Do fungus-farming leaf cutter ants smell like fungus?

Figure 1

Superficial view of an Atta bisphaerica leaf-cutting ant nest, in the Brazilian savannah. Photo: Lohan Valadares

A blog post highlighting the article by L. Valaderes and F.S. do Nascimento in Insectes Sociaux

By Lohan Valadares  (@valadareslohan, @lo.han.792)

About 30 million years ago, in the South American savannahs, ants that cultivate fungus have over evolutionary time become the leaf-cutting ants (genera Atta and Acromyrmex). These ants reside in large subterranean nests of the Neotropical region, with galleries and fungus chambers that shelter thousands to millions of individuals, where they farm the fungus Leucoagaricus gongylophorus. The fungus has never been found free-living without the ants, and it is generally accepted that both organisms have co-evolved into an obligate mutualism, which means that one organism cannot live without the other. For the ants, the fungus is priceless as it serves as a rearing site and unique nutritional substrate for brood development. In exchange, the ants protect the fungus against parasites and forage for fresh vegetation, which is used as substrate for fungal growth. Recently, chemical analyses of the fungus gardens of Acromyrmex species have revealed the presence of hydrocarbons, a class of chemical compound found commonly on the cuticle of insects (Viana et al., 2001, Richard et al., 2007). These two species are so intertwined, however, that whether it is the ants or the fungus which are the ultimate producers of these hydrocarbons is not yet clear.

Cuticular hydrocarbons (CHCs) are of particular importance for social insects, and decades of studies have demonstrated that they are important as nestmate recognition cues. Consequently, variations in these substances have been associated with caste, age, dominance hierarchy, worker subcastes, and developmental stages. Furthermore, the leaves used as the fungus substrate seem to be an important source of CHCs for leaf-cutting ants (Richard et al., 2004, Valadares et al., 2015). This leads us to hypothesise that the intermediate organism in this process – the symbiotic fungus – is actually the one promoting the transference of these substances, or may actually be involved in the synthesis of hydrocarbons itself.

To explore the resemblance between the chemical compounds of both organisms involved in this symbiotic association, we studied the resemblance between the CHC profile of fungal cultivars with those of the ants, focusing on the changes in the CHC composition during the ants’ larval-to-adult moulting in the leaf-cutting ant Atta sexdens and its associated fungus. Our main question was: how do the changes in the CHC profile associated with the moulting cycle correlate with the hydrocarbon profile of the fungus cultivated by the ant Atta sexdens? To assess it, we collected pieces of fungal mycelium, and ant larvae, pupae and adult workers from laboratory populations, and soaked them in a solvent to extract their hydrocarbons, which later on were injected into a gas chromatography–mass spectrometry (GC–MS) for separation and identification of hydrocarbons.

Figure 2

An excavated fungus chamber of an Acromyrmex subterraneus leaf-cutting ant nest, in the Brazilian savannah. Photo: Lohan Valadares

Our analyses demonstrated that ant and fungus shared 58% of hydrocarbons, and the rest were all ant-specific hydrocarbons that were absent from the symbiotic fungus. Our comparative analyses revealed a great similarity between the hydrocarbon profiles of larva and fungus, due to the fact that both groups shared mainly highly concentrated linear hydrocarbons. As individuals progressed through developmental stages, the chemical profiles between ant and fungus became increasingly different. As the moulting cycle progressed, the hydrocarbon profile was characterised by a great shift from the ‘less pronounced’ state of the brood’s chemical profile towards a more diverse chemical profile of adult workers comprised of highly concentrated branched hydrocarbons.

Our findings coincide with the observations made for Acromyrmex leaf-cutting ants (Vianna et al. 2001, Richard et al. 2007), strengthening the evidence for the intimate relationship between brood and fungus as a variable shaping the hydrocarbon profile of both species. However, because we don’t know in detail how the hydrocarbons are biologically synthesised and transferred between these organisms, it is difficult to interpret whether the ants or the fungus (or even both organisms) are the ones actually synthesising hydrocarbons. However, we believe that our descriptive and comparative analyses indicating such strong resemblances has suggested new exciting opportunities of research tha­t will help us to understand how this co-evolutionary relationship has shaped the chemical profile of both organisms. If we consider the optimistic scenario where the fungus is actually the one to be transferring these compounds to the ants, how does this transference impact colony odour? Do these substances play a role in nestmate recognition? Several questions remain to be answered and hopefully our work has contributed to the journey towards the understanding of the chemo-ecology between leaf-cutting ants and their fungal cultivars.



Branstetter, M.G., Jesovnik, Aa, Sosa-Calvo, J., Lloyd, M.W., Faircloth, B.G., Brady, S.G., Schultz, T.R. (2017). Dry habitats were crucibles of domestication in the evolution of agriculture. Proceedings Royal Society B. 284: 20170095.

Richard, F.J., Heftz, A., Christides, J.P., Errard, C. (2004) Food influence on colonial recognition and chemical signature between nestmates in the fungus-growing ant Acromyrmex subterraneus subterraneus. J Chem Ecol 14: 9–16

Richard, F.J., Poulsen, M., Hefetz, E.C., Nash, D.R., Boomsma, J.J. (2007). The origin of the chemical profiles of fungal symbionts and their significance for nestmate recognition in Acromyrmex leaf-cutting ants. Behav Ecol Sociobiol 61(11):1637-1649

Valadares, L., Nascimento, D., Nascimento, F.S. (2015) Foliar substrate affects cuticular hydrocarbon profiles and intraspecific aggression in the leafcutter ant Atta sexdens. Insects 6: 141-151

Viana, A.M., Frézard, A., Malosse, C., Della Lucia, T.M., Errard, C., Lenoir, A. (2001) Colonial recognition of fungus in the fungus-growing ant Acromyrmex subterraneus subterraneus (Hymenoptera: Formicidae). Chemoecol 11(1): 29-36

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