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Applied and Environmental Microbiology, July 2005, p. 3959-3965, Vol. 71, No. 7
0099-2240/05/$08.00+0     doi:10.1128/AEM.71.7.3959-3965.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Improvement of the Fungal Biocontrol Agent Trichoderma atroviride To Enhance both Antagonism and Induction of Plant Systemic Disease Resistance

Kurt Brunner,1 Susanne Zeilinger,1 Rosalia Ciliento,2 Sheridian L. Woo,2 Matteo Lorito,2 Christian P. Kubicek,1 and Robert L. Mach1*

Abteilung für Gentechnologie und Angewandte Biochemie, Institut für Verfahrenstechnik, Umwelttechnik und Technische Biowissenschaften, Technische Universität Wien, Getreidemarkt 9/166/6, A-1060 Vienna, Austria,1 Dipartimento di Arbricoltura, Botanica e Patologia Vegetale, Sezione di Patologia Vegetale, Università degli Studi di Napoli "Frederico II" and Centro di Studio CNR per le Tecniche di Lotta Biologica (CETELOBI), Via Università, 100, I-80050 Portici (NA), Italy2

Received 17 August 2004/ Accepted 6 January 2005

Biocontrol agents generally do not perform well enough under field conditions to compete with chemical fungicides. We determined whether transgenic strain SJ3-4 of Trichoderma atroviride, which expresses the Aspergillus niger glucose oxidase-encoding gene, goxA, under a homologous chitinase (nag1) promoter had increased capabilities as a fungal biocontrol agent. The transgenic strain differed only slightly from the wild-type in sporulation or the growth rate. goxA expression occurred immediately after contact with the plant pathogen, and the glucose oxidase formed was secreted. SJ3-4 had significantly less N-acetylglucosaminidase and endochitinase activities than its nontransformed parent. Glucose oxidase-containing culture filtrates exhibited threefold-greater inhibition of germination of spores of Botrytis cinerea. The transgenic strain also more quickly overgrew and lysed the plant pathogens Rhizoctonia solani and Pythium ultimum. In planta, SJ3-4 had no detectable improved effect against low inoculum levels of these pathogens. Beans planted in heavily infested soil and treated with conidia of the transgenic Trichoderma strain germinated, but beans treated with wild-type spores did not germinate. SJ3-4 also was more effective in inducing systemic resistance in plants. Beans with SJ3-4 root protection were highly resistant to leaf lesions caused by the foliar pathogen B. cinerea. This work demonstrates that heterologous genes driven by pathogen-inducible promoters can increase the biocontrol and systemic resistance-inducing properties of fungal biocontrol agents, such as Trichoderma spp., and that these microbes can be used as vectors to provide plants with useful molecules (e.g., glucose oxidase) that can increase their resistance to pathogens.

* Corresponding author. Mailing address: Abteilung für Gentechnologie und Angewandte Biochemie, Institut für Verfahrenstechnik, Umwelttechnik und Technische Biowissenschaften, TU Wien, Getreidemarkt 9/166/6, A-1060 Vienna, Austria. Phone: 43 1 58801 17251. Fax: 43 1 5816266. E-mail: rmach{at}mail.zserv.tuwien.ac.at.

Applied and Environmental Microbiology, July 2005, p. 3959-3965, Vol. 71, No. 7
0099-2240/05/$08.00+0     doi:10.1128/AEM.71.7.3959-3965.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.



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