Mode of action of Trichoderma asperellum SKT-1, a biocontrol agent against Gibberella fujikuroi

Abstract

Trichoderma asperellum SKT-1 and Gibberella fujikuroi, known as causal agents of "Bakanae" disease, were both transformed with genes encoding green fluorescent protein (GFP) and hygromycin B (hygB) by restriction enzyme-mediated integration (REMI). Rice seeds inoculated with GFP-tagged G. fujikuroi showed "Bakanae" symptoms. GFP-tagged SKT-1 maintained biocontrol activity against the pathogen by soaking seeds in SKT-1 spore suspension. Then, we monitored in situ interactions between SKT-1 and G. fujikuroi on rice seeds using GFP-tagged transformations under confocal scanning laser stereomicroscopy. G. fujikuroi disappeared from the embryo of rice seeds after treatment with SKT-1, whereas SKT-1 was observed on the embryo 24 hr after initiation of germination. In addition, the hyphae of G. fujikuroi were penetrated by the hyphae of SKT-1, and degradation of the cell walls of G. fujikuroi was observed under SEM in co-culture. The cell wall of G. fujikuroi on the embryo of rice seeds was lysed, suggesting that mycoparasitism is the mode of action of T. asperellum SKT-1.

Use of fluorescent proteins to visualize interactions between the Bakanae disease pathogen Gibberella fujikuroi and the biocontrol agent Talaromyces sp. KNB-422

“…KNB-422 might sense some biological materials such as secondary metabolites from G. fujikuroi hyphae and grow toward the pathogen. Watanabe et al (2007) reported that co-culture of Trichoderma asperellum SKT-1, another biocontrol agent against Bakanae disease, with G. fujikuroi on PDA resulted in loss of GFP fluorescence in G. fujikuroi hyphae. Trichoderma asperellum acts parasitically toward the pathogen colonizing the mutual regions in rice seed embryos.…”

“…Plasmid pMK412 (Watanabe et al 2007) carried a GFP gene (egfp) driven by the translation elongation factor (TEF) promoter from Aureobasidium pullulans and the glucoamylase (gla) terminator from Aspergillus awamori, and the hygromycin B phosphotransferase gene (hph) under control of trifunctional polypeptide (trpC) promoter and terminator from Aspergillus nidulans. Plasmid pDsRed2 (Takara Bio, Ohtsu, Japan) carried dsred2.…”

“…were prepared and used for REMI (restriction enzyme-mediated integration) transformation as described by Watanabe et al (2007). Protoplasts of G. fujikuroi or Talaromyces sp.…”

“…Understanding cell-cell interactions between the pathogen and the biocontrol agent can offer clues to the molecular mechanism underlying biocontrol, which might then be relevant to practical application (Watanabe et al 2007). …”

A Eukaryotic Molecular Target Candidate of Roxithromycin: Fungal Differentiation as a Sensitive Drug Target Analysis System

“…The Ptef was amplified from pMK412 24) with the Ptef-F and Ptef-R primers. Ptef fragment (0.8 kb) was digested by EcoRV and ligated with EcoRV and HindIII digested pCSN43 blunted by the Klenow fragment of Escherichia coli DNA polymeraseI.…”