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Isolation and characterization of endophytic taxol-producing fungi from Taxus chinensis

  • Kaihui Liu
  • Xiaowei Ding
  • Baiwan DengEmail author
  • Wenqiang Chen
Original Paper

Abstract

This study investigated the endophytic fungi diversity of Taxus chinensis and screened the taxol-producing fungi in the host. A total of 115 endophytic fungi isolates obtained from bark segments of T. chinensis were grouped into 23 genera based on the morphological traits and sequence analysis of the internal transcribed spacers (ITS1-5.8S-ITS2), indicating endophytic fungi in T. chinensis are diverse and abundant. Diaporthe, Phomopsis (anamorph of Diaporthe), Acremonium, and Pezicula were the dominant genera, whereas the remaining genera were infrequent groups. The 13 representative species of the distinct genera were capable of producing taxol verified by reverse-phase high performance liquid chromatography (HPLC). Among the taxol-producing fungi, the yield of taxol produced by the Metarhizium anisopliae, H-27 was 846.1 μg l−1 in reformative potato dextrose liquid medium, and the fungal taxol was further validated by mass spectrometry (MS). The taxol-producing fungi (92.3%) were infrequent communities, suggesting that infrequent fungi associated with T. chinensis might be a fascinating reservoir of taxol-generating fungi.

Keywords

Endophytic fungi ITS sequence Phylogenetic diversity Taxol Taxus chinensis 

Notes

Acknowledgments

The work was co-supported by funds from the Shaanxi Educational Committee (08JZ21, 08JK244, and 08JK248) and partially financed by grants from Shaanxi University of Technology (SLGQDO712, SLGQDO713). We are grateful to Dr. Alan T. Bull and the anonymous reviewer for their helpful comments on our manuscript.

References

  1. 1.
    Barber PA, Burgess TJ, Hardy GE, Slippers B, Keane PJ, Wingfield MJ (2005) Botryosphaeria species from Eucalyptus in Australia are pleoanamorphic, producing Dichomera synanamorphs in culture. Mycol Res 109:1347–1363. doi: 10.1017/S0953756205003989 PubMedCrossRefGoogle Scholar
  2. 2.
    Carroll G (1988) Fungal endophytes in stems and leaves: from latent pathogens to mutualistic symbiont. Ecology 69:2–9. doi: 10.2307/1943154 CrossRefGoogle Scholar
  3. 3.
    Caruso M, Colombo AL, Fedeli L, Pavesi A, Quaroni S, Saracchi M, Ventrella G (2000) Isolation of endophytic fungi and actinomycetes taxane producers. Ann Microbiol 50:3–13Google Scholar
  4. 4.
    Deng BW, Liu KH, Chen WQ, Ding XW, Xie XC (2009) Fusarium solani, Tax-3, a new endophytic taxol-producing fungus from Taxus chinensis. World J Microbiol Biotechnol 25:139–143. doi: 10.1007/s11274-008-9876-2 CrossRefGoogle Scholar
  5. 5.
    Fisher PJ, Petrini O (1987) Location of fungal endophytes in tissue of Suaeda fruticosa: a preliminary study. Trans Br Mycol Soc 89:246–249CrossRefGoogle Scholar
  6. 6.
    Gangadevi V, Muthumary J (2008) Taxol, an anticancer drug produced by an endophytic fungus Bartalinia robillardoides Tassi, isolated from a medicinal plant, Aegle marmelos Correa ex Roxb. World J Microbiol Biotechnol 24:717–724. doi: 10.1007/s11274-007-9530-4 CrossRefGoogle Scholar
  7. 7.
    Ganley RJ, Brunsfeld SJ, Newcombe G (2004) A community of unknown, endophytic fungi in western white pine. Proc Natl Acad Sci USA 101:10107–10112. doi: 10.1073/pnas.0401513101 PubMedCrossRefGoogle Scholar
  8. 8.
    Higgins KL, Arnold AE, Miadlikowska J, Sarvate SD, Lutzoni F (2007) Phylogenetic relationships, host affinity, and geographic structure of boreal and arctic endophytes from three major plant lineages. Mol Phylogenet Evol 42:543–555. doi: 10.1016/j.ympev.2006.07.012 PubMedCrossRefGoogle Scholar
  9. 9.
    Huang Y, Wang J, Li G, Zheng Z, Su W (2001) Antitumor and antifungal activities in endophytic fungi isolated from pharmaceutical plants Taxus mairei, Cephalataxus fortunei and Torreyagrandis. FEMS Immunol Med Microbiol 31:163–167. doi: 10.1111/j.1574-695X.2001.tb00513.x PubMedCrossRefGoogle Scholar
  10. 10.
    Kumar S, Tamura K, Nei M (2004) MEGA3: integrated software for molecular evolutionary genetics analysis and sequence alignment. Brief Bioinform 5:150–163. doi: 10.1093/bib/5.2.150 PubMedCrossRefGoogle Scholar
  11. 11.
    Lin X, Lu CH, Huang YJ, Zheng ZH, Su WJ, Shen YM (2007) Endophytic fungi from a pharmaceutical plant, Camptotheca acuminata: isolation, identification and bioactivity. World J Microbiol Biotechnol 23:1037–1040. doi: 10.1007/s11274-006-9329-8 CrossRefGoogle Scholar
  12. 12.
    Miller JD, Mackenzie S, Foto M, Adams GW, Findlay JA (2002) Needles of white spruce inoculated with rugulosin-producing endophytes contain rugulosin reducing spruce budworm growth rate. Mycol Res 106:471–479. doi: 10.1017/S0953756202005671 CrossRefGoogle Scholar
  13. 13.
    Paul NC, Kim WK, Woo SK, Park MS, Yu SH (2007) Fungal endophytes in roots of Aralia species and their antifungal activity. Plant Pathol J 23:287–294Google Scholar
  14. 14.
    Phongpaichit S, Rungjindamai N, Rukachaisirikul V, Sakayaroj J (2006) Antimicrobial activity in cultures of endophytic fungi isolated from Garcinia species. FEMS Immunol Med Microbiol 48:367–372. doi: 10.1111/j.1574-695X.2006.00155.x PubMedCrossRefGoogle Scholar
  15. 15.
    Porras-Alfaro A, Herrera J, Sinsabaugh RL, Odenbach KJ, Lowrey T, Natvig DO (2008) Novel root fungal consortium associated with a dominant desert grass. Appl Environ Microbiol 74:2805–2813. doi: 10.1128/AEM.02769-07 PubMedCrossRefGoogle Scholar
  16. 16.
    Redman RS, Sheehan KB, Stout RG, Rodriquez RJ, Henson JM (2002) Thermotolerance generated by plant/fungal symbiosis. Science 298:1581. doi: 10.1126/science.1072191 PubMedCrossRefGoogle Scholar
  17. 17.
    Rubini MR, Silva-Ribeiro RT, Pomella AW, Maki CS, Araújo WL, Dos Santos DR, Azevedo JL (2005) Diversity of endophytic fungal community of cacao (Theobroma cacao L.) and biological control of Crinipellis perniciosa, causal agent of Witches’ Broom Disease. Int J Biol Sci 1:24–33PubMedGoogle Scholar
  18. 18.
    Saitou N, Nei M (1987) The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol Biol Evol 4:406–425PubMedGoogle Scholar
  19. 19.
    Stierle A, Strobei G, Stierle D (1993) Taxol and taxane production by Taxomyces andreanae, an endophytic fungus of pacific yew. Science 260:214–216. doi: 10.1126/science.8097061 PubMedCrossRefGoogle Scholar
  20. 20.
    Strobel G, Daisy B (2003) Bioprospecting for microbial endophytes and their natural products. Microbiol Mol Biol Rev 67:491–502. doi: 10.1128/MMBR.67.4.491-502.2003 PubMedCrossRefGoogle Scholar
  21. 21.
    Wang YT, Lo HS, Wang PH (2008) Endophytic fungi from Taxus mairei in Taiwan: first report of Colletotrichum gloeosporioides as an endophyte of Taxus mairei. Bot Stud (Taipei, Taiwan) 49:39–43Google Scholar
  22. 22.
    White TJ, Bruns T, Lee S, Taylor J (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 application. Academic Press Inc., San Diego, pp 315–322Google Scholar
  23. 23.
    Tan RX, Zou WX (2001) Endophytes: a rich source of functional metabolites. Nat Prod Rep 18:448–459. doi: 10.1039/b100918o PubMedCrossRefGoogle Scholar
  24. 24.
    Tejesvi M, Nahesh B, Nalini M, Prakash H, Kini K, Subbiah V, Shetty H (2005) Endophytic fungal assemblages from inner bark and twig of Terminalia arjuna W. & A. (Combr taceae). World J Microbiol Biotechnol 21:1535–1540. doi: 10.1007/s11274-005-7579-5 CrossRefGoogle Scholar
  25. 25.
    Thompson JD, Gibson TJ, Plewniak F, Jeanmougin F, Higgins DG (1997) The Clustal X windows interface: flexible strategies for multiple sequence alignment aided by quality analysis tools. Nucleic Acids Res 24:4876–4882. doi: 10.1093/nar/25.24.4876 CrossRefGoogle Scholar
  26. 26.
    Zhang CL, Liu SP, Lin FC, Kubicek CP, Druzhinina IS (2007) Trichoderma taxi sp. nov., an endophytic fungus from Chinese yew Taxus mairei. FEMS Microbiol Lett 270:90–96. doi: 10.1111/j.1574-6968.2007.00659.x PubMedCrossRefGoogle Scholar
  27. 27.
    Zhang D, Yang Y, Castlebury LA, Cerniglia CE (1996) A method for the large scale isolation of high transformation efficiency fungal genomic DNA. FEMS Microbiol Lett 145:261–265. doi: 10.1111/j.1574-6968.1996.tb08587.x PubMedCrossRefGoogle Scholar

Copyright information

© Society for Industrial Microbiology 2009

Authors and Affiliations

  • Kaihui Liu
    • 1
  • Xiaowei Ding
    • 1
  • Baiwan Deng
    • 1
    Email author
  • Wenqiang Chen
    • 1
  1. 1.School of Biological Sciences and EngineeringShaanxi University of TechnologyHanzhongPeople’s Republic of China

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