Screening Of Antagonistic Microorganism Against Mildew In Protectorate Vegetables And Studying Of Agents

Since the international symposium on Botrytis cinerea was held once every four years，Botrytis cinerea of a worldwide disease had attracted great attention all over the world.The antagonistic microorganisms and their metabolic products were used against Botrytis cinerea,reducing the using of chemical pesticides,solving the problem of pesticide residues, delaying the emergence of drug-resistant,and providing the guarantee for the sustainable development of the agricultural economy.In this article, the bacterial CNY-04of having strong antagonism and stability genetic was getted by screening and rescreening.It was identified and its antibacterial mechanism was determined.The antibacterial substances were separated and extracted.Its physicochemical properties were also studied.The culture conditions were optimized.At the same time,the colonization of the strain CNY-04and the controlling effect in field were studied.we draw the following conclusions:1Screening and identification of antagonistic microorganismsThe antagonistic bacteria CNY-04(KC167881) with greaat antagonistic effect and stability genetic was screened from the vegetable base in Liaoning and Shandong Province and was identified as Serratia grimesii.2Study on mechanism of the antagonistic bacteria CNY-04against Botrytis cinereaThe antibacterial substance produced by the strain CNY-04hadn’t been found in the ferment filtrate.But the antagonistic phenomenon was showed when the bacteria and the pathogen spores was cultured tegether.So the antimicrobial substances could be produced though inducing or stimulating by Botrytis cinerea.That was reported for the fisrt time.And the CNY-04didn’t produce pyrrolnitrin, hydrocyanic acid, and hydrolases,but it could produce siderophore and proteinase. The bacteriostatic mechanism was completely different with the Serratia reported.3Separating ang extracting of antibacterial substances and reseaching of its physical and chemical characteristicsThe study showed that:the antibacterial substances could dissolved in water and methanol,but it insolubled in ethyl acetate and n-hexane.Soit was polar substances.Analysised by the liquid chromatography-mas spectrometry,the size of the molecular weight of the antibacterial substances was approximately200Da-400Da.Treated with30min under20℃-100℃, the antibacterial substances was stability. Treated with60min under100℃, the antibacterial substances had75.3%of antibacterial activity compared with the control group.Under the environment of pH2-pH10,. the antibacterial activity was stability.when pH greater than10, antibacterial activity declined sharply.4Optimizating of culture conditions for the antagonistic bacteria CNY-04The growth conditions of the biocontrol strain CNY-04were optimized by Response Surface Methodology basing on single factor tests. The optimal culture conditions of the biocontrol strain CNY-04were determined for2%peptone,0.1%yeast extract,0.5%beef extract, temperature of32℃, pH8.0, speeding of150r/min, the incubation time of48h, inoculation amount of4%, liquid volume of75mL(250mL). The antibacterial circle diameter was44.5mm, improving by30.9%.5Study on colonization capacity and field efficiency for the antagonistic bacteria CNY-04The test results showed that:the strain CNY-4could colonizate on tomato foliar and soil,and the amount of colonization were102-103cfu/leaf and105cfu/g, respectively; The inhibitory activity of tomatoes isolated and Botrytis cinerea was67.27%and71.14%respectively.Compared with chemical pesticides, the difference was not significant.So the strain CNY-04had a good potential.