Screening Of Zearalenone Degrading Bacteria And Its Degradation Effect

Zearalenone(ZEN),also known as F-2 toxin,is a harmful mycotoxin with estrogen-like effect produced by Fusarium fungi.The toxin is widespread in mildewed barley,wheat,corn and sorghum,which are commonly eaten crops.Due to the toxic effects of ZEN,including reproductive toxicity,genetic toxicity and cytotoxicity,direct consumption or accumulation through the food chain can cause acute or chronic poisoning to human body.Therefore,the detoxification method of ZEN has been receiving extensive attention.Methods of removing ZEN include traditional physical and chemical methods as well as biological detoxification methods.However,it is difficult for physical methods to deal with a large number of grains,meanwhile chemical methods may bring other chemical reagents contaminate,destroy the nutritional content of grains,and the taste of chemically-treated grains will deteriorate.So biological detoxification is the best way to deal with ZEN contaminated grains.The biodegradation method uses microorganisms and the enzymes they produce to interact with ZEN to destroy their toxin-producing structure,which has high specificity,and these enzymes will not affect the nutritional value of grains.In this paper,by taking ZEN as the sole carbon source,we screened out the degrading bacteria H35 which has good degradation effect on ZEN,and its physiological and biochemical identification and molecular identification were carried out.Subsequently,the growth and degradation characteristics of the degrading bacteria were studied.Finally,the degradation mechanism of the degrading bacteria was preliminarily studied and the extracellular enzymes secreted by the degrading bacteria H35 were used to degrade ZEN in corn and feed.The main results of the research are as follows:A strain of degrading bacteria H35 with good degradation effect on zearalenone was isolated from wheat fields infected with head blight.Through the morphological observation,physiological and biochemical identification and 16 S rDNA sequence analysis of H35,the bacterium was identified as Curtobacterium.This is the first time that bacteria of the Curtobacterium have been identified as having the ability to degrade ZEN.The growth and degradation characteristics of H35 showed that strain H35 grew well in LB medium and was not sensitive to polymyxin B.The optimal growth conditions of H35 were temperature 30 ?,initial pH 7.0.The more aeration,the greater the growth of the strain.The optimal degradation conditions of H35 were temperature 35? and initial pH 7.0.When the added amount of bacteria liquid is 5%and the initial concentration of ZEN is 20 ?g/m L,the degradation rate of ZEN by strain H35 could reach more than 70% within 5 days under the optimal degradation conditions.The preliminary study on the degradation mechanism of H35 showed that the active substances of strain H35 were mainly distributed in the fermentation supernatant.The fermentation supernatant is treated with proteinase K,proteinase K and SDS,and heating,respectively.The degradation activity of the supernatant is significantly reduced.Therefore,it was speculated that the active substances were extracellular enzymes.The active substance was extracted by the ammonium sulfate precipitation method.It was found that the content of the active substance precipitated by ammonium sulfate with concentration of 60%,65%,70%,75% and 80% had little different.The content of degradation active substances with molecular weight of 30 kDa and 38 kDa was the highest.The effect of H35 degradation products of ZEN on the proliferation of human breast cancer MCF-7 cells indicated that the estrogenic effect of ZEN degradation products was significantly less than that of ZEN.Finally,the extracellular enzyme precipitated by ammonium sulfate was added to the mixed corn sample containing ZEN and the three feeds of DDGS,soybean meal and bran contaminated with ZEN,and it was found that the H35 extracellular enzyme had a good degradation effect on ZEN in corn and feed.It shows that the extracellular enzyme of strain H35 can be used as a potential feed additives in practical production.