| Summary of the safety assessment (food safety): |
Bt11xMIR162xGA21 maize Biosafety, resulting from breeding, by means of the classical genetic improvement, from parental plants linked to genetically modified maize, to their release into the environment, marketing, consumption and any other activities related to such maize and progeny derived from it. Every single event, has already obtained a favourable opinion for marketing in Brazil. The Bt11 event has the gene cry1Ab Bacillus thuringiensis, which confers resistance to certain lepidopteran insects, and the pat gene, derived from the soil micro-organism Streptomyces viridochromogenes, used as selection marker during the transformation process. The Cry1Ab protein and proteolytically cleaved in the alkaline gut of lepidopteran insects, becoming insecticidally active. The MIR162 maize was obtained from vip3Aal9 gene, which confers resistance to lepidopteran insects, and from manA gene, which encodes the enzyme phosphomannose isomerase (PMI), used as a selection marker. A modification caused by the transformation process resulted in a difference in two codons of vip3Aa19 inserted gene, being then called vip3Aa20 for the MIR162 maize. This difference resulted in the modification of only one amino acid, located beyond the proteolytic cleavage site of the protein then called Vip3Aa2O, expressed in MIR162 maize, thus maintaining its insecticidal properties against several lepidopterous pest of maize. The manA gene was obtained from Escherichia coli strain K-12 and PMI protein expression was used as a selection marker during the MIR162 maize transformation process. The GA21 Event contains the gene mepsps that expresses the enzyme 5-Synthase enolpyruvyl-shikimate-3-phosphate (mEPSPS). In the process of shikimic acid, the EPSPS is a key enzyme, involved in the biosynthesis of aromatic amino acids (phenylalanine, tyrosine and tryptophan), found naturally in plants, in fungi and bacteria, and absent in animals. The EPSPS is highly sensitive to herbicides containing glyphosate. Maize plants transformed with the mutant gene epsps (mepsps), such as those from GA21 event, synthesize the mEPSPS protein that confers tolerance to herbicides containing glyphosate. The comparative molecular analysis confirmed the genetic integrity of the inserts Bt11, MIR162 and GA21 during the process of classical genetic improvement to obtain the Bt11xMIR162xGA21 maize. The hybridised fragments showed the expected size and demonstrated the integrity of the inserts. Furthermore, an analysis of genetic segregation was conducted. The results of phenotypic segregation obtained by ELISA and genotypic segregation obtained by qPCR were tested for adhesion and indicated that the loci of individual events and of combination were segregated stably and independently. The efficacy and agronomic evaluations indicated that there is differential expression in any other characteristic beyond those expected, i.e. insect resistance and herbicide tolerance. A comparative study was conducted on the concentration of the proteins Cry1Ab, VIP3Aa20 and mEPSPS, and the separated events was performed in Bt11xMIR162xGA21e hybrid maize. The proteins from plant tissues were quantified by ELISA immunoenzymatic tests in various stages of development. For most assessed tissues at different stages of development, there was not a statistically significant difference between the expression of each protein expressed in maize Bt11xMIR162xGA21 and maize containing events alone. Although significant differences have been identified in the expression of Vip3Aa20 in root and pollen at the stage of anthesis, there is no evidence for the results indicating a trend for change in the expression levels of proteins expressed in function of the combination of events. A study of the Bt11xMIR162xGA21 maize influence on the insect community was conducted to verify the possibility of interference concerning (a) primary pests of maize crop in soil and in aerial parts and (b) secondary pests. The results indicated that the combined maize had no adverse effect for the insect community or for the predator Doru luteipes, when compared to its isogenic, unmodified hybrid. Moreover, CTNBio consulted independent scientific literature to assess the occurrence of any unexpected effect from the breeding among these events.
TECHNICAL REPORT
1. Identification of GMOs
Name of GMO: maize
Applicant: Syngenta Seeds Ltda.
Species: Zea mays
Inserted feature: insect resistance and herbicide tolerance
Method to introduce the feature: The Bt11xMIR162xGA21 maize, classified as Risk Class I, was developed by classical genetic improvement, by sexual reproduction among genetically modified maize lines, containing Bt11, GA21, and MIR162 events.
Proposed use: Free registration, use, tests, planting, transport, storage, marketing, consumption, release, disposal, and any other activities related to the GMO.
II. General information
The maize Zea mays L. is a species from Gramineae family, tribe Maydeae, subfamily Panicoidae that is separate from subgenus Zea and presents chromosome number 2n = 20,21,22,24 (FAO/WHO 2000). The closest wild species of maize is the teosinte, which is found in Mexico and in parts of Central America, where it can be bred with cultivated maize in the production field.
The Bt11xMIR162xGA21 maize is a genetically modified product that provides resistance to several lepidopteran pests and for tolerance to glyphosate herbicide, developed by means of traditional breeding by sexual reproduction among maize lines containing the Bt11 event, the MIR162 event, and the GA21 event. This maize contains no other event in genetic transformation, besides the events Bt11, MIR162, and GA21. The Bt11 maize, MIR162 maize, and GA21 maize have been approved for commercial release by the National Technical Commission on Biosafety (CTNBio), based on Proceedings 01200.002109/2000-04, 01200.007493/2007-08, and 01200.000062/2006-21 as theTechnical Reports 1255/2008, 2042/2009, and 1597/2008, respectively.
The 'combined' or 'pyramided' products, as in the case of Bt11xMIR162xGA21 maize, represent a future trend to cultivate genetically modified plants to meet the demand of producers to combine different characteristics important for agronomy in a single hybrid, allowing to control pests better (Aragão and Andrade, 2010). Besides the tolerance to the herbicide glyphosate by the expression of the GA21 event, the combination of different modes of action of Cry1Ab and VIP3A protein from the events (Bt11, MIR162) allowed to preserve insect technology resistance for a longer period, thereby ensuring its employment to control pests in an integrated manner for maize harvest in Brazil.
Maize hybrids containing the events Bt11, MIR162, and GA21 alone and/or combined by classical genetic improvement so approved in several countries and in several other assessments (CERA, 2010). The Bt11 maize is approved for cultivation in Argentina, Brazil, Colombia, Canada, Japan, Philippines, South Africa, USA and Uruguay; and approved for animal and/or human consumption in Argentina, Australia, Brazil, Canada, China, Colombia, EU, Japan, Korea, Mexico, Philippines, Russia, South Africa, Switzerland, Taiwan, UK, USA, and Uruguay. The GA21 maize is approved for cultivation in the USA, Canada, Japan, Argentina, and Brazil; and approved for animal and/or human consumption in Argentina, Australia, Brazil, Canada, China, EU, Japan, Korea, Mexico, Philippines, Russia, Taiwan, South Africa, and the USA. The MIR162 maize is approved for cultivation in Brazil and for animal and/or human consumption in Brazil, Australia, USA, and Taiwan; and is still under investigation in several other countries, including Indonesia, Japan, Korea, Philippines, Russia, Argentina, Colombia , Canada, Mexico, and USA (for cultivation).
The Bt11xGA21 maize, obtained by classical genetic improvement from the sexual reproduction of maize lines, containing these events separately, is approved for cultivation in Brazil, USA, Canada, and Japan; and approved for animal and/or human consumption in Brazil, USA, Canada, Japan, Mexico, Philippines, and Korea. The Bt11xMIR162xGA21 maize is under assessment in Japan, Taiwan, Europe, South Africa, Switzerland, Argentina, Colombia, and Uruguay.
III. Expressed proteins
By using classical genetic improvement, the Bt11xMIR162xGA21 maize was developed by means of sexual reproduction of maize lines, individually containing the events BT11, MIR162, and GA21.
The Bt11 event includes cry1Ab gene from Bacillus thuringiensis, which confers resistance to certain lepidopteran insects and the pat gene, derived from the soil micro-organism Streptomyces viridochromogenes, used as selection marker during the transformation process. The Cry1Ab protein is proteolytically cleaved in the alkaline gut of lepidopteran insects, resulting in insecticide active form, which, interacting with a receptor molecule present only in cells of the midgut epithelium of susceptible insects, generates pores in cell membranes, leading to its lysis. Several specific docking sites with high affinity for several Bt proteins have been identified in the midgut epithelium of susceptible insects, showing that the insecticidal protein of cry1Ab gene is highly specific to some lepidopteran insects (Hofte and Whiteley, 1989; Melin and Cozzi, 1990). The molecular description of the Bt11 event and the information for maize risk assessment were presented earlier in the process 01200.002109/2000-04, for commercial release of Bt11 maize.
The MIR162 maize was obtained from vip3Aal9 gene, which confers resistance to lepidopteran insects, and from manA gene, which encodes the enzyme phosphomannose isomerase (PMI), used as a selection marker. A modification caused by the transformation process resulted in a difference in two codons vip3Aa19 inserted gene, then was called vip3Aa20 in maize MIR162 (Entrez Accession number DQ539888; NCBI 2006). This difference resulted in the modification of only one amino acid, located beyond the proteolytic cleavage site of the protein then called Vip3Aa20, expressed in MIR162 maize, thus maintaining its insecticidal properties against several lepidopterous pest of maize harvest. The manA gene was obtained from Escherichia coli strain K-12 and PMI protein expression was used as a selection marker during the MIR162 maize transformation process. The molecular description of the MIR162 event and the information for MIR162 maize risk assessment were presented earlier in the process 0 1200.007493/2007-08, for commercial release of MIR162 maize.
The GA21 event contains the mepsps gene that expresses the enzyme 5-Synthase enolpyruvyl-shikimate-3-phosphate (mEPSPS). The EPSPS is a key enzyme for the shikimic acid process, involved in the biosynthesis of aromatic amino acids (phenylalanine, tyrosine and tryptophane), found naturally in plants, fungi and bacteria, and absent in animals. The EPSPS is highly sensitive to herbicides containing glyphosate. Maize plants transformed with the mutant gene epsps (mepsps), such as those from GA21 event, synthesise the mEPSPS protein that confers tolerance to herbicides containing glyphosate. The molecular description of the GA21 event and the information for GA21 maize risk assessment were presented earlier in the process 01200.002293/2004-16, for commercial release of GA21 maize.
A comparative molecular analysis was performed in order to (a) confirm the genetic integrity of the inserts Bt11, MIR162, and GA21 and (b) to obtain Bt11xMIR162xGA21 maize by using sexual reproduction between maize lines containing these events along the process of classical genetic improvement used. The hybridized fragments Bt11xMIR162xGA21 maize showed the expected size for the events Bt11, MIR162, and GA21. It demonstrates that the integrity of the inserts was maintained during the process of classical genetic improvement to combine such events. These findings are results from Southern blot analysis for all events presented by the applicant. A comparative analysis for the events was conducted. The genomic DNA isolated from leaf tissue of plants for each maize was used. The DNA was digested with restriction enzymes specific for cassettes, as detailed below:
• probe specific for cry1Ab, of 1848 PB. The maize genomic DNA was digested with restriction enzymes NdeI , SphI e BglII+EcoRI, and after electrophoresis and membrane transfer, hybridized with a probe specific for cry1Ab (1848 bp).
• with the probe specific for vip3Aa19 of 2370 bp. The maize genomic DNA was digested with restriction enzymes KpnI, EcoRV and NcoI and, after electrophoresis and membrane transfer, hybridized with a probe specific for vip3Aa19 (2370 bp).
• probe specific for mepsps, of 1338 pb. The maize genomic DNA was digested with restriction enzymes HindIII, SacI and SphI and after electrophoresis and transfer to membrane, hybridized with a probe specific for mepsps (1338 bp).
The probe specific for cry1Ab resulted in identical hybridisation bands for Bt11 maize and for Bt11xMIR162xGA21 maize, showing that the Bt11 event was inherited stably in combined maize.
The probe for vip3Aa20, of which gene has a single change in a amino acid, hybridized completely and resulted in identical banding patterns for MIR162 maize and Bt11xMIR162xGA21 maize. Thus, it was found that the gene was inherited stably.
The probe resulted in mepsps hybridizing bands specific for the gene, identical to the GA21 maize and Bt11xMIR162xGA21 maize, revealing the stable form of inheritance.
As shown, all events hybridized, as expected.
A segregation analysis by using specific test event was conducted in genetically modified maize plants to conserve the isolated and combined events. The results of segregation obtained by ELISA (phenotypic) and qPCR (genotype) were subjected to adherence test (X2). The data indicate that the loci of isolated events, as well as combined events, segregated independently, with typical Mendelian proportions.
IV. Aspects related to human and animal health
The safety for animal/human food of Cry proteins was confirmed by several authors, also by Xu et at. (2009). It was observed that the protein was rapidly degraded by Cry1Ab/Ac gastric and intestinal fluids, and showed no adverse effects in mice subjected to an acute dose of 5g (Cry1Ab/Ac protein)/kg of body weight. Again, it was observed that this protein has no sequence homology with known toxins or allergy, or N-linked glycosylation sites, confirming that no harm resulted from the inclusion of protein Cry1Ab/Ac in food or animal feed.
Another twenty-eight-day study in rats conducted by Onose et al. (2008) showed no adverse effects can be attributed to diet containing Cry1Ab, whereas the administration of diet containing Cry1Ab protein had no significant effect on any biochemical or physiological parameter, but a lower concentration of aspartate aminotransferase - AST in serum of animals receiving such maize, when compared with control. However, no changes in weight or histopathology were observed in organs such as heart, liver, and kidneys. Also, generally is observed that serum levels of AST are elevated with tissue injury, but the interpretation of relatively small changes in the levels of AST in toxicology studies should be done sparingly, since the range of variation of this parameter can be widespread in healthy animals. The reduction in AST in this experiment, therefore, it is not considered to be toxicologically significant. In addition, Paul et al. (2009) studied the degradation of Cry1Ab protein in GM maize regarding the total protein in the digestion of dairy cows. The results indicated that Cry1Ab protein is increasingly degraded during digestion in these animals in small fragments of 42kDa, 34kDa and 17kDa.
As the EPSPS protein, similar to that expressed by the GA21 maize, new studies on the biosafety of the EPSPS protein were released in recent years. Lundry et al. (2009) investigated the differences in the levels of nutrients and anti-nutrients from soya beans expressing EPSPS protein and conventional soy beans, with similar genetic profiles. The results of the comparisons showed that GM soya beans is equivalent in composition and nutritionally equivalent to conventional soya beans marketed currently. Recent studies in mice and cattle showed results consistent with the risk assessment conducted for the EPSPS protein. Healy et al. (2008) presented the results of a study of thirteen weeks of feeding rats with GM maize, expressing CryBb1 and CP4 EPSPS. The responses of rats fed with diets containing this variety of maize were comparable to those of rats fed with a diet containing grains of its nearly isogenic control variety, confirming that GM maize is as safe and nutritious as grain from existing commercial maize hybrids. As EPSPS protein, similar to that expressed by the GA21 maize, new studies on the biosafety of the EPSPS protein were released in recent years. Lundry et al. (2009) investigated the differences in the levels of nutrients and anti-nutrients from soya bean protein expressing EPSPS and conventional soy beans, with similar genetic profiles. The results of the comparisons showed that GM soya beans is equivalent in composition and nutritionally equivalent to conventional soya beans marketed currently. Recent studies in mice and cattle showed results consistent with the risk assessment conducted for the EPSPS protein.
The effects of Vip3Aa20 protein in 10 species of non-target organisms were assessed employing parts of maize plants that accumulate the Vip3Aa20 protein or Vip3Aal9 protein. These proteins differ in a single amino acid located at position 129. However, this difference does not change the protein action in its final form: this position is out of place and region of the triptych final peptide that has cytotoxic activity (cleavage of the peptide and performed at the initial position 199). In all cases, samples of maize were tested against a sensitive insect species to be sure of the presence of toxic levels of VIP proteins. Tests with water fleas, ladybugs, lacewings (larvae and adults), earthworms, catfish, bees, and beetles did not reveal significant differences between control and treated plants containing Vip3Aa20 or Vip3Aal9 (Chen et al 2008).
V. Environmental aspects
The Bt11xMIR162xGA21 maize and a GM product that provides resistance to several lepidopteran pests and for tolerance to glyphosate herbicide, developed by classical breeding by sexual reproduction among maize lines with the Bt11 event, MIR162 event, and GA21 event. Thus, this product contains no other event in genetic transformation, besides the events mentioned. The Bt11 maize, MIR162 maize, and GA21 maize have been approved for commercial release by CTNBio. Every individual matters of biosafety for all such events have already been adequately treated by CTNBio at during the assessment of individual events. Regarding the process of sexual reproduction of these events, there are some important aspects that must be disclosed. A comparative molecular analysis of Bt11xMIR162xGA21 maize showed that the integrity of the inserts was maintained during the process of classical genetic improvement with the aim of combining the events. The segregation analysis of the Bt11xMIR162xGA21 maize showed that genes of Bt11 event, MIR162 event and GA21 event are independent. The agronomic assessment and Bt11xMIR162xGA21 maize efficiency assessment indicated that the combination of these events by classical genetic improvement methods did not lead to expression of any other characteristic other than those expected, i.e. resistance to certain insects and tolerance to the herbicide glyphosate. The expression of proteins Cry1Ab, VIP3Aa20, and mEPSPS observed in Bt11xMIR162xGA21 maize did not show a trend to changes in expression levels. It occurred due to the combination of these events by classical genetic improvement. In the tests, the Bt11xMIRI62xGA21 maize did not present adverse effect on the community of non-target insects. Results showed the effectiveness in controlling Bt11xMIR162xGA21 maize attack by Spodoptera frugiperda, Agrotis ipsilon Elasmopalpus lignosellus and Diatraea saccharalis, when compared with their isogenic non-GM hybrid.
Agronomic parameters were assessed as female and male flowering, leaf diseases, plant height, and height of insertion of the ears. The data presented by the applicant showed no differential expression of any other characteristic. The applicant stated that there were no effects different from those expected. The applicant considered the data sets of three locations and the average of the treatments were bought by the Tukey test at 5% significance level.
A study was conducted to assess the influence of Bt11xMIR162xGA21 maize on the insect community involving environmental conditions in Brazil during the 2008/2009 harvest. Insect traps were used, samples were taken fortnightly. The abundance and diversity were assessed. The data indicated that BtllxMIR162xGA21 maize did not present adverse effect on the insect community or on the predator Doru luteipes, when compared to its isogenic hybrid, non-GMO.
VI. Restrictions on use of GMOs and their derivatives
As established in art. 1 of Law 11,460 of 21 March 2007, 'the research and cultivation of GM organisms are banned for indigenous lands and for areas of conservation'.
The report submitted by the applicant showed that there was no synergistic effect between events, resulting from crosses conventional in terms of agronomic characteristics, way to reproduce, to disseminate, or to survive. Thus, the cultivation and consumption of Bt11xMIR162xGA21 maize are not potentially cause of significant environmental degradation or risk to human and animal health. For these reasons, there are no restrictions on the use of maize and its derivatives, except in places covered by Law 11,460, of 21 March 2007.
It must be emphasized that the lack of negative effects resulting from cultivation of transgenic maize does not mean they cannot happen. Zero risk and absolute security does not exist in the biological world, although there is an accumulation of reliable scientific information and a history of safe use of transgenic varieties in agriculture. Thus, the applicant should lead monitoring post-commercial release in accordance with Normative Resolution No 5 of CTNBio and in accordance with such opinion.
VII. Considerations on particularities of different regions of the country (subsidies to monitoring agencies)
As established in art. 1 of Law 11,460, of 21 March 2007, 'research and cultivation of GM organisms are banned for indigenous lands and for areas of conservation'.
VIII. Conclusion
Based on the evidence of current scientific literature available to the proteins Cry1Ab, VIP3Aa20, and mEPSPS and evidence presented on the lack of interaction among the genes for insect resistance in Bt11xMIR162xGA21 maize, CTNBio held that they are as safe as their conventional equivalent, compared to their food and environmental security. Agronomic and effectiveness assessment conducted in Bt11xMIR162xGA21 maize, indicated no consistent pattern to suggest biologically significant changes resulting from the combination of these events, different from those expected, i.e. insect resistance and combined herbicide tolerance. There is a body of evidence obtained in the results of comparative molecular analysis from analysis of the pattern of genetic inheritance and from comparative analysis of expression levels of proteins Cry1Ab, Vip3Aa20, and mEPSPS in Bt11xMIR162xGA2 maize. Thus, this body indicates that exposure levels to non-target organisms and in animal and human food to these proteins are the same as for Bt11 maize, GA21 maize, and MIR162 maize, separately. Moreover, the results from the influence assessment of Bt11xMIR162xGA21 maize on the insect community did not indicate the existence of adverse effects in these non-target organisms.
The pyramid in question is the result of conventional breeding of three events already approved for planting and consumption by CTNBio. Every individual matters of biosafety have been properly addressed by the committee.
There are four aspects to be assessed in the case of pyramiding, as indicated by regulatory agencies in the world: the stability of building in the breeding, the interaction among metabolic pathways, in which products of inserted genes take part, the change in weediness by the accumulation of phenotypes generated by pyramided genes and identification of events in pyramiding, via PCR with primers, shaping as a ring in flanker regions.
In the first case, the experimental data of proponent indicates that the three genetic markers are stable and inherited Mendelianly, as expected.
In the second case, the three roads are quite different: only two of them involve the synthesis of products that will not be employed by the plant but swallowed by the target insect, being two toxins with very different mechanisms of action. The third way involves resistance to glyphosate, as can be seen in the review article by Tan et al. (2006). There is no theoretical reason to expect any interaction and the agronomic results of hybrid also do not suggest so.
In the third case, resistance to glyphosate did not increase the competitiveness of the hybrid in non-agricultural and insect resistance, per se, has already been discussed in this context in the case of commercial release of the events Bt11 and MIR162.
In the fourth case, there are descriptions of primers that uniquely identify the three events pyramided. Recently, Xu et al (2009) described an improvement of real-time PCR, called UP-M-PCR, and demonstrated its applicability in this event on the agenda. The process can, in principle, be applied to any other event pyramid.
Taking into account that this genotype was built by using classical genetic improvement, and that it resulted in the inheritance of a stable and functional copies of genes cry1Ab, vip3Aa20, and mEPSPS, which provided resistance to insects and tolerance to glyphosate;
Taking into account that composition data showed no significant differences between GM and conventional varieties, suggesting the nutritional equivalence between
them;
Taking into account that the CTNBio assessed the events separately and gave its favourable opinion to its commercial release;
Taking into account also that:
1. The events Bt11, MIR162, and GA21 were characterised during the individual approval, being attested to maintain the integrity of the gene constructs inherited from their parents during the process of classical genetic improvement;
2. There is no evidence of indication among the metabolic pathways that act on proteins Cry1Ab, Vip3A, and mEPSPS
3. There were no reported pleiotropy or epistasis in the parental events and together;
4. The expression of proteins in maize pyramid is not significantly different from the expression events observed in the parental separately;
5. No evidence that the expressed proteins to cause allergic or toxic effect on humans and animals was found;
6. The efficacy and agronomic evaluations of maize indicated that the combination of events by sexual reproduction did not lead to expression of any other features that no one expected, i.e. resistance to some insects and tolerance to glyphosate;
7. There were no botanical changes in the Bt11xMIR162xGA21 maize that may confer adaptive advantages;
8. Internationally accepted criteria in the process of risk analysis of GM raw materials in relation to events pyramided;
It is possible to conclude that BtlIxMIR162xGA21 maize is as safe as its conventional equivalent.
CTNBio considers that this activity does not potentially cause significant degradation of the environment or of harm to human and animal health. Restrictions on the use of GMOs in analysis and its derivatives are subject to the provisions of Law 11,460 of 21 March 2007.
Regarding the monitoring plan post-commercial release, CTNBio determines that instructions are followed and implemented techniques to monitor the actions listed below:
I) – Instructions
a) Monitoring should be conducted in commercial fields and not in experimental fields. The areas chosen to be monitored should not be isolated from the others, have borders or any situation that is out of standard business.
b) Monitoring should be carried out in comparison model between the conventional system of cultivation and cropping system of GMOs, and the data collection done by sampling.
c) Monitoring should be conducted in representative biomes of the main areas of cultivation of GMOs and, where possible, involve different types of producers.
d) Monitoring should be conducted for at least five years.
e) For all monitoring, the proponent should detail information on all activities performed in the pre-planting and planting, on their implementation, with reports of activities conducted in the areas of monitoring during the crop cycle, about the activities of harvest and weather conditions.
f) There should also be monitoring for any adverse effects to human and animal health systems by means of official reporting of adverse effects, such as the SINEPS System (Adverse Event Reporting Related to Health Products), regulated by ANVISA.
g) The analytical methods, results and their interpretations must be developed in accordance with the principles of independence and transparency, subject to commercial confidentiality issues previously defined and justified as such.
h) Based on scientific and technical justifications, CTNBio reserves the right to revise this opinion at any time.
II) – Monitoring technique actions to be performed:
1 – Regarding the cp4 epsps gene, which confers resistance to the herbicide should be monitored:
a) Nutritional status and health of GM plants.
b) Chemical and physical attributes related to soil fertility and other basic soil characteristics.
c) Soil microbial diversity. d) Bank of diaspores in the soil.
e) Community of weed.
f) Development of herbicide resistance in weeds.
g) Glyphosate residues in soil, in grain and in aerial parts.
h) Gene Flow.
2 – With respect to genes cry1Ab and Vip3A, which confer resistance to insects must be monitored:
a) Impact on target insects and non-target insects.
b) Impact on soil invertebrates of indicators, not belonging to the class Insecta.
c) Residues of insecticidal proteins in decomposing organic matter, soil and waterways near the tracking area.
d) Development of resistance among target insects.
e) Gene Flow of the two inserted genes.
The assessment of CTNBio considered the opinions expressed by members of the Commission, by ad hoc consultants, contributed documents in the Executive Secretariat of CTNBio by the applicant; results of planned releases in the environment, lectures, texts, etc. Independent scientific studies and publications of the applicant were considered and consulted, and also held by third parties.
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