Abstract
Effect of tobramycin on bacteriostatic and bactericidal activities of β-lactam antibiotics was studied by agar plate method. Two sets of agar plate series containing graded concentrations of tobramycin and a β-lactam antibiotic, in combination and separately, were prepared. Plates were inoculated with 103-104 (order of magnitude) CFUs/inoculum of Escherichia coli, Klebsiella sp. and Pseudomonas aeruginosa. One set of the plates was incubated at 35°C for 20 hours, and the minimal concentrations A and B of either tobramycin and the β-lactam drug yielding no bacterial growth on the plates were designated as minimal inhibitory concentrations (MICs).
The concentrations A' and B' of the two drugs in combination producing the most effective inhibition were determined. The sum of A'/A and B'/B (fractional inhibitory concentrations (FICs)) was defined as FIC index. After reading these bacteriostatic data, the surface of the plates was treated with β-lactamase spray to inactivate the β-lactam antibiotic, and the plates were further incubated at 35°C overnight. The minimal concentration C of the β-lactam drug which produced a 99.9% kill, designated as minimal lethal concentration (MLC), was determined. The concentrations C' and D' of either the β-lactam drug and tobramycin used in combination which produced the best result in terms of a 99.9% kill activity were also determined. The sum of D'/MIC of tobramycin (designated as “fractional lethal concentration” (“FLC”)) and C'/C of the β-lactam drug (FLC) was defined as FLC index.
Another set of the plates was incubated at 35°C for 3 hours. The surface was then treated with β-lactamase spray, and these plates were further incubated at 35°C overnight. The minimal concentration E of the β-lactam drug which produced a 99.9% kill (designated as 3h-MLC) and the concentrations E' and F' of either the β-lactam drug and tobramycin producing the best result in terms of a 99.9% kill activity were determined. The sum of F'/MIC of tobramycin (designated as “3h-FLC”) and E'/E of the β-lactam drug (3h-FLC) was defined as 3h-FLC index.
If the FIC index, FLC index and/or 3h-FLC index was <0.8, ≥0.8-<1.2 or ≥1.2, the combined effect was interpreted as synergistic, additive or antagonistic, respectively. When the combined effect of tobramycin and cephalothin was assessed at bacteriostatic (MIC) level, synergy was detected for 20 of 26 strains of E. coli, 15 of 27 strains of Kiebsiella sp. If assessed at bactericidal (MLC) level, much more strains showed synergistic effect. Most of the remainder of these strains yielded additive effect. Almost all these strains showed synergy if assessed at bactericidal level with 3 hours exposure to the β-lactam antibiotic (3h-MLC level). The combined effect of tobramycin and carbenicillin on 36 strains of P. aeruginosa showed synergism for most of these strains, both at bacteriostatic and bactericidal levels. Antagonism also occurred among some of these test strains.
