Development of an in-house capture ELISA: An attempt to detect CagA antigen in sera of Helicobacter pylori infected patients.

The CagA protein one of the key virulence factors of Helicobacter pylori plays an important role in the pathogenesis of peptic ulcer diseases. Unfortunately the cagA gene status can only be determined by PCR while serology is an alternative approach to detect antigens or antibodies. Our aim is to detect the CagA antigen in sera of infected subjects by the development of an in-house capture ELISA test. Gastric antral biopsies and serum samples were collected from 63 patients. PCR was used to determine the cagA status. Our previously developed recombinant CagA protein and monoclonal antibody were used for setting up the capture ELISA test. H. pylori positive [(38 gastritis, 14 duodenal ulcers (DU), 11 gastric ulcer (GU)] patients were determined by PCR. The cagA gene was detected in 21 (55%) of gastritis, 11 (78%) of DU and 7 (60%) of GU patients. The reagents used in setting up the capture ELISA test following optimization displayed high performance. This study showed that our developed in-house capture ELISA has the potential to detect the CagA antigen at very low concentrations even though not detected in our H. pylori infected patients sera but we are also intended to use it in saliva and stool samples.

Cloning, expression and characterization of recombinant CagA protein of Helicobacter pylori using monoclonal antibodies: Its potential in diagnostics.

Helicobacter pylori CagA protein plays an important role in the severity of the gastric diseases. Our aims were to clone the cagA 5'- conserved region of the gene, characterize the recombinant CagA (rCagA) protein by monoclonal antibodies (mAbs) and to use this protein for the detection of anti-CagA antibodies by an ELISA test. Our developed rCagA protein (67 kDa) showed an amino acid sequence homology of 83% and 80% with Western and East Asian type strains respectively. Two anti-rCagA (BS-53, CK-02) mAbs and 2 additional rCagA proteins of smaller sizes (60 kDa, 28 kDa) were developed for epitope determination. The BS-53 mAb recognized all 3 rCagA proteins while CK-02 mAb recognized only 2 of them indicating recognition of different epitopes. An in-house indirect ELISA using rCagA was developed to detect anti-CagA antibodies in sera of 59 patients. The ELISA results obtained when compared to those of the PCR gave a sensitivity, specificity and accuracy of 81%, 100% and 88% respectively. We have developed for the first time: a rCagA protein that showed high sequence homology with both Western and East Asian type strains and an indirect ELISA of high performance which can be used to detect anti-CagA antibodies in sera of infected patients worldwide.

Evaluation of immobilized metal affinity chromatography kits for the purification of histidine-tagged recombinant CagA protein.

Immobilized metal affinity chromatography (IMAC) technique is used for fast and reliable purification of histidine(His)-tagged recombinant proteins. The technique provides purification under native and denaturing conditions. The aim of this study is to evaluate three commercially available IMAC kits (Thermo Scientific, GE Healthcare and Qiagen) for the purification of a 6xHis-tagged recombinant CagA (cytotoxin-associated gene A) protein from IPTG-induced Escherichia coli BL21(DE3) culture. The kits were tested according to the manufacturer instructions and the protein was purified with only GE Healthcare and Qiagen kits under denaturing conditions. 1% (w/v) SDS was used as denaturing agent in PBS instead of extraction reagent of Thermo Scientific kit to lyse bacterial cells from 100ml culture. The 6xHis-tagged recombinant protein was purified by the three kits equally.

Evaluation of the effect of cagPAI genes of Helicobacter pylori on AGS epithelial cell morphology and IL-8 secretion.

Helicobacter pylori cagPAI genes play an important role in pathogenesis, however little is known about their functions in isolates from Turkish patients. We aimed to evaluate the intactness and the effect of the cagPAI genes (cagT, cagM, cagE, cagA) and cagA EPIYA motifs on the AGS morphological changes and IL-8 induction. Of 53 patients 38 were found infected with H. pylori. PCR amplification of the cagPAI genes showed 42.1 % intact, 39.5 % partially deleted and 18.4 % with complete deletions. Isolates from gastritis, duodenal and gastric ulcer patients with intact and partially deleted cagPAI genes induced higher IL-8 secretion than those with complete deletions. Isolates from gastritis patients had higher deletion frequencies of the cagT and cagM genes than the other two genes. Infection of AGS cells with isolates that possess intact cagPAI and EPIYA-ABC resulted in the formation of the hummingbird phenotype. The cagA positive isolates induced higher IL-8 secretion than cagA negative isolates. Isolates from DU patients with more than one EPIYA-C motif induced higher concentrations of IL-8 than those with EPIYA-ABC. In conclusion, the intactness of the cagPAI in our isolates from different patients was not conserved. An intact cagPAI was found to play an important role in the pathogenesis of DU but not GU or gastritis. The cagA gene, but not other cagPAI genes, was associated with the induction of IL-8 and the morphological changes of the AGS cells. An increase in the number of EPIYA-C motifs had noticeable effect on the formation of the hummingbird phenotype.

Phylogenetic analysis of Helicobacter pylori cagA gene of Turkish isolates and the association with gastric pathology.

BACKGROUND: The cagA gene is one of the important virulence factors of Helicobacter pylori. The diversity of cagA 5' conserved region is thought to reflect the phylogenetic relationships between different H. pylori isolates and their association with peptic ulceration. Significant geographical differences among isolates have been reported. The aim of this study is to compare Turkish H. pylori isolates with isolates from different geographical locations and to correlate the association with peptic ulceration. METHODS: Total of 52 isolates of which 19 were Turkish and 33 from other geographic locations were studied. Gastric antral biopsies collected from 19 Turkish patients (Gastritis = 12, ulcer = 7) were used to amplify the cagA 5' region by PCR then followed by DNA sequencing. RESULTS: The phylogenetic tree displayed 3 groups: A) a mix of 2 sub-groups "Asian" and "African/Anatolian/Asian/European", B) "Anatolian/European" and C) "American-Indian". Turkish H. pylori isolates clustered in the mixed sub-group A were mostly from gastritis patients while those clustered in group B were from peptic ulcer patients. A phylogenetic tree constructed for our Turkish isolates detected distinctive features among those from gastritis and ulcer patients. We have found that 2/3 of the gastritis isolates were clustered alone while 1/3 was clustered together with the ulcer isolates. Several amino acids were found to be shared between the later groups but not with the first group of gastritis. CONCLUSIONS: This study provided an additional insight into the profile of our cagA gene which implies a relationship in geographic locations of the isolates.

A study on the effect of Helicobacter pylori infection on p53 expression in gastric cancer and gastritis tissues.

INTRODUCTION: Helicobacter pylori cause damage to gastric epithelial cells and alterations in the p53 gene that lead to cancer development. This study aimed to determine the correlation of p53 expression with H. pylori using immunohistochemistry, RFLP-PCR, and histopathology. METHODOLOGY: Gastric biopsy samples from gastric cancer (GC) (n = 54) and gastritis (n = 31) patients were examined for histopathological changes and expression of p53 protein by immunohistochemistry. RESULTS: Immunohistochemical analysis of p53 protein expression in H. pylori-positive GC sections showed an average of 44.3% positive cells in tumors and 6.9% in normal tissues, as compared to 16.4% and 4.4% in H. pylori-negative sections. P53 expression showed significant association with H. pylori (P = 0.005), invasion depth (P = 0.029) and inflammation reaction (P = 0.008). In gastritis sections, no difference in the average p53 staining in H. pylori-positive or -negative sections was seen. PCR-RFLP results also showed no difference in genotype frequencies of p53 in H. pylori-positive or -negative gastritis sections. Histopathology study of H. pylori-positive GC sections showed that 97.2% were the intestinal type and 2.8% the diffuse type, while in H. pylori-negative sections 35.2% were the intestinal type and 64.8% the diffuse type. Biopsy sections from H. pylori-positive gastritis patients revealed more severe inflammation than those of H. pylori-negative patients. CONCLUSION: Our results show that H. pylori infection affects p53 expression in GC. The average p53 expression was significantly higher in tumor than in normal tissues. In gastritis sections p53 expression was significantly associated with H. pylori.

Comparison of the lateral flow immunoassays (LFIA) for the diagnosis of Helicobacter pylori infection.

Helicobacter pylori infection is the most common human infection where approximately 50% of the world populations are infected. The diagnosis of such infection is mainly done by endoscopy where gastric biopsies are examined for the presence of H. pylori. Such invasive approach is costly, time consuming and generally requires more than one test to confirm the infection. Serology on the other hand is a non-invasive approach that can detect H. pylori exposure. The lateral flow immunoassays (LFIA) support the serological approach and have the advantage of being fast, economic and require no additional equipment or experience. In this review the principles, components of the LFIA, sensitivities and specificities of the commercially available H. pylori test strips were compared and discussed.

DNA sequence analysis of cagA 3' motifs of Helicobacter pylori strains from patients with peptic ulcer diseases.

The Helicobacter pylori cagA gene is a major virulence factor that plays an important role in gastric pathologies. DNA sequence data for the cagA 3' region of Western isolates differ markedly in their EPIYA motifs from those of East Asian isolates. An increase in the number of these motifs is known to be associated with gastric cancer. Whether such an association is also the case for peptic ulceration was investigated in this study. Gastric biopsies were collected from 96 patients with duodenal ulcer (DU), gastric ulcer (GU) and gastritis. The types of EPIYA motif detected by PCR among 28 DU strains were 13 ABC, eight ABCC, six ABCCC, and in one patient both ABC and ABCCCCC; among nine GU strains were two ABC, five ABCC and two ABCCC; and among 40 gastritis strains were 35 ABC and five ABCC. DNA sequencing was carried out to confirm the detection of the EPIYA motif types and to analyse their peptide sequences. A significant association was found between the number of the EPIYA-C motifs (>or=2) and peptic ulceration (P=0.00001) compared with gastritis. In conclusion, this study shows that our patients harboured cagA-positive H. pylori strains with EPIYA motifs of the Western type and that the increase in the number of EPIYA-C motifs was significantly associated with DU and GU but not with gastritis, indicating predictive association with the severity of the disease.

Helicobacter pylori infection in developing countries: the burden for how long?

Approximately 50% (over 3 billion) of the world populations are known to be infected with Helicobacter pylori , mainly in the developing countries . Among those, hundreds of millions of people develop peptic ulceration during their lifetime and still tens of millions might progress to gastric cancer. Possible modes of H. pylori transmission generally described are through direct contact between family members and also through contaminated water and food. Because the high prevalence of infection occurs mainly in developing countries and because the test-and-treat strategy puts a huge economic burden on many of these countries, it is time to take an immediate action toward this bacterial infection and adopt a strategy to prevent it. To address this issue, an updated prevalence of infection, modes of transmission, economics of infection and preventative measures to block the infection process have been discussed.

Helicobacter pylori DNA in gallbladder tissue of patients with cholelithiasis and cholecystitis.

BACKGROUND: Earlier reports on the detection of H. pylori DNA in gallbladder tissue of patients with cholelithiasis and cholecystitis gave discordant results. Our aim was to detect the presence of H. pylori DNA and to determine whether a correlation can be established with the biliary diseases. METHODOLOGY: The study included a total of 68 patients 20 to 79 years of age. Fifty-three of the participants were females, of whom 33 had cholelithiasis and 20 did not. Out of the 15 male patients, 8 were had cholelithiasis and 7 did not. Gallbladder tissue specimens were taken from all patients undergoing cholecystectomy and processed immediately for histology, culture and PCR. RESULTS: Histological examination revealed that 36 (68%) of 53 females and 9 (60%) of 15 males had cholecystitis. PCR results detected H. pylori DNA in 15 (22%) of 68 samples but no bacteria were isolated in culture. The presence of few bacteria, the geographic distribution of H. pylori strains, and the bile milieu inhibitory effect might be some of the reasons for growth failure. CONCLUSIONS: In this study although H. pylori DNA was detected by PCR in gallbladder tissue of patients with cholecystitis, a clinical correlation with biliary disease could not be established because several conditions were difficult to meet as discussed in the text.

H pylori infection and other risk factors associated with peptic ulcers in Turkish patients: a retrospective study.

AIM: To identify and evaluate the relative impact of H pylori infection and other risk factors on the occurrence of gastric ulcer (GU), duodenal ulcer (DU) and gastritis in Turkish patients. METHODS: A total of 4471 patients (48.3% female) out of 4863 attended the Samatya hospital in Istanbul (June 1999-October 2003) were included. The records of H pylori status (CLO-test), endoscopic findings of GU, DU and gastritis, personal habits (smoking, alcohol intake) and medication [non-steroidal anti-inflammatory drugs (NSAIDs), aspirin intake] were analyzed using multi-way frequency analysis. RESULTS: We have found that GU in the presence of H pylori had significant association with aspirin (P=0.0001), alcohol (P=0.0090) and NSAIDs (P=0.0372). DU on the other hand had significant association with aspirin/smoking/NSAIDs (P=0.0259), aspirin/alcohol (P=0.0002) and aspirin/smoking (P=0.0233), also in the presence of H pylori. In the absence of H pylori GU had significant association with alcohol/NSAIDs (P=0.0431), and NSAIDs (P=0.0436). While DU in the absence of H pylori had significant association with smoking/alcohol/ NSAIDs (P=0.0013), aspirin/NSAIDs (P=0.0334), aspirin/alcohol (P=0.0360). CONCLUSION: In the presence of H pylori, aspirin, alcohol and NSAIDs intake act as an independent risk factors that had an augmenting impact on the occurrence of GU and only together on the occurrence of DU in Turkish patients.

A preliminary study on the genetic profile of cag pathogenicity-island and other virulent gene loci of Helicobacter pylori strains from Turkey.

Helicobacter pylori genetic diversity affects the function and antigenicity of virulence factors associated with the disease outcome. Gene profile was done to identify the distribution of gene loci within and outside the cag pathogenicity-island (PAI). H. pylori strains from 35 patients [21 gastritis, 14 peptic ulcer diseases (PUD)] were analyzed using PCR. The profile of the cag PAI was evaluated using primers spanning the 3' end, cagA, promoter region of the cagA, cagE, cagT, 5' end (LEC), extreme right end, plasticity region open reading frames (ORFs), oipA (Hp0638) and vacA alleles. We found few intact cag PAI in the strains examined. Deletions were found in LEC1 (9.5% versus 14.3%), LEC2 (4.8% versus 14.3%), cagT (33.3% versus 28.6%), cagE (28.6% versus 28.6%) and the promoter region of the cagA (19.0% versus 42.9%) of gastritis and PUD strains, respectively. The cagA gene was detectable in 57.1% of gastritis and 92.9% of PUD-associated strains. The cagRJ region also showed deletions for many of its genes. The oipA (Hp0638) gene was detected in 80.9% of gastritis and in 92.9% of PUD strains. The plasticity region ORFs JHP912 and JHP931 were predominant in PUD strains. The vacA-s1a-m1a genotype was predominant in PUD, while s2m2 in gastritis strains. This comprehensive analysis showed deletions in several genes within and outside the cag PAI. However, cagA, oipA, JHP912, JHP931 and vacA-s1a-m1a were more predominant in PUD strains than gastritis-associated strains, suggesting the importance of genetic diversity on the disease progression and clinical outcome.

Genotyping of Helicobacter pylori strains from gastric biopsies by multiplex polymerase chain reaction. How advantageous is it?

Recent application of multiplex polymerase chain reaction (PCR) for genotyping Helicobacter pylori direct from biopsies revealed variable results (detection of amplicons from DNA extracted by boiling biopsies, variable size amplicons and deletions, uniform intensity of amplicon bands). We aimed to look at how applicable the technique is for determining cagA and vacA genotypes and to correlate the results with the severity of the disease. H. pylori strains from 52 patients (35 duodenal ulcers [DUs], 7 gastric ulcers [GUs], 10 gastritis) were included. Three antral biopsies were obtained for Campylobacter-like organism (CLO) and PCR. Primers for cagA, vacA s1s2, and m1m2 alleles were used. No PCR amplicons were detected from boiling biopsies; thus, DNA was extracted by QIAamp kit. H. pylori was positive in 84.6% of the patients (85.7% DU, 100% GU, and 70% gastritis). The cagA gene was detected in 86.6% DU, 71.4% GU, and 57.0% gastritis patients. The vacA allelic distribution among cagA-positive strains was 80.7% s1m1 in DU and 60.0% in GU patients, whereas 75.0% of gastritis had s1m2. No variability in the amplicon sizes was found, and the intensity of the amplicon bands was not uniform. A deleted band of approximately 420 bp below the m1 band was detected in strains from 2 DU and 1 GU patients. Although the multiplex PCR is a rapid and an effective tool for detecting several genes in a single-step system, one has to adjust for optimization of the technique when genotyping H. pylori direct from biopsies. A significant association was found between the cagA-positive vacA-s1m1 genotype and peptic ulcers.

Canonical correlation analysis of factors involved in the occurrence of peptic ulcers.

The impact of risk factors on the development of peptic ulcers has been shown to vary among different populations. We sought to establish a correlation between these factors and their involvement in the occurrence of peptic ulcers for which a canonical correlation analysis was applied. We included 7,014 patient records (48.6% women, 18.4% duodenal ulcer [DU], 4.6% gastric ulcer [GU]) of those underwent upper gastroendoscopy for the last 5 years. The variables measured are endoscopic findings (DU, GU, antral gastritis, erosive gastritis, pangastritis, pyloric deformity, bulbar deformity, bleeding, atrophy, Barret esophagus and gastric polyp) and risk factors (age, gender, Helicobacter pylori infection, smoking, alcohol, and nonsteroidal anti-inflammatory drugs [NSAIDs] and aspirin intake). We found that DU had significant positive correlation with bulbar deformity (P=2.6 x 10(-23)), pyloric deformity (P=2.6 x 10(-23)), gender (P=2.6 x 10(-23)), H. pylori (P=1.4 x 10(-15)), bleeding (P=6.9 x 10(-15)), smoking (P=1.4 x 10(-7)), aspirin use (P=1.1 x 10(-4)), alcohol intake (P=7.7 x 10(-4)), and NSAIDs (P=.01). GU had a significantly positive correlation with pyloric deformity (P=1,6 x 10(-15)), age (P=2.6 x 10(-14)), bleeding (P=3.7 x 10(-8)), gender (P=1.3 x 10(-7)), aspirin use (P=1.1 x 10(-6)), bulbar deformity (P=7.4 x 10(-4)), alcohol intake (P=.03), smoking (P=.04), and Barret esophagus (P=.03). The level of significance was much higher in some variables with DU than with GU and the correlations with GU in spite of being highly significant the majority, were small in magnitude. In conclusion, Turkish patients with the following endoscopic findings bulbar deformity and pyloric deformity are high-risk patients for peptic ulcers with the risk of the occurrence of DU being higher than that of GU. Factors such as H. pylori, smoking, alcohol use, and NSAIDs use (listed in a decreasing manner) are risk factors that have significant impact on the occurrence of DU; aspirin has a significant impact on both DU and GU.

A follow-up study on the effect of Helicobacter pylori eradication on the severity of gastric histology.

Helicobacter pylori genetic diversity and geographic distribution affect the severity of gastric histology; while eradication heals gastritis, the improvement of atrophy and intestinal metaplasia (IM) is still controversial. We investigated whether H. pylori infection and genotypes (cagA-vacA) influence the histological changes and whether eradication resolves these changes. Twenty-one patients (11 duodenal ulcer, 2 gastric ulcer, 8 gastritis) received treatment. Biopsies for CLO, PCR, histology, and culture were collected before and at 1 and 12 months after treatment, and serum samples at 0, 1, 2, 6, and 12 months. H. pylori eradication was achieved in 71% of the patients. Histological scores for H. pylori densities were significantly higher in the antrum and incisura angularis. Scores for mononuclear cell and neutrophil infiltration were significantly higher in regions with a high H. pylori density and improved progressively after eradication. Eight patients with atrophy including five with IM showed no significant changes 12 months after eradication. The cagA gene, detected in 13 (62%), the vacA-sla gene, in 20 (95%), and the vacA-m1 gene, in 12 (57%) of 21 patients were significantly associated with duodenal ulcer. A gradual decline in antibody titer reached an average of 67% 12 months after eradication. H. pylori infection and the associated genotypes (cagA of Western type) affect the severity of the gastric histology (mild forms of atrophy and IM) and the disease outcome. Eradication of H. pylori resulted in healing of gastritis, but with no significant improvement in atrophy or IM.

Enzyme immunoassay and immunoblotting analysis of Helicobacter pylori infection in Turkish asymptomatic subjects.

Approximately half of the world population is infected with Helicobacter pylori, particularly in developing countries. The aims of the study were to detect H. pylori infection in asymptomatic Turkish subjects, correlate the infection with the associated risk factors, and to evaluate the cytotoxin-associated gene (CagA) status and other H. pylori antigens. Three hundred nine asymptomatic subjects (124 female) 1-82 years of age (average: 31 years) were serologically tested by enzyme immunoassay and immunoblotting. The enzyme immunoassay detected IgG anti-H. pylori antibodies in sera of 216 (70%) out of 309 subjects, 132 (61%) male. Infection rates of 42% in subjects <10 years of age, 55% in 10-19 years, 66% in 20-29 years, 78% in 30-39 years, 79% in 40-49 years, 91% in 50-59 years, 100% in 60-69 years, and 80% in those >70 years of age were detected. Subjects >45 years of age had significantly higher antibody responses, odds ratio = 0.16 (95% confidence interval: 0.07-0.37), than those <45 years. H. pylori infection was significantly higher in married subjects, odds ratio = 0.38 (95% confidence interval: 0.20-0.73), and those with low socioeconomic status. No correlation between gender, education, smoking, and nonsteroidal anti-inflammatory drug intake and infection was detected. Immunoblots revealed antibodies to CagA in 58 (83%) of 70 samples tested. H. pylori infection is prevalent in the asymptomatic Turkish subjects. Marital and socioeconomic status was significantly associated with the acquisition of H. pylori. Antibodies to CagA antigen were highly prevalent in these subjects.

The role of the putative virulence markers (cagA and vacA ) of Helicobacter pylori in peptic ulcer disease.

Helicobacter pylori is genetically diverse and certain strains are more virulent and cause more severe diseases than others and such diversity is reflected on the clinical outcome. The cytotoxin-associated gene (cagA) and vacuolating cytotoxin (vacA) gene are 2 putative markers that were associated with peptic ulcer disease. The basis for the epidemiological association between the cagA and vacA genes is not known. In this review, the molecular characteristics of these markers, and its role in the pathogenesis of peptic ulcer disease and gastric cancer are discussed.

Rubella immune status of pregnant and non-pregnant women in Istanbul, Turkey.

OBJECTIVE: Rubella immunization rates are not optimal and infections during pregnancy still occur since many countries incorporate no rubella vaccine in their national immunization program. The evaluation of immunity to rubella virus relies on the presence of specific antibodies. This study was undertaken to determine in a cross-sectional survey whether rubella virus circulation in the Istanbul city, induces detectable immunoglobulin G (IgG) antibodies with a protective level, in a random group of pregnant and non-pregnant women. METHODS: One hundred and sixty women of 20-41-years of age (average 24-years) were grouped as follows: 1. Forty-eight married women. Among these were 41 pregnant women (33 delivered normally, 8 aborted). 2. One hundred and twelve single women. Samples were collected during the periods from October 2000 through to March 2001 and from November 2001 through to May 2002. Rubella specific IgG antibodies were detected (by the ELISA test) in all women tested. RESULTS: Quantitative analysis of the IgG levels showed noticeable variability that ranged between 24-143 IU/ml (average 94). One hundred and forty-five (91%) out of 160 women had rubella IgG levels of above 50 IU/ml with a range of 54-143 IU/ml (average 92) while 15 (9%) had a level between 24-46 IU/ml (average 38). Rubella IgG-avidity test revealed that 116 (73%) of women had high IgG avidity, 22 (14%) had intermediate avidity and 20 (13%) showed low avidity. Two women who were IgM positive, each had either high or intermediate IgG avidity. CONCLUSION: All women tested were seropositive for rubella specific IgG antibodies suggestive of natural virus circulation within the community. Although the majority appeared to possess protective level of such antibodies, screening for protective immunity appears always to be a necessity for future protection against reinfection.

Analysis of Helicobacter pylori genotypes and correlation with clinical outcome in Turkey.

The predominant Helicobacter pylori strains circulating among geographic locations differ in regard to genomic structure. The association of the cagA-positive, vacA s1 genotypes with peptic ulcer disease (PUD) and gastric cancer was reported in Western countries but not in East Asian countries. Strains from Western countries predominantly possessed cagA type 2a, vacA s1a or s1b/m1a, or vacA m2a genotypes, whereas strains from East Asia possessed cagA type 1a, vacA s1c/m1b, or vacA m2b genotypes. Whether the Turkish strains possessed such genotypes was investigated and correlated with the disease outcome. Seventy-three patients from Turkey were enrolled. H. pylori was detected in 65 (89%) patients (22 with gastritis, 33 with PUD, and 10 with gastric cancer) by any of the following tests: Campylobacter-like organism test, culture, or PCR. Among the H. pylori-positive patients, presence of the cagA gene (78%) was significantly associated with PUD (P < 0.00001), gastric cancer (P < 0.001), and vacA s1a genotypes (P < 0.0001). Multiple vacA genotypes were more prevalent in PUD and gastric cancer patients than in patients with gastritis. Restriction fragment length polymorphism analysis of the cagA gene revealed three different patterns with no significant association with clinical outcome. Turkish strains examined predominantly possessed cagA type 2a, vacA s1a/m1a, or vacA m2a genotypes, which were typical genotypes in strains from Western countries. This fact might be one of the reasons for the low prevalence of severe gastroduodenal diseases in Turkey compared to the East Asian countries.