Determining the clarithromycin-resistance of helicobacter pylori in first-line therapy with melting curve analysis

Abstract

Aims: We aimed to determine the usefulness of melting curve analysis with the PCR technique in predicting the primary clarithromycin resistance of H. pylori before antibiotic use. Methods: 46 H. pylori positive clarithromycin based therapy naïve patients were included in the study. After proving H. pylori positivity with a real-time test, histopathological examination and rapid urease test, patients were treated with clarithromycinbased triple therapy for 14 days. Then patients were underwent a control endoscopy and tissue samples were taken 8 weeks after the initial endoscopy. The presence of H. pylori was investigated by using TaqMan-based real-time PCR and clarithromycin resistance was determined using two different molecular methods:melting curve analysis and DNA sequencing. Results: In total 18 patients (39.1%) were diagnosed as H. pylori positive on histopathological examination and 26 (56.5%) were diagnosed as H. pylori positive with TaqMan-based PCR, after triple therapy. According to results of melting curve analysis, primary resistance was found in 4 samples before treatment and secondary resistance was found in a different 4 samples after treatment. The DNA analysis showed that 9 patients had primary mutations in the pre-treatment group:4 patients had A2142CA2143G(T), 3 patients had T2182C, and 2 patients had both mutations, and also showed that 6 patients had mutations in the post-treatment group:4 patients had A2142C-A2143G, 2 patients had T2182C mutations. Conclusion: PCR based melting curve analysis may fail in determining clarithromycin resistance before eradication therapy. Hence, this method may also be used in combination with another molecular method such as DNA sequence analysis for improved detection of clarithromycin resistance.