Nonphagocytic clearance of Staphylococcus aureus from murine lungs

Several investigations of host and bacterial factors critical to staphylococcal clearance from lungs suggest that alveolar macrophages may not provide the principal defense against inhaled staphylococci. We evaluated possible contributions of extracellular bactericidal activities in lungs with a standard aerosol challenge model by using methods which allowed recovery of macrophages for in vitro bactericidal assays and recovery of intrapulmonary staphylococci for clearance studies. Macrophages recovered by a gentle lavage technique immediately after aerosol exposure contained 6.0 +/- 2.3 colony-forming units of viable staphylococci per 100 glass-adherent macrophages. These intracellular staphylococci were killed in vitro with a half-life of 10.8 +/- 2.1 h, which is identical to our results with a completely in vitro system for ingestion and killing. However, 99.4 +/- 0.2% and 94.9 +/- 1.5% of the viable cocci recovered in bronchoalveolar lavage at 0.5 and 6.0 h after aerosol exposure were sensitive to lysostaphin, a rapidly bactericidal enzyme with no demonstrable activity against intracellular organisms and therefore, presumably extracellular. Photomicrographs from lavage pellets obtained 0.5, 1.5, 3.0, and 5.5 h after aerosol exposure confirmed the presence of numerous extracellular cocci. These extracellular cocci were eliminated at the same rate as whole lung cocci (half-life = 3.07 and 3.14 h, respectively) and at a much faster rate than intracellular cocci. In summary, we found large numbers of extracellular staphylococci in bronchoalveolar spaces during the first 6 h after aerosol exposure that are inactivated at the same rate as the whole lung bacterial population. Since only a small number of staphylococci are ingested by macrophages and intracellular bactericidal activity appears too slow to explain intrapulmonary killing, we conclude that an as yet unidentified extracellular killing process contributed to staphylococcal clearance.