A new species of Teucrium sect. Scordium (Lamiaceae) from SE of Turkey

Turkish Journal of Botany Turk J Bot (2015) 39: 310-317 © TÜBİTAK doi:10.3906/bot-1402-93 http://journals.tubitak.gov.tr/botany/ Research Article A new species of Teucrium sect. Scordium (Lamiaceae) from SE of Turkey 1 1, 2 1 1 Taner ÖZCAN , Tuncay DİRMENCİ *, Fatih COŞKUN , Ekrem AKÇİÇEK , Özal GÜNER Department of Biology Education, Necatibey Education Faculty, Balıkesir University, Balıkesir, Turkey 2 Department of Biology, College of Arts and Sciences, Balıkesir University, Çağış Campus, Balıkesir, Turkey 1 Received: 28.02.2014 Accepted: 09.09.2014 Published Online: 16.03.2015 Printed: 10.04.2015 Abstract: A new species of Teucrium L. from Southeast Anatolia (Turkey), Teucrium sirnakense Özcan & Dirmenci, is described and illustrated. The new species belongs to the sect. Scordium Boiss. and is closely related to Teucrium melissoides Boiss. & Hausskn. and Teucrium scordium L. s.l. The differences between the new species and its allies are discussed. A description, distribution map, ITS nrDNA phylogeny, and taxonomic comments on the new species are provided. Key words: ITS, nrDNA, phylogeny, Lamiaceae, Şırnak, Teucrium, Turkey 1. Introduction Teucrium L. is one of the largest genera belonging to Lamiaceae, subfamily Ajugoideae (Harley et al., 2004). The genus is very large, polymorphic, and cosmopolitan consisting of about 260 species (more than 370 taxa) (Tutin and Wood, 1972; Govaerts et al., 2013). Approximately 250 species in the genus are distributed around the Mediterranean basin, which is the major center of diversity (Cantino et al., 1992; Navarro and El Oualidi, 2000; Govaerts, 2013). Teucrium is distinguished from the other members of Lamiaceae by a lack of corolla upper lip and nongynobasic style (Ekim, 1982; Navarro and El Oualidi, 2000; De Martino et al., 2010). The genus Teucrium has been divided into sections distinguishable by general habit, leaf characters, calyx shape, and inflorescence structure (Ekim, 1982). Teucrium comprises 34 species (with 46 taxa) in Turkey, and 16 of these taxa are endemic (Ekim, 1982; Duman, 2000; Dönmez, 2006; Dönmez et al., 2010; Dinç and Doğu, 2012; Dirmenci, 2012). The species are classified into 8 sections in the Flora of Turkey (Ekim, 1982): sect. Teucrium Benth., with 13 taxa; sect. Chamaedrys (Mill.) Schreb., with 13 taxa; sect. Polium (Mill.) Schreb., with 2 taxa; sect. Isotriodon Boiss., with 10 taxa; sect. Scorodonia (Hill) Schreb., with 1 taxon; sect. Stachybotrys Benth., with 3 taxa; sect. Scordium (Mill.) Benth., with 3 taxa; and sect. Spinularia Boiss., with 1 taxon. Sect. Scordium is represented by 4 taxa in the Mediterranean area (Navarro and El Oualidi, 2000). In Turkey, T. scordium L. (T. scordium subsp. scordium and * Correspondence: 310 T. scordium subsp. scordioides (Schreb.) Arcang.) and T. melissoides Boiss. & Hausskn. are placed in this section (Ekim, 1982; Dönmez et al., 2010). These taxa are either stoloniferous or erect, leaves are conspicuously toothed, crenate, subcordate at base, not petiolate or subpetiolate; verticillasters 2–8 flowers in upper leaf axils, calyx tubular-campanulate, subgibbose at base and calyx tooth triangular-acuminate; nutlet with glandular hairs (Zohary, 1963; Ekim, 1982). During the revisionary study of the Turkish Teucrium species some interesting specimens were collected from Şırnak Province in Turkey in June 2013 (Figure 1). After detailed studies, it was concluded that the specimens represented here are species new to science, and that they show a close relationship with T. melissoides and T. scordium s.l. In this study a new species from the genus Teucrium is described and investigated based on morphologic and molecular phylogenetic data. 2. Materials and methods 2.1. Plant collection The specimens were identified using the relevant literature (Boissier, 1879; Post, 1932; Feinbrun, 1978; Ekim, 1982; Rechinger, 1982) and compared with material found in the following herbaria, ANK, B, BCN, E, EGE, GAZI, HUB, INONU, ISTE, ISTF, ISTO, K, KNYA, L, LE, MA, MARE, RO, TO, VANF, W, WIR, and WU and with our own herbarium specimens at Balıkesir University (Appendix; on the journal’s website). ÖZCAN et al. / Turk J Bot Figure 1. Distribution map of Teucrium sirnakense (■), T. scordium subsp. scordium (●), T. scordium subsp. scordioides (○), T. melissoides (★) in Turkey. 2.2. DNA isolation DNA isolation was performed using the modified phenol/ chloroform/isoamyl alcohol method of Dellaporta et al. (1983). First, dried silica gel leaf samples were powdered using liquid nitrogen. Then 600 µL of isolation buffer and 500 µL of phenol/chloroform/isoamyl alcohol solution (24:24:1) were added to the powdered plant tissue. The solution was mixed carefully and centrifuged. The supernatant was taken into a new tube. Then 3 M NaAc (10% of supernatant) was added, followed by an isopropanol addition (in this step, the DNA was visible and so we continued the process). This solution was centrifuged, and the pellet was visible to the eye at the bottom of the tube. The pellet was dissolved using 500 µL of TE (10 mM, pH 8). Then 5 µL of RNase A was added and mixed in gently. This was incubated at 37 °C for 30 min. After 30 min, 50 µL of NaAc (3 M) was added and mixed in. Then 1 mL of 90% ethyl alcohol was added to the solution and pipetted. It was refrigerated at –80 °C for about 10 min (or at –20 °C for about 30 min). After refrigeration the tube was centrifuged, and the supernatant was discarded; 70% ethyl alcohol was added to the pellet and centrifuged. After centrifugation ethyl alcohol was removed, and the pellet was dried on blotting paper. As a last step, the dried pellet was dissolved using 50 µL of TE or 200 µL of H2O. Finally, the DNA solution was stored at –20 °C. 2.3. PCR amplification and editing of the ITS nrDNA data In this study the internal transcribed spacer (ITS) region of the nuclear ribosomal DNA (nrDNA) sequences was used for molecular analysis of the Teucrium species. Polymerase chain reaction (PCR) amplifications of the ITS nrDNA were performed using ITS5A (5’-CCTTATCATTTAGAGGAAGGAG-3’) (Stanford et al., 2000) and ITS4 (5’-TCCTCCGCTTATTGATATGC-3’) (White et al., 1990) primers. Total volume of each PCR tube was 25 µL and comprised 2.5 µL of CoralLoad PCR Buffer, 1.5 µL of MgCl2, 2.5 µL of Q-solution, 0.3 µL of Taq DNA polymerase (Qiagen), 2.5 µL of ITS5A and 2.5 µL of ITS4 (Sigma-Aldrich), 0.4 µL of 20 µm dNTP solution (Qiagen), and autoclaved deionized water. During the PCR amplification, a thermal cycler machine (Techne-Prime) was used for the routine protocol. Initial denaturation was performed for 5 min at 95 °C. The following 35 cycles were carried out for 1 min at 94 °C for denaturation, 1 min at 51 °C for annealing, and 2 min at 72 °C for extension. A final extension cycle (8 min at 72 °C) followed. PCR products were sent to the RefGen (Gene Research and Biotechnology Company) for sequencing. The sequenced DNAs were edited using Sequencher ver (...truncated)