Transgenic Cotton for Insect Control Peter C Ellsworth

Transgenic Cotton for Insect Control Peter C. Ellsworth, Ph. D. IPM Specialist, University of Arizona Maricopa Agricultural Center Maricopa, AZ, USA Ellsworth/UA

Disclosure • Those engaged in the dialog on biotechnology should fully disclose their relationships and opinions “up front” so that audiences can consider the context. • Partial support for my research comes from companies with interests in biotechnology. • The balance of support comes from state and federal sources of competitively available public funds. Ellsworth/UA

Disclosure (continued) • Biotechnology and its products are neither inherently good nor bad. • The specific process and each of its products should be scientifically and independently evaluated. Ellsworth/UA

Transgenic Cotton for Insect Control • What is available now & in the future? • Origin, identity & development • Insect target(s) in the U. S. • Efficacy & utility in the Arizona system (benefits) • Safety (risks) – – Resistance Impact of gene on plant Biodiversity non-target effects Ellsworth/UA

Products Available for Cotton Insect Control • Only 1 ‘trans’-gene has been commercialized • Based on the crystalline protein produced by Bacillus thuringiensis (Bt) • Developed by Monsanto as Bollgard® and incorporated into commercial varieties by several cotton seed companies (e. g. , Delta Pineland Co. & Stoneville Pedigreed Seed Co. ) • Sold in the U. S. , Australia, Mexico, South Africa, India, China, Argentina, Indonesia Ellsworth/UA

Bacillus thuringiensis (Bt) • Common soil bacterium • Present in nature in a variety of forms (species & strains) • Produces proteins that are toxic to insects • Commonly used in garden sprays & for commercial agriculture, including organic farming • Extremely well-known toxin in terms of human health & environmental safety Ellsworth/UA

Bacillus thuringiensis (Bt) • Crystalline proteins are classified according to structure & have a specific nomenclature (e. g. , Cry 1 Ac) • Cotton has been transformed with Cry 1 Ac (narrow spectrum; Lepidoptera only) • Protein binds with receptors in the insect gut causing pores which perforate the midgut & lead to cell leakage & insect death Ellsworth/UA

The Transformation • The gene of interest is spliced Coker 312 out of the bacterium using a vector, like Agrobacterium tumefasciens, & transferred to cotton cells grown in tissue culture • The cells are grown into a plant & then, after testing, plants are back-crossed into commercial lines to make new varieties Recurrent back-crossing Ellsworth/UA

Spectrum of Activity for BG Excellent Control Tobacco No Budworm, the Control Trichoplusia niprincipal pest Spodoptera exigua in the South Heliothis virescens Spodoptera frugiperda Spodoptera ornithogalli Pectinophora gossypiella Bucculatrix thurberiella Pink Bollworm Estigmene acrea (PBW), our Helicoverpa zea principal pest (pre-bloom) Marmara spp. Helicoverpa zea (post-bloom) Beneficial Insects Agrotis & Feltia spp. Pseudoplusia includens Ellsworth/UA

AZ’s Primary Lepidopteran Pest • Pink Bollworm • Multiple generations • Adult lays eggs on bolls or susceptible squares (SS) • Larvae hatch & penetrate bolls within 24 hrs Ellsworth/UA

Alternatives for PBW Control • Repeated, broad-spectrum sprays are required to prevent moths from invading fields • No effective larvicides or ovicides • Biological controls are limited by the biology of this pest – Little impact of parasitoid or predators • Cultural controls can be very effective – Requires early termination & areawide compliance with plowdown requirements Ellsworth/UA

Secondary Lepidopteran Pests • Occasional pests Helicoverpa zea Heliothis virescens Trichoplusia ni Estigmene acrea (Arctiidae) • Induced pests Spodoptera exigua Bucculatrix thurberiella Ellsworth/UA

Bt Cotton Questions • Efficacy & economic studies – How effective is the gene? – Are oversprays required for lepidopteran control? – If so, are there new scouting & threshold considerations? Ca. 100% for PBW Not for PBW Search for large larvae • Agronomic studies – Impacts (+/-) on yield & fiber qualities? No unintended effects • Product integrity & stability studies – High-dose through life of plant? – High-dose in all varieties? – Purity? Yes, actively growing No, some not marketed > 98% (? ) • Ecological studies – Impact on non-target organisms (NTO) No unintended effects Ellsworth/UA

BG Cotton Efficacy • Young larvae present regardless of cotton type • Little difference between Bt & non-Bt (-) varieties Ellsworth/UA

BG Cotton Kills Small Larvae • PBW larvae must feed in order to be killed. • Large larvae survive mainly in non-Bt varieties. Ellsworth/UA

Impact on Arizona Cotton • In 1990, > 6. 8 sprays were made against PBW; still, > 5% yield loss • Since 1996 when Bt cotton was introduced, it has never required oversprays for PBW control, AND • Since 1997, only 0. 5 sprays have been made against PBW over all cotton acreage (Bt and non. Bt); i. e. , an areawide reduction of PBW has occurred • The net reduction in insecticide use has resulted in huge savings to farmers, and large improvements to the agroecosystem in terms of beneficial insect communities & IPM Ellsworth/UA

Safety - Resistance • Given time & exposure, insects have the capacity to overcome most insecticides. Bt cotton may be no different, however, there are safeguards: • Refugia • High-Dose Strategy • Development of additional proteins Ellsworth/UA

Refugia • Objective: provide harborage for susceptible moth production to reduce the chance of resistant (R) moths mating with each other RR RR RR SS RS SS RR • U. S. growers are required to plant a proportion of their acreage to non-Bt cotton – 5% Refuge, if no lepidopteran-active insecticides are used on it, or else – 20% Refuge Ellsworth/UA

High-Dose Strategy, Depends on: • The production of a dose high enough to kill: >99. 9% of a susceptible (SS) population, and Yes >95% of the heterozygous (RS) individuals, Yes? • A recessive resistance, Yes • Random mating, Yes, *refuges • A low initial frequency of the ‘R’ allele. No (? ) Ellsworth/UA

Development of Additional Transgenes (Bt’s) • Bollgard II® – 2 Bt gene product, original Bollgard (Cry 1 Ac) + Cry 2 Ab – Final stages of US-EPA approval – Limited commercial production in 2003 – Full replacement of BG varieties by 2008? • Bollgard III – Little information on this available at this time; research stages only • Cry 1 F – Under development by Dow Agrosciences in combination with Cry 1 Ac Ellsworth/UA

Impact of Gene on Plant • Isogenic lines were developed for testing the impact of the gene(s) on agronomic and efficacy characteristics of the plant C 312 B DP 50 Particle gun DP 50 B DP 50 II (Cry 1 Ac) (Cry 1 AC+Cry 2 Ab) DP 50 Cry 1 Ac Cry 2 Ab Lines 1. Cry 1 Ac+ Cry 2 Ab 2. Cry 1 Ac only 3. Cry 2 Ab only 4. Null Ellsworth/UA

Isoline Studies of BG & BGII • Replicated studies • Artificial & natural PBW infestations • Sprayed & Unsprayed conditions Ellsworth/UA

Warts are often formed at the site of PBW attack Dead 1 st instar in Bt cotton Ellsworth/UA

BGII Results - PBW, 1 st Instars Dead 1 st Instars Live 1 st Instars Ellsworth/UA

BGII Results - PBW, All Instars Ellsworth/UA

BGII Results - B. thurberiella • BGII prevented cotton leafperforator development better than BG • Leaves at top of plant (younger) express highest doses of Bt • Older leaves (bottom) have reduced doses of Bt Ellsworth/UA

Marmara sp. • Citrus Peel Miner is an incidental lepidopteran that mines the main stem and boll surfaces • Cry 2 Ab alone (‘X’) is more effective than Cry 1 Ac (‘B’) Ellsworth/UA

Spectrum of Activity for BG (Cry 1 Ac) Excellent Control No Control Pectinophora gossypiella Heliothis virescens Bucculatrix thurberiella Marmara spp. Helicoverpa zea Estigmene acrea Trichoplusia ni Beneficial Insects Agrotis & Feltia spp. Pseudoplusia includens Spodoptera exigua Spodoptera frugiperda Spodoptera ornithogalli Ellsworth/UA

Spectrum of Activity for BGII (Cry 1 Ac + Cry 2 Ab) Excellent Control Pectinophora gossypiella Heliothis virescens Bucculatrix thurberiella Marmara spp. Helicoverpa zea Estigmene acrea Trichoplusia ni Pseudoplusia includens No Control Beneficial Insects Agrotis & Feltia spp. Spodoptera exigua Spodoptera frugiperda Spodoptera ornithogalli Ellsworth/UA

High Dose and % Efficacy? • Throughout our early work with BG cotton, we often would find low levels of “survivors” from our field plots Ellsworth/UA

Source of Survivors • Low expression of Bt in plants? • Low levels of non-Bt contaminants? – In the seedbag – From volunteer seed • Resistance? Ellsworth/UA

Before plants are tested for presence of Bt After PBW from non-Bt plants are discarded % Efficacy Against PBW Cry 1 Ac 100% Cry 2 Ab 99. 67% Both Genes 100% Ellsworth/UA

Biodiversity / NTO Studies • The reports of Bt effects on Monarch butterflies have fueled much emotional debate on the use of biotech crops. • Monarch Butterfly, symbol of nature and “wildness” in North America. Ellsworth/UA

Non-Target Organisms (NTO) • Over 370 arthropod species have been tracked in 2 years of field studies using a variety of methodologies. • So far, no major or functional differences have been found in Arizona between BG, BGII, and conventional cotton communities… • Except where harsh PBW sprays are needed in conventional cottons. • Thus, Bt cotton ecosystems are not only safe, but safer than conventional cotton ecosystems where insecticidal inputs are higher. Ellsworth/UA

Conclusions • The use of Bt cottons in Arizona has provided the first larvicidal and selective approach to controlling PBW. • The control provided by Bt cottons approaches immunity. No survivors have been found in field studies. • Bt cotton has revolutionized our ability to implement IPM in AZ cotton & reduced our insecticide inputs by over 60%. • Future transgenic products for insect control in cotton should be independently & scientifically tested. • Other than new Bt genes/events, there are few, if any, development plans for insect contol products. Ellsworth/UA

Information • All University of Arizona crop production & crop protection information is available on our web site, A C I S • Arizona Crop Information Site (ACIS), at • http: //ag. arizona. edu/crops Ellsworth/UA