Bacterial interaction with the microvasculature, a target for therapeutic intervention during septicaemia (BactInfectERA)

Bacterial septicaemia remains a prominent cause of death in developed countries. An essential feature in the pathogenesis of invasive systemic bacteria is their ability to interact with the microvasculature, which allows dissemination, local activation of coagulation pathways and vascular leakage. Paradoxically, these aspects remain poorly explored. Among major issues to be solved are the host molecular and cellular events leading to vascular dysfunction, thrombosis, organ failure and immune escape by these bacterial pathogens. Neisseria meningitidis (meningococcus) is an example of invasive bacterial pathogen responsible for dissemination into different organs including meninges and skin. Specific interaction of this bacterium with the human microvasculature is responsible for thrombotic lesions, which, in the most extreme cases, lead to purpura fulminans, a life-threatening syndrome associating septic shock with vascular leakage and extensive thrombosis leading to death in 30% of the patients. Taking N. meningitidis as a paradigm for invasive extracellular pathogens interacting with human microvessels, the goal of this proposal is to integrate high throughput sequencing methods, combined with molecular, immunological and imaging approaches, to decipher the initial events of meningococcal infection in vivo and to understand the mechanisms by which this pathogen subverts the innate immune response and adapts to the human host. This collaborative project will be conducted by 4 groups with extensive and complementary experience in the fields of vascular biology and infection. In WP1 we will identify the mechanisms by which meningococcus adapts to its host. Utilizing a whole-genome transposon mutant library in combination with massively parallel sequencing we will analyse bacterial genes involved in vascular colonization in vivo. For bacteria at different stages of in vivo infection we will perform a longitudinal large scale profiling of the transcriptional and genomic modifications. In WP2 we will characterize the mechanisms involved in vascular dysfunction and thrombosis. For endothelial and surrounding cells collected by laser capture microdissection from non- infected and infected human skin grafts we will analyze the transcriptomes for gene expression profiling. Computational analysis will lead to the identification of key factors involved in host responses. In WP4 we will establish a mathematical modelling of the meningococus-host interaction. Taking advantage of the genomic and transcriptomic data from WP1 and WP2 and connecting it to the knowledge from public sequence and pathway databases, we will establish a mathematical model of bacterial virulence and host-pathogen interactions. Computational interpretation of the data will be confronted to the experimental investigations performed by the other teams. This consortium will investigate the transcriptional control of meningococcal virulence factors, identify the bacterial genomic changes that occur upon infection and provide a global overview of the host response with a special focus on pathways that give rise to the characteristic symptoms of vascular damage. Genetic targeting and/or pharmacologic blockade of the implicated pathways will be a future strategy for therapeutic adjunctive measures to improve outcome and may hold substantial promise, in concert with antimicrobial agents, in humans with septicaemia.

Neisseria meningitidis is a cause of meningitis epidemics worldwide and of rapidly progressing fatal septic shock. A crucial step in the pathophysiology of invasive bloodborne meningococci is their adhesion to and proliferation in both peripheral and brain blood microvessels, a process referred to as vascular colonization. Hence, the overall objective of this project was to analyse the mechanisms and consequences in vivo of such colonization on vascular remodelling and alteration. To address these specific questions, we took advantage of a novel and pertinent in vivo model of meningococcal infection we developed. Combined with novel next-generation deep-sequencing approaches, we have conducted an unbiased and global characterization of the associated gene expression changes in both the pathogen and the host.