2012 Annual Report 1a.Objectives (from AD-416): Objective 1: Assess the risk of endemic arthropod vectors to transmit introduced exotic arboviruses in North America. • Sub-objective 1.A. Determine the vector competence of the primary U.S. bluetongue virus (BTV) vector, Culicoides sonorensis, for EU-BTV-8. • Sub-objective 1.B. Create models to assess potential population densities for biting insects that might be involved if Rift Valley fever virus was introduced to North America using ecologic and climatic factors. Objective 2: Identify targets and evaluate tools for vector control and interruption of transmission cycles to protect livestock and humans from vector-borne pathogens. • Sub-objective 2.A. Identify molecular components in insects that can be targeted to interrupt orbivirus transmission cycles. • Sub-objective 2.B. Evaluate insecticide resistance of Culicoides sonorensis to common pesticides used in livestock and agricultural operations. • Sub-objective 2.C. Provide livestock entomologists improved identification tools for North American Culex tarsalis and Aedes vexans. 1b.Approach (from AD-416): Livestock are often heavily exposed to biting arthropods, causing a number of animal health issues and making them vulnerable to infection with a wide range of insect-borne pathogens. This research program will focus on:. 1)improving risk assessments of the potential for introduction of foreign disease agents into the U.S.,. 2)interrupting transmission cycles at the vector level,. 3)identifying viable pesticides for control of vectors, and. 4)improving vector identification and understanding of population dynamics to enable more efficient vector control. Determining the vector competence of Culicoides sonorensis for an exotic bluetongue virus (BTV-8) will give an indication of the potential risk for the spread of exotic BTV should it be introduced into North America. Targets for controlling C. sonorensis infection with orbiviruses will come through identification of insect cell receptor(s) for BTV and verification of specific genes associated with orbivirus infection in the insect using RNA interference. Determining the susceptibility of C. sonorensis to common insecticides will identify effective treatments for control of this important livestock pest. Development of predictive models to determine risk of arbovirus transmission, such as Rift Valley fever virus, based on predicted mosquito population densities and distributions will give weeks or months advance notice, allowing preventive measures to reduce or prevent animal and human disease. Understanding the population structures of Aedes vexans and Culex tarsalis through molecular data will provide useful information for field entomologists and agencies developing control strategies and for use in developing models to predict risk of arbovirus transmission. 3.Progress Report: EU-BTV-8 from The Netherlands was inoculated into two sheep, blood was harvested at 8 days post infection and approximately 500 mls of washed blood cell inoculum was made. This will serve as the virus source for upcoming vector competence studies. Permit to ship the virus inoculum from Colorado State University to the Kansas State University Biosecurity Research Institute (BRI) was applied for and received. Virus was shipped and is now in storage at the BRI. An insect-secure arthropod containment laboratory at BRI has been completed and is pending APHIS and CDC approval for use with infected arthropods. RNA has been collected from Culicoides sonorensis unmated AK colony females for baseline transcriptome generation. This includes two biological replicates, each, of (1) unfed teneral, (2) blood fed at 36 h post-meal and (3) 10% sucrose fed at 36 h post-meal. RNA was quantified and processed and sent to Clemson University Genomics Institute for RNAseq transcriptomics. Arthropod Containment Lab at ABADRU has been recently approved (May 2012) and work could not begin before construction of the lab or approval. IBC proposal has been granted for insect infections and micromanipulations in BSL-2. In collaboration with the USDA-ARS-CMAVE, acquired satellite data from NASA then formatted, and analyzed monthly normalized difference vegetation index (NDVI) environmental data for North America, including the calculation of anomaly NDVI values from a 25-year mean NDVI baseline. Long term anomaly NDVI data, including data products derived with the accumulated anomaly approach developed by NASA, are being used to analyze historical eco-climatic variability patterns and linkages with population dynamics of mosquitoes of medical and veterinary importance in the U.S. In collaboration with faculty at the University of Wyoming, a degree day model was developed and placed online. The model is real time and uses temperature to assess risk of Rift Valley fever virus transmission and establishment in the continental USA. In collaboration with faculty at Kansas State University, a hierarchical network modeling approach was used to predict the outcomes of introductions of Rift Valley fever virus from infected mosquito eggs, adult mosquitoes, and cattle. The model combined GIS, environmental, and livestock and human census data to predict the number of infections based on stochastic weather fluctuations and population densities of cattle, humans and mosquitoes in Texas. To control disease vectoring midge populations, an integrated pest management solution was developed that would target the breeding area, aerial application, and on host feeding using a combination of new and established methods and insecticidal products. Several farmers have committed their property for the testing of new methods and products on their animals. Collected Culex tarsalis and Aedes vexans from around the United States and Canada for population genetics experiment. In collaboration with the Floragenex, sequencing of the first 96 populations of mosquitoes for full genome single nucleotide polymorphism discovery has started. 4.Accomplishments 1. The North American Mosquito Project Collection Network. ARS researchers in Manhattan, Kansas led a network of mosquito collectors from the general public, mosquito control districts, and public health agencies worked together to collect two species of mosquitoes throughout the continental United States in the summer of 2011. More than 86 entities (individuals or agencies) collected 454 unique populations making this one of the largest coordinated collections of disease vector mosquitoes in North America. These mosquitoes will be used for genetic studies to determine the historic spread of the species and to determine the migration rates between populations of mosquitoes which will help researchers understand the spread of insecticide resistance genes and vector-borne diseases. 2. Biting midge (Culicoides sonorensis) gene expression analysis. The genome has not yet been sequenced for the biting midge (Culicoides sonorensis), an important disease vector. Likewise, we do not have a good understanding about the products of these genes (‘gene expression’) and their role in the biology of Culicoides midges. ARS scientists in Manhattan, Kansas, in collaboration with Clemson University Genomics Institute are generating the first catalogs of gene products involved in the biological processes of female midges. Analysis of these gene catalogs reveals gene expression profiles associated with basic processes such as feeding and digestion. The evolution of these genes, the frequency at which their products are used, and alterations of gene products are being determined. Comparative studies between the expression profiles across different life conditions (blood feeding, fasted, etc.) should reveal genes uniquely associated with each process, as well as down- or up-regulation. These data will be used in future approaches to reveal genes associated with early and late antiviral responses when compared to profiles for virus-infected midges. Review Publications Cohnstaedt, L.W., Rochon, K., Duehl, A.J., Anderson, J., Barrera, R., Su, N., Gerry, A., Obenauer, P., Campbell, J.F., Lysyk, T., Allan, S.A. 2012. Arthropod surveillance programs: Basic components, strategies, and analysis. Annals of the Entomological Society of America. 105:135-149. Tchouassi, D.P., Sang, R., Sole, C., Bastos, A.D., Cohnstaedt, L.W., Baldwyn, T. 2012. Trapping of Rift Valley Fever (RVF) vectors using Light Emitting Diode (LED) CDC traps in two arboviral disease hot spots in Kenya. Parasites & Vectors. 5:94. Gupta, A.K., Nayduch, D., Verma, P., Shah, B., Ghate, H., Patole, M.S., Shouche, Y.S. 2011. Phylogenetic characterization of bacteria in the gut of house flies (Musca domestica L.). Federation of European Microbiological Societies Microbiology Letters. 78:581-593.