Jianghua Huang¹, Zide Jiang²

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¹ Institute of Plant Pathology, Zhongkai University of Agricultue and Engineering, Guangzhou, China.
² Department of Plant Pathology, College of Natural Resources and Environment, South China Agriculture University, Guangzhou, China.
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Abstract:

This study aims to isolate and identify taxonomic characterization of endophytic fungi from bitter melon in Guangdong province, China. A total of 1172 endophytic fungi are isolated from roots, stems, leaves, flowers, and fruits of healthy plants, and they are classified to 49 taxa based on morphological and molecular features. The results show that endophytic fungi from bitter melon plants exhibit high biodiversity. Arthrinium aureum, A. marii, A. sphaerospermum, Corynascus verrucosus, Curvularia borreriae and C. protuberate have not been recorded in any plants in China. Basidiomycetous endophytes, such as Ceratobasidium sp., and C. cornigerum, are reported in this study for the first time in bitter melon. According to the results of phylogenetic analysis, the rDNA ITS (Internal Transcribed Spacer) sequences can successfully separate species, such as F. solani, F. kyushuense, C. verrucosus, C. globosum, E. rostratum, C. brachyspora, C. verruculosus, C. affinis, P. bougainvilleicola, P. longicolla, P. glabrae, P. verruculosum, P. oxalicum, P. citrinum, P. chermesinum, P. glomerata, A. fumigatus and A. japonicas. Some isolates belonging to A. alternata, C. gloeosporioides, C. cladosporioides, C. brasiliense, C. convolutum, F. proliferatum, F. oxysporum, F. verticillioides, F. equiseti, F. camptoceras and Xylaria, however, require the analysis of others molecular markers to provide better taxonomic resolution. Molecular analyses of rDNA ITS sequences are useful for identification and classification of endophytes of bitter melon, but it is desirable to consider an integrated approach, such as molecular phylogeny, host, colony growth rate, symptoms, and shape of conidia.

Keywords: Endophytic Fungi, Momordica charantia, Phylogenetic Analysis

Cite this paper: Huang, J. and Jiang, Z. (2015) Phylogenetic analysis of Endophytes from Bitter Melon (Momordica charantia) in Guangdong Province. Agricultural Sciences, 6, 609-621. doi: 10.4236/as.2015.66060.

References

[1]   Arnold, A.E., Maynard, Z., Gilbert, G.S., Coley, P.D. and Kursar, T.A. (2000) Are Tropical Fungal Endophytes Hyper-diverse? Ecology Letters, 3, 267-274.
http://dx.doi.org/10.1046/j.1461-0248.2000.00159.x

[2]   Suryanarayanan, T.S., Murali, T.S. and Venkatesan, G. (2002) Occurrence and Distribution of Fungal Endophytes in Tropical Forests across a Rainfall Gradient. Canadian Journal of Botany, 80, 818-826.
http://dx.doi.org/10.1139/b02-069

[3]   Promputtha, I., Lumyong, S., Dhanasekaran, V., McKenzie, E.H.C., Hyde, K.D. and Jeewon, R. (2007) A Phylogenetic Evaluation of Whether Endophytes Become Saprotrophs at Host Senescence. Microbial Ecology, 53, 579-590.
http://dx.doi.org/10.1007/s00248-006-9117-x

[4]   Rosa, L.H., Aline, B.M.V., Caligiorne, R.B., Campolina, S. and Rosa, C.A. (2009) Endophytic Fungi Associated with the Antarctic Grass Deschampsia antarctica Desv. (Poaceae). Polar Biology, 32, 161-167.
http://dx.doi.org/10.1007/s00300-008-0515-z

[5]   de Siqueira, V.M., Conti, R., de Araújo, J.M. and Souza-Motta, C.M. (2011) Endophytic Fungi from the Medicinal Plant Lippia sidoides Cham. and Their Antimicrobial Activity. Symbiosis, 53, 89-95.
http://dx.doi.org/10.1007/s13199-011-0113-7

[6]   Lin, X., Lu, C.H., Huang, Y.J., Zheng, Z.H., Su, W.J. and Shen, Y.M. (2007) Endophytic Fungi from a Pharmaceutical Plant, Camptotheca acuminata: Isolation, Identification and Bioactivity. World Journal of Microbiology and Biotechnology, 23, 1037-1040.
http://dx.doi.org/10.1007/s11274-006-9329-8

[7]   Hyde, K.D. (2001) Where Are the Missing Fungi? Does Hong Kong Have Any Answers? Mycological Research, 105, 1514-1518.
http://dx.doi.org/10.1017/S0953756201004889

[8]   Photita, W., Lumyong, S., Lumyong, P. and Hyde, K.D. (2001) Endophytic Fungi of Wild Banana (Musa acuminata) at Doi Suthep Pui National Park, Thailand. Mycological Research, 105, 1508-1513.
http://dx.doi.org/10.1017/S0953756201004968

[9]   Bérdy, J. (2005) Bioactive Microbial Metabolites. Journal of Antibiotics, 58, 1-26.
http://dx.doi.org/10.1038/ja.2005.1

[10]   Fernandes, N.P., Lagishetty, C.V., Panda, V.S. and Naik, S.R. (2007) An Experimental Evaluation of the Antidiabetic and Antilipidemic Properties of a Standardized Momordica charantia Fruit Extract. BMC Complement. BMC Complementary and Alternative Medicine, 7, 29.
http://dx.doi.org/10.1186/1472-6882-7-29

[11]   Limtrakul, P., Khantamat, O. and Pintha, K. (2004) Inhibition of P-Glycoprotein Activity and Reversal of Cancer Multidrug Resistance by Momordica charantia Extract. Cancer Chemotherapy and Pharmacology, 54, 525-530.
http://dx.doi.org/10.1007/s00280-004-0848-4

[12]   Sette, L.D., Passarini, M.R.Z., Delarmelina, C., Salati, F. and Duarte, M.C.T. (2006) Molecular Characterization and Antimicrobial Activity of Endophytic Fungi from Coffee Plants. World Journal of Microbiology and Biotechnology, 22, 1185-1195.
http://dx.doi.org/10.1007/s11274-006-9160-2

[13]   Chen, Y.X., Zhang, L.P. and Lu, Z.T. (2008) Analysis of the Internal Transcribed Spacer (ITS) Sequences in rDNA of 10 Strains of Fusarium spp. Journal of Anhui Agricultural Sciences, 36, 4886-4887.

[14]   Schulz, B., Wanke, U. and Draeger, S. (1993) Endophytes from Herbaceous and Shrubs: Effectiveness of Surface Sterilization Methods. Mycological Research, 97, 1447-1450.
http://dx.doi.org/10.1016/S0953-7562(09)80215-3

[15]   Song, Z.H., Ding, L.X., Ma, B.J. and Li, W.Z. (1999) Studies on the Population and Dynamic Analysis of Peanut Endophytes. Acta Phytophylacica Sinica, 26, 309-314.

[16]   Lee, S.B. and Taylor, J.W. (1990) Isolation of DNA from Fungal Mycelia and Single Spores. In: Innis, M.A., Gelfand, D.H., Sninsky, J.J. and White, T.J., Eds., PCR Protocols: A Guide to Methods and Applications, Academic Press, San Diego, 282-287.
http://dx.doi.org/10.1016/b978-0-12-372180-8.50038-x

[17]   White, T.J., Bruns, T., Lee, S. and Taylor, J. (1990) Amplification and Direct Sequencing of Fungal Ribosomal RNA Genes for Phylogeentics. In: Innis, M.A., Gelfand, D.H., Sninsky, J.J. and White, T.J., Eds., PCR Protocols: A Guide to Methods and Applications, Academic Press, San Diego, 315-322.

[18]   Thompson, J.D., Gibson, T.J., Plewniak, F., Jeanmougin, F. and Higgins, D.G. (1997) The CLUSTALX Windows Interface: Flexible Strategies for Multiple Sequence Alignment Aided by Quality Analysis Tools. Nucleic Acids Research, 25, 4876-4882.
http://dx.doi.org/10.1093/nar/25.24.4876

[19]   Tamura, K., Dudley, J., Nei, M. and Kumar, S. (2007). MEGA4: Molecular Evolutionary Genetics Analysis (MEGA) Software. Version 4.0. Molecular Biology and Evolution, 24, 1596-1599.
http://dx.doi.org/10.1093/molbev/msm092

[20]   Kumar, S., Tamura, K. and Nei, M. (2004) MEGA3: Intergrated Software for Molecular Evolutionary Genetics Analysis and Sequence Alignment. Brief Bioinformatics, 5, 150-163.
http://dx.doi.org/10.1093/bib/5.2.150

[21]   Guo, L.D., Hyde, K.D. and Liew, E.C.Y. (2001) Detection and Identification of Endophytic Fungi within Frond Tissues of Livistona chinensis Based on rDNA Sequence. Molecular Phylogenetics and Evolution, 20, 1-13.
http://dx.doi.org/10.1006/mpev.2001.0942

[22]   Guo, L.D., Huang, G.R., Wang, Y., He, W.H., Zheng, W.H. and Hyde, K.D. (2003) Molecular Identification of White Morphotype Strains of Endophytic Fungi from Pinus tabulaeformis. Mycological Research, 107, 680-688.
http://dx.doi.org/10.1017/S0953756203007834

[23]   Promputtha, L., Jeewon, R., Lumyong, S., Mckenzie, E.H.C. and Hyde, K.D. (2005) Ribosomal DNA Fingerprinting in the Identification of Non-Sporuating Endophytes from Magnolia liliifera (Magnoliaceae). Fungal Diversity, 20, 167-186.

[24]   Wang, Y., Guo, L.D. and Hyde, K.D. (2005) Taxonomic Placement of Sterile Morphotypes of Endophytic Fungi from Pinus tabulaeformis (Pinaceae) in Northeast China Based on rDNA Sequences. Fungal Diversity. 20, 235-260.

[25]   Wang, Y.T., Lo, H.S. and Wang, P.H. (2008) Endophytic Fungi from Taxus mairei in Taiwan: First Report of Colletotrichum Gloeosporioides as an Endophyte of Taxus mairei. Botanical Studies, 49, 39-43.

[26]   Liu, K.H., Ding, X.W., Deng, B.W. and Chen, W.Q. (2009) Isolation and Characterization of Endophytic Taxol-Producing Fungi from Taxus chinensis. Journal of Industrial Microbiology & Biotechnology, 36, 1171-1177.
http://dx.doi.org/10.1007/s10295-009-0598-8

[27]   Caruso, M., Colombo, A.L., Fedeli, L., Pavesi, A., Quaroni, S., Saracchi, M. and Bentrella, G. (2000) Isolation of Endophytic Fungi and Actinomycetes Taxane Producers. Annals of Microbiology, 50, 3-13.

[28]   Rivera-Orduna, F.N., Suarez-Sanchez, R.A., Flores-Bustamante, Z.R., Gracida-Rodriguez, J.N. and Flores-Cotera, L.B. (2011) Diversity of Endophytic Fungi of Taxus globosa (Mexican Yew). Fungal Diversity, 47, 65-74.
http://dx.doi.org/10.1007/s13225-010-0045-1

[29]   Kharwar, R.N., Gond, S.K., Kumar, A. and Mishra, A. (2010) A Comparative Study of Endophytic and Epiphytic Fungal Associated with Leaf of Eucalyptus citriodora Hook, and Their Antimicrobial Activity. World Journal of Microbiology and Biotechnology, 26, 1941-1948.
http://dx.doi.org/10.1007/s11274-010-0374-y

[30]   Kharwar, R.N., Verma, S.K., Mishra, A., Gond, S.K., Sharma, V.K., Afreen, T. and Kumar, A. (2011) Assessment of Diversity, Distribution and Antibacterial Activity of Endophytic Fungi Isolated from a Medicinal Plant Adenocalymma alliaceum Miers. Symbiosis, 55, 39-46.
http://dx.doi.org/10.1007/s13199-011-0142-2

[31]   Sánchez Márquez, S., Bills, G.F. and Zabalgogeazcoa, I. (2007) The Endophytic Mycobiota of the Grass Dactylis glomerata. Fungal Diversity, 27, 171-195.

[32]   Sánchez Márquez, S., Bills, G.F., Dominguez, A.L. and Zabalgogea, I. (2010) Endophytic Mycobiota of Leaves and Roots of the Grass Holcus lanatus. Fungal Diversity, 41, 115-123.
http://dx.doi.org/10.1007/s13225-009-0015-7

[33]   Thomas, S.E., Crozier, J., Aime, M.C., Evans, H.C. and Holmes, K.A. (2008) Molecular Characterization of Fungal Endophytic Morphospecies Associated with the Indigeneous Forest Tree, Theobroma gileri, in Ecuador. Mycological Research, 112, 852-860.
http://dx.doi.org/10.1016/j.mycres.2008.01.008

[34]   Pinruan, U., Rungjindamai, N., Choeyklin, R., Lumyong, S., Hyde, K.D. and Jones, E.B.G. (2010) Occurrence and Diversity of Basidiomycetous Endophytes from the Oil Palm, Elaeis guineensis in Thailand. Fungal Diversity, 41, 71-88.
http://dx.doi.org/10.1007/s13225-010-0029-1

[35]   Beever, R.E. and Weeds, P.L. (2004) Taxonomy and Genetic Variation of Botrytis and Botryotinia. In: Elad, Y., Williamson, B., Tudzynski, P. and Delan, N., Eds., Botrytis: Biology, Pathology and Control, Kluwer Academic Publishers, Dordrecht, 29-52.

[36]   White, T.J., Bruns, T.D., Lee, S. and Taylor, J. (1990) Amplification and Sequencing of Fungal Ribosomal RNA Genes for Phylogenetics. In: Innis, M.A., Gelfand, D.H., Sninsky, J.J. and White J.J., Eds., PCR Protocols: A Guide to Methods and Applications, Academic Press, San Diego, 315-322.
http://dx.doi.org/10.1016/b978-0-12-372180-8.50042-1