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Most studies of adaptive immunity to SARS-CoV-2 infection focus on peripheral blood, which may not fully reflect immune responses at the site of infection. Using samples from 110 children undergoing tonsillectomy and adenoidectomy during the COVID-19 pandemic, we identified 24 samples with evidence of previous SARS-CoV-2 infection, including neutralizing antibodies in serum and SARS-CoV-2-specific germinal center and memory B cells in the tonsils and adenoids. Single-cell B cell receptor (BCR) sequencing indicated virus-specific BCRs were class-switched and somatically hypermutated, with overlapping clones in the two tissues. Expanded T cell clonotypes were found in tonsils, adenoids and blood post-COVID-19, some with CDR3 sequences identical to previously reported SARS-CoV-2-reactive T cell receptors (TCRs). Pharyngeal tissues from COVID-19-convalescent children showed persistent expansion of germinal center and antiviral lymphocyte populations associated with interferon (IFN)-γ-type responses, particularly in the adenoids, and viral RNA in both tissues. Our results provide evidence for persistent tissue-specific immunity to SARS-CoV-2 in the upper respiratory tract of children after infection.
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COVID-19 , SARS-CoV-2 , Humanos , Niño , Pandemias , Inmunidad Adaptativa , Tonsila Palatina , Anticuerpos AntiviralesRESUMEN
There is limited understanding of the viral antibody fingerprint following severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection in children. Herein, SARS-CoV-2 proteome-wide immunoprofiling of children with mild/moderate or severe coronavirus disease 2019 (COVID-19) versus multisystem inflammatory syndrome in children versus hospitalized control patients revealed differential cytokine responses, IgM/IgG/IgA epitope diversity, antibody binding and avidity. Apart from spike and nucleocapsid, IgG/IgA recognized epitopes in nonstructural protein (NSP) 2, NSP3, NSP12-NSP14 and open reading frame (ORF) 3a-ORF9. Peptides representing epitopes in NSP12, ORF3a and ORF8 demonstrated SARS-CoV-2 serodiagnosis. Antibody-binding kinetics with 24 SARS-CoV-2 proteins revealed antibody parameters that distinguish children with mild/moderate versus severe COVID-19 or multisystem inflammatory syndrome in children. Antibody avidity to prefusion spike correlated with decreased illness severity and served as a clinical disease indicator. The fusion peptide and heptad repeat 2 region induced SARS-CoV-2-neutralizing antibodies in rabbits. Thus, we identified SARS-CoV-2 antibody signatures in children associated with disease severity and delineate promising serodiagnostic and virus neutralization targets. These findings might guide the design of serodiagnostic assays, prognostic algorithms, therapeutics and vaccines in this important but understudied population.
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Prueba Serológica para COVID-19/métodos , COVID-19/complicaciones , COVID-19/inmunología , SARS-CoV-2/inmunología , Síndrome de Respuesta Inflamatoria Sistémica/inmunología , Adolescente , Anticuerpos Neutralizantes/metabolismo , Anticuerpos Antivirales/metabolismo , COVID-19/diagnóstico , Niño , Preescolar , Progresión de la Enfermedad , Epítopos/metabolismo , Femenino , Hospitalización , Humanos , Inmunidad Humoral , Inmunoglobulina A/metabolismo , Inmunoglobulina G/metabolismo , Inmunoglobulina M/metabolismo , Masculino , Pronóstico , Proteoma , Índice de Severidad de la Enfermedad , Síndrome de Respuesta Inflamatoria Sistémica/diagnósticoRESUMEN
Acute viral infections can have durable functional impacts on the immune system long after recovery, but how they affect homeostatic immune states and responses to future perturbations remain poorly understood1-4. Here we use systems immunology approaches, including longitudinal multimodal single-cell analysis (surface proteins, transcriptome and V(D)J sequences) to comparatively assess baseline immune statuses and responses to influenza vaccination in 33 healthy individuals after recovery from mild, non-hospitalized COVID-19 (mean, 151 days after diagnosis) and 40 age- and sex-matched control individuals who had never had COVID-19. At the baseline and independent of time after COVID-19, recoverees had elevated T cell activation signatures and lower expression of innate immune genes including Toll-like receptors in monocytes. Male individuals who had recovered from COVID-19 had coordinately higher innate, influenza-specific plasmablast, and antibody responses after vaccination compared with healthy male individuals and female individuals who had recovered from COVID-19, in part because male recoverees had monocytes with higher IL-15 responses early after vaccination coupled with elevated prevaccination frequencies of 'virtual memory'-like CD8+ T cells poised to produce more IFNγ after IL-15 stimulation. Moreover, the expression of the repressed innate immune genes in monocytes increased by day 1 to day 28 after vaccination in recoverees, therefore moving towards the prevaccination baseline of the healthy control individuals. By contrast, these genes decreased on day 1 and returned to the baseline by day 28 in the control individuals. Our study reveals sex-dimorphic effects of previous mild COVID-19 and suggests that viral infections in humans can establish new immunological set-points that affect future immune responses in an antigen-agnostic manner.
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COVID-19 , Inmunidad Innata , Memoria Inmunológica , Vacunas contra la Influenza , Caracteres Sexuales , Linfocitos T , Vacunación , Femenino , Humanos , Masculino , Linfocitos T CD8-positivos/inmunología , COVID-19/inmunología , Vacunas contra la Influenza/inmunología , Gripe Humana/inmunología , Gripe Humana/prevención & control , Interleucina-15/inmunología , Receptores Toll-Like/inmunología , Linfocitos T/citología , Linfocitos T/inmunología , Monocitos , Inmunidad Innata/genética , Inmunidad Innata/inmunología , Análisis de la Célula Individual , Voluntarios SanosRESUMEN
Passive antibody immunotherapeutics directed against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are promising countermeasures for protection and treatment of coronavirus disease 2019 (COVID-19). SARS-CoV-2 variants of concern (VOCs) and variants of interest (VOIs) can impact the clinical efficacy of immunotherapeutics. A fully human polyclonal antibody immunotherapeutic purified from plasma of transchromosomic (Tc) bovines hyperimmunized with SARS-CoV-2 WA-1 spike (SAB-185) is being assessed for efficacy in a phase 2/3 clinical trial when different circulating SARS-CoV-2 variants predominated. We evaluated antibody binding, avidity maturation, and SARS-CoV-2 VOCs/VOIs virus-neutralizing capacity of convalescent plasma compared with different lots of SAB-185 and individual Tc bovine sera sequentially obtained after each vaccination against Alpha, Epsilon, Iota, Gamma, Beta, Kappa, and Delta variants. In contrast to convalescent plasma, sera and SAB-185 derived from hyperimmunized Tc bovines demonstrated higher antibody avidity and more potent cross-neutralizing activity of VOCs/VOIs. Thus, SAB-185 is a potential promising therapeutic candidate for the treatment of patients infected with SARS-CoV-2 variants.
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COVID-19 , SARS-CoV-2 , Animales , Anticuerpos Neutralizantes , Anticuerpos Antivirales , Afinidad de Anticuerpos , COVID-19/terapia , Bovinos , Humanos , Inmunización Pasiva , Inmunoglobulina G , Pruebas de Neutralización , Glicoproteína de la Espiga del Coronavirus , Sueroterapia para COVID-19RESUMEN
Convalescent plasma (CP) have been used for treatment of coronavirus disease 2019 (COVID-19), but their effectiveness varies significantly. Moreover, the impact of CP treatment on the composition of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antibodies in COVID-19 patients and antibody markers that differentiate between those who survive and those who succumb to the COVID-19 disease are not well understood. Herein, we performed longitudinal analysis of antibody profile on 115 sequential plasma samples from 16 hospitalized COVID-19 patients treated with either CP or standard of care, only half of them survived. Differential antibody kinetics was observed for antibody binding, immunoglobulin M/immunoglobulin G/immunoglobulin A (IgM/IgG/IgA) distribution, and affinity maturation in "survived" versus "fatal" COVID-19 patients. Surprisingly, CP treatment did not predict survival. Strikingly, marked decline in neutralization titers was observed in the fatal patients prior to death, and convalescent plasma treatment did not reverse this trend. Furthermore, irrespective of CP treatment, higher antibody affinity to the SARS-CoV-2 prefusion spike was associated with survival outcome. Additionally, sustained elevated IgA response was associated with fatal outcome in these COVID-19 patients. These findings propose that treatment of COVID-19 patients with convalescent plasma should be carefully targeted, and effectiveness of treatment may depend on the clinical and immunological status of COVID-19 patients, as well as the quality of the antibodies in the convalescent plasma.
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COVID-19 , SARS-CoV-2 , Anticuerpos Neutralizantes , Anticuerpos Antivirales , COVID-19/terapia , Humanos , Inmunización Pasiva , Sueroterapia para COVID-19RESUMEN
Our study demonstrates that children who developed severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccination-induced myocarditis and may not receive another vaccination, could be susceptible to infection with Omicron and emerging variants. We observed higher neutralizing antibody titers in myocarditis patients vs. healthy vaccinated children, but significantly lower neutralization titers against Omicron in both groups.
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COVID-19 , Miocarditis , Niño , Humanos , SARS-CoV-2 , Pruebas de Neutralización , Anticuerpos Antivirales , Miocarditis/etiología , COVID-19/prevención & control , Vacunación/efectos adversos , Anticuerpos NeutralizantesAsunto(s)
COVID-19 , SARS-CoV-2 , COVID-19/terapia , Humanos , Inmunización Pasiva , Inmunoglobulinas , Sueroterapia para COVID-19Asunto(s)
COVID-19/complicaciones , Leucemia Mieloide Aguda/complicaciones , Adenosina Monofosfato/análogos & derivados , Adenosina Monofosfato/uso terapéutico , Adolescente , Alanina/análogos & derivados , Alanina/uso terapéutico , Antimaláricos/uso terapéutico , Antineoplásicos/uso terapéutico , Antivirales/uso terapéutico , COVID-19/diagnóstico , COVID-19/inmunología , Manejo de la Enfermedad , Humanos , Hidroxicloroquina/uso terapéutico , Hidroxiurea/uso terapéutico , Leucemia Mieloide Aguda/tratamiento farmacológico , Leucemia Mieloide Aguda/inmunología , Masculino , SARS-CoV-2/efectos de los fármacos , SARS-CoV-2/inmunología , SARS-CoV-2/aislamiento & purificación , Adulto Joven , Tratamiento Farmacológico de COVID-19RESUMEN
There is minimal knowledge regarding the durability of neutralization capacity and level of binding antibody generated against the highly transmissible circulating Omicron subvariants following SARS-CoV-2 infection in children with acute COVID-19 and those diagnosed with multisystem inflammatory syndrome in children (MIS-C) in the absence of vaccination. In this study, SARS-CoV-2 neutralization titers against the ancestral strain (WA1) and Omicron sublineages were evaluated in unvaccinated children admitted for COVID-19 (n = 32) and MIS-C (n = 32) at the time of hospitalization (baseline) and at six to eight weeks post-discharge (follow-up) between 1 April 2020, and 1 September 2022. In addition, antibody binding to the spike receptor binding domain (RBD) from WA1, BA.1, BA.2.75, and BA.4/BA.5 was determined using surface plasmon resonance (SPR). At baseline, the children with MIS-C demonstrated two-fold to three-fold higher binding and neutralizing antibodies against ancestral WA1 compared to those with COVID-19. Importantly, in children with COVID-19, the virus neutralization titers against the Omicron subvariants at six to eight weeks post-discharge reached the same level as those with MIS-C had at baseline but were higher than titers at 6-8 weeks post-discharge for MIS-C cases. Cross-neutralization capacity against recently emerged Omicron BQ.1, BQ.1.1, and XBB.1 variants was very low in children with either COVID-19 or MIS-C at all time points. These findings about post-infection immunity in children with either COVID-19 or MIS-C suggest the need for vaccinations in children with prior COVID-19 or MIS-C to provide effective protection from emerging and circulating SARS-CoV-2 variants.
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Currently, only Palivizumab and Nirsevimab that target the respiratory syncytical virus (RSV) fusion protein are licensed for pre-treatment of infants. Glycoprotein-targeting antibodies may also provide protection against RSV. In this study, we generate monoclonal antibodies from mice immunized with G proteins from RSV-A2 and RSV-B1 strains. These monoclonal antibodies recognize six unique antigenic classes (G0-G5). None of the anti-G monoclonal antibodies neutralize RSV-A2 or RSV-B1 in vitro. In mice challenged with either RSV-A2 line 19 F or RSV-B1, one day after treatment with anti-G monoclonal antibodies, all monoclonal antibodies reduce lung pathology and significantly reduce lung infectious viral titers by more than 2 logs on day 5 post-RSV challenge. RSV dissemination in the lungs was variable and correlated with lung pathology. We demonstrate new cross-protective anti-G monoclonal antibodies targeting multiple sites including conformation-dependent class G0 MAb 77D2, CCD-specific class G1 MAb 40D8, and carboxy terminus of CCD class G5 MAb 7H11, to support development of G-targeting monoclonal antibodies against RSV.
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Infecciones por Virus Sincitial Respiratorio , Vacunas contra Virus Sincitial Respiratorio , Virus Sincitial Respiratorio Humano , Humanos , Ratones , Animales , Anticuerpos Monoclonales , Infecciones por Virus Sincitial Respiratorio/prevención & control , Anticuerpos Antivirales , Proteínas Virales de Fusión , Proteínas de Unión al GTP , Anticuerpos NeutralizantesRESUMEN
Emergence of highly transmissible Omicron subvariants led to increased SARS-CoV-2 infection and disease in children. However, minimal knowledge exists regarding the neutralization capacity against circulating Omicron BA.4/BA.5, BA.2.75, BQ.1, BQ.1.1 and XBB.1 subvariants following SARS-CoV-2 vaccination in children versus during acute or convalescent COVID-19, or versus multisystem inflammatory syndrome (MIS-C). Here, we evaluate virus-neutralizing capacity against SARS-CoV-2 variants in 151 age-stratified children ( <5, 5-11, 12-21 years old) hospitalized with acute severe COVID-19 or MIS-C or convalescent mild (outpatient) infection compared with 62 age-stratified vaccinated children. An age-associated effect on neutralizing antibodies is observed against SARS-CoV-2 following acute COVID-19 or vaccination. The primary series BNT162b2 mRNA vaccinated adolescents show higher vaccine-homologous WA-1 neutralizing titers compared with <12 years vaccinated children. Post-infection antibodies did not neutralize BQ.1, BQ.1.1 and XBB.1 subvariants. In contrast, monovalent mRNA vaccination induces more cross-neutralizing antibodies in young children <5 years against BQ.1, BQ.1.1 and XBB.1 variants compared with ≥5 years old children. Our study demonstrates that in children, infection and monovalent vaccination-induced neutralization activity is low against BQ.1, BQ.1.1 and XBB.1 variants. These findings suggest a need for improved SARS-CoV-2 vaccines to induce durable, more cross-reactive neutralizing antibodies to provide effective protection against emerging variants in children.
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COVID-19 , SARS-CoV-2 , Adolescente , Niño , Humanos , Preescolar , SARS-CoV-2/genética , COVID-19/prevención & control , Vacunas contra la COVID-19 , Vacuna BNT162 , Vacunación , Anticuerpos Neutralizantes , Anticuerpos ampliamente neutralizantes , ARN Mensajero , Anticuerpos AntiviralesRESUMEN
We recently reported that members of group 1 influenza A virus (IAV) containing H2, H5, H6, and H11 hemagglutinins (HAs) are resistant to lung surfactant protein D (SP-D). H3 viruses, members of group 2 IAV, have high affinity for SP-D, which depends on the presence of high-mannose glycans at glycosite N165 on the head of HA. The low affinity of SP-D for the group 1 viruses is due to the presence of complex glycans at an analogous glycosite on the head of HA, and replacement with high-mannose glycan at this site evoked strong interaction with SP-D. Thus, if members of group 1 IAV were to make the zoonotic leap to humans, the pathogenicity of such strains could be problematic since SP-D, as a first-line innate immunity factor in respiratory tissues, could be ineffective as demonstrated in vitro. Here, we extend these studies to group 2 H4 viruses that are representative of those with specificity for avian or swine sialyl receptors, i.e., those with receptor-binding sites with either Q226 and G228 for avian or recent Q226L and G228S mutations that facilitate swine receptor specificity. The latter have increased pathogenicity potential in humans due to a switch from avian sialylα2,3 to sialylα2,6 glycan receptor preference. A better understanding of the potential action of SP-D against these strains will provide important information regarding the pandemic risk of such strains. Our glycomics and in vitro analyses of four H4 HAs reveal SP-D-favorable glycosylation patterns. Therefore, susceptibilities to this first-line innate immunity defense respiratory surfactant against such H4 viruses are high and align with H3 HA glycosylation.
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There is limited knowledge on durability of neutralization capacity and antibody affinity maturation generated following two versus three doses of SARS-CoV-2 mRNA vaccines in naïve versus convalescent individuals (hybrid immunity) against the highly transmissible Omicron BA.1, BA.2 and BA.3 subvariants. Virus neutralization titers against the vaccine-homologous strain (WA1) and Omicron sublineages are measured in a pseudovirus neutralization assay (PsVNA). In addition, antibody binding and antibody affinity against spike proteins from WA1, BA.1, and BA.2 is determined using surface plasmon resonance (SPR). The convalescent individuals who after SARS-CoV-2 infection got vaccinated develop hybrid immunity that shows broader neutralization activity and cross-reactive antibody affinity maturation against the Omicron BA.1 and BA.2 after either second or third vaccination compared with naïve individuals. Neutralization activity correlates with antibody affinity against Omicron subvariants BA.1 and BA.2 spikes. Importantly, at four months post-third vaccination the neutralization activity and antibody affinity against the Omicron subvariants is maintained and trended higher for the individuals with hybrid immunity compared with naïve adults. These findings about hybrid immunity resulting in superior immune kinetics, breadth, and durable high affinity antibodies support the need for booster vaccinations to provide effective protection from emerging SARS-CoV-2 variants like the rapidly spreading Omicron subvariants.
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COVID-19 , SARS-CoV-2 , Adulto , Anticuerpos Neutralizantes , Anticuerpos Antivirales , Afinidad de Anticuerpos , COVID-19/prevención & control , Humanos , Pruebas de Neutralización , ARN Mensajero , SARS-CoV-2/genética , VacunaciónRESUMEN
The COVID-19 pandemic caused by SARS-CoV-2 infection has led to socio-economic shutdowns and the loss of over 5 million lives worldwide. There is a need for the identification of therapeutic targets to treat COVID-19. SARS-CoV-2 spike is a target of interest for the development of therapeutic targets. We developed a robust SARS-CoV-2 S spike expression and purification protocol from insect cells and studied four recombinant SARS-CoV-2 spike protein constructs based on the original SARS-CoV-2 sequence using a baculovirus expression system: a spike protein receptor-binding domain that includes the SD1 domain (RBD) coupled to a fluorescent tag (S-RBD-eGFP), spike ectodomain coupled to a fluorescent tag (S-Ecto-eGFP), spike ectodomain with six proline mutations and a foldon domain (S-Ecto-HexaPro(+F)), and spike ectodomain with six proline mutations without the foldon domain (S-Ecto-HexaPro(-F)). We tested the yield of purified protein expressed from the insect cell lines Spodoptera frugiperda (Sf9) and Trichoplusia ni (Tni) and compared it to previous research using mammalian cell lines to determine changes in protein yield. We demonstrated quick and inexpensive production of functional glycosylated spike protein of high purity capable of recognizing and binding to the angiotensin converting enzyme 2 (ACE2) receptor. To further confirm functionality, we demonstrate binding of eGFP fused construct of the spike ectodomain (S-Ecto-eGFP) to surface ACE2 receptors on lung epithelial cells by flow cytometry analysis and show that it can be decreased by means of receptor manipulation (blockade or downregulation).
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Enzima Convertidora de Angiotensina 2 , COVID-19 , Enzima Convertidora de Angiotensina 2/genética , Animales , Humanos , Insectos/metabolismo , Mamíferos , Pandemias , Peptidil-Dipeptidasa A/genética , Peptidil-Dipeptidasa A/metabolismo , Prolina , SARS-CoV-2/genética , Glicoproteína de la Espiga del Coronavirus/genéticaRESUMEN
BACKGROUND: Limited knowledge exists in post-partum women regarding durability of SARS-CoV-2 vaccine-induced antibody responses and their neutralising ability against SARS-CoV-2 variants of concern (VOC). METHODS: We elucidated longitudinal mRNA vaccination-induced antibody profiles of 13 post-partum and 13 non-post-partum women (control). FINDINGS: The antibody neutralisation titres against SARS-CoV-2 WA-1 strain were comparable between post-partum and non-post-partum women and these levels were sustained up to four months post-second vaccination in both groups. However, neutralisation titers declined against several VOCs, including Beta and Delta. Higher antibody binding was observed against SARS-CoV-2 receptor-binding domain (RBD) mutants with key VOC amino acids when tested with post-second vaccination plasma from post-partum women compared with controls. Importantly, post-vaccination plasma antibody affinity against VOCs RBDs was significantly higher in post-partum women compared with controls. INTERPRETATION: This study demonstrates that there is a differential vaccination-induced immune responses in post-partum women compared with non-post-partum women, which could help inform future vaccination strategies for these groups. FUNDING: The antibody characterisation work described in this manuscript was supported by FDA's Medical Countermeasures Initiative (MCMi) grant #OCET 2021-1565 to S.K and intramural FDA-CBER COVID-19 supplemental funds.
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Vacunas contra la COVID-19 , COVID-19 , Anticuerpos Antivirales , Afinidad de Anticuerpos , COVID-19/prevención & control , Femenino , Humanos , Inmunoglobulina G , Periodo Posparto , SARS-CoV-2/genética , Vacunas Sintéticas , Vacunas de ARNmRESUMEN
Viral infections can have profound and durable functional impacts on the immune system. There is an urgent need to characterize the long-term immune effects of SARS-CoV-2 infection given the persistence of symptoms in some individuals and the continued threat of novel variants. Here we use systems immunology, including longitudinal multimodal single cell analysis (surface proteins, transcriptome, and V(D)J sequences) from 33 previously healthy individuals after recovery from mild, non-hospitalized COVID-19 and 40 age- and sex-matched healthy controls with no history of COVID-19 to comparatively assess the post-infection immune status (mean: 151 days after diagnosis) and subsequent innate and adaptive responses to seasonal influenza vaccination. Identification of both sex-specific and -independent temporally stable changes, including signatures of T-cell activation and repression of innate defense/immune receptor genes (e.g., Toll-like receptors) in monocytes, suggest that mild COVID-19 can establish new post-recovery immunological set-points. COVID-19-recovered males had higher innate, influenza-specific plasmablast, and antibody responses after vaccination compared to healthy males and COVID-19-recovered females, partly attributable to elevated pre-vaccination frequencies of a GPR56 expressing CD8+ T-cell subset in male recoverees that are "poised" to produce higher levels of IFNγ upon inflammatory stimulation. Intriguingly, by day 1 post-vaccination in COVID-19-recovered subjects, the expression of the repressed genes in monocytes increased and moved towards the pre-vaccination baseline of healthy controls, suggesting that the acute inflammation induced by vaccination could partly reset the immune states established by mild COVID-19. Our study reveals sex-dimorphic immune imprints and in vivo functional impacts of mild COVID-19 in humans, suggesting that prior COVID-19, and possibly respiratory viral infections in general, could change future responses to vaccination and in turn, vaccines could help reset the immune system after COVID-19, both in an antigen-agnostic manner.
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Understanding early innate immune responses to coronavirus disease 2019 (COVID-19) is crucial to developing targeted therapies to mitigate disease severity. Severe acute respiratory syndrome coronavirus (SARS-CoV)-2 infection elicits interferon expression leading to transcription of IFN-stimulated genes (ISGs) to control viral replication and spread. SARS-CoV-2 infection also elicits NF-κB signaling which regulates inflammatory cytokine expression contributing to viral control and likely disease severity. Few studies have simultaneously characterized these two components of innate immunity to COVID-19. We designed a study to characterize the expression of interferon alpha-2 (IFNA2) and interferon beta-1 (IFNB1), both type-1 interferons (IFN-1), interferon-gamma (IFNG), a type-2 interferon (IFN-2), ISGs, and NF-κB response genes in the upper respiratory tract (URT) of patients with mild (outpatient) versus severe (hospitalized) COVID-19. Further, we characterized the weekly dynamics of these responses in the upper and lower respiratory tracts (LRTs) and blood of severe patients to evaluate for compartmental differences. We observed significantly increased ISG and NF-κB responses in the URT of mild compared with severe patients early during illness. This pattern was associated with increased IFNA2 and IFNG expression in the URT of mild patients, a trend toward increased IFNB1-expression and significantly increased STING/IRF3/cGAS expression in the URT of severe patients. Our by-week across-compartment analysis in severe patients revealed significantly higher ISG responses in the blood compared with the URT and LRT of these patients during the first week of illness, despite significantly lower expression of IFNA2, IFNB1, and IFNG in blood. NF-κB responses, however, were significantly elevated in the LRT compared with the URT and blood of severe patients during peak illness (week 2). Our data support that severe COVID-19 is associated with impaired interferon signaling in the URT during early illness and robust pro-inflammatory responses in the LRT during peak illness.
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SARS-CoV-2 infection triggers adaptive immune responses from both T and B cells. However, most studies focus on peripheral blood, which may not fully reflect immune responses in lymphoid tissues at the site of infection. To evaluate both local and systemic adaptive immune responses to SARS-CoV-2, we collected peripheral blood, tonsils, and adenoids from 110 children undergoing tonsillectomy/adenoidectomy during the COVID-19 pandemic and found 24 with evidence of prior SARS-CoV-2 infection, including detectable neutralizing antibodies against multiple viral variants. We identified SARS-CoV-2-specific germinal center (GC) and memory B cells; single cell BCR sequencing showed that these virus-specific B cells were class-switched and somatically hypermutated, with overlapping clones in the adenoids and tonsils. Oropharyngeal tissues from COVID-19-convalescent children showed persistent expansion of GC and anti-viral lymphocyte populations associated with an IFN-γ-type response, with particularly prominent changes in the adenoids, as well as evidence of persistent viral RNA in both tonsil and adenoid tissues of many participants. Our results show robust, tissue-specific adaptive immune responses to SARS-CoV-2 in the upper respiratory tract of children weeks to months after acute infection, providing evidence of persistent localized immunity to this respiratory virus.
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Neutralization capacity of antibodies against Omicron after a prior SARS-CoV-2 infection in children and adolescents is not well studied. Therefore, we evaluated virus-neutralizing capacity against SARS-CoV-2 Alpha, Beta, Gamma, Delta and Omicron variants by age-stratified analyses (<5, 5-11, 12-21 years) in 177 pediatric patients hospitalized with severe acute COVID-19, acute MIS-C, and in convalescent samples of outpatients with mild COVID-19 during 2020 and early 2021. Across all patients, less than 10% show neutralizing antibody titers against Omicron. Children <5 years of age hospitalized with severe acute COVID-19 have lower neutralizing antibodies to SARS-CoV-2 variants compared with patients >5 years of age. As expected, convalescent pediatric COVID-19 and MIS-C cohorts demonstrate higher neutralization titers than hospitalized acute COVID-19 patients. Overall, children and adolescents show some loss of cross-neutralization against all variants, with the most pronounced loss against Omicron. In contrast to SARS-CoV-2 infection, children vaccinated twice demonstrated higher titers against Alpha, Beta, Gamma, Delta and Omicron. These findings can influence transmission, re-infection and the clinical disease outcome from emerging SARS-CoV-2 variants and supports the need for vaccination in children.