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1.
BMC Infect Dis ; 20(1): 670, 2020 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-32933492

RESUMO

BACKGROUND: The 2014/15 Ebola outbreak in West Africa resulted in 11,000 deaths and massive strain on local health systems, and the ongoing outbreak in Democratic Republic of Congo has afflicted more than 3000 people. Accurate, rapid Ebola diagnostics suitable for field deployment would enable prompt identification and effective response to future outbreaks, yet remain largely unavailable. The purpose of this study was to assess the accuracy of three novel rapid diagnostic tests (RDTs): an Ebola, an Ebola-Malaria, and a Fever Panel test that includes Ebola, all from a single manufacturer. METHODS: We evaluated the three RDTs in 109 Ebola-positive and 96 Ebola-negative stored serum samples collected during the outbreak in Guinea in 2014/15, and tested by real-time polymerase chain reaction (RT-PCR). Sensitivity, specificity, and overall percent agreement were calculated for each RDT using RT-PCR as a reference standard, stratified by Ct value ranges. RESULTS: All tests performed with high accuracy on samples with low Ct value (high viral load). The Fever Panel test performed with the highest accuracy, with a sensitivity of 89.9% and specificity of 90.6%. The Ebola and Ebola-Malaria tests performed comparably to each other: sensitivity was 77.1 and 78% respectively, and specificity was 91.7% for the Ebola test and 95.8% for the Ebola-Malaria test. CONCLUSIONS: This study evaluated the accuracy of three novel rapid diagnostic tests for Ebola. The tests may have significant public health relevance, particularly the Fever Panel test, which detects seven pathogens including Ebola. Given limitations to the study resulting from uncertain sample quality, further evaluation is warranted. All tests performed with highest accuracy on samples with low Ct value (high viral load), and the data presented here suggests that these RDTs may be useful for point-of-care diagnosis of cases in the context of an outbreak. Restrictions to their use in non-severe Ebola cases or for longitudinal monitoring, when viral loads are lower, may be appropriate. Highlighting the challenge in developing and evaluating Ebola RDTs, there were concerns regarding sample integrity and reference testing, and there is a need for additional research to validate these assays.


Assuntos
Doença pelo Vírus Ebola/diagnóstico , Sistemas Automatizados de Assistência Junto ao Leito , Surtos de Doenças , Ebolavirus/genética , Ebolavirus/isolamento & purificação , Guiné/epidemiologia , Doença pelo Vírus Ebola/epidemiologia , Doença pelo Vírus Ebola/virologia , Humanos , RNA Viral/análise , RNA Viral/metabolismo , Kit de Reagentes para Diagnóstico , Reação em Cadeia da Polimerase em Tempo Real/métodos , Sensibilidade e Especificidade
2.
Nat Commun ; 11(1): 4131, 2020 08 17.
Artigo em Inglês | MEDLINE | ID: mdl-32807807

RESUMO

Recent outbreaks of viral hemorrhagic fevers (VHFs), including Ebola virus disease (EVD) and Lassa fever (LF), highlight the urgent need for sensitive, deployable tests to diagnose these devastating human diseases. Here we develop CRISPR-Cas13a-based (SHERLOCK) diagnostics targeting Ebola virus (EBOV) and Lassa virus (LASV), with both fluorescent and lateral flow readouts. We demonstrate on laboratory and clinical samples the sensitivity of these assays and the capacity of the SHERLOCK platform to handle virus-specific diagnostic challenges. We perform safety testing to demonstrate the efficacy of our HUDSON protocol in heat-inactivating VHF viruses before SHERLOCK testing, eliminating the need for an extraction. We develop a user-friendly protocol and mobile application (HandLens) to report results, facilitating SHERLOCK's use in endemic regions. Finally, we successfully deploy our tests in Sierra Leone and Nigeria in response to recent outbreaks.


Assuntos
Ebolavirus/patogenicidade , Doença pelo Vírus Ebola/diagnóstico , Febre Lassa/diagnóstico , Vírus Lassa/patogenicidade , Anticorpos Antivirais , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas/genética , Ebolavirus/genética , Doença pelo Vírus Ebola/virologia , Febre Lassa/virologia , Vírus Lassa/genética
3.
PLoS Negl Trop Dis ; 14(7): e0008496, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32735587

RESUMO

BACKGROUND: The unprecedented 2013/16 outbreak of Zaire ebolavirus (Ebola virus) in West Africa has highighted the need for rapid, high-throughput and POC diagnostic assays to enable timely detection and appropriate triaging of Ebola Virus Disease (EVD) patients. Ebola virus is highly infectious and prompt diagnosis and triage is crucial in preventing further spread within community and healthcare settings. Moreover, due to the ecology of Ebola virus it is important that newly developed diagnostic assays are suitable for use in both the healthcare environment and low resource rural locations. METHODOLOGY/PRINCIPLE FINDINGS: A LAMP assay was successfully developed with three detection formats; a real-time intercalating dye-based assay, a real-time probe-based assay to enable multiplexing and an end-point colourimetric assay to simplify interpretation for the field. All assay formats were sensitive and specific, detecting a range of Ebola virus strains isolated in 1976-2014; with Probit analysis predicting limits of detection of 243, 290 and 75 copies/reaction respectively and no cross-detection of related strains or other viral haemorrhagic fevers (VHF's). The assays are rapid, (as fast as 5-7.25 mins for real-time formats) and robust, detecting Ebola virus RNA in presence of minimally diluted bodily fluids. Moreover, when tested on patient samples from the 2013/16 outbreak, there were no false positives and 93-96% of all new case positives were detected, with only a failure to detect very low copy number samples. CONCLUSION/SIGNIFICANCE: These are a set of robust and adaptable diagnostic solutions, which are fast, easy-to-perform-and-interpret and are suitable for use on a range of platforms including portable low-power devices. They can be readily transferred to field-laboratory settings, with no specific equipment needs and are therefore ideally placed for use in locations with limited resources.


Assuntos
Ebolavirus/isolamento & purificação , Doença pelo Vírus Ebola/diagnóstico , Doença pelo Vírus Ebola/virologia , Técnicas de Amplificação de Ácido Nucleico/métodos , Surtos de Doenças , Doença pelo Vírus Ebola/sangue , Doença pelo Vírus Ebola/epidemiologia , Humanos , RNA Viral , Sensibilidade e Especificidade , Serra Leoa/epidemiologia
4.
Nat Commun ; 11(1): 3730, 2020 07 24.
Artigo em Inglês | MEDLINE | ID: mdl-32709840

RESUMO

Long-term follow up studies from Ebola virus disease (EVD) survivors (EBOV_S) are lacking. Here, we evaluate immune and gene expression profiles in 35 Guinean EBOV_S from the last West African outbreak, a median of 23 months (IQR [18-25]) after discharge from treatment center. Compared with healthy donors, EBOV_S exhibit increases of blood markers of inflammation, intestinal tissue damage, T cell and B cell activation and a depletion of circulating dendritic cells. All survivors have EBOV-specific IgG antibodies and robust and polyfunctional EBOV-specific memory T-cell responses. Deep sequencing of the genes expressed in blood reveals an enrichment in 'inflammation' and 'antiviral' pathways. Integrated analyses identify specific immune markers associated with the persistence of clinical symptoms. This study identifies a set of biological and genetic markers that could be used to define a signature of "chronic Ebola virus disease (CEVD)".


Assuntos
Ebolavirus/imunologia , Doença pelo Vírus Ebola/complicações , Doença pelo Vírus Ebola/imunologia , Doenças do Sistema Imunitário/complicações , Doenças do Sistema Imunitário/imunologia , Adulto , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Antivirais/farmacologia , Linfócitos B/imunologia , Citocinas/sangue , Ebolavirus/efeitos dos fármacos , Ebolavirus/genética , Feminino , Marcadores Genéticos , Doença pelo Vírus Ebola/tratamento farmacológico , Doença pelo Vírus Ebola/virologia , Humanos , Doenças do Sistema Imunitário/genética , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Inflamação/genética , Ativação Linfocitária , Masculino , Sobreviventes , Linfócitos T/imunologia , Transcriptoma , Adulto Jovem
5.
Nat Commun ; 11(1): 3736, 2020 07 27.
Artigo em Inglês | MEDLINE | ID: mdl-32719371

RESUMO

A replication-competent vesicular stomatitis virus vaccine expressing the Ebola virus (EBOV) glycoprotein (GP) (rVSV-ZEBOV) was successfully used during the 2013-16 EBOV epidemic. Additionally, chimeric and human monoclonal antibodies (mAb) against the EBOV GP have shown promise in animals and humans when administered therapeutically. Uncertainty exists regarding the efficacy of postexposure antibody treatments in the event of a known exposure of a recent rVSV-ZEBOV vaccinee. Here, we model a worst-case scenario using rhesus monkeys vaccinated or unvaccinated with the rVSV-ZEBOV vaccine. We demonstrate that animals challenged with a uniformly lethal dose of EBOV one day following vaccination, and then treated with the anti-EBOV GP mAb MIL77 starting 3 days postexposure show no evidence of clinical illness and survive challenge. In contrast, animals receiving only vaccination or only mAb-based therapy become ill, with decreased survival compared to animals vaccinated and subsequently treated with MIL77. These results suggest that rVSV-ZEBOV augments immunotherapy.


Assuntos
Anticorpos Monoclonais/uso terapêutico , Anticorpos Antivirais/uso terapêutico , Vacinas contra Ebola/imunologia , Doença pelo Vírus Ebola/imunologia , Doença pelo Vírus Ebola/prevenção & controle , Profilaxia Pós-Exposição , Vacinação , Vírus da Estomatite Vesicular Indiana/imunologia , Animais , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais/imunologia , Anticorpos Antivirais/administração & dosagem , Anticorpos Antivirais/imunologia , Doença pelo Vírus Ebola/patologia , Doença pelo Vírus Ebola/virologia , Humanos , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Estimativa de Kaplan-Meier , Macaca mulatta , Resultado do Tratamento , Carga Viral/imunologia
6.
Trends Biotechnol ; 38(9): 943-947, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32600777

RESUMO

Vaccine solutions rarely reach the public until after an outbreak abates; an Ebola vaccine was approved 5 years after peak outbreak and SARS, MERS, and Zika vaccines are still in clinical development. Despite massive leaps forward in rapid science, other regulatory bottlenecks are hamstringing the global effort for pandemic vaccines.


Assuntos
Infecções por Coronavirus/prevenção & controle , Aprovação de Drogas/organização & administração , Doença pelo Vírus Ebola/prevenção & controle , Influenza Humana/prevenção & controle , Pandemias/prevenção & controle , Pneumonia Viral/prevenção & controle , Vacinas Virais/biossíntese , Betacoronavirus/efeitos dos fármacos , Betacoronavirus/imunologia , Betacoronavirus/patogenicidade , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/virologia , Vacinas contra Ebola/administração & dosagem , Vacinas contra Ebola/biossíntese , Ebolavirus/efeitos dos fármacos , Ebolavirus/imunologia , Ebolavirus/patogenicidade , Europa (Continente)/epidemiologia , Saúde Global/tendências , Regulamentação Governamental , Doença pelo Vírus Ebola/epidemiologia , Doença pelo Vírus Ebola/imunologia , Doença pelo Vírus Ebola/virologia , Humanos , Vírus da Influenza A Subtipo H1N1/genética , Vírus da Influenza A Subtipo H1N1/imunologia , Vacinas contra Influenza/administração & dosagem , Vacinas contra Influenza/biossíntese , Influenza Humana/epidemiologia , Influenza Humana/imunologia , Influenza Humana/virologia , Coronavírus da Síndrome Respiratória do Oriente Médio/efeitos dos fármacos , Coronavírus da Síndrome Respiratória do Oriente Médio/imunologia , Coronavírus da Síndrome Respiratória do Oriente Médio/patogenicidade , Pneumonia Viral/epidemiologia , Pneumonia Viral/imunologia , Pneumonia Viral/virologia , Vírus da SARS/efeitos dos fármacos , Vírus da SARS/imunologia , Vírus da SARS/patogenicidade , Síndrome Respiratória Aguda Grave/epidemiologia , Síndrome Respiratória Aguda Grave/imunologia , Síndrome Respiratória Aguda Grave/prevenção & controle , Síndrome Respiratória Aguda Grave/virologia , Estados Unidos/epidemiologia , Vacinas Virais/administração & dosagem , Zika virus/efeitos dos fármacos , Zika virus/imunologia , Zika virus/patogenicidade , Infecção por Zika virus/epidemiologia , Infecção por Zika virus/imunologia , Infecção por Zika virus/prevenção & controle , Infecção por Zika virus/virologia
7.
PLoS Negl Trop Dis ; 14(7): e0007960, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32687500

RESUMO

Intracerebral microhemorrhages (CMHs) are small foci of hemorrhages in the cerebrum. Acute infections induced by some intracellular pathogens, including rickettsia, can result in CMHs. Annexin a2 (ANXA2) has been documented to play a functional role during intracellular bacterial adhesion. Here we report that ANXA2-knockout (KO) mice are more susceptible to CMHs in response to rickettsia and Ebola virus infections, suggesting an essential role of ANXA2 in protecting vascular integrity during these intracellular pathogen infections. Proteomic analysis via mass spectrometry of whole brain lysates and brain-derived endosomes from ANXA2-KO and wild-type (WT) mice post-infection with R. australis revealed that a variety of significant proteins were differentially expressed, and the follow-up function enrichment analysis had identified several relevant cell-cell junction functions. Immunohistology study confirmed that both infected WT and infected ANXA2-KO mice were subjected to adherens junctional protein (VE-cadherin) damages. However, key blood-brain barrier (BBB) components, tight junctional proteins ZO-1 and occludin, were disorganized in the brains from R. australis-infected ANXA2-KO mice, but not those of infected WT mice. Similar ANXA2-KO dependent CMHs and fragments of ZO-1 and occludin were also observed in Ebola virus-infected ANXA2-KO mice, but not found in infected WT mice. Overall, our study revealed a novel role of ANXA2 in the formation of CMHs during R. australis and Ebola virus infections; and the underlying mechanism is relevant to the role of ANXA2-regulated tight junctions and its role in stabilizing the BBB in these deadly infections.


Assuntos
Anexina A2/metabolismo , Hemorragia Cerebral/metabolismo , Ebolavirus/fisiologia , Doença pelo Vírus Ebola/metabolismo , Infecções por Rickettsia/metabolismo , Rickettsia/fisiologia , Animais , Anexina A2/genética , Antígenos CD/genética , Antígenos CD/metabolismo , Caderinas/genética , Caderinas/metabolismo , Hemorragia Cerebral/genética , Hemorragia Cerebral/microbiologia , Hemorragia Cerebral/virologia , Endossomos/genética , Endossomos/metabolismo , Doença pelo Vírus Ebola/genética , Doença pelo Vírus Ebola/virologia , Humanos , Camundongos , Camundongos Knockout , Rickettsia/genética , Infecções por Rickettsia/genética , Infecções por Rickettsia/microbiologia
9.
Appl Environ Microbiol ; 86(17)2020 08 18.
Artigo em Inglês | MEDLINE | ID: mdl-32591388

RESUMO

The infection of health care workers during the 2013 to 2016 Ebola outbreak raised concerns about fomite transmission. In the wake of the coronavirus disease 2019 (COVID-19) pandemic, investigations are ongoing to determine the role of fomites in coronavirus transmission as well. The bacteriophage phi 6 has a phospholipid envelope and is commonly used in environmental studies as a surrogate for human enveloped viruses. The persistence of phi 6 was evaluated as a surrogate for Ebola virus (EBOV) and coronaviruses on porous and nonporous hospital surfaces. Phi 6 was suspended in a body fluid simulant and inoculated onto 1-cm2 coupons of steel, plastic, and two fabric curtain types. The coupons were placed at two controlled absolute humidity (AH) levels: a low AH of 3.0 g/m3 and a high AH of 14.4 g/m3 Phi 6 declined at a lower rate on all materials under low-AH conditions, with a decay rate of 0.06-log10 PFU/day to 0.11-log10 PFU/day, than under the higher AH conditions, with a decay rate of 0.65-log10 PFU/h to 1.42-log10 PFU/day. There was a significant difference in decay rates between porous and nonporous surfaces at both low AH (P < 0.0001) and high AH (P < 0.0001). Under these laboratory-simulated conditions, phi 6 was found to be a conservative surrogate for EBOV under low-AH conditions in that it persisted longer than Ebola virus in similar AH conditions. Additionally, some coronaviruses persist longer than phi 6 under similar conditions; therefore, phi 6 may not be a suitable surrogate for coronaviruses.IMPORTANCE Understanding the persistence of enveloped viruses helps inform infection control practices and procedures in health care facilities and community settings. These data convey to public health investigators that enveloped viruses can persist and remain infective on surfaces, thus demonstrating a potential risk for transmission. Under these laboratory-simulated Western indoor hospital conditions, we assessed the suitability of phi 6 as a surrogate for environmental persistence research related to enveloped viruses, including EBOV and coronaviruses.


Assuntos
Bacteriófago phi 6/isolamento & purificação , Bacteriófago phi 6/fisiologia , Coronavirus/fisiologia , Ebolavirus/fisiologia , Microbiologia Ambiental , Fômites/virologia , Inativação de Vírus , Betacoronavirus/fisiologia , Coronavirus/isolamento & purificação , Infecções por Coronavirus/transmissão , Infecções por Coronavirus/virologia , Ebolavirus/isolamento & purificação , Doença pelo Vírus Ebola/transmissão , Doença pelo Vírus Ebola/virologia , Hospitais , Humanos , Umidade , Pandemias , Pneumonia Viral/transmissão , Porosidade , Temperatura
11.
PLoS Negl Trop Dis ; 14(5): e0008327, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32407312

RESUMO

BACKGROUND: Against the background of the international public health emergency related to the Ebola outbreak in the Democratic Republic of Congo, in addition to other recent large Ebola epidemics, the issue of transmission due to viral persistence from survivors' body fluids is becoming increasingly urgent. Clinical research in which body fluids play a role is critical and semen testing programs are part of the suggested response to the outbreak. Broad acceptance and understanding of testing programs and research, often in resource poor settings, is essential for the success and sustainability of clinical studies and an accurate epidemic response. Study participants' perceptions on the collection of body fluids are therefore relevant for the programmatic planning and implementation of clinical studies. STUDY AIM AND METHODS: In this qualitative study we aimed to explore the perceptions on bio-sampling in the Sierra Leone Ebola Virus Persistence Study (SLEVP study). We were interested to understand how norms on gender and sexuality related to perceptions and experiences of study participants and staff, specifically, in what way perceptions of the body, on intimacy and on body fluids related to the study process. We purposively sampled former study participants for in-depth interviews and focus-group discussions. We conducted 56 in-depth interviews and eight focus group discussions with 93 participants. In a participatory approach we included study participants in the analysis of data. RESULTS: Overall the SLEVP study was well perceived by study participants and study staff. Study participants conceived the testing of their body fluids positively and saw it as a useful means to know their status. However, some study participants were ambivalent and sometimes reluctant towards sampling of certain body fluids (especially semen, blood and vaginal fluid) due to religious or cultural reasons. Self-sampling was described by study participants as a highly unusual phenomenon. Several narratives were related to the loss of body fluids (especially semen) that would make men weak and powerless, or women dizzy and sick (especially blood). Some rumors indicated mistrust related to study aims that may have been expressions of broader societal challenges and historical circumstances. These reservations could eventually be overcome by guaranteeing confidentiality and privacy and by comprehensive professional counseling. CONCLUSION: In the course of the sampling exercise, study participants were often obliged to transgress cultural and intimate boundaries. It is therefore important to understand the potential importance some of these perceptions have on the recruitment of study participants and the acceptability of studies, on a symbolic as well as a structural level. In order to capture any reservations it is necessary to provide plenty of possibilities of information sharing and follow-up of continuous consent.


Assuntos
Pesquisa Biomédica/métodos , Líquidos Corporais/virologia , Transmissão de Doença Infecciosa/prevenção & controle , Ebolavirus/isolamento & purificação , Doença pelo Vírus Ebola/transmissão , Aceitação pelo Paciente de Cuidados de Saúde/psicologia , Manejo de Espécimes/psicologia , Adolescente , Adulto , Idoso , Feminino , Doença pelo Vírus Ebola/virologia , Humanos , Entrevistas como Assunto , Masculino , Pessoa de Meia-Idade , Serra Leoa , Adulto Jovem
12.
Am J Pathol ; 190(7): 1449-1460, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32275904

RESUMO

Zaire ebolavirus (EBOV) causes Ebola virus disease (EVD), which carries a fatality rate between 25% and 90% in humans. Liver pathology is a hallmark of terminal EVD; however, little is known about temporal disease progression. We used multiplexed fluorescent immunohistochemistry and in situ hybridization in combination with whole slide imaging and image analysis (IA) to quantitatively characterize temporospatial signatures of viral and host factors as related to EBOV pathogenesis. Eighteen rhesus monkeys euthanized between 3 and 8 days post-infection, and 3 uninfected controls were enrolled in this study. Compared with semiquantitative histomorphologic ordinal scoring, quantitative IA detected subtle and progressive features of early and terminal EVD that was not feasible with routine approaches. Sinusoidal macrophages were the earliest cells to respond to infection, expressing proinflammatory cytokine interleukin 6 (IL6) mRNA, which was subsequently also observed in fibrovascular compartments. The mRNA of interferon-stimulated gene-15 (ISG-15), also known as ISG15 ubiquitin like modifier (ISG15), was observed early, with a progressive and ubiquitous hybridization signature involving mesenchymal and epithelial compartments. ISG-15 mRNA was prominent near infected cells, but not in infected cells, supporting the hypothesis that bystander cells produce a robust interferon gene response. This study contributes to our current understanding of early EVD progression and illustrates the value that digital pathology and quantitative IA serve in infectious disease research.


Assuntos
Biomarcadores/análise , Doença pelo Vírus Ebola/patologia , Doença pelo Vírus Ebola/virologia , Interações Hospedeiro-Patógeno/fisiologia , Fígado/virologia , Animais , Ebolavirus , Feminino , Doença pelo Vírus Ebola/imunologia , Fígado/imunologia , Fígado/patologia , Estudos Longitudinais , Macaca mulatta , Masculino
14.
Infect Dis Poverty ; 9(1): 40, 2020 Apr 21.
Artigo em Inglês | MEDLINE | ID: mdl-32312320

RESUMO

BACKGROUND: Following the West Africa Ebola virus disease (EVD) outbreak (2013-2016), WHO developed a preparedness checklist for its member states. This checklist is currently being applied for the first time on a large and systematic scale to prepare for the cross border importation of the ongoing EVD outbreak in the Democratic Republic of Congo hence the need to document the lessons learnt from this experience. This is more pertinent considering the complex humanitarian context and weak health system under which some of the countries such as the Republic of South Sudan are implementing their EVD preparedness interventions. MAIN TEXT: We identified four main lessons from the ongoing EVD preparedness efforts in the Republic South Sudan. First, EVD preparedness is possible in complex humanitarian settings such as the Republic of South Sudan by using a longer-term health system strengthening approach. Second, the Republic of South Sudan is at risk of both domestic and cross border transmission of EVD and several other infectious disease outbreaks hence the need for an integrated and sustainable approach to outbreak preparedness in the country. Third, a phased and well-prioritized approach is required for EVD preparedness in complex humanitarian settings given the costs associated with preparedness and the difficulties in the accurate prediction of outbreaks in such settings. Fourth, EVD preparedness in complex humanitarian settings is a massive undertaking that requires effective and decentralized coordination. CONCLUSION: Despite a very challenging context, the Republic of South Sudan made significant progress in its EVD preparedness drive demonstrating that it is possible to rapidly scale up preparedness efforts in complex humanitarian contexts if appropriate and context-specific approaches are used. Further research, systematic reviews and evaluation of the ongoing preparedness efforts are required to ensure comprehensive documentation and application of the lessons learnt for future EVD outbreak preparedness and response efforts.


Assuntos
Emigração e Imigração/estatística & dados numéricos , Doença pelo Vírus Ebola/transmissão , Surtos de Doenças/prevenção & controle , Surtos de Doenças/estatística & dados numéricos , Ebolavirus/genética , Ebolavirus/fisiologia , Doença pelo Vírus Ebola/epidemiologia , Doença pelo Vírus Ebola/prevenção & controle , Doença pelo Vírus Ebola/virologia , Humanos , Socorro em Desastres/estatística & dados numéricos , Sudão do Sul/epidemiologia
15.
PLoS Negl Trop Dis ; 14(3): e0008117, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32130210

RESUMO

Ebolaviruses pose a substantial threat to wildlife populations and to public health in Africa. Evolutionary analyses of virus genome sequences can contribute significantly to elucidate the origin of new outbreaks, which can help guide surveillance efforts. The reconstructed between-outbreak evolutionary history of Zaire ebolavirus so far has been highly consistent. By removing the confounding impact of population growth bursts during local outbreaks on the free mixing assumption that underlies coalescent-based demographic reconstructions, we find-contrary to what previous results indicated-that the circulation dynamics of Ebola virus in its animal reservoir are highly uncertain. Our findings also accentuate the need for a more fine-grained picture of the Ebola virus diversity in its reservoir to reliably infer the reservoir origin of outbreak lineages. In addition, the recent appearance of slower-evolving variants is in line with latency as a survival mechanism and with bats as the natural reservoir host.


Assuntos
Doenças dos Animais/epidemiologia , Quirópteros/virologia , Reservatórios de Doenças/virologia , Ebolavirus/isolamento & purificação , Doença pelo Vírus Ebola/veterinária , África , Doenças dos Animais/virologia , Animais , Ebolavirus/classificação , Ebolavirus/genética , Genótipo , Doença pelo Vírus Ebola/epidemiologia , Doença pelo Vírus Ebola/virologia , Humanos , Filogenia
16.
PLoS One ; 15(3): e0230322, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32182277

RESUMO

First reported in March 2014, an Ebola epidemic impacted West Africa, most notably Liberia, Guinea and Sierra Leone. We demonstrate the value of social media for automated surveillance of infectious diseases such as the West Africa Ebola epidemic. We experiment with two variations of an existing surveillance architecture: the first aggregates tweets related to different symptoms together, while the second considers tweets about each symptom separately and then aggregates the set of alerts generated by the architecture. Using a dataset of tweets posted from the affected region from 2011 to 2014, we obtain alerts in December 2013, which is three months prior to the official announcement of the epidemic. Among the two variations, the second, which produces a restricted but useful set of alerts, can potentially be applied to other infectious disease surveillance and alert systems.


Assuntos
Mineração de Dados/métodos , Epidemias/prevenção & controle , Monitoramento Epidemiológico , Doença pelo Vírus Ebola/epidemiologia , Mídias Sociais/estatística & dados numéricos , Conjuntos de Dados como Assunto , Ebolavirus , Epidemias/estatística & dados numéricos , Guiné/epidemiologia , Doença pelo Vírus Ebola/diagnóstico , Doença pelo Vírus Ebola/virologia , Humanos , Libéria/epidemiologia , Serra Leoa/epidemiologia
17.
J Virol ; 94(8)2020 03 31.
Artigo em Inglês | MEDLINE | ID: mdl-32051271

RESUMO

Given that the Ebola virus (EBOV) infects a wide array of organs and cells yet displays a relative lack of neurotropism, we asked whether a chimeric vesicular stomatitis virus (VSV) expressing the EBOV glycoprotein (GP) might selectively target brain tumors. The mucin-like domain (MLD) of the EBOV GP may enhance virus immune system evasion. Here, we compared chimeric VSVs in which EBOV GP replaces the VSV glycoprotein, thereby reducing the neurotoxicity associated with wild-type VSV. A chimeric VSV expressing the full-length EBOV GP (VSV-EBOV) containing the MLD was substantially more effective and safer than a parallel construct with an EBOV GP lacking the MLD (VSV-EBOVΔMLD). One-step growth, reverse transcription-quantitative PCR, and Western blotting assessments showed that VSV-EBOVΔMLD produced substantially more progeny faster than VSV-EBOV. Using immunodeficient SCID mice, we focused on targeting human brain tumors with these VSV-EBOVs. Similar to the findings of our previous study in which we used an attenuated VSV-EBOV with no MLD that expressed green fluorescent protein (GFP) (VSV-EBOVΔMLD-GFP), VSV-EBOVΔMLD without GFP targeted glioma but yielded only a modest extension of survival. In contrast, VSV-EBOV containing the MLD showed substantially better targeting and elimination of brain tumors after intravenous delivery and increased the survival of brain tumor-bearing mice. Despite the apparent destruction of most tumor cells by VSV-EBOVΔMLD, the virus remained active within the SCID mouse brain and showed widespread infection of normal brain cells. In contrast, VSV-EBOV eliminated the tumors and showed relatively little infection of normal brain cells. Parallel experiments with direct intracranial virus infection generated similar results. Neither VSV-EBOV nor VSV-EBOVΔMLD showed substantive infection of the brains of normal immunocompetent mice.IMPORTANCE The Ebola virus glycoprotein contains a mucin-like domain which may play a role in immune evasion. Chimeric vesicular stomatitis viruses with the EBOV glycoprotein substituted for the VSV glycoprotein show greater safety and efficacy in targeting brain tumors in immunodeficient mice when the MLD was expressed within the EBOV glycoprotein than when EBOV lacked the mucin-like domain.


Assuntos
Neoplasias Encefálicas/metabolismo , Ebolavirus/imunologia , Glicoproteínas/imunologia , Doença pelo Vírus Ebola/virologia , Mucinas/imunologia , Animais , Neoplasias Encefálicas/patologia , Neoplasias Encefálicas/virologia , Linhagem Celular Tumoral , Modelos Animais de Doenças , Ebolavirus/genética , Glioblastoma/virologia , Glioma/patologia , Glioma/virologia , Proteínas de Fluorescência Verde , Xenoenxertos , Humanos , Camundongos , Camundongos SCID , Mucinas/genética , Vírus da Estomatite Vesicular Indiana/imunologia
18.
mBio ; 11(1)2020 02 25.
Artigo em Inglês | MEDLINE | ID: mdl-32098814

RESUMO

Ebola virus (EBOV) causes a severe and often fatal disease for which no approved vaccines or antivirals are currently available. EBOV VP30 has been described as a viral phosphoprotein, and nonphosphorylated VP30 is essential and sufficient to support secondary transcription in an EBOV-specific minigenome system; however, phosphorylatable serine residues near the N terminus of VP30 are required to support primary viral transcription as well as the reinitiation of VP30-mediated transcription at internal EBOV genes. While the dephosphorylation of VP30 by the cellular phosphatase PP2A was found to be mediated by nucleoprotein, the VP30-specific kinases and the role of phosphorylation remain unknown. Here, we report that serine-arginine protein kinase 1 (SRPK1) and SRPK2 phosphorylate serine 29 of VP30, which is located in an N-terminal R26xxS29 motif. Interaction with VP30 via the R26xxS29 motif recruits SRPK1 into EBOV-induced inclusion bodies, the sites of viral RNA synthesis, and an inhibitor of SRPK1/SRPK2 downregulates primary viral transcription. When the SRPK1 recognition motif of VP30 was mutated in a recombinant EBOV, virus replication was severely impaired. It is presumed that the interplay between SRPK1 and PP2A in the EBOV inclusions provides a comprehensive regulatory circuit to ensure the activity of VP30 in EBOV transcription. Thus, the identification of SRPK1 is an important mosaic stone that completes our picture of the players involved in Ebola virus transcription regulation.IMPORTANCE The largest Ebola virus (EBOV) epidemic in West Africa ever caused more than 28,000 cases and 11,000 deaths, and the current EBOV epidemic in the Democratic Republic of the Congo continues, with more than 3,000 cases to date. Therefore, it is essential to develop antivirals against EBOV. Recently, an inhibitor of the cellular phosphatase PP2A-mediated dephosphorylation of the EBOV transcription factor VP30 has been shown to suppress the spread of Ebola virus. Here, we identified the protein kinase SRPK1 as a VP30-specific kinase that phosphorylates serine 29, the same residue that is dephosphorylated by PP2A. SRPK1-mediated phosphorylation of serine 29 enabled primary viral transcription. Mutation of the SRPK1 recognition motif in VP30 resulted in significant growth inhibition of EBOV. Similarly, elevation of the phosphorylation status of serine 29 by overexpression of SRPK1 inhibited EBOV growth, highlighting the importance of reversible phosphorylation of VP30 as a potential therapeutic target.


Assuntos
Ebolavirus/fisiologia , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Virais/metabolismo , Replicação Viral/fisiologia , África Ocidental , Animais , Linhagem Celular , Sobrevivência Celular , Chlorocebus aethiops , Ebolavirus/genética , Células HEK293 , Doença pelo Vírus Ebola/virologia , Humanos , Corpos de Inclusão Viral/metabolismo , Nucleoproteínas , Fosforilação , RNA Viral/genética , Fatores de Transcrição/metabolismo , Células Vero , Replicação Viral/genética
19.
Proc Natl Acad Sci U S A ; 117(7): 3768-3778, 2020 02 18.
Artigo em Inglês | MEDLINE | ID: mdl-32015126

RESUMO

Antibody-based therapies are a promising treatment option for managing ebolavirus infections. Several Ebola virus (EBOV)-specific and, more recently, pan-ebolavirus antibody cocktails have been described. Here, we report the development and assessment of a Sudan virus (SUDV)-specific antibody cocktail. We produced a panel of SUDV glycoprotein (GP)-specific human chimeric monoclonal antibodies (mAbs) using both plant and mammalian expression systems and completed head-to-head in vitro and in vivo evaluations. Neutralizing activity, competitive binding groups, and epitope specificity of SUDV mAbs were defined before assessing protective efficacy of individual mAbs using a mouse model of SUDV infection. Of the mAbs tested, GP base-binding mAbs were more potent neutralizers and more protective than glycan cap- or mucin-like domain-binding mAbs. No significant difference was observed between plant and mammalian mAbs in any of our in vitro or in vivo evaluations. Based on in vitro and rodent testing, a combination of two SUDV-specific mAbs, one base binding (16F6) and one glycan cap binding (X10H2), was down-selected for assessment in a macaque model of SUDV infection. This cocktail, RIID F6-H2, provided protection from SUDV infection in rhesus macaques when administered at 50 mg/kg on days 4 and 6 postinfection. RIID F6-H2 is an effective postexposure SUDV therapy and provides a potential treatment option for managing human SUDV infection.


Assuntos
Anticorpos Antivirais/administração & dosagem , Ebolavirus/imunologia , Doença pelo Vírus Ebola/tratamento farmacológico , Animais , Anticorpos Monoclonais/administração & dosagem , Modelos Animais de Doenças , Ebolavirus/genética , Feminino , Glicoproteínas/imunologia , Doença pelo Vírus Ebola/virologia , Humanos , Imunoterapia , Macaca mulatta , Masculino , Camundongos , Proteínas Virais/imunologia
20.
EBioMedicine ; 52: 102601, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31953031

RESUMO

BACKGROUND: TKM-130803 is a specific anti-EBOV therapeutic comprised of two small interfering RNAs (siRNA) siLpol-2 and siVP35-2. The pharmacokinetics (PK) of these siRNAs was defined in Ebola virus disease (EVD) patients, with reference to efficacy (ET) and toxicology thresholds (TT). The relationship between PK and patient survival was explored. METHODS: Pharmacokinetic (PK) and pharmacodynamic (PD) data were available for seven participants with EVD in Sierra Leone who received 0·3 mg/kg of TKM-130803 by intravenous infusion over 2 h daily for up to 7 days. Plasma concentration of siRNA was compared to survival at 14 days. PK data were fitted to two-compartment models then Monte Carlo simulated PK profiles were compared to ET (Cmax 0·04-0·57 ng/mL and mean concentration 1·43 ng/mL), and TT (3000 ng/mL). FINDINGS: Viral loads (VL) were not significantly different at treatment onset or during treatment (p = 0·1) in subjects who survived or died. siRNA was in quantitative excess of virus genomes throughout treatment, but the 95% percentile exceeded TT. The maximum AUC for which the 95% percentile remained under TT was a continuous infusion of 0·15 mg/kg/day. Plasma concentration of both siRNAs were higher in subjects who died compared to subjects who survived (p<0·025 both siRNAs). INTERPRETATION: TKM-130803 was circulating in molar excess of circulating virus; a level considered needed for efficacy. Given extremely high viral loads it seems likely that the patients died because they were physiologically beyond the point of no return. Subjects who died exhibited some indication of impaired drug clearance, justifying caution in dosing strategies for such patients. This analysis has given a useful insight into the pharmacokinetics of the siRNA in the disease state and illustrates the value of designing PKPD studies into future clinical trials in epidemic situations. FUNDING: This work was supported by the Wellcome Trust of Great Britain (grant number 106491/Z/14/Z and 097997/Z/11/A) and by the EU FP7 project PREPARE (602525). The PHE laboratory was funded by the UK Department for International Development. The funders had no role in trial design, data collection or analysis. The views expressed are those of the authors and not necessarily those of Public Health England, the Department of Health, or the EU. TRIAL REGISTRATION: Pan African Clinical Trials Registry PACTR201501000997429.


Assuntos
Antivirais/farmacocinética , Doença pelo Vírus Ebola/tratamento farmacológico , Doença pelo Vírus Ebola/virologia , RNA Interferente Pequeno/farmacocinética , Algoritmos , Antivirais/administração & dosagem , Simulação por Computador , Relação Dose-Resposta a Droga , Avaliação Pré-Clínica de Medicamentos , Monitoramento de Medicamentos , Doença pelo Vírus Ebola/diagnóstico , Doença pelo Vírus Ebola/mortalidade , Humanos , Modelos Teóricos , RNA Interferente Pequeno/administração & dosagem , Índice de Gravidade de Doença , Serra Leoa , Resultado do Tratamento , Carga Viral
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