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United States Department of Agriculture

Agricultural Research Service

Research Project: Molecular Analysis of Salmonella Virulence, Antibiotic Resistance, and Host Response
2012 Annual Report


1a.Objectives (from AD-416):
1)Identify molecular mechanisms in Salmonella that impact virulence and antimicrobial resistance in the gastrointestinal environment of cattle and swine. 2)Characterize the genetic pathways in swine that respond to Salmonella infection, their relationship to carrier status, and the ability to predict and control Salmonella prevalence and ultimately food safety. 3)Investigate interactions between Salmonella and host microbiota that influence Salmonella colonization and persistence, alter the commensal composition in the host, and provide novel intervention strategies to decrease Salmonella in swine populations.


1b.Approach (from AD-416):
1) Use phenotypic assays, recombineering, and global transcriptional analysis to identify the genes and genetic pathways involved in regulating Salmonella DT104 hyper-invasion in response to a) rumen protozoa and b) sub-inhibitory concentrations of antibiotics..
2)Use an attenuated, LPS/flagella-deficient S. Typhimurium strain to re-focus the pig’s immune response from serovar-specific antigens to conserved antigens during vaccination against Salmonella, thereby providing cross-protection against multiple Salmonella serovars..
3)Classify the porcine gene expression differences between low and high shedders of Salmonella during colonization..
4)Compare the gastrointestinal microbial communities of pigs before, during and after Salmonella colonization to assess correlations with Salmonella shedding status in the pigs.


3.Progress Report:
The incidence of multidrug-resistant (MDR) Salmonella is an important health and food safety concern in humans as 33.1 percent of Salmonella Typhimurium isolates are resistant to at least one class of antibiotics, and 20.8 percent are resistant to five or more classes. Because the response of drug resistant bacteria exposed to antibiotics can affect virulence, we investigated the effect of tetracycline (a commonly used antibiotic in animal feed) on isolates of MDR Salmonella Typhimurium. We found that exposure to tetracycline induced invasion in several Salmonella isolates. These results indicate that the invasive capabilities of a subset MDR Salmonella Typhimurium are enhanced in the presence of tetracycline.

Salmonella-shedding pigs contaminate other animals, meat products during processing, and crops when Salmonella-containing manure is used as a soil fertilizer. To identify factors that may influence Salmonella shedding in pigs, we are examining the bacteria residing in the intestinal tract of pigs, referred to as commensal bacteria or the intestinal microbiota. We have sequenced a common gene within this “community” of bacteria in pigs to determine which bacteria are present before and after Salmonella challenge that might affect Salmonella shedding levels. Our goal is to identify Salmonella antagonists (or their bio-products) that reduce Salmonella shedding in pigs to decrease environmental contamination and increase food safety.


4.Accomplishments
1. Linking genetic variations in porcine genes to colonization and shedding of Salmonella in pigs. To address this strategy, ARS researchers at Ames, IA and collaborators at Iowa State University searched greater than 3,000 porcine genes to identify genetic variations in pig genes that respond to Salmonella. DNA segments know as single nucleotide polymorphisms (SNPs) were identified in 31 pig genes whose expression is altered in response to Salmonella. To determine if the identified porcine SNPs were associated with tissue colonization or fecal shedding of Salmonella, four independent pig populations were genotyped for the SNPs. Statistical analysis revealed 13 SNPs that associated with Salmonella fecal shedding or tissue colonization. These genetic differences in pigs associated with Salmonella shedding provide insight in identifying and selecting for pigs with increased resistance to Salmonella colonization to improve food safety (for example, development of diagnostic tests to screen pig lines for the desired trait).


Review Publications
Uthe, J.J., Bearson, S.M., Qu, L., Dekkers, J.C., Nettleton, D., Y Rodriguez, T., O'Connor, A., Mckean, J., Tuggle, C.K. 2011. Integrating comparative expression profiling data with association of SNPs to Salmonella shedding for improved food safety and porcine disease resistance. Animal Genetics. 42(5):521-534.

Uthe, J.J., Qu, L., Couture, O., Bearson, S.M., O'Connor, A., Mckean, J., Rodriguez, T.Y., Dekkers, J.C., Nettleton, D., Tuggle, C.K. 2011. Use of bioinformatic SNP predictions in differentially expressed genes to find SNPs associated with Salmonella colonization in swine. Journal of Animal Breeding and Genetics. 128(5):354-365.

Brunelle, B.W., Sensabaugh, G.F. 2012. Nucleotide and phylogenetic analyses of the Chlamydia trachomatis ompA gene indicates it is a hotspot for mutation. BMC Research Notes [serial online]. 5(53). Available: http://www.biomedcentral.com/1756-0500/5/53/abstract.

Huang, T., Uthe, J.J., Bearson, S.M., Demirkale, C.Y., Nettleton, D., Knetter, S.M., Christian, C.M., Ramer-Tait, A.E., Wannemuehler, M.J., Tuggle, C.K. 2011. Distinct peripheral blood RNA responses to Salmonella in pigs differing in Salmonella shedding levels: intersection of IFNG, TLR and miRNA pathways. PLoS One [serial online]. 6(12). Available: http://www.plosone.org/article/info%3Adoi%2F10.1371%2Fjournal.pone.0028768.

Last Modified: 11/20/2014
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