Project description:This study characterised the Ser/Thr/Tyr phosphoproteome of classical Bordetella species and examined its role in Bordetella biology and virulence. This study found 70 unique phosphorylated proteins in the classical bordetellae group with a high degree of conservation observed and phosphorylation was a key regulator of Bordetella metabolism with proteins involved in gluconeogenesis, TCA cycle, amino acid and nucleotide synthesis significantly enriched. We also identified the phosphorylation of three key virulence pathways which separates classical from non-classical bordetellae including the type III secretion system, alcaligin synthesis (the primary siderophore produced by Bordetella) and the BvgAS master transcriptional regulatory system for virulence genes in Bordetella. Seven new phosphosites were identified in BvgA with 6 located in the DNA binding domain. Of the 7 new phosphosites, four were not detected in non-classical bordetellae. This suggests that serine/threonine phosphorylation may play an important role in stabilising or destabilising BvgA binding to DNA for fine-tuning of virulence gene expression and that BvgA phosphorylation may be an important factor that separates classical from non-classical bordetellae. This study provides the first insight into the phosphoproteome of classical Bordetella species and the role Ser/Thr/Tyr phosphorylation plays in Bordetella biology and virulence.
Project description:The genomes of three closely related bordetellae are currently being sequenced, thus providing an opportunity for comparative genomic approaches driven by an understanding of the comparative biology of these three bacteria. Although the other strains being sequenced are well studied, the strain of Bordetella parapertussis chosen for sequencing is a recent human clinical isolate (strain 12822) that has yet to be characterized in detail. This investigation reports the first phenotypic characterization of this strain, which will likely become the prototype for this species in comparison with the prototype strains of B. pertussis (Tohama I), B. bronchiseptica (RB50), and other isolates of B. parapertussis. Multiple in vitro and in vivo assays distinguished each species. B. parapertussis was more similar to B. bronchiseptica than to B. pertussis in many assays, including in BvgS signaling characteristics, presence of urease activity, regulation of urease expression by BvgAS, virulence in the respiratory tracts of immunocompromised mice, induction of anti-Bordetella antibodies, and serum antimicrobial resistance. In other assays, B. parapertussis was distinct from all other species (in pigment production) or more similar to B. pertussis (by lack of motility and cytotoxicity to a macrophage-like cell line). These results begin to provide phenotypes that can be related to genetic differences identified in the genomic sequences of bordetellae.
Project description:Comparative genomic hybridization (CGH) of 71 Bordetella parapertussis strains isolated from either humans or sheep, and 1 B. bronchiseptica strain isolated from a rabbit
Project description:The classical bordetellae (Bordetella pertussis, B. parapertussis, and B. bronchiseptica) are obligate aerobes that use only oxygen as their terminal electron acceptor for electron transport-coupled oxidative phosphorylation. Therefore, access to oxygen is critical for these bacteria to survive. To better understand how B. bronchiseptica changes its gene regulation when faced with different levels of oxygen, we grew liquid cultures of B. bronchiseptica RB50 in ambient air, 5% oxygen, and 2% oxygen. We also measured how the presence of 5% carbon dioxide affected gene expression in these bacteria, since they are respiratory pathogens and therefore get exposed to higher carbon dioxide levels during infection than are found in ambient air.
