Submitted to: Veterinary Microbiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 10/28/2005
Publication Date: 4/16/2006
Citation: Uthe, J.J., Stabel, T.J., Zhao, S., Tuggle, C.K., Bearson, S.M. 2006. Analysis of porcine differential gene expression following challenge with Salmonella enterica serotype Choleraesuis using suppression subtractive hybridization. Veterinary Microbiology. 114(1-2):60-71. Interpretive Summary: Swine infected with Salmonella enterica serovar Choleraesuis (S. Choleraesuis) are a concern for two major reasons: first, Salmonella-infected swine are a food safety concern, and second, swine salmonellosis is an important animal health issue resulting in millions of dollars of lost income to the pork industry annually. Thus, characterizing the porcine response to infection with Salmonella may identify targets for diagnostic and therapeutic development. The aim of this study was to analyze swine genes with differential gene expression during infection with S. Choleraesuis. Among the genes with increased expression, several genes involved in host immunity and survival during infectious stress were identified. An increase in expression was also observed for a gene that regulates rearrangements in the host cytoskeleton, a system that Salmonella manipulates to gain entry into host cells. A decrease in expression was observed for genes playing a role in cytoskeleton sequestering and the anti-inflammatory response. This research contributes to understanding the reponse in pigs to infection with S. Choleraesuis that may assist in developing more effective methods in disease diagnosis and prophylaxis. Improving swine health will benefit both swine producers and swine consumers.
Technical Abstract: An important aspect of salmonellosis that continues to elude our understanding is the mechanism(s) of Salmonella enterica serotype-host specific response. Salmonella enterica subsp. enterica serotype Choleraesuis (S. Choleraesuis) is a swine-adapted strain that may affect host gene expression in a species-specific manner. To characterize the porcine transcriptional response to S. Choleraesuis infection, the RNA profiles from mesenteric lymph nodes of swine experimentally infected with S. Choleraesuis for 24 hours were analyzed by Suppression Subtractive Hybridization (SSH). Forward and reverse subtractions of pooled mRNA samples from 3 infected and 3 non-infected pigs were performed to identify differentially expressed genes. Differential cDNA screening by Southern hybridization was performed on 384 forward and 288 reverse subtracted cDNA clones, revealing 44 up-regulated and 44 down-regulated genes. The DNA sequence of the cDNA clones identified genes with a role in a variety of cellular functions as well as gene products of unknown function. Seven up-regulated genes (CXCL10, CXCR4, Syntenin, DNAJA1, HSP105, HSP90 and Annexin A5) and two down-regulated genes (Thymosin-beta-4 and GNB2L1) as well as two functionally related genes (HSP70 and DNAJA4:pDJA1) were selected for further analysis based on their predicted roles in infection and immunity. Real-time RT-PCR was performed using RNA collected from a time course of infection spanning from the acute phase (8 h) to the chronic phase (21 d). Correlating with the clinical signs of infection (fever, diarrhea and lethargy), the most dramatic induction of gene expression occurred at 48 h p.i. This was preceded by the down-regulation of the inflammatory suppression protein, Thymosin-beta-4 at 24 h p.i. Thus, the investigation revealed the differential expression of genes with a role in a variety of host cellular functions in response to S. Choleraesuis infection including genes involved in innate immunity and cytoskeleton regulation, thereby further defining the porcine response to a host-adapted strain of Salmonella.