ARTICLE IN PRESS Flora ] (]]]]) ]]]–]]] www.elsevier.de/flora Seed-coat microsculpturing and its systematic application in Isatis (Brassicaceae) and allied genera in Iran Hamid Moazzenia, Shahin Zarrea,Ã, Ihsan A. Al-Shehbazb, Klaus Mummenhoffc aDepartment of Botany, School of Biology, University College of Science, University of Tehran, P.O. Box 14155-6455, Tehran, Iran bMissouri Botanical Garden, P.O. Box 299, St. Louis, MO 63166-0299, USA cUniversita¨t Osnabru¨ck, Spezielle Botanik, Barbarastrasse 11, 49076 Osnabru¨ck, Germany Received 2 June 2006; accepted 19 October 2006 Abstract In order to examine the systematic application of seed-coat microsculpturing in Isatis, seed surfaces of 23 species (41 populations) in four genera of tribe Isatideae were examined using scanning electron microscopy (SEM). Eight types of basic ornamentation patterns were recognized among the studied specimens. Of these, the reticulate–areolate type was the most common and was found in the genera Isatis, Pachypterygium, Samerari and Tauscheria and 15 species (e.g., I. cappodocica, I. kotschyana and I. tinctoria). The reticulate type, the second most frequent, occurred in 7 species while other types each were represented by only one or 2 species. Although different populations of a given species show similar seed-surface sculpturing in most cases, in some polymorphic species like I. cappadocica and I. kotschyana these patterns were variable among populations. To some extent the variation corresponds to infraspecific taxa for some species, but the differences are not significant enough to be useful in the delimitation of the subspecies recognized by previous workers. Moreover, seed-coat characters do not support the separation of genera Isatis, Pachypterygium, Sameraria and Tauscheria. r 2007 Published by Elsevier GmbH. Keywords: Brassicaceae; Isatis; Sameraria; Intraspecific classification; Seed-coat microsculpturing; Iran Introduction even in fruits, which provide the most valuable diagnostic characters (Davis, 1965; Hedge, 1968; Jafri, Isatis L. comprises approximately 79 species (Al- 1973). This pattern of variation might suggest that Shehbaz et al., 2006) and the genus is distributed hybridization may have played a role in the evolution of primarily in the northern hemisphere, especially in the the genus. Notes on intermediate morphological char- Irano-Turanian region where nearly 90% of its species acters of Isatis specimens have been frequently made, grow (Appel and Al-Shehbaz, 2003; Davis, 1965). Isatis even among some taxa that are morphologically distinct species are sometimes highly polymorphic morphologi- (e.g., in the case of I. buschiana Schischik. which can be cally and morphological differences are often obscure, erroneously determined as I. tinctoria L. and I. lusitanica L., Davis, 1965; Hedge, 1968), and thus numerous ÃCorresponding author. Tel.: +98 21 61112482; specimens have to be analyzed for the delimitation of fax: +98 21 66405141. species. Very closely related to Isatis are Pachypterygium E-mail address: [email protected] (S. Zarre). Bunge, Tauscheria Fisch. ex DC., Sameraria Desv. and 0367-2530/$ - see front matter r 2007 Published by Elsevier GmbH. doi:10.1016/j.flora.2006.10.004 Please cite this article as: Moazzeni, H., et al., Seed-coat microsculpturing and its systematic application in Isatis (Brassicaceae) and allied genera in.... Flora (2007), doi:10.1016/j.flora.2006.10.004 ARTICLE IN PRESS 2 H. Moazzeni et al. / Flora ] (]]]]) ]]]–]]] the Central Asian Chartoloma Bunge (not reported thus not congruent with the traditional classification of far from Iran). Most of these genera are placed in the Schulz (1936) based on fruit form. Several lineages in tribe Isatideae by De Candolle (1821), but transferred the molecular tree include Thlaspi species with diverse into Arabideae subtribe Isatidinae by Hayek (1911) and fruit types, whereas species with the same fruit type subsequently into Lepidieae subtribe Isatidinae by belong to different clades, thus providing strong Schulz (1936). The differences between these genera evidence for convergence in fruit traits. Vaughan and are often based on single fruit characters. For example, Whitehouse (1971) investigated the seed-coat anatomy Sameraria differs from Isatis by its distinct (instead of of 200 species of Brassicaceae, but only a few Isatis obsolete) style. The thickened (vs. thin) fruit margin is species have been studied using the scanning electron used to separate Pachypterygium from Isatis (Hedge, microscopy (SEM). 1968). However, some botanists (e.g., Jafri, 1973; The present study is the first to investigate the seed Rechinger, 1958; Sajedi et al., 2005) reduce Pachypter- surface of Isatis and its allies in a taxonomic context. It ygium to synonymy of Isatis and consider the thickened deals with 41 populations of 23 species in four genera of fruit margin not a reliable character to separate these the tribe Isatideae sensu Al-Shehbaz et al. (2006). genera apart. Based strictly on the overall morphology, it is highly likely that the Isatideae sensu Al-Shehbaz (Al-Shehbaz Material and methods et al., 2006) also include the genera Pachypterygium (3 spp.), Sameraria (9 spp.) and the monotypic Boreava Information on the species analyzed, collection data Jaub. & Spach, Chartoloma, Glastaria Boiss., Schimpera and the vouchers used are given in Table 1. Seeds were Hochst. & Steud. ex Endl., Spirorrhynchus Kar. & collected from living plants whenever possible, otherwise Kir. and Tauscheria. Both Myagrum L. and Tauscheria from herbarium specimens. The species belong to Isatis were placed by De Candolle (1821) with Isatis (inclu- (15 of 17 species in Iran), Sameraria (4 of 5 species in ding Sameraria) in the Isatideae. This morphologically Iran), Pachypterygium (all 3 species) and Tauscheria well-defined group of primarily central Asian genera (monotypic, distributed in Iran and neighboring coun- is characterized by having indehiscent, 1- or 2-seeded, tries). Iranian species which are not included in this primarily angustiseptate fruits, yellow or rarely whitish analysis were previously recorded as doubtfully occur- flowers, sessile and often auriculate cauline leaves, ring in Iran, or they are rare or known only from the and simple or no trichomes (Appel and Al-Shehbaz, type collections that possess no seeds. 2003). Preliminary molecular data (Beilstein et al., For the SEM studies, clean seeds were fixed on 2006) show that Isatis and Myagrum (1 sp.) form aluminum stubs using double-sided adhesive and were a monophyletic group (93% bootstrap support), coated with a thin layer (ca. 25 nm) of gold-palladium. sister to a clade including the tribes Brassiceae, The SEM micrographs were taken in a Zeiss SEM-960A Schizopetaleae and Sisymbrieae. However, the (Germany) (Figs. 1–6) or Leo SEM-4401 (England) phylogenic relationships and the delimitation of the (Figs. 7–15) from the middle of the seed, mostly at a Isatideae are still awaiting a comprehensive molecular magnification of 100–700 Â . At least four seeds from analysis. each specimen were scanned to ensure the consistency of Some of the published accounts on the seed coat of seed-coat characters. For recording gross morphology Brassicaceae (e.g., Barton, 1967) dealt with the species and size parameters, at least 10 seeds were measured. of economic importance (e.g., Brassica L., Sinapis L. Although the terminology of Barthlott (1981) was and Raphanus L.) and were primarily concerned with based on SEM observations of epidermal and seed-coat species identification. Other studies used the ultra- surface in 5000 species of seed plants and has been used structural pattern as a reliable approach for assessing in several studies, we found those of Murley (1951) and phenetic relationships and resolving taxonomic pro- Koul et al. (2000) to be more applicable to the patterns blems in the Brassicaceae (Barthlott, 1981; Bengoechea in Isatis and its relatives: and Go´ mez-Campo, 1975; Bouman, 1975; Buth and Roshan Ara, 1983, 1987; Cˇernohorsky` , 1950; Francois, Colliculate: having small hill-like eminences or 1937; Koul et al., 2000; McCugan, 1948; Meyer, 1973, rounded broad elevations, closely spaced, covering 1979, 1991; Murley, 1951; Musil, 1948; Vaughan and throughout the seed coat (Fig. 4); Whitehouse, 1971). For Thlaspi L., it was clearly Lineolate: marked with broken fine lines (Fig. 7); demonstrated that molecular data (Koch and Mum- Ocellate: having eye-like depressions, each with a menhoff, 2001; Mummenhoff and Koch, 1994; Mum- raised circular border (Figs. 14 and 15); menhoff et al., 1997a, b; Zunk et al., 1996) provide Reticulate: with a reticulum or net and interspaces strong support for the recognition of several segregates characterized with raised walls. The interspaces can as proposed by Meyer (1973, 1979, 1991) based on seed- be distinct (Fig. 8) and relatively large or more or less coat anatomy. The molecular phylogeny of Thlaspi is indistinct and small (Fig. 10); Please cite this article as: Moazzeni, H., et al., Seed-coat microsculpturing and its systematic application in Isatis (Brassicaceae) and allied genera in.... Flora (2007), doi:10.1016/j.flora.2006.10.004 ARTICLE IN PRESS H. Moazzeni et al. / Flora ] (]]]]) ]]]–]]] 3 Table 1. List of taxa, voucher and some characteristic seed features of Isatis and its allies used in the present study Taxon Place of origin and herbarium Voucher Seed Seed-surface number length pattern (mm) I. buschiana Kordistan, Bijar to Zanjan, 5 km up to Bianlou 973-KOHa 3.1270.25 Reticulate