Fungal community analysis by large-scale sequencing of environmental samples - PubMed (original) (raw)

Fungal community analysis by large-scale sequencing of environmental samples

Heath E O'Brien et al. Appl Environ Microbiol. 2005 Sep.

Abstract

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. All sequences were deposited in GenBank, with accession numbers AY 969316 to AY 970290 for the ITS sequences and AY 969135 to AY 969315 for the SSU sequences.

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Figures

FIG. 1.

FIG. 1.

Proportions of SSU sequences belonging to different fungal phyla and other eukaryotic kingdoms. The width of triangles corresponds to the number of sequences in each, and the height corresponds to the maximum amount of sequence divergence within clades (see scale bars).

FIG. 2.

FIG. 2.

Proportional distribution of different taxonomic groups in ITS clone libraries. The x axis indicates the proportion of sequences assigned to each taxonomic group, and bar width corresponds to the proportion of all sequences belonging to each library. A. Proportion of all ITS sequences belonging to each fungal kingdom. B. Proportion of Ascomycota ITS sequences belonging to each subclass. C. Proportion of Basidiomycota ITS sequences belonging to each order.

FIG. 3.

FIG. 3.

Species-effort curves of richness for fungal OTUs. A. Mixed hardwood plot. B. Pine plot. C. Pooled data from both plots.

FIG. 4.

FIG. 4.

Species-effort curves for observed and estimated fungal OTU richness in each plot.

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