Controversy persists over the role that the capsular polysaccharide plays in the pathogenesis of Staphylococcus aureus infections. To address this issue, we compared the mouse virulence of S. aureus Reynolds and capsule-defective mutant strains cultivated under conditions of high or low capsule expression. Strain Reynolds cells cultivated on Columbia salt agar plates expressed approximately 100-fold more type 5 capsular polysaccharide than did cells cultivated in Columbia salt broth. The relative virulence of strain Reynolds and its capsule-defective mutants after growth on either solid or liquid medium was examined in mice challenged intraperitoneally or intravenously. The results indicated that agar-grown Reynolds cells were cleared from the bloodstream of mice less readily than broth-grown Reynolds cells. When the parental and mutant strains were cultivated on solid medium, strain Reynolds sustained a higher level of bacteremia than did the capsular mutants. We performed in vitro opsonophagocytic killing assays to determine whether staphylococcal virulence for mice correlated with resistance to phagocytosis. S. aureus Reynolds cultivated on solid medium was susceptible to phagocytic killing only in the presence of specific capsular antibodies and complement. Strain Reynolds grown in broth showed opsonic requirements for phagocytic killing that were similar to those of the capsular mutants (grown in broth or on agar); i.e., the bacteria were opsonized for phagocytosis by nonimmune serum with complement activity. These studies indicate that optimal expression of capsule enhances bacterial virulence in the mouse model of bacteremia, probably by rendering the organisms resistant to opsonophagocytic killing by leukocytes.