Botrytis cinerea is a necrotrophic model fungal plant pathogen that causes grey mould, a devastating disease responsible for large losses in the agriculture sector. As important targets of fungicides, membrane proteins are hot spots in the research and development of fungicide products. We previously found that membrane protein Bcest may be closely related to the pathogenicity of Botrytis cinerea. Herein, we further explored its function. We generated and characterised ΔBcest deletion mutants of B. cinerea and constructed complemented strains. The ΔBcest deletion mutants exhibited reduced conidia germination and germ tube elongation. The functional activity of ΔBcest deletion mutants was investigated by reduced necrotic colonisation of B. cinerea on grapevine fruits and leaves. Targeted deletion of Bcest also blocked several phenotypic defects in aspects of mycelial growth, conidiation and virulence. All phenotypic defects were restored by targeted-gene complementation. The role of Bcest in pathogenicity was also supported by reverse-transcriptase real-time quantitative PCR results indicating that melanin synthesis gene Bcpks13 and virulence factor Bccdc14 were significantly downregulated in the early infection stage of the ΔBcest strain. Taken together, these results suggest that Bcest plays important roles in the regulation of various cellular processes in B. cinerea.

Grey mould caused by Botrytis cinerea is a devastating disease responsible for large losses to agricultural production, and B. cinerea is a necrotrophic model fungal plant pathogen. Membrane proteins are important targets of fungicides and hotspots in the research and development of fungicide products. Wuyiencin affects the permeability and pathogenicity of B. cinerea, parallel reaction monitoring revealed the association of membrane protein Bcsdr2, and the bacteriostatic mechanism of wuyiencin was elucidated. In the present work, we generated and characterised ΔBcsdr2 deletion and complemented mutant B. cinerea strains. The ΔBcsdr2 deletion mutants exhibited biofilm loss and dissolution, and their functional activity was illustrated by reduced necrotic colonisation on strawberry and grape fruits. Targeted deletion of Bcsdr2 also blocked several phenotypic defects in aspects of mycelial growth, conidiation and virulence. All phenotypic defects were restored by targeted gene complementation. The roles of Bcsdr2 in biofilms and pathogenicity were also supported by quantitative real-time RT-PCR results showing that phosphatidylserine decarboxylase synthesis gene Bcpsd and chitin synthase gene BcCHSV II were downregulated in the early stages of infection for the ΔBcsdr2 strain. The results suggest that Bcsdr2 plays important roles in regulating various cellular processes in B. cinerea. Key points: • The mechanism of wuyiencin inhibits B. cinerea is closely associated with membrane proteins. • Wuyiencin can downregulate the expression of the membrane protein Bcsdr2 in B. cinerea. • Bcsdr2 is involved in regulating B. cinerea virulence, growth and development.