Targeting microbial pathogenic mechanisms as a novel therapeutic strategy in IBD.

BACKGROUND: Current therapy for patients suffering from inflammatory bowel diseases (IBD) is focused on inflammatory mechanisms exclusively and not the dysbiotic microbiota, despite growing evidence implicating a role for intestinal microbes in disease. MAIN BODY: Ongoing research into the intestinal microbiota of IBD patients, using new technologies and/or deeper application of existing ones, has identified a number of microorganisms whose properties and behaviors warrant consideration as causative factors in disease. Such studies have implicated both bacteria and fungi in the pathogenesis of disease. Some of these organisms manifest mechanisms that should be amenable to therapeutic intervention via either conventional or novel drug discovery platforms. Of particular note is a deeper characterization of microbial derived proteases and their destructive potential. CONCLUSION: Given the steady progress on the mechanistic role of the microbiota in inflammatory diseases, it is reasonable to anticipate a future in which therapeutics targeting microbial derived pathogenic factors play an important role in improving the lives of IBD patients.

Emergence of rare and low abundant anaerobic gut Firmicutes is associated with a significant downfall of Klebsiella in human colon cancer.

Gut bacterial dysbiosis has been linked to several gastrointestinal diseases, including deadly colorectal cancer (CRC), a leading cause of mortality in cancer patients. However, perturbation in gut bacteriome during colon cancer (CC, devoid of colorectal malignancy) remains poorly explored. Here, 16S rRNA gene amplicon sequencing was carried out for fecal DNA samples targeted to hypervariable V3-V4 region by employing MiSeq platform to explore the gut bacterial community shift in CC patients. While alpha diversity indices predicted high species richness and diversity, beta diversity showed marked gut bacterial compositional dissimilarity in CC versus healthy controls (HC, n = 10 each). We observed a significant (p < 0.05, Wilcoxon Rank-Sum test) emergence of low-abundant anaerobic taxa, including Parvimonas and Peptostreptococcus, in addition to Subdoligranulum, Coprococcus, Holdemanella, Solobacterium, Bilophila, Blautia, Dorea, Moryella and several unidentified taxa, mainly affiliated to Firmicutes, in CC patients. In addition, we also traced the emergence of putative probiotic taxon Slackia, belonging to Actinomycetota, in CC patients. The emergence of anaerobic Firmicutes in CC is accompanied by a significant (p < 0.05) decline in the Klebsiella, as determined through linear discriminant analysis effect size (LEfSe) and heat tree analyses. Shifts in core microbiome and variation in network correlation were also witnessed. Taken together, this study highlighted a significant and consistent emergence of rare anaerobic Firmicutes suggesting possible anaerobiosis driving gut microbial community shift, which could be exploited in designing diagnostic and therapeutic tools targeted to CC.

Macrophage-depleted young mice are beneficial in vivo models to assess the translocation of Klebsiella pneumonia from the gastrointestinal tract to the liver in the elderly.

Pathobionts are commensal intestinal microbiota capable of causing systemic infections under specific conditions, such as environmental changes or aging. However, it is unclear how pathobionts are recognized by the intestinal mucosal immune system under physiological conditions. This study demonstrates that the gut pathobiont Klebsiella pneumoniae causes injury to the epithelium and translocates to the liver in specific pathogen-free mice treated with clodronate-liposomes that depleted macrophages. In the clodronate-liposome-treated mice, indigenous classical K. pneumoniae (cKp) with non-K1/K2 capsular serotypes were isolated from the liver, indicating that gut commensal cKp translocated from the gastrointestinal tract to the liver due to the depletion of intestinal macrophages. Oral inoculation of isolated cKp to clodronate-liposome-treated mice significantly reduced the survival rates compared to that of non-treated mice. Our findings demonstrate that intestinal mucosal macrophages play a pivotal role in sensing commensal cKp and suppressing their translocation to the liver. This study demonstrates that clodronate-liposome-treated mouse models are effective for screening and evaluating drugs that prevent the translocation of cKp to the liver, providing new insights into the development of preventive protocols against K. pneumoniae infection.

CCR2-dependent CX3CR1+ colonic macrophages promote Enterococcus faecalis dissemination.

Enterococci are common commensal bacteria that colonize the gastrointestinal tracts of most mammals, including humans. Importantly, these bacteria are one of the leading causes of nosocomial infections. This study examined the role of colonic macrophages in facilitating Enterococcus faecalis infections in mice. We determined that depletion of colonic phagocytes resulted in the reduction of E. faecalis dissemination to the gut-draining mesenteric lymph nodes. Furthermore, we established that trafficking of monocyte-derived CX3CR1-expressing macrophages contributed to E. faecalis dissemination in a manner that was not reliant on CCR7, the conventional receptor involved in lymphatic migration. Finally, we showed that E. faecalis mutants with impaired intracellular survival exhibited reduced dissemination, suggesting that E. faecalis can exploit host immune cell migration to disseminate systemically and cause disease. Our findings indicate that modulation of macrophage trafficking in the context of antibiotic therapy could serve as a novel approach for preventing or treating opportunistic infections by disseminating enteric pathobionts like E. faecalis.

How infection-triggered pathobionts influence virulence evolution.

Host-pathogen interactions can be influenced by the host microbiota, as the microbiota can facilitate or prevent pathogen infections. In addition, members of the microbiota can become virulent. Such pathobionts can cause co-infections when a pathogen infection alters the host immune system and triggers dysbiosis. Here we performed a theoretical investigation of how pathobiont co-infections affect the evolution of pathogen virulence. We explored the possibility that the likelihood of pathobiont co-infection depends on the evolving virulence of the pathogen. We found that, in contrast to the expectation from classical theory, increased virulence is not always selected for. For an increasing likelihood of co-infection with increasing pathogen virulence, we found scenario-specific selection for either increased or decreased virulence. Evolutionary changes, however, in pathogen virulence do not always translate into similar changes in combined virulence of the pathogen and the pathobiont. Only in one of the scenarios where pathobiont co-infection is triggered above a specific virulence level we found a reduction in combined virulence. This was not the case when the probability of pathobiont co-infection linearly increased with pathogen virulence. Taken together, our study draws attention to the possibility that host-microbiota interactions can be both the driver and the target of pathogen evolution. This article is part of the theme issue 'Sculpting the microbiome: how host factors determine and respond to microbial colonization'.

Metabolic network of the gut microbiota in inflammatory bowel disease.

Gut dysbiosis is closely linked to the pathogenesis of inflammatory bowel disease (IBD). Emerging studies highlight the relationship between host metabolism and the modulation of gut microbiota composition through regulating the luminal microenvironment. In IBD, various disease-associated factors contribute to the significant perturbation of host metabolism. Such disturbance catalyzes the selective proliferation of specific microbial populations, particularly pathobionts such as adherent invasive Escherichia coli and oral-derived bacteria. Pathobionts employ various strategies to adapt better to the disease-associated luminal environments. In addition to the host-microbe interaction, recent studies demonstrate that the metabolic network between commensal symbionts and pathobionts facilitates the expansion of pathobionts in the inflamed gut. Understanding the metabolic network among the host, commensal symbionts, and pathobionts provides new insights into the pathogenesis of IBD and novel avenues for treating IBD.

Temperature influences commensal-pathogen dynamics in a nasal epithelial cell co-culture model.

Chronic rhinosinusitis (CRS) is an inflammatory disease of the paranasal sinuses, and microbial dysbiosis associated with CRS is thought to be a key driver of host inflammation that contributes to disease progression. Staphylococcus aureus is a common upper respiratory tract (URT) pathobiont associated with higher carriage rates in CRS populations, where S. aureus-secreted toxins can be identified in CRS tissues. Although many genera of bacteria colonize the URT, few account for the majority of sequencing reads. These include S. aureus and several species belonging to the genus Corynebacterium, including Corynebacterium propinquum and Corynebacterium pseudodiphtheriticum, which are observed at high relative abundance in the healthy URT. Studies have examined bacterial interactions between major microbionts of the URT and S. aureus, but few have done so in the context of a healthy versus diseased URT environment. Here, we examine the role of temperature in commensal, pathogen, and epithelial dynamics using an air-liquid interface cell culture model mimicking the nasal epithelial environment. Healthy URT temperatures change from the nares to the nasopharynx and are increased during disease. Temperatures representative of the healthy URT increase persistence and aggregate formation of commensal C. propinquum and C. pseudodiphtheriticum, reduce S. aureus growth, and lower epithelial cytotoxicity compared to higher temperatures correlating with the diseased CRS sinus. Dual-species colonization revealed species-specific interactions between Corynebacterium species and S. aureus dependent on temperature. Our findings suggest URT mucosal temperature plays a significant role in mediating polymicrobial and host-bacterial interactions that may exacerbate microbial dysbiosis in chronic URT diseases.IMPORTANCEChronic rhinosinusitis is a complex inflammatory disease with a significant healthcare burden. Although presence of S. aureus and microbial dysbiosis are considered mediators of inflammation in CRS, no studies have examined the influence of temperature on S. aureus interactions with the nasal epithelium and the dominant genus of the healthy URT, Corynebacterium. Interactions between Corynebacterium species and S. aureus have been documented in several studies, but none to date have examined how environmental changes in the URT may alter their interactions with the epithelium or each other. This study utilizes a polarized epithelial cell culture model at air-liquid interface to study the colonization and spatial dynamics of S. aureus and clinical isolates of Corynebacterium from people with CRS to characterize the role temperature has in single- and dual-species dynamics on the nasal epithelium.

Enterococci facilitate polymicrobial infections.

Enterococci are ubiquitous members of the gut microbiota in human beings and animals and are among the most important nosocomial organisms. Due to their opportunistic pathogenicity, enterococci are referred to as pathobionts and play decisive roles in a diverse array of polymicrobial infections. Enterococci can promote the colonization, pathogenesis, and persistence of various pathogens, compromise the efficacy of drugs, and pose a severe threat to public health. Most current treatments tend to focus on the sole pathogenic bacteria, with insufficient attention to the driving role of enterococci. In this review, we summarize the characteristics of enterococci in infections, the factors facilitating their outgrowth, as well as the sites and types of enterococci-associated polymicrobial infections. We present an overview of the underlying mechanisms of enterococci-mediated pathogenesis in polymicrobial infections. Furthermore, we discuss alternative strategies and potential intervention approaches to restrict such infections, shedding light on the discovery and development of new therapies against polymicrobial infections.

Butyrate reduces adherent-invasive E. coli-evoked disruption of epithelial mitochondrial morphology and barrier function: involvement of free fatty acid receptor 3.

Gut bacteria provide benefits to the host and have been implicated in inflammatory bowel disease (IBD), where adherent-invasive E. coli (AIEC) pathobionts (e.g., strain LF82) are associated with Crohn's disease. E. coli-LF82 causes fragmentation of the epithelial mitochondrial network, leading to increased epithelial permeability. We hypothesized that butyrate would limit the epithelial mitochondrial disruption caused by E. coli-LF82. Human colonic organoids and the T84 epithelial cell line infected with E. coli-LF82 (MOI = 100, 4 h) showed a significant increase in mitochondrial network fission that was reduced by butyrate (10 mM) co-treatment. Butyrate reduced the loss of mitochondrial membrane potential caused by E. coli-LF82 and increased expression of PGC-1α mRNA, the master regulator of mitochondrial biogenesis. Metabolomics revealed that butyrate significantly altered E. coli-LF82 central carbon metabolism leading to diminished glucose uptake and increased succinate secretion. Correlating with preservation of mitochondrial network form/function, butyrate reduced E. coli-LF82 transcytosis across T84-cell monolayers. The use of the G-protein inhibitor, pertussis toxin, implicated GPCR signaling as critical to the effect of butyrate, and the free fatty acid receptor three (FFAR3, GPR41) agonist, AR420626, reproduced butyrate's effect in terms of ameliorating the loss of barrier function and reducing the mitochondrial fragmentation observed in E. coli-LF82 infected T84-cells and organoids. These data indicate that butyrate helps maintain epithelial mitochondrial form/function when challenged by E. coli-LF82 and that this occurs, at least in part, via FFAR3. Thus, loss of butyrate-producing bacteria in IBD in the context of pathobionts would contribute to loss of epithelial mitochondrial and barrier functions that could evoke disease and/or exaggerate a low-grade inflammation.

Dysbiosis by Eradication of Helicobacter pylori Infection Associated with Follicular Gastropathy and Pangastropathy.

Dysbiosis plays an important role in the development of bacterial infections in the gastric mucosa, particularly Helicobacter pylori. The international guidelines for the treatment of H. pylori infections suggest standard triple therapy (STT). Nevertheless, because of the increasing resistance rates to clarithromycin, metronidazole has been widely considered in several countries. Unfortunately, the non-justified administration of antibiotics induces dysbiosis in the target organ. We characterized the gastric microbiota of patients diagnosed with follicular gastropathy and pangastropathy attributed to H. pylori infection, before and after the administration of STT with metronidazole. Dominant relative abundances of Cutibacterium were observed in pre-treatment patients, whereas H. pylori was observed at <11%, suggesting the multifactor property of the disease. The correlation of Cutibacterium acnes and H. pylori with gastric infectious diseases was also evaluated using quantitative real-time polymerase chain reaction. The dominance of C. acnes over H. pylori was observed in gastritis, gastropathies, and non-significant histological alterations. None of the microorganisms were detected in the intestinal metaplasia. Post-treatment alterations revealed an increase in the relative abundances of Staphylococcus, Pseudomonas, and Klebsiella. Non-H. pylori gastrointestinal bacteria can be associated with the initiation and development of gastric diseases, such as pathobiont C. acnes.

Fiber-deficient diet inhibits colitis through the regulation of the niche and metabolism of a gut pathobiont.

Exclusive enteral nutrition (EEN) with fiber-free diets is an effective steroid-sparing treatment to induce clinical remission in children with Crohn's disease (CD). However, the mechanism underlying the beneficial effects of EEN remains obscure. Using a model of microbiota-dependent colitis with the hallmarks of CD, we find that the administration of a fiber-free diet prevents the development of colitis and inhibits intestinal inflammation in colitic animals. Remarkably, fiber-free diet alters the intestinal localization of Mucispirillum schaedleri, a mucus-dwelling pathobiont, which is required for triggering disease. Mechanistically, the absence of dietary fiber reduces nutrient availability and impairs the dissimilatory nitrate reduction to ammonia (DNRA) metabolic pathway of Mucispirillum, leading to its exclusion from the mucus layer and disease remission. Thus, appropriate localization of the specific pathobiont in the mucus layer is critical for disease development, which is disrupted by fiber exclusion. These results suggest strategies to treat CD by targeting the intestinal niche and metabolism of disease-causing microbes.

Draft Genome Sequence of a Primate Isolate of Kazachstania pintolopesii.

Kazachstania pintolopesii is an opportunistic mammalian pathobiont from the K. telluris species complex. No draft genomes of this species are currently available. Here, we report the first draft genome sequence of a primate isolate of K. pintolopesii (NCYC 4417).

Rise of the guardians: Gut microbial maneuvers in bacterial infections.

AIMS: Bacterial infections are one of the major causes of mortality globally. The gut microbiota, primarily comprised of the commensals, performs an important role in maintaining intestinal immunometabolic homeostasis. The current review aims to provide a comprehensive understanding of how modulation of the gut microbiota influences opportunistic bacterial infections. MATERIALS AND METHODS: Primarily centered around mechanisms related to colonization resistance, nutrient, and metabolite-associated factors, mucosal immune response, and commensal-pathogen reciprocal interactions, we discuss how gut microbiota can promote or prevent bacterial infections. KEY FINDINGS: Opportunistic infections can occur directly due to obligate pathogens or indirectly due to the overgrowth of opportunistic pathobionts. Gut microbiota-centered mechanisms of altered intestinal immunometabolic and metabolomic homeostasis play a significant role in infection promotion and prevention. Depletion in the population of commensals, increased abundance of pathobionts, and overall decrease in gut microbial diversity and richness caused due to prolonged antibiotic use are risk factors of opportunistic bacterial infections, including infections from multidrug-resistant spp. Gut commensals can limit opportunistic infections by mechanisms including the production of antimicrobials, short-chain fatty acids, bile acid metabolism, promoting mucin formation, and maintaining immunological balance at the mucosa. Gut microbiota-centered strategies, including the administration of probiotics and fecal microbiota transplantation, could help attenuate opportunistic bacterial infections. SIGNIFICANCE: The current review discussed the gut microbial population and function-specific aspects contributing to bacterial infection susceptibility and prophylaxis. Collectively, this review provides a comprehensive understanding of the mechanisms related to the dual role of gut microbiota in bacterial infections.

Die Kämpfe únd schláchten-the struggles and battles of innate-like effector T lymphocytes with microbes.

The large majority of lymphocytes belong to the adaptive immune system, which are made up of B2 B cells and the αß T cells; these are the effectors in an adaptive immune response. A multitudinous group of lymphoid lineage cells does not fit the conventional lymphocyte paradigm; it is the unconventional lymphocytes. Unconventional lymphocytes-here called innate/innate-like lymphocytes, include those that express rearranged antigen receptor genes and those that do not. Even though the innate/innate-like lymphocytes express rearranged, adaptive antigen-specific receptors, they behave like innate immune cells, which allows them to integrate sensory signals from the innate immune system and relay that umwelt to downstream innate and adaptive effector responses. Here, we review natural killer T cells and mucosal-associated invariant T cells-two prototypic innate-like T lymphocytes, which sense their local environment and relay that umwelt to downstream innate and adaptive effector cells to actuate an appropriate host response that confers immunity to infectious agents.

Diet prevents the expansion of segmented filamentous bacteria and ileo-colonic inflammation in a model of Crohn's disease.

BACKGROUND: Crohn's disease (CD) is associated with changes in the microbiota, and murine models of CD-like ileo-colonic inflammation depend on the presence of microbial triggers. Increased abundance of unknown Clostridiales and the microscopic detection of filamentous structures close to the epithelium of Tnf ΔARE mice, a mouse model of CD-like ileitis pointed towards segmented filamentous bacteria (SFB), a commensal mucosal adherent bacterium involved in ileal inflammation. RESULTS: We show that the abundance of SFB strongly correlates with the severity of CD-like ileal inflammation in two mouse models of ileal inflammation, including Tnf ΔARE and SAMP/Yit mice. SFB mono-colonization of germ-free Tnf ΔARE mice confirmed the causal link and resulted in severe ileo-colonic inflammation, characterized by elevated tissue levels of Tnf and Il-17A, neutrophil infiltration and loss of Paneth and goblet cell function. Co-colonization of SFB in human-microbiota associated Tnf ΔARE mice confirmed that SFB presence is indispensable for disease development. Screening of 468 ileal and colonic mucosal biopsies from adult and pediatric IBD patients, using previously published and newly designed human SFB-specific primer sets, showed no presence of SFB in human tissue samples, suggesting a species-specific functionality of the pathobiont. Simulating the human relevant therapeutic effect of exclusive enteral nutrition (EEN), EEN-like purified diet antagonized SFB colonization and prevented disease development in Tnf ΔARE mice, providing functional evidence for the protective mechanism of diet in modulating microbiota-dependent inflammation in IBD. CONCLUSIONS: We identified a novel pathogenic role of SFB in driving severe CD-like ileo-colonic inflammation characterized by loss of Paneth and goblet cell functions in Tnf ΔARE mice. A purified diet antagonized SFB colonization and prevented disease development in Tnf ΔARE mice in contrast to a fiber-containing chow diet, clearly demonstrating the important role of diet in modulating a novel IBD-relevant pathobiont and supporting a direct link between diet and microbial communities in mediating protective functions. Video Abstract.

Comparative analysis of the red-complex organisms and recently identified periodontal pathogens in the subgingival plaque of diabetic and nondiabetic patients with severe chronic periodontitis.

Aim: This analytical case-control study sought to evaluate the presence of the recently established putative periodontal pathogen organisms, Filifactor alocis and Fretibacterium fastidiosum, against the levels of the already established red-complex pathogens, Porphyromonas gingivalis, Tannerella forsythia, and Treponema denticola, in chronic periodontitis sites of patients with and without diabetes mellitus. Materials and Methods: Fifty-six subgingival plaque samples were obtained from the deepest sites of subjects diagnosed with severe chronic periodontitis with and without diabetes mellitus. These patients were categorized into two groups of 28 each. Clinical parameters were recorded and microbial analysis was done with quantitative polymerase chain reaction, and the bacterial counts of F. alocis and F. fastidiosum were determined and then compared with that of the red-complex organisms. Results: The bacterial counts were found to be higher in the diabetic group than that in the nondiabetic group, which was statistically significant for T. forsythia (P < 0.037) and T. denticola (P < 0.003). The study found very less number of F. alocis, which was slightly higher in the diabetic group. When correlating the bacterial levels within the nondiabetic groups, the red complex species had a strong positive correlation both individually with F. alocis (P < 0.0001) and F. fastidiosum (P < 0.001) and also when the newer species was clubbed together as a cohort (P < 0.0001). Whereas, in the diabetic group, although there was a positive correlation, there was no statistical significance. Conclusion: The results of this study highlighted the presence of a definite difference in the subgingival microbiota of both the patient groups evaluated. They also indicate that of the newly identified microorganisms, both the cohorts had higher levels of F. fastidiosum, suggesting a pathobiont-like role of this bacteria among both these periodontitis groups. F. alocis was comparatively lesser in number among the cohorts evaluated, and the cause for this decreased level of F. alocis needs to be further evaluated. The results of the present study depict a higher bacterial load in the diabetic group when compared to the nondiabetic group. Further, the study demonstrates a strong correlation between the red-complex species and the newer organisms in the nondiabetic group.

Adherent-invasive E. coli - induced specific IgA limits pathobiont localization to the epithelial niche in the gut.

Background and aim: Adherent-invasive E. coli (AIEC) has been identified as a pathobiont associated with Crohn's disease (CD), that prefers to grow in inflammatory conditions. Although the colonization by AIEC is implicated in the progression of the disease and exacerbates inflammation in murine colitis models, the recognition and response of host immunity to AIEC remains elusive. Methods: Antibiotic treated female C57BL/6 mice were inoculated by commensal E. coli and LF82 AIEC strains. Luminal-IgA fractions were prepared from feces and their binding to AIEC and other strains was assessed to confirm specificity. IgA binding to isogenic mutant strains was performed to identify the functional molecules that are recognized by AIEC specific IgA. The effect of IgA on epithelial invasion of LF82 strain was confirmed using in vitro invasion assay and in vivo colonization of the colonic epithelium. Results: Persistent colonization by AIEC LF82 induced secretion of luminal IgA, while commensal E. coli strain did not. Induced anti-LF82 IgA showed specific binding to other AIEC strains but not to the commensal, non-AIEC E. coli strains. Induced IgA showed decreased binding to LF82 strains with mutated adhesin and outer membrane proteins which are involved in AIEC - epithelial cell interaction. Consistently, LF82-specific IgA limited the adhesion and invasion of LF82 in cultured epithelial cells, which seems to be required for the elimination in the colonic epithelium in mice. Conclusion: These results demonstrate that host immunity selectively recognizes pathobiont E. coli, such as AIEC, and develop specific IgA. The induced IgA specific to pathobiont E. coli, in turn, contributes to preventing the pathobionts from accessing the epithelium.

Association of nine pathobionts with periodontitis in four South American and European countries.

Aim: Our aim was to compare the prevalence and load of nine pathobionts in subgingival samples of healthy individuals and periodontitis patients from four different countries. Methods: Five hundred and seven subgingival biofilm samples were collected from healthy subjects and periodontitis patients in Belgium, Chile, Peru and Spain. The prevalence and load of Eubacterium brachy, Filifactor alocis, Fretibacterium fastidiosum, Porphyromonas endodontalis, Porphyromonas gingivalis, Selenomonas sputigena, Treponema denticola, Tannerella forsythia and Treponema socranskii were measured by quantitative PCR. Results: The association with periodontitis of all species, except for T. socranskii, was confirmed in all countries but Peru, where only P. endodontalis, P. gingivalis and T. denticola were found to be significantly associated. Moreover, most species showed higher loads at greater CAL and PPD, but not where there was BOP. Through Principal Component Analysis, samples showed clearly different distributions by diagnosis, despite observing a smaller separation in Peruvian samples. Conclusions: Unlike prevalence, relative load was found to be a reliable variable to discriminate the association of the species with periodontitis. Based on this, F. alocis, P. endodontalis, P. gingivalis, T. denticola and T. forsythia may be biomarkers of disease in Belgium, Chile and Spain, due to their significantly higher abundance in periodontitis patients.

Metagenome data-based phage therapy for intestinal bacteria-mediated diseases.

Improvements in genome analysis technology using next-generation sequencing have revealed that abnormalities in the composition of the intestinal microbiota are important in numerous diseases. Furthermore, intestinal commensal pathogens that are directly involved in the onset and exacerbation of disease have been identified. Specific control of them is strongly desired. However, antibiotics are not appropriate for the control of intestinal commensal pathogens because they may kill beneficial bacteria as well. The intestinal tract contains many viruses: most are bacteriophages (phages) that infect intestinal bacteria rather than viruses that infect human cells. Phages have very high specificity for their host bacteria. Therefore, phage therapy is considered potentially useful for controlling intestinal commensal pathogens. However, the intestinal tract is a specialized, anaerobic environment, and it is impossible to isolate phages that infect host intestinal bacteria if the bacteria cannot be cultured. Furthermore, genomic analysis methods for intestinal phages have not been well established, so until recently, a complete picture of the intestinal phage has not been clear. In this review, I summarize the importance of next-generation phage therapy based on metagenomic data and describe a novel therapy against Clostridioides difficile developed using such data.