Abstract
Avian coronavirus infectious bronchitis virus (IBV) is the etiological agent of infectious bronchitis, an acute highly contagious economically relevant respiratory disease of poultry. Vaccination is used to control IBV infections, with live-attenuated vaccines generated via serial passage of a virulent field isolate through embryonated hens' eggs. A fine balance must be achieved between attenuation and the retention of immunogenicity. The exact molecular mechanism of attenuation is unknown, and vaccines produced in this manner present a risk of reversion to virulence as few consensus level changes are acquired. Our previous research resulted in the generation of a recombinant IBV (rIBV) known as M41-R, based on a pathogenic strain M41-CK. M41-R was attenuated in vivo by two amino acid changes, Nsp10-Pro85Leu and Nsp14-Val393Leu; however, the mechanism of attenuation was not determined. Pro85 and Val393 were found to be conserved among not only IBV strains but members of the wider coronavirus family. This study demonstrates that the same changes are associated with a temperature-sensitive (ts) replication phenotype at 41°C in vitro, suggesting that the two phenotypes may be linked. Vaccination of specific-pathogen-free chickens with M41-R induced 100% protection against clinical disease, tracheal ciliary damage, and challenge virus replication following homologous challenge with virulent M41-CK. Temperature sensitivity has been used to rationally attenuate other viral pathogens, including influenza, and the identification of amino acid changes that impart both a ts and an attenuated phenotype may therefore offer an avenue for future coronavirus vaccine development. IMPORTANCE Infectious bronchitis virus is a pathogen of economic and welfare concern for the global poultry industry. Live-attenuated vaccines against are generated by serial passage of a virulent isolate in embryonated eggs until attenuation is achieved. The exact mechanisms of attenuation are unknown, and vaccines produced have a risk of reversion to virulence. Reverse genetics provides a method to generate vaccines that are rationally attenuated and are more stable with respect to back selection due to their clonal origin. Genetic populations resulting from molecular clones are more homogeneous and lack the presence of parental pathogenic viruses, which generation by multiple passage does not. In this study, we identified two amino acids that impart a temperature-sensitive replication phenotype. Immunogenicity is retained and vaccination results in 100% protection against homologous challenge. Temperature sensitivity, used for the development of vaccines against other viruses, presents a method for the development of coronavirus vaccines.
Original language | English |
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Article number | e01100-22 |
Number of pages | 21 |
Journal | Journal of Virology |
Volume | 96 |
Issue number | 17 |
Early online date | 16 Aug 2022 |
DOIs | |
Publication status | Published - 14 Sept 2022 |
Bibliographical note
Funding Information:This study was funded by UKRI Biotechnology and Biological Sciences Research Council (BBSRC) grants BB/R019576/1, BB/P019137/1, BBS/E/I/00007034, and BBS/E/I/ 00007030 and strategic funding to The Pirbright Institute. We thank the Central Services Unit and the Animal Services department at The Pirbright Institute for their help with preparation of primary chicken kidney cells and the in vivo experiment. A patent has been filed by The Pirbright Institute to protect the intellectual property of this work (EP3172319B1). S.K. and E.B. devised the experimental design and study. S.K., E.B., P.S.-L., G.D., K.F., I.W., A.F., K.L., and H.E. performed the in vitro work. P.S.-L., E.B., S.K., G.D., and I.W. performed the in vivo studies. All authors contributed to the writing of the manuscript.
Funding Information:
This study was funded by UKRI Biotechnology and Biological Sciences Research Council (BBSRC) grants BB/R019576/1, BB/P019137/1, BBS/E/I/00007034, and BBS/E/I/ 00007030 and strategic funding to The Pirbright Institute.
Publisher Copyright:
Copyright © 2022 Keep et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license.