Agaricus bisporus is a globally important edible fungus. The occurrence of ginger blotch caused by Pseudomonas 'gingeri' during A. bisporus growth and post-harvest stages results in significant economic losses. The biotoxin monoacetylphloroglucinol (MAPG) produced by P. 'gingeri' is responsible for inducing ginger blotch on A. bisporus. However, the understanding of the toxic mechanisms of MAPG on A. bisporus remains limited, which hinders the precise control of ginger blotch disease in A. bisporus and the breeding of disease-resistant varieties. Integrating transcriptomic, metabolomic, and physiological data revealed that MAPG led to an increase in intracellular superoxide anion (O2 -) levels and lipid peroxidation in A. bisporus. MAPG changed the cellular membrane composition of A. bisporus, causing to damage membrane permeability. MAPG inhibited the expression of genes associated with the 19s subunit of the proteasome, thereby impeding cellular waste degradation in A. bisporus. Unlike melanin, MAPG stimulated the synthesis of flavonoids in A. bisporus, which might explain the manifestation of ginger-colored symptoms rather than browning. Meanwhile, the glutathione metabolism pathway in A. bisporus played a pivotal role in counteracting the cytotoxic effects of MAPG. Additionally, enhanced catalase activity and up-regulation of defense-related genes, including cytochrome P450s, Major Facilitator Superfamily (MFS), and ABC transporters, were observed. This study provides comprehensive insights into MAPG toxicity in A. bisporus and uncovers the detoxification strategies of A. bisporus against MAPG. The findings offer valuable evidence for precise control and breeding of resistant varieties against ginger blotch in A. bisporus. © 2024 Society of Chemical Industry.