A plasmid-based vector system for the cloning and expression of Helicobacter pylori genes encoding outer membrane proteins

Helicobacter pylori produces a number of proteins associated with the outer membrane, including adhesins and the vacuolating cytotoxin. We observed that the functional expression of such proteins is deleterious to Escherichia coli, the host bacterium used for gene cloning. Therefore, a general method was developed for the functional expression of such genes on a shuttle vector in H. pylori, which has been termed SOMPES (Shuttle vector-based Outer Membrane Protein Expression System). The intact, active gene is reconstituted by recombination in H. pylori from partial gene sequences cloned on an E. coli-H. pylori shuttle vector. This system was established in an H. pylori strain carrying a precise, unmarked chromosomal deletion of the vacA gene, which was constructed by adapting the streptomycin sensitivity system to H. pylori. It is based on the expression of the H. pylori rpsL gene as a counterselectable marker in the genetic background of an rpsL mutant. The utility of this approach is demonstrated by the expression of a recombinant gene encoding vacuolating cytotoxin (vacA) and a recombinant gene encoding an adherence-associated outer membrane protein (alpA) in H. pylori. Received: 10 May 1999 / Accepted: 7 July 1999     

Review: Impact of Helicobacter pylori on Alzheimer's disease: What do we know so far?

Background

Helicobacter pylori has changed radically gastroenterologic world, offering a new concept in patients' management. Over time, more medical data gave rise to diverse distant, extragastric manifestations and interactions of the “new” discovered bacterium. Special interest appeared within the field of neurodegenerative diseases and particularly Alzheimer's disease, as the latter and Helicobacter pylori infection are associated with a large public health burden and Alzheimer's disease ranks as the leading cause of disability. However, the relationship between Helicobacter pylori infection and Alzheimer's disease remains uncertain.

Methods

We performed a narrative review regarding a possible connection between Helicobacter pylori and Alzheimer's disease. All accessible relevant (pre)clinical studies written in English were included. Both affected pathologies were briefly analyzed, and relevant studies are discussed, trying to focus on the possible pathogenetic role of this bacterium in Alzheimer's disease.

Results

Data stemming from both epidemiologic studies and animal experiments seem to be rather encouraging, tending to confirm the hypothesis that Helicobacter pylori infection might influence the course of Alzheimer's disease pleiotropically. Possible main mechanisms may include the bacterium's access to the brain via the oral‐nasal‐olfactory pathway or by circulating monocytes (infected with Helicobacter pylori due to defective autophagy) through disrupted blood‐brain barrier, thereby possibly triggering neurodegeneration.

Conclusions

Current data suggest that Helicobacter pylori infection might influence the pathophysiology of Alzheimer's disease. However, further large‐scale randomized controlled trials are mandatory to clarify a possible favorable effect of Helicobacter pylori eradication on Alzheimer's disease pathophysiology, before the recommendation of short‐term and cost‐effective therapeutic regimens against Helicobacter pylori‐related Alzheimer's disease.     

A meta‐analysis of the association between Helicobacter pylori (H. pylori) infection and hyperemesis gravidarum

Background

Hyperemesis gravidarum remains a common, distressing, and significant yet poorly understood disorder during pregnancy. The association between maternal Helicobacter pylori (H. pylori) infection and hyperemesis gravidarum has been increasingly recognized and investigated. This study thus aimed to provide an updated review and meta‐analysis of the topic.

Methods

Using the search terms (H. pyloriOR Helicobacter ORHelicobacter pyloriOR infection) AND (pregnancy OR emesis OR hyperemesis gravidarum OR nausea OR vomiting), a preliminary search on the PubMed, Ovid, Web of Science, Google Scholar, and WanFang database yielded 372 papers published in English between January 1st, 1960 and June 1st, 2017.

Results

A total of 38 cross‐sectional and case‐control studies, with a total of 10 289 patients were eligible for review. Meta‐analysis revealed a significant association between H. pylori infection and hyperemesis gravidarum during pregnancy, with a pooled odds ratio of 1.348 (95% CI: 1.156‐1.539, P < .001). Subgroup analysis found that serologic and stool antigen tests were comparable methods of detecting H. pylori as they yielded similar odds ratios.

Limitations

Although the studies did not have high heterogeneity (I² = 28%), publication bias was observed, and interstudy discrepancies in the diagnostic criteria adopted for hyperemesis gravidarum limit the reliability of findings. Also, 15 of the included studies were from the same country (Turkey), which could limit the generalizability of current findings. The prevalence of H. pylori infection varies throughout the world, and there may also be pathogenic differences as most strains of H. pylori in East Asia carry the cytotoxin‐associated gene A gene.

Conclusion

H. pylori infection was associated with an increased likelihood of hyperemesis gravidarum during pregnancy. Given the high prevalence of H. pylori infections worldwide, detecting H. pylori infection and the eradication of maternal H. pylori infection could be part of maternal hyperemesis gravidarum management. Further confirmation with robust longitudinal studies and mechanistic investigations are needed.     

Peptic ulcers and erosions are common in Israeli children undergoing upper endoscopy

Background: Peptic ulcers and erosions (PU&E) are thought to be uncommon in children. Patients with early exposure to Helicobacter pylori may be at a higher risk for early onset PU&E. Children in Israel have a high prevalence and early acquisition of Helicobacter pylori (H. pylori) and have easy access to pediatric gastroenterologists and endoscopy. Our aim was to describe the prevalence and characteristics of PU&E in this population referred by Pediatric Gastroenterologists for an upper endoscopy. Methods: We conducted a retrospective study over the years January 2003–May 2006. Over these years we had information on 751 diagnostic upper endoscopies. PU&E was regarded as erosive gastritis/duodenitis or ulcer in either the stomach or duodenum. H. pylori status was assessed using rapid urease test and gastric biopsies. Results: PU&E was detected in 169 (22.5%) patients (ulcers 51 (6.8%), erosions 118 (15.7%)). One hundred twenty‐four had gastric PU&E and 58 had duodenal PU&E. H. pylori was positive in 112 (66.3%). H. pylori‐associated PU&E becomes common after age 10 years, with gastric PU&E presenting much earlier than duodenal disease. Most of the H. pylori‐negative PU&E were idiopathic and improved symptomatically on PPI treatment. Interestingly, 43% of patients with PU&E in our cohort were either immigrants from the former Soviet Union or of Israeli Arab origin. Conclusions: PU&E appears to be common in this selected population with a relatively high incidence of gastric PU&E. H. pylori associated PU&E becomes common after age 10 years with gastric PU&E presenting much earlier than duodenal disease. Non H. pylori PU&E in children comprises approximately a third of all PU&E, are mostly idiopathic and appear earlier than H. pylori associated PU&E.     

Potential antigen candidates for subunit vaccine development against Helicobacter pylori infection

Helicobacter pylori (H. pylori) is a resident bacterium in the stomach that accounts for 75% cases of gastric cancer. In this review, we comprehensively studied published papers on H. pylori vaccines using Google Scholar and NCBI databases to gather information about vaccines against H. pylori. Considering the pivotal roles of the enzyme urease (in production of NH₃ and neutralization of the acidic medium of the stomach), cytotoxin-associated gene A, and vacuolating cytotoxin A proteins in H. pylori infection, they could be the best candidates for the construction of recombinant vaccines. The outer membrane porins (Hop), blood group antigen-binding adhesin (BabA), sialic acid-binding adhesin (SabA), and outer inflammatory protein A, play significant roles in binding of bacterium to human gastric tissues, and because binding is the first step in bacterial fixation and colonization, these antigens also can be considered as suitable candidates for designing vaccines. Likely, other significant bacterial antigens, such as NapA (chemotactic factor for recruitment of human neutrophils and monocytes to the site of infection), duodenal ulcer promoting protein A (to promote duodenal ulcer), and Hsp60 (as a molecular chaperon for activation of urease enzyme), can be used in the construction of subunit vaccines. New vaccines in use currently, such as DNA vaccines and subunit vaccines, can efficiently replace the dead and attenuated vaccines. Nonetheless, the results show that urease enzyme is most used compared with bacterial components in the designing and construction of recombinant vaccines. The BabA and SabA antigens belong to the outer membrane porins family in H. pylori and are required for binding and fixation of the bacterium to the human gastric tissues.     

Presence of Helicobacter pylori in a Sibling is Associated with a Long‐Term Increased Risk of H. pylori Infection in Israeli Arab Children

Background and Objectives: We examined the dynamics of Helicobacter pylori infection between pre‐school and school ages and compared the determinants of late acquisition of H. pylori infection with determinants of early and persistent H. pylori infection. Methods: ELISA was used to detect H. pylori antigens in stool specimens collected from children at preschool age (3–5 years) and from their mothers and siblings in 2004. The children were tested again for H. pylori at school age (6–9 years) in 2007–2009. Household and socioeconomic characteristics were obtained by interviews. Results: The prevalence of H. pylori infection increased from 49.7% (95% CI 42.8, 56.7) in 2004 to 58.9% (95% CI 51.8, 65.6) in 2007–2009. Among children tested in both examinations, 69 (49.3%) had persistent infection, 14 (10.0%) were new cases, 56 (40.0%) remained uninfected, and one (0.7%) had lost H. pylori infection. The approximate annual incidence of infection during 2004–2009 was 5%. Sibling’s H. pylori positivity at baseline increased the risk for late acquisition of H. pylori infection; adjusted prevalence ratio (PR) 4.62 (95% CI 0.76, 28.23) (p = .09), while maternal education lowered the risk; adjusted PR 0.84 (95% CI 0.69, 1.01) (p = .06). Sibling’s H. pylori positivity was the only significant variable associated with early and persistent H. pylori infection in multivariate analysis. Conclusions: Most H. pylori infections are acquired at preschool age and transient infection beyond this age is uncommon in this population. Helicobacter pylori‐infected siblings are the major reservoir of H. pylori in early and late childhood demonstrating sustained intra‐familial transmission of H. pylori.     

Helicobacter pylori infection in connective tissue disorders is associated with high levels of antibodies to mycobacterial hsp65 but not to human hsp60

Background. To investigate whether the Helicobacter pylori status influences levels of antibodies against mycobacterial heat shock protein (hsp) 65 and human hsp60 in systemic autoimmune diseases and to study the concentration of anti‐H. pylori antibodies in autoimmune patients and healthy controls. Materials and Methods. Antibodies against human heat‐shock protein hsp60, mycobacterial heat‐shock protein hsp65 were analyzed by ELISA. Anti‐Helicobacter antibodies were determined by enzyme immunoassay. Results. There was a markedly higher prevalence of H. pylori infection in undifferentiated connective tissue disease (82%) (n = 33) and systemic sclerosis (78%) (n = 55) but not in systemic lupus erythematosus (n = 49), polymyositis/dermatomyositis (n = 14), rheumatoid arthritis (n = 21) or primary Raynaud's syndrome (n = 26) compared with controls (59%) (n = 349). In autoimmune diseases H. pylori infection was associated with elevated levels of antihsp65 (p = .008) but not of antihsp60. Anti‐hsp65 levels were significantly higher in H. pylori‐infected (n = 129) than in uninfected patients (n = 69) (p = .0007). Conclusions. These findings indicate that in autoimmune diseases the infection with the H. pylori bacterium is associated with increased concentration of antimycobacterial hsp65.     

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.     

Helicobacter pylori Infection – A Boon or a Bane: Lessons from Studies in a Low‐Prevalence Population

Helicobacter pylori (H. pylori) infection is etiologically associated with gastric cancer and peptic ulcer diseases which are both important public health burdens which could be largely eliminated by H. pylori eradication. However, some investigators urge caution based on the hypothesis that eradication of H. pylori may result in an increase in the incidence of gastroesophageal reflux disease, esophageal adenocarcinoma, and childhood asthma. The ethnic Malays of northeastern Peninsular Malaysia have long had a low prevalence of H. pylori infection and, as expected, the incidence of gastric cancer and its precursor lesions is exceptionally low. The availability of a population with a low H. pylori prevalence and generally poor sanitation allows separation of H. pylori from the hygiene hypothesis and direct testing of whether absence of H. pylori is associated with untoward consequence. Contrary to predictions, in Malays, erosive esophagitis, Barrett's esophagus, distal esophageal cancers, and childhood asthma are all of low incidence. This suggests that H. pylori is not protective rather the presence of H. pylori infection is likely a surrogate for poor hygiene and not an important source of antigens involved in the hygiene hypothesis. Helicobacter pylori in Malays is related to transmission from H. pylori‐infected non‐Malay immigrants. The factors responsible for low H. pylori acquisition, transmission, and burden of H. pylori infection in Malays remain unclear and likely involves a combination of environmental, host (gene polymorphisms), and strain virulence factors. Based on evidence from this population, absence of H. pylori infection is more likely to be boon than a bane.     

Dynamics of the Cag‐type IV secretion system of Helicobacter pylori as studied by bacterial co‐infections

Many pathogenic Gram‐negative bacteria possess type IV secretion systems (T4SS) to inject effector proteins directly into host cells to modulate cellular processes to their benefit. The human bacterial pathogen Helicobacter pylori, a major aetiological agent in the development of chronic gastritis, duodenal ulcer and gastric carcinoma, harbours the cag‐T4SS to inject the cytotoxin associated Antigen (CagA) into gastric epithelial cells. This results in deregulation of major signalling cascades, actin‐cytoskeletal rearrangements and eventually gastric cancer. We show here that a pre‐infection with live H. pylori has a dose‐dependent negative effect on the CagA translocation efficiency of a later infecting strain. This effect of the ‘first’ strain was independent of any of its T4SS, the vacuolating cytotoxin (VacA) or flagella. Other bacterial pathogens, e.g. pathogenic Escherichia coli, Campylobacter jejuni, Staphylococcus aureus, or commensal bacteria, such as lactobacilli, were unable to interfere with H. pylori's CagA translocation capacity in the same way. This interference was independent of the β1 integrin receptor availability for H. pylori, but certain H. pylori outer membrane proteins, such as HopI, HopQ or AlpAB, were essential for the effect. We suggest that the specific interference mechanism induced by H. pylori represents a cellularresponse to restrict and control CagA translocation into a host cell to control the cellular damage.     

Detection of Helicobacter pylori in sewage and water using a new quantitative PCR method with SYBR green

Aims: To demonstrate the application of a new quantitative polymerase chain reaction (qPCR) technique for the determination of Helicobacter pylori concentrations in water, and to use this method to investigate the occurrence of the bacteria in sewage. The other aim was to study the survival capacity and detectability of the bacteria in artificially contaminated groundwater at different temperatures of 4 and 15°C. Methods and Results: The detection of H. pylori in water was aided by PCR using specific primers designed for the amplification of a fragment within the major vacuolating cytotoxin gene. Conventional culture was compared with conventional PCR and the new real-time (RT) qPCR approach for the quantification of the bacterium. Helicobacter pylori remained culturable for 120 h at 4°C as opposed to only 24 h at 15°C. RT qPCR demonstrated a 100-fold greater sensitivity for the detection of H. pylori DNA in comparison with conventional PCR. Scanning electron microscopic (SEM) observation showed that the normal spiral form changed to a coccoid form after 24 and 72 h at 15 and 4°C, respectively. Helicobacter pylori was found at 2–28 cells ml⁻¹ in sewage, of the 23 sewage samples – 84% were positive for H. pylori species-specific vacuolating cyctotoxin gene (vacA) by RT qPCR, but were negative by conventional PCR. Conclusions: The RT qPCR assay provided a specific, sensitive and rapid method for the quantitative detection of H. pylori in sewage. This molecular method would be valuable in studying the prevalence of H. pylori as required by the United States Environmental Protection Agency Contaminant Candidate List, particularly in nondisinfected ground waters, in sewage as a source of contamination, and for addressing the possible presence of viable but nonculturable of H. pylori. Significance and Impact of the Study: The quantitative detection of H. pylori by rapid and less-expensive methods than the TaqMan Assay using SYBR green could be an important tool to monitor infection in community by measuring the concentrations in sewage and to meet the new regulatory and risk-based frameworks for water supplies.     

Development of Immunoreagents for Diagnostics of CagA‐Positive Helicobacter pylori Infections

Background: Helicobacter pylori strains expressing cytotoxic CagA protein are more likely to provoke severe gastric mucosal pathology and cause adenocarcinoma development than that lacking CagA. Determination of the CagA‐status of a pathogen, therefore, is regarded as informative approach in H. pylori infection diagnostics and disease risk prediction. Materials and Methods: Molecular cloning, recombinant protein expression in Escherichia coli, affinity chromatography, electrophoresis and commonly used techniques of hybridoma production and screening were used as well as different immunosorbent assays and Western blot procedures. Results: Four overlapping N‐terminally His₆‐tagged recombinant fragments of CagA that covered the entire CagA sequence were produced and purified. An ELISA for specific anti‐CagA serum antibodies detection was developed and evaluated. Utilizing recombinant fragments, the first set of monoclonal antibodies against CagA‐antigen was produced and characterized. Three antibodies recognized distinct linear epitopes inside conserved regions of the cytotoxin whereas the epitope of the forth antibody was mapped in the variable area of CagA. The monoclonal antibodies allowed discriminating CagA‐positive and CagA‐negative H. pylori strains by means of Western blot and immunosorbent assays. Conclusions: The use of recombinant protein technology allowed obtaining pure CagA antigen, thus providing new perspectives for development of immunodiagnostic reagents. The set of monoclonal antibodies is a valuable tool for determination of CagA‐status of H. pylori infection and for the investigation of cytotoxin molecule as well.     

Methods to monitor autophagy in H. pylori vacuolating cytotoxin A (VacA)-treated cells

Helicobacter pylori is a gram negative pathogen that infects at least half of the world’s population and is associated not only with gastric cancer but also with other diseases such as gastritis and peptic ulcers. Indeed, H. pylori is considered the single most important risk factor for the development of gastric cancer. The vacuolating cytotoxin, VacA, secreted by H. pylori promotes intracellular survival of the bacterium and modulates host immune responses. In a recent study, we reported that VacA induces autophagy. Multilamellar autophagosomes are detected in gastric epithelial cells that are distinct from the large vacuoles formed by VacA. Furthermore, inhibition of autophagy stabilizes VacA and reduces vacuolation in the cells indicating that the toxin is being degraded by autophagy, thus limiting toxin-induced host cell damage. Many of the methods that were used for this study are commonly employed techniques that were adapted for H. pylori infection and VacA intoxication. In this paper, we describe the various methods and specific protocols used for the assessment and monitoring of autophagy during H. pylori infection.     

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.     

Inverse association between Helicobacter pylori and pediatric asthma in a high-prevalence population

Background: Helicobacter pylori‐associated disease has led to aggressive diagnostic and eradication protocols that are partially responsible for the decrease in prevalence of H. pylori carriage. Recent evidence indicates that in low‐prevalence populations, H. pylori may have protective effects on allergic diseases. The aim of this study was to explore the relationship between pediatric asthma and H. pylori infection in a population with high prevalence of H. pylori infection. Materials and Methods: A national referral laboratory was screened for all ¹³C urea breath tests performed in children aged 5–18 years between 2007 and 2008, for patient demographics and physician‐diagnosed asthma. Data concerning asthma‐associated medication usage were extracted from electronic medical records and databases. Data were analyzed using a stepwise logistic regression model. Results: During the study period, 6959 patients underwent urea breath testing (average age 12.4 ± 3.5 years). Of these, 3175/6959 (45.6%) were positive for H. pylori, and 578/6959 (8.3%) had asthma. Rates of asthma in H. pylori‐positive and H. pylori‐negative children were 7.3 and 9.1%, respectively (odds ratio 0.82; 95% confidence interval (CI) 0.69–0.98; p = .032). We also confirmed that male gender, urban residence, and age are associated with childhood asthma. Conclusions: We demonstrate an inverse association between H. pylori and pediatric asthma in a population with a high prevalence of H. pylori.     

Detection of Helicobacteraceae in intestinal biopsies of children with Crohn's disease

Background: Given that members of Helicobacteraceae family colonize the intestinal mucus layer, it has been hypothesized that they may play a role in Crohn’s disease. This study investigated the presence of Helicobacteraceae DNA in biopsies collected from children with Crohn’s disease and controls. Materials and Methods: The presence of Helicobacteraceae DNA was investigated in intestinal biopsies collected from 179 children undergoing colonoscopy (Crohn’s disease n = 77, controls n = 102) using a Helicobacteraceae‐specific PCR. Results: Members of the Helicobacteraceae were detected in 32/77 children with Crohn’s disease (41.5%) and 23/102 controls (22.5%). Statistical analysis showed the prevalence of Helicobacteraceae detected in patients to be significantly higher than that in controls (p = .0062). Analysis of non‐pylori Helicobacteraceae showed that their prevalence was also significantly higher in patients than in controls (p = .04). Helicobacter pylori was detected in 14.0% of the biopsies across all groups. Given that all children tested were negative for gastric H. pylori, this was a surprising finding. Phylogenetic analysis of H. pylori sequences detected in the biopsies showed that the H. pylori strains identified in the patients did not group with gastric H. pylori included in the analysis, but rather with other H. pylori strains detected in the intestine, gall bladder, and liver. Conclusions:  The higher prevalence of Helicobacteraceae DNA in Crohn’s disease patients would suggest that members of this family may be involved in this disease. In addition, phylogenetic analysis of H. pylori strains showed that extragastric sequences clustered together, indicating that different H. pylori strains may adapt to colonize extragastric niches.     

Haem oxygenase‐1 inhibits phosphorylation of the Helicobacter pylori oncoprotein CagA in gastric epithelial cells

The cytotoxin‐associated gene A protein (CagA) plays a pivotal role in the aetiology of Helicobacter pylori‐associated gastric diseases. CagA is injected into the cytoplasm of host cells by a type IV secretion system, and is phosphorylated on tyrosine residues by the host enzyme c‐Src. We previously reported that the enzyme haem oxygenase‐1 (HO‐1) inhibits IL‐8 secretion by H. pylori‐infected cells. However, the cellular mechanism by which HO‐1 regulates the innate immune function of infected cells remains unknown. We now show that nitric oxide and haemin, two inducers of HO‐1, decrease the level of phosphorylated CagA (p‐CagA) in H. pylori‐infected gastric epithelial cells and this is blocked by either pharmacological inhibition of HO‐1 or siRNA knockdown of hmox‐1. Moreover, forced expression of HO‐1 by transfection of a plasmid expressing hmox‐1 also results in a strong attenuation of CagA phosphorylation. This occurs through the inhibition of H. pylori‐induced c‐Src phosphorylation/activation by HO‐1.Consequently, H. pylori‐induced cytoskeletal rearrangements and activation of the pro‐inflammatory response mediated by p‐CagA are inhibited in HO‐1‐expressing cells. These data highlight a mechanism by which the innate immune response of the host can restrict the pathogenicity of H. pylori by attenuating CagA phosphorylation in gastric epithelial cells.     

The vacuolating cytotoxin of Helicobacter pylori

Helicobacter pylori, the causative agent of chronic superficial gastritis and duodenal ulcer disease in humans, produces a unique cytotoxin (VacA) that induces cytoplasmic vacuolation in eukaryotic cells. The structural organization and processing of the vacuolating cytotoxin are characteristic of a family of proteins exemplified by Neisseria gonorrhoeae IgA protease. Although only 50% of H. pylori isolates produce detectable cytotoxin activity in vitro, vacA homologues are present in virtually all isolates. Several families of vacA alleles have been identified, and there is a strong correlation between presence of specific vacA genotypes, cytotoxin activity, and peptic ulceration. Experiments in a mouse model of H. pylori-induced gastric damage indicate that the cytotoxin plays an important role in inducing gastric epithelial necrosis.     

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.