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Applied and Environmental Microbiology, September 2005, p. 5544-5550, Vol. 71, No. 9
0099-2240/05/$08.00+0     doi:10.1128/AEM.71.9.5544-5550.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Fungal Community Analysis by Large-Scale Sequencing of Environmental Samples{dagger}

Heath E. O'Brien,1* Jeri Lynn Parrent,1 Jason A. Jackson,1 Jean-Marc Moncalvo,2 and Rytas Vilgalys1

Department of Biology, Duke University, Durham, North Carolina 27708,1 Centre for Biodiversity and Conservation Biology, Royal Ontario Museum, and Department of Botany, University of Toronto, Toronto, Canada2

Received 10 November 2004/ Accepted 15 April 2005

Fungi are an important and diverse component of soil communities, but these communities have proven difficult to study in conventional biotic surveys. We evaluated soil fungal diversity at two sites in a temperate forest using direct isolation of small-subunit and internal transcribed spacer (ITS) rRNA genes by PCR and high-throughput sequencing of cloned fragments. We identified 412 sequence types from 863 fungal ITS sequences, as well as 112 ITS sequences from other eukaryotic microorganisms. Equal proportions of Basidiomycota and Ascomycota sequences were present in both the ITS and small-subunit libraries, while members of other fungal phyla were recovered at much lower frequencies. Many sequences closely matched sequences from mycorrhizal, plant-pathogenic, and saprophytic fungi. Compositional differences were observed among samples from different soil depths, with mycorrhizal species predominating deeper in the soil profile and saprophytic species predominating in the litter layer. Richness was consistently lowest in the deepest soil horizon samples. Comparable levels of fungal richness have been observed following traditional specimen-based collecting and culturing surveys, but only after much more extensive sampling. The high rate at which new sequence types were recovered even after sampling 863 fungal ITS sequences and the dominance of fungi in our libraries relative to other eukaryotes suggest that the abundance and diversity of fungi in forest soils may be much higher than previously hypothesized.

* Corresponding author. Mailing address: Department of Biology, Duke University, Durham NC 27708. Phone: (919) 660-7285. Fax: (919) 660-7293. E-mail: heo3{at}duke.edu.

{dagger} Supplemental material for this article may be found at http://aem.asm.org.

Applied and Environmental Microbiology, September 2005, p. 5544-5550, Vol. 71, No. 9
0099-2240/05/$08.00+0     doi:10.1128/AEM.71.9.5544-5550.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.



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