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1.
Int J Antimicrob Agents ; 57(2): 106272, 2021 Feb.
Article in English | MEDLINE | ID: covidwho-1385674

ABSTRACT

INTRODUCTION: Genomic alterations in a viral genome can lead to either better or worse outcome and identifying these mutations is of utmost importance. Here, we correlated protein-level mutations in the SARS-CoV-2 virus to clinical outcome. METHODS: Mutations in viral sequences from the GISAID virus repository were evaluated by using "hCoV-19/Wuhan/WIV04/2019" as the reference. Patient outcomes were classified as mild disease, hospitalization and severe disease (death or documented treatment in an intensive-care unit). Chi-square test was applied to examine the association between each mutation and patient outcome. False discovery rate was computed to correct for multiple hypothesis testing and results passing FDR cutoff of 5% were accepted as significant. RESULTS: Mutations were mapped to amino acid changes for 3,733 non-silent mutations. Mutations correlated to mild outcome were located in the ORF8, NSP6, ORF3a, NSP4, and in the nucleocapsid phosphoprotein N. Mutations associated with inferior outcome were located in the surface (S) glycoprotein, in the RNA dependent RNA polymerase, in ORF3a, NSP3, ORF6 and N. Mutations leading to severe outcome with low prevalence were found in the ORF3A and in NSP7 proteins. Four out of 22 of the most significant mutations mapped onto a 10 amino acid long phosphorylated stretch of N indicating that in spite of obvious sampling restrictions the approach can find functionally relevant sites in the viral genome. CONCLUSIONS: We demonstrate that mutations in the viral genes may have a direct correlation to clinical outcome. Our results help to quickly identify SARS-CoV-2 infections harboring mutations related to severe outcome.


Subject(s)
COVID-19/drug therapy , COVID-19/etiology , Mutation , SARS-CoV-2/genetics , Coronavirus Nucleocapsid Proteins/genetics , Female , Hospitalization , Humans , Male , Mutation Rate , Viral Nonstructural Proteins/genetics , Viral Proteins/genetics , Viroporin Proteins/genetics
2.
Gene Rep ; 25: 101044, 2021 Dec.
Article in English | MEDLINE | ID: covidwho-1385601

ABSTRACT

SARS-CoV-2 is mutating and creating divergent variants by altering the composition of essential constituent proteins. Pharmacologically, it is crucial to understand the diverse mechanism of mutations for stable vaccine or anti-viral drug design. Our current study concentrates on all the constituent proteins of 469 SARS-CoV-2 genome samples, derived from Indian patients. However, the study may easily be extended to the samples across the globe. We perform clustering analysis towards identifying unique variants in each of the SARS-CoV-2 proteins. A total of 536 mutated positions within the coding regions of SARS-CoV-2 proteins are detected among the identified variants from Indian isolates. We quantify mutations by focusing on the unique variants of each SARS-CoV-2 protein. We report the average number of mutation per variant, percentage of mutated positions, synonymous and non-synonymous mutations, mutations occurring in three codon positions and so on. Our study reveals the most susceptible six (06) proteins, which are ORF1ab, Spike (S), Nucleocapsid (N), ORF3a, ORF7a, and ORF8. Several non-synonymous substitutions are observed to be unique in different SARS-CoV-2 proteins. A total of 57 possible deleterious amino acid substitutions are predicted, which may impact on the protein functions. Several mutations show a large decrease in protein stability and are observed in putative functional domains of the proteins that might have some role in disease pathogenesis. We observe a good number of physicochemical property change during above deleterious substitutions.

3.
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.

4.
J Biomol Struct Dyn ; 39(12): 4243-4255, 2021 08.
Article in English | MEDLINE | ID: covidwho-1317834

ABSTRACT

Recent outbreak of novel Coronavirus disease () pandemic around the world is associated with severe acute respiratory syndrome. The death toll associated with the pandemic is increasing day by day. SARS-CoV-2 is an enveloped virus and its N terminal domain (NTD) of Nucleocapsid protein (N protein) binds to the viral (+) sense RNA and results in virus ribonucleoprotien complex, essential for the virus replication. The N protein is composed of a serine-rich linker region sandwiched between NTD and C terminal (CTD). These terminals play a role in viral entry and its processing post entry. The NTD of SARS-CoV-2 N protein forms orthorhombic crystals and binds to the viral genome. Therefore, there is always a quest to target RNA binding domain of nucleocapsid phosphoprotein (NTD-N-protein which in turn may help in controlling diseases caused by SARS-CoV-2 in humans. The role of Chloroquine and Hydroxychloroquine as potential treatments for is still under debate globally because of some side effects associated with it. This study involves the In silico interactions of Chloroquine and Hydroxychloroquine with the NTD-N-protein of SARS-CoV-2. With the help of various computational methods, we have explored the potential role of both of these antiviral drugs for the treatment of patients by comparing the efficacy of both of the drugs to bind to NTD-N-protein. In our research Hydroxychloroquine exhibited potential inhibitory effects of NTD-N-protein with binding energy -7.28 kcal/mol than Chloroquine (-6.30 kcal/mol) at SARS-CoV-2 receptor recognition of susceptible cells. The outcomes of this research strongly appeal for in vivo trials of Hydroxychloroquine for the patients infected with . Furthermore, the recommended doses of Hydroxychloroquine may reduce the chances of catching to the healthcare workers and staff who are in contact with or delivering direct care to coronavirus patients as long as they have not been diagnosed with . We further hypothesize that the comparative NTD-N-protein -drug docking interactions may help to understand the comparative efficacy of other candidate repurposing drugs until discovery of a proper vaccine.Communicated by Ramaswamy H. Sarma.


Subject(s)
COVID-19 , Hydroxychloroquine , Antiviral Agents/pharmacology , COVID-19/drug therapy , Chloroquine/pharmacology , Computer Simulation , Drug Repositioning , Humans , Hydroxychloroquine/pharmacology , Nucleocapsid , Nucleocapsid Proteins , RNA-Binding Motifs , SARS-CoV-2
5.
J Clin Invest ; 131(14)2021 07 15.
Article in English | MEDLINE | ID: covidwho-1249496

ABSTRACT

The SARS-CoV-2 virus, which causes COVID-19, has been associated globally with substantial morbidity and mortality. Numerous reports over the past year have described the clinical and immunological profiles of COVID-19 patients, and while some trends have emerged for risk stratification, they do not provide a complete picture. Therefore, efforts are ongoing to identify genetic susceptibility factors of severe disease. In this issue of the JCI, Povysil et al. performed a large, multiple-country study, sequencing genomes from patients with mild and severe COVID-19, along with population controls. Contrary to previous reports, the authors observed no enrichment of predicted loss-of-function variants in genes in the type I interferon pathway, which might predispose to severe disease. These studies suggest that more evidence is needed to substantiate the hypothesis for a genetic immune predisposition to severe COVID-19, and highlights the importance of considering experimental design when implicating a monogenic basis for severe disease.


Subject(s)
COVID-19 , Interferon Type I , Genetic Predisposition to Disease , Humans , SARS-CoV-2
6.
J Virol ; 95(15): e0049621, 2021 07 12.
Article in English | MEDLINE | ID: covidwho-1243690

ABSTRACT

The Severe acute respiratory syndrome coronavirus (SARS-CoV) and SARS-CoV-2 originated in bats and adapted to infect humans. Several SARS-CoV-2 strains have been identified. Genetic variation is fundamental to virus evolution and, in response to selection pressure, is manifested as the emergence of new strains and species adapted to different hosts or with novel pathogenicity. The combination of variation and selection forms a genetic footprint on the genome, consisting of the preferential accumulation of mutations in particular areas. Properties of betacoronaviruses contributing to variation and the emergence of new strains and species are beginning to be elucidated. To better understand their variation, we profiled the accumulation of mutations in all species in the genus Betacoronavirus, including SARS-CoV-2 and two other species that infect humans: SARS-CoV and Middle East respiratory syndrome coronavirus (MERS-CoV). Variation profiles identified both genetically stable and variable areas at homologous locations across species within the genus Betacoronavirus. The S glycoprotein is the most variable part of the genome and is structurally disordered. Other variable parts include proteins 3 and 7 and ORF8, which participate in replication and suppression of antiviral defense. In contrast, replication proteins in ORF1b are the least variable. Collectively, our results show that variation and structural disorder in the S glycoprotein is a general feature of all members of the genus Betacoronavirus, including SARS-CoV-2. These findings highlight the potential for the continual emergence of new species and strains with novel biological properties and indicate that the S glycoprotein has a critical role in host adaptation. IMPORTANCE Natural infection with SARS-CoV-2 and vaccines triggers the formation of antibodies against the S glycoprotein, which are detected by antibody-based diagnostic tests. Our analysis showed that variation in the S glycoprotein is a general feature of all species in the genus Betacoronavirus, including three species that infect humans: SARS-CoV, SARS-CoV-2, and MERS-CoV. The variable nature of the S glycoprotein provides an explanation for the emergence of SARS-CoV-2, the differentiation of SARS-CoV-2 into strains, and the probability of SARS-CoV-2 repeated infections in people. Variation of the S glycoprotein also has important implications for the reliability of SARS-CoV-2 antibody-based diagnostic tests and the design and deployment of vaccines and antiviral drugs. These findings indicate that adjustments to vaccine design and deployment and to antibody-based diagnostic tests are necessary to account for S glycoprotein variation.


Subject(s)
Betacoronavirus/genetics , Evolution, Molecular , Genetic Variation , Genome, Viral , Spike Glycoprotein, Coronavirus/genetics , Genome-Wide Association Study , Humans
7.
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
8.
Proc Natl Acad Sci U S A ; 118(17)2021 04 27.
Article in English | MEDLINE | ID: covidwho-1172591

ABSTRACT

In order to understand the transmission and virulence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), it is necessary to understand the functions of each of the gene products encoded in the viral genome. One feature of the SARS-CoV-2 genome that is not present in related, common coronaviruses is ORF10, a putative 38-amino acid protein-coding gene. Proteomic studies found that ORF10 binds to an E3 ubiquitin ligase containing Cullin-2, Rbx1, Elongin B, Elongin C, and ZYG11B (CRL2ZYG11B). Since CRL2ZYG11B mediates protein degradation, one possible role for ORF10 is to "hijack" CRL2ZYG11B in order to target cellular, antiviral proteins for ubiquitylation and subsequent proteasomal degradation. Here, we investigated whether ORF10 hijacks CRL2ZYG11B or functions in other ways, for example, as an inhibitor or substrate of CRL2ZYG11B While we confirm the ORF10-ZYG11B interaction and show that the N terminus of ORF10 is critical for it, we find no evidence that ORF10 is functioning to inhibit or hijack CRL2ZYG11B Furthermore, ZYG11B and its paralog ZER1 are dispensable for SARS-CoV-2 infection in cultured cells. We conclude that the interaction between ORF10 and CRL2ZYG11B is not relevant for SARS-CoV-2 infection in vitro.


Subject(s)
COVID-19/metabolism , Cell Cycle Proteins/metabolism , Cullin Proteins/metabolism , Multiprotein Complexes/metabolism , Open Reading Frames , SARS-CoV-2/metabolism , Viral Proteins/metabolism , COVID-19/genetics , Cell Cycle Proteins/genetics , Cullin Proteins/genetics , HEK293 Cells , Humans , Multiprotein Complexes/genetics , SARS-CoV-2/genetics , Viral Proteins/genetics
9.
Front Genet ; 12: 586569, 2021.
Article in English | MEDLINE | ID: covidwho-1170081

ABSTRACT

Humanity has seen numerous pandemics during its course of evolution. The list includes several incidents from the past, such as measles, Ebola, severe acute respiratory syndrome (SARS), and Middle East respiratory syndrome (MERS), etc. The latest edition to this is coronavirus disease 2019 (COVID-19), caused by the novel coronavirus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). As of August 18, 2020, COVID-19 has affected over 21 million people from 180 + countries with 0.7 million deaths across the globe. Genomic technologies have enabled us to understand the genomic constitution of pathogens, their virulence, evolution, and rate of mutation, etc. To date, more than 83,000 viral genomes have been deposited in public repositories, such as GISAID and NCBI. While we are writing this, India is the third most affected country by COVID-19, with 2.7 million cases and > 53,000 deaths. Gujarat is the 11th highest affected state with a 3.48% death rate compared to the national average of 1.91%. In this study, a total of 502 SARS-CoV-2 genomes from Gujarat were sequenced and analyzed to understand its phylogenetic distribution and variants against global and national sequences. Further variants were analyzed from diseased and recovered patients from Gujarat and the world to understand its role in pathogenesis. Among the missense mutations present in the Gujarat SARS-CoV-2 genomes, C28854T (Ser194Leu) had an allele frequency of 47.62 and 7.25% in deceased patients from the Gujarat and global datasets, respectively. In contrast, the allele frequency of 35.16 and 3.20% was observed in recovered patients from the Gujarat and global datasets, respectively. It is a deleterious mutation present in the nucleocapsid (N) gene and is significantly associated with mortality in Gujarat patients with a p-value of 0.067 and in the global dataset with a p-value of 0.000924. The other deleterious variant identified in deceased patients from Gujarat (p-value of 0.355) and the world (p-value of 2.43E-06) is G25563T, which is located in Orf3a and plays a potential role in viral pathogenesis. SARS-CoV-2 genomes from Gujarat are forming distinct clusters under the GH clade of GISAID. This study will shed light on the viral haplotype in SARS-CoV-2 samples from Gujarat, India.

10.
Emerg Microbes Infect ; 9(1): 221-236, 2020.
Article in English | MEDLINE | ID: covidwho-1169480

ABSTRACT

A mysterious outbreak of atypical pneumonia in late 2019 was traced to a seafood wholesale market in Wuhan of China. Within a few weeks, a novel coronavirus tentatively named as 2019 novel coronavirus (2019-nCoV) was announced by the World Health Organization. We performed bioinformatics analysis on a virus genome from a patient with 2019-nCoV infection and compared it with other related coronavirus genomes. Overall, the genome of 2019-nCoV has 89% nucleotide identity with bat SARS-like-CoVZXC21 and 82% with that of human SARS-CoV. The phylogenetic trees of their orf1a/b, Spike, Envelope, Membrane and Nucleoprotein also clustered closely with those of the bat, civet and human SARS coronaviruses. However, the external subdomain of Spike's receptor binding domain of 2019-nCoV shares only 40% amino acid identity with other SARS-related coronaviruses. Remarkably, its orf3b encodes a completely novel short protein. Furthermore, its new orf8 likely encodes a secreted protein with an alpha-helix, following with a beta-sheet(s) containing six strands. Learning from the roles of civet in SARS and camel in MERS, hunting for the animal source of 2019-nCoV and its more ancestral virus would be important for understanding the origin and evolution of this novel lineage B betacoronavirus. These findings provide the basis for starting further studies on the pathogenesis, and optimizing the design of diagnostic, antiviral and vaccination strategies for this emerging infection.


Subject(s)
Betacoronavirus/genetics , Coronavirus Infections/virology , Genome, Viral , Pneumonia, Viral/virology , Amino Acid Sequence , Betacoronavirus/isolation & purification , COVID-19 , China , Humans , Phylogeny , SARS-CoV-2 , Sequence Analysis, Protein , Travel , Viral Proteins/chemistry , Viral Proteins/genetics
11.
Travel Med Infect Dis ; 40: 101980, 2021.
Article in English | MEDLINE | ID: covidwho-1096252

ABSTRACT

BACKGROUND: In Marseille, France, the COVID-19 incidence evolved unusually with several successive epidemic phases. The second outbreak started in July, was associated with North Africa, and involved travelers and an outbreak on passenger ships. This suggested the involvement of a new viral variant. METHODS: We sequenced the genomes from 916 SARS-CoV-2 strains from COVID-19 patients in our institute. The patients' demographic and clinical features were compared according to the infecting viral variant. RESULTS: From June 26th to August 14th, we identified a new viral variant (Marseille-1). Based on genome sequences (n = 89) or specific qPCR (n = 53), 142 patients infected with this variant were detected. It is characterized by a combination of 10 mutations located in the nsp2, nsp3, nsp12, S, ORF3a, ORF8 and N/ORF14 genes. We identified Senegal and Gambia, where the virus had been transferred from China and Europe in February-April as the sources of the Marseille-1 variant, which then most likely reached Marseille through Maghreb when French borders reopened. In France, this variant apparently remained almost limited to Marseille. In addition, it was significantly associated with a milder disease compared to clade 20A ancestor strains, in univariate analysis. CONCLUSION: Our results demonstrate that SARS-CoV-2 can genetically diversify rapidly, its variants can diffuse internationally and cause successive outbreaks.


Subject(s)
COVID-19/virology , SARS-CoV-2/classification , SARS-CoV-2/genetics , Adult , Africa South of the Sahara/epidemiology , Aged , Amino Acid Substitution , COVID-19/epidemiology , China/epidemiology , Coronavirus Papain-Like Proteases/genetics , Coronavirus RNA-Dependent RNA Polymerase/genetics , Female , France/epidemiology , Genome, Viral , Humans , Male , Middle Aged , Mutation , Phylogeny , Travel , Viral Nonstructural Proteins/genetics , Viral Proteins/genetics , Viroporin Proteins/genetics
12.
Indian J Clin Biochem ; 36(4): 416-426, 2021 Oct.
Article in English | MEDLINE | ID: covidwho-1092844

ABSTRACT

Nutritional deficiency is associated with impaired immunity and increased susceptibility to infections. The complex interactions of trace elements with the macromolecules trigger the effective immune response against the viral diseases. The outcome of various viral infections along with susceptibility is affected by trace elements such as zinc, selenium, iron, copper, etc. due to their immuno-modulatory effects. Available electronic databases have been comprehensively searched for articles published with full text available and with the key words "Trace elements", "COVID-19", "Viral Infections" and "Immune Response" (i.e. separately Zn, Se, Fe, Cu, Mn, Mo, Cr, Li, Ni, Co) appearing in the title and abstract. On the basis of available articles we have explored the role of trace elements in viral infections with special reference to COVID-19 and their interactions with the immune system. Zinc, selenium and other trace elements are vital to triggerTH1 cells and cytokine-mediated immune response for substantial production of proinflammatory cytokines. The antiviral activity of some trace elements is attributed to their inhibitory effect on viral entry, replication and other downstream processes. Trace elements having antioxidants activity not only regulate host immune responses, but also modify the viral genome. Adequate dietary intake of trace elements is essential for activation, development, differentiation and numerous functions.

13.
J Gen Virol ; 102(3)2021 03.
Article in English | MEDLINE | ID: covidwho-1081877

ABSTRACT

The novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causing COVID-19 has rapidly turned into a pandemic, infecting millions and causing 1 157 509 (as of 27 October 2020) deaths across the globe. In addition to studying the mode of transmission and evasion of host immune system, analysing the viral mutational landscape constitutes an area under active research. The latter is expected to impart knowledge on the emergence of different clades, subclades, viral protein functions and protein-protein and protein-RNA interactions during replication/transcription cycle of virus and response to host immune checkpoints. In this study, we have attempted to bring forth the viral genomic variants defining the major clade(s) as identified from samples collected from the state of Telangana, India. We further report a comprehensive draft of all genomic variations (including unique mutations) present in SARS-CoV-2 strain in the state of Telangana. Our results reveal the presence of two mutually exclusive subgroups defined by specific variants within the dominant clade present in the population. This work attempts to bridge the critical gap regarding the genomic landscape and associate mutations in SARS-CoV-2 from a highly infected southern region of India, which was lacking to date.


Subject(s)
COVID-19/virology , Genome, Viral , SARS-CoV-2/genetics , COVID-19/epidemiology , Genomics , Humans , India/epidemiology , Mutation , Phylogeny , SARS-CoV-2/isolation & purification , Sequence Analysis, RNA , Viral Nonstructural Proteins/genetics , Viral Proteins/genetics
14.
Mol Biol Evol ; 38(6): 2428-2445, 2021 05 19.
Article in English | MEDLINE | ID: covidwho-1069279

ABSTRACT

COVID-19 can lead to acute respiratory syndrome, which can be due to dysregulated immune signaling. We analyze the distribution of CpG dinucleotides, a pathogen-associated molecular pattern, in the SARS-CoV-2 genome. We characterize CpG content by a CpG force that accounts for statistical constraints acting on the genome at the nucleotidic and amino acid levels. The CpG force, as the CpG content, is overall low compared with other pathogenic betacoronaviruses; however, it widely fluctuates along the genome, with a particularly low value, comparable with the circulating seasonal HKU1, in the spike coding region and a greater value, comparable with SARS and MERS, in the highly expressed nucleocapside coding region (N ORF), whose transcripts are relatively abundant in the cytoplasm of infected cells and present in the 3'UTRs of all subgenomic RNA. This dual nature of CpG content could confer to SARS-CoV-2 the ability to avoid triggering pattern recognition receptors upon entry, while eliciting a stronger response during replication. We then investigate the evolution of synonymous mutations since the outbreak of the COVID-19 pandemic, finding a signature of CpG loss in regions with a greater CpG force. Sequence motifs preceding the CpG-loss-associated loci in the N ORF match recently identified binding patterns of the zinc finger antiviral protein. Using a model of the viral gene evolution under human host pressure, we find that synonymous mutations seem driven in the SARS-CoV-2 genome, and particularly in the N ORF, by the viral codon bias, the transition-transversion bias, and the pressure to lower CpG content.


Subject(s)
COVID-19/genetics , CpG Islands , Evolution, Molecular , Genome, Viral , RNA, Viral/genetics , SARS-CoV-2 , Humans , SARS-CoV-2/genetics , SARS-CoV-2/pathogenicity
15.
mBio ; 12(1)2021 01 19.
Article in English | MEDLINE | ID: covidwho-1066817

ABSTRACT

Viral genome sequencing has guided our understanding of the spread and extent of genetic diversity of SARS-CoV-2 during the COVID-19 pandemic. SARS-CoV-2 viral genomes are usually sequenced from nasopharyngeal swabs of individual patients to track viral spread. Recently, RT-qPCR of municipal wastewater has been used to quantify the abundance of SARS-CoV-2 in several regions globally. However, metatranscriptomic sequencing of wastewater can be used to profile the viral genetic diversity across infected communities. Here, we sequenced RNA directly from sewage collected by municipal utility districts in the San Francisco Bay Area to generate complete and nearly complete SARS-CoV-2 genomes. The major consensus SARS-CoV-2 genotypes detected in the sewage were identical to clinical genomes from the region. Using a pipeline for single nucleotide variant calling in a metagenomic context, we characterized minor SARS-CoV-2 alleles in the wastewater and detected viral genotypes which were also found within clinical genomes throughout California. Observed wastewater variants were more similar to local California patient-derived genotypes than they were to those from other regions within the United States or globally. Additional variants detected in wastewater have only been identified in genomes from patients sampled outside California, indicating that wastewater sequencing can provide evidence for recent introductions of viral lineages before they are detected by local clinical sequencing. These results demonstrate that epidemiological surveillance through wastewater sequencing can aid in tracking exact viral strains in an epidemic context.


Subject(s)
COVID-19/virology , SARS-CoV-2/genetics , SARS-CoV-2/isolation & purification , Sewage/virology , Base Sequence , COVID-19/epidemiology , California/epidemiology , Environmental Microbiology , Genome, Viral , Genotype , Humans , Metagenome , Metagenomics , Polymorphism, Single Nucleotide , RNA, Viral/genetics , Real-Time Polymerase Chain Reaction , Transcriptome
16.
Virology ; 556: 62-72, 2021 04.
Article in English | MEDLINE | ID: covidwho-1065649

ABSTRACT

Members of the APOBEC family of cytidine deaminases show antiviral activities in mammalian cells through lethal editing in the genomes of small DNA viruses, herpesviruses and retroviruses, and potentially those of RNA viruses such as coronaviruses. Consistent with the latter, APOBEC-like directional C→U transitions of genomic plus-strand RNA are greatly overrepresented in SARS-CoV-2 genome sequences of variants emerging during the COVID-19 pandemic. A C→U mutational process may leave evolutionary imprints on coronavirus genomes, including extensive homoplasy from editing and reversion at targeted sites and the occurrence of driven amino acid sequence changes in viral proteins. If sustained over longer periods, this process may account for the previously reported marked global depletion of C and excess of U bases in human seasonal coronavirus genomes. This review synthesizes the current knowledge on APOBEC evolution and function and the evidence of their role in APOBEC-mediated genome editing of SARS-CoV-2 and other coronaviruses.


Subject(s)
APOBEC Deaminases/metabolism , Coronavirus/genetics , Evolution, Molecular , Genome, Viral/genetics , RNA Editing , APOBEC Deaminases/chemistry , APOBEC Deaminases/genetics , Animals , Coronavirus Infections/virology , Humans , Mutation , SARS-CoV-2/genetics
17.
Viruses ; 13(2)2021 02 02.
Article in English | MEDLINE | ID: covidwho-1059964

ABSTRACT

Infections with SARS-CoV-2 can progress toward multiple clinical outcomes, and the identification of factors associated with disease severity would represent a major advance to guide care and improve prognosis. We tested for associations between SARS-CoV-2 genomic variants from an international cohort of 2508 patients and mortality rates. Findings were validated in a second cohort. Phylogenetic analysis of SARS-CoV-2 genome sequences revealed four well-resolved clades which had significantly different mortality rates, even after adjusting for patient demographic and geographic characteristics. We further identified ten single-nucleotide polymorphisms (SNPs) in the SARS-CoV-2 genome that were associated with patient mortality. Three SNPs remained associated with mortality in a generalized linear model (GLM) that also included patient age, sex, geographic region, and month of sample collection. Multiple SNPs were confirmed in the validation cohort. These SNPs represent targets to assess the mechanisms underlying COVID-19 disease severity and warrant straightforward validation in functional studies.


Subject(s)
COVID-19/mortality , COVID-19/virology , Genome, Viral , Polymorphism, Single Nucleotide , SARS-CoV-2/genetics , Adult , Age Distribution , Aged , Aged, 80 and over , Cohort Studies , Female , Humans , Male , Middle Aged , Phylogeny
18.
Nucleic Acids Res ; 48(20): 11270-11283, 2020 11 18.
Article in English | MEDLINE | ID: covidwho-1060299

ABSTRACT

Specific elements of viral genomes regulate interactions within host cells. Here, we calculated the secondary structure content of >2000 coronaviruses and computed >100 000 human protein interactions with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The genomic regions display different degrees of conservation. SARS-CoV-2 domain encompassing nucleotides 22 500-23 000 is conserved both at the sequence and structural level. The regions upstream and downstream, however, vary significantly. This part of the viral sequence codes for the Spike S protein that interacts with the human receptor angiotensin-converting enzyme 2 (ACE2). Thus, variability of Spike S is connected to different levels of viral entry in human cells within the population. Our predictions indicate that the 5' end of SARS-CoV-2 is highly structured and interacts with several human proteins. The binding proteins are involved in viral RNA processing, include double-stranded RNA specific editases and ATP-dependent RNA-helicases and have strong propensity to form stress granules and phase-separated assemblies. We propose that these proteins, also implicated in viral infections such as HIV, are selectively recruited by SARS-CoV-2 genome to alter transcriptional and post-transcriptional regulation of host cells and to promote viral replication.


Subject(s)
Genome, Viral , Protein Interaction Maps , SARS-CoV-2/genetics , Angiotensin-Converting Enzyme 2/metabolism , COVID-19/virology , Humans , Protein Binding , SARS-CoV-2/pathogenicity , SARS-CoV-2/physiology , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/metabolism , Virulence/genetics , Virus Internalization , Virus Replication
19.
Front Genet ; 11: 572702, 2020.
Article in English | MEDLINE | ID: covidwho-1021887

ABSTRACT

The emergence of a new coronavirus (CoV), severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), responsible for severe respiratory disease in humans termed coronavirus disease of 2019 (COVID-19), became a new global threat for health and the economy. The SARS-CoV-2 genome is about a 29,800-nucleotide-long plus-strand RNA that can form functionally important secondary and higher-order structures called cis-acting RNA elements. These elements can interact with viral proteins, host proteins, or other RNAs and be involved in regulating translation and replication processes of the viral genome and encapsidation of the virus. However, the cis-acting RNA elements and their biological roles in SARS-CoV-2 as well as their comparative analysis in the closely related viral genome have not been well explored, which is very important to understand the molecular mechanism of viral infection and pathogenies. In this study, we used a bioinformatics approach to identify the cis-acting RNA elements in the SARS-CoV-2 genome. Initially, we aligned the full genomic sequence of six different CoVs, and a phylogenetic analysis was performed to understand their evolutionary relationship. Next, we predicted the cis-acting RNA elements in the SARS-CoV-2 genome using the structRNAfinder tool. Then, we annotated the location of these cis-acting RNA elements in different genomic regions of SARS-CoV-2. After that, we analyzed the sequence conservation patterns of each cis-acting RNA element among the six CoVs. Finally, the presence of cis-acting RNA elements across different CoV genomes and their comparative analysis was performed. Our study identified 12 important cis-acting RNA elements in the SARS-CoV-2 genome; among them, Corona_FSE, Corona_pk3, and s2m are highly conserved across most of the studied CoVs, and Thr_leader, MAT2A_D, and MS2 are uniquely present in SARS-CoV-2. These RNA structure elements can be involved in viral translation, replication, and encapsidation and, therefore, can be potential targets for better treatment of COVID-19. It is imperative to further characterize these cis-acting RNA elements experimentally for a better mechanistic understanding of SARS-CoV-2 infection and therapeutic intervention.

20.
Stem Cell Res Ther ; 11(1): 514, 2020 11 30.
Article in English | MEDLINE | ID: covidwho-949106

ABSTRACT

Coronavirus disease 2019 (COVID-19) caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a global pandemic. The prevalence/severity of COVID-19 is higher among patients with cardiovascular risk factors. Despite the expression of angiotensin-converting enzyme 2 (ACE2), a receptor for SARS-CoV-2 infection, in cardiomyocytes, there has been no conclusive evidence of direct viral infection although the presence of viral genome within COVID-19 patients' hearts has been reported. Here, we overexpressed SARS-CoV-2 genes in A549 lung epithelial cells. We then isolated extracellular vesicles (EVs) and detected the presence of viral RNA within these EVs. We observed that human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) are receptive to these EVs, and viral genes were detectable in the cardiomyocytes. Accordingly, the uptake of viral RNA-harboring EVs led to an upregulation of inflammation-related genes in hiPSC-CMs. Thus, our findings indicate that SARS-CoV-2 RNA containing EVs represents an indirect route of viral RNA entry into cardiomyocytes.


Subject(s)
COVID-19/virology , Extracellular Vesicles/virology , Myocytes, Cardiac/virology , SARS-CoV-2/pathogenicity , Virus Internalization , A549 Cells , Humans , Induced Pluripotent Stem Cells , RNA, Viral
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