Blackwell Science, LtdOxford, UKBOJBotanical Journal of the Linnean Society0024-4074The Linnean Society of London, 2003? 2003 1422 125186 Original Article PHYLOGENETICS OF MAGNOLIALES H. SAUQUET ET AL Botanical Journal of the Linnean Society, 2003, 142, 125–186. With 14 figures Phylogenetic analysis of Magnoliales and Myristicaceae based on multiple data sets: implications for character evolution HERVÉ SAUQUET1*, JAMES A. DOYLE2, TANYA SCHARASCHKIN2, THOMAS BORSCH3, KHIDIR W. HILU4, LARS W. CHATROU5 and ANNICK LE THOMAS1 1Laboratoire de Biologie et Évolution des Plantes vasculaires EPHE, Muséum national d’Histoire naturelle, 16, rue Buffon, 75005 Paris, France 2Section of Evolution and Ecology, University of California, Davis, CA 95616, USA 3Abteilung Systematik und Biodiversität, Botanisches Institut und Botanischer Garten, Friedrich- Wilhelms-Universität Bonn, Meckenheimer Allee 170, 53115 Bonn, Germany 4Department of Biology, Virginia Tech, Blacksburg, VA 24061, USA 5National Herbarium of the Netherlands, Utrecht University branch, Heidelberglaan 2, 3584 CS Utrecht, The Netherlands Received September 2002; accepted for publication January 2003 Magnoliales, consisting of six families of tropical to warm-temperate woody angiosperms, were long considered the most archaic order of flowering plants, but molecular analyses nest them among other eumagnoliids. Based on sep- arate and combined analyses of a morphological matrix (115 characters) and multiple molecular data sets (seven variable chloroplast loci and five more conserved genes; 14 536 aligned nucleotides), phylogenetic relationships were investigated simultaneously within Magnoliales and Myristicaceae, using Laurales, Winterales, and Piperales as outgroups. Despite apparent conflicts among data sets, parsimony and maximum likelihood analyses of combined data converged towards a fully resolved and well-supported topology, consistent with higher-level molecular analyses except for the position of Magnoliaceae: Myristicaceae + (Magnoliaceae + ((Degeneria + Galbulimima) + (Eupomatia + Annonaceae))). Based on these results, we discuss morphological evolution in Magnoliales and show that several supposedly plesiomorphic traits are synapomorphies of Magnoliineae, the sister group of Myristicaceae (e.g. laminar stamens). Relationships within Annonaceae are also resolved with strong support (Anaxagorea basal, then ambav- ioids). In contrast, resolution of relationships within Myristicaceae is difficult and still incomplete, due to a very low level of molecular divergence within the family and a long stem lineage. However, our data provide good evidence that Mauloutchia is nested among other Afro-Malagasy genera, contradicting the view that its androecium and pol- len are plesiomorphic © 2003 The Linnean Society of London, Botanical Journal of the Linnean Society, 2003, 142, 125–186. ADDITIONAL KEYWORDS: Annonaceae – data combination – Magnoliaceae – matK – morphological evolution – ndhF – trnK intron – trnL intron – trnL-trnF – trnT-trnL. INTRODUCTION primitiveness were listed first in most major pre-cla- distic systems of angiosperm classification (including Magnoliales were long thought to be ‘the most archaic Cronquist, 1968, 1981, 1988; Thorne, 1974, 1992; existing order of flowering plants’, as stated by Cron- Takhtajan, 1980, 1997). This interpretation was based quist (1981, 1988), and because of their presumed on a special concentration of characters assumed to be ancestral in angiosperms, including simple, entire leaves with pinnate venation and monosulcate pollen with granular exine structure. Additional presumed *Corresponding author. E-mail:[email protected] plesiomorphic traits included laminar stamens, cond- © 2003 The Linnean Society of London, Botanical Journal of the Linnean Society, 2003, 142, 125–186 125 126 H. SAUQUET ET AL. uplicate carpels, the spiral arrangement of fertile as a paraphyletic basal grade or the sister-group of all parts in Magnoliaceae, and the absence of vessels in other angiosperms, all higher-level phylogenetic anal- the wood of Winteraceae. Although the concept that yses have contradicted a basal position for Magno- Magnoliales are the most primitive angiosperms has liales. Other analyses of morphological data placed been refuted by recent higher-level phylogenetic anal- different taxa at the base of the angiosperms: Caly- yses (as detailed below), they remain one of the most canthaceae (sensu APG, 1998; Renner, 1999) (Loconte important lineages in the early radiation of & Stevenson, 1991), Chloranthaceae (Taylor & Hickey, angiosperms, with a fossil record extending back to 1992; Nixon et al., 1994), Nymphaeales plus monocots the Cenomanian (Dilcher & Crane, 1984; Frumin & (Doyle, Donoghue & Zimmer, 1994), or Nymphaeales Friis, 1999) and possibly the Albian (Crane, Friis & (Doyle, 1996). These variations reflect different out- Pedersen, 1994: stamen type 1) and Aptian (Lethom- group relationships and difficulties in assessment of asites: Ward, Doyle & Hotton, 1989; Doyle & Hotton, morphological homologies between angiosperms and 1991). other seed plants. However, except for studies of rbcL (Chase et al., 1993; Qiu et al., 1993), which placed Ceratophyllum at the base of the angiosperms, analy- CIRCUMSCRIPTION OF MAGNOLIALES ses of molecular data have converged on trees in which The taxa assigned to Magnoliales have varied consid- Amborella, Nymphaeales, Illiciales, Trimeniaceae and erably in recent classification systems (Table 1), Austrobaileya (the ANITA taxa) form a basal, para- although the order has always included a central phyletic grade (Hamby & Zimmer, 1992; Doyle et al., group of three families: Degeneriaceae, Himantan- 1994; Goremykin et al., 1996; D. Soltis et al., 1997; draceae and Magnoliaceae. However, recent progress 2000; P. Soltis et al., 1999; 2000; Mathews & Dono- in angiosperm phylogenetics (Chase et al., 1993; Qiu ghue, 1999, 2000; Parkinson, Adams & Palmer, 1999; et al., 1993, 1999, 2000; Mathews & Donoghue, 1999, Qiu et al., 1999, 2000, 2001; Barkman et al., 2000; 2000; Soltis, Soltis & Chase, 1999; Barkman et al., Borsch, 2000; Graham & Olmstead, 2000; Graham 2000; Doyle & Endress, 2000; Savolainen et al., 2000; et al., 2000; Savolainen et al., 2000; Zanis et al., 2002; D. Soltis et al., 2000; P. Soltis et al., 2000; Zanis et al., Borsch et al. in press; K. W. Hilu et al., unpubl. data). 2002) has led to a new, stable circumscription of the All these analyses place Magnoliales in a more derived order as a monophyletic group of six families (Table 2), (nested) position in the angiosperms, although they as formalized by APG (1998), including the three fam- are still located in the magnoliid grade that is basal to ilies listed above and Myristicaceae, Annonaceae and eudicots and monocots. Eupomatiaceae. It is this definition that we adopt in Analyses of rbcL (Chase et al., 1993; Qiu et al., the present study. 1993), rbcL and atpB (Savolainen et al., 2000), rbcL, Annonaceae, Magnoliaceae and Myristicaceae each atpB and 18S rDNA (P. Soltis et al., 2000), morphology include many species and have pantropical or Asian- (Doyle & Endress, 2000), and morphology plus rbcL, American distributions, whereas Degeneriaceae, atpB and 18S rDNA sequences (Doyle & Endress, Eupomatiaceae and Himantandraceae each consist of 2000) have identified Winteraceae and Canellaceae as a single genus with two species almost entirely the sister-group of Magnoliales. The second outgroup restricted to the Australasian area (Table 2). Unlike of Magnoliales was Laurales (sensu APG, 1998; Annonaceae and Magnoliaceae, none of the genera of Renner, 1999) (Doyle & Endress, 2000; P. Soltis et al., Myristicaceae have widely disjunct distributions. All 2000), Laurales plus Piperales (sensu APG, 1998) genera of Myristicaceae are endemic to one of the four (Savolainen et al., 2000; P. Soltis et al., 2000), or Chlo- following areas: South and Central America (six gen- ranthaceae plus the ANITA taxa (Chase et al., 1993; era), continental Africa (five genera), Madagascar Qiu et al., 1993). The clade consisting of Winteraceae (four genera), and South-east Asia to the western and Canellaceae has been supported by all molecular Pacific (six genera). Most Magnoliales are trees, or analyses, but it was not formally named by APG occasionally shrubs (with some lianas in the Annon- (1998). Chase et al. (1993) treated it as part of Mag- aceae), and usually bear conspicuous, bisexual flow- noliales, but because its position is still unresolved ers, with the notable exception of Myristicaceae, which (see below), we believe it is better considered a distinct are instead characterized by very small, always uni- order, for which several recent papers have used the sexual flowers, with a single perianth cycle of usually name Winterales (Donoghue, 1994; Qiu et al., 1999, three tepals. 2000; P. Soltis et al., 1999, 2000; D. Soltis et al., 2000; Barkman et al., 2000; Doyle & Endress, 2000; Kuzoff & Gasser, 2000). RELATIONSHIPS OF MAGNOLIALES A larger number of molecular analyses have identi- Except for the morphological cladistic analysis of fied Laurales as the sister group of Magnoliales and a Donoghue & Doyle (1989), which placed Magnoliales clade consisting of Winterales and Piperales as their © 2003 The Linnean Society of London, Botanical Journal of the Linnean Society, 2003, 142, 125–186 © 2003TheLinneanSocietyof London, Table 1. Evolution of the family content of Magnoliales through various classification systems prior to APG (1998). The six families underlined are those included in