Biocontrol Potential of Forest Tree Endophytes

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Expression of cmg1, an Exo-β-1,3-Glucanase Gene from Coniothyrium minitans, Increases during Sclerotial Parasitism

During sclerotial infection of Sclerotinia sclerotiorum the mycoparasite Coniothyrium minitans penetrates through the host cell wall, which contains β-1,3-glucan as its major component. A PCR-based strategy was used to clone a β-1,3-glucanase-encoding gene, designated cmg1, from a cDNA library of the fungus. The nucleotide and deduced amino acid sequences of this gene showed high levels of similarity to the sequences of other fungal exo-β-1,3-glucanase genes. The calculated molecular mass of the deduced protein (without the predicted 24-amino-acid N-terminal secretion signal peptide) was 83,346 Da, and the estimated pI was 4.73. Saccharomyces cerevisiae INVSc1 expressing the cmg1 gene secreted a ∼100-kDa β-1,3-glucanase enzyme (as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis) into the culture medium. N-terminal sequence analysis of the purified recombinant enzyme revealed that the secreted enzyme starts at Ala-32, seven amino acids downstream from the predicted signal peptidase cleavage site. The purified recombinant glucanase inhibited in vitro mycelial growth of S. sclerotiorum by 35 and 85% at concentrations of 300 and 600 μg ml(−1), respectively. A single copy of the cmg1 gene is present in the genome of C. minitans. Northern analyses indicated increases in the transcript levels of cmg1 due to both carbon starvation and the presence of ground sclerotia of S. sclerotiorum; only slight repression was observed in the presence of 2% glucose. Expression of cmg1 increased during parasitic interaction with S. sclerotiorum

Use of Coniothyrium minitans as a biocontrol agent and some molecular aspects of sclerotial mycoparasitism

The use of the sclerotial mycoparasite Coniothyrium minitans as a biological control agent of diseases caused by sclerotium-forming pathogens especially Sclerotinia sclerotiorum is briefly reviewed. A number of studies have examined production and application methods, integrated control, ecology, and modes of action in order to understand the biology of the mycoparasite and enhance activity and reproducibility of use. Recently, development of a number of molecular-based techniques has begun to allow the examination of genes involved in mycoparasitism. Some of these procedures have been applied to identify pathogenicity genes involved in the infection of sclerotia of S. sclerotiorum by C. minitans and this work is discussed