The isolation of cytomegalovirus from peripheral blood.

D URING A STUDY designed to detect the presence of transforming agents in the blood of patients with leukemia, a cytomegalovirus was isolated from one patient. The essence of the series of experiments was to co-cultivate leukemic cells and human embryonic fibroblasts and to maintain these latter for a long period of time to observe any evidence of transformation. In none of these experiments, except the one to be considered here, is there any evidence so far to suggest the presence of virus; nor is there any evidence of transformation. The patient was a boy, aged 8 years, in whom acute lymphatic leukemia was first diagnosed in May 1965. By the time the first blood sample was taken on June 1, 1966, the child had been treated with steroids, 6-mercaptopurine, methotrexate, cyclophosphamide, and x-irradiation. Vincristine was used between the first and second blood samples. The child died on July 21, 1966, one day after the second blood sample was taken. The family history is of some interest: The mother had an x-ray pelvimetry during the pregnancy and the child was delivered by Caesarian section. An older brother is alive and well, but a younger sister has a neuroblastoma. Blood was taken into heparin, spun, and the buffy coat removed; since the white count was low, the buffy coat also included some red cells. This suspension of red and white cells was then washed three times is tissue culture medium ( medium F 101, 80 per cent; agamma bovine serum, 10 per cent; and tryptose phosphate broth, 10 per cent ) . A suspension of low-passage human embryo lung fibroblasts was prepared and half of the suspension mixed with the washed blood cells. Four dishes (with approx. 10 fibroblasts) were then plated out from the mixed suspension and four from the unmixed suspension. The medium was changed the following day. No cvtopathic effect was noted in the treated cultures, but they did remain inactive for a few days. Controls and treated cultures were routinely medium changed and subcultured. Four weeks later small foci of abnormal cells were noted in the cultures treated with leukemic cells, and with time these became more extensive. The control plates were consistently negative in all observations. Attempts to transmit the effect using filtered supernatants were unsuccessful. However, transmission was demonstrated by x-irradiating affected cells with