Objective Black spot of broccoli is a common disease caused by Alternaria brassicicola. The antibacterial activity of clove extract and its main active ingredient eugenol against A. brassicicola was investigated to provide a theoretical basis for the green prevention and control of black spot of broccoli.

Method The optimal bacteriostatic concentration of clove extract and its main component eugenol on A. brassicicola was determined by plate bacteriostatic test. The inhibitory activity of clove extract and its main component eugenol on A. brassicicola spore was analyzed by spore germination method. The damage of pathogen cell membrane was observed by fluorescein diacetate (FDA) and propidium iodide (PI) staining. The effects of clove extract and eugenol on the relative conductivity of A. brassicicola were determined. The bacteriostatic effect of clove extract and eugenol on black spot of broccoli leaves was explored by in vivo bacteriostatic test.

Result Both clove extract and eugenol significantly inhibited the mycelial growth of A. brassicicola in a concentration-dependent manner. Considering the cost and antibacterial effect, the optimum inhibitory concentrations of clove extract and eugenol were determined to be 10 g/L and 300 μg/mL, and the colony growth inhibition rates were 93% and 88%, respectively. Both 10 g/Lclove extract and 300 μg/mL eugenol inhibited spore germination, and the germination inhibition rates were 87% and 96%, respectively. Fluorescence staining experiments showed that clove extract and eugenol could destroy the integrity of mycelial cell membrane and lead to the outflow of cell contents. The relative conductivity results showed that the conductivity of 10 g/Lclove extract and 300 μg/mL eugenol treatment groups increased by 61% and 56%, respectively, compared with the control group. In vivo bacteriostatic test showed that clove extract and eugenol had obvious bacteriostatic effect on black spot disease of broccoli, compared to T3, the Fv/Fm values of leaves in the clove extract (T4) and eugenol (T5) treatment groups increased significantly by 70% and 68%, respectively, on the third day of treatment, and clove extract could effectively alleviate leaf cell necrosis caused by pathogen infection.

Conclusion Clove extract and eugenol can inhibit the growth of A. brassicicola mycelium, reduce the germination rate of spores, destroy the integrity of the cell membrane of mycelium, and have a good inhibitory effect on black spot disease of broccoli. It provides a theoretical basis for the development of plant-derived biological pesticides for the prevention and control of black spot disease of broccoli.