Non-mycorrhizal fungal endophytes in two orchids of Kaiga forest (Western Ghats), India

Abstract

We used standard isolation protocols to explore the endophytic fungal communities in three tissue types of two dominant orchids (Bulbophyllum neilgherrense and Vanda testacea) of the Kaiga forest of the Western Ghats. We surface sterilized and assessed 90 segments of each orchid for the occurrence and diversity of endophytic fungal taxa. The 118 fungal isolates were obtained from root, bulb and leaves of B. neilgherrense, consisting of 17 anamorphic taxa (range, 10–15 taxa) with 1.3 fungal taxa per segment (range, 1.2–1.4 taxa). Four taxa (Aspergillus flavus, A. niger, Penicillium sp. and morpho sp. 1) belonged to the core group (11.1%–32.2%). The relative abundance of A. flavus and morpho sp. 1 was more than 10%. A total of 130 fungal isolates from roots, stems and leaves of V. testacea yielded 20 anamorphic taxa (range, 11–15 taxa) with 1.4 fungal taxa per segment (range, 1.4–1.5 taxa). Aspergillus flavus, A. niger, A. ochraceus, Gliocladium viride, Penicillium sp. and morpho sp. 1 belonged to the core group. Relative abundance exceeded 10% for A. flavus, A. niger, and morpho sp. 1. The Simpson and Shannon diversity indices were higher in leaf than root or bulb/stem of both orchids. Jaccard’s similarity coefficient was higher between root and leaf in both orchids (56.3%-60%) than between other pairs. Our study revealed that the endophytic fungal assemblage and diversity of B. neilgherrense and V. testacea of Kaiga forest of the Western Ghats were relatively similar between orchids and their tissues.

Keywords

orchids / Bulbophyllum / Vanda / endophytic fungi / Western Ghats

References

[1] Abadie J.C., Püttsepp U., Gebauer G., Faccio A., Bonfante P., Selosse M.A.. Cephalanthera longifolia (Neottieae, Orchidaceae) is mixotrophic: A comparative study between green and non-photosynthetic individuals. Canadian Journal of Botany, 2006, 84: 1462-1477.

[2] Abraham A., Vatsala P.. Introduction to Orchids. 1981, Trivandrum, India: St Joseph’s Press, 220.

[3] Alias S.A., Kuthubutheen A.J., Jones E.B.G.. Frequency of occurrence of fungi on wood in Malaysian mangroves. Hydrobiologia, 1995, 295: 97-106.

[4] Vaz A. B.M., Mota R. C., Bomfim M. R. Q., Mariana L.A., Vieira, Zani C. L., Rosa C. A., Rosa L. H.. Antimicrobial activity of endophytic fungi associated with Orchidaceae in Brazil. Canadian Journal of Microbiology, 2009, 55: 1381-1391.

[5] Ananda K., Sridhar K.R.. Diversity of endophytic fungi in the roots of mangrove species on the west coast of India. Canadian Journal of Microbiology, 2002, 48: 871-878.

[6] Arnold A.E., Lutzoni F.. Diversity and host range of foliar fungal endophytes: are tropical leaves biodiversity hotspots?. Ecology, 2007, 88: 541-549.

[7] Arnold A.E.. Understanding the diversity of foliar endophytic fungi: progress, challenges, and frontiers. Fungal Biology Review, 2007, 21: 51-66.

[8] Bae H., Kim S., Sicher R.C. Jr, Kim M.S., Strem M.D., Bailey B.A.. The beneficial endophyte, Trichoderma hamatum, delays the onset of drought stress in Theobroma cacao. Biological Control, 2008, 46: 24-35.

[9] Bagyalakshmi G., Muthukumar T., Sathiyadash K., Muniappan V.. Mycorrhizal and dark septate fungal associations in shola species of Western Ghats, southern India. Mycoscience, 2010, 9: 297-313.

[10] Bayman P., Lebron L.L., Trembly R.L., Lodge D.J.. Variation in endophytic fungi from roots and leaves of Lepanthes (Orchidaceae). New Phytologist, 1997, 135: 143-149.

[11] Bayman P., Otero J.T.. Schulz B., Boyle C., Sieber T.. Microbial endophytes of orchid roots. Microbial Root Endophytes. 2006, New York: Springer, 153 173

[12] Bhat D.J.. Fascinating Microfungi (Hyphomycetes) of Western Ghats — India. 2010, Goa: Broadway Publishers, 221.

[13] Bills G.F., Polishook J.D.. Microfungi from Carpinus caroliniana. Canadian Journal of Botany, 1991, 69: 1477-1482.

[14] Bonnardeaux Y., Brundrett M., Batty A., Dixon K., Koch J., Sivasithamparam K.. Diversity of mycorrhizal fungi of terrestrial orchids: compatibility webs, brief encounters, lasting relationships and alien invasions. Mycological Research, 2007, 111: 51-61.

[15] Cai L., Hyde K.D., Tsui C.K.M.. Genera of Freshwater Fungi. Fungal Diversity Research Series # 18. 2006, Hong Kong: Fungal Diversity Press, 261.

[16] Carmichael J.W., Kendrick W.B., Conners I.L., Sigler L.. Genera of Hyphomycetes. 1980, Edmonton, Canada: The University of Alberta Press, 386.

[17] Chen J., Wang H., Gua S.-X.. Isolation and identification of endophytic and mycorrhizal fungi from seeds and roots of Dendrobium (Orchidaceae). Mycorrhiza, 2012, 22(4): 297-307.

[18] Chen X.M., Dong H.L., Hu K.X., Sun Z.R., Chen J., Guo S.-X.. Diversity and antimicrobial and plant-growth-promoting activities of endophytic fungi in Dendrobium loddigesii Rolfe. Journal of Plant Growth Regulation, 2010, 29: 328-337.

[19] Chen X.M., Guo S.X.. Effects of four species of endophytic fungi on the growth and polysaccharide and alkaloid contents of Dendrobium nobile. China Journal of Chinese Materia Medica, 2005, 30: 253-257.

[20] Christensen M.. A view of fungal ecology. Mycologia, 1989, 81: 1-19.

[21] Dearnaley J.D.W.. Further advances in orchid mycorrhizal research. Mycorrhiza, 2007, 17: 475-486.

[22] Elena V., Gastaldo A., Tondello A., Baldan B., Villani M., Squartini A.. Identification of two fungal endophytes associated with the endangered orchid Orchis militaris L. Journal of Microbiology and Biotechnology, 2010, 20: 630-636.

[23] Ellis M.B., Ellis J.P.. Microfungi on Land Plants: An Identification Handbook. 1987, London: Croom Helm, 818.

[24] Ellis M.B.. Dematiaceous Hyphomycetes. 1971, Kew: Commonwealth Mycological Institute, 608.

[25] Ellis M.B.. More Dematiaceous Hyphomycetes. 1976, Kew: Commonwealth Mycological Institute

[26] Ganesan R., Livingstone C.. Checklist of orchids from a mid-elevation evergreen forest at Kakachip Kodayar, Kalakkad-Mundanthurai tiger reserve Agasthyamalai, Southern Western Ghats. Zoos’ Print Journal, 2001, 16: 445-446.

[27] Gardes M.. An orchid-fungus marriage: physical promiscuity, conflict and cheating. New Phytologist, 2002, 154: 4-7.

[28] Gunatilaka A.A.L.. Natural products from plant-associated microorganisms: Distribution, structural diversity, bioactivity, and implications of their occurrence. Journal of Natural Products, 2006, 69: 509-526.

[29] Hawksworth D.L.. Fungal diversity and its implications for genetic resource collections. Studies in Mycology, 2004, 50: 9-18.

[30] Hyde K.D., Soytong K.. The fungal endoophyte dilemma. Fungal Diversity, 2008, 33: 163-173.

[31] Irwin M.J., Bougoure J.J., Dearnaley J.D.W.. Pterostylis nutans (Orchidaceae) has a specific association with two Ceratobasidium root associated fungi across its range in Eastern Australia. Mycoscience, 2007, 48: 231-239.

[32] Jones D.L.. A Complete Guide to Native Orchids of Australia including the Island Territories. 2006, Sydney: Reed New Holland, 363.

[33] Kaliamoorthy S.. Pattern of mycorrhizal infections in the roots of Aerides maculosum Lindl. and Calanthe triplicate (Willem.) Ames. Mycorrhiza News, 2007, 19: 14-18.

[34] Kaveriappa K.M., Shetty B.V.. Biodiversity of the Western Ghats with special reference to conservation of plant diversity at Kaiga. Nu Power, 2001, 15: 40-42.

[35] Kenkel N.C., Booth T.. Carroll G.C., Wicklow D.T.. Multivariate analysis in fungal ecology. The Fungal Community: its Organization and Role in the Ecosystem. 1992, New York: Dekker, 209 227

[36] Kobayashi D.Y., Palumbo J.D.. Bacon C.W., White J.F.. Bacterial endophytes and their effects on plants and uses in agriculture. Microbial Endophytes. 2000, New York: Marcel Dekker, 199 236

[37] Kumar C.S., Shetty B.V., Bennet S.S.R., Rao T.A., Molur S., Walker S.. Endemic Orchids of the Western Ghats. 2001, Coimbatore, India: Zoo Outreach Organization, 139.

[38] Ludwig J.A., Reynolds J.F.. Statistical Ecology — A Primer on Methods and Computing. 1988, New York: John Wiley and Sons, 337.

[39] Magurran A.E.. Ecological Diversity and its Measurement. 1988, New Jersey: Princeton University Press, 192.

[40] McCormick M.K., Whigham D.F., O’Neill J.. Mycorrhizal diversity in photosynthetic terrestrial orchids. New Phytologist, 2004, 163: 425-438.

[41] Pereira O.L., Kasuya M.C.M., Borges A.C., de Araújo E.F.. Morphological and molecular characterization of mycorrhizal fungi isolated from neotropical orchids in Brazil. Canadian Journal of Botany, 2005, 83: 54-65.

[42] Pereira O.L., Rollemberg C.L., Borges A.C., Matsuoka K., Kasuya M.C.M.. Epulorhiza epiphytica sp nov isolated from mycorrhizal roots of epiphytic orchids in Brazil. Mycoscience, 2003, 44: 153-155.

[43] Peters A.F.. Field and culture studies of Streblonema-Macrocystis new species Ectocarpales Phaeophyceae from Chile, a sexual endophyte of giant kelp. Phycologia, 1991, 30: 365-377.

[44] Petrini O.. Endophytische Pilze in Epiphytischen Araceae, Bromeliaceae und Orchidiaceae. Sydowia, 1981, 34: 135-148.

[45] Petrini O.. Andrews J., Hirano S.. Fungal endophytes of tree leaves. Microbiol Ecology of Leaves. 1991, New York: Springer Verlag, 179 197

[46] Pielou F.D.. Ecological Diversity. 1975, New York: Wiley InterScience, 165.

[47] Pinuran U., Rungjindamai N., Choeyklin R., Lumyong S., Hyde K.D., Jones E.B.G.. Occurrence and diversity of badidiomycetous endophytes from the oil palm, Elaeis guineensis in Thailand. Fungal Diversity, 2010, 41: 71-88.

[48] Rasmussen H.N.. Recent developments in the study of orchid mycorrhiza. Plant and Soil, 2002, 244: 149-163.

[49] Redman R.S., Sheehan K.B., Stout T.G., Rodriguez R.J., Henson J.M.. Thermotolerance generated by plant/fungal symbiosis. Science, 2002, 298 1581

[50] Rungjindamai N., Pinuran U., Cheyklin R., Hattori T., Jones E.B.G.. Molecular characterization of basidiomycetous endophytes isolated from leaves, rachis and petioles of the oil palm, Elaeis guineensis, in Thailand. Fungal Diversity, 2008, 33: 139-161.

[51] Sarma V.V., Hyde K.D.. A review on frequently occurring fungi in mangroves. Fungal Diversity, 2001, 8: 1-34.

[52] Schulz B., Boyle C., Draeger S., Römmert A.-K., Krohn K.. Endophytic fungi: a source of novel biologically active secondary metabolites. Mycological Research, 2002, 106: 996-1004.

[53] Schulz B., Boyle C.. The endophytic continuum. Mycological Research, 2005, 109: 661-686.

[54] Seena S., Sridhar K.R.. Endophytic fungal diversity of 2 sand dune wild legumes from the southwest coast of India. Canadian Journal of Microbiology, 2004, 50: 1015-1021.

[55] Selosse M., Vohník M., Chauvet E.. Out of rivers: Are some aquatic hypohomycetes plant endophytes?. New Phytologist, 2008, 198: 3-7.

[56] Shefferson R.P., Weiß M., Kull T., Taylor D.L.. High specificity generally characterizes mycorrhizal association in rare lady’s slipper orchids, genus Cypripedium. Molecular Ecology, 2005, 14: 613-626.

[57] Sieber T.N.. Waisel Y., Eshel A., Kafkafi U.. Fungal root endophytes. The Hidden Half. 2002, New York: Marcel Dekker, 887 917

[58] Smith S.E., Read D.J.. Mycorrhizal Symbiosis. 1997, Cambridge: Academic Press, 312.

[59] Sokolski S., Piché Y., Chauvet E., Bérubé J.A.. A fungal endophyte of black spruce (Picea mariana) needles is also an aquatic hyphomycete. Molecular Ecology, 2006, 15: 1955-1962.

[60] Sridhar K.R., Bärlocher F.. Endophytic aquatic hyphomycetes in spruce, birch and maple. Mycological Research, 1992, 96: 305-308.

[61] Stark C., Babik W., Durka W.. Fungi from the roots of the common terrestrial orchid Gymnadenia conopsea. Mycological Research, 2009, 13: 952-959.

[62] Stone J.K., Bacon C.W., White J.F.. Bacon C.W., White J.F.. An overview of endophytic microbes: endophytism defined. Microbial Endophytes. 2000, New York: Dekker, 3 30

[63] Suarez J.P., Weiß M., Abele A., Garnica S., Oberwinkler F., Kottke I.. Diverse tulasnelloid fungi form mycorrhizas with epiphytic orchids in an Andean cloud forest. Mycological Research, 2006, 110: 1257-1270.

[64] Suryanarayanan T.S., Thirunavukkarasu N., Govindarajulu M.B., Sasse F., Jansen R., Murali T.S.. Fungal endophytes and bioprospecting. Fungal Biology Review, 2009, 23: 72-85.

[65] Swamy K.K., Kumar K.N.K., Ramakrishna T.M., Ramaswamy S.N.. Studies on seed morphometry of epiphytic orchids from Western Ghats of Karnataka. Taiwania, 2004, 49: 124-140.

[66] Swarts N.D., Sinclair E.A., Francis A., Dixon K.W.. Ecological specialization in mycorrhizal symbiosis leads to rarity in an endangered orchid. Molecular Ecology, 2010, 19: 3226-3242.

[67] Tan R.X., Zou W.X.. Endophytes: a rich source of functional metabolites. Natural Product Report, 2001, 18: 448-459.

[68] Tao G., Liu Z.Y., Hyde K.D., Lui X.Z., Yu Z.N.. Whole rDNA analysis reveals novel and endophytic fungi in Bletilla ochracea (Orchidaceae). Fungal Diversity, 2008, 33: 101-122.

[69] Taylor D.L., Bruns T.D., Szaro T.M., Hodges S.A.. Divergence in mycorrhizal specialization within Hexalectris spicata (Orchidaceae), a nonphotosynthetic desert orchid. American Journal of Botany, 2003, 90: 1168-1179.

[70] Taylor J.E., Hyde K.D., Jones E.B.G.. Endophytic fungi associated with the temperate palm, Trachycarpus fortunei, within and outside its natural geographic range. New Phytologist, 1999, 142: 335-346.

[71] Vega F.E., Posada F., Catherine A.M., Pava-Ripoll M., Infante F., Rehner S.A.. Entomopathogenic fungal endophytes. Biological Control, 2008, 46: 72-82.

[72] Zettler L.W., Sharma J., Rasmussen F.. Dixon K., Cribb P., Kell S., Barrett R.. Mycorrhizal diversity. Orchid Conservation. 2004, Malaysia: Natural History Publications, 185 203

[73] Zhi-lin Y., Yi-cum C., Yun Yang.. Diverse non-mycorrhizal fungal endophytes inhabiting an epiphytic, medicinal orchid (Dendrobium nobile): estimation and characterization. World Journal of Microbiology and Biotechnology, 2009, 25: 295-303.