Abstract
Herbivores gain access to nutrients stored in plant biomass largely by harnessing the metabolic activities of microbes. Leaf-cutter ants of the genus Atta are a hallmark example; these dominant neotropical herbivores cultivate symbiotic fungus gardens on large quantities of fresh plant forage. As the external digestive system of the ants, fungus gardens facilitate the production and sustenance of millions of workers. Using metagenomic and metaproteomic techniques, we characterize the bacterial diversity and physiological potential of fungus gardens from two species of Atta. Our analysis of over 1.2 Gbp of community metagenomic sequence and three 16S pyrotag libraries reveals that in addition to harboring the dominant fungal crop, these ecosystems contain abundant populations of Enterobacteriaceae, including the genera Enterobacter, Pantoea, Klebsiella, Citrobacter and Escherichia. We show that these bacterial communities possess genes associated with lignocellulose degradation and diverse biosynthetic pathways, suggesting that they play a role in nutrient cycling by converting the nitrogen-poor forage of the ants into B-vitamins, amino acids and other cellular components. Our metaproteomic analysis confirms that bacterial glycosyl hydrolases and proteins with putative biosynthetic functions are produced in both field-collected and laboratory-reared colonies. These results are consistent with the hypothesis that fungus gardens are specialized fungus–bacteria communities that convert plant material into energy for their ant hosts. Together with recent investigations into the microbial symbionts of vertebrates, our work underscores the importance of microbial communities in the ecology and evolution of herbivorous metazoans.
Similar content being viewed by others
Log in or create a free account to read this content
Gain free access to this article, as well as selected content from this journal and more on nature.com
or
References
Abril AB, Bucher EH . (2002). Evidence that the fungus cultured by leaf-cutting ants does not metabolize cellulose. Ecol Lett 5: 325–328.
Allgaier M, Reddy A, Park JI, Ivanova N, D’Haeseleer P, Lowry S et al. (2009). Targeted discovery of glycoside hydrolases from a switchgrass-adapted compost community. PLoS One 5: e8812.
Angly FE, Willner D, Prieto-Davo A, Edwards RA, Schmieder R, Vega-Thurber R et al. (2009). The GAAS metagenomic tool and its estimations of viral and microbial average genome size in four major biomes. PLoS Comput Biol 5: e1000593.
Bacci M, Ribeiro SB, Casarotto MEF, Pagnocca FC . (1995). Biopolymer-degrading bacteria from nests of the leaf-cutting ant Atta Sexdens Rubropilosa. Braz J Med Biol Res 28: 79–82.
Belt T . (1874). The Naturalist in Nicaragua. E. Bumpus: London.
Brulc JM, Antonopoulos DA, Miller ME, Wilson MK, Yannarell AC, Dinsdale EA et al. (2009). Gene-centric metagenomics of the fiber-adherent bovine rumen microbiome reveals forage specific glycoside hydrolases. Proc Natl Acad Sci USA 106: 1948–1953.
Burnum KE, Callister SJ, Nicora CD, Purvine SO, Hugenholtz P, Warnecke F et al. (2011). Proteome insights into the symbiotic relationship between a captive colony of Nasutitermes corniger and its hindgut microbiome. ISME J 5: 161–164.
Cantarel BL, Coutinho PM, Rancurel C, Bernard T, Lombard V, Henrissat B . (2009). The Carbohydrate-Active EnZymes database (CAZy): an expert resource for glycogenomics. Nucleic Acids Res 37 (Database issue): D233–D238.
Chapela IH, Rehner SA, Schultz TR, Mueller UG . (1994). Evolutionary history of the symbiosis between fungus-growing ants and their fungi. Science 266: 1691–1694.
Costa AN, Vasconcelos HL, Vieira-Neto EHM, Bruna EM . (2009). Do herbivores exert top-down effects in Neotropical savannas? Estimates of biomass consumption by leaf-cutter ants. J Veg Sci 19: 849–854.
Currie CR, Scott JA, Summerbell RC, Malloch D . (1999). Fungus-growing ants use antibiotic-producing bacteria to control garden parasites. Nature 398: 701–704.
Currie CR, Stuart AE . (2001). Weeding and grooming of pathogens in agriculture by ants. Proc R Soc Lond B Biol Sci 268: 1033–1039.
De Fine Licht HH, Schiott M, Mueller UG, Boomsma JJ . (2010). Evolutionary transitions in enzyme activity of ant fungus gardens. Evolution 64: 2055–2069.
Eng J, McCormack A, Yates J . (1994). An approach to correlate tandem mass spectral data of peptides with amino acid sequences in a protein database. J Am Soc Mass Spectrom 5: 976–989.
Fernández-Marín H, Zimmerman JK, Rehner SA, Wcislo WT . (2006). Active use of the metaplural glands by ants in controlling fungal infection. Proc R Soc London B Biol Sci 273: 1689–1695.
Gill SR, Pop M, Deboy RT, Eckburg PB, Turnbaugh PJ, Samuel BS et al. (2006). Metagenomic analysis of the human distal gut microbiome. Science 312: 1355–1359.
Gomes De Siqueira C, Bacci Jr M, Pagnocca FC, Bueno OC, Hebling MJA . (1998). Metabolism of plant polysaccharides by Leucoagaricus gongylophorus, the symbiotic fungus of the leaf-cutting ant Atta sexdens. Appl Environ Microbiol 64: 4820–4822.
Haeder S, Wirth R, Herz H, Spiteller D . (2009). Candicidin-producing Streptomyces support leaf-cutting ants to protect their fungus garden against the pathogenic fungus Escovopsis. Proc Natl Acad Sci USA 106: 4742–4746.
Hölldobler B, Wilson EO . (1990). The Ants. Harvard University Press: Cambridge, MA.
Hölldobler B, Wilson EO . (2008). The Superorganism: The Beauty, Elegance, and Strangeness of Insect Societies. W. W. Norton & Company: New York, NY.
Hölldobler B, Wilson EO . (2010). The Leafcutter Ants: Civilization by Instinct. W. W. Norton & Company: New York, NY.
Kalyuzhnaya MG, Lapidus A, Ivanova N, Copeland AC, McHardy AC, Szeto E et al. (2008). High-resolution metagenomics targets specific functional types in complex microbial communities. Nat Biotechnol 26: 1029–1034.
Kanehisa M, Araki M, Goto S, Hattori M, Hirakawa M, Itoh M et al. (2008). KEGG for linking genomes to life and the environment. Nucleic Acids Res 36 (Database issue): D480–D484.
Kim S, Gupta N, Pevzner PA . (2008). Spectral probabilities and generating functions of tandem mass spectra: a strike against decoy databases. J Proteome Res 7: 3354–3363.
Margulies M, Egholm M, Altman WE, Attiya S, Bader JS, Bemben LA et al. (2005). Genome sequencing in microfabricated high-density picolitre reactors. Nature 437: 376–380.
Markowitz VM, Ivanova NN, Szeto E, Palaniappan K, Chu K, Dalevi D et al. (2008). IMG/M: a data management and analysis system for metagenomes. Nucleic Acids Res 36 (Database issue): D534–D538.
Martin MM, Weber NA . (1969). The cellulose-utilizing capability of the fungus cultured by the attine ant Atta colombica tonsipes. Ann Entomol Soc Am 62: 1386–1387.
Mueller U, Gerardo N, Aanen D, Six D, Schultz T . (2005). The evolution of agriculture in insects. Annu Rev Ecol Evol Syst 36: 563–595.
Pinto-Tomas AA, Anderson MA, Suen G, Stevenson DM, Chu FS, Cleland WW et al. (2009). Symbiotic nitrogen fixation in the fungus gardens of leaf-cutter ants. Science 326: 1120–1123.
Pope PB, Denman SE, Jones M, Tringe SG, Barry K, Malfatti SA et al. (2010). Adaptation to herbivory by the Tammar wallaby includes bacterial and glycoside hydrolase profiles different from other herbivores. Proc Natl Acad Sci USA 107: 14793–14798.
Powell RJ, Stradling DJ . (1986). Factors influencing the growth of Attamyces bromatificus, a symbiont of the attine ants. Trans Br Mycol Soc 87: 205–213.
Ram RJ, Verberkmoes NC, Thelen MP, Tyson GW, Baker BJ, Blake II RC et al. (2005). Community proteomics of a natural microbial biofilm. Science 308: 1915–1920.
Santos AV, Dillon RJ, Dillon VM, Reynolds SE, Samuels RI . (2004). Ocurrence of the antibiotic producing bacterium Burkholderia sp. in colonies of the leaf-cutting ant Atta sexdens rubropilosa. FEMS Microbiol Lett 239: 319–323.
Schiott M, De Fine Licht HH, Lange L, Boomsma JJ . (2008). Towards a molecular understanding of symbiont function: identification of a fungal gene for the degradation of xylan in the fungus gardens of leaf-cutting ants. BMC Microbiol 8: 40.
Schiott M, Rogowska-Wrzesinska A, Roepstorff P, Boomsma JJ . (2010). Leaf-cutting ant fungi produce cell wall degrading pectinase complexes reminiscent of phytopathogenic fungi. BMC Biol 8: 156.
Schultz TR, Brady SG . (2008). Major evolutionary transitions in ant agriculture. Proc Natl Acad Sci USA 105: 5435–5440.
Scott JJ, Budsberg KJ, Suen G, Wixon DL, Balser TC, Currie CR . (2010). Microbial community structure of leaf-cutter ant fungus gardens and refuse dumps. PLoS One 5: e9922.
Semenova TA, Hughes DP, Boomsma JJ, Schiott M . (2011). Evolutionary patterns of proteinase activity in attine ant fungus gardens. BMC Microbiol 11: 15.
Silva A, Bacci Jr M, Pagnocca FC, Bueno OC, Hebling MJ . (2006a). Production of polysaccharidases in different carbon sources by Leucoagaricus gongylophorus Moller (Singer), the symbiotic fungus of the leaf-cutting ant Atta sexdens Linnaeus. Curr Microbiol 53: 68–71.
Silva A, Bacci Jr M, Pagnocca FC, Bueno OC, Hebling MJ . (2006b). Starch metabolism in Leucoagaricus gongylophorus, the symbiotic fungus of leaf-cutting ants. Microbiol Res 161: 299–303.
Suen G, Scott JJ, Aylward FO, Adams SM, Tringe SG, Pinto-Tomas AA et al. (2010). An insect herbivore microbiome with high plant biomass-degrading capacity. PLoS Genet 6: e1001129.
Tatusov RL, Natale DA, Garkavtsev IV, Tatusova TA, Shankavaram UT, Rao BS et al. (2001). The COG database: new developments in phylogenetic classification of proteins from complete genomes. Nucleic Acids Res 29: 22–28.
Verberkmoes NC, Russell AL, Shah M, Godzik A, Rosenquist M, Halfvarson J et al. (2009). Shotgun metaproteomics of the human distal gut microbiota. ISME J 3: 179–189.
Warnecke F, Luginbuhl P, Ivanova N, Ghassemian M, Richardson TH, Stege JT et al. (2007). Metagenomic and functional analysis of hindgut microbiota of a wood-feeding higher termite. Nature 450: 560–565.
Weber NA . (1966). Fungus-growing ants. Science 153: 587–604.
Wilmes P, Wexler M, Bond PL . (2008). Metaproteomics provides functional insight into activated sludge wastewater treatment. PLoS One 3: e1778.
Woyke T, Teeling H, Ivanova NN, Huntemann M, Richter M, Gloeckner FO et al. (2006). Symbiosis insights through metagenomic analysis of a microbial consortium. Nature 443: 950–955.
Acknowledgements
We thank the staff of the Joint Genome Institute, Pacific Northwest National Laboratories, and the Smithsonian Tropical Research Institute for their expertise and support in the collection and processing of all samples, in particular S Malfatti, L Seid, Y Clemons, R Urriola, M Paz and O Arosemena. We thank all members of the Currie lab for their comments on the manuscript. We also thank three anonymous reviewers for their comments on the manuscript. The US Department of Energy Joint Genome Institute effort was supported by the Office of Science of the US Department of Energy under Contract No. DE-AC02-05CH11231. Proteomic work was performed in the Environmental Molecular Sciences Laboratory, a US Department of Energy (DOE) Office of Biological and Environmental Research national scientific user facility on the Pacific Northwest National Laboratory (PNNL) campus. Portions of this research were supported by the US Department of Energy's (DOE) Office of Biological and Environmental Research (OBER) Panomics program. PNNL is a multiprogram national laboratory operated by Battelle for the DOE under Contract DE-AC05-76RL01830. This work is also supported by the National Science Foundation (grants DEB-0747002, MCB-0702025, and MCB-0731822 to CRC) and the DOE Great Lakes Bioenergy Research Center (DOE Office of Science BER DE-FC02-07ER64494).
Author information
Authors and Affiliations
Corresponding author
Additional information
Supplementary Information accompanies the paper on The ISME Journal website
Rights and permissions
About this article
Cite this article
Aylward, F., Burnum, K., Scott, J. et al. Metagenomic and metaproteomic insights into bacterial communities in leaf-cutter ant fungus gardens. ISME J 6, 1688–1701 (2012). https://doi.org/10.1038/ismej.2012.10
Received:
Revised:
Accepted:
Published:
Issue date:
DOI: https://doi.org/10.1038/ismej.2012.10
Keywords
This article is cited by
-
The Hyphosphere of Leaf-Cutting Ant Cultivars Is Enriched with Helper Bacteria
Microbial Ecology (2023)
-
Gut symbiotic bacteria are involved in nitrogen recycling in the tephritid fruit fly Bactrocera dorsalis
BMC Biology (2022)
-
Fungus-growing insects host a distinctive microbiota apparently adapted to the fungiculture environment
Scientific Reports (2020)
-
Symbiotic bacterial communities in rainforest fungus-farming ants: evidence for species and colony specificity
Scientific Reports (2020)
-
Soil metaproteomics as a tool for monitoring functional microbial communities: promises and challenges
Reviews in Environmental Science and Bio/Technology (2020)
