| Background:Helicobacter pylori is not only an important pathogenic factor of chronic gastritis, duodenal ulcer, gastric cancer, gastric mucosa associated lymphoid tissue tumor, but also closely related to the development of chronic gastritis, duodenal ulcer and peptic ulcer in children. Helicobacter pylori infection is a global problem, and the infection rate is 50% in the whole world. As known, eradication of Hp has been proved to reduce the incidence of gastric cancer, and thus improve ulcer healing. However, due to the abuse of antibiotics in the treatment of Hp, the caused antibiotics resistance will lead a failure to Hp treatment. For the treatment of Hp, standard triple therapy is currently widely used as the main treatment option, while this therapy includes three kinds of antibiotics, namely clarithromycin, amoxicillin or metronidazole, and proton pump inhibitors. However, with the increase of clarithromycin resistance, it greatly reduces the effect of this therapy. Recent reports suggest that the potential cause for antibiotic resistance is mainly due to the genetic mutations of Hp. Therefore, detection of antibiotics resistant Hp gene mutation, could avoid the abuse of antibiotics in children. The aim of this study is to understand the initial drug resistance of Hp in children, and to study the gene mutation of antibiotics. In terms of the detection of clarithromycin resistance, sensitivity tests should be conducted including basic culture medium and the nucleic acid analysis experiment. Most of previous studies were based on the culture medium based sensitivity test, which can’t provide generic information. With the rapid development of generic techniques, it is more meaningful to apply PCR to detect Hp. In this study, we applied PCR method to detect genetic mutations between drug-resistance Hp strains and sensitive Hp strains, and thus determine the genetic information of clarithromycin, metronidazole and fluoroquinolone-resistant Hp strains.Purpose:Our study aims to investigate the relationship between genes mutation and antibiotic resistant of Hp, which can provide a theoretical basis for the profound research of childhood Hp infection.Method:Antibiotic resistance associated genes were amplified from 80 Hp strains which were isolated from children samples by using PCR method.280bpDNA fragment of 23SrDNA was amplified from clarithromycin resistance strains; rdxA and frxA genes were amplified from metronidazole resistance strains; gyr A and gyrB genes were amplified from levofloxacin resistance strains. Based on the sequencing and bioinformatic analysis of associated genes mutations, the relationship between gene mutation and antibiotic resistance was investigated.Results:Among the 80 antibiotic resistant strains,29 strains were resistant to clarithromycin with the occurrence frequency of 36.2%. There were no regular mutations of 23SrDNA gene, which focused on A2056G,T2095C and T2157C,and the rates were 90%,93% and 100%.But these mutation sites also emerged in several sensitivity strains.58 strains were resistant to metronidazole, the percentage attained 72.5%. Mutation of rdxA gene in metronidazole resistance strains focused on G47A and T184G, mutation sites of G60A, C273T and C49T were only detected in several resistance strains. Meanwhile, T184G mutation was found in metronidazole sensitivity strains. A48G mutation site of frxA gene was detected in both sensitivity strains and resistance strains.12 strains were resistant to levofloxacin with the percentage of 15%, D91Y mutation of gyrA gene occurred in resistance strains. N87K and nonsense mutations were detected both in sensitivity strains and in resistance strains.Conclusion:There were no regular mutations of 23SrDNA gene in clarithromycin resistant strains.G47A mutation of rdxA gene was found in11 metronidazole resistance strains, but no detection in sensitivity strains. D91Y mutation of gyrA gene was found in 3 levofloxacin resistance strains, but no detection in sensitive strains. |
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