关键词: 油茶, 果生炭疽菌, O-甘露糖基转移酶CfPmt2, 致病力

Abstract:

Objective: As a member of the O-mannosyltransferase family, Pmt2 is a key factor regulating fungal growth, development, and pathogenicity. This study aims to investigate the biological function of CfPmt2 in Colletotrichum fructicola (the dominant pathogen of Camellia oleifera anthracnose), thereby laying a theoretical foundation for deciphering the regulatory roles of O-mannosyltransferases during the pathogenesis of C. fructicola anthracnose and providing valuable references for the management of this disease. Method: A CfPMT2 gene knockout vector was constructed based on the principle of homologous recombination. The CfPMT2 deletion mutant (ΔCfpmt2) and complemented strain (ΔCfpmt2/PMT2) were successfully obtained by PEG-mediated protoplast transformation. Key biological phenotypes, such as vegetative growth, conidiation and appressorium formation, adhesion ability, sensitivity to cell wall and oxidative stresses, mycelial growth and development, and pathogenicity, were assayed for the wild-type (WT), mutant and complemented strains. Result: 1) Compared with the WT strain, the mycelial growth rate of ΔCfpmt2 on PDA, CM, and MM media decreased by 69.23%, 74.77%, and 81.55%, respectively. The conidia yield, germination rate, and appressorium formation rate were reduced by approximately 96%, 17%, and 30%, respectively, while the appressorium malformation rate increased to 56%. The morphogenesis of mycelia, conidia, and appressoria was impaired, and the mutant’s adhesion ability was significantly decreased to only 46.12% and 50.84% of that of the WT. 2) Quantitative real-time PCR (qRT-PCR) analysis showed that, compared with the WT strain, the relative expression levels of adhesion-related genes GAA1, GPI8, and MSB2 in the mutant were downregulated by 31.19%, 30.81%, and 25.36%, respectively. 3) The growth inhibition rate of ΔCfpmt2 on PDA media supplemented with cell wall or oxidative stressors was significantly higher than that of the WT, and the homeostasis of chitin and reactive oxygen species (ROS) in hyphae, conidia, and appressoria was disrupted. 4) Pathogenicity assays revealed that the mutant’s virulence to both wounded and unwounded leaves was significantly reduced, with lesion areas accounting for only 11.12% and 19.2% of those in the WT, respectively. The complementation strain ΔCfpmt2/PMT2 restored the defects in growth, development, and pathogenicity, consistent with the WT. Conclusion: CfPmt2 is involved in regulating the formation and development of hyphae, conidia and appressoria, as well as adhesion and chitin metabolism, ROS homeostasis, stress response, and pathogenicity of C. fructicola.

Key words: Camellia oleifera, Colletotrichum fructicola, O-mannosyltransferase CfPmt2, pathogenicity