Clinical trials in surgery suggest that some failures of antibiotic prophylaxis are related to the in vivo degradation of β-lactams by Staphylococcus aureus β-lactamase. To explore this issue further, isogeneic isolates of S. aureus differing only in whether they contained the structural gene for type A staphylococcal β-lactamase were constructed and compared for their ability to establish an abscess in a guinea pig model. With ampicillin prophylaxis, the ID50 was 870 cfu for the β-lactamasenegative isolate VK7114 and 240 cfu for the β-lactamase-producing isolate VK7115 (P < .001). Similarly, the ID50 was greater for the β-lactamase-negative isolate when cefazolin prophylaxis was administered (599 vs. 128 cfu, VK7114 and VK7115; P < .001). In the setting of prophylaxis with β-lactamase-susceptible antibiotics, β-lactamase contributes to the pathogenesis of S. aureus wound infections.