Dataset Information

Induction of attachment-independent biofilm formation and repression of Hfq expression by low-fluid-shear culture of Staphylococcus aureus.

ABSTRACT: The opportunistic pathogen Staphylococcus aureus encounters a wide variety of fluid shear levels within the human host, and they may play a key role in dictating whether this organism adopts a commensal interaction with the host or transitions to cause disease. By using rotating-wall vessel bioreactors to create a physiologically relevant, low-fluid-shear environment, S. aureus was evaluated for cellular responses that could impact its colonization and virulence. S. aureus cells grown in a low-fluid-shear environment initiated a novel attachment-independent biofilm phenotype and were completely encased in extracellular polymeric substances. Compared to controls, low-shear-cultured cells displayed slower growth and repressed virulence characteristics, including decreased carotenoid production, increased susceptibility to oxidative stress, and reduced survival in whole blood. Transcriptional whole-genome microarray profiling suggested alterations in metabolic pathways. Further genetic expression analysis revealed downregulation of the RNA chaperone Hfq, which parallels low-fluid-shear responses of certain Gram-negative organisms. This is the first study to report an Hfq association with fluid shear in a Gram-positive organism, suggesting an evolutionarily conserved response to fluid shear among structurally diverse prokaryotes. Collectively, our results suggest S. aureus responds to a low-fluid-shear environment by initiating a biofilm/colonization phenotype with diminished virulence characteristics, which could lead to insight into key factors influencing the divergence between infection and colonization during the initial host-pathogen interaction.

SUBMITTER: Castro SL

PROVIDER: S-EPMC3187170 | biostudies-literature | 2011 Sep

REPOSITORIES: biostudies-literature

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Induction of attachment-independent biofilm formation and repression of Hfq expression by low-fluid-shear culture of Staphylococcus aureus.

Castro Sarah L SL Nelman-Gonzalez Mayra M Nickerson Cheryl A CA Ott C Mark CM

Applied and environmental microbiology 20110729 18

The opportunistic pathogen Staphylococcus aureus encounters a wide variety of fluid shear levels within the human host, and they may play a key role in dictating whether this organism adopts a commensal interaction with the host or transitions to cause disease. By using rotating-wall vessel bioreactors to create a physiologically relevant, low-fluid-shear environment, S. aureus was evaluated for cellular responses that could impact its colonization and virulence. S. aureus cells grown in a low-f ...[more]

PMID: 21803898

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Project description:The opportunistic pathogen, Staphylococcus aureus, encounters a wide variety of fluid shear levels within the human host, which may play a key role in dictating whether this organism adopts a commensal interaction with the host or transitions to cause disease. Using rotating-wall vessel bioreactors to create a physiologically-relevant, low fluid shear environment, S. aureus was evaluated for cellular responses that could impact its colonization and virulence. S. aureus cells grown in a low fluid shear environment initiated a novel attachment-independent biofilm phenotype and were completely encased in extracellular polymeric substances. Compared to controls, low-shear cultured cells displayed slower growth and repressed virulence characteristics, including decreased carotenoid production, increased susceptibility to oxidative stress, and reduced survival in whole blood. Transcriptional whole genome microarray profiling suggested alterations in metabolic pathways. Further genetic expression analysis revealed the down-regulation of the RNA chaperone Hfq, which parallels low fluid shear responses of certain Gram negative organisms. This is the first study to report an Hfq association to fluid shear in a Gram positive organism, suggesting an evolutionarily conserved response to fluid shear among structurally diverse prokaryotes. Collectively, our results suggest S. aureus responds to a low fluid shear environment by initiating a biofilm/colonization phenotype with diminished virulence characteristics, which could lead to insight into key factors influencing the divergence between infection and colonization during initial host pathogen interaction. Genetic expression profiles of Staphylococcus aureus cultured under low fluid shear conditions was compared to control cultures of S. aureus which was cultured in identical hardware in an orientation disrupting the low fluid shear effect. Samples from the same date of culture were compared (control 21:low 21 and control 30: low 30). S. aureus was cultured for 20 hours in either the low fluid shear or control orientated rotating wall vessel (RWV) bioreactor at which point the cells were removed and RNA extracted. At 20 hours, both cultures were in the same stage of growth (stationary phase) and at this point phenotypic differences between control and low fluid shear cultures were noted.

Project description:Periprosthetic joint infections (PJIs) are a devastating complication that occurs in 2% of patients following joint replacement. These infections are costly and difficult to treat, often requiring multiple corrective surgeries and prolonged antimicrobial treatments. The Gram-positive bacterium Staphylococcus aureus is one of the most common causes of PJIs, and it is often resistant to a number of commonly used antimicrobials. This tolerance can be partially attributed to the ability of S. aureus to form biofilms. Biofilms associated with the surface of indwelling medical devices have been observed on components removed during chronic infection, however, the development and localization of biofilms during PJIs remains unclear. Prior studies have demonstrated that synovial fluid, in the joint cavity, promotes the development of bacterial aggregates with many biofilm-like properties, including antibiotic resistance. We anticipate these aggregates have an important role in biofilm formation and antibiotic tolerance during PJIs. Therefore, we sought to determine specifically how synovial fluid promotes aggregate formation and the impact of this process on surface attachment. Using flow cytometry and microscopy, we quantified the aggregation of various clinical S. aureus strains following exposure to purified synovial fluid components. We determined that fibrinogen and fibronectin promoted bacterial aggregation, while cell free DNA, serum albumin, and hyaluronic acid had minimal effect. To determine how synovial fluid mediated aggregation affects surface attachment, we utilized microscopy to measure bacterial attachment. Surprisingly, we found that synovial fluid significantly impeded bacterial surface attachment to a variety of materials. We conclude from this study that fibrinogen and fibronectin in synovial fluid have a crucial role in promoting bacterial aggregation and inhibiting surface adhesion during PJI. Collectively, we propose that synovial fluid may have conflicting protective roles for the host by preventing adhesion to surfaces, but by promoting bacterial aggregation is also contributing to the development of antibiotic tolerance.

Project description:The viscoelasticity of a biofilm's EPS (extracellular polymeric substance) matrix conveys protection against mechanical challenges, but adaptive responses of biofilm inhabitants to produce EPS are not well known. Here, we compare the responses of a biofilm of an EPS-producing (ATCC 12600) and a non-EPS producing (5298) Staphylococcus aureus strain to fluid shear and mechanical challenge. Confocal laser scanning microscopy confirmed absence of calcofluor-white-stainable EPS in biofilms of S. aureus 5298. Attenuated total reflection Fourier transform infrared (ATR-FTIR) spectroscopy combined with tribometry indicated that polysaccharide production per bacterium in the initial adhering layer was higher during growth at high shear than at low shear and that this increased EPS production extended to entire biofilms, as indicated by tribometrically measured coefficients of friction (CoF). CoF of biofilms grown under high fluid shear were higher than those when grown under low shear, likely due to wash-off polysaccharides. Measurement of a biofilm's CoF implies application of mechanical pressure that yielded an immediate increase in the polysaccharide band area of S. aureus ATCC 12600 biofilms due to their compression. Compression decreased after relief of pressure to the level observed prior to mechanical pressure. For biofilms grown under high shear, this coincided with a higher percent whiteness in optical coherence tomography-images indicative of water outflow, returning back into the biofilm during stress relaxation. Biofilms grown under low shear, however, were stimulated during tribometry to produce EPS, also after relief of stress. Knowledge of factors that govern EPS production and water flow in biofilms will allow better control of biofilms under mechanical challenge and better understanding of the barrier properties of biofilms against antimicrobial penetration.IMPORTANCE Adaptive responses of biofilm inhabitants in nature to environmental challenges such as fluid shear and mechanical pressure often involve EPS production with the aim of protecting biofilm inhabitants. EPS can assist biofilm bacteria in remaining attached or can impede antimicrobial penetration. The TriboChemist is a recently introduced instrument, allowing the study of initially adhering bacteria to a germanium crystal using ATR-FTIR spectroscopy, while simultaneously allowing measurement of the coefficient of friction of a biofilm, which serves as an indicator of the EPS content of a biofilm. EPS production can be stimulated by both fluid shear during growth and mechanical pressure, while increased EPS production can continue after pressure relaxation of the biofilm. Since EPS is pivotal in the protection of biofilm inhabitants against mechanical and chemical challenges, knowledge of the factors that make biofilm inhabitants decide to produce EPS, as provided in this study, is important for the development of biofilm control measures.

Dataset Information

Induction of attachment-independent biofilm formation and repression of Hfq expression by low-fluid-shear culture of Staphylococcus aureus.

Publications

Induction of attachment-independent biofilm formation and repression of Hfq expression by low-fluid-shear culture of Staphylococcus aureus.

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