Turkish Journal of Botany Turk J Bot (2016) 40: 87-96 http://journals.tubitak.gov.tr/botany/ © TÜBİTAK Research Article doi:10.3906/bot-1407-71 New species and a synonym of the genus Hesperis (Brassicaceae) from Turkey Ahmet DURAN, Özlem ÇETİN* Department of Biology, Faculty of Science, Selçuk University, Selçuklu, Konya, Turkey Received: 18.07.2014 Accepted/Published Online: 12.04.2015 Final Version: 01.01.2016 Abstract: Hesperis tosyaensis A.Duran, a new species from northern Anatolia, is described and illustrated. It grows under open Quercus forest. The new species is distinct from the closely allied H. ozcelikii A.Duran by the following features: stem erect, 45–90 cm tall, hispid, hairs 2–2.5 mm, and fruit glabrous. Genetic differentiation between closely related Hesperis L. taxa and Matthiola W.T.Aiton was examined by molecular analyses of the ISSR. The chromosome number for Hesperis tosyaensis is 2n = 14. Information is also presented on its ecology and conservation status. In addition, Hesperis aspera E.Fourn. is treated as a synonym of Hesperis bicuspidata (Willd.) Poir. Key words: Hesperis, new species, ISSR, Cruciferae, systematics 1. Introduction on the infrageneric and infraspecific classification of the Hesperis L. (Brassicaceae) is represented by many taxa at genus Hesperis, ecological and geographic variations of the junctions of the Irano-Turanian, Mediterranean, and some Hesperis taxa cause some confusion in taxonomy Euro-Siberian phytogeographic regions; these regions and nomenclature, especially among Hesperis matronalis meet in Anatolia. Towards the outer boundaries of each L., H. bicuspidata (Willd.) Poir., H. pendula DC., and phytogeographic region, Hesperis is represented by fewer certain other taxa. Hesperis bicuspidata is localized in the taxa, which is also indicated by the number of Hesperis taxa Irano-Turanian phytogeographical region and the species in various floras. There are 14 species in Europe, 11 species has a widespread distribution in Turkey. H. bicuspidata in Iran, 9 species in Romania, 5 species in Iraq, 3 species in grows in different habitats with different features. Hesperis Italy, and 1 species in Palestine (Săvulescu, 1955; Ball, 1964; bicuspidata specimens were collected from different Zohary, 1966; Dvořák, 1968, 1980; Pignatti, 1982; Duran localities in Turkey. and Hamzaoğlu, 2004; Duran and Ocak, 2005; Duran, 2005; Hesperis aspera, which is endemic to Turkey and Parolly and Tan, 2006; Duran, 2008, 2009). is known only from Tosya (Kastamonu Province), was The genus Hesperis is represented by 31 species in the described as a new species by Fournuer (1866). Our flora of Turkey (Cullen, 1965; Davis et al., 1988). Five new examinations of type specimens and herbarium specimens species have been described from Turkey and 10 species of H. aspera have revealed that it is a synonym of H. are treated as synonymous (Duran and Ocak, 2005; Parolly bicuspidata. Based on our study, H. aspera is treated as a and Tan, 2006; Duran, 2008, 2009; Duran et al., 2011). synonym of H. bicuspidata. According to the latest study, Hesperis is represented by 27 Some interesting Hesperis specimens were collected species recorded from Turkey (Duran, 2012). from the Tosya district (Kastamonu). Herbarium studies The genus Hesperis in Turkey has been studied in terms and field observations showed that our specimen is a of morphology, floristic features, palynology, molecular new taxon. Diagnostic morphological characters from characteristics, anatomy, and chemotaxonomy (Duran, closely similar Hesperis species are discussed in this study. 2005; Duran and Ocak, 2005; Duran, 2008; Ünal et al., Ecology, conservation status, and biogeography of the new 2008; Yücel et al., 2008; Aras et al., 2009; Büyükkartal et al., species are presented. Genetic differentiation of the new 2009; Duran, 2009; Pınar et al., 2009; Kırımer et al., 2010). species and related species were analyzed using NTSYSpc Despite the fact that many studies have been performed 2.1 software and dendrograms were obtained. * Correspondence: [email protected] 87 DURAN and ÇETİN / Turk J Bot 2. Materials and methods performed in a mix containing 2.5 μL of PCR buffer, 3 μL 2.1. Morphology of 25 mM MgCl2, 1 μL of primer, 0.5 μL of dNTP mix, 0.3 We followed the descriptions of taxa according to the μL of Taq DNA polymerase, 4 μL of each diluted DNA, and Flora of Turkey and the eastern Aegean Islands and 13.7 units of PCR water. After a predenaturation step of 3 other floras (Halácsy, 1900; Hayek, 1927; Busch, 1939; min at 94 °C, amplification reactions were cycled 40 times Săvulescu, 1955; Tzvelev, 1959; Ball, 1964; Cullen, 1965; at 94 °C for 1 min, at annealing temperature (Table 1) for 1 Zohary, 1966; Dvorák, 1968; Dvorák, 1980; Pignatti, 1982; min, and at 72 °C for 1 min in an Eppendorf Mastercycler Davis et al., 1988; Tan and Iatrou, 2001) were examined. gradient thermocycler. A final extension was performed The specimens present in the AEF, ANK, BM, E, EGE, G, for 10 min at 72 °C. Amplified fragments were visualized GAZI, HUB, ISTF, K, KNYA, P, and WU herbaria were under a UV transilluminator and photographed using examined. All measurements were made on 10 randomly a gel documentation system (Vilbert Lourmat, Infinity selected specimens. Measurements were made of plant model). All amplified fragments were treated as dominant parts using a ruler and an ocular micrometer under a genetic markers. Each DNA band generated was visually microscope. Localities of studied taxa are given in the scored as an independent character or locus. Analyses Appendix. were performed with the NTSYS-pc 2.1 software package (Rohlf, 2001). The dendrogram was constructed using SM 2.2. Karyology coefficient and UPGMA clustering methods. Karyological observations were carried out on root tips obtained from germinating seeds. Root tips were 3. Results pretreated for 16 h in α-monobromonaphthalene at 4 °C Hesperis bicuspidata (Willd.) Poir., Encycl. Suppl. 3: 195 and fixed in Carnoy solution (3:1 absolute ethanol and (1813) glacial acetic acid), and then the root tips were hydrolyzed Sect. Hesperis with 1 N HCl at room temperature and stained with 2% Type: Tournefort, no. 16 (B, P! KNYA photo.!, GAZI aceto-orcein. Stained root tips were flattened in a drop photo!). Described from Turkey. of 45% acetic acid, and permanent slides were made by Synonyms ≡ Cheiranthus bicuspidatus Willd., Sp. Pl., 3, mounting in Depex. 1: 519 (1800). = Hesperis kotschyana Fenzl, Pug. PI. Nov. 2.3. Molecular analyses Syr., 13 (1842); H. violacea Boiss., Diagn. PI. Orient., ser. Total DNA was obtained from 50–75 mg of dried leaf 1, 5: 80 (1844); H. reuteriana Boiss. & A.Huet, Diagn. PI. tissue from 11 different individuals. DNAs were extracted Orient., ser. 2, 5: 20 (1856); H. unguicularis Boiss., Diagn. using phenol-chloroform (Sambrook et al., 1989) after PI. Orient., ser. 2, 5: 21 (1856); H. aspera E. Fourn., Bull. concentrations were determined by NanoDrop. Sample Soc. Bot. Fr. 13: 352 (1866) syn. nov.; H. tauricola Kotschy DNAs were diluted to 25 ng/µL. Stock DNAs were kept & Boiss., Fl. Orient. 1:232 (1867); H. sintenisii Hausskn. at –86 °C. ISSR primers were used for PCR amplification; & Bornm., Repert. Spec. Nov. Regni Veg. Beih. 89(1): 38 the characteristics of the primers used are given in Table (1936); H. stellata Dvorak, Oesterr. Bot. Z. 112: 610 (1965). 1. Each reaction contained 25 μL. PCR reactions were Hesperis tosyaensis A.Duran sp. nov. (Figures 1–6) Table 1. The characteristics of the ISSR primers. Primer Sequences Tm (°C) Length (bp) GC ratio (%) Total bands M1 5ʹ-AGC AGC AGC AGC AGC AGC G-3ʹ 63.10 19 68.4 5 M2 5ʹ-ACC ACC ACC ACC ACC ACC G-3ʹ 63.10 19 68.4 9 M3 5ʹ-AGC AGC AGC AGC AGC AGC C-3ʹ 63.10 19 68.4 9 M5 5ʹ-GAG AGA GAG AGA GAG AGA C-3ʹ 56.70 19 52.6 16 M7 5ʹ-AGA GAG AGA GAG AGA GAG C-3ʹ 56.70 19 52.6 15 M9 5ʹ-ACA CAC ACA CAC ACA CCG-3ʹ 56.00 18 55.6 8 Diplo1 5ʹ-CAC ACA CAC ACA CAC AG-3ʹ 57.19 17 52.9 7 Cucurbita 5ʹ-GAG CAA CAA CAA CAA CAA -3ʹ 53.07 18 33.3 25 Cucurbita2 5ʹ-GAG AGA GAG AGA GAG AGA T-3ʹ 54.78 17 52.9 10 88 DURAN and ÇETİN / Turk J Bot Figure 1. Hesperis tosyaensis: A- general view, B- mature fruit. Scale bar = 2 cm. Subgen. Hesperis Sect. Hesperis Description: Perennial, rarely biennial herb. Roots Type: Turkey. A5 Kastamonu: between Tosya-Sekiler thickened taproot, (2.5–)4–16 mm diameter. Stem village, 7 km, 1000 m, under an open Quercus forest, ±erect, 45–90(–110) cm tall (including flowering part), 40°55.62′N, 34°02.06′E, 12.5.2001, A.Duran 5657 & generally purplish below or rarely completely, solitary or Hamzaoğlu (holotype: KNYA, isotypes: GAZI, ANK, 2–4-branched flowering part, terete, smooth, 1.5–9 mm ISTE, ISTF, E, K). in diameter at base; hispid especially in lower part, simple Diagnosis: Hesperis tosyaensis differs from H. ozcelikii hairs at bottom; bifurcate and simple hairs in middle part; by its erect stem, 45–90 cm (not ascending to erect, 25–60 bifurcate, a few trifid, simple, and rarely glandular hairs cm), simple hairs especially in lower part and hispid (not in upper part, long simple hairs 2–2.5 mm long. Leaves with only densely articulated glandular hairs or densely crowded at bottom and middle part; basal leaves ±narrowly glandular, sparsely bifurcate hairs below), hairs 2–2.5 mm lyrate, 5–10(–14) × 0.6–2(–3.5) cm (including 2.5–5 cm long (not ±1.5 mm), pedicel glabrous or sparsely bifurcate petiole), remotely denticulate; cauline leaves decreasing and rarely trifid hairs (not only densely glandular hairs, or to flowering part, all with main midrib ±conspicuous; bifurcate-stellate and glandular hairs), fruit glabrous (not lower cauline leaves bifurcate, glandular, a few simple and densely glandular, sparsely bifurcate hairs), valve narrower 3 branched hairs, simple hairs especially on main midrib than septum (not valve wider than septum).