Screening Of T-2 Toxin Detoxification Strains And Study Of The Detoxification Characteristics

T-2 toxin is a kind of trichothecene produced by Fusarium,which mainly exists in moldy crops and feeds.The content of toxin increases with the increase of environmental humidity.T-2 toxin can be used for bioaccumulation and transmission through food chain.It is harmful to the safety and health of people and animals.In this experiment,the microorganisms with high effective scavenging effect on T-2 toxin were obtained from soil samples where severe contamination of Fusarium occurred in Jiangsu and Zhejiang Provinces.The detoxification characteristics of the positive strains were analyzed,and their detoxification mechanism was compared.The main results were as follows:?1?231 soil samples were divided into 50 groups.These mixture fluid were high-throughput screened with 96 well plates.Screening group had 15 positive of fluid.Further screening from the positive of liquid,six strains?A24,B7,B14,B22,B25 and B26?that could efficiently eliminate T-2 toxins in the system of single strain were obtained.Six detoxicating strains were identified by 16S rDNA.The results showed that A24,B7,B14,B22,B25,B14 and B26 were Microbacterium paraoxydans,Ochrobactrum anthropi,Microbacillus oxydans,Achromobacter xylosoxidans,Enterobacter cancerogenus and Bacillus siamensis.Among them B7,B22,B25 were pathogenic bacteria.?2?B14 and B26 reached the stable period was 14.0h,and A24 needed 16.0h.The OD₆₀₀00 of B26 in stable period was about 7.60.The OD₆₀₀ of B14 in the stable period was 4.50.In addition,A24,B14,B26 removed more than 95.00%T-2 toxins from the system at15.0h,12.0h and 16.0h respectively.?3?The three strains of A24,B14 and B26 could adsorb T-2 toxin,and B14 strain had the strongest ability to adsorb T-2 toxin.The detoxification effect of B26 on T-2 toxin was mainly derived from intracellular enzymes,and the effects of A24 and B14 by extracellular enzymes.?4?The HPLC spectra of three strains of T-2 toxin after detoxification only B14 showed two new peaks.BLYR was used to detect the toxicity of T-2 toxin virus-free product of strain B14.The IC₂₀ value of co-cultured samples was 0.7747 at 0h,and it changed to 4.4363 at24.0h.This showed that the toxicity of detoxification product was less than 1/4000 of the toxicity of T-2 toxin.?5?There are serine proteases or mercapto proteases in the degradation enzymes of T-2toxin.The enzymes of B14 degrading enzyme contain metal ions.The supernatant of strain B14 had the best detoxification effect on T-2 toxin at 24.0-37.0?,pH=7.0-8.0.?6?The results of LC-MS analysis showed that strain B14 metabolized T-2 toxin into a less toxic product by deacetylation and deepoxidation.A total of seven kinds of metabolites were detected,including four kinds of products that did not have the literature.?7?On the basis of Yeast extract 0.50%,tryptone 1.00%,NaCl 1.00%,the the optimum combination was tsucrose 2.00%,soy peptone 0.50%,dibasic potassium phosphate 5.00mM and pH=7.0 of B14 by medium optimization experiments.?8?The enzyme of metabolic T-2 toxin in the supernatant of B14 bacteria could be separated from the AKTA by anion column in ph=8.0 and 10%of salt concentration.