Leishmania infantum is the causative agent of canine leishmaniasis in the Mediterranean area. The aspects of the canine immune response which may explain the chronic severe disease in these animals have not been well investigated. Therefore, in this work we have evaluated the parasite killing ability by circulating polymorphonuclear (PMN) cells and monocytes in dogs with spontaneous leishmaniasis and in healthy dogs. Moreover, in order to analyse the mechanisms responsible for killing activity, we evaluated the oxidative burst of circulating PMN cells, in terms of O2- and H2O2 production. The killing ability of infected dog PMN cells and monocytes was significantly lower than in a group of healthy dogs and a group of infected dogs treated with meglumine antimoniate and without symptoms of leishmaniasis. Superoxide anion and hydrogen peroxide production by PMN cells was significantly lower in a group of Leishmania-infected dogs than in healthy dogs, suggesting that these mediators may be involved in the reduced killing activity of infected dogs.