For over thirty years, Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) has been a major contributor to morbidity and mortality in the commercial swine industry across the globe. This highly mutagenic RNA virus causes significant economic losses wherever it is prevalent, leading to $664 million in annual losses in the United States. Unfortunately, the current prevention and diagnostic techniques available have proven to be insufficient in controlling the spread of this disease. We describe an alternative diagnostic method exploiting the rapid turnaround time and long-read capacity of Oxford Nanopore Technology's MinION next-generation sequencer. We have developed a novel primer set designed to span Open Reading Frames 3 through 7 of the PRRSV genome, which has allowed for multiplexing of samples, thus reducing individual cost of testing, while yielding significantly more information than previously available. This novel primer pair and sequencing technique have distinguished mixed infections within individual animals and may be used to determine vaccination status. This new approach will help producers and veterinarians make better-informed decisions about co-mingling of animals and vaccination strategies, thus reducing the emergence of new, pathogenic strains of PRRSV. / Master of Science / Porcine reproductive and respiratory syndrome virus (PRRSV) is a common, economically important pathogen in commercial swine production. The virus was first identified in the late 1980's during outbreaks in the United States and Europe. In female pigs, the disease is characterized by abortion storms, and the delivery of mummified fetuses or very weak, ill piglets. Neonates often display signs of pneumonia, respiratory distress, and many die from hypoxia. Surviving piglets are highly susceptible to other diseases and are poor growers compared to other, unaffected piglets. Boars may show signs of respiratory disease and can also have decreased libido and reproductive success for months at a time. The virus is prone to mutating once a pig is infected, preventing herds from mounting sufficient immunity to protect against new, mutant strains. Identifying infected pigs early and accurately is crucial to managing PRRSV outbreaks. Currently available diagnostic tests for PRRSV have many limitations, thus we have developed a new diagnostic test using next-generation sequencing technology. Oxford Nanopore Technology provides a commercially available nanopore sequencer, the MinION, that can read long DNA strands in real-time. With this technology we have expanded the area of the PRRSV genome that can be sequenced, which allows us to better identify and distinguish strains of PRRSV in infected, and vaccinated pigs. This new testing method will allow veterinarians and practitioners across the country to better identify and predict outbreaks in their herds, helping them develop better management strategies against PRRSV.
| Identifer | oai:union.ndltd.org:VTETD/oai:vtechworks.lib.vt.edu:10919/117321 |
| Date | 08 January 2024 |
| Creators | Buman Ruiz Diaz, Maria Paz |
| Contributors | Biomedical and Veterinary Sciences, Lahmers, Kevin K., Slade, Daniel Joseph, LeRoith, Tanya |
| Publisher | Virginia Tech |
| Source Sets | Virginia Tech Theses and Dissertation |
| Language | English |
| Detected Language | English |
| Type | Thesis |
| Format | ETD, application/pdf, application/pdf |
| Rights | In Copyright, http://rightsstatements.org/vocab/InC/1.0/ |
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