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1.
Cell Host Microbe ; 25(4): 617-629.e7, 2019 Apr 10.
Artigo em Inglês | MEDLINE | ID: mdl-30926239

RESUMO

The complement system is vital for anti-microbial defense. In the classical pathway, pathogen-bound antibody recruits the C1 complex (C1qC1r2C1s2) that initiates a cleavage cascade involving C2, C3, C4, and C5 and triggering microbial clearance. We demonstrate a C4-dependent antiviral mechanism that is independent of downstream complement components. C4 inhibits human adenovirus infection by directly inactivating the virus capsid. Rapid C4 activation and capsid deposition of cleaved C4b are catalyzed by antibodies via the classical pathway. Capsid-deposited C4b neutralizes infection independent of C2 and C3 but requires C1q antibody engagement. C4b inhibits capsid disassembly, preventing endosomal escape and cytosolic access. C4-deficient mice exhibit heightened viral burdens. Additionally, complement synergizes with the Fc receptor TRIM21 to block transduction by an adenovirus gene therapy vector but is partially restored by Fab virus shielding. These results suggest that the complement system could be altered to prevent virus infection and enhance virus gene therapy efficacy.

2.
Innate Immun ; 24(2): 112-121, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-29433372

RESUMO

Viral infection is a major cause of morbidity and mortality following allogeneic hematopoietic stem cell transplant (HSCT), with up to one in four deaths directly linked to viral disease. Whilst awaiting lymphocyte reconstitution post-HSCT, the innate antiviral immune response is the first line of defense against invading viruses. Several novel innate viral-sensing pathways have recently been characterized, but their physiological importance in humans is poorly understood. We analyzed a panel of innate viral-sensor genes in HSCT patients, and assessed whether differences in innate antiviral responses could account for variation in susceptibility to viral infections. Expression levels of innate viral sensors in HSCT patients with active viral infections, HSCT patients without active infections and healthy volunteers were highly homogenous. Although IFN-α expression was up-regulated in actively infected patients relative to controls, a corresponding up-regulation of innate viral sensor expression was not observed. IFN-α stimulation of patient PBMCs in vitro showed intact IFN-α signaling, but actively infected patients' PBMCs had reduced up-regulation of innate viral sensors. We show that the aberrant IFN-α responses in HSCT patients were not due to calcineurin inhibition. Our data therefore raises the possibility of an intrinsic defect in innate viral sensor up-regulation in HSCT patients following viral infection.

3.
Nature ; 544(7650): 309-315, 2017 04 20.
Artigo em Inglês | MEDLINE | ID: mdl-28405027

RESUMO

The 2013-2016 West African epidemic caused by the Ebola virus was of unprecedented magnitude, duration and impact. Here we reconstruct the dispersal, proliferation and decline of Ebola virus throughout the region by analysing 1,610 Ebola virus genomes, which represent over 5% of the known cases. We test the association of geography, climate and demography with viral movement among administrative regions, inferring a classic 'gravity' model, with intense dispersal between larger and closer populations. Despite attenuation of international dispersal after border closures, cross-border transmission had already sown the seeds for an international epidemic, rendering these measures ineffective at curbing the epidemic. We address why the epidemic did not spread into neighbouring countries, showing that these countries were susceptible to substantial outbreaks but at lower risk of introductions. Finally, we reveal that this large epidemic was a heterogeneous and spatially dissociated collection of transmission clusters of varying size, duration and connectivity. These insights will help to inform interventions in future epidemics.


Assuntos
Ebolavirus/genética , Ebolavirus/fisiologia , Genoma Viral/genética , Doença pelo Vírus Ebola/transmissão , Doença pelo Vírus Ebola/virologia , Clima , Surtos de Doenças/estatística & dados numéricos , Ebolavirus/isolamento & purificação , Geografia , Doença pelo Vírus Ebola/epidemiologia , Humanos , Internacionalidade , Modelos Lineares , Epidemiologia Molecular , Filogenia , Viagem/legislação & jurisprudência , Viagem/estatística & dados numéricos
4.
Mol Cell Proteomics ; 16(4 suppl 1): S215-S229, 2017 04.
Artigo em Inglês | MEDLINE | ID: mdl-28087593

RESUMO

Noroviruses produce viral RNAs lacking a 5' cap structure and instead use a virus-encoded viral protein genome-linked (VPg) protein covalently linked to viral RNA to interact with translation initiation factors and drive viral protein synthesis. Norovirus infection results in the induction of the innate response leading to interferon stimulated gene (ISG) transcription. However, the translation of the induced ISG mRNAs is suppressed. A SILAC-based mass spectrometry approach was employed to analyze changes to protein abundance in both whole cell and m7GTP-enriched samples to demonstrate that diminished host mRNA translation correlates with changes to the composition of the eukaryotic initiation factor complex. The suppression of host ISG translation correlates with the activity of the viral protease (NS6) and the activation of cellular caspases leading to the establishment of an apoptotic environment. These results indicate that noroviruses exploit the differences between viral VPg-dependent and cellular cap-dependent translation in order to diminish the host response to infection.


Assuntos
Infecções por Caliciviridae/genética , Norovirus/metabolismo , Proteômica/métodos , Capuzes de RNA/metabolismo , RNA Mensageiro/metabolismo , Proteínas Virais/metabolismo , Apoptose , Infecções por Caliciviridae/imunologia , Infecções por Caliciviridae/virologia , Interações Hospedeiro-Patógeno , Humanos , Imunidade Inata , Marcação por Isótopo/métodos , Espectrometria de Massas/métodos , Norovirus/genética , RNA Viral/metabolismo
5.
PLoS Med ; 13(10): e1002139, 2016 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-27727279

RESUMO

BACKGROUND: Interleukin-2 (IL-2) has an essential role in the expansion and function of CD4+ regulatory T cells (Tregs). Tregs reduce tissue damage by limiting the immune response following infection and regulate autoreactive CD4+ effector T cells (Teffs) to prevent autoimmune diseases, such as type 1 diabetes (T1D). Genetic susceptibility to T1D causes alterations in the IL-2 pathway, a finding that supports Tregs as a cellular therapeutic target. Aldesleukin (Proleukin; recombinant human IL-2), which is administered at high doses to activate the immune system in cancer immunotherapy, is now being repositioned to treat inflammatory and autoimmune disorders at lower doses by targeting Tregs. METHODS AND FINDINGS: To define the aldesleukin dose response for Tregs and to find doses that increase Tregs physiologically for treatment of T1D, a statistical and systematic approach was taken by analysing the pharmacokinetics and pharmacodynamics of single doses of subcutaneous aldesleukin in the Adaptive Study of IL-2 Dose on Regulatory T Cells in Type 1 Diabetes (DILT1D), a single centre, non-randomised, open label, adaptive dose-finding trial with 40 adult participants with recently diagnosed T1D. The primary endpoint was the maximum percentage increase in Tregs (defined as CD3+CD4+CD25highCD127low) from the baseline frequency in each participant measured over the 7 d following treatment. There was an initial learning phase with five pairs of participants, each pair receiving one of five pre-assigned single doses from 0.04 × 106 to 1.5 × 106 IU/m2, in order to model the dose-response curve. Results from each participant were then incorporated into interim statistical modelling to target the two doses most likely to induce 10% and 20% increases in Treg frequencies. Primary analysis of the evaluable population (n = 39) found that the optimal doses of aldesleukin to induce 10% and 20% increases in Tregs were 0.101 × 106 IU/m2 (standard error [SE] = 0.078, 95% CI = -0.052, 0.254) and 0.497 × 106 IU/m2 (SE = 0.092, 95% CI = 0.316, 0.678), respectively. On analysis of secondary outcomes, using a highly sensitive IL-2 assay, the observed plasma concentrations of the drug at 90 min exceeded the hypothetical Treg-specific therapeutic window determined in vitro (0.015-0.24 IU/ml), even at the lowest doses (0.040 × 106 and 0.045 × 106 IU/m2) administered. A rapid decrease in Treg frequency in the circulation was observed at 90 min and at day 1, which was dose dependent (mean decrease 11.6%, SE = 2.3%, range 10.0%-48.2%, n = 37), rebounding at day 2 and increasing to frequencies above baseline over 7 d. Teffs, natural killer cells, and eosinophils also responded, with their frequencies rapidly and dose-dependently decreased in the blood, then returning to, or exceeding, pretreatment levels. Furthermore, there was a dose-dependent down modulation of one of the two signalling subunits of the IL-2 receptor, the ß chain (CD122) (mean decrease = 58.0%, SE = 2.8%, range 9.8%-85.5%, n = 33), on Tregs and a reduction in their sensitivity to aldesleukin at 90 min and day 1 and 2 post-treatment. Due to blood volume requirements as well as ethical and practical considerations, the study was limited to adults and to analysis of peripheral blood only. CONCLUSIONS: The DILT1D trial results, most notably the early altered trafficking and desensitisation of Tregs induced by a single ultra-low dose of aldesleukin that resolves within 2-3 d, inform the design of the next trial to determine a repeat dosing regimen aimed at establishing a steady-state Treg frequency increase of 20%-50%, with the eventual goal of preventing T1D. TRIAL REGISTRATION: ISRCTN Registry ISRCTN27852285; ClinicalTrials.gov NCT01827735.


Assuntos
Diabetes Mellitus Tipo 1/prevenção & controle , Interleucina-2/análogos & derivados , Linfócitos T Reguladores/efeitos dos fármacos , Adolescente , Adulto , Biomarcadores , Quimiocinas/biossíntese , Relação Dose-Resposta a Droga , Eosinófilos/efeitos dos fármacos , Feminino , Humanos , Imunofenotipagem , Mediadores da Inflamação/metabolismo , Interleucina-2/efeitos adversos , Interleucina-2/farmacologia , Células Matadoras Naturais/efeitos dos fármacos , Células Matadoras Naturais/imunologia , Contagem de Linfócitos , Masculino , Pessoa de Meia-Idade , Proteínas Recombinantes/efeitos adversos , Proteínas Recombinantes/farmacologia , Adulto Jovem
6.
Genome Announc ; 4(5)2016 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-27634993

RESUMO

Hepatitis E virus (HEV) genotype 3 is endemic in the United Kingdom but the complete sequence of HEV, generated directly from a clinical sample, is lacking. We report a near full-length genome sequence of genotype 3 HEV from the serum of a patient with acute hepatitis.

7.
Euro Surveill ; 20(40)2015.
Artigo em Inglês | MEDLINE | ID: mdl-26539753

RESUMO

The Magazine Wharf area, Freetown, Sierra Leone was a focus of ongoing Ebola virus transmission from late June 2015. Viral genomes linked to this area contain a series of 13 T to C substitutions in a 150 base pair intergenic region downstream of viral protein 40 open reading frame, similar to the Ebolavirus/H.sapiens-wt/SLE/2014/Makona-J0169 strain (J0169) detected in the same town in November 2014. This suggests that recently circulating viruses from Freetown descend from a J0169-like virus.


Assuntos
Surtos de Doenças , Ebolavirus/genética , Doença pelo Vírus Ebola/epidemiologia , Ebolavirus/isolamento & purificação , Genoma Viral , Genótipo , Doença pelo Vírus Ebola/diagnóstico , Humanos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Serra Leoa
8.
PLoS One ; 10(6): e0128703, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26076364

RESUMO

Hepatitis E virus (HEV) genotypes 3 and 4 are zoonotic pathogens, with pigs predominantly implicated in disease transmission. The rapid rise in human cases in developed countries over the past decade indicates a change in epidemiology of HEV, and it has been suggested that additional animal species may be involved in transmission of infection. Multiple studies have identified contact with dogs as a risk factor for HEV infection in industrialised nations, and a low seroprevalence to HEV has previously been reported in dogs in low-income countries. In this study we aimed to evaluate the possibility that dogs are susceptible to HEV, and determine the frequency with which this occurs. Serum samples from UK dogs with and without hepatitis were screened for HEV-specific antibodies, and canine liver and stool samples were analysed by qPCR for the presence of HEV RNA. We describe evidence to show HEV infection occurs at low levels in dogs in the UK, but the strain of origin is undetermined. The low seroprevalence level of HEV in dogs implies the risk of zoonotic disease transmission is likely to be limited, but further investigations will be required to determine if HEV-infected dogs can transmit HEV to man.


Assuntos
Doenças do Cão/imunologia , Doenças do Cão/virologia , Anticorpos Anti-Hepatite/imunologia , Vírus da Hepatite E/imunologia , Hepatite E/veterinária , Animais , Cães , Genótipo , Anticorpos Anti-Hepatite/sangue , Vírus da Hepatite E/genética , Humanos , Norovirus/imunologia , RNA Viral , Estudos Soroepidemiológicos , Reino Unido
9.
J Clin Microbiol ; 53(6): 1873-83, 2015 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-25832298

RESUMO

Human noroviruses (HuNoVs) are a major cause of viral gastroenteritis, with an estimated 3 million cases per year in the United Kingdom. HuNoVs have recently been isolated from pet dogs in Europe (M. Summa, C.-H. von Bonsdorff, and L. Maunula, J Clin Virol 53:244-247, 2012, http://dx.doi.org/10.1016/j.jcv.2011.12.014), raising concerns about potential zoonotic infections. With 31% of United Kingdom households owning a dog, this could prove to be an important transmission route. To examine this risk, canine tissues were studied for their ability to bind to HuNoV in vitro. In addition, canine stool samples were analyzed for the presence of viral nucleic acid, and canine serum samples were tested for the presence of anti-HuNoV antibodies. The results showed that seven different genotypes of HuNoV virus-like particles (VLPs) can bind to canine gastrointestinal tissue, suggesting that infection is at least theoretically possible. Although HuNoV RNA was not identified in stool samples from 248 dogs, serological evidence of previous exposure to HuNoV was obtained in 43/325 canine serum samples. Remarkably, canine seroprevalence for different HuNoV genotypes mirrored the seroprevalence in the human population. Though entry and replication within cells have not been demonstrated, the canine serological data indicate that dogs produce an immune response to HuNoV, implying productive infection. In conclusion, this study reveals zoonotic implications for HuNoV, and to elucidate the significance of this finding, further epidemiological and molecular investigations will be essential.


Assuntos
Infecções por Caliciviridae , Cães/virologia , Norovirus , Zoonoses , Animais , Anticorpos Antivirais/sangue , Infecções por Caliciviridae/epidemiologia , Infecções por Caliciviridae/veterinária , Infecções por Caliciviridae/virologia , Fezes/virologia , Feminino , Trato Gastrointestinal/metabolismo , Humanos , Norovirus/genética , Norovirus/isolamento & purificação , Norovirus/metabolismo , Saliva/metabolismo , Saliva/virologia , Estudos Soroepidemiológicos , Reino Unido/epidemiologia , Vírion/metabolismo , Zoonoses/epidemiologia , Zoonoses/virologia
10.
Vet Microbiol ; 177(1-2): 206-13, 2015 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-25818578

RESUMO

Astroviruses are a common cause of gastroenteritis in children worldwide. These viruses can also cause infection in a range of domestic and wild animal species. Canine astrovirus (CaAstV) was first identified in the USA, and has since been reported in dogs from Europe, the Far East and South America. We sought to determine whether CaAstV is circulating in the UK dog population, and to characterise any identified strains. Stool samples were collected from pet dogs in the UK with and without gastroenteritis, and samples were screened for CaAstV by qPCR. Four CaAstV positive samples were identified from dogs with gastroenteritis (4/67, 6.0%), whereas no samples from healthy dogs were positive (p<0.001). Sequencing of the capsid sequences from the four CaAstV strains found significant genetic heterogeneity, with only 80% amino acid identity between strains. The full genome sequence of two UK CaAstV strains was then determined, confirming that CaAstV conforms to the classic genome organisation of other astroviruses with ORF1a and ORF1b separated by a frameshift and ORF2 encoding the capsid protein. This is the first report describing the circulation of CaAstV in UK dogs with clinical signs of gastroenteritis, and the first description of the full-length genomes of two CaAstV strains.


Assuntos
Infecções por Astroviridae/veterinária , Astroviridae/genética , Doenças do Cão/epidemiologia , Doenças do Cão/virologia , Gastroenterite/veterinária , Genoma Viral/genética , Animais de Estimação/virologia , Sequência de Aminoácidos , Animais , Infecções por Astroviridae/epidemiologia , Sequência de Bases , Proteínas do Capsídeo/genética , Cães , Fezes/virologia , Gastroenterite/virologia , Dados de Sequência Molecular , Reação em Cadeia da Polimerase/veterinária , Análise de Sequência de DNA/veterinária , Homologia de Sequência , Especificidade da Espécie , Reino Unido
11.
Curr Protoc Microbiol ; 33: 15K.2.1-61, 2014 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-24789596

RESUMO

Murine norovirus (MNV) is a positive-sense, plus-stranded RNA virus in the Caliciviridae family. It is the most common pathogen in biomedical research colonies. MNV is also related to the human noroviruses, which cause the majority of nonbacterial gastroenteritis worldwide. Like the human noroviruses, MNV is an enteric virus that replicates in the intestine and is transmitted by the fecal-oral route. MNV replicates in murine macrophages and dendritic cells in cells in culture and in the murine host. This virus is often used to study mechanisms in norovirus biology, because human noroviruses are refractory to growth in cell culture. MNV combines the availability of a cell culture and reverse genetics system with the ability to study infection in the native host. Herein, we describe a panel of techniques that are commonly used to study MNV biology.


Assuntos
Norovirus/crescimento & desenvolvimento , Cultura de Vírus , Animais , Linhagem Celular , Centrifugação com Gradiente de Concentração , Césio , Cloretos , Ensaio de Imunoadsorção Enzimática , Genoma Viral , Humanos , Camundongos , Mutagênese , Norovirus/genética , Reação em Cadeia da Polimerase , Recombinação Genética , Genética Reversa , Replicação Viral
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