Essential Oils and Hydrolates: Potential Tools for Defense against Bacterial Plant Pathogens

Abstract

The essential oils (EOs) of Origanum compactum and Satureja montana chemotyped (CT) at carvacrol, two Thymus vulgaris CT at thujanol and thymol, and Hydrolates (Hys) of S. montana and Citrus aurantium var. amara were chosen for studying their bactericidal efficacy against few phytobacterial pathogens. The Minimal Inhibitory Concentration (MIC) and Bactericidal Concentration (MBC) were found by microdilution assay. The essential oils of O. compactum (MBC 0.06% v/v), T. vulgaris CT thymol (MBC 0.06% v/v), and Hy of C. aurantium (MBC 6.25% v/v) resulted in being the most effective against Erwinia amylovora; thus, they were used as starting concentrations for ex vivo assays. Despite the great in vitro effectiveness, the disease incidence and the population dynamic ex vivo assays showed no significant results. On the other hand, EO of O. compactum and Hy of C. aurantium (at 0.03% and 4.5% v/v, respectively) showed resistance induction in tomato plants against Xanthomonas vesicatoria infections; both treatments resulted in approximately 50% protection. In conclusion, EOs and Hys could be promising tools for agricultural defense, but further studies will be necessary to stabilize the EOs emulsions, while Hys application could be an effective method to prevent bacterial diseases when used as resistance inducer by pre-transplantation treatment at roots.

Keywords: antibacterial activity; essential oil; hydrolate; induced resistance; sustainable agriculture defence.

Figure 1

Pear flowers disease incidence. Flowers cv. Williams, inoculated with E. amylovora, pre-treated with streptomycin sulphate at 100 μg/mL (SM, positive control), sterile distilled water (SDW, negative control), O. compactum CT carvacrol 0.06% and 0.12% (OC 0.06%), T. vulgaris CT Thymol 0.06% (TM 0.06%). The phytopathometric assessments were performed 4 and 6 dpi. The bars represent the standard deviations, and the letters indicate the statistical categories (SNK, p < 0.05).

Figure 2

Fire blight incidence on pear flowers cv. Williams. Treatment: O. compactum CT carvacrol at 0.06% (OC 0.06%), T. vulgaris CT Thymol at 0.06% (TM 0.06%), streptomycin sulphate at 100 μg/mL (SM, positive control), sterile distilled water (SDW, negative control).

Figure 3

Holed pear fruitlets disease incidence. Immature pear fruits cv. Abate Fétel inoculated with E. amylovora, pre-treated with streptomycin sulphate at 100 μg/mL (SM, positive control); sterile distilled water (SDW, negative control); O. compactum CT carvacrol at 0.12% prepared in accordance with M1 (OC-M10.12%), O. compactum CT carvacrol at 0.12% prepared in accordance with M2 (OC-M2 0.12%), T. vulgaris CT thymol at 0.12% prepared in accordance with M2 (TM-M2 0.12%). Symptoms were recorded at 3 and 6 dpi. The bars represent standard deviation and the letters the statistical categories (Tukey HSD, p < 0.05).

Figure 4

Punctured pear fruitlets disease incidence. Immature pears cv. Abate Fétel were pierced, inoculated with E. amylovora, and pre-treated with streptomycin sulphate at 100 μg/mL (SM, positive control), sterile distilled water (SDW, negative control); O. compactum at 0.12% prepared according to M2 (OC 0.12%). The observations were performed from 3 to 6 dpi. The bars indicate standard deviations, and the letters represent the statistical categories (Tukey HSD, p < 0.05).

Figure 5

Population dynamics of E. amylovora rifampicin-resistant strain Ea 273R1, on apple blossoms cv. Rome Beauty pre-treated with sterile distilled water (SDW, negative control); O. compactum at 0.12% (OC 0.12%); O. compactum at 0.5% (OC 0.50%); the pathogen population was monitored for 96 h. Empty circles indicate the E. amylovora 273R1 strain population on each flower. The filled black circles and the black line indicate the mean population present in 5 flowers/day. Standard deviation bars are shown for each time point.

Figure 6

Apple blossoms, cv. Rome Beauty, treated to test population dynamics with EOs: O. compactum at 0.5%; note complete drying of the flowers (OC 0.5%), O. compactum at 0.12%; note slight darkening of the petals (OC 0.12%), sterile distilled water; note healthy flowers without darkening (SDW, negative control).

Figure 7

SEM observation of the E. amylovora 273R1 population, 48 h after its inoculation. Apple flowers hypanthium treated by nebulization of (A) sterile distilled water (negative control) and (B) O. compactum at 0.12% that shows a higher concentration of bacteria than negative treatment. The bars indicate the magnifications of 0.01 mm.

Figure 8

SEM observations of the apple flowers hypanthium surface treated and after 1 h sprayed with E. amylovora 273R1: (A) untreated and uninoculated flower; (B) flower treated with sterile distilled water (negative control) and inoculated; (C,D) flowers treated with O. compactum at 0.12% and 0.5% v/v, respectively, and inoculated. The red squares indicate the observation site. Bars indicate the magnifications and correspond to 1 mm, 0.1 mm, 0.01 mm, respectively.

Figure 9

SEM observation of apple flower hypanthium surface after treatment with O. compactum at 0.5%, 48 h after inoculation with E. amylovora 273R1. The red arrows indicate tissue cracks caused by the treatment; blue arrows indicate bacterial cells. The bars indicate a magnification of 0.01 mm.

Figure 10

In planta experiments on bacterial leafspot control (X. vesicatoria) of tomato plants (cv. VF-10) pre-treated at the roots with sterile distilled water (SDW, negative control); acibenzolar-S-methyl (ASM, 75 µg/mL, positive control); O. compactum at 0.03% (OC 0.03%); O. compactum at 0.015% (OC 0.015%); C. aurantium var. amara Hy at 4.5% (AA 4.5%); C. aurantium var. amara Hy at 2.2% (AA 2.2%). The bars indicate standard deviations, and different letters show significant differences according to Duncan’s test (p < 0.05).