Role of the microbiota in inflammatory bowel diseases


  • Nabeetha A. Nagalingam PhD,

    1. Colitis and Crohn's Disease Microbiome Research Core, Division of Gastroenterology, University of California, San Francisco, Calfornia
    Search for more papers by this author
  • Susan V. Lynch PhD

    Corresponding author
    1. Colitis and Crohn's Disease Microbiome Research Core, Division of Gastroenterology, University of California, San Francisco, Calfornia
    • Colitis and Crohn's Disease Microbiome Research Core, Division of Gastroenterology, University of California, San Francisco, 513 Parnassus Ave., Med Sci S-357, San Francisco, CA 94143-0538
    Search for more papers by this author


Studying the role of the human microbiome as it relates to human health status has revolutionized our view of microbial community contributions to a large number of diseases, particularly chronic inflammatory disorders. The lower gastrointestinal (GI) tract houses trillions of microbial cells representing a large diversity of species in relatively well-defined phylogenetic ratios that are associated with maintenance of key aspects of host physiology and immune homeostasis. It is not surprising, therefore, that many GI inflammatory diseases, including inflammatory bowel disease (IBD), are associated with substantial changes in the composition of these microbial assemblages, either as a cause or consequence of host inflammatory response. Here we review current knowledge in the emerging field of human microbiome research as it relates to IBD, specifically focusing on Crohn's disease (CD) and ulcerative colitis (UC). We discuss bacteriotherapeutic efforts to restore GI microbial assemblage integrity via probiotic supplementation of IBD patients, and speculate on future directions for the field. (Inflamm Bowel Dis 2011;)

“True friendship is like sound health, the value of it is seldom known until it is lost”

—Charles Caleb Colton (1780–1832)

Although the sentiments expressed by Colton are now centuries old, they are perhaps even more relevant today in the context of the gastrointestinal (GI) microbiota and inflammatory bowel disease (IBD). Encompassing a spectrum of diseases characterized by chronic GI inflammation, IBD is typified by disintegration of the partnership between host and microbial community, including a significant loss of bacterial diversity. The outcome is an approximate 1.4 million, clinician-diagnosed, IBD patients in the U.S. alone, resulting in an annual healthcare expenditure in excess of 1.7 billion dollars, primarily for hospital visits and clinical disease management.1, 2

The two most common IBD patient populations are those with Crohn's disease (CD) and ulcerative colitis (UC). The former is characterized by a “cobblestone-like” pattern of inflammation, with affected regions interrupted by healthy tissue that can occur anywhere along the length of the GI tract (mouth to anus). It is also typified by transmural ulcerations resulting in fissures that may perforate the intestinal wall and impact other organs such as the kidney or uterus. The latter, UC, typically manifests as contiguous inflammation involving only the superficial mucosal and submucosal layers. It is primarily localized in the colon and most commonly originates at the rectum.3

Although the etiology of IBD remains unclear, several factors are believed to play a role in its development and progression, including host genotype, immune disequilibrium, and the composition of microbial communities resident in the GI tract.1 Here we focus primarily on the role of the last component, the GI microbiome, as we review evidence for its role in these diseases and provide a discussion of recent data in the larger system-based context of the human host.1

Figure 1.

Some of the known contributions of intestinal microbiota to host health. Changes in the microbial community have been linked to alterations in host physiology and immune responses that affect both local and extraintestinal sites.


Multiple studies have demonstrated an association between microbiota composition and various aspects of host health, including physiological development, metabolism, and immunological response.4–7 Appropriate development of the immune response is dependent on GI colonization. For example, sterilely derived rabbits, hand raised in isolation from conventional animals, and thus acquiring a distinct GI microbiota, exhibit underdeveloped GALT and a substantial decrease in antibody repertoire diversification.8 It has also been demonstrated that gnotobiotic mice, devoid of an appropriate diversity of microbial colonizers, exhibit aberrant immune development.9 Clearly, early development of the mammalian system is highly dependent on microbial colonization; however, even in established (adult) mammalian systems, microbial manipulation of the immune response persists. In several cases, it has been demonstrated that specific GI bacterial species activate host immune responses to facilitate their own survival and competitive fitness.6, 10 For example, Bacteriodetes thetaiotaomicron induces host production of defensins, presumably to eliminate potential competing organisms (including pathogens) and reduce competition for available space; however, in doing so it also plays a key role in regulation of postnatal angiogenesis.10 Other species such as the GI pathogen Salmonella enteritica Typhimurium induce host inflammation to improve their fitness. This species has recently been shown to proliferate under conditions of GI inflammation due to its ability to use the unusual terminal electron acceptor, tetrathionate, generated upon the interaction of reactive oxygen species and luminal thiosulfate.11 Evidently, there is a critical need for the human host to strike a delicate balance between recognizing pathogenic species and accommodating commensal species that modulate a variety of physiological and immunological aspects critical to maintenance of host health. When this alliance is broken, due to immune and/or microbiota disequilibrium, disease, particularly chronic inflammatory diseases, ensue. However, to understand the role of the microbiota in disease development it is important to first appreciate its role in promoting host health.


There is a wealth of information on the benefits of intestinal microbiota on host development. A review by Wostmann,12 published in the early 1980s before the advent of high-resolution culture-independent microbiome profiling tools, summarized a multitude of studies performed describing physiological differences observed between germfree and conventionally raised animals. These included increased ceca size, decreased intestinal weight, and colon surface area in germfree mice compared to their conventionally raised counterparts.13, 14 With the advent of microbial profiling, hypotheses have been put forward to explain these phenomena, including the absence of butyrate-producing bacterial species. Since this short chain fatty acid is the sole energy source for colonocytes,15 its absence putatively leads to poor cellular development in germfree animals. Indeed, germfree mice also exhibit reduced fat deposition indicating that the GI microbiota represent an important facet of lipid metabolism,16 an observation that has been borne out in microbiota-based human obesity studies.17–19

An independent study also noted physiological differences in germfree and conventional mice, specifically, the latter animals exhibited shorter crypt lengths and increased numbers of mucin-secreting goblet cells.20 Mucin plays a critical barrier function in the lower GI tract and these observations suggest that members of the resident microbiota stimulate production of this protective polysaccharide, thus enhancing GI epithelial protection or, conversely, in their absence, depleted mucin coverage and susceptibility to the harmful effects of pathogenic effector molecules or toxic metabolic byproducts that damage the epithelial layer. Interestingly, germfree mice have also been shown to exhibit leaky tight junctions when compared to mice colonized with bacteria.21 Tight junctions are composed of a protein mesh that seals intercellular spaces in the epithelial layer and epithelial permeability is a characteristic of several chronic inflammatory diseases, including CD and UC.22, 23 This is particularly pertinent given a previous study demonstrating that mice colonized with the probiotic Escherichia coli Nissle 1917 caused increased expression of the protein zonula occludens 1, involved in the formation of tight junctions.21 Indeed, other probiotic species such as Lactobacillus plantarum MB452 have been shown to enhance epithelial integrity, specifically through induction of genes involved in tight junction formation, including occludin.24 This suggests that this phenomenon is not restricted to a specific bacterial species, but may represent a hallmark of several commensal organisms that promote epithelial barrier function, which likely confers a fitness advantage on these species in this niche putatively through reducing host inflammatory responses. These findings underscore the contribution of “beneficial bacteria” to their host and the need to better understand the functional interplay between key GI microbes and the host in an effort to harness these species as potential bacteriotherapeutic species.


The GI tract, particularly the lower GI tract where bacterial cell density can reach 1 × 1014 cells, represents a crucial interface between host and environment and, as mentioned, plays a key role in both educating and modulating host responses. Several lines of evidence support this; germfree mice possess fewer Peyer's patches, a thinner lamina propria, fewer plasma cells in germinal centers, and fewer lymphoid follicles (reviewed by Round and Mazmanian25). These animals also exhibit fewer T cells, and Paneth cells that exhibit reduced gene expression, as well as B cells that produce less IgA compared to conventionally raised mice. The collective impact of these changes is large, given the key functions and influence these individual cell types exert on the GI microbiota. For example, Paneth cells, located in the crypts of the small intestine, secrete defensins, small antimicrobial peptides (AMPs) that are largely believed to protect overlying epithelial stem cell populations, crucial for epithelial barrier renewal. Some of these AMPs are quite specific in their microbial targets; for example, RegIIIγ, a C-lectin AMP, specifically targets Gram-positive bacteria. Loss of this fraction from the GI microbiota could have dramatic effects on both the bacterial assemblage and host, since several of the key species known to contribute to immune homeostasis are Gram-positive species, e.g., segmented filamentous bacteria, which has recently been shown to impact Th17 cell proliferation in the terminal ileum of mice.26

There are several mechanisms known for the immunological response elicited by pathogenic bacteria, such as effector systems on pathogenic islands, e.g., the Type III secretion system possessed by many pathogens such as Yersinia pestis or the flagella of Salmonella.27, 28 However, limited information exists explaining why resident beneficial bacteria in the GI tract fail to trigger the host's immune response and thereby can coexist with the host within this niche. Studies have cited various mechanisms that particular bacteria have evolved to suppress the immunological response.29, 30 Several bacteria inhibit nuclear factor kappaB (NF-κB) activation; Bacteriodetes thetaiotaomicron, for instance, is known to act downstream of Toll-like receptor (TLR) signaling and NF-κB activation.31 Other studies demonstrate alternative mechanisms for tolerance of commensal bacteria.32, 33 Some of these are described by Cario and Podolsky,34 who review the tolerance of intestinal immunity through TLRs. Consequently, the host is not simply blind to the presence of indigenous bacteria but employs a two-way communication resulting in a tolerant mutually beneficial arrangement.


As discussed above, the gut microbiota is key to appropriate development of local adaptive and innate response against microbial and nonmicrobial exposures. However, it is becoming apparent that these communities also have far-reaching effects at sites outside of the GI tract, e.g., respiratory tract (reviewed by Rauch and Lynch35). Early events in GI colonization have, for example, been linked to childhood allergic disease development. Children with a high abundance of GI E. coli or Clostridium difficile at 3 weeks of age have a significantly higher risk of developing childhood asthma.36 The GI microbiota has also recently been shown to play a role in liver function, including deconjugation of bile acids (by some Lactobacilli), and insulin resistance.37 Dumas et al37 conducted a study using a murine model susceptible to dietary-induced impaired glucose homeostasis and nonalcoholic fatty liver disease (NAFLD). The investigation demonstrated, using metabolic profiling, that the genetic predisposition of these mice to develop disease was associated with disruption of choline metabolism, a process highly dependent on the GI microbiota. The study indicated that the GI microbiota may, therefore, play an active role in the development of insulin resistance in human populations (Figure 1).

To underscore the far-reaching effects of the gut microbiota, one recent article also reported the effects of this assemblage on the ocular lens.38 The authors measured lipid content of the lens and retina of mice with a conventional gut microbiota and compared it to that of germfree animals. They demonstrated that conventional mice possessed significantly less lens-associated phosphatidylcholines, indicating exposure to oxidative stress. Although association does not necessarily infer causality, this observation is particularly pertinent when one considers the extraintestinal manifestations of IBD, which have definitively been associated with a GI microbiota dysbiosis.39 Extraintestinal disorders afflicting up to 36% of IBD patients include ocular, hepatobiliary, musculoskeletal, dermatologic, genitourinary, vascular and hematologic, cardiac, pulmonary, endocrine, and metabolic syndromes.40 There is also emerging evidence for local microbiota dysbiosis in many of these sites that are involved in other disorders.41–43 However, what is unclear is whether patients who exhibit these disorders, independent of IBD development, also possess a GI microbiota dysbiosis. Nonetheless, what is clear is that the intestinal microbiome may impact host physiology far beyond local digestive functions, and considering this as a critical ancillary organ in the context of the human host, is necessary to truly understand its role in the fundamentals of human physiology and health status.


Like many other diseases, IBD has been shown to have a genetic component. Through genome-wide association studies (GWAS), over 99 nonoverlapping genetic risk loci have been associated with IBD, as reviewed by Khor et al.44 These risk loci can be categorized into distinct functional groups involved in, for example, epithelial barrier function, lymphocyte activation, and multiple other categories of immune functioning. In fact, a large number of the risk loci identified play key, and sometimes multifaceted, roles in host immune responses. NOD2 (also known as CARD15), a member of the caspase recruitment domain protein family, is necessary for recognition of muramyl dipeptide, a cell wall constituent of many Gram-positive and -negative bacteria. In addition, mutations in this gene result in significantly reduced interleukin 10 (IL-10) expression,45 leading to enhanced inflammatory responses. NOD2 encodes a protein expressed constitutively in Paneth cells,46–48 a subset of epithelial cells at the base of intestinal crypts that predominate in the terminal ileum. Paneth cells are important in shaping the microbiota as they produce antimicrobial defensins, which cull the intestinal microbial community.10, 49 Hence, mutations in this gene undoubtedly impact the microbiota composition, uncoupling the delicate balance between immune and microbial community homeostasis. This is supported by observations that individuals with IBD-associated NOD2 mutations exhibit abnormally close adherence of intestinal microbes to the epithelial layer,50 and CD patients with a NOD2 variant exhibit serologic responses to microbial antigens that are absent in patients without this variant.51

More recent studies have begun to uncover the specific impact of such mutations on the GI microbiota composition; CD patients harboring mutations in NOD2 and ATG16L1 possessed a microbiota that specifically exhibited shifts in the relative abundance of members of the Faecalibacterium and Escherichia genera.52 Moreover, a recent study demonstrated a clear role for NOD2 in the development of the microbial community in weaning mice,53 supporting an emerging hypothesis that the predisposition to develop IBD may be laid down during the initial stage of GI microbiota development in infancy. This certainly fits with the paradigm proposed for other inflammatory diseases such as asthma, in which early dysbiosis in GI microbiota (at 3 weeks of age) is associated with a significantly higher risk of development of allergic disease in childhood.36 Collectively, these data suggest that mutations in the NOD2 gene, one of 99 known risk loci associated with IBD, can affect innate sensing of microbes, microbial community structure, and redefine early events in GI colonization, factors that likely play key roles in both initiating or perpetuating inflammation in IBD patients.

As studies uncover the roles of other risk loci in IBD, the complexity and multitude of pathways involved and their association with the GI microbiota is becoming more apparent. For example, genetic variants in protein regulators, single immunoglobulin IL-1R-related molecule (SIGIRR) of TLRs (membrane-bound receptors for bacterial ligands), are also located on an IBD susceptibility locus. SIGIRR-deficient mice have been shown to develop more severe colitis following dextran sodium sulfate (DSS) administration compared with wildtype animals,54 indicating a protective role for the microbiota against disease development. Variants of genes for both TLR2 and TLR9, key membrane-bound receptors critical to bacterial recognition, have also been associated with CD.55, 56 It has also been documented that IBD is associated with defects in autophagy, which can involve genes such as ATG16L1.57 Not only have ATG16L1 mutations been linked with the inability of hosts to clear infections by intracellular pathogens, but reductions in expression levels of ATG16L1 have also been linked to defects in Paneth cells,58 which clearly can impact microbiota membership through altered antimicrobial peptide production. Increased cytoplasmic vesicles were observed in Paneth cells of ATG16L1 hypomorphic cells, via electron microscopy, a phenomenon previously reported in CD patients.59 This may explain, in part, why patients with mutations in ATG16L exhibit a distinct microbiota compared to those who did not possess these genotypes.52 Such associations raise questions about the extent of the role to which an altered microbiota plays in disease development. However, data demonstrating that CD patients homozygous for ATG16L1 have deformed Paneth cells in uninflamed regions of the GI mucosa60 implies that although mutations in ATG16L1 can alter the microbiota, other factors may be necessary to induce inflammation at a particular site, emphasizing the multivariate nature of IBD.

Although genetic risk loci are important factors in understanding IBD disease mechanisms, they cannot explain the disease in its entirety, and thus, it has been suggested that IBD also possesses an environmental component. Twin studies support this hypothesis61; disease concordance between both monozygotic and dizygotic twin pairs is 10%–15% for UC and 30%–35% for CD in monozygotic twins.62 These data provide evidence of the interplay between the environment and genetic components, and also suggest that environmental factors (including microbial exposures) may play a larger role in development of UC compared with CD.


GWAS studies and subsequent investigations delineating the role of target risk loci and IBD development have underlined the complex interplay between host immunity and the microbiota in IBD. Further support for the role of the GI microbiota–host immune response interplay in IBD development comes from the observation that disease does not develop in immunocompromised animals raised under germfree conditions, while those reared in the presence of microbes develop inflammation.63 Several recent studies have begun to deconstruct host–microbe interactions and demonstrated a clear role for specific members of the GI microbiota in modulating inflammatory responses relevant to IBD. IL-17, a proinflammatory cytokine produced specifically by the Th-17 arm of the lymphocytic response, is significantly increased in expression in patients with IBD compared with healthy controls. In addition, increased expression was detected in biopsies from inflamed regions compared to healthy ones originating from the same patient,64 demonstrating focal induction of this potent proinflammatory cytokine at sites of inflammation along the GI mucosa. Recently, using culture-independent methods to profile isogenic mice from two different suppliers, who were fed distinct chow and exhibited differential Th17 cell populations in the lamina propria of the terminal ileum, Ivanov and colleagues identified a specific member of the GI microbiota, segmented filamentous bacteria (SFB), as a critical inducer of Th-17 cell proliferation at this site.26, 65, 66 More recently, Salzman et al67 demonstrated that mice expressing a human-derived antimicrobial peptide (DEFA-5), exhibit dramatic depletion of SFB, and, as a consequence, fewer IL-17-expressing T cells. This study both confirms the key role for this species in Th-17 cell induction and demonstrates the key importance of antimicrobial peptides in shaping microbiota membership. It also illustrates that loss of specific bacterial species can dramatically shift immunological status and provides a possible explanation for the observation that, while SFB are found in the GI tracts of many mammalian species, they are absent in humans who express DEFA-5 AMP. Other AMPs, such as RegIIIγ produced by Paneth cells, have been associated with IBD. Patients with ileal CD exhibit a reduction in Paneth cell numbers and, by extension, RegIIIγ production.68 This was observed only in patients with inflammation confined to the ileum, implying that the effect on the resident microbiota may be localized, leading to focal inflammation of the small intestine.

Collectively, these findings suggest that factors influencing the relative abundance of such species at key immunomodulatory sites along the GI tract may drive aberrant inflammatory responses and that strategies aimed at rehabilitating the microbiota composition and, specifically, the appropriate abundance of these species may represent novel therapeutic strategies to combat chronic inflammatory disease, both in the GI tract and beyond.


IBS clearly involves a breakdown in relations between the host immune response and microbial population resident in the GI tract. Although multiple studies to date have failed to reveal a single etiological pathogenic species responsible for IBD, the current view is that while individual species may play significant roles in immunomodulation, collateral damage to the microbiome due to their loss or overabundance, plays a key role in persistence of inflammatory responses in chronic disease. Evidence for the role of the microbiota in pathogenesis is provided through studies demonstrating that antibiotic use can reduce or prevent inflammation both in patients and in murine models of disease.69, 70 In humans, fecal diversion have been successful in ameliorating disease in patients with CD71; prevention of the fecal stream from passing through inflamed sections of the intestine has successfully ameliorated disease. However, disease recurred when fecal passage resumed. Fecal bacteriotherapy,72 primarily indicated for recurrent C. difficile-associated diarrhea (CDAD), has more recently been indicated as a potential therapeutic strategy for patients with UC.73 The approach involving transfer of fecal material from a healthy donor to a patient with chronic inflammatory disease has demonstrated efficacy in CDAD patients.74 That the strategy is effective suggests that restoration of microbiome diversity that undoubtedly restores microbial community functionality and appropriate host interactions may lead to highly efficacious outcomes in treating IBD. Indeed, initial studies of UC patients inoculated with stool collected from healthy donors exhibited disease remission within a week of receiving their fecal transfer, with complete recovery noted after 4 months.72

Given what we know of the dysbiotic nature of the GI microbiota in IBD, an obvious question is which key members and functions are depleted in the disease state. Dramatic reductions in the relative abundance of members of the Firmicutes, a major phylum present in the intestinal microbiota,75 have been observed in IBD patients. Particular interest has been placed on members of this group since they are known producers of important short chain fatty acid metabolites such as acetate and butyrate, substrates with potent antiinflammatory properties.76–78 Specifically, Clostridial clusters IV and XIV exhibit relatively lower abundance in patients with IBD compared to healthy controls, suggesting that their loss may deplete the microbiota of key antiinflammatory metabolites or other cell-associated immunomodulatory ligands.

Several investigators have documented alterations in the microbiota associated with CD, particularly a decrease in members of the phylum Firmicutes and concomitant increase in Proteobacteria.52, 79, 80 A reduction in the Clostridium leptum group, particularly Faecalibacterium prausnitzii,81–83 has been described in these patients. Anti-inflammatory properties of this bacterium, e.g., production of butyrate, lend support to its positive association with GI health and suggest that its depletion may disrupt host tolerance to the intestinal microbial community, which may precede or perpetuate intestinal inflammation.

Through sequencing of the full-length 16S rRNA phylogenetic biomarker gene, inflamed and noninflamed sites have been performed within a cohort of IBD patients; significant differences were evident between these sites.52 There was a reduction in bacterial diversity in inflamed regions, even within the same patient. UC patients also typically exhibit a higher bacterial load than those suffering from CD; however, bacterial burden was lower than those observed in the healthy controls.79 Furthermore, CD patients exhibit lower bacterial loads, specifically at regions of inflammation.52 Such differences were also observed by others using terminal restriction fragment length polymorphism (T-RFLP)84, 85 and fluorescent in situ hybridization (FISH).86 Collectively, these data suggest that induction of host inflammation contributes to the loss of diversity characteristic of IBD patients. This is particularly pertinent given the clear ability of particular organisms, e.g., S. enteritica Typhimurium, to not only induce inflammatory responses, but to proliferate under these conditions due to the inherent fitness advantage inflammation confers on this species. That particular members of the microbiota have a pronounced presence in the diseased state, e.g., Peptostreptococcus, Eubacteria, and Coprococcus, in higher relative abundance in CD patients,87 has been confirmed by the detection of elevated serum antibodies for these species and suggests that these community members may play individual or collective roles in disease pathology, although this remains to be determined.

Significant differences between IBD patients and non-IBD subjects have been noted, and some studies have demonstrated compositional differences between UC and CD patients81, 88 (Table 1). However, other studies have reported indistinguishable microbiota between these two patient groups.89, 90 These discrepancies may be due to the depth of microbiota profiling performed, the tools employed to analyze these microbial communities, or the stage of disease investigated. In addition clear niche-specificity in microbiota composition exists along the length of the GI tract, thus the site of sample acquisition, if insufficiently standardized, may introduce sufficient variability in community composition as to mask the key differentials underlying patient groups.

Table 1. Documented Microbial Alterations in IBD Patients
Changes in MicrobiotaDiseaseSample SourceMethodYearAuthor
  1. ↑Increase in bacteria compared to healthy controls.

  2. ↓Decrease in bacteria compared to healthy controls.

  3. UC, ulcerative colitis; CD, Crohn's disease; qPCR, quantitative PCR; Cl, clones; FISH, fluorescent in situ hybridization; Pyro, 454 pyrosequencing; DGGE, denaturing gradient gel electrophoresis; TGGE, temperature gradient gel electrophoresis; T-RFLP, terminal restriction fragment length polymorphism; ARISA, automated ribosomal spacer analysis; PMA, phylogenetic microarray.

Escherichia coliCD, UCTissueCulture2011Thomazini et al136
Enterobacteriaceae, Rumicoccus gnavusCDTissueCl2011Frank et al52
ClostridiumCD, UCFecesT-RFLP2011Andoh et al88
BacteriodetesCDFecesT-RFLP2011Andoh et al88
FirmicutesCD, UCTissueCl, qPCR2011Walker et al79
BacteriodetesCD, UCTissueCl, qPCR2011Walker et al79
EnterobacteriaceaeCDTissueCl, qPCR2011Walker et al79
Bacteriodes vulgatusCD, UCTissueCulture, qPCR2011Walker et al79
Dialister invisus, Clostridium cluster XIVa, Faecalibacterium prausnitzii, Bifidobacterium adolescentisCDFecesDGGE2011Joossens et al137
Ruminococcus gnavusCDFecesDGGE2011Joossens et al137
Actinobacteria, ProteobacteriaUCTissueCl2011Lepage et al138
Faecalibacterium, RoseburiaCDFecesPyro2010Willing et al139
Enterobacteriaceae, Ruminococcus gnavusCDFecesPyro2010Willing et al139
ClostridiumUCTissueqPCR2010Verma et al140
EubacteriumCDTissueqPCR2010Verma et al140
MethanobrevibacterCD, UCTissueqPCR2010Verma et al140
↑ Sulfur reducing bacteria (SRB)CD, UCTissueqPCR2010Verma et al140
Ruminococcus, Lactobacillus, Bifidobacterium, BacteriodetesCD, UCTissueqPCR2010Verma et al140
Faecalibacterium praunitizii, BifidobacteriaCDFecesqPCR2010Schwiertz et al141
Escherichia coliCDFecesqPCR2010Schwiertz et al141
BifidobacteriaUCFecesqPCR2010Schwiertz et al141
Eubacterium rectale, Bacteroides fragilis group, B. vulgatus, Ruminococcus albus, R. callidus, R. bromii, Faecalibacterium prausnitziiCDFecesPMA, qPCR2010Kang et al142
Enterococcus sp., Clostridium difficile, Escherichia coli, Shigella flexneri, Listeria sp.CDFecesPMA, qPCR2010Kang et al142
Bacteroides vulgatus, B. ovatus, B. uniformis, Parabacteroides sp.UCFecesDGGE2010Noor et al143
StreptococcusCDTissueCulture2009Fyderek et al144
LactobacillusUCTissueCulture2009Fyderek et al144
BifidobacteriaCD, UCTissueCulture2009Fyderek et al144
Faecalibacterium praunitiziiCDTissueT-RFLP, Cl, qPCR2009Willing et al82
Escherichia coliCDTissueT-RFLP, Cl, qPCR2009Willing et al82
Ruminococcus obeum, R. gnavusUCTissueT-RFLP2009Nishikawa et al145
Bacteroides, EnterobacterialesCDFecesT-RFLP2009Andoh et al148
Clostridium cluster IV, XIVa,CDFecesT-RFLP2009Andoh et al148
Bacteroides fragilis, B. vulgatus, B. ovatus, Clostiridium coccoides, C. leptum, Atopobium, Bacteroidaceceae, Bifidobacteria, VeillonellaCD, UCFecesFISH, qPCR, culture2008Takaishi et al146
Lactobacillus, EnterococcusCD, UCFecesCulture2008Takaishi et al146
Faecalibacterium praunitiziiUCFecesqPCR2008Sokol et al83
Enterobacteria, ActinobacteriaCDTissueFISH2008Sokol et al83
Firmicutes, BacteriodetesCDTissueFISH2008Sokol et al83
Bacteroides uniformCDFecesT-RFLP2008Dicksved et al147
Bacteroides ovatus, B. vulgatusCDFecesT-RFLP2008Dicksved et al147
ClostridiaCD, UCTissueARISA, T-RFLP2007Sepehri et al85
BacteriodetesCD, UCTissueARISA, T-RFLP2007Sepehri et al85
Clostridium leptum, C. coccoidesCDTissueTTGE2007Sokol et al
LachnospiraceaeCD, UCTissueCl2007Frank et al81
ProteobacteriaCD, UCTissueCl2007Frank et al81
Faecalibacterium praunitiziiCDFecesFISH2007Swidsinki et al149
Faecalibacterium praunitiziiUCFecesFISH2007Swidsinki et al149
Enterobacteriaceae, Bacteroides fragilis, Faecalibacterium prausnitzii-like “Butyrate-producing bacterium” L2-6, Pseudomonas aeruginosaUCTissueCl2007Wang et al150
Firmicutes (C. leptum)CDFecesFISH, Cl2006Manichanh et al89
↓ Bacteriodes, lactic acid bacteria (LAB)CDFecesDGGE2006Scanlan et al151
Clostridium coccoides, C. leptumCDFecesFISH2006Sokol et al
BacteroidetesCDTissueFISH, Cl2006Bibiloni et al152
PorphyromonadaceaeUCTissueFISH, Cl2006Bibiloni et al152
VerrucomicrobiaCDTissueFISH, Cl2006Bibiloni et al152
Bacteriodetes, ProteobacteriaCDTissueCl2006Gophna et al80
ClostridiaUCTissueCl2006Gophna et al80
Eubacteria rectaleCD, UCTissueFISH2005Swidsinski et al70
Bacteroides fragilis, Bacteriodetes-ProvetellaCD, UCTissueFISH2005Swidsinski et al70
Bacteriodetes, Eubacterium, LactobacillusCDTissueCl2004Ott et al75
Clostridium coccoidesCDFecesTGGE2003Seksik et al153
EnterobacteriaCDFecesTGGE2003Seksik et al153

In addition to loss of key microbiome members, relative increases in the abundance of potentially pathogenic members of the community may also contribute to disease.52, 79, 80, 91, 92 There is precedence for this: for example, C. difficile proliferates under conditions of antimicrobial administration when the background microbial diversity has been depleted.93, 94 These data emphasize the importance of interactions within the microbiota itself and that in addition to microbial–host interactions, microbe–microbe interactions likely play a major role in community self-regulation and defining the physiology and functionality of members of the assemblage.


Although human studies have indicated a role for the microbiota in IBD development, to further understand this relationship between microbiota and host immunity and its degradation in inflammatory disease of the intestine, animal models represent an attractive avenue of investigation. Rodent studies have been useful in attempting to tease apart components involved in disease development. For example, IL-10 deficient (IL-10−/−) mice have played a large role in defining the role of microbes in the onset of IBD; conventionally raised IL-10−/− mice, or those gavaged with a fecal slurry (and not sterile filtrate of the fecal slurry) developed disease,95 indicating that the microbial-associated molecular patterns (MAMPS) are responsible for stimulation of the immune response.96

It has also been postulated that resident bacteria may cause colitis by compromising the colonic epithelial barrier.97 Ohkusa et al97 demonstrated that several indigenous bacteria were capable of inducing varying levels of cytokine production from host intestinal tissue. The activity of these inflammatory molecules suggests that these microbes could induce and maintain inflammation and that their lack of inflammation-inducing activity in vivo may be due to host responses or, indeed, the activities of neighboring species in the microbiota.

DSS is commonly used as a means to reliably induce colitis.98 A vast number of studies have examined the effects of DSS on the host tissue and immune response as it relates to UC.99, 100 Using this model, Medzhitov and colleagues101 have shown that TLR4 knockout mice did not develop colitic lesions upon treatment with DSS. MyD88 knockout mice (which are unable to signal through TLRs) also fail to develop the characteristic lesions of UC following DSS administration.32 And, more recently, Ungaro et al102 showed that TLR4 antagonist antibody ameliorates inflammation in colitic mice. These studies and others clearly demonstrate that intestinal bacteria play a role in the development of UC via TLR signaling pathways.

Some studies have shown that germfree mice administered DSS develop more severe disease than their conventional counterparts suggesting a protective effect of the microbiome.103 This indicates that the microbiota's beneficial effects may be due to production of metabolites such as short chain fatty acids (SCFA) that are necessary for colonocyte development and epithelial cell health.15 However, others have shown that administration of anaerobic bacterial antigens reduced the severity of disease in mice treated with DSS,104 leading the authors to speculate that ingested antigens increased tolerance to the intestinal bacteria. This suggests that altered homeostasis between the microbiota and host immune response may be the driver for immune reaction to DSS.104 The authors also postulated that the effects may be due to antigens altering the microbial community by competing for binding sites on epithelial surfaces, altering the immune system that may have subsequently exhibited effects of the microbiota.

Antibiotic studies revealed that certain antimicrobial therapies reduce inflammation,105, 108 indicating both a protective and detrimental effect of the microbiota on colitis. Araki et al106 treated Caco-2 cells in vitro with DSS and demonstrated that H2S, which is associated with DSS-induced colitis in mice and UC in humans, was not produced by the host in response to exposure to the chemical, emphasizing that microbial metabolism is necessary to produce this toxic substance.107 Others have tried to explain the negative role of microbiota by postulating that microbes act on DSS and metabolize it to a form that causes inflammation. Kitajima et al103 incubated DSS with luminal contents for 24 hours before the components were examined using spectroscopy. They demonstrated that the microbes did not metabolize DSS, dispelling this potential mechanism of action. However, it is possible that members of the mucosal microbiota and not the luminal communities (which exhibit distinct compositions) are involved in DSS metabolism, a factor not examined in that study, particularly since multiple other studies indicate such an important role for the microbiota in this model of IBD.109–111

An additional useful model of IBD is infection of IL-10−/− mouse by Helicobacter hepaticus.112 There have been several studies showing the development of colitis in IL-10−/− mice infected with this microbe,113–115 which involves T-cell responses.112 Recent evidence has also linked IL-23-induced Th 17 cells in the development and modulation of this infection.116 It is tempting to speculate that this may be due to shifts in the relative abundance of GI species such as SFB, which clearly plays a role in the induction of this T-cell population. Dieleman et al63 demonstrated that disease only ensues in the presence of a resident microbiota and not in H. hepaticus monoassociated mice. It should be kept in mind that germfree animals are not “normal,” in that they do not have a developed immune system and would not react as conventional animals. Also, disease does not occur in mice with an intact immune system, since several studies with this bacterium in wildtype animals have not resulted in inflammation.117 Furthermore, Helicobacter spp. have been found to cause the dysregulation of TLR2 and NOD1 in epithelial cells,118 receptors for bacterial antigens. Investigation of the role of TLR4 and IL-10 in the H. hepaticus-triggered model of colitis show that TLR/IL-10 deficient mice have elevated interferon-gamma (INFγ), IL-17, tumor necrosis factor alpha (TNFα), IL-1, and IL-6 when infected with the bacterium, emphasizing the role of resident bacteria in induction of disease.

Investigators have also identified “colitiogenic” bacterial communities.119, 120 TRUC mice (Rag deficient and T-Bet deficient), for example, induced shifts in the intestinal bacterial community simultaneously with IBD development. Subsequent transfer of this altered community induced inflammation in wildtype mice, suggesting a causative role for the microbiota.


Given the preponderance of human microbiota studies and the importance of GI communities in maintenance of immune homeostasis and host health, it is unsurprising that the field of probiotic research has experienced recent renewed interest. Probiotics are defined as “living microorganisms which, when administered in adequate concentration, confer a health benefit on the host.”15 These benefits may arise through multiple mechanisms. Probiotic species can colonize the GI mucosa and occupy niches, preventing their colonization by potential pathogens in a phenomenon known as colonization resistance (reviewed by Callaway et al121). This strategy of competitive microbial exclusion may be achieved through depletion of available nutritional resources, or altering the environment (e.g., pH, mucus production), thus creating ecosystem conditions that are hostile to species unable to compete under these circumstances.122 The beneficial bacteria may also alter gene expression profiles of potential pathogens, resulting in decreased expression of virulence factors.123 Many of the genera used, such as Lactobacillus and Bifidobacteria, are resident members of the healthy GI community, and putatively functionally important in maintaining health status. These organisms work synergistically with the intestinal community to maintain health, and are therefore considered viable therapeutic strategies for management of IBD.

Specific probiotic species have also been shown to modulate host immune responses. Lactobacillus rhamnosus GG, for example, has been shown to activate NF-κB in macrophages.124 Conversely, L. plantarum, can actively block NF-κB signaling, resulting in suppression of inflammation.125 Moreover, it appears that the factor responsible for this phenomenon is not cell-associated. A cell-free solution from cultures of L. plantarum resulted in inhibition of NF-κB expression from murine intestinal epithelial cells, specifically through both MyD88-dependent and -independent pathways via the TNF receptor. Clearly, the latter case highlights that, in addition to direct contact with microbial ligands, bacterial exoproducts may also mediate their effect on host immune responses.

Martin et al126 studied the effects of two probiotics, Lactobacillus paracasei and L. rhamnosus in mice colonized with human baby flora (HBF). The presence of these bacteria altered the blood and tissue metabolic profiles of these animals. L. paracasei impacted energy metabolism pathways, increasing accumulation of bile acids, leading to increased lipid absorption in the intestine.126 Bacteria change the lipid composition of the feces and the vitamin and mineral uptake in the gut. Also, complex carbohydrates are broken down into SCFAs with the aid of gut microbes. These functions indicate that it is a product of the bacterium and not the microbe itself that has the beneficial effects, exemplifying the various ways the microorganisms affects host physiology.

Despite the increasing numbers of potential probiotics cited as possible treatments for IBD,24, 125, 127 few have been proven to be effective against UC, and none against CD (reviewed by Haller et al128). E. coli Nissle 1917 and VSL#3 (a mix of L. casei, L. plantarum, L. acidophilus, L. delbrueckii subsp. bulgaricus, B. longum, B. breve, B. infantis, and S. salivarius subsp. thermophilus) were both successful in maintaining remission of UC,129, 130 but not CD. E. coli Nissle 1917 also appeared to be even more effective when used in combination with other agents, such as balsalazide (an anti-inflammatory drug), in the treatment of mild to moderate colitis.131 In the case of VSL#3, the role of each bacterium is still to be elucidated. Although many trials yield neutral outcomes, factors such as low power, insufficient bacterial numbers per dose and use of inappropriate probiotic species or strains to target the specific gastrointestinal sites affected by the disease, may be confounding. Thus, species tested with unfavorable results may still hold hope for IBD treatment.


Developing therapies for IBD is particularly challenging since this disease is clearly complex and a result of a multifaceted host–microbial interactions.61 Investigators have identified genetic variants, immunologic dysfunction, and a role of the GI microbiome in this disease. The major challenge in this field is, as for other chronic inflammatory diseases, elucidating whether changes in the microbial assemblages represent cause, or consequence, of host inflammation and disease state. Indeed, both scenarios may occur in initiation and perpetuation of the inflammation present in IBD. It is possible that alterations in the microbial community initiates inflammation and, once host mediators storm intestinal tissues, they exert pressure on the intestinal microbiota that further shifts community structure, thus creating a feedback loop that perpetuates inflammation. In characterizing these changes in community structure and function, it may be possible to alter these events to circumvent or ameliorate IBD.

In characterizing the microbial community, we also need to consider other members of the microbiota such as fungi and viruses.132 Many of the techniques used presently utilize the 16S rRNA gene, specifically used to profile bacterial communities. Being such a complex network of interactions, it is not difficult to imagine that fluctuations in the relative abundance and presence of fungal and viral species could have consequences on the composition and function of the overall community and host response. With the advent of deep sequencing techniques, it is becoming more apparent that not only are global shifts in bacterial community composition associated with disease, but that more subtle changes are important factors in defining community functionality,133 which is ultimately central to host–microbiome interactions. This is not surprising, since it is accepted that microbial keystone species are not necessarily the dominant members of a given community.134 Therefore, not only is it necessary to quantify the dominant members of the community, but it is also necessary to survey the community for less abundant, “rare” members that may contribute key functions pivotal to microbial–microbial and microbial–host interactions.

Since organisms that share similar traits tend to coexist, some communities are predisposed to invasion by pathogens or, conversely, to colonization by beneficial microbes.135 Such microbiota predictors may be useful prognostic or diagnostic indicators of disease development, recurrence, or treatment response. As the field of human microbiome research expands, integrated studies that examine microbiome–host interactions are revealing avenues for novel therapeutic development and improved diagnostics that ultimately will improve our ability to combat chronic inflammatory diseases, including IBD.