The genome of the leaf-cutting ant Acromyrmex echinatior suggests key adaptations to advanced social life and fungus farming.

We present a high-quality (>100× depth) Illumina genome sequence of the leaf-cutting ant Acromyrmex echinatior, a model species for symbiosis and reproductive conflict studies. We compare this genome with three previously sequenced genomes of ants from different subfamilies and focus our analyses on aspects of the genome likely to be associated with known evolutionary changes. The first is the specialized fungal diet of A. echinatior, where we find gene loss in the ant's arginine synthesis pathway, loss of detoxification genes, and expansion of a group of peptidase proteins. One of these is a unique ant-derived contribution to the fecal fluid, which otherwise consists of "garden manuring" fungal enzymes that are unaffected by ant digestion. The second is multiple mating of queens and ejaculate competition, which may be associated with a greatly expanded nardilysin-like peptidase gene family. The third is sex determination, where we could identify only a single homolog of the feminizer gene. As other ants and the honeybee have duplications of this gene, we hypothesize that this may partly explain the frequent production of diploid male larvae in A. echinatior. The fourth is the evolution of eusociality, where we find a highly conserved ant-specific profile of neuropeptide genes that may be related to caste determination. These first analyses of the A. echinatior genome indicate that considerable genetic changes are likely to have accompanied the transition from hunter-gathering to agricultural food production 50 million years ago, and the transition from single to multiple queen mating 10 million years ago.

Insects associated with tropical foliage produced in the coffee growing region of Colombia

We conducted a survey of insects and pest management practices on 34 farms growing ornamental tropical foliage plants in the central coffee region of Colombia over two years. Tropical foliage provided habitat for a diverse range of insects. In total, phytophagous or detritivorous insects from six orders, 40 families and 62 genera were collected. The most common were Hemiptera (29 genera from 16 families), followed by Coleoptera (17 genera from 4 families), Diptera (5 genera from 5 families), Lepidoptera (5 genera from 4 families), Hymenoptera (3 genera from 2 families) and Orthoptera (2 genera from 2 families). The most common phytophagous species were leaf cutting ants (Atta and Acromyrmex spp.), leaf beetles (Chrysomelidae), leafhoppers (Cicadellidae), stinkbugs (Pentatomidae), squash bugs (Coreidae), tree hoppers (Membracidae) and plant hoppers (Fulgoridae). Beneficial insects identified from tropical foliage included predators and parasitoids amongst 5 orders, 12 families and 22 genera. The most abundant were predators among the Coccinellidae, Chrysopidae, Reduviidae, Lycidae and Formicidae but only low numbers of parasitoids (Ichneumonidae, Braconidae and Tachinidae) were collected. A pest management questionnaire given to growers revealed a preponderance of reliance on broad spectrum insecticides with a smaller number of growers (approximately one third) also using some biological control methods. Our survey contributes basic information regarding diversity of Neotropical insects associated with ornamental foliage plants.

Tissue-adapted invasion strategies of the rice blast fungus Magnaporthe oryzae.

Magnaporthe oryzae causes rice blast, the most serious foliar fungal disease of cultivated rice (Oryza sativa). During hemibiotrophic leaf infection, the pathogen simultaneously combines biotrophic and necrotrophic growth. Here, we provide cytological and molecular evidence that, in contrast to leaf tissue infection, the fungus adopts a uniquely biotrophic infection strategy in roots for a prolonged period and spreads without causing a loss of host cell viability. Consistent with a biotrophic lifestyle, intracellularly growing hyphae of M. oryzae are surrounded by a plant-derived membrane. Global, temporal gene expression analysis used to monitor rice responses to progressive root infection revealed a rapid but transient induction of basal defense-related gene transcripts, indicating perception of the pathogen by the rice root. Early defense gene induction was followed by suppression at the onset of intracellular fungal growth, consistent with the biotrophic nature of root invasion. By contrast, during foliar infection, the vast majority of these transcripts continued to accumulate or increased in abundance. Furthermore, induction of necrotrophy-associated genes during early tissue penetration, previously observed in infected leaves, was not seen in roots. Collectively, our results not only report a global characterization of transcriptional root responses to a biotrophic fungal pathogen but also provide initial evidence for tissue-adapted fungal infection strategies.

Variation in virulence in the rice blast fungus Magnaporthe grisea in São Paulo State

Resistant varieties have been the preferred means to control Magnaporthe grisea, the causal organism of the rice blast disease. The objective of this study was to examine the degree of diversity of the pathogen in different rice growing regions of São Paulo State, Brazil. Blast samples collected from rice varieties in three different regions (Tremembé, Mococa and José Bonifácio) were analyzed for race structure employing the Japanese rice differentials. The highest degree of virulence diversity was observed in Tremembé with 22 different races in three different varieties. Furthermore, no resistance gene in the Japanese differentials was effective to all isolates of M. grisea from São Paulo State.

Wood ants use resin to protect themselves against pathogens

Social life is generally associated with an increased exposure to pathogens and parasites, due to factors such as high population density, frequent physical contact and the use of perennial nest sites. However, sociality also permits the evolution of new collective behavioural defences. Wood ants, Formica paralugubris, commonly bring back pieces of solidified coniferous resin to their nest. Many birds and a few mammals also incorporate green plant material into their nests. Collecting plant material rich in volatile compounds might be an efficient way to fight bacteria and fungi. However, no study has demonstrated that this behaviour has a positive effect on survival. Here, we provide the first experimental evidence that animals using plant compounds with antibacterial and antifungal properties survive better when exposed to detrimental micro-organisms. The presence of resin strongly improves the survival of F. paralugubris adults and larvae exposed to the bacteria Pseudomonas fluorescens, and the survival of larvae exposed to the entomopathogenic fungus Metarhizium anisopliae. These results show that wood ants capitalize on the chemical defences which have evolved in plants to collectively protect themselves against pathogens.

Low variation in ribosomal DNA and internal transcribed spacers of the symbiotic fungi of leaf-cutting ants (Attini: Formicidae)

Leaf-cutting ants of the genera Atta and Acromyrmex (tribe Attini) are symbiotic with basidiomycete fungi of the genus Leucoagaricus (tribe Leucocoprineae), which they cultivate on vegetable matter inside their nests. We determined the variation of the 28S, 18S, and 5.8S ribosomal DNA (rDNA) gene loci and the rapidly evolving internal transcribed spacers 1 and 2 (ITS1 and ITS2) of 15 sympatric and allopatric fungi associated with colonies of 11 species of leafcutter ants living up to 2,600 km apart in Brazil. We found that the fungal rDNA and ITS sequences from different species of ants were identical (or nearly identical) to each other, whereas 10 GenBank Leucoagaricus species showed higher ITS variation. Our findings suggest that Atta and Acromyrmex leafcutters living in geographic sites that are very distant from each other cultivate a single fungal species made up of closely related lineages of Leucoagaricus gongylophorus. We discuss the strikingly high similarity in the ITS1 and ITS2 regions of the Atta and Acromyrmex symbiotic L. gongylophorus studied by us, in contrast to the lower similarity displayed by their non-symbiotic counterparts. We suggest that the similarity of our L. gongylophorus isolates is an indication of the recent association of the fungus with these ants, and propose that both the intense lateral transmission of fungal material within leafcutter nests and the selection of more adapted fungal strains are involved in the homogenization of the symbiotic fungal stock.

Metabolism of plant polysaccharides by Leucoagaricus gongylophorus, the symbiotic fungus of the leaf-cutting ant Atta sexdens L.

Atta sexdens L, ante feed on the Fungus they cultivate on cut leaves inside their nests. The fungus, Leucoagaricus gongylophorus, metabolizes plant polysaccharides, such as xylan, starch, pectin, and cellulose, mediating assimilation of these compounds lay the ants, This metabolic integration may be an important part of the ant-fungus symbiosis, and it involves primarily xylan and starch, both of which support rapid fungal growth. Cellulose seems to be less important for symbiont nutrition, since it is poorly degraded and assimilated by the fungus. Pectin is rapidly degraded but slowly assimilated by L. gongylophorus, and its degradation may occur so that the fungus can more easily access other polysaccharides in the leaves.

Activity of Ricinus communis (Euphorbiaceae) and ricinine against the leaf-cutting ant Atta sexdens rubropilosa (Hymenoptera : Formicidae) and the symbiotic fungus Leucoagaricus gongylophorus

The focus of this study was the identification of compounds from plant extracts for use in crop protection. This paper reports on the toxic activity of fractions of leaf extracts of Ricinus communis L (Euphorbiaceae) and isolated active compounds in the leaf-cutting ant Atta sexdens rubropilosa Forel and its symbiotic fungus Leucoagaricus gongylophorus (Singer) Moller. The main compounds responsible for activity against the fungus and ant in leaf extracts of R communis were found to be fatty acids for the former and ricinine for the ants. (C) 2004 Society of Chemical Industry.

Synthetic amides toxic to the leaf-cutting ant Atta sexdens rubropilosa L. and its symbiotic fungus

1 Nine synthetic amides similar to natural N-piperidine-3-(4,5-methylenedioxyphenyl)-2-(E)-propenainide and N-pyrrolidine-3-(4,5-methylenedyoxiphenyl)2-(E)-propenamide were synthesized and identified by their spectroscopic data.2 the toxicity of these synthetic amides to the Atta sexdens rubropilosa workers and the antifungal activity against Leticoagaricus gongylophorus, the symbiotic fungus of the leaf-cutting ants, were determined.3 Workers ants that were fed daily on an artificial diet to which these compounds were added had a higher mortality rate than the controls for N-pyrrolidine-3(3',4'-methylenedioxyphenyl)-2-(E)-propenamide and N-benzyl-3-(3',4'-methylenedioxyphenyl)-2-(E)-propenamide at a concentration of 100 mu g/mL.4 the completely inhibition (100%) of the fungal growth was observed with N-piperldine-3-(3',4'-methylenedioxyphenyl)-2-(E)-propenamide and N,N-diethyl-3-(3',4'-methylenedioxyphenyl)-2-(E)-propenamide at concentrations of 50 and 100 mu g/mL and N-pirrolidine-3-(3',4'-methylenedioxyphenyl)-2-(E)-propenamide at a concentration of 100 mu g/mL.5 the possibility of controlling these insects in the future using synthetic piperamides that can simultaneously target both organisms is discussed.

Isolation and maintenance of symbiotic fungi of ants in the tribe Attini (Hymenoptera: Formicidae)

O isolamento e a manutenção de fungos basidiomicetos simbiontes de formigas da tribo Attini tem sido dificultado pela baixa velocidade de crescimento desses fungos, bem como pela presença de muitos microrganismos que vivem na superfície do material que as formigas mantêm no interior nos ninhos como substrato para o crescimento dos seus fungos simbiontes. No presente trabalho nós descrevemos um método que aumenta em mais de sete vezes a eficiência de isolamento desses fungos, quando comparada àquela obtida por procedimentos tradicionais. Ninhos subterrâneos de formigas atíneas dos gêneros Atta, Acromyrmex, Trachymyrmex e Mycetarotes foram localizados e deles foram coletadas amostras contendo fungos simbiontes e formigas, que foram transportadas para o laboratório, onde as formigas foram capazes de limpar a cultura do fungo e estimular o seu crescimento. em seguida, porções dos micélios foram assepticamente coletadas e transferidas para meio Yeast Nitrogen Base contendo glicose e cloranfenicol. Para facilitar a manutenção dos isolados em culturas de laboratório, diferentes nutrientes foram analisados para a elaboração de um meio de cultivo complexo, que possibilitou aumentar a velocidade de crescimento dos fungos e estocá-los por longos períodos. O método foi aplicado com sucesso para os fungos simbiontes de todos os gêneros de formigas estudados, gerando, assim, um procedimento extremamente útil para a formação e manutenção de uma coleção representativa de diferentes fungos simbiontes de formigas da tribo Attini.

Low variation in ribosomal DNA and internal transcribed spacers of the symbiotic fungi of leaf-cutting ants (Attini: Formicidae)

Leaf-cutting ants of the genera Atta and Acromyrmex (tribe Attini) are symbiotic with basidiomycete fungi of the genus Leucoagaricus (tribe Leucocoprineae), which they cultivate on vegetable matter inside their nests. We determined the variation of the 28S, 18S, and 5.8S ribosomal DNA (rDNA) gene loci and the rapidly evolving internal transcribed spacers 1 and 2 (ITS1 and ITS2) of 15 sympatric and allopatric fungi associated with colonies of 11 species of leafcutter ants living up to 2,600 km apart in Brazil. We found that the fungal rDNA and ITS sequences from different species of ants were identical (or nearly identical) to each other, whereas 10 GenBank Leucoagaricus species showed higher ITS variation. Our findings suggest that Atta and Acromyrmex leafcutters living in geographic sites that are very distant from each other cultivate a single fungal species made up of closely related lineages of Leucoagaricus gongylophorus. We discuss the strikingly high similarity in the ITS1 and ITS2 regions of the Atta and Acromyrmex symbiotic L. gongylophorus studied by us, in contrast to the lower similarity displayed by their non-symbiotic counterparts. We suggest that the similarity of our L. gongylophorus isolates is an indication of the recent association of the fungus with these ants, and propose that both the intense lateral transmission of fungal material within leafcutter nests and the selection of more adapted fungal strains are involved in the homogenization of the symbiotic fungal stock.

Activity of sesame leaf extracts against the symbiotic fungus of Atta sexdens L.

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Evaluation of the adequacy of diflubenzuron and dechlorane in toxic baits for leaf-cutting ants (Hymenoptera : Formicidae) based on formicidal activity

Studies assessing the characteristics of active ingredients (AIs) of toxic baits for leaf-cutting ants are still scarce, although the need for a delayed action of these compounds on adult workers has been well accepted (mortality <= 15% at 24 h and <= 90% at 21 days). Therefore, we determined the insecticidal action of two AIs used in commercial baits, diflubenzuron and dechlorane, over time in workers, and discussed the control of colonies in relation to the existing literature. Dechlorane presented excellent insecticidal activity with a delayed action at all concentrations tested, although its commercial use has been prohibited due to its organochlorine nature. In contrast, diflubenzuron did not cause significant mortality or symptoms of intoxication, indicating that the successful cases reported with the use of baits containing this AI were in fact due to an accidental contamination with dechlorane. We comment about the ineffectiveness of diflubenzuron on alternative targets, i.e., young forms and the mutualistic fungus, supporting the concept that the AI needs to a have a delayed action on adult workers.

Larval isolation and brood care in Acromyrmex leaf-cutting ants

The larvae of leaf-cutting ants are maintained within the fungus gardens of their colonies and are fed pieces of fungus by the adult workers. However, little else is known about the nature of the worker-larva interaction in these ecologically important ants. To examine whether workers can gauge the needs of individual larvae, we isolated larvae without adult workers for different lengths of time. We then placed workers with the larvae and recorded the type and frequency of the subsequent behaviours of the workers. Workers scraped the mouthparts of larvae, ingested their faecal fluid, fed them with fungal hyphae, transported them around the fungus garden and, most frequently, licked their bodies. The workers were also observed to 'plant' fungal hyphae on the bodies of larvae. Workers interacted more frequently with larvae that had been isolated without workers than with those that had not, but there was no effect of the length of isolation. The results suggest that the interactions are complex, involving a number of behaviours that probably serve different functions, and that workers are to some extent able to assess the individual needs of larvae.

Filamentous fungi found on foundress queens of leaf-cutting ants (Hymenoptera: Formicidae)

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)