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1.
EuropePMC; 2022.
Preprint in English | EuropePMC | ID: ppcovidwho-335620

ABSTRACT

SARS-CoV-2 is a highly contagious respiratory virus and the causative agent for COVID-19. The severity of disease varies from mildly symptomatic to lethal and shows an extraordinary correlation with increasing age, which represents the major risk factor for severe COVID-19 1 . However, the precise pathomechanisms leading to aggravated disease in the elderly are currently unknown. Delayed and insufficient antiviral immune responses early after infection as well as dysregulated and overshooting immunopathological processes late during disease were suggested as possible mechanisms. Here we show that the age-dependent increase of COVID-19 severity is caused by the disruption of a timely and well-coordinated innate and adaptive immune response due to impaired interferon (IFN) responses. To overcome the limitations of mechanistic studies in humans, we generated a mouse model for severe COVID-19 and compared the kinetics of the immune responses in adult and aged mice at different time points after infection. Aggravated disease in aged mice was characterized by a diminished IFN-γ response and excessive virus replication. Accordingly, adult IFN-γ receptor-deficient mice phenocopied the age-related disease severity and supplementation of IFN-γ reversed the increased disease susceptibility of aged mice. Mimicking impaired type I IFN immunity in adult and aged mice, a second major risk factor for severe COVID-19 2–4 , we found that therapeutic treatment with IFN-λ in adult and a combinatorial treatment with IFN-γ and IFN-λ in aged Ifnar1 -/- mice was highly efficient in protecting against severe disease. Our findings provide an explanation for the age-dependent disease severity of COVID-19 and clarify the nonredundant antiviral functions of type I, II and III IFNs during SARS-CoV-2 infection in an age-dependent manner. Based on our data, we suggest that highly vulnerable individuals combining both risk factors, advanced age and an impaired type I IFN immunity, may greatly benefit from immunotherapy combining IFN-γ and IFN-λ.

2.
Sci Adv ; 7(49): eabk0172, 2021 Dec 03.
Article in English | MEDLINE | ID: covidwho-1546430

ABSTRACT

Vaccines are instrumental and indispensable in the fight against the COVID-19 pandemic. Several recent SARS-CoV-2 variants are more transmissible and evade infection- or vaccine-induced protection. We constructed live attenuated vaccine candidates by large-scale recoding of the SARS-CoV-2 genome and showed that the lead candidate, designated sCPD9, protects Syrian hamsters from a challenge with ancestral virus. Here, we assessed immunogenicity and protective efficacy of sCPD9 in the Roborovski dwarf hamster, a nontransgenic rodent species that is highly susceptible to SARS-CoV-2 and severe COVID-19­like disease. We show that a single intranasal vaccination with sCPD9 elicited strong cross-neutralizing antibody responses against four current SARS-CoV-2 variants of concern, B.1.1.7 (Alpha), B.1.351 (Beta), B.1.1.28.1 (Gamma), and B.1.617.2 (Delta). The sCPD9 vaccine offered complete protection from COVID-19­like disease caused by the ancestral SARS-CoV-2 variant B.1 and the two variants of concern B.1.1.7 and B.1.351.

3.
Science ; 374(6571): 1099-1106, 2021 Nov 26.
Article in English | MEDLINE | ID: covidwho-1467657

ABSTRACT

Molecular virology tools are critical for basic studies of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and for developing new therapeutics. Experimental systems that do not rely on viruses capable of spread are needed for potential use in lower-containment settings. In this work, we use a yeast-based reverse genetics system to develop spike-deleted SARS-CoV-2 self-replicating RNAs. These noninfectious self-replicating RNAs, or replicons, can be trans-complemented with viral glycoproteins to generate replicon delivery particles for single-cycle delivery into a range of cell types. This SARS-CoV-2 replicon system represents a convenient and versatile platform for antiviral drug screening, neutralization assays, host factor validation, and viral variant characterization.


Subject(s)
RNA, Viral/genetics , Replicon/physiology , SARS-CoV-2/genetics , Animals , Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , Antiviral Agents/pharmacology , Cell Line , Humans , Interferons/pharmacology , Microbial Sensitivity Tests , Mutation , Plasmids , RNA, Viral/metabolism , Replicon/genetics , Reverse Genetics , SARS-CoV-2/drug effects , SARS-CoV-2/physiology , Saccharomyces cerevisiae/genetics , Spike Glycoprotein, Coronavirus/genetics , Viral Nonstructural Proteins/genetics , Viral Nonstructural Proteins/metabolism , Virion/genetics , Virion/physiology , Virus Replication
4.
Emerg Infect Dis ; 27(7): 1811-1820, 2021 07.
Article in English | MEDLINE | ID: covidwho-1278358

ABSTRACT

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has spread globally, and the number of worldwide cases continues to rise. The zoonotic origins of SARS-CoV-2 and its intermediate and potential spillback host reservoirs, besides humans, remain largely unknown. Because of ethical and experimental constraints and more important, to reduce and refine animal experimentation, we used our repository of well-differentiated airway epithelial cell (AEC) cultures from various domesticated and wildlife animal species to assess their susceptibility to SARS-CoV-2. We observed that SARS-CoV-2 replicated efficiently only in monkey and cat AEC culture models. Whole-genome sequencing of progeny viruses revealed no obvious signs of nucleotide transitions required for SARS-CoV-2 to productively infect monkey and cat AEC cultures. Our findings, together with previous reports of human-to-animal spillover events, warrant close surveillance to determine the potential role of cats, monkeys, and closely related species as spillback reservoirs for SARS-CoV-2.


Subject(s)
Animals, Wild , COVID-19 , Animals , Epithelial Cells , Humans , Respiratory System , SARS-CoV-2
5.
Nature ; 592(7852): 122-127, 2021 04.
Article in English | MEDLINE | ID: covidwho-1104508

ABSTRACT

During the evolution of SARS-CoV-2 in humans, a D614G substitution in the spike glycoprotein (S) has emerged; virus containing this substitution has become the predominant circulating variant in the COVID-19 pandemic1. However, whether the increasing prevalence of this variant reflects a fitness advantage that improves replication and/or transmission in humans or is merely due to founder effects remains unknown. Here we use isogenic SARS-CoV-2 variants to demonstrate that the variant that contains S(D614G) has enhanced binding to the human cell-surface receptor angiotensin-converting enzyme 2 (ACE2), increased replication in primary human bronchial and nasal airway epithelial cultures as well as in a human ACE2 knock-in mouse model, and markedly increased replication and transmissibility in hamster and ferret models of SARS-CoV-2 infection. Our data show that the D614G substitution in S results in subtle increases in binding and replication in vitro, and provides a real competitive advantage in vivo-particularly during the transmission bottleneck. Our data therefore provide an explanation for the global predominance of the variant that contains S(D614G) among the SARS-CoV-2 viruses that are currently circulating.


Subject(s)
COVID-19/transmission , COVID-19/virology , Mutation , SARS-CoV-2/genetics , SARS-CoV-2/physiology , Spike Glycoprotein, Coronavirus/genetics , Virus Replication/genetics , Angiotensin-Converting Enzyme 2/genetics , Angiotensin-Converting Enzyme 2/metabolism , Animals , Bronchi/cytology , Bronchi/virology , COVID-19/epidemiology , Cell Line , Cells, Cultured , Cricetinae , Disease Models, Animal , Epithelial Cells/virology , Female , Ferrets/virology , Founder Effect , Gene Knock-In Techniques , Genetic Fitness , Humans , Male , Mesocricetus , Mice , Nasal Mucosa/cytology , Nasal Mucosa/virology , Protein Binding , RNA, Viral/analysis , Receptors, Coronavirus/metabolism , SARS-CoV-2/metabolism , SARS-CoV-2/pathogenicity
7.
bioRxiv ; 2020 Oct 27.
Article in English | MEDLINE | ID: covidwho-915978

ABSTRACT

During the evolution of SARS-CoV-2 in humans a D614G substitution in the spike (S) protein emerged and became the predominant circulating variant (S-614G) of the COVID-19 pandemic 1 . However, whether the increasing prevalence of the S-614G variant represents a fitness advantage that improves replication and/or transmission in humans or is merely due to founder effects remains elusive. Here, we generated isogenic SARS-CoV-2 variants and demonstrate that the S-614G variant has (i) enhanced binding to human ACE2, (ii) increased replication in primary human bronchial and nasal airway epithelial cultures as well as in a novel human ACE2 knock-in mouse model, and (iii) markedly increased replication and transmissibility in hamster and ferret models of SARS-CoV-2 infection. Collectively, our data show that while the S-614G substitution results in subtle increases in binding and replication in vitro , it provides a real competitive advantage in vivo , particularly during the transmission bottle neck, providing an explanation for the global predominance of S-614G variant among the SARS-CoV-2 viruses currently circulating.

8.
Methods Mol Biol ; 2203: 167-184, 2020.
Article in English | MEDLINE | ID: covidwho-761352

ABSTRACT

The Escherichia coli and vaccinia virus-based reverse genetics systems have been widely applied for the manipulation and engineering of coronavirus genomes. These systems, however, present several limitations and are sometimes difficult to establish in a timely manner for (re-)emerging viruses. In this chapter, we present a new universal reverse genetics platform for the assembly and engineering of infectious full-length cDNAs using yeast-based transformation-associated recombination cloning. This novel assembly method not only results in stable coronavirus infectious full-length cDNAs cloned in the yeast Saccharomyces cerevisiae but also fosters and accelerates the manipulation of their genomes. Such a platform is widely applicable for the scientific community, as it requires no specific equipment and can be performed in a standard laboratory setting. The protocol described can be easily adapted to virtually all known or emerging coronaviruses, such as Middle East respiratory syndrome coronavirus (MERS-CoV).


Subject(s)
Coronavirus/genetics , DNA, Complementary/genetics , Genomics/methods , Saccharomyces cerevisiae/genetics , Animals , Cell Line , Coronavirus/pathogenicity , Homologous Recombination , Middle East Respiratory Syndrome Coronavirus/genetics , Middle East Respiratory Syndrome Coronavirus/pathogenicity
9.
Emerg Infect Dis ; 26(7): 1592-1595, 2020 07.
Article in English | MEDLINE | ID: covidwho-712728

ABSTRACT

Infection control instructions call for use of alcohol-based hand rub solutions to inactivate severe acute respiratory syndrome coronavirus 2. We determined the virucidal activity of World Health Organization-recommended hand rub formulations, at full strength and multiple dilutions, and of the active ingredients. All disinfectants demonstrated efficient virus inactivation.


Subject(s)
Alcohols/pharmacology , Betacoronavirus/drug effects , Coronavirus Infections/prevention & control , Disinfectants/pharmacology , Hand Disinfection/methods , Pandemics/prevention & control , Pneumonia, Viral/prevention & control , Virus Inactivation , COVID-19 , Humans , SARS-CoV-2 , World Health Organization
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