No correlation of babA2 with vacA and cagA genotypes of Helicobacter pylori and grading of gastritis from peptic ulcer disease patients in Brazil

BACKGROUND: The babA2 gene, which encodes a blood-group antigen-binding adhesin that mediates attachment of Helicobacter pylori to human Lewis(b) antigens on gastric epithelial cells, has been associated with a higher risk of peptic ulcer and gastric cancer. The purpose of this study was to ascertain the frequency of babA2 genotype in H. pylori strains of patients with peptic ulcer and to correlate with other virulence factors. MATERIALS AND METHODS: vacA, cagA, and babA2 genotypes of H. pylori were determined by using polymerase chain reaction (PCR). DNA was extracted from positive urease test gastric samples of 150 patients with peptic ulcer. Antrum and corpus biopsies were taken for histologic examination according to the updated Sydney system classification. RESULTS: babA2 genotype was present in 104 (69.3%) and cagA in 113 (75.3%) gastric samples. No significant correlation was observed between babA2 and vacAs1 genotype or between babA2 and cagA status. The correlation of vacAs1 genotype with positive cagA was statistically significant ( p < .001). The babA2-positive strain was more frequently found from the gastric samples of men, than of women (p = .01). Strains harboring cagA, vacAs1, and babA2 genotypes had no association to the grading of gastritis, presence of glandular atrophy, or intestinal metaplasia. The simultaneous presence of cagA, vacAs1, and babA2 was found in 32.6% of the H. pylori strains. CONCLUSIONS: babA2 genotype is frequently found in H. pylori strains from peptic ulcer disease in Brazil, although it has no significant correlation to the worsening of the gastritis and to other virulence markers such as vacAs1 and cagA.     

Correlation of the Helicobacter pylori adherence factor BabA with duodenal ulcer disease in four European countries

Helicobacter pylori strains harboring the vacAs1, cagA and babA2 have been associated with ulcer disease (UD). We compared the prevalence of these different genotypes and adhesive properties in H. pylori infected patients with UD in four European countries. Genomic DNA was isolated from 314 H. pylori strains: Germany (GER; n=92), Sweden (SWE, n=74), Portugal (POR, n=91) and Finland (FIN, n=57). The frequencies of babA2 genotype varied from 35% to 60%. Triple-positive strains (vacAs1+, cagA+ and babA2+) were significantly associated with UD in GER and POR and were closely correlated with UD in FIN, but not in SWE. Classification as triple-positive strains had a higher specificity for detection of UD in GER, POR and FIN than type1 or cagA+ strains. In vitro adhesion assays revealed that Swedish strains showed high adhesion properties and were thus correlated with the diagnosis of UD, although PCR detected the babA2 gene at lower frequencies and failed to show a correlation with UD. This finding appears to reflect allelic variations of the babA2 gene in SWE, although adhesive properties of the strains are retained.     

Prevalence of cagA, vacA, babA2 and iceA genes in H. pylori strains isolated from Colombian patients with functional dyspepsia

The clinical outcome of Helicobacter pylori infection has been particularly associated with virulence genotypes. These genotypes are useful as molecular markers in the identification of patients that are infected and at high risk for developing more severe gastric pathologies. Our main objective was to determine the prevalence of virulence genotypes cagA, vacA, iceA and babA2 of H. pylori, in patients with functional dyspepsia who are infected with the bacteria. H. pylori genotypes babA2 and cagA as well as vacA and iceA allelic variants were identified by PCR in 122 isolates resulting from 79 patients with functional dyspepsia. A high prevalence of genes cagA+ (71%), vacAs1am1 (34%), babA2 (57%) and iceA1 (87%) was found. The most frequent combined genotype found were cagA+/vacAs1am1/babA2+/iceA1 and cagA-/vacAs1am1/babA2+/iceA1, regardless of any family history of gastric cancer or MALT lymphoma. The very virulent genotype cagA+/vacAs1am1/babA2+/iceA1 prevailed in the studied patients with functional dyspepsia. Our results provide information about the prevalence of four of the more important virulent factors and constitute new evidence on the prevalence of the most virulent H. pylori genotype in patients with functional dyspepsia.     

淮南地区幽门螺杆菌感染个体菌株基因多态性与感染结局的影响

目的:探讨淮南地区幽门螺杆菌感染个体菌株基因多态性及其与感染结局的影响。方法:选取125例幽门螺杆菌(H.pylori,HP)感染的慢性胃炎、消化性溃疡患者,常规获取胃窦、胃体部粘膜,进行HP分离、培养,提取HP基因组DNA,采用随机扩增多态性DNA(RAPD)指纹分析法检测菌株基因多态性;125例患者均给予质子泵抑制、H2受体拮抗剂、铋剂为基础的三联或四联疗法治疗,治疗后4~6周进行14C-尿素呼气试验评估Hp根除情况;获取HP根除失败患者的胃窦、胃体黏膜进行HP分离、培养、鉴定,并采用RAPD指纹分析法检测菌株来源,评估HP基因多态性对治疗结局的影响。结果:cagA、iceA1、iceA2、vacAs1、vacAm1、babA2阳性率分别为92.80%、36.00%、93.60%、93.60%、29.50%、53.50%,cagA、iceA2、vacAs阳性率均高于其他基因类型阳性率(P0.05或P0.01),其他基因类型阳性率比较差异无统计学意义(P0.05)。经治疗后HP根除率为86.4%(107/125),14.4%(18/125)根除失败;18例根除失败患者中,15例患者治疗前后的菌株具有相同的指纹图谱,证实为原菌株复发,其中cagA、iceA1、iceA2、vacAs1、vacAm1、babA2阳性率分别为93.33%、13.33%、86.67%、93.33%、6.67%、20.00%,cagA、iceA2、vacAs阳性率均高于其他基因类型阳性率(P0.05或P0.01)。结论:cagA+、vacAs+、iceA2+为淮南地区HP感染的优势基因型,该基因型易导致HP根除失败;未发现babA2与HP感染结局存在相关性。     

Lower prevalence of Helicobacter pylori infection with vacAs1a, cagA-positive, and babA2-positive genotype in erosive reflux esophagitis disease

BACKGROUND: Increased prevalence of esophagitis has been recognized in the West. Helicobacter pylori infection, particularly virulent strains, is proposed as a protective factor against the development of gastroesophageal reflux disease. To evaluate the relationship of reflux esophagitis with virulent H. pylori infection, we studied the prevalence of reflux esophagitis among H. pylori-infected and -uninfected patients and the genotype of isolates in Taiwan. METHODS: Patients who had routine physical examination were investigated. The severity of esophagitis was evaluated using the Los Angeles grading system. H. pylori status was assessed by histology, rapid urease test, and bacterial culture. Genotyping of vacA, cagA, and babA2 was determined by polymerase chain reaction (PCR). Risk factors for severe esophagitis were evaluated. RESULTS: Reflux esophagitis was found in 21.2% of 1622 patients. The prevalence of H. pylori infection was found in 33.0% of 276 patients with reflux esophagitis compared with 67.5% of 378 patients with normal esophagus (p < .001). Esophagitis occurred in a significantly lower rate among H. pylori-positive patients with peptic ulcer than those without peptic ulcer. cagA, babA2, and vacAs1a were detected in 100% of 143 isolates. Factors that predicted severe esophagitis included age, gender, and hiatus hernia but not H. pylori infection. CONCLUSIONS: Our study suggests significantly lower incidence of H. pylori infection with the triple-positive virulent genotype in patients with reflux esophagitis in Taiwan.     

Helicobacter pylori induces apoptosis of rat gastric parietal cells

Gastric Helicobacter pylori infection may lead to multifocal atrophic corpus gastritis associated with loss of epithelial cells as well as glandular structures. The current work investigated H. pylori effects on cell death of isolated, nontransformed rat parietal cells (PC). Highly enriched rat PC (>97%) were isolated from gastric mucosa and cultured in serum-free medium over 24 h. The cells were cocultured over 8 h with cytotoxin-associated immunodominant protein (cagA)(+)/vacuolating toxin (vacA)(+) or with cagA(-)/vacA(-) H. pylori laboratory strains and also with H. pylori mutants deleted in several genes of the cag pathogenicity island. Staphylococcus aureus or Campylobacter jejuni were used as controls. Apoptosis was determined by terminal deoxynucleotidyl transferase dUTP nick-end labeling staining and electron microscopy. Interleukin (IL)-8 and cytokine-induced neutrophil chemoattractant (CINC)-1 secretion was measured by ELISA. Activation of nuclear factor-kappaB (NF-kappaB) was studied in nuclear extracts of PC by electrophoretic mobility shift assay. Apoptosis of PC was induced in a concentration- and time-dependent manner by cagA(+)/vacA(+) H. pylori strains but not by cagA(-)/vacA(-) negative strains or by the cagE knockout mutant. S. aureus and C. jejuni had no effect. PC showed no IL-8 or CINC-1 secretion on exposure to cagA(+)/vacA(+) H. pylori. cagA(+)/vacA(+) strains induced activation of NF-kappaB complexes in nuclear extracts of PC, which were composed of p65 and p50 subunits. No significant stimulation of NF-kappaB activation was detected by incubation of PC with the cagE knockout mutant. Preincubation of PC with antisense but not missense oligodeoxynucleotides against the p65 subunit significantly reduced DNA binding to the kappaB recognition sequence. The p65 oligonucleotides as well as the proteasome inhibitor N-CBZ-isoleucin-glutamin-(o-t-butyl-)-alanin-leucin and the nitric oxide synthase inhibitor N(G)-monomethyl-L-arginine completely prevented PC apoptosis induced by cagA(+)/vacA(+) strains. In summary, cagE presence appears to be essential for H. pylori-induced apoptosis of gastric parietal cells, and this effect is dependent on the activation of NF-kappaB and production of nitric oxide.     

Helicobacter pylori infection interferes with epithelial Stat6-mediated interleukin-4 signal transduction independent of cagA,cagE, or VacA

Helicobacter pylori is a bacterial pathogen evolved to chronically colonize the gastric epithelium, evade immune clearance by the host, and cause gastritis, peptic ulcers, and even gastric malignancies in some infected humans. In view of the known ability of this bacterium to manipulate gastric epithelial cell signal transduction cascades, we determined the effects of H. pylori infection on epithelial IL-4-Stat6 signal transduction. HEp-2 and MKN45 epithelial cells were infected with H. pylori strains LC11 or 8823 (type 1; cagA(+)/cagE(+)/VacA(+)), LC20 (type 2; cagA(-), cagE(-), VacA(-)), and cagA, cagE, and vacA isogenic mutants of strain 8823, with some cells receiving subsequent treatment with the Th2 cytokine IL-4, a known Stat6 activator. Immunofluorescence showed a disruption of Stat6-induced nuclear translocation by IL-4 in LC11-infected HEp-2 cells. IL-4-inducible Stat6 DNA binding in HEp-2 and MKN45 cells was abrogated by infection, but MKN45 cell viability was unaffected. A decrease in IL-4-mediated Stat6 tyrosine phosphorylation in nuclear and whole cell lysates was also observed following infection with strains LC11 and LC20, while neither strain altered IL-4 receptor chain alpha or Janus kinase 1 protein expression. Furthermore, parental strain 8823 and its isogenic cagA, cagE, and vacA mutants also suppressed IL-4-induced Stat6 tyrosine phosphorylation to comparable degrees. Thus, H. pylori did not directly activate Stat6, but blocked the IL-4-induced activation of epithelial Stat6. This may represent an evolutionarily conserved strategy to disrupt a Th2 response and evade the host immune system, allowing for successful chronic infection.     

上海市某三甲医院幽门螺杆菌耐药性和毒力与其生物膜形成能力的相关性研究

为探讨复旦大学附属华东医院(以下简称本院)分离培养的幽门螺杆菌(Helicobacter pylori,H. pylori)的耐药性、毒力和感染特征与其生物膜形成能力的相关性,收集2014年12月-2015年6月于本院消化内镜中心的胃活检组织标本及相应临床病例资料,分离培养获得幽门螺杆菌,分析菌株的耐药性、毒力基因型、临床病例特征。结果显示,从胃活检组织样本中共分离培养28株幽门螺杆菌,对左氧氟沙星(levofloxacin,LEV)、甲硝唑(metronidazole,MTZ)和克拉霉素(clarithromycin,CLA)的耐药率分别为32%、75%和11%,未发现阿莫西林(amoxicillin,AMX)耐药。单一药物耐药17株(17/28,61%),双重耐药10株(10/28,36%)。毒力基因cagA、oipA和vacAs1检出率为100%,未检出vacAs2。基因型vacAs1m1占39%(11/28),vacAs1m2占61%(17/28);iceA1占54%(15/28),iceA2占21%(6/28),iceA1A2占25%(7/28);dupA^＋占36%(10/28)。28株菌株均能形成生物膜,但能力不尽相同。单因素及独立样本t检验分析显示,45～59岁、iceA1^＋dupA^－基因型和甲硝唑敏感菌株形成生物膜的能力较强。结果提示,本院分离的幽门螺杆菌对甲硝唑耐药率最高,双重耐药不容忽视。菌株主要毒力基因型为cagA、oipA、vacAs1m2。幽门螺杆菌的生物膜形成能力与患者年龄有关,45～59岁组较强;携带毒力基因iceA1的菌株生物膜形成能力强;dupA基因型及甲硝唑耐药与菌株生物膜形成呈负相关。     

幽门螺杆菌毒力基因型与胃癌的相关性研究

目的:研究中国东部地区幽门螺杆菌(H.pylori)cagA、vacA和iceA1毒力基因型的分布状况及其与胃癌的相关性。方法:从52例病人胃黏膜活检组织(31例慢性浅表性胃炎,21例胃癌)中分离培养H.pylori,用PCR方法扩增分离菌株的cagA、iceA1、vacAs,i,m区毒力基因片段,统计并分析上述毒力因素与胃癌发生的相关性。结果:在52例菌株中,cagA、vacAs1/i1/m1、vacAs1/i1/m2和iceA1的阳性率分别是92.3%(48/52),48.1%(25/52),48.1%(25/52),90.4%(47/52);所有的胃癌分离株中均为cagA(+)vacAs1/i1(+)型,vacAm1、vacAm2和iceA1在胃癌组中的阳性率分别是47.6%(10/21),52.4%(11/21),95.2%(20/21),与胃炎组相比,上述基因型的阳性率差异均无显著统计学意义(P>0.05)。结论:cagA、vacAs1/i1/m1、vacAs1/i1/m2和iceA1是中国东部地区H.pylori的优势基因型;cagA、vacAs1、vacAi1和iceA1毒力基因型的存在与胃癌的发生无相关性。     

Redefinition of the Carbohydrate Binding Specificity of Helicobacter pylori BabA Adhesin

Certain Helicobacter pylori strains adhere to the human gastric epithelium using the blood group antigen-binding adhesin (BabA). All BabA-expressing H. pylori strains bind to the blood group O determinants on type 1 core chains, i.e. to the Lewis b antigen (Fucα2Galβ3(Fucα4)GlcNAc; Le(b)) and the H type 1 determinant (Fucα2Galβ3GlcNAc). Recently, BabA strains have been categorized into those recognizing only Le(b) and H type 1 determinants (designated specialist strains) and those that also bind to A and B type 1 determinants (designated generalist strains). Here, the structural requirements for carbohydrate recognition by generalist and specialist BabA were further explored by binding of these types of strains to a panel of different glycosphingolipids. Three glycosphingolipids recognized by both specialist and generalist BabA were isolated from the small intestine of a blood group O pig and characterized by mass spectrometry and proton NMR as H type 1 pentaglycosylceramide (Fucα2Galβ3GlcNAcβ3Galβ4Glcβ1Cer), Globo H hexaglycosylceramide (Fucα2Galβ3GalNAcβ3Galα4Galβ4Glcβ1Cer), and a mixture of three complex glycosphingolipids (Fucα2Galβ4GlcNAcβ6(Fucα2Galβ3GlcNAcβ3)Galβ3GlcNAcβ3Galβ4Glcβ1Cer, Fucα2Galβ3GlcNAcβ6(Fucα2Galβ3GlcNAcβ3)Galβ3GlcNAcβ3Galβ4Glcβ1Cer, and Fucα2Galβ4(Fucα3)GlcNAcβ6(Fucα2Galβ3GlcNAcβ3)Galβ3GlcNAcβ3Galβ4Glcβ1Cer). In addition to the binding of both strains to the Globo H hexaglycosylceramide, i.e. a blood group O determinant on a type 4 core chain, the generalist strain bound to the Globo A heptaglycosylceramide (GalNAcα3(Fucα2)Galβ3GalNAcβ3Galα4Galβ4Glcβ1Cer), i.e. a blood group A determinant on a type 4 core chain. The binding of BabA to the two sets of isoreceptors is due to conformational similarities of the terminal disaccharides of H type 1 and Globo H and of the terminal trisaccharides of A type 1 and Globo A.     

Clinical and pathological importance of vacA allele heterogeneity and cagA status in peptic ulcer disease in patients from North Brazil

We have examined the prevalence of gene cagA and vacA alleles in 129 patients, 69 with gastritis and 60 with peptic ulcer diseases from North Brazil and their relation with histopathological data. vacA and cagA genotype were determined by polymerase chain reaction. Hematoxylin-eosin staining was used for histological diagnosis. 96.6% of the patients were colonized by Helicobacter pylori strains harboring single vacA genotype (nont-mixed infection). Among them, 11.8% had subtype s1a, 67.8% had subtype s1b, and 17% subtype s2. In regard to the middle region analysis, m1 alleles were found in 75.4% and m2 in 21.2% of patients. The cagA gene was detected in 78% patients infected with H. pylori and was associated with the s1-m1 vacA genotype. The H. pylori strains, vacA s1b m1/cagA-positive, were associated with increased risk of peptic ulcer disease and higher amounts of lymphocytic and neutrophilic infiltrates and the presence of intestinal metaplasia. These findings show that cagA and vacA genotyping may have clinical relevance in Brazil.     

Prevalence of Helicobacter pylori vacA, cagA, cagE, iceA, babA2 genotypes and correlation with clinical outcome in Turkish patients with dyspepsia

BACKGROUND: Distinct virulence factors of Helicobacter pylori have been associated with clinical outcome of the infection; however, considerable variations have been reported from different geographic regions and data on genotypes of Turkish H. pylori isolates are sparse. AIM: To determine the prevalence of specific genotypes of H. pylori in Turkish patients with dyspepsia. MATERIALS AND METHODS: Ninety-three H. pylori-positive patients [30 with non-ulcer dyspepsia (NUD), 30 with duodenal ulcer (DU), and 33 with gastric cancer (GC)] who were admitted to our endoscopy unit due to dyspepsia were enrolled in the study. H. pylori infection was confirmed in all patients by histology and rapid urease test (RUT). The presence of vacA alleles, cagA, cagE, iceA, and babA2 genotypes were determined by polymerase chain reaction (PCR). Chi-squared test and Fisher's exact test were used for statistical comparisons and multivariate regression analysis was performed to find out independent predictors of different clinical outcomes. RESULTS: Turkish strains examined predominantly possessed the vacA s1,m2 (48.4%) and s1,m1 (40.7%) genotypes. The vacA s1a genotype was detected in 66.7, 96.4, and 87.9% of isolates from patients with NUD, DU, and GC, respectively, and its presence was significantly associated with that of DU (p = .004), GC (p = .043), and cagA gene (p = .021). None of the cases was found to harbor the s1c genotype. The frequencies of the cagA and cagE genes among studied isolates were 73.6 and 59.3%, respectively. The cagA gene was significantly associated with the presence of DU (p = .004) and GC (p = .003), and the cagE gene, too, was significantly associated with the presence of DU (p = .002) and GC (p = .000). All H. pylori isolates possessed the iceA gene. In all, 68 isolates (74.7%) were positive for iceA1 and 23 (25.3%) for iceA2. The frequency of icea1 gene was significantly higher in cases with GC (85%) than in cases with NUD (60%) (p = .026). The frequency of babA2 gene was 23.3, 46.4, and 87.9% in isolates of patients with NUD, DU, and GC, respectively. When compared to cases with NUD (p = .000) and DU (p = .000), the presence of babA2 gene was significantly higher in cases with GC. Multivariate regression analysis disclosed cagE (p = .006) and vacA s1a (p = .027) genotypes to be independent predictors of DU and babA2 (p = .000) and cagE (p = .013) genotypes to be independent predictors of GC. CONCLUSIONS: H. pylori vacA s1a, cagA, cagE genotypes have significant relations with the presence of DU and GC, and iceA1, babA2 with GC in Turkish patients with dyspepsia, whereas cagE and vacA s1a genotypes are independent predictors of DU, and babA2 and cagE genotypes are independent predictors of GC.     

Human dendritic cells respond to Helicobacter pylori, promoting NK cell and Th1-effector responses in vitro

Helicobacter pylori infection leads to chronic gastric inflammation. The current study determined the response of human APCs, NK cells, and T cells toward the bacteria in vitro. Human monocyte-derived dendritic cells (DC) were incubated with bacteria for 48 h. Intact H. pylori at a multitude of infection 5 stimulated the expression of MHC class II (4- to 7-fold), CD80, and CD86 B7 molecules (10- to 12-fold) and the CD83 costimulatory molecule (>30-fold) as well as IL-12 secretion (>50-fold) in DCs, and thereby, strongly induced their maturation and activation. CD56(+)/CD4(-) NK cells, as well as CD4(+)/CD45RA(+) naive T cells, were isolated and incubated with DCs pulsed with intact bacteria or different cellular fractions. Coculture of H. pylori-pulsed DCs with NK cells strongly potentiated the secretion of TNF-alpha and IFN-gamma. Coculture of naive T cells with H. pylori-pulsed DCs significantly enhanced TNF-alpha, IFN-gamma, and IL-2 secretion as well as T-bet mRNA levels, while GATA-3 mRNA was lowered. However, the effect appeared attenuated compared with coculture with Escherichia coli. A greater stimulation was seen with naive T cells and DCs pulsed with H. pylori membrane preparations. Intact H. pylori potently induced the maturation and activation of human monocyte-derived DC and thereby promote NK and Th1 effector responses. The strong activation of NK cells may be important for the innate immune response. Th1-polarized T cells were induced especially by incubation with membrane preparations of H. pylori, suggesting that membrane proteins may account for the specific adaptive immune response.     

A simple multiplex PCR assay for diagnosing virulent Helicobacter pylori infection in human gastric biopsy specimens from subjects with gastric carcinoma and other gastro-duodenal diseases

AIM: To evaluate and develop a multiplex polymerase chain reaction (PCR) assay for diagnosing and specific identification of virulent Helicobacter pylori strains and their main virulence genes cagA, cagE, cagT, vacA and hrgA. METHODS AND RESULTS: Genomic DNA from 82 gastric tissues was screened. A master pool of all the ingredients of multiplex reaction was prepared for amplification. Amplicons were sequenced to confirm the amplification of each target genes. Multiplex PCR assay was able to detect all the five target genes in 81.7% and deletions in one or more loci among 18.3%. Genotype cagT +ve/hrgA +ve/cagA +ve/cagE +ve/vacAs1 +ve was more predominant in this study population (67.07%). hrgA, cagT, cagE and cagA genes were present in 100%, 92.7%, 85.4% and 81.7% of the subjects, respectively. The vacAs1 subtype had higher prevalence frequency in patients with overt gastrointestinal disease (78.57%) than with GERD (gastro-esophageal reflux disease) and NUD (non-ulcer dispepsia) (50%). CONCLUSIONS: The multiplex PCR assay developed herein was able to genotype H. pylori isolates based on the main virulence genes. SIGNIFICANCE AND IMPACT OF THE STUDY: The ability to identify H. pylori and the majority of their virulence gene markers by multiplex PCR assay represents a considerable advancement over other PCR-based methods for genotyping H. pylori from large population, and can be explored to gain insights at the genotypic variability exhibited by this pathogen.     

Distinctiveness of genotypes of Helicobacter pylori in Calcutta, India

The genotypes of 78 strains of Helicobacter pylori from Calcutta, India (55 from ulcer patients and 23 from more-benign infections), were studied, with a focus on putative virulence genes and neutral DNA markers that were likely to be phylogenetically informative. PCR tests indicated that 80 to 90% of Calcutta strains carried the cag pathogenicity island (PAI) and potentially toxigenic vacAs1 alleles of the vacuolating cytotoxin gene (vacA), independent of disease status. This was higher than in the West (where cag PAI(+) vacAs1 genotypes are disease associated) but lower than in east Asia. The iceA2 gene was weakly disease associated in Calcutta, whereas in the West the alternative but unrelated iceA1 gene at the same locus is weakly disease associated. DNA sequence motifs of vacAm1 (middle region) alleles formed a cluster that was distinct from those of east Asia and the West, whereas the cagA sequences of Calcutta and Western strains were closely related. An internal deletion found in 20% of Calcutta iceA1 genes was not seen in any of approximately 200 strains studied from other geographic regions and thus seemed to be unique to this H. pylori population. Two mobile DNAs that were rare in east Asian strains were also common in Calcutta. About 90% of Calcutta strains were metronidazole resistant. These findings support the idea that H. pylori gene pools differ regionally and emphasize the potential importance of studies of Indian and other non-Western H. pylori populations in developing a global understanding of this gastric pathogen and associated disease.     

Helicobacter pylori-induced mucosal inflammation is Th1 mediated and exacerbated in IL-4, but not IFN-gamma, gene-deficient mice

To elucidate the pathogenesis of Helicobacter pylori-associated gastritis, we studied immune responses of C57BL/6J wild-type (WT), SCID, and gene deficient (IFN-gamma-/- and IL-4-/-) mice following infection with a pathogenic isolate of H. pylori (SPM326). During early infection in WT mice, mononuclear and polymorphonuclear cells accumulated in the gastric lamina propria, and the numbers of cells in the inflamed mucosa expressing IFN-gamma, but not IL-4, mRNA rose significantly (p < 0.005), consistent with a local Th1 response. Splenic T cells from the same infected WT mice produced high levels of IFN-gamma, no detectable IL-4, and low amounts of IL-10 following in vitro H. pylori urease stimulation, reflecting a systemic Th1 response. Infected C57BL/6J SCID mice did not develop gastric inflammation despite colonization by many bacteria. Infected C57BL/10J and BALB/c mice also did not develop gastric inflammation and displayed a mixed Th1/Th2 splenic cytokine profile. These data imply a major role for the Th1 cytokine IFN-gamma in H. pylori-associated gastric inflammation in C57BL/6J mice. Compared with WT animals, infected IL-4-/- animals had more severe gastritis and higher levels of IFN-gamma production by urease-stimulated splenocytes (p < 0.01), whereas IFN-gamma-/- mice exhibited no gastric inflammation and higher levels of IL-4 production by stimulated splenocytes. These findings establish C57BL/6J mice as an important model for H. pylori infection and demonstrate that up-regulated production of IFN-gamma, in the absence of the opposing effects of IL-4 (and possibly IL-10), plays a pivotal role in promoting H. pylori-induced mucosal inflammation.     

Helicobacter pylori picB, a homologue of the Bordetella pertussis toxin secretion protein, is required for induction of IL-8 in gastric epithelial cells

Approximately 60% of Helicobacter pylori strains are cagA ⁺ and this genotype is more frequently associated with duodenal ulcer disease. Although most wild-type cagA ⁺ strains are both cytotoxigenic and induce enhanced Interleukin-8 (IL-8) secretion in gastric epithelial cells, isogenic cagA ⁻ mutants retain full activity in these assays; thus, cagA appears to be a marker of enhanced virulence. Delineation of the nucleotide sequence of a 4 kb region upstream of cagA allowed the identification of 966 bp ( picA ) and 2655 bp ( picB ) open reading frames encoding 36 kDa and 101 kDa polypeptides, respectively. picA and picB constitute an operon in opposite orientation to cagA . The deduced picB product showed significant homology (26% identity and 50% similarity) with the Bordetella pertussis toxin secretion protein (PtlC). Of 55 H. pylori clinical isolates, the picA and picB segment was conserved exclusively in cagA ⁺ strains and present in all isolates from patients with duodenal ulceration, versus 59% of isolates from patients with gastritis alone ( P =0.01). Using gene-replacement techniques, we constructed picA and picB mutant H. pylori strains and demonstrated that the picB gene product is involved in the induction of IL-8 expression in gastric epithelial cells. Further, Northern blot hybridization and RT-PCR data showed that picA and picB are co-transcribed and an insertional mutation in picA ablates picB expression. These studies indicate a role of picA and picB in the induction of an inflammatory response in gastric epithelial cells either directly or by enabling secretion of an unidentified product, and suggest a mechanism for the overrepresentation of strains possessing these genes in patients with peptic ulceration.     

The role of CagA status in gastric and extragastric complications of Helicobacter pylori.

Two major markers of virulence have been described in H. pylori. The first is a secreted protein (VacA) that is toxic to human cells in tissue culture. This cytotoxin causes vacuolation of epithelial cells in vitro and induces epithelial cell damage in mice. The second is a 40-Kb pathogenicity island for which the gene cagA (cytotoxin-associated gene A) is a marker. Approximately 60% of H. pylori isolates in Western countries are cagA+. The protein encoded by cagA+ has a molecular weight of 120-140 kDa and exhibits sequence heterogeneity among strains isolated from Western and Eastern countries. Although no specific function has been identified for CagA, there is increasing evidence that cagA+ strains are associated with increased intensity of gastric inflammation and increased mucosal concentration of particular cytokines including interleukin 8. Inactivation of picB (Hp 0544) or any of several other genes in the cag island ablates the enhanced IL-8 secretion of human gastric epithelial cells in tissue culture. Furthermore, persons colonized with cagA+ strains have an increased risk of developing more severe gastric diseases such as peptic ulcer and distal (non-cardia) gastric cancer than those harboring cagA- strains. We investigated the role of cagA status in both gastroduodenal and extragastroduodenal disease with H. pylori. Among the diseases limited to the antrum and body of the stomach and the duodenum, we demonstrated a correlation between CagA seropositivity and peptic ulcer disease. We also showed correlation between distal gastric cancer rated and CagA prevalence in populations in both developed and developing countries. In addition, we found that for several Asian populations, the relationship between CagA seropositivity and gastroduodenal diseases was complex. For extragastroduodenal diseases, our results confirmed previous reports that demonstrated that CagA status did not play a role in diseases such as rheumatoid arthritis and hyperemesis gravidarum. However, we found a clear negative association between the presence of a positive response to CagA and esophageal diseases. Therefore, CagA seropositivity (and thus gastric carriage) is associated with increased risks of certain diseases (involving the lower stomach and duodenum) and decreased risks of GERD and its sequelae. This apparent paradox can best be explained by differences in the interaction of cagA+ and cagA- strains with their hosts.