Study Of Plasmid-mediated Quinolone Resistance

Quinolones is the first complete synthesis antimicrobial agents. Because of their strongly activity against gram-negative and gram-positive bacteria, including anaerobes, they are extensively used in various types of infections and in the veterinary environment together with other antibacterial agents. Extensive use of them has generated the higher bacteria resistance. The resistant mechanism is quite complex. To date, there are two main mechanisms implied in this resistance (both by elements chromosomal): alterations in the targets of the quinolones and decrease in the accumulation of the antibiotic in the bacterial interior by making the membrane impermeable (loss of porins or alterations of the lipopolysaccharide) or by expression of active expulsion systems. In 1998, horizontally transferable resistance to quinolones was described for the first time. The qnr gene is genetically responsible for resistance and is found inside a mobile element. The horizontal dissemination of mechanisms resistant to quinolones opens up the possibility of a rapid expansion of resistance to such antimicrobials, in animal as well as human pathogens, and even more so with the extensive use made of quinolones.This study analyzed drug resistance of the clinical isolates of Escherichia coli and Klebsiella pneumoniae from 2000 to 2003 in the Affliated Hospital of Anhui Medical University and investigate the resistant rates of common antimicrobial agents. We found the resistant rate of ciprofloxacin is higher than others reports and the resistant rate of ciprofloxacin of ESBLs producing strains was significant higher than ESBLs nonproducing strains. In order to study the molecule mechanism of this difference, we studied the qnr gene of the clinical isolates of Escherichia coli and Klebsiella and detected the spectrum beta-lactamases and extended spectrum beta-lactamases in qnr gene positive strains.OBJECTIVE To investigate the resistance in clinical isolates of Escherichia coli and Klebsiella pneumoniae for reasonable antimicrobial agents, and to analyze incidences of multi-drug resistance between ciprofloxacin and other common clinical antimicrobial. METHODS The drug sensitivity of 1414 clinical isolates of Escherichia coli and 392 clinical isolates of Klebsiella pneumoniae were tested by micro-dilution method. Extended spectrum beta-lactamases(ESBLs) were screened by micro-dilution method and confirmed by ceftazidime, ciftazidime-clavulanic acid and cefotaxime, cefotaxime-clavulanic acid. RESULTS Resistant rates of E.coli over 30% added up to 10 among the 18 tested antimicrobial agents. Resistant rates of Escherichia coli against ampicillin, ampicillin-sulbactam, cephalothin, piperacillin, ciprofloxacin, trimethoprim-sulfamethoxazole, gentamicin, tobramycin were more than 50%, respectively. The lowest of resistant rates was imipenem(0%) and the second was piperacillin-tazobactam (5.2%). Resistant rates of E.coli against cephalosporins tended to increasing. Incidences of ESBLs producing strains of Escherichia coli were from 25.5% to 39.8%. The average was 32.7%. Resistant rates of ESBLs producing strains of E.coli to most antimicrobial agents were much higher than ESBLs nonproducing strains (p<0.05). Resistant rates of Klebsiella pneumoniae over 30% added up to 11 among the 18 antimicrobial agents. Resistant rates of Klebsiella pneumoniae against ampicillin, ampicillin-Sulbactam, cephalothin, piperacillin, trimethoprim-sulfamethoxazole, cefacidal were more than 50%, respectively. Resistance rates of four antimicrobial agents were lower than 10 percentage, and they were ceftriaxone(7.7%), cefetaxime(7.4%), aztreonam(6.9%), imipenem(0%). Incidences of ESBLs producingstrains of Klebsiella pneumoniae were from 32.9% to 45.8%. The average was 39.8%. Resistant rates of ESBLs producing strains of K.pneumoniae against most of antimicrobial agents were much higher than ESBLs nonproducing strains (p<0.05). CONCLUSION Resistant rates of clinical isolates of Escherichia coli and Klebsiella pneumoniae were rather high and most multidrug-resistance, especially in ESBLs producing strains. We should give more attention to the surveillance of antimicrobial resistance in these isolates and prevent transmission and epidemic of those resistant strains.Part II Detection of qnr Gene in Escherichia coli and KlebsiellaAbstract: OBJECTIVE To detect qnr gene in Escherichia coli and Klebsiella from the First Affiliated Hospital of Anhui Medical Unversity, and to study the mechanism of resistance and multi-drug resistance in qnr gene positive strains. METHODS Primers were designed by Primer Premier 5.0. Purpose genes were amplified by PCR. Antimicrobial susceptibility testing was detected by K-B methods breakpoint methods and agar dilution methods, respectively. Plasmid DNA is isolated from small-scale (1-2 ml) bacterial cultures by treatment with alkali and SDS. Preparation and Transformation of Competent E. coli Using Calcium Chloride. RESULTS Of the 227 E.coli and 36 Klebsiella strains, qnr gene were found in 6 clinical isolates that included 2 E.coli, 1 K.oxytoca and 3 K.pneumoniae. Mutation of qnr gene was found in K.oxytoca, inducing 36 bases and 4 amino acids. The sequence was submitted to the Genebank database (accession numbers AY675584). There are 6 Strains with qnr gene were ESBLs producing and multidrug-resistance strains. CONCLUSION In strains with qnr...