Introduction: Since the first outbreaks of Ebola virus disease 1976 in the Democratic Republic of the Congo (DRC) and Sudan there has been recurrent epidemics. The biggest outbreak so far hit West Africa between 2013-2016 when at least 28 600 people were estimated to have been infected and approximately 11 300 died in the most severely affected countries Guinea, Sierra Leone and Liberia. Ebola hemorrhagic fever (EHF) or Ebola virus disease (EVD) is caused by a virus that together with the Marburg virus belongs to the Filovirus family. The Ebola virus can cause hemorrhagic bleeding, has a rapid disease progression rate and a high mortality (25-90%). The symptoms are initially headache, fever, sore throat and muscle pain but during the latter part of the disease the characteristic bleedings from all bodily orifices occurs. Many internal organs do damage and survivors often have persistent sequels and long convalescense. EVD is a zoonosis and might infect antelopes, monkeys, porcupines among others. The natural reservoir is believed to be different species of bats. In august 2018, another outbreak of Ebola was reported in DRC. This was the tenth time the disease hit the country. However, for the first time there was candidate vaccine in clinical trial that could be used during an emergency. Experts within the WHO recommended that vaccination was initiated using Merck`s Ebolavaccine rVSV-∆G-ZEBOV-GP that was ready for a phase III clinical trial. This vaccine had already been used during the West African outbreak and had been known to provide a good safety profile as well as immunogenicity. The second vaccine to be used was a two-dose heterologous prime-boost vaccine regimen called Ad26.ZEBOV/MVA-BN-Filo which was marketed by Johnson&Johnson. Ad26.ZEBOV was given as a first dose to prime the immune system and then MVA-BN-Filo was administered as a second dose approximately 8 weeks later as a booster. This vaccine regimen had also been showing promising results in phase I and II clinical trials. Objective: This degree project was made in attempt to answer the issue: How effective and safe are the new Ebola vaccines against EVD? Methods: To answer this question, a literature study was conducted. The outbreak in DRC was ongoing by the time of this degree project and there were no phase III trials conducted for the Ad26.ZEBOV/MVA-BN-Filo vaccine regimen. A lot of information was therefore retrieved from WHO, Doctors without borders, and so on. Merck`s vaccine had been prequalified and received the generic name Ervebo in the fall of 2018. Randomised clinical trials (RCT) and clinical trials regarding Ervebo was downloaded from the internet using the search engine PubMed that`s accessing different databases. Results: Both Ervebo and the 2-dose vaccine regimen Ad26.ZEBOV/MVA-BN-Filo initiated an immune response against ebolavirus that was almost 100%. The most frequent solicited local and systemic adverse events were injection site pain and headache, myalgia and fever. All adverse events were reported to be mild and resolved in a short period of time. No severe adverse events were reported. Discussion: The biggest challenge in the future will concern how to initiate vaccination strategies in countries with a lack of financial resources and infrastructure as well as ongoing armed conflicts (mainly DRC). Conclusion: The new vaccines against Ebola virus infection are all effective with a good safety profile. / Bakgrund: Sedan de första utbrotten av ebolafeber 1976 i Demokratiska republiken Kongo (DRK) och Sudan har epidemier blossat upp med jämna mellanrum. Det hittills största utbrottet drabbade Västafrika 2013-2016 då minst 28 600 beräknades ha smittats och minst 11 300 avled i de hårdast drabbade länderna Guinea, Sierra Leone och Liberia. Ebolafeber, eller ebola hemorragisk feber, orsakas av ett virus som tillsammans med Marburgviruset hör till familjen Filovirus. Ebolaviruset kan orsaka hemorragiska blödningar, har snabbt sjukdomsförlopp och hög dödlighet (25-90%). Symptomen är inledningsvis influensaliknande men under den senare delen av sjukdomsförloppet förekommer de karaktäristiska blödningarna från alla kroppsöppningar. Många inre organ tar skada och överlevare har ofta bestående men och lång konvalescens. Ebolafeber är en zoonos och kan drabba antiloper, olika sorters apor m fl. Den naturliga reservoaren tros vara olika arter av fladdermöss. I augusti 2018 rapporterades om ännu ett utbrott av ebolafeber i DRK, det 10:e i ordningen. En avgörande skillnad vid detta utbrott var att det nu fanns kandidatvaccin i kliniska studier att tillgå. En expertgrupp inom WHO rekommenderade att vaccination inledddes med rVSV-∆G-ZEBOV, ett ebolavaccin tillverkat av Merck redo för kliniska studier fas III. Detta vaccin hade använts redan under utbrottet i Västafrika och visat goda resultat vad gällde immunogenicitet och säkerhet. Det andra vaccinet att sättas in var Ad26.ZEBOV/MVA-BN-Filo, tillhandahållet av Johnson&Johnson. Detta bestod av två olika doser som gavs med ca 8 veckors mellanrum, ofta kallat ”prime/boost”-vaccin. Även för Ad26.ZEBOV/MVA-BN-Filo fanns dokumenterade positiva resultat. Syfte: Detta examensarbetet gjordes med syftet att svara på frågeställningen: Hur effektiva och säkra är de nya vaccinen mot ebolafeber? Metod: För att svara på frågan gjordes en litteraturstudie. När detta examensarbete författades pågick fortfarande utbrottet i DRK och fas III studierna för Ad26.ZEBOV/MVA-BN-Filo var inte avslutade. Mycket information fick hämtas från olika hemsidor, bl a WHO och Läkare utan gränser. Mercks vaccin prekvalificerades hösten 2019 och gavs handelsnamnet Ervebo. För detta vaccin fanns fler RCT-studier tillgängliga, vilka uteslutande valdes m h a sökmotorn PubMed som är länkad till olika databaser. Resultat: Både singeldosvaccinet Ervebo och dubbeldosvaccinen Ad26.ZEBOV/MVA-BN-Filo konstaterades ge ett nästan 100%-igt immunsvar mot ebolavirus. I den mån biverkningar förekom var de milda och övergående. Den mest rapporterade lokala biverkningen var ömhet vid injektionsstället och de vanligaste systemiska biverkningarna var huvudvärk, myalgi och feber. Diskussion: Den största utmaningen i framtiden blir att inleda vaccinationsprogram i länder med bristande finansiella resurser och infrastruktur samt pågående väpnade konflikter (främst DRK). Slutsatsen blev att de nya vaccinen mot ebolafeber är effektiva och säkra vid behandling mot ebolavirusinfektion.

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The outbreak of the Ebola virus was declared a Public Health Emergency of International Concern by the World Health Organisation (WHO). Due to the complex nature of the outbreak, the Centers for Disease Control and Prevention (CDC) had created interim guidance for monitoring people potentially exposed to Ebola and for evaluating their intended travel and restricting the movements of carriers when needed. Tools to evaluate the risk of individuals and groups of individuals contracting the disease could mitigate the growing anxiety and fear. The goal is to understand and analyze the nature of risk an individual would face when he/she comes in contact with a carrier. This thesis presents a tool that makes use of contextual data intelligence to predict the risk factor of individuals who come in contact with the carrier.

SOCIOECONOMIC FACTORS AND THE 2014-16 EBOLA VIRUS DISEASE OUTBREAK IN GUINEA, LIBERIA, AND SIERRA LEONE INTRODUCTION: Ebola virus disease (EVD) is an infectious disease transmitted by close contact with an estimated case fatality rate fluctuating around 50%. The most affected countries by the 2013-16 West African Ebola outbreak were Guinea, Liberia, and Sierra Leone. These countries reported a total of 28616 probable, suspected and confirmed cases. However, we are still learning about the sociodemographic factors that contributed to the outbreak characteristics at the subnational level. METHODS: Data were collected from the World Health Organization, Demographic Health Surveys, and Global Data Lab for 37 districts (8 for Guinea, 15 for Liberia, and 14 for Sierra Leone). The outcome of interest was epidemic size at the district level for Guinea, Liberia, and Sierra Leone (cumulative number of EVD patient confirmed and probable cases). Socio-demographic predictors included household density, sanitation level, mobility, and wealth status. We also controlled for the timing of the start of the outbreak across districts. Pearson’s correlation and multiple linear regression were employed in our analyses. Model building was informed by a review of the relevant literature. Sensitivity analyses were conducted to assess the impact of potential outliers. RESULTS: In the final multivariable regression model, wealth status and household density were positively associated with the epidemic size while sanitation level and the difference in the outbreak start dates were negatively associated with the outcome. These results did not change in the sensitivity analyses. The regression model explained 57% of the variance in epidemic size (Adj R-Sq=0.57), with the largest contribution from the international wealth index (semi-partial R-square=0.22). CONCLUSION: District sociodemographic characteristics such as household density, wealth and sanitation levels contributed to the EVD outbreak in Guinea, Liberia, and Sierra Leone, which is in agreement with recent studies. However, further research should consider other sociodemographic indicators as well as the role of migration and connectivity among regions.

Ebola viruses cause sever disease in humans and non-human primates. The resulting disease, Ebola virus disease (EVD), can have hemorrhagic manifestations and has mortality rates ranging from 20-90%. There is a strong need for better understanding of the disease as well as improved diagnostics and prognostics. One approach to improving diagnostic and prognostics for severe viral diseases such as EVD is to define how the host response to infection develops and produces indicators of disease and outcome. To create a better means to understand if a patient is likely to survive or succumb to Ebola (EBOV) infection, I have sought to develop an understanding of the host response to EBOV infection in humans from the recent outbreak. I analyzed RNA-Seq samples from the 2013-2016 West Africa outbreak. I identified that the innate immune pathways are in general over activated in EVD and is stronger in patients who succumbed to disease. Furthermore, I developed a set of 10 genes that can perform as a prognostic indicator of disease independent of the viral load. This is the first demonstration that the circulating transcriptional immune response to EBOV infection can be used to predict infection outcome. To work towards a diagnostic platform of disease, I analyzed multiple studies of time-resolved datasets in animal models of disease. I analyzed RNA-Seq and NanoString data coupled with telemetry data in EBOV-challenged macaques. The earliest and strongest changes seen in the pre-symptomatic stage of disease is the up-regulation of many innate immune genes. I used this information to develop a NanoString codeset that can act as a pre-symptomatic indicator of disease that was tested in further animal studies as a diagnostic in pre-symptomatic stages of disease. Together, this work has identified a sets of genes that can work as a diagnostic for pre-symptomatic patients of EBOV and act as a prognostic indicators of disease. In future outbreaks, this type of information will be important to help track primary contacts of infected individuals and first responders, as well as better inform clinical management of patients. This lays the groundwork for similar analysis to be performed on other severe viral diseases such as Lassa Fever and Marburg Fever. / 2019-11-27T00:00:00Z

Label-free imaging of viruses and nanoparticles directly in complex solutions is important for virology, vaccine research, and rapid diagnostics. These fields would all benefit from tools that allow for more rapid and sensitive characterization of viruses. Traditionally, light microscopy has been used in laboratories for detection of parasites, fungi, and bacteria for both research and clinical diagnosis because it is portable and simple to use. However, virus particles typically cannot be explored using light microscopy without the use of secondary labels due to their small size and low contrast. Characterization and detection of virus particles therefore rely on more complex approaches such as electron microscopy, ELISA, or plaque assay. These approaches require a significant level of expertise, purification of the virus from its natural environment, and often offer indirect verification of the virus presence. A successful virus visualization technique should be rapid, sensitive, and inexpensive, while needing minimal sample preparation or user expertise. We have developed a disposable cartridge based platform for real-time, sensitive, and label free visualization of viruses and nanoparticles directly in complex solutions such as serum. To create this platform we combined an interference reflectance imaging technique (SP-IRIS) with a sealable microfluidic cartridge. Through empirical testing and numeric modelling, the cartridge parameters were optimized and a flow rate of ~3 µL/min was established as optimal. A complex 2-dimensional paper based capillary pump was incorporated into the polymer cartridge to achieve a constant flow rate. Using this platform we were able to reliably show virus detection in a 20 minute experiment. We demonstrate sensitivity comparable to laboratory-based assays such as ELISA and plaque assay, and equal or better sensitivity compared to paper based rapid diagnostic tests. These results display a platform technology that is capable of rapid multiplexed detection and visualization of viruses and nanoparticles directly in solution. This disposable cartridge based platform represents a new approach for sample-to-answer label-free detection and visualization of viruses and nanoparticles. This technology has the potential to enable rapid and high-throughput investigation of virus particle morphology, as well as be used as a rapid point-of-care diagnostic tool where imaging viruses directly in biological samples would be valuable.

Gene delivery via lentiviruses can yield long term expression of transgenes. Specificity of host cell targeting by viral vectors occurs primarily through viral glycoprotein (GP)/cellular receptor interactions. Ebola virus (EBOV) GP has broad tropism for a variety of cell types making this viral GP a potentially useful reagent for delivery of gene therapy. However, titers of EBOV GP pseudotyped lentiviruses are insufficient for practical use in clinical applications. Enhancement of EBOV-GP pseudotyped titers by as little as half a log might yield clinically applicable titers. In an alanine scanning study, we identified 19 residues in EBOV-GP1 that increased transduction efficiency two to three fold. When mapped onto the crystal structure of EBOV GP, these residues were primarily located at the interface of GP1/GP2 suggesting these residue substitutions may confer conformational changes in the protein structure thereby enhancing transduction efficiency. To determine if combinations of these alanine substitutions might further enhance transduction, we have introduced the changes into EBOV GP in a stepwise manner. To date, introduction of some combinations of alanine substitutions resulted in as much as an eight-fold increase in transduction over WT GP, this being our super mutant combination, whereas other combinations eliminated transduction. Identification of 5 additional mutations via 3D modeling of the glycoprotein uncovered an additional mutation in GP2, located at the GP1/GP2 interface, which also enhances EBOV GP transduction. Transduction of cell lines important for gene therapy including hepatocytes and porcine airway cells confirmed an enhancement in transduction as well. Other cell populations, specifically fibroblasts and renal cells, were also transduced but enhanced transduction was not observed indicating this phenomenon may be cell type specific. The in vivo studies were inconclusive because no expression was detected from any of the EBOV GP pseudovirions. Even expression of the positive control, GP64 particles, waned after 3 weeks post inoculation indicating insufficient quantities or poor quality pseudovirions were used. These EBOV GPs should prove useful for future gene therapy studies by providing an alternate glycoprotein that is as effective as GP64 at producing high titer lentiviral vectors.

In early 2014, 3 West African states of Guinea, Liberia, and Sierra Leone made news headlines when Ebola virus disease (EVD) ravaged the sub-region. The Liberian government was ill-equipped to efficiently contain EVD outbreak due to inadequate training for hospitals and healthcare workers. The government's mandatory cremation policies and the banning of public gatherings significantly contributed to the spread of EVD. EVD infected 10,666 and 4,808 died from the disease in the first 6 months of the epidemic. Using Bandura's Social Cognitive Theory (SCT) as the theoretical framework, the purpose of this case study research was to examine the social, economic, and policy factors that contributed to the spread of EVD in the city of Monrovia, Liberia. Utilizing snowball sampling to identify participants, data were collected through in-depth interviews with 30 participants that included 10 EVD survivors, 10 family caregivers, 2 government officials, 4 nongovernmental organization staff, 2 academicians, and 2 members of the media. All data were inductively coded and analyzed using Braun and Clarke's thematic analysis procedure. Two key themes were identified through data analysis. First, participants noted that a better understanding of cultural traditions may have created opportunities for intervention that prevented unnecessary exposure to the virus. Second, survivors and caregivers experienced a 'hope for the best, but expect the worst' mentality throughout the experience that guided faith. The positive social change implications stemming include recommendations to the government of Monrovia to implement culturally sensitive policies related to pandemic containment, including training of healthcare workers and the public in the event of disease outbreak.

The Ebola viruses cause sporadic outbreaks of Ebola hemorrhagic fever (EHF) where origins have been traced to the continent of Africa and the Philippines. Since the initial discovery of Zaire and Sudan ebolavirus in 1976, the Ebola viruses have been responsible for severe hemorrhagic fever outbreaks in Africa with case fatality rates between 40-90%. The natural reservoir(s) of the Ebola viruses is currently unknown, but there is mounting evidence that fruit bats may play a key role. The goal of the current study is to screen a large variety of bat species from Africa and Asia where Ebola is known to be endemic for the presence of IgG specific antibody to Ebola virus in order to see which bat species may show evidence of past Ebola virus infection. Ebola virus would not be expected to cause lethal disease in its natural reservoir; therefore the presence of IgG antibody would be present. Identifying the species of bats that have been infected will allow researchers to hopefully isolate Ebola virus from bats adding to the evidence that bats are a reservoir species. The knowledge gained may also provide clues to new species of bats yet to be identified as possible natural reservoir(s) as well as expand the known geographical range of known Ebola virus outbreaks. Knowing which species of bats as well as their geographic range may help prevent future Ebola outbreaks by minimizing human-reservoir contact.