Discovery and functional validation of diapause-related genes in the brown stink bug complex (Hemiptera: Pentatomidae)

The Euschistus (Hemiptera: Pentatomidae) stink bug complex is a significant crop pest in the Americas. They survive harsh winter conditions by entering diapause induced by short days, synchronizing development and reproduction with resource availability, often crops. However, diapause in Euschistus has not been characterized molecularly, and there is a critical need to do so. Thus, we aimed to 1) identify diapause-associated genes from ovaries of E. taurulus via RNA-seq and 2) functionally validate the resulting candidate genes via RNAi, in the Neotropical species Euschistus taurulus, E. crenator, and E. heros, and the Nearctic species E. servus exposed to either short daylength (SD) [10h-light and 14h-dark] or long daylength (LD) [14h-light and 10h-dark]. Employing next-generation sequencing and de novo transcriptome assembly, we successfully identified and validated three candidate genes: nuclear receptor fushi-tarazu factor-1 (ftz-f1), upregulated in SD, juvenile hormone epoxide hydrolase (jheh), upregulated in LD, and farnesyl pyrophosphate synthase (fpps), upregulated in LD. The knockdown of ftz-f1 caused high mortality and induced diapause phenotype under long daylength conditions in Euschistus species. The genes fpps and jheh did not cause significant lethality; however, they induced a diapause phenotype in E. taurulus and E. servus but not in E. heros and E. crenator. We conclude that the selected candidate genes play a crucial role in regulating the diapause phenotype in Euschistus. Furthermore, ftz knockdown leads to high mortality of stink bugs, which could potentially be used to manage Euschistus in the Americas.