Helicobacter pylori colonization critically depends on postprandial gastric conditions.

The risk of Helicobacter pylori infection is highest in childhood, but the colonization process of the stomach mucosa is poorly understood. We used anesthetized Mongolian gerbils to study the initial stages of H. pylori colonization. Prandial and postprandial gastric conditions characteristic of humans of different ages were simulated. The fraction of bacteria that reached the deep mucus layer varied strongly with the modelled postprandial conditions. Colonization success was weak with fast gastric reacidification typical of adults. The efficiency of deep mucus entry was also low with a slow pH decrease as seen in pH profiles simulating the situation in babies. Initial colonization was most efficient under conditions simulating the postprandial reacidification and pepsin activation profiles in young children. In conclusion, initial H. pylori colonization depends on age-related gastric physiology, providing evidence from an in vivo infection model that suggests an explanation why the bacterium is predominantly acquired in early childhood.

Specific therapeutic schemes of omeprazole affect the orientation of Helicobacter pylori.

Until now, it has been unclear how proton pump inhibitors (PPIs) support Helicobacter pylori therapy. We tested whether the PPI omeprazole acts on the spatial orientation of H. pylori in the gastric mucus of infected Mongolian gerbils. Following repetitive PPI administration once daily but not following single doses or administration every 8 h, the bacterial spatial distribution changed, indicating a loss of orientation. Therefore, the therapeutic scheme of PPI administration may affect efficiency of treatment.

Pathophysiological mechanisms of extraesophageal reflux in otolaryngeal disorders.

As more than 40% of adults experience symptoms of the gastroesophageal reflux (GER) and 26% are affected by the extraesophageal reflux (EER), the aim of this article was to review the literature concerning pathophysiological mechanisms contributing to these common diseases. Reflux symptoms are ascribable for nearly one-third of otolaryngeal disorders. In contrast to patients solely affected by GERD, patients with otolaryngeal disorders attributable to extraesophageal reflux have a relatively good esophageal acid clearance but for unknown reasons increased amounts of laryngeal reflux. Transient upper esophageal sphincter relaxations are discussed as the pathophysiological mechanism, as the resting tone of the upper esophageal sphincter is not affected. When exposed to gastroduodenal contents, the ciliated epithelium of otolaryngeal structures is more susceptible to damage, and thereby even a few reflux episodes are suggested to cause extraesophageal reflux disease (EERD). Particularly active pepsin contributes to laryngeal lesions and eustachian tube dysfunction. Despite the importance of EER in laryngeal diseases, the causative role in other otolaryngeal disorders like sinusitis and otitis media with effusion remains unresolved.

Functional characterization and mutagenesis of the proposed behavioral sensor TlpD of Helicobacter pylori.

Helicobacter pylori requires flagellar motility and chemotaxis to establish and maintain chronic infection of the human stomach. The pH gradient in the stomach mucus is essential for bacterial orientation and guides the bacterium toward a narrow layer of the mucus, suggesting that H. pylori is capable of energy sensing or taxis. In the present study, H. pylori wild-type behavior in a temporal swimming assay could be altered by electron transport inhibitors, indicating that a connection between metabolism and behavior exists. In order to elucidate mechanisms of behavioral responses of H. pylori related to energy sensing, we investigated the phenotypes of single and multiple mutants of the four proposed chemotaxis sensor proteins. All sensor mutants were motile, but they diverged in their behavior in media supporting different energy yields. One proposed intracellular sensor, TlpD, was crucial for behavioral responses of H. pylori in defined media which did not permit growth and led to reduced bacterial energy levels. Suboptimal energetic conditions and inhibition of electron transport induced an increased frequency of stops and direction changes in the wild type but not in tlpD mutants. Loss of metabolism-dependent behavior in tlpD mutants could be reversed by complementation but not by electron donors bypassing the activity of the electron transport chain, in contrast to the case for the wild type. TlpD, which apparently lacks transmembrane domains, was detected both in the bacterial cytoplasm and at the bacterial periphery. The proposed energy sensor TlpD was found to mediate a repellent tactic response away from conditions of reduced electron transport.

In situ measurement of pH in the secreting canaliculus of the gastric parietal cell and adjacent structures.

The gastric H(+)/K(+)-ATPase is located within an infolding (secretory canaliculus) of the apical plasma membrane of gastric parietal cells. Our aim was to measure the pH values in the cytosol and canaliculus of the acid-secreting parietal cell and the adjacent gland lumen in situ. We used ultrafine double-barreled tip-sealed microelectrodes at high acceleration rates for intracellular and canalicular measurements. Immunohistochemical staining of the parietal cells was used to identify the track of the electrode and to estimate the position of the electrode tip at the time of the last intracellular measurement. En route to the deepest regions of the mucosa, where the average gland lumen pH was approximately 3, and on advancing in steps of 2 mum, the electrode entered the cytosol of the parietal cells, where the pH value was 7.4. Advancing the electrode further resulted, in several instances, in a sharp decrease in pH to an average value of 1.7 +/- 0.2, which we interpreted as the measurement within the canaliculus. When the electrode was advanced even further, the pH reading returned to the cytosolic value. From the difference in pH between the secreting canaliculus and the adjacent gland lumen, we concluded that the released acid was immediately buffered. Thus, the only cellular structure directly exposed to the highly acidic canalicular content is the apical membrane forming the canaliculus in the parietal cell.

Tracing of gastric reflux into the middle ear in a mongolian gerbil model.

OBJECTIVES: The purpose of this study was to trace induced gastric reflux and to examine whether it reaches the middle ear in a Mongolian gerbil model. BACKGROUND: Otitis media with effusion is the most frequent middle ear disease in childhood. Gastroesophageal reflux is suspected to be a possible factor in its pathogenesis. METHODS: Seventeen Mongolian gerbils were assigned to three groups: the control (phosphate-buffered saline application to the lower esophageal sphincter) and two experimental groups (Aquo-Trinitrosan [Merck, Darmstadt, Germany] application to the lower esophageal sphincter, low gastric pressure and Aquo-Trinitrosan application, higher gastric pressure). We injected Chinese ink into the stomach to trace the path of a potential gastroesophageal reflux in all three groups. The traces of ink were investigated by ear and larynx endoscopy and histology. RESULTS: There were no signs of gastroesophageal reflux based on the data obtained from the control group. In animals with traceable laryngeal reflux, the ink was also shown to advance through the eustachian tube and reach the middle ear. In addition, we found that when reflux reaches the middle ear on one side, it also reaches the contralateral middle ear in most cases. CONCLUSION: Gastroesophageal reflux induced by relaxation of the lower esophageal sphincter was shown to reach the middle ear in our Mongolian gerbil model. These results support recent hypotheses linking gastroesophageal reflux to the development of otitis media with effusion.

Vaccination prevents Helicobacter pylori-induced alterations of the gastric flora in mice.

Molecular analysis of the gastric microflora in mice revealed that Helicobacter pylori infection causes an increase in microbial diversity. The stomachs of H. pylori-infected animals were colonized by bacteria which are naturally restricted to the lower intestinal tract. Clostridia, Bacteroides/Prevotella spp., Eubacterium spp., Ruminococcus spp., streptococci and Escherichia coli were detected exclusively in the stomachs of infected animals, whereas lactobacilli dominated the gastric flora in noninfected mice. The H. pylori-induced shifts in the gastric microbiota were independent from histological pathology and from changes in the gastric pH but were prevented by immunization of mice with live Salmonella expressing H. pylori urease. Immunized mice displayed reduced H. pylori levels in the gastric epithelium and developed a normal gastric microflora, indicating that vaccination may be protective against H. pylori-induced changes in the gastric flora.

Gastric antibacterial efficiency is different for pepsin A and C.

The gastric lumen represents a bactericidal barrier, whose major components are an acidic pH and a family of isoenzymes of the gastric aspartate protease, pepsin. To evaluate whether specific pepsins are specialized in antibacterial protection, we tested their effects on the gastric pathogen Helicobacter pylori. In a recent study we found pepsin to affect the motility of the bacteria, one of its most important virulence factors. We were able to show that the antibacterial effect of pepsin occurs in two phases: rapid loss of motility and subsequent destruction. In the present study we used the rapid pepsin-induced bacterial immobilization as a marker of antibacterial efficiency. The proteolytic activity of different pepsins was normalized to values between 2 and 200 U/ml in the hemoglobin degradation test of Anson, performed at pH 2 and 5. We found that pepsin C completely inactivates H. pylori at proteolytic activities of 2 (pH 5) and 20 (pH 2) U/ml. In contrast, the activities of pepsin A and chymosin required to affect Helicobacter motility were ten times higher.

Rapid loss of motility of Helicobacter pylori in the gastric lumen in vivo.

The human pathogen Helicobacter pylori has infected more than half of the world's population. Nevertheless, the first step of infection, the acute colonization of the gastric mucus, is poorly understood. For successful colonization, H. pylori must retain active motility in the gastric lumen until it reaches the safety of the mucus layer. To identify the factors determining the acute colonization, we inserted bacteria into the stomach of anesthetized Mongolian gerbils. We adjusted the gastric juice to defined pH values of between 2.0 and 6.0 by using an autotitrator. Despite the fact that Helicobacter spp. are known to survive low pH values for a certain time in vitro, the length of time that H. pylori persisted under the assay conditions within the gastric juice in vivo was remarkably shorter. In the anesthetized animal we found H. pylori to be irreversibly immotile in less than 1 min at lumen pH values of 2 and 3. At pH 4 motility was lost after 2 min. However, the period of motility increased to more than 15 min at pH 6. Blocking pepsins in the gastric lumen in vivo by using pepstatin significantly increased the period of motility. It was possible to simulate the rapid in vivo immotilization in vitro by adding pepsins. We conclude that pepsin limits the persistence of H. pylori in the gastric chymus to only a few minutes by rapidly inhibiting active motility. It is therefore likely that this short period of resistance in the gastric lumen is one of the most critical phases of Helicobacter infection.

The spatial orientation of Helicobacter pylori in the gastric mucus.

The highly motile human pathogen Helicobacter pylori lives deep in the gastric mucus layer. To identify which chemical gradient guides the bacteria within the mucus layer, combinations of luminal perfusion, dialysis, and ventilation were used to modify or invert transmucus gradients in anaesthetized Helicobacter-infected mice and Mongolian gerbils. Neither changes in lumen or arterial pH nor inversion of bicarbonate/CO2 or urea/ammonium gradients disturbed Helicobacter orientation. However, elimination of the mucus pH gradient by simultaneous reduction of arterial pH and bicarbonate concentration perturbed orientation, causing the bacteria to spread over the entire mucus layer. H. pylori thus uses the gastric mucus pH gradient for chemotactic orientation.