Delayed rejection in a leaf-cutting ant after foraging on plants unsuitable for the symbiotic fungus

Leaf-cutting ants culture a mutualistic fungus for which they collect and process a great diversity of fresh plant material as substrate. It has previously been observed that workers show delayed rejection behavior toward substrate that is harmful for the fungus but not for the ants: workers initially accept such materials but thereafter avoid its collection. In this study, we investigated delayed rejection behavior toward natural leaves in several 2-choice experiments in laboratory subcolonies of Acromyrmex lundi. We experimentally manipulated leaf suitability for the fungus by infiltrating them with a fungicide (cycloheximide) not detectable to the ants. The ants' delayed rejection behavior was specific toward the respective fungicide-treated plant species. Delayed rejection was also observed in naive ants after contact with the fungus garden containing treated leaves, confirming previous results with artificial bait. The onset of delayed rejection occurred 10 h after incorporation of treated leaves into the fungus garden. Rejection behavior was maintained for at least 9 weeks when incorporation of the previously unsuitable plant species was precluded. However, acceptance resumed after 3 weeks when ants were forced to feed on untreated leaves of the previously treated plant species. The observed species-specific, rapidly expressed, and flexible rejection of unsuitable substrate may be a mechanism to successfully avoid the provisioning of the fungus garden with plants containing harmful compounds as they occur in the highly diverse natural habitat of the colonies.