Loss of the flagellum happened only once in the fungal lineage: phylogenetic structure of kingdom Fungi inferred from RNA polymerase II subunit genes

Abstract

Background: At present, there is not a widely accepted consensus view regarding the phylogenetic structure of kingdom Fungi although two major phyla, Ascomycota and Basidiomycota, are clearly delineated. Regarding the lower fungi, Zygomycota and Chytridiomycota, a variety of proposals have been advanced. Microsporidia may or may not be fungi; the Glomales (vesicular-arbuscular mycorrhizal fungi) may or may not constitute a fifth fungal phylum, and the loss of the flagellum may have occurred either once or multiple times during fungal evolution. All of these issues are capable of being resolved by a molecular phylogenetic analysis which achieves strong statistical support for major branches. To date, no fungal phylogeny based upon molecular characters has satisfied this criterion.

Results: Using the translated amino acid sequences of the RPB1 and RPB2 genes, we have inferred a fungal phylogeny that consists largely of well-supported monophyletic phyla. Our major results, each with significant statistical support, are: (1) Microsporidia are sister to kingdom Fungi and are not members of Zygomycota; that is, Microsporidia and fungi originated from a common ancestor. (2) Chytridiomycota, the only fungal phylum having a developmental stage with a flagellum, is paraphyletic and is the basal lineage. (3) Zygomycota is monophyletic based upon sampling of Trichomycetes, Zygomycetes, and Glomales. (4) Zygomycota, Basidiomycota, and Ascomycota form a monophyletic group separate from Chytridiomycota. (5) Basidiomycota and Ascomycota are monophyletic sister groups.

Conclusion: In general, this paper highlights the evolutionary position and significance of the lower fungi (Zygomycota and Chytridiomycota). Our results suggest that loss of the flagellum happened only once during early stages of fungal evolution; consequently, the majority of fungi, unlike plants and animals, are nonflagellated. The phylogeny we infer from gene sequences is the first one that is congruent with the widely accepted morphology-based classification of Fungi. We find that, contrary to what has been published elsewhere, the four morphologically defined phyla (Ascomycota, Basidiomycota, Zygomycota and Chytridiomycota) do not overlap with one another. Microsporidia are not included within kingdom Fungi; rather they are a sister-group to the Fungi. Our study demonstrates the applicability of protein sequences from large, slowly-evolving genes to the derivation of well-resolved and highly supported phylogenies across long evolutionary distances.

Figure 1

Some of fungi used in this study. a-d, Chytridiomycota, Allomyces macrogynus (a), Coelomomyces stegomyiae (b), Monoblepharis sp. (c), Chytriomyces hyalinus (d); e, Rhizopus oryzae (Zygomycota); f, Neolecta vitellina (Ascomycota); g-i, Basidiomycota,Amanita phalloides (g),Hydnum repandum (h),Ustilago maydis (i). Photographs are courtesy of Howard Whisler for b and c, Christopher Skory for e, Raymond Boyer for f, Steven Trudell for g and h, and Joe Ammirati for i.

Figure 2

Phylogenies of fungi based on 18S rDNA sequences (A), RPB1 (B) and RPB2 (C) protein sequences. The phylogenies shown are the consensus trees of Bayesian Inference with maximum likelihood branch lengths evaluated using TREE-PUZZLE 5.2. The dots (•)above branches represent the braches with significant statistical support (>95% posterior probabilities of Bayesian inferences and >70% bootstrap values of parsimony analyses)

Figure 3

The phylogeny based on the combined RPB1 and RPB2 protein sequences. This phylogeny is obtained by Bayesian inference. Bayesian posterior probabilities (% are noted above individual branches and bootstrap values below the branches. The chart on the right side of the phylogeny depicts the %G+C of coding sequences of RPB1 and RPB2 with error bars for each taxon. The unique gains and losses of certain characters are mapped on the phylogeny.

Figure 4

RPB1 and RPB2 primers used in the study. The long bars show the extent of the coding regions of RPB1 and RPB2, while the boxes with letters or numbers in them represent the amino acid motifs that are conserved throughout the eukaryotes. The arrows above show the positions of the primers used in this study, and their amino acid sequences and degenerate oligonucleotide sequences are listed below.