Poster Presentation 14th Lorne Infection and Immunity 2024

Assessment of growth dynamics over time and dispersal responses to nitric oxide by P. aeruginosa biofilms (#113)

Xavier Bertran i Forga 1 , Jilong Qin 1 , Makrina Totsika 1
  1. Queensland University of Technology, South Brisbane, QLD, Australia

The opportunistic pathogen P. aeruginosa is a main causative agent of chronic infections in hospital settings due to its readiness to form biofilms in a multitude of tissues and surfaces. In this state, their tolerance to antimicrobials becomes greatly enhanced. To sensitise biofilms to treatment, dispersal agents such as nitric oxide (NO) have been explored to support the action of antimicrobials due to their ability to revert bacteria to their planktonic phenotype. However, studies exploring the activity and molecular mechanisms activated in P. aeruginosa during NO-mediated dispersal have been performed utilising a wide variety of study designs. This lack of consensus over study design spans over factors such as the utilised platform, biofilm growth time, type of NO donor, concentration and treatment time, all variables that could induce changes in biofilm responses. Due to their high throughput design, closed systems are widely used platforms to monitor changes in biofilm biomass, but due to the limited quantity of nutrients and aeration, biofilm growth can be severely impacted over time, potentially affecting how NO-mediated dispersal is interpreted. In this study, the biofilm growth of two strains of Pseudomonas aeruginosa (PAO1 and PA14) in M9, BM2 and Mueller-Hinton (MH) media was monitored by CFU counting or crystal violet staining over 24h using common plate-based platforms for in vitro biofilm assays in either shaking or static conditions to capture the evolution of biofilm biomass over time in closed systems. Prior to reaching a point of innate dispersal, biofilms were treated with different concentrations of NO donors, and biomass reductions were recorded under different sets of conditions over a span of 2 hours. The goal of this ongoing investigation is to illustrate how growth and treatment conditions can compromise the outcomes that have been collectively identified as dispersal to this day.