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BackgroundSARS-CoV-2 utilizes the ACE2 transmembrane peptidase as cellular entry receptor. However, whether SARS-CoV-2 in the alveolar compartment is strictly ACE2-dependent and to what extent virus-induced tissue damage and/or direct immune activation determines early pathogenesis is still elusive.MethodsSpectral microscopy, single-cell/-nucleus RNA sequencing or ACE2 ‘gain-of-function’ experiments were applied on infected human lung explants and adult stem cell-derived human lung organoids to correlate ACE2 and related host factors with SARS-CoV-2 tropism, propagation, virulence and immune activation compared to SARS-CoV, influenza and MERS-CoV. COVID-19 autopsy material was used to validate ex vivo results.ResultsWe provide evidence that alveolar ACE2 expression must be considered scarce, thereby limiting SARS-CoV-2 propagation and virus-induced tissue damage in the human alveolus. Instead, ex vivo infected human lungs and COVID-19 autopsy samples showed that alveolar macrophages were frequently positive for SARS-CoV-2. Single-cell/-nucleus transcriptomics further revealed non-productive virus uptake and a related inflammatory and anti-viral activation, especially in ‘inflammatory alveolar macrophages’, comparable to those induced by SARS-CoV and MERS-CoV but different from NL63 or influenza virus infection.ConclusionsCollectively, our findings indicate that severe lung injury in COVID-19 likely results from a macrophage triggered immune activation rather than direct viral damage of the alveolar compartment.

Severe acute respiratory syndrome (SARS)-CoV and SARS-CoV-2 infections are characterized by remarkable differences, including infectivity and case fatality rate. The underlying mechanisms are not well understood, illustrating major knowledge gaps of coronavirus biology. In this study, protein expression of the SARS-CoV- and SARS-CoV-2-infected human lung epithelial cell line Calu-3 was analyzed using data-independent acquisition–mass spectrometry. This resulted in a comprehensive map of infection-related proteome-wide expression changes in human cells covering the quantification of 7478 proteins across four time points. Most notably, the activation of interferon type-I response was observed, which is surprisingly absent in several proteome studies. The data reveal that SARS-CoV-2 triggers interferon-stimulated gene expression much stronger than SARS-CoV, which reflects the already described differences in interferon sensitivity. Potentially, this may be caused by the enhanced abundance of the viral M protein of SARS-CoV in comparison to SARS-CoV-2, which is a known inhibitor of type I interferon expression. This study expands the knowledge on the host response to SARS-CoV-2 infections on a global scale using an infection model, which seems to be well suited to analyze the innate immunity.

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Temperate bacteriophages possess a genetic switch which regulates the lytic and lysogenic cycle. The genomes of the enterobacterial telomere phages N15, PY54 and fKO2 harbor a primary immunity region (immB) comprising genes for the prophage repressor, the lytic repressor and a putative antiterminator, similar to CI, Cro and Q of lambda, respectively. Moreover, N15 and fKO2 contain 3 related operator (OR) sites between cI and cro, while only one site (O R 3) has been detected in PY54. Marine telomere phages possess a putative cI gene but not a cro-like gene. Instead, a gene is located at the position of cro, whose product shows some similarity to the PY54 ORF42 product, the function of which is unknown. We have determined the transcription start sites of the predicted repressor genes of N15, PY54, fKO2 and of the marine telomere phage VP58.5. The influence of the genes on phage propagation was analyzed in E. coli, Y. enterocolitica and V. parahaemolyticus. We show that the repressors and antiterminators of N15, fKO2 and PY54 exerted their predicted activities. However, while the proteins of both N15 and fKO2 affected lysis and lysogeny by N15, they did not affect PY54 propagation. On the other hand, the respective PY54 proteins exclusively influenced the propagation of this phage. The immB region of VP58.5 contains 2 genes that revealed prophage repressor activity, while a lytic repressor gene could not be identified. The results indicate an unexpected diversity of the growth regulation mechanisms in these temperate phages.

Middle East respiratory syndrome coronavirus (MERS-CoV) is enzootic in dromedary camels across the Middle East and Africa. Virus-induced pneumonia in humans results from animal contact, with a potential for limited onward transmission. Phenotypic changes have been suspected after a novel recombinant clade (lineage 5) caused large nosocomial outbreaks in Saudi Arabia and South Korea in 2016. However, there has been no functional assessment. Here we perform a comprehensive in vitro and ex vivo comparison of viruses from parental and recombinant virus lineages (lineage 3, n = 7; lineage 4, n = 8; lineage 5, n = 9 viruses) from Saudi Arabia, isolated immediately before and after the shift toward lineage 5. Replication of lineage 5 viruses is significantly increased. Transcriptional profiling finds reduced induction of immune genes IFNB1, CCL5, and IFNL1 in lung cells infected with lineage 5 strains. Phenotypic differences may be determined by IFN antagonism based on experiments using IFN receptor knock out and signaling inhibition. Additionally, lineage 5 is more resilient against IFN pre-treatment of Calu-3 cells (ca. 10-fold difference in replication). This phenotypic change associated with lineage 5 has remained undiscovered by viral sequence surveillance, but may be a relevant indicator of pandemic potential.

SARS-CoV-2 and its emerging variants of concern remain a major threat for global health. Here we introduce an infection model based upon polarized human Alveolar Epithelial Lentivirus immortalized (hAELVi) cells grown at the air–liquid interface to estimate replication and epidemic potential of respiratory viruses in the human lower respiratory tract. hAELVI cultures are highly permissive for different human coronaviruses and seasonal influenza A virus and upregulate various mediators following virus infection. Our analysis revealed a significantly reduced capacity of SARS-CoV-2 Omicron BA.1 and BA.2 variants to propagate in this human model compared to earlier D614G and Delta variants, which extends early risk assessments from epidemiological and animal studies suggesting a reduced pathogenicity of Omicron.

Summary Background The mRNA vaccine BNT162b2 (Comirnaty, BioNTech/Pfizer) and the vaccine candidate CVnCoV (Curevac) each encode a stabilized spike protein of SARS‐CoV2 as antigen but differ with respect to the nature of the mRNA (modified versus unmodified nucleotides) and the mRNA amount (30 μg versus 12 μg RNA). This study characterizes antisera elicited by these two vaccines in comparison to convalescent sera. Methods Sera from BNT162b2 vaccinated healthcare workers, and sera from participants of a phase I trial vaccinated with 2, 4, 6, 8, or 12 μg CVnCoV and convalescent sera from hospitalized patients were analyzed by ELISA, neutralization tests, surface plasmon resonance (SPR), and peptide arrays. Results BNT162b2‐elicited sera and convalescent sera have a higher titer of spike‐RBD‐specific antibodies and neutralizing antibodies as compared to the CVnCoV‐elicited sera. For all analyzed sera a reduction in binding and neutralizing antibodies was found for the lineage B.1.351 variant of concern. SPR analyses revealed that the CVnCoV‐elicited sera have a lower fraction of slow‐dissociating antibodies. Accordingly, the CVnCoV sera almost fail to compete with the spike‐ACE2 interaction. The significance of common VOC mutations K417N, E484K, or N501Y focused on linear epitopes was analyzed using a peptide array approach. The peptide arrays showed a strong difference between convalescent sera and vaccine‐elicited sera. Specifically, the linear epitope at position N501 was affected by the mutation and elucidates the escape of viral variants to antibodies against this linear epitope. Conclusion These data reveal differences in titer, neutralizing capacity, and affinity of the antibodies between BNT162b2‐ and CVnCoV‐elicited sera, which could contribute to the apparent differences in vaccine efficacy.