Plasmodium falciparum (P. falciparum) and Plasmodium knowlesi (P. knowlesi) are protozoan pathogens that cause malaria. Of the five species of Plasmodium that infect humans, P. falciparum and P. knowlesi both involve the sequestration of parasitized red blood cells in the deep tissue microvasculature and both cause a range of disease severity. The goal of the proposed study is to understand how fluctuations in the oxygen concentration in the microenvironment, which varies across tissue sites in the body, affect the multiplication rate of P. falciparum and P. knowlesi. The findings from this study are significant toward our understanding of antimalarial drug resistance, as some of the most important antimalarial drugs have been shown to work by triggering increased reactive oxygen species (ROS) in the parasite, and in vitro studies of drug-resistant Plasmodium under different oxygen concentrations have shown variable results. While it is well-known that Plasmodium grows best in the laboratory under low oxygen conditions, little is known about the biology of the parasite under higher oxygen conditions. Our preliminary in vitro data suggests that P. falciparum replicates significantly slower at 13% oxygen (mimicking the oxygen concentration in lungs and liver) versus 1% oxygen (mimicking the oxygen concentration in brain and bone marrow). Confocal imaging suggested lower mitochondrial activity in the 13% condition. Here in Aim 1, we will characterize the cellular and molecular mechanism(s) underlying the variability in multiplication rate under different oxygen conditions, studying this phenomenon in diverse species and strains of Plasmodium. We hypothesize that as oxygen in the microenvironment increases, an increase in intracellular ROS and cellular damage follows, leading to a decline in multiplication rate. In Aim 2, we will directly test whether the parasite’s redox balance affects Plasmodium multiplication rate, through manipulation of ROS and P. falciparum’s glutathione antioxidant system. While much focus exists on characterizing genetic mutations underlying drug resistance, very little work explores the host environmental factors that may also play a role in the redox balance of the parasites, which may in turn contribute to antimalarial drug efficacy in vivo.