Molecular Phylogenetics and Evolution xxx (2006) xxx–xxx www.elsevier.com/locate/ympev 1055-7903/$ - see front matter  2006 Published by Elsevier Inc. doi:10.1016/j.ympev.2006.11.001 ARTICLE IN PRESS Please cite this article in press as: Moreira, D. et al., Global eukaryote phylogeny: Combined small- and large-subunit ribosomal DNA trees support monophyly of Rhizaria, Retaria and Excavata, Mol. Phylogenet. Evol. (2006), doi:10.1016/j.ympev.2006.11.001 Global eukaryote phylogeny: Combined small- and large-subunit ribosomal DNA trees support monophyly of Rhizaria, Retaria and Excavata David Moreira a,¤ , Sophie von der Heyden b,1 , David Bass b , PuriWcación López-García a , Ema Chao b , Thomas Cavalier-Smith b a Unité d’Ecologie, Systématique et Evolution, UMR CNRS 8079, Université Paris-Sud, Bâtiment 360, 91405 Orsay Cedex, France b Department of Zoology, University of Oxford, South Parks Road, Oxford OX1 3PS, UK Received 26 July 2006; revised 11 October 2006; accepted 2 November 2006 Abstract Resolution of the phylogenetic relationships among the major eukaryotic groups is one of the most important problems in evolution- ary biology that is still only partially solved. This task was initially addressed using a single marker, the small-subunit ribosomal DNA (SSU rDNA), although in recent years it has been shown that it does not contain enough phylogenetic information to robustly resolve global eukaryotic phylogeny. This has prompted the use of multi-gene analyses, especially in the form of long concatenations of numer- ous conserved protein sequences. However, this approach is severely limited by the small number of taxa for which such a large number of protein sequences is available today. We have explored the alternative approach of using only two markers but a large taxonomic sam- pling, by analysing a combination of SSU and large-subunit (LSU) rDNA sequences. This strategy allows also the incorporation of sequences from non-cultivated protists, e.g., Radiozoa ( D radiolaria minus Phaeodarea). We provide the Wrst LSU rRNA sequences for Heliozoa, Apusozoa (both Apusomonadida and Ancyromonadida), Cercozoa and Radiozoa. Our Bayesian and maximum likelihood analyses for 91 eukaryotic combined SSU+LSU sequences yielded much stronger support than hitherto for the supergroup Rhizaria (Cercozoa plus Radiozoa plus Foraminifera) and several well-recognised groups and also for other problematic clades, such as the Retaria (Radiozoa plus Foraminifera) and, with more moderate support, the Excavata. Within opisthokonts, the combined tree strongly conWrms that the Wlose amoebae Nuclearia are sisters to Fungi whereas other Choanozoa are sisters to animals. The position of some bikont taxa, notably Heliozoa and Apusozoa, remains unresolved. However, our combined trees suggest a more deeply diverging position for Ancyromonas, and perhaps also Apusomonas, than for other bikonts, suggesting that apusozoan zooXagellates may be central for understanding the early evolution of this huge eukaryotic group. Multiple protein sequences will be needed fully to resolve basal bikont phylogeny. Nonetheless, our results suggest that combined SSU+LSU rDNA phylogenies can help to resolve several ambiguous regions of the eukaryotic tree and identify key taxa for subsequent multi-gene analyses.  2006 Published by Elsevier Inc. Keywords: SSU rDNA; LSU rDNA; Eukaryotic phylogeny; Combined analysis; Rhizaria; Retaria; Excavata 1. Introduction For nearly two decades, eukaryote phylogeny has been studied using SSU rDNA 2 as preferred, almost exclusive, * Corresponding author. Fax: +33 1 6915 4697. E-mail address: david.moreira@ese.u-psud.fr (D. Moreira). 1 Present address: Department of Botany and Zoology, University of Stellenbosch, Private Bag X1, Matieland 7602, South Africa. 2 Abbreviations used: BP; bootstrap proportion; LSU rDNA; large-sub- unit ribosomal DNA; ML; maximum likelihood; PP; posterior probability; SSU rDNA; small-subunit ribosomal DNA.