Evidence for a true post-β-lactamase-inhibitor effect of clavulanic acid against Klebsiella pneumoniae and Haemophilus influenzae

The purpose of this study was to investigate and characterize in vitro the post-β-lactamase inhibitor effect (PLIE) of clavulanic acid against two β-lactamase-producing species of bacteria. The PLIE was investigated against one strain of Klebsiella pneumoniae and one strain of Haemophilus influenzae. A stationary-phase inoculum of about 107 colony-forming units per mL of each bacterium was pre-exposed for 2 h to clavulanic acid, either alone or in combination with amoxicillin at various concentrations. After pre-exposure, the dilution required to remove the β-lactamase inhibitor was 1:100 or 1:1000 according to the bacterial species and their susceptibilities to clavulanic acid. Bacteria were counted hourly after drug removal, on solid agar medium. Control cultures exposed to amoxicillin alone after dilution, showed a delay in growth, which may be inherent to the time required to synthesize sufficient β-lactamase after the dilution steps. Control experiments clearly distinguished the post-antibiotic effect and the growth delay from the PLIE. The PLIE could be one of several factors explaining why β-lactam/β-lactamase inhibitor combinations remain effective throughout the dosing interval, even if a few hours after in vivo administration, serum concentrations of β-lactamase inhibitor fall below levels that are active in vitro.