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Quorum-sensing antagonistic activities of azithromycin in Pseudomonas aeruginosa PAO1: a global approach.

ABSTRACT: The administration of macrolides such as azithromycin for chronic pulmonary infection of cystic fibrosis patients has been reported to be of benefit. Although the mechanisms of action remain obscure, anti-inflammatory effects as well as interference of the macrolide with Pseudomonas aeruginosa virulence factor production have been suggested to contribute to an improved clinical outcome. In this study we used a systematic approach and analyzed the impact of azithromycin on the global transcriptional pattern and the protein expression profile of P. aeruginosa PAO1 cultures versus those in untreated controls. The most remarkable result of this study is the finding that azithromycin exhibited extensive quorum-sensing antagonistic activities. In accordance with the inhibition of the quorum-sensing systems, virulence factor production was diminished and the oxidative stress response was impaired, whereas the type III secretion system was strongly induced. Moreover, P. aeruginosa motility was reduced, which probably accounts for the previously observed impaired biofilm formation capabilities of azithromycin-treated cultures. The interference of azithromycin with quorum-sensing-dependent virulence factor production, biofilm formation, and oxidative stress resistance in P. aeruginosa holds great promise for macrolide therapy in cystic fibrosis. Clearly quorum-sensing antagonist macrolides should be paid more attention in the management of chronic P. aeruginosa infections, and as quorum-sensing antagonists, macrolides might gain vital importance for more general application against chronic infections.

SUBMITTER: Nalca Y

PROVIDER: S-EPMC1472232 | biostudies-literature | 2006 May

REPOSITORIES: biostudies-literature

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Quorum-sensing antagonistic activities of azithromycin in Pseudomonas aeruginosa PAO1: a global approach.

Nalca Yusuf Y Jänsch Lothar L Bredenbruch Florian F Geffers Robert R Buer Jan J Häussler Susanne S

Antimicrobial agents and chemotherapy 20060501 5

The administration of macrolides such as azithromycin for chronic pulmonary infection of cystic fibrosis patients has been reported to be of benefit. Although the mechanisms of action remain obscure, anti-inflammatory effects as well as interference of the macrolide with Pseudomonas aeruginosa virulence factor production have been suggested to contribute to an improved clinical outcome. In this study we used a systematic approach and analyzed the impact of azithromycin on the global transcriptio ...[more]

PMID: 16641435

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Project description:Two clinical isolates of the opportunist pathogen Pseudomonas aeruginosa named PAO1 and PA14 are commonly studied in research laboratories. Despite the isolates being closely related, PA14 exhibits increased virulence compared to that of PAO1 in various models. To determine which players are responsible for the hypervirulence phenotype of the PA14 strain, we elected a transcriptomic approach through RNA sequencing. We found 2,029 genes that are differentially expressed between the two strains, including several genes that are involved with or regulated by quorum sensing (QS), known to control most of the virulence factors in P. aeruginosa Among them, we chose to focus our study on QslA, an antiactivator of QS whose expression was barely detectable in the PA14 strain according our data. We hypothesized that lack of expression of qslA in PA14 could be responsible for higher QS expression in the PA14 strain, possibly explaining its hypervirulence phenotype. After confirming that QslA protein was highly produced in PAO1 but not in the PA14 strain, we obtained evidence showing that a PAO1 deletion strain of qslA has faster QS gene expression kinetics than PA14. Moreover, known virulence factors activated by QS, such as (i) pyocyanin production, (ii) H2-T6SS (type VI secretion system) gene expression, and (iii) Xcp-T2SS (type II secretion system) machinery production and secretion, were all lower in PAO1 than in PA14, due to higher qslA expression. However, biofilm formation and cytotoxicity toward macrophages, although increased in PA14 compared to PAO1, were independent of QslA control. Together, our findings implicated differential qslA expression as a major determinant of virulence factor expression in P. aeruginosa strains PAO1 and PA14.IMPORTANCE Pseudomonas aeruginosa is an opportunistic pathogen responsible for acute nosocomial infections and chronic pulmonary infections. P. aeruginosa strain PA14 is known to be hypervirulent in different hosts. Despite several studies in the field, the underlining molecular mechanisms sustaining this phenotype remain enigmatic. Here we provide evidence that the PA14 strain has faster quorum sensing (QS) kinetics than the PAO1 strain, due to the lack of QslA expression, an antiactivator of QS. QS is a major regulator of virulence factors in P. aeruginosa; therefore, we propose that the hypervirulent phenotype of the PA14 strain is, at least partially, due to the lack of QslA expression. This mechanism could be of great importance, as it could be conserved among other P. aeruginosa isolates.

Project description:The Pseudomonas aeruginosa quorum sensing (QS) network plays a key role in the adaptation to environmental changes and the control of virulence factor production in this opportunistic human pathogen. Three interlinked QS systems, namely las, rhl, and pqs, are central to the production of pyocyanin, a phenazine virulence factor which is typically used as phenotypic marker for analysing QS. Pyocyanin production in P. aeruginosa is a complex process involving two almost identical operons termed phzA1B1C1D1E1F1G1 (phz1) and phzA2B2C2D2E2F2G2 (phz2), which drive the production of phenazine-1-carboxylic acid (PCA) which is further converted to pyocyanin by two modifying enzymes PhzM and PhzS. Due to the high sequence conservation between the phz1 and phz2 operons (nucleotide identity > 98%), analysis of their individual expression by RNA hybridization, qRT-PCR or transcriptomics is challenging. To overcome this difficulty, we utilized luminescence based promoter fusions of each phenazine operon to measure in planktonic cultures their transcriptional activity in P. aeruginosa PAO1-N genetic backgrounds impaired in different components of the las, rhl, and pqs QS systems, in the presence or absence of different QS signal molecules. Using this approach, we found that all three QS systems play a role in differentially regulating the phz1 and phz2 phenazine operons, thus uncovering a higher level of complexity to the QS regulation of PCA biosynthesis in P. aeruginosa than previously appreciated.ImportanceThe way the P. aeruginosa QS regulatory networks are intertwined creates a challenge when analysing the mechanisms governing specific QS-regulated traits. Multiple QS regulators and signals have been associated with the production of phenazine virulence factors. In this work we designed experiments where we dissected the contribution of specific QS switches using individual mutations and complementation strategies to gain further understanding of the specific roles of these QS elements in controlling expression of the two P. aeruginosa phenazine operons. Using this approach we have teased out which QS regulators have either indirect or direct effects on the regulation of the two phenazine biosynthetic operons. The data obtained highlight the sophistication of the QS cascade in P. aeruginosa and the challenges in analysing the control of phenazine secondary metabolites.

Dataset Information

Quorum-sensing antagonistic activities of azithromycin in Pseudomonas aeruginosa PAO1: a global approach.

Publications

Quorum-sensing antagonistic activities of azithromycin in Pseudomonas aeruginosa PAO1: a global approach.

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