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Background: Environmental enteric dysfunction (EED) is an enigmatic disorder of the small intestine postulated to play a role in childhood undernutrition, a pressing global health problem. Defining the incidence of EED, its pathophysiology, and its contribution to impaired linear and ponderal growth has been hampered by the difficulty in directly sampling the small intestinal mucosa and microbial community (microbiota).Methods: Slum-dwelling Bangladeshi children aged 18±2 months, with linear growth-faltering (stunting) who failed a nutritional intervention underwent endoscopy to obtain duodenal biopsies and aspirates. Levels of 4077 plasma proteins and 2619 duodenal proteins were quantified in 80 children with histopathologic evidence of EED, and the abundances of bacterial strains in their duodenal microbiota were determined using culture-independent methods. Young germ-free mice, fed a Bangladeshi diet, were colonized with bacterial strains cultured from the duodenal aspirates.Results: The absolute abundances of a shared group of 14 bacterial strains recovered from the duodenums of children with EED and not typically classified as enteropathogens were negatively correlated with linear growth (length-for-age Z-score;β=-0.38±0.12(SEM);ρ=-0.49;p=0.003), and positively correlated with duodenal proteins involved in immunoinflammatory responses. Representation of these 14 duodenal taxa was significantly different in fecal microbiota from EED versus healthy children (p<0.001;PERMANOVA). Gnotobiotic mice colonized with cultured EED-donor duodenal strains develop a small intestinal enteropathy.Conclusions: These results provide evidence of a causal relationship between components of the small intestinal microbiota, enteropathy and stunting and offer a rationale for developing therapeutics that target what must no longer remain terra incognita-the small intestinal microbiota. ClinicalTrials.gov identifier: NCT02812615. [ABSTRACT FROM AUTHOR]

Background: The microbiome regulates the respiratory epithelium's immunomodulatory functions. To explore how the microbiome's biodiversity affects microbe-epithelial interactions, we screened 58 phylogenetically diverse microbes for their transcriptomic effect on human primary bronchial air-liquid interface (ALI) cell cultures., Results: We found distinct species- and strain-level differences in host innate immunity and epithelial barrier response. Strikingly, we found that host interferon, an antiviral response, was one of the most variable host processes. This variability was not driven by microbial phylogenetic diversity, bioburden, nor by the microbe's ability to stimulate other innate immunity pathways., Conclusions: Microbial colonization differentially stimulates host gene expression with variations observed across phylogenetically diverse microbes and across different strains of the same species. Our study provides a foundation for understanding how the respiratory microbiome's biodiversity affects epithelial, and particularly antiviral, innate immunity., Competing Interests: Declarations. Ethics approval and consent to participate: All genomic sequencing was performed in the context of the U19AI142733 grant at the Jackson Laboratory and has been approved by the Human Subjects Institutional Review Board at Hartford Hospital, Hartford, CT, USA (IRB #E-HHC-2019–0048). Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Salmonella enterica can persist asymptomatically within tissues for extended periods. This remarkable feat is achieved through intricate host-pathogen interactions in immune cell aggregates called granulomas, wherein Salmonella find favorable cellular niches to exploit while the host limits its expansion and tissue dissemination. Here, using a mouse model of persistent Salmonella infection, we identify a host-protective role of eosinophils in control of Salmonella Typhimurium ( S Tm) infection within the mesenteric lymph nodes (MLN), the main lymphoid tissue of S Tm persistence. Combining spatial transcriptomics and experimental manipulations, we found that macrophages responding to S Tm infection recruited eosinophils in a C-C motif chemokine ligand 11 (CCL11)-dependent manner and enhanced their activation. Eosinophil deficiencies increased Salmonella burdens, which was associated with altered granuloma size and impaired type-1 immunity in the MLN. Thus, eosinophils play a vital role in restraining Salmonella exploitation of granuloma macrophages at a key site of bacterial persistence., Competing Interests: Additional Declarations: There is NO Competing Interest.

Environmental enteric dysfunction (EED), a small intestinal disorder found at a high prevalence in stunted children, is associated with gut mucosal barrier disruption and decreased absorptive capacity due to reduced intact small intestinal villi 1-4 . To test the hypothesis that intergenerational transmission of a perturbed small intestinal microbiota contributes to undernutrition by inducing EED 5 , we characterized two consortia of bacterial strains cultured from duodenal aspirates from stunted Bangladeshi children with EED - one of which induced local and systemic inflammation in gnotobiotic female mice. Offspring of dams that received this inflammatory consortium exhibited immunologic changes along their gut that phenocopied features of EED in children. Single nucleus plus bulk RNA-sequencing revealed alterations in inter-cellular signaling pathways related to intestinal epithelial cell renewal, barrier integrity and immune function while analyses of cerebral cortex disclosed alterations in glial- and endothelial-neuronal signaling pathways that regulate neural growth/axonal guidance, angiogenesis and inflammation. Analysis of ultrasonic vocalization calls in gnotobiotic P5-P9 pups indicated increased arousal and perturbed neurodevelopment in the offspring of dams harboring the inflammation-inducing consortium. Cohousing experiments and follow-up screening of candidate disease-promoting bacterial isolates identified a strain typically found in the oral microbiota ( Campylobacter concisus ) as a contributor to enteropathy. Given that fetal growth was also impaired in the dams with the consortium that induced enteropathy, this preclinical model allows the effects of the human small intestinal microbiota on both pre- and postnatal development to be ascertained, setting the stage for identification of small intestinal microbiota-targeted therapeutics for (intergenerational) undernutrition., Competing Interests: Competing Interests - D.R. and A.O. are co-founders of Phenobiome Inc., a company pursuing development of computational tools for predictive phenotype profiling of microbial communities.

Macrophages provide a crucial environment for Salmonella enterica serovar Typhi (S. Typhi) to multiply during typhoid fever, yet our understanding of how human macrophages and S. Typhi interact remains limited. In this study, we delve into the dynamics of S. Typhi replication within human macrophages and the resulting heterogeneous transcriptomic responses of macrophages during infection. Our study reveals key factors that influence macrophage diversity, uncovering distinct immune and metabolic pathways associated with different stages of S. Typhi intracellular replication in macrophages. Of note, we found that macrophages harboring replicating S. Typhi are skewed towards an M1 pro-inflammatory state, whereas macrophages containing non-replicating S. Typhi exhibit neither a distinct M1 pro-inflammatory nor M2 anti-inflammatory state. Additionally, macrophages with replicating S. Typhi were characterized by the increased expression of genes associated with STAT3 phosphorylation and the activation of the STAT3 transcription factor. Our results shed light on transcriptomic pathways involved in the susceptibility of human macrophages to intracellular S. Typhi replication, thereby providing crucial insight into host phenotypes that restrict and support S. Typhi infection., (© 2024. The Author(s).)

The gut microbiota and tumor-associated macrophages (TAMs) affect tumor responses to anti-programmed cell death protein 1 (PD-1) immune checkpoint blockade. Reprogramming TAM by either blocking or deleting the macrophage receptor triggering receptor on myeloid cells 2 (TREM2) attenuates tumor growth, and lack of functional TREM2 enhances tumor elimination by anti-PD-1. Here, we found that anti-PD-1 treatment combined with TREM2 deficiency in mice induces proinflammatory programs in intestinal macrophages and a concomitant expansion of Ruminococcus gnavus in the gut microbiota. Gavage of wild-type mice with R. gnavus enhanced anti-PD-1-mediated tumor elimination, recapitulating the effect occurring in the absence of TREM2. A proinflammatory intestinal environment coincided with expansion, increased circulation, and migration of TNF-producing CD4 + T cells to the tumor bed. Thus, TREM2 remotely controls anti-PD-1 immune checkpoint blockade through modulation of the intestinal immune environment and microbiota, with R. gnavus emerging as a potential probiotic agent for increasing responsiveness to anti-PD-1.

Interleukin 22 (IL-22) promotes intestinal barrier integrity, stimulating epithelial cells to enact defense mechanisms against enteric infections, including the production of antimicrobial peptides. IL-22 binding protein (IL-22BP) is a soluble decoy encoded by the Il22ra2 gene that decreases IL-22 bioavailability, attenuating IL-22 signaling. The impact of IL-22BP on gut microbiota composition and functioning is poorly understood. We found that Il22ra2 -/- mice are better protected against Clostridioides difficile and Citrobacter rodentium infections. This protection relied on IL-22-induced antimicrobial mechanisms before the infection occurred, rather than during the infection itself. Indeed, the gut microbiota of Il22ra2 -/- mice mitigated infection of wild-type (WT) mice when transferred via cohousing or by cecal microbiota transplantation. Indicator species analysis of WT and Il22ra2 -/- mice with and without cohousing disclosed that IL22BP deficiency yields a gut bacterial composition distinct from that of WT mice. Manipulation of dietary fiber content, measurements of intestinal short-chain fatty acids and oral treatment with acetate disclosed that resistance to C. difficile infection is related to increased production of acetate by Il22ra2 -/- -associated microbiota. Together, these findings suggest that IL-22BP represents a potential therapeutic target for those at risk for or with already manifest infection with this and perhaps other enteropathogens., Competing Interests: Competing interests statement:The authors declare no competing interest.

The aryl-hydrocarbon receptor (AHR) is a ligand-activated transcription factor that buoys intestinal immune responses. AHR induces its own negative regulator, the AHR repressor (AHRR). Here, we show that AHRR is vital to sustaining intestinal intraepithelial lymphocytes (IELs). AHRR deficiency reduced IEL representation in a cell-intrinsic fashion. Single-cell RNA sequencing revealed an oxidative stress profile in Ahrr -/- IELs. AHRR deficiency unleashed AHR-induced expression of CYP1A1, a monooxygenase that generates reactive oxygen species, increasing redox imbalance, lipid peroxidation, and ferroptosis in Ahrr -/- IELs. Dietary supplementation with selenium or vitamin E to restore redox homeostasis rescued Ahrr -/- IELs. Loss of IELs in Ahrr -/- mice caused susceptibility to Clostridium difficile infection and dextran sodium-sulfate-induced colitis. Inflamed tissue of inflammatory bowel disease patients showed reduced Ahrr expression that may contribute to disease. We conclude that AHR signaling must be tightly regulated to prevent oxidative stress and ferroptosis of IELs and to preserve intestinal immune responses., Competing Interests: Declaration of interests M. Colonna receives research support from Pfizer., (Copyright © 2023 Elsevier Inc. All rights reserved.)

The gut microbiota exerts a variety of positive effects on the intestinal homeostasis, including the production of beneficial molecules, control of the epithelial barrier integrity and the regulation of the balance between host's cell death and proliferation. The interactions between commensal bacteria and intestinal cells are still under-investigated and is then of paramount importance to address such interactions at the molecular and cellular levels. We report an in vitro analysis of the effects of molecules secreted by Lactobacillus gasseri SF1183 on HCT116 cells, selected as a model of intestinal epithelial cells. SF1183 is a L. gasseri strain isolated from an ileal biopsy of a human healthy volunteer, able to prevent colitis symptoms in vivo. Expanding previous findings, we show that bioactive molecules secreted by SF1183 reduce the proliferation of HCT116 cells in a reversible manner determining a variation in cell cycle markers (p21WAF, p53, cyclin D1) and resulting in the protection of HCT116 cells from TNF-alfa induced apoptosis, an effect potentially relevant for the protection of the epithelial barrier integrity and reconstitution of tissue homeostasis. Consistently, SF1183 secreted molecules increase the recruitment of occludin, a major component of TJ, at the cell-cell contacts, suggesting a reinforcement of the barrier function., (© 2022. The Author(s).)

C-type lectin domain family 4, member a4 (Clec4a4) is a C-type lectin inhibitory receptor specific for glycans thought to be exclusively expressed on murine CD8α− conventional dendritic cells. Using newly generated Clec4a4-mCherry knock-in mice, we identify a subset of Clec4a4-expressing eosinophils uniquely localized in the small intestine lamina propria. Clec4a4+ eosinophils evinced an immunomodulatory signature, whereas Clec4a4− eosinophils manifested a proinflammatory profile. Clec4a4+ eosinophils expressed high levels of aryl hydrocarbon receptor (Ahr), which drove the expression of Clec4a4 as well as other immunomodulatory features, such as PD-L1. The abundance of Clec4a4+ eosinophils was dependent on dietary AHR ligands, increased with aging, and declined in inflammatory conditions. Mice lacking AHR in eosinophils expanded innate lymphoid cells of type 2 and cleared Nippostrongylus brasiliensis infection more effectively than did wild-type mice. These results highlight the heterogeneity of eosinophils in response to tissue cues and identify a unique AHR-dependent subset of eosinophils in the small intestine with an immunomodulatory profile.

Innate lymphocytes encompass a diverse array of phenotypic identities with specialized functions. DNA methylation and hydroxymethylation are essential for epigenetic fidelity and fate commitment. The landscapes of these modifications are unknown in innate lymphocytes. Here, we characterized the whole-genome distribution of methyl-CpG and 5-hydroxymethylcytosine (5hmC) in mouse innate lymphoid cell 3 (ILC3), ILC2 and natural killer (NK) cells. We identified differentially methylated regions (DMRs) and differentially hydroxymethylated regions (DHMRs) between ILC and NK cell subsets and correlated them with transcriptional signatures. We associated lineage-determining transcription factors (LDTFs) with demethylation and demonstrated unique patterns of DNA methylation/hydroxymethylation in relationship to open chromatin regions (OCRs), histone modifications and TF-binding sites. We further identified an association between hydroxymethylation and NK cell superenhancers (SEs). Using mice lacking the DNA hydroxymethylase TET2, we showed the requirement for TET2 in optimal production of hallmark cytokines by ILC3s and interleukin-17A (IL-17A) by inflammatory ILC2s. These findings provide a powerful resource for studying innate lymphocyte epigenetic regulation and decode the regulatory logic governing their identity., (© 2022. Springer Nature America, Inc.)

Lymphoid tissue inducer (LTi)-like cells are tissue resident innate lymphocytes that rapidly secrete cytokines that promote gut epithelial integrity and protect against extracellular bacterial infections.Here, we report that the retention of LTi-like cells in conventional solitary intestinal lymphoid tissue (SILT) is essential for controlling LTi-like cell function and is maintained by expression of the chemokine receptor CXCR5. Deletion of Cxcr5 functionally unleashed LTi-like cells in a cell intrinsic manner, leading to uncontrolled IL-17 and IL-22 production. The elevated production of IL-22 in Cxcr5 -deficient mice improved gut barrier integrity and protected mice during infection with the opportunistic pathogen Clostridium difficile Interestingly, Cxcr5 -/- mice developed LTi-like cell aggregates that were displaced from their typical niche at the intestinal crypt, and LTi-like cell hyperresponsiveness was associated with the local formation of this unconventional SILT. Thus, LTi-like cell positioning within mucosa controls their activity via niche-specific signals that temper cytokine production during homeostasis., Competing Interests: Competing interest statement: M. Colonna receives research support from Pfizer.

Crohn's disease (CD) is a chronic transmural inflammation of intestinal segments caused by dysregulated interaction between microbiome and gut immune system. Here, we profile, via multiple single-cell technologies, T cells purified from the intestinal epithelium and lamina propria (LP) from terminal ileum resections of adult severe CD cases. We find that intraepithelial lymphocytes (IEL) contain several unique T cell subsets, including NKp30 + γδT cells expressing RORγt and producing IL-26 upon NKp30 engagement. Further analyses comparing tissues from non-inflamed and inflamed regions of patients with CD versus healthy controls show increased activated T H 17 but decreased CD8 + T, γδT, T FH and Treg cells in inflamed tissues. Similar analyses of LP find increased CD8 + , as well as reduced CD4 + T cells with an elevated T H 17 over Treg/T FH ratio. Our analyses of CD tissues thus suggest a potential link, pending additional validations, between transmural inflammation, reduced IEL γδT cells and altered spatial distribution of IEL and LP T cell subsets.

Undernourished children in low-income countries often exhibit poor responses to oral vaccination. Perturbed microbiota development is linked to undernutrition, but whether and how microbiota changes affect vaccine responsiveness remains unclear. Here, we show that gnotobiotic mice colonized with microbiota from undernourished Bangladeshi children and fed a Bangladeshi diet exhibited microbiota-dependent differences in mucosal IgA responses to oral vaccination with cholera toxin (CT). Supplementation with a nutraceutical consisting of spirulina, amaranth, flaxseed, and micronutrients augmented CT-IgA production. Mice initially colonized with a microbiota associated with poor CT responses exhibited improved immunogenicity upon invasion of bacterial taxa from cagemates colonized with a more "responsive" microbiota. Additionally, a consortium of five cultured bacterial invaders conferred augmented CT-IgA responses in mice fed the supplemented diet and colonized with the "hypo-responsive" community. These results provide preclinical proof-of-concept that diet and microbiota influence mucosal immune responses to CT vaccination and identify a candidate synbiotic formulation., Competing Interests: Declaration of Interests J.I.G. is a co-founder of Matatu, Inc., a company characterizing the role of diet-by-microbiota interactions in animal health. N.F.D is an employee of the GSK group of companies and reports ownership of GSK shares and/or restricted GSK shares. GSK and Washington University have filed a joint patent application describing the potential utility of the prebiotic and probiotic formulations., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Antibiotic-induced dysbiosis is a key predisposing factor for Clostridium difficile infections (CDIs), which cause intestinal disease ranging from mild diarrhea to pseudomembranous colitis. Here, we examined the impact of a microbiota-derived metabolite, short-chain fatty acid acetate, on an acute mouse model of CDI. We found that administration of acetate is remarkably beneficial in ameliorating disease. Mechanistically, we show that acetate enhances innate immune responses by acting on both neutrophils and ILC3s through its cognate receptor free fatty acid receptor 2 (FFAR2). In neutrophils, acetate-FFAR2 signaling accelerates their recruitment to the inflammatory sites, facilitates inflammasome activation, and promotes the release of IL-1β; in ILC3s, acetate-FFAR2 augments expression of the IL-1 receptor, which boosts IL-22 secretion in response to IL-1β. We conclude that microbiota-derived acetate promotes host innate responses to C. difficile through coordinate action on neutrophils and ILC3s., Competing Interests: Disclosures: The authors declare no competing interests exist., (© 2019 Fachi et al.)

Although innate lymphoid cells (ILCs) functionally analogous to T helper type 1 (Th1), Th2, and Th17 cells are well characterized, an ILC subset strictly equivalent to IL-10-secreting regulatory T cells has only recently been proposed. Here, we report the absence of an intestinal regulatory ILC population distinct from group 1 ILCs (ILC1s), ILC2s, and ILC3s in (1) mice bred in our animal facility; (2) mice from The Jackson Laboratory, Taconic Biosciences, and Charles River Laboratories; and (3) mice subjected to intestinal inflammation. Instead, a low percentage of intestinal ILC2s produced IL-10 at steady state. A screen for putative IL-10 elicitors revealed that IL-2, IL-4, IL-27, IL-10, and neuromedin U (NMU) increased IL-10 production in activated intestinal ILC2s, while TL1A suppressed IL-10 production. Secreted IL-10 further induced IL-10 production in ILC2s through a positive feedback loop. In summary, ILC2s provide an inducible source of IL-10 in the gastrointestinal tract, whereas ILCregs are not a generalizable immune cell population in mice., (© 2019 Bando et al.)

Group 3 innate lymphoid cells (ILC3s) are the innate counterparts of Th17 that require the transcription factor RORγt for development and contribute to the defense against pathogens through IL-22 and IL-17 secretion. Proliferation and effector functions of Th17 require a specific mTOR-dependent metabolic program that utilizes high-rate glycolysis, while mitochondrial lipid oxidation and production of reactive oxygen species (mROS) support alternative T reg cell differentiation. Whether ILC3s employ a specific metabolic program is not known. Here, we find that ILC3s rely on mTOR complex 1 (mTORC1) for proliferation and production of IL-22 and IL-17A after in vitro activation and Citrobacter rodentium infection. mTORC1 induces activation of HIF1α, which reprograms ILC3 metabolism toward glycolysis and sustained expression of RORγt. However, in contrast to Th17, ILC3 activation requires mROS production; rather than inducing an alternative regulatory fate as it does in CD4 T cells, mROS stabilizes HIF1α and RORγt in ILC3s and thereby promotes their activation. We conclude that ILC3 activation relies on a metabolic program that integrates glycolysis with mROS production., (© 2019 Di Luccia et al.)

The gastrointestinal tract hosts the largest collection of commensal microbes in the body. Infections at this site can cause significant perturbations in the microbiota, known as dysbiosis, that facilitate the expansion of pathobionts, and can elicit inappropriate immune responses that impair the intestinal barrier function. Dysbiosis typically occurs during intestinal infection with Toxoplasma gondii . Host resistance to T. gondii depends on a potent Th1 response. In addition, a Th17 response is also elicited. How Th17 cells contribute to the host response to T. gondii remains unclear. Here we show that class I-restricted T cell-associated molecule (CRTAM) expression on T cells is required for an optimal IL-17 production during T. gondii infection. Moreover, that the lack of IL-17, results in increased immunopathology caused by an impaired antimicrobial peptide production and bacterial translocation from the intestinal lumen to the mesenteric lymph nodes and spleen.

Up until the first half of the 20th century, silver found significant employment in medical applications, particularly in the healing of open wounds, thanks to its antibacterial and antifungal properties. Wound repair is a complex and dynamic biological process regulated by several pathways that cooperate to restore tissue integrity and homeostasis. To facilitate healing, injuries need to be promptly treated. Recently, the interest in alternatives to antibiotics has been raised given the widespread phenomenon of antibiotic resistance. Among these alternatives, the use of silver appears to be a valid option, so a resurgence in its use has been recently observed. In particular, in contrast to ionic silver, colloidal silver, a suspension of metallic silver particles, shows antibacterial activity displaying less or no toxicity. However, the human health risks associated with exposure to silver nanoparticles (NP) appear to be conflicted, and some studies have suggested that it could be toxic in different cellular contexts. These potentially harmful effects of silver NP depend on various parameters including NP size, which commonly range from 1 to 100 nm. In this study, we analyzed the effect of a colloidal silver preparation composed of very small and homogeneous nanoparticles of 0.62 nm size, smaller than those previously tested. We found no adverse effect on the cell proliferation of HaCaT cells, even at high NP concentration. Time-lapse microscopy and indirect immunofluorescence experiments demonstrated that this preparation of colloidal silver strongly increased cell migration, re-modeled the cytoskeleton, and caused recruitment of E-cadherin at cell-cell junctions of human cultured keratinocytes.

The small intestine contains CD4 + CD8αα + double-positive intraepithelial lymphocytes (DP IELs), which originate from intestinal CD4 + T cells through down-regulation of the transcription factor Thpok and have regulatory functions. DP IELs are absent in germ-free mice, which suggests that their differentiation depends on microbial factors. We found that DP IEL numbers in mice varied in different vivaria, correlating with the presence of Lactobacillus reuteri This species induced DP IELs in germ-free mice and conventionally-raised mice lacking these cells. L. reuteri did not shape the DP-IEL-TCR (TCR, T cell receptor) repertoire but generated indole derivatives of tryptophan that activated the aryl-hydrocarbon receptor in CD4 + T cells, allowing Thpok down-regulation and differentiation into DP IELs. Thus, L. reuteri , together with a tryptophan-rich diet, can reprogram intraepithelial CD4 + T cells into immunoregulatory T cells., (Copyright © 2017 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.)

Objective : The link between metabolic derangement of the gut-2013liver-visceral white adipose tissue (v-WAT) axis and gut microbiota was investigated. Methods : Rats were fed a fructose-rich diet and treated with an antibiotic mix. Inflammation was measured in portal plasma, ileum, liver, and v-WAT, while insulin signalling was analysed by measuring levels of phosphorylated kinase Akt. The function and oxidative status of hepatic mitochondria and caecal microbiota composition were also evaluated. Results : Ileal inflammation, increase in plasma transaminases, plasma peroxidised lipids, portal concentrations of tumour necrosis factor alpha, lipopolysaccharide, and non-esterified fatty acids, were induced by fructose and were reversed by antibiotic. The increased hepatic ceramide content, inflammation and decreased insulin signaling in liver and v-WAT induced by fructose was reversed by antibiotic. Antibiotic also blunted the increase in hepatic mitochondrial efficiency and oxidative damage of rats fed fructose-rich diet. Three genera, Coprococcus, Ruminococcus, and Clostridium, significantly increased, while the Clostridiaceae family significantly decreased in rats fed a fructose-rich diet, and antibiotic abolished these variations Conclusions : When gut microbiota modulation by fructose is prevented by antibiotic, inflammatory flow from the gut to the liver and v-WAT are reversed.

A novel metal ion-sensitive fluorescent peptidyl-probe has been designed based on the most common five-residue repeat in mammalian histidine rich glycoproteins (HRGs). A dansyl-amide moiety at the N-terminus and a tryptophan residue at the C-terminus of the peptide were added as they can act as a FRET (fluorescence resonance energy transfer) pair. The dansyl fluorophore was chosen also because it frequently shows strong CHEF (chelation enhanced fluorescence) and solvatochromic effects. The designed peptide, dansyl-HPHGHW-NH 2 (dH3w), showed a selective fluorescence turn-on response to Zn 2+ in aqueous solutions at pH 7.0 when excited at both 295 nm and 340 nm, thus indicating that both FRET and CHEF or solvatochromic effects are active in the metal/peptide complex. Steady-state fluorescence and isothermal titration calorimetry (ITC) measurements demonstrated that two peptide molecules bind to one zinc ion with an association constant K a = 5.7 × 10 5 M -1 at 25 °C and pH 7.0. The fluorescence response to Zn 2+ was not influenced by Pb 2+ , Cd 2+ , Mn 2+ , Fe 2+ , Fe 3+ , Mg 2+ , Ca 2+ , K + and Na + ions and only slightly influenced by Co 2+ and Ni 2+ . Copper(ii), at concentrations as low as 5 μM, caused a strong quenching of both free and Zn 2+ complexed dH3w. The determination of the binding parameters for Cu 2+ has shown that one copper ion binds to one dH3w molecule with an association constant of 1.2 × 10 6 M -1 thus confirming the higher affinity of peptide for Cu 2+ than for Zn 2+ . Finally, we demonstrated that dH3w can penetrate into HeLa cells and could thus be used for the determination of intracellular Zn 2+ and Cu 2+ concentrations.

The ability to produce an extracellular matrix and form multicellular communities is an adaptive behavior shared by many bacteria. In Bacillus subtilis, the model system for spore-forming bacteria, matrix production is one of the possible differentiation pathways that a cell can follow when vegetative growth is no longer feasible. While in B. subtilis the genetic system controlling matrix production has been studied in detail, it is still unclear whether other spore formers utilize similar mechanisms. We report that SF214, a pigmented strain of Bacillus pumilus isolated from the marine environment, can produce an extracellular matrix relying on orthologs of many of the genes known to be important for matrix synthesis in B. subtilis. We also report a characterization of the carbohydrates forming the extracellular matrix of strain SF214. The isolation and characterization of mutants altered in matrix synthesis, pigmentation, and spore formation suggest that in strain SF214 the three processes are strictly interconnected and regulated by a common molecular mechanism.

A fructose-rich diet can induce metabolic syndrome, a combination of health disorders that increases the risk of diabetes and cardiovascular diseases. Diet is also known to alter the microbial composition of the gut, although it is not clear whether such alteration contributes to the development of metabolic syndrome. The aim of this work was to assess the possible link between the gut microbiota and the development of diet-induced metabolic syndrome in a rat model of obesity. Rats were fed either a standard or high-fructose diet. Groups of fructose-fed rats were treated with either antibiotics or faecal samples from control rats by oral gavage. Body composition, plasma metabolic parameters and markers of tissue oxidative stress were measured in all groups. A 16S DNA-sequencing approach was used to evaluate the bacterial composition of the gut of animals under different diets. The fructose-rich diet induced markers of metabolic syndrome, inflammation and oxidative stress, that were all significantly reduced when the animals were treated with antibiotic or faecal samples. The number of members of two bacterial genera, Coprococcus and Ruminococcus, was increased by the fructose-rich diet and reduced by both antibiotic and faecal treatments, pointing to a correlation between their abundance and the development of the metabolic syndrome. Our data indicate that in rats fed a fructose-rich diet the development of metabolic syndrome is directly correlated with variations of the gut content of specific bacterial taxa.

Bacillus pumilus SF214 is a spore forming bacterium, isolated from a marine sample, able to produce a matrix and a orange-red, water soluble pigment. Pigmentation is strictly regulated and high pigment production was observed during the late stationary growth phase in a minimal medium and at growth temperatures lower than the optimum. Only a subpopulation of stationary phase cells produced the pigment, indicating that the stationary culture contains a heterogeneous cell population and that pigment synthesis is a bimodal phenomenon. The fraction of cells producing the pigment varied in the different growth conditions and occurred only in cells not devoted to sporulation. Only some of the pigmented cells were also able to produce a matrix. Pigment and matrix production in SF214 appear then as two developmental fates both alternative to sporulation. Since the pigment had an essential role in the cell resistance to oxidative stress conditions, we propose that within the heterogeneous population different survival strategies can be followed by the different cells.