Azithromycin Blocks Quorum Sensing and Alginate Polymer Formation and Increases the Sensitivity to Serum and Stationary-Growth-Phase Killing ofPseudomonas aeruginosaand Attenuates ChronicP. aeruginosaLung Infection inCftr^−/−Mice

Abstract

The consequences of O-acetylated alginate-producing Pseudomonas aeruginosa biofilms in the lungs of chronically infected cystic fibrosis (CF) patients are tolerance to both antibiotic treatments and effects on the innate and the adaptive defense mechanisms. In clinical trials, azithromycin (AZM) has been shown to improve the lung function of CF patients. The present study was conducted in accordance with previous in vitro studies suggesting that the effect of AZM may be the inhibition of alginate production, blockage of quorum sensing (QS), and increased sensitivity to hydrogen peroxide and the complement system. Moreover, we show that AZM may affect the polymerization of P. aeruginosa alginate by the incomplete precipitation of polymerized alginate and high levels of readily dialyzable uronic acids. In addition, we find that mucoid bacteria in the stationary growth phase became sensitive to AZM, whereas cells in the exponential phase did not. Interestingly, AZM-treated P. aeruginosa lasI mutants appeared to be particularly resistant to serum, whereas bacteria with a functional QS system did not. We show in a CF mouse model of chronic P. aeruginosa lung infection that AZM treatment results in the suppression of QS-regulated virulence factors, significantly improves the clearance of P. aeruginosa alginate biofilms, and reduces the severity of the lung pathology compared to that in control mice. We conclude that AZM attenuates the virulence of P. aeruginosa, impairs its ability to form fully polymerized alginate biofilms, and increases its sensitivity to complement and stationary-phase killing, which may explain the clinical efficacy of AZM.Mucoid Pseudomonas aeruginosa plays an important role in chronic lung infections among patients with CF due to its ability to form alginate-producing biofilms, which enable the bacteria to resist treatments with antibiotics and which affect the actions of the cells of the immune system (23). Macrolides such as azithromycin (AZM) are normally used against grampositive bacteria, whereas many gram-negative rods, such as Pseudomonas spp., are inherently resistant to macrolides (56). However, several clinical studies from Japan (36, 39, 65) have highlighted the beneficial effects of different macrolides, including AZM, in the treatment of patients with diffuse panbronchiolitis, a disease which is very similar to CF. The first indication that AZM therapy could also benefit the lung function in CF patients was given by Jaffe et al. (27) and was subsequently confirmed by other investigators (9,16,58,74). The mechanisms of action macrolides are multiple and may include an anti-inflammatory effect (36, 40), inhibition of a key enzyme in the alginate synthesis pathway (47), modulation of the production of quorum-sensing (QS) bacterial virulence factors (33,44,69), and/or inhibition of protein synthesis after prolonged exposure (67, 68).In P. aeruginosa, the production of several virulence factors is regulated by cell-to-cell communication, based on the lasand rhl-encoded QS systems (6), w...

Azithromycin Reduces the Production of α-hemolysin and Biofilm Formation in Staphylococcus aureus

“…Macrolide antibiotics have been shown to act as QS inhibitors at subinhibitory concentrations. For example, erythromycin has been reported to suppress production of P. aeruginosa hemagglutinins, protease, hemolysin and AHL signals [14], and studies have shown that azithromycin at sub-MIC concentrations affects QS-regulated virulence genes and biofilm formation in vitro [15][16][17] and in in vivo models of disease [18]. In the present study, we demonstrate that sub-inhibitory concentrations of azithromycin could dose-dependently reduce the haemolytic activity and biofilm formation in S. aureus.…”

Exploiting Quorum Sensing To Confuse Bacterial Pathogens

“…In a cystic fibrosis mouse model (utilizing Cftr Ϫ/Ϫ mice [322]), treatment with AZM reduced the bacterial load in lungs of mice, and gene expression of the QSregulated lasB gene was downregulated in vivo (323). AZM treatment also downregulated production and polymerization of alginate, which, combined with decreasing QS responses, was a major contribution to enhanced sensitivity to H 2 O 2 and complement and loss of viability in the stationary phase of growth.…”

Infections associées aux biofilms

“…Une approche proposée pour lutter contre les biofilms formés par P. aeruginosa repose sur l'inhibition du quorum-sensing en utilisant des analogues structuraux des homosérine lactones, comme les furanones, qui réduisent le biofilm de P. aeruginosa, in vitro et in vivo [15,47]. L'utilisation de l'azithromycine, qui n'a qu'une activité faible contre P. aeruginosa, altère les signaux de communication de la bactérie in vitro et in vivo et permet une amélioration de la fonction respiratoire et une réduction du nombre d'exacerbations chez les patients atteints de mucoviscidose [48,49]. Néanmoins, il a été démontré que l'azithromycine pouvait avoir un effet immunologique direct chez l'homme dont les conséquences sont, pour l'instant, mal connues [50].…”