Chapare virus (CHAPV) is an emerging arenavirus first discovered in Bolivia. Clinical cases have a high case-fatality rate and a concerning capacity for person-to-person spread. Animal models of Chapare hemorrhagic fever are needed to study pathogenesis and development of medical counter measures. Here, we present a narrowly focused study describing lethal infection of strain 13 guinea pigs with CHAPV. Animals challenged with CHAPV had progressive weight loss, lymphocytopenia, neutropenia, thrombocytopenia, hepatitis, vascular leakage, and gastrointestinal hemorrhage, resulting in uniform lethality between 7 and 16 days following challenge. This work lays the foundation for development of a small animal model of CHAPV infection. Animal models of the emerging mammarenavirus, Chapare virus (CHAPV), are needed to study pathogenesis and development of medical counter measures. Here, we present a narrowly focused study describing lethal infection of strain 13 guinea pigs with CHAPV.

Hendra virus (HeV) is a medically important, zoonotic paramyxovirus that emerged over thirty years ago which causes severe, often fatal disease in humans and animals. There are presently no approved medical countermeasures to prevent or treat human HeV disease, although many are in various stages of development. Critical to the stringent evaluation of these experimental countermeasures are nonhuman primate models of HeV disease which accurately recapitulate the pathogenesis of human infection. The continued emergence of HeV since its initial discovery in 1994 has recently expanded to include a second genotype. Although this variant HeV produced fatal equine disease, its pathogenesis and lethality are unknown in humans. Here, we investigated the pathogenesis of clinically relevant and contemporary HeV isolates from genotype 1 (HeV/Australia/Horse/2008/Redlands) and genotype 2 (HeV-var/Australia/Horse/2015/Gympie) in the African green monkey (AGM) model of henipavirus disease. AGMs challenged with HeV genotype 1 (HeV-g1) or genotype 2 (HeV-g2) isolates via the combined intranasal/intratracheal route of exposure produced divergent survival outcomes, with four of five AGMs infected with the HeV-g2 isolate surviving. All five HeV-g1 infected subjects developed acute HeV disease which accurately recapitulated HeV pathogenesis reported in humans. Our findings revealed that HeV-g2 is less pathogenic than HeV-g1 in the AGM model and suggests that HeV-g2 may be less pathogenic in humans.

Obeldesivir (ODV; GS-5245) is an orally administered ester prodrug of the parent nucleoside GS-441524 that has broad spectrum antiviral activity inhibiting viral RNA-dependent RNA polymerases. We recently showed that ODV completely protects cynomolgus macaques against lethal infection with Sudan virus when given 24 hours after parenteral exposure. Here, we report that once daily oral ODV treatment of cynomolgus and rhesus macaques for 10 days confers 80 and 100% protection, respectively, against lethal Ebola virus infection when treatment is initiated 24 hours after mucosal (conjunctival) exposure. ODV treatment delayed viral replication to abate excessive inflammation and promote adaptive immunity. For outbreak response, oral antivirals might present substantial advantages over now approved intravenous drugs, such as easy supply, storage, distribution, and administration. Furthermore, these results support the potential of ODV as an oral postexposure prophylaxis with broad spectrum activity across filoviruses.

Microtubule (MT) dynamic instability, a cycle of growth, catastrophe, shrinkage and rescue, is driven by the switching of tubulin between two structural states, one stabilised by GTP and the other by GDP. Recent work has uncovered the ancient origins of this structural switch and revealed further fundamental elements of microtubule dynamic instability, whereby switching can be brought about by a range of allosteric effectors, propagate deep within the lattice of assembled MTs, and profoundly affect MT function. Here, we review evidence for structural switching within the MT lattice and discuss current ideas about its mechanisms.

Since the emergence of Junín virus in 1953, pathogenic New World arenaviruses have remained a public health concern. These viruses, which also include Machupo virus, Guanarito virus, Sabiá virus, and Chapare virus, cause acute viral hemorrhagic fever and neurological complications, resulting in significant morbidity and mortality. Given the dearth of licensed therapeutics or vaccines against these pathogens, animal models of infection that recapitulate human manifestations of disease remain critically important to the development of efficacious medical countermeasures. Rodents and non-human primates have been successfully used to model human New World arenaviral infections, with guinea pigs, rhesus macaques, and cynomolgus macaques being the most successful models of infection for most major pathogenic New World arenaviruses. Here, we provide a highly comprehensive review of publicly reported animal models of pathogenic New World arenavirus infections, with a discussion of advantages and disadvantages for each model.

Marburg virus (MARV) has caused sporadic outbreaks of severe hemorrhagic fever in Africa in humans and nonhuman primates (NHPs) and has the potential to be used as a biological weapon. Currently, there are no licensed vaccines or therapeutics to respond to outbreaks or deliberate misuse. Vaccine and therapeutic efficacy testing against MARV requires animal models that accurately mimic human disease. In vitro testing in cell culture cannot appropriately model the complex immunological host responses required to accurately predict efficacy in humans, which will ultimately be required for licensure of a medical countermeasure (MCM). While small animal models for MARV have been valuable for dissecting disease processes and the screening of vaccine and drug candidates, there are several caveats to their use including required adaptation of the virus, lack of host-specific reagents, or the need of an immunocompromised host. Conversely, the NHP MARV disease model addresses all shortcomings of small animal models and closely recapitulates all hallmark features of human disease. As such, NHPs have served as the "gold standard" for testing filovirus MCMs and will most likely be required for regulatory approval. Here, we describe the use of NHPs for vaccine and therapeutic evaluation against MARV.

Filoviruses, including Ebola and Marburg viruses, have caused periodic outbreaks of severe hemorrhagic disease in humans and nonhuman primates (NHP), resulting in major public health emergencies primarily in endemic areas. Filovirus disease has also been exported to developed nations, where it has been equally disruptive. There are four ebolaviruses (Ebola virus, Sudan virus, Bundibugyo virus, and Taï Forest virus) and two marburgviruses [Marburg virus (MARV) and Ravn virus] known to cause disease in humans, yet vaccines and therapeutics have only been approved for Ebola virus. NHPs have long served as the "gold standard" for medical countermeasure (MCM) evaluation and will most likely be required for regulatory approval for use in humans. However, screening and refinement of MCM dosing regimens are more efficiently and ethically performed in lower-order species such as rodents. However, mouse, hamster, and guinea pig models of filovirus infection require virus adaptation to cause disease in these animal models. Nonadapted filovirus strains can be used in immune-compromised rodent systems (genetic knockouts or humanized mice), but the immune defect must be accounted for when interpreting MCM efficacy. Recently, several groups have described the use of the domestic ferret (Mustela putorius furo) as a model for several ebolaviruses using wild-type (nonadapted) virus, with disease largely reflecting what has been observed in humans and NHPs. Interestingly, no disease has been observed in ferrets challenged with MARV. Here, we describe the use of the domestic ferret for vaccine and therapeutic evaluation against ebolaviruses.

Nipah virus (NiV) is a highly pathogenic emerging viral zoonotic disease which causes severe respiratory disease and febrile encephalitis in humans. The virus is associated with a 40-70% case fatality rate and has the potential for aerosol transmission. There are currently no licensed vaccines or treatments available to combat disease. Recombinant vesicular stomatitis virus-vectored vaccines are safe and rapidly efficacious with a single dose, which makes them ideal for deployment during outbreaks and in remote areas as seen with the success of Ervebo® during the 2013-2016 Zaire ebolavirus epidemic in West Africa. In this study, we evaluated the minimum dose needed to provide protective efficacy for a single dose of rVSVΔG-NiV-G vaccine after intratracheal/intranasal or mucosal atomization device challenge with NiV. Notably, some vaccinated animals receiving low doses in both groups succumbed to disease, enabling us to elucidate systemic mechanisms of protection in survivors. We employed advanced techniques including bead-based immunoassays and ultra-high-parameter spectral flow cytometry to profile immune cell subsets including functional and intracellular targets. This work highlights the VSV-based immune mechanisms required to protect against high-risk zoonotic henipaviruses. This work was funded by US Army Medical Research Acquisition Activity contract W81XWH1910028 to TWG Vaccines and Immunotherapy (VAC)

A cocktail of human monoclonal antibodies 1C3 and 1C11 previously protected macaques from a lethal exposure to either Ebola virus (EBOV) or Sudan virus (SUDV). 1C3 is of particular interest because its paratope strongly binds with unique stoichiometry to the glycoprotein head of several orthoebolaviruses, resulting in neutralization of EBOV and SUDV. Therefore, we evaluated the protective activity of 1C3 as a standalone therapeutic in macaques exposed to either EBOV or SUDV. Two doses of 1C3 monotherapy, administered 4 and 7 days post-exposure, did not protect SUDV-exposed macaques and partially protected EBOV-exposed macaques. Notably, in a macaque that succumbed to EBOV infection, we identified two mutually exclusive escape mutations that emerged immediately after the first dose and resulted in two amino acid changes at the 1C3 binding site. We also detected a subconsensus treatment-emergent mutation likely affecting the 1C3 binding site in all three deceased SUDV-exposed macaques. Our findings highlight combination treatment with 1C11 as critical for protection, particularly against SUDV, and in vivo activity of unpartnered 1C3 as susceptible to rapid EBOV and SUDV escape under therapeutic pressure. IMPORTANCE:A cocktail of human monoclonal antibodies 1C3 and 1C11 previously protected macaques exposed to a lethal dose of either Ebola virus (EBOV) or Sudan virus (SUDV). Since the unique binding characteristics of 1C3 are of particular interest, we evaluated its protective activity as monotherapy in macaques exposed to either EBOV or SUDV. Two doses of 1C3 alone did not protect SUDV-exposed macaques and only partially protected EBOV-exposed macaques. Importantly, failure to protect was associated with the rapid emergence of previously in vitro-identified escape mutations at the 1C3 binding site, highlighting the importance of its use in combination with 1C11 for protection against fatal disease outcome and avoiding rapid EBOV and SUDV escape. Findings have broader implications for the wise use of combination-based monoclonal antibody therapeutics to improve outcomes and prevent resistance in filovirid diseases.

Sapovirus (SaV) infections have been linked with moderate-to-severe acute gastroenteritis (AGE) in animals and humans and represent a significant risk to public health. SaVs from animals including pigs, chimpanzees, and rodents have been reported to be closely related with human SaVs, indicating the possibility of cross-species transmission. Divergent SaVs have been reported in various bat species across various continents including Asia, Europe, Oceania and Africa. However, little is known about the evolutionary history of SaVs across various bat species and their zoonotic potential. In this report, we describe the findings of a surveillance study across various bat species in Nigeria. Samples were pooled and subjected to metagenomics sequencing and analyses. Nine of 57 sample pools (containing 223 rectal swabs from five bat species) had SaV reads from which we assembled a total of four complete and three near-complete (having complete coding sequences) genomes. The bat SaV (BtSaV) strains from this study formed five distinct lineages of which four represented novel genogroups. BtSaV lineages clustered mainly according to bat families, which might suggest a likely virus-host-specific evolution. The BtSaV VP1 capsid protein structure prediction confirmed three main domains (S, P1, and P2) as reported for Human SaV (HuSaV). We found that the P2 subdomain of the VP1 protein contains a degree of homology to known immunoreactive epitopes suggesting these conserved regions may be valuable for diagnostics or medical countermeasure development. This study expands our understanding of reservoir hosts, provides information on the genetic diversity and continuous evolution of SaVs in bats.

Summary: Background: Lassa fever (LF) is a zoonotic haemorrhagic disease caused by Lassa virus (LASV), which is endemic in West African countries. The multimammate rat is the main animal reservoir and its geographic range is expected to expand due to influences like climate change and land usage, and this will place larger parts of Africa at risk. We conducted preclinical development on a promising experimental vaccine that allowed its advancement into human trials. Methods: The LF vaccine is based on a vesicular stomatitis virus (VSV) vector in which the VSV glycoprotein (G) was replaced with the LASV glycoprotein complex (GPC). Earlier studies showed that this vaccine (VSVΔG-LASV-GPC) was efficacious in macaques, thus we regenerated VSVΔG-LASV-GPC using laboratory and documentation practices required to support vaccine manufacturing and human trials. The efficacy of the clinical vaccine candidate was assessed in cynomolgus macaques and more extensive immunologic analysis was performed than previously to investigate immune responses associated with protection. Findings: A single VSVΔG-LASV-GPC vaccination elicited innate, humoural and cellular immune responses, prevented development of substantial LASV viraemia, and protected animals from disease. Vaccinated macaques developed polyfunctional antibodies and serum was shown to neutralize virus expressing GPCs representative of geographically diverse LASV lineages. Interpretation: The VSVΔG-LASV-GPC clinical candidate elicited immunity that protected 10 of 10 vaccinated macaques from disease supporting its use in a clinical development program, which recently progressed to phase 2 clinical trials. Moreover, immunologic analysis showed that virus-neutralizing serum antibodies likely played a role in preventing LASV disease in vaccinated macaques. Funding: This work was supported by the Coalition for Epidemic Preparedness Innovations (CEPI), The National Institute of Allergy and Infectious Diseases (NIAID)/National Institutes of Health (NIH), The Bill and Melinda Gates Global Vaccine Accelerator Program, the Burroughs Wellcome Fund, and financial gifts and support by Nancy Zimmerman, Mark and Lisa Schwartz, and Terry and Susan Ragon.

The recent outbreak of Marburg virus (MARV) in Rwanda underscores the need for effective countermeasures against this highly fatal pathogen, with case fatality rates reaching 90

In 2018, a clinical trial of four investigational therapies for Ebola virus disease (EVD), known as the PALM trial, was conducted in the Democratic Republic of Congo. All patients received either the antiviral remdesivir (RDV) or a monoclonal antibody product: ZMapp, mAb114 (Ebanga), or REGN-EB3 (Inmazeb). The study concluded that both mAb114 and REGN-EB3 were superior to ZMapp and RDV in reducing mortality from EVD. However, the data suggested that some patients in the RDV and ZMapp groups might have been sicker at the time of treatment initiation. Here, we assessed the efficacy of each of these therapies in a uniformly lethal rhesus monkey model of EVD when treatment was initiated 5 days after Ebola exposure. Treatment with RDV, mAb114, REGN-EB3, and ZMapp each resulted in similar survival (approximately 40%). Survival was associated with circulating viral load at treatment initiation. A trend of more escape mutants in the GP1 and GP2 domains was observed for the mAb114 group. Our data show similar suboptimal efficacy of individual therapeutics in the uniformly lethal NHP model of EVD, supporting further clinical investigation of therapeutic combinations to maximize the overall therapeutic effect and improve patient outcomes, particularly for the treatment of advanced stage EVD.

Chapare virus (CHAPV) is an emerging New World arenavirus that is the causative agent of Chapare haemorrhagic fever (CHHF) responsible for recent outbreaks with alarmingly high case fatality rates in Bolivia near the Brazilian border. Here, we describe a nonhuman primate (NHP) model of CHHF infection which represents an essential tool to understand this emerging biological threat agent. Cynomolgus macaques challenged intravenously with CHAPV develop clinical disease, which recapitulates several key features of human CHHF. All subjects lost weight and had clinical scores following the CHAPV challenge. Notably, one of four NHPs developed a lethal disease with viral hepatitis and haemorrhagic features. Clinical chemistry and haematology revealed leukopoenia, anaemia, thrombocytopenia, and increased transaminase levels. In all four subjects, viremia was detectable for the first week following the challenge and viral RNA was detectable in serum and many tissues persisting 35 days post-challenge. Several medical countermeasures (MCM) have efficacy against CHAPV infection in vitro, but the current model for MCM testing and approval of new drugs is reliant on the availability of animal models. This work lays the foundation for future CHHF MCM development.

Lassa virus (LASV) causes significant human morbidity and mortality in endemic areas of West Africa. Previous studies have shown that cynomolgus macaques of non-Mauritius Asian origin best reproduce clinical features of human Lassa fever (LF). Because of the shortage of macaques caused by the COVID-19 pandemic, research on high-consequence pathogens including LASV has been severely hindered. We assessed the pathogenic potential of LASV in Mauritius-origin cynomolgus macaques (MCMs) and African green monkeys (AGMs) to find a more available alternative species to model LF. Importantly, we show similarity in transcriptomic host responses related to interferon signaling, cytokinemia, and immune cell dysregulation; however, AGMs more consistently reproduced hallmark features of LF, developing hemorrhagic manifestations closer to those seen in humans. We further show that the lethal dose 50 (LD50) of LASV in mucosally exposed AGMs is approximately 27 plaque-forming units (PFU). This low LD50 highlights the concern about the public health threat posed by LASV.

Various animal models have been established to gain a better understanding of the pathogenesis of Marburg virus (MARV) and Ravn virus (RAVV), and to develop medical countermeasures (MCMs) against them. Of these models, which range from rodents to nonhuman primates (NHPs), the macaque model most closely mimics the severe disease displayed in humans. Nevertheless, rodent models mirror many key aspects of human infection and are frequently used for the initial assessment of experimental vaccines and treatments. Due to the less restrictive housing and husbandry requirements for these models, large-scale experiments can be performed to evaluate a number of test articles and/or dosing regimens.Adaptation of MARV and RAVV by serial passaging is necessary to cause disease in immunocompetent rodent species. While mice provide limited predictive value of vaccine and therapeutic efficacy against these viruses, guinea pigs have emerged as a dependable indicator of outcomes in late-stage NHP testing. Additionally, the larger size of guinea pigs compared to mice permits more frequent and substantial blood sample collection. This chapter outlines the essential procedures to conduct intraperitoneal challenge, blood collection, and the administration of MCMs in MARV and RAVV guinea pig models using biosafety level 4 practices.

Lassa virus (LASV) is a World Health Organization (WHO) priority pathogen that causes high morbidity and mortality. Recently, we showed that a combination of three broadly neutralizing human monoclonal antibodies known as Arevirumab-3 (8.9F, 12.1F, 37.2D) based on the lineage IV Josiah strain protected 100% of cynomolgus macaques against heterologous challenge with lineage II and III strains of LASV when therapy was initiated beginning at day 8 after challenge. LASV strains from Benin and Togo represent a new lineage VII that are more genetically diverse from lineage IV than strains from lineages II and III. Here, we tested the ability of Arevirumab-3 to protect macaques against a LASV lineage VII Togo isolate when treatment was administered beginning 8 days after exposure. Unexpectedly, only 40% of treated animals survived challenge. In a subsequent study we showed that Arevirumab-3 protected 100% of macaques from lethal challenge when treatment was initiated 7 days after LASV Togo exposure. Based on our transcriptomics data, successful Arevirumab-3 treatment correlated with diminished neutrophil signatures and the predicted development of T cell responses. As the in vitro antiviral activity of Arevirumab-3 against LASV Togo was equivalent to lineage II and III strains, the reduced protection in macaques against Togo likely reflects the faster disease course of LASV Togo in macaques than other strains. This data causes concern regarding the ability of heterologous vaccines and treatments to provide cross protection against lineage VII LASV isolates.