Title: Strategy for enhanced transgenic strain development for embryonic conditionnal lethality in Anastrepha suspensa Authors
|Schetelig, Marc Florian|
Submitted to: Proceedings of the National Academy of Sciences
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: April 12, 2012
Publication Date: June 12, 2012
Citation: Schetelig, M.A., Handler, A.M. 2012. Strategy for enhanced transgenic strain development for embryonic conditionnal lethality in Anastrepha suspensa. Proceedings of the National Academy of Sciences. 109(24):9348-9353. Interpretive Summary: The ability to create transgenic strains of economically and medically important insect species has the potential to greatly improve existing biological control methods, which is a major goal of our laboratory at the Center for Medical, Agricultural and Veterinary Entomology, USDA, Agricultural Research Service, Gainesville, FL. Development of effective and safe transgenic insect strains for biological control will depend on robust and reliable expression of lethal effectors and rapid evaluation strategies to identify the best systems in the shortest possible time-frame. But thus far, transgenic systems have been developed only in few insect species with development times of 2-3 years in Drosophila melanogaster and the Mediterranean fruitfly Ceratitis capitata. Here, we developed an in-vitro evaluation system (based on functional cell death assays and qPCR) to test transgenic lethality systems. Using the identified “best” components in-vivo in the caribfly A. suspensa, we were able to obtain a mostly embryonic lethal strain within 10 month. This is the first report of an early lethality system in Anastrepha suspensa and the first successful strategy to drastically reduce the developmental time for transgenic insects in general.
Technical Abstract: Here the first reproductive sterility system for the tephritid pest, Anastrepha suspensa, is presented, based on lethality primarily in embryos heterozygous for a lethal conditional transgene combination. The tetracycline-suppressible system uses the cellularization-specific A. suspensa serendipity a gene promoter to drive a phospho-mutated variant of the hid pro-apoptotic cell death gene from A. ludens. The AlhidAla2 variant was validated as having the highest cell death activity compared to orthologous hid genes in cell culture assays. These cell death assays also allowed a determination of the most efficient driver-effector cassette combination for use in A. suspensa transformants, resulting in 100% lethality in first instar larvae (96% in embryos) in the absence of tetracycline. Here we demonstrate that the isolation and in vitro validation of species-specific promoters and lethal effector genes can greatly improve the efficiency of creating high-performance conditional lethality strains, that may be extended to other insect pest species.