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Protection against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and risk of long COVID has been associated with the depletion or over-abundance of specific taxa within the gut microbiome. However, the microbial mechanisms mediating these effects are not yet known. We hypothesized that altered microbial production of tryptophan and its downstream derivatives might contribute to inappropriate immune responses to viral infection. In patients hospitalized with COVID-19 (n = 172), serum levels of tryptophan and indole-3-propionate (IPA) negatively correlated with serum levels of many proinflammatory mediators (including C-reactive protein and Serum amyloid A), while C-glycosyltryptophan (C-Trp), indole-3-lactic acid (ILA) and indole-3-acetic acid (IAA) levels were positively correlated with levels of acute phase proteins, proinflammatory cytokines, alarmins and chemokines. A similar pattern was observed in long COVID patients (n = 20) where tryptophan and IPA were negatively associated with a large number of serum cytokines, while C-Trp and IAA were positively associated with circulating cytokine levels. Metagenomic analysis of the fecal microbiota showed the relative abundance of genes encoding the microbial enzymes required for tryptophan production (e.g. anthranilate synthase) and microbial tryptophan metabolism was significantly lower in patients hospitalized with COVID-19 (n = 380) compared to healthy controls (n = 270). Microbial tryptophan metabolites reduced innate cell proinflammatory responses to cytosolic DNA sensor Stimulator of interferon genes (STING), toll-like receptor (TLR)-3 and TLR-4 stimulation in vitro, while IL-10 secretion was enhanced. Microbial tryptophan metabolites also modified ex vivo human lymphocyte responses by limiting the production of TH1 and TH17 associated cytokines, while enhancing secretion of IL-22. These data suggest that lower levels of tryptophan production and tryptophan metabolism by gut microbes may increase the risk of severe and chronic outcomes to SARS-CoV-2 infection due to impaired innate and adaptive responses to infection. Screening patients for lower-level microbiome capacity for tryptophan metabolism may help identify at-risk individuals.

Abstract The term long COVID (LC) effectively describes the broad long-term disease burden of severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) infections, encompassing individual suffering and significant socioeconomic impacts. However, its general use hampers precise epidemiological research, diagnostics and therapeutic strategies. Misinterpretations occur, for example, when population surveys are compared to studies using health record data, because both refer to these data as LC. This also emphasizes the need for different terminology. The National Institute for Health and Care Excellence (NICE) rapid guideline differentiates ongoing symptomatic COVID-19 from post-COVID conditions, yet real-world observations challenge these two subgroup definitions. We propose refining the term LC into three subgroups: ongoing symptomatic COVID-19, SARS-CoV-2 induced or exacerbated diseases and post-acute COVID condition. This stratification aids targeted diagnostics, treatment and epidemiological research. Subgroup-specific documentation using the International Classification of Diseases, Tenth Revision (ICD-10) codes ensures accurate tracking and understanding of long-term effects. The subgroup of post-acute COVID condition again includes various symptoms, syndromes and diseases like post-exertional malaise (PEM), dysautonomia or cognitive dysfunctions. In this regard, differentiation, especially considering PEM, is crucial for effective diagnostics and treatment.

Abstract Introduction Guidelines recommend treating asthma exacerbations (AAEs) with bronchodilators combined with inhaled and/or systemic corticosteroids. Indications for antibiotic prescriptions for AAEs are usually not incorporated although the literature shows antibiotics are frequently prescribed. Aim To investigate the antibiotic prescription rates in AAEs and explore the possible determining factors of those practices. Methods A digital survey was created to determine the antibiotic prescription rates in AAEs and the influencing factors for the prescription practices. The survey was distributed among European academy of allergy and clinical immunology (EAACI) members by mass emailing and through regional/national societies in the Netherlands, Italy, Greece, and Poland. Furthermore, we retrieved local antibiotic prescription rates. Results In total, 252 participants completed the survey. Respondents stated that there is a lack of guidelines to prescribe antibiotics in AAEs. The median antibiotic prescription rate in this study was 19% [IQR: 0%–40%] and was significantly different between 4 professions: paediatrics 0% [IQR: 0%–37%], pulmonologists 25% [IQR: 10%–50%], general practitioners 25% [IQR: 0%–50%], and allergologists 17% [IQR: 0%–33%]) (p = 0.046). Additional diagnostic tests were performed in 71.4% of patients before prescription and the most common antibiotic classes prescribed were macrolides (46.0%) and penicillin (42.9%). Important clinical factors for health care providers to prescribe antibiotics were colorised/purulent sputum, abnormal lung sounds during auscultation, fever, and presence of comorbidities. Conclusion In 19% of patients with AAEs, antibiotics were prescribed in various classes with a broad range among different subspecialities. This study stresses the urgency to compose evidence‐based guidelines to aim for more rational antibiotic prescriptions for AAE.

Abstract Background Allergic diseases, including respiratory and food allergies, as well as allergic skin conditions have surged in prevalence in recent decades. In allergic diseases, the gut microbiome is dysbiotic, with reduced diversity of beneficial bacteria and increased abundance of potential pathogens. Research findings suggest that the microbiome, which is highly influenced by environmental and dietary factors, plays a central role in the development, progression, and severity of allergic diseases. The microbiome generates metabolites, which can regulate many of the host’s cellular metabolic processes and host immune responses. Aims and Methods Our goal is to provide a narrative and comprehensive literature review of the mechanisms through which microbial metabolites regulate host immune function and immune metabolism both in homeostasis and in the context of allergic diseases. Results and Discussion We describe key microbial metabolites such as short‐chain fatty acids, amino acids, bile acids and polyamines, elucidating their mechanisms of action, cellular targets and their roles in regulating metabolism within innate and adaptive immune cells. Furthermore, we characterize the role of bacterial metabolites in the pathogenesis of allergic diseases including allergic asthma, atopic dermatitis and food allergy. Conclusion Future research efforts should focus on investigating the physiological functions of microbiota‐derived metabolites to help develop new diagnostic and therapeutic interventions for allergic diseases.

ABSTRACTAlthough many recent studies have examined associations between the gut microbiome and COVID-19 disease severity in individual patient cohorts, questions remain on the robustness across international cohorts of the biomarkers they reported. Here, we performed a meta-analysis of eight shotgun metagenomic studies of COVID-19 patients (comprising 1,023 stool samples) and 23 > 16S rRNA gene amplicon sequencing (16S) cohorts (2,415 total stool samples). We found that disease severity (as defined by the WHO clinical progression scale) was associated with taxonomic and functional microbiome differences. This alteration in gut microbiome configuration peaks at days 7–30 post diagnosis, after which the gut microbiome returns to a configuration that becomes more similar to that of healthy controls over time. Furthermore, we identified a core set of species that were consistently associated with disease severity across shotgun metagenomic and 16S cohorts, and whose abundance can accurately predict disease severity category of SARS-CoV-2 infected subjects, with Actinomyces oris abundance predicting population-level mortality rate of COVID-19. Additionally, we used relational diet-microbiome databases constructed from cohort studies to predict microbiota-targeted diet patterns that would modulate gut microbiota composition toward that of healthy controls. Finally, we demonstrated the association of disease severity with the composition of intestinal archaeal, fungal, viral, and parasitic communities. Collectively, this study has identified robust COVID-19 microbiome biomarkers, established accurate predictive models as a basis for clinical prognostic tests for disease severity, and proposed biomarker-targeted diets for managing COVID-19 infection.

ABSTRACTA high-fat (HF) diet reduces resistance to the foodborne pathogen Listeria monocytogenes. We demonstrate that short-term gavage with A. muciniphila increases resistance to oral and systemic L. monocytogenes infection in mice fed a HF diet. A. muciniphila reduced inflammation in the gut and liver of mice fed a high-fat diet prior to infection and reduced inflammatory cell infiltration in the ileum to levels similar to mice fed a low-fat (LF) diet. Akkermansia administration had minimal impacts upon the microbiota and microbial metabolites and did not affect individual taxa or impact the Bacteroidetes to Firmicutes ratio. In summary, A. muciniphila increased resistance to L. monocytogenes infection in mice fed a HF diet by moderating immune/physiological effects through specific interaction between A. muciniphila and the host gut.

Abstract Rhinoviruses and allergens, such as house dust mite are major agents responsible for asthma exacerbations. The influence of pre-existing airway inflammation on the infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is largely unknown. We analyse mechanisms of response to viral infection in experimental in vivo rhinovirus infection in healthy controls and patients with asthma, and in in vitro experiments with house dust mite, rhinovirus and SARS-CoV-2 in human primary airway epithelium. Here, we show that rhinovirus infection in patients with asthma leads to an excessive RIG-I inflammasome activation, which diminishes its accessibility for type I/III interferon responses, leading to their early functional impairment, delayed resolution, prolonged viral clearance and unresolved inflammation in vitro and in vivo. Pre-exposure to house dust mite augments this phenomenon by inflammasome priming and auxiliary inhibition of early type I/III interferon responses. Prior infection with rhinovirus followed by SARS-CoV-2 infection augments RIG-I inflammasome activation and epithelial inflammation. Timely inhibition of the epithelial RIG-I inflammasome may lead to more efficient viral clearance and lower the burden of rhinovirus and SARS-CoV-2 infections.

Protection against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and associated clinical sequelae requires well-coordinated metabolic and immune responses that limit viral spread and promote recovery of damaged systems. However, the role of the gut microbiota in regulating these responses has not been thoroughly investigated. In order to identify mechanisms underpinning microbiota interactions with host immune and metabolic systems that influence coronavirus disease 2019 (COVID-19) outcomes, we performed a multi-omics analysis on hospitalized COVID-19 patients and compared those with the most severe outcome (i.e. death, n = 41) to those with severe non-fatal disease (n = 89), or mild/moderate disease (n = 42), that recovered. A distinct subset of 8 cytokines (e.g. TSLP) and 140 metabolites (e.g. quinolinate) in sera identified those with a fatal outcome to infection. In addition, elevated levels of multiple pathobionts and lower levels of protective or anti-inflammatory microbes were observed in the fecal microbiome of those with the poorest clinical outcomes. Weighted gene correlation network analysis (WGCNA) identified modules that associated severity-associated cytokines with tryptophan metabolism, coagulation-linked fibrinopeptides, and bile acids with multiple pathobionts, such as Enterococcus. In contrast, less severe clinical outcomes are associated with clusters of anti-inflammatory microbes such as Bifidobacterium or Ruminococcus, short chain fatty acids (SCFAs) and IL-17A. Our study uncovered distinct mechanistic modules that link host and microbiome processes with fatal outcomes to SARS-CoV-2 infection. These features may be useful to identify at risk individuals, but also highlight a role for the microbiome in modifying hyperinflammatory responses to SARS-CoV-2 and other infectious agents.

COVID-19, a systemic multi-organ disease resulting from infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is known to result in a wide array of disease outcomes, ranging from asymptomatic to fatal. Despite persistent progress, there is a continued need for more accurate determinants of disease outcomes, including post-acute symptoms after COVID-19. In this study, we characterised the serum metabolomic changes due to hospitalisation and COVID-19 disease progression by mapping the serum metabolomic trajectories of 71 newly hospitalised moderate and severe patients in their first week after hospitalisation. These 71 patients were spread out over three hospitals in Switzerland, enabling us to meta-analyse the metabolomic trajectories and filter consistently changing metabolites. Additionally, we investigated differential metabolite–metabolite trajectories between fatal, severe, and moderate disease outcomes to find prognostic markers of disease severity. We found drastic changes in serum metabolite concentrations for 448 out of the 901 metabolites. These results included markers of hospitalisation, such as environmental exposures, dietary changes, and altered drug administration, but also possible markers of physiological functioning, including carboxyethyl-GABA and fibrinopeptides, which might be prognostic for worsening lung injury. Possible markers of disease progression included altered urea cycle metabolites and metabolites of the tricarboxylic acid (TCA) cycle, indicating a SARS-CoV-2-induced reprogramming of the host metabolism. Glycerophosphorylcholine was identified as a potential marker of disease severity. Taken together, this study describes the metabolome-wide changes due to hospitalisation and COVID-19 disease progression. Moreover, we propose a wide range of novel potential biomarkers for monitoring COVID-19 disease course, both dependent and independent of the severity.

Abstract The growing world population and increased pressure on agricultural resources are driving a shortage of dietary protein sources. As a result, industry is developing more sustainable novel food protein sources such as insects, algae and duckweed and using new processing techniques. Consumer exposure to these novel or processed proteins, could cause new food allergies, exacerbating a public health issue which is already directly affecting an estimated 20 million Europeans. Introduction of novel foods should not add to the burden of food allergy and this calls for a reliable, harmonised, evidence-based and validated allergenicity risk assessment strategy. The COST (Cooperation in Science and Technology) Action ImpARAS (Improved Allergenicity Risk Assessment Strategy), a four-year networking project, identified gaps in current allergy risk assessment, and proposed new ideas and plans for improving it. Here, we report on the lessons learned from the ImpARAS network and suggestions for future research. The safe introduction of novel and more sustainable food protein sources, while protecting humans from food allergy, calls for a multidisciplinary approach based on an improved understanding of what determines the relative allergenic potency of proteins, novel testing and assessment methodologies, harmonized decision-making criteria, and a clear ranking approach to express the allergenicity of novel product relative to that of existing known allergenic proteins: (from ‘non’/to weakly and to strongly allergenic proteins).

Biomechanical stability plays an important role in fracture healing, with unstable fixation being associated with healing disturbances. A lack of stability is also considered a risk factor for fracture-related infection (FRI), although confirmatory studies and an understanding of the underlying mechanisms are lacking. In the present study, we investigate whether biomechanical (in)stability can lead to altered immune responses in mice under sterile or experimentally inoculated conditions. In non-inoculated C57BL/6 mice, instability resulted in an early increase of inflammatory markers such as granulocyte-colony stimulating factor (G-CSF), keratinocyte chemoattractant (KC) and interleukin (IL)-6 within the bone. When inoculated with Staphylococcus epidermidis, instability resulted in a further significant increase in G-CSF, IL-6 and KC in bone tissue. Staphylococcus aureus infection led to rapid osteolysis and instability in all animals and was not further studied. Gene expression measurements also showed significant upregulation in CCL2 and G-CSF in these mice. IL-17A was found to be upregulated in all S. epidermidis infected mice, with higher systemic IL-17A cell responses in mice that cleared the infection, which was found to be produced by CD4+ and γδ+ T cells in the bone marrow. IL-17A knock-out (KO) mice displayed a trend of delayed clearance of infection (P=0.22, Fisher’s exact test) and an increase in interferon (IFN)-γ production. Biomechanical instability leads to a more pronounced local inflammatory response, which is exaggerated by bacterial infection. This study provides insights into long-held beliefs that biomechanics are crucial not only for fracture healing, but also for control of infection.

Here, the authors characterize immunological and microbiome alterations in a cohort of obese asthmatics, finding that disease severity negatively correlates with fecal abundance of Akkermansia muciniphila, and show in a mouse model that administration of A. muciniphila reduces airway hyper-reactivity and airway inflammation.

Short-chain fatty acids (SCFAs) produced by the gut microbiota have previously been demonstrated to play a role in numerous chronic inflammatory diseases and to be key mediators in the gut-bone signaling axis. However, the role of SCFAs in bone fracture healing and its impact on systemic inflammation during the regeneration process has not been extensively investigated yet. The aim of this study was to first determine the effects of the SCFA butyrate on key cells involved in fracture healing in vitro, namely, osteoclasts and mesenchymal stromal cells (MSCs), and second, to assess if butyrate supplementation or antibiotic therapy impacts bone healing, systemic immune status, and inflammation levels in a murine osteotomy model. Butyrate significantly reduced osteoclast formation and resorption activity in a dose-dependent manner and displayed a trend for increased calcium deposits in MSC cultures. Numerous genes associated with osteoclast differentiation were differentially expressed in osteoclast precursor cells upon butyrate exposure. In vivo, antibiotic-treated mice showed reduced SCFA levels in the cecum, as well as a distinct gut microbiome composition. Furthermore, circulating proinflammatory TNFα, IL-17a, and IL-17f levels, and bone preserving osteoprotegerin (OPG), were increased in antibiotic-treated mice compared to controls. Antibiotic-treated mice also displayed a trend towards delayed bone healing as revealed by reduced mineral apposition at the defect site and higher circulating levels of the bone turnover marker PINP. Butyrate supplementation resulted in a lower abundance of monocyte/macrophages in the bone marrow, as well as reduced circulating proinflammatory IL-6 levels compared to antibiotic- and control-treated mice. In conclusion, this study supports our hypothesis that SCFAs, in particular butyrate, are important contributors to successful bone healing by modulating key cells involved in fracture healing as well as systemic inflammation and immune responses.

Abstract The mucosal immune system is in constant communication with the vast diversity of microbes present on body surfaces. The discovery of novel molecular mechanisms, which mediate host-microbe communication, have highlighted the important roles played by microbes in influencing mucosal immune responses. Dendritic cells, epithelial cells, ILCs, T regulatory cells, effector lymphocytes, NKT cells and B cells can all be influenced by the microbiome. Many of the mechanisms being described are bacterial strain- or metabolite-specific. Microbial dysbiosis in the gut and the lung is increasingly being associated with the incidence and severity of asthma. More accurate endotyping of patients with asthma may be assisted by further analysis of the composition and metabolic activity of an individual’s microbiome. In addition, the efficacy of specific therapeutics may be influenced by the microbiome and novel bacterial-based therapeutics should be considered in future clinical studies.

Rural lifestyle has been shown to be highly protective against the development of allergies. Contact to farm-animals or pets and early-life consumption of milk products turned out to be important. These exposures provide contact to N-glycolylneuraminic acid (Neu5Gc), a sialic acid naturally expressed in mammalians but not in humans or microbes although both are able to incorporate exogenously provided Neu5Gc and induce thereby an anti-Neu5Gc antibody response. Farmers' children had elevated levels of anti-Neu5Gc antibodies associated with increased contact to Neu5Gc. Farm-related exposures that were associated with protection against allergies such as exposure to farm-animals or pets and consumption of milk were also associated with an antibody response to Neu5Gc in children. Exposure to cats was associated with increased anit-Neu5Gc IgG levels at different timepoints assessed between 1 year of age and school-age. Moreover, consumption of non-pasteurized milk in the first year of life was associated with increased anti-Neu5Gc IgG levels. Neu5Gc-providing exposures that were associated with protection against allergies were reflected in an elevated anti-Neu5Gc IgG level in children. Exposure to Neu5Gc was associated with anti-inflammation and protection of asthma development in children and mice without contribution of anti-Neu5Gc antibodies.

The focus of this research was to explore school leadership in post-primary schools using an adapted Schmidt Toxic Leadership Scale ©, which the authors recalibrated to examine both constructive and destructive leadership, the impact on individuals professional and personal lives, and on staff morale. Using a mixed methods approach, data were collected from 111 teaching professionals via online survey. Findings indicated a notable emergence of toxic leadership experiences which is reported in this paper. In addition, participants reported various and concerning negative consequences including: decreased job satisfaction, professional agency, and staff morale; reduced performance; increased attrition; increased negative behaviours including incivility; stifled career development; reduced self-confidence; depression; stress and anxiety; fear; tearfulness; humiliation; anger; mistrust; exhaustion; burnout; health issues; migraines; weight gain; substance abuse, suicidal thoughts, as well as, negative consequences on personal/home life. The results indicate that the quality of leadership was perceived to influence the health of respondents and had a bearing on their occupational wellbeing. Further research is needed to understand the nature of toxic leadership in education and its effects on organisational members.

Staphylococcus epidermidis is a permanent member of the normal human microbiota, commonly found on skin and mucous membranes. By adhering to tissue surface moieties of the host via specific adhesins, S. epidermidis is capable of establishing a lifelong commensal relationship with humans that begins early in life. In its role as a commensal organism, S. epidermidis is thought to provide benefits to human host, including out-competing more virulent pathogens. However, largely due to its capacity to form biofilm on implanted foreign bodies, S. epidermidis has emerged as an important opportunistic pathogen in patients receiving medical devices. S. epidermidis causes approximately 20% of all orthopedic device-related infections (ODRIs), increasing up to 50% in late-developing infections. Despite this prevalence, it remains underrepresented in the scientific literature, in particular lagging behind the study of the S. aureus. This review aims to provide an overview of the interactions of S. epidermidis with the human host, both as a commensal and as a pathogen. The mechanisms retained by S. epidermidis that enable colonization of human skin as well as invasive infection, will be described, with a particular focus upon biofilm formation. The host immune responses to these infections are also described, including how S. epidermidis seems to trigger low levels of pro-inflammatory cytokines and high levels of interleukin-10, which may contribute to the sub-acute and persistent nature often associated with these infections. The adaptive immune response to S. epidermidis remains poorly described, and represents an area which may provide significant new discoveries in the coming years.

The Bifibobacterium longum subsp. longum 35624™ strain (formerly named Bifidobacterium longum subsp. infantis) is a well described probiotic with clinical efficacy in Irritable Bowel Syndrome clinical trials and induces immunoregulatory effects in mice and in humans. This paper presents (a) the genome sequence of the organism allowing the assignment to its correct subspeciation longum; (b) a comparative genome assessment with other B. longum strains and (c) the molecular structure of the 35624 exopolysaccharide (EPS624). Comparative genome analysis of the 35624 strain with other B. longum strains determined that the sub-speciation of the strain is longum and revealed the presence of a 35624-specific gene cluster, predicted to encode the biosynthetic machinery for EPS624. Following isolation and acid treatment of the EPS, its chemical structure was determined using gas and liquid chromatography for sugar constituent and linkage analysis, electrospray and matrix assisted laser desorption ionization mass spectrometry for sequencing and NMR. The EPS consists of a branched hexasaccharide repeating unit containing two galactose and two glucose moieties, galacturonic acid and the unusual sugar 6-deoxy-L-talose. These data demonstrate that the B. longum 35624 strain has specific genetic features, one of which leads to the generation of a characteristic exopolysaccharide.

The microbiota is required for optimal host development and ongoing immune homeostasis. Lactobacilli are common inhabitants of the mammalian large intestine and immunoregulatory effects have been described for certain, but not all, strains. The mechanisms underpinning these protective effects are beginning to be elucidated. One such protective organism is Lactobacillus rhamnosus JB-1 (Lb. rhamnosus JB-1). Lb. murinus has no such anti-inflammatory protective effects and was used as a comparator organism. Human monocyte-derived dendritic cells (MDDCs) were co-incubated with bacteria and analysed over time for bacterial adhesion and intracellular processing, costimulatory molecule expression, cytokine secretion and induction of lymphocyte polarization. Neutralising antibodies were utilized to identify the responsible MDDC receptors. Lb. rhamnosus JB-1 adhered to MDDCs, but internalization and intracellular processing was significantly delayed, compared to Lb. murinus which was rapidly internalized and processed. Lb. murinus induced CD80 and CD86 expression, accompanied by high levels of cytokine secretion, while Lb. rhamnosus JB-1 was a poor inducer of costimulatory molecule expression and cytokine secretion. Lb. rhamnosus JB-1 primed MDDCs induced Foxp3 expression in autologous lymphocytes, while Lb. murinus primed MDDCs induced Foxp3, T-bet and Ror-γt expression. DC-SIGN was required for Lb. rhamnosus JB-1 adhesion and influenced IL-12 secretion, while TLR-2 influenced IL-10 and IL-12 secretion. Here we demonstrate that the delayed kinetics of bacterial processing by MDDCs correlates with MDDC activation and stimulation of lymphocytes. Thus, inhibition or delay of intracellular processing may be a novel strategy by which certain commensals may avoid the induction of proinflammatory responses.

Unfortunately this article [1] was published with an error in the Funding section. The BM4SIT project is not acknowledged. This section should be corrected to the below: Funding EAACI and the BM4SIT project (Grant Number 601763) in the European Union’s Seventh Framework Programme FP7.

In regions with a high infectious disease burden, concerns have been raised about the safety of iron supplementation because higher iron concentrations in the gut lumen may increase risk of enteropathogen infection. The aim of this study was to investigate interactions of the enteropathogen Salmonella enterica ssp. enterica Typhimurium with intestinal cells under different iron concentrations encountered in the gut lumen during iron deficiency and supplementation using an in vitro colonic fermentation system inoculated with immobilized child gut microbiota combined with Caco-2/HT29-MTX co-culture monolayers. Colonic fermentation effluents obtained during normal, low (chelation by 2,2'-dipyridyl) and high iron (26.5 mg iron/L) fermentation conditions containing Salmonella or pure Salmonella cultures with similar iron conditions were applied to cellular monolayers. Salmonella adhesion and invasion capacity, cellular integrity and immune response were assessed. Under high iron conditions in pure culture, Salmonella adhesion was 8-fold increased compared to normal iron conditions while invasion was not affected leading to decreased invasion efficiency (-86%). Moreover, cellular cytokines IL-1β, IL-6, IL-8 and TNF-α secretion as well as NF-κB activation in THP-1 cells were attenuated under high iron conditions. Low iron conditions in pure culture increased Salmonella invasion correlating with an increase in IL-8 release. In fermentation effluents, Salmonella adhesion was 12-fold and invasion was 428-fold reduced compared to pure culture. Salmonella in high iron fermentation effluents had decreased invasion efficiency (-77.1%) and cellular TNF-α release compared to normal iron effluent. The presence of commensal microbiota and bacterial metabolites in fermentation effluents reduced adhesion and invasion of Salmonella compared to pure culture highlighting the importance of the gut microbiota as a barrier during pathogen invasion. High iron concentrations as encountered in the gut lumen during iron supplementation attenuated Salmonella invasion efficiency and cellular immune response suggesting that high iron concentrations alone may not lead to an increased Salmonella invasion.

Appropriate dendritic cell processing of the microbiota promotes intestinal homeostasis and protects against aberrant inflammatory responses. Mucosal CD103(+) dendritic cells are able to produce retinoic acid from retinal, however their role in vivo and how they are influenced by specific microbial species has been poorly described. Bifidobacterium infantis 35624 (B. infantis) feeding to mice resulted in increased numbers of CD103(+)retinaldehyde dehydrogenase (RALDH)(+) dendritic cells within the lamina propria (LP). Foxp3(+) lymphocytes were also increased in the LP, while TH1 and TH17 subsets were decreased. 3,7-dimethyl-2,6-octadienal (citral) treatment of mice blocked the increase in CD103(+)RALDH(+) dendritic cells and the decrease in TH1 and TH17 lymphocytes, but not the increase in Foxp3(+) lymphocytes. B. infantis reduced the severity of DSS-induced colitis, associated with decreased TH1 and TH17 cells within the LP. Citral treatment confirmed that these effects were RALDH mediated. RALDH(+) dendritic cells decreased within the LP of control inflamed animals, while RALDH(+) dendritic cells numbers were maintained in the LP of B. infantis-fed mice. Thus, CD103(+)RALDH(+) LP dendritic cells are important cellular targets for microbiota-associated effects on mucosal immunoregulation.

Host defence against infection requires a range of innate and adaptive immune responses that may lead to tissue damage. Such immune-mediated pathologies can be controlled with appropriate T regulatory (Treg) activity. The aim of the present study was to determine the influence of gut microbiota composition on Treg cellular activity and NF-kappaB activation associated with infection. Mice consumed the commensal microbe Bifidobacterium infantis 35624 followed by infection with Salmonella typhimurium or injection with LPS. In vivo NF-kappaB activation was quantified using biophotonic imaging. CD4+CD25+Foxp3+ T cell phenotypes and cytokine levels were assessed using flow cytometry while CD4+ T cells were isolated using magnetic beads for adoptive transfer to naïve animals. In vivo imaging revealed profound inhibition of infection and LPS induced NF-kappaB activity that preceded a reduction in S. typhimurium numbers and murine sickness behaviour scores in B. infantis-fed mice. In addition, pro-inflammatory cytokine secretion, T cell proliferation, and dendritic cell co-stimulatory molecule expression were significantly reduced. In contrast, CD4+CD25+Foxp3+ T cell numbers were significantly increased in the mucosa and spleen of mice fed B. infantis. Adoptive transfer of CD4+CD25+ T cells transferred the NF-kappaB inhibitory activity. Consumption of a single commensal micro-organism drives the generation and function of Treg cells which control excessive NF-kappaB activation in vivo. These cellular interactions provide the basis for a more complete understanding of the commensal-host-pathogen trilogue that contribute to host homeostatic mechanisms underpinning protection against aberrant activation of the innate immune system in response to a translocating pathogen or systemic LPS.