The Impact of Essential Oils Derived from Citrus Species to Control Botrytis cinerea and Their Potential Physiological Actions

Abstract

Botrytis cinerea is one of the phytopathogenic fungi of the greatest economic importance worldwide. Essential oils (EOs) have been proposed as a sustainable alternative to reduce the growth of phytopathogenic fungi. Nevertheless, few studies exist about its mechanisms of action. This study evaluated the antifungal activity of EOs from Citrus reticulata, Citrus limon, Citrus sinensis, and Citrus paradisi peels and their encapsulation inside solid lipid nanoparticles (SLNs). Accordingly, Citrus EOs were mainly constituted by monoterpene hydrocarbons, where limonene was the most abundant in all EOs. C. reticulata and C. limon EOs reduced the mycelial growth at above 54% after 96 h. The other EOs did not significantly impact the phytopathogen. C. reticulata EO increased the hyphae damage by 40%, but the spore germination was reduced by only 8.34%. It also significantly increased the pH, the electrical conductivity, and the release of intracellular absorbing material and soluble proteins in B. cinerea cultures. Contrary, the esterase, mitochondrial, and succinate dehydrogenase activities decreased at above 50%. C. reticulata EO into SLN reduced the mycelial growth of B. cinerea by 90-97%. These results show that the EO of C. reticulata alters the physiological and metabolic activities of B. cinerea to reduce its growth.

Keywords: Botrytis cinerea; citrus essential oil; physiological mechanisms; solid lipid nanoparticles.

Figure 1

The impact of different concentrations of essential oils of (a) Citrus reticulata, (b) Citrus limon, (c) Citrus paradisi, and (d) Citrus sinesis on mycelial growth of Botrytis cinerea. (mean values ± standard error n = 4).

Figure 2

Inhibition of spore germination of B. cinerea exposed to different concentrations of C. reticulata EO. Different letters above bars indicate significant differences according to the Tukey test (p < 0.05) (mean values ± standard error, n = 3).

Figure 3

Effects of C. reticulata EO on (a) dry weight and (b,c) release of cell constituents of B. cinerea measurement through extracellular pH and conductivity. Different letters above bars indicate significant differences according to the Tukey test (p < 0.05) (mean values ± standard error, n = 3).

Figure 4

The impact of essential oil from C. reticulata peels on (a) intracellular absorbing material OD_260nm and (b) extracellular soluble proteins. Different letters above bars indicate significant differences according to the Tukey test (p < 0.05) (mean values ± standard error, n = 3). *ns indicate no statistically significant differences.

Figure 5

The potential action mechanisms of the C. reticulata EO to suppress the growth of B. cinerea were measured through (a) mitochondrial activity, (b) viability, (c) hyphae damage, and (d) intracellular esterases measured by the MTT technique. Different letters above bars indicate significant differences according to the Tukey test (p < 0.05) (mean values ± standard error, n = 3).

Figure 6

Physicochemical characterization and antifungal activity of solid lipid nanoparticles (SLNs) loaded with C. reticulata EO. (a,c) Hydrodynamic size distribution and (b,d) ζ-potential of SLNs, (e) representative photograph of SLN captured by scanning transmission electron microscopy (STEM), and (f) the mycelial growth inhibition of B. cinerea modulated the controlled release of the EO of C. reticulata from SLN (mean values ± standard error, n = 3).