Risk Assessment and Risk Assessment and Management in Transgenic Management in Transgenic

CropsCrops

P. BalasubramanianP. BalasubramanianDirectorDirector

Centre for Plant Molecular BiologyCentre for Plant Molecular BiologyTamil Nadu Agricultural UniversityTamil Nadu Agricultural University

Coimbatore 641 003Coimbatore 641 003

Status of Development Status of Development of Bt of Bt BrinjalBrinjal at TNAUat TNAU

Bt Bt BrinjalBrinjal ProgrammeProgrammeNetwork Network programmeprogramme involving involving •• TNAU, Coimbatore, TNAU, Coimbatore, •• University of Agricultural Sciences, University of Agricultural Sciences, DharwadDharwad, , •• Indian Vegetables Research Institute, Indian Vegetables Research Institute, VaranasiVaranasi and and •• MahycoMahyco, , JalnaJalna

Project funded by Project funded by •• Agricultural Biotechnology Support Agricultural Biotechnology Support ProgrammeProgramme II of II of

USAID and USAID and •• Department of Biotechnology, Government of IndiaDepartment of Biotechnology, Government of India

MahycoMahyco provides the Bt geneprovides the Bt genePublic sector institutes develop Bt Public sector institutes develop Bt brinjalbrinjal varietiesvarieties

ObjectiveObjective

Development of Bt Development of Bt brinjalbrinjal varieties varieties expressing resistance to fruit and expressing resistance to fruit and shoot borershoot borerDevelopment of Bt Development of Bt brinjalbrinjal lines in lines in local elite genetic background local elite genetic background keeping in view the local consumer keeping in view the local consumer preferencepreferenceFacilitating resourceFacilitating resource--poor farmers to poor farmers to save seeds for their future usesave seeds for their future use

ProgrammeProgrammeMahycoMahyco brinjalbrinjal event EE1 event EE1 expressing expressing cryIAccryIAc used as donor used as donor to to introgressintrogress the gene into local the gene into local elite backgrounds through backelite backgrounds through back--cross breeding cross breeding programmeprogramme

With a view to developing Bt With a view to developing Bt versionsversions•• Co2, Co2,

•• MDU1, MDU1,

•• PLR1 and PLR1 and

•• KKM1 KKM1

are used as recurrent parentsare used as recurrent parents

Progress and future Progress and future programmeprogrammeBCBC22FF11 progenies are progenies are being evaluated in being evaluated in transgenic greenhouse transgenic greenhouse BCBC33FF11 will be produced will be produced Superior plant types Superior plant types will be identified in will be identified in BCBC33FF2 2 andand BCBC33FF3 3 generationsgenerationsBioassay on Bioassay on homozygous homozygous transgenic linestransgenic linesField testingField testing

Co2 PLR1

MDU1KKM1

Bt eggplant research at TNAUBt eggplant research at TNAU

Resistance to Innovations Related to FoodResistance to Innovations Related to FoodPasteurizationPasteurization

Change is InevitableProgress is Optional

History of Technology AdoptionHistory of Technology Adoption

Resistance to Innovations Related to FoodResistance to Innovations Related to FoodCanningCanning

Change is InevitableProgress is Optional

History of Technology AdoptionHistory of Technology Adoption

Resistance to Innovations Related to FoodResistance to Innovations Related to FoodFreezingFreezing

Change is InevitableProgress is Optional

History of Technology AdoptionHistory of Technology Adoption

Certain Innovations Not Readily AcceptedCertain Innovations Not Readily AcceptedRecalcitrance to Adopt (Dvorak Recalcitrance to Adopt (Dvorak v/sv/s QWERTY)QWERTY)

Change is InevitableProgress is Optional

History of Technology AdoptionHistory of Technology Adoption

Certain Innovations Not Readily AcceptedCertain Innovations Not Readily AcceptedEntrenched Economic Interests (Metric in US)Entrenched Economic Interests (Metric in US)

Change is InevitableProgress is Optional

History of Technology AdoptionHistory of Technology Adoption

Risk, definedRisk, defined

The probability of a hazardThe probability of a hazardThe probability of exposure to hazardThe probability of exposure to hazard

RISKRISK

Risk = f (hazard, probability)Risk = f (hazard, probability)Risk = f (exposure x hazard), Risk = f (exposure x hazard),

Where Where •• exposure is the frequency at which an exposure is the frequency at which an

event occurs and event occurs and •• hazard is the magnitude of the impact hazard is the magnitude of the impact

of that eventof that event

Risks, examplesRisks, examplesRisks for animal and human healthRisks for animal and human health•• Toxicity & food quality safetyToxicity & food quality safety•• Pathogen drug resistancePathogen drug resistance

Risks for the environmentRisks for the environment•• Persistency of gene of transgene (volunteers, Persistency of gene of transgene (volunteers,

increased fitness, invasiveness) or transgenic increased fitness, invasiveness) or transgenic products (accumulative effect)products (accumulative effect)

•• Susceptibility of nonSusceptibility of non--target organismstarget organisms•• Increased use of chemicals in agricultureIncreased use of chemicals in agriculture•• Unpredictable gene expression or transgene Unpredictable gene expression or transgene

instabilityinstability

Risks, examplesRisks, examples

Risks in agricultureRisks in agriculture•• Resistance or tolerance of target organismResistance or tolerance of target organism•• Weeds or super weedsWeeds or super weeds•• Alteration of nutritional value (attractiveness Alteration of nutritional value (attractiveness

of the organism to pests)of the organism to pests)•• Horizontal gene transferHorizontal gene transfer

Genetic pollution through pollen or seed dispersalGenetic pollution through pollen or seed dispersalTransfer of foreign gene to microorganism (DNA Transfer of foreign gene to microorganism (DNA uptake) or generation or new live viruses by uptake) or generation or new live viruses by recombinationrecombinationInteraction among different Interaction among different GMOsGMOs

Risk analysis, three steps inRisk analysis, three steps inRisk assessmentRisk assessment•• evaluates and compares scientific evidence evaluates and compares scientific evidence

regarding the risks associated with alternative regarding the risks associated with alternative activitiesactivities

Risk managementRisk management•• develops strategies to prevent and control develops strategies to prevent and control

risks within acceptable limits and relies on risk risks within acceptable limits and relies on risk assessmentassessment

Risk communicationRisk communication•• involves an oninvolves an on--going dialogue between going dialogue between

regulators and the public about risk and regulators and the public about risk and options to manage risk so that appropriate options to manage risk so that appropriate decisions can be madedecisions can be made

General risk assessment of General risk assessment of GMOsGMOs is is different from that of chemicalsdifferent from that of chemicals

GMO are living organismsGMO are living organismsPotential to disperse to new habitats, Potential to disperse to new habitats, colonize these sites and multiplycolonize these sites and multiplyTheir novel activity (production of Their novel activity (production of metabolic products, enzymes, toxins) will metabolic products, enzymes, toxins) will occur as long as occur as long as GMOsGMOs remain remain metabolically activemetabolically activeOnce established, living organism cannot Once established, living organism cannot be recalled be recalled

Risk assessmentRisk assessment

Identifying and evaluating possible Identifying and evaluating possible dangers dangers predicting the chances the danger predicting the chances the danger will occurwill occurassessing the extent of damage assessing the extent of damage should the danger occurshould the danger occur

Risk assessmentRisk assessment

ScienceScience--basedbasedTransparentTransparentCase by case by transformation Case by case by transformation eventeventAbsence of scientific evidence is not Absence of scientific evidence is not construed to imply safety nor dangerconstrued to imply safety nor dangerRisk assessment is revisited if new Risk assessment is revisited if new evidence is foundevidence is found

Risk assessmentRisk assessmentIn order to be able to assess the environmental In order to be able to assess the environmental safety of a genetically engineered plant, safety of a genetically engineered plant, one one must be familiar with both the biology of the must be familiar with both the biology of the plant itselfplant itself, , This concept of This concept of familiarityfamiliarity is a key approach is a key approach used in identifying and evaluating environmental used in identifying and evaluating environmental risks (i.e., hazards) that may be associated with risks (i.e., hazards) that may be associated with the release of a genetically engineered plant and the release of a genetically engineered plant and also in informing management practices that may also in informing management practices that may be needed to manage recognized risks. be needed to manage recognized risks. Familiarity considers the Familiarity considers the biology of the plant biology of the plant speciesspecies, the , the traittrait, and the , and the agricultural practicesagricultural practicesused in the production of the crop.used in the production of the crop.

The Biology of the Plant SpeciesThe Biology of the Plant Species

AA MonographMonograph may be used to identify may be used to identify speciesspecies--specific characteristics that may specific characteristics that may be affected by the novel trait so as to be affected by the novel trait so as to permit the transgenic plant to become permit the transgenic plant to become ““weedyweedy””, invasive of natural habitats, or , invasive of natural habitats, or be otherwise harmful to the environment. be otherwise harmful to the environment. It can also provide details on significant It can also provide details on significant interactions between the plant and other interactions between the plant and other lifelife--formsforms that must be evaluated in the that must be evaluated in the impact analysis. impact analysis.

The Biology of the Plant SpeciesThe Biology of the Plant Species

Typically such a document includes the following:Typically such a document includes the following:•• Taxonomic descriptionTaxonomic description•• Consumption and uses of the crop plantConsumption and uses of the crop plant•• Regional/national breeding, seed production, and agronomic Regional/national breeding, seed production, and agronomic

practicespractices•• Reproductive biology of the crop plant, including details on Reproductive biology of the crop plant, including details on

pollination, mechanisms for dispersal of pollen and seed, and pollination, mechanisms for dispersal of pollen and seed, and any other means of gene escapeany other means of gene escape

•• Occurrence and viability of Occurrence and viability of intraspecificintraspecific, , interspecificinterspecific, and , and intergenericintergeneric hybridshybrids

•• Details on the Details on the centrescentres of origin and genetic diversity for the of origin and genetic diversity for the plant speciesplant species

•• Details on the Details on the ploidyploidy of the cultivated crop, its progenitors and of the cultivated crop, its progenitors and any sexually compatible speciesany sexually compatible species

•• Distribution and ecology of related species or feral biotypes, Distribution and ecology of related species or feral biotypes, including any evidence of including any evidence of weedinessweediness

•• Common diseases and pestsCommon diseases and pests•• Potential interactions with other organisms such as pollinators,Potential interactions with other organisms such as pollinators,

mycorrhizalmycorrhizal fungi, animal browsers, birds, soil microbes and fungi, animal browsers, birds, soil microbes and soil insectssoil insects

Novel TraitNovel Trait

The environmental risk of releasing a The environmental risk of releasing a genetically modified plant is genetically modified plant is notnotdetermined by the determined by the novelty of the genetic novelty of the genetic elementselements used to transform it, but by the used to transform it, but by the novelty of the plant itselfnovelty of the plant itself. . Useful information can be gained from Useful information can be gained from evaluating whether a trait expressed in a evaluating whether a trait expressed in a transgenic plant is similar to traits already transgenic plant is similar to traits already introduced into that plant species. introduced into that plant species. •• For example, the potential impact of For example, the potential impact of

introducing a introducing a fungal resistance genefungal resistance gene on the on the persistence of a persistence of a transgenictransgenic plant may be plant may be addressed, in part, by evaluating the effect addressed, in part, by evaluating the effect that that conventionally bredconventionally bred varieties with the varieties with the same fungal resistance trait had when they same fungal resistance trait had when they were released. were released.

Substantial EquivalenceSubstantial Equivalence

The concept of familiarity is The concept of familiarity is coupledcoupled with with that of substantial equivalencethat of substantial equivalenceSubstantial equivalence is based on the principle Substantial equivalence is based on the principle that novel that novel plants can be compared with their nonplants can be compared with their non--transformed transformed counterparts that have an established history of safe usecounterparts that have an established history of safe useThe The objective is to determineobjective is to determine if the novel plant presents if the novel plant presents any new or any new or greater risksgreater risks in comparison in comparison with its traditional with its traditional counterpartcounterpart, or whether it can be used , or whether it can be used interchangeably with interchangeably with its traditional counterpart without negatively affecting the its traditional counterpart without negatively affecting the environmentenvironment in which it is grownin which it is grownThe goal is not to establish an absolute level of safetyThe goal is not to establish an absolute level of safety, but , but rather a relative level of safety, so that there is a rather a relative level of safety, so that there is a reasonable certainty that no undue risk to the environment reasonable certainty that no undue risk to the environment will result from the cultivation of the novel plantwill result from the cultivation of the novel plant under under anticipated conditions of productionanticipated conditions of production

Ancient AgricultureAncient Agriculture(Domestication of Wild Species)(Domestication of Wild Species) Modern AgricultureModern Agriculture

(Traditional Plant Breeding)(Traditional Plant Breeding)

Modern AgricultureModern Agriculture(Genetic Engineering)(Genetic Engineering)

From a trichological point of view

Safety ConsiderationsSafety ConsiderationsThe The goalgoal of environmental risk assessment of of environmental risk assessment of genetically modified plants is to identify and genetically modified plants is to identify and evaluate the risks associated with the release and evaluate the risks associated with the release and cultivation of these plants cultivation of these plants in comparison with a in comparison with a counterpart that has a history of safe usecounterpart that has a history of safe use. . In addition to information on the host organism, In addition to information on the host organism, and the donor organism, particularly if and the donor organism, particularly if the donor the donor or other members of its genus normally exhibit or other members of its genus normally exhibit characteristics of characteristics of pathogenicitypathogenicity or environmental or environmental toxicology, or have other traits that affect human toxicology, or have other traits that affect human health, these concerns include:health, these concerns include:•• Molecular characterization and stability of genetic Molecular characterization and stability of genetic

modificationmodification•• Gene transfer to related plantsGene transfer to related plants•• Gene transfer to unrelated organismsGene transfer to unrelated organisms•• WeedinessWeediness potentialpotential•• Secondary and nonSecondary and non--target adverse effectstarget adverse effects

Bacillus thuringiensis VsBacillus anthracis

Molecular characterization and Molecular characterization and stability of genetic modificationstability of genetic modification

Characterization of a transgenic plant at the Characterization of a transgenic plant at the molecular level is used to provide information molecular level is used to provide information about: about: •• the composition and integrity of the inserted DNAthe composition and integrity of the inserted DNA•• the number of copies of the inserted DNA the number of copies of the inserted DNA •• the number of sites of insertion and the number of sites of insertion and •• the level of expression of the novel the level of expression of the novel protein(sprotein(s) over time ) over time

and in different tissues.and in different tissues.It is very important that a It is very important that a rigorous molecular rigorous molecular characterization of each transgenic plant characterization of each transgenic plant submittedsubmitted for review be completed, it is equally for review be completed, it is equally important to recognize the limitations of this important to recognize the limitations of this approach in predicting the safety of a novel food approach in predicting the safety of a novel food or plant.or plant.

On an individual event basis

Gene Transfer to Related PlantsGene Transfer to Related PlantsIntrogression from one plant to another Introgression from one plant to another is is only significant if the two plants are only significant if the two plants are sexually compatible and if their hybrid sexually compatible and if their hybrid offspring are viable. offspring are viable. In order to assess potential environmental In order to assess potential environmental risks associated with risks associated with outcrossingoutcrossing from from transgenic plants, transgenic plants, •• the reproductive biology of the plant and the reproductive biology of the plant and

distribution of sexually compatible relatives distribution of sexually compatible relatives must be knownmust be known, and , and

•• the the impactimpact of the introduced trait, of the introduced trait, should it be should it be introgressedintrogressed into other plant speciesinto other plant species, must be , must be understood. understood.

Bt cotton Vs wild cotton

Gene Transfer to Unrelated OrganismsGene Transfer to Unrelated Organisms

Horizontal gene transfer (HGT) is the nonHorizontal gene transfer (HGT) is the non--sexual sexual exchange of genetic material between organisms exchange of genetic material between organisms belonging to the same or different species belonging to the same or different species The possibility of HGT between plants and bacteria in The possibility of HGT between plants and bacteria in either the soil or gut either the soil or gut -- particularly as this relates to particularly as this relates to the possible transfer of genes encoding antibiotic the possible transfer of genes encoding antibiotic resistance resistance -- has been seen as a hazard associated has been seen as a hazard associated with genetically engineered plants with genetically engineered plants HGT is a natural process of crossHGT is a natural process of cross--species gene species gene movement responsible for effecting genetic change, movement responsible for effecting genetic change, and that genes introduced into transgenic plants are and that genes introduced into transgenic plants are no more likely to be transferred to other organisms no more likely to be transferred to other organisms than are other plant genesthan are other plant genes

CaMV 35S promoter and safety of Cauliflower

WeedinessWeediness PotentialPotentiala measure of a planta measure of a plant’’s ability to successfully s ability to successfully colonize an ecosystem, especially when it may colonize an ecosystem, especially when it may also lead to the displacement of other speciesalso lead to the displacement of other species

To evaluate if a transgenic plant has altered To evaluate if a transgenic plant has altered weedinessweediness potential in comparison with its potential in comparison with its conventional counterpart, the following may be conventional counterpart, the following may be examined: examined: •• Dissemination of seedDissemination of seed•• Dormancy of seedDormancy of seed•• Germination of seed/survivalGermination of seed/survival•• CompetitivenessCompetitiveness•• Agronomic characteristics Agronomic characteristics

time to maturity, disease and pest resistancetime to maturity, disease and pest resistance•• Stress toleranceStress tolerance

e.g.: the Congress weed (Parthenium)

Secondary and NonSecondary and Non--Target Adverse EffectsTarget Adverse Effects

Environmental risk assessment must Environmental risk assessment must consider the consider the unintended consequencesunintended consequences of of the environmental release of a transgenic the environmental release of a transgenic plant, particularly as this may impact on plant, particularly as this may impact on existing agricultural practices and the existing agricultural practices and the agroagro--ecosystemecosystemA A nonnon--target organismtarget organism is any plant, is any plant, animal or microorganism that is animal or microorganism that is unintentionally impacted by the novel, or unintentionally impacted by the novel, or transgenic, plant. transgenic, plant.

Risk assessment, the OECD guidelinesRisk assessment, the OECD guidelines

According to the According to the OrganisationOrganisation for Economic Cofor Economic Co--operation operation and Developmentand Development (OECD):(OECD):•• Safety in biotechnology is achieved by the appropriate Safety in biotechnology is achieved by the appropriate

application of risk/safety analysis and risk management. application of risk/safety analysis and risk management. Risk/safety analysis comprises hazard identification and, if a Risk/safety analysis comprises hazard identification and, if a hazard has been identified, risk assessment. hazard has been identified, risk assessment.

•• Risk/safety analysis is based on the characteristics of the Risk/safety analysis is based on the characteristics of the organism, the introduced trait, the environment into which the organism, the introduced trait, the environment into which the organism is introduced, the interaction between these, and the organism is introduced, the interaction between these, and the intended application. intended application.

•• Risk/safety analysis is conducted prior to an intended action Risk/safety analysis is conducted prior to an intended action and is typically a routine component of research, development and is typically a routine component of research, development and testing of new organisms, whether performed in a and testing of new organisms, whether performed in a laboratory or a field setting. laboratory or a field setting.

•• Risk/safety analysis is a scientific procedure which does not Risk/safety analysis is a scientific procedure which does not imply or exclude regulatory oversight or imply that every case imply or exclude regulatory oversight or imply that every case will necessarily be reviewed by a national or other authority.will necessarily be reviewed by a national or other authority.

In short, risk assessment of genetically engineered plants In short, risk assessment of genetically engineered plants should be based on sound science and should be applied on should be based on sound science and should be applied on a casea case--byby--case basis.case basis.

Case study Case study –– Bt corn Mon 810 Bt corn Mon 810

Host OrganismHost Organism

Genetic and Phenotypic VariabilityGenetic and Phenotypic VariabilityThe assessment of the interaction of MON 810 The assessment of the interaction of MON 810 with the environment has included studies on:with the environment has included studies on:•• susceptibility to insects and diseases; susceptibility to insects and diseases; •• survival capacity (volunteers); survival capacity (volunteers); •• seed multiplication capacity (yields); seed multiplication capacity (yields); •• Cry1Ab protein expression in leaves and grain; Cry1Ab protein expression in leaves and grain; •• seed composition analysis; seed composition analysis; •• safety for birds; and safety for birds; and •• safety for mammals safety for mammals

No significant differences have been observed No significant differences have been observed between MON810 and other maize varietiesbetween MON810 and other maize varieties apart apart from protection from certain from protection from certain lepidopteranlepidopteran insectsinsects

Host OrganismHost Organism

No differencesNo differences in seed or plant maturity in seed or plant maturity have been observedhave been observedA comparison of a nonA comparison of a non--transgenic hybrid transgenic hybrid with the same hybrid in which one parent with the same hybrid in which one parent was a backcross derived MON 810 line was a backcross derived MON 810 line showed showed no significant differenceno significant difference between between these hybrids in yield these hybrids in yield No differencesNo differences in agronomic quality, in agronomic quality, disease, or insect susceptibility other than disease, or insect susceptibility other than European corn borer control were detected European corn borer control were detected between MON 810 and nonbetween MON 810 and non--transgenic transgenic plants. plants.

Donor organismDonor organism

MON 810 contains DNA sequences derived MON 810 contains DNA sequences derived from the following donor organisms:from the following donor organisms:•• Bacillus Bacillus thuringiensisthuringiensis cry1Abcry1Ab genegene•• Cauliflower mosaic virus (Cauliflower mosaic virus (CaMVCaMV) enhanced 35S ) enhanced 35S

promoter with the duplicated enhancer region promoter with the duplicated enhancer region •• The The intronintron from the maize from the maize hsp70hsp70 gene for an gene for an

increased level of transcription increased level of transcription •• The 3' The 3' untranslateduntranslated region of the region of the nopalinenopaline

synthasesynthase gene (NOS 3') from the Ti plasmid of gene (NOS 3') from the Ti plasmid of Agrobacterium tumefaciensAgrobacterium tumefaciens. .

NoneNone of the inserted sequences are known of the inserted sequences are known to have any pathogenic or harmful to have any pathogenic or harmful characteristics. characteristics.

Transformation MethodTransformation Method

Particle bombardmentParticle bombardmentGenes encoding for Genes encoding for glyphosateglyphosate tolerance (CP4 tolerance (CP4 EPSPS and EPSPS and goxgox genes used for selection but not genes used for selection but not present in MON810present in MON810•• CoCo--transformation of pVtransformation of pV--ZMBKO7 (contains the ZMBKO7 (contains the cry1Abcry1Ab

gene) and gene) and pVpV--ZMGTlOZMGTlO (containing the CP4 EPSPS and (containing the CP4 EPSPS and goxgox genes) genes)

•• nptIInptII for bacterial selection, origin of replication from for bacterial selection, origin of replication from pUCpUC

Plant expression vector Plant expression vector •• cry1Abcry1Ab gene under the control of the enhanced gene under the control of the enhanced CaMVCaMV

35S promoter (0.6 kb)35S promoter (0.6 kb)•• 0.8 kb 0.8 kb intronintron from the maize from the maize hsp70hsp70 located between the located between the

thethe promoter and the promoter and the cry1Abcry1Ab genegene•• IntronIntron followed by 3.47 kb followed by 3.47 kb cryIAbcryIAb gene gene –– modified to modified to

increase the levels of expression in maize increase the levels of expression in maize •• 0.27 kb 0.27 kb nopalinenopaline synthasesynthase 3' 3' nontranslatednontranslated sequence, sequence,

NOS 3NOS 3

Molecular Molecular CharacterisationCharacterisation

Southern Blot Analysis Southern Blot Analysis •• Probes homologous to the Probes homologous to the cryIAbcryIAb, CP4 EPSPS, , CP4 EPSPS,

goxgox, , nptIInptII, and , and oriori--pUCpUC genetic regions genetic regions •• the insert number (number of integration sites the insert number (number of integration sites

within the maize genome), and the copy within the maize genome), and the copy number and integrity of each gene was number and integrity of each gene was examined examined

Insert Number (Insert Number (cryIAbcryIAb)) –– one one MON 810 MON 810 does not containdoes not contain the CP4 EPSPS the CP4 EPSPS and and goxgox gene gene No backbone integrationNo backbone integration

Genetic stability of introduced traitGenetic stability of introduced trait

Segregation Analysis of MON 810 Segregation Analysis of MON 810 –– a a single active insert single active insert segregating segregating according to according to MendelianMendelian genetics genetics StabilityStability of this insertion of this insertion demonstrated through demonstrated through seven seven generationsgenerations of crossing of crossing Southern blot analysis demonstrated Southern blot analysis demonstrated that the that the cry1Abcry1Ab insertion insertion was stable was stable through threethrough three generationsgenerations

Expression of transgeneExpression of transgeneField trials of MON 810 were conducted in major maize Field trials of MON 810 were conducted in major maize growing regions of US, Italy, and France (representing a growing regions of US, Italy, and France (representing a variety of environmental conditions) variety of environmental conditions) The identity and levels of expression of the Cry1Ab protein The identity and levels of expression of the Cry1Ab protein in plant tissue samples collected from these sites in plant tissue samples collected from these sites determined by ELISAdetermined by ELISAThe Cry1Ab protein levels in maize line The Cry1Ab protein levels in maize line MON 810 were lowMON 810 were lowrelative to total protein in maize leaf, grain and whole plant relative to total protein in maize leaf, grain and whole plant tissues, but tissues, but sufficient to provide season long controlsufficient to provide season long control of ECB of ECB CP4 EPSPS and GOX protein were CP4 EPSPS and GOX protein were not detectablenot detectable in maize in maize leaf, grain and whole plant tissue leaf, grain and whole plant tissue MON 810 was shown by Western blot analysis to contain a MON 810 was shown by Western blot analysis to contain a Cry1Ab Cry1Ab trypsintrypsin--resistant core protein that was equivalent to resistant core protein that was equivalent to the the E. coliE. coli--produced Cry1Ab produced Cry1Ab trypsintrypsin--resistant core protein resistant core protein used for safety assessment studies. The equivalence used for safety assessment studies. The equivalence established by this study served as the justification for established by this study served as the justification for using the safety data generated with using the safety data generated with E. coliE. coli--produced produced Cry1Ab Cry1Ab trypsintrypsin--resistant core protein to support the safety resistant core protein to support the safety of the Cry1Ab of the Cry1Ab trypsintrypsin--resistant core protein expressed in resistant core protein expressed in this insect protected maize line this insect protected maize line

Gene transfer to related plants Gene transfer to related plants For gene flow to occur via normal sexual For gene flow to occur via normal sexual transmission, certain conditions must exist:transmission, certain conditions must exist:•• the two parents must be sexually compatiblethe two parents must be sexually compatible•• their fecundity must coincidetheir fecundity must coincide•• a suitable pollen vector must be present and capable of a suitable pollen vector must be present and capable of

transferring pollen between the two parents and transferring pollen between the two parents and •• resulting progeny must be fertile and ecologically fit for resulting progeny must be fertile and ecologically fit for

the environment in which they are situatedthe environment in which they are situatedMaize easily crosses with Maize easily crosses with teosinteteosinte, but , but teosinteteosinte is is not present in the United Statesnot present in the United StatesOutcrossingOutcrossing with with TripsacumTripsacum species is not knownspecies is not knownto occur in the wild and it is only with extreme to occur in the wild and it is only with extreme difficulty that maize can be crossed with difficulty that maize can be crossed with TripsacumTripsacumNo cases of gene flow between maize and its wild No cases of gene flow between maize and its wild relatives are known in either Canada or the relatives are known in either Canada or the United States United States

Gene transfer to related plants Gene transfer to related plants OutcrossingOutcrossing to Cultivated to Cultivated ZeaZea VarietiesVarieties

Gene exchange between varieties of cultivated and Gene exchange between varieties of cultivated and genetically modified maize will be similar to that which genetically modified maize will be similar to that which occurs naturally between cultivated maize varieties. occurs naturally between cultivated maize varieties. WindWind--borne pollen will move between plants within the borne pollen will move between plants within the same field and to maize plants in nearby fields. same field and to maize plants in nearby fields. The transfer of the transgenic trait to other cultivated maize The transfer of the transgenic trait to other cultivated maize will not impart any additional safety concernswill not impart any additional safety concerns to those to those already identified and addressed for the original transgenic already identified and addressed for the original transgenic maize maize The environmental impact of the novel trait in cultivated The environmental impact of the novel trait in cultivated maize will have been considered during the environmental maize will have been considered during the environmental risk assessment of the genetically modified line and risk assessment of the genetically modified line and transferring the trait, either intentionally or unintentionally,transferring the trait, either intentionally or unintentionally,to other maize varieties to other maize varieties will not result in additional or new will not result in additional or new risks risks

WeedinessWeediness potentialpotential

Dissemination of seed Dissemination of seed -- Maize cannot survive Maize cannot survive without human assistance due to past selection in without human assistance due to past selection in its evolution and consequently, seed dispersal of its evolution and consequently, seed dispersal of individual kernels naturally does not occur individual kernels naturally does not occur because of the structure of the ears of maize. because of the structure of the ears of maize. The introduced trait, insectThe introduced trait, insect--protection, had no protection, had no influence on reproductive morphology and hence influence on reproductive morphology and hence no changes in seed dissemination would be no changes in seed dissemination would be expectedexpectedthere are there are no differencesno differences in vegetative vigour or in vegetative vigour or adaptation to environmental stress factors adaptation to environmental stress factors including drought, heat, and frost between MON including drought, heat, and frost between MON 810 and the parental control or other maize lines 810 and the parental control or other maize lines of similar genetic background of similar genetic background

Secondary and NonSecondary and Non--Target Adverse EffectsTarget Adverse Effects

Plant pest potential Plant pest potential -- A plant may be A plant may be considered a pest but not a weed. For considered a pest but not a weed. For example, a plant that produces an example, a plant that produces an allelopathicallelopathic substance may be considered substance may be considered a pest if the toxin produced has an a pest if the toxin produced has an undesirable environmental effect. undesirable environmental effect. Transgenic plants expressing novel toxins Transgenic plants expressing novel toxins or potential allergens must be assessed or potential allergens must be assessed accordinglyaccordingly•• There is no evidenceThere is no evidence to indicate that MON 810 to indicate that MON 810

has altered plant pest potential in comparison has altered plant pest potential in comparison with its nonwith its non--transformed counterpart.transformed counterpart.

Effect on nonEffect on non--target organismtarget organismThe The insecticidallyinsecticidally active active trypsintrypsin--resistant core protein (HDresistant core protein (HD--1) produced in MON810, is 1) produced in MON810, is identical toidentical to the respective full the respective full length and length and trypsintrypsin--resistant core Cry1Ab proteins contained resistant core Cry1Ab proteins contained in commercial microbial formulationsin commercial microbial formulationsCry1Ab protein is Cry1Ab protein is extremely selectiveextremely selective for the for the lepidopteranlepidopteraninsects bind specifically to receptors on the midinsects bind specifically to receptors on the mid--gut of gut of lepidopteranlepidopteran insects and have no deleterious effect on insects and have no deleterious effect on beneficial/nonbeneficial/non--target insects target insects Cry1AbCry1Ab--expressing maize had expressing maize had neither a direct nor an neither a direct nor an indirect effect on the beneficial arthropod speciesindirect effect on the beneficial arthropod species studied. studied. InsectInsect--protected maize had protected maize had no effect on spiders, no effect on spiders, coccinellidcoccinellid, , chrysopidchrysopid which are also known to be important predators which are also known to be important predators of the European corn borer as well as of other economically of the European corn borer as well as of other economically important pests of maize. important pests of maize.

Effect on nonEffect on non--target organismtarget organism

Dietary feeding trials of key nonDietary feeding trials of key non--target target indicator speciesindicator species•• Honey bee larvae and adultsHoney bee larvae and adults•• Green lacewingGreen lacewing•• Parasitic Parasitic HymenopteranHymenopteran•• Ladybird beetlesLadybird beetles•• Daphnia Daphnia •• EarthwormEarthworm•• CollembolaCollembola•• Northern Bobwhite Quail Northern Bobwhite Quail

Resistance ManagementResistance ManagementThe potential development of insect The potential development of insect resistance to resistance to BtBt, as a consequence of , as a consequence of largelarge--scale commercial plantings of scale commercial plantings of BtBtcrops, is also a concern to farmerscrops, is also a concern to farmers““high dose/refuge strategyhigh dose/refuge strategy””Exposing a portion of the pest population Exposing a portion of the pest population to to BtBt plants with an extremely high plants with an extremely high concentration of toxin [25 times the concentration of toxin [25 times the amount needed to kill 99% of the amount needed to kill 99% of the susceptible insects], while maintaining susceptible insects], while maintaining another part of the population in a refuge another part of the population in a refuge where the pests do not encounter any where the pests do not encounter any BtBttoxin toxin

Best Management Principles for Best Management Principles for BtBt CropsCrops

A specific Insect Resistance Management plan is A specific Insect Resistance Management plan is necessary to ensure longnecessary to ensure long--term resistance term resistance management. elements of the IRM plan are: management. elements of the IRM plan are: •• high dosehigh dose•• structured refugestructured refuge•• susceptible pest biology and ecology datasusceptible pest biology and ecology data•• impact on secondary pests impact on secondary pests •• impact on pests affecting multiple impact on pests affecting multiple BtBt crops crops •• crosscross--resistance potential resistance potential •• resistance mechanismsresistance mechanisms•• monitoring/surveillance and monitoring/surveillance and •• remedial actionremedial action

Best Management Principles for Best Management Principles for BtBt CropsCrops

A high dose/structured refuge strategy is necessary to ensure A high dose/structured refuge strategy is necessary to ensure longlong--term resistance management.term resistance management.Grower education, adoption, and compliance are essential to the Grower education, adoption, and compliance are essential to the implementation and success of a longimplementation and success of a long--term resistance term resistance management strategy.management strategy.BtBt crops are to be used as part of an integrated pest management crops are to be used as part of an integrated pest management program to enhance pest management goals.program to enhance pest management goals.Coordinated annual performance monitoring and surveillance is Coordinated annual performance monitoring and surveillance is necessary to detect or follow resistance development.necessary to detect or follow resistance development.Immediate and coordinated remedial action for suspected and Immediate and coordinated remedial action for suspected and confirmed incidents of resistance is necessary.confirmed incidents of resistance is necessary.IRM strategies should be tailored to address specific regional IRM strategies should be tailored to address specific regional resistance management concerns, as appropriate.resistance management concerns, as appropriate.Deployment of IPM tactics with different modes of action, Deployment of IPM tactics with different modes of action, including conventional pesticides, including conventional pesticides, BtBt toxins expressed in crops toxins expressed in crops with different modes of action, biological control methods, and with different modes of action, biological control methods, and other control methods, is essential for sustainable pest other control methods, is essential for sustainable pest management goalsmanagement goalsContinued resistance management research should be conducted Continued resistance management research should be conducted to evaluate the effectiveness of, and be used to modify, as to evaluate the effectiveness of, and be used to modify, as necessary, IRM strategies for necessary, IRM strategies for BtBt crops crops

Think it over!!!Think it over!!!

Popular assumption: Popular assumption: ““Whatever we Whatever we get in nature is get in nature is goodgood””Fact: Whatever we get in nature is Fact: Whatever we get in nature is neutralneutral..•• Viewing it as Viewing it as goodgood or or badbad depends upon depends upon

makemake--up of our mind and popular up of our mind and popular opinion or the conditioned opinion or the conditioned behaviourbehaviour

Oliver GoldsmithOliver Goldsmith

Oliver GoldsmithOliver Goldsmith’’s famous lines in his s famous lines in his poem poem ““the deserted villagethe deserted village””

•• 55: But a bold peasantry, their country's pride,55: But a bold peasantry, their country's pride,•• 56: When once 56: When once destroy'ddestroy'd, can never be supplied., can never be supplied.

-- The King of The King of BrobdingnagBrobdingnag, Gulliver's , Gulliver's Travels by Jonathan Swift, 1727.Travels by Jonathan Swift, 1727.

"Whoever could make two ears of "Whoever could make two ears of corn, or two blades of grass grow corn, or two blades of grass grow upon a spot of ground where only upon a spot of ground where only one grew before would deserve one grew before would deserve better of Mankind, and do more better of Mankind, and do more

essential service to his country, than essential service to his country, than the whole race of politicians put the whole race of politicians put

together." together."