Oncidium Orchids Research Articles

Advances in the Improvement of Oncidium History and Perspectives

Oncidium orchids, commonly known as “dancing lady orchids,” represent a diverse genus with significant horticultural and commercial value. Breeding programs aimed at improving Oncidium species and hybrids have developed over the past few decades, driven by their ornamental characteristics such as flower colour, size, form, fragrance and disease resistance. This review explores the traditional and modern approaches used in Oncidium orchid breeding, including classical hybridisation, mutagenesis, polyploidy and some biotechnological advancements such as tissue culture and molecular markers. Furthermore, the challenges and potential future directions in breeding Oncidium orchids, including the development of novel traits are discussed.

Environmental biosafety assessment on transgenic Oncidium orchid modified by RNA interference of Phytoene Synthase genes.

Hybrid Oncidium orchids, such as Oncidium Gower Ramsey and Oncidium "Honey Angel," are popular cut flowers in Japan and Taiwan. Due to pollen sterility, no new varieties have been created by conventional breeding methods. Recently, we employed RNA interference (RNAi) technology to suppress phytoene synthase and successfully modified floret hue from yellow to white (Liu et al. 2019). Transgenic white Oncidium orchids, Honey Snow MF-1, have been grown to test their genetic stability, and their environmental biosafety was assessed for approximately one year under government regulatory instructions from the Council of Agriculture, Taiwan. In the present study, pollen sterility was demonstrated by cytological observation of the microsporogenesis step, pollen morphology abortion, and failure of pollen germination. Assays on allelopathic effect on the other plants and the soil rhizospheric microbial flora-revealed that transgenic Oncidium orchids are potentially safe with regard to environmental biodiversity. Therefore, the general release permissions have been granted and an application for licensing for commercial production is under way.

A direct gene transferring system for Oncidium orchids, a difficult crop for genetic transformation

Oncidium orchids are susceptible to damage through viral infection caused by the Cymbidium mosaic virus (CymMV). Generating a virus-resistant plant is a major challenge. Unfortunately, gene transformation in Oncidium orchids is not easy. This study developed selection steps for a direct gene transferring system through particle bombardment. Optimized hygromycin screening was reported in the transformed protocorm-like bodies (PLBs) of two commercial Oncidium hybrids (Onc. Gower Ramsey and Onc. Sweet Sugar) with the RNAi construction of the coat protein gene of CymMV. The transgenic Onc. Sweet Sugar PLBs were regenerated on a modified Murashige and Skoog medium containing hygromycin. The effective selection system in exterminating non-transformed PLBs and the functional transgene in the transgenic lines was analyzed using reverse transcription polymerase chain reaction and quantitative polymerase chain reaction. The application of this study may help the gene transformation system of other orchid plant species.

RAPID MICROPROPAGATION OF VU NU ORCHID (ONCIDIUM SP.) BY USING TISSUE CULTURE TECHNIQUE

: The high demand of Oncidium orchids leads us to find out efficient methods of propagating them. However, the propagation rate of traditional methods are low in nature and a hybrid seed is not genetically stable. Thus, plant cell biotechnology is examined as the most effective way to resolve the barrier of elite clone production. Shoot clusters were cultured on MS medium supplemented with 2,4-D (1 mg/l) for callus induction (76.19%) before induced callus was favoured for PLBs regeneration (98 PLBs/callus cluster) on MS medium supplemented with NAA (0.75 mg/l); the combination of BA (0.5 mg/l) and NAA (0.5 mg/l) was favoured for PLBs regeneration (28.18 PLBs/shoot cluster) from shoots cultivation. The PLBs (79.21 PLBs/PLB cluster) were then proliferated on MS medium supplemented with NAA (1 mg/l) and BA (1 mg/l) for shoot regeneration (12.42 shoots/PLBs cluster). Multiple-shoots were divided to 3-4 shoots/cluster for micropropagation on the MS medium supplemented with the combination of BA (0.25 mg/l) and NAA (0.25 mg/l) to reach 11.66 shoots/cluster. Shoots were finally separated to single-shoot for rooting on the MS medium supplemented with NAA (0.75 mg/l). A scheme for Oncidium micropropagation using PLBs culture techniques was set up.

The C-Terminal Sequence and PI motif of the Orchid (OncidiumGower Ramsey)PISTILLATA(PI) Ortholog Determine its Ability to Bind AP3 Orthologs and Enter the Nucleus to Regulate Downstream Genes Controlling Petal and Stamen Formation

This study focused on the investigation of the effects of the PI motif and C-terminus of the Oncidium Gower Ramsey MADS box gene 8 (OMADS8), a PISTILLATA (PI) ortholog, on floral organ formation. 35S::OMADS8 completely rescued and 35S::OMADS8-PI (with the PI motif deleted) partially rescued petal/stamen formation, whereas these deficiencies were not rescued by 35S::OMADS8-C (C-terminal 29 amino acids deleted) in pi-1 mutants. OMADS8 could interact with Arabidopsis APETALA3 (AP3) and enter the nucleus. The nuclear entry efficiency was reduced for OMADS8-PI/AP3 and OMADS8-C/AP3. OMADS8 could also interact with OMADS5/OMADS9 (the Oncidium AP3 ortholog) and enter the nucleus with an efficiency only slightly affected by the deletion of the C-terminal sequence or PI motif. However, the stability of the OMADS8/OMADS5 and OMADS8/OMADS9 complexes was significantly reduced by deletion of the C-terminal sequence or PI motif. Further analysis indicated that the expression of genes downstream of AP3/PI (BNQ1/BNQ2/GNC/At4g30270) was compensated by 35S::OMADS8 and 35S::OMADS8-PI to a level similar to wild-type plants but was not affected by 35S::OMADS8-C in the pi-1 mutants. A similar FRET (fluorescence resonance energy transfer) efficiency was observed for Arabidopsis AGAMOUS (AG) and the Oncidium AG ortholog OMADS4 for OMADS8, OMADS8-PI and OMADS8-C. These results indicated that the OMADS8 PI motif and C-terminus were valuable for the interaction of OMADS8 with the AP3 orthologs to form higher order heterotetrameric complexes that regulated petal/stamen formation in both Oncidium orchids and transgenic Arabidopsis. However, the C-terminal sequence and PI motif were dispensable for the interaction of OMADS8 with the AG orthologs.

Pectobacterium chrysanthemi as the dominant causal agent of bacterial soft rot in Oncidium “Grower Ramsey”

Oncidium orchids are important ornamental flowers for export in Taiwan, but their production is limited by bacterial soft rot disease. The common pathogens of bacterial soft rot include Pectobacterium carotovorum subsp. carotovorum (Pcc) and P. chrysanthemi (Pch). Pch is a unique pathogen that can be isolated from Oncidium. Thus, the possible reasons for that are need to be studied. In this study, we sought to determine the possible factors affecting the pathogenicity of Pcc and Pch in Oncidium Gower Ramsey. Results showed that both Pcc and Pch are pathogenic to Oncidium orchid and two other common hosts of soft rot bacteria, potato and radish. However, Pch is more virulent in Oncidium orchid than Pcc. Moreover, temperature is not the dominant factor contributing to differences in soft rot development in Oncidium caused by Pcc and Pch, suggesting that host selection is more important than temperature. We further found that Pcc growth is not inhibited by Oncidium leaf extract and that the inhibitory effect of Oncidium on Pcc may be attributed to the defense response of living cells. In addition, stronger reactive oxygen species accumulation was triggered by Pcc rather than Pch. This defense response against Pcc is related to plant immune signal pathways, including calcium influx and NADPH oxidase activation. Through this study, we have provided evidences that Pch, not Pcc, is the dominant soft rot pathogen in Oncidium. The immune response triggered by bacterial recognition explains the inhibition of Pcc growth in Oncidium Gower Ramsey.

Incidence of Cymbidium mosaic virus and Odontoglossum ringspot virus affecting Oncidium orchids in Hainan Island, China

Oncidium is the most popular cut flower for the orchid industry in Hainan Island and several viruses have been reported to negatively impact their growth and yield. A survey of Cymbidium mosaic virus (CymMV) and Odontoglossum ringspot virus (ORSV) in Oncidium orchids cultivated in Hainan Island was conducted during 2009–2012 with indirect enzyme-linked immunosorbent assay (ELISA). A total of 171,585 samples from adult plant, tissue-culture flask, seedling, and medium seedling were tested for CymMV and ORSV. Of the 24,125 adult plant samples from eighteen commercial orchid farms, 77.58% were infected with CymMV, 16.04% with ORSV, and 10.19% with both CymMV and ORSV. Of the 67,080 samples cultivated in tissue-culture flasks from eight tissue culture laboratories, 41.18% were infected with CymMV, 6.11% with ORSV, and 4.46% with both CymMV and ORSV. Of the 54,450 seedling samples from eight seedling nurseries, 53.20% were infected with CymMV, 9.00% with ORSV, and 5.23% with both CymMV and ORSV. And of the other 25,930 medium seedling samples from twelve medium seedling nurseries, 44.96% were infected with CymMV, 7.90% with ORSV, and 5.15% with both CymMV and ORSV. Plant material must be tested for the presence of viruses before being used for mass production by tissue culture, and sanitation during propagation of orchid plants and harvesting of flowers is essential in order to prevent the virus from spreading. These findings emphasize the need to implement a virus-free certification programme that would greatly enhance the management of these two prevalent viruses in the Hainan orchid industry.

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.

Misting cooling technique for protected culture of Oncidium orchids in subtropical regions

Oncidium are planted in open protected culture or under shading nets in subtropical regions. Because of the lack of cooling techniques for the inside microclimates, the production period is limited, and the quantity and quality of flowers is unstable. Without ventilating, high humidity and temperature induces severe growth problems. In this study, mechanical ventilation fans with intermittent misting were applied to modify the internal microclimate of the Oncidium greenhouse. Nine experimental setting of misting intervals were tested. The misting efficiency is defined and served as the performance index. The misting intervals of 3 min on to 3 min off or 5 min on to 5 min off had the higher efficiency. The cooling technique can keep the interior air temperature lower than that of outside air temperature. The temperature of plants could be maintained close to the greenhouse air temperature. The intermittent misting function with the best evaporative cooling efficiency was controlled with interval operation. This technique can be applied to other protected structures of Oncidium and other orchids. Key words : Shading nets, misting cooling, tunnel greenhouse, Oncidium .

Characterization of Oncidium ‘Gower Ramsey’ Transcriptomes using 454 GS-FLX Pyrosequencing and Their Application to the Identification of Genes Associated with Flowering Time

Oncidium 'Gower Ramsey' is a valuable and successful commercial orchid for the floriculture industry in Taiwan. However, no genome reference for entire sequences of the transcribed genes currently exists for Oncidium orchids, to facilitate the development of molecular biological studies and the breeding of these orchids. In this study, we generated Oncidium cDNA libraries for six different organs: leaves, pseudobulbs, young inflorescences, inflorescences, flower buds and mature flowers. We utilized 454-pyrosequencing technology to perform high-throughput deep sequencing of the Oncidium transcriptome, yielding >0.9 million reads with an average length of 328 bp, for a total of 301 million bases. De novo assembly of the sequences yielded 50,908 contig sequences with an average length of 493 bp from 796,463 reads and 120,219 singletons. The assembled sequences were annotated using BLAST, and a total of 12,757 and 13,931 unigene transcripts from the Arabidopsis and rice genomes were matched by TBLASTX, respectively. A Gene Ontology (GO) analysis of the annotated Oncidium contigs revealed that the majority of sequenced genes were associated with 'unknown molecular function', 'cellular process' and 'intracellular components'. Furthermore, a complete flowering-associated expressed sequence that included most of the genes in the photoperiod pathway and the 15 CONSTANS-LIKE (COL) homologs with the conserved CCT domain was obtained in this collection. These data revealed that the Oncidium expressed sequence tag (EST) database generated in this study has sufficient coverage to be used as a tool to investigate the flowering pathway and various other biological pathways in orchids. An OncidiumOrchidGenomeBase (OOGB) website has been constructed and is publicly available online (http://predictor.nchu.edu.tw/oogb/).

Bacterial Soft Rot of Oncidium Orchids Caused by a Dickeya sp. (Pectobacterium chrysanthemi) in Florida.

Oncidium orchids have been subjected to extensive cultivation in the pot-plant and cut flower industries because of their attractive and numerous flowers. In August 2008, approximately 50 Oncidium 'Gower Ramsey' orchids were discovered at a commercial orchid nursery in South Florida with brown, macerated leaves typical of soft rot disease reported in other orchids. Ten plants were selected, and sections were removed from the edge of symptomatic tissue and bacteria were isolated according to the method described by Schaad et al. (3). All isolates were gram negative, anaerobic, degraded pectate, grew at 37°C, produced blue-to-brown pigment on nutrient agar-glycerol-manganese chloride (NGM) medium (1), were sensitive to erythromycin, oxidase negative, and positive for phosphatase and indole production. Further analyses were performed on four of the isolates. MIDI analysis (Sherlock version TSBA 4.10; Microbial Identification, Newark, DE) identified the isolates as Erwinia chrysanthemi (SIM 0.880 to 0.929). Polymerase chain reactions were performed with the 16S primers 27f and 1495r (4) and 1,423 bp of the 16S rDNA gene showed 98 to 99% sequence identity to Pectobacterium chrysanthemi (GenBank Accession No. FM946179). Sequences were deposited in GenBank (Nos. HQ287572-HQ287575). Pathogenicity tests were performed by injecting 10 Oncidium 'Gower Ramsey' orchids with 100 μl of a bacterial suspension at 1 × 108 CFU/ml. Ten plants were inoculated with 100 μl of sterile water as controls. Plants were placed in a greenhouse at 26.0°C to 30.0°C and 50 to 83% relative humidity. Soft rot symptoms were observed on all inoculated plants within 24 h while control plants appeared normal. A Dickeya sp. was reisolated and identified according to the method described above. Oncidium orchids are known to be highly susceptible to P. carotovora (= E. carotovora) and soft rot caused by P. carotovora is known to occur frequently on Oncidium orchids (2). Although, an Erwinia sp. has been reported to cause soft rot symptoms on Oncidium aureum, to our knowledge, this is the first report of a Dickeya sp. (= P. chrysanthemi) causing soft rot symptoms on Oncidium orchids grown in large-scale commercial production in the United States.

Hydrogen peroxide mediates the expression of ascorbate-related genes in response to methanol stimulation in Oncidium

We investigated the signaling role of hydrogen peroxide (H 2O 2) in regulating the ascorbate (AsA) level after exogenous methanol (MeOH) application. The endogenous H 2O 2 and AsA levels as well as the expression of related genes were monitored after MeOH treatment of cultures of Oncidium protocorm-like bodies (PLB). A high MeOH concentration was deleterious and caused irreversible consumption of endogenous AsA. However, a low MeOH concentration (50 mM) triggered the synthesis of H 2O 2 and was effective in enhancing the expression of AsA-biosynthetic genes of the Smirnoff–Wheeler and galacturonate (GalUA) pathways. The increased expression of these genes could be blocked by the addition of hydroxylamine, an inhibitor of alcohol oxidase (EC: 1.1.3.13), and diphenyleneiodonium chloride (DPI), an inhibitor of NADPH oxidase (EC: 1.6.3.1). Thus, the H 2O 2 generated by MeOH application is a product of MeOH detoxification through alcohol oxidase and NADPH oxidase activation. In this chain, H 2O 2 acts as a secondary messenger for the activation of AsA-related genes. Our results reveal the signaling function of H 2O 2 and cellular AsA homeostasis in Oncidium orchids in response to MeOH stimulation. A mechanism for the MeOH effect on AsA production is suggested.

Potential of invasive and native solitary specialist bee pollinators to help restore the rare cowhorn orchid ( Cyrtopodium punctatum) in Florida

Cyrtopodium punctatum is a rare epiphytic orchid in southern Florida, made rare by historical over-collection. We examined the potential pollination of this orchid by the recently naturalized orchid bee ( Euglossa viridissima), recorded as a pollinator of the orchid in tropical America, and found that this orchid bee is not a pollinator of the plant. We sought to learn what is responsible for relatively heavy fruit set in a Fairchild Tropical Botanic Garden population of C. punctatum, and determined that the native oil-collecting bee, Centris errans, is the most important pollinator. C. punctatum flowers at Fairchild have 18 times the fruit set of flowers in Everglades National Park. The difference is probably due to the many species and individuals of oil-reward plants in the Malpighiaceae in the garden, compared to one uncommon native species in the park. Female C. errans visit these oil-reward flowers to obtain edible oils to provision their brood. Cyrtopodium flowers appear to mimic the oil-reward flowers of the Malpighiaceae to attract Centris bee pollinators, much as many Oncidium orchids do in tropical America. We recommend that Brysonima lucida, a rare native malpighiaceous shrub, and C. punctatum be planted together in Everglades National Park and other natural areas to attempt to increase C. errans pollination, to restore and enhance the long-term survival of the orchid. Planting model malpighiaceous plants to enhance Centris bee pollination may be a useful restoration tactic for other rare orchid mimics in the American tropics, including Atlantic Forest in Brazil.

Contact dermatitis due to orchids (Cymbidium and Oncidium)

A 64-year-old male who cultivated orchids as a hobby, had noticed itchy erythematous lesions on both hands since 6 months earlier. His eruptions had gradually worsened and scales and fissures had appeared. Although he was treated with topical corticosteroids by a doctor, his erythema showed no improvement. Patch tests with the orchids he cultivated, fertilizers, antiseptics and insecticides showed positive reactions to the stems of Cymbidium and Oncidium orchids. A diagnosis of contact dermatitis attributable to the cultivated orchids was made.