Degree Type

Thesis

Date of Award

2017

Degree Name

Master of Science

Department

Veterinary Microbiology and Preventive Medicine

Major

Veterinary Microbiology

First Advisor

James A. Roth

Second Advisor

Eric M. Vaughn

Abstract

A recombinant subunit vaccine against Seneca Valley virus (SVV) would be valuable to the swine industry. Recent SVV outbreaks have raised concerns with national biosecurity. This is due to the potential of foreign vesicular animal diseases entering the United States undetected because of confounding clinical signs of SVV. Research with the baculovirus expression vector system (BEVS) has produced virus-like particles (VLP) that elicited neutralizing antibodies and protected against challenge for many Picornaviridae viruses. Building on this previous research, attempts of SVV VLP assembly for a vaccine candidate were performed using the BEVS.

All baculovirus constructs were designed to encode the SVV P1 region, 2A protease, portions of the 2B and 3B genes and 3C protease. Recombinant SVV proteins VP1, VP2 and VP3 were expressed in several baculovirus construct iterations. Peptide specific antibodies detected each protein by means of Western blot analysis. Sucrose gradient fractionation and electron microscopy (EM) were performed to verify VLP formation. However, VLP production was not confirmed by either method.

Several factors influence the assembly of SVV VLPs. One outcome providing insight of the complete cleavage of the recombinant capsid proteins is the identification of a 55kDa protein band. This band was detected in the α-SVV VP1 and α-SVV VP3 Western blots. Also, the 3C protease and its impact in the BEVS need further investigation to determine its role in complete cleavage of the recombinant capsid proteins. Lastly, there were indications the recombinant capsid proteins were aggregating instead of folding properly into VLPs. The gaps in information noted here support the need for further research into capsid formation in Picornaviridae viruses to improve VLP assembly using the BEVS.

Copyright Owner

Jennifer Lynn English

Language

en

File Format

application/pdf

File Size

75 pages

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