Technical Opinion no. 2040/2009

Proceedings:  01200.000925/2009-11
Applicant:  Syngenta Seeds Ltda.
CNPJ:   49.156.326/0001-00
Address: Avenida Nações Unidas, 18.001, 4º Andar, São Paulo, SP.
Matter: Commercial release of Genetically Modified Cotton.
Previous summary: 1810/2009. Published on 04.29.2009
Meeting: 126th CTNBio Regular Meeting, held on 09.17.2009.
Decision:  GRANTED.

CTNBio, following approval of an application for commercial release of genetically modified insect resistant and herbicide tolerant corn (Bt11 x GA21), as well as all progenies originated from the Bt11 x BA21 event and its derivatives  obtained from crossing non-transgenic corn lineages and populations with lineages carriers of Event Bt11 x GA21 decided favorably to the  GRANTING, under the terms of this Technical Opinion.
Syngenta Seeds Ltda. requested CTNBio a Technical  Opinion related to biosafety of the genetically modified insect resistant and herbicide tolerant corn (Zea mays), namely Bt11 x GA21 corn, Event BT11 x GA21 for purpose of free registration, use in the environment, human and animal consumption, marketing and industrial use and any other use and activity related  to  this  GMO including derivative lineages and  cultivars as well  as byproducts, in compliance with  other regulations and requirements applicable to any use  of cultivated species of the genus Zea effective in Brazil. Bt11 x GA21 corn  was generated by classic  genetic development, through sexual crossing between genetically modified lineages containing either event BT11 or GA2, previously approved by the National Biosafety Technical Commission – CTNBio to be commercially released. Bt11 x GA21 corn was already approved in the United States, Canada, Japan, Mexico, Korea and the Philippines for human and animal consumption. In the process of developing Bt11 x GA21 corn there were no genetic modifications besides the introgression of both genes in corn lineages. Compared to conventional corn, Bt11 x GA21 corn has no higher ability to survive as a pest. The presence of genes granting resistance to Lepidoptera insects and tolerance to the glyphosate insecticide give Bt11 x GA21  corn selective advantage when exposed to the herbicide and in the presence of target insects. However, these features are not sufficient to turn the product into a pest in corn cultures. The use of corn containing the stacked event, Bt11 x GA21, represents a future trend   meeting corn producers’ demand – by combining two important agronomic features into a same hybrid plant. Event Bt11 contains gene cry1A(b) – coming from  Bacillus thuringiensis, granting resistance to certain insects, and gene pat, derived from Streptomyces  viridochromogenes, a soil bacterium used as a selection marker during the transformation process. Gene cry1A(b) is responsible for producing protein Btk that is proteolytically cleavated in the alkaline intestine of Lepidoptera insects in an active insecticide form. This insecticide protein, when active, interacts with a receptor molecule that is present solely in the epithelial cells of the middle intestine of susceptible insects, generating pores in cell membranes. When the pores are formed, the cell osmotic balance becomes disrupted, cells swell and undergo lysis. The larvae of  susceptible insects, when submitted to the Cry protein stop feeding and later die. Several bonding sites featuring high affinity for Bt proteins were already identified in the middle intestine of susceptible insects. Studies showed that the insecticide protein codified by gene cry1A(b) is highly specific for Lepidoptera insects. Event GA21 contains the mutant gene mepsps that is responsible for the expression of protein 5 enolpyruvyl-shikimate-3-phosphate-syntase (mEPSPS). The EPSPS protein is a key enzyme in processing the shikimic acid, which is involved in the biosynthesis of aromatic amino acids and is commonly found in plants, fungi and bacteria. The enzyme does not exist in animals. The EPSPS enzyme is highly sensitive to herbicide products containing the glyphosate active principle. Results from hybridation data show that Bt11 x GA21 corn maintains its hybridation standard in the same way as the respective parental events for each corresponding transgene. The set of evidence obtained based on results of comparative molecular analysis, analysis of genetic heritage standards and comparative analysis of level of expression for proteins Cry1A(b) and mEPSPS in Bt11 x GA21 corn suggest that the levels of exposure to non-target organisms and in human and animal feeding are the same as to either Bt11 or GA21 measured individually. The presence of genes pat and mepsps make the Bt11 x GA21 corn resistant to a pair of chemical products with herbicide properties available in the market: glyphosate and gluphosinate ammonium. Should corn have potential to turn into a pest plant, its control where such products were used could increase its invasive ability. However, the high degree of domestication of corn made the species highly dependent on man. This may be ratified by the absence of pest or even feral populations in agricultural and natural environments, even after millennia of cultivation in Brazil. Therefore, the risk that corn changes into a pest plant, if such risk exists, is negligible. According to Annex I to Ruling Resolution nº 5, of March 12, 2009, the applicant shall have a term of thirty (30) days from the publication of this Technical Opinion to adjust its proposal to the post-commercial release monitoring  plan.  Under Article 14 of Law  no. 11,105/2005, CTNBio found that the request complies with  the applicable rules and legislation securing the biosafety of environment, agriculture, human and animal health and reached a conclusion that the stacked corn Bt11 x GA21 is substantially equivalent to conventional corn and its consumption is safe for human and animal health. Regarding the environment, CTNBio’s conclusion is that cultivation of Bt11 x GA21 corn is not a potential cause of significant environmental degradation, keeping with the biota a relation identical to that of conventional corn.
 
 TECHNICAL OPINION
I. GMO Identification
GMO name:    Bt11 x GA21 corn.
Applicant:   Syngenta Seeds  Ltda.
Species:   Zea Mays L.
Inserted characteristics:  Tolerance to glyphosate herbicide and insect resistance
Method of insertion:  Bt11 x GA21 corn, ranked as Risk Class I, was developed by classical genetic improvement, through sexual crossing between genetically modified lineages containing event Bt11 and event GA21.
Prospective use:  Free registration, use, essays, tests, sowing, transport, storage, marketing, consumption, import, release and discarding.   
II. General  Information
Corn, Zea Mays L. is a species of the family Gramineae, tribe Maydae, sub-family Panicoidae that is separated within the sub-genus Zea and has chromosome number 2n = 20,21,22,24(1). The wild species closest to corn is the teosinte found in Mexico and some regions of Central America, where it is able to cross with cultivated corn in the production fields.
Corn has a history of over eight thousand years in the Americas, and is cultivated since the pre-Columbian era. Among higher plants, corn is the best scientifically characterized and is currently the cultivated species that reached the highest degree of domestication and is unable to survive in nature but when cultivated by man(2). There are currently over 300 identified races of corn and, within each such race, thousands of cultivars.
One of the most important sources of food in the world, corn is an input in the production of a wide range of foodstuff, rations and industrial products. Brazil is one of the largest producers of corn over the world, and corn is cultivated nearly all over the national territory(3).
Occurrence of insects in Earth is larger in the tropics than in temperate regions, where the damages caused by such animals are more noticeable. Among the most damaging corn pests an important place is taken by the fall armyworm, Spodoptera frugiperda. Cruz et al. (4) estimated that the losses in Brazil caused by infestations of S. frugiperda reach 400 million Dollars each year. Other species of the order Lepidoptera are also important pests in the culture of corn, such as the corn earworm (Helicoverpa zea) and stalk borer (Ditraea sacharalis).
The main measure to control insects in the corn cultivation has been the use of insecticides. In some areas of the Brazilian Center-West, for instance, tenths of insecticide sprays are needed in a single culture cycle. Another measure to control pests would be the use of resistant cultivars.
Bt11 x GA21 corn was developed through classic genetic improvement, by sexual crossing between genetically modified lineages containing separately event Bt11 and event GA21(5,6). Bt11 x GA21 corn was already approved in the United States, Canada, Japan, Korea and the Philippines for animal and human consumption. In the course of developing Bt11 x GA21 corn, there were no other genetic changes in addition to the introgression of both events contained in corn lineages Bt11 and GA21 separately(7).
Compared with conventional corn, Bt11 x GA21 corn fails to display greater ability to survive as a pest. The presence of gens granting resistance to Lepidoptera insects and tolerance to glyphosate herbicide give a selective advantage to Bt11 x GA21 corn when it is exposed to the herbicide and submitted to the presence of target-insects. However, such characteristics are not sufficient to make this corn a pest in corn cultivars(8).
The use of corn featuring the stacked event, Bt11 x GA21, represents a future trend – that meets the demand of farmers – by combining two agronomically important features in a single hybrid. Corns with events combined by classic genetic improvement are approved in Japan, Korea, Philippines and are under analysis in a number of other countries(7).
III. Description of  GMO and Proteins Expressed
The corn containing event Bt11 was obtained by direct transfer of DNA  in protoplasts of corn lineage H8540 by inserting plasmid pZO1502 containing genes cry1Ab and pat. The expression product of gene cry1Ab is protein Cry1Ab that has insecticide activity on target-pests, protecting the plants  from damages caused by such pests. Gene cry1Ab was isolated from bacterium B. thuringiensis subspecies kurstaki strain HD-1(3,8,9). Gene pat is derived from Streptomyces  viridochromogenes  strain Tu494 and is responsible for codifying enzyme phosphinothricin N-acetyltransferase (PAT). This enzyme is able to chemically inactivate herbicides derived from phosphinotricin, such as glyphosinate ammonium, making resistant cells and plants containing the enzyme. The PAT enzyme activity has its activity described and well known(9).
Event GA2 contains gene mepsps that expresses enzyme 5-enolpyruvyl-shikimate-3-phosphate-synthase (mEPSPS). Protein mEPSPS differs from feral  EPSPS in two amino acids and is a key enzyme in the shikimic acid process, involved in the biosynthesis of aromatic amino acids. mEPSPS is  highly sensitive to herbicide products containing glyphosate. Corn plants transformed with mEPSPS gene, such as those derived from event GA2, synthesize protein mEPSPS  that grants tolerance to herbicide products containing glyphosate(10,11).
Proteins mEPSPS and Cry1Ab mode of action and biologic activities expressed in Bt11 x GA21 corn are different and do not posses known interaction mechanisms  that are able to cause adverse effects to human and animal health, and to the environment. mEPSPS and  Cry1Ab proteins present in Bt11 x GA21 corn are accumulated in different cell compartments and display distinct and non-interactive metabolic functions. Therefore, protein mEPSPS is directed to the chloroplast while protein Cry1Ab is accumulated in the cytoplasm(5,6).
Expression level of proteins mEPSPS and Cry1Ab is low in individual events (GA21 corn and Br11 corn) and therefore the likelihood that such proteins would interact between them is held improbable, a fact that is macroscopically confirmed through the analysis of agronomic and phenotypical characteristics related to efficacy and selectivity of Bt11 x GA21 corn in the  fields(5,6).
IV. Aspects Related to Human and Animal Health
Safety aspects of proteins Cry1Ab and EPSPS were thoroughly assessed by CTNBio and protein Cry1Ab mode of action is well clarified by scientific literature(5,6). In vitro tests were used to assay increased digestibility of foodstuffs containing pre-heated proteins Cry1Ab and mEPSPS. The study showed that pre-heating increases the protein digestibility in simulated gastric and intestine fluids, suggesting that the likelihood of an eventual allergenic potential of protein Cry1Ab is extremely low, for the easy of its digestion(5,6). Further, in vivo and in vitro studies confirmed that proteins Cry1Ab expressed in B. thuringiensis and Bt11 x GA21 corn are highly selective and do not act on mammals(13,14,15,16,17,18,19).
Protein mEPSPS is an enzyme that is present in all plants and in a large number of microorganisms(17), while protein Cry1Ab does not display enzymatic activity in plants and therefore fails to affect plant metabolism.
The likelihood that biochemical interaction takes place between proteins mEPSPS and Cry1Ab in the complex matrix of a plant is limited, since such proteins accumulate in different places of the cells and in a low level of expression. With this, a potential exposure to such proteins is extremely low in human and animal feeding.
Considering that proteins mEPSPS and Cry1Ab fail to produce toxicity in the maximum doses tested, it is highly unlikely that an interaction able to cause additive or synergic effects occurs between such proteins in the normal doses found in foodstuffs. The literature in the area of toxicology of chemical mixtures provides information showing that such interactions are inexistent when the substances are administered in doses substantially below the levels of unobserved adverse effect(20,2,22,23).
Due to the rigorous specificity for substrates, enzymes EPSPS link just S3P, PEP and glyphosate. The only known metabolic product is the 5-enolpyruvyl shikimic-3-phosphate, which corresponds to the penultimate product of the shikimic acid pathway. Shikimic acid is a precursor for biosynthesis of amino acids (phenylalanine, tyrosine and triptophane) and a number of secondary metabolits, such as tetrahydrofolate, ubiquinone and K vitamin(24). Though the shikimic acid (or shikimate) pathway and proteins EPSPS do not occur in mammals, fish, birds and insects, they are important to plants. It is reckoned that aromatic molecules, all of them derived from shikimic acid, represent no less than 35% of a plant’s dry weight(25,26).
In vitro assays performed with simulated digestive fluids are widely used tools as a model for animal digestion. This simulated system was used to probe into digestibility of plant proteins(27,28), animal proteins(29), and food additives(30), as well as to assay the protein quality(31) and the allergenicity potential through absorption of the proteins by the digestive system(32).
The knowledge on the mode of action, specificity and safe use history of protein EPSPS, potential toxic and allergenic effects of such proteins to humans and other mammals were assayed through in vitro digestion tests. The studies used simulated gastric fluids (pH 1.2) and intestinal fluids (pH 7.5). The degradation rate of protein mEPSPS (mature protein with no transit peptide) was assessed through Western blot analyses. The study showed that protein mEPSPS and peptides degraded in less than 15 seconds after exposed to the gastric fluid. In the simulated intestinal fluid, degradation of protein mEPSPS occurred in a period shorter than 10 minutes(33).
Finally, enzyme EPSPS expressed in corn containing event GA21 has no typical characteristics of known allergens, since the behavior of allergenic proteins in the digestive tract is well described(34,35,36). There are no homology regions when the introduced sequence is compared with known allergen sequences. Besides, several alimentary allergens are known to be stable under heat.
V. Environmental Aspects
Corn is a monoic, allogamic and annual plant, pollinated mostly by the wind. Distances that may be covered by the pollen depend on wind patterns, humidity and temperature. Corn pollen is freely dispersed close to the cultivated area, and may reach styli-stigmas of the same or different genotypes and, under adequate conditions, starts its germination, generating the pollinic tube and promoting the ovule fecundation within an average term of 24 hours.
Studies on pollen dispersion have been conducted and some of them show that pollen may travel long distances, though the majority is deposited close to the cultivated area with a very low translocation rate. Over 95% of the pollen may reach distances within 60 m of the pollen source(37). Luna et al. (38) investigated pollen isolation distance and control, where it was shown that crossed pollination occurs within 200 m. However, no crossed pollination, under conditions of non-detasseling, was noticed in distances higher than 300 m from the pollen source. The results indicate that pollen viability is maintained for two hours and that crossed pollination was not observed in distances of 300 m from the pollen source.
When compared with concentrations at one meter from the source culture under low-to-moderate winds it was estimated that about 2% of the pollen reaches 60 meters, 1.1% reaches 200 meters, and 0.75% to 0.5% reaches a distance of 500 meters. At a distance of ten meters from the field, the number of pollen grains per unit of area is tenfold lower than the number observed at one meter from the border. Therefore, if established separation distances developed for the production of corn seeds are observed, it may be expected that pollen transfer to surrounding varieties is minimized and that the presence of glyphosate-tolerant genetic materials is unlikely.
There are no kindred species of corn naturally distributed within Brazil. However, though the gene flow to local varieties of open pollination is possible, it poses the same risk than the one caused by commercial genotypes available in the marketplace. In the specific case of crossing between GA21 corn and creole varieties, selected pressure from the management by small farmers is not expected; the transgene will not be incorporated to the genome of creole varieties because in practice a small farmer does not use herbicides.
From the agronomic viewpoint, coexistence between cultivars of conventional corn (improved or creole) and transgenic corn is possible(39,40). Old communities and modern farmers have learned how to live on without problems with different corn cultivars, while keeping their genetic identities along time.
The likelihood of a transgenic plant becoming an invading species, as well as the likelihood that the crossing of GA21 corn with other corn plants generating an invading plant is negligible, in view of the biologic characteristics of the species and the fact that corn cannot survive without human intervention, a result of the selection made during the plant evolution.  Corn is held as the species reaching the highest degree of domestication among cultivated plants, and has lost its ability to survive in nature, such as elimination of thrashing. Therefore, corn is a plant that is unable to survive under natural conditions without technical assistance. Therefore, one expects GA21 corn to exhibit an environmental behavior similar to ordinary corn, being hence negligible the possibility of changing into an invading or pest plant.
Introduction of gene elements did not change the plant reproductive features, and the odds of crossed fecundation between hybrids and conventional corn lineages will persist between event Bt11 x GA21 and other corn plants.
Gene flow in corn may take place through transfer of pollen and dispersion of seeds – which is easily controlled – since corn domestication eliminated the ancient mechanisms of seed dispersion and the movement of pollen is the only effective form for genes to escape from corn plants.
The likelihood that a gene mepsps of a transgenic plant  passes over to other organisms, such as, for instance, soil microorganisms, is practically zero(41,42). Naturally, gene  epsps  is common in plants, fungi and microorganisms, occurring abundantly in nature and does not results in significant risk to the soil microbiota. Besides, there is no evidence that plant genes have in some way been transferred to bacteria under natural conditions.
Agronomic parameters and efficacy of controlling pest Lepidoptera in Bt11 hybrid corn were compared with isogenic lineages in assays conducted in five locations: City of Uberlândia, State of Minas Gerais; City of Ituiutaba, State of Minas Gerais; City of Iraí de Minas, State of Minas Gerais; City of Campo Mourão, State of Paraná; and City of Pinhalzinho, State of Santa Catarina. Plant height, ear insertion height, date of  male and female flowering, percentage of erect plants, type and color of seed-corn, humidity content, yield and ear damage were the parameters observed in agronomic assays. In order to assay the efficacy of event Bt11 in controlling pest Lepidoptera, damages of fall armyworm (S. frugiperda); corn borer (D. saccharalis); and corn earworm (H. zea) (5,6) were examined.
Hybrids containing event Bt11 were efficient to control the assayed pest Lepidoptera and exceeded the agronomic parameters related to corn-seed and grain damage. According to available information, the favorable performance difference was mainly related to the efficient protection against attacks from pests studied. As for other agronomic parameters assayed, Bt11 hybrids exhibited a performance  statistically similar to the respective isogenic non-genetically modified hybrids. The results confirm the equivalence of agronomic performance between Bt11 hybrids and isogenous corn non-genetically modified under Brazilian cultivation conditions(5,6).
Frequent use of chemical insecticides contributes towards environment degradation, environmental pollution and disruption of the whole ecosystem in the corn culture and even of other cultures in rotation. With the adoption of insect resistant genetically modified plants, reduction of insecticides was considerable in countries where the technology has been used for over ten years. In 2001 alone, United States farmers, for instance, obtained a reduction of over 8,000 tons of active insecticide ingredient in (43,44,45).
In China, the use of insecticides were reduced by 67% on average, and the reduction in terms of active insecticide ingredient reached 80%(46). In South Africa, the reduction was about 66%(42). These examples suggest that the use of Bt technology in Brazil may contribute for a diminished use of insecticides and, therefore, reduction of the impacts resulting from the use of such pesticides to the environment and human and animal health. Besides, the use of Bt technologies may have positive effects in preserving populations of non-target organisms and beneficial insects, making an integrated management of crop pests easier(47,48,49). Besides, the use of technologies that reduce spraying chemicals on crops may bring secondary benefits with the reduced use of raw-material in the production of pesticides, reduction of fuels used to produce, distribute and apply pesticides and elimination of the need to use and discard pesticide packaging(50).
Compared with conventional corn, Bt11 x GA21 corn has no higher ability to survive as a pest. The presence of genes for Lepidoptera insect resistance and glyphosate herbicide tolerance imparts a selective advantage to Bt11 x GA21 corn when exposed to the herbicide and in the presence of target insects. However, such features are not enough for Bt11 x GA21 corn change into a pest to corn cultures(8).
Comparison of amino acid sequences of the new proteins expressed by Bt11 x GA21 corn against sequences of toxic or allergenic proteins show that there is no homology likely to indicate similarities. Therefore, toxic or allergenic effects are not expected as a result of contact or consumption of foodstuffs containing event Bt11 x GA21(8).
VI. Restrictions to the Use of the GMO and its Derivatives
As established by Article 11  of Law nº 11,460, of March 21, 2007 “research and cultivation of genetically modified organisms may not be conducted in indigenous lands and areas of  conservation units.”
The studies submitted by applicant showed that there was no significant difference between corn hybrids derived from unmodified lineages and Bt11 x GA21 corn regarding agronomic characteristics; reproduction and dissemination modes; and survival ability. All evidences submitted with  the application documents and bibliographic references confirm the risk level of the transgenic variety as being equivalent to the risk level of non transgenic varieties regarding soil microbiota, other plants and human and animal health. Therefore, cultivation and consumption of Bt11 x GA21 corn are not potential causes of significant degradation of the environment, nor of risks to human and animal health. For these reasons, there are no restrictions to the use of such corn and its derivatives, except in locations mentioned by Law nº 11,460, of March 21, 2007.
Vertical gene flow to local varieties (the so-called creole corns) of open pollination is possible and poses the same risk as the one caused by commercial genotypes available in the market (80% of conventional corn planted in Brazil comes from commercial seeds that underwent a genetic improvement process). Coexistence of conventional corn (either improved or creole) cultivars and transgenic corn cultivars is possible from the agronomic viewpoint(39,40) and shall comply with the provisions of CTNBio Ruling Resolution nº 4.
After being used for ten years in other countries, no problems were detected to human and animal health and the environment that may be ascribed to transgenic corns. It shall be emphasized that the lack of negative effects resulting from farming transgenic corn plants is far from a guarantee that such problems may not occur in the future. Zero risk and absolute safety do not exist in the biologic world, although there is a significant amount of reliable scientific information and a safe history of ten years underlying the fact that Bt11 x GA21 corn is as safe as the traditional corn versions. Therefore, applicant shall conduct the post-commercial release monitoring according to CTNBio Ruling Resolution nº 3.
VII. Consideration on the Particulars of Different Regions of the Country (Information to supervisory agencies)
As established by Article 11  of Law nº 11,460, of March 21, 2007 “research and cultivation of genetically modified organisms may not be conducted in indigenous lands and areas of  conservation units.”
VIII. Conclusion
Whereas the corn (Zea mays) variety Bt11 x GA21 belongs to a well characterized species with a solid history of safety for human consumption and that genes cry1A(b), pat, mepsps introduced in this variety codify proteins that are ubiquitous in nature and are present in plants, fungi and microorganisms that are part of human and animal alimentary diet;
Whereas insertion of this genotype took place through classic genetic improvement and resulted in insertion of a stable and functional copy of cry1A(b), pat, mepsps genes that granted to the plants tolerance to glyphosate herbicide and resistance to insects;
Whereas data on centesimal composition failed to show significant differences between genetically modified and conventional varieties, suggesting a nutritional equivalence between them;
Whereas CTNBio conducted a separate assay on the events and issued an opinion favorable to commercial release of the separate events;
Whereas:
1. Bt11 x GA21 corn is a genetically modified product, displaying resistance to a number of Lepidoptera pests and tolerance to glyphosate herbicide, developed through classic improvement by sexual crossing between lineages containing event Bt11 and event GA21, previously approved for commercial release;
2. Comparative molecular analysis of Bt11 x GA21 corn evidenced that integrity of inserts was maintained during the classic improvement with the purpose of combining both events;
3. Segregation analysis and genetic heritance standards of Bt11 x GA21 corn  showed that genes of events Bt11 and GA21 are independent and segregate on a stable manner along successive generations;
4. Agronomic and efficacy assays of Bt11 x GA21 corn indicate that combination of such events by classic genetic improvement methods (sexual crossings) did not lead to expression of any other characteristics, except those already expected, that is to say, resistance to certain insects and tolerance to glyphosate herbicide;
5. Expressions of proteins Cry1Ab and mEPSPS  in Bt11 x GA21 corn are not significantly different from their expression in corns containing the separate events;
Therefore, considering internationally accepted criteria in the process of analyzing risks in genetically modified raw-material it is possible to conclude that Bt11 x GA21 corn is as safe as its conventional equivalents. In the context of the competences granted to it under Article 14 of Law nº 11,105/05, CTNBio considered that the request complied with the rules and legislation in effect that intend to guaranty environmental and agricultural biosafety and human and animal health, reaching a conclusion that Bt11 x GA21 corn is substantially equivalent to conventional corn, being its consumption safe for human and animal health. Regarding the environment, CTNBio’s conclusion was that the Bt11 x GA21 corn is not a potential cause of significant degradation to the environment, keeping with the biota a relation identical to that of conventional corn.
CTNBio advocates that this activity is not a potential cause of significant degradation to the environment or of harm to human and animal health. Restrictions to the use of the GMO analyzed and its derivatives are conditioned to the provisions of Law nº 11,460, of March 21, 2007, and to CTNBio Ruling Resolution nº 03 and Ruling Resolution nº 04.
CTNBio assay took into consideration opinions issued by the Commission members; ad hoc consultants; documents delivered to CTNBio Executive Secretary by applicant; results of planned releases to the environment; and discussions, lectures and papers related to the public hearing held on 03.20.2007. Besides, independent studies and scientific literature of applicant, conducted by third parties were also considered.
According to Annex I to Ruling Resolution nº 5, of March 12, 2009, the applicant shall have a term of thirty (30) days from publication of this Technical Opinion to adjust its proposal to the post-commercial release monitoring  plan.
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Walter Colli
President of CTNBio

Dissenting Votes:
CTNBio members Doctor Pedro Binsfeld, Doctor Rodrigo Roubach, Doctor Graziele Almeida  da Silva, and Doctor Leonardo Melgarejo (Permanent Sector Subcommission: Human and Animal Health, Environment, respectively) voted against the  commercial release of Bt11 x GA21 corn.
Doctor Paulo Yoshio Kageyama (Permanent Sector Subcommission for the Environment) issued an opinion against the approval of the product on the following bases:
1. A full risk assessment would be necessary, both for human and animal health and the environment;
2. Events that were already commercially released were always partially based on unconfirmed hypothesis and aggregating two or more sources of uncertainty generally results in uncertainties that exceed the sum or product of such source uncertainties;
3. The Annex III to the Cartagena Protocol to Biosafety, by which a decision on commercial release should depend on risk assessment, was discarded.
4. Risk assessment of Genetically Modified Plants stacked events should be conducted under CTNBio Ruling Resolution nº 5 and other complementary rulings, mentioned in Doctor Kageyama’s opinion.
The author of the Request for Examination, under Article 22 of MCT Directive nº 146, Paragraph 22, Doctor Leonardo Melgarejo (Environment Permanent Sector Subcommission) issued an opinion against approval of Bt11 x GA21 corn considering that:
1. The technical foundations submitted by applicant are out of date;
2. Applicant failed to take into account the possible pleiotropic effects and interactions between the genomes and the environment;
3. The stacked events deserve full risk assessment and attention to aspects of human and animal health and the environment;
4. Economic and social aspects should not be discarded;
5. Events up to now released for commercial fail to comply in full with the provisions of CTNBio Ruling Resolution nº 5;
6. It is the author’s understanding that there are not studies or research supporting the similarity of risks in case of singular events and their stacked composition, according to the Precaution Principle;
7. The applicant should supply the event molecular characterization to confirm preservation of the insert features and map the homology between the genetically modified parents and the stacked event;
8. Applicant should conduct studies to submit an analytic comparison of the stacked event taking into account a standard group of compositional and agronomic parameters;
9. Applicant should conduct studies analyzing potential interactions of the characteristics incorporated to the stacked event;
10. The author does not agree that the methods used in molecular characterization are sufficiently accurate.
11. The author believes that the potential interactions between transgenic characteristics could not be assayed with the available tools.