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Providing a toolbox for genomic engineering of Trichoderma aggressivum

Summary

Scientists have developed a set of techniques to genetically modify the fungus Trichoderma aggressivum, which is usually known for ruining mushroom crops. This genetic toolkit allows researchers to edit genes in this fungus to study how it produces various compounds and why it affects mushrooms. By using modern gene-editing technology called CRISPR, researchers can now create specific mutations and study the fungus’s useful properties, such as its potential to protect crops or promote plant growth.

Background

Trichoderma aggressivum is a fungus known primarily as a causal agent of green mold disease in cultivated mushrooms (Agaricus bisporus), causing significant crop losses. However, T. aggressivum also has potential applications as a biocontrol agent and plant growth promoter. Understanding the fundamental biology and secondary metabolites of T. aggressivum requires efficient genetic manipulation tools.

Objective

To develop and establish a comprehensive genetic engineering toolkit for T. aggressivum, including transformation protocols using plasmid DNA and CRISPR-Cas9-based genome editing. The study aims to validate selection markers and demonstrate gene disruption capabilities for future research.

Results

Transformation efficiency with the hph gene as selection marker was 26.63 CFU/µg DNA. Four candidate strains with disrupted pyr4 genes were obtained, resulting in uridine auxotrophy phenotype. The pyr4 gene from T. reesei successfully complemented the deletion and functioned as an effective selection marker in the auxotrophic strain.

Conclusion

A practical and reproducible genetic engineering toolkit has been established for T. aggressivum, including validated transformation protocols and selection markers. These tools enable future investigations of biosynthetic gene clusters, secondary metabolite production, and the fundamental biology of mycoparasitism in this fungus.
  • Published in:Microbiology Spectrum,
  • Study Type:Technical/Methods Paper,
  • Source: 10.1128/spectrum.00966-25, PMID: 40744866
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