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Exploring the Potential for Actinobacteria as Defensive Symbionts in Fungus-Growing Termites

  • Host Microbe Interactions
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Abstract

In fungus-growing termites, fungi of the subgenus Pseudoxylaria threaten colony health through substrate competition with the termite fungus (Termitomyces). The potential mechanisms with which termites suppress Pseudoxylaria have remained unknown. Here we explore if Actinobacteria potentially play a role as defensive symbionts against Pseudoxylaria in fungus-growing termites. We sampled for Actinobacteria from 30 fungus-growing termite colonies, spanning the three main termite genera and two geographically distant sites. Our isolations yielded 360 Actinobacteria, from which we selected subsets for morphological (288 isolates, grouped in 44 morphotypes) and for 16S rRNA (35 isolates, spanning the majority of morphotypes) characterisation. Actinobacteria were found throughout all sampled nests and colony parts and, phylogenetically, they are interspersed with Actinobacteria from origins other than fungus-growing termites, indicating lack of specificity. Antibiotic-activity screening of 288 isolates against the fungal cultivar and competitor revealed that most of the Actinobacteria-produced molecules with antifungal activity. A more detailed bioassay on 53 isolates, to test the specificity of antibiotics, showed that many Actinobacteria inhibit both Pseudoxylaria and Termitomyces, and that the cultivar fungus generally is more susceptible to inhibition than the competitor. This suggests that either defensive symbionts are not present in the system or that they, if present, represent a subset of the community isolated. If so, the antibiotics must be used in a targeted fashion, being applied to specific areas by the termites. We describe the first discovery of an assembly of antibiotic-producing Actinobacteria occurring in fungus-growing termite nests. However, due to the diversity found, and the lack of both phylogenetic and bioactivity specificity, further work is necessary for a better understanding of the putative role of antibiotic-producing bacteria in the fungus-growing termite mutualistic system.

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Acknowledgements

We thank Thomas W. Kuyper for valuable comments on the manuscript. We are grateful to Michael J. Wingfield, Z. Wilhelm de Beer and colleagues at the Forestry and Agricultural Biotechnology Institute (FABI), South Africa, for hosting and welcoming us to use the laboratory facilities at FABI. Thanks go to Jannette D. Mitchell for showing us sampling sites and to the Oerlemans family for allowing us to sample termite mounds on their property. A.A.V. was supported by a fellowship from the C.T. de Wit Graduate School of Production Ecology & Resource Conservation (PE&RC), Wageningen University, the Netherlands; D.K.A. was funded by a Vidi grant by the Dutch Science Foundation (NWO-ALW) and a grant of the C.T. de Wit Graduate School PE&RC; T.N. was funded by a Marie Curie Intra-European Fellowship within the 7th European Community Framework Programme (IEF Project No. 220077), C.R.C. and M.P. were supported by the US DOE Great Lakes Bioenergy Research Center (DOE BER Office of Science DE-FC02-07ER64494) and by a National Science Foundation grant DEB-0747002 awarded to C.R.C, and M.P. was supported by the Carlsberg Foundation.

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Authors and Affiliations

  1. Laboratory of Genetics, Wageningen University, P.O. Box 309, 6700 AH, Wageningen, The Netherlands

    Anna A. Visser, Tânia Nobre & Duur K. Aanen

  2. Department of Bacteriology, University of Wisconsin, 6145 Microbial Sciences Building, 1550 Linden Dr., Madison, WI, 53706, USA

    Cameron R. Currie & Michael Poulsen

  3. DOE Great Lakes Bioenergy Research Center, University of Wisconsin, Madison, WI, 53706, USA

    Cameron R. Currie & Michael Poulsen

  4. Section for Ecology and Evolution, Department of Biology, University of Copenhagen, Universitetsparken 15, Building 12, 2100, Copenhagen East, Denmark

    Michael Poulsen

  5. Droevendaalsesteeg 1, 6708 PB, Wageningen, The Netherlands

    Anna A. Visser

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  1. Anna A. Visser
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Correspondence to Anna A. Visser.

Electronic supplementary material

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Supplementary TableS1

Overview of sampled termite colonies and isolated strains. *Not sampled for Actinobacteria. (DOC 76 kb)

Supplementary Table S2

Overview of Actinobacteria showing isolation details, assigned morphotype (see main text for details), and strain codes used in the bioassays. (DOC 740 kb)

Supplementary Table S3

Complete data of screening bioassay. Effects (in millimetres) of Actinobacteria AV151-AV288 on Pseudoxylaria (P2) and Termitomyces (T1). Strains that were also tested in the detailed bioassay, selected because of their effect in this screening assay on either Pseudoxylaria (P), Termitomyces (T) or both (P and T), are shown in bold. (DOC 355 kb)

Supplementary Table S4

Sequenced Actinobacteria strains from fungus-growing termite included in the estimation of the Neighbour Joining tree: First BLAST-hit of sequenced strains with strain name, ecological and geographical origins (if known). The table also shows the closest match to type strains from an RDP Type strain search. (DOC 128 kb)

Supplementary Table S5

Complete data for the detailed bioassay. Average effects of Actinobacteria on Pseudoxylaria and Termitomyces. Page 1 shows the average effects of Actinobacteria on Pseudoxylaria and Termitomyces in millimetres zone of inhibition and zone of effect. Page 2 gives the effects per individual fungal strain, in addition to the total zone of effect per strain, all in millimetres. (DOC 251 kb)

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Visser, A.A., Nobre, T., Currie, C.R. et al. Exploring the Potential for Actinobacteria as Defensive Symbionts in Fungus-Growing Termites. Microb Ecol 63, 975–985 (2012). https://doi.org/10.1007/s00248-011-9987-4

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