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1.
Sci Rep ; 11(1): 19930, 2021 10 07.
Artigo em Inglês | MEDLINE | ID: mdl-34620923

RESUMO

Transmission of SARS-CoV-2 by aerosols has played a significant role in the rapid spread of COVID-19 across the globe. Indoor environments with inadequate ventilation pose a serious infection risk. Whilst vaccines suppress transmission, they are not 100% effective and the risk from variants and new viruses always remains. Consequently, many efforts have focused on ways to disinfect air. One such method involves use of minimally hazardous 222 nm far-UVC light. Whilst a small number of controlled experimental studies have been conducted, determining the efficacy of this approach is difficult because chamber or room geometry, and the air flow within them, influences both far-UVC illumination and aerosol dwell times. Fortunately, computational multiphysics modelling allows the inadequacy of dose-averaged assessment of viral inactivation to be overcome in these complex situations. This article presents the first validation of the WYVERN radiation-CFD code for far-UVC air-disinfection against survival fraction measurements, and the first measurement-informed modelling approach to estimating far-UVC susceptibility of viruses in air. As well as demonstrating the reliability of the code, at circa 70% higher, our findings indicate that aerosolized human coronaviruses are significantly more susceptible to far-UVC than previously thought.


Assuntos
Coronavirus Humano 229E/efeitos da radiação , Infecções por Coronavirus/prevenção & controle , Coronavirus Humano OC43/efeitos da radiação , Desinfecção/métodos , Raios Ultravioleta , Inativação de Vírus/efeitos da radiação , Aerossóis/isolamento & purificação , Microbiologia do Ar , COVID-19/prevenção & controle , Simulação por Computador , Coronavirus Humano 229E/isolamento & purificação , Coronavirus Humano 229E/fisiologia , Coronavirus Humano OC43/isolamento & purificação , Coronavirus Humano OC43/fisiologia , Desinfecção/instrumentação , Desenho de Equipamento , Humanos , Modelos Biológicos
2.
Viruses ; 13(8)2021 08 10.
Artigo em Inglês | MEDLINE | ID: mdl-34452443

RESUMO

The novel coronavirus SARS-CoV-2 is the seventh identified human coronavirus. Understanding the extent of pre-existing immunity induced by seropositivity to endemic seasonal coronaviruses and the impact of cross-reactivity on COVID-19 disease progression remains a key research question in immunity to SARS-CoV-2 and the immunopathology of COVID-2019 disease. This paper describes a panel of lentiviral pseudotypes bearing the spike (S) proteins for each of the seven human coronaviruses (HCoVs), generated under similar conditions optimized for high titre production allowing a high-throughput investigation of antibody neutralization breadth. Optimal production conditions and most readily available permissive target cell lines were determined for spike-mediated entry by each HCoV pseudotype: SARS-CoV-1, SARS-CoV-2 and HCoV-NL63 best transduced HEK293T/17 cells transfected with ACE2 and TMPRSS2, HCoV-229E and MERS-CoV preferentially entered HUH7 cells, and CHO cells were most permissive for the seasonal betacoronavirus HCoV-HKU1. Entry of ACE2 using pseudotypes was enhanced by ACE2 and TMPRSS2 expression in target cells, whilst TMPRSS2 transfection rendered HEK293T/17 cells permissive for HCoV-HKU1 and HCoV-OC43 entry. Additionally, pseudotype viruses were produced bearing additional coronavirus surface proteins, including the SARS-CoV-2 Envelope (E) and Membrane (M) proteins and HCoV-OC43/HCoV-HKU1 Haemagglutinin-Esterase (HE) proteins. This panel of lentiviral pseudotypes provides a safe, rapidly quantifiable and high-throughput tool for serological comparison of pan-coronavirus neutralizing responses; this can be used to elucidate antibody dynamics against individual coronaviruses and the effects of antibody cross-reactivity on clinical outcome following natural infection or vaccination.


Assuntos
Anticorpos Antivirais/imunologia , Anticorpos Amplamente Neutralizantes/imunologia , COVID-19/imunologia , Coronavirus/imunologia , SARS-CoV-2/imunologia , Glicoproteína da Espícula de Coronavírus/imunologia , Animais , Anticorpos Antivirais/sangue , Anticorpos Amplamente Neutralizantes/sangue , Linhagem Celular , Coronavirus Humano 229E/imunologia , Coronavirus Humano 229E/fisiologia , Coronavirus Humano NL63/imunologia , Coronavirus Humano NL63/fisiologia , Coronavirus Humano OC43/imunologia , Coronavirus Humano OC43/fisiologia , Reações Cruzadas , Humanos , Lentivirus/genética , Coronavírus da Síndrome Respiratória do Oriente Médio/imunologia , Coronavírus da Síndrome Respiratória do Oriente Médio/fisiologia , Testes de Neutralização , Plasmídeos , SARS-CoV-2/fisiologia , Transfecção , Internalização do Vírus
3.
Viruses ; 13(8)2021 08 23.
Artigo em Inglês | MEDLINE | ID: mdl-34452529

RESUMO

An escalating pandemic of the novel SARS-CoV-2 virus is impacting global health, and effective antivirals are needed. Umifenovir (Arbidol) is an indole-derivative molecule, licensed in Russia and China for prophylaxis and treatment of influenza and other respiratory viral infections. It has been shown that umifenovir has broad spectrum activity against different viruses. We evaluated the sensitivity of different coronaviruses, including the novel SARS-CoV-2 virus, to umifenovir using in vitro assays. Using a plaque assay, we revealed an antiviral effect of umifenovir against seasonal HCoV-229E and HCoV-OC43 coronaviruses in Vero E6 cells, with estimated 50% effective concentrations (EC50) of 10.0 ± 0.5 µM and 9.0 ± 0.4 µM, respectively. Umifenovir at 90 µM significantly suppressed plaque formation in CMK-AH-1 cells infected with SARS-CoV. Umifenovir also inhibited the replication of SARS-CoV-2 virus, with EC50 values ranging from 15.37 ± 3.6 to 28.0 ± 1.0 µM. In addition, 21-36 µM of umifenovir significantly suppressed SARS-CoV-2 virus titers (≥2 log TCID50/mL) in the first 24 h after infection. Repurposing of antiviral drugs is very helpful in fighting COVID-19. A safe, pan-antiviral drug such as umifenovir could be extremely beneficial in combating the early stages of a viral pandemic.


Assuntos
Antivirais/farmacologia , Coronavirus Humano 229E/efeitos dos fármacos , Coronavirus Humano OC43/efeitos dos fármacos , Indóis/farmacologia , Vírus da SARS/efeitos dos fármacos , SARS-CoV-2/efeitos dos fármacos , Animais , Antivirais/administração & dosagem , Sobrevivência Celular/efeitos dos fármacos , Chlorocebus aethiops , Coronavirus Humano 229E/fisiologia , Coronavirus Humano OC43/fisiologia , Efeito Citopatogênico Viral/efeitos dos fármacos , Humanos , Indóis/administração & dosagem , Testes de Sensibilidade Microbiana , Vírus da SARS/fisiologia , SARS-CoV-2/fisiologia , Células Vero , Carga Viral/efeitos dos fármacos , Ensaio de Placa Viral , Replicação Viral/efeitos dos fármacos
4.
J Photochem Photobiol B ; 222: 112282, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34404018

RESUMO

Emerging evidence suggests that blue light has the potential to inactivate viruses. Therefore, we investigated the effect of 405 nm, 410 nm, 425 nm and 450 nm pulsed blue light (PBL) on human alpha coronavirus HCoV-229 E and human beta coronavirus HCoV-OC43, using Qubit fluorometry and RT-LAMP to quantitate the amount of nucleic acid in irradiated and control samples. Like SARS-CoV-2, HCoV-229E and HCoV-OC43 are single stranded RNA viruses transmitted by air and direct contact; they have similar genomic sizes as SARS-CoV-2, and are used as surrogates for SARS-CoV-2. Irradiation was carried out either at 32.4 J cm-2 using 3 mW cm-2 irradiance or at 130 J cm-2 using 12 mW cm-2 irradiance. Results: (1) At each wavelength tested, PBL was antiviral against both coronaviruses. (2) 405 nm light gave the best result, yielding 52.3% (2.37 log10) inactivation against HCoV-OC43 (p < .0001), and a significant 1.46 log 10 (44%) inactivation of HCoV-229E (p < .01). HCoV-OC43, which like SARS-CoV-2 is a beta coronavirus, was more susceptible to PBL irradiation than alpha coronavirus HCoV-229E. The latter finding suggests that PBL is potentially antiviral against multiple coronavirus strains, and that, while its potency may vary from one virus to another, it seems more antiviral against beta coronaviruses, such as HCoV-OC43. (3) Further, the antiviral effect of PBL was better at a higher irradiance than a lower irradiance, and this indicates that with further refinement, a protocol capable of yielding 100% inactivation of viruses is attainable.


Assuntos
Coronavirus Humano 229E/efeitos da radiação , Coronavirus Humano OC43/efeitos da radiação , Terapia com Luz de Baixa Intensidade/métodos , SARS-CoV-2/efeitos da radiação , Coronavirus Humano 229E/fisiologia , Coronavirus Humano OC43/fisiologia , Relação Dose-Resposta à Radiação , Humanos , SARS-CoV-2/fisiologia
5.
Science ; 373(6557): 931-936, 2021 08 20.
Artigo em Inglês | MEDLINE | ID: mdl-34285133

RESUMO

There is an urgent need for antiviral agents that treat severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. We screened a library of 1900 clinically safe drugs against OC43, a human beta coronavirus that causes the common cold, and evaluated the top hits against SARS-CoV-2. Twenty drugs significantly inhibited replication of both viruses in cultured human cells. Eight of these drugs inhibited the activity of the SARS-CoV-2 main protease, 3CLpro, with the most potent being masitinib, an orally bioavailable tyrosine kinase inhibitor. X-ray crystallography and biochemistry show that masitinib acts as a competitive inhibitor of 3CLpro. Mice infected with SARS-CoV-2 and then treated with masitinib showed >200-fold reduction in viral titers in the lungs and nose, as well as reduced lung inflammation. Masitinib was also effective in vitro against all tested variants of concern (B.1.1.7, B.1.351, and P.1).


Assuntos
Antivirais/farmacologia , COVID-19/tratamento farmacológico , Proteases 3C de Coronavírus/antagonistas & inibidores , Coronavirus Humano OC43/efeitos dos fármacos , Inibidores de Cisteína Proteinase/farmacologia , SARS-CoV-2/efeitos dos fármacos , Tiazóis/farmacologia , Células A549 , Animais , Antivirais/química , Antivirais/metabolismo , Antivirais/uso terapêutico , COVID-19/virologia , Domínio Catalítico , Proteases 3C de Coronavírus/química , Proteases 3C de Coronavírus/metabolismo , Coronavirus Humano OC43/fisiologia , Inibidores de Cisteína Proteinase/química , Inibidores de Cisteína Proteinase/metabolismo , Células HEK293 , Humanos , Concentração Inibidora 50 , Camundongos , Camundongos Transgênicos , Testes de Sensibilidade Microbiana , SARS-CoV-2/enzimologia , SARS-CoV-2/fisiologia , Tiazóis/química , Tiazóis/metabolismo , Tiazóis/uso terapêutico , Carga Viral/efeitos dos fármacos , Replicação Viral/efeitos dos fármacos
6.
Genes Dev ; 35(13-14): 1005-1019, 2021 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-34168039

RESUMO

N6-methyladenosine (m6A) is an abundant internal RNA modification, influencing transcript fate and function in uninfected and virus-infected cells. Installation of m6A by the nuclear RNA methyltransferase METTL3 occurs cotranscriptionally; however, the genomes of some cytoplasmic RNA viruses are also m6A-modified. How the cellular m6A modification machinery impacts coronavirus replication, which occurs exclusively in the cytoplasm, is unknown. Here we show that replication of SARS-CoV-2, the agent responsible for the COVID-19 pandemic, and a seasonal human ß-coronavirus HCoV-OC43, can be suppressed by depletion of METTL3 or cytoplasmic m6A reader proteins YTHDF1 and YTHDF3 and by a highly specific small molecule METTL3 inhibitor. Reduction of infectious titer correlates with decreased synthesis of viral RNAs and the essential nucleocapsid (N) protein. Sites of m6A modification on genomic and subgenomic RNAs of both viruses were mapped by methylated RNA immunoprecipitation sequencing (meRIP-seq). Levels of host factors involved in m6A installation, removal, and recognition were unchanged by HCoV-OC43 infection; however, nuclear localization of METTL3 and cytoplasmic m6A readers YTHDF1 and YTHDF2 increased. This establishes that coronavirus RNAs are m6A-modified and host m6A pathway components control ß-coronavirus replication. Moreover, it illustrates the therapeutic potential of targeting the m6A pathway to restrict coronavirus reproduction.


Assuntos
Coronavirus Humano OC43/fisiologia , Processamento Pós-Transcricional do RNA/genética , SARS-CoV-2/fisiologia , Replicação Viral/genética , Adenosina/análogos & derivados , Adenosina/genética , Adenosina/metabolismo , Linhagem Celular , Infecções por Coronavirus/metabolismo , Infecções por Coronavirus/virologia , Regulação da Expressão Gênica/efeitos dos fármacos , Interações Hospedeiro-Patógeno/efeitos dos fármacos , Humanos , Metiltransferases/antagonistas & inibidores , Metiltransferases/metabolismo , Proteínas do Nucleocapsídeo , RNA Viral/metabolismo , Proteínas de Ligação a RNA/metabolismo , Replicação Viral/efeitos dos fármacos
7.
mSphere ; 6(3)2021 05 12.
Artigo em Inglês | MEDLINE | ID: mdl-33980675

RESUMO

Human coronavirus (HCoV)-OC43 rarely shows a cytopathic effect (CPE) after infection of various cell lines, and the indirect immunoperoxidase assay (IPA), a relatively complex procedure, has long been used as an alternative assay. Because HCoV-OC43 uses cell-surface transmembrane protease serine 2 (TMPRSS2) for cell entry, VeroE6 cells expressing TMPRSS2 may show a clear CPE after HCoV-OC43 infection. The aim of this study was to construct a 50% tissue culture infectious dose (TCID50) assay for HCoV-OC43 based on CPE evaluation using VeroE6/TMPRSS2 cells. VeroE6/TMPRSS2 cells showed clear CPEs 3 to 4 days after low-titer HCoV-OC43 infection. Evaluation of viral kinetics indicated that the viral titer in the culture supernatant of VeroE6/TMPRSS2 cells in the early stages of infection was higher than that of other cells. In comparison, between the CPE-based and the IPA-based (i.e., the reference titer) methods, the titer measured with CPE evaluation 4 to 5 days after infection using VeroE6/TMPRSS2 cells showed a much smaller difference from the reference titer than that measured using other cells. Thus, the TCID50 assay using CPE evaluation with VeroE6/TMPRSS2 cells provides the correct titer value and will greatly contribute to future research on HCoV-OC43.IMPORTANCE HCoV-OC43 rarely shows a cytopathic effect (CPE) in infected cell lines, and thus the plaque and TCID50 assays by CPE observation are not applicable for titration; the indirect immunoperoxidase assay (IPA) is used instead. However, the IPA is relatively complex, time-consuming, costly, and not suitable for simultaneous titration of many samples. We developed a TCID50 assay using CPE evaluation with TMPRSS2-expressing VeroE6/TMPRSS2 cells that provides the same accuracy as the conventional IPA-based viral titration and does not require any staining procedures using antibodies or substrates. This titration method will greatly contribute to future research on HCoV-OC43 by allowing simple, low-cost, and accurate titration of this virus.


Assuntos
Coronavirus Humano OC43/fisiologia , Efeito Citopatogênico Viral , Receptores Virais/metabolismo , Serina Endopeptidases/metabolismo , Carga Viral/métodos , Animais , Linhagem Celular Tumoral , Chlorocebus aethiops , Coronavirus Humano OC43/isolamento & purificação , Humanos , Técnicas Imunoenzimáticas , Receptores Virais/genética , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Serina Endopeptidases/genética , Células Vero/virologia , Cultura de Vírus , Internalização do Vírus , Replicação Viral
8.
Viruses ; 13(2)2021 02 03.
Artigo em Inglês | MEDLINE | ID: mdl-33546185

RESUMO

The long-term control strategy of SARS-CoV-2 and other major respiratory viruses needs to include antivirals to treat acute infections, in addition to the judicious use of effective vaccines. Whilst COVID-19 vaccines are being rolled out for mass vaccination, the modest number of antivirals in use or development for any disease bears testament to the challenges of antiviral development. We recently showed that non-cytotoxic levels of thapsigargin (TG), an inhibitor of the sarcoplasmic/endoplasmic reticulum (ER) Ca2+ ATPase pump, induces a potent host innate immune antiviral response that blocks influenza A virus replication. Here we show that TG is also highly effective in blocking the replication of respiratory syncytial virus (RSV), common cold coronavirus OC43, SARS-CoV-2 and influenza A virus in immortalized or primary human cells. TG's antiviral performance was significantly better than remdesivir and ribavirin in their respective inhibition of OC43 and RSV. Notably, TG was just as inhibitory to coronaviruses (OC43 and SARS-CoV-2) and influenza viruses (USSR H1N1 and pdm 2009 H1N1) in separate infections as in co-infections. Post-infection oral gavage of acid-stable TG protected mice against a lethal influenza virus challenge. Together with its ability to inhibit the different viruses before or during active infection, and with an antiviral duration of at least 48 h post-TG exposure, we propose that TG (or its derivatives) is a promising broad-spectrum inhibitor against SARS-CoV-2, OC43, RSV and influenza virus.


Assuntos
Antivirais/farmacologia , Betacoronavirus/efeitos dos fármacos , Coronavirus Humano OC43/efeitos dos fármacos , Vírus da Influenza A Subtipo H1N1/efeitos dos fármacos , Vírus Sincicial Respiratório Humano/efeitos dos fármacos , SARS-CoV-2/efeitos dos fármacos , Tapsigargina/farmacologia , Animais , Antivirais/uso terapêutico , Betacoronavirus/fisiologia , Linhagem Celular , Linhagem Celular Tumoral , Células Cultivadas , Coronavirus Humano OC43/fisiologia , Estresse do Retículo Endoplasmático , Humanos , Vírus da Influenza A Subtipo H1N1/fisiologia , Camundongos , Testes de Sensibilidade Microbiana , Infecções por Orthomyxoviridae/tratamento farmacológico , Infecções por Orthomyxoviridae/virologia , Vírus Sincicial Respiratório Humano/fisiologia , Ribavirina/farmacologia , SARS-CoV-2/fisiologia , Tapsigargina/uso terapêutico , Replicação Viral/efeitos dos fármacos
9.
Biochem Biophys Res Commun ; 547: 23-28, 2021 04 02.
Artigo em Inglês | MEDLINE | ID: mdl-33588235

RESUMO

COVID-19 pandemic results in record high deaths in many countries. Although a vaccine for SARS-CoV-2 is now available, effective antiviral drugs to treat coronavirus diseases are not available yet. Recently, EGCG, a green tea polyphenol, was reported to inhibit SARS-CoV-2 3CL-protease, however the effect of EGCG on coronavirus replication is unknown. In this report, human coronavirus HCoV-OC43 (beta coronavirus) and HCoV-229E (alpha coronavirus) were used to examine the effect of EGCG on coronavirus. EGCG treatment decreases 3CL-protease activity of HCoV-OC43 and HCoV-229E. Moreover, EGCG treatment decreased HCoV-OC43-induced cytotoxicity. Finally, we found that EGCG treatment decreased the levels of coronavirus RNA and protein in infected cell media. These results indicate that EGCG inhibits coronavirus replication.


Assuntos
Coronavirus Humano 229E/efeitos dos fármacos , Coronavirus Humano OC43/efeitos dos fármacos , Polifenóis/farmacologia , Chá/química , Replicação Viral/efeitos dos fármacos , Sequência de Aminoácidos , Linhagem Celular Tumoral , Coronavirus Humano 229E/fisiologia , Proteases 3C de Coronavírus/antagonistas & inibidores , Coronavirus Humano OC43/fisiologia , Humanos , SARS-CoV-2/efeitos dos fármacos , SARS-CoV-2/fisiologia
10.
Cell ; 184(1): 120-132.e14, 2021 01 07.
Artigo em Inglês | MEDLINE | ID: mdl-33382968

RESUMO

The coronavirus disease 2019 (COVID-19) pandemic has claimed the lives of over one million people worldwide. The causative agent, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is a member of the Coronaviridae family of viruses that can cause respiratory infections of varying severity. The cellular host factors and pathways co-opted during SARS-CoV-2 and related coronavirus life cycles remain ill defined. To address this gap, we performed genome-scale CRISPR knockout screens during infection by SARS-CoV-2 and three seasonal coronaviruses (HCoV-OC43, HCoV-NL63, and HCoV-229E). These screens uncovered host factors and pathways with pan-coronavirus and virus-specific functional roles, including major dependency on glycosaminoglycan biosynthesis, sterol regulatory element-binding protein (SREBP) signaling, bone morphogenetic protein (BMP) signaling, and glycosylphosphatidylinositol biosynthesis, as well as a requirement for several poorly characterized proteins. We identified an absolute requirement for the VMP1, TMEM41, and TMEM64 (VTT) domain-containing protein transmembrane protein 41B (TMEM41B) for infection by SARS-CoV-2 and three seasonal coronaviruses. This human coronavirus host factor compendium represents a rich resource to develop new therapeutic strategies for acute COVID-19 and potential future coronavirus pandemics.


Assuntos
Infecções por Coronavirus/genética , Estudo de Associação Genômica Ampla , SARS-CoV-2/fisiologia , Células A549 , Linhagem Celular , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Coronavirus Humano 229E/fisiologia , Infecções por Coronavirus/virologia , Coronavirus Humano NL63/fisiologia , Coronavirus Humano OC43/fisiologia , Técnicas de Inativação de Genes , Células HEK293 , Interações Hospedeiro-Patógeno/efeitos dos fármacos , Humanos , Proteínas de Membrana/metabolismo , Redes e Vias Metabólicas/efeitos dos fármacos , Mapeamento de Interação de Proteínas
11.
Proc Natl Acad Sci U S A ; 117(41): 25759-25770, 2020 10 13.
Artigo em Inglês | MEDLINE | ID: mdl-32994342

RESUMO

Human coronaviruses OC43 and HKU1 are respiratory pathogens of zoonotic origin that have gained worldwide distribution. OC43 apparently emerged from a bovine coronavirus (BCoV) spillover. All three viruses attach to 9-O-acetylated sialoglycans via spike protein S with hemagglutinin-esterase (HE) acting as a receptor-destroying enzyme. In BCoV, an HE lectin domain promotes esterase activity toward clustered substrates. OC43 and HKU1, however, lost HE lectin function as an adaptation to humans. Replaying OC43 evolution, we knocked out BCoV HE lectin function and performed forced evolution-population dynamics analysis. Loss of HE receptor binding selected for second-site mutations in S, decreasing S binding affinity by orders of magnitude. Irreversible HE mutations led to cooperativity in virus swarms with low-affinity S minority variants sustaining propagation of high-affinity majority phenotypes. Salvageable HE mutations induced successive second-site substitutions in both S and HE. Apparently, S and HE are functionally interdependent and coevolve to optimize the balance between attachment and release. This mechanism of glycan-based receptor usage, entailing a concerted, fine-tuned activity of two envelope protein species, is unique among CoVs, but reminiscent of that of influenza A viruses. Apparently, general principles fundamental to virion-sialoglycan interactions prompted convergent evolution of two important groups of human and animal pathogens.


Assuntos
Coronavirus/fisiologia , Hemaglutininas Virais/genética , Glicoproteína da Espícula de Coronavírus/genética , Proteínas Virais de Fusão/genética , Vírion/metabolismo , Animais , Evolução Biológica , Linhagem Celular , Coronavirus/genética , Coronavirus/metabolismo , Infecções por Coronavirus/virologia , Coronavirus Humano OC43/genética , Coronavirus Humano OC43/metabolismo , Coronavirus Humano OC43/fisiologia , Coronavirus Bovino/genética , Coronavirus Bovino/metabolismo , Coronavirus Bovino/fisiologia , Hemaglutininas Virais/química , Hemaglutininas Virais/metabolismo , Humanos , Lectinas/genética , Lectinas/metabolismo , Camundongos , Mutação , Ligação Proteica , Domínios Proteicos , Receptores Virais/metabolismo , Seleção Genética , Ácidos Siálicos/metabolismo , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/metabolismo , Proteínas Virais de Fusão/química , Proteínas Virais de Fusão/metabolismo , Vírion/genética , Ligação Viral , Liberação de Vírus
12.
Science ; 368(6493): 860-868, 2020 05 22.
Artigo em Inglês | MEDLINE | ID: mdl-32291278

RESUMO

It is urgent to understand the future of severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2) transmission. We used estimates of seasonality, immunity, and cross-immunity for human coronavirus OC43 (HCoV-OC43) and HCoV-HKU1 using time-series data from the United States to inform a model of SARS-CoV-2 transmission. We projected that recurrent wintertime outbreaks of SARS-CoV-2 will probably occur after the initial, most severe pandemic wave. Absent other interventions, a key metric for the success of social distancing is whether critical care capacities are exceeded. To avoid this, prolonged or intermittent social distancing may be necessary into 2022. Additional interventions, including expanded critical care capacity and an effective therapeutic, would improve the success of intermittent distancing and hasten the acquisition of herd immunity. Longitudinal serological studies are urgently needed to determine the extent and duration of immunity to SARS-CoV-2. Even in the event of apparent elimination, SARS-CoV-2 surveillance should be maintained because a resurgence in contagion could be possible as late as 2024.


Assuntos
Betacoronavirus/fisiologia , Infecções por Coronavirus/virologia , Modelos Biológicos , Pneumonia Viral/virologia , Betacoronavirus/imunologia , COVID-19 , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/transmissão , Coronavirus Humano OC43/fisiologia , Surtos de Doenças , Transmissão de Doença Infecciosa , Humanos , Pandemias , Pneumonia Viral/epidemiologia , Pneumonia Viral/transmissão , SARS-CoV-2 , Estações do Ano
13.
Infect Dis (Lond) ; 52(6): 419-422, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32067542

RESUMO

A child with pre-B acute lymphoblastic leukaemia (ALL) developed fatal encephalitis associated with human coronavirus OC43 (HCoV-OC43). During chemotherapy the child had a persistent HCoV-OC43 respiratory infection and later developed progressive encephalitis. Cerebrospinal fluid was negative for pathogens including HCoV-OC43, but a brain biopsy was HCoV-OC43-positive by metagenomic next-generation sequencing.


Assuntos
Infecções por Coronavirus/virologia , Coronavirus Humano OC43/fisiologia , Encefalite/virologia , Encéfalo/patologia , Encéfalo/virologia , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/mortalidade , Infecções por Coronavirus/patologia , Coronavirus Humano OC43/isolamento & purificação , Encefalite/imunologia , Encefalite/mortalidade , Encefalite/patologia , Evolução Fatal , Humanos , Hospedeiro Imunocomprometido , Lactente
14.
J Clin Virol ; 122: 104206, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31783264

RESUMO

BACKGROUND: While respiratory viral infections are recognized as a frequent cause of illness in hematopoietic stem cell transplantation (HSCT) recipients, HCoV-OC43 infections have rarely been investigated as healthcare-associated infections in this population. OBJECTIVES: In this report, HCoV-OC43 isolates collected from HSCT patients were retrospectively characterized to identify potential clusters of infection that may stand for a hospital transmission. STUDY DESIGN: Whole-genome and S gene sequences were obtained from nasal swabs using next-generation sequencing and phylogenetic trees were constructed. Similar identity matrix and determination of the most common ancestor were used to compare clusters of patient's sequences. Amino acids substitutions were analysed. RESULTS: Genotypes B, E, F and G were identified. Two clusters of patients were defined from chronological data and phylogenetic trees. Analyses of amino acids substitutions of the S protein sequences identified substitutions specific for genotype F strains circulating among European people. CONCLUSIONS: HCoV-OC43 may be implicated in healthcare-associated infections.


Assuntos
Infecções por Coronavirus/virologia , Coronavirus Humano OC43/genética , Infecção Hospitalar/virologia , Genoma Viral/genética , Adulto , Idoso , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/transmissão , Coronavirus Humano OC43/isolamento & purificação , Coronavirus Humano OC43/fisiologia , Infecção Hospitalar/epidemiologia , Infecção Hospitalar/transmissão , Europa (Continente)/epidemiologia , Feminino , Genótipo , Transplante de Células-Tronco Hematopoéticas , Humanos , Masculino , Pessoa de Meia-Idade , Epidemiologia Molecular , Filogenia , Estudos Retrospectivos , Sequenciamento Completo do Genoma , Adulto Jovem
15.
Antiviral Res ; 173: 104646, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31705922

RESUMO

Human coronaviruses (HCoVs) are important pathogens that cause upper respiratory tract infections and have neuroinvasive abilities; however, little is known about the dynamic infection process of CoVs in vivo, and there are currently no specific antiviral drugs to prevent or treat HCoV infection. Here, we verified the replication ability and pathogenicity of a reporter HCoV-OC43 strain expressing Renilla luciferase (Rluc; rOC43-ns2DelRluc) in mice with different genetic backgrounds (C57BL/6 and BALB/c). Additionally, we monitored the spatial and temporal progression of HCoV-OC43 through the central nervous system (CNS) of live BALB/c mice after intranasal or intracerebral inoculation with rOC43-ns2DelRluc. We found that rOC43-ns2DelRluc was fatal to suckling mice after intranasal inoculation, and that viral titers and Rluc expression were detected in the brains and spinal cords of mice infected with rOC43-ns2DelRluc. Moreover, viral replication was initially observed in the brain by non-invasive bioluminescence imaging before the infection spread to the spinal cord of BALB/c mice, consistent with its tropism in the CNS. Furthermore, the Rluc readout correlated with the HCoV replication ability and protein expression, which allowed quantification of antiviral activity in live mice. Additionally, we validated that chloroquine strongly inhibited rOC43-ns2DelRluc replication in vivo. These results provide new insights into the temporal and spatial dissemination of HCoV-OC43 in the CNS, and our methods provide an extremely sensitive platform for evaluating the efficacy of antiviral therapies to treat neuroinvasive HCoVs in live mice.


Assuntos
Sistema Nervoso Central/virologia , Infecções por Coronavirus/virologia , Coronavirus Humano OC43/fisiologia , Animais , Antivirais/administração & dosagem , Encéfalo/diagnóstico por imagem , Encéfalo/virologia , Sistema Nervoso Central/diagnóstico por imagem , Cloroquina/administração & dosagem , Infecções por Coronavirus/diagnóstico por imagem , Infecções por Coronavirus/tratamento farmacológico , Coronavirus Humano OC43/genética , Genes Reporter , Humanos , Luciferases de Renilla/genética , Luciferases de Renilla/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Imagem Molecular , Replicação Viral/efeitos dos fármacos
16.
Biomolecules ; 9(11)2019 11 04.
Artigo em Inglês | MEDLINE | ID: mdl-31690059

RESUMO

Stephania tetrandra and other related species of Menispermaceae are the major sources of the bis-benzylisoquinoline alkaloids tetrandrine (TET), fangchinoline (FAN), and cepharanthine (CEP). Although the pharmacological properties of these compounds include anticancer and anti-inflammatory activities, the antiviral effects of these compounds against human coronavirus (HCoV) remain unclear. Hence, the aims of the current study were to assess the antiviral activities of TET, FAN, and CEP and to elucidate the underlying mechanisms in HCoV-OC43-infected MRC-5 human lung cells. These compounds significantly inhibited virus-induced cell death at the early stage of virus infection. TET, FAN, and CEP treatment dramatically suppressed the replication of HCoV-OC43 as well as inhibited viral S and N protein expression. The virus-induced host response was reduced by compound treatment as compared with the vehicle control. Taken together, these findings demonstrate that TET, FAN, and CEP are potential natural antiviral agents for the prevention and treatment of HCoV-OC43 infection.


Assuntos
Antivirais/farmacologia , Benzilisoquinolinas/farmacologia , Infecções por Coronavirus/virologia , Coronavirus Humano OC43/efeitos dos fármacos , Coronavirus Humano OC43/fisiologia , Extratos Vegetais/farmacologia , Stephania tetrandra/química , Benzilisoquinolinas/química , Linhagem Celular , Infecções por Coronavirus/genética , Infecções por Coronavirus/metabolismo , Coronavirus Humano OC43/genética , Citocinas/genética , Citocinas/metabolismo , Humanos , Extratos Vegetais/química
17.
Nat Struct Mol Biol ; 26(6): 481-489, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31160783

RESUMO

Coronaviruses cause respiratory tract infections in humans and outbreaks of deadly pneumonia worldwide. Infections are initiated by the transmembrane spike (S) glycoprotein, which binds to host receptors and fuses the viral and cellular membranes. To understand the molecular basis of coronavirus attachment to oligosaccharide receptors, we determined cryo-EM structures of coronavirus OC43 S glycoprotein trimer in isolation and in complex with a 9-O-acetylated sialic acid. We show that the ligand binds with fast kinetics to a surface-exposed groove and that interactions at the identified site are essential for S-mediated viral entry into host cells, but free monosaccharide does not trigger fusogenic conformational changes. The receptor-interacting site is conserved in all coronavirus S glycoproteins that engage 9-O-acetyl-sialogycans, with an architecture similar to those of the ligand-binding pockets of coronavirus hemagglutinin esterases and influenza virus C/D hemagglutinin-esterase fusion glycoproteins. Our results demonstrate these viruses evolved similar strategies to engage sialoglycans at the surface of target cells.


Assuntos
Infecções por Coronavirus/metabolismo , Coronavirus Humano OC43/fisiologia , Ácido N-Acetilneuramínico/metabolismo , Receptores de Superfície Celular/metabolismo , Glicoproteína da Espícula de Coronavírus/metabolismo , Infecções por Coronavirus/virologia , Coronavirus Humano OC43/química , Microscopia Crioeletrônica , Células HEK293 , Humanos , Modelos Moleculares , Ácido N-Acetilneuramínico/análogos & derivados , Multimerização Proteica , Glicoproteína da Espícula de Coronavírus/química , Internalização do Vírus
18.
Viruses ; 11(4)2019 04 05.
Artigo em Inglês | MEDLINE | ID: mdl-30959796

RESUMO

Despite high similarity of canine respiratory coronavirus (CRCoV), bovine coronavirus, (BCoV) and human coronavirus OC43 (HCoV-OC43), these viruses differ in species specificity. For years it was believed that they share receptor specificity, utilizing sialic acids for cell surface attachment, internalization, and entry. Interestingly, careful literature analysis shows that viruses indeed bind to the cell surface via sialic acids, but there is no solid data that these moieties mediate virus entry. In our study, using a number of techniques, we showed that all three viruses are indeed able to bind to sialic acids to a different extent, but these molecules render the cells permissive only for the clinical strain of HCoV-OC43, while for others they serve only as attachment receptors. CRCoV and BCoV appear to employ human leukocyte antigen class I (HLA-1) as the entry receptor. Furthermore, we identified heparan sulfate as an alternative attachment factor, but this may be related to the cell culture adaptation, as in ex vivo conditions, it does not seem to play a significant role. Summarizing, we delineated early events during CRCoV, BCoV, and HCoV-OC43 entry and systematically studied the attachment and entry receptor utilized by these viruses.


Assuntos
Coronavirus Humano OC43/fisiologia , Coronavirus Bovino/fisiologia , Coronavirus Canino/fisiologia , Receptores Virais/análise , Ligação Viral , Células Cultivadas , Heparitina Sulfato/metabolismo , Antígenos de Histocompatibilidade Classe I/metabolismo , Humanos , Ácidos Siálicos/metabolismo
19.
Proc Natl Acad Sci U S A ; 116(7): 2681-2690, 2019 02 12.
Artigo em Inglês | MEDLINE | ID: mdl-30679277

RESUMO

Human betacoronaviruses OC43 and HKU1 are endemic respiratory pathogens and, while related, originated from independent zoonotic introductions. OC43 is in fact a host-range variant of the species Betacoronavirus-1, and more closely related to bovine coronavirus (BCoV)-its presumptive ancestor-and porcine hemagglutinating encephalomyelitis virus (PHEV). The ß1-coronaviruses (ß1CoVs) and HKU1 employ glycan-based receptors carrying 9-O-acetylated sialic acid (9-O-Ac-Sia). Receptor binding is mediated by spike protein S, the main determinant of coronavirus host specificity. For BCoV, a crystal structure for the receptor-binding domain S1A is available and for HKU1 a cryoelectron microscopy structure of the complete S ectodomain. However, the location of the receptor-binding site (RBS), arguably the single-most important piece of information, is unknown. Here we solved the 3.0-Å crystal structure of PHEV S1A We then took a comparative structural analysis approach to map the ß1CoV S RBS, using the general design of 9-O-Ac-Sia-binding sites as blueprint, backed-up by automated ligand docking, structure-guided mutagenesis of OC43, BCoV, and PHEV S1A, and infectivity assays with BCoV-S-pseudotyped vesicular stomatitis viruses. The RBS is not exclusive to OC43 and related animal viruses, but is apparently conserved and functional also in HKU1 S1A The binding affinity of the HKU1 S RBS toward short sialoglycans is significantly lower than that of OC43, which we attribute to differences in local architecture and accessibility, and which may be indicative for differences between the two viruses in receptor fine-specificity. Our findings challenge reports that would map the OC43 RBS elsewhere in S1A and that of HKU1 in domain S1B.


Assuntos
Coronavirus Humano OC43/fisiologia , Fusão de Membrana , Ácido N-Acetilneuramínico/metabolismo , Receptores Virais/metabolismo , Glicoproteína da Espícula de Coronavírus/metabolismo , Acetilação , Animais , Sítios de Ligação , Humanos , Ratos , Receptores Virais/química
20.
J Virol ; 92(17)2018 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-29925652

RESUMO

Human coronaviruses (HCoVs) are recognized respiratory pathogens for which accumulating evidence indicates that in vulnerable patients the infection can cause more severe pathologies. HCoVs are not always confined to the upper respiratory tract and can invade the central nervous system (CNS) under still unclear circumstances. HCoV-induced neuropathologies in humans are difficult to diagnose early enough to allow therapeutic interventions. Making use of our already described animal model of HCoV neuropathogenesis, we describe the route of neuropropagation from the nasal cavity to the olfactory bulb and piriform cortex and then the brain stem. We identified neuron-to-neuron propagation as one underlying mode of virus spreading in cell culture. Our data demonstrate that both passive diffusion of released viral particles and axonal transport are valid propagation strategies used by the virus. We describe for the first time the presence along axons of viral platforms whose static dynamism is reminiscent of viral assembly sites. We further reveal that HCoV OC43 modes of propagation can be modulated by selected HCoV OC43 proteins and axonal transport. Our work, therefore, identifies processes that may govern the severity and nature of HCoV OC43 neuropathogenesis and will make possible the development of therapeutic strategies to prevent occurrences.IMPORTANCE Coronaviruses may invade the CNS, disseminate, and participate in the induction of neurological diseases. Their neuropathogenicity is being increasingly recognized in humans, and the presence and persistence of human coronaviruses (HCoV) in human brains have been proposed to cause long-term sequelae. Using our mouse model relying on natural susceptibility to HCoV OC43 and neuronal cell cultures, we have defined the most relevant path taken by HCoV OC43 to access and spread to and within the CNS toward the brain stem and spinal cord and studied in cell culture the underlying modes of intercellular propagation to better understand its neuropathogenesis. Our data suggest that axonal transport governs HCoV OC43 egress in the CNS, leading to the exacerbation of neuropathogenesis. Exploiting knowledge on neuroinvasion and dissemination will enhance our ability to control viral infection within the CNS, as it will shed light on underlying mechanisms of neuropathogenesis and uncover potential druggable molecular virus-host interfaces.


Assuntos
Axônios/metabolismo , Infecções por Coronavirus/virologia , Coronavirus Humano OC43/fisiologia , Animais , Axônios/virologia , Infecções por Coronavirus/metabolismo , Humanos , Camundongos , Cavidade Nasal/metabolismo , Cavidade Nasal/virologia , Bulbo Olfatório/metabolismo , Bulbo Olfatório/virologia , Córtex Piriforme/metabolismo , Córtex Piriforme/virologia , Proteínas Virais/metabolismo , Montagem de Vírus
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