Screening And Antibiotic Characterization Of Volatile Organic Compounds Produced By Antagonistic Yeasts Against Grape Grey Mold

Grape grey mold caused by Botrytis cinerea is particularly prominent during post-harvest storage of grapes and is regarded as a devastating disease that causes significant economic losses to the grape industry.Control of grape grey mold mainly based on chemical control,combined with agricultural cultivation measures,preservation technology,and so on.The long-term use of chemical fungicides increases the drug resistance of B.cinerea,and it easily pollutes the fruit and ecological environment and causes harmful to human and animal health.Therefore,the search for a safe and effective control measure for grape gray mold has become an urgent demand in the grape industry.The purpose of this study was to screen and obtain yeast strains that were effective against B.cinerea in the natural world,and explore the antibacterial mechanism of the yeast strains in order to provide the basis for the biological control of grape grey mold.Objective:Through screening antagonistic yeasts with inhibitory effect on grape gray mold and their taxonomic status was determined;The optimal culture conditions of the VOCs produced by yeast 0939-5were determined;The components of the VOCs were analyzed,initially detected the inhibitory effect of the main VOCs of yeast on B.cinerea;Revealed the main antagonistic mechanism of yeast 0939-5 against grape gray mold.Methods:Antagonistic yeasts were obtained by in vitro and in vivo screening of the 334 yeast strains tested by the plate isolation confrontation method and inoculation method in grape berries;The antagonistic yeast 0939-5 was identified by morphological,physiological and biochemical characteristics and molecular biology;Using double dish clasp method determined different environmental conditions and different nutrients on the production of VOCs in yeast;The inhibition effect of adding activated carbon on the production of VOCs and the inhibition effect of VOCs on the growth of hyphae and spore germination were detected by plate isolation confrontation and double dish clasping;The VOCs produced by yeast were analyzed and detected by headspace solid phase microextraction and gas chromatography?GC-MS?.Results:1)Screening from 334 strains of yeast obtained an antagonistic yeast 0939-5 producing VOCs and had remarkable inhibition effect.The yeast strain 0939-5 was identified as W.anomalus based on morphological,physiological and biochemical characteristics and molecular biology.2)It was found that the production of VOCs by strain 0939-5 had the best inhibitory activity against Botrytis cinerea on YPD medium,with an inhibition rate of 64.49%.MCDA medium with sucrose as carbon source and ammonium sulfate as nitrogen source had a pH of 7 and a temperature of 25?favored the production of VOCs by yeast 0939-5.3)The non-volatile substances produced by yeast strain 0939-5 did not inhibit the mycelium growth of B.cinerea,but yeasts could inhibit the mycelium growth of B.cinerea by competition action;the inhibition rate of mycelial growth by the VOCs was 71.59%,and the inhibition rate of spore germination was 99.33%,resulted in mycelial malformation,distortion,and increased mycelial branching.With the activated carbon,the inhibition effect of the VOCs on B.cinerea was significantly reduced.In addition,the VOCs produced by yeast 0939-5 also had different degrees inhibition effect on other 7 pathogenic fungi.4)Using GC-MS to test,there were a total of 27 VOCs produced by yeast 0939-5 whose peak area was more than 0.1%.Among them,Styrene had the highest content of 27.39%.After testing the pure products of the five VOCs the results showed that Styrene had the best inhibitory effect on the mycelial growth of B.cinerea,IC₅₀0 less than 5?L/L;the inhibitory effect of Benzocyclobutene was the best on spore germination,IC₅₀0 less than 15?L/L.Conclusion:The yeast strain 0939-5 was isolated and screened to have a significant inhibitory effect on the mycelial growth and spore germination of B.cinerea,it was identified as W.anomalus according to morphological characteristics,physiological and biochemical characteristics and molecular biology;the production of VOCs by strain 0939-5 had the best inhibitory effect against B.cinerea on YPD medium,MCDA medium with sucrose as carbon source and ammonium sulfate as nitrogen source had a pH of 7 and a temperature of 25?favored the production of VOCs by yeast 0939-5.The highest content of Styrene in the VOCs produced by yeast 0939-5,and the inhibition effect of Styrene on the growth of B.cinerea mycelium was the best;Benzocyclobutene was the best on inhibition of spore germination.This study provides a theoretical basis for the control of grey mold and other diseases by using VOCs produced by yeast in the future.