**2.** *Begonia* **classification**

Morphological and molecular data firmly place the Begoniaceae within the order Cucurbitales, which also includes the large and economically important family Cucurbitaceae (950-980 species), and the small families Anisophylleaceae (29-34 species), Apodanthaceae (19 species), Coriariaceae (15-20 species), Corynocarpaceae (5-6 species), Datiscaceae (2 species) and Tetramelaceae (2 species) (APG, 2009; Matthews & Endress, 2004; Schaefer & Renner, 2011; Zhang et al., 2006). Analyses of DNA sequence data from 14

2005; Neale et al., 2006). The distribution of *Begonia* diversity is uneven throughout tropical regions, with the greatest diversity in America and Asia (>600 species each), whilst being relatively species poor in Africa (160 species) and absent in Australia (Goodall-Copestake et al., 2010). The genus is thought to have originated in Africa, while South American and South East Asian species are the results of parallel radiations over the last 20 - 50 million years (Goodall Copestake et al., 2010; Plana et al., 2004; Thomas et al., 2011). Long distance dispersal is rare, for example *Begonia* species have failed to cross

Begoniaceae are easily recognizable by diagnostic characters such as asymmetrical leaves, unisexual monoecious flowers, twisted-, papillose stigmas, and dry-, three-winged capsules (Doorenbos, 1998). However, there are numerous deviations from these typical character states. Within the genus *Begonia* there is a large range of morphological diversity, particularly in vegetative form, and this is linked to adaptation to a variety of ecological conditions. Vegetative adaptations such as the evolution of perennating rhizomes, leaf micromorphology optimised for low, scattered light; or stomatal clustering may underlie their ability to thrive in diverse niches. Phenotypic polymorphism within populations occurs, most frequently in the anthocyanin patterns on the leaves, which although striking have not been shown to have measurable effects on light capture (Hughes et al., 2008). The genetic and morphological diversity of the genus *Begonia* has been exploited through cultivation to produce over 10,000 cultivars. These are horticulturally divided into 5 classes: a) the tuberous begonias (*B*. x *tuberhybrida*), a complex group derived from crosses between species such as *B. boliviensis* or *B. pearcei*, b) Elatior begonias (*B*. x *hiemalis*), a cross between tuberous begonias and *B. socotrana*, c) Lorraine begonias (*B.* x *cheimanta*), a cross between tuberous hybrids and *B. dregei*, d) semperflorens begonias (*B. semperflorens*-cultorum), with *B. cucullata* and *B. schmidtiana* as important ancestors, and e) begonias grown for their ornamental foliage (*B*. *rex*-cultorum), Asiatic in origin (Haegeman, 1979; Hvoslef-Eide et al., 2007). Commercial interest in this group has promoted research into a variety of topics,

Phylogenetic and cytological research in the last decade has significantly increased our knowledge of diversity within the genus *Begonia*. In this review, the current classification and the evolution of species diversity is discussed with reference to recent progress in: a) population genetic and phylogenetic techniques using genetic markers in association with morphological characters and b) cytological techniques such as mitotic or meiotic chromosome visualisation, linked to genome size studies. We also present new data on barriers to hybridisation between *Begonia* species. We focus on how genetic, cytological and local ecological effects may contribute to diversity in this genus, particularly the evolution of

Morphological and molecular data firmly place the Begoniaceae within the order Cucurbitales, which also includes the large and economically important family Cucurbitaceae (950-980 species), and the small families Anisophylleaceae (29-34 species), Apodanthaceae (19 species), Coriariaceae (15-20 species), Corynocarpaceae (5-6 species), Datiscaceae (2 species) and Tetramelaceae (2 species) (APG, 2009; Matthews & Endress, 2004; Schaefer & Renner, 2011; Zhang et al., 2006). Analyses of DNA sequence data from 14

the Torres strait from Papua New Guinea to Australia.

including hybridisation and polyploidy.

species diversity.

**2.** *Begonia* **classification** 

nuclear, mitochondrial and plastid markers strongly support a close relationship of Begoniaceae with Datiscaceae and Tetramelaceae (Schaefer & Renner, 2011).

Only two genera are currently recognized in the Begoniaceae: the monotypic genus *Hillebrandia*, and the species-rich and morphologically diverse genus *Begonia* (Doorenbos et al., 1998; Forrest & Hollingsworth, 2003). A third genus, *Symbegonia*, was previously included in Begoniaceae, and separated from *Begonia* by floral characters (syntepalous perianth and a monadelphous androecium). Based on molecular data, the genus *Symbegonia* has been shown to be nested within *Begonia* section *Petermannia* (Forrest & Hollingsworth, 2003). *Hillebrandia sandwicensis*, which is endemic to Hawaii, can be differentiated from *Begonia* by a suite of morphological characters. These include more differentiated segments of the perianth, semi-inferior ovaries (inferior in *Begonia*), and fruit dehiscence between the styles in contrast to the usually loculicidal dehiscence in *Begonia* (Clement et al., 2004; Forrest & Hollingsworth, 2003; Forrest et al., 2005).

A reliable infrageneric classification and subdivision of large genera such as *Begonia* is crucial in order to inform taxonomic monographs, biogeographic and evolutionary studies. A revision of circumscriptions of *Begonia* sections by Doorenbos et al. (1998) provides a foundation for the subdivision of the genus. In this revision 63 sections were recognized, and another three sections have been subsequently proposed (de Wilde & Plana, 2003; Forrest & Hollingsworth, 2003; Shui et al., 2002). The distributions of all but one of the currently accepted *Begonia* sections are limited to single continental regions, i.e. Africa, Asia, or America. Only section *Tetraphila* can be found in multiple continents and a single, recently discovered, and still to be named species in this predominantly African section is indigenous to continental Southeast Asia (de Wilde, 2011).

DNA sequence data from non-coding regions plays an important role in plant classification and barcoding (CBOL Plant working group, 2009), and has widely been used to resolve relationships at the species and sectional level in *Begonia*. A framework phylogeny of *Begonia* based on analyses of c. 13000 bases of plastid and mitochondrial DNA of 30 *Begonia* species (Goodall-Copestake et al., 2010; Fig. 1) indicates that African taxa form the earliest divergent clades in the *Begonia* phylogeny and that both Asian and American *Begonia* lineages are derived from African ancestors. The phylogenetic relationships within the relatively small group of African *Begonia*, which comprises around 160 species subdivided into 17 sections (de Wilde & Plana, 2003; Doorenbos et al., 1998), are relatively well understood. African *Begonia* species are not retrieved as monophyletic, but South African species placed in section *Augustia* were shown to be closely related to a clade of American taxa, and Socotran *Begonia* species (section *Peltaugustia*) were shown to form a monophyletic clade with Asian taxa (Forrest et al., 2005; Goodall-Copestake et al., 2010; Plana et al., 2004; Thomas et al., 2011). Revisions exist for the majority of the African sections (see references in Plana, 2003), and the intersectional relationships of African *Begonia* species have been studied using molecular systematic approaches and have been discussed in some detail in Plana (2003) and Plana et al. (2004). Most African sections are well circumscribed and seem to represent monophyletic taxa, but section *Mezierea* is polyphyletic (Forrest et al., 2005; Plana, 2003; Plana et al., 2004); and the lack of resolution or support in phylogenies makes the assessment of the monophyly of some sections problematic. Apart from the grade of continental African taxa, a major Madagascan radiation can be detected. Only one of the c. 50 Madagascan (incl. Comores and Mascarenes) *Begonia* species, *Begonia oxyloba*, is also widespread on the African continent (Keraudren-Aymonin, 1983). The other Madagascan species seem to be the result of a single dispersal event from continental Africa and a subsequent radiation (Plana, 2003; Plana et al., 2004).

The Origin of Diversity in *Begonia*:

Thomas, 2010).

(Thomas, 2010; Thomas et al., 2011).

**3. Historical biogeography** 

delimitations and investigate intersectional relationships.

Genome Dynamism, Population Processes and Phylogenetic Patterns 31

Asian and Socotran *Begonia* species form a well supported clade in recent analyses of nuclear, mitochondrial and plastid DNA sequence data (Goodall-Copestake et al., 2010; Thomas, 2010; Thomas et al., 201)). The c. 750 Asian *Begonia* species have been divided into 20 sections (Doorenbos et al., 1998; Forrest & Hollingsworth, 2003; Shui et al., 2002; Thomas et al., 2011). However, the circumscriptions of several of these sections are questionable and phylogenetic analyses of sequence data provide evidence for the non-monophyly of most species-rich Asian sections (Forrest & Hollingsworth, 2003; Tebbitt et al., 2006; Thomas, 2010; Thomas et al., 2011). Moreover, several small or monotypic sections were shown to be nested within clades of species assigned to the larger, species-rich sections. For example, section *Baryandra* is nested within Philippine *Diploclinum* (Rubite, 2010; Thomas, 2010); sections *Alicida* and *Putzeysia* fall into a grade of continental Asian species predominantly assigned to section *Diploclinium* (Rajbhandary, 2010; Thomas, 2010; Thomas et al., 2011); and section *Monopteron* is nested within section *Platycentrum* (Forrest, 2001; Rajbhandary, 2010;

Within Asian *Begonia* two well supported major clades can be differentiated based on molecular data. The first is dominated by continental Asian taxa, and mainly includes species of section *Parvibegonia*, continental Asian species of the polyphyletic section *Diploclinium*, and species in sections *Platycentrum* and *Sphenanthera*. The second major clade contains the predominantly Chinese section *Coelocentrum* that forms the sister clade to a clade of Malesian species placed in sections *Ridleyella*, *Bracteibegonia*, *Petermannia*, and Malesian species placed in the polyphyletic sections *Diploclinium* and *Reichenheimia* 

American *Begonia* comprise more than 600 species divided into 29 sections (Burt-Utley, 1985; Doorenbos et al., 1998). American *Begonia* species are retrieved as monophyletic in analyses of nuclear ribosomal and mitochondrial DNA sequence data, but form two distinct clades with African sister groups in analyses of plastid DNA sequence data (Forrest & Hollingsworth, 2003; Goodall-Copestake et al., 2010). This phylogenetic incongruence may indicate an ancient hybridisation event in this group (Goodall-Copestake et al., 2010). In contrast to African and Asian *Begonia*, the phylogenetics of American *Begonia* are still poorly understood, and the circumscriptions of several neotropical sections are highly problematic. Doorenbos et al. (1998: 181) emphasized that several neotropical sections "shade off into each other," i.e. sectional delimitations often lack autapomorphic characters and morphologically transitional species are present. Low-density taxon samples of 9-11 American species were included in the analyses by Clement et al. (2004), Forrest & Hollingsworth (2003), Forrest et al. (2005) and Goodall-Copestake et al. (2010), but further studies including a much wider and geographically robust sampling of American *Begonia* species are needed to identify major subdivisions within American *Begonia*, clarify sectional

Molecular divergence age estimates based on DNA sequence data and the calibration of molecular changes with a dimension of time, allow inference of: i) the time of origin of monophyletic groups, and ii) the timing of dispersal to a geographic area (Renner, 2005). However, molecular divergence age estimates for clades of Begoniaceae is problematic, as the family has a poor fossil record, and suitable fossils for direct calibration are lacking

Fig. 1. Cladograms showing relationships supported by ≥ 50% parsimony bootstrap and ≥ 50% Bayesian posterior probability support after analysis of the genome-level DNA sequence datasets with indel data (Goodall-Copestake et al., 2010). The geographic origin of taxon samples is given in brackets and the African, American, and Asian continental species assemblages are indicated by pale, mid, and dark grey shading, respectively. Seasonallyadapted African *Begonia* species are indicated by asterisks.

Fig. 1. Cladograms showing relationships supported by ≥ 50% parsimony bootstrap and ≥ 50% Bayesian posterior probability support after analysis of the genome-level DNA sequence datasets with indel data (Goodall-Copestake et al., 2010). The geographic origin of taxon samples is given in brackets and the African, American, and Asian continental species assemblages are indicated by pale, mid, and dark grey shading, respectively. Seasonally-

adapted African *Begonia* species are indicated by asterisks.

Asian and Socotran *Begonia* species form a well supported clade in recent analyses of nuclear, mitochondrial and plastid DNA sequence data (Goodall-Copestake et al., 2010; Thomas, 2010; Thomas et al., 201)). The c. 750 Asian *Begonia* species have been divided into 20 sections (Doorenbos et al., 1998; Forrest & Hollingsworth, 2003; Shui et al., 2002; Thomas et al., 2011). However, the circumscriptions of several of these sections are questionable and phylogenetic analyses of sequence data provide evidence for the non-monophyly of most species-rich Asian sections (Forrest & Hollingsworth, 2003; Tebbitt et al., 2006; Thomas, 2010; Thomas et al., 2011). Moreover, several small or monotypic sections were shown to be nested within clades of species assigned to the larger, species-rich sections. For example, section *Baryandra* is nested within Philippine *Diploclinum* (Rubite, 2010; Thomas, 2010); sections *Alicida* and *Putzeysia* fall into a grade of continental Asian species predominantly assigned to section *Diploclinium* (Rajbhandary, 2010; Thomas, 2010; Thomas et al., 2011); and section *Monopteron* is nested within section *Platycentrum* (Forrest, 2001; Rajbhandary, 2010; Thomas, 2010).

Within Asian *Begonia* two well supported major clades can be differentiated based on molecular data. The first is dominated by continental Asian taxa, and mainly includes species of section *Parvibegonia*, continental Asian species of the polyphyletic section *Diploclinium*, and species in sections *Platycentrum* and *Sphenanthera*. The second major clade contains the predominantly Chinese section *Coelocentrum* that forms the sister clade to a clade of Malesian species placed in sections *Ridleyella*, *Bracteibegonia*, *Petermannia*, and Malesian species placed in the polyphyletic sections *Diploclinium* and *Reichenheimia*  (Thomas, 2010; Thomas et al., 2011).

American *Begonia* comprise more than 600 species divided into 29 sections (Burt-Utley, 1985; Doorenbos et al., 1998). American *Begonia* species are retrieved as monophyletic in analyses of nuclear ribosomal and mitochondrial DNA sequence data, but form two distinct clades with African sister groups in analyses of plastid DNA sequence data (Forrest & Hollingsworth, 2003; Goodall-Copestake et al., 2010). This phylogenetic incongruence may indicate an ancient hybridisation event in this group (Goodall-Copestake et al., 2010). In contrast to African and Asian *Begonia*, the phylogenetics of American *Begonia* are still poorly understood, and the circumscriptions of several neotropical sections are highly problematic. Doorenbos et al. (1998: 181) emphasized that several neotropical sections "shade off into each other," i.e. sectional delimitations often lack autapomorphic characters and morphologically transitional species are present. Low-density taxon samples of 9-11 American species were included in the analyses by Clement et al. (2004), Forrest & Hollingsworth (2003), Forrest et al. (2005) and Goodall-Copestake et al. (2010), but further studies including a much wider and geographically robust sampling of American *Begonia* species are needed to identify major subdivisions within American *Begonia*, clarify sectional delimitations and investigate intersectional relationships.
