The efficacy of antimicrobial agents applied topically in the oropharynx and trachea with and without intravenous antibiotics in preventing bacterial pneumonias during prolonged (7 to 10 days) mechanical ventilation was studied in 35 baboons, 30 of which had acute lung injury induced by either oleic acid or hyperoxia. In 12 animals receiving no antibiotics, only topical application of polymyxin B (PB), or only intravenous penicillin and gentamicin (IV PCN/GM), moderate or severe pneumonia was found in 81% of lobes examined at necropsy; no lobes were sterile. Pneumonias were polymicrobial in the absence of antibiotics, due to PCN-sensitive organisms in the topical PB group, and due to gram-negative bacilli in the IV PCN/GM group. Combinations of topical PB or GM or both plus IV PCN were highly efficacious in preventing pneumonia in 23 animals as only 15% of the lobes contained moderate to severe pneumonia and 52% of lobes were sterile. In these groups, histologically evident pneumonias were associated with low concentrations of bacteria in lung tissue, principally gram-negative bacilli resistant to the topical agent being used. Resistance to PB appeared to be solely due to selection of intrinsically resistant species, whereas resistance to GM may have developed through additional mechanisms as well. Although this approach to pneumonia prevention is clearly efficacious in this animal model, clinical studies are needed to define the frequency and significance of microbial resistance in human subjects.