South African Journal of Botany 88 (2013) 10–16 Contents lists available at ScienceDirect South African Journal of Botany journal homepage: www.elsevier.com/locate/sajb Bertilia — A new monotypic genus in the Senecioneae (Asteraceae) from South Africa☆ G.V. Cron ⁎ School of Animal, Plant & Environmental Sciences, University of the Witwatersrand, Private Bag 3, WITS 2050, South Africa article info abstract Article history: The unusual disjunct distribution of the yellow-flowered, radiate, Northern Cape endemic Emilia hantamensis Received 5 March 2013 J.C.Manning & Goldblatt, relative to the other Emilia species in southern Africa, prompted investigation into Received in revised form 18 April 2013 its phylogenetic position and relationships. Phylogenies based on the ITS and trnL–trnF regions reveal that Accepted 24 April 2013 it is not a member of the genus Emilia but belongs in a well-supported clade with Bolandia, a genus compris- Available online 5 June 2013 ing five species mainly from the Western and Northern Cape, and the monotypic genera Stilpnogyne and Edited by JC Manning Mesogramma. It is placed sister to Bolandia and its cypselas match those of Bolandia in shape, colour and indumentum, except for a distinct ridge of hairs on the inner rib of the ray cypselas. It is further distinguished Keywords: from Bolandia by its annual habit and conical disc floret style apices and obtuse to rounded ray floret style Emilia hantamensis apices. It is recognised and described here as a new monotypic genus, Bertilia Cron with the single species Molecular phylogeny B. hantamensis (J.C.Manning & Goldblatt) Cron. Northern Cape endemic © 2013 The Author. Published by Elsevier B.V. on behalf of SAAB. All rights reserved. Systematics 1. Introduction The disjunct distribution of Emilia hantamensis relative to the other Emilia species in southern Africa prompted investigation into relation- Emilia hantamensis J.C.Manning & Goldblatt was first recognised as ships within the genus, notably the relationship of E. hantamensis to a distinct new taxon by Snijman and Perry (1987) during a survey of the other radiate Emilias in sect. Spathulatae of Jeffrey (1986) versus the Nieuwoudtville Wildflower Reserve in the Northern Cape, South the discoid species in South Africa and/or Namibia in sect. Emilia.The Africa. It was initially thought to be an Othonna L. but was subse- geographically nearest member of sect. Spathulatae is the yellow- quently identified as an Emilia (Cass.) Cass. by Bertil Nordenstam flowered E. discifolia in Zimbabwe, whereas E. transvaalensis is the (at S). It was recollected and described by Manning & Goldblatt as nearest discoid species, with its most southerly population from E. hantamensis (Manning and Goldblatt, 2001). It is now known to Mkuze, KwaZulu-Natal (Hilliard, 1977), though mainly occurring occur across the Hantam and along the Roggeveld, from north of in Limpopo Province. Manning and Goldblatt (2001: 48) noted that Loeriesfontein to Middelpos in the Northern Cape (Fig. 1). although E. hantamensis resembled species of sect. Spathulatae in its Emilia is a mainly tropical genus of ca. 100 species — most occur in “radiate yellow-flowered capitula and exappendiculate style branches, Africa, with a few in Asia and 14 in Madagascar (Humbert, 1963; …the rather narrow corolla lobes of its disc florets are not typical Jeffrey, 1986; Bremer, 1994; Nordenstam, 2007). It is characterised of the section ”. This study therefore examined the relationships of by its usually annual (occasionally perennial) habit, glabrous or E. hantamensis in the Senecioneae and tested its placement in Emilia scanty indumentum, uniseriate ecalyculate involucre, persistent pap- using a molecular phylogenetic approach. pus bristles and a basic chromosome number of n = 5, 8, 10, 11, 15, or 20 (Jeffrey, 1986; Nordenstam, 2007). Most members are discoid, 2. Material and methods only a few are radiate, and flower colours vary from white, mauve and purple, to yellow, orange and red. The description of E. hantamensis Morphological features of specimens of Emilia hantamensis from brought to seven the number of species of Emilia occurring in southern J, PRE and NBG were examined using a Zeiss V12 Discovery stereo mi- Africa, with E. hantamensis the only one in the winter rainfall region. croscope and photographed using an AxioCam MRc camera. The spec- imens examined are listed in Section 5.2. ☆ This is an open-access article distributed under the terms of the Creative Commons 2.1. Taxon sampling Attribution-NonCommercial-No Derivative Works License, which permits non-commercial use, distribution, and reproduction in any medium, provided the original author and source Six of the species of Emilia from southern Africa were included in are credited. ⁎ Tel.: +27 11 7176480; fax: +27 11 7176494. the phylogenetic analyses, including the radiate E. discifolia (type of E-mail address: [email protected]. sect. Spathulatae), as well as a range of related African taxa in the 0254-6299/$ – see front matter © 2013 The Author. Published by Elsevier B.V. on behalf of SAAB. All rights reserved. http://dx.doi.org/10.1016/j.sajb.2013.04.014 G.V. Cron / South African Journal of Botany 88 (2013) 10–16 11 Fig. 1. Known distribution of Bertilia hantamensis (previously Emilia hantamensis) in the Northern Cape, South Africa. Senecioneae (Appendix A), based on relationships in the subtribe the phylogeny based on nuclear ITS and ETS sequence data (Pelser et Senecioninae evident in Pelser et al. (2010). Representatives of al., 2010, Fig. 2). The mainly Eurasian genus Tephroseris (Rchb.) Rchb. Pericallis D.Don from the Canary Islands were included as it is placed from the subtribe Tussilagininae was used to root the trees. Se- (with BS b50%) sister to a clade comprising E. coccinea and Packera in quences for Mesogramma DC. and Tephroseris were obtained from Tephroseris 40/3 Euryops brownei 9/4 Dendrosenecio kilimanjari 78 14/0 3/3 Oresbia heterocarpa 4/0 Kleinia galpinii 54/4 83 Cineraria geifolia 74 4/1 Cineraria saxifraga 1/0 Cineraria deltoidea 88 94 96 3/0 Cineraria cyanomontana 8/1 1/2 1/0 Cineraria mollis 1/0 Bolandia argillacea 1/0 68 Bolandia pedunculosa 74 72 1/0 Bolandia pinnatifida 69 1/0 2/1 Bolandia elongata 3/0 75 Bolandia glabrifolia 2/0 1/1 E hantamensis 1 97 100 4/0 8/2 10/2 80 E hantamensis 2 Mesogramma apiifolium 1/2 20/2 Stilpnogyne bellodioides 7/1 95 Emilia ambifaria 100 3/1 Emilia marlothiana 26/4 100 Emilia transvaalensis 71 4/0 37/3 62 Emilia discifolia 7/0 63 14/7 6/2 Emilia protracta 6/1 39/1 100 7/1 Pericallis multiflora 23/1 15/1 Pericallis murrayi 66 Senecio cordifolius 8/1 2/0 Senecio deltoideus 20/0 Fig. 2. One of 20 equally most parsimonious trees (592 steps), based on the nuclear ITS region with Tephroseris as outgroup. Arrows indicate where tree collapses in the strict con- sensus tree. CI (excluding uninformative characters) = 0.64, RI = 0.79. Bootstrap percentages above the branches, minimum branch length below (point mutations/indels). 12 G.V. Cron / South African Journal of Botany 88 (2013) 10–16 Genbank. Sequence data were unavailable for the flanking portion of nuclear and plastid phylogenies, with Stilpnogyne retrieved as sister to the trnF exon, the small subunit and the 5.8 S genes of Tephroseris, the rest of the clade in the ITS phylogeny (Fig. 2)butwithMesogramma and these regions were coded as missing data. retrieved in this position in the trnL–trnF analysis. The six species of Emilia included in this study are in a clade sister 2.2. Molecular methods and phylogenetic analysis to Pericallis D.Don (Figs. 2 and 3), sister to two species of Senecio L. be- longing to the Senecio segregates group of Pelser et al. (2010), i.e. not Leaf material was collected and dried in silica gel, with voucher Senecio sensu stricto. The placement of E. transvaalensis varies between specimens (Appendix A) placed in J and KMG. DNA was extracted the two phylogenies: it is placed within Emilia s.s. in a very strongly using the DNeasy Plant Minikit (Qiagen) and the nuclear ribosomal supported clade (BS 100%) sister to E. ambifaria + E. marlothiana in internal transcribed spacer (ITS) regions and the plastid trnL–trnF re- the ITS-based phylogeny (Fig. 2), but is placed with Kleinia galpinii gion were amplified using the AB101 and AB102 primers of Sun et al. and Oresbia Cron & B. Nord. in a well supported clade (BS 89%) in the (1994) and the ‘c’ and ‘f’ primers of Taberlet et al. (1991) respectively. trnL–trnF analysis (Fig. 3). The two species of Pericallis form a monophy- Polymerase chain reactions (PCR) for the ITS regions were carried out letic group (BS 100%) in the ITS phylogeny (Fig. 2), but are members of a in 50 μℓ total volume with 0.4 μℓ of Truestart Taq (Fermentas®), grade in the same clade in the plastid phylogeny (Fig. 3). Details of the 5 μℓ buffer and 4.5 μℓ MgCl2, 0.5 μℓ primers (10 mM) and 1 μℓ DNA sequence matrices and the tree statistics are presented in Table 1. dNTPs (10 mM) together with the 0.5 or 1 μℓ of DNA. Amplification of the ITS region was as follows: premelting at 95 °C for 2 min, 35 cy- 4. Discussion cles of denaturation at 95 °C for 50 s, annealing at 54 °C for 45 s, ex- tension at 72 °C for 1.30 min, followed by a final extension at 72 °C Molecular analyses of ITS and trnL–trnF sequence data indicate for 7 min. Pyrostart PCR Fast Mastermix (Fermentas®) was used to that Emilia hantamensis is not correctly placed in Emilia as it does amplify the trnL–trnF region with 0.25 μℓ primers (10 mM) and not group with the other six Emilia species included in the nuclear 0.5 μℓ DNA in a total reaction volume of 20 μℓ: premelt at 95 °C and plastid phylogenies.