Helminth exposure and immune response to the two-dose heterologous Ad26.ZEBOV, MVA-BN-Filo Ebola vaccine regimen.

BACKGROUND: The exposure to parasites may influence the immune response to vaccines in endemic African countries. In this study, we aimed to assess the association between helminth exposure to the most prevalent parasitic infections, schistosomiasis, soil transmitted helminths infection and filariasis, and the Ebola virus glycoprotein (EBOV GP) antibody concentration in response to vaccination with the Ad26.ZEBOV, MVA-BN-Filo vaccine regimen in African and European participants using samples obtained from three international clinical trials. METHODS/PRINCIPAL FINDINGS: We conducted a study in a subset of participants in the EBL2001, EBL2002 and EBL3001 clinical trials that evaluated the Ad26.ZEBOV, MVA-BN-Filo vaccine regimen against EVD in children, adolescents and adults from the United Kingdom, France, Burkina Faso, Cote d'Ivoire, Kenya, Uganda and Sierra Leone. Immune markers of helminth exposure at baseline were evaluated by ELISA with three commercial kits which detect IgG antibodies against schistosome, filarial and Strongyloides antigens. Luminex technology was used to measure inflammatory and activation markers, and Th1/Th2/Th17 cytokines at baseline. The association between binding IgG antibodies specific to EBOV GP (measured on day 21 post-dose 2 and on Day 365 after the first dose respectively), and helminth exposure at baseline was evaluated using a multivariable linear regression model adjusted for age and study group. Seventy-eight (21.3%) of the 367 participants included in the study had at least one helminth positive ELISA test at baseline, with differences of prevalence between studies and an increased prevalence with age. The most frequently detected antibodies were those to Schistosoma mansoni (10.9%), followed by Acanthocheilonema viteae (9%) and then Strongyloides ratti (7.9%). Among the 41 immunological analytes tested, five were significantly (p < .003) lower in participants with at least one positive helminth ELISA test result: CCL2/MCP1, FGFbasic, IL-7, IL-13 and CCL11/Eotaxin compared to participants with negative helminth ELISA tests. No significant association was found with EBOV-GP specific antibody concentration at 21 days post-dose 2, or at 365 days post-dose 1, adjusted for age group, study, and the presence of any helminth antibodies at baseline. CONCLUSIONS/SIGNIFICANCE: No clear association was found between immune markers of helminth exposure as measured by ELISA and post-vaccination response to the Ebola Ad26.ZEBOV/ MVA-BN-Filo vaccine regimen. TRIAL REGISTRATION: NCT02416453, NCT02564523, NCT02509494. ClinicalTrials.gov.

Computational and experimental identification of keystone interactions in Ebola virus matrix protein VP40 dimer formation.

The Ebola virus (EBOV) is a lipid-enveloped virus with a negative sense RNA genome that can cause severe and often fatal viral hemorrhagic fever. The assembly and budding of EBOV is regulated by the matrix protein, VP40, which is a peripheral protein that associates with anionic lipids at the inner leaflet of the plasma membrane. VP40 is sufficient to form virus-like particles (VLPs) from cells, which are nearly indistinguishable from authentic virions. Due to the restrictions of studying EBOV in BSL-4 facilities, VP40 has served as a surrogate in cellular studies to examine the EBOV assembly and budding process from the host cell plasma membrane. VP40 is a dimer where inhibition of dimer formation halts budding and formation of new VLPs as well as VP40 localization to the plasma membrane inner leaflet. To better understand VP40 dimer stability and critical amino acids to VP40 dimer formation, we integrated computational approaches with experimental validation. Site saturation/alanine scanning calculation, combined with molecular mechanics-based generalized Born with Poisson-Boltzmann surface area (MM-GB/PBSA) method and molecular dynamics simulations were used to predict the energetic contribution of amino acids to VP40 dimer stability and the hydrogen bonding network across the dimer interface. These studies revealed several previously unknown interactions and critical residues predicted to impact VP40 dimer formation. In vitro and cellular studies were then pursued for a subset of VP40 mutations demonstrating reduction in dimer formation (in vitro) or plasma membrane localization (in cells). Together, the computational and experimental approaches revealed critical residues for VP40 dimer stability in an alpha-helical interface (between residues 106-117) as well as in a loop region (between residues 52-61) below this alpha-helical region. This study sheds light on the structural origins of VP40 dimer formation and may inform the design of a small molecule that can disrupt VP40 dimer stability.

Recent advances in the treatment of Ebola disease: A brief overview.

Ebola disease (EBOD) remains a significant and ongoing threat to African countries, characterized by a mortality rate of 25% to 90% in patients with high viral load and significant transmissibility. The most recent outbreak, reported in Uganda in September 2022, was declared officially over in January 2023. However, it was caused by the Sudan Ebola virus (SUDV), a culprit species not previously reported for a decade. Since its discovery in 1976, the management of EBOD has primarily relied on supportive care. Following the devastating outbreak in West Africa from 2014 to 2016 secondary to the Zaire Ebola virus (EBOV), where over 28,000 lives were lost, dedicated efforts to find effective therapeutic agents have resulted in considerable progress in treating and preventing disease secondary to EBOV. Notably, 2 monoclonal antibodies-Ebanga and a cocktail of monoclonal antibodies, called Inmazeb-received Food and Drug Administration (FDA) approval in 2020. Additionally, multiple vaccines have been approved for EBOD prevention by various regulatory bodies, with Ervebo, a recombinant vesicular stomatitis virus-vectored vaccine against EBOV being the first vaccine to receive approval by the FDA in 2019. This review covers the key signs and symptoms of EBOD, its modes of transmission, and the principles guiding supportive care. Furthermore, it explores recent advancements in treating and preventing EBOD, highlighting the unique properties of each therapeutic agent and the ongoing progress in discovering new treatments.

Ebola virus VP35 interacts non-covalently with ubiquitin chains to promote viral replication.

Ebolavirus (EBOV) belongs to a family of highly pathogenic viruses that cause severe hemorrhagic fever in humans. EBOV replication requires the activity of the viral polymerase complex, which includes the cofactor and Interferon antagonist VP35. We previously showed that the covalent ubiquitination of VP35 promotes virus replication by regulating interactions with the polymerase complex. In addition, VP35 can also interact non-covalently with ubiquitin (Ub); however, the function of this interaction is unknown. Here, we report that VP35 interacts with free (unanchored) K63-linked polyUb chains. Ectopic expression of Isopeptidase T (USP5), which is known to degrade unanchored polyUb chains, reduced VP35 association with Ub and correlated with diminished polymerase activity in a minigenome assay. Using computational methods, we modeled the VP35-Ub non-covalent interacting complex, identified the VP35-Ub interacting surface, and tested mutations to validate the interface. Docking simulations identified chemical compounds that can block VP35-Ub interactions leading to reduced viral polymerase activity. Treatment with the compounds reduced replication of infectious EBOV in cells and in vivo in a mouse model. In conclusion, we identified a novel role of unanchored polyUb in regulating Ebola virus polymerase function and discovered compounds that have promising anti-Ebola virus activity.

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.

Translational success of fundamental virology: a VSV-vectored Ebola vaccine.

Ebola virus disease (EVD) caused by Ebola virus (EBOV) is a severe, often fatal, hemorrhagic disease. A critical component of the public health response to curb EVD epidemics is the use of a replication-competent, recombinant vesicular stomatitis virus (rVSV)-vectored Ebola vaccine, rVSVΔG-ZEBOV-GP (ERVEBO). In this Gem, we will discuss the past and ongoing development of rVSVΔG-ZEBOV-GP, highlighting the importance of basic science and the strength of public-private partnerships to translate fundamental virology into a licensed VSV-vectored Ebola vaccine.

Evaluation of fendiline treatment in VP40 system with nucleation-elongation process: a computational model of Ebola virus matrix protein assembly.

Ebola virus (EBOV) infection is threatening human health, especially in Central and West Africa. Limited clinical trials and the requirement of biosafety level-4 laboratories hinder experimental work to advance our understanding of EBOV and the evaluation of treatment. In this work, we use a computational model to study the assembly and budding process of EBOV and evaluate the effect of fendiline on these processes in the context of fluctuating host membrane lipid levels. Our results demonstrate for the first time that the assembly of VP40 filaments may follow the nucleation-elongation theory, as this mechanism is critical to maintaining a pool of VP40 dimers for the maturation and production of virus-like particles (VLPs). We further find that this nucleation-elongation process is likely influenced by fluctuating phosphatidylserine (PS), which can complicate the efficacy of lipid-targeted therapies like fendiline, a drug that lowers cellular PS levels. Our results indicate that fendiline-induced PS reduction may actually increase VLP production at earlier time points (24 h) and under low fendiline concentrations (≤2 µM). However, this effect is transient and does not change the conclusion that fendiline generally decreases VLP production. In the context of fluctuating PS levels, we also conclude that fendiline can be more efficient at the late stage of VLP budding relative to earlier phases. Combination therapy with a VLP budding step-targeted drug may therefore further increase the treatment efficiency of fendiline. Finally, we also show that fendiline-induced PS reduction more effectively lowers VLP production when VP40 expression is high. Taken together, our results provide critical quantitative information on how fluctuating lipid levels (PS) affect EBOV assembly and egress and how this mechanism can be disrupted by lipid-targeting molecules like fendiline. IMPORTANCE: Ebola virus (EBOV) infection can cause deadly hemorrhagic fever, which has a mortality rate of ~50%-90% without treatment. The recent outbreaks in Uganda and the Democratic Republic of the Congo illustrate its threat to human health. Though two antibody-based treatments were approved, mortality rates in the last outbreak were still higher than 30%. This can partly be due to the requirement of advanced medical facilities for current treatments. As a result, it is very important to develop and evaluate new therapies for EBOV infection, especially those that can be easily applied in the developing world. The significance of our research is that we evaluate the potential of lipid-targeted treatments in reducing EBOV assembly and egress. We achieved this goal using the VP40 system combined with a computational approach, which both saves time and lowers cost compared to traditional experimental studies and provides innovative new tools to study viral protein dynamics.

Bivalent VSV Vectors Mediate Rapid and Potent Protection from Andes Virus Challenge in Hamsters.

Orthohantaviruses may cause hemorrhagic fever with renal syndrome or hantavirus cardiopulmonary syndrome. Andes virus (ANDV) is the only orthohantavirus associated with human-human transmission. Therefore, emergency vaccination would be a valuable public health measure to combat ANDV-derived infection clusters. Here, we utilized a promising vesicular stomatitis virus (VSV)-based vaccine to advance the approach for emergency applications. We compared monovalent and bivalent VSV vectors containing the Ebola virus (EBOV), glycoprotein (GP), and ANDV glycoprotein precursor (GPC) for protective efficacy in pre-, peri- and post-exposure immunization by the intraperitoneal and intranasal routes. Inclusion of the EBOV GP was based on its favorable immune cell targeting and the strong innate responses elicited by the VSV-EBOV vaccine. Our data indicates no difference of ANDV GPC expressing VSV vectors in pre-exposure immunization independent of route, but a potential benefit of the bivalent VSVs following peri- and post-exposure intraperitoneal vaccination.

2020 Ebola virus disease outbreak in Équateur Province, Democratic Republic of the Congo: a retrospective genomic characterisation.

BACKGROUND: The Democratic Republic of the Congo has had 15 Ebola virus disease (EVD) outbreaks, from 1976 to 2023. On June 1, 2020, the Democratic Republic of the Congo declared an outbreak of EVD in the western Équateur Province (11th outbreak), proximal to the 2018 Tumba and Bikoro outbreak and concurrent with an outbreak in the eastern Nord Kivu Province. In this Article, we assessed whether the 11th outbreak was genetically related to previous or concurrent EVD outbreaks and connected available epidemiological and genetic data to identify sources of possible zoonotic spillover, uncover additional unreported cases of nosocomial transmission, and provide a deeper investigation into the 11th outbreak. METHODS: We analysed epidemiological factors from the 11th EVD outbreak to identify patient characteristics, epidemiological links, and transmission modes to explore virus spread through space, time, and age groups in the Équateur Province, Democratic Republic of the Congo. Trained field investigators and health professionals recorded data on suspected, probable, and confirmed cases, including demographic characteristics, possible exposures, symptom onset and signs and symptoms, and potentially exposed contacts. We used blood samples from individuals who were live suspected cases and oral swabs from individuals who were deceased to diagnose EVD. We applied whole-genome sequencing of 87 available Ebola virus genomes (from 130 individuals with EVD between May 19 and Sept 16, 2020), phylogenetic divergence versus time, and Bayesian reconstruction of phylogenetic trees to calculate viral substitution rates and study viral evolution. We linked the available epidemiological and genetic datasets to conduct a genomic and epidemiological study of the 11th EVD outbreak. FINDINGS: Between May 19 and Sept 16, 2020, 130 EVD (119 confirmed and 11 probable) cases were reported across 13 Équateur Province health zones. The individual identified as the index case reported frequent consumption of bat meat, suggesting the outbreak started due to zoonotic spillover. Sequencing revealed two circulating Ebola virus variants associated with this outbreak-a Mbandaka variant associated with the majority (97%) of cases and a Tumba-like variant with similarity to the ninth EVD outbreak in 2018. The Tumba-like variant exhibited a reduced substitution rate, suggesting transmission from a previous survivor of EVD. INTERPRETATION: Integrating genetic and epidemiological data allowed for investigative fact-checking and verified patient-reported sources of possible zoonotic spillover. These results demonstrate that rapid genetic sequencing combined with epidemiological data can inform responders of the mechanisms of viral spread, uncover novel transmission modes, and provide a deeper understanding of the outbreak, which is ultimately needed for infection prevention and control during outbreaks. FUNDING: WHO and US Centers for Disease Control and Prevention.

RNA binding motif 4 inhibits the replication of ebolavirus by directly targeting 3'-leader region of genomic RNA.

Ebola virus (EBOV) belongs to Filoviridae family possessing single-stranded negative-sense RNA genome, which is a serious threat to human health. Nowadays, no therapeutics have been proven to be successful in efficiently decreasing the mortality rate. RNA binding proteins (RBPs) are reported to participate in maintaining cell integrity and regulation of viral replication. However, little is known about whether and how RBPs participate in regulating the life cycle of EBOV. In our study, we found that RNA binding motif protein 4 (RBM4) inhibited the replication of EBOV in HEK293T and Huh-7 cells by suppressing viral mRNA production. Such inhibition resulted from the direct interaction between the RRM1 domain of RBM4 and the "CU" enrichment elements located in the PE1 and TSS of the 3'-leader region within the viral genome. Simultaneously, RBM4 could upregulate the expression of some cytokines involved in the host innate immune responses to synergistically exert its antiviral function. The findings therefore suggest that RBM4 might serve as a novel target of anti-EBOV strategy.

A Flexible, Quantitative Plasmonic-Fluor Lateral Flow Assay for the Rapid Detection of Orthoebolavirus zairense and Orthoebolavirus sudanense.

Filoviruses comprise a family of single-stranded, negative-sense RNA viruses with a significant impact on human health. Given the risk for disease outbreaks, as highlighted by the recent outbreaks across Africa, there is an unmet need for flexible diagnostic technologies that can be deployed in resource-limited settings. Herein, we highlight the use of plasmonic-fluor lateral flow assays (PF-LFA) for the rapid, quantitative detection of an Ebolavirus-secreted glycoprotein, a marker for infection. Plasmonic fluors are a class of ultrabright reporter molecules that combine engineered nanorods with conventional fluorophores, resulting in improved analytical sensitivity. We have developed a PF-LFA for Orthoebolavirus zairense (EBOV) and Orthoebolavirus sudanense (SUDV) that provides estimated limits of detection as low as 0.446 and 0.641 ng/mL, respectively. Furthermore, our assay highlights a high degree of specificity between the two viral species while also maintaining a turnaround time as short as 30 min. To highlight the utility of our PF-LFA, we demonstrate the detection of EBOV infection in non-human primates. Our PF-LFA represents an enormous step forward in the development of a robust, field-deployable assay for filoviruses.

Reverse Genetics Systems for Filoviruses.

Filoviruses are causative agents of severe hemorrhagic fevers with high case fatality rates in humans. For studies of virus biology and the subsequent development of countermeasures, reverse genetic systems, and especially those facilitating the generation of recombinant filoviruses, are indispensable. Here, we describe the generation of recombinant filoviruses from cDNA.

Sequence optimized diagnostic assay for Ebola virus detection.

Rapid pathogen identification is a critical first step in patient isolation, treatment, and controlling an outbreak. Real-time PCR is a highly sensitive and specific approach commonly used for infectious disease diagnostics. However, mismatches in the primer or probe sequence and the target organism can cause decreased sensitivity, assay failure, and false negative results. Limited genomic sequences for rare pathogens such as Ebola virus (EBOV) can negatively impact assay performance due to undiscovered genetic diversity. We previously developed and validated several EBOV assays prior to the 2013-2016 EBOV outbreak in West Africa, and sequencing EBOV Makona identified sequence variants that could impact assay performance. Here, we assessed the impact sequence mismatches have on EBOV assay performance, finding one or two primer or probe mismatches resulted in a range of impact from minimal to almost two log sensitivity reduction. Redesigning this assay improved detection of all EBOV variants tested. Comparing the performance of the new assay with the previous assays across a panel of human EBOV samples confirmed increased assay sensitivity as reflected in decreased Cq values with detection of three positive that tested negative with the original assay.

Baseline gene signatures of reactogenicity to Ebola vaccination: a machine learning approach across multiple cohorts.

Introduction: The rVSVDG-ZEBOV-GP (Ervebo®) vaccine is both immunogenic and protective against Ebola. However, the vaccine can cause a broad range of transient adverse reactions, from headache to arthritis. Identifying baseline reactogenicity signatures can advance personalized vaccinology and increase our understanding of the molecular factors associated with such adverse events. Methods: In this study, we developed a machine learning approach to integrate prevaccination gene expression data with adverse events that occurred within 14 days post-vaccination. Results and Discussion: We analyzed the expression of 144 genes across 343 blood samples collected from participants of 4 phase I clinical trial cohorts: Switzerland, USA, Gabon, and Kenya. Our machine learning approach revealed 22 key genes associated with adverse events such as local reactions, fatigue, headache, myalgia, fever, chills, arthralgia, nausea, and arthritis, providing insights into potential biological mechanisms linked to vaccine reactogenicity.

Effect of Interferon Gamma on Ebola Virus Infection of Primary Kupffer Cells and a Kupffer Cell Line.

Ebola virus disease (EVD) represents a global health threat. The etiological agents of EVD are six species of Orthoebolaviruses, with Orthoebolavirus zairense (EBOV) having the greatest public health and medical significance. EVD pathogenesis occurs as a result of broad cellular tropism of the virus, robust viral replication and a potent and dysregulated production of cytokines. In vivo, tissue macrophages are some of the earliest cells infected and contribute significantly to virus load and cytokine production. While EBOV is known to infect macrophages and to generate high titer virus in the liver, EBOV infection of liver macrophages, Kupffer cells, has not previously been examined in tissue culture or experimentally manipulated in vivo. Here, we employed primary murine Kupffer cells (KC) and an immortalized murine Kupffer cell line (ImKC) to assess EBOV-eGFP replication in liver macrophages. KCs and ImKCs were highly permissive for EBOV infection and IFN-γ polarization of these cells suppressed their permissiveness to infection. The kinetics of IFN-γ-elicited antiviral responses were examined using a biologically contained model of EBOV infection termed EBOV ΔVP30. The antiviral activity of IFN-γ was transient, but a modest ~3-fold reduction of infection persisted for as long as 6 days post-treatment. To assess the interferon-stimulated gene products (ISGs) responsible for protection, the efficacy of secreted ISGs induced by IFN-γ was evaluated and secreted ISGs failed to block EBOV ΔVP30. Our studies define new cellular tools for the study of EBOV infection that can potentially aid the development of new antiviral therapies. Furthermore, our data underscore the importance of macrophages in EVD pathogenesis and those IFN-γ-elicited ISGs that help to control EBOV infection.

Recombinant Sudan virus and evaluation of humoral cross-reactivity between Ebola and Sudan virus glycoproteins after infection or rVSV-ΔG-ZEBOV-GP vaccination.

Ebola disease outbreaks are major public health events because of human-to-human transmission and high mortality. These outbreaks are most often caused by Ebola virus, but at least three related viruses can also cause the disease. In 2022, Sudan virus re-emerged causing more than 160 confirmed and probable cases. This report describes generation of a recombinant Sudan virus and demonstrates its utility by quantifying antibody cross-reactivity between Ebola and Sudan virus glycoproteins after human infection or vaccination with a licensed Ebola virus vaccine.

Ebola virus VP35 perturbs type I interferon signaling to facilitate viral replication.

As one of the deadliest viruses, Ebola virus (EBOV) causes lethal hemorrhagic fevers in humans and nonhuman primates. The suppression of innate immunity leads to robust systemic virus replication of EBOV, leading to enhanced transmission. However, the mechanism of EBOV-host interaction is not fully understood. Here, we identified multiple dysregulated genes in early stage of EBOV infection through transcriptomic analysis, which are highly clustered to Jak-STAT signaling. EBOV VP35 and VP30 were found to inhibit type I interferon (IFN) signaling. Moreover, exogenous expression of VP35 blocks the phosphorylation of endogenous STAT1, and suppresses nuclear translocation of STAT1. Using serial truncated mutations of VP35, N-terminal 1-220 amino acid residues of VP35 were identified to be essential for blocking on type I IFN signaling. Remarkably, VP35 of EBOV suppresses type I IFN signaling more efficiently than those of Bundibugyo virus (BDBV) and Marburg virus (MARV), resulting in stable replication to facilitate the pathogenesis. Altogether, this study enriches understanding on EBOV evasion of innate immune response, and provides insights into the interplay between filoviruses and host.

Antibody responses to recombinant vesicular stomatitis virus-Zaire Ebolavirus vaccination for Ebola virus disease across doses and continents: 5-year durability.

OBJECTIVES: To report 5-year persistence and avidity of antibodies produced by the live-attenuated recombinant vesicular stomatitis virus (rVSV) expressing the Zaire Ebolavirus (ZEBOV) glycoprotein (GP), known as rVSV-ZEBOV (Ervebo®). METHODS: Healthy adults vaccinated with 300,000 or 10-50 million plaque-forming units of rVSV-ZEBOV in the WHO-coordinated trials of 2014-2015 were followed for up to 4 (Lambaréné, Gabon) and 5 (Geneva, Switzerland) years. We report seropositivity rates, geometric mean titres (GMTs), and population distribution of ZEBOV-GP ELISA IgG antibodies, neutralizing antibodies (pseudovirus and live-virus neutralization) and antibody avidity; the primary outcome was ZEBOV-GP ELISA IgG GMTs at 4 or 5 years compared with 1 year (Y1) after immunization. RESULTS: Among the 168 eligible vaccinees (Geneva: 97 and Lambaréné: 71) enrolled 1 year post-immunization, 146 (87%) remained enrolled at 4 years (Geneva: n = 88, Lambaréné: n = 58), and 84 (87%, Geneva) at 5 years post-vaccination. ZEBOV-GP ELISA IgG GMTs plateaued, with no declining trend from 1 year through the last time point assessed (1147.8 [95% CI 874.3-1507.0] at Y1 versus 1548.1 [95% CI 1136.6-2108.5] at Y5 in Geneva volunteers receiving ≥10 million plaque-forming units of rVSV-ZEBOV), their avidity matching that of ZEBOV convalescents. Live-virus neutralizing antibodies were detected for shorter periods and in fewer vaccinees (53/95 [56%] at Y1 versus 35/84 [42%] at Y5 in Geneva volunteers, all dose levels). DISCUSSION: Titres at Y1 emerged as a correlate of antibody persistence at Y5. The findings of persistent ZEBOV-GP ELISA IgG titres yet shorter-lasting, lower titres of live-virus neutralizing antibodies suggest the contribution of antibody-mediated protective mechanisms other than neutralization. Long-term clinical efficacy of rVSV-ZEBOV, however, requires further study.

Ebolavirus VP40 redux. Or rather, redox.

The virion protein 40 (VP40) forms the viral matrix, mediates budding, and downregulates viral RNA synthesis in ebolaviruses. In this issue of Structure, Werner et al. present a structure of VP40 from Sudan ebolavirus with a previously unresolved disulfide bridge that enables regulation of VP40 functions via human thioredoxin.

Molecular characterization of the 2022 Sudan virus disease outbreak in Uganda.

IMPORTANCE: Ebola disease (EBOD) is a public health threat with a high case fatality rate. Most EBOD outbreaks have occurred in remote locations, but the 2013-2016 Western Africa outbreak demonstrated how devastating EBOD can be when it reaches an urban population. Here, the 2022 Sudan virus disease (SVD) outbreak in Mubende District, Uganda, is summarized, and the genetic relatedness of the new variant is evaluated. The Mubende variant exhibited 96% amino acid similarity with historic SUDV sequences from the 1970s and a high degree of conservation throughout the outbreak, which was important for ongoing diagnostics and highly promising for future therapy development. Genetic differences between viruses identified during the Mubende SVD outbreak were linked with epidemiological data to better interpret viral spread and contact tracing chains. This methodology should be used to better integrate discrete epidemiological and sequence data for future viral outbreaks.