Antifungal Activity and Mechanisms of 2-Nonanone, a Volatile Organic Compound Produced by Bacillus velezensis TCS001, against Botrytis cinerea.

Botrytis cinerea is a phytopathogenic fungus notorious for causing gray mold diseases worldwide. The biocontrol agent (BACs) Bacillus velezensis TCS001 inhibits B. cinerea via volatile organic compounds (VOCs). 2-Nonanone was identified as the predominant VOC and strongly suppressed mycelial growth (minimal inhibitory concentration (MIC) 160 μL/L, minimal fungicidal concentration (MFC) 1280 μL/L) and reduced gray mold on cherry tomatoes. It induced morphological and ultrastructural abnormalities, impaired cell wall and membrane integrity, decreased ergosterol content and ATPase, succinate dehydrogenase (SDH), malate dehydrogenase (MDH) activities, and increased malondialdehyde (MDA) levels. Transcriptomic analysis revealed modulation of genes involved in ribosome biogenesis, amino acid and secondary metabolite biosynthesis, including 23 down-regulated pathogenicity-related genes. Drug affinity responsive target stability (DARTS) and microscale thermophoresis (MST) identified COP9 signalosome subunit 6 (BcCsn6) as a direct binding target of 2-nonanone. BcCsn6 knockout mutants showed impaired growth, abolished pathogenicity, and reduced sensitivity to 2-nonanone, confirming it as a key functional target. These findings highlight TCS001 as a promising biocontrol agent against gray mold.