Colonization of the cardiovascular endothelium by viridans group streptococci can result in infective endocarditis and possibly atherosclerosis; however, the mechanisms of pathogenesis are poorly understood. We investigated the ability of selected oral streptococci to infect monolayers of human umbilical vein endothelial cells (HUVEC) in 50% human plasma and to produce cytotoxicity. Planktonic Streptococcus gordonii CH1 killed HUVEC over a 5-h period by peroxidogenesis (alpha-hemolysin) and by acidogenesis but not by production of protein exotoxins. HUVEC were protected fully by addition of supplemental buffers and bovine liver catalase to the culture medium. Streptococci were also found to invade HUVEC by an endocytic mechanism that was dependent on polymerization of actin microfilaments and on a functional cytoskeleton, as indicated by inhibition with cytochalasin D and nocodazole. Electron microscopy revealed streptococci attached to HUVEC surfaces via numerous fibrillar structures and bacteria in membrane-encased cytoplasmic vacuoles. Following invasion by S. gordonii CH1, HUVEC monolayers showed 63% cell lysis over 4 h, releasing 64% of the total intracellular bacteria into the culture medium; however, the bacteria did not multiply during this time. The ability to invade HUVEC was exhibited by selected strains of S. gordonii, S. sanguis, S. mutans, S. mitis, and S. oralis but only weakly by S. salivarius. Comparison of isogenic pairs of S. gordonii revealed a requirement for several surface proteins for maximum host cell invasion: glucosyltransferase, the sialic acid-binding protein Hsa, and the hydrophobicity/coaggregation proteins CshA and CshB. Deletion of genes for the antigen I/II adhesins, SspA and SspB, did not affect invasion. We hypothesize that peroxidogenesis and invasion of the cardiovascular endothelium by viridans group streptococci are integral events in the pathogenesis of infective endocarditis and atherosclerosis.Viridans group streptococci comprise a large proportion of the commensal bacteria that colonize oral surfaces (20,24,25). These bacteria frequently enter the bloodstream following trauma to oral tissues (12,17,41,58) and can then adhere to surfaces of abnormal or previously damaged heart valves (15,21,29,47) or become implanted in arterial atherosclerotic plaques (11). Streptococci growing on heart valve surfaces (causing infective endocarditis) become encased in a matrix of fibrin and platelets, which form macroscopic verrucous lesions and can lead to valve perforation, abnormalities in cardiac conduction, valve ring abscesses, pericarditis, aneurysm of the sinus of Valsalva, and release of peripheral emboli (21, 56). Viridans group streptococci are the most common cause of native valve endocarditis in humans, accounting for 45 to 80% of cases (5, 55). A variety of virulence factors have been implicated in the initial colonization of bacteria to cardiac valve surfaces (1, 32, 49, 50, 54), but those responsible for the ultimate destruction of underlying tissues are not well unders...