Investigation of β-lactamases in clinical isolates of Staphylococcus aureus for further explanation of borderline methicillin resistance

Background: Borderline methicillin resistance in Staphylococcus aureus is due to β-lactamase overproduction and/or specific methicillinases. Methods: β-Lactamase activity in culture supernatants and in cytoplasmic membrane fractions was estimated by bioassay and by SDS-PAGE combined with nitrocefin assay. Results: During the investigation of borderline methicillin-resistant Staphylococcus aureus (BORSA) strains VU94 and 822 two β-lactamases were detected in the membranes, with molecular weights of 13 and 30 kDa. The latter could be found in the culture supernatants, too. In the presence of globomycin, this enzyme disappeared from the membrane, and the oxacillin-hydrolyzing activity of the membrane decreased to the level of susceptible strains. Both β-lactamases were detected in the methicillin-resistant Staphylococcus aureus strain studied, but the susceptible strains possessed only the first enzyme. Conclusions: The 30-kDa β-lactamase proved to be a methicillinase, and it can be one of the main causes of the borderline phenotype of BORSA strains. The other enzyme is one of the smallest β-lactamases published to date.