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BACKGROUND:Intestinal permeability can be assessed by monitoring renal excretion of orally administered radioactively 51Cr-labeled ethylenediaminetetraacetic acid (51Cr-EDTA). Although considered safe, patient participation in using radio-labeled tracers is low. Here, we used orally administered 52Cr-EDTA as non-radioactive alternative to assess intestinal permeability in CD and analyzed the association with disease activity, disease location and gut microbial dysbiosis. MATERIALS AND METHODS:60 CD patients with low (n = 25) and increased (n = 35) fecal calprotectin levels (cut-off: 100 μg/g feces) ingested 20 mL 52Cr-EDTA (20 mmol/L) solution whereafter 24-h urine was collected. Urinary 52Cr-EDTA concentrations were quantified using Inductively Coupled Plasma Mass Spectrometry (ICP-MS). Fecal Enterobacteriaceae and Faecalibacterium prausnitzii were quantified using FISH. Correlations between urinary 52Cr-EDTA excretion and other parameters were established using nonparametric Spearman's correlation coefficients (ρ). RESULTS:CD patients with increased fecal calprotectin levels (> 100 μg/g) demonstrated an elevated urinary 52Cr-EDTA/creatinine ratio (772 vs. 636 μmol/mol, P = 0.132). Patients with primarily colonic disease showed the highest 52Cr-EDTA excretion. Importantly, a positive correlation was observed for the urinary 52Cr-EDTA/creatinine ratio and fecal calprotectin levels (ρ = 0.325, P < 0.05). Finally, urinary 52Cr-EDTA/creatinine ratio negatively correlated with the relative abundance of Faecalibacterium prausnitzii (ρ = -0.221, P = 0.092), while positively correlating with Enterobacteriaceae (ρ = 0.202, P = 0.126). CONCLUSIONS:Orally administered and renal excreted 52Cr-EDTA may be used to assess intestinal permeability in CD and correlates with fecal calprotectin levels and bacterial species relevant to CD. This test may improve non-invasive detection of disease exacerbations and help monitor disease activity.

For some microbial species, such as Bacillus anthracis, the etiologic agent of the disease anthrax, correct detection and identification by molecular methods can be problematic. The detection of virulent B. anthracis is challenging due to multiple virulence markers that need to be present in order for B. anthracis to be virulent and its close relationship to Bacillus cereus and other members of the B. cereus group. This is especially the case in environments where build-up of Bacillus spores can occur and several representatives of the B. cereus group may be present, which increases the chance for false-positives. In this study we show the presence of B. anthracis-like bacteria and other members of the B. cereus group in a microbial community within the human environment of the International Space Station and their preliminary identification by using conventional culturing as well as molecular techniques including 16S rDNA sequencing, PCR and real-time PCR. Our study shows that when monitoring the microbial hygiene in a given human environment, health risk assessment is troublesome in the case of virulent B. anthracis, especially if this should be done with rapid, easy to apply and on-site molecular methods.

A decrease in the abundance and biodiversity of intestinal bacteria within the Firmicutes phylum has been associated with inflammatory bowel disease (IBD). In particular, the anti-inflammatory bacterium Faecalibacterium prausnitzii, member of the Firmicutes phylum and one of the most abundant species in healthy human colon, is underrepresented in the microbiota of IBD patients. The aim of this study was to investigate the immunomodulatory properties of F. prausnitzii strain A2-165, the biofilm forming strain HTF-F and the extracellular polymeric matrix (EPM) isolated from strain HTF-F. For this purpose, the immunomodulatory properties of the F. prausnitzii strains and the EPM were studied in vitro using human monocyte-derived dendritic cells. Then, the capacity of the F. prausnitzii strains and the EPM of HTF-F to suppress inflammation was assessed in vivo in the mouse dextran sodium sulphate (DSS) colitis model. The F. prausnitzii strains and the EPM had anti-inflammatory effects on the clinical parameters measured in the DSS model but with different efficacy. The immunomodulatory effects of the EPM were mediated through the TLR2-dependent modulation of IL-12 and IL-10 cytokine production in antigen presenting cells, suggesting that it contributes to the anti-inflammatory potency of F. prausnitzii HTF-F. The results show that F. prausnitzii HTF-F and its EPM may have a therapeutic use in IBD.

OBJECTIVE: Pregnant individuals of many species, including humans, are more sensitive to various bacteria or their products as compared with non-pregnant individuals. Pregnant individuals also respond differently to different bacteria or their products. Therefore, in the present study, we evaluated whether the increased sensitivity of pregnant women to bacterial products and their heterogeneous response to different bacteria was associated with differences in whole blood cytokine production upon stimulation with bacteria or their products. METHODS: Blood samples were taken from healthy pregnant and age-matched non-pregnant women and ex vivo stimulated with bacteria or LPS from Porphyromonas Gingivalis (Pg) or E-coli for 24 hrs. TNFα, IL-1ß, IL-6, IL-12 and IL-10 were measured using a multiplex Luminex system. RESULTS: We observed a generally lower cytokine production after stimulation with Pg bacteria or it's LPS as compared with E-coli bacteria. However, there was also an effect of pregnancy upon cytokine production: in pregnant women the production of IL-6 upon Pg stimulation was decreased as compared with non-pregnant women. After stimulation with E-coli, the production of IL-12 and TNFα was decreased in pregnant women as compared with non-pregnant women. CONCLUSION: Our results showed that cytokine production upon bacterial stimulation of whole blood differed between pregnant and non-pregnant women, showing that the increased sensitivity of pregnant women may be due to differences in cytokine production. Moreover, pregnancy also affected whole blood cytokine production upon Pg or E-coli stimulation differently. Thus, the different responses of pregnant women to different bacteria or their products may result from variations in cytokine production.

The beneficial human gut microbe Faecalibacterium prausnitzii is a 'probiotic of the future' since it produces high amounts of butyrate and anti-inflammatory compounds. However, this bacterium is highly oxygen-senstive, making it notoriously difficult to cultivate and preserve. This has so far precluded its clinical application in the treatment of patients with inflammatory bowel diseases. The present studies were therefore aimed at developing a strategy to keep F. prausnitzii alive at ambient air. Our previous research showed that F. prausnitzii can survive in moderately oxygenized environments like the gut mucosa by transfer of electrons to oxygen. For this purpose, the bacterium exploits extracellular antioxidants, such as riboflavin and cysteine, that are abundantly present in the gut. We therefore tested to what extent these antioxidants can sustain the viability of F. prausnitzii at ambient air. The present results show that cysteine can facilitate the survival of F. prausnitzii upon exposure to air, and that this effect is significantly enhanced the by addition of riboflavin and the cryoprotectant inulin. The highly oxygen-sensitive gut bacterium F. prausnitzii can be kept alive at ambient air for 24 h when formulated with the antioxidants cysteine and riboflavin plus the cryoprotectant inulin. Improved formulations were obtained by addition of the bulking agents corn starch and wheat bran. Our present findings pave the way towards the biomedical exploitation of F. prausnitzii in redox-based therapeutics for treatment of dysbiosis-related inflammatory disorders of the human gut.

The opportunistic pathogen Staphylococcus aureus is known to interfere with wound healing and represents a significant risk factor for wound infections and invasive disease. It is generally assumed that one individual is predominantly colonized by one S. aureus type. Nevertheless, patients with the genetic blistering disease epidermolysis bullosa (EB) often carry multiple S. aureus types. We therefore investigated whether different S. aureus types are present in individual wounds of EB patients and, if so, how they are spatially distributed. The staphylococcal topography in chronic wounds was mapped by replica-plating of used bandages and subsequent typing of S. aureus isolates. Individual chronic wounds of five patients contained up to six different S. aureus types. Unexpectedly, distinct S. aureus types formed micro-colonies that were located in close proximity and sometimes even overlapped. While some adjacent S. aureus isolates were closely related, others belonged to distinct molecular complexes. We conclude that the general assumption that one individual is predominantly colonized by one type of S. aureus does not apply to chronic wounds of EB patients. We consider this observation important, not only for EB patients, but also for other patients with chronic wounds in view of the potential risk for severe staphylococcal infections.

Mucositis, also referred to as mucosal barrier injury, is one of the most debilitating side effects of radiotherapy and chemotherapy treatment. Clinically, mucositis is associated with pain, bacteremia, and malnutrition. Furthermore, mucositis is a frequent reason to postpone chemotherapy treatment, ultimately leading towards a higher mortality in cancer patients. According to the model introduced by Sonis, both inflammation and apoptosis of the mucosal barrier result in its discontinuity, thereby promoting bacterial translocation. According to this five-phase model, the intestinal microbiota plays no role in the pathophysiology of mucositis. However, research has implicated a prominent role for the commensal intestinal microbiota in the development of several inflammatory diseases like inflammatory bowel disease, pouchitis, and radiotherapy-induced diarrhea. Furthermore, chemotherapeutics have a detrimental effect on the intestinal microbial composition (strongly decreasing the numbers of anaerobic bacteria), coinciding in time with the development of chemotherapy-induced mucositis. We hypothesize that the commensal intestinal microbiota might play a pivotal role in chemotherapy-induced mucositis. In this review, we propose and discuss five pathways in the development of mucositis that are potentially influenced by the commensal intestinal microbiota: 1) the inflammatory process and oxidative stress, 2) intestinal permeability, 3) the composition of the mucus layer, 4) the resistance to harmful stimuli and epithelial repair mechanisms, and 5) the activation and release of immune effector molecules. Via these pathways, the commensal intestinal microbiota might influence all phases in the Sonis model of the pathogenesis of mucositis. Further research is needed to show the clinical relevance of restoring dysbiosis, thereby possibly decreasing the degree of intestinal mucositis.

Periodontitis and caries are infectious diseases of the oral cavity in which oral biofilms play a causative role. Moreover, oral biofilms are widely studied as model systems for bacterial adhesion, biofilm development, and biofilm resistance to antibiotics, due to their widespread presence and accessibility. Despite descriptions of initial plaque formation on the tooth surface, studies on mature plaque and plaque structure below the gum are limited to landmark studies from the 1970s, without appreciating the breadth of microbial diversity in the plaque. We used fluorescent in situ hybridization to localize in vivo the most abundant species from different phyla and species associated with periodontitis on seven embedded teeth obtained from four different subjects. The data showed convincingly the dominance of Actinomyces sp., Tannerella forsythia, Fusobacterium nucleatum, Spirochaetes, and Synergistetes in subgingival plaque. The latter proved to be new with a possibly important role in host-pathogen interaction due to its localization in close proximity to immune cells. The present study identified for the first time in vivo that Lactobacillus sp. are the central cells of bacterial aggregates in subgingival plaque, and that Streptococcus sp. and the yeast Candida albicans form corncob structures in supragingival plaque. Finally, periodontal pathogens colonize already formed biofilms and form microcolonies therein. These in vivo observations on oral biofilms provide a clear vision on biofilm architecture and the spatial distribution of predominant species.