Characterization of heme homeostasis and impact on virulence and colonization of GRAM positive bacteria ; Caractérisation de l'homéostasie et de l'impact de l'hème sur les capacités de virulence et de colonisation de bactéries à GRAM positif

Heme is a redox-reactive molecule with essential function in bacterial metabolism. However, this molecule generates reactive oxygen species responsible for its toxicity. We characterized the mechanism of heme homeostasis involving the efflux transporter HrtBA. In L. lactis, we demonstrated that HrtBA prevents from membrane and intracellular accumulation of internalized exogenous heme thanks to a menaquinone dependent mechanism. HrtBA is also present in several pathogens. In S. agalactiae, the transcription of HrtBA is regulated by a two-component system HssRS. The HssS sensor recognizes internalized exogenous heme. To clarify the role of heme of the host in the virulence of S. agalactiae, a systemic infection model in mice using luminescence (lux) and in vivo imaging (IVIS) has been set up. The monitoring of luminescence generated by heme hypersensitive (ΔhrtBA) bacteria shows that heme of host is toxic and that the capability of S. agalactiae to control heme homeostasis is crucial for infection. In the same way, by demonstrating that respiratory metabolism is crucial for infection (ΔcydA), we demonstrated that S. agalactiae depends on its capacity to acquire the heme of the host to become infectious. By using the HrtBA promoter coupled with lux operon, we studied the capacity of S. agalactiae to detect and to acquire heme in vivo during the infection. Our results show that host heme is especially biodisponible in the liver. On the contrary, heme is not detected by bacteria in the brain. Our results prove that heme of the host is an important parameter for the adaptation of S. agalactiae to its host during infection. Blocking HrtBA or heme sensor HssS could so be a target for antibiotic research against S. agalactiae and other pathogens. Finally, we show in E. faecalis that HrtBA expression also depends on a two-component system. We used the same strategy as in S. agalactiae to create a specific heme sensor that allowed us to demonstrate for the first time that E. faecalis meets and uses heme in the digestive ...