Recombinant Lloviu virus as a model to study inaccessible zoonotic viruses

Next generation sequencing has revealed the presence of many RNA viruses in animal reservoir hosts, including many closely related to known human pathogens. Despite their zoonotic potential, many of these viruses remain understudied due to not yet being cultured. While reverse genetic systems can facilitate virus rescue, this is often hindered by missing viral genome ends. A prime example is Lloviu virus (LLOV), an uncultured filovirus that is closely related to the highly pathogenic Ebola virus. Using minigenome systems, we complemented the missing LLOV genomic ends and identified cis-acting elements required for LLOV replication that were lacking in the published sequence. We leveraged these data to generate recombinant full-length LLOV clones and rescue infectious virus. Recombinant LLOV (rLLOV) displays typical filovirus features, as shown by electron microscopy. Known target cells of Ebola virus, including macrophages and hepatocytes, are permissive to rLLOV infection, suggesting that humans could be potential hosts. However, inflammatory responses in human macrophages, a hallmark of Ebola virus disease, are not induced by rLLOV. We also used rLLOV to test antivirals targeting multiple facets of the replication cycle. Rescue of uncultured viruses of pathogenic concern represents a valuable tool in our arsenal against pandemic preparedness.

Download Full-text

Extinction pathways and outbreak vulnerability in a stochastic Ebola model

Journal of The Royal Society Interface ◽

10.1098/rsif.2016.0847 ◽

2017 ◽

Vol 14 (127) ◽

pp. 20160847 ◽

Cited By ~ 15

Author(s):

Garrett T. Nieddu ◽

Lora Billings ◽

James H. Kaufman ◽

Eric Forgoston ◽

Simone Bianco

Keyword(s):

Ebola Virus ◽

Reproduction Number ◽

Virus Disease ◽

Zoonotic Diseases ◽

Animal Reservoir ◽

Human Host ◽

Zoonotic Viruses ◽

Disease Agent ◽

Dynamic Population ◽

Modelling Approach

A zoonotic disease is a disease that can be passed from animals to humans. Zoonotic viruses may adapt to a human host eventually becoming endemic in humans, but before doing so punctuated outbreaks of the zoonotic virus may be observed. The Ebola virus disease (EVD) is an example of such a disease. The animal population in which the disease agent is able to reproduce in sufficient number to be able to transmit to a susceptible human host is called a reservoir. There is little work devoted to understanding stochastic population dynamics in the presence of a reservoir, specifically the phenomena of disease extinction and reintroduction. Here, we build a stochastic EVD model and explicitly consider the impacts of an animal reservoir on the disease persistence. Our modelling approach enables the analysis of invasion and fade-out dynamics, including the efficacy of possible intervention strategies. We investigate outbreak vulnerability and the probability of local extinction and quantify the effective basic reproduction number. We also consider the effects of dynamic population size. Our results provide an improved understanding of outbreak and extinction dynamics in zoonotic diseases, such as EVD.

Download Full-text

Protective population behavior change in outbreaks of emerging infectious disease

BMC Infectious Diseases ◽

10.1186/s12879-021-06299-x ◽

2021 ◽

Vol 21 (1) ◽

Author(s):

Evans K. Lodge ◽

Annakate M. Schatz ◽

John M. Drake

Keyword(s):

Behavior Change ◽

Ebola Virus ◽

Virus Disease ◽

Population Level ◽

Emerging Infections ◽

Emerging Pathogens ◽

Susceptible Population ◽

Zoonotic Infections ◽

Zoonotic Viruses ◽

And Behavior

Abstract Background During outbreaks of emerging and re-emerging infections, the lack of effective drugs and vaccines increases reliance on non-pharmacologic public health interventions and behavior change to limit human-to-human transmission. Interventions that increase the speed with which infected individuals remove themselves from the susceptible population are paramount, particularly isolation and hospitalization. Ebola virus disease (EVD), Severe Acute Respiratory Syndrome (SARS), and Middle East Respiratory Syndrome (MERS) are zoonotic viruses that have caused significant recent outbreaks with sustained human-to-human transmission. Methods This investigation quantified changing mean removal rates (MRR) and days from symptom onset to hospitalization (DSOH) of infected individuals from the population in seven different outbreaks of EVD, SARS, and MERS, to test for statistically significant differences in these metrics between outbreaks. Results We found that epidemic week and viral serial interval were correlated with the speed with which populations developed and maintained health behaviors in each outbreak. Conclusions These findings highlight intrinsic population-level changes in isolation rates in multiple epidemics of three zoonotic infections with established human-to-human transmission and significant morbidity and mortality. These data are particularly useful for disease modelers seeking to forecast the spread of emerging pathogens.

Download Full-text

Peripheral Neuronopathy Associated With Ebola Virus Infection in Rhesus Macaques: A Possible Cause of Neurological Signs and Symptoms in Human Ebola Patients

The Journal of Infectious Diseases ◽

10.1093/infdis/jiaa304 ◽

2020 ◽

Vol 222 (10) ◽

pp. 1745-1755

Author(s):

David X Liu ◽

Donna L Perry ◽

Timothy K Cooper ◽

Louis M Huzella ◽

Randy J Hart ◽

...

Keyword(s):

Ebola Virus ◽

Neuronal Degeneration ◽

Virus Disease ◽

Rhesus Macaques ◽

Signs And Symptoms ◽

Ebola Virus Disease ◽

Target Cells ◽

Neurological Signs ◽

Enteric Ganglia ◽

Transmission Electron

Abstract Neurological signs and symptoms are the most common complications of Ebola virus disease. However, the mechanisms underlying the neurologic manifestations in Ebola patients are not known. In this study, peripheral ganglia were collected from 12 rhesus macaques that succumbed to Ebola virus (EBOV) disease from 5 to 8 days post exposure. Ganglionitis, characterized by neuronal degeneration, necrosis, and mononuclear leukocyte infiltrates, was observed in the dorsal root, autonomic, and enteric ganglia. By immunohistochemistry, RNAscope in situ hybridization, transmission electron microscopy, and confocal microscopy, we confirmed that CD68+ macrophages are the target cells for EBOV in affected ganglia. Further, we demonstrated that EBOV can induce satellite cell and neuronal apoptosis and microglial activation in infected ganglia. Our results demonstrate that EBOV can infect peripheral ganglia and results in ganglionopathy in rhesus macaques, which may contribute to the neurological signs and symptoms observed in acute and convalescent Ebola virus disease in human patients.

Download Full-text

Macaque Monoclonal Antibodies Targeting Novel Conserved Epitopes within Filovirus Glycoprotein

Journal of Virology ◽

10.1128/jvi.02172-15 ◽

2015 ◽

Vol 90 (1) ◽

pp. 279-291 ◽

Cited By ~ 48

Author(s):

Zhen-Yong Keck ◽

Sven G. Enterlein ◽

Katie A. Howell ◽

Hong Vu ◽

Sergey Shulenin ◽

...

Keyword(s):

Monoclonal Antibodies ◽

Nonhuman Primates ◽

Ebola Virus ◽

Virus Disease ◽

Protective Efficacy ◽

Hemorrhagic Fever ◽

Marburg Virus ◽

Human Pathogens ◽

Content Type ◽

The Core

ABSTRACTFiloviruses cause highly lethal viral hemorrhagic fever in humans and nonhuman primates. Current immunotherapeutic options for filoviruses are mostly specific to Ebola virus (EBOV), although other members ofFiloviridaesuch as Sudan virus (SUDV), Bundibugyo virus (BDBV), and Marburg virus (MARV) have also caused sizeable human outbreaks. Here we report a set of pan-ebolavirus and pan-filovirus monoclonal antibodies (MAbs) derived from cynomolgus macaques immunized repeatedly with a mixture of engineered glycoproteins (GPs) and virus-like particles (VLPs) for three different filovirus species. The antibodies recognize novel neutralizing and nonneutralizing epitopes on the filovirus glycoprotein, including conserved conformational epitopes within the core regions of the GP1 subunit and a novel linear epitope within the glycan cap. We further report the first filovirus antibody binding to a highly conserved epitope within the fusion loop of ebolavirus and marburgvirus species. One of the antibodies binding to the core GP1 region of all ebolavirus species and with lower affinity to MARV GP cross neutralized both SUDV and EBOV, the most divergent ebolavirus species. In a mouse model of EBOV infection, this antibody provided 100% protection when administered in two doses and partial, but significant, protection when given once at the peak of viremia 3 days postinfection. Furthermore, we describe novel cocktails of antibodies with enhanced protective efficacy compared to individual MAbs. In summary, the present work describes multiple novel, cross-reactive filovirus epitopes and innovative combination concepts that challenge the current therapeutic models.IMPORTANCEFiloviruses are among the most deadly human pathogens. The 2014-2015 outbreak of Ebola virus disease (EVD) led to more than 27,000 cases and 11,000 fatalities. While there are five species ofEbolavirusand several strains of marburgvirus, the current immunotherapeutics primarily target Ebola virus. Since the nature of future outbreaks cannot be predicted, there is an urgent need for therapeutics with broad protective efficacy against multiple filoviruses. Here we describe a set of monoclonal antibodies cross-reactive with multiple filovirus species. These antibodies target novel conserved epitopes within the envelope glycoprotein and exhibit protective efficacy in mice. We further present novel concepts for combination of cross-reactive antibodies against multiple epitopes that show enhanced efficacy compared to monotherapy and provide complete protection in mice. These findings set the stage for further evaluation of these antibodies in nonhuman primates and development of effective pan-filovirus immunotherapeutics for use in future outbreaks.

Download Full-text

Ebola virus disease: epidemiology, clinical feature and the way forward

International Journal of Community Medicine and Public Health ◽

10.18203/2394-6040.ijcmph20171744 ◽

2017 ◽

Vol 4 (5) ◽

pp. 1372 ◽

Cited By ~ 1

Author(s):

Lawan Gana Balami ◽

Suriani Ismail ◽

Saliluddin S. M. ◽

Garba S. H.

Keyword(s):

Ebola Virus ◽

Virus Disease ◽

Hypovolemic Shock ◽

Multiple Organ ◽

Ebola Virus Disease ◽

Clinical Feature ◽

Human Pathogens ◽

Disease Epidemiology ◽

Global Threat ◽

The Way

The Ebola virus disease is a zoonotic, acute viral syndrome which occurs by infection with one the strains of the Ebola virus. It is primarily endemic in Africa however the recent outbreak in the year 2014 spanned from West Africa all the way to Europe and America. This shows the virus possess a global threat and should not be considered localized to only certain parts of the world. The social and economic impact of zoonotic diseases today is high as 80% of human pathogens are of zoonotic origin. Human to human transmission happens when there is contact with bodily fluids of infected humans during the infectious phase of the disease. This spread could be through nosocomial means or community spread. Poor knowledge of the syndrome among health care workers coupled with lack of funding and deficient resources has crippled their ability to diagnose and break the chain of transmission of the disease at its early stages. The virus undergoes pathogenesis by immune evasion, immune suppression, coagulopathy, and hypovolemic shock, multiple organ failure and death in up to 90% of cases. The unavailability of a cure or vaccine for this syndrome makes it a recurrent threat due to high risk behavior practiced in endemic countries such as bush meat consumption. Thus this study gives the reader a review of current literature on this deadly disease with the aim of increasing knowledge and aiding its prevention and control.

Download Full-text

Ebola and Other Filoviruses

10.1093/med/9780190604813.003.0002 ◽

2018 ◽

Author(s):

Lisa M. Bebell

Keyword(s):

Ebola Virus ◽

Inflammatory Responses ◽

Virus Disease ◽

Treatment Strategies ◽

Disease Outbreaks ◽

Marburg Virus ◽

Residual Effects ◽

Long Term Outcomes ◽

Pediatric Vaccination

Congenital and pediatric Ebola virus disease (EVD) and Marburg virus disease (MVD) are severe, even lethal infections. Historically, children have been underrepresented in filovirus disease outbreaks, and evidence-based treatment strategies are lacking. Existing data suggest that case fatalities are highest among children under four years of age, which is partially explained by higher virus concentrations in young children. Prevention and aggressive resuscitation, nutrition, and supportive care are the mainstays of management until filovirus-specific therapies can be developed. Differences in pediatric immune and inflammatory responses may necessitate unique approaches to pediatric vaccination and treatment. There are minimal safety or immunogenicity data in children, a crucial knowledge gap that must be addressed in future trials. Studying pediatric survivors of the 2014–2016 West Africa EVD outbreak will provide much-needed data on long-term outcomes and residual effects of filovirus disease while we await effective filovirus-specific vaccines and therapies.

Download Full-text

The Tetherin Antagonism of the Ebola Virus Glycoprotein Requires an Intact Receptor-Binding Domain and Can Be Blocked by GP1-Specific Antibodies

Journal of Virology ◽

10.1128/jvi.01563-16 ◽

2016 ◽

Vol 90 (24) ◽

pp. 11075-11086 ◽

Cited By ~ 12

Author(s):

Constantin Brinkmann ◽

Inga Nehlmeier ◽

Kerstin Walendy-Gnirß ◽

Julia Nehls ◽

Mariana González Hernández ◽

...

Keyword(s):

Receptor Binding ◽

Ebola Virus ◽

Virus Disease ◽

Human Cells ◽

Binding Domain ◽

Cell Protein ◽

Target Cells ◽

Receptor Binding Domain ◽

Northern Spain ◽

Tetherin Antagonism

ABSTRACT The glycoprotein of Ebola virus (EBOV GP), a member of the family Filoviridae , facilitates viral entry into target cells. In addition, EBOV GP antagonizes the antiviral activity of the host cell protein tetherin, which may otherwise restrict EBOV release from infected cells. However, it is unclear how EBOV GP antagonizes tetherin, and it is unknown whether the GP of Lloviu virus (LLOV), a filovirus found in dead bats in Northern Spain, also counteracts tetherin. Here, we show that LLOV GP antagonizes tetherin, indicating that tetherin may not impede LLOV spread in human cells. Moreover, we demonstrate that appropriate processing of N-glycans in tetherin/GP-coexpressing cells is required for tetherin counteraction by EBOV GP. Furthermore, we show that an intact receptor-binding domain (RBD) in the GP1 subunit of EBOV GP is a prerequisite for tetherin counteraction. In contrast, blockade of Niemann-Pick disease type C1 (NPC1), a cellular binding partner of the RBD, did not interfere with tetherin antagonism. Finally, we provide evidence that an antibody directed against GP1, which protects mice from a lethal EBOV challenge, may block GP-dependent tetherin antagonism. Our data, in conjunction with previous reports, indicate that tetherin antagonism is conserved among the GPs of all known filoviruses and demonstrate that the GP1 subunit of EBOV GP plays a central role in tetherin antagonism. IMPORTANCE Filoviruses are reemerging pathogens that constitute a public health threat. Understanding how Ebola virus (EBOV), a highly pathogenic filovirus responsible for the 2013-2016 Ebola virus disease epidemic in western Africa, counteracts antiviral effectors of the innate immune system might help to define novel targets for antiviral intervention. Similarly, determining whether Lloviu virus (LLOV), a filovirus detected in bats in northern Spain, is inhibited by innate antiviral effectors in human cells might help to determine whether the virus constitutes a threat to humans. The present study shows that LLOV, like EBOV, counteracts the antiviral effector protein tetherin via its glycoprotein (GP), suggesting that tetherin does not pose a defense against LLOV spread in humans. Moreover, our work identifies the GP1 subunit of EBOV GP, in particular an intact receptor-binding domain, as critical for tetherin counteraction and provides evidence that antibodies directed against GP1 can interfere with tetherin counteraction.

Download Full-text

A Polymorphism within the Internal Fusion Loop of the Ebola Virus Glycoprotein Modulates Host Cell Entry

Journal of Virology ◽

10.1128/jvi.00177-17 ◽

2017 ◽

Vol 91 (9) ◽

Cited By ~ 17

Author(s):

Markus Hoffmann ◽

Lisa Crone ◽

Erik Dietzel ◽

Jennifer Paijo ◽

Mariana González-Hernández ◽

...

Keyword(s):

Amino Acid ◽

West Africa ◽

Cell Lines ◽

Ebola Virus ◽

Virus Disease ◽

Cell Entry ◽

Target Cells ◽

Fusion Loop ◽

Entry Efficiency ◽

Acid Exchange

ABSTRACT The large scale of the Ebola virus disease (EVD) outbreak in West Africa in 2013-2016 raised the question whether the host cell interactions of the responsible Ebola virus (EBOV) strain differed from those of other ebolaviruses. We previously reported that the glycoprotein (GP) of the virus circulating in West Africa in 2014 (EBOV2014) exhibited reduced ability to mediate entry into two nonhuman primate (NHP)-derived cell lines relative to the GP of EBOV1976. Here, we investigated the molecular determinants underlying the differential entry efficiency. We found that EBOV2014-GP-driven entry into diverse NHP-derived cell lines, as well as human monocyte-derived macrophages and dendritic cells, was reduced compared to EBOV1976-GP, although entry into most human- and all bat-derived cell lines tested was comparable. Moreover, EBOV2014 replication in NHP but not human cells was diminished relative to EBOV1976, suggesting that reduced cell entry translated into reduced viral spread. Mutagenic analysis of EBOV2014-GP and EBOV1976-GP revealed that an amino acid polymorphism in the receptor-binding domain, A82V, modulated entry efficiency in a cell line-independent manner and did not account for the reduced EBOV2014-GP-driven entry into NHP cells. In contrast, polymorphism T544I, located in the internal fusion loop in the GP2 subunit, was found to be responsible for the entry phenotype. These results suggest that position 544 is an important determinant of EBOV infectivity for both NHP and certain human target cells. IMPORTANCE The Ebola virus disease outbreak in West Africa in 2013 entailed more than 10,000 deaths. The scale of the outbreak and its dramatic impact on human health raised the question whether the responsible virus was particularly adept at infecting human cells. Our study shows that an amino acid exchange, A82V, that was acquired during the epidemic and that was not observed in previously circulating viruses, increases viral entry into diverse target cells. In contrast, the epidemic virus showed a reduced ability to enter cells of nonhuman primates compared to the virus circulating in 1976, and a single amino acid exchange in the internal fusion loop of the viral glycoprotein was found to account for this phenotype.

Download Full-text

Bayesian uncertainty quantification for transmissibility of influenza, norovirus and Ebola using information geometry

Journal of The Royal Society Interface ◽

10.1098/rsif.2016.0279 ◽

2016 ◽

Vol 13 (121) ◽

pp. 20160279 ◽

Cited By ~ 7

Author(s):

Thomas House ◽

Ashley Ford ◽

Shiwei Lan ◽

Samuel Bilson ◽

Elizabeth Buckingham-Jeffery ◽

...

Keyword(s):

Inverse Problem ◽

Ebola Virus ◽

Virus Disease ◽

Information Geometry ◽

Disease Dynamics ◽

Human Pathogens ◽

Live Virus ◽

Viral Shedding ◽

Monte Carlo Algorithms ◽

Over Time

Infectious diseases exert a large and in many contexts growing burden on human health, but violate most of the assumptions of classical epidemiological statistics and hence require a mathematically sophisticated approach. Viral shedding data are collected during human studies—either where volunteers are infected with a disease or where existing cases are recruited—in which the levels of live virus produced over time are measured. These have traditionally been difficult to analyse due to strong, complex correlations between parameters. Here, we show how a Bayesian approach to the inverse problem together with modern Markov chain Monte Carlo algorithms based on information geometry can overcome these difficulties and yield insights into the disease dynamics of two of the most prevalent human pathogens—influenza and norovirus—as well as Ebola virus disease.

Download Full-text

Suppressed anti-inflammatory heat shock response in high-risk COVID-19 patients: lessons from basic research (inclusive bats), light on conceivable therapies

Clinical Science ◽

10.1042/cs20200596 ◽

2020 ◽

Vol 134 (15) ◽

pp. 1991-2017 ◽

Cited By ~ 2

Author(s):

Thiago Gomes Heck ◽

Mirna Stela Ludwig ◽

Matias Nunes Frizzo ◽

Alberto Antonio Rasia-Filho ◽

Paulo Ivo Homem de Bittencourt

Keyword(s):

Heat Shock ◽

Heat Shock Response ◽

Inflammatory Responses ◽

Fatal Outcome ◽

Basic Research ◽

Target Cells ◽

Cytokine Storm ◽

Zoonotic Viruses ◽

Shock Response ◽

Anti Inflammatory

Abstract The major risk factors to fatal outcome in COVID-19 patients, i.e., elderliness and pre-existing metabolic and cardiovascular diseases (CVD), share in common the characteristic of being chronic degenerative diseases of inflammatory nature associated with defective heat shock response (HSR). The molecular components of the HSR, the principal metabolic pathway leading to the physiological resolution of inflammation, is an anti-inflammatory biochemical pathway that involves molecular chaperones of the heat shock protein (HSP) family during homeostasis-threatening stressful situations (e.g., thermal, oxidative and metabolic stresses). The entry of SARS coronaviruses in target cells, on the other hand, aggravates the already-jeopardized HSR of this specific group of patients. In addition, cellular counterattack against virus involves interferon (IFN)-mediated inflammatory responses. Therefore, individuals with impaired HSR cannot resolve virus-induced inflammatory burst physiologically, being susceptible to exacerbated forms of inflammation, which leads to a fatal “cytokine storm”. Interestingly, some species of bats that are natural reservoirs of zoonotic viruses, including SARS-CoV-2, possess an IFN-based antiviral inflammatory response perpetually activated but do not show any sign of disease or cytokine storm. This is possible because bats present a constitutive HSR that is by far (hundreds of times) more intense and rapid than that of human, being associated with a high core temperature. Similarly in humans, fever is a physiological inducer of HSR while antipyretics, which block the initial phase of inflammation, impair the resolution phase of inflammation through the HSR. These findings offer a rationale for the reevaluation of patient care and fever reduction in SARS, including COVID-19.

Download Full-text