Identification of small molecules that interfere with c-di-GMP signaling and induce dispersal of Pseudomonas aeruginosa biofilms

Microbial biofilms are involved in a number of infections that in many cases cannot be resolved, since microbes in biofilms display resistance to host immune defenses and antibiotic therapy. Accordingly, novel drugs for efficient combat of biofilm-based infections are urgently needed. We employed a high throughput screening approach for identification of chemical compounds that can reduce the intracellular c-di-GMP content in Pseudomonas aeruginosa. This led to the identification of a small molecule that efficiently deplete P. aeruginosa for c-di-GMP, inhibit biofilm formation and disperse established biofilm. A combination of the anti-biofilm compound and antibiotic was demonstrated to result in efficient eradication of biofilms in vitro, as well as in a murine biofilm infection model. Genetic analysis provided evidence that the c-di-GMP phosphodiesterase BifA in P. aeruginosa is central for the activity of our anti-biofilm compound. RNA-seq analysis of the effect of the anti-biofilm compound on the transcriptome of P. aeruginosa demonstrated that known c-di-GMP regulated genes were down-regulated, and that no virulence genes were up-regulated. The effect of the anti-biofilm compound was restricted to P. aeruginosa, and it did not show activity on a range of other bacterial species. The identified compound might serve as lead compound for the development of drugs that can be used in combination with antibiotics for the treatment of P. aeruginosa biofilm infections. The work constitutes proof of concept for c-di-GMP phosphodiesterase activation as an anti-biofilm strategy.