Bibliography

Can faecal calprotectin predict relapse in inflammatory bowel disease: a mini review

Crohn’s disease and ulcerative colitis are chronic inflammatory disorders affecting the gastrointestinal tract. Faecal calprotectin is a protein complex of the S-100 family of calcium-binding proteins present in inflammatory cells that can be measured in stool samples, which act as a biomarker for bowel inflammation. Elevated faecal calprotectin has been shown to reflect the presence of ongoing mucosal inflammation, which improves with mucosal healing. The aim of this review was to evaluate the available evidence on the ability of faecal calprotectin to predict a relapse in inflammatory bowel disease. Multiple retrospective studies have shown that patients who relapse have significantly higher levels of calprotectin in their stool compared with non-relapsers, especially in ulcerative colitis. Elevated faecal calprotectin postoperatively in Crohn’s disease was also shown to be indicative of a relapse. However, the association of a raised faecal calprotectin and relapse is not universal and may be explained by the different patterns of mucosal inflammatory activity that exist. In conclusion, we put forward our hypothesis that changes such as a rise in faecal calprotectin levels may be more predictive of a relapse than absolute values.

Serum Calprotectin: A Potential Biomarker for Neonatal Sepsis

Introduction: The correct diagnosis of neonatal sepsis is a relevant problem because sepsis is one of the most important causes of neonatal morbidity, mortality, and prolonged hospital stay. Calprotectin is an antimicrobial, calcium and zinc binding heterocomplex protein that could be used as a nonspecific marker for activation of granulocytes and mononuclear phagocytes. Calprotectin has been proposed for the diagnosis of inflammatory conditions. Our aim is to study serum calprotectin as a biomarker for neonatal sepsis diagnosis.

Methods: 41 (20 females, 21 males) infants who underwent blood culture due to suspected sepsis were enrolled in the study. Serum calprotectin was measured by a commercial ELISA assay (Calprest, Eurospital, Trieste, Italy). Statistical analysis was performed using the statistical software package Stata 13.1 (Stata Corporation, College Station, Texas, USA).

Results: 8 neonates (19.51%) showed sepsis with positive culture and 33 (80.49%) showed suspected sepsis. The optimal cut-off for calprotectin is 2.2 μg/mL with a sensitivity of 62.5% and a specificity of 69.7%.

Conclusions: Calprotectin may be considered a promising early, sensitive, specific marker of sepsis thanks to the importance of calprotectin in defense mechanisms and physiological functions of the immune system.

Specificity of a Polyclonal Fecal Elastase ELISA for CELA3

Introduction: Elastase is a proteolytic pancreatic enzyme that passes through the gastrointestinal tract undergoing only limited degradation. ELISA tests to determine stool elastase concentrations have therefore been developed for the diagnosis of exocrine pancreatic insufficiency. Five different isoforms of pancreatic elastase (CELA1, CELA2A, CELA2B, CELA3A, CELA3B) are encoded in the human genome. We have investigated three different polyclonal antisera that are used in a commercial fecal elastase ELISA to determine their specificity for different pancreatic elastase isoforms.

Material and methods: Different polyclonal rabbit antisera against human elastase peptides (BIOSERV Diagnostics GmbH, Germany) were tested by Western blot analysis of human pancreatic juice, in HEK-293 cells expressing Elastase constructs, and in the protein content of porcine pancreatin, used for treatment of exocrine pancreatic insufficiency.

Results: In human pancreatic juice the polyclonal antisera detected proteins at the corresponding size of human pancreatic elastase isoforms (~29kDa). Transiently expressed GFP fusion protein of elastase isoform CELA3A (CELA3A-GFP), but not CELA2A (CELA2A-GFP) could be precipitated from HEK-293 cell lysates with the elastase antisera. We detected no cross-reactivity with elastases in the porcine pancreatic extracts (pancreatin) used for enzyme replacement therapy.

Conclusion: The polyclonal antisera used in a commercial fecal elastase ELISA are specific for the human pancreatic elastase isoform CELA3 and do not cross-react with elastase contained in pig pancreatin. While pancreatic elastase 1 (CELA1) is not expressed in the adult human pancreas, possible differences between the other isoforms regarding their cellular expression, pathophysiological role and relevance in exocrine pancreatic insufficiency deserve further investigation.

Fecal Calprotectin: Diagnostic Accuracy of the Immunochromatographic CalFast Assay in a Pediatric Population

Background: Fecal calprotectin is a noninvasive marker for bowel diseases and it is high valuable to follow disease activity in Crohn’s disease (CD) and ulcerative colitis (UC). In this study, we evaluated the diagnostic performance of the recently introduced immunochromatographic assay CalFast in comparison to the well-known ELISA tests for calprotectin assay to obtain a rapid diagnosis of bowel inflammation in pediatric patients.

Methods: CalFast was tested in parallel to the classic ELISA tests CalPrest and PhiCal (gold standards for the calprotectin determination) on 148 fecal samples from pediatric subjects including 104 healthy subjects, 29 with CD, and 15 with UC.

Results: In this study, the sensitivity and specificity of CalFast, CalPrest, and PhiCal were 86.4%, 88.6%, and 93.2% and 86.6%, 74%, and 64.4%, respectively. The area under the curve, obtained from receiver operating characteristic analysis, indicated the lack of significant difference among all the kits used.

Conclusion: The immunochromatographic assay demonstrated good diagnostic predictive values, comparable to those of the ELISA methods, and may represent a valid alternative in order to save operators’ time. The test, in fact, has a short turnaround time and does not need a specific ELISA instrumentation.

Calprotectin in Daily Practice: Where Do We Stand in 2017?

Background: To make a distinction between organic and functional disease is essential for gastroenterologists in their daily practice, but it may be challenging, given the variety and aspecificity of gastrointestinal symptoms among the general population. The clinician aim is to avoid diagnostic delay and to restrict unnecessary invasive and expensive exams.

Summary: Faecal markers, in particular faecal calprotectin (FC), have given proof of being reliable markers of intestinal inflammation with good clinical sensitivity. Calprotectin is useful in the differential diagnosis between inflammatory bowel disease and irritable bowel syndrome, as well as in the follow-up of inflammatory bowel disease patients and in predicting treatment response, with an excellent correlation with endoscopic activity. Its role in collagenous colitis and infectious colitis is less clear and still under investigation. Key Message: Despite the growing evidence supporting its use, many clinicians are uncomfortable in dosing FC, due to its low specificity and the variability of cut-off values. Indeed there are no clear guidelines about how to manage patients with intermediate levels of FC. The aim of this article is to review recent literature on calprotectin and its use. The strong points and the limits of FC measurement will be analysed, and a practical approach in the daily clinical routine will be proposed.

P201 Monitoring histological activity in ulcerative colitis: correlation of faecal biomarkers with the Riley Score and the Nancy Index

Background: Histological healing in ulcerative colitis (UC) may be a better predictor than macroscopic appearance or clinical criteria for time to relapse. Indicators of acute mucosal inflammation which are used in the Riley Score or the Nancy index are associated with a two- to threefold increase in the risk of UC relapse during 12 months’ follow-up. However histologic assessment requires invasive endoscopy and gaining of biopsy. Non-invasive surrogates of mucosal healing would help to lower risks, costs and might increase patient acceptance. Fecal biomarkers are frequently used in UC and might be of help in this endeavour. The study aimed to investigate the performance of non-invasive faecal biomarkers compared with the Riley Score and Nancy index in patients with UC.

Methods: Colonoscopy was performed in every patient and a faecal sample was harvested within 72 h. Three biopsies were taken from the macroscopically most inflamed location or random from each colonic segment and Riley Score and Nancy index were calculated. For each patient the highest calculated score was compared with the faecal biomarkers Lactoferrin (LF), Calprotectin (CALPREST – CalP), PMN elastase (PMN-E), S100 calcium-binding protein A12 (S100A12) and Eosinophil-derived Neurotoxin (EDN). It was evaluated if the median levels of the fecal markers differ significantly between the grades 0–4 of the Nancy index.

Results: 50 patients (32 female), mean age 42.9 ± 12.3 years (range 23–67) with diagnosed UC were included. The Riley score and the Nancy index correlated with EDN (r (49) = 0.56; p = .001/ r (49) = 0.45; p = .001), S100A12 (r (49) = 0.33; p = .022/ r (49) = 0.35; p = .015), PMN-e (r (49) = 0.31; p = .028/ r (49) = 0.29; p = .044) and LF (r (49) = 0.45; p = .001/ r (49) = 0.35; p = .015), but not with CalP (p > .05). The median levels of the faecal markers of the Nancy index and the results of the Kruskal–Wallis test are presented in Table 1. LF, EDN and CalP differed significantly between the grades. Post-hoc tests showed that LF differed significantly between the grades 2 and 3 (z = −3.13, p = .011), EDN between the grades 2 and 4 (z = −3.01, p = .016) and S100A12 between the grades 2 and 3 (z = −3.97, p = .000) and 2 and 4 (z = −3.07, p = .013).

Conclusions: The fecal biomarkers LF, EDN, S100A12 and PMN-e were correlated significantly with the Riley and Nancy index, LF, EDN and CalP differed significantly between the grades of the Nancy index. The results support the utility of fecal biomarkers for detecting active histologic inflammation in patients with ulcerative colitis.

Practical guidance on the use of faecal calprotectin

Differentiation between inflammatory bowel disease (IBD) and functional gut disorders, and the determination of mucosal disease activity in established cases of IBD remain the cornerstones of disease diagnosis and management. Non-invasive, accurate biomarkers of gut inflammation are needed due to the variability of symptoms, the inaccuracies of currently available blood markers and the cost and invasive nature of endoscopy. Numerous biomarkers have been used and/or considered with some in current use. This article reviews the current evidence base around the indications for using biomarkers and their limitations, with a particular focus on faecal calprotectin.

Expert opinion for use of faecal calprotectin in diagnosis and monitoring of inflammatory bowel disease in daily clinical practice

Background: Despite many publications regarding the role of faecal calprotectin (FC) in inflammatory bowel disease (IBD), clear recommendations for its use in clinical practice are currently lacking in the literature.

Aim: The aim of this article is to provide practical guidance for clinicians for the use of FC in the detection and management of patients with IBD.

Methods: All relevant publications were analysed and practical statements were proposed based on a Delphi consensus approach.

Results: Different commercial assays have been developed but international standardisation is lacking. FC can help in the diagnosis process of IBD. In IBD, FC can predict response to therapy, detect subclinical inflammation and help to drive treatment decisions to achieve better endoscopic and clinical outcomes. After Crohn’s surgery FC can identify patients with early endoscopic recurrence.

Conclusion: Although major therapeutic changes should not be based on FC alone, FC is a valuable tool to optimise the care for IBD patients.

Faecal Calprotectin

Calprotectin is a calcium- and zinc-binding protein of the S-100 protein family which is mainly found within neutrophils and throughout the human body. The presence of calprotectin in faeces is a consequence of neutrophil migration into the gastrointestinal tissue due to an inflammatory process. Faecal calprotectin concentrations demonstrate good correlation with intestinal inflammation and faecal calprotectin is used as a biomarker in gastrointestinal disorders. Faecal calprotectin is a very sensitive marker for inflammation in the gastrointestinal tract, and useful for the differentiation of inflammatory bowel disease (IBD) from irritable bowel syndrome (IBS). Faecal calprotectin is used for the diagnosis, monitoring disease activity, treatment guidance and prediction of disease relapse and post-operative recurrence in IBD. There may also potentially be a role for faecal calprotectin in the management of infectious gastroenteritis, acute appendicitis, peptic ulcer disease, cystic fibrosis, coeliac disease, transplant rejection and graft versus host disease. Further studies are needed to confirm its utility in these conditions. Analysis of faecal calprotectin consists of an extraction step followed by quantification by immunoassay. Over the past few decades, several assays and extraction devices including point-of-care methods have been introduced by manufacturers. The manufacturer-quoted cut-off values for different faecal calprotectin assays are generally similar. However, the sensitivities and specificities at a given cut-off, and therefore the optimum cut-off values, are different between assays. A reference standard for calprotectin is lacking. Therefore, assay standardisation is required for more accurate and traceable test results for faecal calprotectin.

Serum Calprotectin Level in Children: Marker of Obesity and its Metabolic Complications

Aim: Elevated calprotectin levels have been reported in obese adults but have not been evaluated in pediatric population. We investigated the characteristics of serum calprotectin in overweight and obese children and its association with metabolic comorbidities.

Methods: We enrolled 131 children (11.7 ± 4.1 years). According to body mass index (BMI), the subjects were divided into 3 groups: obese > 95th percentile; overweight BMI 75th-95th percentile, and normal weight BMI < 75th percentile. Patients were classified as having Metabolic Syndrome if they met 3 or more of the following criteria for age and sex: BMI > 97th percentile, triglycerides > 95th percentile, high-density lipoprotein cholesterol < 5th percentile, systolic and/or diastolic blood pressure > 95th percentile, and impaired glucose tolerance. In all patients, calprotectin serum levels were also detected.

Results: Calprotectin was higher in obese and overweight children than normal weight subjects (p < 0.001), with calprotectin in females being significantly higher than in males (p = 0.04). Increased calprotectin was related to pathological fasting blood glucose (p < 0.001) and insulin resistance (p = 0.03), while BMI (p = 0.001), and diastolic pressure (p = 0.001) are independent factors for increased calprotectin.

Conclusions: Our findings confirm the association between increased calprotectin and obesity also in children and suggest the potential utility of this biomarker in the monitoring of its metabolic complications.

Characterisation of gut, lung, and upper airways microbiota in patients with non-small cell lung carcinoma: Study protocol for case-control observational trial

Background: Several studies have confirmed the important role of the gut microbiota in the regulation of immune functions and its correlation with different diseases, including cancer. While brain-gut and liver-gut axes have already been demonstrated, the existence of a lung-gut axis has been suggested more recently, with the idea that changes in the gut microbiota could affect the lung microbiota, and vice versa. Likewise, the close connection between gut microbiota and cancer of proximal sites (intestines, kidneys, liver, etc.) is already well established. However, little is known whether there is a similar relation when looking at world’s number one cause of death from cancer-lung cancer.

Objective: Firstly, this study aims to characterise the gut, lung, and upper airways (UAs) microbiota in patients with non-small cell lung cancer (NSCLC) treated with surgery or neoadjuvant chemotherapy plus surgery. Secondly, it aims to evaluate a chemotherapy effect on site-specific microbiota and its influence on immune profile. To our knowledge, this is the 1st study that will analyse multi-site microbiota in NSCLC patients along with site-specific immune response.

Methods: The study is a case-controlled observational trial. Forty NSCLC patients will be divided into 2 groups depending on their anamnesis: Pchir, patients eligible for surgery, or Pct-chir, patients eligible for neoadjuvant chemotherapy plus surgery. Composition of the UAs (saliva), gut (faeces), and lung microbiota (from broncho-alveolar lavage fluid (BALF) and 3 lung pieces: “healthy” tissue distal to tumour, peritumoural tissue and tumour itself) will be analysed in both groups. Immune properties will be evaluated on the local (evaluation of the tumour immune cell infiltrate, tumour classification and properties, immune cell phenotyping in BALF; human neutrophil protein (HNP) 1-3, β-defensin 2, and calprotectin in faeces) and systemic level (blood cytokine and immune cell profile). Short-chain fatty acids (SCFAs) (major products of bacterial fermentation with an effect on immune system) will be dosed in faecal samples. Other factors such as nutrition and smoking status will be recorded for each patient. We hypothesise that smoking status and tumour type/grade will be major factors influencing both microbiota and immune/inflammatory profile of all sampling sites. Furthermore, due to non-selectivity, the same effect is expected from chemotherapy.

Faecal calprotectin delivers on convenience, cost reduction and clinical decision-making in inflammatory bowel disease: a real-world cohort study

Background: Faecal calprotectin (FC) is an accurate biomarker of disease activity in inflammatory bowel disease (IBD), yet the cost/resource implications of incorporating FC into ‘real-world’ practice remain uncertain.

Aim: To evaluate the utility of FC in clinical decision-making and on healthcare costs in IBD.

Methods: Retrospective data, including colonoscopy/other investigations, medication, admission and surgical data, were collected from hospital records and compared between two groups: pre-FC historical cohort (2005-2009) where colonoscopy was used to assess IBD activity versus the cohort where FC was used first instead (2010-2014). Post-test costs were also compared.

Results: A total of 357 FC tests (246 patients, 2010-2014) and 450 colonoscopies (268 patients, 2005-2009) were performed. On subsequent review, both FC and colonoscopy (in their respective cohorts) were associated with changes in management in 50.7 versus 56.2% (P = 0.14), respectively, with similar proportions of subsequent IBD-related investigations within 6 months (21.8 vs 21.9%, P = 1.0). Prior to FC availability (2005-2009), a colonoscopy for disease reassessment cost AU$606 578 (cost per patient-year $1887.34) versus AU$282 048 (cost per patient-year $968.60) when FC ± colonoscopy was used (2010-2014). Within the FC cohort, 73.6% did not proceed to colonoscopy within 6 months post-FC, and 60.6% had not undergone colonoscopy post-FC by the end of follow up (median 1.8 years (0.1, 4.6) post-FC). Those with FC ≥ 250 were scoped earlier than those with FC < 100 μg/mL (median 0.49 vs 1.0 years, P = 0.03).

Conclusion: Introduction of FC into routine IBD care aided changes in clinical management in a similar proportion, yet at potentially half the total cost, compared to a historical colonoscopy-only cohort at the same centre.

Faecal calprotectin to detect inflammatory bowel disease: a systematic review and exploratory meta-analysis of test accuracy

Objective: Test accuracy of faecal calprotectin (FC) testing in primary care is inconclusive. We aimed to assess the test accuracy of FC testing in primary care and compare it to secondary care estimates for the detection of inflammatory bowel disease (IBD).

Methods: Systematic review and meta-analysis of test accuracy using a bivariate random effects model. We searched MEDLINE, EMBASE, Cochrane Library and Web of Science until 31 May 2017 and included studies from auto alerts up until 31 January 2018. Eligible studies measured FC levels in stool samples to detect IBD in adult patients with chronic (at least 6-8 weeks) abdominal symptoms in primary or secondary care. Risk of bias and applicability were assessed using the Quality Assessment of Diagnostic Accuracy Studies-2 criteria. We followed the protocol registered as PROSPERO CRD 42012003287.

Results: 38 out of 2168 studies were eligible including five from primary care. Comparison of test accuracy by setting was precluded by extensive heterogeneity. Overall, summary estimates of sensitivity and specificity were not recorded. At a threshold of 50 µg/g, sensitivity from separate meta-analysis of four assay types ranged from 0.85 (95% CI 0.75 to 0.92) to 0.94 (95% CI 0.75 to 0.90) and specificity from 0.67 (95% CI 0.56 to 0.76) to 0.88 (95% CI 0.77 to 0.94). Across three different definitions of disease, sensitivity ranged from 0.80 (95% CI 0.76 to 0.84) to 0.97 (95% CI 0.91 to 0.99) and specificity from 0.67 (95% CI 0.58 to 0.75) to 0.76 (95% CI 0.66 to 0.84). Sensitivity appears to be lower in primary care and is further reduced at a revised threshold of 100 µg/g.

Conclusions: Conclusive estimates of sensitivity and specificity of FC testing in primary care for the detection of IBD are still missing. There is insufficient evidence in the published literature to support the decision to introduce FC testing in primary care. Studies evaluating FC testing in an appropriate primary care setting are needed.

Faecal calprotectin in inflammatory bowel diseases: a review focused on meta-analyses and routine usage limitations

A growing body of evidence has been published about the usefulness of measuring calprotectin in faecal samples (FCAL) in inflammatory bowel disease (IBD) assessment, including diagnosis, monitoring of disease activity and relapse prediction. Several systematic reviews with meta-analyses compiling studies for each particular clinical setting have been carried out in recent years. Most of these were focused on the use of FCAL in IBD diagnosis and showed a relevant role for this marker in selecting patients with gastrointestinal symptoms who would not need a further examination by endoscopy. Although a lesser number of meta-analyses have been performed on the use of FCAL as a surrogate marker of disease activity, a close correlation between FCAL and endoscopic activity of IBD has been shown. With respect to the predictive capacity of FCAL for IBD relapse, a single meta-analysis published indicates that this role is more limited. Furthermore, FCAL thresholds vary considerably depending on the clinical setting and, what is more concerning, among different commercially available assays due to a lack of FCAL concentration interchangeability. Here, we summarise recent publications about the role and limitations of FCAL in IBD, with a special focus on meta-analyses, and give an overview of alternative faecal biomarkers.

Phylogenetic and pathotype analysis of Escherichia coli stool isolates from Egyptian patients with inflammatory bowel disease

Introduction: The role of Escherichia coli in the pathogenesis of inflammatory bowel disease (IBD) is still controversial. The study aimed to investigate the pathotypes and the phylogenetic groups of E. coli in Egyptian patients with IBD in an attempt to find an association between any type or group with the severity of the disease.

Methods: Thirty ulcerative colitis (UC), 30 Crohn’s disease (CD), and 20 control subjects with normal colonoscopy were included in a cross-sectional study. E. coli were isolated from stool samples by culture. Eight intestinal virulence genes coding for diarrheagenic E. coli were investigated using multiplex PCR. Phylogenetic grouping was performed by a triplex PCR. Antimicrobial susceptibility of all isolates was done using disc diffusion method.

Results: Enteroaggregative E. coli (EAEC) were identified in 25% (15/60) of IBD cases and in none of the controls (p=0.013). Out the 60 IBD cases, 30 (50%) were from phylogenetic group B2. No statistically significant differences in the distribution of E. coli phylogenetic groups were found between study groups. However, 80% of EAEC were assigned to group B2 and D. No statistically significant differences in calprotectin level or in disease severity scores were reported between the four phylogenetic groups. E. coli from both UC and CD patients showed a high rate of resistance to most antimicrobials when compared to the control group.

Conclusions: The identification of EAEC belonging mainly to group B2 and D in IBD cases may indicate the importance of this pathotype in the pathogenesis of IBD in Egyptian patients.

Synovial Fluid Calprotectin for the Preoperative Diagnosis of Chronic Periprosthetic Joint Infection

Background: The diagnosis of periprosthetic joint infection (PJI) represents a challenge in clinical practice and the analysis of synovial fluid is a useful diagnostic tool. Calprotectin is an inflammatory biomarker widely used in the evaluation of chronic inflammatory diseases; however, little is known about its role in PJI. The purpose of this study is to determine the reliability of synovial calprotectin in the diagnosis of PJI.

Methods: Seventy-six patients with painful knee arthroplasty were included in this prospective observational study. Synovial fluid was analyzed for cell count, percentage of polymorphonuclear neutrophils, microbiological culture, leukocyte esterase strip test, alpha-defensin rapid test, and calprotectin immunoassay dosage. The 2018 Consensus Statements criteria for PJI were used as standard reference to define the presence of infection. Sensitivity, specificity, positive and negative likelihood ratio, and receiver-operation characteristic curve were calculated for calprotectin immunoassay test.

Results: By 2018 Consensus Statements criteria for PJI, 28 patients were considered infected, 44 patients were considered not infected, and 4 patients were classified as inconclusive. The calprotectin synovial fluid test resulted in 2 false-positive results and no false-negative results. The calprotectin synovial fluid test demonstrated a sensitivity of 100% (95% confidence interval [CI] 99.96-100) and specificity of 95% (95% CI 89.4-100) for the diagnosis of PJI. The positive likelihood ratio was 22 (95% CI 5.680-85.209) and the negative likelihood ratio was 0 (95% CI 0-0.292). The area under the receiver-operation characteristic curve was 0.996 (95% CI 94.3-100).

Conclusion: The present study suggests that synovial calprotectin immunoassay test has a high sensitivity and specificity in the diagnosis of knee PJI. Moreover, it is easily applied, quick and valuable in clinical practice.

Faecal calprotectin and gut microbiota do not predict enteropathy in very preterm infants

Aim: Very preterm birth is associated with a high risk of enteropathies. Diagnosis is challenging, especially in mild forms, leading to unnecessary periods of cessation of enteral feeding. This study aimed at establishing a prognosis score of enteropathy combining clinical parameters and faecal calprotectin concentration.

Methods: This prospective multicentric study included preterm neonates born at a gestational age of 33 weeks or less. Stools were collected weekly until hospital discharge, and daily in case of digestive events for calprotectin measurement (ELISA and immunochromatography) and microbiota analyses (16S rRNA gene sequencing).

Results: Among the 121 neonates included, 21 experienced at least one episode of enteropathy, mainly mild forms. By ELISA testing, median faecal calprotectin was 88 (8-798) µg/g faeces. No statistically significant association was found between the outset of enteropathy and maternal and neonatal characteristics, and calprotectin levels. The agreement between ELISA and immunochromatography assay was moderate (intra-class correlation coefficient 0.58, 95%CI [0.47-0.66]). Comparison of species diversity and relative bacterial abundance profiles between infants with or without enteropathy revealed no specific alterations associated with enteropathy.

Conclusion: The study failed to propose a prognostic score of enteropathy, probably due the large inter- and intra-individual variability of faecal calprotectin in very preterm neonates.

Synovial Biomarkers to Detect Chronic Periprosthetic Joint Infection: A Pilot Study to Compare Calprotectin Rapid Test, Calprotectin ELISA Immunoassay and Leukocyte Esterase Test

Background: Periprosthetic joint infection (PJI) is a devastating complication after joint replacement surgery, and making diagnosis is often far from obvious. Calprotectin was recently proposed as a promising synovial biomarker to detect PJI. To our knowledge, no comparative study exists between enzyme-linked immunosorbent assay (ELISA) and rapid calprotectin test (CalFAST). Our purpose was to compare these methods with leukocyte esterase (LE) test from synovial fluid of painful knee arthroplasty subjected to infectious workup.

Methods: Ninety-three patients were included in this prospective observational study. They underwent synovial fluid aspiration that was analyzed for cell count, microbiological culture, LE test, calprotectin rapid test, and calprotectin immunoassay dosage. The 2018 Consensus Statements criteria for PJI were used to diagnose PJI. Sensitivity, specificity, positive and negative likelihood ratio, and receiver operating characteristic were calculated for detection methods and compared.

Results: We categorized 39 patients as infected and 50 patients as not infected. The sensitivity comparing the ELISA test and CalFAST test was similar, 92.3% and 97.4%, respectively. LE rapid test showed 46% of sensitivity and 94% of specificity. The highest specificity was found with ELISA test (100%). Comparing the receiver operating characteristic curves by z-test, there were statistically significant differences between LE strip test and the other two methods. Otherwise, no statistically significant differences were present between ELISA and CalFAST test.

Conclusion: Synovial calprotectin detection has high accuracy in knee PJI diagnosis, both ELISA and rapid test. LE strip test remains a good test to confirm the diagnosis of PJI in case of positivity. In clinical practice, the calprotectin rapid test can be considered an excellent point-of-care test.

Calprotectin as a novel diagnostic approach to screen male infertility risk: A pilot study

Research question: Diagnosis of male infertility is essentially based on the evaluation of semen quality (sperm concentration, motility, viability, and morphology). However, there is a lack of knowledge about possible molecules used as candidates for the early identification of male infertility risk. Calprotectin is a biological marker for inflammation, measured prevalently in stool specimens, widely used to discriminate between inflammatory bowel disease (IBD) and irritable bowel syndrome (IBS), and for the subsequent monitoring of gastrointestinal diseases’ development. Would it be possible to use calprotectin determination to identify also male infertility risk?

Design: Cross-sectional pilot study investigated calprotectin concentration in the seminal fluid of 45 men (range: 23-51 yrs) that were under evaluation for semen quality at our Center for Reproductive Medicine. Calprotectin concentration was determined with a commercially available immuno-chromatographic test and successfully detected in 37 of the 45 analyzed men (age: 37.38 ± 6.59). A correlation with semen quality (concentration, motility, morphology) was assessed.

Results: Higher calprotectin concentration seemed to indicate a better quality of the seminal fluid. Normozoospermic subjects (Group A) had on average a calprotectin value of 0.215 ± 0.162 µg/ml (mean ± SD), while subjects with at least one of the semen parameters below reference values (Group B) showed lower calprotectin concentration (mean ± SD: 0.126 ± 0.068, p-value < 0.05). A significant difference was clearly evident between calprotectin concentration measured in seminal fluids with physiological sperm morphology (≥4%) as compared with teratozoospermic samples (<4%) (p-value < 0.05). Indeed, the developed ROC curves showed a good diagnostic accuracy (around 67 %) using calprotectin concentration (threshold value: 0.121 μg/ml) as a preliminary test to discriminate subjects with and without abnormal semen parameters, especially morphology.

Conclusions: Calprotectin determination in the seminal fluid may be proposed as a biological marker for preliminary screening in male subjects at risk of infertility due to one or more alterations of semen quality.

The Role of Serum Calprotectin in Defining Disease Outcomes in Non-Systemic Juvenile Idiopathic Arthritis: A Pilot Study

Serum calprotectin (MRP8/14) is currently being studied as a promising biomarker of disease activity and outcome in patients with juvenile idiopathic arthritis (JIA) but the data in the literature are conflicting. The aim of our study was to investigate the potential role of serum calprotectin as biomarker of disease activity and flare/remission in a group of nsJIA patients during a follow-up period of 18 months. In this prospective longitudinal study, two groups of patients with ns-JIA (55 active patients and 56 patients in remission according to Wallace’s criteria) and a control group (50 children) were recruited at baseline from January 2020 to September 2021. JIA patients were followed for up to 18 months at four timepoints: 3 months (T1), 6 months (T2), 12 months (T3) and 18 months (T4). At each timepoint, the following were recorded: JADAS27, blood counts, ESR, CRP, albumin, ferritin and serum calprotectin. To illustrate the performance of calprotectin, Kaplan-Meier curves were constructed from baseline to relapse/remission, dichotomizing patients at baseline in positive/negative on the basis progressive calprotectin cut-offs. Associations between baseline factors and relapse were determined using Cox regression models. Multivariate models were constructed to analyze the effect of covariates. Comparing baseline clinical and laboratory data of the three groups (active vs. inactive JIA vs. controls), only serum calprotectin reached statistical significance (active patients vs. inactive (p = 0.0016) and vs. controls (p = 0.0012)). In the inactive group, during the 18 months of follow up, 31 patients (55.3%) had a relapse. Comparing the baseline data of relapsers vs. non-relapsers, serum calprotectin showed higher levels (p = 0.001) in relapsers. In survival analysis, a log rank test showed significant differences of up to 12 ng/mL (p = 0.045). Multivariate Cox regression confirmed that only baseline calprotectin levels were independently associated with disease recurrence. In the active group, in the 12 months of follow-up, 19 patients (38%) entered remission of the disease. In addition, in this group, the only statistical difference at the baseline was the value of MPR8/14 (p = 0.0001). Log rank test showed significant differences up to 10 ng/mL (p = 0.003). In the multivariate Cox regression, serum calprotectin levels at baseline were independently associated with remission. In conclusion, our study would suggest a dual role for calprotectin in predicting future relapse and treatment response in patients with nsJIA, thus influencing therapeutic decisions and management of these patients during follow up.