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Eastern equine encephalitis virus (EEEV), Madariaga virus (MADV), and Venezuelan equine encephalitis virus complex (VEEV) are New World alphaviruses transmitted by mosquitoes. They cause febrile and sometimes severe neurological diseases in human and equine hosts. Detecting them during the acute phase is hindered by non-specific symptoms and limited diagnostic tools. We designed and clinically assessed real-time reverse transcription polymerase chain reaction assays (rRT-PCRs) for VEEV complex, MADV, and EEEV using whole-genome sequences. Validation involved 15 retrospective serum samples from 2015 to 2017 outbreaks, 150 mosquito pools from 2015, and 118 prospective samples from 2021 to 2022 surveillance in Panama. The rRT-PCRs detected VEEV complex RNA in 10 samples (66.7%) from outbreaks, with one having both VEEV complex and MADV RNAs. VEEV complex RNA was found in five suspected dengue cases from disease surveillance. The rRT-PCR assays identified VEEV complex RNA in three Culex (Melanoconion) vomerifer pools, leading to VEEV isolates in two. Phylogenetic analysis revealed the VEEV ID subtype in positive samples. Notably, 11.9% of dengue-like disease patients showed VEEV infections. Together, our rRT-PCR validation in human and mosquito samples suggests that this method can be incorporated into mosquito and human encephalitic alphavirus surveillance programs in endemic regions.
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Alphavirus , Culicidae , Dengue , Vírus da Encefalite Equina do Leste , Encefalomielite Equina do Leste , Encefalomielite Equina Venezuelana , Humanos , Animais , Cavalos/genética , Vírus da Encefalite Equina do Leste/genética , Encefalomielite Equina Venezuelana/diagnóstico , Encefalomielite Equina Venezuelana/epidemiologia , Culicidae/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Filogenia , Estudos Prospectivos , Vigilância em Saúde Pública , Estudos Retrospectivos , Alphavirus/genética , RNARESUMO
INTRODUCTION: Eastern equine encephalitis virus (EEEV) and Venezuelan equine encephalitis virus (VEEV) viruses are zoonotic pathogens affecting humans, particularly equines. These neuroarboviruses compromise the central nervous system and can be fatal in different hosts. Both have significantly influenced Colombia; however, few studies analyse its behaviour, and none develop maps using geographic information systems to characterise it. OBJECTIVE: To describe the temporal-spatial distribution of those viruses in Colombia between 2008 and 2019. METHODS: Retrospective cross-sectional descriptive study, based on weekly reports by municipalities of the ICA, of the surveillance of both arboviruses in equines, in Colombia, from 2008 to 2019. The data were converted into databases in Microsoft Access 365®, and multiple epidemiological maps were generated with the Kosmo RC1®3.0 software coupled to shape files of all municipalities in the country. RESULTS: In the study period, 96 cases of EEE and 70 of VEE were reported, with 58% of EEE cases occurring in 2016 and 20% of EEV cases in 2013. The most affected municipalities for EEE corresponded to the department of Casanare: Yopal (20), Aguazul (16), and Tauramena (10). In total, 40 municipalities in the country reported ≥1 case of EEE. CONCLUSIONS: The maps allow a quick appreciation of groups of neighbouring municipalities in different departments (1° political division) and regions of the country affected by those viruses, which helps consider the expansion of the disease associated with mobility and transport of equines between other municipalities, also including international borders, such as is the case with Venezuela. In that country, especially for EEV, municipalities in the department of Cesar are bordering and at risk for that arboviral infection. there is a high risk of equine encephalitis outbreaks, especially for VEE. This poses a risk also, for municipalities in the department of Cesar, bordering with Venezuela.
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Vírus da Encefalite Equina Venezuelana , Encefalomielite Equina Venezuelana , Cavalos , Animais , Colômbia/epidemiologia , Estudos Transversais , Encefalomielite Equina Venezuelana/epidemiologia , Sistemas de Informação Geográfica , Cavalos/virologia , Estudos RetrospectivosRESUMO
The purpose of this study was to evaluate the effects of the route of administration on the immunogenicity and efficacy of a combined western, eastern, and Venezuelan equine encephalitis (WEVEE) virus-like replicon particle (VRP) vaccine in cynomolgus macaques. The vaccine consisted of equal amounts of WEEV, EEEV, and VEEV VRPs. Thirty-three animals were randomly assigned to five treatment or control groups. Animals were vaccinated with two doses of WEVEE VRPs or the control 28 days apart. Blood was collected 28 days following primary vaccination and 21 days following boost vaccination for analysis of the immune response to the WEVEE VRP vaccine. NHPs were challenged by aerosol 28 or 29 days following second vaccination with WEEV CBA87. Vaccination with two doses of WEVEE VRP was immunogenic and resulted in neutralizing antibody responses specific for VEEV, EEEV and WEEV. None of the vaccinated animals met euthanasia criteria following aerosol exposure to WEEV CBA87. However, one NHP control (total of 11 controls) met euthanasia criteria after infection with WEEV CBA87. Statistically significant differences in median fever hours were noted in control NHPs compared to vaccinated NHPs, providing a quantitative measure of infection and efficacy of the vaccine against a WEEV challenge. Alterations in lymphocytes, monocytes, and neutrophils were observed. Lymphopenia was observed in control NHPs.
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Vírus da Encefalite Equina Venezuelana , Encefalomielite Equina Venezuelana , Vacinas Virais , Aerossóis , Animais , Anticorpos Antivirais , Modelos Animais de Doenças , Vírus da Encefalite Equina Venezuelana/genética , Encefalomielite Equina Venezuelana/prevenção & controle , Cavalos , Macaca fascicularis , RepliconRESUMO
Vaccination against infectious diseases is a cornerstone of veterinary medicine in the prevention of disease transmission, illness severity, and often death in animals. In North American equine medicine, equine vaccines protecting against tetanus, rabies, Eastern and Western equine encephalomyelitis, and West Nile are core vaccines as these have been classified as having a heightened risk of mortality, infectiousness, and endemic status. Some guidelines differ from the label of vaccines, to improve the protection of patients or to decrease the unnecessary administration to reduce potential side effects. In North America, resources for the equine practitioners are available on the American Association of Equine Practitioners (AAEP) website. Conversely, in small companion animals, peer review materials are regularly published in open access journals to guide the vaccination of dogs and cats. The aims of this review are to present how the vaccine guidelines have been established for small companion animals and horses in North America, to review the equine literature to solidify or contrast the current AAEP guidelines of core vaccines, and to suggest future research directions in the equine vaccine field considering small companion animal strategies and the current available resources in equine literature.
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INTRODUCTION: Arthropod-borne viruses (arboviruses) cause morbidity and mortality in humans and domestic animals worldwide. The percentage of population immunity or susceptibility to these viruses in Ecuador is unknown. OBJECTIVES: To investigate the proportion of Ecuadorian populations with IgG antibodies (Abs) (past exposure/immunity) and IgM Abs (current exposure) against flaviviruses and alphaviruses and to study the activity of these viruses in Ecuador. MATERIALS AND METHODS: During 2009-2011, we conducted a serosurvey for selected arboviruses in humans (n=1,842), equines (n=149), and sentinel hamsters (n=84) at two coastal locations and one in the Amazon basin (Eastern Ecuador) using enzyme-linked immunosorbent assay and hemagglutination inhibition test. RESULTS: From 20.63% to 63.61% of humans showed IgG-antibodies for the flaviviruses: Dengue virus (DENV), yellow fever virus (YFV) Saint Louis encephalitis virus, and West Nile virus (WNV); from 4.67% to 8.63% showed IgG-Abs for the alphaviruses: Venezuelan equine encephalitis virus, eastern equine encephalitis virus, and western equine encephalitis virus. IgM-Abs were found for DENV and WNV. Equines and hamsters showed antibodies to alphaviruses in all locations; two hamsters seroconverted to YFV in the Amazonia. CONCLUSIONS: The results show a YFV vaccination history and suggest the activity of arboviruses not included in the current surveillance scheme. Enhanced arbovirus and mosquito surveillance, as well as continued YFV vaccination and evaluation of its coverage/effectiveness, are recommended.
Introducción. Los virus transmitidos por artrópodos (arbovirus) causan morbilidad y mortalidad en humanos y animales domésticos mundialmente. Se desconoce el porcentaje de inmunidad o vulnerabilidad de la población ecuatoriana ante estos virus. Objetivos. Investigar la proporción de poblaciones ecuatorianas con anticuerpos IgG (exposición o inmunidad pasada) y anticuerpos IgM (exposición reciente) contra flavivirus y alfavirus, e investigar su actividad en Ecuador. Materiales y métodos. Entre 2009 y 2011, se llevó a cabo una encuesta serológica para arbovirus en humanos (n=1.842), equinos (n=149) y hámsters centinela (n=84) en dos localidades costeras y en una en la Amazonía, utilizando la prueba ELISA (Enzyme-Linked ImmunoSorbent Assay) y la prueba de inhibición de la hemaglutinación. Resultados. Entre el 20,63 y el 63,61 % de los humanos registraron IgG contra el virus del dengue (DENV), el de la fiebre amarilla (YFV), el de la encefalitis de San Luis y el del Nilo Occidental (WNV); entre 4,67 y 8,63 % tenían IgG para los virus de la encefalitis equina venezolana, de la encefalitis equina del este y de la encefalitis equina del oeste. Se encontró IgM para DENV y WNV. En los equinos y en los hámsters se encontraron anticuerpos contra alfavirus en todas las localidades muestreadas; dos hámsters mostraron seroconversión a YFV en la Amazonía. Conclusiones. Los resultados del estudio evidenciaron los antecedentes de vacunación contra el YFV y sugieren la actividad de arbovirus no incluidos en el esquema de vigilancia actual. Se recomienda ampliar la vigilancia de arbovirus y mosquitos, continuar con la vacunación contra el YFV, y evaluar su cobertura y efectividad.
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Arbovírus , Vírus do Nilo Ocidental , Animais , Anticorpos Antivirais , Arbovírus/imunologia , Cricetinae , Equador/epidemiologia , Cavalos , Humanos , Imunoglobulina G , Imunoglobulina M , Estudos Soroepidemiológicos , Vírus do Nilo Ocidental/imunologiaRESUMO
Abstract | Introduction: Arthropod-borne viruses (arboviruses) cause morbidity and mortality in humans and domestic animals worldwide. The percentage of population immunity or susceptibility to these viruses in Ecuador is unknown. Objectives: To investigate the proportion of Ecuadorian populations with IgG antibodies (Abs) (past exposure/immunity) and IgM Abs (current exposure) against flaviviruses and alphaviruses and to study the activity of these viruses in Ecuador. Materials and methods: During 2009-2011, we conducted a serosurvey for selected arboviruses in humans (n=1,842), equines (n=149), and sentinel hamsters (n=84) at two coastal locations and one in the Amazon basin (Eastern Ecuador) using enzyme-linked immunosorbent assay and hemagglutination inhibition test. Results: From 20.63% to 63.61% of humans showed IgG-antibodies for the flaviviruses: Dengue virus (DENV), yellow fever virus (YFV) Saint Louis encephalitis virus, and West Nile virus (WNV); from 4.67% to 8.63% showed IgG-Abs for the alphaviruses: Venezuelan equine encephalitis virus, eastern equine encephalitis virus, and western equine encephalitis virus. IgM-Abs were found for DENV and WNV. Equines and hamsters showed antibodies to alphaviruses in all locations; two hamsters seroconverted to YFV in the Amazonia. Conclusions: The results show a YFV vaccination history and suggest the activity of arboviruses not included in the current surveillance scheme. Enhanced arbovirus and mosquito surveillance, as well as continued YFV vaccination and evaluation of its coverage/ effectiveness, are recommended.
Resumen | Introducción. Los virus transmitidos por artrópodos (arbovirus) causan morbilidad y mortalidad en humanos y animales domésticos mundialmente. Se desconoce el porcentaje de inmunidad o vulnerabilidad de la población ecuatoriana ante estos virus. Objetivos. Investigar la proporción de poblaciones ecuatorianas con anticuerpos IgG (exposición o inmunidad pasada) y anticuerpos IgM (exposición reciente) contra flavivirus y alfavirus, e investigar su actividad en Ecuador. Materiales y métodos. Entre 2009 y 2011, se llevó a cabo una encuesta serológica para arbovirus en humanos (n=1.842), equinos (n=149) y hámsters centinela (n=84) en dos localidades costeras y en una en la Amazonía, utilizando la prueba ELISA (Enzyme-Linked ImmunoSorbent Assay) y la prueba de inhibición de la hemaglutinación. Resultados. Entre el 20,63 y el 63,61 % de los humanos registraron IgG contra el virus del dengue (DENV), el de la fiebre amarilla (YFV), el de la encefalitis de San Luis y el del Nilo Occidental (WNV); entre 4,67 y 8,63 % tenían IgG para los virus de la encefalitis equina venezolana, de la encefalitis equina del este y de la encefalitis equina del oeste. Se encontró IgM para DENV y WNV. En los equinos y en los hámsters se encontraron anticuerpos contra alfavirus en todas las localidades muestreadas; dos hámsters mostraron seroconversión a YFV en la Amazonía. Conclusiones. Los resultados del estudio evidenciaron los antecedentes de vacunación contra el YFV y sugieren la actividad de arbovirus no incluidos en el esquema de vigilancia actual. Se recomienda ampliar la vigilancia de arbovirus y mosquitos, continuar con la vacunación contra el YFV, y evaluar su cobertura y efectividad.
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Arbovírus , Vírus do Nilo Ocidental , Vírus da Febre Amarela , Vírus da Dengue , Vírus da Encefalite Equina do Leste , Vírus da Encefalite Equina VenezuelanaRESUMO
During the 2019 Eastern equine encephalitis virus (EEEV) outbreak in Michigan, two 2-month old Mexican wolf pups experienced neurologic signs, lymphohistiocytic neutrophilic meningoencephalitis with neuronal necrosis and neuronophagia, and acute death. We identified EEEV by reverse transcription real-time PCR and in situ hybridization. Vector mosquitoes were trapped at the zoo.
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Vírus da Encefalite Equina do Leste , Encefalomielite Equina do Leste , Encefalomielite Equina , Lobos , Animais , Cavalos , Michigan , Mosquitos VetoresRESUMO
Serum samples from 89 equids were analyzed (75 horses, 9 donkeys, and 5 mules) from the municipality of Viseu, Pará state, Brazil. Samples were collected in November 2014 and August 2015. The antibody prevalence against the following alphaviruses was estimated: Eastern equine encephalitis virus, Western equine encephalitis virus, Mucambo virus, and Mayaro virus. Seroprevalence was determined by the hemagglutination inhibition (HI) technique. Sera that exhibited HI antibodies with heterotypic reactions for the analyzed viruses were subjected to the 90% plaque reduction neutralization test (PRNT90). The HI prevalence of monotypic reactions to EEEV was 7.9%, and that of WEEV was 1.1%, as confirmed by PRNT90. Viral isolation attempts were negative for all tested blood samples. Our results suggest the circulation of equine encephalitis complex viruses. Future studies should evaluate the possible involvement of arthropod hosts and residents in the viral transmission in the study area.
Foram analisadas amostras de soros de 89 equídeos (75 equinos, 9 asininos e 5 muares) oriundos do município de Viseu, estado do Pará. As amostras foram coletadas no período de novembro de 2014 e agosto de 2015. Foram estimadas as prevalências de anticorpos contra os seguintes Alfavírus: vírus da Encefalite equina leste (EEEV), vírus da Encefalite equina oeste (WEEV), vírus Mucambo (MUCV), and vírus Mayaro (MAYV). A soroprevalência foi determinada por técnicas de Inibição da Hemaglutinação (IH), porém os soros que apresentaram anticorpos IH com reações heterotípicas para os vírus analisados foram submetidos ao PRNT90. A prevalência de reações monotípicas por IH para EEEV foi de 7, 9% e para WEEV foi de 1,1%, confirmadas por PRNT90. A tentativa de isolamento viral foi negativa em todas as amostras de sangue testadas. Nossos resultados sugerem a circulação de vírus do complexo das encefalites equinas. Estudos futuros devem ser realizados para avaliar possível envolvimento de hospedeiros artrópodes e residentes na área de abrangência do estudo.
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Venezuelan, eastern, and western equine encephalitis viruses (VEEV, EEEV, and WEEV) are mosquito-borne viruses in the Americas that cause central nervous system (CNS) disease in humans and equids. In this study, we directly characterized the pathogenesis of VEEV, EEEV, and WEEV in cynomolgus macaques following subcutaneous exposure because this route more closely mimics natural infection via mosquito transmission or by an accidental needle stick. Our results highlight how EEEV is significantly more pathogenic compared to VEEV similarly to what is observed in humans. Interestingly, EEEV appears to be just as neuropathogenic by subcutaneous exposure as it was in previously completed aerosol exposure studies. In contrast, subcutaneous exposure of cynomolgus macaques with WEEV caused limited disease and is contradictory to what has been reported for aerosol exposure. Several differences in viremia, hematology, or tissue tropism were noted when animals were exposed subcutaneously compared to prior aerosol exposure studies. This study provides a more complete picture of the pathogenesis of the encephalitic alphaviruses and highlights how further defining the neuropathology of these viruses could have important implications for the development of medical countermeasures for the neurovirulent alphaviruses.
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Vírus da Encefalite Equina do Leste/patogenicidade , Vírus da Encefalite Equina Venezuelana/patogenicidade , Vírus da Encefalite Equina do Oeste/patogenicidade , Encefalomielite Equina/patologia , Encefalomielite Equina Venezuelana/patologia , Macaca fascicularis/virologia , Animais , Anticorpos Neutralizantes/sangue , Anticorpos Antivirais/sangue , Feminino , Masculino , Replicação ViralRESUMO
A total of 102 free-range wild boars, 170 hunting dogs, and 49 hunters from 3 Brazilian regions were sampled and tested for antibodies to eastern equine encephalitis virus (EEEV), western equine encephalitis virus, and Venezuelan equine encephalitis virus. Three of the 102 (2.9%) wild boars were positive for antibodies against EEEV by microplate serum neutralization test. Based on our data, free-range wild boars from central-western Brazil may be exposed to EEEV, and further studies are needed to evaluate the potential of incorporating serosurveys in routine arbovirus activity surveillance specifically to identify arbovirus activity foci and to help establish thresholds for epidemic transmission.
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Doenças do Cão/virologia , Vírus da Encefalite Equina do Leste , Vírus da Encefalite Equina Venezuelana , Vírus da Encefalite Equina do Oeste , Encefalomielite Equina/veterinária , Doenças dos Suínos/virologia , Animais , Anticorpos Antivirais/sangue , Brasil/epidemiologia , Doenças do Cão/epidemiologia , Cães , Encefalomielite Equina/epidemiologia , Encefalomielite Equina/virologia , Humanos , Estudos Soroepidemiológicos , Sus scrofa , Suínos , Doenças dos Suínos/epidemiologia , Cães TrabalhadoresRESUMO
Vaccinations are a crucial intervention in combating infectious diseases. The three neurotropic Alphaviruses, Eastern (EEEV), Venezuelan (VEEV), and Western (WEEV) equine encephalitis viruses, are pathogens of interest for animal health, public health, and biological defense. In both equines and humans, these viruses can cause febrile illness that may progress to encephalitis. Currently, there are no licensed treatments or vaccines available for these viruses in humans. Experimental vaccines have shown variable efficacy and may cause severe adverse effects. Here, we outline recent strategies used to generate vaccines against EEEV, VEEV, and WEEV with an emphasis on virus-vectored and plasmid DNA delivery. Despite candidate vaccines protecting against one of the three viruses, few studies have demonstrated an effective trivalent vaccine. We evaluated the potential of published vaccines to generate cross-reactive protective responses by comparing DNA vaccine sequences to a set of EEEV, VEEV, and WEEV genomes and determining the vaccine coverages of potential epitopes. Finally, we discuss future directions in the development of vaccines to combat EEEV, VEEV, and WEEV.
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Introducción: En junio de 2019 surgió un nuevo brote de infección por virus de encefalitis equina del este (EEE) y encefalitis equina venezolana (EEV) en Panamá, en pacientes provenientes de la misma área. La infección tiene un amplio espectro de sintomatología pudiendo provocar graves cuadros de encefalitis. En este artículo se pretenden analizar las características de los casos en los que hubo confirmación serológica de la infección, centrándose en la población menor a 16 años atendida en el Hospital del Niño Dr. José Renán Esquivel. Material y métodos: Se realizó un estudio observacional retrospectivo de los casos reportados como encefalitis equina por el sistema de vigilancia epidemiológica del servicio de Epidemiología del Hospital del Niño Dr. José Renán Esquivel de Panamá en los meses de junio y julio de 2019. Los datos fueron analizados con el programa estadístico SPSS. Resultados: Se notificaron 12 casos sospechosos de encefalitis, 6 de ellos fueron confirmados como encefalitis equina del este y/o venezolana. En todos los casos, los pacientes tuvieron una fase prodrómica con fiebre y presentaron convulsiones. Un 66.6% de los pacientes precisó ventilación mecánica. Hubo un fallecimiento. Conclusiones: La infección por el virus de la encefalitis equina puede dar lugar a cuadros neurológicos severos e incluso conducir a la muerte. Dado que no existe un tratamiento específico, es necesaria la vigilancia epidemiológica así como investigación en la prevención de esta infección a través de nuevas vacunas.
Introduction: In June 2019 a new Eastern equine encephalitis (EEE) and Venezuelan equine encephalitis (VEE) outbreak emerged in Panamá in patients coming from the same area. The infection has a large spectrum of symptoms being able to lead to severe encephalitis. This article aims to analyze the characteristics of the cases in which it was serological confirmation, focusing on the population under 16 years old attended at the Hospital del Niño Dr. José Renán Esquivel. Material and methods: An observational retrospective study was carried out among the cases reported as encephalitis by the surveillance system of the Epidemiology Service at Hospital del Niño Dr. José Renán Esquivel from Panama in June and July 2019. Results: 12 cases were notified as suspected encephalitis cases, 6 of them were confirmed with serology tests as Eastern equine encephalitis (EEE) and/or Venezuelan equine encephalitis (VEE). All cases had a prodromic phase with fever and in all cases patients had seizures. A 66.6% of patients needed mechanic ventilation. There was one decease. Conclusions: Equine encephalitis virus can cause severe neurological symptoms and even lead to death. Since there is no specific treatment, it's necessary a close epidemiologic surveillance as well as more investigation in the prevention of this infection trough new vaccines.
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Currently, there are no licensed human vaccines or antivirals for treatment of or prevention from infection with encephalitic alphaviruses. Because epidemics are sporadic and unpredictable, and endemic disease is common but rarely diagnosed, it is difficult to identify all populations requiring vaccination; thus, an effective post-exposure treatment method is needed to interrupt ongoing outbreaks. To address this public health need, we have continued development of ML336 to deliver a molecule with prophylactic and therapeutic potential that could be relevant for use in natural epidemics or deliberate release scenario for Venezuelan equine encephalitis virus (VEEV). We report findings from in vitro assessments of four analogs of ML336, and in vivo screening of three of these new derivatives, BDGR-4, BDGR-69 and BDGR-70. The optimal dosing for maximal protection was observed at 12.5â¯mg/kg/day, twice daily for 8 days. BDGR-4 was tested further for prophylactic and therapeutic efficacy in mice challenged with VEEV Trinidad Donkey (TrD). Mice challenged with VEEV TrD showed 100% and 90% protection from lethal disease when treated at 24 and 48â¯h post-infection, respectively. We also measured 90% protection for BDGR-4 in mice challenged with Eastern equine encephalitis virus. In additional assessments of BDGR-4 in mice alone, we observed no appreciable toxicity as evaluated by clinical chemistry indicators up to a dose of 25â¯mg/kg/day over 4 days. In these same mice, we observed no induction of interferon. Lastly, the resistance of VEEV to BDGR-4 was evaluated by next-generation sequencing which revealed specific mutations in nsP4, the viral polymerase.
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Benzamidas , Benzamidinas , Farmacorresistência Viral/genética , Vírus da Encefalite Equina do Leste/efeitos dos fármacos , Vírus da Encefalite Equina Venezuelana/efeitos dos fármacos , Piperazinas , Animais , Antivirais/síntese química , Antivirais/farmacologia , Benzamidas/síntese química , Benzamidas/farmacologia , Benzamidinas/síntese química , Benzamidinas/farmacologia , Linhagem Celular , Encefalomielite Equina do Leste/tratamento farmacológico , Encefalomielite Equina do Leste/prevenção & controle , Encefalomielite Equina Venezuelana/tratamento farmacológico , Encefalomielite Equina Venezuelana/prevenção & controle , Genes Virais , Camundongos , Mutação , Piperazinas/síntese química , Piperazinas/farmacologiaRESUMO
The New World alphaviruses -Venezuelan, eastern, and western equine encephalitis viruses (VEEV, EEEV, and WEEV respectively) - cause a febrile disease that is often lethal in equines and children and leads to long-term neurological sequelae in survivors. Endemic to the Americas, epizootic outbreaks of the three viruses occur sporadically in the continental United States. All three viruses aerosolize readily, replicate to high titers in cell culture, and have low infectious doses. Additionally, there are no FDA-approved vaccines or therapeutics for human use. To address the therapeutic gap, a high throughput assay utilizing a luciferase reporter virus, TC83-luc, was performed to screen a library of commercially available, FDA-approved drugs for antiviral activity. From a group of twenty compounds found to significantly decrease luminescence, the carcinoma therapeutic sorafenib inhibited replication of VEEV-TC83 and TrD in vitro. Additionally, sorafenib inhibited replication of EEEV and two Old World alphaviruses, Sindbis virus and chikungunya virus, at 8 and 16â¯h post-infection. Sorafenib caused no toxicity in Vero cells, and coupled with a low EC50 value, yielded a selectivity index of >19. Mechanism of actions studies suggest that sorafenib inhibited viral translation through dephosphorylation of several key proteins, including eIF4E and p70S6K, leading to a reduction in viral protein production and overall viral replication.
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Alphavirus/efeitos dos fármacos , Antineoplásicos/farmacologia , Antivirais/farmacologia , Reposicionamento de Medicamentos , Sorafenibe/farmacologia , Replicação Viral/efeitos dos fármacos , Alphavirus/crescimento & desenvolvimento , Animais , Linhagem Celular , Avaliação Pré-Clínica de Medicamentos/métodos , Genes Reporter , Ensaios de Triagem em Larga Escala , Luciferases/análise , Luciferases/genética , Medições Luminescentes , Genética ReversaRESUMO
Venezuelan equine encephalitis virus (VEEV) is a New World alphavirus that is vectored by mosquitos and cycled in rodents. It can cause disease in equines and humans characterized by a febrile illness that may progress into encephalitis. Like the capsid protein of other viruses, VEEV capsid is an abundant structural protein that binds to the viral RNA and interacts with the membrane-bound glycoproteins. It also has protease activity, allowing cleavage of itself from the growing structural polypeptide during translation. However, VEEV capsid protein has additional nonstructural roles within the host cell functioning as the primary virulence factor for VEEV. VEEV capsid inhibits host transcription and blocks nuclear import in mammalian cells, at least partially due to its complexing with the host CRM1 and importin α/ß1 nuclear transport proteins. VEEV capsid also shuttles between the nucleus and cytoplasm and is susceptible to inhibitors of nuclear trafficking, making it a promising antiviral target. Herein, the role of VEEV capsid in viral replication and pathogenesis will be discussed including a comparison to proteins of other alphaviruses.
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Proteínas do Capsídeo/metabolismo , Capsídeo/metabolismo , Vírus da Encefalite Equina Venezuelana/patogenicidade , RNA Viral/metabolismo , Replicação Viral , Transporte Ativo do Núcleo Celular , Animais , Capsídeo/química , Proteínas do Capsídeo/genética , Linhagem Celular , Vírus da Encefalite Equina do Leste , Vírus da Encefalite Equina Venezuelana/genética , Vírus da Encefalite Equina Venezuelana/metabolismo , Vírus da Encefalite Equina do Oeste , Cavalos , Humanos , Carioferinas/genética , Carioferinas/metabolismo , Proteínas de Transporte Nucleocitoplasmático/metabolismo , Receptores Citoplasmáticos e Nucleares/genética , Receptores Citoplasmáticos e Nucleares/metabolismo , Fatores de Virulência , Replicação Viral/genética , Proteína Exportina 1RESUMO
Madariaga virus (MADV), the new species designation for the South American isolates of eastern equine encephalitis virus (EEEV), is genetically divergent and substantially different in ecology and pathogenesis from North American EEEV strains. We isolated and characterized a MADV isolate obtained from a horse in Brazil. Our results support previous phylogenetic studies showing there are three genetically distinct MADV lineages. The MADV isolate from Paraíba State belongs to the South American lineage III and is closely related to Peruvian, Colombian and Venezuelan isolates.
Assuntos
Vírus da Encefalite Equina do Leste , Encefalomielite Equina/veterinária , Doenças dos Cavalos/virologia , Aedes/citologia , Aedes/virologia , Animais , Encéfalo/virologia , Brasil , Células Cultivadas , Vírus da Encefalite Equina do Leste/classificação , Vírus da Encefalite Equina do Leste/genética , Vírus da Encefalite Equina do Leste/isolamento & purificação , Encefalomielite Equina/virologia , Cavalos , Camundongos , FilogeniaRESUMO
Eastern, Venezuelan and western equine encephalitis viruses (EEEV, VEEV, and WEEV) are mosquito-borne viruses that cause substantial disease in humans and other vertebrates. Vaccines are limited and current treatment options have not proven successful. In this report, we vaccinated outbred mice with lipid-antigen-nucleic acid-complexes (LANACs) containing VEEV E1+WEEV E1 antigen and characterized protective efficacy against lethal EEEV, VEEV, and WEEV challenge. Vaccination resulted in complete protection against EEEV, VEEV, and WEEV in CD-1 mice. Measurements of bioluminescence and plaque reduction neutralization tests (PRNTs) indicate that LANAC VEEV E1+WEEV E1 vaccination is sterilizing against VEEV and WEEV challenge; whereas immunity to EEEV is not sterilizing. Passive transfer of rabbit VEEV E1+WEEV E1 immune serum to naive mice extended the mean time to death (MTD) of EEEV challenged mice and provided significant protection from lethal VEEV and WEEV challenge.
Assuntos
Alphavirus/imunologia , Antígenos Virais/imunologia , Reações Cruzadas/imunologia , Vírus da Encefalite Equina Venezuelana/imunologia , Vírus da Encefalite Equina do Oeste/imunologia , Proteínas Virais/imunologia , Infecções por Alphavirus/imunologia , Infecções por Alphavirus/mortalidade , Infecções por Alphavirus/prevenção & controle , Infecções por Alphavirus/virologia , Animais , Anticorpos Antivirais/imunologia , Antígenos Virais/administração & dosagem , Antígenos Virais/genética , Linhagem Celular , Modelos Animais de Doenças , Vírus da Encefalite Equina Venezuelana/genética , Vírus da Encefalite Equina Venezuelana/patogenicidade , Vírus da Encefalite Equina do Oeste/genética , Vírus da Encefalite Equina do Oeste/patogenicidade , Feminino , Expressão Gênica , Genes Reporter , Imunidade Humoral , Imunização , Lipossomos , Camundongos , Ácidos Nucleicos , Homologia de Sequência , Proteínas Virais/administração & dosagem , Proteínas Virais/genética , Virulência/genética , Replicação ViralRESUMO
The encephalitic alphaviruses encode nsP2 protease (nsP2pro), which because of its vital role in virus replication, represents an attractive target for therapeutic intervention. To facilitate the discovery of nsP2 inhibitors we have developed a novel assay for quantitative measurement of nsP2pro activity in a cell-based format. The assay is based on a substrate fusion protein consisting of eGFP and Gaussia luciferase (Gluc) linked together by a small peptide containing a VEEV nsp2pro cleavage sequence. The expression of the substrate protein in cells along with recombinant nsP2pro results in cleavage of the substrate protein resulting in extracellular release of free Gluc. The Gluc activity in supernatants corresponds to intracellular nsP2pro-mediated substrate cleavage; thus, providing a simple and convenient way to quantify nsP2pro activity. Here, we demonstrate potential utility of the assay in identification of nsP2pro inhibitors, as well as in investigations related to molecular characterization of nsP2pro.
Assuntos
Técnicas de Cultura de Células , Cisteína Endopeptidases/metabolismo , Vírus da Encefalite Equina Venezuelana/enzimologia , Ensaios Enzimáticos/métodos , Sequência de Aminoácidos , Antivirais/farmacologia , Linhagem Celular , Cisteína Endopeptidases/química , Cisteína Endopeptidases/genética , Descoberta de Drogas/métodos , Vírus da Encefalite Equina Venezuelana/efeitos dos fármacos , Ativação Enzimática/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Expressão Gênica , Ordem dos Genes , Genes Reporter , Vetores Genéticos , Ensaios de Triagem em Larga Escala , Humanos , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismoRESUMO
Most previous studies of interferon-alpha/beta (IFN-α/ß) response antagonism by alphaviruses have focused upon interruption of IFN-α/ß induction and/or receptor signaling cascades. Infection of mice with Venezuelan equine encephalitis alphavirus (VEEV) or Sindbis virus (SINV) induces serum IFN-α/ß, that elicits a systemic antiviral state in uninfected cells successfully controlling SINV but not VEEV replication. Furthermore, VEEV replication is more resistant than that of SINV to a pre-existing antiviral state in vitro. While host macromolecular shutoff is proposed as a major antagonist of IFN-α/ß induction, the underlying mechanisms of alphavirus resistance to a pre-existing antiviral state are not fully defined, nor is the mechanism for the greater resistance of VEEV. Here, we have separated viral transcription and translation shutoff with multiple alphaviruses, identified the viral proteins that induce each activity, and demonstrated that VEEV nonstructural protein 2-induced translation shutoff is likely a critical factor in enhanced antiviral state resistance of this alphavirus.
Assuntos
Resistência à Doença , Vírus da Encefalite Equina Venezuelana/fisiologia , Encefalomielite Equina Venezuelana/genética , Encefalomielite Equina Venezuelana/virologia , Interações Hospedeiro-Patógeno , Biossíntese de Proteínas , Proteínas não Estruturais Virais/metabolismo , Animais , Antivirais/metabolismo , Antivirais/farmacologia , Linhagem Celular , Vírus da Encefalite Equina Venezuelana/efeitos dos fármacos , Encefalomielite Equina Venezuelana/metabolismo , Encefalomielite Equina Venezuelana/mortalidade , Cavalos , Humanos , Interferons/biossíntese , Interferons/farmacologia , Camundongos , Mutação , Fenótipo , RNA Viral , Proteínas não Estruturais Virais/genéticaRESUMO
The State of Pará comprises 26% of Brazilian Amazon region, where a large diversity of arboviruses has been described. This study sought to assess the prevalence and distribution of hemagglutination inhibition (HI) antibodies against antigens of four alphaviruses (Togaviridae: Alphavirus ) from the species: Eastern equine encephalitis (EEEV), Western equine encephalitis (WEEV), Mayaro virus (MAYV), and Mucambo virus (MUCV) in 753 serum samples of horses in Pará State, Brazil. All investigated arboviruses were detected and indicate that horses are susceptible to these alphaviruses, and show evidences of their active circulation in farm animals in the Brazilian Amazon.(AU)
O estado do Pará corresponde a 26% da Amazônia brasileira, onde uma grande diversidade de arbovírus foi descrita. Este estudo procurou avaliar a prevalência e a distribuição de anticorpos inibidores da hemaglutinação (IH) contra antígenos de quatro alfavirus (Togaviridae: Alphavirus ), das espécies: Vírus da encefalite equina do leste (EEEV), Vírus da encefalite equina do oeste (WEEV), Vírus mayaro (MAYV) e Vírus mucambo (MUCV), de 753 amostras de soro de equinos no estado do Pará, Brasil. Todos os arbovirus pesquisados foram detectados, indicando que os equinos são suscetíveis a esses Alphavirus e mostrando evidências de sua circulação ativa em animais de fazenda na Amazônia brasileira.(AU)