Phenazine production enhances extracellular DNA release via hydrogen peroxide generation in Pseudomonas aeruginosa

Abstract

In Pseudomonas aeruginosa eDNA is a crucial component essential for biofilm formation and stability. In this study we report that release of eDNA is influenced by the production of phenazine in P. aeruginosa. A ∆phzA-G mutant of P. aeruginosa PA14 deficient in phenazine production generated significantly less eDNA in comparison with the phenazine producing strains. The relationship between eDNA release and phenazine production is bridged via hydrogen peroxide (H2O2) generation and subsequent H2O2 mediated cell lysis and ultimately release of chromosomal DNA into the extracellular environment as eDNA.

Keywords: Pseudomonas aeruginosa; extracellular DNA; hydrogen peroxide; phenazines.

Figure 1. Schematic represents quorum sensing controlled expression of genes for phenazine production. P. aeruginosa synthesizes acylated homoserine lactones (AHLs) and Pseudomonas quinone signaling (PQS) as their primary and secondary quorum sensing signaling molecules. PQS regulates the synthesis of phenazine-1-carboxylic acid (PCA) through a set of primary phenazine producing genes phzA1-G1 and phzA2-G2. PCA then converts into various kinds of specific phenazine encoded by specific genes.

Figure 2. Different kinds of phenazine production and its influence in eDNA release. Comparison of bacterial colony formation on LB agar plate after 48 h incubation with non-filtered (upper half) and filtered (bottom half) bacterial supernatant (A). Production of various kinds of phenazine molecules by P. aeruginosa PAO1 strains (B). Quantification of eDNA release in supernatants of various kinds of phenazine producing P. aeruginosa PAO1 (C) and PA14 (D) strains. Error bars represents standard deviations from the mean (n = 3). Asterisks indicate statistically significant (p < 0.05) differences in eDNA concentration in comparison to the PAO1 wildtype (C) and mutant strain ∆phzA-G (D). Schematic represents the relationship between eDNA release and phenazine production is bridged via H₂O₂ generation and subsequent H₂O₂ mediated cell lysis (E).