Background. Continuous post-operative infusion of local anaesthetic solutions has been implicated as the causative factor in many cases of chondrolysis. Recent in-vitro studies have shown that even a single exposure to local anaesthetic can cause apoptosis and mitochondrial dysfunction leading to chondrocyte death. Glucosamine has been shown to have a protective and reparative effect on articular cartilage. Aims. To compare the effect of a single exposure of different local anaesthetic solutions on human articular cartilage and to investigate the protective and reparative effects of Glucosamine on articular cartilage exposed to 0.5% Bupivacaine. Methods. Chondral explants (n=354) were obtained from femoral heads of hip fracture patients undergoing hemiarthroplasty. Each specimen was exposed to one of 8 test solutions for one hour. The specimens were then incubated in culture medium containing radio-labelled 35-sulphur for 16 hours. The uptake of 35-S by each specimen was measured to give an estimate of proteoglycan metabolism. Test solutions. 1. 1% Lidocaine 2. 2% Lidocaine 3. 0.25% Bupivacaine, 4. 0.5% Bupivacaine, 5. 0.5% Levo-Bupivacaine 6. Control solution of M199 culture medium. 7. To investigate its protective effect, 100 micrograms of Glucosamine was added along with 0.5% Bupivacaine 8. To investigate its reparative effect, Glucosamine was added after exposure to Bupivacaine for an hour. Results. Compared to the control solution, the inhibition of proteoglycan metabolism was 64% with 1% Lidocaine(p< 0.001), 79% with 2% Lidocaine(p< 0.001), 61% with 0.25% Bupivacaine(p< 0.001), 85% with 0.5% Bupivacaine(p< 0.001) and 77% with 0.5% Levo-Bupivacaine(p< 0.001). Adding Glucosamine reduced Bupivacaine toxicity to 43%(p< 0.001). Glucosamine marginally repaired the damage caused by Bupivacaine, with inhibition of proteoglycan metabolism at 70%(p=0.004). Conclusion. All local anaesthetic solutions were toxic to articular cartilage. The addition of Glucosamine to 0.5% Bupivacaine protected against its toxicity to articular cartilage

Introduction. Day stay surgery for anterior cruciate ligament (ACL) reconstructions is an increasingly common practice and has driven clinicians to come up with postoperative pain regimes that allow same day mobilisation and a safe and timely discharge. There is a paucity of literature surrounding the use of intraosseous (IO) ropivacaine used as a Bier's block to provide both intraoperative and postoperative analgesia in lower limb surgery. Methods. This patient blinded, pilot study randomised 15 patients undergoing ACL reconstruction to receive either IO ropivacaine 1.5 or 2.0 mg/kg; or 300 mg of ropivacaine as local infiltration (standard of care). Toxic plasma levels of ropivacaine have been defined in the literature and therefore the primary outcome for this study was arterial plasma concentration of ropivacaine as a means to determine its safety profile. Samples were taken via an arterial line at prespecified times after tourniquet deflation. Secondary outcomes that we were interested in included immediate postoperative pain scores using the visual analogue scale (VAS) and perioperative opioid equivalent consumption. Results. Participants had a mean age of 27.8 (SD 9.2) years and 87% (13/15) were male. All patients in the intervention group receiving IO ropivacaine had plasma concentrations well below the threshold for central nervous system (CNS) toxicity (0.60 µg/ml). The highest plasma concentration was achieved in the intervention group receiving 1.5 mg/kg dose of ropivacaine reaching 3.59 mg/ml. This would equate to 0.22 µg/ml of free plasma ropivacaine. There were no differences across the three groups regarding pain scores or perioperative opioid consumption. Conclusions. This study demonstrates that IO administration of 0.2% ropivacaine is both safe and effective in reducing perioperative pain in patients undergoing ACL reconstruction. There may be scope to increase the IO dose further or utilise other analgesics via the IO regional route to improve perioperative pain relief

Aim. Infections represent a serious threat to the successful utilization of implants in modern medicine. Implant-associated infections are difficult to treat, because they involve biofilms that protect bacteria from the immune system and harbour antibiotic-tolerant persister cells. In this work, we developed an antibody-drug conjugate (ADC) containing the anti-neoplastic drug mitomycin C (MMC) as a novel treatment paradigm for implant-associated infections. MMC was chosen as it is a potent antimicrobial against biofilms and its synthesis into an ADC was chosen to alleviate toxicity. Following development and synthesis of the ADC, stability and release of MMC was measured. We then used the ADC to kill bacteria in suspension and in biofilms, in vitro and in vivo. Method. Mitomycin C was conjugated to a commercially available antibody against S. aureus via a disulfide linkage, with a drug release occurred via thiol-disulfide exchange. ADC as tested against S. aureus under various growth conditions (planktonic, persisters and biofilm). In vitro toxicity of ADC vs MMC was measured using a human cell line (MOLT-4). Finally, two independent in vivo experiments were performed in a murine implant-associated osteomyelitis model. In experiment one ADC treatment was compared NaCl, vancomycin and vancomycin + ADC (n=10 for all groups). Subsequently, ADC was compared to NaCl, the antibody used in the ADC construction, MMC and a novel ADC constructed with a non-S. aureus antibody (n=10 for all groups). All treatments were started day 7 post inoculation and were administered for 3 days. CFU enumeration was done following sonication to quantify bacterial load. Results. Drug release could be triggered on demand with N-acetyl cysteine and release occurred, once in contact with free thiols on S. aureus cell surface. The ADCs exhibited a concentration-dependent antimicrobial effect against S. aureus with doses exceeding 0.5 mg/l reducing amount of CFU to below detection limit (p< 0.001). 15 minutes exposure to ADC resulted in an approx. 2 log CFU/ml reduction compared to untreated biofilms (p < 0.01). In vivo ADC treatment was effective compared to NaCl treatment and the vancomycin treatment (p≤ 0.001). Further ADC and MMC treatment were comparable in efficacy, but both were superior than NaCl, pure antibody and the non-specific ADC (p≤ 0.05). Finally, in vitro cytotoxicity was significantly lower for ADC than MMC. Conclusions. In this study we have demonstrated that ADCs can be a novel treatment approach to combat implant-associated infections caused by S. aureus

Aim. Chronic bone infections and infected fractures are often treated with excision of the dead bone and implantation of biomaterials which elute antibiotics. Gentamicin has been a preferred drug for local delivery, but this could induce renal dysfunction due to systemic toxicity. This is a particular concern in patients with pre-existing chronic renal disease treated with new antibiotic carriers which achieve very high peak levels of gentamicin in the first few days after surgery. Method. 163 patients (109 males; average age 51.6 years) with Cierny-Mader Type 3 or 4 chronic osteomyelitis had a single-stage operation with excision of the dead bone, filling of the osseous defect with a calcium sulphate-hydroxyapatite carrier, containing gentamicin and immediate soft tissue closure. 2. No patient was given systemic gentamicin or other renal toxic antibiotics. Mean carrier volume was 10.9mls (range 1–30mls) and mean gentamicin dosing was 190.75mg (maximum 525mg). Seven patients had pre-existing renal disease (4 diabetic nephropathy, 1 nephrotic syndrome, 1 renal transplant and 1 previous acute kidney injury). Serum creatinine levels were collected pre-operatively and during the first seven days post-operatively. Glomerular filtration rate (GFR) was calculated using the CKD-epi creatinine equation. Renal function was defined using the Chronic Kidney Disease (CKD) Staging system. Results. 155 cases had adequate data to allow calculation of pre- and post-operative GFR. Pre-operative CKD staging demonstrated 118 Class I (normal renal function), 30 Class II, 3 Class IIIa, 3 Class IIIb, and 1 Class V disease. Mean pre-operative GFR (99.7ml/min/1.73m. 2. , SD 21.0) was no different to post-operative GFR (103.2ml/min/1.73m. 2. , SD 21.3), p= 0.0861. Four cases had a >10% decline in GFR below normal, with only one case dropping a CKD stage, from I (normal) to II (mildly decreased). Only 1/7 cases with pre-existing renal disease had a GFR drop of >10% (from 11ml/min/1.73m. 2. to 8ml/min/1.73m. 2. ). 70/155 (45.2%) had a temporary GFR drop post-operatively, with the biggest drop occurring a mean 3.06 days following surgery (SD 2.1). No patient had clinical signs of new acute renal impairment post-operatively. Conclusions. Renal function is not significantly affected by local implantation of gentamicin up to 525mg. The presence of pre-existing renal disease is not a contraindication to local gentamicin therapy

Aim. Treatment of prosthetic joint infection (PJI) by systemic administration of high doses of long-term antibiotics often proves ineffective, causing severe side effects. Thus, we presented the phage Sb-1, which coding extracellular polymeric substances (EPS) degradation depolymerases, conjugated with rifampicin-loaded liposomes (Lip-RIF@Phage) by bio-orthogonal functionalization strategy to target biofilm (Figure1). Method. Methicillin-resistant Staphylococcus aureus (MRSA) biofilm was grown on porous glass beads for 24 h in vitro. After the biofilm formation, beads were exposed to 0.9% saline, then sonication. Quantitative and qualitative biofilm analyses were performed by colony counting, scanning electron microscopy and isothermal microcalorimetry. A rat model of total knee arthroplasty infected with the bioluminescent MRSA strain was developed as the PJI model to evaluate the efficacy of Lip-RIF@Phage anti-biofilm therapy in vivo, then the creatinine, alanine transaminase, and aspartate transaminase values were evaluated throughout the entire treatment process. Results. After treatment with Lip-RIF@Phage, no bacterial colonies were observed, consistent with findings from scanning electron microscopy. Similarly, isothermal microcalorimetry revealed no detectable heat following Lip-RIF@Phage treatment, aligning with these observations. In vivo experiments demonstrated a significant reduction in biofilm cell load compared to all other tested conditions, with no evidence of systemic toxicity on renal and liver functions attributed to Lip-RIF@Phage. Conclusions. The innovative depolymerase-phagobot nanosystem (Lip-RIF@Phage) exhibits remarkable efficacy in completely eliminating biofilm cells in vitro. It serves as an excellent carrier for antibiotic delivery, enhancing antibiotic penetration through biofilms and improving biofilm eradication efficacy. Furthermore, it enables personalized treatment strategies against biofilm-associated multidrug-resistant (MDR) infections by maximizing the effectiveness of any remaining sensitive antibiotics. For any tables or figures, please contact the authors directly

Staphylococcus aureus is the most frequently isolated organism in periprosthetic joint infections. The mechanism by which synovial fluid (SF) kills bacteria has not yet been elucidated, and a better understanding of its antibacterial characteristics is needed. We sought to analyze the antimicrobial properties of exogenous copper in human SF against S. aureus. SF samples were collected from patients undergoing total elective knee or hip arthroplasty. Different S. aureus strains previously found to be sensitive and resistant, UAMS-1 and USA300 WT, respectively, were used. We performed in-vitro growth and viability assays to determine the capability of S. aureus to survive in SF with the addition of 10µM of copper. We determined the minimum bactericidal concentration of copper (MBC-Cu) and evaluated the sensitivity to killing, comparing WT and CopAZB-deficient USA300 strains. UAMS-1 evidenced a greater sensitivity to SF when compared to USA300 WT, at 12 (p=0.001) and 24 hours (p=0.027). UAMS-1 significantly died at 24 hours (p=0.017), and USA300 WT survived at 24 hours. UAMS-1 was more susceptible to the addition of copper at 4 (p=0.001), 12 (p=0.005) and 24-hours (p=0.006). We confirmed a high sensitivity to killing with the addition of exogenous copper on both strains at 4 (p=0.011), 12 (p=0.011), and 24 hours (p=0.011). Both WT and CopAZB-deficient USA300 strains significantly died in SF, evidencing a MBC-Cu of 50µM against USA300 WT (p=0.011). SF has antimicrobial properties against S. aureus, and UAMS-1 was more sensitive than USA300 WT. The addition of 10µM of copper was highly toxic for both strains, confirming its bactericidal effect. We evidenced CopAZB-proteins involvement in copper effluxion by demonstrating the high sensitivity of the mutant strain to lower copper concentrations. Thus, we propose CopAZB-proteins as potential targets and the use of exogenous copper as possible treatment alternatives against S. aureus

Hypochlorous acid (HOCl) is a potent anti-bacterial agent which could reduce periprosthetic joint infection. Early infection complications in joint replacements are often considered to be due to local contamination at the time of surgery and result in a significant socioeconomic cost. Current theatre cleaning procedures produce “clean” operating theatres which still contain bacteria (colony forming units, CFU). Reducing this bacterial load may reduce local contamination at the time of surgery. HOCl is produced naturally in the human neutrophil and has been implicated as the primary agent involved in bacterial killing during this process. In vitro research confirms its efficacy against essentially all clinically relevant bacteria. The recent advent of commercial production of HOCl, delivered as a fog, has resulted in extensive use in the food industry. Reported lack of corrosion and high anti-bacterial potency are seen as two key factors for the use of HOCl in the orthopaedic environment. Prior work by the authors comparing human cell toxicity of HOCl, chlorhexidine and iodine solutions shows favourable results. This study evaluates use of neutral HOCl applied as a dry room fog to decrease bacteria in the operating theatre environment. Using an animal operating theatre as the test site, bacterial swabs were taken from ten 100cm. 2. sample areas before standard cleaning with detergent, after standard cleaning, and again after 60 minutes exposure to HOCl fog. After standard cleaning, 6 of 10 sample sites recorded significant bacterial growth (>10 CFU/100cm. 2. ). After exposure to HOCl fog, growth in all 10 sites was below detection limits (<10 CFU/100cm. 2. ). This was repeated with specific exposure to Staphylococcus aureus and Escherichia coli. We can conclude that HOCl is effective when used as a fogging agent to reduce bacterial loading within an operating theatre environment and as such has significant potential to reduce intraoperative contamination and periprosthetic infection

Aim. The utilization of silver as an anti-infective agent is a subject of debate within the scientific community, with recurring discussions surrounding its biocompatibility. Presently, galvanic silver coating finds widespread clinical application in mitigating infection risks associated with large joint arthroplasties. While some instances have linked this coating to sporadic cases of localized argyria, these occurrences have not exhibited systematic or functional limitations. To address concerns regarding biocompatibility, a novel approach has been devised for anti-infective implant coatings: encapsulating silver nitrate within a biopolymer reservoir for non-articulating surfaces. This poly-L-lactic acid layer releases silver ions gradually, thereby circumventing biocompatibility concerns. Method. Female C57BL/6 mice were utilized as an experimental model, with 6x2 mm Ti6Al4V discs, coated with or without the biopolymer-protected silver coating, implanted subcutaneously on both sides of the vertebrae. Daily blood samples were collected, and serum was analyzed for C-reactive protein (CRP) and silver concentration. After three days, histopathological analyses were conducted on the surrounding soft tissue pouch. Results. Maximum CRP levels in the silver group (4.80 mg/L; Median: 3.29 mg/L; IQR: 2.38 to 3.73) did not significantly differ from the control group (4.58 mg/L; Median: 2.93 mg/L; IQR: 1.91 to 3.78) over the study period. Silver levels in serum 24 hours post-implantation were 64 µg/L (IQR: 35 to 78) and decreased subsequently over three days to 23 µg/L (IQR: 13 to 28). Histopathological examinations revealed a similarly strong expression of inflammation signs in tissue samples from the two groups. Conclusions. Despite evidence of local inflammation indicated by CRP and histopathological analysis, no significant difference was observed between the coated and uncoated groups. This suggests that any inflammation may be attributed to the implantation procedure rather than silver influence. Furthermore, silver levels remained below the toxic limit, indicating the efficacy of the biopolymer-protected reservoir in aiding biocompatibility. This study underlines the potential of biopolymer-protected silver reservoirs in enhancing the safety profile of anti-infective silver implant coatings, warranting further investigation into their clinical application

Aim. The use of medical devices has grown significantly over the last decades, and has become a major part of modern medicine and our daily life. Infection of implanted medical devices (biomaterials), like titanium orthopaedic implants, can have disastrous consequences, including removal of the device. For still not well understood reasons, the presence of a foreign body strongly increases susceptibility to infection. These so-called biomaterial-associated infections (BAI) are mainly caused by Staphylococcus aureus and Staphylococcus epidermidis. Formation of biofilms on the biomaterial surface is generally considered the main reason for these persistent infections, although bacteria may also enter the surrounding tissue and become internalized within host cells. To prevent biofilm formation using a non-antibiotic based strategy, we aimed to develop a novel permanently fixed antimicrobial coating for titanium devices based on stable immobilized quaternary ammonium compounds (QACs). Method. Medical grade titanium implants (10×4×1 mm) were dip-coated in a solution of 10% (w/v) hyperbranched polymer, subsequently in a solution of 30% (w/v) polyethyleneimine and 10 mM sodium iodide, using a dip-coater, followed by a washing step for 10 min in ethanol. The QAC-coating was characterized using water contact angle measurements, scanning electron microscopy, FTIR, AFM and XPS. The antimicrobial activity of the coating was evaluated against S. aureus strain JAR060131 and S. epidermidis strain ATCC 12228 using the JIS Z 2801:2000 surface microbicidal assay. Lastly, we assessed the in vivo antimicrobial activity in a mouse subcutaneous implant infection model with S. aureus administered locally on the QAC-coated implants prior to implantation to mimic contamination during surgery. Results. Detailed material characterization of the titanium samples showed the presence of a homogenous and stable coating layer at the titanium surface. Moreover, the coating successfully killed S. aureus and S. epidermidis in vitro. The QAC-coating strongly reduced S. aureus colonization of the implant surface as well as of the surrounding tissue, with no apparent macroscopic signs of toxicity or inflammation in the peri-implant tissue at 1 and 4 days after implantation. Conclusions. An antimicrobial coating with stable quaternary ammonium compounds on titanium has been developed which holds promise to prevent BAI. Non-antibiotic-based antimicrobial coatings have great significance in guiding the design of novel antimicrobial coatings in the present, post-antibiotic era

Aim. Rifampicin and fluoroquinolone based therapy is generally considered as first-choice targeted oral antimicrobial therapy for staphylococcal prosthetic joint infections (PJI) treated with debridement, antibiotics and implant retention (DAIR). Alternative equally effective antimicrobial strategies are urgently needed due to toxicity and drug-drug interactions that frequently occur with this strategy. Data from recent clinical studies suggests equipoise for other antimicrobial treatment regimens. The objective of the Rifampicin Combination Therapy versus Targeted Antimicrobial Monotherapy in the Oral Antimicrobial Treatment Phase of Staphylococcal Prosthetic Joint Infection (RiCOTTA)-trial is to evaluate whether monotherapy with clindamycin is non-inferior to rifampicin/fluoroquinolone combination therapy in patients with staphylococcal PJI that are treated with DAIR. Method. The RiCOTTA-trial is a multicenter, non-inferiority, open-label, randomized controlled trial evaluating clindamycin versus rifampicin/fluoroquinolone combination therapy in the oral treatment phase in patients with staphylococcal PJI managed with DAIR. The trial is performed in 16 hospitals in the Netherlands. Eligible patients are adults with staphylococcal knee or hip PJI managed by DAIR. Patients are included one to six days before antibiotic treatment is switched from intravenous to oral therapy. Patients with a contraindication for rifampicin, with a megaprosthesis or who receive intravenous antibiotics for more than three weeks after initial debridement are excluded. Primary outcome is treatment success one year after finishing antimicrobial treatment. Success is defined as the absence of: i. Infection related re-surgery, ii. New episode of antibiotic treatment for infection of the index joint after the initial treatment phase of 12 weeks, iii. Ongoing use of antibiotics for the index joint at the end of follow-up, iv. Death. The estimated treatment success of rifampicin combination therapy is 85% and the monotherapy strategy is considered not inferior when the difference in treatment success will be less than 10%. Enrolment of 158 patients per group (316 in total) is needed to confirm non-inferiority of monotherapy with a power of 80%. The trial is currently open for enrolment. The study is approved by the Medical Ethics Committee Leiden, the Hague, Delft, the Netherlands and registered under EU trial number 2022-501620-26-00 in Clinical Trial Information System. Conclusions. Currently, the RiCOTTTA study is the largest randomised clinical trial that compares targeted oral monotherapy with rifampicin combination treatment for staphylococcal PJI. Noninferiority of monotherapy would result in a change in national PJI guidelines and enable clinicians to use a more patient-tailored approach when considering antibiotics for patients during the oral treatment phase of PJI

Aim. Prosthetic joint infections pose a major clinical challenge. Developing novel material surface technologies for orthopedic implants that prevent bacterial adhesion and biofilm formation is essential. Antimicrobial coatings applicable to articulating implant surfaces are limited, due to the articulation mechanics inducing wear, coating degradation, and toxic particle release. Noble metals are known for their antimicrobial activity and high mechanical strength and could be a viable coating alternative for orthopaedic implants [1]. In this study, the potential of thin platinum-based metal alloy coatings was developed, characterized, and tested on cytotoxicity and antibacterial properties. Method. Three platinum-based metal alloy coatings were sputter-coated on medical-grade polished titanium discs. The coatings were characterized using optical topography and scanning electron microscopy with energy dispersive spectroscopy (SEM/EDS). Ion release was measured using inductively coupled plasma optical emission spectrometry (ICP-OES). Cytotoxicity was tested according to ISO10993-5 using mouse fibroblasts (cell lines L929 and 3T3). Antibacterial surface activity, bacterial adhesion, bacterial proliferation, and biofilm formation were tested with gram-positive Staphylococcus aureus ATCC 25923 and gram-negative Escherichia coli ATCC 25922. Colony forming unit (CFU) counts, live-dead fluorescence staining, and SEM-EDS images were used to assess antibacterial activity. Results. Three different platinum-based metal alloys consisting of platinum-iridium, platinum-copper, and platinum-zirconium. The coatings were found 80 nm thick, smooth (roughness average < 60 nm), and non-toxic. The platinum-copper coating showed a CFU reduction larger than one logarithm in adherent bacteria compared to uncoated titanium. The other coatings showed a smaller reduction. This data was confirmed by SEM and live-dead fluorescence images, and accordingly, ICP-OES measurements showed low levels of metal ion release from the coatings. Conclusions. The platinum-copper coating showed low anti-adhesion properties, even with extremely low metal ions released. These platinum-based metal alloy coatings cannot be classified as antimicrobial yet. Further optimization of the coating composition to induce a higher ion release based on the galvanic principle is required and copper looks most promising as the antimicrobial compound of choice. Acknowledgments. This publication is supported by the DARTBAC project (with project number NWA.1292.19.354) of the research program NWA-ORC which is (partly) financed by the Dutch Research Council (NWO); and the AMBITION project (with project number NSP20–1-302), co-funded by the PPP Allowance made available by Health-Holland, Top Sector Life Sciences & Health to ReumaNederland

Aim. Antibacterial activity of coatings based on metal and metal oxide nanoparticles (NPs) often depends on materials and biotic targets resulting in a material-specific killing activity of selected Gram-positive and Gram-negative bacteria, including drug-resistant strains. In this perspective, the NPs loading amount, the relative elemental concentration inside the nanogranular building blocks and the deposition method are of paramount importance when the goal is to widen the antimicrobial spectrum, but at the same time to avoid high levels of metal content to limit undesired toxic effects. Aim of the present study was evaluation of the antimicrobial properties of two multielement nanogranular coatings composed of Titanium-Silver and Copper and of Magnesium-Silver and Copper. Method. Ti-Ag-Cu and Mg-Ag-Cu NPs were deposited on circular cover glasses (VWR) by Supersonic Cluster Beam Deposition. Biofilm-producer strains of Staphylococcus aureus (methicillin susceptible and resistant), Staphylococcus epidermidis (methicillin susceptible and resistant), Escherichia coli (fully susceptible and producer of extended spectrum beta lactamases), and Pseudomonas aeruginosa (susceptible and multidrug-resistant) were selected. The abilities of the selected strains to adhere, colonize and produce biofilm on the discs coated with Ti-Ag-Cu or Mg-Ag-Cu NPs were compared to uncoated circular cover glasses which were used as growth control. Cytotoxicity was also evaluated in order to assess the biocompatibility of the newly synthesized NPs. Results. In comparison to uncoated controls, both coatings showed significant anti-adhesive properties against S. aureus, S. epidermidis, and E. coli. Reduction in adhesion to Mg-Ag-Cu coated discs was observed also for P. aeruginosa isolates, although differences vs uncoated controls did not reach statistical significance. Biofilm formation was reduced on discs coated with Mg-Ag-Cu compared to Ti-Ag-Cu and, again, coatings had a milder effect on P. aeruginosa, probably due to its exceptional capability of attachment and matrix production. These results were confirmed by the evaluation of bacterial colonization on nanoparticles-coated discs by means of confocal laser scanning microscopy. A viability of 95.8% and 89.4% of cells cultured in the presence of Ti-Ag-Cu and Mg-Ag-Cu discs, respectively, when compared to negative controls was observed, thus excluding cytotoxic effects on eukaryotic cells. Conclusions. The newly synthesized Ti-Ag-Cu and Mg-Ag-Cu coatings are able to limit bacterial adhesion colonization and biofilm production, thus highlighting the safe use of multi-element families of NPs as new strategies against bacterial attachment to the surface of biomedical implants. However, further studies addressing activity against P. aeruginosa and including a wide number of isolates are warranted

Aim. The use of bone substitutes such as calcium sulfate (CaSO4) and hydroxyapatite with local antibiotics are crucial in the treatment of osteomyelitis. They allow the treatment of the dead space and locally provide large concentrations of antibiotics. However, it is unknown whether use of local vancomycin may elute and influence on vancomycin plasma levels. The aim of this study is to assess whether the addition of vancomycin to CaSO4 with hydroxyapatite may increase vancomycin plasma concentrations in in patients with osteomyelitis and therefore alter dosage adjustments. Method. The present study investigates the vancomycin plasma concentrations at 72–94 h post-surgery after the application of local vancomycin within CaSO4 (660mg vancomycin/10cc) and hydroxyapatite bone substitute in patients treated with empiric intravenous vancomycin and surgically treated for osteomyelitis. Vancomycin plasma concentrations were analyzed in twelve patients with osteomyelitis surgically treated with local release of vancomycin by CaSO4 and hydroxyapatite and undergoing therapeutic drug monitoring (TDM) of their vancomycin plasma concentrations as it is routinely done in our hospital. From 2019 to 2022, demographic data, microbiology, type of osteomyelitis, amount of local vancomycin applied, alteration of renal function, and vancomycin levels were retrospectively analyzed. Results. Twelve patients were included: 9(75%) were men. Median (range) demographic and clinical data: age: 51(26–67) years; body mass index: 27.7(18–46.4) kg/m2;baseline serum creatinine: 0.85 (0.7–1.24)mg/dl and 5(41.7%) with and glomerular filtration rate < 90ml/min(CPD-EPI, ml/min). Most frequently isolated microorganisms were Staphylococci (58%). Seven (54%) patients were classified as Cierny-Mader Osteomyelitis type III, 3(23%) as type IV and 2(23%) as type I. Treatment data: initial dose of vancomycin: 1g/8h in 9(75.0%) and 1g/12h in 3(25%) patients, total daily dose/body weight: 35.3(15.9–46.2) mg/kg. Pharmacokinetic data:days of iv vancomycin treatment until first TDM measurement: 3(3–4) days; minimum and maximum vancomycin plasma concentrations: 9.4(3–17.3) mg/L and 19.6(11.3–33.4) mg/L, respectively; patients with therapeutic concentrations: 6(50%); infratherapeutic: 4(33.3%) and supratherapeutic/potentially toxic: 2(16.7%). These 2 patients were young, had a baseline conserved renal function and were receiving the higher dose of 1g/8h. Conclusions. Vancomycin incorporated into the bone substitute appears not to increase blood concentrations of the glycopeptide in patients with osteomyelitis treated surgically and with intravenous vancomycin. However, 2 of the 12 patients presented supratherapeutic and potentially nephrotoxic vancomycin concentrations in the first TDM measurement, even though they were young and without renal impairment and needed and unexpected dose reduction. These results suggest the need to confirm the safety of local vancomycin in further larger clinical studies

Aim. Vancomycin is frequently used for bone and joint infections (BJI) because of the main role of Gram-positive bacteria as potential causal agents. It is crucial to achieve optimal vancomycin plasma concentrations since the first day to maximize treatment clinical and microbiological efficacy. The aim was to describe the patients’ profile that are more likely to achieve an optimal pharmacokinetic/pharmacodynamics (PK/PD) vancomycin target in the first therapeutic drug monitoring (TDM) sample. Methods. Retrospective study (March 2018-January 2022) in a university hospital including all patients treated with vancomycin for a BJI and undergoing TDM. Initial dose (1g/8-12h) was selected by the responsible clinician. Vancomycin plasma concentrations were obtained pre-dose (Cmin,ss) and 60-minutes after the infusion on day 2 of treatment. Global exposure measured by the area under the curve of plasma concentrations during 24h (AUC024h) was estimated using a bicompartmental PK model. An AUC024h/CMI=400–600mg*h/L was considered optimal, <400 infratherapeutic and >600 supratherapeutic, based on recent guidelines, and patients were classified into these 3 groups. A value of CMI=1 mg/L was considered, following guidelines recommendations. Categorial data: percentages and quantitative data as mean (standard deviation). Results. Ninety-five patients were included: 22(23.2%), 43(45.3%) and 30(31.6%) presented an infratherapeutic, optimal and supratherapeutic PKPD target, respectively. Medium age was 75,8(13,5) in the supratherapeutic group versus 57,2(16,3) in the infratherapeutic group. Weight (kg) was higher in the infratherapeutic group 80,8(18,4) versus 66,8(15,5) in the supratherapeutic group. Vancomycin dose (mg/kg/d) was 43,5(12,4) in the supratherapeutic group versus 34,5(10,8) in the infratherapeutic group. There were 17(56,7) patients who received 1g/8h of vancomycin in the supratherapeutic group and 6 (27,3) in the infratherapeutic group. Baseline glomerular filtration rate (BGF (CKD-EPI) (mL/min/1.73m2) was 71,5(20,1) in the supratherapeutic group and 100,0 (19,9) in the infratherapeutic group. The AUC24h/CMI was 788,0(186,1) in the supratherapeutic group and 323,7(55,4) in the infratherapeutic group. Significant differences observed in age, body weight (BW), baseline renal function and dose/frequency of vancomycin. Dosage adjustments recommendations were made in 62(65.3%) patients: 31(32.6%) dose-increase, 29(30.5%) reduction and 2 cases (2.1%) a temporary suspension. Conclusions. Less than 50% of patients achieved an optimal exposure of vancomycin on day 2 of treatment. Patients with infratherapeutic levels had a younger age and a higher body weight and glomerular filtration rate. In addition, they had received a lower vancomycin initial dose. On the contrary, a potentially toxic exposure was observed within older patients with impaired baseline renal function. These data suggest the relevance of an early vancomycin TDM for optimizing the treatment of BJI

The burden of periprosthetic joint infection (PJI) continues to rise and the management of this dreaded complication continues to pose challenges to the orthopaedic community. Dr Buchholz from the Endo Klinik has been credited for reporting the initial observation that addition of antibiotic to polymethylmethacrylate (PMMA) cement lead to better ability to deliver higher concentrations of antibiotic to the joint milieu and avoid administration of high doses of systemic antibiotics with potential for systemic toxicity. Addition of antibiotics to PMMA cement has continued to be an important aspect of managing patients with chronic PJI. The rationale for this practice is that higher doses of local antibiotics can be reached without placing the patients at risk of systemic toxicity. Whether a one-stage or a two-stage exchange arthroplasty is being performed, antibiotics that can withstand the exothermic reaction of PMMA and are able to elude from cement are added at various doses to the PMMA for later delivery. Although this practice continues to be almost universal, there are a few unknowns. First of all, a recent study raised a valid question regarding this practice. Though intuitively logical, addition of antibiotics to PMMA spacers has not been scrutinised by any level 1 study and hence one is not able to prove that this practice does indeed accomplish its intended objectives of reducing recurrence or persistence of infection. Orthopaedic community is advised to seek avenues to generate this much-needed evidence. The other main unknown is how much, and in some instances which antibiotic, needs to be added to the PMMA cement. Some authorities have declared that antibiotics can be added at high doses, with an average total dose of 10.5 g of vancomycin (range, 3–16 g) and 12.5 g of gentamicin (range, 3.6–19.2 g) in one study, to PMMA cement without the fear of systemic toxicity. In recent years, renal toxicity and other systemic adverse effects have been attributed to addition of high doses of antibiotics to cement. I have personally witnessed such adverse reactions in a few patients. Although initially I was inclined to “blame” the concurrent administration of systemic antibiotics for the renal toxicity that patients developed following insertion of spacer, selective nephrotoxicity (i.e. reaction to aminoglycoside that was only present in the spacer and not systemically administered) and resolution of the nephrotoxicity upon removal of antibiotic spacer, convinced me that our nephrology colleagues have a valid reason to be concerned about addition of high doses of antibiotics to PMMA spacers. What has become clear is that high viscosity cements containing MA-MMA copolymers have been shown to have better antibiotic elution profiles than other PMMA formulations. So when fashioning a spacer in the operating room the surgeon needs to be aware of the differences in elution profile of antibiotics from PMMA and individualise the dose of antibiotics being added to spacer based on the type and viscosity of cement being used and the renal status of the patient. Thus, systemic toxicity caused by addition of antibiotics to cement spacer appears to be a real issue in some circumstances and this needs to be born in mind when managing patients with PJI. There are numerous other issues related to the use of antibiotic cement spacers. In the hip, the lack of adequate offset and limited portfolio of products result in laxity in the soft tissue and subsequent dislocation of the hip. In addition, the dose and type of antibiotic in the premanufactured spacers, at least in the US, are inadequate to lead to a substantial delivery of antibiotics in the local tissues. Because of these issues, I prefer to fabricate “customised” spacers for each patient that I operate on

Tungsten has been increasing in demand for use in manufacturing and recently, medical devices, as it imparts flexibility, strength, and conductance of metal alloys. Given the surge in tungsten use, our population may be subjected to elevated exposures. For instance, embolism coils made of tungsten have been shown to degrade in some patients. In a cohort of breast cancer patients who received tungsten-based shielding for intraoperative radiotherapy, urinary tungsten levels remained over tenfold higher 20 months post-surgery. In vivo models have demonstrated that tungsten exposure increases tumor metastasis and enhances the adipogenesis of bone marrow-derived mesenchymal stem cells while inhibiting osteogenesis. We recently determined that when mice are exposed to tungsten [15 ppm] in their drinking water, it bioaccumulates in the intervertebral disc tissue and vertebrae. This study was performed to determine the toxicity of tungsten on intervertebral disc. Bovine nucleus pulposus (bNP) and annulus fibrosus (bAF) cells were isolated from bovine caudal tails. Cells were expanded in flasks then prepared for 3D culturing in alginate beads at a density of 1×10. ∧. 6 cells/mL. Beads were cultured in medium supplemented with increasing tungsten concentrations in the form of sodium tungstate [0, 0.5, 5, 15 ug/mL] for 12 days. A modified GAG assay was performed on the beads to determine proteoglycan content and Western blotting for type II collagen (Col II) synthesis. Cell viability was determined by counting live and dead cells in the beads following incubation with the Live/Dead Viability Assay kit (Thermo Fisher Scientific). Cell numbers in beads at the end of the incubation period was determined using Quant-iT dsDNA Assay Kit (Thermo Fisher Scientific). Tungsten dose-dependently decreased the synthesis of proteoglycan in IVD cells, however, the effect was significant at the highest dose of 15 ug/mL. (n=3). Furthermore, although tungsten decreased the synthesis of Col II in IVD cells, it significantly increased the synthesis of Col I. Upregulation of catabolic enzymes ADAMTS4 and −5 were also observed in IVD cells treated with tungsten (n=3). Upon histological examination of spines from mice treated with tungsten [15 ug/mL] in their drinking water for 30 days, disc heights were diminished and Col I upregulation was observed (n=4). Cell viability was not markedly affected by tungsten in both bNP and bAF cells, but proliferation of bNP cells decreased at higher concentration. Surprisingly, histological examination of IVDs and gene expression analysis demonstrated upregulation of NGF expression in both NP and AF cells. In addition, endplate capillaries showed increases in CGRP and PGP9.5 expression as determined on histological sections of mouse IVDs, suggesting the development of sensory neuron invasion of the disc. We provide evidence that prolonged tungsten exposure can induce disc fibrosis and increase the expression of markers associated with pain. Tungsten toxicity may play a role in disc degeneration disease

Aim. Periprosthetic joint infection (PJI) is a devastating complication of total joint arthroplasty. While research has focused on developing better tests for disease diagnosis, treatment options have stayed relatively constant over the years with high failure rates ranging from 30%–50% and are due in part to the protective biofilm produced by some bacterial species. Current treatment options are compromised by the presence of biofilm, emphasizing the need for novel treatment strategies to be developed. Our group has developed a novel treatment (PhotothermAA) which has demonstrated in vitro its ability to target bacterial biofilm. The purpose of this study was to test this PhotothermAA technology in vivo in a rabbit model of PJI for its efficacy in eradicating biofilm. Method. Rabbits were fitted with a titanium implant into the tibial plateau and inoculated with 5×10. 6. CFU Xen36 (luminescent Staphylococcus aureus). At two weeks, rabbits underwent irrigation and debridement and treatment with PhotothermAA gel for two hours and subsequently laser heated using an 808 nm laser for 10 minutes. Gel was washed out and implant was removed for quantitative biofilm coverage analysis via scanning electron microscopy (SEM, n=3 for control and n=2 for PhotothermAA treated). Periprosthetic tissue was collected before and after treatment for toxicity studies via hemotoxylin and eosin (H&E) staining and scored for necrosis by three blinded reviewers (n=5 per group). Student's t-test was used for statistical analysis. Results. Implants isolated after PhotothermAA gel treatment had less biofilm coverage on the surface of the implant compared to non-treated control via SEM analysis (36.9% vs. 55.2%, p<0.14). PhotothermAA gel treatment and subsequent laser treatment was not harmful to surrounding tissue as no increase in necrotic tissue was observed. Conclusions. PhotothermAA gel and laser treatment safely decreases biofilm coverage on infected knee implants in a rabbit PJI model

Objectives. The purpose of this study was to evaluate in vivo biocompatibility of novel single-walled carbon nanotubes (SWCNT)/poly(lactic-co-glycolic acid) (PLAGA) composites for applications in bone and tissue regeneration. Methods. A total of 60 Sprague-Dawley rats (125 g to 149 g) were implanted subcutaneously with SWCNT/PLAGA composites (10 mg SWCNT and 1gm PLAGA 12 mm diameter two-dimensional disks), and at two, four, eight and 12 weeks post-implantation were compared with control (Sham) and PLAGA (five rats per group/point in time). Rats were observed for signs of morbidity, overt toxicity, weight gain and food consumption, while haematology, urinalysis and histopathology were completed when the animals were killed. Results. No mortality and clinical signs were observed. All groups showed consistent weight gain, and the rate of gain for each group was similar. All groups exhibited a similar pattern for food consumption. No difference in urinalysis, haematology, and absolute and relative organ weight was observed. A mild to moderate increase in the summary toxicity (sumtox) score was observed for PLAGA and SWCNT/PLAGA implanted animals, whereas the control animals did not show any response. Both PLAGA and SWCNT/PLAGA showed a significantly higher sumtox score compared with the control group at all time intervals. However, there was no significant difference between PLAGA and SWCNT/PLAGA groups. Conclusions. Our results demonstrate that SWCNT/PLAGA composites exhibited in vivo biocompatibility similar to the Food and Drug Administration approved biocompatible polymer, PLAGA, over a period of 12 weeks. These results showed potential of SWCNT/PLAGA composites for bone regeneration as the low percentage of SWCNT did not elicit a localised or general overt toxicity. Following the 12-week exposure, the material was considered to have an acceptable biocompatibility to warrant further long-term and more invasive in vivo studies. Cite this article: Bone Joint Res 2015;4:70–7

Aim. Fungal orthopaedic infections most commonly affect people with complex surgical histories and existing comorbidities. Recurrence and re-infection rates are high, even with optimal surgical and systemic antifungal treatment. AmBisome liposomal amphotericin B has been suggested for local antifungal therapy, as an adjunctive treatment for fungal osteoarticular infections. Few case series have examined its clinical use when combined with polymethylmethacrylate cement PMMA), or with absorbable local antibiotic carriers. We aimed to evaluate the clinical use of local antifungal therapy with AmBisome liposomal amphotericin B (ABlaB), including tolerated doses, serious adverse events, and treatment outcomes. Method. A retrospective cohort of all patients treated with local antifungal therapy with ABlaB between January 2016 and January 2021 in a specialist orthopaedic hospital was identified using pharmacy records. Renal function, serious adverse events during treatment, surgical outcomes including spacer fracture and infection recurrence, were identified from electronic clinical records. The project was approved by the Institutional Review Board (clinical audit 6871). Results. 13 operations involving local antifungal therapy with ABlaB, in 12 patients, were identified. Eleven were infected with Candida species and one with Aspergillus. Mean follow-up was 22 months (range 4–46). Ten first stage arthroplasty revisions, 2 second stage arthroplasty revisions, and one debridement and removal of metalwork for fracture-related infection were performed. Locally implanted doses of ABlaB ranged from 100mg to 3600mg (50–400mg per 40g mix of PMMA). Six patients received ABlaB in absorbable antibiotic carriers containing calcium sulphate. This was noted to delay carrier setting. Patients were also given systemic antifungal therapy. No patients experienced serious adverse events related to toxicity from local antifungal therapy with ABlaB. There were no spacer fractures. Overall treatment success was 54% at final follow-up, although there were no recurrent fungal infections identified in patients experiencing treatment failure. Conclusions. Local antifungal therapy with liposomal amphotericin B, when combined with surgery and systemic therapy, appears to be a safe and well tolerated intervention in the management of complex fungal osteoarticular infections

Aim. Periprosthetic joint infection (PJI) is a feared complication of total joint arthroplasty of hip (THA) or knee (TKA). Debridement, antibiotic treatment, and implant retention (DAIR) is an effective treatment of early PJI. In the Netherlands, cefazolin resistance in early PJI after primary arthroplasty is low. Little is known about causative micro-organisms and resistance patterns in PJI after revision arthroplasty. No recommendations for empirical treatment are described in the current guidelines. The aim of this study is to describe the characteristics of PJI after revision arthroplasty and to evaluate whether the used empirical treatment regimens are adequate, based on microbiology data. Method. In this retrospective study we included patients with early PJI after aseptic revision of THA or TKA, treated with DAIR between 2012 and 2020. Success rate was defined as implant retention and no persistent or recurrent infection during one year follow-up. Results. We identified 96 patients with PJI. PJI was most frequently caused by Staphylococcus spp. (n=73), Gram-negative bacilli (n=31) or Enterococcus spp. (n=13). Polymicrobial infection was diagnosed in 38 PJIs. Mismatches were present in 72 (75%) of the PJIs (95% CI: 0.66–0.84). Table 1 shows the number of mismatches per empirical treatment regimen. Figure 1 shows the responsible micro-organisms for the mismatches. Success rate of PJI treatment was significant reduced for patients with mismatching compared to matching empirical therapy: 62% vs. 95% respectively (OR: 0.09, 95% CI: 0.01–0.68, p=0.004). If vancomycin would have been the empirical treatment, mismatches would have been reduced to 31 (32%) (95% CI: 0.23–0.42). With vancomycin-ciprofloxacin combination therapy the mismatches would have been reduced to 1% (95% CI: −0.01–0.03). Conclusions. There is a high number of mismatches in empirical treatment in early PJI after revision arthroplasty, which have significant influence on the outcome. Based on our data cefazolin should not be recommended as empirical treatment for this specific group. Our data shows that review of local data is necessary to improve treatment strategies, that eventually might improve outcome. Besides changing Gram-positive coverage, a prospective study is needed to assess the benefits of broader spectrum empiric antimicrobial treatment taken into account toxicity and other side effects such as antimicrobial resistance. For any tables or figures, please contact the authors directly