Identification of virulence factors as novel vaccine targets for contagious bovine pleuropneumonia by whole genome saturated mutagenesis
Project summary
There is an urgent need for improved vaccines for contagious bovine pleuropneumonia (CBPP). CBPP is one of the most infectious and highly contagious diseases of cattle in Africa accounting for more than $38.8 million annual loss in cattle productivity in 12 endemic countries in sub-Saharan Africa. Currently, control of CBPP relies on a live vaccine of limited efficacy and occasional severe side effects. Achieving breakthroughs in developing effective tools, including vaccines, for CBPP intervention requires deeper insights into host-bacteria interactions.
Like other diseases caused by mycoplasma, CBPP is characterized by immunopathology, i.e. dysregulation of the host’s immune response. In vitro data indicates that this dysregulation begins at the very early stages of infection, at the initial site of bacteria-host cell interaction in the lung. We aim towards identifying vaccine candidates that can disrupt the bacteria-induced dysregulation of the host’s immune response.
To pump-prime this objective, we propose to generate a random mutant library of the bacteria which can be used to identify bacterial genes that play an important role in the initial bacteria-host cell encounters and the dysregulation of the host’s immune response. Within the scope of the initial project, we will use the mutant library in established in vitro assays to screen for bacteria that fail to induce the typical response (compared to the wild type bacteria). This will allow a deeper understanding of host-pathogen interactions, and the identified antigens will be tested in future in vivo experiments for their suitability as vaccine candidates.
Project outcomes
Contagious bovine pleuropneumonia (CBPP), caused by Mycoplasma mycoides subsp. mycoides (Mmm), is the most important infectious disease affecting cattle in Africa, causing an estimated loss in cattle productivity of $40 million annually. Currently, control of CBPP relies on a live vaccine of limited efficacy and occasional severe side effects, and there is an urgent need for improved vaccines. However, achieving breakthroughs in developing effective tools such as vaccines for CBPP intervention requires deeper insights into host-bacteria interactions. Like other diseases caused by mycoplasma, CBPP is characterized by immunopathology, i.e. dysregulation of the host’s immune response. We aim towards identifying Mmm genes that are involved in the misleading of the host’s immune response, to later be able to use this information to develop vaccines that can disrupt the process and instead allow the host to induce an immune reaction that can clear the infection.
During the pump-priming project we have generated a limited genome-wide transposon mutant library of more than 800 individual mutants, using random integration of a transposon containing plasmid. We were able to show that integration was successful as the mutant colonies were fluorescent as designed, and the integration site of one of the mutants was identified. This work is continuing with the aim of generating at least 3000 individual mutants, to ensure that every non-essential gene in the Mmm genome will be disrupted in at least one mutant.
This library will be used to identify bacterial genes that play an important role in the initial bacteria-host cell encounters and the dysregulation of the host’s immune response. A host cell cytotoxicity assay has been established and will be used to screen the library for Mmm genes that are involved in killing lung epithelial cells.