988 AMPICILLIN (A)/NAFCILLIN (N) SYNERGY AGAINST AMPICILLIN-RESISTANT HAEMOPHILUS INFLUENZAE TYPE B (Hib^R)

Abstract

Three infants with Hib^R bacteremic infections had good clinical responses to A and N therapy. This observation prompted in vitro evaluation of this combination against Hib^R clinical isolates. Minimal inhibitory concentrations (MICs) in μg/ml of A and N alone and in combination were determined by a microdilution method for 5 Hib^R isolates; A: 8-32, N: 8-16, A/N: 0.5-1/1-2 at a 10⁴ colony forming unit/ml inoculum. Results indicate synergy since the MIC of each drug alone is at least 4 times greater than the MIC of the drug in combination. The proposed mechanism of A/N synergy is N inhibition of β-lactamase which protects A from hydrolysis allowing it to exert its antibacterial effect. A spectrophotometric assay of β-lactamase activity was used to study this proposal. The supernatant of sonified Hib^R cell preparations was the β-lactamase source; A served as substrate; and N, methicillin(M) and oxacillin(O) were studied as potential inhibitors. The hydrolysis of A in the absence and presence of the inhibitors obeyed Michaelis-Menten kinetics. N, M, and 0 acted as purely competitive inhibitors of A hydrolysis with N being the most effective and 0 the least as evidenced by K_I values of 7.10 × 10⁻², 3.16 × 10⁻¹, and 3.35 μM respectively. These findings provide an explanation for the observed in vitro A/N synergy which in vivo may have been responsible for favorable outcomes in 3 infants with Hib^R bacteremic infections treated with A and N.

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