|Title:||A Systems Biology Approach to Understanding the Salmonella-Host Interactome in Poultry and Swine|
|Objective:||Objective 1: Conduct research on the differential host-pathogen interactions of Salmonella in human, chicken, and swine intestinal key host immune cells using emerging genomic technologies. |
Objective 2: Analyze and characterize both host and Salmonella proteins that are modulated in expression during infection using quantitative proteomics.
Objective 3: Research on the molecular and cellular details of the host-microbe interactions will be used to identify virulence-associated microbial genes and host defense strategies. Identify potential intervention targets (e.g., host kinases) for Salmonella infections in food animals.
Objective 4: Develop strategies for the reduction of foodborne pathogens by targeting the host innate immune system (by identifying the use of immunomodulatory antimicrobial or host-defense peptides) and targeting and identifying virulence factors. Sub-objective 4A: Molecular characterization of anti-infectives that target the host innate immune system to facilitate pathogen-specific immune responses. Sub-objective 4B: Develop a high-throughput assay to screen a series of commercial libraries of small molecules for their ability to inhibit virulence factors produced by S. typhimurium.
|More Info:||Approach: Objective 1: Utilize deep sequencing and mutagenesis technologies to dissect the differential host-pathogen interactions of Salmonella in human, chicken, and swine intestinal epithelial cells and macrophages. Specifically, we will determine the differential transcriptome of S. Typhimurium in mammalian versus chicken epithelial cells and develop gene-deletion mutants of S. Typhimurium to elucidate the differential mechanisms of intestinal pathogenicity of S. Typhimurium in humans compared to chickens and swine. Objective 2: A newly described technique of purifying live Salmonella expressing green fluorescent protein from either infected tissues or cell cultures using flow cytometry will be used to analyze and characterize both host and Salmonella proteins that are modulated in expression during infection. Quantitative proteomics including conventional two-dimensional electrophoresis, difference gel electrophoresis (DIGE), and mass spectrometry will be used to facilitate accomplishment of this objective. Objective 3: Using a matched comparative model (Salmonella characterized by contrasting degrees of pathogenicity and/or gene-deletion mutants of S. Typhimurium), newly developed peptide arrays will be used for studying the kinome of chicken and swine intestinal epithelial cells and macrophages. Cell lysates will be analyzed on a kinomics array containing 1,024 peptides derived from known phosphorylation sites annotated with reported upstream kinases. In addition, reverse chemical genetics will be used to identify host kinases that are essential in controlling intracellular Salmonella infections. This procedure will enable us to identify a class of kinases using selective chemical inhibitors of kinases with relevant biological activities to control in vitro and in vivo infections. Objective 4: We will develop a high-throughput assay to search for inhibitors of the Type 3 secretion systems. We will screen a series of commercial libraries of small molecules for their ability to inhibit type 3 secretion by S. Typhimurium. A systems approach will be employed to understand and characterize the host-pathogen interactions that are manipulated in food animals using novel therapeutic approaches with BT peptides and CpG oligonucleotides without engendering antimicrobial resistance. Microarray analyses of avian and porcine peripheral blood granulocytes and monocytes following treatment with BT peptides or CpG oligonucleotides will be performed. Using InnateDB, bioinformatic interrogation of gene ontology, signaling pathways and transcription factor binding sites will be undertaken; confirmation will be achieved experimentally by qRT-PCR and inhibitor studies of in vitro functional biological assays, and followed up by direct biochemical confirmation. Collectively, these will lead to substantial advances in understanding the complexity of signaling pathways and transcription factors involved in the responses to BT peptides and CpG modulation.|
|Funding Source:||United States Department of Agriculture (USDA), Agricultural Research Service (ARS)|
|Institutions:||USDA/ARS - Southern Plains Area|
|Project Reports:||2011 Annual Report|
|Published USDA ARS Articles|
Modulation of chicken intestinal gene expression by small cationic peptides as feed additives during the first week post-hatch
Kogut MH, Genovese KJ, He H, Swaggerty CL, Jiang Y.
Clin Vaccine Immunol. 2013 Sep;20(9):1440-8.
The gut microbiota and host innate immunity: Roles in host metabolism and metablic diseases
J Appl Poult Res. 2013 Sep;22(3):637-46.
Salmonella enterica Typhimurium infection causes metabolic changes in chicken muscle involving AMPK, fatty acid and insulin/mTOR signaling
Arsenault RJ, Napper S, Kogut MH .
Vet Res. 2013 May 17;44:35.
Obligate brood parasites show more functionally effective innate immune responses: An eco-immunological hypothesis
Hahn DC, Summers S, Genovese KJ, He LH, Kogut MH.
J Evol Biol. 2013 Apr. [Epub ahead of print]
|Food Safety Categories:||Methodology|
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