Synergistic antifungal activity and mechanism of carvacrol/citral combination against fusarium oxysporum in Dendrobium officinale

Abstract

The soft rot disease caused by Fusarium oxysporum leads to a significant reduction in the yield of Dendrobium officinale. However, research into the synergistic inhibitory effect of essential oils in D. officinale is extremely limited. In this study, we systematically investigated the direct and indirect inhibitory activity of carvacrol, citral and their combination against F. oxysporum, and their synergistic inhibitory mechanism. Carvacrol and citral exhibited significant direct and indirect inhibitory activity against F. oxysporum with EC₅₀ values of 54.37 mg/L and 119.32 mg/L (direct), 18.01 μL/(L•air) and 48.70 μL/(L•air) (indirect). Synergistic analysis revealed that the optimal synergistic toxicity of carvacrol and citral combination (Ca•Ci) against F. oxysporum was 10:1, with co-toxicity coefficient (CTC) of 131.57 and EC₅₀ value of 44.24 mg/L. Microscopy confirmed that the Ca•Ci led to more significant tip constriction, uneven surfaces and serious rupture of F. oxysporum mycelia than single compound. Transmission electron microscopy (TEM) showed that Ca•Ci also caused significant ultrastructural alterations to F. oxysporum, manifesting as cytoplasmic disorganization and partial organellar disintegration. Moreover, Ca•Ci dramatically increased the sensitivity of F. oxysporum to calcofluor white compared with a single compound. Ca•Ci also considerably upregulated the expression of chitinase-related gene (FOXG_12882) and β-1,3-glucanase-related gene (FOXG_10637) in F. oxysporum, resulting in higher chitinase and β-1,3-glucanase activity. However, it should be noted that carvacrol exerted a greater contribution than citral. In conclusion, the combination of carvacrol and citral greatly disrupted the cell wall integrity of F. oxysporum, thereby exhibiting a synergistic effect.