Staphylococcal pyoderma is rarely contagious between dogs and humans, or humans and dogs. This study investigated the hypothesis that there are species differences in adherence of Staphylococcus intermedius (the most common isolate from dogs) and Staphylococcus aureus (the most common isolate from humans) to canine and human corneocytes. Sheets of corneocytes were collected from the ventral abdomen of 10 dogs and the medial forearm of 10 humans (all healthy and without any history or physical signs of skin disease) using double-sided tape. Staphylococcus intermedius from a case of canine bacterial pyoderma and a human strain of S. aureus were prepared in phosphate buffered saline (PBS) and applied in duplicate, respectively, to the canine and human corneocyte-covered tapes using PBS as negative control. After incubation, rinsing, and staining with crystal violet, quantification of the adherent bacteria was carried out blindly by computerized image analysis. Staphylococcus intermedius was found to adhere significantly more to canine corneocytes than S. aureus (P = 0.0006), whereas S. aureus showed greater adherence to human corneocytes than S. intermedius (P < 0.0001). In addition, the pattern of adherence differed between the two organisms, with S. intermedius adhering to the entire surface and S. aureus adhering mainly to the periphery of both canine and human corneocytes. Preference for adherence to these two hosts may explain, in part, why S. intermedius and S. aureus are uncommonly isolated from human and canine skin infections, respectively.