1a.Objectives (from AD-416)
Develop chicken intestinal microarray chips which contains 10,000 non-redundant EST sequences and investigate the molecular basis for the macrophage-mediated innate immune responses in the gut to Salmonella and Eimeria. Identify the nature of host genes which control the protective immunty against Salmonella and Eimeria by conducting high throughput gene expression assays using chicken intestinal and macrophage chips.
1b.Approach (from AD-416)
Chicken intestinal microarray chips will be first developed by PCR reaction of 10,000 non-redundant intestinal genes selected from chicken intestinal cDNA library. Chickens will be infected with coccidia or salmonella, and RNA samples will be prepared from infected intestinal lymphocytes. Primary macrophages will be prepared from broiler chickens and infected with Eimeria or Salmonella for probe RNA preparation. Molecular basis for the protective innate immune responses in the gut to Salmonella and Eimeria infections will be investigated by carrying out high-throughput gene expression analysis using intestinal and macrophage cDNA microarrays. The role of host immune factors which were identified in the Objective 1 will be validated by real time PCR and northern blot analysis and host factors which are responsible for the intracellular killing of Salmonella and Eimeria will be identified.
This research investigates cell-mediated immunity against coccidia and Salmonella using chicken cDNA microarray technology and investigates host-pathogen interactions using functional genomics technologies. Using chicken macrophage and intestinal cDNA microarrays, collaborations with University of Delaware scientists identified many novel genes which are significantly expressed during coccidiosis and salmonellosis. Many host genes which were modulated by Eimeria and Salmonella included defensins and cytokines and controlled local innate immune responses. Real-time RT-PCR was used to validate the expression of many innate immune response genes.
The progress of this grant is monitored by monthly teleconferences, annual progress reports to the CSREES NRI program manager and presentation of new findings at national and international meetings.