Association between Pseudonocardia symbionts and Atta leaf-cutting ants suggested by improved isolation methods

Fungus-growing ants associate with multiple symbiotic microbes, including Actinobacteria for production of antibiotics.The best studied of these bacteria are within the genus Pseudonocardia, which in most fungus-growing ants are conspicuouslyvisible on the external cuticle of workers. However, given that fungus-growing ants in the genus Atta do not carry visibleActinobacteria on their cuticle, it is unclear if this genus engages in the symbiosis with Pseudonocardia. Here we explore whetherimproving culturing techniques can allow for successful isolation of Pseudonocardia from Atta cephalotes leaf-cutting ants. Weobtained Pseudonocardia from 9 of 11 isolation method/colony component combinations from all 5 colonies intensively sampled.The most efficient technique was bead-beating workers in phosphate buffer solution, then plating the suspension on carboxymethylcellulosemedium. Placing these strains in a fungus-growing ant-associated Pseudonocardia phylogeny revealed that while somestrains grouped with clades of Pseudonocardia associated with other genera of fungus-growing ants, a large portion of the isolatesfell into two novel phylogenetic clades previously not identified from this ant-microbe symbiosis. Our findings suggest thatPseudonocardia may be associated with Atta fungus-growing ants, potentially internalized, and that localizing the symbiont andexploring its role is necessary to shed further light on the association. [Int Microbiol 2013; 16(1):17-25

Escovopsis trichodermoides sp. nov., isolated from a nest of the lower attine ant Mycocepurus goeldii

Currently, five species are formally described in Escovopsis, a specialized mycoparasitic genus of fungus gardens of attine ants (Hymenoptera: Formicidae: tribe Attini). Four species were isolated from leaf-cutting ants in Brazil, including Escovopsis moelleri and Escovopsis microspora from nests of Acromyrmex subterraneus molestans, Escovopsis weberi from a nest of Atta sp. and Escovopsis lentecrescens from a nest of Acromyrmex subterraneus subterraneus. The fifth species, Escovopsis aspergilloides was isolated from a nest of the higher attine ant Trachymyrmex ruthae from Trinidad. Here, we describe a new species, Escovopsis trichodermoides isolated from a fungus garden of the lower attine ant Mycocepurus goeldii, which differs from the five other species by highly branched, trichoderma-like conidiophores lacking swollen vesicles, with reduced conidiogenous cells and distinctive conidia morphology. Phylogenetic analyses based on partial tef1 gene sequences support the distinctiveness of this species. A portion of the internal transcribed spacers of the nuclear rDNA was sequenced to serve as a DNA barcode. Future molecular and morphological studies in this group of fungi will certainly unravel the taxonomic diversity of Escovopsis associated with fungus-growing ants.Instituto de Botánica "Dr. Carlos Spegazzini

Escovopsis trichodermoides sp. nov., isolated from a nest of the lower attine ant Mycocepurus goeldii

Currently, five species are formally described in Escovopsis, a specialized mycoparasitic genus of fungus gardens of attine ants (Hymenoptera: Formicidae: tribe Attini). Four species were isolated from leaf-cutting ants in Brazil, including Escovopsis moelleri and Escovopsis microspora from nests of Acromyrmex subterraneus molestans, Escovopsis weberi from a nest of Atta sp. and Escovopsis lentecrescens from a nest of Acromyrmex subterraneus subterraneus. The fifth species, Escovopsis aspergilloides was isolated from a nest of the higher attine ant Trachymyrmex ruthae from Trinidad. Here, we describe a new species, Escovopsis trichodermoides isolated from a fungus garden of the lower attine ant Mycocepurus goeldii, which differs from the five other species by highly branched, trichoderma-like conidiophores lacking swollen vesicles, with reduced conidiogenous cells and distinctive conidia morphology. Phylogenetic analyses based on partial tef1 gene sequences support the distinctiveness of this species. A portion of the internal transcribed spacers of the nuclear rDNA was sequenced to serve as a DNA barcode. Future molecular and morphological studies in this group of fungi will certainly unravel the taxonomic diversity of Escovopsis associated with fungus-growing ants.la fecha de publicacion corresponde a la primera versión on line del manuscrit

Specialized Fungal Parasites and Opportunistic Fungi in Gardens of Attine Ants

Ants in the tribe Attini (Hymenoptera: Formicidae) comprise about 230 described species that share the same characteristic: all coevolved in an ancient mutualism with basidiomycetous fungi cultivated for food. In this paper we focused on fungi other than the mutualistic cultivar and their roles in the attine ant symbiosis. Specialized fungal parasites in the genus Escovopsis negatively impact the fungus gardens. Many fungal parasites may have small impacts on the ants' fungal colony when the colony is balanced, but then may opportunistically shift to having large impacts if the ants' colony becomes unbalanced

Foraging of psilocybe basidiocarps by the leafcutting ant acromyrmex lobicornis in Santa Fé, Argentina

Background It is generally accepted that material collected by leaf-cutting ants of the genus Acromyrmex consists solely of plant matter, which is used in the nest as substrate for a symbiotic fungus providing nutrition to the ants. There is only one previous report of any leaf-cutting ant foraging directly on fungal basidiocarps. Findings Basidiocarps of Psilocybe coprophila growing on cow dung were actively collected by workers of Acromyrmex lobicornis in Santa Fé province, Argentina. During this behaviour the ants displayed typical signals of recognition and continuously recruited other foragers to the task. Basidiocarps of different stages of maturity were being transported into the nest by particular groups of workers, while other workers collected plant material. Conclusions The collection of mature basidiocarps with viable spores by leaf-cutting ants in nature adds substance to theories relating to the origin of fungiculture in these highly specialized social insects. © 2013 Masiulionis et al

Association between Pseudonocardia symbionts and Atta leaf-cutting ants suggested by improved isolation methods

Fungus-growing ants associate with multiple symbiotic microbes, including Actinobacteria for production of antibiotics. The best studied of these bacteria are within the genus Pseudonocardia, which in most fungus-growing ants are conspicuously visible on the external cuticle of workers. However, given that fungus-growing ants in the genus Atta do not carry visible Actinobacteria on their cuticle, it is unclear if this genus engages in the symbiosis with Pseudonocardia. Here we explore whether improving culturing techniques can allow for successful isolation of Pseudonocardia from Atta cephalotes leaf-cutting ants. We obtained Pseudonocardia from 9 of 11 isolation method/colony component combinations from all 5 colonies intensively sampled. The most efficient technique was bead-beating workers in phosphate buffer solution, then plating the suspension on carboxymethylcellulose medium. Placing these strains in a fungus-growing ant-associated Pseudonocardia phylogeny revealed that while some strains grouped with clades of Pseudonocardia associated with other genera of fungus-growing ants, a large portion of the isolates fell into two novel phylogenetic clades previously not identified from this ant-microbe symbiosis. Our findings suggest that Pseudonocardia may be associated with Atta fungus-growing ants, potentially internalized, and that localizing the symbiont and exploring its role is necessary to shed further light on the association

Cultivated fungi and gongylidia observed in gardens of <i>Mycocepurus smithii</i> (A, B, C, D) and some species of higher Attini (E, F, G, H).

<p>(A) Nest chamber and pendant fungus garden of <i>Mycocepurus smithii</i>; (B) gongylidia organized in staphylae in the fungus garden of <i>M. smithii</i> (8x magnification); (C) staphylae in a <i>M. smithii</i> cultivar. Gongylidia in the fungus gardens of <i>M. smithii</i> (D), <i>T. fuscus</i> (E), <i>Ac. disciger</i> (F), <i>A. laevigata</i> (G), and <i>A. sexdens</i> (H). The scale bar in figure C represents 100 µm; scale bars in figures D, E, F, G, and H represent 50 µm.</p

ITS phylogeny of Clade 1 of the fungal tribe Leucocoprineae.

<p>Clade 1 of the Leucocoprineae includes four primary clades of attine cultivars and closely related free-living fungi, here indicated as subclades A–D (sensu Mehdiabadi et al. <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0103800#pone.0103800-Mehdiabadi1" target="_blank">[17]</a>). The clade within subclade B that contains the gongylidia-bearing fungal cultivar of <i>M. smithii</i> from Rio Claro is indicated by dashed lines and is enlarged in the inset. Within the inset, the name of the gongylidia-bearing cultivar is indicated in red. The names of other cultivars of <i>M. smithii</i> in subclades A and B are indicated in red; <i>M. smithii</i> additionally cultivates fungi in Clade 2 (not shown here; see Mehdiabadi et al. <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0103800#pone.0103800-Mehdiabadi1" target="_blank">[17]</a>). The phylogram results from maximum-likelihood analyses of ITS sequence data of 305 fungal taxa. In the inset, numbers above branches are Bayesian posterior probabilities; numbers below branches are maximum-likelihood bootstrap proportions.</p