| Objective: To investigate the drug resistance of clinical isolates from people who failed to eradicate Helicobacter pylori(Helicobacterpylori,Hp)in Xi’an Central Hospital to common antibiotics and the factors that may lead to drug resistance in Hp,and to further analyze the consistency between the detection of 23 Sr RNAV gene mutations related to clarithromycin resistance and drug sensitivity test in order to guide clinical rational drug use and improve the eradication rate of Hp.Methods: 1.Collect the gastric antrum and gastric mucous membrane samples of patients with positive 13C-UBT/14 CUBT breath test and gastroscopy who had previously eradicated Hp in Xi’an Central Hospital from October 2020 to December2022.2.The collected gastric mucosa samples were isolated and cultured under micro-aerobic conditions.for the successfully cultured strains,the colony morphology was observed by naked eye,identified under light microscope after Gram staining,and rapid urease test.the strains were identified by catalase test and oxidase test.3.The clinical isolates identified as Hp were tested for drug sensitivity to metronidazole,clarithromycin,levofloxacin,amoxicillin,furazolidone and tetracycline by E-test method.The drug resistance of Hp to common antibiotics and the changes of Hp resistance in Xi’an in different years were statistically analyzed.4.To analyze the correlation between sex,age,stomach disease,smoking,drinking,BMI and Hp resistant strains.5.The 23 Sr RNAV gene fragments of some clinical isolates were amplified by polymerase chain reaction(PCR)and sequenced,and the consistency between drug resistance gene detection and drug sensitivity test was further analyzed.6.The statistical analysis software is SPSS24.0,the counting data are expressed by statistical tables and statistical charts,and the metrological data are expressed by mean±standard deviation.Chi-square test and Fisher exact probability method are used to analyze the correlation between age,sex,BMI,smoking,drinking,stomach diseases and Hp antibiotic resistance and the changes of Hp drug resistance in different years.Kappa consistency test was used to analyze the consistency between drug resistance gene detection and drug sensitivity test,and the difference was statistically significant.Results: 1.A total of 100 patients with positive Hp isolation and culture were followed up,the positive rate was 71.43%,48 males(48.0%)and 52 females(52.0%).The average age of males and females was 50.62 ±13.31 and 47.90 ±11.16 years respectively.32(32.0%)smokers,18(18.0%)drinkers,54(54.0%)BMI < 24.00Kg/m2,46(46.0%)BMI ≥ 24.00Kg/m2,70(70%)chronic atrophic gastritis,17(17%)chronic non-atrophic gastritis and 13(13.0%)peptic ulcers.2.The resistance rates of Hp clinical isolates from 100 people in Xi’an area to metronidazole,clarithromycin and levofloxacin were 85.0%,62.0% and 38.0%,respectively,while those to amoxicillin,furazolidone and tetracycline were 6.0%,7.0%and 4.0%,respectively.3.The situation of multiple drug resistance of Hp in Xi’an is grim,and no sensitive strains to all antibiotics were found.Among the Hp isolates from 100 people,27 strains(27.0%)were resistant to one antibiotic,and the single drug resistance rate of metronidazole was the highest in 20 cases(20%).46 isolates(46.0%)were resistant to two antibiotics,of which 27(27%)were double resistant to metronidazole and clarithromycin,25(25.0%)were resistant to 3 antibiotics,and 23(23.0%)were triple resistant to metronidazole + clarithromycin + levofloxacin.Two(2.0%)of the clinical isolates developed quadruple drug resistance.4.The drug resistance of Hp to 6 antibiotics was not significantly different from that in the groups of sex,stomach disease,smoking,drinking and BMI.In the age group,the resistance rate of clinical isolates of patients aged 60 and above to levofloxacin was66.67%(12max 18),which was significantly higher than that of clinical isolates of patients less than 60 years old(41.94%).The difference was statistically significant(P <0.05).There was no significant difference in the resistance rates of metronidazole,clarithromycin,amoxicillin,furazolidone and tetracycline in age groups(P > 0.05).5.There was significant difference in the resistance rate of Hp to metronidazole and clarithromycin in 2014 and from 2020 to 2022,while the resistance rate of Hp to levofloxacin,amoxicillin and tetracycline had no significant difference between the two years.The multidrug resistance rate of Hp was also statistically different in two different years,and the multidrug resistance rate was higher than that before.6.There was no significant difference in the multidrug resistance rate of Hp clinical isolates among patients with sex,age,smoking,drinking and BMI,but in the grouping of gastric diseases,the multidrug resistance rate of clinical isolates of chronic atrophic gastritis was significantly higher than that of chronic non-atrophic gastritis and peptic ulcer group.7.In this study,50 clinical isolates of Hp were sequenced and clarithromycin resistance genes were detected in Hp isolated from gastric antrum and gastric body.All the strains with point mutations were A2143 G point mutations in 23 Sr RNAV region.Of the 50 patients,47 completed the sequencing of drug resistance genes in the gastric body,of which 42 had completed the Hp isolation and culture and drug sensitivity test in the gastric body.After the Kappa consistency test,the Kappa value was 0.837,suggesting that the two methods were in good agreement.Of the 50 patients,47 completed the detection and sequencing of drug resistance genes in the gastric antrum,of which 41 completed Hp isolation and culture and drug sensitivity test in the gastric antrum.After Kappa consistency test,the Kappa value was 0.651,suggesting that the two methods were consistent.Conclusion: 1.At present,the clinical isolates of failed sterilization of Hp in Xi’an central nosocomial infection have the highest resistance rate to metronidazole,followed by clarithromycin and levofloxacin.The resistance rate to amoxicillin,tetracycline and furazolidone is still low,and the resistance rate and multidrug resistance rate to metronidazole and clarithromycin are higher than those in previous years.Therefore,the sterilization regimen containing metronidazole,clarithromycin and levofloxacin is not recommended in the remedial treatment of Hp infection.2.People older than 60 years old with failed sterilization are prone to Hp resistance to levofloxacin,while people with chronic atrophic gastritis are prone to multidrug resistance to Hp.3.In this study,the drug resistance of Hp to clarithromycin was mainly caused by A2143 G site mutation of drug resistance gene 23 Sr RNA.The detection of clarithromycin resistance gene of Hp was consistent with drug sensitivity test,which could be used in clinical practice. |
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