A novel MARV glycoprotein-specific antibody with potentials of broad-spectrum neutralization to filovirus.

Marburg virus (MARV) is one of the filovirus species that cause deadly hemorrhagic fever in humans, with mortality rates up to 90%. Neutralizing antibodies represent ideal candidates to prevent or treat virus disease. However, no antibody has been approved for MARV treatment to date. In this study, we identified a novel human antibody named AF-03 that targeted MARV glycoprotein (GP). AF-03 possessed a high binding affinity to MARV GP and showed neutralizing and protective activities against the pseudotyped MARV in vitro and in vivo. Epitope identification, including molecular docking and experiment-based analysis of mutated species, revealed that AF-03 recognized the Niemann-Pick C1 (NPC1) binding domain within GP1. Interestingly, we found the neutralizing activity of AF-03 to pseudotyped Ebola viruses (EBOV, SUDV, and BDBV) harboring cleaved GP instead of full-length GP. Furthermore, NPC2-fused AF-03 exhibited neutralizing activity to several filovirus species and EBOV mutants via binding to CI-MPR. In conclusion, this work demonstrates that AF-03 represents a promising therapeutic cargo for filovirus-caused disease.

Production and Purification of Filovirus Glycoproteins.

Ebola (EBOV) and Marburg (MARV) viruses cause hemorrhagic fever disease in humans and non-human primates (NHPs) with case-fatality rates as high as 90%. The 2013-2016 Ebola virus disease (EVD) outbreak led to over 28,000 cases and 11,000 deaths and took an enormous toll on the economy of West African nations, in the absence of any vaccine or therapeutic options. Like EVD, there have been at least 6 outbreaks of MVD with ~88% case-fatality and the most recent cases emerging in Equatorial Guinea in February 2023. These outbreaks have spurred an unprecedented global effort to develop vaccines and therapeutics for EVD and MVD and led to an approved vaccine (ERVEBO™) and two monoclonal antibody (mAb) therapeutics for EBOV. In contrast to EVD, therapeutic options against Marburg and another Ebola-relative Sudan virus (SUDV) are lacking. The filovirus glycoprotein (GP), which mediates host cell entry and fusion, is the primary target of neutralizing antibodies. In addition to its pre- and post-fusion trimeric states, the protein is highly glycosylated making production of pure and homogeneous trimers on a large scale, a requirement for subunit vaccine development, a challenge. In efforts to address this roadblock, we have developed a unique combination of structure-based design, selection of expression system, and purification methods to produce uniform and stable EBOV and MARV GP trimers at scales appropriate for vaccine production.

Transcriptomics of Marburg virus-infected primary proximal tubular cells reveals negative correlation of immune response and energy metabolism.

Marburg virus, a member of the Filoviridae, is the causative agent of Marburg virus disease (MVD), a hemorrhagic fever with a case fatality rate of up to 90 %. Acute kidney injury is common in MVD and is associated with increased mortality, but its pathogenesis in MVD remains poorly understood. Interestingly, autopsies show the presence of viral proteins in different parts of the nephron, particularly in proximal tubular cells (PTC). These findings suggest a potential role for the virus in the development of MVD-related kidney injury. To shed light on this effect, we infected primary human PTC with Lake Victoria Marburg virus and conducted transcriptomic analysis at multiple time points. Unexpectedly, infection did not induce marked cytopathic effects in primary tubular cells at 20 and 40 h post infection. However, gene expression analysis revealed robust renal viral replication and dysregulation of genes essential for different cellular functions. The gene sets mainly downregulated in PTC were associated with the targets of the transcription factors MYC and E2F, DNA repair, the G2M checkpoint, as well as oxidative phosphorylation. Importantly, the downregulated factors comprise PGC-1α, a well-known factor in acute and chronic kidney injury. By contrast, the most highly upregulated gene sets were those related to the inflammatory response and cholesterol homeostasis. In conclusion, Marburg virus infects and replicates in human primary PTC and induces downregulation of processes known to be relevant for acute kidney injury as well as a strong inflammatory response.

ICTV Virus Taxonomy Profile: Filoviridae 2024.

Filoviridae is a family of negative-sense RNA viruses with genomes of about 13.1-20.9 kb that infect fish, mammals and reptiles. The filovirid genome is a linear, non-segmented RNA with five canonical open reading frames (ORFs) that encode a nucleoprotein (NP), a polymerase cofactor (VP35), a glycoprotein (GP1,2), a transcriptional activator (VP30) and a large protein (L) containing an RNA-directed RNA polymerase (RdRP) domain. All filovirid genomes encode additional proteins that vary among genera. Several filovirids (e.g., Ebola virus, Marburg virus) are pathogenic for humans and highly virulent. This is a summary of the International Committee on Taxonomy of Viruses (ICTV) Report on the family Filoviridae, which is available at www.ictv.global/report/filoviridae.

Bat-Borne Pathogens and Public Health in Rural African Artisanal Gold Mines.

Marburg virus, the first filovirus discovered and a close cousin to the Ebola virus, is carried by the Egyptian rousette bat, a common cave-dwelling fruit bat endemic to sub-Saharan Africa whose populations can exceed 50 000 individuals. Community outbreaks of Marburg virus can result in high morbidity rates. In eastern Africa, favorite habitats of these bats include rural subterranean gold mines-sometimes worked illegally-that create environments conducive to zoonotic virus transmission. This commentary on a case describes how outbreaks of Marburg virus disease among people exposed to sub-Saharan African caves and mines containing these bats cause tensions among miners, companies, public health officials, and conservationists.

Coordinated inflammatory responses dictate Marburg virus control by reservoir bats.

Bats are increasingly recognized as reservoirs of emerging zoonotic pathogens. Egyptian rousette bats (ERBs) are the known reservoir of Marburg virus (MARV), a filovirus that causes deadly Marburg virus disease (MVD) in humans. However, ERBs harbor MARV asymptomatically, likely due to a coadapted and specific host immunity-pathogen relationship. Recently, we measured transcriptional responses in MARV-infected ERB whole tissues, showing that these bats possess a disease tolerant strategy that limits pro-inflammatory gene induction, presumably averting MVD-linked immunopathology. However, the host resistant strategy by which ERBs actively limit MARV burden remains elusive, which we hypothesize requires localized inflammatory responses unresolvable at bulk-tissue scale. Here, we use dexamethasone to attenuate ERB pro-inflammatory responses and assess MARV replication, shedding and disease. We show that MARV-infected ERBs naturally mount coordinated pro-inflammatory responses at liver foci of infection, comprised of recruited mononuclear phagocytes and T cells, the latter of which proliferate with likely MARV-specificity. When pro-inflammatory responses are diminished, ERBs display heightened MARV replication, oral/rectal shedding and severe MVD-like liver pathology, demonstrating that ERBs balance immunoprotective tolerance with discreet MARV-resistant pro-inflammatory responses. These data further suggest that natural ERB immunomodulatory stressors like food scarcity and habitat disruption may potentiate viral shedding, transmission and therefore outbreak risk.

Marburg Virus Outbreak in Equatorial Guinea: Need for Speed.

The co-existence of deadly viral pandemics can be considered a nightmare for public health authorities. The surge of a Marburg virus disease (MVD) outbreak in Africa at a time when the coronavirus-19 (COVID-19) pandemic is partially controlled with its limited resources is an urgent call for concern. Over the past decades, several bouts of MVD outbreaks have occurred in Africa with an alarming case fatality rate. Despite this, little has been done to end its recurrence, and affected countries essentially depend on preventative rather than curative measures of management. The recent outbreak of MVD declared by the health officials of Equatorial Guinea, causing several deaths in the context of the COVID-19 pandemic, signals the need for speed in the establishment and the implementation of appropriate health policies and health system strategies to contain, destroy, and prevent the spread of this deadly virus to other neighboring countries.

Functional characterization of AF-04, an afucosylated anti-MARV GP antibody.

Marburg virus (MARV), one member of the Filoviridae family, cause sporadic outbreaks of hemorrhagic fever with high mortality rates. No countermeasures are currently available for the prevention or treatment of MARV infection. Monoclonal antibodies (mAbs) are promising candidates to display high neutralizing activity against MARV infection in vitro and in vivo. Recently, growing evidence has shown that immune effector function including antibody-dependent cell-mediated cytotoxicity (ADCC) is also required for in vivo efficacy of a panel of antibodies. Glyco-engineered methods are widely utilized to augment ADCC function of mAbs. In this study, we generated a fucose-knockout MARV GP-specific mAb named AF-04 and showed that afucosylation dramatically increased its binding affinity to polymorphic FcγRIIIa (F176/V176) compared with the parental AF-03. Accordingly, AF-04-mediated NK cell activation and NFAT expression downstream of FcγRIIIa in effector cells were also augmented. In conclusion, this work demonstrates that AF-04 represents a novel avenue for the treatment of MARV-caused disease.

Thermostable bivalent filovirus vaccine protects against severe and lethal Sudan ebolavirus and marburgvirus infection.

Although two vaccines for Zaire ebolavirus (EBOV) have been licensed and deployed successfully to combat recurring outbreaks of Ebolavirus Disease in West Africa, there are no vaccines for two other highly pathogenic members of the Filoviridae, Sudan ebolavirus (SUDV) and Marburg marburgvirus (MARV). The results described herein document the immunogenicity and protective efficacy in cynomolgus macaques of a single-vial, thermostabilized (lyophilized) monovalent (SUDV) and bivalent (SUDV & MARV) protein vaccines consisting of recombinant glycoproteins (GP) formulated with a clinical-grade oil-in-water nanoemulsion adjuvant (CoVaccine HT™). Lyophilized formulations of the vaccines were reconstituted with Water for Injection and used to immunize groups of cynomolgus macaques before challenge with a lethal dose of a human SUDV or MARV isolate. Sera collected after each of the three immunizations showed near maximal GP-binding IgG concentrations starting as early as the second dose. Most importantly, the vaccine candidates (monovalent or bivalent) provided 100% protection against severe and lethal filovirus disease after either SUDV or MARV infection. Although mild, subclinical infection was observed in a few macaques, all vaccinated animals remained healthy and survived the filovirus challenge. These results demonstrate the value that thermostabilized protein vaccines could provide for addressing an important gap in preparedness for future filovirus outbreaks.

Development of an Immunochromatography Assay to Detect Marburg Virus and Ravn Virus.

The recent outbreaks of Marburg virus disease (MVD) in Guinea, Ghana, Equatorial Guinea, and Tanzania, none of which had reported previous outbreaks, imply increasing risks of spillover of the causative viruses, Marburg virus (MARV) and Ravn virus (RAVV), from their natural host animals. These outbreaks have emphasized the need for the development of rapid diagnostic tests for this disease. Using monoclonal antibodies specific to the viral nucleoprotein, we developed an immunochromatography (IC) assay for the rapid diagnosis of MVD. The IC assay was found to be capable of detecting approximately 102-4 50% tissue culture infectious dose (TCID50)/test of MARV and RAVV in the infected culture supernatants. We further confirmed that the IC assay could detect the MARV and RAVV antigens in the serum samples from experimentally infected nonhuman primates. These results indicate that the IC assay to detect MARV can be a useful tool for the rapid point-of-care diagnosis of MVD.

Characterization of the Cynomolgus Macaque Model of Marburg Virus Disease and Assessment of Timing for Therapeutic Treatment Testing.

Marburg virus (MARV) causes severe disease and high mortality in humans. The objective of this study was to characterize disease manifestations and pathogenesis in cynomolgus macaques exposed to MARV. The results of this natural history study may be used to identify features of MARV disease useful in defining the ideal treatment initiation time for subsequent evaluations of investigational therapeutics using this model. Twelve cynomolgus macaques were exposed to a target dose of 1000 plaque-forming units MARV by the intramuscular route, and six control animals were mock-exposed. The primary endpoint of this study was survival to Day 28 post-inoculation (PI). Anesthesia events were minimized with the use of central venous catheters for periodic blood collection, and temperature and activity were continuously monitored by telemetry. All mock-exposed animals remained healthy for the duration of the study. All 12 MARV-exposed animals (100%) became infected, developed illness, and succumbed on Days 8-10 PI. On Day 4 PI, 11 of the 12 MARV-exposed animals had statistically significant temperature elevations over baseline. Clinically observable signs of MARV disease first appeared on Day 5 PI, when 6 of the 12 animals exhibited reduced responsiveness. Ultimately, systemic inflammation, coagulopathy, and direct cytopathic effects of MARV all contributed to multiorgan dysfunction, organ failure, and death or euthanasia of all MARV-exposed animals. Manifestations of MARV disease, including fever, systemic viremia, lymphocytolysis, coagulopathy, and hepatocellular damage, could be used as triggers for initiation of treatment in future therapeutic efficacy studies.

A nonlinear fractional epidemic model for the Marburg virus transmission with public health education.

In this study, a deterministic model for the dynamics of Marburg virus transmission that incorporates the impact of public health education is being formulated and analyzed. The Caputo fractional-order derivative is used to extend the traditional integer model to a fractional-based model. The model's positivity and boundedness are also under investigation. We obtain the basic reproduction number [Formula: see text] and establish the conditions for the local and global asymptotic stability for the disease-free equilibrium of the model. Under the Caputo fractional-order derivative, we establish the existence-uniqueness theory using the Banach contraction mapping principle for the solution of the proposed model. We use functional techniques to demonstrate the proposed model's stability under the Ulam-Hyers condition. The numerical solutions are being determined through the Predictor-Corrector scheme. Awareness, as a form of education that lowers the risk of danger, is reducing susceptibility and the risk of infection. We employ numerical simulations to showcase the variety of realistic parameter values that support the argument that human awareness, as a form of education, considerably lowers susceptibility and the risk of infection.

A mathematical model of Marburg virus disease outbreaks and the potential role of vaccination in control.

BACKGROUND: Marburg virus disease is an acute haemorrhagic fever caused by Marburg virus. Marburg virus is zoonotic, maintained in nature in Egyptian fruit bats, with occasional spillover infections into humans and nonhuman primates. Although rare, sporadic cases and outbreaks occur in Africa, usually associated with exposure to bats in mines or caves, and sometimes with secondary human-to-human transmission. Outbreaks outside of Africa have also occurred due to importation of infected monkeys. Although all previous Marburg virus disease outbreaks have been brought under control without vaccination, there is nevertheless the potential for large outbreaks when implementation of public health measures is not possible or breaks down. Vaccines could thus be an important additional tool, and development of several candidate vaccines is under way. METHODS: We developed a branching process model of Marburg virus transmission and investigated the potential effects of several prophylactic and reactive vaccination strategies in settings driven primarily by multiple spillover events as well as human-to-human transmission. Linelist data from the 15 outbreaks up until 2022, as well as an Approximate Bayesian Computational framework, were used to inform the model parameters. RESULTS: Our results show a low basic reproduction number which varied across outbreaks, from 0.5 [95% CI 0.05-1.8] to 1.2 [95% CI 1.0-1.9] but a high case fatality ratio. Of six vaccination strategies explored, the two prophylactic strategies (mass and targeted vaccination of high-risk groups), as well as a combination of ring and targeted vaccination, were generally most effective, with a probability of potential outbreaks being terminated within 1 year of 0.90 (95% CI 0.90-0.91), 0.89 (95% CI 0.88-0.90), and 0.88 (95% CI 0.87-0.89) compared with 0.68 (0.67-0.69) for no vaccination, especially if the outbreak is driven by zoonotic spillovers and the vaccination campaign initiated as soon as possible after onset of the first case. CONCLUSIONS: Our study shows that various vaccination strategies can be effective in helping to control outbreaks of MVD, with the best approach varying with the particular epidemiologic circumstances of each outbreak.

Filoviruses: Scientific Gaps and Prototype Pathogen Recommendation.

Viruses in the family Filoviridae, including the commonly known Ebola (EBOV) and Marburg (MARV) viruses, can cause severe hemorrhagic fever in humans and nonhuman primates. Sporadic outbreaks of filovirus disease occur in sub-Saharan Africa with reported case fatality rates ranging from 25% to 90%. The high mortality and increasing frequency and magnitude of recent outbreaks along with the increased potential for spread from rural to urban areas highlight the importance of pandemic preparedness for these viruses. Despite their designation as high-priority pathogens, numerous scientific gaps exist in critical areas. In this review, these gaps and an assessment of potential prototype pathogen candidates are presented for this important virus family.

Micro‒Global Positioning Systems for Identifying Nightly Opportunities for Marburg Virus Spillover to Humans by Egyptian Rousette Bats.

Marburg virus disease, caused by Marburg and Ravn orthomarburgviruses, emerges sporadically in sub-Saharan Africa and is often fatal in humans. The natural reservoir is the Egyptian rousette bat (ERB), which sheds virus in saliva, urine, and feces. Frugivorous ERBs discard test-bitten and partially eaten fruit, potentially leaving infectious virus behind that could be consumed by other susceptible animals or humans. Historically, 8 of 17 known Marburg virus disease outbreaks have been linked to human encroachment on ERB habitats, but no linkage exists for the other 9 outbreaks, raising the question of how bats and humans might intersect, leading to virus spillover. We used micro‒global positioning systems to identify nightly ERB foraging locations. ERBs from a known Marburg virus‒infected population traveled long distances to feed in cultivated fruit trees near homes. Our results show that ERB foraging behavior represents a Marburg virus spillover risk to humans and plausibly explains the origins of some past outbreaks.

Identification of CCZ1 as an essential lysosomal trafficking regulator in Marburg and Ebola virus infections.

Marburg and Ebola filoviruses are two of the deadliest infectious agents and several outbreaks have occurred in the last decades. Although several receptors and co-receptors have been reported for Ebola virus, key host factors remain to be elucidated. In this study, using a haploid cell screening platform, we identify the guanine nucleotide exchange factor CCZ1 as a key host factor in the early stage of filovirus replication. The critical role of CCZ1 for filovirus infections is validated in 3D primary human hepatocyte cultures and human blood-vessel organoids, both critical target sites for Ebola and Marburg virus tropism. Mechanistically, CCZ1 controls early to late endosomal trafficking of these viruses. In addition, we report that CCZ1 has a role in the endosomal trafficking of endocytosis-dependent SARS-CoV-2 infections, but not in infections by Lassa virus, which enters endo-lysosomal trafficking at the late endosome stage. Thus, we have identified an essential host pathway for filovirus infections in cell lines and engineered human target tissues. Inhibition of CCZ1 nearly completely abolishes Marburg and Ebola infections. Thus, targeting CCZ1 could potentially serve as a promising drug target for controlling infections caused by various viruses, such as SARS-CoV-2, Marburg, and Ebola.

Design and development of new inhibitors against breast cancer, Monkeypox and Marburg virus by modification of natural Fisetin via in silico and SAR studies.

The natural Fisetin and its derivatives have been shown to have effective bioactivity and strong pharmacological profile, which is continuously drawing the interest of therapeutic applications to the development of new biomolecules against Breast cancer and Monkeypox, and Marburg viral infection, while computational approaches and the study of their structure-activity relationship (SAR) are the most eloquent and reliable platform for performing their hypothetical profile renovation. So, the main perspective of this investigation is to evaluate dual function of Fisetin and its derivatives against both virus and cancerous target. First and foremost, the prediction of activity spectra for materials (PASS) valuation has provided preliminary data on the antiviral, antibacterial, antiparasitic, and anti-cancer possibilities of the mentioned compounds. According to the evidence, PASS predicted scores were shown to perform better in antineoplastic and antiviral than antibacterial, and antiparasitic efficiency; as evidenced by their higher PASS scores in antineoplastic and antiviral drug tests. Breast cancer, Monkeypox, and Marburg virus have been selected as targeted pathogens, and different in silico studies were conducted to determine the dual function of mention derivatives. The "Lipinski five rules," on the other hand, has been subjected to extensive testing for drug-like characteristics. Molecular docking against Breast cancer, Monkeypox, and Marburg virus have been accomplished after confirmation of their bioactivity. The molecular docking evaluation against targeted disease displayed re-markable binding affinity and non-bonding engagement, with most of the results indicating that derivatives are more effective than the FDA approved standard antiviral, and antineoplastic drugs. Finally, the ADMET characteristics have been computed, and they indicate that the substance is suitable to use and did not have any chance to produce adverse effects on aquatic or non-aquatic environment, as well as having a highly soluble capacity in water medium, high G.I absorption rate, with outstanding bioavailability index. Therefore, these mentioned Fisetin derivatives could be suggested as potential medication against Breast cancer and newly reported Monkeypox, and Marburg virus, and may further proceed for laboratory experiment, synthesis, and clinical trials to evaluate their practical value.

Detection of Filoviruses in Bats in Vietnam.

A new filovirus named Menglà virus was found in bats in southern China in 2015. This species has been assigned to the new genus Dianlovirus and has only been detected in China. In this article, we report the detection of filoviruses in bats captured in Vietnam. We studied 248 bats of 15 species caught in the provinces of Lai Chau and Son La in northern Vietnam and in the province of Dong Thap in the southern part of the country. Filovirus RNA was found in four Rousettus leschenaultii and one Rousettus amplexicaudatus from Lai Chau Province. Phylogenetic analysis of the polymerase gene fragment showed that three positive samples belong to Dianlovirus, and two samples form a separate clade closer to Orthomarburgvirus. An enzyme-linked immunosorbent assay showed that 9% of Rousettus, 13% of Eonycteris, and 10% of Cynopterus bats had antibodies to the glycoprotein of marburgviruses.

Contrasting effects of filamin A and B proteins in modulating filovirus entry.

Ebola (EBOV) and Marburg viruses (MARV) cause severe hemorrhagic fever associated with high mortality rates in humans. A better understanding of filovirus-host interactions that regulate the EBOV and MARV lifecycles can provide biological and mechanistic insight critical for therapeutic development. EBOV glycoprotein (eGP) and MARV glycoprotein (mGP) mediate entry into host cells primarily by actin-dependent macropinocytosis. Here, we identified actin-binding cytoskeletal crosslinking proteins filamin A (FLNa) and B (FLNb) as important regulators of both EBOV and MARV entry. We found that entry of pseudotype psVSV-RFP-eGP, infectious recombinant rVSV-eGP-mCherry, and live authentic EBOV and MARV was inhibited in filamin A knockdown (FLNaKD) cells, but was surprisingly enhanced in filamin B knockdown (FLNbKD) cells. Mechanistically, our findings suggest that differential regulation of macropinocytosis by FLNa and FLNb likely contributes to their specific effects on EBOV and MARV entry. This study is the first to identify the filamin family of proteins as regulators of EBOV and MARV entry. These findings may provide insight into the development of new countermeasures to prevent EBOV and MARV infections.