RESUMO
Mitochondria are critical modulators of antiviral tolerance through the release of mitochondrial RNA and DNA (mtDNA and mtRNA) fragments into the cytoplasm after infection, activating virus sensors and type-I interferon (IFN-I) response1-4. The relevance of these mechanisms for mitochondrial diseases remains understudied. Here we investigated mitochondrial recessive ataxia syndrome (MIRAS), which is caused by a common European founder mutation in DNA polymerase gamma (POLG1)5. Patients homozygous for the MIRAS variant p.W748S show exceptionally variable ages of onset and symptoms5, indicating that unknown modifying factors contribute to disease manifestation. We report that the mtDNA replicase POLG1 has a role in antiviral defence mechanisms to double-stranded DNA and positive-strand RNA virus infections (HSV-1, TBEV and SARS-CoV-2), and its p.W748S variant dampens innate immune responses. Our patient and knock-in mouse data show that p.W748S compromises mtDNA replisome stability, causing mtDNA depletion, aggravated by virus infection. Low mtDNA and mtRNA release into the cytoplasm and a slow IFN response in MIRAS offer viruses an early replicative advantage, leading to an augmented pro-inflammatory response, a subacute loss of GABAergic neurons and liver inflammation and necrosis. A population databank of around 300,000 Finnish individuals6 demonstrates enrichment of immunodeficient traits in carriers of the POLG1 p.W748S mutation. Our evidence suggests that POLG1 defects compromise antiviral tolerance, triggering epilepsy and liver disease. The finding has important implications for the mitochondrial disease spectrum, including epilepsy, ataxia and parkinsonism.
Assuntos
Alelos , DNA Polimerase gama , Vírus da Encefalite Transmitidos por Carrapatos , Herpesvirus Humano 1 , Tolerância Imunológica , SARS-CoV-2 , Animais , Feminino , Humanos , Masculino , Camundongos , Idade de Início , COVID-19/imunologia , COVID-19/virologia , COVID-19/genética , DNA Polimerase gama/genética , DNA Polimerase gama/imunologia , DNA Polimerase gama/metabolismo , DNA Mitocondrial/imunologia , DNA Mitocondrial/metabolismo , Vírus da Encefalite Transmitidos por Carrapatos/imunologia , Encefalite Transmitida por Carrapatos/genética , Encefalite Transmitida por Carrapatos/imunologia , Encefalite Transmitida por Carrapatos/virologia , Efeito Fundador , Técnicas de Introdução de Genes , Herpes Simples/genética , Herpes Simples/imunologia , Herpes Simples/virologia , Herpesvirus Humano 1/imunologia , Tolerância Imunológica/genética , Tolerância Imunológica/imunologia , Imunidade Inata/genética , Imunidade Inata/imunologia , Interferon Tipo I/imunologia , Doenças Mitocondriais/enzimologia , Doenças Mitocondriais/genética , Doenças Mitocondriais/imunologia , Mutação , RNA Mitocondrial/imunologia , RNA Mitocondrial/metabolismo , SARS-CoV-2/imunologiaRESUMO
Tick-borne flaviviruses (TBFs) are transmitted to humans through milk and tick bites. Although a case of possible mother-to-child transmission of tick-borne encephalitis virus (TBEV) through breast milk has been reported, this route has not been confirmed in experimental models. Therefore, in this study, using type I interferon receptor-deficient A129 mice infected with Langat virus (LGTV), we aimed to demonstrate the presence of infectious virus in the milk and mammary glands of infected mice. Our results showed viral RNA of LGTV in the pup's stomach milk clots (SMCs) and blood, indicating that the virus can be transmitted from dam to pup through breast milk. In addition, we observed that LGTV infection causes tissue lesions in the mammary gland, and viral particles were present in mammary gland epithelial cells. Furthermore, we found that milk from infected mice could infect adult mice via the intragastric route, which has a milder infection process, longer infection time, and a lower rate of weight loss than other modes of infection. Specifically, we developed a nano-luciferase-LGTV reporter virus system to monitor the dynamics of different infection routes and observed dam-to-pup infection using in vivo bioluminescence imaging. This study provides comprehensive evidence to support breast milk transmission of TBF in mice and has helped provide useful data for studying TBF transmission routes.IMPORTANCETo date, no experimental models have confirmed mother-to-child transmission of tick-borne flavivirus (TBF) through breastfeeding. In this study, we used a mouse model to demonstrate the presence of infectious viruses in mouse breast milk and mammary gland epithelial cells. Our results showed that pups could become infected through the gastrointestinal route by suckling milk, and the infection dynamics could be monitored using a reporter virus system during breastfeeding in vivo. We believe our findings have provided substantial evidence to understand the underlying mechanism of breast milk transmission of TBF in mice, which has important implications for understanding and preventing TBF transmission in humans.
Assuntos
Vírus da Encefalite Transmitidos por Carrapatos , Encefalite Transmitida por Carrapatos , Transmissão Vertical de Doenças Infecciosas , Glândulas Mamárias Animais , Leite , Animais , Feminino , Camundongos , Vírus da Encefalite Transmitidos por Carrapatos/crescimento & desenvolvimento , Vírus da Encefalite Transmitidos por Carrapatos/fisiologia , Encefalite Transmitida por Carrapatos/transmissão , Encefalite Transmitida por Carrapatos/virologia , Glândulas Mamárias Animais/virologia , Leite/virologia , Animais Recém-Nascidos/virologiaRESUMO
Powassan virus is a tick-borne flavivirus that can cause severe neuroinvasive disease, with areas of endemicity in the Northeast and Midwest United States, Canada, and Russia. Diagnosis is challenging and relies on a high index of suspicion and choosing the right test based on duration of infection and the patient's immune status. This review covers laboratory testing for Powassan virus, including historical considerations, modern options, and methods being developed in the research space.
Assuntos
Vírus da Encefalite Transmitidos por Carrapatos , Encefalite Transmitida por Carrapatos , Humanos , Encefalite Transmitida por Carrapatos/diagnóstico , Encefalite Transmitida por Carrapatos/epidemiologia , Encefalite Transmitida por Carrapatos/virologia , Vírus da Encefalite Transmitidos por Carrapatos/isolamento & purificação , Técnicas de Laboratório Clínico/métodos , História do Século XXI , História do Século XX , Animais , Canadá/epidemiologia , Anticorpos Antivirais/sangueRESUMO
Ticks are important vectors of disease, particularly in the context of One Health, where tick-borne diseases (TBDs) are increasingly prevalent worldwide. TBDs often involve co-infections, where multiple pathogens co-exist in a single host. Patients with chronic Lyme disease often have co-infections with other bacteria or parasites. This study aimed to create a co-infection model with Borrelia afzelii and tick-borne encephalitis virus (TBEV) in C3H mice and to evaluate symptoms, mortality, and pathogen level compared to single infections. Successful co-infection of C3H mice with B. afzelii and TBEV was achieved. Outcomes varied, depending on the timing of infection. When TBEV infection followed B. afzelii infection by 9 days, TBEV symptoms worsened and virus levels increased. Conversely, mice infected 21 days apart with TBEV showed milder symptoms and lower mortality. Simultaneous infection resulted in mild symptoms and no deaths. However, our model did not effectively infect ticks with TBEV, possibly due to suboptimal dosing, highlighting the challenges of replicating natural conditions. Understanding the consequences of co-infection is crucial, given the increasing prevalence of TBD. Co-infected individuals may experience exacerbated symptoms, highlighting the need for a comprehensive understanding through refined animal models. This study advances knowledge of TBD and highlights the importance of exploring co-infection dynamics in host-pathogen interactions.
Assuntos
Coinfecção , Modelos Animais de Doenças , Vírus da Encefalite Transmitidos por Carrapatos , Encefalite Transmitida por Carrapatos , Doença de Lyme , Camundongos Endogâmicos C3H , Animais , Coinfecção/microbiologia , Coinfecção/virologia , Camundongos , Vírus da Encefalite Transmitidos por Carrapatos/fisiologia , Vírus da Encefalite Transmitidos por Carrapatos/patogenicidade , Doença de Lyme/microbiologia , Encefalite Transmitida por Carrapatos/virologia , Grupo Borrelia Burgdorferi , FemininoRESUMO
Tick-borne encephalitis (TBE) virus is the most prevalent tick-transmitted orthoflavivirus in Europe. Due to the nonspecific nature of its symptoms, TBE is primarily diagnosed by ELISA-based detection of specific antibodies in the patient serum. However, cross-reactivity between orthoflaviviruses complicates the diagnosis. Specificity issues may be mitigated by serum neutralization assays (SNT), although the handling of clinically relevant orthoflaviviruses requires biosafety level (BSL) 3 conditions and they have highly divergent viral kinetics and cell tropisms. In the present study, we established a reporter virus particle (RVP)-based SNT in which the infectivity is measured by luminescence and that can be performed under BSL-2 conditions. The RVP-based SNT for TBEV exhibited a highly significant correlation with the traditional virus-based SNT (R2 = 0.8637, p < 0.0001). The RVP-based assay demonstrated a sensitivity of 92.3% (95% CI: 79.7%-97.4%) and specificity of 100% (95% CI: 81.6%-100%). We also tested the cross-reactivity of serum samples in RVP-based assays against other orthoflaviviruses (yellow fever virus, dengue virus type 2, Zika virus, West Nile virus and Japanese encephalitis virus). Interestingly, all serum samples which had tested TBEV-positive by ELISA but negative by RVP-based SNT were reactive for antibodies against other orthoflaviviruses. Thus, the RVP-based seroneutralization assay provides an added value in clinical diagnostics as well as in epidemiological studies.
Assuntos
Anticorpos Antivirais , Reações Cruzadas , Vírus da Encefalite Transmitidos por Carrapatos , Encefalite Transmitida por Carrapatos , Ensaio de Imunoadsorção Enzimática , Testes de Neutralização , Sensibilidade e Especificidade , Vírus da Encefalite Transmitidos por Carrapatos/imunologia , Humanos , Anticorpos Antivirais/sangue , Testes de Neutralização/métodos , Encefalite Transmitida por Carrapatos/diagnóstico , Encefalite Transmitida por Carrapatos/virologia , Ensaio de Imunoadsorção Enzimática/métodos , Vírion/imunologia , Anticorpos Neutralizantes/sangue , Anticorpos Neutralizantes/imunologia , AnimaisRESUMO
BACKGROUND: Tick-borne encephalitis (TBE) is a severe human neuroinfection caused by TBE virus (TBEV). TBEV is transmitted by tick bites and by the consumption of unpasteurized dairy products from infected asymptomatic ruminants. In France, several food-borne transmission events have been reported since 2020, raising the question of the level of exposure of domestic ungulates to TBEV. In this study, our objectives were (i) to estimate TBEV seroprevalence and quantify antibodies titres in cattle in the historical endemic area of TBEV in France using the micro virus neutralisation test (MNT) and (ii) to compare the performance of two veterinary cELISA kits with MNT for detecting anti-TBEV antibodies in cattle in various epidemiological contexts. A total of 344 cattle sera from four grid cells of 100 km² in Alsace-Lorraine (endemic region) and 84 from western France, assumed to be TBEV-free, were investigated. RESULTS: In Alsace-Lorraine, cattle were exposed to the virus with an overall estimated seroprevalence of 57.6% (95% CI: 52.1-62.8%, n = 344), varying locally from 29.9% (95% CI: 21.0-40.0%) to 92.1% (95% CI: 84.5-96.8%). Seroprevalence did not increase with age, with one- to three-year-old cattle being as highly exposed as older ones, suggesting a short-life duration of antibodies. The proportion of sera with MNT titres lower than 1:40 per grid cell decreased with increased seroprevalence. Both cELISA kits showed high specificity (> 90%) and low sensitivity (less than 78.1%) compared with MNT. Sensitivity was lower for sera with neutralising antibodies titres below 1:40, suggesting that sensitivity of these tests varied with local virus circulation intensity. CONCLUSIONS: Our results highlight that cattle were highly exposed to TBEV. Screening strategy and serological tests should be carefully chosen according to the purpose of the serological study and with regard to the limitations of each method.
Assuntos
Anticorpos Antivirais , Doenças dos Bovinos , Vírus da Encefalite Transmitidos por Carrapatos , Encefalite Transmitida por Carrapatos , Animais , Bovinos , Encefalite Transmitida por Carrapatos/epidemiologia , Encefalite Transmitida por Carrapatos/veterinária , Encefalite Transmitida por Carrapatos/virologia , Vírus da Encefalite Transmitidos por Carrapatos/imunologia , Vírus da Encefalite Transmitidos por Carrapatos/isolamento & purificação , França/epidemiologia , Estudos Soroepidemiológicos , Doenças dos Bovinos/epidemiologia , Doenças dos Bovinos/virologia , Anticorpos Antivirais/sangue , Feminino , Masculino , Testes de Neutralização/veterinária , Doenças Endêmicas/veterináriaRESUMO
Tick-borne encephalitis virus (TBEV) targets the central nervous system (CNS), leading to potentially severe neurological complications. The neurovascular unit plays a fundamental role in the CNS and in the neuroinvasion of TBEV. However, the role of human brain pericytes, a key component of the neurovascular unit, during TBEV infection has not yet been elucidated. In this study, TBEV infection of the primary human brain perivascular pericytes was investigated with highly virulent Hypr strain and mildly virulent Neudoerfl strain. We used Luminex assay to measure cytokines/chemokines and growth factors. Both viral strains showed comparable replication kinetics, peaking at 3 days post infection (dpi). Intracellular viral RNA copies peaked at 6 dpi for Hypr and 3 dpi for Neudoerfl cultures. According to immunofluorescence staining, only small proportion of pericytes were infected (3% for Hypr and 2% for Neudoerfl), and no cytopathic effect was observed in the infected cells. In cell culture supernatants, IL-6 production was detected at 3 dpi, together with slight increases in IL-15 and IL-4, but IP-10, RANTES and MCP-1 were the main chemokines released after TBEV infection. These chemokines play key roles in both immune defense and immunopathology during TBE. This study suggests that pericytes are an important source of these signaling molecules during TBEV infection in the brain.
Assuntos
Encéfalo , Quimiocina CCL5 , Quimiocina CXCL10 , Vírus da Encefalite Transmitidos por Carrapatos , Encefalite Transmitida por Carrapatos , Pericitos , Pericitos/virologia , Pericitos/metabolismo , Humanos , Vírus da Encefalite Transmitidos por Carrapatos/fisiologia , Vírus da Encefalite Transmitidos por Carrapatos/patogenicidade , Encéfalo/virologia , Encéfalo/metabolismo , Encéfalo/patologia , Quimiocina CXCL10/metabolismo , Encefalite Transmitida por Carrapatos/virologia , Encefalite Transmitida por Carrapatos/metabolismo , Quimiocina CCL5/metabolismo , Células Cultivadas , Replicação Viral , Citocinas/metabolismoRESUMO
Powassan viruses (POWVs) are neurovirulent tick-borne flaviviruses emerging in the northeastern United States, with a 2% prevalence in Long Island (LI) deer ticks (Ixodes scapularis). POWVs are transmitted within as little as 15 min of a tick bite and enter the central nervous system (CNS) to cause encephalitis (10% of cases are fatal) and long-term neuronal damage. POWV-LI9 and POWV-LI41 present in LI Ixodes ticks were isolated by directly inoculating VeroE6 cells with tick homogenates and detecting POWV-infected cells by immunoperoxidase staining. Inoculated POWV-LI9 and LI41 were exclusively present in infected cell foci, indicative of cell to cell spread, despite growth in liquid culture without an overlay. Cloning and sequencing establish POWV-LI9 as a phylogenetically distinct lineage II POWV strain circulating in LI deer ticks. Primary human brain microvascular endothelial cells (hBMECs) and pericytes form a neurovascular complex that restricts entry into the CNS. We found that POWV-LI9 and -LI41 and lineage I POWV-LB productively infect hBMECs and pericytes and that POWVs were basolaterally transmitted from hBMECs to lower-chamber pericytes without permeabilizing polarized hBMECs. Synchronous POWV-LI9 infection of hBMECs and pericytes induced proinflammatory chemokines, interferon-ß (IFN-ß) and proteins of the IFN-stimulated gene family (ISGs), with delayed IFN-ß secretion by infected pericytes. IFN inhibited POWV infection, but despite IFN secretion, a subset of POWV-infected hBMECs and pericytes remained persistently infected. These findings suggest a potential mechanism for POWVs (LI9/LI41 and LB) to infect hBMECs, spread basolaterally to pericytes, and enter the CNS. hBMEC and pericyte responses to POWV infection suggest a role for immunopathology in POWV neurovirulence and potential therapeutic targets for preventing POWV spread to neuronal compartments. IMPORTANCE We isolated POWVs from LI deer ticks (I. scapularis) directly in VeroE6 cells, and sequencing revealed POWV-LI9 as a distinct lineage II POWV strain. Remarkably, inoculation of VeroE6 cells with POWV-containing tick homogenates resulted in infected cell foci in liquid culture, consistent with cell-to-cell spread. POWV-LI9 and -LI41 and lineage I POWV-LB strains infected hBMECs and pericytes that comprise neurovascular complexes. POWVs were nonlytically transmitted basolaterally from infected hBMECs to lower-chamber pericytes, suggesting a mechanism for POWV transmission across the blood-brain barrier (BBB). POWV-LI9 elicited inflammatory responses from infected hBMEC and pericytes that may contribute to immune cell recruitment and neuropathogenesis. This study reveals a potential mechanism for POWVs to enter the CNS by infecting hBMECs and spreading basolaterally to abluminal pericytes. Our findings reveal that POWV-LI9 persists in cells that form a neurovascular complex spanning the BBB and suggest potential therapeutic targets for preventing POWV spread to neuronal compartments.
Assuntos
Vetores de Doenças , Vírus da Encefalite Transmitidos por Carrapatos/fisiologia , Encefalite Transmitida por Carrapatos/virologia , Ixodes/virologia , Animais , Células Cultivadas , Vírus da Encefalite Transmitidos por Carrapatos/classificação , Vírus da Encefalite Transmitidos por Carrapatos/efeitos dos fármacos , Vírus da Encefalite Transmitidos por Carrapatos/isolamento & purificação , Encefalite Transmitida por Carrapatos/imunologia , Encefalite Transmitida por Carrapatos/transmissão , Células Endoteliais , Ordem dos Genes , Genoma Viral , Interações Hospedeiro-Patógeno/imunologia , Humanos , Interferons/farmacologia , Pericitos/virologia , Filogenia , Replicação Viral/efeitos dos fármacosRESUMO
Tick-borne encephalitis virus (TBEV), of the genus Flavivirus, is a causative agent of severe encephalitis in regions of endemicity of northern Asia and central and northern Europe. Interferon-induced transmembrane proteins (IFITMs) are restriction factors that inhibit the replication cycles of numerous viruses, including flaviviruses such as West Nile virus, dengue virus, and Zika virus. Here, we demonstrate the role of IFITM1, IFITM2, and IFITM3 in the inhibition of TBEV infection and in protection against virus-induced cell death. We show that the most significant role is that of IFITM3, including the dissection of its functional motifs by mutagenesis. Furthermore, through the use of CRISPR-Cas9-generated IFITM1/3-knockout monoclonal cell lines, we confirm the role and additive action of endogenous IFITMs in TBEV suppression. However, the results of coculture assays suggest that TBEV might partially escape interferon- and IFITM-mediated suppression during high-density coculture infection when the virus enters naive cells directly from infected donor cells. Thus, cell-to-cell spread may constitute a strategy for virus escape from innate host defenses. IMPORTANCE TBEV infection may result in encephalitis, chronic illness, or death. TBEV is endemic in northern Asia and Europe; however, due to climate change, new centers of endemicity have arisen. Although effective TBEV vaccines have been approved, vaccination coverage is low, and due to the lack of specific therapeutics, infected individuals depend on their immune responses to control the infection. IFITM proteins are components of the innate antiviral defenses that suppress cell entry of many viral pathogens. However, no studies on the role of IFITM proteins in TBEV infection have been published thus far. Understanding antiviral innate immune responses is crucial for the future development of antiviral strategies. Here, we show the important role of IFITM proteins in the inhibition of TBEV infection and virus-mediated cell death. However, our data suggest that TBEV cell-to-cell spread may be less prone to both interferon- and IFITM-mediated suppression, potentially facilitating escape from IFITM-mediated immunity.
Assuntos
Vírus da Encefalite Transmitidos por Carrapatos/fisiologia , Encefalite Transmitida por Carrapatos/metabolismo , Encefalite Transmitida por Carrapatos/virologia , Interações Hospedeiro-Patógeno , Interferons/metabolismo , Proteínas de Membrana/metabolismo , Sequência de Aminoácidos , Linhagem Celular , Efeito Citopatogênico Viral , Resistência à Doença/genética , Resistência à Doença/imunologia , Suscetibilidade a Doenças , Encefalite Transmitida por Carrapatos/genética , Encefalite Transmitida por Carrapatos/imunologia , Expressão Gênica , Técnicas de Silenciamento de Genes , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/imunologia , Humanos , Proteínas de Membrana/química , Proteínas de Membrana/genética , Família Multigênica , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , Replicação ViralRESUMO
Kyasanur Forest disease virus (KFDV) and the closely related Alkhurma hemorrhagic disease virus (AHFV) are emerging flaviviruses that cause severe viral hemorrhagic fevers in humans. Increasing geographical expansion and case numbers, particularly of KFDV in southwest India, class these viruses as a public health threat. Viral pathogenesis is not well understood and additional vaccines and antivirals are needed to effectively counter the impact of these viruses. However, current animal models of KFDV pathogenesis do not accurately reproduce viral tissue tropism or clinical outcomes observed in humans. Here, we show that pigtailed macaques (Macaca nemestrina) infected with KFDV or AHFV develop viremia that peaks 2 to 4 days following inoculation. Over the course of infection, animals developed lymphocytopenia, thrombocytopenia, and elevated liver enzymes. Infected animals exhibited hallmark signs of human disease characterized by a flushed appearance, piloerection, dehydration, loss of appetite, weakness, and hemorrhagic signs including epistaxis. Virus was commonly present in the gastrointestinal tract, consistent with human disease caused by KFDV and AHFV where gastrointestinal symptoms (hemorrhage, vomiting, diarrhea) are common. Importantly, RNAseq of whole blood revealed that KFDV downregulated gene expression of key clotting factors that was not observed during AHFV infection, consistent with increased severity of KFDV disease observed in this model. This work characterizes a nonhuman primate model for KFDV and AHFV that closely resembles human disease for further utilization in understanding host immunity and development of antiviral countermeasures.
Assuntos
Modelos Animais de Doenças , Vírus da Encefalite Transmitidos por Carrapatos/patogenicidade , Encefalite Transmitida por Carrapatos/virologia , Febres Hemorrágicas Virais/virologia , Macaca nemestrina , Animais , Chlorocebus aethiops , Citocinas/sangue , Vírus da Encefalite Transmitidos por Carrapatos/genética , Vírus da Encefalite Transmitidos por Carrapatos/imunologia , Encefalite Transmitida por Carrapatos/imunologia , Encefalite Transmitida por Carrapatos/patologia , Feminino , Células HEK293 , Febres Hemorrágicas Virais/imunologia , Febres Hemorrágicas Virais/patologia , Humanos , Linfonodos/virologia , Células Vero , ViremiaRESUMO
The Tick-borne encephalitis virus (TBEV) causes different disease symptoms varying from asymptomatic infection to severe encephalitis and meningitis suggesting a crucial role of the human host immune system in determining the fate of the infection. There is a need to understand the mechanisms underpinning TBEV-host interactions leading to protective immunity. To this aim, we studied the response of human peripheral blood mononuclear cells (PBMC) to the whole formaldehyde inactivated TBEV (I-TBEV), the drug substance of Encepur, one of the five commercially available vaccine. Immunophenotyping, transcriptome and cytokine profiling of PBMC revealed that I-TBEV generates differentiation of a sub-population of plasmacytoid dendritic cells (pDC) that is specialized in type I interferon (IFN) production. In contrast, likely due to the presence of aluminum hydroxide, Encepur vaccine was a poor pDC stimulus. We demonstrated I-TBEV-induced type I IFN together with Interleukin 6 and BAFF to be critical for B cell differentiation to plasmablasts as measured by immunophenotyping and immunoglobulin production. Robust type I IFN secretion was induced by pDC with the concerted action of both viral E glycoprotein and RNA mirroring previous data on dual stimulation of pDC by both S. aureus and influenza virus protein and nucleic acid that leads to a type I IFN-mediated sustained immune response. E glycoprotein neutralization or high temperature denaturation and inhibition of Toll-like receptor 7 signalling confirmed the importance of preserving the functional integrity of these key viral molecules during the inactivation procedure and manufacturing process to produce a vaccine able to stimulate strong immune responses.
Assuntos
Células Dendríticas/imunologia , Vírus da Encefalite Transmitidos por Carrapatos/imunologia , Encefalite Transmitida por Carrapatos/prevenção & controle , Interações entre Hospedeiro e Microrganismos , Interferon Tipo I/metabolismo , Vacinas Virais/imunologia , Antivirais/imunologia , Diferenciação Celular , Quimiocinas/metabolismo , Citocinas/metabolismo , Células Dendríticas/virologia , Encefalite Transmitida por Carrapatos/virologia , Humanos , Leucócitos Mononucleares/imunologia , Ativação Linfocitária , RNA Viral/genética , Proteínas do Envelope Viral/genética , Proteínas do Envelope Viral/metabolismoRESUMO
Tick-borne encephalitis (TBE) is an emerging zoonosis that may cause long-term neurological sequelae or even death. Thus, there is a growing interest in understanding the factors of TBE pathogenesis. Viral genetic determinants may greatly affect the severity and consequences of TBE. In this study, nonstructural protein 1 (NS1) of the tick-borne encephalitis virus (TBEV) was tested as such a determinant. NS1s of three strains with similar neuroinvasiveness belonging to the European, Siberian and Far-Eastern subtypes of TBEV were studied. Transfection of mouse cells with plasmids encoding NS1 of the three TBEV subtypes led to different levels of NS1 protein accumulation in and secretion from the cells. NS1s of TBEV were able to trigger cytokine production either in isolated mouse splenocytes or in mice after delivery of NS1 encoding plasmids. The profile and dynamics of TNF-α, IL-6, IL-10 and IFN-γ differed between the strains. These results demonstrated the involvement of TBEV NS1 in triggering an immune response and indicated the diversity of NS1 as one of the genetic factors of TBEV pathogenicity.
Assuntos
Vírus da Encefalite Transmitidos por Carrapatos , Encefalite Transmitida por Carrapatos , Proteínas não Estruturais Virais , Animais , Camundongos , Vírus da Encefalite Transmitidos por Carrapatos/classificação , Vírus da Encefalite Transmitidos por Carrapatos/fisiologia , Encefalite Transmitida por Carrapatos/imunologia , Encefalite Transmitida por Carrapatos/virologia , Interleucina-10/genética , Zoonoses , Proteínas não Estruturais Virais/metabolismoRESUMO
Tick-borne encephalitis virus (TBEV), a major tick-borne viral pathogen of humans, is known to cause neurological diseases such as meningitis, encephalitis, and meningoencephalitis. However, the life cycle and pathogenesis of TBEV are not well understood. Here, we show that the knockdown or knockout of ADAM15 (a disintegrin and metalloproteinase 15), a host protein involved in neuroblastoma diseases, leads to TBEV replication and assembly defects. We characterized the disintegrin domain in ADAM15 and found that the ADAM15 subcellular localization was changed following TBEV infection. RNA interference (RNAi) screen analysis confirmed ADAM's nonredundant functions and identified a specific role for ADAM15 in TBEV infection. An RNA-sequencing analysis was also conducted to understand the causal link between TBEV infection and the cellular endomembrane network, namely, the generation of replication organelles promoting viral genome replication and virus production. Our data demonstrated that TBEV infection changes ADAM15 cellular localization, which contributes to membrane reorganization and viral replication.IMPORTANCE Tick populations are increasing, and their geographic ranges are expanding. Increases in tick-borne disease prevalence and transmission are important public health issues. Tick-borne encephalitis virus (TBEV) often results in meningitis, encephalitis, and meningoencephalitis. TBEV causes clinical disease in more than 20,000 humans in Europe and Asia per year. An increased incidence of TBE has been noted in Europe and Asia, as a consequence of climate and socioeconomic changes. The need to investigate the mechanism(s) of interaction between the virus and the host factors is apparent, as it will help us to understand the roles of host factors in the life cycle of TBEV. The significance of our research is in identifying the ADAM15 for TBEV replication, which will greatly enhance our understanding of TBEV life cycle and highlight a target for pharmaceutical consideration.
Assuntos
Proteínas ADAM/fisiologia , Vírus da Encefalite Transmitidos por Carrapatos/fisiologia , Encefalite Transmitida por Carrapatos/virologia , Interações entre Hospedeiro e Microrganismos , Proteínas de Membrana/fisiologia , Animais , Chlorocebus aethiops , Cricetinae , Células HEK293 , Humanos , Células Vero , Replicação ViralRESUMO
Tick-borne encephalitis (TBE) is a viral infection of the CNS caused by TBE virus. With no specific treatment available, the only protection is a formalin-inactivated whole virus vaccine. Primary immunization with European TBE vaccines, as recommended by the manufacturers, consists of three vaccine doses administered within a 1-y period. Protection from vaccination is believed to be mediated by Abs, yet T cells may also have a protective role. We set out to characterize the human CD4+ T cell response throughout primary TBE immunization. The responses were evaluated before vaccination and 1 mo after each vaccine dose. A heterogeneous magnitude of CD4+ T cell-mediated memory responses was observed in regard to lymphoblast expansion and cytokine production (IFN-γ, IL-2, and TNF), with the highest median magnitude detected after the second dose of vaccine. Stimulation with an overlapping peptide library based on structural TBE virus proteins E and C revealed that CD4+ T cells concomitantly producing IL-2 and TNF dominated the responses from vaccinees after each vaccine dose, whereas a control cohort of TBE patients responded mainly with all three cytokines. CD107a expression was not upregulated upon peptide stimulation in the vaccinees. However, CD154 (CD40L) expression on cytokine-positive memory CD4+ T cells significantly increased after the second vaccine dose. Taken together, TBE vaccination induced CD4+ T cell responses dominated by IL-2 and TNF production together with CD154 upregulation and a lower IFN-γ response compared with TBE patients. This response pattern was consistent after all three doses of TBE vaccine.
Assuntos
Linfócitos T CD4-Positivos/imunologia , Vírus da Encefalite Transmitidos por Carrapatos/imunologia , Encefalite Transmitida por Carrapatos/prevenção & controle , Imunogenicidade da Vacina , Memória Imunológica , Vacinas Virais/imunologia , Adulto , Linfócitos T CD4-Positivos/metabolismo , Ligante de CD40/imunologia , Ligante de CD40/metabolismo , Estudos de Casos e Controles , Encefalite Transmitida por Carrapatos/sangue , Encefalite Transmitida por Carrapatos/imunologia , Encefalite Transmitida por Carrapatos/virologia , Feminino , Humanos , Esquemas de Imunização , Interferon gama/imunologia , Interferon gama/metabolismo , Interleucina-2/imunologia , Interleucina-2/metabolismo , Masculino , Pessoa de Meia-Idade , Fator de Necrose Tumoral alfa/imunologia , Fator de Necrose Tumoral alfa/metabolismo , Regulação para Cima/imunologia , Vacinas de Produtos Inativados/administração & dosagem , Vacinas de Produtos Inativados/imunologia , Vacinas Virais/administração & dosagem , Adulto JovemRESUMO
Tick-borne encephalitis virus (TBEV), a member of the genus Flavivirus, is common in Europe and Asia and causes a severe disease of the central nervous system. A promising approach in the development of therapy for TBEV infection is the search for small molecule antivirals targeting the flavivirus envelope protein E, particularly its ß-n-octyl-d-glucoside binding pocket (ß-OG pocket). However, experimental studies of candidate antivirals may be complicated by varying amounts and different forms of the protein E in the virus samples. Viral particles with different conformations and arrangements of the protein E are produced during the replication cycle of flaviviruses, including mature, partially mature, and immature forms, as well as subviral particles lacking genomic RNA. The immature forms are known to be abundant in the viral population. We obtained immature virion preparations of TBEV, characterized them by RT-qPCR, and assessed in vivo and in vitro infectivity of the residual mature virions in the immature virus samples. Analysis of the ß-OG pocket structure on the immature virions confirmed the possibility of binding of adamantylmethyl esters of 5-aminoisoxazole-3-carboxylic acid in the pocket. We demonstrated that the antiviral activity of these compounds in plaque reduction assay is significantly reduced in the presence of immature TBEV particles.
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Adamantano/farmacologia , Antivirais/farmacologia , Vírus da Encefalite Transmitidos por Carrapatos/efeitos dos fármacos , Vírus da Encefalite Transmitidos por Carrapatos/fisiologia , Encefalite Transmitida por Carrapatos/virologia , Isoxazóis/farmacologia , Vírion/fisiologia , Adamantano/metabolismo , Animais , Antivirais/metabolismo , Linhagem Celular , Vírus da Encefalite Transmitidos por Carrapatos/crescimento & desenvolvimento , Vírus da Encefalite Transmitidos por Carrapatos/patogenicidade , Glucosídeos/metabolismo , Isoxazóis/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Simulação de Acoplamento Molecular , Ligação Proteica , Conformação Proteica , Suínos , Proteínas do Envelope Viral/química , Proteínas do Envelope Viral/metabolismo , Ensaio de Placa Viral , Vírion/imunologia , Vírion/patogenicidade , Vírion/ultraestruturaRESUMO
Tick-borne encephalitis (TBE) is a relatively severe and clinically variable central nervous system (CNS) disease with a significant contribution of a secondary immunopathology. Monocytes/macrophages play an important role in the CNS inflammation, but their pathogenetic role and migration mechanisms in flavivirus encephalitis in humans are not well known. We have retrospectively analyzed blood and cerebrospinal fluid (CSF) monocyte counts in 240 patients with TBE presenting as meningitis (n = 110), meningoencephalitis (n = 114), or meningoencephalomyelitis (n = 16), searching for associations with other laboratory parameters, clinical presentation, and severity. We have measured concentrations of selected monocytes-attracting chemokines (CCL7, CXCL12, CCL20) in serum and CSF of the prospectively recruited patients with TBE (n = 15), with non-TBE aseptic meningitis (n = 6) and in non-infected controls (n = 8). The data were analyzed with non-parametric tests, p < 0.05 considered significant. Monocyte CSF count correlated with other CSF inflammatory parameters, but not with the peripheral monocytosis, consistent with an active recruitment into CNS. The monocyte count did not correlate with a clinical presentation. The median CSF concentration of CCL7 and CXCL12 was increased in TBE, and that of CCL7 was higher in TBE than in non-TBE meningitis. The comparison of serum and CSF concentrations pointed to the intrathecal synthesis of CCL7 and CXCL12, but with no evident concentration gradients toward CSF. In conclusion, the monocytes are recruited into the intrathecal compartment in concert with other leukocyte populations in TBE. CCL7 and CXCL12 have been found upregulated intrathecally but are not likely to be the main monocyte chemoattractants.
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Quimiocina CCL7/genética , Quimiocina CXCL12/genética , Encefalite Transmitida por Carrapatos/genética , Macrófagos/virologia , Meningoencefalite/genética , Monócitos/virologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Barreira Hematoencefálica/metabolismo , Barreira Hematoencefálica/virologia , Estudos de Casos e Controles , Sistema Nervoso Central/imunologia , Sistema Nervoso Central/metabolismo , Sistema Nervoso Central/virologia , Quimiocina CCL20/sangue , Quimiocina CCL20/líquido cefalorraquidiano , Quimiocina CCL20/genética , Quimiocina CCL7/sangue , Quimiocina CCL7/líquido cefalorraquidiano , Quimiocina CXCL12/sangue , Quimiocina CXCL12/líquido cefalorraquidiano , Quimiotaxia/imunologia , Encefalite Transmitida por Carrapatos/sangue , Encefalite Transmitida por Carrapatos/líquido cefalorraquidiano , Encefalite Transmitida por Carrapatos/virologia , Feminino , Regulação da Expressão Gênica , Humanos , Macrófagos/imunologia , Masculino , Meningoencefalite/sangue , Meningoencefalite/líquido cefalorraquidiano , Meningoencefalite/virologia , Pessoa de Meia-Idade , Monócitos/imunologia , Estudos RetrospectivosRESUMO
OBJECTIVES: This review article summarizes what has been published on Alkhumra hemorrhagic fever virus (AHFV), a novel flavivirus that was discovered in Saudi Arabia in 1995. METHODS: PubMed was used to search for studies published from January 1995 to June 2019 using the key words Alkhumra virus, Alkhurma virus, novel flavivirus, and tick-borne encephalitis virus. Additionally, records of the Saudi Ministry of Health were reviewed. RESULTS: Thirty-two articles on AHFV were identified. Acute febrile flu-like illness, hepatitis, hemorrhagic manifestations, and, less commonly, encephalitis are the main clinical features. The virus seems to be transmitted from livestock animals to humans by direct contact with these animals or their raw meat, or perhaps by tick or mosquito bites. The ability of ticks and mosquitoes to serve as vectors for AHFV needs to be confirmed by biological studies. The exact role of animals such as sheep, goats, camels, and other mammals in the transmission and maintenance of the virus remains to be elucidated. Preventive measures require an interdisciplinary approach involving the human and veterinary health sectors, the municipality, the ministry of agriculture, the vector control sector, and academic and research institutes. CONCLUSIONS: AHFV has been well characterized; nevertheless, some aspects remain to be elucidated.
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Vírus da Encefalite Transmitidos por Carrapatos , Encefalite Transmitida por Carrapatos , Animais , Anticorpos Antivirais , Bases de Dados Factuais , Vírus da Encefalite Transmitidos por Carrapatos/classificação , Encefalite Transmitida por Carrapatos/diagnóstico , Encefalite Transmitida por Carrapatos/epidemiologia , Encefalite Transmitida por Carrapatos/transmissão , Encefalite Transmitida por Carrapatos/virologia , Humanos , Filogenia , Arábia Saudita , Doenças Transmitidas por Carrapatos/prevenção & controle , Doenças Transmitidas por Carrapatos/transmissão , Doenças Transmitidas por Vetores/prevenção & controle , Doenças Transmitidas por Vetores/transmissãoRESUMO
Tick-borne encephalitis (TBE) is an acute disease caused by the tick-borne encephalitis virus. Due to the viral nature of the condition, there is no effective causal treatment for full-blown disease. Current and nonspecific TBE treatments only relieve symptoms. Unfortunately, the first phase of TBE is characterized by flu-like symptoms, making diagnosis difficult during this period. The second phase is referred to as the neurological phase as it involves structures in the central nervous system-most commonly the meninges and, in more severe cases, the brain and the spinal cord. Therefore, it is important that early markers of TBE that will guide clinical decision-making and the choice of treatment are established. In this review, we performed an extensive search of literature reports relevant to biomarkers associated with TBE using the MEDLINE/PubMed database. We observed that apart from routinely determined specific immunoglobulins, free light chains may also be useful in the evaluation of intrathecal synthesis in the central nervous system (CNS) during TBEV infection. Moreover, selected metalloproteinases, chemokines, or cytokines appear to play an important role in the pathogenesis of TBE as a consequence of inflammatory reactions and recruitment of white blood cells into the CNS. Furthermore, we reported promising findings on tau protein or Toll-like receptors. It was also observed that some people may be predisposed to TBE. Therefore, to understand the role of selected tick-borne encephalitis biomarkers, we categorized these factors and discussed their potential application in the diagnosis, prognosis, monitoring, or management of TBE.
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Vírus da Encefalite Transmitidos por Carrapatos , Encefalite Transmitida por Carrapatos/sangue , Encefalite Transmitida por Carrapatos/líquido cefalorraquidiano , Animais , Anticorpos Antivirais/sangue , Anticorpos Antivirais/líquido cefalorraquidiano , Biomarcadores/sangue , Biomarcadores/líquido cefalorraquidiano , Barreira Hematoencefálica/patologia , Quimiocinas/sangue , Quimiocinas/líquido cefalorraquidiano , Encefalite Transmitida por Carrapatos/diagnóstico , Encefalite Transmitida por Carrapatos/genética , Encefalite Transmitida por Carrapatos/virologia , Predisposição Genética para Doença , Humanos , Imunoglobulina G/sangue , Imunoglobulina G/líquido cefalorraquidiano , Imunoglobulina M/sangue , Imunoglobulina M/líquido cefalorraquidiano , Cadeias lambda de Imunoglobulina/sangue , Cadeias lambda de Imunoglobulina/líquido cefalorraquidiano , Metaloproteinase 9 da Matriz/sangue , Metaloproteinase 9 da Matriz/líquido cefalorraquidianoRESUMO
The tick-borne pathogen Powassan virus is a rare cause of encephalitis in North America and the Russian Far East. The number of documented cases described since the discovery of Powassan virus in 1958 may be <150, although detection of cases has increased over the past decade. In the United States, the incidence of Powassan virus infections expanded from the estimated 1 case per year prior to 2005 to 10 cases per year during the subsequent decade. The increased detection rate may be associated with several factors, including enhanced surveillance, the availability of improved laboratory diagnostic methods, the expansion of the vector population, and, perhaps, altered human activities that lead to more exposure. Nonetheless, it remains unclear whether Powassan virus is indeed an emerging threat or if enzootic cycles in nature remain more-or-less stable with periodic fluctuations of host and vector population sizes. Despite the low disease incidence, the approximately 10% to 15% case fatality rate of neuroinvasive Powassan virus infection and the temporary or prolonged sequelae in >50% of survivors make Powassan virus a medical concern requiring the attention of public health authorities and clinicians. The medical importance of Powassan virus justifies more research on developing specific and effective treatments and prevention and control measures.
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Encefalite Transmitida por Carrapatos/epidemiologia , Flavivirus , Doenças Transmitidas por Carrapatos/virologia , Vírus da Encefalite Transmitidos por Carrapatos , Encefalite Transmitida por Carrapatos/diagnóstico , Encefalite Transmitida por Carrapatos/prevenção & controle , Encefalite Transmitida por Carrapatos/virologia , Humanos , Fatores de RiscoRESUMO
Tick-borne encephalitis virus (TBEV) is a zoonotic virus in the genus Flavivirus, family Flaviviridae. TBEV is widely distributed in northern regions of the Eurasian continent, including Japan, and causes severe encephalitis in humans. Tick-borne encephalitis (TBE) was recently reported in central Hokkaido, and wild animals with anti-TBEV antibodies were detected over a wide area of Hokkaido, although TBEV was only isolated in southern Hokkaido. In this study, we conducted a survey of ticks to isolate TBEV in central Hokkaido. One strain, designated Sapporo-17-Io1, was isolated from ticks (Ixodes ovatus) collected in Sapporo city. Sequence analysis revealed that the isolated strain belonged to the Far Eastern subtype of TBEV and was classified in a different subcluster from Oshima 5-10, which had previously been isolated in southern Hokkaido. Sapporo-17-Io1 showed similar growth properties to those of Oshima 5-10 in cultured cells and mouse brains. The mortality rate of mice infected intracerebrally with each virus was similar, but the survival time of mice inoculated with Sapporo-17-Io1 was significantly longer than that of mice inoculated with Oshima 5-10. These results indicate that the neurovirulence of Sapporo-17-Io1 was lower than that of Oshima 5-10. Using an infectious cDNA clone, the replacement of genes encoding non-structural genes from Oshima 5-10 with those from Sapporo-17-Io1 attenuated the neuropathogenicity of the cloned viruses. This result indicated that the non-structural proteins determine the neurovirulence of these two strains. Our results provide important insights for evaluating epidemiological risk in TBE-endemic areas of Hokkaido.