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Abstracts 1001-1250 305 and contains several ORs, OBPs and IRs. These genes are candidates for this gene encodes a key regulator of secretory function in the Ae. aegypti being involved in the adaptation of An. gambiae to its human host and salivary gland. These studies highlight the need for further analysis of were sequenced in six anopheline species to identify those that show mosquito developmental genetics and may foster comparative studies of evidence of positive selection. salivary gland development in additional vector insect species. 1001 1003 DENGUE 2 INFECTION ALTERS MICRORNA EXPRESSION IN MEDUSA: A NOVEL GENE DRIVE SYSTEM FOR CONFINED AEDES AEGYPTI SUPPRESSION OF MOSQUITO POPULATIONS Corey Campbell, Ann Hess, Gregory D. Ebel John M. Marshall1, Bruce A. Hay2 Colorado State University, Fort Collins, CO, United States 1Imperial College London, London, United Kingdom, 2California Institute of Emerging studies show that important avenues in the post-transcriptional Technology, Pasadena, CA, United States regulation of gene expression occur via small RNA regulatory pathways. Following successful field trials of sterile GM mosquitoes designed to Non-coding RNAs (ncRNAs) are key features of these pathways, and control dengue fever, interest is now growing in the use of gene drive critical molecules involved in their biogenesis and function are conserved systems, such as X-shredders, capable of inducing a population crash as in plants, insects and mammals. To identify products of anti-viral RNA they spread. These systems hold much promise for wide-scale disease interference in vector mosquitoes, deep sequencing small RNA (sRNA) control; however issues arise from their potential to spread across libraries were prepared from DENV2-fed Ae. aegypti females (RexD strain) international borders. We propose a novel gene drive system, Medusa, and matched un-infected controls at 2, 4, and 9 days post-infection (dpi). capable of inducing a local population crash without spreading into An earlier publication described DENV2-derived viral sRNAs (viRNAs) across neighboring populations. Medusa consists of four components - two at a three size classes: unusually small RNAs (usRNAs) (14-19nts), canonical locus on the X chromosome and two at a locus on the Y chromosome. A siRNAs (20-24nts), and piRNAs (25-30nts) (Hess et al, BMC Microbiology, maternally-expressed, X-linked toxin and a zygotically-expressed, Y-linked 2011). In the present analysis, these libraries were mined to determine antidote suppress the female population because only males can protect whether substantive changes occur in mosquito microRNA (miRNA) levels themselves against the effects of the toxin. A zygotically-expressed, during DENV2 infection. Reads were aligned to mIRBase release 17 hairpin Y-linked toxin and a zygotically-expressed, X-linked antidote ensure database (mirbase.org). MiRNAs with over 50 reads across treatment that the two constructs are always inherited together. We use simple groups and showing > 2 fold-changes were chosen for further study. Our population genetic models to explore the dynamics of the Medusa system. analysis reveals that significant changes to specific miRNA levels occur in An all-male release is preferred since males don’t bite and, if released DENV2-infected mosquitoes compared to un-infected controls. Moreover, over two generations, Medusa is expected to induce a population crash some miRNAs showed coordinated enrichment or depletion at both 2 within seven generations for modest release sizes. Re-invasion of wild and 4 dpi, substantiating the hypothesis that they are important to the mosquitoes can lead to the population eventually rebounding; however establishment of virus infection, whether by being exploited by the virus or this can be prevented by small, regular releases of Medusa males. The as part of an anti-viral mechanism. Coordinately co-regulated miRNAs or Medusa system could serve as a proof of principle for invasive population *miRNAs include miR155, mIR2755, miR281 and miR277, among others. suppression systems such as X-shredders. We describe molecular solutions miRNAs were classified by type: conserved (homologous to previously to chromosomal anomalies that could interfere with Medusa dynamics. reported miRNAs), *miRNA (complementary to miRNA), non-canonical or unclassified. Although the precise part played by each differentially 1004 expressed miRNA remains to be elucidated, orthologous miRNAs in other animals are important effectors of cellular differentiation, neurogenesis, VACCINATION WITH EXCRETORY/SECRETORY PRODUCTS transcriptional regulation, nutritional metabolism and the regulation of CONFERS PARTIAL PROTECTION IN A MURINE MODEL OF apoptosis. Our results show an intriguing new way in which major cellular FILARIASIS processes of mosquitoes respond to arbovirus infection. C. Paul Morris, Marina Torrero, Kristin Killoran, Edward Mitre 1002 Uniformed Services University of the Health Sciences, Bethesda, MD, United States GENETIC REGULATION OF VECTOR MOSQUITO SALIVARY Infection with filarial worms can cause severe and debilitating diseases GLAND DEVELOPMENT in both humans and animals. While many vaccine candidates have been studied in filariasis, our understanding of protective immune responses in Molly Duman Scheel1, Chilinh Nguyen2, Zeinab Annan1, Emily a permissive model to filariasis is incomplete. In this study, we evaluated Andrews1, Christy Le2, Longhua Sun2, Anthony Clemons2, David preparations of worm antigens for protection against challenge infection W. Severson2 in the BALB/c Litomosoides sigmodontis model. The fractions included 1 Indiana University School of Medicine South Bend at Notre Dame, South LS, soluble antigens of a homogenate of adult worms, and ES, excretory/ 2 Bend, IN, United States, University of Notre Dame, Notre Dame, IN, secretory products of adult female worms. 6-8 week old female BALB/c United States mice were given 3 intraperitoneal injections of 10 micrograms LS or ES Understanding mosquito salivary gland development is critical given the with CpG/Alum and subsequently challenged with 40 infectious larvae importance of this tissue in blood feeding and pathogen transmission. subcutaneously. 8 weeks after infection mice were euthanized and Our recent survey of the mosquito genomes indicated that mosquitoes parasite burdens were determined. Mice that were vaccinated with LS have orthologs of many genes that regulate embryonic salivary gland antigen showed no significant protection against challenge infection development in Drosophila melanogaster, a well-characterized insect compared to control mice. Mice vaccinated with ES product, however, genetic model organism. The expression patterns of a large subset harbored 60% fewer adult worms than control mice. While mice of these genes were assessed during development of Aedes aegypti, vaccinated with LS and ES produced similar levels of IgG antibodies to an emerging model for vector mosquito development. These studies both antigen preparations, analysis by western blot demonstrated that revealed that the early stages of Ae. aegypti salivary gland development ES vaccinated mice recognized different ES proteins than LS-vaccinated significantly differ from that of D. melanogaster. We are now using an mice. Currently, we are in the process of conducting mass spectroscopy RNAi knockdown strategy to investigate the roles of genes expressed in to identify a ~160kda protein that was strongly preferentially recognized the developing Ae. aegypti salivary gland. Functional characterization of by ES-vaccinated mice. No substantially large differences were observed cyclic-AMP response element binding protein A (crebA) indicates that between the two vaccinated groups with regards to lymphocyte www.astmh.org 306 proliferation or Th1 and Th2 cytokine production in response to ES or LS. a number of different IL-17 responses in the CIA model. Firstly, it acts However, IL-10 responses to parasite antigens were substantially decreased to inhibit priming and polarisation of IL-17 responses by targeting a in ES vaccinated mice. Analysis of worm counts at early timepoints suggest complex IL-17-producing network, involving signalling between dendritic that ES vaccination does not disrupt L3 migration through tissues. Work cells and/or CD4+ T cells. Secondly, ES-62 directly targets Th17 cells by is underway testing whether ES vaccination blocks the ability of filarial down-regulating expression of MyD88 to suppress responses mediated worms to induce immune regulation. by IL-1 and TLR ligands. Further, ES-62 interferes with migration of T cells and this is reflected by direct suppression of CD44 up-regulation 1005 and, as evidenced by in situ analysis, dramatically reduced levels of IL-17- producing cells, including lymphocytes, infiltrating the joint. Finally, there IMMUNO-PROPHYLACTIC EVALUATION OF RECOMBINANT is strong suppression of IL-17 production by cells resident in the joint, CUTICULAR COLLAGEN (COL-4) AND ABUNDANT LARVAL such as osteoclasts within the bone areas. Targeting of IL-17 responses by TRANSCRIPT (ALT) USING MULTIVALENT STRATEGY IN ES-62 presumably reflects the need of the parasite to dampen down host EXPERIMENTAL FILARIASIS inflammatory responses that could be dangerous to parasite or even host. However, the multi-site manipulation of the initiation and effector phases 1 1 1 Kaliraj Perumal , Arunkumar Chakkaravarthi , Prince R. Prabhu , of the IL-17 inflammatory network is notable and we believe further 2 Reddy Maryada Venkata Rami strengthens the argument
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