The mucin O-glycosylation of 10 individuals with and without gastric disease was examined in depth in order to generate a structural map of human gastric glycosylation. In the stomach, these mucins and their O-glycosylation protect the epithelial surface from the acidic gastric juice and provide the first point of interaction for pathogens such as Helicobacter pylori, reported to cause gastritis, gastric and duodenal ulcers and gastric cancer. The rational of the present study was to map the O-glycosylation that the pathogen may come in contact with. An enormous diversity in glycosylation was found, which varied both between individuals and within mucins from a single individual: mucin glycan chain length ranged from 2-13 residues, each individual carried 34-103 O-glycan structures and in total over 258 structures were identified. The majority of gastric O-glycans were neutral and fucosylated. Blood group I antigens, as well as terminal α1,4-GlcNAc-like and GalNAcβ1-4GlcNAc-like (LacdiNAc-like), were common modifications of human gastric O-glycans. Furthemore, each individual carried 1-14 glycan structures that were unique for that individual. The diversity and alterations in gastric O-glycosylation broaden our understanding of the human gastric O-glycome and its implications for gastric cancer research and emphasize that the high individual variation makes it difficult to identify gastric cancer specific structures. However, despite the low number of individuals, we could verify a higher level of sialylation and sulfation on gastric O-glycans from cancerous tissue than from healthy stomachs.

h Helicobacter heilmannii naturally colonizes the stomachs of dogs and cats and has been associated with gastric disorders in humans. Nine feline Helicobacter strains, classified as H. heilmannii based on ureAB and 16S rRNA gene sequences, were divided into a highly virulent and a low-virulence group. The genomes of these strains were sequenced to investigate their phylogenetic relationships, to define their gene content and diversity, and to determine if the differences in pathogenicity were associated with the presence or absence of potential virulence genes. The capacities of these helicobacters to bind to the gastric mucosa were investigated as well. Our analyses revealed that the low-virulence strains do not belong to the species H. heilmannii but to a novel, closely related species for which we propose the name Helicobacter ailurogastricus. Several homologs of H. pylori virulence factors, such as IceA1, HrgA, and jhp0562-like glycosyltransferase, are present in H. heilmannii but absent in H. ailurogastricus. Both species contain a VacA-like autotransporter, for which the passenger domain is remarkably larger in H. ailurogastricus than in H. heilmannii. In addition, H. ailurogastricus shows clear differences in binding to the gastric mucosa compared to H. heilmannii. These findings highlight the low-virulence character of this novel Helicobacter species. Helicobacter pylori is considered one of the most successful human pathogens. Infection with this agent has been associated with a wide range of gastric disorders. However, H. pylori is not the only Helicobacter species causing gastric disease in humans. Helicobacter heilmannii (sensu stricto), a zoonotic bacterium naturally colonizing the stomachs of cats and dogs, has been associated with gastritis, peptic and duodenal ulcers, and mucosa-associated lymphoid tissue (MALT) lymphoma in humans (1-6). This Helicobacter species is highly prevalent in the stomachs of clinically healthy cats and dogs as well as in those of animals showing chronic active gastritis (1, 4). Its pathogenic significance in these animals remains unclear and is probably strain dependent or related to host differences (1).Little information is available regarding the pathogenesis of H. heilmannii infections in humans (1, 7). A recent experimental infection study, using Mongolian gerbils as an in vivo model to study Helicobacter-related gastric pathology in humans, investigated the colonization capacities and virulence of nine different Helicobacter strains (8). These helicobacters had been isolated from the gastric mucosae of stray cats and had been classified as H. heilmannii on the basis of the ureAB and 16S rRNA gene sequences (9). At 9 weeks postinfection, the induction of an antrum-dominant chronic active gastritis associated with the formation of lymphocytic aggregates and upregulation of the proinflammatory cytokine interleukin 1␤ (IL-1␤) was shown for seven strains. However, differences in the expression of IL-1␤ were noted, together with differences in the intensity of the obser...

Helicobacter suis colonizes the stomach of most pigs and is the most prevalent non-Helicobacter pylori Helicobacter species found in the human stomach. In the human host, H. suis contributes to the development of chronic gastritis, peptic ulcer disease and MALT lymphoma, whereas in pigs it is associated with gastritis, decreased growth and ulcers. Here, we demonstrate that the level of H. pylori and H. suis binding to human and pig gastric mucins varies between individuals with species dependent specificity. The binding optimum of H. pylori is at neutral pH whereas that of H. suis has an acidic pH optimum, and the mucins that H. pylori bind to are different than those that H. suis bind to. Mass spectrometric analysis of mucin O-glycans from the porcine mucin showed that individual variation in binding is reflected by a difference in glycosylation; of 109 oligosaccharide structures identified, only 14 were present in all examined samples. H. suis binding to mucins correlated with glycans containing sulfate, sialic acid and terminal galactose. Among the glycolipids present in pig stomach, binding to lactotetraosylceramide (Galβ3GlcNAcβ3Galβ4Glcβ1Cer) was identified, and adhesion to Galβ3GlcNAcβ3Galβ4Glc at both acidic and neutral pH was confirmed using other glycoconjugates. Together with that H. suis bound to DNA (used as a proxy for acidic charge), we conclude that H. suis has two binding modes: one to glycans terminating with Galβ3GlcNAc, and one to negatively charged structures. Identification of the glycan structures H. suis interacts with can contribute to development of therapeutic strategies alternative to antibiotics.

The mucin O-glycosylation of 10 individuals with and without gastric disease was examined in depth in order to generate a structural map of human gastric glycosylation. In the stomach, these mucins and their O-glycosylation protect the epithelial surface from the acidic gastric juice and provide the first point of interaction for pathogens such as Helicobacter pylori, reported to cause gastritis, gastric and duodenal ulcers and gastric cancer. The rational of the present study was to map the O-glycosylation that the pathogen may come in contact with. An enormous diversity in glycosylation was found, which varied both between individuals and within mucins from a single individual: mucin glycan chain length ranged from 2-13 residues, each individual carried 34 -103 O-glycan structures and in total over 258 structures were identified. The majority of gastric O-glycans were neutral and fucosylated. Blood group I antigens, as well as terminal ␣1,4-GlcNAc-like and GalNAc␤1-4GlcNAc-like (LacdiNAc-like), were common modifications of human gastric O-glycans. Gastric cancer is the second most common cause of cancer-associated death and fourth most commonly diagnosed cancer worldwide (1). Annually, 0.7 million patients with gastric cancer die globally (2). The cancer is associated with glycosylation changes, but how alteration of gastric mucins relates to gastric cancer pathogenesis remains unknown. Despite the protection by mucins and the acidic gastric juice and proteolytic enzymes, the bacterium Helicobacter pylori manage to thrive in the gastric lining, infecting about half of the world's population (3). There is a direct correlation between infection and gastric cancer, where 0.1-3% of infected individuals develop gastric adenocarcinoma or mucosa-associated lymphoid tissue lymphoma and another 10 -15% develop symptomatic gastritis or gastric and duodenal ulcers, whereas the majority show no symptoms (4).In the stomach, MUC5AC and MUC6 are the major secreted mucins, whereas MUC1 is the dominant membraneassociated mucin. MUC5AC is produced by the surface epithelium, whereas MUC6 is secreted from the deep glands of the gastric mucosa (5, 6). Both MUC5AC and MUC6 are large oligomeric mucins that occur as distinct glycoforms (7). In gastric precancerous lesions and cancer, altered expression of MUC5AC, MUC6, MUC2, and MUC5B has been described, with MUC2 being a marker for intestinal metaplasia (8, 9). The gastric surface and foveolar epithelium are formed by a single layer of tall columnar mucin-producing cells that have a basal nucleus below an apical cup of mucin. These cells have a turnover rate of 3-6 days, but the mucus layer produced in these cells have an even shorter life span: the production rate from start of glycosylation until release at the apical side is about 6 h (10), demonstrating that both the mucin repertoire and glycosylation theoretically can change rapidly. The carbohydrate structures present on mucosal surfaces vary according to cell lineage, tissue location, and developmental stage (11). The massive...

Gastrointestinal infections cause significant challenges and economic losses in animal husbandry. As pathogens becoming resistant to antibiotics are a growing concern worldwide, alternative strategies to treat infections in farmed animals are necessary in order to decrease the risk to human health and increase animal health and productivity. Mucosal surfaces are the most common route used by pathogens to enter the body. The mucosal surface that lines the gastrointestinal tract is covered by a continuously secreted mucus layer that protects the epithelial surface. The mucus layer is the first barrier the pathogen must overcome for successful colonization, and is mainly composed of densely glycosylated proteins called mucins. The vast array of carbohydrate structures present on the mucins provide an important setting for host-pathogen interactions. This review summarizes the current knowledge on gastrointestinal mucins and their role during infections in farmed animals. We examine the interactions between mucins and animal pathogens, with a focus on how pathogenic bacteria can modify the mucin environment in the gut, and how this in turn affects pathogen adhesion and growth. Finally, we discuss analytical challenges and complexities of the mucus-based defense, as well as its potential to control infections in farmed animals.