Blackwell Publishing LtdOxford, UKBOJBotanical Journal of the Linnean Society0024-4074The Linnean Society of London, 2006? 2006 1513 411419 Original Article KARYOLOGY OF IRANIAN COUSINIA S. M. GHAFFARI Botanical Journal of the Linnean Society, 2006, 151, 411–419. With 32 figures New chromosome counts in the genus Cousinia (Asteraceae) from Iran SEYED MAHMOOD GHAFFARI1, NÚRIA GARCIA-JACAS2* and ALFONSO SUSANNA2 1 Institute of Biochemistry and Biophysics, University of Tehran, PO Box 13145-1384, Tehran, Iran Botanic Institute of Barcelona (CSIC-ICUB), Passeig del Migdia s.n., E-08038 Barcelona, Spain 2 Received April 2005; accepted for publication October 2005 Chromosome counts are reported of 38 populations representing 24 species of the genus Cousinia from Iran, one of the main centres of speciation of the genus. Sixteen of the counts are new to science. Our results confirm that Cousinia exhibits a complex dysploid series ranging from x = 13 to x = 9. One of the basic chromosome numbers, x = 9, is new for the genus and must be confirmed. Some considerations on the correlation between chromosome numbers and section classification are made. © 2006 The Linnean Society of London, Botanical Journal of the Linnean Society, 2006, 151, 411–419. ADDITIONAL KEYWORDS: Cardueae – Compositae – dysploidy – karyology. INTRODUCTION The Irano-Turanian and Central Asian genus Cousinia Cass. totals 662 species, 385 of which are in the Flora Iranica area (Rechinger, 1972, 1979, 1986; Knapp, 1987). No fewer than 220 species grow specifically in Iran, 165 of which are endemic (Ghahreman & Attar, 1999). These numbers are growing every year, with many new species being described recently (Ghahreman, Iranshahr & Attar, 1999; Attar & Ghahreman, 2000, 2002; Attar, Ghahreman & Assadi, 2000a, 2000b, 2001, 2002; Attar & Joharchi, 2002; Mehregan, Assadi & Attar, 2003; Mirtadzadini & Attar, 2004). The genus is remarkable in various respects. After Senecio (c. 1500 species) and Vernonia (c. 1000 species), Cousinia is the third largest genus in the Asteraceae; it is the largest in the tribe Cardueae and it causes astonishment amongst botanists due to the variability in its morphological characters. The shape and texture of phyllaries vary considerably, in some cases recalling other genera of the tribe Cardueae, sometimes unarmed, as in Centaurea L. or Jurinea ET AL. *Corresponding author. E-mail: ngarciajacas@ibb.csic.es Cass., but more often spiny. In addition, the achene characters vary to such an extent that one can hardly believe that the relevant species belong to the same genus when faced with the extremes of their variation. The generic delineation of Cousinia is a question that remains debatable, especially after the molecular study by Susanna et al. (2003a), and the distinction between Arctium (15 species only) and Cousinia is unclear. The sectional classification has also been questioned, partly on the same molecular basis (Susanna et al., 2003a) and partly on karyological grounds (Susanna et al., 2003b). Two classifications have been proposed so far, one by Tscherneva (1962) in her treatment for the Flora of the SSSR (revised later in Tscherneva, 1988a, 1988b), mainly based on the species of Central Asia, and one by Rechinger (1972, 1979, 1986), mainly on Iranian species. In fact, no botanist has worked more intensively on Irano-Turanian species of Cousinia than K. H. Rechinger. He has devoted a great deal of interest in this extraordinarily speciesrich genus over the last half century. His Flora Iranica (Rechinger, 1972, 1979, 1986) encompasses the greater part of Cousinia, with 385 species and 57 sections. It is worth mentioning that 153 species and 23 sections were described by him, and two species, C. rechingerae Bornm. and C. rechingerorum Bornm., © 2006 The Linnean Society of London, Botanical Journal of the Linnean Society, 2006, 151, 411–419 Downloaded from https://academic.oup.com/botlinnean/article-abstract/151/3/411/2420402 by guest on 28 July 2018 411 412 S. M. GHAFFARI ET AL. were named after him and his wife (see Plant Systematics and Evolution, 155, which was dedicated to him on the occasion of his 80th birthday). Rechinger was quoted as saying: ‘The question may arise as to whether Cousinia could be divided into several genera’ (Rechinger, 1986). He believed that the answer is no, at least at the present state of knowledge. This point of view, as far as Cousinia subgenus Cousinia is concerned, agrees with the preliminary results of molecular analyses in the complex (Susanna et al., 2003a). We should wait for more research combining achene and pollen characters, molecular analyses, and chromosome studies. Based on information found in the chromosome indexes (Ornduff, 1968, 1969; Fedorov, 1969; Moore, 1970, 1971, 1972, 1973, 1974, 1977; Goldblatt, 1981, 1984, 1985, 1988; Goldblatt & Johnson, 1990, 1991, 1994, 1996, 1998, 2000, 2003), it is evident that Cousinia is poorly known cytologically. Taking recent papers into account, chromosome numbers have been determined for approximately 114 species in the genus (Ghaffari & Djavadi, 1998; Djavadi & Ghaffari, 1999; Ghaffari, Attar & Ghahreman, 2000; Susanna et al., 2003b), which represents roughly 19% of total species. In this study, meiotic chromosome numbers (with one somatic count for C. boisseri Buhse) are reported for 24 species representing 16 sections and four species described by Rechinger (1972) without indication of section. MATERIAL AND METHODS Floral buds of appropriate size were fixed in absolute ethanol–chloroform–propionic acid (6 : 3 : 2) for 24 h, transferred to 70% alcohol, and stored under refrigeration until analysis. Anthers were squashed and stained in 2% acetocarmine. Chromosome counts were carried out from microsporocytes in various stages of meiosis. Slides were made permanent by the venetian turpentine method (Wilson, 1945). Voucher specimens were deposited in the Central Herbarium of Tehran University (TUH) or in the IRAN Herbarium. RESULTS AND DISCUSSION In this study, all sections and species follow the classification by Rechinger (1972). COUSINIA SECT. BADGHYSIA TSCHERNEVA Cousinia multiloba DC. Tehran: Haraz road, Emamzadeh Hashem, Ghaffari 3275, 2650 m, n = 13. Tehran: Damavand, between Deh-e-Chenar and Tar lake, Ghaffari 2660 m, n = 13. Rechinger (1972, 1979) mentioned this taxon as a widespread species endemic to Iran, Afghanistan, Pakistan, and Turkmenistan. According to Tscherneva (1988b), C. multiloba belongs to section Alpinae Bunge. The count agrees with a previous count of 2n = 26 for this species by Podlech & Bader (1974). Meiosis was observed in two collections, with 13 bivalents at first metaphase (Fig. 1). Chromosome segregation at anaphase I was a regular 13–13 (Fig. 2). Mean chiasma frequency was the same in the two collections and showed 1.30 per bivalent at metaphase I. On the basis of this and other previous counts (Afzal-Rafii, 1980), sect. Badghysia has x = 13. COUSINIA SECT. CHRYSOPTERA TSCHERNEVA Cousinia turkmenorum Bornm. & Gauba Khorasan: Mashhad, towards Nayshabour, Ghaffari 1756, n = 11. This taxon is a common species endemic to Iran, Turkmenistan, and Afghanistan. Meiosis was regular and showed 11 bivalents at metaphase I. Also, 11–11 chromosome segregation at anaphase I was observed (Figs 3, 4). In diakinesis, two bivalents were associated with the nucleolus. According to our data, this is the first chromosome count for this species. The only other count within this section was 2n = 18 in C. bipinnata (Chuksanova in Fedorov, 1969), a doubtful result. If this record is confirmed, section Chrysoptera has two basic chromosome numbers: x = 9 and 11. COUSINIA SECT. CONGESTAE BUNGE Cousinia congesta Bunge Qom: 90 km towards Arak, 1560 m, Ghaffari 11364, n = 12. Khorasan: east of Mashhad, 990 m, Ghaffari 1856, n = 12. Figures 1–16. Figs 1–15. Meiosis in Cousinia spp. Fig. 1. C. multiloba, metaphase I, n = 13. Fig. 2. C. multiloba, anaphase I (13–13). Fig. 3. C. turkmenorum, metaphase I, n = 11. Fig. 4. C. turkmenorum, anaphase I (11–11). Fig. 5. C. congesta, metaphase I, n = 12. Fig. 6. C. wilhelminae, metaphase I, n = 12. Fig. 7. C. behboudiana, metaphase I, n = 12. Fig. 8. C. keredjensis, metaphase I, n = 12, showing two quadrivalents (arrows). Fig. 9. C. onopordioides, metaphase I, n = 12. Fig. 10. C. gmelini, diakinesis, n = 12. Fig. 11. C. heliantha, metaphase I, n = 13. Fig. 12. C. aitchisonii, metaphase I, n = 13. Fig. 13. C. lasiandra, metaphase I, n = 13. Fig. 14. C. lasiandra, anaphase I (13–13). Fig. 15. C. arctotidifolia, metaphase I, n = 12. Fig. 16. C. boissieri, prophase of mitosis, 2n = 24. Scale bar = 10 µm. © 2006 The Linnean Society of London, Botanical Journal of the Linnean Society, 2006, 151, 411–419 Downloaded from https://academic.oup.com/botlinnean/article-abstract/151/3/411/2420402 by guest on 28 July 2018 413 KARYOLOGY OF IRANIAN COUSINIA 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 © 2006 The Linnean Society of London, Botanical Journal of the Linnean Society, 2006, 151, 411–419 Downloaded from https://academic.oup.com/botlinnean/article-abstract/151/3/411/2420402 by guest on 28 July 2018 414 S. M. GHAFFARI ET AL. Meiosis in this taxon was regular and showed 12 bivalents at metaphase I (Fig. 5). In diakinesis, two bivalents of chromosomes were associated with the nucleolus. This is the only species from this section in Iran, the rest occurring in Afghanistan and Turkmenistan. Previous chromosome counts for this taxon are n = 12 from Iran (Aryavand, 1975) and 2n = 26 from Uzbekistan (Susanna et al., 2003b), discordant results that suggest a misinterpretation of poor-quality somatic metaphase plates in Susanna et al. (2003b). COUSINIA SECT. COUSINIA Cousinia wilhelminae Rech. f. Azarbaijan-e-Garbi: Salmas, 10 km towards Khoy, 1420 m, Ghaffari 18271, n = 12. Twelve bivalents at metaphase I and 12–12 chromosome segregation at anaphase I were observed (Fig. 6). According to our data, this is the first chromosome count for this species. Eight endemic species of this section are present in Iran. C. wilhelminae is a narrow endemic to Iran, distributed in a restricted area of western Azarbaijan. There are six chromosome counts for sect. Cousinia in the literature, all the same as ours, showing x = 12 to be the basic chromosome number for this section. However, there is a count of 2n = 18 for C. horridula Juz. (Chuksanova in Fedorov, 1969), not confirmed by other authors (Tscherneva, 1985; Ghaffari & Djavadi, 1998; Susanna et al., 2003b). Cousinia onopordioides Ledeb. Tehran: Firuzkuh, 17 km towards Semnan, 2050 m, Ghaffari 2875, n = 12. Section Cynaroideae comprises more than 50 species, mainly in the western area of the Flora Iranica region, except for C. onopordioides which is widespread in this range. We found 12 bivalents at metaphase I in this taxon (Fig. 9). This count agrees with a previous report of 2n = 24 by Tscherneva (1985). All previous counts in species of sect. Cynaroideae indicate the same basic number of x = 12 (AfzalRafii, 1980; Ghaffari, 1984, 1986; Tscherneva, 1985; Ghaffari et al., 2000; Susanna et al., 2003b). COUSINIA SECT. Cousinia gmelini C. Winkl. Mazandaran: Pol-e-zanguleh, 2065 m, Ghaffari 2976, n = 12. This taxon is endemic to a limited area in south Mazandaran Province. Meiosis in this species showed 12 bivalents (Fig. 10). According to our data, this is the first chromosome count for this species. The other two studied species of this section are C. franchetii C. Winkl., with 2n = 26 (Tscherneva, 1985), and C. caespitosa C. Winkl., with 2n = 22 (Susanna et al., 2003b). Thus, sect. Eriocousinia seems to have three different basic numbers (x = 11, 12 and 13), which merits further investigation. COUSINIA COUSINIA SECT. ERIOCOUSINIA TSCHERNEVA SECT. HELIANTHAE BUNGE CYNAROIDEAE BUNGE Cousinia behboudiana Rech. & Esfand. Tehran: Damavand, towards Tar lake, 2320 m, Ghaffari 2775, n = 12. Tehran: Haraz road, between Rineh and Larijan, 2100 m, Ghaffari 2675, n = 12. This species is endemic to Iran, and is distributed only in the north of Tehran province. Two collections of this taxon were studied and both had n = 12 (Fig. 7). This is the first chromosome count for this species. Cousinia keredjensis Bornm. & Gauba Tehran: Kuh-Dshtae, between Tehran and Karaj, 1730 m, Ghaffari 1575, n = 12. This taxon is endemic to Iran and is found in a limited area of Karaj and Qazvin (towards Karaj). We found 12 bivalents at first metaphase, but in some cells one or two quadrivalents were observed (Fig. 8). In diakinesis, two bivalents were associated with the nucleolus. According to our data, this is the first chromosome count for this species. Cousinia heliantha Bunge Khorasan: between Kashmar and Nayshabur, 1150 m, Ghaffari 5656, n = 13. This section comprises only two species: C. heliantha in Iran (Rechinger, 1972, 1979) and C. spryginii Kult. in the ancient Soviet Union (Tscherneva, 1962). C. heliantha is a narrow endemic to Iran, growing in a small area of Khorasan. A previous somatic count of 2n = 26 (Ghaffari, 1984) is the same as our present result of n = 13 (Fig. 11). To date, sect. Helianthae has the basic chromosome number of x = 13. COUSINIA SECT. LASIANDRA BUNGE Cousinia aitchisonii C. Winkl. Cistan and Baluchestan: Taybad, 40 km towards Torbat-e-Jaam, 830 m, Ghaffari 2356, n = 13. Section Lasiandra comprises five species, most of them growing in Afghanistan (Rechinger, 1972). Meiosis in this taxon showed 13 bivalents at metaphase I © 2006 The Linnean Society of London, Botanical Journal of the Linnean Society, 2006, 151, 411–419 Downloaded from https://academic.oup.com/botlinnean/article-abstract/151/3/411/2420402 by guest on 28 July 2018 KARYOLOGY OF IRANIAN COUSINIA (Fig. 12). According to our data, this is the first chromosome count for this species. Cousinia lasiandra Bunge Khorasan: Mashhad, 53 km towards Sarakhs, 507 m, Ghaffari 1256, n = 13. This taxon is endemic to Iran. Meiosis in this species showed 13 bivalents at metaphase I (Fig. 13). Chromosome segregation at anaphase I was 13–13 (Fig. 14). According to our data, this is the first chromosome count for this species. To date, sect. Lasiandra has the basic number of x = 13. COUSINIA SECT. LEIOCAULES BUNGE Cousinia arctotidifolia Bunge Khorasan: between Taybad and Torbat-e-Jaam, 830 m, Ghaffari 2456, n = 12. According to Rechinger (1972), Iran has six species of this section. Meiosis in this species was regular, with 12 bivalents at metaphase I (Fig. 15). In diakinesis, three bivalents were associated with the nucleolus. Previous somatic chromosome counts of 2n = 24 (Ghaffari, 1984, on Iranian populations; Tscherneva, 1985, on a sample from Turkmenistan) agree with the present count of n = 12. (Aryavand, 1976; Tscherneva, 1985; Susanna et al., 2003b). Discordances in the delineation of sect. Microcarpae are evident from the molecular study by Susanna et al. (2003a). COUSINIA SECT. BOISSIERI BUHSE Zanjan: 20 km towards Miyaneh, 1550 m, Ghaffari 19175, 2n = 24. This taxon is endemic to north-west Iran. Mitotic prophase of parenchyma cells of the anthers indicates 2n = 24 (Fig. 16). This is the first chromosome count for this species. The basic chromosome number of x = 12 for sect. Leiocaules is a confirmation of other counts (Ghaffari, 1984; Tscherneva, 1985; Susanna et al., 2003b). COUSINIA SECT. MICROCARPAE BUNGE Cousinia microcarpa Boiss. Tehran: Damavand, Simindasht, 1800 m, Ghaffari 1970, n = 13. Khorasan: between Fariman and Torbat-e-Jaam, 1030 m, Ghaffari 2156, n = 13. Previous counts for this taxon are 2n = 26 (Koul, 1964; Podlech & Dieterle, 1969; Ghaffari, 1984; Tscherneva, 1985; Susanna et al., 2003b) and n = 10 by Mehra & Remanandan (1969), which is questionable. Two collections of this species were examined, both showing 13 bivalents at first metaphase (Fig. 17). Also, chromosome segregation at anaphase I was 13– 13. Other counts in sect. Microcarpa indicate three different basic numbers for this section, x = 11, 12 and 13 MICROCOUSINIA TSCHERNEVA Cousinia prolifera Jaub. & Spach Khorasan: Torbat-e-Jaam, 930 m, Ghaffari 2256, n = 10. Tehran: Karaj, Mardabad, 1190 m, Ghaffari 1475, n = 10. A previous report of n = 9 in this species (Ghaffari, 1986) was probably erroneous. Meiosis in the two collections of this study was regular and showed 10 bivalents at metaphase I (Fig. 18). Chromosome segregation at anaphase I was also 10–10. The chiasma frequency determined from 15 cells was 1.30 per bivalent at metaphase I. The other purported species of this section, C. minuta Boiss., which also has x = 10 (Mehra et al., 1965), is a synonym of C. prolifera according to Rechinger (1972, 1979). Thus, the monotypic section Microcousinia has the basic chromosome number x = 10. COUSINIA COUSINIA 415 SECT. MYRIOTOMAE RECH. F. Cousinia candolleana Jaub. & Spach Zanjan: between Abgarm and Avaj, 1950 m, Ghaffari 7167, n = 9. This species is endemic to Iran. Its meiosis was regular and showed nine bivalents at metaphase I (Fig. 19). This is the first time the basic number x = 9 has been reported in Cousinia, because previous reports of x = 9 by Ghaffari & Djavadi (1998) were corrected to other basic numbers. As the only other count in this section was 2n = 26 in C. glandulosa Kult. (Chuksanova in Fedorov, 1969), our result suggests that there are two different basic numbers in sect. Myriotomae: x = 9 and 13. More studies are needed in this section to confirm the new basic number. COUSINIA SECT. PLATYACANTAE RECH. F. Cousinia trachyphyllaria Bornm. & Rech. f. Khorasan: between Mashhad and Torbat-Haydariyeh, 1215 m, Ghaffari 3656, n = 13. This section has seven representatives in Iran, five of which are endemic. Thirteen bivalents were observed at metaphase I (Fig. 20), which agrees with a previous somatic report of 2n = 26 (Ghaffari, 1984). © 2006 The Linnean Society of London, Botanical Journal of the Linnean Society, 2006, 151, 411–419 Downloaded from https://academic.oup.com/botlinnean/article-abstract/151/3/411/2420402 by guest on 28 July 2018 416 S. M. GHAFFARI ET AL. 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 © 2006 The Linnean Society of London, Botanical Journal of the Linnean Society, 2006, 151, 411–419 Downloaded from https://academic.oup.com/botlinnean/article-abstract/151/3/411/2420402 by guest on 28 July 2018 KARYOLOGY OF IRANIAN COUSINIA COUSINIA SECT. SCIADOCOUSINIA TSCHERNEVA Cousinia eryngioides Boiss. Tehran: Sorkh-e-Hesar, 1300 m, Ghaffari 1576, n = 11. Hamadan: 10 km towards Malayear, 1560 m, Ghaffari 4275, n = 11. Khorasan: between Bojnourd and Shirvan, 1100 m, Ghaffari 256, n = 11. This taxon is distributed in Iran and Turkmenistan. The three collections of this species showed 11 bivalents at metaphase I (Fig. 21), which agrees with a previous report of 2n = 22 (Ghaffari, 1984). Chromosome segregation at first anaphase was 11–11 (Fig. 22). In diakinesis, three bivalents were associated with the nucleolus. The only other count for this taxon, of 2n = 18 by Chuksanova (in Fedorov, 1969), is questionable. COUSINIA SECT. STENOCEPHALAE BUNGE Cousinia alexeenkoana Bornm. Hamadan: between Razan and Avaj, Soltanboolagh, 1750 m, Ghaffari 2975, n = 13. This species is endemic to Iran. Meiosis in this taxon showed 13 bivalents at metaphase I (Fig. 23). At diakinesis, three bivalents were associated with the nucleolus. According to our information, this is the first chromosome count for this species. Cousinia lucida DC. Loristan: Azna, 15 km towards Dorud, 1600 m. Ghaffari 5877, n = 13. This taxon is endemic to western Iran. Meiosis in this species showed 13 bivalents, with three associated with the nucleolus in diakinesis (Fig. 24). Our results agree with a previous report of 2n = 26 (AfzalRafii, 1980). All counts indicate x = 13 for section Stenocephalae. COUSINIA SECT. 417 Cousinia irritans Rech. f. Tehran: Damavand, between Deh-e-chenar and Tar lake, 2750 m, Ghaffari 3075, n = 13. This species is endemic to Iran. The gametic chromosome number in this taxon was n = 13, with regular monad segregation at anaphase II of 13–13 + 13–13 (Fig. 25). This is also the first chromosome count for this species. Cousinia hypoleuca Boiss. Tehran: Karaj, Shahrestanak, 2300 m, Ghaffari 3975, n = 13. Tehran: between Ab-ali and Emamzadeh-Hashem, 2600 m, Ghaffari 3675, n = 13. This taxon is endemic to Iran. Two collections of this species were studied, both with n = 13 (Fig. 26). Chromosome segregation at anaphase I and II indicates 13 dyads and monads at each pole, respectively. The chiasma frequency determined from 28 cells was 1.35 per bivalent at metaphase I. Our count agrees with a previous count of 2n = 26 by Afzal-Rafii (1980). Cousinia crispa Jaub. & Spach Mazandaran: Kandavan Mt., 2750 m, Ghaffari 4175, n = 13. Tehran: between Polur and Rineh, 2305 m, Ghaffari 3375, n = 13. This taxon is endemic to Iran. Two collections of this species were studied and showed 13 bivalents at metaphase I and diakinesis. Occasionally, in some cells, quadrivalents or hexavalents were observed at metaphase I (Fig. 27). This count agrees with one previous report of 2n = 26 by Afzal-Rafii (1980). Cousinia pinarocephala Boiss. Tehran: between Semshak and Dizin, 2350 m, Ghaffari 4075, n = 12. This species is endemic to Iran. Meiosis in this taxon was regular and showed 12 bivalents at first metaphase (Fig. 28). According to our records, this is the first chromosome count for this species. SERRATULOIDEAE BUNGE The flora of Iran has 11 species of this section, ten of which are endemic. This section is restricted to the north of Iran. Our results indicate two different basic numbers for sect. Serratuloideae: x = 12 and 13. COUSINIA SPECIES OF INDETERMINATE SECTION Four species were examined, our counts being the first in every case. Figures 17–32. Meiosis in Cousinia spp. Fig. 17. C. microcarpa, metaphase I, n = 13. Fig. 18. C. prolifera, metaphase I, n = 10. Fig. 19. C. candolleana, metaphase I, n = 9. Fig. 20. C. trachyphyllaria, metaphase I, n = 13. Figs 21 and 22. C. eryngioides, metaphase I and anaphase I, n = 11. Fig. 23. C. alexeenkoana, metaphase I, n = 13. Fig. 24. C. lucida, diakinesis, n = 13. Fig. 25. C. irritans, anaphase II, showing 13–13 + 13–13 monad segregation. Fig. 26. C. hypoleuca, metaphase I, n = 13. Fig. 27. C. crispa, metaphase I, n = 13. Fig. 28. C. pinarocephala, metaphase I, n = 12. Fig. 29. C. meshhedensis, metaphase I, n = 13. Fig. 30. C. pichleriana, metaphase I, n = 12. Fig. 31. C. raphiostegia, anaphase I, showing 13–13 segregation. Fig. 32. C. trachylepis, metaphase I, n = 13, showing two hexavalents (arrows). Scale bar = 10 µm. © 2006 The Linnean Society of London, Botanical Journal of the Linnean Society, 2006, 151, 411–419 Downloaded from https://academic.oup.com/botlinnean/article-abstract/151/3/411/2420402 by guest on 28 July 2018 418 S. M. GHAFFARI ET AL. Cousinia meshhedensis Bornm. & Rech. f. Khorasan: Nayshabour, 60 km towards Kashmar, 1195 m, Ghaffari 6156, n = 13. This taxon is endemic to Iran. Meiosis in this species showed 13 bivalents at metaphase I (Fig. 29). Cousinia pichleriana Bornm. ex Rech. f. Markazi: Arak, Senejan, 1800 m, Ghaffari 3875, n = 12. This taxon is endemic to Iran. Twelve bivalents at metaphase I were observed. Anaphase I showed 12–12 chromosome segregation (Fig. 30). Cousinia raphiostegia Rech. f. Khorasan: Ghaen, 5 km towards Gonabad, 1440 m, Ghaffari 4256, n = 13. This taxon is a narrow endemic to Iran. The gametic number in this species was n = 13, which was observed at various stages of meiosis (Fig. 31). Cousinia trachylepis Bunge Semnan: Shahrud, Mojen, 1850 m, Ghaffari 3276, n = 13. This species is a narrow endemic, distributed in a limited area around Shahrud. Meiosis in this species showed 13 bivalents at metaphase I, but, in some cells, one or two quadrivalents or hexavalents were observed (Fig. 32), which could indicate a hybridization event in this population. CONCLUDING REMARKS Basic chromsome numbers ranging from x = 13 to x = 11 were most frequently found in our study, as was the case in previous reports. The two other basic numbers, x = 10 and x = 9, were infrequent, and the latter should be confirmed by extending the studies to the rest of the species of sect. Myriotomae. With regard to chromosome evolution in Cousinia, Susanna et al. (2003b) suggested some correlations between molecular data and dysploidy. However, the confirmation of the basic numbers x = 10 and x = 9 makes their correlations and their outlined karyological evolution hypothesis highly provisional, because no species with either of these two low chromosome numbers has been included so far in any molecular analysis. Descending dysploidy has been confirmed in other groups, as in the closely related subtribe Centaureinae (Garcia-Jacas et al., 2001), and this is also probably the general trend in Cousinia. However, in Cousinia, we are very far from the level of karyological and molecular knowledge that we have in the Centaureinae. It is mandatory to extend our karyological studies as much as possible, given the potential of karyology in the systematics of the genus Cousinia. ACKNOWLEDGEMENTS This work was supported by a grant (project 6401011/ 1/01) from the Research Council of the University of Tehran. 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