al., "Characterization of adherent-invasive *Escherichia coli* isolated from pediatric patients with inflammatory bowel disease," *Inflammatory Bowel Diseases*, vol. 18, no. 5, pp. 913-924, 2012.

[65] S. Schippa, V. Iebba, V. Totino et al., "A potential role of *Escherichia coli* pathobionts in the pathogenesis of pediatric inflammatory bowel disease," *Canadian Journal of Microbiology*, vol. 58, no. 4, pp. 426-432, 2012.

[66] H. T. T. Nguyen, G. Dalmasso, S. Müller, J. Carrière, F. Seibold, and A. Darfeuille-Michaud, "Crohn's disease-associated adherent invasive *Escherichia coli* modulate levels of microRNAs in intestinal epithelial cells to reduce autophagy," *Gastroenterology*, vol. 146, no. 2, pp. 508-519, 2014.

[67] J. Boudeau, A.-L. Glasser, E. Masseret, B. Joly, and A. Darfeuille-Michaud, "Invasive ability of an *Escherichia coli* strain isolated from the ileal mucosa of a patient with Crohn's disease," *Infection and Immunity*, vol. 67, no. 9, pp. 4499-4509, 1999.

[68] T. Eaves-Pyles, C. A. Allen, J. Taormina et al., "*Escherichia coli* isolated from a Crohn's disease patient adheres, invades, and induces inflammatory responses in polarized intestinal epithelial cells," *International Journal of Medical Microbiology*, vol. 298, no. 5-6, pp. 397-409, 2008.

[69] A.-L. Glasser, J. Boudeau, N. Barnich, M.-H. Perruchot, J.-F. Colombel, and A. Darfeuille-Michaud, "Adherent invasive *Escherichia coli* strains from patients with Crohn's disease survive and replicate within macrophages without inducing host cell death," *Infection and Immunity*, vol. 69, no. 9, pp. 5529-5537, 2001.

[70] M.-A. Bringer, A.-L. Glasser, C.-H. Tung, S. Méresse, and A. Darfeuille-Michaud, "The Crohn's disease-associated adherent-invasive *Escherichia coli* strain LF82 replicates in mature phagolysosomes within J774 macrophages," *Cellular Microbiology*, vol. 8, no. 3, pp. 471-484, 2006.

[71] S. Meconi, A. Vercellone, F. Levillain et al., "Adherent-invasive *Escherichia coli* isolated from Crohn's disease patients induce granulomas in vitro," *Cellular Microbiology*, vol. 9, no. 5, pp. 1252-1261, 2007.

[72] M. de la Fuente, L. Franchi, D. Araya et al., "*Escherichia coli* isolates from inflammatory bowel diseases patients survive in macrophages and activate NLRP3 inflammasome," *International Journal of Medical Microbiology*, vol. 304, no. 3-4, pp. 384-392, 2014.

[73] K. W. Simpson, B. Dogan, M. Rishniw et al., "Adherent and invasive *Escherichia coli* is associated with granulomatous colitis in Boxer dogs," *Infection and Immunity*, vol. 74, no. 8, pp. 4778-4792, 2006.

[74] N. Barnich, F. A. Carvalho, A.-L. Glasser et al., "CEACAM6 acts as a receptor for adherent-invasive *E. coli*, supporting ileal mucosa colonization in Crohn disease," *Journal of Clinical Investigation*, vol. 117, no. 6, pp. 1566-1574, 2007.

[75] B. Chassaing, N. Rolhion, A. de Vallée et al., "Crohn disease-associated adherent-invasive *E. coli* bacteria target mouse and human Peyer's patches via long polar fimbriae," *Journal of Clinical Investigation*, vol. 121, no. 3, pp. 966-975, 2011.

[76] F. A. Carvalho, N. Barnich, P. Sauvanet, C. Darcha, A. Gelot, and A. Darfeuille-Michaud, "Crohn's disease-associated *Escherichia coli* LF82 aggravates colitis in injured mouse colon via signaling by flagellin," *Inflammatory Bowel Diseases*, vol. 14, no. 8, pp. 1051-1060, 2008.

[77] C. H. F. Chan and C. P. Stanners, "Novel mouse model for carcinoembryonic antigen-based therapy," *Molecular Therapy*, vol. 9, no. 6, pp. 775-785, 2004.

[78] F. A. Carvalho, N. Barnich, A. Sivignon et al., "Crohn's disease adherent-invasive *Escherichia coli* colonize and induce strong gut inflammation in transgenic mice expressing human CEA-CAM," *Journal of Experimental Medicine*, vol. 206, no. 10, pp. 2179-2189, 2009.

[79] C.-L. N. Small, S. A. Reid-Yu, J. B. McPhee, and B. K. Coombes, "Persistent infection with Crohn's disease-associated adherent-invasive *Escherichia coli* leads to chronic inflammation and intestinal fibrosis," *Nature Communications*, vol. 4, p. 1957, 2013.

[80] J. B. McPhee, C. L. Small, S. A. Reid-Yu, J. R. Brannon, H. Le Moual, and B. K. Coombes, "Host defense peptide resistance contributes to colonization and maximal intestinal pathology by Crohn's disease-associated adherent-invasive *Escherichia coli*," *Infection and Immunity*, vol. 82, no. 8, pp. 3383-3393, 2014.

[81] M. Martinez-Medina, J. Denizot, N. Dreux et al., "Western diet induces dysbiosis with increased *E. coli* in CEABAC10 mice, alters host barrier function favouring AIEC colonisation," *Gut*, vol. 63, no. 1, pp. 116-124, 2014.

[82] B. Chassaing, O. Koren, F. A. Carvalho, R. E. Ley, and A. T. Gewirtz, "AIEC pathobiont instigates chronic colitis in susceptible hosts by altering microbiota composition," *Gut*, vol. 63, no. 7, pp. 1069-1080, 2014.

[83] M. Craven, C. E. Egan, S. E. Dowd et al., "Inflammation drives dysbiosis and bacterial invasion in murine models of ileal Crohn's Disease," *PLoS ONE*, vol. 7, no. 7, Article ID e41594, 2012.

[84] J. L. Round and S. K. Mazmanian, "The gut microbiota shapes intestinal immune responses during health and disease," *Nature Reviews Immunology*, vol. 9, no. 5, pp. 313-323, 2009.

[85] D. Low, H. T. Tran, I.-A. Lee et al., "Chitin-binding domains of *Escherichia coli* ChiA mediate interactions with intestinal epithelial cells in mice with colitis," *Gastroenterology*, vol. 145, no. 3, pp. 602.e9-612.e9, 2013.

[86] N. Rolhion, N. Barnich, M.-A. Bringer et al., "Abnormally expressed ER stress response chaperone Gp96 in CD favours adherent-invasive *Escherichia coli* invasion," *Gut*, vol. 59, no. 10, pp. 1355-1362, 2010.

[87] A. D. Garg, A. Kaczmarek, O. Krysko, P. Vandenabeele, D. V. Krysko, and P. Agostinis, "ER stress-induced inflammation: does it aid or impede disease progression?" *Trends in Molecular Medicine*, vol. 18, no. 10, pp. 589-598, 2012.

[88] K. A. Dunne, A. Allam, A. McIntosh et al., "Increased S-nitrosylation and proteasomal degradation of caspase-3 during
infection contribute to the persistence of adherent invasive *Escherichia coli* (AIEC) in immune cells,” *PLoS ONE*, vol. 8, no. 7, Article ID e68386, 2013.

[89] I. Cleynen, E. Vazeille, M. Artieda et al., “Genetic and microbial factors modulating the ubiquitin proteasome system in inflammatory bowel disease,” *Gut*, vol. 63, no. 8, pp. 1265–1274, 2013.

[90] P. Lapaquette, M.-A. Bringer, and A. Darfeuille-Michaud, “Defects in autophagy favour adherent-invasive *Escherichia coli* persistence within macrophages leading to increased pro-inflammatory response,” *Cellular Microbiology*, vol. 14, no. 6, pp. 791–807, 2012.

[91] A. Chargui, A. Cesaro, S. Mimouna et al., “Subversion of autophagy in adherent invasive *Escherichia coli*-infected neutrophils induces inflammation and cell death,” *PLoS ONE*, vol. 7, no. 12, Article ID e51727, 2012.

[92] C. Lu, J. Chen, H.-G. Xu et al., “MIR106B and MIR93 prevent removal of bacteria from epithelial cells by disrupting ATG16L1-mediated autophagy,” *Gastroenterology*, vol. 146, no. 1, pp. 188–199, 2014.

[93] K. P. Nickerson and C. McDonald, “Crohn’s disease-associated adherent-invasive *Escherichia coli* adhesion is enhanced by exposure to the ubiquitous dietary polysaccharide maltodextrin,” *PLoS ONE*, vol. 7, no. 12, Article ID e52132, 2012.

[94] C. D. Packey and R. B. Sartor, “Commensal bacteria, traditional and opportunistic pathogens, dysbiosis and bacterial killing in inflammatory bowel diseases,” *Current Opinion in Infectious Diseases*, vol. 22, no. 3, pp. 292–301, 2009.

[95] S. Miquel, E. Peyretaillade, L. Claret et al., “Complete genome sequence of crohn’s disease-associated adherent-invasive *E. coli* strain LF82,” *PLoS ONE*, vol. 5, no. 9, Article ID e12714, pp. 1–16, 2010.

[96] J. H. E. Nash, A. Villegas, A. M. Kropinski et al., “Genome sequence of adherent-invasive *Escherichia coli* and comparative genomic analysis with other *E. coli* pathotypes,” *BMC Genomics*, vol. 11, article no. 667, 2010.

[97] N. Dreux, J. Denizot, M. Martinez-Medina et al., “Point mutations in FimH adhesin of Crohn’s disease-associated adherent-invasive *Escherichia coli* enhance intestinal inflammatory response,” *PLoS Pathogens*, vol. 9, no. 1, Article ID e1003141, 2013.

[98] E. Vazeille, M.-A. Bringer, A. Gardarin et al., “Role of meprins to protect ileal Mucosa of Crohn’s disease patients from colonization by adherent-invasive *E. coli*,” *PLoS ONE*, vol. 6, no. 6, Article ID e21199, 2011.

[99] L. J. John, M. Fromm, and J.-D. Schulzke, “Epithelial barriers in intestinal inflammation,” *Antioxidants and Redox Signaling*, vol. 15, no. 5, pp. 1255–1270, 2011.

[100] A. V. Keita and J. D. Söderholm, “Barrier dysfunction and bacterial uptake in the follicle-associated epithelium of ileal Crohn’s disease,” *Annals of the New York Academy of Sciences*, vol. 1258, no. 1, pp. 125–134, 2012.

[101] L. Antoni, S. Nuding, J. Wehkamp, and E. F. Stange, “Intestinal barrier in inflammatory bowel disease,” *World Journal of Gastroenterology*, vol. 20, no. 5, pp. 1165–1179, 2014.

[102] J. Benjamin, G. K. Makharia, V. Ahuja, M. Kalaivani, and Y. K. Joshi, “Intestinal permeability and its association with the patient and disease characteristics in Crohn’s disease,” *World Journal of Gastroenterology*, vol. 14, no. 9, pp. 1399–1405, 2008.

[103] E. Wine, J. C. Ossa, S. D. Gray-Owen, and P. M. Sherman, “Adherent-invasive *Escherichia coli*, strain LF82 disrupts apical junctional complexes in polarized epithelia,” *BMC Microbiology*, vol. 9, article 180, 2009.

[104] L. C.-H. Yu, J.-T. Wang, S.-C. Wei, and Y.-H. Ni, “Host-microbial interactions and regulation of intestinal epithelial barrier function: from physiology to pathology,” *World Journal of Gastrointestinal Pathophysiology*, vol. 3, no. 1, pp. 27–43, 2012.

[105] M. E. V. Johansson, D. Ambort, T. Pelaseyed et al., “Composition and functional role of the mucus layers in the intestine,” *Cellular and Molecular Life Sciences*, vol. 68, no. 22, pp. 3635–3641, 2011.

[106] M. A. McGuckin, S. K. Lindén, P. Sutton, and T. H. Florin, “Mucin dynamics and enteric pathogens,” *Nature Reviews Microbiology*, vol. 9, no. 4, pp. 265–278, 2011.

[107] C. W. Png, S. K. Lindén, K. S. Gilshenan et al., “Mucolytic bacteria with increased prevalence in IBD mucosa augment in vitro utilization of mucin by other bacteria,” *The American Journal of Gastroenterology*, vol. 105, no. 11, pp. 2420–2428, 2010.

[108] J. A. Guttman and B. B. Finlay, “Tight junctions as targets of infectious agents,” *Biochimica et Biophysica Acta*, vol. 1788, no. 4, pp. 832–841, 2009.

[109] S. Zeissig, N. Bürgel, D. Günzel et al., “Changes in expression and distribution of claudin 2, 5 and 8 lead to discontinuous tight junctions and barrier dysfunction in active Crohn’s disease,” *Gut*, vol. 56, no. 1, pp. 61–72, 2007.

[110] J. Denizot, A. Sivignon, F. Barreau et al., “Adherent-invasive *Escherichia coli* induce claudin-2 expression and barrier defect in CEABAC10 mice and Crohn’s disease patients,” *Inflammatory Bowel Diseases*, vol. 18, no. 2, pp. 294–304, 2012.

[111] B. K. Reuter and T. T. Pizarro, “Mechanisms of tight junction dysregulation in the SAMP1YitFc model of Crohn’s disease-like ileitis,” *Annals of the New York Academy of Sciences*, vol. 1165, pp. 301–307, 2009.

[112] T. S. Olson, B. K. Reuter, K. G.-E. Scott et al., “The primary defect in experimental ileitis originates from a nonhematopoietic source,” *The Journal of Experimental Medicine*, vol. 203, no. 3, pp. 541–552, 2006.

[113] C. Jung, J.-P. Hugot, and F. Barreau, “Peyers patches: the immune sensors of the intestine,” *International Journal of Inflammation*, vol. 2010, Article ID 823710, 12 pages, 2010.

[114] A. Marra and R. R. Isberg, “Invasin-dependent and invasin-independent pathways for translocation of *Yersinia pseudotuberculosis* across the Peyer’s patch intestinal epithelium,” *Infection and Immunity*, vol. 65, no. 8, pp. 3412–3421, 1997.

[115] R. P. Martins, V. Lorenzi, C. Arce, C. Lucena, A. Carvajal, and J. J. Garrido, “Innate and adaptive immune mechanisms are effectively induced in ileal Peyer’s patches of *Salmonella typhimurium* infected pigs,” *Developmental & Comparative Immunology*, vol. 41, no. 1, pp. 100–104, 2013.

[116] H. Lelouard, M. Fallet, B. de Bovis, S. Méresse, and J. Gorvel, “Peyer’s patch dendritic cells sample antigens by extending dendrites through M cell-specific transcellular pores,” *Gastroenterology*, vol. 142, no. 3, pp. 592.e3–601.e3, 2012.

[117] E. Gullberg and J. D. Söderholm, “Peyer’s patches and M cells as potential sites of the inflammatory onset in Crohn’s disease,” *Annals of the New York Academy of Sciences*, vol. 1072, pp. 218–232, 2006.

[118] N. Kamada, S.-U. Seo, G. Y. Chen, and G. Núñez, “Role of the gut microbiota in immunity and inflammatory disease,” *Nature Reviews Immunology*, vol. 13, no. 5, pp. 321–335, 2013.

[119] G. F. Sonnenberg and D. Artis, “Innate lymphoid cell interactions with microbiota: implications for intestinal health and disease,” *Immunity*, vol. 37, no. 4, pp. 601–610, 2012.

[120] M. J. Molloy, N. Bouladoux, and Y. Belkaid, “Intestinal microbiota: shaping local and systemic immune responses,” *Seminars in Immunology*, vol. 24, no. 1, pp. 58–66, 2012.
[121] A. Cebula, M. Seweryn, G. A. Rempala et al., "Thymus-derived regulatory T cells contribute to tolerance to commensal microbiota," *Nature*, vol. 497, no. 7448, pp. 258-262, 2013.

[122] M. R. Hepworth, L. A. Monticelli, T. C. Fung et al., "Innate lymphoid cells regulate CD4⁺ T-cell responses to intestinal commensal bacteria," *Nature*, vol. 498, no. 7452, pp. 113-117, 2013.

[123] L. V. Hooper and A. J. MacPherson, "Immune adaptations that maintain homeostasis with the intestinal microbiota," *Nature Reviews Immunology*, vol. 10, no. 3, pp. 159-169, 2010.

[124] A. Konrad, Y. Cong, W. Duck, R. Borlaza, and C. O. Elson, "Tight mucosal compartmentation of the murine immune response to antigens of the enteric microbiota," *Gastroenterology*, vol. 130, no. 7, pp. 2050-2059, 2006.

[125] E. Mazzini, L. Massimiliano, G. Penna, and M. Rescigno, "Oral tolerance can be established via gap junction transfer of fed antigens from CX3CR1⁺ macrophages to CD103⁺ dendritic cells," *Immunity*, vol. 40, no. 2, pp. 248-261, 2014.

[126] R. Marques and I. G. Boneca, "Expression and functional importance of innate immune receptors by intestinal epithelial cells," *Cellular and Molecular Life Sciences*, vol. 68, no. 22, pp. 3661-3673, 2011.

[127] T. W. Hand, L. M. Dos Santos, N. Bouladoux et al., "Acute gastrointestinal infection induces long-lived microbiota-specific T cell responses," *Science*, vol. 337, no. 6101, pp. 1553-1556, 2012.

[128] A. Haas, K. Zimmermann, F. Graw et al., "Systemic antibody responses to gut commensal bacteria during chronic HIV-1 infection," *Gut*, vol. 60, no. 11, pp. 1506-1519, 2011.

[129] C. Pearson, H. H. Uhlig, and F. Powrie, "Lymphoid microenvironment and innate lymphoid cells in the gut," *Trends in Immunology*, vol. 33, no. 6, pp. 289-296, 2012.

[130] K.atarashi, T. Tanoue, T. Shima et al., "Induction of colonic regulatory T cells by indigenous *Clostridium* species," *Science*, vol. 331, no. 6015, pp. 337-341, 2011.

[131] K.atarashi, T. Tanoue, K. Oshima et al., "Treg induction by a rationally selected mixture of *Clostridia* strains from the human microbiota," *Nature*, vol. 500, no. 7461, pp. 232-236, 2013.

[132] M. Livingston, D. Loach, M. Wilson, G. W. Tannock, and M. Baird, "Gut commensal *Lactobacillus reuteri* 100-23 stimulates an immunoregulatory response," *Immunology and Cell Biology*, vol. 88, no. 1, pp. 99-102, 2010.

[133] K. M. Maslowski, A. T. Vieira, A. Ng et al., "Regulation of inflammatory responses by gut microbiota and chemoattractant receptor GPR43," *Nature*, vol. 461, no. 7268, pp. 1282-1286, 2009.

[134] Y. Furusawa, Y. Obata, S. Fukuda et al., "Commensal microbiota-derived butyrate induces the differentiation of colonic regulatory T cells," *Nature*, vol. 504, no. 7480, pp. 446-450, 2013.

[135] P. M. Smith, M. R. Howitt, N. Panikov et al., "The microbial metabolites, short-chain fatty acids, regulate colonic T reg cell homeostasis," *Science*, vol. 341, no. 6145, pp. 569-573, 2013.

[136] J. Ochoa-Repáraz, D. W. Mielcarz, Y. Wang et al., "A polysaccharide from the human commensal *Bacteroides fragilis* protects against CNS demyelinating disease," *Mucosal Immunology*, vol. 3, no. 5, pp. 487-495, 2010.

[137] J. L. Round and S. K. Mazmanian, "Inducible Foxp3⁺ regulatory T-cell development by a commensal bacterium of the intestinal microbiota," *Proceedings of the National Academy of Sciences of the United States of America*, vol. 107, no. 27, pp. 12204-12209, 2010.

[138] I. I. Ivanov, K. Atarashi, N. Manel et al., "Induction of intestinal Th17 cells by segmented filamentous bacteria," *Cell*, vol. 139, no. 3, pp. 485-498, 2009.

[139] V. Gaboriau-Routhiau, S. Rakotobe, E. Lécuyer et al., "The key role of segmented filamentous bacteria in the coordinated maturation of gut helper T cell responses," *Immunity*, vol. 31, no. 4, pp. 677-689, 2009.

[140] J. A. Hall, N. Bouladoux, C. M. Sun et al., "Commensal DNA limits regulatory T cell conversion and is a natural adjuvant of intestinal immune responses," *Immunity*, vol. 29, no. 4, pp. 637-649, 2008.

[141] H.-J. Wu, I. I. Ivanov, J. Darce et al., "Gut-residing segmented filamentous bacteria drive autoimmune arthritis via T helper 17 cells," *Immunity*, vol. 32, no. 6, pp. 815-827, 2010.

[142] S. Wu, K.-J. Rhee, E. Albesiano et al., "A human colonic commensal promotes colon tumorigenesis via activation of T helper type 17 T cell responses," *Nature Medicine*, vol. 15, no. 9, pp. 1016-1022, 2009.

[143] M. Lochner, C. Ohnmacht, L. Presley et al., "Microbiota-induced tertiary lymphoid tissues aggravate inflammatory disease in the absence of RORγt and LTI cells," *The Journal of Experimental Medicine*, vol. 208, no. 1, pp. 125-134, 2011.

[144] M. J. McGeachy, K. S. Bak-Jensen, Y. Chen et al., "TGF-β and IL-6 drive the production of IL-17 and IL-10 by T cells and restrain TH-17 cell-mediated pathology," *Nature Immunology*, vol. 8, no. 12, pp. 1390-1397, 2007.

[145] S. Hapfelmeier, M. A. E. Lawson, E. Slack et al., "Reversible microbial colonization of germ-free mice reveals the dynamics of IgA immune responses," *Science*, vol. 328, no. 5986, pp. 1705-1709, 2010.

[146] W. S. Garrett, G. M. Lord, S. Punit et al., "Communicable ulcerative colitis induced by T-bet deficiency in the innate immune system," *Cell*, vol. 131, no. 1, pp. 33-45, 2007.

[147] S. Saha, X. Jing, S. Y. Park et al., "Peptidoglycan recognition proteins protect mice from experimental colitis by promoting normal gut flora and preventing induction of interferon-γ," *Cell Host and Microbe*, vol. 8, no. 2, pp. 147-162, 2010.

[148] A. Couturier-Maillard, T. Secher, A. Rehman et al., "NOD2-mediated dysbiosis predisposes mice to transmissible colitis and colorectal cancer," *The Journal of Clinical Investigation*, vol. 123, no. 2, pp. 700-711, 2013.

[149] L. Jostins, S. Ripke, R. K. Weersma et al., "Host-microbe interactions have shaped the genetic architecture of inflammatory bowel disease," *Nature*, vol. 491, no. 7422, pp. 119-124, 2012.

[150] S. Brand, "Crohn's disease: Th1, Th17 or both? The change of a paradigm: new immunological and genetic insights implicate Th17 cells in the pathogenesis of Crohn's disease," *Gut*, vol. 58, no. 8, pp. 1152-1167, 2009.

[151] C. Abraham and J. H. Cho, "Inflammatory bowel disease," *The New England Journal of Medicine*, vol. 361, no. 21, pp. 2066-2078, 2009.

[152] K. L. Wallace, L.-B. Zheng, Y. Kanazawa, and D. Q. Shih, "Immunopathology of inflammatory bowel disease," *World Journal of Gastroenterology*, vol. 20, no. 1, pp. 6-21, 2014.

[153] R. H. Duerr, K. D. Taylor, S. R. Brant et al., "A genome-wide association study identifies IL23R as an inflammatory bowel disease gene," *Science*, vol. 314, no. 5804, pp. 1461-1463, 2006.

[154] A. Raza, W. Yousaf, R. Giannella, and M. T. Shata, "Th17 cells: interactions with predisposing factors in the immunopathogenesis of inflammatory bowel disease," *Expert Review of Clinical Immunology*, vol. 8, no. 2, pp. 161-168, 2012.

[155] S. Buonocore, P. P. Ahern, H. H. Uhlig et al., "Innate lymphoid cells drive interleukin-23-dependent innate intestinal pathology," *Nature*, vol. 464, no. 7293, pp. 1371-1375, 2010.
[156] A. Geremia, C. V. Arancibia-Cárcamo, M. P. P. Fleming et al., "IL-23-responsive innate lymphoid cells are increased in inflammatory bowel disease," *Journal of Experimental Medicine*, vol. 208, no. 6, pp. 1127-1133, 2011.

[157] F. Annunziato, L. Cosmi, V. Santarlasci et al., "Phenotypic and functional features of human Th17 cells," *The Journal of Experimental Medicine*, vol. 204, no. 8, pp. 1849-1861, 2007.

[158] A. A. Elkadri, J. M. Stempak, T. D. Walters et al., "Serum antibodies associated with complex inflammatory bowel disease," *Inflammatory Bowel Diseases*, vol. 19, no. 7, pp. 1499-1505, 2013.

[159] F. D. M. van Schaik, B. Oldenburg, A. R. Hart et al., "Serological markers predict inflammatory bowel disease years before the diagnosis," *Gut*, vol. 62, no. 5, pp. 683-688, 2013.

[160] M. Papp, I. Altorjay, G. L. Norman et al., "Seroreactivity to microbial components in Crohn's disease is associated with ileal involvement, noninflammatory disease behavior and NOD2/CARD15 genotype, but not with risk for surgery in a Hungarian cohort of IBD patients," *Inflammatory Bowel Diseases*, vol. 13, no. 8, pp. 984-992, 2007.

[161] M. Camus, S. Esses, B. Pariente et al., "Oligoclonal expansions of mucosal T cells in Crohn's disease predominate in NKG2D-expressing CD4 T cells," *Mucosal Immunology*, vol. 7, no. 2, pp. 325-334, 2014.

[162] S. Danese, "New therapies for inflammatory bowel disease: from the bench to the bedside," *Gut*, vol. 61, no. 6, pp. 918-932, 2012.

[163] J. Torres, S. Danese, and J.-F. Colombel, "New therapeutic avenues in ulcerative colitis: thinking out of the box," *Gut*, vol. 62, no. 11, pp. 1642-1652, 2013.

[164] M. F. Neurath, "New targets for mucosal healing and therapy in inflammatory bowel diseases," *Mucosal Immunology*, vol. 7, no. 1, pp. 6-19, 2014.

[165] R. N. Fedorak, A. Gangl, C. O. Elson et al., "Recombinant human interleukin 10 in the treatment of patients with mild to moderately active Crohn's disease," *Gastroenterology*, vol. 119, no. 6, pp. 1473-1482, 2000.

[166] J. D. Lewis, G. R. Lichtenstein, J. J. Deren et al., "Rosiglitazone for active ulcerative colitis: a randomized placebo-controlled trial," *Gastroenterology*, vol. 134, no. 3, pp. 688-695, 2008.

[167] J. Motta, L. Magne, D. Descamps et al., "Modifying the protease, antiprotease pattern by Elafin overexpression protects mice from colitis," *Gastroenterology*, vol. 140, no. 4, pp. 1272-1282, 2011.

[168] H. Braat, P. Rottiers, D. W. Hommes et al., "A phase I trial with transgenic bacteria expressing interleukin-10 in Crohn's disease," *Clinical Gastroenterology and Hepatology*, vol. 4, no. 6, pp. 754-759, 2006.

[169] J.-P. Motta, L. G. Bermúdez-Humarán, C. Deraison et al., "Food-grade bacteria expressing elafin protect against inflammation and restore colon homeostasis," *Science Translational Medicine*, vol. 4, no. 158, Article ID 158ra144, 2012.

[170] W. J. Sandborn, J.-F. Colombel, B. E. Sands et al., "Abatacept for Crohn's disease and ulcerative colitis," *Gastroenterology*, vol. 143, no. 1, pp. 62-e4-69-e4, 2012.

[171] W. J. Sandborn, B. G. Feagan, P. Rutgeerts et al., "Vedolizumab as induction and maintenance therapy for Crohn's disease," *The New England Journal of Medicine*, vol. 369, no. 8, pp. 711-721, 2013.

[172] P. Desreumaux, A. Foussat, M. Allez et al., "Safety and efficacy of antigen-specific regulatory T-cell therapy for patients with refractory Crohn's disease," *Gastroenterology*, vol. 143, no. 5, pp. 1207-1217.e2, 2012.

[173] R. K. Burt, R. M. Craig, F. Milanetti et al., "Autologous nonmyeloablative hematopoietic stem cell transplantation in patients with severe anti-TNF refractory Crohn disease: long-term follow-up," *Blood*, vol. 116, no. 26, pp. 6123-6132, 2010.

[174] M. Clerici, A. Cassinotti, F. Onida et al., "Immunomodulatory effects of unselected haematopoietic stem cells autotransplantation in refractory Crohn's disease," *Digestive and Liver Disease*, vol. 43, no. 12, pp. 946-952, 2011.

[175] A. Cassinotti, C. Annaloro, S. Ardizzone et al., "Autologous haematopoietic stem cell transplantation without CD34 + cell selection in refractory Crohn's disease," *Gut*, vol. 57, no. 2, pp. 211-217, 2008.

[176] M. L. Scribano and C. Prantera, "Use of antibiotics in the treatment of Crohn's disease," *World Journal of Gastroenterology*, vol. 19, no. 5, pp. 648-653, 2013.

[177] K. J. Khan, T. A. Ullman, A. C. Ford et al., "Antibiotic therapy in inflammatory bowel disease: a systematic review and meta-analysis," *The American Journal of Gastroenterology*, vol. 106, no. 4, pp. 661-673, 2011.

[178] C. Prantera, H. Lochs, M. Grimaldi, S. Danese, M. L. Scribano, and P. Gionchetti, "Rifaximin-extended intestinal release induces remission in patients with moderately active crohn's disease," *Gastroenterology*, vol. 142, no. 3, pp. 473.e4-481.e4, 2012.

[179] G. R. D'Haens, S. Vermeire, G. van Assche et al., "Therapy of metronidazole with azathioprine to prevent postoperative recurrence of Crohn's disease: a controlled randomized trial," *Gastroenterology*, vol. 135, no. 4, pp. 1123-1129, 2008.

[180] M. Mañosa, E. Cabré, I. Bernal et al., "Addition of metronidazole to azathioprine for the prevention of postoperative recurrence of Crohn's disease: a randomized, double-blind, placebo-controlled trial," *Inflammatory Bowel Diseases*, vol. 19, no. 9, pp. 1889-1895, 2013.

[181] L. J. Brandt, O. C. Aroniadis, M. Mellow et al., "Long-term follow-up of colonoscopic fecal microbiota transplant for recurrent *Clostridium difficile* infection," *American Journal of Gastroenterology*, vol. 107, no. 7, pp. 1079-1087, 2012.

[182] C. R. Kelly, L. de Leon, and N. Jasutkar, "Fecal microbiota transplantation for relapsing clostridium difficile infection in 26 patients: methodology and results," *Journal of Clinical Gastroenterology*, vol. 46, no. 2, pp. 145-149, 2012.

[183] J. L. Anderson, R. J. Edney, and K. Whelan, "Systematic review: faecal microbiota transplantation in the management of inflammatory bowel disease," *Alimentary Pharmacology and Therapeutics*, vol. 36, no. 6, pp. 503-516, 2012.

[184] H. Gordon and M. Harbord, "A patient with severe Crohn's colitis responds to Faecal Microbiota Transplantation," *Journal of Crohn's and Colitis*, vol. 8, no. 3, pp. 256-257, 2014.

[185] B. Cui, Q. Feng, H. Wang et al., "Fecal microbiota transplantation through mid-gut for refractory Crohn's disease: safety, feasibility and efficacy trial results," *Journal of Gastroenterology and Hepatology*, 2014.

[186] K. Tsilingiri and M. Rescigno, "Postbiotics: what else?" *Beneficial Microbes*, vol. 4, no. 1, pp. 101-107, 2013.

[187] P. Mallon, D. McKay, S. Kirk, and K. Gardiner, "Probiotics for induction of remission in ulcerative colitis," *Cochrane Database of Systematic Reviews*, no. 4, Article ID CD005573, 2007.

[188] L. X. Sang, B. Chang, W. L. Zhang, X. M. Wu, X. H. Li, and M. Jiang, "Remission induction and maintenance effect of probiotics on ulcerative colitis: a meta-analysis," *World Journal of Gastroenterology*, vol. 16, no. 15, pp. 1908-1915, 2010.
[189] G. A. Doherty, G. C. Bennett, A. S. Cheifetz, and A. C. Moss, "Meta-analysis: targeting the intestinal microbiota in prophylaxis for post-operative Crohn's disease," *Alimentary Pharmacology and Therapeutics*, vol. 31, no. 8, pp. 802-809, 2010.

[190] A. Sivignon, A. de Vallée, N. Barnich et al., "Saccharomyces cerevisiae CNCM I-3856 prevents colitis induces by AIEC bacteria in transgenic mouse model mimicking Crohn's disease," *Inflammatory Bowel Diseases*, In Press.

[191] R. Orel and T. Kamhi Trop, "Intestinal microbiota, probiotics and prebiotics in inflammatory bowel disease," *World Journal of Gastroenterology*, vol. 20, no. 33, pp. 11505-11524, 2014.

[192] K. Tsilingiri, T. Barbosa, G. Penna et al., "Probiotic and postbiotic activity in health and disease: comparison on a novel polarised ex-vivo organ culture model," *Gut*, vol. 61, no. 7, pp. 1007-1015, 2012.

[193] M.-A. von Schillde, G. Hörmannsperger, M. Weiher et al., "Lactocepin secreted by *Lactobacillus* exerts anti-inflammatory effects by selectively degrading proinflammatory chemokines," *Cell Host and Microbe*, vol. 11, no. 4, pp. 387-396, 2012.

[194] J. Wagner, J. Maksimovic, G. Farries et al., "Bacteriophages in gut samples from pediatric Crohn's disease patients: metagenomic analysis using 454 pyrosequencing," *Inflammatory Bowel Diseases*, vol. 19, no. 8, pp. 1598-1608, 2013.

[195] A. Reyes, M. Wu, N. P. McNulty, F. L. Rohwer, and J. I. Gordon, "Gnotobiotic mouse model of phage-bacterial host dynamics in the human gut," *Proceedings of the National Academy of Sciences of the United States of America*, vol. 110, no. 50, pp. 20236-20241, 2013.

[196] D. Berry and W. Reinisch, "Intestinal microbiota: a source of novel biomarkers in inflammatory bowel diseases?" *Best Practice and Research: Clinical Gastroenterology*, vol. 27, no. 1, pp. 47-58, 2013.
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