Small molecules with antibiofilm, antivirulence and antibiotic synergy activities against Pseudomonas aeruginosa.

Abstract

Biofilm formation is a universal bacterial strategy for long-term survival in nature and during infections. Biofilms are dense microbial communities enmeshed within a polymeric extracellular matrix that protects bacteria from antibiotic exposure and the immune system and thus contribute to chronic infections. Pseudomonas aeruginosa is an archetypal biofilm-forming organism that utilizes a biofilm growth strategy to cause chronic lung infections in Cystic Fibrosis (CF) patients. The extracellular matrix of P. aeruginosa biofilms is comprised mainly of exopolysaccharides (EPS) and DNA. Both mucoid and non-mucoid isolates of P. aeruginosa produces the Pel and Psl EPS, each of which have important roles in antibiotic resistance, biofilm formation and immune evasion. Given the central importance of the Pel and Psl EPS in biofilm structure, they are attractive targets for novel anti-infective compounds. In this study we used a high throughput gene expression screen to identify compounds that repress expression of pel and psl genes as measured by transcriptional lux fusions. Testing of the pel/psl repressors demonstrated an antibiofilm activity against microplate and flow chamber biofilms formed by wild type and hyperbiofilm forming strains. To determine the potential role of EPS in virulence, mutants in pel/psl were shown to have reduced virulence in the feeding behavior and slow killing virulence assays in Caenorhabditis elegans. The antibiofilm molecules also reduced P. aeruginosa PAO1 virulence in the nematode slow killing model. Importantly, the combination of antibiotics and antibiofilm compounds were synergistic in killing P. aeruginosa biofilms. These small molecules represent a novel anti-infective strategy for the possible treatment of chronic P. aeruginosa infections.\n\nAuthor summaryBacteria use the strategy of growing as a biofilm to promote long-term survival and therefore to cause chronic infections. One of the best examples is Pseudomonas aeruginosa and the chronic lung infections in individuals with Cystic Fibrosis (CF). Biofilms are generally a dense community of bacteria enmeshed in an extracellular matrix that protects bacteria from numerous environmental stresses, including antibiotics and the immune system. In this study we developed an approach to identify P. aeruginosa biofilm inhibitors by repressing the production of the matrix exopolysaccharide (EPS) polymers. Bacteria treated with compounds and then fed to the nematode also had showed reduced virulence by promoting nematode survival. To tackle the problem of biofilm tolerance of antibiotics, the compounds identified here also had the beneficial property of increasing the biofilm sensitivity to different classes of antibiotics. The compounds disarm bacteria but they do not kill or limit growth like antibiotics. We provide further support that disarming P. aeruginosa may be a critical anti-infective strategy that limits the development of antibiotic resistance, and provides a new way for treating chronic infections.