Regeneration of Virus Myxomatosum (Sanarelli) in the Presence of Cells of Exudates Surviving in Vitro.

Abstract

For a number of years the relation of viruses to cells has been under investigation in our laboratory. In pursuance of this study, attempts to cultivate the virus of infectious myxomatosis of rabbits in different kinds of tissue cultures were made. At the beginning of the work, bits of rabbit testicle suspended in a mixture of rabbit serum (1 part) and Tyrode's solution (3 parts) were used as a medium. Such preparations, however, proved unsuitable for the in vitro cultivation of the virus. Previous observations indicated that large mononuclear wandering cells are involved in this disease. Therefore, it seemed not unlikely that the inadequacy of the above methods of cultivation was referable to the failure of mobilization of these cells in excised tissues. To test this hypothesis, attempts to cultivate the virus in the presence of mononuclear cells obtained according to the method of Gay and Clark 1 were made.

A sterile mixture (6 cc.) of beef extract and gum acacia was injected into the right pleural cavity of a normal rabbit. Seventy-two hours later the cavity was opened aseptically and the fluid contents were aspirated. Heparin (1 cc. of a 1:1000 solution for each 10 cc. of pleural fluid) was added to the exudate to prevent clotting and the mixture was centrifuged for 5 minutes at low speed. The sediment was resuspended in Tyrode's solution and again centrifuged. Finally, the cells from 5 to 10 cc. of exudate were suspended in 2 cc. amounts of a mixture of rabbit serum (1 part) and Tyrode's solution (3 parts) and placed in 3 cm. Carrel flasks. To each flask with its 2 cc. of medium, 0.2 cc. of sterile tissue juice containing myxoma virus was added.