Investigation and characterisation of viral and host factors that impact intraspecies EBOV infectivity using a viral pseudotyping assay
Zaire Ebola Virus (EBOV) is the etiological agent of Ebola Virus Disease, which is characterised by inflammation and damage to endothelial cell linings that can rapidly advance to internal haemorrhaging and multi-organ failure, with mortality rates observed as high as 90%. As a zoonotic virus, curre...
| Main Author: | |
|---|---|
| Format: | Thesis (University of Nottingham only) |
| Language: | English |
| Published: |
2023
|
| Subjects: | |
| Online Access: | https://eprints.nottingham.ac.uk/76475/ |
| _version_ | 1848800906710089728 |
|---|---|
| author | Dey, Jayasree |
| author_facet | Dey, Jayasree |
| author_sort | Dey, Jayasree |
| building | Nottingham Research Data Repository |
| collection | Online Access |
| description | Zaire Ebola Virus (EBOV) is the etiological agent of Ebola Virus Disease, which is characterised by inflammation and damage to endothelial cell linings that can rapidly advance to internal haemorrhaging and multi-organ failure, with mortality rates observed as high as 90%. As a zoonotic virus, current evidence suggests that bats may be the natural reservoir for EBOV. With outbreaks becoming more frequent, EBOV remains an ongoing threat to public health. Further work investigating the intraspecies variation present within EBOV and the viral and host genetic factors that impact viral fitness, is needed to provide insight into potential genetic drivers of virus spillover and pathogenesis.
This project developed the first EBOV GP pseudotyping panel that is representative of the intraspecies variation the virus displayed in outbreaks between 1976 and 2016. The panel was then tested in HuH-7 and Hyp/Lu 45.1 cells to determine the differences in infectivity profiles between the EBOV variants. The project demonstrated that EBOV outbreak strains display differential infectivity in both cell lines, with the Mbandza 2003 strain exhibiting the lowest infectivity in multiple cell lines. This characteristic was deemed a product of the T527 residue that is present in the internal fusion loop of the Mbandza 2003 GP. The assay also confirmed that the tissue culture adaptation, T544I, significantly impacted the infectivity of the Mayinga 1976 strain, increasing it relative to wild-type strains in both cell lines. The panel was further tested alongside sequences representative of theoretical ancestral GPs to determine the impact of the genetic differences between these sequences on viral fitness and also provide more insight into the theory that bats are the reservoir for the virus. Investigations showed that ancestral sequences are more infectious than outbreak sequences in both HuH-7 and Hyp/Lu 45.1 cells, suggesting that the H. monstrosus bat species that the Hyp/Lu 45.1 cells were derived from, may not be a natural reservoir for EBOV and that other selective pressures may be driving evolution.
Finally, the project investigated the impact of single nucleotide polymorphisms (SNPs) that were present in the NPC1 receptor of African populations to determine their impact on the susceptibility to EBOV. It was determined that the NPC1 containing the P434S and M642I SNPs was more permissive to EBOV infection relative to its wild-type homologue. In conclusion, this project established that variation in both the viral GP and host receptor impacts the entry fitness of EBOV. |
| first_indexed | 2025-11-14T20:59:00Z |
| format | Thesis (University of Nottingham only) |
| id | nottingham-76475 |
| institution | University of Nottingham Malaysia Campus |
| institution_category | Local University |
| language | English |
| last_indexed | 2025-11-14T20:59:00Z |
| publishDate | 2023 |
| recordtype | eprints |
| repository_type | Digital Repository |
| spelling | nottingham-764752025-02-28T15:19:09Z https://eprints.nottingham.ac.uk/76475/ Investigation and characterisation of viral and host factors that impact intraspecies EBOV infectivity using a viral pseudotyping assay Dey, Jayasree Zaire Ebola Virus (EBOV) is the etiological agent of Ebola Virus Disease, which is characterised by inflammation and damage to endothelial cell linings that can rapidly advance to internal haemorrhaging and multi-organ failure, with mortality rates observed as high as 90%. As a zoonotic virus, current evidence suggests that bats may be the natural reservoir for EBOV. With outbreaks becoming more frequent, EBOV remains an ongoing threat to public health. Further work investigating the intraspecies variation present within EBOV and the viral and host genetic factors that impact viral fitness, is needed to provide insight into potential genetic drivers of virus spillover and pathogenesis. This project developed the first EBOV GP pseudotyping panel that is representative of the intraspecies variation the virus displayed in outbreaks between 1976 and 2016. The panel was then tested in HuH-7 and Hyp/Lu 45.1 cells to determine the differences in infectivity profiles between the EBOV variants. The project demonstrated that EBOV outbreak strains display differential infectivity in both cell lines, with the Mbandza 2003 strain exhibiting the lowest infectivity in multiple cell lines. This characteristic was deemed a product of the T527 residue that is present in the internal fusion loop of the Mbandza 2003 GP. The assay also confirmed that the tissue culture adaptation, T544I, significantly impacted the infectivity of the Mayinga 1976 strain, increasing it relative to wild-type strains in both cell lines. The panel was further tested alongside sequences representative of theoretical ancestral GPs to determine the impact of the genetic differences between these sequences on viral fitness and also provide more insight into the theory that bats are the reservoir for the virus. Investigations showed that ancestral sequences are more infectious than outbreak sequences in both HuH-7 and Hyp/Lu 45.1 cells, suggesting that the H. monstrosus bat species that the Hyp/Lu 45.1 cells were derived from, may not be a natural reservoir for EBOV and that other selective pressures may be driving evolution. Finally, the project investigated the impact of single nucleotide polymorphisms (SNPs) that were present in the NPC1 receptor of African populations to determine their impact on the susceptibility to EBOV. It was determined that the NPC1 containing the P434S and M642I SNPs was more permissive to EBOV infection relative to its wild-type homologue. In conclusion, this project established that variation in both the viral GP and host receptor impacts the entry fitness of EBOV. 2023-12-13 Thesis (University of Nottingham only) NonPeerReviewed application/pdf en cc_by_nc https://eprints.nottingham.ac.uk/76475/1/DEY_JAYASREE_4303643_Final_Submission_Minor_Corrections_OCT_2023.pdf Dey, Jayasree (2023) Investigation and characterisation of viral and host factors that impact intraspecies EBOV infectivity using a viral pseudotyping assay. PhD thesis, University of Nottingham. Zaire Ebola Virus; Intraspecies variation; Viral fitness; Virus spillover; Pathogenesis |
| spellingShingle | Zaire Ebola Virus; Intraspecies variation; Viral fitness; Virus spillover; Pathogenesis Dey, Jayasree Investigation and characterisation of viral and host factors that impact intraspecies EBOV infectivity using a viral pseudotyping assay |
| title | Investigation and characterisation of viral and host factors that impact intraspecies EBOV infectivity using a viral pseudotyping assay |
| title_full | Investigation and characterisation of viral and host factors that impact intraspecies EBOV infectivity using a viral pseudotyping assay |
| title_fullStr | Investigation and characterisation of viral and host factors that impact intraspecies EBOV infectivity using a viral pseudotyping assay |
| title_full_unstemmed | Investigation and characterisation of viral and host factors that impact intraspecies EBOV infectivity using a viral pseudotyping assay |
| title_short | Investigation and characterisation of viral and host factors that impact intraspecies EBOV infectivity using a viral pseudotyping assay |
| title_sort | investigation and characterisation of viral and host factors that impact intraspecies ebov infectivity using a viral pseudotyping assay |
| topic | Zaire Ebola Virus; Intraspecies variation; Viral fitness; Virus spillover; Pathogenesis |
| url | https://eprints.nottingham.ac.uk/76475/ |