The agmatinase MrAGM1 is required for Metarhizium robertsii virulence by modulating putrescine‐associated appressorium development

BACKGROUND: Fungal polyamine metabolism plays an integral role in cellular development and host-pathogen interactions. However, the biological functions of agmatinase, a key enzyme that hydrolyzes agmatine to putrescine and urea, remain poorly characterized in entomopathogenic fungi. RESULTS: In this study, we characterized the agmatinase gene, MrAgm1, in Metarhizium robertsii. Transcriptional analysis showed that MrAgm1 was upregulated during appressorium development. Fusion protein of MrAGM1-GFP was localized to vacuoles and cell membranes. Although the deletion of MrAgm1 did not alter vegetative growth, conidiation, or tolerance to various abiotic stresses, it significantly attenuated fungal virulence against Galleria mellonella larvae via topical inoculation, as evidenced by a 1.94-day delay in the median lethal time. Conversely, intrahemocoel injection bioassays revealed no significant difference in virulence between the WT and ΔMrAgm1 strains, indicating that MrAgm1 is specifically required for host cuticle penetration rather than postinfection colonization. Further phenotypic assays demonstrated that ΔMrAgm1 exhibited compromised conidial adhesion, reduced appressorium formation and impaired penetration through cicada wings. These defects correlated with the significant downregulation of key infection-related genes encoding adhesins (mad1), proteases (pr1A, pr1B, pr1C and pr1F) and a chitinase (chit30). Notably, the addition of exogenous putrescine successfully rescued the appressorium formation defect of the ΔMrAgm1 strain. CONCLUSION: MrAGM1 regulates conidial adhesion, appressorium morphogenesis and cuticle penetration via the putrescine pathway, reflecting its specialized function in early infection of M. robertsii. These findings advance mechanistic understanding of agmatinase function in polyamine metabolism of entomopathogenic fungi, and provide a molecular target to boost their biocontrol efficacy. © 2026 Society of Chemical Industry.