Annals of Botany 109: 1341–1357, 2012 doi:10.1093/aob/mcs054, available online at www.aob.oxfordjournals.org Phylogenetic relationships and generic delimitation of Eurasian Aster (Asteraceae: Astereae) inferred from ITS, ETS and trnL-F sequence data Wei-Ping Li1,*, Fu-Sheng Yang2, Todorka Jivkova3 and Gen-Shen Yin4 1College of Life Sciences, Hunan Normal University, Changsha 410081, China, 2State Key Laboratory of Systematic and Evolutionary Botany, Institute of Botany, The Chinese Academy of Sciences, Beijing 100093, China, 3Department of Structural Biology, Faculty of Natural Sciences, Shumen University, Shumen 9700, Bulgaria and 4Key Laboratory of Biodiversity and Biogeography, Kunming Institute of Botany, The Chinese Academy of Sciences, Kunming 650204, China * For correspondence. E-mail [email protected] Downloaded from Received: 18 September 2011 Returned for revision: 7 November 2011 Accepted: 13 February 2012 Published electronically: 19 April 2012 † Background and Aims The classification and phylogeny of Eurasian (EA) Aster (Asterinae, Astereae, Asteraceae) remain poorly resolved. Some taxonomists adopt a broad definition of EA Aster, whereas others favour a narrow generic concept. The present study aims to delimit EA Aster sensu stricto (s.s.), elucidate the phylogenetic relationships of EA Aster s.s. and segregate genera. http://aob.oxfordjournals.org/ † Methods The internal and external transcribed spacers of nuclear ribosomal DNA and the plastid DNA trnL-F region were used to reconstruct the phylogeny of EA Aster through maximum parsimony and Bayesian analyses. † Key Results The analyses strongly support an Aster clade including the genera Sheareria, Rhynchospermum, Kalimeris (excluding Kalimeris longipetiolata), Heteropappus, Miyamayomena, Turczaninowia, Rhinactinidia, eastern Asian Doellingeria, Asterothamnus and Arctogeron. Many well-recognized species of Chinese Aster s.s. lie outside of the Aster clade. † Conclusions The results reveal that EA Aster s.s. is both paraphyletic and polyphyletic. Sheareria, Rhynchospermum, Kalimeris (excluding K. longipetiolata), Heteropappus, Miyamayomena, Turczaninowia, Rhinactinidia, eastern Asian Doellingeria, Asterothamnus and Arctogeron should be included in Aster, at Kunming Institute of Botany,CAS on July 25, 2012 whereas many species of Chinese Aster s.s. should be excluded. The recircumscribed Aster should be divided into two subgenera and nine sections. Kalimeris longipetiolata, Aster batangensis, A. ser. Albescentes, A. series Hersileoides, a two-species group composed of A. senecioides and A. fuscescens, and a six-species group including A. asteroides, should be elevated to generic level. With the Aster clade, they belong to the Australasian lineages. The generic status of Callistephus should be maintained. Whether Galatella (including Crinitina) and Tripolium should remain as genera or be merged into a single genus remains to be determined. In addition, the taxonomic status of A. auriculatus and the A. pycnophyllus–A. panduratus clade remains unre- solved, and the systematic position of some segregates of EA Aster requires further study. Key words: Asteraceae, Astereae, ETS, Eurasian Aster, generic delimitation, infrageneric classification, ITS, molecular phylogeny, trnL-F. INTRODUCTION Nesom (1994b) that a fundamental difference exists between NA and EA Aster. Furthermore, ITS data indicate that EA Aster sensu lato (s.l.; Asterinae, Astereae, Asteraceae) has Aster is nested in the Southern Hemisphere grade and does been a taxonomic dumping ground for large numbers of mor- not form a monophyletic group (Noyes and Rieseberg, 1999, phologically similar but distantly related taxa (Noyes and Brouillet et al., 2001, 2009b; Fiz et al., 2002), and African Rieseberg, 1999; Dorn, 2003). Aster s.l. occurs mainly in the Aster should be separated from Aster s.s. (Brouillet et al., Northern Hemisphere in both Eurasia (EA) and North 2009b). The classification and phylogeny of EA Aster have America (NA) and is estimated to comprise 250–1000 remained poorly resolved, however, because of insufficient species (Ling et al., 1985; Nesom, 1994b; Ito and Soejima, sampling in these studies. 1995; Noyes and Rieseberg, 1999). Based primarily on The circumscription of EA Aster has confused botanists for achene morphology and cytology, Nesom (1994b) segregated several decades. Many taxonomists have adopted a broad def- NA Aster species from Aster s.l. and redistributed them inition of EA Aster.InFlora Europaea, Merxmu¨ller et al. among generic segregates Symphyotrichum, Doellingeria, (1976) maintained Doellingeria, Galatella, Crinitaria (the Eucephalus, etc. At the same time, he kept the remainder, name Crinitaria is a synonym of Galatella and species that about 180 species, as Aster sensu stricto (s.s.), typified by are considered part of Crinitaria should be included in A. amellus. Consequently, Aster, containing approx. 180 Crinitina) and Tripolium in Aster. Similarly, Grieson (1975) species, is restricted to the Northern Hemisphere of the Old accepted Aster s.l. in Flora of Turkey and the East Aegean World. Internal transcribed spacer (ITS) sequence phylogenet- Islands because he did not recognize Kemulariella and ic data (Noyes and Rieseberg, 1999) support the viewpoint of Tripolium as segregate genera. In Flora of Japan, Ito and # The Author 2012. Published by Oxford University Press on behalf of the Annals of Botany Company. All rights reserved. For Permissions, please email: [email protected] 1342 Li et al. — Molecular phylogenetics of Eurasian Aster (Asteraceae: Astereae) Soejima (1995) merged Tripolium as section Tripolium into majority of EA Aster s.s. Thus, a reliable phylogenetic analysis Aster, placed Heteropappus in section Pseudocalimeris, based on extensive taxon sampling is essential to determine included Kalimeris within section Asteromoea, and the inter- and intrageneric relationships of EA Aster. associated Doellingeria and Miyamayomena into section Principally based on nuclear ribosomal DNA (nrDNA) ITS, Teretiachaenium. ETS and plastid trnL-F sequence data of Sheareria nana, Other taxonomists have favoured a narrow generic concept Rhynchospermum verticillatum and 62 species of EA Aster of EA Aster and have recognized small genera endemic to s.l., the present study aims to (1) reconstruct the phylogeny eastern Asia. Tamamschyan (1959) segregated two new of EA Aster s.l.; (2) redelimit the genus Aster and discuss its genera (Kemulariella and Conyzanthus)fromAster and recog- infrageneric classification; and (3) discuss the systematic pos- nized many small genera such as Doellingeria, Kalimeris, ition of EA Aster segregates. Asterothamnus, Krylovia, Turczaninowia, Galatella, Linosyris (¼ Crinitina) and Tripolium. Czerepanov (1995) fol- MATERIALS AND METHODS lowed Tamamschyan (1959) except that he placed Galatella ¼ and Linosyris under the genus name Crinitaria ( Generic circumscriptions and nomenclature of Astereae follow Downloaded from Crinitina). Nesom (1994a, b) made Aster largely equal to Nesom and Robinson (2007) except for Turczaninowia, which EA Aster s.s. and EA Aster s.l. almost equal to sub-tribe follows Ling et al. (1985), and Crinitina, which is substituted Asterinae Dumort. for Crinitaria (a synonym of Galatella). The name Aster setch- Ling et al. (1985) treated Chinese Asterinae in the narrow uenensis follows the International Plant Names Index. The div- sense of Aster, recognizing generic status for Gymnaster (¼ ision of phylogenetic lineages of Astereae refers to Brouillet Miyamayomena), Kalimeris, Callistephus, Heteropappus, et al. (2009b). Voucher DBY9206 was deposited in the http://aob.oxfordjournals.org/ Doellingeria, Turczaninowia, Krylovia (¼ Rhinactinidia), Wenzhou University Herbarium (WZU) and the others in Asterothamnus, Galatella, Linosyris (¼ Crinitina), the Hunan Normal University Herbarium (HNNU; see the Arctogeron and Tripolium. These treatments were followed Appendix). completely for floras of Chinese provinces (e.g. Zhuang, 2004; Lin, 2007). Despite this, Chinese Aster s.s. remains a Taxon sampling large genus with approx. 100 species, of which 75 are endemic to China (Fu, 1983; Ling et al., 1985; Chen, 1988, Seventy-six species of Astereae and three outgroup species 1990; Zhu and Min, 1990; Li and Liu, 2002; Li and Zhang, were collected from China and Bulgaria and examined for se- at Kunming Institute of Botany,CAS on July 25, 2012 2004). Therefore, China, especially south-western China quence variations in nrDNA ITS, ETS and plastid DNA trnL-F (the Qinghai–Tibetan and Yunnan–Guizhou Plateaux and (GenBank accession numbers are given in the Appendix). The Sichuan Province), is the diversity centre of Aster,asitis vouchers of all accessions were identified using published keys for many genera (Huang, 2011). and compared with herbarium specimens in the Institute of Molecular markers, especially ITS and the external tran- Botany, Chinese Academy of Sciences Herbarium (PE), scribed spacer (ETS) of 35S ribosomal DNA, have frequently Northwest Agriculture and Forestry University Herbarium been used to investigate phylogenetic relationships in Astereae (WUK), Sichuan University Herbarium (SZ), Chengdu (e.g. Noyes and Rieseberg, 1999; Lowrey et al., 2001; Markos Institute of Biology Herbarium (CDBI), HNNU, Herbarium and Baldwin, 2001; Cross et al., 2002; Fiz et al., 2002; of Kunming Institute of Botany, the Chinese Academy of Roberts, 2002; Lowell et al., 2003; Urbatsch and Roberts, Sciences (KUN), the Herbarium of the South China 2003; Urbatsch