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Fumonisin Content in Conventional Versus Bt Maize and Implications Of Iowa State University Capstones, Theses and Graduate Theses and Dissertations Dissertations 2013 Fumonisin content in conventional versus Bt maize and implications of fumonisin contamination on fuel ethanol yield and DDGS quality Erin Louise Bowers Iowa State University Follow this and additional works at: https://lib.dr.iastate.edu/etd Part of the Agriculture Commons, and the Toxicology Commons Recommended Citation Bowers, Erin Louise, "Fumonisin content in conventional versus Bt maize and implications of fumonisin contamination on fuel ethanol yield and DDGS quality" (2013). Graduate Theses and Dissertations. 13596. https://lib.dr.iastate.edu/etd/13596 This Dissertation is brought to you for free and open access by the Iowa State University Capstones, Theses and Dissertations at Iowa State University Digital Repository. It has been accepted for inclusion in Graduate Theses and Dissertations by an authorized administrator of Iowa State University Digital Repository. For more information, please contact [email protected]. Fumonisin content in conventional versus Bt maize and implications of fumonisin contamination on fuel ethanol yield and DDGS quality by Erin Louise Bowers A dissertation submitted to the graduate faculty in partial fulfillment of the requirements for the degree of DOCTOR OF PHILOSOPHY Major: Toxicology Program of Study Committee: Gary P. Munkvold, Major Professor Patricia A. Murphy Richard L. Hellmich Steve M. Ensley Lawrence A. Johnson Felicia Wu Anthony L. Pometto III Iowa State University Ames, Iowa 2013 Copyright © Erin Louise Bowers, 2013. All rights reserved. ii TABLE OF CONTENTS Page ABSTRACT………………………………. .............................................................. iv CHAPTER 1. INTRODUCTION ........................................................................ 1 Dissertation Organization .................................................................................... 1 Literature Review................................................................................................. 1 References…… .................................................................................................... 24 Tables and Figures ............................................................................................... 35 CHAPTER 2. VIP3AA AND CRY1AB PROTEINS IN MAIZE REDUCE FUSARIUM EAR ROT AND FUMONISINS BY DETERRING KERNEL INJURY FROM MULTIPLE LEPIDOPTERAN PESTS ......................................... 41 Abstract……. ....................................................................................................... 41 Introduction. ......................................................................................................... 42 Materials and Methods ......................................................................................... 46 Results……...... .................................................................................................... 49 Discussion…… .................................................................................................... 52 Literature Cited .................................................................................................... 56 Tables and Figures ............................................................................................... 60 CHAPTER 3. COMPARISON OF FUMONISIN CONTAMINATION USING HPLC AND ELISA METHODS IN BT AND NEAR-ISOGENIC MAIZE HYBRIDS INFESTED WITH EUROPEAN CORN BORER OR WESTERN BEAN CUTWORM ................................................................................................... 66 Abstract……. ....................................................................................................... 66 Introduction. ......................................................................................................... 67 Materials and Methods ......................................................................................... 71 Results……...... .................................................................................................... 78 Discussion…… .................................................................................................... 85 References…… .................................................................................................... 91 Tables and Figures ............................................................................................... 95 CHAPTER 4. FUMONISINS IN CONVENTIONAL AND TRANSGENIC, INSECT-RESISTANT MAIZE INTENDED FOR FUEL ETHANOL PRODUCTION: IMPLICATIONS FOR FERMENTATION EFFICIENCY AND DDGS CO-PRODUCT QUALITY .................................................................. 105 Abstract……. ....................................................................................................... 105 Introduction. ......................................................................................................... 106 Materials and Methods ......................................................................................... 112 iii Results……...... .................................................................................................... 118 Discussion…… .................................................................................................... 123 References…… .................................................................................................... 129 Tables and Figures ............................................................................................... 134 CHAPTER 5. GENERAL CONCLUSIONS ........................................................ 143 iv ABSTRACT Genetic engineering of maize plants with genes (Bt genes) for resistance to Lepidopteran insects reduces the risk of grain contamination with fumonisin mycotoxins. Fumonisins are secondary metabolites of certain Fusarium fungal species that may be present in maize grain and, when consumed, lead to a variety to detrimental health impacts in both humans and animals. Changing insect populations, newly available transgenes, and trends in maize utilization have raised new questions about the scope of Bt effects on fumonisins. Field trials were conducted at two locations in 2008-2011 to assess the effects of expression of Bt insecticidal proteins Cry1Ab, Cry1F, or Cry1Ab x Vip3Aa on fumonisin contamination in grain following infestations with European corn borer (ECB), Western bean cutworm (WBC), corn earworm (CEW), and natural insect infestation (no insects applied). These maize pests are common in the major maize-producing regions of the United States and their feeding can reduce yields and increase levels of fungal contamination and mycotoxins. Fumonisins have also demonstrated toxic effects on certain yeast strains used in fermentation of maize grain for fuel ethanol production. Additionally, because fumonisins are heat-stable and non-volatile under relevant conditions, they can be enriched up to three- fold in dried distillers’ grains with solubles (DDGS), the residual non-fermentable co-product of fuel ethanol processing which is fed to livestock. Therefore, fumonisin contamination in maize is a significant consideration for maize and maize-based ethanol producers. Grain quality was assessed by visual inspection to determine the extent of kernel injury and Fusarium ear rot, and by ELISA or HPLC to determine the fumonisin content. Visual assessment of maize obtained from field trials revealed significant reductions in kernel injury and Fusarium ear rot in Bt versus non-Bt maize. Both HPLC and ELISA v measurements of maize grain demonstrated that fumonisin levels were significantly reduced in Bt as compared with non-Bt maize. Protection against multiple Lepidopteran pests provided by Cry1F or Cry1Ab x Vip3Aa resulted in the most noteworthy improvements in grain quality and reduced fumonisin levels. Maize obtained from these field trials was used to examine the impacts of grain quality and fumonisin contamination on fermentation outputs including ethanol and DDGS. Naturally and artificially contaminated maize samples ranging from 0-37 mg kg-1 total fumonisins were fermented, ethanol yield determined, and DDGS was collected and analyzed for fumonisin content. Ethanol yield was not affected by fumonisin contamination of maize in the range of concentrations examined. Fumonisins in DDGS derived from Bt hybrids averaged 2.04 mg kg-1 whereas those from non-Bt hybrids averaged 8.25 mg kg-1. Enrichment factors (DDGS fumonisin level/ground grain fumonisin level) did not differ significantly from 3.0 for 50 out of 57 hybrid x insect infestation treatment combinations. The seven that differed significantly from 3.0 were <3.0 (1.56 to 2.56x). The present study provides evidence for the efficacy of Cry1Ab, Cry1F, and Cry1Ab x Vip3Aa Bt maize reducing feeding from Lepidopteran insects ECB, CEW, and WBC, which resulted in indirect fumonisin reductions in Bt maize compared with non-Bt maize. The present study also provides laboratory-scale validation for the industry assumption of three-fold enrichment of fumonisins in DDGS, with fumonisin measurements traceable to individual samples. 1 CHAPTER I INTRODUCTION Modified from a paper to be submitted to a journal yet to be determined Erin Bowers 1 and Gary Munkvold 2 1Primary author, 2Major Professor Dissertation Organization This dissertation begins with a literature review describing the impacts of transgenic, insect-resistant maize (Bt maize) on insect feeding, fungal contamination, and Fusarium mycotoxin content of grain. Following the literature review are three manuscripts detailing
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