Characterization of the T-cell subsets in intestinal lesions in sheep with paratuberculosis may provide important information on understanding the pathogenesis of this disease. To determine the phenotype and distribution of lymphocytes in the normal sheep intestinal mucosa and in Mycobacterium paratuberculosis infected sheep, immunohistochemistry was performed on 12 normal and 12 naturally infected, clinically diseased sheep. Immunoperoxidase staining was carried out on frozen sections of ileum using monoclonal antibodies against ovine CD4, CD8, gamma T-cell receptor (TCR). In both sample groups, cells appeared to be non-randomly distributed throughout the lamina propria. Higher densities of lymphocytes were present in villus than in crypt areas. CD8+ cells were located principally around the epithelial basement membrane, whereas CD4+ cells were localized towards the central villus area of the lamina propria. Very few CD4+ cells were present within the epithelium. Lymphocytes bearing the gamma T-cell receptor were more widely distributed, both in epithelial and lamina propria compartments. The mean relative percentages of total positively staining cells by monoclonal antibody were CD4+ 37.1/37.9, CD8+ 52.0/46.3, gamma TCR 11.0/15.8 percent in normal and diseased ileum respectively. This indicates a significant relative increase in the percentage of gamma TCR bearing cells in paratuberculosis infection (P less than 0.05).