U.S. Department of Energy

Pacific Northwest National Laboratory

The Ants March To Keep Their Fungus Gardens Thriving

Figure 1 Leaf cutter ants carrying various substrates (A) a leaf, (B) a flower and (C) an oat. Ants tending to their fungus garden with newly incorporated leaf material (D) (photographs by Don Parsons).

Leaf cutter ants area classic example of foraging herbivores that use symbiotic microbes to get energy and nutrients from plant material. According to researchers, a spongy fungal species known as L. gongylophorus,has cooperated with leaf cutter ants for the last 23 million years.

The ants live in colonies and forage up to 20 plant species to create “fungus gardens” that are sometimes referred to as the “gut” of the ants. The ant-fungus relationship is simple: The ants bring various plant material into the garden, combine it with fecal matter and saliva, and form it into a mass that L. gongylophorus uses as a substrate for growth. The ants then feed off of swollen tips, called gongylidia, that form on the fungus. Repeat.

In a recent paper, published in Molecular Ecology,PNNL researchers and collaborators investigate the metabolic flexibility of these leaf cutter fungal gardens.Using feeding experiments and a novel approach in metaproteomics, they examined the enzymatic response of L. gongylophorusto leaves, flowers, oats, and a mixture of all three.


Researchers excavated five colonies from Panama between December 2012 and January 2013. Five fungus chambers were excised from each colony, and each of the five chambers were further split into four sub-colonies. Each colony was assigned to one of four feeding treatments: leaves, flowers, Quaker instant oatmeal, or a mixture of all three.

Of the three substrates leaves are the most recalcitrant. Flowers have a similar wall structure to leaves, but overall they’re easier to digest. Oats are highly processed, but have the most accessible energy.

Scientists hypothesized that L. gongylophorus would respond to different plant substrates by producing specific proteins that can break down whatever food is provided. They were right.


Using metaproteomic data processing and statistical analysis, the scientists identified and quantified 1,766 different fungal proteins within the gardens, including 161 putative biomass-degrading enzymes. Furthermore, they found significant differences in protein profiles in fungus gardens that were treated with different plant substrates.  They found the gardens produced a higher abundance of proteins when they needed to break down the tougher substrates, such as leaves and flowers. But there was a low variability of proteins amongst the leaf- and flower-treated colonies. The opposite was true for the gardens treated with oats and mixed substrates: There was a greater variability, with some overlap in proteins. 

The sub-colonies fed exclusively with oats experienced a significantly lower survivorship in the gardens. Scientists hypothesized that an exclusive diet of oats lacks the necessary micronutrients that ants, fungus, and bacteria take from fresh plant material. Grain-fed cows show similar responses to such a diet. They experience rapid weight gain, but often suffer from a metabolic disease known as ruminal acidosis. 

The scientists were surprised when they examined the differences between the gardens treated with leaves and flowers and those receiving mixed treatments. For example, when treated with only leaves and flowers, the gardens produced more proteins expected to break down cellulose. That supports the argument for the role of fungus in recalcitrant biomass degradation.  The mixed-treatment gardens, however, didn’t generate a lot of biomass-degrading enzymes. Instead, these gardens first derived energy from the easily digestible food (oats) before tackling the more recalcitrant substrates. This metabolic flexibility, the scientists say, contributes to the ability of leaf cutter ants to be herbivores of the “dominant generalist” type – that is, herbivores that thrive in changing seasons and locations. 


The breakdown of biomass by L. gongylophorus is central to the success of leaf cutter ant colonies and for the function of the ant-fungus mutualism. In a broader sense, though, the paper’s results are an important step in understanding microbial mediation of a generalist herbivore system. This study details the substrate-specific enzymatic response of L. gongylophorus thereby giving unprecedented insight into molecules responsible for digesting cellulose and other recalcitrant material in this symbiotic system. 


This work was funded in part by the Department of Energy Great Lakes Bioenergy Research Center and the DOE Office of Biological and Environmental Research, Genomic Science Program.


Khadempour, L., Burnum-Johnson, K. E., Baker, E. S., Nicora, C. D., Webb-Robertson, B.-J. M., White, R. A., Monroe, M. E., Huang, E. L., Smith, R. D. and Currie, C. R. (2016), "The fungal cultivar of leaf-cutter ants produces specific enzymes in response to different plant substrates". Mol Ecol, 25: 5795–5805. doi:10.1111/mec.13

October 2016
| Pacific Northwest National Laboratory