A Study On Antimicrobial Activity And Persistent Effects Of Combinations Consisted Of β-Lactams And β-Lactamase Inhibitors Against β-Lactamase-producing E.coli And K.pneumonia

BackgroundsIt is well known to all that the β-lactams (penicillins, cephalosporins, and atypical p-lactams) are at the lead position in antibiotic market in the world. Because of the abuse of β-lactams, bacterial resistance has become a growing phenomenon. The main mechanisms of bacterial resistance include the production of inactivating enzymes, the change of targets structure, decrease of permeability in outer membrane, abnormalities in active efflux and forming of bacterial biofilm. The production of β-lactamases is still the main mechanism for resistance of Gram-negative bacilli to β-lactams, which are hydrolyzed by β-lactamas. The spread of bacterial resistance induced by β-lactamase production has led to the use of β-lactamase inhibitors. The inhibitors have strong affinity with β-lactamases, protecting β-lactams from hydrolysis. It is important to consider the pharmacokinetic/pharmacodynamic profiles of these combinations consisting of β-lactams and β-lactams inhibitors. Recently the research of the PAEs of β-Lactams/β-lactamase inhibitors combinations has attracted attention around the world. PLIE was described for the first time in 1992 by Thorburn and confirmed later by others. So far, PLIE has not been investigated in China. The research of PASME in vitro analogues the course of pharmacodynamics in vito, soPASME should be a better pharmacodynamic parameter than the others. Based on the views above, the aims of this work were to study the antibacterial activities of β-lactams/β-lactamase inhibitors combinations on β-lactamase-producing Gram-negative bacilli in vitro, and PAEs, PLIEs and PASMEs of these combinations were also measured. These combinations included cefoperazone with clavulanic acid, sulbactam, tazobactam, respectively. Piperacillin was the same as cefoperazone. We tried to provide useful data for the evaluation of the new antibiotics and the determination of the optimal dosing regimen, to control the infection of β-lactamase-producing strains. ObjectsTo investingate the antibacterial activity of combinations of β-lactams and β-lactamase inhibitors against β-lactamase-producing Gram-negative bacilli in vitro, draw Time-Kill curves of combinations, obtain the minimal saturation bactericidal concentration, observe whether or not the change of the antibacterial activity depending on the time or concentration, compare PAEs, PLIEs, PASMEs of combinations with β-lactams, and to evaluate the value of persistant effects of combinations in clinical administration. Material and methodsThe antimicrobial agents: cefoperazone, piperacillin, clavulanic acid, sulbactam, tazobactamOrganisms: TEM-1 -producing strian of Escherichia coli ATCC35218, ESBL-producing strian of Klebsiella pneumonia ATCC700603, β-lactamase-negative strian of Escherichia coli ATCC 25922 Inocula: For all the stains, the broth cultures incubated at 35℃ for 2⁶h were determined by VITEX Colorimeter, and then were diluted in Mueller-Hinton broth to obtain a starting inoculum with concentration between 10⁶ and 10⁷ CFU/mL. MICs determinations: MICs were determined in Mueller-Hinton broth byusing a two-fold serial dilution.PAE, PLIE and PASME determinations: Organisms were exposed to antibiotics at ten times the MICs in Mueller-Hinton broth for 2h. Then the bacteria were washed and resuspended in drug-free medium to determine the PAE; and the medium containing cefoperazone or piperacillin alone with the same concentration as before was used to determine the PLIE, and the medium containing combination of 0.1 times MIC(T₁), 0.2 times MIC(T₂), or 0.3 times MIC(T₃) to determine the PASME. The bacteria in control tube for PAE were exposed to drug-free medium before and after centrifligation, the bacteria in control tube for PLIE were exposed to cefoperazone or piperacillin alone at the same concentration before and after centrifugation, and the bacteria in control tube for PASME exposed to drug-free medium, and then exposed to combination of 0.1 times MIC(C₁), 0.2 times MIC(C₂), or 0.3 times MIC(C₃) after centrifu...