BackgroundInflammatory bowel diseases (IBD) are a group of complex and multifactorial disorders with unknown etiology. Chronic intestinal inflammation develops against resident intestinal bacteria in genetically susceptible hosts. We hypothesized that host intestinal immunoglobulin (Ig) G can be used to identify bacteria involved in IBD pathogenesis.ResultsIgG-bound and -unbound microorganisms were collected from 32 pediatric terminal ileum aspirate washes during colonoscopy [non-IBD (n = 10), Crohn disease (n = 15), and ulcerative colitis (n = 7)], and composition was assessed using the Illumina MiSeq platform. In vitro analysis of invasive capacity was evaluated by fluorescence in situ hybridization and gentamicin invasion assay; immune activation was measured by qPCR. Despite considerable inter-individual variations, IgG binding favored specific and unique mucosa-associated species in pediatric IBD patients. Burkholderia cepacia, Flavonifractor plautii, and Rumminococcus sp. demonstrated increased IgG binding, while Pseudomonas ST29 demonstrated reduced IgG binding, in IBD. In vitro validation confirmed that B. cepacia, F. plautii, and Rumminococcus display invasive potential while Pseudomonas protogens did not.ConclusionUsing IgG as a marker of pathobionts in larger patient cohorts to identify microbes and elucidate their role in IBD pathogenesis will potentially underpin new strategies to facilitate development of novel, targeted diagnostic, and therapeutic approaches. Interestingly, this method can be used beyond the scope of this manuscript to evaluate altered gut pathobionts in a number of diseases associated with altered microbiota including arthritis, obesity, diabetes mellitus, alcoholic liver disease, cirrhosis, metabolic syndrome, and carcinomas.Electronic supplementary materialThe online version of this article (10.1186/s40168-018-0604-3) contains supplementary material, which is available to authorized users.

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Background & Aims Very early onset inflammatory bowel diseases (VEOIBD), including infant disorders, are a diverse group of diseases found in children less than 6 years of age. They have been associated with several gene variants. We aimed to identify genes that cause VEOIBD. Methods We performed whole-exome sequencing of DNA from 1 infants with severe enterocolitis and her parents. Candidate gene mutations were validated in 40 pediatric patients and functional studies were carried out using intestinal samples and human intestinal cell lines. Results We identified compound heterozygote mutations in the tetratricopeptide repeat domain 7 (TTC7A) gene in an infant from non-consanguineous parents with severe exfoliative apoptotic enterocolitis; we also detected the mutations in 2 unrelated families, each with 2 affected siblings. TTC7A interacts with EFR3 homolog B (EFR3B) to regulate phosphatidylinositol 4-kinase (PI4KA) at the plasma membrane. Functional studies demonstrated that TTC7A is expressed in human enterocytes. The mutations we identified in TTC7A result in either mislocalization or reduced expression of TTC7A. PI4KA was found to co-immunoprecipitate with TTC7A; the identified TTC7A mutations reduced this binding. Knockdown of TTC7A in human intestinal-like cell lines reduced their adhesion, increased apoptosis, and decreased production of phosphatidylinositol 4-phosphate. Conclusion In a genetic analysis, we identified loss of function mutations in TTC7A in 5 infants with VEOIBD. Functional studies demonstrated that the mutations cause defects in enterocytes and T cells that lead to severe apoptotic enterocolitis. Defects in the PI4KA–TTC7A–EFR3B pathway are involved in the pathogenesis of VEOIBD.

These findings suggest an underlying defect in the UC-afflicted intestinal tract even in the absence of inflammation, implicating barrier and microbial changes as primary abnormalities in UC that may play a causative role in disease development.

Helicobacter pylori activates the transcription factor NF-B, leading to proinflammatory cytokine production by gastric epithelial cells. However, the receptors for the initial bacterial interaction with host cells which activate downstream signaling events have not been completely defined. Recently, it has been shown that microbial components activate Toll-like receptors (TLRs), thereby leading to AP-1-and NF-B-dependent transcription and resulting in the production of proinflammatory cytokines. Helicobacter pylori is a gram-negative bacterium that plays an etiologic role in the development of gastritis, peptic ulceration, and gastric adenocarcinoma (2). Several bacterial factors are proposed to play a role in disease pathogenesis. Type I H. pylori strains contain a pathogenicity island, which carries a number of virulence factors, including cagA and cagE (7), and is associated with more severe gastroduodenal disease (2). Studies using isogenic mutants demonstrate that certain genes carried on the cag pathogenicity island, including cagE but not cagA, are responsible for nuclear factor-B (NF-B) activation resulting in the transcription of a number of proinflammatory genes such as interleukin-8 (IL-8), IL-1␤, gamma interferon, and tumor necrosis factor alpha (20,29). However, the eukaryotic receptors involved in H. pylori activation of the innate immune response have not been clearly defined.Toll-like receptors (TLRs) play a crucial role in host innate and adaptive immune responses to microbial pathogens and their products (1). TLRs have leucine-rich motifs in their extracellular domains similar to those of other pattern-recognition proteins that promote ligand binding (1). TLR proteins also contain a cytoplasmic tail that is homologous to the IL-1 and IL-18 receptor and hence can trigger intracellular signaling pathways (23). To date, 10 TLRs have been described (31), with TLR2 and TLR4 the two best characterized. TLR2 responds to peptidoglycan, lipoteichoic acid (24), and bacterial lipoproteins (19). TLR4 is activated by the lipopolysaccharide (LPS) of gram-negative bacteria (3). Recently, it has been demonstrated that TLR2 and TLR4 are expressed on human intestinal epithelial cell lines (4, 5) and that Escherichia coli (O26:B6)-derived LPS induces TLR4 trafficking in epithelial cells (3). Maeda et al. (20) showed that TLR4 mRNA is also expressed on gastric epithelial MKN45 cells. In contrast to macrophages, TLR4 is not involved in H. pylori-induced NF-B activation in gastric epithelia (21). Therefore, the precise function(s) of TLR4 in gastric epithelial cells is still not known. MATERIALS AND METHODSReagents. H. pylori-derived LPS was kindly provided by Mario Monteiro (Institute for Biological Sciences, National Research Council, Ottawa, Ontario, Canada). Polyclonal anti-TLR4 and anti-actin were purchased from Santa Cruz Biotechnology (Santa Cruz, Calif.), and monoclonal anti-TLR4 (HTA125) antibody was purchased from eBioscience (San Diego, Calif.). Polyclonal H. pylori immune serum was purchased from DAKO (...