IN VIVO DIGESTION OF GENETICALLY MODIFIED PROTEINS IN THE CHICKEN AND FATE OF SUCH PROTEINS

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1 IN VIVO DIGESTION OF GENETICALLY MODIFIED PROTEINS IN THE CHICKEN AND FATE OF SUCH PROTEINS Dr. Sheila E. Purdum, Professor University of Nebraska, Lincoln, Nebraska

2 PROTEIN DIGESTION REVIEW Function of the Gut to digest proteins Metabolism of Peptides and amino acids post absorption

3 AVIAN GASTROINTESTINAL SYSTEM

4

5

6 DISPOSAL OF PROTEIN N AA in surplus to body s requirement are degraded Liver is major site of AA degradation N disposal can be divided into two main stages Removal of amino group from AA Formation of uric acid or urea

7 URIC ACID METABOLISM Taken from Nutrition of the Chicken by Scott, Nesheim and Young, 1976

8 GENETICALLY MODIFIED PROTEINS (GMO S) GMO s are just another protein as far as the chicken s gut is concerned and will be digested and metabolized by normal processes in the avian

9 Objective To test presence of the CP4 EPSPS protein from genetically modified protein in Roundup Ready soybean from source (soybeans) to tissue and eggs in the laying hen STUDY ONE. FATE OF GMP FROM ROUNDUP READY SOYBEANS IN LAYING HENS

10 SAMPLES COLLECTED 2 commercial egg producers Whole soybeans (Round-up Ready) Soybean meal (from Round-up Ready Soybeans) Layer diets Whole egg, egg albumen Hen liver and excreta

11 TESTING FOR GENETICALLY MODIFIED PROTEIN Immunoassay with antibodies specific to CP4EPSPS, coupled to a color reagent and incorporated into a lateral flow strip to produce an antibody sandwich around the modified protein if present Accurate to 0.1% of the CP4 EPSPS protein Verification by ELISA by an independent laboratory

12 TABLE 1. IMMUNOASSAY TEST RESULTS FOR THE CP4 EPSPS PROTEIN FROM ROUNDUP READY SOYBEANS IN FEED AND POULTRY TISSUES (ASH ET.AL., 2003) Company 1 Company 2 Rep 1 Rep 2 Rep 1 Rep2 RR Soybeans Yes Yes Yes Yes Soybean Meal Yes Yes Yes Yes Complete Diet Yes Yes Yes Inconclusive Whole Egg No No No No Egg Albumen No No No No Liver No No No No Feces No No No No

13 SUMMARY FOR STUDY ONE Roundup Ready soybean GMO protein is detectable by ELISA tests in SBM and layer diets at concentrations > 0.1% No Roundup Ready soybean protein was detected in whole egg, albumen, liver or excreta

14 Objective Two trials were conducted to assess the fate of the Cry3Bb1 protein from YieldGard rootworm corn (MON 863) when fed to laying hens. In the first trial, 2 diets, 1 formulated with MON 863 and 1 with conventional corn, were fed to laying hens (12 replicate cages with 4 hens/cage per treatment) for 8 wk. STUDY TWO: IN VIVO DIGESTIVE FATE OF CRY3BB1 PROTEIN IN LAYING HENS FED MON 863 CORN

15 INTRODUCTION TO MON 863 CORN The MON 863 plants are resistant to the larval feeding damage of the coleopteran insect, corn rootworm (Coleoptera, Chrysomelidae, Diabrotica sp.). The digestibility of Cry3 proteins has been thoroughly evaluated in vitro (OECD, 2007) and as part of the safety assessment requirements for product registration with the US Environmental Protection Agency and Food and Drug Administration. The Cry proteins, including Cry3Bb1, are unstable in simulated gastric conditions and are rapidly degraded by pepsin (FSANZ, 2005).

16 EXPERIMENTAL DESIGN MON 863 and its near-isogenic control were obtained from Monsanto Company (St. Louis, MO) and shipped to the University of Nebraska. Before the start of the study, MON 863 and control corn were sampled and checked for Cry3Bb1 protein using Cry3Bb1 lateral flow test strips Trial 1. Feces were collected 1 d before the start of study (designated as time zero), and then feces and eggs were collected at 4 and 8 wk for a 24-h test period and then saved for Cry3Bb1 protein analysis. At the end of the 8- wk feeding period, 12 hens per treatment were killed and hepatic and breast tissue samples were taken and tested for the presence of Cry3Bb1 protein.

17 TABLE 2. NUMBER OF SAMPLES POSITIVE FOR CRY3BB1 PROTEIN IN FECAL, LIVER AND BREAST TISSUE (12 SAMPLES) - TRIAL 1. Feces Liver Breast Time 0 4 wk 8 wk Time 0 4 wk 8 wk Time 0 4 wk 8 wk Control Test

18 TRIAL 2 MEASUREMENTS After 7 d, 6 hens per treatment were sampled for blood, killed by cervical dislocation, and surgically eviscerated. Immediately after collection, test strips were inserted directly into the blood tubes. The crop, small intestine, cecum, and large intestine were excised from each hen, and digesta samples were taken from each section. Digesta samples were tested for the presence of Cry3Bb1 protein using Cry3Bb1 lateral flow test strips. The remaining birds were sampled and tested, as described, at the end of the 4-wk period.

19 FIGURE 1. AMOUNT OF TRYPSIN-RESISTANT CORE CRY3BB1 DETECTED PROTEIN IN FECAL EXTRACTS

20 Objective: To compare Herculex *1 (TC1507) Corn from Pioneer Hy-Bred to its Iso-line equivalent and 2 other conventional corn sources in the diets of laying hens over a 4 month time period STUDY 3. AMOUNT OF NUTRITIONAL EQUIVALENCY OF CORN GRAIN FROM DAS (HERCULEX *1) IN DIETS OF LAYING HENS

21 TABLE 3. DIETARY CORN TREATMENT EFFECTS ON PERFORMANCE VARIABLES Diet Corn Type Feed Intake g/hen/day Egg Prod % Feed Conv. (g egg/gfeed) Hen Wt. Gain grams 1 Isoline TC Conv Conv p-value NS

22 TABLE 4. DIETARY CORN TREATMENT EFFECTS ON EGG VARIABLES Diet Corn Type Egg Wt. grams Haugh Unit Roche Color Score 1 Isoline TC Conv Conv p-value NS

23 SUMMARY 1. Roundup Ready soybean GMO protein is detectable by ELISA tests in soybean meal and complete diets fed to layer chickens at concentrations above 0.1%. 2. No Roundup Ready soybean GM protein was detectable in whole egg, egg albumen, liver, or excreta tissue. These results were confirmed by two separate labs for egg protein. 3. The laying hen is efficient at digesting soybean meal protein and corn GMO to a stage at which no detectable intact GMO material is absorbed or deposited in tissues such as liver and eggs. 4. GMO corn is nutrient equivalent to conventional in value and performance in laying hens.

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