PORTEFEUILLE

The airways of patients with Cystic Fibrosis (CF) are colonized early in their lives with the opportunistic pathogen Pseudomonas aeruginosa. Although P. aeruginosa infections can be treated with antibiotics, full clearance of the infection is not possible. This results in chronic infections and a gradual decline of lung function representing the main cause of mortality in CF patients. The molecular basis for the remarkable persistence of P. aeruginosa is poorly understood. Recent findings from my lab introduced a fundamentally novel concept for bacterial persistence. We postulate that the bacterial second messenger c-di-GMP is a key determinant of chronic infections of CF airways and that continuous genetic adaptations of the c-di-GMP network facilitate P. aeruginosa persistence during lung colonization. This discovery offers the potential to specifically interfere with chronic infections in CF lungs.

We propose a multidisciplinary approach to analyze the role of c-di-GMP in persistence in more detail. We will analyze how c-di-GMP contributes to the emergence of persistent morphotypes of P. aeruginosa that are strongly linked to chronic behavior and poor lung function. Deep sequencing of clinical isolates and strains from experimental evolution experiments will reveal genetic changes within the c-di-GMP network facilitating persistence. Analysis of samples from patients undergoing chemotherapy will lend insights into the selective forces shaping P. aeruginosa adaptation to the chronic CF environment. In parallel, we will follow up on recent observations that have linked c-di-GMP directly to reduced metabolic activity and exopolysaccharide biosynthesis in P. aeruginosa, two prime candidate persistent factors. Finally, we will exploit our expertise in c-di-GMP signaling to develop, in collaboration with partners in the biotech industry, diguanylate cyclase inhibitors as potential anti-persistence drugs. These studies will provide an entry point into defining the molecular mechanisms of bacterial persistence in CF airways and will pave the way for novel and innovative approaches to interfere with chronic infections and infection relapses.

Despite of recent progress in treating CF patients with available antibiotics, eradication of chronic forms of P. aeruginosa is not possible. Our studies introduce and validate a novel molecular concept for persistence of P. aeruginosa in long-term CF infections. By contributing to a better understanding of the causes for this remarkable form of microbial persistence, our research will help to develop diagnostic tools for chronic behavior of P. aeruginosa and will lay the foundation for chemical interference with the primary cause of CF mortality.

During the initial phase of this project we focused on the characterization of novel c-di-GMP effector proteins in Pseudomonas aeruginosa. These studies led to the identification of a novel pathway involved in P. aeruginosa surface colonization and biofilm formation and its regulation by c-di-GMP. Undergoing efforts attempt to fully define this pathway on the molecular and cellular level and to decipher its significance for the chronic potential of P. aeruginosa.

In parallel, we are studying the genetic determinants responsible for persistent behavior of P. aeruginosa in CF lungs. This includes determinants of chronic virulence and biofilm formation as well as mechanisms of drug tolerance, the ability of pathogenic bacteria to survive the exposure to high doses of antibiotics without developing resistance. These studies exposed the central importance of c-di-GMP for persistence of this pathogen during infections in humans. Our studies also disclosed drug tolerance as an important mechanism of in-patient adaptation that might largely contribute to treatment failure in chronically infected patients.

Broder, U. et al. LadS is a Calcium-Responsive Kinase that induces Acute-to-Chronic Virulence Switch in Pseudomonas aeruginosa. Nature Microbiology, in press;
Malone, J. G. et al. The YfiBNR Signal Transduction Mechanism Reveals Novel Targets for the Evolution of Persistent Pseudomonas aeruginosa in Cystic Fibrosis Airways. PLoS Pathog 8, e1002760 (2012).