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

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

Dermal injury drives a skin to gut axis that disrupts the intestinal microbiome and intestinal immune homeostasis in mice.

The composition of the microbial community in the intestine may influence the functions of distant organs such as the brain, lung, and skin. These microbes can promote disease or have beneficial functions, leading to the hypothesis that microbes in the gut explain the co-occurrence of intestinal and skin diseases. Here, we show that the reverse can occur, and that skin directly alters the gut microbiome. Disruption of the dermis by skin wounding or the digestion of dermal hyaluronan results in increased expression in the colon of the host defense genes Reg3 and Muc2, and skin wounding changes the composition and behavior of intestinal bacteria. Enhanced expression Reg3 and Muc2 is induced in vitro by exposure to hyaluronan released by these skin interventions. The change in the colon microbiome after skin wounding is functionally important as these bacteria penetrate the intestinal epithelium and enhance colitis from dextran sodium sulfate (DSS) as seen by the ability to rescue skin associated DSS colitis with oral antibiotics, in germ-free mice, and fecal microbiome transplantation to unwounded mice from mice with skin wounds. These observations provide direct evidence of a skin-gut axis by demonstrating that damage to the skin disrupts homeostasis in intestinal host defense and alters the gut microbiome.

A serum-induced gene signature in hepatocytes is associated with pediatric nonalcoholic fatty liver disease.

OBJECTIVE: Pediatric nonalcoholic fatty liver disease (NAFLD) is a growing problem, but its underlying mechanisms are poorly understood. We used transcriptomic reporter cell assays to investigate differences in transcriptional signatures induced in hepatocyte reporter cells by the sera of children with and without NAFLD. METHODS: We studied serum samples from 45 children with NAFLD and 28 children without NAFLD. The sera were used to induce gene expression in cultured HepaRG cells and RNA-sequencing was used to determine gene expression. Computational techniques were used to compare gene expression patterns. RESULTS: Sera from children with NAFLD induced the expression of 195 genes that were significantly differentially expressed in hepatocytes compared to controls with obesity. NAFLD was associated with increased expression of genes promoting inflammation, collagen synthesis, and extracellular matrix remodeling. Additionally, there was lower expression of genes involved in endobiotic and xenobiotic metabolism, and downregulation of peroxisome function, oxidative phosphorylation, and xenobiotic, bile acid, and fatty acid metabolism. A 13-gene signature, including upregulation of TREM1 and MMP1 and downregulation of CYP2C9, was consistently associated with all diagnostic categories of pediatric NAFLD. CONCLUSION: The extracellular milieu of sera from children with NAFLD induced specific gene profiles distinguishable by a hepatocyte reporter system. Circulating factors may contribute to inflammation and extracellular matrix remodeling and impair xenobiotic and endobiotic metabolism in pediatric NAFLD.

Tuft cells mediate commensal remodeling of the small intestinal antimicrobial landscape.

Succinate produced by the commensal protist Tritrichomonas musculis (T. mu) stimulates chemosensory tuft cells, resulting in intestinal type 2 immunity. Tuft cells express the succinate receptor SUCNR1, yet this receptor does not mediate antihelminth immunity nor alter protist colonization. Here, we report that microbial-derived succinate increases Paneth cell numbers and profoundly alters the antimicrobial peptide (AMP) landscape in the small intestine. Succinate was sufficient to drive this epithelial remodeling, but not in mice lacking tuft cell chemosensory components required to detect this metabolite. Tuft cells respond to succinate by stimulating type 2 immunity, leading to interleukin-13-mediated epithelial and AMP expression changes. Moreover, type 2 immunity decreases the total number of mucosa-associated bacteria and alters the small intestinal microbiota composition. Finally, tuft cells can detect short-term bacterial dysbiosis that leads to a spike in luminal succinate levels and modulate AMP production in response. These findings demonstrate that a single metabolite produced by commensals can markedly shift the intestinal AMP profile and suggest that tuft cells utilize SUCNR1 and succinate sensing to modulate bacterial homeostasis.

Gut microbiota and metabolites drive chronic sickle cell disease pain.

Pain is a debilitating symptom and leading reason for hospitalization of individuals with sickle cell disease. Chronic sickle cell pain is poorly managed because the biological basis is not fully understood. Using transgenic sickle cell mice and fecal material transplant, we determined that the gut microbiome drives persistent sickle cell pain. In parallel patient and mouse analyses, we identified bilirubin as one metabolite that induces sickle cell pain by altering vagus nerve activity. Furthermore, we determined that decreased abundance of the gut bacteria Akkermansia mucinophila is a critical driver of chronic sickle cell pain. These experiments demonstrate that the sickle cell gut microbiome drives chronic widespread pain and identify bacterial species and metabolites that should be targeted for chronic sickle cell disease pain management.

Lactiplantibacillus plantarum 299v supplementation modulates ß-cell ER stress and antioxidative defense pathways and prevents type 1 diabetes in gluten-free BioBreeding rats.

The increasing incidence of Type 1 diabetes has coincided with the emergence of the low-fiber, high-gluten Western diet and other environmental factors linked to dysbiosis. Since Lactiplantibacillus plantarum 299 v (Lp299v) supplementation improves gut barrier function and reduces systemic inflammation, we studied its effects in spontaneously diabetic DRlyp/lyp rats provided a normal cereal diet (ND) or a gluten-free hydrolyzed casein diet (HCD). All rats provided ND developed diabetes (62.5±7.7 days); combining ND with Lp299v did not improve survival. Diabetes was delayed by HCD (72.2±9.4 days, p = .01) and further delayed by HCD+Lp299v (84.9±14.3 days, p < .001). HCD+Lp299v pups exhibited increased plasma propionate and butyrate levels, which correlated with enriched fecal Bifidobacteriaceae and Clostridiales taxa. Islet transcriptomic and histologic analyses at 40-days of age revealed that rats fed HCD expressed an autophagy profile, while those provided HCD+Lp299v expressed ER-associated protein degradation (ERAD) and antioxidative defense pathways, including Nrf2. Exposing insulinoma cells to propionate and butyrate promoted the antioxidative defense response but did not recapitulate the HCD+Lp299v islet ERAD transcriptomic profile. Here, both diet and microbiota influenced diabetes susceptibility. Moreover, Lp299v supplement modulated antioxidative defense and ER stress responses in ß-cells, potentially offering a new therapeutic direction to thwart diabetes progression and preserve insulin secretion.

Group 3 innate lymphoid cells require BATF to regulate gut homeostasis in mice.

Group 3 innate lymphoid cells (ILC3s) are crucial for the maintenance of host-microbiota homeostasis in gastrointestinal mucosal tissues. The mechanisms that maintain lineage identity of intestinal ILC3s and ILC3-mediated orchestration of microbiota and mucosal T cell immunity are elusive. Here, we identified BATF as a gatekeeper of ILC3 homeostasis in the gut. Depletion of BATF in ILC3s resulted in excessive interferon-γ production, dysbiosis, aberrant T cell immune responses, and spontaneous inflammatory bowel disease (IBD), which was considerably ameliorated by the removal of adaptive immunity, interferon-γ blockade, or antibiotic treatment. Mechanistically, BATF directly binds to the cis-regulatory elements of type 1 effector genes, restrains their chromatin accessibility, and inhibits their expression. Conversely, BATF promotes chromatin accessibility of genes involved in MHCII antigen processing and presentation pathways, which in turn directly promotes the transition of precursor ILC3s to MHCII+ ILC3s. Collectively, our findings reveal that BATF is a key transcription factor for maintaining ILC3 stability and coordinating ILC3-mediated control of intestinal homeostasis.

Probiotic normalization of systemic inflammation in siblings of type 1 diabetes patients: an open-label pilot study.

The incidence of type 1 diabetes (T1D) has increased, coinciding with lifestyle changes that have likely altered the gut microbiota. Dysbiosis, gut barrier dysfunction, and elevated systemic inflammation consistent with microbial antigen exposure, have been associated with T1D susceptibility and progression. A 6-week, single-arm, open-label pilot trial was conducted to investigate whether daily multi-strain probiotic supplementation could reduce this familial inflammation in 25 unaffected siblings of T1D patients. Probiotic supplementation was well-tolerated as reflected by high participant adherence and no adverse events. Community alpha and beta diversity were not altered between the pre- and post-supplement stool samplings. However, LEfSe analyses identified post-supplement enrichment of the family Lachnospiraceae, producers of the anti-inflammatory short chain fatty acid butyrate. Systemic inflammation was measured by plasma-induced transcription and quantified with a gene ontology-based composite inflammatory index (I.I.com). Post-supplement I.I.com was significantly reduced and pathway analysis predicted inhibition of numerous inflammatory mediators and activation of IL10RA. Subjects with the greatest post-supplement reduction in I.I.com exhibited significantly lower CD4+ CD45RO+ (memory):CD4+ CD45RA+ (naïve) T-cell ratios after supplementation. Post-supplement IL-12p40, IL-13, IL-15, IL-18, CCL2, and CCL24 plasma levels were significantly reduced, while post-supplement butyrate levels trended 1.4-fold higher. Probiotic supplementation may modify T1D susceptibility and progression and warrants further study.

Skin inflammation activates intestinal stromal fibroblasts and promotes colitis.

Inflammatory disorders of the skin are frequently associated with inflammatory bowel diseases (IBDs). To explore mechanisms by which these organs communicate, we performed single-cell RNA-Seq analysis on fibroblasts from humans and mice with IBD. This analysis revealed that intestinal inflammation promoted differentiation of a subset of intestinal stromal fibroblasts into preadipocytes with innate antimicrobial host defense activity. Furthermore, this process of reactive adipogenesis was exacerbated if mouse skin was inflamed as a result of skin wounding or infection. Since hyaluronan (HA) catabolism is activated during skin injury and fibroblast-to-adipocyte differentiation is dependent on HA, we tested the hypothesis that HA fragments could alter colon fibroblast function by targeted expression of human hyaluronidase-1 in basal keratinocytes from mouse skin. Hyaluronidase expression in the skin activated intestinal stromal fibroblasts, altered the fecal microbiome, and promoted excessive reactive adipogenesis and increased inflammation in the colon after challenge with dextran sodium sulfate. The response to digested HA was dependent on expression of TLR4 by preadipocytes. Collectively, these results suggest that the association between skin inflammation and IBD may be due to recognition by mesenchymal fibroblasts in the colon of HA released during inflammation of the skin.

Pediatric nonalcoholic fatty liver disease and the microbiome: Mechanisms contributing to pathogenesis and progression.

Nonalcoholic fatty liver disease (NAFLD) is the most common form of pediatric liver disease in the United States, and often associated with obesity and metabolic syndrome. NAFLD comprises a broad spectrum of liver diseases, from hepatic steatosis to steatohepatitis, fibrosis and cirrhosis. Disease progression is considered a multi-modal process of liver injury. The intestinal microbiome has been implicated in several aspects of NAFLD pathophysiology. Pediatric studies associating the intestinal microbiome with NAFLD have been limited in number and complicated by inconsistencies in study design and approach. Nevertheless, they provide support for involvement of the intestinal microbiome in NAFLD development and progression and point to common mechanisms shared by microbiome-associated inflammatory diseases with potential to inform future therapeutic intervention.

Lactobacillus plantarum 299v probiotic supplementation in men with stable coronary artery disease suppresses systemic inflammation.

Recent trials demonstrate that systemic anti-inflammatory therapy reduces cardiovascular events in coronary artery disease (CAD) patients. We recently demonstrated Lactobacillus plantarum 299v (Lp299v) supplementation improved vascular endothelial function in men with stable CAD. Whether this favorable effect is in part due to anti-inflammatory action remains unknown. Testing this hypothesis, we exposed plasma obtained before and after Lp299v supplementation from these subjects to a healthy donor's PBMCs and measured differences in the PBMC transciptome, performed gene ontological analyses, and compared Lp299v-induced transcriptome changes with changes in vascular function. Daily alcohol users (DAUs) (n = 4) had a significantly different response to Lp299v and were separated from the main analyses. Non-DAUs- (n = 15) showed improved brachial flow-mediated dilation (FMD) and reduced circulating IL-8, IL-12, and leptin. 997 genes were significantly changed. I.I.com decreased (1.01 ± 0.74 vs. 0.22 ± 0.51; P < 0.0001), indicating strong anti-inflammatory effects. Pathway analyses revealed downregulation of IL-1ß, interferon-stimulated pathways, and toll-like receptor signaling, and an increase in regulator T-cell (Treg) activity. Reductions in GBP1, JAK2, and TRAIL expression correlated with improved FMD. In non-DAU men with stable CAD, post-Lp299v supplementation plasma induced anti-inflammatory transcriptome changes in human PBMCs that could benefit CAD patients. Future studies should delineate changes in circulating metabolites responsible for these effects.

Microbiome Signatures Associated With Steatohepatitis and Moderate to Severe Fibrosis in Children With Nonalcoholic Fatty Liver Disease.

BACKGROUND & AIMS: The intestinal microbiome might affect the development and severity of nonalcoholic fatty liver disease (NAFLD). We analyzed microbiomes of children with and without NAFLD. METHODS: We performed a prospective, observational, cross-sectional study of 87 children (age range, 8-17 years) with biopsy-proven NAFLD and 37 children with obesity without NAFLD (controls). Fecal samples were collected and microbiome composition and functions were assessed using 16S ribosomal RNA amplicon sequencing and metagenomic shotgun sequencing. Microbial taxa were identified using zero-inflated negative binomial modeling. Genes contributing to bacterial pathways were identified using gene set enrichment analysis. RESULTS: Fecal microbiomes of children with NAFLD had lower α-diversity than those of control children (3.32 vs 3.52, P = .016). Fecal microbiomes from children with nonalcoholic steatohepatitis (NASH) had the lowest α-diversity (control, 3.52; NAFLD, 3.36; borderline NASH, 3.37; NASH, 2.97; P = .001). High abundance of Prevotella copri was associated with more severe fibrosis (P = .036). Genes for lipopolysaccharide biosynthesis were enriched in microbiomes from children with NASH (P < .001). Classification and regression tree model with level of alanine aminotransferase and relative abundance of the lipopolysaccharide pathway gene encoding 3-deoxy-d-manno-octulosonate 8-phosphate-phosphatase identified patients with NASH with an area under the receiver operating characteristic curve value of 0.92. Genes involved in flagellar assembly were enriched in the fecal microbiomes of patients with moderate to severe fibrosis (P < .001). Classification and regression tree models based on level of alanine aminotransferase and abundance of genes encoding flagellar biosynthesis protein had good accuracy for identifying case children with moderate to severe fibrosis (area under the receiver operating characteristic curve, 0.87). CONCLUSIONS: In an analysis of fecal microbiomes of children with NAFLD, we associated NAFLD and NASH with intestinal dysbiosis. NAFLD and its severity were associated with greater abundance of genes encoding inflammatory bacterial products. Alterations to the intestinal microbiome might contribute to the pathogenesis of NAFLD and be used as markers of disease or severity.

Fecal Microbial Transplant Capsules Are Safe in Hepatic Encephalopathy: A Phase 1, Randomized, Placebo-Controlled Trial.

Hepatic encephalopathy (HE) can cause major morbidity despite standard of care (SOC; rifaximin/lactulose). Fecal microbial transplant (FMT) enemas postantibiotics are safe, but the effect of FMT without antibiotics using the capsular route requires investigation. The aim of this work was to determine the safety, tolerability, and impact on mucosal/stool microbiota and brain function in HE after capsular FMT in a randomized, single-blind, placebo-controlled clinical trial in Virginia. Patients with cirrhosis with recurrent HE with MELD (Model for End-Stage Liver Disease) <17 on SOC were randomized 1:1 into receiving 15 FMT capsules versus placebo from a single donor enriched in Lachnospiraceae and Ruminococcaceae. Endoscopies with duodenal and sigmoid biopsies, stool analysis, cognition, serum lipopolysaccharide-binding protein (LBP), and duodenal antimicrobial peptide (AMP) expression at baseline were used. Clinical follow-up with SOC maintenance was performed until 5 months. FMT-assigned patients underwent repeat endoscopies 4 weeks postenrollment. Twenty subjects on lactulose/rifaximin were randomized 1:1. MELD score was similar at baseline (9.6 vs. 10.2) and study end (10.2 vs. 10.5). Six patients in the placebo group required hospitalizations compared to 1 in FMT, which was deemed unrelated to FMT. Infection/HE episodes were similar between groups. Baseline microbial diversity was similar in all tissues between groups. Post-FMT, duodenal mucosal diversity (P = 0.01) increased with higher Ruminococcaceae and Bifidobacteriaceae and lower Streptococcaceae and Veillonellaceae. Reduction in Veillonellaceae were noted post-FMT in sigmoid (P = 0.04) and stool (P = 0.05). Duodenal E-cadherin (P = 0.03) and defensin alpha 5 (P = 0.03) increased whereas interleukin-6 (P = 0.02) and serum LBP (P = 0.009) reduced post-FMT. EncephalApp performance improved post-FMT only (P = 0.02). Conclusion: In this phase 1 study, oral FMT capsules are safe and well tolerated in patients with cirrhosis and recurrent HE. FMT was associated with improved duodenal mucosal diversity, dysbiosis, and AMP expression, reduced LBP, and improved EncephalApp performance. Further studies are needed to prove efficacy.

Challenges in IBD Research: Preclinical Human IBD Mechanisms.

Preclinical human IBD mechanisms is part of five focus areas of the Challenges in IBD research document, which also include environmental triggers, novel technologies, precision medicine and pragmatic clinical research. The Challenges in IBD research document provides a comprehensive overview of current gaps in inflammatory bowel diseases (IBD) research and delivers actionable approaches to address them. It is the result of a multidisciplinary input from scientists, clinicians, patients, and funders, and represents a valuable resource for patient centric research prioritization. In particular, the preclinical human IBD mechanisms manuscript is focused on highlighting the main research gaps in the pathophysiological understanding of human IBD. These research gap areas include: 1) triggers of immune responses; 2) intestinal epithelial homeostasis and wound repair; 3) age-specific pathophysiology; 4) disease complications; 5) heterogeneous response to treatments; and 6) determination of disease location. As an approach to address these research gaps, the prioritization of reverse translation studies is proposed in which clinical observations are the foundation for experimental IBD research in the lab, and for the identification of new therapeutic targets and biomarkers. The use of human samples in validating basic research findings and development of precision medicine solutions is also proposed. This prioritization aims to put emphasis on relevant biochemical pathways and humanized in vitro and in vivo models that extrapolate meaningfully to human IBD, to eventually yield first-in-class and effective therapies.

Longitudinal changes in the gut microbiome of infants on total parenteral nutrition.

BACKGROUND: Animal studies suggest that total parenteral nutrition (TPN) may alter bacterial colonization of the intestinal tract and contribute to complications. Progressive changes in gut microbiome of infants receiving TPN are not well understood. METHODS: Infants with and without TPN/soy lipid were enrolled in a prospective, longitudinal study. Weekly fecal samples were obtained for the first 4 weeks of life. High throughput pyrosequencing of 16S rDNA was used for compositional analysis of the gut microbiome. RESULTS: 47 infants were eligible for analyses, 25 infants received TPN, and 22 infants did not (control). Although similar between TPN and control groups in the first week, fecal bacterial alpha diversity was significantly lower in the TPN group compared to controls at week 4 (Shannon index 1.0 vs 1.5, P-value = 0.03). The TPN group had significantly lower Bacteroidetes and higher Verrucomicrobia abundance compared to controls (P-values < 0.05), and these differences became more pronounced over time. At the genus level, TPN was associated with lower abundance of Bacteroides and Bifidobacterium in all weeks. CONCLUSIONS: TPN is associated with significant loss of biodiversity and alterations in the pattern of gut microbial colonization of infants over time. TPN-associated dysbiosis may predispose infants to adverse NICU outcomes.

Sortase-Dependent Proteins Promote Gastrointestinal Colonization by Enterococci.

The human gastrointestinal tract (GIT) is inhabited by a dense microbial community of symbionts. Enterococci are among the earliest members of this community and remain core members of the GIT microbiota throughout life. Enterococci have also recently emerged as opportunistic pathogens and major causes of nosocomial infections. Although recognized as a prerequisite for infection, colonization of the GIT by enterococci remains poorly understood. One way that bacteria adapt to dynamic ecosystems like the GIT is through the use of their surface proteins to sense and interact with components of their immediate environment. In Gram-positive bacteria, a subset of surface proteins relies on an enzyme called sortase for covalent attachment to the cell wall. Here, we show that the housekeeping sortase A (SrtA) enzyme promotes intestinal colonization by enterococci. Furthermore, we show that the enzymatic activity of SrtA is key to the ability of Enterococcus faecalis to bind mucin (a major component of the GIT mucus). We also report the GIT colonization phenotypes of E. faecalis mutants lacking selected sortase-dependent proteins (SDPs). Further examination of the mucin binding ability of these mutants suggests that adhesion to mucin contributes to intestinal colonization by E. faecalis.

Colonization of the mammalian intestinal tract by enterococci.

Enterococci are colonizers of the mammalian gastrointestinal tract (GIT) and normally live in healthy association with their human host. However, enterococci are also major causes of healthcare-acquired infections, prompting the US Centers for Disease Control and Prevention to declare vancomycin-resistant enterococci (VRE) a serious threat to public health. Because of both intrinsic and acquired antibiotic resistance, enterococci proliferate in the GIT during antibiotic therapy, leading to dissemination and disease. The recognition that colonization of the GIT is a pre-requisite for enterococcal infections has prompted research to study mechanisms used by enterococci to colonize this niche. This review discusses major findings of recent research to understand GIT colonization by enterococci using diverse experimental models, each of which exhibits unique strengths. This work has revealed enterococcal transcriptional reprogramming in the GIT, contributions of specific enterococcal genes encoded by the core genome to GIT colonization, the impact of genome plasticity, and roles for intra-species and inter-species interactions in modulation of GIT colonization.

Engineered E. coli Nissle 1917 for the reduction of vancomycin-resistant Enterococcus in the intestinal tract.

Vancomycin-resistant Enterococcus (VRE) poses a serious threat in hospitals where they densely colonize the intestinal tracts of patients. In vulnerable hosts, these pathogens may translocate to the bloodstream and become lethal. The ability to selectively reduce VRE in the intestinal tracts of patients could potentially prevent many of these translocation events and reduce the spread of the pathogen. Herein, we have engineered Escherichia. coli Nissle 1917 to produce and secrete three antimicrobial peptides, Enterocin A, Enterocin B, and Hiracin JM79, to specifically target and kill Enterococcus. These peptides exhibited potent activity against both Enterococcus faecium and Enterococcus faecalis, the two most prominent species responsible for VRE infections. We first discuss the optimization of the system used to express and secrete the peptides. We then show that by simultaneously expressing these peptides, both E. faecium and E. faecalis were drastically inhibited. We then demonstrate a suppression of the development of resistance when supernatant from the E. coli producer strains was used to treat E. faecium. Finally, we tested the efficacy of the probiotic in a VRE colonization model in mice. These studies showed that administration of the engineered probiotic significantly reduced the levels of both E. faecium and E. faecalis in the feces of male Balb/cJ mice.

Ceftriaxone Administration Disrupts Intestinal Homeostasis, Mediating Noninflammatory Proliferation and Dissemination of Commensal Enterococci.

Enterococci are Gram-positive commensals of the mammalian intestinal tract and harbor intrinsic resistance to broad-spectrum cephalosporins. Disruption of colonization resistance in humans by antibiotics allows enterococci to proliferate in the gut and cause disseminated infections. In this study, we used Enterococcus faecalis (EF)-colonized mice to study the dynamics of enterococci, commensal microbiota, and the host in response to systemic ceftriaxone administration. We found that the mouse model recapitulates intestinal proliferation and dissemination of enterococci seen in humans. Employing a ceftriaxone-sensitive strain of enterococci (E. faecalis JL308), we showed that increased intestinal abundance is critical for the systemic dissemination of enterococci. Investigation of the impact of ceftriaxone on the mucosal barrier defenses and integrity suggested that translocation of enterococci across the intestinal mucosa was not associated with intestinal pathology or increased permeability. Ceftriaxone-induced alteration of intestinal microbial composition was associated with transient increase in the abundance of multiple bacterial operational taxonomic units (OTUs) in addition to enterococci, for example, lactobacilli, which also disseminated to the extraintestinal organs. Collectively, these results emphasize that ceftriaxone-induced disruption of colonization resistance and alteration of mucosal homeostasis facilitate increased intestinal abundance of a limited number of commensals along with enterococci, allowing their translocation and systemic dissemination in a healthy host.

Intestinal Microbiota Disruption Reduces Regulatory T Cells and Increases Respiratory Viral Infection Mortality Through Increased IFNγ Production.

Alterations in gastrointestinal microbiota indirectly modulate the risk of atopic disease, but effects on respiratory viral infections are less clear. Using the murine paramyxoviral virus type 1, Sendai virus (SeV), we examined the effect of altering gastrointestinal microbiota on the pulmonary antiviral immune response. C57BL6 mice were treated with streptomycin before or during infection with SeV and resulting immune response studied. Ingestion of the non-absorbable antibiotic streptomycin led to a marked reduction in intestinal microbial diversity without a significant effect on lung microbiota. Reduction in diversity in the gastrointestinal tract was followed by greatly increased mortality to respiratory viral infection (p < 0.0001). This increase in mortality was associated with a dysregulated immune response characterized by decreased lung (p = 0.01) and intestinal (p = 0.03) regulatory T cells (Tregs), and increased lung IFNγ (p = 0.049), IL-6 (p = 0.015), and CCL2 (p = 0.037). Adoptive transfer of Treg cells or neutralization of IFNγ prevented increased mortality. Furthermore, Lin-CD4+ cells appeared to be a potential source of the increased IFNγ. Together, these results demonstrate gastrointestinal microbiota modulate immune responses at distant mucosal sites and have the ability to significantly impact mortality in response to a respiratory viral infection.