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Information about genetic engineering (GE) for vector control in the United States is disseminated primarily in English, though non-English speakers are equally, and in some geographic regions even more affected by such technologies. Non-English-speaking publics should have equal access to such information, which is especially critical when the technology in question may impact whole communities. We convened an interdisciplinary workgroup to translate previously developed narrated slideshows on gene drive mosquitoes from English into Spanish, reviewing each iteration for scientific accuracy and accessibility to laypeople. Using the finalised stimuli, we conducted five online, chat-based focus groups with Spanish-speaking adults from California. Overall, participants expressed interest in the topic and were able to summarise the information presented in their own words. Importantly, participants asked for clarification and expressed scepticism about the information presented, indicating critical engagement with the material. Through collaboration with Spanish-speaking scientists engaged in the development of GE methods of vector control, we translated highly technical scientific information into Spanish that successfully engaged Spanish-speaking participants in conversations about this topic. In this manuscript, we document the feasibility of consulting Spanish-speaking publics about a complex emerging technology by drawing on the linguistic diversity of the scientific teams developing the technology. [ABSTRACT FROM AUTHOR]

Aedes albopictus mosquitoes are important vectors for a number of human pathogens including the Zika, dengue, and chikungunya viruses. Capable of displacing Aedes aegypti populations, this mosquito adapts to cooler environments which increases its geographical range and transmission potential. There are limited control strategies for Aedes albopictus mosquitoes which is likely attributed to the lack of comprehensive biological studies on this emerging vector. To fill this void, here using RNAseq we characterized Aedes albopictus mRNA expression profiles at 34 distinct time points throughout development providing the first high-resolution comprehensive view of the developmental transcriptome of this worldwide human disease vector. This enabled us to identify several patterns of shared gene expression among tissues as well as sex-specific expression patterns. To illuminate the similarities and differences with Aedes aegypti, a related human disease vector, we also performed a comparative analysis between the two developmental transcriptomes, identifying life stages where the two species exhibit similar and distinct gene expression patterns. These findings provide insights into the similarities and differences between Aedes albopictus and Aedes aegypti mosquito biology. In summary, the results generated from this study should form the basis for future investigations on the biology of Aedes albopictus and provide a gold mine resource for the development of transgene-based vector control strategies. [ABSTRACT FROM AUTHOR]

With dengue virus (DENV) becoming endemic in tropical and subtropical regions worldwide, there is a pressing global demand for effective strategies to control the mosquitoes that spread this disease. Recent advances in genetic engineering technologies have made it possible to create mosquitoes with reduced vector competence, limiting their ability to acquire and transmit pathogens. Here we describe the development of Aedes aegypti mosquitoes synthetically engineered to impede vector competence to DENV. These mosquitoes express a gene encoding an engineered single-chain variable fragment derived from a broadly neutralizing DENV human monoclonal antibody and have significantly reduced viral infection, dissemination, and transmission rates for all four major antigenically distinct DENV serotypes. Importantly, this is the first engineered approach that targets all DENV serotypes, which is crucial for effective disease suppression. These results provide a compelling route for developing effective genetic-based DENV control strategies, which could be extended to curtail other arboviruses. Author summary: With limited success of traditional vector control methods to curb dengue infections and more than half of the world's population still at risk, there is a need for novel strategies to reduce its impact on public health. Recent advances in genetic technologies has allowed for precise modifications of mosquito genome to make them resistant to infections, thus breaking the transmission cycle. Here we generated engineered Ae. aegypti mosquitoes efficiently expressing a DENV-targeting single-chain variable fragment (scFv) derived from a previously characterized broadly neutralizing human antibody, which blocked infection and transmission in these mosquitoes. To our knowledge, this is the first example of an engineered transgene capable of rendering Ae. aegypti mosquitoes 100% refractory to all four serotypes of DENV. The engineered mosquitoes, in future, could easily be paired with a gene drive, capable of spreading the transgene throughout wild disease-transmitting mosquito populations and preventing further DENV transmission. Since a number of diverse and well-characterized antibodies exist against other arboviruses (eg chikungunya and Zika, this work also provides a proof-of-concept principle for developing similar genetic strategies for reducing the impact of these arboviruses. [ABSTRACT FROM AUTHOR]

Recent Zika virus (ZIKV) outbreaks have highlighted the necessity for development of novel vector control strategies to combat arboviral transmission, including genetic versions of the sterile insect technique, artificial infection with Wolbachia to reduce population size and/or vectoring competency, and gene drive-based methods. Here, we describe the development of mosquitoes synthetically engineered to impede vector competence to ZIKV. We demonstrate that a polycistronic cluster of engineered synthetic small RNAs targeting ZIKV is expressed and fully processed in Aedes aegypti, ensuring the formation of mature synthetic small RNAs in the midgut where ZIKV resides in the early stages of infection. Critically, we demonstrate that engineered Ae. aegypti mosquitoes harboring the anti-ZIKV transgene have significantly reduced viral infection, dissemination, and transmission rates of ZIKV. Taken together, these compelling results provide a promising path forward for development of effective genetic-based ZIKV control strategies, which could potentially be extended to curtail other arboviruses. [ABSTRACT FROM AUTHOR]

In Fig 1B, the columns corresponding to infection with DENV-3 and DENV-4 are duplicated and both denote the DENV-4 data. Reference 1 Buchman A, Gamez S, Li M, Antoshechkin I, Li H-H, Wang H-W, et al. (2020) Broad dengue neutralization in mosquitoes expressing an engineered antibody. [Extracted from the article]

Anopheles gambiae is amajor mosquito vector ofmalaria in Africa. Although increased use of insecticide-based vector control tools hasdecreased malaria transmission, elimination is likely torequire novelgenetic control strategies. It canbearguedthat theabsenceof an A. gambiae inbred line has slowed progress toward genetic vector control. In order to empower genetic studies and enable precise and reproducible experimentation, we set out to create an inbred line of this species. We found that amenability to inbreeding varied between populations of A. gambiae. After full-sib inbreeding for ten generations, we genotyped 112 individuals-56 saved prior to inbreeding and 56 collected after inbreeding-at a genome-wide panel of single nucleotide polymorphisms (SNPs). Although inbreeding dramatically reduced diversity across much of the genome, we discovered numerous, discrete genomic blocks that maintained high heterozygosity. For one large genomic region, we were able to definitively show that high diversity is due to the persistent polymorphismof a chromosomal inversion. Inbred lines in other eukaryotes often exhibit a qualitatively similar retention of polymorphism when typed at a small number ofmarkers. Our whole-genome SNP data provide the first strong, empirical evidence supporting associative overdominance as the mechanism maintaining higher than expected diversity in inbred lines. Although creation of A. gambiae lines devoid of nearly all polymorphism may not be feasible, our results provide critical insights into how more fully isogenic lines can be created. [ABSTRACT FROM AUTHOR]

Animals face the dual threat of virus infections hijacking cellular function and transposons proliferating in germline genomes. For insects, the deeply conserved RNA interference (RNAi) pathways and other chromatin regulators provide an important line of defense against both viruses and transposons. For example, this innate immune system displays adaptiveness to new invasions by generating cognate small RNAs for targeting gene silencing measures against the viral and genomic intruders. However, within the Dipteran clade of insects, Drosophilid fruit flies and Culicids mosquitoes have evolved several unique mechanistic aspects of their RNAi defenses to combat invading transposons and viruses, with the Piwi-piRNA arm of the RNAi pathways showing the greatest degree of novel evolution. Whereas central features of Piwi-piRNA pathways are conserved between Drosophilids and Culicids, multiple lineage-specific innovations have arisen that may reflect distinct genome composition differences and specific ecological and physiological features dividing these two branches of Dipterans. This perspective review focuses on the most recent findings illuminating the Piwi/piRNA pathway distinctions between fruit flies and mosquitoes, and raises open questions that need to be addressed in order to ameliorate human diseases caused by pathogenic viruses that mosquitoes transmit as vectors. [ABSTRACT FROM AUTHOR]

Ongoing speciation in the most important African malaria vectors gives rise to cryptic populations, which differ remarkably in their behavior, ecology, and capacity to vector malaria parasites. Understanding the population structure and the drivers of genetic differentiation among mosquitoes is crucial for effective disease control because heterogeneity within vector species contributes to variability in malaria cases and allow fractions of populations to escape control efforts. To examine population structure and the potential impacts of recent large-scale control interventions, we have investigated the genomic patterns of differentiation in mosquitoes belonging to the Anopheles nili group-a large taxonomic group that diverged ~3 Myr ago. Using 4,343 single nucleotide polymorphisms (SNPs), we detected strong population structure characterized by high- FST values between multiple divergent populations adapted to different habitats within the Central African rainforest. Delineating the cryptic species within the Anopheles nili group is challenging due to incongruence between morphology, ribosomal DNA, and SNP markers consistent with incomplete lineage sorting and/or interspecific gene flow. A very high proportion of loci are fixed ( FST = 1) within the genome of putative species, which suggests that ecological and/or reproductive barriers are maintained by strong selection on a substantial number of genes. [ABSTRACT FROM AUTHOR]

Malaria vectors are exposed to intense selective pressures due to large-scale intervention programs that are underway in most African countries. One of the current priorities is therefore to clearly assess the adaptive potential of Anopheline populations, which is critical to understand and anticipate the response mosquitoes can elicit against such adaptive challenges. The development of genomic resources that will empower robust examinations of evolutionary changes in all vectors including currently understudied species is an inevitable step toward this goal. Here we constructed double-digest Restriction Associated DNA (ddRAD) libraries and generated 6461 Single Nucleotide Polymorphisms (SNPs) that we used to explore the population structure and demographic history of wild-caught Anopheles moucheti from Cameroon. The genome-wide distribution of allelic frequencies among samples best fitted that of an old population at equilibrium, characterized by a weak genetic structure and extensive genetic diversity, presumably due to a large long term effective population size. Estimates of F ST and Linkage Disequilibrium (LD) across SNPs reveal a very low genetic differentiation throughout the genome and the absence of segregating LD blocks among populations, suggesting an overall lack of local adaptation. Our study provides the first investigation of the genetic structure and diversity in An. moucheti at the genomic scale. We conclude that, despite a weak genetic structure, this species has the potential to challenge current vector control measures and other rapid anthropogenic and environmental changes thanks to its great genetic diversity. [ABSTRACT FROM AUTHOR]

Aedes aegypti is the principal mosquito vector for many arboviruses that increasingly infect millions of people every year. With an escalating burden of infections and the relative failure of traditional control methods, the development of innovative control measures has become of paramount importance. The use of gene drives has sparked significant enthusiasm for genetic control of mosquitoes; however, no such system has been developed in Ae. aegypti. To fill this void, here we develop several CRISPR-based split gene drives for use in this vector. With cleavage rates up to 100% and transmission rates as high as 94%, mathematical models predict that these systems could spread anti-pathogen effector genes into wild populations in a safe, confinable and reversible manner appropriate for field trials and effective for controlling disease. These findings could expedite the development of effector-linked gene drives that could safely control wild populations of Ae. aegypti to combat local pathogen transmission. [ABSTRACT FROM AUTHOR]