Strategy for enhanced transgenic strain development for embryonic conditional lethality in Anastrepha suspensa

Abstract

Here the first reproductive sterility system for the tephritid fruit fly pest, Anastrepha suspensa, is presented, based on lethality primarily limited to embryos heterozygous for a conditional lethal transgene combination. This tetracycline (Tet)-suppressible system uses a driver construct having the promoter from the newly isolated embryo-specific A. suspensa serendipity α gene linked to the Tet-transactivator. This was used to drive expression of a phosphomutated variant of the pro-apoptotic cell death gene, hid, from A. ludens, that was isolated, based on its identity to A. suspensa hid. The Alhid Ala2 variant was shown to have the highest cell death activity in an in vitro A. suspensa cell death assay compared to the orthologous genes Ashid, Dmhid, and the variant Dmhid Ala5 . These cell death assays also allowed a determination of the most-efficient driver-effector cassette combinations for use in A. suspensa transformants, resulting in two hybrid strains exhibiting 100% lethality. One strain was 96% lethal in embryos in the absence of tetracycline, with none surviving past the first larval instar, which is critical for pests that are most damaging in late-larval stages. 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.apoptosis | insect pest management | sterile insect technique

The Anopheles arabiensis genetic sexing strain ANO IPCL1 and its application potential for the sterile insect technique in integrated vector management programmes

“…In the molecular approaches, priority is being given to develop transgenic sexing through a conditional lethal system which would eliminate females as early as possible during development, preferably at the embryonic stage. Similar systems have been developed in fruit flies (Franz, 2005;Heinrich and Scott, 2000;Horn and Wimmer, 2003;Schetelig and Handler, 2012;Ogaugwu et al, 2013).…”

Cloning and functional characterizations of an apoptogenic Hid gene in the Scuttle Fly, Megaselia scalaris (Diptera; Phoridae)

“…2). In contrast, AsHID contains only two such sites and elimination of them enhanced slightly its killing activity in a cell-based assay (Schetelig and Handler, 2012). There is only one putative MAPK phosphorylation site (PPSP) found in MsHID, which might also contribute to the cell death resulting from the deprivation of survival signal.…”

The Orthologue of the Fruitfly Sex Behaviour Gene Fruitless in the Mosquito Aedes aegypti: Evolution of Genomic Organisation and Alternative Splicing

“…aegypti (AAEL009224, AAEL006416, AAEL009222 and AAEL004293) as well as two putative homologs within the genome of An. gambiae (AGAP006798 and a second unannotated putative tra-2 homolog, identified starting from the EST gb|BM620287.1) The presence of multiple copies of the tra-2 gene in these two dipteran genomes is a new feature, since tra-2 is a single copy gene in Drosophilidae and apparently also in Tephritidae [75], [77], [78].…”