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1.
BMC Infect Dis ; 19(1): 986, 2019 Nov 21.
Artigo em Inglês | MEDLINE | ID: mdl-31752731

RESUMO

BACKGROUND: Zika virus (ZIKV) infection gained public health concern after the 2015 outbreak in Brazil, when microcephaly rates increased in babies born from infected mothers. It was demonstrated that ZIKV causes a congenital Zika virus syndrome, including various alterations in the development of the central nervous system. Although the infection of cells from the nervous system has been well documented, less is known in respect of ZIKV ability to infect immune cells. Herein, we investigated if peripheral blood mononuclear cells (PBMCs), freshly-isolated from healthy donors, could be infected by ZIKV. METHODS: PBMCs from healthy donors were isolated and cultured in medium with ZIKV strain Rio-U1 (MOI = 0.1). Infection was analyzed by RT-qPCR and flow cytometry. RESULTS: We detected the ZIKV RNA in PBMCs from all donors by RT-qPCR analysis. The detection of viral antigens by flow cytometry revealed that PBMC from more than 50% the donors were infected by ZIKV, with CD3+CD4+ T cells, CD3-CD19+ B cells and CD3+CD8+ T cells being, respectively, the most frequently infected subpopulations, followed by CD14+ monocytes. Additionally, we observed high variability in PBMC infection rates among different donors, either by numbers or type infected cells. CONCLUSIONS: These findings raise the hypothesis that PBMCs can act as a reservoir of the virus, which may facilitate viral dissemination to different organs, including immune-privileged sites.


Assuntos
Leucócitos Mononucleares/virologia , Infecção por Zika virus/virologia , Zika virus/isolamento & purificação , Antígenos CD19/genética , Antígenos CD19/imunologia , Linfócitos B/imunologia , Brasil , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/virologia , Células Cultivadas , Humanos , Leucócitos Mononucleares/imunologia , Monócitos/imunologia , Monócitos/virologia , Reação em Cadeia da Polimerase em Tempo Real , Zika virus/genética , Zika virus/fisiologia , Infecção por Zika virus/diagnóstico , Infecção por Zika virus/genética , Infecção por Zika virus/imunologia
2.
Emerg Microbes Infect ; 8(1): 1668-1678, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31735122

RESUMO

Since its emergence in Yap Island in 2007, Zika virus (ZIKV) has affected all continents except Europe. Despite the hundreds of cases imported to European countries from ZIKV-infested regions, no local cases have been reported in localities where the ZIKV-competent mosquito Aedes albopictus is well established. Here we analysed the vector competence of European Aedes (aegypti and albopictus) mosquitoes to different genotypes of ZIKV. We demonstrate that Ae. albopictus from France was less susceptible to the Asian ZIKV than to the African ZIKV. Critically we show that effective crossing of anatomical barriers (midgut and salivary glands) after an infectious blood meal depends on a viral load threshold to trigger: (i) viral dissemination from the midgut to infect mosquito internal organs and (ii) viral transmission from the saliva to infect a vertebrate host. A viral load in body ≥4800 viral copies triggered dissemination and ≥12,000 viral copies set out transmission. Only 27.3% and 18.2% of Ae. albopictus Montpellier mosquitoes meet respectively these two criteria. Collectively, these compelling results stress the poor ability of Ae. albopictus to sustain a local transmission of ZIKV in Europe and provide a promising tool to evaluate the risk of ZIKV transmission in future outbreaks.


Assuntos
Aedes/fisiologia , Mosquitos Vetores/fisiologia , Infecção por Zika virus/transmissão , Zika virus/fisiologia , Aedes/genética , Aedes/virologia , Animais , Europa (Continente) , Feminino , Humanos , Mosquitos Vetores/genética , Mosquitos Vetores/virologia , Carga Viral , Zika virus/genética , Infecção por Zika virus/virologia
3.
Nat Commun ; 10(1): 4430, 2019 09 27.
Artigo em Inglês | MEDLINE | ID: mdl-31562326

RESUMO

Zika virus (ZIKV) invades and persists in the central nervous system (CNS), causing severe neurological diseases. However the virus journey, from the bloodstream to tissues through a mature endothelium, remains unclear. Here, we show that ZIKV-infected monocytes represent suitable carriers for viral dissemination to the CNS using human primary monocytes, cerebral organoids derived from embryonic stem cells, organotypic mouse cerebellar slices, a xenotypic human-zebrafish model, and human fetus brain samples. We find that ZIKV-exposed monocytes exhibit higher expression of adhesion molecules, and higher abilities to attach onto the vessel wall and transmigrate across endothelia. This phenotype is associated to enhanced monocyte-mediated ZIKV dissemination to neural cells. Together, our data show that ZIKV manipulates the monocyte adhesive properties and enhances monocyte transmigration and viral dissemination to neural cells. Monocyte transmigration may represent an important mechanism required for viral tissue invasion and persistence that could be specifically targeted for therapeutic intervention.


Assuntos
Moléculas de Adesão Celular/metabolismo , Monócitos/metabolismo , Monócitos/virologia , Neurônios/metabolismo , Migração Transendotelial e Transepitelial/fisiologia , Infecção por Zika virus/metabolismo , Zika virus/fisiologia , Zika virus/patogenicidade , Animais , Adesão Celular/fisiologia , Sobrevivência Celular , Sistema Nervoso Central/metabolismo , Sistema Nervoso Central/patologia , Sistema Nervoso Central/virologia , Cerebelo/patologia , Cerebelo/virologia , Modelos Animais de Doenças , Células-Tronco Embrionárias , Endotélio/virologia , Feminino , Humanos , Monócitos/patologia , Neurônios/patologia , Neurônios/virologia , Organoides/metabolismo , Organoides/patologia , Peixe-Zebra , Infecção por Zika virus/patologia , Infecção por Zika virus/virologia
4.
Acta Virol ; 63(3): 316-321, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31507198

RESUMO

The recent Zika virus (ZIKV) outbreaks and rapid spread in tropical Latin America since introduction to Brazil in 2014, and now appearing cases in the USA, are alarming. World Health Organization (WHO) has considered transmission of ZIKV, a serious public health problem because of the increasing number of outbreaks. There are currently no drugs approved for the treatment of ZIKV infection. Discovery of safe and effective drugs are hampered by the risk in treating pregnant woman and toxicity to the fetus. Sweet basil, known as Ocimum basilicum in the scientific community, is a very well-known medicinal herb. Numerous studies have documented its beneficial activity against a great variety of human pathogens ranging from bacteria and virus to fungus and protozoans. Although, basil extracts and oils have been tested successfully against other viruses, its application to tackle ZIKV infection has not been exploited at all. In this study, we report for the first time that highly diluted ethanol extracts prepared from basil leaves can effectively inhibit ZIKV replication in Vero E6 cells with a half maximal inhibitory concentration (IC50) value of 1:134. The diluted extract as well as the amount of ethanol that goes into its preparation have been found to be completely non-toxic to the above mentioned cell line. The extract seems to inhibit the virus at the step of attachment and entry into the host cell. The specific inhibition of ZIKV observed using the basil leaf extract suggests a new alternative mode of treatment against flavivirus. Keywords: Zika virus; basil extract; antiviral.


Assuntos
Ocimum basilicum , Extratos Vegetais , Internalização do Vírus , Infecção por Zika virus , Animais , Sobrevivência Celular/efeitos dos fármacos , Etanol/química , Ocimum basilicum/química , Extratos Vegetais/farmacologia , Folhas de Planta/química , Plantas Medicinais/química , Células Vero , Internalização do Vírus/efeitos dos fármacos , Zika virus/efeitos dos fármacos , Zika virus/fisiologia
5.
Nat Commun ; 10(1): 4344, 2019 09 25.
Artigo em Inglês | MEDLINE | ID: mdl-31554802

RESUMO

Innate immune responses to Zika virus (ZIKV) are dampened in the lower female reproductive tract (LFRT) compared to other tissues, but the mechanism that underlies this vulnerability is poorly understood. Using tissues from uninfected and vaginally ZIKV-infected macaques and mice, we show that low basal expression of RNA-sensing pattern recognition receptors (PRRs), or their co-receptors, in the LFRT contributes to high viral replication in this tissue. In the LFRT, ZIKV sensing provides limited protection against viral replication, and the sensors are also minimally induced after vaginal infection. While IFNα/ß receptor signaling offers minimal protection in the LFRT, it is required to prevent dissemination of ZIKV to other tissues, including the upper FRT. Our findings support a role for RNA-sensing PRRs in the dampened innate immunity against ZIKV in the LFRT compared to other tissues and underlie potential implications for systemic dissemination upon heterosexual transmission of ZIKV in women.


Assuntos
Genitália Feminina/imunologia , Imunidade Inata/imunologia , RNA Viral/imunologia , Infecção por Zika virus/imunologia , Zika virus/imunologia , Animais , Feminino , Regulação Viral da Expressão Gênica , Genitália Feminina/metabolismo , Genitália Feminina/virologia , Humanos , Imunidade Inata/genética , Macaca mulatta , Camundongos Endogâmicos C57BL , Camundongos Knockout , RNA Viral/genética , Receptor de Interferon alfa e beta/genética , Receptor de Interferon alfa e beta/imunologia , Receptor de Interferon alfa e beta/metabolismo , Receptores de Reconhecimento de Padrão/genética , Receptores de Reconhecimento de Padrão/imunologia , Receptores de Reconhecimento de Padrão/metabolismo , Receptor 3 Toll-Like/genética , Receptor 3 Toll-Like/imunologia , Receptor 3 Toll-Like/metabolismo , Vagina/imunologia , Vagina/metabolismo , Vagina/virologia , Replicação Viral/genética , Replicação Viral/imunologia , Zika virus/genética , Zika virus/fisiologia , Infecção por Zika virus/genética , Infecção por Zika virus/virologia
6.
PLoS Negl Trop Dis ; 13(9): e0007695, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31527907

RESUMO

Zika virus infection is associated with the development of Guillain-Barré syndrome (GBS), a neurological autoimmune disorder caused by immune recognition of gangliosides and other components at nerve membranes. Using a high-throughput ELISA, we have analyzed the anti-glycolipid antibody profile, including gangliosides, of plasma samples from patients with Zika infections associated or not with GBS in Salvador, Brazil. We have observed that Zika patients that develop GBS present higher levels of anti-ganglioside antibodies when compared to Zika patients without GBS. We also observed that a broad repertoire of gangliosides was targeted by both IgM and IgG anti-self antibodies in these patients. Since Zika virus infects neurons, which contain membrane gangliosides, antigen presentation of these infected cells may trigger the observed autoimmune anti-ganglioside antibodies suggesting direct infection-induced autoantibodies as a cause leading to GBS development. Collectively, our results establish a link between anti-ganglioside antibodies and Zika-associated GBS in patients.


Assuntos
Gangliosídeos/imunologia , Síndrome de Guillain-Barré/sangue , Infecção por Zika virus/sangue , Zika virus/fisiologia , Autoanticorpos , Brasil , Síndrome de Guillain-Barré/diagnóstico , Síndrome de Guillain-Barré/virologia , Humanos , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Infecção por Zika virus/diagnóstico , Infecção por Zika virus/virologia
7.
Int J Infect Dis ; 88: 49-59, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31499212

RESUMO

INTRODUCTION: While death due to Zika virus (ZIKV) infection has been described, reports of fatal cases have been infrequent and no systematic reviews on the subject have been published. METHODS: A systematic review of the literature in four databases was performed to assess fatal outcomes of postnatal ZIKV infection and the available evidence that links ZIKV infection to death. RESULTS: Three hundred and eleven articles were retrieved; 20 of them were epidemiological reports from surveillance agencies and ministries of health. After screening by abstract and title, 59 articles were selected for full-text assessment. Of these, 35 were excluded (with reasons) and 24 were finally included for qualitative analysis. A total of 51 reported deaths associated with ZIKV infection in nine countries were identified. The majority of cases (56.9%) were not related to Guillain-Barré syndrome. Cases from three countries accounted for 67.6% of the deaths. ZIKV infection was laboratory-confirmed in the majority of cases (64.7%). DISCUSSION: ZIKV was not considered to be a dangerous, and much less a lethal pathogen, until very recently. However, an increasing number of fatalities have been published in the literature since the first death was reported in 2016. Additional research is needed to elucidate factors that may mediate the pathogenesis of severe, atypical, and fatal disease.


Assuntos
Infecção por Zika virus/epidemiologia , Zika virus/fisiologia , Américas/epidemiologia , Humanos , Infecção por Zika virus/mortalidade , Infecção por Zika virus/virologia
8.
Genomics Proteomics Bioinformatics ; 17(3): 319-331, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31494268

RESUMO

Proteins usually associate with other molecules physically to execute their functions. Identifying these interactions is important for the functional analysis of proteins. Previously, we reported the parallel analysis of translated ORFs (PLATO) to couple ribosome display of full-length ORFs with affinity enrichment of mRNA/protein/ribosome complexes for the "bait" molecules, followed by the deep sequencing analysis of mRNA. However, the sample processing, from extraction of precipitated mRNA to generation of DNA libraries, includes numerous steps, which is tedious and may cause the loss of materials. Barcoded PLATO (PLATO-BC), an improved platform was further developed to test its application for protein interaction discovery. In this report, we tested the antisera-antigen interaction using serum samples from patients with inclusion body myositis (IBM). Tripartite motif containing 21 (TRIM21) was identified as a potentially new IBM autoantigen. We also expanded the application of PLATO-BC to identify protein interactions for JQ1, single ubiquitin peptide, and NS5 protein of Zika virus. From PLATO-BC analyses, we identified new protein interactions for these "bait" molecules. We demonstrate that Ewing sarcoma breakpoint region 1 (EWSR1) binds to JQ1 and their interactions may interrupt the EWSR1 association with acetylated histone H4. RIO kinase 3 (RIOK3), a newly identified ubiquitin-binding protein, is preferentially associated with K63-ubiquitin chain. We also find that Zika NS5 protein interacts with two previously unreported host proteins, par-3 family cell polarity regulator (PARD3) and chromosome 19 open reading frame 53 (C19orf53), whose attenuated expression benefits the replication of Zika virus. These results further demonstrate that PLATO-BC is capable of identifying novel protein interactions for various types of "bait" molecules.


Assuntos
Fases de Leitura Aberta/genética , Mapeamento de Interação de Proteínas/métodos , Anticorpos/metabolismo , Células HEK293 , Humanos , Peptídeos/metabolismo , Ligação Proteica , Ubiquitina/metabolismo , Zika virus/fisiologia , Infecção por Zika virus/genética , Infecção por Zika virus/virologia
9.
Nat Commun ; 10(1): 3890, 2019 09 05.
Artigo em Inglês | MEDLINE | ID: mdl-31488835

RESUMO

Neurological complications affecting the central nervous system have been reported in adult patients infected by Zika virus (ZIKV) but the underlying mechanisms remain unknown. Here, we report that ZIKV replicates in human and mouse adult brain tissue, targeting mature neurons. ZIKV preferentially targets memory-related brain regions, inhibits hippocampal long-term potentiation and induces memory impairment in adult mice. TNF-α upregulation, microgliosis and upregulation of complement system proteins, C1q and C3, are induced by ZIKV infection. Microglia are found to engulf hippocampal presynaptic terminals during acute infection. Neutralization of TNF-α signaling, blockage of microglial activation or of C1q/C3 prevent synapse and memory impairment in ZIKV-infected mice. Results suggest that ZIKV induces synapse and memory dysfunction via aberrant activation of TNF-α, microglia and complement. Our findings establish a mechanism by which ZIKV affects the adult brain, and point to the need of evaluating cognitive deficits as a potential comorbidity in ZIKV-infected adults.


Assuntos
Encéfalo/virologia , Sinapses/virologia , Replicação Viral , Infecção por Zika virus/virologia , Zika virus/fisiologia , Animais , Comportamento Animal , Encéfalo/metabolismo , Encéfalo/patologia , Proteínas do Sistema Complemento/metabolismo , Modelos Animais de Doenças , Hipocampo/metabolismo , Humanos , Inflamação , Aprendizagem , Masculino , Memória , Transtornos da Memória , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Microglia/metabolismo , Microglia/patologia , Neurônios/virologia , Terminações Pré-Sinápticas/metabolismo , Receptores Tipo I de Interleucina-1/genética , Sinapses/metabolismo , Fator de Necrose Tumoral alfa/metabolismo
10.
Virol J ; 16(1): 99, 2019 08 08.
Artigo em Inglês | MEDLINE | ID: mdl-31395061

RESUMO

BACKGROUND: Both vector borne and sexual transmission of Zika virus (ZIKV) involve infection of epithelial cells in the initial stages of infection. Epithelial cells are unique in their ability to form polarized monolayers and their barrier function. Cell polarity induces an asymmetry in the epithelial monolayer, which is maintained by tight junctions and specialized sorting machinery. This differential localization can have a potential impact of virus infection. Asymmetrical distribution of a viral receptor can restrict virus entry to a particular membrane while polarized sorting can lead to a directional release of virions. The present study examined the impact of cell polarity on ZIKV infection and release. METHODS: A polarized Caco-2 cell model we described previously was used to assess ZIKV infection. Transepithelial resistance (TEER) was used to assess epithelial cell polarity, and virus infection was measured by immunofluorescence microscopy and qRT-PCR. Cell permeability was measured using a fluorescein leakage assay. Statistical significance was calculated using one-way ANOVA and significance was set at p < 0.05. RESULTS: Using the Caco-2 cell model for polarized epithelial cells, we report that Zika virus preferentially infects polarized cells from the apical route and is released vectorially through the basolateral route. Our data also indicates that release occurs without disruption of cell permeability. CONCLUSIONS: Our results show that ZIKV has directional infection and egress in a polarized cell system. This mechanism of directional infection may be one of the mechanisms that enables the cross the epithelial barrier effectively without a disruption in cell monolayer integrity. Elucidation of entry and release characteristics of Zika virus in polarized epithelial cells can lead to better understanding of virus dissemination in the host, and can help in developing effective therapeutic interventions.


Assuntos
Polaridade Celular , Células Epiteliais/virologia , Internalização do Vírus , Zika virus/fisiologia , Células CACO-2 , Humanos , Microscopia de Fluorescência , Receptores Virais/fisiologia
11.
Emerg Microbes Infect ; 8(1): 1003-1016, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31282298

RESUMO

Zika virus (ZIKV) is a mosquito-borne Flavivirus that causes Zika disease with particular neurological complications, including Guillain-Barré Syndrome and congenital microcephaly. Although ZIKV has been shown to directly infect human neural progenitor cells (hNPCs), thereby decreasing their viability and growth, it is as yet unknown which of the cellular pathways involved in the disruption of neurogenesis are affected following ZIKV infection. By comparing the effect of two ZIKV strains in vitro on hNPCs, the differentiation process of the latter cells was found to lead to a decreased susceptibility to infection and cell death induced by each of the ZIKV strains, which was associated with an earlier and stronger antiviral innate immune response in infected, differentiated hNPCs, as compared to undifferentiated cells. Moreover, ZIKV modulated, both in hNPCs and in vivo in fetal brain in an experimental mouse model, the expression of the Notch pathway which is involved in cellular proliferation, apoptosis and differentiation during neurogenesis. These results show that the differentiation state of hNPCs is a significant factor contributing to the outcome of ZIKV infection and furthermore suggest that ZIKV infection might initiate early activation of the Notch pathway resulting in an abnormal differentiation process, implicated in ZIKV-induced brain injury.


Assuntos
Células-Tronco Neurais/virologia , Neurogênese , Receptor Notch1/metabolismo , Infecção por Zika virus/virologia , Zika virus/fisiologia , Animais , Apoptose , Feminino , Humanos , Camundongos , Células-Tronco Neurais/citologia , Células-Tronco Neurais/metabolismo , Receptor Notch1/genética , Transdução de Sinais , Zika virus/genética , Infecção por Zika virus/genética , Infecção por Zika virus/metabolismo , Infecção por Zika virus/fisiopatologia
12.
Emerg Microbes Infect ; 8(1): 1098-1107, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31340725

RESUMO

Studies in mice showed that African Zika virus (ZIKV) strains cause more damage in embryos. These studies, however, were limited to the mouse-adapted African MR766 strain or infection at early gestation. Here, we compared infection of Asian and African strains in the fetal pig model at midgestation. Both strains caused fetal infection. ZIKV was detected in placenta, amniotic membrane, amniotic fluid, fetal blood, and brain. The African strain produced more vigorous in utero infection as represented by more efficient virus transmission between siblings, and higher viral loads in fetal organs and membranes. Infection with both strains was associated with reduced fetal brain weight and increased number of placental CD163-positive cells, as well as elevated in utero interferon alpha and cortisol levels. This is the first large animal model study which demonstrated that African strain of ZIKV, with no passage history in experimental animals, can cause persistent infection in fetuses and fetal membranes at midgestation. Our studies also suggest that similar to Asian strains, ZIKV of African lineage might cause silent pathology which is difficult to identify in deceptively healthy fetuses. The findings emphasize the need for further studies to highlight the impact of ZIKV heterogeneity on infection outcomes during pregnancy.


Assuntos
Doenças Fetais/virologia , Complicações Infecciosas na Gravidez/virologia , Infecção por Zika virus/virologia , Zika virus/fisiologia , Animais , Encéfalo/virologia , Modelos Animais de Doenças , Feminino , Humanos , Placenta/virologia , Gravidez , Suínos , Útero/virologia , Zika virus/classificação , Zika virus/genética , Infecção por Zika virus/transmissão
13.
Emerg Infect Dis ; 25(8): 1477-1484, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31310224

RESUMO

We characterized natural vertical transmission of Zika virus in pools of Aedes aegypti larvae hatched from eggs collected in Jojutla, Morelos, Mexico. Of the 151 pools analyzed, 17 tested positive for Zika virus RNA; infectious Zika virus was successfully isolated from 1 of the larvae pools (31N) in C6/36 cells. Real-time quantitative PCR and indirect immunofluorescence assays confirmed the identity of the isolate, named Zika virus isolate 31N; plaque assays in Vero cells demonstrated the isolate's infectivity in a mammalian cell line. We obtained the complete genome of Zika virus isolate 31N by next-generation sequencing and identified 3 single-nucleotide variants specific to Zika virus isolate 31N using the meta-CATS tool. These results demonstrate the occurrence of natural vertical transmission of Zika virus in wild Ae. aegypti mosquitoes and suggest that this transmission mode could aid in the spread and maintenance of Zika virus in nature.


Assuntos
Aedes/virologia , Mosquitos Vetores/virologia , Zika virus/fisiologia , Animais , Linhagem Celular , Células Cultivadas , Microbiologia Ambiental , Genoma Viral , Humanos , Transmissão Vertical de Doença Infecciosa , Larva , México/epidemiologia , Filogenia , Vigilância em Saúde Pública , Células Vero , Carga Viral , Ensaio de Placa Viral , Sequenciamento Completo do Genoma , Zika virus/classificação , Infecção por Zika virus/transmissão , Infecção por Zika virus/virologia
14.
Cells ; 8(7)2019 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-31311201

RESUMO

Zika virus (ZIKV) has emerged as an important human pathogen that can cause congenital defects in the fetus and neurological conditions in adults. The interferon (IFN) system has proven crucial in restricting ZIKV replication and pathogenesis. The canonical IFN response is triggered by the detection of viral RNA through RIG-I like receptors followed by activation of the adaptor protein MAVS on mitochondrial membranes. Recent studies have shown that a second organelle, peroxisomes, also function as a signaling platforms for the IFN response. Here, we investigated how ZIKV infection affects peroxisome biogenesis and antiviral signaling. We show that ZIKV infection depletes peroxisomes in human fetal astrocytes, a brain cell type that can support persistent infection. The peroxisome biogenesis factor PEX11B was shown to inhibit ZIKV replication, likely by increasing peroxisome numbers and enhancing downstream IFN-dependent antiviral signaling. Given that peroxisomes play critical roles in brain development and nerve function, our studies provide important insights into the roles of peroxisomes in regulating ZIKV infection and potentially neuropathogenesis.


Assuntos
Interações Hospedeiro-Patógeno , Peroxissomos/virologia , Zika virus/patogenicidade , Animais , Astrócitos/imunologia , Astrócitos/virologia , Linhagem Celular Tumoral , Células Cultivadas , Células HEK293 , Humanos , Imunidade Inata , Interferons/metabolismo , Proteínas de Membrana/metabolismo , Transdução de Sinais , Células Vero , Replicação Viral , Zika virus/fisiologia
15.
Int J Mol Sci ; 20(12)2019 Jun 21.
Artigo em Inglês | MEDLINE | ID: mdl-31234341

RESUMO

Zika virus (ZIKV) transmission can cause serious fetal neurological abnormalities. ZIKV persistence in various human cells and tissues can serve as infectious reservoirs and post serious threats to public health. The human embryonic kidney (HEK293) cell line with known neuronal developmental properties was readily infected by ZIKV in a strain-dependent fashion. Significant cytopathic effect in HEK293 cells infected by the prototype MR 766 strain of ZIKV resulted in complete loss of cells, while small numbers of HEK293 cells infected by contemporary ZIKV isolates (PRV or FLR strain) continued to survive and regrow to confluency in the culture around two months after initial infection. Most, if not all, of the cells in the two resulting persistently ZIKV-infected HEK293 cell lines tested positive for ZIKV antigen. Compared to HEK293 control cells, the persistently ZIKV-infected HEK293 cells had slower growth rates with some cells undergoing apoptosis in culture. The "persistent ZIKVs" produced constitutively by both PRV and FLR strains ZIKV-infected HEK293 cells had significantly attenuated cell infectivity and/or cytopathogenicity. Comparative genome sequence analyses between the persistent ZIKVs and the original inoculum ZIKVs showed no clonal selection with specific gene mutations in the prolonged process of establishing persistently PRV strain ZIKV-infected HEK293 cells; while selection of ZIKV subclones with mutations in the envelope, protein pr and multiple NS genes was evident in developing persistently FLR strain ZIKV-infected HEK293 cell line. Our study provides molecular insights into the complex interplays of ZIKV and human host cells in establishing ZIKV persistence.


Assuntos
Infecção por Zika virus/patologia , Zika virus/fisiologia , Animais , Apoptose , Efeito Citopatogênico Viral , Genoma Viral , Genômica , Células HEK293 , Humanos , Mutação , Células Vero , Zika virus/genética , Zika virus/patogenicidade , Infecção por Zika virus/virologia
16.
Microbes Infect ; 21(8-9): 353-360, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31158508

RESUMO

Since the ZIKV outbreak in Brazil in 2015, the scientific community has joined efforts to gather more information on the epidemiology, clinical features and pathogenicity of the virus. Here, we summarize the most important advances made recently and discuss promising, innovative approaches to understand and control ZIKV infection.


Assuntos
Infecção por Zika virus/prevenção & controle , Infecção por Zika virus/virologia , Zika virus/fisiologia , Zika virus/patogenicidade , Animais , Antivirais/farmacologia , Arbovirus/patogenicidade , Arbovirus/fisiologia , Brasil/epidemiologia , Humanos , Imunidade Inata , Mosquitos Vetores/virologia , Vacinas Virais/imunologia , Zika virus/efeitos dos fármacos , Infecção por Zika virus/epidemiologia , Infecção por Zika virus/imunologia
17.
Insect Biochem Mol Biol ; 111: 103169, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31103782

RESUMO

The yellow fever mosquito, Aedes aegypti, serves as the primary vector for epidemic transmission of yellow fever, dengue, Zika (ZIKV), and chikungunya viruses to humans. Control of Ae. aegypti is currently limited to insecticide applications and larval habitat management; however, to combat growing challenges with insecticide resistance, novel genetic approaches for vector population reduction or transmission interruption are being aggressively pursued. The objectives of this study were to assess the ability of the Ae. aegypti antiviral exogenous-small interfering RNA (exo-siRNA) response to inhibit ZIKV infection and transmission, and to identify the optimal RNA interference (RNAi) target region in the ZIKV genome. We accomplished these objectives by in vitro transcription of five long double-stranded RNAs (dsRNAs) from the genome region spanning the NS2B-NS3-NS4A genes, which were the most highly conserved among ZIKV RNA sequences representing both East and West African and Asian-American clades, and evaluation of the ability of these dsRNAs to trigger an effective antiviral exo-siRNA response after intrathoracic injection into Ae. aegypti. In a pilot study, five ZIKV dsRNAs were tested by intrathoracic inoculation of 250 ng dsRNA into groups of approximately 5-day-old mosquitoes. Three days post-inoculation, mosquitoes were provided an infectious blood-meal containing ZIKV strain PRVABC59 (Puerto Rico), MR766 (Uganda), or 41525 (Senegal). On days 7 and 14 post-infection individual whole mosquito bodies were assessed for ZIKV infectious titer by plaque assays. Based on the results of this initial assessment, three dsRNAs were selected for further evaluation of viral loads of matched body and saliva expectorants using a standardized infectious dose of 1 × 107 PFU/mL of each ZIKV strain. Fourteen days post-exposure to ZIKV, paired saliva and carcass samples were harvested from individual mosquitoes and assessed for ZIKV RNA load by qRT-PCR. Injection of each of the three dsRNAs resulted in significant inhibition of replication of all three strains of ZIKV in mosquito bodies and saliva. This study lays critical groundwork for pursuing ZIKV transmission-blocking strategies that exploit the Ae. aegypti exo-siRNA response for arbovirus suppression in natural populations.


Assuntos
Aedes/virologia , Interferência de RNA , Infecção por Zika virus/transmissão , Zika virus/genética , Animais , Bovinos , Mosquitos Vetores/virologia , Projetos Piloto , RNA de Cadeia Dupla , RNA Interferente Pequeno , Saliva/virologia , Análise de Sequência de RNA , Células Vero , Carga Viral , Replicação Viral , Zika virus/fisiologia , Infecção por Zika virus/virologia
18.
Virology ; 533: 59-67, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31112915

RESUMO

Zika virus (ZIKV) is an emerging pathogen with global health and economic impacts. ZIKV circulates as two major lineages, Asian or African. The Asian lineage has recently been associated with significant disease in humans. Numerous studies have revealed differences between African and Asian ZIKV strains with respect to cellular infectivity, pathogenesis, and immune activation. Less is known about the mechanism of ZIKV entry and whether viral entry differs between strains. Here, we characterized ZIKV entry with two Asian and two African strains. All viruses exhibited a requirement for clathrin-mediated endocytosis and Rab5a function. Additionally, all ZIKV strains tested were sensitive to pH in the range of 6.5-6.1 and were reliant on endosomal acidification for infection. Finally, we provide direct evidence that ZIKV primarily fuses with late endosomes. These findings contribute new insight into the ZIKV entry process and suggest that divergent ZIKV strains enter cells in a highly conserved manner.


Assuntos
Internalização do Vírus , Infecção por Zika virus/virologia , Zika virus/fisiologia , África , Ásia , Endocitose , Endossomos/virologia , Humanos , Zika virus/classificação , Zika virus/genética , Infecção por Zika virus/fisiopatologia
19.
PLoS Negl Trop Dis ; 13(5): e0007443, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-31107912

RESUMO

BACKGROUND: Wolbachia's ability to restrict arbovirus transmission makes it a promising tool to combat mosquito-transmitted diseases. Wolbachia-infected Aedes aegypti are currently being released in locations such as Brazil, which regularly experience concurrent outbreaks of different arboviruses. A. aegypti can become co-infected with, and transmit multiple arboviruses with one bite, which can complicate patient diagnosis and treatment. METHODOLOGY/PRINCIPLE FINDINGS: Using experimental oral infection of A. aegypti and then RT-qPCR, we examined ZIKV/DENV-1 and ZIKV/DENV-3 co-infection in Wolbachia-infected A. aegypti and observed that Wolbachia-infected mosquitoes experienced lower prevalence of infection and viral load than wildtype mosquitoes, even with an extra infecting virus. Critically, ZIKV/DENV co-infection had no significant impact on Wolbachia's ability to reduce viral transmission. Wolbachia infection also strongly altered expression levels of key immune genes Defensin C and Transferrin 1, in a virus-dependent manner. CONCLUSIONS/SIGNIFICANCE: Our results suggest that pathogen interference in Wolbachia-infected A. aegypti is not adversely affected by ZIKV/DENV co-infection, which suggests that Wolbachia-infected A. aegypti will likely prove suitable for controlling mosquito-borne diseases in environments with complex patterns of arbovirus transmission.


Assuntos
Aedes/microbiologia , Aedes/virologia , Vírus da Dengue/fisiologia , Controle de Mosquitos/métodos , Mosquitos Vetores/microbiologia , Mosquitos Vetores/virologia , Wolbachia/fisiologia , Zika virus/fisiologia , Animais , Brasil , Vírus da Dengue/genética , Feminino , Masculino , Wolbachia/genética , Zika virus/genética
20.
Parasit Vectors ; 12(1): 204, 2019 May 03.
Artigo em Inglês | MEDLINE | ID: mdl-31053164

RESUMO

BACKGROUND: A number of mosquito-borne viruses such as dengue virus (DENV), Usutu virus (USUV), West Nile virus (WNV) are autochthonously transmitted in Europe and six invasive mosquito species have been detected in this temperate region. This has increased the risk for the emergence of further mosquito-borne diseases. However, there is a paucity of information on whether European populations of invasive mosquito species are competent to transmit arboviruses. In this study, the susceptibility of Aedes albopictus originating from Spain and a laboratory-adapted colony of Aedes aegypti, was assessed for infection with, and transmission of Zika virus (ZIKV). Vertical transmission in both species was also assessed. METHODS: Aedes albopictus colonised from eggs collected in Spain and an existing colony of Ae. aegypti were fed infectious blood meals containing ZIKV (Polynesian strain) at 1.6 × 107 PFU/ml. Blood-fed mosquitoes were separated and maintained at 20 °C or 25 °C. Legs, saliva and bodies were sampled from specimens at 7, 14 and 21 days post-infection (dpi) in order to determine infection, dissemination and transmission rates. All samples were analysed by real-time RT-PCR using primers targeting the ZIKV NS1 gene. RESULTS: At 14 dpi and 21 dpi, ZIKV RNA was detected in the bodies of both species at both temperatures. However, live virus only was detected in the saliva of Ae. aegypti at 25 °C with a transmission rate of 44%. No evidence for virus expectoration was obtained for Ae. albopictus under any condition. Notably, ZIKV RNA was not detectable in the saliva of Ae. aegypti at 20 °C after 21 days. No vertical transmission of ZIKV was detected in this study. CONCLUSIONS: Experimental infection of Ae. albopictus colonized from Spain with ZIKV did not result in expectoration of virus in saliva in contrast to results for Ae. aegypti. No evidence of vertical transmission of virus was observed in this study. This suggests that this strain of Ae. albopictus is not competent for ZIKV transmission under the conditions tested.


Assuntos
Aedes/virologia , Mosquitos Vetores/virologia , Infecção por Zika virus/transmissão , Zika virus/fisiologia , Aedes/crescimento & desenvolvimento , Aedes/fisiologia , Animais , Feminino , Humanos , Masculino , Mosquitos Vetores/crescimento & desenvolvimento , Mosquitos Vetores/fisiologia , Saliva/virologia , Espanha , Temperatura Ambiente , Zika virus/genética , Zika virus/isolamento & purificação , Infecção por Zika virus/virologia
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