Malignant catarrhal fever is an important disease of cattle and certain wild ruminants. It occurs in sporadic and epizootic forms in Colorado cattle. Specimens from 15 cattle with naturally occurring malignant catarrhal fever and 5 cattle with induced disease were examined for cell-free and cell-associated viruses. Enteroviruses were isolated from leukocytes of 2 cattle with field cases of the disease. A herpesvirus with characteristics of the "Movar"-type, isolated from the spleen of another steer with a field case could be propagated optimally in bovine fetal spleen cells. A cell-associated virus, forming polykaryons, was isolated in adrenal and thyroid cells from 3 cattle with experimentally induced malignant catarrhal fever. It was cultured from leukocytes, ependymal tissue, spleen, lymph node, kidney and thyroid and adrenal glands of affected cattle and remained cell-associated in 48 subsequent passages. It was inactivated by freezing and thawing and by treatment with ultrasound; its polykaryon-forming activity was inhibited by 25 mug of 5 fluoro-2' deoxyuridine/ml. This viral agent replicated in bovine fetal adrenal, thyroid, and spleen cells, but not in primary kidney cells or in MDBK, HeLa, or L cells. A representative isolate was identified electronmicroscopically as an enveloped virus, 120 to 150 nm in diameter. Structural analysis indicated that it had properties of the bovine syncytial viruses. Attempts to induce MCF in cattle with 1 of the isolates failed. A parvovirus was isolated from the jejunal lymph node of calf 72-P-293, which had the experimentally induced disease and was also infected with the syncytial virus.