Propagation of Costus speciosus (Koen.) Sm. through in vitro rhizome production

Rhizomes developed in vitro on shoots of Costus speciosus (Koen.) Sm. which were initiated from zygotic embryos. The effect of different hormonal and nutritional additions to Schenk and Hildebrandt' s (SH) basal nutrient medium on rhizome development was studied. Rhizomes developed on SH basal salts but formed with increased frequency on medium supplemented with adenine sulfate, casamino acids (CA) and various combinations of cytokinins and auxins. Incubation in light was necessary for rhizome formation. Isolated basal as well as nodal (aerial) rhizomes formed and produced new shoots. In basal medium without any growth additives (control) the average number of shoots produced per inoculum was 3.3 ±0.73 whereas in the best suited medium i.e. supplemented with 250 mg l^–1 CA the number of shoots obtained was 22.7±1.92. There was no dormancy period for rhizomes under the cultural conditions investigated. Rhizome-sprouts were easily transplanted from the in vitro conditions to the field. More than five hundred propagules (i.e. sprouted-rhizomes) were obtained within 5 months and it has been estimated that more than 2.4 × 10⁵ propagules could be achieved per year through four subsequent in vitro rhizomes-generation cycles. Thus, a rapid method of propagation has been established.Abbreviations AdS Adenine sulphate - BA benzyladenine - CA casamino acids (vitamin free) - 2,4-D 2,4-dichlorophenoxyacetic acid - IAA indoleacetic acid - IBA indolebutyric acid - Kn Kinetin - NAA naphthaleneacetic acid - SH Schenk and Hildebrandt's medium (1972)     

Chromosome number and DNA content in callus culture ofCostus speciosus (Koen.) Sm.

Karyological changes in the callus tissue ofCostus speciosus (Koen.) Sm. at different ages have been analysed both by counting chromosome number and quantitating DNA content through cytophotometry. The cultures had been established from the tuber and maintained in modified Murashige and Skoog's basal medium (Physiol. Plant. 15: 473–497, 1962) supplemented with 2,4-D, NAA and BAP. Abnormality in chromosome behaviour leading to the formation of hypo- and hyperdiploid cells along with the diploid cells was observed. The abnormalities reached an optimum at different ages of the callus followed by a decline which varied amongst three populations. The frequency of hyperdiploid cells was higher than that of the hypodiploids. The role of endomitotic replication as well as non-disjunction of chromosomes resulting in the variation in chromosome number has been suggested. This was supported by the nuclear DNA value in successive passages. A difference in average amount of nuclear DNA with increase in age of the callus was recorded and the value also differed amongst these populations.     

Antioxidant activity of costunolide and eremanthin isolated from Costus speciosus (Koen ex. Retz) Sm.

Antioxidant properties of many medicinal plants have been widely recognized and some of them have been commercially exploited. Plant derived antioxidants play a very important role in alleviating problems related to oxidative stress. The present study was aimed at assessing the antioxidant property of costunolide and eremanthin isolated from a medicinal plant Costus speciosus (Koen ex. Retz) Sm. rhizome. Experimental diabetes was induced by a single dose of STZ (60 mg/kg, i.p.) injection. The oxidative stress was measured by tissue thiobarbituric acid reactive substances (TBARS), reduced glutathione (GSH) content and enzymatic activities of superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx) in brain, liver, heart, kidney and pancreas. An increase in TBARS level, a significant reduction in GSH content along with decreased enzymatic activities of SOD, CAT, and GPx were seen in untreated diabetic rats. Administration of either costunolide (20 mg/kg day) or eremanthin (20 mg/kg day) for 60 days caused a significant reduction in TBARS level and a significant increase in GSH content along with increased enzymatic activities of SOD, CAT and GPx in the treated rats when compared to untreated diabetic rats. Acute toxicity test revealed the non-toxic nature of the compounds. The results indicated for the first time the protective effect of costunolide and eremanthin from oxidative stress, thus opening the way for their use in medication.     

A simple and rapid method for selection of high diosgenin yielding clones ofDioscorea deltoidea callus

Summary Antimony pentachloride has been used to detect high diosgenin producing callus and cell clones ofDioscorea deltcidea grownin vitro. Using this method high yielding cultures, with a potential to produce up to 1.86% diosgenin, were selected.     

Antidiabetic and antilipidemic effect of eremanthin from Costus speciosus (Koen.)Sm., in STZ-induced diabetic rats

The increasing prevalence of diabetes mellitus worldwide is an issue of major socio-economic concern. Diabetes mellitus is a complex and a multifarious group of disorders that disturbs the metabolism of carbohydrates, fat and protein. Medicinal plants play an important role in the management of diabetes mellitus especially in developing countries. Costus speciosus is widely used in Indian medicine to treat various diseases. Eremanthin was isolated from C. speciosus. The structure was identified using gas chromatography–mass spectrometry (GC–MS) analysis. Eremanthin was administered to streptozotocin (STZ) (50 mg/kg bw) induced diabetic male Wistar rats at different doses (5, 10, 20 mg/kg bw) for 60 days. Plasma glucose level was significantly (p < 0.05) reduced in a dose dependent manner when compared to the control. In addition, oral administration of eremanthin (20 mg/kg bw) significantly decreased glycosylated hemoglobin (HbA_1c), serum total cholesterol, triglyceride, LDL-cholesterol and at the same time markedly increased plasma insulin, tissue glycogen, HDL-cholesterol and serum protein. Eremanthin also restored the altered plasma enzyme (aspartate aminotransferase, alanine aminotransferase, lactate dehydrogenase, alkaline phosphatase and acid phosphatase) levels to near normal. Results of this experimental study indicated that eremanthin possessed hypoglycemic and hypolipidemic activities and hence it could be used as a drug for treating diabetes.     

Effect of triacontanol on the micropropagation of <Emphasis Type="Italic">Costus speciosus</Emphasis> (Koen.) SM. Using rhizome thin sections

Summary The highest percentage of shoot regeneration of Costus speciosus was achieved using thin rhizome sections and triacontanol (TRIA). Factors affecting the rate of shoot multiplication and rooting with TRIA have been investigated. Initiation of shoot buds was observed when rhizome thin sections were cultured on B₅ basal medium supplemented with 5μgl⁻¹ TRIA. Shoots with two to three leaves produced roots when cultured on B₅ basad medium supplemented with 2 μgl⁻¹ TRIA. The well-rooted shoots were hardened and transferred to soil where they showed normal growth and a 100% survival rate was achieved. Results of this study showed that TRIA can be used as an effective growth regulator in the micropropagation and conservation of C. speciosus.     

Homaliodendron decompositum (Brid.) Wagn. reduced to synonymy with H. flabellaturn (Sm.) Fleisch

Abstract

Four small moss families found in Malawi are reviewed, and keys are provided to all taxa known in Malawi (and usually to all known in Africa). Regmatodontaceae is shown to contain only one species in Africa, but both known species are keyed. Rhachitheciaceae, contains three genera in Africa (which are keyed), only one of which occurs in Malawi, and for this genus a key is provided for all three African species. Rhacocarpaceae contains only two species in Africa, which are keyed, only one of which is known from Malawi. Rhizogoniaceae contains two genera and three species in Africa.     

Growth of callus and suspension culture cells from cotton varieties (Gossypium hirsutum L.) resistant and susceptible toxanthomonas malvacearum (E. F. Sm.) dows

Summary In vitro conditions are defined for starting and maintaining callus and suspension, cells from two cotton (Gossypium hirsitum L.) varieties, Im 216 and Acala 44, which are resistant and susceptible, respectively, to the bacterial pathogenXanthomonas malvacearum (E. F. Sm.) Dows. A light, friable callus was easily obtained and has been maintained for over 4 years. Whether stems or leaves, the explant source for callus initiation made no difference for growth of callus tissue. Acala 44 callus had a fresh-weight doubling time of 4 to 5 days, and Im 216 callus had a fresh-weight doubling time of 4 to 9 days; however, in suspension culture the fresh-weight doubling times for Im 216 and Acala 44 were 6 days. The pH of the suspension medium dropped to 4.7 during the exponential growth phase and rose to 5.4 at the stationary phase. Attempts to induce root and shoot initiation from these callus cells were unsuccessful; however, greening of the callus tissue did occur. Schenk and Hildebrandt medium was used for both callus and suspension cultures. Inoculation of Im 216 and Acala 44 callus tissues with two races ofX. malvacearum resulted in a resistant and susceptible response, respectively. This research was supported in part by C. S. R. S. Grant 315-16-96 and the Agricultural Experiment Station of Oklahoma State University.     

Growth responses of selected clones of birch (Betula pendula roth.,B. pubescens Ehrh.) and willow (Salix caprea L.,S. cinerea L.) to nitrogen in solution culture

Summary Growth of selected clones of birch and willow obtained from nitrogen deficient soils was compared with that of unselected controls in pot experiments using three levels of nitrogen. Unselected controls of both genera continued shoot growth, albeit very slowly, with a very low level of nitrogen (5 ppm), while selected clones of birch grew significantly more than the controls. Selected clones of willow, on the other hand, ceased shoot growth after 10 weeks with this low nitrogen treatment. Nevertheless, they remained healthy, their leaves containing similar concentrations of nitrogen to those of the plants which continued stem growth throughout the experiment. Furthermore, they had very high root: shoot ratios compared with those of control willows and both selected and unselected birch. The two genera may have developed different mechanisms for tolerating low nitrogen, birch producing a small, relatively efficient root system; willow a larger but less efficient one. Both appear equally effective in ensuring survival on low-nitrogen sites in the field since all the selected clones were obtained from such sites and have survived well in field trials on similar sites. Both birch and willow responded toincreased nitrogen availability with increasing shoot growth and a relative decline in root growth. However, whereas in willow the unselected plants responded significantly more than selected clones, a similar but less markeddifference was found in birch. It appears that in both genera, as in herbaceous plants originating from nitrogen deficient sites, selected clones are less able to respond to increasing nitrogen supply than control plants from more fertile habitats. Attempts to correlate the response of the selected clones to nitrogen in this experiment with that to added nitrogen fertilizer in field trials has been unsuccessful. Further work is required to determine the importance of the many interacting factors which influence the response of young trees to nitrogen under the unusual field conditions associated with restored mineral workings.     

Embryo culture of Medicago scutellata and M. sativa

Resistance to the alfalfa weevil (Hypera postica (Gyllenhal)) and the potato leafhopper (Empoasco fabae (Harris)) is lacking in cultivated alfalfa. However, a closely related annual Medicago, Medicago scutellata, possesses dense glandular stem and leaf hairs which provides a mechanism for resistance. Several attempts have been made at transfering the glandular haired traint from M. scutellata to perennial alfalfa with limited success. Earlier studies have shown that one reason for the lack of success is embryo abortion. Therefore, this study was initiated to observe zygotic embryo-genesis and to develop an embryo rescue technique for M. scutellata and M. sativa. Observations of zygotic embryogenesis showed that the two species are similar in morphology and can be described from youngest to oldest as globular, heart, torpedo, and hook shaped embryos. M. sativa embryos are smaller than M. scutellata embryos and develop three to four days later. Self pollinated M. scutellata (PI 307446) and sib mated M. sativa (Saranac AR) embryos were cultivated on Murashige and (2,4-D), indolacetic acid (IAA), 6-benzylaminopurine (BAP), and kinetic (KIN). Embryos from both species were also cultured on Schenk and Hildebrandt's (SH) basal medium with the addition of L-glutamine and L-proline. The experimental design was a completely randomized factorial for each experiment. Heart and torpedo shaped embryos from M. scutellata grew best (27.5% plantlet recovery) when cultured on MS medium with 0.05 mgl^-1 of both IAA and BAP. After 15 to 30 days on this medium, the embryos had only developed shoots. Therefore, it was necessary to transfer the shoots to MS basal medium without phytohormones for rooting. Rooting occurred in 15 to 30 days and the plantlets could be acclimatized to soil within 2 to 4 weeks. M. sativa embryos grew best (31% plantlet recovery) on SH medium with 50 mM L-glutamine. M. sativa embryos developed both shoots and roots on this medium. This information may now be applied to the development of an embryo culture method for recovering insect resistant hybrids between M. scutellata and M. sativa.Disclaimer statement: Mention of a trademark or proprietary product does not constitute a guarantee or warranty of the product by the USDA, and does not imply its approval to the exclusion of other products that may also be suitable.     

Anther culture of kale (Brassica oleracea L. convar.acephala (DC.) Alef.)

The pollen development and androgenic ability of 18 kale (Brassica oleracea convar.acephala) genotypes was observed during an anther culture study. Anther culture was successful in 6 of the genotypes and the highest yield obtained was 17 embryos per 100 anthers plated. Two stages of anther development were identified as being responsive to anther culture. The first and most responsive was that corresponding to the late uninucleated stage and the second to the late binucleated stage. These stages correspond with the onset of mitotic events in the microspores. Pollen viability was studied and low viability was noted which declined to zero after 9 days of anther culture. The initial viability level however was not clearly related to androgenic ability. The significance of the production of haploid and dihaploid kale genotypes in the study and breeding of resistance to clubroot is discussed.     

Taxonomic revision of Lepisorus （J. Sm.） Ching sect. Lepisorus （Polypodiaceae） from China

The taxonomy of Lepisorus （J. Sm.） Ching sect. Lepisorus in China was revised based on herbarium specimen examinations, field observations, and microscopic study of rhizome scales, soral paraphyses, leaf epidermis, and spores. As a result, nine species were recognized： Lepisorus macrosphaerus （Baker） Ching, Lepisorus asterolepis （Baker） Ching, Lepisorus marginatus Ching, Lepisorus kuchenensis （Y.C. Wu） Ching, Lepisorus megasorus （C. Chr.） Ching, Lepisorus kawakamii （Hayata） Tagawa, Lepisorus subsessilis Ching ＆ Y.X. Lin, Lepisorus affinis Ching, and Lepisorus nudus （Hook.） Ching. Lepisorus kawakamii （Hayata） Tagawa was reinstated; Lepisorus gyirongensis Ching ＆ S.K. Wu and Lepisorus longus Ching were reduced to synonyms ofL. nudus and L. affinis, respectively. The subdivision ofLepisorus macrosphaerus was not accepted. Rhizome scales and paraphyses are the most useful characters for species delimitation as well as for infrageneric classification. Characteristics of the leaf epidermis and spore ornamentation are usually stable and thus of great significance in understanding the relationships among groups within the genus.     

Ovule and Embryo Ontogenesis in Bombacopsis glabra (Pasq.) A. Robyns

Ontogeny of the ovule and development of the embryo in Bombacopsisglabra (Pasq.) A. Robyns were examined. The ovule is bitegmic,crassinucellate, and anatropous. The exostome is eccentric relativeto the endostome; stomata occur on the outer integument. Thesingle archesporial cell functions directly as the megasporemother cell. The embryo-sac is bisporic. The organization ofthe nuclei in the mature embryo-sac is normal. The antipodalcells disintegrate soon after formation. Double fertilization takes place; the zygote undergoes a longperiod of dormancy, but the primary endosperm nucleus dividesimmediately to produce first a nuclear-type, later a cellular-typeendosperm. The zygote is of the caryophyllad type. Adventive embryos arise from single cells of the nucellus inthe vicinity of the micropyle, and develop faster than the sexuallyproduced embryo; this leads to anomictic renroduction.     

荚果蕨胚胎发育的研究

利用石蜡切片法研究了荚果蕨（Matteuccia struthiopteris(L.) Todaro）胚胎发育过程。合子第一次分裂,分裂面垂直于原叶体纵轴且平行于颈卵器颈部;第二次分裂面平行于原叶体纵轴且垂直于颈卵器颈部;第三次分裂面同时平行于原叶体纵轴和颈卵器颈部。经多次分裂的球形胚胎,胚胎的外上和外下区域几乎同时分别发育出第一叶顶端细胞和第一根顶端细胞。随着发育的进行,它们分别斜向分裂产生第一叶原基和第一根原基。随后,第一叶原基迅速分裂,突破帽状体形成第一幼叶;而第一根原基的分裂速度稍慢,第一根发育速度稍慢于第一叶。     

Performance ofin vitro propagatedAlnus glutinosa (L.) Gaertn. clones inoculated withFrankiae

Summary ThreeAlnus glutinosa (L.) Gaertn. clones, obtained byin vitro propagation techniques, were inoculated with four strains ofFrankia. The ability of these clones to nodulate and fix nitrogen was previously reported; this study deals with the performance of 12 different combinations of pairs of symbionts.Shoot fresh weight, shoot height and collar diameter were measured 60 and 82 days after inoculation. Shoot fresh weight seems to be more sensitive and reliable than the other parameters. Nitrogenase activity, measured by the acetylene reduction assay, was assayed 78 days after inoculation and was consistent with the biomass measurements.Better growth was observed when type N strains were used. Significant growth differences were observed between clones AG-2 and AG-8 on the one hand and clone AG-4 on the other. Thus, the use of genetically defined host plants and microsymbionts permitted the demonstration of significant performance variation even among cloned plants from the same provenance (AG-4 and AG-8).The duration of the experiment influenced the results with differences becoming less significant with time. This might be caused by an external limiting factor such as the pot size, competition for light,etc. But it could also be indicative of differences in nodulation speed among the treatments.