The microbicidal activities of freeze-thaw and high-salt extracts of human and bovine polymorphonuclear leukocyte (PMN) granules were tested against a smooth intermediate strain (45/0) and a rough strain (45/20) of Brucella abortus which differ in virulence and survival within PMNs. Freeze-thaw extracts of human PMN granules were more brucellacidal than high-salt extracts when supplemented with hydrogen peroxide (H2O2) and potassium iodide (KI), whereas the opposite was found with freeze-thaw and high-salt extracts of bovine PMN granules. There was no oxygen-independent killing of either the smooth or rough strain of B. abortus by amounts of granule extracts which caused 100% killing of a deep rough mutant (Re) of Salmonella typhimurium. The oxygen-dependent brucellacidal activity of granule extracts was dependent on concentrations of myeloperoxidase (MPO) units, H2O2, and KI. Maximal brucellacidal activity was observed at pH 5.5 to 6.0. The smooth strain, 45/0, was more resistant to oxygen-dependent killing by granule extracts than was the rough strain, 45/20. Granule extracts were more brucellacidal than purified MPO at equivalent levels of MPO enzyme units, suggesting that at least one other reaction enhances killing by the MPO-H2O2-I- system.