Apoptosis stalks Plasmodium falciparum maintained in continuous culture condition

Growth kinetic of Plasmodium falciparum in culture or in the host fall short of expected growth rate considering that there are 4 x 10(6)/µL red blood cell (RBCs) available for invasion and about 16 merozoites growing in each infected RBC. This study determined whether apoptotic machinery is operable to keep the parasite population under check.

A synchronized culture of P. falciparum (Dd2 strain) was initiated at 0.5% ring stage parasitaemia and kept under conditions not limiting for RBCs and nutrient by adjusting hematocrit to 5% at each schizogony and changing growth media daily. Parasite growth pattern and morphology was evaluated by blood smear microscopy and flow-cytometry using SYBR green. The apoptotic processes were evaluated for evidence of: DNA fragmentation by TUNEL, collapse of mitochondria membrane potential (ΔΨm) by TMRE, expression of metacaspase gene by RT-qPCR and by probing parasite proteins with anti-caspase antibodies.

From the seeding parasitaemia of 0.5%, the parasites doubled every 48 hours to a parasitaemia of 4%. Thereafter, the growth stagnated and the culture consistently crashed at about 6% parasitaemia. ΔΨm potential collapsed as the parasite density increased and DNA fragmentation increased steadily from 0.2% to ~6%. The expression of metacaspase gene and protein was observed in all stages, but their abundance was variable among the stages.

These findings suggest existence of P. falciparum quorum sensing that keep the parasite population under check.