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Leaf-cutter ants cultivate and feed on the mutualistic fungus, Leucoagaricus gongylophorus, which is threatened by parasitic fungi of the genus Escovopsis. The mechanism of Escovopsis parasitism is poorly understood. Here, we assessed the nature of the antagonism of different Escovopsis species against its host. We also evaluated the potential antagonism of Escovopsioides, a recently described fungal genus from the attine ant environment whose role in the colonies of these insects is unknown. We performed dual-culture assays to assess the interactions between L. gongylophorus and both fungi. We also evaluated the antifungal activity of compounds secreted by the latter on L. gongylophorus growth using crude extracts of Escovopsis spp. and Escovopsioides nivea obtained either in (1) absence or (2) presence of the mutualistic fungus. The physical interaction between these fungi and the mutualistic fungus was examined under scanning electron microscopy (SEM). Escovopsis spp. and E. nivea negatively affected the growth of L. gongylophorus, which was also significantly inhibited by both types of crude extract. These results indicate that Escovopsis spp. and E. nivea produce antifungal metabolites against the mutualistic fungus. SEM showed that Escovopsis spp. and E. nivea maintained physical contact with the mutualistic fungus, though no specialised structures related to mycoparasitism were observed. These results showed that Escovopsis is a destructive mycoparasite that needs physical contact for the death of the mutualistic fungus to occur. Also, our findings suggest that E. nivea is an antagonist of the ant fungal cultivar.

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A termite colony is usually founded by a pair of alates, the primary reproductives, which produce all the nestmates. In some species, secondary reproductives appear to either replace the primaries or supplement colony reproduction. In termites, secondary reproductives are generally ergatoids derived from workers or nymphoids derived from nymphs. Silvestritermes euamignathus is a termite species that forms multiple nymphoid reproductives, and to date it was hypothesized that these secondary reproductives were the progeny of the primary founding reproductives. We developed markers for 12 microsatellite loci and used COI mitochondrial DNA (mtDNA) to genotype 59 nymphoid neotenics found in a colony of S. euamignathus to test this hypothesis. Our results showed that nymphoids of S. euamignathus are not all siblings. The microsatellite analysis suggests that the secondary reproductives derived from a minimum of four different pairs of reproductives belonging to at least two different matrilines. This is the first record of non-sibling secondary reproductives occupying the same nest in a higher termite. These unrelated reproductives might be the result of either pleometrotic colony foundation or colony fusion.