Porcine epidemic diarrhea Virus (PEDV) is a virus effecting swine. It is the cause of disease that manifests with symptoms ranging from depression, to severe dehydration and death. Young piglets are particularly susceptible to the virus, which can reach mortality rates of 100%. Presence of the virus on a swine farm can therefore cause severe economic losses. Treatments currently exist for PEDV, but are mostly generated from the virus itself. There has recently been renewed interest in a vaccine that is made from a different source, which might help eliminate some of the side effects of those that currently exist on the market. This project outlines three experiments performed in animals. During the first experiment, a structural protein from the Hepatitis B virus was genetically altered to include important structural portions of PEDV. This new protein is generated in E. coli and purified. After purification, the protein assembles into a virus-like particle (VLP). VLPs are structural proteins of existing viruses that are expressed and assembled to mimic the virus. By doing so, the immune system recognizes the protein as a potential threat, and launches a response in the form of antibodies. Manipulations of the VLPs as describe herein allow the new vaccine to generate antibodies toward other diseases such as PEDV. Although all five of the vaccines used in the first experiment were able to generate appropriate antibodies, only two of them were effective at preventing PEDV from entering susceptible cells (virus neutralization). A second experiment, with three newly designed vaccines was therefore performed. This experiment, like the first, was successful in producing antibodies to several of the included PEDV protein sections, but none were able to neutralize the virus. These results led to a third experiment, during which further design improvements were made to the basic vaccine structure in an attempt to increase the neutralization capabilities of the vaccines. The results from the third experiment indicated that several changes to the vaccine increased the immune response to the structural portions of PEDV, providing a better overall vaccine candidate. This also led to the conclusion that one specific sequence from PEDV has a better ability to neutralize the virus than the other sections. / PHD

Porcine reproductive and respiratory syndrome virus (PRRSV) and porcine epidemic diarrhea virus (PEDV) are two of the most prevalent swine pathogens that have impacted the global swine industry for decades. Both are RNA viruses with increasing heterogeneity over the years, making a vaccine solution ever so challenging. Modified live-attenuated vaccines (MLVs) have been the most common approach, but the long-term safety regarding their potential for pathogenic reversion still needs to be addressed. Subunit based vaccines have been the focus of numerous development studies around the world with renewed interest in their promising prospects in both safety and efficacy. Our lab has developed a unique approach to use hepatitis B virus core capsid protein (HBcAg) as a vaccine delivery vehicle for either PRRSV or PEDV viral epitope antigens. Recombinantly produced HBcAg forms an icosahedral capsid virus-like particle (VLP) that has 240 repeats in a single assembled particle. By inserting different epitope antigens from these porcine pathogens into the particle, we can achieve repetitive antigen presentation to the host's immune system by taking advantage of the polymeric nature of VLP. The first animal study evaluated the efficacy of 4 VLP based vaccine candidates against PRRSV in mice. These 4 vaccines incorporated 2 B-cell epitopes (61QAAIEVYEPGRS72 and 89ELGFVVPPGLSS100) and 2 T-cell epitopes (117LAALICFVIRLAKNC131 and 149KGRLYRWRSPVIIEK163) from PRRSV structural proteins GP3 and GP5 respectively. Candidate GP3-4 was able to stimulate a significant viral neutralizing response in mouse sera against two PRRSV strains, one being heterologous, demonstrating its potential of cross-protection against PRRSV. The second animal study took an optimized VLP vaccine candidate against PEDV from previous development studies in mice, and assessed its efficacy through a comprehensive pregnant gilt vaccination and neonatal piglet challenge model. The vaccine candidate incorporated B-cell epitope 748YSNIGVCK755 from the PEDV spike protein. It was able to elicit significant viral neutralization antibody titer in gilt milk at 3 days post-farrowing (DPF), and provided nursing piglets with clinical relief in terms of morbidity, viral shedding, small intestinal lesions, and 10 days post-challenge (DPC) survival rate. / Doctor of Philosophy / Porcine reproductive and respiratory syndrome virus (PRRSV) and porcine epidemic diarrhea virus (PEDV) are two pathogens that infect pigs, resulting in immense economic losses to the global pork production industry every year. Both viruses have large diversity with various strains due to mutations that have occurred over the years. This makes vaccine development that aims at combating the pathogens even more challenging. One common vaccine strategy has been immunizing animals with modified live viruses with decreased pathogenicity. Naturally, long term safety of this option has been a concern. A much safer vaccine approach that is purely protein based has attracted renewed interest around the world. Protein based vaccines lack genetic materials from the viruses and are not able to replicate inside the host. Our lab has developed a platform that uses protein-based particles (VLPs) originated from the hepatitis B virus (HBV), and incorporates short pieces of proteins from either PRRSV or PEDV to train host's immune system to recognize these pathogens, and hopefully to prevent future infection. For the first animal study, we tested 4 VLP vaccine candidates against PRRSV in mice and discovered that mouse serum from one candidate GP3-4 was able to prevent infection of 2 distinct PRRSV strains in petri dishes, paving the way for further development. For the second animal study, we took an optimized VLP vaccine candidate against PEDV from previous mouse studies, and evaluated its performance in pigs. We immunized pregnant mother pigs with the vaccine before they gave birth, then experimentally infected newborn piglets with the virus. Piglets from the vaccinated mothers showed improved clinical signs and faster recovery from the infection.