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Dengue, Zika, chikungunya and yellow fever viruses continue to be a major public health burden. Aedes mosquitoes, the primary vectors responsible for transmitting these viral pathogens, continue to flourish due to local challenges in vector control management. Yeast interfering RNA-baited larval lethal ovitraps are being developed as a novel biorational control tool for Aedes mosquitoes. This intervention circumvents increasing issues with insecticide resistance and poses no known threat to non-target organisms. In an effort to create public awareness of this alternative vector control strategy, gain stakeholder feedback regarding product design and acceptance of the new intervention, and build capacity for its potential integration into existing mosquito control programs, this investigation pursued community stakeholder engagement activities, which were undertaken in Trinidad and Tobago. Three forms of assessment, including paper surveys, community forums, and household interviews, were used with the goal of evaluating local community stakeholders' knowledge of mosquitoes, vector control practices, and perceptions of the new technology. These activities facilitated evaluation of the hypothesis that the ovitraps would be broadly accepted by community stakeholders as a means of biorational control for Aedes mosquitoes. A comparison of the types of stakeholder input communicated through use of the three assessment tools highlighted the utility and merit of using each tool for assessing new global health interventions. Most study participants reported a general willingness to purchase an ovitrap on condition that it would be affordable and safe for human health and the environment. Stakeholders provided valuable input on product design, distribution, and operation. A need for educational campaigns that provide a mechanism for educating stakeholders about vector ecology and management was highlighted. The results of the investigation, which are likely applicable to many other Caribbean nations and other countries with heavy arboviral disease burdens, were supportive of supplementation of existing vector control strategies through the use of the yeast RNAi-based ovitraps., Competing Interests: I have read the journal’s policy and the authors of this manuscript have the following competing interests: Scheel and Severson are inventors on a related patent submission, US patent application 62/361,704 / European patent application EP 17828458.4. This does not alter their adherence to PLOS ONE policies on sharing data and materials. The other authors have no competing interests.

Increasing reports of insecticide resistance continue to hamper the gains of vector control strategies in curbing malaria transmission. This makes identifying new insecticide targets or alternative vector control strategies necessary. CLassifier of Essentiality AcRoss EukaRyote (CLEARER), a leave-one-organism-out cross-validation machine learning classifier for essential genes, was used to predict essential genes in Anopheles gambiae and selected predicted genes experimentally validated. The CLEARER algorithm was trained on six model organisms: Caenorhabditis elegans, Drosophila melanogaster, Homo sapiens, Mus musculus, Saccharomyces cerevisiae and Schizosaccharomyces pombe, and employed to identify essential genes in An. gambiae. Of the 10,426 genes in An. gambiae, 1,946 genes (18.7%) were predicted to be Cellular Essential Genes (CEGs), 1716 (16.5%) to be Organism Essential Genes (OEGs), and 852 genes (8.2%) to be essential as both OEGs and CEGs. RNA interference (RNAi) was used to validate the top three highly expressed non-ribosomal predictions as probable vector control targets, by determining the effect of these genes on the survival of An. gambiae G3 mosquitoes. In addition, the effect of knockdown of arginase (AGAP008783) on Plasmodium berghei infection in mosquitoes was evaluated, an enzyme we computationally inferred earlier to be essential based on chokepoint analysis. Arginase and the top three genes, AGAP007406 (Elongation factor 1-alpha, Elf1), AGAP002076 (Heat shock 70kDa protein 1/8, HSP), AGAP009441 (Elongation factor 2, Elf2), had knockdown efficiencies of 91%, 75%, 63%, and 61%, respectively. While knockdown of HSP or Elf2 significantly reduced longevity of the mosquitoes (p<0.0001) compared to control groups, Elf1 or arginase knockdown had no effect on survival. However, arginase knockdown significantly reduced P. berghei oocytes counts in the midgut of mosquitoes when compared to LacZ-injected controls. The study reveals HSP and Elf2 as important contributors to mosquito survival and arginase as important for parasite development, hence placing them as possible targets for vector control. [ABSTRACT FROM AUTHOR]