Your browser doesn't support javascript.
Show: 20 | 50 | 100
Results 1 - 8 de 8
Filter
2.
Innovation (Camb) ; 3(2): 100221, 2022 Mar 29.
Article in English | MEDLINE | ID: covidwho-1713028

ABSTRACT

The highly pathogenic and readily transmissible SARS-CoV-2 has caused a global coronavirus pandemic, urgently requiring effective countermeasures against its rapid expansion. All available vaccine platforms are being used to generate safe and effective COVID-19 vaccines. Here, we generated a live-attenuated candidate vaccine strain by serial passaging of a SARS-CoV-2 clinical isolate in Vero cells. Deep sequencing revealed the dynamic adaptation of SARS-CoV-2 in Vero cells, resulting in a stable clone with a deletion of seven amino acids (N679SPRRAR685) at the S1/S2 junction of the S protein (named VAS5). VAS5 showed significant attenuation of replication in multiple human cell lines, human airway epithelium organoids, and hACE2 mice. Viral fitness competition assays demonstrated that VAS5 showed specific tropism to Vero cells but decreased fitness in human cells compared with the parental virus. More importantly, a single intranasal injection of VAS5 elicited a high level of neutralizing antibodies and prevented SARS-CoV-2 infection in mice as well as close-contact transmission in golden Syrian hamsters. Structural and biochemical analysis revealed a stable and locked prefusion conformation of the S trimer of VAS5, which most resembles SARS-CoV-2-3Q-2P, an advanced vaccine immunogen (NVAX-CoV2373). Further systematic antigenic profiling and immunogenicity validation confirmed that the VAS5 S trimer presents an enhanced antigenic mimic of the wild-type S trimer. Our results not only provide a potent live-attenuated vaccine candidate against COVID-19 but also clarify the molecular and structural basis for the highly attenuated and super immunogenic phenotype of VAS5.

3.
Nat Commun ; 12(1): 5654, 2021 09 27.
Article in English | MEDLINE | ID: covidwho-1440471

ABSTRACT

There is an urgent need for animal models to study SARS-CoV-2 pathogenicity. Here, we generate and characterize a novel mouse-adapted SARS-CoV-2 strain, MASCp36, that causes severe respiratory symptoms, and mortality. Our model exhibits age- and gender-related mortality akin to severe COVID-19. Deep sequencing identified three amino acid substitutions, N501Y, Q493H, and K417N, at the receptor binding domain (RBD) of MASCp36, during in vivo passaging. All three RBD mutations significantly enhance binding affinity to its endogenous receptor, ACE2. Cryo-electron microscopy analysis of human ACE2 (hACE2), or mouse ACE2 (mACE2), in complex with the RBD of MASCp36, at 3.1 to 3.7 Å resolution, reveals the molecular basis for the receptor-binding switch. N501Y and Q493H enhance the binding affinity to hACE2, whereas triple mutations at N501Y/Q493H/K417N decrease affinity and reduce infectivity of MASCp36. Our study provides a platform for studying SARS-CoV-2 pathogenesis, and unveils the molecular mechanism for its rapid adaptation and evolution.


Subject(s)
COVID-19/diagnosis , SARS-CoV-2/pathogenicity , Spike Glycoprotein, Coronavirus/metabolism , Amino Acid Substitution , Angiotensin-Converting Enzyme 2/metabolism , Animals , Binding Sites/genetics , COVID-19/mortality , COVID-19/virology , Disease Models, Animal , Female , Humans , Male , Mice , Protein Binding/genetics , Protein Domains/genetics , SARS-CoV-2/genetics , Severity of Illness Index , Spike Glycoprotein, Coronavirus/genetics
6.
Science ; 369(6511): 1603-1607, 2020 09 25.
Article in English | MEDLINE | ID: covidwho-690532

ABSTRACT

The ongoing coronavirus disease 2019 (COVID-19) pandemic has prioritized the development of small-animal models for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). We adapted a clinical isolate of SARS-CoV-2 by serial passaging in the respiratory tract of aged BALB/c mice. The resulting mouse-adapted strain at passage 6 (called MASCp6) showed increased infectivity in mouse lung and led to interstitial pneumonia and inflammatory responses in both young and aged mice after intranasal inoculation. Deep sequencing revealed a panel of adaptive mutations potentially associated with the increased virulence. In particular, the N501Y mutation is located at the receptor binding domain (RBD) of the spike protein. The protective efficacy of a recombinant RBD vaccine candidate was validated by using this model. Thus, this mouse-adapted strain and associated challenge model should be of value in evaluating vaccines and antivirals against SARS-CoV-2.


Subject(s)
Betacoronavirus/immunology , Coronavirus Infections/prevention & control , Disease Models, Animal , Mice , Pandemics/prevention & control , Pneumonia, Viral/prevention & control , Viral Vaccines/immunology , Administration, Intranasal , Angiotensin-Converting Enzyme 2 , Animals , Betacoronavirus/genetics , Betacoronavirus/pathogenicity , COVID-19 , COVID-19 Vaccines , Coronavirus Infections/immunology , Female , High-Throughput Nucleotide Sequencing , Humans , Immunogenicity, Vaccine , Lung/virology , Lung Diseases, Interstitial/virology , Mice, Inbred BALB C , Mice, Transgenic , Mutation , Peptidyl-Dipeptidase A/genetics , SARS-CoV-2 , Spike Glycoprotein, Coronavirus/genetics , Vaccines, Synthetic/administration & dosage , Vaccines, Synthetic/immunology , Viral Vaccines/administration & dosage , Virulence/genetics
7.
Cell ; 182(5): 1271-1283.e16, 2020 09 03.
Article in English | MEDLINE | ID: covidwho-666099

ABSTRACT

There is an urgent need for vaccines against coronavirus disease 2019 (COVID-19) because of the ongoing SARS-CoV-2 pandemic. Among all approaches, a messenger RNA (mRNA)-based vaccine has emerged as a rapid and versatile platform to quickly respond to this challenge. Here, we developed a lipid nanoparticle-encapsulated mRNA (mRNA-LNP) encoding the receptor binding domain (RBD) of SARS-CoV-2 as a vaccine candidate (called ARCoV). Intramuscular immunization of ARCoV mRNA-LNP elicited robust neutralizing antibodies against SARS-CoV-2 as well as a Th1-biased cellular response in mice and non-human primates. Two doses of ARCoV immunization in mice conferred complete protection against the challenge of a SARS-CoV-2 mouse-adapted strain. Additionally, ARCoV is manufactured as a liquid formulation and can be stored at room temperature for at least 1 week. ARCoV is currently being evaluated in phase 1 clinical trials.


Subject(s)
RNA, Messenger/genetics , RNA, Viral/genetics , Vaccines, Synthetic/immunology , Viral Vaccines/immunology , Animals , Antibodies, Neutralizing/immunology , Binding Sites , COVID-19 Vaccines , Chlorocebus aethiops , Coronavirus Infections/genetics , Coronavirus Infections/immunology , Coronavirus Infections/prevention & control , Female , HEK293 Cells , HeLa Cells , Humans , Immunogenicity, Vaccine , Injections, Intramuscular , Macaca fascicularis , Male , Mice , Mice, Inbred ICR , Nanoparticles/chemistry , RNA, Messenger/metabolism , RNA, Viral/metabolism , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/metabolism , Th1 Cells/immunology , Vaccine Potency , Vaccines, Synthetic/administration & dosage , Vaccines, Synthetic/genetics , Vero Cells , Viral Vaccines/administration & dosage , Viral Vaccines/genetics
8.
Chin. J. Microbiol. Immunol. ; 2(40): 103-109, 20200229.
Article in Chinese | WHO COVID, ELSEVIER | ID: covidwho-23278

ABSTRACT

Objective: To study the effects of different pre-sequencing sample processing modes on the results of whole genome sequencing with high-throughput sequencing (HTS) by taking the largest RNA virus (human coronavirus, HCoV) as the representative. Methods: Cell-cultured human coronavirus HCoV-OC43 strains were used as the representative samples and divided into different groups based on pre-sequencing processing modes as follows: untreated group, DNase and RNase treatment before nucleic acid extraction group, DNase treatment after nucleic acid extraction group, and DNase and RNase treatment before nucleic acid extraction and DNase treatment after nucleic acid extraction group. Nucleic acid samples of each group were analyzed by direct RNA sequencing (without amplification) and DNA sequencing after sequence independent single primer amplification (SISPA), respectively. Results: No significant difference in viral genome coverage rates was observed between different groups. The highest genome coverage and sequencing accuracy were obtained in DNase treatment after nucleic acid extraction group by direct RNA sequencing, and the ratio of viral reads and the sequencing depth of each locus were effectively improved by SISPA amplification. Conclusions: This study provided an optimized technical strategy for whole genome sequencing of RNA viruses such as coronavirus.

SELECTION OF CITATIONS
SEARCH DETAIL