In vitro and in vivo generation and characterization of Pseudomonas aeruginosa biofilm–dispersed cells via c-di-GMP manipulation

Chua, Song Lin; Hultqvist, Louise Dahl; Yuan, Mingjun; Rybtke, Morten; Nielsen, Thomas E.; Givskov, Michael; Tolker‐Nielsen, Tim; Yang, Liang

doi:10.1038/nprot.2015.067

Cited by 67 publications

(61 citation statements)

References 56 publications

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“…In total, 200 µl of cell culture was then transferred into each well of a 96-well plate (triplicates). As previously described [13], the OD 600 and pyoverdine fluorescence (excitation 400 nm/emission 460 nm) were measured using the Tecan microplate reader, and the relative fluorescence intensity count calculated by dividing pyoverdine fluorescence values by OD 600 values. Three independent experiments were performed in triplicate, one-way ANOVA and Student's t -tests were used to determine statistical significance and the results were shown as the mean ± s.d.…”

Section: Methodsmentioning

confidence: 99%

Reactive oxygen species drive evolution of pro-biofilm variants in pathogens by modulating cyclic-di-GMP levels

et al. 2016

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The host immune system offers a hostile environment with antimicrobials and reactive oxygen species (ROS) that are detrimental to bacterial pathogens, forcing them to adapt and evolve for survival. However, the contribution of oxidative stress to pathogen evolution remains elusive. Using an experimental evolution strategy, we show that exposure of the opportunistic pathogen Pseudomonas aeruginosa to sub-lethal hydrogen peroxide (H2O2) levels over 120 generations led to the emergence of pro-biofilm rough small colony variants (RSCVs), which could be abrogated by l-glutathione antioxidants. Comparative genomic analysis of the RSCVs revealed that mutations in the wspF gene, which encodes for a repressor of WspR diguanylate cyclase (DGC), were responsible for increased intracellular cyclic-di-GMP content and production of Psl exopolysaccharide. Psl provides the first line of defence against ROS and macrophages, ensuring the survival fitness of RSCVs over wild-type P. aeruginosa. Our study demonstrated that ROS is an essential driving force for the selection of pro-biofilm forming pathogenic variants. Understanding the fundamental mechanism of these genotypic and phenotypic adaptations will improve treatment strategies for combating chronic infections.

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Section: Methodsmentioning

confidence: 99%

Reactive oxygen species drive evolution of pro-biofilm variants in pathogens by modulating cyclic-di-GMP levels

et al. 2016

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“…By monitoring the expression level of biomarkers specific to the P. aeruginosa dispersal state ( pvdA and cdrA ), Chua et al . showed that the dispersal state can be maintained in harsh nutritional conditions, but that addition of fresh medium induces reversion to the planktonic mode of growth within 2 hours (Chua et al ., ).…”

Section: Methodological Guidelines For Characterization Of Biofilm DImentioning

confidence: 99%

Biofilm dispersal: multiple elaborate strategies for dissemination of bacteria with unique properties

Guilhen

Forestier

Balestrino

2017

Molecular Microbiology

182

143

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Summary In most environments, microorganisms evolve in a sessile mode of growth, designated as biofilm, which is characterized by cells embedded in a self‐produced extracellular matrix. Although a biofilm is commonly described as a “cozy house” where resident bacteria are protected from aggression, bacteria are able to break their biofilm bonds and escape to colonize new environments. This regulated process is observed in a wide variety of species; it is referred to as biofilm dispersal, and is triggered in response to various environmental and biological signals. The first part of this review reports the main regulatory mechanisms and effectors involved in biofilm dispersal. There is some evidence that dispersal is a necessary step between the persistence of bacteria inside biofilm and their dissemination. In the second part, an overview of the main methods used so far to study the dispersal process and to harvest dispersed bacteria was provided. Then focus was on the properties of the biofilm‐dispersed bacteria and the fundamental role of the dispersal process in pathogen dissemination within a host organism. In light of the current body of knowledge, it was suggested that dispersal acts as a potent means of disseminating bacteria with enhanced colonization properties in the surrounding environment.

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“…In P. aeruginosa, dispersion upon exposure to NO and elevated nutrient concentrations has been linked to the reduction of the cellular c-di-GMP levels, requiring the PDEs DipA, NbdA, and RbdA [40,51,52,59]. The finding of dispersed cells being characterized by reduced c-di-GMP levels has led to the hypothesis that dispersed cells can be generated by reducing the intracellular c-di-GMP content through modulation of PDEs [60]. In P. putida and P. fluorescens , the large adhesive outer-membrane protein LapA mediates attachment to surfaces and matrix components and is required for the transition from reversible to irreversible attachment [41,61-63].…”

Section: A Tale Of C-di-gmp Modulationmentioning

confidence: 99%

Escaping the biofilm in more than one way: desorption, detachment or dispersion

Petrova

Sauer

2016

Current Opinion in Microbiology

221

163

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Biofilm bacteria have developed escape strategies to avoid stresses associated with biofilm growth, respond to changing environmental conditions, and disseminate to new locations. An ever-expanding body of research suggests that cellular release from biofilms is distinct from a simple reversal of attachment and reversion to a planktonic mode of growth, with biofilm dispersion involving sensing of specific cues, regulatory signal transduction, and consequent physiological alterations. However, dispersion is only one of many ways to escape the biofilm mode of growth. The present review is aimed at distinguishing this active and regulated process of dispersion from the passive processes of desorption and detachment by highlighting the regulatory processes and distinct phenotypes specific to dispersed cells.

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In vitro and in vivo generation and characterization of Pseudomonas aeruginosa biofilm–dispersed cells via c-di-GMP manipulation

Cited by 67 publications

References 56 publications

Reactive oxygen species drive evolution of pro-biofilm variants in pathogens by modulating cyclic-di-GMP levels

Reactive oxygen species drive evolution of pro-biofilm variants in pathogens by modulating cyclic-di-GMP levels

Biofilm dispersal: multiple elaborate strategies for dissemination of bacteria with unique properties

Escaping the biofilm in more than one way: desorption, detachment or dispersion

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