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Background and Aims: Fistulizing Crohn’s Disease (fCD) affects up to 50% of people with Crohn’s Disease over their lifetime. Despite this high prevalence, the burden of disease, treatment and natural history in the current biologic era are poorly described. This study explores demographic, disease and treatment factors in a real-world Australasian cohort. Methods: A large real-world cohort of people with inflammatory bowel disease under routine care was interrogated in August 2023. Current fCD was defined as fistula(e) on most recent clinical, radiologic or endoscopic investigation; prior fCD was defined as the resolution of fistula(e) on most recent documentation. Results: Of 3075 people with Crohn’s Disease, 7.4% had current and 10.1% prior fCD (n = 224 & 311). Most patients were in Australia (77%), where 19.3% had current or previous fCD compared to 11% in New Zealand (P < .001). Patients with current or previous fCD were younger compared to those without (P = .003 & P < .001). Males were more commonly affected (P = .021). Current or prior fCD were more likely to be on biologic therapy (P < .001), with anti-tumor necrosis factor agents most frequently utilized. Conversely, those without fCD were more likely on Ustekinumab or Vedolizumab compared to current and prior fCD groups. People with fistulizing disease had higher hospitalization rates, while the prior fCD cohort had longer hospital admissions and more frequently required surgical intervention. Conclusion: People with fCD used more health-care resources, making this an important area for further research into care gaps to improve outcomes and optimal treatment approaches.

Abstract Background and Aim Pouchitis is a common complication after restorative ileal pouch–anal anastomosis following proctocolectomy for ulcerative colitis. Antibiotic‐dependent or antibiotic‐refractory chronic pouchitis (CP), which is a common cause of pouch failure affecting 15–20% of patients, is challenging to treat. The efficacy of second‐line immunomodulator and biologic therapy remains poorly defined. We present a pooled analysis of real‐world efficacy data from peer‐reviewed full‐text manuscripts, focusing on immunomodulator and biologic therapies in CP. Methods Embase and PubMed databases were searched for full‐text articles describing the treatment of CP. We performed a systematic review and pooled analysis of published studies to assess the efficacy of immunomodulators, including thiopurines and methotrexate, and biologics including antitumor necrosis factor, anti‐integrin, and interleukin‐12/23 antagonists. Clinical and endoscopic response and remission rates were combined for pooled analyses. Rates of treatment discontinuation and safety were also assessed. Results Pooled analysis comprised 20 full‐text articles (485 patients). Overall clinical response rate was 46% (95% CI: 35–59%) and clinical remission rate was 35% (95% CI: 21–52%). Overall endoscopic response and remission rates were 41% (95% CI: 18–68%) and 15% (95% CI: 5–39%), respectively. Individual agents' safety profile was reassuring, with vedolizumab being the most favorable. Conclusion The real‐world efficacy data of immunomodulators in the treatment of CP is insufficient. Vedolizumab and ustekinumab appeared effective and safe for CP, whereas anti‐TNFs showed higher rates of adverse events. The high heterogeneity within the studies is attributed to the real‐world study design, obfuscating drug efficacy comparisons across the studies. Further studies are required to define the comparative effectiveness of available treatments of CP.

Conclusion: EBD is a highly effective and safe procedure in both de novo and anastomotic strictures < 4 cm that can prevent or delay the need for surgical treatment.

Background & Aims: MUC13 cell surface mucin is highly expressed on the mucosal surface throughout the intestine, yet its role against bacterial infection is unknown. We investigated how MUC13 impacts Salmonella typhimurium (S Tm) infection and elucidated its mechanisms of action. Methods: Muc13-/- and wild-type littermate mice were gavaged with 2 isogenic strains of S Tm after pre-conditioning with streptomycin. We assessed clinical parameters, cecal histology, local and systemic bacterial load, and proinflammatory cytokines after infection. Cecal enteroids and epithelial cell lines were used to evaluate the mechanism of MUC13 activity after infection. The interaction between bacterial SiiE and MUC13 was assessed by using siiE-deficient Salmonella. Results: S Tm-infected Muc13-/- mice had increased disease activity, histologic damage, and higher local and systemic bacterial loads. Mechanistically, we found that S Tm binds to MUC13 through its giant SiiE adhesin and that MUC13 acts as a pathogen-binding decoy shed from the epithelial cell surface after pathogen engagement, limiting bacterial invasion. In addition, MUC13 reduces epithelial cell death and intestinal barrier breakdown by enhancing nuclear factor kappa B signaling during infection, independent of its decoy function. Conclusions: We show for the first time that MUC13 plays a critical role in antimicrobial defense against pathogenic S Tm at the intestinal mucosal surface by both acting as a releasable decoy limiting bacterial invasion and reducing pathogen-induced cell death. This further implicates the cell surface mucin family in mucosal defense from bacterial infection.

Background and Aims: There is clinical interest in the sustainability or otherwise of prebiotic, microbial, and antibiotic treatments to both prevent and treat inflammatory bowel diseases. This study examined the role of antibiotic manipulation of the gut microbiome to treat spontaneous and induced murine models of colitis. Methods: Symptomatic, histological, molecular, and microbial ecology and bioinformatic readouts were used to study the effect of a 10-day antibiotic cocktail and then follow-up over 2 months in the spontaneous Winnie colitis mouse preclinical model of ulcerative colitis and also the indirect antibiotic and Winnie microbiotic gavage effects in an acute dextran sodium sulfate–induced colitis model in wild-type mice. Results: The antibiotics elicited a striking reduction in both colitis symptoms and blinded histological colitis scores, together with a convergence of the microbial taxonomy of the spontaneous colitis and wild-type control mice, toward a taxonomic phenotype usually considered to be dysbiotic. The improvement in colitis was sustained over the following 8 weeks although the microbial taxonomy changed. In vitro, fecal waters from the antibiotic-treated colitis and wild-type mice suppressed the inflammatory tenor of colonic epithelial cells, and gavaged cecal slurries from these mice moderated the acute induced colitis. Conclusion: The results clearly show the possibility of a sustained remission of colitis by microbial manipulation, which is relevant to clinical management of inflammatory bowel diseases. The beneficial effects appeared to depend on the microbial metabolome rather than its taxonomy.

Background: There are limited treatment options for mental health comorbidities associated with Inflammatory Bowel Disease (IBD), although they have been shown to negatively affect the course of IBD and multiple important areas of functioning. Photobiomodulation (PBM) is a new therapeutic intervention using laser-generated low-powered light therapy that has shown early promise in alleviating fatigue, depression, and pain in chronic illness. Methods: This prospective, single-arm pilot study aims to assess the feasibility and efficacy of PBM in the treatment of fatigue, depression, and pain in youth with IBD. We will recruit 28 young adults with IBD who will receive PBM in addition to treatment as usual. The primary outcome will be fatigue, while secondary outcomes will include depression, pain, quality of life, inflammatory markers, alterations in microbiome composition, physical activity, and functioning. Outcome measures will be assessed at baseline, after a 10-week control period (pre-PBM), at 20 weeks (post-PBM), and at 30 weeks. Feasibility will be assessed by attendance, recruitment rates, and participants’ views of PBM. Mixed-effects linear regression modelling will be used to assess the PBM effect on continuous outcomes (fatigue, depression, anxiety and stress scores, and inflammation levels). Results: The study will provide preliminary indicators of PBM feasibility and efficacy in IBD.

Background & Aims: Chronic bowel inflammation increases the risk of colon cancer; colitis-associated cancer (CAC). Thiopurine treatments are associated with a reduction in dysplasia and CAC in inflammatory bowel disease (IBD). Abnormal Wnt/β-catenin signalling is characteristic of >90% of colorectal cancers. Immunosuppression by thiopurines is via Rac1 GTPase, which also affects Wnt/β-catenin signalling. Autophagy is implicated in colonic tumors, and topical delivery of the thiopurine thioguanine (TG) is known to alleviate colitis and augment autophagy. This study investigated the effects of TG in a murine model of CAC and potential mechanisms. Methods: Colonic dysplasia was induced by exposure to azoxymethane (AOM) and dextran sodium sulfate (DSS) in wild-type (WT) mice and mice harboring intestinal epithelial cell–specific deletion of autophagy related 7 gene (Atg7ΔIEC). TG or vehicle was administered intrarectally, and the effect on tumor burden and β-catenin activity was assessed. The mechanisms of action of TG were investigated in vitro and in vivo. Results: TG ameliorated DSS colitis in wild-type but not Atg7ΔIEC mice, demonstrating that anti-inflammatory effects of locally delivered TG are autophagy-dependent. However, TG inhibited CAC in both wild-type and Atg7ΔIEC mice. This was associated with decreased β-catenin activation/nuclear translocation demonstrating that TG’s inhibition of tumorigenesis occurred independently of anti-inflammatory and pro-autophagic actions. These results were confirmed in cell lines, and the dependency on Rac1 GTPase was demonstrated by siRNA knockdown and overexpression of constitutively active Rac1. Conclusions: Our findings provide evidence for a new mechanism that could be exploited to improve CAC chemoprophylactic approaches.

Background and Aim Inflammatory bowel diseases (IBD) are chronic inflammatory conditions of the gut resulting in a significant risk for malnutrition. The reported prevalence of malnutrition in inflammatory disease patients varies from 5.7 to 82.8%. The aim of this study was to measure the prevalence of malnutrition and its association with quality of life (QOL) in a cohort of Australian IBD outpatients. Methods A total of 107 consecutive patients (68 Crohn's disease, 35 ulcerative colitis, 4 indeterminate colitis) were enrolled in this cross‐sectional study. Demographic data were collected, and patients underwent a malnutrition assessment using the patient‐generated subjective global assessment. The RAND 36‐item health survey was used to measure QOL. Results Mild to moderate malnutrition was detected in 17 patients (16%). Malnourished patients were more likely to be underweight (P ≤ 0.01), have active disease (P ≤ 0.01), and have been admitted to hospital in the preceding 12 months (P ≤ 0.05). Malnourished patients had a significantly lower QOL in physical (P ≤ 0.01) and mental (P ≤ 0.01) health components. Patients with active or recently active disease had reduced QOL compared to patients in remission. Malnutrition factors predictive of poor physical health‐related QOL were pain (odds ratio [OR] = 12.8, 95% confidence interval [CI] 2.0–80.4) and unintentional weight loss (OR = 3.1 per kg lost, 95% CI 1.2–7.9). The predictor of poor mental health‐related QOL was early satiety (OR = 7.7, 95% CI 1.7–33.9). Conclusions The malnutrition prevalence for this population was 16%. Malnutrition was associated with being underweight, active disease, and increased number of hospital admissions. Disease activity and malnutrition were associated with poorer QOL.

Abstract Background and Aim Objective monitoring of disease activity is integral to therapeutic decision‐making in inflammatory bowel disease (IBD). Data are sparse on patients' perspectives of tools used to monitor disease activity in IBD. To evaluate patients' perspectives of gastrointestinal ultrasound (GIUS) performed during routine IBD clinical care, along with its impact on IBD‐specific knowledge. Methods Patients with a formal diagnosis of IBD who underwent GIUS at two tertiary IBD services between March 2017 and January 2019 participated in this prospective study. Participants completed a questionnaire measuring the acceptability, tolerability, and usefulness of GIUS using a visual analogue scale (VAS) from 0 (disagree) to 10 (strongly agree). Comparative acceptability of IBD monitoring tools and the impact of GIUS on IBD‐specific knowledge was measured. Results A total of 121 participants completed the questionnaire, with a mean age of 42 years (range 17–78), 54 (45%) males, and 79 (65%) Crohn's disease patients. In the overall population, GIUS was scored as highly acceptable for monitoring IBD (mean 9.20 ± 1.37) compared to colonoscopy (7.94 ± 2.30), stool sampling (8.17 ± 1.96), blood sampling (8.87 ± 1.62), and imaging (8.67 ± 1.60); P

A hallmark of Listeria (L.) monocytogenes pathogenesis is bacterial escape from maturing entry vacuoles, which is required for rapid bacterial replication in the host cell cytoplasm and cell-to-cell spread. The bacterial transcriptional activator PrfA controls expression of key virulence factors that enable exploitation of this intracellular niche. The transcriptional activity of PrfA within infected host cells is controlled by allosteric coactivation. Inhibitory occupation of the coactivator site has been shown to impair PrfA functions, but consequences of PrfA inhibition for L. monocytogenes infection and pathogenesis are unknown. Here we report the crystal structure of PrfA with a small molecule inhibitor occupying the coactivator site at 2.0 Å resolution. Using molecular imaging and infection studies in macrophages, we demonstrate that PrfA inhibition prevents the vacuolar escape of L. monocytogenes and enables extensive bacterial replication inside spacious vacuoles. In contrast to previously described spacious Listeria-containing vacuoles, which have been implicated in supporting chronic infection, PrfA inhibition facilitated progressive clearance of intracellular L. monocytogenes from spacious vacuoles through lysosomal degradation. Thus, inhibitory occupation of the PrfA coactivator site facilitates formation of a transient intravacuolar L. monocytogenes replication niche that licenses macrophages to effectively eliminate intracellular bacteria. Our findings encourage further exploration of PrfA as a potential target for antimicrobials and highlight that intra-vacuolar residence of L. monocytogenes in macrophages is not inevitably tied to bacterial persistence. Author summary Listeria monocytogenes is a bacterial pathogen that causes severe, often fatal disease in vulnerable individuals. Listeria monocytogenes infects host cells, where it needs to escape from its entry vacuole to replicate inside the host cell cytoplasm and spread to neighboring cells. Inability of Listeria to escape from vacuoles has been associated with long-term bacterial survival inside infected immune cells and chronic infection. We discovered that inhibition of the master regulator of the bacterial factors that orchestrate the intracellular lifestyle of Listeria monocytogenes traps the bacteria inside their entry vacuoles and facilitates extensive bacterial replication in spacious vacuoles. Yet, the majority of infected host immune cells survive the infection and ultimately clear the bacteria. Thus, our data demonstrate that intra-vacuolar existence of Listeria monocytogenes in immune cells is not inevitably tied to bacterial persistence and chronic infection. We provide high resolution structural insights into how an effective inhibitory molecule interacts with the master regulator of Listeria virulence, which may be exploited for rational drug design.

The gut microbiome is made up of hundreds of trillions of microorganisms that reside in a state of homeostatic balance within the healthy individual. Next generation sequencing has provided insight into the diversity of these microorganisms that reside within our gastrointestinal tract; despite developments in metabolomics and culturing techniques, the functions of many of these bacteria remain largely elusive. As such, research into the capacity of the gut microbiome to regulate immune homeostasis has revealed the importance of bacteria in human health, with the potential for exploiting these bacteria only now coming into focus.

Introduction Perianal fistulising Crohn’s disease (pfCD) can be somewhat treatment refractory. Higher infliximab trough levels (TLIs) may improve fistula healing rates; however, it remains unclear whether escalating infliximab therapy to meet higher TLI targets using proactive, or routine, therapeutic drug monitoring (TDM) improves outcomes. This randomised controlled trial aimed to assess whether infliximab therapy targeting higher TLIs guided by proactive TDM improves outcomes compared with standard therapy.Methods and analysis Patients with active pfCD will be randomised 1:1 to either the proactive TDM arm or standard dosing arm and followed up for 54 weeks. Patients in the proactive TDM arm will have infliximab dosing optimised to target higher TLIs. The targets will be TLI ≥ 25 µg/mL at week 2, ≥ 20 µg/mL at week 6 and ≥ 10 µg/mL during maintenance therapy. The primary objective will be fistula healing at week 32. Secondary objectives will include fistula healing, fistula closure, radiological fistula healing, patient-reported outcomes and economic costs up to 54 weeks. Patients in the standard dosing arm will receive conventional infliximab dosing not guided by TLIs (5 mg/kg at weeks 0, 2 and 6, and 5 mg/kg 8 weekly thereafter). Patients aged 18–80 years with pfCD with single or multiple externally draining complex perianal fistulas who are relatively naïve to infliximab treatment will be included. Patients with diverting ileostomies or colostomies and pregnant or breast feeding will be excluded. Fifty-eight patients per arm will be required to detect a 25% difference in the primary outcome measure, with 138 patients needed to account for an estimated 6.1% primary non-response rate and 10% dropout rate.Ethics and dissemination Results will be presented in peer-reviewed journals and international conferences. Ethics approval has been granted by the South Western Sydney Local Health District Human Research Ethics Committee in Australia.Trial registration number Australian New Zealand Clinical Trials Registry (ACTRN12621000023853); Pre-results.

Introduction The perinatal–postnatal family environment is associated with childhood outcomes including impacts on physical and mental health and educational attainment. Family longitudinal cohort studies collect in-depth data that can capture the influence of an era on family lifestyle, mental health, chronic disease, education and financial stability to enable identification of gaps in society and provide the evidence for changes in government in policy and practice.Methods and analysis The Queensland Family Cohort (QFC) is a prospective, observational, longitudinal study that will recruit 12 500 pregnant families across the state of Queensland (QLD), Australia and intends to follow-up families and children for three decades. To identify the immediate and future health requirements of the QLD population; pregnant participants and their partners will be enrolled by 24 weeks of gestation and followed up at 24, 28 and 36 weeks of gestation, during delivery, on-ward, 6 weeks postpartum and then every 12 months where questionnaires, biological samples and physical measures will be collected from parents and children. To examine the impact of environmental exposures on families, data related to environmental pollution, household pollution and employment exposures will be linked to pregnancy and health outcomes. Where feasible, data linkage of state and federal government databases will be used to follow the participants long term. Biological samples will be stored long term for future discoveries of biomarkers of health and disease.Ethics and dissemination Ethical approval has been obtained from the Mater Research Ethics (HREC/16/MHS/113). Findings will be reported to (1) QFC participating families; (2) funding bodies, institutes and hospitals supporting the QFC; (3) federal, state and local governments to inform policy; (4) presented at local, national and international conferences and (5) disseminated by peer-review publications.

Intestinal epithelial cell endoplasmic reticulum (ER) stress has been implicated in intestinal inflammation. It remains unclear whether ER stress is an initiator of or a response to inflammation. Winnie mice, carrying a Muc2 gene mutation resulting in intestinal goblet cell ER stress, develop spontaneous colitis with a depleted mucus barrier and increased bacterial translocation. This study aims to determine whether the microbiota was required for the development of Winnie colitis, and whether protein misfolding itself can initiate inflammation directly in absence of the microbiota. To assess the role of microbiota in driving Winnie colitis, WT and Winnie mice on the same background were rederived into the germ-free facility and housed in the Trexler-type soft-sided isolators. The colitis phenotype of these mice was assessed and compared to WT and Winnie mice housed within a specific pathogen-free facility. We found that Winnie colitis was substantially reduced but not abolished under germ-free conditions. Expression of inflammatory cytokine genes was reduced but several chemokines remained elevated in absence of microbiota. Concomitantly, ER stress was also diminished, although mucin misfolding persisted. RNA-Seq revealed that Winnie differentiated colon organoids have decreased expression of the negative regulators of the inflammatory response compared to WT. This data along with the increase in Mip2a chemokine expression, suggests that the epithelial cells in the Winnie mice are more responsive to stimuli. Moreover, the data demonstrate that intestinal epithelial intrinsic protein misfolding can prime an inflammatory response without initiating the unfolded protein response in the absence of the microbiota. However, the microbiota is necessary for the amplification of colitis in Winnie mice. Genetic predisposition to mucin misfolding in secretory cells initiates mild inflammatory signals. However, the inflammatory signal sets a forward-feeding cycle establishing progressive inflammation in the presence of microbiota. Abbreviations: Endoplasmic Reticulum: ER; Mucin-2: Muc-2; GF: Germ-Free; Inflammatory Bowel Disease: IBD

Abstract Objective Adoptive regulatory T cell (Treg) therapy is being trialled for the treatment of different autoimmune disorders, including inflammatory bowel diseases (IBD). In‐depth understanding of the biological variability of Treg in the human blood may be required to improve IBD immune monitoring and treatment strategies. Methods Through a combination of quantitative proteomic, multiparametric flow cytometry, RNA‐sequencing data analysis and functional assays on Treg enriched from the blood of ulcerative colitis (UC) patients and healthy controls, we investigated the association between CD49f expression, Treg phenotype and function, and UC disease activity. Results High‐dimensional analysis and filtering defined two distinct subsets of human Treg based on the presence or absence of CD49f with divergent transcriptional landscape and functional activities. CD49f negative (CD49f−) Treg are enriched for functional Treg markers and present significantly increased suppressive capacity. In contrast, CD49fhigh Treg display a pro‐inflammatory Th17‐like phenotype and accumulate in the blood of patients with UC. Dysregulation on CD49f Treg subsets in patients with UC correlate with disease activity. Conclusion Overall, our findings uncover the importance of CD49f expression on Treg in physiological immunity and in pathological autoimmunity.

Abstract Objectives Tumor‐associated autoantibodies (AAbs) in individuals with cancer can precede clinical diagnosis by several months to years. The objective of this study was to determine whether the primary immune response in form of IgM and gut mucosa‐associated IgA can aid IgG AAbs in the detection of early‐stage colorectal cancer (CRC). Methods We developed a novel protein array comprising 492 antigens seropositive in CRC. The array was used to profile IgG, IgM and IgA antibody signatures in 99 CRC patients and 99 sex‐ and age‐matched non‐cancer controls. A receiver operating curve (ROC), Kaplan–Meier survival analysis and univariate and multivariate Cox regression analyses were conducted. Results We identified a panel of 16 multi‐isotype AAbs with a cumulative sensitivity of 91% and specificity of 74% (AUC 0.90, 95% CI: 0.850–0.940) across all CRC stages. IgM and IgG isotypes were conversely associated with disease stage with IgM contributing significantly to improved stage I and II sensitivity of 96% at 78% specificity (AUC 0.928, 95% CI: 0.884–0.973). A single identified IgA AAb reached an overall sensitivity of 5% at 99% specificity (AUC 0.520, 95% CI: 0.440–0.601) balanced across all CRC stages. Kaplan–Meier analysis revealed that se33‐1 (ZNF638) IgG AAbs were associated with reduced 5‐year overall survival (log‐rank test, P = 0.012), whereas cumulative IgM isotype signatures were associated with improved 5‐year overall survival (log‐rank test, P = 0.024). Conclusion IgM AAbs are associated with early‐stage colorectal cancer. Combining IgG, IgM and IgA AAbs is a novel strategy to improve early diagnosis of cancers.

Enterococcus faecalis is an early coloniser of the human infant gut and contributes to the development of intestinal immunity. To better understand the functional capacity of E. faecalis, we constructed a broad host range RP4 mobilizable vector, pEHR513112, that confers chloramphenicol resistance and used a metaparental mating approach to isolate E. faecalis AHG0090 from a fecal sample collected from a healthy human infant. We demonstrated that E. faecalis AHG0090 is genetically tractable and could be manipulated using traditional molecular microbiology approaches. E. faecalis AHG0090 was comparable to the gold-standard anti-inflammatory bacterium Faecalibacterium prausnitzii A2-165 in its ability to suppress cytokine-mediated nuclear factor kappa B (NF-κB) activation in human gut-derived LS174T goblet cell like and Caco-2 enterocyte-like cell lines. E. faecalis AHG0090 and F. prausnitzii A2-165 produced secreted low molecular weight NF-κB suppressive peptidic bioactives. Both bioactives were sensitive to heat and proteinase K treatments although the E. faecalis AHG0090 bioactive was more resilient to both forms of treatment. As expected, E. faecalis AHG0090 suppressed IL-1β-induced NF-κB-p65 subunit nuclear translocation and expression of the NF-κB regulated genes IL-6, IL-8 and CXCL-10. Finally, we determined that E. faecalis AHG0090 is distantly related to other commensal strains and likely encodes niche factors that support effective colonization of the infant gut.

The polymorphism ATG16L1 T300A, associated with increased risk of Crohn’s disease, impairs pathogen defense mechanisms including selective autophagy, but specific pathway interactions altered by the risk allele remain unknown. Here, we use perturbational profiling of human peripheral blood cells to reveal that CLEC12A is regulated in an ATG16L1-T300A-dependent manner. Antibacterial autophagy is impaired in CLEC12A-deficient cells, and this effect is exacerbated in the presence of the ATG16L1∗300A risk allele. Clec12a−/− mice are more susceptible to Salmonella infection, supporting a role for CLEC12A in antibacterial defense pathways in vivo. CLEC12A is recruited to sites of bacterial entry, bacteria-autophagosome complexes, and sites of sterile membrane damage. Integrated genomics identified a functional interaction between CLEC12A and an E3-ubiquitin ligase complex that functions in antibacterial autophagy. These data identify CLEC12A as early adaptor molecule for antibacterial autophagy and highlight perturbational profiling as a method to elucidate defense pathways in complex genetic disease.

Genome-wide association studies and follow-up meta-analyses in Crohn's disease (CD) and ulcerative colitis (UC) have recently identified 163 disease-associated loci that meet genome-wide significance for these two inflammatory bowel diseases (IBD). These discoveries have already had a tremendous impact on our understanding of the genetic architecture of these diseases and have directed functional studies that have revealed some of the biological functions that are important to IBD (e.g. autophagy). Nonetheless, these loci can only explain a small proportion of disease variance (~14% in CD and 7.5% in UC), suggesting that not only are additional loci to be found but that the known loci may contain high effect rare risk variants that have gone undetected by GWAS. To test this, we have used a targeted sequencing approach in 200 UC cases and 150 healthy controls (HC), all of French Canadian descent, to study 55 genes in regions associated with UC. We performed follow-up genotyping of 42 rare non-synonymous variants in independent case-control cohorts (totaling 14,435 UC cases and 20,204 HC). Our results confirmed significant association to rare non-synonymous coding variants in both IL23R and CARD9, previously identified from sequencing of CD loci, as well as identified a novel association in RNF186. With the exception of CARD9 (OR = 0.39), the rare non-synonymous variants identified were of moderate effect (OR = 1.49 for RNF186 and OR = 0.79 for IL23R). RNF186 encodes a protein with a RING domain having predicted E3 ubiquitin-protein ligase activity and two transmembrane domains. Importantly, the disease-coding variant is located in the ubiquitin ligase domain. Finally, our results suggest that rare variants in genes identified by genome-wide association in UC are unlikely to contribute significantly to the overall variance for the disease. Rather, these are expected to help focus functional studies of the corresponding disease loci.

Staphylococcus epidermidis and Staphylococcus aureus are leading causes of hospital-acquired infections that have become increasingly difficult to treat due to the prevalence of antibiotic resistance in these organisms. The ability of staphylococci to produce biofilm is an important virulence mechanism that allows bacteria both to adhere to living and artificial surfaces and to resist host immune factors and antibiotics. Here, we show that the icaADBC locus, which synthesizes the biofilm-associated polysaccharide intercellular adhesin (PIA) in staphylococci, is required for the formation of a lethal S. epidermidis infection in the intestine of the model nematode Caenorhabditis elegans. Susceptibility to S. epidermidis infection is influenced by mutation of the C. elegans PMK-1 p38 mitogen-activated protein (MAP) kinase or DAF-2 insulin-signaling pathways. Loss of PIA production abrogates nematocidal activity and leads to reduced bacterial accumulation in the C. elegans intestine, while overexpression of the icaADBC locus in S. aureus augments virulence towards nematodes. PIA-producing S. epidermidis has a significant survival advantage over ica-deficient S. epidermidis within the intestinal tract of wild-type C. elegans, but not in immunocompromised nematodes harboring a loss-of-function mutation in the p38 MAP kinase pathway gene sek-1. Moreover, sek-1 and pmk-1 mutants are equally sensitive to wild-type and icaADBC-deficient S. epidermidis. These results suggest that biofilm exopolysaccharide enhances virulence by playing an immunoprotective role during colonization of the C. elegans intestine. These studies demonstrate that C. elegans can serve as a simple animal model for studying host-pathogen interactions involving staphylococcal biofilm exopolysaccharide and suggest that the protective activity of biofilm matrix represents an ancient conserved function for resisting predation.