Fusobacterium nucleatum Transports Noninvasive Streptococcus cristatus into Human Epithelial Cells

Analysis of human buccal epithelial cells frequently reveals an intracellular polymicrobial consortium of bacteria. Although several oral bacteria have been demonstrated to invade cultured epithelial cells, several others appear unable to internalize. We hypothesized that normally noninvasive bacteria may gain entry into epithelial cells via adhesion to invasive bacteria. Fusobacterium nucleatum is capable of binding to and invading oral epithelial cells. By contrast, Streptococcus cristatus binds weakly to host cells and is not internalized. F. nucleatum and S. cristatus coaggregate strongly via an arginine-sensitive interaction. Coincubation of KB or TERT-2 epithelial cells with equal numbers of F. nucleatum and S. cristatus bacteria led to significantly increased numbers of adherent and internalized streptococci. F. nucleatum also promoted invasion of KB cells by other oral streptococci and Actinomyces naeslundii. Dissection of fusobacterial or streptococcal adhesive interactions by using sugars, amino acids, or antibodies demonstrated that this phenomenon is due to direct attachment of S. cristatus to adherent and invading F. nucleatum. Inhibition of F. nucleatum host cell attachment and invasion with galactose, or fusobacterial-streptococcal coaggregation by the arginine homologue L-canavanine, abrogated the increased S. cristatus adhesion to, and invasion of, host cells. In addition, polyclonal antibodies to F. nucleatum, which inhibited fusobacterial attachment to both KB cells and S. cristatus, significantly decreased invasion by both species. Similar decreases were obtained when epithelial cells were pretreated with cytochalasin D, staurosporine, or cycloheximide. These studies indicate that F. nucleatum may facilitate the colonization of epithelial cells by bacteria unable to adhere or invade directly.