Transglutaminase 3 crosslinks the secreted gel-forming mucus component Mucin-2 and stabilizes the colonic mucus layer

The colonic mucus layer is organized as a two-layered system providing a physical barrier against pathogens and simultaneously harboring the commensal flora. The factors contributing to the organization of this gel network are not well understood. In this study, the impact of transglutaminase activity on this architecture was analyzed. Here, we show that transglutaminase TGM3 is the major transglutaminase-isoform expressed and synthesized in the colon. Furthermore, intrinsic extracellular transglutaminase activity in the secreted mucus was demonstrated in vitro and ex vivo. Absence of this acyl-transferase activity resulted in faster degradation of the major mucus component the MUC2 mucin and changed the biochemical properties of mucus. Finally, TGM3-deficient mice showed an early increased susceptibility to Dextran Sodium Sulfate-induced colitis. Here, we report that natural isopeptide cross-linking by TGM3 is important for mucus homeostasis and protection of the colon from inflammation, reducing the risk of colitis.

Short peptides derived from EntV inhibit virulence and biofilm formation in Candida albicans

Candida albicans exists as a member of the commensal flora of the skin and gut where many complex polymicrobial interactions occur with genera such as Pseudomonas, Staphylococcus, and Streptococcus. Some of these interactions potentiate or inhibit virulence. The bacterial gastrointestinal commensal speciesEnterococcus faecalisproduces a small peptide, EntV, that modulates C. albicans virulence. The active 68 amino acid EntV peptide inhibits biofilm formationin vitro; biofilm-related infections are difficult to treat with current therapeutics. EntV also attenuates fungal virulence in a Caenorhabditis elegansinfection model and a murine oral candidiasis model. We sought to identify the regions of EntV responsible for the anti-fungal activity, and based on structural information, we hypothesized that it could be localized to a single helix of the mature peptide. In this study, we report that smaller peptides derived from this helix ranging from 12 to 16 amino acids have equal to improved efficacy in inhibiting C. albicans virulence andbiofilm formation. These smaller peptides attenuate virulence in the C. elegans infection model, inhibit initial adhesion to abiotic surfaces, and reduce the size of mature biofilms measured by confocal microscopy. Further trimming of these peptides to fewer than 11 amino acids reduces and eventually eliminates activity. These data indicate that EntV-derived peptides warrant further investigation as potential non-fungicidal additives to medical devices and antifungal therapeutics.

Cross-Talk between Probiotic Nissle 1917 and Human Colonic Epithelium Affects the Metabolite Composition and Demonstrates Host Antibacterial Effect

Colonic epithelium–commensal interactions play a very important role in human health and disease development. Colonic mucus serves as an ecologic niche for a myriad of commensals and provides a physical barrier between the epithelium and luminal content, suggesting that communication between the host and microbes occurs mainly by soluble factors. However, the composition of epithelia-derived metabolites and how the commensal flora influences them is less characterized. Here, we used mucus-producing human adult stem cell-derived colonoid monolayers exposed apically to probiotic E. coli strain Nissle 1917 to characterize the host–microbial communication via small molecules. We measured the metabolites in the media from host and bacterial monocultures and from bacteria-colonoid co-cultures. We found that colonoids secrete amino acids, organic acids, nucleosides, and polyamines, apically and basolaterally. The metabolites from host-bacteria co-cultures markedly differ from those of host cells grown alone or bacteria grown alone. Nissle 1917 affects the composition of apical and basolateral metabolites. Importantly, spermine, secreted apically by colonoids, shows antibacterial properties, and inhibits the growth of several bacterial strains. Our data demonstrate the existence of a cross-talk between luminal bacteria and human intestinal epithelium via metabolites, which might affect the numbers of physiologic processes including the composition of commensal flora via bactericidal effects.

Microbiota Signals Suppress B Lymphopoiesis With Aging in Mice

Aging is associated with significant changes in hematopoiesis that include a shift from lymphopoiesis to myelopoiesis and an expansion of phenotypic hematopoietic stem cells (HSCs) with impaired self-renewal capacity and myeloid-skewed lineage differentiation. Signals from commensal flora support basal myelopoiesis in young mice; however, their contribution to hematopoietic aging is largely unknown. Here, we characterize hematopoiesis in young and middle-aged mice housed under specific pathogen free (SPF) and germ-free (GF) conditions. The marked shift from lymphopoiesis to myelopoiesis that develops during aging of SPF mice is mostly abrogated in GF mice. Compared with aged SPF mice, there is a marked expansion of B lymphopoiesis in aged GF mice, which is evident at the earliest stages of B cell development. The expansion of phenotypic and functional HSCs that occurs with aging is similar in SPF and GF mice. However, HSCs from young GF mice have increased lymphoid lineage output, and the aging-associated expansion of myeloid-biased HSCs is significantly attenuated in GF mice. Consistent with these data, RNA expression profiling of phenotypic HSCs from aged GF mice show enrichment for non-myeloid biased HSCs. Surprisingly, the RNA expression profiling data also suggest that inflammatory signaling is increased in aged GF HSCs compared with aged SPF HSCs. Collectively, these data suggest that microbiota-related signals suppress B lymphopoiesis at multiple stages of development and contribute to the expansion of myeloid-biased HSCs that occurs with aging.

Case report of an unusual hepatic abscess caused by Actinomyces odontolyticus in a patient with human immunodeficiency virus infection

Background Actinomyces odontolyticus is not commonly recognized as a causative microbe of liver abscess. The detection and identification of A. odontolyticus in laboratories and its recognition as a pathogen in clinical settings can be challenging. However, in the past decades, knowledge on the clinical relevance of A. odontolyticus is gradually increasing. A. odontolyticus is the dominant oropharyngeal flora observed during infancy [Li et al. in Biomed Res Int 2018:3820215, 2018]. Herein we report a case of severe infection caused by A. odontolyticus in an immunocompromised patient with disruption of the gastrointestinal (GI) mucosa. Case presentation We present a unique case of a patient with human immunodeficiency virus infection who was admitted due to liver abscess and was subsequently diagnosed as having coinfection of A. odontolyticus, Streptococcus constellatus, and Candida albicans during the hospital course. The empirical antibiotics metronidazole and ceftriaxone were replaced with the intravenous administration of fluconazole and ampicillin. However, the patient’s condition deteriorated, and he died 3 weeks later. Conclusion This report is one of the first to highlight GI tract perforation and its clinical relevance with A. odontolyticus infection. A. odontolyticus infection should be diagnosed early in high-risk patients, and increased attention should be paid to commensal flora infection in immunocompromised individuals.

Tbet expression by Tregs is needed to protect against Th1-mediated immunopathology during Toxoplasma infection in mice

T. gondii infection has proven to be an ideal model to understand the delicate balance between protective immunity and immune-mediated pathology during infection. Lethal infection causes a collapse of Tregs mediated by loss of IL-2, and conversion of Tregs to IFNγ producing cells. Importantly, these Tregs highly express the Th1 transcription factor Tbet. To determine the role of Tbet in Tregs, we infected Tbx21f/f-Foxp3YFPCre and control Foxp3YFPCre mice with the type II strain of T. gondii, ME49. The majority of Tbx21f/f-Foxp3YFPCre mice succumb to a non-lethal acute infection. Notably, parasite burden is comparable between Tbx21f/f-Foxp3YFPCre and Foxp3YFPCre control mice. We found that Tbx21f/f-Foxp3YFPCre mice have significantly higher serum levels of proinflammatory cytokines IFNγ and TNFα, suggestive of a heightened immune response. To test if CD4+ T cells were driving immunopathology, we treated Tbx21f/f-Foxp3YFPCre mice with anti-CD4 depleting antibody and partially rescued these mice. Broad spectrum antibiotic treatment also improved survival, demonstrating a role for commensal flora in immunopathology in Tbx21f/f-Foxp3YFPCre mice. RNA-seq analysis reinforced that Tbet regulates several key cellular pathways, including chromosome segregation, cytokine receptor activity and cell cycle progression, that help to maintain fitness in Tregs during Th1 responses. Taken together, our data shows an important role for Tbet in Tregs in preventing lethal immunopathology during Toxoplasma gondii infection, further highlighting the protective role of Treg plasticity to self and microbiota.

Hospitalization and colonization by methicillin-resistant Staphylococcus aureus in the surgical department of 03 health facilities in the Ndé division, West-Cameroon

Background Commensal flora colonization during hospitalization by bacteria is the first step for nosocomial infections while antibiotic resistance reduces therapeutic options. In aim to control this phenomenon, we initiated this study to describe the impact of hospitalization on colonization by methicillin-resistant Staphylococcus aureus in the surgical department of 03 health facilities in the Ndé division, West-Cameroon. Methods This study was carried out on patients admitted for surgery in 03 health facilities of the Ndé division, West-Cameroon (District Hospital of Bangangté, Protestant Hospital of Bangwa and Cliniques Universitaires des Montagnes). After obtaining ethical clearance and authorizations, nasal swabs were performed at admission and discharge, with the aim of isolating bacteria and performing their antibiotic susceptibility tests. Informations on each participant's antibiotic therapy were recorded. Laboratory investigations were carried out according to standard protocols (CASFM, 2019). Results The most commonly used antibiotics were β-lactams. A total of 104 nasal swabs were performed on 52 patients who agreed to participate to the study. From the analysis, 110 (57 at admission versus 53 at discharge) Staphylococcus isolates were obtained. Overall, susceptibility testing showed that antibiotic resistance rates were higher at discharge than at admission; with significant differences between the susceptibility profiles obtained at admission and discharge for β-lactams and not significant for fluoroquinolones and aminoglycosides. Globally, frequency of nasal carriage of methicillin-resistant Staphylococcus aureus at discharge 16 (30.77%) was significantly higher than at admission 07 (13.46%) with Chi-2 = 4.52 and p = 0.0335. Conclusion The high rates of antibiotic resistance of bacteria isolated at discharge compared to those isolated at admission obtained in the present investigation, highlights the important role that hospitalization plays in the selection and dissemination of methicillin-resistant Staphylococcus aureus and colonization by these bacteria in health structures of Ndé division. As a result, further investigations to find the factors that promote this phenomenon should be carried out.

Transglutaminase 3 crosslinks secreted MUC2 and stabilizes the colonic mucus layer

The colonic mucus layer is organized as a two-layered system providing a physical barrier against pathogens and simultaneously harboring the commensal flora. The factors contributing to the organization of this gel network are not well understood. In this study, the impact of transglutaminase activity on this architecture was analyzed. Here, we show that transglutaminase TGM3 is the major TGM isoform expressed and synthesized in the colon. Furthermore, intrinsic extracellular TGM activity in the secreted mucus was demonstrated in vitro and ex vivo. Absence of this acyl-transferase activity resulted in faster degradation of the major mucus component the MUC2 mucin and changed the biochemical properties of mucus. Finally, TGM3-deficient mice showed an early increased susceptibility to DSS-induced colitis. Thus, these observations suggest that natural isopeptide cross-linking by TGM3 is important for mucus homeostasis and protection of the colon from inflammation, a suggested pre-stage of colon carcinoma.

Understanding and exploiting the response of EHEC O157:H7 to human gastrointestinal chemical signals

Enterohemorrhagic Escherichia coli (EHEC) is a clinically relevant foodborne pathogen, resulting in over 95,000 cases of EHEC-associated illness and 60 deaths each year in the US alone. Since EHEC is a continuous global issue with new outbreaks constantly occurring, the development of new therapeutic strategies is vital to minimizing the cases of infection seen each year. A key aspect in new drug development is the identification of vulnerabilities in EHEC’s pathogenicity, in particular, during its transit through the human gastrointestinal (GI) tract. As EHEC passes through the human GI tract to its site of colonization in the large intestine, it faces a multitude of host assaults including acute acid stress in the stomach, bile salt stress and cationic antimicrobial peptide exposure in the small intestine, and short chain fatty acid (SCFA) stress in the large intestine. The research carried out in this doctoral dissertation focuses on understanding how EHEC senses chemical cues from the host’s innate immune responses and how this knowledge can be exploited to develop effective antimicrobial strategies. Our findings successfully demonstrate that a novel antimicrobial peptide ameliorates infection in a mouse model of infection by enhancing acid-induced pathogen killing during gastric passage, and that the DNAbinding protein, Dps, plays a significant role in protecting EHEC against peptide killing. Moreover, this research successfully shows that varying concentrations of SCFAs result in differential modulation of EHEC virulence – a finding that contributes to our understanding of the role of diet and commensal flora in host susceptibility to infection. Together the findings of this research demonstrate how the selected innate host defences throughout the human GI tract can be exploited and/or manipulated to effectively prevent infection by the human pathogen EHEC.

Understanding and exploiting the response of EHEC O157:H7 to human gastrointestinal chemical signals

Enterohemorrhagic Escherichia coli (EHEC) is a clinically relevant foodborne pathogen, resulting in over 95,000 cases of EHEC-associated illness and 60 deaths each year in the US alone. Since EHEC is a continuous global issue with new outbreaks constantly occurring, the development of new therapeutic strategies is vital to minimizing the cases of infection seen each year. A key aspect in new drug development is the identification of vulnerabilities in EHEC’s pathogenicity, in particular, during its transit through the human gastrointestinal (GI) tract. As EHEC passes through the human GI tract to its site of colonization in the large intestine, it faces a multitude of host assaults including acute acid stress in the stomach, bile salt stress and cationic antimicrobial peptide exposure in the small intestine, and short chain fatty acid (SCFA) stress in the large intestine. The research carried out in this doctoral dissertation focuses on understanding how EHEC senses chemical cues from the host’s innate immune responses and how this knowledge can be exploited to develop effective antimicrobial strategies. Our findings successfully demonstrate that a novel antimicrobial peptide ameliorates infection in a mouse model of infection by enhancing acid-induced pathogen killing during gastric passage, and that the DNAbinding protein, Dps, plays a significant role in protecting EHEC against peptide killing. Moreover, this research successfully shows that varying concentrations of SCFAs result in differential modulation of EHEC virulence – a finding that contributes to our understanding of the role of diet and commensal flora in host susceptibility to infection. Together the findings of this research demonstrate how the selected innate host defences throughout the human GI tract can be exploited and/or manipulated to effectively prevent infection by the human pathogen EHEC.