Screening,Identification And Application Of Vomiting Toxin Degrading Bacteria

Deoxynivalenol(DON)is a toxin produced by Fusarium nigrum,Fusarium oxysporum,Fusarium moniliforme,etc.It is widely found in animal feed and feedstuff,such as wheat and corn.It is a mycotoxin with the highest detection rate and over-standard rate,which is harmful to animals and people.As a safe and environmentally friendly treatment,microbial fermentation treatment has become a hot spot for the study of vomiting toxin degradation.The aim of this study was to screen out microbial strains with high degrade ability to vomiting toxin,identify them and apply them to biological detoxification treatment of feed ingredients.10 samples were collected from soil,sludge,moldy straw,chicken intestinal contents,pig manure,and cow dung from different environments.DON was added to the solid medium of LB,NA and MRS to prepare the improved solid screening medium for the initial screening of the strain.The strains obtained by the initial screening were cultured in different modified liquid medium,and the DON degradation rates in the modified medium was determined and calculated.According to the culture medium,culture mode and degradation rate,some decomposing bacteria were selected for morphological observation,Gram staining,microscopic examination and 16 S rDNA conservative sequence analysis to identify their species.The selected and identified strains were combined and used to degrade the natural DON in the bran and soybean meal in optimized the fermentation conditions.The results showed that 80 strains capable of growing in the primary screening medium were screened by primary screening,and 16 strains with DON degradation rate ranging from 9.7% to 66.7% were screened by rescreening.Three strains with degradation rates of 19.1%,51.3%,and 66.7%,respectively,were selected and identified by morphological observation of colony characteristics,bacterial characteristics,and 16 S rDNA conserved sequence alignment.Among them,the colony of strain LB01 was white opaque,the surface was rough,the edge was irregular,dry and wrinkled,the cells were long rod-shaped,with capsules and spores,and the similarity with EnB-alf13(KP792638.1)in Genebank reached 92%.The degrading bacteria are Bacillus subtilis;the colony of strain NA06 is milky white,the surface is dry and opaque,the edges of the middle fold are not neat,the ends of the cells areobtuse,and the ends are long rod-shaped,without parasporal crystals,in pairs or chains.Arrangement,the cysts have no obvious expansion,and the degrading bacteria with 99% similarity with MSC46(KM222187.1)in Genebank is Bacillus amyloliquefaciens;the strain MRS13 is milky white translucent,with a diameter of0.4~2.0 mm,and the edges are neat and convex.It is smooth,smooth and moist,and the cells are short rod-shaped,and the arrangement of the cells is single and chain-like,without spores,and the degrading bacteria with 99% similarity with Sourdough-H04(MG754699.1)in Genebank is Lactobacillus plantarum.The optimal fermentation parameters for degradation of DON in degraded bran and soybean meal were determined by single factor experiment: inoculation quantity was 10 mL/100 g,fermentation temperature was 37°C,fermentation for 48 h.Through the different fermentation substrate comparison experiments,the combination of Bacillus subtilis and Lactobacillus plantarum had the highest degradation rate of vomiting toxin in bran when fermenting bran,reaching 71%.Under the same fermentation conditions,B.amyloliquefaciens had the highest degradation rate of vomiting toxin in soybean meal,reaching 82%.