Anchoring of Staphylococcus aureus surface protein to the cell wall is catalyzed by sortase, a transpeptidase. The contribution of staphylococcal surface proteins to establishment of infection was examined using a murine septic arthritis model. Intravenous inoculation of mice with the sortase-deficient mutant S. aureus strain SMK3 did not result in weight loss or severe septic arthritis, in contrast to the parent strain, S. aureus Newman. Direct inoculation of the sortase mutant into joint cavities also failed to cause severe synovitis or erosive arthritis. Furthermore, intravenous inoculation with staphylococci resulted in the rapid clearing of the sortase mutant from the bloodstream. This phenomenon demonstrates the involvement of host neutrophils; when these cells were depleted, sortase mutant staphylococci caused severe systemic infection, although not septic arthritis. These results suggest that sortase mutant staphylococci are significantly less virulent than the parent strain, Newman: the sortase mutant has decreased ability to reach target organs and, once there, to induce an inflammatory response.