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1.
J Virol ; 94(12)2020 06 01.
Article in English | MEDLINE | ID: covidwho-1723543

ABSTRACT

The novel coronavirus severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that recently emerged in China is thought to have a bat origin, as its closest known relative (BatCoV RaTG13) was described previously in horseshoe bats. We analyzed the selective events that accompanied the divergence of SARS-CoV-2 from BatCoV RaTG13. To this end, we applied a population genetics-phylogenetics approach, which leverages within-population variation and divergence from an outgroup. Results indicated that most sites in the viral open reading frames (ORFs) evolved under conditions of strong to moderate purifying selection. The most highly constrained sequences corresponded to some nonstructural proteins (nsps) and to the M protein. Conversely, nsp1 and accessory ORFs, particularly ORF8, had a nonnegligible proportion of codons evolving under conditions of very weak purifying selection or close to selective neutrality. Overall, limited evidence of positive selection was detected. The 6 bona fide positively selected sites were located in the N protein, in ORF8, and in nsp1. A signal of positive selection was also detected in the receptor-binding motif (RBM) of the spike protein but most likely resulted from a recombination event that involved the BatCoV RaTG13 sequence. In line with previous data, we suggest that the common ancestor of SARS-CoV-2 and BatCoV RaTG13 encoded/encodes an RBM similar to that observed in SARS-CoV-2 itself and in some pangolin viruses. It is presently unknown whether the common ancestor still exists and, if so, which animals it infects. Our data, however, indicate that divergence of SARS-CoV-2 from BatCoV RaTG13 was accompanied by limited episodes of positive selection, suggesting that the common ancestor of the two viruses was poised for human infection.IMPORTANCE Coronaviruses are dangerous zoonotic pathogens; in the last 2 decades, three coronaviruses have crossed the species barrier and caused human epidemics. One of these is the recently emerged SARS-CoV-2. We investigated how, since its divergence from a closely related bat virus, natural selection shaped the genome of SARS-CoV-2. We found that distinct coding regions in the SARS-CoV-2 genome evolved under conditions of different degrees of constraint and are consequently more or less prone to tolerate amino acid substitutions. In practical terms, the level of constraint provides indications about which proteins/protein regions are better suited as possible targets for the development of antivirals or vaccines. We also detected limited signals of positive selection in three viral ORFs. However, we warn that, in the absence of knowledge about the chain of events that determined the human spillover, these signals should not be necessarily interpreted as evidence of an adaptation to our species.


Subject(s)
Betacoronavirus/genetics , Evolution, Molecular , Selection, Genetic , Amino Acid Sequence , Animals , Betacoronavirus/classification , COVID-19 , Chiroptera/virology , Coronavirus Infections/virology , Genome, Viral/genetics , Humans , Models, Molecular , Open Reading Frames/genetics , Pandemics , Phylogeny , Pneumonia, Viral/virology , SARS-CoV-2 , Viral Proteins/chemistry , Viral Proteins/genetics
2.
J Infect Dis ; 224(3): 415-419, 2021 08 02.
Article in English | MEDLINE | ID: covidwho-1526165

ABSTRACT

Mutagenic ribonucleosides can act as broad-based antiviral agents. They are metabolized to the active ribonucleoside triphosphate form and concentrate in genomes of RNA viruses during viral replication. ß-d-N4-hydroxycytidine (NHC, initial metabolite of molnupiravir) is >100-fold more active than ribavirin or favipiravir against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), with antiviral activity correlated to the level of mutagenesis in virion RNA. However, NHC also displays host mutational activity in an animal cell culture assay, consistent with RNA and DNA precursors sharing a common intermediate of a ribonucleoside diphosphate. These results indicate highly active mutagenic ribonucleosides may hold risk for the host.


Subject(s)
Antiviral Agents/pharmacology , Cytidine/analogs & derivatives , Mutagens/pharmacology , SARS-CoV-2/drug effects , Animals , Antiviral Agents/adverse effects , CHO Cells/drug effects , Cells, Cultured , Cricetulus , Cytidine/adverse effects , Cytidine/pharmacology , Dose-Response Relationship, Drug , Mutagenesis/drug effects , Mutagens/adverse effects , SARS-CoV-2/genetics , Virus Replication/drug effects
3.
Int J Mol Sci ; 21(19)2020 Oct 01.
Article in English | MEDLINE | ID: covidwho-1389385

ABSTRACT

Tom70 is a versatile adaptor protein of 70 kDa anchored in the outer membrane of mitochondria in metazoa, fungi and amoeba. The tertiary structure was resolved for the Tom70 of yeast, showing 26 α-helices, most of them participating in the formation of 11 tetratricopeptide repeat (TPR) motifs. Tom70 serves as a docking site for cytosolic chaperone proteins and co-chaperones and is thereby involved in the uptake of newly synthesized chaperone-bound proteins in mitochondrial biogenesis. In yeast, Tom70 additionally mediates ER-mitochondria contacts via binding to sterol transporter Lam6/Ltc1. In mammalian cells, TOM70 promotes endoplasmic reticulum (ER) to mitochondria Ca2+ transfer by association with the inositol-1,4,5-triphosphate receptor type 3 (IP3R3). TOM70 is specifically targeted by the Bcl-2-related protein MCL-1 that acts as an anti-apoptotic protein in macrophages infected by intracellular pathogens, but also in many cancer cells. By participating in the recruitment of PINK1 and the E3 ubiquitin ligase Parkin, TOM70 can be implicated in the development of Parkinson's disease. TOM70 acts as receptor of the mitochondrial antiviral-signaling protein (MAVS) and thereby participates in the corresponding system of innate immunity against viral infections. The protein encoded by Orf9b in the genome of SARS-CoV-2 binds to TOM70, probably compromising the synthesis of type I interferons.


Subject(s)
Immunity, Innate , Mitochondrial Membrane Transport Proteins/chemistry , Animals , Betacoronavirus/genetics , Binding Sites , Humans , Mitochondrial Membrane Transport Proteins/metabolism , Open Reading Frames , Protein Binding , Protein Transport , SARS-CoV-2
4.
Enferm Infecc Microbiol Clin (Engl Ed) ; 2021 Jan 19.
Article in English, Spanish | MEDLINE | ID: covidwho-1385464
5.
Biochem Biophys Res Commun ; 545: 75-80, 2021 03 19.
Article in English | MEDLINE | ID: covidwho-1385064

ABSTRACT

Antiviral drug discovery continues to be an essential complement to vaccine development for overcoming the global pandemic caused by SARS-CoV-2. The genomic RNA of SARS-CoV-2 contains structural elements important for viral replication and/or pathogenesis making them potential therapeutic targets. Here we report on the stem-loop II motif, a highly conserved noncoding RNA element. Based on our homology model we determined that the G to U transversion in the SARS-CoV-2 stem-loop II motif (S2MG35U) forms a C-U base-pair isosteric to the C-G base-pair in the early 2000's SARS-CoV (S2M). In addition, chemo-enzymatic probing and molecular dynamics simulations indicate the S2MG35U conformational profile is altered compared to S2M in the apical loop region. We explored S2MG35U as a potential drug target by docking a library of FDA approved drugs. Enzymatic probing of the best docking ligands (aminoglycosides and polymyxins) indicated that polymyxin binding alters the conformational profile and/or secondary structure of the RNA. The SARS-CoV-2 stem-loop II motif conformational differences due to nucleotide transversion and ligand binding are highly significant and provide insight for future drug discovery efforts since the conformation of noncoding RNA elements affects their function.


Subject(s)
RNA, Viral/chemistry , SARS-CoV-2/genetics , Antiviral Agents/chemistry , Antiviral Agents/metabolism , Base Pairing , Binding Sites , Ligands , Models, Molecular , Molecular Docking Simulation , Molecular Dynamics Simulation , Nucleic Acid Conformation , RNA, Untranslated/chemistry , RNA, Untranslated/metabolism , RNA, Viral/metabolism , SARS-CoV-2/chemistry , SARS-CoV-2/metabolism
6.
Angew Chem Int Ed Engl ; 60(21): 11884-11891, 2021 05 17.
Article in English | MEDLINE | ID: covidwho-1384108

ABSTRACT

2D NOESY plays a central role in structural NMR spectroscopy. We have recently discussed methods that rely on solvent-driven exchanges to enhance NOE correlations between exchangeable and non-exchangeable protons in nucleic acids. Such methods, however, fail when trying to establish connectivities within pools of labile protons. This study introduces an alternative that also enhances NOEs between such labile sites, based on encoding a priori selected peaks by selective saturations. The resulting selective magnetization transfer (SMT) experiment proves particularly useful for enhancing the imino-imino cross-peaks in RNAs, which is a first step in the NMR resolution of these structures. The origins of these enhancements are discussed, and their potential is demonstrated on RNA fragments derived from the genome of SARS-CoV-2, recorded with better sensitivity and an order of magnitude faster than conventional 2D counterparts.


Subject(s)
Nuclear Magnetic Resonance, Biomolecular/methods , Protons , RNA, Viral/analysis , SARS-CoV-2/chemistry , Magnetic Phenomena , RNA, Viral/chemistry
7.
Front Med (Lausanne) ; 8: 631769, 2021.
Article in English | MEDLINE | ID: covidwho-1389197

ABSTRACT

Background: SARS-CoV-2 infection may not provide long lasting post-infection immunity. While hundreds of reinfections have reported only a few have been confirmed. Whole genome sequencing (WGS) of the viral isolates from the different episodes is mandatory to establish reinfection. Methods: Nasopharyngeal (NP), oropharyngeal (OP) and whole blood (WB) samples were collected from paired samples of four individuals who were suspected of SARS-CoV-2 reinfection based on distinct clinical episodes and RT-PCR tests. Details from their case record files and investigations were documented. RNA was extracted from the NP and OP samples and subjected to WGS, and the nucleotide and amino acid sequences were subjected to genome and protein-based functional annotation analyses. Serial serology was performed for Anti-N IgG, Anti- S1 RBD IgG, and sVNT (surrogate virus neutralizing test). Findings: Three patients were more symptomatic with lower Ct values and longer duration of illness. Seroconversion was detected soon after the second episode in three patients. WGS generated a genome coverage ranging from 80.07 to 99.7%. Phylogenetic analysis revealed sequences belonged to G, GR and "Other" clades. A total of 42mutations were identified in all the samples, consisting of 22 non-synonymous, 17 synonymous, two in upstream, and one in downstream regions of the SARS-CoV-2 genome. Comparative genomic and protein-based annotation analyses revealed differences in the presence and absence of specific mutations in the virus sequences from the two episodes in all four paired samples. Interpretation: Based on the criteria of genome variations identified by whole genome sequencing and supported by clinical presentation, molecular and serological tests, we were able to confirm reinfections in two patients, provide weak evidence of reinfection in the third patient and unable to rule out a prolonged infection in the fourth. This study emphasizes the importance of detailed analyses of clinical and serological information as well as the virus's genomic variations while assessing cases of SARS-CoV-2 reinfection.

8.
Brief Bioinform ; 22(5)2021 09 02.
Article in English | MEDLINE | ID: covidwho-1387714

ABSTRACT

In a recently published paper, we have found that SARS-CoV-2 hot-spot mutations are significantly associated with inverted repeat loci and CG dinucleotides. However, fast-spreading strains with new mutations (so-called mink farm mutations, England mutations and Japan mutations) have been recently described. We used the new datasets to check the positioning of mutation sites in genomes of the new SARS-CoV-2 strains. Using an open-access Palindrome analyzer tool, we found mutations in these new strains to be significantly enriched in inverted repeat loci.


Subject(s)
Mutation , SARS-CoV-2/genetics , COVID-19/virology , Genome, Viral , Humans
9.
Bioinformatics ; 2020 Dec 21.
Article in English | MEDLINE | ID: covidwho-1387720

ABSTRACT

SUMMARY: While over 150 thousand genomic sequences are currently available through dedicated repositories, ad hoc methods for the functional annotation of SARS-CoV-2 genomes do not harness all currently available resources for the annotation of functionally relevant genomic sites. Here we present CorGAT, a novel tool for the functional annotation of SARS-CoV-2 genomic variants. By comparisons with other state of the art methods we demonstrate that, by providing a more comprehensive and rich annotation, our method can facilitate the identification of evolutionary patterns in the genome of SARS-CoV-2. AVAILABILITY: Galaxy: http://corgat.cloud.ba.infn.it/galaxy; software: https://github.com/matteo14c/CorGAT/tree/Revision_V1; docker: https://hub.docker.com/r/laniakeacloud/galaxy_corgat. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

10.
IDCases ; 25: e01187, 2021.
Article in English | MEDLINE | ID: covidwho-1267690

ABSTRACT

BACKGROUND: SARS-CoV-2 uses the human cell receptor angiotensin-converting enzyme (ACE2). ACE2 is widely present in the cardiovascular system including the myocardium and the conduction system. COVID-19 patients that present severe symptoms have been reported to have complications involving myocardial injuries caused by the virus. Here we report the detection of SARS-CoV-2 by whole genome sequencing in the endocardium of a patient with severe bradycardia. CASE PRESENTATION: We report a case of a 34-year-old male patient with COVID-19 tested by PCR, he started with gastrointestinal symptoms, however, he quickly deteriorated his hemodynamic state by means of myocarditis and bradycardia. After performing an endocardium biopsy, it was possible to identify the presence of SARS-CoV-2 in the heart tissue and to sequence its whole genome using the ARTIC-Network protocol and a modified tissue RNA extraction method. The patient's outcome was improved after a permanent pacemaker was implanted. CONCLUSIONS: It was possible to identify a SARS-CoV-2 clade 20A in the endocardium of the reported patient.

11.
Sci Rep ; 11(1): 11773, 2021 06 03.
Article in English | MEDLINE | ID: covidwho-1258597

ABSTRACT

Since the first report of SARS-CoV-2 in China in 2019, there has been a huge debate about the origin. In this work, using a different method we aimed to strengthen the observation that no evidence of genetic manipulation has been found by (1) detecting classical restriction site (RS) sequence in human SARS-CoV-2 genomes and (2) comparing them with other recombinant SARS-CoV-like virus created for experimental purposes. Finally, we propose a novel approach consisting in the generation of a restriction endonucleases site map of SARS-CoV-2 and other related coronavirus genomes to be used as a fingerprint to trace the virus evolution.


Subject(s)
Biological Evolution , DNA Barcoding, Taxonomic/methods , DNA Restriction Enzymes/genetics , SARS-CoV-2/genetics , Animals , Chiroptera/virology , DNA Restriction Enzymes/metabolism , Genetic Markers , Genome, Viral , Humans , Restriction Mapping , SARS-CoV-2/isolation & purification , Spike Glycoprotein, Coronavirus/genetics
12.
J Biol Chem ; 297(1): 100821, 2021 07.
Article in English | MEDLINE | ID: covidwho-1240418

ABSTRACT

Viral proteins are known to be methylated by host protein arginine methyltransferases (PRMTs) necessary for the viral life cycle, but it remains unknown whether severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) proteins are methylated. Herein, we show that PRMT1 methylates SARS-CoV-2 nucleocapsid (N) protein at residues R95 and R177 within RGG/RG motifs, preferred PRMT target sequences. We confirmed arginine methylation of N protein by immunoblotting viral proteins extracted from SARS-CoV-2 virions isolated from cell culture. Type I PRMT inhibitor (MS023) or substitution of R95 or R177 with lysine inhibited interaction of N protein with the 5'-UTR of SARS-CoV-2 genomic RNA, a property required for viral packaging. We also defined the N protein interactome in HEK293 cells, which identified PRMT1 and many of its RGG/RG substrates, including the known interacting protein G3BP1 as well as other components of stress granules (SGs), which are part of the host antiviral response. Methylation of R95 regulated the ability of N protein to suppress the formation of SGs, as R95K substitution or MS023 treatment blocked N-mediated suppression of SGs. Also, the coexpression of methylarginine reader Tudor domain-containing protein 3 quenched N protein-mediated suppression of SGs in a dose-dependent manner. Finally, pretreatment of VeroE6 cells with MS023 significantly reduced SARS-CoV-2 replication. Because type I PRMT inhibitors are already undergoing clinical trials for cancer treatment, inhibiting arginine methylation to target the later stages of the viral life cycle such as viral genome packaging and assembly of virions may represent an additional therapeutic application of these drugs.


Subject(s)
Arginine/metabolism , COVID-19/metabolism , COVID-19/virology , Nucleocapsid Proteins/chemistry , Nucleocapsid Proteins/metabolism , RNA, Viral/metabolism , SARS-CoV-2/physiology , Amino Acid Motifs , COVID-19/genetics , Cytoplasmic Granules/genetics , Cytoplasmic Granules/metabolism , HEK293 Cells , Humans , Methylation , Nucleocapsid Proteins/genetics , RNA Stability , RNA, Viral/chemistry , RNA, Viral/genetics , SARS-CoV-2/chemistry , SARS-CoV-2/genetics , Virus Replication
13.
Basic Clin Pharmacol Toxicol ; 129(2): 104-129, 2021 Aug.
Article in English | MEDLINE | ID: covidwho-1223466

ABSTRACT

BACKGROUND: The COVID-19 pandemic has demanded effective therapeutic protocol from researchers and clinicians across the world. Currently, a large amount of primary data have been generated from several preclinical studies. At least 300 clinical trials are underway for drug repurposing against COVID-19; the clinician needs objective evidence-based medication to treat COVID-19. OBSERVATIONS: Single-stranded RNA viral genome of SARS-CoV-2 encodes structural proteins (spike protein), non-structural enzymatic proteins (RNA-dependent RNA polymerase, helicase, papain-like protease, 3-chymotrypsin-like protease) and other accessory proteins. These four enzymatic proteins on spike protein are rate-limiting steps in viral replications and, therefore, an attractive target for drug development against SARS-CoV-2. In silico and in vitro studies have identified various potential epitomes as candidate sequences for vaccine development. These studies have also revealed potential targets for drug development and drug repurposing against COVID-19. Clinical trials utilizing antiviral drugs and other drugs have given inconclusive results regarding their clinical efficacy and side effects. The need for angiotensin-converting enzyme (ACE-2) inhibitors/angiotensin receptor blockers and corticosteroids has been recommended. Western countries have adopted telemedicine as an alternative to prevent transmission of infection in the population. Currently, no proven, evidence-based therapeutic regimen exists for COVID-19. CONCLUSION: The COVID-19 pandemic has put tremendous pressure on researchers to evaluate and approve drugs effective against the disease. Well-controlled randomized trials should assess medicines that are not marketed with substantial evidence of safety and efficacy and more emphasis on time tested approaches for drug evaluation.


Subject(s)
Antiviral Agents/therapeutic use , COVID-19/drug therapy , Drug Repositioning , COVID-19/epidemiology , COVID-19/virology , Computer Simulation , Humans , Pandemics , SARS-CoV-2/drug effects
14.
Proc Natl Acad Sci U S A ; 118(21)2021 05 25.
Article in English | MEDLINE | ID: covidwho-1223143

ABSTRACT

The genome of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) coronavirus has a capping modification at the 5'-untranslated region (UTR) to prevent its degradation by host nucleases. These modifications are performed by the Nsp10/14 and Nsp10/16 heterodimers using S-adenosylmethionine as the methyl donor. Nsp10/16 heterodimer is responsible for the methylation at the ribose 2'-O position of the first nucleotide. To investigate the conformational changes of the complex during 2'-O methyltransferase activity, we used a fixed-target serial synchrotron crystallography method at room temperature. We determined crystal structures of Nsp10/16 with substrates and products that revealed the states before and after methylation, occurring within the crystals during the experiments. Here we report the crystal structure of Nsp10/16 in complex with Cap-1 analog (m7GpppAm2'-O). Inhibition of Nsp16 activity may reduce viral proliferation, making this protein an attractive drug target.


Subject(s)
RNA Caps/metabolism , RNA, Messenger/metabolism , RNA, Viral/metabolism , SARS-CoV-2/chemistry , Crystallography , Methylation , Methyltransferases/chemistry , Methyltransferases/metabolism , Multiprotein Complexes/chemistry , Multiprotein Complexes/metabolism , RNA Cap Analogs/chemistry , RNA Cap Analogs/metabolism , RNA Caps/chemistry , RNA, Messenger/chemistry , RNA, Viral/chemistry , S-Adenosylhomocysteine/chemistry , S-Adenosylhomocysteine/metabolism , S-Adenosylmethionine/chemistry , S-Adenosylmethionine/metabolism , SARS-CoV-2/genetics , SARS-CoV-2/metabolism , Synchrotrons , Viral Nonstructural Proteins/chemistry , Viral Nonstructural Proteins/metabolism , Viral Regulatory and Accessory Proteins/chemistry , Viral Regulatory and Accessory Proteins/metabolism
15.
Adv Exp Med Biol ; 1318: 355-368, 2021.
Article in English | MEDLINE | ID: covidwho-1222724

ABSTRACT

During the COVID-19 pandemic associated with high incidence, transmissibility, and mortality, this chapter focuses on three phases of the disease: initial exposure, initiation of the immune response to the agent, and finally, an inflammatory/autoimmune-like presentation with pulmonary, neurological, and renal failure and disseminated intravascular coagulation which occurs in a small proportion of the patients. The elegant demonstration of the site of interaction between the spike (S) protein of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which is the causative agent of COVID-19, and the ACE (angiotensin-converting enzyme) 2 receptor of cells distributed throughout the body has enabled research efforts to develop pharmacological and immune countermeasures to the viral phase of the disease. This chapter rapidly reviews the molecular and structural organization of SARS-CoV-2 and its interaction with ACE2. It is followed by a discussion over the role of the major histocompatibility complex (MHC) in recognition of the virus. The importance of rapid compartmentation of the viral genome into the target cells as opposed to the binding constant of the virus for the ACE receptor is discussed. Host factors affecting the immune response to the virus are examined, and the subsequent inflammatory dysregulation enabling the cytokine storm leading to system organ failure is described. Finally, the similarities of the clinical effects of the murine hepatitis virus-JHM (a coronavirus) on multi-organ systems (liver, brain, clotting cascade) as described by Perlman and colleagues permit insights regarding the role of the interaction between the host and the virus in developing the clinical presentation of the inflammatory/autoimmune disorders that occur in multiple sclerosis, neuromyelitis optica, and more interestingly, during the third phase of COVID-19.


Subject(s)
COVID-19 , Pandemics , Animals , Humans , Lung , Mice , Peptidyl-Dipeptidase A/genetics , SARS-CoV-2
16.
Proc Natl Acad Sci U S A ; 118(21)2021 05 25.
Article in English | MEDLINE | ID: covidwho-1220249

ABSTRACT

Prolonged detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA and recurrence of PCR-positive tests have been widely reported in patients after recovery from COVID-19, but some of these patients do not appear to shed infectious virus. We investigated the possibility that SARS-CoV-2 RNAs can be reverse-transcribed and integrated into the DNA of human cells in culture and that transcription of the integrated sequences might account for some of the positive PCR tests seen in patients. In support of this hypothesis, we found that DNA copies of SARS-CoV-2 sequences can be integrated into the genome of infected human cells. We found target site duplications flanking the viral sequences and consensus LINE1 endonuclease recognition sequences at the integration sites, consistent with a LINE1 retrotransposon-mediated, target-primed reverse transcription and retroposition mechanism. We also found, in some patient-derived tissues, evidence suggesting that a large fraction of the viral sequences is transcribed from integrated DNA copies of viral sequences, generating viral-host chimeric transcripts. The integration and transcription of viral sequences may thus contribute to the detection of viral RNA by PCR in patients after infection and clinical recovery. Because we have detected only subgenomic sequences derived mainly from the 3' end of the viral genome integrated into the DNA of the host cell, infectious virus cannot be produced from the integrated subgenomic SARS-CoV-2 sequences.


Subject(s)
COVID-19/genetics , COVID-19/virology , SARS-CoV-2/genetics , Virus Integration/genetics , Animals , COVID-19/metabolism , Chlorocebus aethiops , Genome, Viral , HEK293 Cells , Humans , RNA, Viral/genetics , SARS-CoV-2/metabolism , Vero Cells , Virus Integration/physiology , Virus Replication/genetics , Virus Replication/physiology
17.
Database (Oxford) ; 20212021 05 08.
Article in English | MEDLINE | ID: covidwho-1219730

ABSTRACT

Numerous studies demonstrate frequent mutations in the genome of SARS-CoV-2. Our goal was to statistically link mutations to severe disease outcome. We used an automated machine learning approach where 1594 viral genomes with available clinical follow-up data were used as the training set (797 'severe' and 797 'mild'). The best algorithm, based on random forest classification combined with the LASSO feature selection algorithm, was employed to the training set to link mutation signatures and outcome. The performance of the final model was estimated by repeated, stratified, 10-fold cross validation (CV) and then adjusted for multiple testing with Bootstrap Bias Corrected CV. We identified 26 protein and Untranslated Region (UTR) mutations significantly linked to severe outcome. The best classification algorithm uses a mutation signature of 22 mutations as well as the patient's age as the input and shows high classification efficiency with an area under the curve (AUC) of 0.94 [confidence interval (CI): [0.912, 0.962]] and a prediction accuracy of 87% (CI: [0.830, 0.903]). Finally, we established an online platform (https://covidoutcome.com/) that is capable to use a viral sequence and the patient's age as the input and provides a percentage estimation of disease severity. We demonstrate a statistical association between mutation signatures of SARS-CoV-2 and severe outcome of COVID-19. The established analysis platform enables a real-time analysis of new viral genomes.


Subject(s)
COVID-19/genetics , COVID-19/pathology , Genome, Viral , Mutation , SARS-CoV-2/genetics , Severity of Illness Index , Area Under Curve , COVID-19/virology , Datasets as Topic , Humans , Machine Learning , Probability , Untranslated Regions
18.
J Infect Dev Ctries ; 15(4): 470-477, 2021 04 30.
Article in English | MEDLINE | ID: covidwho-1218644

ABSTRACT

INTRODUCTION: Coronaviruses which are single-stranded RNAs, are members of a large family of viruses that may be important pathogens for humans. SARS-CoV-2 was found to cause the severe respiratory syndrome, and on January 22, 2020 first human-to-human transmission was reported. We aimed to reveal the complete genomes of 19 SARS-CoV-2 isolates from Denizli province and identify Turkish patients' genetic similarities. METHODOLOGY: 15 samples with the highest viral loads resulting from RT-PCR were selected for NGS analysis. Fifteen SARS-CoV-2 complete genome sequences were then subjected to phylogenetic analysis and uploaded to the GISAID database. Phylogenetic trees were constructed by the Neighbor-Joining method using MEGAX software. RESULTS: Whole-genome sequencing of the viral RNA samples revealed 32 missense, 21 synonymous, and 4 non-coding alleles. In all samples c.1-25C>T (5'UTR), c.14144C>T (ORF1ab), c.2772C>T (ORF1ab) and c.1841A>G(S) mutations were detected. Phylogenetic analysis revealed that most of the present study's genomes are in 20B clade while the two are in 20A. The phylogenetic tree constructed with all complete SARS-CoV-2 genomes of Turkey showed that the viruses were spread nearly homogenous on eastern (around Kars) and western (around Istanbul) sides. CONCLUSIONS: Here, we reported the viral genomes in Denizli comprehensively for the first time. We identified 11 rare missense mutations in the virus compared to the reference genome. Phylogenetic analysis revealed that while most of our isolates were similar to European sequences, some had different sublineages depending on their genomic variants.


Subject(s)
Phylogeny , SARS-CoV-2/genetics , COVID-19/virology , Genome, Viral , Humans , Mutation , SARS-CoV-2/isolation & purification , Whole Genome Sequencing
19.
Mol Biol Evol ; 38(8): 3046-3059, 2021 07 29.
Article in English | MEDLINE | ID: covidwho-1214648

ABSTRACT

Global sequencing of genomes of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has continued to reveal new genetic variants that are the key to unraveling its early evolutionary history and tracking its global spread over time. Here we present the heretofore cryptic mutational history and spatiotemporal dynamics of SARS-CoV-2 from an analysis of thousands of high-quality genomes. We report the likely most recent common ancestor of SARS-CoV-2, reconstructed through a novel application and advancement of computational methods initially developed to infer the mutational history of tumor cells in a patient. This progenitor genome differs from genomes of the first coronaviruses sampled in China by three variants, implying that none of the earliest patients represent the index case or gave rise to all the human infections. However, multiple coronavirus infections in China and the United States harbored the progenitor genetic fingerprint in January 2020 and later, suggesting that the progenitor was spreading worldwide months before and after the first reported cases of COVID-19 in China. Mutations of the progenitor and its offshoots have produced many dominant coronavirus strains that have spread episodically over time. Fingerprinting based on common mutations reveals that the same coronavirus lineage has dominated North America for most of the pandemic in 2020. There have been multiple replacements of predominant coronavirus strains in Europe and Asia as well as continued presence of multiple high-frequency strains in Asia and North America. We have developed a continually updating dashboard of global evolution and spatiotemporal trends of SARS-CoV-2 spread (http://sars2evo.datamonkey.org/).


Subject(s)
COVID-19/genetics , SARS-CoV-2/genetics , Biological Evolution , COVID-19/metabolism , Computational Biology/methods , Contact Tracing/methods , Evolution, Molecular , Genome, Viral , Humans , Mutation , Pandemics , Phylogeny , SARS-CoV-2/metabolism , SARS-CoV-2/pathogenicity , Sequence Analysis, DNA/methods
20.
EMBO Mol Med ; 13(6): e14062, 2021 06 07.
Article in English | MEDLINE | ID: covidwho-1210029

ABSTRACT

Scientists and the public were alarmed at the first large viral variant of SARS-CoV-2 reported in December 2020. We have followed the time course of emerging viral mutants and variants during the SARS-CoV-2 pandemic in ten countries on four continents. We examined > 383,500 complete SARS-CoV-2 nucleotide sequences in GISAID (Global Initiative of Sharing All Influenza Data) with sampling dates extending until April 05, 2021. These sequences originated from ten different countries: United Kingdom, South Africa, Brazil, United States, India, Russia, France, Spain, Germany, and China. Among the 77 to 100 novel mutations, some previously reported mutations waned and some of them increased in prevalence over time. VUI2012/01 (B.1.1.7) and 501Y.V2 (B.1.351), the so-called UK and South Africa variants, respectively, and two variants from Brazil, 484K.V2, now called P.1 and P.2, increased in prevalence. Despite lockdowns, worldwide active replication in genetically and socio-economically diverse populations facilitated selection of new mutations. The data on mutant and variant SARS-CoV-2 strains provided here comprise a global resource for easy access to the myriad mutations and variants detected to date globally. Rapidly evolving new variant and mutant strains might give rise to escape variants, capable of limiting the efficacy of vaccines, therapies, and diagnostic tests.


Subject(s)
COVID-19/prevention & control , Genome, Viral , SARS-CoV-2/genetics , COVID-19/pathology , COVID-19/therapy , COVID-19/virology , COVID-19 Vaccines/administration & dosage , COVID-19 Vaccines/immunology , Humans , Mutation , SARS-CoV-2/isolation & purification , Spike Glycoprotein, Coronavirus/genetics , Viral Nonstructural Proteins/genetics
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