Chlorogenic Acid From <i>Pseudopestalotiopsis camelliae‐sinensis</i> , Regulated by <scp> <i>PscC3H</i> </scp> , Inhibits the Growth of Tea Anthracnose Pathogen <i>Colletotrichum camelliae</i>

ABSTRACT Tea anthracnose, caused predominantly by Colletotrichum camelliae , is a major disease affecting the tea plant ( Camellia sinensis ), with no effective, environmentally friendly control measures. Pseudopestalotiopsis camelliae‐sinensis produces multiple bioactive secondary metabolites. A culture of P. camelliae‐sinensis in 20% tea broth (TB) medium for 20 days (20% TB‐ Psc ) significantly inhibited the vegetative growth of C. camelliae . Untargeted metabolomics analysis indicated a marked upregulation of secondary metabolites in 20% TB‐ Psc compared to a control broth inoculated with sterile potato dextrose agar (20% TB‐ck). Among these metabolites, chlorogenic acid (CGA) was significantly upregulated in 20% TB‐ Psc and significantly inhibited the mycelial growth of C. camelliae in vitro. A key gene named PscC3H , encoding a cytochrome P450 monooxygenase, was identified as a potential contributor to CGA biosynthesis in P. camelliae‐sinensis . To confirm its role in CGA biosynthesis, we established a polyethylene glycol (PEG)‐mediated protoplast transformation system for P. camelliae‐sinensis and generated the gene knock‐out mutant ∆ PscC3H through homologous recombination. Deletion of PscC3H did not affect the vegetative growth, conidiation or pathogenicity of P. camelliae‐sinensis on detached tea leaves. However, CGA content and antifungal activity in 20% TB inoculated with ∆ PscC3H (20% TB‐ Psc ∆) was significantly lower than in 20% TB‐ Psc . Collectively, these results demonstrate that CGA produced by P. camelliae‐sinensis plays a significant role in conferring resistance against C. camelliae , and PscC3H plays a crucial role in CGA biosynthesis in P. camelliae‐sinensis , but not by altering its growth or pathogenicity.