A persistently replicating SARS-CoV-2 variant derived from an asymptomatic individual.
Amino Acid Substitution
Animals
Betacoronavirus
/ genetics
COVID-19
Chlorocebus aethiops
Coronavirus Infections
/ epidemiology
Cytopathogenic Effect, Viral
/ genetics
Genetic Variation
Genome, Viral
Humans
Italy
/ epidemiology
Mutation
Pandemics
Phylogeny
Pneumonia, Viral
/ epidemiology
Polymorphism, Single Nucleotide
SARS-CoV-2
Translational Research, Biomedical
Vero Cells
Viral Proteins
/ genetics
Virus Cultivation
/ methods
Virus Replication
/ genetics
Whole Genome Sequencing
Asymptomatic infection
COVID-19 epidemic
Genetic variation
SARS-CoV-2
Virus isolate
Virus persistence
Journal
Journal of translational medicine
ISSN: 1479-5876
Titre abrégé: J Transl Med
Pays: England
ID NLM: 101190741
Informations de publication
Date de publication:
23 09 2020
23 09 2020
Historique:
received:
04
07
2020
accepted:
15
09
2020
entrez:
24
9
2020
pubmed:
25
9
2020
medline:
3
10
2020
Statut:
epublish
Résumé
Since the first outbreak of SARS-CoV-2, the clinical characteristics of the Coronavirus Disease 2019 (COVID-19) have been progressively changed. Data reporting a viral intra-host and inter-host evolution favouring the appearance of mild SARS-CoV-2 strains are since being accumulating. To better understand the evolution of SARS-CoV-2 pathogenicity and its adaptation to the host, it is therefore crucial to investigate the genetic and phenotypic characteristics of SARS-CoV-2 strains circulating lately in the epidemic. Nasopharyngeal swabs have been analyzed for viral load in the early (March 2020) and late (May 2020) phases of epidemic in Brescia, Italy. Isolation of SARS-CoV-2 from 2 high viral load specimens identified on March 9 (AP66) and on May 8 (GZ69) was performed on Vero E6 cells. Amount of virus released was assessed by quantitative PCR. Genotypic characterization of AP66 and GZ69 was performed by next generation sequencing followed by an in-depth in silico analysis of nucleotide mutations. The SARS-CoV-2 GZ69 strain, isolated in May from an asymptomatic healthcare worker, showed an unprecedented capability of replication in Vero E6 cells in the absence of any evident cytopathic effect. Vero E6 subculturing, up to passage 4, showed that SARS-CoV-2 GZ69 infection was as productive as the one sustained by the cytopathic strain AP66. Whole genome sequencing of the persistently replicating SARS-CoV-2 GZ69 has shown that this strain differs from the early AP66 variant in 9 nucleotide positions (C2939T; C3828T; G21784T; T21846C; T24631C; G28881A; G28882A; G28883C; G29810T) which lead to 6 non-synonymous substitutions spanning on ORF1ab (P892S; S1188L), S (K74N; I95T) and N (R203K, G204R) proteins. Identification of the peculiar SARS-CoV-2 GZ69 strain in the late Italian epidemic highlights the need to better characterize viral variants circulating among asymptomatic or paucisymptomatic individuals. The current approach could unravel the ways for future studies aimed at analyzing the selection process which favours viral mutations in the human host.
Sections du résumé
BACKGROUND
Since the first outbreak of SARS-CoV-2, the clinical characteristics of the Coronavirus Disease 2019 (COVID-19) have been progressively changed. Data reporting a viral intra-host and inter-host evolution favouring the appearance of mild SARS-CoV-2 strains are since being accumulating. To better understand the evolution of SARS-CoV-2 pathogenicity and its adaptation to the host, it is therefore crucial to investigate the genetic and phenotypic characteristics of SARS-CoV-2 strains circulating lately in the epidemic.
METHODS
Nasopharyngeal swabs have been analyzed for viral load in the early (March 2020) and late (May 2020) phases of epidemic in Brescia, Italy. Isolation of SARS-CoV-2 from 2 high viral load specimens identified on March 9 (AP66) and on May 8 (GZ69) was performed on Vero E6 cells. Amount of virus released was assessed by quantitative PCR. Genotypic characterization of AP66 and GZ69 was performed by next generation sequencing followed by an in-depth in silico analysis of nucleotide mutations.
RESULTS
The SARS-CoV-2 GZ69 strain, isolated in May from an asymptomatic healthcare worker, showed an unprecedented capability of replication in Vero E6 cells in the absence of any evident cytopathic effect. Vero E6 subculturing, up to passage 4, showed that SARS-CoV-2 GZ69 infection was as productive as the one sustained by the cytopathic strain AP66. Whole genome sequencing of the persistently replicating SARS-CoV-2 GZ69 has shown that this strain differs from the early AP66 variant in 9 nucleotide positions (C2939T; C3828T; G21784T; T21846C; T24631C; G28881A; G28882A; G28883C; G29810T) which lead to 6 non-synonymous substitutions spanning on ORF1ab (P892S; S1188L), S (K74N; I95T) and N (R203K, G204R) proteins.
CONCLUSIONS
Identification of the peculiar SARS-CoV-2 GZ69 strain in the late Italian epidemic highlights the need to better characterize viral variants circulating among asymptomatic or paucisymptomatic individuals. The current approach could unravel the ways for future studies aimed at analyzing the selection process which favours viral mutations in the human host.
Identifiants
pubmed: 32967693
doi: 10.1186/s12967-020-02535-1
pii: 10.1186/s12967-020-02535-1
pmc: PMC7509824
doi:
Substances chimiques
Viral Proteins
0
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
362Références
Infect Genet Evol. 2020 Sep;83:104351
pubmed: 32387564
Antiviral Res. 2020 Jul;179:104811
pubmed: 32360182
J Mol Biol. 2003 Aug 29;331(5):991-1004
pubmed: 12927536
Front Cell Dev Biol. 2020 May 22;8:410
pubmed: 32574318
Bioinformatics. 2014 Aug 1;30(15):2114-20
pubmed: 24695404
BMJ. 2020 Feb 19;368:m606
pubmed: 32075786
Virology. 2018 Nov;524:78-89
pubmed: 30165309
J Virol. 2005 Sep;79(17):11476-86
pubmed: 16103198
Nat Methods. 2012 Mar 04;9(4):357-9
pubmed: 22388286
Viruses. 2020 Jul 24;12(8):
pubmed: 32722343
Mol Biol Evol. 2015 Jan;32(1):268-74
pubmed: 25371430
Curr Opin Infect Dis. 2001 Jun;14(3):251-6
pubmed: 11964840
J Virol. 2015 Feb;89(3):1523-36
pubmed: 25428866
Clin Chem Lab Med. 2020 Jun 29;58(9):1573-1577
pubmed: 32598306
Genome Biol. 2019 Nov 28;20(1):257
pubmed: 31779668
JAMA. 2020 Apr 7;323(13):1239-1242
pubmed: 32091533
J Virol. 2008 Jun;82(11):5137-44
pubmed: 18367528
Virology. 2015 Oct;484:313-322
pubmed: 26149721
Viruses. 2020 Mar 31;12(4):
pubmed: 32244383
Sci Rep. 2019 Feb 22;9(1):2606
pubmed: 30796243
Bioinformatics. 2014 Nov 15;30(22):3276-8
pubmed: 25095880
Cell Res. 2020 Mar;30(3):269-271
pubmed: 32020029
PLoS One. 2010 Jun 25;5(6):e11147
pubmed: 20593022
Nature. 2016 Oct 12;538(7624):193-200
pubmed: 27734858
J Biol Chem. 2006 Apr 21;281(16):10669-81
pubmed: 16431923
Proc Natl Acad Sci U S A. 2006 Mar 28;103(13):5108-13
pubmed: 16549795
Protein Sci. 2020 Sep;29(9):1890-1901
pubmed: 32654247
Environ Microbiol. 2020 Jun;22(6):2001-2006
pubmed: 32367648
BMJ. 2020 Mar 12;368:m1036
pubmed: 32165426
Emerg Microbes Infect. 2020 Dec;9(1):837-842
pubmed: 32301390
Cell Host Microbe. 2020 May 13;27(5):841-848.e3
pubmed: 32289263
J Virol. 2008 Dec;82(24):12325-34
pubmed: 18922871
J Med Virol. 2020 Jun;92(6):584-588
pubmed: 32083328
Emerg Microbes Infect. 2020 Dec;9(1):1474-1488
pubmed: 32543348
PLoS Pathog. 2007 Jan;3(1):e5
pubmed: 17222058
Natl Sci Rev. 2020 Jun;7(6):1012-1023
pubmed: 34676127
Brief Bioinform. 2019 Jul 19;20(4):1160-1166
pubmed: 28968734
Virulence. 2017 Oct 3;8(7):1229-1244
pubmed: 28112573
J Virol. 1968 Oct;2(10):955-61
pubmed: 4302013
mBio. 2016 Dec 13;7(6):
pubmed: 27965448
J Transl Med. 2020 Apr 22;18(1):179
pubmed: 32321524
Nat Microbiol. 2020 Nov;5(11):1403-1407
pubmed: 32669681
Cell Mol Life Sci. 2016 Dec;73(23):4433-4448
pubmed: 27392606