Inhibitory effect of a fava bean component on the in vitro development of Plasmodium falciparum in normal and glucose-6-phosphate dehydrogenase deficient erythrocytes

Abstract

We examined the hypothesis that G-6-PD deficiency associated with fava bean ingestion confers resistance to malaria by studying the in vitro interactions between malaria parasites (Plasmodium falciparum), human erythrocytes with varying degrees of G-6-PD deficiency, and isouramil (IU), a fava bean extract that is known to cause oxidant stress and hemolysis of G-6-PD-deficient erythrocytes. Untreated G-6-PD-deficient and normal erythrocytes supported the in vitro growth of P. falciparum equally well. However, after pretreatment with IU, G-6-PD-deficient erythrocytes did not support parasite growth in vitro, whereas growth remained high in normal erythrocytes. Parasite growth was proportional to the G-6-PD activity of the IU-treated erythrocytes. In contrast, when parasitized erythrocytes were exposed to IU, parasites even in normal erythrocytes were destroyed. Ring forms were much less sensitive than late trophozoites and schizonts. The results suggest that there are two modes by which IU affects the development of P. falciparum and demonstrate in vitro that G-6-PD deficiency confers resistance against malaria under conditions of fava-bean-associated oxidant stress.

Resistance of glucose-6-phosphate dehydrogenase deficiency to malaria: effects of fava bean hydroxypyrimidine glucosides onPlasmodium falciparumgrowth in culture and on the phagocytosis of infected cells

“…We find that vicine and convicine inhibit the growth of P. falciparum inside normal RBC and more so in G6PD" cells. This inhibition was further exacerbated when /?-glucosidase was added to the culture medium, implicating the aglycones in the anti-malarial activity, in agreement with previous reports on the anti-malarial effect of the aglycones and related to the diminished antioxidant capacity of G6PD^ RBC (Clark et al 1984;Golenser et al 1983Golenser et al , 1988. To explain inhibition in the absence of added enzyme, we were tempted to suggest that the glucosides as such could penetrate into infected cells through the new permeability pathways induced by the parasite in the membrane of their host cell (Ginsburg, 1994).…”

“…Such selection may be mediated by the faster elimination of infected cells or by more efficient killing of parasites residing in G6PD" RBC. Of the fava bean aglycones, divicine was shown to be toxic to P. vinckei in vivo (Clark et al 1984) and isouramil toxic to P. falciparum in culture (Golenser et al 1983(Golenser et al , 1988. Toxicity may be mediated by the oxidation of pyridine nucleotides and GSH and the production of H 2 O 2 (Chevion et al 1982;Benatti et al 1985;Winterbourn, Benatti & De Flora, 1986).…”

“…Toxicity may be mediated by the oxidation of pyridine nucleotides and GSH and the production of H 2 O 2 (Chevion et al 1982;Benatti et al 1985;Winterbourn, Benatti & De Flora, 1986). More importantly, G6PD" but not normal RBC pre-treated with isouramil did not support parasite growth (Golenser et al 1983), probably due to the depletion of GSH and ATP, the limited ability of G6PD" RBC to replenish GSH (Mager et al 1965) and the increase of superoxide dismutase and reduction of glutathione peroxidase activities (Mavelli et al 1984). Parasites in G6PD~ RBC were more sensitive to isouramil than those thriving in normal RBC (Golenser et al 1988).…”

“…Since partial inhibition of parasite growth in vitro on its own is incapable of explaining the high frequency of the otherwise potentially deleterious G6PD deficiency in human populations, one must invoke a combination of host response, and possibly with dietary oxidants such as the aglycones of fava beans pro-oxidant hydroxypyrimidines vicine and convicine (Clark et al 1984;Golenser et al 1983Golenser et al , 1988Greene, 1993). The pyridine-^-glucosides vicine (2,6-diamino-4,5-dihydroxypyrimidine 5-(/?-D-glucopyranoside) and convicine (2,4,5-trihydroxy-6-aminopyrimidine 5-(/?-D-glucopyranoside) constitute the toxic compounds of fava beans, responsible for the haemolytic crisis in bearers of G6PD~ (Arese, 1982;Luzzatto & Mehta, 1989;Arese & De Flora, 1990).…”

“…Plasmodium falciparum that grows in G6PD~ cells is more sensitive than that which inhabits normal RBC to isouramil (Golenser et al 1983), and divicine kills P. vinckei in mice (Clark et al 1984). It has therefore been suggested that the balanced polymorphism of G6PD" that has been acquired under the pressure of malaria must have invoked a combination of host response and dietary oxidants such as those found in fava beans which serve as staple food in some endemic malarious areas (Huheey & Martin, 1975;Golenser et al 1983;Clark et al 1984;Clark & Cowden, 1985;Greene, 1993). In the present investigation we have attempted to shed further light on the mechanistic details that may account for the resistance of G6PD" RBC to malaria.…”