Development and Commercialization
of Insect Resistant Transgenic Crops
700 Chesterfield Parkway, North
St. Louis, MO 63918
(prepared from the videotaped presentation*)
Biotechnology, the topic of this presentation, is just one key technology for improving agricultural productivity. Other important techniques are plant breeding, agrochemical discovery, biological control, and farm management.
In agricultural biotechnology, my field of study, insect resistance, is a prime research area that has the potential to greatly improve agricultural productivity. Other agronomic traits to which biotechnology is being applied include viral, bacterial, and fungal resistance, herbicide tolerance, and stress tolerance. Biotechnology is also being applied to post-harvest traits, such as extended shelf life, carbohydrate and protein modification, and improved taste and texture, and to using plants as bioreactors which produce substances such as polymers and antibodies.
Biotechnology allows researchers to make specific, precise changes which are not limited to genes within a species or even to genes of closely related species.
Three areas in which great progress has been made toward bringing insect-protected crops to the commercial phase are
- Transformation/regeneration technology. This includes Agrobacterium transformation and microprojectile bombardment which have allowed major advances. The latter has been particularly important in the transformation of monocots.
- Efficient selection of transformants, for which a selectable marker is needed.
- Modification of Bt endotoxin coding sequences to produce sequences that are better expressed by plant cells. Bt expression in plants has increased dramatically in the past 10 years.
Plants that have been transformed to produce Bt protein include potatoes, cotton, and corn, broccoli, tobacco, sweet potato, tomato, rice, rutabaga, soybean, walnut, poplar, larch, and apple. This list, prepared only weeks ago, is probably already outdated due to the fast pace of research being done. Monsanto has worked on potatoes, corn, and cotton, and I will speak about those.
The major limiting factor for potato-growing in many areas is control of the Colorado potato beetle because it is so resistant to all classes of classical pesticides. The benefits of the product developed by Monsanto are that it provides excellent larvae control - there are essentially no survivors - and it also inihibits reproduction in the adult beetles.
The European corn borer is a major pest of corn. It has two generations per year. One borer per stalk can decrease corn yield 4-7%. Bt corn products control both foliar damage and stalk-boring activities. In addition, there are yield benefits with transgenic plants.
Cotton is one of the most heavily treated (with insecticides) crops in the U.S. Annual costs for control are 200-300 million dollars, and there are also yield losses. The pests are mostly lepidopterous insects and the boll weevil. With transgenic cotton, the larvae die before they reach the 2nd instar.
To produce Bt cotton, the Bt gene is developed and introduced into a cotton plant. The lines with the best resistance are identified. This sometimes takes hundreds or thousands of transformants to get effective expression levels with appropriate and acceptable agronomics. With a plant like cotton, an extensive backcrossing program is necessary to get high yielding cotton lines of interest. There is also lots of regulatory input.
In the field the transgenic cotton plant provides excellent bollworm and budworm control - equivalent to or better than currrent chemical insecticide programs. The transgenic cotton also provides yields equivalent to the best insecticide treatments, and sometimes better.
Bt proteins are generally regarded as safe and have been looked at from a microbial standpoint with no safety issues arising. The issues that have been looked at regarding Bt are food and feed safety, effects on non-target organisms, weediness potential of transgenics, and the potential for pest resistance development.
There are three agencies involved with the regulatory process: USDA, which determines if a transgenic plant is or could become a pest (a "regulated article"), the FDA, which consults on the food and feed safety of transgenic plant products, and the EPA, which judges the environmental safety of transgenic plants and the active ingredients (the proteins). We work primarily with the EPA.
We provide a molecular analysis to the EPA on our products. This includes the genetic elements involved, the DNA inserted into the plant and the proteins produced, and the insertion event.
When one deals with a microprojectile bombardment of a cell, a single copy of the gene doesn't always go into the plant cell cleanly. Sometimes multiple copies result, sometimes partial copies, and these all must be defined and described for the EPA.
I do studies on host range, which is part of "product characterization." We look at the target pest species but also at other insect groups to verify that the proteins are indeed selective. Our challenge is to find good ways to get these proteins into the insect feeding.
Human health related studies.
Other procedures required by the regulatory agency are homology searches. Allergenicity to determine if the Bt protein is like known toxins or allergens is a current focus of much interest in research.
Digestive fate studies.
At Monsanto, we also perform in vitro gastric- and intestinal-fluids studies at different pH's to be sure the proteins will digest. In all these cases it's gone within 30 seconds.
Ecological and environmental risks.
We must determine the potential for adverse effects on non-target organisms: avian, mammalian, aquatic, beneficial insects, and endangered species. What happens to the gene? Does it transfer to other plants? Does it cause problems in the plant itself?
We do biological fate analysis and biochemical fate analysis. What happens to the protein in the environment? Does it accumulate and become a problem to non-targets, especially in the soil? Insects that we test are honeybee (adults and larvae), ladybeetle, hymenoptera (parasitoids), and lacewing.
We do a biochemical analysis of CryIA(b) protein. It degrades extremely rapidly when corn tissue in which it is expressed is brought into contact with the soil.
Insect Resistance Management.
There is much discussion within academic and regulatory circles and also within companies such as Monsanto on the topic of insect resistance. The two approaches currently used are:
- Having the highest dose of protein possible expressed in a plant that is also consistent with good agronomic qualities.
- Combining use of the product with a refugia (alternative hosts and non Bt plants).
Approaches coming along are:
- Use of multiple insect control proteins (pyramiding).
- Education on sound IPM practices.
- Monitoring for resistance.
The optimum dose instead of the highest dose will control all targeted pests and allow minimal survival of resistant insects; resistance is most likely in heterozygous insects, so we want to kill as many of those as possible with the dose expressed in the transgenic plant.
Establishing a refugia will make it likely that the heterozygous insects that develop resistance will mate with homozygous, susceptible insects from the refugia. Therefore, after the selection, the insects will be heterozygous and susceptible to the transgenic plant.
Monitoring is done from a product stewardship standpoint and also to comply with EPA regulations. We have a test that should be able to detect resistant insects before there is a control failure, and other people are developing biochemical techniques as well. These should assist in monitoring. Monitoring tests the effectiveness of resistance management tactics, provides an early warning of impending resistance problems, andevaluates changes in distribution and the severity of the resistance problem.
Multiple insect control proteins (pyramiding) in the same lines is similar doses of at least 2 proteins with different modes of action. This is aimed at redundant killing of insects so that resistance is much less likely to occur. I.e., it is much less likely that the same insect will simultaneously have resistance to both proteins. Pyramiding also increases the total insecticidal protein concentration in the plant.
Affect on other natural enemies.
In one potato field in Oregon, the spider, Nabis, and Geocoris were the 3 most predominant predators. A study showed that plants do not have any morphological changes that will harm natural enemies or reduce their effectiveness. In fact, the natural enemies present more effectively controlled the aphid population than in non-transgenic fields.
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