The Ebola virus (EBOV), a member of the Filoviridae family, is a highly pathogenic agent responsible for severe hemorrhagic fever in humans. Understanding the molecular mechanisms governing its replication is critical for developing effective antiviral strategies. VP35-TurboID immunosuppression coupled with quantitative mass spectrometry identified Septin9, the host GTP-binding protein which played a role in cytoskeletal regulation, as a novel interactor of VP35. Western blotting and Far-Western blotting confirmed the direct interaction and demonstrated that the C-terminal region of VP35 was the critical binding domain. Functionally, EBOV replication as well as the formation of viral inclusion bodies (VIBs) was demonstrated to be significantly suppressed by Septin9 knockdown and depletion, as shown by the EBOV minigenome (EBOV MG) and the transcription- and replication-competent virus-like particles (trVLPs) system. This study reveals that VP35 engages in a specific interaction with the GTP-binding protein Septin9, thereby impeding EBOV replication through the disruption of inclusion bodies. The overarching objective of this study is to significantly enhance our understanding about the pathogenic mechanism of EBOV and offer a robust theoretical foundation and solid empirical support for the formulation of innovative therapeutic strategies against EBOV.
Virus Replication/physiology* ; Septins/physiology* ; Humans ; Ebolavirus/physiology* ; Inclusion Bodies, Viral/metabolism* ; Viral Regulatory and Accessory Proteins/metabolism* ; Hemorrhagic Fever, Ebola/virology*

Ebola virus (EBOV) is an enveloped negative-sense RNA virus and a member of the filovirus family. Nucleoprotein (NP) expression alone leads to the formation of inclusion bodies (IBs), which are critical for viral RNA synthesis. The matrix protein, VP40, not only plays a critical role in virus assembly/budding, but also can regulate transcription and replication of the viral genome. However, the molecular mechanism by which VP40 regulates viral RNA synthesis and virion assembly/budding is unknown. Here, we show that within IBs the N-terminus of NP recruits VP40 and is required for VLP-containing NP release. Furthermore, we find four point mutations (L692A, P697A, P698A and W699A) within the C-terminal hydrophobic core of NP result in a stronger VP40-NP interaction within IBs, sequestering VP40 within IBs, reducing VP40-VLP egress, abolishing the incorporation of NC-like structures into VP40-VLP, and inhibiting viral RNA synthesis, suggesting that the interaction of N-terminus of NP with VP40 induces a conformational change in the C-terminus of NP. Consequently, the C-terminal hydrophobic core of NP is exposed and binds VP40, thereby inhibiting RNA synthesis and initiating virion assembly/budding.
Ebolavirus/physiology* ; HEK293 Cells ; HeLa Cells ; Humans ; Nucleocapsid Proteins/metabolism* ; RNA, Viral/metabolism* ; Viral Matrix Proteins/metabolism* ; Virion/metabolism* ; Virus Assembly

OBJECTIVES: This study aimed to extend an epidemiological model (SEIHFR) to analyze epidemic trends, and evaluate intervention efficacy. METHODS: SEIHFR was modified to examine disease transmission dynamics after vaccination for the Ebola outbreak. Using existing data from Liberia, sensitivity analysis of various epidemic scenarios was used to inform the model structure, estimate the basic reproduction number ℜ₀ and investigate how the vaccination could effectively change the course of the epidemic. RESULTS: If a randomized mass vaccination strategy was adopted, vaccines would be administered prophylactically or as early as possible (depending on the availability of vaccines). An effective vaccination rate threshold for Liberia was estimated as 48.74% among susceptible individuals. If a ring vaccination strategy was adopted to control the spread of the Ebola virus, vaccines would be given to reduce the transmission rate improving the tracing rate of the contact persons of an infected individual. CONCLUSION: The extended SEIHFR model predicted the total number of infected cases, number of deaths, number of recoveries, and duration of outbreaks among others with different levels of interventions such as vaccination rate. This model may be used to better understand the spread of Ebola and develop strategies that may achieve a disease-free state.
Africa, Western ; Basic Reproduction Number ; Disease Outbreaks ; Ebolavirus ; Humans ; Liberia ; Mass Vaccination ; Vaccination ; Vaccines

In this paper, we propose a window-based mechanism visualization approach as an alternative way to measure the seriousness of the difference among data-insights extracted from a composite biodata point. The approach is based on two components: undirected graph and Mosaab-metric space. The significant application of this approach is to visualize the segmented genome of a virus. We use Influenza and Ebola viruses as examples to demonstrate the robustness of this approach and to conduct comparisons. This approach can provide researchers with deep insights about information structures extracted from a segmented genome as a composite biodata point, and consequently, to capture the segmented genetic variations and diversity (variants) in composite data points.
Ebolavirus ; Genetic Variation ; Genome ; Influenza, Human

Adenoviridae ; genetics ; Ebolavirus ; genetics ; pathogenicity ; Gene Transfer Techniques ; Genetic Vectors ; therapeutic use ; Glycoproteins ; genetics ; HEK293 Cells ; Hemorrhagic Fever, Ebola ; genetics ; pathology ; virology ; Humans ; Inflammation ; genetics ; pathology ; virology ; Mucins ; genetics ; Transfection ; Viral Envelope Proteins ; genetics

PURPOSE: The goal of this study was to purify and characterize Ebola virus glycoprotein (GP)-specific IgG antibodies from hybridoma clones. MATERIALS AND METHODS: For hybridoma production, mice were injected by intramuscular-electroporation with GP DNA vaccines, and boosted with GP vaccines. The spleen cells were used for producing GP-specific hybridoma. Enzyme-linked immunosorbent assay, Western blot assay, flow cytometry, and virus-neutralizing assay were used to test the ability of monoclonal IgG antibodies to recognize GP and neutralize Ebola virus. RESULTS: Twelve hybridomas, the cell supernatants of which displayed GP-binding activity by enzyme-linked immunosorbent assay and the presence of both IgG heavy and light chains by Western blot assay, were chosen as a possible IgG producer. Among these, five clones (C36-1, D11-3, D12-1, D34-2, and E140-2) were identified to secrete monoclonal IgG antibodies. When the monoclonal IgG antibodies from the 5 clones were tested for their antigen specificity, they recognized GP in an antigen-specific and IgG dose-dependent manner. They remained reactive to GP at the lowest tested concentrations (1.953–7.8 ng/mL). In particular, IgG antibodies from clones D11-3, D12-1, and E140-2 recognized the native forms of GP expressed on the cell surface. These antibodies were identified as IgG1, IgG2a, or IgG2b kappa types and appeared to recognize the native forms of GP, but not the denatured forms of GP, as determined by Western blot assay. Despite their GP-binding activity, none of the IgG antibodies neutralized Ebola virus infection in vitro, suggesting that these antibodies are unable to neutralize Ebola virus infection. CONCLUSION: This study shows that the purified IgG antibodies from 5 clones (C36-1, D11-3, D12-1, D34-2, and E140-2) possess GP-binding activity but not Ebola virus-neutralizing activity.
Animals ; Antibodies* ; Antibody Formation ; Blotting, Western ; Clone Cells ; Ebolavirus* ; Enzyme-Linked Immunosorbent Assay ; Flow Cytometry ; Glycoproteins* ; Hemorrhagic Fever, Ebola ; Hybridomas ; Immunoglobulin G* ; In Vitro Techniques ; Mice ; Sensitivity and Specificity ; Spleen ; Vaccines ; Vaccines, DNA

Ebola virus (EBOV) is an extremely contagious pathogen first discovered in Africa associated with severe hemorrhagic disease in humans and nonhuman primates, which has resulted in at least 28 500 suspected cases and 11 300 confirmed deaths in 2014-2016 Ebola epidemic in West Africa. Rapid and sensitive detection of EBOV is the key to increasing the probability of survival and reducing infection rates in pandemic regions. Here, we report an ultrasensitive and instrument-free EBOV detection assay based on colloidal carbon immunochromatography. Carbon nanoparticle-labeled rabbit anti-EBOV-VP40 IgG were concentrated in the conjugate pad, monoclonal antibody (McAb, 4B7F9) against EBOV-VP40 and goat anti-rabbit IgG were immobilized on the nitrocellulose membrane with 2 μL/cm at a concentration of 1 mg/mL as test and control lines, respectively. Then the sample application pad, conjugate release pad, nitrocellulose membrane and absorbent pad were assembled into a lateral flow test strip. The test strip shows strong specificity against related viruses that share similar clinical symptoms and geographic range with EBOV, including marburg virus, influenza virus, yellow fever virus and dengue virus. In addition, 1 500 negative serums were tested with false-positive rate of 1.3‰ which significantly lower than that of ReEBOV™ colloidal gold test kit recommended by World Health Organization (WHO). The sensitivity of this strip was analyzed using inactivated EBOV with detection limit of 100 ng/mL (10⁶ copies/mL) which clearly higher than that of ReEBOV™ dipstick (10⁸ copies/mL). Furthermore, the strip showed excellent thermal stability characteristics in room temperature and could be as a point-of-care (POC), ultra-sensitive and specific promising candidate for EBOV serological screening in rural Africa or entry/exit ports.
Animals ; Carbon ; Ebolavirus ; Hemorrhagic Fever, Ebola ; Humans ; Nanoparticles ; Rabbits

OBJECTIVES: In light of the dramatic spread of Ebola virus in some parts of Africa and the 2014 outbreak in Nigeria, a study was conducted to evaluate bushmeat dealers' knowledge and attitudes about zoonotic infections and the risk of transmission to humans. METHODS: A cross-sectional survey was conducted in a community in Nsukka, southeast Nigeria. Hunters (n=34) and bushmeat traders (n=42) were interviewed. A semi-structured questionnaire was used to generate the data. The Fisher exact test was used to evaluate the significance of differences between these groups. RESULTS: Only 11.8% of the hunters, as compared to 35.7% of the traders, had no knowledge of possible causes of zoonotic infections (p < 0.05). However, 64.7% of the hunters, compared to 38.1% of the traders, were ignorant regarding the responsibility of public health personnel and veterinarians (p < 0.05), and 76.5% of the hunters compared to 42.9% of the traders were ignorant regarding the existence of zoonoses in Nigeria (p < 0.05). A statistically significant difference was also found between these groups regarding the risk of contracting an infection from ectoparasites (p < 0.05). The attitudes of respondents towards zoonotic diseases did not differ significantly between the groups. CONCLUSION: The level of awareness about zoonotic diseases was low in this area, underscoring the need for interventions.
Africa ; Cross-Sectional Studies ; Ebolavirus ; Humans ; Nigeria* ; Public Health ; Risk Factors* ; Surveys and Questionnaires ; Veterinarians ; Zoonoses*

OBJECTIVES: In light of the dramatic spread of Ebola virus in some parts of Africa and the 2014 outbreak in Nigeria, a study was conducted to evaluate bushmeat dealers' knowledge and attitudes about zoonotic infections and the risk of transmission to humans.METHODS: A cross-sectional survey was conducted in a community in Nsukka, southeast Nigeria. Hunters (n=34) and bushmeat traders (n=42) were interviewed. A semi-structured questionnaire was used to generate the data. The Fisher exact test was used to evaluate the significance of differences between these groups.RESULTS: Only 11.8% of the hunters, as compared to 35.7% of the traders, had no knowledge of possible causes of zoonotic infections (p < 0.05). However, 64.7% of the hunters, compared to 38.1% of the traders, were ignorant regarding the responsibility of public health personnel and veterinarians (p < 0.05), and 76.5% of the hunters compared to 42.9% of the traders were ignorant regarding the existence of zoonoses in Nigeria (p < 0.05). A statistically significant difference was also found between these groups regarding the risk of contracting an infection from ectoparasites (p < 0.05). The attitudes of respondents towards zoonotic diseases did not differ significantly between the groups.CONCLUSION: The level of awareness about zoonotic diseases was low in this area, underscoring the need for interventions.
Africa ; Cross-Sectional Studies ; Ebolavirus ; Humans ; Nigeria ; Public Health ; Risk Factors ; Surveys and Questionnaires ; Veterinarians ; Zoonoses

PURPOSE: The goal of this study was to investigate the utility of DNA vaccines encoding Ebola virus glycoprotein (GP) as a vaccine type for the production of GP-specific hybridomas and antibodies. MATERIALS AND METHODS: DNA vaccines were constructed to express Ebola virus GP. Mice were injected with GP DNA vaccines and their splenocytes were used for hybridoma production. Enzyme-linked immunosorbent assays (ELISAs), limiting dilution subcloning, antibody purification methods, and Western blot assays were used to select GP-specific hybridomas and purify monoclonal antibodies (MAbs) from the hybridoma cells. RESULTS: Twelve hybridomas, the cell supernatants of which displayed GP-binding activity, were selected by ELISA. When purified MAbs from 12 hybridomas were tested for their reactivity to GP, 11 MAbs, except for 1 MAb (from the A6-9 hybridoma) displaying an IgG2a type, were identified as IgM isotypes. Those 11 MAbs failed to recognize GP. However, the MAb from A6-9 recognized the mucin-like region of GP and remained reactive to the antigen at the lowest tested concentration (1.95 ng/mL). This result suggests that IgM-secreting hybridomas are predominantly generated by DNA vaccination. However, boosting with GP resulted in greater production of IgG-secreting hybridomas than GP DNA vaccination alone. CONCLUSION: DNA vaccination may preferentially generate IgM-secreting hybridomas, but boosting with the protein antigen can reverse this propensity. Thus, this protein boosting approach may have implications for the production of IgG-specific hybridomas in the context of the DNA vaccination platform. In addition, the purified monoclonal IgG antibodies may be useful as therapeutic antibodies for controlling Ebola virus infection.
Animals ; Antibodies ; Antibodies, Monoclonal ; Antibody Formation ; Blotting, Western ; Clinical Coding* ; DNA* ; Ebolavirus* ; Enzyme-Linked Immunosorbent Assay ; Glycoproteins* ; Hemorrhagic Fever, Ebola ; Hybridomas* ; Immunization* ; Immunoglobulin G ; Immunoglobulin M ; Mice ; Vaccination ; Vaccines, DNA*