The role of Staphylococcus aureus collagen binding in the development of experimental endocarditis was studied. Two isogenic strains of S. aureus, 1 carrying an insertional inactivation of the gene encoding collagen-binding protein, were compared in a rat model of catheter-induced infective endocarditis (IE). Separate groups of rats with traumatized aortic valves were intravenously challenged with 1 of the strains. In rats sacrificed 24 h after inoculation, the collagen-binding strain significantly outnumbered the mutant strain (P < .001); however, 1 h after challenge, there was no difference in numbers of the 2 strains. The results were substantiated, using a 1:1 mixture of the parent strain and the mutant as an inoculate. Our findings suggest that collagen binding of S. aureus is important in the sustenance of experimental IE and plays a limited role during the initial attachment of the microorganism to traumatized aortic valves.