Molecular Phylogenetics of the Macaronesian-Endemic Genus

Home , Bystropogon, Echium simplex, Gonospermum, Mentheae, Minthostachys, Pycnanthemum incanum

Molecular Ecology (2005) 14, 1177–1189 doi: 10.1111/j.1365-294X.2005.02487.x

BlackwellMolecular Publishing, Ltd. phylogenetics of the Macaronesian-endemic genus Bystropogon (Lamiaceae): palaeo-islands, ecological shifts and interisland colonizations

JENNIFER L. TRUSTY,*¶ RICHARD G. OLMSTEAD,† ARNOLDO SANTOS-GUERRA,‡ SUSANA SÁ-FONTINHA,§ and JAVIER FRANCISCO-ORTEGA* *Department of Biological Sciences, Florida International University, University Park Campus, Miami, FL 33199, USA †Department of Biology, University of Washington, PO Box 355325, Seattle, WA 98195, USA, ‡Jardín de Aclimatación de La Orotava, Calle Retama Num. 2, Puerto de La Cruz, E-38400, Tenerife, Canary Islands, Spain, §Parque Natural da Madeira/CEM, Caminho do Meio, Bom Sucesso, PT-9050-251, Funchal, Madeira, Portugal, ¶Fairchild Tropical Botanic Garden, 11935 Old Cutler Road, Coral Gables, FL 33156, USA

Abstract A molecular phylogenetic study of Bystropogon L’Hèr. (Lamiaceae) is presented. We per- formed a cladistic analysis of nucleotide sequences of the internal transcribed spacers (ITS), of the nuclear ribosomal DNA, and of the trnL gene and trnL-trnF intergenic spacer of the chloroplast DNA. Bystropogon odoratissimus is the only species endemic to the Canary Islands that occurs in the three palaeo-islands of Tenerife. This species is not part of an early diverging lineage of Bystropogon and we suggest that it has a recent origin. This phylogenetic pattern is followed by most of the species endemic to the palaeo-islands of Tenerife. The two sections currently recognized in Bystropogon form two monophyletic groups. Taxa belonging to the section Bystropogon clade show interisland colonization limited to the Canary Islands with ecological shifts among three ecological zones. Taxa from the section Canariense clade show interisland colonization both within the Canary Islands and between the Canary Islands and Madeira. Speciation events within this clade are mostly limited to the laurel forest. The genus has followed a colonization route from the Canaries towards Madeira. This route has also been followed by at least five other plant genera with species endemic to Macaronesia. Major incongruences were found between the current infrasectional classification and the molecular phylogeny, because the varieties of Bystropogon origanifolius and Bystropogon canariensis do not form two monophyletic groups. The widespread B. origanifolius appears as progenitor of the other species in section Bystropogon with a more restricted distribution. Keywords: adaptive radiation, biogeography, evolution, molecular phylogenetics, oceanic islands, quantum speciation Received 20 October 2004; revision received 6 January 2005; accepted 6 January 2005

plant speciation processes in archipelagos worldwide Introduction (Baldwin et al. 1998; Juan et al. 2000; Emerson 2002; Silvertown The Macaronesian Islands comprise the Atlantic archipel- 2004; Valido et al. 2004). Research into the interisland rela- agos of the Azores, Madeira, Selvagens, Canaries, and Cape tionships of Macaronesian endemics gives insight into Verde. In the last 10 years, many evolutionary biology the relative roles of dispersal and ecological adaptation studies have focused on this region, and these island systems as evolutionary avenues (Francisco-Ortega et al. 1996, 2002; have played an important role in the understanding of Panero et al. 1999). However, none of these studies have focused on interpreting phylogenetic patterns in the frame- Correspondence: Javier Francisco-Ortega, Fax: 1305 3481986; E-mail: work of the geological history of Macaronesia. The island [email protected] of Tenerife (Canary Islands) has a complex geological history

1178 J. L. TRUSTY ET AL.

Fig. 1 Geographical distribution of Bystro- pogon in the Canary and the Madeira Islands. The palaeo-islands of Adeje, Anaga, and Teno are indicated by hatched shading.

and is a good model system to investigate these kind of The Macaronesian-endemic genus Bystropogon L’Hèr. questions. Three of the mountain systems of this island, (Lamiaceae) presents an opportunity to address the role Anaga, Adeje, and Teno, are considered ‘palaeo-islands’ played by the geologically old island areas in the diversi- (Ancochea et al. 1990; Marrero & Francisco-Ortega 2001; fication of island endemics. Bystropogon has seven species Marrero 2004; Fig. 1). These regions are estimated to be and four varieties (La Serna-Ramos 1984; Fig. 1). Two of between 4 (Anaga) and 12 (Adeje) million years (Myr) the species are restricted to the island of Madeira (Bystro- old; Teno is approximately 6 Myr old (Thirlwall et al. pogon maderensis and Bystropogon punctatus), while the rest 2000; Guillou et al. 2004). Volcanic activity during the late of the species are endemic to the Canary Islands. Among Tertiary and early Quaternary led to the merger of these the Canary Islands, only Tenerife and La Palma harbour three palaeo-islands, which eventually formed the current single-island endemics of this genus. Bystropogon odoratis- island of Tenerife approximately 1 million years ago (Ma) simus and Bystropogon plumosus are restricted to Tenerife, (Ancochea et al. 1990). These three regions have a high whereas Bystropogon wildpretii is endemic to La Palma. All number of endemics (Bramwell & Bramwell 1974; Santos- the species of Bystropogon are abundant with the exception Guerra & Fernández-Galván 1983; Martín et al. 1999), and of B. odoratissimus and B. wildpretti. some authors have suggested that they have some of the Bystropogon odoratissimus is restricted to the three palaeo- oldest plant lineages of the archipelago (reviewed by islands of Tenerife. This species has a disjunct distribution Marrero & Francisco-Ortega 2001; Marrero 2004). and is found only in Teno, Anaga, and Adeje. This unique

BYSTROPOGON PHYLOGENY 1179 biogeographical pattern is not known for any other plant an extremely common understory species in the pine forest species of Tenerife and has led some authors to consider belt. Occasionally, this species can be found in the lowland B. odoratissimus to be part of an ancient lineage of the genus scrub and laurel forests areas of Tenerife. (Hernández & García 1996). In this study we present a molecular phylogenetic study Two widespread Bystropogon species, B. canariensis and of the genus Bystropogon based on nucleotide sequences B. origanifolius, are highly polymorphic and form two spe- of the nuclear and chloroplast genomes. The three major cies complexes. Bystropogon origanifolius has three varieties objectives of our study were (i) to determine if B. odora- restricted to Gran Canaria (var. canariae), El Hierro (var. tissimus is part of a early diverging lineage of Bystropogon ferrensis), and La Palma (var. palmensis). Bystropogon origan- and investigate the biogeographical role played by the ifolius var. origanifolius is endemic to the islands of La Gomera three palaeo-islands of Tenerife; (ii) to discuss the main and Tenerife. Bystropogon canariensis has two varieties. The ecological and geographical avenues followed by the genus type variety canariensis occurs on the islands of Gran Canaria, during its evolutionary history in the context of other La Palma, and Tenerife, while B. canariensis var. smithianus Macaronesian-endemic groups; and (iii) to determine if the occurs on four islands, La Gomera, El Hierro, La Palma, molecular phylogeny of the genus is concordant with the and Tenerife. latest taxonomic classification of Bystropogon. The latest taxonomic revision of Bystropogon produced by La Serna-Ramos (1984) recognized two sections: Bystro- Materials and methods pogon section Bystropogon and Bystropogon section Canariense La Serna. Bystropogon canariensis and the two Madeiran Plant materials species, B. maderensis and B. punctatus, were placed in Bystropogon section Canariense. The remaining four species The ingroup consisted of 17 taxa representing all the species comprise Bystropogon section Bystropogon. and varieties of Bystropogon in Macaronesia from the two The three species of Bystropogon section Canariense are sections currently recognized by La Serna-Ramos (1984). primarily restricted to the laurel forest ecological zone When a species was found on multiple islands, a single (Pruno hixae-Lauretea novocanariensis) (La Serna-Ramos 1984; representative from each island was included in the analysis. Capelo et al. 1999; Jardim & Francisco 2000). This vegetation Mentha L., Minthostachys (Benth.) Spach, Pycnanthemum zone is found on the northern slopes of the islands which Michx., and Ziziphora L. were chosen as the outgroups. The are under the direct influence of the humid and cool north- outgroup was selected based on a previous phylogenetic eastern trade winds and receive approximately 1000 mm study of the tribe Mentheae (Trusty et al. 2004). Details of of average rainfall per year (Fernández-Palacios 1999). the plant material, including voucher information, accession Two species of Bystropogon section Bystropogon (i.e. B. provenance, and nucleotide sequence identification, are origanifolius and B. wildpretii) are confined mostly to the pine listed in Table 1. forest ecological zone (Chamaecytiso-Pinetea canariensis) (La Serna-Ramos 1984). This ecological zone only occurs in the DNA extraction, PCR amplification, and sequencing Canary Islands and is situated above the laurel forest (on the northern slopes of the islands) or above the lowland DNA was extracted from either fresh or silica-gel-dried scrub (on the southern slopes). Rainfall in this forest ranges material using the DNeasy protocol (QIAGEN). Both strands between 400 and 800 mm per year (Fernández-Palacios of the two nuclear ribosomal DNA internal transcribed 1999). spacer (ITS) regions including the 5.8S gene were amplified The two other species of this section, B. odoratissimus using primers ITS4 (White et al. 1990) and ITS5 (Downie and B. plumosus, are primarily found in the lowland scrub & Katz-Downie 1996). Polymerase chain reaction (PCR) (Rhamno crenulatae-Oleetea cerasiformis) (La Serna-Ramos amplification conditions are described in Kim & Jansen 1984; Hernández & García 1996). The lowland scrub is (1994). Difficulty in amplifying the ITS region of some taxa situated above the coastal xerophytic belt (Kleinio-Euphorbietea was ameliorated through the use of the Ready-To-Go PCR canariensis) and receives up to 550 m of rainfall per year Bead Kit (Amersham/Pharmacia Biotech). (Fernández-Palacios 1999). The chloroplast trnL gene and trnL-trnF intergenic Bystropogon origanifolius var. origanifolius has the widest spacer (trnL/F) were amplified using the ‘C’ and ‘F’ ecological amplitude of any of Bystropogon species. This primers according to the protocol described by Taberlet taxon is reported in the few natural pine forests of La et al. (1991). Gomera (Fernández-Galván 1983; Del Arco-Aguilar et al. PCR products were cleaned using the QIAquick silica 1990); however, it is much more abundant in the lowland columns (QIAGEN) following the manufacturer’s instruc- scrub vegetation zone and in sunny areas of the laurel for- tions. The purified PCR products were cycle-sequenced est of this island (La Serna-Ramos 1984). B. origanifolius var. in both directions using the ABI Prism BigDye, Terminator origanifolius has a different ecology in Tenerife, where it is Cycle Sequencing Ready Reaction Kit (Perkin-Elmer, Applied

Table 1 List of plant material used in this study

GenBank Accession nos Taxon Island Voucher ITS/trnL-F

Bystropogon canariensis (L.) L′ Hèr. Gran Canaria Los Marteles; Marrero, Caujapé & Francisco-Ortega (LPA) AY04590/AY704605 var. canariensis La Palma Sobre Tagoja; Santos-Guerra 6–1999 (ORT) AY04583/AY704607 Tenerife Pedro Alvarez; Santos-Guerra 7-6-1999 (ORT) AY506634/AY506597 Bystropogon canariensis (L.) L′ Hèr. La Gomera Chorros de Epina; Santos-Guerra 10-3-2000 (ORT) ------/AY704603 var. smithianus Christ El Hierro El Golfo; Santos-Guerra 8-2-2000 (ORT) AY704592/AY704595 La Palma Breña Baja, Santos-Guerra 6-10-2002 (ORT) AY704593/AY704606 Tenerife Miradores de la Cumbre; Santos-Guerra 8-8-1999 (ORT) AY706475/AY704594 Bystropogon maderensis Webb Madeira Ribeiro Frio; Fontinha and Roberto s. n. (MADJ) AY506633/AY506596 Bystropogon odoratissimus C. Bolle Tenerife Punta Hidalgo, Tenerife; Santos-Guerra 6-20-1999 (ORT) AY704589/AY704596 Bystropogon origanifolius L′ Hèr. Tenerife Montaña de Los Poleos; Santos-Guerra 8-6-1999 (ORT) AY506635/AY506598 var. origanifolius La Gomera Vallehermoso; Santos-Guerra 5-3-1997 (ORT) AY704591/AY704604 Bystropogon origanifolius L′ Hèr. Gran Canaria Riscos de Tamadaba; Marrero, Caujapé and AY704587/AY704600 var. canariae I. La-Serna Francisco-Ortega (LPA) Bystropogon origanifolius L′ Hèr. El Hierro Isora; Santos Guerra 8-2-2000 (ORT) AY704585/AY704597 var. ferrensis (Ceb. and Ort.) I. La-Serna Bystropogon origanifolius L′ Hèr. La Palma Roque de los Muchachos; Santos Guerra 7–1999 (ORT) AY704588/AY704599 var. palmensis L′ Hèr. Bystropogon plumosus (L. f.) L′ Hèr. Tenerife Articosa; Santos-Guerra 8-31-2000 (ORT) AY704586/AY704598 Bystropogon punctatus L′ Hèr. Madeira Folhadal; Fontinha and Roberto s. n. (MADJ) AY704582/AY704602 Bystropogon wildpretii I. La-Serna La Palma Tijarafe; Santos-Guerra 6–1999 (ORT) AY704584/AY704601 Outgroup species: Mentha rotundifolia (L.) Huds. Wagstaff 88–026 (BHO) AY506645/AY506610 Minthostachys mollis (Kunth) Tulcan, Ecuador; Thompson and Rawlins 942 (BHO) AY506638/AY506601 Griseb. Pycnanthemum incanum (L.). Bloomington, Indiana; Olmstead 90–06 (WTU) AY506640/AY506604 Michx. Ziziphora hispanica L. Madrid, Spain; Sánchez-Mata and Gavilán 100 (MSC) AF369166/AY506595

Herbaria (Holmgren et al. 1990; www.nybg.org/bsci/ih/ih.html) where vouchers are deposited are indicated in brackets.

Biosystems Division) with AmpliTaq DNA polymerase. The Data analysis sequencing reactions were conducted using the same primers that were used for the PCR amplifications. Sequences were assembled, edited, and aligned using Dye-terminator reactions were carried out in 10-µL reactions, sequencher 4.1 (Genecodes) and clustal x (Thompson diluted 50:50 using AmpliTaq FS buffer (Perkin-Elmer, et al. 1997), respectively. Minor adjustments were made Applied Biosystems Division) and amplified according to manually to the final alignment using se-al version 2a08 manufacturer’s protocol. Cycle sequencing products were (A. Rambaut, University of Oxford, UK). Fewer than 0.1% separated on an ABI 377 automated sequencer at the Florida of data matrix cells were scored as missing. Gaps were International University (FIU) DNA Core Facility. coded as binary characters following the ‘simple indel coding’ procedure of Simmons & Ochoterena (2000). The aligned data matrices are deposited in the treebase MFOLD analysis database (Accession no. SN1952). Thermodynamic stability of the ITS sequences were Heuristic phylogenetic parsimony analyses were per- analysed using mfold version 3.1 (Zuker 2003). The par- formed using paup* (Swofford 2002). Heuristic searches ameters were set with the folding temperature of 37 °C with 1000 random taxon addition replicates (using the TBR and the sodium concentration of 1 m. Comparisons of and MulPars options of paup*) to look for multiple optimal free energy estimates (dG values) among ITS sequences tree islands (Maddison 1991). Phylogenetic support for allowed us to identify putative nonfunctional paralogue each clade was evaluated through bootstrap analysis copies (pseudogenes) which were consequently excluded (Felsenstein 1985) of 1000 replicates with one random from the phylogenetic analyses. sequence addition per replicate and the TBR and MulPars

© 2005 Blackwell Publishing Ltd, Molecular Ecology, 14, 1177–1189 BYSTROPOGON PHYLOGENY 1181 options (DeBry & Olmstead 2000). In the trnL/F bootstrap analysis, the protocol was modified to save only 20 trees per replicate because of computer memory limitations.

Results

ITS phylogenetic analyses The ITS sequence from Bystropogon canariensis var. smithianus from La Gomera had an unusually high number of changes when compared with the other sequences. In addition, alignment of this sequence with the rest of the sequences was difficult to achieve. Uncorrected ‘p’ distances (Swofford 2002) were determined to estimate levels of nucleotide divergence among sequences. Distance p values between outgroup and ingroup varied between 0.06 and 0.09 for sequences with dG values lower than −50. The unusual sequence of B. canariensis var. smithianus from La Gomera had dG values of −35.9 (for the ITS1 region) and −28.1 (for the ITS2 region). The sequence diverged from the two other complete ITS Bystropogon sequences of the same variety by 0.13 and 0.17. Pairwise P values within the ingroup (excluding this sequence from La Gomera) varied between 0.02 and 0.07. This unusual ITS sequence was therefore identified as a putative nonfunctional paralogue and was subsequently excluded from the phylogenetic analysis. We were able to successfully sequence only the ITS2 region for Fig. 2 Nuclear and chloroplast DNA phylogenies of Bystropogon. (a) One of the nine most parsimonious trees from the ITS analysis B. canariensis var. smithianus from La Palma. The dG value − (184 steps; CI = 0.859; RI = 0.865). This tree was identical to the ( 64.7) for this ITS2 sequence indicated that it belonged to strict consensus tree. (b) One of the 28 most-parsimonious trees a functional copy. This ITS2 sequence was included in the from the trnL/F analysis analysis [68 steps; CI = 0.897; RI = 0.829). phylogenetic analyses. Branches which collapse in the strict consensus tree are indicated The final aligned ITS matrix was 648 nucleotide positions by dashed lines. Bootstrap values are below branches. Branch in length and included only one informative coded gap. lengths are above branches. There were 140 variable characters, with 39% (55) of these being parsimony informative. The search yielded nine clade. One was moderately supported with a 72% boot- equally most-parsimonious trees [length = 184 steps; con- strap value and contains Bystropogon wildpretti, Bystropogon sistency index (CI) = 0.859; retention index (RI) = 0.865]. origanifolius var. ferrensis and Bystropogon origanifolius var. One of the nine most-parsimonious trees (equivalent to the palmensis. The second was strongly supported (95%) and strict consensus tree) is shown in Fig. 2. When the phylo- contains the rest of the taxa of the section, including the genetic analysis was conducted using only the ITS2 data, palaeo-island-endemic Bystropogon odoratissimus. 21 trees were found. The strict consensus of the ITS2 tree was consistent with the strict consensus of the complete ITS, trnL/F phylogenetic analyses except for reduced resolution within the two major clades. The ITS strict consensus tree supported a basal split of The final aligned matrix used for parsimony analyses of Bystropogon into two major clades (Fig. 2). One of these trnL/F was of 832 DNA characters in length and included major clades (98% bootstrap value) contained all the five parsimony informative gaps. There were 60 variable members of Bystropogon section Canariense. B. canariensis characters, with 30% (18) of these being parsimony infor- var. smithianus from La Palma was sister to the rest of this mative. Parsimony analysis yielded 28 most-parsimonious clade. This basal split was also found in the ITS2 topology. trees (length = 68 steps each; CI = 0.897; RI = 0.829). One of The rest of the taxa of this section formed a monophyletic these 28 trees and the strict consensus tree are shown in assemblage weakly supported by a bootstrap value of 69%. Fig. 2. The second major clade (99% bootstrap value) contained all The trnL/F strict consensus tree identified three major the members of Bystropogon section Bystropogon. Two major clades. One of these clades (68% bootstrap value) comprised lineages were identified at the base of this second major the outgroup genera Minthostachys and Pycnanthemum.

Fig. 3 One of the 48 most-parsimonious trees from the combined analyses of ITS and trnL/F data (257 steps; CI = 0.852; RI = 0.837). Branches which collapse in the strict consensus tree are indicated by dashed lines. Bootstrap values are below branches. Branch lengths are above branches. Open circles indicate taxa found in the pine forest, closed circles indicate taxa found to the laurel forest, and closed squares indicate taxa found in the lowland scrub.

Another clade consisted of Bystropogon section Canariense; not include B. canariensis var. smithianus from La Gomera this clade had low bootstrap support below 50%. The third because we were unable to obtain ITS sequences for this major clade comprised all of the members of Bystropogon accession. There were 190 variable characters, with 38% (73) section Bystropogon (61% bootstrap value). Two major lin- of these being parsimony informative. Forty-eight equally eages were distinguished in this clade, the first lineage had most-parsimonious trees were obtained (length = 257 steps, only the Gran Canaria endemic B. origanifolius var. canariae. CI = 0.852; RI = 0.837). One of the most-parsimonious trees The remaining six taxa of Bystropogon sect. Bystropogon, and the strict consensus of all trees are shown in Fig. 3. The including B. odoratissimus, formed the second lineage combined ITS – trnL/F strict consensus tree also showed with no resolution among them. the two major clades obtained after the phylogenetic analyses of ITS data. The five Canary Islands where multiple taxa of Bystropogon Combined analyses occur have at least one taxon from each of the two major The final aligned matrix obtained from the combination of clades of the genus. Unfortunately we could not verify this ITS and trnL/F data was 1480 positions in length and had result for the island of La Gomera, because functional ITS six phylogenetically informative gaps. This matrix did sequences of B. canariensis var. smithianus from this island

© 2005 Blackwell Publishing Ltd, Molecular Ecology, 14, 1177–1189 BYSTROPOGON PHYLOGENY 1183 were not available for the combined data set. However, species endemic to Macaronesia. A phylogenetic study based on the chloroplast DNA (cpDNA) phylogeny, this of this genus shows that the relatively common Canarian accession is part of the section Canariense clade. The clade endemic L. acerifolia Cav. belongs to one of the crown groups comprising section Bystropogon does not have any species of the Lavatera–Malva L. complex (Fuertes-Aguilar et al. on the island of Madeira. 2002). The palaeo-island endemic Lavatera phoenicea is sister The clade comprising section Canariense was mostly to the rest of the members of the complex. limited to the laurel forest, and only one ecological shift was The woody Sonchus alliance is the only taxonomic group identified within this clade. In contrast section Bystropogon with more than 20 species endemic to Macaronesia that has shows ecological shifts among three ecological zones: the an early diverging species restricted to one of the palaeo- laurel forest, the lowland scrub, and the pine forest. islands of Tenerife. The ITS phylogeny of this alliance The current species-level classification of B. origanifolius shows that the Teno-endemic S. tuberifer is part of an early and B. canariensis is not supported in the combined analyses. diverging lineage of the woody Sonchus alliance. However The four varieties of B. origanifolius did not form a mono- most of the nodes of this phylogenetic tree are weakly phyletic group. Varieties canariae and origanifolius formed a supported with bootstrap values between 50% and 60% clade with B. odoratissimus and B. plumosus. The other (Kim et al. 1996). varieties of B. origanifolius (i.e. var. ferrensis and var. palmensis) Finally, a molecular phylogenetic study of tribe Inuleae grouped with B. wildpretii. Together, the two varities of shows that Vierea and the North African genus Perralderia Bystropogon canariensis formed a paraphyletic group with Coss. (three species) form an assemblage that is sister to a respect to B. maderensis and B. punctatus. large clade of 10 genera with a predominantly Mediterranean distribution, however, this relationship is weakly supported (Francisco-Ortega et al. 2001b). Discussion There are at least seven plant genera with species restricted to the old island of La Gomera and at least to one of the The phylogenetic position of Bystropogon three palaeo-islands of Tenerife (Sventenius 1948; Santos- odoratissimus — phylogenetic patterns of plant species Guerra and Fernández-Galván 1983; Gómez-Campo 1996; occurring in the three palaeo-islands of Tenerife Hernández & Bañares 1996; Beltrán-Tejera et al. 1999; Del Approximately 55 species are endemic to at least one of the Arco-Aguilar 2000; Varios Autores 2000). La Gomera is three palaeo-islands of Tenerife (Sventenius 1948; Santos- approximately 9 Myr old (Guillou et al. 2004) and together Guerra & Fernández-Galván 1983; Gómez-Campo 1996; with the three palaeo-islands of Tenerife formed a small Hernández & Bañares 1996; Beltrán-Tejera et al. 1999; Del archipelago located in the westernmost Canary Islands until Arco-Aguilar 2000; Varios Autores 2000). This number of the early Quaternary (Marrero & Francisco-Ortega 2001; endemic species represents approximately 10% of the Marrero 2004). It was only then that submarine volcanism flora endemic to the Canary Islands. Thirty-seven of these lead to the formation of the islands of La Palma and El species have been included in molecular phylogenetic Hierro. Three of the species restricted to La Gomera and studies (Table 2), therefore there are phylogenies available the palaeo-islands of Tenerife (i.e. Convolvulus volubilis, for almost 67% of these palaeo-island endemics. Dicheranthus plocamoides, Teline pallida) have been the Although Bystropogon odoratissimus is restricted to the subject of molecular phylogenetic studies (Table 2). Both three palaeo-islands of Tenerife, our phylogenetic study D. plocamoides and T. pallida follow the pattern detected does not support B. odoratissimus as an early diverging for most of the palaeo-island endemics and are part of lineage of Bystropogon. This species is nested inside the terminal tips of their respective phylogenies (Oxelman section Bystropogon clade in a late branching position. The et al. 2002; Percy & Cronk 2002; Smissen et al. 2002). phylogenetic patterns of B. odoratissimus are not unique Convolvulus volubilis has a unique biogeographical pattern, and 84% of the endemic species from the palaeo-islands in that it is the only Canarian species endemic to Adeje, follow similar patterns (Table 2). Anaga, La Gomera, and Teno. This species represents the Species restricted to Anaga/Adeje/Teno from the genera first branching lineage of endemic climbing species of Aeonium Webb and Berthel., Argyranthemum Sch. Bip., Cheir- Convolvulus and therefore is another example of an early olophus Cass., Crambe L., Echium L., Lotus L., Lugoa DC., diverging taxon with a palaeo-island (including La Gomera) Monanthes Haw., Sideritis L., Teline Medicus, and Tolpis distribution. Adanson are part of crown groups in their respective The paucity of ancient, or early diverging groups (16%, phylogenies, although in many cases the relevant nodes Table 2) restricted to the palaeo-islands of Tenerife supports were supported by bootstrap values below 75%. the view that several of the speciation events in the Canary In contrast, species from only three genera, Lavatera L., Islands are relatively recent and that most of the species the woody Sonchus alliance, and Vierea Webb and Berthel. which are endemic to these palaeo-islands are not the are part of early diverging lineages. Lavatera has two result of ancient phylogenetic splits. Indeed, most of these

Table 2 Molecular phylogenetic studies of species endemic in the palaeo-islands of Adeje, Anaga, or Teno

Geographical distribution

Species Adeje Anaga Teno Reference for phylogenetic study

Aeonium ciliatum Webb and Berthel. (Crassulaceae) + Mort et al. (2002) Aeonium haworthii (Webb and Berthel.) Webb and Berthel. (Crassulaceae) + Mort et al. (2002) Aeonium mascaense Bramwell (Crassulaceae) + Mort et al. (2002) Aeonium pseudourbicum Bañares (Crassulaceae) + Mort et al. (2002) Aeonium volkeri H. Hernández and Bañares (Crassulaceae) + Mort et al. (2002) Argyranthemum coronopifolium (Willd.) Humphries (Asteraceae) + + Francisco-Ortega et al. (1996)*† Argyranthemum foeniculaceum (Willd.) Sch. Bip. (Asteraceae) + Francisco-Ortega et al. (1996)*† Argyranthemum lemsii Humphries (Asteraceae) + Francisco-Ortega et al. (1996)* Argyranthemum sundingii L. Borgen (Asteraceae) + Francisco-Ortega et al. (1996)*† Bystropogon odoratissimus Bolle (Lamiaceae) + + + Present study Cheirolophus burchardii Susanna (Asteraceae) + Susanna et al. (1999)*† Cheirolophus canariensis (Willd.) Holub (Asteraceae) + Susanna et al. (1999)*† Cheirolophus tagananensis (Svent.) Holub (Asteraceae) + Susanna et al. (1999)*† Convolvulus volubilis Link (Convolvulaceae) ++ + Carine et al. (2004)*¶ Crambe laevigata Christ (Brassicaceae) + Francisco-Ortega et al. (2002)*† Dicheranthus plocamoides Webb (Caryophyllaceae) + Oxelman et al. (2001); Smissen et al. (2002)*†¶ Echium leuchophaeum Sprague and Hutch. (Boraginaceae) + Böhle et al. (1996)*† Echium simplex DC. (Boraginaceae) + Böhle et al. (1996)*† Lavatera phoenicea Vent. (Malvaceae) ++ Fuertes-Aguilar et al. (2002) Hypochaeris oligocephala (Bramwell) Lack (Asteraceae) + Cerbah et al. (1998)* Lotus dumetorum Murr. (Fabaceae) + + Allan et al. (2004)*† Lotus mascaensis Burch. (Fabaceae) + Allan et al. (2004)*† Lugoa revoluta DC. (Asteraceae) + Francisco-Ortega et al. (2001a)*† Monanthes anagensis Praeger (Crassulaceae) + Mort et al. (2002) Monanthes minima (Bolle) Christ (Crassulaceae) + Mort et al. (2002) Sideritis brevicaulis Mend.-Heuer (Lamiaceae) + Barber et al. (2000)*† Sideritis cystosiphon Svent. (Lamiaceae) + Barber et al. (2000)*† Sideritis infernalis Bolle (Lamiaceae) + Barber et al. (2000)*† Sideritis macrostachya Poir. (Lamiaceae) + Barber et al. (2000)*† Sideritis nervosa Poir. (Lamiaceae) + Barber et al. (2000)*† Sonchus fauces-orci Knoche (Asteraceae) + + Kim et al. (1996)*†§ Sonchus tuberifer Svent. (Asteraceae) + Kim et al. (1996)*† Teline salsoloides Arco and Acebes (Fabaceae) + Percy & Cronk (2002)*† Teline pallida Arco and Acebes (Fabaceae) + + Percy & Cronk (2002)*¶ Tolpis crassiuscula Svent. (Asteraceae) + + Moore et al. (2002)*‡ Tolpis glabrescens Kämmer. (Asteraceae) + Moore et al. (2002)* Vierea laevigata Webb (Asteraceae) + Francisco-Ortega et al. (2001b)*†

Geographical distribution compiled from Sventenius (1948), Santos-Guerra & Fernández-Galván (1983), Gómez-Campo (1996), Hernández & Bañares (1996), Beltrán-Tejera et al. (1999), Del Arco-Aguilar (2000), and Varios Autores (2000). Species in bold represent early diverging lineages in phylogenetic studies. *Phylogenetic study based on DNA data from one genomic compartment only. †Phylogenies show low bootstrap support (< 75%) for relevant nodes. ‡The presence of Tolpis crassiuscula in Adeje (Gómez-Campo 1996) needs further confirmation. §The presence of Sonchus fauces-orci outside Adeje and Teno (Gómez-Campo 1996) needs further confirmation. ¶Species is also present in La Gomera.

palaeo-island endemics appear to have a relatively recent terranean (Cerbah et al. 1998). Hypochaeris provides the only origin and are terminal tips of their respective phylogenies. known phylogenetic evidence for a floristic connection Finally, Hypochaeris oligocephala has a unique pattern, is between Macaronesian and the east Mediterranean. the only species of this genus endemic to one of the palaeo- The unusual phylogenetic patterns detected for Hypochaeris, islands of Tenerife and its sister species, Hypochaeris cretensis Lavatera, Sonchus, and Vierea suggest that the three palaeo- (L.) Chaub. and Bory, is distributed in the eastern Medi- islands of Tenerife harbour some old endemic lineages

© 2005 Blackwell Publishing Ltd, Molecular Ecology, 14, 1177–1189 BYSTROPOGON PHYLOGENY 1185 and also provide evidence of floristic connections between Aeonium alliance, Crambe, Lotus, Pericallis, Sideritis, and the Macaronesia and distant regions of the Mediterranean. woody Sonchus alliance (Kim et al. 1996; Panero et al. 1999; Barber et al. 2000; Francisco-Ortega et al. 2002; Mort et al. 2002; Allan et al. 2004). These genera follow a common Interisland colonizations and ecological shifts pattern in which the majority of their clades show both The clades formed by sections Bystropogon and Canariense interisland colonization and several ecological shifts. differ concerning their patterns of interisland colonization The two different modes of speciation shown by Bystro- and ecological shifts. Speciation events within the section pogon are not unique in Macaronesia. A phylogenetic Canariense clade are mostly restricted to the laurel forest. study of the Canarian endemic Gonospermum Less. alliance Populations of Bystropopon canariensis var. smithianus from (Asteraceae) showed that this group is also composed of Tenerife are limited to the southeastern sector of this island two major clades (Francisco-Ortega et al. 2001a). One of where they are linked both to the laurel and pine forest these clades has three species restricted to Gran Canaria, ecological zones. This clade shows extensive interisland and they have radiated into three different ecological colonizations; indeed this is the only clade that has reached zones. The second clade has two major lineages and does the northern archipelago of Madeira. Our data suggest that not have any representative in Gran Canaria; it shows B. canariensis var. smithianus which is endemic to La Palma interisland colonizations among La Gomera, El Hierro, is the sister taxon to the rest of this clade. These results Tenerife, and La Palma. One of the two lineages is restricted imply that Bystropogon originated in the Canary Islands to the lowland scrub, the second lineage is confined to the and dispersed to Madeira, probably as a single coloniza- pine forest. tion event. Bystropogon provides the sixth known example of a colonization route from the Canaries toward Madeira. Evolutionary implications and morphological The other five taxa that appear to have followed this differentiation dispersal route are the Aeonium alliance (Mort et al. 2002; Fairfield et al. 2004), Convolvulus L. (Convolvulaceae) (Carine Barriers to gene flow between the various species of et al. 2004); Crambe (Francisco-Ortega et al. 2002), Pericallis Bystropogon appear to be eco-geographical rather than (Panero et al. 1999), and the woody Sonchus alliance (Kim genetic. Several hybrids between the two sections of this et al. 1996). genus have been found in the wild and formally described Interisland dispersal is common in the section Canariense (La Serna-Ramos 1984). In addition, during our field clade, since each of the Canarian taxa are distributed on studies we have also observed several morphological forms more than one island and there has been at least one dis- that seem to be hybrids between Bystropogon plumosus and persal to Madeira. Previous phylogenetic studies of the Bystropogon origanifolius in Tenerife (A. Santos-Guerra, Macaronesian species have demonstrated that interisland unpublished). colonization between similar ecological zones is one of the Both the ITS and trees are congruent in supporting modes of species diversification in the Canary Islands the sectional classification of Bystropogon proposed by La (Francisco-Ortega et al. 2001a). However, very few studies Serna-Ramos (1984). It seems that gene flow between these have found that this pattern also can involve species from two sections has not been a major factor in the evolutionary the Canary and Madeira archipelagos. Indeed Bystropogon history of this genus, even though natural intersectional and Crambe are the only known cases where sister relation- hybrids are known to occur. Despite the congruence in ships between Canarian and Madeira taxa are confined sectional classification, disagreements between taxonomy to only one ecological zone (Francisco-Ortega et al. 2002). and species-level phylogeny were detected within each of Crambe sventeni Bramwell & Sunding from the island the sections. Multiple accessions from the two varieties of of Fuerteventura is sister to the Madeira-endemic Crambe B. canariensis did not form two separate clades. In addition, fruticosa L. f. Both species thrive in the lowland scrub of the two species from Madeira were nested within a clade their respective islands. formed by the six accessions of B. canariensis. A similar In contrast, speciation events within the section Bystro- pattern was detected within the section Bystropogon clade, pogon clade are defined by ecological shifts among three where the four varieties of B. origanifolius did not form a different ecosystems. This clade does not have any end- monophyletic group. The three other species of this section emic species in Madeira; and each island of the Canaries, were nested inside the clade with the four varieties of with the exception of La Gomera, has at least one single- B. origanifolius. island endemic taxon. The section Bystropogon clade has Molecular phylogenetic studies for other Macaronesian followed an evolutionary route in which radiation into dif- genera have found major incongruences between taxo- ferent ecological zones seems as important as interisland nomic classifications and phylogenetic patterns. These colonization. This mode of plant speciation is followed by discrepancies have been considered to be the result of most of the Canary Island species. Good examples are the hybridization events. The two best examples are provided

© 2005 Blackwell Publishing Ltd, Molecular Ecology, 14, 1177–1189 1186 J. L. TRUSTY ET AL. by Argyranthemum and Sideritis (Francisco-Ortega et al. a particular species gives rise to other species leading to a 1996; Barber et al. 2000). Natural interspecific hybrids are progenitor-derivative pattern. As far as we are aware this known to occur in these two genera and it has been sug- speciation mode has not been reported previously in any gested that hybridization may be the cause of incongruence other oceanic island group. Although, a recent molecular between groups obtained from the taxonomic classification phylogeny for Macaronesian species of Aichryson Webb and and those detected in the phylogenetic trees. Berthel. (Crassulaceae) also shows a common and highly Morphological differences between the two Bystropogon polymorphic species (A. pachycaulon) forming a para- sections are based on reproductive characters concerning phyletic group with respect to three other taxa with more calyx teeth, corolla shape, inflorescence architecture, and restricted distributions (Fairfield et al. 2004). seed coat structure. In contrast, species boundaries within In Bystropogon peripheral populations of widespread each section rely mostly on leaf characters such as leaf species are prone to genetic and morphological changes hairiness, shape, and plant fragrance. We believe that through genetic drift and inbreeding. Most Bystropogon these few leaf traits are more prone to respond rapidly taxa are restricted to single islands, therefore these two to selection and are likely to be highly homoplasious. We evolutionary processes may act to accelerate speciation cannot rule out that these morphological traits are under events. simple genetic control as it has been proven for the Hawai- ian endemic species of Tetramolopium Nees (Asteraceae). Conclusions and future molecular perspectives Sex expression differences among island endemics of this genus are mostly controlled by two loci (Whitkus et al. With only seven species, Bystropogon is a small genus com- 2000). pared with other taxonomic groups with species endemic La Serna-Ramos (1984) distinguished two morphological to Macaronesia. These groups include the Aeonium alliance groups in Bystropogon origanifolius. The first of these (63 endemic species), Argyranthemum (26 endemic species), groups has the two varieties from La Palma and El Hierro Echium (27 endemic species), Sideritis (26 endemic species), (i.e. var. ferrensis and palmensis), and they have spatulate or the woody Sonchus alliance (21 endemic species) (Böhle leaves that are attenuated at their bases. The second group et al. 1996; Kim et al. 1996; Santos-Guerra 2001; Mort et al. has the other three varieties; plants from this group 2002; Marrero & Navarro 2003). These groups not only have leaves which are neither spatulate nor attenuated at have large number of endemic species, but they also their bases. occur in many of the Macaronesian islands and show Bystropogon section Bystropogon is split into two major extensive radiations in all of the ecosystems of Macaronesia. clades which concord with the morphological groups Despite its small size, Bystropogon provides unique suggested by La Serna-Ramos (1984). These clades had insights into the evolutionary history of plants endemic bootstrap support of 70% and 78%. However, there were to the Macaronesian islands, particularly on the phylo- other single-island endemics nested in each of these major genetic position of taxa endemic to geologically ancient clades. Bystropogon wildpretii from La Palma grouped with regions, and how interisland colonizations and ecological B. origanifolius var. ferrensis and B. origanifolius var. palmensis. shifts shaped plant biodiversity in Macaronesia. Further The leaves of Bystropogon wildpretii are also spatulate and population genetic studies are needed to understand slightly attenuated at their bases (I. La Serna-Ramos, better the potential occurrence of multiple dispersal events personal communication). The second major clade has between islands, and to what extent populations of taxa the remaining varieties of B. origanifolius together with occurring in the palaeo-islands of Tenerife have unique B. odoratissimus and B. plumosus. Taxa from this clade do not patterns of genetic diversity not found in the rest of the have spatulate leaves, although plants of B. odoratissimus archipelago. and B. plumosus have leaves which are sometimes slightly attenuated at their bases. Our phylogenetic results show the two species with Acknowledgements the most restricted distribution (B. odoratissimus and We dedicate this paper to Henrique Miguel Costa Neves, pioneer B. wildpretii) are terminal branches within the clade of the of Biological Conservation in the archipelago of Madeira. Without widespread and polymorphic species B. origanifolius. This his perseverance, enthusiasm, and tireless work the few remain- suggests B. origanifolius as progenitor of the two other ing Macaronesian individuals of the Mediterranean monk seal species. This phylogenetic pattern has been previously would not have found a secure refuge in the Desertas Islands at reported for continental taxa, and there are several Parque Natural da Madeira. Mark Carine, T. Collins, J. Geiger, C. Lewis, A. Marrero, M. Maunder, I. La Serna-Ramos and S. Zona examples from the California flora (Baldwin 2003; Gottlieb critically read and provided valuable suggestions to an early draft 2003). ‘Quantum speciation’(Grant 1981) is considered as of this paper. Irene La Serna-Ramos provided helpful discussions the evolutionary mechanism behind these phylogenetic on the morphology, ecology, and evolutionary patterns of Bystro- results (Rieseberg & Brouilet 1994). In this speciation mode pogon. Aguedo Marrero and J. Caujapé-Castells kindly helped JFO

© 2005 Blackwell Publishing Ltd, Molecular Ecology, 14, 1177–1189 BYSTROPOGON PHYLOGENY 1187 during field studies in the island of Gran Canaria. Carl Lewis pro- chromosomal evolution. Molecular Biology and Evolution, 15, vided valuable insights on the interpretation of dG values. Bruce 345–354. Baldwin guided us on relevant literature on quantum speciation DeBry RW, Olmstead RG (2000) A simulation study of reduced and shared with us unpublished results from his research with tree-search effort in bootstrap resampling analysis. Systematic Californian tarweeds. This work was supported by an FIU Pre- Biology, 49, 171–179. sidential Fellowship, a United States Environmental Protection Del Arco-Aguilar MJ (2000) Notes on the taxonomy and nomen- Agency STAR Fellowship, and a Tropical Biology Program (TBP- clature of Teline pallida (Poir.) G. Kunkel. Taxon, 49, 17–25. FIU) research assistantship to JT. Additional support was pro- Del Arco-Aguilar MJ, Pérez-de-Paz PL, Wildpret-de-La-Torre W, vided by start-up funds from the Tropical Biology Program of FIU, Lucia-Sauquillo V, Salas-Pascual M (1990) Atlas Cartográfico de research funds from Fairchild Tropical Botanic Garden (FTBG), an Los Pinares Canarios: La Gomera y El Hierro. Dirección General de FIU Provost’s Office Summer Research Competition grant, and a Medio Ambiente y Conservación de La Naturaleza, Consejería Summer Research Position from FTBG to JFO. This is contribution de Política Territorial, Gobierno de Canarias, Santa Cruz de number 045 of the TBP-FIU. Tenerife, Spain. Downie SR, Katz-Downie DS (1996) A molecular phylogeny of Apiaceae subfamily Apoideae: evidence from nuclear ribosomal References DNA internal transcribed spacer sequences. American Journal of Botany, 83, 234–251. Allan GJ, Francisco-Ortega J, Santos-Guerra A, Boerner E, Zimmer Emerson BC (2002) Evolution on oceanic islands: molecular EA (2004) Molecular phylogenetic evidence for the geographic phylogenetic approaches to understanding pattern and process. origin and classification of Canary Island Lotus (Fabaceae: Molecular Ecology, 11, 951–966. Loteae). Molecular Phylogenetics and Evolution, 32, 122–138. Fairfield KN, Mort ME, Santos-Guerra A (2004) Phylogenetics and Ancochea E, Fuster JM, Ibarrola E et al. (1990) Volcanic evolution evolution of the Macaronesian members of the genus Aichryson of the island of Tenerife (Canary Islands) in light of new (Crassulaceae) inferred from nuclear and chloroplast sequence K-Ar data. Journal of Volcanology and Geothermal Research, 44, data. Plant Systematics and Evolution, 248, 71–83. 231–249. Felsenstein J (1985) Confidence limits on phylogenies: an approach Baldwin BG (2003) A phylogenetic perspective on the origin and using bootstrap. Evolution, 39, 783–791. evolution of Madiinae. In: Tarweeds and Silverswords (eds Car- Fernández-Galván M (1983) Esquema de la vegetación potencial lquist S, Baldwin BG, Carr GD), pp. 193–228. Missouri Botanical de la Isla de Gomera. In: Comunicações Apresentadas ao II Congreso Garden Press, St. Louis, Missouri. International pro Flora Macaronesica. 19–25 Junho de 1977 (ed. Baldwin BG, Crawford DJ, Francisco-Ortega J, Kim S-C, Sang T, Anonymous), pp. 269–293. Funchal, Portugal. Stuessy TF (1998) Molecular phylogenetic insights on the Fernández-Palacios JM (1999) Marco ecológico de las Islas Canarias. origin and evolution of oceanic island plants. In: Molecular In: Ecología y Cultura en Canarias (eds Fernández-Palacios JM, Systematics of Plants II: DNA Sequencing (eds Soltis DE, Soltis PS, Bacallado JJ, Belmonte JA), pp. 83–105. Organismo Autónomo Doyle JJ), pp. 410–441. Kluwer Academic Publishers, Boston, Complejo Insular de Museos y Centros, Cabildo de Tenerife, Massachusetts. Santa Cruz de Tenerife, Spain. Barber JC, Francisco-Ortega J, Santos-Guerra A, Marrero A, Jansen RK Francisco-Ortega J, Jansen RK, Santos-Guerra A (1996) Chloroplast (2000) Evolution of endemic Sideritis (Lamiaceae) in Macaronesia: DNA evidence of colonization, adaptive radiation, and hybrid- insights from a chloroplast DNA restriction site analysis. ization in the evolution of the Macaronesian flora. Proceedings of Systematic Botany, 25, 633–647. the National Academy of Sciences of the United States of America, 93, Beltrán-Tejera E, Wildpret-De-La-Torre W, León-Arencibia MC, 4085–4090. García-Gallo A, Reyes-Hernández J (1999) Libro Rojo de Las Especies Francisco-Ortega J, Fuertes-Aguilar J, Kim S-C et al. (2002) Phylogeny de la Flora Canaria Incluidas en el Anexo II de la Directiva 92/43/CEE of the Macaronesian endemic Crambe section Dendrocrambe del Consejo. Dirección General de Conservación de la Naturaleza, (Brassicaceae) based on internal transcribed spacer sequences of La Laguna, Tenerife, Spain. nuclear ribosomal DNA. American Journal of Botany, 89, 1984–1990. Böhle U-R, Hilger HH, Martin WF (1996) Island colonization and Francisco-Ortega J, Barber J, Santos-Guerra A, Febles-Hernández R, evolution of the insular woody habit in Echium L. (Boragi- Jansen RK (2001a) Origin and evolution of the endemic genera naceae). Proceedings of the National Academy of Sciences of the of Gonosperminae (Asteraceae: Anthemideae) from the Canary United States of America, 93, 11740–11745. Islands: evidence from nucleotide sequences of the internal Bramwell D, Bramwell ZI (1974) Wild Flowers of the Canary transcribed spacers of the nuclear ribosomal DNA. American Islands. Cabildo Insular de Tenerife, Santa Cruz de Tenerife, Journal of Botany, 88, 161–169. Spain. Francisco-Ortega J, Park S-J, Santos-Guerra A, Benabid A, Jansen Capelo J, Costa JC, Lousã M et al. (1999) Vegetação da Madeira RK (2001b) Origin and evolution of the endemic Macaronesian (Portugal): I – Aproximação à tipologia fitossociológica. Silva Inuleae (Asteraceae): evidence from the internal transcribed Lusitana, 7, 257–286. spacers of nuclear ribosomal DNA. Biological Journal of the Linnean Carine MA, Russell SJ, Santos-Guerra A, Francisco-Ortega J (2004) Society, 72, 77–97. Relationships of the Macaronesian and Mediterranean floras: Fuertes-Aguilar J, Ray MF, Francisco-Ortega J, Santos-Guerra A, molecular evidence for multiple colonizations into Macaronesia Jansen RK (2002) Molecular evidence from chloroplast and and back-colonization of the continent in Convolvulus L. (Con- nuclear markers for multiple colonizations of Lavatera (Malvaceae) volvulaceae). American Journal of Botany, 91, 1070–1085. in the Canary Islands. Systematic Botany, 27, 74–83. Cerbah M, Souza-Chies T, Jubier MF, Lejeune B, Siljak-Yakovlev S Gómez-Campo C (ed.) (1996) Libro Rojo de las Especies Vegetales (1998) Molecular phylogeny of the genus Hypochaeris using Amenazadas de Las Islas Canarias. Gobierno de Canarias, Tenerife, internal transcribed spacers of nuclear rDNA: inference for Spain.

Gottlieb LD (2003) Rethinking classic examples of recent speciation Oxelman B, Ahlgren B, Thulin M (2002) Circumscription and in plants. New Phytologist, 161, 71–82. phylogenetic relationships of Gymnocarpos (Caryophyllaceae – Grant V (1981) Plant Speciation, 2nd edn. Columbia University Paranychioideae). Edinburgh Journal of Botany, 59, 221–237. Press, New York. Panero JL, Francisco-Ortega J, Jansen RK, Santos-Guerra A (1999) Guillou H, Carracedo JC, Paris R, Pérez-Torrado FJ (2004) Impli- Molecular evidence for multiple origins of woodiness and a cations for the early shield-stage evolution of Tenerife from K/ New World biogeographic connection of the Macaronesian Ar ages and magnetic stratigraphy. Earth and Planetary Science Island endemic Pericallis (Asteraceae: Senecioneae). Proceedings Letters, 222, 599–614. of the National Academy of Sciences of the United States of America, Hernández E, Bañares A (1996) Aeonium volkerii sp. nov., nuevo 96, 13886–13891. endemismo de la isla de Tenerife, Islas Canarias (Crassulaceae). Percy DM, Cronk QCB (2002) Different fates of island brooms: Vieraea, 25, 159–168. contrasting evolution in Adenocarpus, Genista, and Teline (Geni- Hernández E, García D (1996) Bystropogon odoratissimus Bolle en el steae, Fabaceae) in the Canary Islands and Madeira. American Macizo de Teno (Tenerife, Islas Canarias). Botánica Macaronésica, Journal of Botany, 89, 854–864. 23, 313–315. Rieseberg LH, Brouilet L (1994) Are many plant species para- Holmgren PK, Holmgren NH, Barnett LC (1990) Index herbariorum. phyletic? Taxon, 43, 21–32. Part I: the herbaria of the world, 8th edn. Regnum Vegetabile, 120, Santos-Guerra A (2001) Flora vascular nativa. In: Naturaleza de las 1–693. Islas Canarias Ecología y Conservación (eds Fernández-Palacios Jardim R, Francisco D (2000) Flora Endémica da Madeira. Múchia, JM, Martín-Esquivel JL), pp. 185–192. Publicaciones Turquesa, Funchal, Portugal. Santa Cruz de Tenerife, Spain. Juan C, Emerson BC, Oromi P, Hewitt GM (2000) Colonization Santos-Guerra A, Fernández-Galván M (1983) Vegetación del and diversification: towards a phylogeographic synthesis for Macizo de Teno. Datos para su conservación. In: Comunicações the Canary Islands. Trends in Ecology and Evolution, 15, 104– Apresentadas ao II Congreso International Pro Flora Macaronesica. 109. 19–25 Junho de 1977 (ed. Anonymous), pp. 385–424. Funchal, Kim SC, Crawford DJ, Francisco-Ortega J, Santos-Guerra A (1996) Portugal. A common origin for woody Sonchus and five related genera in Silvertown J (2004) The ghost of competition past in the phylogeny the Macaronesian Islands: molecular evidence for extensive of island endemic plants. Journal of Ecology, 92, 168–173. radiation. Proceedings of the National Academy of Sciences of the Simmons MP, Ochoterena H (2000) Gaps as characters in United States of America, 93, 7743–7748. sequence-based phylogenetic analyses. Systematic Biology, 49, Kim K-J, Jansen RK (1994) Comparisons of phylogenetic hypothesis 369–381. among different data sets in dwarf dandelions (Krigia): Addi- Smissen RD, Clement JC, Garnock-Jones PJ, Chambers GK (2002) tional information from internal transcribed spacers sequences Subfamilial relationships within Caryophyllaceae as inferred of nuclear DNA. Plant Systematics and Evolution, 190, 157–185. from 5′ ndhF sequences. American Journal of Botany, 89, 1336– La Serna-Ramos IE (1984) Revisión del género Bystropogon L’Hér., 1341. nom. cons. (Lamiaceae-Stachyoideae): endemismo de la región Susanna A, Garnatje T, García-Jacas N (1999) Molecular phylogeny Macaronésica. Phanerogamum Monographiaea, 18, 1–380. of Cheirolophus (Asteraceae–Centaureinae) based on ITS sequences Maddison DR (1991) The discovery and importance of multiple of nuclear ribosomal DNA. Plant Systematics and Evolution, 214, islands of most-parsimonious trees. Systematic Zoology, 40, 315– 147–160. 328. Sventenius ERS (1948) Plantas nuevas o poco conocidas de Ten- Marrero A (2004) Procesos evolutivos en plantas insulares, el caso erife. Boletín del Instituto Nacional de Investigaciones Agronómicas, de Canarias. In: Ecología Insular / Island Ecology (eds Fernández- 18, 273–291. Palacios JM, Morici C), pp. 305–356. Asociación Española de Swofford DL (2002) PAUP*: Phylogenetic Analysis Using Parsimony Ecología Terrestre & Cabildo Insular de La Palma, Spain. (*and Other Methods), Version 4.0b10. Sinauer Associates, Marrero A, Francisco-Ortega J (2001) Evolución en islas: la metá- Sunderland, Massachusetts. fora espacio — tiempo — forma. In: Naturaleza de las Islas Canarias. Taberlet P, Gielly L, Puatou G, Bouvet J (1991) Universal primers Ecología y Conservación (eds Fernández-Palacios JM, Martín- for amplification of three non-coding regions of chloroplast Esquivel JL), pp. 133–140. Publicaciones Turquesa, Santa Cruz DNA. Plant Molecular Biology, 17, 1105–1109. de Tenerife, Spain. Thirlwall MF, Singer BS, Marriner GF (2000) 39Ar–40Ar ages and Marrero A, Navarro B (2003) Sideritis amagroi sp. nov (Lamiaceae) geochemistry of the basaltic shield stage of Tenerife, Canary una nueva especie para Gran Canaria (Islas Canarias). Botánica Islands, Spain. Journal of Volcanology and Geothermal Research, Macaronésica, 24, 57–66. 103, 247–297. Martín JL, Vera MA, Arechavaleta M (1999) Biodiversidad Thompson JD, Gibson TJ, Plewniak F, Jeanmougin F, Higgins DG taxonómica y análisis de prioridad para el establecimiento de (1997) The clustal x window interface: flexible strategies for áreas protegidas. Vieraea, 27, 245–253. multiple sequence alignment aided by quality analysis tools. Moore MJ, Francisco-Ortega J, Santos-Guerra A, Jansen RK (2002) Nucleic Acids Research, 24, 4876. Chloroplast DNA evidence for the roles of island colonization Trusty J, Olmstead RG, Bogler DJ, Santos-Guerra A, Francisco- and extinction in Tolpis (Asteraceae: Lactuceae). American Journal Ortega J (2004) Using molecular data to test a biogeographic of Botany, 89, 518–526. connection of the Macaronesian genus Bystropogon (Lamiaceae) Mort ME, Soltis DE, Soltis PS, Francisco-Ortega J, Santos-Guerra A to the New World: a case of conflicting phylogenies. Systematic (2002) Phylogenies and evolution of the Macaronesian clade of Botany, 29, 702–715. Crassulaceae inferred from nuclear and chloroplast sequence Valido A, Dupont YL, Olesen JM (2004) Bird–flower interactions in data. Systematic Botany, 27, 271–288. the Macaronesian islands. Journal of Biogeography, 31, 1945–1953.

Varios Autores (2000) Lista roja de la flora vascular española. Conservación Vegetal, 6 (extra), 11–38. Jennifer Trusty is a graduate student at FIU/FTBG, her research White TJ, Bruns T, Lee S, Taylor J (1990) Amplification and direct focuses on the origin, evolution, and conservation genetics of sequencing of fungal ribosomal RNA genes for phylogenetics. island plants, particularly from the Antilles, Cocos Island (Pacific In: PCR Protocols: a Guide to Methods and Application (eds Innis M, Ocean), and Macaronesia. Richard Olmstead is Professor of Gelfand D, Sninsky J, White T), pp. 315–312. Academic Press, Biology at University of Washington. His main research concerns San Francisco. plant systematics, particularly within the Lamiales and Solanales. Whitkus R, Doan H, Lowrey TK (2000) Genetics of adaptive Arnoldo Santos-Guerra is Director of Research of ORT. His radiation in Hawaiian species of Tetramolopium (Asteraceae). main research concerns plant biodiversity in Macaronesia. Susana III. Evolutionary genetics of sex expression. Heredity, 85, 37– Sa-Fontinha is Director of PNM, she specializes on conservation 42. and taxonomy of Macaronesian plants. Javier Francisco-Ortega Zuker M (2003) mfold web server for nucleic acid folding and is Associate Professor at FIU, he is interested in island plant hybridization prediction. Nucleic Acids Research, 31, 3406– biodiversity and conservation. 3415.