Novel Endophytic Fungi from Euchresta tubulosa Dunn: Characterization of Their Bioactive Secondary Metabolites and Extracellular Enzymes

The endangered ethnomedicinal plant Euchresta tubulosa harbors a valuable community of endophytic fungi, demonstrating significant potential for biotechnological applications. Endophytic fungi were isolated from E. tubulosa and identified to characterize their secondary metabolites and extracellular enzyme activities. Endophytic fungi were isolated from E. tubulosa using tissue explant culture and identified by morphological and molecular (ITS) analysis. The chemical profiles of strain fermentation products were analyzed by LC–MS/MS, while extracellular enzyme production (cellulase, protease, amylase) was assessed through chromogenic plate assays and liquid fermentation. The results indicated that a total of 55 endophytic fungi were isolated from E. tubulosa, assigned to 17 genera. Among these, three genera (Colletotrichum, Fusarium, and Penicillium) constituted the dominant groups, while four strains (including three novel species) represented potential new taxa. LC–MS/MS analysis revealed that fermentation products of the three novel endophytic fungal species contained bioactive compounds such as flavonoids and alkaloids; furthermore, bioactivity assays indicate that they exhibited significant degrees of antibacterial and antioxidant activity. Extracellular enzyme assays demonstrated that three E. tubulosa-derived endophytic strains exhibited multi-hydrolytic enzyme production capabilities. Notably, strain ETG-1-2-1 showed the highest amylase and cellulase activities (10.95 U/mL and 9.68 U/mL, respectively), while strain ETXG-1-1-1 displayed the highest protease activity (2.34 U/mL). This study provides the first systematic report on the diversity of endophytic fungi in E. tubulosa, their secondary metabolite profiles, and extracellular enzyme activities, establishing a theoretical foundation for discovering novel bioactive compounds and developing microbial resources, while also highlighting their ecological roles and biotechnological potential.