### NATIONAL ENVIRONMENTALPOLICYACT DECISION AND FINDING OFNO SIGNIFICANT IMPACT

Permitapplication 11-052-lOlrm

received from ArborGen LLC

Field testing ofgenetically engineered Eucalyptusgrandis X Eucalyptus urophylla



### United States DepartmentofAgriculture Animal and PlantHealth Inspection Service Biotechnology Regulatory Services

United States Department ofAgriculture (USDA) and the Animal and Plant Health Inspection Service (APHIS) have developed a decision documentto complywith the requirements ofthe National Environmental Policy Actof 1969, as amended, the Council ofEnvironmental Quality's (CEQ) regulations implementing NEPA, and the USDA and APHIS NEPA implementing regulations and procedures. This NEPA decision document is intended to state APHIS' NEPA decision and presentthe rationale for its selection.

In accordance with APHIS procedures implementingthe NEPARegulations (7 CFRpart 372), APHIS has prepared an Environmental Assessment (EA) to evaluate and determine ifthere are anypotentially significant impacts to the quality ofthe human environment in response to a permit application (11-052-101rm) received from ArborGen for the confined environmental release and interstate movement. Ifissued, the permitwould authorize the planting of genetically engineered (GE) Eucalyptus hybrid clone (Eucalyptus grandis x Eucalyptus urophylla) to supportproposed field research studies on six research sites in Alabama, Florida, Mississippi, and South Carolina. The permit application requests that APHIS permitall the trees exceptthe trees ontwo ofthe sites in South Carolinato flower. The purpose ofthe confined environmental release is for ArborGen to assess the effectiveness ofintroduced gene constructs which are intended to confer coldtolerance; to test the efficacy ofgenes to alter lignin biosynthesis; to testthe efficacy ofgenes to alter growth; andto test the efficacy ofgenes designed to alter flowering. In addition the trees have been engineered with a selectable marker gene which confers resistance to the antibiotic kanamycin. ArborGen has also requested under this permit application that all genetically engineered Eucalyptus trees authorized underpermits 09-070-101rm and 11-041-101rm be incorporated into this newpermit and allowed to flower. Newtrees will also be moved interstate from ArborGen's nursery and greenhouse facilities located in South Carolinato the release locations identified in the permitapplication where they will be planted.

The EA assesses the potential impacts ofAPHIS issuing or denying a confined environmental release and interstate movementpermitwith supplemental permit conditions to allow the interstate movement, planting, field testing and flowering ofa GEEucalyptus hybrid clone (Eucalyptus grandis x Eucalyptus urophylla) engineered to express various genes at 6 confined field site locations within in the States ofAlabama, Florida, Mississippi, and South Carolina encompassing a total of 14.7 acres and analyzes the potential environmental and social effects thatresult from the proposed action and the alternatives. The proposed action ofUSDA APHIS, Page 1 of28Biotechnology Regulatory Services (BRS) is to issue the APHIS confined field release and
movementpermit for a hybrid ofEucalyptus grandis X Eucalyptus urophylla with supplemental permit conditions (see Appendices VI and VII ofthe EA). The movement authorization would be valid for one year and the release would be valid for a three-yearperiod. Comments from the public involvementprocess were reviewed for substantive issues which were considered in developing this NEPA decision.

Since 1986, the United States governmenthas regulated GE organisms pursuantto a regulatory framework known as the Coordinated Framework for the Regulation ofBiotechnology (Coordinated Framework) (51 FR23302, 57 FR 22984). The Coordinated Framework, published bythe Office ofScience and Technology Policy, describes the comprehensive federal regulatorypolicy for ensuringthe safety ofbiotechnology research and products and explains how federal agencies will use existing Federal statutes in a mannerto ensure public health and environmental safety while maintaining regulatory flexibility to avoid impedingthe growth of the biotechnology industry. The Coordinated Framework is based on several important guiding principles: (1) agencies should define those transgenic organisms subjectto review to the extent permitted bytheirrespective statutory authorities; (2) agencies are required to focus on the characteristics and risks ofthe biotechnology product, not the process by which it is created; and (3) agencies are mandated to exercise oversight ofGE organisms only when there is evidence of "unreasonable" risk.

The Coordinated Frameworkexplains the regulatory roles and authorities for the three major agencies involved in regulating GE organisms: USDA'sAPHIS, the Food and Drug Administration (FDA), and the Environmental Protection Agency (EPA).

APHIS is responsible for regulating GE organisms and plants under the plantpestprovisions in the PlantProtection Act of2000, as amended (7 USC 

### § 7701 et seq.) to ensure that they do not pose a plantpest risk to the environment. APHIS regulations atTitle 7 ofthe Code ofFederal Regulations (CFR) part 340, which were promulgated pursuantto authority granted bythe Plant Protection Act, as amended (7 United States Code (U.S.C.) 7701-7772), regulate the introduction (importation, interstate movement, orrelease into the environment) ofcertain GE organisms and products. A GE organism is considered a regulated article ifthe donor organism, recipient organism, vector, or vector agentused in engineering the organism belongs to one of the taxa listed in the regulation (7 CFR 340.2) and is also considered a plantpest. A GE organism is also regulated under Part 340 when APHIS has reason to believe thatthe GE organism may be a plantpest orAPHIS does nothave information to determine ifthe GE organism is unlikely to pose a plantpest risk.

When APHIS receives an application for a permit for environmental release and movement, the application is evaluatedto determine whetherthe environmental release and movement, with appropriate conditions imposed, can be carried outwhile preventing the dissemination and establishment ofplantpests. The receipt ofa permit application to introduce a genetically engineered organism requires a response from the Administrator:

Administrative action on applications. After receipt and review byAPHIS of the application and the data submittedpursuantto paragraph (a) ofthis section, including any additional information requested by APHIS, a permit shall be granted or denied (7 CFR 340.4(e)).
Page 2 of28The applicant has provided the required information associated with this request in the permit application. This information has been reviewed byAPHIS-BRS and is analyzed in the EA.

FDA regulates underthe authority ofthe Federal Food, Drug and Cosmetic Act. The FDA policy statement concerning regulation ofproducts derived from newplant varieties, including those genetically engineered, waspublished in the Federal Register on May 29, 1992 (57 FR 22984-23005). Underthis policy, FDA uses what is termed a consultation process to ensure that human food and animal feed safety issues or other regulatory issues (e.g., labeling) are resolved priorto commercial distribution ofbioengineered food. The EPA regulates plant-incorporated protectants underthe Federal Insecticide, Fungicide and Rodenticide Act (FIFRA) and certain biological control organisms underthe Toxic Substances Control Act (TSCA). This GE Eucalyptus hybrid is not used for food orfeed purposes and does not contain any genetically engineeredpesticides ortolerance to herbicides.



### PublicInvolvement In a notice published in the Federal Register on February 10, 2012, (77 FR28 7123-7124, Docket No. APHIS-2011-0130), APHIS announced the availability ofan EA for public review and
comment for a proposed controlled field release ofa genetically engineered clone ofa Eucalyptus hybrid. Comments on the environmental assessmentwere required to be received on

or before March 12, 2012. APHIS received a total of246 comments during the 30-day comment period. All respondents expressed concerns about the permitoropposed granting the permit. Comment documents may be viewed at http://www.regulations.gov/#!searchResults;rpp=25:po=O;s=APHIS%25E2%2580%25932011% 25E2%2580%25930130. All comments were analyzed to identify new issues, alternatives, or information. Responses to the substantive comments are included as an attachmentto this Finding ofNo Significant Impact.



### MajorIssues Addressed in theEA The EA describes the alternatives considered and evaluated using the identified issues. Issues considered in the EA were developed based on APHIS' determination to issue a confined environmental release and interstate movementpermit with supplemental permit conditions to allow the interstate movement, planting, field testing and flowering ofa GEEucalyptus hybrid clone (Eucalyptus grandis x Eucalyptus urophylla) engineered to express various genes at 6 confined field site locations within the States ofAlabama, Florida, Mississippi, and South Carolina. The following issues were identified as importantto the scope ofthe analysis (40 CFR 1508.25):

• Interstate Movementofthe trees underpermit

• Alteration in Susceptibilityto Disease orInsects- Potential ofthe Eucalyptus to Harbor PlantPests

• Expression ofthe Gene Products, New Enzymes, orChanges to PlantMetabolism- Risk ofthe Gene Products on the Environment

• Alteration in Weediness characteristics

- Potential ofthe Engineered Eucalyptus to be Invasive

• Possibility ofGene Flow Withinthe Field Test

• Possibility ofGene Flow Outside ofthe Field Test

• Possibility ofVegetative Propagation I Persistence Outside ofthe Field Test Page 3 of28• Potential ofthe Eucalyptus in the Field Tests to Become an Invasive Species that Threatens Native Plant and Animal Communities

• Impact on Existing Agricultural Practices

• Potential Impacts to Wildlife

• Potential Impacts by Fire

• Potential Impacts to Human Health

• Transfer ofGenetic Information to Organisms with which it Cannot Interbreed

- Horizontal Gene Transfer to Other Organisms

• Potential Effects ofGrowingEucalyptus on Soil Hydrology

• Potential Allelopathic Effects ofEucalyptus

• Risks to Threatened and Endangered Species

• Impacts on Unique Characteristics ofProtected Areas

Affected Environment: The confined field tests would take place on land controlled by ArborGen orthrough private
contracts established by ArborGen for field testing. The exact locations are claimed as CBI and have been submitted as part ofthe APHIS permitapplication, however the States and Counties where the confined field tests would occur are Baldwin and Escambia Counties, Alabama; Highlands County, Florida; Pearl River County Mississippi; and Berkeley and Dorchester Counties, South Carolina. Underthe permit application submitted by ArborGen, there are six research sites where trees would be planted. A total of 14.7 acres (all 6 sites combined) is being requested bythe applicant. Five ofthese locations currently have active APHIS permits (08- 0ll-106rm, 08-014-lOlrm, 09-070-lOrm, 10-112-lOlr, and 11-041-lOlrm) for environmental release ofGE Eucalyptus hybrid clone (Eucalyptus grandis x Eucalyptus urophylla) in Alabama, Florida, Mississippi, and South Carolina. The remaining site in Dorchester County, South Carolina has been listed in previous APHIS permits/notifications as a holding area for transgenic
trees. ArborGen is requesting thattrees be allowed to flower at four locations in Alabama, Florida and Mississippi. Atthe two locations in Berkeley and Dorchester Counties, SC, they have requested to release trees in containers and have indicatedthey will not allow these trees to flower at these locations. The size ofeach individual confined field test sites identified in ArborGen's permit application ranges from 0.5 to 7.7 acres, which is less than the 20 acres analyzed in APHIS EA and subsequentFONSI prepared for Permits 08-0ll-106rm, 08-014- lOlrm and 10-112-lOlr (http://www.aphis.usda.gov/brs/aphisdocs/06 325lllr ea.pdf) and (http://www.aphis.usda.gov/brs/aphisdocs/08 014101r ea.pdf). Forthe five research sites that

are currently authorized by APHIS to plant GE Eucalyptus hybrid clone (Eucalyptus grandis x Eucalyptus urophylla), ArborGen has indicated in their permit application that any new planting authorized by APHIS under 11-052-lOlrm would not expandthe current field site locations beyond the areas currently authorized byAPHIS.

All the confined field test sites listed in the permit application by ArborGen are either on privately owned and managedplantation forests and agricultural farm lands or experimental research stations managed by academic institutions and industry, and have been in managed agricultural production for more than 5 years. The standard agricultural and silvicultural practices for land preparation, planting, irrigation, and harvesting ofplants have been routinely used onthese sites. Sitesthat include managed pastures have had intense activity including the

use ofheavy machinery for general upkeep, irrigation, fertilization, controlled grazing and management ofgrasses. Standard silvicultural practices would be used atthese sites for the duration ofthe field tests. Surveys conducted bythe applicant ateach ofthese locations indicate Page 4 of28thatthere are not any old growth forests orundisturbed natural areas in the immediate surroundings ofthe test sites. ArborGen has indicatedthatthe trees would be planted on individual research sites ranging from 0.5 up to 7.7 acres, depending on the location with planting density ranging from 300

- 600 trees per acre 1•

The EA analyzes the potential environmental consequences ofa proposal for APHIS to issue a confined environmental release and interstate movementpermitwith supplementalpermit conditions to allow the interstate movement, planting, field testing and flowering ofa GE Eucalyptus hybrid clone (Eucalyptus grandis x Eucalyptus urophylla) engineeredto express various genes at 6 confined field site locations within the States ofAlabama, Florida, Mississippi, and South Carolina. Based upon the permit application submitted by ArborGen, two alternatives are considered and analyzed in the EA: (1) deny the permit and (2) issue the APHIS permit.



### AlternativeA: No Action- Deny thePermit Underthe No Action alternative APHIS would deny the permitapplication (11-052-101rm) submitted by ArborGen. ArborGen would not be authorized to move andplantthe GE Eucalyptus trees with new constructs and allow them to reach maturity and flower. Confined field release under permits 09-070-101rm and 11-041-101rm would continue to not allow flowering.

There are currently six active permits underwhich ArborGen is authorized by APHIS to grow GE Eucalyptus which include 32 unique locations within 7 States: 08-011-106rm, 08-014- 101rm, 09-070-101rm, 10-112-101r, 11-041-101rm and 11-201-103r. Noplantings at any ofthe 32 locations are authorized by APHIS to exceed 20 acres in size. Trees are allowed to flower under four permits: 08-011-106rm, 08-014-101rm, 10-112-101r and 11-201-103r (two ofthe locations in Berkeley and Dorchester Counties, SC trees are not authorized to flower). Under permits 09-070-101rm and 11-041-101rm trees are not authorized to flower. As ofSeptember 2011, ArborGen has reported to APHIS that they are growing a total ofapproximately 67 acres oftrees on 18 ofthe 32 permitted locations. Underthe No Action alternative, ArborGen could continue to grow GEEucalyptus as authorized by APHIS under these six active permits.



### AlternativeB: Preferred Alternative

- Issue theAPHIS Permit The Preferred Alternative is to approve ArborGen permit application request (11-052-101rm) and issue the APHIS confined field release and movementpermit for a hybrid ofEucalyptus grandis X Eucalyptus urophylla with supplemental permit conditions (see Appendices VI and VII ofthe EA). The movement authorization would be valid for one year and the release would be valid for a three-yearperiod. The permitwill need to be renewed by ArborGen and subsequently approved by APHIS to allow the transgenic Eucalyptus plants to remain in the ground beyondthe three-yeartime period specified in the permit application.

Underthis alternative, APHIS would issue a confined environmental release permitto ArborGen in accordance with 7 CFRpart 340 to allow the movement, planting, field testing and flowering

1 Planting density typically refers tothe number oftrees per acre. Planting densities can vary greatly depending upon the tree species and the environment, but densities ofshort rotation hardwood trees in the southeastern US

are typically in the range of30Q-800 trees per acre. Therefore sites ranging from 10to 20 acres can have from 3000 to 16,000total trees planted in the ground. Twenty acres, as defined by forest plantation standards in the southeast, is considered a small planting. Page 5 of28ofa GE Eucalyptus hybrid clone engineered to express various genes at 6 confined field site locations within in the States ofAlabama, Florida, Mississippi, and South Carolina encompassing a total of 14.7 acres. The permit would allow all the trees exceptthe trees ontwo ofthe sites in South Carolinato flower. All genetically engineered Eucalyptus trees authorized under permits 09-070-101rm and 11-041-101rm would be incorporated into this newpermit and allowed to flower. This permit would authorize ArborGen to move and plantthe GE Eucalyptus
trees with new constructs and allow them to reach maturity and flower. Under this alternative, the applicant would be allowed to gather data on performance ofthe transgenic trees over a multiyearperiod and the efficacy ofthe genes in a wide variety ofenvironments for multiple years. This alternative would allow the safe development and use ofGE organisms under the mission ofBRS.

Environmental Consequences ofAPHIS' Selected Action The EA contains a full analysis ofthe alternatives to which we refer the reader for specific details. The following table briefly summarizes the results for each ofthe issues fully analyzed in the Environmental Consequences section ofthe EA.

Page 6 of28

### Table 1.Issue AlternativeA AlternativeB NoAction Pre{j!rredAlternative Denv thePermit IssueAPHISPermit Interstate Movementofthe trees under No increased risk No increased risk permit Alteration in Susceptibilityto Disease or No increased risk No increased risk Insects

- Potential ofthe Eucalyptus to Harbor PlantPests Expression ofthe Gene Products, New No expected risk No expected risk Enzymes, or Changes to Plant Metabolism

- Risk ofthe Gene Products on the Environment Alteration in Weediness characteristics

- Not likely Not likely Potential ofthe EngineeredEucalyptus to be Invasive Possibility ofGene Flow within the Field Minimal Minimal Test Possibility ofGene Flow Outside ofthe Minimal Minimal Field Test Possibility ofVegetative Propagation I Not likely Not likely Persistence Outside ofthe Field Test Potential ofthe Eucalyptus in the Field Not likely Not likely Tests to Become an Invasive Species that Threatens Native Plant and Animal Communities Impact onExistingAgricultural Practices No change No change Potential Impacts to Wildlife No adverse impacts No adverse impacts Potential Impacts by Fire Minimal risk Minimal risk Potential Impacts to Human Health Negligible Negligible Transfer ofGenetic Informationto No risk No risk Organisms with which it Cannot Interbreed

- Horizontal Gene Transferto Other Organisms Potential Effects ofGrowingEucalyptus on No negative impacts No negative impacts Soil Hydrology Potential Allelopathic Effects ofEucalyptus Minimal Minimal Risks to Threatened and Endangered No effect No effect Species Impacts on Unique Characteristics of No effect No effect ProtectedAreas

Page 7 of28Finding ofNo Significant Impact The analysis in the EA indicates thatthere will not be a significant impact, individually or cumulatively, on the quality ofthe human environment as a result ofthis proposed action. I agree with this conclusion andtherefore find that an EIS need not be prepared. This NEPA determination is based on the following context and intensity factors (40 CFR 1508.27):



### Context- The term "context" recognizes potentially affected resources, as well as the location and setting in which the environmental impact would occur. This action would be limited to the interstate movement and environmental release ofa GE Eucalyptus hybrid clone at 6 well dispersed confined field site locations in the States ofAlabama, Florida, Mississippi, and South Carolina encompassing a total of 14.7 acres and has limited potential to affect resources outside ofthe confined field test sites. Supplemental permit conditions 

### (see Appendices VI and VII of the EA) established for the permitwill effectively limit the reproductive capabilities and establishment ofthis GE 

### Eucalyptus outside the confined field trial locations and reduce the possibility ofunintended exposure from moving GE Eucalyptus trees to be negligible to nonexistent.



### Intensity- Intensity is a measure ofthe degree or severity ofan impact based upon the ten factors. The following factors were used as a basis for this decision:



### 1. Impacts that maybe both beneficialandadverse. According to the applicant, genetically engineered cold tolerant

### Eucalyptus would enable the production ofthis hardwood species for pulping and for biofuel applications in managed plantation forests in the southeastern U.S. APHIS issuance ofthe interstate movement and confined field release permitwould allow ArborGen to conductresearch to assess the efficacy ofthe introduced coldtolerance genes, genes to alter lignin biosynthesis in 

### Eucalyptus, genes to alter growth andto research mechanisms for altered fertility. The confined release ofthe trees at six well dispersed confined field site locations in the States ofAlabama, Florida, Mississippi, and South Carolinawill allow the applicant to obtain data

onperformance ofthe transgenic trees and the efficacy ofthe inserted genes in a wide variety ofenvironments. The establishment and growth ofthese small confined field tests will not have any impact on existing agricultural practices because they are solely for research purposes. Current agricultural practices will essentially remain unchanged. As identifiedbythe applicant, the field sites that are beingproposed under the permithave been used as forest tree plantations, pastures, or for forestry and agriculture research and are specifically designed for field testing crop plants orforest trees. As discussed in Section V ofthe EA, APHIS considers the possibility ofunintended exposure from moving GE Eucalyptus trees underthe proposed action to be negligible to non-existent. 

### 2. The degree to which theproposedaction affectspublic health orsafety. The proposed action to issue the APHIS interstate movement and confined field release permit should notpose an unnecessaryrisk to human health and therefore would have no significant impacts onhuman health. This GE 

### Eucalyptus hybrid is not used for food or feed purposes and does not contain any genetically engineered pesticides ortolerance to herbicides. As described in Section V ofthe EA and in the response to comments, the risk thatthese field trials will result in a higher incidence ofa fungal pathogen in the U.S. and thereby pose a risk to human health is considered to be negligible.

Page 8 of283. Unique characteristics of the geographic area such asproximity to historic or cultural
resources,parklands, prime farmlands, wetlands, wildandscenic rivers, orecologically critical areas. There are no unique characteristics ofgeographic area such as park lands, prime farm lands, wetlands, wild and scenic areas, or ecologically critical areas that would be significantly affected. As described in Section V and Appendix V ofthe EA, issuance ofthe permitto allow flowering ofthese additional constructs is not expected to impact unique characteristics ofprotected areas such as park lands, wetlands, wild and scenic areas, or ecologically critical areas. All the confined field test sites listed in the permit application

are either onprivately owned and managedplantation forests and agricultural farm lands or experimental research stations managed by academic institutions and industry. The standard agricultural and silvicultural practices for land preparation, planting, irrigation, and harvesting ofplants have been routinely used on these sites. Surveys conducted bythe applicant at each ofthese locations indicate thatthere are not any old growth forests or undisturbed natural areas in the immediate surroundings ofthe test sites. In addition, supplemental permit conditions (see Appendix VII ofthe EA) established for the permitwill effectively limit the reproductive capabilities and establishment ofthis GE Eucalyptus outside the confined field trial locations and reduce the possibility ofunintended exposure from moving GE Eucalyptus trees to be negligible to non-existent. 4. The degree to which the effects on the qualityof the human environmentare likely to be highly controversial. The effects onthe quality ofthe human environment are not highly controversial. Although there is some oppositionto APHIS issuing the interstate movement and confined field release permit, this action is nothighly controversial in terms ofsize, nature or effect. This action would be limitedto 6 well dispersed confined field site locations in the States of Alabama, Florida, Mississippi, and South Carolinaencompassing a total of 14.7 acres on individual research sites ranging from 0.5 up to 7.7 acres, depending onthe location with planting densityranging from 300

- 600 trees per acre. Duringthe public commentperiod, APHIS received comments opposingAPHIS' issuance ofthe permit. No new issues, alternatives or substantive new information were identified in any ofthe comments received by APHIS. APHIS has addressed substantive comments in the response to public comments document attached to this FONSI based on scientific evidence found in peer-reviewed, scholarly, and scientificjournals. 5. The degree to which thepossible effects on the human environmentare highly uncertain or involve unique or unknown risks. The effects ofthe proposed action to issue the APHIS interstate movement and confined field release permit are nothighly uncertain and do not involve unique orunknown risks. Based onthe analysis documented in the EA, the effects onthe human environment would
not be significant. APHIS has no evidence for any unknown risks ofthis GE hybrid plant species when released for confined field testing orfrom moving GE Eucalyptus trees. This GEEucalyptus hybrid has been previouslyreleased for confined field testing under APHIS issued notifications and permits in similar geographic locations without any known adverse effects. As described in Section V ofthe EA and response to comments, the issuance ofthe confined field release permitto allow the applicantto plant additional trees and to allow the
trees to flower at four ofthe six locations does notpresent any unforeseen risks. Since there is no increase in the number ofsites or acreage where trees will be allowed to reach maturity and flower, overthose already authorized by APHIS to flower underpermits 08-011-106rm, 08-014-101rm, 10-112-101rand 11-201-103rwith similar supplemental permit conditions,
Page 9 of28APHIS has concluded thatthere is no substantial greaterrisk ofloss ofconfinement and risk
to the environment from the proposed action. As presented in Appendix I ofthe EA, information that has been collected bythe applicant and providedto APHIS as part of ArborGen's annual reporting requirements under the existingpermits that allow flowering confirmedthat cold tolerant translines grown in these field tests did notproduce anypollen, thatthe applicant has not observed any volunteer seedlings in or around the test sites where
trees have flowered and produced seed capsules, and monthly field test monitoring observations have not identified any differences in diseases and insects or othernon-target organisms between the transgenic and non-transgenic trees in the field test. Based on the analysis and information provided in the EA and supportingpermit application, the new genes that are engineered into the trees should notpose any greater risk ofloss of confinement and risk to the environmentthan those that are currently authorized by APHIS for confined field release. The addition ofnew genes to increase cold tolerance, alter lignin and growth should not compromise the ability ofthese to remain confined field tests. Adherence to the supplemental permit conditions established for the permit will effectively limit anypotential adverse impacts to the human environment. As discussed in Section V of the EA, APHIS considers the possibility ofunintended exposure from moving GE Eucalyptus trees underthe proposed action to be negligible to non-existent. 6. The degree to which the action mayestablish aprecedent for future actions with significant effects orrepresents a decision inprinciple abouta future consideration. Theproposed action would not establish a precedent for future actions with significant effects orrepresent a decision in principle about a future decision. Similarto pastpermit applications reviewed and approved byAPHIS, the decision on whether or not to issue a permit for interstate movement and confined environmental release will be based upon information provided in the permit application. APHIS regulations at 7 CFRpart 340, regulate the introduction (importation, interstate movement, orrelease into the environment) ofcertain GE organisms and products. In accordance withthese regulations, when APHIS receives an application for apermit for environmental release ormovement, the application is evaluated to determine whetherthe environmental release ormovement, with appropriate conditions imposed, can be carried outwhile preventing the dissemination and establishment ofplantpests. The applicant has provided the information associated with this request in the permit application and APHIS now must make a determination to either grant or deny the permit. Eachpermit application that APHIS receives undergoes this independentreview to determine ifAPHIS should grant ordeny the individual permit. 

### 7. Whether the action is relatedto otheractions with individually insignificant but cumulativelysignificant impacts. No significant cumulative effects were identified through this assessment. As discussed in the cumulative effects analysis presented in Section V ofthe EA, APHIS has determined thatthere are no past, present, orreasonably foreseeable actions that would aggregate with effects ofthe proposed action to create cumulative impacts orreduce the long-term productivity or sustainability ofany ofthe resources (soil, water, ecosystem quality, biodiversity, etc.) associated with the release sites orthe ecosystem in which they are situated. No resources will be significantly impacted due to cumulative impacts resulting from the proposed action. 8. The degree to which the action mayadverselyaffectdistricts, sites, highways, structures, or objects listed in oreligible for listing in the NationalRegisterof Historic Places or may

cause loss or destruction of significantscientific, cultural, or historical resources.

Page 10 of28The issuance ofthe interstate movement and confined field release permitwould have no impact on districts, sites, highways, structures, or objects listed in or eligible for listing in the National Register ofHistoric Places, nor would they likely cause any loss ordestruction ofsignificant scientific, cultural, orhistorical resources. All the confined field test sites listed in the permit application are either on privately owned managed plantation forests and agricultural farm lands orexperimental research stations managed by academic institutions and industry. The standard agricultural and silvicultural practices for land preparation, planting, irrigation, and harvesting ofplants have been routinely used on these sites. Similar types ofagricultural and silvicultural practices would be used bythe applicant for managing
trees associated with the permit. In addition, supplemental permit conditions (see Appendix VII ofthe EA) established for the permit will effectively limit the reproductive capabilities and establishment ofthis GE Eucalyptus hybrid outside the confined field trial locations and reduce the possibility ofunintended exposure from moving GE Eucalyptus trees to be negligible to non-existent. 9. The degree to which the action may adversely affectan endangeredor threatenedspecies or its habitatthathas been determinedto be critical under the EndangeredSpeciesActof 1973. APHIS evaluated the potential for negative effects on federal threatened and endangered species as listed bythe U.S. Fish and Wildlife Service from the issuance ofthe interstate
movement and confined field release permit and determined thatthe movement and confined environmental release ofthis GE Eucalyptus hybrid would have no effect on federally listed threatened or endangered species or species proposed for listing, oron designated critical habitat orhabitatproposed for designation (see section on Threatened and Endangered Species in Section V and Appendix IV ofthe EA). 10. Whether the action threatens a violation of Federal, State, orlocal law or requirements imposed for theprotection of the environment. The proposed action would be in compliance with all federal, state, and local laws. The proposed action to issue the APHIS interstate movement and confined field release permit would be carried out in accordance with 7 CFRpart 340. This GE Eucalyptus hybrid is not used for food orfeed purposes and does not contain any genetically engineered pesticides

ortolerance to herbicides. There are no other Federal, state, or local permits that are needed priorto the implementation ofthis action.

NEPA Decision and Rationale I have carefully reviewed the EAprepared for this NEPA determination and the input from the public involvementprocess and have selected Alternative B-Issue the APHIS Permit. This alternative would fulfill APHIS' statutory mission and responsibilities to allow the safe development and use ofgenetically engineered organisms consistent withthe plantpest provisions ofthe PlantProtection Act.

As stated in the CEQ regulations, "the agency's preferred alternative is the alternative which the agency believes would fulfill its statutory mission and responsibilities, giving consideration to economic, environmental, technical and otherfactors." The preferred alternative has been selected for implementation based on consideration ofa number ofenvironmental, regulatory, and social factors. Based upon our evaluation and analysis, Alternative B is selected because (l) it allows APHIS to fulfill its statutory mission to protectAmerica's agriculture and environment using a dynamic and science-based regulatory framework that allows for the safe development and use ofgenetically engineered organisms; and (2) it allows APHIS to fulfill its regulatory
Page ll of28obligations. Therefore, it is my decision to implementthe preferred alternative as described in the EA. Based on all ofthe analysis and reasons above, I have determined thatthere would be

no significant impactto the quality ofthe human environment from the implementation ofthe chosen alternative (Alternative B) and therefore, no EIS needs to be prepared.

Michael C. Gregoire Deputy Administrator Biotechnology Regulatory Services Animal and Plant Health Inspection Services U.S. Department ofAgriculture

Page 12 of28 DateAttachment Finding ofno significant impact Response to comments APHIS No. 11-052-lOlrm

In a notice published in the Federal Register (77FR28 7123-7124, DocketNo. APHIS-2011- 0130), APHIS announced the availability ofan EA for public review and comment for a proposed controlled field release ofa genetically engineered clone ofa Eucalyptus hybrid. Comments on the environmental assessmentwere required to be received on orbefore March 12, 2012. APHIS received a total of246 comments during the 30-day commentperiod. All respondents expressed concerns about the permit or opposed granting the permit. Comment documents may be viewed at http://www.regulations.gov/#!searchResults;rpp=25;po=O;s=APHIS%25E2%2580%259320II% 25E2%2580%25930130.

Respondents opposingAPHIS' issuance ofthe permit came from nine non-governmental organizations (NGOs) or individuals apparently representing them, two State government entities, and individuals. The majority ofrespondents opposing the issuance ofthe permit came from one NGO, whose members submitted nearly identical form letters with some minor modifications. The NGO also submitted a petition bearing I,957 signatories that opposed the issuance ofthe permit. The majority ofrespondents opposed believe thatAPHIS failed to comprehensively analyze all relevant issues related to the proposed field trial, that an Environmental Impact Statement should have beenprepared and that cumulative impacts were not considered; thatthere is greatpotential for these Eucalyptus hybrids to become invasive or

cross breed with non-transgenic Eucalyptus trees; Eucalyptus could be a host for Cryptococcus neoformans gattii, a fungi that couldpotentially cause mycoses in humans and animals; that Eucalyptus plantations have an impact on hydrology and cause or exacerbate drought situations; Eucalyptus trees are known to be an incendiary risk and tend to bumvery hot during forest fires that are likely to occur in the drought prone southeastern U.S.; and could have significant impacts on biodiversity. APHIS' responses to these and all relevant comments submitted are as follows:

Eucalyptus trees are introduced organisms in theU.S. and aredocumented as invasive pests in California and Florida. Thecold tolerancetrait could vastly expand the range ofthis GE Eucalyptus tree-and hence enhance its abilityto invade native ecosystems. Experience in California and otherparts oftheworld has clearly demonstrated thatwhenEucalyptus escape, it is next to impossibleto eradicatethem. Courtdecisions on genetically engineered perennial organisms including GE bentgrass and GE alfalfa, demonstrate a growing legal foundation around thepotentialescape ofperennial GE organisms, even in field trials. The above comments were submitted in the form letter.

Response: As covered in the EA, the genes introduced to affect cold tolerance could make the engineeredEucalyptus more adapted to cold temperatures in the southern United States, butthis trait alone would not impart invasive or weediness characteristics to the engineeredplants. The trees could be considered weedy or invasive ifthey were to produce many seedlings that were readily spread away from the field test sites. Where the non-engineered hybrid Eucalyptus (EH1) has been grown in Brazil, on an estimated 400,000 acres planted over 15 years, there has been no indication that large numbers ofseedlings are beingproduced and are becoming invasive
Page 13 of28from the commercial plantations. ArborGen is required to report any unusual occurrences, such

as increased flowering or seed productionto APHIS during the field testing period.

TheEucalyptus being released is similarto the Veridian phytomining operation One commenter compared the release ofthe Eucalyptus underthis permitto the Veridian phytomining operation that released Yellowtuft (Alyssum murale andA. corsicum) in southwest Oregon. TheAlyssum escaped from cultivation and has been listed as a noxious weed in the State ofOregon. It is now a top priority for eradication in the State.

Response: The situation under which these GE hybrid Eucalyptus trees would be grown and tested is very different from the situationthat existed in Oregon underthe Veridian project where the Yellowtuft species were allowed to escape from cultivation.

Yellowtuftproduces masses ofyellow flowers and is a prolific seeder; and its height, from ten to thirty-six inches tall, makes it possible to harvest it by mechanical means. In addition, it is well adapted to the long, hot summers and relatively mild winters ofsouthwest Oregon (http://www.oregonencyclopedia.org/entry/view/alyssum/). As discussed in the EA, the soil conditions in the southeastern U.S. are not conducive to germination and growth ofEucalyptus seedlings. TheAlyssum species used in Oregon produces a large number ofseedlings. The serpentine soils where they are being detected are highly conducive to the growth ofthe species. Supplemental permit conditions (see Appendix VII ofthe EA) established for the permitwill effectively limit the reproductive capabilities and establishment ofthis GE Eucalyptus hybrid outside the confined field trial locations and reduce the possibility ofunintended exposure from moving GEEucalyptus trees to be negligible to non-existent. Underthe supplemental permit conditions established for the permitArborGen is required to monitor for any seedlings that are produced, remove them and report this to APHIS. The seedlings are easyto detect since they are distinct from othernative species and they are readily controlled bythe application ofEPA approved herbicides. In addition, the GE hybrid Eucalyptus trees that would be grown in these confined field tests are similarto those already authorized by APHIS to flower underpermits 08- 011-106rm, 08-014-101rm, 10-112-101r and 11-201-103rwith similar supplemental permit conditions. As presented in Appendix I ofthe EA, information that has been collected bythe applicant and providedto APHIS as partoftheir annual reporting requirements underthe existingpermits indicate no volunteer seedlings during the four years thatthey have been grown and allowed to flower.

State ofFlorida Official-flowering oftrees An official from the FloridaDepartmentofAgriculture and Consumer Services' Division of Plant Industry (DPI) expressed a concern that unlike otherpermits approved by DPI, these trees will be allowed to flower and that monitoring at the time offlowering and the subsequent information gathered atthat time will be crucial to later decisions concerningthe release ofthis cultivar on a larger scale.

Response: APHIS contactedthe official and discussed the misunderstanding about the permit with him. APHIS explained that GE Eucalyptus trees have been allowed to flower underpermits in Florida since 2008 and thatreproductive information associated with these permits has been collected bythe applicant and provided to APHIS as part oftheir annual reporting requirements. To date no volunteers have been reported bythe applicant. We also discussed the details ofthe supplemental permit conditions and explained thatthe permittee is required to scout for and
Page 14 of28report the presence ofany volunteer seedlings. We also explained that the individual research sites identified in the permit application would be similar in size to the confined field release permits thathave been issued by APHIS in Florida. None ofwhich have been over 20 acres in size. The official did not indicate that the agency had had any disagreement or concerns with the analysis provided in the EA.

State ofGeorgia, District Supervisor Soil & WaterDistrict- potential invasive species This individual was concerned that the Eucalyptus that are being field tested could be another invasive species. He states that "until enough controlled tests are run and the Americanpeople

can know for sure, USDA can control this invasive species, it should not be allowed to be massedplanted." He indicates that he is not aware ofhow many years this has been field tested and underwhat conditions.

Response: Thepermit application under consideration by APHIS is for the movement and confined field testing ofGE Eucalyptus trees on a limited area. APHIS is considering issuing a confined environmental release and interstate movementpermitwith supplemental permit conditions to allow the interstate movement, planting, field testing and flowering ofa GE Eucalyptus hybrid clone (Eucalyptus grandis x Eucalyptus urophylla) engineered to express various genes at 6 confined field site locations within the States ofAlabama, Florida, Mississippi, and South Carolina encompassing a total of 14.7 acres. ArborGen has indicated that the trees would be planted on individual research sites ranging from 0.5 up to 7.7 acres, depending on the location with planting density ranging from 300

- 600 trees per acre. The GE hybrid Eucalyptus trees that would be grown in these confined field tests are similarto those already authorized by APHIS to flower underpermits 08-011-106rm, 08-014-101rm, 1 0-112- 101r and 11-201-103rwith similar supplemental permit conditions. As presented in Appendix I ofthe EA, information thathas been collected bythe applicant and provided to APHIS as partof their annual reporting requirements under the existing permits have produced no volunteer seedlings during the four years that they have been grown and allowed to flower. As reported by the applicant, these cold tolerant translines grown in these field tests did not produce anypollen, the applicant has not observed any volunteer seedlings in or around the test sites where trees have flowered and produced seed capsules, and monthly field test monitoring observations have
not identified any differences in diseases and insects or other non-target organisms between the transgenic and non-transgenic trees in the field test. Based on the analysis and information provided in the EA and supportingpermit application, the new genes that are engineered into the
trees should notpose any greater risk ofloss ofconfinement and risk to the environmentthan those that are currently authorized by APHIS for confined field release. The addition ofnew genes to increase cold tolerance, alter lignin and growth should not compromise the ability of these to remain confined field tests. Adherence to the supplemental permit conditions established for the permit by the applicant will effectively limit any potential adverse impacts to the human environment. As discussed in Section V ofthe EA, APHIS considers the possibility ofunintended exposure from moving GE Eucalyptus trees underthe proposed action to be negligible to non-existent. Nothing in the comment indicates thatthere was any disagreement with the analysis provided in the EA.

Genetic engineering alters genomes in a random and unpredictablefashion One commenter cited a paperbyLatham (2006) which says that "Despite the fact that confidence in the safety and dependability ofcrop species rests significantly on their genetic integrity, the frequency oftransformation-induced mutations and their importance as potential
Page 15 of28biosafety hazards are poorly understood." The commenter goes on to state that "Releasing GE
trees will mean releasing trees that have never existed before with totally unknown qualities. This is damaging to biodiversity and since they are living organisms this pollution is uncontainable and has the potential to grow overtime."

Response: The engineeredEucalyptus trees in the proposed confined field tests have been observed overthe course ofnine years in the laboratory, greenhouse and in confined field tests to identify any unusual or unintended effects due to the transformationprocess. As confirmed in field tests conducted bythe applicant since 2006, onlythe traits for which the trees have been transformed have been expressed in these trees. In most cases any transformation-induced mutations would have been eliminated early in the transformation process as trees were regenerated in the laboratory and grown for field testing. The permit application under consideration byAPHIS is for the movement and confined field testing ofGE Eucalyptus trees in a limited area. APHIS is considering issuing a confined environmental release and interstate
movementpermit with supplemental permit conditions which includes specific monitoring and reporting requirements. In addition, APHIS would also conduct yearly inspections to look for unintended effects. The GE hybrid Eucalyptus trees that would be grown in these confined field
tests are similarto those already authorized by APHIS to flower underpermits 08-011-106rm, 08-014-101rm, 10-112-101r and 11-201-103rwith similar supplemental permit conditions. As presented in Appendix I ofthe EA, information thathas been collected bythe applicant and provided to APHIS as part oftheir annual reporting requirements under the existingpermits have
not identified any differences in diseases and insects orothernon-target organisms betweenthe transgenic and non-transgenic trees in the field test. Given the small size ofthe confined field
test sites identified in the permit application and the supplemental permitconditions that would be imposed onthe applicant and the fact that the permits can be revoked at any time byAPHIS and the trees removed ifany unintended effects are found, APHIS does not anticipate that issuance ofthe confined environmental release and interstate movement permit would impact biodiversity.

Excessive claims ofCBI and failure ofArborGen to reveal critical information A couple ofcommenters indicated thatthe designation ofthe majority oftransgenic modifications as CBI prevents any rational evaluation ofthe impact ofthose genes and modified Eucalyptus onthe environment, humans and animals. And because the locations are claimed as CBI it is impossible to create a creditable environmental assessment for the permit application. Also a commenter indicatedthat normally consideration would be limitedto only those species and critical habitat identified bythe United States Fish and Wildlife Service and/orNational Marine Fisheries Service likely to be found in the area ofthe field tests. The commenter believed that, withoutthis information, such consideration is impossible. There was a concern that "bystanders exposed to the CBI GM Eucalyptus pollen, dust from decaying leafdetritus and pollution ofsurface and ground water from pollutingplantproducts" would suffer adverse side effects.

Response: APHIS takes the use and designation ofCBI very seriously. Like all Federal agencies, APHIS is subjectto the Trade Secrets Act (18 U.S.C. § 1905) (TSA). The TSA prohibits officers and employees offederal agencies from publishing ordisclosing trade secrets and otherconfidential business information "to any extent not authorized by law." Information that is claimed as CBI by applicants is evaluated by APHIS to ensure thatthe claims are legitimate and APHIS evaluated ArborGen's claims for CBIprotection in this instance. It would
Page 16 of28be a clearviolation ofthe Act for APHIS to reveal Confidential Business Information to the public that legitimately falls under the Act. Federal employees can be fined and/or imprisoned for violating the TSA. Every gene construct component in the permit submission is evaluated for any plantpestrisk as well as any potential effects on the environment. None ofthe genes that have been engineered into these trees (to affect coldtolerance, lignin formation, growth and pollen formation) are expected to have any effects on "bystanders exposed" to the Eucalyptus
trees orpotentiallypollute surface and ground water. No pollen is beingproduced bythe trees in the field tests due to the presence ofthe barnase gene. In addition a thorough analysis for threatened and endangered species and critical habitat are made for every location and ifAPHIS determines that there could be a species present at the field test site, appropriate contacts are made with the USFWS. A thorough discussion ofpotential impacts ofthe proposed action on human health, waterresources and threatened and endangered species are presented in Section V and Appendix IV ofthe EA.

Concern thatthe CBFgenes could have pleiotropic effects and affect allelopathy One commenter indicated thatthe C-Repeat Binding Factor (CBF) genes when over expressed display improvement in cold tolerance, waterretention, higher oil gland density, wax deposition and over expression ofanthocyaninpigments (Navarro et al., 2011). The gene network influenced by CBF gene modification produced highlypleiotropic. The commenterwas concernedthatthe production ofallelochemicals in the modified trees did not appearto have been investigated.

Response: The reference cited in the comment (Navarro et al., 2011) does indicate thatthe gene
can have multiple phenotypic effects. As a transcription factor this is not unexpected. As the publication indicates, the multiple changes observed are consistent with the correlation between coldtolerance and drought tolerance and reinforce the hypothesis that, bytriggering metabolic changes directed towards limiting cell dehydration, the DREB transcription factors are partly responsible for these observed correlations. As covered in the EA, allelopathy tends to be an inexact science and many studies in allelopathy are inconclusive and difficult to interpret due to potential interactions with other aspects ofthe environment. Even though the Eucalyptus under this permit could demonstrate allelopathic properties, the presence ofany allelochemicals is not going to make the Eucalyptus planted under these permits more invasive orpresent aplantpest risk. Since all these field tests are confined and limited in size, any allelopathic effects should be small. As a standard silvicultural practice, herbicides will also be used within the field test sites and any oftheir effects on understory vegetation will be as severe ormore severe than any allelopathic effects.

Pollen spread One commenterwas concerned aboutpollen spread from the trees and cites anAustralian study that found thatremnantpopulations ofEucalyptus were connected bypollen dispersal to pollen

sources up to 1.94 kilometers away (Sampson and Byrne, 2008). The concern is thatthe distance between these GE Eucalyptus and otherEucalyptus that are 1000 meters away would lead to the spread ofthe GE trees.
Response: As indicated in the EA, the observations made to date by the applicant on APHIS approved confined field tests thathave allowed GE Eucalyptus hybrid trees to flower indicate thatthe flowering ofthe hybrid does not overlap the flowering ofotherEucalyptus in Florida including one oftheparents ofthe hybrid, i.e. E. grandis. Also given thatthe transgenic hybrid
Page 17 of28is not producing any pollen, there is little ifany potential for outcrossing ofthe hybrid to other species. One ofthe requirements ofthe existingpermits and proposedpermit is that the applicant mustreport any overlap in flowering ifit occurs with otherEucalyptus tree species in the area. Ifany overlap in flowering were to occur, the only seeds that could be formed from crossing would be on trees within the testplots whose female flowers could be susceptible to incomingpollen. Monitoring ofany seeds and seedling formation in the permitted plots bythe applicant would show ifany seedlings were being formed as hybrids from incomingpollen

sources. In accordance with the supplemental permit conditions, monitoring for and removal of volunteers within lOOm from the edge oftransgenictestplotby the applicant would effectively eliminate any seedlingsthat may be produced and thereby effectively reduce the possibilitythat any Eucalyptus hybrid trees would become established.

Concerns about Cryptococcus One commenter and the form letter was concerned about the spread ofCryptococcus neoformans gattii bythe transgenic Eucalyptus trees indicatingthatthe spread ofthis "deadly yeast" is mainly brought aboutbybirds and their feces. The commenter appears to take issue with the
statement in the EA that the scale ofthe field tests is miniscule comparedto the vast expanses of native trees that could potentially harbor the pathogen and says that "There was no vast expanse ofnative species homes for the toxic yeast in the peer reviewed scientific publications."

Response: As covered in the EA, it is well documented in the literature that manytrees, other than Eucalyptus, can harbor Cryptococcus in North America. Accordingto the APHIS report cited in the EA (APHIS, 2004): "Inthe endemic area ofVancouver Island, trees that have tested positive for C. gattii include alder, bitter cherry, cedar, Douglas Fir, and Garry Oak." Many years after its first recovery from clinical specimens, C. gattii was isolated from the environment for the first time by sampling ofEucalyptus camaldulensis trees (Ellis and Pfeiffer, 1990). This
tree species was initially thought to bethe exclusive natural habitat ofC. gattii both in its native Australia and around the world given that the tree has been exported extensively. Howevernow it is clearthat more than 50 tree species have been reported as harboring the fungus (Springer and Chaturvedi, 2010). Although Eucalyptus is present in many ofthe areas known to have C. gattii-associated cryptococcosis, the actual isolation ofC. gattii from Eucalyptus trees is rare outside Australia, despite extensive sampling. ImportedEucalyptus has not been associated with the environmental presence ofC. gattii in Spain, central Africa, orCanada; and mostEucalyptus
trees tested in PapuaNew Guinea, Egypt, and Italy were negative for C. gattii. When C. gattii

was first associated with cryptococcosis, early environmental surveys for C. gattii in imported Eucalyptus spp. rarely included other local tree species for testing. Although understandable, this was unfortunate because C. gattii is now known to have extensive associations with other
tree species (Springer and Chaturvedi, 2010).

C. gattii is clearly not specific to Eucalyptus trees in North America and other areas ofthe world and is most likelybeing spread by birds and other animals (Springer and Chaturvedi, 2010) throughout temperate tree species since very little Eucalyptus is present in these areas. Therefore, the planting GE Eucalyptus trees in confined field tests in southeast would not be expectedto increase the incidence ofthe disease any more than planting any othertree species, since many species can hostthe fungus. Also it is importantto note that the recent literature has begun to sort outthe difference between C. neoformans and C. gattii. Until recently, these two fungi were classified as varieties ofthe same species but are now considered two different spectes. The pathogen C. gattii is believed to be clinically more virulent than C. neoformans and
Page 18 of28causes more severe symptoms ofcryptococcosis. For a review see (Chaturvedi and Chaturvedi, 2011).

Concerns aboutthe alteration in susceptibility to diseases orinsects Commenters were concernedthatthe susceptibility ofthe trees to insects and disease were not adequately addressed in the EA, particularlythe trees that have altered lignin.

Response: The GE hybrid Eucalyptus trees that would be grown in these confined field tests are similarto those already authorized by APHIS to flower underpermits 08-011-106rm, 08-014- 101rm, 10-112-101r and 11-201-103rwith similar supplemental permit conditions. As presented in Appendix I ofthe EA, information that has been collected bythe applicant and provided to APHIS as part oftheir annual reporting requirements under the existingpermits have not identified any differences in diseases and insects or other non-target organisms between the transgenic and non-transgenic trees in the field test. Growth measurements have indicated that
trees had normal to a moderately reduced growth phenotype. The trees have been visually inspected on a monthly basis for the presence ofany insect and disease damage. As reported in the applicant's annual reports, the degree ofinsect and disease infection has been minimal. Adherence to the supplemental permit conditions established for the permit bythe applicant will effectively limit any potential adverse impacts to the human environment. However, ifduring the tests there is evidence ofincrease disease or insect susceptibility, the applicant is required to report this to APHIS. The permittee is required to report any such unanticipated effects (including excessive mortality ormorbidity) to APHIS under the terms ofthe permit- see 7 CFR 340.4(f)(1O)(ii). An assessment ofdisease and insect susceptibilities is one ofthe primary

reasons the applicant is conducting the confined field testing ofthe trees.

Kanamycin resistance One commenter raised an issue aboutthe risks from the kanamycin resistance gene. The
commenter indicated thatkanamycin is still in clinical use and also kanamycin resistance crossreacts with new antibiotics.

Response: In 1998, FDA developed draft guidance on the use ofantibiotic resistance markers in transgenic plants http://www.fda.gov/food/guidancecomplianceregulatoryinformationlguidancedocuments/biotech nology/ucm096135.htm. In this document, FDA concluded that the likelihood ofthe transfer of antibiotic resistance is remote. In this draft guidance the FDA stated, "FDA acknowledges that the likelihood oftransfer ofan antibiotic resistance marker from plants to microorganisms in the gut or in the environment is remote and that, such transfer, ifany, would likely be insignificant when comparedto transfer between microorganisms, and in most cases, would not add to existing levels ofresistance in bacterial populations in any meaningful way." Further, to evaluate safety, the European Food SafetyAuthority (EFSA) reviewed the antibiotic selection markers used in genetically engineered plants http://www.efsa.europa.eu/enlefsajoumal/doc/opinion gmo 05 en1.pdf. In this 2004 document, various antibiotic resistance genes were assigned into groups based on the criteria oftherapeutic

use in humans and in animals and presence in the environment; Group I is composed of kanamycin and hygromycin resistance genes. The opinion states that because ofthe frequency ofhorizontal transfer plants to other organisms is veryrare, previous existence in the environment and the history ofuse ofthe kanamycin resistance, thatthere is no rationale for restricting Group I antibiotics. In addition, the EFSA 2004 document states: Kanamycin is
Page 19 of28rarely used today because ofits considerable side effects. Only under conditions ofmultiple mycobacterial resistance to otherdrugs is kanamycin still used as a reserve tuberculostatic agent. Forthe same reason as kanamycin, neomycin, which is poorly absorbed orally, is also rarely used intravenously/intramuscularly to treat infections. These antibiotics are rarely administered orally, which minimizes the selective pressure for antibiotic resistance in the gut. Theiruse in the
treatment ofhumans has been superseded by more effective aminoglycoside antibiotics that are not substrates for APH(3')-II. The antibiotics are rarely used in agriculture or aquaculture and thereby do notprovide selective pressure for a possible transfer ofthe resistance genes from genetically modified plants to soil microorganisms.

Gene for altered fertility Commenters were concerned aboutthe "dangers ofthe barnase gene" and stated that it is a known cytotoxic protein that breaks down RNA. "Because they are from a soil bacterium, and unrelated to any mammalian RNAses, they are not susceptible to eukaryotic 3 RNAse inhibitors. Consequently, they are highly toxic, and are actually being engineered currently as a means of killing cancercells. Although the barnase is being used to preventpollen formation, this is not 100 [%]effective, and many beneficial insects and other wild-life could well be affected."

Response: As indicated in the EA, the transgenic Eucalyptus hybrid identified in the permit application utilizes barnase to produce male sterile plants, reducing the potential for gene flow into the environment. Male sterility is achieved through the localized production ofbarnase in pollenproducing cells. Barnase is a ribonuclease, an enzyme that degrades RNA, thereby regulating protein synthesis. Ribonucleases are highly ubiquitous molecules found in all living cells (Worrall and Luisi, 2007). The transgenic Eucalyptus trees identified in the permit application and otherpermits already authorized byAPHIS were engineered using barnase gene mutants with reduced toxicity to minimize tissue damage (Rottmann et al., 2008). The barnase production is controlled by atissue specific promoter. As with all genes, the gene encoding barnase is present in every cell ofthe transgenic plant, however, the promoter acts like an "on" switch that controls when and how stronglythe gene is expressed. The PrMC2 promoter used by ArborGen, originally identified in pine (Pinus radiata), restricts barnase productionto the tapetum, a small layer ofcells within the male floral organ, or anther (Walden et al., 1999; Hofig
et al., 2003; Rottmann, et al., 2008). Within the anther, the tapetum surrounds developing pollen grains. Mariani, et al. (1990) developed transgenic tobacco plants using the barnase gene also controlled by a tapetum-specific promoter (Tap29). They observedthat in transgenic plants tapetal cells senesced early in their development, preventing pollen formation. All otherfloral organs, including the anthers, formed normallyproving tapetal cells which could be eliminated without affecting flower development. Similarresults were observed using PrMC2 in Eucalyptus
trees (Rottmann et al., 2008). After several years ofresearch, no pollen has been observed in transgenic Eucalyptustrees as reported in the permit application and in annual reports submitted by ArborGen up through 2011 (see Appendix I ofthe EA). As with other ribonucleases, barnase degrades quickly afterthe destruction oftapetal cells, and does not accumulate within the plant tissues (Mariani et al., 1990); http://www.fda.gov/Food/Biotechnology/Submissions /ucml61162.htm). Therefore barnase is onlyproduced for a short period oftime during floral development, is limited to a discrete cell layer within the anthers ofthe flower, and is rapidly degraded (Mariani et al., 1990; Rottmann et al., 2008; Hofig et al., 2003). In previous studies using transgenic radicchio containingthe barnase gene, researchers were unable to detect accumulation ofbarnase within floral tissue (http://www.fda.gov/Food/Biotechnology/Submissions/ucml61162.htm). Therefore it is highly
Page 20 of28unlikely that consumption of, orexposure to, transgenic Eucalyptus could contain enough bamase to cause mammalian orinsecttoxicity. Directexposure oforgan tissue, is also highly unlikely to occur in nature. In addition the FDA has previously reported that consumption of bamase would likely degrade quickly during digestion further reducing the risk ofbamase exposure (http://www.fda.gov/Food/Biotechnology/Submissions/ucm161162.htm).

With regard to toxicity ofbamase to bees and otherpollinating insects, there is no clear evidence indicatingpollinators would be adversely affected by ingestion ofbamase. Combined with the facts thatbamase is only produced during the shortperiod oftapetum formation and is quickly degraded, as well as the lack ofpollen produced by the Eucalyptus hybrids, it can be concluded the tissue specific production ofbarnase is unlikely to adversely affect pollinators or other insects. APHIS has made a determination ofnonregulated status ofthree plant species, com (petitions 95-288-01p, 97-342-01p, 98-349-01p), rapeseed (petitions 98-278-01p, 01-206-01p) and chicory (petition 97-148-01p) since 1995 that have included the barnase gene. APHIS is

unaware ofany reported cases ofmammalian or insect toxicityresulting from barnase consumption orexposure occurring within the past 17 years.

Based on these findings, no hazard orexposure associated withthe use ofbarnase has been identified, therefore APHIS concludes thatthere is no foreseeable risk associated with the use of bamase in transgenic Eucalyptus trees in the proposed confined field tests.

Mode oftransformation and hazards ofhorizontal gene transfer One commenter expressed concerns about horizontal gene transfer between the Agrobacterium used to transform the plants to wild-typeAgrobacterium in the soil, and to otherbacteria and fungi in the soil, citing a publication by (Knight, 2010).

Response: It is highly unlikely that any Agrobacterium remains on the plants that are generated in the laboratory since antibiotics are used to remove them priorto release into soil. During the Eucalyptus transformation process, the transformed tissue is kept on selection medium containing antibiotics to kill any residual Agrobacterium for at least four months, following which individual events are transferred to medium without selection. As indicated bythe applicant, Agrobacterium growth has neverbeen observed in the cultures atthis stage. ArborGen has also collected vegetative tissue from selecttissue cultures as well as field-grown transgenic plants and analyzed for the VirD gene which would indicate the presence of Agrobacterium. The lack ofdetectable VirD signal in samples indicates that no live or dormant Agrobacterium remains in the transgenic plants.

High flammability ofEucalyptus Several commenters including the form letter noted the high flammability ofEucalyptus which allows Eucalyptus plantations to be susceptible to initiation orrapid spread ofwildfire.

Response: As indicated in the EA, it is not clearthatEucalyptus plantations present a greater fire hazard than do the pine plantations which are common in the Southeastern USA. To assess the risk ofwildfire in live and dead material, (Nufiez Regueira et al., 2002a; Nufiez Regueira et al., 2002b) in northern Spain used calorimetry analysis combined with multidimensional
assessment ofclimatic and physical characteristics in plantations ofa Pinus species (P. pinaster) and ofa Eucalyptus species (E. globulus). Forpine, the fire risk index for live biomass varied from extremelyhigh in July to little risk in some winter and spring months. Pine residues,
Page 21 of28however, varied between no risk and extremely high risk. ForEucalyptus, fire risk index for live biomass varied from a middle risk in July to little risk in fall and winter. Dead matter of Eucalyptus varied between a middle riskto extremely high risk in August. Ganteaume et al. (2009) undertook avariety ofassays and comparisons offuel bed flammability and firebrand capacity (the material that can ignite the fuel bed). Pinus litter flammability was higher than that ofEucalyptus, which in tum was higherthan that ofa hardwood species (Ulex). While differences between southeastern pines and the proposed Eucalyptus species and these analyses may be important, basic similarities within the genera are likely relevant. For example, studies ofdifferent Eucalyptus species have shown that despite the differing climactic specificities ofE. globulus and E. nitens and differences ofvolatile content and solid mass, the total caloric content ofwastes after pulping are similar (Perez et al., 2006). Due to the small size ofthese field trials, and the fact thatthey are isolated from one another by many miles, there is no reason to believe that theypose any more ofa fire risk than do other forest tree plantings

Wildfires in California and Australia Some ofthe comments and the form letter cited two large wildfires in arid areas ofAustralia and California involving Eucalyptus. These fires were mentioned as examples ofdeleterious consequences ofEucalyptusplantings.

Response: These fires occurred in areas ofextensive plantings ofEucalyptus, but it is likelythat many factors are important for the frequency and season ofoccurrence offires, notjustthe species oftrees within a burned area. Onepost-fire report on the Oakland, California fire noted that extended drought conditions and freeze-killed Eucalyptus contributed large amounts of residue for the fire. However, a wide variety ofgrasses, brush, trees, and houses combined to provide fuel that sustainedthis fire, and the fire had not initiated in the Eucalyptus stands (FEMA, 1991). As noted, one ofthemost importantrisk factors for Eucalyptusproduction is the presence ofaccumulating fuels within a stand (Nufiez Regueira, et al., 2002a). The duration of these field trials will be limited (no more than seven to nine years). ArborGen has seen little accumulation in field trials extending through five years (Les Pearson, ArborGen, personal communication) and consequently little to no accumulating fuel is expected to arise within the proposed confined field trials.

Failureto ConsidertheEnvironmentalEffects ofEucalyptusgrandisX Eucalyptus urophylla Pollen on Bee Populations One commenter was concerned thatthe EA did not considerthe effect ofpollen on bee populations. There was a concern that when the flower pollen becomes genetically modified or sterile, the bees will potentially go malnourished and die ofillness due to the lack ofnutrients and the interruption ofthe digestive capacity ofwhatthey feed on through the summer and over the winter hibernation process. He also indicates that there already exist sufficient acreage and plotsto capture and testpollen from GMO Eucalyptus and to monitor its effect on bee populations in proximityto them. And ifthis process is shown to have no deleterious effects on existing bee populations, over a series ofyears should any proposal to expandthe range ofGMO Eucalyptus be considered.

Response: It is importantto note thatthe trees are not producing anypollen, therefore the studies that are proposed by the commentercannot be performed. Also, as discussed above and within the EA, there is no reason to believe that the products ofthe introduced genes would have any negative effects on insects such as bees. ArborGen has observed many bees visiting the
Page 22 of28Eucalyptus flowers in the field test because the flowers are very attractive to bees and they feed

onthe nectarproduced by the flowers. As noted above no products (including barnase) produced by the inserted genes would be expected to affect bees. No dead bees have been observed in the vicinity ofthe field tests.
UnacceptableEnvironmentalRisks: WaterResources Several comments including the form letter noted the potential impacts ofEucalyptus on hydrology. The comments centered on concerns thattree plantations have been documented to deplete ground water and cause or exacerbate drought situations. They also citedthe information that the Forest Service provided upon APHIS' request as an appendix to the EA to indicate that the risk is unacceptable.

Response: The proposed action would be limited to 6 well dispersed confined field site locations in the States ofAlabama, Florida, Mississippi, and South Carolina encompassing a total of 14.7 acres on individual research sites ranging from 0.5 up to 7.7 acres. ArborGen has supplied data to APHIS indicatingthe maximum size ofeach ofthe plantings at each site, the individual watersheds where the plantings occur, the area ofthe watershed, how much ofthe watershed will be occupied bythe field tests, the location ofthe closestprimary and secondary streams, and the location ofany critical habitat for Federally listed threatened and endangered species within the watershed. The data provided by ArborGen show that none ofthe potential planting sites identified in the permit application occupy more than 0.005% ofany given watershed.

The potential impacts by Eucalyptus on hydrology relate to the scale ofthe plantings. The proposed confined field trial plantings are very small in terms ofscale in forestry practices and watersheds. Individual forest plantings in the southern U.S. range in size but can typically be up to 120 acres at a single site in a given year (SFI standards: http://www.sfiprogram.org/files/pdf/sfi requirements 2010-2014.pdf). A 20 acre planting is considered small in these terms. The issue ofhydrology is thoroughly addressed in Section V of the EA. As noted in the section under Potential Effects ofGrowing Eucalyptus on Soil Hydrology in the EA, the Forest Service has pointed outthatthe significance ofthe impact on groundwater and stream flow will depend greatly on the area extent, size, and spatial distribution ofthe plantations. For example, a few small (less than 10 hectares, i.e. approximately 25 acres) and well-dispersedplantations may only have very localized impacts and negligible impacts at the watershed scale. Based on the very small footprint ofthese proposed confined field tests and the weight ofevidence, APHIS concludes thatthe impacts ofthese field trials on hydrology will be negligible.

Unacceptableenvironmental risks: Native Flora and Fauna One commenternoted a commentmade on aprevious EA for GE Eucalyptus permits. The
commenter states that "The Georgia Department ofNatural Resources, Wildlife Resources Division has recommended that ArborGen's permit requests be denied because the Department has "serious concerns aboutpotential impacts on hydrology, soil chemistry, native biodiversity and ecosystem functions regardless ofwhetherthis nonnative hybrid turns outto be invasive in a plantation setting." They further note that Eucalyptus plantations "will be extremely inhospitable environments for native flora and fauna." Moreover, noting the high wateruse for Eucalyptus plantations, the Department expressed concern regarding the increasedpotential for significant impacts on waterresources and aquatic communities.
Page 23 of28Response: The potential impacts ofthe proposed action on native flora and fauna is thoroughly analyzed in Section V ofthe EA and discussed in several ofthe responses above.

The GE hybrid Eucalyptus trees that would be grown in these confined field tests are similarto those already authorized by APHIS to flower underpermits 08-011-106rm, 08-014-101rm, 10- 112-101r and 11-201-103r with similar supplemental permit conditions. As presented in Appendix I ofthe EA, information that has been collected bythe applicant and provided to APHIS as part oftheir annual reporting requirements under the existing permits have produced

no volunteer seedlings during the four years that they have been grown and allowed to flower. As reported bythe applicant, these cold tolerant translines grown in these field tests did not produce any pollen, the applicant has not observed any volunteer seedlings in oraround the test sites where trees have flowered and produced seed capsules, and monthly field test monitoring observations have not identified any differences in diseases and insects orothernon-target organisms between the transgenic and non-transgenic trees in the field test. Based onthe analysis and information provided in the EA and supportingpermit application, the new genes that are engineered into the trees should not pose any greaterrisk ofloss ofconfinement and risk to the environmentthan those that are currently authorized by APHIS for confined field release. The addition ofnew genes to increase cold tolerance, alter lignin and growth should not compromise the ability ofthese to remain confined field tests. Adherence to the supplemental permitconditions established for the permitbythe applicant will effectively limit any potential adverse impacts to the human environment. As discussed in Section V ofthe EA, APHIS considers the possibility ofunintended exposure from moving GE Eucalyptus trees underthe proposed action to be negligible to non-existent.

Failureto Assess CumulativeEffects ofall existing ArborGen permits One commenter expressed the concernthat since there are now six active permits underwhich ArborGen is authorized by APHIS to grow GE Eucalyptus which include 32 unique locations within 7 States where up to atotal of654.7 acres ofGE Eucalyptus could be planted, thatthe cumulative impacts ofthis proposal are substantial and significant and should be considered.

Response: A cumulative impacts analysis is provided in Section V ofthe EA. As identified in the permit application, GE Eucalyptus trees with the new constructs would be released at 6 locations. The new permit application does not expand the number oftrees ortotal acreage that could be planted at any ofthe confined release sites currently approved byAPHIS. As described in the EA and otherEAs prepared by APHIS for authorizing ArborGen permits (http://www.aphis.usda.gov/brs/aphisdocs/06 32511lr ea.pdfand http://www.aphis.usda.gov/brs/aphisdocs/08 01410lr ea.pdf), the 32 field test sites are widely dispersed across the southeastern US. The sites are 900 miles apart from the farthermost east and westplantings. They average approximately 50 miles apart. The closestplantings have at least a mile separation between them, no individual planting is greater than 20 acres in size and there is no reason to assume thatthere will be any interaction between these distant sites. The small size ofthe field plots and the distance betweentest sites spread overthousands ofmiles indicate no cumulative effects would result from APHIS issuing the proposed field release permits. Therefore the onlypast, present, and reasonably foreseeable actions associated with the locations for the proposed releases under permit are those related to agricultural orforest tree production. Based onthe analysis provided in the EA, APHIS has determined thatthere are no past, present, orreasonably foreseeable actions that would aggregate with effects ofthe proposed
Page 24 of28action to create cumulative impacts orreduce the long-term productivity or sustainability ofany ofthe resources (soil, water, ecosystem quality, biodiversity, etc.) associated with the release sites orthe ecosystem in which they are situated. No resources will be significantly impacted due to cumulative impacts resulting from the proposed action.

Failureto Consider a ReasonableRange ofAlternatives One commenter said: "While the DEA contains only two alternatives, it actually only has one: the proposal contained in Arborgen's application. The other alternative is the "no action" alternative, which is to deny the application. Ittherefore fails to include a range ofalternatives. The commenter suggests two additional alternatives should be analyzed in the DEA:

• One which excludes the change in conditions, the allowed flowering, ofgenetically engineeredEucalyptus trees authorized under permits 09-070-101rm and 11-041-101rm, and

• One which both excludes the change in conditions, the allowed flowering, ofgenetically engineered Eucalyptus trees authorized underpermits 09-070-101rm and 11-041-101rm, but also does not allow the flowering ofgenetically engineered Eucalyptus trees at the 6

new sites.

Response: APHIS has prepared this EA in response to confined environmental release permit applications (APHIS Number 11-052-101rm) received from ArborGen LLC. This permit application was submitted in accordance with APHIS' regulations in 7 CFRpart 340. In accordance with these regulations when APHIS receives an application for a permit for environmental release, the application is evaluatedto determine whetherthe environmental release, with appropriate conditions imposed, can be carried outwhile preventingthe dissemination and establishment ofplantpests. The receipt ofa permit application to introduce a genetically engineered organism requires a response from the Administrator:

Administrative action on applications. Afterreceipt and review by APHIS ofthe application and the data submitted pursuantto paragraph (a) ofthis section, including any additional information requested byAPHIS, a permit shall be granted or denied (7 CFR 340.4(e)).

The applicanthas providedthe information associated withthis request in the permit application and APHIS now mustmake a determination to either grant or deny the permit. Since APHIS decision is to either grant or deny the permit application, APHIS analysis ofthe two alternatives (No Action- Deny the permit and Preferred Alternative- Issue the APHIS Permit) identified in the EA are reasonable. The EA analyzes the potential cumulative effects ofthe proposed field
test locations and has shownthatthe issuance ofthe permits will not have a significant impact on the human environment. Further analysis on a site by site basis orover a more restricted space

ortime will notprovide any additional information oranalysis that would be necessary to make

an informed NEPA decision onthe issuance ofthese permits.

The alternatives suggested by the commenterhave been analyzed in the EA as eitherthe No Action Alternative orthe Preferred Alternative. As clearly stated in Section V ofthe EA, GE Eucalyptus trees grown underthe confined field release permits 09-070-101rm and 11-041- 101rm would notbe allowed to flower underthe No Action Alternative. An additional alternative that analyzes the potential impacts ofnot allowing GEEucalyptus trees to flower at Page 25 of28any ofthe proposed 6 confined release sites would notprovide any additional informationto the decision makerthan what is already provided under the Preferred Alternative. In effect, the supplemental permit conditions under the Preferred Alternative will have the same impact on the environment as would an alternative that does not allow the trees to flower. As described in the EA, the supplemental permit conditions (see Appendix VII ofthe EA) established for the permit will effectively limitthe reproductive capabilities and establishment ofthis GE Eucalyptus outside the confined field trial locations.



### Failure to prepare an EA or EISon permits 09-070-lOlrm and 11-041-lOlrm A commenter said: "Becausepermits 09-070-101rm and 11-041-101rm and all previous permits

were approved without completing either an EIS or an EA, there has notbeen an environmental
assessment ofany sort on the environmental effects ofintroducingEucalyptusgrandisX Eucalyptus urophylla. This includes, but is not limited to critical habitat analysis under ESA, effects on hydrology and effects ofpollen on bee populations, as referenced in previous sections ofthis document. APHIS did notprovide public notice or an opportunity for public comments or request an ESA determination from FWS. Therefore the environmental effects ofall ofthese permits mustbe considered in any environmental analysis, immediate, cumulative orotherwise."



### Response: This comment is outside the scope ofthis EA. APHIS has prepared this EA specificallyto evaluate and determine ifthere are any potentially significant impacts to the human environment in response to a confined environmental release and interstate movement permit application (11-052-101rm) received from ArborGen to authorize the planting of genetically engineered (GE) Eucalyptus hybrid clone (Eucalyptus grandis x Eucalyptus urophylla) to supportproposed field research studies on six research sites in Alabama, Florida, Mississippi, and South Carolina. A discussion ofpotential impacts associated withpermits authorized by APHIS for GE Eucalyptus is presented in the cumulative impacts analysis provided in Section V ofthe EA and the "cumulative effects" response above.

All regulatory actions taken by APHIS, including the authorization ofpermits 09-070-101rm and 11-041-101rm, are conducted in accordance with: (1) The National Environmental PolicyAct of 1969 (NEPA), as amended (42 U.S.C. 

### §4321 et seq.), (2) regulations ofthe Council on Environmental Quality for implementingthe procedural provisions ofNEPA (40 CFRparts 1500-1508), (3) USDA regulations implementingNEPA (7 CFRpart 1b), and (4) APHIS' NEPA Implementing Procedures (7 CFRpart 372).



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