Determination of H+/K+-ATPase Activity in Human Gastric Biopsy Specimens

A reliable diagnostic test for Helicobacter pylori is important in clinical practice and research. The ideal diagnostic test for H. pylori should be sensitive, specific, and cost-effective. Helicobacter pylori resistance to clarithromycin is a common reason for failure of eradication therapy. The aim of this study was to evaluate the fluorescent in situ hybridization (FISH) method to detect H. pylori and determine clarithromycin resistance in formalin-fixed, paraffin-embedded gastric biopsy specimens. One hundred seventeen gastric biopsy specimens from patients with dyspepsia were examined for the presence of H. pylori by conventional culture, FISH, and histopathological methods. A set of fluorescent-labeled oligonucleotide probes binding to either H. pylori 16S rRNA or 23S rRNA sequences were used for FISH analysis. Phenotypic antibiotic susceptibilities of the isolates were tested using the Epsilometer test method (E test). Helicobacter pylori was detected in 70 of 117 biopsy specimens by histopathological examination and FISH, whereas it was detected in 47 specimens by culturing. Histopathology and FISH techniques failed to identify H. pylori in 1 biopsy sample isolated by culture. Clarithromycin resistance was found in 11 of 46 H. pylori isolates using the E test method. All of the phenotypic resistance measurements of isolates were correlated with genotypic clarithromycin resistance. Eleven clarithromycin-resistant strains were identified by FISH. The diagnosis of H. pylori infection and the determination of clarithromycin resistance in formalin-fixed, paraffin-embedded specimens using FISH is promising because it provides a rapid, reliable, and culture-independent diagnosis.Key words: Helicobacter pylori, clarithromycin resistance, FISH.

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Determination of Helicobacter pylori Virulence Genes in Gastric Biopsies by PCR

ISRN Gastroenterology ◽

10.1155/2013/606258 ◽

2013 ◽

Vol 2013 ◽

pp. 1-4 ◽

Cited By ~ 8

Author(s):

Tamer Essawi ◽

Wail Hammoudeh ◽

Israr Sabri ◽

Walid Sweidan ◽

Mohammad A. Farraj

Keyword(s):

Helicobacter Pylori ◽

Virulence Genes ◽

Gastric Biopsy ◽

Strong Indication ◽

Specific Primers ◽

Biopsy Specimens ◽

Pcr Identification ◽

Gastric Biopsies ◽

H Pylori

Aim. The aim of this study was to identify the presence of H. pylori in biopsy specimens from symptomatic patients by PCR. In addition, the rate of cagA, vacA, iceA1, and iceA2 virulence genes was determined. Materials and Methods. One hundred antral gastric biopsy specimens were collected during endoscopy from patients suffering from gastroduodenal symptoms. The samples were collected by the gastroenterologists in their own clinics in Ramallah, Palestine. DNA was extracted from the biopsies and subsequently used for PCR identification of H. pylori and the virulence genes using specific primers. Results. The rate of positive H. pylori in the collected biopsies was 44%. The rates of the virulence genes in this sample: cagA, vacA, iceA1, and iceA2 were 65.9%, 40.9%, 63.6%, and 84.1%, respectively. Conclusion. The iceA2 gene was the most frequent in this study. Much research is necessary to determine the presence of an association of this gene with gastric pathology. Variation in the rates of the iceA gene in different countries is a strong indication of its geographical distribution. This study would provide important information regarding the prevalence of virulence genes (vacA, cagA, iceA1, and iceA2) in H. pylori strains in the sample tested in this country.

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Direct detection of Helicobacter pylori resistance to macrolides by a polymerase chain reaction/DNA enzyme immunoassay in gastric biopsy specimens

Gut ◽

10.1136/gut.44.4.463 ◽

1999 ◽

Vol 44 (4) ◽

pp. 463-467 ◽

Cited By ~ 65

Author(s):

A Marais ◽

L Monteiro ◽

A Occhialini ◽

M Pina ◽

H Lamouliatte ◽

...

Keyword(s):

Polymerase Chain Reaction ◽

Gastric Biopsy ◽

23S Rrna ◽

Wild Type ◽

Chain Reaction ◽

Biopsy Specimens ◽

Resistant Strains ◽

Polymerase Chain ◽

H Pylori

BACKGROUNDThe increasing use of macrolides especially in the treatment ofHelicobacter pylori infection has led to an increase in resistant strains. The resistance of H pylori to macrolides, especially clarithromycin, is one of the major causes of eradication failure. In H pylori, clarithromycin resistance is due to point mutations localised in domain V of 23S rRNA.AIMTo develop a molecular technique based on amplification of a relevant fragment of the 23S rRNA and colorimetric hybridisation in liquid phase to detect directly in biopsy specimens the type of mutation associated with resistance of H pylori to clarithromycin.METHODSGastric biopsy samples from 61 patients were submitted to this test. The results were compared with standard methods (determination of minimal inhibition concentration, polymerase chain reaction/restriction fragment length polymorphism, and/or DNA sequencing) in order to evaluate the test and to define the cut off values, specificity, and sensitivity.RESULTSThe 14 biopsy samples in which H pylori was not detected did not give a positive result in any assay, and the 14 samples harbouring strains susceptible to clarithromycin gave a positive result with the wild type probe as expected. The 33 biopsy specimens containing resistant strains always gave a positive signal with one of the probes detecting resistant organisms, but in eight cases they also reacted with the wild type probe, indicating that a mixture of resistant and susceptible organisms was present.CONCLUSIONThe importance of this new assay is that it allows the detection of multiple genotypes corresponding to either heterogeneous genotypes or mixed infections. Moreover, it allows in a single step not only the detection of H pylori but also the determination of its susceptibility to clarithromycin directly in biopsy specimens without the need for culture.

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