Galactosaminogalactan orchestrates Verticillium dahliae virulence and rhizosphere microbial ecology through multi-partite interactions.

The infection of plants by soil-borne fungal pathogens is a complex process and depends on their adhere ability to host surfaces and interactions with the rhizosphere bacteria. In this study, we identify galactosaminogalactan (GAG) as a pivotal virulence determinant in the pathogenesis of Verticillium dahliae. By characterizing the VdGAG biosynthetic gene cluster, we show that the glycosyltransferase VdGtb is essential for GAG synthesis. The knockout of GAG increased fungal sensitivity to the cell wall-perturbing agent calcofluor white and reduced mycelial ball formation in liquid culture. The absence of GAG polysaccharides reduced root-binding capacity by 50% and increased cotton immune responses 1 day post fungal infection. The ΔVdGtb mutant exhibited a significant 30.5% decrease in the pathogenicity toward cotton seedlings compared with the wild type V991. Microbiome and bacterial enrichment analysis indicate that the GAG polysaccharides promote the enrichment of soil bacteria and alter the bacterial community structure in the plant rhizosphere. Several bacteria enriched by GAG-contained fungal cells, including Achromobacter animicus, Pseudomonas aeruginosa, and Acinetobacter pittii exhibited strong growth-inhibitory effects against V. dahliae and showed distinct effects on fungal virulence in a GAG-dependent manner. Together, these results reveal that GAG is not merely a cell wall component but a multi-functional molecule that orchestrates fungal protection, host infection, and inter-kingdom microbial communication.