Characteristics and roles of severe acute respiratory syndrome coronavirus 2-specific antibodies in patients with different severities of coronavirus 19

The diagnosis of coronavirus 19 (COVID-19) relies mainly upon viral nucleic acid detection, but false negatives can lead to missed diagnosis and misdiagnosis; severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-specific antibody detection is convenient, safe and highly sensitive. Immunoglobulin (Ig)M and IgG are commonly used to serologically diagnose COVID-19; however, the role of IgA is not well known. We aimed to quantify the levels of SARS-CoV-2-specific IgM, IgA and IgG antibodies, identify changes in them based on COVID-19 severity, and establish the significance of combined antibody detection. COVID-19 patients, divided into a severe and critical group and a moderate group, and non-COVID-19 patients with respiratory disease were included in this study. A chemiluminescence method was used to detect the levels of SARS-CoV-2-specific IgM, IgA and IgG in the blood samples from the three groups. Epidemiological characteristics, symptoms, blood test results and other data were recorded for all patients. Compared to the traditional IgM–IgG combined antibodies, IgA–IgG combined antibodies are more effective for diagnosing COVID-19. During the disease process, IgA appeared first and disappeared last. All three antibodies had significantly higher levels in COVID-19 patients than in non-COVID-19 patients. IgA and IgG were also higher for severe and critical disease than for moderate disease. All antibodies were at or near low levels at the time of tracheal extubation in critical patients. Detection of SARS-CoV-2-specific combined IgA–IgG antibodies is advantageous in diagnosing COVID-19. IgA detection is suitable during early and late stages of the disease. IgA and IgG levels correspond to disease severity.     

Characterization of SARS-CoV-2-specific humoral immunity and its potential applications and therapeutic prospects

Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is an ongoing pandemic that poses a great threat to human health worldwide. As the humoral immune response plays essential roles in disease occurrence and development, understanding the dynamics and characteristics of virus-specific humoral immunity in SARS-CoV-2-infected patients is of great importance for controlling this disease. In this review, we summarize the characteristics of the humoral immune response after SARS-CoV-2 infection and further emphasize the potential applications and therapeutic prospects of SARS-CoV-2-specific humoral immunity and the critical role of this immunity in vaccine development. Notably, serological antibody testing based on the humoral immune response can guide public health measures and control strategies; however, it is not recommended for population surveys in areas with very low prevalence. Existing evidence suggests that asymptomatic individuals have a weaker immune response to SARS-CoV-2 infection, whereas SARS-CoV-2-infected children have a more effective humoral immune response than adults. The correlations between antibody (especially neutralizing antibody) titers and protection against SARS-CoV-2 reinfection should be further examined. In addition, the emergence of cross-reactions among different coronavirus antigens in the development of screening technology and the risk of antibody-dependent enhancement related to SARS-CoV-2 vaccination should be given further attention.     

IgG autoantibodies against ACE2 in SARS-CoV-2 infected patients

How frequently autoantibodies against angiotensin-converting enzyme 2 (ACE2) occur in patients infected by SARS-CoV-2 is understudied and limited to investigations on a small sample size. The presence of these antibodies may contribute to the long-lasting effects of COVID-19 observed in some individuals, particularly if IgG-class antibodies would emerge in patients. This study assessed the prevalence of IgG autoantibodies against ACE2 in 1139 patients infected with SARS-CoV-2 and examined their relationship with severity, demographic characteristics, and status of vaccination against influenza. The overall prevalence of anti-ACE IgG antibodies in our cohort was 1.5%. Most of these individuals were men (76.5%) and underwent mild COVID-19, but some severe and asymptomatic cases were also observed. Patients with severe infection had twofold higher titers than mild and asymptomatic cases. Age, comorbidities, and influenza vaccination status were not related to antibody prevalence. The prevalence of IgG anti-SARS-CoV-2 antibodies (against nucleocapsid protein and S2 subunit, but not against receptor-binding domain) was higher in the subset with ACE2 autoantibodies. Further research is required to understand the potential spectrum and duration of effects of IgG autoantibodies against ACE2 in patients after SARS-CoV-2 infection, particularly concerning long COVID-19.     

Development of chemiluminescent lab-on-fiber immunosensors for rapid point-of-care testing of anti-SARS-CoV-2 antibodies and evaluation of longitudinal immune response kinetics following three-dose inactivation virus vaccination

Developing reliable, rapid, and quantitative point-of-care testing (POCT) technology of SARS-CoV-2-specific antibodies and understanding longitudinal vaccination response kinetics are highly required to restrain the ongoing coronavirus disease 2019 (COVID-19) pandemic. We demonstrate a novel portable, sensitive, and rapid chemiluminescent lab-on-fiber detection platform for detection of anti-SARS-CoV-2 antibodies: the chemiluminescent lab-on-fiber immunosensor (c-LOFI). Using SARS-CoV-2 Spike S1 RBD protein functionalized fiber bio-probe, the c-LOFI can detect anti-SARS-CoV-2 immunoglobulin G (IgG) and immunoglobulin M (IgM) antibodies with high sensitivity based on their respective horseradish peroxidase-labeled secondary antibodies. The limits of detection of anti-SARS-CoV-2 IgG and IgM antibodies were 0.6 and 0.3 ng/ml, respectively. The c-LOFI was successfully applied for direct detection of anti-SARS-CoV-2 antibodies in whole blood samples with simple dilution, which can serve as a finger prick test to rapidly detect antibodies. Furthermore, the longitudinal immune response (>12 months) kinetics following three-dose inactivated virus vaccines was evaluated based on anti-SARS-CoV-2 IgG detection results, which can provide important significance for understanding the immune mechanism against COVID-19 and identify individuals who may benefit from the vaccination and booster vaccination. The c-LOFI has great potential to become a sensitive, low-cost, rapid, high-frequency POCT tool for the detection of both SARS-CoV-2-specific antibodies and other biomarkers.     

Retest positive for SARS-CoV-2 RNA of “recovered” patients with COVID-19: Persistence,sampling issues,or re-infection?

“Retest Positive” for severe acute respiratory syndrome-related coronavirus-2 (SARS-CoV-2) from “recovered” coronavirus disease-19 (COVID-19) has been reported and raised several important questions for this novel coronavirus and COVID-19 disease. In this commentary, we discussed several questions: (a) Can SARS-CoV-2 re-infect the individuals who recovered from COVID-19? This question is also associated with other questions: whether or not SARS-CoV-2 infection induces protective reaction or neutralized antibody? Will SARS-CoV-2 vaccines work? (b) Why could some recovered patients with COVID-19 be re-tested positive for SARS-CoV-2 RNA? (c) Are some recovered pwith atients COVID-19 with re-testing positive for SARS-CoV-2 RNA infectious? and (d) How should the COVID-19 patients with retest positive for SARS-CoV-2 be managed?     

Antibody isotype epitope mapping of SARS-CoV-2 Spike RBD protein: Targets for COVID-19 symptomatology and disease control

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) still poses a challenge for biomedicine and public health. To advance the development of effective diagnostic, prognostic, and preventive interventions, our study focused on high-throughput antibody binding epitope mapping of the SARS-CoV-2 spike RBD protein by IgA, IgM and IgG antibodies in saliva and sera of different cohorts from healthy uninfected individuals to SARS-CoV-2-infected unvaccinated and vaccinated asymptomatic, recovered, nonsevere, and severe patients. Identified candidate diagnostic (455-LFRKSNLKPFERD-467), prognostic (395-VYADSFVIRGDEV-407-C-KLH, 332-ITNLCPFGEV-342-C-KLH, 352-AWNRKRI-358-C-KLH, 524-VCGPKKSTNLVKN-536-KLH), and protective (MKLLE-487-NCYFPLQSYGFQPTNGVG-504-GGGGS-446-GGNYNYLYRLFRKSNLKPFERD-467) epitopes were validated with sera from prevaccine and postvaccine cohorts. The results identified neutralizing epitopes and support that antibody recognition of linear B-cell epitopes in RBD protein is associated with antibody isotype and disease symptomatology. The findings in asymptomatic individuals suggest a role for anti-RBD antibodies in the protective response against SARS-CoV-2. The possibility of translating results into diagnostic interventions for the early diagnosis of asymptomatic individuals and prognosis of disease severity provides new tools for COVID-19 surveillance and evaluation of risks in hospitalized patients. These results, together with other approaches, may contribute to the development of new vaccines for the control of COVID-19 and other coronavirus-related diseases using a quantum vaccinomics approach through the combination of protective epitopes.     

Long-term coexistence of SARS-CoV-2 with antibody response in COVID-19 patients

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection causing coronavirus disease 2019 (COVID-19) has spread worldwide. Whether antibodies are important for the adaptive immune responses against SARS-CoV-2 infection needs to be determined. Here, 26 cases of COVID-19 in Jinan, China, were examined and shown to be mild or with common clinical symptoms, and no case of severe symptoms was found among these patients. Strikingly, a subset of these patients had SARS-CoV-2 and virus-specific IgG coexist for an unexpectedly long time, with two cases for up to 50 days. One COVID-19 patient who did not produce any SARS-CoV-2–bound IgG successfully cleared SARS-CoV-2 after 46 days of illness, revealing that without antibody-mediated adaptive immunity, innate immunity alone may still be powerful enough to eliminate SARS-CoV-2. This report may provide a basis for further analysis of both innate and adaptive immunity in SARS-CoV-2 clearance, especially in nonsevere cases.     

SARS-CoV-2 antibody dynamics and B-cell memory response over time in COVID-19 convalescent subjects

ObjectivesThe worldwide spread of coronavirus disease 2019 (COVID-19) highlights the need for assessment of long-term humoral immunity in convalescent subjects. Our objectives were to evaluate long-term IgG antibody response to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and B-cell memory response in COVID-19 convalescent subjects.MethodsBlood samples were collected from a cohort of subjects recovering from COVID-19 and from healthy subjects who donated blood. SARS-CoV-2 IgG antibodies were quantitatively detected by ELISA using anti-S1 spike IgG. SARS-CoV-2 spike-specific IgG memory B cells were evaluated by reversed B-cell FluroSpot based on human IgG SARS-CoV-2 receptor-binding domain in a randomly selected group of subjects recovering from COVID-19. Statistical analysis was performed with clinical variables and time post COVID-19 infection.ResultsAntibody response was not detected in 26 of 392 COVID-19 convalescent subjects (6.6%). Over a period of 9 months, the level of antibodies decreased by 50% but stabilized at 6 months, and a protective level prevailed for up to 9 months. No differences were found regarding IgG SARS-CoV-2 antibody levels for age, gender, and major blood types over time. Over time, asymptomatic COVID-19 subjects did not differ in antibody level from subjects with mild to severe disease. Repeated paired IgG SARS-CoV-2 antibody level analyses disclosed that, over 6 and 9 months, 15.3% (nine of 59) and 15.8% (three of 19) of subjects became SARS-CoV-2 IgG-seronegative, respectively, all with a low antibody level at 3 months. Rate of antibody decline was not affected by age, gender, or clinical symptomatology. In a subgroup of recovering subjects, memory B-cell response up to 9 months post-COVID-19 infection was undetectable in 31.8% of subjects (14/44), and there was no correlation with age, SARS-CoV-2 antibody level, or time post infection.ConclusionsThe majority of convalescent COVID-19 subjects develop an IgG SARS-CoV-2 antibody response and a protective level prevails over a period of up to 9 months, regardless of age, gender, major blood types or clinical symptomatology.     

A surrogate cell-based SARS-CoV-2 spike blocking assay

To monitor infection by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and successful vaccination against coronavirus disease 2019 (COVID-19), the kinetics of neutralizing or blocking anti-SARS-CoV-2 antibody titers need to be assessed. Here, we report the development of a quick and inexpensive surrogate SARS-CoV-2 blocking assay (SUBA) using immobilized recombinant human angiotensin-converting enzyme 2 (hACE2) and human cells expressing the native form of surface SARS-CoV-2 spike protein. Spike protein-expressing cells bound to hACE2 in the absence or presence of blocking antibodies were quantified by measuring the optical density of cell-associated crystal violet in a spectrophotometer. The advantages are that SUBA is a fast and inexpensive assay, which does not require biosafety level 2- or 3-approved laboratories. Most importantly, SUBA detects blocking antibodies against the native trimeric cell-bound SARS-CoV-2 spike protein and can be rapidly adjusted to quickly pre-screen already approved therapeutic antibodies or sera from vaccinated individuals for their ACE2 blocking activities against any emerging SARS-CoV-2 variants.     

Role of the humoral immune response during COVID-19: guilty or not guilty?

Systemic and mucosal humoral immune responses are crucial to fight respiratory viral infections in the current pandemic of COVID-19 caused by the SARS-CoV-2 virus. During SARS-CoV-2 infection, the dynamics of systemic and mucosal antibody infections are affected by patient characteristics, such as age, sex, disease severity, or prior immunity to other human coronaviruses. Patients suffering from severe disease develop higher levels of anti-SARS-CoV-2 antibodies in serum and mucosal tissues than those with mild disease, and these antibodies are detectable for up to a year after symptom onset. In hospitalized patients, the aberrant glycosylation of anti-SARS-CoV-2 antibodies enhances inflammation-associated antibody Fc-dependent effector functions, thereby contributing to COVID-19 pathophysiology. Current vaccines elicit robust humoral immune responses, principally in the blood. However, they are less effective against new viral variants, such as Delta and Omicron. This review provides an overview of current knowledge about the humoral immune response to SARS-CoV-2, with a particular focus on the protective and pathological role of humoral immunity in COVID-19 severity. We also discuss the humoral immune response elicited by COVID-19 vaccination and protection against emerging viral variants.     

Intranasal Sendai virus-based SARS-CoV-2 vaccine using a mouse model

The coronavirus disease 2019 (COVID-19) epidemic remains worldwide. The usefulness of the intranasal vaccine and boost immunization against severe acute respiratory syndrome-related coronavirus (SARS-CoV-2) has recently received much attention. We developed an intranasal SARS-CoV-2 vaccine by loading the receptor binding domain of the S protein (S-RBD) of SARS-CoV-2 as an antigen into an F-deficient Sendai virus vector. After the S-RBD-Fd antigen with trimer formation ability was intranasally administered to mice, S-RBD-specific IgM, IgG, IgA, and neutralizing antibody titers were increased in serum or bronchoalveolar lavage fluid for 12 weeks. Furthermore, in mice that received a booster dose at week 8, a marked increase in neutralizing antibodies in the serum and bronchoalveolar lavage fluid was observed at the final evaluation at week 12, which neutralized the pseudotyped lentivirus expressing the SARS-CoV-2 spike protein, indicating the usefulness of the Sendai virus-based SARS-CoV-2 intranasal vaccine.     

新型冠状病毒感染者和灭活疫苗受种者抗体水平研究

目的:通过观测不同时期新型冠状病毒(severe acute respiratory syndrome coronavirus 2,SARS-CoV-2)感染者和SARS-CoV-2灭活疫苗受种者血清病毒特异性IgM和IgG抗体水平,增进对SARS-CoV-2和SARS-CoV-2灭活疫苗进入机体后免疫学特征的了解。方...     

Divergent SARS-CoV-2-specific T- and B-cell responses in severe but not mild COVID-19 patients

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of the current coronavirus disease 2019 (COVID-19) pandemic. Understanding the immune response that provides specific immunity but may also lead to immunopathology is crucial for the design of potential preventive and therapeutic strategies. Here, we characterized and quantified SARS-CoV-2-specific immune responses in patients with different clinical courses. Compared to individuals with a mild clinical presentation, CD4⁺ T-cell responses were qualitatively impaired in critically ill patients. Strikingly, however, in these patients the specific IgG antibody response was remarkably strong. Furthermore, in these critically ill patients, a massive influx of circulating T cells into the lungs was observed, overwhelming the local T-cell compartment, and indicative of vascular leakage. The observed disparate T- and B-cell responses could be indicative of a deregulated immune response in critically ill COVID-19 patients.     

Immune repertoire sequencing reveals an abnormal adaptive immune system in COVID-19 survivors

Accumulating evidence suggests that severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) impairs the adaptive immune system during acute infection. Still, it remains largely unclear whether the frequency and functions of T and B cells return to normal after the recovery of Coronavirus Disease 2019 (COVID-19). Here, we analyzed immune repertoires and SARS-CoV-2-specific neutralization antibodies in a prospective cohort of 40 COVID-19 survivors with a 6-month follow-up after hospital discharge. Immune repertoire sequencing revealed abnormal T- and B-cell expression and function with large T cell receptor/B cell receptor clones, decreased diversity, abnormal class-switch recombination, and somatic hypermutation. A decreased number of B cells but an increased proportion of CD19⁺CD138⁺ B cells were found in COVID-19 survivors. The proportion of CD4⁺ T cells, especially circulating follicular helper T (cTfh) cells, was increased, whereas the frequency of CD3⁺CD4⁻ T cells was decreased. SARS-CoV-2-specific neutralization IgG and IgM antibodies were identified in all survivors, especially those recorded with severe COVID-19 who showed a higher inhibition rate of neutralization antibodies. All severe cases complained of more than one COVID-19 sequelae after 6 months of recovery. Overall, our findings indicate that SARS-CoV-2-specific antibodies remain detectable even after 6 months of recovery. Because of their abnormal adaptive immune system with a low number of CD3⁺CD4⁻ T cells and high susceptibility to infections, COVID-19 patients might need more time and medical care to fully recover from immune abnormalities and tissue damage.     

Antibody response using six different serological assays in a completely PCR-tested community after a coronavirus disease 2019 outbreak—the CoNAN study

ObjectivesDue to a substantial proportion of asymptomatic and mild courses, many severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections remain unreported. Therefore, assessment of seroprevalence may detect the real burden of disease. We aimed to determine and characterize the rate of SARS-CoV-2 infections and the resulting seroprevalence in a defined population. The primary objective of the study was to assess SARS-CoV-2 antibody seroprevalence using six different IgG-detecting immunoassays. Secondary objectives of the study were: (a) to determine potential risk factors for symptomatic versus asymptomatic coronavirus disease 2019 courses, and (b) to investigate the rate of virus RNA-persistence.MethodsCoNAN is a population-based cohort study performed in the community Neustadt am Rennsteig, Germany, which was quarantined from 22 March to 5 April after six SARS-CoV-2 cases were detected in the village's population. The SARS-CoV-2 outbreak comprised 51 cases and 3 deaths. The CoNAN study was performed from 13 May to 22 May 2020, 6 weeks after a SARS-CoV-2 outbreak.ResultsWe enrolled a total of 626 participants (71% of the community population) for PCR and antibody testing in the study. All actual SARS-CoV-2 PCR tests were negative. Fifty-two out of 620 (8.4%) participants had antibodies against SARS-CoV-2 in at least two different assays. There were 38 participants with previously PCR-confirmed SARS-CoV-2 infection. Of those, only 19 (50%) displayed anti-SARS-CoV-2 antibodies. We also show that antibody-positive participants with symptoms compatible with a respiratory tract infection had significantly higher antibody levels then asymptomatic participants (EU-assay: median 2.9 versus 7.2 IgG-index, p 0.002; DS-assay: median 45.2 versus 143 AU/mL, p 0.002). Persisting viral replication was not detected.ConclusionsOur data question the relevance and reliability of IgG antibody testing to detect past SARS-CoV-2 infections 6 weeks after an outbreak. We conclude that assessing immunity for SARS-CoV-2 infection should not rely on antibody tests alone.     

Humoral response to spike S1 and S2 and nucleocapsid proteins on microarray after SARS-CoV-2 infection

Many aspects of the humoral immune response to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), such as its role in protection after natural infection, are still unclear. We evaluated IgA and IgG response to spike subunits 1 and 2 (S1 and S2) and Nucleocapsid proteins of SARS-COV-2 in serum samples of 109 volunteers with viral RNA detected or seroconversion with different clinical evolution (asymptomatic, mild, moderate, and severe coronavirus disease 2019), using the ViraChip® Test Kit. We observed that the quantification of antibodies to all antigens had a positive correlation to disease severity, which was strongly associated with the presence of comorbidities. Seroreversion was not uncommon even during the short (median of 77 days) observation, occurring in 15% of mild-asymptomatic cases at a median of 55 days for IgG and 46 days for IgA. The time to reach the maximal antibody response did not differ significantly among recovered and deceased volunteers. Our study illustrated the dynamic of anti-S1, anti-N, and anti-S2 IgA and IgG antibodies, and suggests that high production of IgG and IgA does not guarantee protection to disease severity and that functional responses that have been studied by other groups, such as antibody avidity, need further attention.     

One-month humoral response following two or three doses of messenger RNA coronavirus disease 2019 vaccines as primary vaccination in specific populations in France: first results from the Agence Nationale Recherche contre le Sida (ANRS)0001S COV-POPART cohort

Objectives: We aimed to investigate the 1-month humoral response to two or three doses of a messenger RNA coronavirus disease 2019 (COVID-19) vaccine as a primary vaccination regimen in specific populations compared with that in healthy adults.MethodsAgence Nationale Recherche contre le Sida (ANRS)0001S–COV-POPART (NCT04824651) is a French nation-wide, multi-centre, prospective, observational cohort study assessing the immune response to COVID-19 vaccines routinely administered to 11 sub-groups of patients with chronic conditions and two control groups. Patients and controls who received at least two vaccine doses and whose results 1 month after the second dose were available were included. The humoral response was assessed 1 month after the first, second and third doses (if applicable) based on the percentage of responders (positive for anti-Spike severe acute respiratory syndrome coronavirus 2 [SARS-CoV-2] IgG antibodies), geometric means of anti-Spike SARS-CoV-2 IgG antibodies (enzyme-linked immunosorbent assay) and proportion of participants with anti-SARS-CoV-2-specific neutralizing antibodies (in vitro neutralization assay for the original SARS-CoV-2 strain). All analyses were centralized.ResultsWe included 4091 participants in this analysis: 2979 participants from specific sub-populations and 1112 controls. Only 522 (17.5%) participants from the specific populations received three doses as a primary vaccination regimen. Patients living with human immunodeficiency virus, cancer and diabetes had high percentages of responders after two doses, whereas patients with solid organ transplants, allogeneic hematopoietic stem cell transplants and hypogammaglobulinaemia had the lowest percentage of responders (35.9% [95% CI, 29.2–43.0], 57.4% [95% CI, 48.1–66.3] and 77.1% [95% CI, 65.6–86.3], respectively). In those who received the third dose, the percentage of responders reached 54.2% (95% CI, 42.9–65.2) (vs. 32.3% [95% CI, 16.7–51.4] after 2 doses) among those with solid organ transplants and 73.9% (95% CI, 58.9–85.7) (vs. 56.1% [95% CI, 46.2–65.7] after 2 doses) among those with hematopoietic stem cell transplants. Similar results were found with anti-SARS-CoV-2-specific neutralizing antibodies.ConclusionsA lower humoral response to COVID-19 vaccines was observed in the specific populations compared with that in the controls. The third dose of this vaccine in the primary regimen had a positive effect on the percentages of patients who developed anti-Spike IgG antibodies and specific neutralizing antibodies.     

Antibody Responses One Year after Mild SARS-CoV-2 Infection

Understanding the long-term kinetics of antibodies in coronavirus disease 2019 (COVID-19) is essential in interpreting serosurvey data. We investigated the antibody response one year after infection in 52 mildly symptomatic patients with severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) infection, using three commercial immunoassays and a surrogate virus neutralization test (sVNT) kit. Anti-N pan-immunoglobulin (Ig), anti-S IgG, and anti-S1 IgG were detected in 43 (82.7%), 44 (84.6%), and 30 (57.7%), respectively. In 49 (94.2%), the antibody could be detected by either anti-N pan-Ig or anti-S IgG assay. In the sVNT, 30 (57.7%) had positive neutralizing activity. Despite waning immunity, SARS-CoV-2 antibodies can be detected up to one year after infection, even in mild COVID-19 patients.