Aim. One of the most accurate and inexpensive tests in detection of prosthetic joint infection (PJI) is synovial fluid white blood cell (WBC) count and differential. Since leukocytes produce many different interleukins (IL) in situation of PJI, we hypothesized that ILs could be even more accurate in detection of PJI. The aim of the study was to test, if the synovial fluid IL-6 level is superior to WBC count and differential in detection of PJI. Methods. Unselected patients undergoing total hip or knee revision surgery were prospectively included. In perioperative assessment phase, WBC count, differential and IL-6 levels of synovial fluid were measured. Patients were labelled as positive or negative according to the predefined cut-off values for IL-6 (230pg/ml) and WBC count with differential (1,7 × 10. 9. WBC/ml with ≥65% of granulocytes). During the surgery, at least 4 intraoperative samples for microbiological and one for histopathological analysis were obtained. PJI was defined as presence of sinus tract, inflammation in histopathological samples, and growth of the same microorganism in at least two or more samples of periprosthetic tissue or synovial fluid. Binary diagnostic test was performed to check the diagnostic strength of both methods in detection of PJI. Results. 49 joints of 48 patients (mean age, 71 years; 53% females) undergoing artificial hip (n = 24) or knee (n = 25) revision surgery were included. 11 joints (22%) were infected. Sensitivity of synovial fluid WBC count with differential was 82%, specificity 97%, accuracy 94%, positive and negative predictive values were 90% and 95%, respectively. Sensitivity of IL-6 level was 73%, specificity 95%, accuracy 90%, positive and negative predictive values were 80% and 92%, respectively. There was a strong statistical agreement between both tests (Kappa value=0.749) and consequently there was no significant difference in detection of PJI comparing both tests (P=0.171). Conclusion. Our study revealed that synovial fluid IL-6 level is not superior to synovial fluid WBC count with differential in detecting PJI

Aims. The diagnosis of periprosthetic joint infection (PJI) remains a challenge, as no single diagnostic test shows high diagnostic accuracy. Recently, the measurement of synovial biomarkers has shown promising results. The aim of this study was to present a novel multiplex micro-enzyme-linked immunosorbent assay (ELISA) method for the rapid and simultaneous measurement of alpha-defensin, interleukin (IL)-6, and calprotectin developed in a model buffer system and human spiked synovial fluid. Methods. A microfluidics- and chemiluminescence-based multiplex micro-ELISA point-of-care testing system was employed for the rapid and simultaneous measurement of alpha-defensin, calprotectin, and IL-6 developed in a model buffer system and spiked human synovial fluid. Cut-off values of 1.56 µg/ml for alpha-defensin, 50 µg/ml for calprotectin, and 0.031 µg/ml for IL-6 were extracted from the literature as optimal cut-off values for the diagnosis of PJI, and were used for comparison. Results. Limit of detection (LoD) was determined for each individual biomarker by means of calibration curves with serial dilutions in a model buffer system. LoDs of 0.008, 0.002, and 0.00014 µg/ml were determined for alpha-defensin, calprotectin, and IL-6, respectively. The spiked synovial fluid assay determined LoDs of 0.08, 0.31, and 0.004 µg/ml for alpha-defensin, calprotectin, and IL-6, respectively. Conclusion. These findings highlight the proposed platform’s unique features, including simultaneous measurement of three key synovial biomarkers, minimal sample volume requirements (5 to 50 µl), lower LoDs compared to conventional tests, and a short processing turnaround time of 22 minutes. Further validation studies are necessary to confirm its clinical utility. Cite this article: Bone Joint Res 2025;14(3):176–184

The paramount importance of synovial fluid in lubrication and protection of articular joints has long been recognized. Synovial fluid, a dialysate of plasma, forms an interface with both the synovium and cartilage and plays a crucial role in joint lubrication and bearing functions. In an osteoarthritic joint, damage to the articular cartilage causes modifications in the rheological properties of synovial fluid and, reducing the viscoelasticity and increasing the friction between articular surfaces. Viscosupplementation is a treatment for osteoarthritis that uses hyaluronic acid as a (visco)supplement to the diseased joint. The aim of this treatment is to restore the rheological properties of synovial fluid. Osteoarthritis is the most common disease affecting the joints in human population and among the most important causes of pain, disability and economic loss. Therefore, innovative methods are needed to more effectively treat osteoarthritis, directly addressing the disease process. Among various locomotor mechanisms that could serve to illustrate the integrated nature of functional morphology, perhaps none is more complex than the equine locomotor system. Confronting the need for evaluating the current methods to control joint disease, the horse provides an excellent animal model. As it suffers similar clinical manifestations to those seen in human, it may provide tentative biomedical extrapolations

Aim. Prosthetic joint infection (PJI) represents the second most frequent complication of total joint arthroplasty (TJA) with up to 20% of low-grade PJI treated as aseptic failure. Sensitive diagnostic criteria have been provided by EBJIS. However, to date there is no single test to reliably diagnose all PJIs. Studies of Mazzucco et al. and Fu et al. suggest that synovial fluid (SF) viscosity could be considered as an important marker for PJI. The primary aim of our study was to determine if SF viscosity is a more reliable diagnostic criterion of PJI than the SF cell count with differential (CCD), and the combined diagnostic value of SF viscosity and CCD. Method. We prospectively analysed the viscosity of SF samples obtained during TJA of hip and knee revisions. We sampled 2.5–5mL of SF for viscosity and CCD. Intraoperatively, 1mL of the sample was analysed for the CCD. The remaining SF was centrifuged for 4min at 7000rpm. The viscosity of the supernatant was determined on Ostwald viscometer as the time required to pass the viscometer at 20°C. During each surgery at least 5 microbiological and multiple histopathological samples were harvested, and explant sonication was performed. The diagnosis was based on EBJIS definition. The viscosity threshold for detecting PJI was set at 65 seconds. Results. Between December 2020 and January 2023, we analysed 65 knee and 47 hip TJA revision procedures. There were 55 septic and 57 aseptic diagnoses. As a diagnostic marker of PJI, SF viscosity achieved 100% sensitivity and 82.5% specificity, with area under the receiver operating characteristic curve (AUC) of 0.832 (95% CI 0.739, 0.925). The specificity and sensitivity of SF CCD were 98.2% and 78.2%, respectively, with AUC of 0.921 (95% CI 0.869, 0.974). Of the 10 cases incorrectly diagnosed as aseptic based on SF viscosity, 2 were acute traumas and 8 metalloses. The SF CCD in all these cases was <0.5. Of the 12 cases incorrectly diagnosed as aseptic based on SF CCD, 6 cases were culture negative, 4 C. acnes and 2 S. epidemidis isolates in microbiology. Taken together, SF viscosity and CCD achieved a combined AUC of 0.953 (95% CI 0.919, 0.987). Conclusions. Our study is the first to report that SF viscosity is more sensitive but slightly less specific for PJI than SF CCD. The study demonstrates diagnostic value of combining SF viscosity with CCD in decision making in TJA revision surgery

Aims. This study aimed to explore the diagnostic value of synovial fluid neutrophil extracellular traps (SF-NETs) in periprosthetic joint infection (PJI) diagnosis, and compare it with that of microbial culture, serum ESR and CRP, synovial white blood cell (WBC) count, and polymorphonuclear neutrophil percentage (PMN%). Methods. In a single health centre, patients with suspected PJI were enrolled from January 2013 to December 2021. The inclusion criteria were: 1) patients who were suspected to have PJI; 2) patients with complete medical records; and 3) patients from whom sufficient synovial fluid was obtained for microbial culture and NET test. Patients who received revision surgeries due to aseptic failure (AF) were selected as controls. Synovial fluid was collected for microbial culture and SF-WBC, SF-PNM%, and SF-NET detection. The receiver operating characteristic curve (ROC) of synovial NET, WBC, PMN%, and area under the curve (AUC) were obtained; the diagnostic efficacies of these diagnostic indexes were calculated and compared. Results. The levels of SF-NETs in the PJI group were significantly higher than those of the AF group. The AUC of SF-NET was 0.971 (95% confidence interval (CI) 0.903 to 0.996), the sensitivity was 93.48% (95% CI 82.10% to 98.63%), the specificity was 96.43% (95% CI 81.65% to 99.91%), the accuracy was 94.60% (95% CI 86.73% to 98.50%), the positive predictive value was 97.73%, and the negative predictive value was 90%. Further analysis showed that SF-NET could improve the diagnosis of culture-negative PJI, patients with PJI who received antibiotic treatment preoperatively, and fungal PJI. Conclusion. SF-NET is a novel and ideal synovial fluid biomarker for PJI diagnosis, which could improve PJI diagnosis greatly. Cite this article: Bone Joint Res 2023;12(2):113–120

Aim. Synovial fluid D-lactate may be useful for diagnosing septic arthritis (SA) as this biomarker is almost exclusively produced by bacteria. We evaluated the performance of synovial fluid D-lactate and determined its optimal cut-off value for diagnosing SA. Method. Consecutive patients with suspicion of septic arthritis were prospectively included. They underwent joint aspiration and synovial fluid was collected for culture, leukocyte count and D-lactate concentration (by spectrophotometry). Youden's J statistic was used for determining optimal D-lactate cut-off value on the receiver operating characteristic (ROC) curve by maximizing sensitivity and specificity. Results. A total of 155 patients were included. Using institutional criteria, 21 patients (14%) were diagnosed with SA and 134 (86%) patients with aseptic arthropathy, out of which 43 (27%) had osteoarthrosis, 80 (52%) had rheumatic arthropathy and 11 (7%) reactive arthritis. The optimal cut-off of synovial fluid D-lactate to differentiate SA from aseptic cases was 0,035 mmol/l. Synovial fluid D-lactate had a sensitivity 90% (95% CI: 70–99%) and specificity 87% (95% CI: 80–92%) compared to leukocyte count with sensitivity 81% (95% CI: 60–95%) and specificity 83% (95% CI: 76–90%). Culture was positive in only 17 (80%) out of 21 patients with SA. Conclusions. The synovial fluid D-lactate showed high sensitivity and specificity for diagnosis of SA which was higher than the current gold standard of diagnosis (culture and leukocyte count). The high sensitivity makes this biomarker useful as a point-of-care screening test for SA

Aim. Evaluate the metabolites composition of the synovial fluid from patients with PJI or aseptic failure of total knee arthroplasties. Method. The synovial fluids from 21 patients scheduled for revision total knee arthroplasty (11 with the diagnosis of PJI and 10 with aseptic failures) were analyzed using 1D 1H NMR spectroscopy. Univariate and multivariate statistical analyses were used to identify metabolites that were differentially abundant between those groups. Results. A total of 28 metabolites were identified and five of them found to be differentially abundant between infected and non-infected synovial fluids. Lactate, acetate and 3-hydroxybutyrate were found to be in a higher concentration, and glucose and creatine were found reduced in the synovial fluid from PJI patients. Conclusions. Synovial fluid from patients with PJI exhibit a distinct metabolic profile, possibly reflecting metabolic adaptation that occurs in the infected periprosthetic microenvironment. Further research and studies are warranted to gain a broader insight into the metabolic pathways engaged by both pathogen and immune cells in the context of a PJI

Aims. This study aimed to investigate the role and mechanism of meniscal cell lysate (MCL) in fibroblast-like synoviocytes (FLSs) and osteoarthritis (OA). Methods. Meniscus and synovial tissue were collected from 14 patients with and without OA. MCL and FLS proteins were extracted and analyzed by liquid chromatography‒mass spectrometry (LC‒MS). The roles of MCL and adenine nucleotide translocase 3 (ANT3) in FLSs were examined by enzyme-linked immunosorbent assay (ELISA), flow cytometry, immunofluorescence, and transmission electron microscopy. Histological analysis was performed to determine ANT3 expression levels in a male mouse model. Results. We discovered for the first time that MCL was substantially enriched in the synovial fluid of OA patients and promoted the release of inflammatory cytokines from FLSs through MCL phagocytosis. Through LC‒MS, ANT3 was identified and determined to be significantly upregulated in MCL and OA-FLSs, corresponding to impaired mitochondrial function and cell viability in OA-FLSs. Mitochondrial homeostasis was restored by ANT3 suppression, thereby alleviating synovial inflammation. Furthermore, elevated ANT3 levels inhibited ERK phosphorylation. Specifically, silencing ANT3 prevented inhibition of ERK phosphorylation and significantly reduced the elevation of reactive oxygen species (ROS) and JC1 membrane potential in MCL-induced synovial inflammation. Conclusion. This study revealed the important roles of MCL and ANT3 in FLS mitochondria. Silencing ANT3 rescued ERK phosphorylation, thereby restoring mitochondrial homeostasis in FLSs and alleviating synovitis and OA development, offering a potential target for treating synovitis and preventing early-stage OA. Cite this article: Bone Joint Res 2023;12(4):274–284

Aims. This study aimed to evaluate calprotectin in synovial fluid for diagnosing chronic prosthetic joint infection (PJI) . Methods. A total of 63 patients who were suspected of PJI were enrolled. The synovial fluid calprotectin was tested by an enzyme-linked immunosorbent assay (ELISA). Laboratory test data, such as ESR, CRP, synovial fluid white blood cells (SF-WBCs), and synovial fluid polymorphonuclear cells (SF-PMNs), were documented. Chi-squared tests were used to compare the sensitivity and specificity of calprotectin and laboratory tests. The area under the curve (AUC) of the receiver operating characteristic (ROC) curve was calculated to determine diagnostic efficacy. Results. The median calprotectin level was 776 μg/ml (interquartile range (IQR) 536.5 to 1132) in the PJI group and 54.5 μg/ml (IQR, 38.75 to 78.25) in the aseptic failure (AF) group (p < 0.05). Using a threshold of 173 ug/ml, the sensitivity was 95.2%, with a 97.6% specificity, and the AUC was 0.993. The sensitivity of calprotectin of the antibiotic-treated PJI group was 100% versus 90.9% of the non-antibiotic-treated PJI group. Although 47.6% (ten cases) of the patients in the PJI group received antibiotics before aspiration, the diagnostic efficacy of calprotectin was not affected. The sensitivity and specificity of ESR, CRP, SF-WBCs, and SF-PMNs ranged from 76.2% to 90.5% and 64.3% to 85.7%, respectively. Conclusion. Calprotectin in synovial fluid has great diagnostic efficacy for PJI diagnosisand outperformed ESR, CRP, SF-WBCs, and SF-PMNs. Cite this article: Bone Joint Res 2020;9(8):450–456

Aims. Synovial fluid white blood cell (WBC) count and percentage of polymorphonuclear cells (%PMN) are elevated at periprosthetic joint infection (PJI). Leucocytes produce different interleukins (IL), including IL-6, so we hypothesized that synovial fluid IL-6 could be a more accurate predictor of PJI than synovial fluid WBC count and %PMN. The main aim of our study was to compare the predictive performance of all three diagnostic tests in the detection of PJI. Methods. Patients undergoing total hip or knee revision surgery were included. In the perioperative assessment phase, synovial fluid WBC count, %PMN, and IL-6 concentration were measured. Patients were labeled as positive or negative according to the predefined cut-off values for IL-6 and WBC count with %PMN. Intraoperative samples for microbiological and histopathological analysis were obtained. PJI was defined as the presence of sinus tract, inflammation in histopathological samples, and growth of the same microorganism in a minimum of two or more samples out of at least four taken. Results. In total, 49 joints in 48 patients (mean age 68 years (SD 10; 26 females (54%), 25 knees (51%)) were included. Of these 11 joints (22%) were infected. The synovial fluid WBC count and %PMN predicted PJI with sensitivity, specificity, accuracy, PPV, and NPV of 82%, 97%, 94%, 90%, and 95%, respectively. Synovial fluid IL-6 predicted PJI with sensitivity, specificity, accuracy, PPV, and NPV of 73%, 95%, 90%, 80%, and 92%, respectively. A comparison of predictive performance indicated a strong agreement between tests. Conclusions. Synovial fluid IL-6 is not superior to synovial fluid WBC count and %PMN in detecting PJI. Level of Evidence: Therapeutic Level II. Cite this article: Bone Jt Open 2020;1-12:737–742

Aims. To analyze the potential role of synovial fluid peptidase activity as a measure of disease burden and predictive biomarker of progression in knee osteoarthritis (KOA). Methods. A cross-sectional study of 39 patients (women 71.8%, men 28.2%; mean age of 72.03 years (SD 1.15) with advanced KOA (Ahlbäck grade ≥ 3 and clinical indications for arthrocentesis) recruited through the (Orthopaedic Department at the Complejo Asistencial Universitario de León, Spain (CAULE)), measuring synovial fluid levels of puromycin-sensitive aminopeptidase (PSA), neutral aminopeptidase (NAP), aminopeptidase B (APB), prolyl endopeptidase (PEP), aspartate aminopeptidase (ASP), glutamyl aminopeptidase (GLU) and pyroglutamyl aminopeptidase (PGAP). Results. Synovial fluid peptidase activity varied significantly as a function of clinical signs, with differences in levels of PEP (p = 0.020), ASP (p < 0.001), and PGAP (p = 0. 003) associated with knee locking, PEP (p = 0.006), ASP (p = 0.001), GLU (p = 0.037), and PGAP (p = 0.000) with knee failure, and PEP (p = 0.006), ASP (p = 0.001), GLU (p = 0.037), and PGAP (p < 0.001) with knee effusion. Further, patients with the greatest functional impairment had significantly higher levels of APB (p = 0.005), PEP (p = 0.005), ASP (p = 0.006), GLU (p = 0.020), and PGAP (p < 0.001) activity, though not of NAP or PSA, indicating local alterations in the renin-angiotensin system. A binary logistic regression model showed that PSA was protective (p = 0.005; Exp (B) 0.949), whereas PEP (p = 0.005) and GLU were risk factors (p = 0.012). Conclusion. These results suggest synovial fluid peptidase activity could play a role as a measure of disease burden and predictive biomarker of progression in KOA. Cite this article: Bone Joint Res 2020;9(11):789–797

Aim. Periprosthetic joint infections (PJI) are severe complications after total joint arthroplasty (TJA). Up to now, a gold standard in the diagnostics of PJI is missing. Small extracellular vesicles (sEVs) are secreted by all types of cells and play a key role in immune response in presence of infection (1). In this prospective study, the diagnostic accuracy of sEVs in the synovial fluid to detect PJI of knee, hip and shoulder joints was investigated. We hypothesized increased surface markers of sEVs in PJI compared to aseptic complications (e.g. implant loosening, stress shielding related pain). Method. Synovial fluid from 48 patients with painful arthroplasty was examined. The distinction between aseptic and infectious cases was made on the basis of the 2018 Definition of Periprosthetic Hip and Knee Infection (2). 35 (72,9%) probands assigned to aseptic and 13 patients (27,1%) to PJI group. Immuno-fluorescence flow cytometry served to document the concentrations of CD9, CD63, CD66b, CD82 and HLA-DR on sEVs. Results. The concentration of CD9 surface marker on sEVs in synovial fluid was significantly lower (p=0.002) in PJI group than in aseptic group. In contrast, the levels of CD82 on sEVs in synovial fluid was significantly higher (p<0.0001) in the PJI group than in aseptic group. The concentrations of CD63, CD66b and HLA-DR on sEVs in synovial fluid did not differ significantly between the two cohorts (CD63: p=0.372; CD66b: p=0.634; HLA-DR: p=0.558). Conclusions. Overall, the significance of sEVs in the diagnostics of PJI is not well enough understood and the subject of current research and scientific discussion. Our data suggest, that CD82 and CD9 on sEVs in synovial fluid are promising biomarkers to differentiate between PJI and aseptic complications

Aim. Evaluate if Neutrophil Extracellular Traps related biomarkers (citrullinated histone H3 [H3Cit], cellfree DNA [cfDNA], and myeloperoxidase) are increased in synovial fluid of patients with PJI and investigate the diagnostic accuracy of NET formation biomarkers for PJI. Method. Patients who underwent hip or knee revision total joint arthroplasty were categorised into two groups according to the Second International Consensus Meeting on Musculoskeletal Infection (2018) criteria. Sixteen patients were classified as infected and 16 as non-infected. cf-DNA, myeloperoxidase and H3Cit were measured in synovial fluid collected during surgery. Sensitivity, specificity, and receiver operating characteristic (ROC) curve were calculated. Results. Patients with PJI presented significantly higher levels of synovial fluid cf-DNA (105.5 ng/ml ± 58.3 vs 1.9 ± 1.2, p>0.0001), myeloperoxidase (1575 pg/ml ± 826 vs 50.16 ± 100, p<0.0001) and citrullinated histone H3 (1.688 ± 1.214 vs 13.88 ± 24.4, p < 0.0001). In the ROC curve analyses, the area under the curve for cf-DNA, myeloperoxidase and H3cit were 1 [0.89 – 1], 0.98 [0.86 – 1], and 0.94 [0.8 – 0.99], respectively. The sensitivity for detecting PJI using synovial fluid was 100% for cf-DNA, 93,7% for myeloperoxidase, and 87,5% for H3cit. The sensitivity for cf-DNA and myeloperoxidase was 100%, and 87,5 % for H3cit. Conclusions. Our results show that neutrophils within periprosthetic microenvironment release NETs as part of the bactericidal arsenal to fight infection. These results allow a better understanding of the cellular and molecular processes that occur in this microenvironment, enabling the design of more assertive strategies for the identification of new biomarkers and for a better use of the available ones. Furthermore, novel studies are needed to define whether and how NET-related biomarkers can be useful for the diagnosis of PJI

Aim. Prosthetic joint infection (PJI) presents the second most common complication of total joint arthroplasty (TJA). Accumulating evidence suggests that up to 20% of aseptic failures are low-grade PJI. However, there is still no single test to reliably diagnose all PJI. In his thesis, Mazzucco emphasized the viscosity differences between normal, osteoarthritic, and rheumatic synovial fluid. Similarly, a recent study by Fu et al. reported significantly lower viscosity in patients with PJI compared to the aseptic failure cohort. The primary aim of our study was to determine whether synovial fluid viscosity is a more reliable diagnostic criterion for PJI compared to the synovial fluid cell count with differential and serum C-reactive protein (CRP) levels. Method. We prospectively analyzed the viscosity of synovial fluid samples obtained during TJA of hip and knee joint revision procedures. We sampled 2.5–5 mL of synovial fluid for viscosity measurement. The samples were centrifuged (4 min at 7000 rpm) and the resulting supernatant was immediately transferred into the Ostwald viscometer. Viscosity was derived from the time required for a given volume of synovial fluid to pass the viscometer at 20 °C. The synovial fluid samples were also analysed for their cell count with differential and serum CRP was measured. The definite diagnosis of PJI was established on basis of EBJIS criteria. For the viscosity, the threshold for detecting PJI was set at 65 seconds. Results. Between December 2020 and March 2021, we analyzed 12 knee and 11 hip TJA revision samples. These included 14 septic and 9 aseptic synovial fluid samples. The average viscometer time in the PJI group was 31s (range 20–48s) compared to 247s (range 68–616s) in the group of aseptic revision procedures. The specificity and sensitivity of our viscosity measurements were 100%. The sensitivity and specificity of cell count was 100% and 85.7%, for the synovial fluid differential they were 100% and 85.7%, and for the CRP they were 88.9% and 71.4%, respectively. Conclusions. Our study is the first to report a significant difference in synovial fluid viscosity between the PJI and the aseptic cohort. It points towards the diagnostic superiority of viscosity measurements over conventional synovial fluid cell count, synovial fluid differential, and serum CRP levels. Albeit currently limited by small sample size, the study remains ongoing

Aim. Periprosthetic joint infection (PJI) is one of the most frequent and devastating complications of total knee arthroplasty (TKA). Accurate diagnosis and proper treatment are essential to prevent functional loss and progression to systemic infection. However, the correct diagnosis of PJI is still a challenge since there is no accurate diagnostic method and the existing diagnostic criteria are based on serological, histological and microbiological tests that are imprecise and time-consuming. Recently, it was demonstrated that cell-free DNA is increased in the synovial fluid of patients with PJI. Therefore, this study aims to evaluate a new point-of-care methodology for quantifying free DNA in synovial fluid. Method. A prospective study was carried out with patients undergoing TKA revision surgery, from whom it was possible to collect synovial fluid (SF) during the surgical procedure. Cell-free DNA quantification was performed directly from the SF, using a portable fluorimeter. Sensitivity, specificity and receiver operating characteristic (ROC) curve were calculated. Results. Fifty-four patients were included in the study, of which 25 were diagnosed with PJI. Cell-free DNA levels measured immediately after collection were increased in the synovial fluid of patients with PJI (26.3 ± 14.8) in comparison with the uninfected group (4.6 ng/ml ± 3.8, p< 0.0001). The area under the receiver operating characteristic curve (AUC ROC) was 0.981 (95% CI 0.914 to 0.999). Conclusions. From the results presented, we can conclude that the quantification of free DNA with a portable fluorimeter proved to be a test with high sensitivity and specificity for the diagnosis of PJI

Aims. The aim of this study was to evaluate the performance of metagenomic next-generation sequencing (mNGS) in detecting pathogens from synovial fluid of prosthetic joint infection (PJI) patients. Methods. A group of 75 patients who underwent revision knee or hip arthroplasties were enrolled prospectively. Ten patients with primary arthroplasties were included as negative controls. Synovial fluid was collected for mNGS analysis. Optimal thresholds were determined to distinguish pathogens from background microbes. Synovial fluid, tissue, and sonicate fluid were obtained for culture. Results. A total of 49 PJI and 21 noninfection patients were finally included. Of the 39 culture-positive PJI cases, mNGS results were positive in 37 patients (94.9%), and were consistent with culture results at the genus level in 32 patients (86.5%) and at the species level in 27 patients (73.0%). Metagenomic next-generation sequencing additionally identified 15 pathogens from five culture-positive and all ten culture-negative PJI cases, and even one pathogen from one noninfection patient, while yielding no positive findings in any primary arthroplasty. However, seven pathogens identified by culture were missed by mNGS. The sensitivity of mNGS for diagnosing PJI was 95.9%, which was significantly higher than that of comprehensive culture (79.6%; p = 0.014). The specificity is similar between mNGS and comprehensive culture (95.2% and 95.2%, respectively; p = 1.0). Conclusion. Metagenomic next-generation sequencing can effectively identify pathogens from synovial fluid of PJI patients, and demonstrates high accuracy in diagnosing PJI. Cite this article: Bone Joint Res 2020;9(7):440–449

Aims. The lack of disease-modifying treatments for osteoarthritis (OA) is linked to a shortage of suitable biomarkers. This study combines multi-molecule synovial fluid analysis with machine learning to produce an accurate diagnostic biomarker model for end-stage knee OA (esOA). Methods. Synovial fluid (SF) from patients with esOA, non-OA knee injury, and inflammatory knee arthritis were analyzed for 35 potential markers using immunoassays. Partial least square discriminant analysis (PLS-DA) was used to derive a biomarker model for cohort classification. The ability of the biomarker model to diagnose esOA was validated by identical wide-spectrum SF analysis of a test cohort of ten patients with esOA. Results. PLS-DA produced a streamlined biomarker model with excellent sensitivity (95%), specificity (98.4%), and reliability (97.4%). The eight-biomarker model produced a fingerprint for esOA comprising type IIA procollagen N-terminal propeptide (PIIANP), tissue inhibitor of metalloproteinase (TIMP)-1, a disintegrin and metalloproteinase with thrombospondin motifs 4 (ADAMTS-4), monocyte chemoattractant protein (MCP)-1, interferon-γ-inducible protein-10 (IP-10), and transforming growth factor (TGF)-β3. Receiver operating characteristic (ROC) analysis demonstrated excellent discriminatory accuracy: area under the curve (AUC) being 0.970 for esOA, 0.957 for knee injury, and 1 for inflammatory arthritis. All ten validation test patients were classified correctly as esOA (accuracy 100%; reliability 100%) by the biomarker model. Conclusion. SF analysis coupled with machine learning produced a partially validated biomarker model with cohort-specific fingerprints that accurately and reliably discriminated esOA from knee injury and inflammatory arthritis with almost 100% efficacy. The presented findings and approach represent a new biomarker concept and potential diagnostic tool to stage disease in therapy trials and monitor the efficacy of such interventions. Cite this article: Bone Joint Res 2020;9(9):623–632

Aims. The mechanism by which synovial fluid (SF) kills bacteria has not yet been elucidated, and a better understanding is needed. We sought to analyze the antimicrobial properties of exogenous copper in human SF against Staphylococcus aureus. Methods. We performed in vitro growth and viability assays to determine the capability of S. aureus to survive in SF with the addition of 10 µM of copper. We determined the minimum bactericidal concentration of copper (MBC-Cu) and evaluated its sensitivity to killing, comparing wild type (WT) and CopAZB-deficient USA300 strains. Results. UAMS-1 demonstrated a greater sensitivity to SF compared to USA300 WT at 12 hours (p = 0.001) and 24 hours (p = 0.027). UAMS-1 died in statistically significant quantities at 24 hours (p = 0.017), and USA300 WT survived at 24 hours. UAMS-1 was more susceptible to the addition of copper at four (p = 0.001), 12 (p = 0.005), and 24 hours (p = 0.006). We confirmed a high sensitivity to killing with the addition of exogenous copper on both strains at four (p = 0.011), 12 (p = 0.011), and 24 hours (p = 0.011). WT and CopAZB-deficient USA300 strains significantly died in SF, demonstrating a MBC-Cu of 50 µM against USA300 WT (p = 0.011). Conclusion. SF has antimicrobial properties against S. aureus, and UAMS-1 was more sensitive than USA300 WT. Adding 10 µM of copper was highly toxic, confirming its bactericidal effect. We found CopAZB proteins to be involved in copper effluxion by demonstrating the high sensitivity of mutant strains to lower copper concentrations. Thus, we propose CopAZB proteins as potential targets and use exogenous copper as a treatment alternative against S. aureus. Cite this article: Bone Joint Res 2024;13(11):632–646

Aim. The cut-off values for synovial fluid leukocyte count and neutrophils differential (%PMN) for differentiating aseptic from septic failure in total knee arthroplasties were already defined in the past. Our goal was to determine the cut-off values for synovial fluid leukocyte count and %PMN in failed total hip arthroplasties (THA). Method. Patients undergoing revision THA were prospectively included. In perioperative assessment phase, synovial fluid leukocyte count and %PMN were determined. During the surgery, at least 4 intraoperative samples for microbiological and one for histopathological analysis were obtained. Infection was defined as presence of sinus tract, inflammation in histopathological samples, and ≥2 tissue and/or synovial fluid samples growing the same microorganism. Exclusion criteria were systemic inflammatory diseases, revision surgery performed less than 3 months from index surgery and insufficient tissue sampling. Receiver operating characteristic (ROC) curves were constructed to assess the diagnostic performance and Youden's J statistic was computed to identify optimal cut-off values. Results. During the study period (between June 2006 and June 2011) 227 revision THAs were performed by the senior author. 31 patients were excluded. 196 patients (mean age, 69 years; 68% females) with THA failure were included. Aseptic failure was diagnosed in 150 patients (76,5%) and THA infection was diagnosed in 46 patients (23,5%). Synovial fluid leukocyte counts were significantly higher in the infected group (median, 5.50×10. 6. leukocytes/ml range, 0.05 to 143.9×10. 6. leukocytes/mL) than in the aseptic group (median, 0.23×10. 6. cells/ml; range, 0 to 21.3×10. 6. leukocytes/ml, P<0,0001). The %PMN was also significantly higher in the infected group (median, 83%; range, 6% to 97%) than in the aseptic group (median, 27,5%; range, 0% to 94%, P<0,0001). A synovial fluid leukocyte count of > 1.54×10. 6. leukocytes/ml, had a sensitivity of 63%, specificity of 95%, positive and negative predictive values of 78% and 89%, respectively. A synovial fluid %PMN of > 64%, had a sensitivity of 65%, specificity of 93%, positive and negative predictive values of 73% and 90%, respectively. Conclusion. The synovial fluid leukocyte count of > 1.54×10. 6. leukocytes/ml and %PMN of > 64% are useful and reliable tests for excluding THA infection, having a negative predictive value of around 90%. This tests and calculated cut-off values are highly recommended in the diagnostic process of failed THAs

Abstract. OBJECTIVE. Knee varus malalignment increases medial knee compartment loading and is associated with knee osteoarthritis (OA) progression and severity. 1. Altered biomechanical loading and dysregulation of joint tissue biology drive OA progression, but mechanistic links between these factors are lacking. Subchondral bone structural changes are biomechanically driven, involve bone resorption, immune cell influx, angiogenesis, and sensory nerve invasion, and contribute to joint destruction and pain. 2. We have investigated mechanisms underlying this involving RANKL and alkaline phosphatase (ALP), which reflect bone resorption and mineralisation respectively. 3. and the axonal guidance factor Sema3A. Sema3A is osteotropic, expressed by mechanically sensitive osteocytes, and an inhibitor of sensory nerve, blood vessel and immune cell invasion. 4. Sema3A is also differentially expressed in human OA bone. 5. HYPOTHESIS: Medial knee compartment overloading in varus knee malalignment patients causes dysregulation of bone derived Sema3A signalling directly linking joint biomechanics to pathology and pain. METHODS. Synovial fluid obtained from 30 subjects with medial knee OA (KL grade II-IV) undergoing high tibial osteotomy surgery (HTO) was analysed by mesoscale discovery and ELISA analysis for inflammatory, neural and bone turnover markers. 11 of these patients had been previously analysed in a published patient-specific musculoskeletal model. 6. of gait estimating joint contact location, pressure, forces, and medial-lateral condyle load distribution in a published data set included in analyses. Data analysis was performed using Pearson's correlation matrices and principal component analyses. Principal Components (PCs) with eigenvalues greater than 1 were analysed. RESULTS. PC1 (32.94% of variation) and PC2 (25.79% of variation) from PCA analysis and correlation matrices separated patients according to correlated clusters of established inflammatory markers of OA pain and progression (IL6/IL8, r=0.754, p<0.001) and anti-inflammatory mediators (IL4/IL10, r=0.469, p=0.005). Bone turnover marker ALP was positively associated with KL grade (r=0.815, p=0.002) and negatively associated with IL10 (r=−0.402, p=0.018) and first peak knee loading pressures (r=−0.688, p=0.019). RANKL was positively associated with IL4 (r=0.489, p=0.003). Synovial fluid Sema3A concentrations showed separate clustering from all OA progression markers and was inversely correlated with TNF-α (r=−0.423, p=0.022) in HTO patients. Sema3A was significantly inversely correlated with total predicted force in the medial joint compartment (r=−0.621, p=0.041), mean (r=−0.63, p=0.038) and maximum (r=−0.613, p=0.045) calculated medial compartment joint pressures during the first phase and mean (r=−0.618, p=0.043) and maximum (r=−0.641, p=0.034) medial compartment joint pressures during midstance outputs of patient-specific musculoskeletal model. CONCLUSIONS. This study shows joint inflammatory status and mechanical overloading influence subchondral bone-remodelling. Synovial Sema3A concentrations are inversely correlated to patient-specific musculoskeletal model estimations of pathological medial overloading. This study reveals Sema3A as a biological mediator with capacity to induce OA pain and disease progression that is directly regulated by gait mechanical loading. Declaration of Interest. (b) declare that there is no conflict of interest that could be perceived as prejudicing the impartiality of the research reported:I declare that there is no conflict of interest that could be perceived as prejudicing the impartiality of the research project