The occurrence of rosetting of Plasmodium falciparum-infected human red blood cells (IRBC) with uninfected red blood cells (RBC) and its potential pathophysiologic consequences were investigated under flow conditions using the perfused rat mesocecum vasculature. Perfusion experiments were performed using two knobby (K+) lines of P falciparum, ie, rosetting positive (K+R+) and rosetting negative (K+R-). The infusion of K+R+ IRBC resulted in higher peripheral resistance (PRU) than K+R- IRBC (P less than .0012). Video microscopy showed that under conditions of flow, in addition to cytoadherence of K+R+ IRBC to the venular endothelium, rosette formation was also restricted to venules, especially in the areas of slow flow. Rosettes were absent in arterioles and were presumably dissociated by higher wall shear rates. The presence of rosettes in the venules must therefore reflect their rapid reformation after disruption. Cytoadherence of K+R+ IRBC was characterized by formation of focal clusters along the venular wall. In addition, large aggregates of RBC were frequently observed at venular junctions, probably as a result of interaction between flowing rosettes, free IRBC, and uninfected RBC. In contrast, the infusion of K+R+ IRBC resulted in diffuse cytoadherence of these cells exclusively to the venular endothelium but not in rosetting or large aggregate formation. The cytoadherence of K+R+ IRBC showed strong inverse correlation with the venular diameter (r = -.856, P less than .00001). Incubation of K+R+ IRBC with heparin and with monoclonal antibodies to glycoprotein IV/CD36 abolished the rosette formation and resulted in decreased PRU and microvascular blockage. These findings demonstrate that rosetting of K+R+ IRBC with uninfected RBC enhances vasocclusion, suggesting an important in vivo role for rosetting in the microvascular sequestration of P falciparum-infected RBC.