Detection of synovial sepsis in horses using enzymes as biomarkers

Abstract

Background: Synovial sepsis is a commonly occurring, potentially career-ending or even lifethreatening orthopaedic emergency. Diagnosis of synovial sepsis is currently primarily based on synovial fluid analysis, which often leaves diagnostic ambiguity due to overlap of clinicopathologic parameters between septic and aseptic inflammatory synovitis. Objectives:To evaluate the reliability of lysozyme (LYS), myeloperoxidase (MPO) and elastase (ELT) as biomarkers for synovial sepsis in horses using a photometric assay to measure increased enzyme activity.Study design: Prospective, single-blinded, analytical, clinical study.Methods: Equine synovial samples were assigned to one of three groups: 1) healthy controls (n=10); 2) aseptic (n=27) and 3) septic synovitis (n=30). The enzyme activity assays (LYS, MPO, ELT) were compared with standard synovial fluid parameters and broad-range bacterial 16S rDNA PCR.Results: LYS and MPO activities were significantly different between septic synovial samples, and both aseptic and control samples (p<0.001, LYS: confidence interval [CI]: 2.25-3.41 resp. 2.21-3.8, MPO: CI 0.752-1.6 resp. 0.639-1.81). LYS achieved a 100% sensitivity and 100% specificity in differentiating between septic and aseptic (cut-off value 751.4) or control (cut-off: 484.6) samples (p<0.001). MPO reached 93.33% sensitivity, 100% specificity for distinguishing septic from control (cut-off value: 0.1254) synovial samples and 93.33% sensitivity, 81.48% specificity for discriminating between septic and aseptic (cut-off value: 0.1305) synovial samples (p<0.001).ELT activity could not be measured in any synovial sample. Both the LYS and the MPO measurements showed a highly significant correlation with PCR (LYS r= 0.79, MPO r = 0.69), synovial leukocyte count (LYS r= 0.752, MPO r = 0.571), % neutrophils (LYS r=0.751, MPO r = 0.663) and each other (r=0.744, all p<0.001).Main limitations: Variation in horses' signalment, affected synovial structures, and synovial fluid freezing times may have affected the discriminative power of this study. Conclusions:Increased MPO and LYS activities allow reliable, rapid diagnosis of synovial sepsis with high sensitivity and specificity.

Synovial Fluid Metabolome Can Differentiate between Healthy Joints and Joints Affected by Osteoarthritis in Horses

“…In recent years, omics sciences, from genomics downwards to metabolomics, have granted a wealth of new diagnostic approaches in clinical and biomedical studies thanks to continuous developments in analytical techniques and bioinformatics [ 12 , 13 ], these approaches have also impacted the study of joint diseases through the observation of the synovial fluid in equines as well as in humans [ 14 , 15 , 16 , 17 , 18 ].…”

Visualizing neutrophil extracellular traps in septic equine synovial and peritoneal fluid samples using immunofluorescence microscopy

“…Myeloperoxidase (MPO), which assists with the unfolding of chromatin and forms a major component of NETs, 66 , 76 has been studied in SF and PF samples of horses. Septic SF had significantly elevated MPO compared to aseptic synovitis and healthy control samples, 28 and MPO was also higher in PF from horses with necrotic intestines. 25 But MPO is not specific for NETs; it is normally present in resting neutrophils, 54 and can be increased with neutrophil degranulation and necrosis.…”

“…Bacterial culture is slow and can result in false negatives, and cytology has poor sensitivity for detecting infection. 1,2,28,64 There is also a delay in normalization of laboratory results despite resolution of infection; this has been shown for neutrophil percentage and total protein (TP) in SF samples, 38 and serum amyloid A in horses with synovial infection. 75 Myeloperoxidase (MPO), which assists with the unfolding of chromatin and forms a major component of NETs, 66,76 has been studied in SF and PF samples of horses.…”

“…25 But MPO is not specific for NETs; it is normally present in resting neutrophils, 54 and can be increased with neutrophil degranulation and necrosis. 28 , 42 Evaluation of septic peritoneal effusions from dogs has demonstrated increased NET markers, 46 and NETs were visualized with immunofluorescence. 42 We retrieved no cases of septic SF samples containing NETs in a search of Google Scholar, PubMed, CAB Abstracts, Web of Science, and Scopus, using the search terms: neutrophil extracellular traps, synovial fluid, synovitis, septic, infection, veterinary, canine, feline, ruminant, suggesting that this condition has not been reported in other veterinary species.…”

Review of glucocorticoid therapy in horses: Intra‐articular corticosteroids

“…A positive synovial fluid culture is the gold standard of diagnosis but is obtained in less than 50% of confirmed cases (Taylor et al, 2010). Differentiation between septic and nonseptic inflammatory arthritis is somewhat confused by the overlap between clinicopathological values; however, the presence of synovial fluid leucocytosis (>30 ×10 9 nucleated cells/L), neutrophilia (>80–90% neutrophils), hyperproteinaemia (>30 g/L), cytological evidence of sepsis such as neutrophil degeneration, presence of intracellular bacteria and increased serum amyloid A concentration all provide convincing evidence for diagnosis (Haralambus et al, 2021; Ludwig et al, 2016; Sinovich et al, 2020). Treatment strategies include arthroscopic evaluation, through‐and‐through needle lavage, arthrotomy and systemic and local antibiotic administration (Crosby et al, 2019).…”