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Objectives Patients with Systemic Lupus Erythematosus are known to have dysregulated immune responses and may have reduced response to vaccination against COVID-19 while being at risk of severe COVID-19 disease. The aim of this study was to identify whether vaccine responses were attenuated in SLE and to assess disease- and treatment-specific associations. Methods Patients with SLE were matched by age, sex and ethnic background to healthcare worker healthy controls (HC). Anti-SARS-CoV-2 spike glycoprotein antibodies were measured at 4–8 weeks following the second COVID-19 vaccine dose (either BNT162b2 or ChAdOx1 nCoV-19) using a CE-marked combined ELISA detecting IgG, IgA and IgM (IgGAM). Antibody levels were considered as a continuous variable and in tertiles and compared between SLE patients and HC and associations with medication, disease activity and serological parameters were determined. Results Antibody levels were lower in 43 SLE patients compared to 40 HC ( p < 0.001). There was no association between antibody levels and medication, lupus disease activity, vaccine type or prior COVID infection. Higher serum IgA, but not IgG or IgM, was associated with being in a higher anti-SARS-CoV-2 antibody level tertile (OR [95% CI] 1.820 [1.050, 3.156] p = 0.033). Similarly, higher lymphocyte count was also associated with being in a higher tertile of anti-SARS-CoV-2 (OR 3.330 [1.505, 7.366] p = 0.003) Conclusion Patients with SLE have lower antibody levels following 2 doses of COVID-19 vaccines compared to HC. In SLE lower lymphocyte counts and serum IgA levels are associated with lower antibody levels post vaccination, potentially identifying a subgroup of patients who may therefore be at increased risk of infection.

In this population-based cohort of 7538 adults, combined immunoglobulin (Ig) G, IgA, and IgM (IgG/A/M) anti-spike titers measured after severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccination were predictive of protection against breakthrough SARS-CoV-2 infection. Discrimination was significantly improved by adjustment for factors influencing risk of SARS-CoV-2 exposure, including household overcrowding, public transport use, and visits to indoor public places. Anti-spike IgG/A/M titers showed positive correlation with neutralizing antibody titers (rs = 0.80 [95% confidence interval, .72–.86]; P < .001) and S peptide–stimulated interferon-γ concentrations (rs = 0.31 [.13–.47]; P < .001).

Purpose To define the burden of morbidity and mortality arising from COVID-19 in individuals with primary (PID) and secondary immunodeficiency (SID) in the United Kingdom. Methods In March 2020, the United Kingdom Primary Immunodeficiency Network (UKPIN) established a registry of cases to collate the outcomes of individuals with PID and SID following SARS-CoV-2 infection and treatment. Anonymised demographic data, pre-SARS-CoV-2 infection lymphocyte counts, co-morbidities, targeted treatments and outcomes were collected. Three groups were analysed in further detail: individuals with common variable immunodeficiency (CVID), individuals with any PID, including CVID, receiving immunoglobulin replacement therapy (IgRT) and individuals with secondary immunodeficiency. Results A total of 310 cases of SARS-CoV-2 infection in individuals with PID or SID have now been reported in the UK. The overall mortality within the cohort was 17.7% (n = 55/310). Individuals with CVID demonstrated an infection fatality rate (IFR) of 18.3% (n = 17/93), individuals with PID receiving IgRT had an IFR of 16.3% (n = 26/159) and individuals with SID, an IFR of 27.2% (n = 25/92). Individuals with PID and SID, had higher inpatient mortality and died at a younger age than the general population. Increasing age, low pre-SARS-CoV-2 infection lymphocyte count and the presence of common co-morbidities increased the risk of mortality in PID. Access to specific COVID-19 treatments in this cohort was limited: only 22.9% (n = 33/144) of patients admitted to hospital received dexamethasone, remdesivir, an anti-SARS-CoV-2 antibody-based therapeutic (e.g. REGN-COV2 or convalescent plasma) or tocilizumab as a monotherapy or in combination. Dexamethasone, remdesivir and anti-SARS-CoV-2 antibody-based therapeutics appeared efficacious in PID and SID. Conclusion Compared to the general population, individuals with PID or SID are at high risk of mortality following SARS-CoV-2 infection. Increasing age, low baseline lymphocyte count and the presence of co-morbidities are additional risk factors for poor outcome in this cohort.

Aims Thrombotic complications and vasculopathy have been extensively associated with severe COVID-19 infection; however, the mechanisms inducing endotheliitis and the disruption of endothelial integrity in the microcirculation are poorly understood. We hypothesized that within the vessel wall, pericytes preferentially take up viral particles and mediate the subsequent loss of vascular integrity. Methods and results Immunofluorescence of post-mortem patient sections was used to assess pathophysiological aspects of COVID-19 infection. The effects of COVID-19 on the microvasculature were assessed using a vascular organoid model exposed to live viral particles or recombinant viral antigens. We find increased expression of the viral entry receptor angiotensin-converting enzyme 2 on pericytes when compared to vascular endothelium and a reduction in the expression of the junctional protein CD144, as well as increased cell death, upon treatment with both live virus and/or viral antigens. We observe a dysregulation of genes implicated in vascular permeability, including Notch receptor 3, angiopoietin-2, and TEK. Activation of vascular organoids with interleukin-1β did not have an additive effect on vascular permeability. Spike antigen was detected in some patients’ lung pericytes, which was associated with a decrease in CD144 expression and increased platelet recruitment and von Willebrand factor (VWF) deposition in the capillaries of these patients, with thrombi in large vessels rich in VWF and fibrin. Conclusion Together, our data indicate that direct viral exposure to the microvasculature modelled by organoid infection and viral antigen treatment results in pericyte infection, detachment, damage, and cell death, disrupting pericyte-endothelial cell crosstalk and increasing microvascular endothelial permeability, which can promote thrombotic and bleeding complications in the microcirculation.

B-cell-depleting agents are among the most commonly used drugs to treat haemato-oncological and autoimmune diseases. They rapidly induce a state of peripheral B-cell aplasia with the potential to interfere with nascent vaccine responses, particularly to novel antigens. We have examined the relationship between B-cell reconstitution and SARS-CoV-2 vaccine responses in two cohorts of patients previously exposed to B-cell-depleting agents: a cohort of patients treated for haematological B-cell malignancy and another treated for rheumatological disease. B-cell depletion severely impairs vaccine responsiveness in the first 6 months after administration: SARS-CoV-2 antibody seroprevalence was 42.2% and 33.3% in the haemato-oncological patients and rheumatology patients, respectively and 22.7% in patients vaccinated while actively receiving anti-lymphoma chemotherapy. After the first 6 months, vaccine responsiveness significantly improved during early B-cell reconstitution; however, the kinetics of reconstitution was significantly faster in haemato-oncology patients. The AstraZeneca ChAdOx1 nCoV-19 vaccine and the Pfizer BioNTech 162b vaccine induced equivalent vaccine responses; however, shorter intervals between vaccine doses (36 m previously), vaccine non-responsiveness was independent of peripheral B-cell reconstitution. The findings have important implications for primary vaccination and booster vaccination strategies in individuals clinically vulnerable to SARS-CoV-2., B cell depleting drugs including rituximab are commonly used to treat haematological malignancy and autoimmune diseases but may impair the immunological response to vaccination. This study investigates SARS-CoV-2 vaccine responses in individuals with haematological and rheumatological disease with previously exposure to B cell depleting agents. Vaccination within the first 6 months of B cell depletion is associated with significant impairment of vaccine responsiveness; however, rheumatology and haemato-oncological patients display different kinetics of B cell reconstitution corresponding to differential vaccine responsiveness over time., Graphical Abstract Graphical Abstract

Background Lung cancer is the leading cause of cancer-related death worldwide. Surgical resection remains the definitive curative treatment for early-stage disease offering an overall 5-year survival rate of 62%. Despite careful case selection, a significant proportion of early-stage cancers relapse aggressively within the first year post-operatively. Identification of these patients is key to accurate prognostication and understanding the biology that drives early relapse might open up potential novel adjuvant therapies. Methods We performed an unsupervised interrogation of >1600 serum-based autoantibody biomarkers using an iterative machine-learning algorithm. Results We identified a 13 biomarker signature that was highly predictive for survivorship in post-operative early-stage lung cancer; this outperforms currently used autoantibody biomarkers in solid cancers. Our results demonstrate significantly poor survivorship in high expressers of this biomarker signature with an overall 5-year survival rate of 7.6%. Conclusions We anticipate that the data will lead to the development of an off-the-shelf prognostic panel and further that the oncogenic relevance of the proteins recognised in the panel may be a starting point for a new adjuvant therapy.

ObjectiveTo determine the effect of population level implementation of a test-and-treat approach to correction of suboptimal vitamin D status (25-hydroxyvitamin D (25(OH)D) DesignPhase 3 open label randomised controlled trial.SettingUnited Kingdom.Participants6200 people aged ≥16 years who were not taking vitamin D supplements at baseline.InterventionsOffer of a postal finger prick test of blood 25(OH)D concentration with provision of a six month supply of lower dose vitamin D (800 IU/day, n=1550) or higher dose vitamin D (3200 IU/day, n=1550) to those with blood 25(OH)D concentration Main outcome measuresThe primary outcome was the proportion of participants with at least one swab test or doctor confirmed acute respiratory tract infection of any cause. A secondary outcome was the proportion of participants with swab test confirmed covid-19. Logistic regression was used to calculate odds ratios and associated 95% confidence intervals. The primary analysis was conducted by intention to treat.ResultsOf 3100 participants offered a vitamin D test, 2958 (95.4%) accepted and 2674 (86.3%) had 25(OH)D concentrations ConclusionsAmong people aged 16 years and older with a high baseline prevalence of suboptimal vitamin D status, implementation of a population level test-and-treat approach to vitamin D supplementation was not associated with a reduction in risk of all cause acute respiratory tract infection or covid-19.Trial registrationClinicalTrials.govNCT04579640.

Background: In several countries, extended interval COVID-19 vaccination regimens are now used to accelerate population coverage, but the relative immunogenicity of different vaccines in older people remains uncertain. In this study we aimed to assess the antibody and cellular responses of older people after a single dose of either the BNT162b2 vaccine (tozinameran; Pfizer-BioNTech) or ChAdOx1 nCoV-19 vaccine (Oxford University-AstraZeneca). Methods: Participants aged 80 years or older, who did not live in a residential or care home or require assisted living, and had received a single dose of either the BNT162b2 vaccine or ChAdOx1 nCoV-19 vaccine were eligible to participate. Participants were recruited through local primary care networks in the West Midlands, UK. Blood samples and dried blood spots were taken 5-6 weeks after vaccination to assess adaptive immune responses using Elecsys electrochemiluminescence immunoassay and cellular responses by ELISpot. Primary endpoints were percentage response and quantification of adaptive immunity. Findings: Between Dec 29, 2020, and Feb 28, 2021, 165 participants were recruited and included in the analysis. 76 participants had received BNT162b2 (median age 84 years, IQR 82-89; range 80-98) and 89 had received ChAdOx1 nCoV-19 (median age 84 years, 81-87; 80-99). Antibody responses against the spike protein were detectable in 69 (93%) of 74 BNT162b2 vaccine recipients and 77 (87%) of 89 ChAdOx1 nCoV-19 vaccine recipients. Median antibody titres were of 19·3 U/mL (7·4-79·4) in the BNT162b2 vaccine recipients and 19·6 U/mL (6·1-60·0) in the ChAdOx1 nCoV-19 vaccine recipients (p=0·41). Spike protein-specific T-cell responses were observed in nine (12%) of 73 BNT162b2 vaccine recipients and 27 (31%) of 88 ChAdOx1 nCoV-19 vaccine recipients, and median responses were three-times higher in ChAdOx1 nCoV-19 vaccine recipients (24 spots per 1 × 106 peripheral blood mononuclear cells) than BNT162b2 vaccine recipients (eight spots per 1 × 106 peripheral blood mononuclear cells; p

Post-acute cardiac sequelae, following SARS-CoV-2 infection, are well recognized as complications of COVID-19. We have previously shown the persistence of autoantibodies against antigens in skin, muscle, and heart in individuals following severe COVID-19; the most common staining on skin tissue displayed an inter-cellular cement pattern consistent with antibodies against desmosomal proteins. Desmosomes play a critical role in maintaining the structural integrity of tissues. For this reason, we analyzed desmosomal protein levels and the presence of anti-desmoglein (DSG) 1, 2, and 3 antibodies in acute and convalescent sera from patients with COVID-19 of differing clinical severity. We find increased levels of DSG2 protein in sera from acute COVID-19 patients. Furthermore, we find that DSG2 autoantibody levels are increased significantly in convalescent sera following severe COVID-19 but not in hospitalized patients recovering from influenza infection or healthy controls. Levels of autoantibody in sera from patients with severe COVID-19 were comparable to levels in patients with non-COVID-19-associated cardiac disease, potentially identifying DSG2 autoantibodies as a novel biomarker for cardiac damage. To determine if there was any association between severe COVID-19 and DSG2, we stained post-mortem cardiac tissue from patients who died from COVID-19 infection. This confirmed DSG2 protein within the intercalated discs and disruption of the intercalated disc between cardiomyocytes in patients who died from COVID-19. Our results reveal the potential for DSG2 protein and autoimmunity to DSG2 to contribute to unexpected pathologies associated with COVID-19 infection.

SUMMARYBackground & AimsVitamin D deficiency has been reported to associate with impaired development of antigen-specific responses following vaccination. We aimed to determine whether vitamin D supplements might boost immunogenicity and efficacy of SARS-CoV-2 vaccination.MethodsWe conducted three sub-studies nested within the CORONAVIT randomised controlled trial, which investigated effects of offering vitamin D supplements at a dose of 800 IU/day or 3200 IU/day vs. no offer on risk of acute respiratory infections, including COVID-19, in UK adults with circulating 25-hydroxyvitamin D concentrations Results1945/2808 (69.3%) sub-study 1 participants received two doses of ChAdOx1 nCoV-19 (Oxford–AstraZeneca); the remainder received two doses of BNT162b2 (Pfizer). Vitamin D supplementation did not influence risk of breakthrough SARS-CoV-2 infection (800 IU/day vs. no offer: adjusted hazard ratio 1.28, 95% CI 0.89 to 1.84; 3200 IU/day vs. no offer: 1.17, 0.81 to 1.70). Neither did it influence IgGAM anti-Spike titres, neutralising antibody titres or IFN-γ concentrations in supernatants of S peptide-stimulated whole blood.ConclusionsAmong adults with sub-optimal baseline vitamin D status, vitamin D replacement at a dose of 800 or 3200 IU/day did not influence protective efficacy or immunogenicity of SARS-CoV-2 vaccination.Clinical Trial RegistrationClinicalTrials.govNCT04579640.

BACKGROUND: Vaccination prevents severe morbidity and mortality from COVID-19 in the general population. The immunogenicity and efficacy of SARS-CoV-2 vaccines in patients with antibody deficiency is poorly understood. OBJECTIVES: COVID-19 in patients with antibody deficiency (COV-AD) is a multi-site UK study that aims to determine the immune response to SARS-CoV-2 infection and vaccination in patients with primary or secondary antibody deficiency, a population that suffers from severe and recurrent infection and does not respond well to vaccination. METHODS: Individuals on immunoglobulin replacement therapy or with an IgG less than 4 g/L receiving antibiotic prophylaxis were recruited from April 2021. Serological and cellular responses were determined using ELISA, live-virus neutralisation and interferon gamma release assays. SARS-CoV-2 infection and clearance were determined by PCR from serial nasopharyngeal swabs. RESULTS: A total of 5.6% (n = 320) of the cohort reported prior SARS-CoV-2 infection, but only 0.3% remained PCR positive on study entry. Seropositivity, following two doses of SARS-CoV-2 vaccination, was 54.8% (n = 168) compared with 100% of healthy controls (n = 205). The magnitude of the antibody response and its neutralising capacity were both significantly reduced compared to controls. Participants vaccinated with the Pfizer/BioNTech vaccine were more likely to be seropositive (65.7% vs. 48.0%, p = 0.03) and have higher antibody levels compared with the AstraZeneca vaccine (IgGAM ratio 3.73 vs. 2.39, p = 0.0003). T cell responses post vaccination was demonstrable in 46.2% of participants and were associated with better antibody responses but there was no difference between the two vaccines. Eleven vaccine-breakthrough infections have occurred to date, 10 of them in recipients of the AstraZeneca vaccine. CONCLUSION: SARS-CoV-2 vaccines demonstrate reduced immunogenicity in patients with antibody deficiency with evidence of vaccine breakthrough infection.

Dental care professionals (DCPs) are thought to be at enhanced risk of occupational exposure to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). However, robust data to support this from large-scale seroepidemiological studies are lacking. We report a longitudinal seroprevalence analysis of antibodies to SARS-CoV-2 spike glycoprotein, with baseline sampling prior to large-scale practice reopening in July 2020 and follow-up postimplementation of new public health guidance on infection prevention control (IPC) and enhanced personal protective equipment (PPE). In total, 1,507 West Midlands DCPs were recruited into this study in June 2020. Baseline seroprevalence was determined using a combined IgGAM enzyme-linked immunosorbent assay and the cohort followed longitudinally for 6 mo until January/February 2021 through the second wave of the coronavirus disease 2019 pandemic in the United Kingdom and vaccination commencement. Baseline seroprevalence was 16.3%, compared to estimates in the regional population of 6% to 7%. Seropositivity was retained in over 70% of participants at 3- and 6-mo follow-up and conferred a 75% reduced risk of infection. Nonwhite ethnicity and living in areas of greater deprivation were associated with increased baseline seroprevalence. During follow-up, no polymerase chain reaction–proven infections occurred in individuals with a baseline anti–SARS-CoV-2 IgG level greater than 147.6 IU/ml with respect to the World Health Organization international standard 20-136. After vaccination, antibody responses were more rapid and of higher magnitude in those individuals who were seropositive at baseline. Natural infection with SARS-CoV-2 prior to enhanced PPE was significantly higher in DCPs than the regional population. Natural infection leads to a serological response that remains detectable in over 70% of individuals 6 mo after initial sampling and 9 mo from the peak of the first wave of the pandemic. This response is associated with protection from future infection. Even if serological responses wane, a single dose of the Pfizer-BioNTech 162b vaccine is associated with an antibody response indicative of immunological memory.

Summary Coronavirus 19 (COVID‐19) has been associated with both transient and persistent systemic symptoms that do not appear to be a direct consequence of viral infection. The generation of autoantibodies has been proposed as a mechanism to explain these symptoms. To understand the prevalence of autoantibodies associated with severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) infection, we investigated the frequency and specificity of clinically relevant autoantibodies in 84 individuals previously infected with SARS‐CoV‐2, suffering from COVID‐19 of varying severity in both the acute and convalescent setting. These were compared with results from 32 individuals who were on the intensive therapy unit (ITU) for non‐COVID reasons. We demonstrate a higher frequency of autoantibodies in the COVID‐19 ITU group compared with non‐COVID‐19 ITU disease control patients and that autoantibodies were also found in the serum 3–5 months post‐COVID‐19 infection. Non‐COVID patients displayed a diverse pattern of autoantibodies; in contrast, the COVID‐19 groups had a more restricted panel of autoantibodies including skin, skeletal muscle and cardiac antibodies. Our results demonstrate that respiratory viral infection with SARS‐CoV‐2 is associated with the detection of a limited profile of tissue‐specific autoantibodies, detectable using routine clinical immunology assays. Further studies are required to determine whether these autoantibodies are specific to SARS‐CoV‐2 or a phenomenon arising from severe viral infections and to determine the clinical significance of these autoantibodies., Acute infection with COVID is associated with a pattern of autoantibodies including a high proportion with epidermal antibodies. This autoantibody pattern is more common in severe COVID and is persistent up to 6 months.

Background: Post-operative recurrence in lung cancer continues to pose a major clinical challenge. B cell and plasma cell presence has been shown to correlate with improved survival and lower relapse rates in solid cancers and with improved response to checkpoint blockade. The aim of this study was to identify distinct immune cell compositions in the circulation and tumor microenvironment and make correlations with clinical and disease specific outcomes. Methods: In this study, utilizing high-dimensional deep immunophenotyping techniques (mass cytometry), multispectral immunofluorescence based imaging and bulk RNA sequencing, we explored the B cell repertoire in the circulation and tumor microenvironment in early-stage lung cancer patients undergoing radical surgical resection. Results: We demonstrate using high-dimensional deep phenotyping, clear differences in B cell composition between the circulation and the intratumoural microenvironment. Intratumoural predominant populations include plasma cells, from CD38var immature to CD138+ mature cells. B cells undergoing germinal center and follicular maturation as well as class-switched memory cells were predominantly found in the circulation. Eight key populations were significantly differentially abundant between compartments on generalized linear mixed modelling (p Conclusions: Our data has demonstrated the unique roles of effector and suppressive B cell subsets in NSCLC and the clear difference in surrogacy between tissue compartments. The lesser infiltration of suppressive phenotypes into the tumor nest may be to prevent the subsequent infiltration of effector plasma and effector B cells into the tumor nest. This tumor line of defense would be in keeping with the observation that higher levels of suppressive B cell subsets in the circulation links to higher rates of recurrence and a higher tumor effector plasma cell infiltrate correlates with protection against recurrent disease. The ambiguous role of B cells in the tumor environment predetermines the multidirectional development of immunotherapeutic approaches, either supporting positive B cell types or inhibiting the suppressive B cell phenotypes. Citation Format: Akshay J. Patel, Naeem Khan, Alex G. Richter, Babu Naidu, Mark T. Drayson, Gary W. Middleton. Plasma cell repertoire in the lung cancer tumor microenvironment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5943.

Background Patients with chronic kidney disease (CKD) are more prone to severe infection. Vaccination is a key strategy to reduce this risk. Some studies suggest vaccine efficacy may be reduced in patients with CKD, despite preserved maintenance of long-term responses to some pathogens and vaccines. Here, we investigated immune responses to 2 vaccines in patients with CKD to identify predictors of immunological responsiveness. Methods Individuals >65 years old, with or without nondialysis CKD (n = 36 and 29, respectively), were vaccinated with a nonadjuvanted seasonal influenza vaccine (T-dependent) and Pneumovax23 (23-valent pneumococcal polysaccharide [PPV23], T-independent). Humoral responses were measured at baseline, day 28, and 6 months. Lymphocyte subset and plasma cell/blast analyses were performed using flow cytometry. Cytomegalovirus (CMV) serotyping was assessed by enzyme-linked immunosorbent assay. Results Only modest responsiveness was observed to both vaccines, independent of CKD status (25% adequate response in controls vs. 12%–18% in the CKD group). Unexpectedly, previous immunization with PPV23 (median 10-year interval) and CMV seropositivity were associated with poor PPV23 responsiveness in both study groups (P < .001 and .003, respectively; multivariable linear regression model). Patients with CKD displayed expanded circulating populations of T helper 2 and regulatory T cells, which were unrelated to vaccine responses. Despite fewer circulating B cells, patients with CKD were able to mount a similar day 7 plasma cell/blast response to controls. Conclusion Patients with nondialysis CKD can respond similarly to vaccines as age- and sex-matched healthy individuals. CKD patients display an immune signature that is independent of vaccine responsiveness. Prior PPV23 immunization and CMV infection may influence responsiveness to vaccination. Clinical Trials Registration. NCT02535052, Patients with nondialysis Chronic Kidney Disease (CKD) can mount similar responses to vaccination as controls. Subtle changes in lymphocyte phenotype are seen in CKD when controlled for effect of Cytomegalovirus (CMV). Latent CMV infection and previous PPV23 vaccination are associated with poorer vaccine responses.

BACKGROUND AND AIMS A variety of auto-antibody assays are available as part of the clinical care of patients with liver disease. We sought to better understand the clinical utility of immune serological testing in patients with primary biliary cholangitis (PBC). METHODS We retrospectively analysed data from 2846 patients investigated for liver disease at a UK liver centre between 2001 and 2017. A total of 499 patients with PBC were identified. Immune serology results were examined for their diagnostic utility and prognostic significance to predict transplant-free survival. RESULTS Antimitochondrial antibodies (AMAs) were specific (94.5%) and sensitive (85.6%) for PBC; antinuclear antibodies (ANAs) against glycoprotein 210 (gp210) and sp100 were specific (>98%) but not sensitive (

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infections have a devastating effect on patients receiving hemodialysis. To what extent infection-induced antibody responses are maintained, or protective, is unknown. This study describes the evolution of antibodies against the SARS-CoV-2 spike protein in a cohort of 990 patients on hemodialysis. During the first wave of the pandemic, 26% of patients had developed antispike SARS-CoV-2 antibodies. Fewer PCR-confirmed second-wave infections were observed in patients with pre-existing antibodies (4.2%) than those without antibodies (11.4%). This study shows that SARS-CoV-2 antibodies in patients on hemodialysis are well maintained and associate with reduced risk of subsequent SARS-CoV-2 infection.

Background The role of tracheostomy in coronavirus disease 2019 (COVID-19) is unclear, with several consensus guidelines advising against this practice. We developed both a dedicated airway team and coordinated education programme to facilitate ward management of tracheostomised COVID-19 patients. Here, we report outcomes in the first 100 COVID-19 patients who underwent tracheostomy at our institution. Methods This was a prospective observational cohort study of patients confirmed to have COVID-19 who required mechanical ventilation at Queen Elizabeth Hospital, Birmingham, UK. The primary outcome measure was 30-day survival, accounting for severe organ dysfunction (Acute Physiology and Chronic Health [APACHE]-II score>17). Secondary outcomes included duration of ventilation, ICU stay, and healthcare workers directly involved in tracheostomy care acquiring COVID-19. Results A total of 164 patients with COVID-19 were admitted to the ICU between March 9, 2020 and April 21, 2020. A total of 100 patients (mean [standard deviation] age: 55 [12] yr; 29% female) underwent tracheostomy; 64 (age: 57 [14] yr; 25% female) did not undergo tracheostomy. Despite similar APACHE-II scores, 30-day survival was higher in 85/100 (85%) patients after tracheostomy, compared with 27/64 (42%) non-tracheostomised patients {relative risk: 3.9 (95% confidence intervals [CI]: 2.3–6.4); P

Dried blood spot (DBS) samples can be used for the detection of severe acute respiratory syndrome coronavirus 2 spike antibodies. DBS sampling is comparable to matched serum samples with a relative 98.1% sensitivity and 100% specificity. Thus, DBS sampling offers an alternative for population-wide serologic testing in the coronavirus pandemic.

Multiple myeloma (MM) is associated with increased risk of infection, but little is known regarding antibody levels against specific bacteria. We assessed levels of polyclonal immunoglobulin and antibacterial antibodies in patients recruited to the TEAMM trial, a randomised trial of antibiotic prophylaxis at the start of anti-myeloma treatment. Polyclonal IgG, IgA and IgM levels were below the reference range in 71%, 83% and 90% of 838 MM patients at diagnosis. Anti-vaccine targeted tetanus toxoid antibodies were protective in 95% of 193 healthy controls but only 41% of myeloma patients. In healthy controls, protective antibodies against 6/12 pneumococcal serotypes, haemophilus and meningococcus A were present in 67%, 41% and 56% compared to just 15%, 21% and 17% of myeloma patients. By 1 year, myeloma patients IgG levels had recovered for 57% of patients whilst the proportion with protective levels of IgG against thymus-dependent protein antigen tetanus toxoid had changed little. In contrast the proportions of patients with protective levels against thymus independent polysaccharide antigens pneumococcus, haemophilus and meningococcus had fallen from 15 to 7%, 21 to 0% and 17 to 11%. Findings highlight the need for strategies to protect patients against bacterial infections during therapy and vaccination programmes during remission.

ObjectiveTo determine the rates of asymptomatic viral carriage and seroprevalence of SARS-CoV-2 antibodies in healthcare workers.DesignA cross-sectional study of asymptomatic healthcare workers undertaken on 24/25 April 2020.SettingUniversity Hospitals Birmingham NHS Foundation Trust (UHBFT), UK.Participants545 asymptomatic healthcare workers were recruited while at work. Participants were invited to participate via the UHBFT social media. Exclusion criteria included current symptoms consistent with COVID-19. No potential participants were excluded.InterventionParticipants volunteered a nasopharyngeal swab and a venous blood sample that were tested for SARS-CoV-2 RNA and anti-SARS-CoV-2 spike glycoprotein antibodies, respectively. Results were interpreted in the context of prior illnesses and the hospital departments in which participants worked.Main outcome measureProportion of participants demonstrating infection and positive SARS-CoV-2 serology.ResultsThe point prevalence of SARS-CoV-2 viral carriage was 2.4% (n=13/545). The overall seroprevalence of SARS-CoV-2 antibodies was 24.4% (n=126/516). Participants who reported prior symptomatic illness had higher seroprevalence (37.5% vs 17.1%, χ2=21.1034, pConclusions and relevanceWe identify differences in the occupational risk of exposure to SARS-CoV-2 between hospital departments and confirm asymptomatic seroconversion occurs in healthcare workers. Further investigation of these observations is required to inform future infection control and occupational health practices.

Children were relatively spared during COVID-19 pandemic. However, the recently reported hyperinflammatory syndrome with overlapping features of Kawasaki disease and toxic shock syndrome-"Paediatric Inflammatory Multisystem Syndrome-temporally associated with SARS-CoV-2" (PIMS-TS) has caused concern. We describe cardiac findings and short-term outcomes in children with PIMS-TS at a tertiary children's hospital. Single-center observational study of children with PIMS-TS from 10th April to 9th May 2020. Data on ECG and echocardiogram were retrospectively analyzed along with demographics, clinical features and blood parameters. Fifteen children with median age of 8.8 (IQR 6.4-11.2) years were included, all were from African/Afro-Caribbean, South Asian, Mixed or other minority ethnic groups. All showed raised inflammatory/cardiac markers (CRP, ferritin, Troponin I, CK and pro-BNP). Transient valve regurgitation was present in 10 patients (67%). Left Ventricular ejection fraction was reduced in 12 (80%), fractional shortening in 8 (53%) with resolution in all but 2. Fourteen (93%) had coronary artery abnormalities, with normalization in 6. ECG abnormalities were present in 9 (60%) which normalized in 6 by discharge. Ten (67%) needed inotropes and/or vasopressors. None needed extracorporeal life support. Improvement in cardiac biochemical markers was closely followed by improvement in ECG/echocardiogram. All patients were discharged alive and twelve (80%) have been reviewed since. Our entire cohort with PIMS-TS had cardiac involvement and this degree of involvement is significantly more than other published series and emphasizes the need for specialist cardiac review. We believe that our multi-disciplinary team approach was crucial for the good short-term outcomes.

BACKGROUND: Infections remain a part of the natural course of cancer, and lung cancer patients often present with some form of respiratory infection that can lead to their ultimate demise. METHODS: Data was gathered concerning all unplanned hospital admissions (UHAs) to our centre from three separate patient cohorts; non-small cell lung cancer (NSCLC) patients (cohort 1), “other cancer” patients (breast, prostate, colon) (cohort 2) and all non-cancer patients (cohort 3). RESULTS: Across the three cohorts, there were 455, 1,190 and 54,158 individual patient UHAs to our centre respectively. Within the NSCLC cohort, 164 UHAs were as a direct result of pneumonia (36.0%), compared to 1.3% and 2.2% in the other two cohorts (P

Monoclonal gammopathy of undetermined significance (MGUS) affects 3·2% of adults aged >50 years. MGUS carries a life-long risk of progression to multiple myeloma and causes complications including infection and renal impairment; common causes of hospital admission. This study aimed to assess MGUS prevalence in emergency medical hospital admissions. Patients were recruited from unselected emergency medical admissions in a hospital in the United Kingdom. Serum protein electrophoresis was performed, with immunofixation of abnormal results. Reason for admission and routine test results were recorded. After education about MGUS and myeloma, patients chose whether they wished to be informed of new diagnoses. A total of 660 patients were tested and 35 had a paraprotein suggestive of MGUS. The overall rate of MGUS was 5·3%. MGUS prevalence in those aged >50 years was 6·94%, higher than the previously published rate of 3·2% (P < 0·0005). There were higher rates in those with chronic kidney disease (13·75% vs. 4·14%, P = 0·002), heart failure (14% vs. 4·59%, P = 0·012), anaemia (8·96% vs. 3·41%, P = 0·003) or leucocytosis (9·33% vs. 3·04%, P = 0·002). In all, 96% of patients wished to be informed of their screening results. The prevalence of MGUS in emergency hospital admissions is higher than expected based on previous population-based rates. This may suggest a selected population for screening.

Tumour-infiltrating immune cells have been widely implicated to play a significant role in carcinogenesis, through both pro- or anti-tumour effects. The multi-faceted effects of lung cancer associated T lymphocytes have been extensively studied, and yet, the role of B lymphocytes remains an area less studied. In this review, we will describe the current understanding of the role of tumour-infiltrating B lymphocytes in NSCLC, discuss their prognostic significance, their functionality within the tumour microenvironment and ultimately how we might harness B-cell biology to develop B-cell therapeutic strategies in cancer.

OBJECTIVESTo determine whether population-level implementation of a test-and- treat approach to correction of sub-optimal vitamin D status (25-hydroxyvitamin D [25(OH)D] DESIGNPhase 3 open-label randomised controlled trial (CORONAVIT) utilising trials-within-cohorts (TwiCs) methodology.SETTINGUnited Kingdom.PARTICIPANTS6200 adults aged 16 years or older, who were not already taking vitamin D supplements at baseline.INTERVENTIONSOffer of a postal finger-prick test of blood 25(OH)D concentration with provision of a 6-month supply of higher-dose vitamin D (3200 IU/day, n=1550) or lower-dose vitamin D (800 IU/day, n=1550) to those with blood 25(OH)D concentration th December 2020 to 16th June 2021.MAIN OUTCOME MEASURESThe primary outcome was the proportion of participants experiencing at least one doctor- or swab test-confirmed ARI of any cause. Secondary outcomes included the proportion of participants developing swab test-confirmed COVID-19. Logistic regression was used to calculate odds ratios and associated 95% confidence intervals.RESULTSOf 3100 participants offered 25(OH)D testing, 2958 (95.4%) accepted, and 2690 (86.8%) had 25(OH)D CONCLUSIONSAmong adults with a high baseline prevalence of sub-optimal vitamin D status, implementation of a population-level test-and-treat approach to vitamin D replacement did not reduce risk of all-cause ARI or COVID-19.TRIAL REGISTRATIONClinicalTrials.gov no. NCT04579640SUMMARY BOXWhat is already known on this topic?Vitamin D metabolites support innate immune responses to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and other respiratory pathogens. Sub-optimal vitamin D status (25-hydroxyvitamin D What this study addsThis phase 3 randomised controlled trial, including 6200 participants, shows that implementation of a population-level test-and-treat approach to oral vitamin D replacement at a dose of 800 IU or 3200 IU per day did not reduce risk of all-cause ARI or COVID-19 among adults with a high baseline prevalence of sub-optimal vitamin D status.

In this population-based cohort of 7530 adults, combined IgG/A/M anti-Spike titres measured after SARS-CoV-2 vaccination were predictive of protection against breakthrough SARS-CoV-2 infection. Discrimination was significantly improved by adjustment for factors influencing risk of SARS-CoV-2 exposure including household overcrowding, public transport use, and visits to indoor public places.

Extensive glycosylation of viral glycoproteins is a key feature of the antigenic surface of viruses and yet glycan processing can also be influenced by the manner of their recombinant production. The low yields of the soluble form of the trimeric spike (S) glycoprotein from SARS-CoV-2 has prompted advances in protein engineering that have greatly enhanced the stability and yields of the glycoprotein. The latest expression-enhanced version of the spike incorporates six proline substitutions to stabilize the prefusion conformation (termed SARS-CoV-2 S HexaPro). Although the substitutions greatly enhanced expression whilst not compromising protein structure, the influence of these substitutions on glycan processing has not been explored. Here, we show that the site-specific N-linked glycosylation of the expression-enhanced HexaPro resembles that of an earlier version containing two proline substitutions (2P), and that both capture features of native viral glycosylation. However, there are site-specific differences in glycosylation of HexaPro when compared to 2P. Despite these discrepancies, analysis of the serological reactivity of clinical samples from infected individuals confirmed that both HexaPro and 2P protein are equally able to detect IgG, IgA, and IgM responses in all sera analysed. Moreover, we extend this observation to include an analysis of glycan engineered S protein, whereby all N-linked glycans were converted to oligomannose-type and conclude that serological activity is not impacted by large scale changes in glycosylation. These observations suggest that variations in glycan processing will not impact the serological assessments currently being performed across the globe.

Background Vaccination prevents severe morbidity and mortality from COVID-19 in the general population. The immunogenicity and efficacy of SARS-CoV-2 vaccines in patients with antibody deficiency is poorly understood. Objectives COVID in patients with antibody deficiency (COV-AD) is a multi-site United Kingdom study that aims to determine the immune response to SARS-CoV-2 infection and vaccination in patients with primary or secondary antibody deficiency, a population that suffers from severe and recurrent infection and does not respond well to vaccination. Methods Individuals on immunoglobulin replacement therapy or with an IgG less than 4g/L receiving antibiotic prophylaxis were recruited from April 2021. Serological and cellular responses were determined using ELISA, live-virus neutralisation and interferon gamma release assays. SARS-CoV-2 infection and clearance were determined by PCR from serial nasopharyngeal swabs. Results 5.6% (n=320) of the cohort reported prior SARS-CoV-2 infection, but only 0.3% remained PCR positive on study entry. Seropositivity, following two doses of SARS-CoV-2 vaccination, was 54.8% (n=168) compared with 100% of healthy controls (n=205). The magnitude of the antibody response and its neutralising capacity were both significantly reduced compared to controls. Participants vaccinated with the Pfizer/BioNTech vaccine were more likely to be seropositive (65.7% vs 48.0%, p=0.03) and have higher antibody levels compared with the AstraZeneca vaccine (IgGAM ratio 3.73 vs 2.39, p=0.0003). T cell responses post vaccination were demonstrable in 46.2% of participants, were associated with better antibody responses but there was no difference between the two vaccines. Eleven vaccine-breakthrough infections have occurred to date, 10 of them in recipients of the AstraZeneca vaccine. Conclusion SARS-CoV-2 vaccines demonstrate reduced immunogenicity in patients with antibody deficiency with evidence of vaccine breakthrough infection.

Variants of SARS-CoV-2 may evade natural and vaccine induced immunity and monoclonal antibody immunotherapeutics. There is an urgent need to know how well antibodies, induced by healthy and Clinically Extremely Vulnerable (CEV) patients, will bind and thus help reduce transmission and severity of infection from variants of concern (VOC). This study determines the cross-reactive binding of serum antibodies obtained prior to and 28 days after a third vaccination in three cohorts; a health care worker cohort who received three doses of Pfizer-BioNtech (PPP), a cohort of CEV patients received two doses of the AstraZeneca-ChAdOx1-nCoV-19 (AAP) vaccine, followed by a third PFZ vaccine and a haemodialysis cohort that had a mixture of two AZ or PFZ vaccines followed by a PFZ booster. Six months post second vaccine there was evidence of antibody waning with 58.9% of individuals in the HD cohort seropositive against Wuhan, 34.4% Delta and 62.2% Omicron strains. For the AAP cohort, equivalent figures were 62.5%, 45.8% and 91.7% and the PPP cohort 92.2%, 90% and 91.1%. Post third dose vaccination there were universal increases in seropositivity and median optical density. For the HD cohort, 98.8% were seropositive to the Wuhan strain, 97.6% against Delta and 100% against Omicron strains. For the PPP and AAP cohorts, 100% were seropositive against all 3 strains. Lastly, we examined the WHO NIBSC 20/136 standard and there was no loss of antibody binding to either VOC. Similarly, a dilution series of Sotrovimab (GSK) found this therapeutic monoclonal antibody bound similarly to all VOC.HighlightsIgG anti-SARS-CoV-2 Omicron spike glycoprotein antibody levels were high in 100% of health care workers (HCW), a general practice population considered clinically extremely vulnerable (CEV) and haemodialysis patients (HD) 4 weeks after a third SARS-CoV-2 vaccine dose (Pfizer-BioNtech-PFZ).For both Delta and Omicron variant spike glycoproteins these antibody levels were highest in the CEV cohort who had previously received two doses of AstraZeneca ChAdOx1 nCoV-19 vaccine (AAP), lower in HCW who had previously received two doses of PFZ (PPP) and lowest in HD who had a mix of vaccines for the first and second dosePrior to this third vaccine dose and 6 months post second vaccine dose there was evidence of significant waning of antibodies against VOC.

SummaryBackgroundAntibody responses to SARS-CoV-2 vaccination vary for reasons that remain poorly understood.MethodsWe tested for presence of combined IgG, IgA and IgM (IgGAM) anti-spike antibodies before and after administration of two doses of ChAdOx1 nCoV-19 (ChAdOx1, Oxford-AstraZeneca) or BNT162b2 (Pfizer-BioNTech) in UK adults participating in a population-based longitudinal study who received their first dose of vaccine from December 15, 2020 to July 10, 2021. Information on sixty-six potential sociodemographic, behavioural, clinical, pharmacological and nutritional determinants of serological response to vaccination was captured using serial online questionnaires. We used logistic regression to estimate multivariable-adjusted odds ratios (aORs) for associations between independent variables and risk of seronegativity following two vaccine doses. Participants who were seronegative after receiving two vaccine doses were offered an additional antibody test following subsequent administration of a ‘booster’ dose of BNT162b2 or mRNA-1273 (Moderna) from September 23 to December 12, 2021.FindingsSerology results following two vaccine doses were available for 9,101 participants, of whom 5,770 (63.4%) received ChAdOx1 and 3,331 (36.6%) received BNT162b2. Anti-spike IgGAM was undetectable in 378 (4.2%) participants at a median of 8.6 weeks (IQR 6.4-10.7 weeks) after their second dose of vaccine. Seronegativity following two doses of SARS-CoV-2 vaccination was associated with administration of ChAdOx1 vs BNT162b2 (aOR 7.03, 95% CI 4.39-11.24), shorter interval between first and second vaccine doses (aOR 2.37, 1.06-5.26, for vs >10 weeks; aOR 1.59, 1.18-2.13, for 6-10 weeks vs >10 weeks), poorer self-assessed general health (aOR 3.33, 1.49-7.46, for poor vs excellent), immunodeficiencies (aOR 6.75, 2.63-17.35) and prescription of systemic immunosuppressants (aOR 3.76, 2.44-5.78). By contrast, pre-vaccination SARS-CoV-2 seropositivity (aOR 0.16, 0.04-0.70, for symptomatic seropositives vs seronegatives) and supplemental vitamin D intake (aOR 0.73, 0.53-0.99) were associated with reduced risk of post-vaccination seronegativity. 247/378 (65.3%) of participants who were seronegative after two doses of ChAdOx1 vs BNT162b2 provided a third sample at a median of 7.8 weeks (IQR 5.8-10.4) after receiving a booster dose of BNT162b2 or mRNA-1273: eight (3.2%) of them remained seronegative after three vaccine doses, all of whom either had a primary immunodeficiency or were taking systemic immunosuppressant drugs.InterpretationWe identify multiple determinants of antibody responses to two doses of ChAdOx1 or BNT162b2, many of which are potentially modifiable. Booster doses of BNT162b2 or mRNA-1273 were highly effective in achieving seroconversion in those who failed to mount antibody responses following two doses of ChAdOx1 or BNT162b2.Study registrationhttps://clinicaltrials.gov/ct2/show/NCT04330599FundingBarts Charity, Fischer Family Trust, The Exilarch’s Foundation, DSM Nutritional Products, Health Data Research UKResearch in contextEvidence before this studyWe searched PubMed, medRxiv, and Google Scholar for papers published from January 1, 2020, to February 1, 2022, using the search terms (antibody OR humoral OR serologic* OR immunogenic*) AND (SARS-CoV-2 vaccine OR ChAdOx1 or BNT162b2 coronavirus), with no language restrictions. Population-based studies investigating multiple potential determinants of vaccine immunogenicity in people with known pre-vaccination SARS-CoV-2 serostatus are lacking.Added value of this studyThis large population-based study, conducted in a population with known pre-vaccination SARS-CoV-2 serostatus, examines a comprehensive range of potential sociodemographic, behavioural, clinical, pharmacological and nutritional determinants of antibody responses to administration of two major SARS-CoV-2 vaccines (i.e., ChAdOx1 or BNT162b2), many of which have not previously been investigated. It is also the first population-based study to characterise antibody responses to booster doses of SARS-CoV-2 vaccines in adults who were seronegative after their primary course of vaccination.Implications of all the available evidenceIncreased risk of seronegativity following two doses of SARS-CoV-2 vaccines was associated with administration of ChAdOx1 vs BNT162b2, shorter interval between first and second vaccine doses, poorer self-assessed general health, immunocompromise and SARS-CoV-2 seronegativity pre-vaccination. Regular intake of vitamin D supplements was associated with reduced risk of post-vaccination seronegativity. Randomised controlled trials are now needed to test for causality. Booster doses of BNT162b2 or mRNA-1273 were highly effective in achieving seroconversion in the majority of people who failed to mount antibody responses following a primary course of vaccination, the few exceptions being a subset of those with primary immunodeficiency or systemic immunosuppressant drugs.

Background: \ud The SARS-CoV-2 delta (B.1.617.2) variant was first detected in England in March, 2021. It has since rapidly become the predominant lineage, owing to high transmissibility. It is suspected that the delta variant is associated with more severe disease than the previously dominant alpha (B.1.1.7) variant. We aimed to characterise the severity of the delta variant compared with the alpha variant by determining the relative risk of hospital attendance outcomes.\ud \ud Methods: \ud This cohort study was done among all patients with COVID-19 in England between March 29 and May 23, 2021, who were identified as being infected with either the alpha or delta SARS-CoV-2 variant through whole-genome sequencing. Individual-level data on these patients were linked to routine health-care datasets on vaccination, emergency care attendance, hospital admission, and mortality (data from Public Health England's Second Generation Surveillance System and COVID-19-associated deaths dataset; the National Immunisation Management System; and NHS Digital Secondary Uses Services and Emergency Care Data Set). The risk for hospital admission and emergency care attendance were compared between patients with sequencing-confirmed delta and alpha variants for the whole cohort and by vaccination status subgroups. Stratified Cox regression was used to adjust for age, sex, ethnicity, deprivation, recent international travel, area of residence, calendar week, and vaccination status.\ud \ud Findings: \ud Individual-level data on 43 338 COVID-19-positive patients (8682 with the delta variant, 34 656 with the alpha variant; median age 31 years [IQR 17–43]) were included in our analysis. 196 (2·3%) patients with the delta variant versus 764 (2·2%) patients with the alpha variant were admitted to hospital within 14 days after the specimen was taken (adjusted hazard ratio [HR] 2·26 [95% CI 1·32–3·89]). 498 (5·7%) patients with the delta variant versus 1448 (4·2%) patients with the alpha variant were admitted to hospital or attended emergency care within 14 days (adjusted HR 1·45 [1·08–1·95]). Most patients were unvaccinated (32 078 [74·0%] across both groups). The HRs for vaccinated patients with the delta variant versus the alpha variant (adjusted HR for hospital admission 1·94 [95% CI 0·47–8·05] and for hospital admission or emergency care attendance 1·58 [0·69–3·61]) were similar to the HRs for unvaccinated patients (2·32 [1·29–4·16] and 1·43 [1·04–1·97]; p=0·82 for both) but the precision for the vaccinated subgroup was low.\ud \ud Interpretation: \ud This large national study found a higher hospital admission or emergency care attendance risk for patients with COVID-19 infected with the delta variant compared with the alpha variant. Results suggest that outbreaks of the delta variant in unvaccinated populations might lead to a greater burden on health-care services than the alpha variant.\ud \ud Funding: \ud Medical Research Council; UK Research and Innovation; Department of Health and Social Care; and National Institute for Health Research.

Antibodies specific for the spike glycoprotein (S) and nucleocapsid (N) SARS-CoV-2 proteins are typically present during severe COVID-19, and induced to S after vaccination. The binding of viral antigens by antibody can initiate the classical complement pathway. Since complement could play pathological or protective roles at distinct times during SARS-CoV-2 infection we determined levels of antibody-dependent complement activation along the complement cascade. Here, we used an ELISA assay to assess complement protein binding (C1q) and the deposition of C4b, C3b, and C5b to S and N antigens in the presence of antibodies to SARS-CoV-2 from different test groups: non-infected, single and double vaccinees, non-hospitalised convalescent (NHC) COVID-19 patients and convalescent hospitalised (ITU-CONV) COVID-19 patients. C1q binding correlates strongly with antibody responses, especially IgG1 levels. However, detection of downstream complement components, C4b, C3b and C5b shows some variability associated with the subject group from whom the sera were obtained. In the ITU-CONV, detection of C3b-C5b to S was observed consistently, but this was not the case in the NHC group. This is in contrast to responses to N, where median levels of complement deposition did not differ between the NHC and ITU-CONV groups. Moreover, for S but not N, downstream complement components were only detected in sera with higher IgG1 levels. Therefore, the classical pathway is activated by antibodies to multiple SARS-CoV-2 antigens, but the downstream effects of this activation may differ depending the disease status of the subject and on the specific antigen targeted.

Antibodies specific for the spike glycoprotein (S) and nucleocapsid (N) SARS-CoV-2 proteins are typically present during severe COVID-19, and induced to S after vaccination. The binding of viral antigens by antibody can initiate the classical complement pathway. Since complement could play pathological or protective roles at distinct times during SARS-CoV-2 infection we determined levels of antibody-dependent complement activation along the complement cascade. Here, we used an ELISA assay to assess complement protein binding (C1q) and the deposition of C4b, C3b, and C5b to S and N antigens in the presence of anti-SARS-CoV-2 antibodies from different test groups: non-infected, single and double vaccinees, non-hospitalised convalescent (NHC) COVID-19 patients and convalescent hospitalised (ITU-CONV) COVID-19 patients. C1q binding correlates strongly with antibody responses, especially IgG1 levels. However, detection of downstream complement components, C4b, C3b and C5b shows some variability associated with the antigen and subjects studied. In the ITU-CONV, detection of C3b-C5b to S was observed consistently, but this was not the case in the NHC group. This is in contrast to responses to N, where median levels of complement deposition did not differ between the NHC and ITU-CONV groups. Moreover, for S but not N, downstream complement components were only detected in sera with higher IgG1 levels. Therefore, the classical pathway is activated by antibodies to multiple SARS-CoV-2 antigens, but the downstream effects of this activation may differ depending on the specific antigen targeted and the disease status of the subject.