Location: Endemic Poultry Viral Diseases Research
Project Number: 6040-32000-083-017-I
Project Type: Interagency Reimbursable Agreement
Start Date: May 1, 2023
End Date: Apr 30, 2026
Objective:
We propose to test an in ovo vaccination strategy to protect against both infectious laryngotracheitis virus (ILTV), Marek's disease virus serotype 1 and Newcastle disease virus (NDV) as a trivalent vaccine. This is based on the expression of glycoprotein D(gD) of ILTV infectious and glycoprotein B (gB) individually and, if possible, together delivered by rNDV. Our goal is to insert gD and gB in different intergenic regions of the NDV genomes and test for maximal expression using western blot analysis and immunofluorescence assays. The hypothesis of this proposal is that insertion of the genes encoding the major antigens of ILTV and MDV1within an LaSota-IL-4R genome will generate recombinants that are protective and safe after in ovo administration. Avian immune responses and protective outcomes induced by the delivery of these antigens following vaccination will be evaluated before and during challenges with virulent NDV and ILT viruses. The specific objectives are (1) create new ILTV/NDV vaccines for in ovo administration (2) evaluate the safety in embryonated eggs 3) evaluate the safety and immunogenicity of the vaccines in specific pathogen-free (SPF) chickens; and (3) evaluate the efficacy of the vaccines in chicken in vaccination-challenge models for ND, MD, and ILT. The recombinants will be generated using assembly strategies involving a series of synthetic DNA fragments and the recombination machinery of yeast.
Approach:
Our approach will be to utilize the ability of Newcastle disease virus (NDV) to replicate in then embryonated eggs of chicks to deliver avian alphaherpesvirus antigens. The gene encoding glycoprotein D of infectious laryngotracheitis virus (ILTV) and glycoprotein B of Marek's disease virus-1 (MDV1), as known as serotype 1 will be inserted into different intragenic regions within the genome of the LaSota strain of Newcastle disease virus NDV. To accomplish this an independent transcription unit containing the glycoprotein D gene of ILTV and glycoprotein B of MDV1 will be chemically synthesized and containing flanking sequences necessary for homologous recombination. Similarly, overlapping DNA fragments presenting the complete genome of NDV will be synthesized. Both ILTV, MDV1 and NDV fragments will be assembly in yeast using homologous recombination. NDV recombinants will individually contain an alphaherpesvirus gene (either gB of MDV1) or gD of ILTV. A recombinant containing both will also be attempted. Recombinants will be tested for genetic stability, safety in eggs and their ability to protect against matched challenges following in ovo vaccination.
