Immunity and survival response of Atta cephalotes (Hymenoptera: Myrmicinae) workers to Metarhizium anisopliae infection: Potential role of their associated microbiota.

Leaf-cutting ants of the genera Atta and Acromyrmex are at constant risk of epizootics due to their dense living conditions and frequent social interactions between genetically related individuals. To help mitigate the risk of epizootics, these ants display individual and collective immune responses, including associations with symbiotic bacteria that can enhance their resistance to pathogenic infections. For example, Acromyrmex spp. harbor actinobacteria that control infection by Escovopsis in their fungal gardens. Although Atta spp. do not maintain symbiosis with protective actinobacteria, the evidence suggests that these insects are colonized by bacterial microbiota that may play a role in their defense against pathogens. The potential role of the bacterial microbiome of Atta workers in enhancing host immunity remains unexplored. We evaluated multiple parameters of the individual immunity of Atta cephalotes (Linnaeus, 1758) workers, including hemocyte count, encapsulation response, and the antimicrobial activity of the hemolymph in the presence or absence of bacterial microbiota. Experiments were performed on ants reared under standard conditions as well as on ants previously exposed to the entomopathogenic fungus Metharrizium anisopliae. Furthermore, the effects of the presence/absence of bacteria on the survival of workers exposed to M. anisopliae were evaluated. The bacterial microbiota associated with A. cephalotes workers does not modulate the number of hemocytes under control conditions or under conditions of exposure to the fungal pathogen. In addition, infection by M. anisopliae, but not microbiota, increases the encapsulation response. Similarly, the exposure of workers to this fungus led to increased hemolymph antimicrobial activity. Conversely, the removal of bacterial microbiota did not have a significant impact on the survival of workers with M. anisopliae. Our results suggest that the bacterial microbiota associated with the cuticle of A. cephalotes workers does not play a role as a modulator of innate immunity, either at baseline or after exposure to the entomopathogen M. anisopliae. Further, upon infection, workers rely on mechanisms of humoral immunity to respond to this threat. Overall, our findings indicate that the bacterial microbiota associated with A. cephalotes workers does not play a defensive role.

A simple, rapid method for the extraction of whole fire ant venom (Insecta: Formicidae: Solenopsis).

The invasive fire ant Solenopsis invicta is medically important because its venom is highly potent. However, almost nothing is known about fire ant venom proteins because obtaining even milligram-amounts of these proteins has been prohibitively challenging. We present a simple and fast method of obtaining whole venom compounds from large quantities of fire ants. For this, we separate the ants are from the nest soil, immerse them in dual-phase mixture of apolar organic solvent and water, and evaporate each solvent phase in separate. The remaining extract from the aqueous phase is largely made up of ant venom proteins. We confirmed this by using 2D gel electrophoresis while also demonstrating that our new approach yields the same proteins obtained by other authors using less efficient traditional methods.

Survival of Atta sexdens workers on different food sources.

Leaf-cutting ants belonging to the tribe Attini are major herbivores and important agriculture pests in the neotropics, these ants being thought to feed on the sap which exudes from the plant material which they cut and also on the mycelium of a symbiotic fungus that grows on plant material inside their nests in what is called "the fungus garden". However, we have found that the survival of Atta sexdens worker ants on leaves, on mycelium of the ants' symbiotic fungus, Leucoagaricus gongylophorus, or on plant polysaccharides was the same as that of starved A. sexdens, while, conversely, significantly longer survival was achieved by ants fed on the fungus garden material or on some of the products (especially glucose) of the hydrolysis of plant polysaccharides. We found that the fungus garden contained glucose at a higher concentration than that found in leaves or fungal mycelium, and that this glucose was consumed by the ant to the extent that it was probably responsible for up to 50% of the nutritional needs of the workers. The fungus garden contained polysaccharide degrading enzymes (pectinase, amylase, xylanase and cellulase) in proportions similar to that observed in laboratory cultures of L. gongylophorus. It thus appears that A. sexdens workers obtain a significant part of their nutrients from plant polysaccharide hydrolysis products produced by the action of extracellular enzymes released by L. gongylophorus. In this paper we discuss the symbiotic nutrition strategy of A. sexdens workers and brood and the role played by plant polysaccharides in the nutrition of attine ants.