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Leaf-cutting ants are the most important pests in several cropping systems in the Neotropics. Granulated baits containing active ingredients, considered hazardous by the Stockholm Convention, are the usual method to control these ants. Isocycloseram is a new insecticide molecule with high safety margin for mammals, but without registration for the ants in general. Thus, this study investigated the effectiveness of granulated baits with isocycloseram in leaf-cutting ants control under laboratory and field conditions. Initially, the mortality of Atta sexdens workers, fed with dehydrated citrus pulp paste containing different concentrations of isocycloseram was evaluated in the laboratory for 21 days, for toxicological classification. Subsequently, the loading, devolution, and incorporation of baits with different concentrations of isocycloseram and the mortality of A. sexdens colonies were evaluated in the laboratory. After that, the percentages of loading and devolution of baits, foraging activity, and colony mortality treated with 0.05, 0.1, 0.2, and 0.3% of isocycloseram were evaluated for the species A. sexdens, A. laevigata, and Acromyrmex lundii in field conditions. All concentrations of isocycloseram killed more than 15% of ants in 24 h and more than 90% in 21 days in the laboratory, being classified as a fast-acting and highly effective active ingredient. Baits with 0.001 to 0.03% of isocycloseram were highly loaded and exhibited low rate of devolution. The mortality of A. sexdens colony was higher at concentrations between 0.075 and 0.3%, in the laboratory. Baits containing isocycloseram at concentrations of 0.2 and 0.3% were highly loaded, presented low devolution rates, and were highly efficient in controlling A. sexdens, A. laevigata, and A. lundii in the field, at dosages of 6, 10, and 12 g/m² of nest. This is the first report of the use of isocycloseram against leaf-cutting ants, contributing to the development of efficient and toxicologically safer ant baits.