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Elbohy, Ola Assem Mohamed Saleh (2023) Plant-based vaccines for poultry viral diseases: Avian influenza. PhD thesis, University of Nottingham.

PDF (Thesis. Elbohy Ola. 28.07.2023) (Thesis - as examined) - Repository staff only until 28 July 2025. Subsequently available to Repository staff only - Requires a PDF viewer such as GSview, Xpdf or Adobe Acrobat Reader Available under Licence Creative Commons Attribution. Download (5MB)

Abstract

Avian influenza virus (AIV) is a significant reason for mortality in poultry,causing substantial economic loss. The importance of AIV is increased due to the ability of several subtypes to cause zoonotic infection, with, in the case of H5N1, mortality rates approaching 60%. Haemagglutinin (HA) is the main antigen of AIV, and protecting antibodies are elicited against it in the host after viral infection. Control of viral diseases in poultry depends mainly on vaccination and high biosecurity levels.

Commercial vaccines are available for chickens’ most common viral infections; however, viruses are still causing high economic losses. Vaccines that can be made at a cheap cost, are stable, and can be adapted to local virus strains are urgently needed. Such benefits are provided by plant-based vaccinations, which are appropriate for use in developing countries. The most predominant AIV strains in developing countries, especially Egypt, are H9N2 and H5N8. Virus-like particles (VLPs) have the same structure as the virus without inner nucleic acid. VLPs vaccines are safe and can stimulate both cellular and humoral immune responses in the host.

This project used Agrobacterium tumefaciens to introduce a plasmid encoding the codon optimised (HA) of H9N2 and H5N8 AIV into Nicotiana benthamiana leaves. Recombinant proteins and VLPs were transiently expressed and purified. The H2N3HA was transiently expressed in plants without codon optimisation of the gene but at lower levels, which suggested that codon optimisation is important for enhancing the gene expression levels. VLPs for both H5 and H9 were identified by western blot and transmission electron microscopy (TEM). Each VLP preparation was formulated with an adjuvant for immunisation trials in mice and chickens and HA-specific antibodies were detected by enzyme-linked immunosorbent assay (ELISA).

Furthermore, sera were tested for their capacity to effectively neutralise heterologous pseudotype viruses presenting HA from heterologous strains in the H5 and H9 subtypes. In addition, cellular immune responses in mice were estimated by enzyme-linked immunosorbent spot (ELISpot). Mice were immunised with three H9 and H5 VLPs doses and elicited excellent haemagglutinin-specific antibodies, completely neutralising the heterologous pseudotypes. Additionally, a high number of interferon-γ secreting splenocytes were detected on restimulation of cells from immunised mice with H9 and H5 VLPs through the ELISpot technique. Chickens were immunised with one dose of H5 VLPs eliciting haemagglutinin-specific antibodies, partially neutralizing the heterologous H5 pseudotypes. Plant-based VLP vaccines are an excellent alternative to the traditional vaccine with significant advantages for producing a vaccine candidate quickly with high potential for the poultry industry.

Item Type:	Thesis (University of Nottingham only) (PhD)
Supervisors:	Dunham, Stephen Daly, Janet
Keywords:	Avian influenza virus, AIV, poultry diseases, VLPs, vaccines, plasmids, Agrobacterium tumefaciens
Subjects:	Q Science > QR Microbiology > QR180 Immunology S Agriculture > SF Animal culture
Faculties/Schools:	UK Campuses > Faculty of Medicine and Health Sciences > School of Veterinary Medicine and Science
Item ID:	74282
Depositing User:	ElBohy, Ola
Date Deposited:	09 Aug 2023 13:33
Last Modified:	09 Aug 2023 13:33
URI:	https://eprints.nottingham.ac.uk/id/eprint/74282

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