Multivalent rHVT Vaccine Genome Insertion for Genetic Stability
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Solution Overview
Problem
Existing multivalent recombinant herpesvirus of turkeys (rHVT) vector vaccines face challenges with genetic stability and effective expression of multiple heterologous genes, leading to instability and reduced efficacy in protecting against multiple avian diseases.
Innovation Solution
The rHVT vector vaccine expresses IBDV VP2 and NDV F genes from the Us genome region and ILTV gD and gI genes from the UL genome region, specifically inserting the ILTV gD and gI genes between the UL54 and LORF3 genes, achieving stable replication and expression.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Adaptability or versatility
If multiple heterologous genes are inserted into the HVT vector genome, then the vaccine can protect against multiple diseases (multivalence), but the genetic stability and replication capacity of the vector is compromised
Solution Approach 1:
The patent divides the multiple heterologous genes into separate expression cassettes, each with its own promoter and regulatory elements. These cassettes are inserted at different locations in the HVT genome (Us region for NDV and IBDV genes, UL region for ILTV genes), preventing mutual interference and maintaining genetic stability while achieving multivalence
Solution Approach 2:
The patent applies different promoter types (immediate-early, early, late) from the HVT genome to different heterologous genes based on their expression requirements. This localized optimization of promoter-gene pairing ensures appropriate expression levels for each antigen while maintaining overall vector stability
2Adaptability or versatility
If multiple heterologous genes are inserted into the HVT vector, then protection against multiple pathogens is achieved, but the expression level and duration of foreign genes is insufficient
Solution Approach 1:
The patent uses immediate-early promoters (such as IEprom or mCMV-IE1) that are activated very early in the viral replication cycle to drive expression of heterologous genes. This preliminary action ensures that antigen expression begins immediately upon infection, establishing high expression levels from the start and maintaining them throughout the infection period
Solution Approach 2:
The patent designs expression cassettes that remain active throughout different phases of viral replication by utilizing promoters that are constitutively active or can be activated at multiple stages. This ensures continuous expression of heterologous genes throughout the productive infection period, maintaining protective antigen levels
3Adaptability or versatility
If heterologous genes are inserted into the HVT genome, then the vaccine can express foreign antigens, but the replication capacity of the vector is affected
Solution Approach 1:
The patent extracts heterologous genes from their native viral genomes and places them into the HVT vector under the control of HVT promoters. This separation allows the heterologous genes to be expressed without carrying the regulatory burden of their original viral genomes, minimizing interference with HVT replication while maintaining antigen expression
Solution Approach 2:
The patent uses universal promoter elements from the HVT genome that can drive expression of any heterologous gene. These promoters (IEprom, Eprom, Lprom) serve multiple functions: they activate transcription of viral genes and heterologous genes, and their activity can be modulated by viral regulatory proteins, allowing coordinated expression without compromising replication
4Adaptability or versatility
If the ILTV gD and gI genes are inserted into the HVT vector, then protection against ILTV is achieved, but the genetic stability is reduced
Solution Approach 1:
The patent separates the ILTV gD and gI genes into distinct expression cassettes with individual promoters, rather than inserting them as a single unit. This segmentation allows independent optimization of each gene's expression and reduces the metabolic burden on the viral replication machinery, maintaining genetic stability while achieving ILTV protection
Solution Approach 2:
The patent optimizes the promoter-gene pairing parameters by selecting appropriate HVT promoters (such as Eprom or Lprom for envelope glycoproteins) that match the expression requirements of ILTV antigens. This parameter optimization ensures stable expression without overwhelming the viral replication system
Data Source
AI summary
The present invention regards a new recombinant HVT (rHVT) construct, useful as multivalent vector vaccine for poultry. The rHVT comprises 4 heterologous genes from poultry pathogens: the VP2 gene from IBDV, the F gene from NDV, and the gD and gI genes from ILTV. The VP2 and F genes are inserted in the Us genome region of the rHVT. The gD-gI genes are inserted in the UL genome region, between the UL54 and the LORF3 genes. The new rHVT-VP2-F-gD-gI proved to be genetically stable in vitro and in vivo, and expressed all inserted genes well. Also it was an effective vaccine against severe challenge infections with NDV, IBDV, and ILTV.