Journal article
Fluorescence-Based Reporter for Gauging Cyclic Di-GMP Levels in Pseudomonas aeruginosa
The increased tolerance toward 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 of biofilm formation is believed to be a key aspect in the development of novel antipathogenic 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 antipathogenic 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 cyclic di-GMP level reporters by transcriptionally fusing the cyclic di-GMP-responsive cdrA promoter to genes encoding green fluorescent protein. We show that the reporter constructs give a fluorescent readout 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 that biofilm formation is 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 antipathogenic compounds targeting cyclic di-GMP signaling, as well as for use in research aiming at understanding the biofilm biology of P. aeruginosa.
Language: | English |
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Publisher: | American Society for Microbiology |
Year: | 2012 |
Pages: | 5060-5069 |
ISSN: | 10985336 and 00992240 |
Types: | Journal article |
DOI: | 10.1128/AEM.00414-12 |
ORCIDs: | 0000-0002-6218-6774 , 0000-0001-5004-8609 and 0000-0002-9751-474X |