Your browser doesn't support javascript.
Show: 20 | 50 | 100
Results 1 - 4 de 4
Filter
3.
Nat Commun ; 12(1): 3431, 2021 06 08.
Article in English | MEDLINE | ID: covidwho-1262001

ABSTRACT

The current COVID-19 pandemic is caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). We demonstrate that despite the large size of the viral RNA genome (~30 kb), infectious full-length cDNA is readily assembled in vitro by a circular polymerase extension reaction (CPER) methodology without the need for technically demanding intermediate steps. Overlapping cDNA fragments are generated from viral RNA and assembled together with a linker fragment containing CMV promoter into a circular full-length viral cDNA in a single reaction. Transfection of the circular cDNA into mammalian cells results in the recovery of infectious SARS-CoV-2 virus that exhibits properties comparable to the parental virus in vitro and in vivo. CPER is also used to generate insect-specific Casuarina virus with ~20 kb genome and the human pathogens Ross River virus (Alphavirus) and Norovirus (Calicivirus), with the latter from a clinical sample. Additionally, reporter and mutant viruses are generated and employed to study virus replication and virus-receptor interactions.


Subject(s)
Reverse Genetics , SARS-CoV-2/genetics , Amino Acid Sequence , Animals , Base Sequence , Chlorocebus aethiops , Culicidae/virology , Furin/metabolism , Genome, Viral , HEK293 Cells , Humans , Mice , Mutation/genetics , NIH 3T3 Cells , Polymerase Chain Reaction , RAW 264.7 Cells , Receptors, Virus/metabolism , Vero Cells , Viral Proteins/chemistry , Virus Replication
4.
Mol Biol Evol ; 37(9): 2706-2710, 2020 09 01.
Article in English | MEDLINE | ID: covidwho-641314

ABSTRACT

Due to the scope and impact of the COVID-19 pandemic there exists a strong desire to understand where the SARS-CoV-2 virus came from and how it jumped species boundaries to humans. Molecular evolutionary analyses can trace viral origins by establishing relatedness and divergence times of viruses and identifying past selective pressures. However, we must uphold rigorous standards of inference and interpretation on this topic because of the ramifications of being wrong. Here, we dispute the conclusions of Xia (2020. Extreme genomic CpG deficiency in SARS-CoV-2 and evasion of host antiviral defense. Mol Biol Evol. doi:10.1093/molbev/masa095) that dogs are a likely intermediate host of a SARS-CoV-2 ancestor. We highlight major flaws in Xia's inference process and his analysis of CpG deficiencies, and conclude that there is no direct evidence for the role of dogs as intermediate hosts. Bats and pangolins currently have the greatest support as ancestral hosts of SARS-CoV-2, with the strong caveat that sampling of wildlife species for coronaviruses has been limited.


Subject(s)
Alphacoronavirus/genetics , Betacoronavirus/genetics , Coronavirus Infections/epidemiology , Genome, Viral , Pandemics , Pneumonia, Viral/epidemiology , Reassortant Viruses/genetics , Alphacoronavirus/classification , Alphacoronavirus/pathogenicity , Animals , Betacoronavirus/classification , Betacoronavirus/pathogenicity , Biological Evolution , COVID-19 , Chiroptera/virology , Coronavirus Infections/immunology , Coronavirus Infections/transmission , Coronavirus Infections/virology , CpG Islands , Dogs , Eutheria/virology , Humans , Immune Evasion/genetics , Pneumonia, Viral/immunology , Pneumonia, Viral/transmission , Pneumonia, Viral/virology , Protein Binding , RNA, Viral/genetics , RNA, Viral/metabolism , RNA-Binding Proteins/genetics , RNA-Binding Proteins/immunology , RNA-Binding Proteins/metabolism , Reassortant Viruses/classification , Reassortant Viruses/pathogenicity , SARS-CoV-2 , Virus Replication
SELECTION OF CITATIONS
SEARCH DETAIL