Experimental and computational approaches to understand collective behaviors of bacterial pathogen Pseudomonas aeruginosa

Abstract

Pseudomonas aeruginosa is a Gram-negative opportunistic pathogen, estimated to account for 15-20% of hospital-acquired infection-related deaths around the globe in a year. This bacterium exhibits two distinct collective behaviors, biofilm formation and swarming motility. During biofilm formation, the P. aeruginosa population is comprised of sessile cells - covered in a self-produced polysaccharide matrix. Swarming, on the other hand, is group motility facilitated by flagella and bio-surfactant. Such cooperative behaviors help P. aeruginosa to thrive in hostile environmental conditions. However, the triggers that regulate these behaviors remain unknown and form the focus of my Ph.D. thesis. P. aeruginosa forms biofilms on indwelling medical devices such as endotracheal tubes (ETTs), urinary catheters, vascular access devices, tracheostomies, and feeding tubes, often leading to hospital-acquired infections. Pseudomonas aeruginosa is one of the four frequently encountered bacteria causing pneumonia. In the current work, we have established an in vitro model mimicking the biofilm formation on the endotracheal tube. We have identified two-component system (TCS) genes contributing to this process. The TCS comprises a membrane-associated sensor kinase and an intracellular response regulator. We have found that out of 112 TCS mutants studied, 56 had altered biofilm biomass on ETTs. Some of these are novel ETT-specific TCSs that could serve as targets to prevent biofilm formation on indwelling devices frequently used in clinical settings. Swarming in P. aeruginosa is a collective movement of the bacterial population over a semisolid surface, but specific signals that trigger this motility are unclear. We hypothesized that specific environmental signals could induce swarming in P. aeruginosa. Our data show that a low ethanol concentration under nutrient-limiting conditions provides a strong ecological motivation for swarming in P. aeruginosa PA14. Ethanol serves as a signal and not a carbon source under these conditions. Moreover, ethanol-driven swarming relies on the ability of the bacteria to metabolize ethanol to acetaldehyde using a periplasmic quinoprotein alcohol dehydrogenase, ExaA. We found that ErdR, an orphan response regulator linked to ethanol oxidation, is necessary for the transcriptional regulation of a cluster of 17 genes, including exaA, during swarm lag. Finally, we show that as a volatile, ethanol could induce swarming in P. aeruginosa at a distance, suggesting long-range spatial effects of ethanol as a signaling molecule. P. aeruginosa exists in multispecies consortia in the environment and during the infection of various hosts, including humans. The physicochemical properties which mediate interactions between P. aeruginosa and its neighbors remain elusive. We began our study using P. aeruginosa and Cryptococcus neoformans, a pathogenic yeast species, in a surface-based co-culture assay. We found that the P. aeruginosa colony spread more on the lawn of C. neoformans. Upon microscopic investigation, we found that P. aeruginosa shows exploratory behavior in proximity to C. neoformans cells, and this exploratory behavior does not require metabolic active yeast cells. We hypothesize that the fluid accumulation near C. neoformans cells plays an essential role in the microscopic interaction leading to the macroscopic growth of the P. aeruginosa colony. To test this hypothesis, we have developed an individual-based model with experimentally motivated constraints such as microbial division time, fluid accumulation near yeast cells, and the motility of individual P. aeruginosa and scaled the parameters to simulate macroscopic behavior using the cellular automata modeling approach. We found that the presence of yeast lawn allowed P. aeruginosa to cover more area by utilizing the fluid accumulated around yeast cells.