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Overexpression of an Endochitinase Gene (ThEn-42) in Trichoderma atroviride for Increased Production of Antifungal Enzymes and Enhanced Antagonist Action Against Pathogenic Fungi

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Abstract

Trichoderma is one of the most promising biocontrol agents against plant fungal diseases. In this study, a transgenic strain of Trichoderma atroviride was characterized. The transgenic strain contains an endochitinase gene (ThEn-42) driven by the cellulase promoter cbh1 of T. reesei for overexpression of ThEn-42. The culture filtrates of the transformant and the parental strain grown in eight different media were evaluated for chitinase and antifungal enzyme production based on activity gels, protein profiles, and antifungal activities. Results demonstrated that chitinases are important components and synergistic interactions play a key role in the antagonistic action of T. atroviride. Moreover, altering medium nutrient concentration and composition led to enhanced production of antifungal enzymes, a potential strategy for mass production. Two of the culture filtrates contained almost pure endochitinase, and could be excellent commercial sources for this enzyme. Several culture filtrates were highly antifungal. Two filtrates were so effective in biocontrol of a fungal pathogen, Penicillium digitatum, that they not only inhibited spore germination but destroyed the spores completely when 20 μl of culture filtrate (corresponding to approximately 104 μg of total protein) was applied in a total volume of 150 μl (approximately 0.7 mg protein ml−1).

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Abbreviations

PAGE:

Polyacrylamide gel electrophoresis

SSC:

Sodium chloride–sodium citrate

SDS:

sodium dodecyl sulfate

PDB:

potato dextrose broth

References

  1. Chet, I. (1987). In I. Chet (Ed.), Innovative approaches to plant disease control (pp. 137–160). New York: John Wiley & Sons.

  2. Haran, S., Schickler, H., Oppenheim, A., & Chet, I. (1995). Mycological Research, 99, 441–446.

    CAS  Google Scholar 

  3. Lorito, M., Di Pietro, A., Hayes, C. K., Woo, S. L., & Harman, G. E. (1993). Phytopathology, 83, 721–728.

    Article  CAS  Google Scholar 

  4. Lorito, M., Harman, G. E., Hayes, C. K., Broadway, R. M., Tronsmo, A., Woo, S. L., et al. (1993). Phytopathology, 83, 302–307.

    Article  CAS  Google Scholar 

  5. Lorito, M., Hayes, C. K., Di Pietro, A., & Harman, G. E. (1993). Current Genetics, 24, 349–356.

    Article  CAS  Google Scholar 

  6. Bartnicki-Garcia, S. (1968). Annual Review of Microbiology, 22, 87–108.

    Article  CAS  Google Scholar 

  7. Harman, G. E., Hayes, C. K., Lorito, M., Broadway, R. M., Di Pietro, A., Peterbauer, C., et al. (1993). Phytopathology, 83, 313–318.

    Article  CAS  Google Scholar 

  8. Hayes, C. K., Klemsdal, S., Lorito, M., Di Pietro, A., Peterbauer, C., Nakas, J. P., et al. (1994). Gene, 138, 143–148.

    Article  CAS  Google Scholar 

  9. De la Cruz, J., Hidalgo-Gallego, A., Lora, J. M., Benitez, T., Pintor-Toro, J. A., & Llobell, A. (1992). European Journal of Biochemistry, 206, 859–867.

    Article  Google Scholar 

  10. Ulhoa, C. J., & Peberdy, J. F. (1991). Current Microbiology, 23, 285–289.

    Article  CAS  Google Scholar 

  11. Lorito, M., Hayes, C. K., Di Pietro, A., Woo, S. L., & Harman, G. E. (1994). Phytopathology, 84, 398–405.

    Article  CAS  Google Scholar 

  12. Ali, G. S., Harman, G. E., & Reisch, B. I. (2003). European Journal of Plant Pathology, 109, 639–644.

    Article  CAS  Google Scholar 

  13. Kleman-Leyer, K. M., Siika-aho, M., Teeri, T. T., & Kirk, T. K. (1996). Applied and Environmental Microbiology, 62, 2883–2887.

    CAS  Google Scholar 

  14. Lorito, M., Peterbauer, C., Hayes, C. K., & Harman, G. E. (1994). Microbiology, 140, 623–629.

    CAS  Google Scholar 

  15. Schirmböck, M., Lorito, M., Wang, Y-L., Hayes, C. K., Arisan-Atac, I., Scala, F., et al. (1994). Applied and Environmental Microbiology, 60, 4364–4370.

    Google Scholar 

  16. Limón, M. C., Chacón, M. R., Mejías, R., Delgado-Jarana, J., Rincón, A. M., Codón, A. C., et al. (2004). Applied Microbiology and Biotechnology, 64, 675–685.

    Article  CAS  Google Scholar 

  17. Sandhya, C., Adapa, L. K., Nampoothiri, K. M., Binod, P., Szakacs, G., & Pandey, A. (2004). Journal of Basic Microbiology, 44, 49–58.

    Article  CAS  Google Scholar 

  18. Margolles-Clark, E., Hayes, C. K., Harman, G. E., & Penttilä, M. (1996). Applied and Environmental Microbiology, 62, 2145–2151.

    CAS  Google Scholar 

  19. Margolles-Clark, E., Harman, G. E., & Pentillä, M. (1996). Applied and Environmental Microbiology, 62, 2152–2155.

    CAS  Google Scholar 

  20. Tronsmo, A. (1991). Biological Control, 1, 59–62.

    Article  Google Scholar 

  21. Tronsmo, A., & Harman, G. E. (1993). Analytical Biochemistry, 208, 74–79.

    Article  CAS  Google Scholar 

  22. Ohtakara, A. (1988) In W. A. Wood & S. T. Kellogg (Eds.), Methods in enzymology, (vol. 161, pp. 462–470). San Diego: Academic Press.

  23. Roberts, E. K., & Selitrennikoff, C. P. (1988). Journal of General Microbiology, 134, 169–176.

    CAS  Google Scholar 

  24. Vessey, J. C., & Pegg, G. F. (1973). Transactions of the British Mycological Society, 60, 133–143.

    Article  CAS  Google Scholar 

  25. Blum, H., Beier, H., & Gross, H. J. (1987). Electrophoresis, 8, 93–99.

    Article  CAS  Google Scholar 

  26. Carsolio, C., Gutierrez, A., Jimenez, B., Van Montagu, M., & Herrera-Estrella, A. (1994). Proceedings of the National Academy of Sciences of the United States of America, 91, 10903–10907.

    Article  CAS  Google Scholar 

  27. Di Pietro, A., Lorito, M., Hayes, C. K., Broadway, R. M., & Harman, G. E. (1993). Phytopathology, 83, 308–313.

    Article  Google Scholar 

  28. Peterbauer, C., Lorito, M., Hayes, C. K., Harman, G. E., & Kubicek, C. P. (1996). Current Genetics, 30, 325–331.

    Article  CAS  Google Scholar 

  29. Donzelli, B. G. G., & Harman, G. E. (2001). Applied and Environmental Microbiology, 67, 5643–5647.

    Article  CAS  Google Scholar 

  30. Brunner, K., Peterbauer, C. K., Mach, R. L., Lorito, M., Zeilinger, S., & Kubicek, C. P. (2003). Current Genetics, 43, 289–295.

    Article  CAS  Google Scholar 

  31. Stasz, T. E., Harman, G. E., & Weeden, N. E. (1988). Mycologia, 80, 141–150.

    Article  Google Scholar 

  32. Lorito, M. (1998). In G. E. Harman & C. P. Kubicek (Eds.), Trichoderma and gliocladium (vol. 2, pp. 73–99). London, United Kingdom: Taylor & Francis.

  33. Donzelli, B. D. D., Seibert, K. J., & Harman, G. E. (2005). Enzyme and Microbial Technology, 38, 1823–1833.

    Google Scholar 

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Acknowledgments

The authors would like to thank Seur Kee Park for technical assistance, Thomas Björkman for assistance with microscopy, Jyothi P. Bolar for providing the nag1 primers, and Kristen L. Ondik for her editorial assistance.

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Authors and Affiliations

  1. Department of Plant and Soil Sciences, Oklahoma State University, Stillwater, OK, 74078, USA

    Shiping Deng

  2. Instituto di Patologia Vegetale, Universita degli Studi di Napoli “Federico II” e Centro CNR per lo Studio delle Techniche di Lotta Biologica, 80055, Portici, Naples, Italy

    Matteo Lorito

  3. VTT Biotechnology and Food Research, FIN-02044 VTT, Espoo, Finland

    Merja Penttilä

  4. Departments of Horticultural Sciences and Plant Pathology, Cornell University, Geneva, New York, 14456, USA

    Gary E. Harman

Authors
  1. Shiping Deng
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  2. Matteo Lorito
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  3. Merja Penttilä
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  4. Gary E. Harman
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Correspondence to Shiping Deng.

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Deng, S., Lorito, M., Penttilä, M. et al. Overexpression of an Endochitinase Gene (ThEn-42) in Trichoderma atroviride for Increased Production of Antifungal Enzymes and Enhanced Antagonist Action Against Pathogenic Fungi. Appl Biochem Biotechnol 142, 81–94 (2007). https://doi.org/10.1007/s12010-007-0012-9

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  • DOI: https://doi.org/10.1007/s12010-007-0012-9

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