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Highly compatible Epa-01 strain promotes seed germination and protocorm development of Papilionanthe teres (Orchidaceae)

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Abstract

Symbiotic seed germination serves as a preferred method for orchid multiplication related to conservation and reintroduction programs, which involves isolation, identification, and germination-enhancing testing of symbiotic fungi. This study uses seeds of Papilionanthe teres, a locally endangered and medicinally valuable epiphytic orchid, to attract germination-enhancing fungi on its four host plants. Only one common and highly compatible fungus (Epa-01 strain, Epulorhiza sp.), isolated from seed baiting near three host plants (Averrhoa carambola, Lagerstroemia villosa, Callistemon rigidus), enhanced seed germination by more than 80 % and promoted protocorm development to reach stage 5 (with two leaves). Seeds cocultured with the Epa-01 strain and oat meal agar medium significantly outperformed in germination and growth speed compared with those cocultured with asymbiotic germination medium only, indicating that symbiotic seed germination is an effective method for P. teres seedling production. Bark substrate types have profound effects on symbiotic seed germination and protocorm development possibly due to different abundance and growth vitality of the Epa-01 strain on the four host plants. A significant difference was found in the developmental speed of symbiotic seeds between A. carambola and the other three host plants under ex situ and ex vitro seed germination treatments (all P < 0.05). The results suggest that in situ seed baiting may be used to effectively capture germination-enhancing fungi in epiphytic orchids, and testing the effects of bark substrate types on seed germination and protocorm development contributes to selecting appropriate host plants for its reintroduction into natural habitats.

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References

  • Adhikari YP, Fischer A, Fischer HS (2012) Micro-site conditions of epiphytic orchids in a human impact gradient in Kathmandu valley. J Mt Sci 9(3):31–342

    Article  Google Scholar 

  • Arditti J (1992) Fundamentals of orchid biology. Wiley, New York

    Google Scholar 

  • Athipunyakom P, Manoch L, Piluek C (2004) Isolation and identification of mycorrhizal fungi from eleven terrestrial orchids. Nat Sci 38:216–228

    Google Scholar 

  • Batty AL, Dixon KW, Brundrett M, Sivasithamparam K (2001) Constraints to symbiotic germination of terrestrial orchid seed in a mediterranean bushland. New Phytol 152(3):511–520

    Article  Google Scholar 

  • Batty AL, Dixon KW, Brundrett MC, Sivasithamparam K (2002) Orchid conservation and mycorrhizal associations. In: Sivasithamparam K, Dixon KW, Barrett RL (eds) Microorganisms in plant conservation and biodiversity. Kluwer Academic Publishers, Dordrecht, pp 195–226

    Google Scholar 

  • Benzing DH (1981) Bark surfaces and the origin and maintenance of diversity among angiosperm epiphytes: a hypothesis. Selbyana 5:248–255

    Google Scholar 

  • Bidartondo MI, Read DJ (2008) Fungal specificity bottlenecks during orchid germination and development. Mol Ecol 17:3707–3716

    PubMed  Google Scholar 

  • Bonnardeaux Y, Brundrett M, Batty A, Dixon K, Koch J, Sivasithamparam K (2007) Diversity of mycorrhizal fungi of terrestrial orchids: compatibility webs, brief encounters, lasting relationships and alien invasions. Mycol Res 111:51–61

    Article  PubMed  Google Scholar 

  • Brundrett MC, Scade A, Batty AL, Dixon KW, Sivasithamparam K (2003) Development of in situ and ex situ seed baiting techniques to detect mycorrhizal fungi from terrestrial orchid habitats. Mycol Res 107:1210–1220

    Article  PubMed  Google Scholar 

  • Callaway R, Reinhart K, Moore G, Moore D, Pennings S (2002) Epiphyte host preferences and host traits: mechanisms for species-specific interactions. Oecologia 132:221–230

    Article  Google Scholar 

  • Chen XQ (1999) Chinese Flora (volume 18). In: Lang KY, Chen XQ, Luo YB, Zhu GH (eds) Orchidaceae. Science Press, Beijing, pp 1–12

  • Chen W, Li Z, Li K (2007) Tissue culture and rapid propagation of Papilionanthe teres (Roxb.) Schltr. Plant Physiol Commun 43(5):882

    Google Scholar 

  • Chen XQ, Liu ZJ, Zhu GH, Lang KY, Ji ZH, Luo YB, Jin XH, Cribb PJ, Wood JJ, Gale SW, Ormerod P, Vermeulen JJ, Wood HP, Clayton D, Bell A (2009) Flora of China: Orchidaceae. In: Chen XQ, Wood JJ (eds) 175. PAPILIONANTHE Schlechter, Orchis 9: 78. 1915. Science Press, Beijing, Orchis, pp 477

    Google Scholar 

  • Chen J, Wang H, Guo SX (2012) Isolation and identification of endophytic and mycorrhizal fungi from seeds and roots of Dendrobium (Orchidaceae). Mycorrhiza 22:297–307

    Article  PubMed  Google Scholar 

  • Chutima R, Dell B, Lumyong S (2011) Effects of mycorrhizal fungi on symbiotic seed germination of Pecteilis susannae (L.) Rafin (Orchidaceae), a terrestrial orchid in Thailand. Symbiosis 53:149–156

    Article  CAS  Google Scholar 

  • Clark K, Nadkarni N, Gholz H (1998) Growth, net production, litter decomposition, and net nitrogen accumulation by epiphytic bryophytes in a tropical montane forest. Biotropica 30:12–23

    Article  Google Scholar 

  • Crain B (2012) On the relationship between bryophyte cover and the distribution of Lepanthes spp. Lankesteriana 12(1):13–18

    Article  Google Scholar 

  • Cruz-Higareda JB, Luna-Rosales BS, Barba-Álvarez A (2015) A novel seed baiting technique for the epiphytic orchid Rhynchostele cervantesii, a means to acquire mycorrhizal fungi from protocorms. Lankesteriana 15(1):67–76

    Article  Google Scholar 

  • Diez JM (2007) Hierarchical patterns of symbiotic orchid germination linked to adult proximity and environmental gradients. J Ecol 95(1):159–170

    Article  Google Scholar 

  • Fracchia S, Silvani V, Flachsland E, Terada G, Sede S (2013) Symbiotic seed germination and protocorm development of Aa achalensis Schltr., a terrestrial orchid endemic from Argentina. Mycorrhiza 24:35–43

    Google Scholar 

  • Frei JK, Dodson CH (1972) The chemical effect of certain bark substrates on the germination and early growth of epiphytic orchids. Bull Torrey Bot Club 99:301–307

    Article  Google Scholar 

  • Gowland KM, van der Merwe MM, Linde CC, Clements MA, Nicotra AB (2013) The host bias of three epiphytic Aeridinae orchid species is reflected, but not explained, by mycorrhizal fungal associations. Am J Bot 100(4):764–777

    Article  PubMed  Google Scholar 

  • Illyés Z, Halász K, Rudnóy S, Ouanphanivanh N, Garay T, Bratek Z (2009) Changes in the diversity of the mycorrhizal fungi of orchids as a function of the water supply of the habitat. J Appl Bot Food Qual 83:28–36

    Google Scholar 

  • Johnson TR, Stewart SL, Dutra D, Kane ME, Richardson L (2007) Asymbiotic and symbiotic seed germination of Eulophia alta (Orchidaceae)-preliminary evidence for the symbiotic culture advantage. Plant Cell, Tissue Organ Cult 90(3):313–323

    Article  Google Scholar 

  • Kauth PJ, Dutra D, Johnson TR, Stewart SL, Kane ME, Vendrame WA (2008) Techniques and applications of in vitro orchid seed germination. In: Teixeira DA, Silva JA (eds) Floriculture, ornamental and plant biotechnology: advances and topical issues. Global Science Books, Isleworth, pp 375–391

    Google Scholar 

  • Keel BG, Zettler LW, Kaplin BA (2011) Seed germination of Habenaria repens (Orchidaceae) in situ beyond its range, and its potential for assisted migration imposed by climate change. Castanea 76:43–54

    Article  Google Scholar 

  • Leake JR (2004) Mycoheterotroph/epiparasitic plant interaction with ectomycorrhizal and arbuscular mycorrhizal fungi. Curr Opin Plant Biol 7:422–428

    Article  CAS  PubMed  Google Scholar 

  • Ma YX, Liu YH, Zhang KY (1998) On microclimate edge of tropical rainforest fragments in Xishuangbanna. Acta Phytoecol Sin 22(3):250–255

    Google Scholar 

  • Ma M, Tan TK, Wong SM (2003) Identification and molecular phylogeny of Epulorhiza isolates from tropical orchids. Mycol Res 107:1041–1049

    Article  CAS  PubMed  Google Scholar 

  • Massey EE, Zettler LW (2007) An expanded role for in vitro symbiotic seed germination as a conservation tool: two case studies in North America (Platanthera leucophaea and Epidendrum nocturnum). Lankesteriana 7(1–2):303–308

    Google Scholar 

  • Mazumder PB, Sharma GD, Choudhury MD, Nath D, Talukdar AD, Mazumder B (2010) In vitro propagation and phytochemical screening of Papilionanthe teres (Roxb.) Schltr. Assam Univ J Sci Technol 5(1):37–42

    Google Scholar 

  • Moore RT (1987) The genera Rhizoctonia-like fungi: Ascorhizoctonia, Ceratorhizagen. Nov., Eupulorhiza gen. nov., Moniliopsis, and Rhizoctonia. Mycotaxon 29:91–99

    Google Scholar 

  • Nikabadi S, Bunn E, Stevens J, Newman B, Turner SR, Dixon KW (2014) Germination responses of four native terrestrial orchids from south-west Western Australia to temperature and light treatments. Plant Cell, Tissue Organ Cult 118(3):559–569

    Article  Google Scholar 

  • Nontachaiyapoom S, Sasirat S, Manoch L (2011a) Symbiotic seed germination of Grammatophyllum speciosum Blume and Dendrobium draconis Rchb. f., native orchids of Thailand. Sci Hortic 130(1):303–308

    Article  Google Scholar 

  • Nontachaiyapoom S, Sasirat S, Manoch L (2011b) Isolation and identification of Rhizoctonia-like fungi from roots of three orchid genera, Paphiopedilum, Dendrobium, and Cymbidium, collected in Chiang Rai and Chiang Mai provinces of Thailand. Mycorrhiza 20:459–471

    Article  Google Scholar 

  • Øien DI, O’Neill JP, Whigham DF, McCormick MK (2007) Germination ecology of the boreal-alpine terrestrial orchid Dactylorhiza lapponica (Orchidaceae). Annales Botanici Fennici 45:161–172

    Article  Google Scholar 

  • Osori-Gil EM, Forero-Montana J, Otero T (2008) Variation in mycorrhizal infection of the epiphytic orchid Ionopsis utriculariodes (Orchidiaceae) on different substrata. Carib J Sci 44(1):130–132

    Article  Google Scholar 

  • Otero JT, Thrall PH, Clements M, Burdon JJ, Miller JT (2011) Codiversification of orchids (Pterostylidinae) and their associated mycorrhizal fungi. Aust J Bot 59(5):480–497

    Article  Google Scholar 

  • Pereira OL, Rollemberg CL, Kasuya MCM (2003) Association des mycorhizies dans les orchidees–perspectives d’utilisation dans les programmes de propagation symbiotique. Orchidees 55:24–27

    Google Scholar 

  • Rasmussen HN (1995) Terrestrial orchids from seed to mycotrophic plant. Cambridge University Press, Cambridge

    Book  Google Scholar 

  • Rasmussen HN (2002) Recent developments in the study of orchid mycorrhiza. Plant Soil 244(1–2):149–163

    Article  CAS  Google Scholar 

  • Rasmussen HN, Whigham DF (1993) Seed ecology of dust seeds in situ: a new study technique and its application in terrestrial orchids. Am J Bot 80:1374–1378

    Article  Google Scholar 

  • Sánchez MS, Bills GF, Zabalgogeazcoa I (2008) Diversity and structure of the fungal endophytic assemblages from two sympatric coastal grasses. Fungal Divers 33:87–100

    Google Scholar 

  • Sathiyadash K, Muthukumar T, Murugan SB, Sathishkumar R, Pandey RR (2014) In vitro symbiotic seed germination of South Indian endemic orchid Coelogyne nervosa. Mycoscience 55(3):183–189

    Article  Google Scholar 

  • Scheffknecht S, Winkler M, Hülber K, Rosas MM, Hietz P (2010) Seedling establishment of epiphytic orchids in forests and coffee plantations in Central Veracruz, Mexico. J Trop Ecol 26(01):93–102

    Article  Google Scholar 

  • Sharma J, Zettler LW, Sambeek JW (2003) A survey of mycobionts of federally threatened Platanthera praeclara (Orchidaceae). Symbiosis 34:145–155

    Google Scholar 

  • Sheng CL, Lee YI, Gao JY (2012) Ex situ symbiotic seed germination, isolation and identification of effective symbiotic fungus in Cymbidium mannii (Orchidaceae). Chin J Plant Ecol 36:859–869

    Article  Google Scholar 

  • Smith SE, Read DJ (1997) Mycorrhizal symbiosis. Academic Press, Cambridge

    Google Scholar 

  • Smith ZF, James EA, McLean CB (2007) Experimental reintroduction of the threatened terrestrial orchid Diuris fragrantissima. Lankesteriana 7:377–380

    Google Scholar 

  • Stewart SL (2008) Orchid reintroduction in the United States: a mini-review. N Am Native Orchid J 14:54–59

    Google Scholar 

  • Stewart SL, Kane ME (2006) Symbiotic seed germination and in vitro seedling development of Habenaria macroceratitis (Orchidaceae), a rare Florida terrestrial orchid. Plant Cell, Tissue Organ Cult 86:147–158

    Article  CAS  Google Scholar 

  • Stewart SL, Zettler LW (2002) Symbiotic germination of three semi-aquatic rein orchids (Habenaria repens, H. quinquiseta, H. macroceratitis) from Florida. Aquat Bot 72(1):25–35

    Article  Google Scholar 

  • Stewart SL, Zettler LW, Minso J, Brown PM (2003) Symbiotic germination and reintroduction of Spiranthes brevilabris Lindley, an endangered orchid native to Florida. Selbyana 24:64–70

    Google Scholar 

  • Swangmaneecharern P, Serivichyaswat P, Nontachaiyapoom S (2012) Promoting effect of orchid mycorrhizal fungi Epulorhiza isolates on seed germination of Dendrobium orchids. Sci Hortic 148:55–58

    Article  Google Scholar 

  • Swarts ND, DixonKW (2009a) Terrestrial orchid conservation in the age of extinction. Ann Bot 104:543–556

    Article  PubMed  PubMed Central  Google Scholar 

  • Swarts ND, Dixon KW (2009b) Perspectives on orchid conservation in botanic gardens. Trends Plant Sci 14(11):590–598

    Article  CAS  PubMed  Google Scholar 

  • Tan XM, Cheng XM, Wang CL, Jin XH, Cai JL, Chen J, Guo SX, Zhao LF (2012) Isolation and identification of endophytic fungi in roots of nine Holcoglossum plants (Orchidaceae) collected from Yunnan, Guangxi and Hainan Provinces of China. Curr Microbiol 64:140–147

    Article  CAS  PubMed  Google Scholar 

  • Venturieri GA, Arbieto EAMD (2011) Ex-vitro establishment of Phalaenopsis amabilis seedlings in different substrates. Acta Sci Agron 33(3):495–501

    Article  Google Scholar 

  • Wang S, Xie Y (2004) China species red list. In: Wang S, Xie Y (eds) China species red list, vol 1, chap. 6. Higher Education Press, Beijing, p 459

    Google Scholar 

  • Wang D, Jia SH, Zhang ZX, Cai YP, Lin Y (2007) Isolation and culture of an endophytic fungus associated with Dendrobium huoshanense and its effects on the growth of plantlets. J Fungal Res 5:84–88

    CAS  Google Scholar 

  • Wang H, Fang H, Wang Y, Duan L, Guo S (2011) In situ seed baiting techniques in Dendrobium officinale Kimuraet Migo and Dendrobium nobile Lindl.: the endangered Chinese endemic Dendrobium (Orchidaceae). World J Microbiol Biotechnol 27(9):2051–2059

    Article  Google Scholar 

  • Warcup JH, Talbot PHB (1966) Perfect states of some Rhizoctonia. Trans Br Mycol Soc 49:427–435

    Article  Google Scholar 

  • Warcup JH, Talbot PHB (1971) Perfect states of Rhizoctonia associated with orchids III. New Phytol 70:35–40

    Article  Google Scholar 

  • White TJ, Bruns T, Lee S, Taylor JW (1990) Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. In: Innis MA, Gelfand DH, Sninsky JJ, White TJ (eds) PCR protocols: a guide to methods and applications. Academic, New York, pp 315–322

    Google Scholar 

  • Wu J, Ma H, Lü M, Han S, Zhu Y, Jin H, Xu J (2010) Rhizoctonia fungi enhance the growth of the endangered orchid Cymbidium goeringii. Botany 88:20–29

    Article  CAS  Google Scholar 

  • Yam TW, Arditti J (2009) History of orchid propagation: a mirror of the history of biotechnology. Plant Biotechnol Rep 3(1):1–56

    Article  Google Scholar 

  • Zelmer CD, Cuthbertson L, Currah RS (1996) Fungi associated with terrestrial orchid mycorrhizas, seeds and protocorms. Mycoscience 37(4):439–448

    Article  Google Scholar 

  • Zettler LW, Piskin KA (2011) Mycorrhizal fungi from protocorms, seedlings and mature plants of the eastern prairie fringed orchid, Platanthera leucophaea (Nutt.) Lindley: a comprehensive list to augment conservation. Am Midl Nat 166(1):29–39

    Article  Google Scholar 

  • Zettler LW, Piskin KA, Stewart SL, Hartsock JJ, Bowles ML, Bell TJ (2005) Protocorm mycobionts of the federally threatened eastern prairie fringed orchid, Platanthera leucophaea (Nutt.) Lindley, and a technique to prompt leaf elongation in seedlings. Stud Mycol 53:163–171

    Article  Google Scholar 

  • Zhang J, Cao M (1995) Tropical forest vegetation of Xishuangbanna, SW China and its secondary changes, with special reference to some problems in local nature conservation. Biol Conserv 73(3):229–238

    Article  Google Scholar 

  • Zi XM, Sheng CL, Goodale UM, Shao SC, Gao JY (2014) In situ seed baiting to isolate germination-enhancing fungi for an epiphytic orchid, Dendrobium aphyllum (Orchidaceae). Mycorrhiza 24(7):487–499

    Article  CAS  PubMed  Google Scholar 

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Acknowledgments

This study was supported by the Natural Science Foundation of Yunnan Province (grant no. 2013FB080) and National Natural Science Foundation of China (grant no. 31470450). The authors gratefully acknowledge the Central Laboratory of Xishuangbanna Tropical Botanical Garden for providing the research facilities.

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Zhou, X., Gao, J. Highly compatible Epa-01 strain promotes seed germination and protocorm development of Papilionanthe teres (Orchidaceae). Plant Cell Tiss Organ Cult 125, 479–493 (2016). https://doi.org/10.1007/s11240-016-0964-y

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