Effect of CFEM proteins on pathogenicity, patulin accumulation and host immunity of postharvest apple pathogens Penicillium expansum

Penicillium expansum is a significant post-harvest pathogenic fungi on most pome fruits. Common fungal extracellular membrane (CFEM) proteins, as effectors, contribute to virulence and manipulate host immunity. However, the CFEM proteins in P. expansum have not been identified and functionally studied. In this study, we screened two P. expansum CFEM proteins, PeCFEM5 and PeCFEM8, whose expression was highly up-regulated during postharvest apple infection. Growth and pathogenicity of P. expansum were characterized by knockout and complementary of PeCFEM5 and PeCFEM8. Deletion of PeCFEM5 and PeCFEM8 resulted in changes in spore development and increased resistance to cell wall integrity stress. The lesion spots on apple and pear fruit inoculated with P. expansum gradually expanded and deepened in color. The ΔPeCFEM5 and ΔPeCFEM8 strains reduced lesion diameter on apple fruit by 47 % and 29 %, respectively, compared with the WT strains. Detection of patulin accumulation by high-performance liquid chromatography (HPLC) revealed that deletion of PeCFEM5 or PeCFEM8 suppressed patulin content in medium and apples, and patulin biosynthesis-related genes were down-regulated. The PeCFEM5 and PeCFEM8 were also confirmed as effector proteins capable of suppressing the cell death triggered by BAX and the expression of plant defense genes in Nicotiana benthamiana. Phytohormone ELISA assays showed that jasmonic acid levels were reduced, but salicylic acid levels were increased by transient expression of PeCFEM5 or PeCFEM8 in the host plant. These results indicate that PeCFEM5 and PeCFEM8 effectors are crucial for pathogenicity, patulin biogenesis, and modulating host plant immunity.