How clinical laboratories select and use Analytical Performance Specifications (APS) in Italy.

OBJECTIVES: Even if the topic of the analytical quality required to provide laboratory results "fit for purpose" exists since the beginning of the modern medical laboratory, there is the suspect that the expression "Analytical Performance Specifications" (APS) is not well-known. To investigate this aspect a survey was conducted. METHODS: A questionnaire with seven questions related to the knowledge about the topic, the sources of information and the criteria used by the laboratories to set the APS and their applications was prepared. It was distributed to all the clinical pathology laboratories of Lombardy Region (143) and to the members of SIBioC Laboratory Medicine (excluding Lombardy). RESULTS: We received 201 replies: 127 from Lombardy and 74 from the rest of Italy. Fifteen percent declared to ignore the meaning of APS and only 64 % of those knowing the meaning of the term declared to use them in the daily practice. The state-of-the-art was the principle used more frequently to set APS (about 48 %) followed by biological variation (41 %), and APS were typically applied to set goals for Internal Quality Control for selected measurands. Usually imprecision or total error APS were used, much less frequently uncertainty APS. In fact only 27 % of the laboratories declared to have calculated the measurement uncertainty for part or the majority of their measurands. CONCLUSIONS: Even considering the limits of a survey that relies upon self-declarations, it appears clearly that, at list in Italy, there is some work to be done to promote the concept and the use of APS.

Enhanced laboratory surveillance of respiratory infection disclosed the rapid rise of enterovirus D68 cases, northern Italy, August to September 2024.

We report a considerable increase in enterovirus D68 (EV-D68) cases since July 2024, culminating in an ongoing outbreak of acute respiratory infections in northern Italy, accounting for nearly 90% of all enterovirus infections. The outbreak was identified by community- and hospital-based surveillance systems, detecting EV-D68 in individuals with mild-to-severe respiratory infections. These strains belonged to B3 and a divergent A2 lineage. An increase in adult cases was observed. Enhanced surveillance and molecular characterisation of EV-D68 across Europe are needed.

Digital transition in pathology lab: a survey from the Lombardy region.

Objective: Digital pathology is an opportunity to revise the routine and old artisanal workflow, moving to standard operating procedures, quality control and reproducibility. Here the results of a survey promoted by the Coordinamento della Medicina di Laboratorio (CRC Med Lab) of the Lombardy region in Italy are reported to shed light on the current situation of digital adoption in the country. Methods: The survey composed of 58 questions was sent to 60 pathology laboratories. The results were collected and most significant answers were reported and discussed. Results: Answers were received from 57 (95%) laboratories, a minority organized in spoke-hub networks (16%) with a centralized processing phase (11%). Hybrid manual/computer-assisted traceability was prevalent (36%), with QR/barcode labeling starting within the pathology lab (23%). Different laboratory information systems (LIS) were employed, mostly with alert functions and/or multimedial file attachments (56% and 46%, respectively). The majority opted for a semi-automated tracking management (44, 77%) and 18 centers (32%) were partly digitizing the routine (¾ scanning < 25% of slides). Whole slide images were retained for 3.7 years in average; in-house blocks/slides archiving was still preferred (30, 53%), with 1838 (±1551) and 1798 (±1950) days (5 years) internal permanence for blocks and slides that are stored in out-source (mean turnaround time for return on-demand 3.7±2.1, range 1-10 days). Conclusions: The advantages of digital pathology must be balanced against the challenges faced in the structural revision of the pathology workflow. This regional scouting can represent the foundation to build an efficient and connected digital pathology system in the territory.

A territory-wide opportunistic, hospital-based HCV screening in the general population from northern Italy: The 1969-1989 birth-cohort.

BACKGROUND AND AIM: The World Health Organization (WHO) goal of Hepatitis C Virus (HCV) elimination by 2030 rose awareness about the need of screening plans, worldwide. In Italy, graduated screening starting from people born in 1969-1989 might be the most-effective strategy. We performed an opportunistic HCV screening study in the general population attending health facilities in Lombardy region, Northern Italy. METHODS: This is a prospective, multicenter, territory-wide, opportunistic study supported by the Regional Government of Lombardy, Italy. Between June 2022 and December 2022, all subjects born in 1969-1989, hospitalized or accessing blood collection centres were offered anti-HCV and HCV-RNA tests. Patients with known anti-HCV positivity and/or previous anti-HCV treatment were excluded. Demographic features were uploaded into a regional web-based platform. RESULTS: In total, 120 193 individuals were screened in 75 centres. Mean age was 44 (±6) years, 65.2% were females, 83.7% were tested at blood collection centres. Anti-HCV tested positive in 604 (0.50%) subjects: mean age 47 (±5), 51.1% females. HCV seroprevalence was higher in males (p < 0.00001), elderly (p < 0.00001) and in- vs. outpatients (p = 0.0009). HCV-RNA was detectable in 125 out of 441 (28.3%) anti-HCV positive subjects. Actively infected patients were 46 (±6) years old, mainly males (56.8%). The overall prevalence of active HCV infection was 0.10%, higher in elderly (p = 0.0003) and in in-patients (p = 0.0007). Among 93 HCV-RNA positive patients, the median age was 48 years, 58% males, 62% Italian born, median HCV-RNA levels were 6,1 log IU/mL, liver stiffness measurement (LSM) values 5.5 (3.1-29.9) kPa and ALT levels 48 U/L. CONCLUSIONS: The prevalence of active HCV infection in the 1969-1989 population attending health facilities in Lombardy was low. Most viremic patients were Italian-born, with mild liver disease but high-HCV-RNA levels. Due to the higher prevalence in the elderly, the extension of such opportunistic screening programs to lower birth cohorts would be warranted.

Covid-19 and gender: lower rate but same mortality of severe disease in women-an observational study.

BACKGROUND: Gender-related factors might affect vulnerability to Covid-19. The aim of this study was to describe the role of gender on clinical features and 28-day mortality in Covid-19 patients. METHODS: Observational study of Covid-19 patients hospitalized in Bergamo, Italy, during the first three weeks of the outbreak. Medical records, clinical, radiological and laboratory findings upon admission and treatment have been collected. Primary outcome was 28-day mortality since hospitalization. RESULTS: 431 consecutive adult patients were admitted. Female patients were 119 (27.6%) with a mean age of 67.0 ± 14.5 years (vs 67.8 ± 12.5 for males, p = 0.54). Previous history of myocardial infarction, vasculopathy and former smoking habits were more common for males. At the time of admission PaO2/FiO2 was similar between men and women (228 [IQR, 134-273] vs 238 mmHg [150-281], p = 0.28). Continuous Positive Airway Pressure (CPAP) assistance was needed in the first 24 h more frequently in male patients (25.7% vs 13.0%; p = 0.006). Overall 28-day mortality was 26.1% in women and 38.1% in men (p = 0.018). Gender did not result an independent predictor of death once the parameters related to disease severity at presentation were included in the multivariable analysis (p = 0.898). Accordingly, the Kaplan-Meier survival analysis in female and male patients requiring CPAP or non-invasive ventilation in the first 24 h did not find a significant difference (p = 0.687). CONCLUSION: Hospitalized women are less likely to die from Covid-19; however, once severe disease occurs, the risk of dying is similar to men. Further studies are needed to better investigate the role of gender in clinical course and outcome of Covid-19.

Platelet Transfusion Thresholds: How Low Can We Go in Respect to Platelet Counting?

Platelet transfusion is conventionally used to prevent or treat bleeding in patients with low platelet counts or impaired platelet function. The identification of accurate thresholds of platelet count for guiding platelet transfusion practices is a crucial aspect in health care to prevent adverse events, side effects, unwarranted costs for the health care service, and deprivation of supplies. This article is therefore aimed at providing a narrative overview on current guidelines and recommendations for platelet transfusion across many clinical settings, including platelet function disorders, and critically analyzing the available platelet transfusion thresholds according to the current analytical performance of platelet counting with automated hematological analyzers. Overall, universal agreement on the definition of platelet transfusion thresholds has not been reached. The degree of accuracy and imprecision of many fully automated hematological analyzers appears also unsatisfactory, especially at the lower thrombocytopenic range, and this may thus jeopardize the managed care of patients who are candidates for platelet transfusions. Potential solutions to overcome the current shortcomings of automated platelet counting are also discussed, encompassing the use of alternative tests for guiding platelet transfusion (e.g., thrombin generation assays or thromboelastography) along with innovative approaches for platelet enumeration (e.g., fluorescent labeling and flow cytometry).

Age-specific SARS-CoV-2 infection fatality ratio and associated risk factors, Italy, February to April 2020.

We analysed 5,484 close contacts of coronavirus disease (COVID-19) cases in Italy, all tested for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Infection fatality ratio was 0.43% (95% confidence interval (CI): 0.21-0.79) for individuals younger than 70 years and 10.5% (95% CI: 8.0-13.6) for older individuals. Risk of death after infection was 62% lower (95% CI: 31-80) in clusters identified after 16 March 2020 and 1.8-fold higher for males (95% CI: 1.03-3.16).

Preliminary evaluation of a new flow cytometry method for the routine hematology workflow.

Background In a generalist laboratory, the integration of the data obtained from hematology analyzers (HAs) with those from multiparametric flow cytometry (FMC) could increase the specificity and sensitivity of first level screening to identify the pathological samples. The aim of this study was to perform a preliminary evaluation of a new simple hybrid method (HM). The method was obtained by integration between HAs reagents into FCM, with a basic monoclonal antibodies panel for the leukocytes differential count. Methods Eighty-one peripheral blood samples, collected in K3EDTA tubes, were analyzed by XN-module, and CyFlow Space System, using both standard MoAbs and HM method analysis, and with the optical microscopy (OM). Within-run imprecision was carried out using normal samples, the carryover was evaluated, data comparison was performed with Passing-Bablok regression and Bland-Altman plots. Results The within-run imprecision of HM methods ranged between 1.4% for neutrophils (NE) and 10.1% for monocytes (MO) always equal or lower to the OM. The comparison between HM methods vs. OM shows Passing-Bablok regression slopes comprised between 0.83 for lymphocyte (LY) and 1.14 for MO, whilst the intercepts ranged between -0.18 for NE and 0.25 for LY. Bland-Altman relative bias was comprised between -12.43% for NE, and 19.77% for eosinophils. In all 11 pathological samples the agreement between the methods was 100%. Conclusions The new hybrid method generates a leukocytes differential count suitable for routine clinical use and it is also useful for identifying morphological abnormalities with a reduction in cost and improvement of screening for first level hematology workflow.

Harmonization of interpretative comments in laboratory hematology reporting: the recommendations of the Working Group on Diagnostic Hematology of the Italian Society of Clinical Chemistry and Clinical Molecular Biology (WGDH-SIBioC).

The goal of harmonizing laboratory testing is contributing to improving the quality of patient care and ultimately ameliorating patient outcome. The complete blood and leukocyte differential counts are among the most frequently requested clinical laboratory tests. The morphological assessment of peripheral blood cells (PB) through microscopic examination of properly stained blood smears is still considered a hallmark of laboratory hematology. Nevertheless, a variable inter-observer experience and the different terminology used for characterizing cellular abnormalities both contribute to the current lack of harmonization in blood smear revision. In 2014, the Working Group on Diagnostic Hematology of the Italian Society of Clinical Chemistry and Clinical Molecular Biology (WGDH-SIBioC) conducted a national survey, collecting responses from 78 different Italian laboratories. The results of this survey highlighted a lack of harmonization of interpretative comments in hematology, which prompted the WGDH-SIBioC to develop a project on "Harmonization of interpretative comments in the laboratory hematology report", aimed at identifying appropriate comments and proposing a standardized reporting system. The comments were then revised and updated according to the 2016 revision of the World Health Organization classification of hematologic malignancies. In 2016, the Working Group on Diagnostic Hematology of the Italian Society of Clinical Chemistry and Clinical Molecular Biology (WGDH SIBioC) published its first consensus based recommendation for interpretative comments in laboratory hematology reporting whit the purpose of evaluating comments and the aim to (a) reducing their overall number, (b) standardizing the language, (c) providing information that could be easily comprehended by clinicians and patients, (d) increasing the quality of the clinical information, and (e) suggesting additional diagnostic tests when necessary. This paper represents a review of the recommendations of the former document.

Short- and medium-term biological variation estimates of red blood cell and reticulocyte parameters in healthy subjects.

BACKGROUND: The integrated evaluation of traditional and innovative red blood cell (RBC) and reticulocyte parameters is a rapid, inexpensive and non-invasive diagnostic tools for differential diagnosis and follow-up of anemia and other pathological conditions needing bone marrow erythropoiesis assessment. Therefore, estimating the biological variation (BV) of these parameters is essential for evaluating the analytical performance of hematological analyzers, and for enabling accurate data interpretation and appropriate clinical management. This study aims to define short- and medium-term BV estimates and reference change value (RCV) of RBC and reticulocyte parameters. METHODS: Twenty-one healthy volunteers participated in the assessment of medium-term BV (blood sampling once/week, five consecutive weeks) and 22 volunteers in the assessment of short-term BV (blood sampling once/day, five consecutive days) using Sysmex XN. Outlier analysis was performed before CV-ANOVA, to determine BV estimates with confidence intervals (CI). RESULTS: Medium- and short-term within-subject BV were between 0.3% and 16.4% and 0.2%-10.4% (MCH and IRF), respectively, whereas medium and short-term between-subjects BV ranged between 0.9% and 66.6% (MCHC and Micro-R) and 1.4%-43.6% (MCHC and IRF), respectively. The RCVs were similar for all parameters in both arms of the study, except for hemoglobin, RDW-CV and MCV. CONCLUSIONS: This study allowed for estimating the BV of many RBC and reticulocyte parameters, some of which have not been currently explored. For RBC, hemoglobin, RDW-CV and MCV it seems advisable to use RCV calculated according to monitoring time and/or differentiated by sex. As regards analytical goals, we suggest using the most stringent targets found in the short-term arm of this study.

Validation rules for blood smear revision after automated hematological testing using Mindray CAL-8000.

BACKGROUND: This article was aimed to test the use of validation rules for blood smear review after automated hematological testing using Mindray CAL-8000 (two hematological analyzers and one autoslider). METHODS: This study was based on 1013 peripheral blood samples (PB) referred for routine hematological testing. Results of testing on CAL-8000 were analyzed using both locally derived and International Consensus Group for Hematology (ICGH) validation rules, and then compared with data obtained by optical microscopy (OM). A workflow analysis was also completed. RESULTS: The overall agreement with locally derived and ICGH criteria was 91% and 85%, but a higher sensitivity was observed for locally derived criteria (0.97 vs 0.95). The percentage of false negative and false positive samples was 2.1% and 7.1% using ICGH criteria, and was 1.4% and 14% using locally defined rules. The throughput of CAL-8000 system was 208 samples/h, with a percentage of OM analysis comprised between 14% and 17%, and sensitivity of 0.97. As regards personnel activity, we estimated 0.8 full-time equivalent (FTE) of technical staff and 0.7 FTE of personnel for clinical validation of data and blood smear review. CONCLUSION: These results show that customization of validation rules is necessary for enhancing the quality of hematological testing and optimizing workflow.

Evaluation of Mindray BC-6800 body fluid mode for automated cerebrospinal fluid cell counting.

BACKGROUND: Cellular analysis in cerebrospinal fluid (CSF) provides important diagnostic information in various medical conditions. The aim of this study was to evaluate the application of Mindray BC-6800 body fluid (BF) mode in cytometric analysis of CSF compared to light microscopy (LM). METHODS: One hundred and twenty-nine consecutive CSF samples were analyzed by BC-6800-BF mode as well as by LM. The study also included limits of blank (LoB), limit of detection (LoD), limit of quantitation (LoQ), carryover and linearity. Results White blood cells LoQ was 4.0×106 cells/L. Linearity was good and carryover was negligible. As for the total and white blood cells, the BC-6800-BF parameters vs. LM showed both bias ranged from -10.28 to 0.06×106 cells/L. Polymorphonuclear and mononuclear cells ranged from 6.64 to 10.90%. For white blood cell the diagnostic agreement was 93% at the cut-off >5.0×106 cells/L, and for polymorphonuclear and mononuclear at the cut-off >50% was 91% and 92%, respectively. CONCLUSIONS: BC-6800-BF offers rapid and accurate counts in clinically relevant concentration ranges, replacing LM for most samples. However, in samples with abnormal cell counts or with abnormal white blood cell differential scattergrams the need to microscopic review for a correct clinical outcome remains.

Automated Cerebrospinal Fluid Cell Counts Using the New Body Fluid Mode of Sysmex UF-1000i.

BACKGROUND: We evaluated the new body fluid module on Sysmex UF1000-i (UF1000i-BF) for analysis of white blood cell (WBC) and red blood cell (RBC) in cerebrospinal fluid (CSF). METHODS: WBC and RBC counting were compared between UF1000i-BF and Fuchs-Rosenthal counting chamber in 67 CSF samples. This study also included the evaluation of between-day precision, limit of blank (LoB), limit of detection (LoD), functional sensitivity (limit of quantitation, LoQ), carryover and linearity. Diagnostic agreement for differentiation between normal and increased WBC counts (≥5.0 × 10(6) /L) was also assessed. RESULTS: The agreement between UF1000i-BF and manual WBC counts was otpiaml in all CSF samples (r = 0.99; y = 1.05x + 0.09). A modest overestimation was noticed in samples with WBC < 30 × 10(6) /L (r = 0.95; y = 1.21x - 0.15). A good agreement was observed for RBC counts (r = 0.98; y = 1.15x + 0.55), particularly in samples with RBC ≥ 18 × 10(6) /L (r = 0.98; y = 1.01x + 8.90). Between-day precision was good, with coefficient of variations (CVs) lower than 7.2% for both WBC and RBC. The LoBs were 0.1 × 10(6) WBC/L and 1.2 × 10(6) RBC/L, the LoDs were 0.7 × 10(6) WBC/L and 5.5 × 10(6) RBC/L, the LoQs were 2.4 × 10(6) WBC/L and 18.0 × 10(6) RBC/L, respectively. Linearity was excellent (r = 1.00 for both WBC and RBC). Carryover was negligible. Excellent diagnostic agreement was obtained at 4.5 × 10(6) WBC/L cut-off (sensitivity, 100%; specificity, 97.4%). CONCLUSION: The UF1000i-BF provides rapid and accurate WBC and RBC counts in clinically relevant values of CSF cells. The use of UF1000i-BF may hence allow to replace routine optical counting, except for samples displaying abnormal WBC counts or abnormal scattergram distribution, for which differential cell counts may still be required.

Opportunities and Drawbacks of Digitalized Morphologic Analysis of Body Fluids.

Body fluid analysis has become a critical component of diagnostic and clinical decision-making for a wide spectrum of human pathologies. An automated microscope, a high-quality digital camera, and a software designed to identify and automatically preclassify cells and other features in stained smears comprise the most recent generation of digital morphologic analyzers. The time necessary for expert operator reclassification is another aspect that must be considered at this stage of development, because identifying and sorting distinct elements in body fluids still necessitates the involvement of an expert morphologist.

JAK2V617F mutation and hydroxyurea treatment as determinants of immature platelet parameters in essential thrombocythemia and polycythemia vera patients.

Immature platelets (IPFs), which are hemostatically more active than mature platelets, have been found elevated in essential thrombocythemia and polycythemia vera, 2 myeloproliferative neoplasms (MPN) characterized by an increased risk of thrombosis. It is not known whether the IPF levels are influenced by pathogenetic factors, including JAK2V617F mutational status, or by treatment regimen. To address this point, in 46 essential thrombocythemia and 38 polycythemia vera consecutive patients, we measured IPF and correlated the results to JAK2V617F mutation and myelosuppressive treatment with hydroxyurea. This analysis provides 2 new elements regarding IPF and MPN. The first finding is that the JAK2V617F mutation is linked to the quantity of IPF in patients with MPN, which might contribute to the prothrombotic phenotype in these patients. The second finding is that IPF is susceptible to myelosuppressive treatment, which may additionally explain the favorable effect of hydroxyurea therapy on MPN outcome as well as the associated thrombotic risk.