Optimization of real-time PCR method to assess the direct sensitivity of clarithromycin in Helicobacter pylori

Subject Areas : Molecular Microbiology

Sadegh Ghorbani-Dalini ¹ , Mohammad Kargar ² , Abbas Doosti ³ , Pejman Abbasi ⁴ , Negar Souod ⁵

1 - Department of Microbiology, Jahrom Branch, Young Researcher’s Club, Islamic Azad University, Jahrom, Iran
2 - Department of Microbiology, Jahrom Branch, Islamic Azad University, Jahrom, Iran
3 - Biotechnology Research Center, Shahrekord Branch, Islamic Azad University, Shahrekord, Iran
4 - Department of Microbiology, Jahrom Branch, Islamic Azad University, Jahrom, Iran
5 - Department of Microbiology, Jahrom Branch, Young Researcher’s Club, Islamic Azad University, Jahrom, Iran

Received: 2013-09-16 Accepted : 2013-11-16 Published : 2009-10-01

Keywords: Helicobacter pylori, Real-time PCR, Clarithromycin,

Abstract :

Background and Objectives: The increasing rate of clarithromycin resistance in H. pylori is recognized as a significant contributing factor for treatment failure. The aim of this study was to representing a real-time PCR assay in comparison with disk diffusion method for detection of clarithromycin resistance in H. pylori. Materials and Methods: This Experimental study was performed on 45 H. pylori specimens isolated from gastrointestinal disorders patients. In all samples, presence of H. pylori was confirmed by using culture, rapid urease test and conventional PCR. Also, clarithromycin resistance were assessed by using CLSI (Clinical and Laboratory Standard Institute) protocol and real-time PCR using specific primers and probe for H. pylori peptidylteransferas region of 23S rRNA gene. Results: Disk diffusion method identified 20 (44.44%) samples as susceptible and 25 (55.56%) samples as resistance to clarithromycin. But, real time PCR were identified samples as 20 (44.44%) with susceptible genotype, 5 (11.11%) with mix genotype and 20 (44.44%) as resistance to clarithromycin genotype. Also results showed that sensitivity of this assay was equal to 40 bacteria or 80 copy number of 23S rRNA gene. Conclusion: Results showed that Real-time PCR assay has high accuracy for identifying clarithromycin resistance in short time.

References:
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