A novel conjugative transposon carrying an autonomously amplified plasmid.

Tetracyclines serve as broad-spectrum antibiotics to treat bacterial infections. The discovery of new tetracycline resistance genes has led to new questions about the underlying mechanisms of resistance, gene transfer, and their relevance to human health. We tracked changes in the abundance of a 55-kbp conjugative transposon (CTn214) carrying tetQ, a tetracycline resistance gene, within a Bacteroides fragilis metagenome-assembled genome derived from shotgun sequencing of microbial DNA extracted from the ileal pouch of a patient with ulcerative colitis. The mapping of metagenomic reads to CTn214 revealed the multi-copy nature of a 17,044-nt region containing tetQ in samples collected during inflammation and uninflamed visits. B. fragilis cultivars isolated from the same patient during periods of inflammation harbored CTn214 integrated into the chromosome or both a circular, multi-copy, extrachromosomal region of the CTn214 containing tetQ and the corresponding integrated form. The tetracycline-dependent mechanism for the transmission of CTn214 is nearly identical to a common conjugative transposon found in the genome of B. fragilis (CTnDOT), but the autonomously amplified nature of a circular 17,044-nt region of CTn214 that codes for tetQ and the integration of the same sequence in the linear chromosome within the same cell is a novel observation. Genome and transcriptome sequencing of B. fragilis cultivars grown under different concentrations of tetracycline and ciprofloxacin indicates that tetQ in strains containing the circular form remains actively expressed regardless of treatment, while the expression of tetQ in strains containing the linear form increases only in the presence of tetracycline.IMPORTANCEThe exchange of antibiotic production and resistance genes between microorganisms can lead to the emergence of new pathogens. In this study, short-read mapping of metagenomic samples taken over time from the illeal pouch of a patient with ulcerative colitis to a Bacteroides fragilis metagenome-assembled genome revealed two distinct genomic arrangements of a novel conjugative transposon, CTn214, that encodes tetracycline resistance. The autonomous amplification of a plasmid-like circular form from CTn214 that includes tetQ potentially provides consistent ribosome protection against tetracycline. This mode of antibiotic resistance offers a novel mechanism for understanding the emergence of pathobionts like B. fragilis and their persistence for extended periods of time in patients with inflammatory bowel disease.

Microbiota-dependent proteolysis of gluten subverts diet-mediated protection against type 1 diabetes.

Diet and commensals can affect the development of autoimmune diseases like type 1 diabetes (T1D). However, whether dietary interventions are microbe-mediated was unclear. We found that a diet based on hydrolyzed casein (HC) as a protein source protects non-obese diabetic (NOD) mice in conventional and germ-free (GF) conditions via improvement in the physiology of insulin-producing cells to reduce autoimmune activation. The addition of gluten (a cereal protein complex associated with celiac disease) facilitates autoimmunity dependent on microbial proteolysis of gluten: T1D develops in GF animals monocolonized with Enterococcus faecalis harboring secreted gluten-digesting proteases but not in mice colonized with protease deficient bacteria. Gluten digestion by E. faecalis generates T cell-activating peptides and promotes innate immunity by enhancing macrophage reactivity to lipopolysaccharide (LPS). Gnotobiotic NOD Toll4-negative mice monocolonized with E. faecalis on an HC + gluten diet are resistant to T1D. These findings provide insights into strategies to develop dietary interventions to help protect humans against autoimmunity.

Early-Life Microbial Restitution Reduces Colitis Risk Promoted by Antibiotic-Induced Gut Dysbiosis in Interleukin 10-/- Mice.

BACKGROUND & AIMS: Perturbations in the early-life gut microbiome are associated with increased risk for complex immune disorders like inflammatory bowel diseases. We previously showed that maternal antibiotic-induced gut dysbiosis vertically transmitted to offspring increases experimental colitis risk in interleukin (IL) 10 gene deficient (IL10-/-) mice, a finding that may result from the loss/lack of essential microbes needed for appropriate immunologic education early in life. Here, we aimed to identify key microbes required for proper development of the early-life gut microbiome that decrease colitis risk in genetically susceptible animals. METHODS: Metagenomic sequencing followed by reconstruction of metagenome-assembled genomes was performed on fecal samples of IL10-/- mice with and without antibiotic-induced dysbiosis to identify potential missing microbial members needed for immunologic education. One high-value target strain was then engrafted early and/or late into the gut microbiomes of IL10-/- mice with antibiotic-induced dysbiosis. RESULTS: Early-, but not late-, life engraftment of a single dominant Bacteroides strain of non-antibiotic-treated IL10-/- mice was sufficient to restore the development of the gut microbiome, promote immune tolerance, and prevent colitis in IL10-/- mice that had antibiotic-induced dysbiosis. CONCLUSIONS: Restitution of a keystone microbial strain missing in the early-life antibiotic-induced gut dysbiosis results in recovery of the microbiome, proper development of immune tolerance, and reduced risk for colitis in genetically prone hosts.

Biogeochemical dynamics and microbial community development under sulfate- and iron-reducing conditions based on electron shuttle amendment.

Iron reduction and sulfate reduction are two of the major biogeochemical processes that occur in anoxic sediments. Microbes that catalyze these reactions are therefore some of the most abundant organisms in the subsurface, and some of the most important. Due to the variety of mechanisms that microbes employ to derive energy from these reactions, including the use of soluble electron shuttles, the dynamics between iron- and sulfate-reducing populations under changing biogeochemical conditions still elude complete characterization. Here, we amended experimental bioreactors comprised of freshwater aquifer sediment with ferric iron, sulfate, acetate, and the model electron shuttle AQDS (9,10-anthraquinone-2,6-disulfonate) and monitored both the changing redox conditions as well as changes in the microbial community over time. The addition of the electron shuttle AQDS did increase the initial rate of FeIII reduction; however, it had little effect on the composition of the microbial community. Our results show that in both AQDS- and AQDS+ systems there was an initial dominance of organisms classified as Geobacter (a genus of dissimilatory FeIII-reducing bacteria), after which sequences classified as Desulfosporosinus (a genus of dissimilatory sulfate-reducing bacteria) came to dominate both experimental systems. Furthermore, most of the ferric iron reduction occurred under this later, ostensibly "sulfate-reducing" phase of the experiment. This calls into question the usefulness of classifying subsurface sediments by the dominant microbial process alone because of their interrelated biogeochemical consequences. To better inform models of microbially-catalyzed subsurface processes, such interactions must be more thoroughly understood under a broad range of conditions.

Reduction of Sb(V) by coupled biotic-abiotic processes under sulfidogenic conditions.

Increasing use and mining of antimony (Sb) has resulted in greater concern involving its fate and transport in the environment. Antimony(V) and (III) are the two most environmentally relevant oxidation states, but little is known about the redox transitions between the two in natural systems. To better understand the behavior of antimony in anoxic environments, the redox transformations of Sb(V) were studied in biotic and abiotic reactors. The biotic reactors contained Sb(V) (2 mM as KSb(OH)6), ferrihydrite (50 mM Fe(III)), sulfate (10 mM), and lactate (10 mM), that were inoculated with sediment from a wetland. In the abiotic reactors, The interaction of Sb(V) with green rust, magnetite, siderite, vivianite or mackinawite was examined under abiotic conditions. Changes in the concentrations of Sb, Fe(II), sulfate, and lactate, as well as the microbial community composition were monitored over time. Lactate was rapidly fermented to acetate and propionate in the bioreactors, with the latter serving as the primary electron donor for dissimilatory sulfate reduction (DSR). The reduction of ferrihydrite was primarily abiotic, being driven by biogenic sulfide. Sb and Fe K-edge X-ray absorption near edge structure (XANES) analysis showed reduction of Sb(V) to Sb(III) within 4 weeks, concurrent with DSR and the formation of FeS. Sb K-edge extended X-ray absorption fine structure (EXAFS) spectroscopy analysis indicated that the reduced phase was a mixture of S- and O-coordinated Sb(III). Reduction of Sb(V) was not observed in the presence of magnetite, siderite, or green rust, and limited reduction occurred with vivianite. However, reduction of Sb(V) to amorphous Sb(III) sulfide occurred with mackinawite. These results are consistent with abiotic reduction of Sb(V) by biogenic sulfide and reveal a substantial influence of Fe oxides on the speciation of Sb(III), which illustrates the tight coupling of Sb speciation with the biogeochemical cycling of S and Fe.

Metagenomic Alterations in Gut Microbiota Precede and Predict Onset of Colitis in the IL10 Gene-Deficient Murine Model.

BACKGROUND & AIMS: Inflammatory bowel diseases (IBD) are chronic inflammatory disorders where predictive biomarkers for the disease development and clinical course are sorely needed for development of prevention and early intervention strategies that can be implemented to improve clinical outcomes. Since gut microbiome alterations can reflect and/or contribute to impending host health changes, we examined whether gut microbiota metagenomic profiles would provide more robust measures for predicting disease outcomes in colitis-prone hosts. METHODS: Using the interleukin (IL) 10 gene-deficient (IL10 KO) murine model where early life dysbiosis from antibiotic (cefoperozone [CPZ]) treated dams vertically transferred to pups increases risk for colitis later in life, we investigated temporal metagenomic profiles in the gut microbiota of post-weaning offspring and determined their relationship to eventual clinical outcomes. RESULTS: Compared to controls, offspring acquiring maternal CPZ-induced dysbiosis exhibited a restructuring of intestinal microbial membership in both bacteriome and mycobiome that was associated with alterations in specific functional subsystems. Furthermore, among IL10 KO offspring from CPZ-treated dams, several functional subsystems, particularly nitrogen metabolism, diverged between mice that developed spontaneous colitis (CPZ-colitis) versus those that did not (CPZ-no-colitis) at a time point prior to eventual clinical outcome. CONCLUSIONS: Our findings provide support that functional metagenomic profiling of gut microbes has potential and promise meriting further study for development of tools to assess risk and manage human IBD.

Fecal microbiota transplant rescues mice from human pathogen mediated sepsis by restoring systemic immunity.

Death due to sepsis remains a persistent threat to critically ill patients confined to the intensive care unit and is characterized by colonization with multi-drug-resistant healthcare-associated pathogens. Here we report that sepsis in mice caused by a defined four-member pathogen community isolated from a patient with lethal sepsis is associated with the systemic suppression of key elements of the host transcriptome required for pathogen clearance and decreased butyrate expression. More specifically, these pathogens directly suppress interferon regulatory factor 3. Fecal microbiota transplant (FMT) reverses the course of otherwise lethal sepsis by enhancing pathogen clearance via the restoration of host immunity in an interferon regulatory factor 3-dependent manner. This protective effect is linked to the expansion of butyrate-producing Bacteroidetes. Taken together these results suggest that fecal microbiota transplantation may be a treatment option in sepsis associated with immunosuppression.

Polymorphic Immune Mechanisms Regulate Commensal Repertoire.

Environmental influences (infections and diet) strongly affect a host's microbiota. However, host genetics may influence commensal communities, as suggested by the greater similarity between the microbiomes of identical twins compared to non-identical twins. Variability of human genomes and microbiomes complicates the understanding of polymorphic mechanisms regulating the commensal communities. Whereas animal studies allow genetic modifications, they are sensitive to influences known as "cage" or "legacy" effects. Here, we analyze ex-germ-free mice of various genetic backgrounds, including immunodeficient and major histocompatibility complex (MHC) congenic strains, receiving identical input microbiota. The host's polymorphic mechanisms affect the gut microbiome, and both innate (anti-microbial peptides, complement, pentraxins, and enzymes affecting microbial survival) and adaptive (MHC-dependent and MHC-independent) pathways influence the microbiota. In our experiments, polymorphic mechanisms regulate only a limited number of microbial lineages (independently of their abundance). Our comparative analyses suggest that some microbes may benefit from the specific immune responses that they elicit.

Healthy infants harbor intestinal bacteria that protect against food allergy.

There has been a striking generational increase in life-threatening food allergies in Westernized societies1,2. One hypothesis to explain this rising prevalence is that twenty-first century lifestyle practices, including misuse of antibiotics, dietary changes, and higher rates of Caesarean birth and formula feeding have altered intestinal bacterial communities; early-life alterations may be particularly detrimental3,4. To better understand how commensal bacteria regulate food allergy in humans, we colonized germ-free mice with feces from healthy or cow's milk allergic (CMA) infants5. We found that germ-free mice colonized with bacteria from healthy, but not CMA, infants were protected against anaphylactic responses to a cow's milk allergen. Differences in bacterial composition separated the healthy and CMA populations in both the human donors and the colonized mice. Healthy and CMA colonized mice also exhibited unique transcriptome signatures in the ileal epithelium. Correlation of ileal bacteria with genes upregulated in the ileum of healthy or CMA colonized mice identified a clostridial species, Anaerostipes caccae, that protected against an allergic response to food. Our findings demonstrate that intestinal bacteria are critical for regulating allergic responses to dietary antigens and suggest that interventions that modulate bacterial communities may be therapeutically relevant for food allergy.

PATRIC as a unique resource for studying antimicrobial resistance.

The Pathosystems Resource Integration Center (PATRIC, www.patricbrc.org) is designed to provide researchers with the tools and services that they need to perform genomic and other 'omic' data analyses. In response to mounting concern over antimicrobial resistance (AMR), the PATRIC team has been developing new tools that help researchers understand AMR and its genetic determinants. To support comparative analyses, we have added AMR phenotype data to over 15 000 genomes in the PATRIC database, often assembling genomes from reads in public archives and collecting their associated AMR panel data from the literature to augment the collection. We have also been using this collection of AMR metadata to build machine learning-based classifiers that can predict the AMR phenotypes and the genomic regions associated with resistance for genomes being submitted to the annotation service. Likewise, we have undertaken a large AMR protein annotation effort by manually curating data from the literature and public repositories. This collection of 7370 AMR reference proteins, which contains many protein annotations (functional roles) that are unique to PATRIC and RAST, has been manually curated so that it projects stably across genomes. The collection currently projects to 1 610 744 proteins in the PATRIC database. Finally, the PATRIC Web site has been expanded to enable AMR-based custom page views so that researchers can easily explore AMR data and design experiments based on whole genomes or individual genes.

Parallelized, Aerobic, Single Carbon-Source Enrichments from Different Natural Environments Contain Divergent Microbial Communities.

Microbial communities that inhabit environments such as soil can contain thousands of distinct taxa, yet little is known about how this diversity is maintained in response to environmental perturbations such as changes in the availability of carbon. By utilizing aerobic substrate arrays to examine the effect of carbon amendment on microbial communities taken from six distinct environments (soil from a temperate prairie and forest, tropical forest soil, subalpine forest soil, and surface water and soil from a palustrine emergent wetland), we examined how carbon amendment and inoculum source shape the composition of the community in each enrichment. Dilute subsamples from each environment were used to inoculate 96-well microtiter plates containing triplicate wells amended with one of 31 carbon sources from six different classes of organic compounds (phenols, polymers, carbohydrates, carboxylic acids, amines, amino acids). After incubating each well aerobically in the dark for 72 h, we analyzed the composition of the microbial communities on the substrate arrays as well as the initial inocula by sequencing 16S rRNA gene amplicons using the Illumina MiSeq platform. Comparisons of alpha and beta diversity in these systems showed that, while the composition of the communities that grow to inhabit the wells in each substrate array diverges sharply from that of the original community in the inoculum, these enrichment communities are still strongly affected by the inoculum source. We found most enrichments were dominated by one or several OTUs most closely related to aerobes or facultative anaerobes from the Proteobacteria (e.g., Pseudomonas, Burkholderia, and Ralstonia) or Bacteroidetes (e.g., Chryseobacterium). Comparisons within each substrate array based on the class of carbon source further show that the communities inhabiting wells amended with a carbohydrate differ significantly from those enriched with a phenolic compound. Selection therefore seems to play a role in shaping the communities in the substrate arrays, although some stochasticity is also seen whereby several replicate wells within a single substrate array display strongly divergent community compositions. Overall, the use of highly parallel substrate arrays offers a promising path forward to study the response of microbial communities to perturbations in a changing environment.

Natural polyreactive IgA antibodies coat the intestinal microbiota.

Large quantities of immunoglobulin A (IgA) are constitutively secreted by intestinal plasma cells to coat and contain the commensal microbiota, yet the specificity of these antibodies remains elusive. Here we profiled the reactivities of single murine IgA plasma cells by cloning and characterizing large numbers of monoclonal antibodies. IgAs were not specific to individual bacterial taxa but rather polyreactive, with broad reactivity to a diverse, but defined, subset of microbiota. These antibodies arose at low frequencies among naïve B cells and were selected into the IgA repertoire upon recirculation in Peyer's patches. This selection process occurred independent of microbiota or dietary antigens. Furthermore, although some IgAs acquired somatic mutations, these did not substantially influence their reactivity. These findings reveal an endogenous mechanism driving homeostatic production of polyreactive IgAs with innate specificity to microbiota.

Tracking microbial colonization in fecal microbiota transplantation experiments via genome-resolved metagenomics.

BACKGROUND: Fecal microbiota transplantation (FMT) is an effective treatment for recurrent Clostridium difficile infection and shows promise for treating other medical conditions associated with intestinal dysbioses. However, we lack a sufficient understanding of which microbial populations successfully colonize the recipient gut, and the widely used approaches to study the microbial ecology of FMT experiments fail to provide enough resolution to identify populations that are likely responsible for FMT-derived benefits. METHODS: We used shotgun metagenomics together with assembly and binning strategies to reconstruct metagenome-assembled genomes (MAGs) from fecal samples of a single FMT donor. We then used metagenomic mapping to track the occurrence and distribution patterns of donor MAGs in two FMT recipients. RESULTS: Our analyses revealed that 22% of the 92 highly complete bacterial MAGs that we identified from the donor successfully colonized and remained abundant in two recipients for at least 8 weeks. Most MAGs with a high colonization rate belonged to the order Bacteroidales. The vast majority of those that lacked evidence of colonization belonged to the order Clostridiales, and colonization success was negatively correlated with the number of genes related to sporulation. Our analysis of 151 publicly available gut metagenomes showed that the donor MAGs that colonized both recipients were prevalent, and the ones that colonized neither were rare across the participants of the Human Microbiome Project. Although our dataset showed a link between taxonomy and the colonization ability of a given MAG, we also identified MAGs that belong to the same taxon with different colonization properties, highlighting the importance of an appropriate level of resolution to explore the functional basis of colonization and to identify targets for cultivation, hypothesis generation, and testing in model systems. CONCLUSIONS: The analytical strategy adopted in our study can provide genomic insights into bacterial populations that may be critical to the efficacy of FMT due to their success in gut colonization and metabolic properties, and guide cultivation efforts to investigate mechanistic underpinnings of this procedure beyond associations.

Mutual reinforcement of pathophysiological host-microbe interactions in intestinal stasis models.

Chronic diseases arise when there is mutual reinforcement of pathophysiological processes that cause an aberrant steady state. Such a sequence of events may underlie chronic constipation, which has been associated with dysbiosis of the gut. In this study we hypothesized that assemblage of microbial communities, directed by slow gastrointestinal transit, affects host function in a way that reinforces constipation and further maintains selection on microbial communities. In our study, we used two models - an opioid-induced constipation model in mice, and a humanized mouse model where germ-free mice were colonized with stool from a patient with constipation-predominant irritable bowel syndrome (IBS-C) in humans. We examined the impact of pharmacologically (loperamide)-induced constipation (PIC) and IBS-C on the structural and functional profile of the gut microbiota. Germ-free (GF) mice were colonized with microbiota from PIC donor mice and IBS-C patients to determine how the microbiota affects the host. PIC and IBS-C promoted changes in the gut microbiota, characterized by increased relative abundance of Bacteroides ovatus and Parabacteroides distasonis in both models. PIC mice exhibited decreased luminal concentrations of butyrate in the cecum and altered metabolic profiles of the gut microbiota. Colonization of GF mice with PIC-associated mice cecal or human IBS-C fecal microbiota significantly increased GI transit time when compared to control microbiota recipients. IBS-C-associated gut microbiota also impacted colonic contractile properties. Our findings support the concept that constipation is characterized by disease-associated steady states caused by reinforcement of pathophysiological factors in host-microbe interactions.

Interleukin-15 promotes intestinal dysbiosis with butyrate deficiency associated with increased susceptibility to colitis.

Dysbiosis resulting in gut-microbiome alterations with reduced butyrate production are thought to disrupt intestinal immune homeostasis and promote complex immune disorders. However, whether and how dysbiosis develops before the onset of overt pathology remains poorly defined. Interleukin-15 (IL-15) is upregulated in distressed tissue and its overexpression is thought to predispose susceptible individuals to and have a role in the pathogenesis of celiac disease and inflammatory bowel disease (IBD). Although the immunological roles of IL-15 have been largely studied, its potential impact on the microbiota remains unexplored. Analysis of 16S ribosomal RNA-based inventories of bacterial communities in mice overexpressing IL-15 in the intestinal epithelium (villin-IL-15 transgenic (v-IL-15tg) mice) shows distinct changes in the composition of the intestinal bacteria. Although some alterations are specific to individual intestinal compartments, others are found across the ileum, cecum and feces. In particular, IL-15 overexpression restructures the composition of the microbiota with a decrease in butyrate-producing bacteria that is associated with a reduction in luminal butyrate levels across all intestinal compartments. Fecal microbiota transplant experiments of wild-type and v-IL-15tg microbiota into germ-free mice further indicate that diminishing butyrate concentration observed in the intestinal lumen of v-IL-15tg mice is the result of intrinsic alterations in the microbiota induced by IL-15. This reconfiguration of the microbiota is associated with increased susceptibility to dextran sodium sulfate-induced colitis. Altogether, this study reveals that IL-15 impacts butyrate-producing bacteria and lowers butyrate levels in the absence of overt pathology, which represent events that precede and promote intestinal inflammatory diseases.

Patient-Specific Bacteroides Genome Variants in Pouchitis.

A 2-year longitudinal microbiome study of 22 patients who underwent colectomy with an ileal pouch anal anastomosis detected significant increases in distinct populations of Bacteroides during 9 of 11 patient visits that coincided with inflammation (pouchitis). Oligotyping and metagenomic short-read annotation identified Bacteroides populations that occurred in early samples, bloomed during inflammation, and reappeared after antibiotic treatment. Targeted cultivation of Bacteroides isolates from the same individual at multiple time points and from several patients detected subtle genomic changes, including the identification of rapidly evolving genomic elements that differentiate isogenic strains of Bacteroides fragilis from the mucosa versus lumen. Each patient harbored Bacteroides spp. that are closely related to commonly occurring clinical isolates, including Bacteroides ovatus, B. thetaiotaomicron, B. vulgatus, and B. fragilis, which contained unique loci in different patients for synthesis of capsular polysaccharides. The presence of unique Bacteroides capsular polysaccharide loci within different hosts and between the lumen and mucosa may represent adaptations to stimulate, suppress, and evade host-specific immune responses at different microsites of the ileal pouch. IMPORTANCE: This longitudinal study provides an opportunity to describe shifts in the microbiomes of individual patients who suffer from ulcerative colitis (UC) prior to and following inflammation. Pouchitis serves as a model for UC with a predictable incidence of disease onset and enables prospective longitudinal investigations of UC etiology prior to inflammation. Because of insufficient criteria for predicting which patients will develop UC or pouchitis, the interpretation of cross-sectional study designs suffers from lack of information about the microbiome structure and host gene expression patterns that directly correlate with the onset of disease. Our unique longitudinal study design allows each patient to serve as their own control, providing information about the state of the microbiome and host prior to and during the course of disease. Of significance to the broader community, this study identifies microbial strains that may have genetic elements that trigger the onset of disease in susceptible hosts.

Transplanting a Microbial Organ: the Good, the Bad, and the Unknown.

Fecal microbiota transplantation (FMT) has received increased attention as a therapy for correcting intestinal dysbiosis and restoring a state of health in patients suffering from either recalcitrant infection by Clostridium difficile or more complex disease states, such as inflammatory bowel disease (IBD). The "gut microbial organ" from the donor that is used in these transplants may serve to transfer genetic material between donor and recipient via virus-like particles, specifically bacteriophages, that infect the bacterial component of the microbiota. The recently published study by Chehoud et al. provides evidence for not only the transfer of bacteriophages during FMT but also the transfer of multiple populations of bacteriophages to recipients from the donor microbiota used (C. Chehoud et al., mBio 7:e00322-16, 2016, http://dx.doi.org/10.1128/mBio.00322-16). While the clinical significance of these findings remains unclear, nothing short of a diligent and persistent effort is needed to define the intended and unintended consequences of FMT.

Insights into the pathogenesis of ulcerative colitis from a murine model of stasis-induced dysbiosis, colonic metaplasia, and genetic susceptibility.

Gut dysbiosis, host genetics, and environmental triggers are implicated as causative factors in inflammatory bowel disease (IBD), yet mechanistic insights are lacking. Longitudinal analysis of ulcerative colitis (UC) patients following total colectomy with ileal anal anastomosis (IPAA) where >50% develop pouchitis offers a unique setting to examine cause vs. effect. To recapitulate human IPAA, we employed a mouse model of surgically created blind self-filling (SFL) and self-emptying (SEL) ileal loops using wild-type (WT), IL-10 knockout (KO) (IL-10), TLR4 KO (T4), and IL-10/T4 double KO mice. After 5 wk, loop histology, host gene/protein expression, and bacterial 16s rRNA profiles were examined. SFL exhibit fecal stasis due to directional motility oriented toward the loop end, whereas SEL remain empty. In WT mice, SFL, but not SEL, develop pouchlike microbial communities without accompanying active inflammation. However, in genetically susceptible IL-10-deficient mice, SFL, but not SEL, exhibit severe inflammation and mucosal transcriptomes resembling human pouchitis. The inflammation associated with IL-10 required TLR4, as animals lacking both pathways displayed little disease. Furthermore, germ-free IL-10 mice conventionalized with SFL, but not SEL, microbiota populations develop severe colitis. These data support essential roles of stasis-induced, colon-like microbiota, TLR4-mediated colonic metaplasia, and genetic susceptibility in the development of pouchitis and possibly UC. However, these factors by themselves are not sufficient. Similarities between this model and human UC/pouchitis provide opportunities for gaining insights into the mechanistic basis of IBD and for identification of targets for novel preventative and therapeutic interventions.

Spatial scale drives patterns in soil bacterial diversity.

Soil microbial communities are essential for ecosystem function, but linking community composition to biogeochemical processes is challenging because of high microbial diversity and large spatial variability of most soil characteristics. We investigated soil bacterial community structure in a switchgrass stand planted on soil with a history of grassland vegetation at high spatial resolution to determine whether biogeographic trends occurred at the centimeter scale. Moreover, we tested whether such heterogeneity, if present, influenced community structure within or among ecosystems. Pronounced heterogeneity was observed at centimeter scales, with abrupt changes in relative abundance of phyla from sample to sample. At the ecosystem scale (> 10 m), however, bacterial community composition and structure were subtly, but significantly, altered by fertilization, with higher alpha diversity in fertilized plots. Moreover, by comparing these data with data from 1772 soils from the Earth Microbiome Project, it was found that 20% of bacterial taxa were shared between their site and diverse globally sourced soil samples, while grassland soils shared approximately 40% of their operational taxonomic units with the current study. By spanning several orders of magnitude, the analysis suggested that extreme patchiness characterized community structure at smaller scales but that coherent patterns emerged at larger length scales.

Genetic and Metabolic Signals during Acute Enteric Bacterial Infection Alter the Microbiota and Drive Progression to Chronic Inflammatory Disease.

Chronic inflammatory disorders are thought to arise due to an interplay between predisposing host genetics and environmental factors. For example, the onset of inflammatory bowel disease is associated with enteric proteobacterial infection, yet the mechanistic basis for this association is unclear. We have shown previously that genetic defiency in TLR1 promotes acute enteric infection by the proteobacteria Yersinia enterocolitica. Examining that model further, we uncovered an altered cellular immune response that promotes the recruitment of neutrophils which in turn increases metabolism of the respiratory electron acceptor tetrathionate by Yersinia. These events drive permanent alterations in anti-commensal immunity, microbiota composition, and chronic inflammation, which persist long after Yersinia clearence. Deletion of the bacterial genes involved in tetrathionate respiration or treatment using targeted probiotics could prevent microbiota alterations and inflammation. Thus, acute infection can drive long term immune and microbiota alterations leading to chronic inflammatory disease in genetically predisposed individuals.