Nicotinamide is a vitamin provided by food that is already used in human therapy. In Leishmania protozoan parasites, this molecule shows toxic activity against parasites and has synergistic activity with antimonials, the main drugs used to treat leishmaniasis. By investigating the mode of action of this cheap vitamin, we discovered that nicotinamide is in fact the main precursor of NAD+ synthesis in Leishmania, a redox cofactor essential for all living cells. Leishmania are indeed devoid of a de novo NAD+ pathway and must synthesize it by scavenging precursors from their environment (nicotinamide, nicotinic acid and nicotinamide riboside). This NAD+ auxotrophy reveals a mixed pattern of activity of nicotinamide in Leishmania, i.e. toxic at high concentrations but also essential for parasite survival through its role in NAD+ synthesis. All enzymes of the Leishmania NAD+ salvage pathway were then identified from genome databases. We focused on a putative nicotinamidase, which has no homolog in mammals and governs the conversion of nicotinamide to nicotinic acid, the first step in the NAD+ salvage pathway. Since this enzyme could be considered as an attractive therapeutic target to develop specific parasite inhibitors, we performed a functional analysis of the corresponding gene. Targeted deletion of the nicotinamidase encoding gene induced a marked drop in parasite NAD+ content and a phenotype with strongly delayed growth. Additionally, these mutants are unable to establish durable infections in mice. The crystal structure of the nicotinamidase from L. infantum will allow us to develop specific inhibitors against this new therapeutic target.