Development and evaluation of screwworm, <i>Cochliomyia hominivorax</i> (Diptera: Calliphoridae), transgenic sexing strains with embryonic gene promoters for a genetic control program

The New World screwworm, Cochliomyia hominivorax (Coquerel, 1858; Diptera: Calliphoridae), was eradicated from North and Central America through the first application of the sterile insect technique. The sterile screwworm adult fly releases were mixed sex, but releasing only males could be much more effective. Here, we describe screwworm transgenic sexing strains (TSSs) with expected embryo lethality. Strains were developed with a Tetracycline-off (Tet-off) system utilizing either the Lucilia cuprina (Wiedemann, 1830) nullo (DR6) or Cochliomyia macellaria (Fabricius, 1755) CG14427 (DR7) gene promoter and a tTA-activated effector to promote female lethality. The promoter activity was highest in 2 to 3 h embryos and low in larvae and adults. However, these strains also had unexpected high expression in pupae. Evaluation on two doxycycline (Dox) suppression regimens found that inclusion of Dox in the last larval feeding rescued females of the subsequent generation, likely by maternal transfer of Dox. All TSSs produced only males on a reduced Dox feeding regimen, but the female lethal period for the DR6 TSS was too late in development to reduce diet costs. Production parameters were met by all strains in colony, but strains had lower male fly survival than current production strains after removing Dox. In noncompetitive mating success trials, DR6 strains performed poorly, but DR7 performed equally to production strain males. However, males from all TSSs fared poorly in mating competition tests against production males. Our study highlights the importance of tightly regulated gene promoters and stage-specific antibiotic feeding schemes for the development and evaluation of TSSs based on the Tet-off system.