Citral-Based Acylthiourea Derivative as Antifungal Synergistic Agent: E14 Enhances the Antifungal Activity of Chlorothalonil against Colletotrichum fructicola by Interfering with Mitochondrial Complex I

The citral-based acylthiourea derivative e14 was identified as a potent synergist for chlorothalonil (CT) against Colletotrichum fructicola. The optimized mixture (ECS, 25% e14) exhibited strong synergistic activity (synergistic ratio (SR) = 2.30), significantly enhancing disease control efficacy while enabling reduced fungicide dosage. Mechanistic studies revealed that e14 specifically targets mitochondrial complex I by binding to the NADH dehydrogenase subunit 5 (ND5), thereby disrupting oxidative phosphorylation and inducing energy depletion and oxidative stress. This distinct mode of action differs from that of CT, minimizing the risk of cross-resistance. Importantly, e14 exhibits low mammalian cytotoxicity. This study elucidates the synergistic mechanism of ECS and the enhancing role of e14, thereby providing a solid foundation for the development of effective fungicide synergists.