Unknown,Transcriptomics,Genomics,Proteomics

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Peripheral blood transcriptome responses to Salmonella Typhimurium in pigs differing in Salmonella shedding phenotypes

ABSTRACT: Salmonella species infect many vertebrate species, and pigs colonized with Salmonella enterica serovar Typhimurium (ST) are usually asymptomatic, making detection of these Salmonella-carrier pigs difficult. The variable fecal shedding of this gram-negative bacteria in such pigs is an important cause of foodborne illness and zoonotic disease. To investigate gene pathways and biomarkers associated with the variance in Salmonella shedding following experimental inoculation, we have initiated the first analysis of the whole blood transcriptional response induced by Salmonella. A population of pigs (n=40) was inoculated with ST and the peripheral blood and feces were collected between 2 and 20 days post-inoculation. Two groups of pigs with either low shedding (LS) or persistent shedding (PS) phenotypes were identified. The global transcriptional changes in response to ST inoculation were identified by Affymetrix Genechip?analysis of peripheral blood RNA at day 0 and day 2 post-inoculation. Forty pigs (n=40) was inoculated with ST. Four low shedding (LS) pigs and six persistent shedding (PS) pigs were identified. Transcriptom of peripheral blood collected at 0 and 2 dpi were identified by Affymetrix Genechip analysis.

ORGANISM(S): Sus scrofa

SUBMITTER: Chris Tuggle

PROVIDER: E-GEOD-27000 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:A first generation Affymetrix GeneChipÂ® Porcine genome array was used to profile the gene expression in porcine mesenteric lymph nodes over a time course of infection with S. Typhimurium, including the acute (8 hours post inoculation (hpi), 24 hpi, 48 hpi) and chronic (21 days post-inoculation (dpi)) stages of infection. Our objectives were to 1) identify and examine the stereotypical gene expression response within host MLN to S. Typhimurium infection, 2) characterize global host responses by revealing the specific features of the hostâs innate immunity pathways, and 3) explore if and how S. Typhimurium may escape the host immune response and develop into a carrier state. Our study has attempted to investigate the features of host gene expression profiling during S. Typhimurium infection at the acute and chronic infection stages and to explore the mechanism by which S. Typhimurium can escape from the host immune response and develop a carrier state in the host. In conclusion, by using the Affymetrix porcine GeneChip, 848 differentially expressed genes were identified in porcine MLN during infection and several specific features of host response were revealed by gene cluster and pathway analysis. Our data are the first report to investigate global host responses to S. Typhimurium in porcine MLN, and this new study provides data applicable for studying enteric salmonellosis of pigs, as well as humans. Experiment Overall Design: Fifteen piglets from Salmonella spp.-free sows were weaned at 10 days (d) of age, shipped to the National Animal Disease Center, Ames, IA and raised in isolation facilities. To confirm that all piglets were free of Salmonella spp. prior to challenge, bacteriological cultures were performed on rectal swabs twice. Seven week old pigs were randomly divided into 2 groups, 3 non-infected pigs and 12 infected pigs. Three non-infected control pigs were necropsied 2 days prior to experimental infection. On day 0, pigs in the infected groups were intranasally challenged with 1 billion CFU of Salmonella enterica serovar Typhimurium. Three infected pigs were necropsied at 8 hours post-inoculation (hpi), 24 hpi, 48 hpi and 21 days post-inoculation (dpi). Tissue samples from the mesenteric lymph nodes (MLN) were collected and immediately frozen in liquid nitrogen for RNA isolation.

Project description:The early interaction of Salmonella enterica serovar typhimurium DT104 with intact small intestinal mucosa was studied in a Small Intestinal Segment Perfusion (SISP) model. Intestinal segments were infected with or without Salmonella. Scrapings from jejunal segments were collected after perfusion for 0, 2, 4, or 8 hours. Details of the SISP experiment are described in: Niewold TA, Veldhuizen EJ, van der Meulen J, Haagsman HP, de Wit AA, Smits MA, Tersteeg MH, Hulst MM. Using the Operon 13K pig oligonucleotide array differences in host gene expression were recorded between infected and uninfected segments within a single pig (isogenic comparisons), and between identical treated segments collected from 3 individual SISP pigs, all responding markedly different to infection with Salmonella (inter-animal comparisons). A Small Intestinal Segment Perfusion (SISP) test was performed with 3 pigs (pig no. 2, 3, and 4) (cross-bred YorkshireM-CM-^W(Large WhiteM-CM-^WLandrace)). Two adjacent segments prepared in the mid-jejunum of each pig were perfused for 1 hour with Salmonella enterica serovar typhimurium DT104 suspended in peptone solution (10E-09 CFU/ml) or with peptone solution alone (mock infected segment) respectively. Subsequently, segments were perfused with peptone solution alone for a maximum period of 8 hours. At 2, 4, and 8 hours a part of the infected segment was dissected to obtain mucosal scrappings. The same was done at 0 and 8 hours for the uninfected (mock) control segment. RNA isolated from scrappings was used for microarray comparisons using the Operon 13K pig oligonucleotide array. 9 comparisons were done. For each of the 3 SISP pigs, expression in the 8 hours perfused infected segment, perfused for 8 hours, was compared to expression in its adjacent mock infected segment (3 isogenic comparisons, 8 hpi.). Expression in the infected segment of each SISP pig, dissected after 2 or 4 hours of perfusion, was compared to expression in an infected segment dissected from another SISP pig (2 versus 3, 2 versus 4, and 3 versus 4 / 3 comparisons at 2 hpi., and 3 comparisons at 4 hpi.). Dye-swaps were performed for each comparison. jejunum pig, host-microbe interaction, Salmonella enterica serovar typhimurium DT104.

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Peripheral blood transcriptome responses to Salmonella Typhimurium in pigs differing in Salmonella shedding phenotypes

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