Polymorphism of flagellin A gene in Helicobacter pylori.
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ABSTRACT:
SUBMITTER: Ji WS
PROVIDER: S-EPMC4695594 | biostudies-literature | 2001 Dec
REPOSITORIES: biostudies-literature
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Project description:BackgroundFlagella-mediated motility is both a crucial virulence determinant of Helicobacter pylori and a factor associated with gastrointestinal diseases. Flagellar formation requires flagellins to be glycosylated with pseudaminic acid (Pse), a process that has been extensively studied. However, the transfer of Pse to flagellins remains poorly understood. Therefore, the aim of this study is to characterize a putative glycosyltransferase jhp0106 in flagellar formation.Materials and methodsWestern blotting and chemical deglycosylation were performed to examine FlaA glycosylation. Protein structural analyses were executed to identify the active site residues of Jhp0106, while the Jhp0106-FlaA interaction was examined using a bacterial two-hybrid assay. Lastly, site-directed mutants with mutated active site residues in the jhp0106 gene were generated and investigated using a motility assay, Western blotting, cDNA-qPCR analysis, and electron microscopic examination.ResultsLoss of flagellar formation in the Δjhp0106 mutant was confirmed to be associated with non-glycosylated FlaA. Furthermore, three active site residues of Jhp0106 (S350, F376, and E415) were identified within a potential substrate-binding region. The interaction between FlaA and Jhp0106, Jhp0106::S350A, Jhp0106::F376A, or Jhp0106::E415A was determined to be significant. As well, the substitution of S350A, F376A, or E415A in the site-directed Δjhp0106 mutants resulted in impaired motility, deficient FlaA glycosylation, and lacking flagella. However, these phenotypic changes were regardless of flaA expression, implying an indefinite proteolytic degradation of FlaA occurred.ConclusionsThis study demonstrated that Jhp0106 (PseE) binds to FlaA mediating FlaA glycosylation and flagellar formation. Our discovery of PseE has revealed a new glycosyltransferase family responsible for flagellin glycosylation in pathogens.
| S-EPMC7988653 | biostudies-literature
Project description:The Helicobacter pylori protein HP0958 is essential for flagellum biogenesis. It has been shown that HP0958 stabilizes the sigma(54) factor RpoN. The aim of this study was to further investigate the role of HP0958 in flagellum production in H. pylori. Global transcript analysis identified a number of flagellar genes that were differentially expressed in an HP0958 mutant strain. Among these, the transcription of the major flagellin gene flaA was upregulated twofold, suggesting that HP0958 was a negative regulator of the flaA gene. However, the production of the FlaA protein was significantly reduced in the HP0958 mutant, and this was not due to the decreased stability of the FlaA protein. RNA stability analysis and binding assays indicated that HP0958 binds and destabilizes flaA mRNA. The HP0958 mutant was successfully complemented, confirming that the mutant phenotype described was due to the lack of HP0958. We conclude that HP0958 is a posttranscriptional regulator that modulates the amount of the flaA message available for translation in H. pylori.
| S-EPMC2593243 | biostudies-literature
Project description:Helicobacter mustelae causes chronic gastritis and ulcer disease in ferrets. It is therefore considered an important animal model of human Helicobacter pylori infection. High motility even in a viscous environment is one of the common virulence determinants of Helicobacter species. Their sheathed flagella contain a complex filament that is composed of two distinctly different flagellin subunits, FlaA and FlaB, that are coexpressed in different amounts. Here, we report the cloning and sequence determination of the flaA gene of H. mustelae NCTC12032 from a PCR amplification product. The FlaA protein has a calculated molecular mass of 53 kDa and is 73% homologous to the H. pylori FlaA subunit. Isogenic flaA and flaB mutants of H. mustelae F1 were constructed by means of reverse genetics. A method was established to generate double mutants (flaA flaB) of H. mustelae F1 as well as H. pylori N6. Genotypes, motility properties, and morphologies of the H. mustelae flagellin mutants were determined and compared with those of the H. pylori flaA and flaB mutants described previously. The flagellar organizations of the two Helicobacter species proved to be highly similar. When the flaB genes were disrupted, motility decreased by 30 to 40%. flaA mutants retained weak motility by comparison with strains that were devoid of both flagellin subunits. Weakly positive motility tests of the flaA mutants correlated with the existence of short truncated flagella. In H. mustelae, lateral as well as polar flagella were present in the truncated form. flaA flaB double mutants were completely nonmotile and lacked any form of flagella. These results show that the presence of both flagellin subunits is necessary for complete motility of Helicobacter species. The importance of this flagellar organization for the ability of the bacteria to colonize the gastric mucosa and to persist in the gastric mucus remains to be proven.
| S-EPMC176987 | biostudies-other
Project description:Interleukin-18 (IL-18) is a pleiotropic, pro-inflammatory cytokine that is capable of promoting the Th1 response. A predominant Th1 response induces chronic and persistent inflammatory changes in the gastric mucosa in response to Helicobacter pylori (H. pylori) infection. The aim of this study was to investigate the potential association between IL-18 gene polymorphisms and susceptibility to H. pylori infection in the Korean population. A total of 678 subjects who underwent a routine health check-up were enrolled. The IL-18 gene polymorphisms at positions -656, -607, -137, +113, and +127 were genotyped. H. pylori positivity was demonstrated in 456 subjects (67.3%). The allele frequencies of IL-18 gene polymorphisms at position -137 (rs187238) were different based on the status of H. pylori infection (G vs. C, adjusted OR 0.64 CI: 0.47-0.87, P = 0.005). The results indicate that the genetic variants in the IL-18 gene may be associated with susceptibility to H. pylori infection in the Korean population, suggesting that IL-18 plays a role in the pathogenesis of H. pylori-associated diseases. However, this finding requires further replication and validation.
| S-EPMC4476145 | biostudies-literature
Project description:The global cancer burden of new cases of various types rose with millions of death in 2018. Based on the data extracted by GLOBOCAN 2018, gastric cancer (GC) is the third leading cause of mortality related to cancer across the globe. Carcinogenic or oncogenic infections associated with Helicobacter pylori (Hp) are regarded as one of the essential risk factors for GC development. It contributes to the increased production of cytokines that cause inflammation prior to their growth in the host cells. Hp infections and specific types of polymorphisms within the host cells encoding cytokines are significant contributors to the host's increased susceptibility in terms of the development of GC. Against the backdrop of such an observation is that only a small portion of the cells infected can become malignant. The diversities are a consequence of the differences in the pathogenic pathway of the Hp, susceptibility of the host, environmental conditions, and interplay between these factors. It is evident that hosts carrying cytokine genes with high inflammatory levels and polymorphism tend to exhibit an increased risk of development of GC, with special emphasis being placed on the host cytokines gene polymorphisms.
| S-EPMC8177986 | biostudies-literature
Project description:AIM:To assess the variability of adhesin gene hpaA between different Helicobacter pylori (H pylori) strains with PCR-restriction fragment length polymorphism (RFLP). METHODS:Twelve different H pylori strains were chosen to amplify the 710-bp segments of gene hpaA. These strains were NCTC11637, SS1; Chongqing clinical isolates CCS9801, CCS9802, CCS9803, CCS9806, CCS9809, CCS9810, CCS9813, which were gained from patients of gastritis; Mongolia gerbil adapted H pylori strains (abbreviation MG), which were gained from the following steps: gastric mucosal specimens of Mongolia gerbils infected by clinical isolate CCS9803 were cultured and detected, the positive H pylori strains were named as the first generation of Mongolia gerbil adapted H pylori strains (abbreviation MG1) and then were subcultured with healthy Mongolia gerbil to generate MG2, in turn to gain the ninth generation (abbreviation MG9). All hpaA segments, obtained from 12 different H pylori strains, were digested by HhaI and HaeIII individually and analyzed by agarose gel electrophoresis. RESULTS:In all 12 strains, the 710-bp PCR products were successfully amplified and products were cloned to pMD18-T vector respectively, then the recombinant plasmids were digested simultaneously with NcoI and XhoI to recover the small fragments. The objective fragments from 12 different H pylori strains digested with Hae III could be seen as 4 types of bands and 5 types with Hha I. According to the hpaA RFLP patterns, the 12 H pylori strains could be divided into 5 groups: group I, NCTC11637 and SS1; group II, CCS9809, which RFLP type digested with HaeIII was the same as strains of group I, but HhaI RFLP showed difference compared with the other groups; group III, CCS9810; group IV, CCS9803; group V: CCS9801, CCS9802, CCS9806, CCS9813, MG1, MG3 and MG9. The sequence data of 12 hpaA segments were analyzed by DNAsis software and it was observed that: (1) The homologies of base pair and amino acid sequence between strains NCTC11637, SS1, CCS9809 were 99.6% and 98.9%, respectively; (2) The homology of base pair and amino acid sequence between CCS9803 and CCS9810 was 97.7% and 99.1%; (3) That of the rest strains, CCS9801, CCS9802, CCS9806, CCS9813, MG1, MG3, MG9 reached 99.4% and 98.4%; (4) The base pair homologies between all hpaA fragments of different sources were higher than 94.6%, therefore the correspondence of deduced amino acid sequence was higher than 96.8% between each other. CONCLUSION:The gene hpaA from different H pylori strains revealed variation, and this might provide an effective method for molecular epidemiological survey of H pylori.
| S-EPMC4305759 | biostudies-literature
Project description:To assess the role of Helicobacter pylori infection and interleukin 6 polymorphism -174 (rs1800795) in dyslipidemia.Case-control study comparing serum lipids between H. pylori positive and negative patients and controlling for IL-6 -174 polymorphism, age, sex and smoking.3 hospitals performing outpatient endoscopies in the city of Oulu, Finland.199 adult patients with dyspepsia symptoms fulfilling Rome criteria originating from ethnically Finnish population. Patients with an immunosuppressive disorder or malignant disease, treated H. pylori infection, immunosuppressive or anticoagulant medication, previous gastric surgery or ongoing antibiotic treatment were excluded.Association of H. pylori infection and serum lipid concentrations in the whole group or in genotype-based subgroups. The associations between peptic ulcer, gastric mucosal inflammation and serum lipid concentrations were assessed as secondary outcomes.The median high-density lipoprotein (HDL) serum concentration was significantly lower in the H. pylori positive group (0.81 mmol/L) than in the negative group (0.95 mmol/L; p<0.001). In the genotype subgroup analyses, a similar association between H. pylori infection and HDL serum levels was seen within the IL-6 -174 CC genotype group (HDL 0.72 vs 1.06 mmol/L, respectively; p<0.001), but no significant associations were seen in the GC or GG genotype groups. Additionally, patients with peptic ulcer demonstrated lower HDL levels (0.75 mmol/L) than H. pylori positive patients without ulcer (0.86 mmol/L; p=0.010).H. pylori infection associated significantly with low serum levels of HDL in the IL-6 -174 CC genotype patients but not in the other genotypes. This suggests that the association between H. pylori infection and serum HDL could be transmitted through IL-6. We suggest that the role of IL-6 genotype should also be studied in relation to other associations between gastrointestinal microbiome and cardiovascular risk factors.
| S-EPMC4735314 | biostudies-literature
Project description:Background & aimsIdentification of a disease-specific H pylori virulence factors predictive of the outcome of infection remains unachieved.MethodsWe used the polymerase chain reaction and Southern blot to compare the presence of 14 vir homologue genes with clinical presentation of H pylori infection, mucosal histology, and mucosal interleukin (IL)-8 levels.ResultsWe examined 500 H pylori strains from East Asia and South America, including 120 with gastritis, 140 with duodenal ulcer (DU), 110 with gastric ulcer (GU), and 130 with gastric cancer. Only 1 gene that encompassed both jhp0917 and jhp0918 called dupA (duodenal ulcer promoting gene) was associated with a specific clinical outcome. dupA was present in 42% of DU vs. 21% of gastritis (adjusted odds ratio [OR] = 3.1, 95% confidence interval [CI]: 1.7-5.7). Its presence was also associated with more intense antral neutrophil infiltration and IL-8 levels and was a marker for protection against gastric atrophy, intestinal metaplasia, and gastric cancer (OR for gastric cancer = 0.42, 95% CI: 0.2-0.9 compared with gastritis). In vitro studies in gastric epithelial cells using dupA -deleted and -complemented mutants showed that the dupA plays roles in IL-8 production, in activation of transcription factors responsible for IL-8 promoter activity, and in increased survivability at low pH.ConclusionsdupA is a novel marker associated with an increased risk for DU and reduced risk for gastric atrophy and cancer. Its association with DU-promoting and -protective effects against atrophy/cancer was evident in both Asian and Western countries.
| S-EPMC3130061 | biostudies-literature
Project description:AimTo analyze the influence of the -31 C/T polymorphism of the interleukin-1β gene on Helicobacter pylori eradication therapy success in patients with functional dyspepsia.MethodsFunctional dyspepsia was diagnosed according to the Rome III criteria. All patients underwent upper gastrointestinal endoscopy, and gastric biopsies were obtained at screening and 12 months after randomization (last follow-up visit). Urease test and histological examination were performed to define the H. pylori status. Patients received twice-daily amoxicillin, clarithromycin and omeprazole for 10 days. Genotyping of the interleukin-1beta -31 C/T polymorphism (rs1143627) was performed using polymerase chain reaction-restriction fragment length polymorphism.ResultsOne hundred forty-nine patients received treatment with triple therapy for H. pylori eradication. Only one patient was lost to follow-up, and adherence to study medication was 94.6%. A total of 148 patients (mean age 46.08 ± 12.24 years; 81.8% women) were evaluated for the influence of the interleukin-1beta -31 C/T polymorphism on the outcome of H. pylori eradication therapy. After treatment, bacteria were eradicated in 87% of patients (129/148). Genotype frequencies of the polymorphism were as follows: CC, 38/148 (25.7%); CT, 71/148 (47.9%); and TT, 39/148 (26.4%). Successful eradication rate was 78.9%, 94.4% and 82.1% for the CC, CT and TT genotypes, respectively. The CT genotype was significantly associated with successful H. pylori eradication (p = 0.039).ConclusionThis study suggests that the CT genotype of the interleukin-1beta -31 C/T polymorphism plays a role in the successful eradication of H. pylori among patients with functional dyspepsia.
| S-EPMC9428014 | biostudies-literature
Project description:In this study, a Helicobacter pylori-Escherichia coli shuttle vector was constructed for transferring DNA into H. pylori. The smallest cryptic plasmid (1.2 kb), pHP489, among those harbored by 77 H. pylori isolates was selected as a base replicon for constructing vectors. HindIII-digested pHP489 was ligated with a kanamycin resistance gene [aph(3')-III], which originated from Campylobacter jejuni, to produce the recombinant plasmid pHP489K. pHP489K was efficiently transformed into and stably maintained in H. pylori strains. The shuttle vector pBHP489K (3.6 kb) was constructed by the recombination of pHP489, ColE1, and aph(3')-III sequences. pBHP489K was reciprocally transformed into and maintained in both H. pylori and E. coli. Introduction of the shuttle vector clone DNA (pBHP489K/AB; 6.7 kb), containing the ureA and ureB genes of H. pylori, into urease-negative mutants of H. pylori led to the restoration of their urease activity. The transformants were confirmed to contain the incoming plasmid DNA. pBHP489K satisfied the requirements for an H. pylori-E. coli shuttle vector, implying that it might be a useful vector for investigating pathogenicity and restriction-modification systems of H. pylori.
| S-EPMC168813 | biostudies-other