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Splenic filtration of Plasmodium falciparum-infected red blood cells has been hypothesized to influence malaria pathogenesis. We have developed a minimum cylindrical diameter (MCD) filtration model which estimates physical splenic filtration during malaria infection. The key parameter in the model is the MCD, the smallest tube or cylinder that a red blood cell (RBC) can traverse without lysing. The MCD is defined by a relationship between the RBC surface area and volume. In the MCD filtration model, the MCD filtration function represents the probability of a cell becoming physically removed from circulation. This modelling approach was implemented at a field site in Blantyre, Malawi. We analysed peripheral blood samples from 120 study participants in four clinically defined groups (30 subjects each): cerebral malaria, uncomplicated malaria, aparasitaemic coma and healthy controls. We found statistically significant differences in the surface area and volumes of uninfected RBCs when healthy controls were compared with malaria patients. The estimated filtration rates generated by the MCD model corresponded to previous observations in ex vivo spleen experiments and models of red blood cell loss during acute malaria anaemia.There were no differences in the estimated splenic filtration rates between cerebral malaria and uncomplicated malaria patients. The MCD filtration model estimates that at time of admission, one ring-stage infected RBC is physically filtered by the spleen for each parasite that remains in peripheral circulation. This field study is the first to use microfluidic devices to identify rheological diversity in RBC populations associated with malaria infection and illness in well-characterized groups of children living in a malaria endemic area., (© 2012 Blackwell Publishing Ltd.)

The cellular events leading to severe and complicated malaria in some Plasmodium falciparum infections are poorly understood. Additional tools are required to better understand the pathogenesis of this disease. In this technical report, we describe a microfluidic culture system and image processing algorithms that were developed to observe cytoadhesion interactions of P. falciparum parasitized erythrocytes rolling on primary brain microvascularendothelial cells. We isolated and cultured human primary microvascular brain endothelial cells in a closed loop microfluidic culture system where a peristaltic pump and media reservoirs were integrated onto a microscope stage insert. We developed image processing methods to enhance contrast of rolling parasitized erythrocytes on endothelial cells and to estimate the local wall shear stress. The velocity of parasitized erythrocytes rolling on primary brain microvascularendothelial cells was then measured under physiologically relevant wall shear stresses. Finally, we deployed this method successfully at a field site in Blantyre, Malawi. The method is a promising new tool for the investigation of the pathogenesis of severe malaria.

Splenic filtration of infected red blood cells (RBCs) may contribute to innate immunity and variable outcomes of malaria infections. We show that filterability of individual RBCs is well predicted by the minimum cylindrical diameter (MCD) which is calculated from a RBC's surface area and volume. The MCD describes the smallest diameter tube or smallest pore that a cell may fit through without increasing its surface area. A microfluidic device was developed to measure the MCD from thousands of individual infected RBCs (IRBCs) and uninfected RBCs (URBCs). Average MCD changes during the blood-stage cycle of Plasmodium falciparum were tracked for the cytoadherent strain ITG and the knobless strain Dd2. The MCD values for IRBCs and URBCs raise several new intriguing insights into how the spleen may remove IRBCs: some early-stage ring-IRBCs, and not just late-stage schizont-IRBCs, may be highly susceptible to filtration. In addition, knobby parasites may limit surface area expansions and thus confer high MCDs on IRBCs. Finally, URBCs, in culture with IRBCs, show higher surface area loss which makes them more susceptible to filtration than naive URBCs. These findings raise important basic questions about the variable pathology of malaria infections and metabolic process that affect volume and surface area of IRBCs.

Malaria is a major poverty-related human infectious disease of the world. Over a billion individuals are under threat and several million die from malaria every year. The nature of disease, especially fatal disease, has been the subject of many studies. The consensus is that parasite-induced cytoadherance of red blood cells precipitates capillary blockage and inflammatory responses in affected organs. Reduced deformability of infected erythrocytes may also contribute to disease. What is not very clear is why people with significant parasite burdens display large variations in disease outcomes. Technologies which allow a detailed description of the cytoadherance properties of infected erythrocytes in individual patients, and which allow a complete description of the flow capabilities of red blood cell populations in that patient, would be very useful. Here we review the recent introduction of microfluidic technology to study malaria pathogenesis, including the fabrication processes. The devices are cheap, versatile, portable and require very small patient samples. With greater use in research laboratories and field sites, we eventually expect microfluidic methods to play important roles in malaria diagnosis, as well as prognosis.