Aim. Biomaterial-associated infections (BAI) present a formidable clinical challenge. Bioactive glasses (BG) have proven highly successful in diverse clinical applications, especially in dentistry and orthopaedics. In this study, we aimed to determine the effect of three commonly used BG composition and particle sizes on cell and bacterial attachment and growth. Our focus is on understanding the changes in pH and osmotic pressure in the surrounding environment during glass degradation. Method. First, three different melt-derived glasses were characterized by analyzing particle size and glass network structure using Raman and NMR. The different glasses were then tested in vitro by seeding 4x 10. 4. cells/well (SaOS Cell line) in a 48 well plate. After a pre-incubation period of 72 hours, the different BGs and particle sizes were added to the cells and the pH value, ion release and live/dead staining was measured every hour. The effect of BG against bacteria (S. epidermidis) was analyzed after 24 and 72 hours of treatment by using XTT viability assay and CFU counting by plating out the treated aliquot agar to estimate the viable bacteria cells. Results. All three BG compositions tested showed a significant increase in pH, which was highest in BG composition 45S5 with a value of 11 compared to the other BG compositions 10 and 9 in S53P4 and 13-93 respectively. This strong increase in the pH in all BG samples tested results in a strongly reduced cell viability rate of more than 75% compared to the untreated control and 6-fold reduction in bacterial viability compared to the untreated control. The live/ dead assay also showed an increased cell viability with increasing glass particle size (i. e smallest glass particle < 25% viable cell and largest glass particle> 65% viable cell). The ion release concentration over 50 h showed an increase in sodium ions to 0.25 mol/L, calcium to 0.003 mol/L and a decrease in phosphorus. Conclusions. These results show that the composition of the bioactive glass and the choice of particle size have a major influence on subsequent applications. In addition to the different compositions of the BG, particle size and additional medium change also influence the pH and ion release, and therefore also on cells or bacteria viability. The sizes of the bioactive glass particle are inversely proportional to it. Further tests are necessary to develop custom design BG compositions, which simultaneously stimulate osteoblasts proliferation and prevent microbial adhesion

Osteomyelitis is an infectious process in bone occasionally leading to bone destruction. Traditionally a two-stage operation is performed using PMMA + antibiotic beads or a spacer. In the second operation the void filler is removed and the defect is filled with autologous bone. Bioactive glass (BAG) S53P4 is an antibacterial biodegradable bone substitute. This feature is based on an increase in pH and the osmotic pressure around the BAG, a phenomenon which has been shown to kill both planktonic bacteria and bacteria in biofilm in-vitro. We analyzed retrospectively our early results of osteomyelitis patients treated with BAG from the patient's clinical history. The diagnosis was stated in addition to bacterial samples by MRI, CT and plain radiographs or by a combination of these. Between 2007–2013 we applied BAG as a void-filler in 20 cases (15 male and 5 female) of osteomyelitis in the lower (19) or the upper (1) limb in one-stage procedure. The patients had been suffering from symptoms of osteomyelitis a mean 3,5 months (0,25–24,00) and had a history of mean 3,5 (1–11) earlier operations. Osteomyelitis was estimated to be healed when the enclosed systemic antibiotic treatment and clinical controls were carried out and the patient didn't have symptoms of a persisting disease. The average postoperative follow up was 7,8 (3,0–59,0) months. Fifteen (75%) of the patients healed. One patient run out of controls, but was symptom free during his last visit. In four cases we had to remove the bioactive glass because of continuous secretion. In three cases the debridement was incomplete and one had a poor soft tissue cover and a candida infection. Adjuvant systemic antibiotic treatment was prescribed postoperatively 7,3 (4–19) weeks. Bioactive glass is an effective void filling material in the treatment of osteomyelitis. Proper debridement and a soft tissue cover should be performed. Main reason for that the five patients did not heal is, that this procedure is new and we were looking for the right indications and techniques

Aim. We aimed to compare the in vitro antibacterial activity of Bioactive Glass (BAG) S53P4, which is a compound showing local antibacterial activity, to that of antibiotic-loaded polymethylmethacrylate (PMMA) against multidrug resistant bacteria from osteomyelitis (OM) and prosthetic joint infection (PJI) isolates. Method. We studied convenience samples of multidrug resistant (MDR) microorganisms obtained from patients presenting OM and prosthetic joint infection (PJI). Mixtures containing tryptic soy broth (TSB) and inert glass beads (2mm), BAG-S53P4 granules (0.5–0.8mm and <45 mm) and Gentamicin or Vancomycin-loaded PMMA beads were inoculated with methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-resistant coagulase-negative Staphylococcus (MR-CoNS), Pseudomonas aeruginosa or Klebsiella pneumoniae isolates. Glass beads (2.0mm) were used as a control. Antibacterial activity was evaluated by means of time-kill curve, through seeding the strains on blood agar plates, and subsequently performing colony counts after 24, 48, 72, 96, 120 and 168 hours of incubation. Differences between groups were evaluated by means of two-way analysis of variance (ANOVA) and Bonferroni's t test. Results. Inhibition of bacterial growth started soon after 48 hours of incubation, reached zero CFU/ml between 120 and 168 hours of incubation for both antibiotic-loaded PMMA and BAG S53P4 groups, in comparison with inert glass (p< 0.05). No difference regarding time-kill curves between antibiotic-loaded PMMA and BAG S53P4 was observed. Moreover, despite no difference was observed between both Vancomycin - or Gentamicin-loaded PMMA and BAG groups, there was statistical difference between the effectiveness of all treatments (BAG included) against gram-positive cocci and gram-negative bacilli, the latter of which requiring longer time frames for the cultures to yield no bacterial growth. Conclusions. BAG S53P4 presented antibacterial properties as much as antibiotic-loaded PMMA for MDR bacteria producing OM and PJI, although presenting differences between its effectiveness against different bacterial groups

Introduction

Significant donor site morbidity (3–61%) has been associated with tri-cortical iliac crest bone graft harvesting and reconstruction of the defect has been shown to reduce it. Chitra-HABG (Chitra-Hydroxyapatite-Bio-active glass ceramic composite) is an indigenously developed ceramic which has been evaluated as a bone graft substitute.

Introduction. Various biomaterials and bone graft substitute technologies for use in osteomyelitis treatment are currently used in clinal practice. They vary in mode of action (with or without antibiotics) and clinical application (one-stage or two-stage surgery). This systematic review aims to compare the clinical evidence of different synthetic antimicrobial bone graft substitutes and antibiotic-loaded carriers in eradicating infection and clinical outcome in patients with chronic osteomyelitis. Methods. Systematic review according to PRISMA statement on publications 2002-2023. MESH terms: osteomyelitis and bone substitutes. FREE terms: chronic osteomyelitis, bone infection. A standardized data extraction form was be used to extract data from the included papers. Results. Publications with increased methodological quality and clinical evidence for biomaterials in osteomyelitis treatment were published in the last decades. High 85-95% eradication rates of osteomyelitis were observed for various resorbable Ca-P and/or Ca-S biomaterials combined with antibiotics and S53P4 bioactive glass. Level of evidence varies significantly between products. Antibiotic pharmacokinetic release profiles vary between resorbable Ca-P and/or Ca-S biomaterials. Conclusion. Given the high 85-95% eradication rates of osteomyelitis by various resorbable Ca-P and/or Ca-S biomaterials combined with antibiotics and S53P4 bioactive glass, one-stage treatment is preferred. Surgeons should be aware of variations in mechanical properties and antibiotic pharmacokinetic release profiles between Ca-P and CA-s products. Mechanical, biological and antimicrobial properties of bioactive glass are formulation dependent. Currently, only S53P4 bioactive glass has proven antimicrobial properties. Based on this systematic review antibiotic loaded fleeces should be used with caution and restraint

Biomaterials used in regenerative medicine should be able to support and promote the growth and repair of natural tissues. Bioactive glasses (BGs) have a great potential for applications in bone tissue engineering [1, 2]. As it is well known BGs can bond to host bone and stimulate bone cells toward osteogenesis. Silicate BGs, e.g. 45S5 Bioglass® (composition in wt.%: 45 SiO. 2. , 6 P. 2. O. 5. , 24, 5 Na. 2. O and 24.5 CaO), exhibit positive characteristics for bone engineering applications considering that reactions on the material surface induce the release of critical concentrations of soluble Si, Ca, P and Na ions, which can lead to the up regulation of different genes in osteoblastic cells, which in turn promote rapid bone formation. BGs are also increasingly investigated for their angiogenic properties. This presentation is focused on cell behavior of osteoblast-like cells and osteoclast-like cells on BGs with varying sample geometry (including dense discs for material evaluation and coatings of highly porous Al. 2. O. 3. -scaffolds as an example of load-bearing implants). To obtain mechanically competent porous samples with trabecular architecture analogous to those of cancellous bone, in this study Al. 2. O. 3. scaffolds were fabricated by the well-known foam replication method and coated with Bioglass® by dip coating. The resulted geometry and porosity were proven by SEM and μCT. Originating from peripheral blood mononuclear cells formed multinucleated giant cells, i.e. osteoclast-like cells, after 3 weeks of stimulation with RANKL and M-CSF. Thus, the bioactive glass surface can be considered a promising material for bone healing, providing a surface for bone remodeling. Osteoblast-like cells and bone marrow stromal cells were seeded on dense bioactive glass substrates and coatings showing an initial inhibited cell attachment but later a strong osteogenic differentiation. Additionally, cell attachment and differentiation studies were carried out by staining cytoskeleton and measuring specific alkaline phosphatase activity. In this context, 45S5 bioactive glass surfaces can be considered a highly promising material for bone tissue regeneration, providing very fast kinetics for bone-like hydroxyapatite formation (mineralization). Our examinations revealed good results in vitro for cell seeding efficacy, cell attachment, viability, proliferation and cell penetration onto dense and porous Bioglass®-coated scaffolds. Recent in vivo investigations [3] have revealed also the angiogenic potential of bioactive glass both in particulate form and as 3D scaffolds confirming the high potential of BGs for bone regeneration strategies at different scales. Implant surfaces based on bioactive glasses offer new opportunities to develop these advanced biomaterials for the next generation of implantable devices and tissue scaffolds with desired tissue-implant interaction

Aim. The rise of multidrug-resistant bacteria and the decreasing efficacy of antibiotic therapy in successfully treating biofilm-associated infections are prompting the exploration of alternative treatment options. This study investigates the efficacy of different bioactive glass (BAG) formulations - alone or combined with vancomycin - to eradicate biofilm. Further, we study the influence of BAG on pH and osmotic pressure as important factors limiting bacterial growth. Method. Different BAG-S53P4 formulations were used for this study, including (a) BAG-powder (<45 μm), (b) BAG-granules (500–800 μm), (c) a cone-shaped BAG-scaffold and (d) two kinds of BAG-putty containing granules, with no powder (putty-A) or with additional powder (putty-B), and a synthetic binder. Inert glass beads were included as control. All formulations were tested in a concentration of 1750 g/ml in Müller-Hinton-Broth. Targeted bacteria included methicillin-resistant Staphylococcus aureus (MRSA) and epidermidis (MRSE). Vancomycin was tested at the minimum-inhibitory-concentration for each strain (1 µg/ml for MRSA; 2 μg/ml for MRSE). To investigate the antibiofilm effect of BAG alone or combined with vancomycin, 3 hour-old MRSA or MRSE biofilms were formed on porous glass beads and exposed to BAG ± vancomycin for 24h, 72h and 168h. After co-incubation, biofilm-beads were deep-washed in phosphate-buffered saline and placed in glass vials containing fresh medium. Recovering biofilm bacteria were detected by measuring growth-related heat production at 37°C for 24h by isothermal microcalorimetry. Changes in pH and osmotic pressure over time were assessed after co-incubation of each BAG formulation in Müller-Hinton-Broth for 0h, 24h, 72h and 168h. Results. All BAG formulations showed antibiofilm activity against MRSA and MRSE in a time-dependent manner, where longer incubation times revealed higher antibiofilm activity. BAG-powder and BAG-putty-B were the most effective formulations suppressing biofilm, followed by BAG-granules, BAG-scaffold and finally BAG-putty-A. The addition of vancomycin had no substantial impact on biofilm suppression. An increase in pH and osmotic pressure over time could be observed for all BAG formulations. BAG-powder reached the highest pH value of 12.5, whereas BAG-putty-A resulted in the lowest pH of 9. Both BAG-putty formulations displayed the greatest increase on osmotic pressure. Conclusions. BAG-S53P4 has demonstrated efficient biofilm suppression against MRSA and MRSE, especially in powder-containing formulations. Our data indicates no additional antibiofilm improvement with addition of vancomycin. Moreover, high pH appears to have a larger antimicrobial impact than high osmolarity. Acknowledgements. This work was supported by PRO-IMPLANT Foundation (Berlin, Germany). The tested materials were provided by Bonalive Biomaterials Ltd (Turku, Finland)

Despite the increasing availability of bone grafting materials, the regeneration of large bone defects remains a challenge. Especially infection prevention while fostering regeneration is a crucial issue. Therefore, loading of grafting material with antibiotics for direct delivery to the site of need is desired. This study evaluates the concept of local delivery using in vitro and in vivo investigations. We aim at verifying safety and reliability of a perioperative enrichment procedure of demineralized bone matrix (DBM) with gentamicin. DBM (DBMputty, DIZG, Germany) was mixed with antibiotic using a syringe with an integrated mixing propeller (Medmix Systems, Switzerland). Gentamicin, as powder or solution, was mixed with DBM at different concentrations (25 −100 mg/g DBM), release and cytotoxicity was analyzed. For in vivo analysis, sterile drill hole defects (diameter: 6 mm, depth: 15 mm) were created in diaphyseal and metaphyseal bones of sheep (Pobloth et al. 2016). Defects (6 – 8 per group and time point) were filled with DBM or DBM enriched with gentamicin (50 mg/g DBM) or left untreated. After three and nine weeks, defect regeneration was analyzed by µCT and histology. The release experiments revealed a burst release of gentamicin from DBM independent of the used amount, the sampling strategy, or the formulation (powder or solution). Gentamicin was almost completely released after three days in all set-ups. Eluates showed an antimicrobial activity against S. aureus over at least three days. Eluates had no negative effect on viability and alkaline phosphatase activity of osteoblast-like cells (partially published Bormann et al. 2014). µCT and histology of the drill hole defects revealed a reduced bone formation with gentamicin loaded DBM. After nine weeks significantly less mineralized tissue was detectable in metaphyseal defects of the gentamicin group. Histological evaluation revealed new bone formation starting at the edges of the drill holes and growing into the center over time. The amount of DBM decreased over time due to the active removal by osteoclasts while osteoblasts formed new bone. Using this mixing procedure, loading of DBM was fast, reliable and possible during surgical setting. In vitro experiments revealed a burst and almost complete release after three days, antimicrobial activity and good biocompatibility of the eluates. Gentamicin/DBM concentration was in the range of clinically used antibiotic-loaded-cement for prophylaxis and treatment in joint replacement (Jiranek et al. 2006). The delayed healing seen in vivo was unexpected due to the good biocompatibility found in vitro. A reduced healing was also seen in spinal fusion where DBM was mixed with vancomycin (Shields et al. 2017), whereas DBM with gentamicin or DBM/bioactive glass with tobramycin had no negative effect on osteoinductivity or femur defect healing, respectively (Lewis et al. 2010, Shields et al. 2016). In conclusion, loading of DBM with gentamicin showed a proper antibiotic delivery over several days, covering the critical phase shortly after surgery. Due to the faster and complete release of the antibiotic compared to antibiotic loaded cement, the amount of antibiotic should be much lower in the DBM compared to cement

Aim. The treatment of osteomyelitis often requires extensive surgical debridement and removal of all infected tissues and foreign bodies. Resulting bone loss can then eventually be managed with antibacterial bone substitutes, that may also serve as a regenerative scaffold. Aim of the present study is to report the clinical results of a continuous series of patients, treated at our centre with an antibacterial bioglass*. Method. From November 2010 to May 2016, a total of 106 patients, affected by osteomyelitis, were included in this prospective, single centre, observational study. Inclusion criteria were the presence of osteomyelitis with a contained bone defect or segmental defects < 10 mm, with adequate soft tissue coverage. All patients underwent a one-stage procedure, including surgical debridement and bone void filling with the bioactive glass*, with systemic antibiotic therapy and no local antibiotics. Clinical, radiographic and laboratory examinations were performed at 3, 6 and 12 months and yearly thereafter. Results. Two patients were lost to follow-up, hence a total of 104 patients (65 males, 39 females; mean age: 46 ± 17 years, min 6 – max 81) were available at an average follow-up of 38 ± 26 months (range: 12 – 68); forty-eight patients (46.1%) were classified as Type A, 48 (46.1%) as Type B and 8 (7.7%) as Type C hosts, according to McPherson classification. Tibia (N=61) and femur (N=33) were the most common involved bones. On average patients had undergone 2.1 ± 1.3 (min 0 – max 7) previous surgical operations, with a mean infection duration of 18.7 ± 16.6 months (min 2 – max 120). Infection recurrence was observed in 10 patients (9.6%), most often within one year from surgery (8/10). Negative prognostic factors included infection duration > 2 years, Gram negative or mixed flora or negative cultural examination, Type B or C hosts and soft tissue defect. No side effects or complications related to bioglass were noted. Conclusions. This is to our knowledge the longest and the largest single centre consecutive series of patients, affected by bone infections of the long bones, treated according to a one-stage procedure using bioactive glass. Our results confirm, on a larger population and at a longer follow-up, previous reports. Early treatment, pathogen identification and adequate management of soft tissues should be considered to further reduce infection recurrence rate. *BonAlive®

The aim of the study was to assess an effectiveness of S53P4 bioglass in reconstruction of postinflammatory bone loss. We have also evaluated wound healing after the surgical dead space management with use of the bioglass. A group of 7 patients with bone loss due to active osteomyelitis and with purulent fistula treated with use of S53P4 bioglass is presented in the paper. All the treated patients were male with mean of age 40,5 years. Mean time of an active inflammatory process with purulent discharge from the wound prior the surgery was 587 weeks. Wound healing pattern with an X-ray evaluation of reconstructed void was performed in postoperative period as well as in 1, 3, 6 and 12 mounth follow-up. In 6 out of 7 cases we did not observed any signs of infection recurrence in 1-year follow-up. Starting from 1-month follow-up inflammatory serum markers remained in their reference values. In all the successfully treated cases wound healing was assessed by two independent surgeons as excellent or good. Starting from 3-month follow-up we have observed gradual blurring of granular bioglass structure on an X-ray scans. In 1 complicated case we observed recurrence of septic inflammatory process with purulent fistula that required revision procedure with removal of the bioglass and extended debridement of inflammatory focus. In this case we have faced posttraumatic malunion of the femur that substantially complicated surgical access to the inflammatory focus during primary procedure. S53P4 bioactive glass is an effective solution in reconstruction of postinflammatory bone loss. Properties of this biomaterial efficiently prevent from focal infection recurrence by inhibiting of bacterial bone growth and reduction of dead space. The product requires however meticulous debridement and the access to a vital bone as a source of osteoblast cells. Underestimation of surgical debridement will likely result in reopening of the fistula due to reinfection. The study group requires further evaluation

Currently, no clinical options are available to prevent infections on uncemented orthopedic implants. Therefore we investigated the efficacy of DAC-hydrogel (disposable antibacterial coating(1), Novagenit, Italy) as carrier for various agents to prevent infections in an in vivo implant-model. Titanium rods were implanted in the left tibiae in New Zealand White rabbits. Prior to implantation, the implant bed was contaminated with 10∧5 colony forming units S. aureus. In the experimental groups, the hydrogel was loaded prior to be coated on the rods with: 2%(w/v) vancomycin (Van2 group, N=6), 5%(w/v) vancomycin (Van5 group, N=6), 10%(w/v) bioactive glass (BonAlive, Finland) (BAG group, N=6), which is antibacterial(2) and osteoconductive(3), or 0.5%(w/v) N-acetyl cysteine (NAC group, N=6), which inhibits bacterial growth and decreases biofilm formation(4). In the control group, empty hydrogel was applied (Gel group, N=12). Blood values were measured weekly. Following explantation on day 28, the anterior tibia was processed for bacterial culture. The posterior tibia and rod were used for measuring bone-implant contact using micro-CT and for histopathology. Results of the experimental groups were compared to the Gel group results. The blood values in the Van2 and Van5 groups were lower on day 7. Moreover, culture results demonstrated less animals with an infection in both groups at day 28. In accordance, these groups showed lower grades for infection. Further, the Van2 group demonstrated more bone-implant contact. These results suggest that infection was reduced in the Van2 and Van5 groups. In contrast, blood values, histological grades, and bone-implant contact of the BAG and NAC groups were comparable with the Gel group. These results suggest that infection was not prevented in the BAG and NAC groups. Local application of vancomycin-loaded DAC-hydrogel successfully reduced implant-related infections. Loading of the hydrogel with BAG or NAC did not prevent infection. It is possible that BAG in powder form, as used in the present study, dissolved before the antibacterial effect could take place. Instead, BAG granules may be a viable alternative. Next, it is possible that the NAC concentration was too low to prevent infections in an in vivo environment, although this concentration was proven effective in vitro for its antibacterial properties

Aims

The standard of wide tumour-like resection for chronic osteomyelitis (COM) has been challenged recently by adequate debridement. This paper reviews the evolution of surgical debridement for long bone COM, and presents the outcome of adequate debridement in a tertiary bone infection unit.