Fluorescence-based reporter for gauging cyclic di-GMP Levels in Pseudomonas aeruginosa.

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The increased tolerance towards the host immune system and antibiotics displayed by biofilm forming Pseudomonas aeruginosa and other bacteria in chronic infections such as cystic fibrosis bronchopneumonia is of major concern. Targeting biofilm formation is believed to be a key aspect in the development of novel anti-pathogenic drugs that can augment the effect of classic antibiotics by decreasing antimicrobial tolerance. The second messenger cyclic di-GMP is a positive regulator of biofilm formation, and cyclic di-GMP-signaling is now regarded as a potential target for the development of anti-pathogenic compounds. Here we describe the development of fluorescent monitors that can gauge the cellular level of cyclic di-GMP in P. aeruginosa. We have created c-di-GMP level reporters by transcriptionally fusing the cyclic di-GMP responsive cdrA promoter to genes encoding Gfp. We show that the reporter constructs give a fluorescent read-out of the intracellular level of cyclic di-GMP in P. aeruginosa strains with different levels of cyclic di-GMP. Furthermore, we show that the reporters are able to detect increased turnover of cyclic di-GMP mediated by treatment of P. aeruginosa with the phosphodiesterase inducer nitric oxide. Considering biofilm formation a necessity for the subsequent development of a chronic infection and therefore a pathogenicity trait, the reporters display a significant potential for use in the identification of novel anti-pathogenic compounds targeting cyclic di-GMP-signaling, as well as for use in research aiming at understanding the biofilm biology of P. aeruginosa.
Original languageEnglish
JournalApplied and Environmental Microbiology
Issue number15
Pages (from-to)5060-5069
Number of pages10
Publication statusPublished - 2012

ID: 38229970