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Keywords:

  • antagonistic Fusarium oxysporum;
  • biological control;
  • genetic transformation;
  • hygromycin B resistance;
  • mitotic stability;
  • Fusarium wilt of carnation

Abstract

The stability of a genetically modified strain of Fusarium oxysporum used as antagonist against phytopathogenic formae speciales of F. oxysporum was evaluated both in vitro and in microcosm assays. The Escherichia coli hygromycin B phosphotransferase gene (hph), conferring hygromycin B resistance, was introduced by genetic transformation into a recipient strain marked by benomyl resistance and a dark red pigmentation. Hybridization with the complete plasmid suggested that the integration had generally occurred in a multiple-tandem array at multiple sites. Among nine independent transformants tested, only three of them were mitotically stable after four rounds of vegetative growth with no selective pressure, while six showed various changes in the integration pattern. One transformant had lost the ability to grow in the presence of hygromycin B. In soil microcosms all the transformants maintained the hygromycin B resistant phenotype, but six of them showed rearrangement of transforming DNA. Only one strain (coded T26.40) underwent no obvious rearrangement both after in vitro growth and after recovery from the soil microcosm. The nine transformants were used in three biological control experiments against Fusarium wilt of carnation in comparison to two untransformed reference strains and to the recipient mutant. A high degree of variability in the biocontrol activity was observed throughout the experiments and only transformant T26.40 consistently controlled the incidence of disease. The results are discussed in relation to risk assessment of the release of transgenic antagonistic fungi.