Clostridioides difficile ferrosome organelles combat nutritional immunity.

Iron is indispensable for almost all forms of life but toxic at elevated levels1-4. To survive within their hosts, bacterial pathogens have evolved iron uptake, storage and detoxification strategies to maintain iron homeostasis1,5,6. Recent studies showed that three Gram-negative environmental anaerobes produce iron-containing ferrosome granules7,8. However, it remains unclear whether ferrosomes are generated exclusively by Gram-negative bacteria. The Gram-positive bacterium Clostridioides difficile is the leading cause of nosocomial and antibiotic-associated infections in the USA9. Here we report that C. difficile undergoes an intracellular iron biomineralization process and stores iron in membrane-bound ferrosome organelles containing non-crystalline iron phosphate biominerals. We found that a membrane protein (FezA) and a P1B6-ATPase transporter (FezB), repressed by both iron and the ferric uptake regulator Fur, are required for ferrosome formation and play an important role in iron homeostasis during transition from iron deficiency to excess. Additionally, ferrosomes are often localized adjacent to cellular membranes as shown by cryo-electron tomography. Furthermore, using two mouse models of C. difficile infection, we demonstrated that the ferrosome system is activated in the inflamed gut to combat calprotectin-mediated iron sequestration and is important for bacterial colonization and survival during C. difficile infection.

Fecal microbiota transplantation stimulates type 2 and tolerogenic immune responses in a mouse model.

OBJECTIVES: Clostridioides difficile infection (CDI) is the leading hospital-acquired infection in North America. While previous work on fecal microbiota transplantation (FMT), a highly effective treatment for CDI, has focused on colonization resistance mounted against C. difficile by FMT-delivered commensals, the effects of FMT on host gene expression are relatively unexplored. This study aims to identify transcriptional changes associated with FMT, particularly changes associated with protective immune responses. METHODS: Gene expression was assessed on day 2 and day 7 after FMT in mice after antibiotic-induced dysbiosis. Flow cytometry was also performed on colon and mesenteric lymph nodes at day 7 to investigate changes in immune cell populations. RESULTS: FMT administration after antibiotic-induced dysbiosis successfully restored microbial alpha diversity to levels of donor mice by day 7 post-FMT. Bulk RNA sequencing of cecal tissue at day 2 identified immune genes, including both pro-inflammatory and Type 2 immune pathways as upregulated after FMT. RNA sequencing was repeated on day 7 post-FMT, and expression of these immune genes was decreased along with upregulation of genes associated with restoration of intestinal homeostasis. Immunoprofiling on day 7 identified increased colonic CD45+ immune cells that exhibited dampened Type 1 and heightened regulatory and Type 2 responses. These include an increased abundance of eosinophils, alternatively activated macrophages, Th2, and T regulatory cell populations. CONCLUSION: These results highlight the impact of FMT on host gene expression, providing evidence that FMT restores intestinal homeostasis after antibiotic treatment and facilitates tolerogenic and Type 2 immune responses.

The impact of existing total anti-toxin B IgG immunity in outcomes of recurrent Clostridioides difficile infection.

Late anti-toxin-B humoral immunity acquired after treatment is important for preventing recurrent Clostridioides difficile infection. We prospectively-measured anti-toxin-B IgG and neutralization titers at diagnosis as potential early predictors of recurrence. High anti-toxin-B-IgG/neutralizing antibodies were associated with short-lasting protection within 6-weeks, however, no difference in recurrence risk was observed by 90-days post-infection.

Complement and IL-22: partnering up for border patrol.

Intestinal pathobionts that escape into the periphery can cause serious morbidity and death. In this issue of Immunity, Hasegawa et al. (2014) show that the host's protective measures against such events include interleukin-22-driven systemic elimination of pathobionts via complement regulation.

Interleukin-22 regulates the complement system to promote resistance against pathobionts after pathogen-induced intestinal damage.

Pathobionts play a critical role in disease development, but the immune mechanisms against pathobionts remain poorly understood. Here, we report a critical role for interleukin-22 (IL-22) in systemic protection against bacterial pathobionts that translocate into the circulation after infection with the pathogen Clostridium difficile. Infection with C. difficile induced IL-22, and infected Il22(-/-) mice harbored high numbers of pathobionts in extraintestinal organs despite comparable pathogen load and intestinal damage in mutant and wild-type mice. Pathobionts exhibited increased resistant against complement-mediated phagocytosis, and their intravenous administration resulted in high animal mortality. Selective removal of translocated commensals rescued Il22(-/-) mice, and IL-22 administration enhanced the elimination of pathobionts. Mechanistically, IL-22 augmented bacterial phagocytosis by increasing the expression and bacterial binding of complement C3. Our study demonstrates an unexpected role for IL-22 in controlling the elimination of pathobionts that enter the systemic circulation through the regulation of the complement system.

The Butyrate-Producing Bacterium Clostridium butyricum Suppresses Clostridioides difficile Infection via Neutrophil- and Antimicrobial Cytokine-Dependent but GPR43/109a-Independent Mechanisms.

Short-chain fatty acids, such as butyrate, are major gut microbial metabolites that are beneficial for gastrointestinal health. Clostridium butyricum MIYAIRI588 (CBM588) is a bacterium that produces a robust amount of butyrate and therefore has been used as a live biotherapeutic probiotic in clinical settings. Clostridioides difficile causes life-threatening diarrhea and colitis. The gut resident microbiota plays a critical role in the prevention of C. difficile infection (CDI), as the disruption of the healthy microbiota by antibiotics greatly increases the risk for CDI. We report that CBM588 treatment in mice significantly improved clinical symptoms associated with CDI and increased the number of neutrophils and Th1 and Th17 cells in the colonic lamina propria in the early phase of CDI. The protective effect of CBM588 was abolished when neutrophils, IFN-γ, or IL-17A were depleted, suggesting that induction of the immune reactants is required to elicit the protective effect of the probiotic. The administration of tributyrin, which elevates the concentration of butyrate in the colon, also increased the number of neutrophils in the colonic lamina propria, indicating that butyrate is a potent booster of neutrophil activity during infection. However, GPR43 and GPR109a, two G protein-coupled receptors activated by butyrate, were dispensable for the protective effect of CBM588. These results indicate that CBM588 and butyrate suppress CDI, in part by boosting antimicrobial innate and cytokine-mediated immunity.

Fecal Mycobiota Combined With Host Immune Factors Distinguish Clostridioides difficile Infection From Asymptomatic Carriage.

BACKGROUND & AIMS: Although the role of gut microbiota in Clostridioides difficile infection (CDI) has been well established, little is known about the role of mycobiota in CDI. Here, we performed mycobiome data analysis in a well-characterized human cohort to evaluate the potential of using gut mycobiota features for CDI diagnosis. METHODS: Stool samples were collected from 118 hospital patients, divided into 3 groups: CDI (n = 58), asymptomatic carriers (Carrier, n = 28), and Control (n = 32). The nuclear ribosomal DNA internal transcribed spacer 2 was sequenced using the Illumina HiSeq platform to assess the fungal composition. Downstream statistical analyses (including Alpha diversity analysis, ordination analysis, differential abundance analysis, fungal correlation network analysis, and classification analysis) were then performed. RESULTS: Significant differences were observed in alpha and beta diversity between patients with CDI and Carrier (P < .05). Differential abundance analysis identified 2 genera (Cladosporium and Aspergillus) enriched in Carrier. The ratio of Ascomycota to Basidiomycota was dramatically higher in patients with CDI than in Carrier and Control (P < .05). Correlations between host immune factors and mycobiota features were weaker in patients with CDI than in Carrier. Using 4 fungal operational taxonomic units combined with 6 host immune markers in the random forest classifier can achieve very high performance (area under the curve ∼92.38%) in distinguishing patients with CDI from Carrier. CONCLUSIONS: Our study provides specific markers of stool fungi combined with host immune factors to distinguish patients with CDI from Carrier. It highlights the importance of gut mycobiome in CDI, which may have been underestimated. Further studies on the diagnostic applications and therapeutic potentials of these findings are warranted.

Fusobacteriumnucleatum Adheres to Clostridioides difficile via the RadD Adhesin to Enhance Biofilm Formation in Intestinal Mucus.

BACKGROUND & AIMS: Although Clostridioides difficile infection (CDI) is known to involve the disruption of the gut microbiota, little is understood regarding how mucus-associated microbes interact with C difficile. We hypothesized that select mucus-associated bacteria would promote C difficile colonization and biofilm formation. METHODS: To create a model of the human intestinal mucus layer and gut microbiota, we used bioreactors inoculated with healthy human feces, treated with clindamycin and infected with C difficile with the addition of human MUC2-coated coverslips. RESULTS: C difficile was found to colonize and form biofilms on MUC2-coated coverslips, and 16S rRNA sequencing showed a unique biofilm profile with substantial cocolonization with Fusobacterium species. Consistent with our bioreactor data, publicly available data sets and patient stool samples showed that a subset of patients with C difficile infection harbored high levels of Fusobacterium species. We observed colocalization of C difficile and F nucleatum in an aggregation assay using adult patients and stool of pediatric patients with inflammatory bowel disease and in tissue sections of patients with CDI. C difficile strains were found to coaggregate with F nucleatum subspecies in vitro; an effect that was inhibited by blocking or mutating the adhesin RadD on Fusobacterium and removal of flagella on C difficile. Aggregation was shown to be unique between F nucleatum and C difficile, because other gut commensals did not aggregate with C difficile. Addition of F nucleatum also enhanced C difficile biofilm formation and extracellular polysaccharide production. CONCLUSIONS: Collectively, these data show a unique interaction of between pathogenic C difficile and F nucleatum in the intestinal mucus layer.

Gastrointestinal microbiota-mediated control of enteric pathogens.

The gastrointestinal (GI) microbiota is a complex community of microorganisms residing within the mammalian gastrointestinal tract. The GI microbiota is vital to the development of the host immune system and plays a crucial role in human health and disease. The composition of the GI microbiota differs immensely among individuals yet specific shifts in composition and diversity have been linked to inflammatory bowel disease, obesity, atopy, and susceptibility to infection. In this review, we describe the GI microbiota and its role in enteric diseases caused by pathogenic Escherichia coli, Salmonella enterica, and Clostridium difficile. We discuss the central role of the GI microbiota in protective immunity, resistance to enteric pathogens, and resolution of enteric colitis.

Endotoxin Acts Synergistically With Clostridioides difficile Toxin B to Increase Interleukin 1ß Production: A Potential Role for the Intestinal Biome in Modifying the Severity of C. difficile Colitis.

BACKGROUND: Inflammation is a crucial driver of host damage in patients with Clostridioides difficile colitis. We examined the potential for the intestinal microbiome to modify inflammation in patients with C. difficile colitis via the effects of gut-derived endotoxin on cytokine production. METHODS: Endotoxin from Escherichia coli and Pseudomonas aeruginosa as well as stool-derived endotoxin were tested for their ability to enhance interleukin 1ß (IL-1ß) and tumor necrosis factor alpha (TNF-α) production by toxin B-stimulated peripheral blood mononuclear cells. Inflammasome and Toll-like receptor 4 (TLR4) blocking studies were done to discern the importance of these pathways, while metagenomic studies were done to characterize predominant organisms from stool samples. RESULTS: Endotoxin significantly enhanced the ability of C. difficile toxin B to promote IL-1ß production but not TNF-α. The magnitude of this effect varied by endotoxin type and was dependent on combined inflammasome and TLR4 activation. Stool-derived endotoxin exhibited a similar synergistic effect on IL-1ß production with less synergy observed for stools that contained a high proportion of γ-proteobacteria. CONCLUSIONS: The ability of endotoxin to enhance IL-1ß production highlights a manner by which the microbiome can modify inflammation and severity of C. difficile disease. This information may be useful in devising new therapies for severe C. difficile colitis.

α-Galactosylceramide-Reactive NKT Cells Increase IgG1 Class Switch against a Clostridioides difficile Polysaccharide Antigen and Enhance Immunity against a Live Pathogen Challenge.

All clinical Clostridioides difficile strains identified to date express a surface capsule-like polysaccharide structure known as polysaccharide II (PSII). The PSII antigen is immunogenic and, when conjugated to a protein carrier, induces a protective antibody response in animal models. Given that CD1d-restricted natural killer T (NKT) cells promote antibody responses, including those against carbohydrates, we tested the hypothesis that immunization with PSII and a CD1d-binding glycolipid adjuvant could lead to enhanced protection against a live C. difficile challenge. We purified PSII from a clinical isolate of C. difficile and immunized B6 mice with PSII alone or PSII plus the CD1d-binding glycolipid α-galactosylceramide (α-GC). PSII-specific IgM and IgG titers were evident in sera from immunized mice. The inclusion of α-GC had a modest influence on isotype switch but increased the IgG1/IgG2c ratio. Enhanced protection against C. difficile disease was achieved by inclusion of the α-GC ligand and was associated with reduced bacterial numbers in fecal pellets. In contrast, NKT-deficient Traj18-/- mice were not protected by the PSII/α-GC immunization modality. Absence of NKT cells similarly had a modest effect on isotype switch, but ratios of IgG1/IgG2c decreased. These results indicate that α-GC-driven NKT cells move the humoral immune response against C. difficile PSII antigen toward Th2-driven IgG1 and may contribute to augmented protection. This study suggests that NKT activation represents a pathway for additional B-cell help that could be used to supplement existing efforts to develop vaccines against polysaccharides derived from C. difficile and other pathogens.

The Murine Neonatal Fc Receptor Is Required for Transport of Immunization-Induced C. difficile-Specific IgG to the Gut and Protection against Disease but Does Not Affect Disease Susceptibility.

The pathology associated with Clostridioides difficile disease is caused in large part by TcdB, an intracellular bacterial toxin that inactivates small GTPases. Despite C. difficile causing enteric disease, antitoxin IgG is a clear correlate of protection against infection-associated pathology. Immunization with TcdB-based immunogens or passive transfer of monoclonal antibodies specific for the TcdB carboxy-terminal domain (CTD) confers protection following C. difficile infection. Whether the mechanism by which circulating IgG is delivered to the gut depends on specific receptor-mediated transport or is solely reflective of infection-induced damage to the gut remains unclear. Here, we tested the hypothesis that neonatal Fc receptor (FcRn) is required for the delivery of systemic TcdB-specific IgG to the gut and protection against C. difficile-associated pathology. FcRn-expressing mice and FcRn-deficient littermates were immunized subcutaneously with Alhydrogel adjuvant-adsorbed CTD before challenge with live C. difficile spores. FcRn was required for the delivery of systemic TcdB-specific IgG to the gut and for vaccine-induced protection against C. difficile-associated disease. The lack of FcRn expression had minimal effects on the composition of the gut microbiome and did not affect susceptibility to C. difficile infection in nonimmunized mice. In further experiments, intraperitoneal injection of immune sera in FcRn-deficient mice led to the transport of protective IgG to the gut independently of infection, confirming a reported method of bypassing the FcRn. Our results reveal an FcRn-dependent mechanism by which systemic immunization-induced IgG protects the gut during enteric C. difficile infection. These findings may be beneficial for the targeting of C. difficile-specific IgG to the gut.

Clostridioides difficile Toxin A Remodels Membranes and Mediates DNA Entry Into Cells to Activate Toll-Like Receptor 9 Signaling.

BACKGROUND & AIMS: Clostridioides difficile toxin A (TcdA) activates the innate immune response. TcdA co-purifies with DNA. Toll-like receptor 9 (TLR9) recognizes bacterial DNA to initiate inflammation. We investigated whether DNA bound to TcdA activates an inflammatory response in murine models of C difficile infection via activation of TLR9. METHODS: We performed studies with human colonocytes and monocytes and macrophages from wild-type and TLR9 knockout mice incubated with TcdA or its antagonist (ODN TTAGGG) or transduced with vectors encoding TLR9 or small-interfering RNAs. Cytokine production was measured with enzyme-linked immunosorbent assay. We studied a transduction domain of TcdA (TcdA57-80), which was predicted by machine learning to have cell-penetrating activity and confirmed by synchrotron small-angle X-ray scattering. Intestines of CD1 mice, C57BL6J mice, and mice that express a form of TLR9 that is not activated by CpG DNA were injected with TcdA, TLR9 antagonist, or both. Enterotoxicity was estimated based on loop weight to length ratios. A TLR9 antagonist was tested in mice infected with C difficile. We incubated human colon explants with an antagonist of TLR9 and measured TcdA-induced production of cytokines. RESULTS: The TcdA57-80 protein transduction domain had membrane remodeling activity that allowed TcdA to enter endosomes. TcdA-bound DNA entered human colonocytes. TLR9 was required for production of cytokines by cultured cells and in human colon explants incubated with TcdA. TLR9 was required in TcdA-induced mice intestinal secretions and in the survival of mice infected by C difficile. Even in a protease-rich environment, in which only fragments of TcdA exist, the TcdA57-80 domain organized DNA into a geometrically ordered structure that activated TLR9. CONCLUSIONS: TcdA from C difficile can bind and organize bacterial DNA to activate TLR9. TcdA and TcdA fragments remodel membranes, which allows them to access endosomes and present bacterial DNA to and activate TLR9. Rather than inactivating the ability of DNA to bind TLR9, TcdA appears to chaperone and organize DNA into an inflammatory, spatially periodic structure.

Effective definition of low humoral response to Clostridioides difficile infection.

BACKGROUND: Determination of the humoral response to Clostridioides difficile (CD) toxins could be of great value in the management of patients with CD infection (CDI). METHODS: A prospective study was conducted on the clinical characteristics and humoral response in patients with CDI. Determination of ELISA IgG CD anti-toxin B (tgcBiomics, Germany) was performed. The following dilutions were planned for each patient, 1:100, 1: 200, 1: 400, 1: 800: 1: 1600. A significant concentration of antibody was considered to be present in each dilution if an optical density 0.2 units higher than the negative control of the technique was evident. RESULTS: Eighty-five patients were included during the study period, November 2018-February 2020. The median age was 73 years (interquartile range: 62.5-85 years), with female predominance (45 patients, 52.9%). Thirty-nine patients (45.9%) had a severe infection. Seven patients (8.2%) had suffered an episode of CDI in the previous three months. Seventeen patients (20%) had one or more recurrent episodes during the three-month follow-up: No patient died during admission or required surgery for severe-complicated infection. The incidence of recurrence in patients with no antibody detected at 1:400 dilution was 25.4% (16 patients) while it was 4.3% (one patient) in patients with antibody present at that dilution (p = 0.03). Liver cirrhosis was associated with higher humoral response against CD. CONCLUSIONS: Antibodies IgG CD anti-toxin B detection at a dilution of 1:400, using a B ELISA technique, effectively identified patients at increased risk of recurrence. This information could help assist in the management of patients.

Clostridioides difficile spores stimulate inflammatory cytokine responses and induce cytotoxicity in macrophages.

Clostridioides difficile is a gram-positive, spore-forming anaerobic bacterium, and the leading cause of antibiotic-associated diarrhea worldwide. During C. difficile infection, spores germinate in the presence of bile acids into vegetative cells that subsequently colonize the large intestine and produce toxins. In this study, we demonstrated that C. difficile spores can universally adhere to, and be phagocytosed by, murine macrophages. Only spores from toxigenic strains were able to significantly stimulate the production of inflammatory cytokines by macrophages and subsequently induce significant cytotoxicity. Spores from the isogenic TcdA and TcdB double mutant induced significantly lower inflammatory cytokines and cytotoxicity in macrophages, and these activities were restored by pre-exposure of the spores to either toxins. These findings suggest that during sporulation, spores might be coated with C. difficile toxins from the environment, which could affect C. difficile pathogenesis in vivo.

Molecular features of lipoprotein CD0873: A potential vaccine against the human pathogen Clostridioides difficile.

Clostridioides difficile is the primary cause of antibiotic-associated diarrhea and colitis, a healthcare-associated intestinal disease resulting in a significant fatality rate. Colonization of the gut is critical for C. difficile pathogenesis. The bacterial molecules essential for efficient colonization therefore offer great potential as vaccine candidates. Here we present findings demonstrating that the C. difficile immunogenic lipoprotein CD0873 plays a critical role in pathogen success in vivo We found that in a dixenic colonization model, a CD0873-positive strain of C. difficile significantly outcompeted a CD0873-negative strain. Immunization of mice with recombinant CD0873 prevented long-term gut colonization and was correlated with a strong secretory IgA immune response. We further present high-resolution crystal structures of CD0873, at 1.35-2.50 Å resolutions, offering a first view of the ligand-binding pocket of CD0873 and provide evidence that this lipoprotein adhesin is part of a tyrosine import system, an amino acid key in C. difficile infection. These findings suggest that CD0873 could serve as an effective component in a vaccine against C. difficile.

Clostridioides difficile Infection Induces an Inferior IgG Response to That Induced by Immunization and Is Associated with a Lack of T Follicular Helper Cell and Memory B Cell Expansion.

The intracellularly active bacterial toxin TcdB is a major Clostridioides difficile virulence factor that contributes to inflammation and tissue damage during disease. Immunization with an inactive TcdB fragment prevents C. difficile infection (CDI)-associated pathology. The protective immune response against inactive TcdB involves development of antigen-specific memory B cells and long-lived plasma cells that encode TcdB-neutralizing antibodies. Unlike the response to inactive TcdB, very little is known about the host humoral immune response to C. difficile and TcdB during primary and recurrent infection. Here, we used a murine model of C. difficile disease recurrence to demonstrate that an initial infection induced a serum IgM and mucosal IgA response against the toxin, but a low serum IgG response, which is associated with a lack of protection against disease during reinfection. Infection induced a partial expansion of the T follicular helper cell compartment, essential for B cell memory responses, and, consistent with that, failed to significantly expand the memory B cell compartment. Further, infection failed to stimulate the memory B cell compartment in preimmunized mice, although they were protected against associated disease. These results delineate the key humoral immune events that follow primary and recurrent C. difficile infection and provide a compelling inverse correlation between B cell memory and disease recurrence.

A Phase 2 Study Evaluating the Safety, Tolerability, and Immunogenicity of Two 3-Dose Regimens of a Clostridium difficile Vaccine in Healthy US Adults Aged 65 to 85 Years.

BACKGROUND: Clostridium difficile causes toxin-mediated nosocomial diarrhea and community-acquired infections; no preventive vaccine is licensed. In this phase 2 study, we explored safety, tolerability, and immunogenicity in older US adults of an investigational bivalent C. difficile vaccine that contains equal dosages of genetically and chemically detoxified toxins A and B. METHODS: Conducted from July 2015 through March 2017, 855 healthy adults aged 65-85 years from 15 US centers were randomized 3:3:1 to receive vaccine (100 or 200 µg) or placebo at 0, 1, and 6 months (month regimen) or 1, 8, and 30 days (day regimen). Serum toxin A- and B-specific neutralizing antibodies were measured. Participant-reported local reactions (LRs) and systemic events (SEs), adverse events (AEs), serious AEs, newly diagnosed chronic medical conditions, and immediate AEs were recorded. RESULTS: The 200-µg dose level elicited higher immune responses than the 100-µg dose level across regimens. Compared with the day regimen, the month regimen induced stronger and more persistent immune responses that remained elevated 12 months after dose 3. Responses peaked at month 7 (month regimen) and day 37 (day regimen). LRs (primarily injection site pain) were more frequent in vaccine recipients than controls; SE frequency was similar across groups. More related AEs were reported in the day regimen group than the month regimen group. CONCLUSIONS: The C. difficile vaccine was safe, well tolerated, and immunogenic in healthy US adults aged 65-85 years. Immune responses were particularly robust in the 200-µg month regimen group. These results support continued vaccine development. CLINICAL TRIALS REGISTRATION: NCT02561195.

Epithelium-specific MyD88 signaling, but not DCs or macrophages, control acute intestinal infection with Clostridium difficile.

Infection with Clostridium difficile is one of the major causes of health care acquired diarrhea and colitis. Signaling though MyD88 downstream of TLRs is critical for initiating the early protective host response in mouse models of C. difficile infection (CDI). In the intestine, MyD88 is expressed in various tissues and cell types, such as the intestinal epithelium and mononuclear phagocytes (MNP), including DC or macrophages. Using a genetic gain-of-function system, we demonstrate here that restricting functional MyD88 signaling to the intestinal epithelium, but also to MNPs is sufficient to protect mice during acute CDI by upregulation of the intestinal barrier function and recruitment of neutrophils. Nevertheless, we also show that mice depleted for CD11c-expressing MNPs in the intestine display no major defects in mounting an effective inflammatory response, indicating that the absence of these cells is irrelevant for inducing host protection during acute infection. Together, our results highlight the importance of epithelial-specific MyD88 signaling and demonstrate that although functional MyD88 signaling in DC and macrophages alone is sufficient to correct the phenotype of MyD88-deficiency, these cells do not seem to be essential for host protection in MyD88-sufficient animals during acute infection with C. difficile.

Clostridium difficile-associated disease and Helicobacter pylori seroprevalence: A case-control study.

BACKGROUND: Helicobacter pylori inhabits the stomach and causes persistent inflammation, with changes in gastric acidity. However, it is unclear whether the presence of H pylori plays a role in Clostridium difficile-associated disease (CDAD). The study's aim was to examine relationships of H pylori seroprevalence and serum pepsinogens (PGs), as markers of gastric inflammation, with CDAD. MATERIALS AND METHODS: A case-control study was conducted among 49 CDAD cases and 54 controls (median age 82 years). Using enzyme-linked immunosorbent assays, sera were tested for H pylori IgG antibody, and PGI and PGII levels. Helicobacter pylori-positive samples were tested for IgG antibody to recombinant cytotoxin-associated gene A (CagA) virulent protein. Logistic regression models were fitted. RESULTS: Cases and controls were comparable in age (P = .5) and sex distribution (females 62% vs 57%, P = .6). Helicobacter pylori IgG seroprevalence was 47%, of whom 23% were CagA seropositives. Among cases compared to controls, 43% vs 28% were H pylori seropositive but lacking CagA IgG antibody: adjusted odd ratio (OR) 3.43 (95% confidence intervals [CI] 1.29-9.10); 18% vs 4% were positive for CagA phenotype: adjusted OR 9.32 (95% CI 1.61-53.76). This association was not affected by PG levels. CONCLUSIONS: Helicobacter pylori infection, especially with CagA virulent phenotype, might predispose to C difficile infection in elderly patients.