Location: Foreign Arthropod Borne Animal Disease Research
2024 Annual Report
Objectives
OBJECTIVE 1: Identify factors associated with Flavivirus infections, pathogenesis, and maintenance in vectors and animal hosts to inform prevention and mitigation strategies.
• Identify factors associated with JEV maintenance in relevant insect vectors.
• Characterize susceptibility, pathogenesis, and clinical disease of JEV in swine.
• Characterize vector-virus-host interactions associated with JEV transmission.
Sub-objective 1.A. Evaluate the ability of an emerging JEV genotype to infect and replicate in North American domestic swine and mosquito vectors.
Sub-objective 1.B. Investigate potential roles of North American feral swine and biting midges in JEV transmission.
OBJECTIVE 2: Identify and develop JEV control measures in swine.
• Develop detection measures fit for JEV surveillance in swine.
• Develop JEV vaccines for swine that will prevent virus amplification.
• Develop control measures to protect swine from JEV-infected Culex mosquitoes.
Sub-objective 2.A. Qualification of point-of-need diagnostics for Japanese encephalitis virus infections.
Sub-Objective 2.B. Develop and evaluate novel vaccine platforms to prevent JEV transmission from swine.
Sub-Objective 2.C. Develop control measures to protect swine from JEV-infected Culex mosquitoes.
Approach
Japanese encephalitis virus (JEV) is a zoonotic arthropod-borne pathogen native to Asia and the Pacific Rim, where it is a significant cause of reproductive and neonatal loss in swine and severe encephalitis and death in humans. JEV is transmitted to vertebrate hosts by infected mosquito vectors and has demonstrated an ability to emerge in new geographic regions that contain competent vectors and susceptible hosts. JEV does not currently circulate in the United States (U.S.); however, the risk of its introduction has been assessed as high (Oliveira et al. 2020). Significant research gaps exist regarding U.S. vulnerability following an introduction of JEV, including the range of native vectors and hosts capable of sustaining transmission and whether U.S. mosquitoes and livestock are vulnerable to emerging genotypes of JEV. This project will address these, and other gaps, by evaluating the ability of an emerging JEV genotype to infect and replicate in domestic swine and mosquitoes and by investigating the potential roles of previously uncharacterized wildlife hosts and insect vectors in JEV transmission. These studies will use in vitro and in vivo infection models to investigate the effects of wild-type JE viruses on insect vectors and mammalian hosts (Objective 1). Next generation sequencing and genomic analyses will be used to study vector-virus-host interactions to determine effects that hosts and vectors have on virus populations. Additionally, measures to protect swine from JEV will be developed including molecular diagnostic assays, novel vaccines, and a spatial insect repellent device (Objective 2). The knowledge gained will be used to inform risk assessments and predictive models and help identify target points to guide diagnostic development, surveillance programs, and control strategies. Together, these measures will help strengthen the U.S. disease prevention and response framework for rapidly stopping foreign animal disease incursions to protect the health and profitability of U.S. livestock.
Progress Report
Vector-borne disease transmission is dependent on competent vertebrate hosts and mosquito vectors to maintain and transmit the virus; disrupting the links will stop viral pathogen transmission. Objective 1a focuses on factors associated with Flavivirus infections, pathogenesis, and maintenance. Japanese encephalitis virus (JEV) has five genotypes and the recent emergence of genotype 4 in Australia and genotype 5 in Korea has resulted in questions about the ability of these emerging JEV genotypes to infect and replicate in North American domestic swine and mosquito vectors. To evaluate the susceptibility of North American mosquitoes for JEV competences beyond the established common genotypes 1 and 3 circulating in Asia, in vitro studies with Culex mosquito cell lines was conducted and the replication kinetics on three JEV genotypes recorded. Various virus concentrations of 0.01, 0.1, 1, and 5, or mock infected with media were evaluated and at 24 hours post-infection, cultures were observed for cell death and culture supernatants were collected. Viral titers for each concentration were quantified using a standard plaque assay.
Objective 1b focused on the potential hosts in North America and biting midges and their roles in transmission. Host cell lines were initiated to look for susceptibility to the emerging JEV genotypes and studies with North American feral swine cells and javelinas were initiated. Evaluation of the biting midge cell lines was also started.
Objective 2 aims to develop control measures and surveillance to detect JEV on farms. Rural areas are particularly susceptible for pathogen and invasive species introductions, but they have little to no surveillance. Therefore, a sustainable method to monitor these areas using crowd sourcing was developed as part of the ARSX and ARS Insects as Food and Feed Grand Challenge projects. Intensive agricultural settings such as rice fields or animal operations tend to have large numbers of pestiferous insects which may also be disease vector insects. A novel biomass harvesting trap was invented to trap in mass the pest insects and/or disease vectors which benefits the farmers by reducing the insect burden on crops or animals without the use of harmful pesticides. An additional benefit is these harvested insects can then be disinfected and fed as part of the daily ration to the animals as a protein supplement that is high in essential vitamins and essential amino acids. Researchers in Manhattan, Kansas, can take a small sample of the harvested insects and do metagenomic sequencing on them to look for invasive species DNA or pathogen RNA. The USDA-Biomass Harvest Trap (USDA-BHT) was designed, built, and evaluated for insect collection and use as a surveillance device. Furthermore, studies were conducted to evaluate the quality and safety of the harvested insects and use of those insects in a pilot study with poultry.
Accomplishments
1. Sustainable crowdsourced pathogen and insect surveillance for rural communities. ARS researchers in Manhattan, Kansas, developed the USDA-Biomass Harvest Trap (USDA-BHT) to help rural farmers supplement traditional animal protein sources with harvested pest and disease vector insects from intensive agricultural settings where they are naturally abundant. Simultaneously the harvested insects are used by ARS to conduct insect and pathogen surveillance in rural areas via these crowdsourced insect collections. This research empowers farmers to sustainably remove pestiferous and harmful insects that damage livestock or row crops and turn them into a protein supplement that can be added to the daily rations. As part of the ARSX and Insects as Food and Feed Grand Challenge (MINIstock) projects, the insect harvest team worked with other ARS researchers throughout the country to describe the concept and benefits of insect harvesting, develop and build the USDA-BHT, describe the trap capabilities and uses, evaluate the trap as a surveillance device, define the benefits to the farmers, quantify the nutritional benefits and hazards of harvested insects, and evaluate in a pilot study the benefits of harvested insects as poultry feed.
2. Development of nanoparticle pesticides. Pest and disease vector insects develop insecticide resistance rapidly requiring the continual discovery and evaluation of new pesticides. Novel nano- and microparticle synthesis offers a means to construct environmentally safe yet targeted insecticidal particles for immature insects. ARS researchers in Manhattan, Kansas, identified target sites in disease vector insects and then synthesized biodegradable food particles with silver particle coatings to create a food that is foraged by the aquatic larval stages. The edible particles take advantage of the immature insects’ natural filter feeding behaviors to gather food from the environment which concentrates the particles in the insect gut. Once ingested, the particles are not easily removed by grooming or detoxified with enzymes, rather the particles degrade, and the silver quickly punches holes in the larval gut resulting in high mortality. The combination of edible particles, concentrated in the gut, and an inability to remove or degrade them results in target insect mortality at incredibly low concentrations of active ingredient (parts per billion). New particles are continually being developed and a patent for the composition of the particles is pending.
Review Publications
Tsafrakidou, P., Papoti, V., Giannakakis, E., Christaki, A., Miaoulis, M., Oppert, B.S., Cohnstaedt, L.W., Arsi, K., Donoghue, A.M., Vergos, E., Zinoviadou, K., Chaskopoulou, A. 2024. Mosquitoes harvested from rice-fields as alternative protein ingredient in broiler feed: Insights from the first pilot study. Journal of Economic Entomology. 1-12. https://doi.org/10.1093/jee/toae096.
Lado, P., Rogers, D., Cernicchiaro, N., Swistek, S., Van Nest, K., Shults, P.T., Ewing, R.D., Okeson, D., Brabec, D.L., Cohnstaedt, L.W. 2024. Assessment of the USDA biomass harvest trap device as an insect harvest and mosquito surveillance tool. Journal of Economic Entomology. https://doi.org/10.1093/jee/toae095.
Norton, A.E., Ewing, R.D., Tilley, M., Whitworth, J., Cohnstaedt, L.W. 2023. Fatal food: Silver-coated grain particles display larvicidal activity in Culex quinquefasciatus. ACS Agricultural Science and Technology. 8(37):33437-33443. https://doi.org/10.1021/acsomega.3c03210.
Robinson, K., Duffield, K.R., Ramirez, J.L., Cohnstaedt, L.W., Ashworth, A.J., Jesudhasan, P., Arsi, K., Morales Ramos, J.A., Rojas, M.G., Crippen, T.L., Shanmugasundaram, R., Vaughan, M.M., Webster, C.D., Sealey, W.M., Purswell, J.L., Oppert, B.S., Neven, L.G., Cook, K.L., Donoghue, A.M. 2024. MINIstock: Model for INsect Inclusion in sustainable agriculture: USDA-ARS's research approach to advancing insect meal development and inclusion in animal diets. Journal of Economic Entomology. 117(4):1199-1209. https://doi.org/10.1093/jee/toae130.
Osborne, C.J., Cohnstaedt, L.W., Su, T., Silver, K.S. 2024. Variable gut pH as a potential mechanism of tolerance to Bacillus thuringiensis subsp. israelensis toxins in the biting midge Culicoides sonorensis. Pest Management Science. https://doi.org/10.1002/ps.8104.
Hudson, A.R., McGregor, B.L., Shults, P.T., England, M., Silbernegal, C., Mayo, C., Carpenter, M., Sherman, T., Cohnstaedt, L.W. 2023. Orbivirus epidemiology in a changing climate. Journal of Medical Entomology. 60(6):1221-1229. https://doi.org/10.1093/jme/tjad098.