No Substantial Histopathologic Changes in Mops condylurus Bats Naturally Infected with Bombali Virus, Kenya.

We found similar mild perivascular inflammation in lungs of Bombali virus-positive and -negative Mops condylurus bats in Kenya, indicating the virus is well-tolerated. Our findings indicate M. condylurus bats may be a reservoir host for Bombali virus. Increased surveillance of these bats will be important to reduce potential virus spread.

Bombali Ebolavirus in Mops condylurus Bats (Molossidae), Mozambique

We detected Bombali ebolavirus RNA in 3 free-tailed bats (Mops condylurus, Molossidae) in Mozambique. Sequencing of the large protein gene revealed 98% identity with viruses previously detected in Sierra Leone, Kenya, and Guinea. Our findings further support the suspected role of Mops condylurus bats in maintaining Bombali ebolavirus

Resurgence of Ebola virus in 2021 in Guinea suggests a new paradigm for outbreaks.

Seven years after the declaration of the first epidemic of Ebola virus disease in Guinea, the country faced a new outbreak-between 14 February and 19 June 2021-near the epicentre of the previous epidemic1,2. Here we use next-generation sequencing to generate complete or near-complete genomes of Zaire ebolavirus from samples obtained from 12 different patients. These genomes form a well-supported phylogenetic cluster with genomes from the previous outbreak, which indicates that the new outbreak was not the result of a new spillover event from an animal reservoir. The 2021 lineage shows considerably lower divergence than would be expected during sustained human-to-human transmission, which suggests a persistent infection with reduced replication or a period of latency. The resurgence of Zaire ebolavirus from humans five years after the end of the previous outbreak of Ebola virus disease reinforces the need for long-term medical and social care for patients who survive the disease, to reduce the risk of re-emergence and to prevent further stigmatization.

Development and implementation of the Ebola Exposure Window Calculator: A tool for Ebola virus disease outbreak field investigations.

During an Ebola virus disease (EVD) outbreak, calculating the exposure window of a confirmed case can assist field investigators in identifying the source of infection and establishing chains of transmission. However, field investigators often have difficulty calculating this window. We developed a bilingual (English/French), smartphone-based field application to assist field investigators in determining the exposure window of an EVD case. The calculator only requires the reported date of symptoms onset and the type of symptoms present at onset or the date of death. Prior to the release of this application, there was no similar electronic capability to enable consistent calculation of EVD exposure windows for field investigators. The Democratic Republic of the Congo Ministry of Health endorsed the application and incorporated it into trainings for field staff. Available for Apple and Android devices, the calculator continues to be downloaded even as the eastern DRC outbreak resolved. We rapidly developed and implemented a smartphone application to estimate the exposure window for EVD cases in an outbreak setting.

Cyanovirin-N Binds Viral Envelope Proteins at the Low-Affinity Carbohydrate Binding Site without Direct Virus Neutralization Ability.

Glycan-targeting antibodies and pseudo-antibodies have been extensively studied for their stoichiometry, avidity, and their interactions with the rapidly modifying glycan shield of influenza A. Broadly neutralizing antiviral agents bind in the same order when they neutralize enveloped viruses regardless of the location of epitopes to the host receptor binding site. Herein, we investigated the binding of cyanovirin-N (CV-N) to surface-expressed glycoproteins such as those of human immunodeficiency virus (HIV) gp120, hemagglutinin (HA), and Ebola (GP)1,2 and compared their binding affinities with the binding response to the trimer-folded gp140 using surface plasmon resonance (SPR). Binding-site knockout variants of an engineered dimeric CV-N molecule (CVN2) revealed a binding affinity that correlated with the number of (high-) affinity binding sites. Binding curves were specific for the interaction with N-linked glycans upon binding with two low-affinity carbohydrate binding sites. This biologically active assembly of a domain-swapped CVN2, or monomeric CV-N, bound to HA with a maximum KD of 2.7 nM. All three envelope spike proteins were recognized at a nanomolar KD, whereas binding to HIV neutralizing 2G12 by targeting HA and Ebola GP1,2 was measured in the µM range and specific for the bivalent binding scheme in SPR. In conclusion, invariant structural protein patterns provide a substrate for affinity maturation in the membrane-anchored HA regions, as well as the glycan shield on the membrane-distal HA top part. They can also induce high-affinity binding in antiviral CV-N to HA at two sites, and CVN2 binding is achieved at low-affinity binding sites.

Pooled Analysis of the Accuracy of Xpert Ebola Assay for Diagnosing Ebola Virus Infection.

BACKGROUND: West Africa has witnessed the unprecedented outbreak of Ebola virus disease (EVD). The Ebola virus (EBOV) can cause Ebola hemorrhagic fever, which is documented as the most deadly viral hemorrhagic fever in the world. RT-PCR had been suggested to be employed in the detection of Ebola virus; however, this method has high requirements for laboratory equipment and takes a long time to determine Ebola infection. Although Xpert Ebola is a fast and simple instrument for the detection of Ebola virus, its effect is still unclear. This study is aimed at evaluating the accuracy of Xpert Ebola in diagnosing Ebola virus infection. METHODS: Using the keywords "Xpert" and "Ebola virus", relevant studies were retrieved from the database of PubMed, Embase, Web of Science, and Cochrane. RT-PCR was employed as a reference standard to evaluate whether the study is eligible to be included in the meta-analysis. Data from these included studies were extracted by two independent assessors and were then analyzed by the Meta-DiSc 1.4 software to produce the heterogeneity of sensitivity (SEN), specificity (SP), positive likelihood ratio (PLR), negative likelihood ratio (NLR), and diagnostic advantage ratio (DOR) of the study. The results of pooled analysis were plotted, together with the summary receiver operating characteristic (SROC) curve plotted by calculating the area under the curve (AUC). Generated pooled summary estimates (95% CIs) were calculated for the evaluation of the overall accuracy of this study. RESULTS: Five fourfold tables were made from the four studies that were included in the meta-analysis. The pooled sensitivity of Xpert Ebola was 0.98 (95% confidence interval (CI) (0.95, 0.99)), and the pooled specificity was 0.98 (95% CI (0.97, 0.99)). The pooled values of positive likelihood ratio was 53.91 (95% CI (12.82, 226.79)), with negative likelihood ratio being 0.04 (95% CI (0.02, 0.08)) and diagnostic odds ratio being 2649.45 (95% CI (629.61, 11149.02)). The AUC was 0.9961. CONCLUSIONS: Compared with RT-PCR, Xpert Ebola has high sensitivity and specificity. Therefore, it is a valued alternative method for the clinical diagnosis of Ebola virus infection. However, the Xpert Ebola test is a qualitative test that does not provide quantitative testing of EBOV concentration. Whether it can completely replace other methods or not calls for further evidences.

Baseline Asymptomatic Malaria Infection and Immunogenicity of Recombinant Vesicular Stomatitis Virus-Zaire Ebola Virus Envelope Glycoprotein.

BACKGROUND: The effect of malaria infection on the immunogenicity of the recombinant vesicular stomatitis virus-Zaire Ebola virus envelope glycoprotein (GP) vaccine (rVSVΔG-ZEBOV-GP) (ERVEBO) is unknown. METHODS: The Sierra Leone Trial to Introduce a Vaccine Against Ebola (STRIVE) vaccinated 7998 asymptomatic adults with rVSVΔG-ZEBOV-GP during the 2014-2016 Ebola epidemic. In STRIVE's immunogenicity substudy, participants provided blood samples at baseline and at 1, 6, and 9-12 months. Anti-GP binding and neutralizing antibodies were measured using validated assays. Baseline samples were tested for malaria parasites by polymerase chain reaction. RESULTS: Overall, 506 participants enrolled in the immunogenicity substudy and had ≥1 postvaccination antibody titer. Of 499 participants with a result, baseline malaria parasitemia was detected in 73 (14.6%). All GP enzyme-linked immunosorbent assay (ELISA) and plaque reduction neutralization test (PRNT) geometric mean titers (GMTs) at 1, 6, and 9-12 months were above baseline, and 94.1% of participants showed seroresponse by GP-ELISA (≥2-fold rise and ≥200 ELISA units/mL), while 81.5% showed seroresponse by PRNT (≥4-fold rise) at ≥1 postvaccination assessment. In participants with baseline malaria parasitemia, the PRNT seroresponse proportion was lower, while PRNT GMTs and GP-ELISA seroresponse and GMTs showed a trend toward lower responses at 6 and 9-12 months. CONCLUSION: Asymptomatic adults with or without malaria parasitemia had robust immune responses to rVSVΔG-ZEBOV-GP, persisting for 9-12 months. Responses in those with malaria parasitemia were somewhat lower.

Antibody affinity as a driver of signal generation in a paper-based immunoassay for Ebola virus surveillance.

During epidemics, such as the frequent and devastating Ebola virus outbreaks that have historically plagued regions of Africa, serological surveillance efforts are critical for viral containment and the development of effective antiviral therapeutics. Antibody serology can also be used retrospectively for population-level surveillance to provide a more complete estimate of total infections. Ebola surveillance efforts rely on enzyme-linked immunosorbent assays (ELISAs), which restrict testing to laboratories and are not adaptable for use in resource-limited settings. In this manuscript, we describe a paper-based immunoassay capable of detecting anti-Ebola IgG using Ebola virus envelope glycoprotein ectodomain (GP) as the affinity reagent. We evaluated seven monoclonal antibodies (mAbs) against GP-KZ52, 13C6, 4G7, 2G4, c6D8, 13F6, and 4F3-to elucidate the impact of binding affinity and binding epitope on assay performance and, ultimately, result interpretation. We used biolayer interferometry to characterize the binding of each antibody to GP before assessing their performance in our paper-based device. Binding affinity (KD) and on rate (kon) were major factors influencing the sensitivity of the paper-based immunoassay. mAbs with the best KD (3-25 nM) exhibited the lowest limits of detection (ca. µg mL-1), while mAbs with KD > 25 nM were undetectable in our device. Additionally, and most surprisingly, we determined that observed signals in paper devices were directly proportional to kon. These results highlight the importance of ensuring that the quality of recognition reagents is sufficient to support desired assay performance and suggest that the strength of an individual's immune response can impact the interpretation of assay results.

Ebola Virus Transmission Initiated by Relapse of Systemic Ebola Virus Disease.

During the 2018-2020 Ebola virus disease (EVD) outbreak in North Kivu province in the Democratic Republic of Congo, EVD was diagnosed in a patient who had received the recombinant vesicular stomatitis virus-based vaccine expressing a ZEBOV glycoprotein (rVSV-ZEBOV) (Merck). His treatment included an Ebola virus (EBOV)-specific monoclonal antibody (mAb114), and he recovered within 14 days. However, 6 months later, he presented again with severe EVD-like illness and EBOV viremia, and he died. We initiated epidemiologic and genomic investigations that showed that the patient had had a relapse of acute EVD that led to a transmission chain resulting in 91 cases across six health zones over 4 months. (Funded by the Bill and Melinda Gates Foundation and others.).

Assessing anxiety, depression and insomnia symptoms among Ebola survivors in Africa: A meta-analysis.

BACKGROUND: During health disaster events such as the current devastating havoc being inflicted on countries globally by the SARS-CoV-19 pandemic, mental health problems among survivors and frontline workers are likely concerns. However, during such health disaster events, stakeholders tend to give more precedence to the socio-economic and biomedical health consequences at the expense of mental health. Meanwhile, studies show that regardless of the kind of disaster/antecedent, all traumatic events trigger similar post-traumatic stress symptoms among survivors, families, and frontline workers. Thus, our study investigated the prevalence of anxiety, depression and insomnia symptoms among survivors of the 2014-2016 Ebola virus disease that plagued the West African sub-region. METHODS: We systematically retrieved peer-reviewed articles published between 1970 and 2019 from seven electronic databases, including Google Scholar, MEDLINE, PsychInfo, PubMed, Scopus, Springer Link, Web of Science on Ebola and post-traumatic stress disorder symptoms. A comprehensive hand search complemented this literature search. Of the 87 articles retrieved, only 13 met the inclusion criteria for this meta-analysis. RESULTS: After heterogeneity, influence, and publication bias analysis, our meta-analysis pooled proportion effects estimates showed a moderate to a high prevalence of anxiety (14%; 99% CI: 0.05-0.30), depression (15%; 99% CI: 0.11-0.21), and insomnia (22%; 99% CI: 0.13-0.36). Effect estimates ranging from (0.13; 99% CI: 0.05, 0.28) through to (0.11; 99% CI: 0.05-0.22), (0.15; 99% CI: 0.09-0.25) through to (0.13; 99% CI: 0.08-0.21) and (0.23; 99% CI: 0.11-0.41) to (0.23; 99% CI: 0.11-0.41) were respectively reported for anxiety, depression and insomnia symptoms. These findings suggest a significant amount of EVD survivors are struggling with anxiety, depression and insomnia symptoms. CONCLUSION: Our study provided the first-ever meta-analysis evidence of anxiety, depression, and insomnia symptoms among EVD survivors, and suggest that the predominant biomedical health response to regional and global health disasters should be complemented with trauma-related mental health services.

Investigation of Ebolavirus exposure in pigs presented for slaughter in Uganda.

In 2008, an outbreak of Reston ebolavirus (RESTV) in pigs in the Philippines expanded our understanding of the host range of ebolaviruses. Subsequent experimental infections with the human-pathogenic species Zaire ebolavirus (EBOV) confirmed that pigs are susceptible to African species of ebolaviruses. Pig keeping has become an increasingly important livelihood strategy throughout parts of sub-Saharan Africa, driven by increasing demand for pork. The growth in pig keeping is particularly rapid in Uganda, which has the highest per capita pork consumption in East Africa and a history of sporadic human outbreaks of Ebola virus disease (EVD). Using a systematic sampling protocol, we collected sera from 658 pigs presented for slaughter in Uganda between December 2015 and October 2016. Forty-six pigs (7%) were seropositive based on ELISA tests at two different institutions. Seropositive pigs had antibodies that bound to Sudan NP (n = 27), Zaire NP (Kikwit; n = 8) or both NPs (n = 11). Sera from 4 of the ELISA-positive pigs reacted in Western blot (EBOV NP = 1; RESTV NP = 2; both NPs = 2), and one sample had full neutralizing antibody against Sudan ebolavirus (SUDV) in virus neutralization tests. Pigs sampled in June 2016 were significantly more likely to be seropositive than pigs sampled in October 2016 (p = .03). Seropositive pigs were sourced from all regions except Western region. These observed temporal and spatial variations are suggestive of multiple introductions of ebolaviruses into the pig population in Uganda. This is the first report of exposure of pigs in Uganda to ebolaviruses and the first to employ systematic abattoir sampling for ebolavirus surveillance during a non-outbreak period. Future studies will be necessary to further define the role pigs play (if any) in ebolavirus maintenance and transmission so that potential risks can be mitigated.

Presence of Ebola virus in breast milk and risk of mother-to-child transmission: synthesis of evidence.

To help inform global guidelines on infant feeding, this systematic review synthesizes evidence related to the presence of the Ebola virus (EBOV) in breast milk and its potential risk of viral transmission to the infant when breastfeeding. We relied on a comprehensive search strategy to identify studies including women with suspected, probable, or confirmed EBOV infection, intending to breastfeed or give breast milk to an infant. Our search identified 10,454 records, and after deduplication and screening, we assessed 148 full texts. We included eight studies reporting on 10 breastfeeding mothers and their children (one mother with twins), who provided breast milk samples for assessment. EBOV was detected via RT-PCR or viral culture in seven out of ten breast milk samples. Four out of the five-breastfed infants with EBOV-positive breast milk were found positive for EBOV infection, and all of these EBOV-positive infants died. Since previous reports have detected EBOV in tears, saliva, sweat, and contaminated surfaces, with the current evidence, it is not possible to conclude with certainty that breast milk was the main route of EBOV transmission.

Ebola-negative neonates born to Ebola-infected mothers after monoclonal antibody therapy: a case series.

BACKGROUND: Few fetuses survive childbirth when the mother is positive for Ebola virus, with almost all being miscarried or stillborn, or dying shortly after birth. Before 2019, only two infants had been reported surviving past 28 days, of whom one tested positive for Ebola virus and subsequently received experimental therapies. Little is understood regarding the care of surviving neonates born to Ebola virus-positive mothers in the postnatal period and how novel anti-Ebola virus therapies might affect neonatal outcomes. METHODS: In this case series, we report on two neonates liveborn during the 2018-20 North Kivu Ebola epidemic in the Democratic Republic of the Congo who, along with their Ebola virus-positive mothers, received investigational monoclonal antibody treatment (mAB114 or REGN-EB3) as part of a randomised controlled trial (NCT03719586). FINDINGS: Both infants were born Ebola-negative and progressed well while in the Ebola Treatment Centre. Neither neonate developed evidence of Ebola virus disease during the course of the admission, and both were Ebola-negative at 21 days and remained healthy at discharge. INTERPRETATION: To our knowledge these neonates are the first documented as Ebola virus-negative at birth after being born to Ebola virus-positive mothers, and only the third and fourth neonates ever documented to have survived into infancy. Although no conclusions can be drawn from this small case series, and further research is required to investigate the neonatal effects of antibody therapies, these cases warrant review regarding whether post-delivery antibody therapy should be considered for all liveborn neonates of Ebola virus-positive mothers. In the context of a low resource setting, where survival of low-birthweight infants is poor, these cases also highlight the importance of adequate neonatal care. FUNDING: None.

Analyses of the performance of the Ebola virus disease alert management system in South Sudan: August 2018 to November 2019.

South Sudan implemented Ebola virus disease preparedness interventions aiming at preventing and rapidly containing any importation of the virus from the Democratic Republic of Congo starting from August 2018. One of these interventions was a surveillance system which included an Ebola alert management system. This study analyzed the performance of this system. A descriptive cross-sectional study of the Ebola virus disease alerts which were reported in South Sudan from August 2018 to November 2019 was conducted using both quantitative and qualitative methods. As of 30 November 2019, a total of 107 alerts had been detected in the country out of which 51 (47.7%) met the case definition and were investigated with blood samples collected for laboratory confirmation. Most (81%) of the investigated alerts were South Sudanese nationals. The alerts were identified by health workers (53.1%) at health facilities, at the community (20.4%) and by screeners at the points of entry (12.2%). Most of the investigated alerts were detected from the high-risk states of Gbudwe (46.9%), Jubek (16.3%) and Torit (10.2%). The investigated alerts commonly presented with fever, bleeding, headache and vomiting. The median timeliness for deployment of Rapid Response Team was less than one day and significantly different between the 6-month time periods (K-W = 7.7567; df = 2; p = 0.0024) from 2018 to 2019. Strengths of the alert management system included existence of a dedicated national alert hotline, case definition for alerts and rapid response teams while the weaknesses were occasional inability to access the alert toll-free hotline and lack of transport for deployment of the rapid response teams which often constrain quick response. This study demonstrates that the Ebola virus disease alert management system in South Sudan was fully functional despite the associated challenges and provides evidence to further improve Ebola preparedness in the country.

Local perspectives on Ebola during its tenth outbreak in DR Congo: A nationwide qualitative study.

BACKGROUND: The Democratic Republic of Congo (DR Congo) struggled to end the tenth outbreak of Ebola virus disease (Ebola), which appeared in North Kivu in 2018. It was reported that rumors were hampering the response effort. We sought to identify any rumors that could have influenced outbreak containment and affected prevention in unaffected areas of DR Congo. METHODS: We conducted a qualitative study in DR Congo over a period of 2 months (from August 1 to September 30, 2019) using in-depth interviews (IDIs) and focus group discussions (FGDs). The participants were recruited from five regional blocks using purposeful sampling. Both areas currently undergoing outbreaks and presently unaffected areas were included. We collected participants' opinions, views, and beliefs about the Ebola virus. The IDIs (n = 60) were performed with key influencers (schoolteachers, religious and political leaders/analysts, and Ebola-frontline workers), following a semi-structured interview guide. FGDs (n = 10) were conducted with community members. Interviews were recorded with a digital voice recorder and simultaneous note-taking. Participant responses were categorized in terms of their themes and subthemes. RESULTS: We identified 3 high-level themes and 15 subthemes (given here in parentheses): (1) inadequate knowledge of the origin or cause of Ebola (belief in a metaphysical origin, insufficient awareness of Ebola transmission via an infected corpse, interpretation of disease as God's punishment, belief in nosocomial Ebola, poor hygiene, and bathing in the Congo River). Ebola was interpreted as (2) a plot by multinational corporations (fears of genocide, Ebola understood as a biological weapon, concerns over organ trafficking, and Ebola was taken to be the result of business actions). Finally Ebola was rumored to be subject to (3) politicization (political authorities seen as ambivalent, exclusion of some community leaders from response efforts, distrust of political authorities, and distrust in the healthcare system). CONCLUSIONS: Due to the skepticism against Ebola countermeasures, it is critical to understand widespread beliefs about the disease to implement actions that will be effective, including integrating response with the unmet needs of the population.

Comparative performance study of three Ebola rapid diagnostic tests in Guinea.

BACKGROUND: The 2014/15 Ebola outbreak in West Africa resulted in 11,000 deaths and massive strain on local health systems, and the ongoing outbreak in Democratic Republic of Congo has afflicted more than 3000 people. Accurate, rapid Ebola diagnostics suitable for field deployment would enable prompt identification and effective response to future outbreaks, yet remain largely unavailable. The purpose of this study was to assess the accuracy of three novel rapid diagnostic tests (RDTs): an Ebola, an Ebola-Malaria, and a Fever Panel test that includes Ebola, all from a single manufacturer. METHODS: We evaluated the three RDTs in 109 Ebola-positive and 96 Ebola-negative stored serum samples collected during the outbreak in Guinea in 2014/15, and tested by real-time polymerase chain reaction (RT-PCR). Sensitivity, specificity, and overall percent agreement were calculated for each RDT using RT-PCR as a reference standard, stratified by Ct value ranges. RESULTS: All tests performed with high accuracy on samples with low Ct value (high viral load). The Fever Panel test performed with the highest accuracy, with a sensitivity of 89.9% and specificity of 90.6%. The Ebola and Ebola-Malaria tests performed comparably to each other: sensitivity was 77.1 and 78% respectively, and specificity was 91.7% for the Ebola test and 95.8% for the Ebola-Malaria test. CONCLUSIONS: This study evaluated the accuracy of three novel rapid diagnostic tests for Ebola. The tests may have significant public health relevance, particularly the Fever Panel test, which detects seven pathogens including Ebola. Given limitations to the study resulting from uncertain sample quality, further evaluation is warranted. All tests performed with highest accuracy on samples with low Ct value (high viral load), and the data presented here suggests that these RDTs may be useful for point-of-care diagnosis of cases in the context of an outbreak. Restrictions to their use in non-severe Ebola cases or for longitudinal monitoring, when viral loads are lower, may be appropriate. Highlighting the challenge in developing and evaluating Ebola RDTs, there were concerns regarding sample integrity and reference testing, and there is a need for additional research to validate these assays.