Utility of bedside leucocyte esterase testing to rule out septic arthritis.

INTRODUCTION: Suspected septic arthritis is a common presentation to EDs. The underlying diagnosis is often non-infective pathology. Differentiating between aetiologies is difficult. A bedside test with high negative predictive value (NPV) may allow safe discharge of patients, reduce the time in the ED, hospital admission and associated costs. This study aims to evaluate the NPV of bedside leucocyte esterase (LE) in the assessment of these patients. METHODS: A prospective multicentre observational study of ED adult patients referred to orthopaedics with suspected native joint septic arthritis between October 2015 and April 2016. At three hospital sites in the Bristol region, the results of the LE test exposed to aspirated synovial fluid were recorded along with Gram stain, culture, haematinics and length of stay. A positive LE test was considered 2+ or 3+ leucocytes based on the test strip colour. Data were analysed to establish sensitivity, specificity, NPV and positive predictive value (PPV) against the gold standard 48-hour culture. We determined the potential number of inpatient bed-days that might be avoided using this bedside test. RESULTS: Eighty patients underwent joint aspiration. Five cases had positive 48-hour culture. All (5/5) infected cases showed ≥2+ LE, sensitivity of 100% (95% CI 47.8% to 100%) while the Gram stain was positive in only one case (sensitivity 20%, 95% CI 0.51% to 71.6%). Twenty-three LE were read negative or 1+, all with negative 48-hour culture results, resulting in an NPV of 100% (95% CI 82.1% to 1.00%) for a negative LE test. Specificity of a positive LE test was 30.7% (95% CI 20.5% to 42.45%) with PPV of 8.77% (95% CI 7.64% to 10.1%). It was calculated that 57 orthopaedic bed-days could have potentially been saved by immediately discharging those with a negative LE test. CONCLUSIONS: LE point-of-care testing for suspected septic arthritis of native joints has a high NPV. Implementation of LE may facilitate more rapid discharge of patients with negative results. This test has the potential to reduce diagnostic uncertainty and costs to the healthcare system.

Leukocyte esterase analysis in the diagnosis of joint infection: can we make a diagnosis using a simple urine dipstick?

PURPOSE: Analysis of joint fluid remains a key factor in the diagnosis of periprosthetic infection. Recent reports have shown that neutrophils in infected joint fluid release esterase, an enzyme that is a reliable marker for infection. Testing for leukocyte esterase is routinely done in the analysis of urine for the presence of urinary tract infection, by a simple "dipstick" method. We report our experience with this technique in the evaluation of patients suspected of having septic arthritis or periprosthetic joint infection (PJI) by comparing results of leukocyte esterase positivity with confirmed joint infection as defined by the American Academy of Orthopaedic Surgeons (AAOS). MATERIALS AND METHODS: We retrospectively reviewed leukocyte esterase test results performed on synovial fluid aspirated from 57 patients with prosthetic (52) and native (5) joints. Patients either presented with unexplained painful arthroplasties, routine testing of PROSTALAC (PROSthesis with Antibiotic-Loaded Acrylic Cement) orthopedic implants, or clinical suspicion of periprosthetic infection or septic arthritis. Synovial fluid was percutaneously aspirated using a standard technique. The patient age range was 31-91 years with a mean age of 69.1 years, consisting of 30 women (52.6 %) and 27 men (47.4 %). The "gold standard" for the presence or absence of infection at our institution and in the study group was based on the most recent recommendations of the AAOS. Positive culture remained the "gold standard" for native joint infection. RESULTS: Of the total 57 joints aspirated and included in the study, 20 (35.1 %) were read as positive (2+) on the leukocyte test strip and 37 (64.9 %) were read as negative (negative, trace, or 1+). PJI was diagnosed in 19 patients and native joint septic arthritis was identified in one patient. Sensitivities were excellent at 100 % with no false negatives in the entire cohort. There was one false positive in the periprosthetic group yielding a specificity, positive predictive value and negative predictive value of 97, 95, and 100 %, respectively. The results for the native joints showed markedly less specificity and positive predictive value at 50 and 33 %; however, its negative predictive value remained at 100 %. CONCLUSIONS: Our test results confirm that the leukocyte esterase test can accurately detect PJI and that it can be used as a part of the traditional PJI workup. In the assessment of native joints, its high negative predictive value suggests that it is a valuable tool in excluding native joint septic arthritis.

Disordered lymphoid purine metabolism contributes to the pathogenesis of persistent Borrelia garinii infection in mice.

Extracellular ATP and adenosine are important regulators of immune responses; however, contribution of purinergic signaling to host defense during persistent microbial infections remains obscure. Lyme borreliosis is a common arthropod-borne infection caused by Borrelia burgdorferi sensu lato. In this study, we investigated whether lymphoid purinergic signaling contributes to the mechanisms by which borreliae species evade the immune system and trigger joint inflammation. Intracutaneous inoculation of Borrelia garinii to C3H/He mice induced symptomatic infection manifested in elevated levels of borrelia-specific IgG Abs, persistent spirochete dissemination into the tissues and joint swelling, as well as approximately 2- to 2.5-fold enlargement of draining lymph nodes with hyperplasia of B cell follicle area and L-selectin shedding from activated T lymphocytes. Purine catabolism was also activated in lymph nodes but not spleen and blood of infected C3H/He mice within the first 4 postinfection weeks, particularly manifested in transient upregulations of adenosine triphosphatase/ectonucleoside triphosphate diphosphohydrolase and ecto-5'-nucleotidase/CD73 on CD4(+)CD8(+) T lymphocytes and adenosine deaminase activity on B220(+) B lymphocytes. Compared with borrelia-susceptible C3H/He strain, lymphocytes from C57BL/6 mice displayed markedly enhanced adenosine-generating capability due to approximately three times higher ratio of ecto-5'-nucleotidase to adenosine deaminase. Borrelia-infected C57BL/6 mice efficiently eradicated the inoculated spirochetes at more chronic stage without any signs of arthritis. Strikingly, deletion of key adenosine-generating enzyme, ecto-5'-nucleotidase/CD73, was accompanied by significantly enhanced joint swelling in borrelia-infected CD73-deficient C57BL/6 mice. Collectively, these data suggest that insufficient basal adenosine level and/or pathogen-induced disordered lymphoid purine homeostasis may serve as important prerequisite for promotion of inflammatory responses and further host's commitment to persistence of bacterial infection and arthritis development.

Potential role of fibroblast-like synoviocytes in joint damage induced by Brucella abortus infection through production and induction of matrix metalloproteinases.

Arthritis is one of the most common complications of human brucellosis, but its pathogenic mechanisms have not been elucidated. Fibroblast-like synoviocytes (FLS) are known to be central mediators of joint damage in inflammatory arthritides through the production of matrix metalloproteinases (MMPs) that degrade collagen and of cytokines and chemokines that mediate the recruitment and activation of leukocytes. In this study we show that Brucella abortus infects and replicates in human FLS (SW982 cell line) in vitro and that infection results in the production of MMP-2 and proinflammatory mediators (interleukin-6 [IL-6], IL-8, monocyte chemotactic protein 1 [MCP-1], and granulocyte-macrophage colony-stimulating factor [GM-CSF]). Culture supernatants from Brucella-infected FLS induced the migration of monocytes and neutrophils in vitro and also induced these cells to secrete MMP-9 in a GM-CSF- and IL-6-dependent fashion, respectively. Reciprocally, culture supernatants from Brucella-infected monocytes and neutrophils induced FLS to produce MMP-2 in a tumor necrosis factor alpha (TNF-α)-dependent fashion. The secretion of proinflammatory mediators and MMP-2 by FLS did not depend on bacterial viability, since it was also induced by heat-killed B. abortus (HKBA) and by a model Brucella lipoprotein (L-Omp19). These responses were mediated by the recognition of B. abortus antigens through Toll-like receptor 2. The intra-articular injection of HKBA or L-Omp19 into the knee joint of mice resulted in the local induction of the proinflammatory mediators MMP-2 and MMP-9 and in the generation of a mixed inflammatory infiltrate. These results suggest that FLS, and phagocytes recruited by them to the infection focus, may be involved in joint damage during brucellar arthritis through the production of MMPs and proinflammatory mediators.

Synovial fluid adenosine deaminase activity to diagnose tuberculous septic arthritis.

There are reports of a correlation between high adenosine deaminase (ADA) levels in body fluid and tuberculosis (TB) infection, but none have evaluated synovial fluid ADA and TB arthritis. The objectives of this study were to determine the proper cut-off level for synovial fluid adenosine deaminase (SF-ADA) and the sensitivity and specificity of SF-ADA to diagnose TB arthritis. Between January 2006 and December 2007, SF-ADA were determined using the modified Giusti's method on patients over 15 years of age with clinically suspected TB arthritis or having an unknown etiology of their arthritis. Synovial fluid culture for TB was performed in all patients as a gold standard test. Forty cases were included in the study, with a female to male ratio of 1.7:1 and a mean age of 52.3 +/- 17.4 years (range, 16-80). The median duration of symptoms was 60 days. The prevalence of TB arthritis was 16.7% (6 cases) while the remaining cases were rheumatoid arthritis (8), non-TB bacterial septic arthritis (3), and miscellaneous (23). The mean SF-ADA levels in patients with TB arthritis and non-TB arthritis were 35.7 +/- 10.4 (range, 20-51) and 15.4 +/- 9 (range, 2-34) U/1, respectively. The cut-off value for the diagnosis of TB arthritis was 31 U/1, with a sensitivity of 83.3% (95% CI 35.9-99.6), a specificity of 96.7% (95% CI 82.8-99.9) and an agreement Kappa of 0.8 (p < 0.001). SF-ADA levels higher than 31 U/1 were highly correlated with a diagnosis of TB arthritis, with a high sensitivity and specificity. SF-ADA may be considered as a less invasive and time-consuming diagnostic tool for TB arthritis.

Streptococcus pyogenes degrades extracellular matrix in chondrocytes via MMP-13.

Group A streptococcus (GAS) causes a wide range of human diseases, including bacterial arthritis. The pathogenesis of arthritis is characterized by synovial proliferation and the destruction of cartilage and subchondral bone in joints. We report here that GAS strain JRS4 invaded a chondrogenic cell line ATDC5 and induced the degradation of the extracellular matrix (ECM), whereas an isogenic mutant of JRS4 lacking a fibronectin-binding protein, SAM1, failed to invade the chondrocytes or degrade the ECM. Reverse transcription-PCR and Western blot analysis revealed that the expression of matrix metalloproteinase (MMP)-13 was strongly elevated during the infection with GAS. A reporter assay revealed that the activation of the AP-1 transcription factor and the phosphorylation of c-Jun terminal kinase participated in MMP-13 expression. These results suggest that MMP-13 plays an important role in the destruction of infected joints during the development of septic arthritis.

Measurement of equine myeloperoxidase (MPO) activity in synovial fluid by a modified MPO assay and evaluation of joint diseases - an initial case study.

The aim of this study was to develop a specific myeloperoxidase (MPO) activity assay in the synovial fluid of horses and investigate whether MPO activity is increased in different forms of joint diseases. Synovial fluid samples were taken from affected joints from horses with osteoarthritis, chronic non-septic arthritis and septic arthritis, and from healthy control horses. MPO activity was measured using a specific modified o-dianisidine-assay containing 4-aminobenzoic acid hydrazide as a potent and specific inhibitor of the MPO. This assay is characterized by high reproducibility. The results reveal only a slight elevation of MPO activity in the synovial fluid of horses with osteoarthritis and chronic non-septic arthritis. However, in the cases of septic arthritis a significant increase in MPO activity was found when compared to the controls. In conclusion the first field study suggests that synovial fluid MPO may be used as a marker for septic arthritis in horses.

Relative expression of matrix metalloproteinase-2 and -9 in synovial fluid from healthy calves and calves with experimentally induced septic arthritis.

OBJECTIVE: To identify changes over time in relative expression of matrix metalloproteinase-2 (MMP-2) and -9 (MMP-9) in synovial fluid from healthy calves and calves with experimentally induced septic arthritis. ANIMALS: 12 Holstein calves. PROCEDURES: In 7 calves, Escherichia coli was injected in the right tarsal joint on day 1. Joint lavage was performed on day 2, and calves were treated with ceftiofur from days 2 through 21. Synovial fluid samples were collected on days 1 (before inoculation), 2 (before joint lavage), 3, 4, 8, 12, 16, 20, and 24. In the remaining 5 calves, joint lavage was performed on day 2 and synovial fluid samples were collected from the left tarsal joint. Relative expression of MMP-2 and MMP-9 was determined by means of gel zymography. RESULTS: On day 1, MMP-2 was detected in all synovial fluid samples but MMP-9 was not detected. In calves with septic arthritis, values for relative expression of MMP-9 monomer and dimer were significantly increased on days 2 through 20 and days 2 through 24, respectively, and relative expression of MMP-2 was significantly increased on days 3 through 20. There were significant linear associations between relative expression of the monomer and dimer forms of MMP-9 and between neutrophil count and relative expression of the MMP-9 monomer and dimer forms. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicated that relative expression of MMP-9 and MMP-2 increased in synovial fluid from calves with experimentally induced septic arthritis, with relative expression remaining high for several days after infection.

Use of matrix metalloproteinases 2 and 9 and white blood cell counts in monitoring the treatment and predicting the survival of horses with septic arthritis.

Thirty-nine samples of synovial fluid were collected from the joints of 32 horses with suspected septic arthritis and 39 samples were collected from horses euthanased for non-orthopaedic conditions. The white blood cell counts (WBCC) were determined and the pro and active forms of matrix metalloproteinases (MMPs) 2 and 9 were measured by gelatin zymography and image analysis in each sample. The initial measurements of the ratio of proMMP9:proMMp2 and WBCC were good prognostic indicators of the survival of the horses. There was no significant relationship between the interval between the injury and the horse being referred for treatment and either the WBCC or the levels of MMP2 and MMP9 initially, and no evidence that this interval significantly affected the chances of the horses surviving.

Effectiveness of leucocyte esterase as a diagnostic test for acute septic arthritis.

BACKGROUND: We hypothesized that leucocyte esterase strip test can aid in diagnosing septic arthritis in native synovial fluid because leucocyte esterase concentrations would be elevated at the infection site because of secretion by recruited neutrophils. METHOD: The cohort included 27 patients (suspected septic arthritis and normal subjects). A standard chemical test strip (graded as negative, trace, +, ++ or +++) was used to detect the presence of leucocyte esterase. Fluid leucocyte count, Gram staining, culture, erythrocyte sedimentation rate and C-reactive protein were also assessed. RESULTS: The leucocyte esterase test with a threshold of ++/+++ had a sensitivity of 79.2% (95% CI [confidence interval], 65.9% to 89.2%), specificity of 80.8% (95% CI, 73.3% to 87.1%), positive predictive value (PPV) of 61.8% (95% CI, 49.2% to 73.3%) and negative predictive value (NPV) of 90.1% (95% CI, 84.3% to 95.4%). CONCLUSION: The leucocyte esterase strip test yielded a high specificity, PPV, NPV, high sensitivity and high diagnostic accuracy. Leucocyte esterase is an accurate, quick and bedside test for septic arthritis and can be used effectively for diagnosing periprosthetic joint infections along with other battery of tests according to the Musculoskeletal Infection Society criteria.

Matrix metalloproteinase-9 (gelatinase B) deficiency leads to increased severity of Staphylococcus aureus-triggered septic arthritis.

Matrix metalloproteinases constitute a family of structurally related endopeptidases that are crucial for the normal turnover of the extracellular matrix. Elevated levels of MMP-9 have been demonstrated in synovial fluids of rheumatoid arthritis patients, and a correlation with the severity of the disease has been described. The aim of this study was to explore the impact of MMP-9 expression on joint inflammation and destruction in a model of bacterially induced septic arthritis. MMP-9 knock-out mice and C57Bl6 congenic controls were inoculated intravenously or intra-articularly with Staphylococcus aureus. Arthritis was evaluated clinically and by means of histology. Zymographic analyses were performed to study ex vivo induction of MMP-9 following exposure to S. aureus. The MMP-9 knock-out mice displayed a significantly higher frequency and severity, but not destructivity, of arthritis than did the wild-type mice. The knock-out mice also proved to harbour an increased number of bacteria locally in joints and systemically in kidneys, possibly by impaired extravasation and recruitment of leukocytes and a deficient early defence against infection. Our findings indicate that deficiency in MMP-9 increases the degree of joint inflammation due to decreased bacterial clearance.

Synovial fluids from infected joints contain metalloproteinase--tissue inhibitor of metalloproteinase (TIMP) complexes.

Samples of synovial fluids aspirated from patients with septic arthritis prior to the commencement of any treatment contained active metalloproteinases but no proteinase inhibitory activity. We therefore assayed these samples for proteinase-inhibitor complexes. Although no biologically active alpha 2-macroglobulin or tissue inhibitor of metalloproteinase (TIMP) was present in the fluids, immunoassay of the samples clearly showed that high molecular weight proteinase-TIMP complexes were present. It is proposed that high levels of active metalloproteinases are released from neutrophils into septic synovial fluids and that these proteinases complex all the available TIMP, forming metalloproteinase-TIMP complexes.

Lipoxygenase products in experimental septic arthritis in dogs.

Induction of an experimental septic arthritis was performed in the juvenile dog knee by an intra-articular injection with Staphylococcus aureus. From plasma, lipoxygenase (LO) products of arachidonic acid were separated by reversed-phase high-performance liquid chromatography (RP-HPLC) after extraction of lipids. The plasma samples did not contain UV-detectable amounts of LO products other than the LTB4 metabolite 20-COOH-LTB4, and 12-HETE. Small amounts of chemokinetically active material were found coeluting with the eluate fraction of 20-COOH-LTB4, LTB4 and 12-HETE after RP-HPLC. After 48 hours of septic arthritis, resulting in a massive acute joint inflammation, no significant changes in LO products were observed.

Up-regulation of inducible nitric oxide synthase mRNA in dogs experimentally infected with Borrelia burgdorferi.

The up-regulation of the inducible nitric oxide synthase (iNOS) mRNA was determined by RT-PCR in 25 tissues each from 22 specific pathogen-free (SPF) dogs experimentally infected with Borrelia burgdorferi by tick exposure and from five uninfected control dogs. Using primers specific for a homologous region of the human and canine iNOS sequence, and canine macrophage mRNA, we isolated and partially sequenced canine iNOS. A sequence of 1775 bases was obtained and primers specific for canine iNOS mRNA constructed to investigate the expression of iNOS in dog tissues in response to infection with B. burgdorferi. In 12 out of 22 dogs infected with B. burgdorferi, acute lameness occurred within 55-82 days after infection whereas the other 10 dogs showed no or only mild clinical signs despite persistent infection up to Day 175. The numbers of iNOS mRNA-positive tissues in dogs with acute lameness were significantly higher than in dogs without lameness, while uninfected dogs showed only negligible iNOS expression. Dogs with acute lameness also had higher numbers of borrelia-positive tissues as well as higher scores in histopathological evaluations than infected dogs without lameness. Our results show that the expression of iNOS mRNA is related to the number of B. burgdorferi-positive tissues and the severity of inflammation as assessed by histopathology. These results implicate an up-regulation of the iNOS mRNA as part of the host's immune response to borrelia infection and a possible role for NO in the pathogenesis of canine Lyme arthritis.

Penicillinase-producing gonococcal arthritis in a community hospital.

Disseminated Neisseria gonorrhoeae gonococcal infections are the leading cause of acute arthritis in young adults. Recent published information indicates that a small proportion of gonococcal arthritis is caused by penicillinase-producing Neisseria gonorrhoeae (PPNG). This article reports three cases of PPNG over a 12-month period and recommends that all suspected cases of gonococcal arthritis be treated as if they were PPNG until proven otherwise.

Proteolytic enzymes in equine joints with infectious arthritis.

Significant amounts of collagenase and caseinase activity were detected in infected synovial fluid samples. Partial characterisation of the enzymes by gel filtration suggested that synovial fluid from cases of infectious arthritis may contain enzymes from both the synovial cells and neutrophils. This finding was also supported by analysis of sequential synovial fluid samples from 4 infected joints. In 3 joints the concentration of caseinase and in 1 joint collagenase paralleled the decline in total nucleated cell count. However, in 3 joints the concentration of collagenase remained high after the total nucleated cell count had returned to normal, suggesting that this enzyme originated from resident articular cells.

Erysipelas arthritis in swine: lysosomal enzyme levels in synovial fluids.

Arthritic and histologically normal joints from swine in which arthritis had been produced by the intravenous inoculation of Erysipelothrix rhusiopathiae were used as a source of fluids for lysosomal enzyme determinations. Mean lysozyme activities in synovias from arthritic and histologically normal joints were 16.60 and 5.79 mug/ml, respectively. Acid phosphatase (ACP) was increased more than 8 times the activity in histologically normal joints, but there was no relationship between lysozyme and ACP, indicating the probability that 1 of these enzymes came from another source. The cytoplasmic enzyme, lactic dehydrogenase (LDH), was increased in proportion to ACP, indicating that cell death and not selective extrusion of lysosomal enzymes during phagocytosis was an important mechanism of enzyme release in arthritic joints. Lysozyme activities in synovias from histologically normal joints were often increased above companion serum concentrations, indicating the enzyme has a special role in the joint. Also, the ratio of activities of lysozyme to ACP in pig buffy coat lysates was different from the ratio of the 2 enzymes in synovias from arthritic joints.

[Importance of enzymes in the synovial fluid. Progress report]. / Importanza degli enzimi nel liquido sinoviale. Nota di aggiornamento

The significance and importance of investigation of the synovial fluid enzymes in the main arthropathies are explanined. Tables are given for the main enzymes studied, the cell compartments of origin, and data for their values in rheumatic diseases (as reported in the literature). Stress is laid on the importance of enzymes belonging to the lysosomial compartment, both in the pathogenesis of the underlying inflammation and in the relation to anatomopathological lesions in the joints. Attention is directed to the most widely accepted hypotheses. These ten to see enzymes increases in breakdown of condrocytes, as inflammatory arthritis attributable to synoviocytes and leukocytes. A personal opinion based on prior research is also presented. Further work in this sector is urged a mean of learning more about the pathology of rheumatic diseases.

[Activity of lysosomal exoglycosidases in serum of patients with chronic borrelia arthritis]. / Aktywnosc egzoglikozydaz lizosomalnych w surowicy pacjentów z boreliozowym przewleklym zapaleniem stawów.

To estimate activities of lisosomal exoglycosidases in serum of patients with chronic borrelia arthritis. Study group consisted of 18 patients aged 18-72 years (x=46) hospitalized in Department of Infectious Diseases and Neuroinfections of Medical Academy in Bialystok with diagnosis of chronic arthritis in course of borreliosis. Control consisted of 20 healthy volunteers (health services employees) aged 25-65 years (x=45), with no detectable anti-Borrelia burgdorferi antibodies in serum. In all borreliosis patients serum activity of: N-acetyl-beta-D-glucosaminidase (HEX), beta-galactosidase and alpha-mannosidase was measured before and after 4 weeks of doxycycline treatment. Results were analyzed with Statistica 6.0 software. P < 0.05 was considered statistically significant. HEX activity was significantly increased in serum of Lyme arthritis patients before treatment compared to controls. It decreased after 4-week treatment, remaining insignificantly higher than in controls. b-galactosidase and a-mannosidase activities in serum of Lyme arthritis patients were insignificantly higher than in controls and fell after treatment to the levels observed in control group. N-acetyl-beta-D-glucosaminidase (HEX) is sensitive enzymatic marker of Lyme arthritis. It may be used to monitor course of the disease and its efficiency of treatment.

Leucocyte esterase in the rapid diagnosis of paediatric septic arthritis.

Septic arthritis may affect any age group but is more common in the paediatric population. Infection is generally bacterial in nature. Prompt diagnosis is crucial, as delayed treatment is associated with lifelong joint dysfunction. A clinical history and application of Kocher's criteria may indicate that there is a septic arthritis. However, definitive diagnosis is made on culture of septic synovial fluid. The culture process can take over 24h for the initial culture to yield bacterial colonies. Leucocyte esterase is released by leucocytes at the site of an infection. We hypothesise that leucocyte esterase can be utilized in the rapid diagnosis of septic arthritis and shorten the time to decisive treatment whilst simultaneously decreasing unnecessary treatment of non-septic joints.