Leukotoxic activity in Actinobacillus(Haemophilus) actinomycetemcomitans isolated from patients with rapidly progressive periodontitis (RP), gingivitis (G), and juvenile periodontitis (JP), and several oral bacteria, was determined by observation of morphological changes in polymorphonuclear leukocytes (PMNs). Many A. actinomycetemcomitans isolates yielded both rough-surfaced and umbonate-shaped colonies (A-type), and smooth-surfaced and convex-shaped colonies (B-type), when stock cultures were streaked on agar medium. Both types of cells were identical in terms of Gram stain, cell morphology, sugar fermentation profile, nitrate reduction and cellular fatty acid composition. Sonic extracts were prepared from 32 A. actinomycetemcomitans strains isolated from patients and from 3 American Type Culture Collection (ATCC) strains. Sonic extracts from 8 isolates and 2 ATCC strains induced sphering of PMNs during a 45-50min period of incubation at 37C. Extracts from the other oral bacteria had no effects on PMN morphology. The sphered PMNs were found by their fluorochromatic-negative reactions to be damaged cells. The leukotoxic substance was heat-sensitive (56C, 30min), trypsin-sensitive and did not induce sphering of PMNs at 4C. There was no clear correlation between colony type and leukotoxicity. Among 8 leukotoxic strains, 5 were isolates from an RP patient.