BACKGROUND: Many Gram-positive pathogens aggregate and activate platelets in vitro and this has been proposed to contribute to virulence. Platelets can also form complexes with neutrophils but little is however known about platelet and platelet-neutrophil responses in bacterial infection. METHODOLOGY/PRINCIPAL FINDINGS: We added isolates of Gram-positive bacteria from 38 patients with a bacteremic infection to blood drawn from the same patient. Aggregometry and flow cytometry were used to assess platelet aggregation and to quantify activation of platelets, neutrophils, and platelet-neutrophils complexes (PNCs) induced by the bacteria. Fifteen healthy persons served as controls. Most isolates of Staphylococcus aureus, beta hemolytic streptococci, and Enterococcus faecalis induced aggregation of platelets from their respective hosts, whereas pneumococci failed to do so. S. aureus isolates induced platelet aggregation more rapidly in patients than in controls, whereas platelet activation by S. aureus was lower in patients than in controls. PNCs were more abundant in baseline samples from patients than in healthy controls and most bacterial isolates induced additional PNC formation and neutrophil activation. CONCLUSION/SIGNIFICANCE: We have demonstrated for the first time that bacteria isolated from patients with Gram-positive bacteremia can induce platelet activation and aggregation, PNC formation, and neutrophil activation in the same infected host. This underlines the significance of these interactions during infection, which could be a target for future therapies in sepsis.