Actinobacillus actinomycetemcomitans may utilize either actin-dependent oractin-independent mechanisms of invasion

Actinobacillus actinomycetemcomitans is an important pathogen implicated in juvenile and adult periodontal diseases. An important virulence factor of A. actinomycetemcomitans is the ability to invade human oral epithelial cel ls. A clinical isolate, A. actinomycetemcomitans SUNY 465, has previously b een shown to enter epithelial cells by an actin-dependent mechanism. The in ternalized bacteria are surrounded by an actin halo upon entry. These data are consistent with the mode of entry associated with many enteric pathogen s. We tested the effects of cytochalasin D, an inhibitor of the actin micro filament network, on bacterial entry to determine whether this mode of entr y was common to other A. actinomycetemcomitans clinical isolates. Cytochala sin D was added prior to infection. A. actinomycetemcomitans SUNY 523 and A . actinomycetemcomitans 4065 exhibited enhanced ability to enter epithelial cells in the presence of cytochalasin D. Immunofluorescent labeling of bac teria and host cell actin confirmed that actin was not being mobilized by t he entry of A. actinomycetemcomitans SUNY 523. Inhibitors of receptor-media ted endocytosis inhibited invasion of A. actinomycetemcomitans SUNY 523 and A. actinomycetemcomitans 4065. Microtubule effecters did not inhibit invas ion of A. actinomycetemcomitans. A. actinomycetemcomitans SUNY 523, but not A. actinomycetemcomitans 4065, was deficient in exit from epithelial cells as determined by the absence of organisms in the assay medium. These data suggest that A. actinomycetemcomitans strains utilize at least two distinct mechanisms for entry into epithelial cells, and that A. actinomycetemcomit ans SUNY 523 may be defective in exit and cell-to-cell spread.