Noxious, invasive weeds cause billions of dollars per year in damage to the environment. Progress in invasive weed control has been retarded by lack of commercially feasible diet to support mass production of specialist insects that control major noxious weeds. Such damage could be reduced by natural enemies of noxious weeds, but a means to mass produce the insects is not available for economical, large scale releases. Large scale releases would allow more timely reduction of weed problems by allowing weed control experts to release large numbers of weed controlling agents over large geographical regions where weeds are currently a problem. The purpose of this project is to use specific observations and analyses of natural feeding biology of several weed-destroying species to develop economically feasible artificial diets and diet-based rearing systems permitting mass production of weed eaters in much the same way that agricultural pests are mass produced for other programs such as sterile pink bollworm management. Scientifically-based techniques will be developed so that weed eating specialists can be reared in existing commercial facilities. The product of these mass rearing efforts would be available in large numbers and lower costs that will make programs of mass-release possible throughout the habits that are currently being destroyed by the invasive, noxious weeds. OBJECTIVES: I. This first objective is to understand the micro-feeding strategies used by weed insects to determine, using biochemical and histological analysis of pre- and post-feeding sites and of gut contents of recently fed larvae the specific feeding targets used by these insects eating their natural hosts. Development of suitable diets requires a thorough grasp of feeding strategies of weed-eating insects that use a combination of root, stem, or leaf tissue selection, and pin-point feeding, which avoids the reservoirs of toxic weed metabolites. These observations will include A. Histological/histochemical demonstration of micro-feeding by larvae, B. Microscopic determination of plant tissues selected by larvae by examination of gut contents, C. Determination of feeding efficacy on natural diet using the indices: approximate digestibility, efficiency of conversion of ingested food to insect biomass, and efficiency of conversion of digested food to larval biomass. II. The second objective is to develop extraction and stabilization protocols for producing weed flours and liquid extracts, including A. Development of pre-harvest techniques to reduce leaf toxins. B. Trials of best method of desiccation, C. Development of size reduction techniques that yield optimal flour/meal, D. Development of optimal extract or combination of extracts that elicit maximal diet consumption and larval growth indices, includes exclusion of microbial contaminants inherent in plant materials, E. Development of particle coating techniques with various hydrocolloids or gums and other stabilizers, including antioxidants, chelating agents, and emulsifiers, F. Optimization of re-drying and size reduction techniques to stabilize, preserve, and make available the natural nutrient, phagostimulant, and secondary metabolite content of fresh weeds. Objective III is the development of a suitable profile of non-host plant diet components using commercially-available nutrients such as soy and wheat products to replace weed nutrients. This will be accomplished by A. determination of nutrient profiles of natural weed host plants and then B. performing feeding trials with various weed substitutes such as soy flour, wheat germ, wheat bran, desiccated lettuce leaves, etc. IV. The fourth objective is to combine the findings of the first three objectives and synthesizing them into a feeding system for diet and presentation techniques, that at once satisfy the nutritional needs of the weed insects while applying the technology of producing stable and bioavailable diets presented in a context of environmental conditions and cages that encourage growth and reproduction. The two major features of this project are 1.the approach to application of the concept that the target weed-eating species, like most other herbivorous insects, are very specific in their sites of feeding and 2. the application of the principles that the diets MUST satisfy all four concepts of insect diet success: palatability, nutritional completeness, bioavailability, and stability. APPROACH: This project approaches the problem of development of a complete rearing system for several species of weed consuming insects, basing the rearing program on designing an artificial diet that is to contain a specific signature of the natural weed hosts. Along with this, the goal is to provide adequate nutrition derived from commercially and readily available food sources such as soy, wheat germ, and other plant products along with other diet functional components that add palatability, bioavailability, and stability to the diet. These aspects are unique because: 1. Contrary to previous efforts to develop diets for weed eaters, this approach is to recognize that phytophagous insects are highly specialized as to the tissues that they select. Rather than feeding, in general on roots, stems, or leaves, these insects feed on root epidermis, parenchyma, or some other special root, stem or leaf tissue. This project further includes plans to explicitly purify the specific tissues to perform analyses of the nutritional and token stimulant composition of these pin-pointed tissues, then to include in the artificial diet these specific components or designed substitutes for these components. 2. Also, contrary to diet development efforts for weed species, we plan to apply bioassay methods and diet design strategies to optimize the nutritional profile, bioavailability characteristics, and stability of the weed material substitutes. This means that a variety of nutrients will be tested with the specific target insects, comparing such substances as soy flour, wheat germ, wheat bran, wheat flour, lima bean meal, yeast, and a variety of other nutritionally rich components. Because our company has extensive experience with comparing the quality of such nutrient materials and exploring issues of palatability, bioavailability, and stability, we can take the product development far beyond the limitations that have imposed problems on previous efforts at diet development for weed insects. Also, our extensive expertise that comes from our company specialization in diet development and diet and rearing system quality assurance, we can apply special design in processing diet components to eliminate or reduce the contamination issues that profoundly and negatively impact insect diets. The flow of efforts to be undertaken are as follows: 1. the establishment of micro-feeding patterns of each species of weed insects on their host plants, 2 analysis of token stimuli and extraction of these substances, 3 concentration and sterilization of these substances, 4 mixing and testing the extracts with various combinations of commercial components such as soy flour or wheat germ, which is regarded as the integration stage of product development, 5 optimization of the nutrients and weed-derived additives, 6 testing the newly developed diets for micro-feeding characteristics and food efficiency indices, 7 testing insect products on original host plants to determine their ability to revert to target weeds prior to their release, suggesting feasibility of this project for Phase II potential.
