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Evaluation of Inactivation Methods for Rift Valley Fever Virus in Mouse Microglia
Margarita V Rangel1, Feliza A Bourguet1, Carolyn I Hall2
1Biosciences and Biotechnology Division, Lawrence Livermore National Laboratory, Livermore, CA 94550, USA.
Abstract:
Rift Valley fever phlebovirus (RVFV) is a highly pathogenic mosquito-borne virus with bioweapon potential due to its ability to be spread by aerosol transmission. Neurological symptoms are among the worst outcomes of infection, and understanding of pathogenesis mechanisms within the brain is limited. RVFV is classified as an overlap select agent by the CDC and USDA; therefore, experiments involving fully virulent strains of virus are tightly regulated. Here, we present two methods for inactivation of live virus within samples derived from mouse microglia cells using commercially available kits for the preparation of cells for flow cytometry and RNA extraction. Using the flow cytometry protocol, we demonstrate key differences in the response of primary murine microglia to infection with fully virulent versus attenuated RVFV.
Insights
Rift Valley fever virus (RVFV) infection in the brain causes severe neurological symptoms. New methods inactivate live RVFV in mouse microglia cells, enabling safer study of the virus and its effects.
Area of Science:
- Virology
- Neuroscience
- Immunology
Background:
- Rift Valley fever phlebovirus (RVFV) is a highly pathogenic, mosquito-borne virus with potential for aerosol transmission and bioweapon use.
- Neurological manifestations are severe outcomes of RVFV infection, yet brain pathogenesis mechanisms remain poorly understood.
- RVFV is a CDC and USDA overlap select agent, necessitating strict regulations for experiments with virulent strains.
Purpose of the Study:
- To develop and present methods for inactivating live RVFV in biological samples from mouse microglia cells.
- To enable safer research into RVFV pathogenesis within the central nervous system.
- To compare the responses of primary murine microglia to both fully virulent and attenuated RVFV strains.
Main Methods:
- Two distinct inactivation protocols were employed using commercially available kits.
- Method 1: Preparation of mouse microglia cells for flow cytometry.
- Method 2: Extraction of RNA from infected mouse microglia cells.
Main Results:
- Successful inactivation of live RVFV in samples derived from mouse microglia cells was achieved.
- The flow cytometry protocol allowed for the observation of differential responses in primary murine microglia.
- Key differences were identified in microglia responses to fully virulent RVFV compared to an attenuated strain.
Conclusions:
- The presented inactivation methods facilitate safer investigation of RVFV in cellular models.
- These methods allow for the study of RVFV-induced changes in microglia, crucial for understanding neuroinflammation.
- Comparative analysis revealed distinct cellular responses to different RVFV virulence levels, advancing neuro-pathogenesis research.
