Within a 1-year period, six surgical-site infections (SSI) caused by Staphylococcus schleiferi were observed in the department of cardiac surgery of Ignatius Hospital, Breda, The Netherlands. Since outbreaks caused by this species of coagulase-negative staphylococci have not been described before, an extensive environmental survey and a case control study were performed in combination with molecular typing of the causative microorganism in order to identify potential sources of infection. Variability, as detected by four different genotyping methods (random amplification of polymorphic DNA [RAPD], conventional and PCR-mediated ribotyping, and pulsed-field gel electrophoresis [PFGE] of DNA macro restriction fragments), appeared to be limited both among the clinical isolates and among several control strains obtained from various unrelated sources. Among unrelated strains, RAPD and PCR-mediated ribotyping identified two types only, whereas seven different types were identified in a relatively concordant manner by conventional ribotyping and PFGE. The latter two procedures proved to be the most useful tools for tracking the epidemiology of S. schleiferi. Four of the outbreak-related strains were identical by both methods, and two isolates showed limited differences. In the search for a potential source of S. schleiferi infection, two slightly different PFGE types were encountered on several occasions in the nose of a single surgeon. These strains were, however, clearly different from the outbreak type. In contrast, S. schleiferi cultures remained negative for two persons identified on the basis of case control analysis. It was demonstrated that SSI caused by S. schleiferi had a clinical impact for patients comparable to that of a wound infection caused by Staphylococcus aureus. This report describes the first well-documented outbreak of S. schleiferi infection. A source of the outbreak was not detected.