REvIEwS

Host–microbiota interactions in
inflammatory bowel disease
Roberta Caruso1,2, Bernard C. Lo1,2 and Gabriel Núñez 1
*
Abstract | The mammalian intestine is colonized by trillions of microorganisms that have
co-evolved with the host in a symbiotic relationship. The presence of large numbers of symbionts
near the epithelial surface of the intestine poses an enormous challenge to the host because
it must avoid the activation of harmful inflammatory responses to the microorganisms while
preserving its ability to mount robust immune responses to invading pathogens. In patients with
inflammatory bowel disease, there is a breakdown of the multiple strategies that the immune
system has evolved to promote the separation between symbiotic microorganisms and the
intestinal epithelium and the effective killing of penetrant microorganisms, while suppressing
the activation of inappropriate T cell responses to resident microorganisms. Understanding the
complex interactions between intestinal microorganisms and the host may provide crucial insight
into the pathogenesis of inflammatory bowel disease as well as new avenues to prevent and treat
the disease.

Pathobionts
Microorganisms that, under
normal circumstances, live as
non-​harmful symbionts but can
induce pathology under certain
conditions, usually involving
environmental and/or genetic
alterations.
1
Department of Pathology
and Rogel Cancer Center,
the University of Michigan
Medical School, Ann Arbor,
Michigan, USA.
2
These authors contributed
equally: Roberta Caruso,
Bernard C. Lo.
*e-​mail:
gabriel.nunez@umich.edu
https://doi.org/10.1038/
s41577-019-0268-7
Inflammatory bowel disease (IBD), which includes
Crohn’s disease and ulcerative colitis, is a chronic and
relapsing inflammatory disorder of the intestine that
affects ~3 million people in the United States1. Although
the pathogenesis of IBD is poorly understood, multiple
lines of evidence suggest that the disease is caused by
a confluence of genetic and environmental factors that
alter gut homeostasis, thereby triggering immune-​
mediated inflammation in genetically susceptible indi-
viduals2. Genetic studies have identified more than 200
loci that regulate IBD risk, most of which are associated
with immunological pathways that regulate microbial
recognition and killing, immune responses to micro-
organisms and the reinforcement of the intestinal bar-
rier3,4. Although the pathogenesis of both Crohn’s disease
and ulcerative colitis involves intestinal inflammation,
these two disorders differ in several features, including
the association with specific susceptibility loci and the
type of immune response and pathology associated with
disease2,4 (Box 1).
A key aspect of both forms of IBD pathogenesis is
their link to the presence of symbiotic microorganisms
living in the intestinal tract. The resident symbionts
contribute to many host physiological processes, includ-
ing digestive and metabolic functions, regulation of
the epithelial barrier, development and regulation of the
host immune system, and protection against patho­gen
colonization5,6. Whereas the great majority of intes­
tinal microorganisms live in a mutualistic relation-
ship with the host, some symbiotic organisms, such as
Mucispirillum schaedleri and Helicobacter species, are
often referred to as pathobionts because they can cause
disease under certain conditions. The composition of
the gut microbiota exhibits broad inter-​individual and
intra-​individual variability7,8 and is thought to be a cru-
cial determinant of host susceptibility to several diseases,
including IBD9–11. A key feature of IBD is alteration of
the composition of the gut microbiota (known as dysbi-
osis)12; however, the precise role of dysbiosis in disease
remains poorly understood.
The close proximity of a large number of symbiotic
microorganisms to the epithelial surface represents
a unique challenge to the mucosal immune system in
that it must preserve the ability to mount an immune
response against an invading pathogen while avoiding
harmful inflammatory responses to the gut micro­
biota. In this Review, we discuss the mechanisms that
limit the inappropriate accumulation of particular
microorganisms near the epithelial surface and pro-
mote the clearance of penetrant organisms to minimize
the activation of harmful inflammatory responses
in the gut. We also discuss how IBD-​associated genetic
mutations can disrupt these protective mechanisms,
which have been referred to as ‘mucosal firewalls’13.
In addition, the immune system has developed effec-
tive regulatory mechanisms to suppress the activ­
ation of inappropriate T cell responses to the resident
microbiota that can be compromised in IBD. We also
describe how IBD-​associated genetic defects can lead
to pathobiont accumulation and penetration into the
intestinal tissue, which further promotes dysbiosis and
inflammation. This Review focuses on bacteria as these

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Reviews
Box 1 | Comparing Crohn’s disease and ulcerative colitis
Crohn’s disease and ulcerative colitis, the two major forms of inflammatory bowel
disease in humans, are chronic relapsing inflammatory disorders. although these two
diseases have historically been studied together and share some clinical characteristics,
it is now clear that they represent two distinct pathophysiological entities. Crohn’s
disease is characterized by segmental inflammation with sharp demarcation between
involved and uninvolved bowel segments (skip lesions). Pathologically, Crohn’s disease
is characterized by transmural inflammation, narrow and penetrating ulcers, and the
presence of non-​caseating granulomas in many patients. although lesions in Crohn’s
disease may affect any site of the gastrointestinal tract, the terminal ileum is most
commonly affected and the earliest mucosal lesions in Crohn’s disease are often
located over Peyer’s patches.
unlike Crohn’s disease, ulcerative colitis is characterized by continuous inflammation
that extends proximally from the rectum to a variable distance along the colon.
inflammation in ulcerative colitis is superficial, limited to the mucosal layer and
characterized by the presence of neutrophils that infiltrate the lamina propria and the
intestinal crypts, where they form micro-​abscesses (known as cryptic abscesses).
Depletion of goblet cells is also a common histological feature in ulcerative colitis2.
Genome-​wide association studies have identified more than 200 susceptibility genes
that are either shared by both diseases or specifically associated with one disease3.
some of the Crohn’s disease-​specific genes are involved in immune responses against
bacteria, whereas genes associated with the risk of developing ulcerative colitis are
mainly involved in T cell signalling and epithelial barrier function203,204. although
dysregulated intestinal inflammation is the hallmark of both Crohn’s disease and
ulcerative colitis, mucosal inflammation associated with these two diseases is mediated
by different types of effector T cell. Whereas Crohn’s disease is mainly characterized by
an imbalance of effector T cells, predominantly T helper 1 (TH1) cells or TH17 cells, versus
regulatory T cells, ulcerative colitis has a cytokine profile associated with TH17 cells and
tH2 cells2,205,206.
microorganisms have been best studied in the context of
intestinal inflammation; however, the role of other com-
ponents of the intestinal microbiota, such as viruses and
fungi, in IBD is mentioned where appropriate. Finally,
we conclude the article by discussing how the microbiota
can be targeted as a potential therapy for IBD.
Mucosal firewalls
The vast population of symbiotic microorganisms in the
Crypts of Lieberkühn intestine is separated from host tissues by a single-​cell
Invaginations of the small layer of epithelium. To maintain its homeostatic relation-
intestine that contain epithelial ship with the intestinal microbiota, the host has evolved
stem cells and Paneth cells.
several strategies to minimize the contact between
Oxidative burst microorganisms and the epithelial surface and to limit
The rapid production of the presence of penetrant symbionts that could trigger
reactive oxygen species by unwanted inflammatory responses.
phagocytes that can directly
kill bacteria.
Segregating symbionts from the epithelial surface. The
Segmented filamentous host restricts the access of potentially harmful micro­
bacteria organisms to the mucosal surface by several mecha-
(SFB). A species of Clostridium-​ nisms, including mucus secretion and the release of
related bacteria that primarily
inhabits the terminal ileum of
antimicrobial proteins and immunoglobulins into the
mice and promotes T helper intestinal lumen13,14 (Fig. 1). The mucus lining the gut
17 cell development. epi­thelium is composed primarily of highly glycosylated
proteins, including mucin 2 (MUC2), that are secre­
Endoplasmic reticulum
ted by goblet cells. The glycosylation of MUC2 protects
stress
A consequence of dysregulated the protein from degradation by host and bacterial
protein processing in the proteases and promotes interaction with water to form
endoplasmic reticulum that the mucus gel15. In the large intestine, two clearly distinct
initiates unfolded protein mucus layers are present: the outer layer, which contains
response pathways; unresolved
endoplasmic reticulum stress is
large numbers of mucus-​dwelling bacteria, and the inner
associated with inflammatory layer near the epithelium, which is largely depleted of
bowel disease. microorganisms16,17. The inner mucus layer provides
a formidable physical barrier and is also a scaffold for
antimicrobial peptides and immunoglobulins, which
prevent microorganisms from contacting the epithelial
surface18. The small intestine lacks a well-​demarcated
inner mucus layer but it contains large numbers of
Paneth cells, specialized intestinal cells that reside at
the base of the crypts of Lieberkühn and are rich in anti-
microbial molecules19. In response to bacterial stimula-
tion, Paneth cells release their antimicrobial molecules,
including α-​defensins, into the intestinal lumen to limit
the accumulation of bacterial symbionts19,20. In addi-
tion, intestinal epithelial cells (IECs) in the small intes-
tine secrete antimicrobial lectins, such as regenerating
islet-derived protein 3γ (REG3γ), that accumulate in the
mucus layer and further promote the segregation of
the microbiota from the host21.
Although the lamina propria is largely devoid of
acute inflammatory cells, some neutrophils can transmi-
grate into the intestinal luminal side at steady state and
kill bacteria near the epithelial surface through several
mechanisms, including induction of an oxidative burst22.
The cytokine IL-22 also has a role in establishing
host–microbial mutualism by acting on epithelial cells
to mediate barrier function and antimicrobial host
defence23. For example, secretion of IL-22 by group 3
innate lymphoid cells (ILC3s) is required for the contain-
ment of commensal bacteria by inducing the expression
of antimicrobial peptides that prevent systemic dissem-
ination of Alcaligenes spp. bacteria24. Innate sources of
IL-22 are also important for controlling the proliferation
of segmented filamentous bacteria (SFB) in the gut and
limiting T helper 17 (TH17) cell-​mediated colitis25. IL-22
also promotes epithelial glycan fucosylation to support
the growth of mutualistic bacteria that are adapted to
use fucose as a nutrient source26–28. Disruption of IL-22
signalling and fucosylation leads to greater susceptibil-
ity to enteric infections and colitis, in part owing to the
dysbiotic overgrowth of opportunistic pathobionts27,28.
The separation of intestinal bacteria from the epi-
thelial surface is also promoted by IgA, which is pro-
duced by plasma cells that reside in gut-​associated
lymphoid tissues. Large amounts of polymeric IgA
are transcytosed across the epithelium into the lumen,
where IgA has a role in maintaining barrier function
by coating bacteria and binding to microbial antigens
and their toxins as well as in shaping the composition
of the microbiota through unresolved mechanisms29,30.
IgA responses occur constitutively during homeosta-
sis through T cell-​independent and T cell-​dependent
processes that preferentially target distinct microbial
species29. IgA antibodies are typically polyreactive and
bind with low affinity to microbial lipopolysaccharides,
DNA and flagellar antigens29. Microbial sensing through
Toll-​like receptor (TLR) engagement can directly stim-
ulate IgA production, although T cell-​independent IgA
can also be induced independently of the microbiota by
endoplasmic reticulum stress in IECs to provide protection
against intestinal inflammation31. In addition, T cell-​
intrinsic sensing of microbial signals through myeloid
differentiation primary response protein 88 (MYD88)
is important for homeostatic IgA responses to prevent
dysbiosis and enteropathy32. Select members of the
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intestinal microbiota, which are heavily coated by IgA
during colitis, confer enhanced susceptibility to colitis
when transferred to germ-​free animals, which indicates
that preferential IgA binding during dysbiosis may iden-
tify bacterial species that are relevant to IBD33. Whereas
most commensals strongly elicit T cell-​independent IgA
binding, ‘atypical’ bacteria, such as SFB, M. schaedleri,
Prevotella spp. and Helicobacter sp. flexispira, can evade
T cell-​independent antibody responses to come in close
proximity to the epithelial surface, where they elicit an
antigen-​dependent, high-​affinity, T cell-​dependent IgA
response33,34. In contrast to its role in protecting barrier
surfaces by microbial exclusion, IgA can be used by some
symbiotic species to stably colonize the intestine35. These
studies indicate that IgA can enforce a ‘healthy’ micro-
biota by promoting the colonization of beneficial com-
mensals but that, in states of dysbiosis, IgA responses are
induced against potentially colitic species29,30.

Small intestine Large intestine

Neutrophil
Mucus layer Intestinal
lumen Intestinal
microbiota

Goblet cell Detection

and killing

MUC2 IgG

Pathobiont
SIgA
M cell

Transcytosis IFNγ
Epithelium
AMPs,
FUT2
DC
TH1
α-Defensins Plasma cell
cell
TLR TFH cell
Peyer’s patch
Paneth IL-1β,
cell IL-6, IL-17,
IL-23 IL-22
TH17 cell

ILC3 Kupffer
cell
Lymphoid tissue Liver and systemic sites
Lamina propria

Preventing symbiont
Segregating symbionts from the epithelial surface Control at systemic sites
accumulation in the mucosa

Fig. 1 | mucosal firewalls. The host has evolved several strategies to mini-
mize the contact between intestinal microorganisms and the epithelial
surface and to limit the presence of penetrant symbionts that could trigger
unwanted inflammatory responses. Goblet cell secretion of glycoproteins,
including mucin 2 (MUC2), forms a mucus barrier that prevents micro­
organisms in the intestinal lumen from making contact with the epithelium.
In the small intestine, a loosely formed mucus layer coats the epi­thelium,
whereas, in the large intestine, the inner mucus layer is largely devoid of
bacteria and mucus-​dwelling symbionts are restricted to the outer mucus
layer. Paneth cells located in the crypts of the small intestine constitutively
express microbicidal α-​defensins. Microfold (M) cells are specialized epi­
thelial cells that overlay lymphoid tissues, such as Peyer’s patches, and facil-
itate antigen sampling. Dendritic cells (DCs) sample antigens in the Peyer’s
patches delivered by M cells or possibly directly in the lumen through pro-
jections that penetrate the epithelial layer. Secretory IgA (SIgA) is produced
by plasma cells located in the Peyer’s patches and is secreted across the
epithelium into the gut lumen by transcytosis. The majority of SIgA is pro-
duced following Toll-​like receptor (TLR) engagement on T cells and B cells
by microbial antigens and binds with low affinity to a broad fraction of
intestinal microorganisms, preventing their translocation across the epi­
thelium. T follicular helper (TFH) cell-​dependent responses further contribute
to homeostasis by supporting the production of high-​affinity IgA , which
binds to select bacteria. Myeloid cell-​derived cytokines, including IL-1β, IL-6
and IL-23, promote T helper 17 (TH17) cell differentiation and group 3 innate
lymphoid cell (ILC3) activation. IL-22 produced by ILC3s and TH17 cells fur-
ther reinforces the segregation of symbionts by inducing the expression of
antimicrobial peptides (AMPs), including regenerating islet-​derived protein
3γ (REG3γ). IL-22 also promotes fucosyltransferase 2 (FUT2)-mediated fuco-
sylation of epithelial glycans to support microbial symbiosis and protect
against the invasion of potentially harmful bacteria. If bacteria are able to
overcome these barriers, IL-17-stimulated neutrophils can eliminate patho-
bionts localized near the apical epithelial surface or those that have reached
the lamina propria. Microbial killing by tissue-​resident macrophages and
DCs is further enhanced by TH1 cell-​derived interferon-​γ (IFNγ). The produc-
tion of bacteria-​specific IgG controls systemic infection by binding patho-
bionts to facilitate opsonization. In addition, penetrant microorganisms
are phagocytosed by Kupffer cells in the liver and splenic macrophages to
control systemic dissemination.

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Reviews
Preventing symbiont accumulation in the mucosa. The
protective mechanisms that promote the segregation
of symbionts from the epithelium are not fool-​proof.
Given the large numbers of microorganisms that reside
near the epithelial surface, some bacteria can breach the
epithelial barrier under conditions of homeostasis36–38.
To counter this situation, the host immune system has
strategies to limit damaging inflammation in the mucosa
and the dissemination of penetrant microorganisms to
systemic tissues (Fig. 1). These strategies include the
killing of bacterial symbionts by specialized lamina
propria macrophages that reside beneath the epithe-
lium at high numbers, which engulf and kill penetrant
microorganisms through several mechanisms, including
the production of antimicrobial molecules and reactive
oxygen species39. Under homeostatic conditions, these
resident macrophages are defective in microbial sens-
ing and hence do not induce inflammatory responses,
a regulatory activity that is mediated through IL-10
stimulation40–42. Intestinal microorganisms can also be
engulfed by dendritic cells (DCs) in the lamina pro-
pria and transported to the mesenteric lymph nodes,
where bacteria-​containing DCs induce protective IgA
and regulatory T (Treg) cells; these DCs do not reach sys-
temic secondary lymphoid structures, which limits their
systemic dissemination43.
Control at systemic sites. Despite the presence of robust
mucosal firewalls, rare intestinal microorganisms dis-
seminate systemically through the venous portal sys-
tem or blood vessels (Fig. 1). These microorganisms
can be engulfed and killed by Kupffer cells in the liver
or macrophages in the spleen39,44. Symbiotic bacteria
can also induce homeostatic IgG responses through
T cell-​dependent and T cell-​independent mechanisms.
IgG antibodies can bind multiple symbionts, including
Proteobacteria and related pathogens, through the rec-
ognition of highly conserved proteins, such as murein
lipoprotein, to limit the systemic dissemination of bac-
teria from the intestine38. In addition, IgG2b and IgG3
isotypes have similar reactivity to symbiotic species
that are bound by IgA and can be transferred to neo-
nates in the breast milk; these maternal IgG isotypes,
together with IgA, bind the neonatal microbiota to limit
aberrant commensal-​specific T cell-​mediated inflam-
mation45. Systemic IgG responses to the microbiota are
enhanced in animals deficient in IgA or the MYD88 and
TIR domain-​containing adaptor-​inducing interferon-​β
(TRIF) signalling adaptors for TLR-​mediated sensing of
bacteria46,47. This enhanced IgG response likely represents
a compensatory adaptation to the increased presence of
symbionts in systemic tissues as the result of impaired
containment and killing of penetrant symbiotic bacteria.
Evasion by pathogens. Enteric pathogens have evolved
multiple strategies to disrupt and evade the mucosal
firewalls that limit the penetration of bacterial sym-
bionts into the mucosal tissues. For example, several
pathogens, including Salmonella enterica, Shigella
flexneri, Yersinia enterocolitica and Vibrio cholerae,
produce mucin-​degrading enzymes48–51. In addition,
enteric pathogens have evolved resistance mechanisms
to counter antimicrobial proteins produced by IECs.
For example, S. enterica can express genes involved in
lipopolysaccharide modifications that regulate resis­
tance to antimicrobial peptides as well as genes involved
in the sequestration, efflux and degradation of anti­
microbial peptides52,53. Likewise, Listeria monocytogenes
deacetylates N-​acetylglucosamine residues in peptido-
glycan to evade the bacteriolytic activity of lysozyme,
an enzyme produced by Paneth cells54. Unlike bacterial
symbionts, S. enterica produces virulence proteins to
evade lysosomal degradation in phagocytic cells55. Upon
entry into enterocytes, S. flexneri ruptures the vacuolar
membrane and escapes into the host cytosol, where it
evades autophagy-​mediated degradation, replicates and
spreads between IECs56. Within neutrophils, S. flexneri
can inhibit reactive oxygen species-​mediated killing
by the expression of superoxide dismutases and cata-
lases57. Thus, enteric pathogens have evolved multiple
mechanisms to evade mucosal firewalls. Perhaps more
pertinent to IBD are the pathobionts that can trigger
disease in a susceptible host deficient for one or more
components of the mucosal firewall.
Breakdown of mucosal firewalls in IBD
Breakdown of the homeostatic processes that reduce
the contact between microorganisms and the epithelial
cell surface may increase susceptibility to IBD devel-
opment13,14. This notion is strengthened by the observ­
ation that multiple IBD susceptibility genes encode
proteins that function to limit the penetration of bacte-
rial symbionts into the mucosa or to promote bacterial
killing (Fig. 2).
Segregating symbionts from the epithelial surface.
In mice, deficiency of MUC2 leads to abnormalities in
the mucus layer that promote the close proximity
of the microbiota to the intestinal epithelial surface and
the development of spontaneous colitis58. The presence
of bacteria deep within crypts in close contact with the
epithelium is a common feature observed in patients
with ulcerative colitis58,59. Although aberrant expres-
sion of MUC2 has been described in patients with
ulcerative colitis60,61, variants in mucin genes have not
been associated with IBD susceptibility. However, loss-​
of-function variants in fucosyltransferase 2 (FUT2),
which encodes a protein that promotes mucosal barrier
function, are associated with increased susceptibility to
Crohn’s disease62,63.
Genetic variants of nucleotide-​binding oligomer-
ization domain 2 (NOD2) were the first gene variants
to be associated with Crohn’s disease and remain the
strongest known genetic risk factors for disease devel-
opment64,65. NOD2 is an intracellular receptor that senses
peptidoglycan-​derived muramyl dipeptide and induces
immune responses against bacteria64. NOD2 is expressed
in Paneth cells and may regulate their antimicrobial
function66,67. However, Paneth cell ablation or deficiency
in matrix metalloproteinase 7 (MMP7), which converts
inactive pro-​α-defensins to bactericidal forms, does not
lead to spontaneous inflammation in mice68. This sug-
gests that IBD pathogenesis requires additional genetic
defects or the presence of particular pathobionts that are
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 Reviews

Small intestine Large intestine

Intestinal Mucus layer

lumen Intestinal
microbiota

Goblet cell

MUC2

Pathobiont
SIgA
M cell

Epithelium Macrophage AMPs,

FUT2
DC
AMPs
Peyer’s
patch
Impaired IL-6,
bacterial IL-1β,
Paneth recognition IL-23 Defective TH17 cell
cell • NOD2
IL-22 autophagy
Neutrophil • ATG16L1 TGFβ
IL-17 • IRGM
• NOD2
IL-10
Decreased secretion of ILC3 TH17 cell Defective
anti-microbial molecules bacterial killing
by Paneth cells Impaired ILC3-dependent • CYBB
• NOD2 and TH17 cell-dependent • CYBA TH1 cell Treg cell
• ATG16L1 immunity • NCF1
• XBP1 • TNFSF15 • NCF2
• LRRK2 • IL23R • NCF4 Defective
• RAC1 immunoregulation
• RAC2 • IL10R
Lamina propria

Fig. 2 | Breakdown of mucosal firewalls in inflammatory bowel disease. Mutations in susceptibility genes for
inflammatory bowel disease can impair the mucosal strategies that are used by the host to prevent harmful
microorganisms from gaining access to the intestinal lamina propria. In the small intestine, variants in nucleotide-​binding
oligomerization domain 2 (NOD2) and autophagy-​related genes (ATG16L1, IRGM, XBP1 and LRRK2) lead to decreased
secretion of Paneth cell-​derived antimicrobial peptides (AMPs). Polymorphisms of TNFSF15 and IL23R genes can affect
group 3 innate lymphoid cells (ILC3s) and T helper 17 (TH17) cells, which segregate bacterial symbionts from the mucosa
through the production of IL-22 and IL-17. Crohn’s disease-​associated loss-​of-function NOD2 mutations in phagocytes
such as macrophages and dendritic cells (DCs) impair bacterial recognition. Mutations in autophagy-​related genes
(ATG16L1, IRGM and NOD2) in intestinal epithelial cells may result in defective bacterial clearance. In the large intestine,
genetic variants in components or regulators of the phagosomal nicotinamide adenine dinucleotide phosphate (NADPH)
oxidase complex (CYBB, CYBA, NCF1, NCF2, NCF4, RAC1 and RAC2) impair the oxidative burst and production of reactive
oxygen species, leading to defective bacterial killing in phagocytes. Loss-​of-function mutations in IL10R alter intestinal
immune homeostasis, leading to dysregulated T cell responses. FUT2, fucosyltransferase 2; M, microfold; MUC2, mucin 2;
SIgA, secretory IgA; TGFβ, transforming growth factor-​β; Treg cell, regulatory T cell.

Unfolded protein response
(UPR). A group of intracellular
signal transduction pathways
that facilitates the folding,
processing, export and
degradation of proteins
derived from the endoplasmic
reticulum during stressed
conditions.
not found in the microbiota of the vast majority of mice
housed under specific-​pathogen-free (SPF) conditions.
Mutations in the Crohn’s disease-​associated protein
autophagy-​related protein 16-like 1 (ATG16L1) may
also contribute to ileal disease by impairing the exocy-
tosis of secretory granules within Paneth cells, an activ-
ity that limits the penetration of bacterial symbionts69.
Gene variants of the unfolded protein response (UPR)
transcription factor X-​box binding protein 1 (XBP1)
have been associated with increased risk for Crohn’s
disease70. Specific epithelial deletion of XBP1 results in
endoplasmic reticulum stress and structural defects
in Paneth cells. Notably, impairment of both the UPR and
autophagy pathways in IECs is associated with spontane-
ous Crohn’s disease-​like transmural ileitis71. Two Crohn’s
disease susceptibility genes, TNFSF15 and IL23R, regu-
late ILC3s and TH17 cells, which have a crucial role in the
containment of symbiotic microorganisms through
the production of IL-17 and IL-22 (ref.72). However,
further investigation is needed to understand how
variants of these genes are linked to Crohn’s disease.
Preventing symbiont accumulation in the mucosa.
Killing of bacterial symbionts by specialized macro­
phages in the lamina propria has a key role in limiting
damaging inflammation in the mucosa and preventing

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Reviews
Very early-​onset IBD
A form of inflammatory bowel
disease (IBD) diagnosed when
symptoms manifest before the
age of 6 years; it presents with
a severe disease course,
extensive colonic involvement,
poor response to therapy and
the frequent need for
abdominal surgery.
the dissemination of penetrant microorganisms. In the Studies using gnotobiotic mice have identified bacterial
intestine, NOD2 is expressed by phagocytes, epithelial species that can induce an adaptive immune response
cells and stromal cells as well as Paneth cells. Notably, during homeostasis. Perhaps the best studied example
NOD2 deficiency together with deficiency of CYBB (also are the SFB, spore-​forming Gram-​p ositive bacteria
known as NOX2) — a component of the phagosomal related to Clostridium spp. that can adhere tightly to the
nicotinamide adenine dinucleotide phosphate (NADPH) epithelium of the terminal ileum in mice and promote
oxidase complex, which kills bacteria through the oxi- IL-17 expression by retinoic acid receptor-​related orphan
dative burst — triggers the bloom of M. schaedleri and receptor-​γt (RORγt+) CD4+ TH17 cells in the lamina pro-
spontaneous Crohn’s disease-​like TH1 cell-​driven colitis pria86. The adhesion of SFB to intestinal epithelia trig-
in mice22. Because Crohn’s disease-​associated NOD2 var- gers the epithelial release of serum amyloid A, which
iants are loss-​of-function mutations, the resulting defec- conditions local DCs to secrete the TH17 cell-​promoting
tive bacterial sensing within phagocytic cells or intestinal cytokines IL-1β and IL-23; these cytokines also enhance
and/or stromal cells may promote luminal accumula- the production of IL-22 by gut-​resident ILC3s, which
tion and mucosal penetration of pathobionts, lead- reinforces the expression of serum amyloid A by epi-
ing to T cell-​mediated intestinal inflammation (Fig. 2). thelial cells87,88 (Fig. 3). Consequently, SFB colonization
However, whether NOD2-associated Crohn’s disease confers host protection against extracellular pathogens
is caused by the accumulation of specific pathobionts but is also linked with autoimmune disorders86,89. In
in humans requires further investigation. SFB-​colonized animals, the majority of lamina propria
NOD2 also has a role in autophagy, a pathway that TH17 cells express T cell receptors (TCRs) that recognize
mediates lysosomal degradation and clearance of intra- SFB antigens90.
cellular bacteria73. NOD2 recruits the Crohn’s disease-​ In addition to epithelial adherence, the ability of
associated protein ATG16L1 to the plasma membrane at symbiotic bacteria to reside in the mucus layer near the
sites of bacterial entry74, but Crohn’s disease-​associated intestinal epithelium may also be an important behav-
NOD2 variants have defective recruitment of ATG16L1 ioural feature to influence host immunity. For example,
and impaired bacteria-​induced intraepithelial auto- intestinal colonization by the mucus-​dwelling com-
phagy75. In addition to ATG16L1, immunity-​related mensal Akkermansia muciniphila can spontaneously
GTPase family M (IRGM) and leucine-​rich repeat kinase induce an adaptive immune response during homeo-
2 (LRRK2) also regulate the autophagy pathway and var- stasis in gnotobiotic mice with a simplified microbiota.
iants of these genes have been associated with Crohn’s A. muciniphila-​specific T cells primarily differentiate
disease risk76,77. As defects in autophagy impair the clear- into T follicular helper cells to mediate an IgG1 response
ance of intracellular bacteria78,79, it is conceivable that against the bacteria but the same TCR-​transgenic CD4+
mutations in autophagy-​associated genes could result T cells can adopt multiple cell fates under SPF condi-
in pathobiont penetration and inflammation in the gut. tions, which indicates that a complex microbiome can
A clear example of the link between bacterial killing affect the localization or behaviour of this symbiont91
and susceptibility to IBD is provided in patients with (Fig. 3). Similarly, M. schaedleri, another mucus-​dwelling
very early-​onset IBD80,81, which is associated with genetic symbiont, triggers specific IgG and IgA responses that
variants in components and regulators of the phagocyte protect neonates from colitis through antibody delivery
NADPH oxidase complex82,83. Similarly, up to 40% of to the intestinal lumen during breastfeeding22.
patients with chronic granulomatous disease, a primary
immunodeficiency disorder caused by loss-​of-function Limiting inappropriate inflammation. Treg cells orig-
mutations in the NADPH oxidase components, develop inate from two distinct ontogenic lineages: thymus-
Crohn’s disease-​like colitis84,85. Collectively, these obser-
derived Treg (tTreg) cells and peripherally derived Treg
vations suggest that defects in the killing of bacterial (pTreg) cells (Fig. 4a). Most pTreg cells develop in the colon
symbionts promote the development of Crohn’s disease. extrathymically in the presence of symbiotic bacteria
as the frequency of pTreg cells in the colon is markedly
Beneficial effects of symbionts decreased in germ-free mice92–94. Treg cells have a crucial
Symbiotic bacteria promote protective immunity by role in the preservation of tissue homeo­stasis by prevent-
reinforcing the mucosal firewalls that limit the pene- ing the induction of inappropriate T cell responses to
tration of symbionts while controlling aberrant T cell microbial antigens (Fig. 4a). For example, mice lacking
responses to microorganisms. Disruption of these Treg cells or factors that are important for their regula-
mutualistic host–symbiont interactions may promote tion or function, including IL-10, transforming growth
inappropriate immune responses against the dysbiotic factor-β (TGFβ) and αVβ8 integrin, develop spontane-
microbiota in patients with IBD. ous colitis, although the precise phenotype depends on
the animal facility95–98. Consistent with the animal data,
Inducing protective immune responses. The enteric young children with mutations in the IL-10 receptor
microbiota has a crucial role in modulating host immu- develop colonic Crohn’s disease99. Furthermore, mice and
nity to establish and maintain intestinal homeostasis. humans with mutations in forkhead box P3 (FOXP3),
Germ-​free animals have several impairments in these the transcription factor required for Treg cell develop-
processes, including defective formation of mucosa-​ ment, have autoimmune disorders, including colitis100,101.
associated lymphoid tissues, incomplete TH17 cell and A role for Treg cells in suppressing immune responses
Treg cell development, and reduced numbers of intraep- to sym­biotic bacteria is also supported by the observ­
ithelial CD8αβ T cells and IgA-​producing plasma cells5. ation that microbiota-​dependent colitis induced by the
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 Reviews

Small intestine Large intestine

Intestinal
microbiota

Akkermansia Symbionts
Symbionts muciniphila

SFB SIgA
AMPs

Intestinal IL-17,
epithelium IL-22
IgG
IL-1β, Neutrophil Plasma
IL-23 TFH cell
cell
TFH cell TLR
SAA
ILC3 TH17 cell
DC
Lamina propria

Fig. 3 | Beneficial effects of symbionts. Adhesion of segmented filamentous bacteria (SFB) to the intestinal epithelium
triggers the release of serum amyloid A (SAA), which acts on dendritic cells (DCs) of the lamina propria to stimulate the
secretion of cytokines, including IL-1β and IL-23, and induce retinoic acid receptor-​related orphan receptor-​γt (RORγt)+
T helper 17 (TH17) cell differentiation and group 3 innate lymphoid cell (ILC3) activation. IL-22 secretion by ILC3s
reinforces SAA production by the epithelium to augment TH17 cell-​mediated mucosal defences, including antimicrobial
peptide (AMP) secretion and neutrophil recruitment. Symbionts also promote intestinal homeostasis through IgA
responses induced by B cell-intrinsic and T cell-intrinsic microbial sensing by Toll-​like receptors (TLRs). Secreted IgA (SIgA)
contributes to barrier function by binding to intestinal microorganisms and preventing epithelial translocation. Similarly,
IgG production can occur directly by TLR engagement on B cells, and intestinal colonization by Akkermansia muciniphila
can induce homeostatic IgG production through antigen-specific T follicular helper (TFH) cell responses. In addition, IgG
contributes to the systemic control of pathobionts that penetrate the epithelial barrier (not shown).

transfer of CD45RBhiCD4+ T cells into lymphopenic
mice is ablated by the co-​transfer of Treg cells102. Although
the selective depletion of pTreg cells by deletion of the
Foxp3 enhancer conserved non-​coding DNA sequence 1
(CNS1) in mice does not lead to the spontaneous multi­
organ autoimmunity observed in complete Foxp3-
null scurfy mice, type 2 immune pathologies can be
observed in the gastrointestinal tract103. This immuno­
pathology of the mucosal barrier in pTreg cell-deficient
animals suggests that tTreg cells are sufficient to maintain
tolerance to self-​antigens systemically, whereas pTreg cells
have a non-​redundant role in restraining inflammation
in the intestine103.
Colonic pTreg cells are profoundly influenced by local
antigens as their TCR repertoire is distinct from analo-
gous Treg cells of the peripheral lymph nodes and spleen
and can recognize antigens expressed by Clostridium and
Parabacteroides spp., which further supports that bac-
teria are important for intestinal Treg cell induction93
(Fig. 4b). A limited consortium of 17 human-derived
Clostridium spp., which can colonize the colonic mucus
layer near the epithelium, can induce Treg cells in germ-​
free mice and can suppress intestinal inflammation
when administered to mice with colitis104. Colonization
with the symbiont Bacteroides fragilis in germ-free
animals can similarly induce the differentiation of
IL-10-producing FOXP3+ pTreg cells and the B. fragilis-​
derived polysaccharide A is sufficient to reverse disease
in experimental colitis105.
Bacterial symbionts can promote the differentiation
of intestinal Treg cells by inducing the local production of
TGFβ by epithelial cells and of retinoic acid by CD103+
DCs106–108 (Fig. 4b). Secretion of IL-10 and retinoic acid
by DCs to support Treg cell proliferative expansion is
further reinforced by granulocyte–macrophage colony-​
stimulating factor (GM-​CSF) produced by ILC3s109.
Although different bacterial species can modulate
intestinal Treg cell activity, the specificity of intestinal
Treg cells for microbial antigens in the context of a com-
plex microbiota remains unclear. Bacteria-​derived short-​
chain fatty acids (SCFAs) produced by the fermentation
of dietary sugars can restore the Treg cell population in
germ-​free animals and protect against T cell-​mediated
colitis, although the mechanism remains unclear110–112.
SCFAs inhibit histone deacetylase silencing of Foxp3
gene expression, thus enhancing Treg cell differentiation
through epigenetic modifications111,112. A population of
intestinal microbiota-​inducible RORγt+ Treg cells seems
to be crucial for suppressing gut immunopathology
mediated by TH1 cells and TH17 cells or by type 2 aller-
gic responses113,114. In the T cell-​transfer model of coli-
tis, germ-​free mice colonized with an IBD-​associated

Nature Reviews | Immunology

Reviews

a b
Symbionts Bacteroides
fragilis (PSA)

Helicobacter
hepaticus

Clostridium spp.

SCFAs

TLR2
Macrophage
IL-10, IL-10, GM-CSF IL-10
TGFβ TGFβ
TH1 TH17
cell cell IL-1β IL-10,
Naive pTreg cell tTreg cell ILC3 TGFβ, Naive pTreg cell
T cell RA T cell

DC

CTLA4-mediated and/or Local symbiont antigens

TCR-mediated inhibition

Fig. 4 | Regulatory T cells support intestinal homeostasis. a | Peripherally derived regulatory T (pTreg) cells and thymus-​
derived Treg (tTreg) cells restrain aberrant inflammation in the intestine. Treg cells produce IL-10 and transforming growth
factor-​β (TGFβ) to suppress effector T helper 1 (TH1) cells and TH17 cells. In addition, myeloid cells such as macrophages
are an important target for IL-10-driven intestinal homeostasis. Intestinal Treg cells also restrain effector T cell responses
by the inhibition of antigen-​presenting cells such as dendritic cells (DCs) through cytotoxic T lymphocyte antigen 4
(CTLA4)-mediated or T cell receptor (TCR)-mediated processes. Self-​reactive tTreg cells may also contribute to antigen-​
specific immunosuppression through TCR cross-​reactivity with microbial antigens. b | Symbionts have a crucial role in the
peripheral education of pTreg cells through antigen-​dependent and antigen-​independent processes. Macrophage sensing
of bacteria leads to the secretion of granulocyte–macrophage colony-​stimulating factor (GM-​CSF) by group 3 innate
lymphoid cells (ILC3s) stimulated by IL-1β, which in turn enhances DC expression of retinoic acid (RA) and IL-10 to
support pTreg cell differentiation and proliferation in the colon. Moreover, symbionts can promote the expression of
TGFβ by epithelial cells and DCs in addition to RA and IL-10. Bacteroides fragilis can enhance Treg cell activity through
polysaccharide A (PSA) binding to Toll-​like receptor 2 (TLR2) expressed by Treg cells and DCs. Select bacterial species can
directly induce pTreg cell differentiation; these pTreg cells have TCR specificity for antigens expressed by Clostridium spp.
and Helicobacter spp., which indicates a role for symbiotic antigens in supporting Treg cell differentiation. Short-​chain fatty
acids (SCFAs) are fermentation by-​products produced by symbiotic bacteria, including Clostridium spp., that can promote
pTreg cell population expansion directly by epigenetic modification or through DCs via enhanced RA production.

microbiota have a selective bias towards the expansion of
TH17 cell populations at the expense of RORγt+ Treg cells,
which accounts for disease severity and is predictive of
human disease status115.
Disruption of beneficial symbionts in IBD
Genetic defects in patients with IBD often result in an
altered composition of the microbiota, further com-
promising the beneficial effects that some microorgan-
isms exert on host immunity. A well-​described example
of a beneficial microbial species that is decreased in
patients with ileal Crohn’s disease is Faecalibacterium
prausnitzii, a member of the Firmicutes phylum that
secretes anti-​inflammatory metabolites. A decreased
abundance of F. prausnitzii in the ileum is associated
with increased risk of postoperative recurrence of ileal
Crohn’s disease and endoscopic recurrence at 6 months116.
In addition, decreased numbers of SCFA-​producing
micro­organisms, such as Clostridium spp., are observed
in patients with IBD117. Microbiota-​derived SCFAs, in
particular butyrate, promote the development of Treg cells
and mucus production to downregulate inflammatory
signalling pathways and to strengthen the epithelial
barrier112,118. Thus, the loss of SCFA-​producing micro­
organisms may have detrimental effects in IBD. Another
example of disruption of the beneficial effects of sym­
bionts is the reduction of tryptophan metabolism, which
is a feature of the IBD microbiome119,120. The tryptophan
metabolite indoleacrylic acid, which is produced by sev-
eral Peptostreptococcus spp., promotes mucosal barrier
function and reduces inflammatory responses119. The
symbiont Peptostreptococcus russellii increases goblet cell
numbers and mucin fucosylation, and mediates protec-
tion from dextran sodium sulfate (DSS)-induced colitis119.

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Faecal stream diversion
A diverting terminal ileostomy
constructed proximally to an
ileocolonic anastomosis,
thereby excluding the
neoterminal ileum, the
anastomosis and the colon
from the faecal intestinal
transit.
Tbx21−/−Rag2−/− mice
Immunocompromised mice
that have no adaptive immune
cells and only innate immune
cells with a deficiency in T-​box
transcription factor 21
(TBX21).
16S ribosomal RNA gene
sequencing
A DNA amplification
technology for the study of a
gene conserved among
bacteria that is used for
species identification and
taxonomic classification of
bacteria.
Nature Reviews | Immunology
Thus, the microbial imbalance associated with IBD may
contribute to disruption of the regulatory mechanisms
that suppress inflammation.
Breakdown of symbiosis in IBD
A large body of evidence suggests that the microbiota
has a crucial role in triggering IBD. Furthermore, genetic
defects linked to IBD together with environmental fac-
tors can induce the accumulation and penetration of
pathobionts into the intestinal tissue, which further
promote dysbiosis and inflammation.
The microbiota as a trigger of IBD. Although no path-
ogen or pathobiont has been consistently identified
as being responsible for IBD, multiple lines of evi-
dence support an important role for the microbiota in
driving intestinal inflammation2. For example, faecal
stream diversion decreases or abolishes the inflammation
of ileal Crohn’s disease121. Furthermore, recurrence of
Crohn’s disease in the neoterminal ileum after curative
ileal resection depends on exposure to luminal con-
tents122. Antibiotic treatment can induce remission in
patients with active ulcerative colitis and Crohn’s disease
as well as prevent relapse in some patients with quiescent
Crohn’s disease123,124. However, the interpretation of anti-
biotic studies is difficult given the effects of these drugs
on multiple bacterial species.
Animal models further support a role for the gut
microbiota in the pathogenesis of IBD. For example,
oral transfer of the faecal microbiota from mice with
colitis into healthy animals is sufficient to trigger dis-
ease125–127. Most importantly, genetically susceptible mice
develop colitis with a conventional microbiota but not
under germ-​free conditions. For example, SPF mice, but
not germ-​free mice, with null mutations in TCR genes
develop colitis128. Furthermore, colitis was not observed
in germ-​f ree TCR-​deficient mice colonized with a
defined commensal community128, which suggests that
specific microorganisms are required to trigger disease.
Similarly, the intestinal inflammation that develops in
Il2–/– mice, Il10–/– mice and rats transgenic for HLA-​B27
and human β2-microglobulin is largely abrogated under
germ-​f ree conditions95,96,129,130. In addition, Tbx21 –/–
Rag2–/– mice develop ulcerative colitis-​like spontaneous
inflammation that is markedly inhibited by antibiotic
treatment125. In models of Crohn’s disease-​like ileitis,
TNF∆ARE mice, which have a deletion in the tumour
necrosis factor (TNF) AU-​rich elements (ARE), develop
TNF-​driven transmural inflammation in the presence of
the microbiota126, and SAMP1/Yit mice, a recombinant-​
inbred line, develop ileitis that is attenuated under
germ-​free conditions131. Although studies of germ-​free
animals suggest a crucial role for the microbiota in
IBD, the interpretation of these experiments is difficult
given that multiple deficiencies are present in germ-​free
mice. Studies with Nod2–/–Cybb–/– mice showed that they
develop Crohn’s disease-​like colitis when harbouring
the microbiota of mice from Taconic Biosciences but
not the microbiota of mice from the Jackson Laboratory22,
which indicates that colitis requires the presence of
particular pathobionts to trigger disease in genetically
susceptible mice.
Reviews
Dysbiosis in patients with IBD. The composition of the
microbiota in patients with IBD has been extensively
studied132. 16S ribosomal RNA gene sequencing analysis of
stool and mucosal samples has revealed the presence of a
dysbiotic microbiota, characterized by decreased com-
munity diversity and a shift in bacterial taxa, including
alterations in certain genera of the phylum Firmicutes
and increased abundance of Enterobacteriaceae spe-
cies117,133; these alterations are typically more pronounced
in patients with Crohn’s disease than in patients with
ulcerative colitis134. Some studies also showed changes
in Bacteroides spp., particularly in patients with Crohn’s
disease135. Differences in the microbiota are also present
between patients with IBD and close relatives, includ-
ing twins136,137, which suggests that dysbiosis correlates
with disease rather than with genetic factors. However,
longitudinal studies are needed to determine whether
dysbiosis precedes the onset of inflammation. The
imbalance in the microbial community towards a dys-
biotic state is more pronounced in mucosal samples
than in faecal samples collected at the time of diagnosis
in untreated patients with Crohn’s disease138. Studies of
mucosal samples also revealed a greater net loss of poten-
tially beneficial bacteria in Crohn’s disease compared
with ulcerative colitis, the correlation of certain taxa,
including Enterobacteriaceae in Crohn’s disease and
Ruminococcus spp. in ulcerative colitis, with disease
activity, and an association of certain taxa with response
to TNF therapy in Crohn’s disease134. Metagenomic
sequencing of stool samples has shown that there is a
marked separation between patients with Crohn’s dis-
ease and those with non-​IBD, whereas patients with
ulcerative colitis were more heterogeneous with a gen-
eral trend towards loss of species diversity139. These stud-
ies also found a loss of metabolite diversity in the faeces
of patients with IBD that is comparable to the loss of
species diversity139. Although metagenomic studies have
been limited so far to the analysis of stool samples, owing
to the high levels of host-​derived DNA that are present
in mucosal biopsy samples, they have provided insight
into the functional disruption of the microbiota in IBD.
Whereas most microbiome studies in IBD have
focused on the bacterial contribution to disease patho-
genesis, a few studies highlight the importance of other
microorganisms in IBD. For example, there is increased
fungal diversity in colonic biopsy samples from patients
with Crohn’s disease compared with healthy individ­
uals140. Similarly, increased fungal diversity and increased
prevalence of Candida albicans, Aspergillus clavatus and
Cryptococcus neoformans were observed in ileal mucosal
specimens and stool samples from patients with Crohn’s
disease141. Additional evidence has confirmed the pres-
ence of fungal dysbiosis in IBD, including a decreased
proportion of Saccharomyces cerevisiae and an increase
of C. albicans compared with healthy subjects142. Notably,
some studies correlated increased diversity of the fungal
microbiota with disease severity and suggested that the
Crohn’s disease environment favours fungi at the expense
of bacteria or that antibiotic treatment creates specific
niches for fungal expansion141–143. Intestinal fungi inter-
act with the host immune receptor dectin 1, which sig-
nals through the adaptor protein caspase recruitment
Reviews
Adherent and invasive
Escherichia coli
E. coli strains that can adhere
to and invade intestinal
epithelial cells.
domain-containing protein 9 (CARD9) to induce the
production of inflammatory molecules and TH17 cell
responses 144. CARD9 variants are associated with
increa­sed risk of developing IBD145. Similarly, Card9-
null mice have altered intestinal fungal communities
and increased susceptibility to chemically induced
colitis146. Furthermore, a polymorphism in the gene
encoding dectin 1 has been linked to medically refractory
ulcerative colitis144.
The intestine also harbours a large community of
viruses that may have a role in IBD pathogenesis. For
example, infection with murine norovirus induces
Paneth cell abnormalities in the presence of a mutation
of ATG16L1, a Crohn’s disease susceptibility gene147.
Furthermore, genetic variation in FUT2 has been impli-
cated in susceptibility to both norovirus infection and
Crohn’s disease62. Other viruses that may be relevant to
IBD are bacteriophages. Metagenomic sequencing of the
enteric virome in patients with IBD revealed an expan-
sion of Caudovirales bacteriophages compared with
control individuals, which did not seem to be second-
ary to changes in bacterial populations148; however, this
expansion was cohort specific and not confirmed across
validation cohorts148. Thus, more studies are needed to
understand the role of fungi and viruses in human IBD.
Cause or consequence of IBD? Although alterations in
the microbiota can occur early in IBD and independently
of therapy, there is no direct evidence for a causal role of
dysbiosis in IBD pathogenesis. In mice, chemically indu­
ced colitis and intestinal infection trigger robust changes
in the composition of the microbiota, some of which
are comparable to those observed in patients with IBD,
including expansion of Proteobacteria populations149,150.
Inflammation-induced increased oxygenation of the
intestinal lumen and increased availability of nitrate
and host-derived electron acceptors can drive anaerobic
respiration and blooming of Enterobacteriaceae149,151.
Thus, many of the microbiota alterations that are
observed in the intestinal mucosa and lumen of patients
with IBD are likely to be secondary to inflammation.
In a genetic model of spontaneous Crohn’s disease,
longitudinal analyses have revealed the population expan-
sion of a single or limited number of pathobionts before
the onset of colitis and this dysbiotic microbiota is suffi-
cient to trigger disease in mice with a complex microbi-
ota22. Based on these observations, dysbiosis may proceed
in two sequential stages. At an early stage, IBD-​associated
genetic and environmental factors may lead to the accu-
mulation of disease-​causing pathobionts, which may
precede the development of clinical disease. Although
the identity and number of pathobionts involved in
early dysbiosis are unknown, limited evidence in animal
models of IBD suggests that the genetic and metabolic
features of the bacteria may be important. For example,
M. schaedleri and Helicobacter hepaticus, two pathobionts
that can trigger spontaneous colitis in genetically suscep-
tible mice, produce virulence factors and live near the
epithelium, although the role of these activities in induc-
ing colitis remains unclear22,152,153. Such bacterial features
may lower the threshold for local penetration when the
mucosal firewalls are compromised by mutations in IBD
susceptibility genes (Fig. 5). During late dysbiosis, intes-
tinal inflammation drives further changes in bacterial
taxa, including the expansion of Proteobacteria popu-
lations (Fig. 5). Given that distinct members of the gut
microbiota have beneficial effects on the host immune
system and the intestinal barrier119,154, depletion of cer-
tain taxa may contribute to the exacerbation or persis-
tence of intestinal inflammation. Blooms of particular
bacteria, such as adherent and invasive Escherichia coli that
accumulate in the inflamed mucosa of patients with IBD,
could further promote inflammation (Fig. 5). Adherent
and invasive E. coli adhere to intestinal epithelia using
the common type 1 pili adhesin FimH, which recog-
nizes the carcinoembryonic antigen-​related cell adhesion
molecule 6 (CEACAM6) that is abnormally expressed
in the ileum of patients with Crohn’s disease155. Well-​
designed longitudinal studies of mucosal samples from
humans with increased IBD susceptibility and/or patients
with IBD before clinical relapse are needed to understand
the role of dysbiosis in disease.
Adaptive immune responses to microbial antigens in
IBD. Increased T cell and antibody responses to micro-
bial antigens are often detected in patients with IBD,
providing further evidence for a role of the microbiota
in disease pathogenesis. Patients with IBD produce large
amounts of IgG antibodies against symbiotic bacteria156
as well as having high serum reactivity against fungal
and/or bacterial peptides and glycans compared with
healthy individuals157. Although antimicrobial antibod-
ies can be used as markers of disease and diagnosis, the
role of such antibodies in IBD remains unclear. Given
that healthy mice and humans develop antibodies against
bacterial symbionts36,38,158, the presence of high titres of
IBD-​associated antibodies may reflect enhanced adaptive
immune responses or increased exposure to microbial
antigens in the inflamed gut of patients with IBD.
In addition to humoral responses, dysregulated T cell
responses against the microbiota are observed in IBD.
Early work showed that T cells or mononuclear cells from
the inflamed mucosa of patients with IBD have increased
reactivity compared with cells from normal mucosa after
stimulation with gut microbial antigens159,160. However,
CD4+ T cells reactive against bacterial symbionts can be
detected in the blood and mucosa of both patients with
IBD and healthy individuals161,162. Microbiota-​responsive
human T cells are mainly of a memory phenotype and
have a diverse TCR Vβ repertoire, which is consistent
with reactivity against a wide array of microbial anti-
gens162. In agreement with human data, T cells reactive
against symbiotic bacteria and bacterial antigens can
also be detected in animal models of colitis163. Transfer
of CD4+ T cells reactive against commensal antigens into
lymphopenic animals is sufficient to trigger colitis164–166.
Similarly, monocolonization of Il10–/– germ-​free mice
with some commensal bacteria, including E. coli and
Enterococcus faecalis or the pathobiont H. hepaticus,
elicits colitis167,168. Interestingly, H. hepaticus-​colonized
immunocompetent animals are healthy because the CD4+
TH cells that express H. hepaticus-​specific TCRs predom-
inantly differentiate into RORγt+ pTreg cells and T folli-
cular helper cells169,170. However, under IL-10-deficient
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 Reviews

a Early dysbiosis b Late dysbiosis

Genetic factors Environmental Inflammation

factors

Decreased Loss of Pathobiont

microbial beneficial expansion
diversity symbionts

Decreased abundance Expansion of

of Firmicutes Proteobacteria

Firmicute
O2– + NO NO3– NO2–
Pathobiont
accumulation
and penetration Iron O2

SCFAs
O2

Neutrophil
TH1 cell
Pathobiont
Monocyte penetration
TH17 cell
DC Treg cell

Macrophage

Early inflammatory response Chronic inflammation

Fig. 5 | Dysbiosis in inflammatory bowel disease. The inflammatory bowel disease-​associated genetic defects, together
with environmental factors such as diet and antibiotic use, can lead to the accumulation and penetration of disease-​
causing pathobionts into the intestinal lamina propria (early dysbiosis), which may precede the development of clinically
overt disease. Inflammation may result in larger alterations of bacterial taxa, including the expansion of Proteobacteria
(late dysbiosis), through enhanced luminal oxygenation and increased availability of nitrate (NO3−), host-​derived oxygen
acceptors and iron in the inflamed gut environment. This late stage of dysbiosis is also characterized by an overall
decrease in microbial diversity with a loss of beneficial symbionts, which may result in increased mucosal adherence and
translocation of symbiotic microorganisms, thus triggering chronic inflammation. DC, dendritic cell; TH cell, T helper cell;
Treg cell, regulatory T cell; SCFAs, short-​chain fatty acids.

conditions, Helicobacter spp.-specific CD4+ T cells differ-
entiate into pathogenic TH17 and TH1 cells169,170. Notably,
antigens expressed by symbionts that have limited access
to the host are not sufficient to initiate T cell activation
and colitis; these include Bacteroides spp., which prefer-
entially reside in the intestinal lumen, and CBir1 flagellin-​
expressing bacteria, which require a severe perturbation
in the mucosal barrier or selective impairment of IgA to
trigger an antigen-​specific T cell response170–173. These
observations suggest that pathogenic CD4+ TH cells
recog­nizing pathobionts can develop under homeostatic
conditions and that the breakdown of Treg cell-​mediated
immunosuppression to these pathobionts is crucial for
colitis development. However, the mechanism by which
Treg cells act on effector T cells to inhibit bacteria-induced
inflammation in the gut remains unclear. Treg cells could
inhibit effector T cells in an antigen-independent man-
ner174 and/or act on antigen-presenting cells through
their TCRs to suppress antigen-​specific effector T cells175.
Targeting the microbiota for therapy
Most current therapies for IBD, including steroids and
biologicals such as anti-​TNF or anti-​integrin therapies,
suppress the host immune system but do not directly
target the microorganisms that cause or contribute to
inflammation. Given that IBD therapies induce com-
plete remission in less than 50% of patients, the devel-
opment of therapeutic approaches that target intestinal
microorganisms may provide a unique approach to treat
IBD. Our increased understanding of dysbiosis and the
immunomodulatory functions of select microorganisms
has presented several innovative strategies for modify-
ing the intestinal microbiota to prevent or ameliorate
IBD (Fig. 6).
Faecal microbiota transplantation. Features of dys­
biosis in IBD and recurrent Clostridium difficile infec-
tion include the loss of phylum-​level diversity and
the overgrowth of facultative anaerobic bacteria of the

Nature Reviews | Immunology

Reviews

Faecal microbiota
transplantation
(FMT). The transfer of the
intestinal microbial community
from a healthy individual to a
patient through infusion of
stool, typically by endoscopy.
Enterobacteriaceae family, including Proteobacteria5.
Therefore, approaches aimed at correcting imbal-
ances in the microbial community may be effective
for the treatment of IBD. The high success rate of
faecal microbiota transplantation (FMT) in resolving
recurrent C. difficile infection by restoring gut micro-
bial community diversity has reinforced the potential
of microbiota-​modulating therapies176. Although the
number of patients with IBD who have been treated
with FMT is limited and the responses to treatment
have been variable, induction of clinical remission has
been achieved in about 30% of patients with ulcerative
colitis177,178. Pretreatment with antibiotics and the use
of multiple faecal infusions seem to improve the effec-
tiveness of FMT in patients with ulcerative colitis177,178.
However, one double-​blinded, randomized, placebo-​
controlled study did not observe any beneficial effect of
FMT in patients with ulcerative colitis179. In addition,
the effectiveness of FMT in patients with Crohn’s disease
remains unclear as current studies reporting potential
efficacy are limited in cohort size and lack a placebo
treatment group180,181. Further studies are needed to
understand the mechanism by which FMT is effective
in some patients with ulcerative colitis and to develop
more effective protocols to treat IBD.
Antibiotics and probiotics. Antibiotic usage in the treat-
ment of IBD remains limited as a result of conflicting
findings but studies using broad-​spectrum antibiotic
cocktails have shown that they may be efficacious in
cases of severe acute colitis and chronic ulcerative coli-
tis182. Meta-​analyses of randomized controlled trials also
indicate the benefits of antibiotics in the treatment of
Crohn’s disease and ulcerative colitis183,184. However, the
use of antibiotics to modify the microbiota is limited
by the inability to selectively eliminate disease-​causing
bacteria without potentially affecting beneficial micro-
organisms, which could lead to unexpected adverse
outcomes. In the future, once the metabolic pathways
of IBD-​causing microorganisms have been identified,

FMT Antibiotics Probiotics Prebiotics Drugs Synthetic Diet

microorganisms
Gut
lumen
Direct Direct Indirect Direct Direct Indirect Direct
• Metabolites Pathobiont Population Population Target Host Shapes
• Bacteriophages killing expansion of expansion of metabolic immune microbiota
• Beneficial beneficial Direct beneficial pathways modulation composition
microorganisms microorganisms Microcins microorgansims

Indirect Symbionts
Host Indirect
immune Host
stimulation immune
modulation
Pathobionts SCFAs
Mucus Indirect
layer MAMPs Bacterial
competition

Epithelial
barrier

TLR

IL-10
Macrophage Naive Treg cell
Lamina T cell
propria

TH17 cell
DC

Fig. 6 | Effects of microbiome-​based therapeutics in inflammatory bowel
disease. Given the crucial role of the gut microbiota in the pathogenesis of
inflammatory bowel disease, the development of therapeutic approaches
that target intestinal microorganisms may provide a unique approach to
treat the disease. Faecal microbiota transplantation (FMT) may have direct
beneficial effects through the transfer of bacteria-​derived metabolites and
bacteriophages or by restoring beneficial microorganisms, or indirect
beneficial effects through host innate immune stimulation by bacterial
components (for example, Toll-​like receptor (TLR) triggering by
microorganism-​associated molecular patterns (MAMPs)). Antibiotics may
have broad effects by directly killing pathobionts or indirectly promoting the
population expansion of beneficial microorganisms. Administration of
probiotics, such as Escherichia coli Nissle 1917, can result in the secretion
of microcins, small polypeptides with antimicrobial activity that directly
suppress closely related Enterobacteriaceae. Supplementation with
prebiotics can foster the population expansion of beneficial symbionts that
in turn outcompete harmful bacteria. Targeting of metabolic pathways used
by bacteria to promote intestinal inflammation may provide a new strategy
to treat inflammatory bowel disease. Administration of synthetically
engineered microorganisms may have beneficial effects on the host through
modulation of the immune system, for example, by secretion of IL-10. Diet
shapes the composition of the gut microbiota by promoting the growth of
beneficial microorganisms or the depletion of pathobionts. Short-​chain fatty
acids (SCFAs) derived from dietary polysaccharides can also support the
differentiation and expansion of intestinal regulatory T (Treg) cell populations.
DC, dendritic cell; TH17 cell, T helper 17 cell.

www.nature.com/nri

it may be possible to specifically target them with a new
generation of antibiotics.
Alternatively, administering live microorganisms
may be advantageous in several ways. Once stable col-
onization is achieved, beneficial microbial factors can
be delivered continuously to the host and live bacteria
may trigger beneficial immune responses. The targeted
modulation of the microbiota by administering probi-
otics, including Lactobacillus spp. or Bifidobacterium
spp., has been successful for some bowel disorders185. In
addition, oral treatment with the probiotic E. coli Nissle
1917 has similar efficacy to the standard treatment with
mesalazine in terms of maintaining disease remission
in patients with ulcerative colitis186. Probiotic E. coli
Nissle 1917 can secrete microcins with antimicrobial
activity to suppress competing Enterobacteriaceae that
can potentially exacerbate intestinal inflammation187.
However, probiotics typically have limited effects on the
overall composition of the microbiome as these bacte-
rial species are unable to persistently colonize healthy
adults188. Concurrent supplementation with prebiot-
ics can improve the engraftment efficiency of exoge-
nous microorganisms in some indications, including
enhanced colonization resistance against C. difficile189.
Advances in synthetic biology have provided inno-
vative approaches for the treatment and management
of IBD (Fig. 6). Oral administration of Lactococcus lactis
engineered to express and secrete IL-10 was sufficient
to protect mice from colitis induced by DSS and IL-10
deficiency190. In a small cohort of patients with Crohn’s
disease, the recombinant IL-10-producing L. lactis strain
was safely administered using a thymine-​limiting con-
tainment strategy but it induced only a small improve-
ment in disease activity191. Owing to the inert properties
of L. lactis, this probiotic has also been modified to
express insulin growth factor 1, haem oxygenase 1 or
serine protease inhibitors; when orally administered to
mice, these synthetic probiotics can alleviate the devel-
opment of experimental colitis192–194. However, such
strategies were not successful in clinical trials, perhaps
owing to the inability of the engineered L. lactis strains
to persistently colonize patients with IBD195.
Targeting microbial metabolism. Precise targeting
of the metabolic pathways that are used by harmful
microorganisms to proliferate during dysbiosis has
been effective in ameliorating colitis in mice. For exam-
ple, Proteobacteria take advantage of increased nitrogen
sources, including nitric oxide, produced in the inflamed
gut to bloom during IBD. Notably, targeting the molyb-
denum cofactor-​dependent enzymes that are required to
use nitric oxide for anaerobic respiration protects mice
from DSS-​induced colitis by blunting the expansion of
Enterobacteriaceae, including Proteobacteria196. Thus,
the development of drugs that target metabolic pathways
used by harmful bacteria may provide a new strategy to
treat IBD.
Diet. Diet has an important role in shaping the compo-
sition of the microbiota and can be modulated for the
management of IBD symptoms. Exclusive enteral nutri-
tion (EEN), which is a nutritionally complete elemental
Nature Reviews | Immunology
Reviews
and polymeric formula diet that contains no solid food,
is one of the few dietary interventions that has been
extensively studied in IBD197. EEN is as effective as cor-
ticosteroids in inducing remission in paediatric patients
with Crohn’s disease, but without the side effects that
are associated with corticosteroid therapy198–200. EEN
rapidly alters the microbiota composition independently
of other environmental factors and effectively reduces
intestinal inflammation in paediatric patients with
Crohn’s disease 201. The mechanism by which EEN
induces remission in Crohn’s disease is unclear but it
may promote the growth of beneficial microorganisms
or the depletion of pathobionts. In experimental DSS-​
induced colitis, a survey of various refined diets identi-
fied the beneficial effects of psyllium fibre, whereas an
increase in dietary proteins, including casein, increased
faecal microbial density and exacerbated colitis sever-
ity through alterations in intestinal permeability202.
Importantly, host susceptibility to colitis is highly
dependent on combinations of specific fibre or protein
components that shape the microbiota, which indicates
the importance of dietary composition in IBD202.
Future directions
In the past decade, there has been an enormous increase
in knowledge regarding microbiota–host interactions
as well as the role of genetics and the immune system
in IBD. However, several aspects of our understanding
of IBD pathogenesis remain unclear.
A major deficiency is the lack of knowledge about the
identity of IBD-​causing pathobionts that trigger inflam-
mation in genetically susceptible individuals. Multiple
species of pathobionts may cause disease and these may
vary according to the specific IBD susceptibility loci
of a patient. Therefore, studies using individuals with
identical or similar genetic defects may be important to
identify such microorganisms.
There is a large intra-​species genetic diversity in the
symbionts that live in the mammalian intestine. Carefully
designed longitudinal studies coupled to metagenomic
analyses of mucosal samples and to new methodologies
that can reveal intra-​species variation may be impor-
tant to understand the role of dysbiosis in disease. The
identification of IBD-​causing microorganisms will be
important for understanding disease pathogenesis, mon-
itoring patients for alterations of these pathobionts and
the rational development of new therapies.
Animal models of IBD have provided important
insights into the roles of the microbiota and the immune
system in the development of disease. However, with few
exceptions, such models are not based on genetic defects
linked to human disease and do not fully recapitulate the
pathology of IBD. Furthermore, animal studies often rely
on the use of gnotobiotic mice with a simplified micro-
biota that does not reflect the complex microbial–host
interactions that occur in the intact gut. In addition,
mouse models do not incorporate the genetic and micro-
bial diversity of human populations or their environ-
mental exposures. Thus, it will be important to develop
new models that are more relevant to human IBD.
Published online xx xx xxxx
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Acknowledgements
The authors apologize to colleagues whose work was not
cited or was cited through other review articles because of
space limitations. The authors thank G. Chen for critical
review of the manuscript. Work in G.N.’s laboratory is sup-
ported by US National Institutes of Health grants. R.C. is
supported by a Career Developments Award from the Crohn’s
and Colitis Foundation. B.C.L. is supported by a Canadian
Institutes of Health Research (CIHR) Fellowship.
Author contributions
The authors contributed equally to all aspects of the article.
Competing interests
The authors declare no competing interests.
Peer review information
Nature Reviews Immunology thanks J. Faith, J. Round and
the other, anonymous, reviewer(s) for their contribution to the
peer review of this work.
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