Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 19 de 19
Filtrar
Mais filtros










Base de dados
Intervalo de ano de publicação
1.
J Gen Virol ; 2020 Jun 22.
Artigo em Inglês | MEDLINE | ID: mdl-32568027

RESUMO

The sudden emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) at the end of 2019 from the Chinese province of Hubei and its subsequent pandemic spread highlight the importance of understanding the full molecular details of coronavirus infection and pathogenesis. Here, we compared a variety of replication features of SARS-CoV-2 and SARS-CoV and analysed the cytopathology caused by the two closely related viruses in the commonly used Vero E6 cell line. Compared to SARS-CoV, SARS-CoV-2 generated higher levels of intracellular viral RNA, but strikingly about 50-fold less infectious viral progeny was recovered from the culture medium. Immunofluorescence microscopy of SARS-CoV-2-infected cells established extensive cross-reactivity of antisera previously raised against a variety of non-structural proteins, membrane and nucleocapsid protein of SARS-CoV. Electron microscopy revealed that the ultrastructural changes induced by the two SARS viruses are very similar and occur within comparable time frames after infection. Furthermore, we determined that the sensitivity of the two viruses to three established inhibitors of coronavirus replication (remdesivir, alisporivir and chloroquine) is very similar, but that SARS-CoV-2 infection was substantially more sensitive to pre-treatment of cells with pegylated interferon alpha. An important difference between the two viruses is the fact that - upon passaging in Vero E6 cells - SARS-CoV-2 apparently is under strong selection pressure to acquire adaptive mutations in its spike protein gene. These mutations change or delete a putative furin-like cleavage site in the region connecting the S1 and S2 domains and result in a very prominent phenotypic change in plaque assays.

2.
Antiviral Res ; 178: 104787, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32251768

RESUMO

Although several clinical trials are now underway to test possible therapies, the worldwide response to the COVID-19 outbreak has been largely limited to monitoring/containment. We report here that Ivermectin, an FDA-approved anti-parasitic previously shown to have broad-spectrum anti-viral activity in vitro, is an inhibitor of the causative virus (SARS-CoV-2), with a single addition to Vero-hSLAM cells 2 h post infection with SARS-CoV-2 able to effect ~5000-fold reduction in viral RNA at 48 h. Ivermectin therefore warrants further investigation for possible benefits in humans.


Assuntos
Antivirais/farmacologia , Betacoronavirus/efeitos dos fármacos , Infecções por Coronavirus/virologia , Aprovação de Drogas , Ivermectina/farmacologia , Ivermectina/uso terapêutico , Pneumonia Viral/virologia , Replicação Viral/efeitos dos fármacos , Animais , Austrália , Betacoronavirus/fisiologia , Chlorocebus aethiops , Infecções por Coronavirus/tratamento farmacológico , Humanos , Pandemias , Pneumonia Viral/tratamento farmacológico , Células Vero
4.
Med J Aust ; 212(10): 459-462, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32237278

RESUMO

OBJECTIVES: To describe the first isolation and sequencing of SARS-CoV-2 in Australia and rapid sharing of the isolate. SETTING: SARS-CoV-2 was isolated from a 58-year-old man from Wuhan, China who arrived in Melbourne on 19 January 2020 and was admitted to the Monash Medical Centre, Melbourne from the emergency department on 24 January 2020 with fever, cough, and progressive dyspnoea. MAJOR OUTCOMES: Clinical course and laboratory features of the first reported case of COVID-19 (the illness caused by SARS-CoV-2) in Australia; isolation, whole genome sequencing, imaging, and rapid sharing of virus from the patient. RESULTS: A nasopharyngeal swab and sputum collected when the patient presented to hospital were each positive for SARS-CoV-2 (reverse transcription polymerase chain reaction). Inoculation of Vero/hSLAM cells with material from the nasopharyngeal swab led to the isolation of SARS-CoV-2 virus in culture. Electron microscopy of the supernatant confirmed the presence of virus particles with morphology characteristic of viruses of the family Coronaviridae. Whole genome sequencing of the viral isolate and phylogenetic analysis indicated the isolate exhibited greater than 99.99% sequence identity with other publicly available SARS-CoV-2 genomes. Within 24 hours of isolation, the first Australian SARS-CoV-2 isolate was shared with local and overseas reference laboratories and major North American and European culture collections. CONCLUSIONS: The ability to rapidly identify, propagate, and internationally share our SARS-CoV-2 isolate is an important step in collaborative scientific efforts to deal effectively with this international public health emergency by developing better diagnostic procedures, vaccine candidates, and antiviral agents.


Assuntos
Betacoronavirus/genética , Infecções por Coronavirus/genética , Disseminação de Informação/métodos , Isolamento de Pacientes/métodos , Pneumonia Viral/genética , Austrália , Infecções por Coronavirus/diagnóstico , Humanos , Masculino , Pessoa de Meia-Idade , Pandemias , Pneumonia Viral/diagnóstico , Sequenciamento Completo do Genoma
6.
Emerg Infect Dis ; 25(12): 2266-2269, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31742504

RESUMO

We report a case of Barmah Forest virus infection in a child from Central Province, Papua New Guinea, who had no previous travel history. Genomic characterization of the virus showed divergent origin compared with viruses previously detected, supporting the hypothesis that the range of Barmah Forest virus extends beyond Australia.


Assuntos
Infecções por Alphavirus/diagnóstico , Infecções por Alphavirus/virologia , Alphavirus/classificação , Alphavirus/genética , Alphavirus/isolamento & purificação , Infecções por Alphavirus/epidemiologia , Infecções por Alphavirus/transmissão , Animais , Teorema de Bayes , Pré-Escolar , Chlorocebus aethiops , Humanos , Masculino , Método de Monte Carlo , Papua Nova Guiné , Filogenia , Células Vero
7.
Viruses ; 11(10)2019 Oct 03.
Artigo em Inglês | MEDLINE | ID: mdl-31623340

RESUMO

Human parechovirus (HPeV), particularly type 3 (HPeV3), is an important cause of sepsis-/meningitis-like illness in young infants. Laboratory records identified a total of ten HPeV-positive cases in Southeastern Australia between January and July 2019. The HPeV present in these cases were typed by Sanger sequencing of the partial viral capsid protein 1 (VP1) region and selected cases were further characterised by additional Sanger or Ion Torrent near-full length virus sequencing. In seven of the ten cases, an HPeV type 5 (HPeV5) was identified, and in the remaining three cases, an HPeV type 1 was identified. The HPeV5-positive cases were infants under the age of 3 months admitted to hospital with fever, rash, lethargy and/or sepsis-like clinical signs. Near full-length virus sequencing revealed that the HPeV5 was most likely a recombinant virus, with structural genes most similar to an HPeV5 from Belarus in 2018, and a polymerase gene most similar to an HPeV3 from Australia in 2013/14. While HPeV5 is not typically associated with severe clinical signs, the HPeV5 identified here may have been able to cause more severe disease in young infants through the acquisition of genes from a more virulent HPeV.

8.
Sci Rep ; 9(1): 8906, 2019 06 20.
Artigo em Inglês | MEDLINE | ID: mdl-31222066

RESUMO

Human parechovirus type 3 (HPeV3) can cause severe sepsis-like illness in young infants and may be associated with long term neurodevelopmental delay later in childhood. We investigated the molecular epidemiology of HPeV infection in thirty three infants requiring hospitalization before, during and after the peak of the 2017/18 HPeV epidemic wave in Australia. During the peak of the epidemic, all cases were infected with an HPeV3, while before and after the peak, HPeV1 was the predominant type detected. The predominant HPeV3 was the recombinant HPeV3 also detected in the 2013/14 and 2015/16 Australian epidemics. Sepsis-like or meningitis-like symptoms were only reported in cases infected with the recombinant HPeV3. Phylogenetic analysis of the recombinant HPeV3 revealed that the virus continued to evolve, also between the Australian outbreaks, thus indicating continued circulation, despite not being detected and reported in Australia or elsewhere in between epidemic waves. The recombinant HPeV3 continued to show a remarkable stability in its capsid amino acid sequence, further strengthening our previous argument for development of a vaccine or immunotherapeutics to reduce the severity of HPeV3 outbreaks due to this virus.

9.
Sex Transm Infect ; 95(4): 307-313, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-30554143

RESUMO

OBJECTIVES: Reports of rising herpes simplex virus type 1 (HSV-1) genital infections relative to HSV-2 have been published up to 2006 in Australia. These changes have been attributed to declining childhood immunity to HSV-1. We described the temporal trends of HSV-1 and HSV-2 up to 2017 in Melbourne, Australia, to determine if the earlier trend is continuing. METHODS: We conducted a retrospective review of the medical records of 4517 patients who were diagnosed with first episode of anogenital HSV infection at the Melbourne Sexual Health Centre, Australia, between January 2004 and December 2017. HSV-1 and HSV-2 were calculated as a proportion of all first episode of anogenital HSV infections. The change in the proportions of HSV-1 and HSV-2 over time was assessed by a χ2 trend test. Risk factors associated with HSV-1 were examined using a multivariable logistic regression model. RESULTS: The proportion of first episode of anogenital herpes due to HSV-1 increased significantly over time in women (from 45% to 61%; ptrend<0.001) and heterosexual men (from 38% to 41%; ptrend=0.01) but not in men who have sex with men (MSM) (ptrend=0.21). After adjusting for condom use, partner number and age, the annual increase remained significant only in women (OR 1.08, 95% CI 1.03 to 1.13, p<0.001). In MSM, HSV-1 caused up to two-thirds of anogenital herpes in most years and HSV-1 was more likely to be diagnosed at an anal site than genital site (OR 1.69, 95% CI 1.23 to 2.32, p<0.001). Younger age (<28 years) was an independent risk factor for HSV-1 in all groups. CONCLUSIONS: The proportion of first-episode anogenital herpes due to HSV-1 has been rising in women since 2004. HSV-1 has become the leading cause of anogenital herpes in younger populations, women and MSM.


Assuntos
Herpes Genital/epidemiologia , Herpesvirus Humano 1 , Herpesvirus Humano 2 , Adulto , Instituições de Assistência Ambulatorial , Feminino , Herpes Genital/diagnóstico , Herpes Genital/etiologia , Humanos , Masculino , Registros Médicos , Prevalência , Estudos Retrospectivos , Fatores de Risco , Fatores Sexuais , Vitória/epidemiologia , Adulto Jovem
10.
Viruses ; 10(6)2018 06 11.
Artigo em Inglês | MEDLINE | ID: mdl-29891797

RESUMO

Murray Valley Encephalitis virus (MVEV) is a mosquito-borne Flavivirus. Clinical presentation is rare but severe, with a case fatality rate of 15⁻30%. Here we report a case of MVEV from the cerebrospinal fluid (CSF) of a patient in the Northern Territory in Australia. Initial diagnosis was performed using both MVEV-specific real-time, and Pan-Flavivirus conventional, Polymerase Chain Reaction (PCR), with confirmation by Sanger sequencing. Subsequent isolation, the first from CSF, was conducted in Vero cells and the observed cytopathic effect was confirmed by increasing viral titre in the real-time PCR. Isolation allowed for full genome sequencing using the Scriptseq V2 RNASeq library preparation kit. A consensus genome for VIDRL-MVE was generated and phylogenetic analysis identified it as Genotype 2. This is the first reported isolation, and full genome sequencing of MVEV from CSF. It is also the first time Genotype 2 has been identified in humans. As such, this case has significant implications for public health surveillance, epidemiology, and the understanding of MVEV evolution.


Assuntos
Líquido Cefalorraquidiano/virologia , Vírus da Encefalite do Vale de Murray/classificação , Vírus da Encefalite do Vale de Murray/isolamento & purificação , Encefalite por Arbovirus/virologia , Sequenciamento Completo do Genoma , Animais , Criança , Chlorocebus aethiops , Vírus da Encefalite do Vale de Murray/genética , Genótipo , Humanos , Northern Territory , Filogenia , Reação em Cadeia da Polimerase , Homologia de Sequência , Células Vero , Cultura de Vírus
11.
Open Forum Infect Dis ; 4(4): ofx203, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-29226169

RESUMO

We describe a fatal case of Japanese encephalitis virus infection following short-term travel to Thailand. Viral RNA was detected in urine and whole blood out to 26 and 28 days, respectively, after the onset of symptoms. Live virus was isolated from a urine specimen from day 14.

12.
Biochem Biophys Res Commun ; 483(1): 64-68, 2017 01 29.
Artigo em Inglês | MEDLINE | ID: mdl-28062184

RESUMO

Respiratory syncytial virus (RSV) is a major cause of respiratory infections in infants and the elderly, leading to more deaths than influenza each year worldwide. With no RSV antiviral or efficacious vaccine currently available, improved understanding of the host-RSV interaction is urgently required. Here we examine the contribution to RSV infection of the host stress-regulated c-Jun N-terminal kinase (JNK), for the first time. Peak JNK1/2 phosphoactivation is observed at ∼24 h post-infection, correlating with the time of virus assembly. The release of infectious RSV virions from infected cells was significantly reduced by either JNK1/2 siRNA knockdown or treatment with the JNK-specific inhibitor, JNK-IN-VIII. High resolution microscopy confirmed RSV accumulation in the host cell cytoplasm. The results implicate JNK1/2 as a key host factor for RSV virus production, raising the possibility of agents targeting JNK activity as potential anti-RSV therapeutics.


Assuntos
Proteína Quinase 8 Ativada por Mitógeno/metabolismo , Proteína Quinase 9 Ativada por Mitógeno/metabolismo , Vírus Sincicial Respiratório Humano/fisiologia , Replicação Viral/fisiologia , Células A549 , Ativação Enzimática , Técnicas de Silenciamento de Genes , Interações Hospedeiro-Patógeno/efeitos dos fármacos , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/fisiologia , Humanos , Proteína Quinase 8 Ativada por Mitógeno/antagonistas & inibidores , Proteína Quinase 8 Ativada por Mitógeno/genética , Proteína Quinase 9 Ativada por Mitógeno/antagonistas & inibidores , Proteína Quinase 9 Ativada por Mitógeno/genética , Inibidores de Proteínas Quinases/farmacologia , RNA Interferente Pequeno/genética , Vírus Sincicial Respiratório Humano/efeitos dos fármacos , Vírus Sincicial Respiratório Humano/patogenicidade , Vírion/fisiologia , Montagem de Vírus/fisiologia , Liberação de Vírus/fisiologia
13.
Methods Mol Biol ; 1442: 93-117, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-27464690

RESUMO

Although respiratory syncytial virus (RSV) is the leading cause of bronchiolitis and pneumonia in infants and the elderly worldwide [1], the protein-protein interactions between the host cell and virus remain poorly understood. We have used a focused small interfering RNA (siRNA) approach to knock-down and examine the role(s) of various host cell proteins. Here, we describe approaches for casein kinase 2α (CK2α) as a key example. We show how to study the effect of host gene (CK2α) knockdown using siRNA on cell-associated and released virus titers, using both quantitative RT-PCR, which measures the level of viral RNA, and plaque assay, which measures infectious virus directly. Both assays identified reduced viral titers with CK2α gene knock-down, indicating that it is likely required for efficient viral assembly and/or release. Effects were confirmed in RSV infected cells using the specific CK2α inhibitor 4,5,6,7-tetrabromobenzotriazole, revealing a similar reduction in viral titers as CK2α specific siRNA. This demonstrates that siRNA can be used to characterize critical host cell-RSV protein-protein interactions, and establishes CK2α as a future druggable target.


Assuntos
RNA Interferente Pequeno/genética , Infecções por Vírus Respiratório Sincicial/genética , Vírus Sincicial Respiratório Humano/fisiologia , Células A549 , Caseína Quinase II/genética , Caseína Quinase II/metabolismo , Técnicas de Silenciamento de Genes , Interações Hospedeiro-Patógeno , Humanos , Reação em Cadeia da Polimerase em Tempo Real , Infecções por Vírus Respiratório Sincicial/virologia , Carga Viral , Ensaio de Placa Viral , Montagem de Vírus
14.
Biochem Biophys Res Commun ; 470(3): 735-740, 2016 Feb 12.
Artigo em Inglês | MEDLINE | ID: mdl-26792716

RESUMO

Nuclear import of the accessory protein Vpr is central to infection by human immunodeficiency virus (HIV). We previously identified the Vpr F72L mutation in a HIV-infected, long-term non-progressor, showing that it resulted in reduced Vpr nuclear accumulation and altered cytoplasmic localisation. Here we demonstrate for the first time that the effects of nuclear accumulation of the F72L mutation are due to impairment of microtubule-dependent-enhancement of Vpr nuclear import. We use high resolution imaging approaches including fluorescence recovery after photobleaching and other approaches to document interaction between Vpr and the dynein light chain protein, DYNLT1, and impaired interaction of the F72L mutant with DYNLT1. The results implicate MTs/DYNLT1 as drivers of Vpr nuclear import and HIV infection, with important therapeutic implications.


Assuntos
Núcleo Celular/metabolismo , Dineínas/metabolismo , Produtos do Gene vpr do Vírus da Imunodeficiência Humana/metabolismo , Transporte Ativo do Núcleo Celular/fisiologia , Animais , Sítios de Ligação , Células COS , Chlorocebus aethiops , Ligação Proteica , Transporte Proteico/fisiologia , Relação Estrutura-Atividade
15.
Front Microbiol ; 6: 848, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26322040

RESUMO

The respiratory diseases caused by rhinovirus, respiratory syncytial virus, and influenza virus represent a large social and financial burden on healthcare worldwide. Although all three viruses have distinctly unique properties in terms of infection and replication, they share the ability to exploit/manipulate the host-cell nucleocytoplasmic transport system in order to replicate effectively and efficiently. This review outlines the various ways in which infection by these viruses impacts on the host nucleocytoplasmic transport system, and examples where inhibition thereof in turn decreases viral replication. The highly conserved nature of the nucleocytoplasmic transport system and the viral proteins that interact with it make this virus-host interface a prime candidate for the development of specific antiviral therapeutics in the future.

16.
Mol Cell Proteomics ; 14(3): 532-43, 2015 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-25556234

RESUMO

Although human respiratory syncytial virus (RSV) is the most common cause of bronchiolitis and pneumonia in infants and elderly worldwide, there is no licensed RSV vaccine or effective drug treatment available. The RSV Matrix protein plays key roles in virus life cycle, being found in the nucleus early in infection in a transcriptional inhibitory role, and later localizing in viral inclusion bodies before coordinating viral assembly and budding at the plasma membrane. In this study, we used a novel, high throughput microfluidics platform and custom human open reading frame library to identify novel host cell binding partners of RSV matrix. Novel interactors identified included proteins involved in host transcription regulation, the innate immunity response, cytoskeletal regulation, membrane remodeling, and cellular trafficking. A number of these interactions were confirmed by immunoprecipitation and cellular colocalization approaches. Importantly, the physiological significance of matrix interaction with the actin-binding protein cofilin 1, caveolae protein Caveolin 2, and the zinc finger protein ZNF502 was confirmed. siRNA knockdown of the host protein levels resulted in reduced RSV virus production in infected cells. These results have important implications for future antiviral strategies aimed at targets of RSV matrix in the host cell.


Assuntos
Proteínas de Transporte/metabolismo , Caveolina 2/metabolismo , Cofilina 1/metabolismo , Técnicas Analíticas Microfluídicas/métodos , Proteínas Nucleares/metabolismo , Vírus Sinciciais Respiratórios/fisiologia , Proteínas da Matriz Viral/metabolismo , Animais , Linhagem Celular , Núcleo Celular/metabolismo , Chlorocebus aethiops , Biblioteca Gênica , Células HEK293 , Humanos , Fases de Leitura Aberta , Células Vero , Replicação Viral
17.
Antiviral Res ; 95(3): 202-6, 2012 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-22750233

RESUMO

A key aspect of the infectious cycle of many viruses is the transport of specific viral proteins into the host cell nucleus to perturb the antiviral response. Examples include a number of RNA viruses that are significant human pathogens, such as human immunodeficiency virus (HIV)-1, influenza A, dengue, respiratory syncytial virus and rabies, as well agents that predominantly infect livestock, such as Rift valley fever virus and Venezuelan equine encephalitis virus. Inhibiting the nuclear trafficking of viral proteins as a therapeutic strategy offers an attractive possibility, with important recent progress having been made with respect to HIV-1 and dengue. The results validate nuclear protein import as an antiviral target, and suggest the identification and development of nuclear transport inhibitors as a viable therapeutic approach for a range of human and zoonotic pathogenic viruses.


Assuntos
Transporte Ativo do Núcleo Celular , Antivirais/farmacologia , Vírus de RNA/fisiologia , Proteínas Virais/metabolismo , Animais , Humanos , Infecções por Vírus de RNA/tratamento farmacológico , Infecções por Vírus de RNA/virologia , Vírus de RNA/patogenicidade
18.
J Fluoresc ; 19(3): 567-73, 2009 May.
Artigo em Inglês | MEDLINE | ID: mdl-19067127

RESUMO

Fluorescent labelling of the highly conserved HIV-1 accessory protein Vpr (Viral Protein R) with GFP or variants thereof has proved a valuable approach to track Vpr and/or HIV-1 subcellular localisation in vivo. Our analysis in transfected mammalian cells expressing GFP-Vpr fusion protein, as well as within virus derived there from, documents site-specific proteolytic cleavage of the GFP-Vpr fusion protein. Western analysis revealed that transfected mammalian cells harbour a C-terminally truncated variant of Vpr in addition to full-length GFP-Vpr. Further, virions derived from these GFP-Vpr expressing cells show protein in which the GFP-tag has been additionally cleaved from the Vpr protein. Endogenous HIV protease (PR) activity was shown to be responsible for the latter, as addition of Saquinavir, a potent PR inhibitor abolished the cleavage. Since many previous studies have relied on imaging the GFP fluorescence of GFP-Vpr, it would appear that the results may not reflect intact GFP-Vpr.


Assuntos
Proteínas de Fluorescência Verde/metabolismo , HIV-1/metabolismo , Espaço Intracelular/metabolismo , Proteínas Recombinantes de Fusão/metabolismo , Vírion/metabolismo , Produtos do Gene vpr do Vírus da Imunodeficiência Humana/metabolismo , Animais , Sítios de Ligação , Linhagem Celular , Núcleo Celular/metabolismo , Sobrevivência Celular , Humanos , Transporte Proteico , Especificidade por Substrato
19.
Retrovirology ; 5: 67, 2008 Jul 18.
Artigo em Inglês | MEDLINE | ID: mdl-18638397

RESUMO

We previously reported an epidemiologically linked HIV-1 infected patient cohort in which a long-term non-progressor (LTNP) infected two recipients who then exhibited normal disease progression. Expression of patient-derived vpr sequences from each of the three cohort members in mammalian cells tagged with GFP revealed a significant reduction in Vpr nuclear import and virion incorporation uniquely from the LTNP, whereas Vpr from the two progressing recipients displayed normal localisation and virion incorporation, implying a link between efficient Vpr nuclear import and HIV disease progression. Importantly, an F72L point mutation in the LTNP was identified for the first time as being uniquely responsible for decreased Vpr nuclear import.


Assuntos
Núcleo Celular/metabolismo , Produtos do Gene vpr/metabolismo , Infecções por HIV/metabolismo , Infecções por HIV/virologia , Sobreviventes de Longo Prazo ao HIV , HIV-1/metabolismo , Animais , Células COS , Núcleo Celular/genética , Chlorocebus aethiops , Progressão da Doença , Expressão Gênica , Produtos do Gene vpr/genética , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , HIV-1/genética , Humanos , Mutação Puntual , Transporte Proteico , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA