RESUMEN
Leaf-cutting ants of the genera Atta and Acromyrmex determine serious agricultural problems and live on symbiosis with Leucoagaricus gongylophorus. The aim of this study is to identify morphological and molecularly, as well as to verify the genotypic variability of the symbiotic fungus cultivated by A. heyeri and A. ambiguus from three different regions of the state of Rio Grande do Sul, Brazil. Fungus gardens were collected and fragments of mycelia were grown in selective medium. Total DNA was extracted and amplification of the ITS region was performed by PCR using universal primers. After DNA sequencing, the chromatograms were assembled and phylogenetic analyzes were performed by the Neighbor-Joining method. A total of six isolates of L. gongylophorus were obtained and their identities were confirmed by molecular analyses. Phylogenetic analysis of the ITS region showed a tree with two distinct groups regarding the fungus isolates from A. heiyeri and A. ambiguous. In this study, it was verified that A. heyeri and A. ambiguous, cultivate the same fungus. Additionally, the molecular marker used in this study showed variations in L. gongylophorus, evidencing two distinct branches in the phylogenetic tree, according to the ant species that cultivate L. gongylophorus. However, other studies involving the inclusion of a great number of isolates of L. gongylophorus, as well as the use of other molecular markers to validate the possible variations in the phylogenetic relationship of this symbiotic fungus are required.
Formigas-cortadeiras dos gêneros Atta e Acromyrmex causam elevados prejuízos à agricultura e são dependentes obrigatórias da simbiose com Leucoagaricus gongylophorus. O objetivo deste estudo foi identificar morfologicamente e molecularmente, bem como verificar a variabilidade genotípica do fungo simbionte,cultivado por Acromyrmex heyeri e Acromyrmex ambiguus em três regiões do RS, Brasil. Jardins de fungo foram coletados do interior de formigueiros e fragmentos de micélio foram cultivados em meio de cultura seletivo. O DNA total foi extraído e a amplificação da região ITS foi realizada por PCR, utilizando primers universais. Após sequenciamento, os cromatogramas foram montados e as análises filogenéticas foram realizadas pelo método de Neighbor-Joining. Dos jardins de fungo, obtiveram-se seis isolados de L. gongylophorus, confirmados por análise molecular. A análise filogenética da região ITS mostrou uma árvore com dois grupos distintosem relação aos isolados do fungo oriundos de ninhos de A. heyeri e A. ambiguus. Neste estudo, evidenciou-se que as espécies de formigas A. heyeri e A. ambiguus cultivam o mesmo fungo. Entretanto, o marcador molecular estudado evidenciou variações de L. gongylophorus que permitiram formar duas ramificações diferentes na árvore filogenética relacionada à espécie de formiga que o cultiva. Todavia, para validar as possíveis variações nas relações filogenéticas deste fungo simbionte, é necessária a inclusão de um maior número de isolados de L. gongylophorus, bem como o emprego de outros marcadores moleculares.
RESUMEN
O objetivo deste estudo foi avaliar o desenvolvimento in vitro de Leucoagaricus gongylophorus, fungo simbionte de Atta sexdens rubropilosa, em meio de cultura acrescido com extratos de 24 espécies vegetais. O fungo foi inoculado no meio BDA, acrescido com os extratos vegetais, além de quatro controles, em placas de Petri. As placas foram mantidas em câmaras climatizadas à temperatura de 25±1°C no escuro, por um período de 42 dias. O crescimento do fungo foi avaliado semanalmente através do diâmetro da colônia. Os resultados mostraram que o fungo simbionte dessas formigas apresenta um crescimento diferenciado em relação aos extratos fornecidos nas placas, sendo que o meio acrescido com os extratos de arica (Physocalymma scaberrimum), cerejeira (Amburana acreana), cedro (Cedrela fissilis), timbó (Magonia pubescens), leucena (Leucaena leucocephala) e mogno (Swietenia macrophylla) prejudicaram o desenvolvimento. No entanto, os extratos de genipapo (Genipa americana), seringueira (Heveaa brasiliensis), ingá (Inga edulis) e cambará (Vochysia divergens) apresentaram melhores condições de desenvolvimento para o fungo simbionte.
The objective of this study was to evaluate the in vitro development of Leucoagaricus gongylophorus, the symbiotic fungus of Atta sexdens rubropilosa in culture medium added with extracts of 24 plant species. The fungus was inoculated in PDA added with vegetables extracts and four controls, in Petri dishes. The dishes were maintained in climate chambers at temperature of 25±1°C in the dark, to period of 42 days. The fungal growth was evaluated weekly by diameter of the colony. The results showed that the symbiotic fungus of these ants has differentiated growth when compared to the extracts provided in the dishes, and the medium supplemented with extracts of aricá (Physocalymma scaberrimum), cerejeira (Amburana acreana), cedro (Cedrela fissilis), timbó (Magonia pubescens), leucena (Leucaena leucocephala) e mogno (Swietenia macrophylla) hampered the development, however the extracts of genipapo (Genipa americana), seringueira (Hevea brasiliensis), ingá (Inga edulis) e cambará (Vochysia divergens) had better conditions for the development of the symbiotic fungus.
RESUMEN
In vitro culture of the mutualistic fungus of leaf-cutting ants is troublesome due to its low growth rate, which leads to storage problems and contaminants accumulation. This paper aims at comparing the radial growth rate of the mutualistic fungus of Atta sexdens rubropilosa Forel in two different culture media (Pagnocca B and MEA LP). Although total MEA LP radial growth was greater all along the bioassay, no significant difference was detected between growth efficiencies of the two media. Previous evidences of low growth rate for this fungus were confirmed. Since these data cannot point greater efficiency of one culture medium over the other, MEA LP medium is indicated for in vitro studies with this mutualistic fungus due its simpler composition and translucent color, making the analysis easier.