Orchid Leaf Research Articles

Diversity induction with gamma-ray irradiation on Phalaenopsis amboinensis var. Common natural orchid generation M1V0

Phalaenopsis is a popular genus in the Orchidaceae family. Phalaenopsis amboinensis orchid is an endangered natural orchid, so it needs to be preserved. Conservation of natural orchid Phalaenopsis amboinensis var. Common way is by increasing genetic diversity. This research aims to determine the characteristics and morphological diversity of Phalaenopsis amboinensis natural orchid irradiated gamma-ray results and without gamma-ray irradiation; this research was conducted in two places: the implementation of gamma-ray irradiation at the PAIR-BATAN Laboratory and in Matesih, Karanganyar with an average height of 450 m above sea level. The research was conducted from December 2022 to May 2023. The study used a simple design of one factor, the doses of gamma-ray irradiation with 5 levels: 0 Gy (R0), 7.5 Gy (R1), 15 Gy (R2), 22.5 Gy (R3), 30 Gy (R4). Each treatment contained five accessions. So, there are 25 experimental units. Observational data were analyzed descriptively by comparing observational data from Phalaenopsis amboinensis var. Common Generation M1V0 orchids irradiated with control plants. The results showed diversity in the morphology of the Phalaenopsis amboinensis var. Common orchid Generation M1V0 results from gamma-ray irradiation based on qualitative morphology with phenotypic observations, showing two orchid leaf shapes: oblanceolate and elliptic. There are accessions with fasciated root conditions.

Induksi Nanobubbles (NBs) untuk Pertumbuhan Anggrek Dendrobium Imelda Marina Masagung X Bumi Menangis

Nanobubbles (NBs), nanotechnology has been applied in several fields, such as health, waste management, agriculture, and others. This nanotechnology utilizes the gas in the bubbles, making it easier for the gas to enter the cell. This study aimed to determine the effect of O2, CO2, N2, H2, and O3 gases in NBs on the growth of the Dendrobium Imelda Marina Masagung ´ Bumi Menangis orchid. The method used was to provide fertilizer and NBs treatment to Dendrobium orchids for six weeks with observations every one week. The data obtained were analyzed by two-way ANOVA and further tested post hoc tests with Ptukey <0.05. The results showed significant differences in stem diameter and orchid plant height. The orchid stem diameter was more significant with the NBs treatment than the fertilizer and control treatments. Orchid plants with NBs treatment were higher than those with fertilizer and control treatments. The appearance of new shoots and roots also indicates plant growth. Orchid leaf colour with fertilizer and NBs treatment changed from the Yellow-Green Group (144) to the Green Group (143), except for the control. Leaves colour affects the process of photosynthesis, the green colour will carry out photosynthesis better than Green-Yellow leaves. This affects plant growth in Dendrobium Imelda Marina Masagung ´ Bumi Menangis orchid. NBs enlarge the stem, increase the height and make the leaves green.

Do photosynthetic metabolism and habitat influence foliar water uptake in orchids?

Epiphytic and rupicolous plants inhabit environments with limited water resources. Such plants commonly use Crassulacean Acid Metabolism (CAM), a photosynthetic pathway that accumulates organic acids in cell vacuoles at night, so reducing their leaf water potential and favouring water absorption. Foliar water uptake (FWU) aids plant survival during drought events in environments with high water deficits. We hypothesized that FWU represents a strategy employed by epiphytic and rupicolous orchids for water acquisition and that CAM will favour increased water absorption. We examined 6 epiphyte, 4 terrestrial and 6 rupicolous orchids that use C3 (n=9) or CAM (n=7) pathways. Five individuals per species were used to evaluate FWU, structural characteristics and leaf water balance. Rupicolous species with C3 metabolism had higher FWU than other species. FWU (Cmax and k) could be related to succulence, SLM and leaf RWC. The results indicated that high orchid leaf densities favoured FWU, as area available for water storage increases with leaf density. Structural characteristics linked to water storage (e.g. high RWC, succulence), on the other hand, could limit leaf water absorption by favouring high internal leaf water potentials. Epiphytic, rupicolous and terrestrial orchids showed FWU. Rupicolous species had high levels of FWU, probably through absorption from mist. However, succulence in plants with CAM appears to mitigate FWU.

Simultaneous detection of odontoglossum ringspot virus (ORSV) and cymbidium mosaic virus (CymMV) infecting orchids in Meghalaya, India by Multiplex RT-PCR

• ORSV was found to be more prevalent than CymMV with the incidence percentages of 53.5% and 46.4% respectively. • Mixed infection of ORSV and CymMV was found to be 35.7%. • The Nucleotide sequences of ORSV and CymMV CP genes from Meghalaya showed high sequence identities of 96.90–99.56% and 96.91–99.66% respectively with other available sequences in Genbank. Odontoglossum ringspot virus (ORSV) and cymbidium mosaic virus (CymMV) are the most prevalent viruses infecting orchids all around the world. These viruses cause a serious threat to the orchid industry as they can spread from infected plants to healthy plants through vegetative propagation, contaminated tools, and human hands. This result in negative vigour of infected plants, showing stunted growth, producing fewer flowers, and decreasing flower quality. In this study, 84 leaf samples of orchids were collected from nine horti-hubs and orchid nurseries in Meghalaya. The orchid leaf samples were subjected to multiplex RT-PCR amplification using gene-specific primers targeting the coat protein (CP) genes of CymMV and ORSV. The most prevalent virus detected was ORSV with a 53.5% incidence while CymMV showed a 46.4% incidence. Mixed infections of ORSV and CymMV were detected in 30 samples (35.7%). The nucleotide sequences of ORSV and CymMV CP genes from Meghalaya showed high sequence identities of 96.90–99.56% and 96.91–99.66%, respectively when compared to other sequences in GenBank. The phylogenetic relationship among ORSV isolates and CymMV, based on infected host or geographical origin, showed no clear-cut clustering of isolates.

Identification and Characterization of Soft Rot Bacterial Pathogens on Phalaenopsis Orchid in Bali

The moth orchid (Phalaenopsis sp.) is one of the most popular orchids due to the various colors with distinctive shapes of the flowers. Soft rot disease caused by Pectobacteriaceae (SRP) family is commonly found infected this plant. The infected orchid showed pale-colored to blackish slimy rot. This research was conducted in three locations namely Denpasar, Badung, and Karangasem, and resulted in 10 candidates for pathogenic bacteria. The pathogenicity test of the pathogen candidates was carried out by injecting bacterial suspension into the orchid leaf tissue with the result of 6 bacterial isolates showing soft rot symptoms. Moreover, two specific primers Dda1F-Dda1R and Pcc3F-Pcc3R for Dickeya spp. and Pectobacterium spp. consecutively were used for the PCR test. The electrophoresis result of the PCR product identified the bacteria isolated from infected plants as Dickeya spp. Dickeya spp. showed white to yellowish-white colony color, with convex and circular colony form on PDA medium.

First Report of Dickeya fangzhongdai causing soft rot in Orchids in Canada.

Dickeya fangzhongdai was originally described as the causal agent of bleeding canker of pear tree in China. Recently, D. fangzhongdai was isolated and identified as the causal agent of soft rot in an orchid plant purchased in a local supermarket in Prince Edward Island, Canada. A water-soaked dark green spot on the leaf surface was observed and later became larger soft rot symptom. The origin of the orchid plants was traced back to a producer in Ontario, Canada who propagated them from with cuttings originally imported from the Netherlands and Taiwan. Bacterial isolations were made from a soft rot lesion on an orchid leaf by surface sterilization of small pieces of marginal tissue of the diseased leaf in 70% alcohol. The small pieces of leaf tissue were then washed three time using sterile water, and immersed in drops of sterile water. Bacterial streaming was observed under the microscope and non-fluorescing bacterial colonies were isolated on King's B and casamino acid-peptone-glucose agar plates and purified as isolates 908, 909, 910 and 911. The DNA samples were extracted from the four isolates, as well as the diseased leaf tissue, and tested by using a qPCR assay with the specific primer/probe set (DfF/DfR/DfP) for D. fangzhongdai (Tian et al. 2020). The assay yielded PCR amplicons of 135 bp with a melting temperature of 86.5±0.6 °C as did two control reactions using genomic DNA from D. fangzhongdai strains JS5T and QZH3 originally isolated in China, providing presumptive identification of the orchid isolates as D. fangzhongdai. To fulfill Koch's postulates, freshly purchased healthy orchid plants (n=4) were inoculated by leaf injection with the bacterial isolates obtained in this study and strains JS5 T and QZH3 at ~107 CFU/ml. Three leaves of the same side of the plants were inoculated with the same strains as triplicates. Sterile water was used as the negative control. Inoculated plants were incubated in a growth chamber with a 16 h photoperiod at 23 °C. Water soaked lesions developed in 3-5 days after inoculation followed by soft rotting in leaves inoculated with the new bacterial strains from orchid plants while strain QZH3 caused soft rot in 10 days after inoculation (Fig. S1). The non-fluorescing bacteria on King's B plates with colony morphology similar to those inoculated were re-isolated from the inoculated leaves and confirmed to be D. fangzhongdai by qPCR. Phylogenetic analysis of the assembled 16S rRNA sequence of isolate 908 (GenBank accession number: MT984340), together with GenBank data of all Dickeya spp. and some Pectobacterium spp, using neighbor-joining (NJ) method inferred with MEGA X software (Kumar et al. 2018) showed that isolate 908 clustered with strains JS5T and QZH3 at a phylogenetic distance of 0.0007. This clearly indicated that isolate 908 and JS5T and QZH3 belong to the same genus. Species-level identification of isolate 908 was achieved by genome sequencing and analysis based on average nucleotide identity (ANI). Genomic DNA of isolate 908 was sequenced with Illumina MiSeq to provide approximately 180X genome coverage. After quality checking using FastQC (Andrews 2010), de-novo assembly was performed with VelvetOptimiser v2.2.6 (Zerbino and Birney 2008). The draft genome size of strain 908 was 4,938,027 bp consisting of 76 contigs with 56.8% G+C content and 63,801 bp as N50. The draft genome was checked for misassembled fragments using QUAST v5.0.2 (Gurevich et al. 2013) and found to be of good quality. The draft genome sequence is deposited in GenBank under the accession number of JADCNJ000000000. The draft genome sequence of strain 908 was compared to that of D. fangzhongdai JS5T type strain genome using FastANI v1.2 (Jain et al. 2018) resulting in an ANI value of 98.9%, which is above the 95% cut-off for the same species. Previously, it was reported that D. fangzhongdai caused soft rot in orchid in Europe (Alič et al. 2018) and in onions in New York (Ma et al. 2020). The difference in virulence among D. fangzhongdai strains warrants further investigation and their pathogenicity on potato is being investigated to evaluate any threat to the potato industry. To our knowledge, this is the first report of D. fangzhongdai causing soft rot disease on orchids in Canada and North America.

Induction of Rhizoctonia resistance to odontoglossum ringspot virus (ORSV) infection on orchid leaf organ viability

Orchid plants are in great demand by the public as ornamental plants that have economic value and the beauty of attractive flower shapes and colors, especially Phalaenopsis and Dendrobium. Related to this, conservation efforts are needed through cultivation. However, these efforts are often hampered by viral infections. One of the viruses that trigger inhibition of orchid cultivation that often appears is ORSV. Control efforts that are relatively safe and efficient can utilize mycorrhizae in orchids which play a role in increasing the absorption of plant nutrients. The use of Phalaenopsis and Dendrobium with the induction of Rhizoctonia sp. against ORSV is a very important alternative biofertilizer and biocontrol agent. The objectives of this study were 1) to determine the effectiveness of orchids induced by Rhizoctonia resistance, 2) to determine the viability of orchid leaf organs to ORSV infection, 3) to determine the viability of orchid root organs resulting from Rhizoctonia resistance to ORSV infection. This study was conducted in January 2021 – March 2021 at the Botanical 2 Laboratory, Department of Biology, Faculty of Mathematics and Natural Sciences, University of Lampung. This study employed a completely randomized factorial with 6 treatments and 4 replications. The variable observed in this study was the effectiveness of mycorrhizae, leaf and root length, number of leaves and roots, and leaf width. The data obtained were homogenized using Levene's test and then analyzed by ANOVA and Tukey's further test at the 5% level. The results showed that the effectiveness of mycorrhizae occurred on the 3rd day of incubation. In leaf organs, Phalaenopsis amabilis showed an increased response and Dendrobium discolour showed a decreased response in leaf length, width, and number. Meanwhile, in the root organ, Dendrobium discolour showed an increased response and Phalaenopsis amabilis showed a decreased response in root length and number of roots.

Determination of total nitrogen content in fresh leaves and leaf powder of Dendrobium orchids using near-infrared spectroscopy

Appropriate application of nitrogen (N) fertilizer promotes plant growth, inflorescence yield, and flower quality of orchids. In this study, we used the near-infrared spectroscopy (NIRS) technique to develop a prediction model of the N content in the leaf of Dendrobium orchid, which is an essential indicator for monitoring plant health. The Dendrobium orchid samples were foliar sprayed in rotation between 20N–20P–20K and 30N–10P–10K fertilizers at a frequency of once or twice a week to create a diverse amount of leaf N content. An application of water was used as the control treatment. After nine months of fertilizer treatments, 150 fresh orchid leaf samples, containing various N contents, were scanned using Fourier-transform near-infrared spectroscopy (FT-NIRS). Then the samples were dried and ground to a fine powder and were again scanned. The absorbance spectra were collected at the 12,000–4000 cm−1 (800–2500 nm) region. Total N content was determined by using the combustion method. The result showed a high proficiency in the estimation of N in leaf powder with the correlation coefficient of prediction (Rp), the root mean square error of prediction (RMSEP), and the ratio of standard deviation of reference data of prediction set to standard error of prediction (RPD) being 0.9882, 0.0637% dry weight (DW), and 6.53, respectively. The fresh leaf sample was successfully predicted with Rp, RMSEP, and RPD of 0.8874, 0.207% DW, and 2.13, respectively. Additionally, external validation confirmed the high reliability of using leaf powder since the correlation coefficient of external validation (rp) and RMSEP of the external validation achieved 0.9651 and 0.1405% DW, respectively, while the external validation of fresh leaf should be improved with rp = 0.7438 and RMSEP = 0.2652% DW. The overall results suggested that NIRS can be used for monitoring the N status in orchids, especially in leaf powder, with high accuracy.

GC-MS analysis reveals Dendrobium candidum is a mosquito-attractant orchid with mosquitocidal compounds

Mosquito, the infamous vector organism, is responsible for outbreaks of several deadly epidemics. Semiochemicals or pheromones and some other compounds released from several plants and animals have ability to attract mosquitoes. Heneicosane is one of such compounds reported from some plants that attracts mosquitoes, especially Aedes spp. (vector of dengue). Through our experiment based on GC-MS analysis, we report that leaf of Dendrobium candidum Wall. ex Lindl, a local orchid of Darjeeling Himalayas, is a source of heneicosane along with several other semiochemicals and odorants that can target mosquitoes. Interestingly, the orchid sample has revealed a number of components having insecticidal activities against mosquitoes which can have made this orchid a complete mosquito-bait. This research invites interest for Dendrobium candidum as a mosquito attractant and killer. For this unique property, we recommend this orchid leaf extract as a bait for dengue vector Aedes mosquito.

Wet Transfer of Inkjet Printed Graphene for Microsupercapacitors on Arbitrary Substrates

Significant research interest is being devoted to exploiting the properties of graphene but the difficult integration on various substrates limits its use. In this regard, we developed a transfer technique that allows the direct deposition of inkjet printed graphene devices on arbitrary substrates, even 3D objects and living plants. With this technique, we fabricated microsupercapacitors, which exhibited good adhesion on almost all substrates and no performance degradation induced by the process. Specifically, the microsupercapacitor on an orchid leaf showed an areal capacitance as high as 441 μF cm–2 and a volumetric capacitance of 1.16 F cm–3. This technique can boost the use of graphene in key technological applications, such as self-powered epidermal electronics and environmental monitoring systems.

Host Tree Identity Influences Leaf Nutrient Relations of the Epiphyte Dendrobium guamense Ames.

Epiphytic orchid species grow in stressful conditions that include nutrient deficiency. A greater understanding of the factors that influence epiphytic orchid nutrition may benefit conservationists and horticulturists. Green and senesced leaf nutrient concentrations of the epiphytic Dendrobium guamense were quantified among six host tree species to more fully understand the role of contrasting host identity. Green leaves for D. guamense plants on species with vertical stems contained greater nitrogen, calcium, iron, manganese, zinc, and boron concentrations. Green leaves for D. guamense plants on species with horizontal stems contained greater phosphorus, magnesium, and copper concentrations. Senesced D. guamense leaves followed similar patterns among the host species for all elements except calcium, manganese, and boron. The stoichiometry among green leaf macronutrients indicated nitrogen was most limiting, and the limitation was more severe for plants on horizontal stems. Carbon:nitrogen of senesced leaves indicated D. guamense plants on vertical stems produced higher quality leaf litter, predicting slower decomposition of orchid leaf litter from plants growing on hosts with horizontal stems. The results may improve horticultural decisions to support conservation efforts and show that host identity is a variable that should be more fully studied to understand epiphyte biology.

Leaf spot on cattleya orchid caused by Neoscytalidium orchidacearum in Thailand

Leaf spots caused by plant pathogenic fungi are among the main diseases affecting orchid plants worldwide. A leaf spot disease was widespread in a nursery of cattleya orchid (Cattleya lueddemanniana var. lueddemanniana) in Lampang Province, Thailand in 2016. A fungus was isolated from leaf lesions and identified as Neoscytalidium orchidacearum based on morphological characteristics and the identification was confirmed using a molecular phylogenetic analysis of the combined internal transcribed spacer and large subunit regions of ribosomal DNA sequences. The isolated fungus caused spot symptoms on inoculated orchid leaves similar to symptoms observed in the field. This is the first report of orchid leaf spot disease caused by N. orchidacearum.

Novel and divergent viruses associated with Australian orchid-fungus symbioses

Terrestrial orchids represent a symbiotic union between plants and mycorrhizal fungi. This study describes the occurrence and nature of viruses associated with one population of wild Pterostylis sanguinea orchids, including their fungal symbionts, over two consecutive years. A generic sequencing approach, which combined dsRNA-enrichment from plant and mycelial tissues, random amplification and high throughput shotgun sequencing was used to identify novel viruses. The majority of the virus-like sequences represent partial genomes, and their identification is based solely on de novo assembly of sequencing data. In orchid leaf tissues we found three isolates of a novel totivirus and an unclassified virus; both resemble fungus-infecting viruses. Two isolates of Ceratobasidium sp that were isolated from orchid underground stems contained at least 20 viruses, 16 of which were previously described as alphapartitiviruses and betapartitiviruses. A novel hypovirus and a mitovirus were genetically distant from existing members of the genera and did not readily fit into recognised subgroups.

Orchid Leaf Search Project – July 9 2016

Orchid Leaf Search Project – July 9 2016

The Segmentation of Orchid Leaf Disease Spot Based On Lab Color Space and Otsu

The Segmentation of Orchid Leaf Disease Spot Based On Lab Color Space and Otsu

Combination of a simple differential medium and toxA-specific PCR for isolation and identification of phytopathogenic Burkholderia gladioli

The NGM medium developed in a previous study was used for differential isolation of Pectobacterium chrysanthemi, Burkholderia gladioli, and B. glumae. P. chrysanthemi developed blue colonies, and all B. gladioli and B. glumae strains tested produced diffusible yellow pigments on the NGM medium, easily distinguishable from other Burkholderia spp. and plant pathogenic bacteria. The produced yellow pigments contained a toxoflavin determined by the TLC and orchid leaf chlorosis tests. A specific oligonucleotide primer pair was designed for the detection of toxA, which is involved in toxoflavin biosynthesis. All B. gladioli and B. glumae strains tested contained toxA as determined by PCR amplification. No amplification was observed with other plant pathogenic bacteria. In addition, the toxA-based species-specific PCR assays, based on the nucleotide sequence differences in the promoter region of toxA, were developed for identification of B. gladioli and B. glumae, respectively. The NGM medium and the toxA-based PCR assays were used to determine the causal agents of leaf rot of Phalaenopsis and Oncidium orchids at three cultivation areas in Taiwan. It was found that both P. chrysanthemi and B. gladioli are important pathogenic bacteria of orchid leaf rot in Taiwan. The results indicate that the combination of NGM medium with toxA-based PCR assays is a newly designed and efficient method for isolation and identification of leaf rot pathogenic bacteria especially from plant hosts on which P. chrysanthemi and B. gladioli (or B. glumae) could cause symptoms.

DISINFECTION OF ODONTOGLOSSUM RINGSPOT VIRUS AND CYMBIDIUM MOSAIC VIRUS FROM TOOLS USED DURING ORCHID CULTIVATION

A novel sanitation approach by using liquid culture filtrate (CF) of a Streptomyces isolate (CA5) that can readily disinfect coat proteins (CP) of Odohtoglossum ringspot virus (ORSV) and Cymbidium mosaic virus (CymMV) and eliminate their infectivities is demonstrated. By treating for only 30 s with a preparation of CA5 CF on the materials or tools commonly used in orchid nurseries including razor blades, plastic clips, labels and gloves coated previously with ORSV and CymMV infected plant sap, it is shown that no ELISA signals and infectivity could be recovered on the treated materials. Similarly, virus contaminated human nails and surfaces of Phalaenopsis orchid leaf tissue can also be treated without detecting any leftover ELISA signal and virus infectivity. We also showed that CA5 CF has the potential to eliminate virus contamination on the surface of orchid seeds leading to the production of clean orchid seedlings. The virus degradation activity of CA5 culture filtrate is heat labile to temperature treatments higher than 60°C, indicating that the activity is possibly due to a proteinaceous substance. Further experiments of dialyzing CA5 culture filtrate using different molecular cut-off membranes showed that the molecular mass of the substance should be larger than 100 kDa.

An investigation to determine variation in marsh orchid (Dactylorhiza) populations at Moses Gate Country Park, Bolton

In June 2006, two populations of Dactylorhiza were examined at Moses Gate Country Park, in Greater Manchester, to determine whether variation exists between the two populations. Careful measurements of orchid characteristics were taken at both sites, in addition to soil pH recordings, and the results compared and tested for significance using one-way ANOVA and Chi-square significance tests. Descriptive statistics and box-plots showed that there were indeed differences in orchid height, leaf width and leaf number between the two populations, with one-way ANOVA confirming their significance as P, 0.001 for all the variables. Further analyses using frequency data and Chi-square tests also showed that there were significant variations in spur size, leaf type, labellum shape and petal colour between the two sites with P, 0.001 for all these characteristics. Soil pH data were also analysed and tested using one-way ANOVA which showed that there were significant variations in soil pH levels at both sites with P, 0.001 for this data. It was concluded that there were indeed significant differences between the two populations of Dactylorhiza at Moses Gate Country Park, therefore the null hypothesis was rejected. It was suggested that further investigations could be undertaken at this site, using similar methods, over a number of years, to support this study and to provide further constructive information on the status of the two populations of orchids, which may increase the conservation value of the site.

Impatiens necrotic spot virus and Tomato spotted wilt virus Diagnosed in Phalaenopsis Orchids from Two Florida Nurseries.

In October 2006 (Arcadia, FL) and January 2007 (Sorrento, FL), several white Phalaenopsis orchids with large chlorotic/necrotic ringspot symptoms were sent to the Division of Plant Industry, Gainesville, FL. Symptomatic leaf tissues were tested with the Agdia immunostick-comb (Agdia, Elkhart, IN) for Impatiens necrotic spot virus (INSV), Tomato spotted wilt virus (TSWV), Cucumber mosaic, and Tobacco mosaic virus. Plants from the nursery in Sorrento, FL tested positive for TSWV, while those from the nursery in Arcadia, FL tested positive for INSV. Symptomless leaves from the infected plants tested negative for the viruses with the immunostick-comb. The plants also were tested for TSWV and INSV by double-antibody (DAS)-ELISA (Agdia Inc.) with the same results. Total RNA was extracted from one symptomatic orchid leaf from each nursery. Reverse transcription (RT)-PCR was performed with the universal tospovirus primer set BR60and BR65 (1). PCR bands of the expected size were amplified from each leaf. PCR products were sequenced directly. The orchid leaf that tested positive for TSWV by ELISA produced a 495-bp sequence with 97% identity to several isolates of the TSWV nucleocapsid protein gene listed in GenBank (Accession Nos. AY744479, AY8770391, DQ376185, and AF02659). The orchid leaf that tested positive for INSV by ELISA produced a 396-bp sequence with 98 to 99% identity to several isolates of the INSV nucleocapsid protein gene (Accession Nos. D00914, DQ425096, X66972, and AD109100). Although these viruses have been reported a few times in orchids previously (2,3), to our knowledge, this is the first time they have been reported in this host in Florida. In addition, white Phalaenopsis spp. appears to be a local lesion host and not a systemic host for these viruses.

Investigations of different strategies for high frequency regeneration of Dendrobium malones ‘Victory’

Regeneration of orchid, Dendrobium malones “Victory”, has been established by direct differentiation and through callus formation. With direct thin leaf section differentiation method, the number of protocorm like bodies (PLBs) formed from Thin Section (TS) explants obtained from a single leaf was 10 times more than that from a single whole leaf, in a short time period. This therefore suggested that the proliferation potential is distributed all along the seedling leaves and morphogenically competent cells were not limited in the basal ends or the tips of the leaves in TS of orchid leaf cultures. Full MS media supplemented with auxins/cytokinins either in single or in combination along with peptone, yeast extract, casein hydrolysate (each at 100 mg/l), banana powder (40 g/l), 15% coconut water and 2% sucrose have shown excellent results for high frequency regeneration in orchid cultures. In many higher plants, regenerating leaves responded by developing proliferative loci in some “Predetermined Regenerative” cells in the dermal layers by tissue culture methods.