RESUMO
Viral hemorrhagic fevers (HF) are a group of acute febrile diseases with high mortality rates. Although hemostatic dysfunction appears to be a major determinant of the severity of the disease, it is still unclear what pathogenic mechanisms lead to it. In clinical studies it is found that arenaviruses, such as Lassa, Machupo, and Guanarito viruses cause HF that vary in symptoms and biological alterations. In this study we aimed to characterize the hemostatic dysfunction induced by arenaviral HF to determine its implication in the severity of the disease and to elucidate the origin of this syndrome. We found that lethal infection with Machupo, Guanarito, and Lassa viruses is associated with cutaneomucosal, cerebral, digestive, and pulmonary hemorrhages. The affected animals developed a severe alteration of the coagulation system, which was concomitant with acute hepatitis, minor deficit of hepatic factor synthesis, presence of a plasmatic inhibitor of coagulation, and dysfunction of the fibrinolytic system. Despite signs of increased vascular permeability, endothelial cell infection was not a determinant factor of the hemorrhagic syndrome. There were also alterations of the primary hemostasis during lethal infection, with moderate to severe thrombocytopenia and platelet dysfunction. Finally, we show that lethal infection is accompanied by a reduced hematopoietic potential of the bone marrow. This study provides an unprecedented characterization of the hemostasis defects induced by several highly pathogenic arenaviruses.
Assuntos
Arenaviridae , Arenavirus , Febres Hemorrágicas Virais , Hemostáticos , Animais , Febres Hemorrágicas Virais/patologia , Hemorragia/etiologia , Hemostasia , MacacaRESUMO
BACKGROUND: Lassa fever is a substantial health burden in west Africa. We evaluated the safety, tolerability, and immunogenicity of a recombinant, live-attenuated, measles-vectored Lassa fever vaccine candidate (MV-LASV). METHODS: This first-in-human phase 1 trial-consisting of an open-label dose-escalation stage and an observer-blinded, randomised, placebo-controlled treatment stage-was conducted at a single site at the University of Antwerp, Antwerp, Belgium, and involved healthy adults aged 18-55 years. Participants in the dose-escalation stage were sequentially assigned to a low-dose group (two intramuscular doses of MV-LASV at 2 × 104 times the median tissue culture infectious dose) or a high-dose group (two doses at 1 × 105 times the median tissue culture infectious dose). Participants in the double-blinded treatment stage were randomly assigned in a 2:2:1 ratio to receive low dose, high dose, or placebo. The primary endpoint was the rate of solicited and unsolicited adverse events up to study day 56 and was assessed in all participants who received at least one dose of investigational product. The trial is registered with ClinicalTrials.gov, NCT04055454, and the European Union Drug Regulating Authorities Clinical Trials Database, 2018-003647-40, and is complete. FINDINGS: Between Sept 26, 2019, and Jan 20, 2020, 60 participants were enrolled and assigned to receive placebo (n=12) or MV-LASV (n=48). All 60 participants received at least one study treatment. Most adverse events occurred during the treatment phase, and frequencies of total solicited or unsolicited adverse events were similar between treatment groups, with 96% of participants in the low-dose group, 100% of those in the high-dose group, and 92% of those in the placebo group having any solicited adverse event (p=0·6751) and 76% of those in the low-dose group, 70% of those in the high-dose group, and 100% of those in the placebo group having any unsolicited adverse event (p=0·1047). The only significant difference related to local solicited adverse events, with higher frequencies observed in groups receiving MV-LASV (24 [96%] of 25 participants in the low-dose group; all 23 [100%] participants in the high-dose group) than in the placebo group (6 [50%] of 12 participants; p=0·0001, Fisher-Freeman-Halton test). Adverse events were mostly of mild or moderate severity, and no serious adverse events were observed. MV-LASV also induced substantial concentrations of LASV-specific IgG (geometric mean titre 62·9 EU/ml in the low-dose group and 145·9 EU/ml in the high-dose group on day 42). INTERPRETATION: MV-LASV showed an acceptable safety and tolerability profile, and immunogenicity seemed to be unaffected by pre-existing immunity against the vector. MV-LASV is therefore a promising candidate for further development. FUNDING: Coalition for Epidemic Preparedness Innovations.
Assuntos
Febre Lassa , Sarampo , Adulto , Humanos , Vacina contra Sarampo , Vacinas Sintéticas , Vacinas Atenuadas , Método Duplo-Cego , Anticorpos AntiviraisRESUMO
Lassa virus (LASV) is responsible for a viral hemorrhagic fever in humans and the death of 3,000 to 5,000 people every year. The immune response to LASV is poorly understood, but type I interferon (IFN-I) and T-cell responses appear to be critical for the host. We studied the response of myeloid dendritic cells (mDC) to LASV, as mDCs are involved in both IFN-I production and T-cell activation. We compared the response of primary human mDCs to LASV and Mopeia virus (MOPV), which is similar to LASV, but non-pathogenic. We showed that mDCs produced substantial amounts of IFN-I in response to both LASV and MOPV. However, only MOPV-infected mDCs were able to activate T cells. More surprisingly, coculture with T cells completely inhibited the activation of LASV-infected mDCs. These differences between LASV and MOPV were mostly due to the LASV nucleoprotein, which has major immunosuppressive properties, but the glycoprotein was also involved. Overall, these results suggest that mDCs may be important for the global response to LASV and play a role in the outcome of Lassa fever.
Assuntos
Células Dendríticas/imunologia , Vírus Lassa/imunologia , Células Mieloides/imunologia , Antivirais , Arenaviridae/imunologia , Células Dendríticas/virologia , Voluntários Saudáveis , Febres Hemorrágicas Virais/virologia , Humanos , Interferon Tipo I , Interferon-alfa/metabolismo , Interferon beta/metabolismo , Febre Lassa/virologia , Vírus Lassa/patogenicidade , Ativação Linfocitária/imunologia , Ativação Linfocitária/fisiologia , Células Mieloides/virologia , Nucleoproteínas/metabolismo , Cultura Primária de Células , Linfócitos T/imunologiaRESUMO
Several Old World and New World arenaviruses are responsible for severe endemic and epidemic hemorrhagic fevers, whereas other members of the Arenaviridae family are nonpathogenic. To date, no approved vaccines, antivirals, or specific treatments are available, except for Junín virus. However, protection of nonhuman primates against Lassa fever virus (LASV) is possible through the inoculation of the closely related but nonpathogenic Mopeia virus (MOPV) before challenge with LASV. We reasoned that this virus, modified by using reverse genetics, would represent the basis for the generation of a vaccine platform against LASV and other pathogenic arenaviruses. After showing evidence of exoribonuclease (ExoN) activity in NP of MOPV, we found that this activity was essential for multiplication in antigen-presenting cells. The introduction of multiple mutations in the ExoN site of MOPV NP generated a hyperattenuated strain (MOPVExoN6b) that is (i) genetically stable over passages, (ii) has increased immunogenic properties compared to those of MOPV, and (iii) still promotes a strong type I interferon (IFN) response. MOPVExoN6b was further modified to harbor the envelope glycoproteins of heterologous pathogenic arenaviruses, such as LASV or Lujo, Machupo, Guanarito, Chapare, or Sabia virus in order to broaden specific antigenicity while preserving the hyperattenuated characteristics of the parental strain. Our MOPV-based vaccine candidate for LASV, MOPEVACLASV, was used in a one-shot immunization assay in nonhuman primates and fully protected them from a lethal challenge with LASV. Thus, our hyperattenuated strain of MOPV constitutes a promising new live-attenuated vaccine platform to immunize against several, if not all, pathogenic arenaviruses.IMPORTANCE Arenaviruses are emerging pathogens transmitted to humans by rodents and responsible for endemic and epidemic hemorrhagic fevers of global concern. Nonspecific symptoms associated with the onset of infection make these viruses difficult to distinguish from other endemic pathogens. Moreover, the unavailability of rapid diagnosis in the field delays the identification of the virus and early care for treatment and favors spreading. The vaccination of exposed populations would be of great help to decrease morbidity and human-to-human transmission. Using reverse genetics, we generated a vaccine platform for pathogenic arenaviruses based on a modified and hyperattenuated strain of the nonpathogenic Mopeia virus and showed that the Lassa virus candidate fully protected nonhuman primates from a lethal challenge. These results showed that a rationally designed recombinant MOPV-based vaccine is safe, immunogenic, and efficacious in nonhuman primates.
Assuntos
Arenaviridae/imunologia , Febres Hemorrágicas Virais/imunologia , Febre Lassa/imunologia , Vírus Lassa/imunologia , Doenças dos Macacos/imunologia , Doenças dos Macacos/prevenção & controle , Vacinas Atenuadas/imunologia , Vacinas Virais/imunologia , Animais , Arenaviridae/genética , Linhagem Celular , Chlorocebus aethiops , Cricetinae , Exorribonucleases/metabolismo , Células HEK293 , Febres Hemorrágicas Virais/patologia , Febres Hemorrágicas Virais/transmissão , Febres Hemorrágicas Virais/virologia , Humanos , Interferon Tipo I/imunologia , Febre Lassa/prevenção & controle , Febre Lassa/virologia , Macaca fascicularis , Doenças dos Macacos/virologia , Vacinação , Células VeroRESUMO
IFITMs are broad antiviral factors that block incoming virions in endosomal vesicles, protecting target cells from infection. In the case of HIV-1, we and others reported the existence of an additional antiviral mechanism through which IFITMs lead to the production of virions of reduced infectivity. However, whether this second mechanism of inhibition is unique to HIV or extends to other viruses is currently unknown. To address this question, we have analyzed the susceptibility of a broad spectrum of viruses to the negative imprinting of the virion particles infectivity by IFITMs. The results we have gathered indicate that this second antiviral property of IFITMs extends well beyond HIV and we were able to identify viruses susceptible to the three IFITMs altogether (HIV-1, SIV, MLV, MPMV, VSV, MeV, EBOV, WNV), as well as viruses that displayed a member-specific susceptibility (EBV, DUGV), or were resistant to all IFITMs (HCV, RVFV, MOPV, AAV). The swapping of genetic elements between resistant and susceptible viruses allowed us to point to specificities in the viral mode of assembly, rather than glycoproteins as dominant factors of susceptibility. However, we also show that, contrarily to X4-, R5-tropic HIV-1 envelopes confer resistance against IFITM3, suggesting that viral receptors add an additional layer of complexity in the IFITMs-HIV interplay. Lastly, we show that the overall antiviral effects ascribed to IFITMs during spreading infections, are the result of a bimodal inhibition in which IFITMs act both by protecting target cells from incoming viruses and in driving the production of virions of reduced infectivity. Overall, our study reports for the first time that the negative imprinting of the virion particles infectivity is a conserved antiviral property of IFITMs and establishes IFITMs as a paradigm of restriction factor capable of interfering with two distinct phases of a virus life cycle.
Assuntos
Antígenos de Diferenciação/metabolismo , Vírion , Replicação Viral , Linhagem Celular , HIV-1/fisiologia , Interações Hospedeiro-Patógeno , Humanos , Internalização do VírusRESUMO
UNLABELLED: Lassa virus is an Old World Arenavirus which causes Lassa hemorrhagic fever in humans, mostly in West Africa. Lassa fever is an important public health problem, and a safe and effective vaccine is urgently needed. The infection causes immunosuppression, probably due to the absence of activation of antigen-presenting cells (dendritic cells and macrophages), low type I interferon (IFN) production, and deficient NK cell function. However, a recombinant Lassa virus carrying D389A and G392A substitutions in the nucleoprotein that abolish the exonuclease activity and IFN activation loses its inhibitory activity and induces strong type I IFN production by dendritic cells and macrophages. We show here that during infection by this mutant Lassa virus, antigen-presenting cells trigger efficient human NK cell responses in vitro, including production of IFN-γ and cytotoxicity. NK cell activation involves close contact with both antigen-presenting cells and soluble factors. We report that infected dendritic cells and macrophages express the NKG2D ligands major histocompatibility complex (MHC) class I-related chains A and B and that they may produce interleukin-12 (IL-12), IL-15, and IL-18, all involved in NK cell functions. NK cell degranulation is significantly increased in cocultures, suggesting that NK cells seem to kill infected dendritic cells and macrophages. This work confirms the inhibitory function of Lassa virus nucleoprotein. Importantly, we demonstrate for the first time that Lassa virus nucleoprotein is involved in the inhibition of antigen-presenting cell-mediated NK cell responses. IMPORTANCE: The pathogenesis and immune responses induced by Lassa virus are poorly known. Recently, an exonuclease domain contained in the viral nucleoprotein has been shown to be able to inhibit the type I IFN response by avoiding the recognition of viral RNA by cell sensors. Here, we studied the responses of NK cells to dendritic cells and macrophages infected with a recombinant Lassa virus in which the exonuclease functions have been abolished and demonstrated that NK cells are strongly activated and presented effective functions. These results show that the strategy developed by Lassa virus to evade innate immunity is also effective on NK cells, explaining the weak NK cell activation observed with the wild-type virus. By providing a better understanding of the interactions between Lassa virus and the host immune system, these results are important for the field of arenavirus biology and may be useful for a vaccine approach against Lassa fever.
Assuntos
Células Apresentadoras de Antígenos/imunologia , Proteínas do Capsídeo/imunologia , Exonucleases/imunologia , Células Matadoras Naturais/imunologia , Vírus Lassa/imunologia , Animais , Proteínas do Capsídeo/genética , Degranulação Celular , Sobrevivência Celular , Células Cultivadas , Técnicas de Cocultura , Células Dendríticas/imunologia , Exonucleases/genética , Humanos , Tolerância Imunológica , Interferon gama/metabolismo , Interleucinas/metabolismo , Macrófagos/imunologia , Proteínas Mutantes/genética , Proteínas Mutantes/imunologia , Mutação de Sentido IncorretoRESUMO
Lassa virus (LASV), which causes a viral hemorrhagic fever, inhibits the innate immune response. The exonuclease (ExoN) domain of its nucleoprotein (NP) is implicated in the suppression of retinoic acid-inducible gene I (RIG-I) signaling. We show here that a LASV in which ExoN function has been abolished strongly activates innate immunity and that this effect is dependent on RIG-I signaling. These results highlight the key role of NP ExoN function in the immune evasion that occurs during LASV infection.
Assuntos
Exonucleases/imunologia , Tolerância Imunológica , Imunidade Inata , Vírus Lassa/imunologia , Vírus Lassa/fisiologia , Nucleoproteínas/imunologia , Transdução de Sinais , Células Cultivadas , Proteína DEAD-box 58 , RNA Helicases DEAD-box/metabolismo , Exonucleases/metabolismo , Humanos , Nucleoproteínas/metabolismo , Receptores ImunológicosRESUMO
Nipah virus (NiV) has been recently ranked by the World Health Organization as being among the top eight emerging pathogens likely to cause major epidemics, whereas no therapeutics or vaccines have yet been approved. We report a method to deliver immunogenic epitopes from NiV through the targeting of the CD40 receptor of antigen-presenting cells by fusing a selected humanized anti-CD40 monoclonal antibody to the Nipah glycoprotein with conserved NiV fusion and nucleocapsid peptides. In the African green monkey model, CD40.NiV induces specific immunoglobulin A (IgA) and IgG as well as cross-neutralizing responses against circulating NiV strains and Hendra virus and T cell responses. Challenge experiments using a NiV-B strain demonstrate the high protective efficacy of the vaccine, with all vaccinated animals surviving and showing no significant clinical signs or virus replication, suggesting that the CD40.NiV vaccine conferred sterilizing immunity. Overall, results obtained with the CD40.NiV vaccine are highly promising in terms of the breadth and efficacy against NiV.
Assuntos
Vacinas Virais , Animais , Chlorocebus aethiops , Linfócitos T , Formação de Anticorpos , Células Apresentadoras de Antígenos , Replicação ViralRESUMO
Lassa virus (LASV) and Mopeia virus (MOPV) are closely related Arenaviruses. LASV causes hemorrhagic fever, whereas MOPV is not pathogenic. Both viruses display tropism for APCs such as DCs and macrophages. During viral infections, NK cells are involved in the clearance of infected cells and promote optimal immune responses by interacting with APCs. We used an in vitro model of human NK and APC coculture to study the role of NK cells and to characterize their interactions with APCs during LASV and MOPV infections. As expected, NK cells alone were neither infected nor activated by LASV and MOPV, and infected DCs did not activate NK cells. By contrast, LASV- and MOPV-infected macrophages activated NK cells, as shown by the upregulation of CD69, NKp30, and NKp44, the downregulation of CXCR3, and an increase in NK-cell proliferation. NK cells acquired enhanced cytotoxicity, as illustrated by the increase in granzyme B (GrzB) expression and killing of K562 targets, but did not produce IFN-γ. Contact between NK cells and infected macrophages and type I IFNs were essential for activation; however, NK cells could not kill infected cells and control infection. Overall, these findings show that MOPV- as well as pathogenic LASV-infected macrophages mediate NK-cell activation.
Assuntos
Células Matadoras Naturais/imunologia , Febre Lassa/imunologia , Vírus Lassa/imunologia , Macrófagos/imunologia , Animais , Processos de Crescimento Celular/imunologia , Chlorocebus aethiops , Técnicas de Cocultura , Proteína Ligante Fas/genética , Proteína Ligante Fas/imunologia , Regulação Viral da Expressão Gênica , Granzimas/genética , Granzimas/imunologia , Humanos , Interferon gama/genética , Interferon gama/imunologia , Células K562 , Células Matadoras Naturais/virologia , Febre Lassa/virologia , Ativação Linfocitária/imunologia , Macrófagos/virologia , RNA/química , RNA/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Estatísticas não Paramétricas , Ligante Indutor de Apoptose Relacionado a TNF/genética , Ligante Indutor de Apoptose Relacionado a TNF/imunologia , Células VeroRESUMO
Pathogenic New World arenaviruses (NWAs) cause haemorrhagic fevers and can have high mortality rates, as shown in outbreaks in South America. Neutralizing antibodies (Abs) are critical for protection from NWAs. Having shown that the MOPEVAC vaccine, based on a hyperattenuated arenavirus, induces neutralizing Abs against Lassa fever, we hypothesized that expression of NWA glycoproteins in this platform might protect against NWAs. Cynomolgus monkeys immunized with MOPEVACMAC, targeting Machupo virus, prevented the lethality of this virus and induced partially NWA cross-reactive neutralizing Abs. We then developed the pentavalent MOPEVACNEW vaccine, expressing glycoproteins from all pathogenic South American NWAs. Immunization of cynomolgus monkeys with MOPEVACNEW induced neutralizing Abs against five NWAs, strong innate followed by adaptive immune responses as detected by transcriptomics and provided sterile protection against Machupo virus and the genetically distant Guanarito virus. MOPEVACNEW may thus be efficient to protect against existing and potentially emerging NWAs.
Assuntos
Arenavirus do Novo Mundo , Animais , Arenavirus do Novo Mundo/metabolismo , Vacinas Combinadas , Macaca fascicularis/metabolismo , Anticorpos Neutralizantes , GlicoproteínasRESUMO
Lassa fever hits West African countries annually in the absence of licensed vaccine to limit the burden of this viral hemorrhagic fever. We previously developed MeV-NP, a single-shot vaccine protecting cynomolgus monkeys against divergent strains one month or more than a year before Lassa virus infection. Given the limited dissemination area during outbreaks and the risk of nosocomial transmission, a vaccine inducing rapid protection could be useful to protect exposed people during outbreaks in the absence of preventive vaccination. Here, we test whether the time to protection can be reduced after immunization by challenging measles virus pre-immune male cynomolgus monkeys sixteen or eight days after a single shot of MeV-NP. None of the immunized monkeys develop disease and they rapidly control viral replication. Animals immunized eight days before the challenge are the best controllers, producing a strong CD8 T-cell response against the viral glycoprotein. A group of animals was also vaccinated one hour after the challenge, but was not protected and succumbed to the disease as the control animals. This study demonstrates that MeV-NP can induce a rapid protective immune response against Lassa fever in the presence of MeV pre-existing immunity but can likely not be used as therapeutic vaccine.
Assuntos
Febre Lassa , Febre Lassa/imunologia , Febre Lassa/prevenção & controle , Vírus Lassa/imunologia , Masculino , Animais , Macaca fascicularis , Vacinas Virais/administração & dosagem , Vacinas Virais/imunologia , Nucleoproteínas/imunologia , Imunidade Humoral , Replicação Viral , Linfócitos T/imunologia , Células Matadoras Naturais/imunologia , TranscriptomaRESUMO
The events leading to death in severe cases of Lassa fever (LF) are unknown. Fatality seems to be linked to high viremia and immunosuppression, and cellular immunity, rather than neutralizing antibodies, appears to be essential for survival. We previously compared Lassa virus (LV) with its genetically close but nonpathogenic homolog Mopeia virus (MV), which was used to model nonfatal LF. We showed that strong and early activation of antigen-presenting cells (APC) may play a crucial role in controlling infection. Here we developed an in vitro model of dendritic-cell (DC)-T-cell coculture in order to characterize human T-cell responses induced by MV- or LV-infected DCs. Our results show very different responses to infection with LV and MV. MV strongly and durably stimulated CD8(+) and CD4(+) T cells, showing early and high activation, a strong proliferative response, and acquisition of effector and memory phenotypes. Furthermore, robust and functional CD4(+) and CD8(+) cytotoxic T lymphocytes (CTL) were generated. LV, however, induced only weak memory responses. Thus, this study allows an improved understanding of the pathogenesis and immune mechanisms involved in the control of human LV.
Assuntos
Arenavirus do Velho Mundo/imunologia , Linfócitos T CD4-Positivos/imunologia , Células Dendríticas/virologia , Linfócitos T Citotóxicos/imunologia , Proliferação de Células , Células Cultivadas , Técnicas de Cocultura , Citocinas/biossíntese , Células Dendríticas/imunologia , Ensaio de Imunoadsorção Enzimática , ELISPOT , Citometria de Fluxo , Humanos , Memória Imunológica , Vírus Lassa/imunologia , Ativação Linfocitária , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Lassa virus (LASV; Arenaviridae) is responsible for severe hemorrhagic fevers in Africa. LASV nucleoprotein (NP) plays important roles in regulating viral transcription and replication and in inhibiting type I interferon (IFN) production. The NP C-terminal domain contains a 3'-to-5' exonuclease activity involved in suppressing IFN induction. We have established a murine polymerase (Pol) I reverse genetics system for LASV, showing that residues D389 and G392 of NP were critical for LASV viability, while the D389A/G392A and D389T/392A double mutants were severely altered in the ability to suppress IFN in macrophages and dendritic cells. Assessing their attenuation in vivo may open new perspectives in vaccinology.
Assuntos
Proteínas do Capsídeo/genética , Proteínas do Capsídeo/imunologia , Células Dendríticas/imunologia , Interferon Tipo I/metabolismo , Vírus Lassa/genética , Vírus Lassa/imunologia , Macrófagos/imunologia , Substituição de Aminoácidos , Animais , Chlorocebus aethiops , Células Dendríticas/virologia , Vírus Lassa/crescimento & desenvolvimento , Macrófagos/virologia , Dados de Sequência Molecular , Proteínas Mutantes/genética , Proteínas Mutantes/imunologia , Mutação de Sentido Incorreto , Análise de Sequência de DNA , Células Vero , Carga Viral , Ensaio de Placa ViralRESUMO
The area of Lassa virus (LASV) circulation is expanding, with the emergence of highly pathogenic new LASV lineages. Benin recently became an endemic country for LASV and has seen the emergence of a new LASV lineage (VII). The first two outbreaks in 2014 and 2016 showed a relatively high mortality rate compared to other outbreaks. We infected cynomolgus monkeys with two strains belonging to lineage II and lineage VII that were isolated from deceased patients during the 2016 outbreak in Benin. The lineage VII strain (L7) caused uniform mortality. Death was associated with uncontrolled viral replication, unbalanced inflammatory responses characterized by increased concentrations of pro- and anti-inflammatory mediators, and the absence of efficient immune responses, resembling the pathogenesis associated with the prototypic Josiah strain in monkeys. The lineage II strain (L2) showed apparently lower virulence than its counterpart, with a prolonged time to death and a lower mortality rate. Prolonged survival was associated with better control of viral replication, a moderate inflammatory response, and efficient T-cell responses. Transcriptomic analyses also highlighted important differences in the immune responses associated with the outcome. Both strains caused strong inflammation in several organs. Notably, meningitis and encephalitis were observed in the cerebral cortex and cerebellum in all monkeys, independently of the outcome. Due to their apparently high pathogenicity, emerging strains from lineage VII should be considered in preclinical vaccine testing. Lineage II would also be beneficial in pathogenesis studies to study the entire spectrum of Lassa fever severity.
Assuntos
Febre Lassa , Vírus Lassa , Animais , Humanos , Vírus Lassa/genética , Macaca fascicularis , Replicação ViralRESUMO
Ebola virus has been responsible for two major epidemics over the last several years and there has been a strong effort to find potential treatments that can improve the disease outcome. Antiviral favipiravir was thus tested on non-human primates infected with Ebola virus. Half of the treated animals survived the Ebola virus challenge, whereas the infection was fully lethal for the untreated ones. Moreover, the treated animals that did not survive died later than the controls. We evaluated the hematological, virological, biochemical, and immunological parameters of the animals and performed proteomic analysis at various timepoints of the disease. The viral load strongly correlated with dysregulation of the biological functions involved in pathogenesis, notably the inflammatory response, hemostatic functions, and response to stress. Thus, the management of viral replication in Ebola virus disease is of crucial importance in preventing the immunopathogenic disorders and septic-like shock syndrome generally observed in Ebola virus-infected patients.
Assuntos
Amidas/farmacologia , Antivirais/farmacologia , Síndrome da Liberação de Citocina/prevenção & controle , Ebolavirus/efeitos dos fármacos , Doença pelo Vírus Ebola/tratamento farmacológico , Pirazinas/farmacologia , Carga Viral/efeitos dos fármacos , Animais , Citocinas/sangue , Modelos Animais de Doenças , Doença pelo Vírus Ebola/patologia , Doença pelo Vírus Ebola/veterinária , Macaca fascicularis , Linfócitos T/imunologia , Viremia/sangue , Viremia/patologia , Replicação Viral/efeitos dos fármacosRESUMO
A safe and protective Lassa virus vaccine is crucially needed in Western Africa to stem the recurrent outbreaks of Lassa virus infections in Nigeria and the emergence of Lassa virus in previously unaffected countries, such as Benin and Togo. Major challenges in developing a Lassa virus vaccine include the high diversity of circulating strains and their reemergence from 1 year to another. To address each of these challenges, we immunized cynomolgus monkeys with a measles virus vector expressing the Lassa virus glycoprotein and nucleoprotein of the prototypic Lassa virus strain Josiah (MeV-NP). To evaluate vaccine efficacy against heterologous strains of Lassa virus, we challenged the monkeys a month later with heterologous strains from lineage II or lineage VII, finding that the vaccine was protective against these strains. A second cohort of monkeys was challenged 1 year later with the homologous Josiah strain, finding that a single dose of MeV-NP was sufficient to protect all vaccinated monkeys. These studies demonstrate that MeV-NP can generate both long-lasting immune responses and responses that are able to protect against diverse strains of Lassa virus.
Assuntos
Febre Lassa , Vacinas Virais/imunologia , África Ocidental , Animais , Febre Lassa/prevenção & controle , Vírus Lassa , Macaca fascicularis , NucleoproteínasRESUMO
Lassa virus (LASV) is endemic in West Africa and induces a viral hemorrhagic fever (VHF) with up to 30% lethality among clinical cases. The mechanisms involved in control of Lassa fever or, in contrast, the ensuing catastrophic illness and death are poorly understood. We used the cynomolgus monkey model to reproduce the human disease with asymptomatic to mild or fatal disease. After initial replication at the inoculation site, LASV reached the secondary lymphoid organs. LASV did not spread further in nonfatal disease and was rapidly controlled by balanced innate and T-cell responses. Systemic viral dissemination occurred during severe disease. Massive replication, a cytokine/chemokine storm, defective T-cell responses, and multiorgan failure were observed. Clinical, biological, immunological, and transcriptomic parameters resembled those observed during septic-shock syndrome, suggesting that similar pathogenesis is induced during Lassa fever. The outcome appears to be determined early, as differentially expressed genes in PBMCs were associated with fatal and non-fatal Lassa fever outcome very early after infection. These results provide a full characterization and important insights into Lassa fever pathogenesis and could help to develop early diagnostic tools.
Assuntos
Modelos Animais de Doenças , Febre Lassa/imunologia , Febre Lassa/virologia , Macaca fascicularis , Imunidade Adaptativa , Animais , Biomarcadores/metabolismo , Feminino , Imunidade Inata , Febre Lassa/sangue , Febre Lassa/patologia , Pulmão/patologia , Tecido Linfoide/patologia , Masculino , TranscriptomaRESUMO
Lassa virus causes a hemorrhagic fever endemic in West Africa. The pathogenesis and the immune responses associated with the disease are poorly understood, and no vaccine is available. We followed virological, pathological, and immunological markers associated with fatal and nonfatal Lassa virus infection of cynomolgus monkeys. The clinical picture was characterized by fever, weight loss, depression, and acute respiratory syndrome. Transient thrombocytopenia and lymphopenia, lymphadenopathy, splenomegaly, infiltration of mononuclear cells, and alterations of the liver, lungs, and endothelia were observed. Survivors exhibited fewer lesions and a lower viral load than nonsurvivors. Although all animals developed strong humoral responses, antibodies appeared more rapidly in survivors and were directed against GP(1), GP(2), and NP. Type I interferons were detected early after infection in survivors but only during the terminal stages in fatalities. The mRNAs for CXCL10 (IP-10) and CXCL11 (I-TAC) were abundant in peripheral blood mononuclear cells and lymph nodes from infected animals, but plasma interleukin-6 was detected only in fatalities. In survivors, high activated-monocyte counts were followed by a rise in the total number of circulating monocytes. Activated T lymphocytes circulated in survivors, whereas T-cell activation was low and delayed in fatalities. In vitro stimulation with inactivated Lassa virus induced activation of T lymphocytes from all infected monkeys, but only lymphocytes from survivors proliferated. Thus, early and strong immune responses and control of viral replication were associated with recovery, whereas fatal infection was characterized by major alterations of the blood formula and, in organs, weak immune responses and uncontrolled viral replication.
Assuntos
Infecções por Arenaviridae/imunologia , Infecções por Arenaviridae/virologia , Vírus Lassa/imunologia , Macaca fascicularis/imunologia , Macaca fascicularis/virologia , Replicação Viral , Animais , Infecções por Arenaviridae/sangue , Infecções por Arenaviridae/patologia , Chlorocebus aethiops , Citocinas/biossíntese , Citocinas/imunologia , Macaca fascicularis/sangue , Masculino , Fatores de Tempo , Células Vero , Viremia/virologiaRESUMO
Lassa virus (LASV) causes a viral haemorrhagic fever in humans and is a major public health concern in West Africa. An efficient immune response to LASV appears to rely on type I interferon (IFN-I) production and T-cell activation. We evaluated the response of plasmacytoid dendritic cells (pDC) to LASV, as they are an important and early source of IFN-I. We compared the response of primary human pDCs to LASV and Mopeia virus (MOPV), which is very closely related to LASV, but non-pathogenic. We showed that pDCs are not productively infected by either MOPV or LASV, but produce IFN-I. However, the activation of pDCs was more robust in response to MOPV than LASV. In vivo, pDC activation may support the control of viral replication through IFN-I production, but also improve the induction of a global immune response. Therefore, pDC activation could play a role in the control of LASV infection.
Assuntos
Células Dendríticas/virologia , Vírus Lassa/imunologia , Ativação Linfocitária , Replicação Viral/imunologia , Células Cultivadas , Humanos , Interferon Tipo I/imunologiaRESUMO
BACKGROUND: The West African Ebola virus epidemic from 2014-2016 highlighted the lack of knowledge about the pathogenicity of the virus and the factors responsible for outcome. A performant and rapid diagnosis is of crucial importance, as is overcoming the difficulty of providing high-quality patient management during such an extensive outbreak. Here, we propose to study the role of the immune mediators during Ebola virus disease and to define some molecules of importance in the outcome. METHODS: Plasma from Guinean patients sampled during the outbreak were analyzed using RT-qPCR, magnetic bead assay, ELISA, and high-quality statistical analyses. We also performed a transcriptomic analysis in leukocytes samples. Therefore, we deeply characterized the immune responses involved in Ebola virus disease. RESULTS: We evaluated the immune patterns depending on the outcome of the disease. Survivors presented an efficient and well-balanced immune response, whereas fatalities were characterized by an intense inflammatory response, overexpression of multiple cytokines, and a "chemokine storm." The plasma concentration of most of the parameters tested increased until death. Statistical analyses also allowed us to define a panel of markers highly predictive of outcome. CONCLUSION: The immune response observed in fatalities was highly similar to that characterizing septic shock syndrome. Our results suggest that immune responses can play a major pathogenic role during severe Ebola virus infection and argue in favor of therapeutic approaches that act on both viral replication and the induction of shock syndrome. FUNDING: French Ministry of Foreign Affairs, the Agence Française de Développement, and the Institut Pasteur.