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1.
Vet Microbiol ; 241: 108549, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31928698

RESUMO

Nipah virus (NiV) is a re-emerging zoonotic pathogen that causes high mortality in humans and pigs. Oral immunization in free-roaming animals is one of the most practical approaches to prevent NiV pandemics. We previously generated a recombinant rabies viruses (RABV) Evelyn-Rokitnicki-Abelseth (ERA) strain, rERAG333E, which contains a mutation from arginine to glutamic acid at residue 333 of glycoprotein (G333E) and serves as an oral vaccine for dog rabies. In this study, we generated two recombinant RABVs, rERAG333E/NiVG and rERAG333E/NiVF, expressing the NiV Malaysian strain attachment glycoprotein (NiV-G) or fusion glycoprotein (NiV-F) gene based on the rERAG333E vector platform. Both rERAG333E/NiVG and rERAG333E/NiVF displayed growth properties similar to those of rERAG333E and caused marked syncytia formation after co-infection in BSR cell culture. Adult and suckling mice intracerebrally inoculated with the recombinant RABVs showed NiV-G and NiV-F expression did not increase the virulence of rERAG333E. Oral vaccination with rERAG333E/NiVG either singularly or combined with rERAG333E/NiVF induced significant NiV neutralizing antibody against NiV and RABV, and IgG to NiV-G or NiV-F in mice and pigs. rERAG333E/NiVG and rERAG333E/NiVF thus appeared to be suitable candidates for further oral vaccines for potential animal targets in endemic areas of NiV disease and rabies.


Assuntos
Infecções por Henipavirus/prevenção & controle , Vírus Nipah/imunologia , Vacinas Virais/imunologia , Vacinas Virais/normas , Administração Oral , Animais , Animais Lactentes , Ensaio de Imunoadsorção Enzimática , Feminino , Vetores Genéticos , Imunidade Humoral , Imunoglobulina G/sangue , Camundongos , Camundongos Endogâmicos BALB C , Testes de Neutralização , Vírus da Raiva/genética , Vírus da Raiva/crescimento & desenvolvimento , Vírus da Raiva/patogenicidade , Suínos , Vacinas Sintéticas/administração & dosagem , Vacinas Sintéticas/imunologia , Vacinas Sintéticas/normas , Proteínas Virais/genética , Vacinas Virais/administração & dosagem , Virulência , Zoonoses
2.
F1000Res ; 82019.
Artigo em Inglês | MEDLINE | ID: mdl-31656582

RESUMO

Nipah virus (NiV) is a highly lethal zoonotic paramyxovirus that emerged at the end of last century as a human pathogen capable of causing severe acute respiratory infection and encephalitis. Although NiV provokes serious diseases in numerous mammalian species, the infection seems to be asymptomatic in NiV natural hosts, the fruit bats, which provide a continuous virus source for further outbreaks. Consecutive human-to-human transmission has been frequently observed during outbreaks in Bangladesh and India. NiV was shown to interfere with the innate immune response and interferon type I signaling, restraining the anti-viral response and permitting viral spread. Studies of adaptive immunity in infected patients and animal models have suggested an unbalanced immune response during NiV infection. Here, we summarize some of the recent studies of NiV pathogenesis and NiV-induced modulation of both innate and adaptive immune responses, as well as the development of novel prophylactic and therapeutic approaches, necessary to control this highly lethal emerging infection.


Assuntos
Imunidade Adaptativa , Infecções por Henipavirus/imunologia , Imunidade Inata , Vírus Nipah/patogenicidade , Animais , Bangladesh , Humanos , Índia , Vírus Nipah/imunologia
4.
PLoS Negl Trop Dis ; 13(6): e0007454, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31166946

RESUMO

The ability to appropriately mimic human disease is critical for using animal models as a tool for understanding virus pathogenesis. In the case of Nipah virus (NiV), infection of humans appears to occur either through inhalation, contact with or consumption of infected material. In two of these circumstances, respiratory or sinusoidal exposure represents a likely route of infection. In this study, intermediate-size aerosol particles (~7 µm) of NiV-Malaysia were used to mimic potential routes of exposure by focusing viral deposition in the upper respiratory tract. Our previous report showed this route of exposure extended the disease course and a single animal survived the infection. Here, analysis of the peripheral immune response found minimal evidence of systemic inflammation and depletion of B cells during acute disease. However, the animal that survived infection developed an early IgM response with rapid development of neutralizing antibodies that likely afforded protection. The increase in NiV-specific antibodies correlated with an expansion of the B cell population in the survivor. Cell-mediated immunity was not clearly apparent in animals that succumbed during the acute phase of disease. However, CD4+ and CD8+ effector memory cells increased in the survivor with correlating increases in cytokines and chemokines associated with cell-mediated immunity. Interestingly, kinetic changes of the CD4+ and CD8bright T cell populations over the course of acute disease were opposite from animals that succumbed to infection. In addition, increases in NK cells and basophils during convalescence of the surviving animal were also evident, with viral antigen found in NK cells. These data suggest that a systemic inflammatory response and "cytokine storm" are not major contributors to NiV-Malaysia pathogenesis in the AGM model using this exposure route. Further, these data demonstrate that regulation of cell-mediated immunity, in addition to rapid production of NiV specific antibodies, may be critical for surviving NiV infection.


Assuntos
Aerossóis , Infecções por Henipavirus/imunologia , Imunidade Humoral , Exposição por Inalação , Vírus Nipah/imunologia , Animais , Anticorpos Neutralizantes/sangue , Anticorpos Antivirais/sangue , Linfócitos B/imunologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Chlorocebus aethiops , Citocinas/análise , Modelos Animais de Doenças , Feminino , Humanos , Imunidade Celular , Imunoglobulina M/sangue , Células Matadoras Naturais/imunologia , Masculino
5.
PLoS Negl Trop Dis ; 13(6): e0007462, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31170144

RESUMO

Nipah virus (NiV) is a highly pathogenic re-emerging virus that causes outbreaks in South East Asia. Currently, no approved and licensed vaccine or antivirals exist. Here, we investigated the efficacy of ChAdOx1 NiVB, a simian adenovirus-based vaccine encoding NiV glycoprotein (G) Bangladesh, in Syrian hamsters. Prime-only as well as prime-boost vaccination resulted in uniform protection against a lethal challenge with NiV Bangladesh: all animals survived challenge and we were unable to find infectious virus either in oral swabs, lung or brain tissue. Furthermore, no pathological lung damage was observed. A single-dose of ChAdOx1 NiVB also prevented disease and lethality from heterologous challenge with NiV Malaysia. While we were unable to detect infectious virus in swabs or tissue of animals challenged with the heterologous strain, a very limited amount of viral RNA could be found in lung tissue by in situ hybridization. A single dose of ChAdOx1 NiVB also provided partial protection against Hendra virus and passive transfer of antibodies elicited by ChAdOx1 NiVB vaccination partially protected Syrian hamsters against NiV Bangladesh. From these data, we conclude that ChAdOx1 NiVB is a suitable candidate for further NiV vaccine pre-clinical development.


Assuntos
Adenovirus dos Símios/genética , Portadores de Fármacos , Infecções por Henipavirus/prevenção & controle , Vírus Nipah/imunologia , Vacinas Virais/imunologia , Estruturas Animais/virologia , Animais , Modelos Animais de Doenças , Feminino , Infecções por Henipavirus/imunologia , Mesocricetus , Vírus Nipah/genética , Análise de Sobrevida , Vacinas Sintéticas/administração & dosagem , Vacinas Sintéticas/genética , Vacinas Sintéticas/imunologia , Vacinas Virais/administração & dosagem , Vacinas Virais/genética
6.
PLoS Negl Trop Dis ; 13(6): e0007393, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31246966

RESUMO

The 2018 outbreak of Nipah virus in Kerala, India, highlights the need for global surveillance of henipaviruses in bats, which are the reservoir hosts for this and other viruses. Nipah virus, an emerging paramyxovirus in the genus Henipavirus, causes severe disease and stuttering chains of transmission in humans and is considered a potential pandemic threat. In May 2018, an outbreak of Nipah virus began in Kerala, > 1800 km from the sites of previous outbreaks in eastern India in 2001 and 2007. Twenty-three people were infected and 21 people died (16 deaths and 18 cases were laboratory confirmed). Initial surveillance focused on insectivorous bats (Megaderma spasma), whereas follow-up surveys within Kerala found evidence of Nipah virus in fruit bats (Pteropus medius). P. medius is the confirmed host in Bangladesh and is now a confirmed host in India. However, other bat species may also serve as reservoir hosts of henipaviruses. To inform surveillance of Nipah virus in bats, we reviewed and analyzed the published records of Nipah virus surveillance globally. We applied a trait-based machine learning approach to a subset of species that occur in Asia, Australia, and Oceana. In addition to seven species in Kerala that were previously identified as Nipah virus seropositive, we identified at least four bat species that, on the basis of trait similarity with known Nipah virus-seropositive species, have a relatively high likelihood of exposure to Nipah or Nipah-like viruses in India. These machine-learning approaches provide the first step in the sequence of studies required to assess the risk of Nipah virus spillover in India. Nipah virus surveillance not only within Kerala but also elsewhere in India would benefit from a research pipeline that included surveys of known and predicted reservoirs for serological evidence of past infection with Nipah virus (or cross reacting henipaviruses). Serosurveys should then be followed by longitudinal spatial and temporal studies to detect shedding and isolate virus from species with evidence of infection. Ecological studies will then be required to understand the dynamics governing prevalence and shedding in bats and the contacts that could pose a risk to public health.


Assuntos
Quirópteros/virologia , Controle de Doenças Transmissíveis/organização & administração , Transmissão de Doença Infecciosa , Monitoramento Epidemiológico , Infecções por Henipavirus/epidemiologia , Vírus Nipah/crescimento & desenvolvimento , Zoonoses/epidemiologia , Animais , Reservatórios de Doenças/virologia , Infecções por Henipavirus/transmissão , Infecções por Henipavirus/veterinária , Humanos , Índia/epidemiologia , Vírus Nipah/imunologia , Vírus Nipah/isolamento & purificação , Medição de Risco , Estudos Soroepidemiológicos , Zoonoses/transmissão
8.
Biochem Biophys Res Commun ; 513(4): 781-786, 2019 06 11.
Artigo em Inglês | MEDLINE | ID: mdl-30995971

RESUMO

Nipah virus (NiV) is a recently emerged paramyxovirus that causes acute respiratory illness and fatal encephalitis in a broad spectrum of vertebrates, including humans. Due to its high pathogenicity and mortality rates, NiV requires handling in biosafety level-4 (BSL-4) containment facilities and no effective vaccines or therapeutic agents are currently available. Since current diagnostic tests for detecting serum neutralizing antibodies against NiV mainly employ live viruses, establishment of more safe and robust alternative diagnostic methods is an essential medical requirement. Here, we have developed a pseudotyped NiV and closely related Hendra virus (HeV) expressing envelope attachment (G) and fusion (F) glycoproteins using the Moloney murine leukemia virus (MuLV) packaging system. We additionally generated polyclonal antibodies (pAbs) against NiV-G and HeV-G and assessed their neutralizing activities for potential utilization in the pseudovirus-based neutralization assay and further application in the serum diagnostic test. To enhance the specificity of neutralizing antibody and sensitivity of the serological diagnostic test, monoclonal antibodies (mAbs) against NiV-G were generated, and among which four out of six mAb clones showed significant reactivity. Specifically, the 7G9 clone displayed the highest sensitivity. The selected mAb clones showed no cross-reactivity with HeV-G and efficient neutralizing activities against pseudotyped NiV. These results validate the safety and specificity of neutralization assays against NiV and HeV and present a useful tool to design effective vaccines and serological diagnosis.


Assuntos
Anticorpos Monoclonais/imunologia , Especificidade de Anticorpos/imunologia , Testes de Neutralização/métodos , Vírus Nipah/imunologia , Sequência de Aminoácidos , Animais , Anticorpos Monoclonais/química , Antígenos/imunologia , Linhagem Celular , Feminino , Glicoproteínas , Vírus Hendra , Camundongos Endogâmicos BALB C , Proteínas do Envelope Viral
9.
Vet Q ; 39(1): 26-55, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31006350

RESUMO

Nipah (Nee-pa) viral disease is a zoonotic infection caused by Nipah virus (NiV), a paramyxovirus belonging to the genus Henipavirus of the family Paramyxoviridae. It is a biosafety level-4 pathogen, which is transmitted by specific types of fruit bats, mainly Pteropus spp. which are natural reservoir host. The disease was reported for the first time from the Kampung Sungai Nipah village of Malaysia in 1998. Human-to-human transmission also occurs. Outbreaks have been reported also from other countries in South and Southeast Asia. Phylogenetic analysis affirmed the circulation of two major clades of NiV as based on currently available complete N and G gene sequences. NiV isolates from Malaysia and Cambodia clustered together in NiV-MY clade, whereas isolates from Bangladesh and India clusterered within NiV-BD clade. NiV isolates from Thailand harboured mixed population of sequences. In humans, the virus is responsible for causing rapidly progressing severe illness which might be characterized by severe respiratory illness and/or deadly encephalitis. In pigs below six months of age, respiratory illness along with nervous symptoms may develop. Different types of enzyme-linked immunosorbent assays along with molecular methods based on polymerase chain reaction have been developed for diagnostic purposes. Due to the expensive nature of the antibody drugs, identification of broad-spectrum antivirals is essential along with focusing on small interfering RNAs (siRNAs). High pathogenicity of NiV in humans, and lack of vaccines or therapeutics to counter this disease have attracted attention of researchers worldwide for developing effective NiV vaccine and treatment regimens.


Assuntos
Infecções por Henipavirus/veterinária , Vírus Nipah/imunologia , Vacinas Virais , Zoonoses , Animais , Doenças do Gato/epidemiologia , Doenças do Gato/prevenção & controle , Doenças do Gato/virologia , Gatos , Doenças do Cão/epidemiologia , Doenças do Cão/prevenção & controle , Doenças do Cão/virologia , Cães , Infecções por Henipavirus/epidemiologia , Infecções por Henipavirus/prevenção & controle , Infecções por Henipavirus/virologia , Humanos , Vírus Nipah/classificação , Vacinas Virais/administração & dosagem , Vacinas Virais/análise , Vacinas Virais/uso terapêutico , Zoonoses/epidemiologia , Zoonoses/prevenção & controle , Zoonoses/virologia
10.
Microbes Infect ; 21(7): 278-286, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30817995

RESUMO

Nipah virus is an emerging zoonotic paramyxovirus that causes severe and often fatal respiratory and neurological disease in humans. The virus was first discovered after an outbreak of encephalitis in pig farmers in Malaysia and Singapore with subsequent outbreaks in Bangladesh or India occurring almost annually. Due to the highly pathogenic nature of NiV, its pandemic potential, and the lack of licensed vaccines or therapeutics, there is a requirement for research and development into highly sensitive and specific diagnostic tools as well as antivirals and vaccines to help prevent and control future outbreak situations.


Assuntos
Infecções por Henipavirus/diagnóstico , Infecções por Henipavirus/prevenção & controle , Vírus Nipah/imunologia , Vírus Nipah/isolamento & purificação , Animais , Antivirais/uso terapêutico , Técnicas de Laboratório Clínico , Surtos de Doenças/prevenção & controle , Infecções por Henipavirus/epidemiologia , Infecções por Henipavirus/terapia , Humanos , Modelos Biológicos , Vírus Nipah/genética , Vírus Nipah/patogenicidade , Vacinas Virais/imunologia , Zoonoses/epidemiologia , Zoonoses/transmissão
11.
Emerg Microbes Infect ; 8(1): 272-281, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30866781

RESUMO

Because of its high infectivity in humans and the lack of effective vaccines, Nipah virus is classified as a category C agent and handling has to be performed under biosafety level 4 conditions in non-endemic countries, which has hindered the development of vaccines. Based on a highly efficient pseudovirus production system using a modified HIV backbone vector, a pseudovirus-based mouse model has been developed for evaluating the efficacy of Nipah vaccines in biosafety level 2 facilities. For the first time, the correlates of protection have been identified in a mouse model. The limited levels of neutralizing antibodies against immunogens fusion protein (F), glycoprotein (G), and combination of F and G (FG) were found to be 148, 275, and 115, respectively, in passive immunization. Relatively lower limited levels of protection of 52, and 170 were observed for immunogens F, and G, respectively, in an active immunization model. Although the minimal levels for protection of neutralizing antibody in passive immunization were slightly higher than those in active immunization, neutralizing antibody played a key role in protection against Nipah virus infection. The immunogens F and G provided similar protection, and the combination of these immunogens did not provide better outcomes. Either immunogen F or G would provide sufficient protection for Nipah vaccine. The Nipah pseudovirus mouse model, which does not involve highly pathogenic virus, has the potential to greatly facilitate the standardization and implementation of an assay to propel the development of NiV vaccines.


Assuntos
Infecções por Henipavirus/prevenção & controle , Vírus Nipah/imunologia , Proteínas do Envelope Viral/imunologia , Vacinas Virais/administração & dosagem , Animais , Anticorpos Neutralizantes/metabolismo , Contenção de Riscos Biológicos , Cães , Infecções por Henipavirus/imunologia , Humanos , Células Madin Darby de Rim Canino , Camundongos , Vacinas Virais/imunologia
12.
Curr Top Med Chem ; 18(26): 2202-2208, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30417788

RESUMO

We briefly review the situations arising out of epidemics that erupt rather suddenly, threatening life and livelihoods of humans. Ebola, Zika and the Nipah virus outbreaks are recent examples where the viral epidemics have led to considerably high degree of fatalities or debilitating consequences. The problems are accentuated by a lack of drugs or vaccines effective against the new and emergent viruses, and the inordinate amount of temporal and financial resources that are required to combat the novel pathogens. Progress in computational, biological and informational sciences have made it possible to consider design of synthetic vaccines that can be rapidly developed and deployed to help stem the damages. In this review, we consider the pros and cons of this new paradigm and suggest a new system where the manufacturing process can be decentralized to provide more targeted vaccines to meet the urgent needs of protection in case of a rampaging epidemic.


Assuntos
Doença pelo Vírus Ebola/epidemiologia , Infecções por Henipavirus/epidemiologia , Peptídeos/imunologia , Vacinas Virais/imunologia , Infecção por Zika virus/epidemiologia , Animais , Doença pelo Vírus Ebola/imunologia , Doença pelo Vírus Ebola/prevenção & controle , Doença pelo Vírus Ebola/virologia , Infecções por Henipavirus/imunologia , Infecções por Henipavirus/prevenção & controle , Humanos , Vírus Nipah/imunologia , Infecção por Zika virus/imunologia , Infecção por Zika virus/prevenção & controle , Infecção por Zika virus/virologia
13.
PLoS Negl Trop Dis ; 12(3): e0006343, 2018 03.
Artigo em Inglês | MEDLINE | ID: mdl-29538374

RESUMO

Henipavirus infection causes severe respiratory and neurological disease in humans that can be fatal. To characterize the pathogenic mechanisms of henipavirus infection in vivo, we performed experimental infections in ferrets followed by genome-wide gene expression analysis of lung and brain tissues. The Hendra, Nipah-Bangladesh, and Nipah-Malaysia strains caused severe respiratory and neurological disease with animals succumbing around 7 days post infection. Despite the presence of abundant viral shedding, animal-to-animal transmission did not occur. The host gene expression profiles of the lung tissue showed early activation of interferon responses and subsequent expression of inflammation-related genes that coincided with the clinical deterioration. Additionally, the lung tissue showed unchanged levels of lymphocyte markers and progressive downregulation of cell cycle genes and extracellular matrix components. Infection in the brain resulted in a limited breadth of the host responses, which is in accordance with the immunoprivileged status of this organ. Finally, we propose a model of the pathogenic mechanisms of henipavirus infection that integrates multiple components of the host responses.


Assuntos
Infecções por Henipavirus/genética , Infecções por Henipavirus/imunologia , Henipavirus/fisiologia , Interações Hospedeiro-Patógeno , Transcriptoma , Animais , Encéfalo/metabolismo , Encéfalo/virologia , Ciclo Celular , Modelos Animais de Doenças , Matriz Extracelular/genética , Furões/virologia , Vírus Hendra/imunologia , Vírus Hendra/patogenicidade , Henipavirus/genética , Infecções por Henipavirus/virologia , Humanos , Inflamação , Interferons/genética , Pulmão/metabolismo , Pulmão/virologia , Vírus Nipah/imunologia , Vírus Nipah/patogenicidade , Eliminação de Partículas Virais
14.
Curr Top Microbiol Immunol ; 419: 191-213, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-28674944

RESUMO

Hendra and Nipah viruses (family Paramyxoviridae, genus Henipavirus) are zoonotic RNA viruses that cause lethal disease in humans and are designated as Biosafety Level 4 (BSL4) agents. Moreover, henipaviruses belong to the same group of viruses that cause disease more commonly in humans such as measles, mumps and respiratory syncytial virus. Due to the relatively recent emergence of the henipaviruses and the practical constraints of performing functional genomics studies at high levels of containment, our understanding of the henipavirus infection cycle is incomplete. In this chapter we describe recent loss-of-function (i.e. RNAi) functional genomics screens that shed light on the henipavirus-host interface at a genome-wide level. Further to this, we cross-reference RNAi results with studies probing host proteins targeted by henipavirus proteins, such as nuclear proteins and immune modulators. These functional genomics studies join a growing body of evidence demonstrating that nuclear and nucleolar host proteins play a crucial role in henipavirus infection. Furthermore these studies will underpin future efforts to define the role of nucleolar host-virus interactions in infection and disease.


Assuntos
Genômica , Vírus Hendra/imunologia , Infecções por Henipavirus/genética , Infecções por Henipavirus/imunologia , Interações Hospedeiro-Patógeno , MicroRNAs/metabolismo , Vírus Nipah/imunologia , Proteínas Nucleares/metabolismo , Infecções por Henipavirus/metabolismo , Infecções por Henipavirus/virologia , Humanos , MicroRNAs/genética , Proteínas Nucleares/genética
15.
J Gen Virol ; 98(10): 2447-2453, 2017 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-28984239

RESUMO

Highly pathogenic Nipah virus (NiV) generally causes severe encephalitis in humans. Respiratory symptoms are infrequently observed, likely reflecting variations in infection kinetics in human airways. Supporting this idea, we recently identified individual differences in NiV replication kinetics in cultured airway epithelia from different human donors. As type III interferons (IFN-λ) represent major players in the defence mechanism against viral infection of the respiratory mucosa, we studied IFN-λ induction and antiviral activity in NiV-infected primary differentiated human bronchial epithelial cells (HBEpCs) cultured under air-liquid interface conditions. Our studies revealed that IFN-λ was upregulated in airway epithelia upon NiV infection. We also show that IFN-λ pretreatment efficiently inhibited NiV replication. Interestingly, the antiviral activity of IFN-λ varied in HBEpCs from two different donors. Increased sensitivity to IFN-λ was associated with higher expression levels of IFN-λ receptors, enhanced phosphorylation of STAT1, as well as enhanced induction of interferon-stimulated gene expression. These findings suggest that individual variations in IFN-λ receptor expression affecting IFN responsiveness can play a functional role for NiV replication kinetics in human respiratory epithelial cells of different donors.


Assuntos
Brônquios/imunologia , Células Epiteliais/imunologia , Interferons/biossíntese , Interferons/farmacologia , Vírus Nipah/imunologia , Receptores de Interferon/biossíntese , Mucosa Respiratória/imunologia , Animais , Brônquios/citologia , Brônquios/virologia , Linhagem Celular , Chlorocebus aethiops , Células Epiteliais/virologia , Humanos , Fosforilação , Mucosa Respiratória/citologia , Mucosa Respiratória/virologia , Fator de Transcrição STAT1/metabolismo , Células Vero , Replicação Viral/efeitos dos fármacos
16.
J Virol ; 91(6)2017 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-28077641

RESUMO

Recombinant vesicular stomatitis virus (VSV)-based chimeric viruses that include genes from other viruses show promise as vaccines and oncolytic viruses. However, the critical safety concern is the neurotropic nature conveyed by the VSV glycoprotein. VSVs that include the VSV glycoprotein (G) gene, even in most recombinant attenuated strains, can still show substantial adverse or lethal actions in the brain. Here, we test 4 chimeric viruses in the brain, including those in which glycoprotein genes from Nipah, chikungunya (CHIKV), and influenza H5N1 viruses were substituted for the VSV glycoprotein gene. We also test a virus-like vesicle (VLV) in which the VSV glycoprotein gene is expressed from a replicon encoding the nonstructural proteins of Semliki Forest virus. VSVΔG-CHIKV, VSVΔG-H5N1, and VLV were all safe in the adult mouse brain, as were VSVΔG viruses expressing either the Nipah F or G glycoprotein. In contrast, a complementing pair of VSVΔG viruses expressing Nipah G and F glycoproteins were lethal within the brain within a surprisingly short time frame of 2 days. Intranasal inoculation in postnatal day 14 mice with VSVΔG-CHIKV or VLV evoked no adverse response, whereas VSVΔG-H5N1 by this route was lethal in most mice. A key immune mechanism underlying the safety of VSVΔG-CHIKV, VSVΔG-H5N1, and VLV in the adult brain was the type I interferon response; all three viruses were lethal in the brains of adult mice lacking the interferon receptor, suggesting that the viruses can infect and replicate and spread in brain cells if not blocked by interferon-stimulated genes within the brain.IMPORTANCE Vesicular stomatitis virus (VSV) shows considerable promise both as a vaccine vector and as an oncolytic virus. The greatest limitation of VSV is that it is highly neurotropic and can be lethal within the brain. The neurotropism can be mostly attributed to the VSV G glycoprotein. Here, we test 4 chimeric viruses of VSV with glycoprotein genes from Nipah, chikungunya, and influenza viruses and nonstructural genes from Semliki Forest virus. Two of the four, VSVΔG-CHIKV and VLV, show substantially attenuated neurotropism and were safe in the healthy adult mouse brain. VSVΔG-H5N1 was safe in the adult brain but lethal in the younger brain. VSVΔG Nipah F+G was even more neurotropic than wild-type VSV, evoking a rapid lethal response in the adult brain. These results suggest that while chimeric VSVs show promise, each must be tested with both intranasal and intracranial administration to ensure the absence of lethal neurotropism.


Assuntos
Encéfalo/patologia , Vesiculovirus/patogenicidade , Vacinas Virais/efeitos adversos , Animais , Vírus Chikungunya/genética , Vírus Chikungunya/imunologia , Interferon Tipo I/metabolismo , Camundongos , Vírus Nipah/genética , Vírus Nipah/imunologia , Orthomyxoviridae/genética , Orthomyxoviridae/imunologia , Vírus da Floresta de Semliki/genética , Vírus da Floresta de Semliki/imunologia , Análise de Sobrevida , Vacinas Atenuadas/efeitos adversos , Vacinas Atenuadas/genética , Vacinas Atenuadas/imunologia , Vacinas Sintéticas/efeitos adversos , Vacinas Sintéticas/genética , Vacinas Sintéticas/imunologia , Vesiculovirus/genética , Vesiculovirus/imunologia , Vacinas Virais/genética , Vacinas Virais/imunologia
17.
Jpn J Infect Dis ; 70(1): 26-31, 2017 Jan 24.
Artigo em Inglês | MEDLINE | ID: mdl-27169942

RESUMO

The genes for Nipah virus (NiV) proteins were amplified from viral RNA, cloned into the plasmid pTriEx-3 Hygro, expressed, and purified using immobilized metal affinity chromatography. The recombinant N, F, and G NiV proteins (rNiV-N, rNiV-F, and rNiV-G), were successfully expressed in Escherichia coli and purified with a yield of 4, 16, and 4 mg/L, respectively. All 3 recombinant viral proteins reacted with all 19 samples of NiV-positive human sera. The rNiV-N and rNiV-G proteins were the most immunogenic. The recombinant viral proteins did not react with any of the 12 NiV-negative sera. However, serum from a patient with a late-onset relapsing NiV infection complication was found to be primarily reactive to rNiV-G only. Additionally, there is a distinctive variation in the profile of antigen-reactive bands between the sample from a case of relapsing NiV encephalitis and that of acute NiV infection. The overall findings of this study suggest that the recombinant viral proteins have the potential to be developed further for use in the detection of NiV infection, and continuous biosurveillance of NiV infection in resource-limited settings.


Assuntos
Anticorpos Antivirais/sangue , Antígenos Virais/imunologia , Vírus Nipah/imunologia , Proteínas Recombinantes/imunologia , Proteínas Estruturais Virais/imunologia , Antígenos Virais/genética , Antígenos Virais/isolamento & purificação , Clonagem Molecular , Escherichia coli/genética , Escherichia coli/metabolismo , Expressão Gênica , Vetores Genéticos , Humanos , Plasmídeos , Proteínas Recombinantes/genética , Proteínas Recombinantes/isolamento & purificação , Soro/imunologia , Proteínas Estruturais Virais/genética , Proteínas Estruturais Virais/isolamento & purificação
18.
PLoS Pathog ; 12(9): e1005880, 2016 09.
Artigo em Inglês | MEDLINE | ID: mdl-27622505

RESUMO

For efficient replication, viruses have developed mechanisms to evade innate immune responses, including the antiviral type-I interferon (IFN-I) system. Nipah virus (NiV), a highly pathogenic member of the Paramyxoviridae family (genus Henipavirus), is known to encode for four P gene-derived viral proteins (P/C/W/V) with IFN-I antagonist functions. Here we report that NiV matrix protein (NiV-M), which is important for virus assembly and budding, can also inhibit IFN-I responses. IFN-I production requires activation of multiple signaling components including the IκB kinase epsilon (IKKε). We previously showed that the E3-ubiquitin ligase TRIM6 catalyzes the synthesis of unanchored K48-linked polyubiquitin chains, which are not covalently attached to any protein, and activate IKKε for induction of IFN-I mediated antiviral responses. Using co-immunoprecipitation assays and confocal microscopy we show here that the NiV-M protein interacts with TRIM6 and promotes TRIM6 degradation. Consequently, NiV-M expression results in reduced levels of unanchored K48-linked polyubiquitin chains associated with IKKε leading to impaired IKKε oligomerization, IKKε autophosphorylation and reduced IFN-mediated responses. This IFN antagonist function of NiV-M requires a conserved lysine residue (K258) in the bipartite nuclear localization signal that is found in divergent henipaviruses. Consistent with this, the matrix proteins of Ghana, Hendra and Cedar viruses were also able to inhibit IFNß induction. Live NiV infection, but not a recombinant NiV lacking the M protein, reduced the levels of endogenous TRIM6 protein expression. To our knowledge, matrix proteins of paramyxoviruses have never been reported to be involved in innate immune antagonism. We report here a novel mechanism of viral innate immune evasion by targeting TRIM6, IKKε and unanchored polyubiquitin chains. These findings expand the universe of viral IFN antagonism strategies and provide a new potential target for development of therapeutic interventions against NiV infections.


Assuntos
Infecções por Henipavirus/imunologia , Quinase I-kappa B/imunologia , Evasão da Resposta Imune , Interferon Tipo I/imunologia , Vírus Nipah/imunologia , Proteínas com Motivo Tripartido/imunologia , Ubiquitina-Proteína Ligases/imunologia , Proteínas Virais/imunologia , Células A549 , Animais , Chlorocebus aethiops , Células HeLa , Infecções por Henipavirus/genética , Humanos , Quinase I-kappa B/genética , Imunidade Inata , Interferon Tipo I/genética , Vírus Nipah/genética , Poliubiquitina/genética , Poliubiquitina/imunologia , Multimerização Proteica/genética , Multimerização Proteica/imunologia , Proteínas com Motivo Tripartido/genética , Ubiquitina-Proteína Ligases/genética , Ubiquitinação/genética , Ubiquitinação/imunologia , Células Vero , Proteínas Virais/genética
19.
Biotechnol Prog ; 32(1): 171-7, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-26519022

RESUMO

Nipah virus (NiV) causes fatal respiratory illness and encephalitis in humans and animals. The matrix (M) protein of NiV plays an important role in the viral assembly and budding process. Thus, an access to the NiV M protein is vital to the design of viral antigens as diagnostic reagents. In this study, recombinant DNA technology was successfully adopted in the cloning and expression of NiV M protein. A recombinant expression cassette (baculovirus expression vector) was used to encode an N-terminally His-tagged NiV M protein in insect cells. A time-course study demonstrated that the highest yield of recombinant M protein (400-500 µg) was expressed from 107 infected cells 3 days after infection. A single-step purification method based on metal ion affinity chromatography was established to purify the NiV M protein, which successfully yielded a purity level of 95.67% and a purification factor of 3.39. The Western blotting and enzyme-linked immunosorbent assay (ELISA) showed that the purified recombinant M protein (48 kDa) was antigenic and reacted strongly with the serum of a NiV infected pig.


Assuntos
Vírus Nipah/química , Proteínas Recombinantes/biossíntese , Proteínas da Matriz Viral/biossíntese , Animais , Baculoviridae/genética , Ensaio de Imunoadsorção Enzimática , Vetores Genéticos , Humanos , Insetos/citologia , Vírus Nipah/imunologia , Proteínas Recombinantes/genética , Proteínas Recombinantes/isolamento & purificação , Suínos/imunologia , Suínos/virologia , Proteínas da Matriz Viral/genética , Proteínas da Matriz Viral/isolamento & purificação
20.
J Virol ; 90(5): 2706-9, 2015 Dec 16.
Artigo em Inglês | MEDLINE | ID: mdl-26676791

RESUMO

Nipah virus (NiV) is a highly pathogenic paramyxovirus that causes pronounced infection of brain endothelia and central nervous system (CNS) inflammation. Using primary porcine brain microvascular endothelial cells, we showed that upregulation of E-selectin precedes cytokine induction and is induced not only by infectious NiV but also by NiV-glycoprotein-containing virus-like particles. This demonstrates that very early events in NiV brain endothelial infection do not depend on NiV replication but can be triggered by the NiV glycoproteins alone.


Assuntos
Citocinas/biossíntese , Células Endoteliais/imunologia , Glicoproteínas/imunologia , Interações Hospedeiro-Patógeno , Vírus Nipah/imunologia , Proteínas Virais/imunologia , Virossomos/imunologia , Animais , Células Cultivadas , Selectina E/biossíntese , Células Endoteliais/efeitos dos fármacos , Suínos , Regulação para Cima
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