DD2007-66: Determination of the Safety of BASF's Imidazolinone-Tolerant Clearfield® Wheat Event BW7
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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 Biotechnology Environmental Release Assessment Unit (BERA) of the Science Strategies Directorate and the Animal Feed Division of the Animal Health Directorate have evaluated information submitted by BASF. This information is in regard to the imidazolinone tolerant wheat event BW7. The CFIA has determined that this plant with a novel trait (PNT) does not present altered environmental risk nor, as a novel feed, does it present livestock feed safety concerns when compared to currently commercialized wheat varieties in Canada.
Taking into account these evaluations, unconfined release into the environment and use as livestock feed of the Clearfield® wheat event BW7 is therefore authorized by the Plant Biosafety Office (PBO) of the Plant Health and Biosecurity Directorate and the Animal Feed Division of the Animal Health and Production Division as of March 21, 2007. Any wheat lines derived from BW7 may also be released into the environment and used as livestock feed, provided (i) no inter-specific crosses are performed, (ii) the intended use(s) 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 event BW7 is subject to the same phytosanitary import requirements as its 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
I. Brief Identification of the Modified Plant
II. Background Information
III. Description and Assessment of the Novel Trait
- Development Method
- Imidazolinone Tolerance
- Stable Expression
IV. Criteria for the Environmental Assessment
- Potential of event BW7 to Become a Weed of Agriculture or Invasive of Natural Habitats
- Potential for Gene Flow from event BW7 to Wild Relatives Whose Offspring May Become More Weedy or More Invasive
- Altered Plant Pest Potential of event BW7
- Potential Impact on Non-Target Organisms of event BW7
- Potential Impact on Biodiversity of event BW7
V. Nutritional Criteria Assessment as Livestock Feed
- Potential Impact of event BW7 on Livestock Nutrition
- Potential Impact of event BW7 on Livestock and Workers/By-standers
VI. New Information Requirements
VII. Regulatory Decision
I. Brief Identification of the Modified Plant
Designation(s) of the Modified Plant: Clearfield® wheat event BW7
Applicant: BASF Canada
Plant Species: Wheat (Triticum aestivum L.)
Novel Traits: Tolerance to imidazolinone herbicides
Trait Introduction Method: Chemically induced seed mutagenesis
Proposed Use of the Modified Plant: Production of wheat for livestock feed and human food
II. Background Information
BASF has developed a winter wheat event tolerant to imidazolinone herbicides. This wheat event was developed to provide an alternative strategy for weed control.
The development of the Clearfield® wheat event BW7 was accomplished using chemically induced seed mutagenesis. Event BW7 is an allohexaploid (42 chromosomes, n=21), belonging to genus and species Triticum aestivum L. Wheat carries three complete genomes designated "A", "B", and "C". Each genome has an AHAS gene (Als3, Als2, and Als1, respectively). The herbicide tolerance in event BW7 results from a single point mutation modification in the Als1 acetohydroxyacid synthase (AHAS) gene in the wheat genome such that the resulting modified enzyme has a single amino acid substitution and is no longer affected by imidazolinone herbicides.
BASF has provided data on the identity of the wheat event, 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.
Event BW7 was field tested in the United States of America in 2002, 2003, 2004, and 2005.
Agronomic characteristics of event BW7 such as seed germination, seedling emergence, seeding vigour, foliar disease susceptibility, insect damage, plant height, days to heading, lodging, harvest moisture, test weight and grain yield were compared to those of unmodified wheat counterparts.
Nutritional components of event BW7 such as proximates, amino acids and fatty acids were compared with unmodified wheat counterparts. Levels and/or absences of 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 Biotechnology Environmental Release Assessment (BERA) Unit of the Science Strategies Division, 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". BERA Unit has considered the:
- potential of event BW7 to become a weed of agriculture or be invasive of natural habitats;
- potential for gene flow from event BW7 to wild relatives whose hybrid offspring may become more weedy or more invasive;
- potential of event BW7 to become a plant pest;
- potential impact of event BW7 or its' gene products on non-target species, including humans; and
- potential impact of event BW7 on biodiversity.
The Animal Feed Division of the Animal Health Directorate, 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 Animal Feed Division has considered the:
- potential impact of event BW7 on livestock nutrition; and
- potential impact of event BW7 on livestock and workers/by-standers.
III. Description and Assessment of the Novel Trait
1. Development Method
The original mutant event BW7 was isolated from populations derived by chemical-induced mutagenesis with sodium azide of wheat seed. Whole plant selection procedures for herbicide tolerance were used (seedlings were sprayed with imidazolinone herbicides; only tolerant plants lived and were used in the following breeding generations).
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 BW7 was submitted.
The tolerance to imidazolinone was demonstrated by comparison of the activity of the AHAS enzyme extracted from BW7 wheat plants to that of conventional type wheat plants. In the absence of the herbicide, the AHAS enzyme activity was similar, but when imidazolinone herbicide was added to the assay, the enzyme activity of the susceptible plants was inhibited much more than enzyme from the tolerant 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 are unaffected.
Unlike known food allergens, AHAS is a minor protein in plant tissue, and it is heat-sensitive and trypsin-susceptible. The AHAS protein from BW7 was shown to be heat sensitive, with no detectable activity of AHAS after 1 minute of heating at 100 °C. AHAS from BW7 was shown to be equivalent to parental controls with respect to trypsin degradation. The unmodified and modified forms of the AHAS protein show 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 BW7 are not altered in comparison with an unmodified comparator. HPLC performed using protein extracts from unmodified and modified wheat indicated 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 novel trait.
3. Stable Expression
Wheat event BW7 demonstrated consistent herbicide tolerance over multiple breeding generations, confirming the stable inheritance of this trait.
IV. Criteria for the Environmental Assessment
1. Potential of event BW7 to Become a Weed of Agriculture or Invasive of Natural Habitats
Wheat possesses few of the characteristics that are common to weeds and invasive plants. It is an annual crop that does not persist in unmanaged ecosystems without human intervention. Wheat plants can grow as volunteers in cultivated fields in the seasons following a wheat crop, but they are usually eliminated by soil cultivation or the use of herbicides. There have been no reports of wheat becoming an unmanageable weed of agriculture or an invasive pest, in North America or the world.
The CFIA evaluated data submitted by BASF on the agronomic characteristics and biology of event BW7, and determined that seedling vigour, foliar disease susceptibility, insect damage, plant height, days to heading, lodging, grain moisture at harvest, test weight and grain yield were within the normal range of expression of these traits currently displayed by the parent variety. The seed germination and seedling emergence of event BW7 were lower than those of the parent variety, but were still within the normal range of expression of these traits in locally-adapted winter wheat varieties.
No competitive advantage was conferred on plants with event BW7, other than that conferred by tolerance to imidazolinone herbicides, as none of the reproductive or growth characteristics of BW7 were increased relative to those of the parent variety. Tolerance to imidazolinone herbicides provides a competitive advantage only when these herbicides are used, and will not, in and of itself, make a plant weedier or more invasive of natural habitats. Imidazolinone tolerance will not cause BW7 to become weedier or more invasive in managed habitats than unmodified T. aestivum. Imidazolinone-tolerant volunteers will not be controlled in subsequent crops if imidazolinone is used as the only 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 this event has no altered weed or invasiveness potential in Canada when compared to conventional varieties.
In the longer term, widespread adoption of several different crop species with specific associated weed management systems (i.e. numerous crop species each with one or more tolerances to different herbicides) may lead to the development of crop volunteers with a combination of novel tolerances to different herbicides. This could result in the loss of the use of these herbicides and any of their potential benefits.
BASF has submitted an agronomic stewardship plan (which contains a herbicide tolerance management plan) to the CFIA which was determined to be satisfactory when evaluated by BERA. The Clearfield® Wheat Herbicide Tolerance Stewardship Plan comprises the Best Management Practice Program for the Clearfield® Winter Wheat Production System. This stewardship plan also applies to durum wheat events DW1 (see Decision Document DD2006-63), DW2, DW6 and DW12 (see Decision Document DD2007-64) as well as bread wheat events BW255-2 and BW238-3 (see Decision Document DD2006-60).
BASF will make this stewardship plan readily available to growers and agriculture extension personnel, in both private and public sectors, to promote careful management practices for BW7. These practices include the use of alternate control tools as appropriate to achieve complete weed and volunteer control (which is also recommended to help minimize the development of resistant weed populations). 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 BW7 to BASF, which will facilitate 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.
2. Potential for Gene Flow from event BW7 to Wild Relatives Whose Offspring May Become More Weedy or More Invasive
No known wild Triticum species exist in North America. The closest known relatives in North America are members of the Aegilops genus. Aegilops cylindrica, jointed goat grass, is present in the United States where it can be found as a weed in winter wheat fields. It is listed as a Class I Prohibited noxious weed seed in Canada's Weed Seeds Order, 2005. It has been reported in Canada in two locations: a non-persistent introduction in British Colombia in 1997, and an introduction in 2006 in Ontario which is being monitored for persistence by the CFIA's Plant Health program. The Ontario site is not an agricultural site. This minimizes the chances of gene flow occurring from imidazolinone-tolerant T. aestivum as most pollen flow from T. aestivum occurs within a short distance from the wheat plant (Biology Document BIO1999-01: The Biology of Triticum aestivum L. (Wheat).
Research by Guadagnuolo et al. (2001) indicates that gene flow does occur between Triticum aestivum L. and A. cylindrica at a low rate, and that a small percentage of gene flow events could result in viable hybrid offspring. However, it is the CFIA's opinion that there is no increased risk of consequences of gene flow between BW7 imidazolinone-tolerant T. aestivum and A. cylindrica, given the non-persistent nature of the first introduction in B.C., the non-agricultural nature of the Ontario site. and the previous authorizations of bread wheat events BW255-2 and BW238-3 (see Decision Document DD2006-60). Additionally, BASF's Clearfield® Wheat Herbicide Tolerance Stewardship Plan contains recommendations for delaying the development of herbicide resistance in weeds.
The only common weed species found in Canada that is closely related to Triticum aestivum is Agropyron repens (quackgrass). Agropyron repens is a troublesome weedy grass of agricultural areas throughout Canada. However, no known naturally-occurring hybrids between Triticum aestivum and Agropyron species have been reported, and published literature on the genetics of Triticum and Agropyron indicate that natural crossing between these two genera is unlikely. Other wild relatives of wheat do occur in Canada, but it is improbable that hybrids between wheat and these relatives would occur in nature (Bio1999-01, The Biology of Triticum aestivum L. (Wheat)).
Triticale is a crop that has been developed by humans by crossing wheat (Triticum aestivum or T. turgidum) with rye (Secale cereale). There have been no reports of triticale serving as a hybridization bridge between wheat and rye.
The CFIA has therefore determined that the consequences of gene flow from BW7 to wild or weedy species in Canada are of minimal concern at this time. However, these conclusions may need to be revisited if A. cylindrica becomes established.
3. Altered Plant Pest Potential of event BW7
T. aestivum is not a plant pest in Canada, and the intended effect of the novel trait is unrelated to plant pest potential. Additionally, the agronomic characteristics of event BW7 (including the responses to diseases) were shown to be within the normal range of conventional wheat varieties.
The CFIA has therefore determined that event BW7 does not present a plant pest concern.
4. Potential Impact on Non-Target Organisms of event BW7
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 event BW7. BASF has submitted data indicating that the modified AHAS is substantially equivalent to its unmodified counterparts with respect to enzyme activity (biosynthesis of the precursor molecule of valine, leucine, and isoleucine) and feedback inhibition (from valine and leucine). The modified AHAS enzyme only differs from the unmodified enzyme by being less inhibited by imidazolinone herbicides. 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.
The detailed compositional analysis has led to the conclusion that wheat event BW7 is substantially equivalent to its parent line. In addition, agronomic characteristics, pathogen interactions, and insect pest susceptibility of event BW7 is within the expected range of values displayed by currently commercialized varieties. The CFIA concluded that there were not likely to be significant unintended changes to wheat event BW7 that could have adverse impacts on non target organisms.
5. Potential Impact on Biodiversity of event BW7
Wheat event BW7 is safe to non-target organisms and does not present altered weediness or plant pest potential. In addition, the novel trait has not altered the ability of this line to persist in the Canadian environment and it is not expected to expand the range of cultivation of Triticum aestivum in Canada. No changes in current agronomic practices for wheat are expected.
The CFIA has therefore concluded that the potential impact on biodiversity of wheat event BW7 is equivalent to that of currently commercialized wheat lines.
V. Criteria for the Livestock Feed Assessment
1. Potential Impact on Livestock Nutrition
Nutritional Composition and Anti-Nutritional Factors
Nutritional composition data was obtained from wheat event BW7 and a parental control. Grain samples were collected from replicated plots in 8 field sites in 2002, 2003 and 2005. Note that the data from 2002 and 2003 was combined. These samples were analyzed for proximates, ADF, NDF, TDF, amino acids, fatty acids, minerals and vitamins. There were no statistically significant differences between BW7 and the parental control for crude fat, protein, crude fibre, ADF, NDF, TDF in the 2002/2003 and 2005 grain samples. No statistically significant differences were observed between BW7 and the parental control for the branched- chain amino acids (valine, isoleucine and leucine). Lysine, methionine, cystine and threonine levels were not statistically significantly different between BW7 and the parental control. Stearic, oleic and linoleic acids in BW7 were statistically significantly lower than the parental control in the 2002/2003 samples, while palmitic and oleic acids in BW7 were significantly higher than the parental control in 2005. However, all fatty acids levels were shown to be comparable to those of conventional bread wheat varieties. No statistically significant differences were observed between BW7 and the parental control for all minerals in 2002/2003 samples. There were significant differences between BW7 and the parental control for phosphorous and zinc in the 2005 data, however these levels were shown to be comparable to those of conventional bread wheat varieties. All other minerals were similar for BW7 and the parental control. In 2002/2003 vitamin B6 was significantly lower in BW7 than the parental control, but the range was within the range of commercial varieties. No statistically significant differences were observed between BW7 and the parental control for all vitamins analyzed in 2005 samples.
Phytic acid and trypsin inhibitor were analyzed in wheat event BW7 and compared to the parental control. There were no statistically significant differences in phytic acid between BW7 and the parental control for both 2002/2003 and 2005 grain samples. The levels of trypsin inhibitor were below the limit of detection of the assay for both BW7 and the parental control.
The evidence provided by BASF supports the conclusion that the nutritional composition of wheat event BW7 is equivalent to conventional bread wheat varieties.
2. Potential Impact 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 base pair change would not be expected to change this. AHAS from event BW7 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.
The evidence provided by BASF supports the conclusion that the potential impact on livestock and workers/by-standers of wheat event BW7 is equivalent to that of currently commercialized wheat lines.
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 event BW7, its 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 wheat event BW7 on the environment, livestock and human health, and may re-evaluate its decision with respect to the livestock feed use and environmental release authorizations of wheat event BW7.
VII. Regulatory Decision
Based on the review of data and information and the stewardship plan submitted by BASF, and through comparisons of wheat event BW7 with unmodified wheat counterparts, the Biotechnology Environmental Release Assessment Unit, CFIA, has concluded that the modified gene and its corresponding traits does not confer to wheat event BW7 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 event BW7 with unmodified wheat counterparts, the Animal Feed Division, 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 event BW7. 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 event BW7 has been assessed and found to be as safe and nutritious as traditional wheat varieties. BW7 and its' products are considered to meet the present ingredient definitions and are approved for use as livestock feed ingredients in Canada.
Taking into account these evaluations, unconfined release into the environment and livestock feed use of the Clearfield® wheat event BW7 is therefore authorized by the Plant Biosafety Office of the Plant Health and Biosecurity Directorate and the Animal Feed Division of the Animal Health Directorate as of March 21, 2007. Any wheat lines derived from this event may be imported and/or released, provided no inter-specific crosses are performed, provided the intended use is 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 Clearfield® wheat event BW7 is subject to the same phytosanitary import requirements as its unmodified counterparts.
Please refer to Health Canada's Decisions on Novel Foods for a description of the food safety assessment of Clearfield® wheat event BW7.
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