Selection Versus Mutation: Reducing the Risk of Vaccine Reversion

Vaccination against numerous endemic pathogens is an essential component of the poultry industry. Without these vaccines chickens would succumb to infection at an early age reducing the productivity of the industry well below sustainable levels. IBV is an endemic virus that causes severe disease outbreaks in chickens worldwide. Effective and economically viable vaccines against IBV are available and mainly produced from pathogenic virus strains by passing in eggs approximately one hundred times. During these passages the virus accumulates multiple sequence variations from the original pathogenic sequence. This ultimately leads to attenuation of the virus and the production of a live attenuated vaccine. These vaccines have lost their ability to cause disease but still elicit a protective immune response in the chicken, thus protecting the bird from future infections. However, as these are live viruses the potential for to revert back to a pathogenic form is considerable considering the few sequence changes between wild and vaccine strains. Despite the importance of these vaccines to the poultry industry and the risk of reversion, the processes that occur and the selective forces that drive virus attenuation during egg passage are unknown. Importantly, the differential contribution of virus sequence mutation compared to the selection of minor variants already present in the virus population has not been determined. Understanding these basic processes is essential to the development of future vaccines to reduce the threat of reversion. This study will use passaged pathogenic IBV strains produced in the same way as vaccines. In parallel we will use a unique system that allows us to passage a single virus clone rather than a mixed virus population. Using contemporary deep sequencing technology we will study the molecular changes that occur at fine resolution during the attenuation process. This will for the first time reveal how a mixed population of virus changes during vaccine manufacture and the extent to which individual viruses can mutate. These results will then inform a series of studies that manipulates the forces that drive virus change. The first will use IBV strains that contain a protein from another strain that influences the immune response in the chicken, and the second will use viruses that mutate much faster than wild type viruses. By passaging and deep sequencing these viruses in the same way as the wild type viruses, we will understand how different forces drive virus sequence mutation. These recombinant passaged viruses will then be tested to determine if this process has led to attenuation and also if they maintain the potential to infect other chickens that are exposed to vaccinated birds. Ultimately this research will reveal how IBV is attenuated by egg passage and identify key regions of the genome that prevent the virus from causing disease but do not impair its potential as a vaccine. Moreover, we will further develop our understanding of how different pressures influence the attenuation process and potentially identify ways to improve the process of vaccine design by engineering attenuated viruses. By understanding and manipulating the processes that govern virus attenuation and vaccine production we aim to identify ways of reducing the danger of vaccine strains reverting and causing damaging disease outbreaks.