[Development of an Agrobacterium tumefaciens-mediated genetic transformation system in aconidial Trichoderma taxi]

Trichodermin, a sesquiterpenoid compound with antifungal, plant growth-regulating, and antitumor activities, demonstrates broad application prospects in both pharmaceutical and agricultural fields. Trichoderma taxi is an efficient trichodermin-producing filamentous fungus isolated from Taxus mairei, while its unique aconidial phenotype poses challenges to genetic transformation. To overcome this barrier and enable targeted engineering, this study established an Agrobacterium tumefaciens-mediated transformation (ATMT) system suitable for aconidial T. taxi, building on proven expertise in genetic transformation of sporulating filamentous fungi. Through systematic optimization of key parameters including mycelium pretreatment (mechanical grinding method), infection time, and co-culture conditions, we successfully achieved efficient transformation of T. taxi by A. tumefaciens. Utilizing this technical platform, we targeted tri3 (a gene encoding acetyltransferase, a key enzyme in trichodermin biosynthesis) to construct tri3-overexpressing strains. The engineered strains exhibited trichodermin production reaching 1.17 g/L, representing a 33.3% increase compared with that of the parent strain. The established transformation system not only provides a reliable technical approach for metabolic pathway engineering and high-yielding strain development of T. taxi, but also opens new avenues for genetic manipulation of other aconidial filamentous fungi in industrial applications.