Ring-stage parasitized erythrocytes (RPEs) were demonstrated to interact with effector cells of the innate immune system. With receptor blockade studies and CD36-null macrophages, human and murine macrophages were shown to phagocytose RPEs through the pattern recognition receptor CD36. These in vitro data implicate scavenger receptors in the clearance of RPEs.Phagocytic clearance of Plasmodium falciparum-parasitized erythrocytes (PEs) is an important line of innate defense against malaria (20,25). Pattern recognition receptors, including CD36, have been implicated in innate clearance of maturestage PEs (MPEs) by monocytes and macrophages (3,12,13,15,21,22,24,26). Ring-stage PEs (RPEs) have recently been shown to express ligands capable of interacting with endothelial cells (5,16,23,29). The objective of this study was to examine whether RPEs might also interact with effector cells of the innate immune system.We demonstrate that murine and human monocyte-derived macrophages can recognize and phagocytose RPEs (see Fig. 1 to 3). To ensure that only ring-stage parasites were used in phagocytosis assays (15, 21), we used procedures (2, 10) to generate highly synchronous ring-stage cultures (purity, Ͼ99% RPEs) and used cryopreserved malaria isolates and fresh clinical isolates that only contained RPEs (confirmed by microscopic criteria) (see Fig. 4). To ensure that only RPEs were quantified in phagocytic assays, we used a strict lysis procedure to remove adherent erythrocytes and morphological criteria (12,23) to ensure that only phagocytosed PE were counted. The slides were read blinded to the experimental conditions and confirmed by a second reader.To investigate the molecular mechanisms underlying RPE phagocytosis, we undertook receptor blockade studies with monoclonal antibodies (MAbs) against macrophage receptors known to interact with PEs and against macrophage pattern recognition receptors (4, 9) (Fig. 1). Only CD36 receptor blockade resulted in a significant decrease in RPE or MPE phagocytosis. Inhibition of phagocytosis was confirmed with two separate parasite lines: 3D7 (30) and ITG (17).To confirm the role of CD36 in mediating RPE uptake, we examined phagocytosis by CD36 wild-type and CD36-null macrophages (7, 15). Phagocytosis of RPEs was reduced by ϳ90% in CD36-null macrophages. To investigate whether other macrophage receptors might cooperate with CD36 or mediate uptake in the absence of CD36 (1,6,14,18,19,28,29), we performed phagocytosis assays with wild-type and CD36-null macrophages in the presence of MAbs to other macrophage surface receptors. MAb blockade of receptors other than CD36 did not significantly decrease uptake of RPEs ( Fig. 2A). Since pattern recognition receptors such as the mannose receptor have also been implicated in opsonin-independent phagocytosis of P. chabaudi (26), we investigated whether the mannose receptor contributed to RPE uptake. As shown in Fig. 2A, mannan did not significantly decrease uptake of RPEs but did inhibit uptake of MPEs compared to -glucan treatment (data no...