Degree Type

Dissertation

Date of Award

2020

Degree Name

Master of Science

Department

Veterinary Microbiology and Preventive Medicine

Major

Immunobiology

First Advisor

Bradley Blitvich

Abstract

Baculovirus expression vector systems are used for the production of an increasing number of licensed commercial subunit vaccines. Though baculoviruses are incapable of replicating outside a narrow host range of lepidopteran insects, a replication-defective baculovirus system or platform provides an additional level of safety without the use of chemical inactivation methods such as binary ethylenimine. Circumventing the need for chemical inactivation protects sensitive proteins from degradation, and use of a live, yet replication-defective baculovirus has the potential to enhance antiviral immune response. These factors together make a replication-defective baculovirus platform highly desirable for subunit vaccine production. Choosing the essential baculovirus envelope protein gp64 as our target, we used recombination to delete gp64 from the baculovirus rendering it replication-defective. We used insect cell based plasmid expression vectors to generate insect cell lines stably expressing gp64 to allow passage of the replication-defective virus. Porcine epidemic diarrhea virus (PEDV) remains an economically important disease in swine, and a highly effective vaccine remains elusive. To test the capacity of the replication-defective baculovirus system to express an antigenic target of interest, we used recombination to replace the gp64 in the baculovirus genome with a chimeric PEDV spike protein. We then infected Sf9 cells with the PEDV spike expressing replication-defective baculovirus. Analysis of insect cell supernatants indicated that yield of PEDV spike was low in non-complementing cells, but shows an association between the baculovirus capsid VP39 and PEDV spike suggesting that the pseudotyped spike chimera restores budding of replication-defective baculovirus particles in the absence of gp64 suggesting that the system also has potential as a vector for gene delivery.

DOI

https://doi.org/10.31274/etd-20210114-1

Copyright Owner

Scott Ryan Ackerman

Language

en

File Format

application/pdf

File Size

57 pages

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