Infection of feeder lambs with Mycoplasma ovipneumoniae was associated with outbreaks of respiratory disease. The disease, a "coughing syndrome" characterized by paroxysmal cough, mucoid nasal discharge, intolerance to exercise, and occasional development of rectal prolapses in some animals, has not been reported previously. Mycoplasma ovipneumoniae was isolated from 70% of 105 lambs examined and persistent isolation of this mycoplasma from diseased lambs was one of the evidences of its involvement in this coughing syndrome. The gross pathology associated with this mycoplasma was focal consolidation and was primarily observed in the cranial lobes of infected lungs. However, when a mixture of M. ovipneumoniae and a Gram-negative hemolytic coccobacillus isolated from a lamb lung were inoculated the lesions included severe consolidation and fibrinous adhesions. The histological lesions observed in lungs infected with M. ovipneumoniae included peribronchiolar and perivascular lymphocytic cuffing, lymphoid hyperplasia, interstitial thickening, and patchy loss of ciliated epithelium. The major infiltrating cells were lymphocytes, plasma cells, and macrophages. The histological lesions in lambs with the coughing syndrome were classical and diagnostic for M. ovipneumoniae infection in lambs. The indirect immunohistochemical staining of formalin-fixed lung tissues verified 84% of M. ovipneumoniae infected lungs. This technique proved to be a sensitive and specific method for in situ detection of M. ovipneumoniae antigens;Antigens of M. ovipneumoniae (heat killed antigen, sodium dodecyl sulfate treated antigen, and M. ovipneumoniae membrane fragments) demonstrated limited nonspecific mitogenic capability to transform ovine lymphocytes to blast cells. This may be correlated with the development of a cell mediated immune response in infected lungs as observed in the form of classical histological lesions;A challenge experiment in which a group of mycoplasma-free lambs were infected with an isolate of M. ovipneumoniae produced mild clinical disease. However, gross and histological lesions observed in the lungs of these lambs correlated with the pathological changes present in the natural disease;Restriction endonuclease DNA analysis indicated extensive heterogeneity among isolates of M. ovipneumoniae. Eight of 30 isolates possessed homologous patterns indicative of a single strain cluster. However, major to minor differences were present among the remaining isolates. Analysis of these isolates indicated 62 percent to 92 percent homology in band patterns when compared to the 8 isolates of one strain cluster. Hybridization of restriction DNA digests with a radiolabeled rRNA probe detected multiple bands of different molecular weights. Based on the homology of lower molecular bands 3 strain clusters of M. ovipneumoniae isolates were identified.