Sequential Statistical Improvement of the Liquid Cultivation of Helicobacter pylori

Background: Colonization of the gastric mucosa by Helicobacter pylori is one of the most important causes of acute and chronic gastric pathologies in humans. Achieving the growth of H. pylori in liquid media is of great importance in the development of clinical studies. In this study, we developed a sequential optimization strategy based on statistical models to improve the conditions of liquid culture of H. pylori. Materials and Methods: Four statistical models were sequentially used. First, a Box‐Behnken design was used to select the best process conditions (shaking speed, inoculum concentration, and final volume of culture). Secondly, a general factorial design was used to evaluate the influence of adding gel blocks or gel beads (shape and composition). Then a D‐optimal reduce design was carried out to allow the selection of the most influential factors in increasing the cell concentration (culture media components). Finally, another Box‐Behnken design was used to optimize the concentration of the culture media components previously selected. Results: After 12 hours of liquid culture a concentration of 25 × 10⁸ cells per mL (9.4 log₁₀ cells per mL) of H. pylori was obtained, compared with a predicted 32 × 10⁸ (9.5 log₁₀ cells per mL), which means between 1 and 5 log₁₀ units higher than some previous reports. Conclusions: The sequential statistical approach increased the planktonic H. pylori cell culture. The final culture media and conditions were: Brain Heart Infusion, blood agarose (1.5% w/v), lamb’s blood (3.18% v/v), DENT (0.11% v/v), and Vitox (0.52% v/v) at 60 rpm and 37 °C with filtered CO₂ (5% v/v) bubbled directly into the culture media in a final volume of 76.22 mL.     

The use of AlbuMAX II(®) as a blood or serum alternative for the culture of Helicobacter pylori

Background: Growth of Helicobacter pyloriin vitro depends on supplementation of the medium with blood or serum. However, these supplements often require frozen storage and can show batch‐to‐batch variation, resulting in differences in bacterial growth. In this study, we introduce the use of a commercially available, lipid‐rich supplement called AlbuMAX II^® (Gibco BRL, Grand Island, NY, USA) for use as a serum/blood replacement for H. pylori culture. Materials and Methods: The growth of H. pylori on solid and liquid media was examined by comparing growth after supplementation with horse blood, fetal calf serum, β‐cyclodextrin or AlbuMAX II^® (Gibco BRL). Human gastric adenocarcinoma (AGS) cellular responses to H. pylori were measured by NF‐κB luciferase assays and IL‐8 ELISA. Results: We show that the growth of H. pylori on both solid and liquid media containing AlbuMAX II^® (Gibco BRL) were comparable to levels obtained on blood agar or liquid media supplemented with serum. Growth was consistently higher in media supplemented with AlbuMAX II^® (Gibco BRL) than media containing β‐cyclodextrin. Furthermore, bacteria grown in AlbuMAX II^® (Gibco BRL) induced proinflammatory responses in AGS cells. Conclusions: AlbuMAX II^® (Gibco BRL) can be used as a serum/blood replacement for the cultivation of H. pylori in solid and liquid media. This medium could be useful for an improved understanding of H. pylori metabolism or for antigen production. Furthermore, AlbuMAX II^® (Gibco BRL) may be suitable for use in remote locations, particularly in areas where frozen storage of serum may be a problem.     

How many protein molecules are secreted by single Helicobacter pylori cells: Quantification of serine protease HtrA

Infection with Helicobacter pylori represents a major risk for developing peptic ulcer disease, gastric adenocarcinoma, and various other gastric and nongastric sicknesses. A series of H. pylori virulence factors can be secreted into the cell culture supernatant, and the secretome contains more than 100 different proteins. However, the quantities of proteins secreted by the bacteria over time are unknown. One of these factors is the serine protease high‐temperature requirement A (HtrA), encoded by an essential bifunctional gene with crucial intracellular and extracellular activities. We have demonstrated recently that secreted HtrA can cleave off the ectodomains of the tight junction proteins occludin and claudin‐8, as well as of the tumour suppressor and adherens junction protein E‐cadherin on polarised gastric epithelial cells. The exact mechanism of secretion and the quantity of secreted HtrA, however, have not been studied in detail. Here, we applied protein purification and quantitative Western blotting to determine the number of HtrA molecules secreted by H. pylori cells in liquid culture during a time course. Over a period of 8 hr, actively dividing bacteria secreted HtrA at a similar rate, on average about 9,600 HtrA molecules per cell. We determined minor variation over time corresponding to 9,931 ± 1,768 at an OD₆₀₀ of 0.4 after 2 hr, 9,403 ± 2,356 2 hr later, and 9,644 ± 2,067 molecules per cell after 8 hr of culturing, when the culture had reached an OD₆₀₀ of 0.8. This is the first report on the quantification of a secreted virulence protein from the important gastric pathogen H. pylori. Because HtrA has been considered as a promising new target for antibacterial therapy, knowledge about secreted protein quantities is crucial for optimising corresponding treatment regimes.     

Characterization and functional activity of murine monoclonal antibodies specific for α1,6-glucan chain of Helicobacter pylori lipopolysaccharide

Background: The outer core region of H. pylori lipopolysaccharide (LPS) contains α1,6‐glucan previously shown to contribute to colonizing efficiency of a mouse stomach. The aim of the present study was to generate monoclonal antibodies (mAbs) specific for α1,6‐glucan and characterize their binding properties and functional activity. Materials and Methods: BALB/c mice were injected intraperitoneally with 10⁸ formalin‐fixed H. pylori O:3 0826::Kan cells 3× over 56 days to achieve significant titer. Anti‐α1,6‐glucan‐producing hybridomas were screened by indirect ELISA using purified H. pylori O:3 0826::Kan LPS. One clone, 1C4F9, was selected for further characterization. The specificities of mAbs were determined by indirect and inhibition ELISA using structurally defined H. pylori LPS and synthetic oligosaccharides, and whole‐cell indirect ELISA (WCE) of clinical isolates. They were further characterized by indirect immunofluorescent (IF) microscopy and their functional activity in vitro determined by serum bactericidal assays against wild‐type and mutant strains of H. pylori. Results: The generated anti‐α1,6‐glucan IgM, 1C4F9, has demonstrated an excellent specificity for the glucan chain containing 5 to 6 α1,6‐linked glucose residues and showed surface accessibility by IF microscopy with H. pylori cells adherent to gastric adenocarcinoma cells monolayers. Of 38 isolates from Chile, 17 strains reacted with antiglucan mAbs in WCE (OD₄₅₀ ≥ 0.2). Bactericidal activity was observed against selective wild‐type and mutant H. pylori strains exhibiting OD₄₅₀ values of ≥0.45 in WCE. Conclusions: Anti‐α1,6‐glucan mAbs could have potential application in typing and surveillance of H. pylori isolates as well as offer insights into structural requirements for the development of LPS‐based vaccine against H. pylori infections.     

Significance of the caspase family in Helicobacter pylori induced gastric epithelial apoptosis

Background and Aims. H. pylori infection results in an increased epithelial apoptosis in gastritis and duodenal ulcer patients. We investigated the role and type of activation of caspases in H. pylori‐induced apoptosis in gastric epithelial cells. Methods. Differentiated human gastric cancer cells (AGS) and human gastric mucous cell primary cultures were incubated with H. pylori for 0.5–24 hours in RPMI 1640 medium, and the effects on cell viability, epithelial apoptosis, and activity of caspases were monitored. Apoptosis was analyzed by detection of DNA‐fragments by Hoechst stain®, DNA‐laddering, and Histone‐ELISA. Activities of caspases were determined in fluorogenic assays and by Western blotting. Cleavage of BID and release of cytochrome c were analyzed by Western blot. Significance of caspase activation was investigated by preincubation of gastric epithelial cells with cell permeable specific caspase inhibitors. Results. Incubation of gastric epithelial cells with H. pylori caused a time and concentration dependent induction of DNA fragmentation (3‐fold increase), cleavage of BID, release of cytochrome c and a concomittant sequential activation of caspase‐9 (4‐fold), caspase‐8 (2‐fold), caspase‐6 (2‐fold), and caspase‐3 (6‐fold). No effects on caspase‐1 and ‐7 were observed. Activation of caspases preceded the induction of DNA fragmentation. Apoptosis could be inhibited by prior incubation with the inhibitors of caspase‐3, ‐8, and ‐9, but not with that of caspase‐1. Conclusions. Activation of certain caspases and activation of the mitochondrial apoptotic pathway are essential for H. pylori induced apoptosis in gastric epithelial cells.     

In vitro and in vivo effects of the Mongolian drug Amu‐ru 7 on Helicobacter pylori growth and viability

Amu‐ru 7, a Mongolian folk medicine, is used to treat digestive diseases such as gastritis and gastric and duodenal ulcers. We examined the effect of Amu‐ru 7 on the growth and viability of Helicobacter pylori in vivo and in vitro. By the agar dilution method, the MIC of Amu‐ru 7 for H. pylori strains was shown to be 100–200 μg/mL with a MIC₉₀ of 200 μg/mL. Two hundred micrograms per milliliter of Amu‐ru 7 exhibited potent bactericidal activity against H. pylori in the stationary phase of growth 6 hr after treatment. Amu‐ru 7 inhibited the growth of both AMPC‐resistant and CAM‐resistant strains, and also had a combined effect with AMPC on AMPC‐resistant strain 403. The Amu‐ru 7 inhibited biofilm formation by H. pylori and induced morphological changes, such as bleb‐like formation and shortening of the cell. Although colonization of the stomach of the Mongolian gerbil by H. pylori was not cured by treatment with Amu‐ru 7, both the mean number of H. pylori colonized and the colonization rate were decreased in Amu‐ru 7 treated gerbils. These results suggest the effectiveness Amu‐ru 7 as an adjunct therapy for eradication therapies consisting of a PPI combined with antibiotics.     

The bifidogenic growth stimulator inhibits the growth and respiration of Helicobacter pylori

Background: Triple therapy with amoxicillin, clarithromycin, and a proton‐pump inhibitor is a common therapeutic strategy for the eradication of Helicobacter pylori (H. pylori). However, frequent appearance of clarithromycin‐resistant strains is a therapeutic challenge. While various quinones are known to specifically inhibit the growth of H. pylori, the quinone 1,4‐dihydroxy‐2‐naphthoic acid (DHNA) produced by Propionibacterium has strong stimulating effect on Bifidobacterium. We were interested to see whether DHNA could inhibit the growth of H. pylori in in vitro or in vivo experimental setting. Materials and Methods: The minimum inhibitory concentration (MIC) of DHNA was determined by the agar dilution method. The inhibitory action of DHNA on the respiratory activity was measured by using an oxygen electrode. Germ‐free mice infected with H. pylori were given DHNA in free drinking water containing 100 μg/mL for 7 days. Results: DHNA inhibited H. pylori growth at low MIC values, 1.6–3.2 μg/mL. Likewise, DHNA inhibited clinical isolates of H. pylori, resistant to clarithromycin. However, DHNA did not inhibit other Gram negative or anaerobic bacteria in the normal flora of the human intestine. Both H. pylori cellular respiration and adenosine 5′‐triphosphate (ATP) generation were dose‐dependently inhibited by DHNA. Similarly, the culture filtrates of propionibacterial strains inhibited the growth of H. pylori, and oral administration of DHNA could eradicate H. pylori in the infected germ‐free mice. Conclusions: The bifidogenic growth stimulator DHNA specifically inhibited the growth of H. pylori including clarithromycin‐resistant strains in vitro and its colonization activity in vivo. The bactericidal activity of DHNA was via inhibition of cellular respiration. These actions of DHNA may have clinical relevance in the eradication of H. pylori.     

Brugia malayi and B. pahangi: Cultivation in vitro of infective larvae to the fourth and fifth stages

Cultivation of fourth stage Brugia pahangi and B. malayi larvae from infective larvae (stage 3) were obtained in culture medium RPMI 1640 supplemented with 10% human AB serum and an LCC-MK₂ rhesus monkey kidney continuous cell line feeder layer. This culture system kept larvae alive in excess of 7 weeks, and served as a source for collection of the worms' secretory, excretory, and moulting antigens.     

Effects of cholesterol on Helicobacter pylori growth and virulence properties in vitro

Background: Colonization of the gastric mucosa by Helicobacter pylori is often associated with chronic gastric pathologies in humans. Development of disease correlates with the presence of distinct bacterial pathogenicity factors, such as the cag type IV secretion system (cag‐T4SS), the vacuolating cytotoxin (VacA), or the ability of the bacteria to acquire and incorporate cholesterol from human tissue. Materials and Methods: The in vitro growth of H. pylori requires media (Brucella broth) complemented with vitamins and horse serum or cyclodextrins, prepared as blood agar plates or liquid cultures. Liquid cultures usually show a slow growth. Here, we describe the successful growth of H. pylori strains 26695, P217, P12, and 60190 on serum‐free media replacing serum components or cyclodextrins with a commercially available cholesterol solution. Results: The effects of cholesterol as a substitute for serum or cyclodextrin were rigorously tested for growth of H. pylori on agar plates in vitro, for its general effects on bacterial protein synthesis (the proteome level), for H. pylori’s natural competence and plasmid DNA transfer, for the production of VacA, and the general function of the cag‐pathogenicity island and its encoded cag‐T4SS. Generally, growth of H. pylori with cholesterol instead of serum supplementation did not reveal any restrictions in the physiology and functionality of the bacteria except for strain 26695 showing a reduced growth on cholesterol media, whereas strain 60190 grew more efficient in cholesterol‐ versus serum‐supplemented liquid medium. Conclusions: The use of cholesterol represents a considerable option to serum complementation of growth media for in vitro growth of H. pylori.     

Cyclodextrins for growth ofHelicobacter pylori and production of vacuolating cytotoxin

Growth ofHelicobacter pylori in liquid culture requires the addition of media supplements that often interfere with subsequent purification of bacterial antigens. In order to determine whether cyclodextrins can substitute for conventionalH. pylori growth supplements, we culturedH. pylori in the presence of five commercially available cyclodextrins. The effect of these compounds on the production of the vacuolating cytotoxin antigen was evaluated. Several cyclodextrins supported flourishing growth and permitted the consistent production of vacuolating cytotoxin. These data suggest that Brucella broth supplemented with cyclodextrins is an improved medium for bacterial culture and industrial production ofH. pylori antigens.     

Nutritional Requirements of Plasmodium falciparum in Culture. I. Exogenously Supplied Dialyzable Components Necessary for Continuous Growth

Continuous cultivation of Plasmodium falciparum presently requires the nutritionally complex medium, RPMI 1640. A basal medium of KCl, NaCl, Na₂HPO₄, Ca(NO₃)₂, MgSO₄, glucose, reduced glutathione, HEPES buffer, hypoxanthine, phenol red (in RPMI 1640 concentrations), and 10% (v/v) exhaustively dialyzed pooled human serum was used to determine which vitamins and amino acids had to be exogenously supplied for continuous cultivation. Supplementation of basal medium with calcium pantothenate, cystine, glutamate, glutamine, isoleucine, methionine, proline, and tyrosine was necessary for continuous growth. This semi-defined minimal medium supported continuous growth of four isolates of P. falciparum at rates slightly less than those obtained with RPMI 1640. Adding any other vitamin or amino acid did not improve growth. Incorporation of several non-essential amino acids, particularly phenylalanine and leucine, into proteins was markedly enhanced in the minimal medium compared to RPMI 1640.     

Growth Medium Containing Cyclodextrin and Low Concentration of Horse Serum for Cultivation of Helicobacter pylori

The growth of Helicobacter pylori, a Gram-negative microaerophilic bacterium, is often difficult and requires complex media with the supplementation of 5% to 10% blood or blood derivatives. We have found that Brucella broth supplemented with 1% heated horse serum and 0.1% β-cyclodextrin supports the good growth of H. pylori. The degree of growth and production of urease and vacuolating cytotoxin in this medium were equal to those in the medium supplemented with 5% horse serum. This medium was found to be suitable for both the routine laboratory culture and primary isolation of H. pylori from biopsy samples.     

Long-term in vitro cultivation of Plasmodium berghei

Long-term in vitro culture of Plasmodium berghei was established using the Petri dish candle jar method of Trager and Jensen (1976). Cultures were established at 22, 27 and 37°C. As optimal growth was observed at 27°C, subsequent cultivation was carried out at this temperature. RPMI 1640 medium was modified by incorporating additional glucose (1 mg ml⁻¹) and bactopeptone (1 mg ml⁻¹) in the medium. This medium was found suitable for maintenance of mouse erythrocytes in vitro. P. berghei cultures were maintained using candle jars and this modified RPMI 1640 medium for 45 weeks.     

Simultaneous identification of T and B cells in blood smears using antibody coated bacteria.

Rabbit lymph nodes produce in vitro more antibodies over longer periods if the culture medium is supplemented with a mixture of horse and fetal bovine serum rather than rabbit serum. Medium RPMI 1640 sustains longer antibody responses and promotes a greater expansion of the cultures than medium 199, both media being supplemented with the serum mixture. Propagation of fast-growing, antibody-producing cultures fed with RPMI 1640 was accomplished by interculture transfer of lymphoid tissue. Round, mostly nonadherent cells grow in RPMI 1640 and medium 199 but in the former a striking proliferation of fibroblast-like cells takes place. This proliferation does not occur in medium 199. Preliminary results indicate that although the fibroblast-like cells do not produce antibodies, they contribute to the response in vitro.     

Up-regulation of inducible nitric oxide synthase and nitric oxide in Helicobacter pylori-infected human gastric epithelial cells: possible role of interferon-gamma in polarized nitric oxide secretion

Background. Nitric oxide (NO) generated by nitric oxide synthase (NOS) is known to be an important modulator of the mucosal inflammatory response. In this study, we questioned whether Helicobacter pylori infection could up‐regulate the epithelial cell inducible NOS (iNOS) gene expression and whether NO production could show polarity that can be regulated by immune mediators. Materials and Methods. Human gastric epithelial cell lines were infected with H. pylori, and the iNOS mRNA expression was assessed by quantitative RT‐PCR. NO production was assayed by determining nitrite/nitrate levels in culture supernatants. To determine the polarity of NO secretion by the H. pylori‐infected epithelial cells, Caco‐2 cells were cultured as polarized monolayers in transwell chambers, and NO production was measured. Results. iNOS mRNA levels were significantly up‐regulated in the cells infected with H. pylori, and expression of iNOS protein was confirmed by Western blot analysis. Increased NO production in the gastric epithelial cells was seen as early as 18 hours postinfection, and reached maximal levels by 24 hours postinfection. The specific MAP kinase inhibitors decreased H. pylori‐induced iNOS and NO up‐regulation. After H. pylori infection of polarized epithelial cells, NO was released predominantly into the apical compartment, and IL‐8 was released predominantly into basolateral compartment. The addition of IFN‐γ to H. pylori‐infected polarized epithelial cells showed a synergistically higher apical and basolateral NO release. Conclusion. These results suggest that apical NO production mediated by MAP kinase in H. pylori‐infected gastric epithelial cells may influence the bacteria and basolateral production of NO and IL‐8 may play a role in the tissue inflammation.     

Age-related alterations in cell division and cell cycle kinetics in control and trimethyltin-treated lymphocytes of human individuals

Trimethyltin chloride induced age-related suppression of cell division and cell cycle kinetics in human peripheral blood lymphocytes cultured in RPMI 1640 culture medium supplemented with human AB serum, phytohemagglutinin and bromodeoxyuridine. A high frequency of M₁ (first metaphase) cells was seen in cultures treated with a high dose (C ₁ = 1.0 g per culture) and in lymphocytes from donors in the age range 40–70 years. The delay in cell division and cell cycle kinetics may indicate a longer duration in DNA synthesis induced by trimethyltin chloride in aged lymphocytes.     

Study of the role of iron in the anticryptococcal activity of human serum and fluconazole

Anticryptococcal activity of human serum and apotransferrin in RPMI 1640 was studied in vitro. The effects of varying concentrations of FeCl₃ on this activity was investigated. Possible synergy of serum and apotransferrin with fluconazole was also measured. The fungistatic activity of human serum, whether lyophilized, stored at 4 °C, fresh frozen or purchased from commercial sources vs. Cryptococcus neoformans was comparable. There was no significant loss of fungistatic activity after freezing and thawing the serum up to 10 times. The fungistatic activity of human serum was similar when tested in different tissue culture media with the exception of Medium 199. The addition of apotransferrin (2.0 or 0.2 mg/ml) to RPMI 1640 had an inhibitory effect on cryptococcal growth. This effect was reversed by 20 M of FeCl₃ at both apotransferrin concentrations. By contrast, addition of FeCl₃ to human serum and RPMI 1640 did not reverse inhibition of growth. Fluconazole synergized with the human serum preparations described, but not with pooled commercial serum, for fungicidal activity. Synergistic activity of fluconazole and human serum was not affected by the addition of FeCl₃. Apotransferrin did not show any synergistic fungicidal activity with fluconazole.     

Complex responses to culture conditions in Pseudomonas syringae pv. tomato DC3000 continuous cultures: The role of iron in cell growth and virulence factor induction

The growth of a model plant pathogen, Pseudomonas syringae pv. tomato DC3000, was investigated using a chemostat culture system to examine environmentally regulated responses. Using minimal medium with iron as the limiting nutrient, four different types of responses were obtained in a customized continuous culture system: (1) stable steady state, (2) damped oscillation, (3) normal washout due to high dilution rates exceeding the maximum growth rate, and (4) washout at low dilution rates due to negative growth rates. The type of response was determined by a combination of initial cell mass and dilution rate. Stable steady states were obtained with dilution rates ranging from 0.059 to 0.086 h^?1 with an initial cell mass of less than 0.6 OD₆₀₀. Damped oscillations and negative growth rates are unusual observations for bacterial systems. We have observed these responses at values of initial cell mass of 0.9 OD₆₀₀ or higher, or at low dilution rates (<0.05 h^?1) irrespectively of initial cell mass. This response suggests complex dynamics including the possibility of multiple steady states. Iron, which was reported earlier as a growth limiting nutrient in a widely used minimal medium, enhances both growth and virulence factor induction in iron‐supplemented cultures compared to unsupplemented controls. Intracellular iron concentration is correlated to the early induction (6 h) of virulence factors in both batch and chemostat cultures. A reduction in aconitase activity (a TCA cycle enzyme) and ATP levels in iron‐limited chemostat cultures was observed compared to iron‐supplemented chemostat cultures, indicating that iron affects central metabolic pathways. We conclude that DC3000 cultures are particularly dependent on the environment and iron is likely a key nutrient in determining physiology. Biotechnol. Bioeng. 2010;105: 955–964. © 2009 Wiley Periodicals, Inc.     

A Potential Association between Helicobacter pylori CagA EPIYA and Multimerization Motifs with Cytokeratin 18 Cleavage Rate during Early Apoptosis

Background and Aims: Helicobacter pylori is a highly diverse pathogen, which encounters epithelial cells as the initial defense barrier during its lifelong infection. The structure of epithelial cells can be disrupted through cleavage of microfilaments. Cytokeratin 18 (CK18) is an intermediate filament, the cleavage of which is considered an early event during apoptosis following activation of effector caspases. Methods: Helicobacter pylori strains were isolated from 76 dyspeptic patients. cagA 3’ variable region and CagA protein status were analyzed by PCR and western blotting, respectively. Eight hours post‐co‐culture of AGS cells with different H. pylori strains, flow cytometric analysis was performed using M30 monoclonal antibody specific to CK18 cleavage‐induced neo‐epitope. Results: Higher rates of CK18 cleavage were detected during co‐culture of AGS cells with H. pylori strains bearing greater numbers of cagA EPIYA‐C and multimerization (CM) motifs. On the other hand, H. pylori strains with greater numbers of EPIYA‐B relative to EPIYA‐C demonstrated a decrease in CK18 cleavage rate. Thus, H. pylori‐mediated cleavage of CK18 appeared proportional to the number of CagA EPIYA‐C and CM motifs, which seemed to be downplayed in the presence of EPIYA‐B motifs. Conclusions: Our observation associating the heterogeneity of cagA variants with the potential of H. pylori strains in the induction of CK18 cleavage as an early indication of apoptosis in gastric epithelial cells supports the fact that apoptosis may be a type‐specific trait. However, additional cagA‐targeted experiments are required to clearly identify the role of EPIYA and CM motifs in apoptosis and/or the responsible effector molecules.     

Polaprezinc attenuates Helicobacter pylori-associated gastritis in Mongolian gerbils

Background. The ammonia‐monochloramine system plays an important role in Helicobacter pylori‐associated gastric mucosal injury. Polaprezinc, a new antiulcer agent, has a scavenging action against monochloramine. The aim of the experiment was to investigate the inhibitory effects of polaprezinc on the H. pylori‐induced gastritis in Mongolian gerbils. Materials and Methods. Mongolian gerbils fasting for 24 hours were orally given culture broth containing 2–4 × 10⁸ colony‐forming units of H. pylori ATCC 43054 per milliliter. From 4 hours after inoculation until the end of the experiment, gerbils were given chow pellets with or without 0.02% polaprezinc. All gerbils were killed 12 weeks later. The grades of H. pylori density and histologic features of gastritis were evaluated in accordance with the Updated Sydney System. The scavenging effect of polaprezinc on monochloramine was investigated spectrophotometrically. Results. Polaprezinc had little or no influence on the H. pylori density in both pyloric and fundic mucosae. However, it significantly attenuated the development of polymorphonuclear neutrophil activity, mononuclear infiltration, and surface epithelial erosion in both pyloric and fundic mucosae compared with those of the control group. H. pylori inoculation significantly increased the heights of both pyloric and fundic mucosae (mainly due to the increased height of foveolar hyperplasia), but polaprezinc inhibited the increase of mucosal thickness in both pyloric and fundic mucasae. No intestinal metaplasia was detected in this study. Spectrophotometric examination revealed that polaprezinc scavenged monochloramine. Conclusions. Polaprezinc inhibited the development of H. pylori‐induced gastritis through its scavenging action against monochloramine.