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Poultry producers suffer major economic losses both from mortality due to infectious
diseases, and from contamination of food products due to transfer of microorganisms.
As a result, they have a keen interest in breeding birds capable of resisting
infection by enteric food-borne pathogens such as Salmonella and Campylobacter, and
the development of novel immunologically based strategies to control pathogenic
microorganisms in the gastrointestinal tract of poultry. Understanding how the
immune system is regulated and responds to infectious agents requires whole system
approaches given that single immunological parameters have been unable to unlock
immune system complexity. It is increasingly important to be able to measure changes
in the expression of multiple genes in a tissue or animal in response to a single
physiological change. The availability of the chicken genome sequence provides the
opportunity to resolve questions concerning the molecular components of the innate
immune system. Key developments in molecular, genetic, and cellular biological
techniques provide us with new approaches to use the genome to investigate the
functional genomics (the study of the how and why a given gene behaves in a certain
way under specific conditions) and pathogenomics (the study of how genes behave when
pathogens interact with their host) of the avian innate response to Salmonella and
Campylobacter. Our primary goal is the use of genomic technologies to understand
prospective control points for modulating innate immunity, thus providing the poultry
industry with novel, pre-harvest intervention tools to control food-borne pathogens
and provide safe food products to the consumer.
<ol> <li> Assess innate immune variability in chickens and turkeys by examining the
frequency of genetic polymorphism in genes related to innate immunity
(cytokines/chemokines, toll-like receptors [TLR]). Initial analysis will be
undertaken in 2 pedigree lines of chickens that we have characterized in terms of
innate immune function and disease susceptibility and commercial lines of turkeys and
their wild turkey counterparts;
<li> Identify differentially expressed or genetically
altered genes in heterophils from the innate immune functionally divergent lines of
chickens and turkeys by use of suppressive subtractive hybridization (SSH).
Normalized, directionally cloned avian heterophil cDNA libraries will be constructed
from pools of mRNA purified from resting, inflammatory, and activated heterophils
following either the in vitro stimulation with inflammatory agonists or in vivo
following infection with Salmonella or Campylobacter; <li> Develop new modulators of
innate immunity such as pathogen associated molecular patterns (PAMPs) as immune
modulators of early colonization, PAMPs as adjuvants for commercial live vaccines,
and the development of new live vaccines that specifically induce a heterophil-
mediated innate immune response; <li> Apply anti-sense and RNAi technologies in poultry
by introducing anti-sense oligonucleotides and RNAi to available cell lines (the
macrophage HD-11 cell line) by transfection, using western-blot and Real Time-PCR to
assess the effectiveness of gene silencing, and characterizing the function of the
target gene by stimulating the cells with PAMPs and measuring immune responses. The
in vivo experimental approach is similar to that stated above, except that in ovo
route will be used to introduce anti-sense oligonucleotides and RNAi; and <li> Evaluate
anti-microbial peptides from chicken and turkey heterophils that have potential as new biotherapeutics for food-borne bacteria.</ol>