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Interim Guidance on Honey Bee Data Requirements

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MEMORANDUM

DATE: October 19, 2011

SUBJECT: Interim Guidance on Honey Bee Data Requirements

FROM: Donald Brady, Director, Environmental Fate and Effects Division (7507P), Office of Pesticide Programs

TO: Environmental Fate and Effects Division (7507P), Office of Pesticide Programs

The following is intended to serve as interim guidance for ecological risk assessors (reviewers) in the Environmental Fate and Effects Division (EFED) in determining whether and what additional honey bee (Apis mellifera) studies should be considered toward characterizing the potential hazard, i.e., adverse effects, of chemicals to honey bees as part of registration review and registration of new chemicals/uses. An abbreviated outline of considerations can be found in Appendix 1 of this document.

At this time, the Environmental Fate and Effects Division (EFED) does not have a formal guidance for routinely quantifying risks to beneficial insects, particularly honey bees. A formal process for quantifying potential risks of pesticides to honey bees and non-Apis bees is evolving and will be informed by the recent Society of Environmental Toxicology and Chemistry (SETAC) global Pellston Conference on Pesticide Risk Assessment for Pollinators (held in January 2011). The Office of Pesticide Programs will consult with its FIFRA Scientific Advisory Panel (SAP) on the proposed process for estimating risks to honey bees and non-Apis bees in 2012 and will base its proposed process on the best available science. In developing its proposed process for quantifying risks to honey bees, EFED will include what it has learned through evaluating potential hazards to honey bees using both guideline and nonguideline studies.

Until a formal risk assessment process is established, this document provides a number of factors to consider when evaluating the need for additional data on honey bees. These considerations include:

Existing 40 CFR Part 158 Guidance

Typically, potential hazards to honey bees are characterized for chemicals and label language is recommended based on the outcome of a suite of tiered studies. Title 40 (protection of the Environmental) of the Code of Federal Regulations, Part 158 (Data Requirements for Pesticides) Subpart G (Ecological Effects) § 158.630 (Terrestrial and Aquatic Nontarget Organism Data Requirements Table)1 specifies current toxicity testing requirements for honey bee testing. Current test requirements include the following:

Under the current guidelines, the honey bee acute contact toxicity test is required for pesticide technical grade active ingredients (TGAI) with terrestrial, forestry and residential outdoor uses and is conditionally required for pesticides with aquatic uses as a Tier 1 screen conducted under laboratory conditions. If the results of the honey bee acute contact toxicity test indicate that a pesticide formulation contains one or more active ingredients with a median acute lethal dose to 50% of the animals tested, i.e., the LD50 value, of less than (<) 11 micrograms (µg) per bee and the use pattern indicates that honey bees may be exposed, then the toxicity of residues on foliage test is required as a Tier 2 laboratory-based test. As specified in CFR40 § 158.6305, field testing of pollinators is required if any of the following conditions are met:

Field studies are intended to represent real world conditions and are considered refined (Tier 3) toxicity tests. Pollinator field study designs have in the past varied considerably and EFED has identified study design elements that should be considered for these studies to improve consistency. Based on the outcome of the tiered toxicity tests, risk assessors have characterized the potential hazard to honey bees and have recommended environmental hazard statements based on the Label Review Manual6 (Table 1).

In determining whether additional data are needed for a pesticide, the first question the reviewer should ask is whether the use of the compound is likely to result in exposure to honey bees. If exposure is not considered likely, then data on effects are not necessary. However, even if exposure may initially be considered likely or at least possible, the reviewer should consider working with risk managers to determine whether the Registration Division (RD) and the Pesticide Re-evaluation Division (PRD) as well as registrants are willing to consider label language which may preclude potential exposure. If exposure cannot be precluded and/or if additional uncertainties remain regarding the potential effects of a pesticide or formulation on honey bees, then additional data may be necessary to address these uncertainties.

Table 1
Honey Bee Acute Toxicity Groups and Precautionary Statement Examples
(Reproduced from Label Review Manual7)
Toxicity Group Precautionary Statement if Extended Residual Toxicity is Displayed Precautionary Statement if Extended Residual Toxicity is not Displayed
I
Product contains any active ingredient with acute LD50 of 2 micrograms/bee or less
This product is highly toxic to bees exposed to direct treatment or residues on blooming crops or weeds. Do not apply this product or allow it to drift to blooming crops or weeds if bees are visiting the treatment area. This product is highly toxic to bees exposed to direct treatment on blooming crops or weeds. Do not apply this product or allow it to drift to blooming crops or weeds while bees are actively visiting the treatment area.
II
Product contains any active ingredient(s) with acute LD50 of greater than 2 micrograms/bee but less than 11 micrograms/bee.
This product is toxic to bees exposed to direct treatment or residues on blooming crops or weeds. Do not apply this product if bees are visiting the treatment area. This product is toxic to bees exposed to direct treatment. Do not apply this product while bees are actively visiting the treatment area.
III
All others.
No bee caution required. No bee caution required.

While current data requirements have proven useful for qualitatively describing potential hazards to honey bees from foliarly applied, nonsystemic pesticides, it is uncertain whether they are sufficient/suitable for characterizing potential hazards to honey bees from systemic compounds. System chemicals may be applied foliarly or through soil/seed treatments or trunk injections and bees may be exposed through ingestion of pollen and/or nectar and not through direct contact. One possible means of reducing uncertainty regarding potential exposure to systemic residues may be through the use of magnitude of residue studies (OCSPP Guideline 860.15008) submitted to the Health Effects Division. At this time, it is unknown whether measures of total foliar residues could be considered protective for potential residues in pollen/nectar; however, they may be more reasonable estimates than those based on plant surface residues. Appendix 2 contains a comparison of the strengths and limitations of various studies that could be considered in evaluating potential exposure to pesticides.

To assess potential effects, the reviewer should consider recommending acute oral toxicity tests with young adult honey bees to address uncertainties regarding the ingestion of systemic residues through pollen/nectar. Depending on the toxicity and persistence of systemic residues, additional "higher tier" exposure and effect studies may be necessary for systemic pesticides.

In determining what additional data may be necessary to characterize the potential hazards of a pesticide to honey bees, the CFR 40 Part 1589 indicates that data from other sources (e.g., Experimental Use Permit program, university research, registrant submittals, etc.) showing potential adverse effects on [honey bee] colonies should be considered. As such, multiple lines of evidence must be used to determine the need for additional information and the nature of that information. EFED reviewers should therefore consider existing guideline toxicity data on the chemical and data which may be available through European (EU) data requirements if the compound is registered in the EU; these data may include acute oral toxicity studies, i.e., Organization of Economic Cooperation and Development (OECD) Guideline 21310. Reviewers should also note that OECD has a test guideline for honey bee acute contact toxicity testing as well, i.e., OECD Guideline 21411.

With the exception of field pollinator toxicity testing (OCSPP Guideline 850.3040), the toxicity tests discussed thus far focus on young adult honey bees as the test organism; however, a pesticide may not be sufficiently toxic to kill forage bees immediately upon contact/consumption, and these compounds, whether they are attached to the outside of the bee as fine particulates (dust) or are in pollen/nectar, may be returned to the colony where other castes of bees (e.g., nurse bees, larvae/pupae (brood), drones, queen) or eggs may be potentially exposed. While young adult honey bees typically used for testing may be considered representative of other adult bees in the colony, it is uncertain whether toxicity values for young adult bees can be considered representative for developing brood that can potentially be exposed to residues through consumption of brood food and/or royal jelly secreted by nurse bees or via residues that have partitioned to comb wax in which the brood develops. If there are data to indicate that developing brood may be adversely affected from the use of a pesticide or formulated products, then consideration should be given to requesting honey bee acute larval toxicity tests. For example, insect growth regulators (IGRs) such as ecdysone agonists should be evaluated for potential developmental effects on larvae. The European Plant Protection Organization (EPPO) has published draft guidance by Oomen et al. (1992)12 on honey bee brood feeding tests; copies of this guidance are available through the Terrestrial Biology Technical Team Pollinator Protection Team folder on the G-share drive. OECD Guidance Number 7513 on honey bee brood testing under semi-field conditions (also available through the G-share drive) is a synthesis of the Oomen et al. (1992)14 study. Other methods to consider include that by Aupinel et al. 200715 method (also available on the G-share drive). Because several methods are available for toxicity testing with honey bee larvae, the reviewer should recommend that the registrant be required to submit a protocol for review and approval by EPA prior to study initiation.

If laboratory tests with adults and/or larvae indicate the potential for adverse effects on the entire colony, consideration should be given to requiring a semi-field study before a full field study is required. Although not specifically identified in the Part 158.63016, semi-field toxicity tests are becoming increasingly available through EU registrations. The field pollinator study (OCSPP Guideline 850.304017) can be sited as a placeholder for semi-field studies. The benefits of these studies is that they facilitate the study of whole colonies (although the colonies are typically reduced in size to nucleus "nuc" colonies) that can be confined in enclosures (tunnels) for limited periods of time. Semi-field studies better ensure exposure to the chemical of interest and limit the confounding effects of other pesticides which can be returned to colonies by unconfined free-foraging bees. OECD Guidance No. 7518 provides a useful description of the study design elements; however, reviewers should request that a protocol be provided in advance for approval before the semi-field study is initiated. Consideration should also be given to ensuring that the study monitors colonies for several weeks following their removal from the enclosures.

Full field studies remain a major challenge in both their conduct and analysis/interpretation. These studies require large amounts of resources both for the regulated community to conduct and for EPA to review. As indicated earlier, these studies represent the highest tier of testing and are intended to address specific uncertainties that have been identified in lower tier testing. Field pollinator studies are also intended to represent more realistic application scenarios and bee foraging environments and typically involve commercial bee colonies where a number of factors may be simultaneously affecting the study. It is critical that EFED reviewers work with the risk managers and the registrant in articulating the need for these studies and in developing a suitable protocol for the studies. Given the large amount of resources needed to conduct full field studies and given that honey bees can forage for considerable distances (up to 8 miles), reviewers may need to be flexible in evaluating study performance and in determining the utility of the data.

As noted in the CFR40 Part 15819, university research can serve as a useful source of information on the potential toxicity of pesticides to honey bees and on whether there is a need for additional data; however, they can also serve as a useful source of information on the toxicity of pesticides to non-Apis bees such as bumble bees (Bombus spp.), alfalfa leaf cutter bees (Megachile rotundata) and blue orchard bees (Osmia lignaria). The U.S. Department of Agriculture (USDA) Agricultural Research Service (ARS) and National Institute of Food and Agriculture (NIFA) provide funding through efforts such as the Managed Pollinator Coordinated Agricultural Project (CAP) to support pesticide-related research on honey bees. Abstracts of these research efforts are published in the Exit EPA Disclaimer Proceedings of the American Bee Research Conference 2011. Progress reports on USDA-funded research on pollinators and pesticides are also available through the USDA-led Colony Collapse Disorder Steering Committee annual progress reports to Congress: Colony Collapse Disorder Progress Report 2010 and Colony Collapse Disorder Progress Report 2009.

Open literature can also provide reviewers an understanding of the extent to which pesticide residues may be moving into different honey bee colony matrices, e.g., honey/pollen (bee bread) and wax. Although open literature studies should be considered in the context of the EFED open literature guidance, articles such as that by Mullin et al. (2011)20 can provide insight into the frequency and quantity of specific pesticides and their degradates that are identified in honey bee colonies. It is important to note that many monitoring studies have associated biases due to the way in which data are collected. Risk assessors need to characterize these potential biases that may limit the utility of the monitoring data for assessing trends nationally.

Reviewers in EFED should make use of multiple lines of evidence in determining whether additional study data should be recommended. An important consideration should be incident data submitted under FIFRA 6(a)221. Incident data can be obtained through the Ecological Incident Information System (EIIS) that can be accessed through the EFED Gateway. As with any incident information, the reviewer should not simply tally the number of incidents but make an effort to review the nature of the incident to determine the strength of the cause-effect relationship.

Although data may not be available for the specific chemical under evaluation, the reviewer should consider whether pollinator-related impacts have been observed for chemicals with similar modes of actions and/or similar structures. These data could potentially be used for bridging purposes to characterize potential hazards in the absence of chemical-specific data.

Reliance on bridging data introduces considerable uncertainty into the effects characterization and this uncertainty should be articulated in the assessment. Bridging data however can serve as an effective rationale for recommending additional data are required on honey bees.

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Footnotes

1 CFR40. 2011. Part 158, subpart G, § 158.630

2 USEPA. 1996a. Ecological Effects Test Guidelines OPPTS 850.3020. Honey bee Acute Contact Toxicity. EPA 712-C-96-147.

3 USEPA 1996b. Ecological Effects Test Guidelines OPPTS 850.3030. Honey bee Toxicity of Residues on Foliage. EPA 712-C-96-148.

4 USEPA 1996c. Ecological Effects Test Guidelines OPPTS 850.3040. Field Testing for Pollinators. EPA 712-C-96-150.

5 Ibid CFR40 2011.

6 USEPA. 2008. Label Review Manual. Chapter 8: Environmental Hazards. Section 4 Nontarget Organism Statements B. Honey-bee Hazard Statements Table 2 Updated November 2008.

7 Ibid USEPA 2008.

8 Residue Chemistry Test Guidelines OPPTS 860.1500 Crop Field Trials

9 Ibid CFR40. 2011.

10 OECD. 1998a. OECD Guidelines for the Testing of Chemicals. Exit EPA Disclaimer Honey bees. Acute Oral Toxicity Test. 213.

11 OECD. 1998b. OECD Guidelines for the Testing of Chemicals. Exit EPA Disclaimer Honey bees. Acute Contact Toxicity Test. 214.

12 Oomen, P. A., A. DeRuijter and J. Van Der Steen. 1992. Method for honey bee brood feeding tests with insect growth-regulating insecticides. Bulletin OEPP/EPPO 22: 613 - 616.

13 OECD. 2007. Guidance document on the honey bee (Apis mellifera L.) brood test under semi-field conditions. Series on Testing and Assessment No. 75. ENV/JM/MONO(2007)22.

14 Ibid Oomen et al. 1992.

15 Aupinel, P., D. Fortini, B. Michaud, F. Marolleau, J-N Tasei and J-F Odoux. 2007. Toxicity of dimethoate and fenoxycarb to honey bee brood (Apis mellifera), using a new in vitro standardized feeding method. Pest Management Science 63: 1090 -1094.

16 Ibid CFR40. 2011.

17 Ibid USEPA 1996c.

18 Ibid OECD. 2007

19 Ibid CFR40. 2011

20 Mullin, C.A., M. Frazier, J.L. Frazier, S. Ashcraft, R. Simonds, D.vanEngelsdorp, and J.S. Pettis. 2010. Exit EPA Disclaimer High levels of Miticides and Agrochemicals in North American Apiaries: Implications for Honey Bee Health. PlosONE 5(3): e9754. doi:10.1371/journal.pone.0009754.

21 USEPA. 2011. Adverse Effects Reporting: FIFRA 6(a)2.

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Appendix 1
Considerations for Calling-In Honey Bee Hazard Assessment Data

Evaluating need for additional data depends on the routes of exposure that are of potential concern.

  1. Direct Contact

  2. Dermal/Foliar residue

  3. Oral ingestion (pollen, nectar, guttation water)

The following is a list of assessment questions that should be addressed when determining the need for additional data to evaluate hazards to honey bees (and in the future, risk to pollinators).

  1. Direct Contact Exposure

    1. Method of application

      Could the method of application result in direct contact of foraging bees? (e.g., foliar spraying)

    2. Attractiveness of crop

      Are crops to which the pesticide is applied known or suspected to be foraging sites for bees?

    3. Timing of application

      Does the timing of application coincide with bloom? Is the pesticide being applied to crops with extrafloral nectaries (e.g., cotton) and/or indeterminant bloom (e.g., pumpkin and other cucurbits)?

    4. Toxicity

      Is the pesticide toxic on an acute contact basis?

  2. Dermal (foliar) Residue Exposure

    1. Contact Toxicity

      Is the pesticide considered moderately to highly toxic to bees on an acute contact basis (e.g., < 11 µg/bee?), and does the foliar residue study show prolonged toxicity?

    2. Use pattern

      Does use pattern indicate likely contact with pesticide via foliar residue (e.g., foliar application, moderate to high persistence)?

    Based on the strength of the various lines of evidence above, consider honey bee language on labels to mitigate direct contact exposure (restrictions on timing of application); other labeling may be necessary for crops where direct contact may not be prevented. If hazards/concerns cannot be effectively mitigated via label restrictions for direct and foliar contact routes of exposure, consideration can be given to a semi-field (tunnel) study (see Appendix 2). Such a study would help document "high-end" exposure and impacts to honey bees and their brood due to adult mortality via direct and/or foliar contact with the pesticide.

  3. Oral Exposure

    1. Exposure from Pollen and Nectar

      What information exists to suggest that the pesticide application may result in appreciable exposure through consumption of pollen and nectar?

      1. Is the pesticide systemic?

        Consider HED plant metabolism, magnitude of residue, rotational crop studies.

        Consider physicochemical properties (Kow, pka)

        Consider mode of application (e.g., seed treatment insecticide)

      2. Is the pesticide sufficiently persistent on or in plants such that appreciable residues in pollen and nectar could result following application?

        Consider HED plant metabolism, magnitude or residue, rotational crop studies, other fate information, timing of application

      3. Are crops attractive to bees?

        Consider target crop and rotational crops

      4. What data exist in the literature or what other lines of evidence are available to support appreciable exposure via pollen and nectar?

      Based on the strength of the various lines of evidence above, consider requesting studies that would document the magnitude of exposure via pesticide residues in pollen and nectar. Such information can be obtained from field crop residue studies and semi-field (tunnel) colony studies. Note: in order to interpret the significance of residues in pollen and nectar, an oral toxicity study (see below and Appendix 2) will likely be needed.

    2. Oral Toxicity

      What information exists to suggest the pesticide is toxic at environmentally relevant concentrations via the oral route of exposure? How likely might the sensitivity of developing brood or other metrics of colony health differ appreciably from currently available information (i.e., toxicity to adult workers)?

      1. Consider OECD oral toxicity study to adult worker bees.

      2. Does the mode of action suggest differential toxicity among adults vs developing larvae (e.g., insect growth regulators, certain neonicotinoids)

      3. Consider available information on the effects to developing brood (e.g., OECD semi-field studies, open literature, efficacy studies comparing larval and adult sensitivity).

      4. Consider evidence of sublethal effects that would directly relate to colony health but would not be captured using the existing data.

      Based on the strength of the various lines of evidence above, consider requesting toxicity studies that would enable interpretation of exposure via pollen and nectar. Such studies would include adult oral toxicity, in vitro larval toxicity, in colony brood toxicity, semi-field (tunnel) brood toxicity, and field colony health studies. See Table 1 for the relative strengths and limitations of these studies.

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Appendix 2
Strengths and Limitations of Various Studies Related to Honey Bee Exposure and Effect Assessments


Defining the Nature of Pesticide Uptake in Plants
(e.g., pesticide translocation, metabolism and persistence in plant tissues)
Study NameDescriptionProtocol Status StrengthsLimitationsReference
HED Plant Metabolism Study Typically 14-d studies of pesticide (and metabolite) residues in various plant tissues following application Approved OSCPP Guideline 860.1300
[conditionally required for food uses]
Identification of parent and metabolite residues in various crop tissues (e.g., radish top vs. root) over time; some inference about potential systemicity can usually be made Generally short duration (e.g., 14 to 28d); tissues sampled are relevant for human health concerns, not pollinators (e.g., no pollen, nectar) Residue Chemistry Test Guidelines: OPPTS 860.1300 Nature of the Residue - Plants, Livestock
HED Rotational Crop Study Typically long duration study (e.g., 1 year) that evaluates the persistence and translocation of soil applied pesticide in plants Approved OSCPP Guideline, 860.1900
[required for food uses]
Identification of parent and metabolite residues in various crop tissues at various intervals after soil application; some inference of systemicity may be made based on translocation from soil to various plant tissues Tissues sampled are relevant for human health concerns, not pollinators (e.g., no pollen, nectar)

Title 40: Protection of Environment, PART 158, Subpart O - Residue Chemistry, 158.1410 Residue chemistry data requirements table

Guideline:

OPPTS 860.1900 Field Accumulation in Rotational Crops

Plant Uptake / Phosphor Imaging Study Phosphor imaging of pesticide residues in plant tissues Non-guideline / special study Identifying mechanism of and timing of translocation (xylem vs. phloem movement) Qualitative descriptions of translocation (Concentrations may not be provided, rather phosphor images showing extent of pesticide movement from point of application) Special Study (e.g., sulfoxaflor plant uptake study: MRID 484468-04)
HED Magnitude of Residue Study   Approved OSCPP Guideline 860.1500
[crop field trials required for food uses]
  Nature of the plant tissues sampled vary depending on the crop; must insure total residues are measured and not just dislodgeable residues

Title 40: Protection of Environment, PART 158, Subpart O - Residue Chemistry,158.1410 Residue chemistry data requirements table

Guideline:

OPPTS 860.1500 Crop Field Trials

Also see:

OPPTS 860.1000 Background


Determining Pesticide Residues in Pollen and Nectar Resulting from Label Application
Study NameDescriptionProtocol Status StrengthsLimitationsReference
OECD semi-field tunnel study Honey bee semi-field (tunnel) study for evaluating effects of foliar-applied pesticide to bee brood, adult survival, and flight intensity; study duration varies, but bees confined to tunnel will show signs of stress within roughly 7 days. OECD Guideline 75: Honey bee (Apis mellifera L) Brood Test Under Semi-Field Conditions; proposed protocol should be submitted for review and approval by EFED prior to study initiation. Realistic exposures and effects on various measures of brood development and bee health. Can also provide measures of residue in pollen and nectar. Bee bread can be spiked and exposure to test material assured. Relatively short exposure (7-days) due to tunnel stress. Representativeness of test flora (usually Phacelia) of agricultural crops may be questionable. Natural variability in brood measures of effect can be high. Number 75 Guidance Document on the Honey bee (Apis mellifera L.) Brood Test Under Semi-field Conditions
(see G drive)
Pollen/nectar residue study Special study for quantifying the amount of pesticide in pollen and nectar resulting from labeled applications. Non-Guideline / special study; proposed protocol should be submitted for review and approval by EFED prior to study initiation. Enables direct estimation of residues in pollen/nectar for specific crops and application rates. Does not quantify effects; residues must be interpreted with separate toxicity studies and consumption estimates; in-hive exposure of larvae may differ from measured residues due to pollen/nectar processing; alternative forage may be available under natural foraging activity. Special study. Registrants must submit protocol in advance for review and approval by EPA.
Field monitoring studies (e.g., Mullin et al. 2010) Literature data on pesticide residues measured in pollen, nectar, bees, and hives Open literature; proposed protocol should be submitted for review and approval by EFED prior to study initiation. Provides some quantification of potential exposure of bees to pesticides in various media. Often, exposure history is unknown (e.g., timing and amount of pesticide application; foraging locations, etc.). Some studies (e.g., Mullin et al. 2010) contain measurements associated with bee kill events and do not necessarily reflect "background." High Levels of Miticides and Agrochemicals in North American Apiaries: Implications for Honey Bee Health

Determining Toxicity to Honey bees via Various Routes of Exposure
Study NameDescriptionProtocol Status StrengthsLimitationsReference
Acute Contact Toxicity Single dose; acute contact study typically conducted for 48 hrs. Guideline study (850.3020; OECD 214) Standard methods. Mortality dose-response is quantified with replication. Endpoints limited to mortality to young adult worker bees and clinical signs of intoxication (i.e., sublethal effects are not typically well quantified although guidelines identify need to report).

OPPTS 850.3020 Honey Bee Acute Contact Toxicity

Test No. 214 Honeybees, Acute Contact Toxicity Test
(see G drive)

Foliar Residue Toxicity Toxicity of foliar residues; typically conducted for 24 hrs and then expanded at 24-hr intervals depending of toxicity of foliar residues Guideline study (850.3030) Standard methods. Toxicity from exposure to treated foliage is determined at various post application intervals. Endpoints limited to mortality to young adult worker bees and clinical signs of intoxication (i.e., sublethal effects are not well quantified); may provide time to effect data. OPPTS 850.3030 Honey Bee Toxicity of Residues on Foliage
Acute Oral Toxicity Acute oral toxicity to honey bee; typically 48 hrs. duration but may be extended to 96 hrs. OECD 213
(no OSCPP guideline)
Standard methods. Mortality dose response from oral ingestion of pesticide with replication. Endpoints limited to mortality to young adult worker bees and clinical signs of intoxication (i.e., sublethal effects are not typically well quantified). Test 213 Honeybees, Acute Oral Toxicity Test
(see G drive)
Larval Toxicity In vitro toxicity to honey bee brood (larvae); 96-hr study duration. Special study; proposed protocol should be submitted for review and approval by EFED prior to study initiation. Quantifying effects on bee larvae and adult emergence; method is currently being ring tested. Non-standard study. Reportedly, a small number of laboratories can successfully conduct this study; exposure of larvae may not be representative of in hive exposure. Aupinel, P., D. Fortini, B. Michaud, F. Marolleau, J-N. Tasei and J-F Odoux. 2007. Toxicity of dimethoate and fenoxycarb to honey bee brood (Apis mellifera), using a new in vitro standardized feeding method. Pest Management Science 63: 1090-1094.
(see G drive)
Brood Feeding Test In-hive exposure of brood to pesticides consumed and fed by adults; study duration varies; Oomen study monitors brood for 3 wks. Special Study; proposed protocol should be submitted for review and approval by EFED prior to study initiation. Quantifying effects on brood from pesticide exposure under natural field conditions; study can support longer durations than semi-field tunnel study because bees are not constrained. Non-standard study. Although exposure in the diet is known, the actual amount fed to brood is not. Bees may forage aloft which may "dilute" exposure to spiked diet and/or increase exposure to other pesticides.

Oomen, P.A., A. De Ruijter and J. Van Der Steen. 1992. Method for honey bee brood feeding tests with insect growth-regulating insecticides. Bulletin OEPP/EPPO 22: 613-616
(see G drive)

OPPTS 860.1000 Background

OECD Semi-field Tunnel See above. See above; proposed protocol should be submitted for review and approval by EFED prior to study initiation. See above. See above. Typically rely on smaller nucleus "nuc" colony. Able to track colony performance once removed from tunnel; however, colonies are typically too small to successfully overwinter unless supplemental food is provided.  
Field Pollinator Study Free foraging bees with full-size colonies (8-9 frames) with - 50,000 bees/colony Guideline study (850.3040); proposed protocol should be submitted for review and approval by EFED prior to study initiation. Intended to provide more realistic estimates of actual exposure conditions and potential effects. Multiple measurement endpoints both at the individual bee and colony level. Brood as well as adult bee measurement endpoints. Provides means of tracking overwintering success. Current methodology not well defined; draft study design elements have been developed in cooperation with USDA, (see G drive). Considerable variability associated with measurement endpoints since studies cannot be heavily replicated. Difficult to adequately separate treatments since bees can forage up to 10 miles. Existing pesticide loads in colonies can confound interpretation of study.

OPPTS 850.3040 Field Testing for Pollinators

Registrants must submit protocol in advance for review and approval by EPA.

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