Aim. The aim of this study was to develop an in-house multiplex PCR real-time assay on the LightCycler 480 system (Roche, Basel, Switzerland) with the aim of rapid detection of common pathogens in prosthetic joint infections (PJI), followed by validation on clinical samples (sonication fluid and tissue biopsies) routinely collected for PJI diagnosis. Methods. Using the PrimerQuest and CLC WorkBench tool, we designed six primer sets with specific fluorescently labelled TaqMan probes for the nuc gene in different Staphylococcus species (S. aureus, S. epidermidis, S. capitis, S. lugdunensis, S. hominis, S. haemolyticus). In addition, primers previously developed by Renz et al. (2022) for C. acnes were integrated into our assay with internal control of isolation, leading to the development of specific mPCR assay with seven included targets. Analytical sensitivity and specificity were evaluated using reference bacterial strains. To determine the assay's limit of detection (LOD), we conducted serial dilutions of eluates containing known concentrations of bacterial DNA copies/µl. The overall LOD in spiked clinical samples, including sample preparation and DNA isolation on MagnaPure24, was measured through 10-fold serial dilutions (from 10. 9. to 10. -1. CFU/ml) including additional dilutions of 5000, 500, 50 and 5 CFU/ml. Results. The results with LOD in serial dilutions of eluates and spiked clinical samples, together with analytical sensitivity and specificity, are shown in Table 1. Conclusion. The mPCR assay showed excellent analytical sensitivity and specificity, but with considerably lower LOD after sample preparation and further DNA isolation in spiked clinical samples. Although still promising in diagnostics of acute infections, the use of mPCR could be challenging in chronic, low-grade infections with lower microbial burden. Nevertheless, PCR offers significant advantages in terms of speed and can shorten the time to result, especially for C. acnes infections. Additionally, it represents a promising complementary approach in patients with suspected PJI on antibiotic therapy with negative culture results. For any tables or figures, please contact the authors directly

Aim. We prospectively evaluated four different microbiological tools for diagnostics of prosthetic joint infections (PJI), and assessed their impact on the categorization of infection according to EBJIS guidelines. We compared culture, in-house real-time mPCR for S. aureus, S. lugdunensis, S. hominis, S. epidermidis, S. capitis, S. haemolyticus, C. acnes (mPCR), broad-spectrum PCR (Molzym) with 16S rRNA V3-V4 amplicon Sanger sequencing (16S PCR), and 16S rRNA V3-V4 amplicon next-generation sequencing (16S NGS) on MiSeq (Ilumina). Methods. A total of 341 samples (sonication fluid, tissue biopsy, synovial fluid) were collected from 32 patients with suspected PJI who underwent 56 revision surgeries at the Orthopaedic Centre University Hospital Ljubljana, between 2022 and 2024. Samples were processed using standard protocols for routine culture, followed by DNA isolation using the MagnaPure24 (Roche). All samples were tested with mPCR, and an additional ≥4 samples from each revision (244 in total) were subjected to further metagenomic analysis. Culture results were considered positive if the same microorganism was detected in ≥2 samples, ≥50 CFU/ml were present in the sonication fluid, or ≥1 sample was positive for a more virulent microorganism or if the patient had received antibiotic treatment. Results. Each tool demonstrated high sensitivity for correct EBJIS categorization (100% culture and 16S NGS, 96.88% mPCR and 16S PCR). The highest specificity was observed with mPCR and 16S PCR (87.5%), while culture (79.17%) and NGS (37.5%) showed lower specificity. In 27% (15/56) of revisions, all microbiological tests were negative, although infection was confirmed with histology in one case, and four cases were classified as infection-likely based on clinical signs. In 20% (11/56) of cases, all microbiological tests were positive; in three cases a combination of other EBJIS criteria (without microbiology) categorized the episodes as infection-likely and one as infection-unlikely, emphasizing the importance of microbiological tests in diagnostic criteria. In 43% (24/56) of revisions categorized as infection-unlikely using a combination of other EBJIS criteria, five had positive culture, and three had positive mPCR and 16S PCR. Fifteen (62%) had positive 16S NGS, 12 due to a low number of reads, which may indicate low-grade infection or possible contamination. Conclusion. To date, no test can be established as the ultimate gold standard. The lack of interpretation criteria can result in low specificity of some methods, as the threshold is difficult to determine. A multidisciplinary approach with combination of microbiological tools is still considered the most efficient

Aims. The efficacy of saline irrigation for treatment of implant-associated infections is limited in the presence of porous metallic implants. This study evaluated the therapeutic efficacy of antibiotic doped bioceramic (vancomycin/tobramycin-doped polyvinyl alcohol composite (PVA-VAN/TOB-P)) after saline wash in a mouse infection model implanted with titanium cylinders. Methods. Air pouches created in female BalBc mice by subcutaneous injection of air. In the first of two independent studies, pouches were implanted with titanium cylinders (400, 700, and 100 µm pore sizes) and inoculated with Staphylococcus aureus (1 × 10. 3. or 1 × 10. 6. colony-forming units (CFU)/pouch) to establish infection and biofilm formation. Mice were killed after one week for microbiological analysis. In the second study, pouches were implanted with 400 µm titanium cylinders and inoculated with S. aureus (1 × 10. 3. or 1 × 10. 6. CFU/pouch). Four groups were tested: 1) no bacteria; 2) bacteria without saline wash; 3) saline wash only; and 4) saline wash plus PVA-VAN/TOB-P. After seven days, the pouches were opened and washed with saline alone, or had an additional injection of PVA-VAN/TOB-P. Mice were killed 14 days after pouch wash. Results. The first part of the study showed that low-grade infection was more significant in 400 µm cylinders than cylinders with larger pore sizes (p < 0.05). The second part of the study showed that saline wash alone was ineffective in eradicating both low- and high-grade infections. Saline plus PVA-VAN/TOB-P eradicated the titanium cylinder-associated infections, as manifested by negative cultures of the washouts and supported by scanning electron microscopy and histology. Conclusion. Porous titanium cylinders were vulnerable to bacterial infection and biofilm formation that could not be treated by saline irrigation alone. Application of PVA-VAN/TOB-P directly into the surgical site alone or after saline wash represents a feasible approach for prevention and/or treatment of porous implant-related infections. Cite this article: Bone Joint Res 2024;13(11):622–631

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Pneumatic tourniquets are often used during the surgical treatment of unstable traumatic ankle fractures. The aim of this study was to assess the risk of reoperation after open reduction and internal fixation of ankle fractures with and without the use of pneumatic tourniquets.

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Serum inflammatory parameters are widely used to aid in diagnosing a periprosthetic joint infection (PJI). Due to their limited performances in the literature, novel and more accurate biomarkers are needed. Serum albumin-to-globulin ratio (AGR) and serum CRP-to-albumin ratio (CAR) have previously been proposed as potential new parameters, but results were mixed. The aim of this study was to assess the diagnostic accuracy of AGR and CAR in diagnosing PJI and to compare them to the established and widely used marker CRP.

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Current diagnostic tools are not always able to effectively identify periprosthetic joint infections (PJIs). Recent studies suggest that circulating microRNAs (miRNAs) undergo changes under pathological conditions such as infection. The aim of this study was to analyze miRNA expression in hip arthroplasty PJI patients.

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Histology is widely used for diagnosis of persistent infection during reimplantation in two-stage revision hip and knee arthroplasty, although data on its utility remain scarce. Therefore, this study aims to assess the predictive value of permanent sections at reimplantation in relation to reinfection risk, and to compare results of permanent and frozen sections.

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A higher failure rate has been reported in haematogenous periprosthetic joint infection (PJI) compared to non-haematogenous PJI. The reason for this difference is unknown. We investigated the outcome of haematogenous and non-haematogenous PJI to analyze the risk factors for failure in both groups of patients.

Aim. Treatment algorithms for fracture-related nonunion depend on the presence or absence of bacterial infection. However, the manifestation of septic nonunion varies. Low-grade infections, unlike manifest infections, lack clinical signs of infection and present similarly to aseptic nonunion. The clinical importance of low-grade infection in nonunion is not entirely clear. Therefore, the aim of this study was to evaluate the clinical relevance of low-grade infection in the development and management of femoral or tibial nonunion. Method. A prospective, multicenter clinical study enrolled patients with nonunion and regular healed fractures. Preoperatively, complete blood count without differential, C-reactive protein (CRP), and procalcitonin were obtained, clinical signs of infection were recorded, and a suspected septic or aseptic diagnosis was made based on history and clinical examination. During surgical nonunion revision or routine implant removal, tissue samples were collected for microbiology and histopathology, and osteosynthesis material for sonication. Nonunion patients were followed for 12 months. Definitive diagnosis of “septic” or “aseptic” nonunion was made according to diagnostic criteria for fracture-related infection, considering the results of any further revision surgery during follow-up. Results. 34 patients with regular healed fractures were included. 62 nonunion patients were diagnosed as aseptic, 22 with manifest, and 23 with low-grade infection. The positive predictive value was 88% and the negative predictive value 72% for the suspected diagnosis. The nonunion groups had significantly higher CRP levels than the regular healer group. Differentiation between septic and aseptic nonunion based on blood values was not possible. Low-grade infection demonstrated less frequently histopathologic signs of infection than manifest infection (22% vs. 50%, p=0.048), with 15% of regular healers having histopathologic signs of infection. Cutibacterium acnes was less present in manifest compared to low-grade infection (p=0.042). Healing rates for septic nonunion involving C. acnes were significantly lower for manifest infection (20%) than for low-grade infection (100%, p=0.002). Patients with low-grade infection were treated with systemic antibiotics less frequently than patients with manifest infection (p=0.026), with no significant difference in healing rate (83% vs. 64%), which was slightly lower for low-grade infection than for aseptic nonunion (90%). Conclusions. Low-grade infections play a significant role in nonunion development and are difficult to diagnose preoperatively due to the lack of clinical signs of infection and unremarkable blood counts. However, our results imply that for low-grade infections, antibiotic therapy may not always be mandatory to heal the nonunion. This study was supported by the German Social Accident Insurance (FF-FR0276)

Aim. In 10% of the presumed aseptic hip or knee revisions, a low-grade infection is unexpectedly diagnosed based on the tissue samples taken during revision. Extended antimicrobial prophylaxis can possibly reduce the failure rate in cases of unexpected PJI, because the prophylaxis can be considered as early empiric treatment. In this randomized controlled study we analysed whether extended antimicrobial prophylaxis compared to a single dose is beneficial to improve the outcome of treatment in unexpected PJI in revision arthroplasty. Method. This study was nested in a randomized clinical trial comparing single-dose cefazolin with prolonged prophylaxis (15 doses of cefazolin over 5 days) for revision arthroplasty of the hip or knee. For this analysis, patients were included if an unsuspected PJI (defined as ≥2 positive intraoperative tissue samples with the same microorganism) was diagnosed. PJI treatment consisted of 12 weeks of a rifampicin-based regimen in Staphylococcal PJI, without removal of the prosthesis. We examined Infection characteristics and success of treatment after one year, defined as the absence of signs or treatment for PJI during follow-up. Results. After randomization of 662 patients, 68 unexpected PJI were diagnosed. In 5 cases no antimicrobial treatment was started. The success rate after one year follow-up for those who received PJI treatment was 96% (28/29) in the single dose group and 91% (31/34) in the extended prophylaxis group (p=1.00). The most frequently identified pathogens in unexpected PJI were Cutibacterium acnes (n=50) and Staphylococcus epidermidis (n=14). The causatives were susceptible for the cefazolin prophylaxis in 61 of the 63 cases. The interval between the stopped prophylaxis and the re-start of antimicrobial treatment was on average 10 days (SD 4) for the single dose and 5 days (SD 4) for the extended group. The mean duration of antimicrobial treatment was 83 days (SD 12) and did not differ between both groups (p=0.16). Conclusions. This is the first randomized controlled trial in which extended prophylaxis showed no benefit on the prosthesis survival for patients with an unexpected PJI after assumed aseptic revision of the hip or knee prosthesis. The results imply that extended prophylaxis should not be given as part of early empiric therapy

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. Method. This prospective multicenter study enrolled patients undergoing revision surgery of femur or tibia nonunion. Patients with implant removal after regular fracture healing (HEAL) were included as a control-group. Preoperative blood samples, intraoperative tissue samples, sonication of osteosynthesis material and 1-year-follow-up questionnaire were taken. Nonunion patients were grouped into INF or AS after assessing bacterial culture and histopathology of retrieved samples. Diagnosis of infection followed the fracture related infection consensus group criteria, with additional consideration of healing one year after revision surgery. Targeted proteomics was used to investigate a predefined panel of 45 cytokines in preoperative blood samples. Statistical differences were calculated with Kruskal Wallis and Dunn's post hoc test. Cytokines with less than 80% of samples being above the lower limit of detection range (LLDR) were excluded for this study. Results. We recruited 62 AS, 43 INF and 32 HEAL patients. Patients in the two nonunion groups (INF and AS) did not differ concerning smoking, diabetes or initial open or closed fracture. Thirty-two cytokines were above LLDR in >80% of patients. INF patients showed a significant difference in expression of 8 cytokines compared to AS, with greatest differences observed for Macrophage Colony Stimulating Factor 1 (MCSF-1) and Hepatocyte Growth Factor (HGF) (p<0.01). In comparing AS with HEAL patients, 9 cytokines displayed significant differences, including interleukin (IL)-6, Vascular Endothelial Growth Factor A (VEGFA), Matrix Metalloproteinase 1 (MMP-1). Comparison of INF with HEAL patients revealed significantly different expression of 20 cytokines, including. IL-6, IL-18, VEGFA or MMP-1. Conclusions. Our study revealed differences in plasma cytokine profile of blood samples from INF and AS patients. Although no single biomarker is sufficient to differentiate these patients preoperatively in isolation, future multivariant analysis of this cytokine data in combination with clinical characteristics may provide valuable diagnostic insights. Funded by German Social Accident Insurance (FF-FR 0276) and AO Trauma (AR2021_04)

Pre-operative definitive diagnosis of infection in painful total hip arthroplasty (THA) is not always easy to be established, making the intra-operative decision-making process crucial in management of revision hip surgery. Calprotectin is a promising point-of-care novel biomarker that has displayed high accuracy in detecting PJIs. From November 2020 to December 2022, 105 patients with painful primary THA were treated with revision THA in 3 orthopaedic departments. Pre-operatively, 23 were considered infected and treated with two-stage revision THA. The remaining 82 were likely infected according to the 2019 EBJIS criteria. The suspicion of low-grade infection was based on clinical (rest and/or night pain), laboratory (CRP, ESR, WBC – normal or slightly elevated) and radiological evaluation (loosening). Hip aspiration under CT imaging was performed in these cases and 34 of them yielded positive culture and were treated with two-stage revision. Aspiration was ineffective in the remaining 48 cases (33 negative, 15 unsuccessful attempts). Intra-operatively, calprotectin was measured with lateral flow immunoassay test in these patients. Cases with calprotectin levels ≥ 50 mg/L were treated with 2-stage revision THA; otherwise, they were considered not-infected and one-stage revision was performed. Synovial fluid and tissue samples were collected for analysis. Implants were sent for sonication fluid cultures. Calprotectin was positive (≥ 50 mg/L) in 27 cases and negative in 21 cases. There was 1 false negative case with positive tissue cultures. Out of the 27 positive cases, 25 had positive tissue cultures and sonication. However, 2 cases with high calprotectin levels (>200 mg/L) were not infected. The false positive result was attributed to severe metallosis. Calprotectin sensitivity was 96.2%, specificity 90.9%, PPV 92.6%, NPV 95.2%, AUC 0.935. The results of this ongoing study indicate that calprotectin seems to be a valuable tool in facilitating the intra-operative decision-making process in cases that low-grade infection is suspected and diagnosis cannot be established pre-operatively

The efficacy of saline irrigation for the treatment of periprosthetic infection (PJI) is limited in the presence of infected implants. This study evaluated the efficacy of vancomycin/tobramycin-doped polyvinyl alcohol (PVA)/ceramic composites (PVA-VAN/TOB-P) after saline irrigation in a mouse pouch infection model. 3D printed porous titanium (Ti) cylinders (400, 700 and 100 µm in pore size) were implanted into mice pouches, then inoculated with S. aureus at the amounts of 1X10. 3. CFU and 1X10. 6. CFU per pouch, respectively. Mice were randomized into 4 groups (n=6 for each group): (1) no bacteria; (2) bacteria without saline wash; 3) saline wash only, and (4) saline wash+PVA-VAN/TOB-P. After seven days, pouches were washed out alone or with additional injection of 0.2 ml of PVA-VAN/TOB-P. Mice were sacrificed 14 days after pouch wash. Bacteria cultures of collected Ti cylinders and washout fluid and histology of pouch tissues were performed. The low-grade infection (1X10. 3. CFU) was more significant in 400 µm Ti cylinders than that in Ti cylinders with larger pore sizes (700 and 1000 µm (p<0.05). A similar pattern of high-grade infection (1X10. 6. CFU) was observed (p<0.05). For the end wash, the bacteria burden (0.49±0.02) in saline wash group was completely eradicated by the addition of PVA-VAN/TOB-P (0.005±0.001, p<0.05). We noticed that 400 µm Ti cylinders have the highest risk of implant infection. Our data supported that the effect of saline irrigation was very limited in the presence of contaminated porous Ti cylinders. PVA-VAN/TOB-P was biodegradable, biocompatible, and was effective in eradicating bacteria retention after saline irrigation in a mouse model of low grade and high-grade infection. We believe that PVA-VAN/TOB-P represents an alternative to reduce the risk of PJI by providing a sustained local delivery of antibiotics

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The aim of this study is to report the implant survival and factors associated with revision of total elbow arthroplasty (TEA) using data from the Dutch national registry.

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The aim of this study was to evaluate the optimal deep tissue specimen sample number for histopathological analysis in the diagnosis of periprosthetic joint infection (PJI).

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Periprosthetic joint infection (PJI) in total hip arthroplasty in the elderly may occur but has been subject to limited investigation. This study analyzed infection characteristics, surgical outcomes, and perioperative complications of octogenarians undergoing treatment for PJI in a single university-based institution.

Periprosthetic joint infection (PJI) remains an extremely challenging complication. We have focused on this issue more over the last decade than previously, but there are still many unanswered questions. We now have a workable definition that everyone should align to, but we need to continue to focus on identifying the organisms involved. Surgical strategies are evolving and care is becoming more patient-centred. There are some good studies under way. There are, however, still numerous problems to resolve, and the challenge of PJI remains a major one for the orthopaedic community. This annotation provides some up-to-date thoughts about where we are, and the way forward. There is still scope for plenty of research in this area.

Cite this article: Bone Joint J 2022;104-B(11):1193–1195.

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Biofilm-related infection is a major complication that occurs in orthopaedic surgery. Various treatments are available but efficacy to eradicate infections varies significantly. A systematic review was performed to evaluate therapeutic interventions combating biofilm-related infections on in vivo animal models.

Aim. Failed consolidation (nonunion) after foot and ankle arthrodesis is a major complication, which can lead to additional revision arthrodesis with increased risk of morbidity. Multiple factors can contribute to developing a nonunion, including a low-grade infection. The aim of this study was to investigate the rate of unsuspected low-grade infection in revision arthrodesis for nonunions after foot and ankle arthrodesis. We also analyzed the outcome of unsuspected low-grade infections. Method. We conducted a retrospective study in The Sint Maartenskliniek, The Netherlands. All patients who underwent revision arthrodesis for assumed aseptic nonunion after foot and ankle arthrodesis between January 2020 and July 2021 were included. Patients were excluded if <5 tissue samples were obtained during revision arthrodesis or if they were treated for infection after the index arthrodesis. For the included patients, at least 5 tissue samples for culture were taken during the revision arthrodesis. The causative microorganisms, antibiotic susceptibility and treatment were assessed. An unsuspected infection was defined as ≥2 positive cultures with phenotypical identical microorganisms. Success was defined as union on imaging during clinical follow-up, without signs of persistent infection after finishing the antibiotic treatment. Results. In total 91 revision arthrodesis due to nonunion were performed. The mean duration between index and revision arthrodesis was 571 days. In 14 patients, an unexpected infection was diagnosed. The most frequent causative bacteria identified were Cutibacterium acnes (n=10) and Staphylococcus spps. (n=5). One infection was caused by a Gram-negative bacilli (Acinetobacter spps.). Two infections were polymicrobial. Of the 14 infections, 12 were treated with antibiotics for 12 weeks, 1 for 6 weeks and 1 was not treated. After one-year follow-up, the success rate was 86% and in one patient re-surgery was performed for a non-infectious reason. Conclusions. In 18% of the revisions for nonunion after foot and ankle arthrodesis, an unexpected low- grade infection was the cause of the nonunion. Nonunion occurring after foot and ankle arthrodesis is a severe complication, leading to additional revision arthrodesis. Low-grade infection should be considered as possible explanation of the nonunion, despite the lack of local inflammatory signs. As 18% of the nonunions were unexpectedly caused by low-grade infection, we strongly recommend obtaining at least 5 tissue samples for culture during revision arthrodesis. The outcome of unexpected infection as cause of nonunion is good, when treated with targeted antibiotics for 12 weeks