Effect of erythrocyte membrane on extracellular development of the erythrocytic cycle of Plasmodium falciparum.

Abstract

We have earlier reported conditions that support the axenic development in vitro of a complete asexual erythrocytic cycle of Plasmodium fakciparum. Up to 30%o of merozoites showed initial differentiation into trophic forms (rings) viable at 14 hr. However, only -1% of the merozoites would develop further into trophozoites and early schizonts viable at 36 hr. In efforts to increase the number of late stage parasites, we have now found a significant favorable effect of the addition of erythrocyte ghosts. Doubling the quantity of erythrocyte membrane in the erythrocyte sonicate medium resulted in approximate doubling of the number of trophozoites and early schizonts. These results indicate that components of the erythrocyte membrane are essential for the complete development of the erythrocytic cycle.We have described a method that supports a complete cycle of extracellular development in vitro of one of the many kinds of obligate intracellular parasitic protozoa, the malaria parasite Plasmodium falciparum (1,2). These results open a way to investigate the detailed relationships between the parasite and its host cell and to understanding of the nature of its dependence on the host cell. The importance of extracellular cultivation of intracellular parasites has been well emphasized by Moulder (3). At present, however, our methods do not provide for continuous extracellular culture. Although up to 30% of erythrocytic merozoites of P. falciparum will differentiate into young rings and be viable at 14 hr into their 48-hr cycle of schizogonic development, only -1% develop into trophozoites and early schizonts viable at 36 hr (1). Most of these go on to complete schizogony with formation of infective merozoites. These merozoites can again develop extracellularly (unpublished results) but their numbers are insufficient for continuous maintenance.Previous work (4) had shown that initial development was better in sonicated human erythrocyte extract, containing disrupted erythrocyte membranes, than in frozen-thawed extract from which the stroma had been removed. It therefore seemed reasonable to attempt to improve the medium by the provision of additional erythrocyte membrane. We present here results showing that constituents of the erythrocyte membrane favor the development of rings into trophozoites and schizonts.MATERIALS AND METHODS Parasites. Stock cultures of P. falciparum were maintained in flow vessels (5). We used two clones, HB-3 (6) of Honduras I/CDC isolate and A-2 of FCR-3/Gambia (7). Cultures were synchronized and purified to produce a 2-5% (vol/vol) schizont suspension as described (2). This suspension was incubated at 37°C for 1.25 hr and agitated at 10-min intervals. Incubated suspension was divided in two equal volumes and then centrifuged at 200 X g for 10 min to sediment schizonts. The supernatant containing merozoites was then centrifuged for 10 min at 1500 x g. The supernatant was discarded and the pellets containing merozoites were resuspended in a minimal amount of control or membrane-suppleme...

Single-cell evaluation of red blood cell bio-mechanical and nano-structural alterations upon chemically induced oxidative stress

“…A protective effect against malaria infection is associated with inherited disorders in RBCs, such as cytoskeleton disorders, surface antigen gene mutations, enzymatic deficiencies or hemoglobin alterations 31 . Several earlier studies also reported the effect of erythrocyte membrane on the growth of malaria parasites 32 33 .…”

Epidemiological studies of spectrin mutations related to hereditary elliptocytosis and spectrin polymorphisms in Benin

“…However, in vitro studies (Lecomte et al, 1991;Schulman et al, 1990) on the malarial infection of erythrocytes carrying genetically defined membrane abnormalities have shown the necessity of a qualitatively and quantitatively normal membrane skeleton for an efficient invasion and a complete maturation of the parasite. Studies of extracellular development of the erythrocyte cycle of Plasmodium falciparum have also shown that constituents of the erythrocyte membrane favour the development of rings into trophozoites and schizonts (Williams et al, 1995).…”

Cultivation of Plasmodium spp

“…Trager et al (90) postulated that contact between parasite and spectrin in the erythrocyte sonicate may be an important factor for development. More recently, Williams et al (99) have increased the yields of schizonts forming extracellularly by addition of erythrocyte ghosts obtained by osmotic lysis, to double the amount of membrane present. These results support the critical role of the erythrocyte membrane in the development of the malarial parasite.…”