Investigation and characterisation of viral and host factors that impact intraspecies EBOV infectivity using a viral pseudotyping assay

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.

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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.

Item Type: Thesis (University of Nottingham only) (PhD)
Supervisors: Ball, Jonathan
McClure, Patrick
Urbanowicz, Richard
Keywords: Zaire Ebola Virus; Intraspecies variation; Viral fitness; Virus spillover; Pathogenesis
Subjects: Q Science > QR Microbiology
Faculties/Schools: UK Campuses > Faculty of Medicine and Health Sciences > School of Life Sciences
Item ID: 76475
Depositing User: Dey, Jayasree
Date Deposited: 13 Dec 2023 04:40
Last Modified: 13 Dec 2023 04:40

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