Abstract
The Brucella genus comprises intracellular pathogenic bacteria from the α-proteobacteria class. The present study focused on Brucella abortus (B. abortus), the causative agent of bovine brucellosis, which is responsible for huge economical losses every year in developing countries. As this bacterium is suspected to face multiple envelope-related stresses, a Tn-seq on deoxycholate (DOC) was performed prior to this work. This identified a two-component system (TCS), CenKR, as being required for growth on DOC. The goal of this master thesis was thus to unravel how this TCS may help B. abortus to deal with DOC and other envelope-related stresses. In this prospect, markerless ΔcenR and ΔcenRΔcenK strains were generated. They displayed growth defects on DOC, suggesting both genes are indeed involved in an envelope stress response (ESR) in B. abortus. We then aimed at further characterizing CenKR by testing other stresses that may impact the bacterial envelope. Our results suggest that CenKR is responsible for resistance to multiple envelope-related stresses in B. abortus.These include surfactants, β-lactam antibiotics and macrophage infection. Surprisingly, the opposite seems true for polymyxin B (PxB) resistance in B. abortus as the absence of CenKR improved survival when PxB was present in the media. The strains morphology was also visualized by phase-contrast microscopy to assess potential defects. Bean-shaped bacteria in the ΔcenR population and even swellings in the ΔcenRΔcenK could be identified. This further indicates CenKR is required for the envelope’s integrity in B. abortus, as shown by another study in Caulobacter crescentus. We then aimed at investigating CenR-regulated genes in B. abortus. Since a putative CenR DNA-binding motif was previously identified in R. sphaeroides, we used it to perform an analysis with RSA tools on B. abortus genome. Two particularly interesting targets were identified by this analysis. First, a gene encoding the transcriptional repressor BepR known to repress the bepDE operon in Brucella suis (B. suis). Second, dnaG, an essential gene that encodes for the DNA primase required for bacterial replication. We therefore hypothesize that CenR might repress the expression of both dnaG and bepR when B. abortus faces envelope-related stresses. This may respectively allow (1) a growth arrest during the envelope stress and (2) the formation of BepCDE, a putative tripartite efflux pump that could be involved in a MDR phenotype as previously describe in B. suis. Still, experimental evidences would be required to further characterize CenKR in B. abortus. Finally, this TCS appear well conserved in α- proteobacteria and particularly in Rhizobiales. It could thus be interesting to get further knowledge of it, so that we may compare its functioning and roles in B. abortus to what was previously characterized in other α-proteobacteria.
| Date of Award | 19 Jan 2023 |
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| Original language | English |
| Awarding Institution |
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| Supervisor | Xavier De Bolle (Supervisor) & Adelie Lannoy (Co-Supervisor) |