Biomarkers of fever: from bench to bedside

__Abstract__ This thesis aims to study biomarkers in inflammation and infection, with a special focus on the distinction between infectious and non-infectious fever. The thesis consists of three parts, part I being this introduction, in which the concept of fever in infectious and non-infectious disease is discussed. Furthermore, in this part we provide an overview of the epidemiology of febrile disease, as studied both in a general hospital in the Netherlands and in a general hospital in Curaçao. Also, a review of current literature on biological markers in non-infectious fever is given. Part II describes our clinical studies with focus on biomarkers in different cohorts of patients with infectious and non-infectious fever. In part III, we summarize the findings of this thesis and discuss future research

Epidemiology of febrile diseases in the emergency department of a Caribbean Island: The Curaçao experience

Objective: The aetiology of febrile diseases in tropical countries often remains poorly characterized. We aim to describe the aetiology and outcome of febrile illnesses at the Emergency Department (ED) in Curaçao. Methods: From April 2008 - April 2009, all adult febrile patients (T > 38.5 oC) at the ED of the St Elisabeth Hospital, Curaçao, Netherlands Antilles, were included. Clinical data were recorded, routine laboratory measurements and blood cultures were taken. Final diagnoses were made at discharge by an independent physician and in retrospect by the main investigator. Results: Four hundred and three patients were included: 223 patients (55.6%) were hospitalized, 32 patients (7.9%) died and 18 patients (4.5%) were admitted to the Intensive Care Unit. In 129 febrile patients (32.0%), infection was proven; 84.4% of patients had bacterial (29.0% urinary tract infection, 23.2% pneumonia infection), 5.6% viral and 10.0% parasitic or fungal infections. Twenty-one patients (5.2%) were discharged with a non-infectious diagnosis and 172 patients (42.7%) without a clear diagnosis. Conclusion: A high mortality rate of 7.9% was observed. We found a high prevalence of bacterial infections, with pneumonia and urinary tract infections as the most common causes of fever. One in 20 patients did not have an infectious disease

Immunomodulation Through Low-Dose Radiation for Severe COVID-19:Lessons From the Past and New Developments

Low-dose radiation therapy (LD-RT) has historically been a successful treatment for pneumonia and is clinically established as an immunomodulating therapy for inflammatory diseases. The ongoing COVID-19 pandemic has elicited renewed scientific interest in LD-RT and multiple small clinical trials have recently corroborated the historical LD-RT findings and demonstrated preliminary efficacy and immunomodulation for the treatment of severe COVID-19 pneumonia. The present review explicates archival medical research data of LD-RT and attempts to translate this into modernized evidence, relevant for the COVID-19 crisis. Additionally, we explore the putative mechanisms of LD-RT immunomodulation, revealing specific downregulation of proinflammatory cytokines that are integral to the development of the COVID-19 cytokine storm induced hyperinflammatory state. Radiation exposure in LD-RT is minimal compared to radiotherapy dosing standards in oncology care and direct toxicity and long-term risk for secondary disease are expected to be low. The recent clinical trials investigating LD-RT for COVID-19 confirm initial treatment safety. Based on our findings we conclude that LD-RT could be an important treatment option for COVID-19 patients that are likely to progress to severity. We advocate the further use of LD-RT in carefully monitored experimental environments to validate its effectiveness, risks and mechanisms of LD-RT

Routine Lupus Anticoagulant Sensitive aPTT Testing Can Prevent Unnecessary LA Testing

Even though routine screening of the general hospital population is discouraged, medical laboratories may use a “lupus sensitive” activated partial thromboplastin time test (aPTT) with phospholipid concentrations that are susceptible to inhibition by lupus anticoagulant (LA), to screen for the presence of LA. If deemed necessary, follow-up testing according to ISTH guidelines may be performed. However, LA testing is a laborious and time-consuming effort that is often not readily available due to a lack of automation and/or temporary unavailability of experienced staff. In contrast, the aPTT is a fully automated test that is available 24/7 in almost all medical laboratories and is easily interpreted with the use of reference ranges. In addition to clinical signs, the result of an LA sensitive aPTT may thus be used to lower the suspicion of the presence of LA and reduce costly follow-up testing. In this study, we show that a normal LA sensitive aPTT result may be safely used to refrain from LA testing in the absence of strong clinical suspicion

Higher diagnostic accuracy and cost-effectiveness using procalcitonin in the treatment of emergency medicine patients with fever (The HiTEMP study)

__Background:__ Fever is a common symptom in the emergency department(ED). Fever can be caused by bacterial infections, which are treated with antibiotics. Often, bacterial infections cannot be ruled out in the ED using standard diagnostics, and empiric antibiotic treatment is started. Procalcitonin(PCT) is a biomarker for bacterial infections, but its role in an undifferentiated ED population remains unclear. We hypothesize that PCT-guided therapy may reduce antibiotics prescription in undifferentiated febrile ED patients. The primary objectives of this study are to determine a) the efficacy, b) the safety of PCT-guided therapy, and c) the accuracy of the biomarker PCT for bacterial infections. The secondary objective is to study the cost-effectiveness of PCT-guided therapy. __Methods/design:__ This is a multicenter noninferiority randomized controlled trial. All adult ED patients with fever(≥38.2 °C) are randomized between standard care with and without the addition of a PCT level, after written informed consent. a) For efficacy, the reduction of patients receiving antibiotics is calculated, using a superiority analysis: differences between the PCT-guided group and control group are assessed using a Fisher's exact test, and a multivariable logistic regression analysis to account for the effects of demographic and medical variables on the percentage of febrile patients receiving antibiotics. b) Safety consists of a composite endpoint, defined as mortality, intensive care admission and ED return visit within 14 days. Noninferiority of PCT will be tested using a one-sided 95 % confidence interval for the difference in the composite safety endpoint between the PCT-guided and control groups using a noninferiority margin of 7.5 %. c) Accuracy of PCT and CRP for the diagnosis of bacterial infections will be reported, using the sensitivity, specificity, and the area under the receiver-operating-characteristic curve in the definitive diagnosis of bacterial infections. The sample size is 550 patients, which was calculated using a power analysis for all primary objectives. Enrollment of patients started in August 2014 and will last 2 years. __Discussion:__ PCT may offer a more tailor-made treatment to the individual ED patient with fever. Prospective costs analyses will reveal the economic consequences of implementing PCT-guided therapy in the ED. This trial is registered in the Dutch trial register:NTR4949

Pregnancy outcome predictors in antiphospholipid syndrome: A systematic review and meta-analysis

Objective: To identify and assess the magnitude of effect of pregnancy outcome predictors in women with antiphospholipid syndrome (APS) by means of systematic review and meta-analysis. Methods: PubMed and Embase were searched (13th June 2020) for studies reporting on pre-pregnancy risk factors of pregnancy outcomes in APS patients. Literature screening and data extraction were conducted by two reviewers independently, in a blinded standardized manner. Pooled univariate odds ratios (OR) were computed using a random effects model. Heterogeneity was assessed by I2%. Results: The search yielded 3013 unique results; 27 records were included in this meta-analysis. Previous thrombosis was associated with a decreased live birth risk (OR 0.60, p < 0.01, I2 = 40%), increased neonatal mortality (OR 15.19, p < 0.01, I2 = 0%), an increased risk of antenatal or postpartum thrombosis (OR 6.26, p < 0.01, I2 = 0%) and an increased risk of delivering a small for gestational age neonate (SGA) (OR 2.60, p = 0.01, I2 = 0%). Patients with an APS laboratory category I (double or triple positivity) profile had a decreased live birth risk (OR 0.66, p < 0.01, I2 = 0%), an increased risk of SGA (OR 1.86, p = 0.01, I2 = 43%) and preterm birth (OR 1.35, p < 0.01, I2 = 49%). Triple positivity was associated with a decreased live birth risk (OR 0.33, p < 0.01, I2 = 68%), an increased risk of preeclampsia (OR 2.43, p = 0.02, I2 = 35%) and SGA (OR 2.47, p = 0.04, I2 = 61%). Patients with lupus anticoagulant positivity had an increased risk of preeclampsia (OR 2.10, p = 0.02, I2 = 48%), SGA (OR 1.78, p < 0.01, I2 = 0%) and preterm birth (OR 3.56, p = 0.01, I2 = 48%). Risk of bias assessment suggested considerable bias on study participation and statistical methods. Conclusions: The results of this meta-analysis identified previous thrombosis, laboratory category I, triple positivity and lupus anticoagulant positivity as the most important predictors of adverse pregnancy outcomes. This up-to-date knowledge, can be used in preconception counseling and tailoring of obstetric care

Interference in point-of-care international normalized ratio monitoring in patients with lupus anticoagulant is correlated with anti–β2-glycoprotein I antibody titers

Background: Patients with antiphospholipid syndrome (APS) receive anticoagulant therapy with vitamin K antagonists (VKAs) to prevent recurrent thrombosis. VKA treatment requires strict monitoring with an international normalized ratio (INR). It is known that lupus anticoagulants (LAs) can lead to elevated INR results with point-of-care-testing (POCT) devices, which could result in inadequate adaptation of anticoagulant therapy. Objective: To determine discrepancies between POCT-INR and laboratory-INR in patients who are LA-positive on VKA therapy. Methods: Paired INR testing was performed with 1 POCT device (CoaguChek XS) and 2 laboratory assays (Owren and Quick method) in 33 patients with LA-positive APS on VKA in a single-center cross-sectional study. Patients were tested for anti–β2-glycoprotein I, anticardiolipin, and antiphosphatidylserine/prothrombin immunoglobulin (Ig) G and IgM antibodies. Agreement between assays was evaluated with Spearman's correlation, Lin's correlation coefficient, and Bland–Altman plots. Agreement limits were considered satisfactory if differences were ≤20% as determined by the Clinical and Laboratory Standards Institute. Results: We found poor agreement between POCT-INR and laboratory-INR based on Lin's concordance correlation coefficient (ρc) of 0.42 (95% CI, 0.26-0.55) between POCT-INR and Owren-INR, a ρc of 0.64 (95% CI, 0.47-0.76) between POCT-INR and Quick-INR, and a ρc of 0.77 (95% CI, 0.64-0.85) between Quick-INR and Owren-INR. High anti-β2-glycoprotein I IgG antibody titers correlated with INR disagreement between POCT-INR and laboratory-INR. Conclusion: There is a disagreement between INR values measured with the CoaguChek XS and laboratory-INR in a proportion of patients with LA. Consequently, laboratory-INR monitoring should be preferred over POCT-INR monitoring in patients with LA-positive APS, especially in patients with high anti-β2-glycoprotein IgG antibody titers

Microarray testing in patients with systemic lupus erythematosus identifies a high prevalence of CpG DNA-binding antibodies

OBJECTIVE: Many autoantibodies are known to be associated with SLE, although their role in clinical practice is limited because of low sensitivity and weak associations with clinical manifestations. There has been great interest in the discovery of new autoantibodies to use in clinical practice. In this study, we investigated 57 new and known antibodies and their potential for diagnostics or risk stratification. METHODS: Between 2014 and 2017, residual sera of all anti-dsDNA tests in the UMC Utrecht were stored in a biobank. This included sera of patients with SLE, patients with a diagnosis of another immune-mediated inflammatory disease (IMID), patients with low (non-IMID) or medium levels of clinical suspicion of SLE but no IMID diagnosis (Rest), and self-reported healthy blood bank donors. Diagnosis and (presence of) symptoms at each blood draw were retrospectively assessed in the patient records with the Utrecht Patient-Oriented Database using a newly developed text mining algorithm. Sera of patients were analysed for the presence of 57 autoantibodies with a custom-made immunofluorescent microarray. Signal intensity cut-offs for all antigens on the microarray were set to the 95th percentile of the non-IMID control group. Differences in prevalence of autoantibodies between patients with SLE and control groups were assessed. RESULTS: Autoantibody profiles of 483 patients with SLE were compared with autoantibody profiles of 1397 patients from 4 different control groups. Anti-dsDNA was the most distinguishing feature between patients with SLE and other patients, followed by antibodies against Cytosine-phosphate-Guanine (anti-CpG) DNA motifs (p<0.0001). Antibodies against CMV (cytomegalovirus) and ASCA (anti-Saccharomyces cerevisiae antibodies) were more prevalent in patients with SLE with (a history of) lupus nephritis than patients with SLE without nephritis. CONCLUSION: Antibodies against CpG DNA motifs are prevalent in patients with SLE. Anti-CMV antibodies are associated with lupus nephritis

Recurrence risk of venous thromboembolism associated with systemic lupus erythematosus:A retrospective cohort study

BACKGROUND: Recurrence risk of systemic lupus erythematosus (SLE)‐associated venous thromboembolism (VTE) is unclear. AIM: To determine the recurrence risk of SLE‐associated VTE overall and by presence of provoking factors and SLE flares. METHODS: A multicenter, retrospective cohort study was conducted among patients with first SLE‐associated VTE who discontinued anticoagulation. SLE flares were defined as Systemic Lupus Erythematosus Disease Activity Index 2000 greater than 4. The primary outcome was recurrent VTE. Incidence rates and cumulative incidences were calculated by presence of provoking factors and antiphospholipid syndrome (APS) at index VTE. The hazard ratio (HR) for recurrence after SLE flare–associated index VTE was estimated with Cox regression, adjusted for provoking factor presence and APS. RESULTS: Eighty patients were included with 21 recurrent VTEs in median 8 years. For provoked index VTE, the recurrence rate in patients without APS was 1.1 per 100 person‐years (PY; 95% confidence interval [CI], 0.1–3.1) and in the presence of APS 3.5 per 100 PY (95% CI, 0.9–8.9), yielding cumulative incidences of 7.5% (95% CI, 1.2%–21.7%) and 31.4% (95% CI, 6.3%–61.6%) respectively. For unprovoked index VTE, these analogous rates were 3.8 per 100 PY (95% CI, 1.2–9.0) and 16.7 per 100 PY (95% CI, 4.5–42.7), with cumulative incidences of 33.7% (95% CI, 10.7%–58.9%) and 54.2% (95% CI, 10.7%–84.5%), respectively. Forty‐six index VTEs were flare associated, and the adjusted HR for recurrence was 0.4 (95% CI, 0.1–1.8) compared to those without flares at their index VTE. CONCLUSION: Antiphospholipid syndrome is the main determinant for recurrence risk of SLE‐associated VTE irrespective of presence of a provoking factor. Future research should attempt to confirm that flare‐associated VTE has a lower recurrence risk

Microarray analysis of autoantibodies can identify future Systemic Lupus Erythematosus patients

OBJECTIVE: Reliable early ascertainment in patients with SLE is important to prevent the accumulation of irreversible organ damage. Autoantibodies are often present in the serum of patients before the first symptoms arise, therefore they are of potential use as early diagnostic tools. METHODS: We used a custom-made antibody microarray containing 57 autoantigens to analyze serum samples of 1519 patients previously tested for anti-dsDNA and 361 samples of self-reported healthy blood bank donors (BBD). The 1519 patients included 483 patients with SLE, 346 patients with other immune mediated inflammatory diseases (IMID), 218 patient controls without relevant clinical symptoms (Non-IMID), and 472 patients that did not fit in any of the previous groups (Rest). The Non-IMID and BBD groups were used individually to create multivariable prediction models to distinguish samples of patients with SLE from these control groups. We subsequently used these models to predict the outcome for samples of patients who developed SLE while in follow-up (pre-SLE). RESULTS: Out of 1036 patients with no diagnosis of SLE at the moment of sample collection, 17 patients developed SLE while in follow-up (mean time to diagnosis 7.2 months). The best performing model (AUC 0.83) identified 9 out of 17 (53%) pre-SLE samples as SLE, with a specificity of 94%. CONCLUSION: Serum samples of patients who will develop SLE in the future already show a shift of the autoantibody profile prior to diagnosis. In this study, we show that these autoantibody profiles can be used to identify these future SLE patients