vacA S1 Research Articles

Serum IgG Antibody against Helicobacter pylori and the CagA-VacA s1 Genotype as a Predictor of Gastric Cancer Development: A Case-control Study

Helicobacter pylori (H. pylori)-induced inflammation increases the risk of developing various upper gastrointestinal conditions which may progress to gastric cancer (CA). Early prediction and detection of infection are crucial for reducing cancer-induced mortality rates. The present case-control study aimed to investigate the combination of serum and molecular markers and H. pylori-associated gastro-duodenal conditions as risk factors for predicting CA development in dyspeptic patients. Consecutive gastric biopsies and blood samples were collected from 100 adult dyspeptic patients. Serum IgG antibody levels against H. pylori were determined, and receiver operating characteristic (ROC) analysis was performed. The expression of the virulence genes cagA and vacA was evaluated by polymerase chain reaction (PCR). A significant association was reported between the disease condition and the status of several risk factors, such as family history, serum IgG antibody concentration, and the virulence genes cagA and vacA. Among the 71 H. pylori-positive patients, 35.2% (25/71) had CA. Both cagA and vacA genes were found in 46 out of 71 (64.7%) patients, and 92% (23/25) of CA patients carried the cagA+vacA s1 gene. ROC analysis of the serum IgG concentrations revealed AUC values of 0.81 and 0.78 for differentiating patients with non-ulcer dyspepsia from those with ulceration/inflammation and CA, respectively. The concordance between the IgG-positive and PCR-positive patients was 84% (k value=0.41). Patients who had a family history of CA with an increased serum IgG concentration and the presence of H. pylori cagA-vacA s1 genotypes may be considered strong predictors of future development of gastric pathologies, including CA.

Investigation of Metronidazole Resistance-Associated Mutations and Virulence Genotypes of Helicobacter pylori Isolated from Egyptian Population: A Cross-Sectional Study

IntroductionThis cross-sectional study assesses the prevalence of metronidazole resistance-associated mutations and virulence genotypes in Helicobacter pylori (H. pylori) strains isolated from the Egyptian population. H. pylori infection is a significant public health concern, with antibiotic resistance challenging its eradication. MethodsGastric biopsy samples were collected from symptomatic patients referred for upper gastrointestinal endoscopy at selected healthcare facilities. The study included 250 participants with symptoms suggestive of H. pylori infection and aged 18 years or older. Biopsy samples were obtained using standard endoscopic techniques, and H. pylori strains were isolated and identified in the laboratory. Antimicrobial susceptibility testing was conducted using standard methods. Molecular analysis, including polymerase chain reaction (PCR) and sequencing, was performed to identify metronidazole resistance-associated mutations (rdxA and frxA) and virulence genotypes (cagA and vacA). ResultsAntimicrobial susceptibility testing revealed that 43.6% of the isolates were resistant to metronidazole, while 11.8%, 4.5%, and 55.4% were resistant to clarithromycin, amoxicillin, and levofloxacin. Molecular analysis identified rdxA and frxA mutations in 36.3% and 31.8% of the isolates, respectively, indicating metronidazole resistance-associated mutations. Additionally, 60.0% of the isolates were positive for the cagA gene, and 80.0% had the vacA s1 type, both associated with increased virulence. A significant association was found between metronidazole resistance and the presence of cagA gene, vacA s1 type, rdxA mutation, and frxA mutation. Statistical analysis revealed associations between specific mutations and virulence genotypes with respective odds ratios, indicating higher likelihoods of metronidazole resistance in isolates exhibiting these genetic characteristics. ConclusionsThis study highlights the prevalence of metronidazole resistance and the association between specific mutations and virulence genotypes in H. pylori strains isolated from the Egyptian population. The findings underscore the importance of monitoring antibiotic resistance patterns and understanding the genetic determinants of virulence in H. pylori for effective management and treatment strategies.

Effect of concurrent infection of Helicobacter pylori with Toxoplasma gondii infection on gastric pathology

BackgroundToxoplasma gondii (T. gondii) and Helicobacter pylori (H. pylori) are among the most prevalent foodborne parasitic and bacterial infections worldwide. However, the concurrent impact of coinfection on gastric pathology has yet to be studied in depth. The effect of coinfection generally either adds a synergetic or antagonistic impact; we aimed in the current work to assess the impact of T. gondii coinfection on the progression of H. pylori-associated gastric pathology and reporting H. pylori virulent strains. The study was conducted on 82 patients complaining of persistent gastrointestinal symptoms with failed treatment response and prone to endoscopy. They were subjected to stool examination to detect H. pylori antigen, serological screening for latent toxoplasmosis, endoscopy, histopathological examination, and molecular detection of H. pylori virulence strains in gastric biopsies. Out of the 82 patients, 62 patients were positive for H. pylori antigen in stool and 55 patients confirmed positivity by histopathology; out of them, 37 patients had isolated Vac As1 variants, 11 patients had combined Vac As1 and Cag A variants, and 7 patients had combined Vac As1, Cag A and VacAs2 variants. Patients with the combined two or three variances showed significantly deteriorated histopathological features than patients with a single Vac As1 variant (P < 0.05). Latent toxoplasmosis was positive among 35/82 patients. Combined H. pylori and Toxoplasma gondii infection had significantly marked inflammation than patients with isolated infection (P < 0.05). Conclusion: Screening for toxoplasmosis among H. pylori-infected patients is recommended as it is considered a potential risk factor for gastric inflammation severity. H. pylori gastric inflammation may be heightened by Toxoplasma coinfection.

Historical and Molecular Perspectives on the Presence of Helicobacter pylori in Latin America: A Niche to Improve Gastric Cancer Risk Assessment.

Helicobacter pylori (H. pylori) is responsible for causing chronic gastritis, which can cause peptic ulcer and premalignant lesions such as atrophic gastritis, intestinal metaplasia, and dysplasia, with the risk of developing gastric cancer. Recent data describe that H. pylori colonizes the gastric mucosa of more than 50% of the world's population; however, this bacterium has been described as infecting the human population since its prehistory. This review focuses on the populations and subpopulations of H. pylori, differentiated by the polymorphisms present in their constitutive and virulence genes. These genes have spread and associated with different human populations, showing variability depending on their geographical distribution, and have evolved together with the human being. The predominant genotypes worldwide, Latin America and Chile, are described to understand the genetic diversity and pathogenicity of H. pylori in different populations and geographic regions. The high similarity in the sequence of virulence genes between H. pylori strains present in Peruvian and Spanish natives in Latin America suggests a European influence. The presence of cagA-positive strains and vacA s1 m1 allelic variants is observed with greater prevalence in Chilean patients with more severe gastrointestinal diseases and is associated with its geographical distribution. These findings highlight the importance of understanding the genetic diversity of H. pylori in different regions of the world for a more accurate assessment of the risk of associated diseases and their potential impact on health.

A rapid and high-throughput multiplex genetic detection assay for detection, semi-quantification and virulence genotyping of Helicobacter pylori in non-invasive oral samples.

This study established a high-throughput multiplex genetic detection assay (HMGA) for rapid identification, semi-quantification and virulence analysis of Helicobacter pylori directly from the clinical non-invasive oral samples. The gastric mucosa and oral samples were collected from 242 patients in Shanghai from 2021 to 2022. All the samples were detected by routine clinical tests for H. pylori and Sanger sequenced for inconsistent results. A new multiplex PCR assay providing results within 4 hours was designed and optimized involving fluorescent dye-labeled specific primers targeted 16S rRNA gene, semi-quantitative gene ureC and 10 virulence genes of H. pylori. Semi-quantification was carried out by simulating the serial 10-fold dilutions of positive oral samples, and the H. pylori loads in different clinical samples were further compared. The mixed plasmids of virulence genes vacA s1, vacA m1 and vacA m2 were used to evaluate the performance on different genotypes. The consistency of 10 virulence genes in gastric mucosa, saliva, mouthwash and dental plaque of H. pylori-positive patients was compared. The non-invasive HMGA was highly specific for detection of all 12 targets of H. pylori and human internal reference gene β-globin, and the sensitivity to all target genes could reach 10 copies/μL. Compared with routine clinical tests and sequencing, non-invasive HMGA has a high level (>0.98) of sensitivity, specificity, accuracy, PPV, NPV and kappa coefficient for direct detection of H. pylori in oral samples. Moreover, by detecting peak area levels of ureC, it was confirmed that the H. pylori loads in gastric mucosa were significantly higher than those of the three kinds of oral samples (p<0.05). We also found that 45.0% (91/202) of patients had different H. pylori virulence genes in different oral samples. The concordance of positive detection rates of each virulence gene between saliva and gastric mucosa was more than 78% (p<0.05). The non-invasive HMGA proved to be a reliable method for the rapid H. pylori identification, semi-quantification and detection of 10 virulence genes directly in oral samples, providing a new idea for non-invasive detection of H. pylori.

RETRACTED ARTICLE: Helicobacter pylori strains isolated from raw poultry meat: frequency and molecular characteristics

Even though Helicobacter pylori (H. pylori) is a serious pathogen, its origin is unknown. Poultry (Chicken, Turkey, Quebec, Goose, and Ostrich) are consumed as a regular protein source by a large number of people across the world; therefore, sanitary ways of delivering poultry for food are important for global health. As a result, we looked at the distribution of the pathogenicity cagA, vacA, babA2, oipA, and iceA in H. pylori isolates in poultry meat, as well as their antimicrobial resistance. Wilkins Chalgren anaerobic bacterial medium was used to cultivate 320 raw poultry specimens. Disk diffusion and Multiplex-PCR were used to investigate antimicrobial resistance and genotyping patterns, separately. H. pylori was found in 20 of 320 (6.25%) raw poultry samples. The highest incidence of H. pylori was found in chicken raw meat (15%), whereas the fewest was found in Goose and Quebec (0.00%). Resistance to ampicillin (85%), tetracycline (85%), and amoxicillin (75%) were greatest in H. pylori isolates. The percentage of H. pylori isolates with a MAR value of more than 0.2 was 17/20 (85%). The most prevalent genotypes discovered were VacA s1a (75%), m1a (75%), s2 (70%) and m2 (65%), and cagA (60%). The most typically discovered genotype patterns were s1am1a (45%), s2m1a (45%), and s2m2 (30%). BabA2, OipA + , and OipA− genotypes were found in 40%, 30%, and 30% of the population. In summary, the poultry flesh was polluted by H. pylori, with the babA2, vacA, and cagA genotypes being more prevalent. The simultaneous occurrence of vacA, cagA, iceA, oipA, and babA2 genotypes in antibiotic-resistant H. pylori bacteria implies a serious public health concern about raw poultry eating. In the future, researchers should look into H. pylori's resistance to multiple antibacterial drugs in Iran.

Prevalence of Helicobacter pylori genotypes: cagA, vacA (m1), vacA (s1), babA2, dupA, iceA1, oipA and their association with gastrointestinal diseases. A cross-sectional study in Quito-Ecuador

BackgroundThe most prevalent stomach infection in the world is caused by Helicobacter pylori (H. pylori). Several pathogenicity genes, including cagA, vacA, babA2, dupA, iceA, and oipA, are associated with an increased risk of gastrointestinal disease such as peptic ulcer and stomach cancer. This research aims to determine the prevalence of different H. pylori genotypes and correlate their risk in the development of gastrointestinal diseases in the Ecuadorian population.MethodsA cross-sectional research of 225 patients at the Calderón Hospital in Quito, Ecuador, was conducted. End point PCRs were run to determine the presence of 16S rRNA, cagA, vacA (m1), vacA (s1), babA2, dupA, iceA1, and oipA virulence genes. Chi-square test, odds ratios (OR) and 95% confidence intervals (CI) were utilized for the statistical analysis.ResultsH. pylori infection was present in 62.7% of people. Peptic ulcers were seen in 22.2% and malignant lesions in 3.6% of patients. Genes oipA (93.6%), vacA (s1) (70.9%), and babA2 (70.2%) were the most prevalent. cagA/vacA (s1m1) and cagA/oipA (s1m1) combinations were found in 31.2% and 22.7% of the cases, respectively. Acute inflammation has a significant correlation with the genes cagA (OR = 4.96 95% CI: 1.1–22.41), babA2 (OR = 2.78 95% CI: 1.06–7.3), and the cagA/oipA combination (OR = 4.78, 95% CI: 1.06–21.62). Follicular hyperplasia was associated with iceA1 (OR = 3.13; 95% CI: 1.2–8.16), babA2 (OR = 2.56; 95% CI: 1.14–5.77), cagA (OR = 2.19; 95% CI: 1.06–4.52), and the cagA/oipA combination (OR = 2.32, 95% CI: 1.12–4.84). The vacA (m1) and vacA (s1m1) genes were associated with gastric intestinal metaplasia (OR = 2.71 95% CI: 1.17–6.29) (OR = 2.33 95% CI: 1.03–5.24). Finally, we showed that cagA/vacA (s1m1) gene combination increased the risk of duodenal ulcer development (OR = 2.89, 95% CI 1.10–7.58).ConclusionThis study makes a significant contribution by offering genotypic information regarding H. pylori infection. The presence of several H. pylori genes was associated with the onset of gastrointestinal illness in the Ecuadorian population.

Clinical relevance and distribution of Helicobacter pylori virulence factors in isolates from Chinese patients.

At present, the relationship between virulence factors of Helicobacter pylori (H. pylori) and gastrointestinal diseases is still under discussion. This study investigated the association between distinct virulence factors in H. pylori and different gastrointestinal diseases. Gastric biopsy specimens were obtained from 160 patients with different gastrointestinal diseases in China, including 77 patients with chronic gastritis (CG), 36 with peptic ulcer disease (PUD), and 38 with gastric carcinoma (GC). The presence of certain virulence genes was determined by polymerase chain reaction (PCR), and the results were analyzed using chi-squared tests. A total of 160 H. pylori strains were successfully isolated from gastric biopsy specimens. Overall, all strains of H. pylori were cagA, cagE positive, and the most common vacA genotypes were s1 (98.8%) and m2 (68.1%). The positive rates of the jhp0562, jhp0563, homA, homB, hopQI, and hopQII genes were 99.4%, 32.5%, 33.1%, 71.3%, 100%, and 6.9%, respectively. There was no significant association between these genes and different disease types. The dominant hpyIIIR-positive genotype was detected in 83.1% of the strains, making it significantly more prevalent than the hrgA-positive genotype (P<0.001). Surprisingly, the mixed genotype of hrgA and hpyIIIR was common and accounted for 41.3%. The hrgA-positive strains were more common in GC patients (71.1%) compared to CG patients (50.7%, P<0.05). The mixed genotype was prevalent and accounted for 55.3% and 31.2% of strains from GC and CG patients, respectively. Multivariate analysis showed that the hrgA gene exhibited a positive correlation with GC and increased the risk of GC [odds ratio (OR) =3.606, P<0.05]. In contrast, the presence of hrgA exhibited a negative correlation with CG (OR =0.499, P<0.05). These results suggested that the universal presence of cagA, cagE, vacA s1, jhp0562, homB, and hopQI made it impossible to examine disease-specific associations with any of these virulence factors. In addition, they may synergistically contribute to more virulent strains and severe diseases in China. Furthermore, there was a strong association between the hrgA gene and progression to GC, indicating the potential application of other virulence factors in clinical detection.

Pattern and trends of Helicobacter pylori genotypes in gastric cancer: A Kenyan 8-year study.

Notable geographic and temporal variations in the prevalence and genotypes of Helicobacter pylori, in relation to gastric pathologies, have been observed; however, their significance and trends in African populations is scarcely described. The aim of this study, was to investigate the association of H. pylori and its respective CagA and vacuolating cytotoxin A (VacA) genotypes with gastric adenocarcinoma, and to describe the trends of H. pylori genotypes over an 8-year period (2012-2019). A total of 286 samples of gastric cancer cases and benign controls (one-to-one matching), from three main cities in Kenya, between 2012 and 2019 were included. Histologic evaluation, and CagA and VacA genotyping using PCR, was performed. Distribution of H. pylori genotypes was presented in proportions. To determine association, a univariate analysis was conducted using a Wilcoxon rank sum test for continuous variables, and a Chi-squared test or Fisher's exact test for categorical data. The VacA s1m1 genotype was associated with gastric adenocarcinoma, {odds ratio (OR) = 2.68 [confidence interval (CI) of 95%: 0.83-8.65]; p = 0.108}, whilst VacA s2m2 was associated with a reduced probability of gastric adenocarcinoma [OR = 0.23 (CI 95%: 0.07-0.78); p = 0.031]. No association between cytotoxin associated gene A (CagA) and gastric adenocarcinoma was observed. Over the study period, an increase in all genotypes of H. pylori was seen, and although no predominant genotype was noted, there was significant year-to-year variation, with VacA s1 and VacA s2 showing the greatest variation. VacA s1m1 and VacA s2m2 were associated with increased, and reduced risk of gastric cancer, respectively. Intestinal metaplasia and atrophic gastritis did not appear to be significant in this population.

Virulence determinants and genotypes of &lt;i&gt;Helicobacter pylori&lt;/i&gt; clinical isolates

Background. H. pylori is the principal causative agent of gastroduodenal disorders in humans. The development and severity of lesions in infected individuals depend on the virulence of H. pylori strains.&#x0D; Aims: Detection of virulence determinants and comparative analysis of H. pylori genotypes in patients with chronic gastritis (CG) and duodenal ulcer (DU).&#x0D; Materials and methods. The 53 H. pylori strains were isolated in St. Petersburg from patients with CG (n = 34) and DU (n = 19). The genetic determinants of virulence cagA, iceA, vacA and H. pylori genotypes in patients with CG and UC were determined using the standard PCR method.&#x0D; Results. The cagA gene was found in 64.1% of H. pylori strains. The proportions of cagA+ isolates from patients with CG and DU was 55.8% (15/34) and 78.9% (15/19), respectively (p 0.05). The iceA1 allele of H. pylori was detected in 47.4% of patients with DU, the iceA2 in 47.1% of patients with CG (p 0.05). The vacAs1 allele was significantly dominant in patients with DU 94.7% versus 70.6% in CG (p 0.05). No significant difference in vacA m1 and m2 alleles was found in H. pylori from different groups of patients (p 0.05). All cagA+ strains were carriers of the vacA s1 allele. The vast majority of strains (10 out of 11) of the cagA/vacAs2 genotype were isolated from patients with CG.&#x0D; Conclusion. The significant association between vacAs1, vacAs2 allelic variants, as well as vacA s1/m2, vacA s2/m2 genotypes of the pathogen and severity of clinical manifestations of H. pylori infection has been established in our study. The vacAs1 and vacA s1/m2 genotypes of the pathogen are associated with duodenal ulcer.

Prevalence, Virulence Genes, Phylogenetic Analysis, and Antimicrobial Resistance Profile of Helicobacter Species in Chicken Meat and Their Associated Environment at Retail Shops in Egypt.

Helicobacter pylori (H. pylori) and Helicobacter pullorum (H. pullorum) are frequently reported pathogens in humans and poultry, respectively. Nevertheless, the source of H. pylori is still unclear. This study aimed to detect Helicobacter spp. in chicken carcasses and to assess the antibiogram and the virulence genes of Helicobacter isolates. Three hundred chicken meat samples (100 each of chicken breast, liver, and gizzard), besides 60 swab samples from chicken processing surfaces, were collected from retail shops in Qalyubia Governorate, Egypt, and examined for the prevalence of H. pylori and H. pullorum. The 16S rRNA of three H. pylori and two H. pullorum isolates were sequenced to determine the genetic relationship between these two Helicobacter spp. Of the 300 chicken samples tested, 16 (5.33%) and 14 (4.67%) were positive for H. pylori and H. pullorum, respectively. Multiplex PCR revealed that the virulence genes vacuolating cytotoxin A (vacA)s1, cytotoxin-associated gene A (cagA), and restriction endonuclease-replacing gene A (hrgA) were detected in 66.7%, 77.8%, and 100% of H. pylori strains tested, respectively. H. pylori showed the highest resistance for clarithromycin, while H. pullorum exhibited the highest resistance towards erythromycin and ciprofloxacin. The study concluded that the chicken meat and giblets are potential sources of the virulent and antimicrobial-resistant strains of H. pylori of human origin.

Genetic polymorphisms of &lt;i&gt;helicobacter pylori&lt;/i&gt; clinical isolates in St. Petersburg, Russia

Introduction. Helicobacter pylori was proved to be the principal causative agent of gastroduodenal disorders in human. Although Russian Federation is among the countries with a high prevalence of H. pylori infection (6090%), currently there is a very limited number of studies evaluating H. pylori genotypes in Russia. Objective. Based on the assessment of virulence-associated cagA, oipA, and vacA genes, our study was aimed to determine H. pylori genotypes associated with the clinical outcomes in patients with H. pylori infection in St. Petersburg, Northwest Russia. Materials and methods. Using PCR for the detection of cagA, oipA, and vacA s, m, i allelic variants, we analyzed 61 H. pylori isolates isolated and cultured from biopsies collected during endoscopy of patients with chronic gastritis (G), duodenal ulcer (DU), and gastric cancer (GC). Results. The genetic diversity of H. pylori clinical isolates has been revealed (HGDI 0.88): 41 (67%) of 61 H. pylori isolates were cagA-positive, 38 (62%) oipA-positive. The proportions of cagA+ isolates differed in patients with G (56.7%) and DU (80.9%) (p = 0.06). The s, m, and i allelic variants of the vacA gene were detected in all strains, although the vacA s1 allele was significantly dominant in patients with DU (95.2%) rather than with G (64.9%) (p = 0.01). The vacA alleles m1 and i1 in the isolates from patients with G and DU were found in almost equal proportions: 45.9% and 42.8% for m1 allele, 45.9% and 47.6% for i1 allele, respectively. Seven isolates (11.5%) were positive for different mixed combinations of vacA alleles s, m, and i. Noteworthy, all vacA s2 strains were cagA-negative and had the m2 allele. OipA+ strains were found in almost equal proportions in patients with G (62.2%) and DU (57.1%) (p = 0.71). All three cagA- and oipA-positive isolates from patients with GC carried vacA s1/m1/i1 alleles. Different combinations of virulence-associated determinants constituted 17 genetic profiles. The most common combined genotype cagA+/oipA+/vacA s1/m1/i1 comprised 18 (29.5%) H. pylori isolates. Conclusion. We have determined predominant genotypes in the H. pylori population in the Northwest of Russia. The significant association between vacA s1 genotype of the pathogen and clinical manifestations of H. pylori infection has been established in our study.

Correlation of Helicobacter pylori virulence genotype & severity of mucosal inflammation in gastric biopsies from two geographically diverse regions in India.

H. pylori-associated gastritis in patients from the high-altitude area of Ladakh showed severe gastritis, mucosal nodularity, atrophy, and cancer in comparison to those from North India. This study served to analyze if differences in the H. pylori virulence genotypes decide the extent of gastric mucosal inflammation. Fifty gastric biopsies each from patients with H. pylori-associated gastritis from Ladakh and a tertiary care center in North India were included. The presence of H. pylori strain was confirmed with Warthin starry stain and polymerase chain amplification of the H. pylori-specific 16S rRNA. The cagA, vacA s1, s2, and m1, m2 alleles, and dupA virulence genotypes were studied in all archival samples, followed by their histological correlations. cagA (P 0.009) and vacAs1 m1 (P 0.009) genes were distinctly more in H. pylori strains colonizing the biopsies of North Indian patients. In contrast, the cagA -ve vacAs2 m2 strains were significantly more in H. pylori strain colonizing the biopsies from Ladakhi patients. dupA genotype was almost similarly present in strains from both regions. Among these, only cagA and dupA virulence genes were associated with severe mucosal neutrophilic activity and deep infiltration of H. pylori strains in North Indian patients. Differences in virulence genotypes of H. pylori in gastric biopsies from North Indian and Ladakhi patients were found not significant in deciding the severity of H. pylori-associated gastritis.

Study on the value of antibiotic-resistant gene detection in Helicobacter pylori in China.

The aim of the present study was to explore the value of detecting antibiotic-resistant genes in Helicobacter pylori (H. pylori) and the association between genotype and antibiotic resistance. Two gastric mucosa samples from each H. pylori-positive patient were collected. Each patient's H. pylori sample was cultured in vitro, and the agar plate dilution method was conducted. In addition, all patient samples were analyzed for the detection of antibiotic resistance-related mutant genes and VacA gene genotypes. The association between VacA genotypes and antibiotic resistance was also determined and the value of mutant gene detection in predicting H. pylori resistance to antibiotics was evaluated. In total, 133 H. pylori-positive patients were enrolled. A total of 22 strains of H. pylori failed to grow in in vitro culture and 25 strains were negative in a H. pylori gene test. Among 108 strains detected by PCR, a total of 39 VacA s1m1 strains, 69 VacA s1m2 strains and no VacA s2 strain were identified. There was no significant association between VacA genotypes and antibiotic resistance. The mutation rates of G616A in the rdxA gene, T87A, G91A, A91G and G91T in the gyrA gene and A2143G and A2142G in the 23S rRNA gene were 32.1, 32.3, 22.6, 12.9, 6.5, 81.8 and 0.0%, respectively. Among these mutant sites, the mutation coincidence rates were as follows, according to the agar plate dilution method: rdxA G616A (81.8%), gyrA G91T (66.7%), gyrA G91A (54.5%), 23 S rRNA A2143G (49.1%), gyrA T87A (45.5%), gyrA A91G (33.3%), penicillin-binding protein 1 (PBP1) C556G (0.0%), PBP1 A562T (0.0%), PBP1 A562G (0.0%) and 16 S rRNA 926-927 (AT-GT) (0.0%). VacA m subtypes were not associated with H. pylori antibiotic resistance. In conclusion, the present findings suggested that the detection of related mutant genes had a clinical application value in predicting the antibiotic resistance of H. pylori, particularly resistance to clarithromycin and levofloxacin.

Clinical relevance of virulence genes in Helicobacter pylori isolates recovered from adult dyspeptic patients in Turkey

PurposeBacterial virulence factors play a major role in the pathogenesis of Helicobacter pylori infection. The aims of this study were to evaluate virulence genes in H. pylori isolates and to compare the presence of these genes and associated clinical pathologies. MethodsA total of 148 H. pylori isolates, recovered from adult dyspeptic patients, were used. The patients, from whom the isolates were obtained, were assigned to two groups by their endoscopic findings, which manifested as chronic gastritis or peptic ulcer. The presence of gastric atrophy and intestinal metaplasia was recorded for each patient, based on histopathological examination. Analyses of the virulence genes were performed by the polymerase chain reaction technique. ResultsThe patients had a mean age of 47 ​± ​15 years and 86 (58%) of them were female. Based on endoscopic examination, 103 (69.6%) patients were diagnosed with chronic gastritis and 45 (30.4%) with peptic ulcer. Histopathological examination revealed intestinal metaplasia in 30 (20%) patients and gastric atrophy in 12 (8%) patients. The prevalence rates of cagA, cagE, iceA1, iceA2, and babA2 were determined to be 87%, 74%, 58%, 26%, and 95%, respectively. The most prevalent vacA alleles were s1/s1a (82%/97%) and the least prevalent allele was s2 (20%). A new vacA genotype (s1as1bs1c) was detected, for the first time, in 18 (12%) isolates. No significant difference was found between the patient groups with chronic gastritis and peptic ulcer for the prevalences of the virulence genes (p ​> ​0.05). Furthermore, intestinal metaplasia and gastric atrophy showed no significant correlation with the virulence genes (p ​> ​0.05). ConclusionsIt is thoughted that H. pylori isolates with predominant cagA, cagE, VacA (s1, s1a), and babA2 virulence genes are associated with gastroduodenal diseases. However, there is no correlation between gastric premalignant lesions and virulence genes.

Association of Helicobacter pylori vacA polymorphisms with the risk of gastric precancerous lesions in a Moroccan population.

Helicobacter pylori infection is the major risk factor of atrophic gastritis and intestinal metaplasia. The vacA gene is one of the most virulence factors of H. pylori and genetic diversity in its s, m, i, and d regions is associated with gastric lesions severity. This study aimed to investigate the association of vacA s, m, i, and d regions with the risk of atrophic gastritis and intestinal metaplasia in a Casablanca population. A total of 210 patients suffering from gastric lesions (chronic gastritis, atrophic gastritis, and intestinal metaplasia) were enrolled. The type of lesion was diagnosed by histological examination. Detection of H. pylori infection and genotyping of vacA regions were carried out by PCR. The prevalence of H. pylori was 95%. The most common vacA genotypes were s2 (51.5%), m2 (77%), i2 (60.5%), and d2 (58.5%). VacA s1, m1, and i1 genotypes were associated with a high risk of intestinal metaplasia, while the vacA d1 genotype increases the risk of atrophic gastritis and intestinal metaplasia. The most common vacA combination was s2/m2/i2/d2 (52%), and it was more detected in chronic gastritis. The moderate virulent vacA combination (s1/m2/i1/d1) increases the risk of atrophic gastritis, while the most virulent vacA combination (s1/m1/i1/d1) increases the risk of intestinal metaplasia. Genotyping of vacA d region might be a reliable marker for the identification of vacA virulent strains that represent a high risk of developing precancerous lesions (atrophic gastritis and intestinal metaplasia).

The crosstalk between H. pylori virulence factors and the PD1:PD-L1 immune checkpoint inhibitors in progression to gastric cancer

BackgroundThe progression to gastric cancer has been linked to chronic infection with Helicobacter pylori (H. pylori). Immune checkpoint inhibitors (programmed cell death -1, PD-1; programmed cell death –ligand 1, PD-L1) have a role in cancer immune escape. The relationship between H. pylori virulence factors with PD-1, PD-L1 T helper 1 (Th1), T helper 17 (Th17), and regulatory T cell (Treg) response genes, has not been thoroughly investigated in the development of gastric cancer. Therefore, we evaluated how H. pylori virulence factors influence the expression levels of immune-related genes in the development of gastric immunopathology. MethodsA total of 92 gastric tissues of normal controls and patients with gastritis, gastric ulcer, and gastric cancer were examined for the expression of immune-checkpoint inhibitor genes (PD-1 PD-L1), Th1 (interferon- γ, IFN-γ), Th17 (interleukin- 17, IL-17, Retinoic-acid-receptor- related orphan nuclear receptor gamma t, RORγ-t), and Treg (Forkhead box P3, FOXP3) response genes with quantitative real-time PCR (qRT-PCR). Furthermore, correlation of H. pylori virulence factors’ (cytotoxin-associated gene A, cagA; vacuolating cytotoxin gene A, vacA (s1,s2,m1,m2); blood group antigen-binding adhesin gene A, babA, duodenal ulcer promoting gene A, dupA; the putative neuraminyllactose-binding hemagglutinin homolog, hpaA; neutrophil-activating protein A napA; outer inflammatory protein A, oipA; urease A, ureA; and urease B, ureB) genotypes with a degree of inflammation and density of H. pylori were investigated. Next, the relationship between H. pylori virulence factors and immune-checkpoint inhibitor genes, and T-cell response genes was evaluated. Eventually, a decision tree model was developed to determine the clinical outcome of patients using expression data. ResultsThe intensity of PD-1 and PD-L1 mRNA expression was increased significantly in gastric tissue of patients with gastric ulcer (PD-1: 2.3 fold, p=0.01; PD-L1: 2.1 fold, p=0.004), and gastric cancer (PD-1: 2 fold, p= 0.04; PD-L1: 1.8 fold, p=0.05) compared with control subjects. Also, PD-1: PD-L1 expression was significantly higher in patients with gastritis, who were infected with a marked density of H. pylori compared with its mildly infected counterparts. Furthermore, a novel negative correlation was found between PD-1 (r= -0.43) and PD-L1 (r= -0.42) with FOXP3 in patients with gastritis. CagA-positive H. pylori strain's negative association with PD-L1 expression (r=-0.34) was detected in patients with gastritis. Interestingly, PD-1 mRNA expression correlated positively with vacA s2/m2, in gastritis (r=0.43) and ulcer (r=0.43) patients. Furthermore, PD-1: PDL1 expression negatively correlated with vacA m1/m2 (r=-0.43 for PD-1; r=-0.38 for PD-L1) in gastritis patients. Moreover, an inverse correlation of PDL1 was present with vacA m1 (r=0.52) and vacA s1/m1 (r=0.46) versus vacA m2 (r=-0.44) and vacA m1 (r=0.52) and vacA s1/m2 (r=-0.14) in ulcer patients, respectively. Also, a correlation of vacA m2 (r=-0.47) and vacA s1/s2 (r= 0.45) with PD-1 was detected in ulcer patients. In addition, a novel negative correlation between FOXP3 mRNA levels and napA was shown in patients with gastritis and ulcer (r=-0.59). Finally, a computer-based model that was developed showed that knowing the expression levels of PD-L1, RORγ-t, and vacA s1/m2 would be useful to detect the clinical outcome of a patient. ConclusionOur results suggested that PD-1:PD-L1 immune checkpoint inhibitors were increased in gastric pre-cancerous lesions that progress to gastric cancer. Herein, we report the relationship between H. pylori virulence factors and expression of host immune checkpoint inhibitors for diagnostic prediction of gastric malignancies using computer-based models.

Genotyping of Helicobacter pylori Virulence Genes cagA and vacA: Regional and National Study.

Helicobacter pylori (H. pylori) plays a crucial role in the pathogenesis of gastritis, peptic ulcer, and gastric cancer. The presence of pathogenicity islands (PAI) genes contributes to the pathogenesis of many gastrointestinal disorders. Cytotoxin-associated gene A (cagA) and vacuolating cytotoxin gene (vacA) are the most known virulence genes in H. pylori. So, our aim was to study H. pylori virulence genes' role in gastric disorders pathogenesis. Our study included 150 adult patients who suffered dyspeptic symptoms and were referred to the GIT endoscopy unit. Gastric biopsies were attained for rapid urease test (RUT) and histopathological examination, and multiplex PCR technique for detection of virulence genes was performed. It was found that 100 specimens were (RUT) positive, of which sixty samples (60%) were PCR positive for H. pylori ureC gene. The vacA and cagA genes were identified in 61.6% and 53% of H. pylori strains, respectively. Only 5 cases were vacA-positive and cagA-negative. The most virulent vacA s1 allele existed in 56.6% of cases. Out of the 60 H. pylori strains, 66% had at least one virulence gene and 34% did not show any virulence gene. H. pylori infection showed significant increase with age. H. pylori are prevalent amid dyspeptic patients in our region. The main genotype combinations were vacA+/cagA+ of s1m1 genotype and they were frequently associated with peptic ulcer diseases, gastritis, and gastroesophageal reflux disease.

Predicting the role of dupA-positive Helicobacter pylori strains in severe gastrointestinal disorders: An updated meta-analysis

BackgroundIn 2005, a novel virulence factor of Helicobacter pylori was discovered that could increase the risk of duodenal ulceration by stimulating the secretion of IL-8. Hence, it was named duodenal ulcer promoting gene A (dupA). According to statistical analysis, the association between dupA and severe clinical outcomes in Asian and Western countries was evaluate. MethodsA systematic search was performed on the association between dupA and its clinical outcomes. Finally, a sensitivity analysis was used to eliminate the significant heterogeneity, as well as random-effect meta-regression was used to assess confounding variables such as cagA and vacA s1. Finally, publication bias was calculated using Egger's and Begg's p value tests. ResultsOf 182 related studies, 31 reliable studies were included in the quantitative analysis according to inclusion criteria. Sensitivity analysis showed that dupA significantly increases the risk of duodenal ulcer and gastric cancer (OR: 1.58; 95% CI: 1.33–1.88 and OR: 1.27; 95% Cl: 0.99–1.63, respectively), while no significant relationship was observed between dupA and gastric ulcer risk (OR: 1.27; 95% Cl: 0.99–1.63). Based on meta-regression results, cagA and vacA s1 (slope: 0.31; p value: 0.05, slope: 0.23; p value: 0.05, respectively) virulence factors could significantly effect on the association between dupA and the risk of gastric cancer. Due to confounding effects of cagA and vacA s1, the virulence effects of dupA are distorted, and it appears that this gene alone does not increase the risk of gastric cancer. ConclusionOur results confirmed the previous studies, and we found that dupA alone does not appear to increase the risk of gastric adenocarcinoma.

VIRULENCE PROFILE OF HELICOBACTER PYLORI DETECTED IN GASTRIC BIOPSIES OF PATIENTS UNDERGOING ENDOSCOPY IN UPPER EGYPT

Background: Helicobacter pylori (H. pylori) is a common cause of gastric ulcers and is a risk factor for gastric carcinoma. Little data are available about the characters of H. pylori causing infections in patients from Upper Egypt. Therefore, the aim of this study was to investigate the frequency of H. pylori infection in Egyptian patients undergoing endoscopy and complaining of persistent upper gastrointestinal symptoms directly in biopsy specimens using PCR technique and to study the associated virulence-related genes in H. pylori positive samples. Materials and Methods: One hundred and twenty gastric biopsy specimens were collected from Egyptian patients admitted to Assiut University Hospital and complaining of persistent upper gastrointestinal symptoms. H. pylori DNA was extracted for molecular identification by PCR. Positive samples were further analyzed to investigate the presence of different virulence-related genes and gene combinations. Results: H. pylori was detected in 92/120 (76.6%) of collected biopsies. All strains carried the vacA s1 subtype. The prevalence of the virulence genes cagA, cagE, iceA1, iceA2 and oipA were 79 (84.8%), 41(44.5%), 69 (75%), 44 (47.8%), and 55(59.7%) of the H. pylori positive samples, respectively. Genetic analysis showed that H. pylori were grouped into 29 different genotype combinations. The s1/m1/i1/d1/c1/cagA/iceA1/oipA genotype was the most predominant. Conclusion: Our results show a high frequency of H. pylori infections among Egyptian patients with gastrointestinal symptoms. The s1/m1/i1 vacA hybrid is the most prevalent subtype. The distribution of the different virulence-related genes shows alarming rates of cagA, cagE, iceA1, iceA2 and oipA. Careful monitory of H. pylori infections and investigation of their genetic characters should be carried out to control the spread of the virulent strains.