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Biomedical subjects

P Stoodley

Publications and source records attributed to P Stoodley.

At least 19 recordsLinked to original sources
PubMed2007

Cardiac fibroblasts influence cardiomyocyte phenotype in vitro.

Cardiac fibroblasts impact myocardial development and remodeling through intercellular contact with cardiomyocytes, but less is known about noncontact, profibrotic signals whereby fibroblasts alter cardiomyocyte behavior. Fibroblasts and cardiomyocytes were harvested from newborn rat ventricles and separated by serial digestion and gradient centrifugation. Cardiomyocytes were cultured in 1) standard medium, 2) standard medium diluted 1:1 with PBS, or 3) standard medium diluted 1:1 with medium conditioned > or =72 h by cardiac fibroblasts. Serum concentrations were held constant under all media conditions, and complete medium exchanges were performed daily. Cardiomyocytes began contracting within 24 h at clonal or mass densities with <5% of cells expressing vimentin. Immunocytochemical analysis revealed progressive expression of alpha-smooth muscle actin in cardiomyocytes after 24 h in all conditions. Only cardiomyocytes in fibroblast-conditioned medium stopped contracting by 72 h. There was a significant, sustained increase in vimentin expression specific to these cultures (means +/- SD: conditioned 46.3 +/- 6.0 vs. control 5.3 +/- 2.9%, P < 0.00025) typically with cardiac myosin heavy chain coexpression. Proteomics assays revealed 10 cytokines (VEGF, GRO/KC, monocyte chemoattractant protein-1, leptin, macrophage inflammatory protein-1alpha, IL-6, IL-10, IL-12p70, IL-17, and tumor necrosis factor-alpha) at or below detection levels in unconditioned medium that were significantly elevated in fibroblast-conditioned medium. Latent transforming growth factor-beta and RANTES were present in unconditioned medium but rose to higher levels in conditioned medium. Only granulocyte-macrophage colony-stimulating factor was present above threshold levels in standard medium but decreased with fibroblast conditioning. These data indicated that under the influence of fibroblast-conditioned medium, cardiomyocytes exhibited marked hypertrophy, diminished contractile capacity, and phenotype plasticity distinct from the dedifferentiation program present under standard culture conditions.

Actins↗
PubMed2005

Can laboratory reference strains mirror "real-world" pathogenesis?

The extraordinary plasticity of bacterial genomes raises concerns about the adequacy of laboratory-adapted reference strains for the study of "real-world" pathogenesis. Some laboratory strains have been sub-cultured for decades since their first isolation and might have lost important pathophysiological characteristics. Evidence is presented that bacteria rapidly adapt to in vitro conditions. Genomic differences between laboratory reference strains and corresponding low-passage clinical isolates are reviewed. It appears that no bacterial strain can truly represent its species. For DNA microarray and proteomic studies, this limitation might be overcome by the summation of individual genomes to produce a species-specific virtual supragenome.

Escherichia coli K12↗
PubMed2005

Survival strategies of infectious biofilms.

Modern medicine is facing the spread of biofilm-related infections. Bacterial biofilms are difficult to detect in routine diagnostics and are inherently tolerant to host defenses and antibiotic therapies. In addition, biofilms facilitate the spread of antibiotic resistance by promoting horizontal gene transfer. We review current concepts of biofilm tolerance with special emphasis on the role of the biofilm matrix and the physiology of biofilm-embedded cells. The heterogeneity in metabolic and reproductive activity within a biofilm correlates with a non-uniform susceptibility of enclosed bacteria. Recent studies have documented similar heterogeneity in planktonic cultures. Nutritional starvation and high cell density, two key characteristics of biofilm physiology, also mediate antimicrobial tolerance in stationary-phase planktonic cultures. Advances in characterizing the role of stress response genes, quorum sensing and phase variation in stationary-phase planktonic cultures have shed new light on tolerance mechanisms within biofilm communities.

Adaptation, Physiological↗
PubMed2005

Flowing biofilms as a transport mechanism for biomass through porous media under laminar and turbulent conditions in a laboratory reactor system.

Fluid flow has been shown to be important in influencing biofilm morphology and causing biofilms to flow over surfaces in flow cell experiments. However, it is not known whether similar effects may occur in porous media. Generally, it is assumed that the primary transport mechanism for biomass in porous media is through convection, as suspended particulates (cells and flocs) carried by fluid flowing through the interstices. However, the flow of biofilms over the surfaces of soils and sediment particles, may represent an important flux of biomass, and subsequently affect both biological activity and permeability. Mixed species bacterial biofilms were grown in glass flow cells packed with 1 mm diameter glass beads, under laminar or turbulent flow (porous media Reynolds number = 20 and 200 respectively). The morphology and dynamic behavior reflected those of biofilms grown in the open flow cells. The laminar biofilm was relatively uniform and after 23 d had inundated the majority of the pore spaces. Under turbulent flow the biofilm accumulated primarily in protected regions at contact points between the beads and formed streamers that trailed from the leeward face. Both biofilms caused a 2 to 3-fold increase in friction factor and in both cases there were sudden reductions in friction factor followed by rapid recovery, suggesting periodic sloughing and regrowth events. Time-lapse microscopy revealed that under both laminar and turbulent conditions biofilms flowed over the surface of the porous media. In some instances ripple structures formed. The velocity of biofilm flow was on the order of 10 mum h(-1) in the turbulent flow cell and 1.0 mum h(-1) in the laminar flow cell.

Biofilms↗
PubMed2005

Phenotypic differentiation and seeding dispersal in non-mucoid and mucoid Pseudomonas aeruginosa biofilms.

There is growing evidence that Pseudomonas aeruginosa biofilms exhibit a multicellular developmental life cycle analogous to that of the myxobacteria. In non-mucoid PAO1 biofilms cultured in glass flow cells the phenotypic differentiation of microcolonies into a motile phenotype in the interior of the microcolony and a non-motile surrounding 'wall phenotype' are described. After differentiation the interior cells coordinately evacuated the microcolony from local break out points and spread over the wall of the flow cell, suggesting that the specialized microcolonies were analogous to crude fruiting bodies. A microcolony diameter of approximately 80 microm was required for differentiation, suggesting that regulation was related to cell density and mass transfer conditions. This phenomenon was termed 'seeding dispersal' to differentiate it from 'erosion' which is the passive removal of single cells by fluid shear. Using the flow cell culturing method, in which reproducible seeding phenotype in PAO1 wild-type was demonstrated, the effects of quorum sensing (QS) and rhamnolipid production (factors previously identified as important in determining biofilm structure) on seeding dispersal using knockout mutants isogenic with PAO1 was investigated. Rhamnolipid (rhlA) was not required for seeding dispersal but las/rhl QS (PAO1-JP2) was, in our system. To assess the clinical relevance of these data, mucoid P. aeruginosa cystic fibrosis isolate FRD1 was also investigated and was seeding-dispersal-negative.

Biofilms↗
PubMed2005

Ultrasonically controlled release of ciprofloxacin from self-assembled coatings on poly(2-hydroxyethyl methacrylate) hydrogels for Pseudomonas aeruginosa biofilm prevention.

Indwelling prostheses and subcutaneous delivery devices are now routinely and indispensably employed in medical practice. However, these same devices often provide a highly suitable surface for bacterial adhesion and colonization, resulting in the formation of complex, differentiated, and structured communities known as biofilms. The University of Washington Engineered Biomaterials group has developed a novel drug delivery polymer matrix consisting of a poly(2-hydroxyethyl methacrylate) hydrogel coated with ordered methylene chains that form an ultrasound-responsive coating. This system was able to retain the drug ciprofloxacin inside the polymer in the absence of ultrasound but showed significant drug release when low-intensity ultrasound was applied. To assess the potential of this controlled drug delivery system for the targeting of infectious biofilms, we monitored the accumulation of Pseudomonas aeruginosa biofilms grown on hydrogels with and without ciprofloxacin and with and without exposure to ultrasound (a 43-kHz ultrasonic bath for 20 min daily) in an in vitro flow cell study. Biofilm accumulation from confocal images was quantified and statistically compared by using COMSTAT biofilm analysis software. Biofilm accumulation on ciprofloxacin-loaded hydrogels with ultrasound-induced drug delivery was significantly reduced compared to the accumulation of biofilms grown in control experiments. The results of these studies may ultimately facilitate the future development of medical devices sensitive to external ultrasonic impulses and capable of treating or preventing biofilm growth via "on-demand" drug release.

Anti-Infective Agents↗
PubMed2004

Commonality of elastic relaxation times in biofilms.

Biofilms, sticky conglomerations of microorganisms and extracellular polymers, are among the Earth's most common life forms. One component for their survival is an ability to withstand external mechanical stress. Measurements indicate that biofilm elastic relaxation times are approximately the same (about 18 min) over a wide sample of biofilms though other material properties vary significantly. A possible survival significance of this time scale is that it is the shortest period over which a biofilm can mount a phenotypic response to transient mechanical stress.

Biofilms↗
PubMed2004

Usnic acid, a natural antimicrobial agent able to inhibit bacterial biofilm formation on polymer surfaces.

In modern medicine, artificial devices are used for repair or replacement of damaged parts of the body, delivery of drugs, and monitoring the status of critically ill patients. However, artificial surfaces are often susceptible to colonization by bacteria and fungi. Once microorganisms have adhered to the surface, they can form biofilms, resulting in highly resistant local or systemic infections. At this time, the evidence suggests that (+)-usnic acid, a secondary lichen metabolite, possesses antimicrobial activity against a number of planktonic gram-positive bacteria, including Staphylococcus aureus, Enterococcus faecalis, and Enterococcus faecium. Since lichens are surface-attached communities that produce antibiotics, including usnic acid, to protect themselves from colonization by other bacteria, we hypothesized that the mode of action of usnic acid may be utilized in the control of medical biofilms. We loaded (+)-usnic acid into modified polyurethane and quantitatively assessed the capacity of (+)-usnic acid to control biofilm formation by either S. aureus or Pseudomonas aeruginosa under laminar flow conditions by using image analysis. (+)-Usnic acid-loaded polymers did not inhibit the initial attachment of S. aureus cells, but killing the attached cells resulted in the inhibition of biofilm. Interestingly, although P. aeruginosa biofilms did form on the surface of (+)-usnic acid-loaded polymer, the morphology of the biofilm was altered, possibly indicating that (+)-usnic acid interfered with signaling pathways.

Anti-Bacterial Agents↗
PubMed2004

Detachment characteristics and oxacillin resistance of Staphyloccocus aureus biofilm emboli in an in vitro catheter infection model.

Catheter-related bloodstream infections due to Staphylococcus aureus are of increasing clinical importance. The pathophysiological steps leading to colonization and infection, however, are still incompletely defined. We observed growth and detachment of S. aureus biofilms in an in vitro catheter-infection model by using time-lapse microscopy. Biofilm emboli were characterized by their size and their susceptibility for oxacillin. Biofilm dispersal was found to be a dynamic process in which clumps of a wide range of diameters detach. Large detached clumps were highly tolerant to oxacillin compared with exponential-phase planktonic cultures. Interestingly, the degree of antibiotic tolerance in stationary-phase planktonic cultures was equal to that in the large clumps. The mechanical disruption of large clumps reduced the minimal bactericidal concentration (MBC) by more than 1,000 times. The MBC for whole biofilm effluent, consisting of particles with an average number of 20 bacteria was 3.5 times higher than the MBC for planktonic cultures. We conclude that the antibiotic resistance of detached biofilm particles depends on the embolus size and could be attributed to nutrient-limited stationary-phase physiology of cells within the clumps. We hypothesize that the detachment of multicellular clumps may explain the high rate of symptomatic metastatic infections seen with S. aureus.

Anti-Bacterial Agents↗
PubMed2003

Biofilm removal from silicone tubing: an assessment of the efficacy of dialysis machine decontamination procedures using an in vitro model.

The aim of this study was to assess the efficacy of 21 decontamination procedures, for the removal of a multispecies biofilm. Experiments were performed on five-day-old biofilms grown inside silicone tubing, using a reactor system that mimics a dialysis machine. The treatments were tested on 5 cm tubing samples. Effects of treatment were measured using direct microscopy following staining. Bacterial viability and endotoxin removal were determined using conventional microbiological methods following biofilm detachment by scraping. The 21 procedures were classified into four groups based on the amount of biofilm removed. The most effective treatment was an acid pre-treatment, followed by use of a concentrated bleach solution. Acid pre-treatment removes calcium and magnesium carbonate crystals that are always found in dialysis biofilms. Treatments performed at high temperature did not increase the efficacy of biofilm removal. Most treatments caused at least a 10(5)-fold reduction in bacterial viability with a few resulting in complete kill. Autoclaved and bleach-treated samples gave the best results for viability reduction, with both treatments providing an equally effective and complete kill. In addition, autoclaving led to a significant decrease in endotoxin level (removal of 99.99%).

Biofilms↗
PubMed2002

Viscoelastic fluid description of bacterial biofilm material properties.

A mathematical model describing the constitutive properties of biofilms is required for predicting biofilm deformation, failure, and detachment in response to mechanical forces. Laboratory observations indicate that biofilms are viscoelastic materials. Likewise, current knowledge of biofilm internal structure suggests modeling biofilms as associated polymer viscoelastic systems. Supporting experimental results and a system of viscoelastic fluid equations with a linear Jeffreys viscoelastic stress-strain law are presented here. This system of equations is based on elements of associated polymer physics and is also consistent with presented and previous experimental results. A number of predictions can be made. One particularly interesting result is the prediction of an elastic relaxation time on the order of a few minutes-biofilm disturbances on shorter time scales produce an elastic response, biofilm disturbances on longer time scales result in viscous flow, i.e., nonreversible biofilm deformation. Although not previously recognized, evidence of this phenomenon is in fact present in recent experimental results.

Biofilms↗
PubMed2002

Biofilms as complex differentiated communities.

Prokaryotic biofilms that predominate in a diverse range of ecosystems are often composed of highly structured multispecies communities. Within these communities metabolic activities are integrated, and developmental sequences, not unlike those of multicellular organisms, can be detected. These structural adaptations and interrelationships are made possible by the expression of sets of genes that result in phenotypes that differ profoundly from those of planktonically grown cells of the same species. Molecular and microscopic evidence suggest the existence of a succession of de facto biofilm phenotypes. We submit that complex cell-cell interactions within prokaryotic communities are an ancient characteristic, the development of which was facilitated by the localization of cells at surfaces. In addition to spatial localization, surfaces may have provided the protective niche in which attached cells could create a localized homeostatic environment. In a holistic sense both biofilm and planktonic phenotypes may be viewed as integrated components of prokaryote life.

Bacteria↗
PubMed2002

Biofilm material properties as related to shear-induced deformation and detachment phenomena.

Biofilms of various Pseudomonas aeruginosa strains were grown in glass flow cells under laminar and turbulent flows. By relating the physical deformation of biofilms to variations in fluid shear, we found that the biofilms were viscoelastic fluids which behaved like elastic solids over periods of a few seconds but like linear viscous fluids over longer times. These data can be explained using concepts of associated polymeric systems, suggesting that the extracellular polymeric slime matrix determines the cohesive strength. Biofilms grown under high shear tended to form filamentous streamers while those grown under low shear formed an isotropic pattern of mound-shaped microcolonies. In some cases, sustained creep and necking in response to elevated shear resulted in a time-dependent fracture failure of the "tail" of the streamer from the attached upstream "head." In addition to structural differences, our data suggest that biofilms grown under higher shear were more strongly attached and were cohesively stronger than those grown under lower shears.

Biofilms↗
PubMed2002

The influence of fluid shear on the structure and material properties of sulphate-reducing bacterial biofilms.

Biofilms of sulphate-reducing Desulfovibrio sp. EX265 were grown in square section glass capillary flow cells under a range of fluid flow velocities from 0.01 to 0.4 m/s (wall shear stress, tau(w), from 0.027 to 1.0 N/m(2)). In situ image analysis and confocal scanning laser microscopy revealed biofilm characteristics similar to those reported for aerobic biofilms. Biofilms in both flow cells were patchy and consisted of cell clusters separated by voids. Length-to-width ratio measurements (l(c):w(c)) of biofilm clusters demonstrated the formation of more "streamlined" biofilm clusters (l(c):w(c)=3.03) at high-flow velocity (Reynolds number, Re, 1200), whereas at low-flow velocity (Re 120), the l(c):w(c) of the clusters was approximately 1 (l(c):w(c) of 1 indicates no elongation in the flow direction). Cell clusters grown under high flow were more rigid and had a higher yield point (the point at which the biofilm began to flow like a fluid) than those established at low flow and some biofilm cell aggregates were able to relocate within a cluster, by travelling in the direction of flow, before attaching more firmly downstream.

Biofilms↗
PubMed2002

Influence of hydrodynamics and cell signaling on the structure and behavior of Pseudomonas aeruginosa biofilms.

Biofilms were grown from wild-type (WT) Pseudomonas aeruginosa PAO1 and the cell signaling lasI mutant PAO1-JP1 under laminar and turbulent flows to investigate the relative contributions of hydrodynamics and cell signaling for biofilm formation. Various biofilm morphological parameters were quantified using Image Structure Analyzer software. Multivariate analysis demonstrated that both cell signaling and hydrodynamics significantly (P < 0.000) influenced biofilm structure. In turbulent flow, both biofilms formed streamlined patches, which in some cases developed ripple-like wave structures which flowed downstream along the surface of the flow cell. In laminar flow, both biofilms formed monolayers interspersed with small circular microcolonies. Ripple-like structures also formed in four out of six WT biofilms, although their velocity was approximately 10 times less than that of those that formed in the turbulent flow cells. The movement of biofilm cell clusters over solid surfaces may have important clinical implications for the dissemination of biofilm subject to fluid shear, such as that found in catheters. The ability of the cell signaling mutant to form biofilms in high shear flow demonstrates that signaling mechanisms are not required for the formation of strongly adhered biofilms. Similarity between biofilm morphologies in WT and mutant biofilms suggests that the dilution of signal molecules by mass transfer effects in faster flowing systems mollifies the dramatic influence of signal molecules on biofilm structure reported in previous studies.

Analysis of Variance↗
PubMed2001

Growth and detachment of cell clusters from mature mixed-species biofilms.

Detachment from biofilms is an important consideration in the dissemination of infection and the contamination of industrial systems but is the least-studied biofilm process. By using digital time-lapse microscopy and biofilm flow cells, we visualized localized growth and detachment of discrete cell clusters in mature mixed-species biofilms growing under steady conditions in turbulent flow in situ. The detaching biomass ranged from single cells to an aggregate with a diameter of approximately 500 microm. Direct evidence of local cell cluster detachment from the biofilms was supported by microscopic examination of filtered effluent. Single cells and small clusters detached more frequently, but larger aggregates contained a disproportionately high fraction of total detached biomass. These results have significance in the establishment of an infectious dose and public health risk assessment.

Biofilms↗
PubMed2001

The influence of fluid shear and AICI3 on the material properties of Pseudomonas aeruginosa PAO1 and Desulfovibrio sp. EX265 biofilms.

An understanding of the material properties of biofilms is important when describing how biofilms physically interact with their environment. In this study, aerobic biofilms of Pseudomonas aeruginosa PAO1 and anaerobic sulfate-reducing bacteria (SRB) biofilms of Desulfovibrio sp. EX265 were grown under different fluid shear stresses (tau g) in a chemostat recycle loop. Individual biofilm microcolonies were deformed by varying the fluid wall shear stress (tau w). The deformation was quantified in terms of strain (epsilon), and the relative strength of the biofilms was assessed using an apparent elastic coefficient (Eapp) and residual strain (epsilon r) after three cycles of deformation. Aluminium chloride (AlCl3) was then added to both sets of biofilm and the tests repeated. Biofilms grown under higher shear were more rigid and had a greater yield shear stress than those grown under lower shear. The addition of AlCl3 resulted in a significant increase in Eapp and also increased the yield point. We conclude that the strength of the biofilm is in part dependent on the shear under which the biofilm was grown and that the material properties of the biofilm may be manipulated through cation cross-linking of the extracellular polysaccharide (EPS) slime matrix.

Aluminum Chloride↗