Asparagaceae Research Articles

Comparative Morphology and Taxonomic Study of Chlorophytum sabiense And Two Other Species in the Genus Chlorophytum Ker-Gawl

Chlorophytum sabiense Omok. (a newly identified species in the family Asparagaceae) is unique, having multiple styles emerging from fused ovaries. This specialized character may be overlooked in conjuring up archetype for the genus Chlorophytum, since the genus is characterized by single and simple style. Comparative morphology and taxonomic study of Chlorophytum sabiense and two other species in the genus (Chlorophytum macrophyllum and C. stenopetalum) was carried out to appraise level of relatedness of Chlorophytum. sabiense and these existing members. Distribution of the three species is sympatric in the North Central part of Nigeria. Chlorophytum sabiense may be confused with either of the two species because of the attributes they have in common. Samples collected were raised in a garden and closely observed for duration of three years. Data collected from morphological studies were analyzed statistically. The three species show similarity in most of the vegetative characters studied, however Chlorophytum sabiense is distinguished from the two others having; multiple styles, smaller fruit size and shorter length of pistil relative to the stamen. Exceptional characteristics exhibited by Chlorophytum sabiense makes it stand out among species in the genus, while its close resemblance with its allies accounts for reason why it had escaped recognition for a long time.

Study the Anatomical Characters for Species of Tow Genera Bellevalia Layper, and Ornithogalum L. of Asparagaceae Family of Middle and Northern Iraq

The study dealt with anatomical characters of 11 plant species that belong two genera Bellevalia Layper and Ornithogalum L. from Asparagaceae family growing in middle and north of Iraq B.kurdistanica, B.longipes, B.macrobotrys, B.parva, B.pycnantha, B.saviczii, O.brachystachys, O.neurostegium, O.pyrenaicum. The prevalence of Aleuron grains and Starch grains in addition to studying the aerial stem epidermal cells or the Scapeous and the leaves. When the cross section was made to the aerial stem, it was clear that all of them were circular except B.chrisii it was ovate for the aerial stem. It was noticed also that there are crystals within the cross section of the aerial stem of these species, but the species B.saviczii had different type of crystals which are Druses crystals at the epidermis layer of the aerial stem section. Also, there was a difference in the vascular bands spread, which were in the shape of two rows arranged similar to the arrangement of the vascular bands in Dicotyledons. Anatomy results of the leaves cross-section showed that they consist of simple epidermis with one row of Uniseriate cells, consisting of upper, lower epidermis and Mesophylla as well as the spread vascular bands.

Bulbous Plants Drimia: "A Thin Line between Poisonous and Healing Compounds" with Biological Activities.

Drimia (synonym Urginea) plants are bulbous plants belonging to the family Asparagaceae (formerly the family Hyacinthaceae) and are distinctive, powerful medicinal plants. Just some species are indigenous to South Africa and have been traditionally utilized for centuries to cure various diseases and/or ailments. They have been recognized among the most famous and used medicinal plants in South Africa. Traditionally, the plants are used for various illnesses such as dropsy, respiratory disease, bone and joint complications, skin disorders, epilepsy and cancer. A number of studies have reported biological properties such as antiviral, antibacterial, antioxidant and anti-inflammatory, immunomodulatory, and anticancer activities. Their bulbs are a popular treatment for colds, measles, pneumonia, coughs, fever and headaches. However, some plant species are regarded as one of the six most common poisonous plants in Southern Africa that are toxic to livestock and humans. Due to the therapeutic effects of the Drimia plant bulb, research has focused on the phytochemicals of Drimia species. The principal constituents isolated from this genus are cardiac glycosides. In addition, phenolic compounds, phytosterols and other phytochemical constituents were identified. This study constitutes a critical review of Drimia species’ bioactive compounds, toxicology, biological properties and phytochemistry, advocating it as an important source for effective therapeutic medicine. For this purpose, various scientific electronic databases such as ScienceDirect, Scopus, Google Scholar, PubMed and Web of Science were researched and reviewed to conduct this study. Despite well-studied biological investigations, there is limited research on the toxic properties and the toxic compounds of certain Drimia species. Searching from 2017 to 2021, Google Scholar search tools retrieved 462 publications; however, only 3 investigated the toxicity and safety aspects of Drimia. The aim was to identify the current scientific research gap on Drimia species, hence highlighting a thin line between poisonous and healing compounds, dotted across numerous publications, in this review paper.

Morphological, phytochemical and genetic diversity of threatened Polygonatum verticillatum (L.) All. populations of different altitudes and habitat types in Himalayan region.

The online version contains supplementary material available at 10.1007/s12298-021-01044-9.

Plant structure specialization in Paraphidippus basalis (Araneae: Salticidae), a jumping spider of the Madrean Sky Islands

Paraphidippus basalis (Banks, 1904) is a large jumping spider that occurs in the sky islands of the southwestern United States and northern Mexico. To date, P. basalis has only been incidentally reported on rosette-forming plants in the family Asparagaceae (yucca, agave, and sotol), even though the sky islands support a rich and diverse vegetation community. This apparent specialization is unusual because jumping spiders do not typically have strong associations with the plants on which they live. However, given that the ecology of P. basalis has yet to be studied, the microhabitat preferences of P. basalis remain unclear. We investigated microhabitat choice in P. basalis in the Patagonia Mountains of southeastern Arizona, to determine whether these spiders were specifically associated with rosette-forming plants. We surveyed 160 plots for jumping spiders, 80 with rosette-forming plants and 80 without. P. basalis was found only in rosette-forming plants, whereas other species of jumping spiders showed no preference for rosette or control plots. Larger rosette plants were more likely to contain P. basalis. This study provides an unusual example of host plant structural specificity in a jumping spider.

Saponins from the rhizomes of Polygonatum nodosum Hua and their chemotaxonomic significance

Phytochemical investigation on the rhizomes of Polygonatum nodosum Hua resulted in the isolation of thirteen saponins, which could be subdivided into eleven spirostanol saponins (1–5, 7–11 and 13) and two triterpenoid saponins (6 and 12). Structure elucidation of these isolates was performed based on MS, CD, 1D and 2D NMR data. All compounds (1–13) were first reported from P. nodosumthis, and compound diosgenin 3-O-α-L-rhamnopyranosyl-(1→2)-[O-α-L-arabinofuranosyl-(1 → 4)]-β-D-glucopyranoside (7) was first reported from the family Asparagaceae. Moreover, the chemotaxonomic significance of isolates was detailedly introduced.

Intracellular DNA transfer events restricted to the genus Convallaria within the Asparagaceae family: Possible mechanisms and potential as genetic markers for biographical studies

Intracellular gene transfer among plant genomes is a common phenomenon. Due to their high conservation and high plastid membrane integrity, chloroplast (cp) genomes incorporate foreign genetic material very rarely. Convallaria is a small monocotyledonous genus consisting of C. keiskei, C. majalis and C. montana. Here, we characterized, analyzed and identified 3.3 and 3.7 kb of mitochondrial DNA sequences in the plastome (MCP) of C. majalis and C. montana, respectively. We identified 6 bp and 23 bp direct repeats and mitochondrial pseudogenes, with rps3, rps19 and rpl10 identified in the MCP region. Additionally, we developed novel plastid molecular genetic markers to differentiate Convallaria spp. based on 21 populations. BEAST and biogeographical analyses suggested that Convallaria separated into Eurasian and North American lineages during the middle Pliocene and originated in East Asia. Vicariance in the genus was followed by dispersal into Europe and southeastern North America. These analyses indicate that the MCP event was restricted to the genus Convallaria of Asparagaceae, in contrast to similar events that occurred in its common ancestors with other families of land plants. However, further mitochondrial and population studies are necessary to understand the integration of the MCP region and gene flow in the genus Convallaria.

Study of chlorophylls and carotenoids in Sansevieria hyacinthoides leaves

Topicality. Sansevieria hyacinthoides (L.) Druce belongs to the Asparagaceae family. The plant is native to African countries. It has been introduced and naturalized in some countries of America, Asia, Australia and Europe. This species is grown as an ornamental plant in many countries, and as a medicinal plant in Africa. Aim. To determine the quantitative content of chlorophylls a and b and the amount of carotenoids in Sansevieria hyacinthoides leaves freshly collected and dried. Materials and methods. The quantitative content was determined by the spectrophotometric method. This method allows determining the amount of chlorophyll a and b and carotenoids by the characteristics of the absorption spectra without their prior separation. Results and discussion. The amount of chlorophyll a and b in the dried leaves was almost 7 times higher than in the freshly collected ones (1.82 ± 0.036 mg/g vs. 0.25 ± 0.005 mg/g, and 2.08 ± 0.038 mg/g vs. 0.29 ± 0.006 mg/g, respectively). The quantitative content of the amount of carotenoids differed by 3.6 times (0.05 ± 0.001 mg/g in freshly collected leaves, and 0.18 ± 0.004 mg/g in the dried raw material). Conclusions. The quantitative content of the amount of carotenoids, chlorophyll a and chlorophyll b in freshly collected and dried leaves of Sansevieria hyacinthoides has been determined by the spectrophotometric method. The dried plant raw material of Sansevieria leaves has a higher content of biologically active substances: chlorophyll a – 1.82 ± 0.036 mg/g, chlorophyll b – 2.08 ± 0.038 mg/g, and the amount of carotenoids – 0.18 ± 0.004 mg/g.

Ontogenetic and vitality structure of Muscari botryoides (L.) Mill. (Asparagaceae Juss.) populations at the northeastern limit of its 7nature area

Досліджено онтогенетичну та віталітетну структуру восьми популяцій Muscari botryoides з території України. M. botryoides – це рідкісний, ендемічний вид з родини Аsparagaceae, який перебуває на північно-східній межі ареалу та занесений до третього видання «Червоної книги України» зі статусом «зникаючий». Подано характеристику основних онтогенетичних станів виду. Побудовано вікові спектри популяцій. Розраховано онтогнетичні індекси за І.М. Коваленко. У складі досліджуваних популяцій M. botryoides виявлено особини восьми вікових станів: насіння (sm), проростки (p), ювенільні (j), іматурні (im), віргінільні (v), молоді генеративні (g1), зрілі генеративні (g2) та старі генеративні (g3). Субсенільні (ss) та сенільні (s) особини не зареєстровані. Вікові спектри всіх досліджуваних популяцій лівосторонні з переважанням рослин ювенільного онтогенетичного стану. За ознаками онтогенетичної структури усі досліджувані популяції – нормальні, неповночленні. Згідно з класифікацією популяцій О.О. Уранова та О.В. Смірнової вони є нормальними молодими, згідно з класифікацією «Дельта-Омега», запропонованою Л.А. Животовським – молодими, згідно з Л.О. Жуковою та Т.А. Полянською – перспективними. Індекси відновлення популяцій дуже високі, а індекси генеративності, старіння та віковості – низькі. Проведено факторний аналіз з урахуванням 17 морфопараметрів M. botryoides, встановлено, що ознаками, які детермінують життєвість особин є: довжина першого листка, ширина цибулини, довжина суцвіття та кількість квіток у суцвітті. Віталітетний аналіз засвідчив, що у більшості досліджуваних популяцій домінують особин середнього рівня життєвості. За віталітетною структурою шість з восьми популяцій відповідають процвітаючому якісному типу, дві популяції перебувають у депресивному стані.

In vitro Propagation and Caulogenesis of Dracaena draco Plants

Dracaena draco is a monocot from family Asparagaceae. The dragon tree is a subtropical plant and features many botanic and Mediterranean gardens worldwide. D. draco has been categorized as vulnerable in Europe and endangered worldwide in the IUCN Red List of Threatened Species (IUCN, 1998). The aim of this study is to develop an efficient protocol for rapid in vitro propagation and caulogensis of D. draco Plants via enhanced axillary bud proliferation from single nodal explants cultured on full strength MS medium, 30g/l sucrose, 4g/l gelrite augmented with various concentrations of plant growth regulators as BA, NAA, IBA and their combinations. In general, the present study concluded that the best results for the initiation stage were recorded on MS medium fortified with NAA and BA at 2.00 and 1.00 mg/l, respectively. Meanwhile, supplemented medium with BA and NAA at 4.00 and 1.00 mg/l, respectively, gave rise to the best results for the multiplication stage. Regarding rhizogenesis stage, the best results were recorded when the neoformed shoots of the multiplication stage were divided singly and cultured on MS medium plus IBA and NAA at 1.00 and 0.50 mg/l, respectively which led to the highest mean number of roots formed per propagule. For callus induction, the internodal segments cultured into MS medium supplemented with BA and NAA at 0.50 and 2.00 mg/l, respectively, gave rise to the best results for the percentage of explants formed callus and callus size. Concerning the percentage of direct organogenesis and number of shoots formed per explants best results obtained from BA and NAA at 0.250 mg/l and 0.00 mg/l, respectively.Neoformed plantlets were acclimatized ex vitro and in vivo vigorously in a mixture of perlite and peat moss at (1:1) or (2: 2) or (2:3) and (3:3) consecutively, in addition to fixed volume (1 portion) of sterile sand; which resulted in the highest mean value of survival percentage/plant (100%) and showed true-to-type plants ex vitro.

Extraction and Processing of Pharmaceutical Grade Microcrystalline Cellulose from Dracaena arborea Stem

Cellulose is an abundant renewable biodegradable polymer regarded as a promising feedstock for chemical productions with its versatility evaluated as a useful structural and functional material for pharmaceutical and industrial applications. It is a straight chain polymer which appears in cell walls of most plant and consists of D. glucose units, with absence of coiling or branching and can be derived from variety of sources including: annual plants, microbes, animals etc. Three basic types of cellulose often exist in nature as - alpha (α), beta (β) and gamma (ϒ). Microcrystalline cellulose (MCC) sourced from cellulose, occurs as a purified and partially depolymerized alpha cellulose from plant parts such as D. arborea stem possibly by severe acid or alkaline hydrolysis. D. arborea plant is a palm – like tree of 1.5m height with several branches, often used as a boundary mark, a non-selective habitat and belongs to the family Asparagaceae, sub family Nolinoideae. With alkali hydrolysis of the D. arborea stem, the percentage yield of MCC from the alpha cellulose is 54.32%. Physicochemical analysis of the MCC reveals it to have a pH of 7.80 and physicotechnical analysis resulted in values as recommended in the official monogragh. Proximate principles of the extracted MCC, depicted percentage fiber content as 65.78% and low lipid and protein content as 0.6 and 0.4% respectively. Elemental analysis shows the composition of sodium and iron as 41% and 35% respectively but absence of lead and other deleterious materials. FTIR analysis suggests the presence of carbonyl groups, 6- membered cyclic ring (aromatic structure) with ortho and meta - OH substitution and long aliphatic chains. The x-ray diffraction study gave a percentage crystalinity index of 6.02 at 2q = 22 and 2q = 34.

The complete plastid genome sequence of Dracaena fragrans (L.) Ker Gawl. (Asparagaceae)

Dracaena fragrans is a shrub native to tropical Africa classified in the family Asparagaceae. It is a common horticultural plant grown as a hedge as well as an ornamental house plant for its green leaves, attractive shape and fragrant flowers. The species has been used for many medicinal purposes including inducing labor, treating malnutrition and fighting illness. Here, we investigated the complete plastid genome of D. fragrans using Illumina 150 bp next-generation sequencing technologies. De novo assembly and annotation showed that the chloroplast genome is a typical quadripartite structure with 37.5% GC content. A pair of inverted repeats (IRs, 26,507 bp each) were separated by one large single-copy region (LSC, 83,703 bp) and one small single-copy region (SSC, 18,466 bp). The chloroplast genome is 155,183 bp in length and contained 131 genes of which 12 are intron-containing genes, comprising 85 protein-coding, 38 tRNA, and 8 rRNA genes. Phylogenetic analysis of D. fragrans and 11 plastomes obtained from GenBank shows that it is closely related to D. elliptica.

Tolerancia de plantas de zonas áridas a metales pesados

En México las zonas áridas representan más de la mitad del territorio, en estos lugares existe erosión debido a la contaminación, por lo que es importante estudiar el nivel de tolerancia de las especies vegetales representativas para restaurar o reforestar. Se evaluaron seis especies de zonas áridas Agave funkiana, A. obscura, A. salmiana de la familia Asparagaceae y Opuntia cochenillifera, O. f icus-indica, Pereskia sacharosa, de la familia Cactaceae. Se propagaron in vitro para la obtención de brotes que fueron establecidos a diferentes concentraciones de cadmio (Cd2+) y plomo (Pb2+) durante la etapa de enraizamiento para determinar su tolerancia. Se utilizó el método Probit para calcular la concentración letal media (CL50) de las seis especies a las diferentes dosis. Las diferentes concentraciones de Cd y Pb, produjeron distintos efectos en las seis especies de plantas de zonas áridas evaluadas en esta investigación. A. salmiana y P. sacharosa obtuvieron una mortalidad de 55% con 0.009 mM de Cd y una mortalidad de 77% con 0.8 mM para Pb, siendo estas concentraciones las más altas. Los bioensayos ecotoxicológicos son útiles para evaluar contaminantes como los metales pesados en plantas tolerantes como A. obscura, O. cochenillifera y O. f icus-indica.

Controlled-release Fertilizer in Cordyline spectabilis Seedlings Production, a Potential Species of the Asparagaceae Family

Abstract The success of seedlings establishment for commercial planting and forest restoration depends on the availability of nutrients during seedlings production. The objective was to evaluate the efficiency in controlled-release fertilizer use in Cordyline spectabilis seedlings production and what is the additional cost per seedling produced. Seedling production was performed with doses of 0.0 (control); 2.5; 5.0; 7.5; 10.0 and 12.5 kg m-³ of Basacote® Plus 9M (16-08-12) controlled-release fertilizer added to the substrate. The experiment was placed in a greenhouse and we evaluated morphological parameters to determine seedlings growth. The dose of 10.7 kg m-3 provided a greater number of leaves (11.4) and leaf length (24.9 cm), higher values of shoot (4.64 g), root (2.32 g) and shoot root ratio (1.99) were obtained above 10.7 kg m-3 fertilizer. We recommend the use of 10.7 kg m-3 controlled-release fertilizer for seedlings production with an increase of R$ 0.05 per seedling produced.

Phylogeny, Diversification Rate, and Divergence Time of Agave sensu lato (Asparagaceae), a Group of Recent Origin in the Process of Diversification.

Agave sensu lato is one of the most diverse and complex genera of Asparagaceae, with more than 250 species. The morphological, ecological, and evolutionary diversity of the group has complicated its taxonomical study. We conducted phylogenetic analyses of DNA sequence data to reconstruct the phylogenetic relationships of the Agave genus. We included 107 species of the Asparagaceae family from which 83 correspond to the Agave sensu lato clade (Agave sensu stricto + Polianthes + Manfreda and Prochnyanthes, which together represent 30% of the genus) and as outgroups the genera Dasylirion, Hesperoyucca, Chlorogalum, Camassia, Hesperaloe, Yucca, Beschorneria, and Furcraea, in order to estimate the age and propose the history of their diversification. Previous studies postulated the relevance of the Miocene in the speciation rates of the agaves, as well as the relevance of the type of inflorescence in its diversification. However, these assertions have not been well supported. The analysis of chloroplast regions resulted in low resolution, which could be the consequence of the few variable sites. On the other hand, the internal transcribed spacer (ITS) implemented in our analysis ensued in higher resolution and better support values. Our phylogenetic analyses recovered five groups; one is the Striatae group, which is the sister group to Agave sensu stricto clade. Within this clade, we found three main groups with high support; these groups are not related with previous morphological proposals. We also analyzed the dates of origin and diversification rates. A Bayesian analysis of macroevolutionary mixtures indicated two significant shifts; the first was identified at 6.18 Ma, where the speciation rate increased to 4.10 species/Mya, this shift occurred during the late Miocene period, characterized by the emergence of arid biomes in North America. The second was identified at a stem age of 2.68 Ma where the speciation rate increased to 6.04 species/Mya. Concerning the ancestral reconstruction state of the inflorescence type in the Agave sensu stricto clade, the spike inflorescence character was predominant in the early-diverging groups, whereas the late-diverging groups present panicle inflorescences as the predominant character and higher speciation rates.

Kellermania macrospora Causes Leaf Dieback of Yucca filifera in Queretaro, Mexico

Yucca is a genus of perennial shrubs and trees (family Asparagaceae subfamily Agavoideae) native to hot, dry parts of America and the Caribbean. Yucca filifera is an important plant with a wide range of uses. In November 2017, in Pena de Bernal, Queretaro (20°44′21.45″ N; 99°56′20.13″ W), leaf dieback was observed in natural areas, affecting up to 15% of plants. The disease started with large brown spots on the apex of leaves (in some cases on one side) that dried out. Lesions were covered with black conidiomata scattered throughout. The disease gradually advanced to the leaf base. The disease does not cause death but affects the valuable leaves and indirectly affects plant development. Whole leaf blades at five different survey sites were collected, with three leaf blades per site. Fractions were cut from symptomatic tissue, including healthy tissue, surface disinfested with 2% NaOCl for 2 min, rinsed in sterile distilled water (SDW), and placed in Petri dishes with three filter paper discs soaked with SDW. They were incubated under fluorescent light at 25°C with 12-h light/dark. After 4 days, conidiomata were examined under both an optical microscope and field emission SEM. Conidia were collected from conidial masses exuding from freshly exposed pycnidia. Masses of conidia were collected using a sterilized needle, placed into 1.5-ml tubes with SDW, spread on PDA Petri dishes, and incubated at 25°C. The presumptive causal agent was isolated; 5 × 5-mm sections were cut from the margin of infected tissue and disinfected, placed on PDA Petri dishes, and incubated. Cultures of similar morphology were consistently recovered from both masses of conidia and infected tissue on PDA. Twenty monoconidial isolates were obtained. A representative one was selected for morphological characterization, multilocus phylogenetic analysis, and pathogenicity tests. Growth conditions were 25°C with 12-h light/dark. Cultures were measured and photographed after 2 weeks; appearance was noted after 2 to 3 weeks. On the host, conidiomata pycnidial, black, immersed, globose, unilocular, 365 to 537 × 270 to 334 μm; ostiole central, nonpapillate, up to 37.8 μm diameter, exuding a hyaline conidial cirrhus. Conidiophores lining the inner cavity, reduced to conidiogenous cells, hyaline, smooth, subcylindrical. Conidia hyaline, guttulate, cylindrical to obclavate, three-septate, conidial size (100 conidia) of 82.2 to 94.5 × 7.7 to 9.5 μm; apex acutely rounded; conidial base truncate, appendages absent. Surface ornamentation of conidia, striate with thin parallel ridges. Culture characteristics: colonies on PDA after 2 weeks; surface near light gray, moderate aerial mycelium; margin irregular, whitish; conidiomata black; reverse uncolored. The rDNA ITS was sequenced (MN509001). The sequence was compared with GenBank and showed 98.17 and 98.05% identity with Kellermania macrospora KF766178.1 and KF777165.1, respectively. The fungus was identified as K. macrospora (Crous et al. 2013; Slippers et al. 2013). To confirm Koch’s postulates, Y. filifera plants (∼3 years) were cut and rooted in pots with sterile substrate and kept confined for 6 months before inoculation. A suspension was prepared from 1-week-old culture (MN509001) in SDW at 8 × 10³ conidia/ml containing 0.02% Tween 20. The suspension was sprayed on healthy leaves of 10 plants; SDW with 0.02% Tween 20 was used as a control (three plants). Plants were covered for 48 h with plastic bags to maintain 100% RH. Bags were removed, and plants were maintained at 25 ± 5°C, 65% RH. Lesions developed within 25 days of inoculation; control plants remained symptomless. The fungus recovered from inoculated tissues showed the same morphological characteristics as the isolate used in inoculations. K. macrospora has been reported on Agave sp. (Minnis et al. 2012) and Yucca rostrata (Crous et al. 2013). The leaf dieback on Y. filifera caused by K. macrospora is the first report from Mexico. Yucca leaf dieback has not been reported elsewhere. It appears to be newly emerging in Mexico. It is recommended to establish a pruning scheme to eliminate affected leaves to reduce inoculum levels.

Productive method for commercial propagation through direct embryogenesis and organogenesis in Ledebouria revoluta—an underutilized medicinal plant with cardioprotective properties

The bulbaceous herb Ledebouria revoluta (family Asparagaceae) is an important cardioprotective plant. Two different pathways of in vitro direct regeneration system consisting of somatic embryogenesis and shoot organogenesis have been established for high-frequency micropropagation. A rectangular piece (6 × 10 mm2) of leaf lamina was used as an explant to study the morphogenetic regulation by exogenously supplied auxins, cytokinins, and polyamine within controlled in vitro environment. The explants were cultured on Murashige and Skoog (MS) medium supplemented with 3.0 % sucrose and various concentrations and combinations of plant growth regulators (PGRs), as per experimental needs. Both β-naphthoxyacetic acid and α-naphthalene acetic acid (NAA) auxins individually induce somatic embryogenesis. Combined application of auxins and cytokinins further improves the embryonic response. The synergistic effect of NAA (2.0 mg L–1), kinetin (1.0 mg L–1), and spermidine (SDN, 217.88 mg L–1) provided the best result with 96.7 ± 1.8 % response and highest 63.4 ± 0.8 embryos per explant. The highest (94.5 ± 2.5 %) conversion of embryos to complete plantlets was noted in presence of 1.0 mg L–1 kinetin. The morphogenic path altered from embryogenic to organogenic when auxin was totally withdrawn, and a higher concentration of cytokinin was applied. The synergism between 6-benzylaminopurine (3.0 mg L–1) and SDN (217.88 mg L–1) proved the best PGR combination for the direct shoot (or bulblet) organogenesis with maximum 96.6 ± 2.2 % response and highest 48.6 ± 0.5 bulblets per explant. Total withdrawal of auxin was essential for direct bulblet organogenesis because the presence of any auxin always shifts the morphogenic path from organogenic to embryogenic. Histological study confirms the bipolar structure of somatic embryo and unipolar bulblets. Highest 12.7 ± 0.54 roots per bulblet were induced on ½ strength MS medium containing 2.0 mg L–1 indole-3-butyric acid. Finally, the plantlets were acclimatized with 96.7 % survival rate and all of the survived plants produced bulbs and 92.5 % of them flowered normally in field conditions. This in vitro system permits a high-frequency regeneration through embryogenic and organogenic paths, which can be used for large-scale commercial micropropagation for round-the-year supply of this important cardioprotective medicinal plant.

Development of the female gametophyte of Agave colimana, subgenus Littaea (Asparagaceae)1

Agave colimana, subgenus Littaea (Asparagaceae) is a wild plant that belongs to the Asparagaceae family, and in this species, the formation of its female reproductive system is unknown, probably because it has no commercial use. However, A. colimana recently has been considered as a candidate for marketing as a succulent plant because of its floral and growth architecture. Knowledge about aspects of the formation of the embryo sac is useful for understanding its sexual reproduction, which is important for propagation by seed and breeding purposes. Megasporogenesis and megagametogenesis were studied using Feulgen staining under confocal microscopy. Megasporogenesis resulted in the formation of a linear tetrad with the functional megaspore attached to the chalazal end. Subsequently, megagametogenesis from the functional megaspore was followed by three mitotic divisions generating an embryo sac with seven cells: two synergid cells, one egg cell, three antipodal cells, and one central cell with a 2n nucleus. The embryo sac of Agave colimana was found to be monosporic of the Polygonum-type considered as the typical type in the majority of angiosperm plants. Also, abnormal embryo sacs were observed, following possible errors during meiosis.

Genetic structure, molecular and phytochemical analysis in Iranian populations of Ruscus hyrcanus (Asparagaceae)

Ruscus hyrcanus Woronow a commercially important ornamental and medicinal plant belonging to the Asparagaceae family, has various applications in Iranian traditional medicine due to its diuretic, appetizing, cathartic, vasoconstrictor, anti-hemorrhage, ant-nephritis, anti-infection, aperient and anti-varicose properties. In the present study, morphological, phytochemical and molecular markers were investigated in twelve R. hyrcanus (RH) populations collected from the Hyrcanian ecoregion in the north of Iran. Phytochemical characteristics were remarkably variable among the studied populations, but little morphological variation was observed. The highest contents of ruscogenin and neoruscogenin in the underground parts of the plant were recorded in populations RH9 (Abbasabad 3.41 mg/g DW) and RH6 (Darab Kola 1.44 mg/g DW), respectively. A total of 66 bands were obtained from 11 inter-simple sequence repeat (ISSR) markers, 62 (93.94 %) of which were polymorphic. Although the ISSR evaluation indicated an inter-population genetic differentiation (GST = 0.27), most of the genetic diversity was intra-populational (PPB = 32.26–75.81 %, He = 0.11−0.24, I = 0.17−0.36). A UPGMA cluster analysis of 120 samples of R. hyrcanus was carried out to categorize the studied populations, based on a molecular data matrix calculated with Jaccard’s similarity coefficient. A multiple regression analysis of the relationships between the ISSR bands and morphological and phytochemical traits demonstrated that eight ISSR fragments were related to ruscogenin in the plant materials. Several ISSR markers showed association with some morphological traits. This study provided the comprehensive preliminary data on morphological, phytochemical, and genetic characteristics of R. hyrcanus populations in order to exploit genetic reserves conservation programs.

REGENERACIÓN DE PLANTAS DE Agave marmorata Roezl, VÍA EMBRIOGÉNESIS SOMÁTICA

<p><strong>Background. </strong><em>Agave marmorata</em> Roezl is a monocot plant of the Asparagaceae family, which is mainly used in the elaboration of the alcoholic drink called mezcal. This species is propagated by seeds or stolons (hijuelos), but its reproduction is slow because its maturation stage can last up to 35 years until flowreing. Therefore, efficient and alternative propagation methods should be used to ensure the permanence of the resource. Biotechnology, in particular plant tissue culture techniques, represents a viable alternative for the propagation of this species, however, until now, there are no previous reports on the spread of <em>A. marmorata</em> Roezl, via somatic embryogenesis. <strong>Objective. </strong>Evaluate various treatments to regenerate seedlings of this species through somatic embryogenesis. <strong>Methodology.</strong> Callus induction was obtained from mature seeds established in MS (Murashige and Skoog, 1962) at 25%, supplemented with vitamins L2 (Phillips and Collins, 1979), 5 mg L<sup>-1</sup> of 2,4-dichlorophenoxyacetic (2 , 4-D), 3 mg L<sup>-1</sup> of 6-benzyladenine (BA), 60 g L<sup>-1</sup> of sucrose, 8 g L<sup>-1</sup> of agar and adjusted to a pH of 5.7 ± 0.1. These calluses were sectioned into small portions weighing approximately 0.25 g and were incubated in an MS medium at 50% of their concentration with 30 g L<sup>-1</sup> of sucrose, 8 g L<sup>-1</sup> of agar and subjected to eight concentrations of two plant growth regulators (RCV): BA (0.0, 2.0, 6.0 and 10 mg L<sup>-1</sup>) and 2,4-D (0.0 and 5 mg L<sup>-1</sup>), alone or combined, giving a total of eight treatments. Those explants that responded to the embryogenic callus formation were subcultured to 50% MS medium, 30 g L<sup>-1</sup> of sucrose and gelled with 8 g L<sup>-1</sup> agar where two RCV were evaluated independently (0.1 mg L<sup>-1</sup> of 2,4-D or 3 mg L<sup>-1</sup> of AG<sub>3</sub>) and two culture conditions: 16 h light and 8 h dark and complete darkness, giving a total of 24 treatments. The variables evaluated in this work were: percentage of callus induction, percentage of embryogenic structures, number and growth of regenerated seedlings of <em>A. marmorata</em> Roezl at 255 ddic. <strong>Results.</strong> The best response was observed 120 days after starting the culture (ddic) in treatment five with 0.1 mg L<sup>-1</sup> of 2,4-D under light conditions where the pre-treatment was with 10 mg L<sup>-1</sup> of BA, obtaining 19.4 somatic embryos per explant. Maturation of somatic embryos in 50% MS medium was achieved with 30 g L<sup>-1</sup> of sucrose and 8 g L<sup>-1</sup> of agar, without RCV. The 100% of the regenerated seedlings survived and grew under greenhouse conditions. <strong>Implications.</strong> The results of this study contribute to a better understanding of the importance of a pretreatment with high concentrations of cytokinin (BA) and culture conditions, for the regeneration of <em>Agave marmorata</em> Roezl plants, via somatic embryogenesis. This can assist in the initiation of germplasm genetic improvement and <em>in vitro</em> conservation programs. <strong>Conclusions. </strong>The factors evaluated here were important in the induction and expression of embryogenic structures of this species, making possible the regeneration of seedlings of <em>A. marmorata </em>Roezl, from the formation of somatic embryos.</p>