RESUMO
Changes in temperature were found to affect the morphology, cell viability, and mechanical properties of Staphylococcus epidermidis bacterial biofilms. S. epidermidis biofilms are commonly associated with hospital-acquired medical device infections. We observed the effect of heat treatment on three physical properties of the biofilms: the bacterial cell morphology and viability, the polymeric properties of the extracellular polymeric substance (EPS), and the rheological properties of the bulk biofilm. After application of a 1 h heat treatment at 45 °C, cell reproduction had ceased, and at 60 °C, cell viability was significantly reduced. Size exclusion chromatography was used to fractionate the extracellular polymeric substance (EPS) based on size. Chemical analysis of each fraction showed that the relative concentrations of the polysaccharide, protein, and DNA components of the EPS were unchanged by the heat treatment at 45 and 60 °C. The results suggest that the EPS molecular constituents are not significantly degraded by the temperature treatment. However, some aggregation on the scale of 100 nm was found by dynamic light scattering at 60 °C. Finally, relative to control biofilms maintained at 37 °C, we observed an order of magnitude reduction in the biofilm yield stress after 60 °C temperature treatment. No such difference was found for treatment at 45 °C. From these results, we conclude that the yield stress of bacterial biofilms is temperature-sensitive and that this sensitivity is correlated with cell viability. The observed significant decrease in yield stress with temperature suggests a means to weaken the mechanical integrity of S. epidermidis biofilms with applications in areas such as the treatment of biofilm-infected medical devices.
Assuntos
Aderência Bacteriana , Biofilmes , Biopolímeros/metabolismo , Fenômenos Mecânicos , Staphylococcus epidermidis/fisiologia , Temperatura , Fenômenos Biomecânicos , Sobrevivência Celular , Módulo de Elasticidade , Espaço Extracelular/metabolismo , Hidrodinâmica , Peso Molecular , Reologia , Staphylococcus epidermidis/citologia , Staphylococcus epidermidis/metabolismo , Estresse MecânicoRESUMO
Proton nuclear magnetic resonance (H-NMR) spectroscopic analysis of cerebral spinal fluid provides a quick, non-invasive modality for evaluating the metabolic activity of brain-injured patients. In a prospective study, we compared the CSF of 44 TBI patients and 13 non-injured control subjects. CSF was screened for ten parameters: ß-glucose (Glu), lactate (Lac), propylene glycol (PG), glutamine (Gln), alanine (Ala), α-glucose (A-Glu), pyruvate (PYR), creatine (Cr), creatinine (Crt), and acetate (Ace). Using mixed effects measures, we discovered statistically significant differences between control and trauma concentrations (mM). TBI patients had significantly higher concentrations of PG, while statistical trends existed for lactate, glutamine, and creatine. TBI patients had a significantly decreased concentration of total creatinine. There were no significant differences between TBI patients and non-injured controls regarding ß- or α-glucose, alanine, pyruvate or acetate. Correlational analysis between metabolites revealed that the strongest significant correlations in non-injured subjects were between ß- and α-glucose (r = 0.74), creatinine and pyruvate (r = 0.74), alanine and creatine (r = 0.62), and glutamine and α-glucose (r = 0.60). For TBI patients, the strongest significant correlations were between lactate and α-glucose (r = 0.54), lactate and alanine (r = 0.53), and α-glucose and alanine (r = 0.48). The GLM and multimodel inference indicated that the combined metabolites of PG, glutamine, α-glucose, and creatinine were the strongest predictors for CMRO2, ICP, and GOSe. By analyzing the CSF of patients with TBI, our goal was to create a metabolomic fingerprint for brain injury.
Assuntos
Aminoácidos/líquido cefalorraquidiano , Lesões Encefálicas/líquido cefalorraquidiano , Propilenoglicol/líquido cefalorraquidiano , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Análise de Variância , Feminino , Glucose/líquido cefalorraquidiano , Humanos , Pressão Intracraniana , Espectroscopia de Ressonância Magnética , Masculino , Metabolômica , Pessoa de Meia-Idade , Prótons , Adulto JovemRESUMO
Central line-associated bloodstream infections (CLABSIs) are not easily treated, and many catheters (e.g., hemodialysis catheters) are not easily replaced. Biofilms (the source of infection) on catheter surfaces are notoriously difficult to eradicate. We have recently demonstrated that modest elevations of temperature lead to increased staphylococcal susceptibility to vancomycin and significantly soften the biofilm matrix. In this study, using a combination of microbiological, computational, and experimental studies, we demonstrate the efficacy, feasibility, and safety of using heat as an adjuvant treatment for infected hemodialysis catheters. Specifically, we show that treating with heat in the presence of antibiotics led to additive killing of Staphylococcus epidermidis with similar trends seen for Staphylococcus aureus and Klebsiella pneumoniae. The magnitude of temperature elevation required is relatively modest (45-50°C) and similar to that used as an adjuvant to traditional cancer therapy. Using a custom-designed benchtop model of a hemodialysis catheter, positioned with tip in the human vena cava as well as computational fluid dynamic simulations, we demonstrate that these temperature elevations are likely achievable in situ with minimal increased in overall blood temperature.
Assuntos
Biofilmes , Infecções Relacionadas a Cateter/terapia , Hipertermia Induzida , Infecções por Klebsiella/terapia , Infecções Estafilocócicas/terapia , Antibacterianos/uso terapêutico , Infecções Relacionadas a Cateter/tratamento farmacológico , Infecções Relacionadas a Cateter/microbiologia , Cateterismo Venoso Central , Cateteres de Demora/microbiologia , Temperatura Alta , Humanos , Hidrodinâmica , Infecções por Klebsiella/fisiopatologia , Klebsiella pneumoniae , Modelos Cardiovasculares , Diálise Renal , Infecções Estafilocócicas/fisiopatologia , Staphylococcus aureus , Staphylococcus epidermidis , Vancomicina/uso terapêuticoRESUMO
Given the increasing evidence of safe application of elevated temperature in other clinical contexts, we consider the potential for supplemental hyperthermia to augment the effects of vancomycin against staphylococci, a major source of postoperative and posttraumatic sepsis. Laboratory reference strains and libraries of clinical blood isolates of Staphylococcus epidermidis and methicillin-resistant Staphylococcus aureus, both as planktonic cells and as established biofilms, were assessed for thermosensitivity and increased susceptibility to vancomycin in the setting of thermal treatment. In addition to viability measures, patterns of stress gene expression were assessed with quantitative polymerase chain reaction, and structural changes were measured using quantitative transmission electron microscopy. Laboratory strains of both species had reduced growth and biofilm viability at 45°C, a temperature commonly used in other domains such as adjuvant treatments of malignancy. Blood isolates of S. epidermidis were consistent in this regard as well, but significant between-isolate variability in thermosensitivity was seen in blood isolates of S. aureus. Expression profiling and ultrastructural measurements confirmed that elevated temperature was a substantial stressor with or without vancomycin treatment. Our findings suggest that temperature elevations shown to be tolerated in humans in other settings hold the potential to be used as an adjuvant to antibiotic therapy against staphylococcal biofilms.