Develop new prevention and control strategies for poultry respiratory diseases.<OL> <LI> Establishment of continuously growing avian cell lines<LI>Development of genetically engineered vaccines<BR>A. Reverse genetic systems for RNA virus <BR>B. Genetically attenuated vaccines for DNA viruses
NON-TECHNICAL SUMMARY: Avian viral disease, such as avian influenza, Newcastle disease, infectious bronchitis, pneumovirus, Marek?s disease, or laryngotracheitis cause economic losses in poultry industries by decrease productivities. Effective detection and prevention of avian viruses are of the utmost importance to maintain the US leadership in world poultry product markets. Development of more rapidly produced and more effective vaccine is required to minimize the loss in poultry production. This project for optimal vaccine production will address a stable and safer baseline for protection of avian infectious viral pathogens. By doing so, the improvement in sustainability of US poultry may be realized by the much safer control of long-term threat of virus spread (as either an epidemic or even pandemic).
APPROACH: Various regulatory factors will be modulated by genetic engineering technologies in primary chicken cells to establish continuously growing cell lines. Negatively regulating factors in both cell cycle and cell lifespan will be down-regulated using gene knock-down technology, while positively regulating factors will be overexpressed using various eukaryotic expression vectors containing strong promoter. For reverse genetic system, RNA genome of specific avian virus will be converted to cDNA by reverse transcriptase, and double stranded genome will be assembled into plasmid vector containing specialized promoter. Further recombinant virus propagation will be perform in continuously growing avian cell substrates. Mutagenesis will be performed on specific regions of the viral genes to be able to determine if these changes are detrimental to the infectivity/virulency of RNA virus. To construct the virulency defective-, but infectious DNA viruses, various viral genes including surface glycoproteins will be removed using homologous recombination between purified viral DNA genome and various reporter plasmids, that contain selectable markers. In the entire processes, the immortal chicken cell lines will be used as cellular substrates for virus propagation and diagnostic uses to detect viruses.