Search

Despite intensive research since the emergence of SARS-CoV-2, it has remained unclear precisely which components of the early immune response protect against the development of severe COVID-19. Here, we perform a comprehensive immunogenetic and virologic analysis of nasopharyngeal and peripheral blood samples obtained during the acute phase of infection with SARS-CoV-2. We find that soluble and transcriptional markers of systemic inflammation peak during the first week after symptom onset and correlate directly with upper airways viral loads (UA-VLs), whereas the contemporaneous frequencies of circulating viral nucleocapsid (NC)-specific CD4+ and CD8+ T cells correlate inversely with various inflammatory markers and UA-VLs. In addition, we show that high frequencies of activated CD4+ and CD8+ T cells are present in acutely infected nasopharyngeal tissue, many of which express genes encoding various effector molecules, such as cytotoxic proteins and IFN-γ. The presence of IFNG mRNA-expressing CD4+ and CD8+ T cells in the infected epithelium is further linked with common patterns of gene expression among virus-susceptible target cells and better local control of SARS-CoV-2. Collectively, these results identify an immune correlate of protection against SARS-CoV-2, which could inform the development of more effective vaccines to combat the acute and chronic illnesses attributable to COVID-19.

Despite intensive research since the emergence of SARS-CoV-2, it has remained unclear precisely which components of the early immune response protect against the development of severe COVID-19. To address this issue, we performed a comprehensive immunogenetic and virologic analysis of nasopharyngeal and peripheral blood samples obtained during the acute phase of infection with SARS-CoV-2. We found that soluble and transcriptional markers of systemic inflammation peaked during the first week after symptom onset and correlated directly with the upper airways viral loads (UA-VLs), whereas the contemporaneous frequencies of circulating viral nucleocapsid (NC)-specific CD4+ and CD8+ T cells correlated inversely with various inflammatory markers and UA-VLs. In addition, we observed high frequencies of activated CD4+ and CD8+ T cells in acutely infected nasopharyngeal tissue, many of which expressed genes encoding various effector molecules, such as cytotoxic proteins and IFN-γ. The presence of functionally active T cells in the infected epithelium was further linked with common patterns of gene expression among virus-susceptible target cells and better local control of SARS-CoV-2. Collectively, these results identified an immune correlate of protection against SARS-CoV-2, which could inform the development of more effective vaccines to combat the acute and chronic illnesses attributable to COVID-19.

Risk factors for disease progression and severity of SARS-CoV-2 infections require an understanding of acute and long-term virological and immunological dynamics. Fifty-one RT-PCR positive COVID-19 outpatients were recruited between May and December 2020 in Munich, Germany, and followed up at multiple defined timepoints for up to one year. RT-PCR and viral culture were performed and seroresponses measured. Participants were classified applying the WHO clinical progression scale. Short symptom to test time (median 5.0 days; p = 0.0016) and high viral loads (VL; median maximum VL: 3∙108 copies/mL; p = 0.0015) were indicative for viral culture positivity. Participants with WHO grade 3 at baseline had significantly higher VLs compared to those with WHO 1 and 2 (p = 0.01). VLs dropped fast within 1 week of symptom onset. Maximum VLs were positively correlated with the magnitude of Ro-N-Ig seroresponse (p = 0.022). Our results describe the dynamics of VLs and antibodies to SARS-CoV-2 in mild to moderate cases that can support public health measures during the ongoing global pandemic.

A number of seroassays are available for SARS-CoV-2 testing; yet, head-to-head evaluations of different testing principles are limited, especially using raw values rather than categorical data. In addition, identifying correlates of protection is of utmost importance, and comparisons of available testing systems with functional assays, such as direct viral neutralisation, are needed.We analysed 6658 samples consisting of true-positives (n=193), true-negatives (n=1091), and specimens of unknown status (n=5374). For primary testing, we used Euroimmun-Anti-SARS-CoV-2-ELISA-IgA/IgG and Roche-Elecsys-Anti-SARS-CoV-2. Subsequently virus-neutralisation, GeneScriptcPass, VIRAMED-SARS-CoV-2-ViraChip, and Mikrogen-recomLine-SARS-CoV-2-IgG were applied for confirmatory testing. Statistical modelling generated optimised assay cut-off thresholds. Sensitivity of Euroimmun-anti-S1-IgA was 64.8%, specificity 93.3% (manufacturer’s cut-off); for Euroimmun-anti-S1-IgG, sensitivity was 77.2/79.8% (manufacturer’s/optimised cut-offs), specificity 98.0/97.8%; Roche-anti-N sensitivity was 85.5/88.6%, specificity 99.8/99.7%. In true-positives, mean and median Euroimmun-anti-S1-IgA and -IgG titres decreased 30/90 days after RT-PCR-positivity, Roche-anti-N titres decreased significantly later. Virus-neutralisation was 80.6% sensitive, 100.0% specific (≥1:5 dilution). Neutralisation surrogate tests (GeneScriptcPass, Mikrogen-recomLine-RBD) were >94.9% sensitive and >98.1% specific. Optimised cut-offs improved test performances of several tests. Confirmatory testing with virus-neutralisation might be complemented with GeneScriptcPassTM or recomLine-RBD for certain applications. Head-to-head comparisons given here aim to contribute to the refinement of testing strategies for individual and public health use.

SARS-CoV-2 Beta variant of concern (VOC) resists neutralization by major classes of antibodies from non-VOC COVID-19 patients and vaccinated individuals. Here, serum of Beta variant infected patients revealed reduced cross-neutralization of non-VOC virus. From these patients, we isolated Beta-specific and cross-reactive receptor-binding domain (RBD) antibodies. The Beta-specificity results from recruitment of novel VOC-specific clonotypes and accommodation of VOC-defining amino acids into a major non-VOC antibody class that is normally sensitive to these mutations. The Beta-elicited cross-reactive antibodies share genetic and structural features with non-VOC-elicited antibodies, including a public VH1-58 clonotype targeting the RBD ridge independent of VOC mutations. These findings advance our understanding of the antibody response to SARS-CoV-2 shaped by antigenic drift with implications for design of next-generation vaccines and therapeutics.One sentence summarySARS-CoV-2 Beta variant elicits lineage-specific antibodies and antibodies with neutralizing breadth against wild-type virus and VOCs.

BackgroundAdaptive immune responses to structural proteins of the virion play a crucial role in protection against coronavirus disease 2019 (COVID-19). We therefore studied T cell responses against multiple SARS-CoV-2 structural proteins in a large cohort using a simple, fast, and high-throughput approach.MethodsAn automated interferon gamma release assay (IGRA) for the Nucleocapsid (NC)-, Membrane (M)-, Spike-C-terminus (SCT)-, and N-terminus-protein (SNT)-specific T cell responses was performed using fresh whole blood from study subjects with convalescent, confirmed COVID-19 (n = 177, more than 200 days post infection), exposed household members (n = 145), and unexposed controls (n = 85). SARS-CoV-2-specific antibodies were assessed using Elecsys® Anti-SARS-CoV-2 (Ro-N-Ig) and Anti-SARS-CoV-2-ELISA (IgG) (EI-S1-IgG).Results156 of 177 (88%) previously PCR confirmed cases were still positive by Ro-N-Ig more than 200 days after infection. In T cells, most frequently the M-protein was targeted by 88% seropositive, PCR confirmed cases, followed by SCT (85%), NC (82%), and SNT (73%), whereas each of these antigens was recognized by less than 14% of non-exposed control subjects. Broad targeting of these structural virion proteins was characteristic of convalescent SARS-CoV-2 infection; 68% of all seropositive individuals targeted all four tested antigens. Indeed, anti-NC antibody titer correlated loosely, but significantly with the magnitude and breadth of the SARS-CoV-2-specific T cell response. Age, sex, and body mass index were comparable between the different groups.ConclusionSARS-CoV-2 seropositivity correlates with broad T cell reactivity of the structural virus proteins at 200 days after infection and beyond. The SARS-CoV-2-IGRA can facilitate large scale determination of SARS-CoV-2-specific T cell responses with high accuracy against multiple targets.

Humanized mice are critical for HIV-1 research, but humanized mice generated from cord blood are inefficient at mucosal HIV-1 transmission. Most mucosal HIV-1 transmission studies in mice require fetal tissue-engraftment, the use of which is highly restricted or prohibited. We present a fetal tissue-independent model called CD34T+ with enhanced human leukocyte levels in the blood and improved T cell homing to the gut-associated lymphoid tissue. CD34T+ mice are highly permissive to intra-rectal HIV-1 infection and also show normal env diversification in vivo despite high viral replication. Moreover, mucosal infection in CD34T+ mice can be prevented by infusion of broadly neutralizing antibodies. CD34T+ mice can be rapidly and easily generated using only cord blood cells and do not require any complicated surgical procedures for the humanization process. Therefore, CD34T+ mice provide a novel platform for mucosal HIV-1 transmission studies as well as rapid in vivo testing of novel prevention molecules against HIV-1.

The immune system of most SARS-CoV-2 infected individuals limits viral spread to the upper airways without pulmonary involvement. This prevents the development of pneumonic COVID-19. However, the protective immunological responses causative of successful viral containment in the upper airways remain unclear. Here, we combine longitudinal single-cell RNA sequencing, proteomic profiling, multidimensional flow cytometry, RNA-Seq of FACS-sorted leukocyte subsets and multiplex plasma interferon profiling to uncover temporally resolved protective immune signatures in non-pneumonic and ambulatory SARS-CoV-2 infected patients.We compare host responses in a high-risk patient population infected with SARS-CoV-2 but without pulmonary involvement to patients with COVID-19 pneumonia. Our data reveal a distinct immunological signature of successful viral containment, characterized by an early prominent interferon stimulated gene (ISG) upregulation across immune cell subsets. In addition, reduced cytotoxic potential of Natural Killer (NK) and T cells, as well as a monocyte phenotype with immune-modulatory potential are hallmarks of protective immunity. Temporal resolution across disease trajectories highlights ISG upregulation as particularly prominent early in the disease and confirms increased expression also in comparison to healthy controls.We validate this distinct temporal ISG signature by in-depth RNA-seq of FACS-sorted leukocyte subsets in a large prospective ambulatory SARS-CoV-2 infected cohort confirming early and robust ISG upregulation particularly in monocytes and T cells. In vitro experiments show that Stimulator of Interferon Genes (STING) agonist treatment of PBMCs recapitulates the identified protective immunological signature and might therefore offer a novel therapeutic approach in early disease, without being affected by previously described anti-interferon antibodies. In conclusion, our data demonstrate a protective ISG phenotype in patients with successful containment of SARS-CoV-2 infection without progression to COVID-19. This early protective interferon response might be exploited as a therapeutic approach and for disease course prediction. Funding: This study was supported by the Deutsche Herzstiftung e.V., Frankfurt a.M. [LN],Deutsche Forschungsgemeinschaft (DFG) SFB 914 (S.M. [B02 and Z01], K.S. [B02]), the DFG SFB 1123 (S.M. [B06], K.S. [A07]), M.J and R.Z [Z02]), the DFG FOR 2033 (S.M.), the DGF SFB1243 (W.E., L.E.W. [A14], the DGF EN 1093/2-1 (W.E., A.J.), the German Centre for Cardiovascular Research (DZHK) (Clinician Scientist Programme [L.N.], MHA 1.4VD [S.M.]), DZIF MD student programme (TI 07.003_Deak [F.D.]), FP7 program (project 260309, PRESTIGE [S.M.]), FoFoLe project 1015/1009 (L.N.), and the DFG Clinician Scientist Programme PRIME (413635475, K.P., R.K.). The work was also supported by the European Research Council (ERC 2018-ADG “IMMUNOTHROMBOSIS” [S.M.] and ERC- “T-MEMORE” [K.S.])The CORKUM cohort study was supported by LMUexcellent, funded by the Federal Ministry of Education and Research (BMBF) and the Free State of Bavaria under the Excellence Strategy of the Federal Government and the Lander.The Koco19-Immu Study is funded by Bavarian State Ministry of Science and the Arts, University Hospital, LMU Munich, Helmholtz Centre Munich, University of Bonn, University of Bielefeld, German Ministry for Education and Research (Project No.: 01KI20271). Conflict of Interest: The authors declare no conflict of interest. Ethical Approval: In accordance with the Declaration of Helsinki, and with the approval of the Ethics Committee of Ludwig-Maximilian University Munich, informed consent of the patients or their guardians was obtained. COVID-19 patients are part of the COVID-19 Registry of the LMU University Hospital Munich (CORKUM, WHO trial ID DRKS00021225). Pseudonymizeddata was used for analysis, the CORKUM and KocoImmu studies were approved by the ethics committee of LMUMunich (No: 20-245 & No: 20-371 respectively).

BackgroundSerosurveys are essential to understand SARS-CoV-2 exposure and enable population-level surveillance, but currently available tests need further in-depth evaluation. We aimed to identify testing-strategies by comparing seven seroassays in a population-based cohort.MethodsWe analysed 6,658 samples consisting of true-positives (n=193), true-negatives (n=1,091), and specimens of unknown status (n=5,374). For primary testing, we used Euroimmun-Anti-SARS-CoV-2-ELISA-IgA/IgG and Roche-Elecsys-Anti-SARS-CoV-2; and virus-neutralisation, GeneScript®cPass™, VIRAMED-SARS-CoV-2-ViraChip®, and Mikrogen-recomLine-SARS-CoV-2-IgG, including common-cold CoVs, for confirmatory testing. Statistical modelling generated optimised assay cut-off-thresholds.FindingsSensitivity of Euroimmun-anti-S1-IgA was 64.8%, specificity 93.3%; for Euroimmun-anti-S1-IgG, sensitivity was 77.2/79.8% (manufacturer’s/optimised cut-offs), specificity 98.0/97.8%; Roche-anti-N sensitivity was 85.5/88.6%, specificity 99.8/99.7%. In true-positives, mean and median titres remained stable for at least 90-120 days after RT-PCR-positivity. Of true-positives with positive RT-PCR (recomLine-RBD) were >94.9% sensitive, >98.1% specific. Seasonality had limited effects; cross-reactivity with common-cold CoVs 229E and NL63 in SARS-CoV-2 true-positives was significant.ConclusionOptimised cut-offs improved test performances of several tests. Non-reactive serology in true-positives was uncommon. For epidemiological purposes, confirmatory testing with virus-neutralisation may be replaced with GeneScript®cPass™ or recomLine-RBD. Head-to-head comparisons given here aim to contribute to the refinement of testing-strategies for individual and public health use.

Humanized mice have been critical for HIV-1 research, but are inefficient at mucosal HIV-1 transmission. We present a fetal tissue-independent model called CD34T+ with enhanced human leukocyte levels in the blood and improved T cell homing to the gut-associated lymphoid tissue. CD34T+ mice are highly permissive to intra-rectal HIV-1 infection, which can be prevented by infusion of broadly neutralizing antibodies. Therefore, CD34T+ mice provide a novel platform for mucosal HIV-1 transmission and prevention studies.

The transition metal manganese (Mn) is indispensable for photoautotrophic growth since photosystem II (PSII) employs an inorganic Mn4CaO5 cluster for water splitting. Here, we show that the Arabidopsis membrane protein CHLOROPLAST MANGANESE TRANSPORTER1 (CMT1) is involved in chloroplast Mn homeostasis. CMT1 is the closest homolog of the previously characterized thylakoid Mn transporter PHOTOSYNTHESIS-AFFECTED MUTANT71 (PAM71). In contrast to PAM71, CMT1 resides at the chloroplast envelope and is ubiquitously expressed. Nonetheless, like PAM71, the expression of CMT1 can also alleviate the Mn-sensitive phenotype of yeast mutant Δpmr1. The cmt1 mutant is severely suppressed in growth, chloroplast ultrastructure, and PSII activity owing to a decrease in the amounts of pigments and thylakoid membrane proteins. The importance of CMT1 for chloroplast Mn homeostasis is demonstrated by the significant reduction in chloroplast Mn concentrations in cmt1-1, which exhibited reduced Mn binding in PSII complexes. Moreover, CMT1 expression is downregulated in Mn-surplus conditions. The pam71 cmt1-1double mutant resembles the cmt1-1 single mutant rather than pam71 in most respects. Taken together, our results suggest that CMT1 mediates Mn2+ uptake into the chloroplast stroma, and that CMT1 and PAM71 function sequentially in Mn delivery to PSII across the chloroplast envelope and the thylakoid membrane.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Beta variant of concern (VOC) resists neutralization by major classes of antibodies from COVID-19 patients and vaccinated individuals. In this study, serum of Beta-infected patients revealed reduced cross-neutralization of wild-type virus. From these patients, we isolated Beta-specific and cross-reactive receptor-binding domain (RBD) antibodies. The Beta-specificity results from recruitment of VOC-specific clonotypes and accommodation of mutations present in Beta and Omicron into a major antibody class that is normally sensitive to these mutations. The Beta-elicited cross-reactive antibodies share genetic and structural features with wild type–elicited antibodies, including a public VH1-58 clonotype that targets the RBD ridge. These findings advance our understanding of the antibody response to SARS-CoV-2 shaped by antigenic drift, with implications for design of next-generation vaccines and therapeutics.