The high reproductive performance of Nilaparvata lugens (Stål) (Hemiptera: Delphacidae) is one of the key factors contributing to its serious damage. Studies have demonstrated that trehalose hydrolysis, catalyzed by trehalase (TRE), plays an important role in the reproductive regulation of N. lugens. However, it remains unclear how TRE regulates the reproduction of N. lugens. To address this question, the current study was designed to investigate the effects of TRE on the reproductive regulatory network of N. lugens. Specifically, RNA interference (RNAi) was conducted by injecting double-stranded RNA (dsRNA) targeting the TRE genes, and a trehalase inhibitor, validamycin, was used to suppress TRE activity. Subsequently, the relative expression levels of key genes involved in the reproductive regulatory pathway, as well as the triglyceride content in the fat body and ovary, were determined. The results indicated that both dsTREs and validamycin treatment decreased the relative expression of Vitellogenin (Vg) without affecting the expression of its receptor, VgR. However, their impact on the reproductive regulatory network differed: dsTREs injection did not affect the relative expression of JHAMT and Met in the juvenile hormone signaling pathway, but resulted in the upregulation of USP and TOR, and inhibition of InR2 and S6K. In contrast, validamycin injection inhibited the expression of Met, USP, EcR, InR1, TOR, and S6K. Both dsTREs and validamycin inhibited the expression of Fas, but only validamycin decreased the ovarian triglyceride content. These findings suggest that the inhibition of TRE leads to alterations in reproductive regulatory signals, thereby reducing the reproductive capacity of N. lugens.

Spodoptera frugiperda (J. E. Smith, 1797), an omnivorous crop pest worldwide, reproduces prolifically. Validamycin, a competitive natural inhibitor of trehalase, is regarded as an effective and safe insecticide. Pupae were injected with a validamycin gradient (0.5–10 µg/µL) to block trehalase; enzyme activity and the Vitellogenin gene (Vg)/its receptor gene (VgR) expression (rpL10 reference) were subsequently quantified to determine the compound’s impact on S. frugiperda ontogeny and fecundity. Validamycin directly inhibited pupal membrane-bound trehalase, sharply lowering glycogen. Both pupal and adult mortality rose with dose, yielding marked abnormalities versus the Control Check (CK) group. At 0.5 μg/μL validamycin, eggs blackened and clumped in the lateral oviduct, blocking release; treated females produced far fewer eggs by day 4, exhibited ovarian atrophy, shorter lifespan, and low hatchability. The expression levels of Vg and VgR in the ovaries of the fall armyworm were consistent with the changes in the ovarian developmental phenotype. Validamycin significantly inhibited the activity of trehalase in S. frugiperda, severely hindering their normal eclosion and lowering the potential reproductive capacity of S. frugiperda. Simultaneously, it directly affects ovarian development and the lifespan of female moths. The results provide data to support the development of new methods for controlling S. frugiperda.