BRC-BIO: Discovery and characterization of novel viral pathogens of Caenorhabditis elegans

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Caenorhabditis elegans and other related free-living nematodes are a popular and powerful model system used to study how pathogens infect host cells and how the host immune system responds to infection on a cellular level. C. elegans offers many advantages for these types of studies: the animals are small and easy to grow in large numbers, have a fast life cycle, are easy to genetically manipulate, and are transparent, thus allowing observation of infections over time in live animals. However, only one virus capable of naturally infecting C. elegans has been identified to date, which severely limits C. elegans as a model system for studying viral infections specifically. This project aims to address this gap by identifying new viruses in wild-caught nematodes that can be passed to C. elegans, and studying the cell biology of the newly identified viral infections. Wild nematode collection will be conducted through the Nematode Hunters program, an outreach program that brings authentic biology research into elementary school classrooms. Nematode Hunters participants will collect wild free-living nematodes from their local environment and submit them for infection screening, which will be conducted by undergraduate students enrolled in a course-based research experience. Newly identified viruses will be studied to determine what tissues they infect and the course of the infection. Ultimately, the new viruses will be made available to the research community, thereby enabling the power of the C. elegans model to be applied to a wider range of questions about how viruses interact with host cells. This project aims to identify novel viruses in wild-caught free-living nematodes that are transmissible to C. elegans and to characterize their genomic structure, infection dynamics, and tissue tropism. A geographically diverse sampling will be collected via the Nematode Hunters outreach program, and screening for viral transmissibility to C. elegans will be conducted by co-culturing with C. elegans expressing fluorescent infection reporters. Potential viral infections will be identified by homogenizing co-cultures showing evidence of infection transmission, filtering the homogenate to size-exclude pathogens larger than a virus, and testing the ability of these filtrates to transmit the infection. This high-throughput method will allow rapid screening of many wild nematode isolates to identify novel viral infections. For potential viral infections, RNA and DNA will be extracted from infectious filtrates and infected worms, and sequenced to identify reads corresponding to viral RNA/DNA. Viral sequences identified will be used to design primers and fluorescence in situ hybridization probes, which will be used to determine the tissue tropism and infection dynamics of newly discovered viral infections. By establishing a pipeline for viral discovery and characterization, the project will uncover new viruses capable of naturally infecting C. elegans and make them available to the research community for comparative studies.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.