| dc.description.abstract | Pseudomonas aeruginosa is an opportunistic pathogen that is known to infect the lungs of Cystic Fibrosis patients, which have abnormally high levels of calcium (Ca2+). Our lab has shown that elevated Ca2+ increases P. aeruginosa plant infectivity and its ability to produce several virulence factors, including pyocyanin, pyoverdine, extracellular proteases, and rhamnolipid. Based on these observations, we hypothesized that elevated Ca2+ enhances P. aeruginosa virulence in an animal host. To test this hypothesis, we have adapted an animal virulence model using larvae of Galleria mellonella, also known as wax worm. First, we aimed to determine if Ca2+ increases the pathogenicity of P. aeruginosa. For this, we determined the half lethal dose (LD50) of P. aeruginosa, which is the number of bacterial cells that cause 50% death rate. This information enables comparison between different testing groups. We also generated killing curves that allow comparing rates of the infection. The worms were injected with PAO1, a wild type of P. aeruginosa, grown in 0 mM, 5 mM, or 10 mM Ca2+. We found the LD50 for 0 mM Ca2+ PAO1 is 2 CFUs, which is two-fold higher than that of PAO1 grown in 5 mM Ca2+. The increase of Ca2+ concentrations to 10 mM Ca2+ further decreased LD50 to 0.3 CFUs. This proves that growth at elevated Ca2+ makes the pathogen more virulent. We also observed that the worms died faster at higher Ca2+ concentrations. We next tested the role of two proteins, CarP and CalC, which previously have been shown to mediate Ca2+ regulation of virulence factor production. The carP::Tn5 mutant exhibited density-dependent and reduced virulence at 10 mM Ca2+. The calC::Tn5 mutant had a slight increase in virulence at 0 mM Ca2+, but a slight reduction at 5 mM Ca2+. These data support the role of CarP and CalC in P. aeruginosa Ca2+ induced virulence. This knowledge enables characterization of Ca2+ regulatory network controlling P. aeruginosa virulence, which is a step towards developing novel strategies to fight P. aeruginosa infections. | |
