DD2007-65: Determination of the Safety of Pioneer Hi-Bred International's Corn Event TUSC1
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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 Bio1994-11, The Biology of Zea mays L. (Corn/Maize), 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 of the Science Strategies Division and the Feed Section of the Animal Health and Production Division, have evaluated information submitted by Pioneer Hi-Bred International. This information is in regard to the corn event TUSC1. The CFIA has determined that this plant with a novel trait (PNT) and novel feed does not present altered environmental risk nor does it present livestock feed safety concerns when compared to currently commercialized corn varieties in Canada.
Taking into account these evaluations, unconfined release into the environment and use as livestock feed of corn event TUSC1 is authorized by the Plant Biosafety Office (PBO) of the Plant Products Directorate and the Feed Section of the Animal Health and Production Division as of March 21, 2007. Any corn lines derived from TUSC1 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 corn, in terms of their potential environmental impact and livestock feed safety.
The corn event TUSC1 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
- Reduced 27 kDa Gamma Zein Expression
- Stable Expression
IV. Criteria for the Environmental Assessment
- Potential of event TUSC1 to Become Weeds of Agriculture or Invasive of Natural Habitats
- Potential for Gene Flow from event TUSC1 to Wild Relatives Whose Offspring May Become More Weedy or More Invasive
- Altered Plant Pest Potential of event TUSC1
- Potential Impact on Non-Target Organisms of event TUSC1
- Potential Impact on Biodiversity of event TUSC1
V. Nutritional Criteria Assessment as Livestock Feed
- Potential Impact of event TUSC1 on Livestock Nutrition
- Potential Impact of event TUSC1 on Livestock and Workers/By-standers
VI. New Information Requirements
VII. Regulatory Decision
I. Brief Identification of the Modified Plant
Designation(s) of the Plant: Corn event TUSC1
Applicant: Pioneed Hi-Bred International
Plant Species: Corn (Zea mays)
Novel Traits: Reduced 27 kDa gamma zein expression
Trait Introduction Method: Traditional breeding
Proposed Use of the Plant: Production of corn for livestock feed and human food
II. Background Information
Pioneer Hi-Bred International has produced a corn event with reduced 27 kilodalton (kDa) gamma zein expression. This trait is intended to improve the digestibility of the corn.
The development of corn event TUSC1 was accomplished using traditional breeding involving the use of Robertson mutator lines of corn known to have high frequency of Mu transposition events. The reduced 27 kDa gamma zein expression results from the insertion of a Mu transposon element into the 27 kDa gamma zein gene, thereby reducing gene expression by disrupting the function of the gene.
Pioneer Hi-Bred International has provided data on the identity of the corn event, a detailed description of the modification method and breeding history, information on the modified gene and the stability of trait expression.
Event TUSC1 was field tested in 2002 in the United States and Canada.
Agronomic characteristics of event TUSC1 such as early population (germination), seedling vigour, time to silking, time to pollen shed, plant height, ear height, stalk lodging, root lodging, stay green, pollen viability, final population, disease incidence, insect damage, and yield were compared to those of unmodified corn counterparts.
Nutritional components of event TUSC1 such as proximates, amino acids and fatty acids were compared with unmodified corn counterparts. Anti-nutritional factors were also determined.
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 has considered the:
- potential of event TUSC1 to become a weed of agriculture or be invasive of natural habitats;
- potential for gene flow from event TUSC1 to wild relatives whose hybrid offspring may become more weedy or more invasive;
- potential of event TUSC1 to become a plant pest;
- potential impact of event TUSC1 or its gene products on non-target species, including humans;
- and potential impact of event TUSC1 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 event TUSC1 on livestock nutrition; and
- potential impact of event TUSC1 on livestock and workers/by-standers.
III. Description and Assessment of the Novel Trait
1. Development Method
Corn event TUSC1 was created through traditional breeding methods involving the use of Robertson mutator lines of corn known to have high frequency of Mu transposition. Initial crossing involving the Robertson mutator lines was followed by backcrossing to return the frequency of Mu transposition in the resulting events to a normal level and thus "fix" the Mu insertions that had occurred. Molecular techniques were employed to identify events which had a Mu element inserted within the 27 kDa gamma zein gene. Further analysis of one of the selected events, TUSC1, revealed that this event contained reduced levels of the 27 kDa gamma zein protein.
2. Reduced 27 kDa Gamma Zein Expression
The 27 kDa gamma zein protein is an abundant seed storage protein found in corn endosperm cells. Studies have indicated that the gamma zein protein plays an important role in the formation of zein protein bodies, which form within the endoplasmic reticulum of endosperm cells. The 27 kDa gamma zein protein contains 15 cysteine amino acid residues that form crosslinks with other proteins in the maize endosperm and is believed to be a major factor that determines the properties of the storage protein matrix. Disruption of the 27 kDa gamma zein gene could modify seed characteristics known to be affected by the zein proteins, including the opaque and vitreous endosperm phenotypes, as well as the extractability of starch and the digestibility of grain. Increased digestibility of the grain may lead to increased availability of energy when the corn grain is used as livestock feed.
Event TUSC1 was characterized by comparing agronomic and compositional traits against a suitable comparator. Based on the characterization data provided it is unlikely that any unintended effects have occurred in the corn genome. Additionally, event TUSC1 is not expected to have altered expression of any allergens or toxins given that this event was modified to have reduced expression of an endogenous protein.
Pioneer Hi-Bred International has provided to the CFIA a method for the detection and identification of corn containing the TUSC1 event.
3. Stable Expression
The stable inheritance of both the modified gene and the associated trait, reduced expression of the 27 kDa gamma zein protein, were demonstrated over multiple generations of TUSC1 plants using molecular and phenotypic analyses respectively. Chi-square statistical analyses of this data showed that the trait segregated in the offspring in the expected ratio.
IV. Criteria for the Environmental Assessment
1. Potential of corn event TUSC1 to Become a Weed of Agriculture or Invasive of Natural Habitats
The biology of corn, described in the CFIA Biology Document BIO1994-11, shows that unmodified plants of this species are not invasive of unmanaged habitats in Canada. Corn does not possess the potential to become weedy due to the lack of seed dormancy, the non-shattering nature of corn cobs, and the poor competitive ability of seedlings. According to the information provided by Pioneer Hi-Bred International Inc., corn event TUSC1 was determined to be similar to its counterparts in this respect.
Corn event TUSC1 was tested in 4 locations in 2002 in the USA and Canada which provided a range of environmental and agronomic conditions representative of major corn growing regions. For these agronomic studies, TUSC1 was compared to maize hybrid CHP38/N46 (the control), which was identified as a T1 hybrid (near-isoline) that does not contain the Mu transposable element in the gene expressing the 27 kDa protein. The presence of the 27 kDa gamma zein protein in CHP38/N46 was confirmed with western blot analysis.
A total of 12 phenotypic characteristics were evaluated in the 2002 field trials: germination/early population, accumulated heat units to 50% pollen shed, accumulated heat units to 50% silking, ear height, plant height, stay green (a measure of resistance to senescence), final population, stalk lodging, root lodging, disease incidence, insect damage, and pollen viability (the number of accumulated heat units is a method of comparing plant growth rates across different years, and the number is calculated based on the maximum and minimum temperatures for each day of the growing season). Yield data was collected in 2003 and 2004 from replicated multi-location trials. Seed dormancy was measured by two types of germination tests. Data generated from these studies represent observations that are typically recorded by plant breeders and agronomists to evaluate the qualities of maize over a broad range of environmental conditions and agronomic practices that corn event TUSC1 likely would encounter. The measured characteristics provide crop biology data useful in establishing a basis to assess phenotypic equivalence and familiarity of corn event TUSC1 compared to conventional maize in the context of an environmental safety assessment. Detected differences were considered alone, in consideration of other observed differences, and for trends across locations.
There were no differences between TUSC1 and the control with respect to accumulated heat units to 50% pollen shed, accumulated heat units to 50% silking, ear height, stay green,stalk lodging, root lodging, disease incidence, insect damage, pollen viability, yield, or seed dormancy. Plant height was slightly shorter in TUSC1 compared with the control, but the height of TUSC1 plants was within the range of heights of a number of comparable hybrids grown under comparable growing conditions. Germination/early population and final population were each statistically significantly lower in TUSC1 compared with the control, but again, the values were within the range of a number of comparable hybrids grown under comparable growing conditions
Overall, the phenotypic characteristic data showed no biologically meaningful differences between corn event TUSC1 and the control (as compared to a selection of conventional maize hybrids grown in comparable locations). The data supports a conclusion of phenotypic equivalence as it relates to familiarity and a lack of increased weed potential. Likewise, assessment of the phenotypic data detected no biologically significant differences between corn event TUSC1 and the control that would indicate a selective advantage that would result from this trait that would in turn result in increased weed potential for corn event TUSC1 or other plants if the trait were transferred to a sexually compatible species.
The novel trait (reduced 27 kDa gamma zein expression ) has no intended or observed effects on weediness or invasiveness. The CFIA has therefore concluded that this corn event has no altered weediness or invasiveness potential in Canada when compared to conventional corn hybrids.
2. Potential for Gene Flow from event TUSC1 to Wild Relatives Whose Offspring May Become More Weedy or More Invasive
There are no wild relatives of corn reported in Canada. One distant relative (Tripsacum dactyloides) is found in the north-eastern United States of America, but hybridization of this species with corn is difficult and only accomplished with significant human intervention. The CFIA therefore concludes that gene flow from corn event TUSC1 to wild relatives of corn is not possible in Canada.
3. Altered Plant Pest Potential of event TUSC1
The method of trait introduction in TUSC1 (selection from a population of corn that is fixed for Mu-induced mutations) does not require the use of transformation vectors; therefore no new pathogenic sequences have been incorporated into the TUSC1 genome. The intended effect of the novel trait (reduced 27 kDa gamma zein expression) is unrelated to plant pest potential, and corn is not considered a plant pest in Canada. The agronomic characteristics of corn event TUSC1 were shown to be within the normal range of conventional corn varieties. Additionally, ecological evaluations did not show any increase or decrease in insect damage or disease severity (including red milkweed beetles, wireworms, corn rootworm beetles, Japanese beetles, common smut, and fusarium).
The CFIA has therefore determined that corn event TUSC1 does not present a plant pest concern.
4. Potential Impact on Non-Target Organisms of corn event TUSC1
Since there is no novel gene product in TUSC1, no new allergens or toxins have been introduced into this corn event relative to conventional corn. Compositional, agronomic, disease and insect response characteristics were observed in USA and Canadian field trials and did not reveal any biologically significant differences, indicating that TUSC1 is unlikely to have an altered impact on non-target organisms relative to conventional corn.
Based on the above information and the information cited below in Section V. Criteria for the Livestock Feed Safety Assessment, the CFIA has determined that the unconfined release of corn event TUSC1 will not result in altered impacts on interacting organisms, including humans, compared to current commercial corn hybrids.
5. Potential Impact on Biodiversity of event TUSC1
Corn event TUSC1 has no novel phenotypic characteristics that would extend its range beyond the current geographic range of corn production in Canada, or that would alter the ability of corn to survive in the Canadian environment. Additionally, this trait is unlikely to introduce any changes in management practices with respect to currently commercialized corn. Corn event TUSC1 is unlikely to have any other impacts on biodiversity relative to currently commercialized corn hybrids as it is considered substantially equivalent in terms of potential to be weedy or invasive, potential and consequences of gene flow to wild relatives, plant pest potential, and potential impacts on non-target organisms. The CFIA has therefore concluded that the potential impact on biodiversity of corn event TUSC1 is equivalent to that of currently commercialized corn hybrids.
V. Criteria for the Livestock Feed Assessment
1. Potential Impact on Livestock Nutrition
Four field trials in the US and Canada were used to generate nutritional composition data for corn event TUSC1 and line CHP38/N46, an isogenic control. Forage and grain samples were analyzed for crude protein, crude fibre, fat, ADF, NDF, amino acids, fatty acids, minerals and vitamins. There were no statistically significant differences between corn event TUSC1 and CHP38/N46 control for crude fat, fibre, ash (grain only) ADF and NDF (forage only). Crude protein in corn event TUSC1 grain and forage was significantly higher than CH38/N46, but was within the reported literature ranges. Palmitic acid was significantly lower in corn event TUSC1 than the control, but levels were within literature ranges. Statistically significant differences were observed between corn event TUSC1 and the control for methionine, histidine, leucine and phenylalanine, however the levels were within the literature ranges. Statistically significant differences in Mg, Na, Zn, beta-carotene and Vitamin B1 were observed between corn event TUSC1 and the control, but the levels were comparable to literature values.
Phytic acid and trypsin inhibitor were analyzed in corn event TUSC1 grain and compared to CHP38/N46. No statistically significant differences were observed between TUSC1 and the control for phytic acid and trypsin inhibitor.
A pig study was conducted to evaluate improved grain digestibility and available energy of corn event TUSC1 diets compared to its isogenic control and other corn diets. Fecal and ileal digesta samples were collected to estimate digestibilities using chromic oxide as a marker. Ileal and whole tract digestibilities of nitrogen were statistically significantly higher for event TUSC1 than its control, while no differences were observed for digestibilities of fat, NDF, starch or energy. Digestible energy content was similar between event TUSC1 and its control. In-vitro dry matter disappearance of TUSC1 F2 corn was greater than isogenic control.
The evidence provided by Pioneer Hi-Bred supports the conclusion that the nutritional composition of TUSC1 corn is equivalent to commercial corn varieties.
2. Potential Impact on Livestock and Workers/By-standers
Corn event TUSC1 was not modified to express a new protein, or a modified form of an endogenous protein, but rather to reduce the expression of an endogenous protein. As such, this modification would not be expected to result in the expression of a novel allergen or toxin or the altered expression of endogenous toxins and allergens.
Based on the characterization data provided, including nutritional composition and agronomic data, it is unlikely that any unintended effects have occurred in the corn genome.
The evidence provided by Pioneer Hi-Bred supports the conclusion that corn event TUSC1 will not result in altered impacts on livestock and workers/by-standers compared to current commercial corn hybrids.
VI. New Information Requirements
If, at any time, Pioneer Hi-Bred International 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 corn event TUSC1, its descendants, or products derived there from, Pioneer Hi-Bred International must immediately provide such information to the CFIA. On the basis of such new information, the CFIA will re-evaluate the potential impact of event TUSC1 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 event TUSC1.
VII. Regulatory Decision
Based on the review of data and information submitted by Pioneer Hi-Bred International, and through comparisons of TUSC1 with unmodified corn counterparts, the Biotechnology Environmental Release Assessment Unit, CFIA, has concluded that the modified gene and its corresponding traits does not confer to corn event TUSC1 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 Pioneer Hi-Bred International, including comparisons of TUSC1 with unmodified corn counterparts, the Feed Section, CFIA, has concluded that the modified gene and its corresponding novel trait will not confer to this plant any characteristic that would raise any concerns regarding the safety or nutritional composition of corn event TUSC1. Grain corn, its byproducts and corn oil, are currently listed in Schedule IV of the Feeds Regulations and are, therefore approved for use in livestock feeds in Canada. Corn event TUSC1 has been assessed and found to be as safe as nutritious as traditional corn varieties. TUSC1 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 corn event TUSC1 is therefore authorized by the Plant Biosafety Office of the Plant Products Directorate and the Feed Section of the Animal Health and Production Division as of March 21, 2007. Any corn lines derived from TUSC1 may also be released into the environment and used as livestock feed, provided no inter-specific crosses are performed, provided the intended use(s) 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 corn, in terms of their potential environmental impact and livestock feed safety.
The corn event TUSC1 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 this corn event.
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