The role of Killer immunoglobulin-like receptors (KIRs) in the genetic susceptibility to non-celiac wheat sensitivity (NCWS).

OBJECTIVES: Non-celiac wheat sensitivity (NCWS) is an emerging clinical condition characterized by gastrointestinal and extraintestinal symptoms following the ingestion of gluten-containing foods in patients without celiac disease (CD) or wheat allergy. Despite the great interest for NCWS, the genetic risk factors still need to be fully clarified. In this study, we first assessed the possible contribution of KIR genes and KIR haplotypes on the genetic predisposition to NCWS. METHODS: Fifty patients with NCWS, 50 patients with CD, and 50 healthy controls (HC) were included in this study. KIR genes and KIR genotyping were investigated in all subjects by polymerase chain reaction with the sequence oligonucleotide probe (PCR-SSOP) method using Luminex technology. RESULTS: We found a statistically different distribution of some KIR genes among NCWS, CD, and HC. Specifically, NCWS showed a decreased frequency of KIR2DL1, -2DL3, -2DL5, -2DS2, -2DS3, -2DS4, -2DS5, and -3DS1 genes, and an increased frequency of -3DL1 gene respect to both CD and HC. No difference was detected in the KIR haplotype expression. At the multivariate analysis, KIR2DL5, -2DS4, and -2DS5 were independent predictors of NCWS. CONCLUSIONS: Our findings suggest a role of KIR genes in NCWS susceptibility, with KIR2DL5, -2DS4, and -2DS5 having a protective effect. Further large-scale multicentric studies are required to validate these preliminary findings.

Prostate Health Index (PHI) as a triage tool for reducing unnecessary magnetic resonance imaging (MRI) in patients at risk of prostate cancer.

INTRODUCTION: The aim of this study is to assess the usefulness of the Prostate Health Index (PHI) as a triage tool for selecting patients at risk of prostate cancer (PCa) who should undergo multiparametric Magnetic Resonance Imaging (mpMRI). MATERIAL AND METHODS: We enrolled 204 patients with suspected PCa. For each patient, a blood sample was collected before mpMRI to measure PHI. Findings on mpMRI were assessed according to the Prostate Imaging Reporting & Data System version 2.0 (PI-RADSv2) category scale. RESULTS: According to PI-RADSv2, patients were classified into two groups: PI-RADS < 3 (48 %) and ≥ 3 (52 %). PHI showed the best performance for predicting PI-RADS ≥ 3 [AUC: 0,747 (0,679-0,815), 0,680(0,607-0,754), and 0,613 (0,535-0,690) for PHI, PSA ratio, and total PSA, respectively]. The best PHI cut-off was 30, with a sensitivity of 90%. At the univariate logistic regression, total PSA (p = 0.007), PSA ratio (p = 0.001), [-2]proPSA (p = 0.019) and PHI (p < 0.001) were associated with PI-RADS ≥ 3; however, at the multivariate analysis, only PHI (p < 0.001) was found to be an independent predictor of PI-RADS ≥ 3. CONCLUSION: PHI could represent a reliable noninvasive tool for selecting patients to undergo mpMRI.

A machine learning strategy to mitigate the inappropriateness of procalcitonin request in clinical practice.

Aim: The aim of this study was to develop machine learning (ML) models to mitigate the inappropriate request of Procalcitonin (PCT) in clinical wards. Material and methods: We built six different ML models based on both demographical data, i.e., sex and age, and laboratory parameters, i.e., cell blood count (CBC) parameters, inclusive of monocyte distribution width (MDW), and C-reactive protein (CRP). The dataset included 1667 PCT measurements of different patients. Based on a PCT cut-off of 0.50 ng/mL, we found 1090 negative (65.4%) and 577 positive (34.6%) results. We performed a 70:15:15 train:validation:test splitting based on the outcome. Results: Random Forest, Support Vector Machine and eXtreme Gradient Boosting showed optimal performances for predicting PCT positivity, with an area under the curve ranging from 0.88 to 0.89. Conclusions: The ML models developed could represent a useful tool to predict PCT positivity, avoiding unusefulness PCT requests. ML models are based on laboratory tests commonly ordered together with PCT but have the great advantage to be easy to measure and low-cost.

The Role of New Morphological Parameters Provided by the BC 6800 Plus Analyzer in the Early Diagnosis of Sepsis.

BACKGROUND: Late diagnosis of sepsis is associated with adverse consequences and high mortality rate. The aim of this study was to evaluate the diagnostic value of hematologic research parameters, that reflect the cell morphology of blood cells, available on the BC 6800 plus automated analyzer (Mindray) for the early detection of sepsis. MATERIALS AND METHODS: A complete blood count (CBC) was performed by Mindray BC 6800 Plus Analyzer in 327 patients (223 with a confirmed diagnosis of sepsis following sepsis-3 criteria, 104 without sepsis), admitted at the Intensive Care Unit of the Novara's Hospital (Italy) and in 56 patients with localized infection. RESULTS: In univariate logistic regression, age, Hb, RDW, MO#, NMR, NeuX, NeuY, NeuZ, LymX, MonX, MonY, MonZ were associated with sepsis (p < 0.005). In multivariate analysis, only RDW, NeuX, NeuY, NeuZ, MonX and MonZ were found to be independent predictors of sepsis (p < 0.005). Morphological research parameters are confirmed to be predictors of sepsis even when analyzing the group with localized infection. CONCLUSIONS: In addition to already established biomarkers and basic CBC parameters, new morphological cell parameters can be a valuable aid in the early diagnosis of sepsis at no additional cost.

Estimation of the reference values and decision limits for growth hormone in newborns using dried blood spots.

OBJECTIVES: Severe deficiency of growth hormone (GHD) of the newborn is a rare but potentially life-threatening disease. GH measured during the first week of life, using dried blood spots (DBS), may offer several advantages. Aim of the study was to estimate the reference values for GH in newborns by a new analytical method using DBS. METHODS: Using a new developed analytical method, GH was estimated from DBS of 1,036 healthy newborns attending the Neonatology Unit of Fondazione IRCCS Ca' Granda Ospedale Maggiore Policlinico of Milan in the period July-October 2021. Reference values for GH deficiency were estimated by the Harrell-Davis bootstrap method, with 90 %CI calculated by the bias-corrected and accelerated bootstrap method. RESULTS: All GH measurements required 33 analytical sessions (8 months) with a CV% for calibration curve slopes equal to 6.9 %. Intermediate precision evaluated by measurement of low (3 µg/L) and high (10 µg/L) quality controls was, respectively, 14 and 6.5 %. GH reference values, estimated at percentiles 1.0st, 2.5th and 5.0th, and their 90 %CI, were, respectively, 4.5 µg/L (90 %CI 3.8-5.1), 5.9 µg/L (90 %CI 5.4-6.4) and 7.0 µg/L (90 %CI 6.7-7.3). GH levels were not associated with sex, standard deviation scores, birth weight, gestational age, type of delivery or mother's variables (age, smoking habit, gestational diabetes). CONCLUSIONS: Validation data suggest that this method can be used to measured GH in newborns using DBS. The reference values estimated in this study are in accordance with previous published works using ELISA and may help confirming the clinical suspicion of neonatal GHD.

Drainage fluid LDH and neutrophil to lymphocyte ratio as biomarkers for early detecting anastomotic leakage in patients undergoing colorectal surgery.

OBJECTIVES: In this study, we investigated the role of several circulating and drainage fluid biomarkers for detecting postoperative complications (PCs) and anastomotic leakage (AL) in patients undergoing colorectal surgery. METHODS: All consecutive patients undergoing colorectal surgery between June 2018 and April 2020 were prospectively considered. On postoperative days (POD) 1, 3, and 5, we measured lactate dehydrogenase (LDH) in drainage fluid, C-reactive protein (CRP) in serum and drainage fluid, and neutrophil to lymphocyte ratio (NLR). RESULTS: We enrolled 187 patients. POD1 patients with AL had higher serum CRP levels, while on POD3 and on POD5 higher NLR and serum CRP. LDH and CRP in drainage fluid were also significantly higher at both time points. The area under the curves (AUCs) of serum and drainage fluid CRP were 0.752 (0.629-0.875) and 0.752 (0.565-0.939), respectively. The best cut-off for serum and drainage fluid CRP was 185.23 and 76 mg/dL, respectively. The AUC of NLR on POD3 was 0.762 (0.662-0.882) with a sensitivity and specificity of 84 and 63 %, respectively, at a cut-off of 6,6. Finally, drainage fluid LDH showed the best diagnostic performance for AL, with an AUC, sensitivity, and specificity of 0.921 (0.849-0.993), 82 %, and 90 % at a cut-off of 2,186 U/L. Trends in serum parameters between patients with or without PCs or AL were also evaluated. Interestingly, we found that NLR decreased faster in patients without PCs than in patients with PCs and patients with AL. CONCLUSIONS: Drainage fluid LDH and NLR could be promising biomarkers of PCs and AL.

Predictors of controlled attenuation parameter in metabolic dysfunction.

BACKGROUND & AIMS: Hepatic fat content can be non-invasively estimated by controlled attenuation parameter (CAP) during transient elastography. The aim of this study was to examine the determinants and predictors of CAP values in individuals with metabolic dysfunction. METHODS: We enrolled 1230 consecutive apparently healthy individuals (Liver-Bible-2022 cohort) with ≥3 metabolic dysfunction features. CAP was measured by Fibroscan. CAP determinants and predictors were identified using backward stepwise analysis and introduced in generalized linear models. RESULTS: Participants were predominantly males (82.9%), mean age was 53.8 ± 6.4 years, 600 (48.8%) had steatosis (CAP ≥ 275 dB/m), and 27 had liver stiffness measurement (LSM) ≥ 8 kPa. CAP values correlated with LSM (p < 10-22). In multivariable analysis, fasting insulin and abdominal circumference (AC) were the main determinants of CAP (p < 10-6), together with body mass index (BMI; p < 10-4), age, diabetes, triglycerides, ferritin, and lower HDL and thyroid stimulating hormone (TSH; p < 0.05 for all). In a subset of 592 participants with thyroid hormone measurement, we found an association between higher free triiodothyronine levels, correlating with lower TSH, and CAP values, independent of TSH and of levothyroxine treatment (p = 0.0025). A clinical CAP score based on age, BMI, AC, HbA1c, ALT, and HDL predicted CAP ≥ 275 dB/m with moderate accuracy (AUROC = 0.73), which was better than that of the Fatty Liver Index and of ALT (AUROC = 0.70/0.61, respectively) and validated it in multiple cohorts. CONCLUSION: Abdominal adiposity and insulin resistance severity were the main determinants of CAP in individuals with metabolic dysfunction and may improve steatotic liver disease risk stratification. CAP values were modulated by the hypophysis-thyroid axis.

Machine learning algorithms in sepsis.

Sepsis remains a significant global health challenge due to its high mortality and morbidity, compounded by the difficulty of early detection given its variable clinical manifestations. The integration of machine learning (ML) into laboratory medicine for timely sepsis identification and outcome forecasting is an emerging field of interest. This comprehensive review assesses the current body of research on ML applications for sepsis within the realm of laboratory diagnostics, detailing both their strengths and shortcomings. An extensive literature search was performed by two independent investigators across PubMed and Scopus databases, employing the keywords "Sepsis," "Machine Learning," and "Laboratory" without publication date limitations, culminating in January 2023. Each selected study was meticulously evaluated for various aspects, including its design, intent (diagnostic or prognostic), clinical environment, demographics, sepsis criteria, data gathering period, and the scope and nature of features, in addition to the ML methodologies and their validation procedures. Out of 135 articles reviewed, 39 fulfilled the criteria for inclusion. Among these, the majority (30 studies) were focused on devising ML algorithms for diagnosis, fewer (8 studies) on prognosis, and one study addressed both aspects. The dissemination of these studies across an array of journals reflects the interdisciplinary engagement in the development of ML algorithms for sepsis. This analysis highlights the promising role of ML in the early diagnosis of sepsis while drawing attention to the need for uniformity in validating models and defining features, crucial steps for ensuring the reliability and practicality of ML in clinical setting.

Customized cutoff limits for the sediMAX-2 automated analyzer reduce the number of urine culture tests.

BACKGROUND: Urinary tract infections are highly prevalent in nosocomial and community settings. Their diagnosis, although costly and time-consuming, is crucial to avoid inappropriate treatments and/or clinical complications. In this context, automated analyzers have been developed and commercialized to screen and rule out negative urine samples. Adjustments of the manufacturers' suggested cutoff values might lead to substantial diagnostic and economic advantages. METHODS: We retrospectively analyzed 776 urine samples from different individuals. 546 samples (training group) were used to optimize develop new cutoffs values. The remaining 230 samples (validation group) were used to validate the optimized cutoffs. All samples were subjected to urine culture, 17% resulted positive. Escherichia coli and Enterococcus faecalis were the two most frequently identified bacteria, 95 and 9 samples, respectively. RESULTS: Two different cutoffs levels were obtained. Cutoff-A (bacteria>110 and/or white blood cells> 15 cell/µL), showed the same sensitivity of the manufacturers' suggested cutoff, yet leads to a large reduction of the samples to be cultured. Cutoff-B (bacteria>50 and/or white blood cells>20 cell/µL), showed an almost 100% sensitivity by subjecting only ~70% of the samples to urine culture. CONCLUSION: Cutoff-A is a good compromise between sensitivity and specificity yet allowing economic advantages by reducing the number of urinary cultures. Cutoff-B relegates urinary tract infection misdiagnosis to a rare event without the need of culturing the entire batch of samples. We believe that clinical implementation of the proposed cutoffs will help other laboratories, using similar instrumentation, to reach their most convenient balance between sensitivity and economical needs.

Analysis of annual distributions of hemoglobin A2 values as a method to test for HbA2 standardization.

BACKGROUND AND AIMS: The monitoring of yearly distributions of HbA2 measured has been indicated as a reliable indicator of worldwide standardization. MATERIALS AND METHODS: Measurements/year of HbA2 have been collected over three consecutive years in 15 Italian laboratories each using the same analytical method over three years period. HbA2 distributions, cleaned of replicated measurements, were compared by the overlapping area of the raw probability density functions expressed by coefficient eta (η), and by comparing the reference intervals for the central part of each distribution estimated by the indirect method refineR using the R package "refineR". RESULTS: According to the overlapping areas analysis the distributions/year of the data provided by 4 centers able to perform at least 1000 measurements/year were similar in 2 consecutive years. Moreover, the reference intervals provided by 2 centers using the same analytical methods in two separate locations over the three consecutive years, were very similar. The highest overlap (99.7 %) was observed in one center over two consecutive years. The overlapping areas were very high (93.6-95.7%) in 8 out of 9 inter-comparisons. CONCLUSION: Despite the limitations of this study the yearly distribution of the HbA2 measured in various centers appears a reliable tool to test HbA2 standardization over different centers using different analytical methods.

Monocyte distribution width (MDW) in sepsis.

Sepsis is a life-threatening syndrome due to a dysregulated host response to infection, which can be caused by bacterial, viral, or fungal infection. Thus, it is crucial to know how the different microorganisms influence the levels of a biomarker. In the last decade, monocyte distribution width (MDW) has emerged as a promising sepsis biomarker, especially in acute settings, such as the Emergency Department and Intensive Care Unit. In this article, we explore the relationship between MDW and the different pathogens causing infection. Noteworthy, MDW is not a biological molecule, but it is calculated by a mathematical formula based on monocyte characteristics. Monocytes represent the first line defence against microorganisms and undergo activation upon infection, independently from the invading pathogen. According to the knowledge on the biomarker biology and the few literatures evidence, MDW may be considered a biomarker of sepsis, independent of the causative pathogen. However, further investigations are warranted before drawing definite conclusion.

Detection of Antibodies against the Acetylcholine Receptor in Patients with Myasthenia Gravis: A Comparison of Two Enzyme Immunoassays and a Fixed Cell-Based Assay.

The detection of serum anti-acetylcholine receptor (AChR) antibodies is currently an important tool for diagnosing myasthenia gravis (MG) since they are present in about 85% of MG patients. Many serological tests are now available. Nevertheless, results from these tests can be different in some patients. The aim of this study is to compare the sensitivity of a commercially available fixed cell-based assay (F-CBA) to that of enzyme-linked immunosorbent assay (ELISA) kits for anti-AChR detection in patients with a diagnosis of MG. Overall, 143 patients with a confirmed MG diagnosis were included in the study. The detection and measurement of serum anti-AChR antibodies were performed by three analytical methods, namely, a competitive ELISA (cELISA), an indirect ELISA (iELISA), and an F-CBA, according to the manufacturers' instructions. Anti-AChR antibody titers were positive in 94/143 (66%) using the cELISA, in 75/143 (52%) using the iELISA and in 61/143 (43%) using the F-CBA (adult and/or fetal). Method agreement, evaluated by concordant pairs and Cohen's kappa, was as follows: cELISA-iELISA: 110/143 (77%), k = 0.53 (95%CI 0.40-0.66); cELISA-F-CBA: 108/143 (76%), k = 0.53 (95%CI 0.41-0.66); iELISA-F-CBA: 121/143 (85%), k = 0.70 (95%CI 0.57-0.80). Our findings show that the cELISA has better analytical performance than the iELISA and F-CBA. However, the iELISA and F-CBA show the highest concordance.

The Role of TAR DNA Binding Protein 43 (TDP-43) as a CandiDate Biomarker of Amyotrophic Lateral Sclerosis: A Systematic Review and Meta-Analysis.

BACKGROUND: TAR DNA-binding protein 43 (TDP-43) aggregation in neuronal cells is recognized as a hallmark of amyotrophic lateral sclerosis (ALS). Although the literature strongly supports the pathogenetic role of TDP-43 in ALS pathogenesis, the role of TDP-43 as a biomarker of ALS is controversial. We performed a systematic review and meta-analysis to assess the diagnostic performance of TDP-43 for ALS. METHODS: Relevant publications were identified by a systematic literature search on PubMed and Web of Science from their inception to 8 April 2022. RESULTS: Seven studies, including 472 individuals, of whom 254 had ALS according to the Revised Amyotrophic Lateral Sclerosis Functional Rating Scale, met the inclusion criteria for our meta-analysis. According to the random-effects model, CSF TDP-43 levels are higher in ALS patients compared with control groups. CONCLUSIONS: CSF TDP-43 could represent a biomarker of ALS, but further studies are mandatory before drawing conclusions.

Reference intervals for glycated albumin during physiological pregnancy of Europid women: Evidences from a prospective observational study.

BACKGROUND AND AIMS: Glycated albumin (GA) may assess glycometabolic control over a short period of time respect to HbA1c, and its use to screen for gestational diabetes in pregnancy has been suggested. To this regard few data on reference intervals (RI) for GA on Europid women have been collected, only from cross-sectional investigations. Aim of this work has been to collect trimester-specific RI for GA in physiological pregnancies, following a longitudinal prospective study. METHODS: Forty-five healthy pregnant Europid women have been enrolled for whom a GDM screening test was scheduled at 24-28 weeks, in 5 different Italian centers. Only those negative to the OGTT were included. The women had 4 successive visits at 6-10 weeks of gestation, at 16-18 weeks, at 24-28 weeks and at the end of pregnancy. ALT, AST, total bilirubin, C-reactive protein, cholinesterase, creatinine, GGT, glycated albumin, iron, total serum proteins, transferrin were measured in duplicate on aliquots of serum samples by a central laboratory. RESULTS: The RI (2.5-97.5 percentiles) for GA were 11.1-14.8 % (I visit), 10.9-15.6 % (II visit), 10.6-14.1 % (III visit) and 10.7-14.3 % (IV visit). The RI of other biomarkers confirmed previously published data. The RI for serum cholinesterase we present are novel, and were 5049-9906 U/L (Iv), 4212-8965 U/L (IIv), 3518-8470 U/L (IIIv) and 3945-8727 U/L (IVv). CONCLUSIONS: Trimester-specific RI are important for using GA and serum cholinesterase in pregnancy. However, considering the high inter-individual variability of both markers, the use of longitudinal interpretations of the individual variations of both proteins during pregnancy should be preferred.

Use of Computer Simulation for Calculating Reference Limits for Distribution Overlap.

BACKGROUND: In laboratory medicine, data collected in different settings or under different conditions are frequently analyzed to obtain meaningful information. Evaluation based only on location or variability measures, although useful, is not enough to extract all the value from data. Other mathematical or statistical approaches are possible, but the specific knowledge required is frequently out of the reach of most laboratorians. Computer simulation may help in solving the problem. The aim of this work was to use computer simulation for calculating reference limits for the overlap of 2 distributions. METHODS: Computer simulation was applied to find a reference value, and its confidence limits, from the overlapping area of 2 distributions when population means were allowed to differ by a pre-set amount. The allowable limits were compared with the overlapping area observed between data distributions reported from 3 laboratories. The simulation was run in R language. A description for the experimental setting was added, with the instructions for the simulation reported in structured English. RESULTS: The simulation allowed estimation of a limit to be used as a reference value when comparing two overlapping areas. Based on these limits, one laboratory in the study was found not to be aligned with the others. CONCLUSIONS: Computer simulation is a low-cost, powerful, and easy to implement tool which may help in solving simple or complex problems, when assumptions are unrealistic or unmet, or when mathematical algorithms are not known or difficult to calculate.

Evaluation of the analytical performance of three chemiluminescence serological assays for detecting anti-SARS-CoV-2 antibodies.

The serology surveillance of SARS-CoV-2 antibodies represents a useful tool for monitoring protective immunity in the population. We compared the performance of three SARS-CoV-2 antibody serological immunoassays in 600 vaccinated subjects after the BNT162b2 mRNA COVID-19 vaccine. All serum samples were evaluated by three different immunoassays for detecting anti-SARS-COV-2 antibodies. All SARS-CoV-2 antibody serological immunoassays could detect, when present, a post-vaccine humoral immune response. Median (interquartile range, IQR) anti-S-RBD IgG, Access SARS-CoV-2 IgG (1st IS) and Access SARS-CoV-2 IgG II levels of the subjects investigated were, respectively, 687 BAU/mL (131-2325), 419 IU/mL (58-1091) and 104 AU/mL (14-274). By studying a cohort of unvaccinated subjects, without previous COVID-19 infection, we found a high specificity for all methods. A high correlation was found between IgG titres. Considering the kinetics of subjects with multiple doses, we observed that percentage decreasing gradients were comparable across methods. Our results suggest that all the SARS-CoV-2 antibody serological immunoassays evaluated in this study are suitable for monitoring IgG titers over time. This study contributes to a better understanding of antibody response in vaccinated subjects using some currently available assays.