Direct determination of Helicobacter pylori vacA genotypes and cagA gene in gastric biopsies and relationship to gastrointestinal diseases (original) (raw)
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Diagnostic microbiology and infectious disease, 2008
Several tests/methods for the infection, detection, and genotyping of Helicobacter pylori have been developed in the past; all these differ in specificity and sensitivity and thereby could not be routinely used in clinical study especially in resource-poor developing countries. In the present study, a novel method based on multiplex polymerase chain reaction (PCR) assay was developed to detect H. pylori in patients suffering from gastrointestinal diseases. This method does not require steps of sonication, boiling, or centrifugation for obtaining DNA from biopsy samples, which are otherwise prerequisite in detecting H. pylori by PCR assay. Two hundred seventy-six patients were examined, of which 182 cases (excluding 36 patients having multiple H. pylori strain infection and 8 showing amplification of 16S rDNA only) were H. pylori positive. The dominant vacA genotype was s1 and m1 being present in 168 (92.3%) and 106 (58.2%) patients, respectively, followed by m2 (41.7%), and the lowest being s2 (7.7%). Detection of H. pylori by this method seems rapid, simpler, and suitable for both types 1 and 2 bacteria. Furthermore, genotyping of vacA and cagA genes could also be routinely performed in a large number of patients for diagnostic purposes.
Basrah Journal of Surgery, 2010
This study aimed to learn the incidence of Helicobacter pylori infection in patients with various gastroduodenal endoscopic lesions and the frequency of virulence H.pylori associated genes CagA and VacA in these patients. One hundred seventy six patients (96 males and 80 females) attending endoscopy units for various dyspeptic symptoms were studied. Antral biopsies were obtained to detect H.pylori by rapid urease test, culturing and histopathologic examination. Twenty five patients with positive H.pylori isolates who were found to be mannose resistant, were tested for cytotoxic associated (CagA) and vacuolating cytotoxin A (VacA) genes. Among studied patients, positive H.pylori detected by rapid urease test, culturing and histopathologic examination (from 50 patients only) were 113 (63%), 127 (71%) and 25 (50%) respectively. Out of 25 patients with positive H.pylori isolates who were found to be mannose resistant, positive genes of either CagA or VacA were detected in 18 (72%) patients with positive isolates, while positivity of both genes were detected in 13(52%) patients with positive isolates. Five (45.4%) and 5 (45.4%) out of patients with duodenal ulcers and gastritis respectively were positive for both (CagA) and (VacA) genes. In conclusion, the highest detection rate of H.pylori infection was by bacterial culture. A correlation between CagA and VacA genes and endoscopic lesions of duodenal ulcers and gastritis was found.
Diyala Journal For Pure Science
The objective of current study was to detect the prevalence of Helicobacter pylori by identifying 16SrDNA and to determine the virulence genes (ureA, cagA and vacA) in biopsy specimens from patients suffering gastroduodenal disease using polymerase chain reaction (PCR). Forty samples were obtained by gastroenterologists during endoscopy from gastric antral of suspected individual attending endoscopy unit at Baqubah Teaching Hospital, Diyala, Iraq, during the period from September 2015 to February 2016. According to the endoscopic finding the patients were allocated into four groups of gastroduodenal diseases and control, which include gastritis (GS), duodenal ulcer (DU), gastric ulcer (GU), gastric cancer (GC), their rates were 30% (12), 20% (8), 17.5% (7), 7.5% (3) and 25% (10), respectively. DNA was extracted from the biopsies and subsequently used for PCR detection of H. pylori and the virulence genes using specific primers. The results shows that 60% of samples were positive for H. pylori, of these positive samples, 91,66%, 66.66%, and 48.83%, were shown to have the virulence genes, ureA, cagA, and vacA, respectively. It is important to mention that cagA shown the highest prevalence rate in gastric cancer cases in comparison with vacA gene. further studies are required to study the link between cagA gene and Detection of, cagA and vacA Helicobacter pylori Virulence Genes in Gastric Biopsies of patients with Gastroduodenal disease using Polymerase chain reaction (PCR) technique
Jundishapur Journal of Microbiology, 2015
Background: Helicobacter pylori infection and related diseases outcome are mediated by a complex interplay between bacterial, host and environmental factors. Several distinct virulence factors of H. pylori have been shown to be associated with different clinical outcomes. Here we focused on vacA and cagA genotypes of H. pylori strains isolated from patients with gastric disorder. Objectives: The aim of this study was to determine the frequency of two toxins and genotypes of VacA toxin in patients referred to a central hospital in the west of Iran (Imam Reza hospital, Kermanshah) during 2011-2012. Patients and Methods: Samples were collected from patients infected with H. pylori. Gastric biopsy specimens from the stomach antrum and corpus were cultured. PCR analysis was performed for genotyping H. pylori vacA and cagA genes. Results: Helicobacter pylori was isolated from 48% (96/200) of patients with gastroduodenal disorders. In 81/96 (84%) cases, the cagA gene was present. Among different genotypes of vacA, two s1m2 and s2m2 genotypes were dominant with frequency of 39.5% and 50%, respectively. The frequency of the s1m1 genotype was 7.2% (7/96), which is much lower than elsewhere. H. pylori isolates with positive results for cagA gene and vacA s1m2 genotypes showed statistically significant correlation with peptic ulcer (s1m2 13/34 [38.2%] P = 0.003). However, isolates of H. pylori infection with cagA gene and vacA s2m2 genotypes were significantly associated with development of gastritis (s2m2 41/42 [97.6%] P = 0.000). Conclusions: About 90% of H. pylori strains potentially contained vacA s2m2 and s1m2 genotypes. Infection with H. pylori strain containing the cagA gene or the vacA s1m1 and s1m2 genotypes was associated with increased incidence of peptic ulcer disease (PUD).
2011
The vacuolating cytotoxin VacA and cytotoxin associated gene product CagA, encoded by vacA and cagA are major virulence determinants associated with pathogenesis of Helicobacter pylori. The presence and prevalence of two major H. pylori virulence associated genes among gastric biopsies of Pakistani children were investigated in the current study. Fifty one gastric biopsy specimens of children were analysed for 16S rRNA, vacA and cagA genes using PCR. The results showed that 21 (41.2%) biopsies were positive for H. pylori as determined by 16S rRNA PCR. In the 21 H. pylori positive gastric biopsies, 19 (90.5%) showed vacA s1a, 1 (4.75%) was vacA s1b and 1 (4.75%) was vacA s2 whereas, 5 (23.8%) were vacA m1 and 16 (76.2%) were vacA m2. None of the H. pylori positive biopsies carried vacA s1c subtype. The cagA gene was found in 13 (61.9%) of H. pylori infected biopsies and different vacA combinations were found with or without cagA gene. H. pylori was detected with high frequency of cagA while vacA s1a and vacA m2 regions with vacA s1a/m2 genotype were predominant in H. pylori infected gastric biopsies of children.
Egyptian Journal of Medical Human Genetics, 2017
Background: Helicobactor pylori (H. pylori) virulence markers would be useful to predict peptic ulcer disease (PUD) or gastric cancer. Aim: In Egypt, since inadequate data are present regarding H. pylori virulence-related genes in different age group patients with gastro-duodenal diseases, it becomes crucial to study the clinical status of cagA, vacA and iceA1 genotypes of H. pylori strains recovered from patients with dyspepsia. Subjects and methods: The study included 113 dyspeptic patients who were exposed to upper gastrointestinal endoscopic examination. Four antral biopsies were obtained from each patient for the analysis of H. pylori infection by rapid urease test and detection of 16S rRNA. Results: Sixty (53.1%) patients were confirmed to be infected with H. pylori. Upon endoscopy, gastritis was revealed in 27 patients (45%) and10 patients (16.7%) had PUD. Of the 60 H. pylori strains, 39 (65%) had at least one virulence gene. Six different genotypic forms were recognized; vacA (9/60), iceA1 (1/60), vacA/cagA (7/60), vacA/iceA1 (13/60), vacA/cagA/iceA1 (8/60) only one of cagA/iceA type and we could not detect cagA. The overall vacA, iceA1and cagA genes identified were 61.6%, 38.8%, 26.6% respectively, by PCR-based molecular testing. The vacA gene status was highly significant related to gastritis patient (P 0.036). The vacA s1m1 and s2m2 alleles were significantly found in 50% of H. pylori infected patients with PUD and with gastritis 57.1% respectively (P 0.01). Conclusion: In conclusion, the main genotype combinations in the studied Egyptian patients were; vacAs2m2/iceA1, vacAs1m1/cagA, mostly associated with gastritis, and vacAs1/cagA/icA, mainly in PUD. The less virulent (s2, s2m2) H. pylori genotypes were found in patients aged over 43 years.
Malaysian Journal of Microbiology, 2011
The vacuolating cytotoxin VacA and cytotoxin associated gene product CagA, encoded by vacA and cagA are major virulence determinants associated with pathogenesis of Helicobacter pylori. The presence and prevalence of two major H. pylori virulence associated genes among gastric biopsies of Pakistani children were investigated in the current study. Fifty one gastric biopsy specimens of children were analysed for 16S rRNA, vacA and cagA genes using PCR. The results showed that 21 (41.2%) biopsies were positive for H. pylori as determined by 16S rRNA PCR. In the 21 H. pylori positive gastric biopsies, 19 (90.5%) showed vacA s1a, 1 (4.75%) was vacA s1b and 1 (4.75%) was vacA s2 whereas, 5 (23.8%) were vacA m1 and 16 (76.2%) were vacA m2. None of the H. pylori positive biopsies carried vacA s1c subtype. The cagA gene was found in 13 (61.9%) of H. pylori infected biopsies and different vacA combinations were found with or without cagA gene. H. pylori was detected with high frequency of cagA while vacA s1a and vacA m2 regions with vacA s1a/m2 genotype were predominant in H. pylori infected gastric biopsies of children.
Indian Journal of Gastroenterology, 2009
Background The vacuolating cytotoxin and the cytotoxinassociated protein, encoded by vacA and cagA, respectively, are important virulence determinants of Helicobacter pylori. Objective The aim of this study was to perform vacA genotyping and evaluate its association with cagA genotype and clinical outcome. Methods One hundred and twenty H. pylori strains were isolated from dyspeptic patients (29 with peptic ulcer, 91 with non-ulcer dyspepsia). Genotype was determined by PCR. Results Seventy-nine (66%) of 120 strains had the vacA signal sequence genotype s1 and 41 (34%) had the type s2. The vacA middle-region types m1 and m2 were detected in 56 (47%) and 64 (53%) strains, respectively. The combinations s1-m1 (n=56 [47%] and s2-m2 (41 [34%]) occurred more frequently than s1-m2 (23 [19.2%]; p=0.001). No strain with the combination s2-m1 was found. All patients with peptic ulcers harbored type s1 strains compared to 75 (82.4%) of 91 patients with non-ulcer dyspepsia (p=0.01). The vacA genotype s1 was associated with the presence of cagA (p <0.0001). The cagA gene was detectable in 38 (31.6%) of 120 isolates and present in all 29 patients with ulcer compared to nine of 91 with non-ulcer dyspepsia (p <0.001). Conclusion Helicobacter pylori strains of vacA type s1 and the combination of s1-m1 were associated with peptic ulceration and the presence of cagA gene.
Journal of Applied Microbiology, 2007
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.