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Molecular Biology

, Volume 52, Issue 5, pp 668–675 | Cite as

Transgenic Expression of chit42 gene from Metarhiziumanisopliae in Trichoderma harzianum Enhances Antagonistic Activity against Botrytis cinerea

  • H. Xia
  • Y. Y. Li
  • Z. C. Liu
  • Y. Q. Li
  • J. Chen
GENOMICS. TRANSCRIPTOMICS

Abstract

Сhitinases expressed by some beneficial fungi are crucial for the biocontrol of phytopathogens. The activity of chitinolytic strains of Trichoderma sp. may be enhanced by increasing the expression of chitinases. We describe the Trichoderma strain Mchit42 which expresses a transgenic chitinase chit42 from Metarhizium anisopliae. Inhibitory effects against plant pathogens were tested. Comparison of WT (T30) and OE (Mchit42) indicated that overexpression of M. anisopliaechit42 did not alter Trichoderma growth, while enhancing the expression of endogenous chitinase, β-1,3-glucanases, and polygalacturonase and increasing the antagonistic activity of Trichoderma against Botrytis cinerea. This work confirmed that the expression of the entomopathogenic fungi-sourced chit42 genes in Trichodermaharzianum enhances the efficiency of Trichoderma biocontrol against targeted pathogens.

Keywords:

chit42 gene Metarhizium anisopliae Trichoderma harzianum antagonistic activity Botrytis cinerea 

Notes

ACKONWLEDGMENTS

This work was supported by National Key Research and Development Program of China (2017YFD0200901 and 2017YFD0200400), National Natural Science Foundation of China (31672072) and Agricultural Extension Project of Shanghai Municipal Agricultural Commission (2017) no. 1-6. We thank the laboratory of Plant Pathology of Shanghai Jiaotong University for providing us Metarhizium anisopliae CY1 and Trichoderma harzianum T30.

REFERENCES

  1. 1.
    Steyaert J.M., Stewart A., Jaspers M., Carpenter M., Ridgway H.J. 2004. Co-expression of two genes, a chitinase (chit42) and proteinase (prb1), implicated in mycoparasitism by Trichoderma hamatum. Mycologia. 96, 1245–1252.CrossRefPubMedGoogle Scholar
  2. 2.
    Carrera E., Ruiz-Rivero O., Peres L.E., Atares A., Garcia-Martinez J.L. 2012. Characterization of the procera tomato mutant shows novel functions of the SlDELLA protein in the control of flower morphology, cell division and expansion, and the auxin-signaling pathway during fruit-set and development. Plant Physiol. 160, 1581–1596.CrossRefPubMedPubMedCentralGoogle Scholar
  3. 3.
    Chet I., Inbar J., Hadar I. 1997. Fungal antagonists and mycoparasites. In: The Mycota IV: Environmental and Microbial Relationships. Berlin: Springer-Verlag, 165–184.Google Scholar
  4. 4.
    Kowsari M., Motallebi M., Zamani R.M. 2014. Construction of new GFP-tagged fusants for Trichoderma harzianum with enhanced biocontrol activity. Plant Prot. Res. 54, 122−131.CrossRefGoogle Scholar
  5. 5.
    Saksirirat W., Chareerak P., Bunyatrachata W. 2009. Induced systemic resistance of biocontrol fungus, Trichoderma spp. against bacterial and gray leaf spot in tomatoes. As. J. Food Ag-Ind. 2, 99–104.Google Scholar
  6. 6.
    Bolar J.P., Norelli J.L., Harman G.E., Brown S.K., Aldwinckle H.S. 2001. Synergistic activity of endochitinase and exochitinase from Trichoderma atroviride (T. harzianum) against the pathogenic fungus (Venturia inaequalis) in transgenic apple plants. Transgenic Res. 10, 533–543.CrossRefPubMedGoogle Scholar
  7. 7.
    Grayston S.J. 1996. Rhizosphere carbon flow in trees, in comparison to annual plants: The importance of root exudation and its impact on microbial activity and nutrient cycling. Appl. Soil Ecol. 5, 29–56.CrossRefGoogle Scholar
  8. 8.
    Cumagun C.J.R. 2012. Managing plant diseases and promoting sustainability and productivity with Trichoderma. J. Agric Sci. Technol. 14, 699–714.Google Scholar
  9. 9.
    Vinale F., Sivasithamparam K., Ghisalberti E.L, Marra R., Woo S.L, Lorito M. 2008. Trichoderma–plant–pathogen interactions. Soil Biol. Biochem. 40, 1−10.CrossRefGoogle Scholar
  10. 10.
    Harman G.E, Howell C.R, Viterbo A., Chet I., Lorito M. 2004. Trichoderma species: Opportunistic, avirulent plant symbionts. Nat. Rev. 2, 43–56.Google Scholar
  11. 11.
    Lopez R.C, Gomez-Gomez L. 2009. Isolation of a new fungi and wound-induced chitinase class in corms of Crocus sativus plant. Physiol. Biochem. 47, 426–434.Google Scholar
  12. 12.
    Collinge D.B., Kragh K.M., Mikkelson J.D., Nielson K.K., Rasmussen U., Vad K. 1993. Plant chitinases. Plant J. 3, 31–40.CrossRefPubMedGoogle Scholar
  13. 13.
    Gokul B., Lee J.H., Song K.B., Rhee S.K., Kim C.H., Panda T. 2000. Characterization and applications of chitinases from Trichoderma harzianum. Bioprocess Eng. 23, 691–694.CrossRefGoogle Scholar
  14. 14.
    Harighi M.J., Zamani M.R., Motallebi M. 2007. Evaluation of antifungal activity of purified chitinase 42 from Trichoderma atroviride PTCC5220. Biotechnology. 6, 28–33.CrossRefGoogle Scholar
  15. 15.
    Lee S.Y., Tindwa H., Lee Y.S., Naing K.W., Hong S.H., Nam Y., Kim K.Y. 2012. Biocontrol of anthracnose in pepper using chitinase, beta-1,3-glucanase, and 2-furancarboxaldehyde produced by Streptomyces cavourensis SY224. J. Microbiol. Biotechnol. 22, 1359–1366.CrossRefPubMedGoogle Scholar
  16. 16.
    Haggag W.M., Abdallh E.G. 2012. Purification and characterization of chitinase produced by endophytic Streptomyces hygroscopicus against some phytopathogens. J. Microbiol. Res. 2, 145–151.CrossRefGoogle Scholar
  17. 17.
    Limón M.C., Chacón M.R., Mejías R., Delgado-Jarana J., Rincón A.M., Codón A.C., Benítez T. 2004. Increased antifungal and chitinase specific activities of Trichoderma harzianum CECT 2413 by addition of a cellulose binding domain. Appl. Microbiol. Biotechnol. 64, 675–685.CrossRefPubMedGoogle Scholar
  18. 18.
    Amin F., Razdan V.K., Mohiddin F.A., Bhat K.A., Banday S. 2010. Potential of Trichoderma species as biocontrol agents of soil borne fungal propagules. J. Phytol. 2, 38–41.Google Scholar
  19. 19.
    Harighi M.J., Motallebi M., Zamani M.R. 2006. Antifungal activity of heterologous expressed chitinase 42 (Chit42) from Trichoderma atroviride PTCC5220. Iran J. Biotech. 4, 95–103.Google Scholar
  20. 20.
    Steyaert J.M., Ridgway H.J., Elad Y. 2003. Genetic basis of mycoparasitism: A mechanism of biological control by species of Trichoderma. New Zeal. J. Crop Hort. Sci. 31, 281–291.CrossRefGoogle Scholar
  21. 21.
    da Silva M.V., Santi L., Staats C.C., da Costa A.M., Colodel E.M., Driemeier D., Vainstein M.H., Schrank A. 2005. Cuticle-induced endo/exoacting chitinase CHIT30 from Metarhizium anisopliae is encoded by an ortholog of the chi3 gene. Res. Microbiol. 156, 382–392.CrossRefPubMedGoogle Scholar
  22. 22.
    Bergemann S.E., Miller S.L., Garbelotto M. 2005. Microsatellite loci from Russula brevipes, a common ectomycorrhizal associate of several tree species in North America. Mol. Ecol. Notes. 5, 472–474.CrossRefGoogle Scholar
  23. 23.
    García I., Lora J.M., de la Cruz J., Benítez T., Llobell A., Pintor-Toro J.A. 1994. Cloning and characterization of a chitinase (CHIT42) cDNA from the mycoparasitic fungus Trichoderma harzianum. Curr. Genet. 27, 83–89.CrossRefPubMedGoogle Scholar
  24. 24.
    Gu J., Ye C. 2011. pYEMF, a pUC18-derived XcmI T‑vector for efficient cloning of PCR products. Mol. Biotechnol. 47, 229–233.CrossRefPubMedGoogle Scholar
  25. 25.
    Kirubakaran S.I., Sakthivel N. 2007. Cloning and overexpression of antifungal barley chitinase gene in Escherichia coli. Protein Expr. Purif. 52, 159–166.CrossRefPubMedGoogle Scholar
  26. 26.
    Mullins E.D., Chen X., Romaine P., Raina R., Geiser D.M., Kang S. 2001. Agrobacterium-mediated transformation of Fusarium oxysporum: An efficient tool for insertional mutagenesis and gene transfer. Phytopathology. 91, 173–180.CrossRefPubMedGoogle Scholar
  27. 27.
    Reissig J., Strominger J., Leloir L. 1955. A modified colorimetric method for the estimation of N-acetylamino sugars. J. Biol. Chem. 217, 959–966.PubMedGoogle Scholar
  28. 28.
    Bonev B., Hooper J., Parisot J. 2008. Principles of assessing bacterial susceptibility to antibiotics using the agar diffusion method. J. Antimicrob. Chemother. 61, 1295–1301.CrossRefPubMedGoogle Scholar
  29. 29.
    Frazzon A.P.G., Vaz I.S., Masuda A., Schrank A., Vainstein M.H. 2000. In vitro assessment of Metarhiaium anisopliae to control the tick Boophilus microplus. Vet. Parasitol. 94, 117–125.CrossRefPubMedGoogle Scholar
  30. 30.
    Keller N.P., Turner G., Bennett J.W. 2005. Fungal secondary metabolism—from biochemistry to genomics. Nat. Rev. Microbiol. 3, 937–947.CrossRefPubMedGoogle Scholar
  31. 31.
    Yu J.H., Keller N. 2005. Regulation of secondary metabolism in filamentous fungi. Annu. Rev. Phytopathol. 43, 437–458.CrossRefPubMedGoogle Scholar

Copyright information

© Pleiades Publishing, Inc. 2018

Authors and Affiliations

  • H. Xia
    • 1
    • 2
  • Y. Y. Li
    • 1
    • 2
  • Z. C. Liu
    • 1
    • 2
  • Y. Q. Li
    • 1
    • 2
  • J. Chen
    • 1
    • 2
    • 3
  1. 1.School of Agriculture and Biology, Shanghai Jiaotong UniversityShanghaiChina
  2. 2.The key Laboratory of Urban Agriculture (South), Ministry of AgricultureShanghaiChina
  3. 3.State Key Laboratory of Microbial Metabolism, Shanghai Jiaotong UniversityShanghaiChina

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