Asymbiotic seed germination and in vitro seedling development of Orchis militaris, an endangered orchid in Siberia

Aleksandra Yurievna Nabieva1
1Department of Biotechnology, Central Siberian Botanical Garden SB RAS, Novosibirsk, Russia

Tóm tắt

Terrestrial orchids belonging to the Orchis genus are difficult to propagate and are under great pressure in their natural habitats. Studies regarding the influence of photoperiod and temperature regimes on Orchis militaris germination and morphological changes during immature seed development in vitro are scarce. Our aim was to identify photoperiod, temperature, and different nutrient media requirements for optimization of O. militaris seed germination and vigorous seedling production. Post-germination morphological changes were recorded with O. militaris seeds collected from 32-day-old fruits, where the percentage of O. militaris seeds without embryo was 38.4%. The highest rate of O. militaris seed germination (82.6%) was obtained on Malmgren modified terrestrial orchid medium (mM), enriched by 5% coconut water, 5% birch sap, and 0.1% AC. Nine percent of seedlings were able to reach the advanced seedling stage (stage 6) after 12 months of maintenance on this medium. In all 3 modified media (Harvais, Knudson С and Malmgren), regeneration was via the production of protocorms and seedlings without callus formation. It was proved that more abundantly vigorous protocorms were formed on the modified Harvais 2 under continuous darkness, while the subculture in Knudson C medium with AC addition could be necessary to stimulate their further development. The regeneration success of the species for in vitro conditions was increased by following its natural seasonal cycle. This study demonstrated an efficient micropropagation system for O. militaris using immature seeds and thus widely opened the perspectives for its conservation in nature. The favorable conditions of seed germination periods for in vitro culture, identified as the definite shift of temperatures and photoperiod regimes intrinsic to the species in nature, could improve seedling survival of this medicinally important orchid.

Tài liệu tham khảo

Kretzschmar H, Eccarius W, Dietrich H (2007) The orchid genera Anacamptis. Phylogeny, taxonomy, morphology, biology, distribution, ecology and hybridization. Echinomedia, Bürgel, Orchis and Neotinea Govaerts R, Bernet P, Kratochvil K, Gerlach G, Carr G, Alrich P, Pridgeon AM, Pfahl J, Campacci MA, Baptista HD, Tigges H, Shaw J, Cribb PJ, George A, Kreuz K, Wood J (2016) World Checklist of Orchidaceae. Royal Botanic Gardens, Kew, London. Available at http://apps.kew.org/wcsp/ Kreziou A, De Boer H, Gravendeel B (2016) Harvesting of salep orchids in north-western Greece continues to threaten natural populations. Oryx 50(3):393–396. https://doi.org/10.1017/S0030605315000265 Jagdale SP, Shimpi S, Chachad D (2009) Pharmacological studies of 'Salep'. J. Herb Med Toxic 3(1):153–156 Ghorbani A, Gravendeel B, Naghibi F, de Boer HJ (2014) Wild orchid tuber collection in Iran: a wake-up call for conservation. Biodivers Conserv 23: 2749–2760. Seaton P, Ramsay M (2005) Growing orchids from seed. Royal Botanic Gardens, Kew, London Jakobsone G (2008) Morphogenesis of wild orchid Dactylorhiza fuchsii in tissue culture. Acta Univer Latv, Biology 745:17–23 Valletta A, Attorre F, Bruno F, Pasqua G (2008) In vitro asymbiotic germination of Orchis mascula L. Plant Biosyst 142(3):65655. https://doi.org/10.1080/11263500802411205 Bektas E, Cuce M, Sokmen A (2013) In vitro germination, protocorm formation, and plantlet development of Orchis coriophora (Orchidaceae), a naturally growing orchid species in Turkey. Turk J Bot 37(2):336–342 Farrell L (1985) Biological flora of the British Isles: Orchis militaris L. J Ecol 73:1041–1053 Evans A, Janssens S, Jacquemyn H (2020) Impact of climate change on the distribution of four closely related Orchis (Orchidaceae) species. Diversity 12(8), 312. https://doi.org/10.3390/d12080312. Averyanov LV (2008) Orchis militaris L. In: Bardunov LV, Novikov VS (eds) The Red Data Book of the Russian Federation (Plants and Fungi). KMK Scientific Press Ltd., Moscow Calevo J, Voyron S, Ercole E, Girlanda M (2020) Is the distribution of two rare Orchis sister species limited by their main mycobiont? Diversity 12(262). https://doi.org/10.3390/d12070262 Fardeeva MB (2007) Ontogeny of Orchis militaris L. In: Ontogenetic Atlas of Plants. 5:238–241. Mari. Gos. Univ, Yoshkar Ola. Tremblay RL, Ackerman JD, Zimmerman JK, Calvo RN (2005) Variation in sexual reproduction in orchids and its evolutionary consequences: a spasmodic journey to diversification. Biol J Linn Soc 84:1–54. https://doi.org/10.1111/j.1095-8312.2004.00400.x Henneresse T, Wesselingh RA, Tyteca D (2017) Effects of floral display, conspecific density and rewarding species on fruit set in the deceptive orchid Orchis militaris (Orchidaceae). Plant Ecol. Evol 150:279–292. doi.org/https://doi.org/10.5091/plecevo.2017.1313. Vakhrameeva MG, Varlygina TI, Tatarenko IV (2014) Orchids of Russia (biology, ecology and protection). KMK Scientific Press, Moscow Pierce S, Cerabolini BEL (2011) Asymbiotic germination of the White Mountain Orchid (Pseudorchis albida) from immature seed on media enriched with complex organics or phytohormones. Seed Sci Technol 39(1):199–203. https://doi.org/10.15258/sst.2011.39.1.17 Kendon JP, Rajaovelona L, Sandford H, Fang R, Bell J, Sarasan V (2017) Collecting near mature and immature orchid seeds for ex situ conservation: 'in vitro collecting' as a case study. Bot Stud. 58(1):34. https://doi.org/10.1186/s40529-017-0187-5 Krasnoborov IM (2000) Determinant of plants of the Novosibirsk region. Nauka, Novosibirsk Harvais G (1973) Growth requirements and development of Cypripedium reginae in axenic culture. Canad J Bot 51(2):327–332 Swarts ND, Dixon KW (2017) Conservation methods for terrestrial orchids. J. Ross Publishing, United States Knudson L (1946) A nutrient for germination of orchid seeds. Am Orchid Soc Bull 15:214–217 Malmgren S (1996) Orchid propagation: theory and practice. In: Allen С (ed) Proceedings of North American native terrestrial orchid conference, Maryland. Rasmussen HN (1995) Terrestrial orchids: From seed to mycotrophic plant. Cambridge University Press, Cambridge Malmgren S, Nyström H (2011) Orchid Propagation. http://www.lidaforsgarden.com/Orchids/engelsk.htm Kulikov PV, Phillippov EG (1998) On propagation of temperate zone orchids by tissue culture methods. Bulletin of the Main botanical garden of Russian Academy of Science 176:125–131 Metsare M, Ilves A, Haldna H, Kull T, Tali K (2015) Four seed-quality measures in orchids with different pollination systems. Acta Bot Gallica 162(4):263–269. https://doi.org/10.1080/12538078.2015.1100549 Ye Z-M, Dai W-K, Jin X-F, Gituru RW, Wang Q-F, Yang C-F (2014) Competition and facilitation among plants for pollination: can pollinator abundance shift the plant-plant interactions? Plant Ecol 215:3–13. https://doi.org/10.1007/s11258-013-0274-y Yeung EC (2017) A perspective on orchid seed and protocorm development. Bot Stud. 58(1):33. https://doi.org/10.1186/s40529-017-0188-4 Hürkan YK, Hürkan K, Aki C (2018) Comparative growth media performances on in vitro propagation of some salep orchids. Anadolu University Journal of Science and Technology - C Life Sciences and Biotechnology 7(1):52–62 Baskin CC, Thompson K, Baskin JM (2006) Mistakes in germination ecology and how to avoid them. Seed Sci Res 16:165–168 Delforge P (2006) Orchids of Europe, North Africa and the Middle East, 3rd edn. A & C Black, London Calevo J, Bazzicalupo M (2020) Less is more: low-cost in vitro . Nat Conserv Res 5(1):172–177. https://doi.org/10.24189/ncr.2020.043 Arditti J. (ed) (1981). Orchid Biology. Reviews and perspectives. Orchid seed germination and seedling culture, vol 11. Cornell University Press, Ithaca, New York Teixeira da Silva JA (2013) Orchids: Advances in tissue culture, genetics, phytochemistry and transgenic biotechnology. Floricult Ornam Biotech 7:1–52 Yamazaki J, Miyoshi K (2006) In vitro asymbiotic germination of immature seed and formation of protocorm by Cephalanthera falcata (Orchidaceae). Ann Bot 98(6):1197–1206. https://doi.org/10.1093/aob/mcl223 Yeung EC, Li YY, Lee YI (2018) Understanding seed and protocorm development in orchids. In: Lee YY, Yeung EC (eds) Orchid Propagation: from laboratories to greenhouses – methods and protocols. Springer Protocols Handbooks, New York, pp 1–24 Huh YS, Lee JK, Nam SY, Paek KY, Suh GU (2016) Improvement of asymbiotic seed germination and seedling development of Cypripedium macranthos Sw. with organic additives. J Plant Biotechnol 43(1):138–145 Burch RM, Chambers CB (2013). Micropropagation and establishment of Cypripedium 'Lady slipper' orchids. Acta Hortic 988:161-166 https://doi.org/10.17660/ActaHortic.2013.988.18 Tawaro S, Suraninpong P, Chanprame S (2008) Germination and regeneration of Cymbidium findlay sonianum Lindl. on a medium supplemented with some organic sources. Walailak J Sci Tech 5:125–135 Tan S, Yong J, Ge L (2014) Analyses of phytohormones in coconut (Cocos nucifera L.) water using capillary electrophoresis-tandem mass spectrometry. Chromatography 1(4):211–226 Kefeli V, Kalevitch MV (2003) Natural growth inhibitors and phytohormones in plants and environment. In: Borsari B (ed) Kluwer Academic Publishers, Dordrech, p.324. Molnar Z, Virag E, Ordog E (2011) Natural substances in tissue culture media of higher plants. Acta Biol Szegedvol 55(1):123–127 Pakum W, Watthana S, Srimuang KO, Kongbangkerd A (2016) Influence of medium component on in vitro propagation of Thai’s endangered orchid: Bulbophyllum nipondii Seidenf. Plant Tissue Cult Biotechnol 26(1):37–46. Zhang Y, Lee YI, Deng L, Zhao S (2013) Asymbiotic germination of immature seeds and seedling development of Cypripedium macranthos Sw., an endangered Lady’s slipper orchid. Sci Hortic 164:130–136 Novak SD, Luna LJ, Gamage RN (2014) Role of auxin in orchid development. Plant Signal Behav 9(10):e972277. https://doi.org/10.4161/psb.32169 Ponert J, Vosolsobě S, Kmecová K, Lipavská H (2011) European orchid cultivation – from seed to mature plant. Eur J Environ Sci 1(2):95–107 De KK, Majumdar S, Sharma R, Sharma B (2006) Green pod culture and rapid micropropagation of Dendrobium chrysanthum Wall. – a horticultural and medicinal orchid. Folia Hortic 18:81–90 Godo T, Komori M, Nakaoki E, Yukawa T, Miyoshi K (2010) Germination of mature seeds of Calanthe tricarinata Lindl., an endangered terrestrial orchid, by asymbiotic culture in vitro. In Vitro Cell Dev Biol - Plant 46:323–328 Baker A, Kaviani B, Nematzadeh G, Negahdar N (2014) Micropropagation of Orchis catasetum – a rare and endangered orchid. Acta Sci Pol, Hortorum Cultus 13:197–205 Bektas E, Sokmen A (2016) In vitro seed germination, plantlet growth, tuberization, and synthetic seed production of Serapias vomeracea (Burm.f.) Briq. Turk J Bot 40:584–594 https://doi.org/10.3906/bot-1512-13 Novotna KW, Vejsadova H, Kindlmann P (2007) Effects of sugars and growth regulators on in vitro growth of Dactylorhiza species. Biol Plantarum 51:198–200 Gutierrez-Sanchez A, Monriboc-Villanueva JL, Cocotle-Ronzón Y, Martinez-Cruz NS, Guerrero-Analco JA (2020) Phenolic profile and antioxidant activity from wild and in vitro cultivated Rhynchostele rossii (Orchidaceae). Acta Bot Mex 127(127), e1665. https://doi.org/10.21829/abm127.2020.1665 Vudala SM, Ribas LLF (2017) Seed storage and asymbiotic germination of Hadrolaelia grandis (Orchidaceae). S Afr J Bot 108:1–7. https://doi.org/10.1016/j.sajb.2016.09.008 Merritt DJ, Hay FR, Swarts ND, Sommerville KD, Dixon KW (2014) Ex situ conservation and cryopreservation of orchid germplasm. Int J Plant Sci 175(1):46–58 Vujanovic V, St-Arnaud M, Barabe D, Thibeault G (2000) Viability testing of orchid seed and the promotion of colouration and germination. Ann Bot 86(1):79–86