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Meiotic and Ethnobotanical Studies on Rheum Species from Kashmir Himalaya

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Meiotic and Ethnobotanical Studies on Rheum Species from Kashmir Himalaya © 2016 The Japan Mendel Society Cytologia 81(3): 295–300 Meiotic and Ethnobotanical Studies on Rheum Species from Kashmir Himalaya Shagoon Tabin1, Kuljit Kaur1, Vijay Singh1, Azra Nahaid Kamili2 and Raghbir Chand Gupta1* 1 Department of Botany, Punjabi University, Patiala 147002, Punjab, India 2 Centre of Research for Development University of Kashmir, Srinagar 190006, J&K, India Received October 23, 2015; accepted June 11, 2016 Summary The Rheum species (R. emodi, R. spiciforme, and R. webbianum) are endangered species and have vanished from lower altitudes. Meiotic studies of the three species show that the polyploidy is reported only in R. webbianum, as it depicts two cytotypes, i.e., diploid (2x) in population from Panzila Top and tetraploid (4x) from Parkhachik and Tangole, of which the diploid cytotype is reported for the first time in this species. Moreover, a diploid cytotype (2x) in R. spiciforme is reported for the first time in India. Further, meiotic course is abnormal in all the studied populations, as all show meiotic abnormalities in the form of cytomixis, chromatin stickiness, unoriented bivalents, chromatin bridges and laggards, leading to abnormal microsporogensis, ultimately affecting pollen viability. Ethnobotanical information depicts the use of different parts of all the Rheum species by local people to control many diseases like piles, constipation, skin eruptions, intestinal infection, wounds, joint pains and also used as food. Key words Cytotype, Endangered species, Ethnobotany, Meiosis, Meiotic abnormality, Rheum. The genus Rheum L. (commonly called Rhubarb) nology may lead to the development of new pharmaceu- belongs to the family Polygonaceae of group Monochla- tical drugs. mydae and is represented by 60 species spread through- out the world (Anjen et al. 2003), of which only seven Materials and methods species have been reported from the Indian subcontinent (Ganie et al. 2014). According to Stewart (1972), all the Meiotic study seven species (including R. emodi, R. spiciforme, and Young flower buds were collected during day time R. webbianum) reported from the Indian subcontinent and fixed in standard Carnoy’s fixative (6 : 3 : 1=abso- are also present in the Kashmir Himalaya. A total of lute alcohol : chloroform : glacial acetic acid v/v/v) for 560 plant species of India are included in the Red List 24 h and preserved in 70% alcohol at 4°C. The collected of Threatened Species by IUCN including Rheum spe- buds were subjected to detailed meiotic analysis for cies (Phartyal et al. 2002). Rheum emodi is included in chromosome number determination, meiotic course, critically endangered species. Rheum, commonly known microsporogenesis, pollen fertility percentage, and pol- as Rhubarb, is a strong vigorous plant and can live long len grain size. In each case more than 20 fresh slides even in tough conditions. Many different species of were prepared from different anthers/flowers of more Rheum are distributed in the temperate and sub-tropical than one plant, using a standard 1% acetocarmine regions of Himalaya from Kashmir to Sikkim (Nautiyal technique. Chromosome numbers were determined at et al. 2003) and growing wild in the mountains of the diakinesis, metaphase-I (M-I), and anaphase I/II (A-I/II) Western and North-Western provinces of China and from freshly prepared slides. For the sake of confirma- adjoining Tibetan territory. In taxonomical focus, inflo- tion of any observation, about 100 pollen mother cells rescence is pyramidal or spherical in which flowers are (PMCs) were analyzed at various stages of meiosis from arranged in panicles and pedicel articulates at or below prophase-I to telophase-II. Pollen fertility was estimated the middle. by mounting pollen grains in 50% glycero–acetocar- The ethnobotanical information provided by tribal mine (1 : 1) as proposed by Marks (1954). Well-filled and local people may provide clues for specific medici- and stained pollen grains were considered as fertile and nal properties and the presence of specific biochemicals, shrivelled or unstained pollen grains as sterile. Measure- which when explored with modern phytochemical tech- ments of pollen grain size were taken using micrometry and a Nikon Eclipse 80i Imaging System. Microphoto- * Corresponding author, e-mail: [email protected] graphs of PMCs depicting various meiotic stages and DOI: 10.1508/cytologia.81.295 pollen grains were taken from freshly prepared tempo- 296 S. Tabin et al. Cytologia 81(3) Fig. 1. Meiotic chromosome number in different species of Rheum from Kashmir Himalaya. a, b: Rheum emodi (2n=22): PMC at M-I showing 11II. PMC at A-I showing 11 : 11 distribution of chromosomes. c: Rheum spiciforme (2n=22): PMC at M-I showing 11II. d, e, f: Rheum webbianum (2n=22, 44): PMC at M-I showing 11II, PMC at A-I showing 11 : 11 distribution of chromosomes, PMC at A-I showing 22 : 22 distribution of chromosomes. rary slides using Leica Qwin and Nikon Eclipse 80i mi- bridges at anaphases/telophases (Fig. 2f, i), and unori- croscope Digital Imaging Systems. The specimens were ented bivalents at M-I (Fig. 2c), leading to abnormal mi- submitted to the Herbarium, Department of Botany, crosporogenesis with the presence of monads, triads, and Punjabi University, Patiala and University of Kashmir, tetrads with micronuclei (Fig. 2n, o, q), which ultimately Kashmir. The data regarding the cytologically known leads to the decrease in pollen fertility (72.80%). species, number of cytotypes, and frequency of poly- ploids of a particular genus has been compiled world- Rheum spiciforme Royle wide as well as in India from various Chromosomal Cytologically, both populations of the species showed Atlases and Indexes to Plant Chromosome Numbers by a diploid chromosome count 2n=22 (Fig. 1c), which was Darlington and Wylie (1955), Fedorov (1974), Moore elucidated cytologically for the first time from India and (1973, 1974, 1977), Goldblatt (1981, 1984, 1985, 1988), is in accordance with the reports by Jaretzky (1928) and Goldblatt and Johnson (1990, 1991, 1994, 1996, 1998, Edman (1929) from outside India (Table 1). The detailed 2000, 2003), Kumar and Subramaniam (1986), and Kha- course of meiosis was abnormal with the presence of toon and Ali (1993), various journals, the Internet, as various irregularities like laggards and chromatin bridg- well as the species in this study. es at A-I/T-I (Fig. 2g, j), chromatin stickiness at M-I (2l), unoriented bivalents at M-I (Fig. 2d), and cytomixis Ethnobotany between different PMCs at M-I (Fig. 2b), which leads to The surveys and collection of Rheum species was abnormal microsporogenesis with the presence of micro- mainly on high altitudes, and information was taken nuclei in tetrads (Fig. 2r) and resulted in decreased pol- from gujjars, bakkerwals, and some local people. For len fertility (78.23%). gaining knowledge and information on medicinal plants, these people were interviewed during the course of the Rheum webbianum Royle present study. During meiotic course, one accession of the species collected from Panzila Top (Zanaskar area) showed the Results presence of diploid cytotype showing 11 bivalents at M-I and 11 : 11 distribution of chromosomes at A-I (Fig. Rheum emodi Wall. 1d, e). Two accessions collected from Parkhachik and The meiotic studies of the present accessions reveal Tangole (Kargil district) are tetraploid showing 22 : 22 the presence of a diploid chromosome count 2n=22 (Fig. distribution of chromosomes at A-I (Fig. 1f). Previously, 1a, b) (Table 1), which is in accordance with the previ- the tetraploid cytotype (2n=44) of the species has also ous reports by various workers from India (Gohil and been reported by Gohil and Rather (1986), and Saggoo Rather 1986, Saggoo and Farooq 2011). Further, the and Farooq (2011) from Kashmir Himalaya (Table 1), meiotic course was abnormal with the presence of chro- whereas, 2n=22 adds a new diploid cytotype for the matin transfer at M-I (Fig. 2a), laggards and chromatin first time on a worldwide basis and reports the presence 2016 Meiotic and Ethnobotanical Studies on Rheum Species from Kashmir Himalaya 297 Fig. 2. Meiotic abnormalities in the studied species of Rheum from Kashmir Himalaya. a, b: PMCs showing chromatin transfer. c–e: PMCs at M-I showing unoriented bivalents. f–h: PMCs showing laggards at A-I and T-I. i–k: PMCs showing chro- matin bridges at A-I and T-I. l, m: PMCs showing chromatin stickiness at M-I. n: Monad. o, p: Triad. q, r: Tetrads with micronuclei. s: Fertile and sterile pollen grains. of intraspecific polyploidy. The detailed meiotic course was collected by interacting with local and tribal people. of the diploid cytotype shows various abnormalities as During the surveys, it was observed that tribal people unoriented bivalents at M-I (Fig. 2e), chromatin laggards used to preserve and conserve the Rheum species for and bridges at anaphases/telophases (Fig. 2h, k), and their living. After interviews of local people of high al- chromatin stickiness at M-I (Fig. 2m) (Table 2). These titude, it was observed that Rheum species are used for abnormalities lead to abnormal microsporogenesis with many diseases listed in Table 4. triads and tetrads with micronuclei (Table 3) and result- ed in the reduction of pollen fertility (69.15%), whereas Discussion the tetraploid cytotype shows a normal meiotic course with high pollen fertility (97.38%). Chromosome number The genus is comprised of 60 taxonomically known Ethnobotany species, out of which 42 species/51 cytotypes are cyto- Information regarding local uses of Rheum species logically elucidated worldwide. It is a monobasic genus 298 S. Tabin et al. Cytologia 81(3) Table 1. Data containing accession numbers, locality with altitude, meiotic chromosome number, ploidy level, meiotic course, pollen size, pollen fertility, and previous reports with the remarks of presently studied three species of Rheum from Kashmir Himalaya. Meiotic Ploidy level S.
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