DD2006-60: Determination of the Safety of BASF's Imidazolinone-Tolerant CLEARFIELD™ Wheat Events BW255-2 and BW238-3
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This Decision Document has been prepared to explain the regulatory decision reached under Directive 94-08 (Dir94-08), entitled "Assessment Criteria for Determining Environmental Safety of Plants with Novel Traits", its companion biology document Bio1999-01, The Biology of Triticum aestivum L. (Wheat), and Directive 95-03 (Dir95-03), entitled "Guidelines for the Assessment of Novel Feeds: Plant Sources".
The Canadian Food Inspection Agency (CFIA), specifically the Plant Biosafety Office (PBO) of the Plant Products Directorate and the Feed Section of the Animal Health and Production Division have evaluated information submitted by BASF . This information is in regard to the imidazolinone tolerant wheat events BW255-2 and BW238-3. The CFIA has determined that these plants with a novel trait (PNT) and novel feed do not present altered environmental risk nor do they present livestock feed safety concerns when compared to currently commercialized wheat varieties in Canada.
Unconfined release into the environment and use as livestock feed of the wheat events BW255-2 and BW238-3 are therefore authorized as of June 22, 2006. Any wheat lines derived from BW255-2 and BW238-3 may also be released into the environment and used as livestock feed, provided (i) no inter-specific crosses are performed, (ii) the intended uses are similar, (iii) it is known, following thorough characterization, that these plants do not display any additional novel traits and are substantially equivalent to currently grown wheat, in terms of their potential environmental impact and livestock feed safety.
The wheat events BW255-2 and BW238-3 are subject to the same phytosanitary import requirements as their unmodified counterpart.
Please note that the assessment of livestock feed safety and environmental safety are critical steps in the potential commercialization of these plant types. Other requirements, such as the evaluation of food safety by Health Canada, have been addressed separately from this review.
Table of Contents
- Development Method
- Imidazolinone Tolerance
- Stable Expression
- Potential of events BW255-2 and BW238-3 to Become Weeds of Agriculture or Invasive of Natural Habitats
- Potential for Gene Flow from events BW255-2 and BW238-3 to Wild Relatives Whose Offspring May Become More Weedy or More Invasive
- Altered Plant Pest Potential of events BW255-2 and BW238-3
- Potential Impact on Non-Target Organisms of events BW255-2 and BW238-3
- Potential Impact on Biodiversity of events BW255-2 and BW238-3
- Potential Impact of events BW255-2 and BW238-3 on Livestock Nutrition
- Potential Impact of events BW255-2 and BW238-3 on Livestock and Workers/By-standers
I. Brief Identification of the Modified Plant
Designation(s) of the Plant: CLEARFIELD™ wheat events BW255-2 and BW238-3
Applicant: BASF Canada
Plant Species: Wheat (Triticum aestivum)
Novel Traits: Tolerance to imidazolinone herbicides
Trait Introduction Method: Chemically induced seed mutagenesis
Proposed Use of Plant: Production of wheat for livestock feed and human food.
II . Background Information
BASF has developed wheat events tolerant to imidazolinone herbicides. These wheat events were developed to provide an alternative strategy for weed control.
The development of the CLEARFIELD™ wheat events BW255-2 and BW238-3 was accomplished using chemically induced seed mutagenesis. The herbicide tolerance results from a single point mutation modification of the acetohydroxyacid synthase (AHAS) gene such that the resulting enzyme has a single amino acid substitution and is no longer affected by imidazolinone herbicides. Previously authorized wheat events SWP965001, Teal 11A, AP602CL and AP205CL contain AHAS genes with the same mutation.
BASF has provided data on the identity of the wheat events, a detailed description of the modification method and breeding history, information on the modified gene, the resulting protein and its mode of action and the stability of trait expression.
Events BW255-2 and BW238-3 were field tested in multiple locations in Canada in 1999 and in the United States in 2003.
Agronomic characteristics of events BW255-2 and BW238-3 such as grain yield, heading date, plant height, foliar disease and test weight were compared to those of unmodified wheat counterparts.
Nutritional components of events BW255-2 and BW238-3 such as proximates, amino acids and fatty acids were compared with unmodified wheat counterparts. Anti-nutritional factors were also determined.
BASF has provided an agronomic stewardship plan for imidazolinone tolerant wheat in the Canadian environment. This plan includes information regarding a safe and sustainable deployment of imidazolinone-tolerant wheat and an efficient mechanism for growers to report agronomic problems with this product to BASF.
The Plant Biosafety Office (PBO) of the Plant Products Directorate, CFIA, has reviewed the above information, in light of the assessment criteria for determining environmental safety of PNTs, as described in the Directive 94-08 (Dir94-08), entitled "Assessment Criteria for Determining Environmental Safety of Plants With Novel Traits". The PBO has considered:
- potential of events BW255-2 and BW238-3 to become weeds of agriculture or be invasive of natural habitats;
- potential for gene flow from events BW255-2 and BW238-3 to wild relatives whose hybrid offspring may become more weedy or more invasive;
- potential of events BW255-2 and BW238-3 to become a plant pest;
- potential impact of events BW255-2 and BW238-3 or their gene products on non-target species, including humans; and
- potential impact of events BW255-2 and BW238-3 on biodiversity.
The Feed Section of the Animal Health and Production Division, CFIA, has also reviewed the above information with respect to the assessment criteria for determining the safety and efficacy of livestock feed, as described in Directive 95-03 (Dir95-03), entitled "Guidelines for the Assessment of Novel Feeds: Plant Sources". The Feed Section has considered:
- potential impact of events BW255-2 and BW238-3 on livestock nutrition; and
- potential impact of events BW255-2 and BW238-3 on livestock and workers/by-standers.
III. Description and Assessment of the Novel Trait
1. Development Method
The original mutant events BW255-2 and BW238-3 were isolated from populations derived by chemical-induced mutagenesis of seed of the wheat varieties BW255 and BW238 respectively, with sodium azide. Whole plant selection procedures for herbicide tolerance were used. One herbicide tolerant mutant of each line was selected and were designated BW255-2 and BW238-3.
The BW255-2 and BW238-3 varieties are hexaploid (42 chromosomes n=14), belonging to genus and species Triticum aestivum.
2. Imidazolinone Tolerance
Imidazolinone herbicides are active against the enzyme acetohydroxyacid synthase (AHAS), also known as acetolactate synthase (ALS). AHAS is an enzyme found in bacteria, certain other micro-organisms and plants. This enzyme catalyses the first step in the biosynthesis of the essential branched chain amino acids isoleucine, leucine and valine. Herbicide induced AHAS inhibition results in a lethal decrease in protein synthesis. Unmodified wheat is not tolerant to imidazolinone herbicides.
A single amino acid substitution in the AHAS enzyme was sufficient to alter the binding site for imidazolinone herbicides, resulting in the tolerant phenotype.
The novel imidazolinone tolerance is under the control of the native AHAS promoter and is believed to be constitutively expressed. Sequence information for the modified AHAS gene in BW255-2 and BW238-3 was submitted.
The tolerance to imdazolinone was demonstrated by comparison of the activity of the AHAS enzyme extracted from BW255-2 and BW238-3 wheat plants to that of conventional type wheat plants.
The levels of valine, leucine and isoleucine produced in wheat are regulated by feedback inhibition of AHAS. BASF provided data to demonstrate that the modified AHAS shows similar feedback inhibition by valine and leucine as compared to unmodified AHAS. The modification of the AHAS does not affect feedback inhibition and hence, the regulation and levels of these amino acids.
Unlike known food allergens, AHAS is a minor protein in plant tissue, it is heat sensitive and trypsin susceptible. The AHAS protein from BW255-2 and BW238-3 was shown to be heat sensitive, with no detectable activity of AHAS after 1 min of heating at 100°C. AHAS from BW255-2 and BW238-3 were shown to be equivalent to parental controls with respect to trypsin degradation. The unmodified form of the AHAS protein shows no amino acid similarity to known allergens. The amino acid sequence of mutated AHAS differs by one amino acid from that of unmodified wheat.
BASF provided evidence to show that the protein components of BW255-2 and BW238-3 are not altered in comparison with an unmodified comparator. HPLC was run on protein extracts from unmodified and modified wheat to indicate that no new major proteins or increased protein expression occurred as a result of the mutagenic event.
BASF has provided to the CFIA a method for the detection and identification of wheat containing this modified AHAS gene.
3. Stable Expression
The segregation of herbicide tolerance in generation three BW255-2 and BW238-3 populations was shown to be consistent with the inheritance of a single gene.
IV. Criteria for the Environmental Assessment
1. Potential of events BW255-2 and BW238-3 to Become Weeds of Agriculture or Invasive of Natural Habitats
The centre of origin of wheat is considered to be in the Middle East. Modern wheat does not have high potential for weediness and its survival outside cultivation is limited to short periods. Wheat plants can grow as volunteers in a cultivated field following a wheat crop and are usually eliminated via cultivation or the use of herbicides. After hundreds of years of cultivation in North America and throughout the world, there have been no reports of wheat becoming an invasive pest.
The CFIA evaluated data submitted by BASF on the biology of wheat events BW255-2 and BW238-3, and determined that vegetative vigour, time to maturity, seed production, as well as resistance to disease, were within the normal range of expression of these traits currently displayed by the parent line.
No competitive advantage was conferred to these plants, other than that conferred by tolerance to imidazolinone herbicide. Tolerance to imidazolinone herbicide will not, in itself, render wheat weedy or invasive of natural habitats since none of the reproductive or growth characteristics were modified. Imidazolinone tolerance will not cause BW255-2 or BW238-3 to become more weedy or invasive in managed habitats than unmodified T. aestivum . Imidazolinone-tolerant wheat volunteers will not be controlled in subsequent crops if imidazolinone is used as the sole weed control tool. However, control of imidazolinone tolerant wheat as a volunteer weed in other crops or in fallow ground, can readily be achieved by the use of classes of herbicides other than imidazolinones, or by mechanical means.
The novel trait has no intended or observed effects on weediness or invasiveness. The CFIA has therefore concluded that these wheat events have no altered weed or invasiveness potential in Canada when compared to conventional wheat varieties.
The agronomic stewardship plan, which contains a herbicide tolerance management plan, submitted by BASF was evaluated by the CFIA and determined to be satisfactory. The herbicide tolerant stewardship plan includes recommendations on agricultural practices concerning imidazolinone tolerant wheat and provides an efficient mechanism for growers to report agronomic problems with this product to BASF, facilitating the ongoing monitoring of imidazolinone tolerant wheat. In addition, BASF is required to monitor grower compliance to determine the effectiveness of the stewardship plan and make any changes to the plan as appropriate.
In the longer term, the continued use of imidazolinones on herbicide tolerant crops may provide additional selection pressure for the development of group-2 herbicide tolerant weeds. This could result in the loss of the use of these herbicides and any of their potential benefits. Therefore, BASF will make their stewardship plan readily available to growers and agriculture extension personnel, in both private and public sectors, to promote the careful management practices, such as use of alternate control tools as appropriate to achieve complete weed and volunteer control, recommended to help minimize the development of resistant weed populations.
2. Potential for Gene Flow from events BW255-2 and BW238-3 to Wild Relatives Whose Offspring May Become More Weedy or More Invasive
No known wild Triticum species exist in North America. The weedy relative Aegilops cylindrica, jointed goat grass, is present in winter wheat crops in the United States, but is not reported in Canada and does not readily produce fertile progeny when hybridized with wheat. A. cylindrica is included in the provincial Noxious Weed List in British Columbia to deter the spread of this weed into Canada.
The only weedy species related to wheat in Canada is Agropyron repens, quack grass. A. repens is a troublesome weedy grass of agricultural areas throughout Canada. However, no known naturally-occurring hybrids between wheat and Agropyron species have been reported.
The CFIA has therefore determined that gene flow from BW255-2 or BW238-3 to wild or weedy species in Canada is very unlikely.
3. Altered Plant Pest Potential of events BW255-2 and BW238-3
The intended effect of the novel trait is unrelated to plant pest potential. T. aestivum is not a plant pest in Canada. Additionally, the agronomic characteristics of wheat events BW255-2 and BW238-3 were shown to be within the normal range of conventional wheat varieties.
The CFIA has therefore determined that wheat events BW255-2 and BW238-3 do not present a plant pest concern.
4. Potential Impact on Non-Target Organisms of events BW255-2 and BW238-3
Single amino acid modification of the AHAS enzyme, which alters the herbicide binding site on the enzyme, is the molecular basis for imidazolinone tolerance in wheat events BW255-2 and BW238-3. BASF has submitted data indicating that the modified AHAS is substantially equivalent to its unmodified counterparts. The AHAS enzyme is not a known toxin, does not confer resistance to agricultural pests and is commonly found in a wide variety of plants and micro-organisms with a history of safe use. The mutant AHAS in BW255-2 and BW238-3 has not significantly affected the biosynthesis of the branched- chain amino acids, valine, leucine and isoleucine. The detailed compositional analysis has led to the conclusion that the wheat events BW255-2 and BW238-3 are substantially equivalent to their parent lines.
The AHAS enzyme is not a known toxin, does not confer resistance to agricultural pests and is commonly found in a wide variety of plants and micro-organisms with a history of safe use. No novel toxins were introduced into this variety. Therefore, no negative interactions with non-target symbiotic or consumer organisms are anticipated.
In addition, agronomic characteristics, pathogen interactions, and insect pest susceptibility of events BW255-2 and BW238-3 are expected to be within the range of values displayed by currently commercialized wheat varieties. The CFIA concluded that there were not likely to be significant unintended changes to wheat events BW255-2 and BW238-3 that could have adverse impacts on non target organisms.
5. Potential Impact on Biodiversity of events BW255-2 and BW238-3
Wheat events BW255-2 and BW238-3 are safe to non-target organisms, do not present altered weediness or plant pest potential and are not intended to be grown in Canada. In addition the novel trait has not altered the ability of this line to persist in the Canadian environment.
The CFIA has therefore concluded that the potential impact on biodiversity of wheat events BW255-2 and BW238-3 is equivalent to that of currently commercialized wheat lines.
V. Criteria for the Livestock Feed Assessment
1. Potential Impact of events BW255-2 and BW238-3 on Livestock Nutrition
Nutritional Composition and Anti-Nutritional Factors
Nutritional composition data was obtained from bread wheat events BW255-2 and BW238-3 and their parental control lines BW255 and BW238, respectively, grown in three locations in the United States. Grain samples were analyzed for protein, fat, fibre, branched-chain amino acids (valine, isoleucine, leucine), essential amino acids (cystine, methionine, threonine and lysine), minerals and vitamins. There were statistically significant differences observed in protein levels between events BW255-2 and BW238-3 and their parental controls, but these levels were comparable to conventional bread wheat varieties. No statistically significant differences were observed in essential amino acids (% of total protein) between events BW255-2 and BW238-3 and their parental controls. The branched-chain amino acids (% of total protein) were not different in BW255-2 compared to its parental control, but significant differences were observed between BW238-3 and its parental control. All amino acids in events BW255-2 and BW238-3 were within the range of conventional bread wheat varieties. Statistically significant differences were observed between BW255-2 and its parental control for Mg and Fe, but no differences in minerals were observed in line BW238-3. Vitamin B1 and pantothenic acid in BW255-2 and BW238-3, respectively, were significantly different from their parental controls. However, mineral and vitamin levels in events BW255-2 and BW238-3 were comparable to levels in conventional bread wheat varieties.
Phytic acid and trypsin inhibitor were analyzed in events BW255-2 and BW238-3 grain samples. There were no statistically significant differences in phytic acid between events BW255-2 and BW238-3 and their parental controls. Trypsin inhibitor was not detected in events BW255-2 and BW238-3 or their parental controls.
The evidence provided by BASF supports the conclusion that the nutritional composition of wheat events BW255-2 and BW238-3 is equivalent to conventional bread wheat varieties.
2. Potential Impact of events BW255-2 and BW238-3 on Livestock and Workers/By-standers
The AHAS enzyme is found in a wide variety of plants and micro-organisms. AHAS is not a known toxin or allergen and a single amino-acid change would not be expected to change this. AHAS from events BW255-2 and BW238-3 is feedback inhibited as is unmodified AHAS, it is present in small amounts in the feed, it is heat labile and it is rapidly degraded under conditions in the gastrointestinal tract. The expression of AHAS is not changed by the modification. Based on the information provided by BASF, the modified AHAS is unlikely to be a novel toxin or allergen.
Based on the detailed characterization provided (nutritional composition, agronomic data and HPLC protein profiles of the modified plant compared to the unmodified comparator) it is unlikely that secondary mutations causing unintended effects have occurred in the wheat genome.
VI. New Information Requirements
If at any time, BASF becomes aware of any information regarding risk to the environment, including risk to human or animal health, that could result from release, in Canada or elsewhere, of wheat events BW255-2 and BW238-3, their descendants, or products derived there from, BASF must immediately provide such information to the CFIA. On the basis of such new information, the CFIA will re-evaluate the potential impact of the proposed feed use and environmental release and will re-evaluate its decision with respect to the livestock feed use and environmental release authorizations of these wheat events.
VII. Regulatory Decision
Based on the review of data and information and stewardship plan, submitted by BASF, and through comparisons of BW255-2 and BW238-3 with unmodified wheat counterparts, the Plant Biosafety Office, CFIA, has concluded that the modified gene and its corresponding traits does not confer to wheat events BW255-2 and BW238-3 any characteristic that would result in intended or unintended significant environmental effects following unconfined release.
Based on the review of data and information submitted by BASF, including comparisons of BW255-2 and BW238-3 with unmodified wheat counterparts, the Feed Section, CFIA, has concluded that the modified gene and its corresponding novel trait will not confer to these plants any characteristic that would raise any concerns regarding the safety or nutritional composition of wheat events BW255-2 and BW238-3. Wheat grain, its byproducts and wheat germ oil, are currently listed in Schedule IV of the Feeds Regulations and are, therefore approved for use in livestock feeds in Canada. Wheat events BW255-2 and BW238-3 have been assessed and found to be as safe as nutritious as traditional wheat varieties. BW255-2 and BW238-3 and their products are considered to meet the present ingredient definitions and are approved for use as livestock feed ingredients in Canada.
Unconfined release into the environment and livestock feed use of the wheat events BW255-2 and BW238-3 is therefore authorized as of June 22, 2006. Any other wheat lines derived from these events may be imported and/or released, provided no inter-specific crosses are performed, provided the intended uses are similar, and provided it is known, following thorough characterization that these plants do not display any additional novel traits and are substantially equivalent to currently grown wheat, in terms of their potential environmental impact and livestock feed safety.
The wheat events BW255-2 and BW238-3 are subject to the same phytosanitary import requirements as their unmodified counterparts.
Please refer to Health Canada's Decisions on Novel Foods for a description of the food safety assessment of these CLEARFIELD™ wheat events. The food safety decisions are available at the following Health Canada web site: http://www.hc-sc.gc.ca/fn-an/gmf-agm/appro/index-eng.php
This bulletin is published by the Canadian Food Inspection Agency. For further information, please contact the Feed Section or Plant Biosafety Office at:
Plant Biosafety Office
Plant Products Directorate
59 Camelot Drive, Ottawa
Ontario K1A 0Y9
Animal Health and Production Division
Animal Products Directorate
59 Camelot Drive, Ottawa
Ontario K1A 0Y9
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