| Objective:This study is aimed to investigate the antibiotic resistance of Helicobacter pylori(Hp)isolated from patients in South-Sichuan to amoxicillin,clarithromycin,metronidazole,levofloxacin,tetracycline and to further analyze mutations in the domain V of 23S r RNA gene that are related with clarithromycin resistance to guide clinical rational drug use and improve Hp eradicate rate.Methods:1.To collect 198 cases of gastric mucosa specimens in the affiliated hospital of southwest medical university patients who undergoing gastroscopy of gastrointestinal symptoms and rapid urease test being positive from May to December,2017.2.The analysis of the biopsies bacterial culture in micro aerobic condition and after culturing the bacteria for 3-14 days,we tested Gram staining,oxidase,urease,and catalaseactivity.3.Cultures that were positive for Hp were tested for their susceptibility to five antibiotics(clarithromycin,amoxicillin,tetracycline,metronidazole,and levofloxacin)by E-test.4.To compare the resistance of Hp strains isolated by different genders,ages and diseases to the above five antibiotics.5.According to the results of drug susceptibility test screened 23 clarithromycin resistant strains,9 sensitive strains and 2 control strains and then PCR was used to amplify domain V of 23S r RNA gene and sequenced.6.Using DNAstar software,nucleotide homology of these sequences were compared to Hp 26695(Gen Bank accession number:NR076155.1)correlation sequences which was a susceptible strain.Results:1.A total of 80 Hp strains were isolated from 198patients recruited,the positive rate was 40.4%(80/198).Among them males 46cases and females 34 cases,with a male/female ratio of 1.35:1.Male average age(51.15±13.09)years,female average age(51.82±12.17)years,F=0.42,P=0.52>0.05,the difference was not statistically significant.2.According to the gender,diagnosis and rapid urease comparison the culture positive rate found that rapid urease reaction(+++)were significantly higher than the positive rate of(+),(++),the difference was statistical significance(χ~2=22.95,P<0.05).3.The resistance to metronidazole,levofloxacin,clarithromycin,tetracycline and amoxicillin were 86.25%(69/80),67.50%(54/80),28.75%(23/80),7.50%(6/80),and 5.00%(4/80),respectively.4.The situation of multiple drug resistance in the region is severe.Only 3.75%strains were sensitive to all antibiotics,30.00%strains were single resistance,66.25%were resistant to more than one antibiotic in 80 strains of this study.5.The double drug resistance rates of clarithromycin and metronidazole were an high as 26.25%(21/80).6.The average MIC of Hp drug-resistant strains increased with the increase of drug resistance species.7.There was no significant difference in the resistance rate of the five antibiotics of Hp for different genders and ages(P>0.05).8.The resistance rates to clarithromycin was significantly higher in strains isolated from patients with gastritis compared to those with peptic ulcer.(χ~2=4.1,P<0.05).9.In this study,there were three types of base mutation in 32 strains of South-Sichuan region,including A2144G,A2224G and T2183C.The analysis of mutations in the Domain V of 23S r RNA only found in resistant strains were A2144G and A2224G,accounting for 78.3%(18/23)and 34.8%(8/23).T2183C mutation was found in both clarithromycin resistant strains and susceptible strains.Conclusion:1.The drug resistance situation of Hp in South-Sichuan is serious.The Hp resistant rates to metronidazole,levofloxacin,and clarithromycin were high,and to amoxicillin,tetracycline were lower,so we need to choose eradication therapy containing amoxicillin and tetracycline in clinical work.2.A2144G and A2224G mutations are the most prevalent mutation types related with clarithromycin resistance in South-Sichuan.3.Importantly,owing to the high clarithromycin resistance in our area,when we empirical choice of treatment program,the standard triple therapy should be abandoned as the first-line treatment,and a 14-day course of the bismuth quadruple therapy(BQT)shuold be used for first-line.4.This study provides a theoretical basis for the rational selection of regimen of Hp eradiction,especially choose antibotics to eradicate Hp infection and improve the eradication rate in South-Sichuan,is of important practical value. |
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