P‐anisaldehyde Suppresses Aspergillus Flavus Infection and Aflatoxin Biosynthesis Via Targeting Cell Structure and Ap1‐regulatory Antioxidant System

Aflatoxin, mainly produced by Aspergillus flavus, is one of the most notorious mycotoxin for its toxicity and carcinogenicity. Despite extensive efforts, effective strategies to control A. flavus and AFB1 contamination remain elusive. Here, we investigate the potential of p-anisaldehyde (AS), an aldehyde derived from plant essential oils, as a natural antifungal agent against A. flavus and its ability to modulate AFB1 biosynthesis. We found that AS exhibited broad-spectrum antifungal activities against Aspergillus spp. and effectively inhibited A. flavus asexual development, AFB1 production, and pathogenicity. AS treatment disrupted the cell surface structure and membrane integrity, as observed by scanning electron microscopy and PI staining. RNA-sequencing analysis revealed that AS significantly altered the expression of genes involved in redox homeostasis, plasma membrane function, and cell cycle progression. Further investigation demonstrated that AS induced a reduction in mitochondrial membrane potential (Delta psi m) and accumulation of reactive oxygen species (ROS), leading to cell cycle arrest at the G2/M phase. The AS-induced ROS accumulation was found to be mitigated by the superoxide dismutase-mediated antioxidant system, which is regulated by transcriptional factor Ap1. Notably, the Ap1-regulatory ROS detoxification system was also found to be involved in A. flavus pathogenicity and AFB1 production. Overall, these findings provide valuable insights into the inhibitory mechanism of AS against A. flavus, paving the way for its potential application as a natural strategy to mitigate AFB1 contamination in both food and agriculture crops.