Vijay Kumar Singhal

Impact of cytomixis on meiosis, pollen viability and pollen size in wild populations of Himalayan poppy (Meconopsis aculeata Royle)

We report the occurrence of cytomixis in wild populations of Himalayan poppy (Meconopsis aculeata Royle), which is considered to be an important and threatened medicinal plant growing in the high hills of the Himalayas. The impact of cytomixis on meiotic behaviour, reduced pollen viability and heterogeneous-sized pollen grains was also studied. Cytological studies in the seven wild populations from the high hills of Himachal Pradesh revealed that all the Himalayan populations exist uniformly at the tetraploid level (2n=56) on x=14. The phenomenon of chromatin transfer among the proximate pollen mother cells (PMCs) in six populations caused various meiotic abnormalities. Chromatin transfer also resulted in the formation of coenocytes, aneuploid, polyploid and anucleated PMCs. Among individuals that showed chromatin transfer, chromosome stickiness and interbivalent connections were frequently observed in some PMCs. The phenomenon of cytomixis in the species seems to be directly under genetic control; it affects the meiotic course considerably and results in reduced pollen viability.

Cytomixis and associated meiotic abnormalities affecting pollen fertility in Clematis orientalis

Present cytological investigations from the cold desert regions of Lahaul-Spiti and Kinnaur (India) record the first ever tetraploid (2n=32) chromosome count and cytomixis in Clematis orientalis L. var. acutifolia Hook. f. et Thoms. The phenomenon of cytomixis (9.33–29.80 %) involving chromatin transfer among 2–3 proximate pollen mother cells (PMCs) during male meiosis occurs through narrow and broad cytoplasmic channels from early prophase to tetrad stage. However, frequency of its occurrence during the later meiotic stages is rather low. Chromatin transfer results into the formation of hypo-, hyperploid and enucleated PMCs. Various meiotic abnormalities associated with cytomixis such as chromatin stickiness, pycnotic chromatin, interbivalent connections, out of plate bivalents, late disjunction of bivalents, and laggards and bridges resulted into some pollen sterility (16.33–49.30 %) and heterogeneous pollen grains size.

Impaired Male Meiosis due to Irregular Synapsis Coupled with Cytomixis in a New Diploid Cytotype of Dianthus angulatus (Caryophyllaceae) from Indian Cold Deserts

Dianthus angulatus (Caryophyllaceae) is cytologically examined here for the first time for the area of India. The diploid chromosome count of 2n = 30, ascertained here, represents a new cytotype, supplementing the earlier report of a hexaploid cytotype with 2n = 90 from outside of India. We report here the occurrence of two plants showing impaired meiosis due to irregular synapsis and cytomixis collected from Kinnaur district, Himachal Pradesh (India). The other plants of this species collected in Lahaul-Spiti region showed normal male meiosis (n = 15) with a high (95%−100%) pollen fertility and normal seed set, and they reproduce sexually. Irregular synapsis in two plants from the Kinnaur region is characterized by the complete absence of chromosome pairing and the presence of 30 univalents at diakinesis and meta-anaphase. In addition, other meiotic irregularities were found, such as unoriented chromosomes, laggards, precocious movements of univalents at anaphase-I and micronuclei at telophase. Microsporogenesis was also abnormal, resulting in the formation of monads, dyads, triads, polyads, and tetrads with micronuclei. The occurrence of intra- and intermicrosporal chromatin material transfer during microsporogenesis was observed, which is a rather rarely observed phenomenon. The synaptic irregularities coupled with chromatin transfer in these plants seem to be responsible for the high pollen sterility (38%−42%) and heterogeneously sized pollen grains. In these plants no seeds were set, and plants reproduced vegetatively through root suckers.

Persistent occurrence of meiotic abnormalities in a new hexaploid cytotype of Thalictrum foetidum from Indian cold deserts

Thalictrum foetidum L. (Ranunculaceae), a morphologically variable and widely distributed species of temperate and alpine Himalayas is worked out cytologically for the first time from India. Earlier studies from outside India were restricted to chromosome counts and karyotypic analysis. We studied the male meiosis, microsporogenesis and pollen viability in the wild accessions from the cold deserts of Lahaul-Spiti, Kinnaur and Pangi Valley of Himachal Pradesh. Present cytomorphological surveys in the species record the existence of two distinct morphotypes involving plant size; colour and size of leaf/leaflet; dentation of leaflet lobes; and degree of leaf pubescence. All the accessions in the two morphovariants share the same meiotic chromosome number (n = 21) and adds a new intraspecific hexaploid cytotype. The accessions show the phenomenon of cytomixis involving transfer of chromatin material among proximate pollen mother cells (PMCs) and associated meiotic abnormalities like, out of plate bivalents, interchromosomal connections, and laggards, bridges and micronuclei at anaphases/telophases. Microsporogenesis results into abnormal sporads (tetrads with micronuclei, dyads, triads and polyads). The products of such sporads resulted into some pollen sterility and pollen grains of heterogeneous sizes. The persistent occurrence of phenomenon of cytomixis and associated meiotic abnormalities and consequently pollen sterility and pollen grains of heterogeneous sizes in the hexaploid cytotype of T. foetidum seems to be under some genetic factors associated with the genome.

Syncytes during male meiosis resulting in 2n pollen grain formation in Lindelofia longiflora var. falconeri

Abstract  Lindelofia longiflora (Royle ex Benth.) Baill. var. falconeri (Cl.) Brand (Family: Boraginaceae) is investigated cytologically (n= 12) for the first time from the cold deserts of Pangi Valley, Chamba District (Himachal Pradesh) in India. We report the formation of syncytes and 2n pollen grains in the species. During meiosis, the majority of the pollen mother cells (PMCs) exhibited 12 bivalents, equal segregation of chromosomes during anaphases, regular tetrads, and normal‐sized pollen grain formation. Occasionally, two proximate PMCs fused during the early stages of prophase‐I and resulted in the formation of syncytes. The frequency of syncytes in the accession is rather low, at 25 out of 1866 (1.33%). Such syncyte PMCs are detectable during meiosis due to their larger size compared to typical PMCs. The syncytes or polyploid cells showed normal 24 bivalents and depicted perfectly regular meiotic course. But the products of such PMCs yield 2n or larger sized pollen grains that are almost double the size of typical normal or n pollen grains. The origin of syncytes as a consequence of the fusion of meiocytes during the early stages of meiosis‐I could be attributed to low temperature stress conditions prevailing in the Pangi Valley, where temperature during May and June dip to below freezing, the time the plants enters the reproductive/flowering bud stage. It is possible that such apparently fertile 2n pollen grains originating from syncytes might play a role in the origin of intraspecific polyploids in the species.

Structural heterozygosity and cytomixis driven pollen sterility in Anemone rivularis Buch.-Ham. ex DC. from Western Himalaya (India)

Meiotic studies have been carried out on 12 accessions of Anemone rivularis from different regions of Western Himalaya. The accessions from Chamba, Kullu and Kinnaur districts of Himachal Pradesh and Uttarkashi district of Uttarakhand showed a normal meiotic course leading to high pollen fertility (87–100%). However, the accession scored from Saat Tal, Uttarakhand showed structural heterozygosity for reciprocal translocations. This is the first report of occurrence of structural heterozygosity in the species. Out of the 604 analysed pollen mother cells (PMCs), 38.41% (232) showed quadrivalent formation at diakinesis and metaphase-I. Comparison of chiasma frequency was also performed among selected populations using one-way analysis of variance (ANOVA), which showed significant differences. The occurrence of cytomixis involving inter PMC transfer of chromatin material among neighbouring meiocytes (45.68%) was also observed. PMCs at anaphases/telophases showed laggards (31.25%) and chromatin bridges at A-I/T-I and A-II/T-II (21.02%) leading to abnormal microsporogenesis. Consequently, abnormal sporads such as triads (66.67%), polyads (3.33%) and triads with micronuclei (0.83%) were formed. Structural heterozygosity and inter PMC chromatin material transfer in the individuals of the species seem to be responsible for the reduction in pollen fertility (67.8%) and formation of pollen grains of heterogeneous sizes.

Reduction in chiasma frequency and pollen fertility due to multiple chromosomal associations and univalents in Saxifraga diversifolia from alpine regions of northwest Himalayas (India)

Two varieties of Saxifraga diversifolia, diversifolia and parnassifolia, collected from the moist alpine slopes around Gauri Kund, 3930 m (Manimahesh Hills, Himachal Pradesh, India), are studied for detailed male meiosis, chiasma frequency and pollen fertility. Both varieties exist at diploid level (based on x = 8), and show a meiotic chromosome count of n = 8 at diakinesis and metaphase-I, and regular chromosome distributions at anaphases-I/II. The meiotic chromosome count of n = 8 ascertained here represents a new aneuploid cytotype, supplementing the earlier report of a diploid cytotype with 2n = 20 from the north-west Himalayas in India and the Nepal Himalayas. Of the two varieties, var. diversifolia showed the presence of multiple chromosomal associations and univalent chromosomes at diakinesis and metaphase-I of meiosis-I. On the other hand, var. parnassifolia does not have multivalent formation but showed only 2–4 univalent chromosomes at diakinesis and metaphase-I. Occurrence of univalents in pollen mother cells of var. diversifolia and parnassifolia reduced the chiasma frequency significantly and also caused some pollen sterility (7–8%). The paper herein discusses for the first time the occurrence of structural heterozygosity and univalent chromosomes and their apparent affect on chiasma frequency and pollen fertility in S. diversifolia.

Erratic Male Meiosis Resulting in 2n Pollen Grain Formation in a 4x Cytotype (2n = 28) of Ranunculus laetus Wall. ex Royle

Two accessions were studied for male meiosis in Ranunculus laetus from the cold regions of Northwest Himalayas. One accession showed the presence of 14 bivalents at diakinesis and regular segregation of bivalents at anaphase I which lead to normal tetrad formation with four n microspores and consequently n pollen grains and 100% pollen fertility. Second accession from the same locality revealed the erratic meiosis characterized by the presence of all the 28 chromosomes as univalents in meiocytes at metaphase I. Univalent chromosomes failed to segregate during anaphases and produced restitution nuclei at meiosis I and II. These restitution nuclei resulted into dyads and triads which subsequently produced two types of apparently fertile pollen grains. On the basis of size, the two types of pollen grains were categorized as n (normal reduced) and 2n (unreduced, 1.5-times larger than the n pollen grains). The estimated frequency of 2n pollen grains from dyads and triads (61.59%) was almost the same as that of the observed one (59.90%), which indicated that 2n pollen grains in R. laetus were the result of dyads and triads. The present paper herein may provide an insight into the mechanisms of the formation of various intraspecific polyploids through sexual polyploidization in R. laetus.

Cytology of woody species

Cytological studies have been made on 59 woody species of Polypetalae. Two genera (Gynocardia odorata,n=23;Pahudia martabanica,n=12) and 9 species (Talauma candollei,n=19;Cratoxylon polyanthum,n=11;Sterculia villosa,n=20;Grewia hirsuta,n=9;Millettia brandisiana, 2n=22+0−2B;Phanera glauca,n=14;Terminalia oliveri,n=12;Psidium coriaceum, 2n=77;P. pumilum,n=11) are counted for the first time. Additional and/or varied chromosome numbers are recorded for 8 species. Presence ofB-Chromosomes has been detected inCrataeva nurvala (n=13+0−3B),Erythrina caffra (n=21+0−3B) andMillettia brandisiana (2n=22+0−2B). Existence of some multivalents in the tetraploid taxa ofEugenia jambolana (2n=44) andHydnocarpus laurifolia (2n=48) indicates their segmental alloploid nature. But the presence of cent per cent trivalents (11III) in the triploid cytotype ofEugenia uniflora (2n=33) reveals the autotriploid nature. Presence of some univalents in diploid taxa ofMillettia brandisiana (2n=22) may be due to asynapsis and/or desynapsis. Variation in chromosome number inPMCs and some pollen sterility inHydnocarpus kurzii seems to be the consequence of cytomixis. Abnormalities in microsporogenesis in one of the cultivated trees ofFirmiana pallens (2n=40) are due to spindle irregularities.

Uses of Local Plant Biodiversity among the Tribal Communities of Pangi Valley of District Chamba in Cold Desert Himalaya, India

Pangi Valley is the interior most tribal area in Himachal Pradesh of Northwest Himalaya. An ethnobotanical investigation is attempted to highlight the traditional knowledge of medicinal plants being used by the tribes of Pangi Valley. Various localities visited in the valley 2-3 times in a year and ethnobotanical information was collected through interviews with elderly people, women, shepherds, and local vaids during May 2009 to September 2013. This paper documented 67 plant species from 59 genera and 36 families along with their botanical name, local name, family name, habit, medicinal parts used, and traditional usage, including the use of 35 plants with new ethnomedicinal and other use from the study area for the first time. Wild plants represent an important part of their medicinal, dietary, handicraft, fuel wood, veterinary, and fodder components. These tribal inhabitants and migrants depend on the wild plant resources for food, medicines, fuel, fibre, timber, and household articles for their livelihood security. The present study documents and contributes significant ethnobotanical information from the remote high altitude and difficult region of the world, which remains cut off from rest of the world for 6-7 months due to heavy snowfall.