Determination of Helicobacter pylori primary resistance to levofloxacin in gastric mucosal biopsy samples using real-time polymerase chain reaction

Abstract

Objective. To study the primary resistance of Helicobacter pylori (H. pylori) to levofloxacin in residents of Gomel region by real-time polymerase chain reaction (RT PCR).Materials and methods. The study included 170 patients diagnosed with gastritis and duodenitis, K29, median age - years (25% and 75% 37 and 61 years). According to the questionnaire data of the patients, eradication therapy with levofloxacin was not performed for them. To determine the resistance of H. pylori to levofloxacin we used RT PCR. Results. Out of 170 DNA samples analyzed, 8 samples had doubtful results and according to the methodology for recording the results are subject to rearrangement from the DNA isolation stage. The remaining 162 samples were positive for the β-actin gene (internal control sample ICS) and were taken into account in further analysis (Ct, CY5 19.6-27.4). 16sRNA gene DNA (Ct, ROX 19.5-30.04), indicative of bacterial infection, was confirmed in 152 samples (93.8%). DNA of the gyrA gene (point mutations A259T, T261C, G261A, G271A, G271T and A272G) was detected in 19 of 152 DNA samples, and H. pylori resistance to levofloxacin was 12.5 %, (Ct, Hex 23.2-30.7). The positive control samples had characteristic curve growth on the corresponding detection channels, while the negative samples showed no curve growth.Conclusion. Primary resistance of H. pylori to levofloxacin in residents of Gomel region amounted to 12.5%. Mutations of gyrA gene are the most sensitive marker for predicting successful eradication when using fluoroquinolones, in particular levofloxacin. RT PCR is a reliable method of mutation detection and allows simultaneous detection of H. pylori DNA and resistance to levofloxacin, which significantly reduces the study time.