Epiphytic Orchid Research Articles

Processes controlling programmed cell death of root velamen radicum in an epiphytic orchid.

Development of the velamen radicum on the outer surface of the root epidermis is an important characteristic for water uptake and retention in some plant families, particularly epiphytic orchids, for survival under water-limited environments. Velamen radicum cells derive from the primary root meristem; however, following this development, velamen radicum cells die by incompletely understood processes of programmed cell death (PCD). We combined the use of transmission electron microscopy, X-ray micro-tomography and transcriptome methods to characterize the major anatomical and molecular changes that occur during the development and death of velamen radicum cells of Cymbidium tracyanum, a typical epiphytic orchid, to determine how PCD occurs. Typical changes of PCD in anatomy and gene expression were observed in the development of velamen radicum cells. During the initiation of PCD, we found that both cell and vacuole size increased, and several genes involved in brassinosteroid and ethylene pathways were upregulated. In the stage of secondary cell wall formation, significant anatomical changes included DNA degradation, cytoplasm thinning, organelle decrease, vacuole rupture and cell wall thickening. Changes were found in the expression of genes related to the biosynthesis of cellulose and lignin, which are instrumental in the formation of secondary cell walls, and are regulated by cytoskeleton-related factors and phenylalanine ammonia-lyase. In the final stage of PCD, cell autolysis was terminated from the outside to the inside of the velamen radicum. The regulation of genes related to autophagy, vacuolar processing enzyme, cysteine proteases and metacaspase was involved in the final execution of cell death and autolysis. Our results showed that the development of the root velamen radicum in an epiphytic orchid was controlled by the process of PCD, which included initiation of PCD, followed by formation of the secondary cell wall, and execution of autolysis following cell death.

Root endophytic fungi promote in vitro seed germination in Pleurothallis coriacardia (Orchidaceae)


 Seeds of many orchids rely on the presence of fungi to trigger the germination process and even to initiate the full development of protocorms. While attention has been directed towards the study of mycorrhizal fungi, the diversity and functions of endophytic fungi from orchid roots remain underexplored, and few studies have verified their ecological role. This is the case of Pleurothallis coriacardia, an endemic green Neotropical orchid with both epiphytic and lithophytic habits growing in High-Andean montane forests. In the present study, we screened the cultivable fraction of the endophytic fungi colonizing the roots of mature plants of P. coriacardia using ITS rDNA markers. We also tested the potential of these endophytic fungi to improve embryo development and seed germination. Most of the isolated endophytes were classified within Psathyrellaceae. Some isolates, identified as members of the Ilyonectria and Coprinellus genera, significantly promoted embryo development in vitro in P. coriacardia seeds, a result that highlights the ecological roles these endophytic fungi may play in nature.
 Keywords: Andean montane forest, embryo development, epiphytic orchids, Ilyonectria, lithophytic orchids, Psathryrellaceae

Effects of light, agar, activated charcoal, and culture medium on the germination and early development of Dendrobium seedlings

The objectives of this study were to determine the optimal light conditions, agar concentration, and quantity of activated charcoal in asymbiotic media to improve the in vitro seed germination rate and early seedling development of the epiphytic orchids Dendrobium nobile Lindl. and Dendrobium phalaenopsis Fitzg. Two independent experiments with complete randomized designs were conducted. (1) Treatments were arranged in a split-plot scheme. Seeds in each sub-plot were exposed to one of four light conditions (dark, white fluorescent, red fluorescent + white fluorescent, or red fluorescent) and grown in one of four types of culture media (MS: Murashige and Skoog, ½ MS: half strength MS, K: Knudson C, and VW: Vacin and Went media). (2) Treatments were arranged in a 4 × 5 × 5 factorial scheme (four types of culture media: MS, ½ MS, K, and VW; five concentrations of agar: 0.0, 2.0, 4.0, 6.0, or 8 g L−1; and five concentrations of activated charcoal: 0.0, 1.5, 3.0, 4.5, or 6.0 g L−1). The highest germination rates and early seedling development were observed 45 days after seeding in the presence of white light for D. nobile and red + white light for D. phalaenopsis in MS and ½ MS culture media. Based on the findings of the present study, the use of MS and ½ MS culture media solidified with 4.0–8.0 g L−1 of agar and without activated charcoal is recommended for the optimal propagation of seeds and seedlings of these Dendrobium species.

In Vitro Propagation of Gastrochilus matsuran (Makino) Schltr., an Endangered Epiphytic Orchid.

Gastrochilus matsuran (Makino) Schltr. (Orchidaceae) populations are declining quickly because of overexploitation, climatic changes, and deforestation; therefore, mass-production protocols are required for this orchid. Natural propagation of this species is often hampered by meager seed germination and slow growth. Thus, our aim was to establish an effective protocol for the in vitro propagation of G. matsuran and reduce the risk of its extinction. We investigated the impacts of culture media, coconut water (CW), and plant hormones (gibberellic acid (GA3), indole-3-acetic acid (IAA), indole-3-butyric acid (IBA), α-naphthaleneacetic acid (NAA), and thidiazuron (TDZ)) on asymbiotic germination, multiplication and conversion of protocorms, and plantlet development. Maximal seed germination (93.3%) was achieved on ½ MS medium without vitamins plus 5% CW, 1 µM NAA, and 1.5 µM GA3. Secondary protocorm formation was best achieved on ½ MS medium without vitamins plus 2 µM TDZ. The conversion of protocorms into seedlings was maximized by supplementation with 2 µM IBA or 1 µM NAA. Acclimatized plantlets that exhibited exuberant growth on sphagnum moss were reintroduced to tree trunks in a natural habitat, with a 67% survival rate. This in vitro propagation procedure would be helpful for the mass production and conservation of this rare epiphytic orchid.

Orchid Diversity and Biomass on a Native Host Tree Species in a Semi-deciduous Rain Forest of Cameroon

ABSTRACT Triplochiton scleroxylon K. Schum is among an intensively exploited tree species in the Congo basin forest including the humid forest of Cameroon. As most tropical tree species, it is a potential host of epiphytic orchids for which little information is available. The aim of this study was to assess the biomass and orchids diversity that colonize T. scleroxylon in a semi-deciduous rain forest of Cameroon. Orchid species were inventoried exclusively on 20 felled T. scleroxylon. Orchid biomass obtained by destructive harvest was expressed in kg/tree. Orchid biomass was estimated in four randomized plots of 250 m × 20 m each, using allometric equations established. Twenty-nine orchid species were recorded on T. scleroxylon, while the mean orchid biomass was 1.23 kg/tree. Minimum T. scleroxylon diameter for orchid occurrence was 68.6 cm. The relationship between orchid biomass and T. scleroxylon diameter was low (Adj.R2˂0.4) and height did not improve the model. According to orchid biomass on T. scleroxylon (14.63 kg/ha), an estimated loss associated with T. scleroxylon’s exploitation was 10.47 kg/ha. This study expanded knowledge of orchid species linked to highly exploited T.scleroxylon in the Congo basin forest that impose conservation strategy, in the light of current degradation of their natural habitat and climate change mitigation.

Endophytic fungi associated with roots of epiphytic orchids in two Andean forests in Southern Ecuador and their role in germination

Orchids are known to establish complex relationships with endophytic fungi throughout their life cycle, and particularly during germination of their reserves-deprived seeds. Characterizing generalist or specialist interactions between orchids and associated fungi is key to supporting orchid conservation efforts. Here, endophytic fungi associated with roots of epiphytic orchids were studied in two montane Andean forests in Southern Ecuador. Orchid root samples were collected from ten ~500 m2 plots distributed between two neighboring forests. Endophytic fungi associated with these roots were then isolated, cultured, and identified by sequencing of rDNA markers. In total, 52 pure isolates were recovered from the roots of 10 orchid species. These isolates were classified into nine taxonomic groups except for one isolate that remained unclassified. Most fungal isolates were found in roots of up to two different orchid species; however, Coprinellus radians was found in the roots of all sampled orchids. The potential of C. radians to promote germination of orchid seeds was tested in a separate assay using seeds from two orchid species different than those found in the experimental forest plots. Of the two C. radians isolates tested, one improved germination in the two orchids evaluated to a level about half of that observed in seeds germinated in nutrient-rich medium (Phytamax) and above the null germination observed in plates without the fungus. Together, these results revealed a generalist relationship between C. radians and all the studied epiphytic orchids and the potential role of this fungus as a promoter of orchid seed germination.
 Key Words: Agaricales, Epidendrum, symbiotic germination

Isolation, characterization, and plant growth-promoting activities of endophytic fungi from a wild orchid Vanda cristata

ABSTRACTEndophytism is one of the widely explored phenomena related to orchids and fungi. Endophytic fungi assist plants by supplementing nutrient acquisition, and synthesis of plant growth regulators. Vanda cristata is an epiphytic orchid that has a great diversity of endophytic fungi. Endophytic fungi were isolated from roots, stems, and leaves of V. cristata and identified by both morphological and molecular study. Furthermore, the isolated endophytic fungi were subjected to auxin synthesis, phosphate solubilization, ammonia synthesis, and elicitor growth test for understanding their growth-promoting effect in a qualitative and quantitative manner. Altogether, 12 different endophytic fungi were isolated from roots, stems, and leaves of V. cristata of which most species belonged to Ascomycota. Unidentified II fungi were found to be most effective for auxin synthesis and phosphate solubilization while Agaricus bisporous and Mycolepto discus were most effective for ammonia synthesis. We have tested the plant growth-promoting activity of the twelve isolated endophytic fungi on Cymbidium aloifolium protocorms (12 weeks old). All the endophytic fungi showed growth-promoting activity. Plant growth of Cymbidium aloifolium was found higher on the MS medium supplemented with all fungal elicitors. Fungal elicitor CVS4, however, showed the highest plant growth-promoting activity toward C. aloifolium.

An assessment of the conservation status of Restrepia (Orchidaceae) reveals the threatened status of the genus

ABSTRACT Background The genus Restrepia occurs throughout Central and South America in areas of montane forest heavily affected by deforestation. Aims The current study was designed to test the feasibility of using available online resources to establish the threats facing these orchids and their conservation status for later inclusion in the IUCN online database. Methods Online resources were searched for primary data on the distribution of species of Restrepia. The Geospatial Conservation Assessment Tool (GeoCAT) was used to produce semi-automated IUCN Red List assessments. Locations of populations were examined in Google Earth to establish habitat loss. A comparison of the data produced a Red List assessment for each species. Results The observed losses of Restrepia habitat were: Venezuela 45% of recorded locations for 15 species, Colombia 28% for 30 species, Ecuador 36% for 18 species, Peru 41% for eight species, Costa Rica 81% and Panama 32% for three species. This habitat loss coincided with the route of the Pan-American Highway in these countries. Conclusions It was possible to establish the Red List Status of Restrepia species even with minimal data. The degree of threat facing these and other epiphytic orchid genera in these habitats was shown to be considerable.

Variation in root morphology of epiphytic orchids along small-scale and large-scale moisture gradients

Root morphology is expected to respond to environmental conditions. Two earlier studies reported predictable changes in the structure of the velamen radicum (common in monocotyledons) along rainfall gradients, which was expected from its assumed role in plant-water-relations. The present study expanded on this idea by analysing nine root traits that can be linked to root function, including velamen structure, along two moisture gradients of different spatial scales: 1) along the vertical gradient of a lowland forest in central Panama and 2) along a strong regional rainfall gradient in western Panama. All studied traits (e.g. velamen width, number of xylem poles, lignification of the exodermis) of the 45 orchid species showed substantial intraspecific and interspecific variation, but none of this (within forest) or very little of this (regional gradient) was related to the gradients. Only the community weighted means of velamen width, stele width and xylem pole number varied significantly with rainfall, with substantial effect sizes only found in the latter two traits. Our results clearly disagree with those of earlier publications, which did not include species abundance and/or rigorous statistics in their analyses. This discrepancy highlights that our understanding of the link between velamen form and function is still rudimentary.

Isolation and screening of extracellular enzymatic activity of endophytic fungi isolated from Thai orchids

Endophytes living inside plant tissues are known to be a rich source of unique bioactive compounds. In the current study, endophytic fungi were isolated from the leaves, stems and roots of five species of epiphytic orchids: Doritis pulcherrima, Dendrobiuma phyllum, Dendrobium anosmum, Ascocentrum curvifolium and Aerides falcata. In total, 52 endophytic isolates were obtained from the orchid samples, with the most (27) being obtained from leaf segments followed by roots and stems. All of the endophytic isolates were assessed for their potential role in producing extracellular enzymes, including amylase, protease, cellulase, pectinase and lipase on solid media. The results showed that 67%, 56%, 49%, 39% and 25% of the endophytic isolates had the ability to produce lipase, cellulase, pectinase, protease and amylase enzymes, respectively. One endophytic isolate, DapR 02, isolated from the root of Dendrobium aphyllum produced significantly more pectinase enzyme secretion with a ratio of the clear zone diameter to the colony diameter of 23. It was identified as Pseudopestalotiopsis theae using morphological characteristics and molecular analysis. This was the first report of the pectinase enzyme production potential of the endophyte Pseudopestalotiopsis theae. Therefore, an experiment was conducted to optimize the pectinase production in the DapR 02 isolate. High pectinase production of 1.524 U/mL was observed under culture conditions of 30 °C and pH 3.

Silicon toxicity induced by different concentrations and sources added to in vitro culture of epiphytic orchids

Silicon has been added to the in vitro culture medium of orchids as a beneficial element that can successfully improve tissue culture. However, although supplemental Si may induce toxicity to the orchid culture depending on the source, there are no reports on the toxic concentrations of this element. To this end, this study conducted two experiments with two species of orchids, whose seedlings were obtained by sowing capsules containing mature seeds under aseptic conditions. At 120 days after sowing (DAS), seedlings of the Cymbidium atropurpureo and Dendrobium secundum orchids were recultivated in MS medium plus treatments in a 3 × 5 factorial scheme, with three silicon sources (nanosilica, potassium silicate and sodium silicate mixture, and monosilicic acid) at five different concentrations: 0.00 (control), 14.15, 28.30, 42.45, and 56.6 mmol L−1 Si following a completely randomized design with four repetitions and 10 seedlings each. The orchid culture ended after 300 DAS and the seedlings were removed from the flasks to evaluate the Si accumulation in plant tissues, the physiological data, pigments, biometric data and the survival percentage after acclimatization of 20 seedlings from each treatment. The results show that Si added to in vitro orchid cultivation may either favor growth or cause toxicity depending on the element source and concentration in the culture medium and the orchid species. Nanosilica at the 21.4 mmol L−1 Si concentration was the most promising source for orchid cultivation for Cymbidium atropurpureo. Further, monosilicic acid was considered the most toxic source due to the physiological damage caused to both Cymbidium atropurpureo and Dendrobium secundum at concentrations higher than 14.1 and 11.2 mmol L−1 Si, respectively.

KEANEKARAGAMAN JENIS ANGGREK ALAM EPIFIT PADA BERBAGAI TUTUPAN TAJUK HUTAN DI DESA BALAI SEBUT KABUPATEN SANGGAU

Epiphytic orchids can be found living naturally in the tropical rainforest. Destruction or degradation of forest area in Balai Sebut District Sanggau Regency due to caused degradation of canopy cover. This condition may affect microclimate which than impacts at the presence of species, one of which is epiphytic orchids that grow naturally in the forest. This study used a survey method with purposive double plot sampling for 1 month (28 Mei – 21 July 2019) in the field. Observation plots were made with a size of 20 x 50 m with a total of 12 plots. The results showed that there where 32 types of epiphytic orchids with a total of 431 individuals. At closed forest canopy cover and had been accomplished (>70%) can be 16 species of orchids with 220 individuals, at forest canopy (50-70%) can be 25 species of orchids with 159 individuals, can be at open forest canopy cover (50%) there were 10 species of orchids with 52 individuals. The dominant orchids species in the three canopy cover Flikingeria bicostata with Celogne peltastes.Keywords: closed forest canopy, epiphytic orchids, medium and open forest canopy, species diversity.

Biodiversity exploration of host trees (phorophyte) of epiphyte orchids in the natural habitat

Epiphyte orcids host trees grow naturally in tropical rainforests. Nowadays tropical rainforest destruction could decreased these specific host trees alongside with the various epiphyte orchids. Little information could be found on epiphyte host trees in Indonesia. The aim of this research was to explore the biodiversity of epiphyte orchid host trees (phorophytes) at Highway Forest Park Raden Soeryo and supported by exploration data in 2003, 2005, 2018 at Bromo Tengger Semeru National Park, and in 2006, 2007 at Highway Forest Park R. Soeryo. Exploration of epiphyte orchid host trees at Jogging Track site and Coban Watu Ondo site, Highway Forest Park R. Soeryo, East Java was conducted in May-July 2019. The research methode was carried out using a purposive sampling methode at Jogging Track and line transect at Coban Watu Ondo. From the main transect 30 plots were made with the area of each plot 20 m x 20 m with a distance between plots of 60 m at Jogging Track site and 100 m at Coban Watu Ondo. From the exploration we found at Jogging Track site 15 species of 173 hosts trees with diversity index 1.57 (medium). While at Coban Watu Ondo 10 species of 171 host trees with diversity index 1.52 (medium). Host trees bark characterizes by rough. grooved, hard skin no latex, tree diameter 20.00-85.00 cm. canopy medium-sparse. The biodiversity of porophyte trees exist in the area of National Park Bromo Tengger Semeru and Highway Forest Raden Soeryo south of Mt Arjuno Lalijiwo and east of Mt Anjasmoro regions is quite diverse.

An Account of Epiphytic Orchids of Nainital in Relation to Their Host Specificity

Orchids are highly evolved and specialized group of flowering plants belonging to family Orchidaceae. It is the second largest family of the flowering plants in the world with more than 24,000 species of perennials that includes terrestrial, saprophytic, lithophytic and epiphytic species. The present work is based on the systematic ephiphytic orchids observations based on habit, habitat, colour of flower, flowering period and detailed morphological features. Twenty-two epiphytic orchid species belonging to fifteen genera were recorded from Nainital area. The genera represented were Dendrobium (four species), Aerides, Bulbophyllum, Eria and Oberonia (two species each) while rests were represented by single species each. The host species such as Quercus leucotrichophora (6), Sapium insigne (5), Syzygium operculatum, Bombax ceiba and Pyrus pashia (4) supported maximum number of epiphytic orchids followed by the species Syzygium cumini and Toona ciliata (3). Four host species reported with single orchid species. Due to high anthropogenic pressure and changing environmental conditions resulted in vulnerability of these species. Therefore, regular monitoring of the species, mass scale propagation and creating awareness among the local inhabitants are suggested.

Comparative Seed Morphology of Tropical and Temperate Orchid Species with Different Growth Habits.

Seed morphology underpins many critical biological and ecological processes, such as seed dormancy and germination, dispersal, and persistence. It is also a valuable taxonomic trait that can provide information about plant evolution and adaptations to different ecological niches. This study characterised and compared various seed morphological traits, i.e., seed and pod shape, seed colour and size, embryo size, and air volume for six orchid species; and explored whether taxonomy, biogeographical origin, or growth habit are important determinants of seed morphology. We investigated this on two tropical epiphytic orchid species from Indonesia (Dendrobium strebloceras and D. lineale), and four temperate species from New Zealand, terrestrial Gastrodia cunnninghamii, Pterostylis banksii and Thelymitra nervosa, and epiphytic D. cunninghamii. Our results show some similarities among related species in their pod shape and colour, and seed colouration. All the species studied have scobiform or fusiform seeds and prolate-spheroid embryos. Specifically, D. strebloceras, G. cunninghamii, and P. banksii have an elongated seed shape, while T. nervosa has truncated seeds. Interestingly, we observed high variability in the micro-morphological seed characteristics of these orchid species, unrelated to their taxonomy, biogeographical origin, or growth habit, suggesting different ecological adaptations possibly reflecting their modes of dispersal.

Genome-wide identification and expression profiling of SET DOMAIN GROUP family in Dendrobium catenatum

BackgroundDendrobium catenatum, as a precious Chinese herbal medicine, is an epiphytic orchid plant, which grows on the trunks and cliffs and often faces up to diverse environmental stresses. SET DOMAIN GROUP (SDG) proteins act as histone lysine methyltransferases, which are involved in pleiotropic developmental events and stress responses through modifying chromatin structure and regulating gene transcription, but their roles in D. catenatum are unknown.ResultsIn this study, we identified 44 SDG proteins from D. catenatum genome. Subsequently, comprehensive analyses related to gene structure, protein domain organization, and phylogenetic relationship were performed to evaluate these D. catenatum SDG (DcSDG) proteins, along with the well-investigated homologs from the model plants Arabidopsis thaliana and Oryza sativa as well as the newly characterized 42 SDG proteins from a closely related orchid plant Phalaenopsis equestris. We showed DcSDG proteins can be grouped into eight distinct classes (I~VII and M), mostly consistent with the previous description. Based on the catalytic substrates of the reported SDG members mainly in Arabidopsis, Class I (E(z)-Like) is predicted to account for the deposition of H3K27me2/3, Class II (Ash-like) for H3K36me, Class III (Trx/ATX-like) for H3K4me2/3, Class M (ATXR3/7) for H3K4me, Class IV (Su (var)-like) for H3K27me1, Class V (Suv-like) for H3K9me, as well as class VI (S-ET) and class VII (RBCMT) for methylation of both histone and non-histone proteins. RNA-seq derived expression profiling showed that DcSDG proteins usually displayed wide but distinguished expressions in different tissues and organs. Finally, environmental stresses examination showed the expressions of DcASHR3, DcSUVR3, DcATXR4, DcATXR5b, and DcSDG49 are closely associated with drought-recovery treatment, the expression of DcSUVH5a, DcATXR5a and DcSUVR14a are significantly influenced by low temperature, and even 61% DcSDG genes are in response to heat shock.ConclusionsThis study systematically identifies and classifies SDG genes in orchid plant D. catenatum, indicates their functional divergence during the evolution, and discovers their broad roles in the developmental programs and stress responses. These results provide constructive clues for further functional investigation and epigenetic mechanism dissection of SET-containing proteins in orchids.

Algal associates and the evidence of cyanobacterial nitrogen fixation in the velamen roots of epiphytic orchids

The hanging velamen roots in epiphytic orchids are unique kind of aerial roots in the plant world. The diverse kinds of microbial associates in it suggest its ecological and functional complexities. In this context, diversity and population characteristics of the entire algal flora in the velamen roots of four tropical epiphytic orchids Acampe praemorsa, Rhynchostylis retusa, Epidendrum radicans and Oncidium sphacelatum from different geographic locations of Kerala state, India were explored and compared. The potential nitrogen-fixing activity (NFA) of cyanobacteria in the velamen roots was analysed by acetylene reduction assay to understand the nitrogen-fixing role of cyanobacterial species in the same as a source of nitrogen to the host. The total nitrogen content of the velamen roots and leaves was compared to generate indirect evidence of accumulation of nitrogen in roots by primary addition through nitrogen fixation. Altogether 24 species of algal associates were isolated from the velamen roots of four epiphytic orchids. A majority of algae were found as cyanobacterial species; many of them are well-known nitrogen-fixing categories. The total tissue nitrogen content of the velamen roots was found significantly higher than that of leaves. The NFA of cyanobacteria in the velamen roots revealed its potential role in nitrogen fixation. This preliminary level empirical evidence is sufficient to consider the velamen roots of epiphytic orchids as an ecologically complex aerial ‘organ ecosystem’ with microbial partnerships for nutritional support, especially nitrogen availability in epiphytic orchids.

Chloroplast characterization and phylogenetic relationship of Cymbidium aloifolium (Orchidaceae)

Cymbidium aloifolium is an epiphytic orchid with high medicinal and ornamental value. In order to get a deeper understanding of C. aloifolium, we determined the complete chloroplast genome of C. aloifolium by Illumina sequencing data. The length of this genome is 157,328 bp, including a couple of inverted repeat (IR) regions of 26,829 bp, a large single-copy (LSC) region of 85,793 bp, and a small single-copy (SSC) region of 17,877 bp. The chloroplast genome comprised of 139 genes, including 78 protein-coding genes, 38 tRNA genes, and 8 rRNA genes. In addition, the phylogenetic analysis based on 17 chloroplast genomes of Orchidaceae indicated that C. mannii was closely related to C. aloifolium. This study will provide more valuable information for the classification and phylogenetic research of Cymbidium genus.

Does the orchid Luisia teres attract its male chafer pollinators (Scarabaeidae: Protaetia pryeri pryeri) by sexual deception?

The epiphytic orchid Luisia teres (Asparagales: Orchidaceae) releases floral scent that attracts males of the cupreous polished chafer Protaetia pryeri pryeri (Coleoptera: Scarabaeidae) for pollination. We analyzed this floral scent to identify the attractant(s). When various flower parts were extracted with diethyl ether and assayed, male chafers were attracted to the extract of petals but not to that of sepals or lips. The petal extracts were chromatographed on silica gel column and the activity was found in ether and methanol fractions. Gas chromatography/mass spectrometry (GC/MS) analyses and subsequent NMR analyses revealed that both active fractions contained 2,3-dihydroxypropyl isovalerate (2,3-DHPiV) as a major compound. Natural 2,3-DHPiV showed identical retention time to that of synthetic (R)-enantiomer in subsequent HPLC analyses using a chiral column, while (S)-enantiomer was not detected. The amount of this compound was ca. 5 µg/flower. In field tests performed in Okinawa, Japan, males of P. pryeri pryeri were observed to fly around and land on a cotton ball impregnated with synthetic (R)-2,3-DHPiV, but no male was attracted to the (S)-enantiomer or the racemic mixture. Thus, the chemical attractant released from the flower was identified to be (R)-2,3-DHPiV. 2,3-DHPiV was also found in a volatile sample collected from the virgin females by GC/MS analyses using achiral column, while this compound did not occur in males. Because of the difficulty in HPLC detection of a small amount of 2,3-DHPiV, the chirality remains to be determined. Nevertheless, these results strongly suggest that the flowers of L. teres release the same compound as females of its pollinator P. pryeri pryeri, indicating that the flower of this orchid chemically mimics the female sex pheromone.

Mycoendophytic diversity and their antimicrobial potential from two epiphytic orchids of the Western Ghats forests of India

Mycoendophytic diversity and their antimicrobial potential from two epiphytic orchids of the Western Ghats forests of India