The Boeremia exigua BeAA9 Lytic Polysaccharide Monooxygenase Is a Key Virulence Factor in Mulberry Infection.

Abstract

Boeremia exigua, a phytopathogen responsible for spot diseases on leaves and stems, significantly threatens the health of a wide range of plants. Understanding its pathogenic mechanisms is essential for effective mulberry disease control. In this study, B. exigua GXH1 was isolated from mulberry leaves exhibiting symptoms of spot disease. Infection of mulberry seedlings by B. exigua led to significantly increased levels of defense enzyme activities (superoxide dismutase, peroxidase, catalase, phenylalanine ammonia-lyase), indicating that B. exigua triggers a strong immune response in the host. B. exigua infection in mulberry leaves resulted in abundant hyphae and vesicular structures in the intercellular spaces and epidermal layers. Whole-genome sequencing of B. exigua unveiled a 34.33 Mb genome containing 12,060 coding genes, including a notable abundance of carbohydrate-active enzymes. Transcriptome analysis during mulberry infection revealed 509 upregulated and 335 downregulated genes, with a particular enrichment in genes related to carbohydrate metabolism and redox processes. Notably, the lytic polysaccharide monooxygenase AUXILIARY ACTIVITY FAMILY 9 gene (BeAA9), which is localized to the plasma membrane, was highly upregulated in mulberry leaves infested by B. exigua. Knockout of BeAA9 led to a significant reduction in the pathogenicity of B. exigua. Furthermore, BeAA9 transient expression in Nicotiana benthamiana suppressed BCL2 Associated X Protein (BAX)-induced cell death and reactive oxygen species bursts, while its overexpression diminished plant resistance to Botrytis cinerea and downregulated the expression of plant immune genes. These findings identify BeAA9 as a key virulence factor in B. exigua GXH1, shedding light on its role in suppressing plant immunity.