NtrBC selectively regulates adaptation and intercellular interactions of Pseudomonas aeruginosa

Pseudomonas aeruginosa is an opportunistic pathogen that causes nosocomial and chronic infections contributing to morbidity and mortality in skin wound and cystic fibrosis patients, respectively. One of the reasons for its success as a pathogen is its ability to adapt to a broad range of circumstances. Here, the regulatory two-component system NtrBC is shown to be involved in adaptive and pathogenic states of P. aeruginosa. Characterization of adaptive lifestyles in vitro confirmed that the double ΔntrBC mutant demonstrated a nearly complete inhibition of swarming motility, a modest decrease and alteration of surfing motility and >40% reduction in biofilm formation; except for swarming, single mutants generally had more subtle or no changes. The P. aeruginosa ΔntrBC mutant also had a major increase (~10-fold) in susceptibility to ciprofloxacin. Transcriptional profiles of deletion mutants were defined using RNA-Seq and unveiled dysregulated expression of hundreds of genes implicated in P. aeruginosa virulence during chronic lung infections, as well as carbon and nitrogen metabolism. The role of NtrBC in host-pathogen and interspecies interactions was also examined. P. aeruginosa mutants exhibited distinct host interactions, including modestly increased cytotoxicity toward human bronchial epithelial cells, reduced virulence factor production and 10-fold increased uptake by macrophages in vitro. In a high-density skin infection model, mutants were reduced in their ability to invade or cause damage to tissue and were more susceptible to ciprofloxacin in vivo. To compare the infectivity of strains across tissues, and pre-clinically screen antimicrobial or immunomodulatory therapies for the treatment of sinusitis, a murine model of upper respiratory tract infection was developed. In contrast to the wild-type levels of colonization observed in the abscess model, P. aeruginosa mutants colonized the respiratory tract less well than wild-type. In contrast to wild-type, ΔntrC and ΔntrBC mutants outcompeted Staphylococcus aureus, a commonly co-isolated species in skin wounds and cystic fibrosis patients, during planktonic and biofilm growth. These results indicate that NtrBC is a global regulatory system involved in both adaptive and pathological processes relevant to the success of P. aeruginosa in infection.Science, Faculty ofMicrobiology and Immunology, Department ofGraduat