Antibiotic-laden bone cement is an important strategy of treatment for an established bone infection. It was aimed to find the safe antibiotic dose intervals of the antibiotic cements soaked in Phosphate Buffered Saline solution and to determine whether there was a difference in terms of mechanical strength between the prepared samples.

This study was done in our institute Microbiology and Metallurgy laboratories. All samples were prepared using manual mixing technique using 40 g radiopaque Biomet® Bone cement (Zimmer Biomet, Indiana, USA) under sterile conditions at 19 ± 2 ºC.

In this study, vancomycin (4 groups − 0.5, 2, 4, 6 g), teicoplanin (4 groups − 0.8, 1.2, 2, 2.4 g), daptomycin (4 groups − 1, 2, 2.5, 3 g), piperacillin-tazobactam (4 groups − 0.125, 0.5, 1, 2 g) and meropenem (4 groups − 0.5, 2, 4, 6 g) were measured in a assay balance and added to the cement powder. Antibiotic levels ranged from the lowest 0.625% to the highest 15%.

80×10×4 mm rectangle prism-shaped sample for mechanical measurements in accordance to ISO 5833 standart and 12×6×1 mm disc-shaped samples for microbiological assesments were used. Four sample for each antibiotic dose and control group was made. Prepared samples were evaluated macroscopically and faulty samples were excluded from the study. Prepared samples were kept in Phosphate Buffered Saline solution renewed every 24 hours at 37 ºC. At the end of 6 weeks, all samples were tested with Instron ® 3369 (Norwood Massachusetts, USA) four point bending test.

Staphylococcus aureus (ATCC 29213) strain was used for samples of antibiotics containing vancomycin, teicoplanin and daptomycin after the samples prepared for antibiotic release were maintained under sterile conditions and kept in Phosphate Buffered Saline solution as appropriate. For samples containing meropenem and piperacillin - tazobactam antibiotics, Pseudomonas aeruginosa (ATCC 27853) strain was used.

The addition of more than 5% antibiotics to the cement powder was significantly reduced mechanical strength in all groups(p <0.05) however the power of significance was changed depending on the type of antibiotic. In general, adding antibiotics with 2.5% and less for cement amount was not cause significant changes in mechanical measurements. There was a negative correlation between the increase in the amount of antibiotics mixed with cement and the durability of the cement (p: <0.001, r: −0.883 to 0.914).

In this study, especially the antibacterial effects of piperacillin-tazobactam, containing 0.25 gr and 0.5 gr antibiotic doses, were found to be low. There was no bacterial growth in all other groups for 21 days. Considering the mechanical properties of groups containing meropenem, vancomycin, daptomycin and teicoplanin, it was observed that all antibiotic cements remained above the limit value of 50 MPa in the bending test at concentrations containing 2.5% and less antibiotics. This was not achieved for the piperacillin-tazobactam group. The findings of the study showed that each antibiotic has different MPa values at different doses. Therefore, it could be concluded that not only the antibiotic dose but also the type oould change the mechanical properties. In the light of these findings, mixing more than 2.5% antibiotics in cement for the antibiotic types included in the study was ineffective in terms of antibacterial effect and mechanically reduces the durability of cement below the standard value of 50 MPa.

In chronically infected fracture non-unions, treatment requires extensive debridement to remove necrotic and infected bone, often resulting in large defects requiring elaborate and prolonged bone reconstruction. One approach includes the induced membrane technique (IMT), although the differences in outcome between infected and non-infectious aetiologies remain unclear. Here we present a new rabbit humerus model for IMT secondary to infection, and, furthermore, we compare bone healing in rabbits with a chronically infected non-union compared to non-infected equivalents. A 5 mm defect was created in the humerus and filled with a polymethylmethacrylate (PMMA) spacer or left empty (n=6 per group). After 3 weeks, the PMMA spacer was replaced with a beta-tricalcium phosphate (chronOs, Synthes) scaffold, which was placed within the induced membrane and observed for a further 10 weeks. The same protocol was followed for the infected group, except that four week prior to treatment, the wound was inoculated with Staphylococcus aureus (4×10. 6. CFU/animal) and the PMMA spacer was loaded with gentamicin, and systemic therapy was applied for 4 weeks prior to chronOs application. All the animals from the infected group were culture positive during the first revision surgery (mean 3×10. 5. CFU/animal, n= 12), while at the second revision, after antibiotic therapy, all the animals were culture negative. The differences in bone healing between the non-infected and infected groups were evaluated by radiography and histology. The initially infected animals showed impaired bone healing at euthanasia, and some remnants of bacteria in histology. The non-infected animals reached bone bridging in both empty and chronOs conditions. We developed a preclinical in vivo model to investigate how bacterial infection influence bone healing in large defects with the future aim to explore new treatment concepts of infected non-union

Serial section electron microscopy (SSEM) was initially developed to map the neural connections in the brain. SSEM eventually led to the term ‘Connectomics’ to be coined to describe process of following a cell or structure through a volume of tissue. This permits the true three-dimensionality to be appreciated and relationships between cells and structures. The purpose of this study was to utilize this methodology to interrogate S. aureus infected bone. Bone samples were harvested from mice tibia infected with S. aureus and were fixed, decalcified, and osmicated. The samples were paraffin embedded and 5-micron sections were cut to identify regions of bacterial invasion into the osteocyte-lacuna-canalicular-network (OLCN). This area was cut from the paraffin block, deparaffinized, post-fixed and reprocessed into epoxy resin. Serial sections were cut at 60nm and collected onto Kapton tape utilizing the Automated Tape-collecting Ultramicrotome (ATUMtome) system. Samples were mounted onto 4” silicon wafers and post-stained with 2% uranyl acetate followed by 0.3% lead citrate and carbon coated. A ZEISS GeminiSEM 450 scanning electron microscope fitted with an electron backscatter diffusion detector was used to image the sections. The image stack was aligned and segmented using the open-source software, VASTlite. 264 serial sections were imaged, representing approximately 40 × 45 × 15-micron (x, y, z) volume of tissue. 70% of the canaliculi demonstrated infiltration by S. aureus. This study demonstrates that SSEM can be applied to the skeletal system and provide a new solution to investigate the OLCN system. It is feasible that this methodology could be implemented to investigate why some canaliculi are resistant to colonization and potentially opens up a new direction for the prevention of chronic osteomyelitis. In order to make this a realistic target, automated segmentation methodologies utilizing machine learning must be developed and applied to the bone tissue datasets

Treatment of bone infection often includes a burdensome two-stage revision. After debridement, contaminated implants are removed and replaced with a non-absorbable cement spacer loaded with antibiotics. Weeks later, the spacer is exchanged with a bone graft aiding bone healing. However, even with this two-stage approach infection persists. In this study, we investigated whether a novel 3D-printed, antibiotic-loaded, osteoinductive calcium phosphate scaffold (CPS) is effective in single-stage revision of an infected non-union with segmental bone loss in rabbits. A 5 mm defect was created in the radius of female New Zealand White rabbits. The bone fragment was replaced, stabilized with cerclage wire and inoculated with Staphylococcus aureus (MSSA). After 4 weeks, the infected bone fragment was removed, the site debrided and a spacer implanted. Depending on group allocation, rabbits received: 1) PMMA spacer with gentamycin; 2) CPS loaded with rifampin and vancomycin and 3) Non-loaded CPS. These groups received systemic cefazolin for 4 weeks after revision. Group 4 received a loaded CPS without any adjunctive systemic therapy (n=12 group1-3, n=11 group 4). All animals were euthanized 8 weeks after revision and assessed by quantitative bacteriology or histology. Covariance analysis (ANCOVA) and multiple regression were performed. All animals were culture positive at revision surgery. Half of the animals in all groups had eliminated the infection by end of study. In a historical control group with empty defect and no systemic antibiotic treatment, all animals were infected at euthanasia. There was no significant difference in CFU counts between groups at euthanasia. Our results show that treating an osteomyelitis with segmental bone loss either with CPS or PMMA has a similar cure rate of infection. However, by not requiring a second surgery, the use of CPS may offer advantages over non-resorbable equivalents such as PMMA

Introduction and Objective. Management of bone loss associated with bone contamination or infection represents a double biological and clinical challenge frequent in traumatology. The advent of new biomaterials can allow a different approach in the treatment of bone gap. The purpose of this study was to evaluate the prophylactic and therapeutic effectiveness of addition of a new absorbable bone substitute (BS) eluting different antibiotics in reconstruction of bone defects after infections and fractures with soft tissue damage. Materials and Methods. We conducted a review of patients with contaminated or infected bone defects treated using a new biomaterial, a porous composite of collagen matrices and Beta tricalcium phosphate (β TCP), able to provide a long-term release of different antibiotics. We have included treatment of osteomyelitis and osteosynthesis of exposed fracture (Gustilo Anderson 1–3b) or fractures with soft tissue damage and high risk of contamination. Surgical technique included debridement filling bone defect with BS eluting antibiotics, osteosynthesis (plate, nail, external fixator, kirschner wire), soft tissue coverage, and systemic antibiotic therapy. Radiographic and clinical data including complications (wound dehiscence, superficial or deep infection, osteomyelitis) were collected. Results. We treated 25 patients (21 male, 4 female) with mean age 47 yrs. (range 21–83). The locations treated (for incidence) was: 9 femurs (7 plates, 2 nail), 7 calcanei (one bilateral), 3 tibias, 2 forearms, 2 metatarsi, 2 hands, 1 elbow. 6 patients had large bone loss. 7 patients had bone infections (4 were Cierny Madern 4); 8 patients had osteosynthesis of exposed fractures Gustilo Anderson 1–3b (9 plate, one bilateral calcaneus). 8 patients had treatment for pseudoarthrosis of exposed fractures (6 femurs, 1 forearm, 1 metatarsus) and 3 patients a prophylactic treatment for calcaneal fractures with soft tissue damage. 4 deep infection were treated with multiple surgical debridement and new filling bone defect with BS eluting antibiotic with infection eradication. We have used a combination of vancomycin and gentamicin on 15 cases, vancomycin alone on 4 cases, combination of vancomycin and amikacin on 1 case and amikacin and Linezolid in a targeted multi drug resistance. At final follow-up functional outcome was good in all cases with bone healing. Conclusions. Extensive debridement is a fundamental requisite for eradication of bone infections and contamination. Filling of the bone void with loaded bio-composite eluting diversifiable local antibiotics with synergistic anti-biofilm activity is desirable. Treatment of this bone defects are advantaged when combining his reconstruction with BS and the possibility of release high antibiotic concentration at least for 10 days. This is an important complementing prophylactic and therapeutic antimicrobial option with adjuvant role to systemic therapy that enlarges the success rate

Intro. Calcium sulphate (CS) is a recent alternative for antibiotic elution in infected bones and joints. The purpose of this study is to evaluate the use of antibiotic impregnated calcium sulphate (AICS) beads in the management of infected tibia and femur, with regards to patient outcomes and complication rates (including reinfection rate, remission rate and union rate). Methods. Searches of AMED, CINAHL, EMBASE, EMCARE, Medline, PubMed and Google Scholar were conducted in June 2020, with the mesh terms: “Calcium sulphate beads” or “Calcium sulfate beads” or “antibiotic beads” or “Stimulan” AND “Bone infection” or “Osteomyelitis” or “Debridement” AND “Tibia” or “Femur”. Risk of bias was assessed using the Risk of Bias in Non-randomised Studies of interventions (ROBINS-i) tool, and quality assessed via the Grading of Recommendations, Assessment, Development and Evaluation (GRADE) criteria. Results. Out of 105 relevant papers, 11 met the inclusion criteria for data extraction. Total infection recurrence rate was 6.8% (range 3.2 – 11.9%, n = 295), which was significantly lower (p < 0.001) than that of polymethylmethacrylate (PMMA; 19.6%, n = 163). Complication rates varied. The main issue regarding AICS use was wound drainage (7.9 – 33.3%), which was considerably higher in studies involving treatment of the tibia only. Studies using PMMA did not experience this issue, but there were a few incidences of superficial pin tract infection following surgery. Conclusions. AICS was consistently effective at infection eradication, despite variation in causative organism and location of bead placement. Additionally, PMMA has many inconvenient properties. AICS is therefore an attractive alternative as an adjunct in treatment of infected tibia and femur. Wound drainage rate varied and was higher in studies regarding tibial cases alone

Bone infection occurring after fractures or orthopedic surgery can progress to the chronic stage and lead to poor results of treatment. Optimal treatment of chronic osteomyelitis are stabilization the fracture, biological recovery of bone defects and destroy bacterial infection. Traditional methods of treatment are systemic administration of antibiotics and surgical treatment of active infection focus. Systemic antibiotics are part of the standard therapy after surgical treatment of infected bone, but their effectiveness is limited due to malnutrition and low absorption at the site of infection. Moreover, long-term treatment and higher doses are associated with serious side effects. The aim of this investigation was to study the results of the complex treatment of patients with chronic osteomyelitis using biodegradable nanomaterials “PerOssal” as antibiotic delivery system. The study was performed at Regional center traumatology and orthopedics, Karaganda, Kazakhstan. A total 20 patient with post-traumatic/post-operative osteomyelitis were included in this open-label, prospective study. Bacteriological examination was taken with the determination of culture and sensitivity test preoperatively, during and postoperatively. After radical surgical debridement and ultrasound cavitation, the bone cavity was full filled with Perosal which can be loaded with different antibiotics depending from the antibiotic sensitivity test. Postoperative wound is completely was sutured. Systemic antibiotic treatment are allowed. The course of infection was monitored by determination leukocyte count and blood sedimentation rate; blood samples were taken befor, 24 hours after surgery, and on days 3, 7, 10, 14. Wound healing was assessed on days 2, 3, 7, 10, and at the time of removal of sutures. Resorption of implanted beads and bone reconstruction were evaluated by X-ray at after operation and at approximately one, three and six months after implantation. A total of 20 patients (mean age 38,1 (26 to 53), 14 male, 6 female) were treated with Perossal pellets (AAP, Germany) from October 2013 to April 2015. Mean leukocyte counts and blood sedimentation rate were within the normal laboratory range and did not indicate infectious complications during the first 21 days after surgery. Primary wound healing occurred in 18 patients and secondary wound healing in two patients. There were two cases of re-infection during the course of the study, one of them related to an incomplete eradication of infected tissue and multidrug-resistant strain occurring during the course of the study, the other is occurred that patient non-compliance. Radiographic analysis six months after surgery showed progressive resorption of the implanted pellets, but only 10 cases have decreasing size of defects on X-ray. This study in adult patients with chronic post-traumatic/post-operative osteomyelitis demonstrated that these biodegradable bone filler pellets which can be loaded with different antibiotics are a clinically useful local antibiotic delivery system and bone substitute which can be used as an alternative to other anti-infective implants. The implantation of the pellets was safety and well tolerated in all patients. This composite can provide adequate protection against bacterial infection during the first weeks after implantation and to support the bone healing process

Summary Statement. A single, locally-delivered injection of a human placental product containing multipotent stromal cells reduced severity of infection in an immunosuppressed murine osteomyelitis model and eliminated infection in 25% of animals compared with 0% of controls without the use of antibiotics. Introduction. Implant–associated osteomyelitis is a serious orthopaedic condition and is particularly difficult to treat in immunosuppressed individuals. Despite great advancement in the field of biomaterials and pharmaceuticals, emerging patterns of antibiotic resistance, complex biofilm production and penetration of therapeutic concentrations of effective antibiotics into bone continue to represent unmet clinical challenges. The promise of adult multipotent stromal cells (MSCs) for tissue regeneration has been of intense interest in recent years. Among their many potential therapeutic uses, MSCs have also been shown to have direct antimicrobial properties. The objective of this study was to evaluate the efficacy of a locally–delivered human placental-based tissue product containing multipotent stromal cells (hAmSC) to reduce the severity of implant-associated Staphylococcus aureus osteomyelitis in an immunosuppressed murine model. We hypothesised that athymic mice with implant-associated osteomyelitis would have diminished infection following treatment with hAmSC as evidenced by decreased bioluminescence intensity and lower histologic scores for infection and bacterial load when compared to saline-treated controls. Methods. An athymic murine model of chronic implant-associated osteomyelitis was developed using luciferase-transfected Staphylococcus aureus to study the antimicrobial effects of a human placental-based product containing multi-potent stromal cells (hAmSC). Sixteen athymic mice had osteomyelitis established in the right femoral diaphysis. Fifteen days after inducing luc S. aureus osteomyelitis, the mice were randomised to receive a single 0.5 cc injection of hAmSC (n=8) or vehicle (0.9% saline) (n=8) into the soft tissues immediately adjacent to the infected bone. No antibiotics were administered throughout the duration of the study. Mice were imaged with an In Vivo Imaging System (IVIS 1000, PerkinElmer) twice weekly for 30 days to assess change in bioluminescence intensity from baseline immediately prior to treatment with either hAmSC or saline. Radiographs were obtained at days −10, 0, 10, 20 and 30 days post-injection and scored for bone changes secondary to osteomyelitis by a reviewer blinded to treatment group. Mice were sacrificed 30 days after treatment and femurs were examined histologically and scored for bacterial load and degree of inflammation by a pathologist blinded to treatment group. Results. Osteomyelitis was successfully established in all mice as evidenced by baseline bioluminescence imaging and radiographs. Mean bioluminescence intensity decreased from baseline in animals receiving hAmSC and remained below baseline for 28 days, whereas vehicle-treated animals showed an increase in mean bioluminescence intensity throughout the study period. Osteomyelitis resolved in 2/8 hAmSC-treated animals and 0/8 vehicle-treated animals as evidenced by bioluminescence imaging and histological examination for bacteria/inflammation at sacrifice. Radiograph scores for secondary bone changes were lower in mice treated with hAmSC than vehicle at 10, 20 and 30 days post injection. Median inflammatory score was lower in the hAmSC-treated mice than vehicle treated controls. Conclusions. A single injection of hAmSC was effective at reducing the severity of S. aureus infection without the use of antibiotics in this chronic implant associated osteomyelitis immunosuppressed murine model. In addition to reduced bioluminescence intensity below baseline for 28 days during the study period, infection was eliminated in 25% of animals in the hAmSC-treated group

Objectives

We have observed clinical cases where bone is formed in the overlaying muscle covering surgically created bone defects treated with a hydroxyapatite/calcium sulphate biomaterial. Our objective was to investigate the osteoinductive potential of the biomaterial and to determine if growth factors secreted from local bone cells induce osteoblastic differentiation of muscle cells.

The treatment of chronic osteomyelitis often includes surgical debridement and filling the resultant void with antibiotic-loaded polymethylmethacrylate cement, bone grafts or bone substitutes. Recently, the use of bioactive glass to treat bone defects in infections has been reported in a limited series of patients. However, no direct comparison between this biomaterial and antibiotic-loaded bone substitute has been performed.

In this retrospective study, we compared the safety and efficacy of surgical debridement and local application of the bioactive glass S53P4 in a series of 27 patients affected by chronic osteomyelitis of the long bones (Group A) with two other series, treated respectively with an antibiotic-loaded hydroxyapatite and calcium sulphate compound (Group B; n = 27) or a mixture of tricalcium phosphate and an antibiotic-loaded demineralised bone matrix (Group C; n = 22). Systemic antibiotics were also used in all groups.

After comparable periods of follow-up, the control of infection was similar in the three groups. In particular, 25 out of 27 (92.6%) patients of Group A, 24 out of 27 (88.9%) in Group B and 19 out of 22 (86.3%) in Group C showed no infection recurrence at means of 21.8 (12 to 36), 22.1 (12 to 36) and 21.5 (12 to 36) months follow-up, respectively, while Group A showed a reduced wound complication rate.

Our results show that patients treated with a bioactive glass without local antibiotics achieved similar eradication of infection and less drainage than those treated with two different antibiotic-loaded calcium-based bone substitutes.

Cite this article: Bone Joint J 2014; 96-B:845–50.

Ciprofloxacin hydrochloride-loaded microspheres were prepared by a spray-drying method using pectin and chitosan. The effects of different polymers and drug ratios were investigated.

The most appropriate carriers were selected by in vitro testing. A rat methicillin-resistant Staphylococcus aureus osteomyelitis model was used to evaluate the effects of the loaded microspheres.

The drug was released rapidly from the pectin carrier but this was more sustained in the chitosan formulation.

Chitosan microspheres loaded with ciprofloxacin hydrochloride were more effective for the treatment of osteomyelitis than equivalent intramuscular antibiotics.