GM-Plants for Animal Feeds: Opportunities and Challenges. Gerhard Flachowsky, Institute of Animal Nutrition, Braunschweig, Germany

Transcription

1 GM-Plants for Animal Feeds: Opportunities and Challenges Gerhard Flachowsky, Institute of Animal Nutrition, Braunschweig, Germany FAO-APHCA Regional Workshop, Bangkok, August 2013

2 Table of Content 1. Introduction 2. Challenges for Plant Breeding 3. Fundamentals of Animal Feeding Studies 4. GM-Plants of the 1st Generation (Plants with Input Traits) 5. GM-Plants of the 2nd Generation (Plants with Output Traits) 6. Conclusions

3 1. Introduction (Global Developments) Population Resources per people Emissions

4 Institute of Animal Nutrition Challenges for animal production or Livestock s long shadow (Steinfeld et al. 2006) Year Presently 2050 Percent to presently World population (bill.) Meat production (Mio. t) Milk production (Mio. t)

5 Examples of important limited global resources Area 1,5 Bill. ha arable land; 3,3 Bill. ha Grassland Fossil Energy C-Sources (e.g. petrol, gas, coal) Water Presently about 71% used by agriculture Phosphorus Known resources for about years

6 2. Challenges for Plant Breeding

7 Rate of change Keeping pace with climate change (by Whitford et al. 2010) Effects of climate change Technology gap Conventional breeding solutions Time

8 Conditions to produce phytogenic biomass and their availability per inhabitant under consideration of the increase of population ( Increase, Decrease, No important influence; based on the Royal Society, 2009) Plant nutrients in the atmosphere (N 2, CO 2 ) Sun energy Agricultural area Water Fossil Energy Mineral plant nutrients Variation of genetic pool

9 Basic wishes to plant breeders incl. plant biotechnology 1. Higher yields and stability through competition for land, climate change, etc. 2. Efficient use of resources with limited availability such as: water crude mineral resources (ie phosphorus) fossil energy (ie N fixation by plants) and use of smaller areas of land, which will require: higher energy yields per limited resource higher resistance to plant and animal pests as well as abiotic factors (ie drought, salinity); and improved storage and preservation characteristics (suitability for ensiling and high aerobic stability of silage) 3. More efficient use of unlimited resources (e.g. sun energy, N 2, CO 2, genetic pool) 4. Changes in composition (more valuable/less undesirable substances)

10 Mio. ha Soybean Cotton Maize Rapeseed Year Global area of transgenic crops (GMP) in Mio. ha (ISAAA 2013)

11 Institute of Animal Nutrition Nutritional view of GM-Crops - Definitions - GMP of the 1 st generation Plants with agrotechnical traits (input traits) Without substantial changes in composition/nutritive value; substantial equivalent Examples: Bt-plants (corn, cotton), RR-plants (soybean), PAT-plants (corn, roots) etc. GMP of the 2 nd generation Plants with output traits With substantial changes in composition/nutritive value; not substantial equivalent Examples: Golden rice, low phytate corn, changes in fatty acids (e.g. MON 87769) or amino acids pattern etc.

12 Institute of Animal Nutrition Substantial Equivalence (S.E.) Quo vadis? S.E. (OECD,1993) embodies the concept that if a new food/feed is found to be substantially equivalent to an existing food/feed, it can be treated in the same manner with respect to safety as its traditional counterpart. Problems Coarse frame work for evaluation; limited safety and nutritional assessment What means traditional counterpart (isogenic plants or local average) Irrational if changes in composition (e.g. lower content in mycotoxins, GMP of the 2 nd generation) Proposal Various compartments should be considered as parts of S.E. as Chemical/analytical equivalence Nutritional equivalence Safety equivalence Exposure equivalence

13 Important food/feed from GM-plants and estimated portion used as food or feed GM-Plant Food % Feed % Soybean Oil, Proteins 25 Soybean (extracted oil) meal, Full fat soybean 75 Maize Starch, maize meal, Oil 15 Maize, Oil, DDGS, Gluten feed, Silage, Straw, 85 Rapeseed Oil 25 Rape seed (extracted oil) meal, Rapeseed expeller/cake, Fullfat rapeseed Cotton Oil 15 Cotton seed (extracted oil) meal, Expeller 75 85

14 Institute of Animal Nutrition Questions to Nutritionists Nutritional and safety assessment of food/feed of the 1 st generation of GMP (Plants with input traits) Nutritional and safety assessment of food/feed of the 2 nd generation of GMP (Plants with output traits) Influence of GM-food/feed on human/animal health and quality of food of animal origin Studies on the degradation of newly expressed protein, foreign DNA, unintended effects etc.

15 Risk Assessment means: Assessment of Safety for Man, Animal and Environment How can animal feeding studies to improve safety for man, animal and environment?

16 Events in commercial GM - crops and pipelines worldwide, by trait (Stein and Rodriguez-Cerzo, 2009) Trait category Commercial in 2008 Commercial pipeline Regulatory pipeline Advanced development Total by 2015 Insect resistance Herbicide tolerance Crop composition Virus resistance Abiotic stress tolerance Disease resistance Nematode resistance Fungus resistance o Other Note: The number of trails can be bigger than the number of GM crops; Abiotic stress tolerance includes drough tolerance

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18 Future tendencies (EFSA 2009, Stein and Rodriguez-Cerzo, 2009) Combination of some stacks in one hybrid Improvement of nutritional properties (e.g. amino acids, fatty acids, vitamins, minerals, enzymes, lower content of undesirable components) More efficient use of resources (e.g. water, fuel, N)

19 3. Fundamentals of Animal Feeding Studies

20 Objectives for Animal Feeding Trials (in vivo) with GM- Products Do we need additional information to compositional analysis and in silico, in vitro and/or in situ studies? Do we expect unindented effects? Do we expect long term effects (incl. effects on health, fertility etc.)? Do we need additional information from GMP with output traits (e.g. bioavailability of nutrients) or GM-animals (e.g. new energy and nutrient requirements)? Do we expect effects on product quality/composition of food of animal origin? Can we expect innovations for safety and nutritional research? Are animal studies important/necessary from the view of public concerns and/or may the studies contribute to more public acceptance? Present recommendation: Case by case decision

21 Advantages of studies for safety and nutritional assessment of GM-plants derived food/feed with laboratory or target animals Laboratory Animals Internat. agreed study protocols Small amounts of feed, higher number of repetitions Lower costs for feed and equipment Target Animals Representative for animal species/categories (extrapolation??) Higher amounts of GM-products are fed to animals All control animals fed with comparators (isogenic, commercial) are available for the market Real studies on the transfer of valueable and/or undesired substances into food of animal origin

22 Important types of feeding studies and animals recommended Type of studies Lab. Animals Target Animals Testing of single substances (28 day study) 90-day rodent feeding study Long-term feeding study X X Multigeneration feeding study Determination of digestibility/availability Efficiency study Tolerance study Studies with GM-animals X X X X X X X X X

23 Influencing Factors on Type of Feeding Studies Scientific question(s) Plants of 1st or 2nd generation Availability of GM-feeds (esp.in early stages of breeding) and adequate comparators Financial budget Availability of equipments, animals and qualified manpower

24 Examples of life spans for target animals (growing/fattening animals, in days) Animal species/categories Chickens for fattening (broilers) Conventional/More intensive Organic/More extensive Turkeys for fattening Growing/fattening pigs Veal calves Growing/fattening bulls Laying hens and dairy cattle are usually used for longer periods: Laying hens: about days for growing (pullets); about days (one year) for the laying period. Dairy cattle: about month for growing (heifers); one to ten years for lactation (average in Europe two to five lactations).

25 Institute of Animal Nutrition Some recommendations from the Best practices for the conduct of animals studies to evaluate crops genetically modified for input traits (GMP of the first generation), (adapted from ILSI 2004 and EFSA 2007) Animals ( species / categories ) Poultry for meat production Poultry for egg production Swine Growing and finishing ruminants Number of animals ( coefficient of variation 4 to 5 %) 10 to 12 pens per treatment with 9 to 12 birds per pen 12 to 15 replications per treatment with 3 to 5 layers per pen 6 to 9 replications per treatment with 4 or more pigs per replication 6 to 10 replications per treatment with 6 or more cattle per replication Duration of experiments 5 weeks or more 18 to 40 weeks of age, at least three 28 - day phases Piglets (7 12 kg), 4 6 weeks Growers (15 25 kg) 6 8 weeks days Composition of dies 1 Balanced diets Balanced diets Balanced diets Balanced diets Measurements Feed intake, gain, feed conversion Feed intake, egg production, feed conversion, egg quality Feed intake, gain, feed conversion, carcass quality Feed intake, gain, feed conversion, carcass data Lactating dairy cows cows per Latin square : 28 day Balanced diets Feed intake, milk treatment periods performances and 28 cows per Randomized block composition, body treatment design weight, Body Condition Score (BCS), cell counts in milk, animal health 1 Feed from GMP should be included in high portions in diets and compared with isogenic counterparts

26 Endpoints of Feeding Studies Feed intake, body weight and weight gain, yield in milk or eggs, feed conversion rate (FCR) Animal behavior, health, mortality Physiological parameters in body samples Weight of organs and tissues Chemical composition of food of animal origin (e.g. milk, eggs, tissues, organs)

27 Proposal of endpoints for comparative analysis and further safety relevant studies of food/feed from GM-plants/animals Group of GM-animals Mammals Birds Aquaculture (e.g. fishs, mollusks) Insects (honey bees) Samples from the animal body Tissues: -Meat, muscle (M.long.dorsi; M.bic.femoris) -Body fat -Blood -Some organs (liver, kidney, spleen, brain?, etc.) -Residue body (Meat and bone meal as feed) Tissues -Meat, muscle (breast, thigh) -Abdominal fat -Blood -Some organs (liver, kidney, spleen etc.) -Residue body (Animal body meal as feed) Edible fraction (e.g. fillet) -Residue body (e.g. fish meal as feed) - Food/feed produced by animals Milk Eggs Caviar Honey

28 4. Assessment of feed from the 1 st generation of GMP (input traits)

29 Recommendations for Nutrient Analysis (Examples for grain for non-ruminants and ruminants; by ILSI 2007) Crops/Grains/ Coproducts Livestock Type Analyte Grain: maize, wheat, barley Grain: maize, wheat, barley Non-ruminants DM, CP, EE, ADF, NDF, Ca, P,Mg, K, S, Na, Cl, Fe, Cu, Mn, Zn, ash, starch, lysine, methionine, cystine, threonine, trytophan, isoleucine, arginine, phenylalanine, histidine, leucine, tyrosine, valine, fatty acids, vitamins Ruminants DM, CP, EE, ADF, NDF, Ca, P,Mg, K, S, Na, Cl, Fe, Cu, Mn, Zn, Mo, ash, starch, ADIN, soluble protein, NPN, degrable protein, NDICP, ADICP, fatty acids, fat soluble vitamins

30 Mycotoxins in % Institute of Animal Nutrition Mycotoxins in isogenic (100 %) and Bt-corn (% of isogenic corn; data from some references) Deoxynivalenol Zearalenone Total fumonisins isogenic Bt-corn

31 Institute of Animal Nutrition Summary of published data to compare feeds from 1 st generation GM plants (with input traits) with their isogenic counterparts (2005) Animal (Species/categories) Ruminants Number of experiments Dairy cows 23 Beef cattle 14 Others 10 Pigs 21 Poultry Laying hens 3 Broilers 28 Others (Fish, rabbits etc.) 8 Nutritional assessment No unintended effects in composition and contamination (except lower mycotoxins concentrations in Bt plants) No biological relevant effects on digestibility and animal health as well as no unintended effects on performances of animals and composition of food of animal origin

32 Hatchability (% of inc. eggs) Laying intensity (%) Body weight (g per animal) Institute of Animal Nutrition (A) Body weight of female quails (age: 6 weeks), (B) laying intensity and (C) hatchability of quails fed with isogenic ( ) and transgenic (Bt, ) corn in a 10 generations experiment A Overall means (Range of generations) Isogenic ( ) Transgenic ( ) B Isogenic Transgenic ( ) 81.4 ( ) C Isogenic 77.4 ( ) Transgenic 76.7 ( ) Generations

33 Institute of Animal Nutrition Conclusion: Degradation of DNA - DNA is a permanent part of food/feed (daily intake: men: g; pig: g; cow: g) - tdna intake amounted to ~0.005% of total DNA-intake, if 50 % of diet come from GM-crops - DNA is mostly degraded during conservation (silage making) and industrial processing as well as in the digestive tract (ph, enzymes) - Small fragments of DNA may pass through the mucosa and may be detected in some body tissues (esp. leucocytes, liver, spleen) - There exist no data, that tdna is characterized by another behaviour as native plant-dna during feed treatment and in the animals

34 Institute of Animal Nutrition Conclusion: Degradation of newly expressed Proteins - In the ruminants feed protein are mostly degraded in the rumen and microbial protein and by-pass protein is degraded by enzymes in the smaller intestine, similar to nonruminants - The chemical and physiological properties (including microbial and enzymatic degradation) of novel proteins have been intensively tested - Intact novel proteins were not detected outside of the digestive tract in target animals - There is no advice, that newly expressed proteins are characterized by other chemical/physical properties as native protein

35 Institute of Animal Nutrition Recommendation by ILSI (2007) for performance studies Most studies should have the test grain transgen, its near isogenic counterpart (control), and, preferably, four or more conventional reference varieties to help explain any unexpected differences or confirm any expected differences observed between the test and the control

36 Effect of the maize event DAS (53 to 70% maize in the diet) on growth performance and organ weight of broilers in comparison to the near isogenic control and three non-transgenic conventional hybrids (120 broiler per treatment, 42 days, McNaughton et al., 2007) Criteria Control DAS Confidence interval (95%) Final weight (g/animal) Feed: gain (g/g) Relative weights of some organs (g/kg body weight) Kidney Liver Institute of Animal Nutrition Post-chill carcass (g/kg body weight) a 37 b a, b Different letter in one line show sign. differences (p < 0.05)

37 Institute of Animal Nutrition 5. GM - Plants of the 2 nd Generation (Substantial changes in composition; Plants with output traits)

38 Institute of Animal Nutrition GM - crops with output traits (GMP of the second generation) Increased content of desirable/valuable substances Nutrient precursors (e. g. β-carotene) Nutrients (amino acids, fatty acids, vitamins, minerals etc.) Substances which may improve nutrient digestibility (e. g. enzymes) Substances with surplus effects (e. g. prebiotics) Improvement of sensoric properties/ palatability (e. g. essential oils, aromas) Decreased content of undesirable substances Inhibiting substances (e. g. lignin, phytate) Toxic substances (e. g. alkaloids, glucosinolates, mycotoxins)

39 Proposal to assess the conversion of nutrient precursors of the 2 nd generation into nutrients (GM-plant with output traits; e.g. conversion of the precursor ß-carotene into vitamin A; by EFSA, 2008) Groups 4 Composition of diets Measurements; endpoints Balanced diet with typical amounts of the isogenic counterparts (unsupplemented control) 2 Balanced diet with adequate amounts of the transgenic counterpart (e.g. rich in ß-carotene) 3 Diet of Group 1 with ß-carotene supplementation adequate to Group 2 4 Diet of Group 1 with vitamin A supplementation adequate to expected ß-carotene conversion into vitamin A Depend on genetical modification of plants, e.g.: - Feed intake, animals growth - Concentration of specific/converted substance(s) in most suitable indicator organs (e.g. vit. A in the liver) 3 - Further metabolic parameters such as depots in further organs or tissues, activities of enzymes and hormones 1 Adequate feed amounts (pair feeding) for all animals; depletion phase for all animals before experimentation 2Depletion of specific nutrient in experimental animals could be necessary 3 Up to the steady state in the specific target organ 4 Some animal groups are fed with commercial/isogenic control feed to find out the biological range of the parameter(s)

40 Institute of Animal Nutrition Experimental design to determine the conversion of β-carotene into vitamin A in maize (60 % of diet, Mongolian gerbils, n = 10, Depletion period: 4 weeks, Feeding: 8 weeks, Howe and Tanumihardjo, 2006) β-carotene (nmol/g) Theoretical retinol intake (nmol/d) Unsupplemented control (Maize poor in carotene) Carotene rich maize Control + β-carotene Control + vitamin A Retinol in serum ( mol/l) 1.23 ± ± ± ± 0.16 Retinol in liver ( mol/g) 0.10 a ± b ± b ± c ± 0.15 a, b, c Means with different letters differ (p < 0.05)

41 Proposal to assess the effects of inhibitors of nutrient availability (e.g. phytate) Groups Diet composition Measurements 1 Balanced diet including typical levels of isogenic parenteral line fed ad lib. 2 Balanced diet including typical levels of transgenic line adequate to group 1 (e.g. low phytate crop) fed ad lib. 3 Diet of group 1 and supplemented with inhibited nutrient (e.g. phosphorus) fed in adequate amounts to group 2 4 Including of further groups with other commercial varieties (unsupplemented and supplemented with inhibited nutrient ) Depends on claim of genetic modification: - feed intake - animal performance, feed and nutrient (e.g. P) efficiency - digestibility of inhibited nutrient - concentration of inhibited nutrient in indicator organs/tissues

42 Institute of Animal Nutrition Conventional and low-phytate maize (78.5 % of the mixture) in the feed of fattening pigs (from Spencer et al., 2000) Control (0.3 g of available P per kg) Low-phytate maize (1.7 g of available P per kg) Inorganic P supplement P content (g/kg) kg live weight ) ) kg live weight ) ) Feed intake (kg/d) 2.23 a 2.50 b 2.53 b 2.51 b Live weight gain (g/d) 730 a 870 b 900 b 880 b Feed per gain (kg/kg) 3.05 a 2.87 b 2.81 b 2.85 b P excreted (g/kg weight gain) 4.6 a 8.9 c 3.8 b 8.8 c Strength (4th metacarpal bone, kg) 79.4 a bc b d Ash content (% in 4 th metacarpal bone) 53.5 a 60.1 bc 59.3 b 61.2 c a, b, c, d Different letters in one line indicate significant differences (p < 0.05) 1) +2.0 g P/kg 2 ) +1.5 g P/kg

43 Proposal to assess the effects of enhanced nutrient utilization (e.g. by enzymes) Groups Diet composition Measurements 1 Balanced diets including typical levels of near isogenic parenteral line fed ad lib. 2 Balanced diets including typical levels of transgenic line (e.g. content of specific enzyme); feeding level of group 1 3 Diets of group 1 plus enhancer adequate to transgenic line (or dose-response studies), feeding level of group 1 4 Diets of groups 2 and 3, fed ad lib. Depends on the claim of genetic modification: - feed intake - animal Performance(s) - digestibility of specific nutrient(s) - metabolic parameters; quality of food of animal origin

44 18:0 9 18: : :3 Stearic acid Oleic acid Linoleic acid -linolenic acid LA ALA :3 -linolenic acid GLA 15 18:4 Stearidonic acid SDA Fatty acid biosynthesis in plants and the new introduced changes to produce Stearidonic acid (C18:4 n-3) and the effects of various desaturases ( 6 and 15; by Ursin 2003 and Whelan 2009)

45 Concentrations (mg/l) of some fatty acids in whole milk on the initial and the final day (10 th day) of supplementation with ruminally protected SDA-oil (SDA intake about 30 g per day; n=5; by Kitessa and Young 2011) Fatty acid Initial value (Day 1) Final values (Day 10) C18:1 n-9; Oleic acid C18:2 n-6, Linoleic acid C18:3 n-3; Linolenic acid C18:4n-3, Stearidonic acid C20:5 n-5, EPA C22:5 n-3, DPA Total n Total n

46 Calculation on the influence of added oils on the intake and the pool size of n-3 fatty acids in edible tissues (breast and leg meat with skin) in broilers (by Rymer et al., 2011) Fatty acids Control Supplemented fish oil Intake of n-3 fatty acids (mg/broiler) C18:3 C18:4 C20:5 C22:5 C22: Supplemented SDA oil Pool size (mg/broiler) of fatty acids in edible tissues C18:3 C18:4 C20:5 C22:5 C22:

47 6. Open Questions and Conclusions

48 Assessment of present modifications of plants from the view of food safety and food security Objectives More resistant against herbicides Mores resistant against insects etc. Present significance Contributions to Food safety Global food security ( ) More valuable ingredients ~ Less undesirable ingredients ( ) More efficient use of resources (water etc.) ( ) extremely high very high high ~ not important

49 Animal nutrition (safety and nutritional assessment of feed) between plant and animal breeding Plant breeding (incl. Genetical modifications) Animal nutrition (Safety and nutritional assessment of feeds) Animal breeding (incl. Clones and GM-animals)

50 Open questions Do we need more feeding trials with feeds from GMP of the 2nd generation? Do we need more feeding trials with stacked events (more tdna, more transgenic protein)? Do we need more feeding trials with GMP, which use resources more efficient (e.g. water, abiotic stress)? Do we need adequate studies with plants (feed) from traditional breeding? Which types of studies do we need with GM-animals and for safety assessment of food from GM-animals?

51 Conclusions Up to now about 1.3 Billion ha of GM-crops have been cultivated all over the world Most animal feeding studies were done with GM-crops of the 1stgeneration (with input traits) No unintended effects in composition (except lower mycotoxins), safety and nutritional properties were registered in more than 100 studies with food producing animals tdna and newly expressed proteins in GM-crops show similar properties during processing and in animals as plant-dna and proteins Some other experimental designs are recommended esp. for nutritional assessment of feeds from GM-crops of the 2nd generation (with output traits, e.g. bioavailability of nutrients) and of GM-animals Furthermore case by case studies seem to be necessary to answer open questions, more groups with isogenic counterparts should be included Feeding studies with food producing animals should be also used for safety assessment

52 Will be published in Oct./Nov 2013