The Evolution of Biomineral Armor in Fungus-farming Ants

Biomineralization has enabled remarkable adaptations across the animal kingdom, including defensive structures and rock-grinding teeth. Although calcium carbonate is a common biogenically formed mineral found in all extant animal phyla, it is only recently known in insects and almost universally occurs with minimal magnesium content. A high-Mg calcite armor layer, covering most of the exoskeleton, was recently discovered in the fungus-farming ant Acromyrmex echinatior. Here, using a combination of scanning electron microscopy, energy-dispersive spectroscopy, and X-ray diffraction, paired with genome-scale phylogenetic analysis based on ultraconserved elements (UCEs), we characterize the evolution of biomineralization in fungus-farming ants. We show the presence of high-Mg calcite biomineral in 35 of 50 fungus-farming ant species studied, including taxa that span all major agricultural lineages. The biomineral is absent in all 16 myrmicine outgroup ant species, indicating an origin of the biomineral within ant agriculture, with ancestral state reconstruction analysis slightly supporting multiple origins within the fungus-farming ant clade. We then show that the occurrence of the biomineral is closely associated with the ancient mutualism with antimicrobial-producing Pseudonocardia, finding biomineral presence to be significantly correlated with the Pseudonocardia symbiont and the cuticular symbiotic structures that maintain these bacteria. Furthermore, we reveal significant inter-species variation in mineral composition and coverage, as well as intra-colony caste-specific differences, highlighting its adaptive complexity. Collectively, this work demonstrates that the innovation of a high-Mg calcite armor is strongly associated with the evolution of ant agriculture, revealing a rare biomineral to be widespread in the fungus-farming ants.