Using synthetic biology to generate novel vaccines against avian infectious bronchitis virus (IBV)

Chickens are vaccinated multiple times against poultry pathogens throughout their lives to protect against different diseases. Infectious bronchitis virus (IBV) causes a respiratory disease in chickens making them more susceptible to bacterial infections. In addition to this, chickens infected with IBV gain less weight and produce fewer high quality eggs. It is important to vaccinate against IBV both for welfare and economic reasons. There are a number of problems with the vaccines available against IBV. There are many different strains of IBV circulating around the globe however currently available vaccines do not cross-protect against all strains. New vaccines are produced by growing a virulent strain of IBV in hen's eggs multiple times. This reduces the pathogenicity of the vaccine strain when it is given to chickens but it increases the pathogenicity of the vaccine strain for chicken embryos. Vaccines against IBV are administered by spraying poultry flocks with the vaccine or by putting the vaccine in the drinking water. It is hard to guarantee that each chicken has received the same dose, which make the vaccines less effective.
We have rationally attenuated three strains of IBV using molecular tools. These strains have not been adapted to grow in eggs so are less pathogenic for chicken embryos. This study proposes to establish whether the rationally attenuated strains of IBV could be used as vaccines, by testing the safety, stability and efficacy of all three potential vaccine strains. These novel vaccines have the potential to be given to chicks while they are still developing in the egg so that the chicks are protected against IBV infection even before they hatch. This would guarantee that each chick received the same dose of vaccine and reduce the possibility of vaccine breakdown, which would be very beneficial to the poultry industry. Safer and more efficacious vaccines against IBV will reduce the amount of antibiotics given to chickens to treat secondary bacterial infections resulting from IBV infection. The prolific use of antibiotics in farming may contribute to the development of antibiotic resistance in bacteria, a potentially devastating problem.
The second part of the study proposes to use molecular tools to insert the surface protein, the spike, from a different strain of IBV into one of the novel vaccine strains. The spike is the major structure on the outside of the virus that chickens develop antibodies against. It is highly variable between different strains of IBV so antibodies produced against one strain of IBV may not cross-protect against a new strain of IBV. The spike of one of the most economically damaging strains of IBV will be inserted into a vaccine strain. We will assess whether vaccinating with these recombinant viruses is able to protect against challenge with pathogenic virus containing the same spike or with a different spike. We will establish whether the two viruses containing different spikes are able to grow in competition with each other in eggs. This will investigate the potential to vaccinate eggs with both viruses; protecting against two or more important strains of IBV in a single dose.
The overall aim of this study is to use synthetic biology to generate a new generation of safer, more efficacious vaccines against the important poultry pathogen, infectious bronchitis virus.

The overall aim of this project is to use synthetic biology to generate a new generation of safer, more efficacious vaccines against the important poultry pathogen, infectious bronchitis virus. We will establish whether recombinant infectious bronchitis viruses have the potential to be safe and efficacious vaccines and/or the have ability to provide cross-protection between different serotypes. We have previously developed a reverse genetics system based on a pathogenic strain of infectious bronchitis virus (IBV), M41, and have used this to generate three attenuated isogenic recombinant IBVs (rIBV). This study proposes to evaluate the vaccine potential of the three rIBV in poultry. Vaccine replication and pathology induced in the trachea, kidney and reproductive tract of infected chickens will be assessed, as will the spread of vaccine to sentinel chickens. Vaccination-challenge experiments will be conducted to monitor vaccine efficacy.
We propose to use reverse genetics to swap the spike glycoprotein of the M41 strain of IBV for that of another economically important strain and will characterise the chimaeric virus in vitro, assessing the serotype and cellular tropism. To investigate how the recombinant viruses replicate in competition with each other in eggs we will use next generation sequencing to assess recombination events between the spike genes and determine the feasibility of vaccinating with both viruses in ovo. In vivo vaccination-challenge experiments will be conducted to assess whether vaccination with rIBV expressing spikes from two different serotypes together induces homologous protection and heterologous cross-protection.
The stability of all of the attenuated rIBV isolates will be assessed in ex vivo tracheal organ cultures and maternally derived antibody (MDA) positive eggs using next generation sequencing to model vaccine persistence and evolution of the viral population in poultry flocks under immune pressure.

Poultry is an important food source worldwide and the global poultry industry has expanded in recent years to accommodate increasing demands for healthy, sustainable protein sources from a rising population. The FAO estimates that poultry accounts for 88% of global meat production. Viral diseases such as infectious bronchitis are a constant threat to the poultry industry through reduced meat production from broiler chickens, production of fewer, good quality eggs and animal welfare issues. In fact, infectious bronchitis is the single most economically important infectious disease affecting chickens in the UK. Improved vaccines against IBV would benefit the UK economy as an estimated £23M is spent on control of IBV per annum.
Vaccine breakdown would have a major effect on the UK poultry industry, not only in terms of bird welfare and production costs, but also associated risks to food security. This work will establish whether our novel approach to vaccine design through rational attenuation and modification of vaccine serotype is capable of producing safe and efficacious vaccines for the control of IBV that are less likely to revert to virulence. The development of safer vaccines will reduce the amount of antibiotics used to treat secondary bacterial infections associated with infectious bronchitis, which would have positive environmental impact.
The results of this study will inform approaches to development of other veterinary and human vaccines. This study will benefit The Pirbright Institute, BBSRC and Zoetis, and the wider academic community including other researchers working in the fields of molecular virology, livestock health and coronavirus research, particularly IBV. The project will provide training in virology, deep sequencing, confocal microscopy and in vivo studies. The results will be of direct benefit to the poultry industry and vaccine developers. Knowledge generated by this project will be widely disseminated to the research community through peer-reviewed journals and presentations at national and international virology conferences and interactions with members of the poultry industry and veterinarians.