From sepsis to COVID-19: the emerging role of Monocyte Distribution Width (MDW)

Immune activation of the monocyte-macrophage population plays a pivotal role in the systemic hyper-inflammatory response, typically observed during severe dysimmune diseases, such as sepsis and COVID-19. In this commentary, we have reviewed the literature data on the novel cytometric marker of monocyte activation, known as MDW (Monocyte Volume Distribution Width), a monocyte dimensional parameter obtainable by blood count examination, which has recently been approved for clinical use as "Early Sepsis Indicator" (ESId), in patients accessing the Emergency Department. In particular, in this Opinion paper, we highlight the main clinical applications and relevant perspectives of this new test: from its use for (early) diagnosis of sepsis, in different hospital settings, to its emerging prognostic role in patients with COVID-19, as a biomarker of disease severity. In view of the reported evidence, we discuss the clinico-pathological notion that, basically, severe COVID-19 can be considered a new form of viral sepsis. Further clinical studies are needed to better understand the pre-analytical and analytical variables of this parameter, correlate MDW dynamics with those of other humoral and cytometric markers, and validate the new diagnostic and prognostic applications of MDW on large multicenter case series. Copyright © 2022 Biomedia. All rights reserved.

Evaluation of SARS-CoV-2 neutralizing antibodies before and after III dose of the vaccine

Introduction We all know that the most important task in combating COVID-19 pandemic is to produce enough effective vaccines and the greatest number of vaccinated subjects within a time frame. Nevertheless, the goal of this worldwide effort should be aligned to raise protective level of neutralizing antibodies (NAb) in vaccinees. It is clear that the NAb can block a viral invasion at the initial access to human receptors. Some methods for NAb measurement are commercially available.In this study, we evaluate the immune response of all employees receiving the III dose of BNT162b2 mRNA vaccine at November 2021. Serological NAb determination was performed before the administration of the III dose of vaccine (T0) and then 1 (T1) and 3 months (T2). Methods We collected serum samples of 46 laboratory workers at T0, T1 and T2. We determined the concentration of Anti-S IgG with SARS-CoV-2 IgG II Quant kit by chemiluminescence method on Alinity i Abbott (cutoff 50 AU/mL) in all samples and of NAb in 11 laboratory workers at T0, T1, T2 using SARS-CoV-2 Anticorpi Neutralizzanti kit (SGM Italia), an immunoturbidimetric method (high concentration >30 AU/mL, high % inibition >56%) on Alinity i Abbott platform. This method is able to detect NAb which bind specifically to the binding domain of the RBD receptor blocking the human ACE2 receptor. Statistical analysis was performed using MedCalc Software Ltd. Results At T0, T1 and T2, the mean+/-standard deviation concentration for anti-S IgG is 1016+/-1462, 22239+/-22480, 41777+/-30778 AU/mL and for NAb is 33+/-24, 100+/-100, 95+/-12, and the mean inibition percentage (%NAb) is 66+/-51, 309+/-105 and 320+/-164, respectively. Comparison between anti-S IgG and NAb at T0 showed a good correlation (R2=0.88);the NAb concentration was upper of linearity an all subjects at T1 and T2. Comparison between anti-S IgG and %NAb showed the same trend. Discussion The anti-S IgG are significantly reduced after 6 months from II dose of vaccine and increased about 30 times at T1. Concentration become half at T2 in all subjects who not affected by SARS-CoV-2 between T1 and T2. Although all subjects had a high %NAb at T0 that becomes 5 times higher at T1 and remains high at T2, 7 out of 11 subjects have been infected by SARS-CoV-2. Further studies are needed to define better the SARSCoV-2 neutralizing activity and the suitable routine test to measure them.

Are polymorphisms of ACE1, ACE2, IFTM3 and TMPRSS2 genes associated with susceptibility and severity of SARS-CoV-2 infection? A systematic review and meta-analysis

Background: Some human polymorphisms of angiotensin converting enzyme 1 (ACE1), ACE2, Type 2 transmembrane serine proteases (TMPRSS2) and interferon-induced transmembrane protein 3 (IFITM3) genes may have an effect on the susceptibility to SARSCoV-2 infection and increase the risk to develop severe COVID-19.We conducted a systematic review of current evidence to investigate the association of genetic variants of these genes with the susceptibility to virus infection and patient prognosis. Method(s): We systematically searched Medline, Embase and The Cochrane Library for articles published until May 2022, and included observational studies covering genetic association of ACE1, ACE2, TMPRSS2 and IFITM3 with COVID-19 susceptibility or prognosis.The study selection and review process were performed by 2 authors independently.We pooled data as convenient in meta-analyses using the Mantel-Haenszel Method (RevMan5.4).Odds ratio(OR) values and 95% confidence intervals(CI) were calculated. Heterogeneity among studies was tested using I2 statistic tests. In presence of substantial heterogeneity (I2>50%), the random-effects model was used to analyse the data. Result(s): The literature search identified 1,968 references.Thirty-three studies (19 on ACE1, 7 on ACE2, 5 on TMPRSS2 and 5 on IFITM3), enrolling 4,067 COVID-19 patients, met the inclusion criteria and were included. ACE1 rs4646994 and rs1799752, ACE2 rs2285666, TMPRSS2 rs12329760 and IFTM3 rs12252 were identifies as common polymorphisms.Our meta-analyses showed an association between genetic polymorphisms and susceptibility to SARS-CoV-2 infection for IFTM3 rs12252 TT (OR 0.56;95%CI 0.39-0.79, p<0.0001, I2=0%) and CT (OR 1.64;95%CI 1.15-2.33, p<0.0001, I2=0%) genotypes and for ACE1 rs4646994 DI (OR 0.57;95%CI 0.34-0.96;p=0.03, I2=.92%).Likewise, meta-analyses uncovered that both ACE1 rs4646994 DD (OR 1.26;95%CI 1.03-1.53, p=0.02, I2=48%) and rs12252 IFITM3 CC (OR 2.26;95%CI 1.05-4.89, p=0.04, I2=0%) genotypes carriers had a significantly increased risk of developing severe COVID-19. Discussion(s): These results provide a critical evaluation of genetic polymorphisms as predictors in SARS-CoV-2 infection.ACE1 DD and IFITM3 CC polymorphisms would lead to a genetic predisposition for severe lung injury in patients with COVID-19.

Comparative Assessment of Maglumi Sars-Cov-2ag Test and Nucleic Acid Apmplification Test (Naat): First Considerations

Introduction: NAAT is still the international reference assay for the diagnosis of Covid-19 due to high sensitivity and specificity;it is able to detect the pathogen even at low viral load, nevertheless a positive PCR result demonstrates the presence of nucleic acid in the sample, but not if it contains an infecting virus;the risk of unnecessarily isolate a person who is no longer infectious and a long TAT can preclude a screening utility. Conversely, immunoassays targeted to detect viral antigens have the potential to fit the requirements for a screening population test. In this study, we evaluated the consistency of results obtained by Maglumi SARS-CoV-2 Ag test in comparison with molecular test. Method(s): 79 positive NAAT nasopharyngeal swabs (NPS) with Ctthreshold cycle (Ct) between 39 and 13 were selected and analysed with MAGLUMI CLIA assay targeted to nucleocapsid SARS-Cov-2 antigen (M) by Snibe Diagnostic. 37 NPS were also analysed with Liaijson XL (L), DiaSorin s.p.a., which is routinary used in our lab. 5 negative NPS were included in the experiment as negative controls. Result(s): all 5 negative NPS were confirmed negative with both CLIA assays. 33 of 79 (41,8%) positive NPS were confirmed positive on M, all these samples had a Ct < 26. The remaining 46 positive NPS, resulting negative on M, had a Ct > 24. Results for 37 NPS tested both with M and L agreed 100%. Conclusion(s): CLIA test demonstrated to be less sensitive to the viral presence than NAAT, in particular M test seems to be able to detect a positivity to SARS-Cov-2 when the viral load is detected at a CT < 24. The grey area, where NAAT and CLIA were not in perfect agreement, seems to be Ct= 24 / 25. In this area M missclassified 67% of positive NPS, but further investigation with a larger number of samples is necessary to confirm this finding. While PCR positivity may persist for several weeks after the onset of the disease and the disappearance of symptoms, Ag tests reach satisfactory sensitivities when infected people are more likely to be contagious;together with the high throughput of the technology, makes them an extremely useful tool for screening population, especially during the pandemic.

suPAR evaluation in patients with SARS-CoV-2 infection: our experience

Introduction Elevated soluble urokinase Plasminogen Activator Receptor (suPAR) is a biomarker associated with adverse outcomes. We aimed to investigate the associations among plasma suPAR levels (testing the cut-offs <=4 and >=6 ng/mL that supports patient discharge/hospitalisation, respectively) with other biomarkers such as PCR, PCT, IL-6 and with sex, age, discharge/death and WHO disease severity in patients tested positive for SARS-CoV-2. Methods We performed an observational cohort study of 99 patients (37 females, 62 males) presenting with COVID-19 symptoms at Department of Infectous and Critical Care of our Hospital in April 2020. Plasma suPAR was measured using suPARnostic kit (Virogates, Denmark), an immunoturbidimetric method on Abbott Alinity i platform. Patients were followed for development of mechanical ventilation, mortality or discharged. Statistical analysis was performed using Principal Component Analysis (PCA) that can be applied to datasets to obtain a simplified model for stratifying patients by reducing the number of variables. PCA weights the variables according to their relative importance. This method, in our case, can aid in determining key variables in management of patients affected by SARS-CoV-2. Results The mean age was 58 years;women had a higher concentration average of suPAR (8.9 vs 8.3 ng/mL) but the subdivision by sex did not determine any clustering. All variables showed a positive correlation with disease severity, better with IL-6 and suPAR (IL-6=25.3%, suPAR=24%, age=16.4%, PCT=15.4%, PCR=17.2%), allowing a subdivision of 3 groups (severe/ critic: IL-6=227.65 pg/mL, suPAR=9.26 ng/mL;moderate: IL-6=48.1 pg/mL, suPAR=7.35 ng/mL, paucisymptomatic: IL-6=3.7 pg/mL, suPAR=2.78 ng/mL). Combining the variables and discharge/death outcome showed positive correlation although this did not result any clear clustering (n.78 discharged: IL-6=214 pg/mL, suPAR=8.23 ng/mL;n.14 dead: IL-6=286 pg/mL, suPAR=11.31 ng/mL). Discussion Our data show that suPAR levels increase as the disease worsens. Statistical analyses demonstrated that suPAR levels are positively correlated with age and IL-6 levels. Therefore, further evaluation of suPAR plasma levels in different symptoms of COVID-19 patients could still provide important indications for early admission and treatment.

Evaluation of SARS-CoV-2 antibodies dinamics after booster dose

Introduction At november 2021 in our public hospital, all employees have been received the III dose of SARS-CoV-2 vaccine with BNT162b2 mRNA. To evaluate the immune response to the vaccine, the antibodies measurement directed against the S protein or, more specifically, against the RBD domain stimulated by vaccination, was performed. The aim of this study is to evaluate SARS-CoV-2 antibody dinamics over 6 months after vaccine (at T0, T1, T2, T3: before III dose, at 1, 3 and 6 months, respectively). Methods We collected serum samples from 46 laboratory workers at T0, T1, T2, T3 (between november 2021 and may 2022). Serologic testing for specific SARS-CoV-2 anti-RBD IgG were performed by chemiluminescence method on Alinity Abbott instrument;according to manifacturer, the cut-off is 50 AU/mL. All samples were tested also for anti-N IgG with chemiluminescence assay on Alinity Abbott instrument (cutoff is 1.4 AU/mL) to evaluate subjects affected by COVID-19. Results At T0, the mean concentration of anti-S IgG is 581+/-303 AU/mL for 42 subjects (91%) that received II dosis until to April 2021 whereas the mean concentration is 3963+/-1386 AU/mL for 4 workers (9%) affected by SARS-CoV-2 between I dosis and november 2021. At T1, the mean concentration of anti-S IgG is 23326+/- 15905 AU/mL (min/ max value 4766 and 69816 AU/mL) without difference among subjects that had the upper concentration at T0. At T2, the mean concentration of anti-S IgG is 27374+/-27057 AU/mL (min/max value 1038 and 80000 AU/mL) with mean concentration of 60561+/-22281 AU/mL in 6 subjects affected by SARS-CoV-2 between T1 and T2.At T3, the mean concentration of anti-S IgG is 20610+/-12403 AU/mL (min/max value 1825 and 39796 AU/mL) with mean concentration of 22525+/-10121 AU/mL in 5 subjects affected by SARS-CoV-2 between T2 and T3. Discussion We found that booster dose of the vaccine triggers robust immune responses in healthy recipients, COVID-19 triggers an earlier and more intense immune response even after 2-3 months from III dose;in all cases, however, antibody titers remain at high levels in COVID-19 recovered patients. Although virus infection among vaccinated subjects is rare, this would seem to promote a more intense immune response after boosting dose, inducing antibody titers significantly higher and likely more durable.

Big Data analysis to evaluate the clinical utility of IgM anti SARS-CoV-2 determination: The Modena experience

Introduction: since the scarce diagnostic accuracy of specific circulating antibodies for SARS-CoV-2, we aimed to assess the clinical utility of IgM detection in SARS-CoV-2 using the Big Data analysis. Methods: this is a retrospective study;all the blood samples collected between March and September 2020 were processed using a lateral flow immunoassay (LFIA) kit for IgG and IgM antybody testing. Positives results were tested again using a chemiluminescent method. Subjects confirmed with a positive result were contacted for a molecular test. Results: more than 69 000 serological tests (from 42 911 subjects) were performed. 94.5% (40 559/42 911) of subjects had negative results for both IgG and IgM. 1.5% (n = 640) subjects had both IgG and IgM positive results, and viral RNA research confirmed positivity in 16% (85/533). Among subjects with IgG negative and IgM positive results (n=271), a positivity was confirmed in 1% (4/270). Conversely, in subjects with IgG positive and IgM negative results, a positivity was confirmed in 8% (97/1 215). Therefore, the analysis suggests that up to 98% of serological test results of IgM positivity and IgG negativity are false positive. Discussion: the study, based on Big Data analysis application, proved the scarce clinical utility of IgM detection in COVID-19 management, and underlines the responsibility of laboratory professionals in highlighting the limitations of the serological tests also due to uncertainty in their interpretation.

Mdw is a novel inflammatory biomarker with prognostic relevance in covid-19 patients

Monocyte Distribution Width (MDW), a new hematologic parameter correlating with cytomorphologic changes occurring during monocyte activation, has recently been described as promising early biomarker of sepsis. Similar to sepsis, in SARS-CoV-2-associated disease (COVID- 19), monocyte/macrophage subsets are considered key mediators of the life-threatening hyperinflammatory disorder -commonly defined as 'cytokine storm'- which is part of the complex infectionassociated immune dysregulation observed in severe COVID-19 cases (possibly, representing a new kind of viral sepsis). Thus, we aimed at investigating possible prognostic roles of MDW testing during monitoring of COVID-19 patients.In this work, we longitudinally measured MDW values in a cohort of 87 patients with molecularly-proven COVID-19 diagnosis, consecutively admitted to our intensive/subintensive clinics in early 2020. Statistical analyses were applied to correlate MDW values with common inflammatory markers, disease severity, clinical trajectories and final outcome.We found significant direct correlations between MDW and different inflammatory markers routinely assessed during hospitalization, namely CRP (p<0.001), fibrinogen (p<0.001) and ferritin (p<0.01). Moreover, high MDW values were remarkably associated with fatal outcome (AUC=0.76, sensitivity 0.75, specificity 0.70, MDW threshold 26.4;RR=4.91, OR=7.14). Furthermore, evaluating MDW dynamics in cases with longer followup, we frequently observed progressive MDW increments in patients with worsening inflammatory conditions, while clinical recoveries were consistently associated with MDW decreases.Our study shows, for the first time, that MDW can be useful in the prognostic monitoring of hospitalized COVID-19 patients, as it is: (i) easy and rapid to obtain, (ii) directly related to the activation state of a fundamental inflammatory cell subset (i.e. monocytes, pivotal both in cytokine storm and in sepsis immunopathogenesis), (iii) strongly correlated with clinical severity of COVID-19- associated inflammatory disorder, and, in turn, (iv) endowed with relevant prognostic significance. Additional studies are needed to define the role of MDW monitoring in other clinical settings, including COVID-19 outpatients.

The big data analysis application to evaluate the clinical utility of igm anti sars-cov-2 determination: the experience of modena

Background: Serological tests identifying SARS-CoV-2 IgG and IgM in serum play an important role in understanding the disease epidemiology. However, their immunological significance are currently undefined. There are many methods available for the detection of specific Abs whit suboptimal diagnostic accuracy and relatively high throughput capacity and less stringent specimen requirements compared to RNA-based assays. We aimed to assess the clinical utility of IgM detection in SARSCoV- 2 using the big data analysis. Methods: We conduct a retrospective study analyzing with a big data analysis all samples collected between 11 March and 30 September 2020. All serum samples received at the laboratory were processed using qualitative and commercially available rapid lateral flow immunoassay tests for 2019-nCoV IgG and IgM. Positive results were confirmed using a chemiluminescent method. Subjects with a positive result were contacted from the Department of Public Health for further tests (viral RNA research or subsequent serological tests) for definitive diagnosis.Results: A total of 69,343 serological tests (in 42,911 subjects) and 140,065 oropharyngeal or nasopharyngeal swabs (in 88,771 subjects) were performed. 94.5% of subjects screened (n=40,559) had negative results for both IgG and IgM. Of the 640 subjects with both IgG and IgM positive results, viral RNA research confirmed positivity in 16%. Of the subjects with IgG negative and IgM positive results, a positivity was confirmed in 1.4% (n=7/478) subjects. Subsequent serological testing confirmed IgG positivity in 8 subjects (1.6%). Conversely, in subjects with IgG positive and IgM negative results, a positivity was confirmed in 7.9%. Therefore, analysis suggests that up to 94% of serological test results of IgM positivity and IgG negativity are false positive whereas, serological test results of IgG positive and IgM negative are confirmed true positives in around 7.9% of subjects. Discussion: Our study, based on big data analysis application, confirms the scarce clinical utility of IgM anti SARS-CoV-2 detection in COVID-19 management, and underlines the responsibility of laboratory medicine professionals to highlight limitations of the SARS-CoV-2 serological tests due to uncertainty in their interpretation.

Implementation and decentralization of the analytical process in molecular biology: The experience of Modena for COVID-19 diagnosis

Background: Real time reverse transcriptase polymerase chain reaction (RT-PCR) on clinical specimens is considered the first line test for the diagnosis of SARS-CoV-2 infection. This test involves different steps: RNA extraction, reverse trancription, PCR setting-up, amplification and analysis of results. To date, several analyzers for extraction and amplification phases are availabe, while few are able to guarantee the automation of the entire workflow. To optimise the allocation of swabs, with the aim of maximising the number of swabs tested and reducing the wait time for results, we applied a decentralysed system.Methods: In Provincia of Modena a network of 5 suburban laboratories referring to central laboratory was developed.The work was focused not only on the implementation of new analyzers, but also on the organization and consolidation of the whole workflow integrating the pre-analytical, analytical and postanalytical phases and including the molecular biology tests report on LIS (Laboratory Information System). Furthermore, all laboratory professionals attended a specific training.Results: Between march 2020 and June 2021 450,000 RT-PCR for RNA of Sars-Cov-2 research were performed. The central laboratory analyzes the majority of swabs (n= 259832, 58%), including those enrolled at drive through (44%). 76% were analyzed with a automated system, and 24% with a manual procedure. The laboratory is open every day including Sunday, from 8:00 a.m. to 8:00 p.m. engaging two biologists and five laboratory technicians per day. Our project allowed to increase significantly the number of swabs tested in a day (from 100 in march 2020 to 4600 in march 2021), guaranteeing their reporting within 3 hours in emergency or 24 hours for routine and drive setting.Conclusions: The employ of automated, user friendly and with a guided interface instrument facilited the entire workflow, reducing the operator work and the reporting time. RT-PCR executed with a manual procedure need of a specific expertise to read the reaction products, but the shorter reporting time makes it useful for swabs made in emergency. However, the presence of the barcode reader and the connectivity with the management LIS facilitated the traceability of the samples for the entire diagnostic pathway and the minimization of human error. The implementation of our workflow has involved an important rationalization and optimization of the staff, while integration of new knowledge about SARSCOV-2 and optimal analytical performance of analyzers has allowed a modern management of the samples maximising the laboratory's test capacity for RT-PCR tests.

Neurological complication of COVID-19: Two different Case Reports

Here we present two different cases of mild and severe neurological manifestation due to Covid-19 infection. The first case is a woman with anosmia and ageusia still present after 30 days after infection. The autoimmunity tests showed a significant alteration for antibodies against myelin-associated glycoprotein (MAG) and N-methyl-Daspartate (NMDA) receptor antibodies. The MR revealed a persistent hyperintensity on the olfactory nucleus, a typical sign of encephalitisl with no nasal obstruction or abnormalities. To promote the recovery of both the senses, the patient began an empirical therapy with vitamin C integrators with improvment of symptoms after ninety days. The second case is a woman admitted in ICU for acute severe respiratory sindrome. D-dimer, fibrinogen, prothrombin time (PT), and activated partial thrombopolastine time (aPTT) were no altered, IL-6 level was 123. 50 U/L. After few days patient presented a sudness loss of consciousness with coma due to an acute ischemia with emorragic infartion and after 48 hours she died. We discuss the possible mechanisms of olfactory bulb demage and encephalitis COVID-19 related as well as ischemic stroke, and intracranial hemorrhage. Consequently we try to understand how the laboratory can help physicians in the management of these patients.

Una diagnosi difficile in un paziente dializzato peritoneale

Introduction: Interleukin-6 (IL-6) is a pro-inflammatory cytokine secreted by lymphocytes, fibroblasts and macrophages involved in B-cell differentiation and stimulation of acute-phase proteins. IL-6 is associated with high serum levels in viral human infections such as hepatitis B and C virus, influenza virus, herpes simplex virus, HIV and also in coronavirus disease 2019. This cytokine is also produced by a wide array of intraperitoneal cells after the exposition to a local noxa patogena.Icodextrin has been associate with sterile peritonitis in patients on peritoneal dialysis (PD): this type of peritonitis is a cause of cloudy effluent and mild abdominal discomfort that resolved with the discontinuation of icodextrin. The diagnosis of icodextrinassociated peritonitis is critical to avoid unnecessary antibiotic prescription.Here, we described a case report of steril icodextrin-associated peritonitis coupled with elevation of IL-6 on peritoneal dialysate occurred in a male 76 years old undergoing PD. Methods: IL-6 (DXI 800 Beckman Coulter) and leucocyte count were performed on peripheral blood sample and peritoneal dialysate before and after icodextrin rechallenge. Serum reactive C protein (PCR), serum procalcitonin (PCT) and peritoneal effluent colture were performed before and after icodextrin rechallenge. Results: After 48 hours from the start of icodextrin rechallenge, peritoneal effluent became cloudy with a slight increase in leucocyte count (178 cells/microlitro with 11% neutrophil granulocytes, vs 62). Dyalisate colture, serum PCR and PCT resulted negative;leucocyte count in peripheral blood resulted normal (7300/μL). IL-6 level increased steeply in peritoneal effluent when compared to baseline (1124 vs 114 pg/mL) and subsided to baseline levels with the withdrawal of the icodextrin solution without increase in serum IL-6 (15.7 vs 12 pg/mL);likewise leucocyte count on peritoneal dialysate decreased (63 cells/μL). Discussion: IL-6 can be used, along with peritoneal leukocyte count, as a precocious and sensitive marker of local inflammation and, in these cases, to discriminate from non-inflammatory peritonitis.

Ad interim SIBioC recommendations for serological assessment of SARS-CoV-2 infection

The recent pandemic outbreak caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and associated with the pathology called COVID-19 (coronavirus disease 2019), has now become one of the most strenuous health care challenges since the emergence of the three pandemics caused by influenza viruses during the past century. Throughout the clinical decision-making of COVID-19, laboratory tests are essential for supporting the screening, diagnosis, prognostication and therapeutic monitoring of this severe infectious disease. Serological testing, that reflects the humoral immune response developing after interaction between the host and the virus (or its components), enables to garner a vast array of clinical information which can be especially used in seroprevalence or seroconversion studies. To this end, the Task Force on COVID-19 of the Italian Society of Clinical Biochemistry and Clinical Molecular Biology (SIBioC) has endorsed a series of technical, practical and clinical ad interim recommendations, aimed at facilitating and optimizing the introduction, clinical usage and governance of SARS-CoV-2 serological immunoassays in routine practice.

Kinetics and biological characteristics of humoral response developing after SARS-CoV-2 infection: Implications for vaccination

With the ongoing coronavirus disease 2019 (COVID-19) pandemic outbreak spreading all around the world, an extensive vaccination against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is now universally regarded as one of the most effective strategies for counteracting the unremittent spread of this novel coronavirus. Nonetheless, the reasonable need to identify segments of the population in which vaccination shall be prioritized for avoiding a possible shortage of vaccines seems to collide with indications provided by many national and international healthcare organizations, that endorse widespread vaccination irrespective of a positive history of prior symptomatic or asymptomatic SARS-CoV-2 infection. To this end, this document provides an ad interim guidance aimed at prioritizing SARS-CoV-2 vaccination in people who are more likely to be infected, re-infected and/or to develop more aggressive COVID-19 illness, essentially based on routine assessment and monitoring of anti-SARS-CoV-2 immune response.

Monocyte Distribution Width (MDW) as a new parameter useful in COVID-19 patients in emergency setting

INTRODUCTION AND AIM: Clinical laboratory plays a role in diagnosis (viral RNA research, antibody response) and evaluation of infection severity, progression and response to treatments of COVID-19 patients. The International Federation of Clinical Chemistry and Laboratory Medicine (IFCC) defined a panel of tests1 recommended for clinical evaluation including blood cell count, coagulation and biochemical tests. MDW (Monocyte Distribution Width) is a new parameter describing the standard deviation of monocytes' volume. MDW is included in the differential white blood cell count (CBC-Diff) and has been shown to be an early indicator of sepsis (Crouser2). The aim of the study is to verify the contribution of MDW in the diagnostic and prognostic pathway of patients with suspected SARSCoV-2 infection in Emergency Department (ED) setting. MATERIALS AND METHODS: The retrospective study included 189 consecutive patients entering the ED of Baggiovara Civil Hospital (Modena) tested for SARSCoV-2 RNA using oro-nasopharyngeal swab. A complete and differential white blood cell count (CBC-Diff) including MDW (K2 EDTA, DxH 900 Beckman Coulter Inc) has been performed for all patients. RESULTS: 123 patients were negative for viral RNA, and 66 were positive. Among the latter 25 were discharged and 41 hospitalized. New MDW parameter showed an AUC of 0.83 (sensitivity 84.9 and specificity 75.6) with a cut off 20.1 between negative and positive. Moreover different median values have been observed in negative (18.26), positive discharged (21.60) and positive hospitalized (23.29) patients. DISCUSSION and CONCLUSIONS: MDW, included in CBC-diff, is a parameter quickly available for the clinician without additional request. The observed differences are associated with different severity of the COVID-19 clinical history indicating a potential valuable prognostic role. MDW could be inserted in a diagnostic algorithm as a support for clinical evaluation. BIBLIOGRAPHY: 1IFCC Information Guide on COVID-19;2Crouser et al. "Improved Early Detection of Sepsis in the ED With a Novel Monocyte Distribution Width Biomarker" Chest 2017 Sep;152(3):518-526.

The Role of Eosinophilic Cationic Protein as a New Biomarker in SARS-CoV-2 positive patients with different severity of illness

Background: Severe acute respiratory syndrome coronavirus (SARS-CoV-2) causes coronavirus disease 2019 (COVID-19), with a clinical outcome ranging from mild to severe, including death. To date, it is unclear why some patients develop severe symptoms. The routine laboratory tests and host immunity in COVID-19 patients with different severity of illness were compared after patient admission and also after discharge from the hospital after some time for follow up. Eosinophilic Cationic Protein (ECP) has various biological activities, including antibacterial, antiviral, antiparasitic and neurotoxic functions, and it contributes to the regulation of fibroblast activity. ECP, also induces airway mucus secretion and interacts with the coagulation and complement systems. ECP has been developed as a marker for eosinophilic disease and quantified in biological fluids including serum, bronchoalveolar lavage and nasal secretions. It is found in diseases such as allergic asthma and allergic rhinitis but also occasionally in other diseases. Methods: We evaluated 59 positive patients who underwent a nasopharyngeal swab PCR analysis for SARS-CoV-2, monitoring them through ECP and many routine laboratory tests such as ferritin, lactate dehydrogenase, 25-hydroxyvitamin D (25(OH)D),C-reactive protein (CRP),IL-6, during the period from 20 March to 25 May 2020. ImmunoCAP FEIA Fluorometric Enzyme Immunocapture Assay (ThermoFisher Scientific) measures the level of ECP in serum. Result: A total of 59 COVID 19-positive patients (64 + or-10 years, male 74%) were classified as having mild(HB) (n = 33), severe (S) (n = 19) and extremely severe (ES) (n = 7) illness. ECP levels were significantly higher in" ES"[64,1 microg/L (18-117), p<0.001];"S" showed [21.8 microg/L (5.5-46.3), p=0.016] and "HB" with level [16,5 microg/L (2,3-30,8), p=0.021]compared to Control Group [9.7 microg/L (6,1-13,4)].Interestingly, 14 patients monitored after discharge from hospital show altered ECP values (X=24,6 microg/L) correlated with poor oxygen saturation. Conclusions: We believe that the serial determination of ECP levels is useful for monitoring most patients with COVID 19 but above all to analyze the patients in the follow up to report a possible dysfunction of the respiratory tract.

The prevalence of antibodies to SARS-CoV-2 in healthcare workers of Modena. the screening on more than 7000 subjects

Introduction:Healthcare workers (HCWs) are considered at high risk of SARS-CoV-2 infection, and during outbreak more than 30 thousand of them were infected. From April, in Emilia-Romagna Region, the screening for all HCWs has started through serological tests. In this retrospective study, we evaluated the prevalence of IgG and IgM antibodies (Abs) anti SARS-CoV-2 in HCWs affiliated to AUSL of Modena. Methods:We analysed data on asymptomatic HCWs, to assess the immunological status for the regional screening, that allows Abs to be detected every two weeks. In case of positivity to serological test, SARSCoV-2 RNA detection in nasopharyngeal/oropharingeal swab takes place by RT-PCR. Results: From 30 March to 31 May 2020, 7711 HCWs were screened using commercial immunocromatography and chemiluminescent tests to detect specific Abs IgG and IgM to SARS-CoV-2. 7361 (95.4%) were IgG and IgM negative while 350(5%) were IgG and/or IgM positive. In particular, 72 (0.9%) were IgG+ and IgM+, 233 (3%) were IgG+ and IgM-, 45 (0.6%) were IgG- A nd IgM+. At follow up, 4.6% of analyzed subjects were positive, of these 4.2% was IgG+ and only the 0.4% was IgM+. The RT-PCR performed on positive HCWs revealed that 7% of them was positive for viral RNA. Conclusions: Serological tests could bridge the gap between the disease and symptoms onset, allowing to detect both active and past infection when used as screening. Results of serological tests were influenced by several factors: In asymptomatic people it is difficult to establish the moment of contact, so it is plausible that the serological tests were negative because the body hasn't had the time to develop the antibodies. Conversely, detection of a very low percentage of IgM in conjunction with high IgG levels could be due to a remote contact with the virus, but also due to a scarce sensibility of tests. Furthermore, it is more likely that the asymptomatic people develop a little or no immune reaction because they didn't developed symptoms. Although the prevalence of SARS-CoV-2 infection is low among HCWs of Modena, Abs detection could be useful to detect subjects who developed an immune response, and to understand the epidemiology of the infection.