DON Occurrence in Wheat in US From 2003 - 2014: > 42,100 wheat samples from various production areas 81.1% of samples were < LOQ (0.3-0.5 mg/kg) Average around 0.5 mg/kg Highest level at 20 mg/kg

Global DON Occurrence From more than 30 publications (including 30 countries) Average DON levels similar to those reported in US Except for Africa: data from Morocco, Tunisia and Kenya Most of contamination: Tunisia (average 18 mg/kg) It is very difficult to directly compare the degree of DON occurrence in North American grains with data from around the world as different sampling schemes, sample preparation techniques, samples types, and analytical methods are used. These papers mostly included field samples of recently harvested grain, with a small percentage of the samples being described as commercial samples.

Global DON Occurrence Republic of South Africa (2009 - 2014) Corn and wheat samples 36 grain production areas 9.5 million tonnes/season for maize 1.85 million tonnes/season for wheat Courtesy of the Maize Trust and the Winter Cereal Trust (African industry organizations) via SAGL.

Effects of Growing Regions on DON in Wheat In the US DON associated with climatic conditions of regions During 2003 – 2014: Soft wheat: 19% US wheat production Highest levels in Midwestern and Northeastern regions Hard wheat: 65% US wheat production Highest levels in Northeastern region For this graphs samples were collected using GIPSA-approved methods.

Effects of Growing Regions on DON in Wheat DON in hard red spring wheat in Canada associated with: Climatic conditions of regions Geographical distribution of DON-producing Fusarium Higher levels in eastern prairies of Manitoba For this graphs samples were collected using GIPSA-approved methods.

Effects of Growing Seasons on DON in Wheat Associated with weather conditions during production In the US during the period from 2003 to 2014: Midwestern and Northeastern regions in certain years: Up to 14% of hard wheat had DON > 2mg/kg Up to 62% of soft wheat had DON > 2mg/kg For this graphs samples were collected using GIPSA-approved methods. Northeast is more prone to variation from year to year, regardless of the type of wheat grown, while the Midwest seems to show more variation for certain classes. Higher levels of DON are usually associated with excessively wet periods close to the flowering stage (6-8 weeks prior to harvest).

DON Levels in Barley in US North Dakota and Minnesota (1993 – 2003) Average DON levels varied from 0.5 to 10.3 mg/kg In most years at least 1/3 of samples had > 3 mg/kg Variation from year to year in DON levels and incidence is evident and illustrates once again the volatility of the natural occurrence of such toxins. The paper shows a little bit of data for corn as well. With on average about 10% of samples showing levels above 2.0ppm. Schwarz et al., 2006.

DON: A Multi-Faceted Issue… Levels of DON in Grain Field Conditions (weather, location) Grain (type, variety) Agricultural Practices (fungicide, rotation) Supply Chain Management Processing (cleaning, milling)

DON: A Multi-Faceted Issue… Levels of DON in Grain Field Conditions (weather, location) Grain (type, variety) Agricultural Practices (fungicide, rotation) Supply Chain Management Processing (cleaning, milling)

Breeding Programs Severe epidemic of FHB in 1993 incentivized breeding for reduced DON Sources of resistance: Sumai #3 from China Native resistance in existing breeding germplasm In resistant varieties FHB severity and/or DON reduced by >50%

Breeding Programs Severe epidemic of FHB in 1993 incentivized breeding for reduced DON Growers have been adopting new cultivars In the HRS wheat growing region: 1999 FHB-susceptible cultivars 76% frequency of FHB-susceptible cultivars sown decreased from 76% in 1999 to 21% in 2011. 2011 FHB-susceptible cultivars 21%

Best Agricultural Practices Crop rotation Improved cultivars Fungicide application at flowering Fungicide efficacy for Fusarium head blight (FHB) index, deoxynivalenol (DON), Fusarium-damaged kernels (FDK), and yield in moderately resistant and susceptible wheat cultivars. Within each variable, least significance difference (LSD) values were calculated at P = 0.05. (Adapted, with permission of The American Phytopathological Society, from Wegulo et al. [84].) Wegulo et al., 2011.

Best Agricultural Practices Fungicide application at flowering TEBU - Tebuconazole PROP - Propiconazole PROT - Prothioconazole METC - Metconazole Efficacy of various fungicides for reducing FHB in the field and suppressiong DON production in harvested grain expressed as percent control compared with untreated wheat. McMullen et al., 2012

Web-Based Disease Forecasting Tools Based on hundreds of weather stations Advises growers of the need for timely application of fungicides Warnings about regions with potentially unacceptable levels of DON Provide strategies to grain buyers to manage potential problems Forecasting systems widely used in the spring and summer in all regions as indicator of FHB likelihood.

Web-Based Disease Forecasting Tools Available tools: Fusarium Risk Assessment Tool FHB Alert System Weather Central Forecasting systems widely used in the spring and summer in all regions as indicator of FHB likelihood.

DON: A Multi-Faceted Issue… Levels of DON in Grain Field Conditions (weather, location) Grain (type, variety) Agricultural Practices (fungicide, rotation) Supply Chain Management Processing (cleaning, milling)

Supply Chain Management The occurrence of DON in wheat taken at earlier stages in the handling chain has also been observed in Canadian wheat (Fig. 4) and durum (Fig. 5). DON was measured in 72% and 68% of wheat and durum samples, respectively, taken on farm and in 58% and 79% of wheat and durum samples, respectively, taken from primary elevator stocks before final blending of grain to meet quality specifications was performed. At later stages of the handling chain, <50% of wheat and Durum samples contained quantifiable concentrations of DON. There were some instances of high concentrations of DON (i.e., >5 mg/kg) observed in individual Canadian samples, as with USA samples (Figs. 2 and 3). These instances occurred in samples taken from on farm, primary elevator stocks, or elevator loadings but not at later stages in the grain-handling chain. The average DON concentrations in exported Canadian wheat and durum were both <1 mg/kg. Similar trends are observed with durum wheat and barley along the Canadian grain-handling chain. Schematic of the North American grain-handling supply chain.

Supply Chain Management Deoxynivalenol (DON) occurrence in Canadian wheat along the grain-handling chain. The occurrence of DON in wheat taken at earlier stages in the handling chain has also been observed in Canadian wheat (Fig. 4) and durum (Fig. 5). DON was measured in 72% and 68% of wheat and durum samples, respectively, taken on farm and in 58% and 79% of wheat and durum samples, respectively, taken from primary elevator stocks before final blending of grain to meet quality specifications was performed. At later stages of the handling chain, <50% of wheat and Durum samples contained quantifiable concentrations of DON. There were some instances of high concentrations of DON (i.e., >5 mg/kg) observed in individual Canadian samples, as with USA samples (Figs. 2 and 3). These instances occurred in samples taken from on farm, primary elevator stocks, or elevator loadings but not at later stages in the grain-handling chain. The average DON concentrations in exported Canadian wheat and durum were both <1 mg/kg. Similar trends are observed with durum wheat and barley along the Canadian grain-handling chain. Deoxynivalenol (DON) occurrence in Canadian barley along the grain-handling chain.

DON: A Multi-Faceted Issue… Levels of DON in Grain Field Conditions (weather, location) Grain (type, variety) Agricultural Practices (fungicide, rotation) Supply Chain Management Processing (cleaning, milling)

Processing: Cleaning and Milling Wheat cleaning and milling process diagram. Cleaning steps in green boxes indicate where reduction of deoxynivalenol (DON) typically occurs (129). * Wheat “Red Dog” consists of the offal from the “tail of the mill” together with some fine particles of wheat bran, germ, and flour. This product must be obtained in the usual process of commercial milling and must contain not more than 4% crude fiber.

Processing: Cleaning Effect of cleaning on removal of deoxynivalenol (DON) from grain.

Processing: Milling Effect of milling on removal and distribution of deoxynivalenol (DON) from grain

Processing: Cleaning and Milling Savi et al., 2016. Deoxynivalenol levels (mg kg/1) in the wheat milling fractions of the positive samples. Statistically significance compared with milled wheat **P <0.01, ***P <0.001,according to ANOVA and Bonferroni post-test. Deoxynivalenol (DON) distribution (%) in milled wheat fractions (total DON amount in cleaned grains = 100%). (Derived from data presented in Tables 3 and 4.)

Are we at risk? “Maximum Levels (MLs) are maximum concentrations of a specific substance recommended in a specific commodity or product, providing guidance to manufacturers and consumers.” MLs are established when the contaminant: May be found in amounts that result in adverse public health impact In certain products has a significant contribution to total dietary exposure Wheat Barley Corn

Maximum Tolerable Daily Intake Daily Food Consumption Are we at risk? Maximum Tolerable Daily Intake (μg toxin/ BW in Kg) Maximum Levels for Consumption [Mycotoxin in food] (μg toxin/Kg food) Daily Food Consumption (g food/ BW in Kg)

Are we at risk? Comparison between corrected maximum levels (MLs) from 2006 WHO Global Environment Monitoring System/Food Contamination Monitoring and Assessment Programme (GEMS/Food) consumption data, 2012 WHO GEMS/Food consumption data, and proposed MLs in the Codex Committee on Contaminants in Foods (CX/CF) 12/6/9 (166) as discussed in their February 2012 meeting.

Commodity/ Product Name Maximum Level (ML; mg/kg) As of 2015… Commodity/ Product Name Maximum Level (ML; mg/kg) Notes/Remarks Cereal-based foods for infants and young children 0.2 For sampling plan, see Annex. All cereal-based foods intended for infants (up to 12 months) and young children (12 to 35 months) Flour, meal, semolina and flakes derived from wheat, maize or barley 1.0 For sampling plan, see Annex. Cereal grains (wheat, maize and barley) destined for further processing 2.0 “Destined for further processing” means intended to undergo an additional processing/treatment that has proven to reduce levels of DON before being used as an ingredient in foodstuff, otherwise processed or offered for human consumption. Codex members may define the processes that have been shown to reduce levels.

Potential Impact… For these reasons, the direct impacts of establishing MLs on DON would be on domestic mills, while there would be indirect impacts on growers and exports and imports (discussed below). In response to this proposed regulation, domestic mills could be impacted most directly, although an increase in cost would impact other sectors in the supply chain as well.

Potential Impact… Changes in strategy: Limited supply? Wheat with greater assurance of conformance to MLs Sourcing wheat from certain production regions Increased cleaning Imposing stricter contract limits/ price differential Limited supply? Due to changes in purchasing strategy - domestic mills may be affected in grain-exporting countries Importers of grains, at least from North America, also have tools available for controlling DON content. In concept these are similar to those of domestic mills and include primarily contract limits for DON content. Additionally, these can include targeting locations, ports, and supplier that have the ability to reduce DON content in years when it is problematic. This is most commonly accomplished by targeting origins and cleaning at the country and export elevator prior to shipment, ultimately to meet specification limits set by importers. It is clear from these strategies, that importers can effectively limit their DON content through contract limits. Of course, tighter controls would accrue greater costs due to testing and potential rejections. Meeting these specifications could impact the available supply of wheat. Finally, importers would also be impacted indirectly. If stricter MLs are imposed, the strategy of domestic mills in grain-exporting countries would change and could reduce (slightly) the available supply of DON-compliant wheat to the export market. For this reason, there would be a minor component of crops with unacceptable (even if precautionary) DON levels that would be channeled elsewhere in the market system. This would include the export market or markets for nonfood uses. For importers, this could increase the risks of receiving shipments containing DON at unacceptable concentrations and, as a result, would require more scrutiny of contract limits and price differentials. The response would be to increase the intensity of cleaning and targeting of origins for Grain prior to exporting, which also would affect costs.

Potential Impact… Change in strategies: Wheat with greater assurance of conformance to MLs Sourcing wheat from certain production regions Imposing stricter contract limits Larger discounts Add preprocessing functions to ensure desired MLs Testing Added cleaning Mills and first processors would have to change their strategies. Mills would seek to purchase wheat with greater assurance of conformance to the MLs and/or add preprocessing functions to assure meeting MLs for DON. This could include a number of strategies: sourcing wheat from production regions that typically experience lower incidence of Fusarium and lower DON levels (where possible); imposing stricter limits on wheat and/or larger discounts for excessive DON; requiring more extensive cleaning prior to shipment from the elevator; and segregation, testing, and cleaning at the mill to assure ML conformance. Ultimately, this would add costs to both purchasing and processing. These added costs would vary through time and geographically. In-mill costs of cleaning, scouring, added storage, etc. would also vary through time but, nevertheless, would be a real added cost to the milling sector. Since such MLs currently are not applied to unprocessed raw grains, major processors have not yet quantified the costs associated with such proposed regulations. To do so would require detailed simulation modeling of these functions and risks. Ultimately, these costs would vary by location and be determined by logistics and the mill’s capability for segregation and cleaning.

Potential Impact… More intensive management: Variety selection Fungicide use Shift to other crops Less risk of DON Crops with greater returns As previously discussed, growers currently seek to control DON through crop rotation strategies, adoption of improved cultivars, and use of fungicides and optimal spray technology at flowering. Tighter MLs on DON would have the following impacts: more intensive management, including variety selection and fungicide use, all of which will add costs. Larger discounts for excessive DON and greater risk for wheat and other crops grown with excessive DON, would lead to a shift to other crops with less risk or greater returns. These impacts would vary geographically and through time. It is important to note that as the risk associated with wheat production increases and concurrently competing crops become more viable and profitable there would be a move away from production of wheat and other affected crops in the USA, Canada, and elsewhere. This likely would reduce supplies and increase prices to all consumers. Importers of grains, at least from North America, also have tools available for controlling DON content. In concept these are similar to those of domestic mills and include primarily contract limits for DON content. Additionally, these can include targeting locations, ports, and supplier that have the ability to reduce DON content in years when it is problematic. This is most commonly accomplished by targeting origins and cleaning at the country and export elevator prior to shipment, ultimately to meet specification limits set by importers. It is clear from these strategies, that importers can effectively limit their DON content through Contract limits. Of course, tighter controls would accrue greater costs due to testing and potential rejections. Meeting these specifications could impact the available supply of wheat. Finally, importers would also be impacted indirectly. If stricter MLs are imposed, the strategy of domestic mills in grain-exporting countries would change And could reduce (slightly) the available supply of DON-compliant wheat to the export market. For this reason, there would be a minor component of crops With unacceptable (even if precautionary) DON levels that would be channeled elsewhere in the market system. This would include the export market or Markets for nonfood uses. For importers, this could increase the risks of receiving shipments containing DON at unacceptable concentrations and, as a result, Would require more scrutiny of contract limits and price differentials. The response would be to increase the intensity of cleaning and targeting of origins for Grain prior to exporting, which also would affect costs.

DON: A Multi-Faceted Issue… Levels of DON in Grain Field Conditions (weather, location) Grain (type, variety) Agricultural Practices (fungicide, rotation) Supply Chain Management Processing (cleaning, milling)

DON Occurrence in Grains: A North American Perspective. Acknowledgments DON Occurrence in Grains: A North American Perspective. CEREAL FOODS WORLD. JANUARY–FEBRUARY 2015, VOL. 60, NO. 1