THE TRANSIENT NATURE OF Trichoderma IN ATTINE ANT COLONIES

Fungus-growing “attine” ants are well-known social insects that farm fungi as food source. Although attine colonies are strictly protected by ant workers, several organisms can gain access into this environment. Fungi in the genus Trichoderma continuously enter colonies along with the material collected by the ants and used to nourish their fungal cultivars. In addition, ant workers also carry these fungi on their bodies. Despite some Trichoderma species have been described from leafcutter ants, their occurrence in non-leafcutter ants was poorly studied. To increase the knowledge on the distribution of Trichoderma in the attines’ environment, 73 isolates of the genus were obtained from the colonies of these insects located in three regions in Brazil. This collection represents the largest sampling of Trichoderma from this biological system. After obtaining single spore cultures, the genomic DNA for all isolates was extracted. Then, three molecular markers were amplified and sequenced: the ITS (Internal Transcribed Spacer) region, partial sequences of the tef1 (elongation factor 1 alpha) and rpb2 (RNA polymerase II) genes. Generated sequences were assembled in contigs and compared with their closest relative sequences deposited in NCBI-GenBank. Phylogenetic analysis with the three markers concatenated in one dataset was estimated with Bayesian Inference. We identified 11 known Trichoderma species belonging to the Brevicompactum, Harzianum, Longibrachiatum, Stromaticum, Trichoderma and Virens clades. These taxa were randomly distributed in several attine ant genera examined. Tricoderma spirale (28.7%, 21 out of 73 isolates), followed by T. virens (16.4%) and T. lentiforme (13.7%) were the most abundant species. The remaing taxa occurred in less then 10% of the isolates. In addition, we recovered six putative new phylogenetic species belonging to the Harzianum, Helicum, Lutea, Stromaticum and Trichoderma clades. Considering the ubiquitous nature of Trichoderma, and that the 11 previously described species were also reported from other environments (like soil, rotten logs, sapwood and leaf-litter), our study supports the hypothesis that these fungi are transient organisms in attine ant colonies. However, the occurrence of those new species in other habitats remains unknown. The discovery of new fungi coupled with the foraging behaviour that allows attines to accumulate biomass in their colonies suggest that a high diversity of microorganisms still awaits discovery in this environment.