In vitro characterization of Actinobacillus pleuropneumoniae outer membrane vesicles

PhD student: Zhuang Zhu, zhuangzhu@sund.ku.dk 
Thesis defended: 16 August 2022

Background

Outer membrane vesicles (OMVs) open up a new avenue different from any other strategies used before in vaccine research. OMVs are non-replicative sphere structures originating from the outer membrane of Gram-negative bacteria. Their inherent adjuvanticity, antigenicity, and flexibility as delivery platforms, which have been investigated in a wide range of bacterial species, endow OMVs with excellent potential in vaccine development. Our research group has previously exploited the potency of A. pleuropneumoniae OMVs as a vaccine prototype. However, the immunization of pigs with OMVs was not able to confer protection in spite of strong antibody response raised, suggesting unknown roles that OMVs play in immune responses.

Purpose of the project

The main objective of this PhD project was to increase our understanding of OMVs from A. pleuropneumoniae in terms of their proteomic information and immunomodulatory ability, laying the foundation for future development of OMV-based vaccines.

Results

The first study presented is a literature review summarizing the potential of OMVs in vaccine development, aiming at comprehensively understanding the underlying mechanisms of the pluripotency of OMVs in vaccine research. In the second study, a quantitative proteomic study was performed to compare the protein compositions and abundance in OMVs extracted from A. pleuropneumoniae MIDG2331, ΔnlpI, ΔpalA, and ΔnlpIΔpalA mutants. In these mutants, relevant genes were deleted to produce more and/or immunodominant antigen-free OMVs. Meanwhile, we demonstrated that A. pleuropneumoniae OMVs can be internalized by porcine alveolar macrophages, and are not cytotoxic to them. Furthermore, for the first time, we demonstrated that A. pleuropneumoniae OMVs exert an inhibitory role in the immune responses of porcine alveolar macrophages stimulated by either dead or live A. pleuropneumoniae cells, suggesting that OMVs may represent a new virulence factor in A. pleuropneumoniae.