Kent Academic Repository Full text document (pdf) Citation for published version BORE, Joseph Akoi (2020) Sero-prevalence of Ebola and Lassa virus in the Republic of Guinea, Macenta prefecture. Doctor of Philosophy (PhD) thesis, University of Kent, Public Health England (PHE), Porton. DOI Link to record in KAR https://kar.kent.ac.uk/82364/ Document Version UNSPECIFIED Copyright & reuse Content in the Kent Academic Repository is made available for research purposes. Unless otherwise stated all content is protected by copyright and in the absence of an open licence (eg Creative Commons), permissions for further reuse of content should be sought from the publisher, author or other copyright holder. Versions of research The version in the Kent Academic Repository may differ from the final published version. Users are advised to check http://kar.kent.ac.uk for the status of the paper. Users should always cite the published version of record. Enquiries For any further enquiries regarding the licence status of this document, please contact: [email protected] If you believe this document infringes copyright then please contact the KAR admin team with the take-down information provided at http://kar.kent.ac.uk/contact.html University of Kent School of Biosciences Sero-prevalence of Ebola and Lassa virus in the Republic of Guinea, Macenta Prefecture. by Joseph Akoi BORE Thesis for the degree of Doctor of Philosophy June 2020 1 Abstract Following the 2013–2016 West Africa Ebolavirus (EBOV) outbreak, a broad range of scientific research has been conducted to understand and prevent future epidemics. Our current study, supported by the United States Food and Drug Administration (FDA), uses sero-epidemiology to look at the footprint in- cidence of EBOV and Lassa virus (LASV) in the prefecture of Macenta located in forested Guinea. Our study assesses seroprevalence of these zoonotic vi- ruses in bushmeat hunters and their household members. These groups of people are at risk of exposure to multiple viral zoonotic infections via constant contact with wildlife, either by hunting, trapping or butchering. In order to in- vestigate potential historical EBOV and/or Lassa virus LASV infections, serum samples were collected from villages that were affected or unaffected by the 2013-2016 EBOV outbreak in Guinea. We performed Enzyme-Linked Im- munosorbent Assay (ELISA), western blot analysis and flow cytometry for the detection of EBOV glycoprotein (GP) and LASV nucleoprotein (NP) antigens specific immunoglobulin G. A number of positive samples were detected for both pathogens, suggesting that these two pathogens are circulating in Guinea and may cause mild or asymptomatic infection in a proportion of cases. Im- portantly, this study suggests EBOV may have been circulating in Guinea be- fore the 2013-2016 outbreak. 2 Hypothesis of the Thesis The primary purpose of this study is to assess to what extent Ebolavirus has been circulating within the forested region of Guinea prior to the 2013-2016 Ebola virus disease (EVD) outbreak in West Africa. Due to the nature of their work, bushmeat hunters and their household family members are deemed to be at high risk of Ebo- lavirus contamination and spill over. We then established a cohort of bushmeat hunt- ers and their immediate families and collected serum samples from volunteers. We looked for the presence of anti-Ebolavirus antibodies to determine previous exposure to the virus. Our cohort includes bushmeat hunters from villages affected by the 2013- 16 Ebolavirus outbreak and villages that were not affected. Therefore, we hypothe- sized that a number of samples from previously non exposed villages to EVD will contain anti-Ebolavirus antibodies, suggesting that the virus has been circulating or still circulate in the zone prior to the past Ebola virus outbreak. 3 Table of Contents Abbreviation list .................................................................................................................. 7 List of Tables ........................................................................................................................ 9 List of Figures .................................................................................................................... 10 Acknowledgements .......................................................................................................... 11 Publication .......................................................................................................................... 12 DECLARATION OF AUTHORSHIP ................................................................................ 13 GENERAL INTRODUCTION .............................................................................. 14 1.1 Ebola virus .................................................................................................................. 15 1.1.1 Discovery of Ebola virus ..................................................................................... 15 1.1.2 Notable historical outbreaks .............................................................................. 16 1.1.3 Ebola; Species and phylogenetics ...................................................................... 20 1.1.4 Ebola; Genome structure and organization ....................................................... 21 1.1.5 Ebola; Replication .............................................................................................. 22 1.1.6 Ebola; Disease and immune response ............................................................... 23 1.1.6.1 Innate immune response to Ebola virus ........................................................ 24 1.1.6.2 Adaptive response to Ebola virus ................................................................... 25 1.1.6.3 B cells responses to Ebola virus ..................................................................... 25 1.1.6.4 T cells responses to Ebola virus...................................................................... 26 1.1.6.5 B and T cells dynamics ................................................................................... 27 1.1.7 Host immune response and outcome to survival .............................................. 28 1.1.8 Ebola; Reservoir and Transmission .................................................................... 31 1.1.8.1 Transmission .................................................................................................. 34 1.1.9 Age factor in Ebola virus infection outcome ...................................................... 35 1.1.10 Co-infection and clinical outcome of EVD .......................................................... 35 1.1.11 Ebola; Diagnosis, prophylaxis and vaccines ....................................................... 36 1.2 The West African EVD Outbreak of 2013-2016 ......................................................... 37 1.2.1 West African Outbreak management ................................................................ 40 1.2.2 Social impact of the West Africa EVD ................................................................ 41 1.3 Sub symptomatic infections ....................................................................................... 42 1.3.1 Potential reason for sub -symptomatic infection .............................................. 43 1.3.2 Historical Filovirus sub symptomatic infection .................................................. 43 1.3.3 Case studies on Ebola virus asymptomatic transmission .................................. 45 1.3.4 Pros and cons of seroprevalence studies ........................................................... 48 1.4 Lassa virus; Introduction ............................................................................................ 49 1.4.1 Discovery of Lassa virus ..................................................................................... 50 1.4.2 Lassa; Species and Phylogeny ............................................................................ 51 1.4.3 Lassa; Genome structure and organization ....................................................... 51 4 1.4.4 Lassa; Replication ............................................................................................... 53 1.4.5 Lassa; Transmission ............................................................................................ 55 1.4.6 Lassa; Immune response .................................................................................... 57 1.4.7 Lassa; Clinical symptoms and treatment ........................................................... 58 1.5 Study aim and objectives ........................................................................................... 60 MATERIALS AND METHODS ........................................................................... 61 2.1 Sample collection ....................................................................................................... 62 2.2 Enzyme-Linked Immunosorbent Assays (ELISA) ........................................................ 62 2.2.1 Indirect ELISA assay procedure .......................................................................... 62 2.2.2 Ebola virus Glycoprotein detection .................................................................... 64 2.2.3 Lassa virus IgG detection.................................................................................... 66 2.2.4 Lassa virus IgG detection ELISA assay procedure............................................... 67 2.3 Ebola virus protein detection by western blot .........................................................