48 research outputs found
Free-floating aggregate and single cell-initiated biofilms of Staphylococcus aureus
No full textPeriprosthetic joint infection (PJI) occurring after artificial joint replacement is a major clinical issue requiring multiple surgeries and antibiotic interventions. Staphylococcus aureus is the common bacteria responsible for PJI. Recent in vitro research has shown that staphylococcal strains rapidly form free-floating aggregates in the presence of synovial fluid (SF) with biofilm-like resistance to antimicrobial agents. However, the development of biofilms formed from these aggregates under shear have not been widely investigated. Thus, in this study, we examined the progression of attached biofilms from free-floating aggregates. Biofilms were grown for 24 hours in flow cells on titanium discs after inoculating with either pre-aggregated or single planktonic cells. Image analysis showed no significant difference between the biofilm formed from aggregates vs. the planktonic cells in terms of biomass, surface area, and thickness. Regarding antibiotic susceptibility there was 1 and 2 log reductions in biofilms formed from single cells and aggregates respectively when treated with vancomycin for 24 hours. Thus, this study demonstrates the formation of biofilm from free-floating aggregates follows a similar developmental time-period and shows similar antibiotic tolerance as more traditionally inoculated in vitro flow cell biofilms
The influence of patterned surface features on the accumulation of bovine synovial fluid induced aggregates of Staphylococcus aureus
No full textPeriprosthetic joint infection (PJI) after joint replacement is a major clinical issue requiring multiple surgeries and antibiotic interventions. Recent in vitro research has shown that PJI staphylococcal strains rapidly form antibiotic resistant free-floating aggregates in the presence of bovine synovial fluid (BSF). Staphylococcal aggregates are also present in human PJI joint fluid. However, the influence of surface roughness and fluid shear on the attachment and retention of such aggregates to surfaces is not known. Our aim was to assess how surface roughness and fluid shear stress influenced the attachment and retention of S. aureus BSF mediated aggregates on smooth and rough patterned titanium in flow cells compared to non-aggregated cells. S. aureus attachment of aggregates was significantly greater than single cells but was independent of surface roughness, however on the patterned surfaces aggregates preferentially accumulated in the grooves. Fibrous components in the BSF were also colocalized with the grooves. After a 24 hr attachment and incubation period different shear stresses were applied. There was significant detachment from flat surface at 1 mL/min (τw = 0.03 Pa) but minimal detachment from the patterned surfaces, even at flow rates as high as 13.9 mL/min (τw = 0.42 Pa). The retention of bacterial aggregates and biofilm by rough surfaces exposed to shear might be an important consideration of the colonization location on orthopedic implants which can have a wide range of roughness and surface features as well as influencing efficacy of shear-based debridement methods such as pulse lavage
Staphylococcus aureus aggregates on orthopedic materials under varying levels of shear stress
No full textPeriprosthetic joint infection (PJI) occurring after artificial joint replacement is a major clinical issue requiring multiple surgeries and antibiotic interventions. Staphylococcus aureus is the bacterium most commonly responsible for PJI. Recent in vitro research has shown that staphylococcal strains rapidly form aggregates in the presence of synovial fluid (SF). We hypothesize that these aggregates provide early protection to bacteria entering the wound site, allowing them time to attach to the implant surface, leading to biofilm formation. Thus, understanding the attachment kinetics of these aggregates is critical in understanding their adhesion to various biomaterial surfaces. In this study, the number, size, and surface area coverage of aggregates as well as of single cells of S. aureus were quantified under various conditions on different orthopedic materials relevant to orthopedic surgery: stainless steel (316L), titanium (Ti), hydroxyapatite (HA), and polyethylene (PE). It was observed that, regardless of the material type, SF-induced aggregation resulted in reduced aggregate surface attachment and greater aggregate size than the single-cell populations under various shear stresses. Additionally, the surface area coverage of bacterial aggregates on PE was relatively high compared to that on other materials, which could potentially be due to the rougher surface of PE. Furthermore, increasing shear stress to 78 mPa decreased aggregate attachment to Ti and HA while increasing the aggregates' average size. Therefore, this study demonstrates that SF induced inhibition of aggregate attachment to all materials, suggesting that biofilm formation is initiated by lodging of aggregates on the surface features of implants and host tissues.</p
Killing of a multispecies biofilm using a gram-negative and gram-positive targeted antibiotic released from high purity calcium sulfate beads
No full textBackground: multispecies biofilm orthopedic infections are more challenging to treat than mono-species infections. In this in-vitro study, we aimed to determine if a multispecies biofilm, consisting of Gram positive and negative species with different antibiotic susceptibilities could be treated more effectively using high purity antibiotic-loaded calcium sulfate beads (HP-ALCSB) containing vancomycin (VAN) and tobramycin (TOB) in combination than alone. Methods: three sets of species pairs from bioluminescent strains of Pseudomonas aeruginosa (PA) and Staphylococcus aureus (SA) and clinical isolates, Enterococcus faecalis (EF) and Enterobacter cloacae were screened for compatibility. PA+EF developed intermixed biofilms with similar cell concentrations and so were grown on 316L stainless steel coupons for 72 h or as 24 h agar lawn biofilms and then treated with HP-ALCSBs with single or combination antibiotics and assessed by viable count or bioluminescence and light imaging to distinguish each species. Replica plating was used to assess viability. Results: the VAN+TOB bead significantly reduced the PA+EF biofilm CFU and reduced the concentration of surviving antibiotic tolerant variants by 50% compared to single antibiotics. Conclusion: the combination of Gram-negative and positive targeted antibiotics released from HP-ALCSBs may be more effective in treating multispecies biofilms than monotherapy alone
Mapping bacterial biofilm on features of orthopedic implants in vitro
No full textImplant-associated infection is a major complication of orthopedic surgery. One of the most common organisms identified in periprosthetic joint infections is Staphylococcus aureus, a biofilmforming pathogen. Orthopedic implants are composed of a variety of materials, such as titanium, polyethylene and stainless steel, which are at risk for colonization by bacterial biofilms. Little is known about how larger surface features of orthopedic hardware (such as ridges, holes, edges, etc.) influence biofilm formation and attachment. To study how biofilms might form on actual components, we submerged multiple orthopedic implants of various shapes, sizes, roughness and material type in brain heart infusion broth inoculated with Staphylococcus aureus SAP231, a bioluminescent USA300 strain. Implants were incubated for 72 h with daily media exchanges. After incubation, implants were imaged using an in vitro imaging system (IVIS) and the metabolic signal produced by biofilms was quantified by image analysis. Scanning electron microscopy was then used to image different areas of the implants to complement the IVIS imaging. Rough surfaces had the greatest luminescence compared to edges or smooth surfaces on a single implant and across all implants when the images were merged. The luminescence of edges was also significantly greater than smooth surfaces. These data suggest implant roughness, as well as large-scale surface features, may be at greater risk of biofilm colonization.</p
Investigation of synovial fluid induced <i>Staphylococcus aureus</i> aggregate development and its impact on surface attachment and biofilm formation
No full textPeriprosthetic 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.</div
Pseudomonas aeruginosa biofilm killing beyond the spacer by antibiotic loaded calcium sulfate beads: an in vitro study
No full text
Introduction: Bacterial biofilms are an important virulence factor in chronic periprosthetic joint infection (PJI) and other orthopedic infection since they are highly tolerant to antibiotics and host immunity. Antibiotics are mixed into carriers such as bone cement and calcium sulfate bone void fillers to achieve sustained high concentrations of antibiotics required to more effectively manage biofilm infections through local release. The effect of antibiotic diffusion from antibiotic-loaded calcium sulfate beads (ALCS-B) in combination with PMMA bone cement spacers on the spread and killing of
Pseudomonas aeruginosa Xen41 (PA-Xen41) biofilm was investigated using a "large agar plate" model scaled for clinical relevance.
Methods: Bioluminescent PA-Xen41 biofilms grown on discs of various orthopedic materials were placed within a large agar plate containing a PMMA full-size mock "spacer" unloaded or loaded with vancomycin and tobramycin, with or without ALCS-B. The amount of biofilm spread and log reduction on discs at varying distances from the spacer was assessed by bioluminescent imaging and viable cell counts.
Results: For the unloaded spacer control, PA-Xen41 spread from the biofilm to cover the entire plate. The loaded spacer generated a 3 cm zone of inhibition and significantly reduced biofilm bacteria on the discs immediately adjacent to the spacer but low or zero reductions on those further away. The combination of ALCS-B and a loaded PMMA spacer greatly reduced bacterial spread and resulted in significantly greater biofilm reductions on discs at all distances from the spacer.
Discussion: The addition of ALCS-B to an antibiotic-loaded spacer mimic increased the area of antibiotic coverage and efficacy against biofilm, suggesting that a combination of these depots may provide greater physical antibiotic coverage and more effective dead space management, particularly in zones where the spread of antibiotic is limited by diffusion (zones with little or no fluid motion).
</p
Application of Non-Thermal Plasma on Biofilm: A Review
No full textThe formation of bacterial biofilm on implanted devices or damaged tissues leads to biomaterial-associated infections often resulting in life-threatening diseases and implant failure. It is a challenging process to eradicate biofilms as they are resistant to antimicrobial treatments. Conventional techniques, such as high heat and chemicals exposure, may not be suitable for biofilm removal in nosocomial settings. These techniques create surface degradation on the treated materials and lead to environmental pollution due to the use of toxic chemicals. A novel technique known as non-thermal plasma has a great potential to decontaminate or sterilize those nosocomial biofilms. This article aims to provide readers with an extensive review of non-thermal plasma and biofilms to facilitate further investigations. A brief introduction summarizes the problem caused by biofilms in hospital settings with current techniques used for biofilm inactivation followed by the literature review strategy. The remainder of the review discusses plasma and its generation, the role played by plasma reactive species, various factors affecting the antimicrobial efficacy of non-thermal plasma and summarizes many studies published in the field
Non-Thermal Plasma in Conjunction With Chlorhexidine (CHX) Digluconate Sterilize the Biofilm Contaminated Titanium Surface
No full text
