Clinical relevance of cagL gene and virulence genotypes with disease outcomes in a Helicobacter pylori infected population from Iran.

Helicobacter pylori infection is common in Iran as in other developing countries. Certain genotypes of H. pylori have been associated with increased occurrence of chronic gastritis, peptic ulcers, and gastric adenocarcinoma. The aim of this study was to investigate the clinical relevance of cagL gene and other virulence genotypes of H. pylori isolates with clinical outcomes in Iranian patients. Totally, 126 symptomatic patients who underwent gastroduodenal endoscopy were enrolled in the study. Sixty-one H. pylori strains were isolated from the patients studied. The presence of the cagL, cagA, vacA, iceA, babA2 and sabA genes in the corresponding H. pylori isolates were determined by polymerase chain reaction and the results were compared with clinical outcomes and histopathology. The cagL, cagA, vacA s1, vacA s2, vacA m1, vacA m2, iceA1, iceA2, babA 2 , and sabA genotypes were detected in 96.7, 85.2, 75.4, 24.6, 29.5, 70.5, 42.6, 23, 96.7, and 83.6% of the isolates, respectively. The three genotypic combinations, cagL/cagA/vacAs1m1/iceA1/babA2/sabA, cagL/cagA/vacAs1m2/iceA1/babA2/sabA, and cagL/cagA/vacAs1m2/iceA2/babA2/sabA were determined as the most prevalent combined genotypes. There was a significant correlation between the presence of cagL gene and cagA positivity (P = 0.02). No significant correlation was found between the various genotypes and clinical outcomes (P > 0.05). The present study showed a very high prevalence of cagL genotype among the H. pylori isolates from Iranian patients. Our results demonstrated that neither single genotype nor combination genotypes of virulence-associated genes was significantly helpful markers for predicting the severity of gastroduodenal disease associated with H. pylori infection in Iranian patients.

Differentiation of non-pylori Helicobacter species based on PCR-restriction fragment length polymorphism of the 23S rRNA gene.

Phenotypic identification of non-pylori Helicobacter species has always been problematic and time-consuming in comparison with many other bacteria. We developed a rapid two-step identification assay based on PCR-restriction fragment length polymorphism (PCR-RFLP) analysis of the 23S rRNA gene for differentiating between non-pylori Helicobacter species. A new genus-specific primer pair based on all available complete and partial 23S rRNA sequences of Helicobacter species was designed. In silico restriction analysis of variable regions of the 23S rRNA gene suggested SmaI and HindIII endonucleases would provide a good level of differentiation. Analysis of the obtained 23S rRNA RFLP patterns divided all Helicobacter study strains into three species groups (groups A-C) and 12 unique restriction patterns. Wolinella succinogenes also gave a unique pattern. Our proposed PCR-RFLP method was found to be as a valuable tool for routine identification of non-pylori Helicobacter species from human or animal samples.

Prevalence of multiple drug-resistant Helicobacter pylori strains among patients with different gastric disorders in Iran.

Emergence of multidrug-resistant (MDR) strains of Helicobacter pylori is a global health concern. This study was aimed to determine the frequency of MDR H. pylori strains in Iran. H. pylori isolates were obtained from cultured gastric biopsy samples on selective culture media after their characterization by PCR and conventional biochemical methods. The minimal inhibitory concentrations of rifampicin, ciprofloxacin, levofloxacin, ampicillin, clarithromycin, erythromycin, metronidazole, and tetracycline were determined for 111 strains that were isolated from 197 dyspeptic patients by the agar dilution method. The primary resistance rates were 61.3% (68/111) for metronidazole, 15.3% (17/111) for ampicillin, and 14.4% (16/111) for rifampicin. Resistance rates for other antimicrobials were as follows: macrolides (erythromycin or clarithromycin) 32.4% (36/111) and quinolones (levofloxacin or ciprofloxacin) 30.6% (34/111). Among the resistant strains, the rates of double and multiple drug resistance phenotypes were 22.6% (19/84) and 34.5% (29/84), respectively. The quadruple drug resistance phenotype encompasses 37.9% of the MDR strains, of which 90% of them was resistant to metronidazole. In conclusion, these results showed a high frequency of MDR phenotypes among the studied H. pylori strains in Iran. The eradication of the H. pylori strains presenting high resistance rates to macrolides, fluoroquinolones, or metronidazole could be achieved by approved tetracycline- or amoxicillin-containing regimens as alternative regimens to standard triple therapy.

Diversity of Helicobacter pylori genotypes in Iranian patients with different gastroduodenal disorders.

AIM: To investigate the diversity of Helicobacter pylori (H. pylori) genotypes and correlations with disease outcomes in an Iranian population with different gastroduodenal disorders. METHODS: Isolates of H. pylori from patients with different gastroduodenal disorders were analyzed after culture and identification by phenotypic and genotypic methods. Genomic DNA was extracted with the QIAamp DNA mini kit (Qiagen, Germany). After DNA extraction, genotyping was done for cagA, vacA (s and m regions), iceA (iceA1 , iceA2 ) and babA with specific primers for each allele using polymerase chain reaction (PCR). All patients' pathologic and clinical data and their relation with known genotypes were analyzed by using SPSS version 19.0 software. χ² test and Fisher's exact test were used to assess relationships between categorical variables. The level of statistical significance was set at P < 0.05. RESULTS: A total of 71 isolates from 177 patients with different gastroduodenal disorders were obtained. Based on analysis of the cagA gene (positive or negative), vacA s-region (s1 or s2), vacA m-region (m1 or m2), iceA allelic type (iceA1 and iceA2 ) and babA gene (positive or negative), twenty different genotypic combinations were recognized. The prevalence of cagA, vacA s1 , vacA s2 , vacA m1 , vacA m2 , iceA1 , iceA2 , iceA1+iceA2 and babA were 62%, 78.9%, 19.7%, 21.1%, 78.9%, 15.5%, 22.5%, 40.8% and 95.8%, respectively. Interestingly, evaluation of PCR results for cagA in 6 patients showed simultaneous existence of cagA variants according to their size diversities that proposed mixed infection in these patients. The most prevalent genotype in cagA-positive isolates was cagA⁺/vacAs1m2 /iceA1 +A2 /babA+ and in cagA-negative isolates was cagA⁻/vacAs1m2 /iceA-/babA+. There were no relationships between the studied genes and histopathological findings (H. pylori density, neutrophil activity, lymphoid aggregation in lamina propria and glandular atrophy). The strains which carry cagA, vacAs1/m1 , iceA2 and babA genes showed significant associations with severe active chronic gastritis (P = 0.011, 0.025, 0.020 and 0.031, respectively). The vacAs1 genotype had significant correlation with the presence of the cagA gene (P = 0.013). Also, babA genotype showed associations with cagA (P = 0.024). In the combined genotypes, only cagA⁺/vacAs1m1 /iceA2 /babA+ genotype showed correlation with severe active chronic gastritis (P = 0.025). CONCLUSION: This genotyping panel can be a useful tool for detection of virulent H. pylori isolates and can provide valuable guidance for prediction of the clinical outcomes.