Enterococcus faecalis is responsible for a large variety of nosocomial infections. The intestinal barrier is thought to be one of the preferential portals of entry of enterococci, and the ability of E. faecalis to survive within peritoneal macrophages may contribute to spreading to distant sites. We examined the ability of a polysaccharide-expressing (biofilm-positive) E. faecalis strain and an isogenic biofilm-negative mutant to enter and survive within professional and nonprofessional phagocytes. Biofilm-positive bacteria survived longer in all cell systems than did biofilm-negative bacteria, through a process of receptor-mediated endocytosis that is dependent on functional reorganization of microtubules and polymerization of microfilament and on activation of protein kinases but not ATPases or protein phosphatases. We suggest that glycosaminoglycans--specifically heparin, heparan sulfate, and chondroitin sulfate A--are the host receptors for enterococci on professional and, possibly, nonprofessional phagocytes, allowing entry of enterococci into cell compartments where killing mechanisms are inhibited.