Mastitis was reproduced experimentally by inoculating the lactating mammary glands of 12 ewes with Mycoplasma californicum. Large numbers of mycoplasmas were recovered from the milk. The mastitis had a lobular distribution and was characterized in the early stages by degeneration of the secretory epithelium and intense leucocytic infiltration of alveolar and ductal lumina. Later there was widespread alveolar involution, and lymphohistiocytic infiltration and fibrosis of the interstitium. A monoclonal antibody-based immunohistochemical technique detected M. californicum antigen in alveolar and ductal lumina but not in epithelium or interstitial tissue. This report is the first description of the pathological features of M. californicum mastitis in ewes.