Cloning and Characterization of a 22 kDa Outer-Membrane Protein (Omp22) from Helicobacter pylori

Helicobacter pylori is a causative agent of gastritis and peptic ulceration in humans. As the first step towards development of a vaccine against H. pylori infection, we have attempted to identify protective antigens. A potential target of vaccine development would be a H. pylori specific protein, which is surface-exposed and highly antigenic. We identified a 22 kDa outer-membrane protein (Omp22) from H. pylori, which was highly immunoreactive. By screening a H. pylori genomic DNA library with rabbit anti-H. pylori outer-membrane protein antibodies, the omp22 gene was cloned and 1.4 kb of the nucleotide sequence was determined. One open reading frame, encoding a 179-residue polypeptide, was identified and the amino acid sequence deduced showed homology with peptidoglycan-associated lipoproteins. The sequence was conserved among other H. pylori strains. Omp22 protein is expressed as a precursor polypeptide of 179 residues and undergoes lipid modification and cleavage of an 18 amino acid signal peptide to yield a mature protein. Omp22 protein in H. pylori as well as recombinant Omp22 protein expressed in E. coli was localized into the outer membrane and exposed on the cell surface. Omp22 may have the potential as a target antigen for the development of a H. pylori vaccine.     

Helicobacter pylori chromosomal DNA replication: current status and future perspectives

Helicobacter pylori causes gastritis, gastric ulcer and gastric cancer. Though DNA replication and its control are central to bacterial proliferation, pathogenesis, virulence and/or dormancy, our knowledge of DNA synthesis in slow growing pathogenic bacteria like H. pylori is still preliminary. Here, we review the current understanding of DNA replication, replication restart and recombinational repair in H. pylori. Several differences have been identified between the H. pylori and Escherichia coli replication machineries including the absence of DnaC, the helicase loader usually conserved in gram-negative bacteria. These differences suggest different mechanisms of DNA replication at initiation and restart of stalled forks in H. pylori.     

Survival of Helicobacter pylori in well water

The mortality of a clinical Helicobacter pylori strain was assessed by inoculating it in untreated well water, filtered well water, and autoclaved well water. Two different temperatures (5 and 25 °C) were used during the experimental period. Because Escherichia coli is commonly used as indicator of faecal pollution of water, we compared the survival of H. pylori using E. coli as indicator of its persistence. H. pylori was not culturable 48 h after inoculation, whereas the population of E. coli, monitored at the same temperature, decreased slowly, especially in filtered water. In untreated water, both H. pylori and E. coli survived less well than in filtered and autoclaved water. In general the survival of H. pylori and E. coli was better in filtered water than in autoclaved water and the ability of H. pylori to survive several days in water at 5 °C is reported, supporting the observation that H. pylori survives better at 5 °C than at higher temperature. This suggests a possible faecal–oral transmission of H. pylori in the presence of a contaminated water.     

Anti-<Emphasis Type="Italic">Helicobacter pylori</Emphasis> substances from endophytic fungal cultures

The human pathogenic bacterium Helicobacter pylori has been ascertained to be an aetiological agent for chronic active gastritis and a significant determinant in peptic and duodenal ulcer diseases. Endophytic metabolites are being recognized as a versatile arsenal of antimicrobial agents, since some endophytes have been shown to possess superior biosynthetic capabilities owing to their presumable gene recombination with the host, while residing and reproducing inside the healthy plant tissues. A total of 32 endophytic fungi isolated from the medicinal herb Cynodon dactylon(Poaceae) were grown in in vitroculture, and the ethyl acetate extracts of the cultures were examined in vitro for the anti-H. pylori activity. As a result, a total of 16 endophyte culture extracts were identified as having potent anti-H. pyloriactivities. Subsequently, a detailed bioassay-guided fractionation of the extract of the most active endophyte (strain number: CY725) identified as Aspergillussp., was performed to afford eventually four anti-H. pylori secondary metabolites. The four isolated compounds were identified through a combination of spectral and chemical methods (IR, MS, ¹H- and ¹³C-NMR) to be helvolic acid, monomethylsulochrin, ergosterol and 3β-hydroxy-5α,8α-epidioxy- ergosta-6,22-diene with corresponding MICs of 8.0, 10.0, 20.0 and 30.0 μg/ml, respectively. The MIC of ampicillin co-assayed as a reference drug against H. pylori was 2.0 μg/ml. Furthermore, preliminary examination of the antimicrobial spectrum of helvolic acid, the most active anti-H. pylori metabolite characterized from the endophyte culture, showed that it was inhibitory to the growth of Sarcina lutea, Staphylococcus aureusand Candida albicans with MICs of 15.0, 20.0 and 30.0 μg/ml, respectively.     

幽门螺杆菌感染与全身各系统疾病的研究最新进展

随着近年来对幽门螺杆菌感染研究的不断深入,学者们发现幽门螺杆菌不仅是消化系统疾病的重要病因之一,还与血液、风湿免疫、神经以及其他甲状腺、妊娠高吐、眼部疾病等全身多系统疾病具有密切相关性。就近几年国内外对幽门螺杆菌与全身各系统疾病的相关性以及其机制的最新研究进展进行综述,为临床的诊断和治疗提供参考。     

Type I Restriction–Modification Loci Reveal High Allelic Diversity in Clinical Helicobacter pylori Isolates

Background: A remarkable variety of restriction‐modification (R‐M) systems is found in Helicobacter pylori. Since they encompass a large portion of the strain‐specific H. pylori genes and therefore contribute to genetic variability, they are suggested to have an impact on disease outcome. Type I R‐M systems comprise three different subunits and are the most complex of the three types of R‐M systems. Aims: We investigated the genetic diversity and distribution of type I R‐M systems in clinical isolates of H. pylori. Material and methods: Sixty‐one H. pylori isolates from a Swedish hospital based case‐control study and 6 H. pylori isolates of a Swedish population‐based study were analyzed using polymerase chain reaction for the presence of the three R‐M systems' subunits. Representative gene variants were sequenced. Results: Although the hsdM and hsdR genes appeared conserved in our clinical H. pylori isolates, the sequences of the hsdS loci were highly variable. Despite their sequence diversity, the genes per se were present at high frequencies. We identified a number of novel allelic hsdS variants, which are distinct from corresponding hsdS loci in the sequenced H. pylori strains 26695, J99 and HPAG1. In analyses of paired H. pylori isolates, obtained from the same individuals with a 4‐year interval, we observed genetic modifications of hsdS genes in patients with atrophic gastric mucosa. Discussion: We propose that the genetic variability of hsdS genes in a bacterial population will give rise to new specificities of these enzymes, which might lead to adaptation to an ever‐changing gastric environment.     

Helicobacter pylori infection promotes methylation of WWOX gene in human gastric cancer

WW domain-containing oxidoreductase (WWOX), a tumor suppressor gene, was reported to be downregulated in gastric cancer and other tumors. However, the mechanism by which WWOX is inactivated remains unclear. In our study, methylation status of WWOX was determined by MSP and sequencing. Our results showed that WWOX hypermethylation was frequently detected in gastric cancer, and also significantly correlated with Helicobacter pylori (H. pylori) infection. Promoter methylation of WWOX was induced in BCG823 and AGS cells co-cultured with H. pylori. Finally, we found that expression of DNMT1 and DNMT3A were enhanced when cells were co-cultured with H. pylori. Our study indicated that H. pylori infection promoted methylation of WWOX gene in gastric cancer.     

Adhesion ofHelicobacter pylori strains toα-2,3-linked sialic acids

Helicobacter pylori is a human pathogen associated with gastritis and peptic ulcer. Adhesion properties ofH. pylori to various structures have been described in the literature, including evidence for sialic acid-binding. To study the specificity and frequency of sialic acid-binding, fourteenH. pylori strains were investigated using haemagglutination with derivatized erythrocytes carrying sialic acids only on defined glycans and using haemagglutination inhibition assays. From these studiesH. pylori strains can be grouped into sialic acid-dependent and sialic acid-independent classes. The sialic acid-dependent strains require -2,3-linked sialic acid for haemagglutination. The potential roles of sialic acid-dependent adhesions forH. pylori-related infections are discussed.Abbreviations Sia sialic acids - Neu5Ac N-acetyl-neuraminic acid - Neu5Gc N-glycolylneuraminic acid - Neu5Fm N-formylneuraminic acid - Neu5TFA N-trifluoroacetylneuraminic acid - RBC human red blood cells (erythrocytes)     

Effects of dexamethasone and FK506 on Helicobacter pylori-induced gastritis and bacterial viability in Mongolian gerbils

FK506 and dexamethasone were used to investigate whether or not immunosuppression affects H. pylori colonization and gastric mucosal damage induced by Helicobacter pylori in Mongolian gerbils. Two weeks after H. pylori infection, FK506 and dexamethasone or vehicle alone were subcutaneously administered once daily for the following 2 weeks. FK506 or vehicle alone was administered subcutaneously once daily for 5 weeks (1 week before and 4 weeks after infection). In H. pylori-infected animals for 4 weeks, hemorrhagic erosions and inflammatory responses (neutrophil infiltration and lymphoid follicle formation) were induced in gastric mucosa at an incidence of 100%. Both FK506 and dexamethasone administered for 2 weeks markedly reduced such mucosal changes. In these animals, H. pylori viability in the stomach was significantly elevated. FK506 administered for 5 weeks also significantly inhibited the hemorrhagic erosions, edema and neutrophil infiltration in the stomach. H. pylori viability was slightly elevated as compared with the control. It was concluded that the host immune responses might play dual roles both by deteriorating gastritis induced by H. pylori and by protecting against H. pylori infection in its early stage.     

Synthesis, molecular docking and biological evaluation of metronidazole derivatives as potent Helicobacter pylori urease inhibitors

Fourteen metronidazole derivatives (compounds 3a–f and 4b–h) have been synthesized by coupling of metronidazole and salicylic acid derivatives. All of them are reported for the first time. Their chemical structures are characterized by ¹H NMR, MS, and elemental analysis. The inhibitory activities against Helicobacter pylori urease have been investigated in vitro and many compounds have showed promising potential inhibitory activities of H. pylori urease. The effect of compounds 4b (IC₅₀ = 26 μM) and 4g (IC₅₀ = 12 μM) was comparable with that of acetohydroxamic acid, a well known H. pylori urease inhibitor used as a positive control. The experimental values of IC₅₀ showed that inhibitor was potent urease inhibitor. A docking analysis using the autodock 4.0 program could explain the inhibitory activities of compound 4g against H. pylori urease.     

Protein Evolutionary Rates Correlate with Expression Independently of Synonymous Substitutions in Helicobacter pylori

In free-living microorganisms, such as Escherichia coli and Saccharomyces cerevisiae, both synonymous and nonsynonymous substitution frequencies correlate with expression levels. Here, we have tested the hypothesis that the correlation between amino acid substitution rates and expression is a by-product of selection for codon bias and translational efficiency in highly expressed genes. To this end, we have examined the correlation between protein evolutionary rates and expression in the human gastric pathogen Helicobacter pylori, where the absence of selection on synonymous sites enables the two types of substitutions to be uncoupled. The results revealed a statistically significant negative correlation between expression levels and nonsynonymous substitutions in both H. pylori and E. coli. We also found that neighboring genes located on the same, but not on opposite strands, evolve at significantly more similar rates than random gene pairs, as expected by co-expression of genes located in the same operon. However, the two species differ in that synonymous substitutions show a strand-specific pattern in E. coli, whereas the weak similarity in synonymous substitutions for neighbors in H. pylori is independent of gene orientation. These results suggest a direct influence of expression levels on nonsynonymous substitution frequencies independent of codon bias and selective constraints on synonymous sites. Electronic Supplementary Material Electronic Supplementary material is available for this article at and accessible for authorised users. [Reviewing Editor: Dr. Nicolas Galtier]     

<Emphasis Type="Italic">PARP-1 Val762Ala</Emphasis> polymorphism,CagA<Superscript>+</Superscript><Emphasis Type="Italic">H. pylori</Emphasis> infection and risk for gastric cancer in Han Chinese population

Introduction PARP-1 plays important role in the BER (base excision repair) and maintenance of genomic integrity. Previous study found the Val762Ala genetic variant in the PARP-1 gene contributed to susceptibility of some cancers and decreased PARP-1 enzyme activity in response to oxidative damage. Helicobacter pylori (H. pylori) infection was thought to be one of the major causes of gastric cancer. In this study, we investigated the association between the PARP-1 Val762Ala polymorphism, CagA⁺ H. pylori infection, and the risk for gastric cancer. Methods This hospital-based, case–control study was performed involving 556 individuals (236 cases with gastric cancer and 320 controls without evidence of neoplasm and gastrointestinal disease) using a PCR-RFLP method. Chi-square test and logistic regression analysis were used to count OR and 95% CI. Results 762Ala/Ala genotype was overrepresented in the cases (16.9%) compared with controls (10.3%), (OR, 1.942; 95% CI, 1.157–3.257, P = 0.011). Multivariate analysis showed that two factors were significantly associated with risk of gastric cancer, including CagA⁺ H. pylori infection (OR, 2.562; 95% CI, 1.174–5.240, P = 0.037), PARP-1 762AA genotype (OR, 1.772; 95% CI, 1.065–3.867; P = 0.042). Stratification analysis indicated that among Cag⁺ H. pylori positive subjects, 762Ala/Ala carriers had higher risk for developing gastric cancer compared with 762Val/Val carrier (OR, 2.337; 95% CI, 1.148–4.758; P = 0.017). Conclusion PARP-1 762Ala/Ala could be a risk factor for gastric cancer in Han Chinese population; PARP-1 762Val/Ala polymorphism and Cag⁺ H. pylori infection jointly contribute to higher risk for gastric cancer.     

Helicobacter pylori infection can change the intensity of gastric Lewis antigen expressions differently between adults and children

This study tested whether there were different expressions of gastric Lewis antigens between children and adults with Helicobacter pylori infection, and whether the difference was related to the infection outcome. About 68 dyspeptic children and 110 dyspeptic adults were enrolled to check H. pylori infection, its colonization density, and the related histology. Gastric Lewis antigens b (Le^b), x (Le^x), and sialyl-Lewis x (sialyl-Le^x) were immunohistochemically stained and scored for the intensity. The H. pylori-infected adults, but not the children, had a lower Le^b intensity over the antrum (p = 0.019) but higher Le^b intensity over the corpus (p = 0.001) than the non-infected ones. Over the antrum, both the H. pylori-infected children and adults had a lower Le^x and higher sialyl-Le^x intensity than those non-infected ones (p < 0.05). The H. pylori-infected adults had a higher bacterial density (p = 0.004) and Le^b intensity (p = 0.016) over the corpus than the H. pylori-infected children. For the H. pylori-infected adults, but not children, the corpus had a higher Le^b (p = 0.038) and lower Le^x (p = 0.005) intensity than the antrum. Furthermore, the H. pylori-infected adults expressed a higher Le^b and had a higher bacterial density than those with weak Le^b (antrum, p < 0.001; corpus, p = 0.001). In conclusion, H. pylori infection is associated with the intensity change of Lewis antigen expressions in the stomach. The changes of gastric Lewis antigen expressions are different between adults and children with H. pylori infection, which may exert different H. pylori colonization over the corpus between adults and children.     

Cytokine expression due to Helicobacter pylori in a tissue culture model

Helicobacter pylori, in recent years, has been recognized as the major causative agent in chronic gastritis and peptic ulcer disease in humans. H. pylori is a ubiquitous organism, with at least half of the world’s population infected. Of those individuals with peptic ulcer disease, it is estimated that 90% of cases are caused by H. pylori. Currently, the efficacy of therapies is starting to decline due to increasing resistance rates, especially towards clarithromycin. Due to this, new therapies are needed to combat this bacterium. It is hypothesized that cytokine release (especially interleukin-1β, -6, -8, and TNF-α) due to H. pylori infection and the subsequent influx of inflammatory cells causes a massive release of reactive oxygen species (ROS) during the inflammatory reaction. The ROS then cause the pathologic changes seen in the infected tissues. In this study, human gastric adenocarcinoma cell line ATCC 1739 (a cell line not previously evaluated) was examined for its production of interleukin-1β, -6, -8, and TNF-α when cocultured in a ratio of 10:1 H. pylori to adenocarcinoma cells, to determine its value as a model to demonstrate the inflammatory response. Results from this study indicated that ATCC 1739 cells only reliably produced IL-8 when cocultured with H. pylori and stimulated with TNF-α. The production of IL-1β, IL-6, and TNF-α by the ATCC 1739 cells was no different in H. pylori-exposed cells than non-exposed cells. It was concluded that the ATCC 1739 cell line is not suitable to study the effects of coculture with H. pylori on cytokine production.     

Dual roles of CagA protein in Helicobacterpylori-induced chronic gastritis in mice

Cytotoxin-associated gene A (CagA) acts directly on gastric epithelial cells. However, the roles of CagA in host adaptive immunity against Helicobacter pylori (H. pylori) infection are not fully understood. In this study, to investigate the roles of CagA in the development of H. pylori-induced chronic gastritis, we used an adoptive-transfer model in which spleen cells from C57BL/6 mice with or without H. pylori infection were transferred into RAG2^−/− mice, with gastric colonization of either CagA⁺H. pylori or CagA⁻H. pylori. Colonization of CagA⁺H. pylori but not CagA⁻H. pylori in the host gastric mucosa induced severe chronic gastritis in RAG2^−/− mice transferred with spleen cells from H. pylori-uninfected mice. In addition, when CagA⁺H. pylori-primed spleen cells were transferred into RAG2^−/− mice, CD4⁺ T cell infiltration in the host gastric mucosa were observed only in RAG2^−/− mice infected with CagA⁺H. pylori but not CagA⁻H. pylori, suggesting that colonization of CagA⁺H. pylori in the host gastric mucosa is essential for the migration of H. pylori-primed CD4⁺ T cells. On the other hand, transfer of CagA⁻H. pylori-primed spleen cells into CagA⁺H. pylori-infected RAG2^−/− mice induced more severe chronic gastritis with less Foxp3⁺ regulatory T-cell infiltration as compared to transfer of CagA⁺H. pylori-primed spleen cells. In conclusion, CagA in the stomach plays an important role in the migration of H. pylori-primed CD4⁺ T cells in the gastric mucosa, whereas CagA-dependent T-cell priming induces regulatory T-cell differentiation, suggesting dual roles for CagA in the pathophysiology of H. pylori-induced chronic gastritis.     

Presence of the cagA Gene in the Majority of Helicobacter pylori Strains Is Independent of Whether the Individual Has Duodenal Ulcer or Asymptomatic Gastritis

Background Helicobacter pylori infection presents as many different diseases, including asymptomatic gastritis, peptic ulcer disease, and gastric cancer. Although the virulence factor(s) responsible for different H. pylori-related diseases have not been identified, several candidate genes are being investigated for such an association. The polymerase chain reaction (PCR) frequently is used to assess the presence of genetic factors associated with pathogenesis of disease; the cagA gene and its product have been postulated to have a disease-specific relationship to peptic ulcer and gastric cancer because of differential expression in these diseases compared to histological gastritis alone. Materials and Methods. Genomic DNA was amplified by PCR, using synthetic oligonucleotide primers to the cagA gene to determine the prevalence of the cagA gene in 60 H. pylori isolates obtained from well-documented duodenal ulcer or asymptomatic gastritis patients (30 each). Results were confirmed by hybridization with a 1.4-Kb cagA probe. Results. The expected PCR product was obtained in 90% of isolates from duodenal ulcer patients, compared to 70% of isolates from individuals with asymptomatic gastritis. The PCR products were polymorphic in size, suggesting cagA gene sequence differences among isolates. Evaluation for the presence of the cagA gene by hybridization with a 1.4-Kb cagA probe showed a homologous product in 29 of 30 strains [96.7%; 95% confidence interval (CI) = 83–100%] from duodenal ulcer patients versus 25 of 30 strains (83.3%; 95% CI = 65–94%) obtained from individuals with asymptomatic gastritis (p= 0.19). Conclusions. The high prevalence of the cagA gene in asymptomatic gastritis suggests that it will not prove to be a useful marker to distinguish more virulent or disease-specific H. pylori strains. The genetic heterogeneity among H. pylori strains makes PCR an unwise choice as the single method to determine prevalence of a putative virulence factor. In evaluation of the prevalence of a gene or genetic factor in a population of H. pylori, hybridization with extended probes might be important to ensure that the results are representative of the organism's genotype.     

幽门螺杆菌分子伴侣GroEL相互作用蛋白分析

【目的】获得幽门螺杆菌(Helicobacter pylori,HP) GroEL结合蛋白质组构成谱,为进一步探究GroEL及其与相互作用蛋白在HP致病机制中的作用提供新思路。【方法】在构建HP GroEL原核表达重组大肠杆菌(Escherichia coli) BL21(DE3)⁽pET-28a(+)-groEL)基础上,纯化带有His标签的GroEL蛋白,与HP全菌蛋白提取液共孵育后,利用Protein G磁珠和抗His标签抗体免疫沉淀法对复合物进行捕获,然后对复合物中GroEL及其结合的蛋白质进行质谱法鉴定,根据主要功能对其进行分类,并完成蛋白质相互关系网络分析。【结果】对GroEL蛋白捕获成分进行分析,共鉴定出59种可能与GroEL结合的蛋白质,其中包括19种代谢酶类(KatA、GltA和AhpC等参与氧化还原相关酶类7种,PepA、RocF和HtrA等肽酶5种,以及2种参与脂肪代谢酶、2种参与ATP合成酶、2种尿素酶和HP17＿08079蛋白等)、15种外膜蛋白(黏附素BabA、SabA、HapA及其他膜蛋白等)、8种转录翻译相关蛋白(Tuf、RpoBC...     

Prevalence of Helicobacter pylori genotypes (vacA, cagA, cagE and virB11) in gastric cancer in Brazilian's patients: an association with histopathological parameters

Purpose: To investigate the frequency and the association of vacA alleles, cagA, cagE and virB11 genes of Helicobacter pylori from patients with gastric cancer, considering the clinic histopathological parameters. Methods: One hundred and one gastric adenocarcinoma tissues were assessed by PCR to detect H. pylori and vacA alleles, cagA, cagE and virB11. Results: The distribution of cases according to the presence of the genes studied showed that the group containing vacA s1m1, cagA, cagE and virB11 H. pylori genes was significantly more frequent, followed by the group with at least one marker on the right side and left of the island. They were also present in the early stages and were the most frequent in nearly all histopathological grades. Conclusions: This study verified that vacAs1m1 and cag-PAI genes, cagA, cagE and virB11 are important H. pylori markers for gastric cancer development. Also, this study corroborates the importance of cagE and cagA together as cag-PAI marker.     

Gastric mucosal changes induced by long term infection with Helicobacter pylori in Mongolian gerbils: effects of bacteria eradication

Helicobacter pylori infection has been reported to induce various mucosal changes, including gastric adenocarcinoma, in Mongolian gerbils 62 weeks after inoculation. Using Mongolian gerbils, this study examined whether or not eradication of the bacteria with drugs at specified times after infection prevents the development of mucosal changes. After orally inoculating with H. pylori (TN2GF4, vacA- and cagA-positive), the animals were killed 18 months later. Four or 8 months after H. pylori inoculation, eradication was performed by concurrent treatment with omeprazole+clarithromycin. Immediately after treatment ended, in both the 5 and 9 month groups, it was verified that H. pylori was completely eradicated. Autopsy performed 18 months after H. pylori inoculation revealed gastric hyperplastic polyps with erosive lesions and ulcers that were grossly visible in the non-treated control group. In addition, atrophic gastritis, intestinal metaplasia, carcinoids, and adenocarcinomas were histologically observed in the animals. In animals eradicated after 4 months and autopsied after 18 months, however, such mucosal changes were not observed. In contrast, intestinal metaplasia and mucosal atrophy was observed in animals eradicated after 8 months and autopsied after 18 months. It was concluded that early eradication of H. pylori infection with drug therapy can prevent severe gastric mucosal changes, to include adenocarcinomas, in Mongolian gerbils.     

A proteomic approach for the identification of bismuth-binding proteins in Helicobacter pylori

Helicobacter pylori is a major human pathogen that can cause peptic ulcers and chronic gastritis. Bismuth-based triple or quadruple therapies are commonly recommended for the treatment of H. pylori infections. However, the molecular mechanisms underlying treatment with bismuth are currently not fully understood. We have conducted a detailed comparative proteomic analysis of H. pylori cells both before and after treatment with colloidal bismuth subcitrate (CBS). Eight proteins were found to be significantly upregulated or downregulated in the presence of CBS (20 μg mL⁻¹). Bismuth-induced oxidative stress was confirmed by detecting higher levels of lipid hydroperoxide (approximately 1.8 times) and hemin (approximately 3.4 times), in whole cell extracts of bismuth-treated H. pylori cells, compared with those from untreated cells. The presence of bismuth also led to an approximately eightfold decrease in cellular protease activities. Using immobilized-bismuth affinity chromatography, we isolated and subsequently identified seven bismuth-binding proteins from H. pylori cell extracts. The intracellular levels of four of these proteins (HspA, HspB, NapA and TsaA) were influenced by the addition of CBS, which strongly suggests that they interact directly with bismuth. The other bismuth-interacting proteins identified were two enzymes (fumarase and the urease subunit UreB), and a translational factor (Ef-Tu). Our data suggest that the inhibition of proteases, modulation of cellular oxidative stress and interference with nickel homeostasis may be key processes underlying the molecular mechanism of bismuth’s actions against H. pylori. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.