Biogenesis of outer-membrane vesicles in Bordetella species

The Gram-negative bacterium Bordetella pertussis causes whooping cough, also known as pertussis, which is an infection of the respiratory tract in humans. Pertussis is most severe in newborns. Currently, babies are vaccinated against pertussis, but these vaccines do not protect sufficiently as appears from the increase in pertussis cases over the last two decades. A closely related bacterium, Bordetella bronchiseptica, is the causative agent of respiratory diseases in animals, such as atrophic rhinitis in pigs and kennel cough in dogs. As for pertussis, the effectiveness of the available vaccines against B. bronchiseptica is under debate. Therefore, new vaccines against these pathogens need to be developed. A promising strategy is the use of outer-membrane vesicles (OMVs). These vesicles are naturally shed by Gram-negative bacteria. OMVs are safe because they are unable to replicate, they contain the most relevant antigens, and they are small which facilitates their uptake by immune cells. Unfortunately, spontaneous OMV production by Bordetella is too low for cost-effective vaccine production. In the work described in this thesis, we investigated the formation of OMVs in Bordetella and explored ways to increase OMV production. We show that OMV production can be increased by applying a heat shock to the bacteria and by genetically modifying the composition of the outer membrane or reducing the anchoring of the outer membrane to the underlying layers. The different approaches to increase OMV production resulted in OMVs with different compositions. In the future, the protection induced by the different OMVs needs to be determined.