(<i>E</i>)-2-Hexenal, as a Potential Natural Antifungal Compound, Inhibits <i>Aspergillus flavus</i> Spore Germination by Disrupting Mitochondrial Energy Metabolism

Abstract

Fungal contamination imposes threats to agriculture and food production and human health. A method to safely and effectively restrict fungal contamination is still needed. Here, we report the effect and mode of action of (E)-2-hexenal, one of the green leaf volatiles (GLVs), on the spore germination of Aspergillus flavus, which can contaminate a variety of crops. The EC50 value, minimum inhibitory concentration (MIC), and minimum fungicidal concentration (MFC) of (E)-2-hexenal were 0.26, 1.0, and 4.0 μL/mL, respectively. As observed by scanning electron microscopy (SEM), the surface morphology of A. flavus spores did not change after treatment with the MIC of (E)-2-hexenal, but the spores were shrunken and depressed upon treatment with the MFC of (E)-2-hexenal. The MIC and MFC of (E)-2-hexenal induced evident phosphatidylserine (PS) externalization of A. flavus spores as detected by double staining with Annexin V-FITC and propidium iodide, indicating that early apoptosis was potentially induced. Furthermore, sublethal doses of (E)-2-hexenal disturbed pyruvate metabolism and reduced the intracellular soluble protein content of A. flavus spores during the early stage of germination, and MIC treatment decreased acetyl-CoA and ATP contents by 65.7 ± 3.7% and 53.9 ± 4.0% (P < 0.05), respectively. Additionally, the activity of mitochondrial dehydrogenases was dramatically inhibited by 23.8 ± 2.2% (P < 0.05) at the MIC of (E)-2-hexenal. Therefore, the disruption of mitochondrial energy metabolism and the induction of early apoptosis are involved in the mechanism of action of (E)-2-hexenal against A. flavus spore germination.