The Development of the Malaria Parasite Plasmodium lophurae in Red Blood Cell Suspensions In vitro

Abstract

In earlier studies (Trager 1941, 1943) on conditions affecting the survival and development in vitro of Plasmodium lophurae, an avian malaria parasite, it was found that red cell extract, suitable concentrations of glucose, glutathione, calcium pantothenate and serum or plasma, a balanced salt solution having a high potassium content, an optimal density of parasites per cubic millimeter, gentle agitation of the suspension, aeration (but not a high oxygen tension), renewal of the medium, and provision of fresh red blood cells, all favored survival. In the best preparations, small increases in the parasite number occurred during the first few days of incubation (at about 400 C) and living infective parasites were demonstrated after 16 days in vitro. In all these experiments the parasite density ranged from 25,000 to 100,000/cu mm and the concentration of red cells from about 150,000 to 350,000 per cu mm. The total number of red cells was kept low in order to facilitate the observation of the effects of changes in the medium, and indeed effects were observed which probably could not have been noted in the presence of large numbers of normal red cells. More recently it has been found by others (Ball et al, 1945; Anfinsen et al, 1946; Geiman et al, 1946; McKee et al, 1946) that Plasmodium knowlesi, a monkey malaria parasite, will multiply in suspensions of monkey red blood cells maintained in vitro. The medium is a balanced salt solution resembling in composition the inorganic constituents of monkey plasma and containing vitamins, purines, pyrimidines and amino acids. Most of the experiments consisted of observations of the effect of various conditions on the extent of multiplication of the parasites during the first 24-hour period after removal from the monkey, but in one series the parasites were kept growing in vitro for 6 days. Good multiplication even during the first 24-hour period was obtained only in the presence of adequate concentrations of glucose, plasma or serum, and p-aminobenzoic acid. The vessels with the parasitized blood suspensions were rocked (a condition which favored survival of P. lophurae) and in addition a stream of 5% CO2-95% air was passed through them. In these experiments a low parasite density (16,000-25,000 per cu mm) was again found favorable. However, much larger amounts of normal red blood cells were used (1 million or more per cu mm) than in the earlier work with P. lophurae. In vitro studies with P. lophurae have now been carried out using combinations of the methods of Trager and of Anfinsen, Ball, Geiman, McKee, and Ormsbee.