RESUMEN
Limited knowledge exists on the quality of polyclonal antibody responses generated following Marburg virus (MARV) infection and its evolution in survivors. In this study, we evaluate MARV proteome-wide antibody repertoire longitudinally in convalescent phase approximately every six months for five years following MARV infection in ten human survivors. Differential kinetics were observed for IgM vs IgG vs IgA epitope diversity, antibody binding, antibody affinity maturation and Fc-receptor interaction to MARV proteins. Durability of MARV-neutralizing antibodies is low in survivors. MARV infection induces a diverse epitope repertoire with predominance against GP, VP40, VP30 and VP24 that persisted up to 5 years post-exposure. However, the IgM and IgA repertoire declines over time. Within MARV-GP, IgG recognize antigenic sites predominantly in the amino-terminus, wing domain and GP2-heptad repeat. Interestingly, MARV infection generates robust durable FcɣRI, FcɣRIIA and FcɣRIIIA IgG-Fc receptor interactions. Immunization with immunodominant MARV epitopes reveals conserved wing region between GP1 and GP2, induces neutralizing antibodies against MARV. These findings demonstrate that MARV infection generates a diverse, long-lasting, non-neutralizing, IgG antibody repertoire that perturbs disease by FcɣR activity. This information, along with discovery of neutralizing immunogen in wing domain, could aid in development of effective therapeutics and vaccines against Marburg virus.
Asunto(s)
Anticuerpos Neutralizantes , Anticuerpos Antivirales , Enfermedad del Virus de Marburg , Marburgvirus , Proteoma , Marburgvirus/inmunología , Humanos , Enfermedad del Virus de Marburg/inmunología , Enfermedad del Virus de Marburg/virología , Anticuerpos Antivirales/inmunología , Anticuerpos Neutralizantes/inmunología , Proteoma/inmunología , Femenino , Vacunas Virales/inmunología , Inmunoglobulina G/inmunología , Masculino , Epítopos/inmunología , Adulto , Inmunoglobulina M/inmunología , Persona de Mediana Edad , Estudios Longitudinales , Inmunoglobulina A/inmunología , Desarrollo de Vacunas , Proteínas del Envoltorio Viral/inmunologíaRESUMEN
INTRODUCTION: In March 2023, a Marburg Virus Disease (MVD) outbreak was declared in Kagera region, Northwestern Tanzania. This was the first MVD outbreak in the country. We describe the epidemiological characteristics of MVD cases and contacts. METHODS: The Ministry of Health activated an outbreak response team. Outbreak investigation methods were applied to cases identified through MVD standard case definitions and confirmed through reverse-transcriptase polymerase chain reaction (RT PCR). All identified case contacts were added into the contact listing form and followed up in-person daily for any signs or symptoms for 21 days. Data collected from various forms was managed and analyzed using Excel and QGIS software for mapping. RESULTS: A total of nine MVD cases were reported with eight laboratory-confirmed and one probable. Two of the reported cases were frontline healthcare workers and seven were family related members. Cases were children and adults between 1-59 years of age with a median age of 34 years. Six were males. Six cases died equivalent to a case fatality rate (CFR) of 66.7%. A total of 212 individuals were identified as contacts and two (2) became cases. The outbreak was localized in two geo-administrative wards (Maruku and Kanyangereko) of Bukoba District Council. CONCLUSION: Transmission during this outbreak occurred among family members and healthcare workers who provided care to the cases. The delay in detection aggravated the spread and possibly the consequent fatality but once confirmed the swift response stemmed further transmission containing the disease at the epicenter wards. The outbreak lasted for 72 days but as the origin is still unknown, further research is required to explore the source of this outbreak.
Asunto(s)
Brotes de Enfermedades , Enfermedad del Virus de Marburg , Humanos , Tanzanía/epidemiología , Masculino , Femenino , Adulto , Persona de Mediana Edad , Enfermedad del Virus de Marburg/epidemiología , Enfermedad del Virus de Marburg/transmisión , Enfermedad del Virus de Marburg/virología , Niño , Adolescente , Lactante , Preescolar , Adulto Joven , Marburgvirus/genética , Marburgvirus/aislamiento & purificación , AnimalesRESUMEN
Filoviruses, like the Marburg (MARV) and Ebola (EBOV) viruses, have caused outbreaks associated with significant hemorrhagic morbidity and high fatality rates. Vaccines offer one of the best countermeasures for fatal infection, but to date only the EBOV vaccine has received FDA licensure. Given the limited cross protection between the EBOV vaccine and Marburg hemorrhagic fever (MHF), we analyzed the protective efficacy of a similar vaccine, rVSV-MARV, in the lethal cynomolgus macaque model. NHPs vaccinated with a single dose (as little as 1.6 × 107 pfu) of rVSV-MARV seroconverted to MARV G-protein prior to challenge on day 42. Vaccinemia was measured in all vaccinated primates, self-resolved by day 14 post vaccination. Importantly, all vaccinated NHPs survived lethal MARV challenge, and showed no significant alterations in key markers of morbid disease, including clinical signs, and certain hematological and clinical chemistry parameters. Further, apart from one primate (from which tissues were not collected and no causal link was established), no pathology associated with Marburg disease was observed in vaccinated animals. Taken together, rVSV-MARV is a safe and efficacious vaccine against MHF in cynomolgus macaques.
Asunto(s)
Macaca fascicularis , Enfermedad del Virus de Marburg , Marburgvirus , Vesiculovirus , Vacunas Virales , Animales , Enfermedad del Virus de Marburg/prevención & control , Enfermedad del Virus de Marburg/inmunología , Enfermedad del Virus de Marburg/virología , Marburgvirus/inmunología , Marburgvirus/genética , Vacunas Virales/inmunología , Vacunas Virales/administración & dosificación , Vacunas Virales/genética , Vesiculovirus/genética , Vesiculovirus/inmunología , Anticuerpos Antivirales/sangre , Anticuerpos Antivirales/inmunología , Vacunas Sintéticas/inmunología , Vacunas Sintéticas/administración & dosificación , Modelos Animales de Enfermedad , Vacunación , Masculino , Femenino , Eficacia de las Vacunas , Vectores Genéticos , Inmunogenicidad VacunalRESUMEN
Little is known regarding the molecular mechanisms that highly pathogenic Marburg virus (MARV) utilizes to transcribe and replicate its genome. Previous studies assumed that dephosphorylation of the filoviral transcription factor VP30 supports transcription, while phosphorylated VP30 reduces transcription. Here, we focused on the role of the host protein phosphatase 2A (PP2A) for VP30 dephosphorylation and promotion of viral transcription. We could show that MARV NP interacts with the subunit B56 of PP2A, as previously shown for the Ebola virus, and that this interaction is important for MARV transcription activity. Inhibition of the interaction between PP2A and NP either by mutating the B56 binding motif encoded on NP, or the use of a PP2A inhibitor, induced VP30 hyperphosphorylation, and as a consequence a decrease of MARV transcription as well as viral growth. These results suggest that NP plays a key role in the dephosphorylation of VP30 by recruiting PP2A. Generation of recombinant (rec) MARV lacking the PP2A-B56 interaction motif on NP was not possible suggesting an essential role of PP2A-mediated VP30 dephosphorylation for the MARV replication cycle. Likewise, we were not able to generate recMARV containing VP30 phosphomimetic mutants indicating that dynamic cycles of VP30 de- and rephosphorylation are a prerequisite for an efficient viral life cycle. As the specific binding motifs of PP2A-B56 and VP30 within NP are highly conserved among the filoviral family, our data suggest a conserved mechanism for filovirus VP30 dephosphorylation by PP2A, revealing the host factor PP2A as a promising target for pan-filoviral therapies. IMPORTANCE: Our study elucidates the crucial role of host protein phosphatase 2A (PP2A) in Marburg virus (MARV) transcription. The regulatory subunit B56 of PP2A facilitates VP30 dephosphorylation, and hence transcription activation, via binding to NP. Our results, together with previous data, reveal a conserved mechanism of filovirus VP30 dephosphorylation by host factor PP2A at the NP interface and provide novel insights into potential pan-filovirus therapies.
Asunto(s)
Marburgvirus , Proteína Fosfatasa 2 , Transcripción Genética , Marburgvirus/fisiología , Marburgvirus/genética , Marburgvirus/metabolismo , Proteína Fosfatasa 2/metabolismo , Proteína Fosfatasa 2/genética , Humanos , Fosforilación , Replicación Viral , Células HEK293 , Animales , Interacciones Huésped-Patógeno , Proteínas Virales/metabolismo , Proteínas Virales/genética , Factores de Transcripción/metabolismo , Factores de Transcripción/genética , Enfermedad del Virus de Marburg/virología , Enfermedad del Virus de Marburg/metabolismo , Unión Proteica , Línea CelularRESUMEN
Egyptian rousette bats (ERBs) are implicated as reservoir hosts for Marburg virus (MARV), but natural mechanisms involved in maintenance of MARV in ERB populations remain undefined. A number of hematophagous ectoparasites, including fleas, parasitize bats. Subcutaneous (SC) inoculation of ERBs with MARV consistently results in viremia, suggesting that infectious MARV could be ingested by blood-sucking ectoparasites during feeding. In our study, MARV RNA was detected in fleas that took a blood meal during feeding on viremic bats on days 3, 7, and 11 after SC inoculation. Virus concentration in individual ectoparasites was consistent with detectable levels of viremia in the blood of infected host bats. There was neither seroconversion nor viremia in control bats kept in close contact with MARV-infected bats infested with fleas for up to 40 days post-exposure. In fleas inoculated intracoelomically, MARV was detected up to 14 days after intracoelomic (IC) inoculation, but the virus concentration was lower than that delivered in the inoculum. All bats that had been infested with inoculated, viremic fleas remained virologically and serologically negative up to 38 days after infestation. Of 493 fleas collected from a wild ERB colony in Matlapitsi Cave, South Africa, where the enzootic transmission of MARV occurs, all tested negative for MARV RNA. While our findings seem to demonstrate that bat fleas lack vectorial capacity to transmit MARV biologically, their role in mechanical transmission should not be discounted. Regular blood-feeds, intra- and interhost mobility, direct feeding on blood vessels resulting in venous damage, and roosting behaviour of ERBs provide a potential physical bridge for MARV dissemination in densely populated cave-dwelling bats by fleas. The virus transfer might take place through inoculation of skin, mucosal membranes, and wounds when contaminated fleas are squashed during auto- and allogrooming, eating, biting, or fighting.
Asunto(s)
Quirópteros , Enfermedad del Virus de Marburg , Marburgvirus , Siphonaptera , Animales , Quirópteros/virología , Marburgvirus/genética , Marburgvirus/fisiología , Siphonaptera/virología , Enfermedad del Virus de Marburg/virología , Enfermedad del Virus de Marburg/transmisión , Reservorios de Enfermedades/virología , Viremia , Infestaciones por Pulgas/veterinaria , Infestaciones por Pulgas/transmisión , Infestaciones por Pulgas/virología , ARN Viral/genética , EgiptoRESUMEN
Marburg virus disease (MVD) is a dreadful but exceptional disease. Formerly mainly identified in Uganda, Angola and the Democratic Republic of Congo, it has recently appeared in the Republic of Guinea, Ghana, Equatorial Guinea and Tanzania, adding West Africa to the affected regions. Humans become infected through exposure to bats Roussettus aegyptiacus or during unprotected care of infected people. Five cases are linked to travellers, the last one dates to 2008 and involved a visit to caves colonized by bats. At present, there is no specific treatment or vaccine. Despite its rarity, adventurous travelers should be aware of the risks of exposure and avoid entering places inhabited by bats.
La maladie à virus Marburg est une maladie redoutable mais exceptionnelle. Autrefois identifiée en Ouganda, Angola et République démocratique du Congo, elle a récemment fait son apparition en République de Guinée, au Ghana, en Guinée équatoriale et en Tanzanie, ajoutant l'Afrique de l'Ouest aux régions touchées. Les humains s'infectent lors d'une exposition avec les chauves-souris roussettes d'Égypte ou lors de la prise en charge sans protection de personnes infectées. Cinq cas sont liés à des voyageurs, le dernier remonte à 2008 et était associé à la visite de grottes colonisées par des roussettes d'Égypte. Actuellement, il n'existe aucun traitement spécifique ni vaccin. Malgré sa rareté, les voyageurs aventureux doivent être informés des risques d'exposition et éviter de pénétrer dans des lieux habités par des chauves-souris.
Asunto(s)
Marburgvirus , Viaje , Femenino , Humanos , Masculino , Enfermedad del Virus de Marburg/epidemiología , Enfermedad del Virus de Marburg/transmisión , Enfermedad del Virus de Marburg/virología , Marburgvirus/aislamiento & purificación , Zoonosis Virales/epidemiología , Zoonosis Virales/transmisión , Zoonosis Virales/virología , Quirópteros/virologíaRESUMEN
The COVID-19 pandemic has not only strained healthcare systems in Africa but has also intensified the impact of emerging and re-emerging diseases. Specifically in Equatorial Guinea, mirroring the situation in other African countries, unique zoonotic outbreaks have occurred during this challenging period. One notable resurgence is Marburg virus disease (MVD), which has further burdened the already fragile healthcare system. The re-emergence of the Marburg virus amid the COVID-19 pandemic is believed to stem from a probable zoonotic spill-over, although the precise transmission routes remain uncertain. Given the gravity of the situation, addressing the existing challenges is paramount. Though the genome sequences from the current outbreak were not available for this study, we analyzed all the available whole genome sequences of this re-emerging pathogen to advocate for a shift towards active surveillance. This is essential to ensure the successful containment of any potential Marburg virus outbreak in Equatorial Guinea and the wider African context. This study, which presents an update on the phylodynamics and the genetic variability of MARV, further confirmed the existence of at least two distinct patterns of viral spread. One pattern demonstrates a slower but continuous and recurring virus circulation, while the other exhibits a faster yet limited and episodic spread. These results highlight the critical need to strengthen genomic surveillance in the region to effectively curb the pathogen's dissemination. Moreover, the study emphasizes the importance of prompt alert management, comprehensive case investigation and analysis, contact tracing, and active case searching. These steps are vital to support the healthcare system's response to this emerging health crisis. By implementing these strategies, we can better arm ourselves against the challenges posed by the resurgence of the Marburg virus and other infectious diseases.
Asunto(s)
Enfermedad del Virus de Marburg , Marburgvirus , Animales , Humanos , África/epidemiología , Población Negra , COVID-19/epidemiología , Marburgvirus/genética , Pandemias , Enfermedad del Virus de Marburg/epidemiología , Enfermedad del Virus de Marburg/genética , Enfermedad del Virus de Marburg/virología , Brotes de Enfermedades , Guinea Ecuatorial/epidemiología , Zoonosis Virales/epidemiología , Zoonosis Virales/genética , Zoonosis Virales/virología , FilogeniaAsunto(s)
Ensayos Clínicos como Asunto , Brotes de Enfermedades , Enfermedad del Virus de Marburg , Marburgvirus , Vacunas Virales , Humanos , Brotes de Enfermedades/prevención & control , Brotes de Enfermedades/estadística & datos numéricos , Enfermedad del Virus de Marburg/epidemiología , Enfermedad del Virus de Marburg/prevención & control , Enfermedad del Virus de Marburg/virología , Factores de TiempoRESUMEN
Marburg virus (MARV) is a lipid-enveloped virus harboring a negative-sense RNA genome, which has caused sporadic outbreaks of viral hemorrhagic fever in sub-Saharan Africa. MARV assembles and buds from the host cell plasma membrane where MARV matrix protein (mVP40) dimers associate with anionic lipids at the plasma membrane inner leaflet and undergo a dynamic and extensive self-oligomerization into the structural matrix layer. The MARV matrix layer confers the virion filamentous shape and stability but how host lipids modulate mVP40 oligomerization is mostly unknown. Using in vitro and cellular techniques, we present a mVP40 assembly model highlighting two distinct oligomerization interfaces: the (N-terminal domain [NTD] and C-terminal domain [CTD]) in mVP40. Cellular studies of NTD and CTD oligomerization interface mutants demonstrate the importance of each interface in matrix assembly. The assembly steps include protein trafficking to the plasma membrane, homo-multimerization that induced protein enrichment, plasma membrane fluidity changes, and elongations at the plasma membrane. An ascorbate peroxidase derivative (APEX)-transmission electron microscopy method was employed to closely assess the ultrastructural localization and formation of viral particles for wildtype mVP40 and NTD and CTD oligomerization interface mutants. Taken together, these studies present a mechanistic model of mVP40 oligomerization and assembly at the plasma membrane during virion assembly that requires interactions with phosphatidylserine for NTD-NTD interactions and phosphatidylinositol-4,5-bisphosphate for proper CTD-CTD interactions. These findings have broader implications in understanding budding of lipid-enveloped viruses from the host cell plasma membrane and potential strategies to target protein-protein or lipid-protein interactions to inhibit virus budding.
Asunto(s)
Enfermedad del Virus de Marburg/virología , Marburgvirus/fisiología , Lípidos de la Membrana/metabolismo , Proteínas de la Matriz Viral/metabolismo , Virión/metabolismo , Animales , Células COS , Membrana Celular/química , Membrana Celular/metabolismo , Chlorocebus aethiops , Células HEK293 , Humanos , Membrana Dobles de Lípidos/química , Membrana Dobles de Lípidos/metabolismo , Enfermedad del Virus de Marburg/metabolismo , Marburgvirus/química , Lípidos de la Membrana/química , Modelos Moleculares , Multimerización de Proteína , Proteínas de la Matriz Viral/química , Virión/química , Ensamble de VirusRESUMEN
BACKGROUND: Marburg virus (MARV) and Ebola virus (EBOV) are acute infections with high case fatality rates. It is of great significance for epidemic monitoring and prevention and control of infectious diseases by the development of a rapid, specific, and sensitive quantitative PCR method to detect two pathogens simultaneously. METHODS: Primers and TaqMan probes were designed according to highly conserved sequences of these viruses. Sensitivity, specificity, linear range, limit of detection, and the effects of hemolysis and lipid on real-time qPCR were evaluated. RESULTS: The linearity of the curve allowed quantification of nucleic acid concentrations in range from 103 to 109 copies/ml per reaction (MARV and EBOV). The limit of detection of EBOV was 40 copies/ml, and MARV was 100 copies/ml. It has no cross-reaction with other pathogens such as hepatitis b virus (HBV), hepatitis c virus (HCV), human papillomavirus (HPV), Epstein-Barr virus (EBV), herpes simplex virus (HSV), cytomegalovirus (CMV), and human immunodeficiency virus (HIV). Repeatability analysis of the two viruses showed that their coefficient of variation (CV) was less than 5.0%. The above results indicated that fluorescence quantitative PCR could detect EBOV and MARV sensitively and specifically. CONCLUSIONS: The TaqMan probe-based multiplex fluorescence quantitative PCR assays could detect EBOV and MARV sensitively specifically and simultaneously.
Asunto(s)
Ebolavirus/genética , Fiebre Hemorrágica Ebola/diagnóstico , Enfermedad del Virus de Marburg/diagnóstico , Marburgvirus/genética , Reacción en Cadena de la Polimerasa Multiplex/métodos , Reacción en Cadena en Tiempo Real de la Polimerasa/métodos , Animales , Fiebre Hemorrágica Ebola/virología , Humanos , Enfermedad del Virus de Marburg/virología , Curva ROCRESUMEN
Filoviruses Ebola (EBOV) and Marburg (MARV) are devastating high-priority pathogens capable of causing explosive outbreaks with high human mortality rates. The matrix proteins of EBOV and MARV, as well as eVP40 and mVP40, respectively, are the key viral proteins that drive virus assembly and egress and can bud independently from cells in the form of virus-like particles (VLPs). The matrix proteins utilize proline-rich Late (L) domain motifs (e.g., PPxY) to hijack specific host proteins that contain WW domains, such as the HECT family E3 ligases, to facilitate the last step of virus-cell separation. We identified E3 ubiquitin ligase Smad Ubiquitin Regulatory Factor 2 (SMURF2) as a novel interactor with VP40 that positively regulates VP40 VLP release. Our results show that eVP40 and mVP40 interact with the three WW domains of SMURF2 via their PPxY motifs. We provide evidence that the eVP40-SMURF2 interaction is functional as the expression of SMURF2 positively regulates VLP egress, while siRNA knockdown of endogenous SMURF2 decreases VLP budding compared to controls. In sum, our identification of novel interactor SMURF2 adds to the growing list of identified host proteins that can regulate PPxY-mediated egress of VP40 VLPs. A more comprehensive understanding of the modular interplay between filovirus VP40 and host proteins may lead to the development of new therapies to combat these deadly infections.
Asunto(s)
Ebolavirus/fisiología , Fiebre Hemorrágica Ebola/enzimología , Enfermedad del Virus de Marburg/enzimología , Marburgvirus/fisiología , Ubiquitina-Proteína Ligasas/metabolismo , Proteínas de la Matriz Viral/metabolismo , Liberación del Virus , Secuencias de Aminoácidos , Animales , Ebolavirus/química , Ebolavirus/genética , Fiebre Hemorrágica Ebola/genética , Fiebre Hemorrágica Ebola/virología , Humanos , Enfermedad del Virus de Marburg/genética , Enfermedad del Virus de Marburg/virología , Marburgvirus/química , Marburgvirus/genética , Unión Proteica , Ubiquitina-Proteína Ligasas/genética , Proteínas de la Matriz Viral/química , Proteínas de la Matriz Viral/genética , Virión/genética , Virión/fisiología , Ensamble de VirusRESUMEN
Marburg virus (MARV) is among the most virulent pathogens of primates, including humans. Contributors to severe MARV disease include immune response suppression and inflammatory gene dysregulation ("cytokine storm"), leading to systemic damage and often death. Conversely, MARV causes little to no clinical disease in its reservoir host, the Egyptian rousette bat (ERB). Previous genomic and in vitro data suggest that a tolerant ERB immune response may underlie MARV avirulence, but no significant examination of this response in vivo yet exists. Here, using colony-bred ERBs inoculated with a bat isolate of MARV, we use species-specific antibodies and an immune gene probe array (NanoString) to temporally characterize the transcriptional host response at sites of MARV replication relevant to primate pathogenesis and immunity, including CD14+ monocytes/macrophages, critical immune response mediators, primary MARV targets, and skin at the inoculation site, where highest viral loads and initial engagement of antiviral defenses are expected. Our analysis shows that ERBs upregulate canonical antiviral genes typical of mammalian systems, such as ISG15, IFIT1, and OAS3, yet demonstrate a remarkable lack of significant induction of proinflammatory genes classically implicated in primate filoviral pathogenesis, including CCL8, FAS, and IL6. Together, these findings offer the first in vivo functional evidence for disease tolerance as an immunological mechanism by which the bat reservoir asymptomatically hosts MARV. More broadly, these data highlight factors determining disparate outcomes between reservoir and spillover hosts and defensive strategies likely utilized by bat hosts of other emerging pathogens, knowledge that may guide development of effective antiviral therapies.
Asunto(s)
Quirópteros/inmunología , Reservorios de Enfermedades/virología , Tolerancia Inmunológica/inmunología , Enfermedad del Virus de Marburg/inmunología , Marburgvirus/inmunología , Animales , Infecciones Asintomáticas , Quirópteros/sangre , Quirópteros/genética , Quirópteros/virología , Femenino , Regulación de la Expresión Génica/inmunología , Interacciones Huésped-Patógeno/genética , Interacciones Huésped-Patógeno/inmunología , Humanos , Tolerancia Inmunológica/genética , Masculino , Enfermedad del Virus de Marburg/virología , Monocitos/inmunologíaRESUMEN
Marburg virus (MARV) disease is lethal, with fatality rates up to 90%. Neutralizing antibodies (Abs) are promising drug candidates to prevent or treat the disease. Current efforts are focused in part on vaccine development to induce such MARV-neutralizing Abs. We analyzed the antibody repertoire from healthy unexposed and previously MARV-infected individuals to assess if naïve repertoires contain suitable precursor antibodies that could become neutralizing with a limited set of somatic mutations. We computationally searched the human Ab variable gene repertoire for predicted structural homologs of the neutralizing Ab MR78 that is specific to the receptor binding site (RBS) of MARV glycoprotein (GP). Eight Ab heavy-chain complementarity determining region 3 (HCDR3) loops from MARV-naïve individuals and one from a previously MARV-infected individual were selected for testing as HCDR3 loop chimeras on the MR78 Ab framework. Three of these chimerized antibodies bound to MARV GP. We then tested a full-length native Ab heavy chain encoding the same 17-residue-long HCDR3 loop that bound to the MARV GP the best among the chimeric Abs tested. Despite only 57% amino acid sequence identity, the Ab from a MARV-naïve donor recognized MARV GP and possessed neutralizing activity against the virus. Crystallization of both chimeric and full-length native heavy chain-containing Abs provided structural insights into the mechanism of binding for these types of Abs. Our work suggests that the MARV GP RBS is a promising candidate for epitope-focused vaccine design to induce neutralizing Abs against MARV.
Asunto(s)
Anticuerpos Antivirales/genética , Regiones Determinantes de Complementariedad/genética , Enfermedad del Virus de Marburg/inmunología , Marburgvirus/inmunología , Animales , Anticuerpos Neutralizantes/genética , Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/inmunología , Regiones Determinantes de Complementariedad/inmunología , Epítopos/genética , Epítopos/inmunología , Glicoproteínas/genética , Glicoproteínas/inmunología , Humanos , Cadenas Pesadas de Inmunoglobulina/genética , Cadenas Pesadas de Inmunoglobulina/inmunología , Enfermedad del Virus de Marburg/tratamiento farmacológico , Enfermedad del Virus de Marburg/genética , Enfermedad del Virus de Marburg/virología , Marburgvirus/patogenicidad , Mutación/genética , Mutación/inmunología , Proteínas del Envoltorio Viral , Vacunas Virales/genética , Vacunas Virales/inmunologíaRESUMEN
OBJECTIVES: This article summarizes the countermeasures for Marburg virus disease, focusing on pathogenesis, clinical features and diagnostics. There is an emphasis on therapies and vaccines that have demonstrated, through their evaluation in nonhuman primates (NHPs) and/or in humans, potential for use in an emergency situation. METHODS: A standardized literature review was conducted on vaccines and treatments for Marburg virus disease, with a focus on human and nonhuman primate data published in the last five years. More detail on the methods that were used is summarized in a companion methods paper. RESULTS: The study identified six treatments and four vaccine platforms that have demonstrated, through their efficacy in NHPs, potential benefit for treating or preventing infection in humans. CONCLUSION: Succinct summaries of Marburg countermeasures are provided to give the busy clinician a head start in reviewing the literature if faced with a patient with Marburg virus disease. Links to other authoritative sources of information are also provided.
Asunto(s)
Enfermedad del Virus de Marburg/terapia , Animales , Humanos , Enfermedad del Virus de Marburg/inmunología , Enfermedad del Virus de Marburg/prevención & control , Enfermedad del Virus de Marburg/virología , Marburgvirus/inmunología , Vacunas Virales/administración & dosificación , Vacunas Virales/inmunologíaRESUMEN
The recent Ebola epidemics in West Africa underscore the great need for effective and practical therapies for future Ebola virus outbreaks. We have discovered a new series of remarkably potent small molecule inhibitors of Ebola virus entry. These 4-(aminomethyl)benzamide-based inhibitors are also effective against Marburg virus. Synthetic routes to these compounds allowed for the preparation of a wide variety of structures, including a conformationally restrained subset of indolines (compounds 41-50). Compounds 20, 23, 32, 33, and 35 are superior inhibitors of Ebola (Mayinga) and Marburg (Angola) infectious viruses. Representative compounds (20, 32, and 35) have shown good metabolic stability in plasma and liver microsomes (rat and human), and 32 did not inhibit CYP3A4 nor CYP2C9. These 4-(aminomethyl)benzamides are suitable for further optimization as inhibitors of filovirus entry, with the potential to be developed as therapeutic agents for the treatment and control of Ebola virus infections.
Asunto(s)
Antivirales/farmacología , Benzamidas/farmacología , Fiebre Hemorrágica Ebola/virología , Enfermedad del Virus de Marburg/virología , Internalización del Virus/efectos de los fármacos , Células A549 , Animales , Antivirales/química , Benzamidas/química , Chlorocebus aethiops , Inhibidores del Citocromo P-450 CYP3A/química , Inhibidores del Citocromo P-450 CYP3A/farmacología , Evaluación Preclínica de Medicamentos , Humanos , Microsomas Hepáticos/efectos de los fármacos , Simulación del Acoplamiento Molecular , Relación Estructura-Actividad , Toremifeno/química , Toremifeno/metabolismo , Toremifeno/farmacología , Células Vero , Proteínas del Envoltorio Viral/química , Proteínas del Envoltorio Viral/metabolismoRESUMEN
Outbreaks of filoviruses, such as those caused by the Ebola (EBOV) and Marburg (MARV) virus, are difficult to detect and control. The initial clinical symptoms of these diseases are nonspecific and can mimic other endemic pathogens. This makes confident diagnosis based on clinical symptoms alone impossible. Molecular diagnostics for these diseases that rely on the detection of viral RNA in the blood are only effective after significant disease progression. As an approach to identify these infections earlier in the disease course, we tested the effectiveness of viral RNA detection combined with an assessment of sentinel host mRNAs that are upregulated following filovirus infection. RNAseq analysis of EBOV-infected nonhuman primates identified host RNAs that are upregulated at early stages of infection. NanoString probes that recognized these host-response RNAs were combined with probes that recognized viral RNA and were used to classify viral infection both prior to viremia and postviremia. This approach was highly successful at identifying samples from nonhuman primate subjects and correctly distinguished the causative agent in a previremic stage in 10 EBOV and 5 MARV samples. This work suggests that unified host response/viral fingerprint assays can enable diagnosis of disease earlier than testing for viral nucleic acid alone, which could decrease transmission events and increase therapeutic effectiveness.IMPORTANCE Current molecular tests that identify infection with high-consequence viruses such as Ebola virus and Marburg virus are based on the detection of virus material in the blood. These viruses do not undergo significant early replication in the blood and, instead, replicate in organs such as the liver and spleen. Thus, virus begins to accumulate in the blood only after significant replication has already occurred in those organs, making viremia an indicator of infection only after initial stages have become established. Here, we show that a multianalyte assay can correctly identify the infectious agent in nonhuman primates (NHPs) prior to viremia through tracking host infection response transcripts. This illustrates that a single-tube, sample-to-answer format assay could be used to advance the time at which the type of infection can be determined and thereby improve outcomes.
Asunto(s)
Genoma Viral , Fiebre Hemorrágica Ebola/diagnóstico , Interacciones Huésped-Patógeno/genética , Enfermedad del Virus de Marburg/diagnóstico , ARN Viral/aislamiento & purificación , Transcriptoma , Animales , Ebolavirus/genética , Fiebre Hemorrágica Ebola/virología , Humanos , Macaca , Enfermedad del Virus de Marburg/virología , Marburgvirus/genética , Análisis por Micromatrices , Proteínas Virales/sangre , Proteínas Virales/genética , ViremiaRESUMEN
Marburg virus (MARV) is a filovirus with documented human case-fatality rates of up to 90%. Here, we evaluated the therapeutic efficacy of remdesivir (GS-5734) in nonhuman primates experimentally infected with MARV. Beginning 4 or 5 days post inoculation, cynomolgus macaques were treated once daily for 12 days with vehicle, 5 mg/kg remdesivir, or a 10-mg/kg loading dose followed by 5 mg/kg remdesivir. All vehicle-control animals died, whereas 83% of animals receiving a 10-mg/kg loading dose of remdesivir survived, as did 50% of animals receiving a 5-mg/kg remdesivir regimen. Remdesivir-treated animals exhibited improved clinical scores, lower plasma viral RNA, and improved markers of kidney function, liver function, and coagulopathy versus vehicle-control animals. The small molecule remdesivir showed therapeutic efficacy in this Marburg virus disease model with treatment initiation 5 days post inoculation, supporting further assessment of remdesivir for the treatment of Marburg virus disease in humans.
Asunto(s)
Antimetabolitos/uso terapéutico , Antivirales/uso terapéutico , Enfermedad del Virus de Marburg/tratamiento farmacológico , Marburgvirus/efectos de los fármacos , Enfermedades de los Monos/tratamiento farmacológico , Adenosina Monofosfato/análogos & derivados , Alanina/análogos & derivados , Animales , Modelos Animales de Enfermedad , Femenino , Estimación de Kaplan-Meier , Macaca fascicularis , Masculino , Enfermedad del Virus de Marburg/mortalidad , Enfermedad del Virus de Marburg/patología , Enfermedad del Virus de Marburg/virología , Enfermedades de los Monos/mortalidad , Enfermedades de los Monos/patología , Enfermedades de los Monos/virología , ARN ViralRESUMEN
Marburg virus (MARV) is a lipid-enveloped negative sense single stranded RNA virus, which can cause a deadly hemorrhagic fever. MARV encodes seven proteins, including VP40 (mVP40), a matrix protein that interacts with the cytoplasmic leaflet of the host cell plasma membrane. VP40 traffics to the plasma membrane inner leaflet, where it assembles to facilitate the budding of viral particles. VP40 is a multifunctional protein that interacts with several host proteins and lipids to complete the viral replication cycle, but many of these host interactions remain unknown or are poorly characterized. In this study, we investigated the role of a hydrophobic loop region in the carboxy-terminal domain (CTD) of mVP40 that shares sequence similarity with the CTD of Ebola virus VP40 (eVP40). These conserved hydrophobic residues in eVP40 have been previously shown to be critical to plasma membrane localization and membrane insertion. An array of cellular experiments and confirmatory in vitro work strongly suggests proper orientation and hydrophobic residues (Phe281, Leu283, and Phe286) in the mVP40 CTD are critical to plasma membrane localization. In line with the different functions proposed for eVP40 and mVP40 CTD hydrophobic residues, molecular dynamics simulations demonstrate large flexibility of residues in the EBOV CTD whereas conserved mVP40 hydrophobic residues are more restricted in their flexibility. This study sheds further light on important amino acids and structural features in mVP40 required for its plasma membrane localization as well as differences in the functional role of CTD amino acids in eVP40 and mVP40.
Asunto(s)
Membrana Celular/metabolismo , Enfermedad del Virus de Marburg/virología , Marburgvirus/fisiología , Mutación , Dominios y Motivos de Interacción de Proteínas , Proteínas de la Matriz Viral/química , Proteínas de la Matriz Viral/metabolismo , Aminoácidos/química , Animales , Células COS , Membrana Celular/química , Chlorocebus aethiops , Células HEK293 , Humanos , Interacciones Hidrofóbicas e Hidrofílicas , Lípidos , Modelos Moleculares , Imagen Molecular , Conformación Proteica , Transporte de ProteínasRESUMEN
Postexposure immunization can prevent disease and reduce transmission following pathogen exposure. The rapid immunostimulatory properties of recombinant vesicular stomatitis virus (rVSV)-based vaccines make them suitable postexposure treatments against the filoviruses Ebola virus and Marburg virus (MARV); however, the mechanisms that drive this protection are undefined. Previously, we reported 60-75% survival of rhesus macaques treated with rVSV vectors expressing MARV glycoprotein (GP) 20-30 minutes after a low dose exposure to the most pathogenic variant of MARV, Angola. Survival in this model was linked to production of GP-specific antibodies and lower viral load. To confirm these results and potentially identify novel correlates of postexposure protection, we performed a similar experiment, but analyzed plasma cytokine levels, frequencies of immune cell subsets, and the transcriptional response to infection in peripheral blood. In surviving macaques (80-89%), we observed induction of genes mapping to antiviral and interferon-related pathways early after treatment and a higher percentage of T helper 1 (Th1) and NK cells. In contrast, the response of non-surviving macaques was characterized by hypercytokinemia; a T helper 2 signature; recruitment of low HLA-DR expressing monocytes and regulatory T-cells; and transcription of immune checkpoint (e.g., PD-1, LAG3) genes. These results suggest dysregulated immunoregulation is associated with poor prognosis, whereas early innate signaling and Th1-skewed immunity are important for survival.
Asunto(s)
Enfermedad del Virus de Marburg/inmunología , Enfermedad del Virus de Marburg/virología , Marburgvirus/inmunología , Profilaxis Posexposición , Vacunas Virales/inmunología , Animales , Anticuerpos Antivirales/biosíntesis , Anticuerpos Antivirales/inmunología , Citocinas/sangre , Citotoxicidad Inmunológica , Relación Dosis-Respuesta Inmunológica , Regulación hacia Abajo/genética , Femenino , Inflamación/sangre , Inflamación/inmunología , Interferones/genética , Interferones/metabolismo , Células Asesinas Naturales/inmunología , Macaca mulatta/inmunología , Macaca mulatta/virología , Masculino , Enfermedad del Virus de Marburg/sangre , Enfermedad del Virus de Marburg/genética , ARN Mensajero/genética , ARN Mensajero/metabolismo , Recombinación Genética/genética , Linfocitos T Colaboradores-Inductores/inmunología , Células TH1/inmunología , Células Th2/inmunología , Transcriptoma/genética , Regulación hacia Arriba/genética , Vesiculovirus/genética , Carga Viral/inmunologíaRESUMEN
Marburg virus (MARV) causes sporadic outbreaks of severe Marburg virus disease (MVD). Most MVD outbreaks originated in East Africa and field studies in East Africa, South Africa, Zambia, and Gabon identified the Egyptian rousette bat (ERB; Rousettus aegyptiacus) as a natural reservoir. However, the largest recorded MVD outbreak with the highest case-fatality ratio happened in 2005 in Angola, where direct spillover from bats was not shown. Here, collaborative studies by the Centers for Disease Control and Prevention, Njala University, University of California, Davis USAID-PREDICT, and the University of Makeni identify MARV circulating in ERBs in Sierra Leone. PCR, antibody and virus isolation data from 1755 bats of 42 species shows active MARV infection in approximately 2.5% of ERBs. Phylogenetic analysis identifies MARVs that are similar to the Angola strain. These results provide evidence of MARV circulation in West Africa and demonstrate the value of pathogen surveillance to identify previously undetected threats.