Chapter 2 - Data Requirements for Single Ingredient Approval and Feed Registration

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2.6 Guidelines for the Assessment of Novel Feeds: Plant Sources (formerly DIR 95-03)

Table of Contents

• 1.0 Introduction
• 2.0 Characterization of the Plant Source
• 3.0 Nutritional Data
• 4.0 Dietary Exposure
• 5.0 Toxicology Data
• 6.0 Allergenicity Data
• 7.0 Laboratory Animal/Livestock Feeding Trials
• 8.0 Evaluation of Environmental Safety
• 9.0 Method of Detection and Identification Requirements
• 10.0 Regulatory Decision
• 11.0 Posting of Decision
• 12.0 New Information Requirements
• 13.0 Definitions
• 14.0 References

1.0 Introduction

1.1 Scope

These guidelines (Section 2.6 of Regulatory Guidance: Feed Registration Procedures and Labelling Standards) apply to novel feeds from plants, or parts or products thereof, which have not previously been used as livestock feed in Canada, i.e. not listed in Schedules IV or V of the Feeds Regulations, and/or have a novel trait making them not substantially equivalent to an approved product already on the market in Canada.

Prior to the release of a novel feed (i.e., the manufacture, sale, import or use as livestock feed) authorization from the Animal Feed Division, CFIA, is required.

These guidelines are a guide for the preparation of an application for the authorization of the release of a novel feed. It includes criteria that will be considered in the assessment of safety and efficacy of a novel feed. This directive is not intended to explicitly define all of the data that could be required in the course of the assessment.

More information on novel feeds, application procedures and Animal Feed Division activities is available on the CFIA Web Site.

These guidelines have been prepared to provide guidance regarding the submission of an application for the authorization of the release of a novel feed, as may be required under the Feeds Regulations. Please be aware that the information provided in these guidelines is not exhaustive and will be updated as appropriate to reflect current scientific knowledge. For further clarification, applicants are strongly recommended to consult with the CFIA's Animal Feed Division. For all purposes of interpreting and applying the law, applicants are invited to consult the official versions of the relevant Acts and Regulations.

1.2 Legal Authorities

The manufacture, sale and import of livestock feeds are regulated in Canada under the Feeds Act and Regulations. All feeds must be safe, to livestock; to humans (via the potential transfer of residues into human food, i.e., meat, milk and eggs, and via worker/bystander exposure); and to the environment. Feeds must also be shown to be effective for their intended purpose. Approved feed ingredients are listed and defined in Schedules IV and V of the Feeds Regulations, with appropriate guarantees, standards, and label requirements. All imported feeds must meet the same standards as domestic feeds.

The legislative authorities under which the CFIA regulates novel feeds are as follows:

For the regulation of novel feeds:

The Feeds Act, R.S. 1985 chapter F-9
The Feeds Regulations, 1983 F-9 SOR/83-593

For the collection of fees:

Canadian Food Inspection Agency Fees Notice, Canada Gazette, Part I (05/13/2000)

1.3 Regulation of Novel Feeds

The CFIA evaluates and regulates all feed ingredients, including novel feeds, in the same manner. Any feed ingredient that is new (i.e., is not already listed in Schedules IV or V of the Feeds Regulations), or has been modified such that it differs significantly from a conventional ingredient, is required to undergo a pre-market assessment and approval. The purpose of all feed assessments is the same: to ensure that the feed ingredient is safe (in terms of animal health, human health via residues in food and worker/by-stander exposure, and the environment) and effective for its intended purpose prior to marketing. The evaluation also ensures that the feed is accurately defined in the Feeds Regulations and is labelled appropriately for its safe, effective use and for consumer protection.

Novel feeds are considered to be those derived from an organism or organisms, or parts or products thereof that:

1. are not approved as livestock feed in Canada (i.e. are not listed in Schedule IV or V of the Feeds Regulations), and/or
2. contain a novel trait.

Schedules IV and V of the Feeds Regulations list feed ingredients approved for use in livestock feed in Canada. Schedule IV comprises a range of ingredients such as forages and roughages, energy feeds, protein sources, vitamins, minerals, fermentation products and other products, while Schedule V is restricted to flavouring ingredients.

A novel trait is a heritable characteristic of a feed that is not substantially equivalent in terms of its specific use and safety to a characteristic of a similar feed that is set out in Schedule IV or V of the Feeds Regulations. In other words, for novel feeds derived from plant sources, a novel trait is a heritable characteristic that is new to a plant species, or is an endogenous trait that has been modified such that it differs from conventional parameters for that plant species.

Note that for feeds with novel traits, it is the presence of the novel trait that triggers regulatory oversight under the Feeds Regulations, and not the method used to introduce the trait; hence the plant, rather than the process, is subject to regulatory oversight. Novel feeds may be created by such methods as traditional breeding, mutagenesis, cell fusion, or recombinant DNA techniques.

It is also important to note that the trigger for regulatory oversight is novelty, not risk. Any potential risk, in terms of safety and efficacy, is determined during the assessment of the product.

Proponents seeking guidance with respect to determining whether or not their product is a novel feed can refer to Appendix II of these guidelines or consult directly with the Animal Feed Division. Consultation with the Animal Feed Division is highly recommended for any proponents experiencing difficulty in determining novelty.

1.4 Assessment Principles

Feed assessments are conducted to ensure that feed ingredients are safe (in terms of animal health, human health via residues in food and worker/by-stander exposure, and the environment) and effective for their intended purposes prior to marketing. The Animal Feed Division conducts livestock feed assessments based on several internationally recognized principles.

1. Assessments are conducted on a case-by-case basis.

   Novel feeds can vary greatly in terms of their individual characteristics. Thus the prescription of one set of data requirements to demonstrate efficacy and safety for all assessments would be impractical and onerous for proponents. Data requirements are, instead, determined on the basis of the characteristics of the product in question. This allows proponents to concentrate on meeting those data requirements relevant for their particular product.

2. Familiarity

   The degree of scientific support required in a data package is based on the complexity of the product in question and the degree of familiarity associated with the product/trait. If the Animal Feed Division has a high degree of familiarity with respect to the characteristics of a particular product and/or trait, in terms of livestock feed use, the level of data required to demonstrate safety and efficacy may be reduced or more narrowly defined.

3. Comparative Approach

   Novel feed assessments are done using a comparative approach. A comparison of a novel feed against products with a history of use as livestock feed highlights the similarities and differences that the novel feed has when compared to feeds already present in the marketplace.

   Comparisons of data made within the submitted data package should make use of an appropriate counterpart or counterparts. Appropriate counterparts have a history of livestock feed use within Canada. For feeds with novel traits, proponents should typically make comparisons using the unmodified counterpart. In other words, proponents should use conventional parental varieties and/or, in some cases for plants derived using recombinant DNA techniques, negative segregants when making comparisons.

4. Valid Scientific Rationale

   Provision of a valid scientific rationale (supported by appropriate data or literature references) may negate requirements for certain studies detailed in these guidelines or allow bridging to other valid scientific studies.

5. Weight of Evidence

   A weight of evidence approach is used when performing assessments. This means that the sum of the overall data submitted provides the context for determining whether or not a novel feed should be authorized.

6. Feed Definitions & Labelling

   A component of the approval process for novel feeds is to ensure that they are accurately defined within the Feeds Regulations and are labelled appropriately for safe, effective use and for consumer protection. In some instances a novel feed may be considered as meeting an existing definition, while in other instances an existing definition may need to be modified, or a new definition may be created.

   Additional principles applying to feeds with novel traits:

7. Substantial Equivalence

   Substantial equivalence is based on the comparison of properties between the modified plant from which a feed with a novel trait is derived and an appropriate counterpart. Taking into consideration both intended and unintended effects, similarities and differences between the modified plant and its counterpart can be identified. Those aspects in the modified plant which are determined to be equivalent to the counterpart are accepted and the assessment then focuses on the differences. The concept of substantial equivalence is endorsed by international groups such as the OECD and FAO/WHO.

   In the feed assessment, molecular, nutritional and toxicological data for the novel feed are compared to those of an appropriate counterpart. The identification and subsequent determination of levels of key nutrients, antinutrients and endogenous toxins and allergens is an important aspect of the determination of substantial equivalence. The OECD Consensus documents on individual crops may be of assistance as a reference to aid in the determination of the key elements to be examined.

8. Unintended Effects

   The assessment process considers the likelihood that unintended effects may be present in the modified plant and that these unintended effects may impact efficacy and/or safety of feed derived from the plant.

1.5 Determining Data Requirements

Section 2-8 of these guidelines outline the categories of data requirements that proponents should consider when compiling a novel feed submission. Proponents should address each section, but the manner in which the requirements can be satisfied will vary. In some instances collection and analysis of raw data is necessary, while in other instances providing a valid scientific rationale supported by existing scientific publications is sufficient.

Applicants are encouraged to consult with the Animal Feed Division in the early stages of the development of a novel feed for clarification on the specific information that is required to conduct a safety and efficacy assessment. Please note that additional regulatory requirements exist if a proponent wishes to make claims associated with their feed once it is authorized. Guidance for feed claims can be found in Chapter 2, Section 2.2, of Regulatory Guidance: Feed Registration Procedures and Labelling Standards.

1.6 Data Quality

Data provided as part of an application must be of a standard that would be acceptable for publication in a peer-reviewed scientific journal. Studies must be designed and conducted in accordance with sound scientific concepts and principles and, where appropriate, Good Laboratory Practices. All test substances used in the generation of data should be representative of the final feed form. Controls should also be appropriate to meet the objective(s) of the study. Studies must be carried out by qualified research personnel using validated methods. Complete descriptions of the methods used to generate submitted data must be provided, including the sensitivity and quality control/quality assurance procedures. When applicable, data must be analyzed using appropriate statistical analysis techniques. Primary data must be made available upon request. All parts of the application must be fully legible, with particular attention paid to analytical gels and blots (e.g., SDS-PAGE gels, Northern blots, Southern blots, and Western blots). Any scientific references used to support the data package should be provided with the application.

A reviewer’s checklist for analytical techniques, developed in collaboration with Health Canada and the United States Department of Agriculture’s Animal and Plant Health Inspection Services, providing guidance on the preparation of quality data.

1.7 Presentation of Information

Proponents should organize their submissions in a manner similar to that of a scientific paper suitable for publication. A typical submission format will consist of:

• An overall introduction for the submission
• Per data element:
  • An introduction
  • Description of data collection (note that raw data is typically required)
  • Results
  • Conclusions
• An overall conclusion for the submission

Note that within the concluding statements proponents should clearly indicate how the results relate to product safety and/or efficacy with respect to use as livestock feed.

1.8 Procedures for Submitting an Application

Applicants are strongly encouraged to apply for authorization of the release of the novel feed well in advance of the anticipated time of commercialization. Applicants are required to follow the procedures for submitting an application as outlined in Chapter 1 of Regulatory Guidance: Feed Registration Procedures and Labelling Standards.

Please note that these guidelines are not intended to explicitly define all of the data that might be required in the course of the assessment. Additional information may be requested by the Animal Feed Division on a case by case basis. Applicants are encouraged to consult with the Animal Feed Division in the early stages of the development of a novel feed for clarification on the specific information that is required by the Animal Feed Division to conduct an assessment.

Applicants must submit a hard copy version of the application and are encouraged to also submit an electronic (compact disc) copy of the application.

1.9 Harmonization With Other Regulatory Parties

Where a novel feed is derived from a plant that could reasonably be expected to be released into the Canadian environment and/or used as human food, the Plant Biosafety Office, CFIA, and/or Novel Foods Section, Health Canada may also require applications for authorization of the Plant with a Novel Trait (PNT) and/or novel food, respectively. It is the responsibility of the applicant, and not of the Animal Feed Division, to contact these offices and determine if an application is required.

Applicants seeking authorization for the release of a novel feed in Canada are encouraged to seek authorization for their product in the United States and Canada simultaneously, if the product will be regulated in the United States. Obtaining such authorizations should minimize the movement of unauthorized material across the border.

When a novel feed is derived from a plant that is considered to also be a PNT and/or a source of a novel food, authorizations for use as a novel feed, novel food and environmental release will be granted in a harmonized fashion. This will minimize the potential for unapproved products to enter the Canadian environment or food or feed supplies. Proponents and importers should be aware that, where applicable, approvals are required from all three regulatory parties prior to environmental release, food use or feed use. Note that harmonization of authorizations is only required when a product triggers more than one type of assessment. In other words, novel feeds that are not novel foods or PNTs do not require harmonized authorizations.

Where plant-derived products are intended for exclusive use as food, or feed or for plant molecular farming, consultations among regulatory authorities will still be required to assess any potential risks, and management thereof, associated with release of the product into a commodity stream for which it is not intended.

Please note that once the assessments have been completed, the applicant is notified in writing by the CFIA and Health Canada (in separate letters) regarding their respective decisions regarding the application.

1.10 Confidential Information

Information provided to the CFIA by applicants for the purposes of obtaining authorization for the release of novel feeds can only be accessed by authorized personnel and is protected under the federal Access to Information Act, Section 20.

All third party requests for information are subject to the federal Access to Information and Privacy (ATIP) Acts. For further information, please consult with CFIA’s ATIP service, at 613-225-2342.

2.0 Characterization of the Plant Source

Sufficient data must be submitted to characterize the novel feed, and the plant from which a novel feed is derived, to permit comparison with appropriate counterparts.

Most of the questions regarding product characterization can be addressed by data generated in the developmental stage.

2.1 Source, Host and Donor Organisms

For feed derived from a plant source with no history of use as feed, characterization of the plant source is essential to assess the efficacy and safety of the novel feed.

For feeds with novel traits, information regarding the host and any donor organisms used in the development of a modified plant can, in part, lead to a better understanding of what needs to be assessed. In some cases, characterizing these organisms is quite straightforward, e.g. for those that are conventional crop species. Conversely, donor organisms that have no history of use as livestock feed, or those known to contain toxins, allergens or antinutrients, will require a more detailed characterization.

Required information for applicable source, host and donor organisms includes, but is not restricted to, the following:

1. Common or usual names; scientific name and taxonomic classification.
2. A critical assessment of the ability of the organism to produce/contain potentially toxic compounds (e.g., heavy metals, pesticides), anti-nutritional compounds and endogenous allergens.
3. Available toxicology data for the organism.
4. Information on history of use of the organism, or any related organisms, in livestock feed.

2.2 Plant Characterization

Information regarding the characteristics and use of a plant as a feed source are core components of an application.

Provide the following information:

1. All designations for the plant used in the application.
2. A description of the intended feed use of the plant.
3. List of other government agencies, in Canada or abroad, which were notified of the development of the novel feed, and description of the purpose of the notification; and the countries in which the novel feed has received regulatory approval.

   For a feed with a novel trait also provide:

4. A description of the novel trait.
5. A description of the intended purpose of the novel trait.
6. A description of how the proponent analyzed the host plant to determine if any anticipated secondary effects or unintended effects were present.

   This assessment takes into consideration the possible presence of unintended effects in the modified plant; whether present as a result of the modification process or due to the modification itself. Overall phenotypic characterization of the modified plant can be used as evidence to show a lack of unintended effects or to identify any such effects.

   There are two ways in which plants are often characterized:

   1. compositional profile comparisons with an appropriate counterpart, and
   2. agronomic/phenotypic parameter comparisons with an appropriate counterpart.

   Note that proponents are not required to provide the Animal Feed Division with agronomic/phenotypic data if this information has already been provided to the CFIA as part of a PNT assessment.

   To satisfy this requirement, proponents may refer to data they have provided to address Section 3 of these guidelines (compositional data) and data they have provided as part of the evaluation of environmental safety of the modified plant.

7. A characterization and evaluation of any secondary effects and unintended effects that are identified in the host plant (in terms of the safety and efficacy of the derived feed).

2.3 Modification/Development History

For feeds with novel traits, provide the following information:

1. Describe the method of trait introduction/selection.

   Different methods of trait introduction have different characteristics associated with them with respect to both intended and unintended effects. Knowledge of the method used to introduce the modification can, in part, lead to a better understanding of what needs to be assessed in the modified plant.

2. An outline of the pedigree of the modified plant. Please indicate which generations were used to produce supporting data.
3. Describe the selection criteria that were employed at both the initial stages of development and during the subsequent breeding process.
4. Evidence to support the inheritance and stability of each genetic modification in the plant.
   • For plants which are either male- or female-fertile, or both, data that demonstrates the pattern and stability of inheritance and the expression of the new traits is required. If the new trait cannot be directly measured by an assay, it may be necessary to examine a) the inheritance of the DNA insert directly, and b) RNA expression.
   • For plants which are either infertile, or for which it is difficult to produce seed (such as vegetatively propagated male-sterile potatoes), data to demonstrate that the trait is stably maintained and expressed during vegetative propagation over a number of cycles appropriate to the crop are required.
5. In the case of an allopolyploid plant, name the parental genome in which the genetic modification occurred.

2.4 The Novel Trait(s)

For feeds with novel traits, provide the following information in regards to the novel trait and any associated newly expressed material that is either exogenous or modified native material:

1. A description of the history of use of the novel trait.
2. Characterize how the novel trait operates.

   Information regarding how a novel trait operates can, in part, lead to a better understanding of what needs to be assessed. Define the genetic basis of the trait (e.g. the inserted gene, mutation) and describe the downstream effect(s) of this genetic basis that result in the novel trait. For products of rDNA, consider all genetic material potentially transferred to plant host genome and characterize DNA shown to be transferred (see Appendix I).

3. Detailed information on the gene product(s) including their activity, breakdown products, byproducts and their affected metabolic pathways (including altered accumulation and storage patterns), resulting from the novel trait.
4. Indicate if gene expression is constitutive or inducible, with a description of the inducing agents. Provide information demonstrating that the trait is expressed in the appropriate tissues in a manner and at levels that are consistent with the associated regulatory sequences driving the expression.
5. Characterize the expression level of the gene product in all plant parts that may be fed to livestock. A determination of the levels of the metabolites of the gene product may be required.
   • If the protein concentration is below the limit of detection, mRNA data may be substituted.
   • For virus resistant plants where the transgenes are derived from a viral genome, transgene RNA levels in tissues consistent with the associated regulatory regions driving expression of the transgene are required in addition to transgene protein analysis.
   • For plants modified to specifically alter the accumulation of a specific mRNA or protein, data on the level of the target protein only may be provided. If the target protein levels are below levels of detection, mRNA levels may be provided.
6. The likelihood of exposure to livestock, humans and the environment, including the estimated level and most likely route of exposure to the novel products, breakdown products, and by products.
7. The potential for adverse health effects to humans and livestock, including exposure to toxins and anti-nutritional factors, as well as irritation and allergenicity to humans (in terms of bystander/occupational exposure).
8. The potential of carry over of any novel product to manure and the potential impact of this carry over.

2.5 Marker Genes

Any transformation markers used in the development of a plant must be identified and characterized as per the novel trait (Section 2.4 of these guidelines). Any secondary effects of these markers on the biochemistry, physiology and secondary metabolism of the host plant, must also be characterized and discussed with respect to consequences regarding efficacy and safety.

Because antimicrobials are routinely mixed with livestock feed, a determination of the potential for their inactivation during storage as a result of the activity of any expressed antimicrobial resistance marker gene should be conducted.

The use of marker genes that confer resistance to clinically important antimicrobials should be avoided.

3.0 Nutritional Data

Novel feed ingredients derived from the plant should be described, including information on processing and estimated feeding rates of the ingredient and by-products.

Nutrient and antinutrient composition of the major plant components and byproducts used in livestock feed must be considered. For example, composition of whole plant corn or corn silage in addition to grain should be determined. Consideration must also be given to by-product streams in which nutrients or antinutrients are concentrated.

A comparison of the composition of the novel feed ingredient to that of the same feed ingredient derived from an appropriate counterpart must be undertaken. These comparisons should be based on analysis of representative samples derived from appropriate growing areas over one or more growing season(s). Comparison to literature ranges for nutritional composition of nearest genetic equivalents or counterparts may be an acceptable alternative. Data should demonstrate the uniformity or variability of the composition of the final product and include the analysis/characterization of the gene products or any metabolites that may have been altered (e.g., in the case of proteins, post translational modifications are of interest).

Scientific investigations presented in support of nutritional evaluation must be: designed to facilitate statistical analysis; analyzed by appropriate statistical methods; and, for those feeds which are able to grow in Canada, conducted under growing/environmental conditions similar to those expected to occur in Canada. As a minimum, the experimental design should include three locations, two treatments (novel feed vs. control), and three replicates. Representative samples should not be pooled for analysis. Nutrient and antinutrient analysis should be done in duplicate, according to accepted AOAC International or other recognized methods.

The raw data and printouts of statistical analysis must accompany the application. Statistical differences in composition should be addressed in terms of biological significance.

3.1 Nutrient Composition

Provide, at a minimum:

1. Crude protein
2. Crude fat and ash
3. Crude fibre or acid detergent fibre (ADF) and neutral detergent fibre (NDF).
4. Levels of naturally occurring or adventitious antinutritional factors which could reasonably be expected to be present (e.g., phytates, trypsin inhibitors, alkaloids, pigments, etc.)

   Based on the preceding evaluation and the intended use of the feed, the following information may also be required:

5. True protein, non-protein nitrogenous material, and amino acids
6. Quantitative and qualitative composition of total lipids, i.e., saponifiable and non-saponifiable components, and fatty acids
7. Composition of the carbohydrate fraction, e.g., sugars, tannins, non-starch polysaccharides and lignins
8. Vitamins
9. Minerals

For feeds with novel traits, the OECD consensus documents, which detail the key nutrients and antinutrients to be analysed in various crop species, should be consulted for guidance.

Unusual or unanticipated results in nutrient or antinutrient analysis must be investigated further.

In the case of feeds with novel traits which have been intentionally modified in terms of nutritional composition (e.g., altered amino acid or fatty acid profile) or functionality (e.g., digestibility, bioavailability), nutrient and antinutrient analysis and comparison with the appropriate controls should be conducted as described above. Data to support any claims associated with the modification must also be provided. A discussion of the potential consequences to safety and efficacy of the intentional nutritional modification should be provided.

4.0 Dietary Exposure

Complete details should be supplied on the predicted amount of the novel feed in the complete livestock diet. Estimates of dietary exposure to the novel feed may play a key role in determining the data required for a safety and efficacy assessment. In addition, data specific to animal age and species with respect to the intended use of the novel feed may be necessary. Feeds that are present at low levels in a complete diet may be of less concern than those intended to be significant components of the diet. Proponents should also discuss any impact the novel feed may have on conventional feed ration formulation.

For feeds with novel traits, the predicted amount of the feed in the complete livestock diet is typically based on the traditional use of the unmodified counterpart. Estimates of dietary exposure should be conservative in that they should consider situations whereby 100% replacement of the unmodified counterpart occurs.

If a feed with a novel trait will, or should, be used in a way that will result in dietary exposure that is different from that of the unmodified counterpart, this should be noted and the significance of the changed pattern of use discussed.

If a feed with a novel trait has intentional changes in a compositional value, discuss how this change will affect animal health and production.

5.0 Toxicology Data

The toxicity of the host and donor organisms and/or the introduced novel traits to livestock through feed intake and humans through ingestion of livestock-derived food products (i.e., meat, milk, and eggs) and bystander/occupational exposure should be considered.

5.1 Endogenous Toxins

1. For novel feeds that are new feed ingredients, provide data on the level of endogenous toxins known to be present in the plant from which the novel feed is derived and compare these with levels found in approved feed ingredients.

   For feeds with novel traits, provide a comparison of the levels of endogenous toxins known to be present in host or donor organisms between the modified plant and the unmodified counterpart.

5.2 Newly Expressed Material

Provide the following information for feeds with novel traits in regards to newly expressed material, which is either introduced exogenous material or modified native material:

1. A comparison of the amino acid sequence of the newly expressed protein to amino acid sequences of known toxins to determine any potential homology. Specify the database(s) used to carry out the search.
2. Description of any similarity of newly expressed material resulting from the modification to products having a history of use.
3. The digestive stability of newly expressed protein(s).
4. The heat stability of newly expressed protein(s).
5. If concerns arise from assessment of the data submitted, additional toxicity studies may be required.

If bacterially produced protein is used in place of plant produced protein in studies of the potential toxicity of the novel protein, biochemical, structural and functional equivalency of the two proteins must be demonstrated.

Please refer to Chapter 2, Sections 2.3 to 2.5, of Regulatory Guidance: Feed Registration Procedures and Labelling Standards for further information on traditional toxicity study requirements for feeds.

6.0 Allergenicity Data

The allergenicity potential of the host and donor organisms and/or the introduced novel traits to humans through ingestion of livestock-derived food products (i.e., meat, milk, and eggs) and bystander/occupational exposure should be considered.

6.1 Endogenous Allergens

1. For novel feeds that are new feed ingredients, provide data on the level of allergens known to be present in the plant from which the novel feed is derived and compare these with levels found in approved feed ingredients.

   For feeds with novel traits, provide a comparison of the levels of endogenous allergens known to be present in host, parental lines, or donor organisms between the modified plant and an appropriate counterpart.

6.2 Newly Expressed Proteins

At present there is no definitive test that can be relied upon to predict allergic response to a newly expressed protein. A case by case, step-wise approach (such as the FAO/WHO 2000 or 2001 decision tree) should be applied in the assessment of possible allergenicity of newly expressed proteins.

Provide the following information for feeds with novel traits in regards to newly expressed proteins which are either introduced exogenous material or modified native material:

1. A comparison of the amino acid sequence of the newly expressed protein to amino acid sequences of known allergens. The size of the contiguous amino acid search should be based on scientifically justified rationale to minimize the potential for false negative or false positive results. Validated search and evaluation procedures should be used. Specify the database(s) used to carry out the search.
2. If the novel protein is derived from a known allergenic source or it has significant sequence homology with a known allergen, the reactivity of the novel protein with immunoglobulin-E (IgE) from the blood serum of appropriate allergic individuals should be evaluated.
3. The digestive stability of newly expressed protein(s).
4. The heat stability of newly expressed protein(s).

7.0 Laboratory Animal/Livestock Feeding Trials

Laboratory animal and/or livestock feeding trials may be necessary to address safety and efficacy assessment requirements for plant-derived novel feeds. These studies should be conducted using materials representative of the material that will be used in Canada. In some cases, conducting whole plant feeding trials for the safety assessment may be impractical due to variations in composition and difficulty in feeding sufficient quantities of the substance in question. As such, every effort should be made to fully characterize the plant, plant parts, or in vitro produced proteins with respect to appropriate counterparts and equivalent comparators.

Laboratory animal and/or livestock feeding trials in support of the safety of the plant derived novel feed may be required to address any toxicological concerns arising from an assessment of data submitted under Sections 2.1 to 6.0 of these guidelines, including acute toxicity data.

Feeding trials may also be needed to address issues related to efficacy of the plant derived novel feed such as evidence of the nutritional adequacy and nutrient bioavailability. The necessity of these studies would be determined based on an assessment of data submitted under Sections 2.1 to 6.0 of these guidelines.

In the case of intentional modification of the nutritional composition or functionality, in vitro and/or in vivo studies to support the safety and efficacy of the feed are required.

Authorization of the release of a novel feed for research purposes must be obtained from the Animal Feed Division prior to conducting livestock feeding trials with novel feeds in Canada. For guidance on applying for authorization of the release for research purposes, please refer to Chapter 5 of this document (Regulatory Guidance: Feed Registration Procedures and Labelling Standards).

8.0 Evaluation of Environmental Safety

An evaluation of the potential impact of the novel feed on the environment is required if any component of the novel feed is released, including excreta from the livestock consuming the feed.

This evaluation may also consider the following criteria, if not reviewed by the CFIA as part of a PNT assessment:

1. Potential of the plant to become a weed of agriculture or be invasive of natural habitats.
2. Potential for gene flow from the plant to wild relatives whose hybrid offspring may become more weedy or more invasive.
3. Potential of the plant to become a plant pest.
4. Potential impact of the plant or its gene products on non-target species, including humans.
5. Potential impact of the plant on biodiversity.

9.0 Method of Detection and Identification Requirements

A decision regarding authorization of the release of a feed with a novel trait will not be made until the following is submitted to the CFIA:

1. An appropriate method for the detection and identification of the feed with a novel trait.
2. Written agreement to provide the CFIA with reference material suitable to support the method.

Consultation with the CFIA is recommended prior to submission to determine appropriate detection and identification methodologies and reference material.

Guidance on what constitutes a suitable method is provided on the CFIA website.

10.0 Regulatory Decision

Based on the information received, and after evaluating the potential impact on livestock, humans and the environment, the Animal Feed Division will:

1. Authorize the release of the novel feed where there is minimal risk (note that conditions may be imposed on authorizations to manage risk);
2. Refuse to authorize the release of the novel feed where its use poses an unacceptable risk, including when there is insufficient data to demonstrate safety and/or efficacy; or
3. Follow up with the applicant for any additional information that the Animal Feed Division requires to complete the assessment (“deficiency letter”), followed by one of the decisions above. The Animal Feed Division will consider the application closed if no response to the deficiency letter is received within one year from the date of the deficiency letter.

Once a novel feed (including those with a novel trait) has received authorization (i.e., it gets listed in Schedule IV or V or is determined to be substantially equivalent to an ingredient already listed in the Schedules), it is no longer considered to be novel.

11.0 Posting of Decision

After a regulatory decision has been made to authorize a novel feed that is a new feed ingredient, a definition for the ingredient will be added to Schedule IV or V of the Feeds Regulations.

After a regulatory decision has been made concerning a feed with a novel trait that is not a new feed ingredient, the Animal Feed Division posts a decision document, which is a record of the assessment, on the CFIA website. The decision document explains the regulatory decision reached by the Animal Feed Division, and the Plant Biosafety Office (PBO) in cases where the novel feed is from a plant source that is also a PNT, following their evaluations.

12.0 New Information Requirements

If at any time the proponent becomes aware of any new information regarding risk to the environment, livestock or human health that could result from the release of the novel feed the proponent must immediately provide such information to the CFIA.

On the basis of new information, regardless of source, the CFIA will re-evaluate the potential impact on and risk to the environment, including the potential impact on and risk to human or animal health with respect to the livestock feed use. The CFIA may impose additional conditions on the release or use as livestock feed, may change the conditions on these authorizations, or cancel the authorizations and require the proponent to stop the release or use as livestock feed and take any appropriate action necessary to eliminate or minimize the risk.

13.0 Definitions

Biotechnology: The application of science and engineering in the direct or indirect use of living organisms or parts or products of living organisms in their natural or modified forms (Canadian Environmental Protection Act).

Carrier DNA: DNA used to expedite the preparation or the transformation of genetic material into a plant but which is itself not part of the construct.

Coding region: A DNA sequence which can be translated to produce a protein.

Construct: An engineered DNA fragment (e.g., plasmid) which contains, but is not limited to, the DNA sequences to be integrated into a target plant's genome.

Counterpart: A feed or feeds selected by the applicant, to which the novel feed will be compared. The chosen counterpart should be, if possible, the closest genetic equivalent to the novel feed and may be a previously assessed and authorized novel feed. Since there will be a range of characteristics among varieties within a species, comparison with several counterparts may show that the novel feed has characteristics within the normal range exhibited by that species. If the closest genetic equivalent to the novel feed is not similar to the novel feed (e.g., in the case of a plant intentionally modified for composition), consideration may also be given to comparison with other products the novel feed was designed to emulate.

Database Citations: Publicly available sources of nucleotide or protein sequence information. Five commonly used databases and their website addresses are found in the references section.

Environment: The components of the earth (including air, land and water, all layers of the atmosphere, all organic and inorganic matter and living organisms) and all interacting natural systems that include components referred to above (Feeds Regulations). This includes natural and managed ecosystems which include agricultural ecosystems.

Familiarity: The knowledge of the characteristics of a plant species and experience with that plant species in Canada.

Feed: Any substance or mixture of substances containing amino acids, anti-oxidants, carbohydrates, condiments, enzymes, fats, minerals, non-protein nitrogen products, proteins or vitamins, or pelletizing, colouring, foaming or flavouring agents and any other substance manufactured, sold or represented for use:

1. for consumption by livestock,
2. for providing the nutritional requirements of livestock, or
3. for the purpose of preventing or correcting nutritional disorders of livestock, or any such substance for use in any such substance or mixture of substances (Feeds Act).

Gene flow: The transfer of genetic material by interbreeding from one population of a species to another population (same or related species), thereby changing the composition of the gene pool of the receiving population.

Genetic Modification: Any intentional change to the heritable traits of an organism. This includes, but is not limited to: traditional breeding, recombinant nucleic acid techniques, somaclonal variation, electroporation, mutagenesis, and similar techniques.

Insert: That part of the construct which is integrated into the recipient plant's genome.

Irritant: Any agent capable of eliciting an abnormally excited or sensitive condition in a body part of a human or other animal.

Livestock: Means horses, cattle, sheep, goats, swine, foxes, mink, fish, rabbits and poultry and includes such other creatures as may be designated by regulation as livestock for the purposes of the Feeds Act.

Noncoding region: DNA sequences which lie outside of an open reading frame and which are not translated to become part of a protein. These might include scaffold attachment regions, promoters, leader sequences, enhancers, introns, terminators, and any other sequences that are used for gene expression either in the plant or other hosts, such as origins of replication, transposon elements, T-DNA borders, lox sequences, etc.

Novel Feed: Means a feed comprising an organism or organisms, or parts or products thereof, that:

1. is not set out in Schedule IV or V, or
2. has a novel trait (Feeds Regulations).

Novel Trait: Means a characteristic of the feed that:

1. has been intentionally selected, created or introduced into the feed through a specific genetic change, and
2. based on valid scientific rationale, is not substantially equivalent in terms of its specific use and safety both for the environment and for human and animal health, to any characteristic of a similar feed that is set out in Schedule IV or V (Feeds Regulations).

Plant with Novel Traits (PNT): A plant containing a trait not present in plants of the same species already existing as stable, cultivated populations in Canada, or containing a trait present at a level significantly outside the range of that trait in stable, cultivated populations of that plant species in Canada.

Safety Assessment: Encompasses hazard identification, risk estimation, and risk evaluation and management.

Stability: The ability of the trait to be expressed in the modified plant line and plant lines derived there from in a consistent, reliable, and predictable manner.

Substantial Equivalence: Equivalence of the feed from a plant, within a particular plant species, in terms of its specific use and safety to the environment and human and animal health, to those in that same species, that are in use and generally considered as safe in Canada, based on valid scientific rationale.

Trait: The phenotypic characteristic(s) conferred to the recipient plant by specific genetic changes.

Transgenic Plant: A plant in which one or more genes, genetic constructs, or traits have been introduced using recombinant DNA techniques, which could be considered to include the insertion of genetic material from the same or different species.

Unintended Effects: In addition to intended changes caused by the insertion/mutation of defined DNA sequences during preparation of a genetically modified plant, additional unintended traits may be acquired or existing traits could be lost or modified. These unintended effects may be deleterious, beneficial or neutral with respect to health of plant and safety to livestock and humans.

Vector: An autonomously replicating DNA molecule into which foreign DNA is inserted and then propagated in a host cell.

14.0 References

GenBank: An annotated collection of all publicly available DNA sequences maintained by the National Institute of Health (NIH).

DNA Data Bank of Japan: The officially certified DNA bank of Japan, which collects DNA sequences from researchers.

EMBL Nucleotide Sequence: A database of DNA and RNA sequences collected from the scientific literature, patent applications, and directly submitted from researchers and sequencing groups.

FAO: The Food and Agriculture Organization of the United Nations leads international efforts to defeat hunger. The FAO acts as a neutral forum where nations meet as equals to negotiate agreements and debate policy. The FAO is also a source of knowledge and information.

FARRP Allergen Database: A database containing a list of unique known and putative allergens that were identified by searching publicly available protein databases.

OECD: The Organisation for Economic Co-operation and Development is an international organisation that accepts the principles of representative democracy and free-market economy. The OECD publishes reports that contain technical information for use in the regulatory assessment of products of biotechnology. These documents are mutually recognized among member countries of the OECD.

SWISS-PROT Protein Sequence Data Bank: A database of protein sequences produced collaboratively by Amos Bairoch (University of Geneva) and the EBI.

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Appendix I: Molecular Characterization Requirements for Feeds with Novel Traits Derived from Plants Developed using Recombinant DNA Techniques

For plants developed using recombinant DNA techniques, please provide the following:

1. For each event derived through transgenic methods, a unique identifier for each line must be designated according to the "OECD Guidance for the Designation of a Unique Identifier for Transgenic Plants".
2. A description of the genetic element responsible for the novel trait.
3. State the objective of the modification, e.g., novel herbicide tolerance, male sterility/restoration, etc.
4. A description of, and references for, the method used to introduce the novel trait(s).
   1. Information on the nature and source of the DNA used to modify the plant (e.g., carrier DNA, vectors, helper plasmids).
   2. For Agrobacterium-mediated transformation, the strain designation of the Agrobacterium used during the transformation process, and the method by which the Ti plasmid based vector was disarmed, and whether Agrobacterium was cleared from the transformed tissue.
   3. For transformation systems other than Agrobacterium:
   • Whether the system utilizes a pathogenic organism or nucleic acid sequences from a pathogen, and, if so;
   • how any pathogenesis-related sequences were removed prior to transformation.
5. A description of the genetic material potentially delivered to the host plant.
   1. A summary of all genetic elements which comprise the vector including coding regions, and non-coding sequences of known function. For each genetic element a citation where these functional sequences were described, isolated, and characterized (publicly available database citations are acceptable), indicate:
      • The size and identity of the genetic element;
      • The location and orientation in the vector;
      • The function in the plant;
      • The source (scientific and common name of the donor organism);
      • Whether the genetic element is responsible for disease or injury to plants or other organisms, or if its products are known toxicants, allergens, pathogenicity factors, or irritants.
   2. If there has been a significant modification that affects the amino acid sequence of the protein to be expressed in the plant, the specific amino acid changes should be identified. Indicate whether the modifications are known or expected to result in changes to post-translational modification or affect sites critical to the structure or function of the gene product.
   3. A map of the vector with the location of sequences described above that is sufficient to be used in the analysis of data supporting the characterization of the DNA, including, as appropriate, the location of restriction sites and regions used as probes and/or primers used for polymerase chain reaction (PCR).
6. A characterization of the DNA inserted in the plant, including:
   1. Number of insertion sites;
   2. Data that demonstrate if complete or partial copies are inserted into the plant's genome
   3. For noncoding regions associated with the expression of coding regions:
      • Whether or not plant promoters are inserted intact with the coding regions whose expression they are designed to regulate;
      • DNA analysis may be necessary for introns, leader sequences, terminators, and enhancers of plant-expressible cassettes;
      • DNA analysis may be necessary for promoters and other regulatory regions associated with bacteria-expressible cassettes
   4. For noncoding regions which have no known plant function and are not associated with expression of coding regions:
      • DNA analysis may be required for some sequences of known function (e.g., ori V and ori-322, T-DNA borders of Agrobacterium, and bacterial transposable elements)
      • DNA analysis is not required for any remaining sequences of the plasmid backbone when the plasmid is well characterized
   5. Where appropriate, sequence data of the inserted material and of the surrounding regions (sequencing information may be informative in some cases, i.e., to fully characterize a partial or rearranged DNA insert).

   Protein or RNA characterization may not be required for fragments of genetic constructs not expected to be functional in the plant (e.g., fragments of selectable marker genes driven by bacterial promoters).

7. Consider if a fusion protein could be produced if a fragment of a coding region designed to be expressed in the plant is detected, and in which tissues it may be located.

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Appendix II: Novelty Determination Guidance

It is the responsibility of the proponent to consider whether they have developed a novel feed or not. When proponents are determining whether a plant-derived feed is novel or not, there are some basic questions they can consider.

Q1. Is the plant-derived feed listed in Schedule IV or V of the Feeds Regulations?

NO, novel feed.

For feeds with novel traits:

Q2. Is the feed derived from a plant that has one or more characteristics that are new or outside of the conventional range for that plant species?

YES, novel feed

Proponents should use their expertise and relevant scientific literature to determine the range of a selected trait based on Canadian experience to determine if they have created a feed with a novel trait. A substantive change in agronomic, nutritional or compositional characteristic(s) of a plant that are new or outside the accepted range for a given species would constitute a novel trait, thus triggering the requirement to apply for authorization of the release of feed derived from the plant. For example, feed derived from a new wheat line with an increase in yield that is similar to increases seen historically in wheat lines cultivated in Canada would not be considered novel. Feed derived from a new insect-resistant canola line with resistance many fold higher than that displayed by plants currently grown in Canada would be a feed with a novel trait. Feed derived from a new soybean line with a substantive increase in methionine would be a novel trait.

Q3. Is the feed derived from a plant that is a descendant, produced through intraspecific crossing, of a plant previously authorized for livestock feed use?

YES, not novel

After a plant is authorized for use as livestock feed, all progeny of the modified plant and sister lines, which have been derived from the original modified plant, are also authorized for livestock feed use provided that the proponent has determined that:

• the intended uses are similar;
• Data that demonstrate if complete or partial copies are inserted into the plant's genome
• based on characterization, these plants do not display any additional novel traits and are substantially equivalent, in terms of their specific use and safety for the environment and for human and animal health, to plants currently in use as livestock feed; and
• the trait(s) previously assessed and authorized are expressed at a level similar to that of the authorized line.

The CFIA may ask the proponent to provide evidence supporting these conclusions.

Q4. Is the feed derived from a plant that is a descendant, produced through interspecific crossing, of a plant previously authorized for livestock feed use?

YES, likely novel

For feeds with novel traits, it is the presence of the novel trait that triggers regulatory oversight under the Feeds Regulations, not the method used to introduce the trait. As such, the transfer of a previously assessed and authorized trait into a recipient plant species through interspecific crossing can result in the creation of a novel feed. This means that the recipient species could require assessment and authorization prior to use as a livestock feed.

Subsequent to authorization by the Animal Feed Division for livestock feed use, the proponent may not be required to apply to the Animal Feed Division for authorization of additional lines derived through similar means (i.e. interspecific crossing), provided that the proponent has determined that:

• the intended uses are similar;
• based on characterization, these plants do not display any additional novel traits and are substantially equivalent, in terms of their specific use and safety for the environment and for human and animal health, to plants currently in use as livestock feed; and
• the traits previously assessed and authorized are expressed at a level similar to that of the authorized line.

The CFIA may ask the proponent to provide evidence supporting these conclusions.

Q5. Is the feed derived from a plant possessing previously authorized traits that have been intentionally stacked?

YES, notification of the Animal Feed Division required

The Animal Feed Division is to be notified at least 60 days prior to the intended livestock feed use of plants having stacked traits resulting from intentional intraspecific or inter-specific crosses between novel plants already authorized for livestock feed use. Following the proponent's notification, the Animal Feed Division will issue a letter (within 60 days of notification) informing the proponent of any concerns the Animal Feed Division may have regarding the livestock feed use of the plant with stacked traits. The Animal Feed Division reserves the right to request and review data to support the safe use of the modified plant in livestock feed. Stacking of traits with potential incompatible management requirements, possible negative synergistic effects, or where feed use of the plant is altered may result in the requirement for an assessment. Until all livestock feed concerns have been resolved, the modified plant must not be used as livestock feed.

Q6. Is the feed derived from a plant that is a re-transformation or re-mutation of a plant previously authorized for use as a livestock feed?

YES, may be novel

Re-transformation is considered to be the transformation of a plant with the identical construct(s) as a previously authorized plant of the same species and confers the same trait. Re-mutation is considered to be the mutation of the same gene in a plant as a previously authorized plant of the same species which confers the same trait. Such plants may not require an application to the Animal Feed Division, provided that:

• the method is identical to that used previously;
• the intended uses are similar;
• based on characterization, these plants do not display any additional novel traits and are substantially equivalent, in terms of their specific use and safety for the environment and for human and animal health, to plants currently in use as livestock feed; and
• the traits previously assessed and authorized are expressed at a level similar to that of the authorized line.

Note that if a proponent determines that a feed is not novel the information to support the determination must be retained and presented to the CFIA on request.

Consultation with the Animal Feed Division is highly recommended for any proponents experiencing difficulty in determining novelty.

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