The overall goal of this project is to further understand the genetic basis of how S. typhimurium is able to cause persistent asymptomatic infections in swine. In this project, the repertoire of genes expressed by an S. typhimurium strain that causes persistent infections will be compared to highly virulent strains (that cause disease) and to an attenuated mutant. In the second specific aim, it will be determined if the same sets of genes are expressed upon exposure of S. typhimurium cells to relevant host cells. Finally, it is believed that the host plays an important role in creating suitable conditions for the development of persistent infections. Consequently, the gene repertoire of the host genes expressed in response to S. typhimurium also will be determined.
A comparative genomics approach will be used to evaluate gene expression. DNA microarrays containing the entire set of S. typhimurium genes have been developed and will be used to analyze S. typhimurium gene expression. The S. typhimurium strains that will be used for this study include two phenotypes of S. typhimurium strain 798 (a strain known to cause persistent infections), 3 virulent strains, and an attenuated, phase locked mutant of strain 798. Four cell types have been selected to measure S. typhimurium genes expressed in response to host target cell. The cells include freshly isolated porcine neutrophils and macrophages, an enterocytes cell line (CaCa-2), and differentiated M-cells resulting from co-cultivation of CaCo-2 cells with mouse lymphocytes isolated from Peyer's patches. Since there isn't a pig microarray available, and since the CaCo-2 cells are of human origin, the measurement of host target cell gene responses will employ DNA microarray chips specific for the mouse and human genomes.
Salmonella enterica serotype Typhimurium is one of the major causes of salmonellosis in humans. Pigs persistently infected with S. typhimurium are one of the major reservoirs of this pathogen. Generally, pigs persistently infected with S. typhimurium are asymptomatic. One means to reduce the risk of food borne infections caused by S. typhimurium is to prevent pigs from becoming persistently infected. This project is designed to understand the mechanisms promoting persistent infections using comparative genomics. We previously found a novel mechanism that allows S. typhimurium to sense its location in the intestines and turn on a unique set of genes that promote its ability to colonize enterocytes. It is believed that this results in persistent infections. In the current proposal, the repertoire of genes that are expressed by S. typhimurium cells that are adapted to grow in intestines will be compared to cells that grow outside of the animal. These results also will be compared to those obtained using highly virulent strains of S. typhimurium. In the second objective, we will determine if the same sets of genes are expressed when the S. typhimurium cells are exposed to target cells: epithelial cells from intestines or white blood cells. In the final objective it will be determined if gene expression of the target (host) cells respond differently to the various S. typhimurium strains being used and thus contribute to the development of persistent infections.