Curcumin induced photodynamic therapy mediated suppression of quorum sensing pathway of Pseudomonas aeruginosa: An approach to inhibit biofilm in vitro

The opportunistic human pathogen Pseudomonas aeruginosa is responsible for a variety of acute infections and is a major cause of mortality in chronically infected cystic fibrosis patients. Due to increased resistance to antibiotics, new therapeutic strategies against P. aeruginosa are urgently needed. In this context, we aimed to develop a simple vertebrate animal model to rapidly assess in vivo drug efficacy against P. aeruginosa. Zebrafish are increasingly considered for modeling human infections caused by bacterial pathogens, which are commonly microinjected in embryos. In the present study, we established a novel protocol for zebrafish infection by P. aeruginosa based on bath immersion in 96-well plates of tail-injured embryos. The immersion method, followed by a 48-hour survey of embryo viability, was first validated to assess the virulence of P. aeruginosa wild-type PAO1 and a known attenuated mutant. We then validated its relevance for antipseudomonal drug testing by first using a clinically used antibiotic, ciprofloxacin. Secondly, we used a novel quorum sensing (QS) inhibitory molecule, N-(2-pyrimidyl)butanamide (C11), the activity of which had been validated in vitro but not previously tested in any animal model. A significant protective effect of C11 was observed on infected embryos, supporting the ability of C11 to attenuate in vivo P. aeruginosa pathogenicity. In conclusion, we present here a new and reliable method to compare the virulence of P. aeruginosa strains in vivo and to rapidly assess the efficacy of clinically relevant drugs against P. aeruginosa, including new antivirulence compounds.

Download Full-text

Antibiotic Resistance Evolution Is Contingent on the Quorum-Sensing Response in Pseudomonas aeruginosa

Molecular Biology and Evolution ◽

10.1093/molbev/msz144 ◽

2019 ◽

Vol 36 (10) ◽

pp. 2238-2251 ◽

Cited By ~ 9

Author(s):

Sara Hernando-Amado ◽

Fernando Sanz-García ◽

José Luis Martínez

Keyword(s):

Pseudomonas Aeruginosa ◽

Antibiotic Resistance ◽

Quorum Sensing ◽

Wild Type ◽

Resistance Mutations ◽

Epistatic Interactions ◽

Adaptive Mutations ◽

Resistance Evolution ◽

Evolutionary Trajectories

Abstract Different works have explored independently the evolution toward antibiotic resistance and the role of eco-adaptive mutations in the adaptation to a new habitat (as the infected host) of bacterial pathogens. However, knowledge about the connection between both processes is still limited. We address this issue by comparing the evolutionary trajectories toward antibiotic resistance of a Pseudomonas aeruginosa lasR defective mutant and its parental wild-type strain, when growing in presence of two ribosome-targeting antibiotics. Quorum-sensing lasR defective mutants are selected in P. aeruginosa populations causing chronic infections. Further, we observed they are also selected in vitro as a first adaptation for growing in culture medium. By using experimental evolution and whole-genome sequencing, we found that the evolutionary trajectories of P. aeruginosa in presence of these antibiotics are different in lasR defective and in wild-type backgrounds, both at the phenotypic and the genotypic levels. Recreation of a set of mutants in both genomic backgrounds (either wild type or lasR defective) allowed us to determine the existence of negative epistatic interactions between lasR and antibiotic resistance determinants. These epistatic interactions could lead to mutual contingency in the evolution of antibiotic resistance when P. aeruginosa colonizes a new habitat in presence of antibiotics. If lasR mutants are selected first, this would constraint antibiotic resistance evolution. Conversely, when resistance mutations (at least those studied in the present work) are selected, lasR mutants may not be selected in presence of antibiotics. These results underlie the importance of contingency and epistatic interactions in modulating antibiotic resistance evolution.

Download Full-text

Calpurnia aurea (Aiton) Benth Extracts Reduce Quorum Sensing Controlled Virulence Factors in Pseudomonas aeruginosa

Molecules ◽

10.3390/molecules25102283 ◽

2020 ◽

Vol 25 (10) ◽

pp. 2283

Author(s):

Sekelwa Cosa ◽

Jostina R. Rakoma ◽

Abdullahi A. Yusuf ◽

Thilivhali E. Tshikalange

Keyword(s):

Pseudomonas Aeruginosa ◽

Molecular Docking ◽

Quorum Sensing ◽

Virulence Factors ◽

Laser Scanning ◽

Laser Scanning Microscopy ◽

Confocal Laser ◽

Promising Alternative ◽

The Individual

Pseudomonas aeruginosa is the causative agent of several life-threatening human infections. Like many other pathogens, P. aeruginosa exhibits quorum sensing (QS) controlled virulence factors such as biofilm during disease progression, complicating treatment with conventional antibiotics. Thus, impeding the pathogen’s QS circuit appears as a promising alternative strategy to overcome pseudomonas infections. In the present study, Calpurnia aurea were evaluated for their antibacterial (minimum inhibitory concentrations (MIC)), anti-quorum sensing/antivirulence (AQS), and antibiofilm potential against P. aeruginosa. AQS and antivirulence (biofilm formation, swimming, and swarming motility) activities of plant extracts were evaluated against Chromobacterium violaceum and P. aeruginosa, respectively. The in vitro AQS potential of the individual compounds were validated using in silico molecular docking. Acetone and ethanolic extracts of C. aurea showed MIC at 1.56 mg/mL. The quantitative violacein inhibition (AQS) assay showed ethyl acetate extracts as the most potent at a concentration of 1 mg/mL. GCMS analysis of C. aurea revealed 17 compounds; four (pentadecanol, dimethyl terephthalate, terephthalic acid, and methyl mannose) showed potential AQS through molecular docking against the CviR protein of C. violaceum. Biofilm of P. aeruginosa was significantly inhibited by ≥60% using 1-mg/mL extract of C. aurea. Confocal laser scanning microscopy correlated the findings of crystal violet assay with the extracts significantly altering the swimming motility. C. aurea extracts reduced the virulence of pseudomonas, albeit in a strain- and extract-specific manner, showing their suitability for the identification of lead compounds with QS inhibitory potential for the control of P. aeruginosa infections.

Download Full-text

Function of the bacteriophytochrome BphP in the RpoS/Las quorum-sensing network of Pseudomonas aeruginosa

Microbiology ◽

10.1099/mic.0.049007-0 ◽

2011 ◽

Vol 157 (6) ◽

pp. 1651-1664 ◽

Cited By ~ 15

Author(s):

Katalin Barkovits ◽

Britta Schubert ◽

Sabrina Heine ◽

Maurice Scheer ◽

Nicole Frankenberg-Dinkel

Keyword(s):

Pseudomonas Aeruginosa ◽

Quorum Sensing ◽

Response Regulator ◽

Red Light ◽

Regulatory System ◽

Global Gene Expression ◽

Opportunistic Pathogen ◽

Pleiotropic Effect ◽

Genes Encoding

The bacterial phytochrome of Pseudomonas aeruginosa (PaBphP) is an in vitro-active red/far-red light sensor histidine kinase of a two-component regulatory system. Despite solid biochemical data, its function in this heterotrophic, opportunistic pathogen is still unknown. Previous studies established that the genes encoding the two necessary phytochrome components BphO, a chromophore-producing haem oxygenase, and BphP, the apo-phytochrome, are co-transcribed in a bicistronic operon. Transcription has been shown to be induced in the stationary phase and to be dependent on the alternative sigma factor RpoS. Here we show an additional regulation of bphP expression through the quorum-sensing (QS) regulator LasR. This regulation is also reflected in a combination of expression profile experiments and proteome analyses of wild-type and phytochrome-deficient strains. While PaBphP has a pleiotropic effect on global gene expression, 66 % of the downregulated genes in the phytochrome mutant display a link to the Las QS system. Most of these genes seem to be indirectly regulated by LasR through BphP and the unknown response regulator BphR. A model of phytochrome function within the Las QS network is presented.

Download Full-text

Pseudomonas aeruginosa tolerance to tobramycin, hydrogen peroxide and polymorphonuclear leukocytes is quorum-sensing dependent

Microbiology ◽

10.1099/mic.0.27463-0 ◽

2005 ◽

Vol 151 (2) ◽

pp. 373-383 ◽

Cited By ~ 312

Author(s):

Thomas Bjarnsholt ◽

Peter Østrup Jensen ◽

Mette Burmølle ◽

Morten Hentzer ◽

Janus A. J. Haagensen ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Quorum Sensing ◽

Polymorphonuclear Leukocytes ◽

Present Report ◽

Cell Communication ◽

A Cell ◽

Opportunistic Human Pathogen ◽

Conventional Antibiotics

The opportunistic human pathogen Pseudomonas aeruginosa is the predominant micro-organism of chronic lung infections in cystic fibrosis (CF) patients. P. aeruginosa colonizes the CF lungs by forming biofilm structures in the alveoli. In the biofilm mode of growth the bacteria are highly tolerant to otherwise lethal doses of antibiotics and are protected from bactericidal activity of polymorphonuclear leukocytes (PMNs). P. aeruginosa controls the expression of many of its virulence factors by means of a cell–cell communication system termed quorum sensing (QS). In the present report it is demonstrated that biofilm bacteria in which QS is blocked either by mutation or by administration of QS inhibitory drugs are sensitive to treatment with tobramycin and H2O2, and are readily phagocytosed by PMNs, in contrast to bacteria with functional QS systems. In contrast to the wild-type, QS-deficient biofilms led to an immediate respiratory-burst activation of the PMNs in vitro. In vivo QS-deficient mutants provoked a higher degree of inflammation. It is suggested that quorum signals and QS-inhibitory drugs play direct and opposite roles in this process. Consequently, the faster and highly efficient clearance of QS-deficient bacteria in vivo is probably a two-sided phenomenon: down regulation of virulence and activation of the innate immune system. These data also suggest that a combination of the action of PMNs and QS inhibitors along with conventional antibiotics would eliminate the biofilm-forming bacteria before a chronic infection is established.

Download Full-text

Sulfane Sulfur Regulates LasR-Mediated Quorum Sensing and Virulence in Pseudomonas aeruginosa PAO1

Antioxidants ◽

10.3390/antiox10091498 ◽

2021 ◽

Vol 10 (9) ◽

pp. 1498

Author(s):

Guanhua Xuan ◽

Chuanjuan Lü ◽

Huangwei Xu ◽

Kai Li ◽

Huaiwei Liu ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Hydrogen Sulfide ◽

Quorum Sensing ◽

Sulfide Oxidation ◽

Pseudomonas Aeruginosa Pao1 ◽

Sulfane Sulfur ◽

Target Dna ◽

Production Of Hydrogen

Sulfane sulfur, such as inorganic and organic polysulfide (HSn− and RSn−, n > 2), is a common cellular component, produced either from hydrogen sulfide oxidation or cysteine metabolism. In Pseudomonas aeruginosa PAO1, LasR is a quorum sensing master regulator. After binding its autoinducer, LasR binds to its target DNA to activate the transcription of a suite of genes, including virulence factors. Herein, we report that the production of hydrogen sulfide and sulfane sulfur were positively correlated in P. aeruginosa PAO1, and sulfane sulfur was able to modify LasR, which generated Cys188 persulfide and trisulfide and produced a pentasulfur link between Cys201 and Cys203. The modifications did not affect LasR binding to its target DNA site, but made it several-fold more effective than unmodified LasR in activating transcription in both in vitro and in vivo assays. On the contrary, H2O2 inactivates LasR via producing a disulfide bond between Cys201 and Cys203. P. aeruginosa PAO1 had a high cellular sulfane sulfur and high LasR activity in the mid log phase and early stationary phase, but a low sulfane sulfur and low LasR activity in the declination phase. Thus, sulfane sulfur is a new signaling factor in the bacterium, adding another level of control over LasR-mediated quorum sensing and turning down the activity in old cells.

Download Full-text

Quorum sensing and virulence of Pseudomonas aeruginosa during urinary tract infections

The Journal of Infection in Developing Countries ◽

10.3855/jidc.2543 ◽

2012 ◽

Vol 6 (06) ◽

pp. 501-507 ◽

Cited By ~ 24

Author(s):

Sezgi Senturk ◽

Seyhan Ulusoy ◽

Gulgun Bosgelmez-Tinaz ◽

Aysegul Yagci

Keyword(s):

Pseudomonas Aeruginosa ◽

Urinary Tract ◽

Quorum Sensing ◽

Virulence Factors ◽

Urinary Tract Infections ◽

Clinical Isolates ◽

Pcr Analysis ◽

Signal Molecules ◽

Tract Infections

Introduction: In the opportunistic pathogen Pseudomonas aeruginosa, the production of several virulence factors depends on quorum sensing (QS) involving N-acylhomoserine lactone signal molecules. In vitro studies have suggested that the QS system is crucial in the pathogenesis of P. aeruginosa. However, it is unclear whether QS systems of P. aeruginosa play the same role during infections. Methodology: In this study, to explore the contribution of QS systems to the pathogenesis of P. aeruginosa during urinary tract infections, we collected 82 clinical isolates. Detection of N-acyl-homoserine lactones (C12-HSL and C4-HSL) was performed on agar plates employing biosensor strains C. violaceum. Elastase and biofilm production were determined spectrophotometrically. QS genes were detected by PCR and subsequently underwent sequencing. Results and conclusion: Six isolates were found to be negative in the production of both C12-HSL and C4-HSL and all virulence factors tested. PCR analysis of these isolates revealed that four isolates contained all four QS genes while one isolate was negative for lasR gene, and one isolate negative for lasI, lasR and rhlR genes. Sequence analyses of these isolates showed that the lasR, lasI, rhlR and rhlI genes had point mutations. The combination of these mutations probably explains their C12-HSL, C4-HSL and virulence factor deficiencies. Results of this study suggest that QS deficient clinical isolates occur and are still capable of causing clinical infections in humans.

Download Full-text