High Epstein-Barr virus capsid antigen IgG level associates with the carriership of CD8+ T cell somatic mutations in the STAT3 SH2 domain.

High carrier prevalence of STAT3 SH2 domain somatic mutations was recently discovered in CD8+ T cells. We found these low-allele-fraction clones in 26% of donors, without difference between multiple sclerosis (MS) patients and controls. Here we tested whether anti-viral antibodies associate with the carriership of these mutant clones. We compared antibody responses against common viruses in mutation carriers vs. non-carriers. Plasma samples of 152 donors (92 MS patients, 60 controls) were analyzed for antibodies against cytomegalovirus (CMV), Epstein-Barr virus (EBV), human herpesvirus-6A and parvovirus B19. The mutation carrier status associated with EBV VCA IgG level (p = 0.005) and remained significant after logistic regression (p = 0.036). This association was contributed similarly by MS patients and controls. These results suggest that EBV contributes to the generation or growth of these clones. The pathogenic role of the STAT3 mutant clones in MS is presently unclear, but their detailed characterization warrants further study.

A Combination of N and S Antigens With IgA and IgG Measurement Strengthens the Accuracy of SARS-CoV-2 Serodiagnostics.

BACKGROUND: Primary diagnosis of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is based on detection of virus RNA in nasopharyngeal swab samples. In addition, analysis of humoral immunity against SARS-CoV-2 has an important role in viral diagnostics and seroprevalence estimates. METHODS: We developed and optimized an enzyme immunoassays (EIA) using SARS-CoV-2 nucleoprotein (N), S1 and receptor binding domain (RBD) of the viral spike protein, and N proteins from SARS, Middle East respiratory syndrome (MERS), and 4 low-pathogenic human CoVs. Neutralizing antibody activity was compared with SARS-CoV-2 IgG, IgA, and IgM EIA results. RESULTS: The sensitivity of EIA for detecting immune response in COVID-19 patients (n = 101) was 77% in the acute phase and 100% in the convalescent phase of SARS-CoV-2 infection when N and RBD were used as antigens in IgG and IgA specific EIAs. SARS-CoV-2 infection significantly increased humoral immune responses against the 229E and NL63 N proteins. S1 and RBD-based EIA results had a strong correlation with microneutralization test results. CONCLUSIONS: The data indicate a combination of SARS-CoV-2 S1 or RBD and N proteins and analysis of IgG and IgA immunoglobulin classes in sera provide an excellent basis for specific and sensitive serological diagnostics of COVID-19.

SARS-CoV-2 infections among healthcare workers at Helsinki University Hospital, Finland, spring 2020: Serosurvey, symptoms and risk factors.

BACKGROUND: Exposure, risks and immunity of healthcare workers (HCWs), a vital resource during the SARS-CoV-2 pandemic, warrant special attention. METHODS: HCWs at Helsinki University Hospital, Finland, filled in questionnaires and provided serum samples for SARS-CoV-2-specific antibody screening by Euroimmun IgG assay in March-April 2020. Positive/equivocal findings were confirmed by Abbott and microneutralization tests. Positivity by two of the three assays or RT-PCR indicated a Covid-19 case (CoV+). RESULTS: The rate of CoV(+) was 3.3% (36/1095) and seropositivity 3.0% (33/1095). CoV(+) was associated with contact with a known Covid-19 case, and working on a Covid-19-dedicated ward or one with cases among staff. The rate in the Covid-19-dedicated ICU was negligible. Smoking and age <55 years were associated with decreased risk. CoV(+) was strongly associated with ageusia, anosmia, myalgia, fatigue, fever, and chest pressure. Seropositivity was recorded for 89.3% of those with prior documented RT-PCR-positivity and 2.4% of those RT-PCR-negative. The rate of previously unidentified cases was 0.7% (8/1067) and asymptomatic ones 0% (0/36). CONCLUSION: Undiagnosed and asymptomatic cases among HCWs proved rare. An increased risk was associated with Covid-19-dedicated wards. Particularly high rates were seen for wards with liberal HCW-HCW contacts, highlighting the importance of social distancing also among HCWs.

Performance of a multiplexed serological microarray for the detection of antibodies against central nervous system pathogens.

Central nervous system (CNS) infections have multiple potential causative agents for which simultaneous pathogen screening can provide a useful tool. This study evaluated a multiplexed microarray for the simultaneous detection of antibodies against CNS pathogens. The performance of selected microarray antigens for the detection of IgG antibodies against herpes simplex virus 1 and 2 (HSV-1 and HSV-2), varicella-zoster virus (VZV), adenovirus, Mycoplasma pneumoniae and Borrelia burgdorferi sensu lato, was evaluated using serum sample panels tested with reference assays used in a routine diagnostic laboratory. The microarray sensitivity for HSV-1, HSV-2, VZV, adenovirus and M. pneumonia ranged from 77% to 100%, and the specificity ranged from 74% to 97%. Very variable sensitivities and specificities were found for borrelial antigens of three different VlsE protein IR(6) peptide variants (IR6p1, IR6p2, IR6p4) and three recombinant decorin binding proteins A (DbpA; DbpAIa, DbpA91, DbpAG40). For single antigens, good specificity was shown for antigens of IR6p4 and DbpAIa (96%), while DbpA91, IR6p1 and IR6p2 were moderately specific (88-92%). The analytical sensitivity of the microarray was dependent on the borrelial IgG concentration of the specimen. The overall performance and technical features of the platform showed that the platform supports both recombinant proteins, whole viruses and peptides as antigens. This study showed diagnostic potential for all six CNS pathogens, including Borrelia burgdorferi sensu lato, using glutaraldehyde based microarray, and further highlighted the importance of careful antigen selection and the requirement for the use of multiple borrelial antigens in order to increase specificity without a major lack of sensitivity.

Surface-activated microtiter-plate microarray for simultaneous CRP quantification and viral antibody detection.

Microarrays are widely used in high-throughput DNA and RNA hybridization tests and recently adopted to protein and small molecule interaction studies in basic research and diagnostics. Parallel detection of serum antibodies and antigens has several potential applications in epidemiologic research, vaccine development, and in the diagnosis of allergies, autoimmunity, and infectious diseases. This study demonstrates an immobilization method for immunoassay-based microarray in conventional 96-well polystyrene plates for a serologic diagnostic method combined with quantitative C-reactive protein (CRP) assay. A synthetic peptide (HIV-1), a recombinant protein (Puumala hantavirus nucleocapsid), and purified virus preparations (Sindbis and adenoviruses) were used as antigens for virus-specific antibody detection and monoclonal anti-CRP antibody for antigen detection. The microarray was based on conventional enzyme immunoassays and densitometry from photographed results. Peptide and recombinant antigens functioned well, while whole virus antigens gave discrepant results in 1 out of 23 samples from the reference method, tested with human sera with various antibody responses. The CRP results were in concordance in the concentration range 0.5-150 mg/L with 2 commercially available CRP assays: ReaScan rapid test (R(2) = 0.9975) and Cobas 6000 analyzer (R(2) =0.9595). The results indicate that microtiter plates provide a promising platform for further development of microarrays for parallel antibody and antigen detection.