Platelet vegetations or thrombi are common findings in subacute bacterial endocarditis. We investigated the hypothesis that human platelets selectively bind or adhere strains of Streptococcus sanguis and Streptococcus mutans and aggregate, as a result, into an in vitro thrombus. Earlier ultrastructural studies suggested that aggregation of platelets over time by Staphylococcus aureus was preceded in order by adhesion and platelet activation. We uncoupled the adhesion step from activation and aggregation in our studies by incubating streptococci with platelet ghosts in a simple, quantitative assay. Adhesion was shown to be mediated by protease-sensitive components on the streptococci and platelet ghosts rather than cell surface carbohydrates or dextrans, plasma components, or divalent cations. The same streptococci were also studied by standard aggregometry techniques. Platelet-rich plasma was activated and aggregated by certain isolates of S. sanguis. Platelet ghosts bound the same strains selectively under Ca2+- and plasma-depleted conditions. Fresh platelets could activate after washing, but Ca2+ had to be restored. Aggregation required fresh platelets in Ca2+-restored plasma and was inducible by washed streptococcal cell walls. These reactions in the binding and aggregometry assays were confirmed by transmission electron microscopy. Surface microfibrils on intact S. sanguis were identified. These appendages appeared to bind S. sanguis to platelets. The selectivity of adhesion of the various S. sanguis strains to platelet ghosts or Ca2+- and plasma-depleted fresh washed platelets was similar for all donors. Thus, the platelet binding site was expressed widely in the population and was unlikely to be an artifact of membrane aging or preparation. Since selective adhesion of S. sanguis to platelets was apparently required for aggregation, it is suggested that functionally defined receptors for ligands on certain strains of S. sanguis may be present on human platelets. Some differences in the selectivity and rate of the aggregation response were noted among platelet donors, although the meaning of the variability requires further study. Nonetheless, these interactions may contribute to platelet accretion in the initiation and development of vegetative lesions in the subacute bacterial endocarditis.