| According to the statistics of the food and Agriculture Organization of the United Nations(FAO),about 25%of the world's grain is polluted by mycotoxins,and zearalenone is one of the most widely polluted mycotoxins.Zearalenone(ZEN)is a mycotoxin isolated from corn with scab by stob et al.in1962.The main strains producing ZEN are Fusarium sp.(such as Fusarium graminearum and Fusarium flavum).Due to the continuous changes of global climate,improper storage environment,pest and rainstorm increase the risk of ZEN pollution.ZEN is a kind of exogenous estrogen,its chemical structure is similar to natural estrogen,its toxicity and pathogenicity are usually related to reproductive disorders of specific animals,not only for livestock and human and brings serious health problems and huge economic losses.Therefore,how to solve the pollution of ZEN in food and feed is of great significance to China's agricultural production and human,animal health.High temperature and acid environment are often needed in food and feed processing.In order to degrade mycotoxins in real life,it is necessary to screen highly tolerant degradation strains.Therefore,screening new toxin degrading strains from special environment has become a hot spot.The purpose of this experiment is to isolate and screen a new strain which is resistant to heat and efficient degradation of ZEN,optimize its growth and degradation conditions,The degradation substance and mechanism of ZEN and the characteristics of degradation products were analyzed,so as to apply to the removal of ZEN in food and feed.This study provides a basis for the purification of ZEN degrading enzyme and the mining of functional genes.The main conclusions of this study are as follows(1)In this study,a heat-resistant strain Y4,which can efficiently degrade ZEN,was selected from hot spring by co culture of strain toxin.It is physiological and biochemical characteristics were analyzed and 16S r RNA was identified as Enterobacter kobei.(2)The optimal growth conditions were LB medium,28°C and p H 7 for 30 h,showing a good growth advantage.(3)The effect of different conditions on the degradation of ZEN by strain Y4 was studied.The effects of reaction time,initial p H,temperature and toxin concentration on the degradation of ZEN by Y4 were analyzed.The degradation rate of ZEN was 100%when the strain was incubated with 2mg·L-1ZEN at p H 6,60°C and 180 r·min-1for 48 h.When p H was 5 and other conditions remained unchanged,the degradation rate of 2 mg·L-1ZEN was 91.43%,and that of 50 mg·L-1ZEN was 88.95%.The results showed that the degradation of ZEN by Y4 was highly efficient and heat resistant.(4)The degradation mechanism and products of ZEN by the strain were studied.Mechanism analysis showed that the degradation of ZEN by Y4 was mainly due to the degradation of extracellular enzymes,and Mn2+could significantly increase the degradation ability of supernatant.LC-MS/MS analysis showed that the degradation products of ZEN did not contain?-ZOL,?-ZOL and other estrogenic substances.(5)LC-QTOF/MS was used to find the differences between the metabolites of ZEN in Y4 and the metabolites of bacteria.After incubation with strain Y4 for 15 min,1 h,48 h and 72 h,the difference peaks were at 3.8 min and 5.4 min,the molecular weights were 291.121 and 216.0896 mz.It is speculated that it may be the degradation product of ZEN by Y4,and the molecular structure needs to be further verified.(6)Fifty nine differentially expressed genes were identified by transcriptomic analysis,including24 up-regulated genes and 35 down regulated genes.The gene related to the degradation of zearalenone is the degradation of aromatic compounds.It is speculated that the ZEN degrading enzyme of zearalenone may also be a peroxidase that acts on the benzene ring structure of ZEN.This study not only provides a new strain resource for the development and utilization of ZEN,but also lays a theoretical foundation for the research and application of ZEN detoxification enzyme preparation. |
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