The symbiotic association of the leaf-cutter ants with the fungus Leucoagaricus gongylophorus implies that the negative impact on the fungi will affect the survival of the ants. After a promising antifungal bioactivity, Carica papaya was studied leading to the bioassay-guided isolation of squalene, 4,5-dihydrohumulen, the flavanol-glycosides quercetin, kaempferol, and rutin, and the alkaloid carpaine. Carpaine and squalene were found to have a moderate individual activity, which increased when evaluated together. Based on the presence of these compounds, standardized extracts were formulated for field tests on nests of Acromyrmex octospinosus. All the nests treated were negatively affected evidencing a promising lead for the integrated management of leaf-cutter ants. © 2019, © 2019 Informa UK Limited, trading as Taylor & Francis Group.