Bacterial Carbohydrate Research Articles

ChemInform Abstract: Carbohydrates and Immunology: Synthetic Oligosaccharide Antigens for Vaccine Formulation

AbstractReview: about 300 refs.

Malabsorption and Malnutrition in the Elderly - An Evolution?

Purpose: 80% of the elderly population presenting for hospital admission are either malnourished or potentially malnourished. In a similar cohort, potential malabsorption was identified using a Lactulose/Rhamnose study of small intestinal permeability. Increased permeability was present in 21%. When potential malabsorption begins has not been identified and we proposed to study (over 2 years) a group independently living to determine at what age this process begins. Methods: 154 active, independent living elderly were nutritionally assessed at 3 monthly intervals using the Nestle Mini Nutritional Assessment (MNA) and anthropometrics (hand grip, mid-arm muscle circumference (MAMC), tricep skin-fold thickness, body mass index (BMI), calf circumference). Carbohydrate absorption was measured at 6 monthly intervals with a Rice Breath Hydrogen Test (RBHT) to identify small intestinal bacterial overgrowth (SIBO) and carbohydrate (CHO) malabsorption. The test utilized 100 g of rice, hydrogen levels were measured at 20min intervals for 1 hour, then at 30min intervals for a further hour. A lactulose/rhamnose study of small intestine permeability was completed. Baseline blood tests identify any nutritional deficiencies or coeliac disease. Results: Average age was 80.8 years (60-90 years), ratio 3:1 females to males. At baseline 34.2% had abnormal RBHT's, 22.2% showing evidence of CHO Malabsorption (a RBHT with an increase more than 20 ppm). 12% showed evidence of SIBO (a RBHT with a basal level above 20 ppm). Nutritional assessment revealed 25.1% were malnourished or at risk at baseline which increased to 33.3% at 6 months. Calculated from 12 months data, mean MAMC for subjects with a normal RBHT was 25.7 cm, and 25.4 cm for subjects with an abnormal RBHT. At baseline the mean BMI for the normal RBHT group was 26.9, compared to 26.2 in the abnormal RBHT group. At 6 months the mean BMI for the normal RBHT group was 26.9, whereas the abnormal RBHT group mean BMI was 26.8. At 12 months the mean BMI had declined in both groups, normal RBHT group BMI was 26.3, and abnormal RBHT group BMI was 26.2. Blood tests and urine permeability tests have shown no abnormal results. Conclusion: Evaluations are not sufficient to identify onset of malnutrition. The data suggests that CHO malabsorption and SIBO is present in our independent living elderly with changes in nutritional status shown by MNA, BMI and MAMC. The sequence of events suggests SIBO is perhaps the first marker of potential malabsorption followed by rice CHO malabsorption. There has been no significant disturbance of mucosal permeability. We need to identify individual aspects in the genesis of this process - poor small intestinal motility followed by SIBO.

Host-adaptation of Francisella tularensis alters the bacterium's surface-carbohydrates to hinder effectors of innate and adaptive immunity.

BackgroundThe gram-negative bacterium Francisella tularensis survives in arthropods, fresh water amoeba, and mammals with both intracellular and extracellular phases and could reasonably be expected to express distinct phenotypes in these environments. The presence of a capsule on this bacterium has been controversial with some groups finding such a structure while other groups report that no capsule could be identified. Previously we reported in vitro culture conditions for this bacterium which, in contrast to typical methods, yielded a bacterial phenotype that mimics that of the bacterium's mammalian, extracellular phase.Methods/FindingsSDS-PAGE and carbohydrate analysis of differentially-cultivated F. tularensis LVS revealed that bacteria displaying the host-adapted phenotype produce both longer polymers of LPS O-antigen (OAg) and additional HMW carbohydrates/glycoproteins that are reduced/absent in non-host-adapted bacteria. Analysis of wildtype and OAg-mutant bacteria indicated that the induced changes in surface carbohydrates involved both OAg and non-OAg species. To assess the impact of these HMW carbohydrates on the access of outer membrane constituents to antibody we used differentially-cultivated bacteria in vitro to immunoprecipitate antibodies directed against outer membrane moieties. We observed that the surface-carbohydrates induced during host–adaptation shield many outer membrane antigens from binding by antibody. Similar assays with normal mouse serum indicate that the induced HMW carbohydrates also impede complement deposition. Using an in vitro macrophage infection assay, we find that the bacterial HMW carbohydrate impedes TLR2-dependent, pro-inflammatory cytokine production by macrophages. Lastly we show that upon host-adaptation, the human-virulent strain, F. tularensis SchuS4 also induces capsule production with the effect of reducing macrophage-activation and accelerating tularemia pathogenesis in mice.ConclusionF. tularensis undergoes host-adaptation which includes production of multiple capsular materials. These capsules impede recognition of bacterial outer membrane constituents by antibody, complement, and Toll-Like Receptor 2. These changes in the host-pathogen interface have profound implications for pathogenesis and vaccine development.

A novel polysaccharide secreted by pal/rim mutants of the phytopathogen fungus Ustilago maydis

Linear (1→3)-β-d-glucans and side-chain-branched β-d-glucans are major constituents of capsular materials, with roles in bacterial aggregation, virulence and carbohydrate storage; whereas branched (1→3),(1→6)-β-d-glucans constitute the most abundant components in the cell walls of fungi. In the present manuscript we describe the chemical characterization of a linear β-d-glucan secreted by mutants of the Basidiomycota biotrophic fungus Ustilago maydis affected in the Pal/Rim pathway involved in pH responses, that in contrast is not made by the wild type strain, at least in measurable amounts. The polysaccharide was obtained from the culture medium of a Δrim13::Cbx mutant of the fungus by ethanol precipitation, and analyzed using FT-IR spectroscopy, NMR spectroscopy in solid and liquid states, HPAEC-PAD chromatography, thin layer chromatography, enzymatic digestion and immunodetection. Our data revealed that, according to its characteristics, the polysaccharide is a (1→3)-β-d-glucan, and that the mutants used represent a relevant resource of this polysaccharide with wide applications.

Clinical and laboratory findings in 220 children with recurrent abdominal pain

To investigate the clinical and laboratory findings in children with recurrent abdominal pain (RAP). Consecutive patients with RAP (Apley criteria), age 4-16 years, referred to a secondary medical centre were evaluated by a standardized history, physical examination and laboratory tests. The tests encompassed Helicobacter pylori (Hp), gastrointestinal bacterial infections, protozoa, coeliac disease, carbohydrate malabsorption, food intolerance, abdominal ultrasound and plain abdominal X-ray. More investigations were obtained if indicated. Patient characteristics were compared with surgical patients without abdominal pain (control group). A total of 220 consecutive patients were included (92 M, mean age 8.8 years [4.1-16.0 years]). In 88% of the patients, abnormalities were found that refer to possible causes. Especially, protozoa were present in 33% of the patients, mostly Dientamoeba fragilis, Yersinia enterocolitica in 12% and endoscopically proven infection with Hp in 11%. In 36%, a plain abdominal X-ray raised suspicion of constipation. In 220 consecutive patients with RAP, referred to secondary care, a standardized work-up yielded abnormal results in a high percentage. The clinical significance of these findings remains to be established.

Bacterial Carbohydrate Structure Database 3: Principles and Realization

Bacterial carbohydrate structure database (BCSDB) is an open-access project that collects primary publication data on carbohydrate structures originating from bacteria, their biological properties, bibliographic and taxonomic annotations, NMR spectra, etc. Almost complete coverage and outstanding data consistency are achieved. BCSDB version 3 and the principles lying behind it, including glycan description language, are reported.

Enhancement of the antibacterial properties of silver nanoparticles using β-cyclodextrin as a capping agent

Silver nanoparticles (AgNPs) were synthesised by reducing silver salts using NaBH 4 followed by capping with varying concentrations of β-cyclodextrin (β-CD) and were physically characterised. Antibacterial activity against Escherichia coli, Pseudomonas aeruginosa and Staphylococcus aureus was determined by a microtitre well method. The AgNPs were spherical under transmission electron microscopy, whilst dynamic light scattering showed average diameters of capped particles to be smaller (4–7 nm) than their uncapped equivalents (17 nm). Capped particles demonstrated superior photostability when exposed to intense ultraviolet radiation for 4 h as well as significantly ( P < 0.05) higher (up to 3.5-fold) antibacterial activity. The influence of β-CD concentration was seen to delay bacterial growth, indicating that a Trojan horse mechanism may be occurring owing to bacterial carbohydrate affinity, thereby enhancing silver ion absorption.

In Vitro Kinetics of Prebiotic Inulin-Type Fructan Fermentation by Butyrate-Producing Colon Bacteria: Implementation of Online Gas Chromatography for Quantitative Analysis of Carbon Dioxide and Hydrogen Gas Production

Kinetic analyses of bacterial growth, carbohydrate consumption, and metabolite production of five butyrate-producing clostridial cluster XIVa colon bacteria grown on acetate plus fructose, oligofructose, inulin, or lactate were performed. A gas chromatography method was set up to assess H2 and CO2 production online and to ensure complete coverage of all metabolites produced. Method accuracy was confirmed through the calculation of electron and carbon recoveries. Fermentations with Anaerostipes caccae DSM 14662(T), Roseburia faecis DSM 16840(T), Roseburia hominis DSM 16839(T), and Roseburia intestinalis DSM 14610(T) revealed similar patterns of metabolite production with butyrate, CO2, and H2 as the main metabolites. R. faecis DSM 16840(T) and R. intestinalis DSM 14610(T) were able to degrade oligofructose, displaying a nonpreferential breakdown mechanism. Lactate consumption was only observed with A. caccae DSM 14662(T). Roseburia inulinivorans DSM 16841(T) was the only strain included in the present study that was able to grow on fructose, oligofructose, and inulin. The metabolites produced were lactate, butyrate, and CO2, without H2 production, indicating an energy metabolism distinct from that of other Roseburia species. Oligofructose degradation was nonpreferential. In a coculture of R. inulinivorans DSM 16841(T) with the highly competitive strain Bifidobacterium longum subsp. longum LMG 11047 on inulin, hardly any production of butyrate and CO2 was detected, indicating a lack of competitiveness of the butyrate producer. Complete recovery of metabolites during fermentations of clostridial cluster XIVa butyrate-producing colon bacteria allowed stoichiometric balancing of the metabolic pathway for butyrate production, including H2 formation.

High-Throughput Screening Identifies Novel Inhibitors of the Acetyltransferase Activity of Escherichia coli GlmU

The bifunctional GlmU protein catalyzes the formation of UDP-N-acetylglucosamine in a two-step reaction using the substrates glucosamine-1-phosphate, acetyl coenzyme A, and UTP. This metabolite is a common precursor to the synthesis of bacterial cell surface carbohydrate polymers, such as peptidoglycan, lipopolysaccharide, and wall teichoic acid that are involved in the maintenance of cell shape, permeability, and virulence. The C-terminal acetyltransferase domain of GlmU exhibits structural and mechanistic features unique to bacterial UDP-N-acetylglucosamine synthases, making it an excellent target for antibacterial design. In the work described here, we have developed an absorbance-based assay to screen diverse chemical libraries in high throughput for inhibitors to the acetyltransferase reaction of Escherichia coli GlmU. The primary screen of 50,000 drug-like small molecules identified 63 hits, 37 of which were specific to acetyltransferase activity of GlmU. Secondary screening and mode-of-inhibition studies identified potent inhibitors where compound binding within the acetyltransferase active site was requisite on the presence of glucosamine-1-phosphate and were competitive with the substrate acetyl coenzyme A. These molecules may represent novel chemical scaffolds for future antimicrobial drug discovery. In addition, this work outlines the utility of catalytic variants in targeting specific activities of bifunctional enzymes in high-throughput screens.

Changes in phenolic acids content, phenylalanine ammonia-lyase and peroxidase activities in sugarcane leaves induced by elicitors isolated fromXanthomonas albilineans

Xanthomonas albilineans is the agent responsible for a serious disease of sugarcane called leaf scald disease. Symptoms of the disease vary from a white line on the leaves to plant death. This paper correlates leaf scald resistance to changes in the concentration of phenolic compounds and phenylalanine ammonia-lyase and peroxidase activities in sugarcane leaves which were induced by a putative elicitor isolated from X. albilineans within different SE-chromatography fractions. The interaction was studied in leaf discs obtained from cultivars Louisiana 55-5 (susceptible to leaf scald) and Mayari 55-14 (resistant to leaf scald). SE-chromatography fractions containing only proteins enhanced the production of hydroxycinnamic acids (HCA) in cv. Mayari whereas fractions containing both proteins and carbohydrates enhanced the production of hydroxybenzoic acids (HBA) in cv. Louisiana. The eliciting protein(s) induced a level of PAL activity that increased the concentration of p-coumaric and caffeic acids, whereas ferulic acid, later synthesised in the host plant defence cycle, is used for the synthesis of syringic acid and for reinforcing the cell walls through peroxidase activation in the resistant cultivar. These facts justify the increase of p-coumaric, caffeic and syringic acids and the decrease of ferulic acid concentrations in this cultivar. Bacterial carbohydrates not only elicit the accumulation of p-coumaric acid, but also induce synthesis and accumulation of some hydroxybenzoic acids (HBA), such as gallic, protocatechuic and syringic acids and impede the production of antibacterial substances in the susceptible cultivar.

In Vitro Kinetic Analysis of Fermentation of Prebiotic Inulin-Type Fructans by Bifidobacterium Species Reveals Four Different Phenotypes

Kinetic analyses of bacterial growth, carbohydrate consumption, and metabolite production of 18 Bifidobacterium strains grown on fructose, oligofructose, or inulin were performed. A principal component analysis of the data sets, expanded with the results of a genetic screen concerning the presence of a beta-fructofuranosidase gene previously encountered in Bifidobacterium animalis subsp. lactis DSM 10140(T), revealed the existence of four clusters among the bifidobacteria tested. Strains belonging to a first cluster could not degrade oligofructose or inulin. Strains in a second cluster could degrade oligofructose, displaying a preferential breakdown mechanism, but did not grow on inulin. Fructose consumption was faster than oligofructose degradation. A third cluster was composed of strains that degraded all oligofructose fractions simultaneously and could partially break down inulin. Oligofructose degradation was substantially faster than fructose consumption. A fourth, smaller cluster consisted of strains that shared high fructose consumption and oligofructose degradation rates and were able to perform partial breakdown of inulin. For all strains, a metabolic shift toward more acetate, formate, and ethanol production, at the expense of lactate production, was observed during growth on less readily fermentable energy sources. No correlation between breakdown patterns and the presence of the beta-fructofuranosidase gene could be detected. These variations indicate niche-specific adaptation of bifidobacteria and could have in vivo implications on the strain specificity of the stimulatory effect of inulin-type fructans on bifidobacteria.

Symbiotic commensal bacteria direct maturation of the host immune system

Although commensal bacteria are known to play an important role in the proper maturation of the immune system of their mammalian hosts, the molecular mechanisms underlying this immunomodulation are poorly characterized. The present review summarizes recent findings in the field and describes new knowledge on the interplay of the innate and adaptive arms of the immune response induced by symbiotic bacterial carbohydrate antigens. Commensal bacteria in the intestine not only interact directly with dendritic cells but also engage in cross-talk with epithelial cells. These interactions lead to the induction of tolerogenic antigen-presenting cells in the lamina propria and ultimately to the regulation of functional maturation of effector T cells. Upon recognition of capsular polysaccharide antigens of commensal bacteria by dendritic cells (through toll-like receptor 2), innate immune responses facilitate and act in conjunction with adaptive responses to promote optimal Th1 polarization. In contrast, adaptive immunoglobulin A responses to symbiotic bacteria regulate the magnitude of oxidative innate immune responses in the mucosa as well as bacterial epitope expression in the lumen. Accumulating evidence is elucidating surface carbohydrate structures of symbiotic bacteria that drive the modulation of the intestinal immune system, resulting in mature, balanced immune responses and oral tolerance.

Bacterial production, glucosidase activity and particle-associated carbohydrates in Dona Paula bay, west coast of India

Size-fractionated bacterial production, abundance and α- and β- glucosidase enzyme activities were studied with respect to changes in hydrography, total suspended matter (TSM), chlorophyll a, particulate organic carbon and nitrogen ratio (POC:PON), 1.5 M NaCl-soluble and 10 mM EDTA-soluble carbohydrates (Sal-PCHO and CPCHO) and transparent exopolymeric particles (TEP) in the surface waters from July 1999–2000 at a shallow coastal station in Dona Paula Bay, west coast of India. The bulk of the total bacterial production and glucosidase activity were associated with particles (75% and >80%, respectively). Total bacterial production was linearly correlated to chlorophyll a ( r = 0.513; p < 0.05) whereas enzyme activity was significantly correlated to TSM (α-glucosidase: r = 0.721 ( p < 0.001); β-glucosidase: r = 0.596 ( p < 0.01)). Both α-glucosidase ( r = 0.514; p < 0.05) and β-glucosidase enzymes ( r = 0.598; p < 0.01) appeared to be involved in the degradation of CPCHO and Sal-PCHO, respectively. Changes in α-glucosidase/β-glucosidase ratios highlighted the varying composition of particulate organic matter. The bacterial uptake of 14C-labeled bacterial extracellular carbohydrate measured over 11 days showed a strong linear correlation between 14C-uptake and bacterial production using tritiated thymidine. The turnover rate of 14C-labeled carbohydrate-C was 0.52 d −1, higher than the estimated annual mean potential carbohydrate carbon turnover rate of 0.33 ± 0.2 d −1. Our study suggests that carbohydrates derived from sediments may serve as an important alternative carbon source sustaining the bacterial carbon demand in the surface waters of Dona Paula Bay.

GlycomeDB – integration of open-access carbohydrate structure databases

BackgroundAlthough carbohydrates are the third major class of biological macromolecules, after proteins and DNA, there is neither a comprehensive database for carbohydrate structures nor an established universal structure encoding scheme for computational purposes. Funding for further development of the Complex Carbohydrate Structure Database (CCSD or CarbBank) ceased in 1997, and since then several initiatives have developed independent databases with partially overlapping foci. For each database, different encoding schemes for residues and sequence topology were designed. Therefore, it is virtually impossible to obtain an overview of all deposited structures or to compare the contents of the various databases.ResultsWe have implemented procedures which download the structures contained in the seven major databases, e.g. GLYCOSCIENCES.de, the Consortium for Functional Glycomics (CFG), the Kyoto Encyclopedia of Genes and Genomes (KEGG) and the Bacterial Carbohydrate Structure Database (BCSDB). We have created a new database called GlycomeDB, containing all structures, their taxonomic annotations and references (IDs) for the original databases. More than 100000 datasets were imported, resulting in more than 33000 unique sequences now encoded in GlycomeDB using the universal format GlycoCT. Inconsistencies were found in all public databases, which were discussed and corrected in multiple feedback rounds with the responsible curators.ConclusionGlycomeDB is a new, publicly available database for carbohydrate sequences with a unified, all-encompassing structure encoding format and NCBI taxonomic referencing. The database is updated weekly and can be downloaded free of charge. The JAVA application GlycoUpdateDB is also available for establishing and updating a local installation of GlycomeDB. With the advent of GlycomeDB, the distributed islands of knowledge in glycomics are now bridged to form a single resource.

Hydrolysis of bacterial wall carbohydrates in the microwave using trifluoroacetic acid

By and large, monosaccharide composition and linkage analyses of bacterial cell-surface carbohydrates are achieved by hydrolysis into the corresponding monomeric constituents, and characterization of these, or their derivatives, by chromatographic and spectrometric methods. Normally, these hydrolyses are carried out conveniently with trifluoroacetic acid (TFA) at high temperatures for long periods of time, for example, in 4 M TFA at 100 °C for 5 h in a heating block. In this study, using a closed-vessel system, we investigated the effectiveness and reliability of microwave-assisted TFA hydrolysis of bacterial lipopolysaccharides, capsule, and teichoic-acid polysaccharides that were variably composed of several glycoses. In all cases, we were able to establish that 5 min of hydrolysis in the microwave at 120 °C with 4 M TFA (measured pressure of 90 psi) was sufficient time to obtain comparable results to those afforded by conventional hydrolysis. The same observation was made when fully methylated carbohydrates were hydrolyzed. The data obtained with our microwave system (Aurora Instruments MW600) showed that microwave-induced hydrolysis can be used with a high degree of confidence to carry out sugar composition analysis of complex bacterial glycans in markedly shorter periods of time. The results also suggested that non-thermal mechanistic factors must also be involved, at least in part, in accelerating the reaction rate of glycosidic hydrolysis.

Selective IgG subclass deficiency: quantification and clinical relevance

Each of the four human IgG subclasses exhibits a unique profile of effector functions relevant to the clearance and elimination of infecting microorganisms. The quantitative response within each IgG subclass varies with the nature of the antigen, its route of entry and, presumably, the form in which it is presented to the immune system. This results in antibody responses to certain antigens being predominantly or exclusively of a single IgG subclass. An inability to produce antibody of the optimally protective isotype can result in a selective immunodeficiency state. This is particularly apparent for responses to certain bacterial carbohydrate antigens that are normally of IgG2 isotype. A failure to produce the appropriate specific antibody response may result in recurrent upper and/or lower respiratory tract infection. Careful patient investigation can identify such deficiencies and suggest appropriate clinical management. In this review we outline the biology and clinical relevance of the IgG subclasses and summarize current rational treatment approaches.

Carbohydrate moieties as vaccine candidates: Meeting summary

In September 2007, a meeting entitled ‘Carbohydrate Moieties as Vaccine Candidates’ was held at the National Institutes of Health (Bethesda, MD). This meeting brought together scientists from a number of disciplines to address issues concerning carbohydrate moieties as targets for vaccines for a variety of pathogens and tumors. In addition, the meeting participants addressed fundamental topics of glycoimmunology including the recognition of glycotopes by B and T lymphocytes, the ontogeny of anti-carbohydrate immune responses, peptide mimicry, carbohydrate antigen processing pathways and adjuvants. One session reported progress in the development of new tools such as computational algorithms, glycan arrays and oligosaccharide synthesis and their application to carbohydrate vaccine research. The session titles were: (1) immune response to bacterial carbohydrate antigens; (2) immune response to glycolipids; (3) immune response to carbohydrate antigens on other microbes and on tumors; (4) novel vaccine approaches; (5) novel tools in carbohydrate vaccine research; (6) bench to bedside: carbohydrate moieties as vaccine immunopotentiators.

The Crystal Structure of Pectate Lyase PelI from Soft Rot Pathogen Erwinia chrysanthemi in Complex with Its Substrate

The crystallographic structure of the family 3 polysaccharide lyase (PL-3) PelI from Erwinia chrysanthemi has been solved to 1.45 A resolution. It consists of an N-terminal domain harboring a fibronectin type III fold linked to a catalytic domain displaying a parallel beta-helix topology. The N-terminal domain is located away from the active site and is not involved in the catalytic process. After secretion in planta, the two domains are separated by E. chrysanthemi proteases. This event turns on the hypersensitive response of the host. The structure of the single catalytic domain determined to 2.1 A resolution shows that the domain separation unveils a "Velcro"-like motif of asparagines, which might be recognized by a plant receptor. The structure of PelI in complex with its substrate, a tetragalacturonate, has been solved to 2.3 A resolution. The sugar binds from subsites -2 to +2 in one monomer of the asymmetric unit, although it lies on subsites -1 to +3 in the other. These two "Michaelis complexes" have never been observed simultaneously before and are consistent with the dual mode of bond cleavage in this substrate. The bound sugar adopts a mixed 2(1) and 3(1) helical conformation similar to that reported in inactive mutants from families PL-1 and PL-10. However, our study suggests that the catalytic base in PelI is not a conventional arginine but a lysine as proposed in family PL-9.

Interaction between seroreactivity to microbial antigens and genetics in Crohn’s disease: is there a role for defensins?

Antibodies against different microbial epitopes are associated with disease phenotype, may be of diagnostic importance and may reflect a loss of tolerance in Crohn's disease (CD). Recently, an association was reported between the presence of these antibodies and mutations in pattern receptor genes. Our aim was to investigate whether mutations in various genes other than NOD2/CARD15 or TLR4 associated with CD (NOD1/CARD4, DLG5 and DEFB1) may influence the presence of antibodies against bacterial proteins and carbohydrates in a Hungarian cohort of CD patients. Three hundred and seventy-six well-characterized, unrelated, consecutive CD patients (male/female: 191/185, age at onset: 29.1 +/- 12.9 years, duration: 7.9 +/- 11.7 years) were investigated. Sera were assayed for anti-Omp, anti-Saccharomyces cerevisiae antibodies (ASCAs) immunoglobulin (Ig) A and IgG, and antibodies against a mannan epitope of S. cerevisiae (gASCA), laminaribioside (ALCA), chitobioside (ACCA), and mannobioside (AMCA). NOD1/CARD4, DLG5 and DEFB1 variants were tested by polymerase chain reaction-restriction fragment length polymorphism, and DEFB1 was genotyped in a subgroup of 160 patients. Detailed clinical phenotypes were determined by reviewing the patients' medical charts. The carriage of DEFB1 20A variant alleles less frequently led to antiglycan positivity compared with patients without (29.6% vs 46.2%, OR: 0.49, 95% CI: 0.25-0.97), regardless of disease location or behavior. Similar tendency was observed for DEFB1 44G (present: 21.6% vs absent: 10.2%, P = 0.06) and ALCA. A gene or serology dosage effect was not observed. However, no association was found between the DEFB1 G52A, DLG5 R30Q, and NOD1/CARD4 E266K variants and any of the serology markers. We found that variants in human beta-defensin 1 gene are inversely associated with antiglycan antibodies, further confirming an important role for innate immunity in the pathogenesis of CD.

M2079 Interaction Between Seroreactivity to Microbial Antigens and Genetics in Crohn's Disease: Is There a Role for Defensins?

Antibodies against different microbial epitopes are associated with disease phenotype, may be of diagnostic importance and may reflect a loss of tolerance in Crohn's disease (CD). Recently, an association was reported between the presence of these antibodies and mutations in pattern receptor genes. Our aim was to investigate whether mutations in various genes other than NOD2/CARD15 or TLR4 associated with CD (NOD1/CARD4, DLG5 and DEFB1) may influence the presence of antibodies against bacterial proteins and carbohydrates in a Hungarian cohort of CD patients. Three hundred and seventy-six well-characterized, unrelated, consecutive CD patients (male/female: 191/185, age at onset: 29.1 +/- 12.9 years, duration: 7.9 +/- 11.7 years) were investigated. Sera were assayed for anti-Omp, anti-Saccharomyces cerevisiae antibodies (ASCAs) immunoglobulin (Ig) A and IgG, and antibodies against a mannan epitope of S. cerevisiae (gASCA), laminaribioside (ALCA), chitobioside (ACCA), and mannobioside (AMCA). NOD1/CARD4, DLG5 and DEFB1 variants were tested by polymerase chain reaction-restriction fragment length polymorphism, and DEFB1 was genotyped in a subgroup of 160 patients. Detailed clinical phenotypes were determined by reviewing the patients' medical charts. The carriage of DEFB1 20A variant alleles less frequently led to antiglycan positivity compared with patients without (29.6% vs 46.2%, OR: 0.49, 95% CI: 0.25-0.97), regardless of disease location or behavior. Similar tendency was observed for DEFB1 44G (present: 21.6% vs absent: 10.2%, P = 0.06) and ALCA. A gene or serology dosage effect was not observed. However, no association was found between the DEFB1 G52A, DLG5 R30Q, and NOD1/CARD4 E266K variants and any of the serology markers. We found that variants in human beta-defensin 1 gene are inversely associated with antiglycan antibodies, further confirming an important role for innate immunity in the pathogenesis of CD.