Revisiting the phylogeny of Bombacoideae (Malvaceae): Novel relationships, morphologically cohesive clades, and a new tribal classification based on multilocus phylogenetic analyses

David A Baum; Aline Mota; Suzana Alcantara; Jefferson Carvalho-Sobrinho

Bombacoideae (Malvaceae) is a clade of deciduous trees with a marked dominance in many forests, especially in the Neotropics. The historical lack of a well-resolved phylogenetic framework for Bombacoideae hinders studies in this ecologically important group. We reexamined phylogenetic relationships in this clade based on a matrix of 6465 nuclear (ETS, ITS) and plastid (matK, trnL-trnF, trnS-trnG) DNA characters. We used maximum parsimony, maximum likelihood, and Bayesian inference to infer relationships among 108 species ($70% of the total number of known species). We analyzed the evolution of selected morphological traits: trunk or branch prickles, calyx shape, endocarp type, seed shape, and seed number per fruit, using ML reconstructions of their ancestral states to identify possible synapomorphies for major clades. Novel phylogenetic relationships emerged from our analyses, including three major lineages marked by fruit or seed traits: the winged-seed clade (Bernoullia, Gyranthera, and Huberodendron), the spongy endocarp clade (Adansonia, Aguiaria, Catostemma, Cavanillesia, and Scleronema), and the Kapok clade (Bombax, Ceiba, Eriotheca, Neobuchia, Pachira, Pseudobombax, Rhodognaphalon, and Spirotheca). The Kapok clade, the most diverse lineage of the subfamily, includes sister relationships (i) between Pseudobombax and ''Pochota fendleri " a historically incertae sedis taxon, and (ii) between the Paleotropical genera Bombax and Rhodognaphalon, implying just two bombacoid dispersals to the Old World, the other one involving Adansonia. This new phylogenetic framework offers new insights and a promising avenue for further evolutionary studies. In view of this information, we present a new tribal classification of the subfamily, accompanied by an identification key.

Molecular Phylogenetics and Evolution 101 (2016) 56–74 Contents lists available at ScienceDirect Molecular Phylogenetics and Evolution journal homepage: www.elsevier.com/locate/ympev Revisiting the phylogeny of Bombacoideae (Malvaceae): Novel relationships, morphologically cohesive clades, and a new tribal classification based on multilocus phylogenetic analyses Jefferson G. Carvalho-Sobrinho a,⇑, William S. Alverson b, Suzana Alcantara c, Luciano P. Queiroz d, Aline C. Mota d, David A. Baum b a Colegiado de Ciências Biológicas, Universidade Federal do Vale do São Francisco – UNIVASF, BR-407, Km 12, Vila CS-01, Petrolina, Pernambuco 56300-990, Brazil Department of Botany, Birge Hall, University of Wisconsin-Madison, Madison, WI 53706, USA c Departamento de Botânica-CCB, Universidade Federal de Santa Catarina – UFSC, Florianópolis, SC 88040900, Brazil d Programa de Pós-Graduação em Botânica, Universidade Estadual de Feira de Santana – UEFS, Av. Transnordestina, s/n, Novo Horizonte, Feira de Santana, Bahia 44036900, Brazil b a r t i c l e i n f o Article history: Received 21 May 2015 Revised 21 April 2016 Accepted 2 May 2016 Available online 3 May 2016 Keywords: Baobab Fruit traits Kapok group Neotropics Tribal classification a b s t r a c t Bombacoideae (Malvaceae) is a clade of deciduous trees with a marked dominance in many forests, especially in the Neotropics. The historical lack of a well-resolved phylogenetic framework for Bombacoideae hinders studies in this ecologically important group. We reexamined phylogenetic relationships in this clade based on a matrix of 6465 nuclear (ETS, ITS) and plastid (matK, trnL-trnF, trnS-trnG) DNA characters. We used maximum parsimony, maximum likelihood, and Bayesian inference to infer relationships among 108 species (70% of the total number of known species). We analyzed the evolution of selected morphological traits: trunk or branch prickles, calyx shape, endocarp type, seed shape, and seed number per fruit, using ML reconstructions of their ancestral states to identify possible synapomorphies for major clades. Novel phylogenetic relationships emerged from our analyses, including three major lineages marked by fruit or seed traits: the winged-seed clade (Bernoullia, Gyranthera, and Huberodendron), the spongy endocarp clade (Adansonia, Aguiaria, Catostemma, Cavanillesia, and Scleronema), and the Kapok clade (Bombax, Ceiba, Eriotheca, Neobuchia, Pachira, Pseudobombax, Rhodognaphalon, and Spirotheca). The Kapok clade, the most diverse lineage of the subfamily, includes sister relationships (i) between Pseudobombax and ‘‘Pochota fendleri” a historically incertae sedis taxon, and (ii) between the Paleotropical genera Bombax and Rhodognaphalon, implying just two bombacoid dispersals to the Old World, the other one involving Adansonia. This new phylogenetic framework offers new insights and a promising avenue for further evolutionary studies. In view of this information, we present a new tribal classification of the subfamily, accompanied by an identification key. Ó 2016 Elsevier Inc. All rights reserved. 1. Introduction Bombacoideae is a lineage of Malvaceae (Alverson et al., 1999; Bayer et al., 1999; Nyffeler et al., 2005) that includes trees with outstanding ecological importance in the tropics. It encompasses about 17 genera and 160 species with ca. 90% of the species distributed in the Neotropics. Bombacoideae comprises some of the most abundant and dominant tree species in many Neotropical forests (Andel, 2001; Ferreira and Prance, 1998; Linares-Palomino and Alvarez, 2005; Pennington and Sarukhán, 1968; Pennington et al., 2004; Prance et al., 1976). In the Paleotropics, it is represented ⇑ Corresponding author. E-mail address: jefferson.sobrinho@univasf.edu.br (J.G. Carvalho-Sobrinho). http://dx.doi.org/10.1016/j.ympev.2016.05.006 1055-7903/Ó 2016 Elsevier Inc. All rights reserved. by fewer than 18 native species in three genera: Adansonia L. (eight or nine species), Bombax L. (three or four species), and Rhodognaphalon (Ulbr.) Roberty (three species). Whether the disjunct distribution of Ceiba pentandra (L.) Gaertn. and Pachira glabra Pasq. in the African and American continents is natural or anthropogenic has long been controversial (Dick et al., 2007; Robyns, 1963). Ignoring these two widely cultivated species, three independent migrations from the Neotropics to the Paleotropics are usually invoked to explain the worldwide distribution of Bombacoideae (Duarte et al., 2011). Most traditional systematic and phylogenetic studies in Bombacoideae have focused on floral characters, especially the androecium (Bentham, 1843, 1862; Carvalho-Sobrinho et al., 2009; Duarte et al., 2011; Gibbs and Alverson, 2006; Gibbs and Semir, 2003; Nyffeler et al., 2005; Robyns, 1963). Genera possessing J.G. Carvalho-Sobrinho et al. / Molecular Phylogenetics and Evolution 101 (2016) 56–74 flowers with many filaments partially connate in a tube and monothecate anthers were historically placed in the tribe Adansonieae (Hutchinson, 1967; Schumann, 1886, 1895; see Fig. 1), which comprised most species of Bombacoideae. According to some classification systems, Adansonieae was also characterized by possession of palmately compound leaves, resulting in the inclusion of Ceiba Mill., Bernoullia Oliv., Gyranthera Pittier, and Spirotheca Ulbr. (e.g., Bakhuizen van den Brink, 1924; Barroso et al., 2002). Such a broadened circumscription of Adansonieae makes the tribe rather variable in flower traits: Ceiba and Spirotheca have five stamens with two or four thecae each (Gibbs and Alverson, 2006; Gibbs and Semir, 2003); whereas Bernoullia and Gyranthera have anthers with many distal, sessile thecae on an elongated staminal tube (von Balthazar et al., 2006). Furthermore, Bernoullia and Gyranthera exhibit distinct capsular fruits enclosing winged seeds that differ from the woody berries of Adansonia and from the capsules with kapok and non-winged seeds of Ceiba and most other Adansonieae genera. Recent studies based on morphological (Carvalho-Sobrinho and Queiroz, 2011) and molecular data (Duarte et al., 2011), however, indicate that Adansonieae as conceived historically is not monophyletic without inclusion of genera with simple or 1-foliolate leaves traditionally placed in the tribes Durioneae, Hampeae, ‘Catostemmateae’, or Matiseae (Fig. 1): Catostemma Benth., Cavanillesia Ruiz & Pav., Huberodendron Ducke, and Scleronema Benth (Edlin, 1935; Hutchinson, 1967; Schumann, 1895; Takhtajan, 1997). However, all the aforementioned genera compose a molecularly well-supported clade named the core Bomba- 57 coideae (Baum et al., 2004; Duarte et al., 2011), which may be defined as corresponding to the smallest monophyletic group containing Gyranthera and Bombax (Baum et al., 2004). At the generic level, previous molecular work has supported monophyly of most currently accepted genera. However, Bombacopsis and Rhodognaphalopsis, erected by Pittier (1916) and Robyns (1963), respectively, have been shown to be embedded within Pachira s. lat. (Duarte et al., 2011). Likewise, a recent molecular study suggested that Eriotheca Schott & Endl. forms a paraphyletic grade relative to Pachira Aubl. (Duarte et al., 2011). This same study also found that one species of Pachira, P. quinata (=Bombacopsis quinata), is not related to the remainder of the genus and was recently transferred to Pochota (Alverson and Duarte, 2015). Finally, the wisdom of fusing Ceiba and Chorisia Kunth into Ceiba has long been controversial (Gibbs and Semir, 2003; Gibbs et al., 1988; Ravenna, 1998), but has not yet been thoroughly assessed in a phylogenetic framework. Despite recent efforts to clarify the phylogeny of Bombacoideae, uncertainty remains, especially in regards to intergeneric relationships. The lack of a well resolved phylogenetic framework hampers development of a coherent tribal classification and investigation of the tempo and mode of evolution of Bombacoideae, which is crucial for understanding the diversification of the group and of the Neotropical flora. Here, we use newly generated DNA sequence data of the trnS-trnG spacer region of plastid DNA (cpDNA) and the External Transcribed Spacer (ETS) of the nuclear ribosomal DNA (nrDNA), in combination with previously studied markers, to better resolve the phylogeny of Bombacoideae. Specifically, we Fig. 1. Taxonomic placement of genera of Bombacoideae (Malvaceae) according to different classification systems. Dotted lines indicate changes in genus circumscription. Genera described after the previous treatment are indicated by an asterisk. Quotation marks indicate tribes not validly published. 58 J.G. Carvalho-Sobrinho et al. / Molecular Phylogenetics and Evolution 101 (2016) 56–74 provide much-improved taxon sampling, including around 70% of the known species and 100% of the genera of Bombacoideae. In addition, we assess the evolution of selected morphological traits in order to find synapomorphies that could aid in the identification of major clades. 2. Material and methods 2.1. Taxon sampling strategy We sampled representatives of all genera of Bombacoideae and 108 of 160 known species, including most species of Ceiba (15 of 18 species, four of which were described in Chorisia: C. chodatii, C. insignis, C. speciosa, and C. ventricosa) and Pseudobombax (23 of 25 species plus Pseudobombax sp. and P. aff. campestre [Mart.] A. Robyns) (Appendix A). Septotheca tessmannii Ulbr., the putative sister lineage to the core Bombacoideae was also included, as were seven former Bombacaceae that are now thought to fall outside Bombacoideae (Baum et al., 2004; Duarte et al., 2011): Chiranthodendron pentadactylon Larreat., Fremontodendron californicum (Torr.) Coville, Hampea appendiculata Standl., Ochroma pyramidale (Cav. ex Lam.) Urb., Patinoa sphaerocarpa Cuatrec., Pentaplaris doroteae L.O.Williams & Standl., and Phragmotheca ecuadorensis W.S. Alverson. In all combined analyses, Sterculia lanceolata Cav. and Sterculia nobilis Sm. were included as more distant outgroups to root the tree. Sequences were obtained from 63 specimens collected in the field, 21 herbarium specimens, and 43 samples used in Duarte et al. (2011) (Appendix A). 2.2. DNA extraction, amplification, and sequencing All sequences of matK and trnL-trnF plus 31 sequences of ITS from previous phylogenetic studies (Alverson et al., 1999; Baum et al., 2004; Duarte et al., 2011) were obtained from GenBank. For new sequences of ITS and the newly explored ETS and trnStrnG, laboratory work was performed at the Plant Molecular Systematics Laboratory (LAMOL) of Feira de Santana State University (UEFS). Total DNA was extracted from leaf tissue using DNeasy plant mini kits (Qiagen, Valencia, California). The manufacturer’s protocol of the DNeasy plant mini kit was followed with the exception of eluting with 50 lL, instead of 100 lL, of AE buffer in order to yield a higher concentration of DNA. Amplification was carried out in a 9700 GeneAmp Thermocycler (Applied Biosystems, Singapore). Polymerase chain reactions (PCR) were performed using the TopTaq Master Mix Kit (QIAGEN GmbH, Hilden, Germany) following the manufacturer’s protocol and 5 pmol of each primer. Quantities of DNA for reactions were not measured. In the approach used for ITS and ETS, the PCR reaction also included 0.2 lL of BSA 0.3% (bovine serum albumin), 2 lL of betaine 5 M, and 0.2 lL of DMSO 99.5% (dimethyl sulfoxide). The ETS region was amplified using the primers 18S-IGS, developed by Baldwin and Markos (1998) and AcR2, designed by J. Miller (CSIRO, Canberra) and used in Acacia (Ariati et al., 2006) and Calliandra (Souza et al., 2014). This gene region was amplified using the following PCR conditions: (1) an initial denaturing step of 97 °C for 1 min; (2) 40 cycles of denaturing at 97 °C for 10 s, annealing 55 °C for 30 s, and extension at 72 °C for 20 s; and (3) a final extension step of 72 °C for 7 min. The ITS region was amplified using the plant specific primers, ITS17 and ITS26 (Sun et al., 1994) and the following PCR conditions: (1) an initial denaturing step of 94 °C for 3 min; (2) 28 cycles of denaturing at 94 °C for 45 s, annealing 54 °C for 60 s, and extension at 72 °C for 90 s; and (3) a final extension step of 72 °C for 7 min. The trnS-trnG spacer region of chloroplast DNA was amplified using primers trnS (GCU) and trnG (UCC), designed by Hamilton (1999). This chloroplast spacer region was amplified using the following PCR conditions: (1) an initial denaturing step of 94 °C for 1 min, (2) 30 cycles of denaturing at 94 °C for 30 min, annealing 55 °C for 40 s, and extension at 72 °C for 60 s; and (3) a final extension step of 72 °C for 5 min. All PCR products were visualized using agarose gel electrophoresis and successfully amplified products were cleaned using QIAquick PCR purification kits (QIAGEN, Valencia, California) or by enzymatic treatment with Exonuclease I and shrimp alkaline phosphatase (ExoSapIT, GE Healthcare, Buckinghamshire, U.K.) using protocols recommended by manufacturers. The QIAquick protocol used a final elution with 30 lL instead of 50 lL. Cleaned PCR products were cycle-sequenced with the same primers as used for amplifications using the Big Dye Terminator kit version 3.1 (Applied Biosystems, Foster City, California, U.S.A.), except for ITS for which we used primers ITS4 (White et al., 1990) and ITS92 (Desfeux and Lejeune, 1996). Complementary strands for each region were sequenced using the automatic sequencers Spectrumedix SCE9624 and ABI3130XL at LAMOLUEFS. All newly generated sequences were uploaded to GenBank (Benson et al., 2010) (see Appendix A). 2.3. Data matrix and alignment Complementary strands were combined and base-calling verified with the Staden package (Staden, 1996). Alignments were performed in Muscle (Goujon et al., 2010) and corrected by eye in Mesquite v3.02 (Maddison and Maddison, 2010), when necessary. Six data matrices were constructed: (1) ETS only, (2) ITS only, (3) trnS-trnG only, (4) ETS and ITS (nuclear matrix), (5) matK, trnStrnG, trnL-trnF (plastid matrix), (6) ETS, ITS, matK, trnS-trnG, trnLtrnF (combined matrix). Incompletely sampled taxa were included in the combined matrices assuming that data sets differing considerably in gene and taxon sampling can be gainfully combined (Cho et al., 2011) and frequently increase the accuracy of phylogenies (Jiang et al., 2014). We ran SATé-II (Liu et al., 2012) iterative alignment program with DendroPy 4.0.0 (Sukumaran and Holder, 2010) using default settings on ITS dataset alone, ETS alone, and the two combined to check the effect of alignments errors. All automatic alignment methods and manual corrections produced similar results. The resulting matrices were submitted to TreeBASE (study number S18447). 2.4. Phylogenetic analyses Phylogenies were inferred from each data set using maximum parsimony, maximum likelihood, and Bayesian analyses. A heuristic parsimony search with 10,000 random taxon-addition replicates was performed on each dataset in PAUP⁄ 4.0b10 (Swofford, 2002) using tree bisection-reconnection (TBR) branch swapping and saving 15 trees per replicate. Trees saved in this first round were used as starting trees in a second search using the same parameters and saving a maximum of 15,000 trees. A total of 1000 bootstrap pseudoreplicates (Felsenstein, 1985) were performed using heuristic searches, TBR branch swapping, simple taxon addition, and saving 15 trees per replicate. Gaps were treated as missing data. MrModeltest version 2.3 was used to determine the model of evolution that best fit the data for maximum likelihood and Bayesian inference analyses (Nylander et al., 2004). Bayesian and maximum likelihood analyses were performed in MrBayes version 3.2.3 (Ronquist et al., 2012) and RAxML version 8.1 (Stamatakis, 2006, 2014), respectively, using the Cyberinfrastructure for Phylogenetic Research (CIPRES) Portal 2.0 (Miller et al., 2010). For Bayesian 59 J.G. Carvalho-Sobrinho et al. / Molecular Phylogenetics and Evolution 101 (2016) 56–74 analyses, two independent MCMC runs were conducted, each composed of four chains (one cold and three heated chains) of 50 million generations with sampling every 10,000 generations. The convergence of the runs was assessed by checking the standard deviation of split frequencies. The program Tracer version 1.6 (Rambaut et al., 2014) was used to confirm that stationarity of likelihood scores was achieved early in the chain, and trees from the initial 25% of generations were discarded as burn-in. Remaining trees were summarized in a consensus that included posterior probabilities as branch support estimates. Maximum likelihood (ML) tree searches were implemented with 1000 fast bootstrap replicates under the GTRCAT model unlinked for each partition. Congruence among data partitions was assessed using the incongruence length difference (ILD) test (Farris et al., 1995) as implemented in PAUP⁄ v4.0b10 (Swofford, 2002) through a heuristic parsimony search with 500 random taxon addition replicates, TBR branch swapping, and saving 10 trees per replicate. The possible presence of ITS pseudogenes was investigated through comparison of length and substitution rates in fast-evolving (ITS 1–2) and conserved (5.8S) regions and the presence of polymorphic specimens, following the recommendation of Bailey et al. (2003). 2.5. Ancestral reconstruction of morphological traits in Bombacoideae We analyzed the evolution of five morphological characters often used to recognize groups in the subfamily (see Supplemental data S1–S2 and S5–S7 with the online version of this article). Although most traditional systematic and phylogenetic studies in Bombacoideae have focused on floral characters (see Section 1), they have been marked by several conflicts (Fig. 1), which seem to be common for taxonomic classifications based on floral traits of Neotropical diverse groups (e.g., Cardoso et al., 2012, 2013; Lohmann and Taylor, 2014). The characters and the states coded were as follows: (i) Prickles on trunk or branches: absent, present; (ii) Calyx shape: lobed, truncate, laciniate; (iii) Endocarp type: undifferentiated, papyraceous, spongy, kapok; (iv) Seed shape: non-winged, winged; (v) Seed number per fruit: 1–4, 10–30, 50–800. The morphological data were compiled from herbarium, observations, and the literature (Baum, 1995; Bentham, 1843, 1862; Cuatrecasas, 1950, 1953, 1954a, 1954b; Ducke, 1935a, 1935b, 1938; Dugand, 1943; Gibbs and Alverson, 2006; Gibbs and Semir, 2003; Oliver, 1876; Paula, 1969; Pittier, 1914, 1916, 1921; Robyns, 1963; Ruiz and Pavon, 1797; Shepherd and Alverson, 1981; Steyermark, 1987; Ulbrich, 1914). We would note that prickles in Bombax, Ceiba, and Spirotheca have a distinctive, sharp-pointed morphology that differs from, for example, Cavanillesia ‘‘spines,” which are corky, irregularly shaped protuberances (see Supplemental data S1, F). Ancestral state reconstructions were performed using ML, implemented in Mesquite v3.02 (Maddison and Maddison, 2010), using the Markov k-state 1 parameter (Mk1) model of evolution (Pagel, 1999; Schluter et al., 1997), which assumes equal rates of change between any two character states. We reconstructed the five characters’ evolution over a sample of 1000 post burn-in trees derived from BI analysis of the combined data. For these analyses, we pruned outgroup taxa in order to avoid ancestral reconstruction being biased by the low diversity sampled for outgroups, taking care to keep the branch lengths of the trees unchanged. The ancestral states supported by a threshold log-likelihood ratio of 2.0 were summarized on the BI 50% majority rule consensus tree. When ancestral state reconstruction was ambiguous (character state was not definitely resolved), proportional likelihoods of the alternative states were provided. 3. Results 3.1. Analysis of individual data sets We generated 83 new sequences of ETS, 52 of ITS, and 61 of trnS-trnG for a total of 196 new sequences. The Akaike Information Criterion in MrModeltest found GTR + C as the best fitting model for ETS and ITS, GTR + I + C for matK and trnS-trnG, and HKY + C for trnL-trnF. Summary data for individual and combined data sets, and parsimony results, are presented in Table 1 and Figs. S3 and S4. The trnS-trnG data set had both the lowest number and percentage of informative characters. The ETS data set had the highest percentage of parsimony informative characters whereas ITS had the highest number of informative characters (Table 1). The ILD test suggested a lack of significant conflict among the three cpDNA data sets (p = 0.052), despite matK yielding a distinct optimal tree. There was also no evidence of discordance between the two nuclear markers (p = 0.876). The ILD test suggested, however, existence of significant conflict between the nuclear and plastid data (p = 0.002). A major cause of conflict related to the placement of the genus Spirotheca. The matK data support Spirotheca as sister to Ceiba, ITS data set support it as sister to Bombax, trnL-trnF placed it in a basal grade with Pochota fendleri and Pseudobombax, the ETS data set resolved it as sister to a clade composed by Ceiba and Pachira s.l., and trnS-trnG lacked relevant resolution. Since matK had the most divergent signal of relationships for Spirotheca (S. rosea), this sequence was excluded from combined analyses. After removing this sequence an ILD test ceased detecting significant discordance between the plastid and nuclear data sets. The five genes supported somewhat different outgroup relationships and also differed in the optimal branching order among Table 1 Matrix and tree statistics for the individual and combined data sets. nrETS (nuclear) nrITS (nuclear) matK (plastid) trnL-F (plastid) trnS-G (plastid) Nuclear Plastid Combined No. taxa included in each matrix Length of aligned matrices (bp) No. variable characters 83 532 344 (64.7%) 91 907 540 (59.8%) 58 2629 412 (15.7%) 59 1356 201 (14.8%) 61 1041 185 (17.8%) No. parsimony informative characters No. MP trees MP tree length Consistency index (CI) CI excluding uninformative characters Retention index 264 (49.6%) 391 (43.3%) 166 (6.3%) 104 (7.7%) 64 (6.1%) 95 5026 798 (15.9%) 334 (6.6%) 118 6465 1686 (26.1%) 994 (15.4%) 102 1065 0.5352 0.4902 12,255 2053 0.4584 0.4066 3105 590 0.7678 0.5798 4587 273 0.8278 0.7235 >15,000 249 0.8193 0.6281 107 1439 884 (61.6%) 655 (45.6%) 821 3143 0.4808 0.4314 5193 1131 0.7807 0.6101 315 4161 0.5703 0.4727 0.8370 0.7709 0.7405 0.8813 0.8490 0.7932 0.7971 0.7946 Notes: MP = Maximum parsimony. 60 J.G. Carvalho-Sobrinho et al. / Molecular Phylogenetics and Evolution 101 (2016) 56–74 the major bombacoid clades. However, few of the deep relationships gained strong support from any one gene, leading us to conclude that the combined data set should provide the best estimate of the phylogenetic relationships of Bombacoideae. This tree is shown in Fig. 2, with ML bootstrap support (BS) and Bayesian posterior probabilities (PP) used to summarize clade robustness. 3.2. Clades of Bombacoideae The combined analysis supported the monophyly of the core Bombacoideae clade (BI = 1.0; ML = 94), identified previously (Baum et al., 2004; Duarte et al., 2011). The following three major clades of the core Bombacoideae emerged from our combined analysis of nrDNA and cpDNA, all with clade PP of 1.0: (1) the ‘‘winged seed clade” containing Bernoullia, Gyranthera, and Huberodendron; (2) the ‘‘spongy endocarp clade” comprising Adansonia, Aguiaria, Catostemma, Cavanillesia, and Scleronema; and (3) the ‘‘Kapok clade” including Bombax, Ceiba, Eriotheca, Pachira, Pseudobombax, Rhodognaphalon, and Spirotheca. Within the Kapok clade, three subordinate clades with PP of 0.97–1.0 were recognized: (i) the ‘‘Paleotropical Bombax clade” including Bombax and Rhodognaphalon; (ii) the ‘‘Pachira s.l. clade” composed of Eriotheca and Pachira; (iii) the ‘‘Striated bark clade” including Ceiba, Neobuchia Urb., Pochota, Pseudobombax, and Spirotheca (Fig. 2; see Figs. S5–S7). 3.3. Ancestral state reconstruction of morphological traits We aimed to reconstruct the evolution of five morphological characters (see Appendix B) in order to identify possible synapomorphies and infer the character state of nine target nodes, as marked on Fig. 2. The optimizations were mostly unambiguous, allowing confident assignment of ancestral state for most of the target nodes and the identification of synapomorphies for the major clades (Table 2; Figs. 3, 4 and S5–S7). Only two nodes showed any ambiguity: the last common ancestor of the core Bombacoideae was ambiguous for endocarp type (proportional likelihood of the probable alternative states: papyraceous: 0.5, spongy: 0.3, kapok: 0.19) and seed number per fruit (proportional likelihood of the probable alternative states: 50–800: 0.52, 10–30: 0.44); and the last common ancestor of the Kapok clade was unresolved for the presence/absence of prickles (proportional likelihood of the probable alternative states: present: 0.856, absent: 0.144). 4. Discussion 4.1. Monophyly and major clades of Bombacoideae Our analyses confirmed the monophyly of the core Bombacoideae and of all genera except for Pachira (see Section 4.1.3.2). Fig. 2. Bayesian inference (BI) consensus cladogram based on the combined (plastid and nuclear) data set showing the three major lineages of Bombacoideae (Malvaceae) recovered: the Winged seed clade (green), the Spongy endocarp clade (orange), and the Kapok clade (blue). Numbers above the branches represent Bayesian posterior probabilities (P0.75) as revealed with MrBayes, numbers below the branches indicate maximum likelihood (ML) bootstrap support values (P50%) as obtained with RAxML; only values for nodes discussed on the text are shown. Numbers within circles indicate target nodes for reconstruction of ancestral states of selected morphological traits using ML. (A) Relationships in the Spongy endocarp clade. Photos on the right show the fruits of (a) Huberodendron swietenioides, (b) Adansonia digitata, and (c) Ceiba jasminodora. Photo credits: (a) University of Michigan Herbarium, (b) J. Pierre, (c) Marlon Machado. (B) Relationships in the Kapok clade. Species names in the Amazonian and extra-Amazonian Pachira clades refer to basionyms, except names described in Bombax, to illustrate historical generic concepts. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.) J.G. Carvalho-Sobrinho et al. / Molecular Phylogenetics and Evolution 101 (2016) 56–74 Fig. 2 (continued) 61 Non-winged 50–800 Non-winged 50–800 Non-winged 50–800: 0.84 1–4: 0.14 Winged 10–30 Non-winged 1–4 Lobed Spongy Lobed Papyraceous Lobed Papyraceous: 0.5 Spongy: 0.3 Kapok: 0.19 Winged 50–800: 0.52 10–30: 0.44 Lobed Spongy Non-winged 50–800 Non-winged 50–800 Non-winged 50–800 Truncate Kapok Truncate Kapok Truncate Kapok Truncate Kapok Present Present Absent Present Absent Absent Absent Absent Present: 0.856 Absent: 0.144 Truncate Kapok 7 – Pachira s.l. clade 3 – Spongy endocarp clade 2 – Winged seed clade 4.74 3.85 1.48 1.48 3.94 Prickles on trunk or branches Calyx shape Endocarp type Seed shape Seed number per fruit 1 – Ancestor of core Bombacoideae Rate of transition Node number and reference clade 4 – ‘Catostemmatae’ 5 – Kapok-Clade 6 – Paleotropical Bombax clade 8 – Striated bark clade 9 – Pochota + Pseudobombax J.G. Carvalho-Sobrinho et al. / Molecular Phylogenetics and Evolution 101 (2016) 56–74 Trait Table 2 Results of the ancestral state reconstruction of selected morphological traits of Bombacoideae (Malvaceae) using a maximum likelihood approach. Ancestral character state inferred using a log-likelihood ratio threshold of 2.0. When ancestral state reconstruction for a node is ambiguous, proportional likelihoods of the alternative states are provided. Rate of transition: Mesquite’s estimated rate of change parameter under the Markov k-state 1 parameter (Mk1) model of evolution, in which any particular change between characters’ states is equally probable. 62 The monospecific Septotheca was well supported as falling outside of the core Bombacoideae, as suggested by previous molecular phylogenies (Baum et al., 2004; Duarte et al., 2011). Septotheca inhabits seasonally flooded Amazonian forests and has simple leaves, very distinct from the palmately compound (very rarely unifoliate) leaves of the core Bombacoideae but similar to outgroups. The three major bombacoid lineages inferred from the combined analysis – the Winged seed clade, the Spongy endocarp clade, and the Kapok clade – were also recovered in a previous analysis based on ITS, matK, and trnL-trnF data sets (Duarte et al., 2011). These lineages are morphologically well characterized in relation to floral and fruit traits, though fruit characters have previously been under-emphasized in bombacoid systematics. Our analyses of five morphological traits identified useful synapomorphies for major clades, since there was little ambiguity in ancestral state reconstructions. Ambiguity regarding prickles in the ancestral of the Kapok clade reflects homoplasy (see Fig. S5) as will be more fully discussed. 4.1.1. The Winged seed clade This clade is sister to the other two lineages of the core Bombacoideae and is composed of about ten species of the genera Bernoullia, Gyranthera, and Huberodendron. Representatives of this clade are huge, buttressed trees inhabiting wet forests in the Neotropics. The fruits are typically dehiscent (contrary to Duarte et al., 2011: 695) and large (to 30 cm long), enclosing 10–30 winged seeds (to 20 cm long) surrounded by a papyraceous endocarp (Fig. 2; Fig. S2, E–G; see also Cuatrecasas, 1950: 88; Gleason, 1934: 109; Oliver, 1876: plate 1170). These fruit and seed characters along with their distinctive scorpioid inflorescences (Fig. S1, H; Ducke, 1935a; Gleason, 1934; Oliver, 1876; Pittier, 1921) represent putative morphological synapomorphies of the Winged seed clade and useful diagnostic characters for identification purposes when combined with other traits. Species in the Winged seed clade also share staminal filaments that are completely connate bearing distal, sessile, ‘‘polythecate” anthers (i.e., with many sessile thecae on an elongated staminal tube), though this may be plesiomorphic for the Malvatheca clade (i.e., Malvoideae + Bombacoideae) (von Balthazar et al., 2006). Representatives of the Winged seed clade also have subaggregate wood parenchyma, with very narrow lines regularly alternating with 2–4 tangential rows of fibers that may extend from ray to ray, in cross section, mostly small flowers (ca. 2.5 cm long), and lobed calyces (Fig. S1, H; Cuatrecasas, 1950; Détienne et al., 1983; Gleason, 1934; Oliver, 1876; Pittier, 1914, 1921). 4.1.2. The Spongy endocarp clade This clade encompasses about 30 species of the mostly Amazonian genera Aguiaria, Catostemma, Cavanillesia, and Scleronema, together with eight or nine species of Adansonia, a genus native to the Paleotropics. The molecular data are ambiguous as to whether Cavanillesia or Adansonia are sister to the rest of the clade (Fig. S3). Due to this ambiguity, the polarity of seed number per fruit for the Spongy endocarp clade was unresolved. The four Neotropical genera form a morphologically cohesive group distinguished from other bombacoids by the relatively small flowers, 1– 2-ovulate locules, and widely spaced bands in wood parenchyma (Détienne et al., 1983; Metcalfe and Chalk, 1950; Paula, 1969, 1975; Record and Hess, 1949). Furthermore, whereas Adansonia has many seeds per fruit and the likely ancestral condition of palmately compound leaves, the Neotropical taxa have 1–4 seeds and simple (Cavanillesia) or 1-foliolate leaves (with the sole exception of Catostemma digitatum which has 3–5-foliolate leaves). The Spongy endocarp clade was recovered in previous phylogenetic assessment (Duarte et al., 2011) except for the inclusion of J.G. Carvalho-Sobrinho et al. / Molecular Phylogenetics and Evolution 101 (2016) 56–74 Aguiaria, which is here evaluated in phylogenetic studies for the first time. The morphological similarities of Aguiaria to Catostemma and Scleronema were first noted by Ducke (1937). Baum et al. (2004) speculated that Aguiaria should be phylogenetically close to these two genera, a view which is supported by the present study. Aguiaria, Catostemma, and Scleronema share the unifoliolate leaves. The latter two genera, however, are distinguished by having the 3-locular ovaries as a possible synapomorphy. These three genera are endemic to Amazonian wet forests whereas Cavanillesia, the other Neotropical genera, inhabits seasonally dry forests. Genera within the clade are rather variable in floral traits, and characterized by flowers from a few centimeters (Aguiaria, Catostemma, Cavanillesia, and Scleronema) to almost 30 cm long (Adansonia) and stamens ranging from 50 to more than one thousand. All species have lobed calyces, but this is a plesiomorphic trait for the core Bombacoideae inherited by the Winged seed clade rather than a synapomorphy for this clade (Fig. S1, G–H). Additional support for the monophyly of the spongy endocarp clade comes from fruit morphology (Figs. 3 and S2). At first glance, species in the Spongy endocarp clade have diverse fruit types (see Fig. S2, A–D). Fruits are woody and indehiscent in Adansonia and Scleronema (Baum, 1995; Bentham, 1862), 63 large samaras with five papyraceous wings in Cavanillesia (Ruiz and Pavon, 1797), and woody, tardily dehiscent capsules in Catostemma (Bentham, 1843; Shepherd and Alverson, 1981). In Aguiaria, the small fruits (less than 4 cm long) are unique among Angiosperms in having a coriaceous, dehiscent exocarp that splits off into five valves, all of which remain attached to the base of an indehiscent endocarp (see Fig. S2, C; see also line drawing in Ducke, 1935b). In spite of these differences, most taxa in this clade exhibit a spongy endocarp (Barroso et al., 1999, 2002; Baum, 1995; Ducke, 1935b; Kubitzki and Bayer, 2003; Paula, 1969; Schumann, 1886). Catostemma may be the exception, however: while fruit-derived fibers cover the seeds, they have been described as mucilaginous rather than spongy (Shepherd and Alverson, 1981). It may also be noteworthy that in Adansonia, Aguiaria, and Catostemma, the endocarp tends to adhere to the seed surface (see Fig. S2, A–C). We hypothesize that spongy endocarps constitute a morphological synapomorphy for this clade, as strongly supported by ancestral state reconstruction (Fig. S3). Developmental and anatomical studies of bombacoid fruits are needed to shed light on homologies of the endocarps among Spongy endocarp clade genera as well as to identify processes that may have generated the diversity of fruits in this group. Fig. 3. Ancestral character states reconstruction of endocarp type as shown on the BI majority rule consensus cladogram. Ancestral states reconstructions were performed with ML optimization as implemented in Mesquite v 3.02. Branch lengths are illustrated here without scale, for aesthetical purposes only. Colors represented in the nodes indicate the proportional likelihood of the states reconstructed for each node (see text). Numbers within circles indicate target nodes. Character transitions discussed in the text are indicated by arrows. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.) 64 J.G. Carvalho-Sobrinho et al. / Molecular Phylogenetics and Evolution 101 (2016) 56–74 Fig. 3 (continued) J.G. Carvalho-Sobrinho et al. / Molecular Phylogenetics and Evolution 101 (2016) 56–74 4.1.3. The Kapok clade This clade constitutes the largest lineage of the core Bombacoideae, including ca. 120 Neotropical species, except for Bombax and Rhodognaphalon, which are Paleotropical. Members of the clade have loculicidal capsules with endocarp modified into woolly tissue (‘‘kapok”) that surrounds non-winged, typically lightweight, and numerous seeds (see Fig. S2, H–K), though the kapok is reduced and seeds larger in a few, derived species. Fruits with kapok and truncate calyces (see Figs. 2, S1-I, S6) are putative synapomorphies for this clade as supported by our ancestral character state reconstructions (Figs. 3 and S6). The Kapok clade is composed of genera historically placed in Bombax s.l. (i.e., Bombax, Bombacopsis Pittier, Eriotheca, Pachira, Pseudobombax, and Rhodognaphalon, Rhodognaphalopsis A.Robyns) plus Ceiba and Spirotheca. In addition to kapok and truncate calyces, most species have seeds that are smooth (i.e., nonstriate), maculate or dotted, and occasionally have a raised hilum. A possible chemical synapomorphy for this clade is the presence of cyanidin-3,5-diglucoside (Paula et al., 1997; Refaat et al., 2013), but studies on several genera are lacking. The presence of prickles on trunks or branches is probably plesiomorphic for the Kapok clade with two subsequent losses, one in the Pachira s.l. clade and one in Pseudobombax (see Fig. S5). 65 The first cladogenesis within the Kapok clade separates the two Paleotropical genera, Bombax and Rhodognaphalon from the much more diverse Neotropical taxa in the Pachira s.l. and Striated bark subordinate clades. 4.1.3.1. The Paleotropical Bombax clade. This clade includes the Paleotropical genera Bombax and Rhodagnaphalon. Hutchinson (1967) argued in favor of placing Rhodognaphalon in synonymy with Pachira based on the shared presence of elongated flowers. Previous molecular analyses rejected this view but the phylogenetic placement of the genus remained uncertain (Duarte et al., 2011). Our combined analysis supports Rhodognaphalon as sister to Bombax, a relationship not previously hypothesized. All other performed analyses of individual or combined markers, however, were equivocal on the phylogenetic placement of Rhodognaphalon. The two genera in the Paleotropical Bombax clade share the presence of prickles on the trunks and/or branches (Robyns, 1963: 15). Although adults of Rhodognaphalon are often unarmed, prickled saplings have been reported (Voorhoeve, 1965: 72). The presence of coriaceous (vs. woody) valves, five-winged columellae that are persistent in fruit, and red petals are probable synapomorphies for this clade, though white or whitish petals have been reported for R. brevicuspe (Sprague) Roberty (Robyns, 1963; Fig. 4. Ancestral character states reconstruction of seed shape as shown on the BI majority rule consensus cladogram. Ancestral states reconstructions were performed with ML optimization as implemented in Mesquite v 3.02. Branch lengths are illustrated here without scale, for aesthetical purposes only. Colors represented in the nodes indicate the proportional likelihood of the states reconstructed for each node (see text). Numbers within circles indicate target nodes. The character transition to winged seeds is indicated by an arrow. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.) 66 J.G. Carvalho-Sobrinho et al. / Molecular Phylogenetics and Evolution 101 (2016) 56–74 Fig. 4 (continued) J.G. Carvalho-Sobrinho et al. / Molecular Phylogenetics and Evolution 101 (2016) 56–74 Voorhoeve, 1965: 71). According to Robyns (1963) and Voorhoeve (1965), Rhodognaphalon can be distinguished from Bombax mainly based on the oblong-linear, persistent calyces (vs. cupuliform and deciduous), the stamens organized in one whorl (vs. two), the reddish brown (vs. white) kapok, and the larger seeds, which are also fewer in number than in Bombax. Further work on the phylogeny and biogeography of Bombax + Rhodognaphalon is warranted. 4.1.3.2. The Pachira s.l. clade. This clade was recovered in most of our analyses and encompasses about 70 Neotropical species of Eriotheca and Pachira. The Pachira s.l. clade corresponds to the Pachira clade of Duarte et al. (2011) and includes various, previously recognized segregates and synonyms of Pachira: Bombacopsis, Pochota Ram.-Goyena, and Rhodognaphalopsis. Due to the lack of distinctive morphological characters, Bombacopsis and Rhodognaphalopsis have often been merged into other genera by recent workers. Steyemark and Stevens (1988) treated them as Pochota, distinguished from the two species they retained in Pachira s. str. by the latter’s very long flowers. However, more frequently Bombacopsis and Rhodognaphalopsis are treated as synonyms of Pachira (Alverson, 1994; Alverson and Mori, 2002; Alverson and Steyermark, 1997; Fernández-Alonso, 1998, 2003), as supported by our analyses. Pochota was re-established as a monotypic genus (Alverson and Duarte, 2015) based on previous molecular evidence (Duarte et al., 2011) and its phylogenetic and morphological affinities will be discussed below. Striate seeds have previously been identified as a possible synapomorphy for the Pachira s.l. clade (Duarte et al., 2011). Additionally, the alternate eophylls (i.e., first leaves produced by seedlings; Robyns, 1963: 15), the lack of prickles on trunks and branches, and leaflets with brochidodromous venation are possible synapomorphies for the Pachira s.l. clade (though brochidodromous venation is also present in Spirotheca). Taken together, the monophyly of the Pachira s.l. clade is well supported. Whereas Duarte et al. (2011) found support for trees in which Eriotheca is paraphyletic relative to Pachira, our data supported a monophyletic Eriotheca embedded in a paraphyletic Pachira. The prior result might be an artifact of its less dense taxon sampling, but further work is needed before concluding that either genus is non-monophyletic. We identified two clades within Pachira, one composed of Amazonian species and the other of extraAmazonian species, with the latter appearing more closely related to Eriotheca (see Fig. 2). In the extra-Amazonian Pachira lineage, P. endecaphylla is sister to the other species and is distinguished by numerous, small seeds surrounded by abundant kapok like in Eriotheca. The other four extra-Amazonian species, in contrast, have few, large seeds, and scarce kapok, similar to the Amazonian Pachira insignis. Within the Amazonian Pachira lineage, two sublineages were supported. The first (P. aquatica–P. mawarinumae) is composed of species inhabiting seasonally flooded (‘‘igapó”) forests with 7–11 leaflets (rarely 5), larger flowers and fruits (both to 30 cm long), and finely hairy exocarps. The second (P. gracilis–P. flaviflora) is composed of species from white-sand vegetation (‘‘campinas” and ‘‘campinaranas,” Anderson, 1981), characterized by a reduced number of leaflets (1–3, rarely 5), small, slender flowers (to 12 cm long), and smaller (to 10 cm long), often glabrous fruits. Eriotheca is a South American genus characterized by flowers that are smaller than in Pachira (typically < 4 cm), staminal tubes that are surmounted by entirely free filaments (vs. filaments organized in phalanges) (Robyns, 1963; Duarte et al., 2011), multiflorous cymes (vs. 1–2-flowered cymes), and more or less reniform anthers when dehisced (vs. oblong anthers). 4.1.3.3. The Striated bark clade. The genera Spirotheca, Neobuchia, Ceiba, Pochota fendleri, and Pseudobombax together form a well sup- 67 ported clade in the combined analysis. Spirotheca appears as sister to Bombax in the nuclear analysis, though this result is not strongly supported (Fig. S3). With exception of Pseudobombax, members of this clade have prickles on the trunk and/or branches. Besides the prickled trunks and/or branches, Spirotheca, Neobuchia, and Ceiba share a similar floral morphology in having just five or fifteen ‘‘filaments” (i.e., lobes at the apex of their staminal columns), stamens with appendages, and 2–4-thecate anthers (Gibbs and Alverson, 2006; Gibbs and Semir, 2003). However, it is not yet clear whether these shared floral traits are homologous. The plastid tree, which includes a Spirotheca–Neobuchia–Ceiba clade, supports the homology of these flowers, but the nuclear tree does not. We suspect that the plastid DNA, which has less homoplasy, reflects the true relationships, but analysis of further nuclear markers is needed. Except for Spirotheca, a genus of hemiepiphytes mostly found in wet upper-elevation forests, species in the Striated bark clade are predominantly deciduous while flowering, living in areas with seasonal climates. Trunks are characteristically swollen above or below the ground and display longitudinal striations that result from chlorophyll-rich underbark being exposed between vertical bands of the periderm, leaving the trunk with a marbled, gray/green aspect (see Fig. S1, A–E). While conspicuous striations can be absent in adults of Pochota fendleri and Spirotheca, they occur in juvenile plants. The eucamptodromous venation of Ceiba, Pochota fendleri, and Pseudobombax is likely apomorphic for these three genera and differs from the putatively plesiomorphic brochidodromous venation seen in Spirotheca and the Pachira s.l. clade. Our data support the merging of Ceiba and Chorisia Kunth as adopted in the last revision of Ceiba (Gibbs and Semir, 2003). The monospecific genus Neobuchia was supported as sister to Ceiba, as previously hypothesized by Duarte et al. (2011). Neobuchia paullinae Urb. is endemic to Haiti, and like many Ceiba species, has crenulate to serrate leaflets, deciduous calyces, and a similar androecium. Pochota fendleri has a distinct wood anatomy (Détienne et al., 1983) and our data strongly supported it as sister to Pseudobombax. This relationship has not previously been hypothesized but is supported by the putative synapomorphy of hippocrepiform anthers. Furthermore, members of the Pseudobombax–Pochota fendleri clade differ from its likely sister clade (Spirotheca–Neobuchia–Ceiba) by having calyces persistent in fruit, petals densely covered abaxially by stellate or tufted trichomes, appearing brownish to nigrescent (never white or red), 100–1500 fertile stamens with monothecate anthers, kapok brown (vs. white), fruit columella entire (vs. 5divided, see Fig. S2, K), and nectariferous glands on the receptacle (absent in Ceiba). Besides the presence of woody prickles on the trunks, Pochota fendleri can be distinguished from all species of Pseudobombax by its heavy wood and the coriaceous valves (vs. woody) that are persistent (vs. caducous) in dehisced fruits. The absence of prickles in Pseudobombax (Carvalho-Sobrinho et al., 2013) represents an unambiguous reversion of this character in the core Bombacoideae. Furthermore, Pseudobombax is characterized by leaflets with entire margins and manifest a distinctive synapomorphy of petioles that are distally dilated or expanded with leaflets having inarticulate petiolules (Carvalho-Sobrinho and Queiroz, 2011; CarvalhoSobrinho et al., 2014; Robyns, 1963). 4.2. A new tribal classification of the Bombacoideae Our data do not support the monophyly of Adansonieae sensu Hutchinson (1967), even including Ceiba as proposed by Bakhuizen van den Brink (1924), leaving most species without an obvious tribal affiliation. Therefore, we propose here a new tribal classification of Bombacoideae based on the combined tree derived 68 J.G. Carvalho-Sobrinho et al. / Molecular Phylogenetics and Evolution 101 (2016) 56–74 from the five nuclear and chloroplast markers (Fig. 2) and supplemented by morphological data. It seems logical to recognize the three major clades of core Bombacoideae at the tribal rank. The Winged seed clade does not match any previously-named tribe; we name it Bernoullieae as typified on the genus first described in the clade. The Spongy endocarp clade corresponds to a recircumscribed Adansonieae because it contains the type species of two previously defined tribes, Adansonieae and ‘Catostemmateae’, but the former was the only validly published. The Kapok clade, on the other hand, corresponds to the Takhtajan (1997) tribes ‘Ceibeae’ and Bombaceae (excluding Adansonia and Gyranthera, Fig. 1), but the latter is the sole name validly published at the tribal rank that includes Kapok clade genera. 4.3. Taxonomic treatment Tribe Adansonieae Horan., Char. Ess. Fam.: 192. 17 Jun 1847 (as ‘Adansoniae’).–T: Adansonia L. (1759). Important treatments of included taxa: Alverson (1994), Alverson and Mori (2002), Alverson and Steyermark (1997), Baum (1995), Ducke (1935a, 1935b, 1937, 1938), Paula (1969), Robyns (1964) and Steyermark (1987). Small to large unarmed trees, often with swollen trunks. Leaves simple and 3–5-nerved at base, or palmately compound, with pinnately-veined leaflets, sometimes reduced to one leaflet. Inflorescences axillary, terminal, or ramiflorous. Flowers usually small (c. 2.5 cm long), reaching c. 30 cm in length (Adansonia). Calyx lobed or laciniate, sometimes completely enclosing the corolla in mature buds, with lobes reflexed or coiled at the base of the flower at anthesis (Adansonia). Stamens usually 15–120 or numerous (Adansonia), filaments partially fused into a cylindrical staminal tube, often apically dilated. Ovary usually 2–9carpellate, ovules per locule few (1–2) to numerous (Adansonia). Anthers monothecate. Fruits samaras, woody berries, or tardily dehiscent capsules, endocarp spongy, rarely undifferentiated (Scleronema). Seeds usually few (1–4), or numerous (Adansonia), not winged. Genera included: Adansonia, Aguiaria, Catostemma, Cavanillesia, Scleronema. Recommended phylogenetic clade definition: The most inclusive clade containing Adansonia digitata L. but not Bernoullia flammea Oliv., Bombax ceiba L., Ceiba pentandra (L.) Gaertn., or Pachira aquatica Aubl. Tribe Bernoullieae Carv.-Sobr., tr. nov.–T: Bernoullia Oliv. (1876). Important treatments of included taxa: Alverson and Mori (2002), Cascante-Marin (1997), Pennington et al. (2004) and Robyns (1964). Diagnosis: Inflorescences scorpioid, fruits woody loculicidal capsules with papyraceous endocarps enclosing numerous winged seeds. Large unarmed trees, often buttressed. Leaves palmately compound, 3–5-foliolate, pinnately-veined, or 1-foliolate (appearing as simple leaves) and 3-veined at base. Inflorescences terminal, scorpioid. Flowers usually small (ca. 2.5 cm in length), but to ca. 15 cm long in Gyranthera. Calyx lobed. Stamens 5–20, filaments completely fused in a staminal tube except for short lobes, often laterally cleft on one side. Anthers ‘‘polythecate” (i.e., with many sessile thecae on an elongated staminal tube), sometimes spirally twisted. Ovary 5-carpellate, ovules ca. 10 per locule. Fruits large (to 30 cm long), woody loculicidal capsules with papyraceous endocarps. Seeds 10–30, large (ca. 6–9 cm long), winged. Genera included: Bernoullia, Gyranthera, Huberodendron. Recommended phylogenetic clade definition: The most inclusive clade containing Bernoullia flammea Oliv. but not Adansonia digitata L., Bombax ceiba L., Ceiba pentandra (L.) Gaertn., or Pachira aquatica Aubl. Tribe Bombaceae Kunth, Syn. Pl. 3: 258. 28 Feb 1824.–T: Bombax L., nom. et typ. cons. (1753). Important recent treatments: Alverson and Mori (2002), Alverson and Steyermark (1997), Fernández-Alonso (1998, 2001), Gibbs and Alverson (2006), Gibbs and Semir (2003) and Robyns (1963, 1964). Trees, rarely shrubs or stranglers (Spirotheca), trunks often swollen and prickled. Leaves palmately compound, 3–11-foliolate, rarely 1-foliolate, leaflets pinnately-veined. Inflorescences axillary or terminal, often reduced to solitary flowers. Flowers from 2.5 cm to 30 cm long. Calyx truncate, rarely lobed (Ceiba and Neobuchia). Stamens 5–1500, fused in a staminal tube, often organized in groups (phalanges). Anthers monothecate, bithecate (Ceiba and Neobuchia), or 4-thecate and spirally twisted (Spirotheca). Ovary 5–8-carpellate, ovules numerous per locule. Fruits loculicidal capsules with kapok. Columella persistent, entire or 5-fid. Seeds numerous, not winged, often maculate or striate, small, rarely 2–3 cm in length. Genera included: Bombax, Ceiba, Eriotheca, Neobuchia, Pachira, Pochota, Pseudobombax, Rhodognaphalon, Spirotheca. Recommended phylogenetic clade definition: The most inclusive clade containing Bombax ceiba L., Ceiba pentandra (L.) Gaertn., and Pachira aquatica Aubl. but not Adansonia digitata L., Bernoullia flammea Oliv., Bombax ceiba L., or Catostemma fragrans Benth. Key to the tribes of Bombacoideae (Malvaceae) 1. Inflorescences scorpioid cymes. Filaments fused into a staminal tube that is often laterally cleft on one side; anthers ‘‘polythecate” (i.e., with many sessile theceae on an elongated staminal tube), sometimes spirally twisted. Fruits loculicidal capsules with papyraceous endocarp. Seeds winged. Bernoullieae Carv.-Sobr., tr. nov. 1. Inflorescences cymose but not scorpioid. Filaments fused for one-third of their length or less, or if completely fused then never cleft. Anthers appearing 1–2-thecate, or if twisted then 4-thecate. Fruits woody berries, samaras, or loculicide capsules with endocarp spongy or modified into kapok. Seeds not winged. 2. Trees, never stranglers. Calyces lobed or laciniate, sometimes completely enclosing the corolla in mature buds. Fruits samaras, woody berries, or loculicidal capsules, with endocarps spongy, rarely undifferentiated, without kapok. Adansonieae Horan. (1847) 2. Trees, rarely shrubs or stranglers. Calyces truncate, if lobed then stamens 5( 15), bithecate. Fruits loculicidal capsules with endocarp modified as kapok. Bombaceae Kunth (1824) Acknowledgments Thanks are due to Charles Zartman, Christine Bacon, Domingos Cardoso, Marlon Machado, and Paulo Kaminski who provided plant material for this study, and Daiane Trabuco, Elvia Souza, and Luane do Carmo for assistance in the molecular laboratory. We thank the directors and staff of the molecular laboratory in UEFS (LAMOL) J.G. Carvalho-Sobrinho et al. / Molecular Phylogenetics and Evolution 101 (2016) 56–74 and the directors, curators, and staff of the herbaria who loaned specimens, especially Cláudia Gonçalves, Marc Pignal, and Jacques Florence (P), Hans-Joachim Esser (M), Martin Cheek and Cátia Canteiro (K), Lorenzo Ramella and Nicolas Fumeaux (G). Jim Reveal provided valuable advice on nomenclatural issues. We also thank the Fundação de Amparo à Pesquisa do Estado da Bahia – FAPESB (process APP0006/2011), the Conselho Nacional de Desenvolvimento Científico e Tecnológico – CNPq (processes 300811/2010-1 and 563546/2010-7-REFLORA), the Programa de Apoio a Núcleos de Excelência (PRONEX, process PNX0014/2009), and the Sistema Nacional de Pesquisa em Biodiversidade (SISBIOTA CNPq 563084/2010-3/FAPESB PES0053/2011) for financial support. This paper is part of the PhD thesis of JGCS prepared in the Programa de Pós-Graduação em Botânica (PPGBot–UEFS) and supported by a grant from the Coordenação de Aperfeiçoamento de Pessoal de Nível Superior – CAPES. JGCS also thanks CNPq (process 158916/2014-0), the Reflora programme (process BEX 5415/13-6) for a sandwich fellowship, and the Department of Botany of the Smithsonian Institution, Washington, D.C., for a José Cuatrecasas Fellowship. Partial funding for DAB was provided by NSF Grant DEB-1354793. Appendix A. Origin and voucher specimens for DNA sequences used in the study. Numbers preceded by KM or KR refer to newly generated sequences. ‘‘.... = missing sequences Taxon, collection locality, voucher collector and number, (Herbarium acronyms [following the Index Herbariorum (http://sweetgum.nybg.org/ih/]), GenBank accession numbers: nrETS, nrITS, matK, trnL-trnF, or trnS-trnG. Adansonia digitata L., Pacific Tropical Garden acc. no. 770032002, KM453023, HQ658372⁄, AY321168⁄, HQ696738⁄, ....; Adansonia gregorii F.Muell., Wendel s.n. (ISC), ...., HQ658374⁄, HQ696688⁄, HQ696740⁄, KM453112; Adansonia grandidieri Baill., D.A.Baum 345 (MO), ...., HQ658373⁄, HQ696687⁄, HQ696739⁄, ....; Adansonia kilima Pettigrew, K.L. Bell, Bhagw., Grinan, Jillani, Jean Mey., Wabuyele & C.E. Vickers, J. Pettigrew 402, . . .., JN400327⁄, . . .., JN400300⁄, . . ..; Adansonia madagascariensis Baill., accession A, . . .., AF028532⁄, . . .., JN300407⁄, . . ..; Adansonia perrieri Capuron, isolate 208, . . .., AF028538⁄, . . .., JN400292⁄, . . ..; Adansonia rubrostipa Jum. & H.Perrier, . . .., AF028531⁄, . . .., . . .., . . ..; Adansonia suazerensis H. Perrier ...., AF028529⁄, . . .., . . .., . . ..; Adansonia za Baill., D.A.Baum 357 (MO), ...., AF028536⁄, HQ696689⁄, HQ696741⁄, ....; Aguiaria excelsa Ducke, D.Cardoso 3343 (HUEFS), KM453024, KM453161, ...., ...., KM453109; Bernoullia flammea Oliv., T.S.Cochrane s.n. (WIS), KM453027, HQ658366⁄, HQ696685⁄, HQ696732⁄, ....; Bombax anceps Pierre, KYUM<174>, . . .., . . .., AB924835⁄, . . .., . . ..; Bombax buonopozense P.Beauv., Pac. Trop. Bot. Gard. acc. no. 770474001, KM453025, HQ658376⁄, AY321171⁄, HQ696742⁄, ....; Bombax ceiba L., J.G.Carvalho-Sobrinho 3073 (HUEFS), KM453026, KM453163, ...., ...., KM453109, W.S.Alverson s.n. (WIS), ...., ...., HQ696690⁄, HQ696743⁄, ....; Catostemma albuquerquei Paula, J.G.CarvalhoSobrinho 3117 (HUEFS), KM453028, KM453165, ...., ...., KM453111; Catostemma fragrans Benth., W.S.Alverson 4030 (WIS), KM453029, HQ658370⁄, AY589069⁄, HQ696736⁄, KM453121; Catostemma milanezii Paula, J.G.Carvalho-Sobrinho 3116 (HUEFS), KM453030, KM453166, ...., ...., ....; Cavanillesia chicamochae Fern. Alonso, D. Castellanos 83, KM245242, ...., ...., KM488630, ....; Cavanillesia platanifolia (Bonpl.) Kunth., Fairchild Botanical Gardens acc. no. FG83343A, KM453031, HQ658371⁄, HQ696686⁄, HQ696737⁄, ....; Cavanillesia umbellata Ruiz & Pav., J.G.CarvalhoSobrinho 2987 (HUEFS), KM453032, ...., ...., ...., ....; Ceiba acuminata (S.Watson) Rose, Fairchild Botanical Gardens acc. no. X-2–206, ...., HQ658385⁄, HQ696700⁄, HQ696752⁄, ....; Ceiba aesculifolia (Kunth) 69 Britten & Baker.f., Fairchild Botanical Gardens acc. no. 83301, KM453033, HQ658384⁄, HQ696699⁄, HQ696751⁄, . . ..; Ceiba chodatii (Hassl.) Ravenna, G.Schmeda 1170 (US), KM453034, ...., ...., ...., ....; Ceiba crispiflora (Kunth) Ravenna, Pacific Tropical Garden acc. no. 750726001, KM453035, HQ658387⁄, AY321169⁄, HQ696754⁄, ....; Ceiba erianthos (Cav.) K.Schum., E.R.Souza 710 (HUEFS), KM453036, KM453167, ...., ...., KM453113; Ceiba glaziovii (Kuntze) K.Schum., J.G.Carvalho-Sobrinho 2967 (HUEFS), KM453037, ...., ...., ...., KM453116; Ceiba insignis (Kunth) P.E.Gibbs & Semir, J.Campos & P.López 4953 (US), KM453038, KM488629, ...., ...., ....; Ceiba jasminodora (A.St.-Hil.) K.Schum., J.G.Carvalho-Sobrinho 3070 (HUEFS), KM453039, KM453168, ...., ...., KM453131; Ceiba pentandra (L.) Gaertn., J.G.Carvalho-Sobrinho s.n. (HUEFS), KM453040, KM453169, ...., ...., KM453117, W.S.Alverson s.n. (WIS), ...., HQ658386⁄, HQ696701⁄, HQ696753⁄, ....; Ceiba pubiflora (A.St.Hil.) K.Schum., J.G.Carvalho-Sobrinho 3066 (HUEFS), KM453041, KM453170, ...., ...., KM453118; Ceiba rubriflora Carv.-Sobr. & L.P. Queiroz, J.G.Carvalho-Sobrinho 574 (HUEFS), KM453042, KM453171, ...., ...., KM453119; Ceiba samauma (Mart.) K.Schum., J. G.Carvalho-Sobrinho s.n. (HUEFS), KM453043, ...., ...., ...., ....; Ceiba schottii Britten & Baker.f., Fairchild Botanical Gardens acc. no. 83302, ...., HQ658389⁄, HQ696703⁄, HQ696756⁄, ....; Ceiba speciosa (A.St.-Hil.) Ravenna, W.S.Alverson s.n. (WIS), KM453044, HQ658388⁄, HQ696702⁄, HQ696755⁄, ....; Ceiba ventricosa (Nees & Mart.) Ravenna, J.G.Carvalho-Sobrinho 3124 (HUEFS), KM453045, KM453172, ...., ...., ....; Chiranthodendron pentadactylon Larreat., Wendt s.n. (WIS), KM453046, HQ658356⁄, AY321164⁄, HQ696722⁄, ....; Eriotheca bahiensis M.C. Duarte & G.L. Esteves, M. C.Duarte 89 (SPF), ...., HQ658398⁄, HQ696712⁄, HQ696766⁄, ....; Eriotheca candolleana (K. Schum.) A.Robyns, M.C.Duarte 99 (CVRD), ...., HQ658394⁄, HQ696718⁄, HQ696772⁄, ....; J.G.Carvalho-Sobrinho 3137 (HUEFS), ...., ...., ...., ...., KM453120; Eriotheca crenulaticalyx A. Robyns, L.P.Queiroz s.n. (HUEFS), KM453047, KM453173, ...., ...., KM453122; Eriotheca discolor (Kunth) A.Robyns, Campo 6110 (MO), ...., . . .., HQ696775, HQ696720, . . ..; Eriotheca dolichopoda A. Robyns, M.C.Duarte 92 (CEPEC), ...., HQ658402⁄, HQ696719⁄, HQ696773⁄, ....; J.G.Carvalho-Sobrinho 3123 (HUEFS), KM453048, KM453174, ...., ...., KM453123; Eriotheca estevesiae Carv.-Sobr., G. Pereira-Silva 5392 (HUEFS), KM488628, KM283224, ...., ...., KM453160; Eriotheca globosa (Aubl.) A.Robyns, R.O.Perdiz 745 (CEPEC), KM453049, ...., ...., ...., ....; Eriotheca gracilipes (K.Schum.) A.Robyns, J.G.Carvalho-Sobrinho 3037 (HUEFS), KM453050, ...., ...., ...., KM453124; M.C.Duarte 120 (SP), ...., ...., HQ696708⁄, HQ696762⁄, ....; Eriotheca longipedicellata (Ducke) A.Robyns, M.C. Duarte 93 (IAN, SP), ...., ...., HQ696716⁄, HQ696770⁄, ....; Eriotheca longitubulosa A.Robyns, M.C.Duarte 96 (SP), ...., ...., HQ696717⁄, HQ696771⁄, ....; Eriotheca macrophylla (K. Schum.) A.Robyns, M.C. Duarte 106 (SP), ...., ...., HQ696713⁄, HQ696767⁄, KM453108; J.G. Carvalho-Sobrinho 2949 (HUEFS), KM453051, KM453175, ...., ...., KM453125; Eriotheca obcordata A.Robyns, B.M.Silva 107 (HUEFS), ...., HQ658403⁄, ...., HQ696774⁄, ....; Eriotheca parvifolia (Mart.) A. Robyns, J.G.Carvalho-Sobrinho 2870 (HUEFS), KM453053, ...., ...., ...., KM453126; M.C.Duarte 109 (SP), ...., HQ658401⁄, HQ696710⁄, HQ696764⁄, ....; Eriotheca pentaphylla (Vell. emend. K. Schum.) A. Robyns, M.C.Duarte 75 (SP), . . .., . . .., HQ696768⁄, HQ696714⁄, . . ..; Eriotheca pubescens (Mart.) A.Robyns, J.G.Carvalho-Sobrinho 2873 (HUEFS), ...., ...., ...., ...., KM453127; J.G.Carvalho-Sobrinho 2878 (HUEFS), KM453054, ...., ...., ...., KM488631; M.C.Duarte 115 (SP), ...., HQ658397⁄, HQ696709⁄, HQ696763⁄, ....; Eriotheca roseorum (Cuatrec.) A.Robyns, Fuentes 1167 (MO), . . .., . . .., HQ696765⁄, HQ696711⁄, . . ..; Eriotheca ruizii (K.Schum.) A.Robyns, P.M.Peterson 9487 (US), . . .., . . .., HQ696777⁄, HQ696721⁄, . . ..; Eriotheca saxicola Carv.-Sobr., J.G.Carvalho-Sobrinho 3165 (HUEFS), KM453055, ...., ...., ...., ....; J.G.Carvalho-Sobrinho 3167 (HUEFS), ...., KM453176, ...., ...., ....; J.G.Carvalho-Sobrinho 3146 (HUEFS), ...., ...., ...., ...., KM453129; 70 J.G. Carvalho-Sobrinho et al. / Molecular Phylogenetics and Evolution 101 (2016) 56–74 Eriotheca squamigera (Cuatrec.) Fern.Alonso., Neill 12522,. . .., . . .., . . .., HQ696776⁄, ....; Eriotheca surinamensis (Uittien) A.Robyns, M. C.Duarte 97 (SP), ...., HQ658400⁄, HQ696715⁄, HQ696769⁄, ....; Eriotheca sp., J.G.Carvalho-Sobrinho 3125 (HUEFS), KM453052, KR076544, . . .., . . .., . . ..; Fremontodendron californicum (Torr.) Coville, Ex. Rancho Santo Ana Bot. Garden, Prop. no. 5996, Herb. no. 12343, KM453056, HQ658357⁄, AY321165⁄, HQ696723⁄, KM453114; Gyranthera caribensis Pittier, Iltis et al. s.n (WIS), ...., HQ658368⁄, AY589071⁄, HQ696734⁄, KM453110; Hampea appendiculata Standl., W.S.Alverson 2179 (WIS). . .., U56781, AY589062, . . .., . . ..; Huberodendron patinoi Cuatrec., W.S.Alverson 2201 (WIS), KM453057, HQ658367⁄, AY589072⁄, HQ696733⁄, ....; Huberodendron swietenioides (Gleason) Ducke, J.G.Carvalho-Sobrinho 3292 (HUEFS), ...., KM453162, ...., ...., KM453129; Neobuchia paulinae Urb., Cult. Jardin Botanico, Santo Domingo, Dominican Republic, . . .., . . .., HQ696707⁄, HQ696760⁄, . . ..; Ochroma pyramidale (Cav. ex Lam.) Urb., J.G.Carvalho-Sobrinho 3077 (HUEFS), KM453058, ...., ...., ...., KM453132; W.S. Alverson s.n. (WIS), ...., HQ658363⁄, AY321172⁄, HQ696729⁄, ....; Pachira aquatica Aubl., J.G.CarvalhoSobrinho s.n. (HUEFS), KM453059, ...., ...., ...., KM453133; W.S. Alverson s.n. (WIS), ...., ...., AY321170⁄, HQ696759⁄, ....; Pachira brevipes (A.Robyns) W.S.Alverson, J.G.Carvalho-Sobrinho 3097 (HUEFS), KM453060, ...., ...., ...., KM453139; P.Fine 1060 (UC), ...., HQ658391⁄, HQ696694⁄, ...., ....; Pachira endecaphylla (Vell.) A. Robyns, J.G.Carvalho-Sobrinho 3130 (HUEFS), KM453061, KM453182, ...., ...., KM453134; Pachira faroensis (Ducke) W.S.Alverson, D.Cardoso 3418 (HUEFS), KM453068, KM453179, ...., ...., ....; Pachira flaviflora (Pulle) Fern.Alonso, P.Fine 1062 (UC), ...., HQ658379⁄, HQ696693⁄, HQ696746⁄, ....; Pachira glabra Pasq., M. C.Duarte 70 (SP), ...., HQ658393⁄, HQ696706⁄, HQ696761⁄, ....; J.G. Carvalho-Sobrinho 2863 (HUEFS), KM453062, KM453177, ...., ...., KM453135; Pachira gracilis (A.Robyns) W.S.Alverson, T.D.M.Barbosa 1296 (INPA), KM453063, ...., ...., ...., ....; Pachira humilis Spruce ex Benth., D.Cardoso 3413 (HUEFS), KM453067, KM453178, ...., ...., ....; Pachira insignis (Sw.) Sw. ex Savigny, J.G.Carvalho-Sobrinho 3106 (HUEFS), KM453064, ...., ...., ...., KM453136; P.Fine 1061 (UC), ...., HQ658390⁄, HQ696704⁄, HQ696757⁄, ....; Pachira mawarinumae (Steyerm.) W.S.Alverson, D.Cardoso 3419 (HUEFS), KM453069, KM453180, ...., ...., ....; Pachira minor (Sims) Hemsl., G. Davidse 4901...., ...., HQ696705⁄, HQ696758⁄, ....; Pachira moreirae Carv.Sobr. & W.S.Alverson, J.G.Carvalho-Sobrinho 2963 (HUEFS), KM453065, KF477294, ...., ...., KM453138; Pochota fendleri (Seem.) W.S. Alverson & M.C.Duarte [ Pachira quinata], W.S.Alverson 2174 (WIS), ...., ...., HQ696692⁄, HQ696745⁄, ....; P.E.Kaminski s.n. (HUEFS), KM453074, KM453184, ...., ...., KM453141; Pachira retusa (Mart.) Fern.Alonso, M.V.Moraes 532 (HUEFS), KM453066, KF477293, ...., ...., KM453140; Pachira aff. mawarinumae (Steyerm.) W.S.Alverson, D.Cardoso 3425 (HUEFS), KM453070, KM453181, ...., ...., ....; Pachira sp., J.G.Carvalho-Sobrinho 3176 (HUEFS), KM453071, ...., ...., ...., ....; Patinoa sphaerocarpa Cuatrec., W.S.Alverson s.n. (WIS), ...., HQ658364⁄, AY589074⁄, HQ696730⁄, ....; Pentaplaris doroteae L. O.Williams & Standl., B.E.Hammel 18736 (MO), ...., HQ658358⁄, AY321163⁄, HQ696724⁄, KM453115; Phragmotheca ecuadorensis W.S.Alverson, W.S.Alverson 2223 (WIS), ...., ...., AY589068⁄, ...., ....; Pseudobombax amapaense A.Robyns, J.G.Carvalho-Sobrinho 3105 (HUEFS), KM453092, KM453200, ...., ...., KM453154; Pseudobombax andicola A.Robyns, W.J.Eyerdam 25320 (F), KM453077, ...., ...., ...., ....; C.Antezana 1122 (NY), ...., KM453186, ...., ...., KM453142; Pseudobombax argentinum (R.E.Fries) A.Robyns, C.Saravia-Toledo 11476 (LPB), KM453078, ...., ...., ...., ....; Pseudobombax cajamarcanus Fern. Alonso, C.Díaz 2189 (MO), KM453079, ...., ...., ...., ....; Pseudobombax calcicola Carv.-Sobr. & L.P.Queiroz, J.G.Carvalho-Sobrinho 2993 (HUEFS), KM453080, KM453187, ...., ...., KM453144; Pseudobombax campestre (Mart.) A.Robyns, J.G.Carvalho-Sobrinho 2872 (HUEFS), KM453081, KM453188, ...., ...., KM453145; Pseudobombax croizatii A.Robyns, Oldham s.n. (WIS), ...., HQ658382⁄, HQ696697⁄, HQ696749⁄, ....; Pseudobombax ellipticoideum A.Robyns, M.J.Balick 3704 (MO), KM453084, ...., ...., ...., ....; M.G.Aguilar 3621 (MO), ...., ...., ...., ...., ....; Pseudobombax ellipticum (Kunth) Dugand, J.G. Carvalho-Sobrinho 3131 (HUEFS), KM453083, KM453190, ...., ...., KM488632; Fairchild Botanical Gardens acc. no. FG X.1-101, ...., KM453189, ...., ...., KM453146; Pseudobombax grandiflorum (Cav.) A.Robyns, J.G.Carvalho-Sobrinho 3198 (HUEFS), KM453085, ...., ...., ...., KM453148, Fairchild Botanical Gardens acc. no. FG-65-35, ...., ...., HQ696698⁄, HQ696750⁄, ...., J.G.Carvalho-Sobrinho 2946 (HUEFS), ...., KM453191, ...., ...., ....; Pseudobombax grandiflorum var. majus A.Robyns, J.G.Carvalho-Sobrinho 3069 (HUEFS), KM453088, KM453195, ...., ...., KM453150; Pseudobombax guayasense A.Robyns, T.D.Pennington 14519 (K), KM453086, . . ., . . .., . . .., . . ..; Pseudobombax longiflorum (Mart.) A.Robyns, J.G.Carvalho-Sobrinho 2880 (HUEFS), ...., ...., ...., ...., ...., J.G.Carvalho-Sobrinho 2875 (HUEFS), KM453087, KM453194, ...., ...., ...., J.G.Carvalho-Sobrinho 2882 (HUEFS), ...., ...., ...., KM453149; Pseudobombax marginatum (A.St.Hil., Juss. & Cambess.) A.Robyns, R.Small s.n. (ISC), ...., ...., HQ696696⁄, HQ696748⁄, KM453147, L.P.Queiroz 14753 (HUEFS), KM453089, KM453197, ...., ...., KM453151; Pseudobombax millei A. Robyns, J.G.Carvalho-Sobrinho s.n. (HUEFS), KM453090, KM453198, ...., ...., ....; Pseudobombax mininum Carv.-Sobr. & L.P. Queiroz, J.G.Carvalho-Sobrinho 2887 (HUEFS), KM453091, KM453199, ...., ...., KM453152; Pseudobombax munguba (Mart.) Dugand, J.G.Carvalho-Sobrinho 3105 (HUEFS), KM453092, KM453200, ...., ...., KM453153; Pseudobombax parvifoliumCarv.Sobr. & L.P.Queiroz, J.G.Carvalho-Sobrinho 3029 (HUEFS), KM453094, KM453201, ...., ...., KM453154; Pseudobombax petropolitanum A.Robyns, J.G.Carvalho-Sobrinho 3071 (HUEFS), KM453096, ...., ...., ...., KM453155, J.G.Carvalho-Sobrinho 3171 (HUEFS), ...., ...., ...., ...., ...., J.G.Carvalho-Sobrinho 3173 (HUEFS), ...., KM453203, ...., ...., ....; Pseudobombax pulchellum Carv.-Sobr., A.H.Gentry 75227 (MO), KM453097, ...., ...., ...., ....; Pseudobombax septenatum (Jacq.) Dugand, E.Villanueva 835 (LPB), KM453098, ...., ...., ...., ....; Pseudobombax simplicifolium A.Robyns, J.G.Carvalho-Sobrinho 3027 (HUEFS), KM453099, ...., ...., ...., KM453156, J.G.Carvalho-Sobrinho 3062 (HUEFS), KM453100, KM453204, ...., ...., ....; Pseudobombax aff. campestre (Mart.) A.Robyns, J.G.Carvalho-Sobrinho 3080 (HUEFS), KM453076, KM453205, ...., ...., KM453143; Pseudobombax tomentosum (Mart.) A.Robyns, J.G.Carvalho-Sobrinho 2874 (HUEFS), KM453101, KM453206, ...., ...., ....; J.G.Carvalho-Sobrinho 2879 (HUEFS), KM488627, KM453207, ...., ...., KM453157; Rhodognaphalon schumannianum A.Robyns, M.W.Chase 5973 (K), ...., HQ658380⁄, HQ696695⁄, HQ696747⁄, ....; Scleronema micranthum (Ducke) Ducke, J.G.Carvalho-Sobrinho 3098 (HUEFS), KM453102, ...., ...., ...., KM453158; W.S.Alverson s.n. (WIS), ...., HQ658369⁄, AY589070⁄, HQ696735⁄, ....; Scleronema praecox (Ducke) Ducke, J. G.Carvalho-Sobrinho 3276 (HUEFS), KM453103, ...., ...., ...., KM453159; Septotheca tessmannii Ulbr., J.Rios 1917 (MO), KM453104, HQ658365⁄, AY589073⁄, HQ696731⁄, ....; Spirotheca elegans Carv.-Sobr., M.C.Machado & L.P.Queiroz, J.G.CarvalhoSobrinho 2964 (HUEFS), KM453105, KM453208, ...., ...., ....; Spirotheca rivierii (Decne.) Ulbr., J.G.Carvalho-Sobrinho s.n. (HUEFS), KM453106, KM453209, ...., ...., ....; Spirotheca rosea (Seem.) P.E. Gibbs & W.S.Alverson, W.S.Alverson 2185 (WIS), KM453107, HQ658378⁄, HQ696691⁄, HQ696744⁄, .....; Sterculia lanceolata Cav., YCC635, ...., AF460184⁄, HQ415311⁄, AY328151⁄, ....; Sterculia nobilis Sm., YCC632, ...., AF460183⁄, ...., JN676078⁄, .... 71 J.G. Carvalho-Sobrinho et al. / Molecular Phylogenetics and Evolution 101 (2016) 56–74 Appendix B. Matrix used in the reconstruction of ancestral states of morphological characters of Bombacoideae (Malvaceae) using a maximum likelihood approach Adansonia digitata Adansonia grandidieri Adansonia gregorii Adansonia kilima Adansonia madagascariensis Adansonia perrieri Adansonia rubrostipa Adansonia suazerensis Adansonia za Aguiaria excelsa Bombax anceps Bombax buonopozense Bombax ceiba Bernoullia flammea Catostemma albuquerquei Catostemma fragrans Catostemma milanezii Cavanillesia chicamochae Cavanillesia platanifolia Cavanillesia umbellata Ceiba acuminata Ceiba aesculifolia Ceiba chodatii Ceiba crispiflora Ceiba erianthos Ceiba glaziovii Ceiba insignis Ceiba jasminodora Ceiba pentandra Ceiba pubiflora Ceiba rubriflora Ceiba samauma Ceiba schottii Ceiba speciosa Ceiba ventricosa Eriotheca bahiensis Eriotheca candolleana Eriotheca crenulaticalyx Eriotheca discolor Eriotheca dolichopoda Eriotheca estevesiae Eriotheca globosa Eriotheca gracilipes Eriotheca longipedicellata Eriotheca longitubulosa Eriotheca macrophylla Eriotheca obcordata Eriotheca parvifolia Eriotheca pentaphylla Eriotheca pubescens Eriotheca roseorum Eriotheca ruizii Eriotheca saxicola Eriotheca sp. CS3125 Eriotheca squamigera Eriotheca surinamensis Gyranthera caribensis Huberodendron patinoi Huberodendron swietenioides Prickles on trunk or branches Calyx shape Endocarp type Seed shape Seed number per fruit 0 0 0 0 0 0 0 0 0 0 1 1 1 0 0 0 0 0 0 0 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 2 2 2 2 2 2 2 2 2 0 1 1 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 1 1 1 1 1 1 1 1 1 1 1 0 1 1 1 1 1 1 1 0 0 0 1 1 1 1 1 1 1 1 1 1 2 2 2 0 1 1 1 1 1 1 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 1 1 2 2 2 2 2 2 2 2 2 0 2 2 2 1 0 0 0 0 0 0 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 1 1 1 (continued on next page) 72 J.G. Carvalho-Sobrinho et al. / Molecular Phylogenetics and Evolution 101 (2016) 56–74 Appendix B (continued) Neobuchia paullinae Pachira aff. mawarinumae Pachira aquatica Pachira brevipes Pachira endecaphylla Pachira faroensis Pachira flaviflora Pachira glabra Pachira gracilis Pachira humilis Pachira insignis Pachira mawarinumae Pachira minor Pachira moreirae Pachira retusa Pachira sp. CA85 Pachira sp. CS3176 Pochota fendleri Pseudobombax aff. campestre Pseudobombax amapaense Pseudobombax andicola Pseudobombax argentinum Pseudobombax cajamarcanus Pseudobombax calcicola Pseudobombax campestre Pseudobombax sp. CB305 Pseudobombax croizatii Pseudobombax ellipticum Pseudobombax ellipsticoideum Pseudobombax grandiflorum Pseudobombax guayasense Pseudobombax longiflorum Pseudobombax grandiflorum var. majus Pseudobombax marginatum Pseudobombax millei Pseudobombax minimum Pseudobombax munguba Pseudobombax parvifolium Pseudobombax petropolitanum Pseudobombax pulchellum Pseudobombax septenatum Pseudobombax simplicifolium Pseudobombax tomentosum Rhodognaphalon schumannianum Scleronema micranthum Scleronema praecox Spirotheca elegans Spirotheca rivieri Spirotheca rosea Prickles on trunk or branches Calyx shape Endocarp type Seed shape Seed number per fruit 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 2 2 1 2 2 2 2 1 2 2 1 2 2 1 1 2 1 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 0 0 0 0 0 0 0 0 0 0 1 0 0 1 1 1 1 1 1 1 1 1 1 1 1 1 1 0 0 1 1 1 2 2 2 2 2 2 2 2 2 2 2 3 3 2 2 2 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 2 2 2 2 2 2 2 2 2 2 2 0 0 2 2 2 Appendix C. Supplementary material Supplementary data associated with this article can be found, in the online version, at http://dx.doi.org/10.1016/j.ympev.2016.05.006. References Alverson, W.S., 1994. New species and combinations of Catostemma and Pachira (Bombacaceae) from the Venezuelan Guayana. Novon 4, 3–8. Alverson, W.S., Duarte, M.C., 2015. Hello again Pochota, farewell Bombacopsis. Novon 24, 115–119. Alverson, W.S., Mori, S.A., 2002. Bombacaceae. In: Alverson, W.S., Cremers, G., Gracie, C.A., de Granville, J.-J., Heald, S.V., Hoff, M., Mitchell, J.D. (Eds.), Guide to the Vascular Plants of Central French Guiana. Mem. New York Bot. Gard., vol. 76, pp. 139–145. Alverson, W.S., Steyermark, J.A., 1997. Bombacaceae. In: Berry, P.E., Holst, B.K., Yatskievych, K. (Eds.), Flora of the Venezuelan Guayana, Araliaceae–Cactaceae, vol. 3. Missouri Botanical Garden, St. Louis, USA, pp. 496–527. Alverson, W.S., Whitlock, B.A., Nyffeler, R., Bayer, C., Baum, D.A., 1999. Phylogeny of the core Malvales: evidence from ndhF sequence data. Am. J. Bot. 86, 1474– 1486. J.G. Carvalho-Sobrinho et al. / Molecular Phylogenetics and Evolution 101 (2016) 56–74 Andel, T.V., 2001. Floristic composition and diversity of mixed primary and secondary forests in northwest Guyana. Biodivers. Conserv. 10, 1645–1682. Anderson, A.B., 1981. White-Sand vegetation of Brazilian Amazonia. Biotropica 13, 199–210. Ariati, S.R., Murphy, D.J., Udovicic, F., Ladiges, P.Y., 2006. Molecular phylogeny of three groups of acacias (Acacia subgenus Phyllodineae) in arid Australia based on the internal and external transcribed spacer regions of nrDNA. Syst. Biodivers. 4, 417–426. http://dx.doi.org/10.1017/S1477200006001952. Bailey, C.D., Carr, T.G., Harris, S.A., Hughes, C.E., 2003. Characterization of angiosperm nrDNA polymorphism, paralogy, and pseudogenes. Mol. Phylogenet. Evol. 29, 435–455. http://dx.doi.org/10.1016/j.ympev.2003.08.021. Bakhuizen Van Den Brink, R.C., 1924. Revisio Bombacacearum. Bull. Jard. Bot. Buitenzorg 6, 1–240. Baldwin, B.G., Markos, S., 1998. Phylogenetic utility of the external transcribed spacer (ETS) of 18S–26S rDNA: congruence of ETS and ITS trees of Calycadenia (Compositae). Mol. Phylogenet. Evol. 10, 449–463. Barroso, G.M., Morim, M.P., Peixoto, A.L., Ichaso, C.L.F., 1999. Frutos e sementes: morfologia aplicada à sistemática de dicotiledôneas. Ed. UFV, Viçosa, Brasil. Barroso, G.M., Peixoto, A.L., Ichaso, C.L.F., Guimarães, E.F., Costa, C.G., 2002. Sistemática de Angiospermas do Brasil, vol. 1, segunda edição. Ed. UFV, Viçosa, Brasil. Baum, D.A., 1995. A systematic revision of Adansonia (Bombacaceae). Ann. Missouri Bot. Gard. 82, 440–471. Baum, D.A., Smith, S.D., Yen, A., Alverson, W.S., Nyffeler, R., Whitlock, B.A., Oldham, R.L., 2004. Phylogenetic relationships of Malvatheca (Bombacoideae and Malvoideae; Malvaceae sensu lato) as inferred from plastid DNA sequences. Am. J. Bot. 91, 1863–1871. http://dx.doi.org/10.1016/j.ode.2004.08.001. Bayer, C., Fay, M., De Bruijn, A., Savolainen, V., Morton, C., Kubitzki, K., Alverson, W. S., Chase, M.W., 1999. Support for an expanded family concept of Malvaceae within a recircumscribed order Malvales: a combined analysis of plastid atpb and rbcL DNA sequences. Bot. J. Linn. Soc. 129, 267–303. Benson, D.A., Karsch-Mizrachi, I., Lipman, D.J., Ostell, J., Sayers, E.W., 2010. GenBank. Nucl. Acids Res. 39 (database issue), D32–D37. http://dx.doi.org/10.1093/nar/ gkq1079. Bentham, G., 1843. Contributions towards a flora of South America – enumeration of plants collected by Mr. Schomburgk in British Guiana. London J. Bot. 2, 359–378. Bentham, G., 1862. Notes on Malvaceae and Sterculiaceae. J. Proc. Linn. Soc. 6, 97– 122. Cardoso, D., Lima, H.C., Rodrigues, R.S., Queiroz, L.P., Pennington, R.T., Lavin, M., 2012. The realignment of Acosmium sensu stricto with the Dalbergioid clade (Leguminosae, Papilionoideae) reveals a proneness for independent evolution of radial floral symmetry among early branching papilionoid legumes. Taxon 61, 1057–1073. Cardoso, D., Queiroz, L.P., Lima, H.C., Suganuma, E., van den Berg, C., Lavin, M., 2013. A molecular phylogeny of the vataireoid legumes underscores floral evolvability that is general to many early-branching papilionoid lineages. Am. J. Bot. 100, 403–421. Carvalho-Sobrinho, J.G., Alverson, W.S., Mota, A.C., Machado, M.C., Baum, D.A., 2014. A new deciduous species of Pachira (Malvaceae: Bombacoideae) from a seasonally dry tropical forest in northeastern Brazil. Syst. Bot. 39, 260–267. http://dx.doi.org/10.1600/036364414X678224. Carvalho-Sobrinho, J.G., Queiroz, L.P., 2011. Morphological cladistic analysis of Pseudobombax Dugand (Malvaceae, Bombacoideae) and allied genera. Rev. Bras. Bot. 34, 197–209. http://dx.doi.org/10.1590/S0100-84042011000200007. Carvalho-Sobrinho, J.G., Queiroz, L.P., Dorr, L.J., 2013. Does Pseudobombax have prickles? Assessing the enigmatic species Pseudobombax endecaphyllum (Malvaceae: Bombacoideae). Taxon 62, 814–818. http://dx.doi.org/10.12705/ 624.30. Carvalho-Sobrinho, J.G., Santos, F.A.R., Queiroz, L.P., 2009. Morfologia dos tricomas das pétalas de espécies de Pseudobombax Dugand (Malvaceae, Bombacoideae) e seu significado taxonômico. Acta Bot. Bras. 23, 929–934. http://dx.doi.org/ 10.1590/S0102-33062009000400003. Cascante-Marin, A., 1997. La familia Bombacaceae (Malvales) en Costa Rica. Brenesia 47, 17–36. Cho, S., Zwick, A., Regier, J.C., Mitter, C., Cummings, M.P., Yao, J., Du, Z., Zhao, H., Kawahara, A.Y., Weller, S., Davis, D.R., Baixeras, J., Brown, J.W., Parr, C., 2011. Can deliberately incomplete gene sample augmentation improve a phylogeny estimate for the advanced moths and butterflies (Hexapoda: Lepidoptera). Syst. Biol. 60, 782–796. Cuatrecasas, J., 1950. Contributions to the flora of South America: Studies in South American Plants – II. Publ. Field. Mus. Nat. Hist. Bot. Ser. 27, 87–93. Cuatrecasas, J., 1953. Um nouveau genre de Bombacées, Patinoa. Rev. Int. Bot. Appl. Agric. Trop. 33, 306–313. Cuatrecasas, J., 1954a. Novelties in the Bombacaceae. Phytologia 4, 465–480. Cuatrecasas, J., 1954b. Disertaciones sobre Bombaceas. Rev. Acad. Colomb. Ci. Exact. 9, 64–177. Desfeux, C., Lejeune, B., 1996. Systematics of Euromediterranean Silene (Caryophyllaceae): evidence from a phylogenetic analysis using ITS sequences. Compt. Rend. Acad. Sci. Paris, Sér. 3, Sci. Vie 319, 351–358. Détienne, P., Loureiro, A.A., Jacquet, P., 1983. Estudo anatômico do lenho da família Bombacaceae da América. Acta Amazon. 13, 831–867. Dick, C.W., Eldredge, B., Lemes, M., R. Gribel, R., 2007. Extreme long-distance dispersal of the lowland tropical rainforest tree Ceiba pentandra L. (Malvaceae) in Africa and the Neotropics. Mol. Ecol. 16, 3039–3049. http://dx.doi.org/ 10.1111/j.1365-294X.2007.03341.x. 73 Duarte, M.C., Esteves, G.L., Salatino, M.L.F., Walsh, K.C., Baum, D.A., 2011. Phylogenetic analyses of Eriotheca and related genera (Bombacoideae, http://dx.doi.org/10.1600/ Malvaceae). Syst. Bot. 36, 690–701. 036364411X583655. Ducke, A., 1935a. Plantes nouvelles ou peu connues de la Région Amazonienne (X série). Arq. Inst. Biol. Veg. 2, 59–73. Ducke, A., 1935b. Aguiaria, novo gênero de Bombacáceas, a árvore maior do Alto Rio Negro. Anais Acad. Bras. Ci. 7, 329–332. Ducke, A., 1937. New forest trees of the Brazilian Amazon. Trop. Woods 50, 37–39. Ducke, A., 1938. Aguiaria, novo gênero de Bombacáceas, a árvore maior do Alto Rio Negro. Anais Acad. Bras. Ci. 10, 11–14. Dugand, A., 1943. Revalidacion de Bombax Ceiba L. como especie tipica del genero Bombax L. y descripcion de Pseudobombax gen. nov. Caldasia 2, 47–68. Edlin, H.L., 1935. A critical revision of certain taxonomic groups of the Malvales. New Phytol. 34, 122–143. Farris, J.S., Källersjö, M., Kluge, A.G., Bult, C., 1995. Testing significance of incongruence. Cladistics 10, 315–319. Felsenstein, J., 1985. Confidence limits on phylogenies: an approach using the bootstrap. Evolution 39, 783–791. Fernández-Alonso, J.L., 1998. Novedades taxonômicas, corológicas y nomenclaturales em el gênero Pachira Aubl. (Bombacaceae). Anales Jard. Bot. Madrid 52, 305–314. Fernández-Alonso, J.L., 2001. Bombacaceae neotropicae novae vel minus cognitae V. Novedades en Pseudobombax Dugand y sinopsis de las especies Colombianas. Rev. Acad. Colomb. Ci. Exact. 25, 467–476. Fernández-Alonso, J.L., 2003. Bombacaceae neotropicae novae vel minus cognitae VI. Novedades en los géneros Cavanillesia, Eriotheca, Matisia y Pachira. Rev. Acad. Colomb. Ci. Exact. 27, 25–37. Ferreira, L.V., Prance, G.T., 1998. Species richness and floristic composition in four hectares in the Jaú National Park in upland forests in Central Amazonia. Biodivers. Conserv. 7, 1349–1364. Gibbs, P.E., Alverson, W.S., 2006. How many species of Spirotheca (Malvaceae s.l., Bombacoideae)? Brittonia 58, 245–258. http://dx.doi.org/10.1663/0007-196X. Gibbs, P.E., Semir, J., 2003. A taxonomic revision of the genus Ceiba Mill. (Bombacaceae). Anales Jard. Bot. Madrid 60, 259–300. http://dx.doi.org/ 10.3989/ajbm.2002.v60.i2.92. Gibbs, P.E., Semir, J., da Cruz, N.D., 1988. A proposal to unite the genera Chorisia Kunth with Ceiba Miller (Bombacaceae). Notes Roy. Bot. Gard. Edinburgh 45, 125–136. Gleason, H.A., 1934. Plantae Krukovianae III. Phytologia 1, 106–111. Goujon, M., McWilliam, H., Li, W., Valentin, F., Squizzato, S., Paern, J., Lopez, R., 2010. A new bioinformatics analysis tools framework at EMBL-EBI. Nucl. Acids Res. 38 (Web Server issue), W695–W699. http://dx.doi.org/10.1093/nar/gkq313. Hamilton, M.B., 1999. Four primer pairs for the amplification of chloroplast intergenic regions with intraspecific variation. Mol. Ecol. 8, 521–523. Horan, P.F., 1847. Adansonieae Horan. Char. Ess. Fam., 142 Hutchinson, J., 1967. The Genera of Flowering Plants. Dicotyledones, vol. 2. Clarendon Press, Oxford, UK. Jiang, W., Chen, S.Y., Wang, H., Li, D.Z., Wiens, J.J., 2014. Should genes with missing data be excluded from phylogenetic analyses? Mol. Phylogenet. Evol. 80, 308– 318. Kubitzki, K., Bayer, C., 2003. Bombacoideae. In: Kubitzki, K., Bayer, C. (Eds.), Flowering Plants, Dicotyledons: Malvales, Capparales, and Non-betalain Caryophyllales. Springer-Verlag, Berlin, Heidelberg, New York, pp. 271–277. Kunth, C.S., 1824. Bombaceae kunth. Synopsis Plantarum, vol. 3, p. 258. Linares-Palomino, R.L., Alvarez, S.I.P., 2005. Tree community patterns in seasonally dry tropical forests in the Cerros de Amotape Cordillera, Tumbes, Peru. Forest Ecol. Manage. 209, 261–272. http://dx.doi.org/10.1016/j.foreco.2005.02.003. Liu, K., Warnow, T.J., Holder, M.T., Nelesen, S., Yu, J., Stamatakis, A., Linder, C.R., 2012. SATé-II: very fast and accurate simultaneous estimation of multiple sequence alignments and phylogenetic trees. Syst. Biol. 61, 90–106. http://dx. doi.org/10.1093/sysbio/syr095. Lohmann, L.G., Taylor, C.M., 2014. A new generic classification of tribe Bignonieae (Bignoniaceae). Ann. Missouri Bot. Gard. 99, 348–489. Maddison, W.P., Maddison, D.R., 2010. Mesquite: A Modular System for Evolutionary Analysis, Version 2.72 http://mesquiteproject.org. Metcalfe, C.R., Chalk, L., 1950. Anatomy of Dicotyledons: Leaves, Stem, and Wood in Relation to Taxonomy with Notes on Economic Uses, vol. 2. Clarendon Press, Oxford, UK. Miller, M.A., Pfeiffer, W., Schwartz, T., 2010. Creating the CIPRES Science Gateway for inference of large phylogenetic trees. In: Proceedings of the Gateway Computing Environments Workshop (GCE), 14 November 2010, New Orleans, Louisiana, pp. 1–8 http://www.phylo.org/index.php/portal/. Nyffeler, R., Bayer, C., Alverson, W.S., Yen, A., Whitlock, B., Chase, M.W., Baum, D.A., 2005. Phylogenetic analysis of the Malvadendrina clade (Malvaceae s.l.) based on plastid DNA sequences. Org. Divers. Evol. 5, 109–123. http://dx.doi.org/ 10.1016/j.ode.2004.08.001. Nylander, J.A.A., Ronquist, F., Huelsenbeck, J.P., Nieves-Aldrey, J.L., 2004. Bayesian phylogenetic analysis of combined data. Syst. Biol. 53, 47–67. http://dx.doi.org/ 10.1080/10635150490264699. Oliver, D., 1876. Bernoullia Oliv., gen. nov. In: Hooker, J.D. (Ed.), Hooker’s Icone Plantarum, third series, vol. 12. Longman, Rees, Orme, Brown, Green and London, UK, pp. 62–63, pl. 1169–1170. Pagel, M., 1999. The maximum likelihood approach to reconstructing ancestral character states of discrete characters on phylogenies. Syst. Biol. 48, 612–622. 74 J.G. Carvalho-Sobrinho et al. / Molecular Phylogenetics and Evolution 101 (2016) 56–74 Paula, J.E.P., 1969. Estudos sobre Bombacaceae – I. Contribuição para o conhecimento dos gêneros Catostemma Benth. e Scleronema Benth. da Amazônia Brasileira. Ci. Cult. 21, 697–705. Paula, J.E.P., 1975. Estudos sobre Bombacaceae V – Investigação anatômica das madeiras de Catostemma commune Sandwith, Catostemma sclerophyllum Ducke e Scleronema micranthum (Ducke) Ducke, com vistas à polpa, papel e taxinomia. Acta Amazon. 6, 155–161. Paula, V.F., Barbosa, L.C.A., Demuner, A.J., Piló-Veloso, D., 1997. A química da família bombacaceae. Quím. Nova 20, 627–630. http://dx.doi.org/10.1590/S010040421997000600010. Pennington, T.D., Reynel, C., Daza, A., 2004. Illustrated Guide to the Trees of Peru. David Hunt, Sherborne, England, 848p. Pennington, T.D., Sarukhán, J., 1968. Manual para la identificación de campo de los principales arboles tropicales de Mexico. Instituto Nacional de Investigaciones Forestales, Organización de la Naciones Unidas para la Agricultura y la Alimentación, 523p. Pittier, H., 1914. Gyranthera Pittier, gen. nov. Bombacacearum. Repert. Spec. Nov. Regni Veg. 13, 318–319. Pittier, H., 1916. Bombacaceae. In: New or Noteworthy Plants from Colombia and Central America – 5. Contr. U.S. Natl. Herb., vol. 18, pp. 159–163. Pittier, H., 1921. II – Acerca del genero Gyranthera Pittier. Contribuiciones para la flora de Venezuela. Typografia Americana, Caracas, Venezuela. Prance, G.T., Rodrigues, W.A., Silva, M.F., 1976. Inventário florestal de um hectare de mata de terra firme km 30 da estrada Manaus-Itacoatiara. Acta Amazon. 6, 9–35. Rambaut, A., Suchard, M.A., Xie, D., Drummond, A.J., 2014. Tracer v1.6 http://beast. bio.ed.ac.uk/Tracer. Ravenna, P., 1998. On the identity, validity, and actual placement in Ceiba of several Chorisia species (Bombacaceae), and description of two new South American species. Onira 3, 42–51. Record, S.J., Hess, R.W., 1949. Timbers of the New World. Yale University Press, New Haven, Connecticut, USA. Refaat, J., Desoky, S.Y., Ramadan, M.A., Kamel, M.S., 2013. Bombacaceae: a phytochemical review. Pharm. Biol. 51, 100–130. http://dx.doi.org/10.3109/ 13880209.2012.698286. Robyns, A., 1963. Essai de Monographie du genre Bombax L. s.l. (Bombacaceae). Bull. Jard. Bot. État. Bruxelles 33, 1–315. Robyns, A., 1964. Flora of Panama. Part IV. Bombacaceae. Ann. Missouri Bot. Gard. 51, 37–68. Ronquist, F., Teslenko, M., Van der Mark, P., Ayres, D.L., Darling, A., Höhna, S., Larget, B., Liu, L., Suchard, M.A., Huelsenbeck, J.P., 2012. MrBayes 3.2: efficient Bayesian phylogenetic inference and model choice across a large model space. Syst. Biol. 61, 539–542. http://dx.doi.org/10.1093/sysbio/sys029. Ruiz, H., Pavon, J., 1797. Cavanillesia. In: Florae Peruvianae et Chilensis Prodromus, editio secunda. Typographio Palearniano, Rome, Italy, pp. 85–86. Schluter, D., Price, T., Mooers, A.O., Ludwig, D., 1997. Likelihood of ancestor states in adaptive radiation. Evolution 51, 1699–1711. Schumann, K., 1886. Bombacaceae. In: Martius, K.F.P., Eichler, A.G., Urban, I. (Eds.), Flora Brasiliensis, vol. 12. Munich & Leipzig, pp. 201–250, tab. 40–50 http:// florabrasiliensis.cria.org.br/search?taxon_id=744. Schumann, K., 1895. Bombacaceae. In: Engler, A., Prantl, K. (Eds.), Die natürlichen Pflanzenfamilien, part. 4, div. 6. Verlag von Wilhelm Engelmann, Leipzig, pp. 53–68. Shepherd, J.D., Alverson, W.S., 1981. A new Catostemma (Bombacaceae) from Colombia. Brittonia 33, 587–590. Souza, E.R., Lewis, G.P., Forest, F., Schnadelbach, A.S., Van den Berg, C., Queiroz, L.P., 2014. Phylogeny of Calliandra (Leguminosae: Mimosoideae) based on nuclear and plastid molecular markers. Taxon 62, 1200–1219. http://dx.doi.org/ 10.12705/626.2. Staden, R., 1996. The Staden sequence analysis package. Mol. Biotechnol. 5, 233– 241. Stamatakis, A., 2006. RAxML-VI-HPC: Maximum Likelihood-based phylogenetic analyses with thousands of taxa and mixed models. Bioinformatics 22, 2688– 2690. http://dx.doi.org/10.1093/bioinformatics/btl446. Stamatakis, A., 2014. RAxML Version 8: a tool for phylogenetic analysis and postanalysis of large phylogenies. Bioinformatics 30, 1312–1313. http://dx.doi.org/ 10.1093/bioinformatics/btu033. Steyermark, J.A., 1987. Notes on Catostemma and Scleronema (Bombacaceae). Ann. Missouri Bot. Gard. 74, 636–646. Steyemark, J.A., Stevens, W.D., 1988. Notes on Rhodognaphalopsis and Bombacopsis (Bombacaceae) in the Guayanas. Ann. Missouri Bot. Gard. 75, 396–398. Sukumaran, J., Holder, M.T., 2010. DendroPy: a Python library for phylogenetic computing. Bioinformatics 26, 1569–1571. http://dx.doi.org/10.1093/ bioinformatics/btq228. Sun, Y., Skinner, D.Z., Liang, G.H., Hulbert, S.H., 1994. Phylogenetic analysis of Sorghum and related taxa using internal transcribed spacer ribosomal DNA. Monogr. Theor. Appl. Genet. 89, 26–32. Swofford, D.L., 2002. PAUP⁄: Phylogenetic Analysis Using Parsimony (⁄and other methods), Version 4.0b10. Sinauer, Sunderland, Massachusetts, USA. Takhtajan, A., 1997. Diversity and Classification of Flowering Plants. Columbia University Press, New York, NY. Ulbrich, E., 1914. Bombacaceae. In: R. Pilger (Ed.), Plantae Uleanae novae vel minus cognitae. Notizbl. Bot. Gart. Berlin-Dahlem, vol. 6, pp. 156–166. Von Balthazar, M., Schonenberger, J., Alverson, W.S., Janka, H., Bayer, C., Baum, D.A., 2006. Structure and evolution of the androecium in the Malvatheca clade (Malvaceae s. l.) and implications for Malvaceae and Malvales. Plant Syst. Evol. 260, 171–197. http://dx.doi.org/10.1007/s00606-006-0442-9. Voorhoeve, A.G., 1965. Liberian High Forest Trees. Centrum voor landbouwpublikaties en landbouwdocumentatie, Wageningen, Germany. White, T.J., Bruns, T., Lee, S., Taylor, J., 1990. Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. In: Innis, M., Gelfand, D., Snisnsky, J.T., White, T. (Eds.), PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, USA, pp. 315–322.

RELATED PAPERS

RELATED TOPICS

Log In

Revisiting the phylogeny of Bombacoideae (Malvaceae): Novel relationships, morphologically cohesive clades, and a new tribal classification based on multilocus phylogenetic analyses

Revisiting the phylogeny of Bombacoideae (Malvaceae): Novel relationships, morphologically cohesive clades, and a new tribal classification based on multilocus phylogenetic analyses

Related Papers

RELATED PAPERS

RELATED TOPICS