Wild deer have an important role in the epidemiology of bovine tuberculosis (bTB). The aims of this study were (1) to compare the pattern of lesions present in wild red (Cervus elaphus) and fallow (Dama dama) deer that were naturally infected with Mycobacterium bovis, and (2) to use this information to develop a sampling strategy for the isolation of M. bovis from the lymphoid tissues of the head of these animals. Culture of head lymphoid tissues demonstrated that 28 of 95 red deer and 22 of 100 fallow deer sampled were infected with M. bovis. Approximately 30% of each deer population had no gross lesions. Fallow deer were significantly more likely to have thoracic lesions than red deer. Lesions were observed in the retropharyngeal lymph nodes of 64% of the culture-positive red deer and 43% of the culture positive fallow deer. One third of the red deer, but none of the fallow deer, had well-encapsulated abscess lesions. There were no microscopical differences in the lesions in the lymph nodes of the red and fallow deer. Bacteriological culture from both the tonsil and retropharyngeal lymph nodes increased the rate of isolation of M. bovis by 22% over culture of the retropharyngeal lymph nodes alone in both species. These findings indicate that investigation of wild deer for bTB-compatible lesions should include examination of the medial retropharyngeal, left tracheobronchial, mediastinal, mesenteric and ileocaecal lymph nodes. Sampling for bacteriological culture from head lymphoid tissues should be from the tonsil and the medial retropharyngeal lymph nodes. These protocols may prove useful in bTB surveillance and control in regions where wild deer contribute to the circulation of M. bovis.