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Brenda Oppert  - Research Summaries     [back]

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Current Research Projects

Functional Genomics Applied To Stored Product Pest Problems
New insect control products for storage pests are urgently needed to replace those that are undergoing deregulation or to circumvent resistant pest populations. With the sequencing of the red flour beetle genome, we now have new tools to develop control products based on vulnerabilities in the biology of the beetle. We developed microarrays that contain representative sequences from each gene in the red flour beetle genome to identify sequences that are highly expressed in a particular tissue. Initial studies have identified gut transcripts as potential new targets for biopesticides. Beetle genome microarrays are also being used to probe the beetle's response to dietary inhibitors, toxins, and current control products, such as phosphine. We've found that beetle larvae fed dietary inhibitors mount a complex genetic response to survive. This information has provided us with details about genes that are transcribed by the beetle to survive inhibitor damage, and also genes that encode enzymes critical to digestion of protein in the diet. Because a closely related beetle, the yellow mealworm, has no sequenced genome, we are using high throughput sequencing to identify highly expressed genes in the gut of mealworm larvae. By comparing genetic transcripts in mealworm larvae exposed or not to a bacterial toxin, we are beginning to understand the complex response of mealworm larvae to toxins. All functional genomics applications are highly dependent on sophisticated bioinformatics algorithms, which we are developing to analyze insect sequences and data. As we identify potential new targets for biopesticides, we are using high throughput bioassays to evaluate purified proteins, peptides, or a process of interfering in the synthesis of genetic transcripts, called RNA interference. Biopesticides have applications in the integrated pest management of storage pests in sprays, formulations, or transgenic cereals. (Cooperators: Drs. Jeff Lord, Jeff Fabrick, Juan Luis Jurat-Fuentes, Elena Elpidina, John Tomich)

Discovery of More Effective Insect Control Proteins
Proteins that negatively impact insect growth and development are useful in insect control, because genes encoding insecticidal proteins can be expressed in plants to enhance host plant resistance. Proteinase inhibitors are good candidates because they disrupt insect digestion. However, insects can compensate for proteinase inhibitors by increasing the expression of digestive proteinases and/or expressing inhibitor-insensitive proteinases. Dr. Oppert was part of a team that discovered that combinations of digestive inhibitors targeting different proteinase classes provided better control of some stored product pests. Testing of other proteins by the research group led to the identification of vitamin-binding proteins that were also effective in the control of some stored product pests. Genes for these proteins can be expressed in wheat, maize, and other cereals to reduce pest damage to these products.

Biochemical Techniques Developed To Evaluate Proteinases In Mixtures
Insect proteinases are being investigated to identify those that may be targeted by biopesticides. The study of stored product insect proteinases is complicated by the small size of the insect, and the collection of sufficient quantities of digestive proteinases for purification is problematic. Dr. Oppert developed two major techniques that facilitate the analysis of complex mixtures of proteinases. One technique is a fast, simple, and economical microplate assay (see Oppert et al., 1997b). The other technique provides both a qualitative and quantitative identification of proteinases in the mixture (see Oppert and Kramer, 1998). These techniques have promoted rapid and efficient identification of digestive proteinases in moth and beetle pests of stored products. Researchers from areas other than agriculture have used these assays to study proteinases prior to purification.

Elucidation of Insect Resistance To Bacillus Thuringiensis
Proteinaceous toxins from the bacterium Bacillus thuringiensis (Bt) have been used for years in spray applications to control insect pests. Genes encoding these toxins are now being expressed in plants to control crop pests. Expression of Bt toxins in plants will increase exposure levels to insects for longer periods, thus providing increased selection pressure for the survival of resistant populations of pests. Insects can adapt to Bt toxins through an alteration in the gut receptor that binds the toxin in the early stages of toxicity. Through her work with digestive proteinases of the Indianmeal moth, a major pest of stored products, Dr. Oppert described a novel mechanism of insect resistance to Bt. Some Bt-resistant Indianmeal moths have reduced digestive proteinase activity that enables them to survive on diets containing Bt toxins. This was the first evidence of multiple adaptations in insects that result in a loss of Bt toxin efficacy. Information from this research is being used in the design of more efficient microbial toxins and in the development of effective resistance management policies.