The control of malaria in Africa is challenged by increasing drug resistance. New artemisinin- based combination therapy (ACT) regimens are generally very effective, and have recently been adopted as standard therapy for uncomplicated malaria by nearly every country in Africa. However, heavy and repeated use of ACTs, as is now occuring, will lead to strong selective pressure for resistance to components of these regimens. Our understanding of mechanisms of resistance to ACTs is incomplete. We suggest that the best means of identifying resistance- mediating mutations before they are widespread will be to evaluate parasites that emerge soon after therapy, while they are under the selective pressure of long-acting ACT partner drugs. We hypothesize that treatment failures and the emergence of new infections soon after treatment with ACTs will be associated with known and unknown mutations in malaria parasites that alter responses to artemisinin partner drugs. To test this hypothesis, we will systematically evaluate parasites from recent and ongoing clinical trials in Uganda and Burkina Faso to identify associations between candidate mutations and clinical responses to ACTs. We further hypothesize that increasing use of ACTs will select for parasites with decreasing drug sensitivity, but also decreased fitness. To test this hypothesis, we will search for associations between treatment outcomes and in vitro measures of drug sensitivity and fitness. We will also study parasites with introduced mutations to test the impact of these alterations on in vitro measures of drug sensitivity and fitness. Our specific aims will be (1) to identify genotypes associated with decreased responses to ACTs in Africa, (2) to assess molecular mechanisms and parasitological consequences of increasing resistance to ACTs, and (3) to characterize the specific impacts of parasite polymorphisms on drug sensitivity and fitness. Our studies will offer important insights into mechanisms of resistance to the most important new antimalarial regimens. They will also have direct practical relevance, as they will identify molecular markers of resistance to key regimens, offer an "early warning system" for resistance by studying genotypes of parasites that emerge soon after treatment, and provide insight into the fitness consequences of increasing resistance. New combination therapies have recently become standard for the treatment of malaria, one of the greatest infectious disease problems in the world, due to increasing resistance to older drugs. This project involves studies of mechanisms of resistance of malaria parasites to new combination antimalarial therapies. These studies will have important public health benefits, as they will identify simple markers to identify resistance to new drugs before this problem is widespread, and they will offer insight into choices of the best new therapies for malaria.