1,720,968 research outputs found
Antibiotic loaded calcium sulfate bead and pulse lavage eradicates biofilms on metal implant materials in vitro
No full textPulse lavage (PL) debridement and antibiotic loaded calcium sulfate beads (CS-B) are both used for the treatment of biofilm related periprosthetic joint infection (PJI). However, the efficacy of these alone and in combination for eradicating biofilm from orthopaedic metal implant surfaces is unclear. The purpose of the study was to understand the efficacy of PL and antibiotic loaded CS-B in eradicating bacterial biofilms on 316L stainless steel (SS) alone and in combination in vitro. Biofilms of bioluminescent strains of Pseudomonas aeruginosa Xen41 and a USA300 MRSA Staphylococcus aureus SAP231 were grown on SS coupons for 3 days. The coupons were either, a) debrided for 3s with PL, b) exposed to tobramycin (TOB) and vancomycin (VAN) loaded CS-B for 24 h, or c) exposed to both. An untreated biofilm served as a control. The amount of biofilm was measured by bioluminescence, viable plate count and confocal microscopy using live/dead staining. PL alone reduced the CFU count of both strains of biofilms by approximately 2 orders of magnitude, from an initial cell count on metal surface of approximately 109 CFU/cm2. The antibiotic loaded CS-B caused an approximate 6 log reduction and the combination completely eradicated viable biofilm bacteria. Bioluminescence and confocal imaging corroborated the CFU data. While PL and antibiotic loaded CS-B both significantly reduced biofilm, the combination of two was more effective than alone in removing biofilms from SS implant surfaces
A commercial SnF2 toothpaste formulation reduces simulated human plaque biofilm in a dynamic typodont model
No full textAims: we present a dynamic typodont biofilm model (DTBM) incorporating 1) human dentition anatomy, 2) fluid flow over intermittently fluid bathed tooth surfaces and 3) an oxic headspace to allow aerobic and anaerobic niches to develop naturally, as a screening tool to assess the effect of stannous fluoride (SnF2) toothpaste against a simulated human plaque biofilm (SPB). Methods and results: first, hydroxyapatite (HA) coupons were inoculated with human saliva/plaque and cultured at 37oC under air. Selected species representative of common commensal and anaerobic pathogens were quantified for relative abundance changes over 4d by PCR densitometry to confirm the culture conditions allowed the proliferation of these species. A continuous culture DTBM reactor on a rocker table was inoculated with saliva/plaque and incubated at 37°C for 24h. Tooth shear stress was estimated by particle tracking. A SnF2 toothpaste solution, or a sham rise was administered twice daily for 3d to mimic routine oral hygiene. SPB biomass was assessed by total bacterial DNA and methylene blue (MB) staining. Early colonizer aerobes and late colonizer anaerobes species were detected in the HA and DTBM, and the trends in changing abundance were consistent with those seen clinically. Conclusions: treatment with the SnF2 solution showed significant reductions of 53.05% and 54.4% in the SPB by MB staining and DNA, respectively. Significance and impact of study: The model has potential for assessing dentition anatomy and fluid flow on the efficacy of antimicrobial efficacy against localized SPB and may be amenable to the plaque index clinical evaluation
Reduction in Pseudomonas aeruginosa and Staphylococcus aureus biofilms from implant materials in a diffusion dominated environment: Diffusion mediated biofilm eradication
No full textAntibiotic-loaded calcium sulfate beads (CS-B) are used to treat biofilm related periprosthetic joint infections (PJI). A previous study has shown that such beads are effective in reducing lawns biofilms grown on agar plates; however, the ability of CS-B to eradicate biofilms grown on solid orthopaedic material surfaces has not been investigated. We grew biofilms of bioluminescent strains of Pseudomonas aeruginosa Xen41 and a USA300 MRSA Staphylococcus aureus SAP231 on an ultra-high molecular weight polyethylene (PE), hydroxyapatite (HA), and 316L stainless steel (SS) coupons for three days under static growth conditions, with daily nutrient exchange. The coupons were rinsed with sterile phosphate buffered saline (PBS) to remove planktonic bacteria and placed in a petri dish, surrounded by four either antibiotic vancomycin and tobramycin loaded (CS-BV+T) or unloaded beads (CS-BU). A thin layer of agar was overlaid to simulate a periprosthetic infection where an implant abuts soft tissue, then incubated for 72 hours. The amount of biofilm was measured by bioluminescence imaging (BLI) for activity and viable cell count (CFUs). Coupons exposed to CS-BV+T showed a significant reduction in the amount of biofilm within 24 hours, regardless of the bacterial strain or material type. Whereas, coupons exposed to control CS-BU had no effect on bacteria over 72 hours. Statement of Clinical Significance: Antibiotic-loaded calcium sulfate beads (CS-B) were effective in significantly reducing mature biofilms of P. aeruginosa and S. aureus from orthopaedic relevant surfaces in our novel in vitro periprosthetic-soft tissue model
Biofilms in orthopaedic infections: a review on laboratory methods
No full textBacterial infection after hardware implantation in orthopaedic surgery is a devastating issue as it often necessitates increased hospital costs and stays, multiple revision surgeries, and prolonged use of antibiotics. Due to the nature of hardware implantation into the body, these infections are commonly in the form of attached biofilms. The current literature on a range of methodologies to study clinically explanted infected orthopaedic hardware, with potential biofilm, in the laboratory setting is limited. General methods include traditional and advanced culturing techniques, microscopy imaging techniques, and techniques that manipulate genetic material. The future of diagnostic techniques for infected implants, innovative hardware design, and treatment solutions for patients all depend on the successful evaluation and characterization of clinical samples in the laboratory setting. This review will provide an overview of current methods to study biofilms associated with orthopaedic infections, as well as provide insight into future directions in the field
Antibiotic loaded β-tricalcium phosphate/calcium sulfate for antimicrobial potency, prevention and killing efficacy of Pseudomonas aeruginosa and Staphylococcus aureus biofilms
No full textThis study investigated the efficacy of a biphasic synthetic β-tricalcium phosphate / calcium
sulfate (β-TCP/CS) bone graft substitute for compatibility with vancomycin (V) in
combination with tobramycin (T) or gentamicin (G) evidenced by the duration of potency and
the prevention and killing efficacies of P. aeruginosa (PAO1) and S. aureus (SAP231)
biofilms in in vitro assays. Antibiotic loaded β-TCP/CS beads were compared with antibiotic
loaded beads formed from a well characterized synthetic calcium sulfate (CS) bone void
filler. β-TCP/CS antibiotic loaded showed antimicrobial potency against PAO1 in a repeated
Kirby-Bauer like zone of inhibition assay for 6 days compared to 8 days for CS. However,
both bead types showed potency against SAP231 for 40 days. Both formulations loaded with
V+T completely prevented biofilm formation (CFU below detection limits) for the 3 days of
the experiment with daily fresh inoculum challenges (P < 0.001). In addition, both antibiotic
loaded materials and antibiotic combinations significantly reduced the bioburden of pre
grown biofilms by between 3 and 5 logs (P < 0.001) with V+G performing slightly better
against PAO1 than V+T. Our data, combined with previous data on osteogenesis suggest that
antibiotic loaded β-TCP/CS may have potential to stimulate osteogenesis through acting as a
scaffold as well as simultaneously protecting against biofilm infection. Future in-vivo
experiments and clinical investigations are warranted to more comprehensively evaluate the
use of β-TCP/CS in the management of orthopaedic infections
Ultrastructure imaging of Pseudomonas aeruginosa lawn biofilms and eradication of the tobramycin-resistant variants under in vitro electroceutical treatment
No full textElectrochemically generated bactericidal compounds have been shown to eradicate bacterial lawn biofilms through electroceutical treatment. However, the ultrastructure of biofilms exposed to these species has not been studied. Moreover, it is unknown if the efficacy of electroceutical treatment extends to antibiotic-resistant variants that emerge in lawn biofilms after antibiotic treatment. In this report, the efficacy of the in vitro electroceutical treatment of Pseudomonas aeruginosa biofilms is demonstrated both at room temperature and in an incubator, with a ~ 4 log decrease (p < 0.01) in the biofilm viability observed on the anode at both conditions. The ultrastructure changes in the lawn biofilms imaged using transmission electron microscopy demonstrate significant bacterial cell damage at the anode after 24 h of electroceutical treatment. A mix of both damaged and undamaged cells was observed at the cathode. Finally, both eradication and prevention of the emergence of tobramycin-resistant variants were demonstrated by combining antibiotic treatment with electroceutical treatment on the lawn biofilms
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
Mapping bacterial biofilms on recovered orthopaedic implants by a novel agar candle dip method
No full textWhile the detrimental effects of periprosthetic joint infections (PJIs) are well-known, the process of biofilm formation on orthopaedic hardware is unclear. Previous work has shown that encasement of explant hardware in agar can aid in identifying biofilms. This study tested the utility of agar “candle dip” method in detecting and mapping the location of biofilm on infected orthopedic components. Explant components from fifteen patients were rinsed, briefly submerged in agar to create a surface coating, and incubated. Larger components were coated by pipetting agar over them. After incubation, colony outgrowth on the component surface was documented (candle dip status). Data was compared with clinical laboratory results (clinical culture status) and PJI diagnosis using Musculoskeletal Infection Society criteria (MSIS status). All six patients classified as MSIS and clinical culture positive were also positive with candle dip technique. Of the nine-candle dip negative cases, four were positive and five were negative for both MSIS and clinical culture status. Candle dip may be negative in few cases due to the residual antibiotic eluting from the spacers, limiting growth of bacterial biofilms on the components. The candle dip method shows promise for biofilm mapping but requires additional testing to evaluate clinical diagnostic potential
Targeting intracellular Staphylococcus aureus to lower recurrence of orthopedic infection
No full textStaphylococcus aureus is often found in orthopaedic infections and may be protected from commonly prescribed antibiotics by forming biofilms or growing intracellularly within osteoblasts. To investigate the effect of non-antibiotic compounds in conjunction with antibiotics to clear intracellular and biofilm forming S. aureus causing osteomyelitis. SAOS-2 osteoblast-like cell lines were infected with S. aureus BB1279. Antibiotics (vancomycin, VAN; and dicloxacillin, DICLOX), bacterial efflux pump inhibitors (piperine, PIP; carbonyl cyanide m-chlorophenyl hydrazone, CCCP) and bone morphogenetic protein (BMP-2) were evaluated individually and in combination to kill intracellular bacteria. We present direct evidence that after gentamicin killed extracellular planktonic bacteria and antibiotics had been stopped, seeding from the infected osteoblasts grew as biofilms. VAN was ineffective in treating the intracellular bacteria even at 10x MIC, however in presence of PIP or CCCP the intracellular S. aureus was significantly reduced. Bacterial efflux pump inhibitors (PIP and CCCP) were effective in enhancing permeability of antibiotics within the osteoblasts and facilitated killing of intracellular S. aureus. Confocal laser scanning microscopy (CLSM) showed increased uptake of propidium iodide within osteoblasts in presence of PIP and CCCP. BMP-2 had no effect on growth of S. aureus either alone or in combination with antibiotics. Combined application of antibiotics and natural agents could help in the treatment of osteoblast infected intracellular bacteria and biofilms associated with osteomyelitis. <br/
Bacterial DNA is associated with tunnel widening in failed ACL reconstructions
No full textPurpose: To determine if tunnel widening, defined as change in maximal tunnel diameter from the time of initial bone tunnel drilling to revision surgery is associated with bacterial deoxyribonucleic acid (DNA) presence and concentration in torn graft tissue from failed anterior cruciate ligament reconstructions (ACLRs). Methods: Thirty-four consecutive revision ACLRs were included (mean age 27.3 years SD 10.9; median time to failure 4.9 years range 105 days–20 years). Graft selection of the failed reconstruction was 68% autograft, 26% allograft, and 6% autograft/allograft hybrid with a mean drilled tunnel diameter of 8.4 mm SD 0.8. Maximal tunnel diameters prior to revision were measured on pre-operative three-dimensional imaging and compared to drilled tunnel diameters at the time of the previous reconstruction. Tissue biopsies of the failed graft were obtained from tibial, femoral, and intraarticular segments. Sterile water left open to air during revision ACLRs and tissue from primary ACLRs were used as negative controls. Clinical cultures were obtained on all revision ACLRs and PCR with universal bacterial primer on all cases and negative controls. Fluorescence microscopy was used to confirm the presence and location of biofilms in two patients with retrieved torn graft tissue and fixation material. Amount of tunnel widening was compared to bacterial DNA presence as well as bacterial DNA concentration via Welch ANOVA. Results: Bacterial DNA was present in 29/34 (85%) revision ACLRs, 1/5 (20%) of primary ACLR controls and 0/3 (0%) sterile water controls. Cultures were positive (coagulase negative Staphylococcus sp.) in one case, which also had the greatest degree of tunnel widening. Femoral widening was greater in cases with detectable bacterial DNA (mean widening 2.6 mm SD 3.0) versus without (mean 0.3 mm SD 0.6) (p = 0.003) but was unaffected by bacterial DNA concentration (p = 0.44). Tibial widening was not associated with the presence of bacterial DNA (n.s.); however, higher bacterial DNA concentrations were observed in cases with tibial widening ≥ 3.0 mm (median 2.47 ng bacterial DNA/µg total DNA) versus widening < 3.0 mm (median 0.97 ng bacterial DNA/µg total DNA) (p = 0.046). Tunnel widening was not associated with time to failure, graft selection, or number of prior surgeries (n.s., all comparisons). Fluorescence microscopy confirmed the presence of biofilms on ruptured tendon graft as well as fixation material in 2/2 cases. Conclusion: Bacterial DNA is commonly encountered on failed ACLR grafts and can form biofilms. Bacterial DNA does not cause clinically apparent infection symptoms but is associated with tunnel widening. Further research is needed to determine whether graft decontamination protocols can reduce graft bacterial colonization rates, ACLR tunnel widening or ACLR failure risk. Level of evidence: Therapeutic III.</p
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