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Summary Recent studies emphasize the role of microbial metabolites in regulating gastrointestinal (GI) physiology through activation of host receptors, highlighting the potential for inter-kingdom signaling in treating GI disorders. In this study, we show that tryptamine, a tryptophan-derived bacterial metabolite, stimulates mucus release from goblet cells via activation of G-protein-coupled receptor (GPCR) 5-HT4R. Germ-free mice colonized with engineered Bacteroides thetaiotaomicron optimized to produce tryptamine (Trp D+) exhibit decreased weight loss and increased mucus release following dextran sodium sulfate treatment when compared with mice colonized with control B. thetaiotaomicron (Trp D-). Additional beneficial effects in preventing barrier disruption and lower disease activity index were seen only in female mice, highlighting sex-specific effects of the bacterial metabolite. This study demonstrates potential for the precise modulation of mucus release by microbially produced 5-HT4 GPCR agonist as a therapeutic strategy to treat inflammatory conditions of the GI tract., Graphical Abstract, Highlights • Tryptamine increases serotonin-receptor4-dependent colonic mucus release • Bacterially derived tryptamine attenuates weight loss in DSS colitis mouse model • Protective effect of tryptamine in DSS colitis is more pronounced in female mice • Tryptamine reduces colitis severity and barrier disruption specifically in female mice, Rodent Gastroenterology; Microbiology

SUMMARY Clostridium difficile infection (CDI) is an enteric bacterial disease that is increasing in prevalence worldwide. C. difficile capitalizes on gut inflammation and microbiome dysbiosis to establish infection, with symptoms ranging from watery diarrhea to toxic megacolon. We reported that the safe-in-human clinical drug ebselen (ClinicalTrials.gov: NCT03013400, NCT01452607, NCT00762671, and NCT02603081) has biochemical, cell-based, and in vivo efficacy against the toxins of C. difficile. Here, we show that ebselen treatment reduces recurrence rates and decreases colitis in a hamster model of relapsing CDI. Furthermore, ebselen treatment does not alter microbiome diversity and promotes recovery back to that of healthy controls after antibiotic-induced dysbiosis in healthy and C. difficile-infected mice. This increased microbiome recovery upon ebselen treatment correlates with a decrease in host-derived inflammatory markers, suggesting that the anti-inflammatory properties of ebselen, combined with its anti-toxin function, help to mitigate the major clinical challenges of CDI, including recurrence, microbial dysbiosis, and colitis., In Brief Garland et al. show in a hamster model that the safe-in-human molecule ebselen, with its anti-toxin and anti-inflammatory activity, decreases colitis, and prevents reoccurrence of C. difficile infection. Ebselen also helps to reduce inflammation and promote recovery of microbiome diversity after antibiotic treatment in mice., Graphical Abstract

Sepsis is a deleterious immune response to infection that leads to organ failure and is the 11th most common cause of death worldwide. Despite plaguing humanity for thousands of years, the host factors that regulate this immunological response and subsequent sepsis severity and outcome are not fully understood. Here we describe how the Western diet (WD), a diet high in fat and sucrose and low in fiber, found rampant in industrialized countries, leads to worse disease and poorer outcomes in an LPS-driven sepsis model in WD-fed mice compared with mice fed standard fiber-rich chow (SC). We find that WD-fed mice have higher baseline inflammation (metaflammation) and signs of sepsis-associated immunoparalysis compared with SC-fed mice. WD mice also have an increased frequency of neutrophils, some with an “aged” phenotype, in the blood during sepsis compared with SC mice. Importantly, we found that the WD-dependent increase in sepsis severity and higher mortality is independent of the microbiome, suggesting that the diet may be directly regulating the innate immune system through an unknown mechanism. Strikingly, we could predict LPS-driven sepsis outcome by tracking specific WD-dependent disease factors (e.g., hypothermia and frequency of neutrophils in the blood) during disease progression and recovery. We conclude that the WD is reprogramming the basal immune status and acute response to LPS-driven sepsis and that this correlates with alternative disease paths that lead to more severe disease and poorer outcomes.

Widely available technologies based on DNA sequencing have been used to describe the kinds of microbes that might correlate with health and disease. However, mechanistic insights might be best achieved through careful study of the dynamic proteins at the interface between the foods we eat, our microbes, and ourselves. Mass spectrometry-based proteomics has the potential to revolutionize our understanding of this complex system, but its application to clinical studies has been hampered by low-throughput and laborious experimentation pipelines. In response, we developed SHT-Pro, the first high-throughput pipeline designed to rapidly handle large stool sample sets. With it, a single researcher can process over one hundred stool samples per week for mass spectrometry analysis, conservatively approximately 10× to 100× faster than previous methods, depending on whether isobaric labeling is used or not. Since SHT-Pro is fairly simple to implement using commercially available reagents, it should be easily adaptable to large-scale clinical studies., Stool-based proteomics is capable of significantly augmenting our understanding of host-gut microbe interactions. However, compared to competing technologies, such as metagenomics and 16S rRNA sequencing, it is underutilized due to its low throughput and the negative impact sample contaminants can have on highly sensitive mass spectrometry equipment. Here, we present a new stool proteomic processing pipeline that addresses these shortcomings in a highly reproducible and quantitative manner. Using this method, 290 samples from a dietary intervention study were processed in approximately 1.5 weeks, largely done by a single researcher. These data indicated a subtle but distinct monotonic increase in the number of significantly altered proteins between study participants on fiber- or fermented food-enriched diets. Lastly, we were able to classify study participants based on their diet-altered proteomic profiles and demonstrated that classification accuracies of up to 89% could be achieved by increasing the number of subjects considered. Taken together, this study represents the first high-throughput proteomic method for processing stool samples in a technically reproducible manner and has the potential to elevate stool-based proteomics as an essential tool for profiling host-gut microbiome interactions in a clinical setting. IMPORTANCE Widely available technologies based on DNA sequencing have been used to describe the kinds of microbes that might correlate with health and disease. However, mechanistic insights might be best achieved through careful study of the dynamic proteins at the interface between the foods we eat, our microbes, and ourselves. Mass spectrometry-based proteomics has the potential to revolutionize our understanding of this complex system, but its application to clinical studies has been hampered by low-throughput and laborious experimentation pipelines. In response, we developed SHT-Pro, the first high-throughput pipeline designed to rapidly handle large stool sample sets. With it, a single researcher can process over one hundred stool samples per week for mass spectrometry analysis, conservatively approximately 10× to 100× faster than previous methods, depending on whether isobaric labeling is used or not. Since SHT-Pro is fairly simple to implement using commercially available reagents, it should be easily adaptable to large-scale clinical studies.

SummaryOur emerging view of the gut microbiome largely focuses on bacteria and less is known about other microbial components such as of bacteriophages (phages). Though phages are abundant in the gut, very few phages have been isolated from this ecosystem. Here, we report the genomes of 27 phages from the United States and Bangladesh that infect the prevalent human gut bacterium Bacteroides thetaiotaomicron. These phages are mostly distinct from previously sequenced phages with the exception of two, which are crAss-like phages. We compare these isolates to existing human gut metagenomes, revealing similarities to previously inferred phages and additional unexplored phage diversity. Finally, we use host tropisms of these phages to identify alleles of phage structural genes associated with infectivity. This work provides a detailed view of the gut’s “viral dark matter” and a framework for future efforts to further integrate isolation- and sequencing-focused efforts to understand gut-resident phages.

Efforts to probe the role of the gut microbiota in disease would benefit from a system in which patient-derived bacterial communities can be studied at scale. We addressed this by validating a strategy to propagate phylogenetically complex, diverse, stable, and highly reproducible stool-derived communities in vitro. We generated hundreds of in vitro communities cultured from diverse stool samples in various media; certain media generally preserved inoculum composition, and inocula from different subjects yielded source-specific community compositions. Upon colonization of germ-free mice, community composition was maintained, and the host proteome resembled the host from which the community was derived. Treatment with ciprofloxacin in vivo increased susceptibility to Salmonella invasion in vitro, and the in vitro response to ciprofloxacin was predictive of compositional changes observed in vivo, including the resilience and sensitivity of each Bacteroides species. These findings demonstrate that stool-derived in vitro communities can serve as a powerful system for microbiota research.

Appreciation is growing of how our gut microbes shape health and disease. Now, a study of ancient human faeces sheds light on how microbial populations in the gut have changed during the past 2,000 years. Insights into gut-microbe changes during the past 2,000 years.

The first rudimentary evidence that the human body harbors a microbiota hinted at the complexity of host-associated microbial ecosystems. Now, almost 400 years later, a renaissance in the study of microbiota spatial organization, driven by coincident revolutions in imaging and sequencing technologies, is revealing functional relationships between biogeography and health, particularly in the vertebrate gut. In this review, we present our current understanding of principles governing the localization of intestinal bacteria, and spatial relationships between bacteria and their hosts. We further discuss important emerging directions that will enable progressing from the inherently descriptive nature of localization and –omics technologies to provide functional, quantitative, and mechanistic insight into this complex ecosystem.

Aging is characterized by a progressive loss of physiological integrity, leading to impaired function and increased vulnerability to death(1). Despite rapid advances over recent years, many of the molecular and cellular processes which underlie progressive loss of healthy physiology are poorly understood(2). To gain a better insight into these processes we have created a single cell transcriptomic atlas across the life span of Mus musculus which includes data from 23 tissues and organs. We discovered cell-specific changes occurring across multiple cell types and organs, as well as age related changes in the cellular composition of different organs. Using single-cell transcriptomic data we were able to assess cell type specific manifestations of different hallmarks of aging, such as senescence(3), genomic instability(4) and changes in the organism’s immune system(2). This Tabula Muris Senis provides a wealth of new molecular information about how the most significant hallmarks of aging are reflected in a broad range of tissues and cell types.

While the gut microbiota's malleability makes it highly responsive to our environment, it also renders it susceptible to rapid selection by factors associated with an industrialized lifestyle. Recently, in Cell, Vangay et al. (2018) have tracked and revealed rapid and profound changes in the gut community of immigrants to one that resembles long-term residents of the U.S.

In this prospective cohort study, we examine the feasibility of a protocol to optimize microbiota for fecal microbiota transplantation (FMT). Donor stool metrics generally accepted as markers of gut health were used to select a stool donor based on superior microbial diversity, balanced constitution of Bacteroidetes versus Firmicutes and high concentration of fecal butyrate. Selected donor microbiota was then administered via FMT. A total of 10 patients with median age of 12 years with recurrent Clostridium difficile infection received the intervention. The rate of recurrence-free resolution with 1-2 FMTs was 100% at Week 10. With a single FMT, 80% of patients cleared Clostridium difficile infection without recurrence, whereas 20% of patients required a single re-treatment. No serious adverse events occurred. Microbiota sequencing revealed that recipients' gut microbiota phylogenic diversity increased by 72-hours post-transplantation, with sustainment over 10-week follow-up. This study highlights the feasibility of purposefully selecting the most ideal microbiota for transplantation.

Clostridial metabolite production The clostridia are Firmicute bacterial commensals commonly found in the mammalian gut. Clostridia produce a range of metabolites that diffuse into the host's circulation and have been difficult to manipulate genetically, but Guo et al. successfully developed a CRISPR-Cas9 deletion system in Clostridium sporogenes (see the Perspective by Henke and Clardy). The authors used deletion mutants and mass spectrometry to elucidate clostridial synthesis of several different branched short-chain fatty acids (SCFAs), including isobutyrate, 2-methylbutyrate, and isovalerate. Germ-free mice colonized with mutants incapable of synthesizing SCFAs showed altered immunoglobulin A production. This finding potentially links bacterial SCFA production and host responses to the presence of the clostridia. Science , this issue p. eaav1282 ; see also p. 1309

It has long been known that proteolytic Clostridia obtain their energy by coupling oxidative and reductive pathways for amino acid metabolism – the Stickland reaction1. The oxidation of one amino acid is coupled with reduction of another, yielding energy in the former step and re-achieving redox balance with the latter. Here, we find that the gut bacterium, Clostridium sporogenes metabolizes amino acids through reductive pathways to produce metabolites that circulate within the host. Measurements in vitro indicate that reductive Stickland pathways are coupled to ATP formation, revealing their role in energy capture by gut bacteria. By probing the genetics of C. sporogenes, we find that the Rnf complex is involved in reductive amino acid metabolism. Rnf complex mutants are attenuated for growth in the mouse gut, demonstrating the importance of energy capture during reductive metabolism for gut colonization. Our findings reveal that the production of high-abundance molecules by a commensal bacterium within the host gut is linked to an energy yielding redox process.

SummaryClostridium difficile infection (CDI) is an enteric bacterial disease that is increasing in prevalence worldwide. C. difficile capitalizes on gut inflammation and microbiome dysbiosis to establish infection, with symptoms ranging from watery diarrhea to toxic megacolon. We recently reported that the safe in human clinical drug candidate ebselen (NCT03013400, NCT01452607, NCT00762671, NCT02603081) has biochemical, cell-based and in vivo efficacy against the bacterial toxins of C. difficile. Here, we show that ebselen treatment reduces recurrence rates and decreases colitis in a hamster relapse model of CDI. Furthermore, ebselen treatment does not alter microbiome diversity but promotes its recovery back to that of healthy controls after antibiotic-induced dysbiosis in both healthy and C. difficile-infected mice. This increased microbiome recovery upon ebselen treatment correlates with a decrease in host-derived inflammatory markers suggesting that the anti-inflammatory properties of ebselen, combined with its anti-toxin function, help to mitigate the major clinical challenges of CDI, including recurrence, microbial dysbiosis, and colitis.

Given the significant impact that gut microbes have on our health, it is essential to identify key host and environmental factors that shape this diverse community. While many studies have highlighted the impact of diet on gut microbiota, little is known about how the host regulates this critical diet-microbiota interaction. In our present study, we discovered that gut IgA targeted a protein complex involved in the utilization of an important dietary polysaccharide: fructan. While the presence of dietary fructans was previously thought to allow unrestricted growth of fructan-utilizing bacteria, our work shows that gut IgA, by targeting proteins responsible for fructan utilization, provides the host with tools that can restrict the microbial utilization of such polysaccharides, thereby controlling their growth., Gut-derived immunoglobulin A (IgA) is the most abundant antibody secreted in the gut that shapes gut microbiota composition and functionality. However, most of the microbial antigens targeted by gut IgA remain unknown, and the functional effects of IgA targeting these antigens are currently understudied. This study provides a framework for identifying and characterizing gut microbiota antigens targeted by gut IgA. We developed a small intestinal ex vivo culture assay to harvest lamina propria IgA from gnotobiotic mice, with the aim of identifying antigenic targets in a model human gut commensal, Bacteroides thetaiotaomicron VPI-5482. Colonization by B. thetaiotaomicron induced a microbe-specific IgA response that was reactive against diverse antigens, including capsular polysaccharides, lipopolysaccharides, and proteins. IgA against microbial protein antigens targeted membrane and secreted proteins with diverse functionalities, including an IgA specific against proteins of the polysaccharide utilization locus (PUL) that are necessary for utilization of fructan, which is an important dietary polysaccharide. Further analyses demonstrated that the presence of dietary fructan increased the production of fructan PUL-specific IgA, which then downregulated the expression of fructan PUL in B. thetaiotaomicron, both in vivo and in vitro. Since the expression of fructan PUL has been associated with the ability of B. thetaiotaomicron to colonize the gut in the presence of dietary fructans, our work suggests a novel role for gut IgA in regulating microbial colonization by modulating their metabolism.

BACKGROUND The collection of trillions of microbes inhabiting the human gut, called the microbiome or microbiota, has captivated the biomedical research community for the past decade. Intimate connections exist between the microbiota and the immune system, central nervous system, and metabolism. The growing realization of the fundamental role that the microbiota plays in human health has been accompanied by the challenge of trying to understand which features define a healthy gut community and how these may differ depending upon context. Such insight will lead to new routes of disease treatment and prevention and may illuminate how lifestyle-driven changes to the microbiota can impact health across populations. Individuals living traditional lifestyles around the world share a strikingly similar microbiota composition that is distinct from that found in industrialized populations. Indeed, lineages of gut microbes have cospeciated with humans over millions of years, passing through hundreds of thousands of generations, and lend credence to the possibility that our microbial residents have shaped our biology throughout evolution. Relative to the “traditional” microbiota, the “industrial” microbiota appears to have lower microbial diversity, with major shifts in membership and functions. Individuals immigrating from nonindustrialized to industrialized settings or living at different intermediate states between foraging and industrialization have microbiota composition alterations that correspond to time and severity of lifestyle change. Industrial advances including antibiotics, processed food diets, and a highly sanitized environment have been shown to influence microbiota composition and transmission and were developed and widely implemented in the absence of understanding their effects on the microbiota. ADVANCES Here, we argue that the microbiota harbored by individuals living in the industrialized world is of a configuration never before experienced by human populations. This “new,” industrial microbiota has been shaped by recent progress in medicine, food, and sanitation. As technology and medicine have limited our exposure to pathogenic microbes, enabled feeding large populations inexpensively, and otherwise reduced acute medical incidents, many of these advances have been implemented in the absence of understanding the collateral damage inflicted on our resident microbes or the importance of these microbes in our health. More connections are being drawn between the composition and function of the gut microbiota and alteration in the immune status of the host. These relationships connect the industrial microbiota to the litany of chronic diseases that are driven by inflammation. Notably, these diseases spread along with the lifestyle factors that are known to alter the microbiota. While researchers have been uncovering the basic tenets of how the microbiota influences human health, there has been a growing realization that as the industrial lifestyle spreads globally, changes to the human microbiota may be central to the coincident spread of non-communicable, chronic diseases and may not be easily reversed. OUTLOOK We suggest that viewing microbiota biodiversity with an emphasis on sustainability and conservation may be an important approach to safeguarding human health. Understanding the services provided by the microbiota to humans, analogous to how ecosystem services are used to place value on aspects of macroecosystems, could aid in assessing the cost versus benefit of specific microbiota dysfunctions that are induced by different aspects of lifestyle. A key hurdle is to establish the impact of industrialization-induced changes to the microbiota on human health. The severity of this impact might depend on the specifics of numerous factors, including health status, diet, human genotype, and lifestyle. Isolating and archiving bacterial strains that are sensitive to industrialization may be required to enable detailed study of these organisms and to preserve ecosystem services that are unique to those strains and potentially beneficial to human health. Determining a path forward for sustainable medical practices, diet, and sanitation that is mindful of the importance and fragility of the microbiota is needed if we are to maintain a sustainable relationship with our internal microbial world.

Intestinal microbiotas contain beneficial microorganisms that protect against pathogen colonization; treatment with antibiotics disrupts the microbiota and compromises colonization resistance. Here, we determine the impact of exchanging microorganisms between hosts on resilience to the colonization of invaders after antibiotic-induced dysbiosis. We assess the functional consequences of dysbiosis using a mouse model of colonization resistance against Escherichia coli. Antibiotics caused stochastic loss of members of the microbiota, but the microbiotas of co-housed mice remained more similar to each other compared with the microbiotas among singly housed animals. Strikingly, co-housed mice maintained colonization resistance after treatment with antibiotics, whereas most singly housed mice were susceptible to E. coli. The ability to retain or share the commensal Klebsiella michiganensis, a member of the Enterobacteriaceae family, was sufficient for colonization resistance after treatment with antibiotics. K. michiganensis generally outcompeted E. coli in vitro, but in vivo administration of galactitol-a nutrient that supports the growth of only E. coli-to bi-colonized gnotobiotic mice abolished the colonization-resistance capacity of K. michiganensis against E. coli, supporting the idea that nutrient competition is the primary interaction mechanism. K. michiganensis also hampered colonization of the pathogen Salmonella, prolonging host survival. Our results address functional consequences of the stochastic effects of microbiota perturbations, whereby microbial transmission through host interactions can facilitate reacquisition of beneficial commensals, minimizing the negative impact of antibiotics.

The gut microbiota is a complex network of diverse organisms that exhibits plasticity and is capable of impacting host immunity. The malleability of the microbiota presents microbial alteration as an avenue for tuning the immune system to different set points, an approach that has the potential to enable a wide range of therapeutic applications, and ultimately enable disease prevention. Despite the tremendous potential in harnessing the microbiome-immune axis to benefit human health, key barriers must be addressed to hasten translation. Studies using mouse models have identified numerous specific interactions between the gut microbiota and both local and systemic immunity, in several cases using gnotobiotics and other highly controlled approaches to establish causal relationships. Recent advances in our ability to perform expansive profiling of both the microbiota and the immune system now enable exploring human-gut microbiota connections more thoroughly than ever before. An important next step in realizing the power of microbiome reprogramming is to elucidate a human-relevant “map” of microbial-immune wiring, while focusing on animal studies to probe the subset of interactions likely to be relevant to human biology. Such efforts have the potential for revealing new paradigms in immune function as it relates to the microbiome, and for harnessing this connection to improve human health. Here we provide an overview of this field’s current status and discuss two approaches for establishing priorities for detailed investigation: (i) longitudinal intervention studies in humans probing the dynamics of both the microbiota and the immune system, and (ii) the study of traditional populations to assess lost features of human microbial identity whose absence may be contributing to the rise of immunological disorders. These human centered approaches offer a judicious path forward to capitalize on the potent power of the microbiota as a driver in immune health.

SummaryThat antibiotics alter microbiota composition and increase infection susceptibility is well known, but their generalizable effects on the gut commensal community and dependence on environmental variables remain open questions. Here, we systematically compared antibiotic responses in gnotobiotic and conventional mice across antibiotics, microbiotas, diets, and housing status. We identify remarkable resilience, whereby a humanized microbiota recovers before drug administration ends, with transient dominance of resistant Bacteroides and taxa-asymmetric reduction in diversity. In other cases, in vitro sensitivities were not predictive of in vivo responses, underscoring the significance of host and community contexts. A fiber-deficient diet exacerbated collapse of the microbiota and delayed recovery, despite the presence of a similar core community across diets at the point of maximal disturbance. Resilience to a second ciprofloxacin treatment was observed via response reprogramming, in which species replacement after ciprofloxacin treatment established resilience to a second treatment, and also through cross housing transmission. Single-housing drastically disrupted recovery, highlighting the importance of environmental microbial reservoirs and suggesting sanitation may exacerbate the duration of antibiotic-mediated disruption. Our findings highlight the ability of the commensal microbiota to deterministically adapt to large perturbations, and the translational potential for modulating diet, sanitation, and microbiota composition during antibiotics.

A reduction in intestinal barrier function is currently believed to play an important role in pathogenesis of many diseases, as it facilitates passage of injurious factors such as lipopolysaccharide, peptidoglycan, whole bacteria, and other toxins to traverse the barrier to damage the intestine or enter the portal circulation. Currently available evidence in animal models and in vitro systems has shown that certain dietary interventions can be used to reinforce the intestinal barrier to prevent the development of disease. The relevance of these studies to human health is unknown. Herein, we define the components of the intestinal barrier, review available modalities to assess its structure and function in humans, and review the available evidence in model systems or perturbations in humans that diet can be used to fortify intestinal barrier function. Acknowledging the technical challenges and the present gaps in knowledge, we provide a conceptual framework by which evidence could be developed to support the notion that diet can reinforce human intestinal barrier function to restore normal function and potentially reduce the risk for disease. Such evidence would provide information on the development of healthier diets and serve to provide a framework by which federal agencies such as the US Food and Drug Administration can evaluate evidence linking diet with normal human structure/function claims focused on reducing risk of disease in the general public. ispartof: AMERICAN JOURNAL OF PHYSIOLOGY-GASTROINTESTINAL AND LIVER PHYSIOLOGY vol:317 issue:1 pages:G17-G39 ispartof: location:United States status: published

Diet is the largest source of plant-derived metabolites that influence human health. The gut microbiota can metabolize these molecules, altering their biological function. However, little is known about the gut bacterial pathways that process plant-derived molecules. Glucosinolates are well-known metabolites in brassica vegetables and metabolic precursors to cancer-preventive isothiocyanates. Here, we identify a genetic and biochemical basis for isothiocyanate formation byBacteroides thetaiotaomicron,a prominent gut commensal species. Using a genome-wide transposon insertion screen, we identified an operon required for glucosinolate metabolism inB. thetaiotaomicron.Expression of BT2159-BT2156 in a non-metabolizing relative,Bacteroides fragilis, resulted in gain of glucosinolate metabolism. We show that isothiocyanate formation requires the action of BT2158 and either BT2156 or BT2157in vitro. Monocolonization of mice with mutantBtΔ2157showed reduced isothiocyanate production in the gastrointestinal tract. These data provide insight into the mechanisms by which a common gut bacterium processes an important dietary nutrient.

The production and handling of serotonin (5-HT) is an important determinant of colonic motility and has been reported to be altered in gastrointestinal (GI) disorders such as irritable bowel syndrome (IBS). Recent studies suggest that the intestinal microbiota and sex of the host can influence expression of genes involved in 5-HT biosynthesis and signaling. While expression of genes in serotonergic pathways has been shown to be variable, it remains unclear whether genes within this pathway are coregulated. As a first step in that direction, we investigated potential correlations in relative mRNA expression of serotonergic genes, in the proximal colon isolated from male and female mice in different states of microbial association: germ-free (GF), humanized (ex-germ-free colonized with human gut microbiota, HM), and conventionally raised (CR) mice. Among the 10 pairwise comparisons conducted between five serotonergic transcripts, Tph1, Chga, Maoa, Slc6a4, and Htr4, we found a strong, positive correlation between colonic expression of Slc6a4 and Htr4 across different colonization states and sexes. We also identified a positive correlation between the expression of Tph1 and Chga; however, there were no correlations observed between any other tested pair of 5-HT-related transcripts. These data suggest that correlated expression of Slc6a4 and Htr4 likely involves coregulation of genes located on different chromosomes which modulate serotonergic activity in the gut. Further work will need to be done to understand the pathways and cell types responsible for this correlated expression, given the important role of 5-HT in gastrointestinal physiology.

A perturbed gut microbiome has recently been linked with multiple disease processes, yet researchers currently lack tools that can provide in vivo, quantitative, and real-time insight into these processes and associated host-microbe interactions. We propose an in vivo wireless implant for monitoring gastrointestinal tract redox states using oxidation-reduction potentials (ORP). The implant is powered and conveniently interrogated via ultrasonic waves. We engineer the sensor electronics, electrodes, and encapsulation materials for robustness in vivo, and integrate them into an implant that endures autoclave sterilization and measures ORP for 12 days implanted in the cecum of a live rat. The presented implant platform paves the way for long-term experimental testing of biological hypotheses, offering new opportunities for understanding gut redox pathophysiology mechanisms, and facilitating translation to disease diagnosis and treatment applications., Minor revision. Supplementary information included at the end of the main paper

A variety of cell surface structures, including capsular polysaccharides (CPS), dictate interactions between bacteria and their environment including their viruses (bacteriophages). Members of the prominent human gut Bacteroidetes characteristically produce several phase-variable CPS, but their contributions to bacteriophage interactions are unknown. We used engineered strains of the human symbiont Bacteroides thetaiotaomicron, which differ only in the CPS they express, to isolate bacteriophages from two locations in the United States. Testing each of 71 bacteriophages against a panel of strains that express wild-type phase-variable CPS, one of eight different single CPS, or no CPS at all, revealed that each phage infects only a subset of otherwise isogenic strains. Deletion of infection-permissive CPS from B. thetaiotaomicron was sufficient to abolish infection for several individual bacteriophages, while infection of wild-type B. thetaiotaomicron with either of two different bacteriophages rapidly selected for expression of non-permissive CPS. Surprisingly, acapsular B. thetaiotaomicron also escapes complete killing by these bacteriophages, but surviving bacteria exhibit increased expression of 8 distinct phase-variable lipoproteins. When constitutively expressed, one of these lipoproteins promotes resistance to multiple bacteriophages. Finally, both wild-type and acapsular B. thetaiotaomicron were able to separately co-exist with one bacteriophage for over two months in the mouse gut, suggesting that phase-variation promotes resistance but also generates sufficient numbers of susceptible revertants to allow bacteriophage persistence. Our results reveal important roles for Bacteroides CPS and other cell surface structures that allow these bacteria to persist despite bacteriophage predation and hold important implications for using bacteriophages therapeutically to target gut symbionts.

The study of traditional populations provides a view of human-associated microbes unperturbed by industrialization, as well as a window into the microbiota that co-evolved with humans. Here we discuss our recent work characterizing the microbiota from the Hadza hunter-gatherers of Tanzania. We found seasonal shifts in bacterial taxa, diversity, and carbohydrate utilization by the microbiota. When compared to the microbiota composition from other populations around the world, the Hadza microbiota shares bacterial families with other traditional societies that are rare or absent from microbiotas of industrialized nations. We present additional observations from the Hadza microbiota and their lifestyle and environment, including microbes detected on hands, water, and animal sources, how the microbiota varies with sex and age, and the shortterm effects of introducing agricultural products into the diet. In the context of our previously published findings and of these additional observations, we discuss a path forward for future work.

Secondary bile acids (SBAs) are derived from primary bile acids (PBAs) in a process reliant on biosynthetic capabilities possessed by few microbes. To evaluate the role of BAs in intestinal inflammation, we performed metabolomic, microbiome, metagenomic, and transcriptomic profiling of stool from ileal pouches (surgically created resevoirs) in colectomy-treated patients with ulcerative colitis (UC) versus controls (familial adenomatous polyposis [FAP]). We show that relative to FAP, UC pouches have reduced levels of lithocholic acid and deoxycholic acid (normally the most abundant gut SBAs), genes required to convert PBAs to SBAs, and Ruminococcaceae (one of few taxa known to include SBA-producing bacteria). In three murine colitis models, SBA supplementation reduces intestinal inflammation. This anti-inflammatory effect is in part dependent on the TGR5 bile acid receptor. These data suggest that dysbiosis induces SBA deficiency in inflammatory-prone UC patients, which promotes a pro-inflammatory state within the intestine that may be treated by SBA restoration.

The gut microbiota plays a critical role in pathogen defense. Studies using antibiotic-treated mice reveal mechanisms that increase susceptibility to Clostridioides difficile infection (CDI), but risk factors associated with CDI in humans extend beyond antibiotic use. Here, we studied the dysbiotic gut microbiota of a subset of patients with diarrhea and modeled the gut microbiota of these patients by fecal transplantation into germ-free mice. When challenged with C. difficile, the germ-free mice transplanted with fecal samples from patients with dysbiotic microbial communities showed increased gut amino acid concentrations and greater susceptibility to CDI. A C. difficile mutant that was unable to use proline as an energy source was unable to robustly infect germ-free mice transplanted with a dysbiotic or healthy human gut microbiota. Prophylactic dietary intervention using a low-proline or low-protein diet in germ-free mice colonized by a dysbiotic human gut microbiota resulted in decreased expansion of wild-type C. difficile after challenge, suggesting that amino acid availability might be important for CDI. Furthermore, a prophylactic fecal microbiota transplant in mice with dysbiosis reduced proline availability and protected the mice from CDI. Last, we identified clinical risk factors that could potentially predict gut microbial dysbiosis and thus greater susceptibility to CDI in a retrospective cohort of patients with diarrhea. Identifying at-risk individuals and reducing their susceptibility to CDI through gut microbiota–targeted therapies could be a new approach to preventing C. difficile infection in susceptible patients.

The gut microbiota produce hundreds of molecules that are present at high concentrations in circulation and whose levels vary widely among humans. In most cases, molecule production has not been linked to specific bacterial strains or metabolic pathways, and unraveling the contribution of each molecule to host biology remains difficult. A general system to ‘toggle’ molecules in this pool on/off in the host would enable interrogation of the mechanisms by which they modulate host biology and disease processes. Such a system has been elusive due to limitations in the genetic manipulability of Clostridium and its relatives, the source of many molecules in this pool. Here, we describe a method for reliably constructing clean deletions in a model commensal Clostridium, C. sporogenes (Cs), including multiply mutated strains. We demonstrate the utility of this method by using it to ‘toggle’ off the production of ten Cs-derived molecules that accumulate in host tissues. By comparing mice colonized by wild-type Cs versus a mutant deficient in the production of branched short-chain fatty acids, we discover a previously unknown IgA-modulatory activity of these abundant microbiome-derived molecules. Our method opens the door to interrogating and sculpting a highly concentrated pool of chemicals from the microbiome.

A 2-day workshop organized by the National Institutes of Health and U.S. Department of Agriculture included 16 presentations focused on the role of diet in alterations of the gastrointestinal microbiome, primarily that of the colon. Although thousands of research projects have been funded by U.S. federal agencies to study the intestinal microbiome of humans and a variety of animal models, only a minority addresses dietary effects, and a small subset is described in sufficient detail to allow reproduction of a study. Whereas there are standards being developed for many aspects of microbiome studies, such as sample collection, nucleic acid extraction, data handling, etc., none has been proposed for the dietary component; thus this workshop focused on the latter specific point. It is important to foster rigor in design and reproducibility of published studies to maintain high quality and enable designs that can be compared in systematic reviews. Speakers addressed the influence of the structure of the fermentable carbohydrate on the microbiota and the variables to consider in design of studies using animals, in vitro models, and human subjects. For all types of studies, strengths and weaknesses of various designs were highlighted, and for human studies, comparisons between controlled feeding and observational designs were discussed. Because of the lack of published, best-diet formulations for specific research questions, the main recommendation is to describe dietary ingredients and treatments in as much detail as possible to allow reproduction by other scientists.

The gut microbiota produce hundreds of molecules that are present at high concentrations in the host circulation. Unraveling the contribution of each molecule to host biology remains difficult. We developed a system for constructing clean deletions in

Summary The intestinal microbiota provides colonization resistance against pathogens, limiting pathogen expansion and transmission. These microbiota-mediated mechanisms were previously identified by observing loss of colonization resistance after antibiotic treatment or dietary changes, which severely disrupt microbiota communities. We identify a microbiota-mediated mechanism of colonization resistance against Salmonella enterica serovar Typhimurium (S. Typhimurium) by comparing high-complexity commensal communities with different levels of colonization resistance. Using inbred mouse strains with different infection dynamics and S. Typhimurium intestinal burdens, we demonstrate that Bacteroides species mediate colonization resistance against S. Typhimurium by producing the short-chain fatty acid propionate. Propionate directly inhibits pathogen growth in vitro by disrupting intracellular pH homeostasis, and chemically increasing intestinal propionate levels protects mice from S. Typhimurium. In addition, administering susceptible mice Bacteroides, but not a propionate-production mutant, confers resistance to S. Typhimurium. This work provides mechanistic understanding into the role of individualized microbial communities in host-to-host variability of pathogen transmission.

Genomic differences between gut-resident bacterial strains likely underlie significant interindividual variation in microbiome function. Traditional methods of determining community composition, such as 16S rRNA gene amplicon sequencing, fail to capture this functional diversity. Metagenomic approaches are a significant step forward in identifying strain-level sequence variants; however, given the current paucity of biochemical information, they too are limited to mainly low-resolution and incomplete functional predictions. Using genomic, biochemical, and molecular approaches, we identified differences in the fructan utilization profiles of two closely related Bacteroides thetaiotaomicron strains. B. thetaiotaomicron 8736 ( Bt-8736 ) contains a fructan polysaccharide utilization locus (PUL) with a divergent susC / susD homolog gene pair that enables it to utilize inulin, differentiating this strain from other characterized Bt strains. Transfer of the distinct pair of susC / susD genes from Bt-8736 into the noninulin using type strain B. thetaiotaomicron VPI-5482 resulted in inulin use by the recipient strain, Bt ( 8736-2 ). The presence of the divergent susC / susD gene pair alone enabled the hybrid Bt ( 8736-2 ) strain to outcompete the wild-type strain in vivo in mice fed an inulin diet. Further, we discovered that the susC / susD homolog gene pair facilitated import of inulin into the periplasm without surface predigestion by an endo-acting enzyme, possibly due to the short average chain length of inulin compared to many other polysaccharides. Our data builds upon recent reports of dietary polysaccharide utilization mechanisms found in members of the Bacteroides genus and demonstrates how the acquisition of two genes can alter the functionality and success of a strain within the gut. IMPORTANCE Dietary polysaccharides play a dominant role in shaping the composition and functionality of our gut microbiota. Dietary interventions using these microbiota-accessible carbohydrates (MACs) serve as a promising tool for manipulating the gut microbial community. However, our current gap in knowledge regarding microbial metabolic pathways that are involved in the degradation of these MACs has made the design of rational interventions difficult. The issue is further complicated by the diversity of pathways observed for the utilization of similar MACs, even in closely related microbial strains. Our current work focuses on divergent fructan utilization pathways in two closely related B. thetaiotaomicron strains and provides an integrated approach to characterize the molecular basis for strain-level functional differences.

Background Microbial conversion of dietary substrate to bioactive molecules represents a potential regulatory mechanism by which gut microbes can alter intestinal physiology. Tryptamine, a monoamin...

The gut is home to trillions of microorganisms that have fundamental roles in many aspects of human biology, including immune function and metabolism. The reduced diversity of the gut microbiota in Western populations compared to that in populations living traditional lifestyles presents the question of which factors have driven microbiota change during modernization. Microbiota-accessible carbohydrates (MACs) found in dietary fibre have a crucial involvement in shaping this microbial ecosystem, and are notably reduced in the Western diet (high in fat and simple carbohydrates, low in fibre) compared with a more traditional diet. Here we show that changes in the microbiota of mice consuming a low-MAC diet and harbouring a human microbiota are largely reversible within a single generation. However, over several generations, a low-MAC diet results in a progressive loss of diversity, which is not recoverable after the reintroduction of dietary MACs. To restore the microbiota to its original state requires the administration of missing taxa in combination with dietary MAC consumption. Our data illustrate that taxa driven to low abundance when dietary MACs are scarce are inefficiently transferred to the next generation, and are at increased risk of becoming extinct within an isolated population. As more diseases are linked to the Western microbiota and the microbiota is targeted therapeutically, microbiota reprogramming may need to involve strategies that incorporate dietary MACs as well as taxa not currently present in the Western gut.

Endogenous intestinal microbiota have wide-ranging and largely uncharacterized effects on host physiology. Here, we used reverse-phase liquid chromatography-coupled tandem mass spectrometry to define the mouse intestinal proteome in the stomach, jejunum, ileum, cecum and proximal colon under three colonization states: germ-free (GF), monocolonized with Bacteroides thetaiotaomicron and conventionally raised (CR). Our analysis revealed distinct proteomic abundance profiles along the gastrointestinal (GI) tract. Unsupervised clustering showed that host protein abundance primarily depended on GI location rather than colonization state and specific proteins and functions that defined these locations were identified by random forest classifications. K-means clustering of protein abundance across locations revealed substantial differences in host protein production between CR mice relative to GF and monocolonized mice. Finally, comparison with fecal proteomic data sets suggested that the identities of stool proteins are not biased to any region of the GI tract, but are substantially impacted by the microbiota in the distal colon.

SummaryGenomic technologies have significantly advanced our understanding of the composition and diversity of host-associated microbial populations. However, their spatial organization and functional interactions relative to the host have been more challenging to study. Here we present a pipeline for the assessment of intestinal microbiota localization within immunofluorescence images of fixed gut cross-sections that includes a flexible software package, BacSpace, for high-throughput quantification of microbial organization. Applying this pipeline to gnotobiotic and human microbiota-colonized mice, we demonstrate that elimination of microbiota-accessible carbohydrates (MACs) from the diet results in thinner mucus in the distal colon, increased proximity of microbes to the epithelium, and heightened expression of the inflammatory marker REG3β. Measurements of microbe-microbe proximity reveal that a MAC-deficient diet alters monophyletic spatial clustering. Furthermore, we quantify the invasion of Helicobacter pylori into the glands of the mouse stomach relative to host mitotic progenitor cells, illustrating the generalizability of this approach.

The composition of the gut microbiome in industrialized populations differs from those living traditional lifestyles. However, it has been difficult to separate the contributions of human genetic and geographic factors from lifestyle/modernization. Here, we characterize the stool bacterial composition of four Himalayan populations to investigate how the gut community changes in response to shifts in human lifestyles. These groups led seminomadic hunting-gathering lifestyles until transitioning to varying dependence upon farming. The Tharu began farming 250-300 years ago, the Raute and Raji transitioned 30-40 years ago, and the Chepang retain many aspects of a foraging lifestyle. We assess the contributions of dietary and environmental factors on their gut microbiota and find that the gut microbiome composition is significantly associated with lifestyle. The Chepang foragers harbor elevated abundance of taxa associated with foragers around the world. Conversely, the gut microbiomes of populations that have transitioned to farming are more similar to those of Americans, with agricultural dependence and several associated lifestyle and environmental factors correlating with the extent of microbiome divergence from the foraging population. For example, our results show that drinking water source and solid cooking fuel are significantly associated with the gut microbiome. Despite the pronounced differences in gut bacterial composition across populations, we found little differences in alpha diversity across populations. These findings in genetically similar populations living in the same geographical region establish the key role of lifestyle in determining human gut microbiome composition and point to the next challenging steps of isolating dietary effects from other factors that change during modernization.

The composition of the gut microbiome in industrialized populations differs from those living traditional lifestyles. However, it has been difficult to separate the contributions of human genetic and geographic factors from lifestyle. Whether shifts away from the foraging lifestyle that characterize much of humanity’s past influence the gut microbiome, and to what degree, remains unclear. Here, we characterize the stool bacterial composition of four Himalayan populations to investigate how the gut community changes in response to shifts in traditional human lifestyles. These groups led seminomadic hunting–gathering lifestyles until transitioning to varying levels of agricultural dependence upon farming. The Tharu began farming 250–300 years ago, the Raute and Raji transitioned 30–40 years ago, and the Chepang retain many aspects of a foraging lifestyle. We assess the contributions of dietary and environmental factors on their gut-associated microbes and find that differences in the lifestyles of Himalayan foragers and farmers are strongly correlated with microbial community variation. Furthermore, the gut microbiomes of all four traditional Himalayan populations are distinct from that of the Americans, indicating that industrialization may further exacerbate differences in the gut community. The Chepang foragers harbor an elevated abundance of taxa associated with foragers around the world. Conversely, the gut microbiomes of the populations that have transitioned to farming are more similar to those of Americans, with agricultural dependence and several associated lifestyle and environmental factors correlating with the extent of microbiome divergence from the foraging population. The gut microbiomes of Raute and Raji reveal an intermediate state between the Chepang and Tharu, indicating that divergence from a stereotypical foraging microbiome can occur within a single generation. Our results also show that environmental factors such as drinking water source and solid cooking fuel are significantly associated with the gut microbiome. Despite the pronounced differences in gut bacterial composition across populations, we found little differences in alpha diversity across lifestyles. These findings in genetically similar populations living in the same geographical region establish the key role of lifestyle in determining human gut microbiome composition and point to the next challenging steps of determining how large-scale gut microbiome reconfiguration impacts human biology., Author summary Although much of humanity’s history has been spent foraging in the forests, the advent of agriculture approximately 10,000 years ago and industrialization approximately 250 years ago mark major shifts in human lifestyle. Several studies have investigated the effect of industrialization on the human gut microbiome—a collection of microbes that inhabit the human gut. However, little is known about whether the gut microbiome changed as humans shifted away from foraging. To investigate how the gut community changes in response to shifts in traditional human lifestyles, we characterized the gut microbial community from four Himalayan populations representing diverse subsistence strategies. We show that the divergence of the gut microbiome from the foraging population is strongly correlated with agricultural dependence in these populations. Many of the taxa that differ across lifestyles are known to be influenced by diet, but we also demonstrate that environmental factors, such as sources of drinking water, are strongly associated with the human gut microbiome. Our findings show that both diet and environment play key roles in shaping the human gut microbiome.

Few microbes are capable of converting primary bile acids (PBAs) to secondary bile acids (SBAs), which are some of the most common intestinal metabolites. To evaluate the role of bile acids in intestinal inflammation, we performed metabolomic, bacterial, and transcriptomic profiling of stool from pouches (surgically-created “rectums”) in colectomy-treated patients with inflammatory bowel disease (IBD)—namely ulcerative colitis (UC)—versus colectomy-treated controls without IBD. We show reductions in the most abundant gut SBAs, the expression of genes needed to perform conversion of PBAs to SBAs, and number of Ruminococcaceae (one of only a few taxa known to include select bacteria able to create SBAs) present in UC compared to control pouches. In a murine colitis model, we show that reconstitution with deficient SBAs reduced intestinal inflammation. These data suggest that dysbiosis induces a SBA deficiency that may promote a pro-inflammatory state and that their restoration may treat intestinal inflammation.

Tryptamine, a tryptophan-derived monoamine similar to 5-hydroxytryptamine (5-HT), is produced by gut bacteria and is abundant in human and rodent feces. However, the physiologic effect of tryptamine in the gastrointestinal (GI) tract remains unknown. Here, we show that the biological effects of tryptamine are mediated through 5-HT(4) receptor (5-HT(4)R), a G-protein coupled receptor (GPCR) uniquely expressed in the colonic epithelium. Tryptamine increases ionic flux across the colonic epithelium and increases fluid secretion in colonoids from germ free (GF) and humanized (ex-GF colonized with human stool) mice consistent with increased intestinal secretion. The secretory effect of tryptamine is dependent on 5-HT(4)R activation and is blocked by 5-HT(4)R antagonist and absent in 5-HT(4)R(−/−) mice. GF mice colonized by Bacteroides thetaiotaomicron engineered to produce tryptamine exhibit accelerated gastrointestinal transit. Our study demonstrates an aspect of host physiology under control of a bacterial metabolite that can be exploited as a therapeutic modality.

Osmotic diarrhea is a prevalent condition in humans caused by food intolerance, malabsorption, and widespread laxative use. Here, we assess the resilience of the gut ecosystem to osmotic perturbation at multiple length and timescales using mice as model hosts. Osmotic stress caused reproducible extinction of highly abundant taxa and expansion of less prevalent members in human and mouse microbiotas. Quantitative imaging revealed decimation of the mucus barrier during osmotic perturbation, followed by recovery. The immune system exhibited temporary changes in cytokine levels and a lasting IgG response against commensal bacteria. Increased osmolality prevented growth of commensal strains in vitro, revealing one mechanism contributing to extinction. Environmental availability of microbiota members mitigated extinction events, demonstrating how species reintroduction can affect community resilience. Our findings (1) demonstrate that even mild osmotic diarrhea can cause lasting changes to the microbiota and host and (2) lay the foundation for interventions that increase system-wide resilience.

Clostridium difficile is an opportunistic diarrhoeal pathogen, and C. difficile infection (CDI) represents a major health care concern, causing an estimated 15,000 deaths per year in the United States alone 1 . Several enteric pathogens, including C. difficile, leverage inflammation and the accompanying microbial dysbiosis to thrive in the distal gut 2 . Although diet is among the most powerful available tools for affecting the health of humans and their relationship with their microbiota, investigation into the effects of diet on CDI has been limited. Here, we show in mice that the consumption of microbiota-accessible carbohydrates (MACs) found in dietary plant polysaccharides has a significant effect on CDI. Specifically, using a model of antibiotic-induced CDI that typically resolves within 12 days of infection, we demonstrate that MAC-deficient diets perpetuate CDI. We show that C. difficile burdens are suppressed through the addition of either a diet containing a complex mixture of MACs or a simplified diet containing inulin as the sole MAC source. We show that switches between these dietary conditions are coincident with changes to microbiota membership, its metabolic output and C. difficile-mediated inflammation. Together, our data demonstrate the outgrowth of MAC-utilizing taxa and the associated end products of MAC metabolism, namely, the short-chain fatty acids acetate, propionate and butyrate, are associated with decreased C. difficile fitness despite increased C. difficile toxin expression in the gut. Our findings, when placed into the context of the known fibre deficiencies of a human Western diet, provide rationale for pursuing MAC-centric dietary strategies as an alternate line of investigation for mitigating CDI. A murine diet high in microbiota-accessible carbohydrates (MACs) reduces Clostridium difficile colonization compared to a low-MAC diet, which is associated with changes in microbiota composition, short-chain fatty acid concentrations and inflammation.

The dense microbial ecosystem in the gut is intimately connected to numerous facets of human biology, and manipulation of the gut microbiota has broad implications for human health. In the absence of profound perturbation, the bacterial strains that reside within an individual are mostly stable over time1. By contrast, the fate of exogenous commensal and probiotic strains applied to an established microbiota is variable, generally unpredictable and greatly influenced by the background microbiota2,3. Therefore, analysis of the factors that govern strain engraftment and abundance is of critical importance to the emerging field of microbiome reprogramming. Here we generate an exclusive metabolic niche in mice via administration of a marine polysaccharide, porphyran, and an exogenous Bacteroides strain harbouring a rare gene cluster for porphyran utilization. Privileged nutrient access enables reliable engraftment of the exogenous strain at predictable abundances in mice harbouring diverse communities of gut microbes. This targeted dietary support is sufficient to overcome priority exclusion by an isogenic strain4, and enables strain replacement. We demonstrate transfer of the 60-kb porphyran utilization locus into a naive strain of Bacteroides, and show finely tuned control of strain abundance in the mouse gut across multiple orders of magnitude by varying porphyran dosage. Finally, we show that this system enables the introduction of a new strain into the colonic crypt ecosystem. These data highlight the influence of nutrient availability in shaping microbiota membership, expand the ability to perform a broad spectrum of investigations in the context of a complex microbiota, and have implications for cell-based therapeutic strategies in the gut. Finely tuned control of strain engraftment and abundance in the mouse gut microbiota was achieved using the marine polysaccharide porphyran, which could exclusively be used by an introduced subset of wild-type or genetically modified Bacteroides strains.

Although humans have cospeciated with their gut-resident microbes, it is difficult to infer features of our ancestral microbiome. Here, we examine the microbiome profile of 350 stool samples collected longitudinally for more than a year from the Hadza hunter-gatherers of Tanzania. The data reveal annual cyclic reconfiguration of the microbiome, in which some taxa become undetectable only to reappear in a subsequent season. Comparison of the Hadza data set with data collected from 18 populations in 16 countries with varying lifestyles reveals that gut community membership corresponds to modernization: Notably, the taxa within the Hadza that are the most seasonally volatile similarly differentiate industrialized and traditional populations. These data indicate that some dynamic lineages of microbes have decreased in prevalence and abundance in modernized populations.

Regulatory Tcells (Tregs) are required to establish immune tolerance to commensal microbes. Tregs accumulate abruptly in the skin during a defined window of postnatal tissue development. However, the mechanisms mediating Treg migration to neonatal skin are unknown. Here we show that hair follicle (HF) development facilitates the accumulation of Tregs in neonatal skin and that upon skin entry these cells localize to HFs, a primary reservoir for skin commensals. Further, germ-free neonates had reduced skin Tregs indicating that commensal microbes augment Treg accumulation. We identified Ccl20 as a HF-derived, microbiota-dependent chemokine and found its receptor, Ccr6, to be preferentially expressed by Tregs in neonatal skin. The Ccl20-Ccr6 pathway mediated Treg migration invitro and invivo. Thus, HF morphogenesis, commensal microbe colonization, and local chemokine production work in concert to recruit Tregs into neonatal skin, thereby establishing this tissue Treg niche early in life.

Summary Consumption of glucosinolates, pro-drug-like metabolites abundant in Brassica vegetables, has been associated with decreased risk of certain cancers. Gut microbiota have the ability to metabolize glucosinolates, generating chemopreventive isothiocyanates. Here, we identify a genetic and biochemical basis for activation of glucosinolates to isothiocyanates by Bacteroides thetaiotaomicron, a prominent gut commensal species. Using a genome-wide transposon insertion screen, we identified an operon required for glucosinolate metabolism in B. thetaiotaomicron. Expression of BT2159-BT2156 in a non-metabolizing relative, Bacteroides fragilis, resulted in gain of glucosinolate metabolism. We show that isothiocyanate formation requires the action of BT2158 and either BT2156 or BT2157 in vitro. Monocolonization of mice with mutant BtΔ2157 showed reduced isothiocyanate production in the gastrointestinal tract. These data provide insight into the mechanisms by which a common gut bacterium processes an important dietary nutrient.

Gut microbiota alterations have been described in several diseases with altered gastrointestinal (GI) motility, and awareness is increasing regarding the role of the gut microbiome in modulating GI function. Serotonin [5-hydroxytryptamine (5-HT)] is a key regulator of GI motility and secretion. To determine the relationship among gut microbes, colonic contractility, and host serotonergic gene expression, we evaluated mice that were germ-free (GF) or humanized (HM; ex-GF colonized with human gut microbiota). 5-HT reduced contractile duration in both GF and HM colons. Microbiota from HM and conventionally raised (CR) mice significantly increased colonic mRNAs Tph1 [(tryptophan hydroxylase) 1, rate limiting for mucosal 5-HT synthesis; P < 0.01] and chromogranin A (neuroendocrine secretion; P < 0.01), with no effect on monoamine oxidase A (serotonin catabolism), serotonin receptor 5-HT4, or mouse serotonin transporter. HM and CR mice also had increased colonic Tph1 protein (P < 0.05) and 5-HT concentrations (GF, 17 ± 3 ng/mg; HM, 25 ± 2 ng/mg; and CR, 35 ± 3 ng/mg; P < 0.05). Enterochromaffin (EC) cell numbers (cells producing 5-HT) were unchanged. Short-chain fatty acids (SCFAs) promoted TPH1 transcription in BON cells (human EC cell model). Thus, gut microbiota acting through SCFAs are important determinants of enteric 5-HT production and homeostasis.—Reigstad, C. S., Salmonson, C. E., Rainey, III, J. F., Szurszewski, J. H., Linden, D. R., Sonnenburg, J. L., Farrugia, G., Kashyap, P. C. Gut microbes promote colonic serotonin production through an effect of short-chain fatty acids on enterochromaffin cells.

Clostridium difficile is a leading cause of antibiotic-associated diarrhea. The mechanisms underlying C.difficile expansion after microbiota disturbance are just emerging. We assessed the gene expression profile of C.difficile within the intestine of gnotobiotic mice to identify genes regulated in response to either dietary or microbiota compositional changes. In the presence of the gut symbiont Bacteroides thetaiotaomicron, C.difficile induces a pathway that metabolizes the microbiota fermentation end-product succinate to butyrate. The low concentration of succinate present in the microbiota of conventional mice is transiently elevated upon antibiotic treatment or chemically induced intestinal motility disturbance, and C.difficile exploits this succinate spike to expand in the perturbed intestine. A C.difficile mutant compromised in succinate utilization is at a competitive disadvantage during these perturbations. Understanding the metabolic mechanisms involved in microbiota-C.difficile interactions may help to identify approaches for the treatment and prevention of C.difficile-associated diseases.

The gut microbiota of a healthy person may not be equivalent to a healthy microbiota. It is possible that the Western microbiota is actually dysbiotic and predisposes individuals to a variety of diseases. The asymmetric plasticity between the relatively stable human genome and the more malleable gut microbiome suggests that incompatibilities between the two could rapidly arise. The Western lifestyle, which includes a diet low in microbiota-accessible carbohydrates (MACs), has selected for a microbiota with altered membership and functionality compared to those of groups living traditional lifestyles. Interactions between resident microbes and host leading to immune dysregulation may explain several diseases that share inflammation as a common basis. The low-MAC Western diet results in poor production of gut microbiota-generated short-chain fatty acids (SCFAs), which attenuate inflammation through a variety of mechanisms in mouse models. Studies focused on modern and traditional societies, combined with animal models, are needed to characterize the connection between diet, microbiota composition, and function. Differentiating between an optimal microbiota, one that increases disease risk, and one that is causative or potentiates disease will be required to further understand both the etiology and possible treatments for health problems related to microbiota dysbiosis.