This project proposes to develop a transformation system to express anti-parasitic genes in tsetse flies with the goal of making such an insect population incapable of carrying disease-causing parasites; ie., exhibiting reduced vector-competence. Instead of introducing anti-parasitic genes directly into insect somatic or germ-line cells, genes will be introduced and expressed in the endosymbiotic bacteria in tsetse flies. The symbionts are constantly associated with the insect midgut epithelial cells. During their establishment in tsetse flies, trypanosomes also .invade the midgut epithelial cells where the bacterial symbionts are harbored. Thus expression of anti-parasitic gene products in midgut epithelial cells may be lethal for parasites. The isolation and characterization of the tsetse fly bacterial symbionts as well as the methods and plasmid constructs used for their genetic transformation are described. Initial studies will concentrate on the development of aposymbiotic tsetse flies in order to reconstitute with the genetically altered bacterial symbionts. Anti-parasitic genes that affect the trypanosomes, but not the symbiotic bacteria, will be characterized and expressed in the symbiotic bacteria. Tsetse flies will be reconstituted with the anti-parasitic gene product expressing bacteria and their ability to transmit trypanosomes will be evaluated. This approach can be used in conjunction with other symbiont based methods for spreading genes throughout insect populations.
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