AbstractFeline coronavirus (FCoV) leads to the fatal disease known as feline infectious peritonitis in a small proportion of infected cats. Research into FCoV has so far been hindered by our inability to culture the most common serotype, Type 1, in vitro.
This project aimed to establish a reverse genetic system for Type 1 FCoV. Such a system would enable a deeper understanding of the role of viral mutations in the pathogenesis of disease and could serve as a platform for rational vaccine design. To this end, cDNA infectious clone and replicon constructs based on Type 1 FCoV were developed and transcribed. During this project, infectious virus or replicon-expressing cells were not recovered following transfection of the constructs into mammalian cells, but a foundation was laid for this to be achieved in future.
Recovery of recombinant Type 1 FCoV would necessitate a cell line capable of supporting this serotype’s growth in vitro, so an aim of this project was to identify a cell entry receptor (CER) for Type 1 FCoV in order to develop a cell line permissive to infection with the virus. To this end, ‘bait proteins’ bearing the spike proteins of Type 1 and 2 FCoV were produced, and Type 2 bait protein was able to recognise its CER. Feline intestinal organoid cultures were established, and an interaction identified between Type 1 bait protein and heat shock 70 kDa protein 1a (HSPA1A) suggested that HSPA1A is a receptor for Type 1 FCoV. Finally, a feline IFN-gamma ELISpot assay was used to measure the cellular immune response to peptides representing a Type 1 FCoV epitope with and without a substitution. No significant difference in the ability of the peptide variants to stimulate a response was identified, but the assay could be used in the future to guide rational vaccine design.
|Date of Award||23 Jan 2020|
|Supervisor||Andrew D Davidson (Supervisor) & Severine Tasker (Supervisor)|