Experimental evidence of a tripartite mutualism: bacteria protect ant fungus gardens from specialized parasites

Symbioses shape all levels of biological organization. Although symbiotic interactions are typically viewed as bipartite associations, with two organisms interacting largely in isolation from other organisms, the presence and importance of additional symbionts is becoming increasingly more apparent. This study examines the importance of a third mutualist within the ancient symbiosis between leaf‐cutting ants and their fungal cultivars. Specifically, we experimentally examine the role of a filamentous bacterium (actinomycete), which is typically carried on the cuticle of fungus‐growing ants, in suppressing the growth of a specialized microfungal parasite ( Escovopsis ) of the fungus garden. We conducted two‐by‐two factorial design experiments crossing the presence/absence of actinomycete with the presence/absence of Escovopsis within small sub‐colonies of Acromyrmex octospinosus . In these experiments, infection by Escovopsis became much more extensive within fungus gardens and had a greater impact on the health of gardens in those sub‐colonies with the bacterium removed from workers as compared to gardens with the bacterium still present on the ants. We establish that the actinomycete bacterium is most abundant on those major workers tending the garden, providing further support that the bacterium is involved in garden hygiene. We also found a significantly higher abundance of actinomycete on workers in colonies experimentally infected with Escovopsis as compared to uninfected control colonies. We suggest that mutualisms between antibiotic‐producing microbes and higher organisms may be common associations that are mostly overlooked and that the role of symbionts in reducing the impact of parasites is likely an important aspect in the cost‐benefit assessment of mutualisms.