Invasion by Porphyromonas gingivalis has been proposed as a possible mechanism of pathogenesis in periodontal and cardiovascular diseases. Porphyromonas gingivalis have direct access to the systemic circulation and endothelium in periodontitis patients by transient bacteremia. Periodontitis can be described as one of the predominant polymicrobial infections of humans. In the present study, P. gingivalis strains were tested for their ability to invade a human gingival epithelial cell line (Ca9-22) and human aortic endothelial cells in coinfection with Fusobacterium nucleatum using antibiotic protection assays. Coinfection with F. nucleatum resulted in 2-20-fold increase in the invasion of host cells by P. gingivalis strains. The invasive abilities of P. gingivalis strains were significantly greater when incubated with a F. nucleatum clinical isolate (which possesses strong biofilm-forming ability), than when incubated with a F. nucleatum-type strain. In inhibition assays with metabolic inhibitors, a difference in inhibition profiles was observed between mono- and polymicrobial infections. Collectively, our results suggest that F. nucleatum facilitates invasion of host cells by P. gingivalis. Investigations of polymicrobial infection of host cells should improve our understanding of the role of P. gingivalis in periodontal infection and proatherogenic mechanisms.