Filamentous fungi from the genus Trichoderma are commonly found in soil. They are considered facultative mycoparasites, and are antagonists of other fungi such as the cultivar of leaf-cutting ants (Leucoagaricus gongylophorus). The aim of the present study was to bioprospect Trichoderma spp. from different soils collected from Gurupi, Tocantins, Brazil, for antagonistic effects against the mutualistic fungus of leaf-cutting ants. To isolate filamentous fungi, samples were collected from six locations. Preliminarily, isolates were identified by morphological analysis as belonging to Trichoderma. Trichoderma spp. had their internal transcribed spacer region (ITS) of ribosomal RNA genes (rRNA) sequenced to confirm species-level taxonomy. L. gongylophorus was isolated from a laboratory ant colony. Antagonistic properties of seven isolates of Trichoderma against L. gongylophorus were measured using paired disks in Petri dishes with potato dextrose agar medium (PDA). All Trichoderma isolates inhibited the growth of L. gongylophorus in Petri dishes. Isolate 2 of Trichoderma spirale group exhibited slow mycelial growth in the Petri dish, and a high rate of inhibition against L. gongylophorus. This isolate is a promising fungus for field tests of biological control methods for leaf-cutting ants.

Resumo. As formigas-cortadeiras, Atta e Acromyrmex, são consideradas as principais pragas no sistema agroflorestal da Região Neotropical. Isso porque cortam material vegetal que servirá de alimento ao fungo simbionte que elas cultivam.  Diversos fungos, que ocorrem em condições naturais no solo, podem ser encontrados associados às formigas-cortadeiras, muitos deles comprovadamente entomopatogênicos. No entanto, esses agentes não têm sido usados em programa de controle biológico dessas formigas. O objetivo deste estudo foi isolar e identificar fungos filamentosos associados às operárias de Atta sexdens e Atta laevigata e testar sua patogenicidade contra operárias da formiga A. sexdens. Para isolamento dos fungos, foi coletado um total de 180 operárias forrageadoras em seis colônias (30 por colônia), sendo 90 de A. sexdens e 90 de A. laevigata. Dentre os fungos isolados das operárias de A. sexdens,foram identificados Metarhizium anisopliae (Metchnikoff) Sorokin, Aspergillus flavus Link, Acremonium sp. 1, Aspergillus sp.1, Colletotrichum sp. e Acremonium sp. 2. Os fungos encontrados em associação com as operárias da A. laevigata foram Mucor sp., Aspergillus flavus,  Fusarium solani (Martius) Saccardo, A. niger van Tieghem. A patogenicidade de três desses fungos (A. flavus, A. niger e M. anisopliae) foi testada em operárias de A. sexdens. Nos testes com A. niger e M. anisopliae, o tempo para causar mortalidade de 50% das operárias foi de cinco dias (TL50 = 5 dias) e, nos dois casos, o TL50 foi significativamente inferior ao tratamento controle. Assim, testes devem prosseguir com esses isolados que apresentam potencial no uso do controle biológico das formigas-cortadeiras.Filamentous Fungi Associated With Atta sexdens (Linnaeus) and Atta laevigata (F. Smith) (Hymenoptera: Formicidae)Abstract. Leaf-cutting ants, Atta and Acromyrmex genera, are considered major pests in a neotropical agroforestry system because they cut plant material that will serve as food for the symbiotic fungus cultivated by them. Several fungi naturally occurring in the soil can be found associated with leaf-cutting ants, many of them are demonstrably entomopathogenic. However, these agents have not been used as biological control of leaf-cutting ants. The aim of this study was to isolate and to identify filamentous fungi associated with forage workers of Atta sexdens (Linnaeus) and Atta laevigata (F. Smith) and to test their pathogenicity against workers from laboratory colonies. To isolate filamentous fungi, it was collected a total of 180 forage workers (30 in each colony) in six field colonies, 90 of A. sexdens and 90 workers of A. laevigata. Six fungi species from A. sexdens were isolated and identified: Metarhizium anisopliae (Metchnikoff) Sorokin, Aspergillus flavus Link, Acremonium sp. 1, Aspergillus sp. 1, Colletotrichum sp. and Acremonium sp. 2. In A. laevigata, it was found four species: Mucor sp., Aspergillus flavus, Fusarium solani (Martius) Saccardo, Aspergillus niger van Tieghem. Three of these fungi were selected for pathogenicity tests against workers of the leaf-cutting ant A. sexdens: A. flavus, A. niger and M. anisopliae. The LT50 (time to cause 50% mortality of workers) in tests with A. niger and M. anisopliae were five days and significantly lower than the control group. Therefore, further tests should proceed with those isolates to demonstrate their potential use in the biological control of leaf-cutting ants.

-The objective of this work was to evaluate if the immunosuppressant Sandimmun Neoral enhances the activity of Metarhizium anisopliae against the leaf-cutting ant Atta sexdens. The vulnerability to the pathogen was measured by comparing the mortality rate of worker ants subjected to the following treatments: immunosuppressant+control, immunosuppressant+M. anisopliae, excipient+control, excipient+M. anisopliae, control+M. anisopliae, and control. Worker ants treated with immunosuppressant+M. anisopliae showed the highest mortality rate in comparison with those subjected to all other treatments. The use of the immunosuppressant together with entomopathogenic fungus controlled leaf-cutting ants in laboratory conditions. Index terms: biological control, Formicidae, immunity, immunosuppression. Suscetibilidade de operárias de Atta sexdens tratadas com o imunossupressor Sandimmun Neoral a Metarhizium anisopliaeResumo -O objetivo deste trabalho foi avaliar se o imunossupressor Sandimmun Neoral favorece a atividade de Metarhizium anisopliae contra a formiga cortadeira Atta sexdens. A vulnerabilidade ao patógeno foi medida ao se comparar a mortalidade de operárias expostas aos seguintes tratamentos: imunossupressor+controle, imunossupressor+M. anisopliae, excipiente+controle, excipiente+M. anisopliae, controle+M. anisopliae e controle. Operárias tratadas com imunossupressor+M. anisopliae apresentaram a maior taxa de mortalidade em comparação às submetidas a todos os outros tratamentos. O uso do imunossupressor juntamente com fungo entomopatogênico controlou as formigas cortadeiras em condições de laboratório.Termos para indexação: controle biológico, Formicidae, imunidade, imunossupressão.

The importance of Simarouba versicolor St. Hil. fruit dispersal by the leaf-cutting ant Atta sexdens (L.) was studied in the Cerrado, Tocantins, Brazil. The trees and nests were located between a forest area and a Brachiaria decumbens Stapf pasture. Seeds were collected in October 2015 along foraging trails and on the anthill of an A. sexdens colony. Germination of three groups of seeds was tested: (1) seeds with the tegument removed by the ants; (2) seeds without tegument, cleaned manually, and (3) seeds with tegument. The germination rates for the three treatments were similar; however, it was verified that the seeds cleaned by ants germinated faster. In addition, it was verified that the ants dispersed the seeds by at least 20 meters in the study area. Simarouba versicolor is a plant studied for its insecticidal properties, and this is the first study to our knowledge reporting its dispersal by ants.

Founder females of the leaf-cutting ant species Atta sexdens experience high mortality during the founding and establishment of their colonies. The foundation site is crucial for the success of a new colony. In this study, we isolated and identified actinobacteria from fungus garden chambers of A. sexdens colony growth in soils from (1) forested areas without leafcutter ant nests and (2) open ground areas close to leafcutter ant nests. The inhibitory effect of these isolates on pathogenic fungi and the mutualistic fungus cultivated by leafcutter ants was evaluated. The 16S rRNA gene sequences were employed to identify nine selected actinobacteria species found in the soil: Streptomyces (6), Nocardia (2), and Kitasatospora (1). One Streptomyces and one Kitasatospora isolate inhibited all the tested fungi. Since there is no evidence of actinobacteria cultivation in the workers' cuticle of the Atta genus, our results corroborate the hypothesis that these workers may establish temporary adaptive symbiosis with soil microorganisms that produce antibiotic substances, living in some parts of their nest, or even inside their bodies.