Cryptosporidium parvum is a zoonotic protozoan parasite that causes cryptosporidiosis. It is characterized by infection of the epithelial cells of the small intestines leading to acute watery diarrhea and consequently to dehydration that may lead to death in severe cases. Cryptosporidiosis represents a true one-health threat affecting humans and veterinary health. There is no vaccine and very limited chemotherapy available for humans and animals. Therefore, there is an urgent need for developing efficient curative compounds to control the disease. To do so, two complementary approaches will be used within this PhD project, with two collaborators, M.H. Hakimi (IAB, Grenoble) and C. Hedberg (Umea University) with the support of two funding sources. The first one has the purpose to chemically optimize an initial lead compound using structure activity relationship (SAR) to be used in ruminants as curative treatment against Cryptosporidiosis. We have generated 2 new Cryptosporidium parvum (INRAE) strains harboring fluorescent proteins (GFP/mCherry) and Nluc using the CRISP-cas9 technique. We are using these parasites in vitro and in vivo for precise/fine detection of parasite development. In our in vitro experiments, we compared the last generation of compounds by performing EC50 showing impressive activities in the low picomolar range and presenting low toxicity (MTS assay) leading to a selectivity index close to 106. The lead compound, administered orally, presents strong efficacy in vivo in the GKO mouse model and in newborn lambs, one of the target species of the parasite. Both parasite load and symptoms were severely reduced and no natural resistance observed so far even in the presence of suboptimal dosage. We have now a first highly promising candidate in hand for which we will determine the precise mechanism of action, impact of the treatment on microbiota, and the presence of secondary metabolites within lamb tissues following repeated oral treatments.