Functional genetics of Pseudomonas aeruginosa stress-response systems and their relevance in bacteriophage defense

Pseudomonas aeruginosa is an increasingly problematic opportunistic human pathogen that causes recalcitrant chronic infections in susceptible individuals. P. aeruginosa’s success as a pathogen is in part owed to its sophisticated stress-response systems and other regulatory circuits that respond to environmental changes, which endows P. aeruginosa with its notorious resilience and ecological versatility. Bacteriophage therapy is a suggested alternative or complementary treatment strategy against this pathogen, as it has some specific advantages such as anti-biofilm activity. However, gaps in understanding of P. aeruginosa bacteriophage defense systems have previously been shown to be problematic in implementation of phage therapy. More research to guide the design and implementation of phage therapy is warranted.

In this thesis, current knowledge of select P. aeruginosa stress-response functional genetics is introduced as well as bacteriophage defense systems in P. aeruginosa and other bacteria. Overlaps between bacteriophage defense and stress-response systems is discussed as a possibly understudied facet of bacteriophage defense. I summarize and discuss my own contributions to this field which included a transposon sequencing screen for bacteriophage defense genes in P. aeruginosa strain PAO1. PA3040 operon, an undercharacterized member of the P. aeruginosa cell envelope stress response, was identified as a putative bacteriophage defense locus. While a stress-related phenotype could not be identified in PAO1, PA3040 operon overexpression was found to cause an aberrant colony morphology and reduced biofilm formation. Promoter induction experiments also revealed PA3040 expression in response to phage infection and high-density culture. In a recently isolated clinical strain designated PAC3, we found that PA3040 operon deletion caused tobramycin tolerance, changes in biofilm formation, and increased pyocyanin production. A phage-related phenotype was not confirmed in either strain, but suitable experiments to investigate this are suggested.