| In recent years,with the intensive development of aquaculture,bacterial diseases of farmed aquatic products have been erupting,causing huge economic losses to the aquaculture industry.Aquatic anesthetic eugenol has good antibacterial and bactericidal effects,not only has a good inhibitory effect on foodborne pathogenic bacteria,but also has potential antibacterial effect on aquatic pathogenic bacteria.The purpose of this paper is to explore the antibacterial and antibacterial mechanism of eugenol on the main pathogenic bacteria Vibrio anguillarum in aquaculture,in order to provide basic data for the application of eugenol as a bacteriostatic agent in aquaculture.In this study,antibacterial effect of eugenol on Vibrio anguillarum was carried out.The inhibition zone diameter was 21.13±0.74 mm when the concentration of eugenol was6400 μg/mL,which indicated that Vibrio anguillarum is extremely sensitive to eugenol at this concentration;The minimum inhibitory concentration(MIC)of eugenol against Vibrio anguillarum was 400 μg/mL by microdiffusion method using ELISA at OD600 nm.The solutions of 400 μg/mL,800 μg/mL and 1600 μg/mL micropores were applied to the culture plates for 24 h.The concentration of Vibrio anguillarum did not grow when the concentration was ≥800 μg/mL,indicating that the minimum bactericidal concentration(MBC)was 800 μg/mL.The results of bacteriostatic aging experiment showed that 400 μg/mL eugenol basically inhibited the growth of Vibrio anguillarum within 18 h.Even after 18 h,Vibrio anguillarum began to grow slowly,but compared with the blank control group,the absorbance at the corresponding time(32-36 h)was only one-fifth of the blank control group,indicating that the eugenol was still against Vibrio anguillarum.The conductivity test was used to verify the effect of eugenol on the cell membrane permeability of Vibrio anguillarum.The conductivity test showed that the conductivity of the MIC group and the MBC group was much higher than that of the blank control group,indicating that the MIC group In the MBC group,the membrane permeability of Vibrio anguillarum was destroyed,and some ions and conductive small substances in the cells flowed outside the cells.The conductivity of the MBC group increased slightly higher thanthat of the MIC group,indicating that 800 μg/mL eugenol damages the cell membrane permeability of Vibrio anguillarum more than 400 μg/mL eugenol,and the eugenol eutropha cell membrane permeability The effect is also affected by the concentration.The effect of MIC concentration of eugenol on Vibrio anguillarum protein,succinate dehydrogenase(SDH)and malate dehydrogenase(NAD-MDH)was investigated by spectrophotometry.The results showed that compared with the blank control group,the MIC group reduced the protein by 39.4%,and the SDH and NAD-MDH enzyme activities decreased by 45.85% and 62.25%,respectively.Finally,SDS-PAGE gel electrophoresis was used to observe the molecular weight of the MIC group and the blank control group.The results showed that the molecular weight ranged from 25 KD to 150 KD.The bacterial protein band of the MIC group was larger than that of the blank control group.The bands are all lighter,that is,the molecular weight of these molecular weights is reduced;a small number of bands disappear,and it may be that these molecular weight proteins are reduced in synthesis or flow out of the bacteria. |
