Research progress concerning the involvement of the intestinal microbiota in the occurrence and development of inflammatory bowel disease

Abstract

Inflammatory bowel disease (IBD), a chronic disorder characterized by intestinal inflammation and mucosal damage, includes mainly Crohn’s disease and ulcerative colitis. However, the cause of its onset remains unclear. The pathogenesis of IBD is closely related to host genetic susceptibility, disorders of the intestinal flora, damage to the intestinal mucosal barrier, and abnormal intestinal mucosal immunity. On the basis of the progress in research on the structure of the intestinal microbiota involved in IBD, the influence of genetics on the intestinal barrier and intestinal microbiota; the metagenomics, metatranscriptomics, and metabolomics of the intestinal microbiota involved in IBD; and treatments such as probiotics and fecal microbiota transplantation are important for the future treatment of IBD and the development of drugs for effective treatment.

Key Words: Inflammatory bowel disease; Intestinal microbiota; Intestinal barrier; Metagenomics; Metabolomics

Core Tip: The etiology of inflammatory bowel disease (IBD) is complex and involves multiple factors working together, including genetic, environmental, dietary, immune and neurological factors. These factors are interrelated through a complex gut microbiome composed of bacteria, fungi, archaea, viruses and protozoa. Gut microbiota play a core role in IBD, such as genetic and serum markers related to host-gut microbiota interactions, alterations in gut microbiota in first-degree relatives, and the dynamic recovery of gut microbiota that accompanies the remission of inflammation. In fact, the rising incidence of IBD worldwide is closely related to changes in a series of environmental factors that typically affect gut microbiota. In recent years, advancements in multi-omics technologies such as metagenomics, metatranscriptomics and metabolomics, as well as their application in mouse models, human, fungal and bacterial cells, have further deepened our understanding of the role of gut microbiota in IBD.

INTRODUCTION

Inflammatory bowel disease (IBD) is a specific intestinal inflammatory disorder that involves the ileum, colon and rectum; IBD includes Crohn’s disease (CD) and ulcerative colitis (UC). The cause of IBD is still unclear. Clinically, it often presents with symptoms such as abdominal pain, diarrhea, stools that contain mucus and pus, and bloody stools, all of which seriously affect the quality of life of patients. The incidence rate is relatively high in developed countries in Europe and America. For instance, the incidence rate of IBD in Denmark increased from 11.5 per 100000 people in 1978 to 51.3 per 100000 people in 2014, and has tended to stabilize in recent years. At present, the incidence rate of IBD in China is approximately 1.96-3.14 per 100000, showing an upward trend[1]. The exact causes of IBD remain unresolved. Genetic factors, immune factors, impairment of intestinal barrier function and environmental factors are involved in the occurrence and development of IBD[2].

IBD is usually treated with medication. The main classes of drugs used in the treatment of IBD include salicylates, glucocorticoids and immunosuppressants. Salicylic acid drugs such as 5-aminosalicylic acid exert anti-inflammatory effects by reducing the release of proinflammatory factors in the intestine. They are often applied in clinical practice for patients with mild to moderate symptoms of enteritis. However, the adverse effects of salicylic acid derivatives on the gastrointestinal tract are relatively serious, and patients may suffer from kidney damage after being treated with these drugs. Glucocorticoids such as hydrocortisone and prednisone relieve inflammation by inhibiting the release of inflammatory substances such as prostaglandins and leukotrienes. They are often employed in clinical practice for patients with moderate to severe symptoms of enteritis. However, long-term or excessive use of glucocorticoids can lead to drug resistance and various adverse reactions. Immunosuppressants such as azathioprine and tumor necrosis factor-α (TNF-α) inhibitors inhibit the proliferation of inflammatory cells. They are mainly used in clinical practice to induce or maintain remission of hormone-dependent CD, which is difficult to treat clinically. However, patients are susceptible to adverse reactions such as nausea, vomiting and diarrhea after taking them[3]. In conclusion, although salicylic acid drugs, hormones and immunosuppressants can relieve the clinical symptoms of IBD, all of these drugs exhibit shortcomings, such as frequent recurrence of symptoms after discontinuation of the drug and low patient compliance with long-term use.

Research has shown that changes in intestinal microecology are involved in the onset and progression of IBD[4]. The intestinal microecology encompasses intestinal microorganisms, intestinal epithelial cells, and immune cells. Among these, intestinal microorganisms play the most important role in shaping the microecology of the intestine. These microorganisms, which consist primarily of bacteria, viruses, fungi and parasites, are distributed on the surface of the intestinal lumen. The number of bacteria present in the intestinal environment is approximately 10¹⁴, approximately ten times the number of cells in the human body. The total mass of intestinal bacteria is approximately 0.2 kg, and they account for 60% of the dry weight of feces. There are more than 50 types of bacteria, representing approximately 1100 species. The majority comprises Bacteroides and Firmicutes (90%), while a small portion comprises Actinobacillus and Proteus[5]. Multiple factors influence the composition of the human intestinal microbiota. The environment to which the fetus and the newborn are exposed directly affects the intestinal microbiota; the birth canal during delivery, the diet provided during infancy, antibiotic use, contact with pets, sex, and the mother’s health are all related to the composition of the intestinal microbiota of infants. In infants younger than one year, the diversity of the intestinal microbiota increases rapidly; diversity then begins to stabilize until the age of three and becomes more stable by the age of five, at which time the intestinal microbiota comprises mainly of Bacteroides[6]. During adulthood, exposure to various environmental factors such as smoking, air pollution, and stress, as well as hygiene habits, diet, and medication, can alter the composition of the intestinal microbiota.

The intestinal microbiota interact with the body in a way that aids in food digestion and infection resistance; they also participate directly in shaping inherent and acquired immune responses that occur in the intestinal mucosa, and they block pathogenic substances from invading the human body. Therefore, they are also known as the second-largest immune system in the human body. Dysbiosis of the intestinal flora leads to impaired microbial barrier function, invasion by pathogenic microorganisms and inflammation, resulting in intestinal and systemic disease, including inflammatory responses, tumors, and metabolic abnormalities.

INFLUENCE OF THE INTESTINAL MICROBIOTA ON INTESTINAL BARRIER FUNCTION

Intestinal microbiota and the intestinal mucosal barrier

The intestinal mucosal barrier comprises a mucus layer that is secreted by intestinal epithelial cells such as Paneth cells and goblet cells and contains symbiotic bacteria. It serves as the first line of defense against pathogenic bacteria. However, the homeostasis between the intestinal microbiota and the mucus layer is easily disrupted by external factors such as invasion by pathogenic bacteria that adhere to epithelial cells, leading to changes in the function of the intestinal mucosal barrier[7]. On the one hand, the metabolic products of the intestinal flora play a significant role in maintaining the normal physiological activities of cells. For instance, short-chain fatty acids (SCFAs) serve as energy sources for intestinal epithelial cells. They can induce the expression of the antimicrobial peptide LL37 (cathelicidin LL37), induce the differentiation and proliferation of regulatory T cells (Tregs), activate the G protein-coupled receptors 43 (GPR43) and 109a (GPR109a), and stimulate activation of the nucleotide-binding oligomerization domain of NOD-like receptor protein 3 (NLRP3), leading to resistance to pathogen infection, causing stress injury and enhancing intestinal barrier function; bile acids (BAs) promote the proliferation of intestinal epithelial cells and reduce intestinal permeability by upregulating the expression of the G protein-coupled BA receptor 1.5 (Takeda G protein-coupled receptor 5)[8]. Wrzosek et al[9] reported that Bacteroides thetaiotaomicron and Faecobacter prausnitzii have beneficial effects on the thickness and composition of the mucus layer through the promotion of goblet cell differentiation and regulation of the expression of mucin-related genes. The intestinal probiotic Akkermansia muciniphila uses mucin as a carbon and nitrogen source for growth; in this way, it can effectively regulate the thickness of the intestinal mucus layer and thereby maintain intestinal barrier function. Nakajima et al[10] reported that feeding mice soluble high-fiber feed increased the abundance of Bacteroides fragilis in the intestinal tract and that Bacteroides fragilis promoted the expression of immunoglobulin A (IgA). The polysaccharide A (PSA) produced promotes secretion of interleukin (IL)-10 and differentiation of Tregs through Toll-like receptor 2 (TLR2), thereby slowing the occurrence of inflammation. Research reported that Faecobacter prausnitzii secretes microbial anti-inflammatory molecules that inhibit the nuclear factor-kappa B signaling pathway and in this way inhibits inflammation. In contrast, when the composition of the intestinal flora is disrupted, pathogenic bacteria can multiply and colonize the intestines, causing damage to the intestinal mucosal barrier as well as inflammation[10]. For example, in IBD patients, the abundance of adherent invasive Escherichia coli (AIEC) increases; these bacteria adhere to and invade colonic epithelial cells and promote the release of proinflammatory factors. Sicard et al[11] reported the presence of Clostridium difficile, Escherichia coli, Listeria monocytogenes, and Salmonella. Both Salmonella typhimurium and Vibrio cholerae can adhere to intestinal mucin, thereby disrupting tight junctions between epithelial cells and the intestinal mucus barrier.

Intestinal microbiota and the intestinal immune barrier

The regulation of intestinal mucosal immune function is related to the presence of various intestinal microorganisms. There are more than 10¹⁴ (more than 1000 types) microorganisms in the gastrointestinal tract, and these are located mainly in the colon. Most intestinal bacteria are difficult to cultivate in vitro. The proportion of thick-walled bacteria (gram-positive bacteria) and Bacteroides (gram-negative bacteria) in the intestinal flora exceeds 90%, whereas Proteus, actinomycetes, viruses, protists and fungi are rare[12]. The gut microbiota, as a target of inflammatory responses, notably influence host immune response. Schaubeck et al[13] transplanted feces from UC model mice into healthy mice, which developed UC symptoms. However, germ-free mice that are genetically susceptible to IBD do not spontaneously develop enteritis when they are raised in a sterile environment[14], indicating that the intestinal flora constitute an important factor in the occurrence of IBD. Deletion of the NOD2 or RIP2 genes can lead to the formation of an inflammatory microenvironment in the intestine, intensify abnormal proliferation of intestinal epithelial cells, and cause damage that leads to colitis. Congenital immune deficiency can also cause changes in the intestinal microbiota. For instance, when the colons of wild-type mice are colonized with the fecal microbiota of NOD2^-/- mice, the risk of enteritis in the wild-type mice increases. However, when NOD2^-/- mice receive microbiota transplantation from healthy mice, the risk of disease is reduced, indicating that deletion of the NOD2 gene alters the composition of the intestinal microbiota in mice[15]. The NLRP6 inflammasome is a complex that is composed of multiple proteins and can serve as a sensor for endogenous or exogenous injury-related molecular signals. Elinav et al[16] reported that NLRP6 deficiency in the colonic epithelial cells of mice led to a decrease in IL-18 levels and that this in turn caused abnormal proliferation of intestinal cells. The increase in the abundance of members of the family Prevotellaceae of the phylum Bacteroidetes in the intestine may trigger the promotion of dextran sulfate sodium-induced colitis symptoms in mice. T helper 1 (Th1) and Th2 cells can be activated by the antigenic characteristics of intestinal bacteria to cause intestinal inflammation, but they are ineffective when activated nonspecifically[17]. The presence of IgA in the intestinal mucosa prevents pathogenic bacteria from invading intestinal epithelial cells and helps maintain a normal intestinal environment. However, the IgA in the mucosa can be degraded by Sutterella. An increase in the abundance of Sutterella in the intestine leads to a decrease in the amount of IgA in the intestinal mucosa, making it easier for pathogens to invade intestinal epithelial cells. Sartre can also facilitate the activation of pattern recognition receptors, resulting in the production of IL-8 and causing inflammatory responses[18]. Schirmer et al[19] reported that Bacteroides regulates the expression of antimicrobial peptides in intestinal epithelial cells by activating TLRs on Paneth cells, thereby influencing the host’s innate immune capacity.

INTESTINAL MICROBIAL ANTIGENS ARE INVOLVED IN IMMUNE REGULATION IN INTESTINAL MUCOSAL TISSUES

Microbial exposure during childhood is beneficial for the development of helper T cells in the human body, especially in the establishment of a Th2-type immune response in the intestine. An imbalance in immune homeostasis caused by insufficient microbial exposure is considered one of the causes of IBD. Epidemiological investigations have revealed that insufficient exposure to microbiota during childhood is associated with the occurrence of immune-related diseases in adulthood; this is referred to as the hygiene hypothesis[20]. The mechanism underlying this proposed relationship may relate to the fact that the gut microbiota can inhibit excessive IgE production; however, this hypothesis is still controversial. Animal experiments revealed that when CD4+CD45RB-high T cells from germ-free mice were transplanted into Rag-1^-/- mice, colitis did not occur, indicating that the intestinal microbiota are involved in the activation of pathogenic T cells and the induction of colitis. After oral administration of antibiotics to mice for 7 days, macrophages in the intestine were abnormally activated, and this affected intestinal mucosal immune homeostasis, especially the immune response regulated by T cells[21]. These findings show that intestinal bacterial antigens exert diverse effects on local immune regulation in the intestinal mucosa.

Some bacterial antigens cause damage to the immune system or provide protective effects by inducing differentiation of Th1/Th17/Treg cells. Transplantation of the inherent Bacteroides fragilis present in the intestine into the intestines of germ-free mice can induce the differentiation of CD4+ T cells in the spleen into Th1 cells[21]. The PSA produced by Bacteroides fragilis is a prerequisite for inducing the differentiation of Th1 cells and can also serve as a ligand for TLR2 on the surface of CD4+ T cells, thereby participating in the activation of CD4+ T cells. Segmented filamentous bacteria can induce the differentiation of Th17 cells in the intestinal tract. When segmented filamentous bacteria infiltrate the mucus layer and come into contact with intestinal epithelial cells, they can increase the expression of Th17-related genes in the epithelial cells, causing proliferation and differentiation of the Th17 cells. Other intestinal symbiotic bacteria, such as Schaedler flora (altered Schaedler flora), can also stimulate differentiation of intestinal Th17 cells. Clostridium difficile, which resides naturally in the intestinal tract, can induce the differentiation of intestinal CD4+ T cells into Tregs[22,23]. Some other intestinal symbiotic bacteria, such as Fusobacterium nucleatum and Streptococcus pneumoniae, can also induce the differentiation of CD4+ T cells in the intestine into Tregs. The mechanism through which bacterial antigens induce Treg differentiation may involve the production of IL-10 by intestinal CX3CR1+ phagocytes induced by contact with antigens present on symbiotic bacteria[24].

Under normal physiological conditions, intestinal symbiotic bacteria and intestinal immune cells interact with each other and maintain a balanced immune state, thereby keeping the internal environment stable. For example, PSA in the outer membrane vesicles of Bacteroides fragilis in the intestinal tract can activate dendritic cells (DCs) and thereby induce Tregs in the intestinal mucosal tissue to release IL-10, which exerts an anti-inflammatory effect. This process depends on the autophagy-related proteins ATG16 L1 and NOD2[25]. In CD patients with defects in the ATG16 L1 or NOD2 genes, DCs in intestinal mucosal tissue are unable to recognize outer membrane vesicles, resulting in reduced differentiation of Tregs and decreased secretion of IL-10. In addition, intestinal symbiotic bacteria maintain the stability of the intestinal innate immune response by restricting colonization and overgrowth by pathogenic bacteria.

REGULATION OF INTESTINAL IMMUNE RESPONSES BY METABOLIC PRODUCTS OF INTESTINAL MICROORGANISMS

The regulatory effect of SCFAs, which are metabolites produced by intestinal microorganisms, on intestinal immune homeostasis has received increasing attention. SCFAs are produced by intestinal bacteria that break down food fiber. The main SCFAs that play a role in shaping intestinal immunity are acetic acid, propionic acid and butyric acid. They regulate the differentiation, proliferation and exocrine activity of immune cells by binding to GPR41/GRP43/GPR109 on the surfaces of the immune cells. For instance, in B cells, SCFAs can promote the production of acetyl-CoA and regulate metabolic receptors, thereby increasing antibody production and inducing the differentiation of B cells into plasma cells[26]. Research determined that in GPR43^-/- mice, the secretion of secretory IgA (sIgA), the binding of bacteria to the intestinal lumen, and the number of IgA + B cells in the intestine decreased. When wild-type mice were fed acetic acid, the levels of sIgA and intestinal IgA + B cells in the feces increased, whereas feeding of acetic acid to GPR43^-/- mice did not change those levels[27]. These findings indicate that the SCFA acetic acid induces intestinal B cells to secrete sIgA through the activation of GPR43. Another group of studies revealed that SCFAs induced the expression of IL-10 in CD4+ T cells in intestinal mucosal tissue, whereas in GPR43^-/-CD4+ T cells its expression decreased, leading to an exacerbated inflammatory response in the colon. These findings indicate that SCFAs regulate T-cell differentiation and promote IL-10 production through GPR43 and thereby fulfill a protective role against colitis[28].

The indole metabolites produced by intestinal bacteria during the decomposition of tryptophan (Trp), including indole, indoleacetic acid, indolepropionic acid and indoleacetaldehyde, play an immunomodulatory role in maintaining the homeostasis of the intestinal internal environment. An analysis of intestinal microbiota metabolites in IBD patients revealed that reduced levels of Trp metabolites (indole, indolepropionic acid, and indoleacetaldehyde) can lead to a decrease in the biological activity of aromatic hydrocarbon receptor (AHR), and that this damages the intestinal inflammatory response. Indole metabolites induce the expression of IL-10R1 in intestinal epithelial cells and enhance intestinal mucosal barrier function, whereas Escherichia coli bearing mutations in the gene that encodes Trp kinase fails to induce IL-10R1 expression in intestinal epithelial cells. Subsequent animal studies revealed that intragastric administration of indole metabolites activates the IL-10 signaling pathway, thereby alleviating colitis. Transplantation of intestinal feces from CARD9^-/- mice into germ-free mice promoted the occurrence of enteritis; the mechanism through which this occurs might be related to the reduced indole content of the intestinal feces of CARD9^-/- mice[29].

STRUCTURAL CHANGES IN THE INTESTINAL MICROBIOTA IN PATIENTS WITH IBD

The composition of the intestinal flora of IBD patients differs significantly from that of healthy people, and this is manifested primarily as a decrease in the abundance of beneficial bacteria and an increase in the abundance of pathogenic bacteria[30]. Morgan et al[31] conducted gene sequencing of the microbiota found in colon tissue and fecal samples from 196 IBD patients (121 CD patients and 75 UC patients) and 27 healthy individuals. The results of the analysis revealed that in UC and CD patients, the abundance of Bordetella, Faecalibacterium prausnitzii, Odoribacter and Phascolarctobacterium was significantly lower, whereas the abundance of Clostridium was significantly higher. In patients with CD, the abundance of Ruminococcaceae was lower, whereas that of Enterobacteriaceae was higher; in particular, the abundance of Escherichia-Shigella, members of the family Enterobacteriaceae, was higher. The abundance of Leuconostocaceae is lower in patients with UC. A reduction in the number of beneficial bacteria leads to a significant decrease in the content of beneficial metabolites such as SCFAs. As important energy sources for intestinal epithelial cells, SCFAs play crucial roles in maintaining the physiological functions of these cells. For instance, Rosacea bacteria can produce acetic acid and propionic acid, rumen bacteria and Candida propioniae can produce acetic acid, Faecobacter propioniae can produce butyric acid[32], and Odoribacters planchnicus can produce acetic acid, propionic acid and butyric acid[32]. An increase in the abundance of pathogenic bacteria can cause excessive inflammatory responses, leading to intestinal damage. For example, Escherichia coli can activate TLR4 and thereby trigger inflammatory responses[33]. Another study revealed that the abundance levels of Roseburia hominis, Dorea formicigenerans and Ruminococcus obeum in the intestines of IBD patients were significantly reduced. Intestinal Ruminococcus gnavus, Escherichia coli and Clostridium clostridioforme were significantly enriched in patients with CD, and Bifidobacterium breve and Clostridium symbiosum were significantly enriched in the intestines of UC patients (Table 1)[34]. Hall et al[35] reported that the abundance of Noctuid pests in IBD patients had increased, leading to significant upregulation of the activity of intestinal oxidative stress-related gene pathways. Alam et al[36] sequenced the organisms found in the feces of 20 IBD patients (11 CD patients and 9 UC patients) and 10 healthy volunteers. They found that the abundances of Burkholderiaceae (members of the phylum Proteobacteria) and Coriobacteriaceae (members of the phylum Actinomycetes) were greater in the patients with IBD than in the healthy individuals. The abundance levels of Firmicutes and Actinobacteria were significantly greater in UC patients than in CD patients. The abundance of Proteobacteria was approximately 3.8-fold higher in CD patients than in healthy people, while in UC patients, it was decreased by approximately one-third. In patients with CD, the abundance of Bacteroidetes was approximately 1/5 of that in the healthy volunteers, while patients with UC exhibited no significant difference from the healthy volunteers in the abundance of Bacteroidetes.

Table 1 Major intestinal flora.

Phyla	Genus	Number
Bacteroidetes	Bacteroides, Porphyromonas, and Puccinia	More, accounting for more than 90%
Firmicutes	Clostridium, Clostridium tenella, Eubacterium, Rumen coccus and Lactobacillus
Actinobacteria	Bifidobacterium and gas-producing Collins	Less quantity
Proteobacteria	Enterobacteriaceae bacteria, Helicobacter pylori
Verrucomicrobia	Verrucomicrobium

Another retrospective study revealed that the use of antibiotics during pregnancy and childhood significantly increases the risk of very early onset of IBD, indicating that changes in the gut microbiota are associated with the occurrence of IBD[37]. Analysis of the composition of the intestinal flora in IBD patients revealed significant changes in the flora, with a decrease in diversity (by 50% in CD and by 30% in UC) and an increase in bacterial instability. Among the differences, the abundance levels of actinomycetes and Proteus increased, especially the abundance of Enterobacterium, which are strongly adherent and invasive, whereas the abundance levels of Bacteroides and thick-walled bacteria decreased[38]. In particular, the abundance of Lactobacillus, which produces SCFAs, decreased significantly. Further analysis of the fecal microbiota of CD patients revealed a significant increase in the number of Enterobacteriaceae. Additionally, in subsequent animal experiments, it was found that using tungstate to inhibit the excessive growth of Enterobacteriaceae could alleviate the inflammatory response. Another study revealed that the abundance of Faecalibacterium prausnitzii in the feces of CD patients was significantly reduced. This bacterium is associated with disease recurrence. Researchers have transplanted Faecalibacterium prausnitzii into the intestines of experimental mice with chronic colitis and found that this can alleviate colonic inflammatory responses[39]. In a subsequent study, researchers treated mice with experimentally induced colitis involving Lactobacillus faecalis expressing anti-inflammatory molecules; they reported that the inflammatory response was significantly reduced, suggesting that the anti-inflammatory substances secreted by Enterococcus faecalis can regulate intestinal inflammatory responses[40].

The numbers of Bacteroides and Candida in the feces of UC patients increased significantly. Another analysis of the microbiota of UC patients revealed a reduction in the abundance of Clostridium butyricum, which produces butyric acid. Butyric acid may play an important protective role in the occurrence of IBD[41]. Fecal metabolomics studies have shown that when the number of symbiotic Escherichia coli in the intestinal tract increases, their respiration and formate oxidation lead to the occurrence of dysbiosis and intestinal inflammatory responses. It is generally considered that activation of the AHR signaling pathway is reduced in the intestinal mucosa of IBD patients with CARD9 gene defects[42]. An analysis of the fecal metabolites of UC patients with defects in this gene revealed that indole, a degradation product of Trp, was present in reduced amounts, leading to decreased activation of the indole-related AHR signaling pathway and decreased ability to produce IL-22. This might be the pathophysiological mechanism through which CARD9 gene defects are involved in the pathogenesis of UC[43].

Research has shown that certain pathogenic bacteria are involved in the progression of IBD. A retrospective study revealed that in the peripheral blood of IBD patients with a longer disease course, the concentrations of Clostridium difficile, Escherichia coli, Salmonella, and Staphylococcus aureus toxins were significantly increased. In the peripheral blood of patients at the active stage of the disease, the concentrations of these bacterial toxins were significantly greater than those in patients who were at the remission stage or in healthy volunteers[44]. These findings indicate that when IBD occurs, the bacterial toxins produced by the intestinal flora are absorbed into the blood through the damaged intestinal mucosal barrier and participate in the disease process; thus, the presence of bacterial toxins in the peripheral blood can be used as an indicator of the progression of IBD. In addition, research revealed that many bacteria can bind to IgA or IgG in the intestinal lumen of patients with active IBD and that the content of soluble IgA/IgG in the feces of such patients is both significantly greater than that in healthy volunteers and positively correlated with IBD disease activity[45].

IBD-RELATED GENES

The genetic basis of IBD has been studied extensively. For example, Khor et al[46] reported that children with IBD and adult patients share the same susceptibility genes, indicating that IBD occurrence is not limited to specific age groups and that there is a genetic predisposition to the disease. Jostins et al[47] analyzed the complete genomes of patients with CD and UC and discovered 71 associated pathways and 163 IBD susceptibility genes. Among the susceptibility genes, 110 were related to the phenotypes of both CD and UC[47]. Thirty susceptibility gene loci were associated only with the CD phenotype, and 23 susceptibility gene loci were associated only with the UC phenotype. Other studies have shown that some genes that are associated with susceptibility to IBD are also associated with susceptibility to other immune-mediated diseases. For instance, type 1 diabetes is associated with mutations in the PTPN2 and PTPN22 genes, psoriasis is associated with mutations in the CDKAL1 and GCKR genes, and type 2 diabetes, ankylosing spondylitis, and systemic lupus erythematosus are associated with mutations in the CDKAL1, IL23R, and PTPN22 genes, respectively. These findings indicate that IBD may share a cellular immune mechanism with other immune-mediated diseases[48].

Influence of IBD-related genes on the intestinal barrier

Huang et al[49] reported that intestinal homeostasis affects the occurrence and development of IBD and that its maintenance is influenced by factors such as genetic susceptibility, the integrity of the intestinal epithelial barrier, and the microbial barrier. The barrier function of intestinal epithelial cells is influenced by the NOD2 gene. The protein encoded by the NOD2 gene recognizes peptidoglycans produced by the decomposition of intestinal bacteria and transmits signals that initiate an immune response that prevents pathogenic bacteria from invading intestinal epithelial cells; thus, it exerts a protective effect on the intestinal barrier. Duerr et al[50] reported that a coding variant of the IL23R gene (rs11209026, c.1142G>A, p.Arg381Gln) is involved in the host immune response and can inhibit the occurrence and development of CD. The antibacterial active peptides secreted by intestinal Paneth cells and secretions such as mucin, intestinal trefoil factor, and resistin-like molecule β produced by goblet cells have important regulatory effects on the intestinal mucosal barrier. The single-nucleotide polymorphism rs11741861, which is found in a variant of the IRGM gene, and the single-nucleotide polymorphism rs12994997, which is found in a variant of the ATG16 L1 gene, are associated with inhibition of selective autophagy by Paneth cells and goblet cells. This affects the relevant signaling molecules secreted by cells and reduces the ability of intestinal epithelial cells to defend against invasion by harmful bacteria[51].

Influence of IBD-related genes on the intestinal flora

Sokol et al[52] reported that the IBD-related gene CARD9 promotes the repair of intestinal epithelial cells. Mouse enteritis models were generated by inducing CARD9^-/- mice and wild-type mice with dextran sulfate sodium. After the model was established, the weight loss of CARD9^-/- mice was greater than that of wild-type mice, colonic shortening was more obvious, and tissue damage was more severe. Moreover, the ability of CARD9^-/- mice to express cytokines that promote epithelial healing, such as IL-17 and IL-22, was reduced[52]. Lamas et al[53] reported that the CARD9 gene plays a role in alleviating colonic inflammation by promoting the secretion of IL-22, while the intestinal microbiota of CARD9^-/- mice cannot metabolize Trp to produce ligands of the endogenous AHR. Therefore, intestinal inflammation in these mice cannot be alleviated by activating AHR signaling. When the intestinal microbiota of CARD9^-/- mice were transplanted into germ-free wild-type mice, the susceptibility of the wild-type mice to colitis increased. Aschard et al[54] reported that the abundance of Roseburia and Faecalibacterium prausnitzii in the intestines of NOD2^-/- mice decreased, as did the abundance of Faecalibacterium prausnitzii in the intestines of CARD9^-/- mice. The ATG16 L1 and NOD2 genes are involved in the protective effect of Bacteroides fragilis against colitis[54]. Ramanan et al[55] reported that deletion of the NOD2 gene promoted the proliferation of Bacteroides vulgatus, a harmful intestinal bacterium, and contributed to the development of colonic inflammation.

INTESTINAL MICROBIOTA IMBALANCE AND IBD

Under normal circumstances, microorganisms that colonize the intestine maintain a state of dynamic equilibrium with the host’s immune system, together resisting the invasion of pathogenic microorganisms and maintaining the intestinal mucosal microenvironment healthy. Intestinal microorganisms participate not only in food metabolism and energy absorption, thereby providing necessary nutrients, but also in the degradation of indigestible compounds. In addition, they play significant roles in the formation of the intestinal barrier and the regulation of immune function. Studies have shown that both the gut microbiota and its metabolic products affect the health of the host’s gut, and an increasing number of metabolic products have been identified and functionally characterized in IBD studies[56].

Metagenomic study of the gut microbiota involved in IBD

Through metagenomic research, the types and relative changes in the abundance of specific populations of microorganisms in the intestines of IBD patients were partially determined. Intestinal microbiota samples from IBD patients showed decreased overall diversity and a reduction in the abundance of anti-inflammatory species[57]. The main manifestations of this are a significant reduction in the abundance of beneficial bacteria such as Clostridium IV and XIVa, Bacteroides, Sutterella, Rosacea, Bifidobacterium, and Clostridium pratricum and a significant increase in the abundance of proinflammatory bacteria such as Proteus, Veillonella, Pasteurella, Fusobacterium, and Ruminococcus[58]. Recent metagenomic data have shown that the abundance of Akmannibacterium mucoproteinophilus is negatively correlated with IBD[14], and it has been shown that the β-N-acetylhexosaminase secreted by Akmannibacterium mucoproteinophilus yields a protective effect against sodium dextrose sulfate-induced colitis in mice[59]; notably, β-N-acetylhexosaminase participates in shaping the host barrier and the host immune response by regulating intestinal ecological imbalance and colonization by other pathogens[60]. To determine whether the changes in the gut microbiota observed in IBD patients are the cause or the result of the disease, Dwijayanti et al[61] used 16S rRNA gene sequencing to detect changes in the microbiota in the feces of newly diagnosed and newly treated IBD patients. Changes in the gut microbiota were found and were detectable at the onset of the disease. Therefore, changes in the composition of the gut microbiota can serve as novel biomarkers for the occurrence of IBD and for the progression in inflammation. The latest research by Mondragon Portocarrero et al[62] revealed that changes in the composition of the gut microbiota are greater in CD patients than in UC patients. In IBD patients, the main change is a significant increase in the population of Enterobacteriaceae, among which Escherichia coli, Klebsiella mutabilis, Klebsiella pneumoniae, Proteus mirabilis, Citrobacter freesiformis and Citrobacter citrobacter show the most obvious increasing trends. The intestinal tracts of UC patients display a greater abundance of Citrobacter aureus, Citrobacter pasteurella, Citrobacter walkerman and Proteus huis than do those of healthy people. The number and abundance of the species Klebsiella acidophilus, Morganella morganii and Citrobacter citrate is significantly increased in patients with CD.

Fungi account for a relatively small proportion of the intestinal microbiota, but they exhibit close antagonistic or synergistic relationships with bacteria and viruses in the intestine and play important roles in the occurrence and progression of IBD[63]. The fungal flora in the human intestinal tract comprise mainly three groups: Ascomycota, Basidiomycota and Chytridomycota. When inflammation occurs, the abundance and diversity of these fungi increase significantly, while the proportion of anti-inflammatory fungi such as yeast decreases. Research observed the composition of bacteria and fungi in the feces of 235 IBD patients and 38 healthy individuals. Their analysis revealed that in the intestines of IBD patients, the proportions of Basidiomycota and Ascomycota, especially the proportion of proinflammatory fungi such as Candida albicans, had significantly increased[64]. Through deep sequencing of the fecal fungal microbiome combined with retrospective studies, it was found that the abundance of yeast and Candida in the intestines of UC patients increased significantly during inflammation, while that of Penicillium increased significantly during remission. After studying the fungal composition of fecal samples obtained from 421 UC patients during clinically active periods and during remission. Research reported that the relative abundance of Candida increased by 3.5-fold during the clinically active period compared with that during the remission period and that the relative abundance of Candida during the remission period was significantly correlated with the relative abundance levels of Bacteroides parasuloides, Faecobacter propani, and Bacteroides multiforme[65]. However, these correlations disappeared during the inflammatory phase of UC. The specific mechanism underlying this effect remains unclear, and its determination requires further research. Park[66] found that Candida albicans may interact with host intestinal immune cells through the dectin-1, TLR2, and TLR4 signaling pathways and thereby participate in the occurrence and development of IBD. An other research reported that after dysregulation of the host’s intestinal fungi, the dectin-1-Syk-nuclear factor-kappa B signaling pathway was activated, promoting the expression in CD4+ T cells of key enzymes and transporters involved in glutamine dissolution and in turn promoting the occurrence of IBD and inflammatory responses[67]. In addition, clinical studies have shown that intestinal supplementation with Saccharomyces boularii can alleviate colonic inflammatory responses in mice by reshaping the intestinal microbiota and subsequently altering the types and levels of metabolites that are present. These findings indicate that fungi play a key role in controlling the progression of intestinal inflammation and provide important clues that can be used in further research on the relationship between fungi and IBD[68].

Viruses, as another important component of the gut microbiota, form the gut virome by infecting both prokaryotic and eukaryotic cells. In healthy individuals, phages of the order Caviridae or the family Microviridae are dominant. They infect host bacteria in a latent state and produce a small number of viral particles that may infect and kill other bacteria. The enteroviriome of IBD patients shows significant changes. Notably, the abundance of bacteriophages in IBD patients is greater, and the population of caudate microbiota is significantly larger, accompanied by an increase in the abundance of bacteriophages that infect bacteria belonging to the genera Escherichia and Enterobacter[69]. In addition, changes in the composition of the virome can reflect changes in the composition of the bacteria. The changes in the virome and biome of patients with CD are more obvious than those in patients with UC and can reflect the severity of the disease[70].

Metatranscriptomics of the intestinal microbiota in IBD

Metatranscriptomic analysis, which has been made possible through the rapid development of high-throughput sequencing technology, enables us to better understand the molecular mechanisms through which the gut microbiota regulate host gene expression and function. Such analysis can facilitate the study of the gut microbiota at the level of its genetic material[71]. The transcriptional activity of the intestinal microbiota is influenced by multiple factors such as the host’s health and disease status, the immune microenvironment, the host’s diet, and the microbial ecosystem[72]. On the basis of comparative metagenomic and metatranscriptomic analyses of healthy individuals and IBD patients, research reported that the functional potential of the gut microbiota, as determined through metagenomic analysis, is usually, but not always, positively correlated with the metatranscriptomic results. For instance, the abundance levels of Desulfovibrio invisus and Parabacteroides merdae are comparable at the DNA level, but the genome of Desulfovibrio invisus is not actively transcribed in the intestine, while that of Parabacteroides merdae is. A related analysis revealed that the abundance of Bacteroides fragilis DNA was significantly lower in UC patients than in a healthy control group higher in CD patients than in healthy controls[73]. However, the RNA abundance of Bacteroides fragilis in the intestines of patients with CD was similar to that in the healthy control group but was significantly decreased in patients with UC. Studies have shown that when an inflammatory response occurs in the host intestine, multiple metabolic pathways within the intestinal microbiota, including the methyl erythritol phosphate pathway, which is dominated by Alistipes putredinis, and the dTDP-L-rhamnose biosynthesis pathway of Faecalibacterium prausnitzii, are differentially expressed[74]. These pathways are involved in functions such as inducing and regulating inflammation and immune responses and altering interactions among species within the gut. Klepinowski et al[75] reported that the differences in the abundance of the transcriptomes of Clostridium hathewayi, Clostridium bolteae and active Ruminococcus were greater than the differences between the genomic abundances of those species, suggesting that their functional differences might be more significant than genomic ones. These findings suggest that the actual functional activity and the functional potential of the intestinal microbiota in IBD patients differ. Such differences yield more diverse functions of the intestinal microbiota. In-depth research is needed to better understand the potential mechanisms of intestinal microbiota imbalance and its regulatory role in the progression of IBD.

Metabolomics of the gut microbiota in IBD patients

The stability of the intestinal microenvironment is maintained by complex interactions among the intestinal microbiota, intestinal epithelial cells and immune cells. One of the main ways through which microorganisms and their hosts interact is through metabolites. Microbial metabolites serve as important feedback signals in this process to ensure normal function of the epithelial barrier and immune cells. Changes in the composition of microorganisms can lead to changes in their metabolic products. The small molecules that represent these intermediate or final products of microbial metabolism originate from the metabolism of food, from modification by host molecules (such as BAs), and directly from bacteria[76]. These microorganism-derived metabolites jointly maintain host immune maturation, immune homeostasis, energy metabolism and mucosal integrity. Among them, SCFAs, tryptophan and BAs have received considerable attention in IBD research.

SCFAs: SCFAs are the metabolic products of specific flora in the human intestinal microbiota and are produced mainly by probiotics through metabolism in the intestinal tract. The SCFAs acetate, propionate and butyrate are important anti-inflammatory bacterial metabolites, and the number of bacteria that produce these metabolites is significantly reduced in the intestines of IBD patients[77]. Bacteroidetes produce the majority of acetate and propionate, while Firmicutes are the main producers of butyrate. SCFAs enhance the immune function of the intestinal mucosa by synergistically regulating intestinal pH, promoting mucus production, and providing energy for epithelial cells[78]. They also regulate the integrity and permeability of the intestinal barrier in various ways. Studies have shown that butyrate present in the intestine can repair the intestinal mucus layer by increasing the expression and secretion of mucin-2 in epithelial cells and simultaneously promoting the polarization of M2 macrophages, thereby promoting mucus layer repair[79]. In vitro studies have confirmed that butyrate inhibits neutrophil migration and the formation of neutrophil extracellular traps in patients with UC and CD. Transcriptome sequencing analysis revealed that the immunomodulatory effect of butyrate on neutrophils in patients with IBD involves signaling pathways related to cell activation, the innate immune regulatory response, and the oxidative stress response[80]. In addition, propionate and butyrate inhibit the maturation of DCs that recognize matrix metalloproteinases, produce proinflammatory cytokines, and interact with T cells in a way that damages the mucosal barrier[81]. Animal experiments have demonstrated that SCFAs can activate histone acetylases through the GPR pathway, regulate the recognition of innate immunity and reduce the secretion of cytokines such as IL-23, IL-17, and IL-1β, thereby regulating the integrity and permeability of the intestinal barrier[82]. In addition, a phenomenon referred to as microbial cross-feeding, in which one microorganism utilizes the final metabolic products of another, occurs in the intestinal tract. For instance, AIEC in IBD patients degrades SCFAs and thereby counteracts their anti-inflammatory effects, ultimately leading to immune dysregulation in the intestinal microenvironment of IBD[83].

Trp: Trp, an essential amino acid in humans, serves as a reaction substrate and binds to active compounds that provide key physiological functions in biosynthesis. Studies have demonstrated that Trp deficiency can lead to the occurrence of IBD and exacerbate active inflammation in individuals with this disease. Combined transcriptomics and metabolomics analysis revealed various changes in intestinal Trp metabolism during inflammatory activity in IBD patients; these changes included reduced intestinal Trp absorption, activation of aromatic receptor signaling, enhancement of the kynurenine pathway, increased interstitial serotonin availability, and changes in the indole pathway[84]. Trp metabolites such as kynurenine and indole, produced through a series of endogenous enzymes or through microbial metabolism, can bind to and activate the AHR. The sequence of processes that is activated in this way is called the Trp-AHR pathway. The activated Trp-AHR pathway subsequently induces the expression of downstream cytokines such as IL-22 and IL-17[85], which regulate the release of antimicrobial peptides by intestinal epithelial cells and thereby regulate intestinal homeostasis. Further studies have revealed that Trp metabolites such as indole-ethanol, indole-3-pyruvate and indole-3-aldehyde maintain the stability of the intestinal epithelial barrier by affecting the integrity of the apical junctional complex and its associated actin regulatory proteins[86].

BAs: BAs, which are metabolites of cholesterol, play important roles in driving physiological processes such as nutrient absorption, glucose homeostasis and the regulation of energy expenditure. They have a bidirectional regulatory role with respect to the intestinal microbiota. The intestinal microbiota regulate BA metabolism in the gut-liver circulation, and BAs can exert direct toxic effects on intestinal bacteria by destroying their cell membrane structure and by other means, thereby regulating the composition and function of the intestinal microbiota[87]. In addition to their direct effects, BAs indirectly affect the intestinal microbiota through BA receptors that stimulate the host to produce antimicrobial peptides[88]; this includes an indirect effect mediated by the farnesoid X receptor, which is encoded by NR1H4, on the composition of the intestinal microbiota[89]. Animal experiments have revealed that BAs can alter the composition of the intestinal microbiome in rats; notably, they increase the number of Clostridium and erysipeloides and reduce the number of species belonging to the phyla Bacteroides and Actinomycetes[89]. Moreover, in mouse models, lithocholic acid can serve as an anti-inflammatory agent to alleviate IBD to a certain extent[90]. In addition, BAs can also inhibit the release of proinflammatory cytokines, regulate immune tolerance and maintain the intestinal barrier. Studies have shown that choline and deoxycholic acid derivatives can serve as important signaling molecules that regulate the differentiation of Th17 cells and Tregs, further influencing the intestinal inflammatory response mediated by immune dysregulation[91].

TREATMENT OF IBD

Probiotic therapy

Recent studies have shown that supplementation of the diet with probiotics has a positive in IBD[92]. Estevinho et al[93] demonstrated that when the diets of 200 IBD patients were supplemented with a mixture of probiotics such as Streptococcus thermophilus, Lactobacillus acidophilus, and Bifidobacterium praxis, the incidence of adverse reactions caused by steroid treatment and surgery decreased, and that the higher the frequency of probiotic administration was, the better the effect. Moreover, symptom relief was more obvious in UC patients than in CD patients. Sang et al[94] randomly divided 40 IBD patients into two groups, with 20 patients in each group. After 90 days of intervention with probiotics or placebo, the oxidative stress levels in the IBD patients who took oral probiotics were found to be safely and effectively regulated[94]. Minhas et al[95] demonstrated that when UC patients were treated with the mixed probiotic VSL#3 (comprising 3 species of Bifidobacterium, 4 species of Lactobacillus, and Streptococcus thermophilus), the levels of TLR2 and IL-12 p40 decreased, the level of IL-10 increased, and the symptoms of patients with mild to moderate UC were effectively relieved. Fetarayani et al[77] reported that the ability of Escherichia coli Nissle 1917 to relieve colitis is similar to that of mesalazine. On the one hand, probiotics can inhibit the growth and proliferation of pathogenic bacteria, such as Citrobacter rodentium of rodents, Streptococcus bovis and Bacteroides, in the intestinal tract[96]. On the other hand, probiotics can promote the expression of tight junction proteins in intestinal epithelial cells, reduce epithelial cell apoptosis and regulate the thickness of the intestinal mucus layer, thereby enhancing intestinal barrier function[97]. Other studies have revealed that probiotic supplements containing a mixture of Lactobacillus, Bifidobacterium, Lactobacillus fermentosus, and other bacteria downregulate the expression of the IL-1, IL-8 and TNF-α genes in the peripheral blood of patients with Parkinson’s disease and exert beneficial effects[98]. When the probiotic Lactobacillus acidophilus La-5 is cocultured with Escherichia coli O157:H7, significant inhibition of the expression of the virulence gene of Escherichia coli O157:H7 occurs, reducing its potential toxic effects[99]. At present, treatment of IBD with probiotics focuses mainly on drug efficacy. The impact of probiotic treatment on gene expression in IBD patients remains to be further explored. In addition, in terms of whether probiotics are able to colonize the intestinal tract, the types of probiotics that are beneficial, and the influence of the host’s intestinal environment, it should be noted that taking probiotics cannot increase the diversity of the host’s microbiota and is not beneficial to recovery of the structure of the intestinal microbiota. The safety and effectiveness of the long-term use of probiotics in IBD patients remain to be studied.

Fecal microbiota transplantation therapy

Fecal microbiota transplantation (FMT) involves specifically treating the feces of healthy individuals and introduction of the treated material into the patient’s intestines through nasogastric tubes or enemas[100]. Zhang et al[101] reported that when UC patients were treated with FMT and a placebo, the number and proportion of UC patients who achieved remission were 11/41 (27%) and 3/40 (8%), respectively. The diversity of the gut microbiota in patients receiving FMT increases and remains stable over a certain period. The abundance of Fusobacterium and Sartre in UC patients is greater than that in healthy people, whereas Fusobacterium nearly disappears from the intestines of UC patients who are in remission[101]. These results suggest that FMT may restore the normal structure of the intestinal flora by reducing the abundance of Fusobacterium and Sartre, thereby alleviating the symptoms of the disease. In a randomized clinical trial in which the clinical manifestations of patients (such as the degree of loose stool and mental state) and the results of colonic endoscopy were considered, Bi et al[102] found that the clinical remission rate was significantly higher in the FMT treatment group than in the placebo group (28% vs 9%); the clinical response rate was also significantly higher in the FMT treatment group than in the placebo group (49% vs 28%). Cheng et al[103] reported that among 459 children and adult IBD patients who received FMT treatment, the clinical remission rate was 28.8% (132/459), the effective response rate was 53% (241/459), and the total incidence of adverse reactions was 28.5%. The remission effect of FMT was more obvious in patients with moderate to severe IBD[103]. Clinical studies have shown that patients who undergo FMT may experience digestive tract symptoms such as fever, abdominal pain, abdominal distension, and diarrhea, as well as extraintestinal symptoms such as nasal congestion, vomiting, and sore throat. The occurrence of various symptoms may be related to individual factors, but they all persist for only a short time and can be rapidly relieved by drug intervention[104]. The mechanism through which FMT alleviates IBD symptoms is that patients receive the intestinal flora of a healthy human body, and this increases the diversity of the intestinal flora in the patient’s body. The beneficial bacteria that are introduced compete for the limited nutrients available in the intestine, inhibit the growth of pathogenic bacteria, and simultaneously produce metabolites such as SCFAs and IgA that help regulate host immune function, an effect that is conducive to restoration of intestinal barrier function[105]. In a randomized controlled study of the use of FMT in the treatment of active UC, the remission rate was significantly greater in the FMT group than in the placebo group at week 7 of treatment (24% vs 5%), while the diversity of intestinal bacteria in the FMT-treated patients was also greater than that in the placebo group. In a double-blind controlled study in which a fecal microbiota enema solution obtained from multiple volunteers was used to treat active UC, the hormone-free remission and endoscopic remission rates at week 8 of treatment were higher in the FMT group than in the placebo group (relative risk value 3.6, 95% confidence interval: 1.1-11.9; P = 0.021), and the diversity of intestinal bacteria in the patients who received FMT was greater than that in the placebo group[106].

FMT has a significant effect in the treatment of recurrent Clostridium difficile infection, but its efficacy for other diseases (such as IBD, metabolic diseases, etc.) is controversial. The indications for FMT have not yet been fully clarified. Although it has been approved for Clostridium difficile infection, more high-quality research is still needed for the treatment of other diseases. For instance, the application of FMT in treating diseases such as obesity and autism is still in the exploratory stage and lacks unified standards.

FMT may cause adverse reactions such as abdominal pain, diarrhea, and infection. Although the overall safety is relatively high, there are still rare and serious incidents reported, such as sepsis and allergic reactions. In addition, quality control issues during donor screening and fecal treatment may also affect safety. The long-term impact of FMT on the gut microbiota remains unclear. Studies have shown that the microbiota after transplantation may change in the short term, but its long-term stability and impact on host health still require further research[107].

However, FMT treatment also exhibits limitations. For instance, the donated material is unstable and difficult to replicate. Currently, there are no scientific or unified standards for its preparation or for the dosage sizes or administration frequencies used. The safety and efficacy of long-term FMT treatment still require further research. Meta-analyses have confirmed the efficacy of FMT in treating active UC, but its long-term tolerability and safety remain unclear. Moreover, there is a lack of evidence for its efficacy in the treatment of CD. Additionally, ethical controversies exist regarding FMT treatment. Studies have shown that FMT can lead to transmission of the virus group of donors to recipients, increasing the risk of disease in patients[107].

APPLICATION VALUE OF MICROECOLOGICAL OMICS RESEARCH IN THE CLINICAL DIAGNOSIS, TREATMENT AND PROGNOSIS OF IBD

In recent years, in clinical translational research on IBD, the study of intestinal microecoomics has been a hotly pursued topic. The changes in the gut microbiota that occur in IBD may provide new clues that can be useful in clinical diagnosis, treatment and prognosis. Fecal bacterial profiles differ between IBD patients and healthy individuals, while fecal bacterial colonization in IBD patients also shows disease selectivity. In a cohort study, researchers performed 16S rRNA analysis of feces collected from 2045 IBD patients; they reported that the microbiota of CD patients were imbalanced, with reduced diversity and decreased quantity. Eight types of microbiota were screened for use in the differentiation of patients with CD from patients with other diseases. Among them, Faecalibacterium, nameless bacteria of the family Streptococcaceae, anaerobic bacilli, Methanobacillus, and nameless bacteria of the family Christensenellaceae were present both in the normal control group and in UC patients. Clostridium and Escherichia coli were enriched in the feces of CD patients, and Collinsella was detected only in the feces of UC patients. When fecal bacteria were used as markers to distinguish individuals with CD from individuals with other diseases, it was found that the accuracy rate for differentiating CD patients from healthy individuals or patients with other functional disease reached 77% (sensitivity: 60%; specificity: 68%), while the accuracy rate for differentiating CD patients from UC patients was 64% (sensitivity: 60%, specificity: 94.8%)[108]. In another cohort study, fecal microbiota analysis was performed on samples collected from 21 patients with CD in remission, 17 of their siblings, and 19 healthy volunteers. The results revealed that IBD patients and their relatives displayed similar changes in their fecal microbiota, but the diversity was lower than that of the healthy control group. The abundance of Faecalibacterium prausnitzii was found to be significantly reduced, and there was a high risk of CD[109]. This result may explain the familial aggregation of IBD incidence. The levels of bacterial metabolites such as SCFAs and indole metabolites in the feces of IBD patients decrease; this is related to changes in the intestinal bacterial spectrum at the onset of the disease and can also be used as an indicator in the clinical diagnosis and assessment of the activity of IBD[110].

Changes in the gut microbiota may play a predictive role in the treatment of IBD. A prospective study involving the treatment of UC with an antibody against TNF-α revealed that the degree of intestinal flora imbalance prior to treatment was lower in the response group than in the nonresponse group and that the abundance of Clostridium faecalis was greater[111]. The content of Clostridium faecalis in the feces of patients in the response group increased during the induction of remission. In another study, patients with IBD were treated with vedolizumab, a monoclonal antibody; in that study, the a-diversity of the fecal microbiota of the CD patients who responded had increased significantly by week 14. Patients with CD who have increased levels of Roseburia and Burkholderiales in their feces prior to treatment experience better therapeutic effects[112]. In patients undergoing surgical treatment, the composition of the gut microbiota can also predict the likelihood of postoperative complications and the risk of recurrence. The abundance of active Ruminococcus gnavus, Bacteroides vulgatus and Clostridium perfringens in the feces of UC patients who experienced complications involving pocket inflammation after ileum pocket anal anastomosis increased, whereas the abundance of the family Lachnospiraceae and that of the genus Roseburia decreased[113]. After ileocolonic surgery for CD, changes in the microbiota at the end of the small intestine and anastomosis can directly affect the recurrence of the postoperative inflammatory response. Clostridium faecalis and Ruminococcus fulfill a protective role in this process[114]. Another study of changes in intestinal flora after surgery in patients with CD revealed that surgery affected the α-diversity of the ileal mucosa-related flora, with significant increases in the abundances of Proteus (especially Proteobacteria), fungi, and Enterococcus[115]. However, the abundance of thick-walled bacteria (especially those belonging to the families Helicobacteraceae and Ruminococcaceae and eubacteria, Ruminococcus, Butyricococcus, Dorea and Rothia, decreased significantly. This change was associated with endoscopic recurrence and can serve as an indicator for predicting postoperative recurrence.

FUNGAL MICROBIOTA AND IBD

Changes in microbiota structure: The microbiota structure of fungi in the intestines of IBD patients undergoes significant changes, manifested as an increase in the ratio of Basidiomycota to Ascomycota, among which the relative abundance of pathogenic fungi such as Candida albicans rises. Fungi induce inflammatory responses by activating immune receptors such as DC-associated C-type lectin. For instance, Candida albicans can enhance the differentiation of Th1 cells and exacerbate intestinal inflammation. Research has found that the abundance of Candida is correlated with the disease activity of UC patients. Machine learning models based on fungal abundance can assist in diagnosing the active and remission phases of the disease.

VIRAL MICROBIOTA AND IBD

The enteroviome of IBD patients is characterized by the expansion of caudate phages (Caudovirales) and the reduction of Microviridae. This dysregulation may indirectly promote inflammation by affecting the bacterial community. Viruses can affect intestinal immune homeostasis by interacting with host cells. For instance, some eukaryotic viruses may disrupt the epithelial barrier function, while bacteriophages can release inflammatory mediators by lysing bacteria. Phage therapy has shown therapeutic potential against specific pathogenic bacteria (such as AIEC) in animal models, but the safety and efficacy of clinical application still need to be further verified. The fungal and viral microbiota are involved in the pathogenesis of IBD by altering the intestinal microecological balance and activating immune responses. Future research needs to further clarify its mechanism of action and explore precise treatment strategies based on the microbiota.

CONCLUSION

IBD is a complex disease that involves multiple factors, including host genes, immune responses and the gut microbiota. The normal structure of the intestinal flora helps maintain the intestinal mucosal barrier and regulate host immune function. When internal or external factors disrupt the balance among the intestinal flora, the intestinal environment becomes conducive to the growth and reproduction of pathogenic bacteria. An increase in the abundance of pathogenic bacteria triggers an immune response in the body, ultimately leading to inflammation. The intestinal microbiota play an important immunomodulatory role in maintaining the homeostasis of the intestinal environment and interact with intestinal immune cells and intestinal epithelial cells, thereby participating in the occurrence of IBD. Studying the regulatory role of intestinal microecology in the pathogenesis of IBD provides a new theoretical basis for research on the pathogenesis of the disease and offers new ideas to guide future clinical diagnosis, disease status assessment, drug efficacy, and prognosis evaluation.

ACKNOWLEDGEMENTS

We would like to thank our colleagues in the Institute of Digestive Disease for their help and support in this research.