The purpose of this study was to determine whether lysostaphin would enhance its anti-staphylococcal efficacy in combination with lysozyme. Minimum inhibitory concentrations (MICs) of lysostaphin and lysozyme were separately determined for 41 strains belonging to 10 different species of human staphylococci. Lysozyme was virtually inactive and showed MICs of 15 mg/ml. On the contrary, all strains were susceptible to lysostaphin and showed MICs ranging from 2.5 to 60 micrograms/ml for the different Staphylococcus species. When the MIC of lysostaphin was determined in media containing submultiples of the MIC of lysozyme, the values obtained were much lower. The reduction of the lysostaphin MIC ranged from 16- to 200-fold in the different species tested. In Staphylococcus aureus, in particular, the combination of lysostaphin with 1.5 mg of lysozyme per ml reduced the MIC of lysostaphin by 25-fold. The activities of two combinations of the two enzymes were evaluated: one combination was expected to be active on S. aureus only, and the other combination was expected to inhibit all Staphylococcus strains. The first combination (0.5 micrograms of lysostaphin plus 0.5 mg of lysozyme per ml) was inhibitory to all of the 84 S. aureus strains tested, whereas 137 of 151 strains of other Staphylococcus species were unaffected. On the contrary, all of the 235 Staphylococcus strains tested were inhibited by the second combination (4 micrograms of lysostaphin plus 5 mg of lysozyme per ml). The possible mechanisms of lysostaphin potentiation by lysozyme are considered, and the potential use of a lysostaphin-lysozyme combination for topical therapy of staphylococcal infections resistant to other antibiotics is discussed.