Genetic Opportunities to Enhance Sustainability of Pork Production in Developing Countries: A Model for Food Animals

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1 Genetic Opportunities to Enhance Sustainability of Pork Production in Developing Countries: A Model for Food Animals Cecil Forsberg, S. Golovan, A. Ajakaiye, J.P. Phillips, R.G. Meidinger, M.Z. Fan, J.M. Kelly and R.R. Hacker The University of Guelph Guelph, Ontario CANADA

2 Trends in Pig Production in Developed and Developing Countries (FAO Stats, 2003) Live Pigs (Millions) Year Developed Countries Developing Countries Linear (Developing Countries) Linear (Developed Countries)

3 Reported Transgenic Pig Projects Gene Tissue Effect Growth hormone. All. Enhanced growth IGF1...Muscle. Muscle growth Plant oleate desaturase. All tissues Improved nutritional value (for human) Lactoferrin Mammary Resistance of piglets gland to infection α-lactalbumin. Mammary Improved piglet gland growth Phytase. Salivary.. Efficient P Utilization gland Reduced P in manure

4 Eutrophication & Fish Mortalities Caused by Manure Contamination PO 4 3- Run-off, tiles N 2 N 2 O Anoxia Algal Toxins

5 Phytate Phosphorus Content of Major Cereal Grains and Protein Supplements O - P- O O - O= - O -P O O O= O - O= P- O - H H O O - O= P-O - O O H H - O -P O O -= H O - O -P O O -= Foodstuff Total P(%) Phytic acid (% of Total P) Barley Corn Wheat Soybean meal Cottonseed meal Sesame meal Wheat bran Phytic acid Nelson et al. Poultry Sci. 47:

6 Action of Phytase + + O Ca O O= P-O O-P= O O O O Protein O= P-O H -+ H O CH O Mg 2 O= P-O - + NH+ O H H 3 O -O-P= O O H O O-P= O OH Starch Phytate + Ca + CH 2 0H Starch P. C. M. Simons et al. Brit.J.Nutr. 64: , H O 2 O = O-C-CH -Protein 2 Phytase H OH OH H H OH H H OH Inositol OH H + 6 H 3 PO 4 Pigs U Phytase/kg feed 24% available P 35% fecal P + Protein Starch Minerals

7 How the Enviropig TM Works Phytase produced in the salivary glands and secreted in the saliva Liver Pancreas Cecum Indigestible Phytate phosphorus in the diet Stomach Phytase Duodenum Phytate Phosphate Small Intestine PHOSPHATE ABSORPTION Colon Manure low in Phosphorus Phytate Passes through the non-transgenic pig Unchanged Phytate is digested in the Enviropig stomach

8 Important Regions of the plama2/appa Plasmid Introduced into Embryos to Create the Enviropig TM Constitutive Promoter Mouse-PSP Promoter E. coli APPA coding region Phytase gene Not I ori plama2/appa with pbluescript II plasmid backbone 20.6 kb Segment deleted prior to injection of transgene β-lactamase coding region Xho I

9 Sampling A Pig For Phytase in the Saliva A 50 kg grower pig produces 0.5 L of saliva when consuming 0.5 kg dry feed

10 Founder and First Generation Enviropig TM Founder Founder First Generation Transgenic Piglets

11 Different Enviropig TM Lines JA Wks 2 nd Gen 160 Kg, June 11/02 Le Wks 1 st Gen. 140 Kg GO Litter Wks 3 rd Gen June 11/02

12 Conventional Diets Fed to Phytase Pigs Lack Supplemental Phosphorus Ingredient Weaner Grower Finisher Corn Barley Wheat Soybean meal Fish meal Whey Dical-PO 4 Nil Nil Nil CaCO

13 Relationship Between Salivary Phytase Activity and Fecal-P Among Non-Tg and Tg Pigs Fecal Phosphate (% of Dry Matter) Control (Non-transgenic) n=18 0 Manon Tg n=10 Gordie Tg n=10 Jacques Tg n= Phytase Activity (U/ml of saliva) Wayne Tg n=10 (45% ) (57% ) (64% ) (64% )

14 Phytase Activity in Saliva of Different Generations of the Cassie Line of Phytase Pigs Activity (U/ml) n=1 F -Founder n=1 n=10 n=1 n=7 F G 1 G 2 G 3 Generation Males Females n=17 n=20 (Assayed morning and evening of day 11)

15 Reproductive Viability of The Cassie Line as Compared to Non-Transgenic Yorkshire Pigs Number / Litter Live Births Stillborns Category Yorkshire Cassie Tg Early Deaths Mummies

16 Offspring From a Homozygous Outcross to Non-Tg BoarL JA173, 5 wk, 6 kg, 3 rd Generation (Documents the generation of a homozygous line of pigs suited for breeding)

17 Nigeria 6 million pigs 97% pigs free range Free Range Pigs Plant materials, scraps and soil Diets Managed Pigs Energy -Corn & cassava Protein supplement -Soybeans meal/cake or peanut meal Phosphorus -Bone meal (Primarily from cattle) No suppl. Phytase!

Brazil 30 million pigs 15 million in other SA countries Pigs usually on large farms Diet Corn (60%) & Soybeans (40%) Supplemental P-either dicalcium P0 4 or phytase Santa

18 Brazil 30 million pigs 15 million in other SA countries Pigs usually on large farms Diet Corn (60%) & Soybeans (40%) Supplemental P-either dicalcium P0 4 or phytase Santa Catarina State 85% of water sources contaminated with fecal organisms due to spreading manure and runoff into streams, ponds and rivers etc. Brazilian legislation forbids GM livestock.

China 465 million pigs 68% meat production Up to 80% pigs raised in back yards Diet corn & soybean meal Other protein supplements peanut meal, rapeseed meal, cottonseed meal, linseed

19 China 465 million pigs 68% meat production Up to 80% pigs raised in back yards Diet corn & soybean meal Other protein supplements peanut meal, rapeseed meal, cottonseed meal, linseed meal, wheat shorts, rice bran (all high phytate). Phosphorus low quality mineral PO 4 or phytase - Bone meal has been banned Serious P & N pollution in some locations (eg. Lake Taihu)

20 Predicted Relation Between Increased Supplemental Phytase and Reduction in Phosphorus Excretion % Reduction in P Excretion % Reduction attained by Enviropigs $1.33 $2.66 $5.33 $3.99 $ Supplemental Phytase (U kg -1 ) Prediction developed from data by Kornegay, E.T P247 Enzymes in Farm Animal Nutrition, M.R. Bedford and G.G. Partridge

21 Effect of Reducing the Crude Protein Content of the Diet on Reduction of Phosphorus in the Fecal Matter From Enviropigs Assumptions: 60% redn in fecal phosphorus % Reduction in Acreage for Spreading Manure One % redn in diet CP P-limit 10% redn fecal N % Reduction of Crude Protein in the Diet Note: PO 4 3- is the primary nutrient causing eutrophication followed by nitrogen

22 Competing Technologies Commercial Phytase Enzymes Requires continuous addition to all diets with feed mixing and preferably pelleted. All phytase to date are partly heat labile and subject to destruction during pelletting. Corn Kernels smaller, 15% yield reduction. Reduced germination. Soybean meal 25% lower yield. Barley Reported good germination and no yield redn, but agronomic characteristics not tested yet.

23 Regulatory Hurdles Canada: Environment Canada Canadian Environmental Protection (Act Schedule 19) -Document safety to environment including wild animals, flora and fauna Health Canada Novel Foods Act -Safety of food for human consumption Canadian Food Inspection Agency -Animal well-being, feed and fertilizer acts. United States: Food and Drug Admin. (Center for Veterinary Med.) -Animal drug EPA USDA

24 Pre-Market Safety Assessment of Food Derived from Genetically Modified Plants (a) Safety of the inserted DNA (potential for transfer to other organisms) (b) Safety of DNA ingested (potential toxicity) Food safety issues of the newly produced protein: (a) Potential for toxicity of the novel protein (b) Potential for allergenicity of the novel protein (c) Safety of any unintended effects Equivalence of composition to the original food Retention of nutritional value The human dietary exposure No well worn path for food animals!

25 Conclusions The Enviropig TM Documented healthy Enviropigs that digest phytate phosphorus efficiently. Major pig breeding companies and other potential investors are concerned by the uncertain time line for societal acceptance of pork from genetically modified animals. However, there is growing global societal acceptance of GM products. Environment Issues Remain World mineral phosphate sources are being depleted. Bone meal is no longer used in pig diets in a growing number of countries due to the concern of disease transmission (BSE). Phytase enzyme issues availability, mixing, pelleting, storage and heat stability? Low phytate crops germination & yield not fully resolved. None adapted to tropical or subtropical climates?

Thank you Research Support Ontario Pork Producers Marketing Board Natural Sciences & Engineering Research Council of Canada Agriculture & Agri-Food Canada Ontario

26 Thank you Research Support Ontario Pork Producers Marketing Board Natural Sciences & Engineering Research Council of Canada Agriculture & Agri-Food Canada Ontario Ministry of Agriculture & Food Food Systems Biotechnology Centre U of G Mars Landing Program

J.A. Lory 1, R.E. Massey 2 and M.C. Shannon 3 1

J.A. Lory 1, R.E. Massey 2 and M.C. Shannon 3 1 Plant Sciences, 2 Agricultural and Applied Economics, 3 Animal Science University of Missouri, Columbia MO 65211 Software is commonly used to optimize diets