Prevention and Pathogenesis of Avian Infectious Bronchitis: The Role of Infectious Bronchitis Virus Spike Proteins in Cell Attachment and Protective Immunity
Type of DegreePhD Dissertation
General Veterinary Medicine
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Infectious bronchitis virus (IBV) evolves by genetic diversity and selection resulting in continuous emergence of new virus phenotypes. Spike (S) protein is responsible for virus entry. The S1 subunit of the S glycoprotein mediates viral attachment to host cells and the S2 subunit is responsible for membrane fusion. Using protein histochemistry, we investigated the tissue binding ability of the S-ectodomain of the ArkDPI-derived vaccine subpopulation previously designed C2 compared to its S1 protein alone. We demonstrated that extension of S1 with the S2 ectodomain, together comprising the S-ectodomain, greatly increases binding to chicken tissues, suggesting that vaccination with the S-ectodomain might improve protection over vaccination with S1 protein alone. Our protection study revealed that chickens immunized with recombinant S-ectodomain protein showed statistically significantly reduced viral loads 5 days post-challenge in both tears and tracheas, and exhibited improved protection of tracheal integrity compared to chickens immunized with recombinant S1 protein. These results indicate that the S2 domain has an important role in inducing protective immunity. IBV 4/91 (serotype 793/B) vaccine has been reported to protect against divergent IBV strains in a prime-boost approach with an IBV Mass vaccine. Thus, we evaluated the protective capabilities of recombinant Newcastle disease virus LaSota (rLS/IBV.S2-4/91) expressing the S2 gene of IBV 4/91, to determine whether this cross protection could be explained by cross-protective properties of 4/91 S2. We found non-significant differences in protection 5 days post-challenge between chickens primed with Mass vaccine and boosted with rLS/IBV.S2-4/91 and chickens vaccinated with Mass only, indicating that the S2 by itself might be insufficient to induce broad cross-protective immunity. The role of S protein domains in attachment and vaccine subpopulation selection were also investigated using S proteins derived from ArkDPI-derived vaccine subpopulations with different selection pattern: C2 (strongly selected), C3 (weakly selected), and major vaccine population (negatively selected). We demonstrated that S1 proteins of vaccine virus subpopulations that are strongly positively selected in chickens bind more efficiently to chicken tissues than that of the negatively selected vaccine population. The S2 domain was required in addition to the S1 domain for binding to lung and kidney tissues. Finally, we determined the receptor binding domain (RBD) of recombinant S1 protein. The N-terminal amino acids 19-258 (NTD258) of S1 is required and sufficient for binding to all chicken tissues tested except lung tissues. Only S-ectodomain bound to lung. The binding efficiency of NTD258 protein is better than full-length S1 protein to all tissues tested, suggesting that the remainder of the S1 protein affects the conformation of the NTD.