Risk assessment and Gene flow

Mike Wilkinson

United Kingdom

Contents

1. Current state of GM technology2. Principles of risk assessment3. Defining hazards4. Measuring exposure5. Conclusions

Current state of GM technology

• Most crops can be genetically modified

• Four crops dominate (Maize, Cotton, Soya, oilseed rape)

• Herbicide tolerance and insect resistance account for nearly all GM cultivars

• Four countries dominate GM production (USA, China, Argentina, Canada)

Future

• More GM crops will be commercialised

• More countries will approve commercial release

• The trend is towards transgene stacking

• The next wave of GM crops will target stress tolerance (drought and salt)

Implications for risk assessment

As the number of crop-cultivar-gene-gene mixture-location-construct mixes grows,

So does the need for generic information to assess risks

•RISK

•HAZARD

•EXPOSURE

Principles of Risk Assessment

Hazard, exposure & risk

• Hazard– Adverse effect

• Exposure– Frequency/intensity of contact with agent

causing adverse effect

• Risk– Magnitude and likelihood of an adverse effect

A ‘bad thing’

Is it likely the ‘bad thing’ will happen?

a judgement based on ‘how bad’ and ‘how likely’

Risk = f (Hazard, Exposure)

Ecological hazards

That a transgene from a GM crop will move into a wild relative

AND

lead to some form of unwanted ecological change

First generation hybrid

Seed

Seed

cross-fertilise

cross-fertilise

Second generation hybrid

Crop

Wild species

Invasion of new habitats

Replacement of existing genotypes

Changed abundance/mix of herbivores

Out-compete other species in same habitat

Change pollinator mix in community

Changed abundance/mix of predators

Categories of ecological hazard

• Relating to the recipient (Direct hazards)– Increased population size within habitat

– Invasion of new habitat

– Replacement of native genotypes

• Relating to other organisms (Indirect hazards)– Decline in sympatric plant species

– Changed pollinator abundance

– Changed herbivore abundance

– Changed predator/parasitoid abundance

Note:

The ‘end point’ species may not be the wild recipient

Having defined the hazard

What is the exposure?

Exposure pathway concept

In chemical toxicology, exposure is to a toxic chemical is simple and direct

The ecological hazards represent endpoints in a pathway or matrix of linked events

GM crop

F1 hybrid in region

Transgene stabilises by introgression

Transgene spreads to most populations

Enhanced resistance to herbivore depresses herbivore numbers

Depressed herbivore numbers cause extinction of specialist parasitoid of the herbivore

Wilkinson et al 2003. Trends in Plant Science

GM crop

F1 hybrid in region

Transgene stabilises by introgression

Transgene spreads to most populations

GM crop

F1 hybrid in region

Transgene stabilises by introgression

Transgene spreads to most populations

Generic exposure elements (applies to most transgenes)Specific exposure elements (applies to 1-few genes)

Hazard

The risk assessment process

1. Specify and rank the hazards2. Quantify generic aspects of exposure3. Evaluate aspects of exposure specific to

one transgene or GM cultivar4. Assess the risks

Stage 1: Specify and rank hazards

1. Identify cross-compatible recipients

2. Rank crudely according to likelihood (sympatry, ease of hybridization)

3. Specify hazards relating to recipient (direct hazards)

4. Specify hazards relating to sympatric organisms (indirect hazards)

1. Which species are interfertile with wheat?

The tribe Triticeae contains around 330 species in 18 genera and shows ‘an exceptional capacity for intergeneric hybridisation’ Clayton and Renvoize (1986)

Breeders define ‘Gene Pool’ groupings of species related to wheat based on taxonomy and ease of crossing

Gene pool 1: Wild species within Triticum

‘Crossing is easy and hybrids fertile’

Triticum monococcum ssp. aegilopoides

Triticum urartu

Triticum turgidum ssp. dicoccoides

Triticum timopheevii ssp. armeniacum

……..Not found in the USA

Gene Pool 2: Closely related Genera

‘All species that will cross with a crop, although with more difficulty than GP1. Hybrids tend to be sterile’

Comprises of species from the following genera:

Aegilops (22 species)

Amblyopyrum (1 species)

Agropyrum (4 species)

Dasypyrum (2 species)

……Several species introduced into USA

Gene Pool 3: ‘Wide hybrids’

‘Gene transfer is not possible without radical techniques’

Includes species from:

Agropyrum

Elymus (eg couch grass)

Hordeum

…….Natural hybrids not recorded

Evidence of natural hybrids in Gene Pool 2 ?

At least 12 species

Which of these occur in USA?

Aegilops triuncialis

Aegilops geniculata

Aegilops tauschii

Aegilops neglecta

Aegilops cylindrica

Wheat production in the USA: Harvested area per county for 2000

Aegilops triuncialis

Aegilops geniculata

Aegilops tauschii

Aegilops neglecta

(positions approximate)

Species interfertile with wheat found in USA

Ffffff

ff

Distribution of Aegilops cylindrica in USA: 1993 USDA survey

Other records (positions approximate)

Present

Moderate to dense infestations

Hazards for Aegilops cylindricawhen exposed to GMHT wheat

1. The transgene (herbicide tolerance) will cause A. cylindrica to become a more aggressive weed (Direct Hazard)

2. Changed herbicide use on A. cylindrica will cause decline in a named plant or animal species (Indirect Hazards)

Prioritising hazards

For any crop-wild relative combination, there are many possible and fewer plausible hazards

But comprehensive risk assessment is both expensive and slow

So, we need to prioritise hazards

Possible options for ranking hazards

1. Scarcity of the ‘end point’ species2. Cultural importance (Bald Eagle in the

USA)3. Ecological importance of ‘end point’

species4. Agronomic or medicinal value of ‘end

point’ species5. Cuddliness of ‘end point’ species

Stage 2: quantify exposure

1. Generic exposure elements

2. Specific exposure elements

Quantify generic elements of exposure

1. Quantify hybridization

2. Quantify introgression

3. Quantify gene spread

Quantifying hybrid formation

Important to define1. Context of contact (weed or adjacent wild

population)2. Distribution of crop and recipient species3. Relationship between hybrid frequency

and separation, donor/recipient population size

4. Crop rotation patterns

Cultivated and wild Helianthus annuus, Nebraska USA, in sympatry

1. Context of contact

2. Crop and recipient distribution

??

Sunflower production in the USA: Harvested area per county for 2000

wild Helianthus annuus

Wild Helianthus annus in the USA (data incomplete) – USDA NRCS 2001

This scale is too crude

So use

• Direct surveys

• Literature

• Remote sensing

• Herbarium specimens

3. Importance of separation and population size on hybrid frequency

• Direct measures of gene flow – Seed collections – Hybrid plant screens

• Modelled gene flow– Pollen dispersal models– Seed dispersal models

Direct measures

• Sampling strategy– Representative sample of the field/population

• Need for markers to handle large numbers– Transgene

– Physiological/ phenotypic screen

– Flow cytometry

• Confirmation of hybrid status– Molecular analysis (microsatellites/ locus-specifc PCR)

Modelling gene flow

• Need pollen dispersal curves

• Population sizes

• Life history details

4. Crop rotation patterns

• Farmer’s records

• Remote sensing

Crop rotation in the target area/nation

2. Quantify Introgression

Locus transmission rates in the field will be influenced by drag imposed by crop genes

• Gametic disequilibrium

• Linkage disequilibrium

Possibly countered by fitness advantage of transgene

Measuring introgression

• Map historic introgression of mapped neutral markers to establish position effects

• Note introgression profiles of mapped markers in glasshouse conditions to study effects of chromosome pairing

3. Transgene spread

• Detailed distribution of recipient

• Proportion of populations exposed to gene flow

• Gene exchange rates between populations

• Demography of recipient

Combine to hybridization introgression and gene spread

data to quantify generic elements of exposure

Stage 3: Quantifying specific exposure elements

• Tiered approach– First tier (worst case scenario conditions)– Second tier (more realistic laboratory

experiments)– Third tier (field/ microcosm experiments)

Losey et al. 1999, Nature

Tier 1

Larvae exposed to GM Bt pollen on milkweed leaves died

Tier 2

Sears et al 2001 PNAS 98 (21): 11937-11942

• Expression levels in most GM maize is low in pollen

• Laboratory experiments exposing larvae to pollen concentrations found in the field showed no acute effects on the larvae

Tier 3

Stanley-Horn etal (2001) PNAS 98: 11931-11936

• Monarch larvae exposed to 3 different Bt pollen sources in field conditions

• Effects varied with event and position

• Negligible effects of Bt11 and Mon810 pollen on larvae survivorship feeding 14 to 22 days on milkweeds in fields

Conclusions

• Quantitative risk assessment require 2 processes – Stage 1: Specify and rank hazards

– Stage 2: Quantify exposure

• Exposure comprises a matrix of linked events• Early elements of exposure relate to hybrid

formation and spread and are largely generic• Late elements are transgene-specific and require a

tiered approach to evaluate

Thanks to

Joel Allainguillaume, Luisa Elliott,

David Mason, Rob Treu, Chris Smith