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.

Introduction

Bone and joint infection (BJI) is often characterized by severe inflammation and progressive bone destruction. Osteocytes are the most numerous and long-lived bone cell type, and therefore represent a potentially important long-term reservoir of bacterial infection. Staphylococcus aureus is known to establish stable intracellular osteocytic infections, however, little is known about the less virulent yet second most prevalent BJI pathogen, S. epidermidis, associated with late-diagnosed, chronic BJI. Thus, this study sought to establish an in vitro model to study the infection characteristics of S. epidermidis in human osteocyte-like cells.

Aim

The time to onset of symptoms after fracture fixation is still commonly used to classify fracture-related infections (FRI). Early infections (<2 weeks) can often be treated with debridement, systemic antibiotics, irrigation, and implant preservation (DAIR). Late infections (>10 weeks) typically require implant removal as mature, antibiotic-tolerant biofilms have formed. However, the recommendations for delayed infections (2–10 weeks) are not clearly defined. Here, infection healing and bone healing in early and delayed FRI is investigated in a rabbit model with a standardized DAIR procedure.

Aim

Bacteriophages are remerging as alternative and adjunctive therapy for fracture-related infection (FRI). However, current administration protocols involve prolonged retention of a percutaneous draining tube with potential risk of developing superinfection. In this study, we applied a cocktail of in vitro evolved biofilm-targeting phages for Methicillin-resistant Staphylococcus aureus (MRSA) in a hydrogel platform co-delivering vancomycin. In vitro synergy and antibiofilm activity was assessed and a subsequent in vivo study was performed in a mouse FRI model with MRSA.

Aim

In this study we investigated the effects of non-steroidal anti-inflammatory drugs (NSAIDs) with different cyclooxygenase (COX) selectivity on orthopaedic device-related infections (ODRIs) in a rat model. Specifically, we aimed to measure the impact of NSAID therapy on bone changes, bacterial load, and cytokine levels after treatment with antibiotics. In addition, we compared the effects of long vs short-term celecoxib (a COX-2 inhibitor) treatment on the same outcomes.

Aim

Debridement, Antibiotics, Irrigation, and implant Retention (DAIR) is a surgical treatment protocol suitable for some patients with fracture related infection (FRI). Clinically relevant pre-clinical models of DAIR are scarce and none have been developed in large animals. Therefore, this project aimed to develop a large animal model for FRI including a DAIR approach and compare outcomes after 2 or 5 weeks of infection.

Aim

Differentiation of infected (INF) nonunion from aseptic (AS) nonunion is crucial for the choice of intra- and postoperative treatment. Preoperative diagnosis of infected nonunion is challenging, especially in case of low-grade infection lacking clinical signs of infection. Standard blood markers such as C-reactive protein or leucocyte count do not aid in preoperative diagnosis. Proteomic profiling has shown promising results for differentiation of numerous chronic disease states, and in this study was applied to preoperative blood samples of patients with nonunion in an attempt to identify potential biomarkers.

We investigated the effects of non-steroidal anti-inflammatory drugs (NSAIDs) with different cyclooxygenase (COX) selectivity on orthopaedic device-related infections (ODRIs) in a rat model. We aimed to measure the impact of NSAID therapy on bone changes, bacterial load, and cytokine levels after treatment with antibiotics. We also compared the effects of long vs short-term celecoxib (a COX-2 inhibitor) treatment on the same outcomes.

Skeletally mature female Wistar rats were implanted with Staphylococcus epidermidis- contaminated polyetheretherketone (PEEK) screws in the proximal right tibia and monitored for 7 days. All animals received subcutaneous antibiotics (rifampicin plus cefazolin) for two weeks from day 7 to 21. In phase I of the study, rats were randomly assigned to receive 28 days of oral treatment with acetylsalicylic acid, ibuprofen, celecoxib, or vehicle control. In phase II, an additional group received seven days of celecoxib treatment from day 0 to 7. Bone changes were monitored using in vivo micro-CT and histology. Quantitative bacteriology was performed at euthanasia. Plasma samples were collected to measure cytokine levels on days 0, 6, 20, and 28.

Combination antibiotic therapy resulted in treatment success in 85.71% of cases, while the addition of long-term celecoxib treatment reduced it to 45.45%. Long-term celecoxib treatment significantly reduced bone loss (33.85% mean difference [95% CI 14.12–53.58], p=0.0004 on day 6 bone fraction) and periosteal reaction (0.2760 μm mean difference [95% CI 0.2073–0.3448], p<0.0001 on day 14 periosteal thickness) during early infection compared to the control group. Short- term celecoxib treatment showed similar radiological results without a reduction in treatment success (88.9%). No differences in the inflammatory markers were observed.

Our findings highlight the potential benefits of short-term use of celecoxib in improving bone fraction during the early post-infection period without impairing the efficacy of antibiotic therapy

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 (4×10⁶ 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 3×10⁵ 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.

Fracture related infection remains a major challenge in musculoskeletal trauma surgery. Despite best practice, treatment strategies suffer from high failure rates due to antibiotic resistance and tolerance. Bacteriophages represent a promising alternative as they retain activity against such bacteria. However, optimal phage administration protocols remain unknown, although injectable hydrogels, loaded with phage and conventional antibiotics, may support conventional therapy.

In this study we tested the activity of meropenem, and two newly isolated bacteriophages (ϕ9 and ϕ3) embedded within alginate-chitosan microbeads and a hydrogel. Antibiotic and phage stability and activity were monitored in vitro, over a period of 10 days. In vivo, the same material was tested in treatment of a 5-day old Pseudomonas aeruginosa infection of a tibial plate osteotomy in mice. Treatment involved debridement and 5 days of systemic antibiotic therapy plus: i- saline, ii-phages in saline, iii-phages and antibiotics loaded into a hydrogel (n=7 mice/group). To assess the efficacy of the treatments, the infection load was monitored during revision surgery with debridement of the infected tissue after 5,10 and 13 days (euthanasia) by CFU and PFU quantification.

In vitro testing confirmed that the stability of meropenem and activity of ϕ9 and ϕ3, was not affected within the alginate beads or hydrogel over 10 days. The in vivo study showed that all mice receiving phages and antibiotics loaded into a hydrogel survived the infection with a reduction of the bacterial load in the soft tissue. Active phages could be recovered from the infected site at euthanasia (10⁴ PFU/g).

The hydrogel loaded with bacteriophages and meropenem showed a positive result in locally reducing the infection load indicating a synergistic effect of the selected antimicrobials. Overall, our new strategy shows encouraging results for improving the treatment of antibiotic-resistant biofilm infections that are related to medical implants.

Aim

Staphylococcus aureus is the leading pathogen in fracture-related infection (FRI). Virulence factors vary between different strains, which may have a decisive influence on the course of infection. Previous in vitro experiments, in vivo testing in wax moth larvae, and genomic analysis of S. aureus isolates from FRI identified a low- and high-virulent strain. These findings correlated with the acute course of FRI induced by the high-virulent pathogen, whereas the low-virulent strain caused a chronic FRI in its human host. However, the role of bacterial virulence in FRI is not completely understood. Therefore, the present study aimed to compare the identified high- and low-virulent S. aureus isolates in a murine FRI model.

Aim

Implant-associated osteomyelitis is a devastating complication with poor outcomes following treatment, especially when caused by antibiotic-resistant bacteria such as methicillin-resistant Staphylococcus aureus (MRSA). A large animal model of a two-stage revision to treat MRSA implant-associated osteomyelitis has been developed to assess novel treatments. A bioresorbable, thermo-responsive hyaluronan hydrogel (THH) loaded with antibiotics has been developed and our aim was to investigate it´s in vivo efficacy as a local antibiotic carrier compared to the current standard of care i.e. antibiotic-loaded polymethylmethacrylate (PMMA) bone cement.

Aim

Non-steroidal anti-inflammatory drugs (NSAIDs) are a cornerstone of perioperative pain management in orthopedic trauma surgery, although concerns persist regarding the potential impact of these drugs on fracture healing. Furthermore, NSAIDs may also exert an influence on host immune defenses, which may also be important in the context of infection treatment. However, this has been very much under-investigated in the clinical and scientific literature. The aim of this study was to determine the impact of NSAIDs on the course of an orthopedic device-related infection (ODRI) and its response to antibiotic therapy in a rat model.

Aim

Alongside debridement and irrigation, soft tissue coverage and osseous stabilization, systemic antibiotic prophylaxis is considered the gold standard in the management of open fractures and considerably reduces the risk of subsequent fracture-related Infections (FRI). The direct application of antibiotics into the surgical field (local antibiotics) has been used for decades as additional prophylaxis in open fractures, although definitive evidence confirming a beneficial effect is scarce. The purpose of the present study was to review the clinical evidence regarding the effect of prophylactic application of local antibiotics in open limb fractures.

Aim

Infection rates after management of open fractures are still high. Existing guidelines regarding prevention of this complication are inhomogeneous. A survey directed to orthopaedic trauma surgeons worldwide aims to give an overview of current practices in the management of open fractures.

Device-associated infection remains a serious clinical problem in orthopaedic and trauma surgery. The emergence of resistant organisms such as methicillin resistant Staphylococcus aureus (MRSA) has further exacerbated this problem by limiting the range of treatment options. Currently, systemic antibiotic therapy is the cornerstone of treatment, alongside surgical resection of infected tissues and implant removal. The potential for antibiotic loaded biomaterials to support the prevention and treatment of infection is significant, although the currently available options are limited in number and often re-purposed from other applications e.g. antibiotic loading of bone cement. The first part of the talk will cover the basic concepts involved in antibiotic treatment, with an emphasis on the ideal antibiotic release kinetics from biomaterials, and how bacterial biofilms and antibiotic resistance influence antimicrobial efficacy. The next generation of biomaterials for antibiotic delivery should be specifically designed with this knowledge in mind. Regulatory approval of antimicrobial combination devices, however, is an evolving process as regulatory bodies seek more robust and clinically relevant efficacy data. Approval will require preclinical efficacy using standardized animal models that recapitulate the key features of the clinical disease. The second part of this talk will cover best practice in this important stage of development.

Aim

Open fractures still have a high risk for fracture-related Infection (FRI). The optimal duration of perioperative antibiotic prophylaxis (PAP) for open fractures remains controversial due to heterogeneous guidelines and highly variable prophylactic regimens in clinical practice. In order to provide further evidence with which to support the selection of antibiotic duration for open fracture care, we performed a preclinical evaluation in a contaminated rabbit fracture model.

Aim

Treatment regimens for fracture-related infection (FRI) often refer to the classification of Willenegger and Roth, which stratifies FRIs based on time of onset of symptoms. The classification includes early (<2 weeks), delayed (2–10 weeks) and late (>10 weeks) infections. Early infections are generally treated with debridement and systemic antibiotics but may not require implant removal. Delayed and late infections, in contrast, are believed to have a mature biofilm on the implant, and therefore, treatment often involves implant removal. This distinction between early and delayed infections has never been established in a controlled clinical or preclinical study. This study tested the hypothesis that early and delayed FRIs respond differently to treatment comprising implant retention.

Aim

Staphylococcus epidermidis has emerged as an important opportunistic pathogen causing orthopedic device-related infections (ODRIs). In this prospective clinical and laboratory study, we have investigated the association of genome variation and phenotypic features of the infecting S. epidermidis isolate with the clinical outcome of the infected patient.

This longitudinal microCT study revealed the osteolytic response to a Staphylococcus epidermidis-infected implant in vivoand also demonstrates how antibiotics and/or a low bone mass state influence the morphological changes in bone and the course of the infection.

Colonisation of orthopaedic implants with Staphylococcus aureusor S. epidermidisis a major clinical concern, since infection-induced osteolysis can drastically impair implant fixation or integration within bone. High fracture incidence in post-menopausal osteoporosis patients means that this patient group are at risk of implant infection. The low bone mass in these patients may exacerbate infection-induced osteolysis, or alter antibiotic efficacy. Therefore, the aims of this study were to examine the bone changes resulting from a S. epidermidisimplant infection in vivousing microCT imaging, and to determine if a low bone mass stateinfluences the course of the infection and the efficacy of antibiotic therapy. An in vivomodel system using microCT scanning [1], involving the implantation of either a sterile or a S. epidermidis-colonised PEEK screw into the proximal tibia of 24 week-old female Wistar rats, was used to investigate the morphological changes in bone following infection over a 28 day period. In addition, the efficacy of a combination antibiotic therapy (rifampin and cefazolin: administered twice daily from days 7–21 post-screw implantation) for affecting osteolysis was also assessed. A subgroup of animals was subjected to ovariectomy (OVX) at 12 weeks of age, allowing for a 12 week period for bone loss prior to screw implantation at 24 weeks. Bone resorption and formation rates, bone-implant contact and peri-implant bone volume in the proximity of the screw were assessed by microCT scanning at days 0, 3, 6, 9, 14, 20 and 28 days post-surgery. Following euthanasia at day 28, the implanted screw, bone and soft tissues were subjected to quantitative bacteriology as a measure of the efficacy of the antibiotic regimen. In non-OVX animals S. epidermidisinfection induced marked osteolysis, which peaked between 9 and 14 days post-screw implantation. Peak bone resorption was detected at day 6, before recovering to baseline levels at day 14. Infection also resulted in extensive deposition of mineralised tissue, initially within the periosteal region (day 9–14), then subsequently in the osteolytic region at day 20–28. Quantitative bacteriology indicated all non-OVX animals remained infected. Rifampin and cefazolin successfully cleared the infection in 5/6 non-OVX animals group although there was no difference observed in CT-derived bone parameters. OVX resulted in extensive loss of trabecular bone but this did not alter the temporal pattern of infection-induced osteolysis, or mineralised tissue deposition, which was similar to that observed in the non-OVX animals. Similarly, there was no difference in bacterial counts between non-OVX and OVX animals (39,005 colony-forming units (CFU) [range: 3,675–156,800] vs 37,665 CFU [range 3,250–84,000], respectively). Interestingly, antibiotic treatment was less effective in the OVX animals (3/5 remained infected), suggesting that antibiotics have reduced efficacy in OVX animals. This study demonstrates S. epidermidis-induced osteolysis displays a similar temporal pattern in both normal and low bone mass states, with comparable bacterial loads present within the localised infection site.