Study On The Mechanism Of High-level Resistance To Aminoglycosides In Acinetobacter Baumannii Isolates

Objective:To survey the drug resistance in Acinetobacter baumanniiclinical separations to 21 commonly used antimicrobial agents in the northeastof Si chuan ; To analyze the distribution and effects of seven 16 S rRNAmethylase genes, three aminoglycoside modifying enzymes genes and onekind of efflux pump gene in Acinetobacter baumannii with high-levelresistance to aminoglycosides or multiple resistance in order to providesignificant experimental data for the treatment of Acinetobacter baumanniiinfection diseases.Methods: The minimum inhibition concentrations (MICs) of21Antimicrobial drugs against to clinical isolates of 110 Acinetobacterbaumannii were determined by agar dilution on the basis of CLSI, then thesestrains were divided into two groups based on MIC≥256μg∕ml ofamikacin：high-level resistance to aminoglycosides（60 strains）and low -levelresistance(50 strains) .The 16S rRNA methylase genes (armA, rmtA，rmtB，rmtC，rmtD，rmtE,and npmA)，aminoglycoside modifying enzymes genes (aac (6’)-Ib,ant(3’’)-Ia, aph(3′)-I) and efflux pump gene（adeB）were analyzed bypolymerase chain reaction (PCR). Using RT-PCR methord to determin themRNA expressions of genes in some strains, and analyze the difference ofgenes positive rates and mRNA expression between two groups.Results: Of the 110 Acinetobacter baumannii isolates, 50.1%-83.8% ofisolates were resistant to aminoglycosides and 46.4%-65.4% were highly resistant (MIC≥256μg/mL). The resistant rate of Polymyxin B, Imipenemand Meropenem was 0, 8.2% and 9.1% respectively. The other durgs werevariable ( 43.6%-93.6%). In the high-level resistance to aminoglycosidesgroup, the armA, aac (6’)-Ib, ant(3’’)-Ia and aph(3′)–I genes were present in66.7%, 51.7%, 81.7% , and 58.3% of the 60 clinical isolates respectively,butin the other group, the aac (6’)-Ib，ant(3’’)-Ia，and aph(3′)–I genes werepresent in 4%, 6% and 4% only, the genes positive rates were significantlydifferent（p<0.05）in two groups. It was shown by RT-PCR that in the high-level resistance group, all the selected strains were amplified with the purposegene. However only parts of purpose genes were amplified in the low-levelresistance group . There were 66 strains with efflux pump adeB and thepositive rate was 60%; The multidrug resistant strains occupied 92.4%( 61isolates)among the strains with adeB .However , there were only 5 strains inrelatively sensitive group. RT-PCR analysed the expression of adeB in twogroups .The differences of mRNA expression levels were statistically signif-icant (p < 0.05).Conclusions:1. Acinetobacter Baumannii clinical isolates were widely resistant to theantibacterial agents with the most commonly used, and especially someaminoglycosides were high-level resistant in the northeast of Sichuan.2. The most active agents against Acinetobacter baumannii infections areMinocycline,Imipenem Polymyxin B and Meropenem.3. The 16 SrRNA methylase enzymes and aminoglycoside modifyingenzymes genes were found frequently in acinetobacter baumannii clinicalisolates and played an important role in the high-level resistance toaminoglycoside antibiotics. 4. The high expression levels of adeB gene are correlated withaminoglycosides resistance and multiple drug resistance in Acinetobacterbaumannii.