The physiological basis for the resistance to falciparum malaria o individuals with sickle cell trait has not been understood. Recent advances in erythrocytic Plasmodium falciparum culture have made possible a direct investigation of the development of the malaria parasite in cells with sickle cell hemoglobin. In a high (18%) oxygen atmosphere, there is no apparent sickling of cells, and the growth and multiplication of P. falciparum is identical in normal (AA), hemoglobin S homozygous (SS), and hemoglobin S heterozygous (SA) erythrocytes. Cultures under low (1-5%) oxygen, however, showed clear inhibition of growth. The sickling of SS red cells killed and lysed most or all of the intracellular parasites. Parasites in SA red cells were killed primarily at the large ring stage, probably as a result of a disruption of the parasite metabolism. Incubation in cyanate prior to culture reversed the resistance of SA erythrocytes to plasmodium growth, but had no effect on SS red cell sickling or resistance. Thus, the mechanism of resistance in vivo may be due solely to intraerythrocytic conditions. In 1949, the elucidation of the genetic basis for sickle cell anemia (SS) and sickle cell trait (SA) (1, 2) and the demonstration of the involvement of an abnormal hemoglobin in the disease (3) formed a link between genes and molecules. At the same time, medical scientists working in Africa suggested that sickle cell gene carriers were resistant to falciparum malaria (4, 5), thereby forging a connection between molecules and selective pressure. It was proposed that the inheritance of this variant hemoglobin gene conferred resistance in the heterozygous state to falciparum malaria, which is often lethal in the early years of life. The greater ability of heterozygotes to survive to the age of reproduction was thought to balance the almost complete mortality of the homozygous state. In the entire chain between genes and evolution, the weakest link has been that describing the intimate relationship between sickle cell hemoglobin (Hb S) and the malaria parasite, Plasmodium falciparum. Population studies have confirmed that children heterozygous for Hb S are protected, statistically, from death, though not infection, by P. falciparum (6). Thus far, however, a direct connection between sickling and resistance has remained suggestive and speculative.The erythrocytic cycle of the parasite proceeds from an infective form, the merozoite. Infection occurs by induced endocytosis, and the growth of the early ring to a trophozoite follows. Nuclear division precedes segmentation of the mature schizont into 16 or more merozoites, which are released upon lysis of the red blood cell. The entire cycle requires 48 hr, the latter two-thirds with the parasitized cell sequestered in the vascular network of the tissues. The periodic fever of malaria results from the synchronous lysis of infected cells upon maturation of the parasite.In the Hb S-containing red cell, the parasite may not find as perfect a host as the one to which it is adapted. ...

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The kinetics of sickling of malaria-infected red cells from humans with sickle cell trait were studied in vitro in an attempt to obtain direct experimental evidence for a selective advantage of the hemoglobin S heterozygote in a malarious region. The sickling rates of cells infected with Plasmodium falciparum and of non-infected cells were studied both in the total absence of oxygen (by dithionite addition) and at several different concentrations of oxyhemoglobin which might obtain in vivo. In all cases, red cells containing small plasmodium parasite forms (ring forms) sickled approximately eight times as readily as uninfected cells. Cells containing large parasitic forms (trophozoites and schizonts) appeared to sickle less readily than uninfected cells, by light microscopy criteria, but electron micrographs demonstrated the presence of polymerized deoxyhemoglobin S with a high frequency. It is concluded that enhanced sickling of plasmodium-infected AS cells may be one mechanism whereby the hemoglobin S polymorphism is balanced in favor of the heterozygote.

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A high frequency of nonhemolytic hereditary ovalocytosis in Malayan aborigines is thought to result from reduced susceptibility of affected individuals to malaria. Indeed, Kidson et al. recently showed that ovalocytes from Melanesians in Papua New Guinea are resistant to infection in culture by the malarial parasite Plasmodium falciparum. In order to determine if protection against parasitic invasion in these ovalocytes might be the result of some altered membrane material property in these unusual cells, we measured their membrane and cellular deformability characteristics using an ektacytometer . Ovalocytic red cells were found to be much less deformable in comparison to normal discoid red cells. Similar measurements on isolated membrane preparations revealed a marked reduction in ovalocytic membrane deformability. To produce equal deformation of ovalocytic and normal membranes, ovalocytes required an 8–10-fold increase in applied shear stress, indicating that their membrane was capable of deforming under sufficient stress. To test the possibility that this increased membrane rigidity might confer resistance to parasitic invasion, we performed an in vitro invasion assay using Plasmodium falciparum merozoites and Malayan ovalocytes of varying deformability from seven different donors. The level of infection of the ovalocytes ranged from 1% to 35% of that in control cells, and the extent of inhibition appeared to be closely related to the reduction in membrane deformability. Moreover, we were able to induce similar resistance to parasitic invasion in nonovalocytic normal red cells by increasing their membrane rigidity with graded exposure to a protein crosslinking agent. Our findings suggest that resistance to parasite invasion of Malayan ovalocytes is the result of a genetic mutation that causes increased membrane rigidity.