Autoantibodies in COVID-19, a possible role in the pathogenesis of the disease.

OBJECTIVES: In this study, we investigated whether SARS-CoV-2 stimulates autoantibody production. METHODS: The study included 91 patients hospitalized due to COVID 19, with no previous history of immunological diseases. Immunofluorescence assays were performed to detect antinuclear antibodies (ANAs) and antineutrophil cytoplasmic antibodies (ANCAs), along with tests for specific autoantibodies. RESULTS: The median age (57% male) was 74 years (range 38-95 years). Autoantibodies were positive in 67 (74%), ANA in 65 (71%), and ANCA in 11 (12%) patients. Female gender (p = 0.01), age (p = 0.005), and Charlson comorbidity index (p = 0.004) were significant predictors for the development of ANA/ANCA antibodies (p = 0.004). Nuclear mitotic apparatus (NuMA)-like, positivity was the strongest predictor of acute kidney injury (AKI), together with noninvasive ventilation and eGFR (χ2 = 49.01, P < 0.001). CONCLUSION: Positive autoantibodies in a large proportion of patients suggest a role of autoimmunity in the pathophysiology of acute COVID-19 disease. NuMA was the strongest predictor of AKI.

Non-Myelofibrosis Chronic Myeloproliferative Neoplasm Patients Show Better Seroconversion Rates after SARS-CoV-2 Vaccination Compared to Other Hematologic Diseases: A Multicentric Prospective Study of KroHem.

Disease- and treatment-mediated immunodeficiency might render SARS-CoV-2 vaccines less effective in patients with hematologic diseases. We performed a prospective non-interventional study to evaluate humoral response after one and two doses of mRNA-1273, BNT162b2, or ChAdOx1 nCoV-19 vaccine in 118 patients with different malignant or non-malignant hematologic diseases from three Croatian treatment centers. An electrochemiluminescent assay was used to measure total anti-SARS-CoV-2 S-RBD antibody titers. After one vaccine dose, 20/66 (33%) achieved seropositivity with a median antibody titer of 6.1 U/mL. The response rate (58/90, 64.4%) and median antibody titer (>250 U/mL) were higher after two doses. Seropositivity varied with diagnosis (overall p < 0.001), with the lowest rates in lymphoma (34.6%) and chronic lymphocytic leukemia (52.5%). The overall response rate in chronic myeloproliferative neoplasms (CMPN) was 81.3% but reached 100% in chronic myeloid leukemia and other non-myelofibrosis CMPN. At univariable analysis, age > 67 years, non-Hodgkin's lymphoma, active treatment, and anti-CD20 monoclonal antibody therapy increased the likelihood of no vaccine response, while hematopoietic stem cell recipients were more likely to respond. Age and anti-CD20 monoclonal antibody therapy remained associated with no response in a multivariable model. Patients with the hematologic disease have attenuated responses to SARS-CoV-2 vaccines, and significant variations in different disease subgroups warrant an individualized approach.

Unusual total anti-SARS-CoV-2 antibody kinetics observed during longitudinal monitoring after BNT162b2 vaccination.

The present study aimed to clarify unusual total antibody kinetics in three female individuals observed during longitudinal monitoring of antibody response to BNT162b2 COVID-19 vaccine in 54 healthy volunteers. Total and IgG antibodies against the SARS-CoV-2 spike glycoprotein were measured using Roche and Abbott quantitative assays, respectively, a day before and 8, 71, 135 and 217 days after the second dose. Samples showing unusual kinetics were additionally tested with Beckman Coulter and Euroimmun IgG assays, as well as IgA assay. Antibody levels peaked 8 days after the second dose (total:2769 U/mL; IgG:20022 AU/mL) and declined to 611 U/mL (total) and 783 AU/mL (IgG), after 217 days. A delayed increase of total but not IgG antibodies evidenced in three females, was in two cases coupled with an increase in IgA antibodies. This study identified a previously unknown contribution of anti-SARS-CoV-2 IgA antibodies to a delayed total antibody increase in a subgroup of vaccinated individuals. It also emphasizes that different commercially available serological assays do not provide uniform information about the post-vaccination immune status and that thorough understanding the assays' features is crucial for the proper interpretation of antibody response monitoring.

Association of polymorphisms in genes encoding prothrombotic and cardiovascular risk factors with disease severity in COVID-19 patients: A pilot study.

The present study aimed to assess the association of 16 polymorphisms in genes encoding prothrombotic and cardiovascular risk factors with COVID-19 disease severity: FV G1691A, FV H1299R, FII G20210A, MTHFR C677T, MTHFR A1298, factor XIII V34L, PAI-1 4G/5G, EPCR haplotypes (A1/A2/A3), eNOS -786 T > C, eNOS G894T, LTA C804A, ACE I/D, ITGB3 PIA1/A2, ITGA2B Baka/b, ß-Fbg -455 G > A and ApoB R3500Q. The study included 30 patients with severe COVID-19 and 49 non-severe COVID-19 patients. All studied polymorphisms except ITGA2B Baka/b were determined using multilocus genotyping assays CVD StripAssays (ViennaLab Diagnostics), while ITGA2B was genotyped using a real-time PCR method based on TaqMan technology. A higher frequency of carriers of at least one ITGB3 PIA2 allele was found in severe COVID-19 patients (p = 0.009). The distribution of genotypes was significantly different for ß-Fbg -455 G > A (p = 0.042), with only three homozygous AA genotypes found among severe COVID-19 patients. The association with an increased risk for severe COVID-19 was found for ITGB3, with carriers of at least one ITGB3 PIA2 allele having a 3.5-fold greater risk of severe COVID-19 (p = 0.011). Genotype distribution differences were obtained for the combinations of FV H1299R and FXIII V34L (p = 0.026), ITGB3 PIA1/A2 and ITGA2B Baka/b (p = 0.024), and ACE I/D and PAI-1 4G/5G (p = 0.046). ITGB3 polymorphism emerged as an independent risk factor for severe COVID-19 and homozygosity for ß-Fbg -455 G > A mutation could contribute to disease severity. The combined effect of polymorphisms in genes encoding prothrombotic and cardiovascular risk factors could further contribute to disease severity.

Humoral and cellular immunity in convalescent COVID-19 people with multiple sclerosis treated with ofatumumab.

OBJECTIVES: To report clinical outcome, development of humoral and T-cell mediated immunity in convalescent COVID-19 people with multiple sclerosis (pwMS) treated with ofatumumab in the ALITHIOS study from a single center. METHODS: Testing for SARS-Cov2 IgG antibodies was performed on two occasions with at least three months apart between the two testing. During the second antibody testing, interferon-γ ELISpot was used to assess cellular immunity. RESULTS: All four subjects had mild COVID-19 infection without any sequelae. In all subjects except subject 2, COVID-19 was confirmed with PCR. Subjects 1, 2 and 4 had normal levels of IgM and IgG without measurable counts of CD19 cells prior to COVID-19. Subject 3 administered the last dose of ofatumumab 24 days prior to COVID-19 symptoms, but had a gap of 28 weeks of ofatumumab application beforehand due to low IgM levels. Subject 4 received COVID-19 vaccinations before second testing, so second testing and T-cell immunity testing were not performed. Subjects who were CD19 depleted did not had measurable levels of SARS-Cov2 IgG antibodies. Subject 3 had first and second SARS-COV2 titer of 118 U/ml and > 250 U/ml, respectively. All three pwMS showed T cell immunity against SARS-CoV-2. Quotient of basal spots divided by interferon-γ secreting spot forming units were 4, 8 and 14.7 SI in subjects 1, 2 and 3, respectively (>3 considered reactive). CONCLUSION: While no antibody response was observed in pwMS who were CD19+ lymphocyte depleted, T cell immunity against SARS-CoV-2 was observed in all three pwMS treated with ofatumumab.

Humoral and cellular immunity in convalescent and vaccinated COVID-19 people with multiple sclerosis: Effects of disease modifying therapies.

OBJECTIVES: To determine anti-SARS-Cov2 antibodies and T-cell immunity in convalescent people with multiple sclerosis (pwMS) and/or pwMS vaccinated against Covid-19, depending on the disease modifying therapy, and in comparison to healthy controls (HC). METHODS: 75 participants were enrolled: Group 1-29 (38.7%) COVID-19 convalescent participants; Group 2-34 (45.3%) COVID-19 vaccinated; Group 3-12 (16.0%) COVID-19 convalescent participants who were later vaccinated against COVID-19. Cellular immunity was evaluated by determination of number of CD4+ and CD8+ cells secreting TNFα, IFNγ, and IL2 after stimulation with SARS-CoV-2 peptides. RESULTS: pwMS treated with ocrelizumab were less likely to develop humoral immunity after COVID-19 recovery or vaccination. No difference was observed in the cellular immunity in all studied parameters between pwMS treated with ocrelizumab compared to HC or pwMS who were treatment naïve or on first line therapies. These findings were consistent in convalescent, vaccinated, and convalescent+vaccinated participants. COVID-19 vaccinated convalescent pwMS on ocrelizumab compared to COVID-19 convalescent HC who were vaccinated did not show statistically difference in the rate of seroconversion nor titers of SARS-CoV-2 antibodies. CONCLUSION: Presence of cellular immunity in pwMS on B-cell depleting therapies is reassuring, as at least partial protection from more severe COVID-19 outcomes can be expected.

Humoral immune response to COVID-19 vaccines in people with secondary progressive multiple sclerosis treated with siponimod.

OBJECTIVE: The aim of this study is to determine a development of humoral response after COVID-19 vaccination in persons with secondary progressive multiple sclerosis (pwSPMS) on siponimod, compared to healthy controls (HC). METHODS: In 13 pwSPMS taking siponimod and 11 HC, testing for SARS-CoV2 antibodies was performed after vaccination against COVID-19. RESULTS: pwSPMS taking siponimod had a significantly lower titer of SARS-CoV2 antibodies compared to healthy controls (19.4 (0-250) vs. 250 (250), p>0.001). Two (15.4%) pwSPMS on siponimod had unmeasurable titers of SARS-CoV2-2 antibodies, while all HC had positive titers. CONCLUSION: Although the results of this study are limited by a small sample size, results have consistently shown low titers of SARS-CoV-2 IgG after COVID-19 vaccinations in pwSPMS on siponimod.

Antibody response and self-reported adverse reactions following vaccination with Comirnaty: a pilot study from a Croatian university hospital.

This study aimed to determine antibody responses against SARS-CoV-2 spike (S) after both BioNTech-Pfizer Comirnaty vaccine doses and study the correlation with self-perceived adverse reactions. Antibodies determination with Elecsys anti-SARS-CoV-2 S assay was performed a day prior to or just before administration of the second dose and 8-13 days after the second dose. Participants selected from a predefined list of the experienced local (injection site reactions) and/or systemic (fatigue, headache, myalgia, arthralgia, chills and fever) post-vaccination adverse reactions. An average 100-fold increase in antibody titre in naive vaccinees was observed between the two time points (median 67 U/mL vs 2841 U/mL, p<0.001). Participants aged below 50 had higher antibody titres (median 99 U/mL vs 26 U/mL, p=0.003 after the first dose; median 3617 U/mL vs 2556 U/mL, p=0.026 after the second dose). All reported adverse reactions were mild-to-moderate, with more participants declaring systemic reactions after the second dose (p=0.001), without a clear correlation with antibody titre.

Extracorporeal blood purification is associated with improvement in biochemical and clinical variables in the critically-ill COVID-19 patients

Abstract This study tried to investigate the impact of oXiris filter on both clinical and laboratory parameters in critically-ill COVID-19 intensive care unit (ICU) patients receiving extracorporeal blood purification and the clinical setting for the initiation of therapy. A consecutive sample of 15 ICU patients with COVID-19 was treated with oXiris membrane for blood purification or for support of renal function due to acute kidney injury. We have included 19 non treated ICU COVID-19 patients as a control group. Two chest x-rays were analyzed for determining the chest x-ray severity score. We have found a significant decrease of SOFA score, respiratory status improved and the chest x-ray severity score was significantly decreased after 72?h of treatment. IL-6 significantly decreased after 72?h of treatment while other inflammatory markers did not. Respiratory status in the control group worsened as well as increase in SOFA score and chest x-ray severity score. Survived patients have shorter time from the onset of symptoms before starting with extracorporeal blood purification treatment and shorter time on vasoactive therapy and invasive respiratory support than deceased patients. Critically-ill patients with COVID-19 treated with extracorporeal blood purification survived significantly longer than other ICU COVID-19 patients. Treatment with oXiris membrane provides significant reduction of IL-6, leads to improvement in respiratory status, chest x-ray severity score, and reduction of SOFA score severity. Our results can suggest that ICU COVID-19 patients in an early course of a disease could be potentially a target group for earlier initiation of extracorporeal blood purification.

Humoral immune response in convalescent COVID-19 people with multiple sclerosis treated with high-efficacy disease-modifying therapies: A multicenter, case-control study.

AIM: To determine the influence of high-efficacy disease modifying therapy (DMT) on the development of IgG SARS-CoV-2 antibody response in COVID-19 convalescent people with multiple sclerosis (pwMS). METHODS: Seventy-four pwMS taking high-efficacy DMTs (specifically natalizumab, fingolimod, alemtuzumab, ocrelizumab, cladribine and ublituximab) and diagnosed with COVID-19 and 44 healthy persons (HC) were enrolled. SARS-CoV2 antibodies were tested with Elecsys® Anti-SARSCoV-2 S assay. RESULTS: pwMS taking high-efficacy DMTs had a significantly higher chance of having negative titer of SARS-CoV2 antibodies compared to healthy controls (33 negative pwMS [44.6%] compared to one negative HC [2.3%], p < 0.001). pwMS taking B-cell depleting therapy (ocrelizumab and ublituximab) had a significantly higher chance of having negative titer of SARS-CoV2 antibodies compared to pwMS on all other DMTs (29 negative pwMS on B-cell therapy [64.4%] compared to four negative pwMS on all other DMTs [13.8%], p < 0.001). Out of other DMTs, two (33.3%) pwMS taking fingolimod and two (16.7%) pwMS taking cladribine failed to develop IgG SARS-COV-2 antibodies. B-cell depleting therapy independently predicted negative titer of IgG SARS-CoV-2 antibody (Exp[B] =0.014, 95%CI 0.002-0.110, p < 0.001). CONCLUSIONS: A significant proportion of convalescent COVID-19 pwMS on high-efficacy DMTs will not develop IgG SARS-CoV-2 antibodies. B-cell depleting therapies independently predict negative and low titer of IgG SARS-CoV-2 antibody.

Laboratory professionals' attitudes towards ISO 15189:2012 accreditation: an anonymous survey of three Croatian accredited medical laboratories.

INTRODUCTION: Effective implementation and continual compliance with ISO 15189:2012 require ongoing commitment and active involvement of laboratory staff. Our aim was to assess attitudes regarding accreditation implementation by conducting a survey in three Croatian accredited medical laboratories. MATERIALS AND METHODS: An anonymous survey consisting of 34 questions was distributed either electronically or in a paper form a week prior to scheduled annual audits. Distributions of answers regarding age, work experience, laboratory workplace, and education level and according to the respective laboratory were compared. RESULTS: The overall response rate was 76% (225/297). Preference towards working in an accredited laboratory and a positive attitude were revealed by 70% and 56% participants, respectively, with better process documentation as the main advantage. Only 14% of responders considered themselves completely familiar with ISO 15189:2012. Total of 68% of responders felt that accreditation increases the usual workload, with excessive paperwork as the main contributor. Half of the responders declared partial agreement that accreditation requirements and expectations were clearly explained and claimed that their suggestions were taken into account only occasionally, which was especially emphasized by technical staff. The vast majority (89%) completely follow the prescribed protocols. Only 27% consider turnaround time monitoring useful. Competence assessment is considered efficient by 41% of responders. The majority (73%) prefer an online audit in times of COVID-19. CONCLUSIONS: Despite an overall positive attitude towards accreditation, further efforts are needed in providing better education about ISO 15189:2012 for technical staff and modifying formats of competence assessment, in order to achieve better adherence to ISO 15189:2012 requirements.

COVID-19 Pandemic Once Again Exposes the Weakest Link in Laboratory Services: Specimen Delivery.

OBJECTIVE: Reorganization of the emergency department (ED) during the COVID-19 pandemic implied closure of the ED-dedicated laboratory and manual transport of all specimens to the dislocated central laboratory. The impact of such reorganization on laboratory turnaround time (TAT) was examined. METHODS: The TAT from blood sampling to specimen reception (TAT1), from specimen reception to test reporting (TAT2), and from sampling to test reporting (TAT3) were compared between the pandemic peak month in 2020 and the same month in 2019. We evaluated whether TAT2 fulfills the recommended 60-minute criteria. RESULTS: A statistically significant difference was observed for all comparisons (P <.001), with TAT1 prominently contributing to TAT3 prolongation (from 48 minutes to 108 minutes) and exceeding the recommended 60-minute criteria. The TAT2 was extended from 33 minutes to 49 minutes. CONCLUSION: An ED reorganization compromised the usual laboratory services for patients in the ED, with manual specimen delivery being the main cause for TAT prolongation.