Serum-mediated release of leukotoxin from the cell surface of the periodontal pathogen Actinobacillus actinomycetemcomitans

The leukotoxin of the periodontopathogen Actinobacillus actinomycetemcomitans is an important virulence factor that lyses human neutrophils and monocytes and thus, it may enable the bacterium to evade the local host defense. The toxin also induces degranulation of neutrophils and cytokine release in monocytes. To trigger these biological activities, leukotoxin has to be released from the bacterium and diffuse into the periodontal tissues. To date, the conditions found to cause toxin release have been artificial and have included high ion concentration and alkaline conditions. To study the release of the toxin under conditions mimicking the natural environment of the periodontium the ability of human serum to enable leukotoxin release from the bacterial surface was examined. Suspensions of leukotoxic A. actinomycetemcomitans strains were incubated with various concentrations of human serum or serum albumin. The suspensions were centrifuged and the leukotoxin in the supernatants or the cell pellets was detected by gel electrophoresis and immunoblotting. Serum was found to cause the rapid release of leukotoxin from the bacteria in a concentration-dependent manner. Pure albumin exhibited a similar effect. The leukotoxin released was active against human neutrophils. Only a minor proportion of it was associated with membranous vesicles produced by the bacteria. The results indicate that serum, a fluid closely related to the exudate in inflamed periodontal pockets, releases leukotoxin from the cell surface of A. actinomycetemcomitans . The process may enable the diffusion of the toxin from the bacterial biofilm into the surrounding tissues, where it can exert its biological effect.