Commensal Research Articles

Intestinal epithelial vitamin D receptor defense against inflammatory bowel disease via regulating microfold cells

Vitamin D receptor (VDR) is involved in the pathogenesis of inflammatory bowel disease (IBD). However, the mechanism of VDR in IBD is still unclear. Microfold cells (M cells) mediated antigen-sampling pathway is central in developing immune responses to pathogenic and commensal bacteria and related to IBD. We found that Intestinal epithelial cell-specific knockdown of VDR(VDRIEC-KO) increases the susceptibility of mice to experimental colitis induced by sodium dextran sulfate(DSS) by producing more M cells. Knockdown VDR in intestinal epithelial cells increased RANKL-induced microfold cells and promoted the ability of microfold cells to uptake S. Typhimurium (S. T.). Mechanistically, we demonstrated that knockdown VDR promoted the differentiation and maturation of M cells via the Spi-B-dependent pathway. We conclude that M cells may be a potential target of VDR for treating intestinal mucosal barrier dysfunction in IBD.

An Enteric Bacterial Infection Triggers Neuroinflammation and Neurobehavioral Impairment in 3xTg-AD Transgenic Mice.

Klebsiella pneumoniae is infamous for hospital-acquired infections and sepsis, which have also been linked to Alzheimer disease (AD)-related neuroinflammatory and neurodegenerative impairment. However, its causative and mechanistic role in AD pathology remains unstudied. A preclinical model of K. pneumoniae enteric infection and colonization is developed in an AD model (3xTg-AD mice) to investigate whether and how K. pneumoniae pathogenesis exacerbates neuropathogenesis via the gut-blood-brain axis. K. pneumoniae, particularly under antibiotic-induced dysbiosis, was able to translocate from the gut to the bloodstream by penetrating the gut epithelial barrier. Subsequently, K. pneumoniae infiltrated the brain by breaching the blood-brain barrier. Significant neuroinflammatory phenotype was observed in mice with K. pneumoniae brain infection. K. pneumoniae-infected mice also exhibited impaired neurobehavioral function and elevated total tau levels in the brain. Metagenomic analyses revealed an inverse correlation of K. pneumoniae with gut biome diversity and commensal bacteria, highlighting how antibiotic-induced dysbiosis triggers an enteroseptic "pathobiome" signature implicated in gut-brain perturbations. The findings demonstrate how infectious agents following hospital-acquired infections and consequent antibiotic regimen may induce gut dysbiosis and pathobiome and increase the risk of sepsis, thereby increasing the predisposition to neuroinflammatory and neurobehavioral impairments via breaching the gut-blood-brain barrier.

Opportunistic Features of Non-Clostridium botulinum Strains Containing bont Gene Cluster.

The cluster of genes determining the production of botulinum toxins is an attribute of not only the Clostridium botulinum species. This cluster is also found in other members of the Clostridium genus, such as C. baratii, C. butyricum, and C. sporogenes. The occurrence of a botulinum-like cluster has also been recorded in strains of other genera, i.e., Enterococcus faecium, as well as in a Gram-negative species isolated from freshwater sediments; however, the biological activity of bont-related genes has not been noted. It can be said that the mentioned species have a dual nature. Another species with a dual nature is C. butyricum. This bacterium is a common human and animal gut commensal bacterium and is also frequently found in the environment. Although non-toxigenic strains are currently used as probiotics in Asia, other strains have been implicated in pathological conditions, such as botulism in infants or necrotizing enterocolitis in preterm neonates. Additionally, C. baratii strains are rare opportunistic pathogens associated with botulism intoxication. They have been isolated from food and soil and can be carried asymptomatically or cause botulism outbreaks in animals and humans. In addition to the mentioned clostridia, the other microorganisms considered as non-toxigenic have also been suspected of carrying botulinum cluster Gram-negative bacteria, such as Chryseobacterium piperi isolated from freshwater sediments; however, the biological activity of bont-related genes has not been noted. Additionally, Enterococcus faecium strains have been discovered carrying BoNT-related clusters (BoNT/En). Literature data regarding the heterogeneity of BoNT-producing strains indicate the requirement to reclassify C. botulinum species and other microorganisms able to produce BoNTs or possess botulinum-like gene clusters. This article aims to show the dual nature of Clostridium strains not belonging to the C. botulinum species that are sporadically able to carry bont clusters, which are usually considered saprophytic and even probiotic, and bont-like clusters in microorganisms from other genera. The aim was also to consider the genetic mechanisms of botulinum cluster expression in strains that are considered opportunistic and the microbiological safety aspects associated with their occurrence in the environment.

ActA-mediated PykF acetylation negatively regulates oxidative stress adaptability of Streptococcus mutans.

Dental caries poses a significant challenge to global oral health, driven by microbial dysbiosis within dental biofilms. The pathogenicity of Streptococcus mutans, a major cariogenic bacterium, is closely linked to its ability to adapt to changing environments and cellular stresses. Our investigation into the protein acetylation mechanisms, particularly through the acetyltransferase ActA, reveals a critical pathway by which S. mutans modulates its adaptability to oxidative stress, the dominant stressor within dental biofilms. By elucidating how ActA affects the oxidative stress adaptability and competitiveness of S. mutans through the regulatory axis of ActA-PykF-pyruvate, our findings provide insights into the dynamic interplay between cariogenic and commensal bacteria within dental biofilms. This work emphasizes the significance of protein acetylation in bacterial stress response and competitiveness, opening avenues for the development of novel strategies to maintain oral microbial balance within dental biofilms.

Regulation of Bacterial Growth and Behavior by Host Plant.

Plants are associated with diverse bacteria in nature. Some bacteria are pathogens that decrease plant fitness, and others are beneficial bacteria that promote plant growth and stress resistance. Emerging evidence also suggests that plant-associated commensal bacteria collectively contribute to plant health and are essential for plant survival in nature. Bacteria with different characteristics simultaneously colonize plant tissues. Thus, plants need to accommodate bacteria that provide service to the host plants, but they need to defend against pathogens at the same time. How do plants achieve this? In this review, we summarize how plants use physical barriers, control common goods such as water and nutrients, and produce antibacterial molecules to regulate bacterial growth and behavior. Furthermore, we highlight that plants use specialized metabolites that support or inhibit specific bacteria, thereby selectively recruiting plant-associated bacterial communities and regulating their function. We also raise important questions that need to be addressed to improve our understanding of plant-bacteria interactions.

The prebiotic effects of fructooligosaccharides enhance thegrowth characteristics of Staphylococcus epidermidis and enhance the inhibition of Staphylococcus aureus biofilm formation.

Oligosaccharides have been shown to enhance the production of short chain fatty acids (SCFAs) by gut probiotics and regulate gut microbiota, to improve intestinal health. Recent research indicates that oligosaccharides may also positively impact skin microbiota by selectively promoting the growth of skin commensal bacteria and inhibiting pathogenic bacteria. However, the specific metabolic and regulatory mechanisms of skin commensal bacteria in response to oligosaccharides remain unclear. This study aims to explore the influence of four oligosaccharides on the growth and metabolism of Staphylococcus epidermidis and further identify skin prebiotics that can enhance its probiotic effects on the skin. Fructooligosaccharides (FOS), isomaltooligosaccharide (IMO), galactooligosaccharides (GOS) and inulin were compared in terms of their impact on cell proliferation, SCFAs production of S. epidermidis CCSM0287 and the biofilm inhibition effect of their fermentation supernatants on Staphylococcus aureus CCSM0424. Furthermore, the effect of FOS on S. epidermidis CCSM0287 was analysed by the transcriptome analysis. All four oligosaccharides effectively promoted the growth of S. epidermidis CCSM0287 cells, increased the production of SCFAs, with FOS demonstrating the most significant effect. Analysis of the SCFAs indicated that S. epidermidis CCSM0287 predominantly employs oligosaccharides to produce acetic acid and isovaleric acid, differing from the SCFAs produced by gut microbiota. Among the four oligosaccharides, the addition of 2% FOS fermentation supernatant significantly inhibited S. aureus CCSM0424 biofilm formation. Furthermore, RNA sequencing revealed 162 differentially expressed genes (84 upregulated and 78 downregulated) of S. epidermidis CCSM0287 upon FOS treatment compared with glucose treatment. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis highlighted differences in the amino acid synthesis pathway, particularly in terms of arginine biosynthesis. FOS promotes cell proliferation, increases the SCFA production of S. epidermidis CCSM0287 and enhance the inhibition of S. aureus biofilm formation, suggesting that FOS serves as a potential prebiotic for strain S. epidermidis CCSM0287.

Cave Bats as Carriers of Extended Spectrum Beta-lactamase Produced by Escherichia coli from the Island of Lombok, Indonesia

Background: Escherichia coli is a Gram-negative commensal bacterium that normally resides in the digestive tract of humans and animals which is used as an indicator of the extended-spectrum beta-lactamase (ESBL) gene in an individual and can carry out gene transfer. Transmission of ESBL E. coli from wild animals has not been given much attention even though wild animals can act as vectors that spread ESBL in the environment so that it can pose a risk to public health. This research aims to identify the blaTEM gene which codes for ESBL in E. coli from bats that live in caves in Lombok, Indonesia. Methods: A total of 135 samples from bat rectal swabs were cultured on Eosin Methylene Blue Agar media and biochemical suspects were identified using the Indole, Methyl red, Voges-Proskauer, Citrate + and H2S (IMVIC) test. The E. coli bacteria obtained were tested for sensitivity using 7 classes of antibiotics, namely amoxicillin, ciprofloxacine, sulfamethoxazole/trimethoprim, tetracycline, gentamicin, cefotaxime and azithromycin. Bacteria that show multidrug resistance are subjected to PCR testing to detect the blaTEM gene. Result: Of the 135 test samples, it was found that 97 (71.85%) samples were positive for E. coli, 12 (12.37%) samples were Multidrug Resistance (MDR) and 2 (2.06%) samples had the blaTEM gene as the ESBL coding gene in bats that live in caves on the island of Lombok, Indonesia. The presence of the blaTEM gene in E. coli from bats can be indicated as a reservoir for MDR and ESBL transmission so that it can have an impact on public health.

EP568 - A rare case of necrotizing fasciitis caused by Streptococcus Anginosus and Staphylococcus Lugdunensis

Abstract Necrotizing fasciitis (NF) is a rare soft tissue infection involving the fascial planes, causing rapid and extensive tissue destruction. We present a unique case of NF in a patient with poor glycemic control caused by atypical pathogens. A 66-year-old female with a recent biopsy for a wart-like lesion developed a wound infection, with greenish-black exudate. Initial observations were: temperature: 39.4 degrees, heart rate: 105, GCS: 15/15. Physical examination revealed a grossly filiform lesion below her abdominal apron measuring 7cm in diameter. The patient was empirically treated with IV Co-Amoxiclav. Abnormal biochemistry results are shown below. Microbiological analysis revealed Streptococcus Anginosus, anaerobes and Staphylococcus Lugdunensis. CT abdomen revealed extensive inflammatory changes with gas and fluid collection and enlarged pelvic and inguinal lymph nodes. Two urgent surgical explorations and debridements, with meticulous wound care using negative pressure wound dressings (Prevena) were done. Intravenous Gentamicin, Benzylpenicillin, Clindamycin, Metronidazole and Flucloxacillin were administered for six weeks. The patient's clinical status gradually improved. Streptococcus Anginosus, that is part of the Streptococcus Milleri group and Staphylococcus Lugdunensis, a coagulase-negative Staphylococci (CoNS) are commensal organisms of the oral and gastrointestinal tracts. These organisms possess virulence factors that facilitate tissue invasion and destruction, causing an aggressive course of NF. This case highlights the importance of considering atypical pathogens, especially in patients with predisposing factors such as diabetes and obesity. Early recognition, prompt surgical intervention and a multidisciplinary approach are essential for improving outcomes in these cases.TestResultWhite Blood Cells (WBC)22,200 [WBCs per microliter]Sodium123 [mmol/L]Chloride88 [mmol/L]C-reactive protein (CRP)318 [mg/L]Blood Glucose19.3 [mmol/L]

Pangenome analysis of Clostridium scindens : a collection of diverse bile acid and steroid metabolizing commensal gut bacterial strains.

Clostridium scindens is a commensal gut bacterium capable of forming the secondary bile acids deoxycholic acid and lithocholic acid from the primary bile acids cholic acid and chenodeoxycholic acid, respectively, as well as converting glucocorticoids to androgens. Historically, only two strains, C. scindens ATCC 35704 and C. scindens VPI 12708, have been characterized in vitro and in vivo to any significant extent. The formation of secondary bile acids is important in maintaining normal gastrointestinal function, in regulating the structure of the gut microbiome, in the etiology of such diseases such as cancers of the GI tract, and in the prevention of Clostridium difficile infection. We therefore wanted to determine the pangenome of 34 cultured strains of C. scindens and a set of 200 metagenome-assembled genomes (MAGs) to understand the variability among strains. The results indicate that the 34 strains of C. scindens have an open pangenome with 12,720 orthologous gene groups, and a core genome with 1,630 gene families, in addition to 7,051 and 4,039 gene families in the accessory and unique (i.e., strain-exclusive) genomes, respectively. The core genome contains 39% of the proteins with predicted metabolic function, and, in the unique genome, the function of storage and processing of information prevails, with 34% of the proteins being in that category. The pangenome profile including the MAGs also proved to be open. The presence of bile acid inducible ( bai ) and steroid-17,20-desmolase ( des ) genes was identified among groups of strains. The analysis reveals that C. scindens strains are distributed into two clades, indicating the possible onset of C. scindens separation into two species, confirmed by gene content, phylogenomic, and average nucleotide identity (ANI) analyses. This study provides insight into the structure and function of the C. scindens pangenome, offering a genetic foundation of significance for many aspects of research on the intestinal microbiota and bile acid metabolism.

The Role of Host Genetics and Intestinal Microbiota and Metabolome as a New Insight into IBD Pathogenesis.

Inflammatory bowel disease (IBD) is an incurable, chronic disorder of the gastrointestinal tract whose incidence increases every year. Scientific research constantly delivers new information about the disease and its multivariate, complex etiology. Nevertheless, full discovery and understanding of the complete mechanism of IBD pathogenesis still pose a significant challenge to today's science. Recent studies have unanimously confirmed the association of gut microbial dysbiosis with IBD and its contribution to the regulation of the inflammatory process. It transpires that the altered composition of pathogenic and commensal bacteria is not only characteristic of disturbed intestinal homeostasis in IBD, but also of viruses, parasites, and fungi, which are active in the intestine. The crucial function of the microbial metabolome in the human body is altered, which causes a wide range of effects on the host, thus providing a basis for the disease. On the other hand, human genomic and functional research has revealed more loci that play an essential role in gut homeostasis regulation, the immune response, and intestinal epithelial function. This review aims to organize and summarize the currently available knowledge concerning the role and interaction of crucial factors associated with IBD pathogenesis, notably, host genetic composition, intestinal microbiota and metabolome, and immune regulation.

Quantifying trade-offs between therapeutic efficacy and resistance dissemination for enrofloxacin dose regimens in cattle

The use of antimicrobial drugs in food-producing animals contributes to the selection pressure on pathogenic and commensal bacteria to become resistant. This study aims to evaluate the existence of trade-offs between treatment effectiveness, cost, and the dynamics of resistance in gut commensal bacteria. We developed a within-host ordinary differential equation model to track the dynamics of antimicrobial drug concentrations and bacterial populations in the site of infection (lung) and the gut. The model was parameterized to represent enrofloxacin treatment for bovine respiratory disease (BRD) caused by Pastereulla multocida in cattle. Three approved enrofloxacin dosing regimens were compared for their effects on resistance on P. multocida and commensal E. coli: 12.5 mg/kg and 7.5 mg/kg as a single dose, and 5 mg/kg as three doses. Additionally, we explored non-FDA-approved regimes. Our results indicated that both 12.5 mg/kg and 7.5 mg/kg as a single dose scenario increased the most the treatment costs and prevalence of P. multocida resistance in the lungs, while 5 mg/kg as three doses increased resistance in commensal E. coli bacteria in the gut the most out of the approved scenarios. A proposed non-FDA-approved scenario (7.5 mg/kg, two doses 24 h apart) showed low economic costs, minimal P. multocida, and moderate effects on resistant E. coli. Overall, the scenarios that decrease P. multocida, including resistant P. multocida did not coincide with those that decrease resistant E. coli the most, suggesting a trade-off between both outcomes. The sensitivity analysis suggests that bacterial populations were the most sensitive to drug conversion factors into plasma (β\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$${\\beta}$$\\end{document}), elimination of the drug from the colon (ϑ\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$\\vartheta$$\\end{document}), fifty percent sensitive bacteria (P. multocida) killing effect (Ls50\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$${\ ext{L}}_{\ ext{s50}}$$\\end{document}), fifty percent of bacteria (E. coli) above ECOFF killing effect (Cr50\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$${\ ext{C}}_{\ ext{r50}}$$\\end{document}), and net drug transfer rate in the lung (γ\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$\\gamma$$\\end{document}) parameters.

Feeding intolerance after pediatric cardiac surgery is associated with dysbiosis, barrier dysfunction, and reduced short-chain fatty acids.

Congenital heart disease (CHD) is the most common birth defect, occurring in roughly 40,000 U.S. births annually. Malnutrition and feeding intolerance (FI) in CHD range from 30% to 42% and are associated with longer hospitalization and increased mortality. Cardiopulmonary bypass (CPB) required for surgical repair of CHD induces a systemic inflammatory response worsening intestinal dysbiosis and leading to intestinal epithelial barrier dysfunction (EBD), possibly contributing to postoperative FI. The objective of this study was to determine the relationship of postoperative FI with intestinal microbiome, short-chain fatty acids (SCFAs), and EBD in pediatric CHD after cardiac surgery. This was a prospective study of patients aged 0-15 years undergoing cardiac surgery with CPB. Samples were collected preoperatively and postoperatively to evaluate the gut microbiome, plasma EBD markers, short-chain fatty acids (SCFAs), and plasma cytokines. Clinical data were collected to calculate a FI score and evaluate patient status postoperatively. We enrolled 26 CPB patients and identified FI (n = 13). Patients with FI had unique microbial shifts with the reduced SCFA-producing organisms Rothia, Clostridium innocuum, and Intestinimonas. Patients who developed FI had associated elevations in the plasma EBD markers claudin-2 (P < 0.05), claudin-3 (P < 0.01), and fatty acid binding protein (P < 0.01). Patients with FI had reduced plasma and stool SCFAs. Mediation analysis showed the microbiome functional shift was associated with reductions in stool butyric and propionic acid in patients with FI. In conclusion, we provide novel evidence that intestinal dysbiosis, markers of EBD, and SCFA depletion are associated with FI. These data will help identify mechanisms and therapeutics to improve clinical outcomes following pediatric cardiac surgery.NEW & NOTEWORTHY Feeding intolerance contributes to postoperative morbidity following pediatric cardiac surgery. The intestinal microbiome and milieu play a vital role in gut function. Short-chain fatty acids are gut and cardioprotective metabolites produced by commensal bacteria and help maintain appropriate barrier function. Depletion of these metabolites and barrier dysfunction contribute to postoperative feeding intolerance following cardiac surgery. Identifying mechanistic targets to improve the intestinal milieu with the goal of improved nutrition and clinical outcomes is critical.

A Microphysiological System with an Anaerobic Air-Liquid Interface and Functional Mucus Layer for Coculture of Intestinal Bacteria and Primary Human Colonic Epithelium.

Coculture of intestinal bacteria with primary human intestinal epithelium provides a valuable tool for investigating host-colon bacterial interactions and for testing and screening therapeutics. However, most current intestinal model systems lack key physiological features of the in vivo colon, such as both a proper oxygen microenvironment and a mucus layer. In this work, a new in vitro colonic microphysiological system is demonstrated with a cell-derived, functional mucus that closely resembles the in vivo colonic mucosa and apical microenvironment by employing an anaerobic air-liquid interface culture. The human primary colon epithelial cells in this new in vitro system exhibit high cell viability (>98%) with ≈100 μm thick functional mucus layer on average. Successful coculture of model anaerobic gut bacterial strains Lactobacillus rhamnosus GG and Anaerobutyricum hallii without loss in human cell viability demonstrates that this new model can be an invaluable tool for future studies of the impact of commensal and pathogenic bacteria on the colon.

Antibacterial activity of Cinnamomum cassia against multiple drug resistant Klebsiella pneumoniae

The irrational use of antibiotics in various fields like medicine, animal husbandry and industrial sectors is probably a major cause of the emergence of antibiotic resistance in pathogens and commensal organisms. It is a big challenge to modern clinical practices, as it could lead to therapeutic failures, financial losses, and the creation of a gene pool that could be passed on to humans. The use of medicinal plants can be considered as an alternative, with a consequent impact on microbial resistance. Spices and hearbs have been traditionally used as coloring agents, flavoring agents, preservatives, food additives and medicine in India. The present work aimed to find out the antimicrobial activity of Cinnamomum cassiaon multi-drug resistant Klebsiella pneumoniae isolated from tap water. Natural spices might have anti-bacterial activity against other MDR bacteria and could be used for prevention of diseases.

Gynostemma pentaphyllum saponins shield mice from peanut allergy by modulation of gut microbiota: A novel approach for peanut allergy management

BackgroundFood allergies, particularly peanut (PN) allergies, are a growing concern, with fatal anaphylaxis incidents often reported. While palforzia is the sole FDA-approved drug for managing PN allergies, it is not universally effective. PurposeThis study aimed to investigate the potential of Gynostemma pentaphyllum saponins (GpS) as a novel therapeutic agent for PN allergy through modulation of gut microbiota, addressing the limitations of current treatments. MethodsTo elucidate the role of GpS on peanut allergy, we first built a PN-sensitized C57BL/6J model mice. Through comprehensive sequencing analysis, we identified Parabacteroides distasonis as a key bacterium triggering PN sensitization. Employing the same mouse model, GpS was evaluated for its effects on anaphylactic symptoms, serum immunoglobulin levels, and allergy-related biomarkers. 16S rRNA sequencing and transcriptomic analysis were applied to investigate the impact of GpS on the host's gut epithelium and microbiome. ResultsGpS treatment effectively reduced anaphylactic symptoms in PN-sensitized mice, as shown by decreased IgG1, total IgE, and PN-specific IgE levels. It also modulated the immune response by suppressing proinflammatory cytokines (IL-1β, IFN-γ, IL-21) and chemokines (CCL5, CCL12, CCL17, CCL22), while enhancing anti-inflammatory cytokines (IL-4, IL-10, IL-12, IL-13). Fecal microbial transplant from GpS-treated Model mice to PN-sensitized mice displayed anti-peanut allergy effects. Additionally, the administration of GpS-enhanced bacteria (Clostridium aldenese or Lactobacillus murinus), alleviated anaphylactic symptoms and reduced serum allergy markers in PN-sensitized mice. ConclusionTo conclude, we revealed the intestinal environment, signaling molecules, mucosal cytokines, and commensal microbial profiles in the peanut-sensitized mouse model. We further presented evidence for the protective effect of GpS against PN allergen sensitization by downregulating a series of food-allergy-associated biomarkers and cytokines via the modulation of gut bacteria. More importantly, supported by both in vitro and in vivo experiments, we demonstrated that the protective effect of GpS against PN-allergy is through the enhancement of two commensal bacteria, Clostridium aldenese, and Lactobacillus murinus.

A Linkable, Polycarbonate Gut Microbiome-Distal Tumor Chip Platform for Interrogating Cancer Promoting Mechanisms.

Gut microbiome composition is tied to diseases ranging from arthritis to cancer to depression. However, mechanisms of action are poorly understood, limiting development of relevant therapeutics. Organ-on-chip platforms, which model minimal functional units of tissues and can tightly control communication between them, are ideal platforms to study these relationships. Many gut microbiome models are published to date but devices are typically fabricated using oxygen permeable polydimethylsiloxane, requiring interventions to support anaerobic bacteria. To address this challenge, a platform is developed where the chips are fabricated entirely from gas-impermeable polycarbonate without tapes or gaskets. These chips replicate polarized villus-like structures of the native tissue. Further, they enable co-cultures of commensal anaerobic bacteria Blautia coccoides on the surface of gut epithelia for two days within a standard incubator. Another complication of commonly used materials in organ-on-chip devices is high ad-/absorption, limiting applications in high-resolution microscopy and biomolecule interaction studies. For future communication studies between gut microbiota and distal tumors, an additional polycarbonate chip design is developed to support hydrogel-embedded tissue culture. These chips enable high-resolution microscopy with all relevant processing done on-chip. Designed for facile linking, this platform will make a variety of mechanistic studies possible.

Clostridium perfringens chitinases, key enzymes during early stages of necrotic enteritis in broiler chickens.

The interaction between bacteria and the intestinal mucus is crucial during the early pathogenesis of many enteric diseases in mammals. A critical step in this process employed by both commensal and pathogenic bacteria focuses on the breakdown of the protective layer presented by the intestinal mucus by mucolytic enzymes. C. perfringens type G, the causative agent of necrotic enteritis in broilers, produces two glycosyl hydrolase family 18 chitinases, ChiA and ChiB, which display distinct substrate preferences. Whereas ChiB preferentially processes linear substrates such as chitin, ChiA prefers larger and more branched substrates, such as carbohydrates presented by the chicken intestinal mucus. Here, we show via crystal structures of ChiA and ChiB in the apo and ligand-bound forms that the two enzymes display structural features that explain their substrate preferences providing a structural blueprint for further interrogation of their function and inhibition. This research focusses on the roles of ChiA and ChiB in bacterial proliferation and mucosal attachment, two processes leading to colonization and invasion of the gut. ChiA and ChiB, either supplemented or produced by the bacteria, led to a significant increase in C. perfringens growth. In addition to nutrient acquisition, the importance of chitinases in bacterial attachment to the mucus layer was shown using an in vitro binding assay of C. perfringens to chicken intestinal mucus. Both an in vivo colonization trial and a necrotic enteritis trial were conducted, demonstrating that a ChiA chitinase mutant strain was less capable to colonize the intestine and was hampered in its disease-causing ability as compared to the wild-type strain. Our findings reveal that the pathogen-specific chitinases produced by C. perfringens type G strains play a fundamental role during colonization, suggesting their potential as vaccine targets.

Changes in Innate Immunity and Microbiome in Different Aging Phenotypes.

The indicators of innate immunity and the composition of the microbiome in the nasopharyngeal mucosa in centenarians with different aging phenotypes were analyzed. A significant increase in the expression of pattern-recognizing receptor genes (TLR2, TLR4, and NLRP3) and proinflammatory cytokines (IL1B, IL18) was shown in the group of centenarians with pathological aging phenotype. In centenarians with successful aging phenotype, increased diversity of the microbiome composition was observed. At the same time, a moderate inverse correlation was found between an increase in the growth of the commensal bacterium Streptococcus salivarius and a decrease in the expression of proinflammatory cytokine genes IL1B and IL18. These findings can serve as biomarkers for the timely identification of the phenotype of aging in senile and elderly people.

Set Up and Utilization of a Three-Dimensional In Vitro Bioreactor System for Human Intestinal Studies and Microbial Co-Cultures.

The study of human intestinal physiology and host-microbe interactions is crucial for understanding gastrointestinal health and disease. Traditional two-dimensional cell culture models lack the complexity of the native intestinal environment, limiting their utility in studying intestinal biology. Here, we present a detailed protocol for the set up and utilization of a three-dimensional (3D) in vitro bioreactor system for human intestinal studies and bacterial co-culture. This article outlines the design and assembly of the bioreactor system, scaffold fabrication, bacterial culture techniques, analysis methods, and troubleshooting tips. By providing step-by-step instructions, the goal is to enable other laboratories to utilize physiologically relevant tissue models of the human intestine, incorporating key features, such as nutrient flow, multiple human cell types, 3D architecture, and microbial communities. The incorporation of commensal bacteria into the bioreactor system allows for the investigation of complex host-microbe interactions, providing insight into gastrointestinal health and pathology. This article serves as a comprehensive resource for scientists seeking to advance their understanding of intestinal biology toward the development of novel therapeutic strategies for gastrointestinal disorders. © 2024 Wiley Periodicals LLC. Basic Protocol 1: Scaffold design Basic Protocol 2: Intestinal cell culture: Caco2 cells Basic Protocol 3: Intestinal cell culture: organoids Basic Protocol 4: Bioreactor design and set up Basic Protocol 5: Bacteria in 3D bioreactor set up Basic Protocol 6: Bacteria and drug dosing.

Liquid crystal nanoparticles for oral combination antibiotic therapies: A strategy towards protecting commensal gut bacteria during treatment

Antibiotics are essential for treating infections and reducing risks during medical interventions. However, many commonly used antibiotics lack the physiochemical properties for an efficient oral administration when treating systemic infection. Instead, we are reliant on intravenous delivery, which presents complications outside of clinical settings. Developing novel formulations for oral administration is a potential solution to this problem. We engineered hexosome and cubosome liquid crystal nanoparticles (LCNPs) characterized by small-angle X-ray scattering and cryogenic transmission electron microscopy, and could encapsulate the antibiotics vancomycin (VAN) and clarithromycin (CLA) with high loading efficiencies. By rationally choosing stable lipid building blocks, the loaded LCNPs demonstrated excellent resilience against enzymatic degradation in an in vitro gut model LCNP stability is crucial as premature antibiotic leakage can negatively impact the gut microbiota. In screens against the representative gut bacteria Enterococcus faecalis and Escherichia coli, our LCNPs provided a protective effect. Furthermore, we explored co-administration and dual loading strategies of VAN and CLA, and demonstrated effective loading, stability and protection for E. faecalis and E. coli. This work represents a proof of concept for the early-stage development of antibiotic-loaded LCNPs to treat systemic infection via oral administration, opening opportunities for combination antibiotic therapies.