Enzymatic degradation of mycotoxins during grain processing

Abstract

Methods, compositions, and systems for steeping, propagation and fermentation, particularly large-scale operations for production of starch and ethanol and fermentation product streams are provided. Addition of mycotoxin mitigating enzymes or microorganisms expressing mycotoxin mitigating enzymes to steeping, propagation, and/or fermentation tanks, and/or to post-fermentation product streams, mitigates mycotoxin levels in fermentation co-products obtained from mycotoxin contaminated feedstocks.

What is claimed is: 
     
         1 . A livestock feed composition comprising distiller's grain sourced from feedstock fermented in the presence of a microorganism expressing an enzyme to mitigate mycotoxin contamination, or a cell lysate comprising a microorganism expressing an enzyme to mitigate mycotoxin contamination, wherein the levels of mycotoxin in the livestock feed composition are decreased relative to the levels of mycotoxin in a livestock feed composition obtained from a feedstock fermented in the absence of a microorganism expressing an enzyme to mitigate mycotoxin contamination or a cell lysate comprising a microorganism expressing an enzyme to mitigate mycotoxin contamination. 
     
     
         2 . A livestock feed composition comprising distiller's grain sourced from post-fermentation stillage, wet cake, and/or syrup stream treated with an enzyme to mitigate mycotoxin contamination, wherein the levels of mycotoxin in the livestock feed composition are decreased relative to the levels of mycotoxin in a livestock feed composition obtained from untreated post-fermentation stillage, wet cake, and/or syrup stream, and wherein the enzyme is obtained by inoculating post-fermentation stillage, wet cake, and/or syrup stream with a microorganism expressing an enzyme to mitigate mycotoxin contamination or by adding to the post-fermentation stillage, wet cake, and/or syrup stream a cell lysate comprising a microorganism expressing an enzyme to mitigate mycotoxin contamination. 
     
     
         3 . A livestock feed composition comprising distiller's grain sourced from beer treated with an enzyme to mitigate mycotoxin contamination, wherein the levels of mycotoxin in the livestock feed composition are decreased relative to the levels of mycotoxin in a livestock feed composition obtained from untreated beer, and wherein the enzyme is obtained by inoculating beer with a microorganism expressing an enzyme to mitigate mycotoxin contamination or by adding to the beer a cell lysate comprising a microorganism expressing an enzyme to mitigate mycotoxin contamination. 
     
     
         4 . The livestock feed composition of  claim 1 , wherein the mycotoxin is selected from the group consisting of aflatoxin, Ochratoxin A, patulin, T-2, HT-2, beauvericin, neosolaniol, nivalenol, deoxynivalenol, 3-ADON, trichothecene, ochratoxin, and zearalenone. 
     
     
         5 . The livestock feed composition of  claim 1 , wherein the enzyme is selected from the group consisting of aflatoxin oxidase, 3-O acetyltransferase, peroxidase, F420H2-dependent reductases, Ery4, UDP-glycosyltransferase, laccase, manganese peroxidase, deoxynivalenol hydroxylase, xylanase, DepA/DepB, lactono hydrolase, Zearalenone hydrolase, and 2cys-peroxiredoxin. 
     
     
         6 . The livestock feed composition of  claim 1 , wherein the microorganism is a yeast. 
     
     
         7 . The livestock feed composition of  claim 1 , wherein the enzyme is expressed by a yeast host cell. 
     
     
         8 . The livestock feed composition of  claim 1 , wherein the levels of mycotoxin are less than about 10 ppm. 
     
     
         9 . A method of fermentation comprising inoculating a feedstock with one or more yeast strains, wherein at least one yeast strain expresses one or more mycotoxin mitigating enzymes. 
     
     
         10 . The method of  claim 9 , further comprising fermenting the feedstock to produce ethanol and distiller's grain. 
     
     
         11 . The method of  claim 9 , wherein the mycotoxin mitigating enzyme is deoxynivalenol hydroxylase. 
     
     
         12 . The method of  claim 11 , wherein the feedstock is contaminated with deoxynivalenol. 
     
     
         13 . The method of  claim 11 , wherein the levels of deoxynivalenol in the distiller's grain are decreased relative to the levels of deoxynivalenol in distiller's grain obtained from a feedstock fermented in the absence of a yeast expressing deoxynivalenol hydroxylase. 
     
     
         14 . A method of mitigating mycotoxin contamination in livestock feed, the method comprising:
 (i) fermenting a feedstock in the presence of a microorganism expressing an enzyme selected from the group consisting of aflatoxin oxidase, 3-O acetyltransferase, peroxidase, F420H2-dependent reductases, Ery4, UDP-glycosyltransferase, laccase, manganese peroxidase, deoxynivalenol hydroxylase, lactono hydrolase, Zearalenone hydrolase, xylanase, DepA/DepB, and 2cys-peroxiredoxin;   (ii) contacting post-fermentation stillage, wet cake, and/or syrup stream with an enzyme selected from the group consisting of aflatoxin oxidase, 3-O acetyltransferase, peroxidase, F420H2-dependent reductases, Ery4, UDP-glycosyltransferase, laccase, manganese peroxidase, deoxynivalenol hydroxylase, lactono hydrolase, Zearalenone hydrolase, xylanase, DepA/DepB, and 2cys-peroxiredoxin; or   (iii) contacting beer with an enzyme selected from the group consisting of aflatoxin oxidase, 3-O acetyltransferase, peroxidase, F420H2-dependent reductases, Ery4, UDP-glycosyltransferase, laccase, manganese peroxidase, deoxynivalenol hydroxylase, lactono hydrolase, Zearalenone hydrolase, xylanase, DepA/DepB, and 2cys-peroxiredoxin.   
     
     
         15 . The method of claim  14 (b), wherein the levels of mycotoxin in the livestock feed composition are decreased relative to the levels of mycotoxin in a livestock feed composition obtained from untreated post-fermentation stillage, wet cake, and/or syrup stream, wherein the enzyme is obtained by inoculating post-fermentation stillage, wet cake, and/or syrup stream with a microorganism expressing an enzyme to mitigate mycotoxin contamination or by adding to the post-fermentation stillage, wet cake, and/or syrup stream a cell lysate comprising a microorganism expressing an enzyme to mitigate mycotoxin contamination. 
     
     
         16 . The method of claim  14 (c), wherein the levels of mycotoxin in the livestock feed composition are decreased relative to the levels of mycotoxin in a livestock feed composition obtained from untreated pre-distillation whole stillage, and wherein the enzyme is obtained by inoculating beer with a microorganism expressing an enzyme to mitigate mycotoxin contamination or by adding to the beer a cell lysate comprising a microorganism expressing an enzyme to mitigate mycotoxin contamination. 
     
     
         17 . The method of  claim 16 , wherein the step of contacting beer with an enzyme comprises lysing yeast expressing the enzyme and adding the lysate to the beer. 
     
     
         18 . A vector comprising:
 (i) a nucleic acid sequence of SEQ ID NO: 1, or a nucleic acid having at least about 95% sequence identity to SEQ ID NO: 1, encoding deoxynivalenol hydroxylase (Ddna); or   (ii) a nucleic acid sequence of SEQ ID NO: 10, or a nucleic acid having at least about 95% sequence identity to SEQ ID NO: 10.   
     
     
         19 . The vector of  claim 18 , further comprising one or more nucleic acid sequences encoding a KdR gene, KdX gene, or Ddna gene. 
     
     
         20 . A microorganism expressing a vector of  claim 18 .