Study On Reversal Of Resistance In Acinetobacter Baumannii By Efflux Pump Inhibitors

Objective:To investigate drug resistance in Acinetobacter Baumannii clinical isolates to 22 commonly used antimicrobial drugs in northeast Sichuan areas and to determine the relationship between the distribution of efflux pumps gene-adeB and the multidrug resistance profile. The effects of efflux pump inhibitors(EPIs) on reversal of drug resistance in Acinetobacter baumannii were further identified for the development of efflux pump inhibitors and the treatment of multiple drug-resistant infections.Methods:A total of 116 non-duplicate clinical isolates of Acinetobacter baumannii were recovered and antimicrobial susceptibility was determined by disk diffusion and agar dilution. The antimicrobial resistance related gene-adeB in the isolates was detected by regular polymerase chain reaction(PCR). The isolates were treated using cefazolin, norfloxacin, chloramphenicol, gentamicin and tetracycline as efflux substrates with efflux pump inhibitors including reserpine (40μg/ml), verapamil (80μg/ml), omeprazole (80μg/ml) and carbonyl cyanide-m-chlorophenylhydrazone (CCCP) (20μg/ml) and antimicrobial susceptibility was determined. The effects of efflux pump inhibitors on reversal of drug resistance in adeB positive or negative Acinetobacter baumannii isolates were analyzed and the efflux effects in the Acinetobacter baumannii isolates were further determined using Rhodamine 6G as the tracer.Results:Acinetobacter Baumannii clinical isolates were mainly isolated from sputum (72.4%) in ICU (32.8%), department of respiratory medicine (16.4%) and department of burn (10.3%). Among 22 kinds of antimicrobial agents, the sensitivity of polymyxin was up to 100% and Rifamycin had the highest resistance rate (97.4%). The resistance rates of ampicillin, cefazolin, cefoxitin and aztreonam were more than 90%. Meropenem (1.7%) and imipenem (11.2%) showed the lowest resistance rate. Of the 116 isolates, 89 strains were multiple drug resistant (76.7%). The adeB gene was identified in 74 strains of bacteria (63.8%), including 72 strains of multiple drug resistant bacteria. Susceptibility studied showed that 87 strains (75%) had positive efflux with treatment of gentamicin with CCCP. The treatment of reserpine with chloramphenicol, omeprazole with chloramphenicol and verapamil with gentamicin showed positive efflux ratios of 46.6%, 33.6% and 17.2%, respectively. The efflux inhibition effects of reserpine, CCCP and omeprazole in adeB positive bacteria were much higher than those of adeB negative bacteria. The accumulation levels of Rhodamine 6G in multidrug resistance isolates also were much lower than those of relatively sensitive isolates.Conclusions:1. Acinetobacter Baumannii clinical isolates in northeast Sichuan areas were extensively resistant to most commonly used antimicrobial agents.2. The detection ratio of adeB gene in Acinetobacter Baumannii is correlated with multiple drug resistance.3. Active efflux of drug is one of the most important multiple drug resistant mechanisms of Acinetobacter Baumanni. The efflux pump inhibitors including reserpine, verapamil, omeprazole and CCCP can partially reverse the resistance of Acinetobacter baumannii. Consistent with susceptibility studies, the specificities of efflux pump inhibitors on substrates are obviously different, and CCCP has the best efflux effects in Acinetobacter baumannii. 4. Using Rhodamine 6G as the model efflux tracer, the efflux amount of multiple drug resistant strains are significantly higher than that of the relatively sensitive strains, which is correlated with the multiple drug resistance.