[Beta-lactam resistance in aquatic Enterobacter cloacae strains using phenotypic and genotypic criteria].

Bacterial resistance by producing of beta-lactamases represents an increasing problem of infections chemotherapy. beta-lactam hydrolyzing activities are detected in virtually all bacteria, from witch Enterobacter cloacae produce chromosomal beta-lactamases included in inducible class AmpC beta-lactamases. The purpose of this preliminary study was to investigate the antibiotic susceptibility of 7 inducible beta-lactamase-producing Enterobacter cloacae strains isolated from aquatic sources (river and polluted waters). The identification to the species level was performed with the API 32E system and susceptibility to antimicrobial agents was tested by the disk diffusion method according to NCCLS recommendations. The following antibiotics were tested: ampicillin, amoxycillin/clavulanic acid, ceftazidime, cefotaxime, cephalotin, cefamandole, cephaclor, imipenem, amikacin, gentamycin, kanamycin, tobramycin, ciprofloxacin, norfloxacin, ofloxacin, nalidixic acid, tetracycline and chloramphenicol. Minimum inhibitory concentrations (MICs) were performed using dilution method in Mueller-Hinton broth with a 0.06-64 micrograms/ml concentration range for all antimicrobials and bacterial inoculum of about 1-2 x 10(8) cfu/ml. For the data analysis NCCLS breakpoints for resistance and sensitivity were used. Interaction of beta-lactamase inhibitor clavulanate with cefotaxime was performed by double-disk synergy test. Detection of inducible beta-lactamase expression was performed by the inductibility disk diffusion test using cefotaxime, ceftazidime and imipenem. Genomic DNA was isolated using CTAB technique and bacterial plasmid isolation was performed by an alkaline lysis method. Genetic characterization was performed by agarose gel electrophoresis and spectrophotometric analysis. The majority of examined E. cloacae strains were sensitive to imipenem, cefamandole, amikacin and quinolones (norfloxacin and ofloxacin), a higher moderate resistance being observed only to nalidixic acid (higher than 50%) and ciprofloxacin (15%). The percentage of resistant strains ranged from 72% (to kanamycin) to 87% (to gentamicin). The best results (resistance about 99%) were obtained with ampicillin, amoxycillin/clavulanic acid, ceftazidime, cefotaxime, cephalotin, cephaclor, tobramycin, tetracycline and chloramphenicol. The disk diffusion tests showed the absence of extended-spectrum beta-lactamases production and the expression of inducible beta-lactamases. Electrophoretic patterns point out the presence of plasmid DNA. Plasmid profile revealed the presence of several different plasmids ranging from 2.5 kpb to more than 30 kpb. The presence of inducible beta-lactamase E. cloacae strains in aquatic media (river and polluted waters) and the closely related pattern of susceptibility among these strains reflect the possible contamination of these sources and the common origin of them.