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1.
Nat Commun ; 12(1): 4144, 2021 07 06.
Artigo em Inglês | MEDLINE | ID: mdl-34230476

RESUMO

To investigate the duration of humoral immune response in convalescent coronavirus disease 2019 (COVID-19) patients, we conduct a 12-month longitudinal study through collecting a total of 1,782 plasma samples from 869 convalescent plasma donors in Wuhan, China and test specific antibody responses. The results show that positive rate of IgG antibody against receptor-binding domain of spike protein (RBD-IgG) to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in the COVID-19 convalescent plasma donors exceeded 70% for 12 months post diagnosis. The level of RBD-IgG decreases with time, with the titer stabilizing at 64.3% of the initial level by the 9th month. Moreover, male plasma donors produce more RBD-IgG than female, and age of the patients positively correlates with the RBD-IgG titer. A strong positive correlation between RBD-IgG and neutralizing antibody titers is also identified. These results facilitate our understanding of SARS-CoV-2-induced immune memory to promote vaccine and therapy development.


Assuntos
Anticorpos Neutralizantes/sangue , Anticorpos Antivirais/sangue , Imunoglobulina G/sangue , Receptores Virais/imunologia , SARS-CoV-2/imunologia , Adulto , Animais , Doadores de Sangue , COVID-19/terapia , Linhagem Celular , China , Chlorocebus aethiops , Convalescença , Feminino , Humanos , Imunidade Humoral/imunologia , Imunização Passiva , Memória Imunológica/imunologia , Estudos Longitudinais , Masculino , Fatores Sexuais , Glicoproteína da Espícula de Coronavírus/imunologia , Células Vero
3.
PLoS One ; 16(7): e0254367, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34242356

RESUMO

COVID-19 serological test must have high sensitivity as well as specificity to rule out cross-reactivity with common coronaviruses (HCoVs). We have developed a quantitative multiplex test, measuring antibodies against spike (S) proteins of SARS-CoV-2, SARS-CoV, MERS-CoV, and common human coronavirus strains (229E, NL63, OC43, HKU1), and nucleocapsid (N) protein of SARS-CoV viruses. Receptor binding domain of S protein of SARS-CoV-2 (S-RBD), and N protein, demonstrated sensitivity (94% and 92.5%, respectively) in COVID-19 patients (n = 53), with 98% specificity in non-COVID-19 respiratory-disease (n = 98), and healthy-controls (n = 129). Anti S-RBD and N antibodies appeared five to ten days post-onset of symptoms, peaking at approximately four weeks. The appearance of IgG and IgM coincided while IgG subtypes, IgG1 and IgG3 appeared soon after the total IgG; IgG2 and IgG4 remained undetectable. Several inflammatory cytokines/chemokines were found to be elevated in many COVID-19 patients (e.g., Eotaxin, Gro-α, CXCL-10 (IP-10), RANTES (CCL5), IL-2Rα, MCP-1, and SCGF-b); CXCL-10 was elevated in all. In contrast to antibody titers, levels of CXCL-10 decreased with the improvement in patient health suggesting it as a candidate for disease resolution. Importantly, anti-N antibodies appear before S-RBD and differentiate between vaccinated and infected people-current vaccines (and several in the pipeline) are S protein-based.


Assuntos
Anticorpos Antivirais , COVID-19 , Quimiocinas , Proteínas do Nucleocapsídeo de Coronavírus , Imunoglobulina G , Imunoglobulina M , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus , Adulto , Animais , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , COVID-19/sangue , COVID-19/imunologia , Quimiocinas/sangue , Quimiocinas/imunologia , Proteínas do Nucleocapsídeo de Coronavírus/sangue , Proteínas do Nucleocapsídeo de Coronavírus/imunologia , Feminino , Humanos , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Imunoglobulina M/sangue , Imunoglobulina M/imunologia , Macaca mulatta , Masculino , Pessoa de Meia-Idade , Fosfoproteínas/sangue , Fosfoproteínas/imunologia , Coelhos , SARS-CoV-2/imunologia , SARS-CoV-2/metabolismo , Glicoproteína da Espícula de Coronavírus/sangue , Glicoproteína da Espícula de Coronavírus/imunologia
5.
Science ; 373(6551): 231-236, 2021 07 09.
Artigo em Inglês | MEDLINE | ID: mdl-34244417

RESUMO

In mammals, early resistance to viruses relies on interferons, which protect differentiated cells but not stem cells from viral replication. Many other organisms rely instead on RNA interference (RNAi) mediated by a specialized Dicer protein that cleaves viral double-stranded RNA. Whether RNAi also contributes to mammalian antiviral immunity remains controversial. We identified an isoform of Dicer, named antiviral Dicer (aviD), that protects tissue stem cells from RNA viruses-including Zika virus and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-by dicing viral double-stranded RNA to orchestrate antiviral RNAi. Our work sheds light on the molecular regulation of antiviral RNAi in mammalian innate immunity, in which different cell-intrinsic antiviral pathways can be tailored to the differentiation status of cells.


Assuntos
RNA Helicases DEAD-box/genética , RNA Helicases DEAD-box/metabolismo , Interferência de RNA , Vírus de RNA/fisiologia , RNA Viral/metabolismo , Ribonuclease III/genética , Ribonuclease III/metabolismo , Células-Tronco/enzimologia , Células-Tronco/virologia , Processamento Alternativo , Animais , Encéfalo/enzimologia , Encéfalo/virologia , Linhagem Celular , RNA Helicases DEAD-box/química , Humanos , Imunidade Inata , Isoenzimas/química , Isoenzimas/genética , Isoenzimas/metabolismo , Camundongos , Organoides/enzimologia , Organoides/virologia , Infecções por Vírus de RNA/enzimologia , Infecções por Vírus de RNA/imunologia , Infecções por Vírus de RNA/virologia , Vírus de RNA/genética , Vírus de RNA/imunologia , RNA de Cadeia Dupla/metabolismo , RNA Interferente Pequeno/metabolismo , Ribonuclease III/química , SARS-CoV-2/genética , SARS-CoV-2/imunologia , SARS-CoV-2/fisiologia , Replicação Viral , Zika virus/genética , Zika virus/imunologia , Zika virus/fisiologia , Infecção por Zika virus/enzimologia , Infecção por Zika virus/imunologia , Infecção por Zika virus/virologia
7.
Cell Rep ; 36(2): 109353, 2021 07 13.
Artigo em Inglês | MEDLINE | ID: mdl-34237283

RESUMO

SARS-CoV-2 is one of three coronaviruses that have crossed the animal-to-human barrier and caused widespread disease in the past two decades. The development of a universal human coronavirus vaccine could prevent future pandemics. We characterize 198 antibodies isolated from four COVID-19+ subjects and identify 14 SARS-CoV-2 neutralizing antibodies. One targets the N-terminal domain (NTD), one recognizes an epitope in S2, and 11 bind the receptor-binding domain (RBD). Three anti-RBD neutralizing antibodies cross-neutralize SARS-CoV-1 by effectively blocking binding of both the SARS-CoV-1 and SARS-CoV-2 RBDs to the ACE2 receptor. Using the K18-hACE transgenic mouse model, we demonstrate that the neutralization potency and antibody epitope specificity regulates the in vivo protective potential of anti-SARS-CoV-2 antibodies. All four cross-neutralizing antibodies neutralize the B.1.351 mutant strain. Thus, our study reveals that epitopes in S2 can serve as blueprints for the design of immunogens capable of eliciting cross-neutralizing coronavirus antibodies.


Assuntos
Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/imunologia , COVID-19/imunologia , COVID-19/prevenção & controle , Glicoproteína da Espícula de Coronavírus/imunologia , Enzima de Conversão de Angiotensina 2/química , Enzima de Conversão de Angiotensina 2/imunologia , Animais , Anticorpos Antivirais/química , Anticorpos Antivirais/imunologia , Sítios de Ligação , Linhagem Celular , Reações Cruzadas , Epitopos/imunologia , Feminino , Células HEK293 , Humanos , Camundongos , Testes de Neutralização , Ligação Proteica/imunologia , Domínios Proteicos , SARS-CoV-2/imunologia , Glicoproteína da Espícula de Coronavírus/química
8.
Cell Rep ; 36(2): 109385, 2021 07 13.
Artigo em Inglês | MEDLINE | ID: mdl-34237284

RESUMO

Administration of convalescent plasma or neutralizing monoclonal antibodies (mAbs) is a potent therapeutic option for coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. However, SARS-CoV-2 variants with mutations in the spike protein have emerged in many countries. To evaluate the efficacy of neutralizing antibodies induced in convalescent patients against emerging variants, we isolate anti-spike mAbs from two convalescent COVID-19 patients infected with prototypic SARS-CoV-2 by single-cell sorting of immunoglobulin-G-positive (IgG+) memory B cells. Anti-spike antibody induction is robust in these patients, and five mAbs have potent neutralizing activities. The efficacy of most neutralizing mAbs and convalescent plasma samples is maintained against B.1.1.7 and mink cluster 5 variants but is significantly decreased against variants B.1.351 from South Africa and P.1 from Brazil. However, mAbs with a high affinity for the receptor-binding domain remain effective against these neutralization-resistant variants. Rapid spread of these variants significantly impacts antibody-based therapies and vaccine strategies against SARS-CoV-2.


Assuntos
Anticorpos Neutralizantes/imunologia , COVID-19/imunologia , COVID-19/terapia , SARS-CoV-2/imunologia , Glicoproteína da Espícula de Coronavírus/imunologia , Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/isolamento & purificação , Anticorpos Antivirais/imunologia , COVID-19/virologia , Linhagem Celular , Células HEK293 , Humanos , Imunização Passiva , Masculino , Mutação , Testes de Neutralização , Domínios Proteicos , Glicoproteína da Espícula de Coronavírus/genética
9.
Cell Rep ; 36(2): 109391, 2021 07 13.
Artigo em Inglês | MEDLINE | ID: mdl-34242574

RESUMO

The immunogenicity of the SARS-CoV-2 proteome is largely unknown, especially for non-structural proteins and accessory proteins. In this study, we collect 2,360 COVID-19 sera and 601 control sera. We analyze these sera on a protein microarray with 20 proteins of SARS-CoV-2, building an antibody response landscape for immunoglobulin (Ig)G and IgM. Non-structural proteins and accessory proteins NSP1, NSP7, NSP8, RdRp, ORF3b, and ORF9b elicit prevalent IgG responses. The IgG patterns and dynamics of non-structural/accessory proteins are different from those of the S and N proteins. The IgG responses against these six proteins are associated with disease severity and clinical outcome, and they decline sharply about 20 days after symptom onset. In non-survivors, a sharp decrease of IgG antibodies against S1 and N proteins before death is observed. The global antibody responses to non-structural/accessory proteins revealed here may facilitate a deeper understanding of SARS-CoV-2 immunology.


Assuntos
COVID-19/imunologia , SARS-CoV-2/imunologia , Glicoproteína da Espícula de Coronavírus/imunologia , Proteínas não Estruturais Virais/imunologia , Proteínas Virais Reguladoras e Acessórias/imunologia , Adulto , Idoso , Anticorpos Antivirais/imunologia , Formação de Anticorpos , Humanos , Imunoglobulina G/imunologia , Imunoglobulina M/imunologia , Masculino , Pessoa de Meia-Idade , Análise Serial de Proteínas
10.
Curr Allergy Asthma Rep ; 21(6): 38, 2021 07 14.
Artigo em Inglês | MEDLINE | ID: mdl-34259961

RESUMO

PURPOSE OF REVIEW: Increasing knowledge of the pathogenesis of the SARS-CoV-2 infection and the complex interaction between host and viral factors have allowed clinicians to stratify the severity of COVID-19 infection. Epidemiological data has also helped to model viral carriage and infectivity. This review presents a comprehensive summary of the pathophysiology of COVID-19, the mechanisms of action of the SARS-CoV-2 virus, and the correlation with the clinical and biochemical characteristics of the disease. RECENT FINDINGS: ACE2 and TMPRSS2 receptors have emerged as a key player in the mechanism of infection of SARS-CoV-2. Their distribution throughout the body has been shown to impact the organ-specific manifestations of COVID-19. The immune-evasive and subsequently immunoregulative properties of SARS-CoV-2 are also shown to be implicated in disease proliferation and progression. Information gleaned from the virological properties of SARS-CoV-2 is consistent with and reflects the clinical behavior of the COVID-19 infection. Further study of specific clinical phenotypes and severity classes of COVID-19 may assist in the development of targeted therapeutics to halt progression of disease from mild to moderate-severe. As the understanding of the pathophysiology and mechanism of action of SARS-CoV-2 continues to grow, it is our hope that better and more effective treatment options continue to emerge.


Assuntos
COVID-19/fisiopatologia , SARS-CoV-2/patogenicidade , Enzima de Conversão de Angiotensina 2/metabolismo , COVID-19/epidemiologia , COVID-19/imunologia , COVID-19/virologia , Progressão da Doença , Humanos , Evasão da Resposta Imune , Especificidade de Órgãos , SARS-CoV-2/imunologia , Serina Endopeptidases/metabolismo , Índice de Gravidade de Doença , Internalização do Vírus
11.
PLoS Pathog ; 17(7): e1009705, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-34265022

RESUMO

COVID-19 (coronavirus disease 2019) caused by SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) infection is a disease affecting several organ systems. A model that captures all clinical symptoms of COVID-19 as well as long-haulers disease is needed. We investigated the host responses associated with infection in several major organ systems including the respiratory tract, the heart, and the kidneys after SARS-CoV-2 infection in Syrian hamsters. We found significant increases in inflammatory cytokines (IL-6, IL-1beta, and TNF) and type II interferons whereas type I interferons were inhibited. Examination of extrapulmonary tissue indicated inflammation in the kidney, liver, and heart which also lacked type I interferon upregulation. Histologically, the heart had evidence of myocarditis and microthrombi while the kidney had tubular inflammation. These results give insight into the multiorgan disease experienced by people with COVID-19 and possibly the prolonged disease in people with post-acute sequelae of SARS-CoV-2 (PASC).


Assuntos
COVID-19/imunologia , Regulação para Baixo/imunologia , Interferon Tipo I/imunologia , Rim/imunologia , Miocárdio/imunologia , Sistema Respiratório/imunologia , SARS-CoV-2/imunologia , Animais , COVID-19/patologia , Cricetinae , Modelos Animais de Doenças , Humanos , Inflamação/imunologia , Inflamação/patologia , Rim/patologia , Rim/virologia , Masculino , Mesocricetus , Miocárdio/patologia , Sistema Respiratório/patologia , Sistema Respiratório/virologia
12.
Signal Transduct Target Ther ; 6(1): 271, 2021 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-34267185

RESUMO

COVID-19 vaccines from multiple manufacturers are needed to cope with the problem of insufficient supply. We did two single-center, randomised, double-blind, placebo-controlled phase 1 and phase 2 trials to assess the safety, tolerability and immunogenicity of a recombinant COVID-19 vaccine (Sf9 cells) in healthy population aged 18 years or older in China. Eligible participants were enrolled, the ratio of candidate vaccine and placebo within each dose group was 3:1 (phase 1) or 5:1 (phase 2). From August 28, 2020, 168 participants were sequentially enrolled and randomly assigned to receive the low dose vaccine, high dose vaccine or placebo with the schedule of 0, 28 days or 0, 14, 28 days in phase 1 trial. From November 18, 2020, 960 participants were randomly assigned to receive the low dose vaccine, high dose vaccine or placebo with the schedule of 0, 21 days or 0, 14, 28 days in phase 2 trial. The most common solicited injection site adverse reaction within 7 days in both trials was pain. The most common solicited systematic adverse reactions within 7 days were fatigue, cough, sore throat, fever and headache. ELISA antibodies and neutralising antibodies increased at 14 days, and peaked at 28 days (phase 1) or 30 days (phase 2) after the last dose vaccination. The GMTs of neutralising antibody against live SARS-CoV-2 at 28 days or 30 days after the last dose vaccination were highest in the adult high dose group (0, 14, 28 days), with 102.9 (95% CI 61.9-171.2) and 102.6 (95% CI 75.2-140.1) in phase 1 and phase 2 trials, respectively. Specific T-cell response peaked at 14 days after the last dose vaccination in phase 1 trial. This vaccine is safe, and induced significant immune responses after three doses of vaccination.


Assuntos
Vacinas contra COVID-19 , COVID-19 , Imunogenicidade da Vacina , SARS-CoV-2/imunologia , Adolescente , Adulto , COVID-19/sangue , COVID-19/imunologia , COVID-19/prevenção & controle , Vacinas contra COVID-19/administração & dosagem , Vacinas contra COVID-19/imunologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade
13.
Mikrochim Acta ; 188(8): 262, 2021 07 20.
Artigo em Inglês | MEDLINE | ID: mdl-34282508

RESUMO

COVID-19 is now a severe threat to global health. Facing this pandemic, we developed a space-encoding microfluidic biochip for high-throughput, rapid, sensitive, simultaneous quantitative detection of SARS-CoV-2 antigen proteins and IgG/IgM antibodies in serum. The proposed immunoassay biochip integrates the advantages of graphene oxide quantum dots (GOQDs) and microfluidic chip and is capable of conducting multiple SARS-CoV-2 antigens or IgG/IgM antibodies of 60 serum samples simultaneously with only 2 µL sample volume of each patient. Fluorescence intensity of antigens and IgG antibody detection at emission wavelength of ~680 nm was used to quantify the target concentration at excitation wavelength of 632 nm, and emission wavelength of ~519 nm was used during the detection of IgM antibodies at excitation wavelength of 488 nm. The method developed has a large linear quantification detection regime of 5 orders of magnitude, an ultralow detection limit of ~0.3 pg/mL under optimized conditions, and less than 10-min qualitative detection time. The proposed biosensing platform will not only greatly facilitate the rapid diagnosis of COVID-19 patients, but also provide a valuable screening approach for infected patients, medical therapy, and vaccine recipients.


Assuntos
Antígenos Virais/sangue , Imunoensaio , Imunoglobulina G/sangue , Imunoglobulina M/sangue , SARS-CoV-2/isolamento & purificação , Reações Antígeno-Anticorpo , Antígenos Virais/imunologia , Humanos , Imunoglobulina G/imunologia , Imunoglobulina M/imunologia , Nanopartículas/química , Tamanho da Partícula , SARS-CoV-2/imunologia , Sensibilidade e Especificidade
14.
Pediatr Infect Dis J ; 40(8): e294-e299, 2021 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-34250968

RESUMO

INTRODUCTION: Antibody response developed within 2-3 weeks after exposure to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been shown to decrease over time; however, there is limited data about antibody levels at 6 months or later postinfection, particularly in children. MATERIALS AND METHOD: A prospective multicenter study was performed using 315 samples of 74 confirmed and 10 probable coronavirus disease 2019 pediatric cases. About 20% of these cases were classified as asymptomatic, 74% as mild/moderate and 6% as severe/critical. Patients were included if at least 2 samples were available. The antibody response was classified as either early-period or late-period (14 days-3 months and after 6 months, respectively) for IgG response whereas IgA response was tested on various time intervals, including as early as 4 days up to 3 months. Euroimmun Anti-SARS-CoV-2 IgG and IgA and Genscript SARS-CoV-2 Surrogate Virus Neutralization Kits were used for antibody detection. RESULTS: There was no difference between the early-period and late-period IgG positivity (P = 0.1). However, the median IgG levels were 11.98 in the early periods and 4.05 in the late periods, with a significance of P < 0.001. Although the decrease in IgG levels was significant in asymptomatic and mild/moderate cases (P < 0.008 and P < 0.001, respectively), the decrease in severe/critical cases was moderate (P = 0.285). The sensitivity of the IgG after 15 days was higher than 94%, and the sensitivity of IgA was 88% on days 8-15. CONCLUSION: SARS-CoV-2 IgG antibody levels decreased after 6 months. The decrease was moderate in severe/critical cases. Overall, 95.8% of the patients remained positive up to 9 months after infection. Although the IgA response may be useful early on, the IgG response is useful after 14 days.


Assuntos
Anticorpos Antivirais/biossíntese , COVID-19/imunologia , SARS-CoV-2/imunologia , Adolescente , Anticorpos Antivirais/imunologia , Formação de Anticorpos , Criança , Pré-Escolar , Feminino , Seguimentos , Humanos , Imunoglobulina A , Imunoglobulina G/biossíntese , Imunoglobulina G/imunologia , Lactente , Estudos Longitudinais , Masculino , Estudos Prospectivos
15.
BMC Med ; 19(1): 163, 2021 07 14.
Artigo em Inglês | MEDLINE | ID: mdl-34256745

RESUMO

BACKGROUND: Few studies had described the health consequences of patients with coronavirus disease 2019 (COVID-19) especially in those with severe infections after discharge from hospital. Moreover, no research had reported the health consequences in health care workers (HCWs) with COVID-19 after discharge. We aimed to investigate the health consequences in HCWs with severe COVID-19 after discharge from hospital in Hubei Province, China. METHODS: We conducted an ambidirectional cohort study in "Rehabilitation Care Project for Medical Staff Infected with COVID-19" in China. The participants were asked to complete three physical examinations (including the tests of functional fitness, antibodies to SARS-CoV-2 and immunological indicators) at 153.4 (143.3, 164.8), 244.3 (232.4, 259.1), and 329.4 (319.4, 339.3) days after discharge, respectively. Mann-Whitney U test, Kruskal-Wallis test, t test, one-way ANOVA, χ2, and Fisher's exact test were used to assess the variance between two or more groups where appropriate. RESULTS: Of 333 HCWs with severe COVID-19, the HCWs' median age was 36.0 (31.0, 43.0) years, 257 (77%) were female, and 191 (57%) were nurses. Our research found that 70.4% (114/162), 48.9% (67/137), and 29.6% (37/125) of the HCWs with severe COVID-19 were considered to have not recovered their functional fitness in the first, second, and third functional fitness tests, respectively. The HCWs showed improvement in muscle strength, flexibility, and agility/dynamic balance after discharge in follow-up visits. The seropositivity of IgM (17.0% vs. 6.6%) and median titres of IgM (3.0 vs. 1.4) and IgG (60.3 vs. 45.3) in the third physical examination was higher than that in the first physical examination. In the third physical examination, there still were 42.1% and 45.9% of the HCWs had elevated levels of IL-6 and TNF-α, and 11.9% and 6.3% of the HCWs had decreased relative numbers of CD3+ T cells and CD4+ T cells. CONCLUSION: The HCWs with severe COVID-19 showed improvement in functional fitness within 1 year after discharge, active intervention should be applied to help their recovery if necessary. It is of vital significance to continue monitoring the functional fitness, antibodies to SARS-CoV-2 and immunological indicators after 1 year of discharge from hospital in HCWs with severe COVID-19.


Assuntos
Anticorpos Antivirais/sangue , Teste Sorológico para COVID-19 , COVID-19 , Teste de Esforço , Pessoal de Saúde/estatística & dados numéricos , SARS-CoV-2/imunologia , Adulto , COVID-19/epidemiologia , COVID-19/imunologia , COVID-19/fisiopatologia , COVID-19/reabilitação , Teste Sorológico para COVID-19/métodos , Teste Sorológico para COVID-19/estatística & dados numéricos , China/epidemiologia , Teste de Esforço/métodos , Teste de Esforço/estatística & dados numéricos , Feminino , Seguimentos , Estado Funcional , Humanos , Interleucina-6/sangue , Masculino , Alta do Paciente/estatística & dados numéricos , Índice de Gravidade de Doença , Fator de Necrose Tumoral alfa/sangue
16.
Int J Nanomedicine ; 16: 4739-4753, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34267520

RESUMO

Background: Serological tests detecting severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) are widely used in seroprevalence studies and evaluating the efficacy of the vaccination program. Some of the widely used serological testing techniques are enzyme-linked immune-sorbent assay (ELISA), chemiluminescence immunoassay (CLIA), and lateral flow immunoassay (LFIA). However, these tests are plagued with low sensitivity or specificity, time-consuming, labor-intensive, and expensive. We developed a serological test implementing flow-through dot-blot assay (FT-DBA) for SARS-CoV-2 specific IgG detection, which provides enhanced sensitivity and specificity while being quick to perform and easy to use. Methods: SARS-CoV-2 antigens were immobilized on nitrocellulose membrane to capture human IgG, which was then detected with anti-human IgG conjugated gold nanoparticle (hIgG-AuNP). A total of 181 samples were analyzed in-house. Within which 35 were further evaluated in US FDA-approved CLIA Elecsys SARS-CoV-2 assay. The positive panel consisted of RT-qPCR positive samples from patients with both <14 days and >14 days from the onset of clinical symptoms. The negative panel contained samples collected from the pre-pandemic era dengue patients and healthy donors during the pandemic. Moreover, the sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of FT-DBA were evaluated against RT-qPCR positive sera. However, the overall efficacies were assessed with sera that seroconverted against either nucleocapsid (NCP) or receptor-binding domain (RBD). Results: In-house ELISA selected a total of 81 true seropositive and 100 seronegative samples. The sensitivity of samples with <14 days using FT-DBA was 94.7%, increasing to 100% for samples >14 days. The overall detection sensitivity and specificity were 98.8% and 98%, respectively, whereas the overall PPV and NPV were 99.6% and 99%. Moreover, comparative analysis between in-house ELISA assays and FT-DBA revealed clinical agreement of Cohen's Kappa value of 0.944. The FT-DBA showed sensitivity and specificity of 100% when compared with commercial CLIA kits. Conclusion: The assay can confirm past SARS-CoV-2 infection with high accuracy within 2 minutes compared to commercial CLIA or in-house ELISA. It can help track SARS-CoV-2 disease progression, population screening, and vaccination response. The ease of use of the assay without requiring any instruments while being semi-quantitative provides the avenue of its implementation in remote areas around the globe, where conventional serodiagnosis is not feasible.


Assuntos
Ouro/química , Immunoblotting/métodos , Imunoglobulina G/análise , Nanopartículas Metálicas/química , Nucleocapsídeo/análise , SARS-CoV-2/isolamento & purificação , Adulto , Anticorpos Antivirais/sangue , Humanos , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Masculino , Valor Preditivo dos Testes , SARS-CoV-2/imunologia , Sensibilidade e Especificidade , Estudos Soroepidemiológicos
17.
Front Immunol ; 12: 684014, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34194438

RESUMO

T cells play a fundamental role in the early control and clearance of many viral infections of the respiratory system. In SARS-CoV-2-infected individuals, lymphopenia with drastically reduced CD4+ and CD8+ T cells correlates with Coronavirus disease 2019 (COVID-19)-associated disease severity and mortality. In this study, we characterized cellular and humoral immune responses induced in patients with mild, severe and critical COVID-19. Peripheral blood mononuclear cells of 37 patients with mild, severe and critical COVID-19 and 10 healthy individuals were analyzed by IFNγ ELISpot and multi-color flow cytometry upon stimulation with peptide pools covering complete immunodominant SARS-CoV-2 matrix, nucleocapsid and spike proteins. In addition SARS-CoV-2 antibody levels, neutralization abilities and anaphylatoxin levels were evaluated by various commercially available ELISA platforms. Our data clearly demonstrates a significantly stronger induction of SARS-CoV-2 specific CD8+ T lymphocytes and higher IFNγ production in patients with mild compared to patients with severe or critical COVID-19. In all patients SARS-CoV-2-specific antibodies with similar neutralizing activity were detected, but highest titers of total IgGs were observed in critical patients. Finally, elevated anaphylatoxin C3a and C5a levels were identified in severe and critical COVID-19 patients probably caused by aberrant immune complex formation due to elevated antibody titers in these patients. Crucially, we provide a full picture of cellular and humoral immune responses of COVID-19 patients and prove that robust polyfunctional CD8+ T cell responses concomitant with low anaphylatoxin levels correlate with mild infections. In addition, our data indicates that high SARS-CoV-2 antibody titers are associated with severe disease progression.


Assuntos
Anafilatoxinas/metabolismo , Linfócitos T CD8-Positivos/imunologia , COVID-19/imunologia , SARS-CoV-2/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , COVID-19/fisiopatologia , Progressão da Doença , ELISPOT , Feminino , Citometria de Fluxo , Humanos , Imunidade Humoral , Interferon gama/sangue , Masculino , Pessoa de Meia-Idade , Gravidade do Paciente
18.
Int J Mol Sci ; 22(13)2021 Jun 23.
Artigo em Inglês | MEDLINE | ID: mdl-34201817

RESUMO

The usefulness of anti-inflammatory drugs as an adjunct therapy to improve outcomes in COVID-19 patients is intensely discussed in this paper. Willow bark (Salix cortex) has been used for centuries to relieve pain, inflammation, and fever. Its main active ingredient, salicin, is metabolized in the human body into salicylic acid, the precursor of the commonly used pain drug acetylsalicylic acid (ASA). Here, we report on the in vitro anti-inflammatory efficacy of two methanolic Salix extracts, standardized to phenolic compounds, in comparison to ASA in the context of a SARS-CoV-2 peptide challenge. Using SARS-CoV-2 peptide/IL-1ß- or LPS-activated human PBMCs and an inflammatory intestinal Caco-2/HT29-MTX co-culture, Salix extracts, and ASA concentration-dependently suppressed prostaglandin E2 (PGE2), a principal mediator of inflammation. The inhibition of COX-2 enzyme activity, but not protein expression was observed for ASA and one Salix extract. In activated PBMCs, the suppression of relevant cytokines (i.e., IL-6, IL-1ß, and IL-10) was seen for both Salix extracts. The anti-inflammatory capacity of Salix extracts was still retained after transepithelial passage and liver cell metabolism in an advanced co-culture model system consisting of intestinal Caco-2/HT29-MTX cells and differentiated hepatocyte-like HepaRG cells. Taken together, our in vitro data suggest that Salix extracts might present an additional anti-inflammatory treatment option in the context of SARS-CoV-2 peptides challenge; however, more confirmatory data are needed.


Assuntos
Anti-Inflamatórios/farmacologia , Aspirina/farmacologia , COVID-19/tratamento farmacológico , COVID-19/imunologia , Extratos Vegetais/farmacologia , Anti-Inflamatórios/química , Álcoois Benzílicos/metabolismo , COVID-19/virologia , Células CACO-2 , Ciclo-Oxigenase 2/efeitos dos fármacos , Citocinas/metabolismo , Dinoprostona/metabolismo , Glucosídeos/metabolismo , Células HT29 , Humanos , Inflamação , Leucócitos Mononucleares/efeitos dos fármacos , Lipopolissacarídeos/imunologia , Casca de Planta/química , Extratos Vegetais/química , SARS-CoV-2/imunologia
19.
Sci Immunol ; 6(61)2021 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-34210785

RESUMO

A central feature of the SARS-CoV-2 pandemic is that some individuals become severely ill or die, whereas others have only a mild disease course or are asymptomatic. Here we report development of an improved multimeric αß T cell staining reagent platform, with each maxi-ferritin "spheromer" displaying 12 peptide-MHC complexes. Spheromers stain specific T cells more efficiently than peptide-MHC tetramers and capture a broader portion of the sequence repertoire for a given peptide-MHC. Analyzing the response in unexposed individuals, we find that T cells recognizing peptides conserved amongst coronaviruses are more abundant and tend to have a "memory" phenotype, compared to those unique to SARS-CoV-2. Significantly, CD8+ T cells with these conserved specificities are much more abundant in COVID-19 patients with mild disease versus those with a more severe illness, suggesting a protective role.


Assuntos
Linfócitos T CD8-Positivos/imunologia , COVID-19/imunologia , Epitopos de Linfócito T/imunologia , SARS-CoV-2/imunologia , Índice de Gravidade de Doença , Linfócitos T CD8-Positivos/patologia , COVID-19/patologia , Feminino , Humanos , Masculino
20.
Viruses ; 13(6)2021 06 05.
Artigo em Inglês | MEDLINE | ID: mdl-34198852

RESUMO

Epithelial characteristics underlying the differential susceptibility of chronic asthma to SARS-CoV-2 (COVID-19) and other viral infections are currently unclear. By revisiting transcriptomic data from patients with Th2 low versus Th2 high asthma, as well as mild, moderate, and severe asthmatics, we characterized the changes in expression of human coronavirus and influenza viral entry genes relative to sex, airway location, and disease endotype. We found sexual dimorphism in the expression of SARS-CoV-2-related genes ACE2, TMPRSS2, TMPRSS4, and SLC6A19. ACE2 receptor downregulation occurred specifically in females in Th2 high asthma, while proteases broadly assisting coronavirus and influenza viral entry, TMPRSS2, and TMPRSS4, were highly upregulated in both sexes. Overall, changes in SARS-CoV-2-related gene expression were specific to the Th2 high molecular endotype of asthma and different by asthma severity and airway location. The downregulation of ACE2 (COVID-19, SARS) and ANPEP (HCoV-229E) viral receptors wascorrelated with loss of club and ciliated cells in Th2 high asthma. Meanwhile, the increase in DPP4 (MERS-CoV), ST3GAL4, and ST6GAL1 (influenza) was associated with increased goblet and basal activated cells. Overall, this study elucidates sex, airway location, disease endotype, and changes in epithelial heterogeneity as potential factors underlying asthmatic susceptibility, or lack thereof, to SARS-CoV-2.


Assuntos
Asma/imunologia , COVID-19/imunologia , Infecções por Coronavirus/imunologia , Células Epiteliais/virologia , Expressão Gênica , Interações entre Hospedeiro e Microrganismos , Influenza Humana/imunologia , Índice de Gravidade de Doença , Asma/genética , Asma/virologia , COVID-19/genética , Coronavirus Humano 229E/genética , Coronavirus Humano 229E/imunologia , Infecções por Coronavirus/genética , Células Epiteliais/classificação , Feminino , Perfilação da Expressão Gênica , Interações entre Hospedeiro e Microrganismos/genética , Interações entre Hospedeiro e Microrganismos/imunologia , Humanos , Influenza Humana/genética , Masculino , Coronavírus da Síndrome Respiratória do Oriente Médio/genética , Coronavírus da Síndrome Respiratória do Oriente Médio/imunologia , Orthomyxoviridae/genética , Orthomyxoviridae/imunologia , SARS-CoV-2/genética , SARS-CoV-2/imunologia , Caracteres Sexuais
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