Background: Attractive Toxic Sugar Baits (ATSBs) are an innovative vector control strategy based on the “attract-and-kill” principle. The core of ATSBs lies in the preparation of attractive and toxic baits through the mixing and proportioning of luring and active ingredients. Although previous studies have investigated the effects of ATSBs on mosquitoes, significant challenges remain for broader field application. Methods: This study evaluated five fruit juices as ATSBs for mosquitoes, focusing on feeding preferences. Preservative concentrations were assessed by measuring antimicrobial activity over time. Two commercial traps were tested for mosquito entry rates. The optimal insecticide species and concentration were determined based on mortality rates. An optimized ATSBs system was developed and tested under a semi-field cage. Statistical analysis was performed using GraphPad Prism. Results: Within 24 h, apple juice-based ATSBs had the highest attractant index for Culex quinquefasciatus and Anopheles sinensis, while a pear juice-based ATSB was most effective for Aedes albopictus. A 0.1% preservative concentration best maintained juice stability. The LC50 values of dinotefuran-based ATSBs for Cx. quinquefasciatus, Ae. albopictus, and An. sinensis were 1.18 × 10⁻³, 4.06 × 10⁻⁴, and 5.20 × 10⁻⁵ g/L, respectively. The Spodoptera frugiperda trap outperformed the Drosophilidae trap. Simulated semi-field cage tests showed 48 h mortality rates of 86.00% for Cx. quinquefasciatus and 95.67% for Ae. albopictus. Conclusion: This study optimized an ATSB system by screening various fruit juices, preservative concentrations, insecticides, and trap devices. The system’s efficacy in mosquito control was evaluated under a semi-field cage. These findings provide a strong foundation for the future application and refinement of ATSB-based mosquito control strategies.

Identifying conserved RNAi targets that remain effective across mosquito species and resistance backgrounds is important for developing broadly applicable mosquito control tools. Here, an engineered Escherichia coli HT115-L4440 system was used to produce β-tubulin double-stranded RNA (dsRNA), and its larvicidal activity was evaluated in susceptible and pyrethroid-resistant strains of Culex quinquefasciatus and Aedes albopictus. Recombinant bacteria produced species-specific β-tubulin dsRNA fragments of approximately 360 bp. Immersion treatment caused high larval mortality in both species. In Cx. quinquefasciatus, mortality reached 85.00% and 89.00% in susceptible and resistant strains, respectively, compared with 8.00% and 10.00% in controls, while β-tubulin transcript abundance showed maximum reductions of 571-fold and 333-fold, respectively. In Ae. albopictus, mortality reached 91.00% and 82.00% in susceptible and resistant strains, compared with 9% and 8% in controls, with maximum reductions in β-tubulin expression of 157-fold and 337-fold, respectively. These comparable effects suggest that β-tubulin dsRNA is not compromised by conventional pyrethroid-resistance mechanisms. Transcriptomic and proteomic analyses showed that β-tubulin knockdown disrupted key cellular processes, including cytoskeletal regulation, metabolism, protein synthesis, intracellular transport, detoxification, oxidative stress, DNA replication, and autophagy. Overall, engineered bacteria-derived β-tubulin dsRNA induced consistent gene silencing and larval mortality in both susceptible and resistant mosquito larvae, supporting β-tubulin as a conserved candidate RNAi target for mosquito control.