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Abstract

We have previously provided evidence for a pathway in Plasmodium falciparum-infected erythrocytes, coined the parasitophorous duct pathway, which provides serum (macro)molecules direct access to intraerythrocytic parasites [1]. The present study addresses the purity of the fluorescent macromolecules used to define the duct pathway and provides ultrastructural evidence for its presence. The fluorescent tracers used to characterize transport remain intact during their incubation with infected erythrocytes. Transport of macromolecules in the external medium or host cell cytosol to the intracellular parasites is shown to occur by two distinct pathways. Fluorescent dextrans in the erythrocyte cytosol are ingested by the parasite via a specialized organelle, the cytostome, and are transported to the parasite food vacuole. Transport through this pathway occurs throughout the asexual life cycle. By contrast, fluorescent dextrans in the external medium bypass the erythrocyte cytosol, and are internalized by the parasite by a process resembling fluid-phase endocytosis. Serial sections of mature parasites fixed and stained by various methods for transmission electron microscopy reveal areas of apparent membrane continuity between the erythrocyte membrane and the parasitophorous vacuolar membrane that surrounds the parasite, that could leave the parasites exposed to the external medium. Using carboxylate and amidine-modified fluorescent latex spheres and laser scanning confocal microscopy, macromolecules up to 50–70 nm in diameter are found to have direct access to intraerythrocytic parasites. This size exclusion is consistent with the dimensions of the parasitophorous duct pathway revealed by electron microscopy. This investigation reports for the first time the existence of two, distinct macromolecular transport pathways in malaria-infected erythrocytes.