Determination Of The Antifungal Effect Of Citronella Essential Oil On Botrytis Cinerea And Research On Its Mechanism

Botrytis cinerea can cause gray mold on over 1 400 species of plants such as tomatoes,strawberries,cucumbers and grapes,and is an important fungal disease in agricultural production and storage.Due to the problems of chemical fungicide resistance,residues,and re-rampant,the development of new and efficient plant-derived fungicides is a current research hotspot for gray mold control.Plant essential oils have been proven to have broad-spectrum fungicidal activity,which,combined with their low toxicity and environmental friendliness,has triggered widespread interest among scholars.In this thesis,we measured the antifungal activity of nine essential oils against B.cinerea,of which citronella essential oil had the highest antifungal activity,and further investigated its antifungal mechanism.The results are as follows:1.To clarify the antifungal effect of essential oils on B.cinerea,the mycelial growth method was used to determine the antifungal activity of nine essential oils of orange,sandalwood,citronella,mugwort,torreya,cypress,sweet orange,artemisia annua and chamomile on B.cinerea,among which citronella essential oil has the best antifungal effect,and the inhibitory rate of citronella essential oil at a concentration of 400μL/L on B.cinerea reached 58.7%.The essential oil of Citronella was analyzed by GC-MS,and 37 compounds were detected.The inhibition activities of its main components citronellol,citronellal,nerolidol and nerolidol were determined and found that citronellol had the best inhibition effect on B.cinerea with MIC and MFC of 250μL/L and 350μL/L,respectively.2.Further studies revealed that citronellol could inhibit spore germination and cell membrane breakage of B.cinerea.200μL/L of citronellol inhibited spore germination up to 97.8%;250μL/L of citronellol caused a 1.96-fold and 1.86-fold increase in the extracellular conductivity and absorbance values at OD_{260 nm} of B.cinerea,respectively,and the malondialdehyde content increased to 1.4 times that of the control.Meanwhile,citronellol led to a 3.63-fold increase in hydrogen peroxide content.The activities of antioxidant enzymes CAT,POD and SOD increased and then decreased with the increase of citronellol treatment,and finally decreased to 57%,45%and 60%of the control.3.In order to investigate the mechanism of citronellol inhibition on B.cinerea,transcriptome sequencing was performed on B.cinerea mycelium.Transcriptomic studies screened a total of 4 051significantly differentially expressed genes,of which 2 074 genes were up-regulated and 1 977 genes were down-regulated.A large number of up-regulated differentially expressed genes were enriched in glycerophospholipid metabolism,which is closely related to biofilm,and peroxisome,which is closely related to oxygen stress.Down-regulated differentially expressed genes were mainly enriched in ribosomes,ribosome biosynthesis in eukaryotes,cell cycle and biosynthesis of amino acid.4.In order to understand more precisely the changes of B.cinerea metabolites by citronellol treatment,metabolomic techniques were used to identify the primary and secondary metabolites in the samples.The results showed that a total of 429 significantly different metabolites were screened,of which192 metabolites were significantly up-regulated and 237 metabolites were down-regulated.The differential metabolites were significantly enriched in glycolytic andβ-alanine metabolic pathways.In summary,citronellol significantly inhibited mycelial growth and spore germination of B.cinerea,disrupting the membrane structure of cells and leading to the accumulation of intracellular reactive oxygen species.The high concentration of citronellol can inhibit the activity of antioxidant enzymes,leading to the intensification of oxidative stress of the fungus and eventually leading to the death of the fungus.This study combines transcriptomic and metabolomic technologies to systematically investigate the inhibition of essential oils,further elaborate the mechanism of essential oil inhibition,and provide a theoretical basis for the development of novel plant-derived fungicides.