Phylogeny, character evolution and spatiotemporal diversification of the species-rich and world-wide distributed tribe Rubieae (Rubiaceae)

Abstract

The Rubiaceae tribe Rubieae has a world-wide distribution with up to 1,000 species. These collectively exhibit an enormous ecological and morphological diversity, making Rubieae an excellent group for macro- and microevolutionary studies. Previous molecular phylogenetic analyses used only a limited sampling within the tribe or missed lineages crucial for understanding character evolution in this group. Here, we analyze sequences from two plastid spacer regions as well as morphological and biogeographic data from an extensive and evenly distributed sampling to establish a sound phylogenetic framework. This framework serves as a basis for our investigation of the evolution of important morphological characters and the biogeographic history of the Rubieae. The tribe includes three major clades, the Kelloggiinae Clade (Kelloggia), the Rubiinae Clade (Didymaea, Rubia) and the most species-rich Galiinae Clade (Asperula, Callipeltis, Crucianella, Cruciata, Galium, Mericarpaea, Phuopsis, Sherardia, Valantia). Within the Galiinae Clade, the largest genera Galium and Asperula are para- and polyphyletic, respectively. Smaller clades, however, usually correspond to currently recognized taxa (small genera or sections within genera), which may be used as starting points for a refined classification in this clade. Life-form (perennial versus annual), flower shape (long versus short corolla tube) and fruit characters (dry versus fleshy, with or without uncinate hairs) are highly homoplasious and have changed multiple times independently. Inference on the evolution of leaf whorls, a characteristic feature of the tribe, is sensitive to model choice. Multi-parted leaf whorls appear to have originated from opposite leaves with two small interpetiolar stipules that are subsequently enlarged and increased in number. Early diversification of Rubieae probably started during the Miocene in western Eurasia. Disjunctions between the Old and the New World possibly are due to connections via a North Atlantic land bridge. Diversification of the Galiineae Clade started later in the Miocene, probably in the Mediterranean, from where lineages reached, often multiple times, Africa, eastern Asia and further on the Americas and Australia.

Fig 1. Phylogenetic relationships of Rubieae inferred from Bayesian analysis with a relaxed clock of molecular data from the reduced data set.

Shown is the 50% majority rule consensus tree (for details of the Galium Clade see Fig 2); values above branches are posterior probabilities of at least 0.5 from a Bayesian analysis with molecular data only and values below branches are posterior probabilities of at least 0.5 from a Bayesian analysis with combined data (molecular data, trait data, biogeographical data). Roman numerals indicate clade designations used by Ehrendorfer and Barfuss [12]. Abbreviations of major clades: A, Asperula Clade; AC, Asperula-Cruciata Clade; AS, Asperula-Sherardia Clade; C, Cymogalia Clade; G, Galiinae Clade; Ga, Galium Clade; K, Kelloggiinae Clade; R, Rubiinae Clade.

Fig 2. Phylogenetic relationships of Rubieae inferred from Bayesian analysis with a relaxed clock of molecular data from the reduced data set:

The Galium Clade. Shown is the 50% majority rule consensus tree; values above branches are posterior probabilities of at least 0.5 from a Bayesian analysis with molecular data only and values below branches are posterior probabilities of at least 0.5 from a Bayesian analysis with combined data (molecular data, trait data, biogeographical data). The Roman numeral indicates clade designation used by Ehrendorfer and Barfuss [12]. Abbreviations of the major clade: Ga, Galium Clade.

Fig 3. Evolution of life form (perennial/annual) in Rubieae.

Ancestral character states reconstructed over a set of posterior trees using maximum parsimony (left pie charts) and Bayesian inference (right pie charts) are shown on a simplified majority rule consensus tree; therefore, terminal pie charts often show reconstructed character states at the crown node of small terminal clades (indicated by names in normal print) and only rarely the character states of a single accession (indicated by names in italics). Abbreviations of major clades: A, Asperula Clade; AC, Asperula-Cruciata Clade; AS, Asperula-Sherardia Clade; C, Cymogalia Clade; G, Galiinae Clade; Ga, Galium Clade; K, Kelloggiinae Clade; R, Rubiinae Clade.

Fig 4. Evolution of stipules/stipular elements with respect to their number (0 to >2) and size (relative to the true leaves) in Rubieae.

Ancestral character states reconstructed over a set of posterior trees using maximum parsimony (left pie charts) and Bayesian inference (right pie charts) are shown on a simplified majority rule consensus tree; therefore, terminal pie charts often show reconstructed character states at the crown node of small terminal clades (indicated by names in normal print) and only rarely the character states of a single accession (indicated by names in italics). Smaller pie charts indicate reconstructions under a model of ordered character state evolution; these are shown only in cases where reconstructions deviate considerably from those using unordered character states (see text for details). Abbreviations of major clades: A, Asperula Clade; AC, Asperula-Cruciata Clade; AS, Asperula-Sherardia Clade; C, Cymogalia Clade; G, Galiinae Clade; Ga, Galium Clade; K, Kelloggiinae Clade; R, Rubiinae Clade.

Fig 5. Evolution of corolla tube in Rubieae.

Ancestral character states reconstructed over a set of posterior trees using maximum parsimony (left pie charts) and Bayesian inference (right pie charts) are shown on a simplified majority rule consensus tree; therefore, terminal pie charts often show reconstructed character states at the crown node of small terminal clades (indicated by names in normal print) and only rarely the character states of a single accession (indicated by names in italics). Abbreviations of major clades: A, Asperula Clade; AC, Asperula-Cruciata Clade; AS, Asperula-Sherardia Clade; C, Cymogalia Clade; G, Galiinae Clade; Ga, Galium Clade; K, Kelloggiinae Clade; R, Rubiinae Clade.

Fig 6

Evolution of (A) pollen type and (B) fruit type in Rubieae. Ancestral character states reconstructed over a set of posterior trees using maximum parsimony (left pie charts) and Bayesian inference (right pie charts) are shown on strongly simplified majority rule consensus trees; note that, therefore, terminal pie charts often show reconstructed character states at the crown node of terminal clades (indicated by names in normal font) and only rarely the character states of a single accession (indicated by names in italics).

Fig 7. Evolution of fruit indumentum in Rubieae.

Ancestral character states reconstructed over a set of posterior trees using maximum parsimony (left pie charts) and Bayesian inference (right pie charts) are shown on a simplified majority rule consensus tree; therefore, terminal pie charts often show reconstructed character states at the crown node of small terminal clades (indicated by names in normal print) and only rarely the character states of a single accession (indicated by names in italics). Abbreviations of major clades: A, Asperula Clade; AC, Asperula-Cruciata Clade; AS, Asperula-Sherardia Clade; C, Cymogalia Clade; G, Galiinae Clade; Ga, Galium Clade; K, Kelloggiinae Clade; R, Rubiinae Clade.

Fig 8. Spatiotemporal evolution of Rubieae.

Ancestral areas reconstructed via Bayesian analysis of combined data (sequence data, trait data, biogeographical data) shown on a simplified maximum clade credibility tree (node heights are median ages); reconstructions are shown for all collapsed clades and for nodes that have posterior probability of at least 0.7. Collapsed clades (the same as used in Figs 3–7) are shown as triangles, whose vertical extension is proportional to sample size; the color of the triangles (colors corresponding to those used in the ancestral areas pie charts) indicates the proportion of geographic areas in the distribution of the included taxa. Scale bar is in million years, the duration of geological epochs is indicated. Abbreviations of major clades: A, Asperula Clade; AC, Asperula-Cruciata Clade; AS, Asperula-Sherardia Clade; C, Cymogalia Clade; G, Galiinae Clade; Ga, Galium Clade; K, Kelloggiinae Clade; R, Rubiinae Clade.