Variations in Helicobacter pylori lipopolysaccharide to evade the innate immune component surfactant protein D.

Helicobacter pylori is a common and persistent human pathogen of the gastric mucosa. Surfactant protein D (SP-D), a component of innate immunity, is expressed in the human gastric mucosa and is capable of aggregating H. pylori. Wide variation in the SP-D binding affinity to H. pylori has been observed in clinical isolates and laboratory-adapted strains. The aim of this study was to reveal potential mechanisms responsible for evading SP-D binding and establishing persistent infection. An escape variant, J178V, was generated in vitro, and the lipopolysaccharide (LPS) structure of the variant was compared to that of the parental strain, J178. The genetic basis for structural variation was explored by sequencing LPS biosynthesis genes. SP-D binding to clinical isolates was demonstrated by fluorescence-activated cell sorter analyses. Here, we show that H. pylori evades SP-D binding through phase variation in lipopolysaccharide. This phenomenon is linked to changes in the fucosylation of the O chain, which was concomitant with slipped-strand mispairing in a poly(C) tract of the fucosyltransferase A (fucT1) gene. SP-D binding organisms are predominant in mucus in vivo (P = 0.02), suggesting that SP-D facilitates physical elimination. Phase variation to evade SP-D contributes to the persistence of this common gastric pathogen.

Chemotactic response of Helicobacter pylori to human plasma and bile.

To clarify further the role of chemotaxis in Helicobacter pylori colonization, the in vitro bacterium response to human plasma and bile (secretions containing chemoeffector compounds that are present in the gastric mucus layer) was examined. Human plasma, after dilution to 1 % (v/v) with buffer, was found to be a chemoattractant for the motile bacillus. Human gall-bladder bile, after dilution to 2 % (v/v) with buffer, was found to be a chemorepellent, but did not cause the motility of the bacillus to be diminished after prolonged exposure. The basis of the chemoattractant effect of plasma was explored by examining how urea and 12 amino acids found in plasma affected the taxis of H. pylori. Urea and the amino acids histidine, glutamine, glycine and arginine were the strongest chemoattractants. Other amino acids were chemoattractants, with the exceptions of aspartic and glutamic acids, which were chemorepellents. The basis of the chemorepellent effect of bile was explored by examining how the six most abundant conjugated bile acids in human bile affected the taxis of H. pylori. All the bile acids were chemorepellents, with the greatest effects being demonstrated by taurocholic and taurodeoxycholic acids. The implications of these findings for H. pylori colonization of gastric epithelium are discussed.

How Helicobacter pylori urease may affect external pH and influence growth and motility in the mucus environment: evidence from in-vitro studies.

BACKGROUND: Survival of Helicobacter pylori is dependent upon urease in the cytoplasm and at the bacterial surface. We have sought to clarify how alkaline ammonium salts, released from urea by this enzyme, might alter mucus pH and so affect growth and motility of the bacterium in the gastric mucus environment. METHODS: Experiments were conducted in vitro to determine how the growth and motility of H. pylori are affected by changes in external pH, and how the bacterium, by hydrolysing urea, alters the pH of the bicarbonate buffer that occurs at the gastric mucosal surface. These data were fitted into experimental models that describe how pH varies within the mucus layer in the acid-secreting stomach. RESULTS: H. pylori was motile between pH 5 and 8, with optimal motility at pH 5. It grew between pH 6 and 8, with optimal growth at pH 6. The bacterium had urease activity between pH 2.7 and 7.4, as evidenced by pH rises in bicarbonate-buffered solutions of urea. Changes in buffer pH were dependent upon initial pH and urea concentration, with the greatest rate of pH change occurring at pH 3. Modelling experiments utilizing these data indicated that (1) in the absence of urease, H. pylori growth and motility in the mucus layer would be restricted severely by low mucus pH in the acid-secreting stomach, and (2) urease will sometimes inhibit H. pylori growth and motility in the mucus layer by elevating the pH of the mucus environment above pH 8. CONCLUSIONS: Urease is essential to the growth and motility of H. pylori in the mucus layer in the acid-secreting stomach, but, paradoxically, sometimes it might suppress colonization by raising the mucus pH above 8. This latter effect may protect the bacteria from the adverse consequences of overpopulation.

Expression of surfactant protein D in the human gastric mucosa and during Helicobacter pylori infection.

Helicobacter pylori establishes persistent infection of gastric mucosa with diverse clinical outcomes. The innate immune molecule surfactant protein D (SP-D) binds selectively to microorganisms, inducing aggregation and phagocytosis. In this study, we demonstrated the expression of SP-D in gastric mucosa by reverse transcription-PCR and immunohistochemical analysis. SP-D is present at the luminal surface and within the gastric pits, with maximal expression at the surface. Levels of expression are significantly increased in H. pylori-associated gastritis compared to those in the normal mucosa. Immunofluorescence microscopy was used to demonstrate binding and agglutination of H. pylori by SP-D in a lectin-specific manner. These activities resulted in a 50% reduction in the motility of H. pylori, as judged on the basis of curvilinear velocity measured by using a Hobson BacTracker. Lipopolysaccharides extracted from three H. pylori strains were shown to bind SP-D in a concentration-dependent manner, and there was marked variation in the avidity of binding among the strains. SP-D may therefore play a significant role in the innate immune response to H. pylori infection.