Development and application of recombinant proteins for improved BPI-3 and BHV-1 sero-diagnosis

Abstract

Bovine Respiratory Disease (BRD) is an economically important multifactorial disease in which an increasing prevalence of viral agents has been identified. This study investigated the application of recombinant viral surface proteins, associated with Bovine parainfluenza type-3 (BPI-3) and Bovine herpes virus type 1 (BHV-1), as capture antigens for the purposes of improving the serodiagnosis and screening of these infections. Newly designed and synthesised recombinant BHV-1 glycoprotein B (gB) and BPI-3 Haemagglutinin Neuraminidase (HN) proteins were incorporated within different immunoassay formats and performance compared to commercial viral lysate-based serological tests. Recombinant BPI-3 HN showed the most potential as a capture antigen, demonstrating versatility in the development of serological, fluorescent and biosensor BPI-3 specific immunoassays. All formats demonstrated high viral specificity and minimal non-specific binding interferences, with the fluorescent BPI-3 immunoassay demonstrating increased assay sensitivity relative to available BPI-3 HN serological assays. An IgM-based BPI-3 HN immunoassay was also developed revealing IgM antibody patterns suggesting that IgM may play a more significant role in anamnestic responses during early vaccination / vaccination in the face of maternal antibodies. Orientated immobilisation of recombinant BHV-1 gB onto SPR biosensor platforms was shown to overcome issues with protein purification, enabling measurement of significant differences between BHV-1 positive and negative animals, as well as other viral-specific sera. Overall, recombinant BPI-3 HN protein showed the best utility as a capture antigen for improved viral serodiagnosis, as well as potential for further use in the development of multiplex/point of care tests. Using oriented immobilisation approaches, recombinant BHV-1 gB protein can be used as a capture antigen, and with further purification has potential for use for immunoassay development on more sensitive assay platforms.

Thesis embargoed until 31st December 2026

Date of Award	Dec 2025
Original language	English
Awarding Institution	Queen's University Belfast
Supervisor	Mark Mooney (Supervisor), Gordon Allan (Supervisor) & Ruramayi Nzuma (Supervisor)