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Sample records for enterica serovar typhimurium

  1. Sensitive and rapid molecular detection assays for Salmonella enterica serovar Typhimurium and Heidelberg

    USDA-ARS?s Scientific Manuscript database

    Salmonella enterica is a significant cause of gastroenteritis worldwide, with S. enterica serovars Typhimurium and Heidelberg being particularly prevalent. S. enterica serovars Typhimurium and Heidelberg have broad host ranges infecting poultry, dairy animals and humans. Traditional methods used fo...

  2. Draft Genome Sequences of Salmonella enterica subsp. enterica Serovars Typhimurium and Nottingham Isolated from Food Products

    PubMed Central

    Zheng, Jie; Ayers, Sherry; Melka, David C.; Curry, Phillip E.; Payne, Justin S.; Laasri, Anna; Wang, Charles; Hammack, Thomas S.; Brown, Eric W.

    2016-01-01

    A quantitative real-time PCR (qPCR) designed to detect Salmonella enterica subsp. enterica serovar Enteritidis, targeting the sdf gene, generated positive results for S. enterica subsp. enterica serovar Typhimurium (CFSAN033950) and S. enterica subsp. enterica serovar Nottingham (CFSAN006803) isolated from food samples. Both strains show pulsed-field gel electrophoresis (PFGE) patterns distinct from those of S. Enteritidis. Here, we report the genome sequences of these two strains. PMID:27445384

  3. Complete Genome and Methylome Sequences of Salmonella enterica subsp. enterica Serovars Typhimurium, Saintpaul, and Stanleyville from the SARA/SARB Collection

    PubMed Central

    Yao, Kuan; Roberts, Richard J.; Allard, Marc W.

    2017-01-01

    ABSTRACT In this announcement, we report the complete genome and methylome sequences of three Salmonella enterica strains from the SARA and SARB collection: S. enterica subsp. enterica serovar Typhimurium (SARA13), S. enterica subsp. enterica serovar Saintpaul (SARA26), and S. enterica subsp. enterica serovar Stanleyville (SARB61). PMID:28302778

  4. Complete, Closed Genome Sequences of 10 Salmonella enterica subsp. enterica Serovar Typhimurium Strains Isolated from Human and Bovine Sources

    PubMed Central

    Nguyen, Scott V.; Bono, James L.; Smith, Timothy P. L.; Fields, Patricia I.; Dinsmore, Blake A.; Santovenia, Monica; Kelley, Christy M.; Wang, Rong; Bosilevac, Joseph M.; Harhay, Gregory P.

    2016-01-01

    Salmonella enterica is a leading cause of enterocolitis for humans and animals. S. enterica subsp. enterica serovar Typhimurium infects a broad range of hosts. To facilitate genomic comparisons among isolates from different sources, we present the complete genome sequences of 10 S. Typhimurium strains, 5 each isolated from human and bovine sources. PMID:27811097

  5. Aspartic Peptide Hydrolases in Salmonella enterica Serovar Typhimurium

    PubMed Central

    Larsen, Rachel A.; Knox, Tina M.; Miller, Charles G.

    2001-01-01

    Two well-characterized enzymes in Salmonella enterica serovar Typhimurium and Escherichia coli are able to hydrolyze N-terminal aspartyl (Asp) dipeptides: peptidase B, a broad-specificity aminopeptidase, and peptidase E, an Asp-specific dipeptidase. A serovar Typhimurium strain lacking both of these enzymes, however, can still utilize most N-terminal Asp dipeptides as sources of amino acids, and extracts of such a strain contain additional enzymatic activities able to hydrolyze Asp dipeptides. Here we report two such activities from extracts of pepB pepE mutant strains of serovar Typhimurium identified by their ability to hydrolyze Asp-Leu. Although each of these activities hydrolyzes Asp-Leu at a measurable rate, the preferred substrates for both are N-terminal isoAsp peptides. One of the activities is a previously characterized isoAsp dipeptidase from E. coli, the product of the iadA gene. The other is the product of the serovar Typhimurium homolog of E. coli ybiK, a gene of previously unknown function. This gene product is a member of the N-terminal nucleophile structural family of amidohydrolases. Like most other members of this family, the mature enzyme is generated from a precursor protein by proteolytic cleavage and the active enzyme is a heterotetramer. Based on its ability to hydrolyze an N-terminal isoAsp tripeptide as well as isoAsp dipeptides, the enzyme appears to be an isoAsp aminopeptidase, and we propose that the gene encoding it be designated iaaA (isoAsp aminopeptidase). A strain lacking both IadA and IaaA in addition to peptidase B and peptidase E has been constructed. This strain utilizes Asp-Leu as a leucine source, and extracts of this strain contain at least one additional, as-yet-uncharacterized, peptidase able to cleave Asp dipeptides. PMID:11325937

  6. Aspartic peptide hydrolases in Salmonella enterica serovar typhimurium.

    PubMed

    Larsen, R A; Knox, T M; Miller, C G

    2001-05-01

    Two well-characterized enzymes in Salmonella enterica serovar Typhimurium and Escherichia coli are able to hydrolyze N-terminal aspartyl (Asp) dipeptides: peptidase B, a broad-specificity aminopeptidase, and peptidase E, an Asp-specific dipeptidase. A serovar Typhimurium strain lacking both of these enzymes, however, can still utilize most N-terminal Asp dipeptides as sources of amino acids, and extracts of such a strain contain additional enzymatic activities able to hydrolyze Asp dipeptides. Here we report two such activities from extracts of pepB pepE mutant strains of serovar Typhimurium identified by their ability to hydrolyze Asp-Leu. Although each of these activities hydrolyzes Asp-Leu at a measurable rate, the preferred substrates for both are N-terminal isoAsp peptides. One of the activities is a previously characterized isoAsp dipeptidase from E. coli, the product of the iadA gene. The other is the product of the serovar Typhimurium homolog of E. coli ybiK, a gene of previously unknown function. This gene product is a member of the N-terminal nucleophile structural family of amidohydrolases. Like most other members of this family, the mature enzyme is generated from a precursor protein by proteolytic cleavage and the active enzyme is a heterotetramer. Based on its ability to hydrolyze an N-terminal isoAsp tripeptide as well as isoAsp dipeptides, the enzyme appears to be an isoAsp aminopeptidase, and we propose that the gene encoding it be designated iaaA (isoAsp aminopeptidase). A strain lacking both IadA and IaaA in addition to peptidase B and peptidase E has been constructed. This strain utilizes Asp-Leu as a leucine source, and extracts of this strain contain at least one additional, as-yet-uncharacterized, peptidase able to cleave Asp dipeptides.

  7. Biofilm formation by Salmonella enterica serovar Typhimurium colonizing solid tumours.

    PubMed

    Crull, Katja; Rohde, Manfred; Westphal, Kathrin; Loessner, Holger; Wolf, Kathrin; Felipe-López, Alfonso; Hensel, Michael; Weiss, Siegfried

    2011-08-01

    Systemic administration of Salmonella enterica serovar Typhimurium to tumour bearing mice results in preferential colonization of the tumours and retardation of tumour growth. Although the bacteria are able to invade the tumour cells in vitro, in tumours they were never detected intracellularly. Ultrastructural analysis of Salmonella-colonized tumours revealed that the bacteria had formed biofilms. Interestingly, depletion of neutrophilic granulocytes drastically reduced biofilm formation. Obviously, bacteria form biofilms in response to the immune reactions of the host. Importantly, we tested Salmonella mutants that were no longer able to form biofilms by deleting central regulators of biofilm formation. Such bacteria could be observed intracellularly in immune cells of the host or in tumour cells. Thus, tumour colonizing S. typhimurium might form biofilms as protection against phagocytosis. Since other bacteria are behaving similarly, solid murine tumours might represent a unique model to study biofilm formation in vivo. © 2011 Blackwell Publishing Ltd.

  8. Complete, closed genome sequences of 10 Salmonella enterica subsp. enterica serovar Typhimurium strains isolated from human and bovine sources

    USDA-ARS?s Scientific Manuscript database

    Salmonella enterica are a leading cause of enterocolitis for humans and animals. S. enterica serovar Typhimurium infects a broad range of hosts. To facilitate genomic comparisons among isolates from different sources, we present the complete genome sequences of ten S. Typhimurium strains, five each...

  9. Polyamines Are Required for Virulence in Salmonella enterica Serovar Typhimurium

    PubMed Central

    Jelsbak, Lotte; Thomsen, Line Elnif; Wallrodt, Inke; Jensen, Peter Ruhdal; Olsen, John Elmerdahl

    2012-01-01

    Sensing and responding to environmental cues is a fundamental characteristic of bacterial physiology and virulence. Here we identify polyamines as novel environmental signals essential for virulence of Salmonella enterica serovar Typhimurium, a major intracellular pathogen and a model organism for studying typhoid fever. Central to its virulence are two major virulence loci Salmonella Pathogenicity Island 1 and 2 (SPI1 and SPI2). SPI1 promotes invasion of epithelial cells, whereas SPI2 enables S. Typhimurium to survive and proliferate within specialized compartments inside host cells. In this study, we show that an S. Typhimurium polyamine mutant is defective for invasion, intracellular survival, killing of the nematode Caenorhabditis elegans and systemic infection of the mouse model of typhoid fever. Virulence of the mutant could be restored by genetic complementation, and invasion and intracellular survival could, as well, be complemented by the addition of exogenous putrescine and spermidine to the bacterial cultures prior to infection. Interestingly, intracellular survival of the polyamine mutant was significantly enhanced above the wild type level by the addition of exogenous putrescine and spermidine to the bacterial cultures prior to infection, indicating that these polyamines function as an environmental signal that primes S. Typhimurium for intracellular survival. Accordingly, experiments addressed at elucidating the roles of these polyamines in infection revealed that expression of genes from both of the major virulence loci SPI1 and SPI2 responded to exogenous polyamines and was reduced in the polyamine mutant. Together our data demonstrate that putrescine and spermidine play a critical role in controlling virulence in S. Typhimurium most likely through stimulation of expression of essential virulence loci. Moreover, our data implicate these polyamines as key signals in S. Typhimurium virulence. PMID:22558361

  10. Proteomic analysis of triclosan resistance in Salmonella enterica serovar Typhimurium.

    PubMed

    Webber, Mark A; Coldham, Nick G; Woodward, Martin J; Piddock, Laura J V

    2008-07-01

    The aim of this study was to determine and compare the proteomes of three triclosan-resistant mutants of Salmonella enterica serovar Typhimurium in order to identify proteins involved in triclosan resistance. The proteomes of three distinct but isogenic triclosan-resistant mutants were determined using two-dimensional liquid chromatography mass separation. Bioinformatics was then used to identify and quantify tryptic peptides in order to determine protein expression. Proteomic analysis of the triclosan-resistant mutants identified a common set of proteins involved in production of pyruvate or fatty acid with differential expression in all mutants, but also demonstrated specific patterns of expression associated with each phenotype. These data show that triclosan resistance can occur via distinct pathways in Salmonella, and demonstrate a novel triclosan resistance network that is likely to have relevance to other pathogenic bacteria subject to triclosan exposure and may provide new targets for development of antimicrobial agents.

  11. Characterization of Salmonella enterica serovar Typhimurium aconitase A.

    PubMed

    Baothman, Othman A S; Rolfe, Matthew D; Green, Jeffrey

    2013-06-01

    Aconitases (Acn) are iron-sulfur proteins that catalyse the reversible isomerization of citrate and isocitrate via the intermediate cis-aconitate in the Krebs cycle. Some Acn proteins are bi-functional and under conditions of iron starvation and oxidative stress lose their iron-sulfur clusters and become post-transcriptional regulators by binding specific mRNA targets. Many bacterial species possess two genetically distinct aconitase proteins, AcnA and AcnB. Current understanding of the regulation and functions of AcnA and AcnB in dual Acn bacteria is based on a model developed in Escherichia coli. Thus, AcnB is the major Krebs cycle enzyme expressed during exponential growth, whereas AcnA is a more stable, stationary phase and stress-induced enzyme, and both E. coli Acns are bi-functional. Here a second dual Acn bacterium, Salmonella enterica serovar Typhimurium (S. Typhimurium), has been analysed. Phenotypic traits of S. Typhimurium acn mutants were consistent with AcnB acting as the major Acn protein. Promoter fusion experiments indicated that acnB transcription was ~10-fold greater than that of acnA and that acnA expression was regulated by the cyclic-AMP receptor protein (CRP, glucose starvation), the fumarate nitrate reduction regulator (FNR, oxygen starvation), the ferric uptake regulator (Fur, iron starvation) and the superoxide response protein (SoxR, oxidative stress). In contrast to E. coli, S. Typhimurium acnA was not induced in the stationary phase. Furthermore, acnA expression was enhanced in an acnB mutant, presumably to partially compensate for the lack of AcnB activity. Isolated S. Typhimurium AcnA protein had kinetic and mRNA-binding properties similar to those described for E. coli AcnA. Thus, the work reported here provides a second example of the regulation and function of AcnA and AcnB proteins in a dual Acn bacterium.

  12. Interaction of Salmonella enterica subspecies enterica serovar Typhimurium and mung bean (Phaseolus aureus) plants.

    PubMed

    Singh, Bhoj Raj; Chandra, Mudit; Agarwal, Ravikant

    2005-03-01

    The effect of Salmonella enterica subspecies enterica serovar Typhimurium, a zoonotic serovar, on mung bean (Phaseolus aureus) cultivar Pant Mung-3 plants was studied. Inoculation of mung bean seeds with Salmonella Typhimurium (7.2 x 10(5) CFU/ml) reduced sprouting rate (P < 0.07). This effect was more pronounced at higher levels of contamination. In the soil inoculated with Salmonella Typhimurium (7.2 x 10(6) CFU/g), germination was retarded and the number of defective sprouts was also significantly higher (P < 0.002). Salmonella Typhimurium grew inside germinating seeds and plant tissues and persisted in seedlings, adult plants, and harvested seedlings dried and stored at room temperature (30 degrees C) up to 45 days. Phaseolus aureus plants grown in sterile soil was resistant to Salmonella Typhimurium infection at 15 days of age and cleared Salmonella from all the aerial parts within 3 h of infection. However, Salmonella Typhimurium could be reisolated from the basal area of the stem and from soil even after 45 days of exposure to the pathogen.

  13. Epidemiology of a Salmonella enterica subsp. enterica Serovar Typhimurium Strain Associated with a Songbird Outbreak

    PubMed Central

    Hernandez, Sonia M.; Keel, Kevin; Sanchez, Susan; Trees, Eija; Gerner-Smidt, Peter; Adams, Jennifer K.; Cheng, Ying; Ray, Al; Martin, Gordon; Presotto, Andrea; Ruder, Mark G.; Brown, Justin; Blehert, David S.; Cottrell, Walter

    2012-01-01

    Salmonella enterica subsp. enterica serovar Typhimurium is responsible for the majority of salmonellosis cases worldwide. This Salmonella serovar is also responsible for die-offs in songbird populations. In 2009, there was an S. Typhimurium epizootic reported in pine siskins in the eastern United States. At the time, there was also a human outbreak with this serovar that was associated with contaminated peanuts. As peanuts are also used in wild-bird food, it was hypothesized that the pine siskin epizootic was related to this human outbreak. A comparison of songbird and human S. Typhimurium pulsed-field gel electrophoresis (PFGE) patterns revealed that the epizootic was attributed not to the peanut-associated strain but, rather, to a songbird strain first characterized from an American goldfinch in 1998. This same S. Typhimurium strain (PFGE type A3) was also identified in the PulseNet USA database, accounting for 137 of 77,941 total S. Typhimurium PFGE entries. A second molecular typing method, multiple-locus variable-number tandem-repeat analysis (MLVA), confirmed that the same strain was responsible for the pine siskin epizootic in the eastern United States but was distinct from a genetically related strain isolated from pine siskins in Minnesota. The pine siskin A3 strain was first encountered in May 2008 in an American goldfinch and later in a northern cardinal at the start of the pine siskin epizootic. MLVA also confirmed the clonal nature of S. Typhimurium in songbirds and established that the pine siskin epizootic strain was unique to the finch family. For 2009, the distribution of PFGE type A3 in passerines and humans mirrored the highest population density of pine siskins for the East Coast. PMID:22885752

  14. Epidemiology of a Salmonella enterica subsp. Enterica serovar Typhimurium strain associated with a songbird outbreak.

    USGS Publications Warehouse

    Blehert, David S.; Hernandez, Sonia M.; Keel, Kevin; Sanchez, Susan; Trees, Eija; ,

    2012-01-01

    Salmonella enterica subsp. enterica serovar Typhimurium is responsible for the majority of salmonellosis cases worldwide. This Salmonella serovar is also responsible for die-offs in songbird populations. In 2009, there was an S. Typhimurium epizootic reported in pine siskins in the eastern United States. At the time, there was also a human outbreak with this serovar that was associated with contaminated peanuts. As peanuts are also used in wild-bird food, it was hypothesized that the pine siskin epizootic was related to this human outbreak. A comparison of songbird and human S. Typhimurium pulsed-field gel electrophoresis (PFGE) patterns revealed that the epizootic was attributed not to the peanut-associated strain but, rather, to a songbird strain first characterized from an American goldfinch in 1998. This same S. Typhimurium strain (PFGE type A3) was also identified in the PulseNet USA database, accounting for 137 of 77,941 total S. Typhimurium PFGE entries. A second molecular typing method, multiple-locus variable-number tandem-repeat analysis (MLVA), confirmed that the same strain was responsible for the pine siskin epizootic in the eastern United States but was distinct from a genetically related strain isolated from pine siskins in Minnesota. The pine siskin A3 strain was first encountered in May 2008 in an American goldfinch and later in a northern cardinal at the start of the pine siskin epizootic. MLVA also confirmed the clonal nature of S. Typhimurium in songbirds and established that the pine siskin epizootic strain was unique to the finch family. For 2009, the distribution of PFGE type A3 in passerines and humans mirrored the highest population density of pine siskins for the East Coast.

  15. Epidemiology of a Salmonella enterica subsp. enterica serovar Typhimurium strain associated with a songbird outbreak.

    PubMed

    Hernandez, Sonia M; Keel, Kevin; Sanchez, Susan; Trees, Eija; Gerner-Smidt, Peter; Adams, Jennifer K; Cheng, Ying; Ray, Al; Martin, Gordon; Presotto, Andrea; Ruder, Mark G; Brown, Justin; Blehert, David S; Cottrell, Walter; Maurer, John J

    2012-10-01

    Salmonella enterica subsp. enterica serovar Typhimurium is responsible for the majority of salmonellosis cases worldwide. This Salmonella serovar is also responsible for die-offs in songbird populations. In 2009, there was an S. Typhimurium epizootic reported in pine siskins in the eastern United States. At the time, there was also a human outbreak with this serovar that was associated with contaminated peanuts. As peanuts are also used in wild-bird food, it was hypothesized that the pine siskin epizootic was related to this human outbreak. A comparison of songbird and human S. Typhimurium pulsed-field gel electrophoresis (PFGE) patterns revealed that the epizootic was attributed not to the peanut-associated strain but, rather, to a songbird strain first characterized from an American goldfinch in 1998. This same S. Typhimurium strain (PFGE type A3) was also identified in the PulseNet USA database, accounting for 137 of 77,941 total S. Typhimurium PFGE entries. A second molecular typing method, multiple-locus variable-number tandem-repeat analysis (MLVA), confirmed that the same strain was responsible for the pine siskin epizootic in the eastern United States but was distinct from a genetically related strain isolated from pine siskins in Minnesota. The pine siskin A3 strain was first encountered in May 2008 in an American goldfinch and later in a northern cardinal at the start of the pine siskin epizootic. MLVA also confirmed the clonal nature of S. Typhimurium in songbirds and established that the pine siskin epizootic strain was unique to the finch family. For 2009, the distribution of PFGE type A3 in passerines and humans mirrored the highest population density of pine siskins for the East Coast.

  16. Natural surface coating to inactivate Salmonella enterica Serovar Typhimurium and maintain quality of cherry tomatoes

    USDA-ARS?s Scientific Manuscript database

    The objectives of the present study were to investigate the effectiveness of zein-based coatings in reducing populations of Salmonella enterica serovar Typhimurium and preserving quality of cherry tomatoes. Tomatoes were inoculated with a cocktail of S. Typhimurium LT2 plus three mutants on the smoo...

  17. Flagella-independent surface motility in Salmonella enterica serovar Typhimurium

    PubMed Central

    Park, Sun-Yang; Pontes, Mauricio H.; Groisman, Eduardo A.

    2015-01-01

    Flagella are multiprotein complexes necessary for swimming and swarming motility. In Salmonella enterica serovar Typhimurium, flagella-mediated motility is repressed by the PhoP/PhoQ regulatory system. We now report that Salmonella can move on 0.3% agarose media in a flagella-independent manner when experiencing the PhoP/PhoQ-inducing signal low Mg2+. This motility requires the PhoP-activated mgtA, mgtC, and pagM genes, which specify a Mg2+ transporter, an inhibitor of Salmonella’s own F1Fo ATPase, and a small protein of unknown function, respectively. The MgtA and MgtC proteins are necessary for pagM expression because pagM mRNA levels were lower in mgtA and mgtC mutants than in wild-type Salmonella, and also because pagM expression from a heterologous promoter rescued motility in mgtA and mgtC mutants. PagM promotes group motility by a surface protein(s), as a pagM-expressing strain conferred motility upon a pagM null mutant, and proteinase K treatment eliminated motility. The pagM gene is rarely found outside subspecies I of S. enterica and often present in nonfunctional allelic forms in organisms lacking the identified motility. Deletion of the pagM gene reduced bacterial replication on 0.3% agarose low Mg2+ media but not in low Mg2+ liquid media. Our findings define a form of motility that allows Salmonella to scavenge nutrients and to escape toxic compounds in low Mg2+ semisolid environments. PMID:25624475

  18. Global Genomic Epidemiology of Salmonella enterica Serovar Typhimurium DT104

    PubMed Central

    Hendriksen, Rene S.; Le Hello, Simon; Weill, François-Xavier; Baggesen, Dorte Lau; Jun, Se-Ran; Lund, Ole; Crook, Derrick W.; Wilson, Daniel J.; Aarestrup, Frank M.

    2016-01-01

    It has been 30 years since the initial emergence and subsequent rapid global spread of multidrug-resistant Salmonella enterica serovar Typhimurium DT104 (MDR DT104). Nonetheless, its origin and transmission route have never been revealed. We used whole-genome sequencing (WGS) and temporally structured sequence analysis within a Bayesian framework to reconstruct temporal and spatial phylogenetic trees and estimate the rates of mutation and divergence times of 315 S. Typhimurium DT104 isolates sampled from 1969 to 2012 from 21 countries on six continents. DT104 was estimated to have emerged initially as antimicrobial susceptible in ∼1948 (95% credible interval [CI], 1934 to 1962) and later became MDR DT104 in ∼1972 (95% CI, 1972 to 1988) through horizontal transfer of the 13-kb Salmonella genomic island 1 (SGI1) MDR region into susceptible strains already containing SGI1. This was followed by multiple transmission events, initially from central Europe and later between several European countries. An independent transmission to the United States and another to Japan occurred, and from there MDR DT104 was probably transmitted to Taiwan and Canada. An independent acquisition of resistance genes took place in Thailand in ∼1975 (95% CI, 1975 to 1990). In Denmark, WGS analysis provided evidence for transmission of the organism between herds of animals. Interestingly, the demographic history of Danish MDR DT104 provided evidence for the success of the program to eradicate Salmonella from pig herds in Denmark from 1996 to 2000. The results from this study refute several hypotheses on the evolution of DT104 and suggest that WGS may be useful in monitoring emerging clones and devising strategies for prevention of Salmonella infections. PMID:26944846

  19. Global Genomic Epidemiology of Salmonella enterica Serovar Typhimurium DT104

    SciTech Connect

    Leekitcharoenphon, Pimlapas; Hendriksen, Rene S.; Le Hello, Simon; Weill, François-Xavier; Baggesen, Dorte Lau; Jun, Se-Ran; Ussery, David W.; Lund, Ole; Crook, Derrick W.; Wilson, Daniel J.; Aarestrup, Frank M.

    2016-03-04

    It has been 30 years since the initial emergence and subsequent rapid global spread of multidrug-resistant Salmonella enterica serovar Typhimurium DT104 (MDR DT104). Nonetheless, its origin and transmission route have never been revealed. In this paper, we used whole-genome sequencing (WGS) and temporally structured sequence analysis within a Bayesian framework to reconstruct temporal and spatial phylogenetic trees and estimate the rates of mutation and divergence times of 315 S. Typhimurium DT104 isolates sampled from 1969 to 2012 from 21 countries on six continents. DT104 was estimated to have emerged initially as antimicrobial susceptible in ~1948 (95% credible interval [CI], 1934 to 1962) and later became MDR DT104 in ~1972 (95% CI, 1972 to 1988) through horizontal transfer of the 13-kb Salmonella genomic island 1 (SGI1) MDR region into susceptible strains already containing SGI1. This was followed by multiple transmission events, initially from central Europe and later between several European countries. An independent transmission to the United States and another to Japan occurred, and from there MDR DT104 was probably transmitted to Taiwan and Canada. An independent acquisition of resistance genes took place in Thailand in ~1975 (95% CI, 1975 to 1990). In Denmark, WGS analysis provided evidence for transmission of the organism between herds of animals. Interestingly, the demographic history of Danish MDR DT104 provided evidence for the success of the program to eradicate Salmonella from pig herds in Denmark from 1996 to 2000. Finally, the results from this study refute several hypotheses on the evolution of DT104 and suggest that WGS may be useful in monitoring emerging clones and devising strategies for prevention of Salmonella infections.

  20. Global Genomic Epidemiology of Salmonella enterica Serovar Typhimurium DT104

    DOE PAGES

    Leekitcharoenphon, Pimlapas; Hendriksen, Rene S.; Le Hello, Simon; ...

    2016-03-04

    It has been 30 years since the initial emergence and subsequent rapid global spread of multidrug-resistant Salmonella enterica serovar Typhimurium DT104 (MDR DT104). Nonetheless, its origin and transmission route have never been revealed. In this paper, we used whole-genome sequencing (WGS) and temporally structured sequence analysis within a Bayesian framework to reconstruct temporal and spatial phylogenetic trees and estimate the rates of mutation and divergence times of 315 S. Typhimurium DT104 isolates sampled from 1969 to 2012 from 21 countries on six continents. DT104 was estimated to have emerged initially as antimicrobial susceptible in ~1948 (95% credible interval [CI], 1934more » to 1962) and later became MDR DT104 in ~1972 (95% CI, 1972 to 1988) through horizontal transfer of the 13-kb Salmonella genomic island 1 (SGI1) MDR region into susceptible strains already containing SGI1. This was followed by multiple transmission events, initially from central Europe and later between several European countries. An independent transmission to the United States and another to Japan occurred, and from there MDR DT104 was probably transmitted to Taiwan and Canada. An independent acquisition of resistance genes took place in Thailand in ~1975 (95% CI, 1975 to 1990). In Denmark, WGS analysis provided evidence for transmission of the organism between herds of animals. Interestingly, the demographic history of Danish MDR DT104 provided evidence for the success of the program to eradicate Salmonella from pig herds in Denmark from 1996 to 2000. Finally, the results from this study refute several hypotheses on the evolution of DT104 and suggest that WGS may be useful in monitoring emerging clones and devising strategies for prevention of Salmonella infections.« less

  1. Draft Genome Sequences of 40 Salmonella enterica Serovar Typhimurium Strains Isolated from Humans and Food in Brazil

    PubMed Central

    Almeida, Fernanda; Medeiros, Marta Inês Cazentini; Rodrigues, Dália Prazeres; Payne, Justin; Timme, Ruth E.

    2016-01-01

    Salmonellosis is an important health problem worldwide and Salmonella enterica serovar Typhimurium is one of the most common isolated serovars. Here, we reported the draft genomes of 40 S. Typhimurium strains isolated from humans and food in Brazil. These draft genomes will improve phylogenetic analysis and will help enhance our understanding of strains of this serovar isolated in Brazil. PMID:27660768

  2. Draft Genome Sequences of 40 Salmonella enterica Serovar Typhimurium Strains Isolated from Humans and Food in Brazil.

    PubMed

    Almeida, Fernanda; Medeiros, Marta Inês Cazentini; Rodrigues, Dália Prazeres; Payne, Justin; Timme, Ruth E; Allard, Marc W; Falcão, Juliana Pfrimer

    2016-09-22

    Salmonellosis is an important health problem worldwide and Salmonella enterica serovar Typhimurium is one of the most common isolated serovars. Here, we reported the draft genomes of 40 S Typhimurium strains isolated from humans and food in Brazil. These draft genomes will improve phylogenetic analysis and will help enhance our understanding of strains of this serovar isolated in Brazil.

  3. Antimicrobial Resistance of Salmonella enterica Serovar Typhimurium in Shanghai, China

    PubMed Central

    Wang, Jinyan; Li, Yongrui; Xu, Xuebin; Liang, Beibei; Wu, Fuli; Yang, Xiaoxia; Ma, Qiuxia; Yang, Chaojie; Hu, Xiaofeng; Liu, Hongbo; Li, Hao; Sheng, Chunyu; Xie, Jing; Du, Xinying; Hao, Rongzhang; Qiu, Shaofu; Song, Hongbin

    2017-01-01

    We aimed to analyze the antimicrobial resistance phenotypes and to elucidate the molecular mechanisms underlying resistance to cephalosporins, ciprofloxacin, and azithromycin in Salmonella enterica serovar Typhimurium isolates identified from patients with diarrhea in Shanghai. The isolates showed high rates of resistance to traditional antimicrobials, and 20.6, 12.7, and 5.5% of them exhibited decreased susceptibility to cephalosporins, ciprofloxacin, and azithromycin, respectively. Notably, 473 (84.6%) isolates exhibited multidrug resistance (MDR), including 161 (28.8%) isolates that showed an ACSSuT profile. Twenty-two MDR isolates concurrently exhibited decreased susceptibility to cephalosporins and ciprofloxacin, and six of them were co-resistant to azithromycin. Of all the 71 isolates with decreased susceptibility to ciprofloxacin, 65 showed at least one mutation (D87Y, D87N, or D87G) in gyrA, among which seven isolates simultaneously had mutations of parC (S80R) (n = 6) or parC (T57S/S80R) (n = 1), while 49 isolates with either zero or one mutation in gyrA contained plasmid-mediated quinolone resistance (PMQR) genes including qnrB, qnrS, and aac(6′)-Ib-cr. Among the 115 cephalosporin-resistant isolates, the most common ESBL gene was blaCTX-M, followed by blaTEM-1, blaOXA-1, and blaSHV -12. Eight subtypes of blaCTX-M were identified and blaCTX-M-14 (n = 22) and blaCTX-M-55 (n = 31) were found to be dominant. To the best of our knowledge, this is the first report of the presence of blaCTX-M-123 and blaCTX-M-125 in S. Typhimurium. Besides, mphA gene was identified in 15 of the 31 azithromycin-resistant isolates. Among the 22 isolates with reduced susceptibility to cephalosporins and ciprofloxacin, 15 contained ESBL and PMQR genes. Coexistence of these genes lead to the emergence of MDR and the transmission of them will pose great difficulties in S. Typhimurium treatments. Therefore, surveillance for these MDR isolates should be enhanced. PMID:28400764

  4. Internal Colonization of Salmonella enterica Serovar Typhimurium in Tomato Plants

    PubMed Central

    Gu, Ganyu; Hu, Jiahuai; Cevallos-Cevallos, Juan M.; Richardson, Susanna M.; Bartz, Jerry A.; van Bruggen, Ariena H. C.

    2011-01-01

    Several Salmonella enterica outbreaks have been traced back to contaminated tomatoes. In this study, the internalization of S. enterica Typhimurium via tomato leaves was investigated as affected by surfactants and bacterial rdar morphotype, which was reported to be important for the environmental persistence and attachment of Salmonella to plants. Surfactants, especially Silwet L-77, promoted ingress and survival of S. enterica Typhimurium in tomato leaves. In each of two experiments, 84 tomato plants were inoculated two to four times before fruiting with GFP-labeled S. enterica Typhimurium strain MAE110 (with rdar morphotype) or MAE119 (without rdar). For each inoculation, single leaflets were dipped in 109 CFU/ml Salmonella suspension with Silwet L-77. Inoculated and adjacent leaflets were tested for Salmonella survival for 3 weeks after each inoculation. The surface and pulp of ripe fruits produced on these plants were also examined for Salmonella. Populations of both Salmonella strains in inoculated leaflets decreased during 2 weeks after inoculation but remained unchanged (at about 104 CFU/g) in week 3. Populations of MAE110 were significantly higher (P<0.05) than those of MAE119 from day 3 after inoculation. In the first year, nine fruits collected from one of the 42 MAE119 inoculated plants were positive for S. enterica Typhimurium. In the second year, Salmonella was detected in adjacent non-inoculated leaves of eight tomato plants (five inoculated with strain MAE110). The pulp of 12 fruits from two plants inoculated with MAE110 was Salmonella positive (about 106 CFU/g). Internalization was confirmed by fluorescence and confocal laser microscopy. For the first time, convincing evidence is presented that S. enterica can move inside tomato plants grown in natural field soil and colonize fruits at high levels without inducing any symptoms, except for a slight reduction in plant growth. PMID:22096553

  5. Genomic heterogeneity of Salmonella enterica serovar Typhimurium bacteriuria from chronic infection.

    PubMed

    Octavia, Sophie; Wang, Qinning; Tanaka, Mark M; Sintchenko, Vitali; Lan, Ruiting

    2017-03-07

    We sequenced the genomes of 14 sequential Salmonella enterica serovar Typhimurium isolates obtained over a five year period from a patient with persistent Salmonella bacteriuria. The isolates formed five distinct lineages; two of which co-existed over four years. We inferred that the observed within-patient variation resulted from mutation events.

  6. Tetracycline promotes the expression of ten fimbrial operons in specific Salmonella enterica serovar Typhimurium isolates

    USDA-ARS?s Scientific Manuscript database

    Multidrug-resistant (MDR) Salmonella is associated with increased morbidity in humans and presents an important food safety concern. Antibiotic resistance among isolates of Salmonella enterica serovar Typhimurium has become especially prevalent as over 27 per cent of isolates from humans in the Unit...

  7. Hydrogen-Stimulated carbon acquisition and conservation in salmonella enterica serovar typhimurium

    USDA-ARS?s Scientific Manuscript database

    Salmonella enterica serovar Typhimurium can utilize molecular hydrogen for growth and amino acid transport during anaerobic growth. Via microarray we identified H2 gas-affected gene expression changes in Salmonella. Addition of H2 caused altered expression of 965 genes; 176 genes were H2-up-regulate...

  8. Swarm and swim motilities of Salmonella enterica serovar Typhimurium and role of osmoregulated periplasmic glucans

    USDA-ARS?s Scientific Manuscript database

    Background: Salmonella enterica serovar Typhimurium strains synthesize osmoregulated periplasmic glucans (OPGs) under low osmolarity conditions (< 70 mos mol l-1). OPG synthesis is not observed when cells are grown in iso- or hyper-osmotic media (> 400 mos mol l-1). Mutation in OPG structural gene...

  9. Antibiotics induce the expression of attachment genes in specific isolates of Salmonella enterica serovar Typhimurium

    USDA-ARS?s Scientific Manuscript database

    More than 27 percent of Salmonella enterica serovar Typhimurium isolates from humans in the United States are resistant to three or more antibiotics. This presents an important food safety concern as multidrug-resistant (MDR) Salmonella is associated with increased morbidity in humans. It has been...

  10. Polynucleotide phosphorlyase (PNPase) is required for Salmonella enterica serovar Typhimurium colonization in swine

    USDA-ARS?s Scientific Manuscript database

    The pnp gene encodes polynucleotide phosphorylase, an exoribonuclease involved in RNA degradation. A mutation in the pnp gene was previously identified by our group in a signature-tagged mutagenesis screen designed to search for Salmonella enterica serovar Typhimurium genes required for survival in...

  11. Intracellular Voyeurism: Examining the Modulation of Host Cell Activities bySalmonella enterica Serovar Typhimurium.

    PubMed

    Szeto, Jason; Brumell, John H

    2005-11-01

    Salmonella spp. can infect host cells by gaining entry through phagocytosis or by inducing host cell membrane ruffling that facilitates bacterial uptake. With its wide host range, Salmonella enterica serovar Typhimurium has proven to be an important model organism for studying intracellular bacterial pathogenesis. Upon entry into host cells, serovar Typhimurium typically resides within a membrane-bound compartment termed the Salmonella-containing vacuole (SCV). From the SCV, serovar Typhimurium can inject several effector proteins that subvert many normal host cell systems, including endocytic trafficking, cytoskeletal rearrangements, lipid signaling and distribution, and innate and adaptive host defenses. The study of these intracellular events has been made possible through the use of various imaging techniques, ranging from classic methods of transmission electron microscopy to advanced livecell fluorescence confocal microscopy. In addition, DNA microarrays have now been used to provide a "snapshot" of global gene expression in serovar Typhimurium residing within the infected host cell. This review describes key aspects of Salmonella-induced subversion of host cell activities, providing examples of imaging that have been used to elucidate these events. Serovar Typhimurium engages specific host cell machinery from initial contact with the host cell to replication within the SCV. This continuous interaction with the host cell has likely contributed to the extensive arsenal that serovar Typhimurium now possesses, including two type III secretion systems, a range of ammunition in the form of TTSS effectors, and a complex genetic regulatory network that coordinates the expression of hundreds of virulence factors.

  12. Analysis of antimicrobial resistance genes detected in multidrug-resistant salmonella enterica serovar typhimurium isolated from food animals

    USDA-ARS?s Scientific Manuscript database

    The development of multi drug resistance (MDR) in foodborne pathogens such as Salmonella enterica is a concern for both animal and human health. MDR Salmonella enterica serovar Typhimurium is the most prevalent penta-resistant serovar isolated from animals as part of the National Antimicrobial Resis...

  13. TolA mediates the differential detergent resistance pattern between the Salmonella enterica subsp. enterica serovars Typhi and Typhimurium.

    PubMed

    Lahiri, Amit; Ananthalakshmi, T K; Nagarajan, Arvindhan G; Ray, Seemun; Chakravortty, Dipshikha

    2011-05-01

    The tol-pal genes are essential for maintaining the outer membrane integrity and detergent resistance in various Gram-negative bacteria, including Salmonella. The role of TolA has been well established for the bile resistance of Salmonella enterica subsp. enterica serovar Typhimurium. We compared the bile resistance pattern between the S. enterica serovars Typhi and Typhimurium and observed that Typhi is more resistant to bile-mediated damage. A closer look revealed a significant difference in the TolA sequence between the two serovars which contributes to the differential detergent resistance. The tolA knockout of both the serovars behaves completely differently in terms of membrane organization and morphology. The role of the Pal proteins and difference in LPS organization between the two serovars were verified and were found to have no direct connection with the altered bile resistance. In normal Luria broth (LB), S. Typhi ΔtolA is filamentous while S. Typhimurium ΔtolA grows as single cells, similar to the wild-type. In low osmolarity LB, however, S. Typhimurium ΔtolA started chaining and S. Typhi ΔtolA showed no growth. Further investigation revealed that the chaining phenomenon observed was the result of failure of the outer membrane to separate in the dividing cells. Taken together, the results substantiate the evolution of a shorter TolA in S. Typhi to counteract high bile concentrations, at the cost of lower osmotic tolerance.

  14. Refined live attenuated Salmonella enterica serovar Typhimurium and Enteritidis vaccines mediate homologous and heterologous serogroup protection in mice.

    PubMed

    Tennant, Sharon M; Schmidlein, Patrick; Simon, Raphael; Pasetti, Marcela F; Galen, James E; Levine, Myron M

    2015-12-01

    Invasive nontyphoidal Salmonella (NTS) infections constitute a major health problem among infants and toddlers in sub-Saharan Africa; these infections also occur in infants and the elderly in developed countries. We genetically engineered a Salmonella enterica serovar Typhimurium strain of multilocus sequence type 313, the predominant genotype circulating in sub-Saharan Africa. We evaluated the capacities of S. Typhimurium and Salmonella enterica serovar Enteritidis ΔguaBA ΔclpX live oral vaccines to protect mice against a highly lethal challenge dose of the homologous serovar and determined protection against other group B and D serovars circulating in sub-Saharan Africa. The vaccines S. Typhimurium CVD 1931 and S. Enteritidis CVD 1944 were immunogenic and protected BALB/c mice against 10,000 50% lethal doses (LD50) of S. Typhimurium or S. Enteritidis, respectively. S. Typhimurium CVD 1931 protected mice against the group B serovar Salmonella enterica serovar Stanleyville (91% vaccine efficacy), and S. Enteritidis CVD 1944 protected mice against the group D serovar Salmonella enterica serovar Dublin (85% vaccine efficacy). High rates of survival were observed when mice were infected 12 weeks postimmunization, indicating that the vaccines elicited long-lived protective immunity. Whereas CVD 1931 did not protect against S. Enteritidis R11, CVD 1944 did mediate protection against S. Typhimurium D65 (81% efficacy). These findings suggest that a bivalent (S. Typhimurium and S. Enteritidis) vaccine would provide broad protection against the majority of invasive NTS infections in sub-Saharan Africa.

  15. Refined Live Attenuated Salmonella enterica Serovar Typhimurium and Enteritidis Vaccines Mediate Homologous and Heterologous Serogroup Protection in Mice

    PubMed Central

    Schmidlein, Patrick; Simon, Raphael; Pasetti, Marcela F.; Galen, James E.; Levine, Myron M.

    2015-01-01

    Invasive nontyphoidal Salmonella (NTS) infections constitute a major health problem among infants and toddlers in sub-Saharan Africa; these infections also occur in infants and the elderly in developed countries. We genetically engineered a Salmonella enterica serovar Typhimurium strain of multilocus sequence type 313, the predominant genotype circulating in sub-Saharan Africa. We evaluated the capacities of S. Typhimurium and Salmonella enterica serovar Enteritidis ΔguaBA ΔclpX live oral vaccines to protect mice against a highly lethal challenge dose of the homologous serovar and determined protection against other group B and D serovars circulating in sub-Saharan Africa. The vaccines S. Typhimurium CVD 1931 and S. Enteritidis CVD 1944 were immunogenic and protected BALB/c mice against 10,000 50% lethal doses (LD50) of S. Typhimurium or S. Enteritidis, respectively. S. Typhimurium CVD 1931 protected mice against the group B serovar Salmonella enterica serovar Stanleyville (91% vaccine efficacy), and S. Enteritidis CVD 1944 protected mice against the group D serovar Salmonella enterica serovar Dublin (85% vaccine efficacy). High rates of survival were observed when mice were infected 12 weeks postimmunization, indicating that the vaccines elicited long-lived protective immunity. Whereas CVD 1931 did not protect against S. Enteritidis R11, CVD 1944 did mediate protection against S. Typhimurium D65 (81% efficacy). These findings suggest that a bivalent (S. Typhimurium and S. Enteritidis) vaccine would provide broad protection against the majority of invasive NTS infections in sub-Saharan Africa. PMID:26351285

  16. Defining the Core Genome of Salmonella enterica Serovar Typhimurium for Genomic Surveillance and Epidemiological Typing

    PubMed Central

    Fu, Songzhe; Octavia, Sophie; Tanaka, Mark M.; Sintchenko, Vitali

    2015-01-01

    Salmonella enterica serovar Typhimurium is the most common Salmonella serovar causing foodborne infections in Australia and many other countries. Twenty-one S. Typhimurium strains from Salmonella reference collection A (SARA) were analyzed using Illumina high-throughput genome sequencing. Single nucleotide polymorphisms (SNPs) in 21 SARA strains ranged from 46 to 11,916 SNPs, with an average of 1,577 SNPs per strain. Together with 47 strains selected from publicly available S. Typhimurium genomes, the S. Typhimurium core genes (STCG) were determined. The STCG consist of 3,846 genes, a set that is much larger than that of the 2,882 Salmonella core genes (SCG) found previously. The STCG together with 1,576 core intergenic regions (IGRs) were defined as the S. Typhimurium core genome. Using 93 S. Typhimurium genomes from 13 epidemiologically confirmed community outbreaks, we demonstrated that typing based on the S. Typhimurium core genome (STCG plus core IGRs) provides superior resolution and higher discriminatory power than that based on SCG for outbreak investigation and molecular epidemiology of S. Typhimurium. STCG and STCG plus core IGR typing achieved 100% separation of all outbreaks compared to that of SCG typing, which failed to separate isolates from two outbreaks from background isolates. Defining the S. Typhimurium core genome allows standardization of genes/regions to be used for high-resolution epidemiological typing and genomic surveillance of S. Typhimurium. PMID:26019201

  17. Salmonella enterica Serovar Typhimurium and Escherichia coli Contamination of Root and Leaf Vegetables Grown in Soils with Incorporated Bovine Manure

    PubMed Central

    Natvig, Erin E.; Ingham, Steven C.; Ingham, Barbara H.; Cooperband, Leslie R.; Roper, Teryl R.

    2002-01-01

    Bovine manure, with or without added Salmonella enterica serovar Typhimurium (three strains), was incorporated into silty clay loam (SCL) and loamy sand (LS) soil beds (53- by 114-cm surface area, 17.5 cm deep) and maintained in two controlled-environment chambers. The S. enterica serovar Typhimurium inoculum was 4 to 5 log CFU/g in manure-fertilized soil. The conditions in the two environmental chambers, each containing inoculated and uninoculated beds of manure-fertilized soil, simulated daily average Madison, Wis., weather conditions (hourly temperatures, rainfall, daylight, and humidity) for a 1 March or a 1 June manure application and subsequent vegetable growing seasons ending 9 August or 28 September, respectively. Core soil samples were taken biweekly from both inoculated and uninoculated soil beds in each chamber. Radishes, arugula, and carrots were planted in soil beds, thinned, and harvested. Soils, thinned vegetables, and harvested vegetables were analyzed for S. enterica serovar Typhimurium and Escherichia coli (indigenous in manure). After the 1 March manure application, S. enterica serovar Typhimurium was detected at low levels in both soils on 31 May, but not on vegetables planted 1 May and harvested 12 July from either soil. After the 1 June manure application, S. enterica serovar Typhimurium was detected in SCL soil on 7 September and on radishes and arugula planted in SCL soil on 15 August and harvested on 27 September. In LS soil, S. enterica serovar Typhimurium died at a similar rate (P ≥ 0.05) after the 1 June manure application and was less often detected on arugula and radishes harvested from this soil compared to the SCL soil. Pathogen levels on vegetables were decreased by washing. Manure application in cool (daily average maximum temperature of <10°C) spring conditions is recommended to ensure that harvested vegetables are not contaminated with S. enterica serovar Typhimurium. Manure application under warmer (daily average maximum

  18. Induction and Resuscitation of Viable but Nonculturable Salmonella enterica Serovar Typhimurium DT104†

    PubMed Central

    Gupte, A. R.; de Rezende, C. L. E.; Joseph, S. W.

    2003-01-01

    Salmonella enterica serovar Typhimurium DT104 11601was tested for its ability to maintain viability in minimal, chemically defined solutions. Periodic monitoring of growth and survival in microcosms of different ion concentrations, maintained at various temperatures, showed a gradual decline in culturable organisms (∼235 days) at 5°C. Organisms maintained at a higher temperature (21°C) showed continuous, equivalent CFU per milliliter (∼106) up to 400 days after inoculation. Fluorescence microscopy with Baclight revealed that nonculturable cells were actually viable, while observations with scanning electron microscopy showed that the cells had retained their structural integrity. Temperature upshift (56°C ± 0.5, 15 s) of the nonculturable organisms (5°C) in Trypticase soy broth followed by immediate inoculation onto Trypticase soy agar (TSA) gave evidence of resuscitation. Interestingly, S. enterica serovar Typhimurium DT104 from the microcosms at either 5°C (1 to 200 days) or 21°C (1 to 250 days) did not show enhanced growth after intermittent inoculation onto catalase-supplemented TSA. Furthermore, cells from 21°C microcosms exposed to oxidative and osmotic stress showed greater resistance to stresses over increasing times of exposure than did recently grown cells. It is possible that the exceptional survivability and resilience of this particular strain may in part reflect the growing importance of this multidrug-resistant organism, in general, as a cause of intestinal disease in humans. The fact that S. enterica serovar Typhimurium DT104 11601 is capable of modifying its physiological characteristics, including entry into and recovery from the viable but nonculturable state, suggests the overall possibility that S. enterica serovar Typhimurium DT104 may be able to respond uniquely to various adverse environmental conditions. PMID:14602627

  19. Draft Genome Sequence of the Tumor-Targeting Salmonella enterica Serovar Typhimurium Strain SL7207

    PubMed Central

    Johnson, Síle A.; Ormsby, Michael J.

    2017-01-01

    ABSTRACT Salmonella enterica serovar Typhimurium strain SL7207 is a genetically modified derivative of strain SL1344, which preferentially accumulates in tumors and can be used as a vehicle for tissue-specific gene delivery in vivo. Here, we report the draft genome sequence of SL7207, confirming a purported aroA deletion and four single-nucleotide polymorphisms compared to SL1344. PMID:28153911

  20. Salmonella enterica Serovar Typhimurium Response Involved in Attenuation of Pathogen Intracellular Proliferation

    PubMed Central

    Cano, David A.; Martínez-Moya, Marina; Pucciarelli, M. Graciela; Groisman, Eduardo A.; Casadesús, Josep; García-Del Portillo, Francisco

    2001-01-01

    Salmonella enterica serovar Typhimurium proliferates within cultured epithelial and macrophage cells. Intracellular bacterial proliferation is, however, restricted within normal fibroblast cells. To characterize this phenomenon in detail, we investigated the possibility that the pathogen itself might contribute to attenuating the intracellular growth rate. S. enterica serovar Typhimurium mutants were selected in normal rat kidney fibroblasts displaying an increased intracellular proliferation rate. These mutants harbored loss-of-function mutations in the virulence-related regulatory genes phoQ, rpoS, slyA, and spvR. Lack of a functional PhoP-PhoQ system caused the most dramatic change in the intracellular growth rate. phoP- and phoQ-null mutants exhibited an intracellular growth rate 20- to 30-fold higher than that of the wild-type strain. This result showed that the PhoP-PhoQ system exerts a master regulatory function for preventing bacterial overgrowth within fibroblasts. In addition, an overgrowing clone was isolated harboring a mutation in a previously unknown serovar Typhimurium open reading frame, named igaA for intracellular growth attenuator. Mutations in other serovar Typhimurium virulence genes, such as ompR, dam, crp, cya, mviA, spiR (ssrA), spiA, and rpoE, did not result in pathogen intracellular overgrowth. Nonetheless, lack of either SpiA or the alternate sigma factor RpoE led to a substantial decrease in intracellular bacterial viability. These results prove for the first time that specific serovar Typhimurium virulence regulators are involved in a response designed to attenuate the intracellular growth rate within a nonphagocytic host cell. This growth-attenuating response is accompanied by functions that ensure the viability of intracellular bacteria. PMID:11553591

  1. Transcriptomic Responses of Salmonella enterica Serovars Enteritidis and Typhimurium to Chlorine-Based Oxidative Stress▿ †

    PubMed Central

    Wang, Siyun; Phillippy, Adam M.; Deng, Kaiping; Rui, Xiaoqian; Li, Zengxin; Tortorello, Mary Lou; Zhang, Wei

    2010-01-01

    Salmonella enterica serovars Enteritidis and Typhimurium are the leading causative agents of salmonellosis in the United States. S. Enteritidis is predominantly associated with contamination of shell eggs and egg products, whereas S. Typhimurium is frequently linked to tainted poultry meats, fresh produce, and recently, peanut-based products. Chlorine is an oxidative disinfectant commonly used in the food industry to sanitize the surfaces of foods and food processing facilities (e.g., shell eggs and poultry meats). However, chlorine disinfection is not always effective, as some S. enterica strains may resist and survive the disinfection process. To date, little is known about the underlying mechanisms of how S. enterica responds to chlorine-based oxidative stress. In this study, we designed a custom bigenome microarray that consists of 385,000 60-mer oligonucleotide probes and targets 4,793 unique gene features in the genomes of S. Enteritidis strain PT4 and S. Typhimurium strain LT2. We explored the transcriptomic responses of both strains to two different chlorine treatments (130 ppm of chlorine for 30 min and 390 ppm of chlorine for 10 min) in brain heart infusion broth. We identified 209 S. enterica core genes associated with Fe-S cluster assembly, cysteine biosynthesis, stress response, ribosome formation, biofilm formation, and energy metabolism that were differentially expressed (>1.5-fold; P < 0.05). In addition, we found that serovars Enteriditis and Typhimurium differed in the responses of 33 stress-related genes and 19 virulence-associated genes to the chlorine stress. Findings from this study suggest that the oxidative-stress response may render S. enterica resistant or susceptible to certain types of environmental stresses, which in turn promotes the development of more effective hurdle interventions to reduce the risk of S. enterica contamination in the food supply. PMID:20562293

  2. Streptomycin Induced Stress Response in Salmonella enterica Serovar Typhimurium Shows Distinct Colony Scatter Signature

    PubMed Central

    Singh, Atul K.; Drolia, Rishi; Bai, Xingjian; Bhunia, Arun K.

    2015-01-01

    We investigated the streptomycin-induced stress response in Salmonella enterica serovars with a laser optical sensor, BARDOT (bacterial rapid detection using optical scattering technology). Initially, the top 20 S. enterica serovars were screened for their response to streptomycin at 100 μg/mL. All, but four S. enterica serovars were resistant to streptomycin. The MIC of streptomycin-sensitive serovars (Enteritidis, Muenchen, Mississippi, and Schwarzengrund) varied from 12.5 to 50 μg/mL, while streptomycin-resistant serovar (Typhimurium) from 125–250 μg/mL. Two streptomycin-sensitive serovars (Enteritidis and Mississippi) were grown on brain heart infusion (BHI) agar plates containing sub-inhibitory concentration of streptomycin (1.25–5 μg/mL) and a streptomycin-resistant serovar (Typhimurium) was grown on BHI containing 25–50 μg/mL of streptomycin and the colonies (1.2 ± 0.1 mm diameter) were scanned using BARDOT. Data show substantial qualitative and quantitative differences in the colony scatter patterns of Salmonella grown in the presence of streptomycin than the colonies grown in absence of antibiotic. Mass-spectrometry identified overexpression of chaperonin GroEL, which possibly contributed to the observed differences in the colony scatter patterns. Quantitative RT-PCR and immunoassay confirmed streptomycin-induced GroEL expression while, aminoglycoside adenylyltransferase (aadA), aminoglycoside efflux pump (aep), multidrug resistance subunit acrA, and ribosomal protein S12 (rpsL), involved in streptomycin resistance, were unaltered. The study highlights suitability of the BARDOT as a non-invasive, label-free tool for investigating stress response in Salmonella in conjunction with the molecular and immunoassay methods. PMID:26252374

  3. Assessment of antibiotic resistance phenotype and integrons in Salmonella enterica serovar Typhimurium isolated from swine.

    PubMed

    Rayamajhi, Nabin; Kang, Sang Gyun; Kang, Mi Lan; Lee, Hee Soo; Park, Kyung Yoon; Yoo, Han Sang

    2008-10-01

    Salmonella enterica serovar Typhimurium (S. Typhimurium) isolated and identified from swine were subjected for the analysis of antibiotic resistance pattern and clinically important class 1 and 2 integrons. In addition, S. Typhimurium isolates exhibiting ampicillin, chloramphenicol, streptomycin, sulfamethoxazole, tetracycline and florfenicol (ACSSuTF) resistance pattern as described in most Salmonella enterica serotype Typhimurium definitive type 104 (DT104) were characterized by polymerase chain reaction. All the isolates were resistant to more than four antibiotics and showed the highest resistance to streptomycin (94.1%), followed by tetracycline (90.1%), ampicillin (64.7%), chloramphenicol (56.8%) and gentamicin (54.9%). MIC value for the ten isolates ranged between 0.125-2 mug/ml for ciprofloxacin. Among the beta-lactams used, only one of the isolate exhibited resistance to ceftiofur (MIC 8 microg/ml). Sixty eight percent of these multi drug resistance (MDR) S. Typhimurium isolates carried clinically important class 1 integron with 1kb (aadA) and/or 2kb (dhfrXII-orfF-aadA2) resistance gene cassettes. This study reports the increasing trend of multi drug resistance (MDR) S. Typhimurium with clinically important class 1 integron in pigs. In addition, emergence of the ACSSuTF-type resistance in S. Typhimurium PT other than DT104 may limit the use of resistance gene markers in its detection methods by PCR.

  4. Differences in the motility phenotype of multidrug-resistant Salmonella enterica serovar Typhimurium exposed to various antibiotics

    USDA-ARS?s Scientific Manuscript database

    Salmonella enterica serovar Typhimurium (S. Typhimurium) is one of the most prevalent foodborne-associated bacteria in humans and livestock, and over 35 per cent of these isolates are resistant to three or more antibiotics. This is a concern as multidrug-resistant (MDR) Salmonella has been associat...

  5. The inositol phosphatase SHIP controls Salmonella enterica serovar Typhimurium infection in vivo.

    PubMed

    Bishop, Jennifer L; Sly, Laura M; Krystal, Gerald; Finlay, B Brett

    2008-07-01

    The SH2 domain-containing inositol 5'-phosphatase, SHIP, negatively regulates various hematopoietic cell functions and is critical for maintaining immune homeostasis. However, whether SHIP plays a role in controlling bacterial infections in vivo remains unknown. Salmonella enterica causes human salmonellosis, a disease that ranges in severity from mild gastroenteritis to severe systemic illness, resulting in significant morbidity and mortality worldwide. The susceptibility of ship(+/+) and ship(-/-) mice and bone marrow-derived macrophages to S. enterica serovar Typhimurium infection was compared. ship(-/-) mice displayed an increased susceptibility to both oral and intraperitoneal serovar Typhimurium infection and had significantly higher bacterial loads in intestinal and systemic sites than ship(+/+) mice, indicating a role for SHIP in the gut-associated and systemic pathogenesis of serovar Typhimurium in vivo. Cytokine analysis of serum from orally infected mice showed that ship(-/-) mice produce lower levels of Th1 cytokines than do ship(+/+) animals at 2 days postinfection, and in vitro analysis of supernatants taken from infected bone marrow-derived macrophages derived to mimic the in vivo ship(-/-) alternatively activated (M2) macrophage phenotype correlated with these data. M2 macrophages were the predominant population in vivo in both oral and intraperitoneal infections, since tissue macrophages within the small intestine and peritoneal macrophages from ship(-/-) mice showed elevated levels of the M2 macrophage markers Ym1 and Arginase 1 compared to ship(+/+) cells. Based on these data, we propose that M2 macrophage skewing in ship(-/-) mice contributes to ineffective clearance of Salmonella in vivo.

  6. Regulation of fucose and 1,2-propanediol utilization by Salmonella enterica serovar Typhimurium.

    PubMed

    Staib, Lena; Fuchs, Thilo M

    2015-01-01

    After ingestion, Salmonella enterica serovar Typhimurium (S. Typhimurium) encounters a densely populated, competitive environment in the gastrointestinal tract. To escape nutrient limitation caused by the intestinal microbiota, this pathogen has acquired specific metabolic traits to use compounds that are not metabolized by the commensal bacteria. For example, the utilization of 1,2-propanediol (1,2-PD), a product of the fermentation of L-fucose, which is present in foods of herbal origin and is also a terminal sugar of gut mucins. Under anaerobic conditions and in the presence of tetrathionate, 1,2-PD can serve as an energy source for S. Typhimurium. Comprehensive database analysis revealed that the 1,2-PD and fucose utilization operons are present in all S. enterica serovars sequenced thus far. The operon, consisting of 21 genes, is expressed as a single polycistronic mRNA. As demonstrated here, 1,2-PD was formed and further used when S. Typhimurium strain 14028 was grown with L-fucose, and the gene fucA encoding L-fuculose-1-phosphate aldolase was required for this growth. Using promoter fusions, we monitored the expression of the propanediol utilization operon that was induced at very low concentrations of 1,2-PD and was inhibited by the presence of D-glucose.

  7. A live attenuated Salmonella enterica serovar Typhimurium vaccine provides cross-protection against Salmonella serovars to reduce disease severity and pathogen transmission

    USDA-ARS?s Scientific Manuscript database

    A live attenuated Salmonella enterica serovar Typhimurium vaccine was developed to confer broad protection against multiple Salmonella serovars to prevent disease and reduce pathogen colonization and shedding. Two vaccine trials were performed in swine to determine the protection afforded by the vac...

  8. Protective immunity conferred by a DNA adenine methylase deficient Salmonella enterica serovar Typhimurium vaccine when delivered in-water to sheep challenged with Salmonella enterica serovar Typhimurium.

    PubMed

    Mohler, V L; Heithoff, D M; Mahan, M J; Walker, K H; Hornitzky, M A; Gabor, L; Thomson, P C; Thompson, A; House, J K

    2011-04-27

    Stimulation of acquired immunity to Salmonella in livestock is not feasible in neonates (which can be infected within 24h of birth) and is challenging in feedlots, which typically source animals from diverse locations and vendors. Induction of innate immune mechanisms through mass vaccination of animals upon arrival to feedlots is an alternative approach. Transport, environmental conditions, changes in social grouping, and further handling during feedlot assembly are significant stressors. These factors, as well as concurrent exposure to a diversity of pathogens, contribute to the risk of disease. We have shown that oral immunization of calves with a modified live Salmonella enterica serovar Typhimurium vaccine strain, which lacks the DNA adenine methylase gene (S. Typhimurium dam), attenuates the severity of clinical disease, reduces fecal shedding, and promotes clearance of salmonellae following virulent homologous and heterologous challenge. This study examines the safety and efficacy of a S. Typhimurium dam vaccine in sheep via oral delivery in drinking water (ad libitum), as a means to effectively vaccinate large groups of animals. Adult merino sheep were vaccinated in drinking water -28 days, -7 days and 24h pre and 24h post-virulent Salmonella Typhimurium challenge which was administered via the oral route. Significant attenuation of clinical disease (temperature, appetite, and attitude) and reduction in mortality and virulent Salmonella Typhimurium fecal shedding and tissue colonization was observed in animals that received the vaccine 28 and 7 days pre-challenge. Further, vaccination did not pose a risk to stock previously infected with virulent salmonellae as mortalities and clinical disease in sheep vaccinated prior to or following virulent challenge did not differ significantly from the non-vaccinated controls. The capacity of S. Typhimurium dam vaccines delivered in drinking water to protect livestock from virulent Salmonella challenge offers an

  9. Innate immune control of Salmonella enterica serovar Typhimurium: mechanisms contributing to combating systemic Salmonella infection.

    PubMed

    Wick, Mary Jo

    2011-01-01

    Infections with Salmonella enterica serovars remain a serious problem worldwide. While serovar Typhi causes significant morbidity and mortality that is restricted to humans, serovar Typhimurium causes gastroenteritidis in humans and can also infect other animals. As mice with the susceptible Nramp1 locus get systemic infection with serovar Typhimurium, murine infection models using this serovar have been widely used to decipher the immune mechanisms required to survive systemic Salmonella infection. This review summarizes recent studies in murine infection models that have advanced our understanding of the events that occur during the first days after oral Salmonella infection. The pathways of bacterial penetration across the intestinal epithelium, bacterial spread to draining (mesenteric) lymph nodes and dissemination to systemic tissues is discussed. The response of myeloid cell populations, including dendritic cells, inflammatory monocytes and neutrophils, during the early stage of infection is also discussed. Finally, the mechanisms driving recruitment of myeloid cells to infected intestinal lymphoid tissues and what is known about Toll-like receptor signaling pathways in innate immunity to Salmonella infection is also discussed.

  10. Interaction of Salmonella enterica Serovar Typhimurium with Intestinal Organoids Derived from Human Induced Pluripotent Stem Cells.

    PubMed

    Forbester, Jessica L; Goulding, David; Vallier, Ludovic; Hannan, Nicholas; Hale, Christine; Pickard, Derek; Mukhopadhyay, Subhankar; Dougan, Gordon

    2015-07-01

    The intestinal mucosa forms the first line of defense against infections mediated by enteric pathogens such as salmonellae. Here we exploited intestinal "organoids" (iHOs) generated from human induced pluripotent stem cells (hIPSCs) to explore the interaction of Salmonella enterica serovar Typhimurium with iHOs. Imaging and RNA sequencing were used to analyze these interactions, and clear changes in transcriptional signatures were detected, including altered patterns of cytokine expression after the exposure of iHOs to bacteria. S. Typhimurium microinjected into the lumen of iHOs was able to invade the epithelial barrier, with many bacteria residing within Salmonella-containing vacuoles. An S. Typhimurium invA mutant defective in the Salmonella pathogenicity island 1 invasion apparatus was less capable of invading the iHO epithelium. Hence, we provide evidence that hIPSC-derived organoids are a promising model of the intestinal epithelium for assessing interactions with enteric pathogens.

  11. Inactivation of Salmonella enterica serovar Typhimurium on fresh produce by cold atmospheric gas plasma technology.

    PubMed

    Fernández, A; Noriega, E; Thompson, A

    2013-02-01

    Cold atmospheric gas plasma treatment (CAP) is an alternative approach for the decontamination of fresh and minimally processed food. In this study, the effects of growth phase, growth temperature and chemical treatment regime on the inactivation of Salmonella enterica serovar Typhimurium (S. Typhimurium) by Nitrogen CAP were examined. Furthermore, the efficacy of CAP treatment for decontaminating lettuce and strawberry surfaces and potato tissue inoculated with S. Typhimurium was evaluated. It was found that the rate of inactivation of S. Typhimurium was independent of the growth phase, growth temperature and chemical treatment regime. Under optimal conditions, a 2 min treatment resulted in a 2.71 log-reduction of S. Typhimurium viability on membrane filters whereas a 15 min treatment was necessary to achieve 2.72, 1.76 and 0.94 log-reductions of viability on lettuce, strawberry and potato, respectively. We suggest that the differing efficiency of CAP treatment on the inactivation of S. Typhimurium on these different types of fresh foods is a consequence of their surface features. Scanning electron microscopy of the surface structures of contaminated samples of lettuce, strawberry and potato revealed topographical features whereby S. Typhimurium cells could be protected from the active species generated by plasma.

  12. Development of triclosan and antibiotic resistance in Salmonella enterica serovar Typhimurium.

    PubMed

    Birosová, Lucia; Mikulásová, Mária

    2009-04-01

    The possible association between the use of triclosan and the development of antibiotic resistance was examined in triclosan-resistant mutants of Salmonella enterica serovar Typhimurium. These mutants were obtained from a sensitive parental strain and from ciprofloxacin-resistant isogenic strains using spontaneous mutagenesis or selection after one short exposure or continuous exposure to low concentrations of triclosan. The results showed that triclosan in the environment does not increase the mutation frequency but selects bacterial strains with reduced antibiotic susceptibility. This property depended on the multiple antibiotic resistance (Mar) phenotype of bacterial strains and on the triclosan concentration.

  13. Genomic Variability of Serial Human Isolates of Salmonella enterica Serovar Typhimurium Associated with Prolonged Carriage.

    PubMed

    Octavia, Sophie; Wang, Qinning; Tanaka, Mark M; Sintchenko, Vitali; Lan, Ruiting

    2015-11-01

    Salmonella enterica serovar Typhimurium is an important foodborne human pathogen that often causes self-limiting but severe gastroenteritis. Prolonged excretion of S. Typhimurium after the infection can lead to secondary transmissions. However, little is known about within-host genomic variation in bacteria associated with asymptomatic shedding. Genomes of 35 longitudinal isolates of S. Typhimurium recovered from 11 patients (children and adults) with culture-confirmed gastroenteritis were sequenced. There were three or four isolates obtained from each patient. Single nucleotide polymorphisms (SNPs) were analyzed in these isolates, which were recovered between 1 and 279 days after the initial diagnosis. Limited genomic variation (5 SNPs or fewer) was associated with short- and long-term carriage of S. Typhimurium. None of the isolates was shown to be due to reinfection. SNPs occurred randomly, and the majority of the SNPs were nonsynonymous. Two nonsense mutations were observed. A nonsense mutation in flhC rendered the isolate nonmotile, whereas the significance of a nonsense mutation in yihV is unknown. The estimated mutation rate is 1.49 × 10(-6) substitution per site per year. S. Typhimurium isolates excreted in stools following acute gastroenteritis in children and adults demonstrated limited genomic variability over time, regardless of the duration of carriage. These findings have important implications for the detection of possible transmission events suspected by public health genomic surveillance of S. Typhimurium infections.

  14. Genomic Analysis of Salmonella enterica Serovar Typhimurium from Wild Passerines in England and Wales.

    PubMed

    Mather, Alison E; Lawson, Becki; de Pinna, Elizabeth; Wigley, Paul; Parkhill, Julian; Thomson, Nicholas R; Page, Andrew J; Holmes, Mark A; Paterson, Gavin K

    2016-11-15

    Passerine salmonellosis is a well-recognized disease of birds in the order Passeriformes, which includes common songbirds such as finches and sparrows, caused by infection with Salmonella enterica serovar Typhimurium. Previous research has suggested that some subtypes of S Typhimurium-definitive phage types (DTs) 40, 56 variant, and 160-are host adapted to passerines and that these birds may represent a reservoir of infection for humans and other animals. Here, we have used the whole-genome sequences of 11 isolates from British passerines, five isolates of similar DTs from humans and a domestic cat, and previously published S Typhimurium genomes that include similar DTs from other hosts to investigate the phylogenetic relatedness of passerine salmonellae to other S Typhimurium isolates and investigate possible genetic features of the distinct disease pathogenesis of S Typhimurium in passerines. Our results demonstrate that the 11 passerine isolates and 13 other isolates, including those from nonpasserine hosts, were genetically closely related, with a median pairwise single nucleotide polymorphism (SNP) difference of 130 SNPs. These 24 isolates did not carry antimicrobial resistance genetic determinants or the S Typhimurium virulence plasmid. Although our study does not provide evidence of Salmonella transmission from passerines to other hosts, our results are consistent with the hypothesis that wild birds represent a potential reservoir of these Salmonella subtypes, and thus, sensible personal hygiene precautions should be taken when feeding or handling garden birds.

  15. Requirement for cobalamin by Salmonella enterica serovars Typhimurium, Pullorum, Gallinarum and Enteritidis during infection in chickens

    PubMed Central

    Vaz, Jacqueline Boldrin; Penha Filho, Rafael Antonio Casarin; Junior, Angelo Berchieri; Lemos, Manoel Victor Franco

    2011-01-01

    Salmonella enterica serovar Typhimurium synthesizes cobalamin (vitamin B12) only during anaerobiosis. Two percent of the S. Typhimurium genome is devoted to the synthesis and uptake of vitamin B12 and to B12-dependent reactions. To understand the requirement for cobalamin synthesis better, we constructed mutants of Salmonella serovars Enteritidis and Pullorum that are double-defective in cobalamin biosynthesis (ΔcobSΔcbiA). We compared the virulence of these mutants to that of their respective wild type strains and found no impairment in their ability to cause disease in chickens. We then assessed B12 production in these mutants and their respective wild type strains, as well as in S. Typhimurium ΔcobSΔcbiA, Salmonella Gallinarum ΔcobSΔcbiA, and their respective wild type strains. None of the mutants was able to produce detectable B12. B12 was detectable in S. Enteritidis, S. Pullorum and S. Typhimurium wild type strains but not in S. Gallinarum. In conclusion, the production of vitamin B12in vitro differed across the tested Salmonella serotypes and the deletion of the cbiA and cobS genes resulted in different levels of alteration in the host parasite interaction according to Salmonella serotype tested. PMID:24031771

  16. Effect of EDTA on Salmonella enterica serovar Typhimurium involves a component not assignable to lipopolysaccharide release.

    PubMed

    Alakomi, H-L; Saarela, M; Helander, I M

    2003-08-01

    The effect of EDTA on Salmonella enterica serovar Typhimurium was studied in different growth phases with cells grown with or without Ca(2+) and Mg(2+) supplementation. EDTA affected the outer membrane much more strongly in the early exponential phase than in the mid- or late exponential phase, as indicated by uptake of 1-N-phenylnaphthylamine (a nonpolar hydrophobic probe, M(r) 219), and detergent (SDS) susceptibility. This effect was, however, not paralleled by LPS release (determined by measuring LPS-specific fatty acids or 14C-labelled LPS in cell-free supernatants, per a standardized cell density), which remained unchanged as a function of the growth curve. The conclusion from these results is that in the early exponential phase the effect of EDTA in S. enterica involves a component that is independent of LPS release.

  17. Interaction between Host Cells and Septicemic Salmonella enterica Serovar Typhimurium Isolates from Pigs ▿

    PubMed Central

    Bergeron, Nadia; Corriveau, Jonathan; Letellier, Ann; Daigle, France; Lessard, Louise; Quessy, Sylvain

    2009-01-01

    Salmonella enterica serovar Typhimurium is an important pathogen in swine and is also a frequently reported zoonotic agent. The objective of this study was to characterize isolates of S. enterica serovar Typhimurium associated with septicemia in swine and to compare them to isolates recovered from clinically healthy pigs. We were particularly interested in comparing the two groups of isolates for their ability to adhere to and invade host cells, to be phagocytized and survive in monocyte cells, to induce apoptosis, and to adhere to intestinal mucus. Their surface properties were also evaluated by interactions with solvents. The isolates recovered from diseased animals were shown to invade intestinal epithelial cell lines at a higher rate (P = 0.003) than isolates from healthy pigs. Septicemic isolates were phagocytized by human monocytes at a higher rate than isolates from healthy pigs (P = 0.009). The mean percentages of phagocytosis were significantly lower for human monocytes than for porcine monocytes (P = 0.02 and P = 0.008, respectively) for isolates from both diseased and healthy animals. Healthy animal isolates were phagocytized more by porcine monocytes at 15 min (P = 0.02) than septicemic isolates. No difference between isolates from septicemic pigs and isolates from healthy pigs was detected for other tested parameters. These results suggest that septicemic isolates have a particular pattern of invasion. PMID:19710281

  18. Salmonella enterica Serovar Typhimurium Exploits Inflammation to Modify Swine Intestinal Microbiota

    PubMed Central

    Drumo, Rosanna; Pesciaroli, Michele; Ruggeri, Jessica; Tarantino, Michela; Chirullo, Barbara; Pistoia, Claudia; Petrucci, Paola; Martinelli, Nicola; Moscati, Livia; Manuali, Elisabetta; Pavone, Silvia; Picciolini, Matteo; Ammendola, Serena; Gabai, Gianfranco; Battistoni, Andrea; Pezzotti, Giovanni; Alborali, Giovanni L.; Napolioni, Valerio; Pasquali, Paolo; Magistrali, Chiara F.

    2016-01-01

    Salmonella enterica serovar Typhimurium is an important zoonotic gastrointestinal pathogen responsible for foodborne disease worldwide. It is a successful enteric pathogen because it has developed virulence strategies allowing it to survive in a highly inflamed intestinal environment exploiting inflammation to overcome colonization resistance provided by intestinal microbiota. In this study, we used piglets featuring an intact microbiota, which naturally develop gastroenteritis, as model for salmonellosis. We compared the effects on the intestinal microbiota induced by a wild type and an attenuated S. Typhimurium in order to evaluate whether the modifications are correlated with the virulence of the strain. This study showed that Salmonella alters microbiota in a virulence-dependent manner. We found that the wild type S. Typhimurium induced inflammation and a reduction of specific protecting microbiota species (SCFA-producing bacteria) normally involved in providing a barrier against pathogens. Both these effects could contribute to impair colonization resistance, increasing the host susceptibility to wild type S. Typhimurium colonization. In contrast, the attenuated S. Typhimurium, which is characterized by a reduced ability to colonize the intestine, and by a very mild inflammatory response, was unable to successfully sustain competition with the microbiota. PMID:26835435

  19. Draft Genome Sequences of Two Strains of Salmonella enterica Serovar Typhimurium Displaying Different Virulence in an Experimental Chicken Model

    PubMed Central

    Blais, Burton; Huang, Hongsheng; Wang, Linru; Elmufti, Mohamed; Allain, Ray; Hazelwood, Jennifer; Grenier, Chris; Amoako, Kingsley; Savic, Mirjana; Fattahi Ghazi, Nashmil

    2017-01-01

    ABSTRACT Salmonella enterica serovar Typhimurium strains 22495 and 22792, obtained from wild birds, were found to display different virulence attributes in an experimental chicken model. Closed genome sequences were assembled after sequencing with the Roche 454 and Illumina MiSeq platforms. An additional plasmid was present in the more virulent strain 22495. PMID:28183752

  20. Correlating Blood Immune Parameters and a CCT7 Genetic Variant with the Shedding of Salmonella enterica Serovar Typhimurium in Swine

    USDA-ARS?s Scientific Manuscript database

    In the current study, 40 crossbred pigs were intranasally inoculated with Salmonella enterica serovar Typhimurium and monitored for Salmonella fecal shedding and blood immune parameters at 2, 7, 14 and 20 days post-inoculation (dpi). Using a multivariate permutation test, a positive correlation was...

  1. Motility revertants of opgGH mutants of Salmonella enterica serovar Typhimurium remain defective in mice virulence

    USDA-ARS?s Scientific Manuscript database

    We recently demonstrated that osmoregulated periplasmic glucans (OPGs) of Salmonella enterica serovar Typhimurium are required for optimal mouse virulence (Bhagwat et al., 2009. Microbiology 155:229-237). However, lack of OPGs also generated pleiotropic phenotypes such as reduced motility and slower...

  2. Tetracycline accelerates the temporally-regulated invasion response in specific isolates of multidrug-resistant Salmonella enterica serovar Typhimurium

    USDA-ARS?s Scientific Manuscript database

    Background Multidrug-resistant (MDR) Salmonella is associated with increased morbidity compared to antibiotic-sensitive strains and is an important health and safety concern in both humans and animals. Salmonella enterica serovar Typhimurium is a prevalent cause of foodborne disease, and a consider...

  3. Molecular profiling: Catecholamine modulation of gene expression in Escherichia coli O157:H7 and Salmonella enterica serovar Typhimurium

    USDA-ARS?s Scientific Manuscript database

    Investigations of Escherichia coli O157:H7 and Salmonella enterica serovar Typhimurium have demonstrated that these bacterial pathogens can respond to the presence of catecholamines including norepinephrine and/or epinephrine in their environment by modulating gene expression and exhibiting various ...

  4. Transcriptomic analysis of swarm motility phenotype of Salmonella enterica serovar Typhimurium mutant defective in periplasmic glucan synthesis

    USDA-ARS?s Scientific Manuscript database

    Movement of food-borne pathogens on moist surfaces enables them to migrate towards more favorable niches and facilitate their survival for extended periods of time. Salmonella enterica serovar Typhimurium mutants defective in OPG synthesis are unable to exhibit motility on moist surfaces (swarming) ...

  5. Inactivation of Salmonella enterica serovar Typhimurium and quality maintenance of cherry tomatoes treated with gaseous essential oils

    USDA-ARS?s Scientific Manuscript database

    The antimicrobial activity of the essential oils (EOs) from cinnamon bark, oregano, mustard and of their major components cinnamaldehyde, carvacrol, and allyl isothiocyanate (AIT) were evaluated as a gaseous treatment to reduce Salmonella enterica serovar Typhimurium in vitro and on tomatoes. In dif...

  6. Global transcriptome and mutagenic analyses of the acid tolerance response of Salmonella enterica serovar Typhimurium.

    PubMed

    Ryan, Daniel; Pati, Niladri Bhusan; Ojha, Urmesh K; Padhi, Chandrashekhar; Ray, Shilpa; Jaiswal, Sangeeta; Singh, Gajinder P; Mannala, Gopala K; Schultze, Tilman; Chakraborty, Trinad; Suar, Mrutyunjay

    2015-12-01

    Salmonella enterica serovar Typhimurium (S. Typhimurium) is one of the leading causative agents of food-borne bacterial gastroenteritis. Swift invasion through the intestinal tract and successful establishment in systemic organs are associated with the adaptability of S. Typhimurium to different stress environments. Low-pH stress serves as one of the first lines of defense in mammalian hosts, which S. Typhimurium must efficiently overcome to establish an infection. Therefore, a better understanding of the molecular mechanisms underlying the adaptability of S. Typhimurium to acid stress is highly relevant. In this study, we have performed a transcriptome analysis of S. Typhimurium under the acid tolerance response (ATR) and found a large number of genes (∼47%) to be differentially expressed (more than 1.5-fold or less than -1.5-fold; P < 0.01). Functional annotation revealed differentially expressed genes to be associated with regulation, metabolism, transport and binding, pathogenesis, and motility. Additionally, our knockout analysis of a subset of differentially regulated genes facilitated the identification of proteins that contribute to S. Typhimurium ATR and virulence. Mutants lacking genes encoding the K(+) binding and transport protein KdpA, hypothetical protein YciG, the flagellar hook cap protein FlgD, and the nitrate reductase subunit NarZ were significantly deficient in their ATRs and displayed varied in vitro virulence characteristics. This study offers greater insight into the transcriptome changes of S. Typhimurium under the ATR and provides a framework for further research on the subject. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  7. Global Genomic Epidemiology of Salmonella enterica Serovar Typhimurium DT104.

    PubMed

    Leekitcharoenphon, Pimlapas; Hendriksen, Rene S; Le Hello, Simon; Weill, François-Xavier; Baggesen, Dorte Lau; Jun, Se-Ran; Ussery, David W; Lund, Ole; Crook, Derrick W; Wilson, Daniel J; Aarestrup, Frank M

    2016-04-01

    It has been 30 years since the initial emergence and subsequent rapid global spread of multidrug-resistant Salmonella entericaserovar Typhimurium DT104 (MDR DT104). Nonetheless, its origin and transmission route have never been revealed. We used whole-genome sequencing (WGS) and temporally structured sequence analysis within a Bayesian framework to reconstruct temporal and spatial phylogenetic trees and estimate the rates of mutation and divergence times of 315S Typhimurium DT104 isolates sampled from 1969 to 2012 from 21 countries on six continents. DT104 was estimated to have emerged initially as antimicrobial susceptible in ∼1948 (95% credible interval [CI], 1934 to 1962) and later became MDR DT104 in ∼1972 (95% CI, 1972 to 1988) through horizontal transfer of the 13-kb Salmonella genomic island 1 (SGI1) MDR region into susceptible strains already containing SGI1. This was followed by multiple transmission events, initially from central Europe and later between several European countries. An independent transmission to the United States and another to Japan occurred, and from there MDR DT104 was probably transmitted to Taiwan and Canada. An independent acquisition of resistance genes took place in Thailand in ∼1975 (95% CI, 1975 to 1990). In Denmark, WGS analysis provided evidence for transmission of the organism between herds of animals. Interestingly, the demographic history of Danish MDR DT104 provided evidence for the success of the program to eradicate Salmonellafrom pig herds in Denmark from 1996 to 2000. The results from this study refute several hypotheses on the evolution of DT104 and suggest that WGS may be useful in monitoring emerging clones and devising strategies for prevention of Salmonella infections.

  8. Mapping the Regulatory Network for Salmonella enterica Serovar Typhimurium Invasion

    PubMed Central

    Smith, Carol; Stringer, Anne M.; Mao, Chunhong; Palumbo, Michael J.

    2016-01-01

    ABSTRACT Salmonella enterica pathogenicity island 1 (SPI-1) encodes proteins required for invasion of gut epithelial cells. The timing of invasion is tightly controlled by a complex regulatory network. The transcription factor (TF) HilD is the master regulator of this process and senses environmental signals associated with invasion. HilD activates transcription of genes within and outside SPI-1, including six other TFs. Thus, the transcriptional program associated with host cell invasion is controlled by at least 7 TFs. However, very few of the regulatory targets are known for these TFs, and the extent of the regulatory network is unclear. In this study, we used complementary genomic approaches to map the direct regulatory targets of all 7 TFs. Our data reveal a highly complex and interconnected network that includes many previously undescribed regulatory targets. Moreover, the network extends well beyond the 7 TFs, due to the inclusion of many additional TFs and noncoding RNAs. By comparing gene expression profiles of regulatory targets for the 7 TFs, we identified many uncharacterized genes that are likely to play direct roles in invasion. We also uncovered cross talk between SPI-1 regulation and other regulatory pathways, which, in turn, identified gene clusters that likely share related functions. Our data are freely available through an intuitive online browser and represent a valuable resource for the bacterial research community. PMID:27601571

  9. Chloramphenicol and tetracycline decrease motility and increase invasion and attachment gene expression in specific isolates of multidrug-resistant Salmonella enterica serovar Typhimurium

    USDA-ARS?s Scientific Manuscript database

    Salmonella enterica serovar Typhimurium (S. Typhimurium) is one of the most common serovars isolated from humans and livestock, and over 35 percent of these isolates are resistant to three or more antibiotics. Multidrug-resistant (MDR) Salmonella is a public health concern as it is associated with i...

  10. Direct ROS Scavenging Activity of CueP from Salmonella enterica serovar Typhimurium

    PubMed Central

    Yoon, Bo-Young; Yeom, Ji-Hyun; Kim, Jin-Sik; Um, Si-Hyeon; Jo, Inseong; Lee, Kangseok; Kim, Yong-Hak; Ha, Nam-Chul

    2014-01-01

    Salmonella enterica serovar Typhimurium (S. Typhimurium) is an intracellular pathogen that has evolved to survive in the phagosome of macrophages. The periplasmic copper-binding protein CueP was initially known to confer copper resistance to S. Typhimurium. Crystal structure and biochemical studies on CueP revealed a putative copper binding site surrounded by the conserved cysteine and histidine residues. A recent study reported that CueP supplies copper ions to periplasmic Cu, Zn-superoxide dismutase (SodCII) at a low copper concentration and thus enables the sustained SodCII activity in the periplasm. In this study, we investigated the role of CueP in copper resistance at a high copper concentration. We observed that the survival of a cueP-deleted strain of Salmonella in macrophage phagosome was significantly reduced. Subsequent biochemical experiments revealed that CueP specifically mediates the reduction of copper ion using electrons released during the formation of the disulfide bond. We observed that the copper ion-mediated Fenton reaction in the presence of hydrogen peroxide was blocked by CueP. This study provides insight into how CueP confers copper resistance to S. Typhimurium in copper-rich environments such as the phagosome of macrophages. PMID:24598994

  11. Outbreak of Salmonella enterica serovar Typhimurium phage type DT41 in Danish poultry production.

    PubMed

    Löfström, Charlotta; Hintzmann, Ann-Sofie; Sørensen, Gitte; Baggesen, Dorte Lau

    2015-07-09

    Salmonella enterica subspecies enterica serovar Typhimurium (S. Typhimurium) is one of the most prevalent serovars in Europe - where both poultry and poultry related products are common sources of human salmonellosis. Due to efficient control programs, the prevalence of S. Typhimurium in Danish poultry production is very low. Despite this, during the past decades there has been a reoccurring problem with infections with S. Typhimurium phage type DT41 in the Danish poultry production without identifying a clear source. In the end of 2013 and beginning of 2014 an increased isolation of S. Typhimurium DT41 was noted mainly in this production, but also in other samples. To investigate this is in more detail, 47 isolates from egg layers (n=5, 1 flock), broilers (n=33, 13 flocks), broiler breeding flocks and hatches (n=5; 2 flocks and 1 environmental hatchery sample), feed (n=1), poultry slaughter house (n=3, environmental sample and meat) were typed with multi locus variable number of tandem repeat analysis (MLVA) and pulsed-field gel electrophoresis (PFGE) to investigate the epidemiology of the outbreak. Based on PFGE results isolates were divided into four groups (Simpson's index of diversity (DI)=0.24±0.15). Due to the low DI, PFGE was not sufficient to provide information to unravel the outbreak. Based on MLVA typing the DT41 (42/47 isolates) and the RDNC isolates (5/47) were split into nine groups (DI=0.65±0.14). When a maximum divergence at one locus was permitted these could be gathered into four groups. Using this criterion, combined with epidemiological information, a spread of one type from broiler breeders to broilers and further to the poultry slaughter house was plausible. In conclusion, although it could be concluded that a spread within the broiler production pyramid had taken place the source of the sudden increase of S. Typhimurium DT41 remains unclear. To investigate this in more detail, further studies using whole genome sequencing to obtain a

  12. An rfaH mutant of Salmonella enterica serovar typhimurium is attenuated in swine and reduces intestinal colonization, fecal shedding, and disease severity due to virulent Salmonella Typhimurium

    USDA-ARS?s Scientific Manuscript database

    Swine are often asymptomatic carriers of Salmonella spp., and interventions are needed to limit colonization of swine to enhance food safety and reduce environmental contamination. We evaluated the attenuation and potential vaccine use in pigs of a Salmonella enterica serovar Typhimurium mutant of r...

  13. Beneficial Effects of Sodium Phenylbutyrate Administration during Infection with Salmonella enterica Serovar Typhimurium

    PubMed Central

    Jellbauer, Stefan; Perez Lopez, Araceli; Behnsen, Judith; Gao, Nina; Nguyen, Thao; Murphy, Clodagh; Edwards, Robert A.

    2016-01-01

    Sodium phenylbutyrate (PBA) is a derivative of the short-chain fatty acid butyrate and is approved for treatment of urea cycle disorders and progressive familial intrahepatic cholestasis type 2. Previously known functions include histone deacetylase inhibitor, endoplasmic reticulum stress inhibitor, ammonia sink, and chemical chaperone. Here, we show that PBA has a previously undiscovered protective role in host mucosal defense during infection. Administration of PBA to Taconic mice resulted in the increase of intestinal Lactobacillales and segmented filamentous bacteria (SFB), as well as an increase of interleukin 17 (IL-17) production by intestinal cells. This effect was not observed in Jackson Laboratory mice, which are not colonized with SFB. Because previous studies showed that IL-17 plays a protective role during infection with mucosal pathogens, we hypothesized that Taconic mice treated with PBA would be more resistant to infection with Salmonella enterica serovar Typhimurium (S. Typhimurium). By using the streptomycin-treated mouse model, we found that Taconic mice treated with PBA exhibited significantly lower S. Typhimurium intestinal colonization and dissemination to the reticuloendothelial system, as well as lower levels of inflammation. The lower levels of S. Typhimurium gut colonization and intestinal inflammation were not observed in Jackson Laboratory mice. Although PBA had no direct effect on bacterial replication, its administration reduced S. Typhimurium epithelial cell invasion and lowered the induction of the proinflammatory cytokine IL-23 in macrophage-like cells. These effects likely contributed to the better outcome of infection in PBA-treated mice. Overall, our results suggest that PBA induces changes in the microbiota and in the mucosal immune response that can be beneficial to the host during infection with S. Typhimurium and possibly other enteric pathogens. PMID:27382022

  14. Receptor Diversity and Host Interaction of Bacteriophages Infecting Salmonella enterica Serovar Typhimurium

    PubMed Central

    Kim, Hyeryen; Choi, Younho; Heu, Sunggi; Ryu, Sangryeol

    2012-01-01

    Background Salmonella enterica subspecies enterica serovar Typhimurium is a Gram-negative pathogen causing salmonellosis. Salmonella Typhimurium-targeting bacteriophages have been proposed as an alternative biocontrol agent to antibiotics. To further understand infection and interaction mechanisms between the host strains and the bacteriophages, the receptor diversity of these phages needs to be elucidated. Methodology/Principal Findings Twenty-five Salmonella phages were isolated and their receptors were identified by screening a Tn5 random mutant library of S. Typhimurium SL1344. Among them, three types of receptors were identified flagella (11 phages), vitamin B12 uptake outer membrane protein, BtuB (7 phages) and lipopolysaccharide-related O-antigen (7 phages). TEM observation revealed that the phages using flagella (group F) or BtuB (group B) as a receptor belong to Siphoviridae family, and the phages using O-antigen of LPS as a receptor (group L) belong to Podoviridae family. Interestingly, while some of group F phages (F-I) target FliC host receptor, others (F-II) target both FliC and FljB receptors, suggesting that two subgroups are present in group F phages. Cross-resistance assay of group B and L revealed that group L phages could not infect group B phage-resistant strains and reversely group B phages could not infect group L SPN9TCW-resistant strain. Conclusions/Significance In this report, three receptor groups of 25 newly isolated S. Typhimurium-targeting phages were determined. Among them, two subgroups of group F phages interact with their host receptors in different manner. In addition, the host receptors of group B or group L SPN9TCW phages hinder other group phage infection, probably due to interaction between receptors of their groups. This study provides novel insights into phage-host receptor interaction for Salmonella phages and will inform development of optimal phage therapy for protection against Salmonella. PMID:22927964

  15. Characterization of Small ColE1-Like Plasmids Conferring Kanamycin Resistance in Salmonella enterica subsp. enterica serovars Typhimurium and Newport

    USDA-ARS?s Scientific Manuscript database

    Multi-antibiotic resistant (MR) Salmonella enterica serovars Typhimurium and Newport are an increasing concern in human and animal health. Many strains are known to carry antibiotic resistance determinants on multiple plasmids, yet detailed information is scarce. Three plasmids conferring kanamycin...

  16. Salmonella enterica serovar Typhimurium ΔmsbB Triggers Exacerbated Inflammation in Nod2 Deficient Mice

    PubMed Central

    Claes, Anne-Kathrin; Steck, Natalie; Schultz, Dorothee; Zähringer, Ulrich; Lipinski, Simone; Rosenstiel, Philip; Geddes, Kaoru; Philpott, Dana J.; Heine, Holger; Grassl, Guntram A.

    2014-01-01

    The intracellular pathogen Salmonella enterica serovar Typhimurium causes intestinal inflammation characterized by edema, neutrophil influx and increased pro-inflammatory cytokine expression. A major bacterial factor inducing pro-inflammatory host responses is lipopolysaccharide (LPS). S. Typhimurium ΔmsbB possesses a modified lipid A, has reduced virulence in mice, and is being considered as a potential anti-cancer vaccine strain. The lack of a late myristoyl transferase, encoded by MsbB leads to attenuated TLR4 stimulation. However, whether other host receptor pathways are also altered remains unclear. Nod1 and Nod2 are cytosolic pattern recognition receptors recognizing bacterial peptidoglycan. They play important roles in the host's immune response to enteric pathogens and in immune homeostasis. Here, we investigated how deletion of msbB affects Salmonella's interaction with Nod1 and Nod2. S. Typhimurium Δ msbB-induced inflammation was significantly exacerbated in Nod2−/− mice compared to C57Bl/6 mice. In addition, S. Typhimurium ΔmsbB maintained robust intestinal colonization in Nod2−/− mice from day 2 to day 7 p.i., whereas colonization levels significantly decreased in C57Bl/6 mice during this time. Similarly, infection of Nod1−/− and Nod1/Nod2 double-knockout mice revealed that both Nod1 and Nod2 play a protective role in S. Typhimurium ΔmsbB-induced colitis. To elucidate why S. Typhimurium ΔmsbB, but not wild-type S. Typhimurium, induced an exacerbated inflammatory response in Nod2−/− mice, we used HEK293 cells which were transiently transfected with pathogen recognition receptors. Stimulation of TLR2-transfected cells with S. Typhimurium ΔmsbB resulted in increased IL-8 production compared to wild-type S. Typhimurium. Our results indicate that S. Typhimurium ΔmsbB triggers exacerbated colitis in the absence of Nod1 and/or Nod2, which is likely due to increased TLR2 stimulation. How bacteria with “genetically detoxified” LPS

  17. Microgravity as a novel environmental signal affecting Salmonella enterica serovar Typhimurium virulence

    NASA Technical Reports Server (NTRS)

    Nickerson, C. A.; Ott, C. M.; Mister, S. J.; Morrow, B. J.; Burns-Keliher, L.; Pierson, D. L.

    2000-01-01

    The effects of spaceflight on the infectious disease process have only been studied at the level of the host immune response and indicate a blunting of the immune mechanism in humans and animals. Accordingly, it is necessary to assess potential changes in microbial virulence associated with spaceflight which may impact the probability of in-flight infectious disease. In this study, we investigated the effect of altered gravitational vectors on Salmonella virulence in mice. Salmonella enterica serovar Typhimurium grown under modeled microgravity (MMG) were more virulent and were recovered in higher numbers from the murine spleen and liver following oral infection compared to organisms grown under normal gravity. Furthermore, MMG-grown salmonellae were more resistant to acid stress and macrophage killing and exhibited significant differences in protein synthesis than did normal-gravity-grown cells. Our results indicate that the environment created by simulated microgravity represents a novel environmental regulatory factor of Salmonella virulence.

  18. The wetting agent required for swarming in Salmonella enterica serovar typhimurium is not a surfactant.

    PubMed

    Chen, Bryan G; Turner, Linda; Berg, Howard C

    2007-12-01

    We compared the abilities of media from agar plates surrounding swarming and nonswarming cells of Salmonella enterica serovar Typhimurium to wet a nonpolar surface by measuring the contact angles of small drops. The swarming cells were wild type for chemotaxis, and the nonswarming cells were nonchemotactic mutants with motor biases that were counterclockwise (cheY) or clockwise (cheZ). The latter strains have been shown to be defective for swarming because the agar remains dry (Q. Wang, A. Suzuki, S. Mariconda, S. Porwollik, and R. M. Harshey, EMBO J. 24:2034-2042, 2005). We found no differences in the abilities of the media surrounding these cells, either wild type or mutant, to wet a low-energy surface (freshly prepared polydimethylsiloxane); although, their contact angles were smaller than that of the medium harvested from the underlying agar. So the agent that promotes wetness produced by wild-type cells is not a surfactant; it is an osmotic agent.

  19. Microgravity as a novel environmental signal affecting Salmonella enterica serovar Typhimurium virulence.

    PubMed

    Nickerson, C A; Ott, C M; Mister, S J; Morrow, B J; Burns-Keliher, L; Pierson, D L

    2000-06-01

    The effects of spaceflight on the infectious disease process have only been studied at the level of the host immune response and indicate a blunting of the immune mechanism in humans and animals. Accordingly, it is necessary to assess potential changes in microbial virulence associated with spaceflight which may impact the probability of in-flight infectious disease. In this study, we investigated the effect of altered gravitational vectors on Salmonella virulence in mice. Salmonella enterica serovar Typhimurium grown under modeled microgravity (MMG) were more virulent and were recovered in higher numbers from the murine spleen and liver following oral infection compared to organisms grown under normal gravity. Furthermore, MMG-grown salmonellae were more resistant to acid stress and macrophage killing and exhibited significant differences in protein synthesis than did normal-gravity-grown cells. Our results indicate that the environment created by simulated microgravity represents a novel environmental regulatory factor of Salmonella virulence.

  20. Complete genome sequence of Salmonella enterica serovar Typhimurium VNP20009, a strain engineered for tumor targeting.

    PubMed

    Broadway, Katherine M; Modise, Thero; Jensen, Roderick V; Scharf, Birgit E

    2014-12-20

    A mutagenized and genetically modified derivative of Salmonella enterica serovar Typhimurium 14028S, VNP20009 (ATCC 202165), is attenuated in host virulence and accumulates preferentially in tumors. Here, we report the complete genome of this anticancer therapy agent consisting of one chromosome and one virulence plasmid. The major genetic features that distinguish VNP20009 from its parental strain are: an engineered msbB deletion, a 3'-extension in pykA, a Tn10-caused 16.6-kbp inversion leading to the disruption of purM, a 108-kbp Suwwan deletion resulting in the loss of 128 genes, and 50 non-synonymous SNPs. Copyright © 2014 Elsevier B.V. All rights reserved.

  1. Microgravity as a Novel Environmental Signal Affecting Salmonella enterica Serovar Typhimurium Virulence

    PubMed Central

    Nickerson, Cheryl A.; Ott, C. Mark; Mister, Sarah J.; Morrow, Brian J.; Burns-Keliher, Lisa; Pierson, Duane L.

    2000-01-01

    The effects of spaceflight on the infectious disease process have only been studied at the level of the host immune response and indicate a blunting of the immune mechanism in humans and animals. Accordingly, it is necessary to assess potential changes in microbial virulence associated with spaceflight which may impact the probability of in-flight infectious disease. In this study, we investigated the effect of altered gravitational vectors on Salmonella virulence in mice. Salmonella enterica serovar Typhimurium grown under modeled microgravity (MMG) were more virulent and were recovered in higher numbers from the murine spleen and liver following oral infection compared to organisms grown under normal gravity. Furthermore, MMG-grown salmonellae were more resistant to acid stress and macrophage killing and exhibited significant differences in protein synthesis than did normal-gravity-grown cells. Our results indicate that the environment created by simulated microgravity represents a novel environmental regulatory factor of Salmonella virulence. PMID:10816456

  2. Genome Sequence of Bacteriophage GG32, Which Can Infect both Salmonella enterica Serovar Typhimurium and Escherichia coli O157:H7.

    PubMed

    Chae, Su-Jin; Kwon, Taesoo; Lee, Sunjin; Kang, Yeon Ho; Chung, Gyung Tae; Kim, Dae-Won; Lee, Deog-Yong

    2016-12-08

    We report here a new virulent Salmonella enterica serovar Typhimurium (S Typhimurium) bacteriophage, GG32, which was isolated from the Guem River in the Republic of Korea. The strain can infect both S Typhimurium and Escherichia coli (E. coli) O157:H7 and may be a good candidate for a bio-control agent. Copyright © 2016 Chae et al.

  3. Genome Sequence of Bacteriophage GG32, Which Can Infect both Salmonella enterica Serovar Typhimurium and Escherichia coli O157:H7

    PubMed Central

    Chae, Su-Jin; Kwon, Taesoo; Lee, Sunjin; Kang, Yeon Ho; Chung, Gyung Tae

    2016-01-01

    We report here a new virulent Salmonella enterica serovar Typhimurium (S. Typhimurium) bacteriophage, GG32, which was isolated from the Guem River in the Republic of Korea. The strain can infect both S. Typhimurium and Escherichia coli (E. coli) O157:H7 and may be a good candidate for a bio-control agent. PMID:27932635

  4. Swarm motility of Salmonella enterica serovar Typhimurium is inhibited by compounds from fruit peel extracts.

    PubMed

    Mahadwar, G; Chauhan, K R; Bhagavathy, G V; Murphy, C; Smith, A D; Bhagwat, A A

    2015-04-01

    Controlling spread of human pathogens on fresh produce is a top priority for public health reasons. Isolation of compounds from agricultural waste that would control spread of human pathogens was explored using Salmonella enterica serovar Typhimurium as a model organism. In the environment, micro-organisms migrate as a 'community' especially when they move on moist surfaces. This type of motility is characterized as swarming motility. We examined extracts from agricultural waste such as soya bean husk, peels of orange, pineapple, avocado and pomegranate for antiswarming activity. Avocado and pineapple peels showed moderate (~40%) inhibition of swarming motility while pomegranate peel extract had high antiswarming activity (~85% inhibition) and was examined in further detail. Although the pomegranate peel extract was acidic, swarm-inhibitory activity was not due to low pH and the peel extract did not inhibit growth of Salmonella. Among the key swarm motility regulatory genes, class II (fliF, fliA, fliT and fliZ) and class III (fliC and fliM) regulators were downregulated upon exposure to pomegranate peel extract. Pomegranate peels offer great potential as a bioactive repellent for pathogenic micro-organisms on moist surfaces. Controlling the spread of food-borne pathogens in moist environments is an important microbial food safety issue. Isolation of compounds from agricultural waste (such as fruit peels) that would control spread of human pathogens was explored using Salmonella enterica serovar Typhimurium as a model organism. Pomegranate peels offer great potential as a bioactive repellent for pathogenic micro-organisms. © 2014 The Society for Applied Microbiology.

  5. Curcumin increases the pathogenicity of Salmonella enterica serovar Typhimurium in murine model.

    PubMed

    Marathe, Sandhya A; Ray, Seemun; Chakravortty, Dipshikha

    2010-07-09

    Curcumin has gained immense importance for its vast therapeutic and prophylactic applications. Contrary to this, our study reveals that it regulates the defense pathways of Salmonella enterica serovar Typhimurium (S. Typhimurium) to enhance its pathogenicity. In a murine model of typhoid fever, we observed higher bacterial load in Peyer's patches, mesenteric lymph node, spleen and liver, when infected with curcumin-treated Salmonella. Curcumin increased the resistance of S. Typhimurium against antimicrobial agents like antimicrobial peptides, reactive oxygen and nitrogen species. This increased tolerance might be attributed to the up-regulation of genes involved in resistance against antimicrobial peptides--pmrD and pmrHFIJKLM and genes with antioxidant function--mntH, sodA and sitA. We implicate that iron chelation property of curcumin have a role in regulating mntH and sitA. Interestingly, we see that the curcumin-mediated modulation of pmr genes is through the PhoPQ regulatory system. Curcumin downregulates SPI1 genes, required for entry into epithelial cells and upregulates SPI2 genes required to intracellular survival. Since it is known that the SPI1 and SPI2 system can be regulated by the PhoPQ system, this common regulator could explain curcumin's mode of action. This data urges us to rethink the indiscriminate use of curcumin especially during Salmonella outbreaks.

  6. Structural and Functional Characterization of Three DsbA Paralogues from Salmonella enterica Serovar Typhimurium*

    PubMed Central

    Heras, Begoña; Totsika, Makrina; Jarrott, Russell; Shouldice, Stephen R.; Gunčar, Gregor; Achard, Maud E. S.; Wells, Timothy J.; Argente, M. Pilar; McEwan, Alastair G.; Schembri, Mark A.

    2010-01-01

    In prototypic Escherichia coli K-12 the introduction of disulfide bonds into folding proteins is mediated by the Dsb family of enzymes, primarily through the actions of the highly oxidizing protein EcDsbA. Homologues of the Dsb catalysts are found in most bacteria. Interestingly, pathogens have developed distinct Dsb machineries that play a pivotal role in the biogenesis of virulence factors, hence contributing to their pathogenicity. Salmonella enterica serovar (sv.) Typhimurium encodes an extended number of sulfhydryl oxidases, namely SeDsbA, SeDsbL, and SeSrgA. Here we report a comprehensive analysis of the sv. Typhimurium thiol oxidative system through the structural and functional characterization of the three Salmonella DsbA paralogues. The three proteins share low sequence identity, which results in several unique three-dimensional characteristics, principally in areas involved in substrate binding and disulfide catalysis. Furthermore, the Salmonella DsbA-like proteins also have different redox properties. Whereas functional characterization revealed some degree of redundancy, the properties of SeDsbA, SeDsbL, and SeSrgA and their expression pattern in sv. Typhimurium indicate a diverse role for these enzymes in virulence. PMID:20233716

  7. Identification of HilD-Regulated Genes in Salmonella enterica Serovar Typhimurium

    PubMed Central

    Petrone, Brianna L.; Stringer, Anne M.

    2014-01-01

    Salmonella enterica serovar Typhimurium (S. Typhimurium) pathogenicity island 1 (SPI-1) encodes a type III secretion system required for invasion of host gut epithelial cells. Expression of SPI-1 virulence genes is controlled by a complex hierarchy of transcription factors encoded within and outside SPI-1. The master regulator of SPI-1, HilA, is itself regulated by three homologous transcription factors, HilD, HilC, and RtsA. HilD activates transcription of hilA and other target genes in response to environmental conditions associated with the intestinal microenvironment of the host. We have mapped the binding of HilD across the S. Typhimurium genome using chromatin immunoprecipitation-sequencing (ChIP-seq). Thus, we have identified 17 regions bound by HilD, including 11 novel targets. The majority of HilD targets are located outside SPI-1. We demonstrate transcription activation of 8 genes by HilD; four of these genes have not been previously described as being regulated by HilD, including lpxR, which encodes a lipid A deacylase important for immune evasion. We also show that HilD-activated genes are frequently activated by HilC and RtsA, indicating extensive overlap of the HilD, HilC, and RtsA regulons. PMID:24375101

  8. Emergence of Clinical Salmonella enterica Serovar Typhimurium Isolates with Concurrent Resistance to Ciprofloxacin, Ceftriaxone, and Azithromycin

    PubMed Central

    Wong, Marcus Ho Yin; Yan, Meiying; Chan, Edward Wai Chi; Biao, Kan

    2014-01-01

    Salmonella infection is an important public health issue for which the needs of antimicrobial treatment are increasing. A total of 546 human clinical S. enterica serovar Typhimurium isolates were recovered from patients in hospitals in China during the period of 2005 to ∼2011. Twenty percent of the isolates exhibited resistance to ciprofloxacin, and 4% were resistant to ceftriaxone. Importantly, for the first time, 12 (2%) S. Typhimurium isolates resistant to both ciprofloxacin and ceftriaxone were recovered; among these 12 isolates, two were also resistant to azithromycin, and one was resistant to all other drugs tested. The combined effects of various transferrable extended-spectrum β-lactamase determinants and a novel efflux-based ciprofloxacin resistance mechanism encoded by the mobile efflux gene oqxAB were responsible for the emergence of these extremely (highly) drug-resistant (XDR) S. Typhimurium isolates. The dissemination of resistance genes, such as those encoding ESBLs and the OqxAB pump, among Salmonella organisms will speed up the selection of XDR Salmonella, posing a huge threat to public health and Salmonella infection control. PMID:24752251

  9. Oxidoreductases that Act as Conditional Virulence Suppressors in Salmonella enterica Serovar Typhimurium

    PubMed Central

    Anwar, Naeem; Sem, Xiao Hui; Rhen, Mikael

    2013-01-01

    In Salmonella enterica serovar Typhimurium, oxidoreductases of the thioredoxin superfamily contribute to bacterial invasiveness, intracellular replication and to the virulence in BALB/c mice as well as in the soil nematode Caenorhabditis elegans. The scsABCD gene cluster, present in many but not all enteric bacteria, codes for four putative oxidoreductases of the thioredoxin superfamily. Here we have analyzed the potential role of the scs genes in oxidative stress tolerance and virulence in S. Typhimurium. An scsABCD deletion mutant showed moderate sensitization to the redox-active transition metal ion copper and increased protein carbonylation upon exposure to hydrogen peroxide. Still, the scsABCD mutant was not significantly affected for invasiveness or intracellular replication in respectively cultured epithelial or macrophage-like cells. However, we noted a significant copper chloride sensitivity of SPI1 T3SS mediated invasiveness that strongly depended on the presence of the scs genes. The scsABCD deletion mutant was not attenuated in animal infection models. In contrast, the mutant showed a moderate increase in its competitive index upon intraperitoneal challenge and enhanced invasiveness in small intestinal ileal loops of BALB/c mice. Moreover, deletion of the scsABCD genes restored the invasiveness of a trxA mutant in epithelial cells and its virulence in C. elegans. Our findings thus demonstrate that the scs gene cluster conditionally affects virulence and underscore the complex interactions between oxidoreductases of the thioredoxin superfamily in maintaining host adaptation of S. Typhimurium. PMID:23750221

  10. Curcumin Increases the Pathogenicity of Salmonella enterica Serovar Typhimurium in Murine Model

    PubMed Central

    Marathe, Sandhya A.; Ray, Seemun; Chakravortty, Dipshikha

    2010-01-01

    Curcumin has gained immense importance for its vast therapeutic and prophylactic applications. Contrary to this, our study reveals that it regulates the defense pathways of Salmonella enterica serovar Typhimurium (S. Typhimurium) to enhance its pathogenicity. In a murine model of typhoid fever, we observed higher bacterial load in Peyer's patches, mesenteric lymph node, spleen and liver, when infected with curcumin-treated Salmonella. Curcumin increased the resistance of S. Typhimurium against antimicrobial agents like antimicrobial peptides, reactive oxygen and nitrogen species. This increased tolerance might be attributed to the up-regulation of genes involved in resistance against antimicrobial peptides - pmrD and pmrHFIJKLM and genes with antioxidant function - mntH, sodA and sitA. We implicate that iron chelation property of curcumin have a role in regulating mntH and sitA. Interestingly, we see that the curcumin-mediated modulation of pmr genes is through the PhoPQ regulatory system. Curcumin downregulates SPI1 genes, required for entry into epithelial cells and upregulates SPI2 genes required to intracellular survival. Since it is known that the SPI1 and SPI2 system can be regulated by the PhoPQ system, this common regulator could explain curcumin's mode of action. This data urges us to rethink the indiscriminate use of curcumin especially during Salmonella outbreaks. PMID:20634977

  11. Biofilm formation ability of Salmonella enterica serovar Typhimurium acrAB mutants.

    PubMed

    Schlisselberg, Dov B; Kler, Edna; Kisluk, Guy; Shachar, Dina; Yaron, Sima

    2015-10-01

    Recent studies offer contradictory findings about the role of multidrug efflux pumps in bacterial biofilm development. Thus, the aim of this study was to investigate the involvement of the AcrAB efflux pump in biofilm formation by investigating the ability of AcrB and AcrAB null mutants of Salmonella enterica serovar Typhimurium to produce biofilms. Three models were used to compare the ability of S. Typhimurium wild-type and its mutants to form biofilms: formation of biofilm on polystyrene surfaces; production of biofilm (mat model) on the air/liquid interface; and expression of curli and cellulose on Congo red-supplemented agar plates. All three investigated genotypes formed biofilms with similar characteristics. However, upon exposure to chloramphenicol, formation of biofilms on solid surfaces as well as the production of curli were either reduced or were delayed more significantly in both mutants, whilst there was no visible effect on pellicle formation. It can be concluded that when no selective pressure is applied, S. Typhimurium is able to produce biofilms even when the AcrAB efflux pumps are inactivated, implying that the use of efflux pump inhibitors to prevent biofilm formation is not a general solution and that combined treatments might be more efficient. Other factors that affect the ability to produce biofilms depending on efflux pump activity are yet to be identified.

  12. Natural surface coating to inactivate Salmonella enterica serovar Typhimurium and maintain quality of cherry tomatoes.

    PubMed

    Yun, Juan; Fan, Xuetong; Li, Xihong; Jin, Tony Z; Jia, Xiaoyu; Mattheis, James P

    2015-01-16

    The objective of the present study was to investigate the effectiveness of zein-based coatings in reducing populations of Salmonella enterica serovar Typhimurium and preserving quality of cherry tomatoes. Tomatoes were inoculated with a cocktail of S. Typhimurium LT2 plus three attenuated strains on the smooth skin surface and stem scar area. The zein-based coatings with and without cinnamon (up to 20%) and mustard essential oil or a commercial wax formulation were applied onto tomatoes and the treated fruits were stored at 10 °C for up to 3 weeks. Populations of S. Typhimurium decreased with increased essential oil concentration and storage duration. S. Typhimurium populations on the smooth skin surface were reduced by 4.6 and 2.8 log colony forming units(CFU)/g by the zein coatings with 20% cinnamon and 20% mustard oil, respectively, 5h after coating. The same coating reduced populations of S. Typhimurium to levels below detection limit (1.0 log CFU/g) on the stem scar area of tomato during 7 days of storage at 10 °C. Salmonella populations were not reduced on fruit coated with the commercial wax. All of the coatings resulted in reduced weight loss compared with uncoated control. Compared with the control, loss of firmness and ascorbic acid during storage was prevented by all of the coatings except the zein coating with 20% mustard oil which enhanced softening. Color was not consistently affected by any of the coating treatments during 21 days of storage at 10°C. The results suggest that the zein-based coating containing cinnamon oil might be used to enhance microbial safety and quality of tomato.

  13. Salmonella enterica serovar Typhimurium RamA, intracellular oxidative stress response, and bacterial virulence.

    PubMed

    van der Straaten, Tahar; Zulianello, Laurence; van Diepen, Angela; Granger, Donald L; Janssen, Riny; van Dissel, Jaap T

    2004-02-01

    Escherichia coli and Salmonella enterica serovar Typhimurium have evolved genetic systems, such as the soxR/S and marA regulons, to detoxify reactive oxygen species, like superoxide, which are formed as by-products of metabolism. Superoxide also serves as a microbicidal effector mechanism of the host's phagocytes. Here, we investigate whether regulatory genes other than soxR/S and marA are active in response to oxidative stress in Salmonella and may function as virulence determinants. We identified a bacterial gene, which was designated ramA (342 bp) and mapped at 13.1 min on the Salmonella chromosome, that, when overexpressed on a plasmid in E. coli or Salmonella, confers a pleiotropic phenotype characterized by increased resistance to the redox-cycling agent menadione and to multiple unrelated antibiotics. The ramA gene is present in Salmonella serovars but is absent in E. coli. The gene product displays 37 to 52% homology to the transcriptional activators soxR/S and marA and 80 to 100% identity to a multidrug resistance gene in Klebsiella pneumoniae and Salmonella enterica serovar Paratyphi A. Although a ramA soxR/S double null mutant is highly susceptible to intracellular superoxide generated by menadione and displays decreased Mn-superoxide dismutase activity, intracellular survival of this mutant within macrophage-like RAW 264.7 cells and in vivo replication in the spleens in Ityr mice are not affected. We concluded that despite its role in the protective response of the bacteria to oxidative stress in vitro, the newly identified ramA gene, together with soxR/S, does not play a role in initial replication of Salmonella in the organs of mice.

  14. Heat tolerance of Salmonella enterica serovars Agona, Enteritidis, and Typhimurium in peanut butter.

    PubMed

    Shachar, Dina; Yaron, Sima

    2006-11-01

    Recent large foodborne outbreaks caused by Salmonella enterica serovars have been associated with consumption of foods with high fat content and reduced water activity, even though their ingredients usually undergo pasteurization. The present study was focused on the heat tolerance of Salmonella enterica serovars Agona, Enteritidis, and Typhimurium in peanut butter. The Salmonella serovars in the peanut butter were resistant to heat, and even at a temperature as high as 90 degrees C only 3.2-log reduction in CFU was observed. The obtained thermal inactivation curves were upwardly concave, indicating rapid death at the beginning (10 min) followed by lower death rates and an asymptotic tail. The curves fitted the nonlinear Weibull model with beta parameters < 1, indicating that the remaining cells have a lower probability of dying. beta at 70 degrees C (0.40 +/- 0.04) was significantly lower than beta at 80 degrees C (0.73 +/- 0.19) and 90 degrees C (0.69 +/- 0.17). Very little decrease in the viable population (less than 2-log decrease) was noted in cultures that were exposed to a second thermal treatment. Peanut butter is a highly concentrated colloidal suspension of lipid and water in a peanut meal phase. We hypothesized that differences in the local environments of the bacteria, with respect to fat content or water activity, explained the observed distribution and high portion of surviving cells (0.1%, independent of the initial cell number). These results demonstrate that thermal treatments are inadequate to consistently destroy Salmonella in highly contaminated peanut butter and that the pasteurization process cannot be improved significantly by longer treatment or higher temperatures.

  15. Genomic Analysis of Salmonella enterica Serovar Typhimurium from Wild Passerines in England and Wales

    PubMed Central

    Mather, Alison E.; Lawson, Becki; de Pinna, Elizabeth; Wigley, Paul; Thomson, Nicholas R.; Page, Andrew J.; Holmes, Mark A.

    2016-01-01

    ABSTRACT Passerine salmonellosis is a well-recognized disease of birds in the order Passeriformes, which includes common songbirds such as finches and sparrows, caused by infection with Salmonella enterica serovar Typhimurium. Previous research has suggested that some subtypes of S. Typhimurium—definitive phage types (DTs) 40, 56 variant, and 160—are host adapted to passerines and that these birds may represent a reservoir of infection for humans and other animals. Here, we have used the whole-genome sequences of 11 isolates from British passerines, five isolates of similar DTs from humans and a domestic cat, and previously published S. Typhimurium genomes that include similar DTs from other hosts to investigate the phylogenetic relatedness of passerine salmonellae to other S. Typhimurium isolates and investigate possible genetic features of the distinct disease pathogenesis of S. Typhimurium in passerines. Our results demonstrate that the 11 passerine isolates and 13 other isolates, including those from nonpasserine hosts, were genetically closely related, with a median pairwise single nucleotide polymorphism (SNP) difference of 130 SNPs. These 24 isolates did not carry antimicrobial resistance genetic determinants or the S. Typhimurium virulence plasmid. Although our study does not provide evidence of Salmonella transmission from passerines to other hosts, our results are consistent with the hypothesis that wild birds represent a potential reservoir of these Salmonella subtypes, and thus, sensible personal hygiene precautions should be taken when feeding or handling garden birds. IMPORTANCE Passerine salmonellosis, caused by certain definitive phage types (DTs) of Salmonella Typhimurium, has been documented as a cause of wild passerine mortality since the 1950s in many countries, often in the vicinity of garden bird feeding stations. To gain better insight into its epidemiology and host-pathogen interactions, we sequenced the genomes of a collection of 11

  16. Molecular characterization of Salmonella enterica subsp. enterica serovar Typhimurium DT1 isolates.

    PubMed Central

    Lindqvist, N.; Heinikainen, S.; Siitonen, A.; Pelkonen, S.

    2004-01-01

    Salmonella Typhimurium DT1 is endemic to Finland and has caused human outbreaks since the 1960s. Domestic DT1 isolates (n=235) from 1972 to 1999 from human cases, animals and other sources, as well as foreign DT1 isolates from human cases (n=20) were analysed by molecular methods. Pulsed-field gel electrophoresis (PFGE) yielded 38 XbaI profiles. Of these, XbaI profile 10 was seen in 49% (125/255) of the isolates. Twelve IS200 profiles were obtained; the most common IS200 profile D was seen in 64% (33/52) of the isolates. Two clusters were formed by compilation of the XhaI-, BlnI- and SpeI-PFGE and IS200 profiles and possession of the serovar-specific virulence plasmid. The major cluster contained eight IS200 profiles, including IS200 profile D and XhaI profile 10, and had no virulence plasmid, and can be regarded as typical of the endemic Typhimurium DT1 infection. PMID:15061501

  17. Characterization of a Monoclonal Antibody Directed against Salmonella enterica Serovar Typhimurium and Serovar [4,5,12:i:−] ▿

    PubMed Central

    Rementeria, A.; Vivanco, A. B.; Ramirez, A.; Hernando, F. L.; Bikandi, J.; Herrera-León, S.; Echeita, A.; Garaizar, J.

    2009-01-01

    Flagellar extracts of Salmonella enterica serovars expressing phase 2 H1 antigenic complex (H:1,2, H:1,5, H:1,6, and H:1,7) and a mutant flagellin obtained by site-directed mutagenesis of the fljB gene from serovar Typhimurium at codon 218, transforming threonine to alanine, expressed in Escherichia coli (fljB218A) were used to analyze the H1 antigenic complex. Cross-reactions were detected by Western blotting and dot blotting using commercial polyclonal antibodies against the different wild-type extracts and mutant FljB218A. Therefore, we produced a monoclonal antibody (MAb), 23D4, isotyped as immunoglobulin M, against H:1,2 S. enterica serovar Typhimurium flagellin. The mutant flagellin was not recognized by this MAb. When a large number of phase 1 and phase 2 flagellin antigens of different serovars were used to characterize the 23D4 MAb, only extracts of serovars Typhimurium and [4,5,12:i:−] reacted. The protein composition of phase 1 and phase 2 extracts and highly purified H:1,2 flagellin from serovar Typhimurium strain LT2 and extract of strain 286 (serovar [4,5,12:i:−]), which reacted with the MAb, was studied. Phase 2 flagellin (FljBH:1,2) was detected in phase 1 and phase 2 flagellar heat extracts of serovar Typhimurium and was the single protein identified in all spots of purified H:1,2 flagellin. FliC, FlgK, and other proteins were detected in some immunoreactive spots and in the flagellar extract of serovar [4,5,12:i:−]. Immunoelectron microscopy of complete bacteria with 23D4 showed MAb attachment at the base of flagella, although the MAb failed to recognize the filament of flagella. Nevertheless, the results obtained by the other immunological tests (enzyme-linked immunosorbent assay, Western blotting, and dot blotting) indicate a reaction against flagellins. The epitopes could also be shared by other proteins on spots where FljB is not present, such as aminopeptidase B, isocitrate lyase, InvE, EF-TuA, enolase, DnaK, and others. In conclusion

  18. Selection of Small-Colony Variants of Salmonella enterica Serovar Typhimurium in Nonphagocytic Eucaryotic Cells

    PubMed Central

    Cano, David A.; Pucciarelli, M. Graciela; Martínez-Moya, Marina; Casadesús, Josep; García-del Portillo, Francisco

    2003-01-01

    Salmonella enterica strains are enteropathogenic bacteria that survive and proliferate within vacuolar compartments of epithelial and phagocytic cells. Recently, it has been reported that fibroblast cells are capable of restricting S. enterica serovar Typhimurium intracellular growth. Here, we show that prolonged residence of bacteria in the intracellular environment of fibroblasts results in the appearance of genetically stable small-colony variants (SCV). A total of 103 SCV isolates, obtained from four independent infections, were subjected to phenotypic analysis. The following phenotypes were observed: (i) δ-aminolevulinic acid auxotrophy; (ii) requirement for acetate or succinate for growth in glucose minimal medium; (iii) auxotrophy for aromatic amino acids; and (iv) reduced growth rate under aerobic conditions not linked to nutrient auxotrophy. The exact mutations responsible for the SCV phenotype in three representative isolates were mapped in the lpd, hemL, and aroD genes, which code for dihydrolipoamide dehydrogenase, glutamate-1-semyaldehyde aminotransferase, and 3-dehydroquinate dehydratase, respectively. The lpd, hemL, and aroD mutants had intracellular persistence rates in fibroblasts that were 3 to 4 logs higher than that of the parental strain and decreased susceptibility to aminoglycoside antibiotics. All three of these SCV isolates were attenuated in the BALB/c murine typhoid model. Complementation with lpd+, hem+, and aroD+ genes restored the levels of intracellular persistence and antibiotic susceptibility to levels of the wild-type strain. However, virulence was not exhibited by any of the complemented strains. Altogether, our data demonstrate that similar to what it has been reported for SCV isolates of other pathogens, S. enterica SCV display enhanced intracellular persistence in eucaryotic cells and are impaired in the ability to cause overt disease. In addition, they also suggest that S. enterica SCV may be favored in vivo. PMID:12819049

  19. The Tricarballylate Utilization (tcuRABC) Genes of Salmonella enterica Serovar Typhimurium LT2

    PubMed Central

    Lewis, Jeffrey A.; Horswill, Alexander R.; Schwem, Brian E.; Escalante-Semerena, Jorge C.

    2004-01-01

    The genes of Salmonella enterica serovar Typhimurium LT2 encoding functions needed for the utilization of tricarballylate as a carbon and energy source were identified and their locations in the chromosome were established. Three of the tricarballylate utilization (tcu) genes, tcuABC, are organized as an operon; a fourth gene, tcuR, is located immediately 5′ to the tcuABC operon. The tcuABC operon and tcuR gene share the same direction of transcription but are independently transcribed. The tcuRABC genes are missing in the Escherichia coli K-12 chromosome. The tcuR gene is proposed to encode a regulatory protein needed for the expression of tcuABC. The tcuC gene is proposed to encode an integral membrane protein whose role is to transport tricarballylate across the cell membrane. tcuC function was sufficient to allow E. coli K-12 to grow on citrate (a tricarballylate analog) but not to allow growth of this bacterium on tricarballylate. E. coli K-12 carrying a plasmid with wild-type alleles of tcuABC grew on tricarballylate, suggesting that the functions of the TcuABC proteins were the only ones unique to S. enterica needed to catabolize tricarballylate. Analyses of the predicted amino acid sequences of the TcuAB proteins suggest that TcuA is a flavoprotein, and TcuB is likely anchored to the cell membrane and probably contains one or more Fe-S centers. The TcuB protein is proposed to work in concert with TcuA to oxidize tricarballylate to cis-aconitate, which is further catabolized via the Krebs cycle. The glyoxylate shunt is not required for growth of S. enterica on tricarballylate. A model for tricarballylate catabolism in S. enterica is proposed. PMID:14996793

  20. The Tricarballylate utilization (tcuRABC) genes of Salmonella enterica serovar Typhimurium LT2.

    PubMed

    Lewis, Jeffrey A; Horswill, Alexander R; Schwem, Brian E; Escalante-Semerena, Jorge C

    2004-03-01

    The genes of Salmonella enterica serovar Typhimurium LT2 encoding functions needed for the utilization of tricarballylate as a carbon and energy source were identified and their locations in the chromosome were established. Three of the tricarballylate utilization (tcu) genes, tcuABC, are organized as an operon; a fourth gene, tcuR, is located immediately 5' to the tcuABC operon. The tcuABC operon and tcuR gene share the same direction of transcription but are independently transcribed. The tcuRABC genes are missing in the Escherichia coli K-12 chromosome. The tcuR gene is proposed to encode a regulatory protein needed for the expression of tcuABC. The tcuC gene is proposed to encode an integral membrane protein whose role is to transport tricarballylate across the cell membrane. tcuC function was sufficient to allow E. coli K-12 to grow on citrate (a tricarballylate analog) but not to allow growth of this bacterium on tricarballylate. E. coli K-12 carrying a plasmid with wild-type alleles of tcuABC grew on tricarballylate, suggesting that the functions of the TcuABC proteins were the only ones unique to S. enterica needed to catabolize tricarballylate. Analyses of the predicted amino acid sequences of the TcuAB proteins suggest that TcuA is a flavoprotein, and TcuB is likely anchored to the cell membrane and probably contains one or more Fe-S centers. The TcuB protein is proposed to work in concert with TcuA to oxidize tricarballylate to cis-aconitate, which is further catabolized via the Krebs cycle. The glyoxylate shunt is not required for growth of S. enterica on tricarballylate. A model for tricarballylate catabolism in S. enterica is proposed.

  1. Salmonella Enterica Serovar Typhimurium BipA Exhibits Two Distinct Ribosome Binding Modes

    SciTech Connect

    deLivron, M.; Robinson, V

    2008-01-01

    BipA is a highly conserved prokaryotic GTPase that functions to influence numerous cellular processes in bacteria. In Escherichia coli and Salmonella enterica serovar Typhimurium, BipA has been implicated in controlling bacterial motility, modulating attachment and effacement processes, and upregulating the expression of virulence genes and is also responsible for avoidance of host defense mechanisms. In addition, BipA is thought to be involved in bacterial stress responses, such as those associated with virulence, temperature, and symbiosis. Thus, BipA is necessary for securing bacterial survival and successful invasion of the host. Steady-state kinetic analysis and pelleting assays were used to assess the GTPase and ribosome-binding properties of S. enterica BipA. Under normal bacterial growth, BipA associates with the ribosome in the GTP-bound state. However, using sucrose density gradients, we demonstrate that the association of BipA and the ribosome is altered under stress conditions in bacteria similar to those experienced during virulence. The data show that this differential binding is brought about by the presence of ppGpp, an alarmone that signals the onset of stress-related events in bacteria.

  2. The Legacy of Genetic Analysis Advances Contemporary Research with Escherichia coli K-12 and Salmonella enterica serovar Typhimurium LT2.

    PubMed

    Stewart, Valley

    2017-04-01

    Escherichia coli K-12 and Salmonella enterica serovar Typhimurium LT2 became standard organisms for genetic analysis during the Truman administration. Half a century later, genetic analysis with these strains had become an art form, interpreted through 23 articles in the ambitious two-volume masterpiece edited by the late Fred Neidhardt and colleagues. These legacy articles now are available through EcoSal Plus, so as to inform and inspire contemporary genetic analyses in these standard organisms and their relatives.

  3. Net replication of Salmonella enterica serovars Typhimurium and Choleraesuis in porcine intestinal mucosa and nodes is associated with their differential virulence.

    PubMed

    Paulin, Susan M; Jagannathan, Aparna; Campbell, June; Wallis, Timothy S; Stevens, Mark P

    2007-08-01

    Salmonella enterica is a facultative intracellular pathogen of worldwide importance and causes a spectrum of diseases depending on serovar- and host-specific factors. Oral infection of pigs with S. enterica serovar Typhimurium strain 4/74 produces acute enteritis but is rarely fatal, whereas serovar Choleraesuis strain A50 causes systemic disease with a high mortality rate. With a porcine ligated ileal loop model, we observed that systemic virulence of serovar Choleraesuis A50 is not associated with enhanced intestinal invasion, secretory responses, or neutrophil recruitment compared to serovar Typhimurium 4/74. The net growth in vivo of serovar Choleraesuis A50 and serovar Typhimurium 4/74 was monitored following oral inoculation of pigs with strains harboring pHSG422, which exhibits temperature-sensitive replication. Analysis of plasmid partitioning revealed that the enteric virulence of serovar Typhimurium 4/74 relative to that of serovar Choleraesuis A50 is associated with rapid replication in the intestinal wall, whereas systemic virulence of serovar Choleraesuis A50 is associated with enhanced persistence in intestinal mesenteric lymph nodes. Faster replication of serovar Typhimurium, compared to that of serovar Choleraesuis, in the intestinal mucosa was associated with greater induction of the proinflammatory cytokines tumor necrosis factor alpha, interleukin-8 (IL-8), and IL-18 as detected by reverse transcriptase PCR analysis of transcripts from infected mucosa. During replication in batch culture and porcine alveolar macrophages, transcription of genes encoding components of type III secretion systems 1 (sipC) and 2 (sseC) was observed to be significantly higher in serovar Typhimurium 4/74 than in serovar Choleraesuis A50, and this may contribute to the differences in epithelial invasion and intracellular proliferation. The rapid induction of proinflammatory responses by strain 4/74 may explain why pigs confine serovar Typhimurium infection to the

  4. Rescuing chemotaxis of the anticancer agent Salmonella enterica serovar Typhimurium VNP20009.

    PubMed

    Broadway, Katherine M; Denson, Elizabeth A P; Jensen, Roderick V; Scharf, Birgit E

    2015-10-10

    The role of chemotaxis and motility in Salmonella enterica serovar Typhimurium tumor colonization remains unclear. We determined through swim plate assays that the well-established anticancer agent S. Typhimurium VNP20009 is deficient in chemotaxis, and that this phenotype is suppressible. Through genome sequencing, we revealed that VNP20009 and four selected suppressor mutants had a single nucleotide polymorphism (SNP) in cheY causing a mutation in the conserved proline residue at position 110. CheY is the response regulator that interacts with the flagellar motor-switch complex and modulates rotational bias. The four suppressor mutants additionally carried non-synonymous SNPs in fliM encoding a flagellar switch protein. The CheY-P110S mutation in VNP20009 likely rendered the protein unable to interact with FliM, a phenotype that could be suppressed by mutations in FliM. We replaced the mutated cheY in VNP20009 with the wild-type copy and chemotaxis was partially restored. The swim ring of the rescued strain, VNP20009 cheY(+), was 46% the size of the parental strain 14028 swim ring. When tested in capillary assays, VNP20009 cheY(+) was 69% efficient in chemotaxis towards the attractant aspartate as compared to 14028. Potential reasons for the lack of complete restoration and implications for bacterial tumor colonization will be discussed. Copyright © 2015 Elsevier B.V. All rights reserved.

  5. Stress-induced prophage DNA replication in Salmonella enterica serovar Typhimurium.

    PubMed

    Garcia-Russell, Nathalie; Elrod, Brandon; Dominguez, Katrina

    2009-09-01

    Salmonella Typhimurium, a foodborne pathogen, is the cause of new outbreaks every year. The virulence of new pathogens is determined by their virulence genes, many of them carried on transferable elements, such as prophages. In bacteria harboring multiple prophages such as Salmonella, the reassortment of these genes plays a major role in the emergence of new pathogens and consequently new epidemics. This gene transfer depends on prophage induction and the initiation of the phage lytic cycle. In the present study we have tested the effects of bacterial extracytoplasmic stress on prophage induction. We developed a quantitative real-time PCR assay to quantify variations in phage genes copy number, representative of phage DNA replication associated with the initiation of the lytic cycle. The induction of the four Salmonella enterica serovar Typhimurium LT2 prophages (Fels-1, Fels-2, Gifsy-1 and Gifsy-2) was measured during exponential growth, stationary phase, starvation, as well as after treatment with Mitomycin C, Ampicillin or heat. Our results show that the four prophages respond differently to each treatment. Gifsy-2 showed constant low level of induction independently of the extracytoplasmic stress, Fels-1 was strongly induced after DNA damage, Fels-2 showed spontaneous induction only during optimal bacterial growth, and Gifsy-1 was repressed in all conditions. These findings show that the transfer of virulence genes can respond to and depend on variations of the bacterial surrounding conditions, and help to explain the appearance of new Salmonella outbreaks.

  6. First Complete Genome Sequence of Salmonella enterica subsp. enterica Serovar Typhimurium Strain ATCC 13311 (NCTC 74), a Reference Strain of Multidrug Resistance, as Achieved by Use of PacBio Single-Molecule Real-Time Technology

    PubMed Central

    Juan, Ayaka; Tamotsu, Hinako; Ashimine, Noriko; Nakano, Kazuma; Shimoji, Makiko; Shiroma, Akino; Teruya, Kuniko; Satou, Kazuhito; Hirano, Takashi

    2014-01-01

    We report the first complete genomic sequence of Salmonella enterica subsp. enterica serovar Typhimurium strain ATCC 13311, the leading food-borne pathogen and a reference strain used in drug resistance studies. De novo assembly with PacBio sequencing completed its chromosome and one plasmid. They will accelerate the investigation into multidrug resistance in Salmonella Typhimurium. PMID:25278532

  7. Characterization of the yehUT two-component regulatory system of Salmonella enterica Serovar Typhi and Typhimurium.

    PubMed

    Wong, Vanessa K; Pickard, Derek J; Barquist, Lars; Sivaraman, Karthikeyan; Page, Andrew J; Hart, Peter J; Arends, Mark J; Holt, Kathryn E; Kane, Leanne; Mottram, Lynda F; Ellison, Louise; Bautista, Ruben; McGee, Chris J; Kay, Sally J; Wileman, Thomas M; Kenney, Linda J; MacLennan, Calman A; Kingsley, Robert A; Dougan, Gordon

    2013-01-01

    Proteins exhibiting hyper-variable sequences within a bacterial pathogen may be associated with host adaptation. Several lineages of the monophyletic pathogen Salmonella enterica serovar Typhi (S. Typhi) have accumulated non-synonymous mutations in the putative two-component regulatory system yehUT. Consequently we evaluated the function of yehUT in S. Typhi BRD948 and S. Typhimurium ST4/74. Transcriptome analysis identified the cstA gene, encoding a carbon starvation protein as the predominantly yehUT regulated gene in both these serovars. Deletion of yehUT had no detectable effect on the ability of these mutant Salmonella to invade cultured epithelial cells (S. Typhi and S. Typhimurium) or induce colitis in a murine model (S. Typhimurium only). Growth, metabolic and antimicrobial susceptibility tests identified no obvious influences of yehUT on these phenotypes.

  8. A comparative study of thermal and acid inactivation kinetics in fruit juices of Salmonella enterica serovar Typhimurium and Salmonella enterica serovar Senftenberg grown at acidic conditions.

    PubMed

    Alvarez-Ordóñez, Avelino; Fernández, Ana; Bernardo, Ana; López, Mercedes

    2009-11-01

    Acid and heat inactivation in orange and apple juices of Salmonella enterica serovar Typhimurium Colección Española de Cultivos Tipo (i.e., Spanish Type Culture Collection) 443 (CECT 443) (Salmonella Typhimurium) and S. enterica serovar Senftenberg CECT 4384 (Salmonella Senftenberg) grown in buffered brain heart infusion (pH 7.0) and acidified brain heart infusion up to pH 4.5 with acetic, citric, lactic, and hydrochloric acids was evaluated. Acid adaptation induced an adaptive response that increased the subsequent resistance to extreme pH conditions (pH 2.5) and to heat, although the magnitude of these responses differed between the two isolates and fruit juices. The acid resistance in orange juice for acid-adapted cells (D-values of 28.3-34.5 min for Salmonella Senftenberg and 30.0-39.2 min for Salmonella Typhimurium) resulted to be about two to three times higher than that corresponding to non-acid-adapted cells. In apple juice, acid-adapted Salmonella Senftenberg cells survived better than those of Salmonella Typhimurium, obtaining mean D-values of 114.8 +/- 12.3 and 41.9 +/- 2.5 min, respectively. The thermotolerance of non-acid-adapted Salmonella Typhimurium in orange (D(58)-value: 0.028 min) and apple juices (D(58)-value: 0.10 min) was approximately double for acid-adapted cells. This cross-protection to heat was more strongly expressed in Salmonella Senftenberg. D(58)-values obtained for non-acid-adapted cells in orange (0.11 min) and apple juices (0.19 min) increased approximately 10 and 5 times, respectively, after their growth in acidified media. The conditions prevailing during bacterial growth and heat treatment did not significantly influence the z-values observed (6.0 +/- 0.3 degrees C for Salmonella Typhimurium and 7.0 +/- 0.3 degrees C for Salmonella Senftenberg). The enhanced acid resistance found for both isolates could enable them to survive for prolonged time periods in the gastrointestinal tract, increasing the risk of illness. Further, it

  9. Increased efficacy of inactivated vaccine candidates prepared with Salmonella enterica serovar Typhimurium strains of predominant genotypes in ducks.

    PubMed

    Youn, S Y; Kwon, Y K; Song, C S; Lee, H J; Jeong, O M; Choi, B K; Jung, S C; Kang, M S

    2016-08-01

    Salmonella enterica serovar Typhimurium has been a major causative agent of food-borne human disease, mainly due to consumption of contaminated food animal products. In particular, ducks serve as a reservoir of serovar Typhimurium, and are one of the common sources of human infection. To prevent infection of ducks, and therefore minimize human infection, it is critical to control the persistent epidemic strains in ducks. Here, we analyzed the genetic diversity and virulence of serovar Typhimurium isolates from ducks in Korea to identify the predominant strains that might be used as efficient vaccine candidates for ducks. Among the isolates, 2 representative isolates (ST26 and ST76) of predominant genotypes were selected as vaccine strains on the basis of genotypic analysis by pulsed-field gel electrophoresis and DNA microarrays. Two-week-old ducks were then injected intramuscularly with inactivated vaccine candidates prepared using ST26 or ST76 (10(8) cfu/0.5 mL/duck or 10(9) cfu/0.5 mL/duck), and oral challenge with a highly virulent serovar Typhimurium strain (10(9) cfu/0.5 mL/duck) was carried out 2 wk later. Shedding of the challenge strain was significantly decreased in group 2 after vaccination. The antibody levels by enzyme-linked immunosorbent assay in all vaccinated groups were enhanced significantly (P < 0.05) compared to the unvaccinated control group. Overall, vaccination with ST26 or ST76 reduced bacterial shedding and colonization in internal organs, and induced elevated antibody response. In particular, serovar Typhimurium ST26 (10(8) cfu/0.5 mL/duck) was the most effective vaccine candidate, which can provide efficient protection against serovar Typhimurium in ducks with higher effectiveness compared to a commercial vaccine currently used worldwide. © 2016 Poultry Science Association Inc.

  10. Infection of Mice by Salmonella enterica Serovar Enteritidis Involves Additional Genes That Are Absent in the Genome of Serovar Typhimurium

    PubMed Central

    Silva, Cecilia A.; Blondel, Carlos J.; Quezada, Carolina P.; Porwollik, Steffen; Andrews-Polymenis, Helene L.; Toro, Cecilia S.; Zaldívar, Mercedes; Contreras, Inés

    2012-01-01

    Salmonella enterica serovar Enteritidis causes a systemic, typhoid-like infection in newly hatched poultry and mice. In the present study, a library of 54,000 transposon mutants of S. Enteritidis phage type 4 (PT4) strain P125109 was screened for mutants deficient in the in vivo colonization of the BALB/c mouse model using a microarray-based negative-selection screening. Mutants in genes known to contribute to systemic infection (e.g., Salmonella pathogenicity island 2 [SPI-2], aro, rfa, rfb, phoP, and phoQ) and enteric infection (e.g., SPI-1 and SPI-5) in this and other Salmonella serovars displayed colonization defects in our assay. In addition, a strong attenuation was observed for mutants in genes and genomic islands that are not present in S. Typhimurium or in most other Salmonella serovars. These genes include a type I restriction/modification system (SEN4290 to SEN4292), the peg fimbrial operon (SEN2144A to SEN2145B), a putative pathogenicity island (SEN1970 to SEN1999), and a type VI secretion system remnant SEN1001, encoding a hypothetical protein containing a lysin motif (LysM) domain associated with peptidoglycan binding. Proliferation defects for mutants in these individual genes and in exemplar genes for each of these clusters were confirmed in competitive infections with wild-type S. Enteritidis. A ΔSEN1001 mutant was defective for survival within RAW264.7 murine macrophages in vitro. Complementation assays directly linked the SEN1001 gene to phenotypes observed in vivo and in vitro. The genes identified here may perform novel virulence functions not characterized in previous Salmonella models. PMID:22083712

  11. HilA gene expression in SCFAs adapted and inorganic acid challenged Salmonella enterica serovar typhimurium.

    PubMed

    Rishi, Praveen; Ricke, Steven

    2007-09-01

    Salmonella enterica serovar typhimurium (S. typhimurium) encounters short chain fatty acids (inorganic acids containing propionate, butyrate and acetate) in the intestine as well as in food preservatives. Short chain fatty acids (SCFAs) exposed organisms have been reported to offer resistance to organic acid resulting into enhanced virulence. However, the role of hilA (hyper invasive loci) gene expression has not been assessed in this context. In the present study, S. typhimurium was grown under SCFAs stress condition simulating the in vivo environment and hilA gene expression was evaluated. The gene expression was measured by beta-galactosidase (beta-gal) assay using a hilA-lacZY fusion strain and calculated as Miller units. hilA gene expression was found to be significantly higher in the SCFAs exposed cells than the unexposed ones, after 2 hrs and 4 hrs of exposure. However, no significant difference was observed between the activities at 2 hrs and 4 hrs. It indicates that hilA gene gets expressed by 2 hrs and persists till 4 hrs at least. The beta-gal activity was also measured in the unadapted / SCFAs adapted organisms followed by acid shock for 1 hr. The gene expression was also found to be higher in the SCFAs adapted--acid (pH 3) challenged as compared to the unadapated acid challenged organisms suggesting that SCFAs adaptation may induce organic acid tolerance by modulating the hilA response. This observation indicates that hilA may be the additional gene contributing to acid resistance and thereby increasing virulence of the organism after SCFAs adaptation.

  12. The Role of RamA on the Development of Ciprofloxacin Resistance in Salmonella enterica Serovar Typhimurium

    PubMed Central

    Sun, Yawei; Dai, Menghong; Hao, Haihong; Wang, Yulian; Huang, Lingli; Almofti, Yassir A.; Liu, Zhenli; Yuan, Zonghui

    2011-01-01

    Active efflux pump is a primary fluoroquinolone resistant mechanism of clinical isolates of Salmonella enterica serovar Typhimurium. RamA is an essential element in producing multidrug resistant (MDR) S.enterica serovar Typhimurium. The aim of the present study was to elucidate the roles of RamA on the development of ciprofloxacin, the first choice for the treatment of salmonellosis, resistance in S. enterica serovar Typhimurium. Spontaneous mutants were selected via several passages of S. enterica serovar Typhimurium CVCC541 susceptible strain (ST) on M-H agar with increasing concentrations of ciprofloxacin (CIP). Accumulation of ciprofloxacin was tested by the modified fluorometric method. The expression levels of MDR efflux pumps were determined by real time RT-PCR. In ST and its spontaneous mutants, the ramA gene was inactivated by insertion of the kan gene and compensated on a recombinant plasmid pGEXΦ(gst-ramA). The mutant prevention concentration (MPC) and mutant frequencies of ciprofloxacin against ST and a spontaneous mutant in the presence, absence and overexpression of RamA were tested. Four spontaneous mutants (SI1-SI4) were obtained. The SI1 (CIP MICs, 0.1 mg/L) without any target site mutation in its quinolone resistant determining regions (QRDRs) and SI3 (CIP MICs, 16 mg/L) harboring the Ser83→Phe mutation in its QRDR of GyrA strains exhibited reduced susceptibility and resistance to multidrugs, respectively. In SI1, RamA was the main factor that controlled the susceptibility to ciprofloxacin by activating MdtK as well as increasing the expression level of acrAB. In SI3, RamA played predominant role in ciprofloxacin resistance via increasing the expression level of acrAB. Likewise, the deficiency of RamA decreased the MPCs and mutant frequencies of ST and SI2 to ciprofloxacin. In conclusion, the expression of RamA promoted the development of ciprofloxacin resistant mutants of S. enterica serovar Typhimurium. The inhibition of RamA could decrease

  13. The role of RamA on the development of ciprofloxacin resistance in Salmonella enterica serovar Typhimurium.

    PubMed

    Sun, Yawei; Dai, Menghong; Hao, Haihong; Wang, Yulian; Huang, Lingli; Almofti, Yassir A; Liu, Zhenli; Yuan, Zonghui

    2011-01-01

    Active efflux pump is a primary fluoroquinolone resistant mechanism of clinical isolates of Salmonella enterica serovar Typhimurium. RamA is an essential element in producing multidrug resistant (MDR) S. enterica serovar Typhimurium. The aim of the present study was to elucidate the roles of RamA on the development of ciprofloxacin, the first choice for the treatment of salmonellosis, resistance in S. enterica serovar Typhimurium. Spontaneous mutants were selected via several passages of S. enterica serovar Typhimurium CVCC541 susceptible strain (ST) on M-H agar with increasing concentrations of ciprofloxacin (CIP). Accumulation of ciprofloxacin was tested by the modified fluorometric method. The expression levels of MDR efflux pumps were determined by real time RT-PCR. In ST and its spontaneous mutants, the ramA gene was inactivated by insertion of the kan gene and compensated on a recombinant plasmid pGEXΦ(gst-ramA). The mutant prevention concentration (MPC) and mutant frequencies of ciprofloxacin against ST and a spontaneous mutant in the presence, absence and overexpression of RamA were tested. Four spontaneous mutants (SI1-SI4) were obtained. The SI1 (CIP MICs, 0.1 mg/L) without any target site mutation in its quinolone resistant determining regions (QRDRs) and SI3 (CIP MICs, 16 mg/L) harboring the Ser83→Phe mutation in its QRDR of GyrA strains exhibited reduced susceptibility and resistance to multidrugs, respectively. In SI1, RamA was the main factor that controlled the susceptibility to ciprofloxacin by activating MdtK as well as increasing the expression level of acrAB. In SI3, RamA played predominant role in ciprofloxacin resistance via increasing the expression level of acrAB. Likewise, the deficiency of RamA decreased the MPCs and mutant frequencies of ST and SI2 to ciprofloxacin. In conclusion, the expression of RamA promoted the development of ciprofloxacin resistant mutants of S. enterica serovar Typhimurium. The inhibition of RamA could decrease

  14. Complete Genome Sequence of Salmonella enterica Serovar Typhimurium Strain YU15 (Sequence Type 19) Harboring the Salmonella Genomic Island 1 and Virulence Plasmid pSTV

    PubMed Central

    Calva, Edmundo; Puente, José L.; Zaidi, Mussaret B.

    2016-01-01

    The complete genome of Salmonella enterica subsp. enterica serovar Typhimurium sequence type 19 (ST19) strain YU15, isolated in Yucatán, Mexico, from a human baby stool culture, was determined using PacBio technology. The chromosome contains five intact prophages and the Salmonella genomic island 1 (SGI1). This strain carries the Salmonella virulence plasmid pSTV. PMID:27081132

  15. Complete Genome Sequencing of a Multidrug-Resistant and Human-Invasive Salmonella enterica Serovar Typhimurium Strain of the Emerging Sequence Type 213 Genotype

    PubMed Central

    Calva, Edmundo; Zaidi, Mussaret B.; Sanchez-Flores, Alejandro; Estrada, Karel; Silva, Genivaldo G. Z.; Soto-Jiménez, Luz M.; Wiesner, Magdalena; Fernández-Mora, Marcos; Edwards, Robert A.

    2015-01-01

    Salmonella enterica subsp. enterica serovar Typhimurium strain YU39 was isolated in 2005 in the state of Yucatán, Mexico, from a human systemic infection. The YU39 strain is representative of the multidrug-resistant emergent sequence type 213 (ST213) genotype. The YU39 complete genome is composed of a chromosome and seven plasmids. PMID:26089426

  16. Complete genome sequencing of a multidrug-resistant and human-invasive Salmonella enterica serovar Typhimurium strain of the emerging sequence type 213 genotype

    DOE PAGES

    Calva, Edmundo; Silva, Claudia; Zaidi, Mussaret B.; ...

    2015-06-18

    Salmonella enterica subsp. enterica serovar Typhimurium strain YU39 was isolated in 2005 in the state of Yucatán, Mexico, from a human systemic infection. The YU39 strain is representative of the multidrug-resistant emergent sequence type 213 (ST213) genotype. The YU39 complete genome is composed of a chromosome and seven plasmids.

  17. Efflux Pump Overexpression Contributes to Tigecycline Heteroresistance in Salmonella enterica serovar Typhimurium.

    PubMed

    Chen, Yi; Hu, Daxing; Zhang, Qijing; Liao, Xiao-Ping; Liu, Ya-Hong; Sun, Jian

    2017-01-01

    Bacterial heteroresistance has been identified in several combinations of bacteria and antibiotics, and it complicated the therapeutic strategies. Tigecycline is being used as one of the optimal options for the treatment of infections caused by multidrug-resistant Salmonella. This study investigated whether heterorresistance to tigecycline exists in a Salmonella enterica serovar Typhimurium strain harboring the oqxAB-bearing IncHI2 plasmid pHXY0908. MIC and population analyses were performed to evaluate population-wide susceptibility to tigecycline. The effects of efflux pumps on MIC levels were assessed using the efflux pump inhibitor Phe-Arg-β-naphthylamide, measuring intracellular tigecycline accumulation as well as mRNA levels of regulatory and efflux pump genes. DNA sequencing of regulatory regions were performed and plasmid curing from a resistant strain provided an appropriate control. Results showed that MICs of a parental strain with and without pHXY0908 as well as a plasmid-cured strain 14028/Δp52 were 0.5, 1, and 1 μg/mL, respectively. Population analysis profiling (PAP) illustrated that only the pHXY0908-containg strain was heteroresistant to tigecycline. A fraction of colonies exhibited stable profiles with 4- to 8-fold increases in MIC. The frequencies of emergence of these isolates were higher in the plasmid-containing strain pHXY0908 than either the parental or the 14028/Δp52 strain. Phe-Arg-β-naphthylamide addition restored tigecycline susceptibility of these isolates and intracellular tigecycline accumulation was reduced. Heteroresistant isolates of the strain containing pHXY0908 also had elevated expression of acrB, ramA, and oqxB. DNA sequencing identified numerous mutations in RamR that have been shown to lead to ramA overexpression. In conclusions, heteroresistance to tigecycline in Salmonella enterica serovar Typhimurium was manifested in a plasmid-bearing strain. Our results suggest that this phenotype was associated with overexpression

  18. Efflux Pump Overexpression Contributes to Tigecycline Heteroresistance in Salmonella enterica serovar Typhimurium

    PubMed Central

    Chen, Yi; Hu, Daxing; Zhang, Qijing; Liao, Xiao-Ping; Liu, Ya-Hong; Sun, Jian

    2017-01-01

    Bacterial heteroresistance has been identified in several combinations of bacteria and antibiotics, and it complicated the therapeutic strategies. Tigecycline is being used as one of the optimal options for the treatment of infections caused by multidrug-resistant Salmonella. This study investigated whether heterorresistance to tigecycline exists in a Salmonella enterica serovar Typhimurium strain harboring the oqxAB-bearing IncHI2 plasmid pHXY0908. MIC and population analyses were performed to evaluate population-wide susceptibility to tigecycline. The effects of efflux pumps on MIC levels were assessed using the efflux pump inhibitor Phe-Arg-β-naphthylamide, measuring intracellular tigecycline accumulation as well as mRNA levels of regulatory and efflux pump genes. DNA sequencing of regulatory regions were performed and plasmid curing from a resistant strain provided an appropriate control. Results showed that MICs of a parental strain with and without pHXY0908 as well as a plasmid-cured strain 14028/Δp52 were 0.5, 1, and 1 μg/mL, respectively. Population analysis profiling (PAP) illustrated that only the pHXY0908-containg strain was heteroresistant to tigecycline. A fraction of colonies exhibited stable profiles with 4- to 8-fold increases in MIC. The frequencies of emergence of these isolates were higher in the plasmid-containing strain pHXY0908 than either the parental or the 14028/Δp52 strain. Phe-Arg-β-naphthylamide addition restored tigecycline susceptibility of these isolates and intracellular tigecycline accumulation was reduced. Heteroresistant isolates of the strain containing pHXY0908 also had elevated expression of acrB, ramA, and oqxB. DNA sequencing identified numerous mutations in RamR that have been shown to lead to ramA overexpression. In conclusions, heteroresistance to tigecycline in Salmonella enterica serovar Typhimurium was manifested in a plasmid-bearing strain. Our results suggest that this phenotype was associated with overexpression

  19. Iron regulated genes of Salmonella enterica serovar Typhimurium in response to norepinephrine and the requirement of fepCDG for norepinephrine-enhanced growth

    USDA-ARS?s Scientific Manuscript database

    The presence of catecholamines in vivo may stimulate enteric bacteria including the foodborne pathogen Salmonella enterica serovar Typhimurium by two mechanisms, acting as a quorum sensing signal and providing iron in the presence of serum. To identify genes of Salmonella Typhimurium that participa...

  20. The Salmonella enterica serovar Typhimurium QseB Response Regulator Negatively Regulates Bacterial Motility and Swine Colonization in the Absence of the QseC Sensor Kinase

    USDA-ARS?s Scientific Manuscript database

    Salmonella enterica serovar Typhimurium (S. Typhimurium) responds to the catecholamine, norepinephrine by increasing bacterial growth and enhancing motility. In this study, iron with or without the siderophore, ferrioxamine E also enhanced bacterial motility. Iron-enhanced motility was growth-rate ...

  1. A Mutation in the PoxA Gene of Salmonella enterica Serovar Typhimurium Results in Altered Protein Production, Elevated Susceptibility to Environmental Challenges, and Decreased Swine Colonization

    USDA-ARS?s Scientific Manuscript database

    Using signature-tagged mutagenesis of Salmonella enterica serovar Typhimurium (S. Typhimurium), a mutation in the poxA gene (STM4344; yjeA; poxR), encoding a putative lysyl-tRNA synthetase, was previously identified by our research group which caused decreased survival in an ex vivo swine stomach co...

  2. 6S RNA is involved in acid resistance and invasion of epithelial cells in Salmonella enterica serovar Typhimurium.

    PubMed

    Ren, Jie; Sang, Yu; Qin, Ran; Cui, Zhongli; Yao, Yu-Feng

    2017-09-01

    Acid is an important environmental condition encountered frequently by Salmonella enterica serovar Typhimurium during its pathogenesis, but the role of small-noncoding RNAs (sRNAs) in response to acid stress is poorly understood. We used RNA sequencing to explore acid-responsive sRNAs in S. Typhimurium. It identified that 6S RNA encoded by the ssrS was significantly upregulated at pH 3.0. The 6S RNA knockout strain showed a reduced ability to survive at pH 3.0. Additionally, genes in Salmonella pathogenicity island-1 were downregulated in the 6S RNA knockout strain. The loss of 6S RNA significantly reduced S. Typhimurium invasion ability in HeLa cells and virulence in a mouse model. These findings demonstrate that 6S RNA plays an important role in S. Typhimurium survival under extremely acid conditions and for invasion of epithelial cells.

  3. Caveolin-1-deficient mice show defects in innate immunity and inflammatory immune response during Salmonella enterica serovar Typhimurium infection.

    PubMed

    Medina, Freddy A; de Almeida, Cecilia J; Dew, Elliott; Li, Jiangwei; Bonuccelli, Gloria; Williams, Terence M; Cohen, Alex W; Pestell, Richard G; Frank, Philippe G; Tanowitz, Herbert B; Lisanti, Michael P

    2006-12-01

    A number of studies have shown an association of pathogens with caveolae. To this date, however, there are no studies showing a role for caveolin-1 in modulating immune responses against pathogens. Interestingly, expression of caveolin-1 has been shown to occur in a regulated manner in immune cells in response to lipopolysaccharide (LPS). Here, we sought to determine the role of caveolin-1 (Cav-1) expression in Salmonella pathogenesis. Cav-1(-/-) mice displayed a significant decrease in survival when challenged with Salmonella enterica serovar Typhimurium. Spleen and tissue burdens were significantly higher in Cav-1(-/-) mice. However, infection of Cav-1(-/-) macrophages with serovar Typhimurium did not result in differences in bacterial invasion. In addition, Cav-1(-/-) mice displayed increased production of inflammatory cytokines, chemokines, and nitric oxide. Regardless of this, Cav-1(-/-) mice were unable to control the systemic infection of Salmonella. The increased chemokine production in Cav-1(-/-) mice resulted in greater infiltration of neutrophils into granulomas but did not alter the number of granulomas present. This was accompanied by increased necrosis in the liver. However, Cav-1(-/-) macrophages displayed increased inflammatory responses and increased nitric oxide production in vitro in response to Salmonella LPS. These results show that caveolin-1 plays a key role in regulating anti-inflammatory responses in macrophages. Taken together, these data suggest that the increased production of toxic mediators from macrophages lacking caveolin-1 is likely to be responsible for the marked susceptibility of caveolin-1-deficient mice to S. enterica serovar Typhimurium.

  4. aroA-Deficient Salmonella enterica Serovar Typhimurium Is More Than a Metabolically Attenuated Mutant

    PubMed Central

    Frahm, Michael; Kocijancic, Dino; Rohde, Manfred; Eckweiler, Denitsa; Bielecka, Agata; Bueno, Emilio; Cava, Felipe; Abraham, Wolf-Rainer; Curtiss, Roy; Häussler, Susanne; Erhardt, Marc; Weiss, Siegfried

    2016-01-01

    ABSTRACT Recombinant attenuated Salmonella enterica serovar Typhimurium strains are believed to act as powerful live vaccine carriers that are able to elicit protection against various pathogens. Auxotrophic mutations, such as a deletion of aroA, are commonly introduced into such bacteria for attenuation without incapacitating immunostimulation. In this study, we describe the surprising finding that deletion of aroA dramatically increased the virulence of attenuated Salmonella in mouse models. Mutant bacteria lacking aroA elicited increased levels of the proinflammatory cytokine tumor necrosis factor alpha (TNF-α) after systemic application. A detailed genetic and phenotypic characterization in combination with transcriptomic and metabolic profiling demonstrated that ΔaroA mutants display pleiotropic alterations in cellular physiology and lipid and amino acid metabolism, as well as increased sensitivity to penicillin, complement, and phagocytic uptake. In concert with other immunomodulating mutations, deletion of aroA affected flagellin phase variation and gene expression of the virulence-associated genes arnT and ansB. Finally, ΔaroA strains displayed significantly improved tumor therapeutic activity. These results highlight the importance of a functional shikimate pathway to control homeostatic bacterial physiology. They further highlight the great potential of ΔaroA-attenuated Salmonella for the development of vaccines and cancer therapies with important implications for host-pathogen interactions and translational medicine. PMID:27601574

  5. Identification and functional analysis of Salmonella enterica serovar Typhimurium PmrA-regulated genes.

    PubMed

    Tamayo, Rita; Prouty, Angela M; Gunn, John S

    2005-02-01

    The PmrA-PmrB two-component regulatory system of Salmonella enterica serovar Typhimurium is activated in vivo and plays an important role in resistance to cationic antimicrobial peptides. Resistance is partly mediated by modifications to the lipopolysaccharide. To identify new PmrA-regulated genes, microarray analysis was undertaken comparing cDNA derived from PmrA-constitutive and PmrA-null strains. A combination of RT-PCR and transcriptional analysis confirmed the inclusion of six new loci in the PmrA-PmrB regulon: STM1253, STM1269, STM4118, STM0459, STM3968 and STM4568. These loci did not affect the ability to grow in high iron conditions, the ability to modify lipid A with aminoarabinose, or virulence. STM4118, a putative phosphoethanolamine phosphotransferase, had a minor effect on polymyxin resistance, whereas the remaining genes had no role in polymyxin resistance. Although several of the identified loci lacked the consensus PmrA binding site, PmrA was demonstrated to bind the promoter of a PmrA-activated gene lacking the consensus site. A more complete definition of the PmrA-PmrB regulon will provide a better understanding of its role in host and non-host environments.

  6. Salmonella enterica Serovar Typhimurium Exploits Inflammation to Compete with the Intestinal Microbiota

    PubMed Central

    Stecher, Bärbel; Westendorf, Astrid M; Barthel, Manja; Kremer, Marcus; Chaffron, Samuel; Macpherson, Andrew J; Buer, Jan; Parkhill, Julian; Dougan, Gordon; von Mering, Christian; Hardt, Wolf-Dietrich

    2007-01-01

    Most mucosal surfaces of the mammalian body are colonized by microbial communities (“microbiota”). A high density of commensal microbiota inhabits the intestine and shields from infection (“colonization resistance”). The virulence strategies allowing enteropathogenic bacteria to successfully compete with the microbiota and overcome colonization resistance are poorly understood. Here, we investigated manipulation of the intestinal microbiota by the enteropathogenic bacterium Salmonella enterica subspecies 1 serovar Typhimurium (S. Tm) in a mouse colitis model: we found that inflammatory host responses induced by S. Tm changed microbiota composition and suppressed its growth. In contrast to wild-type S. Tm, an avirulent invGsseD mutant failing to trigger colitis was outcompeted by the microbiota. This competitive defect was reverted if inflammation was provided concomitantly by mixed infection with wild-type S. Tm or in mice (IL10−/−, VILLIN-HACL4-CD8) with inflammatory bowel disease. Thus, inflammation is necessary and sufficient for overcoming colonization resistance. This reveals a new concept in infectious disease: in contrast to current thinking, inflammation is not always detrimental for the pathogen. Triggering the host's immune defence can shift the balance between the protective microbiota and the pathogen in favour of the pathogen. PMID:17760501

  7. Mechanism and Fitness Costs of PR-39 Resistance in Salmonella enterica Serovar Typhimurium LT2 ▿

    PubMed Central

    Pränting, Maria; Negrea, Aurel; Rhen, Mikael; Andersson, Dan I.

    2008-01-01

    PR-39 is a porcine antimicrobial peptide that kills bacteria with a mechanism that does not involve cell lysis. Here, we demonstrate that Salmonella enterica serovar Typhimurium can rapidly acquire mutations that reduce susceptibility to PR-39. Resistant mutants appeared at a rate of 0.4 × 10−6 per cell per generation. These mutants were about four times more resistant than the wild type and showed a greatly reduced rate of killing. Genetic analysis revealed mutations in the putative transport protein SbmA as being responsible for the observed resistance. These sbmA mutants were as fit as the wild-type parental strain as measured by growth rates in culture medium and mice and by long-term survival in stationary phase. These results suggest that resistance to certain antimicrobial peptides can rapidly develop without an obvious fitness cost for the bacteria and that resistance development could become a threat to the efficacy of antimicrobial peptides if used in a clinical setting. PMID:18519732

  8. Dietary rice bran promotes resistance to Salmonella enterica serovar Typhimurium colonization in mice.

    PubMed

    Kumar, Ajay; Henderson, Angela; Forster, Genevieve M; Goodyear, Andrew W; Weir, Tiffany L; Leach, Jan E; Dow, Steven W; Ryan, Elizabeth P

    2012-07-04

    Dietary rice bran consists of many bioactive components with disease fighting properties; including the capacity to modulate the gut microbiota. Studies point to the important roles of the gut microbiota and the mucosal epithelium in the establishment of protection against enteric pathogens, such as Salmonella. The ability of rice bran to reduce the susceptibility of mice to a Salmonella infection has not been previously investigated. Therefore, we hypothesized that the incorporation of rice bran into the diet would inhibit the colonization of Salmonella in mice through the induction of protective mucosal responses. Mice were fed diets containing 0%, 10% and 20% rice bran for one week prior to being orally infected with Salmonella enterica serovar Typhimurium. We found that mice consuming the 10 and 20% rice bran diets exhibited a reduction in Salmonella fecal shedding for up to nine days post-infection as compared to control diet fed animals (p < 0.05). In addition, we observed decreased concentrations of the pro-inflammatory cytokines, TNF-alpha, IFN-gamma, and IL-12 (p < 0.05) as well as increased colonization of native Lactobacillus spp. in rice bran fed mice (p < 0.05). Furthermore, in vitro experiments revealed the ability of rice bran extracts to reduce Salmonella entry into mouse small intestinal epithelial cells. Increasing rice bran consumption represents a novel dietary means for reducing susceptibility to enteric infection with Salmonella and potentially via induction of native Lactobacillus spp.

  9. Nifedipine Affects the Course of Salmonella enterica Serovar Typhimurium Infection by Modulating Macrophage Iron Homeostasis

    PubMed Central

    Mair, Sabine M.; Nairz, Manfred; Bellmann-Weiler, Rosa; Muehlbacher, Thomas; Schroll, Andrea; Theurl, Igor; Moser, Patrizia L.; Talasz, Heribert; Fang, Ferric C.

    2011-01-01

    Background. Iron overload can adversely influence the course of infection by increasing microbial replication and suppressing antimicrobial immune effector pathways. Recently, we have shown that the calcium channel blocker nifedipine can mobilize tissue iron in mouse models of iron overload. We therefore investigated whether nifedipine treatment affects the course of infection with intracellular bacteria via modulation of iron homeostasis. Methods. The effect of nifedipine on intramacrophage replication of bacteria and modulation of cellular iron homeostasis was investigated in the murine macrophage cell line RAW264.7, and the impact of nifedipine treatment on the course of systemic infection was investigated in C57BL/6 mice in vivo. Results. In RAW264.7 cells, nifedipine treatment significantly reduced intracellular bacterial survival of Salmonella enterica serovar Typhimurium and Chlamydophila pneumoniae. This could be attributed to the induction of the iron exporter ferroportin 1, which limited the availability of iron for intracellular Salmonella. When C57BL/6 mice were infected intraperitoneally with Salmonella and subsequently injected with nifedipine for 3 consecutive days, bacterial counts in livers and spleens were significantly reduced and survival of the mice significantly was prolonged compared with solvent-treated littermates. Nifedipine treatment increased expression of ferroportin 1 in the spleen, whereas splenic levels of the iron storage protein ferritin and serum iron concentrations were reduced. Conclusions. Our data provide evidence for a novel mechanism whereby nifedipine enhances host resistance to intracellular pathogens via limitation of iron availability. PMID:21844295

  10. Identification and characterization of outer membrane vesicle-associated proteins in Salmonella enterica serovar Typhimurium.

    PubMed

    Bai, Jaewoo; Kim, Seul I; Ryu, Sangryeol; Yoon, Hyunjin

    2014-10-01

    Salmonella enterica serovar Typhimurium is a primary cause of enteric diseases and has acquired a variety of virulence factors during its evolution into a pathogen. Secreted virulence factors interact with commensal flora and host cells and enable Salmonella to survive and thrive in hostile environments. Outer membrane vesicles (OMVs) released from many Gram-negative bacteria function as a mechanism for the secretion of complex mixtures, including virulence factors. We performed a proteomic analysis of OMVs that were isolated under standard laboratory and acidic minimal medium conditions and identified 14 OMV-associated proteins that were observed in the OMV fraction isolated only under the acidic minimal medium conditions, which reproduced the nutrient-deficient intracellular milieu. The inferred roles of these 14 proteins were diverse, including transporter, enzyme, and transcriptional regulator. The absence of these proteins influenced Salmonella survival inside murine macrophages. Eleven of these proteins were predicted to possess secretion signal sequences at their N termini, and three (HupA, GlnH, and PhoN) of the proteins were found to be translocated into the cytoplasm of host cells. The comparative proteomic profiling of OMVs performed in this study revealed different protein compositions in the OMVs isolated under the two different conditions, which indicates that the OMV cargo depends on the growth conditions and provides a deeper insight into how Salmonella utilizes OMVs to adapt to environmental changes. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  11. Characterization of Salmonella enterica serovar typhimurium from marine environments in coastal waters of Galicia (Spain).

    PubMed

    Martinez-Urtaza, Jaime; Liebana, Ernesto; Garcia-Migura, Lourdes; Perez-Piñeiro, Pelayo; Saco, Montserrat

    2004-07-01

    Twenty-three Salmonella enterica serovar Typhimurium isolates from marine environments were characterized by phage typing, pulsed-field gel electrophoresis (PFGE) analysis, plasmid analysis, and antibiotic resistance, and the distribution of the different types in the coastal waters were subsequently analyzed. Five phage types were identified among the isolates (PT41, PT135, PT99, DT104, and DT193). PT135 isolates were exclusively detected during the winter months from 1998 to 2000, whereas DT104 and PT41 isolates were detected exclusively in the summer months from 2000 to 2002. XbaI PFGE analysis revealed 9 PFGE types, and plasmid profiling identified 8 plasmid types (with 1 to 6 plasmids) among the isolates. Only three isolates presented multidrug resistance to antibiotics. Two DT104 isolates were resistant to 8 and 7 antibiotics (profiles ACCeFNaSSuT and ACeFNeSSuT), whereas a PT193 isolate presented resistance to 6 antibiotics (profile ACFSSu). In addition, four PT41 isolates were resistant to a single antibiotic. The detection of multidrug-resistant phage types DT104 and DT193 in shellfish emphasizes the importance of monitoring the presence of Salmonella in routine surveillance of live bivalve molluscs.

  12. Characterization of Salmonella enterica Serovar Typhimurium from Marine Environments in Coastal Waters of Galicia (Spain)

    PubMed Central

    Martinez-Urtaza, Jaime; Liebana, Ernesto; Garcia-Migura, Lourdes; Perez-Piñeiro, Pelayo; Saco, Montserrat

    2004-01-01

    Twenty-three Salmonella enterica serovar Typhimurium isolates from marine environments were characterized by phage typing, pulsed-field gel electrophoresis (PFGE) analysis, plasmid analysis, and antibiotic resistance, and the distribution of the different types in the coastal waters were subsequently analyzed. Five phage types were identified among the isolates (PT41, PT135, PT99, DT104, and DT193). PT135 isolates were exclusively detected during the winter months from 1998 to 2000, whereas DT104 and PT41 isolates were detected exclusively in the summer months from 2000 to 2002. XbaI PFGE analysis revealed 9 PFGE types, and plasmid profiling identified 8 plasmid types (with 1 to 6 plasmids) among the isolates. Only three isolates presented multidrug resistance to antibiotics. Two DT104 isolates were resistant to 8 and 7 antibiotics (profiles ACCeFNaSSuT and ACeFNeSSuT), whereas a PT193 isolate presented resistance to 6 antibiotics (profile ACFSSu). In addition, four PT41 isolates were resistant to a single antibiotic. The detection of multidrug-resistant phage types DT104 and DT193 in shellfish emphasizes the importance of monitoring the presence of Salmonella in routine surveillance of live bivalve molluscs. PMID:15240279

  13. Optimizing the restored chemotactic behavior of anticancer agent Salmonella enterica serovar Typhimurium VNP20009.

    PubMed

    Broadway, Katherine M; Suh, Seungbeum; Behkam, Bahareh; Scharf, Birgit E

    2017-06-10

    Bacteria, including strains of Salmonella, have been researched and applied as therapeutic cancer agents for centuries. Salmonella are particularly of interest due to their facultative anaerobic nature, facilitating colonization of differentially oxygenated tumor regions. Additionally, Salmonella can be manipulated with relative ease, resulting in the ability to attenuate the pathogen or engineer vectors for drug delivery. It was recently discovered that the anti-cancer Salmonella enterica serovar Typhimurium strain VNP20009 is lacking in chemotactic ability, due to a non-synonymous single nucleotide polymorphism in cheY. Replacing the mutated copy of cheY with the wild-type sequence restored chemotaxis to 70% of the parental strain. We aimed to investigate further if chemotaxis of VNP20009 can be optimized. By restoring the gene msbB in VNP20009 cheY(+), which confers attenuation by lipid A modification, we observed a 9% increase in swimming speed, 13% increase in swim plate performance, 19% increase in microfluidic device partitioning towards the attractant at the optimum concentration gradient, and mitigation of a non-motile cell subpopulation. We conclude that chemotaxis can be enhanced further but at the cost of changing one defining characteristic of VNP20009. A less compromised strain might be needed to employ for investigating bacterial chemotaxis in tumor interactions. Copyright © 2017 Elsevier B.V. All rights reserved.

  14. Salmonella enterica Serovar Typhimurium Skills To Succeed in the Host: Virulence and Regulation

    PubMed Central

    Fàbrega, Anna

    2013-01-01

    SUMMARY Salmonella enterica serovar Typhimurium is a primary enteric pathogen infecting both humans and animals. Infection begins with the ingestion of contaminated food or water so that salmonellae reach the intestinal epithelium and trigger gastrointestinal disease. In some patients the infection spreads upon invasion of the intestinal epithelium, internalization within phagocytes, and subsequent dissemination. In that case, antimicrobial therapy, based on fluoroquinolones and expanded-spectrum cephalosporins as the current drugs of choice, is indicated. To accomplish the pathogenic process, the Salmonella chromosome comprises several virulence mechanisms. The most important virulence genes are those located within the so-called Salmonella pathogenicity islands (SPIs). Thus far, five SPIs have been reported to have a major contribution to pathogenesis. Nonetheless, further virulence traits, such as the pSLT virulence plasmid, adhesins, flagella, and biofilm-related proteins, also contribute to success within the host. Several regulatory mechanisms which synchronize all these elements in order to guarantee bacterial survival have been described. These mechanisms govern the transitions from the different pathogenic stages and drive the pathogen to achieve maximal efficiency inside the host. This review focuses primarily on the virulence armamentarium of this pathogen and the extremely complicated regulatory network controlling its success. PMID:23554419

  15. Genetic Characterization of the Galactitol Utilization Pathway of Salmonella enterica Serovar Typhimurium.

    PubMed

    Nolle, Nicoletta; Felsl, Angela; Heermann, Ralf; Fuchs, Thilo M

    2017-02-15

    Galactitol degradation by salmonellae remains underinvestigated, although this metabolic capability contributes to growth in animals (R. R. Chaudhuri et al., PLoS Genet 9:e1003456, 2013, https://doi.org/10.1371/journal.pgen.1003456). The genes responsible for this metabolic capability are part of a 9.6-kb gene cluster that spans from gatY to gatR (STM3253 to STM3262) and encodes a phosphotransferase system, four enzymes, and a transporter of the major facilitator superfamily. Genome comparison revealed the presence of this genetic determinant in nearly all Salmonella strains. The generation time of Salmonella enterica serovar Typhimurium strain ST4/74 was higher in minimal medium with galactitol than with glucose. Knockout of STM3254 and gatC resulted in a growth-deficient phenotype of S Typhimurium, with galactitol as the sole carbon source. Partial deletion of gatR strongly reduced the lag phase of growth with galactitol, whereas strains overproducing GatR exhibited a near-zero growth phenotype. Luciferase reporter assays demonstrated strong induction of the gatY and gatZ promoters, which control all genes of this cluster except gatR, in the presence of galactitol but not glucose. Purified GatR bound to these two main gat gene cluster promoters as well as to its own promoter, demonstrating that this autoregulated repressor controls galactitol degradation. Surface plasmon resonance spectroscopy revealed distinct binding properties of GatR toward the three promoters, resulting in a model of differential gat gene expression. The cyclic AMP receptor protein (CRP) bound these promoters with similarly high affinities, and a mutant lacking crp showed severe growth attenuation, demonstrating that galactitol utilization is subject to catabolite repression. Here, we provide the first genetic characterization of galactitol degradation in Salmonella, revealing novel insights into the regulation of this dissimilatory pathway.

  16. The Hyb Hydrogenase Permits Hydrogen-Dependent Respiratory Growth of Salmonella enterica Serovar Typhimurium

    PubMed Central

    Lamichhane-Khadka, Reena; Kwiatkowski, Andrea; Maier, Robert J.

    2010-01-01

    Salmonella enterica serovar Typhimurium contains three distinct respiratory hydrogenases, all of which contribute to virulence. Addition of H2 significantly enhanced the growth rate and yield of S. Typhimurium in an amino acid-containing medium; this occurred with three different terminal respiratory electron acceptors. Based on studies with site-specific double-hydrogenase mutant strains, most of this H2-dependent growth increase was attributed to the Hyb hydrogenase, rather than to the Hya or Hyd respiratory H2-oxidizing enzymes. The wild type strain with H2 had 4.0-fold greater uptake of 14C-labeled amino acids over a period of minutes than did cells incubated without H2. The double-uptake hydrogenase mutant containing only the Hyb hydrogenase transported amino acids H2 dependently like the wild type. The Hyb-only-containing strain produced a membrane potential comparable to that of the wild type. The H2-stimulated amino acid uptake of the wild type and the Hyb-only strain was inhibited by the protonophore carbonyl cyanide m-chlorophenylhydrazone but was less affected by the ATP synthase inhibitor sodium orthovanadate. In the wild type, proteins TonB and ExbD, which are known to couple proton motive force (PMF) to transport processes, were induced by H2 exposure, as were the genes corresponding to these periplasmic PMF-coupling factors. However, studies on tonB and exbD single mutant strains could not confirm a major role for these proteins in amino acid transport. The results link H2 oxidation via the Hyb enzyme to growth, amino acid transport, and expression of periplasmic proteins that facilitate PMF-mediated transport across the outer membrane. PMID:21157514

  17. Analysis of Spleen-Induced Fimbria Production in Recombinant Attenuated Salmonella enterica Serovar Typhimurium Vaccine Strains

    PubMed Central

    Łaniewski, Paweł; Baek, Chang-Ho; Roland, Kenneth L.

    2017-01-01

    ABSTRACT Salmonella enterica serovar Typhimurium genome encodes 13 fimbrial operons. Most of the fimbriae encoded by these operons are not produced under laboratory conditions but are likely to be synthesized in vivo. We used an in vivo expression technology (IVET) strategy to identify four fimbrial operons, agf, saf, sti, and stc that are expressed in the spleen. When any three of these operons were deleted, the strain retained wild-type virulence. However, when all four operons were deleted, the resulting strain was completely attenuated, indicating that these four fimbriae play functionally redundant roles critical for virulence. In mice, oral doses of as low as 1 × 105 CFU of the strain with four fimbrial operons deleted provided 100% protection against challenge with 1 × 109 CFU of wild-type S. Typhimurium. We also examined the possible effect of these fimbriae on the ability of a Salmonella vaccine strain to deliver a guest antigen. We modified one of our established attenuated vaccine strains, χ9088, to delete three fimbrial operons while the fourth operon was constitutively expressed. Each derivative was modified to express the Streptococcus pneumoniae antigen PspA. Strains that constitutively expressed saf or stc elicited a strong Th1 response with significantly greater levels of anti-PspA serum IgG and greater protective efficacy than strains carrying saf or stc deletions. The isogenic strain in which all four operons were deleted generated the lowest anti-PspA levels and did not protect against challenge with virulent S. pneumoniae. Our results indicate that these fimbriae play important roles, as yet not understood, in Salmonella virulence and immunogenicity. PMID:28830946

  18. The ferric enterobactin transporter Fep is required for persistent Salmonella enterica serovar typhimurium infection.

    PubMed

    Nagy, Toni A; Moreland, Sarah M; Andrews-Polymenis, Helene; Detweiler, Corrella S

    2013-11-01

    Most bacterial pathogens require iron to grow and colonize host tissues. The Gram-negative bacterium Salmonella enterica serovar Typhimurium causes a natural systemic infection of mice that models acute and chronic human typhoid fever. S. Typhimurium resides in tissues within cells of the monocyte lineage, which limit pathogen access to iron, a mechanism of nutritional immunity. The primary ferric iron import system encoded by Salmonella is the siderophore ABC transporter FepBDGC. The Fep system has a known role in acute infection, but it is unclear whether ferric iron uptake or the ferric iron binding siderophores enterobactin and salmochelin are required for persistent infection. We defined the role of the Fep iron transporter and siderophores in the replication of Salmonella in macrophages and in mice that develop acute followed by persistent infections. Replication of wild-type and iron transporter mutant Salmonella strains was quantified in cultured macrophages, fecal pellets, and host tissues in mixed- and single-infection experiments. We show that deletion of fepB attenuated Salmonella replication and colonization within macrophages and mice. Additionally, the genes required to produce and transport enterobactin and salmochelin across the outer membrane receptors, fepA and iroN, are needed for colonization of all tissues examined. However, salmochelin appears to be more important than enterobactin in the colonization of the spleen and liver, both sites of dissemination. Thus, the FepBDGC ferric iron transporter and the siderophores enterobactin and salmochelin are required by Salmonella to evade nutritional immunity in macrophages and cause persistent infection in mice.

  19. Contribution of Target Gene Mutations and Efflux to Decreased Susceptibility of Salmonella enterica Serovar Typhimurium to Fluoroquinolones and Other Antimicrobials▿

    PubMed Central

    Chen, Sheng; Cui, Shenghui; McDermott, Patrick F.; Zhao, Shaohua; White, David G.; Paulsen, Ian; Meng, Jianghong

    2007-01-01

    The mechanisms involved in fluoroquinolone resistance in Salmonella enterica include target alterations and overexpression of efflux pumps. The present study evaluated the role of known and putative multidrug resistance efflux pumps and mutations in topoisomerase genes among laboratory-selected and naturally occurring fluoroquinolone-resistant Salmonella enterica serovar Typhimurium strains. Strains with ciprofloxacin MICs of 0.25, 4, 32, and 256 μg/ml were derived in vitro using serovar Typhimurium S21. These mutants also showed decreased susceptibility or resistance to many nonfluoroquinolone antimicrobials, including tetracycline, chloramphenicol, and several β-lactams. The expression of efflux pump genes acrA, acrB, acrE, acrF, emrB, emrD, and mdlB were substantially increased (≥2-fold) among the fluoroquinolone-resistant mutants. Increased expression was also observed, but to a lesser extent, with three other putative efflux pumps: mdtB (yegN), mdtC (yegO), and emrA among mutants with ciprofloxacin MICs of ≥32 μg/ml. Deletion of acrAB or tolC in S21 and its fluoroquinolone-resistant mutants resulted in increased susceptibility to fluoroquinolones and other tested antimicrobials. In naturally occurring fluoroquinolone-resistant serovar Typhimurium strains, deletion of acrAB or tolC increased fluoroquinolone susceptibility 4-fold, whereas replacement of gyrA double mutations (S83F D87N) with wild-type gyrA increased susceptibility >500-fold. These results indicate that a combination of topoisomerase gene mutations, as well as enhanced antimicrobial efflux, plays a critical role in the development of fluoroquinolone resistance in both laboratory-derived and naturally occurring quinolone-resistant serovar Typhimurium strains. PMID:17043131

  20. Regulation of Salmonella enterica serovar typhimurium invasion genes by csrA.

    PubMed

    Altier, C; Suyemoto, M; Lawhon, S D

    2000-12-01

    Penetration of intestinal epithelial cells by Salmonella enterica serovar Typhimurium requires the expression of invasion genes, found in Salmonella pathogenicity island 1 (SPI1), that encode components of a type III secretion apparatus. These genes are controlled in a complex manner by regulators within SPI1, including HilA and InvF, and those outside SPI1, such as the two-component regulators PhoP/PhoQ and BarA/SirA. We report here that epithelial cell invasion requires the serovar Typhimurium homologue of Escherichia coli csrA, which encodes a regulator that alters the stability of specific mRNA targets. A deletion mutant of csrA was unable to efficiently invade cultured epithelial cells and showed reduced expression of four tested SPI1 genes, hilA, invF, sipC, and prgH. Overexpression of csrA from an induced araBAD promoter also negatively affected the expression of these genes, indicating that CsrA can act as both a positive and a negative regulator of SPI1 genes and suggesting that the bacterium must tightly control the level or activity of CsrA to achieve maximal invasion. We found that CsrA affected hilA, a regulator of the other three genes we tested, probably by controlling one or more genetic elements that regulate hilA. We also found that both the loss and the overexpression of csrA reduced the expression of two regulators of hilA, hilC and hilD, suggesting that csrA exerts its control of hilA through one or both of these regulators. We further found, however, that CsrA could affect the expression of both invF and sipC independent of its effects on hilA. One additional striking phenotype of the csrA mutant, not observed in a comparable E. coli mutant, was its slow growth. Phenotypic revertants that had normal growth rates, while maintaining the csrA mutation, were common. These suppressed strains, however, did not recover the ability to invade cultured cells, indicating that the csrA-mediated loss of invasion cannot be attributed simply to poor growth

  1. Interactions of Salmonella enterica subspecies enterica serovar Typhimurium with gut bacteria.

    PubMed

    Avendaño-Pérez, Gaspar; Nueno-Palop, Carmen; Narbad, Arjan; George, Susan M; Baranyi, József; Pin, Carmen

    2015-06-01

    The aim of this study was to evaluate the impact of the gut microbiota on the growth and survival of S. Typhimurium. This was tested in two-species co-cultures and in mixed cultures with a simplified gut model microbiota. Subsequently, interactions between S. Typhimurium and human faecal bacteria were quantified in both batch and continuous culture systems simulating the human colon. The exponential growth of S. Typhimurium was halted when the population of Escherichia coli reached the maximum population density in a two-compartment co-culture system where the two species were separated by a 0.45 μm pore membrane. Furthermore, the growth of some gut bacteria such as Lactobacillus gasseri and Bifidobacterium bifidum was inhibited by the presence of S. Typhimurium in the other compartment. The survival of S. Typhimurium was severely affected in mixed batch cultures with human faecal samples; a reduction of 10(3)-10(4) cfu/ml in the concentration of S. Typhimurium was observed in these cultures. However, no effect on S. Typhimurium survival was observed in mixed batch cultures with a simplified gut model microbiota under the same conditions. The effect of human faecal samples on S. Typhimurium in a three-stage continuous culture was different to that obtained in batch cultures; its growth rather than survival was affected under these conditions. S. Typhimurium growth was inhibited, and the bacterium was therefore eliminated by the continuous flow of the medium. Depending upon culturing conditions, the gut microbiota caused either growth inhibition, inactivation or did not affect S. Typhimurium.

  2. Integrative analysis of Salmonellosis in Israel reveals association of Salmonella enterica Serovar 9,12:l,v:- with extraintestinal infections, dissemination of endemic S. enterica Serovar Typhimurium DT104 biotypes, and severe underreporting of outbreaks.

    PubMed

    Marzel, Alex; Desai, Prerak T; Nissan, Israel; Schorr, Yosef Ilan; Suez, Jotham; Valinsky, Lea; Reisfeld, Abraham; Agmon, Vered; Guard, Jean; McClelland, Michael; Rahav, Galia; Gal-Mor, Ohad

    2014-06-01

    Salmonella enterica is the leading etiologic agent of bacterial food-borne outbreaks worldwide. This ubiquitous species contains more than 2,600 serovars that may differ in their host specificity, clinical manifestations, and epidemiology. To characterize salmonellosis epidemiology in Israel and to study the association of nontyphoidal Salmonella (NTS) serovars with invasive infections, 48,345 Salmonella cases reported and serotyped at the National Salmonella Reference Center between 1995 and 2012 were analyzed. A quasi-Poisson regression was used to identify irregular clusters of illness, and pulsed-field gel electrophoresis in conjunction with whole-genome sequencing was applied to molecularly characterize strains of interest. Three hundred twenty-nine human salmonellosis clusters were identified, representing an annual average of 23 (95% confidence interval [CI], 20 to 26) potential outbreaks. We show that the previously unsequenced S. enterica serovar 9,12:l,v:- belongs to the B clade of Salmonella enterica subspecies enterica, and we show its frequent association with extraintestinal infections, compared to other NTS serovars. Furthermore, we identified the dissemination of two prevalent Salmonella enterica serovar Typhimurium DT104 clones in Israel, which are genetically distinct from other global DT104 isolates. Accumulatively, these findings indicate a severe underreporting of Salmonella outbreaks in Israel and provide insights into the epidemiology and genomics of prevalent serovars, responsible for recurring illness.

  3. Salmonella enterica serovar Typhimurium BaeSR two-component system positively regulates sodA in response to ciprofloxacin

    PubMed Central

    Guerrero, P.; Collao, B.; Álvarez, R.; Salinas, H.; Morales, E. H.; Calderón, I. L.; Saavedra, C. P.

    2013-01-01

    In response to antibiotics, bacteria activate regulatory systems that control the expression of genes that participate in detoxifying these compounds, like multidrug efflux systems. We previously demonstrated that the BaeSR two-component system from Salmonella enterica serovar Typhimurium (S. Typhimurium) participates in the detection of ciprofloxacin, a bactericidal antibiotic, and in the positive regulation of mdtA, an efflux pump implicated in antibiotic resistance. In the present work, we provide further evidence for a role of the S. Typhimurium BaeSR two-component system in response to ciprofloxacin treatment and show that it regulates sodA expression. We demonstrate that, in the absence of BaeSR, the transcript levels of sodA and the activity of its gene product are lower. Using electrophoretic mobility shift assays and transcriptional fusions, we demonstrate that BaeR regulates sodA by a direct interaction with the promoter region. PMID:23918818

  4. Salmonella enterica serovar Typhimurium BaeSR two-component system positively regulates sodA in response to ciprofloxacin.

    PubMed

    Guerrero, P; Collao, B; Álvarez, R; Salinas, H; Morales, E H; Calderón, I L; Saavedra, C P; Gil, F

    2013-10-01

    In response to antibiotics, bacteria activate regulatory systems that control the expression of genes that participate in detoxifying these compounds, like multidrug efflux systems. We previously demonstrated that the BaeSR two-component system from Salmonella enterica serovar Typhimurium (S. Typhimurium) participates in the detection of ciprofloxacin, a bactericidal antibiotic, and in the positive regulation of mdtA, an efflux pump implicated in antibiotic resistance. In the present work, we provide further evidence for a role of the S. Typhimurium BaeSR two-component system in response to ciprofloxacin treatment and show that it regulates sodA expression. We demonstrate that, in the absence of BaeSR, the transcript levels of sodA and the activity of its gene product are lower. Using electrophoretic mobility shift assays and transcriptional fusions, we demonstrate that BaeR regulates sodA by a direct interaction with the promoter region.

  5. Oral administration of Salmonella enterica serovar Typhimurium expressing swine interleukin-18 induces Th1-biased protective immunity against inactivated vaccine of pseudorabies virus.

    PubMed

    Kim, Seong Bum; Kim, Seon Ju; Lee, Byung Min; Han, Young Woo; Rahman, Md Masudur; Uyangaa, Erdenebileg; Kim, Jin Hyoung; Choi, Jin Young; Yoo, Dong Jin; Kim, Koanhoi; Eo, Seong Kug

    2012-03-23

    Enhancing and/or modulating innate and adaptive immunity by cytokines appears to be greatly useful to provide effective protective immunity against infectious diseases. However, an effective delivery system for mass administration in livestock industry is needed because of limitations such as cost, labor, time, and protein stability. Here the immunomodulatory functions of swine interleukine-18 (swIL-18), known as IFN-γ-inducing factor (IGIF), were evaluated in a vaccination model of pseudorabies virus (PrV) using attenuated Salmonella enterica serovar Typhimurium as the oral delivery system. The oral administration of S. enterica serovar Typhimurium expressing swIL-18 prior to vaccination with inactivated PrV vaccine induced enhanced levels of serum PrV-specific IgG and its IgG2 isotype, compared to administration of S. enterica serovar Typhimurium harboring the empty vector. Furthermore, S. enterica serovar Typhimurium expressing swIL-18 mounted Th1-biased cellular immune responses against PrV antigen, as evaluated by the production of IFN-γ and IL-4 from peripheral blood mononuclear cells of piglets. Subsequently, Th1-biased immunity induced by S. enterica serovar Typhimurium expressing swIL-18 showed rapid response and rendered piglets displayed more alleviated clinical signs following the virulent PrV challenge. Also, this alleviation of clinical signs was further confirmed by the reduction of nasal excretion of PrV after challenge. The present study demonstrates the extended use of immunomodulatory functions of swIL-18 orally delivered by attenuated S. enterica serovar Typhimurium. Copyright © 2011 Elsevier B.V. All rights reserved.

  6. Effects of indole on drug resistance and virulence of Salmonella enterica serovar Typhimurium revealed by genome-wide analyses

    PubMed Central

    2012-01-01

    Background Many Gram-positive and Gram-negative bacteria produce large quantities of indole as an intercellular signal in microbial communities. Indole demonstrated to affect gene expression in Escherichia coli as an intra-species signaling molecule. In contrast to E. coli, Salmonella does not produce indole because it does not harbor tnaA, which encodes the enzyme responsible for tryptophan metabolism. Our previous study demonstrated that E. coli-conditioned medium and indole induce expression of the AcrAB multidrug efflux pump in Salmonella enterica serovar Typhimurium for inter-species communication; however, the global effect of indole on genes in Salmonella remains unknown. Results To understand the complete picture of genes regulated by indole, we performed DNA microarray analysis of genes in the S. enterica serovar Typhimurium strain ATCC 14028s affected by indole. Predicted Salmonella phenotypes affected by indole based on the microarray data were also examined in this study. Indole induced expression of genes related to efflux-mediated multidrug resistance, including ramA and acrAB, and repressed those related to host cell invasion encoded in the Salmonella pathogenicity island 1, and flagella production. Reduction of invasive activity and motility of Salmonella by indole was also observed phenotypically. Conclusion Our results suggest that indole is an important signaling molecule for inter-species communication to control drug resistance and virulence of S. enterica. PMID:22632036

  7. Epidemiology of plasmid-mediated quinolone resistance in salmonella enterica serovar typhimurium isolates from food-producing animals in Japan

    PubMed Central

    2010-01-01

    A total of 225 isolates of Salmonella enterica serovar Typhimurium from food-producing animals collected between 2003 and 2007 were examined for the prevalence of plasmid-mediated quinolone resistance (PMQR) determinants, namely qnrA, qnrB, qnrC, qnrD, qnrS, qepA and aac(6')Ib-cr, in Japan. Two isolates (0.8%) of S. Typhimurium DT104 from different dairy cows on a single farm in 2006 and 2007 were found to have qnrS1 on a plasmid of approximately 9.6-kbp. None of the S. Typhimurium isolates had qnrA, qnrB, qnrC, qnrD, qepA and acc(6')-Ib-cr. Currently in Japan, the prevalence of the PMQR genes among S. Typhimurium isolates from food animals may remain low or restricted. The PFGE profile of two S. Typhimurium DT104 isolates without qnrS1 on the farm in 2005 had an identical PFGE profile to those of two S. Typhimurium DT104 isolates with qnrS1. The PFGE analysis suggested that the already existing S. Typhimurium DT104 on the farm fortuitously acquired the qnrS1 plasmid. PMID:21138594

  8. Proteolytic Inhibition of Salmonella enterica Serovar Typhimurium-Induced Activation of the Mitogen-Activated Protein Kinases ERK and JNK in Cultured Human Intestinal Cells

    PubMed Central

    Mynott, Tracey L.; Crossett, Ben; Prathalingam, S. Radhika

    2002-01-01

    Bromelain, a mixture of cysteine proteases from pineapple stems, blocks signaling by the mitogen-activated protein (MAP) kinases extracellular regulated kinase 1 (ERK-1) and ERK-2, inhibits inflammation, and protects against enterotoxigenic Escherichia coli infection. In this study, we examined the effect of bromelain on Salmonella enterica serovar Typhimurium infection, since an important feature of its pathogenesis is its ability to induce activation of ERK-1 and ERK-2, which leads to internalization of bacteria and induction of inflammatory responses. Our results show that bromelain dose dependently blocks serovar Typhimurium-induced ERK-1, ERK-2, and c-Jun NH2-terminal kinase (JNK) activation in Caco-2 cells. Bromelain also blocked signaling induced by carbachol and anisomycin, pharmacological MAP kinase agonists. Despite bromelain inhibition of serovar Typhimurium-induced MAP kinase signaling, it did not prevent subsequent invasion of the Caco-2 cells by serovar Typhimurium or alter serovar Typhimurium -induced decreases in resistance across Caco-2 monolayers. Surprisingly, bromelain also did not block serovar Typhimurium-induced interleukin-8 (IL-8) secretion but synergized with serovar Typhimurium to enhance IL-8 production. We also found that serovar Typhimurium does not induce ERK phosphorylation in Caco-2 cells in the absence of serum but that serovar Typhimurium-induced invasion and decreases in monolayer resistance are unaffected. Collectively, these data indicate that serovar Typhimurium-induced invasion of Caco-2 cells, changes in the resistance of epithelial cell monolayers, and IL-8 production can occur independently of the ERK and JNK signaling pathways. Data also confirm that bromelain is a novel inhibitor of MAP kinase signaling pathways and suggest a novel role for proteases as inhibitors of signal transduction pathways in intestinal epithelial cells. PMID:11748167

  9. Proteolytic inhibition of Salmonella enterica serovar typhimurium-induced activation of the mitogen-activated protein kinases ERK and JNK in cultured human intestinal cells.

    PubMed

    Mynott, Tracey L; Crossett, Ben; Prathalingam, S Radhika

    2002-01-01

    Bromelain, a mixture of cysteine proteases from pineapple stems, blocks signaling by the mitogen-activated protein (MAP) kinases extracellular regulated kinase 1 (ERK-1) and ERK-2, inhibits inflammation, and protects against enterotoxigenic Escherichia coli infection. In this study, we examined the effect of bromelain on Salmonella enterica serovar Typhimurium infection, since an important feature of its pathogenesis is its ability to induce activation of ERK-1 and ERK-2, which leads to internalization of bacteria and induction of inflammatory responses. Our results show that bromelain dose dependently blocks serovar Typhimurium-induced ERK-1, ERK-2, and c-Jun NH(2)-terminal kinase (JNK) activation in Caco-2 cells. Bromelain also blocked signaling induced by carbachol and anisomycin, pharmacological MAP kinase agonists. Despite bromelain inhibition of serovar Typhimurium-induced MAP kinase signaling, it did not prevent subsequent invasion of the Caco-2 cells by serovar Typhimurium or alter serovar Typhimurium -induced decreases in resistance across Caco-2 monolayers. Surprisingly, bromelain also did not block serovar Typhimurium-induced interleukin-8 (IL-8) secretion but synergized with serovar Typhimurium to enhance IL-8 production. We also found that serovar Typhimurium does not induce ERK phosphorylation in Caco-2 cells in the absence of serum but that serovar Typhimurium-induced invasion and decreases in monolayer resistance are unaffected. Collectively, these data indicate that serovar Typhimurium-induced invasion of Caco-2 cells, changes in the resistance of epithelial cell monolayers, and IL-8 production can occur independently of the ERK and JNK signaling pathways. Data also confirm that bromelain is a novel inhibitor of MAP kinase signaling pathways and suggest a novel role for proteases as inhibitors of signal transduction pathways in intestinal epithelial cells.

  10. Three-dimensional tissue assemblies: novel models for the study of Salmonella enterica serovar Typhimurium pathogenesis

    NASA Technical Reports Server (NTRS)

    Nickerson, C. A.; Goodwin, T. J.; Terlonge, J.; Ott, C. M.; Buchanan, K. L.; Uicker, W. C.; Emami, K.; LeBlanc, C. L.; Ramamurthy, R.; Clarke, M. S.; Vanderburg, C. R.; Hammond, T.; Pierson, D. L.

    2001-01-01

    The lack of readily available experimental systems has limited knowledge pertaining to the development of Salmonella-induced gastroenteritis and diarrheal disease in humans. We used a novel low-shear stress cell culture system developed at the National Aeronautics and Space Administration in conjunction with cultivation of three-dimensional (3-D) aggregates of human intestinal tissue to study the infectivity of Salmonella enterica serovar Typhimurium for human intestinal epithelium. Immunohistochemical characterization and microscopic analysis of 3-D aggregates of the human intestinal epithelial cell line Int-407 revealed that the 3-D cells more accurately modeled human in vivo differentiated tissues than did conventional monolayer cultures of the same cells. Results from infectivity studies showed that Salmonella established infection of the 3-D cells in a much different manner than that observed for monolayers. Following the same time course of infection with Salmonella, 3-D Int-407 cells displayed minimal loss of structural integrity compared to that of Int-407 monolayers. Furthermore, Salmonella exhibited significantly lower abilities to adhere to, invade, and induce apoptosis of 3-D Int-407 cells than it did for infected Int-407 monolayers. Analysis of cytokine expression profiles of 3-D Int-407 cells and monolayers following infection with Salmonella revealed significant differences in expression of interleukin 1alpha (IL-1alpha), IL-1beta, IL-6, IL-1Ra, and tumor necrosis factor alpha mRNAs between the two cultures. In addition, uninfected 3-D Int-407 cells constitutively expressed higher levels of transforming growth factor beta1 mRNA and prostaglandin E2 than did uninfected Int-407 monolayers. By more accurately modeling many aspects of human in vivo tissues, the 3-D intestinal cell model generated in this study offers a novel approach for studying microbial infectivity from the perspective of the host-pathogen interaction.

  11. Virulent Salmonella enterica Serovar Typhimurium Evades Adaptive Immunity by Preventing Dendritic Cells from Activating T Cells

    PubMed Central

    Tobar, Jaime A.; Carreño, Leandro J.; Bueno, Susan M.; González, Pablo A.; Mora, Jorge E.; Quezada, Sergio A.; Kalergis, Alexis M.

    2006-01-01

    Dendritic cells (DCs) constitute the link between innate and adaptive immunity by directly recognizing pathogen-associated molecular patterns (PAMPs) in bacteria and by presenting bacterial antigens to T cells. Recognition of PAMPs renders DCs as professional antigen-presenting cells able to prime naïve T cells and initiate adaptive immunity against bacteria. Therefore, interfering with DC function would promote bacterial survival and dissemination. Understanding the molecular mechanisms that have evolved in virulent bacteria to evade activation of adaptive immunity requires the characterization of virulence factors that interfere with DC function. Salmonella enterica serovar Typhimurium, the causative agent of typhoid-like disease in the mouse, can prevent antigen presentation to T cells by avoiding lysosomal degradation in DCs. Here, we show that this feature of virulent Salmonella applies in vivo to prevent activation of adaptive immunity. In addition, this attribute of virulent Salmonella requires functional expression of a type three secretion system (TTSS) and effector proteins encoded within the Salmonella pathogenicity island 2 (SPI-2). In contrast to wild-type virulent Salmonella, mutant strains carrying specific deletions of SPI-2 genes encoding TTSS components or effectors proteins are targeted to lysosomes and are no longer able to prevent DCs from activating T cells in vitro or in vivo. SPI-2 mutant strains are attenuated in vivo, showing reduced tissue colonization and enhanced T-cell activation, which confers protection against a challenge with wild-type virulent Salmonella. Our data suggest that impairment of DC function by the activity of SPI-2 gene products is crucial for Salmonella pathogenesis. PMID:17057096

  12. Three-dimensional tissue assemblies: novel models for the study of Salmonella enterica serovar Typhimurium pathogenesis

    NASA Technical Reports Server (NTRS)

    Nickerson, C. A.; Goodwin, T. J.; Terlonge, J.; Ott, C. M.; Buchanan, K. L.; Uicker, W. C.; Emami, K.; LeBlanc, C. L.; Ramamurthy, R.; Clarke, M. S.; hide

    2001-01-01

    The lack of readily available experimental systems has limited knowledge pertaining to the development of Salmonella-induced gastroenteritis and diarrheal disease in humans. We used a novel low-shear stress cell culture system developed at the National Aeronautics and Space Administration in conjunction with cultivation of three-dimensional (3-D) aggregates of human intestinal tissue to study the infectivity of Salmonella enterica serovar Typhimurium for human intestinal epithelium. Immunohistochemical characterization and microscopic analysis of 3-D aggregates of the human intestinal epithelial cell line Int-407 revealed that the 3-D cells more accurately modeled human in vivo differentiated tissues than did conventional monolayer cultures of the same cells. Results from infectivity studies showed that Salmonella established infection of the 3-D cells in a much different manner than that observed for monolayers. Following the same time course of infection with Salmonella, 3-D Int-407 cells displayed minimal loss of structural integrity compared to that of Int-407 monolayers. Furthermore, Salmonella exhibited significantly lower abilities to adhere to, invade, and induce apoptosis of 3-D Int-407 cells than it did for infected Int-407 monolayers. Analysis of cytokine expression profiles of 3-D Int-407 cells and monolayers following infection with Salmonella revealed significant differences in expression of interleukin 1alpha (IL-1alpha), IL-1beta, IL-6, IL-1Ra, and tumor necrosis factor alpha mRNAs between the two cultures. In addition, uninfected 3-D Int-407 cells constitutively expressed higher levels of transforming growth factor beta1 mRNA and prostaglandin E2 than did uninfected Int-407 monolayers. By more accurately modeling many aspects of human in vivo tissues, the 3-D intestinal cell model generated in this study offers a novel approach for studying microbial infectivity from the perspective of the host-pathogen interaction.

  13. Hydrogen-Stimulated Carbon Acquisition and Conservation in Salmonella enterica Serovar Typhimurium ▿ §

    PubMed Central

    Lamichhane-Khadka, Reena; Frye, Jonathan G.; Porwollik, Steffen; McClelland, Michael; Maier, Robert J.

    2011-01-01

    Salmonella enterica serovar Typhimurium can utilize molecular hydrogen for growth and amino acid transport during anaerobic growth. Via microarray we identified H2 gas-affected gene expression changes in Salmonella. The addition of H2 caused altered expression of 597 genes, of which 176 genes were upregulated and 421 were downregulated. The significantly H2-upregulated genes include those that encode proteins involved in the transport of iron, manganese, amino acids, nucleosides, and sugars. Genes encoding isocitrate lyase (aceA) and malate synthase (aceB), both involved in the carbon conserving glyoxylate pathway, and genes encoding the enzymes of the d-glucarate and d-glycerate pathways (gudT, gudD, garR, garL, garK) are significantly upregulated by H2. Cells grown with H2 showed markedly increased AceA enzyme activity compared to cells without H2. Mutant strains with deletion of either aceA or aceB had reduced H2-dependent growth rates. Genes encoding the glutamine-specific transporters (glnH, glnP, glnQ) were upregulated by H2, and cells grown with H2 showed increased [14C]glutamine uptake. Similarly, the mannose uptake system genes (manX, manY) were upregulated by H2, and cells grown with H2 showed about 2.0-fold-increased [14C]d-mannose uptake compared to the cells grown without H2. Hydrogen stimulates the expression of genes involved in nutrient and carbon acquisition and carbon-conserving pathways, linking carbon and energy metabolism to sustain H2-dependent growth. PMID:21856852

  14. Genomic Comparison of Non-Typhoidal Salmonella enterica Serovars Typhimurium, Enteritidis, Heidelberg, Hadar and Kentucky Isolates from Broiler Chickens.

    PubMed

    Dhanani, Akhilesh S; Block, Glenn; Dewar, Ken; Forgetta, Vincenzo; Topp, Edward; Beiko, Robert G; Diarra, Moussa S

    2015-01-01

    Non-typhoidal Salmonella enterica serovars, associated with different foods including poultry products, are important causes of bacterial gastroenteritis worldwide. The colonization of the chicken gut by S. enterica could result in the contamination of the environment and food chain. The aim of this study was to compare the genomes of 25 S. enterica serovars isolated from broiler chicken farms to assess their intra- and inter-genetic variability, with a focus on virulence and antibiotic resistance characteristics. The genomes of 25 S. enterica isolates covering five serovars (ten Typhimurium including three monophasic 4,[5],12:i:, four Enteritidis, three Hadar, four Heidelberg and four Kentucky) were sequenced. Most serovars were clustered in strongly supported phylogenetic clades, except for isolates of serovar Enteritidis that were scattered throughout the tree. Plasmids of varying sizes were detected in several isolates independently of serovars. Genes associated with the IncF plasmid and the IncI1 plasmid were identified in twelve and four isolates, respectively, while genes associated with the IncQ plasmid were found in one isolate. The presence of numerous genes associated with Salmonella pathogenicity islands (SPIs) was also confirmed. Components of the type III and IV secretion systems (T3SS and T4SS) varied in different isolates, which could explain in part, differences of their pathogenicity in humans and/or persistence in broilers. Conserved clusters of genes in the T3SS were detected that could be used in designing effective strategies (diagnostic, vaccination or treatments) to combat Salmonella. Antibiotic resistance genes (CMY, aadA, ampC, florR, sul1, sulI, tetAB, and srtA) and class I integrons were detected in resistant isolates while all isolates carried multidrug efflux pump systems regardless of their antibiotic susceptibility profile. This study showed that the predominant Salmonella serovars in broiler chickens harbor genes encoding adhesins

  15. Genomic Comparison of Non-Typhoidal Salmonella enterica Serovars Typhimurium, Enteritidis, Heidelberg, Hadar and Kentucky Isolates from Broiler Chickens

    PubMed Central

    Dhanani, Akhilesh S.; Block, Glenn; Dewar, Ken; Forgetta, Vincenzo; Topp, Edward; Beiko, Robert G.; Diarra, Moussa S.

    2015-01-01

    Background Non-typhoidal Salmonella enterica serovars, associated with different foods including poultry products, are important causes of bacterial gastroenteritis worldwide. The colonization of the chicken gut by S. enterica could result in the contamination of the environment and food chain. The aim of this study was to compare the genomes of 25 S. enterica serovars isolated from broiler chicken farms to assess their intra- and inter-genetic variability, with a focus on virulence and antibiotic resistance characteristics. Methodology/Principal Finding The genomes of 25 S. enterica isolates covering five serovars (ten Typhimurium including three monophasic 4,[5],12:i:, four Enteritidis, three Hadar, four Heidelberg and four Kentucky) were sequenced. Most serovars were clustered in strongly supported phylogenetic clades, except for isolates of serovar Enteritidis that were scattered throughout the tree. Plasmids of varying sizes were detected in several isolates independently of serovars. Genes associated with the IncF plasmid and the IncI1 plasmid were identified in twelve and four isolates, respectively, while genes associated with the IncQ plasmid were found in one isolate. The presence of numerous genes associated with Salmonella pathogenicity islands (SPIs) was also confirmed. Components of the type III and IV secretion systems (T3SS and T4SS) varied in different isolates, which could explain in part, differences of their pathogenicity in humans and/or persistence in broilers. Conserved clusters of genes in the T3SS were detected that could be used in designing effective strategies (diagnostic, vaccination or treatments) to combat Salmonella. Antibiotic resistance genes (CMY, aadA, ampC, florR, sul1, sulI, tetAB, and srtA) and class I integrons were detected in resistant isolates while all isolates carried multidrug efflux pump systems regardless of their antibiotic susceptibility profile. Conclusions/Significance This study showed that the predominant

  16. Contribution of the PhoP/Q regulon to survival and replication of Salmonella enterica serovar Typhimurium in macrophages

    PubMed Central

    Thompson, Jessica A.; Liu, Mei

    2011-01-01

    The ability of serovars of Salmonella enterica to cause systemic disease is dependent upon their survival and replication within macrophages. To do this, bacteria must withstand or surmount bacteriostatic and bactericidal responses by the host cell, including the delivery of hydrolytic enzymes from lysosomes to the phagosome. The bacterial two-component regulatory system PhoP/Q has been implicated in avoidance of phagolysosomal fusion by S. enterica serovar Typhimurium (S. Typhimurium) in murine macrophages. In this study, the involvement of PhoP/Q-activated genes in avoidance of phagolysosomal fusion was analysed: of all the S. Typhimurium mutant strains tested, only an mgtC mutant strain partially reproduced the phenotype of the phoP mutant strain. As this gene is required for bacterial growth in magnesium-depleted conditions in vitro, the contributions of PhoP/Q to intramacrophage replication and survival were reappraised. Although PhoP/Q was required for both replication and survival of S. Typhimurium within murine macrophages, subsequent analysis of the kinetics of phagolysosomal fusion, taking account of differences in the replication rates of wild-type and phoP mutant strains, provided no evidence for a PhoP/Q-dependent role in this process. PhoP/Q appeared to act subsequent to the process of phagolysosomal avoidance and to promote replication of those bacteria that had already escaped a phagolysosomal fate. Therefore, we conclude that the PhoP/Q regulon enables S. Typhimurium to adapt to intramacrophage stresses other than phagolysosomal fusion. PMID:21511762

  17. A murine model to study the antibacterial effect of copper on infectivity of Salmonella enterica serovar Typhimurium.

    PubMed

    Sharan, Riti; Chhibber, Sanjay; Reed, Robert H

    2011-01-01

    This study investigated the effect of copper as an antibacterial agent on the infectivity of Salmonella enterica serovar Typhimurium. Mice were infected orally with a standardized dose of unstressed Salmonella Typhimurium and copper-stressed cells of Salmonella Typhimurium. Bacterial counts in ileum, blood, liver and spleen were observed up to 168 h under normal aerobic conditions. Serum sensitivity, phagocytosis, malondialdehyde levels and histopathology were studied for both set of animals. A decreased bacterial count in the organs with mild symptoms of infection and a complete recovery by 48 h was observed in mice infected with copper-stressed bacteria. Histopathological examination of ileum tissue demonstrated regeneration of damaged tissue post-infection with copper-stressed bacteria and no malondialdehyde levels were detected after 24 h in ileum, spleen and liver. Exposure to copper sensitized Salmonella Typhimurium to the lytic action of serum and intracellular killing by peritoneal macrophages. It can be concluded that copper stress confers a decrease in the infectivity of healthy Salmonella Typhimurium in normal mice. This study highlights the significance of use of copper as an antibacterial agent against Salmonella Typhimurium in reducing the risk of incidence of Salmonella infections from contaminated water.

  18. Genomic diversity and adaptation of Salmonella enterica serovar Typhimurium from analysis of six genomes of different phage types

    PubMed Central

    2013-01-01

    Background Salmonella enterica serovar Typhimurium (or simply Typhimurium) is the most common serovar in both human infections and farm animals in Australia and many other countries. Typhimurium is a broad host range serovar but has also evolved into host-adapted variants (i.e. isolated from a particular host such as pigeons). Six Typhimurium strains of different phage types (defined by patterns of susceptibility to lysis by a set of bacteriophages) were analysed using Illumina high-throughput genome sequencing. Results Variations between strains were mainly due to single nucleotide polymorphisms (SNPs) with an average of 611 SNPs per strain, ranging from 391 SNPs to 922 SNPs. There were seven insertions/deletions (indels) involving whole or partial gene deletions, four inactivation events due to IS200 insertion and 15 pseudogenes due to early termination. Four of these inactivated or deleted genes may be virulence related. Nine prophage or prophage remnants were identified in the six strains. Gifsy-1, Gifsy-2 and the sopE2 and sspH2 phage remnants were present in all six genomes while Fels-1, Fels-2, ST64B, ST104 and CP4-57 were variably present. Four strains carried the 90-kb plasmid pSLT which contains several known virulence genes. However, two strains were found to lack the plasmid. In addition, one strain had a novel plasmid similar to Typhi strain CT18 plasmid pHCM2. Conclusion The genome data suggest that variations between strains were mainly due to accumulation of SNPs, some of which resulted in gene inactivation. Unique genetic elements that were common between host-adapted phage types were not found. This study advanced our understanding on the evolution and adaptation of Typhimurium at genomic level. PMID:24138507

  19. Osmoprotection of Salmonella enterica serovar Typhimurium by Ngamma-acetyldiaminobutyrate, the precursor of the compatible solute ectoine.

    PubMed

    García-Estepa, Raul; Cánovas, David; Iglesias-Guerra, Fernando; Ventosa, Antonio; Csonka, Laszlo N; Nieto, Joaquín J; Vargas, Carmen

    2006-12-01

    N(gamma)-acetyl-2,4-diaminobutyrate (NADA), the precursor of the compatible solute ectoine, was shown to function as an osmoprotectant for the non-halophilic bacterium Salmonella enterica serovar Typhimurium. The addition of NADA-containing extracts of an ectoine synthase mutant of the broad salt-growing halophile Chromohalobacter salexigens DSM 3043(T) could alleviate the inhibitory effects of high salinity in S. enterica, which lacks the ectoine biosynthetic pathway. NADA, purified from extracts of the mutant, protected S. enterica against salinity stress. This osmoprotective effect was slightly lower than that of ectoine, but more potent than that of hydroxyectoine. Accumulation of purified NADA by S. enterica was demonstrated by (13)C-NMR spectroscopy and HPLC analysis. In addition, it was shown that NADA was taken up by S. enterica via the ProP and ProU transport systems, which are known to transport glycine betaine and proline. This finding provides evidence that these permeases can recognize a diaminoacid that carries an unsubstituted alpha-amino group. This is the first time that NADA has been connected with osmoprotective functions in non-halophilic bacteria.

  20. Caveolin-1-Deficient Mice Show Defects in Innate Immunity and Inflammatory Immune Response during Salmonella enterica Serovar Typhimurium Infection▿ †

    PubMed Central

    Medina, Freddy A.; de Almeida, Cecilia J.; Dew, Elliott; Li, Jiangwei; Bonuccelli, Gloria; Williams, Terence M.; Cohen, Alex W.; Pestell, Richard G.; Frank, Philippe G.; Tanowitz, Herbert B.; Lisanti, Michael P.

    2006-01-01

    A number of studies have shown an association of pathogens with caveolae. To this date, however, there are no studies showing a role for caveolin-1 in modulating immune responses against pathogens. Interestingly, expression of caveolin-1 has been shown to occur in a regulated manner in immune cells in response to lipopolysaccharide (LPS). Here, we sought to determine the role of caveolin-1 (Cav-1) expression in Salmonella pathogenesis. Cav-1−/− mice displayed a significant decrease in survival when challenged with Salmonella enterica serovar Typhimurium. Spleen and tissue burdens were significantly higher in Cav-1−/− mice. However, infection of Cav-1−/− macrophages with serovar Typhimurium did not result in differences in bacterial invasion. In addition, Cav-1−/− mice displayed increased production of inflammatory cytokines, chemokines, and nitric oxide. Regardless of this, Cav-1−/− mice were unable to control the systemic infection of Salmonella. The increased chemokine production in Cav-1−/− mice resulted in greater infiltration of neutrophils into granulomas but did not alter the number of granulomas present. This was accompanied by increased necrosis in the liver. However, Cav-1−/− macrophages displayed increased inflammatory responses and increased nitric oxide production in vitro in response to Salmonella LPS. These results show that caveolin-1 plays a key role in regulating anti-inflammatory responses in macrophages. Taken together, these data suggest that the increased production of toxic mediators from macrophages lacking caveolin-1 is likely to be responsible for the marked susceptibility of caveolin-1-deficient mice to S. enterica serovar Typhimurium. PMID:16982844

  1. Genomic Analysis of Salmonella enterica Serovar Typhimurium DT160 Associated with a 14-Year Outbreak, New Zealand, 1998-2012.

    PubMed

    Bloomfield, Samuel J; Benschop, Jackie; Biggs, Patrick J; Marshall, Jonathan C; Hayman, David T S; Carter, Philip E; Midwinter, Anne C; Mather, Alison E; French, Nigel P

    2017-06-01

    During 1998-2012, an extended outbreak of Salmonella enterica serovar Typhimurium definitive type 160 (DT160) affected >3,000 humans and killed wild birds in New Zealand. However, the relationship between DT160 within these 2 host groups and the origin of the outbreak are unknown. Whole-genome sequencing was used to compare 109 Salmonella Typhimurium DT160 isolates from sources throughout New Zealand. We provide evidence that DT160 was introduced into New Zealand around 1997 and rapidly propagated throughout the country, becoming more genetically diverse over time. The genetic heterogeneity was evenly distributed across multiple predicted functional protein groups, and we found no evidence of host group differentiation between isolates collected from human, poultry, bovid, and wild bird sources, indicating ongoing transmission between these host groups. Our findings demonstrate how a comparative genomic approach can be used to gain insight into outbreaks, disease transmission, and the evolution of a multihost pathogen after a probable point-source introduction.

  2. Expression of cspH, encoding the cold shock protein in Salmonella enterica serovar Typhimurium UK-1.

    PubMed

    Kim, B H; Bang, I S; Lee, S Y; Hong, S K; Bang, S H; Lee, I S; Park, Y K

    2001-10-01

    Both Salmonella enterica serovar Typhimurium and Escherichia coli contain the cspH gene encoding CspH, one of the cold shock proteins (CSPs). In this study, we investigated the expression of cspH in S. enterica serovar Typhimurium and found that it was induced in response to a temperature downshift during exponential phase. The cspH promoter was activated at 37 degrees C, and its mRNA was more stable than the other csp mRNAs at 37 degrees C. Moreover, lacZ expression of the translational cspH-lacZ fusion was induced at that temperature. Interestingly, the cspH mRNA had a much shorter 5'-untranslated region than those in the other cold-shock-inducible genes, and the promoter sequence, which was only 55 bp, was sufficient for cspH expression. The 14-base downstream box located 12 bases downstream of the initiation codon of cspH mRNA was essential for its cold shock activation.

  3. Gene expression values of pattern-recognition receptors in porcine leukocytes and their response to Salmonella enterica serovar Typhimurium infection.

    PubMed

    Osvaldova, Alena; Stepanova, Hana; Faldyna, Martin; Matiasovic, Jan

    2017-03-01

    Pattern-recognition receptors (PRRs) recognize pathogen-associated molecular patterns and play an important role in triggering innate immune responses. PRRs distribution and function is well documented in mice and humans, but studies in pigs are scarce. Salmonella enterica serovar Typhimurium is common pathogen found in pigs and was used as a model for interaction with PRRs. This study investigated expression of PRRs in porcine leukocyte subpopulations at the mRNA level. Eight subpopulations of leukocytes comprising NK cells, Th, Tc, double positive T cells and γδ T cells, B cells, monocytes and neutrophils were sorted, and the expression of 12 PRRs was measured, including selected Toll-like receptors and their co-receptors, NOD-like receptor NOD2, RP-105, CD14, and dectin. The highest expression rates of most PRRs were observed in monocytes and neutrophils. The B cells expressed high levels of TLR1, TLR6, TLR9, TLR10, and RP-105. Only monocytes and γδ T cells were found to respond to Salmonella enterica serovar Typhimurium infection by intensification of PRRs expression. In Th and B cells, PRRs mRNA down-regulation was detected after infection.

  4. A proteomic approach to study Salmonella enterica serovar Typhimurium putative transporter YjeH associated with ceftriaxone resistance

    SciTech Connect

    Hu, Wensi S. Lin, Y.-H.; Shih, C.-C.

    2007-09-28

    Mutant 6B7 of Salmonella enterica serovar Typhimurium has a transposon inserted in the putative transporter gene yjeH and shows a more-than-fourfold reduction in resistance to ceftriaxone. In this report we have used proteomic analysis to compare outer membrane protein profiles between this mutant and its parental strain R200. Five identified proteins were found to be altered. Of these proteins, the level of expression of the porin OmpD was increased and those of the putative outer membrane proteins STM1530 and STM3031, a subunit of the proton-pumping oxidoreductase NuoB and the heat shock protein MopA were decreased in 6B7 strain. Although the function of the yjeH gene remains unknown, a complementation assay suggested that the OmpD, STM1530, STM3031, NuoB, and MopA proteins are associated with ceftriaxone resistance and the expression of these proteins is influenced by the putative transporter gene yjeH in S. enterica serovar Typhimurium.

  5. Effect of the Escherichia coli EMO strain on experimental infection by Salmonella enterica serovar Typhimurium in gnotobiotic mice.

    PubMed

    Lima-Filho, J V M; Vieira, L Q; Arantes, R M E; Nicoli, J R

    2004-07-01

    An experimental infection with Salmonella enterica subsp. enterica serovar Typhimurium was evaluated in gnotobiotic mice previously exposed to a plasmid-free non-pathogenic Escherichia coli (EMO strain). Mice were exposed to EMO (experimental) or not (control) 10 days before challenge with Salmonella Typhimurium (10(2) colony forming units (CFU)/mouse). Survival after challenge was higher (P < 0.05) in the experimental group (16%) than in the control animals (0%). Histopathological examination of the colon and ileum mucosa of the experimental group showed less extensive lesions such as edema, cell inflammatory infiltration and hyperemia. The epithelial cells of the mucosal surface and the production of the mucous layer were also better preserved in the experimental group. The population levels of Salmonella Typhimurium in the feces were initially 10-fold lower (P < 0.05) in the experimental groups. However, 3 days after challenge both experimental and control groups showed similar population levels ranging from 10(8) to 10(9) CFU/g of feces. The intestinal contents of total and anti-Salmonella Typhimurium sIgA were higher in the experimental groups 10 days after inoculation of E. coli EMO strain. Translocation of Salmonella Typhimurium to the spleen was 10-fold lower (P < 0.05) in the experimental group only on day 3 after infection. This was not related to an increase in the bacterial blood clearance of the animals, as shown by experimental venous challenge with E. coli B41. In conclusion, treatment of mice with E. coli EMO strain promoted a relative protection against experimental infection with Salmonella Typhimurium. This protection was not due to the reduction of the population of pathogens in the intestine but was probably related to stimulation of the immune response.

  6. In Vitro Development of Ciprofloxacin Resistance of Salmonella enterica Serovars Typhimurium, Enteritidis, and Indiana Isolates from Food Animals.

    PubMed

    Zhang, Wen-Hui; Zhang, Chuan-Zhen; Liu, Zhi-Jie; Gu, Xi-Xi; Li, Wan; Yang, Ling; Liu, Ya-Hong; Zeng, Zhen-Ling; Jiang, Hong-Xia

    2017-01-13

    Difference in the development of resistance may be associated with the epidemiological spread and drug resistance of different Salmonella enterica serovar strains. In the present study, three susceptible S. enterica serovars, Typhimurium (ST), Enteritidis (SE), and Indiana (SI) strains, were subjected to stepwise selection with increasing ciprofloxacin concentrations. The results indicated that the mutation frequencies of the SI group were 10(1)-10(4) higher and developed resistance to ciprofloxacin more rapidly compared with the ST and SE groups. Ciprofloxacin accumulation in the SI strain was also higher than the other two strains in the presence of an efflux pump inhibitor. The development of ciprofloxacin resistance was quite different among the three serovar strains. In SI, increasing AcrAB-TolC efflux pump expression and single or double mutations in gyrA with or without a single parC mutation (T57S) were found in the development of ciprofloxacin resistance. In SE, an increase in the AcrAB-TolC efflux pump regulatory gene ramA gradually decreased as resistant bacteria developed; then resistance resulted from gyrA D87G and gyrB E466D mutations and/or in other active efflux pumps besides AcrAB-TolC. For ST, ramA expression increased rapidly along with gyrA D87 N and/or gyrB S464F mutations. In conclusion, persistent use of ciprofloxacin may aggravate the resistance of different S. enterica serovars and prudent use of the fluoroquinolones is needed. The quicker resistance and higher mutation frequency of the SI isolates present a potential public health threat.

  7. Differences in Pathogenesis for Salmonella enterica serovar Typhimurium in the Mouse Versus the Swine Model Identify Bacterial Gene Products Required for Systemic but not Gastrointestinal Disease

    USDA-ARS?s Scientific Manuscript database

    Over the last several decades, the mouse model of Typhoid fever has been an extremely productive model to investigate Salmonella enterica serovar Typhimurium pathogenesis. The mouse is the paradigm for investigating systemic disease due to infection by Salmonella; however, the swine model of gastro...

  8. Complete Genome Sequence of Salmonella enterica Serovar Typhimurium Strain SO3 (Sequence Type 302) Isolated from a Baby with Meningitis in Mexico

    PubMed Central

    Puente, José L.; Calva, Edmundo; Zaidi, Mussaret B.

    2016-01-01

    The complete genome of Salmonella enterica serovar Typhimurium strain SO3 (sequence type 302), isolated from a fatal meningitis infection in Mexico, was determined using PacBio technology. The chromosome hosts six complete prophages and is predicted to harbor 51 genomic islands, including 13 pathogenicity islands (SPIs). It carries the Salmonella virulence plasmid (pSTV). PMID:27103717

  9. Mixed biofilm formation by Shiga toxin-producing Escherichia coli and Salmonella enterica serovar typhimurium enhanced bacterial resistance to sanitization due to extracellular polymeric substances

    USDA-ARS?s Scientific Manuscript database

    Shiga toxin–producing Escherichia coli O157:H7 and Salmonella enterica serovar Typhimurium are important foodborne pathogens capable of forming single-species biofilms or coexisting in multispecies biofilm communities. Bacterial biofilm cells are usually more resistant to sanitization than their pla...

  10. Osmoregulated periplasmic glucans are needed for competitive growth and biofilm formation by Salmonella enterica serovar Typhimurium in leafy-green vegetable wash-waters and colonization in mice

    USDA-ARS?s Scientific Manuscript database

    Osmoregulated periplasmic glucans (OPGs) are major periplasmic constituents of Gram negative bacteria. The role of OPGs has been postulated in symbiotic as well as pathogenic host-microbe interactions. Here we report the role of OPGs from Salmonella enterica serovar Typhimurium during growth and b...

  11. Evidence for Lack of Acquisition of Tolerance in Salmonella enterica Serovar Typhimurium ATCC 14028 after Exposure to Subinhibitory Amounts of Origanum vulgare L. Essential Oil and Carvacrol

    PubMed Central

    Luz, Isabelle da Silva; Gomes Neto, Nelson Justino; Tavares, Adassa Gama; Nunes, Pollyana Campos; Magnani, Marciane

    2012-01-01

    Overnight exposure of Salmonella enterica serovar Typhimurium to sublethal amounts of Origanum vulgare essential oil (OV) and carvacrol (CAR) did not result in direct and cross-bacterial protection. Cells subcultured with increasing amounts of OV or CAR survived up to the MIC of either compound, revealing few significant changes in bacterial susceptibility. PMID:22544235

  12. Complete genome sequencing of a multidrug-resistant and human-invasive Salmonella enterica serovar Typhimurium strain of the emerging sequence type 213 genotype

    SciTech Connect

    Calva, Edmundo; Silva, Claudia; Zaidi, Mussaret B.; Sanchez-Flores, Alejandro; Estrada, Karel; Silva, Genivaldo G. Z.; Soto-Jiménez, Luz M.; Wiesner, Magdalena; Fernández-Mora, Marcos; Edwards, Robert A.; Vinuesa, Pablo

    2015-06-18

    Salmonella enterica subsp. enterica serovar Typhimurium strain YU39 was isolated in 2005 in the state of Yucatán, Mexico, from a human systemic infection. The YU39 strain is representative of the multidrug-resistant emergent sequence type 213 (ST213) genotype. The YU39 complete genome is composed of a chromosome and seven plasmids.

  13. Anti-infective mechanisms induced by a probiotic Lactobacillus strain against Salmonella enterica serovar Typhimurium infection.

    PubMed

    de LeBlanc, Alejandra de Moreno; Castillo, Natalia A; Perdigon, Gabriela

    2010-04-15

    The prevention of pathogen infections is one of the most extensively studied effects of probiotics. L. casei CRL 431 is a probiotic bacterium and its effects on the gut immune cells have been extensively studied. The aim of the present study was to determine, using a mouse model, the preventive and therapeutic effect of L. casei CRL 431 to achieve protection against Salmonella enteritidis serovar Typhimurium infection. In both previous and continuous (previous and post-infection) probiotic administration, the mechanisms induced by this lactic acid bacteria on the first line of intestinal defense (non-specific barrier and the innate immune cells associated to the gut), as a way to understand some of the mechanisms involved in the protection against Salmonella enteritidis serovar Typhimurium, were analyzed. The results obtained demonstrated that 7 days L. casei CRL 431 administration before infection decreased the severity of the infection with Salmonella enteritidis serovar Typhimurium, demonstrating that the continuous administration (even after infection) had the best effect. This continuous administration diminished the counts of the pathogen in the intestine as well as its spread outside this organ. Several mechanisms and cells are involved in this protective effect against Salmonella enteritidis serovar Typhimurium. L. casei CRL 431 acted on cells of the innate and adaptive immune response. The probiotic administration decreased the neutrophil infiltration with the consequent diminution of intestinal inflammation; activated the macrophage phagocytic activity in different sites such as Peyer's patches, spleen and peritoneum; and increased the number of IgA+cells in the lamina propria of the small intestine which was correlated with increased release of s-IgA specific against the pathogen in the intestinal fluids. The mechanism of the inhibition of cellular apoptosis was not involved.

  14. mcr-1−Harboring Salmonella enterica Serovar Typhimurium Sequence Type 34 in Pigs, China

    PubMed Central

    Yi, Linxian; Wang, Jing; Gao, Yanling; Liu, Yiyun; Doi, Yohei; Wu, Renjie; Zeng, Zhenling; Liang, Zisen

    2017-01-01

    We detected the mcr-1 gene in 21 (14.8%) Salmonella isolates from pigs at slaughter; 19 were serovar Typhimurium sequence type 34. The gene was located on IncHI2-like plasmids that also harbored IncF replicons and lacked a conjugative transfer region. These findings highlight the need to prevent further spread of colistin resistance in animals and humans. PMID:28098547

  15. Corrected Genome Annotations Reveal Gene Loss and Antibiotic Resistance as Drivers in the Fitness Evolution of Salmonella enterica Serovar Typhimurium.

    PubMed

    Paul, Sandip; Sokurenko, Evgeni V; Chattopadhyay, Sujay

    2016-12-01

    Horizontal acquisition of novel chromosomal genes is considered to be a key process in the evolution of bacterial pathogens. However, the identification of gene presence or absence could be hindered by the inconsistencies in bacterial genome annotations. Here, we performed a cross-annotation of omnipresent core and mosaic accessory genes in the chromosome of Salmonella enterica serovar Typhimurium across a total of 20 fully assembled genomes deposited into GenBank. Cross-annotation resulted in a 32% increase in the number of core genes and a 3-fold decrease in the number of genes identified as mosaic genes (i.e., genes present in some strains only) by the original annotation. Of the remaining noncore genes, the vast majority were prophage genes, and 255 of the nonphage genes were actually of core origin but lost in some strains upon the emergence of the S Typhimurium serovar, suggesting that the chromosomal portion of the S Typhimurium genome acquired a very limited number of novel genes other than prophages. Only horizontally acquired nonphage genes related to bacterial fitness or virulence were found in four recently sequenced isolates, all located on three different genomic islands that harbor multidrug resistance determinants. Thus, the extensive use of antimicrobials could be the main selection force behind the new fitness gene acquisition and the emergence of novel Salmonella pathotypes.

  16. Higher Storage Temperature Causes Greater Salmonella enterica Serovar Typhimurium Internal Penetration of Artificially Contaminated, Commercially Available, Washed Free Range Eggs.

    PubMed

    Whiley, Alice; Fallowfield, Howard; Ross, Kirstin; McEvoy, Vanessa; Whiley, Harriet

    2016-07-01

    Foodborne salmonellosis is a major public health concern, with contaminated eggs identified as a significant source of infection. In Australia, the most prevalent cause of salmonellosis from eggs is Salmonella enterica subsp. enterica serovar Typhimurium. This study explored the effect of temperature after 1, 7, 14, 21, and 28 days of storage on commercially available washed free range eggs, artificially contaminated with Salmonella Typhimurium on the external surface. At each time point, the external surface of the egg, the crushed eggshell, and the internal egg yolk and albumen were analyzed for Salmonella. After 28 days of storage, 25% of eggs stored at 4°C, 50% of eggs stored at 14°C, and 100% of eggs stored at 23 and 35°C were internally contaminated with Salmonella. After 1 day of storage, more than 50% of all eggs had Salmonella present in the crushed shell after the external surface had been disinfected with ethanol. This is the first study to demonstrate that refrigeration reduced the potential for Salmonella Typhimurium to penetrate the eggshell membrane and internally contaminate table eggs commercially available in Australia. It also suggests that the processes of cracking eggs may be a source of cross-contamination within the kitchen.

  17. Hydrosol of Thymbra capitata Is a Highly Efficient Biocide against Salmonella enterica Serovar Typhimurium Biofilms

    PubMed Central

    Karampoula, Foteini; Deschamps, Julien; Doulgeraki, Agapi I.; Nychas, George-John E.

    2016-01-01

    ABSTRACT Salmonella is recognized as one of the most significant enteric foodborne bacterial pathogens. In recent years, the resistance of pathogens to biocides and other environmental stresses, especially when they are embedded in biofilm structures, has led to the search for and development of novel antimicrobial strategies capable of displaying both high efficiency and safety. In this direction, the aims of the present work were to evaluate the antimicrobial activity of hydrosol of the Mediterranean spice Thymbra capitata against both planktonic and biofilm cells of Salmonella enterica serovar Typhimurium and to compare its action with that of benzalkonium chloride (BC), a commonly used industrial biocide. In order to achieve this, the disinfectant activity following 6-min treatments was comparatively evaluated for both disinfectants by calculating the concentrations needed to achieve the same log reductions against both types of cells. Their bactericidal effect against biofilm cells was also comparatively determined by in situ and real-time visualization of cell inactivation through the use of time-lapse confocal laser scanning microscopy (CLSM). Interestingly, results revealed that hydrosol was almost equally effective against biofilms and planktonic cells, whereas a 200-times-higher concentration of BC was needed to achieve the same effect against biofilm compared to planktonic cells. Similarly, time-lapse CLSM revealed the significant advantage of the hydrosol to easily penetrate within the biofilm structure and quickly kill the cells, despite the three-dimensional (3D) structure of Salmonella biofilm. IMPORTANCE The results of this paper highlight the significant antimicrobial action of a natural compound, hydrosol of Thymbra capitata, against both planktonic and biofilm cells of a common foodborne pathogen. Hydrosol has numerous advantages as a disinfectant of food-contact surfaces. It is an aqueous solution which can easily be rinsed out from surfaces, it

  18. Hydrosol of Thymbra capitata Is a Highly Efficient Biocide against Salmonella enterica Serovar Typhimurium Biofilms.

    PubMed

    Karampoula, Foteini; Giaouris, Efstathios; Deschamps, Julien; Doulgeraki, Agapi I; Nychas, George-John E; Dubois-Brissonnet, Florence

    2016-09-01

    Salmonella is recognized as one of the most significant enteric foodborne bacterial pathogens. In recent years, the resistance of pathogens to biocides and other environmental stresses, especially when they are embedded in biofilm structures, has led to the search for and development of novel antimicrobial strategies capable of displaying both high efficiency and safety. In this direction, the aims of the present work were to evaluate the antimicrobial activity of hydrosol of the Mediterranean spice Thymbra capitata against both planktonic and biofilm cells of Salmonella enterica serovar Typhimurium and to compare its action with that of benzalkonium chloride (BC), a commonly used industrial biocide. In order to achieve this, the disinfectant activity following 6-min treatments was comparatively evaluated for both disinfectants by calculating the concentrations needed to achieve the same log reductions against both types of cells. Their bactericidal effect against biofilm cells was also comparatively determined by in situ and real-time visualization of cell inactivation through the use of time-lapse confocal laser scanning microscopy (CLSM). Interestingly, results revealed that hydrosol was almost equally effective against biofilms and planktonic cells, whereas a 200-times-higher concentration of BC was needed to achieve the same effect against biofilm compared to planktonic cells. Similarly, time-lapse CLSM revealed the significant advantage of the hydrosol to easily penetrate within the biofilm structure and quickly kill the cells, despite the three-dimensional (3D) structure of Salmonella biofilm. The results of this paper highlight the significant antimicrobial action of a natural compound, hydrosol of Thymbra capitata, against both planktonic and biofilm cells of a common foodborne pathogen. Hydrosol has numerous advantages as a disinfectant of food-contact surfaces. It is an aqueous solution which can easily be rinsed out from surfaces, it does not have the

  19. The importance of motility and chemotaxis for extra-animal survival of Salmonella enterica serovar Typhimurium and Dublin.

    PubMed

    Olsen, J E; Hoegh-Andersen, K H; Casadesús, J; Thomsen, L E

    2012-09-01

    This study investigated the importance of flagella and motility of Salmonella enterica serovar Typhimurium and Dublin in models of extra-animal survival. The study was performed using transposon mutants in flagella genes fliC and fljB and in chemotaxis genes cheA, cheB and cheR. Flagella and chemotaxis were found to be of minor importance for attachment to plant leaves, survival in liquid manure and interaction with the nematode C. elegans, while differences were observed between the fliC mutant and the wild-type strain of S. Dublin in interactions with amoebae. The study shows that flagella and chemotaxis play a minor role in extra-animal survival of these two serovars of Salmonella under the conditions tested. Extra-animal survival is important in the full infection cycle for zoonotic salmonellae. Such serovars are motile. Even though the current study was only based on the characterization of two serovars, it strongly suggests that motility and chemotaxis are of minor importance during the spread of Salmonella from one animal to the next through the external environment. © 2012 The Authors Journal of Applied Microbiology © 2012 The Society for Applied Microbiology.

  20. Novel Approach of a Phage-Based Magnetoelastic Biosensor for the Detection of Salmonella enterica serovar Typhimurium in Soil.

    PubMed

    Park, Mi-Kyung; Chin, Bryan A

    2016-12-28

    To date, there has been no employment of a magnetoelastic (ME) biosensor method to detect Salmonella enterica serovar Typhimurium in soil. The ME biosensor method needs to be investigated and modified for its successful performance. The filtration method, cation-exchange resin method, and combinations of both methods were employed for the extraction of S. Typhimurium from soil. The number of S. Typhimurium and the resonant frequency shift of the ME sensor were then compared using a brilliant green sulfa agar plate and an HP 8751A network analyzer. A blocking study was performed using bovine serum albumin (BSA), polyethylene glycol (PEG), and casein powder suspension. Finally, the modified ME biosensor method was performed to detect S. Typhimurium in soil. The number of S. Typhimurium was significantly decreased from 7.10 log CFU/soil to 4.45-4.72 log CFU/soil after introduction of the cation-exchange resin method. The greatest resonant frequency shift of the measurement sensor was found when employing centrifugation and filtration procedures. The resonant frequency shift of the PEG-blocked measurement sensor was 3,219 ± 755 Hz, which was significantly greater than those of the BSA- and casein-blocked ME sensor. The optimum concentration of PEG was determined to be 1.0 mg/ml after considering the resonant shift and economic issue. Finally, the modified ME biosensor method was able to detect S. Typhimurium in soil in a dose-response manner. Although these modifications of the ME biosensor method sacrificed some advantages, such as cost, time effectiveness, and operator friendliness, this study demonstrated a novel approach of the ME biosensor method to detect S. Typhimurium in soil.

  1. Bidirectional Salmonella enterica serovar Typhimurium transfer between bare/glove hands and green bell pepper and its interruption.

    PubMed

    Jimenez, Maribel; Siller, Jorge H; Valdez, Jose B; Carrillo, Armando; Chaidez, Cristobal

    2007-10-01

    The aim of this study was to quantify the amount of Salmonella enterica serovar Typhimurium transferred from volunteers' hands (bare or gloved) to green bell peppers and vice versa; and to assess the effectiveness of hand hygiene techniques. The highest and lowest percentages of bacterial transfer were achieve from green bell peppers to gloved hands (46.56%) and from bare hands to green bell peppers (0.21%), respectively. The highest and lowest Log10 reductions of S. Typhimurium were achieved by the combination of hand washing and alcohol-based gel (4.38 Log10) and iodine solution (2.08 Log10), respectively. This study provides important information concerning the transfer's efficiency of S. Typhimurium from hands to fresh produce and from fresh produce to hands. The study also showed that gloved hands, could be a mean of transfer of S. Typhimurium between green peppers and hands, and the best hand hygiene technique was the combination of hand washing and alcohol-based gel.

  2. Enterobacterial Common Antigen Mutants of Salmonella enterica Serovar Typhimurium Establish a Persistent Infection and Provide Protection against Subsequent Lethal Challenge

    PubMed Central

    Gilbreath, Jeremy J.; Colvocoresses Dodds, Jennifer; Rick, Paul D.; Soloski, Mark J.

    2012-01-01

    Infection with Salmonella spp. is a significant source of disease globally. A substantial proportion of these infections are caused by Salmonella enterica serovar Typhimurium. Here, we characterize the role of the enterobacterial common antigen (ECA), a surface glycolipid ubiquitous among enteric bacteria, in S. Typhimurium pathogenesis. Construction of a defined mutation in the UDP-N-acetylglucosamine-1-phosphate transferase gene, wecA, in two clinically relevant strains of S. Typhimurium, TML and SL1344, resulted in strains that were unable to produce ECA. Loss of ECA did not affect the gross cell surface ultrastructure, production of lipopolysaccharide (LPS), flagella, or motility. However, the wecA mutant strains were attenuated in both oral and intraperitoneal mouse models of infection (P < 0.001 for both routes of infection; log rank test), and virulence could be restored by complementation of the wecA gene in trans. Despite the avirulence of the ECA-deficient strains, the wecA mutant strains were able to persistently colonize systemic sites (spleen and liver) at moderate levels for up to 70 days postinfection. Moreover, immunization with the wecA mutant strains provided protection against a subsequent lethal oral or intraperitoneal challenge with wild-type S. Typhimurium. Thus, wecA mutant (ECA-negative) strains of Salmonella may be useful as live attenuated vaccine strains or as vehicles for heterologous antigen expression. PMID:22025511

  3. Phosphate Groups of Lipid A Are Essential for Salmonella enterica Serovar Typhimurium Virulence and Affect Innate and Adaptive Immunity

    PubMed Central

    Six, David A.; Liu, Qing; Gu, Lillian; Wang, Shifeng; Alamuri, Praveen; Raetz, Christian R. H.

    2012-01-01

    Lipid A is a key component of the outer membrane of Gram-negative bacteria and stimulates proinflammatory responses via the Toll-like receptor 4 (TLR4)-MD2-CD14 pathway. Its endotoxic activity depends on the number and length of acyl chains and its phosphorylation state. In Salmonella enterica serovar Typhimurium, removal of the secondary laurate or myristate chain in lipid A results in bacterial attenuation and growth defects in vitro. However, the roles of the two lipid A phosphate groups in bacterial virulence and immunogenicity remain unknown. Here, we used an S. Typhimurium msbB pagL pagP lpxR mutant, carrying penta-acylated lipid A, as the parent strain to construct a series of mutants synthesizing 1-dephosphorylated, 4′-dephosphorylated, or nonphosphorylated penta-acylated lipid A. Dephosphorylated mutants exhibited increased sensitivity to deoxycholate and showed increased resistance to polymyxin B. Removal of both phosphate groups severely attenuated the mutants when administered orally to BALB/c mice, but the mutants colonized the lymphatic tissues and were sufficiently immunogenic to protect the host from challenge with wild-type S. Typhimurium. Mice receiving S. Typhimurium with 1-dephosphorylated or nonphosphorylated penta-acylated lipid A exhibited reduced levels of cytokines. Attenuated and dephosphorylated Salmonella vaccines were able to induce adaptive immunity against heterologous (PspA of Streptococcus pneumoniae) and homologous antigens (lipopolysaccharide [LPS] and outer membrane proteins [OMPs]). PMID:22753374

  4. Differences in Salmonella enterica serovar Typhimurium strain invasiveness are associated with heterogeneity in SPI-1 gene expression

    PubMed Central

    Clark, Leann; Perrett, Charlotte A.; Malt, Layla; Harward, Caryn; Humphrey, Suzanne; Jepson, Katy A.; Martinez-Argudo, Isabel; Carney, Laura J.; La Ragione, Roberto M.; Humphrey, Tom J.

    2011-01-01

    Most studies on Salmonella enterica serovar Typhimurium infection focus on strains ATCC SL1344 or NTCC 12023 (ATCC 14028). We have compared the abilities of these strains to induce membrane ruffles and invade epithelial cells. S. Typhimurium strain 12023 is less invasive and induces smaller membrane ruffles on MDCK cells compared with SL1344. Since the SPI-1 effector SopE is present in SL1344 and absent from 12023, and SL1344 sopE mutants have reduced invasiveness, we investigated whether 12023 is less invasive due to the absence of SopE. However, comparison of SopE+ and SopE− S. Typhimurium strains, sopE deletion mutants and 12023 expressing a sopE plasmid revealed no consistent relationship between SopE status and relative invasiveness. Nevertheless, absence of SopE was closely correlated with reduced size of membrane ruffles. A PprgH–gfp reporter revealed that relatively few of the 12023 population (and that of the equivalent strain ATCC 14028) express SPI-1 compared to other S. Typhimurium strains. Expression of a PhilA–gfp reporter mirrored that of PprgH–gfp in 12023 and SL1344, implicating reduced signalling via the transcription factor HilA in the heterogeneous SPI-1 expression of these strains. The previously unrecognized strain heterogeneity in SPI-1 expression and invasiveness has important implications for studies of Salmonella infection. PMID:21493681

  5. The role of flagella and chemotaxis genes in host pathogen interaction of the host adapted Salmonella enterica serovar Dublin compared to the broad host range serovar S. Typhimurium

    PubMed Central

    2013-01-01

    Background The importance of flagella and chemotaxis genes in host pathogen interaction in Salmonella enterica is mainly based on studies of the broad host range serovar, S. Typhimurium, while little is known on the importance in host specific and host adapted serovars, such as S. Dublin. In the current study we have used previously characterized insertion mutants in flagella and chemotaxis genes to investigate this and possible differences in the importance between the two serovars. Results fliC (encoding the structural protein of the flagella) was essential for adhesion and fliC and cheB (CheB restores the chemotaxis system to pre-stimulus conformation) were essential for invasion of S. Dublin into epithelial Int407 cells. In S. Typhimurium, both lack of flagella (fliC/fljB double mutant) and cheB influenced adhesion, and invasion was influenced by lack of both cheA (the histidine-kinase of the chemotaxis system), fliC/fljB and cheB mutation. Uptake in J774A.1 macrophage cells was significantly reduced in cheA, cheB and fliC mutants of S. Dublin, while cheA was dispensable in S. Typhimurium. Removal of flagella in both serotypes caused an increased ability to propagate intracellular in J774 macrophage cells and decreased cytotoxicity toward these cells. Flagella and chemotaxis genes were found not to influence the oxidative response. The induction of IL-6 from J774A-1 cells depended on the presence of flagella in S. Typhimurium, whilst this was not the case following challenge with S. Dublin. Addition of fliC from S. Typhimurium in trans to a fliC mutant of S. Dublin increased cytotoxicity but it did not increase the IL-6 production. Flagella were demonstrated to contribute to the outcome of infection following oral challenge of mice in S. Dublin, while an S. Typhimurium fliC/fljB mutant showed increased virulence following intra peritoneal challenge. Conclusions The results showed that flagella and chemotaxis genes differed in their role in host pathogen

  6. Gamma radiation used as hygienization technique for foods does not induce viable but non-culturable state (VBNC) in Salmonella enterica subsp. enterica serovar Typhimurium.

    PubMed

    Saroj, Sunil; Shashidhar, R; Bandekar, Jayant

    2009-10-01

    Gamma radiation has been widely used for hygienization of food products. Whether gamma radiation stress induces VBNC state in Salmonella is of great concern. Therefore, the study was carried out to determine whether gamma radiation exposure induces VBNC state in Salmonella enterica subsp. enterica serovar Typhimurium (S. typhimurium). The parameters tested were culturability on agar medium, transcriptional activity by RT-PCR, cytoplasmic membrane integrity, and direct viable count using LIVE/DEAD BacLight bacterial viability kit. The LIVE/DEAD BacLight counts for S. typhimurium cells treated with 0.5 and 1.0 kGy radiation dose were 0.8 and 0.1% of the control, respectively. Plate counts for S. typhimurium cells treated with 0.5 and 1.0 kGy radiation dose were 0.7 and 0.05% of the control, respectively. No viable cells of S. typhimurium were detected by both plate count and LIVE/DEAD BacLight after radiation treatment with 2 kGy. No transcriptional activity was detected in cells treated with 2 kGy radiation dose. If there were VBNC cells present, then significant differences in the counts between the LIVE/DEAD BacLight microscopic counts and plate agar counts must be observed. No significant difference (P > 0.05) in the counts were observed. Thus, it can be concluded that treatment with 2 kGy results in complete killing and does not induce VBNC state in S. typhimurium.

  7. Fermented rapeseed meal is effective in controlling Salmonella enterica serovar Typhimurium infection and improving growth performance in broiler chicks.

    PubMed

    Ashayerizadeh, Amin; Dastar, Behrouz; Shams Shargh, Mahmoud; Sadeghi Mahoonak, Alireza; Zerehdaran, Saeed

    2017-03-01

    The aim of present experiment was to assess the effects of fermented rapeseed meal (FRSM) on Salmonella enterica serovar Typhimurium (S. Typhimurium) colonization and growth performance in broiler chicks. Two hundred forty day-old male Cobb 500 broiler chicks were divided into six experimental treatments with four replicates and 10 birds per each. The treatments were including two positive and negative controls which birds received a basal corn-soybean diet as well as four others which birds received the diets that rapeseed meal (RSM) or FRSM was replaced with soybean meal at 50 and 100% levels. All chicks except the negative control birds were challenged orally with 10(5) CFU of S. Typhimurium at 3days of age. Results showed that birds were fed FRSM had significantly greater lactic acid bacteria populations and lesser S. Typhimurium colonization in ileal and cecal sections compared to others (P<0.05). The less percentage of liver and bursa of fabricius was belonged to negative control group. At 10day, feeding chicks with diet containing FRSM, but not RSM, significantly decreased the organ invasion by S. Typhimurium (P<0.05). Heterophil to lymphocyte ratio was significantly lesser in chicks were fed FRSM compared to those fed RSM or positive control (P<0.05). Birds were fed FRSM had significantly higher weight gain and better feed conversion ratio compared to those birds were fed RSM (P<0.05). The findings of present experiment concerning positive effects of feeding FRSM on reducing S. Typhimurium and improving growth performance show that this processed protein source can be considered as a nutritional effective strategy to control Salmonella contamination in broiler chicks.

  8. Molecular and Functional Characterization of Salmonella enterica Serovar Typhimurium poxA Gene: Effect on Attenuation of Virulence and Protection

    PubMed Central

    Kaniga, Koné; Compton, Melissa S.; Curtiss, Roy; Sundaram, Preeti

    1998-01-01

    Salmonella enterica poxA mutants exhibit a pleiotropic phenotype, including reduced pyruvate oxidase activity; reduced growth rate; and hypersensitivity to the herbicide sulfometuron methyl, α-ketobutyrate, and amino acid analogs. These mutants also failed to grow in the presence of the host antimicrobial peptide, protamine. In this study, PoxA− mutants of S. enterica serovar Typhimurium (S. typhimurium) were found to be 10,000-fold attenuated in orally inoculated BALB/c mice and 1,000-fold attenuated in intraperitoneally inoculated BALB/c mice, compared to wild-type S. typhimurium UK-1. In addition, poxA mutants were found to be capable of colonizing the spleen, mesenteric lymph nodes, and Peyer’s patches; to induce strong humoral immune responses; and to protect mice against a lethal wild-type Salmonella challenge. A 2-kb DNA fragment was isolated from wild-type S. typhimurium UK-1 based on its ability to complement an isogenic poxA mutant. The nucleotide sequence of this DNA fragment revealed an open reading frame of 325 amino acids capable of encoding a polypeptide of 36.8 kDa that was confirmed in the bacteriophage T7 expression system. Comparison of the translated sequence to the available databases indicated high homology to a family of lysyl-tRNA synthetases. Our results indicate that a mutation of poxA has an attenuating effect on Salmonella virulence. Further, poxA mutants are immunogenic and could be useful in designing live vaccines with a variety of bacterial species. To our knowledge, this is the first report on the effect of poxA mutation on bacterial virulence. PMID:9826331

  9. Molecular characterization of Salmonella enterica serovar Typhimurium isolated from human, food, and animal sources in Malaysia.

    PubMed

    Ngoi, Soo Tein; Lindstedt, Bjørn-Arne; Watanabe, Haruo; Thong, Kwai Lin

    2013-01-01

    Salmonella Typhimurium is an important nontyphoidal Salmonella serovar associated with foodborne diseases in many parts of the world. This organism is the major causative agent of nontyphoidal salmonellosis in Malaysia. We aimed to investigate the genetic profiles of the strains isolated from clinical, zoonotic, and dietary sources in Malaysia using multilocus variable number tandem repeat analysis (MLVA) and pulsed-field gel electrophoresis (PFGE). By focusing on the 5 common variable number tandem repeat (VNTR) loci, we found that PFGE (D = 0.99) was more discriminative than MLVA (D = 0.76). The low MLVA score might be because of a lack of VNTR loci STTR6 (81.0%) and STTR10pl (76.2%). Both subtyping methods suggested that our S. Typhimurium strains were largely endemic with limited genetic variation. Furthermore, we observed that biphasic S. Typhimurium strains were dominant (99%) and multidrug resistance was prevalent (50%) within our sample pool. The most frequently observed phenotypes were resistance to compound sulfonamides (49%), tetracycline (51%), and streptomycin (52%). In this study, we documented the genetic relationship, antimicrobial resistance characteristics, and flagellar-phase dominance among S. Typhimurium strains found in Malaysia.

  10. Expression of STM4467-encoded arginine deiminase controlled by the STM4463 regulator contributes to Salmonella enterica serovar Typhimurium virulence.

    PubMed

    Choi, Younho; Choi, Jeongjoon; Groisman, Eduardo A; Kang, Dong-Hyun; Shin, Dongwoo; Ryu, Sangryeol

    2012-12-01

    Arginine deiminase (ADI), carbamate kinase (CK), and ornithine transcarbamoylase (OTC) constitute the ADI system. In addition to metabolic functions, the ADI system has been implicated in the virulence of certain pathogens. The pathogenic intracellular bacterium Salmonella enterica serovar Typhimurium possesses the STM4467, STM4466, and STM4465 genes, which are predicted to encode ADI, CK, and OTC, respectively. Here we report that the STM4467 gene encodes an ADI and that ADI activity plays a role in the successful infection of a mammalian host by S. Typhimurium. An STM4467 deletion mutant was defective for replication inside murine macrophages and was attenuated for virulence in mice. We determined that a regulatory protein encoded by the STM4463 gene functions as an activator for STM4467 expression. The expression of the ADI pathway genes was enhanced inside macrophages in a process that required STM4463. Lack of STM4463 impaired the ability of S. Typhimurium to replicate within macrophages. A mutant defective in STM4467-encoded ADI displayed normal production of nitric oxide by macrophages.

  11. Expression of STM4467-Encoded Arginine Deiminase Controlled by the STM4463 Regulator Contributes to Salmonella enterica Serovar Typhimurium Virulence

    PubMed Central

    Choi, Younho; Choi, Jeongjoon; Groisman, Eduardo A.; Kang, Dong-Hyun

    2012-01-01

    Arginine deiminase (ADI), carbamate kinase (CK), and ornithine transcarbamoylase (OTC) constitute the ADI system. In addition to metabolic functions, the ADI system has been implicated in the virulence of certain pathogens. The pathogenic intracellular bacterium Salmonella enterica serovar Typhimurium possesses the STM4467, STM4466, and STM4465 genes, which are predicted to encode ADI, CK, and OTC, respectively. Here we report that the STM4467 gene encodes an ADI and that ADI activity plays a role in the successful infection of a mammalian host by S. Typhimurium. An STM4467 deletion mutant was defective for replication inside murine macrophages and was attenuated for virulence in mice. We determined that a regulatory protein encoded by the STM4463 gene functions as an activator for STM4467 expression. The expression of the ADI pathway genes was enhanced inside macrophages in a process that required STM4463. Lack of STM4463 impaired the ability of S. Typhimurium to replicate within macrophages. A mutant defective in STM4467-encoded ADI displayed normal production of nitric oxide by macrophages. PMID:23006851

  12. No beneficial effects evident for Enterococcus faecium NCIMB 10415 in weaned pigs infected with Salmonella enterica serovar Typhimurium DT104.

    PubMed

    Kreuzer, Susanne; Janczyk, Pawel; Assmus, Jens; Schmidt, Michael F G; Brockmann, Gudrun A; Nöckler, Karsten

    2012-07-01

    Salmonella enterica serovar Typhimurium DT 104 is the major pathogen for salmonellosis outbreaks in Europe. We tested if the probiotic bacterium Enterococcus faecium NCIMB 10415 can prevent or alleviate salmonellosis. Therefore, piglets of the German Landrace breed that were treated with E. faecium (n = 16) as a feed additive and untreated controls (n = 16) were challenged with S. Typhimurium 10 days after weaning. The presence of salmonellae in feces and selected organs, as well as the immune response, were investigated. Piglets treated with E. faecium gained less weight than control piglets (P = 0.05). The feeding of E. faecium had no effect on the fecal shedding of salmonellae and resulted in a higher abundance of the pathogen in tonsils of all challenged animals. The specific (anti-Salmonella IgG) and nonspecific (haptoglobin) humoral immune responses as well as the cellular immune response (T helper cells, cytotoxic T cells, regulatory T cells, γδ T cells, and B cells) in the lymph nodes, Peyer's patches of different segments of the intestine (jejunal and ileocecal), the ileal papilla, and in the blood were affected in the course of time after infection (P < 0.05) but not by the E. faecium treatment. These results led to the conclusion that E. faecium may not have beneficial effects on the performance of weaned piglets in the case of S. Typhimurium infection. Therefore, we suggest a critical discussion and reconsideration of E. faecium NCIMB 10415 administration as a probiotic for pigs.

  13. No Beneficial Effects Evident for Enterococcus faecium NCIMB 10415 in Weaned Pigs Infected with Salmonella enterica Serovar Typhimurium DT104

    PubMed Central

    Kreuzer, Susanne; Aßmus, Jens; Schmidt, Michael F. G.; Brockmann, Gudrun A.; Nöckler, Karsten

    2012-01-01

    Salmonella enterica serovar Typhimurium DT 104 is the major pathogen for salmonellosis outbreaks in Europe. We tested if the probiotic bacterium Enterococcus faecium NCIMB 10415 can prevent or alleviate salmonellosis. Therefore, piglets of the German Landrace breed that were treated with E. faecium (n = 16) as a feed additive and untreated controls (n = 16) were challenged with S. Typhimurium 10 days after weaning. The presence of salmonellae in feces and selected organs, as well as the immune response, were investigated. Piglets treated with E. faecium gained less weight than control piglets (P = 0.05). The feeding of E. faecium had no effect on the fecal shedding of salmonellae and resulted in a higher abundance of the pathogen in tonsils of all challenged animals. The specific (anti-Salmonella IgG) and nonspecific (haptoglobin) humoral immune responses as well as the cellular immune response (T helper cells, cytotoxic T cells, regulatory T cells, γδ T cells, and B cells) in the lymph nodes, Peyer's patches of different segments of the intestine (jejunal and ileocecal), the ileal papilla, and in the blood were affected in the course of time after infection (P < 0.05) but not by the E. faecium treatment. These results led to the conclusion that E. faecium may not have beneficial effects on the performance of weaned piglets in the case of S. Typhimurium infection. Therefore, we suggest a critical discussion and reconsideration of E. faecium NCIMB 10415 administration as a probiotic for pigs. PMID:22544257

  14. Functional Analysis of the Genes Encoding Diaminopropionate Ammonia Lyase in Escherichia coli and Salmonella enterica Serovar Typhimurium

    PubMed Central

    Kalyani, J. N.; Ramachandra, Nagaraju; Kachroo, Aashiq H.; Mahadevan, S.

    2012-01-01

    Diaminopropionate ammonia lyase (DAPAL) is a pyridoxal-5′phosphate (PLP)-dependent enzyme that catalyzes the conversion of diaminopropionate (DAP) to pyruvate and ammonia and plays an important role in cell metabolism. We have investigated the role of the ygeX gene of Escherichia coli K-12 and its ortholog, STM1002, in Salmonella enterica serovar Typhimurium LT2, presumed to encode DAPAL, in the growth kinetics of the bacteria. While Salmonella Typhimurium LT2 could grow on dl-DAP as a sole carbon source, the wild-type E. coli K-12 strain exhibited only marginal growth on dl-DAP, suggesting that DAPAL is functional in S. Typhimurium. The expression of ygeX in E. coli was low as detected by reverse transcriptase PCR (RT-PCR), consistent with the poor growth of E. coli on dl-DAP. Strains of S. Typhimurium and E. coli with STM1002 and ygeX, respectively, deleted showed loss of growth on dl-DAP, confirming that STM1002 (ygeX) is the locus encoding DAPAL. Interestingly, the presence of dl-DAP caused a growth inhibition of the wild-type E. coli strain as well as the knockout strains of S. Typhimurium and E. coli in minimal glucose/glycerol medium. Inhibition by dl-DAP was rescued by transforming the strains with plasmids containing the STM1002 (ygeX) gene encoding DAPAL or supplementing the medium with Casamino Acids. Growth restoration studies using media lacking specific amino acid supplements suggested that growth inhibition by dl-DAP in the absence of DAPAL is associated with auxotrophy related to the inhibition of the enzymes involved in the biosynthetic pathways of pyruvate and aspartate and the amino acids derived from them. PMID:22904288

  15. BcsZ inhibits biofilm phenotypes and promotes virulence by blocking cellulose production in Salmonella enterica serovar Typhimurium.

    PubMed

    Ahmad, Irfan; Rouf, Syed Fazle; Sun, Lei; Cimdins, Annika; Shafeeq, Sulman; Le Guyon, Soazig; Schottkowski, Marco; Rhen, Mikael; Römling, Ute

    2016-10-19

    Cellulose, a 1,4 beta-glucan polysaccharide, is produced by a variety of organisms including bacteria. Although the production of cellulose has a high biological, ecological and economical impact, regulatory mechanisms of cellulose biosynthesis are mostly unknown. Family eight cellulases are regularly associated with cellulose biosynthesis operons in bacteria; however, their function is poorly characterized. In this study, we analysed the role of the cellulase BcsZ encoded by the bcsABZC cellulose biosynthesis operon of Salmonella enterica serovar Typhimurium (S. Typhimurium) in biofilm related behavior. We also investigated the involvement of BcsZ in pathogenesis of S. Typhimurium including a murine typhoid fever infection model. In S. Typhimurium, cellulase BcsZ with a putative periplasmic location negatively regulates cellulose biosynthesis. Moreover, as assessed with a non-polar mutant, BcsZ affects cellulose-associated phenotypes such as the rdar biofilm morphotype, cell clumping, biofilm formation, pellicle formation and flagella-dependent motility. Strikingly, although upregulation of cellulose biosynthesis was not observed on agar plate medium at 37 °C, BcsZ is required for efficient pathogen-host interaction. Key virulence phenotypes of S. Typhimurium such as invasion of epithelial cells and proliferation in macrophages were positively regulated by BcsZ. Further on, a bcsZ mutant was outcompeted by the wild type in organ colonization in the murine typhoid fever infection model. Selected phenotypes were relieved upon deletion of the cellulose synthase BcsA and/or the central biofilm activator CsgD. Although the protein scaffold has an additional physiological role, our findings indicate that the catalytic activity of BcsZ effectively downregulates CsgD activated cellulose biosynthesis. Repression of cellulose production by BcsZ subsequently enables Salmonella to efficiently colonize the host.

  16. Influence of a Probiotic Strain of Enterococcus faecium on Salmonella enterica Serovar Typhimurium DT104 Infection in a Porcine Animal Infection Model▿

    PubMed Central

    Szabó, István; Wieler, Lothar H.; Tedin, Karsten; Scharek-Tedin, Lydia; Taras, David; Hensel, Andreas; Appel, Bernd; Nöckler, Karsten

    2009-01-01

    The beneficial effects of probiotic Enterococcus spp. in different hosts, such as mice and humans, have previously been reported in several studies. However, studies of large domestic animals, as well as challenge studies with pathogenic microorganisms, are very rare. Here, we investigated the influence of oral treatment of pigs with the probiotic bacterium Enterococcus faecium NCIMB 10415 on Salmonella enterica serovar Typhimurium DT104 infections in weaning piglets. Clinical symptoms, fecal excretion, the organ distribution of Salmonella, and the humoral immune response (immunoglobulin G [IgG], IgM, and IgA levels) in serum were examined. A pool of 89 piglets was randomly divided into probiotic and control groups. The probiotic group received a feed supplement containing E. faecium starting on day 14 postpartum prior to challenge with Salmonella serovar Typhimurium DT104 at 28 days postpartum. After challenge with Salmonella serovar Typhimurium DT104, piglets in both groups showed no severe clinical signs of salmonellosis. However, fecal excretion and colonization of Salmonella in organs were significantly greater in piglets fed E. faecium. Likewise, the humoral immune response against Salmonella (serum IgM and IgA levels) was significantly greater in the probiotic group animals than in control animals. The results of this study suggest that E. faecium NCIMB 10415 treatment enhanced the course of infection in weaning piglets challenged with Salmonella serovar Typhimurium DT104. However, the probiotic treatment also appeared to result in greater production of specific antibodies against Salmonella serovar Typhimurium DT104. PMID:19270131

  17. Influence of a probiotic strain of Enterococcus faecium on Salmonella enterica serovar Typhimurium DT104 infection in a porcine animal infection model.

    PubMed

    Szabó, István; Wieler, Lothar H; Tedin, Karsten; Scharek-Tedin, Lydia; Taras, David; Hensel, Andreas; Appel, Bernd; Nöckler, Karsten

    2009-05-01

    The beneficial effects of probiotic Enterococcus spp. in different hosts, such as mice and humans, have previously been reported in several studies. However, studies of large domestic animals, as well as challenge studies with pathogenic microorganisms, are very rare. Here, we investigated the influence of oral treatment of pigs with the probiotic bacterium Enterococcus faecium NCIMB 10415 on Salmonella enterica serovar Typhimurium DT104 infections in weaning piglets. Clinical symptoms, fecal excretion, the organ distribution of Salmonella, and the humoral immune response (immunoglobulin G [IgG], IgM, and IgA levels) in serum were examined. A pool of 89 piglets was randomly divided into probiotic and control groups. The probiotic group received a feed supplement containing E. faecium starting on day 14 postpartum prior to challenge with Salmonella serovar Typhimurium DT104 at 28 days postpartum. After challenge with Salmonella serovar Typhimurium DT104, piglets in both groups showed no severe clinical signs of salmonellosis. However, fecal excretion and colonization of Salmonella in organs were significantly greater in piglets fed E. faecium. Likewise, the humoral immune response against Salmonella (serum IgM and IgA levels) was significantly greater in the probiotic group animals than in control animals. The results of this study suggest that E. faecium NCIMB 10415 treatment enhanced the course of infection in weaning piglets challenged with Salmonella serovar Typhimurium DT104. However, the probiotic treatment also appeared to result in greater production of specific antibodies against Salmonella serovar Typhimurium DT104.

  18. Dissemination of clonal Salmonella enterica serovar Typhimurium isolates causing salmonellosis in Mauritius.

    PubMed

    Issack, Mohammad I; Garcia-Migura, Lourdes; Ramsamy, Veemala D; Svendsen, Christina A; Pornruangwong, Srirat; Pulsrikarn, Chaiwat; Hendriksen, Rene S

    2013-07-01

    Salmonella enterica serotype Typhimurium is one of the leading causes of salmonellosis in Mauritius, where it has also been associated with outbreaks of foodborne illness. However, little is known about its molecular epidemiology in the country. This study was therefore undertaken to investigate the clonality and source of Salmonella Typhimurium in Mauritius by studying human, food, and poultry isolates by pulsed-field gel electrophoresis (PFGE) and antibiotic minimum inhibitory concentration determination. Forty-nine isolates collected between 2008 and 2011 were analyzed, including 25 stool isolates from foodborne illness outbreaks and sporadic gastroenteritis cases, four blood isolates, one postmortem colon isolate, 14 food isolates, and five poultry isolates. All isolates were pansusceptible to the 16 antibiotics tested, except for two isolates that were resistant to sulfamethoxazole and trimethoprim. Overall characterization of the isolates by PFGE digested with XbaI and BlnI resulted in eight different patterns. The largest of the clusters in the composite dataset consisted of 20 isolates, including two raw chicken isolates, four poultry isolates, and nine human stool isolates from two outbreaks. A second cluster consisted of 18 isolates, of which 12 originated from human blood and stool samples from both sporadic and outbreak cases. Six food isolates were also found in this cluster, including isolates from raw and grilled chicken, marlin mousse, and cooked pork. One poultry isolate had a closely related PFGE pattern. The results indicate that one clone of Salmonella Typhimurium found in poultry has been causing outbreaks of foodborne illness in Mauritius and another clone that has caused many cases of gastrointestinal illness and bacteremia in humans could also be linked to poultry. Thus, poultry appears to be a major reservoir for Salmonella Typhimurium in Mauritius. Initiating on-farm control strategies and measures against future dissemination may

  19. Novel small RNA (sRNA) landscape of the starvation-stress response transcriptome of Salmonella enterica serovar typhimurium

    PubMed Central

    Amin, Shivam V.; Roberts, Justin T.; Patterson, Dillon G.; Coley, Alexander B.; Allred, Jonathan A.; Denner, Jason M.; Johnson, Justin P.; Mullen, Genevieve E.; O'Neal, Trenton K.; Smith, Jason T.; Cardin, Sara E.; Carr, Hank T.; Carr, Stacie L.; Cowart, Holly E.; DaCosta, David H.; Herring, Brendon R.; King, Valeria M.; Polska, Caroline J.; Ward, Erin E.; Wise, Alice A.; McAllister, Kathleen N.; Chevalier, David; Spector, Michael P.; Borchert, Glen M.

    2016-01-01

    ABSTRACT Small RNAs (sRNAs) are short (∼50–200 nucleotides) noncoding RNAs that regulate cellular activities across bacteria. Salmonella enterica starved of a carbon-energy (C) source experience a host of genetic and physiological changes broadly referred to as the starvation-stress response (SSR). In an attempt to identify novel sRNAs contributing to SSR control, we grew log-phase, 5-h C-starved and 24-h C-starved cultures of the virulent Salmonella enterica subspecies enterica serovar Typhimurium strain SL1344 and comprehensively sequenced their small RNA transcriptomes. Strikingly, after employing a novel strategy for sRNA discovery based on identifying dynamic transcripts arising from “gene-empty” regions, we identify 58 wholly undescribed Salmonella sRNA genes potentially regulating SSR averaging an ∼1,000-fold change in expression between log-phase and C-starved cells. Importantly, the expressions of individual sRNA loci were confirmed by both comprehensive transcriptome analyses and northern blotting of select candidates. Of note, we find 43 candidate sRNAs share significant sequence identity to characterized sRNAs in other bacteria, and ∼70% of our sRNAs likely assume characteristic sRNA structural conformations. In addition, we find 53 of our 58 candidate sRNAs either overlap neighboring mRNA loci or share significant sequence complementarity to mRNAs transcribed elsewhere in the SL1344 genome strongly suggesting they regulate the expression of transcripts via antisense base-pairing. Finally, in addition to this work resulting in the identification of 58 entirely novel Salmonella enterica genes likely participating in the SSR, we also find evidence suggesting that sRNAs are significantly more prevalent than currently appreciated and that Salmonella sRNAs may actually number in the thousands. PMID:26853797

  20. Novel small RNA (sRNA) landscape of the starvation-stress response transcriptome of Salmonella enterica serovar typhimurium.

    PubMed

    Amin, Shivam V; Roberts, Justin T; Patterson, Dillon G; Coley, Alexander B; Allred, Jonathan A; Denner, Jason M; Johnson, Justin P; Mullen, Genevieve E; O'Neal, Trenton K; Smith, Jason T; Cardin, Sara E; Carr, Hank T; Carr, Stacie L; Cowart, Holly E; DaCosta, David H; Herring, Brendon R; King, Valeria M; Polska, Caroline J; Ward, Erin E; Wise, Alice A; McAllister, Kathleen N; Chevalier, David; Spector, Michael P; Borchert, Glen M

    2016-01-01

    Small RNAs (sRNAs) are short (∼50-200 nucleotides) noncoding RNAs that regulate cellular activities across bacteria. Salmonella enterica starved of a carbon-energy (C) source experience a host of genetic and physiological changes broadly referred to as the starvation-stress response (SSR). In an attempt to identify novel sRNAs contributing to SSR control, we grew log-phase, 5-h C-starved and 24-h C-starved cultures of the virulent Salmonella enterica subspecies enterica serovar Typhimurium strain SL1344 and comprehensively sequenced their small RNA transcriptomes. Strikingly, after employing a novel strategy for sRNA discovery based on identifying dynamic transcripts arising from "gene-empty" regions, we identify 58 wholly undescribed Salmonella sRNA genes potentially regulating SSR averaging an ∼1,000-fold change in expression between log-phase and C-starved cells. Importantly, the expressions of individual sRNA loci were confirmed by both comprehensive transcriptome analyses and northern blotting of select candidates. Of note, we find 43 candidate sRNAs share significant sequence identity to characterized sRNAs in other bacteria, and ∼70% of our sRNAs likely assume characteristic sRNA structural conformations. In addition, we find 53 of our 58 candidate sRNAs either overlap neighboring mRNA loci or share significant sequence complementarity to mRNAs transcribed elsewhere in the SL1344 genome strongly suggesting they regulate the expression of transcripts via antisense base-pairing. Finally, in addition to this work resulting in the identification of 58 entirely novel Salmonella enterica genes likely participating in the SSR, we also find evidence suggesting that sRNAs are significantly more prevalent than currently appreciated and that Salmonella sRNAs may actually number in the thousands.

  1. Iron-induced virulence of Salmonella enterica serovar typhimurium at the intestinal epithelial interface can be suppressed by carvacrol.

    PubMed

    Kortman, Guus A M; Roelofs, Rian W H M; Swinkels, Dorine W; de Jonge, Marien I; Burt, Sara A; Tjalsma, Harold

    2014-01-01

    Oral iron therapy can increase the abundance of bacterial pathogens, e.g., Salmonella spp., in the large intestine of African children. Carvacrol is a natural compound with antimicrobial activity against various intestinal bacterial pathogens, among which is the highly prevalent Salmonella enterica serovar Typhimurium. This study aimed to explore a presumed interaction between carvacrol and bacterial iron handling and to assess the potential of carvacrol in preventing the increase of bacterial pathogenicity during high iron availability. S. Typhimurium was cultured with increasing concentrations of iron and carvacrol to study the effects of these combined interventions on growth, adhesion to intestinal epithelial cells, and iron uptake/influx in both bacterial and epithelial cells. In addition, the ability of carvacrol to remove iron from the high-affinity ligand transferrin and an Fe-dye complex was examined. Carvacrol retarded growth of S. Typhimurium at all iron conditions. Furthermore, iron-induced epithelial adhesion was effectively reduced by carvacrol at high iron concentrations. The reduction of growth and virulence by carvacrol was not paralleled by a change in iron uptake or influx into S. Typhimurium. In contrast, bioavailability of iron for epithelial cells was moderately decreased under these conditions. Further, carvacrol was shown to lack the properties of an iron binding molecule; however, it was able to weaken iron-ligand interactions by which it may possibly interfere with bacterial virulence. In conclusion, our in vitro data suggest that carvacrol has the potential to serve as a novel dietary supplement to prevent pathogenic overgrowth and colonization in the large intestine during oral iron therapy.

  2. Iron-Induced Virulence of Salmonella enterica Serovar Typhimurium at the Intestinal Epithelial Interface Can Be Suppressed by Carvacrol

    PubMed Central

    Kortman, Guus A. M.; Roelofs, Rian W. H. M.; Swinkels, Dorine W.; de Jonge, Marien I.; Burt, Sara A.

    2014-01-01

    Oral iron therapy can increase the abundance of bacterial pathogens, e.g., Salmonella spp., in the large intestine of African children. Carvacrol is a natural compound with antimicrobial activity against various intestinal bacterial pathogens, among which is the highly prevalent Salmonella enterica serovar Typhimurium. This study aimed to explore a presumed interaction between carvacrol and bacterial iron handling and to assess the potential of carvacrol in preventing the increase of bacterial pathogenicity during high iron availability. S. Typhimurium was cultured with increasing concentrations of iron and carvacrol to study the effects of these combined interventions on growth, adhesion to intestinal epithelial cells, and iron uptake/influx in both bacterial and epithelial cells. In addition, the ability of carvacrol to remove iron from the high-affinity ligand transferrin and an Fe-dye complex was examined. Carvacrol retarded growth of S. Typhimurium at all iron conditions. Furthermore, iron-induced epithelial adhesion was effectively reduced by carvacrol at high iron concentrations. The reduction of growth and virulence by carvacrol was not paralleled by a change in iron uptake or influx into S. Typhimurium. In contrast, bioavailability of iron for epithelial cells was moderately decreased under these conditions. Further, carvacrol was shown to lack the properties of an iron binding molecule; however, it was able to weaken iron-ligand interactions by which it may possibly interfere with bacterial virulence. In conclusion, our in vitro data suggest that carvacrol has the potential to serve as a novel dietary supplement to prevent pathogenic overgrowth and colonization in the large intestine during oral iron therapy. PMID:24379194

  3. Salmonella enterica serovar Typhimurium utilizes the ClpPX and Lon proteases for optimal fitness in the ceca of chickens

    PubMed Central

    Troxell, Bryan

    2016-01-01

    Salmonella enterica serovar Typhimurium (S. Typhimurium) is a leading cause of salmonellosis. Poultry and poultry products are implicated in transmission of Salmonella to humans. In 2013, an outbreak of S. Typhimurium occurred that comprised 39 states within the United States and was associated with backyard flocks of chickens. Colonization of the avian host by S. Typhimurium requires numerous genetic factors encoded within the bacterium. Of particular interest are genetic factors induced by alternative sigma factors within S. Typhimurium since these genetic elements are important for adaptation to different environmental stresses. The heat shock response is a dedicated change in gene regulation within bacteria in response to several stresses, specifically growth at 42°C. Because chickens have a higher body temperature than other animals (42°C) the hypothesis was tested that components of the heat shock response are important for optimal fitness within the chicken. To this end, deletion of the heat shock proteases clpPX (BTNC0022) or lon (BTNC0021) was accomplished and the bacterial fitness in vivo was compared to the “wild-type” strain (NC1040) using a competition assay. One-day-old chicks were orally gavaged with an equal mixture of NC1040 and either BTNC0022 or BTNC0021. Quantification of viable bacteria over time by using plate counts indicated that deletion of either heat shock protease resulted in significantly reduced colonization of the chicken ceca compared to the wild-type strain. To satisfy the molecular Koch's postulates, clpPX and lon mutants were complemented in trans using a low-copy number plasmid for additional in vivo experiments. Complementation studies confirmed the importance of either heat shock protease to colonization of the chicken ceca. This report demonstrated that both ClpPX and Lon were important for optimal fitness within chickens. Moreover, these results suggested that components of the heat shock may be critical factors used

  4. Influence of rpoS mutations on the response of Salmonella enterica serovar Typhimurium to solar radiation.

    PubMed

    Oppezzo, Oscar J; Costa, Cristina S; Pizarro, Ramón A

    2011-01-10

    Salmonella enterica serovar Typhimurium is an important pathogen, and exhibits considerable resistance to the lethal effects of solar radiation. To evaluate the involvement of the RpoS transcription factor in the defense mechanisms of this organism, the sunlight response of a wild type strain (ATCC14028) was compared with that of an rpoS mutant, which exhibited increased sensitivity. Kinetics of cell death was complex in both strains, probably due to the presence of a variety of targets for the radiation. When ultraviolet radiation was excluded from the incident sunlight, lethal effects were abolished independently of the allelic state of rpoS. Reduction of oxygen concentration in the irradiation medium provided moderate protection to ATCC14028, but notably improved survival of the mutant. Similar assays were developed with another S. enterica strain (DA1468), which is a derivative of strain LT2 and produces low levels of RpoS. In this strain the loss of viability reveals the dependence on solar ultraviolet and oxygen concentration found for ATCC14028, but radiation resistance was slightly reduced. Increased sensitivity was observed in an rpoS mutant derived from DA1468, indicating that RpoS functions related to photoprotection are conserved in this strain. In addition, notable differences in the shape of the survival curves obtained for mutants derived from ATCC14028 and DA1468 were found, suggesting that genes beyond RpoS control are relevant in the sunlight response of these mutants.

  5. Characterization of a laboratory-derived, high-level ampicillin-resistant Salmonella enterica serovar Typhimurium strain that caused meningitis in an infant.

    PubMed

    Chiu, Cheng-Hsun; Chu, Chishih; Su, Lin-Hui; Wu, Wan-Yu; Wu, Tsu-Lan

    2002-05-01

    A Salmonella enterica serovar Typhimurium strain that harbored a plasmid carrying a TEM-1-type beta-lactamase gene was isolated from the blood and cerebrospinal fluid of an infant with meningitis. This 3.2-kb plasmid was further characterized to be a nonconjugative pGEM series cloning vector containing a foreign insert. The strain was likely laboratory derived and contaminated the environment before it caused the infection.

  6. Complete Genome Sequence of Salmonella enterica Serovar Typhimurium Strain SO2 (Sequence Type 302) Isolated from an Asymptomatic Child in Mexico

    PubMed Central

    Calva, Edmundo; Puente, José L.; Zaidi, Mussaret B.

    2016-01-01

    The complete genome sequence of Salmonella enterica serovar Typhimurium strain SO2, isolated from an asymptomatic child in Mexico, was determined using PacBio single-molecule real-time technology. Strain SO2 has six complete chromosomal prophages, namely, ST104, Gifsy-2, ST64B, Gifsy-1, ELPhiS, and FSL SP-004, and carries a Salmonella virulence plasmid. PMID:27081133

  7. Scano-magneto immunoassay based on carbon nanotubes/gold nanoparticles nanocomposite for Salmonella enterica serovar Typhimurium detection.

    PubMed

    Amaro, Moegiratul; Oaew, Sukunya; Surareungchai, Werasak

    2012-01-01

    To improve sensitivity of S. enterica serovar Typhimurium detection, multiwalled carbon nanotubes (MWCNTs) and gold nanoparticles (AuNPs) were combined and used as a label to amplify signal in a scanometric based assay. In this study, the MWCNTs/AuNPs nanocomposite was fabricated by directly assemble of Au(3+) to MWCNTs and allowed growing of AuNPs along the MWCNTs surface. This MWCNTs/AuNPs nanocomposite was then attached to anti-S. typhimurium antibody (MWCNTs/AuNPs/Ab(1)) and used as a detecting molecule. Upon binding to Salmonella, they were pre-concentrated by magenetic beads/antibody (MBs/Ab(2)) forming a sandwich immuno-complex which is later spotted on a nitrocellulose membrane coated slide. Silver reduction was applied to amplify signal. The detection limit of 42CFU/ml was achieved when 2% BSA was used as a blocking agent. Given different types of real samples testing, chicken broth was found to give lowest detection limit, followed by orange juice low fat and whole milk. Selectivity testing was performed by using Escherichia coli as interference and found slightly cross-reactivity which could be due to specificity of the Ab used. By virtue of using a slide for multi-samples spotting and a flatbed scanner for signal-read out acquisition, this scano-magneto immunoassay could enable low-cost detection as well as high throughput screening. Copyright © 2012 Elsevier B.V. All rights reserved.

  8. Genomic Analysis of Salmonella enterica Serovar Typhimurium DT160 Associated with a 14-Year Outbreak, New Zealand, 1998–2012

    PubMed Central

    Benschop, Jackie; Biggs, Patrick J.; Marshall, Jonathan C.; Hayman, David T.S.; Carter, Philip E.; Midwinter, Anne C.; Mather, Alison E.; French, Nigel P.

    2017-01-01

    During 1998–2012, an extended outbreak of Salmonella enterica serovar Typhimurium definitive type 160 (DT160) affected >3,000 humans and killed wild birds in New Zealand. However, the relationship between DT160 within these 2 host groups and the origin of the outbreak are unknown. Whole-genome sequencing was used to compare 109 Salmonella Typhimurium DT160 isolates from sources throughout New Zealand. We provide evidence that DT160 was introduced into New Zealand around 1997 and rapidly propagated throughout the country, becoming more genetically diverse over time. The genetic heterogeneity was evenly distributed across multiple predicted functional protein groups, and we found no evidence of host group differentiation between isolates collected from human, poultry, bovid, and wild bird sources, indicating ongoing transmission between these host groups. Our findings demonstrate how a comparative genomic approach can be used to gain insight into outbreaks, disease transmission, and the evolution of a multihost pathogen after a probable point-source introduction. PMID:28516864

  9. Protein folding failure sets high-temperature limit on growth of phage P22 in Salmonella enterica serovar Typhimurium.

    PubMed

    Pope, Welkin H; Haase-Pettingell, Cameron; King, Jonathan

    2004-08-01

    The high-temperature limit for growth of microorganisms differs greatly depending on their species and habitat. The importance of an organism's ability to manage thermal stress is reflected in the ubiquitous distribution of the heat shock chaperones. Although many chaperones function to reduce protein folding defects, it has been difficult to identify the specific protein folding pathways that set the high-temperature limit of growth for a given microorganism. We have investigated this for a simple system, phage P22 infection of Salmonella enterica serovar Typhimurium. Production of infectious particles exhibited a broad maximum of 150 phage per cell when host cells were grown at between 30 and 39 degrees C in minimal medium. Production of infectious phage declined sharply in the range of 40 to 41 degrees C, and at 42 degrees C, production had fallen to less than 1% of the maximum rate. The host cells maintained optimal division rates at these temperatures. The decrease in phage infectivity was steeper than the loss of physical particles, suggesting that noninfectious particles were formed at higher temperatures. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed a decrease in the tailspike adhesins assembled on phage particles purified from cultures incubated at higher temperatures. The infectivity of these particles was restored by in vitro incubation with soluble tailspike trimers. Examination of tailspike folding and assembly in lysates of phage-infected cells confirmed that the fraction of polypeptide chains able to reach the native state in vivo decreased with increasing temperature, indicating a thermal folding defect rather than a particle assembly defect. Thus, we believe that the folding pathway of the tailspike adhesin sets the high-temperature limit for P22 formation in Salmonella serovar Typhimurium.

  10. Salicylate reduces the antimicrobial activity of ciprofloxacin against extracellular Salmonella enterica serovar Typhimurium, but not against Salmonella in macrophages.

    PubMed

    Hartog, Efrat; Menashe, Ofir; Kler, Edna; Yaron, Sima

    2010-05-01

    Salicylate, a potent inducer of the MarA activator in Salmonella enterica, is the principal metabolite of aspirin, which is often consumed for medicinal and cosmetic uses. Our research was aimed at testing if salicylate activates the mar regulon in macrophage-associated Salmonella (intracellular bacteria), and investigating its effects on bacterial susceptibility to ciprofloxacin extracellularly and intracellularly. J774 macrophages were infected with S. enterica serovar Typhimurium (wild-type and marA null mutant), treated with ciprofloxacin with and without pre-exposure to salicylate, and the surviving bacteria were counted. Similar experiments were conducted with bacteria in broth (extracellular bacteria). Phe-Arg-beta-naphthylamide (PAbetaN) was added to investigate the role of efflux pumps in resistance. The transcriptional regulation of marRAB, acrAB and micF in extracellular and intracellular Salmonella Typhimurium with and without salicylate and ciprofloxacin was investigated using green fluorescent protein as a marker protein and quantitative real time PCR. Pre-exposure of Salmonella to salicylate increased the resistance of extracellular but not intracellular bacteria to ciprofloxacin, although salicylate stimulated the expression of mar genes in intracellular and extracellular bacteria. Using marA mutants and the inhibitor PAbetaN, we showed that the improved resistance in extracellular bacteria is derived from the induction of acrAB by salicylate, which is mediated by MarA. In intracellular bacteria, the expression of acrAB is already higher when compared with extracellular cells; therefore, salicylate does not result in significant acrAB induction intracellularly and subsequent resistance enhancement. Results show that conclusions raised from extracellular studies cannot be applied to intracellular bacteria, although the systems have similar functions.

  11. Detection of Multidrug-Resistant Salmonella enterica Serovar Typhimurium Phage Types DT102, DT104, and U302 by Multiplex PCR

    PubMed Central

    Chiu, Cheng-Hsun; Su, Lin-Hui; Chu, Chi-Hong; Wang, Mei-Hwei; Yeh, Chia-Ming; Weill, Francois-Xavier; Chu, Chishih

    2006-01-01

    Salmonella enterica serovar Typhimurium is a common cause of nontyphoidal salmonellosis in humans and animals. Multidrug-resistant serovar Typhimurium phage type DT104, which emerged in the 1990s, has become widely distributed in many countries. A total of 104 clinical isolates of Salmonella serogroup B were collected from three major hospitals in Taiwan during 1997 to 2003 and were examined by a multiplex PCR targeting the resistance genes and the spv gene of the virulence plasmid. A total of 51 isolates (49%) were resistant to all drugs (ACSSuT [resistance to ampicillin, chloramphenicol, streptomycin, sulfonamide, and tetracycline]), and all contained a 1.25-kb PCR fragment of integron that is part of the 43-kb Salmonella genomic island 1 (SGI1). The second group was resistant to SSu (28%), and the third was susceptible to all five drugs (13%). Fifty-nine isolates were serotyped to be serovar Typhimurium by the tube agglutination method using H antisera. The virulence plasmid was found in 54 (91.5%) of the 59 serovar Typhimurium isolates. A majority (94.1%) of the Salmonella serogroup B isolates with the ACSSuT resistance pattern harbored a virulence plasmid. Phage typing identified three major phage types: DT104, DT120, and U302. Analysis of the isolates by pulsed-field gel electrophoresis showed six genotypes. We found two genotypes in DT104 strains, two in DT120, and the other two in U302. The presence of a monophasic serovar (4,5,12:i:−) has added difficulty in the determination of the serovars of multidrug-resistant Salmonella serogroup B isolates. Nevertheless, the multiplex PCR devised in the present study appears to be efficient and useful in the rapid identification of ACSSuT-type serovar Typhimurium with SGI1, irrespective of their phage types. PMID:16825349

  12. Tetracycline accelerates the temporally-regulated invasion response in specific isolates of multidrug-resistant Salmonella enterica serovar Typhimurium

    PubMed Central

    2013-01-01

    Background Multidrug-resistant (MDR) Salmonella isolates are associated with increased morbidity compared to antibiotic-sensitive strains and are an important health and safety concern in both humans and animals. Salmonella enterica serovar Typhimurium is a prevalent cause of foodborne disease, and a considerable number of S. Typhimurium isolates from humans and livestock are resistant to three or more antibiotics. The majority of these MDR S. Typhimurium isolates are resistant to tetracycline, a commonly used and clinically and agriculturally relevant antibiotic. Because exposure of drug-resistant bacteria to antibiotics can affect cellular processes associated with virulence, such as invasion, we investigated the effect tetracycline had on the invasiveness of tetracycline-resistant MDR S. Typhimurium isolates. Results The isolates selected and tested were from two common definitive phage types of S. Typhimurium, DT104 and DT193, and were resistant to tetracycline and at least three other antibiotics. Although Salmonella invasiveness is temporally regulated and normally occurs during late-log growth phase, tetracycline exposure induced the full invasive phenotype in a cell culture assay during early-log growth in several DT193 isolates. No changes in invasiveness due to tetracycline exposure occurred in the DT104 isolates during early-log growth or in any of the isolates during late-log growth. Real-time PCR was used to test expression of the virulence genes hilA, prgH, and invF, and these genes were significantly up-regulated during early-log growth in most isolates due to tetracycline exposure; however, increased virulence gene expression did not always correspond with increased invasion, and therefore was not an accurate indicator of elevated invasiveness. This is the first report to assess DT193 isolates, as well as the early-log growth phase, in response to tetracycline exposure, and it was the combination of both parameters that was necessary to observe the

  13. Proteins from latex of Calotropis procera prevent septic shock due to lethal infection by Salmonella enterica serovar Typhimurium.

    PubMed

    Lima-Filho, José V; Patriota, Joyce M; Silva, Ayrles F B; Filho, Nicodemos T; Oliveira, Raquel S B; Alencar, Nylane M N; Ramos, Márcio V

    2010-06-16

    The latex of Calotropis procera has been used in traditional medicine to treat different inflammatory diseases. The anti-inflammatory activity of latex proteins (LP) has been well documented using different inflammatory models. In this work the anti-inflammatory protein fraction was evaluated in a true inflammatory process by inducing a lethal experimental infection in the murine model caused by Salmonella enterica Subsp. enterica serovar Typhimurium. Experimental Swiss mice were given 0.2 ml of LP (30 or 60 mg/kg) by the intraperitoneal route 24 h before or after lethal challenge (0.2 ml) containing 10(6) CFU/ml of Salmonella Typhimurium using the same route of administration. All the control animals succumbed to infection within 6 days. When given before bacterial inoculums LP prevented the death of mice, which remained in observation until day 28. Even, LP-treated animals exhibited only discrete signs of infection which disappeared latter. LP fraction was also protective when given orally or by subcutaneous route. Histopathological examination revealed that necrosis and inflammatory infiltrates were similar in both the experimental and control groups on days 1 and 5 after infection. LP activity did not clear Salmonella Typhimurium, which was still present in the spleen at approximately 10(4) cells/g of organ 28 days after challenge. However, no bacteria were detected in the liver at this stage. LP did not inhibit bacterial growth in culture medium at all. In the early stages of infection bacteria population was similar in organs and in the peritoneal fluid but drastically reduced in blood. Titration of TNF-alpha in serum revealed no differences between experimental and control groups on days 1 and 5 days after infection while IL-12 was only discretely diminished in serum of experimental animals on day 5. Moreover, cultured macrophages treated with LP and stimulated by LPS released significantly less IL-1beta. LP-treated mice did not succumb to septic shock when

  14. ProP is required for the survival of desiccated Salmonella enterica serovar typhimurium cells on a stainless steel surface.

    PubMed

    Finn, Sarah; Händler, Kristian; Condell, Orla; Colgan, Aoife; Cooney, Shane; McClure, Peter; Amézquita, Aléjandro; Hinton, Jay C D; Fanning, Séamus

    2013-07-01

    Consumers trust commercial food production to be safe, and it is important to strive to improve food safety at every level. Several outbreaks of food-borne disease have been caused by Salmonella strains associated with dried food. Currently we do not know the mechanisms used by Salmonella enterica serovar Typhimurium to survive in desiccated environments. The aim of this study was to discover the responses of S. Typhimurium ST4/74 at the transcriptional level to desiccation on a stainless steel surface and to subsequent rehydration. Bacterial cells were dried onto the same steel surfaces used during the production of dry foods, and RNA was recovered for transcriptomic analysis. Subsequently, dried cells were rehydrated and were again used for transcriptomic analysis. A total of 266 genes were differentially expressed under desiccation stress compared with a static broth culture. The osmoprotectant transporters proP, proU, and osmU (STM1491 to STM1494) were highly upregulated by drying. Deletion of any one of these transport systems resulted in a reduction in the long-term viability of S. Typhimurium on a stainless steel food contact surface. The proP gene was critical for survival; proP deletion mutants could not survive desiccation for long periods and were undetectable after 4 weeks. Following rehydration, 138 genes were differentially expressed, with upregulation observed for genes such as proP, proU, and the phosphate transport genes (pstACS). In time, this knowledge should prove valuable for understanding the underlying mechanisms involved in pathogen survival and should lead to improved methods for control to ensure the safety of intermediate- and low-moisture foods.

  15. Impairment of Swimming Motility by Antidiarrheic Lactobacillus acidophilus Strain LB Retards Internalization of Salmonella enterica Serovar Typhimurium within Human Enterocyte-Like Cells▿

    PubMed Central

    Liévin-Le Moal, Vanessa; Amsellem, Raymonde; Servin, Alain L.

    2011-01-01

    We report that both culture and the cell-free culture supernatant (CFCS) of Lactobacillus acidophilus strain LB (Lactéol Boucard) have the ability (i) to delay the appearance of Salmonella enterica serovar Typhimurium strain SL1344-induced mobilization of F-actin and, subsequently, (ii) to retard cell entry by S. Typhimurium SL1344. Time-lapse imaging and Western immunoblotting showed that S. Typhimurium SL1344 swimming motility, as represented by cell tracks of various types, was rapidly but temporarily blocked without affecting the expression of FliC flagellar propeller protein. We show that the product(s) secreted by L. acidophilus LB that supports the inhibitory activity is heat stable and of low molecular weight. The product(s) caused rapid depolarization of the S. Typhimurium SL1344 cytoplasmic membrane without affecting bacterial viability. We identified inhibition of swimming motility as a newly discovered mechanism by which the secreted product(s) of L. acidophilus strain LB retards the internalization of the diarrhea-associated pathogen S. enterica serovar Typhimurium within cultured human enterocyte-like cells. PMID:21825295

  16. Leveraging management strategies for seedborne plant diseases to reduce Salmonella enterica Serovar Typhimurium incidence on tomato seed and seedlings.

    PubMed

    Lewis Ivey, Melanie L; Xu, Xiulan; Miller, Sally A

    2014-03-01

    Tomatoes have been linked to many outbreaks of salmonellosis over the last decade, but the routes of contamination have yet to be discerned. Many phytopathogens of tomato are seedborne and are effectively managed using seed sanitizers. Seed sanitizers effective against bacterial phytopathogens were evaluated for their efficacy in killing bioluminescent Salmonella enterica serovar Typhimurium strain SeT-A14 on tomato seed infested with moderately high and high levels of pathogen. SeT-A14 incidence on seedlings produced from contaminated seed following sanitation was also determined. At a moderately high infestation rate (40%), SeT-A14 was eradicated on seed sanitized with 1.2% sodium hypochlorite (NaClO) mixed with 0.03% surfactant for 2 min, hydrochloric acid (HCl) for 30 min, and trichloromelamine for 2 min. At a higher infestation rate (94%), only NaClO and HCl were effective in eradicating SeT-A14 from the seed. At both infestation rates, 2% Virkon-S for 15 min significantly reduced SeT-A14 incidence compared with the nontreated infested controls but did not eradicate the pathogen. Hot water, a commonly used sanitizer for managing seedborne bacterial plant diseases, significantly reduced SeT-A14 on heavily infested seed, but incidence was still moderate at 17.5%. On seedlings produced from moderately highly infested seed, SeT-A14 was not detected using RapidChek Salmonella test strips. Using heavily infested seed, SeT-A14 was detected with the test strips in one of four pooled samples of 14-day-old seedlings produced from nonsanitized seed and from seed sanitized with hot water and trichloromelamine. However, bioluminescence was not observed on 14-day-old seedlings. To our knowledge, this is the first report that provides evidence that S. enterica serovar Typhimurium can be seed transmitted and can lead to the contamination of tomato seedlings. In addition to eliminating important bacterial phytopathogens from tomato seed, NaClO or HCl may mitigate the risk of

  17. Salmonella enterica serovar Typhimurium in Mauritius linked to consumption of marlin mousse.

    PubMed

    Issack, Mohammad I; Hendriksen, Rene S; Lun, Phimy Lan Keng; Lutchun, Ram K S; Aarestrup, Frank M

    2009-01-01

    We report the first outbreak of salmonellosis caused by consumption of contaminated marlin mousse. Between 29 October and 5 November 2008, at least 53 persons developed diarrheal illness, all with a history of eating marlin mousse. Salmonella spp. that did not produce gas from glucose was isolated from stools of 26 affected patients and blood culture from one patient. Salmonella sp. isolates with the same phenotype were isolated in three samples of marlin mousse manufactured on 27 October 2008. The constituents of the mousse were smoked marlin, raw eggs, bovine gelatin, oil, and cream. A laboratory investigation of one sample of marlin mousse manufactured 3 days later, and the individual ingredients sampled a week after production of the contaminated batch were all negative for Salmonella. Serotyping and minimum inhibitory concentration determination were performed on 12 patient isolates related to the outbreak and two mousse isolates. All isolates belonged to Salmonella serovar Typhimurium and were pansusceptible to all antimicrobials tested. Pulsed-field gel electrophoresis revealed that all the isolates were indistinguishable, thus implicating the mousse as the vehicle of the outbreak.

  18. Choice of bacterial growth medium alters the transcriptome and phenotype of Salmonella enterica Serovar Typhimurium.

    PubMed

    Blair, Jessica M A; Richmond, Grace E; Bailey, Andrew M; Ivens, Al; Piddock, Laura J V

    2013-01-01

    The type of bacterial culture medium is an important consideration during design of any experimental protocol. The aim of this study was to understand the impact of medium choice on bacterial gene expression and physiology by comparing the transcriptome of Salmonella enterica SL1344 after growth in the widely used LB broth or the rationally designed MOPS minimal medium. Transcriptomics showed that after growth in MOPS minimal media, compared to LB, there was increased expression of 42 genes involved in amino acid synthesis and 23 genes coding for ABC transporters. Seven flagellar genes had decreased expression after growth in MOPS minimal medium and this correlated with a decreased motility. In both MOPS minimal medium and MEM expression of genes from SPI-2 was increased and the adhesion of S. Typhimurium to intestinal epithelial cells was higher compared to the levels after growth in LB. However, SL1344 invasion was not significantly altered by growth in either MOPs minimal media or MEM. Expression of SPI-2 was also measured using chromosomal GFP reporter fusions followed by flow cytometry which showed, for the first time, that the reduction in SPI-2 transcript after growth in different media related to a reduction in the proportion of the bacterial population expressing SPI-2. These data highlight the profound differences in the global transcriptome after in vitro growth in different media and show that choice of medium should be considered carefully during experimental design, particularly when virulence related phenotypes are being measured.

  19. The transcriptional programme of Salmonella enterica serovar Typhimurium reveals a key role for tryptophan metabolism in biofilms

    PubMed Central

    2009-01-01

    Background Biofilm formation enhances the capacity of pathogenic Salmonella bacteria to survive stresses that are commonly encountered within food processing and during host infection. The persistence of Salmonella within the food chain has become a major health concern, as biofilms can serve as a reservoir for the contamination of food products. While the molecular mechanisms required for the survival of bacteria on surfaces are not fully understood, transcriptional studies of other bacteria have demonstrated that biofilm growth triggers the expression of specific sets of genes, compared with planktonic cells. Until now, most gene expression studies of Salmonella have focused on the effect of infection-relevant stressors on virulence or the comparison of mutant and wild-type bacteria. However little is known about the physiological responses taking place inside a Salmonella biofilm. Results We have determined the transcriptomic and proteomic profiles of biofilms of Salmonella enterica serovar Typhimurium. We discovered that 124 detectable proteins were differentially expressed in the biofilm compared with planktonic cells, and that 10% of the S. Typhimurium genome (433 genes) showed a 2-fold or more change in the biofilm compared with planktonic cells. The genes that were significantly up-regulated implicated certain cellular processes in biofilm development including amino acid metabolism, cell motility, global regulation and tolerance to stress. We found that the most highly down-regulated genes in the biofilm were located on Salmonella Pathogenicity Island 2 (SPI2), and that a functional SPI2 secretion system regulator (ssrA) was required for S. Typhimurium biofilm formation. We identified STM0341 as a gene of unknown function that was needed for biofilm growth. Genes involved in tryptophan (trp) biosynthesis and transport were up-regulated in the biofilm. Deletion of trpE led to decreased bacterial attachment and this biofilm defect was restored by exogenous

  20. Expression of Toll-like receptors, interleukin 8, macrophage migration inhibitory factor, and osteopontin in tissues from pigs challenged with Salmonella enterica serovar Typhimurium or serovar Choleraesuis.

    PubMed

    Burkey, T E; Skjolaas, K A; Dritz, S S; Minton, J E

    2007-02-15

    Two serovars of Salmonella enterica, namely serovar Typhimurium (ST) and serovar Choleraesuis (SC) account for the vast majority of clinical cases of swine salmonellosis worldwide. These serovars are thought to be transmitted among pigs in production settings mainly through fecal-oral routes. Yet, few studies have evaluated effects of these serovars on expression of innate immune targets when presented to pigs via repeated oral dosing in an attempt to model transmission in production settings. Thus, a primary objective of the current experiments was to evaluate expression of Toll-like receptors (TLR) and selected chemoattractive mediators (interleukin 8, IL8; macrophage migration inhibitory factor, MIF; osteopontin, OPN) in tissues from pigs exposed to ST or SC that had been transformed with kanamycin resistance and green (STG) or red (SCR) fluorescent protein to facilitate isolation from pen fecal samples. In vitro studies confirmed that STG and SCR largely (though not completely) retained their ability to upregulate IL8 and CC chemokine ligand 20 (CCL20) in cultured swine jejunal epithelial cells. Transformed bacteria were then fed to pigs in an in vivo study to determine tissue specific effects on mRNA relative expression. Pigs were fed cookie dough inoculated with bacteria on days 0, 3, 7, and 10 with 10(8)CFU STG (n=8) or SCR (n=8), while control (CTL) pigs (n=8) received dough without bacteria. Animals were sacrificed 14 days from the initial bacterial challenge and samples of tonsil, jejunum, ileum, colon, mesenteric lymph node (MLN), spleen, and liver were removed for subsequent RNA isolation. Expression of mRNA in tissues was determined using real-time quantitative PCR and expressed relative to 18S rRNA. Within CTL pigs, when expressed relative to the content in liver, mRNA for all targets demonstrated substantial tissue effects (P<0.001 for all TLR; MIF, and OPN; P<0.05 for IL8). Feeding STG and SCR resulted in significant (P

  1. Characterisation of recently emerged multiple antibiotic-resistant Salmonella enterica serovar typhimurium DT104 and other multiresistant phage types from Danish pig herds.

    PubMed

    Baggesen, D L; Aarestrup, F M

    1998-07-25

    A total of 670 isolates of Salmonella enterica were isolated from Danish pig herds, phage typed and tested for susceptibility to amoxycillin + clavulanate, ampicillin, colistin, enrofloxacin, gentamicin, neomycin, spectinomycin, streptomycin, tetracyclines, and trimethoprim + sulphadiazine. S enterica serovar typhimurium (S typhimurium) isolates resistant to ampicillin, streptomycin and tetracycline and three isolates of S typhimurium DT104, two from 1994 and one from 1995, were further tested for resistance against chloramphenicol and sulphonamide and analysed by pulsed-field gel electrophoresis (PFGE) using the restriction enzyme Xba I. Overall, 66 per cent of the 670 isolates were sensitive to all the antimicrobial agents tested. Eleven isolates of S typhimurium were resistant to ampicillin, streptomycin and tetracycline and also resistant to other antibiotics in different resistance patterns. Seven different multiresistant clones were identified. The most common clones were four isolates of DT104 and three isolates of DT193. Two of the three S typhimurium DT104 from 1994 and 1995 were sensitive to all the antimicrobials tested whereas the remaining isolate from 1994 was resistant to spectinomycin, streptomycin and sulphonamides. All three isolates showed PFGF profiles identical to the four multiresistant DT104 isolates. Compared with most other countries antimicrobial resistance among S enterica isolated from Danish pig herds is uncommon. However, several different multiresistant clones were found.

  2. [Characterisation of a phenotypic monophasic variant belonging to Salmonella enterica subsp. enterica serovar Typhimurium from wild birds and its possible transmission to cats and humans].

    PubMed

    Hauser, Elisabeth; Hühn, Stephan; Junker, Ernst; Jaber, Manuela; Schroeter, Andreas; Helmuth, Reiner; Rabsch, Wolfgang; Winterhoff, Nora; Malorny, Burkhard

    2009-01-01

    In the last two years The National Salmonella Reference Laboratory (NRL-Salm) received an accumulating number of salmonellae with sero-formula 4,12:-:1,2 isolated from perished wild birds, particularly siskins. Within these strains flagellar antigen of the first phase was phenotypically not detectable. By PCR a fragment could be amplified coding specifically for the H:i-flagellar antigen. Consequently, this is a phenotypically monophasic variant of Salmonella enterica subsp. enterica serovar Typhimurium var. copenhagen with the sero-formula 4,12:-:1,2. Phage-typing showed most of the isolates belonged to phage type DT40. Some other strains harboured the same lysis pattern but could not assign to a definite phage type. Those strains are designated as RDNC (react with phages but does not conform with definite or provisorial types). Pulsed field gel-electrophoresis (PFGE) confirmed those two lineages of the monophasic variant. Phage type DT40 isolates from humans or cats, able to express both flagellar antigens, did not differ in genotypic properties from those not able to express the flagellar antigen of the first phase. Salmonella strains with identical genotypic characteristics have been isolated from wild birds, human cases particular infants and also cats. This refers to a direct or indirect transmission of the pathogen from wild bird to human. By eating or getting in contact with contaminated birds domestic cats could play an important role as vehicle between bird and human.

  3. Internalisation potential of Escherichia coli O157:H7, Listeria monocytogenes, Salmonella enterica subsp. enterica serovar Typhimurium and Staphylococcus aureus in lettuce seedlings and mature plants.

    PubMed

    Standing, Taryn-Ann; du Plessis, Erika; Duvenage, Stacey; Korsten, Lise

    2013-06-01

    The internalisation potential of Listeria monocytogenes, Staphylococcus aureus, Escherichia coli O157:H7 and Salmonella enterica subsp. enterica serovar Typhimurium in lettuce was evaluated using seedlings grown in vermiculite in seedling trays as well as hydroponically grown lettuce. Sterile distilled water was spiked with one of the four human pathogenic bacteria (10(5) CFU/mL) and used to irrigate the plants. The potential for pathogen internalisation was investigated over time using light microscopy, transmission electron microscopy and viable plate counts. Additionally, the identities of the pathogens isolated from internal lettuce plant tissues were confirmed using polymerase chain reaction with pathogen-specific oligonucleotides. Internalisation of each of the human pathogens was evident in both lettuce seedlings and hydroponically grown mature lettuce plants. To our knowledge, this is the first report of S. aureus internalisation in lettuce plants. In addition, the levels of background microflora in the lettuce plants were determined by plate counting and the isolates identified using matrix-assisted laser ionisation-time of flight (MALDI-TOF). Background microflora assessments confirmed the absence of the four pathogens evaluated in this study. A low titre of previously described endophytes and soil inhabitants, i.e., Enterobacter cloacae, Enterococcus faecalis, Lysinibacillus fusiformis, Rhodococcus rhodochrous, Staphylococcus epidermidis and Staphylococcus hominis were identified.

  4. Effects of leachate from crumb rubber and zinc in green roofs on the survival, growth, and resistance characteristics of Salmonella enterica subsp. enterica serovar Typhimurium.

    PubMed

    Crampton, Mollee; Ryan, Allayna; Eckert, Cori; Baker, Katherine H; Herson, Diane S

    2014-05-01

    The use of green roofs is a growing practice worldwide, particularly in densely populated areas. In an attempt to find new methods for recycling crumb rubber, incorporation of crumb rubber into artificial medium for plant growth in green roofs and similar engineered environments has become an attractive option for the recycling of waste tires. Though this approach decreases waste in landfills, there are concerns about the leaching of zinc and other heavy metals, as well as nutrient and organic compounds, into the environment. The present study analyzed the impact of leachate from crumb rubber and zinc on the growth and viability of Salmonella enterica subsp. enterica serovar Typhimurium. Zinc was chosen for further studies since it has been previously implicated with other biological functions, including biofilm formation, motility, and possible cross-resistance to antimicrobial agents. The study showed that Salmonella can colonize crumb rubber and that crumb rubber extract may provide nutrients that are usable by this bacterium. Salmonella strains with reduced susceptibility (SRS) to zinc were obtained after subculturing in increasing concentrations of zinc. The SRS exhibited differences in gene expression of flux pump genes zntA and znuA compared to that of the parent when exposed to 20 mM added zinc. In biofilm formation studies, the SRS formed less biofilm but was more motile than the parental strain.

  5. Effects of Leachate from Crumb Rubber and Zinc in Green Roofs on the Survival, Growth, and Resistance Characteristics of Salmonella enterica subsp. enterica Serovar Typhimurium

    PubMed Central

    Crampton, Mollee; Ryan, Allayna; Eckert, Cori; Baker, Katherine H.

    2014-01-01

    The use of green roofs is a growing practice worldwide, particularly in densely populated areas. In an attempt to find new methods for recycling crumb rubber, incorporation of crumb rubber into artificial medium for plant growth in green roofs and similar engineered environments has become an attractive option for the recycling of waste tires. Though this approach decreases waste in landfills, there are concerns about the leaching of zinc and other heavy metals, as well as nutrient and organic compounds, into the environment. The present study analyzed the impact of leachate from crumb rubber and zinc on the growth and viability of Salmonella enterica subsp. enterica serovar Typhimurium. Zinc was chosen for further studies since it has been previously implicated with other biological functions, including biofilm formation, motility, and possible cross-resistance to antimicrobial agents. The study showed that Salmonella can colonize crumb rubber and that crumb rubber extract may provide nutrients that are usable by this bacterium. Salmonella strains with reduced susceptibility (SRS) to zinc were obtained after subculturing in increasing concentrations of zinc. The SRS exhibited differences in gene expression of flux pump genes zntA and znuA compared to that of the parent when exposed to 20 mM added zinc. In biofilm formation studies, the SRS formed less biofilm but was more motile than the parental strain. PMID:24584242

  6. Characterization of small ColE1-like plasmids conferring kanamycin resistance in Salmonella enterica subsp. enterica serovars Typhimurium and Newport.

    PubMed

    Chen, Chin-Yi; Strobaugh, Terence P; Frye, Jonathan G

    2010-05-01

    Multi-antibiotic resistant (MR) Salmonella enterica serovars Typhimurium and Newport are an increasing concern in human and animal health. Many strains are known to carry antibiotic resistance determinants on multiple plasmids, yet detailed information has been scarce. Three plasmids conferring kanamycin (Kan) resistance were isolated and nucleotide sequences were determined. Two Kan(R) plasmids from Salmonella Newport strains, pSN11/00Kan and pSN02/01Kan, were found to be identical and were 5698bp in size. Plasmid pG7601Kan from Salmonella Typhimurium phage type U302 strain G7601 was 3208bp, and was the same as the previously reported pU302S from another U302 strain G8430. All three plasmids carried identical aph(3')-I genes. The plasmids were ColE1-like, containing RNA I/RNA II and the rom gene. Plasmids pSN11/00Kan and pSN02/01Kan also carried mobilization genes mobC and mobABD, similar to those of the pColK-K235 and pColD-157 plasmids from the colicinogenic Escherichia coli strains. All three plasmids were stable without kanamycin selection for approximately 100 generations.

  7. Inhibition of Intracellular Growth of Salmonella enterica Serovar Typhimurium in Tissue Culture by Antisense Peptide-Phosphorodiamidate Morpholino Oligomer ▿

    PubMed Central

    Mitev, Georgi M.; Mellbye, Brett L.; Iversen, Patrick L.; Geller, Bruce L.

    2009-01-01

    Two types of phosphorodiamidate morpholino oligomers (PMOs) were tested for inhibition of growth of Salmonella enterica serovar Typhimurium. Both PMOs have the same 11-base sequence that is antisense to the region near the start codon of acpP, which is essential for lipid biosynthesis and viability. To the 3′ end of each is attached the membrane-penetrating peptide (RXR)4XB (R, X, and B indicate arginine, 6-aminohexanoic acid, and β-alanine, respectively). One peptide-PMO (AcpP PPMO) has no charge on the PMO moiety. The second PPMO has three cations (piperazine) attached to the phosphorodiamidate linkages (3+Pip-AcpP PPMO). A scrambled-sequence PPMO (Scr PPMO) was synthesized for each type of PMO. The MICs of AcpP PPMO, 3+Pip-AcpP PPMO, and either one of the Scr PPMOs were 1.25 μM (7 μg/ml), 0.156 μM (0.94 μg/ml), and >160 μM (>900 μg/ml), respectively. 3+Pip-AcpP PPMO at 1.25 or 2.5 μM significantly reduced the growth rates of pure cultures, whereas AcpP PPMO or either Scr PPMO had no effect. However, the viable cell count was significantly reduced at either concentration of 3+Pip-AcpP PPMO or AcpP PPMO, but not with either Scr PPMO. In other experiments, macrophages were infected intracellularly with S. enterica and treated with 3 μM 3+Pip-AcpP PPMO. Intracellular bacteria were reduced >99% with 3+Pip-AcpP PPMO, whereas intracellular bacteria increased 3 orders of magnitude in untreated or Scr PPMO-treated cultures. We conclude that either AcpP PPMO or 3+Pip-AcpP PPMO inhibited growth of S. enterica in pure culture and that 3+Pip-AcpP PPMO reduced intracellular viability of S. enterica in macrophages. PMID:19581453

  8. Structural and functional characterization of Salmonella enterica serovar Typhimurium YcbL: An unusual Type II glyoxalase

    PubMed Central

    Stamp, Anna L; Owen, Paul; Omari, Kamel El; Nichols, Charles E; Lockyer, Michael; Lamb, Heather K; Charles, Ian G; Hawkins, Alastair R; Stammers, David K

    2010-01-01

    YcbL has been annotated as either a metallo-β-lactamase or glyoxalase II (GLX2), both members of the zinc metallohydrolase superfamily, that contains many enzymes with a diverse range of activities. Here, we report crystallographic and biochemical data for Salmonella enterica serovar Typhimurium YcbL that establishes it as GLX2, which differs in certain structural and functional properties compared with previously known examples. These features include the insertion of an α-helix after residue 87 in YcbL and truncation of the C-terminal domain, which leads to the loss of some recognition determinants for the glutathione substrate. Despite these changes, YcbL has robust GLX2 activity. A further difference is that the YcbL structure contains only a single bound metal ion rather than the dual site normally observed for GLX2s. Activity assays in the presence of various metal ions indicate an increase in activity above basal levels in the presence of manganous and ferrous ions. Thus, YcbL represents a novel member of the GLX2 family. PMID:20669241

  9. Human milk mucin 1 and mucin 4 inhibit Salmonella enterica serovar Typhimurium invasion of human intestinal epithelial cells in vitro.

    PubMed

    Liu, Bo; Yu, Zhuoteng; Chen, Ceng; Kling, David E; Newburg, David S

    2012-08-01

    Many human milk glycans inhibit pathogen binding to host receptors and their consumption by infants is associated with reduced risk of disease. Salmonella infection is more frequent among infants than among the general population, but the incidence is lower in breast-fed babies, suggesting that human milk could contain components that inhibit Salmonella. This study aimed to test whether human milk per se inhibits Salmonella invasion of human intestinal epithelial cells in vitro and, if so, to identify the milk components responsible for inhibition. Salmonella enterica serovar Typhimurium SL1344 (SL1344) invasion of FHs 74 Int and Caco-2 cells were the models of human intestinal epithelium infection. Internalization of fluorescein-5-isothiocyanate-labeled SL1344 into intestinal cells was measured by flow cytometry to quantify infection. Human milk and its fractions inhibited infection; the inhibitory activity localized to the high molecular weight glycans. Mucin 1 and mucin 4 were isolated to homogeneity. At 150 μg/L, a typical concentration in milk, human milk mucin 1 and mucin 4 inhibited SL1344 invasion of both target cell types. These mucins inhibited SL1344 invasion of epithelial cells in a dose-dependent manner. Thus, mucins may prove useful as a basis for developing novel oral prophylactic and therapeutic agents that inhibit infant diseases caused by Salmonella and related pathogens.

  10. Transcriptomic Analysis of the Swarm Motility Phenotype of Salmonella enterica Serovar Typhimurium Mutant Defective in Periplasmic Glucan Synthesis.

    PubMed

    Bhagwat, Arvind A; Young, Lynn; Smith, Allen D; Bhagwat, Medha

    2017-09-01

    Movement of food-borne pathogens on moist surfaces enables them to migrate towards more favorable niches and facilitate their survival for extended periods of time. Salmonella enterica serovar Typhimurium mutants defective in Osmoregulated periplasmic glucans (OPG) synthesis are unable to exhibit motility on moist surfaces (swarming); however, their mobility in liquid (swim motility) remains unaffected. In order to understand the role of OPG in swarm motility, transcriptomic analysis was performed using cells growing on a moist agar surface. In opgGH deletion mutant, lack of OPG significantly altered transcription of 1039 genes out of total 4712 genes (22%). Introduction of a plasmid-borne copy of opgGH into opgGH deletion mutant restored normal expression of all but 30 genes, indicating a wide-range influence of OPG on gene expression under swarm motility condition. Major pathways that were differentially expressed in opgGH mutants were motility, virulence and invasion, and genes related to the secondary messenger molecule, cyclic di-GMP. These observations provide insights and help explain the pleiotropic nature of OPG mutants such as sub-optimal virulence and competitive organ colonization in mice, biofilm formation, and sensitivity towards detergent stress.

  11. Changes in ultrastructure and Fourier transform infrared spectrum of Salmonella enterica serovar typhimurium cells after exposure to stress conditions.

    PubMed

    Alvarez-Ordóñez, A; Prieto, M

    2010-11-01

    The effect of exposure to acid (pH 2.5), alkaline (pH 11.0), heat (55°C), and oxidative (40 mM H₂O₂) lethal conditions on the ultrastructure and global chemical composition of Salmonella enterica serovar Typhimurium CECT 443 cells was studied using transmission electron microscopy and Fourier transform infrared spectroscopy (FT-IR) combined with multivariate statistical methods (hierarchical cluster analysis and factor analysis). Infrared spectra exhibited marked differences in the five spectral regions for all conditions tested compared to those of nontreated control cells, which suggests the existence of a complex bacterial stress response in which modifications in a wide variety of cellular compounds are involved. The visible spectral changes observed in all of the spectral regions, together with ultrastructural changes observed by transmission electron microscopy and data obtained from membrane integrity tests, indicate the existence of membrane damage or alterations in membrane composition after heat, acid, alkaline, and oxidative treatments. Results obtained in this study indicate the potential of FT-IR spectroscopy to discriminate between intact and injured bacterial cells and between treatment technologies, and they show the adequacy of this technique to study the molecular aspects of bacterial stress response.

  12. Discovery of Novel Secreted Virulence Factors from Salmonella enterica Serovar Typhimurium by Proteomic Analysis of Culture Supernatants

    SciTech Connect

    Niemann, George; Brown, Roslyn N.; Gustin, Jean K.; Stufkens, Afke; Shaikh-Kidwai, Afshan S.; Li, Jie; McDermott, Jason E.; Brewer, Heather M.; Schepmoes, Athena A.; Smith, Richard D.; Adkins, Joshua N.; Heffron, Fred

    2011-01-01

    The intracellular pathogen Salmonella enterica serovar Typhimurium is a leading cause of acute gastroenteritis in the world. This pathogen has two type-III secretion systems (TTSS) necessary for virulence that are encoded in Salmonella pathogenicity islands 1 and 2 (SPI-1 and SPI-2) and are expressed during extracellular or intracellular infectious states, respectively, to deliver virulence factors (effectors) to the host cell cytoplasm. While many have been identified and at least partially characterized, the full repertoire of effectors has not been catalogued. In this mass spectrometry-based proteomics study, we identified effector proteins secreted under minimal acidic medium growth conditions that induced the SPI-2 TTSS and its effectors, and compared the secretome from the parent strain to the secretome from strains missing either essential (SsaK) or regulatory components (SsaL) of the SPI-2 secretion apparatus. We identified 75% of the known TTSS effector repertoire. Excluding translocon components, 95% of the known effectors were biased for identification in the ssaL mutant background, which demonstrated that SsaL regulates SPI-2 type III secretion. To confirm secretion to animal cells, we made translational fusions of several of the best candidates to the calmodulin-dependent adenylate cyclase of Bordetella pertussis and assayed cAMP levels of infected J774 macrophage-like cells. From these infected cells we identified six new TTSS effectors and two others that are secreted independent of TTSS. Our results substantiate reports of additional secretion systems encoded by Salmonella other than TTSS.

  13. Changes in Ultrastructure and Fourier Transform Infrared Spectrum of Salmonella enterica Serovar Typhimurium Cells after Exposure to Stress Conditions ▿

    PubMed Central

    Alvarez-Ordóñez, A.; Prieto, M.

    2010-01-01

    The effect of exposure to acid (pH 2.5), alkaline (pH 11.0), heat (55°C), and oxidative (40 mM H2O2) lethal conditions on the ultrastructure and global chemical composition of Salmonella enterica serovar Typhimurium CECT 443 cells was studied using transmission electron microscopy and Fourier transform infrared spectroscopy (FT-IR) combined with multivariate statistical methods (hierarchical cluster analysis and factor analysis). Infrared spectra exhibited marked differences in the five spectral regions for all conditions tested compared to those of nontreated control cells, which suggests the existence of a complex bacterial stress response in which modifications in a wide variety of cellular compounds are involved. The visible spectral changes observed in all of the spectral regions, together with ultrastructural changes observed by transmission electron microscopy and data obtained from membrane integrity tests, indicate the existence of membrane damage or alterations in membrane composition after heat, acid, alkaline, and oxidative treatments. Results obtained in this study indicate the potential of FT-IR spectroscopy to discriminate between intact and injured bacterial cells and between treatment technologies, and they show the adequacy of this technique to study the molecular aspects of bacterial stress response. PMID:20851964

  14. Exposure of Salmonella enterica Serovar Typhimurium to High Level Biocide Challenge Can Select Multidrug Resistant Mutants in a Single Step

    PubMed Central

    Whitehead, Rebekah N.; Overton, Tim W.; Kemp, Caroline L.; Webber, Mark A.

    2011-01-01

    Background Biocides are crucial to the prevention of infection by bacteria, particularly with the global emergence of multiply antibiotic resistant strains of many species. Concern has been raised regarding the potential for biocide exposure to select for antibiotic resistance due to common mechanisms of resistance, notably efflux. Methodology/Principal Findings Salmonella enterica serovar Typhimurium was challenged with 4 biocides of differing modes of action at both low and recommended-use concentration. Flow cytometry was used to investigate the physiological state of the cells after biocide challenge. After 5 hours exposure to biocide, live cells were sorted by FACS and recovered. Cells recovered after an exposure to low concentrations of biocide had antibiotic resistance profiles similar to wild-type cells. Live cells were recovered after exposure to two of the biocides at in-use concentration for 5 hours. These cells were multi-drug resistant and accumulation assays demonstrated an efflux phenotype of these mutants. Gene expression analysis showed that the AcrEF multidrug efflux pump was de-repressed in mutants isolated from high-levels of biocide. Conclusions/Significance These data show that a single exposure to the working concentration of certain biocides can select for mutant Salmonella with efflux mediated multidrug resistance and that flow cytometry is a sensitive tool for identifying biocide tolerant mutants. The propensity for biocides to select for MDR mutants varies and this should be a consideration when designing new biocidal formulations. PMID:21829527

  15. Dormant Intracellular Salmonella enterica Serovar Typhimurium Discriminates among Salmonella Pathogenicity Island 2 Effectors To Persist inside Fibroblasts

    PubMed Central

    Núñez-Hernández, Cristina; Alonso, Ana; Pucciarelli, M. Graciela; Casadesús, Josep

    2014-01-01

    Salmonella enterica uses effector proteins delivered by type III secretion systems (TTSS) to colonize eukaryotic cells. Recent in vivo studies have shown that intracellular bacteria activate the TTSS encoded by Salmonella pathogenicity island-2 (SPI-2) to restrain growth inside phagocytes. Growth attenuation is also observed in vivo in bacteria colonizing nonphagocytic stromal cells of the intestinal lamina propria and in cultured fibroblasts. SPI-2 is required for survival of nongrowing bacteria persisting inside fibroblasts, but its induction mode and the effectors involved remain unknown. Here, we show that nongrowing dormant intracellular bacteria use the two-component system OmpR-EnvZ to induce SPI-2 expression and the PhoP-PhoQ system to regulate the time at which induction takes place, 2 h postentry. Dormant bacteria were shown to discriminate the usage of SPI-2 effectors. Among the effectors tested, SseF, SseG, and SseJ were required for survival, while others, such as SifA and SifB, were not. SifA and SifB dispensability correlated with the inability of intracellular bacteria to secrete these effectors even when overexpressed. Conversely, SseJ overproduction resulted in augmented secretion and exacerbated bacterial growth. Dormant bacteria produced other effectors, such as PipB and PipB2, that, unlike what was reported for epithelial cells, did not to traffic outside the phagosomal compartment. Therefore, permissiveness for secreting only a subset of SPI-2 effectors may be instrumental for dormancy. We propose that the S. enterica serovar Typhimurium nonproliferative intracellular lifestyle is sustained by selection of SPI-2 effectors that are produced in tightly defined amounts and delivered to phagosome-confined locations. PMID:24144726

  16. Effect of chlorate, molybdate, and shikimic acid on Salmonella enterica serovar Typhimurium in aerobic and anaerobic cultures.

    PubMed

    Oliver, Christy E; Beier, Ross C; Hume, Michael E; Horrocks, Shane M; Casey, Thomas A; Caton, Joel S; Nisbet, David J; Smith, David J; Krueger, Nathan A; Anderson, Robin C

    2010-04-01

    Experiments were conducted to determine factors that affect sensitivity of Salmonella enterica serovar Typhimurium to sodium chlorate (5mM). In our first experiment, cultures grown without chlorate grew more rapidly than those with chlorate. An extended lag before logarithmic growth was observed in anaerobic but not aerobic cultures containing chlorate. Chlorate inhibition of growth during aerobic culture began later than that observed in anaerobic cultures but persisted once inhibition was apparent. Conversely, anaerobic cultures appeared to adapt to chlorate after approximately 10h of incubation, exhibiting rapid compensatory growth. In anaerobic chlorate-containing cultures, 20% of total viable counts were resistant to chlorate by 6h and had propagated to 100% resistance (>10(9)CFU mL(-1)) by 24h. In the aerobic chlorate-containing cultures, 12.9% of colonies had detectable resistance to chlorate by 6h, but only 1% retained detectable resistance at 24h, likely because these cultures had opportunity to respire on oxygen and were thus not enriched via the selective pressure of chlorate. In another study, treatment with shikimic acid (0.34 mM), molybdate (1mM) or their combination had little effect on aerobic or anaerobic growth of Salmonella in the absence of added chlorate. As observed in our earlier study, chlorate resistance was not detected in any cultures without added chlorate. Chlorate resistant Salmonella were recovered at equivalent numbers regardless of treatment after 8h of aerobic or anaerobic culture with added chlorate; however, by 24h incubation chlorate sensitivity was completely restored to aerobic but not anaerobic cultures treated with shikimic acid or molybdate but not their combination. Results indicate that anaerobic adaptation of S. Typhimurium to sodium chlorate during pure culture is likely due to the selective propagation of low numbers of cells exhibiting spontaneous resistance to chlorate and this resistance is not reversible by

  17. Structural and Functional Characterization of ScsC, a Periplasmic Thioredoxin-Like Protein from Salmonella enterica Serovar Typhimurium

    PubMed Central

    Shepherd, Mark; Heras, Begoña; Achard, Maud E. S.; King, Gordon J.; Argente, M. Pilar; Kurth, Fabian; Taylor, Samantha L.; Howard, Mark J.; King, Nathan P.

    2013-01-01

    Abstract Aims: The prototypical protein disulfide bond (Dsb) formation and protein refolding pathways in the bacterial periplasm involving Dsb proteins have been most comprehensively defined in Escherichia coli. However, genomic analysis has revealed several distinct Dsb-like systems in bacteria, including the pathogen Salmonella enterica serovar Typhimurium. This includes the scsABCD locus, which encodes a system that has been shown via genetic analysis to confer copper tolerance, but whose biochemical properties at the protein level are not defined. The aim of this study was to provide functional insights into the soluble ScsC protein through structural, biochemical, and genetic analyses. Results: Here we describe the structural and biochemical characterization of ScsC, the soluble DsbA-like component of this system. Our crystal structure of ScsC reveals a similar overall fold to DsbA, although the topology of β-sheets and α-helices in the thioredoxin domains differ. The midpoint reduction potential of the CXXC active site in ScsC was determined to be −132 mV versus normal hydrogen electrode. The reactive site cysteine has a low pKa, typical of the nucleophilic cysteines found in DsbA-like proteins. Deletion of scsC from S. Typhimurium elicits sensitivity to copper (II) ions, suggesting a potential involvement for ScsC in disulfide folding under conditions of copper stress. Innovation and Conclusion: ScsC is a novel disulfide oxidoreductase involved in protection against copper ion toxicity. Antioxid. Redox Signal. 19, 1494–1506. PMID:23642141

  18. Exposure of Salmonella enterica Serovar Typhimurium to Three Humectants Used in the Food Industry Induces Different Osmoadaptation Systems.

    PubMed

    Finn, Sarah; Rogers, Lisa; Händler, Kristian; McClure, Peter; Amézquita, Alejandro; Hinton, Jay C D; Fanning, Séamus

    2015-10-01

    Common salt (NaCl) is frequently used by the food industry to add flavor and to act as a humectant in order to reduce the water content of a food product. The improved health awareness of consumers is leading to a demand for food products with reduced salt content; thus, manufacturers require alternative water activity-reducing agents which elicit the same general effects as NaCl. Two examples include KCl and glycerol. These agents lower the water activity of a food matrix and also contribute to limit the growth of the microbiota, including foodborne pathogens. Little is currently known about how foodborne pathogens respond to these water activity-lowering agents. Here we examined the response of Salmonella enterica serovar Typhimurium 4/74 to NaCl, KCl, and glycerol at three time points, using a constant water activity level, compared with the response of a control inoculum. All conditions induced the upregulation of gluconate metabolic genes after 6 h of exposure. Bacteria exposed to NaCl and KCl demonstrated the upregulation of the osmoprotective transporter mechanisms encoded by the proP, proU, and osmU (STM1491 to STM1494) genes. Glycerol exposure elicited the downregulation of these osmoadaptive mechanisms but stimulated an increase in lipopolysaccharide and membrane protein-associated genes after 1 h. The most extensive changes in gene expression occurred following exposure to KCl. Because many of these genes were of unknown function, further characterization may identify KCl-specific adaptive processes that are not stimulated by NaCl. This study shows that the response of S. Typhimurium to different humectants does not simply reflect reduced water activity and likely involves systems that are linked to specific humectants. Copyright © 2015 Finn et al.

  19. Loss of Multicellular Behavior in Epidemic African Nontyphoidal Salmonella enterica Serovar Typhimurium ST313 Strain D23580

    PubMed Central

    Singletary, Larissa A.; Karlinsey, Joyce E.; Libby, Stephen J.; Mooney, Jason P.; Lokken, Kristen L.; Tsolis, Renée M.; Byndloss, Mariana X.; Hirao, Lauren A.; Gaulke, Christopher A.; Crawford, Robert W.; Dandekar, Satya; Kingsley, Robert A.; Msefula, Chisomo L.; Heyderman, Robert S.

    2016-01-01

    ABSTRACT Nontyphoidal Salmonella enterica serovar Typhimurium is a frequent cause of bloodstream infections in children and HIV-infected adults in sub-Saharan Africa. Most isolates from African patients with bacteremia belong to a single sequence type, ST313, which is genetically distinct from gastroenteritis-associated ST19 strains, such as 14028s and SL1344. Some studies suggest that the rapid spread of ST313 across sub-Saharan Africa has been facilitated by anthroponotic (person-to-person) transmission, eliminating the need for Salmonella survival outside the host. While these studies have not ruled out zoonotic or other means of transmission, the anthroponotic hypothesis is supported by evidence of extensive genomic decay, a hallmark of host adaptation, in the sequenced ST313 strain D23580. We have identified and demonstrated 2 loss-of-function mutations in D23580, not present in the ST19 strain 14028s, that impair multicellular stress resistance associated with survival outside the host. These mutations result in inactivation of the KatE stationary-phase catalase that protects high-density bacterial communities from oxidative stress and the BcsG cellulose biosynthetic enzyme required for the RDAR (red, dry, and rough) colonial phenotype. However, we found that like 14028s, D23580 is able to elicit an acute inflammatory response and cause enteritis in mice and rhesus macaque monkeys. Collectively, these observations suggest that African S. Typhimurium ST313 strain D23580 is becoming adapted to an anthroponotic mode of transmission while retaining the ability to infect and cause enteritis in multiple host species. PMID:26933058

  20. Exposure of Salmonella enterica Serovar Typhimurium to Three Humectants Used in the Food Industry Induces Different Osmoadaptation Systems

    PubMed Central

    Finn, Sarah; Rogers, Lisa; Händler, Kristian; McClure, Peter; Amézquita, Alejandro; Hinton, Jay C. D.

    2015-01-01

    Common salt (NaCl) is frequently used by the food industry to add flavor and to act as a humectant in order to reduce the water content of a food product. The improved health awareness of consumers is leading to a demand for food products with reduced salt content; thus, manufacturers require alternative water activity-reducing agents which elicit the same general effects as NaCl. Two examples include KCl and glycerol. These agents lower the water activity of a food matrix and also contribute to limit the growth of the microbiota, including foodborne pathogens. Little is currently known about how foodborne pathogens respond to these water activity-lowering agents. Here we examined the response of Salmonella enterica serovar Typhimurium 4/74 to NaCl, KCl, and glycerol at three time points, using a constant water activity level, compared with the response of a control inoculum. All conditions induced the upregulation of gluconate metabolic genes after 6 h of exposure. Bacteria exposed to NaCl and KCl demonstrated the upregulation of the osmoprotective transporter mechanisms encoded by the proP, proU, and osmU (STM1491 to STM1494) genes. Glycerol exposure elicited the downregulation of these osmoadaptive mechanisms but stimulated an increase in lipopolysaccharide and membrane protein-associated genes after 1 h. The most extensive changes in gene expression occurred following exposure to KCl. Because many of these genes were of unknown function, further characterization may identify KCl-specific adaptive processes that are not stimulated by NaCl. This study shows that the response of S. Typhimurium to different humectants does not simply reflect reduced water activity and likely involves systems that are linked to specific humectants. PMID:26209672

  1. An rfaH Mutant of Salmonella enterica Serovar Typhimurium is Attenuated in Swine and Reduces Intestinal Colonization, Fecal Shedding, and Disease Severity Due to Virulent Salmonella Typhimurium

    PubMed Central

    Bearson, Bradley L.; Bearson, Shawn M. D.; Kich, Jalusa D.; Lee, In Soo

    2014-01-01

    Swine are often asymptomatic carriers of Salmonella spp., and interventions are needed to limit colonization of swine to enhance food safety and reduce environmental contamination. We evaluated the attenuation and potential vaccine use in pigs of a Salmonella enterica serovar Typhimurium mutant of rfaH, the gene encoding the RfaH antiterminator that prevents premature termination of long mRNA transcripts. Pigs inoculated with wild-type S. Typhimurium exhibited a significant elevation in average body temperature (fever) at 1 and 2 days post-inoculation; rfaH-inoculated pigs did not (n = 5/group). During the 7-day trial, a significant reduction of Salmonella in the feces, tonsils, and cecum were observed in the rfaH-inoculated pigs compared to wild-type inoculated pigs. To determine whether vaccination with the rfaH mutant could provide protection against wild-type S. Typhimurium challenge, two groups of pigs (n = 14/group) were intranasally inoculated with either the rfaH mutant or a PBS placebo at 6 and 8 weeks of age and challenged with the parental, wild-type S. Typhimurium at 11 weeks of age. The average body temperature was significantly elevated in the mock-vaccinated pigs at 1 and 2 days post-challenge, but not in the rfaH-vaccinated pigs. Fecal shedding at 2 and 3 days post-challenge and colonization of intestinal tract tissues at 7 days post-challenge by wild-type S. Typhimurium was significantly reduced in the rfaH-vaccinated pigs compared to mock-vaccinated pigs. Serological analysis using the IDEXX HerdChek Swine Salmonella Test Kit indicated that vaccination with the rfaH mutant did not stimulate an immune response against LPS. These results indicate that vaccination of swine with the attenuated rfaH mutant confers protection against challenge with virulent S. Typhimurium but does not interfere with herd level monitoring for Salmonella spp., thereby allowing for differentiation of infected from vaccinated animals (DIVA). PMID

  2. Influence of Temperature and Predation on Survival of Salmonella enterica Serovar Typhimurium and Expression of invA in Soil and Manure-Amended Soil▿

    PubMed Central

    García, R.; Bælum, J.; Fredslund, L.; Santorum, P.; Jacobsen, C. S.

    2010-01-01

    The effects of three temperatures (5, 15, and 25°C) on the survival of Salmonella enterica serovar Typhimurium in topsoil were investigated in small microcosms by three different techniques: plate counting, invA gene quantification, and invA mRNA quantification. Differences in survival were related to the effect of protozoan predation. Tetracycline-resistant Salmonella serovar Typhimurium was inoculated into soil and manure-amended soil at 1.5 × 108 cells g soil−1. Population densities were determined by plate counting and by molecular methods and monitored for 42 days. Simultaneous extraction of RNA and DNA, followed by quantitative PCR, was used to investigate invA gene levels and expression. Analysis by these three techniques showed that Salmonella serovar Typhimurium survived better at 5°C. Comparing DNA and CFU levels, significantly higher values were determined by DNA-based techniques. invA mRNA levels showed a fast decrease in activity, with no detectable mRNA after an incubation period of less than 4 days in any of the soil scenarios. A negative correlation was found between Salmonella serovar Typhimurium CFU levels and protozoan most probable numbers, and we propose the role of the predator-prey interaction as a factor to explain the die-off of the introduced strain by both culture- and DNA quantification-based methods. The results indicate that temperature, manure, and protozoan predation are important factors influencing the survival of Salmonella serovar Typhimurium in soil. PMID:20562283

  3. H-NS Silencing of the Salmonella Pathogenicity Island 6-Encoded Type VI Secretion System Limits Salmonella enterica Serovar Typhimurium Interbacterial Killing

    PubMed Central

    Brunet, Yannick R.; Khodr, Ahmad; Logger, Laureen; Aussel, Laurent; Mignot, Tâm; Rimsky, Sylvie

    2015-01-01

    The secretion of bacterial toxin proteins is achieved by dedicated machineries called secretion systems. The type VI secretion system (T6SS) is a widespread versatile machine used for the delivery of protein toxins to both prokaryotic and eukaryotic cells. In Salmonella enterica serovar Typhimurium, the expression of the T6SS genes is activated during macrophage or mouse infection. Here, we show that the T6SS gene cluster is silenced by the histone-like nucleoid structuring H-NS protein using a combination of reporter fusions, electrophoretic mobility shift assays, DNase footprinting, and fluorescence microscopy. We further demonstrate that derepression of the S. Typhimurium T6SS genes induces T6SS-dependent intoxication of competing bacteria. Our results suggest that relieving T6SS H-NS silencing may be used as a sense-and-kill mechanism that will help S. Typhimurium to homogenize and synchronize the microbial population to gain efficiency during infection. PMID:25916986

  4. Molecular characterisation and mechanisms of resistance of multidrug-resistant human Salmonella enterica serovar Typhimurium isolated in Amiens (France).

    PubMed

    Biendo, Maurice; Laurans, Geneviève; Thomas, Danièle; Canarelli, Brigitte; Hamdad-Daoudi, Farida; Rousseau, Florence; Castelain, Sandrine; Eb, François

    2005-09-01

    Antimicrobial resistance patterns of Salmonella enterica serovar Typhimurium isolates obtained during the study period were examined. The molecular epidemiology and the mechanisms of resistance to ampicillin, chloramphenicol and tetracycline were investigated. Resistance to ampicillin increased from 59% between 1996 and 1999 to 62.5% in 2000 and to 66.6% in 2001. Of 51 S. Typhimurium isolates studied, 100% were resistant to ampicillin (minimum inhibitory concentration (MIC)>256 mg/L) and sulphonamide (MIC range, 128 to >256 mg/L). Ninety-eight percent of isolates were resistant to streptomycin (MIC range, 48-256 mg/L), 92.2% to tetracycline (MIC range, 32 to >256 mg/L), 88.2% to chloramphenicol (MIC>256 mg/L), 21.5% to sulphamethoxazole/trimethoprim (MIC>32 mg/L), 5.8% to amoxicillin/clavulanic acid (MIC, 32 mg/L) and 1.9% to cefalothin (MIC, 64mg/L). Six resistance phenotypes were found (a-f), with phenotypes a (47%) and b (27.5%) being predominant. Twenty-five (49%) of 51 isolates produced a single beta-lactamase, among which 48% produced PSE-1, 44% produced TEM-1 and 8% produced OXA-1. Among 26 of the 51 isolates, 10 produced PSE-1+OXA-1, 7 produced TEM-1+PSE-1+OXA-1, 6 produced TEM-1+PSE-1, and 3 produced TEM-1+OXA-1. Forty-eight (94.1%) of the 51 isolates had the plasmid-mediated resistance gene flo(ST) to chloramphenicol and tetracycline. Combining enterobacterial repetitive intergenic consensus polymerase chain reaction (ERIC-PCR) and pulsed-field gel electrophoresis (PFGE), 16 distinct patterns were identified, among which patterns IA (35.3%) and IF (27.4%) were considered as epidemic patterns. The dendrogram obtained from S. Typhimurium pulsotypes allowed five clones (S1-S5) to be identified, with two prevalent clones comprising 47.8% (S2) and 27.3% (S4) of the isolates.

  5. Role of SPI-1 secreted effectors in acute bovine response to Salmonella enterica Serovar Typhimurium: a systems biology analysis approach.

    PubMed

    Lawhon, Sara D; Khare, Sangeeta; Rossetti, Carlos A; Everts, Robin E; Galindo, Cristi L; Luciano, Sarah A; Figueiredo, Josely F; Nunes, Jairo E S; Gull, Tamara; Davidson, George S; Drake, Kenneth L; Garner, Harold R; Lewin, Harris A; Bäumler, Andreas J; Adams, Leslie Garry

    2011-01-01

    Salmonella enterica Serovar Typhimurium (S. Typhimurium) causes enterocolitis with diarrhea and polymorphonuclear cell (PMN) influx into the intestinal mucosa in humans and calves. The Salmonella Type III Secretion System (T3SS) encoded at Pathogenicity Island I translocates Salmonella effector proteins SipA, SopA, SopB, SopD, and SopE2 into epithelial cells and is required for induction of diarrhea. These effector proteins act together to induce intestinal fluid secretion and transcription of C-X-C chemokines, recruiting PMNs to the infection site. While individual molecular interactions of the effectors with cultured host cells have been characterized, their combined role in intestinal fluid secretion and inflammation is less understood. We hypothesized that comparison of the bovine intestinal mucosal response to wild type Salmonella and a SipA, SopABDE2 effector mutant relative to uninfected bovine ileum would reveal heretofore unidentified diarrhea-associated host cellular pathways. To determine the coordinated effects of these virulence factors, a bovine ligated ileal loop model was used to measure responses to wild type S. Typhimurium (WT) and a ΔsipA, sopABDE2 mutant (MUT) across 12 hours of infection using a bovine microarray. Data were analyzed using standard microarray analysis and a dynamic bayesian network modeling approach (DBN). Both analytical methods confirmed increased expression of immune response genes to Salmonella infection and novel gene expression. Gene expression changes mapped to 219 molecular interaction pathways and 1620 gene ontology groups. Bayesian network modeling identified effects of infection on several interrelated signaling pathways including MAPK, Phosphatidylinositol, mTOR, Calcium, Toll-like Receptor, CCR3, Wnt, TGF-β, and Regulation of Actin Cytoskeleton and Apoptosis that were used to model of host-pathogen interactions. Comparison of WT and MUT demonstrated significantly different patterns of host response at early time

  6. Role of SPI-1 Secreted Effectors in Acute Bovine Response to Salmonella enterica Serovar Typhimurium: A Systems Biology Analysis Approach

    PubMed Central

    Lawhon, Sara D.; Khare, Sangeeta; Rossetti, Carlos A.; Everts, Robin E.; Galindo, Cristi L.; Luciano, Sarah A.; Figueiredo, Josely F.; Nunes, Jairo E. S.; Gull, Tamara; Davidson, George S.; Drake, Kenneth L.; Garner, Harold R.; Lewin, Harris A.; Bäumler, Andreas J.; Adams, Leslie Garry

    2011-01-01

    Salmonella enterica Serovar Typhimurium (S. Typhimurium) causes enterocolitis with diarrhea and polymorphonuclear cell (PMN) influx into the intestinal mucosa in humans and calves. The Salmonella Type III Secretion System (T3SS) encoded at Pathogenicity Island I translocates Salmonella effector proteins SipA, SopA, SopB, SopD, and SopE2 into epithelial cells and is required for induction of diarrhea. These effector proteins act together to induce intestinal fluid secretion and transcription of C-X-C chemokines, recruiting PMNs to the infection site. While individual molecular interactions of the effectors with cultured host cells have been characterized, their combined role in intestinal fluid secretion and inflammation is less understood. We hypothesized that comparison of the bovine intestinal mucosal response to wild type Salmonella and a SipA, SopABDE2 effector mutant relative to uninfected bovine ileum would reveal heretofore unidentified diarrhea-associated host cellular pathways. To determine the coordinated effects of these virulence factors, a bovine ligated ileal loop model was used to measure responses to wild type S. Typhimurium (WT) and a ΔsipA, sopABDE2 mutant (MUT) across 12 hours of infection using a bovine microarray. Data were analyzed using standard microarray analysis and a dynamic Bayesian network modeling approach (DBN). Both analytical methods confirmed increased expression of immune response genes to Salmonella infection and novel gene expression. Gene expression changes mapped to 219 molecular interaction pathways and 1620 gene ontology groups. Bayesian network modeling identified effects of infection on several interrelated signaling pathways including MAPK, Phosphatidylinositol, mTOR, Calcium, Toll-like Receptor, CCR3, Wnt, TGF-β, and Regulation of Actin Cytoskeleton and Apoptosis that were used to model of host-pathogen interactions. Comparison of WT and MUT demonstrated significantly different patterns of host response at early time

  7. Antimicrobial action of carvacrol at different stages of dual-species biofilm development by Staphylococcus aureus and Salmonella enterica serovar Typhimurium.

    PubMed

    Knowles, J R; Roller, S; Murray, D B; Naidu, A S

    2005-02-01

    The effects of carvacrol, a natural biocide, on dual-species biofilms formed by Staphylococcus aureus and Salmonella enterica serovar Typhimurium were investigated with a constant-depth film fermentor. Biofilm development reached a quasi-steady state in 12 days at 25 degrees C with S. aureus predominance ( approximately 99%). Cryosectional analysis detected viable S. aureus and S. enterica serovar Typhimurium at depths of 320 and 180 mum from the film surface, respectively. Carvacrol pulses (1.0 mmol/h) inhibited S. aureus by 2.5 log CFU/biofilm during the early stages of film formation, ultimately causing a significant reduction (P < 0.001) of the staphylococcal population at quasi-steady state. Initial carvacrol pulsing elicited a 3 log CFU/biofilm reduction in viable S. enterica serovar Typhimurium, and additional periodic carvacrol pulses instigated significant inhibition of salmonellae (1 to 2 log CFU/biofilm) during biofilm development. Carvacrol pulsing reduced protein levels fivefold (P < 0.001) during initial biofilm development. Comparative studies with a peroxide-based commercial sanitizer (Spor-Klenz RTU) revealed that this commercial sanitizer was more biocidal than carvacrol during early biofilm development. When the biofilm reached quasi-steady state, however, periodic pulses with 1 mmol of carvacrol per h (P = 0.021) elicited a significantly higher inhibition than Spor-Klenz RTU (P = 0.772). Dual-species microcolonies formed under the influence of continuously fed low carvacrol concentrations (1.0 mmol/h) but failed to develop into a mature quasi-steady-state biofilm and did not reach any stage of film formation in the presence of high concentrations (5.0 mmol/h). These data show that carvacrol is an effective natural intervention to control dual-species biofilm formation.

  8. Antimicrobial Action of Carvacrol at Different Stages of Dual-Species Biofilm Development by Staphylococcus aureus and Salmonella enterica Serovar Typhimurium

    PubMed Central

    Knowles, J. R.; Roller, S.; Murray, D. B.; Naidu, A. S.

    2005-01-01

    The effects of carvacrol, a natural biocide, on dual-species biofilms formed by Staphylococcus aureus and Salmonella enterica serovar Typhimurium were investigated with a constant-depth film fermentor. Biofilm development reached a quasi-steady state in 12 days at 25°C with S. aureus predominance (≈99%). Cryosectional analysis detected viable S. aureus and S. enterica serovar Typhimurium at depths of 320 and 180 μm from the film surface, respectively. Carvacrol pulses (1.0 mmol/h) inhibited S. aureus by 2.5 log CFU/biofilm during the early stages of film formation, ultimately causing a significant reduction (P < 0.001) of the staphylococcal population at quasi-steady state. Initial carvacrol pulsing elicited a 3 log CFU/biofilm reduction in viable S. enterica serovar Typhimurium, and additional periodic carvacrol pulses instigated significant inhibition of salmonellae (1 to 2 log CFU/biofilm) during biofilm development. Carvacrol pulsing reduced protein levels fivefold (P < 0.001) during initial biofilm development. Comparative studies with a peroxide-based commercial sanitizer (Spor-Klenz RTU) revealed that this commercial sanitizer was more biocidal than carvacrol during early biofilm development. When the biofilm reached quasi-steady state, however, periodic pulses with 1 mmol of carvacrol per h (P = 0.021) elicited a significantly higher inhibition than Spor-Klenz RTU (P = 0.772). Dual-species microcolonies formed under the influence of continuously fed low carvacrol concentrations (1.0 mmol/h) but failed to develop into a mature quasi-steady-state biofilm and did not reach any stage of film formation in the presence of high concentrations (5.0 mmol/h). These data show that carvacrol is an effective natural intervention to control dual-species biofilm formation. PMID:15691933

  9. Identification of multidrug-resistant Salmonella enterica serovar typhimurium isolates that have an antibiotic-induced invasion phenotype

    USDA-ARS?s Scientific Manuscript database

    Multidrug-resistant (MDR) Salmonella is an important food safety issue in humans and animals. The National Antimicrobial Resistance Monitoring System (NARMS) has reported that 27.3% of Salmonella enterica serotype Typhimurium isolates in humans were resistant to three or more classes of antibiotics...

  10. Mutational analysis of the residue at position 48 in the Salmonella enterica Serovar Typhimurium PhoQ sensor kinase.

    PubMed

    Sanowar, Sarah; Martel, Alexandre; Moual, Hervé Le

    2003-03-01

    The PhoP/PhoQ two-component regulatory system of Salmonella enterica serovar Typhimurium plays an essential role in controlling virulence by mediating the adaptation to Mg(2+) depletion. The pho-24 allele of phoQ harbors a single amino acid substitution (T48I) in the periplasmic domain of the PhoQ histidine kinase sensor. This mutation has been shown to increase net phosphorylation of the PhoP response regulator. We analyzed the effect on signaling by PhoP/PhoQ of various amino acid substitutions at this position (PhoQ-T48X [X = A, S, V, I, or L]). Mutations T48V, T48I, and T48L were found to affect signaling by PhoP/PhoQ both in vivo and in vitro. Mutations PhoQ-T48V and PhoQ-T48I increased both the expression of the mgtA::lacZ transcriptional fusion and the net phosphorylation of PhoP, conferring to cells a PhoP constitutively active phenotype. In contrast, mutation PhoQ-T48L barely responded to changes in the concentration of external Mg(2+), in vivo and in vitro, conferring to cells a PhoP constitutively inactive phenotype. By analyzing in vitro the individual catalytic activities of the PhoQ-T48X sensors, we found that the PhoP constitutively active phenotype observed for the PhoQ-T48V and PhoQ-T48I proteins is solely due to decreased phosphatase activity. In contrast, the PhoP constitutively inactive phenotype observed for the PhoQ-T48L mutant resulted from both decreased autokinase activity and increased phosphatase activity. Our data are consistent with a model in which the residue at position 48 of PhoQ contributes to a conformational switch between kinase- and phosphatase-dominant states.

  11. Low-Shear modeled microgravity alters the Salmonella enterica serovar typhimurium stress response in an RpoS-independent manner

    NASA Technical Reports Server (NTRS)

    Wilson, James W.; Ott, C. Mark; Ramamurthy, Rajee; Porwollik, Steffen; McClelland, Michael; Pierson, Duane L.; Nickerson, Cheryl A.

    2002-01-01

    We have previously demonstrated that low-shear modeled microgravity (low-shear MMG) serves to enhance the virulence of a bacterial pathogen, Salmonella enterica serovar Typhimurium. The Salmonella response to low-shear MMG involves a signaling pathway that we have termed the low-shear MMG stimulon, though the identities of the low-shear MMG stimulon genes and regulatory factors are not known. RpoS is the primary sigma factor required for the expression of genes that are induced upon exposure to different environmental-stress signals and is essential for virulence in mice. Since low-shear MMG induces a Salmonella acid stress response and enhances Salmonella virulence, we reasoned that RpoS would be a likely regulator of the Salmonella low-shear MMG response. Our results demonstrate that low-shear MMG provides cross-resistance to several environmental stresses in both wild-type and isogenic rpoS mutant strains. Growth under low-shear MMG decreased the generation time of both strains in minimal medium and increased the ability of both strains to survive in J774 macrophages. Using DNA microarray analysis, we found no evidence of induction of the RpoS regulon by low-shear MMG but did find that other genes were altered in expression under these conditions in both the wild-type and rpoS mutant strains. Our results indicate that, under the conditions of these studies, RpoS is not required for transmission of the signal that induces the low-shear MMG stimulon. Moreover, our studies also indicate that low-shear MMG can be added to a short list of growth conditions that can serve to preadapt an rpoS mutant for resistance to multiple environmental stresses.

  12. Regulation of the AcrAB multidrug efflux pump in Salmonella enterica serovar Typhimurium in response to indole and paraquat.

    PubMed

    Nikaido, Eiji; Shirosaka, Ikue; Yamaguchi, Akihito; Nishino, Kunihiko

    2011-03-01

    Salmonella enterica serovar Typhimurium has at least nine multidrug efflux pumps. Among these, AcrAB is constitutively expressed and is the most efficient, playing a role in both drug resistance and virulence. The acrAB locus is induced by indole, Escherichia coli-conditioned medium, and bile salts. This induction is dependent on RamA through the binding sequence in the upstream region of acrA that binds RamA. In the present study, we made a detailed investigation of the ramA and acrAB induction mechanisms in Salmonella in response to indole, a biological oxidant for bacteria. We found that acrAB and ramA induction in response to indole is dependent on RamR. However, the cysteine residues of RamR do not play a role in the induction of ramA in response to indole, and the oxidative effect of indole is therefore not related to ramA induction via RamR. Furthermore, we showed that paraquat, a superoxide generator, induces acrAB but not ramA. We further discovered that the mechanism of acrAB induction in response to paraquat is dependent on SoxS. The data indicate that there are at least two independent induction pathways for acrAB in response to extracellular signals such as indole and paraquat. We propose that Salmonella utilizes these regulators for acrAB induction in response to extracellular signals in order to adapt itself to environmental conditions.

  13. The effect of recombinant human lactoferrin from the milk of transgenic cows on Salmonella enterica serovar typhimurium infection in mice.

    PubMed

    Wu, Jinghuan; Hu, Yujie; Du, Chunming; Piao, Jianhua; Yang, Lichen; Yang, Xiaoguang

    2016-01-01

    Lactoferrin (LF) is a multifunctional protein with antibacterial and immunomodulatory activities. Given this beneficial effect, transgenic approaches have been used to produce lactoferrin. The aim of the current study was to investigate the in vivo effect of recombinant human lactoferrin (rhLF) from the milk of transgenic cows on Salmonella enterica serovar typhimurium (ST) infection in mice. Two hours before the infection with 0.3 ml at 2 × 10(5) CFU ml(-1) of ST, each animal in the ST + rhLF group received 0.3 ml of rhLF with 20 mg ml(-1) concentration while the ST group received PBS as placebos with the same volume through oral gavage. The mice were infected with ST once only on the first day. After the infection, the mice received 0.3 ml of rhLF with 20 mg ml(-1) (6 mg d(-1)) concentration or PBS, respectively, for 7 days. Mortality and weight were monitored daily. Bacterial enumeration in the blood, liver, and spleen and histopathological analysis of the liver, spleen, kidney and intestine were conducted. The results showed that rhLF decreased the bacterial load in the liver and spleen of mice, reduced the degree of mice hepatomegaly and splenomegaly, and attenuated infectious inflammation with less histopathological abnormalities in the liver, spleen and kidney of mice in the ST infection. This study showed that rhLF with 6 mg per day had antibacterial activity of alleviating the infection caused by ST bacteria, which indicated that rhLF could be used as a supplement in special products.

  14. Low-Shear modeled microgravity alters the Salmonella enterica serovar typhimurium stress response in an RpoS-independent manner.

    PubMed

    Wilson, James W; Ott, C Mark; Ramamurthy, Rajee; Porwollik, Steffen; McClelland, Michael; Pierson, Duane L; Nickerson, Cheryl A

    2002-11-01

    We have previously demonstrated that low-shear modeled microgravity (low-shear MMG) serves to enhance the virulence of a bacterial pathogen, Salmonella enterica serovar Typhimurium. The Salmonella response to low-shear MMG involves a signaling pathway that we have termed the low-shear MMG stimulon, though the identities of the low-shear MMG stimulon genes and regulatory factors are not known. RpoS is the primary sigma factor required for the expression of genes that are induced upon exposure to different environmental-stress signals and is essential for virulence in mice. Since low-shear MMG induces a Salmonella acid stress response and enhances Salmonella virulence, we reasoned that RpoS would be a likely regulator of the Salmonella low-shear MMG response. Our results demonstrate that low-shear MMG provides cross-resistance to several environmental stresses in both wild-type and isogenic rpoS mutant strains. Growth under low-shear MMG decreased the generation time of both strains in minimal medium and increased the ability of both strains to survive in J774 macrophages. Using DNA microarray analysis, we found no evidence of induction of the RpoS regulon by low-shear MMG but did find that other genes were altered in expression under these conditions in both the wild-type and rpoS mutant strains. Our results indicate that, under the conditions of these studies, RpoS is not required for transmission of the signal that induces the low-shear MMG stimulon. Moreover, our studies also indicate that low-shear MMG can be added to a short list of growth conditions that can serve to preadapt an rpoS mutant for resistance to multiple environmental stresses.

  15. Interaction of Saccharomyces boulardii with Salmonella enterica Serovar Typhimurium Protects Mice and Modifies T84 Cell Response to the Infection

    PubMed Central

    Martins, Flaviano S.; Dalmasso, Guillaume; Arantes, Rosa M. E.; Doye, Anne; Lemichez, Emmanuel; Lagadec, Patricia; Imbert, Veronique; Peyron, Jean-François; Rampal, Patrick; Nicoli, Jacques R.; Czerucka, Dorota

    2010-01-01

    Background Salmonella pathogenesis engages host cells in two-way biochemical interactions: phagocytosis of bacteria by recruitment of cellular small GTP-binding proteins induced by the bacteria, and by triggering a pro-inflammatory response through activation of MAPKs and nuclear translocation of NF-κB. Worldwide interest in the use of functional foods containing probiotic bacteria for health promotion and disease prevention has increased significantly. Saccharomyces boulardii is a non-pathogenic yeast used as a probiotic in infectious diarrhea. Methodology/Principal Findings In this study, we reported that S. boulardii (Sb) protected mice from Salmonella enterica serovar Typhimurium (ST)-induced death and prevented bacterial translocation to the liver. At a molecular level, using T84 human colorectal cancer cells, we demonstrate that incubation with Sb before infection totally abolished Salmonella invasion. This correlates with a decrease of activation of Rac1. Sb preserved T84 barrier function and decreased ST-induced IL-8 synthesis. This anti-inflammatory effect was correlated with an inhibitory effect of Sb on ST-induced activation of the MAPKs ERK1/2, p38 and JNK as well as on activation of NF-κB. Electron and confocal microscopy experiments showed an adhesion of bacteria to yeast cells, which could represent one of the mechanisms by which Sb exerts its protective effects. Conclusions Sb shows modulating effects on permeability, inflammation, and signal transduction pathway in T84 cells infected by ST and an in vivo protective effect against ST infection. The present results also demonstrate that Sb modifies invasive properties of Salmonella. PMID:20111723

  16. Low-Shear modeled microgravity alters the Salmonella enterica serovar typhimurium stress response in an RpoS-independent manner

    NASA Technical Reports Server (NTRS)

    Wilson, James W.; Ott, C. Mark; Ramamurthy, Rajee; Porwollik, Steffen; McClelland, Michael; Pierson, Duane L.; Nickerson, Cheryl A.

    2002-01-01

    We have previously demonstrated that low-shear modeled microgravity (low-shear MMG) serves to enhance the virulence of a bacterial pathogen, Salmonella enterica serovar Typhimurium. The Salmonella response to low-shear MMG involves a signaling pathway that we have termed the low-shear MMG stimulon, though the identities of the low-shear MMG stimulon genes and regulatory factors are not known. RpoS is the primary sigma factor required for the expression of genes that are induced upon exposure to different environmental-stress signals and is essential for virulence in mice. Since low-shear MMG induces a Salmonella acid stress response and enhances Salmonella virulence, we reasoned that RpoS would be a likely regulator of the Salmonella low-shear MMG response. Our results demonstrate that low-shear MMG provides cross-resistance to several environmental stresses in both wild-type and isogenic rpoS mutant strains. Growth under low-shear MMG decreased the generation time of both strains in minimal medium and increased the ability of both strains to survive in J774 macrophages. Using DNA microarray analysis, we found no evidence of induction of the RpoS regulon by low-shear MMG but did find that other genes were altered in expression under these conditions in both the wild-type and rpoS mutant strains. Our results indicate that, under the conditions of these studies, RpoS is not required for transmission of the signal that induces the low-shear MMG stimulon. Moreover, our studies also indicate that low-shear MMG can be added to a short list of growth conditions that can serve to preadapt an rpoS mutant for resistance to multiple environmental stresses.

  17. Role of Autocleavage in the Function of a Type III Secretion Specificity Switch Protein in Salmonella enterica Serovar Typhimurium

    PubMed Central

    Monjarás Feria, Julia V.; Lefebre, Matthew D.; Stierhof, York-Dieter

    2015-01-01

    ABSTRACT Type III secretion systems (T3SSs) are multiprotein machines employed by many Gram-negative bacteria to inject bacterial effector proteins into eukaryotic host cells to promote bacterial survival and colonization. The core unit of T3SSs is the needle complex, a supramolecular structure that mediates the passage of the secreted proteins through the bacterial envelope. A distinct feature of the T3SS is that protein export occurs in a strictly hierarchical manner in which proteins destined to form the needle complex filament and associated structures are secreted first, followed by the secretion of effectors and the proteins that will facilitate their translocation through the target host cell membrane. The secretion hierarchy is established by complex mechanisms that involve several T3SS-associated components, including the “switch protein,” a highly conserved, inner membrane protease that undergoes autocatalytic cleavage. It has been proposed that the autocleavage of the switch protein is the trigger for substrate switching. We show here that autocleavage of the Salmonella enterica serovar Typhimurium switch protein SpaS is an unregulated process that occurs after its folding and before its incorporation into the needle complex. Needle complexes assembled with a precleaved form of SpaS function in a manner indistinguishable from that of the wild-type form. Furthermore, an engineered mutant of SpaS that is processed by an external protease also displays wild-type function. These results demonstrate that the cleavage event per se does not provide a signal for substrate switching but support the hypothesis that cleavage allows the proper conformation of SpaS to render it competent for its switching function. PMID:26463164

  18. Genome-guided design of a defined mouse microbiota that confers colonization resistance against Salmonella enterica serovar Typhimurium.

    PubMed

    Brugiroux, Sandrine; Beutler, Markus; Pfann, Carina; Garzetti, Debora; Ruscheweyh, Hans-Joachim; Ring, Diana; Diehl, Manuel; Herp, Simone; Lötscher, Yvonne; Hussain, Saib; Bunk, Boyke; Pukall, Rüdiger; Huson, Daniel H; Münch, Philipp C; McHardy, Alice C; McCoy, Kathy D; Macpherson, Andrew J; Loy, Alexander; Clavel, Thomas; Berry, David; Stecher, Bärbel

    2016-11-21

    Protection against enteric infections, also termed colonization resistance, results from mutualistic interactions of the host and its indigenous microbes. The gut microbiota of humans and mice is highly diverse and it is therefore challenging to assign specific properties to its individual members. Here, we have used a collection of murine bacterial strains and a modular design approach to create a minimal bacterial community that, once established in germ-free mice, provided colonization resistance against the human enteric pathogen Salmonella enterica serovar Typhimurium (S. Tm). Initially, a community of 12 strains, termed Oligo-Mouse-Microbiota (Oligo-MM(12)), representing members of the major bacterial phyla in the murine gut, was selected. This community was stable over consecutive mouse generations and provided colonization resistance against S. Tm infection, albeit not to the degree of a conventional complex microbiota. Comparative (meta)genome analyses identified functions represented in a conventional microbiome but absent from the Oligo-MM(12). By genome-informed design, we created an improved version of the Oligo-MM community harbouring three facultative anaerobic bacteria from the mouse intestinal bacterial collection (miBC) that provided conventional-like colonization resistance. In conclusion, we have established a highly versatile experimental system that showed efficacy in an enteric infection model. Thus, in combination with exhaustive bacterial strain collections and systems-based approaches, genome-guided design can be used to generate insights into microbe-microbe and microbe-host interactions for the investigation of ecological and disease-relevant mechanisms in the intestine.

  19. Molecular Epidemiology of Salmonella enterica Serovar Typhimurium Isolates Determined by Pulsed-Field Gel Electrophoresis: Comparison of Isolates from Avian Wildlife, Domestic Animals, and the Environment in Norway

    PubMed Central

    Refsum, Thorbjørn; Heir, Even; Kapperud, Georg; Vardund, Traute; Holstad, Gudmund

    2002-01-01

    The molecular epidemiology of 142 isolates of Salmonella enterica serovar Typhimurium from avian wildlife, domestic animals, and the environment in Norway was investigated using pulsed-field gel electrophoresis (PFGE) and computerized numerical analysis of the data. The bacterial isolates comprised 79 isolates from wild-living birds, including 46 small passerines and 26 gulls, and 63 isolates of nonavian origin, including 50 domestic animals and 13 environmental samples. Thirteen main clusters were discernible at the 90% similarity level. Most of the isolates (83%) were grouped into three main clusters. These were further divided into 20 subclusters at the 95% similarity level. Isolates from passerines, gulls, and pigeons dominated within five subclusters, whereas isolates from domestic animals and the environment belonged to many different subclusters with no predominance. The results support earlier results that passerines constitute an important source of infection to humans in Norway, whereas it is suggested that gulls and pigeons, based on PFGE analysis, represent only a minor source of human serovar Typhimurium infections. Passerines, gulls, and pigeons may also constitute a source of infection of domestic animals and feed plants or vice versa. Three isolates from cattle and a grain source, of which two were multiresistant, were confirmed as serovar Typhimurium phage type DT 104. These represent the first reported phage type DT 104 isolates from other sources than humans in Norway. PMID:12406755

  20. Gene regulation of Salmonella enterica serovar Typhimurium as influenced by LuxS/AI-2

    USDA-ARS?s Scientific Manuscript database

    Quorum sensing is mediated by small signaling molecules, autoinducer molecules. The luxS gene which is conserved in several Gram-positive and Gram-negative bacteria is involved in the synthesis of the autoinducer molecule-2 (AI-2). Genes controlled by luxS in S. Typhimurium were identified using mic...

  1. Isolation of QseC-regulated genes in Salmonella enterica serovar Typhimurium by transposon mutgagenesis

    USDA-ARS?s Scientific Manuscript database

    Non-typhoidal Salmonella, a leading cause of U.S. foodborne disease and food-related deaths, often asymptomatically colonizes food-producing animals. In fact, >50% of U.S. swine production facilities test positive for Salmonella. The multidrug-resistant (MDR) Salmonella Typhimurium DT104 NCTC13348 c...

  2. Coptidis rhizome and Si Jun Zi Tang can prevent Salmonella enterica serovar Typhimurium infection in mice.

    PubMed

    Chang, Chiung-Hung; Yu, Bi; Su, Chiu-Hsian; Chen, Daniel S; Hou, Yu-Chi; Chen, Yueh-Sheng; Hsu, Yuan-Man

    2014-01-01

    Salmonella, a common zoonotic pathogen, causes gastroenteritis in both humans and animals. Traditional Chinese Medicine (TCM) has been used to improve gastrointestinal dysfunction and to modify the immune response to inflammation for centuries. This study used six herbal plants and four TCM formulae to rate their efficacy in preventing S. Typhimurium infection via mouse model. Minimum bactericidal concentration (MBC) of Coptidis rhizome (CR) against the reference strain tallied 12.5 mg/ml and against clinical isolate ST21 was 25 mg/ml. MBCs of other herbal extracts and formulae on Salmonella Typhimurium strains were above 50 mg/ml. In the mice model, CR and Si Jun Zi Tang (SJZT) could significantly decrease the bacterial load in organs and blood after being challenged, along with body weight loss due to the infection. CR and SJZT alleviated infection-induced interferon-gamma levels in the serum and tissues, and tumor necrosis factor-alpha (TNF-α) levels in intestinal tissues. CR and SJZT serum metabolites could suppress S. Typhimurium invasion and TNF-α expression in RAW264.7 cells. The therapeutic activity of CR and SJZT may involve berberine, ginsenoside Rb1, and glycyrrhizin, interfering with Salmonella when invading macrophages. CR and SJZT has shown potential in preventing S. Typhimurium infection through the regulation of the immune response.

  3. Salmonella enterica serovar Typhimurium and Listeria monocytogenes acid tolerance response induced by organic acids at 20 degrees C: optimization and modeling.

    PubMed

    Greenacre, E J; Brocklehurst, T F; Waspe, C R; Wilson, D R; Wilson, P D G

    2003-07-01

    An acid tolerance response (ATR) has been demonstrated in Listeria monocytogenes and Salmonella enterica serovar Typhimurium in response to low pH poised (i.e., adapted) with acetic or lactic acids at 20 degrees C and modeled by using dynamic differential equations. The ATR was not immediate or prolonged, and optimization occurred after exposure of L. monocytogenes for 3 h at pH 5.5 poised with acetic acid and for 2 h at pH 5.5 poised with lactic acid and after exposure of S. enterica serovar Typhimurium for 2 h at pH 5.5 poised with acetic acid and for 3 h at pH 5.5 poised with lactic acid. An objective mechanistic analysis of the acid inactivation data yielded estimates of the duration of the shoulder (t(s)), the log-linear decline (k(max)), and the magnitude of a critical component (C). The magnitude of k(max) gave the best agreement with estimates of conditions for optimum ATR induction made from the raw data.

  4. The Stringent Response Regulator DksA Is Required for Salmonella enterica Serovar Typhimurium Growth in Minimal Medium, Motility, Biofilm Formation, and Intestinal Colonization

    PubMed Central

    Azriel, Shalhevet; Goren, Alina; Rahav, Galia

    2015-01-01

    Salmonella enterica serovar Typhimurium is a facultative intracellular human and animal bacterial pathogen posing a major threat to public health worldwide. Salmonella pathogenicity requires complex coordination of multiple physiological and virulence pathways. DksA is a conserved Gram-negative regulator that belongs to a distinct group of transcription factors that bind directly to the RNA polymerase secondary channel, potentiating the effect of the signaling molecule ppGpp during a stringent response. Here, we established that in S. Typhimurium, dksA is induced during the logarithmic phase and DksA is essential for growth in minimal defined medium and plays an important role in motility and biofilm formation. Furthermore, we determined that DksA positively regulates the Salmonella pathogenicity island 1 and motility-chemotaxis genes and is necessary for S. Typhimurium invasion of human epithelial cells and uptake by macrophages. In contrast, DksA was found to be dispensable for S. Typhimurium host cell adhesion. Finally, using the colitis mouse model, we found that dksA is spatially induced at the midcecum during the early stage of the infection and required for gastrointestinal colonization and systemic infection in vivo. Taken together, these data indicate that the ancestral stringent response regulator DksA coordinates various physiological and virulence S. Typhimurium programs and therefore is a key virulence regulator of Salmonella. PMID:26553464

  5. A Mutation in the Putative Lysl-tRNA Synthetase Gene, PoxR of Salmonella enterica Serovar Typhimurium Results in Altered Protein Production, Elevated Susceptibility to Environmental Challenges and Decreased Swine Colonization

    USDA-ARS?s Scientific Manuscript database

    Using signature-tagged mutagenesis, a mutation in the poxR gene of Salmonella enterica serovar Typhimurium was identified with decreased survival in an ex vivo swine stomach content assay(Bearson et al. Appl Environ Microbiol. 72:2829-36). Gastrointestinal colonization and fecal shedding of the pox...

  6. Complete Genome Sequence of a Human-Invasive Salmonella enterica Serovar Typhimurium Strain of the Emerging Sequence Type 213 Harboring a Multidrug Resistance IncA/C Plasmid and a blaCMY-2-Carrying IncF Plasmid.

    PubMed

    Silva, Claudia; Calva, Edmundo; Calva, Juan J; Wiesner, Magdalena; Fernández-Mora, Marcos; Puente, José L; Vinuesa, Pablo

    2015-11-12

    Salmonella enterica subsp. enterica serovar Typhimurium strain 33676 was isolated in Mexico City, Mexico, from a patient with a systemic infection, and its complete genome sequence was determined using PacBio single-molecule real-time technology. Strain 33676 harbors an IncF plasmid carrying the extended-spectrum cephalosporin gene blaCMY-2 and a multidrug resistance IncA/C plasmid.

  7. Complete Genome Sequence of a Human-Invasive Salmonella enterica Serovar Typhimurium Strain of the Emerging Sequence Type 213 Harboring a Multidrug Resistance IncA/C Plasmid and a blaCMY-2-Carrying IncF Plasmid

    PubMed Central

    Calva, Edmundo; Calva, Juan J.; Wiesner, Magdalena; Fernández-Mora, Marcos; Puente, José L.

    2015-01-01

    Salmonella enterica subsp. enterica serovar Typhimurium strain 33676 was isolated in Mexico City, Mexico, from a patient with a systemic infection, and its complete genome sequence was determined using PacBio single-molecule real-time technology. Strain 33676 harbors an IncF plasmid carrying the extended-spectrum cephalosporin gene blaCMY-2 and a multidrug resistance IncA/C plasmid. PMID:26564044

  8. Recovery of Salmonella enterica serovars Typhimurium and Tennessee in peanut butter after electron beam exposure.

    PubMed

    Matak, Kristen E; Hvizdzak, Andrea L; Beamer, Sarah; Jaczynski, Jacek

    2010-09-01

    The effect of electron beam (e-beam) radiation on the recovery of Salmonella serotypes Tennessee (ATCC 10722) and Typhimurium (ATCC 14028) in creamy peanut butter over a 14-d storage period at 22 °C was studied. Each Salmonella type was independently inoculated into peanut butter and subjected to e-beam doses that ranged from 0 to 3.1 kGy, confirmed by film dosimetry. After 2-, 4-, 6-, 8-, and 14-d of storage, microbial analyses were conducted. Survivors were recovered on growth and selective media using standard spread-plating methods. Microbial counts (CFU/g) were log-converted and differences were determined by ANOVA and Tukey's Honestly Significant Differences test. When samples were not e-beam-treated, there were no significant changes (P > 0.05) in microbial numbers over time. In e-beamed samples, microbial numbers decreased over time; however, reductions were not always significant. Initial recovery rates (R-rates) 2 d after e-beam treatment were significantly different for the 2 strains of Salmonella and between recovery media (P < 0.05); however, these differences did not persist for the remainder of the storage period (P > 0.05) indicating that injured cells were not able to survive in the high-fat, low-water activity peanut butter environment. R-rates for both strains of Salmonella were maintained until day 14 when there were significant reductions in Salmonella Typhimurium (P < 0.05). These results indicate that Salmonella Tennessee and Salmonella Typhimurium will survive in peanut butter when exposed to nonlethal doses of e-beam irradiation. Electron beam (e-beam) irradiation is an alternative to thermal processing; this technique inactivates microorganisms and insects that might be present in a food by generating radiation by accelerated electrons that inactivate organisms directly because of interaction with cell components and indirectly by producing free radicals that disrupt integrity of the cell membrane. E-beam radiation will reduce the number

  9. Influence of temperature fluctuations on Escherichia coli O157:H7 and Salmonella enterica serovar Typhimurium in cow manure.

    PubMed

    Semenov, Alexander V; van Bruggen, Ariena H C; van Overbeek, Leo; Termorshuizen, Aad J; Semenov, Alexander M

    2007-06-01

    The effects of four average temperatures (7, 16, 23 and 33 degrees C) and daily oscillations with three amplitudes (0, +/-4, +/-7 degrees C) on the survival of the enteropathogens Escherichia coli O157:H7 and Salmonella serovar Typhimurium were investigated in small microcosms. Manure was inoculated with a green fluorescent protein transformed strain of either pathogen at 10(7) cells g(-1) dry weight. Samples were collected immediately after inoculation, and 1 and 2 weeks after inoculation for E. coli O157:H7, and immediately and after 2 and 3 weeks for Salmonella serovar Typhimurium. Population densities were determined by dilution plating and direct counting. In addition, total bacterial CFUs were determined. Growth and survival data were fitted to a modified logistic model. Analysis of the estimated parameter values showed that E. coli O157:H7 survived for shorter periods of time and was more sensitive to competition by the native microbial community than Salmonella serovar Typhimurium. Survival of both pathogens significantly declined with increasing mean temperatures and with increasing amplitude in daily temperature oscillations. The results indicated that responses of enteropathogens to fluctuating temperatures cannot be deduced from temperature relationships determined under constant temperatures.

  10. Global Screening of Salmonella enterica Serovar Typhimurium Genes for Desiccation Survival

    PubMed Central

    Mandal, Rabindra K.; Kwon, Young M.

    2017-01-01

    Salmonella spp., one of the most common foodborne bacterial pathogens, has the ability to survive under desiccation conditions in foods and food processing facilities for years. This raises the concerns of Salmonella infection in humans associated with low water activity foods. Salmonella responds to desiccation stress via complex pathways involving immediate physiological actions as well as coordinated genetic responses. However, the exact mechanisms of Salmonella to resist desiccation stress remain to be fully elucidated. In this study, we screened a genome-saturating transposon (Tn5) library of Salmonella Typhimurium (S. Typhimurium) 14028s under the in vitro desiccation stress using transposon sequencing (Tn-seq). We identified 61 genes and 6 intergenic regions required to overcome desiccation stress. Salmonella desiccation resistance genes were mostly related to energy production and conversion; cell wall/membrane/envelope biogenesis; inorganic ion transport and metabolism; regulation of biological process; DNA metabolic process; ABC transporters; and two component system. More than 20% of the Salmonella desiccation resistance genes encode either putative or hypothetical proteins. Phenotypic evaluation of 12 single gene knockout mutants showed 3 mutants (atpH, atpG, and corA) had significantly (p < 0.02) reduced survival as compared to the wild type during desiccation survival. Thus, our study provided new insights into the molecular mechanisms utilized by Salmonella for survival against desiccation stress. The findings might be further exploited to develop effective control strategies against Salmonella contamination in low water activity foods and food processing facilities.

  11. Transfer of Salmonella enterica Serovar Typhimurium from Beef to Tomato through Kitchen Equipment and the Efficacy of Intermediate Decontamination Procedures.

    PubMed

    Gkana, E; Lianou, A; Nychas, G-J E

    2016-07-01

    It is well established that a high percentage of foodborne illness is caused by failure of consumers to prepare food in a hygienic manner. Indeed, a common practice in households is to use the same kitchen equipment for both raw meat and fresh produce. Such a practice may lead to cross-contamination of fruits and vegetables, which are mainly consumed without further processing, with pathogenic microorganisms originating from raw meat. The present study was performed to examine the transfer of the pathogenic bacterium Salmonella enterica serovar Typhimurium from inoculated beef fillets to tomatoes via contact with high-density polyethylene (PE), stainless steel (SS), and wooden (WD) surfaces and through cutting with SS knives. Furthermore, the following decontamination procedures were applied: (i) rinsing with tap water, (ii) scrubbing with tap water and liquid dish detergent, and (iii) using a commercial antibacterial spray. When surfaces and knives that came into contact with contaminated beef fillets were not cleaned prior to handling tomatoes, the lowest level of pathogen transfer to tomatoes was observed through PE surfaces. All of the decontamination procedures applied were more effective on knives than on surfaces, while among the surface materials tested, WD surfaces were the most difficult to decontaminate, followed by PE and SS surfaces. Mechanical cleaning with tap water and detergent was more efficient in decontaminating WD surfaces than using commercial disinfectant spray, followed by rinsing only with water. Specifically, reductions of 2.07 and 1.09 log CFU/cm(2) were achieved by washing the WD surfaces with water and detergent and spraying the surfaces with an antibacterial product, respectively. Although the pathogen's populations on SS and PE surfaces, as well as on tomatoes, after both aforementioned treatments were under the detection limit, the surfaces were all positive after enrichment, and thus, the potential risk of cross-contamination cannot

  12. Role of autocleavage in the function of a type III secretion specificity switch protein in Salmonella enterica serovar Typhimurium.

    PubMed

    Monjarás Feria, Julia V; Lefebre, Matthew D; Stierhof, York-Dieter; Galán, Jorge E; Wagner, Samuel

    2015-10-13

    Type III secretion systems (T3SSs) are multiprotein machines employed by many Gram-negative bacteria to inject bacterial effector proteins into eukaryotic host cells to promote bacterial survival and colonization. The core unit of T3SSs is the needle complex, a supramolecular structure that mediates the passage of the secreted proteins through the bacterial envelope. A distinct feature of the T3SS is that protein export occurs in a strictly hierarchical manner in which proteins destined to form the needle complex filament and associated structures are secreted first, followed by the secretion of effectors and the proteins that will facilitate their translocation through the target host cell membrane. The secretion hierarchy is established by complex mechanisms that involve several T3SS-associated components, including the "switch protein," a highly conserved, inner membrane protease that undergoes autocatalytic cleavage. It has been proposed that the autocleavage of the switch protein is the trigger for substrate switching. We show here that autocleavage of the Salmonella enterica serovar Typhimurium switch protein SpaS is an unregulated process that occurs after its folding and before its incorporation into the needle complex. Needle complexes assembled with a precleaved form of SpaS function in a manner indistinguishable from that of the wild-type form. Furthermore, an engineered mutant of SpaS that is processed by an external protease also displays wild-type function. These results demonstrate that the cleavage event per se does not provide a signal for substrate switching but support the hypothesis that cleavage allows the proper conformation of SpaS to render it competent for its switching function. Bacterial interaction with eukaryotic hosts often involves complex molecular machines for targeted delivery of bacterial effector proteins. One such machine, the type III secretion system of some Gram-negative bacteria, serves to inject a multitude of structurally

  13. Immunogenicity of recombinant attenuated Salmonella enterica serovar Typhimurium vaccine strains carrying a gene that encodes Eimeria tenella antigen SO7.

    PubMed

    Konjufca, Vjollca; Jenkins, Mark; Wang, Shifeng; Juarez-Rodriguez, Maria Dolores; Curtiss, Roy

    2008-12-01

    Recombinant attenuated Salmonella vaccines against avian coccidiosis were developed to deliver Eimeria species antigens to the lymphoid tissues of chickens via the type 3 secretion system (T3SS) and the type 2 secretion system (T2SS) of Salmonella. For antigen delivery via the T3SS, the Eimeria tenella gene encoding sporozoite antigen SO7 was cloned downstream of the translocation domain of the Salmonella enterica serovar Typhimurium sopE gene in the parental pYA3868 and pYA3870 vectors to generate pYA4156 and pYA4157. Newly constructed T3SS vectors were introduced into host strain chi8879 (Delta phoP233 Delta sptP1033::xylE Delta asdA16), an attenuated derivative of the highly virulent UK-1 strain. The SopE-SO7 fusion protein was secreted by the T3SS of Salmonella. The vector pYA4184 was constructed for delivery of the SO7 antigen via the T2SS. The SO7 protein was toxic to Salmonella when larger amounts were synthesized; thus, the synthesis of this protein was placed under the control of the lacI repressor gene, whose expression in turn was dependent on the amount of available arabinose in the medium. The pYA4184 vector was introduced into host strain chi9242 (Delta phoP233 Delta asdA16 Delta araBAD23 Delta relA198::araC P(BAD) lacI TT [TT is the T4ipIII transcription terminator]). In addition to SO7, for immunization and challenge studies we used the EAMZ250 antigen of Eimeria acervulina, which was previously shown to confer partial protection against E. acervulina challenge when it was delivered via the T3SS. Immunization of chickens with a combination of the SO7 and EAMZ250 antigens delivered via the T3SS induced superior protection against challenge by E. acervulina. In contrast, chickens immunized with SO7 that was delivered via the T2SS of Salmonella were better protected from challenge by E. tenella.

  14. Interaction of Bacillus species and Salmonella enterica serovar Typhimurium in immune or inflammatory signaling from swine intestinal epithelial cells.

    PubMed

    Aperce, C C; Burkey, T E; KuKanich, B; Crozier-Dodson, B A; Dritz, S S; Minton, J E

    2010-05-01

    Previous research evaluated a laboratory strain of Bacillus licheniformis (BL) in a model swine epithelium and found it exerted antiinflammatory effects on Salmonella enterica serovar Typhimurium (Sal)-induced secretion of IL-8. The current investigation evaluated the antiinflammatory actions of Bacillus bacteria available commercially as feed additives for the swine industry. Three isolates were obtained from the product, 2 Bacillus subtilis (BS1 and BS3) and 1 BL (BL2). Swine jejunal epithelial IPEC-J2 cells were seeded into wells on permeable membrane supports and allowed to form confluent monolayers. Treatments included apical pretreatment with BL, BS1, BL2, or BS3 for 17 h without Sal, and the same Bacillus treatments but with 10(8) cfu of Sal added in the final hour of Bacillus incubation. Two additional treatments included negative control wells receiving no bacteria (control) and positive control wells receiving only Sal (10 total treatments). After bacterial incubation, wells were washed and fresh medium containing gentamicin was added. Cells were incubated for an additional 5 h, after which apical and basolateral media were recovered for determination of IL-8 and bacitracin. In addition, inserts with epithelial cells that had received Sal were lysed and lysates were cultured to determine treatment effects on Sal invasion. Exposure to Sal alone provoked an increase in IL-8 secretion from IPEC-J2 cells compared with control wells (P < 0.001 for both the apical and basolateral directions). Pretreatment with each Bacillus isolate followed by challenge with Sal reduced Sal-induced IL-8 secretion in both the apical and basolateral compartments compared with wells receiving only Sal (P < 0.001; except for BS3 apical, P < 0.01). The residual presence of bacitracin could be detected only in BL2 and BL2+Sal. Fewer Sal colonies could be cultured from lysates of BL2+Sal than from the Sal, BS1+Sal, and BS3+Sal treatments (P < 0.001). Results indicate that B. subtilis

  15. Formation of Thiolated Nucleosides Present in tRNA from Salmonella enterica serovar Typhimurium Occurs in Two Principally Distinct Pathways

    PubMed Central

    Leipuviene, Ramune; Qian, Qiang; Björk, Glenn R.

    2004-01-01

    tRNA from Salmonella enterica serovar Typhimurium contains five thiolated nucleosides, 2-thiocytidine (s2C), 4-thiouridine (s4U), 5-methylaminomethyl-2-thiouridine (mnm5s2U), 5-carboxymethylaminomethyl-2-thiouridine (cmnm5s2U), and N-6-(4-hydroxyisopentenyl)-2-methylthioadenosine (ms2io6A). The levels of all of them are significantly reduced in cells with a mutated iscS gene, which encodes the cysteine desulfurase IscS, a member of the ISC machinery that is responsible for [Fe-S] cluster formation in proteins. A mutant (iscU52) was isolated that carried an amino acid substitution (S107T) in the IscU protein, which functions as a major scaffold in the formation of [Fe-S] clusters. In contrast to the iscS mutant, the iscU52 mutant showed reduced levels of only two of the thiolated nucleosides, ms2io6A (10-fold) and s2C (more than 2-fold). Deletions of the iscU, hscA, or fdx genes from the isc operon lead to a similar tRNA thiolation pattern to that seen for the iscU52 mutant. Unexpectedly, deletion of the iscA gene, coding for an alternative scaffold protein for the [Fe-S] clusters, showed a novel tRNA thiolation pattern, where the synthesis of only one thiolated nucleoside, ms2io6A, was decreased twofold. Based on our results, we suggest two principal distinct routes for thiolation of tRNA: (i) a direct sulfur transfer from IscS to the tRNA modifying enzymes ThiI and MnmA, which form s4U and the s2U moiety of (c)mnm5s2U, respectively; and (ii) an involvement of [Fe-S] proteins (an unidentified enzyme in the synthesis of s2C and MiaB in the synthesis of ms2io6A) in the transfer of sulfur to the tRNA. PMID:14729702

  16. Distinct Innate Responses are Induced by Attenuated Salmonella enterica serovar Typhimurium Mutants

    PubMed Central

    Powell, Daniel A.; Roberts, Lydia M.; Ledvina, Hannah E.; Sempowski, Gregory D.; Curtiss, Roy; Frelinger, Jeffrey A.

    2015-01-01

    Upon bacterial infection the host cells generate a wide variety of cytokines. Genetic attenuation of bacterial physiological pathogens can be accomplished not only by disruption of normal bacterial processes, but also by the loss of the ability to redirect the host immune system. We examined nine attenuated Salmonella Typhimurium mutants for their ability to replicate as well as the cytokines produced after infection of Bone Marrow Derived Macrophages (BMDM). Infection of BMDM with attenuated Salmonella mutants led to host cytokine patterns distinct from those that followed WT infection. Surprisingly, each bacterial mutant had a unique cytokine signature. Because some of the mutants induced an IL-10 response not seen in WT, we examined the role of IL-10 on Salmonella replication. Surprisingly, addition of IL-10 before or concurrent with infection restricted growth of WT Salmonella in BMDM. Bacterial attenuation is not a single process and results in attenuated host responses, which result in unique patterns for each attenuated mutants. PMID:26546408

  17. Diversity of Plasmids Encoding Virulence and Resistance Functions in Salmonella enterica subsp. enterica Serovar Typhimurium Monophasic Variant 4,[5],12:i:- Strains Circulating in Europe

    PubMed Central

    García, Patricia; Hopkins, Katie L.; García, Vanesa; Beutlich, Janine; Mendoza, M. Carmen; Threlfall, John; Mevius, Dik; Helmuth, Reiner; Rodicio, M. Rosario; Guerra, Beatriz

    2014-01-01

    Plasmids encoding resistance and virulence properties in multidrug resistant (MDR) Salmonella enterica (S.) serovar Typhimurium monophasic variant 4,[5],12:i:- isolates recovered from pigs and humans (2006-2008) in Europe were characterised. The isolates were selected based on the detection by PCR-amplification of S. Typhimurium virulence plasmid pSLT genes and were analysed by multi-locus sequence typing (MLST). The resistance genes present in the isolates and the association of these genes with integrons, transposons and insertion sequences were characterised by PCR-sequencing, and their plasmid location was determined by alkaline lysis and by S1-nuclease pulsed-field gel electrophoresis (PFGE) Southern-blot hybridisation. Plasmids were further analysed by replicon typing, plasmid MLST and conjugation experiments. The 10 S. 4,[5],12,i:- selected isolates belonged to ST19. Each isolate carried a large plasmid in which MDR with pSLT-associated virulence genes were located. After analysis, eight different plasmids of three incompatibility groups (IncA/C, IncR and IncF) were detected. Two IncA/C plasmids represented novel variants within the plasmid family of the S. 4,[5],12:i:- Spanish clone, and carried an empty class 1 integron with a conventional qacEΔ1-sul1 3′ conserved segment or an In-sul3 type III with estX-psp-aadA2-cmlA1-aadA1-qacH variable region linked to tnpA440-sul3, part of Tn2, Tn21 and Tn1721 transposons, and ISCR2. Four newly described IncR plasmids contained the resistance genes within In-sul3 type I (dfrA12-orfF-aadA2-cmlA1-aadA1-qacH/tnpA440-sul3) and part of Tn10 [tet(B)]. Two pSLT-derivatives with FIIs-ST1+FIB-ST17 replicons carried cmlA1-[aadA1-aadA2]-sul3-dfrA12 and blaTEM-1 genes linked to an In-sul3 type I integron and to Tn2, respectively. In conclusion, three emerging European clones of S. 4,[5],12:i:- harboured MDR plasmids encoding additional virulence functions that could contribute significantly to their evolutionary success. PMID

  18. Diversity of plasmids encoding virulence and resistance functions in Salmonella enterica subsp. enterica serovar Typhimurium monophasic variant 4,[5],12:i:- strains circulating in Europe.

    PubMed

    García, Patricia; Hopkins, Katie L; García, Vanesa; Beutlich, Janine; Mendoza, M Carmen; Threlfall, John; Mevius, Dik; Helmuth, Reiner; Rodicio, M Rosario; Guerra, Beatriz

    2014-01-01

    Plasmids encoding resistance and virulence properties in multidrug resistant (MDR) Salmonella enterica (S.) serovar Typhimurium monophasic variant 4,[5],12:i:- isolates recovered from pigs and humans (2006-2008) in Europe were characterised. The isolates were selected based on the detection by PCR-amplification of S. Typhimurium virulence plasmid pSLT genes and were analysed by multi-locus sequence typing (MLST). The resistance genes present in the isolates and the association of these genes with integrons, transposons and insertion sequences were characterised by PCR-sequencing, and their plasmid location was determined by alkaline lysis and by S1-nuclease pulsed-field gel electrophoresis (PFGE) Southern-blot hybridisation. Plasmids were further analysed by replicon typing, plasmid MLST and conjugation experiments. The 10 S. 4,[5],12,i:- selected isolates belonged to ST19. Each isolate carried a large plasmid in which MDR with pSLT-associated virulence genes were located. After analysis, eight different plasmids of three incompatibility groups (IncA/C, IncR and IncF) were detected. Two IncA/C plasmids represented novel variants within the plasmid family of the S. 4,[5],12:i:- Spanish clone, and carried an empty class 1 integron with a conventional qacEΔ1-sul1 3' conserved segment or an In-sul3 type III with estX-psp-aadA2-cmlA1-aadA1-qacH variable region linked to tnpA440-sul3, part of Tn2, Tn21 and Tn1721 transposons, and ISCR2. Four newly described IncR plasmids contained the resistance genes within In-sul3 type I (dfrA12-orfF-aadA2-cmlA1-aadA1-qacH/tnpA440-sul3) and part of Tn10 [tet(B)]. Two pSLT-derivatives with FIIs-ST1+FIB-ST17 replicons carried cmlA1-[aadA1-aadA2]-sul3-dfrA12 and blaTEM-1 genes linked to an In-sul3 type I integron and to Tn2, respectively. In conclusion, three emerging European clones of S. 4,[5],12:i:- harboured MDR plasmids encoding additional virulence functions that could contribute significantly to their evolutionary success.

  19. Characterization of Salmonella enterica Serovar Typhimurium Isolates from Pigs and Pig Environment-Related Sources and Evidence of New Circulating Monophasic Strains in Spain.

    PubMed

    Andrés-Barranco, Sara; Vico, Juan Pablo; Marín, Clara María; Herrera-León, Silvia; Mainar-Jaime, Raú Carlos

    2016-03-01

    A total of 117 Salmonella enterica serovar Typhimurium and 59 monophasic Salmonella Typhimurium (S. enterica serovar 4,[5],12:i:-) strains isolated between 2008 and 2012 from pig, wild bird, rodent, and farm environment samples from the northeast of Spain were characterized by phage typing, antibiotic susceptibility testing, and multiple-locus variable-number tandem repeat analysis in order to evaluate their phenotypic and genetic relatedness. In Salmonella 4,[5],12:i:-, the most prevalent phage types were U311 (40.7%) and DT195 (22%), which did not correspond with the so-called Spanish clone and generally showed a different resistance pattern (ASSuT). Antibiotic resistance was found in 85.8% of the isolates, with 94.1% of them displaying multidrug resistance. Multiple-locus variable-number tandem repeat analysis identified 92 different profiles, six of them shared by both serovars. The minimum spanning tree showed one major cluster that included 95% of the Salmonella 4,[5],12:i:- isolates, which came from different animal sources, geographic locations, and time periods, suggesting high clonality among those Salmonella strains and the ability to spread among pig farms. Overall, isolates of Salmonella 4,[5],12:i:- were more similar to European strains than to the well-characterized Spanish clone. The spread of these new strains of Salmonella 4,[5],12:i:- would likely have been favored by the important pig trade between this Spanish region and other European countries. The overall high prevalence of multidrug resistance observed in these new strains should be noted.

  20. Absence of Platelet Endothelial Cell Adhesion Molecule 1, PECAM-1/CD31, In Vivo Increases Resistance to Salmonella enterica Serovar Typhimurium in Mice

    PubMed Central

    Lovelace, Michael D.; Yap, May Lin; Yip, Jana; Muller, William; Wijburg, Odilia

    2013-01-01

    PECAM-1/CD31 is known to regulate inflammatory responses and exhibit pro- and anti-inflammatory functions. This study was designed to determine the functional role of PECAM-1 in susceptibility to murine primary in vivo infection with Salmonella enterica serovar Typhimurium and in in vitro inflammatory responses of peritoneal macrophages. Lectin profiling showed that cellular PECAM-1 and recombinant human PECAM-1-Ig chimera contain high levels of mannose sugars and N-acetylglucosamine. Consistent with this carbohydrate pattern, both recombinant human and murine PECAM-1-Ig chimeras were shown to bind S. Typhimurium in a dose-dependent manner in vitro. Using oral and fecal-oral transmission models of S. Typhimurium SL1344 infection, PECAM-1−/− mice were found to be more resistant to S. Typhimurium infection than wild-type (WT) C57BL/6 mice. While fecal shedding of S. Typhimurium was comparable in wild-type and PECAM-1−/− mice, the PECAM-1-deficient mice had lower bacterial loads in systemic organs such as liver, spleen, and mesenteric lymph nodes than WT mice, suggesting that extraintestinal dissemination was reduced in the absence of PECAM-1. This reduced bacterial load correlated with reduced tumor necrosis factor (TNF), interleukin-6 (IL-6), and monocyte chemoattractant protein (MCP) levels in sera of PECAM-1−/− mice. Following in vitro stimulation of macrophages with either whole S. Typhimurium, lipopolysaccharide (LPS) (Toll-like receptor 4 [TLR4] ligand), or poly(I·C) (TLR3 ligand), production of TNF and IL-6 by PECAM-1−/− macrophages was reduced. Together, these results suggest that PECAM-1 may have multiple functions in resistance to infection with S. Typhimurium, including binding to host cells, extraintestinal spread to deeper tissues, and regulation of inflammatory cytokine production by infected macrophages. PMID:23509149

  1. Effect of deletion of genes involved in lipopolysaccharide core and O-antigen synthesis on virulence and immunogenicity of Salmonella enterica serovar typhimurium.

    PubMed

    Kong, Qingke; Yang, Jiseon; Liu, Qing; Alamuri, Praveen; Roland, Kenneth L; Curtiss, Roy

    2011-10-01

    Lipopolysaccharide (LPS) is a major virulence factor of Salmonella enterica serovar Typhimurium and is composed of lipid A, core oligosaccharide (C-OS), and O-antigen polysaccharide (O-PS). While the functions of the gene products involved in synthesis of core and O-antigen have been elucidated, the effect of removing O-antigen and core sugars on the virulence and immunogenicity of Salmonella enterica serovar Typhimurium has not been systematically studied. We introduced nonpolar, defined deletion mutations in waaG (rfaG), waaI (rfaI), rfaH, waaJ (rfaJ), wbaP (rfbP), waaL (rfaL), or wzy (rfc) into wild-type S. Typhimurium. The LPS structure was confirmed, and a number of in vitro and in vivo properties of each mutant were analyzed. All mutants were significantly attenuated compared to the wild-type parent when administered orally to BALB/c mice and were less invasive in host tissues. Strains with ΔwaaG and ΔwaaI mutations, in particular, were deficient in colonization of Peyer's patches and liver. This deficiency could be partially overcome in the ΔwaaI mutant when it was administered intranasally. In the context of an attenuated vaccine strain delivering the pneumococcal antigen PspA, all of the mutations tested resulted in reduced immune responses against PspA and Salmonella antigens. Our results indicate that nonreversible truncation of the outer core is not a viable option for developing a live oral Salmonella vaccine, while a wzy mutant that retains one O-antigen unit is adequate for stimulating the optimal protective immunity to homologous or heterologous antigens by oral, intranasal, or intraperitoneal routes of administration.

  2. Igg Subclasses Targeting the Flagella of Salmonella enterica Serovar Typhimurium Can Mediate Phagocytosis and Bacterial Killing

    PubMed Central

    Goh, Yun Shan; Armour, Kathryn L; Clark, Michael R; Grant, Andrew J; Mastroeni, Pietro

    2016-01-01

    Invasive non-typhoidal Salmonella are a common cause of invasive disease in immuno-compromised individuals and in children. Multi-drug resistance poses challenges to disease control, with a critical need for effective vaccines. Flagellin is an attractive vaccine candidate due to surface exposure and high epitope copy number, but its potential as a target for opsonophacytic antibodies is unclear. We examined the effect of targeting flagella with different classes of IgG on the interaction between Salmonella Typhimurium and a human phagocyte-like cell line, THP-1. We tagged the FliC flagellar protein with a foreign CD52 mimotope (TSSPSAD) and bacteria were opsonized with a panel of humanised CD52 antibodies with the same antigen-binding V-region, but different constant regions. We found that IgG binding to flagella increases bacterial phagocytosis and reduces viable intracellular bacterial numbers. Opsonisation with IgG3, followed by IgG1, IgG4, and IgG2, resulted in the highest level of bacterial uptake and in the highest reduction in the intracellular load of viable bacteria. Taken together, our data provide proof-of-principle evidence that targeting flagella with antibodies can increase the antibacterial function of host cells, with IgG3 being the most potent subclass. These data will assist the rational design of urgently needed, optimised vaccines against iNTS disease. PMID:27366588

  3. Stress enhances the sensitivity of Salmonella enterica serovar Typhimurium to bacteriocins.

    PubMed

    Galvão, M F; Prudêncio, C V; Vanetti, M C D

    2015-05-01

    The aim of this study was to evaluate the potential application of bacteriocins against Gram-negative bacteria when associated with others food preservation methods. Salmonella was subjected to heat, cold, acid and chemical (with ethylenediaminetetracetate and trisodium phosphate) stresses. Then, the cells were recovered and subjected to treatment with bacteriocins (500 AU ml(-1) ) for 6 h. Heat and cold stress were those that promoted more sensitization to bactericidal activity of nisin. Under the same conditions, bovicin HC5 acted more rapidly than nisin reducing the number of viable cells to undetectable levels after 20 min of treatment. Similar results with use of nisin only were observed after 6 h of treatment. Stress conditions used in food industry, such as temperature and pH, and use of chelating agents or membrane disruptors, sensitized Salmonella Typhimurium cells to bacteriocins produced by lactic acid bacteria, such as nisin and bovicin HC5. Food preservation methods sensitized Gram-negative bacteria to bacteriocins activity, which demonstrate the potential of nisin and bovicin HC5 to inhibit the growth of Salmonella. © 2015 The Society for Applied Microbiology.

  4. Igg Subclasses Targeting the Flagella of Salmonella enterica Serovar Typhimurium Can Mediate Phagocytosis and Bacterial Killing.

    PubMed

    Goh, Yun Shan; Armour, Kathryn L; Clark, Michael R; Grant, Andrew J; Mastroeni, Pietro

    2016-05-30

    Invasive non-typhoidal Salmonella are a common cause of invasive disease in immuno-compromised individuals and in children. Multi-drug resistance poses challenges to disease control, with a critical need for effective vaccines. Flagellin is an attractive vaccine candidate due to surface exposure and high epitope copy number, but its potential as a target for opsonophacytic antibodies is unclear. We examined the effect of targeting flagella with different classes of IgG on the interaction between Salmonella Typhimurium and a human phagocyte-like cell line, THP-1. We tagged the FliC flagellar protein with a foreign CD52 mimotope (TSSPSAD) and bacteria were opsonized with a panel of humanised CD52 antibodies with the same antigen-binding V-region, but different constant regions. We found that IgG binding to flagella increases bacterial phagocytosis and reduces viable intracellular bacterial numbers. Opsonisation with IgG3, followed by IgG1, IgG4, and IgG2, resulted in the highest level of bacterial uptake and in the highest reduction in the intracellular load of viable bacteria. Taken together, our data provide proof-of-principle evidence that targeting flagella with antibodies can increase the antibacterial function of host cells, with IgG3 being the most potent subclass. These data will assist the rational design of urgently needed, optimised vaccines against iNTS disease.

  5. Distinct innate responses are induced by attenuated Salmonella enterica serovar Typhimurium mutants.

    PubMed

    Powell, Daniel A; Roberts, Lydia M; Ledvina, Hannah E; Sempowski, Gregory D; Curtiss, Roy; Frelinger, Jeffrey A

    2016-01-01

    Upon bacterial infection the host cells generate a wide variety of cytokines. Genetic attenuation of bacterial physiological pathogens can be accomplished not only by disruption of normal bacterial processes, but also by the loss of the ability to redirect the host immune system. We examined nine attenuated Salmonella Typhimurium mutants for their ability to replicate as well as the cytokines produced after infection of Bone Marrow Derived Macrophages (BMDM). Infection of BMDM with attenuated Salmonella mutants led to host cytokine patterns distinct from those that followed WT infection. Surprisingly, each bacterial mutant had a unique cytokine signature. Because some of the mutants induced an IL-10 response not seen in WT, we examined the role of IL-10 on Salmonella replication. Surprisingly, addition of IL-10 before or concurrent with infection restricted growth of WT Salmonella in BMDM. Bacterial attenuation is not a single process and results in attenuated host responses, which result in unique patterns for each attenuated mutants. Copyright © 2015 Elsevier Inc. All rights reserved.

  6. Protection of mice from Brucella infection by immunization with attenuated Salmonella enterica serovar typhimurium expressing A L7/L12 and BLS fusion antigen of Brucella.

    PubMed

    Zhao, Zhongpeng; Li, Min; Luo, Deyan; Xing, Li; Wu, Shuo; Duan, Yueqiang; Yang, Penghui; Wang, Xiliang

    2009-08-20

    This study describes the potential use of attenuated Salmonella enterica serovar Typhimurium Strains (S. typhimurium) to express and deliver a L7/L12 and BLS fusion antigen of Brucella as a vaccination strategy to prevent Brucella infection in mice. S. typhimurium X4072 that contained a pTrc99A-BLS-L7/L12 plasmid, designated X4072bl, can deliver a L7/L12 and BLS fusion antigen expressed by the bacterium itself, while S. typhimurium X4550 that contained an asd-pVAX1-BLS-L7/L12 (asd-pBL) plasmid, designated X4550bl, can deliver the antigen to be expressed in target eukaryotic cells. When orally administered to BALB/c mice, both attenuated carrier strains were able to elicit mucosal and systemic immunity, which induced protection against B. abortus 544 infection in mice. The immunogenicity and protective efficacy of X4072bl and X4550bl were compared with a recombinant BLS-L7/L12 fusion protein vaccine (rBL) and a pVAX1-BLS-L7/L12 DNA vaccine (pBL) in this study. When rBL and pBL were intramuscularly injected into mice, both vaccines could also elicit comparable humoral and cellular immune responses, but not mucosal immunity, which therefore induced lower protection. Taken together, Salmonella-based subunit vaccines are a promising vaccine strategy in the prevention of Brucella infection.

  7. Enhancement of host immune responses by oral vaccination to Salmonella enterica serovar Typhimurium harboring both FliC and FljB flagella.

    PubMed

    Eom, Jeong Seon; Seok Kim, Jin; Im Jang, Jung; Kim, Bae-Hoon; Young Yoo, So; Hyeon Choi, Ji; Bang, Iel-Soo; Lee, In Soo; Keun Park, Yong

    2013-01-01

    Flagellin, the structural component of the flagellar filament in various motile bacteria, can contribute to the activation of NF-κB and proinflammatory cytokine expression during the innate immune response in host cells. Thus, flagellin proteins represent a particularly attractive target for the development of vaccine candidates. In this study, we investigated the immune response by increasing the flagella number in the iacP mutant strain and the adjuvant activity of the flagellin component FljB of Salmonella enterica serovar Typhimurium. We found that the iacP mutant strain expresses two flagellin proteins (FliC and FljB), which result in increased NF-κB-dependent gene expression in bone marrow derived macrophages. Using an oral immunization mouse model, we observed that the administration of a live attenuated S. typhimurium BRD509 strain expressing the FliC and FljB flagellins induced significantly enhanced flagellin-specific IgG responses in the systemic compartment. The mice immunized with the recombinant attenuated S. typhimurium strain that has two types of flagella were protected from lethal challenge with the Salmonella SL1344 strain. These results indicate that overexpression of flagella in the iacP mutant strain enhance the induction of an antigen-specific immune responses in macrophage cell, and both the FliC and FljB flagellar filament proteins-producing S. typhimurium can induce protective immune responses against salmonellosis.

  8. Enhancement of Host Immune Responses by Oral Vaccination to Salmonella enterica serovar Typhimurium Harboring Both FliC and FljB Flagella

    PubMed Central

    Eom, Jeong Seon; Seok Kim, Jin; Im Jang, Jung; Kim, Bae-Hoon; Young Yoo, So; Hyeon Choi, Ji; Bang, Iel-Soo; Lee, In Soo; Keun Park, Yong

    2013-01-01

    Flagellin, the structural component of the flagellar filament in various motile bacteria, can contribute to the activation of NF-κB and proinflammatory cytokine expression during the innate immune response in host cells. Thus, flagellin proteins represent a particularly attractive target for the development of vaccine candidates. In this study, we investigated the immune response by increasing the flagella number in the iacP mutant strain and the adjuvant activity of the flagellin component FljB of Salmonella enterica serovar Typhimurium. We found that the iacP mutant strain expresses two flagellin proteins (FliC and FljB), which result in increased NF-κB-dependent gene expression in bone marrow derived macrophages. Using an oral immunization mouse model, we observed that the administration of a live attenuated S. typhimurium BRD509 strain expressing the FliC and FljB flagellins induced significantly enhanced flagellin-specific IgG responses in the systemic compartment. The mice immunized with the recombinant attenuated S. typhimurium strain that has two types of flagella were protected from lethal challenge with the Salmonella SL1344 strain. These results indicate that overexpression of flagella in the iacP mutant strain enhance the induction of an antigen-specific immune responses in macrophage cell, and both the FliC and FljB flagellar filament proteins-producing S. typhimurium can induce protective immune responses against salmonellosis. PMID:24069357

  9. Non-Saccharomyces yeasts protect against epithelial cell barrier disruption induced by Salmonella enterica subsp. enterica serovar Typhimurium.

    PubMed

    Smith, I M; Baker, A; Arneborg, N; Jespersen, L

    2015-11-01

    The human gastrointestinal epithelium makes up the largest barrier separating the body from the external environment. Whereas invasive pathogens cause epithelial barrier disruption, probiotic micro-organisms modulate tight junction regulation and improve epithelial barrier function. In addition, probiotic strains may be able to reduce epithelial barrier disruption caused by pathogenic species. The aim of this study was to explore non-Saccharomyces yeast modulation of epithelial cell barrier function in vitro. Benchmarking against established probiotic strains, we evaluated the ability of four nonpathogenic yeast species to modulate transepithelial electrical resistance (TER) across a monolayer of differentiated human colonocytes (Caco-2 cells). Further, we assessed yeast modulation of a Salmonella Typhimurium-induced epithelial cell barrier function insult. Our findings demonstrate distinct patterns of non-Saccharomyces yeast modulation of epithelial cell barrier function. While the established probiotic yeast Saccharomyces boulardii increased TER across a Caco-2 monolayer by 30%, Kluyveromyces marxianus exhibited significantly stronger properties of TER enhancement (50% TER increase). In addition, our data demonstrate significant yeast-mediated modulation of Salmonella-induced epithelial cell barrier disruption and identify K. marxianus and Metschnikowia gruessii as two non-Saccharomyces yeasts capable of protecting human epithelial cells from pathogen invasion. This study demonstrates distinct patterns of non-Saccharomyces yeast modulation of epithelial cell barrier function in vitro. Further, our data demonstrate significant yeast-mediated modulation of Salmonella Typhimurium-induced epithelial cell barrier disruption and identify Kluyveromyces marxianus and Metschnikowia gruessii as two non-Saccharomyces yeasts capable of protecting human epithelial cells from pathogen invasion. This study is the first to demonstrate significant non-Saccharomyces yeast

  10. Coordinated Regulation of Virulence during Systemic Infection of Salmonella enterica serovar Typhimurium

    SciTech Connect

    Yoon, Hyunjin; McDermott, Jason E.; Porwollik, Steffen; Mcclelland, Michael; Heffron, Fred

    2009-02-20

    Salmonella must respond to a myriad of environmental cues during infection of a mouse and express specific subsets of genes in a temporal and spatial manner to subvert the host defense mechanisms but these regulatory pathways are poorly established. To unravel how micro-environmental signals are processed and integrated into coordinated action, we constructed in-frame non-polar deletions of 84 regulators inferred to play a role in Salmonella typhimurium virulence and tested them in three virulence assays (intraperitoneal (i.p.), and intragastric (i.g.) infection in BALB/c mice, and persistence in SvJ129 mice). Overall 36 regulators were identified that were less virulent in at least one assay, and of those, 15 regulators were required for systemic mouse infection in an acute infection model. As a first step towards understanding the interplay between a pathogen and its host from a systems biology standpoint we focused on these 15 genes. Transcriptional profiles were obtained for each of these 15 regulators from strains grown under four different environmental conditions. These results as well as publicly available transcriptional profiles were analyzed using both network inference and cluster analysis algorithms. The analysis predicts a regulatory network in which all 15 regulators control a specific set of genes necessary for Salmonella to cause systemic infection. We tested the regulatory model by expressing a subset of the regulators in trans and monitoring transcription of 7 known virulence factors located within Salmonella pathogenicity island 2 (SPI-2). These experiments validated the regulatory model and showed that, for these 7 genes, the response regulator SsrB and the marR type regulator SlyA co-regulate in a regulatory cascade by integrating multiple signals.

  11. Effects of Physical Properties of Feed on Microbial Ecology and Survival of Salmonella enterica Serovar Typhimurium in the Pig Gastrointestinal Tract

    PubMed Central

    Mikkelsen, Lene Lind; Naughton, Patrick J.; Hedemann, Mette S.; Jensen, Bent Borg

    2004-01-01

    A two-by-two factorial experiment with pigs was conducted to study the effect of feed grinding (fine and coarse) and feed processing (pelleted and nonpelleted) on physicochemical properties, microbial populations, and survival of Salmonella enterica serovar Typhimurium DT12 in the gastrointestinal tracts of pigs. Results demonstrated a strong effect of diet on parameters measured in the stomachs of the pigs, whereas the effect was less in the other parts of the gastrointestinal tract. Pigs fed the coarse nonpelleted (C-NP) diet showed more solid gastric content with higher dry matter content than pigs fed the fine nonpelleted (F-NP), coarse pelleted (C-P), or fine pelleted (F-P) diet. Pigs fed the C-NP diet also showed significantly increased number of anaerobic bacteria (P < 0.05), increased concentrations of organic acids, and reduced pH in the stomach. In addition, pigs fed the C-NP diet showed increased in vitro death rate of S. enterica serovar Typhimurium DT12 in content from the stomach (P < 0.001). Pigs fed the C-NP diet had a significantly higher concentration of undissociated lactic acid in gastric content than pigs fed the other diets (P < 0.001). A strong correlation between the concentration of undissociated lactic acid and the death rate of S. enterica serovar Typhimurium DT12 was found. In the distal small intestine, cecum, and midcolon, significantly lower numbers of coliform bacteria were observed in pigs fed the coarse diets than in pigs fed the fine diets (P < 0.01). Pigs fed the C-NP diet showed the lowest number of coliform bacteria in these segments of the gastrointestinal tract. Pigs fed the coarse diets showed increased concentration of butyric acid in the cecum (P < 0.05) and colon (P < 0.10) compared with pigs fed the fine diets. It was concluded that feeding a coarsely ground meal feed to pigs changes the physicochemical and microbial properties of content in the stomach, which decreases the survival of Salmonella during passage through

  12. Toll-like receptor 4 signalling through MyD88 is essential to control Salmonella enterica serovar typhimurium infection, but not for the initiation of bacterial clearance.

    PubMed

    Talbot, Suzanne; Tötemeyer, Sabine; Yamamoto, Masahiro; Akira, Shizuo; Hughes, Katherine; Gray, David; Barr, Tom; Mastroeni, Pietro; Maskell, Duncan J; Bryant, Clare E

    2009-12-01

    Toll-like receptor-4 (TLR4) is important in protection against lethal Salmonella enterica serovar Typhimurium (S. Typhimurium) infection. Control of the early stages of sublethal S. Typhimurium infection in mice depends on TLR4-dependent activation of macrophages and natural killer (NK) cells to drive an inflammatory response. TLR4 signals through the adapter proteins Mal/MyD88 and TRIF-related adaptor molecule (TRAM)/TIR-domain-containing adaptor-inducing interferon-b (TRIF). In the mouse typhoid model we showed that TLR4 and MyD88, but not Mal or TRIF, are essential for the control of exponential S. Typhimurium growth. TRIF(-/-) mice have a higher bacterial load in comparison with wild-type mice during a sublethal infection because TRIF is important for bacterial killing during the first day of systemic disease. Minimal pro-inflammatory responses were induced by S. Typhimurium infection of macrophages from TLR4(-/-), MyD88(-/-) and TRIF(-/-) mice in vitro. Pro-inflammatory responses from Mal(-/-) macrophages were similar to those from wild-type cells. The pro-inflammatory responses of TRIF(-/-) macrophages were partially restored by the addition of interferon-gamma (IFN-gamma), and TRIF(-/-) mice produced markedly enhanced IFN-gamma levels, in comparison to wild-type mice, probably explaining why bacterial growth can be controlled in these mice. TLR4(-/-), MyD88(-/-), TRIF(-/-) and Mal(-/-) mice all initiated clearance of S. Typhimurium, suggesting that TLR4 signalling is not important in driving bacterial clearance in comparison to its critical role in controlling early bacterial growth in mouse typhoid.

  13. Poultry Body Temperature Contributes to Invasion Control through Reduced Expression of Salmonella Pathogenicity Island 1 Genes in Salmonella enterica Serovars Typhimurium and Enteritidis

    PubMed Central

    Petri, Nicholas; Daron, Caitlyn; Pereira, Rafaela; Mendoza, Mary; Hassan, Hosni M.; Koci, Matthew D.

    2015-01-01

    Salmonella enterica serovars Typhimurium (S. Typhimurium) and Enteritidis (S. Enteritidis) are foodborne pathogens, and outbreaks are often associated with poultry products. Chickens are typically asymptomatic when colonized by these serovars; however, the factors contributing to this observation are uncharacterized. Whereas symptomatic mammals have a body temperature between 37°C and 39°C, chickens have a body temperature of 41°C to 42°C. Here, in vivo experiments using chicks demonstrated that numbers of viable S. Typhimurium or S. Enteritidis bacteria within the liver and spleen organ sites were ≥4 orders of magnitude lower than those within the ceca. When similar doses of S. Typhimurium or S. Enteritidis were given to C3H/HeN mice, the ratio of the intestinal concentration to the liver/spleen concentration was 1:1. In the avian host, this suggested poor survival within these tissues or a reduced capacity to traverse the host epithelial layer and reach liver/spleen sites or both. Salmonella pathogenicity island 1 (SPI-1) promotes localization to liver/spleen tissues through invasion of the epithelial cell layer. Following in vitro growth at 42°C, SPI-1 genes sipC, invF, and hilA and the SPI-1 rtsA activator were downregulated compared to expression at 37°C. Overexpression of the hilA activators fur, fliZ, and hilD was capable of inducing hilA-lacZ at 37°C but not at 42°C despite the presence of similar levels of protein at the two temperatures. In contrast, overexpression of either hilC or rtsA was capable of inducing hilA and sipC at 42°C. These data indicate that physiological parameters of the poultry host, such as body temperature, have a role in modulating expression of virulence. PMID:26386070

  14. Comparative proteomic analysis of Salmonella enterica serovar Typhimurium ppGpp-deficient mutant to identify a novel virulence protein required for intracellular survival in macrophages

    PubMed Central

    2010-01-01

    Background The global ppGpp-mediated stringent response in pathogenic bacteria plays an important role in the pathogenesis of bacterial infections. In Salmonella enterica serovar Typhimurium (S. Typhimurium), several genes, including virulence genes, are regulated by ppGpp when bacteria are under the stringent response. To understand the control of virulence genes by ppGpp in S. Typhimurium, agarose 2-dimensional electrophoresis (2-DE) combined with mass spectrometry was used and a comprehensive 2-DE reference map of amino acid-starved S. Typhimurium strain SH100, a derivative of ATCC 14028, was established. Results Of the 366 examined spots, 269 proteins were successfully identified. The comparative analysis of the wild-type and ppGpp0 mutant strains revealed 55 proteins, the expression patterns of which were affected by ppGpp. Using a mouse infection model, we further identified a novel virulence-associated factor, STM3169, from the ppGpp-regulated and Salmonella-specific proteins. In addition, Salmonella strains carrying mutations in the gene encoding STM3169 showed growth defects and impaired growth within macrophage-like RAW264.7 cells. Furthermore, we found that expression of stm3169 was controlled by ppGpp and SsrB, a response regulator of the two-component system located on Salmonella pathogenicity island 2. Conclusions A proteomic approach using a 2-DE reference map can prove a powerful tool for analyzing virulence factors and the regulatory network involved in Salmonella pathogenesis. Our results also provide evidence of a global response mediated by ppGpp in S. enterica. PMID:21176126

  15. A five-strain probiotic combination reduces pathogen shedding and alleviates disease signs in pigs challenged with Salmonella enterica Serovar Typhimurium.

    PubMed

    Casey, Pat G; Gardiner, Gillian E; Casey, Garrett; Bradshaw, Bernard; Lawlor, Peadar G; Lynch, P Brendan; Leonard, Finola C; Stanton, Catherine; Ross, R Paul; Fitzgerald, Gerald F; Hill, Colin

    2007-03-01

    Salmonella spp. infection is a major cause of gastroenteritis, with many thousands of cases reported in the European Union every year. The use of probiotics offers the potential to improve this situation. Here, we investigate the effects of oral treatment of pigs with a defined lactic acid bacteria culture mixture on both clinical and microbiological signs of Salmonella enterica serovar Typhimurium infection. Fifteen weaned pigs blocked by sex and weight were administered control milk or a mixture of five probiotic strains as either a milk fermentate or milk suspension for a total of 30 days. The mixture consisted of two strains of Lactobacillus murinus and one strain each of Lactobacillus salivarius subsp. salivarius, Lactobacillus pentosus, and Pediococcus pentosaceous. Following probiotic administration for 6 days, animals were challenged orally with serovar Typhimurium; the health of the animals and the microbiological composition of their feces were monitored for 23 days postinfection. Animals treated with probiotic showed reduced incidence, severity, and duration of diarrhea. These animals also gained weight at a greater rate than control pigs administered skim milk. Mean fecal numbers of Salmonella were significantly reduced in probiotic-treated animals at 15 days postinfection (P = 0.01). The administered probiotic bacteria improved both the clinical and microbiological outcome of Salmonella infection. These strains offer significant benefit for use in the food industry and may have potential in human applications.

  16. The Lipopolysaccharide Structures of Salmonella enterica Serovar Typhimurium and Neisseria gonorrhoeae Determine the Attachment of Human Mannose-Binding Lectin to Intact Organisms

    PubMed Central

    Devyatyarova-Johnson, Marina; Rees, Ian H.; Robertson, Brian D.; Turner, Malcolm W.; Klein, Nigel J.; Jack, Dominic L.

    2000-01-01

    Mannose-binding lectin (MBL) is an important component of the innate immune system. It binds to the arrays of sugars commonly presented by microorganisms and activates the complement system independently of antibody. Despite detailed knowledge of the stereochemical basis of MBL binding, relatively little is known about how bacterial surface structures influence binding of the lectin. Using flow cytometry, we have measured the binding of MBL to a range of mutants of Salmonella enterica serovar Typhimurium and Neisseria gonorrhoeae which differ in the structure of expressed lipopolysaccharide (LPS). For both organisms, the possession of core LPS structures led to avid binding of MBL, which was abrogated by the addition of O antigen (Salmonella serovar Typhimurium) or sialic acid (N. gonorrhoeae). Truncation of the LPS within the core led to lower levels of MBL binding. It was not possible to predict the magnitude of MBL binding from the identity of the LPS terminal sugar alone, indicating that the three-dimensional disposition of LPS molecules is probably also of importance in determining MBL attachment. These results further support the hypothesis that LPS structure is a major determinant of MBL binding. PMID:10858200

  17. Long-term dissemination of CTX-M-5-producing hypermutable Salmonella enterica serovar typhimurium sequence type 328 strains in Russia, Belarus, and Kazakhstan.

    PubMed

    Kozyreva, Varvara K; Ilina, Elena N; Malakhova, Maja V; Carattoli, Alessandra; Azizov, Ilya S; Tapalski, Dmitry V; Kozlov, Roman S; Edelstein, Mikhail V

    2014-09-01

    In this paper, we present evidence of long-term circulation of cefotaxime-resistant clonally related Salmonella enterica serovar Typhimurium strains over a broad geographic area. The genetic relatedness of 88 isolates collected from multiple outbreaks and sporadic cases of nosocomial salmonellosis in various parts of Russia, Belarus, and Kazakhstan from 1996 to 2009 was established by multilocus tandem-repeat analysis (MLVA) and multilocus sequence typing (MLST). The isolates belong to sequence type 328 (ST328) and produce CTX-M-5 β-lactamase, whose gene is carried by highly related non-self-conjugative but mobilizable plasmids. Resistance to nalidixic acid and low-level resistance to ciprofloxacin is present in 37 (42%) of the isolates and in all cases is determined by various single point mutations in the gyrA gene quinolone resistance-determining region (QRDR). Isolates of the described clonal group exhibit a hypermutable phenotype that probably facilitates independent acquisition of quinolone resistance mutations.

  18. Semiquantitative Analysis of the Red, Dry, and Rough Colony Morphology of Salmonella enterica Serovar Typhimurium and Escherichia coli Using Congo Red.

    PubMed

    Cimdins, Annika; Simm, Roger

    2017-01-01

    The Congo Red (CR) assay is a standard biofilm test assessing the colony morphology of bacteria growing on agar plates supplemented with the diazo dye Congo Red. Biofilm forming Salmonella enterica serovar Typhimurium and Escherichia coli produce a red, dry, and rough (rdar) morphotype on CR-plates. The phenotype is characterized by staining of the extracellular matrix components curli (brown color) and cellulose (pink color) by CR. This method allows semiquantitative determination of the expression level of the individual matrix components and dissection of the regulatory networks controlling their production in response to c-di-GMP levels. Here, we describe the CR-assay and its variations and discuss the effect of deletion or overexpression of c-di-GMP turnover proteins on colony morphology.

  19. Model-driven discovery of synergistic inhibitors against E. coli and S. enterica serovar Typhimurium targeting a novel synthetic lethal pair, aldA and prpC

    PubMed Central

    Aziz, Ramy K.; Khaw, Valerie L.; Monk, Jonathan M.; Brunk, Elizabeth; Lewis, Robert; Loh, Suh I.; Mishra, Arti; Nagle, Amrita A.; Satyanarayana, Chitkala; Dhakshinamoorthy, Saravanakumar; Luche, Michele; Kitchen, Douglas B.; Andrews, Kathleen A.; Palsson, Bernhard Ø.; Charusanti, Pep

    2015-01-01

    Mathematical models of biochemical networks form a cornerstone of bacterial systems biology. Inconsistencies between simulation output and experimental data point to gaps in knowledge about the fundamental biology of the organism. One such inconsistency centers on the gene aldA in Escherichia coli: it is essential in a computational model of E. coli metabolism, but experimentally it is not. Here, we reconcile this disparity by providing evidence that aldA and prpC form a synthetic lethal pair, as the double knockout could only be created through complementation with a plasmid-borne copy of aldA. Moreover, virtual and biological screening against the two proteins led to a set of compounds that inhibited the growth of E. coli and Salmonella enterica serovar Typhimurium synergistically at 100–200 μM individual concentrations. These results highlight the power of metabolic models to drive basic biological discovery and their potential use to discover new combination antibiotics. PMID:26441892

  20. Multidrug-resistant Salmonella enterica Serovar Typhimurium Monophasic Variant 4,12:i:- Isolated from Asymptomatic Wildlife in a Catalonian Wildlife Rehabilitation Center, Spain.

    PubMed

    Molina-López, Rafael A; Vidal, Anna; Obón, Elena; Martín, Marga; Darwich, Laila

    2015-07-01

    Wildlife can act as long-term asymptomatic reservoirs for zoonotic bacteria, such as Salmonella. The prevalence and antimicrobial-susceptibility profiles of Salmonella spp. were assessed in 263 cases in wildlife from 22 animal orders from a wildlife rehabilitation center in Catalonia (NE Spain), September 2013-May 2014. Eleven of 263 tested animals were positive for Salmonella spp., representing an overall prevalence of 4.2%. Prevalences by taxonomic categories were 2% in mammals, 4.7% in birds, and 4.5% in reptiles. By species, one each of European hedgehog (Erinaceus europeus; from a sample of n = 26), Eurasian Eagle Owl (Bubo bubo; n = 2), Barn Owl (Tyto alba; n = 3), Tawny Owl (Strix aluco; n = 20), Egyptian Vulture (Neophron percnopterus; n = 1), Griffon Vulture (Gyps fulvus; n = 1), and Hoopoe (Upupa epops; n = 2), and two each Common Kestrels (Falco tinnunculus; n = 16) and pond sliders (Trachemys scripta; n = 25) were positive for Salmonella. By serotyping, seven of eleven isolates were classified as S. enterica subsp. enterica serovar Typhimurium, and five of seven belonged to the monophasic variant 4,12:i:-. All the monophasic variants were isolated from birds (4/5 in raptors) and showed a multidrug-resistance (MDR) profile to at least ampicillin, streptomycin, sulfonamide, and tetracycline (R-type ASSuT), and up to 12 antibiotics. The large proportion of S. Typhimurium monophasic MDR strains detected in wildlife never treated with antibiotics, especially in raptors, adds more complexity to the epidemiologic control of one of the most frequent serovars involved in human and livestock infection.

  1. Effects of dam and/or seqA mutations on the fatty acid and phospholipid membrane composition of Salmonella enterica serovar Typhimurium.

    PubMed

    Aloui, Amine; Mihoub, Mouadh; Sethom, Mohamed Marwan; Chatti, Abdelwaheb; Feki, Moncef; Kaabachi, Naziha; Landoulsi, Ahmed

    2010-05-01

    We examined the phospholipids (Phls) and the membrane fatty acid (FA) composition in Salmonella enterica serovar Typhimurium dam and/or seqA mutants. Phosphatidylglycerol, phosphatidylethanolamine (PE), and cardiolipin (CL) are the major Phls present in all the strains and accounted for greater than 95% of the total lipid phosphorus. Phosphatidic acid and phosphatidylserine are the minor ones. The seqA mutant showed a decrease in PE and an increase in CL and phosphatidylglycerol proportion compared with the wild-type strain. The same changes were observed with the seqA dam double mutant. However, the dam mutation caused an unusual accumulation of CL with a significant decrease in the PE content, compared with the isogenic wild-type strain. FA composition of the total lipids and the different fractions containing Phls have been determined. The major saturated FAs (SFAs) and unsaturated FAs (UFAs) found were C(14:0), C(16:0) and C(16:1w7), C(18:1w9), respectively. Cyclic FAs, cyc(17:0) and cyc(19:0), were also present in appreciable amounts. Moreover, dam and/or seqA mutations caused a decrease in UFA/SFA ratio and there was a progressive reduction in the content of C(16:1w7) and C(18:1w9), going through the order seqA, dam/seqA, and dam mutants. This decrease in UFA content was compensated for in all strains by an increase in the corresponding C(17-) and C(19-) cyclic FAs. So these UFAs were converted to their cyclopropane derivatives, which resulted in a low UFA/SFA ratio. SeqA and Dam proteins might regulate FA biosynthesis and Phls composition of Salmonella enterica serovar Typhimurium.

  2. Revival of an old problem: an increase in Salmonella enterica serovar Typhimurium definitive phage type 8 infections in 2010 in England and Northern Ireland linked to duck eggs.

    PubMed

    Noble, D J; Lane, C; Little, C L; Davies, R; De Pinna, E; Larkin, L; Morgan, D

    2012-01-01

    Salmonella enterica serovar Typhimurium definitive phage type (DT) 8 is uncommon in humans in the UK. In July 2010, the Health Protection Agency reported an excess isolation rate of pan-susceptible S. Typhimurium DT8 in England and Northern Ireland. By the end of October, this amounted to 81 laboratory-confirmed human cases for all regions of England and Northern Ireland in 2010, an increase of 26% and 41% on 2009 and 2008, respectively. Descriptive epidemiological investigation found a strong association with infection and consumption of duck eggs. Duck eggs contaminated with S. Typhimurium DT8 were collected from a patient's home and also at farms in the duck-egg supply chain. Although duck eggs form a small part of total UK eggs sales, there has been significant growth in sales in recent years. This is the first known outbreak of salmonellosis linked to duck eggs in the UK since 1949 and highlighted the impact of a changing food source and market on the re-emergence of salmonellosis linked to duck eggs. Control measures by the duck-egg industry should be improved along with a continued need to remind the public and commercial caterers of the potential high risks of contracting salmonellosis from duck eggs.

  3. Gre factors-mediated control of hilD transcription is essential for the invasion of epithelial cells by Salmonella enterica serovar Typhimurium

    PubMed Central

    El Mouali, Youssef; Römling, Ute

    2017-01-01

    The invasion of epithelial cells by Salmonella enterica serovar Typhimurium is a very tightly regulated process. Signaling cascades triggered by different environmental and physiological signals converge to control HilD, an AraC regulator that coordinates the expression of several virulence factors. The expression of hilD is modulated at several steps of the expression process. Here, we report that the invasion of epithelial cells by S. Typhimurium strains lacking the Gre factors, GreA and GreB, is impaired. By interacting with the RNA polymerase secondary channel, the Gre factors prevent backtracking of paused complexes to avoid arrest during transcriptional elongation. Our results indicate that the Gre factors are required for the expression of the bacterial factors needed for epithelial cell invasion by modulating expression of HilD. This regulation does not occur at transcription initiation and depends on the capacity of the Gre factors to prevent backtracking of the RNA polymerase. Remarkably, genetic analyses indicate that the 3’-untranslated region (UTR) of hilD is required for Gre-mediated regulation of hilD expression. Our data provide new insight into the complex regulation of S. Typhimurium virulence and highlight the role of the hilD 3’-UTR as a regulatory motif. PMID:28426789

  4. Salmonella enterica serovar typhimurium colonizing the lumen of the chicken intestine grows slowly and upregulates a unique set of virulence and metabolism genes.

    PubMed

    Harvey, P C; Watson, M; Hulme, S; Jones, M A; Lovell, M; Berchieri, A; Young, J; Bumstead, N; Barrow, P

    2011-10-01

    The pattern of global gene expression in Salmonella enterica serovar Typhimurium bacteria harvested from the chicken intestinal lumen (cecum) was compared with that of a late-log-phase LB broth culture using a whole-genome microarray. Levels of transcription, translation, and cell division in vivo were lower than those in vitro. S. Typhimurium appeared to be using carbon sources, such as propionate, 1,2-propanediol, and ethanolamine, in addition to melibiose and ascorbate, the latter possibly transformed to d-xylulose. Amino acid starvation appeared to be a factor during colonization. Bacteria in the lumen were non- or weakly motile and nonchemotactic but showed upregulation of a number of fimbrial and Salmonella pathogenicity island 3 (SPI-3) and 5 genes, suggesting a close physical association with the host during colonization. S. Typhimurium bacteria harvested from the cecal mucosa showed an expression profile similar to that of bacteria from the intestinal lumen, except that levels of transcription, translation, and cell division were higher and glucose may also have been used as a carbon source.

  5. Toll-Like Receptor Activation by Generalized Modules for Membrane Antigens from Lipid A Mutants of Salmonella enterica Serovars Typhimurium and Enteritidis

    PubMed Central

    Rossi, Omar; Caboni, Mariaelena; Negrea, Aurel; Necchi, Francesca; Alfini, Renzo; Micoli, Francesca; Saul, Allan; MacLennan, Calman A.

    2016-01-01

    Invasive nontyphoidal Salmonella (iNTS) disease is a neglected disease with high mortality in children and HIV-positive individuals in sub-Saharan Africa, caused primarily by Africa-specific strains of Salmonella enterica serovars Typhimurium and Enteritidis. A vaccine using GMMA (generalized modules for membrane antigens) from S. Typhimurium and S. Enteritidis containing lipid A modifications to reduce potential in vivo reactogenicity is under development. GMMA with penta-acylated lipid A showed the greatest reduction in the level of cytokine release from human peripheral blood monocytes from that for GMMA with wild-type lipid A. Deletion of the lipid A modification genes msbB and pagP was required to achieve pure penta-acylation. Interestingly, ΔmsbB ΔpagP GMMA from S. Enteritidis had a slightly higher stimulatory potential than those from S. Typhimurium, a finding consistent with the higher lipopolysaccharide (LPS) content and Toll-like receptor 2 (TLR2) stimulatory potential of the former. Also, TLR5 ligand flagellin was found in Salmonella GMMA. No relevant contribution to the stimulatory potential of GMMA was detected even when the flagellin protein FliC from S. Typhimurium was added at a concentration as high as 10% of total protein, suggesting that flagellin impurities are not a major factor for GMMA-mediated immune stimulation. Overall, the stimulatory potential of S. Typhimurium and S. Enteritidis ΔmsbB ΔpagP GMMA was close to that of Shigella sonnei GMMA, which are currently in phase I clinical trials. PMID:26865597

  6. Removal of the phage-shock protein PspB causes reduction of virulence in Salmonella enterica serovar Typhimurium independently of NRAMP1.

    PubMed

    Wallrodt, Inke; Jelsbak, Lotte; Thomsen, Line E; Brix, Lena; Lemire, Sébastien; Gautier, Laurent; Nielsen, Dennis S; Jovanovic, Goran; Buck, Martin; Olsen, John E

    2014-06-01

    The phage-shock protein (Psp) system is believed to manage membrane stress in all Enterobacteriaceae and has recently emerged as being important for virulence in several pathogenic species of this phylum. The core of the Psp system consists of the pspA-D operon and the distantly located pspG gene. In Salmonella enterica serovar Typhimurium (S. Typhimurium), it has recently been reported that PspA is essential for systemic infection of mice, but only in NRAMP1(+) mice, signifying that attenuation is related to coping with divalent cation starvation in the intracellular environment. In the present study, we investigated the contribution of individual psp genes to virulence of S. Typhimurium. Interestingly, deletion of the whole pspA-D set of genes caused attenuation in both NRAMP1(+) and NRAMP1(-) mice, indicating that one or more of the psp genes contribute to virulence independently of NRAMP1 expression in the host. Investigations of single gene mutants showed that knock out of pspB reduced virulence in both types of mice, while deletion of pspA only caused attenuation in NRAMP1(+) mice, and deletion of pspD had a minor effect in NRAMP1(-) mice, while deletions of either pspC or pspG did not affect virulence. Experiments addressed at elucidating the role of PspB in virulence revealed that PspB is dispensable for uptake to and intracellular replication in cultured macrophages and resistance to complement-induced killing. Furthermore, the Psp system of S. Typhimurium was dispensable during pIV-induced secretin stress. In conclusion, our results demonstrate that removal of PspB reduces virulence in S. Typhimurium independently of host NRAMP1 expression, demonstrating that PspB has roles in intra-host survival distinct from the reported contributions of PspA.

  7. Characterization of the Salmonella enterica Serovar Typhimurium ydcI Gene, Which Encodes a Conserved DNA Binding Protein Required for Full Acid Stress Resistance▿†

    PubMed Central

    Jennings, Matthew E.; Quick, Laura N.; Soni, Anjali; Davis, Richard R.; Crosby, Kathleen; Ott, C. Mark; Nickerson, Cheryl A.; Wilson, James W.

    2011-01-01

    Salmonella enterica serovar Typhimurium possesses a stimulon of genes that are differentially regulated in response to conditions of low fluid shear force that increase bacterial virulence and alter other phenotypes. In this study, we show that a previously uncharacterized member of this stimulon, ydcI or STM1625, encodes a highly conserved DNA binding protein with related homologs present in a range of Gram-negative bacterial genera. Gene expression analysis shows that ydcI is expressed in different bacterial genera and is involved in its autoregulation in S. Typhimurium. We demonstrate that purified YdcI protein specifically binds a DNA probe consisting of its own promoter sequence. We constructed an S. Typhimurium ΔydcI mutant strain and show that this strain is more sensitive to both organic and inorganic acid stress than is an isogenic WT strain, and this defect is complemented in trans. Moreover, our data indicate that ydcI is part of the rpoS regulon related to stress resistance. The S. Typhimurium ΔydcI mutant was able to invade cultured cells to the same degree as the WT strain, but a strain in which ydcI expression is induced invaded cells at a level 2.8 times higher than that of the WT. In addition, induction of ydcI expression in S. Typhimurium resulted in the formation of a biofilm in stationary-phase cultures. These data indicate the ydcI gene encodes a conserved DNA binding protein involved with aspects of prokaryotic biology related to stress resistance and possibly virulence. PMID:21398541

  8. The Bacterial iprA Gene Is Conserved across Enterobacteriaceae, Is Involved in Oxidative Stress Resistance, and Influences Gene Expression in Salmonella enterica Serovar Typhimurium

    PubMed Central

    Herman, Allison; Serfecz, Jacquelyn; Kinnally, Alexandra; Crosby, Kathleen; Youngman, Matthew; Wykoff, Dennis

    2016-01-01

    ABSTRACT The iprA gene (formerly known as yaiV or STM0374) is located in a two-gene operon in the Salmonella enterica serovar Typhimurium genome and is associated with altered expression during spaceflight and rotating-wall-vessel culture conditions that increase virulence. However, iprA is uncharacterized in the literature. In this report, we present the first targeted characterization of this gene, which revealed that iprA is highly conserved across Enterobacteriaceae. We found that S. Typhimurium, Escherichia coli, and Enterobacter cloacae ΔiprA mutant strains display a multi-log-fold increase in oxidative stress resistance that is complemented using a plasmid-borne wild-type (WT) copy of the S. Typhimurium iprA gene. This observation was also associated with increased catalase activity, increased S. Typhimurium survival in macrophages, and partial dependence on the katE gene and full dependence on the rpoS gene. Our results indicate that IprA protein activity is sensitive to deletion of the N- and C-terminal 10 amino acids, while a region that includes amino acids 56 to 80 is dispensable for activity. RNA sequencing (RNA-Seq) analysis revealed several genes altered in expression in the S. Typhimurium ΔiprA mutant strain compared to the WT, including those involved in fimbria formation, spvABCD-mediated virulence, ethanolamine utilization, the phosphotransferase system (PTS) transport, and flagellin phase switching from FlgB to FliC (likely a stochastic event) and several genes of hypothetical or putative function. IMPORTANCE Overall, this work reveals that the conserved iprA gene measurably influences bacterial biology and highlights the pool of currently uncharacterized genes that are conserved across bacterial genomes. These genes represent potentially useful targets for bacterial engineering, vaccine design, and other possible applications. PMID:27246569

  9. Extensive Genetic Variability Linked to IS26 Insertions in the fljB Promoter Region of Atypical Monophasic Variants of Salmonella enterica Serovar Typhimurium

    PubMed Central

    Boland, Cécile; Bertrand, Sophie; Mattheus, Wesley; Dierick, Katelijne; Jasson, Vicky; Rosseel, Toon; Van Borm, Steven; Mahillon, Jacques

    2015-01-01

    Fifty-nine monophasic Salmonella enterica serovar Typhimurium isolates, collected in Belgium during the period from 2008 to 2011, have been serotyped as 4,[5]:i:− and shown to harbor an fljB coding sequence. The genetic differences between these strains and phenotypically biphasic Salmonella Typhimurium were analyzed through PCR and DNA sequencing. Genetic alterations in the fljB promoter region affecting expression of the phase 2 flagellin were observed in 53 isolates. Other genetic events in the invertible region carrying the fljB promoter were observed in 2 isolates. For the remaining 4 isolates, no molecular differences with a reference biphasic Salmonella Typhimurium strain could be observed. Next-generation sequencing of one representative isolate affected in the fljB promoter region revealed a 26-kb IS26 composite transposon insertion along with a local genomic rearrangement. Several other IS26 element-mediated alterations of this genomic region were observed. This group of monophasic Salmonella Typhimurium isolates was genetically heterogeneous, as revealed by multilocus variable-number tandem-repeat analysis (MLVA), PCR, and sequencing. Pigs and pork represented a major source of such monophasic isolates in Belgium, as reported in other countries. Three out of 5 isolates of human origin presented genetic profiles identical to those of food isolates, demonstrating the pathogenic potential of the newly characterized variants and potential dissemination along the food chain. This study highlighted the key role played by IS26 insertions in the loss of phase 2 flagellin expression and the subsequent generation of multiple monophasic variant lineages from biphasic Salmonella Typhimurium ancestors. PMID:25724958

  10. Foodborne Salmonellosis in Italy: Characterization of Salmonella enterica Serovar Typhimurium and Monophasic Variant 4,[5],12:i- Isolated from Salami and Human Patients.

    PubMed

    Andreoli, Giuseppina; Merla, Cristina; Valle, Claudia Dalla; Corpus, Francesco; Morganti, Marina; D'incau, Mario; Colmegna, Silvia; Marone, Piero; Fabbi, Massimo; Barco, Lisa; Carra, Elena

    2017-04-01

    Salmonella enterica serovar Typhimurium (STm) and its monophasic variant 4,[5],12:i:- (VMSTm) have been responsible for an increased number of foodborne infections in humans in Europe in recent years. The aim of this study was to investigate the origin of three foodborne salmonellosis outbreaks that occurred in Pavia Province (Lombardy region, northern Italy) in 2010. Phenotypic and genetic characteristics of the STm and VMSTm isolates from patients and from food that were recovered in the framework of the three outbreaks were evaluated through serotyping, phage typing, antimicrobial susceptibility testing, pulsed-field gel electrophoresis (PFGE), and multiple-locus variable-number tandem repeat analysis (MLVA). Salami from three artisan producers, which had all purchased meat from the same slaughterhouse, was the food source of infection in outbreak I. STm isolates were recovered from salami and patients with symptoms of gastroenteritis. These isolates had the same PFGE type and the same rare MLVA profile (3-18-9-NA-211). The same molecular profiles were found in an STm isolate from a salami, which likely was the source of another family outbreak (II). A VMSTm strain with common phenotypic and molecular profiles was isolated from three hospitalized patients and identified as the cause of another putative outbreak (III). During the following 3 years (2011 through 2013), 360 salami produced in Pavia Province were monitored for the presence of S. enterica . In 2011, no STm and VMSTm isolates were recovered from 159 salami tested. During 2012 and 2013, 13.9% of 201 tested salami harbored S. enterica , and half of the isolates were VMSTm, mainly in salami from those artisan producers involved in the previous outbreaks. These isolates were genetically variable, especially in terms of MLVA profiles. The data collected suggest that from 2012, VMSTm has replaced STm in the environments of the salami producers monitored in this study, and these data confirm the dominance of

  11. wksl3, a New Biocontrol Agent for Salmonella enterica Serovars Enteritidis and Typhimurium in Foods: Characterization, Application, Sequence Analysis, and Oral Acute Toxicity Study

    PubMed Central

    Kang, Hyun-Wol; Kim, Jae-Won; Jung, Tae-Sung

    2013-01-01

    Of the Salmonella enterica serovars, S. Enteritidis and S. Typhimurium are responsible for most of the Salmonella outbreaks implicated in the consumption of contaminated foods in the Republic of Korea. Because of the widespread occurrence of antimicrobial-resistant Salmonella in foods and food processing environments, bacteriophages have recently surfaced as an alternative biocontrol tool. In this study, we isolated a virulent bacteriophage (wksl3) that could specifically infect S. Enteritidis, S. Typhimurium, and several additional serovars. Transmission electron microscopy revealed that phage wksl3 belongs to the family Siphoviridae. Complete genome sequence analysis and bioinformatic analysis revealed that the DNA of phage wksl3 is composed of 42,766 bp with 64 open reading frames. Since it does not encode any phage lysogeny factors, toxins, pathogen-related genes, or food-borne allergens, phage wksl3 may be considered a virulent phage with no side effects. Analysis of genetic similarities between phage wksl3 and four of its relatives (SS3e, vB_SenS-Ent1, SE2, and SETP3) allowed wksl3 to be categorized as a SETP3-like phage. A single-dose test of oral toxicity with BALB/c mice resulted in no abnormal clinical observations. Moreover, phage application to chicken skin at 8°C resulted in an about 2.5-log reduction in the number of Salmonella bacteria during the test period. The strong, stable lytic activity, the significant reduction of the number of S. Enteritidis bacteria after application to food, and the lack of clinical symptoms of this phage suggest that wksl3 may be a useful agent for the protection of foods against S. Enteritidis and S. Typhimurium contamination. PMID:23335772

  12. Distribution of Gifsy-3 and of Variants of ST64B and Gifsy-1 Prophages amongst Salmonella enterica Serovar Typhimurium Isolates: Evidence that Combinations of Prophages Promote Clonality

    PubMed Central

    Hiley, Lester; Fang, Ning-Xia; Micalizzi, Gino R.; Bates, John

    2014-01-01

    Salmonella isolates harbour a range of resident prophages which can influence their virulence and ability to compete and survive in their environment. Phage gene profiling of a range of phage types of Salmonella enterica subsp. enterica serovar Typhimurium (S. Typhimurium) indicates a significant level of correlation of phage gene profile with phage type as well as correlation with genotypes determined by a combination of multi-locus variable-number tandem repeat (VNTR) typing and clustered regularly interspaced short palindromic repeats (CRISPR) typing. Variation in phage gene profiles appears to be partly linked to differences in composition of variants of known prophages. We therefore conducted a study of the distribution of variants of ST64B and Gifsy-1 prophages and coincidently the presence of Gifsy-3 prophage in a range of S. Typhimurium phage types and genotypes. We have discovered two variants of the DT104 variant of ST64B and at least two new variants of Gifsy-1 as well as variants of related phage genes. While there is definite correlation between phage type and the prophage profile based on ST64B and Gifsy-1 variants we find stronger correlation between the VNTR/CRISPR genotype and prophage profile. Further differentiation of some genotypes is obtained by addition of the distribution of Gifsy-3 and a sequence variant of the substituted SB26 gene from the DT104 variant of ST64B. To explain the correlation between genotype and prophage profile we propose that suites of resident prophages promote clonality possibly through superinfection exclusion systems. PMID:24475087

  13. Distinct Roles of the Salmonella enterica Serovar Typhimurium CyaY and YggX Proteins in the Biosynthesis and Repair of Iron-Sulfur Clusters

    PubMed Central

    Velayudhan, Jyoti; Karlinsey, Joyce E.; Frawley, Elaine R.; Becker, Lynne A.; Nartea, Margaret

    2014-01-01

    Labile [4Fe-4S]2+ clusters found at the active sites of many dehydratases are susceptible to damage by univalent oxidants that convert the clusters to an inactive [3Fe-4S]1+ form. Bacteria repair damaged clusters in a process that does not require de novo protein synthesis or the Isc and Suf cluster assembly pathways. The current study investigates the participation of the bacterial frataxin ortholog CyaY and the YggX protein, which are proposed to play roles in iron trafficking and iron-sulfur cluster repair. Previous reports found that individual mutations in cyaY or yggX were not associated with phenotypic changes in Escherichia coli and Salmonella enterica serovar Typhimurium, suggesting that CyaY and YggX might have functionally redundant roles. However, we have found that individual mutations in cyaY or yggX confer enhanced susceptibility to hydrogen peroxide in Salmonella enterica serovar Typhimurium. In addition, inactivation of the stm3944 open reading frame, which is located immediately upstream of cyaY and which encodes a putative inner membrane protein, dramatically enhances the hydrogen peroxide sensitivity of a cyaY mutant. Overexpression of STM3944 reduces the elevated intracellular free iron levels observed in an S. Typhimurium fur mutant and also reduces the total cellular iron content under conditions of iron overload, suggesting that the stm3944-encoded protein may mediate iron efflux. Mutations in cyaY and yggX have different effects on the activities of the iron-sulfur cluster-containing aconitase, serine deaminase, and NADH dehydrogenase I enzymes of S. Typhimurium under basal conditions or following recovery from oxidative stress. In addition, cyaY and yggX mutations have additive effects on 6-phosphogluconate dehydratase-dependent growth during nitrosative stress, and a cyaY mutation reduces Salmonella virulence in mice. Collectively, these results indicate that CyaY and YggX play distinct supporting roles in iron-sulfur cluster biosynthesis

  14. Use of electron beam radiation for the reduction of Salmonella enterica serovars typhimurium and Tennessee in peanut butter.

    PubMed

    Hvizdzak, A L; Beamer, S; Jaczynski, J; Matak, K E

    2010-02-01

    Peanut butter and peanut paste products were implicated as the vehicle of contamination in an outbreak of Salmonella Typhimurium, which began in September 2008, and in the November 2006 outbreak of Salmonella Tennessee. Therefore, this study evaluated the effectiveness of electron beam (e-beam) radiation for the reduction of Salmonella serovars Tennessee (ATCC 10722) and Typhimurium (ATCC 14028) in creamy peanut butter. Each strain was studied independently. Peanut butter samples were inoculated with approximately 8.0 log CFU/g of Salmonella, and exposed to e-beam doses ranging from 0 to 3.1 kGy. Doses were confirmed with film dosimetry. Survivors were enumerated by standard spread plating on nonselective tryptic soy agar (TSA) and selective xylose-lysine-desoxycholate agar (XLD) media. Salmonella Tennessee was more susceptible to e-beam radiation, with 5.00- and 6.75-log reduction of cells on TSA and XLD, respectively, at the approximate e-beam dose of 3.0 kGy. Salmonella Typhimurium was reduced by 4.19 and 4.85 log on TSA and XLD, respectively, at the approximate e-beam dose of 3.0 kGy. D(10)-values show that Salmonella Typhimurium was more resistant (0.82 +/- 0.02 and 0.73 +/- 0.01 kGy on TSA and XLD, respectively) than was Salmonella Tennessee (0.72 +/- 0.02 and 0.60 +/- 0.01 kGy on TSA and XLD, respectively) to e-beam radiation (P < 0.05). The recovery on growth and selective media were different (P < 0.05), indicating cell injury. The results of this study demonstrate that e-beam radiation may be an effective processing step for the nonthermal inactivation of Salmonella in peanut butter.

  15. Genome Expression Analysis of Nonproliferating Intracellular Salmonella enterica Serovar Typhimurium Unravels an Acid pH-Dependent PhoP-PhoQ Response Essential for Dormancy

    PubMed Central

    Núñez-Hernández, Cristina; Tierrez, Alberto; Ortega, Álvaro D.; Pucciarelli, M. Graciela; Godoy, Marta; Eisman, Blanca; Casadesús, Josep

    2013-01-01

    Genome-wide expression analyses have provided clues on how Salmonella proliferates inside cultured macrophages and epithelial cells. However, in vivo studies show that Salmonella does not replicate massively within host cells, leaving the underlying mechanisms of such growth control largely undefined. In vitro infection models based on fibroblasts or dendritic cells reveal limited proliferation of the pathogen, but it is presently unknown whether these phenomena reflect events occurring in vivo. Fibroblasts are distinctive, since they represent a nonphagocytic cell type in which S. enterica serovar Typhimurium actively attenuates intracellular growth. Here, we show in the mouse model that S. Typhimurium restrains intracellular growth within nonphagocytic cells positioned in the intestinal lamina propria. This response requires a functional PhoP-PhoQ system and is reproduced in primary fibroblasts isolated from the mouse intestine. The fibroblast infection model was exploited to generate transcriptome data, which revealed that ∼2% (98 genes) of the S. Typhimurium genome is differentially expressed in nongrowing intracellular bacteria. Changes include metabolic reprogramming to microaerophilic conditions, induction of virulence plasmid genes, upregulation of the pathogenicity islands SPI-1 and SPI-2, and shutdown of flagella production and chemotaxis. Comparison of relative protein levels of several PhoP-PhoQ-regulated functions (PagN, PagP, and VirK) in nongrowing intracellular bacteria and extracellular bacteria exposed to diverse PhoP-PhoQ-inducing signals denoted a regulation responding to acidic pH. These data demonstrate that S. Typhimurium restrains intracellular growth in vivo and support a model in which dormant intracellular bacteria could sense vacuolar acidification to stimulate the PhoP-PhoQ system for preventing intracellular overgrowth. PMID:23090959

  16. Increased Ferroportin-1 Expression and Rapid Splenic Iron Loss Occur with Anemia Caused by Salmonella enterica Serovar Typhimurium Infection in Mice

    PubMed Central

    Nick, Heidi J.; McCoy, Melissa W.; Moreland, Sarah M.; Stepanek, Aaron M.; Benik, Ross; O'Connell, Karyn E.; Pilonieta, Maria C.; Nagy, Toni A.

    2015-01-01

    The Gram-negative intracellular bacterium Salmonella enterica serovar Typhimurium causes persistent systemic inflammatory disease in immunocompetent mice. Following oral inoculation with S. Typhimurium, mice develop a hematopathological syndrome akin to typhoid fever with splenomegaly, microcytic anemia, extramedullary erythropoiesis, and increased hemophagocytic macrophages in the bone marrow, liver, and spleen. Additionally, there is marked loss of iron from the spleen, an unanticipated result, given the iron sequestration reported in anemia of inflammatory disease. To establish why tissue iron does not accumulate, we evaluated multiple measures of pathology for 4 weeks following oral infection in mice. We demonstrate a quantitative decrease in splenic iron following infection despite increased numbers of splenic phagocytes. Infected mice have increased duodenal expression of the iron exporter ferroportin-1, consistent with increased uptake of dietary iron. Liver and splenic macrophages also express high levels of ferroportin-1. These observations indicate that splenic and hepatic macrophages export iron during S. Typhimurium infection, in contrast to macrophage iron sequestration observed in anemia of inflammatory disease. Tissue macrophage export of iron occurs concurrent with high serum concentrations of interferon gamma (IFN-γ) and interleukin 12 (IL-12). In individual mice, high concentrations of both proinflammatory (tumor necrosis factor alpha [TNF-α]) and anti-inflammatory (IL-10) cytokines in serum correlate with increased tissue bacterial loads throughout 4 weeks of infection. These in vivo observations are consistent with previous cell culture studies and suggest that the relocation of iron from tissue macrophages during infection may contribute to anemia and also to host survival of acute S. Typhimurium infection. PMID:25824831

  17. Protective Immunity Elicited by Oral Immunization of Mice with Salmonella enterica Serovar Typhimurium Braun Lipoprotein (Lpp) and Acetyltransferase (MsbB) Mutants

    PubMed Central

    Erova, Tatiana E.; Kirtley, Michelle L.; Fitts, Eric C.; Ponnusamy, Duraisamy; Baze, Wallace B.; Andersson, Jourdan A.; Cong, Yingzi; Tiner, Bethany L.; Sha, Jian; Chopra, Ashok K.

    2016-01-01

    We evaluated the extent of attenuation and immunogenicity of the ΔlppAB and ΔlppAB ΔmsbB mutants of Salmonella enterica serovar Typhimurium when delivered to mice by the oral route. These mutants were deleted either for the Braun lipoprotein genes (lppA and lppB) or in combination with the msbB gene, which encodes an acetyltransferase required for lipid A modification of lipopolysaccharide. Both the mutants were attenuated (100% animal survival) and triggered robust innate and adaptive immune responses. Comparable levels of IgG and its isotypes were produced in mice infected with wild-type (WT) S. typhimurium or its aforementioned mutant strains. The ΔlppAB ΔmsbB mutant-immunized animals resulted in the production of higher levels of fecal IgA and serum cytokines during later stages of vaccination (adaptive response). A significant production of interleukin-6 from T-cells was also noted in the ΔlppAB ΔmsbB mutant-immunized mice when compared to that of the ΔlppAB mutant. On the other hand, IL-17A production was significantly more in the serum of ΔlppAB mutant-immunized mice (innate response) with a stronger splenic T-cell proliferative and tumor-necrosis factor-α production. Based on 2-dimensional gel analysis, alterations in the levels of several proteins were observed in both the mutant strains when compared to that in WT S. typhimurium and could be associated with the higher immunogenicity of the mutants. Finally, both ΔlppAB and ΔlppAB ΔmsbB mutants provided complete protection to immunized mice against a lethal oral challenge dose of WT S. typhimurium. Thus, these mutants may serve as excellent vaccine candidates and also provide a platform for delivering heterologous antigens. PMID:27891321

  18. Inactivation of Salmonella enterica serovar Typhimurium and Escherichia coli O157:H7 in peanut butter cracker sandwiches by radio-frequency heating.

    PubMed

    Ha, Jae-Won; Kim, Sung-Youn; Ryu, Sang-Ryeol; Kang, Dong-Hyun

    2013-05-01

    A multistate outbreak of Salmonella enterica serovar Typhimurium recently occurred in the USA, which was traced back to various food products made with contaminated peanut butter. This study was conducted to investigate the efficacy of radio-frequency (RF) heating to inactivate S. Typhimurium and Escherichia coli O157:H7 in peanut butter cracker sandwiches using creamy and chunky commercial peanut butter and to determine the effect on quality by measuring color changes and sensory evaluation. Samples were treated for a maximum time of 90 s in a 27.12 MHz RF heating system. Samples were prepared in the form of peanut butter cracker sandwiches and placed in the middle of two parallel-plate electrodes. After 90 s of RF treatment, the log reductions of S. Typhimurium and E. coli O157:H7 were 4.29 and 4.39 log CFU/g, respectively, in creamy peanut butter. RF treatment of chunky peanut butter for 90 s also significantly (P < 0.05) reduced levels of S. Typhimurium and E. coli O157:H7 by 4.55 log CFU/g and 5.32 log CFU/g. Color values and sensory characteristics of the RF treated peanut butter and crackers were not significantly (P > 0.05) different from the control. These results suggest that RF heating can be applied to control pathogens in peanut butter products without affecting quality.

  19. Protective Immunity Elicited by Oral Immunization of Mice with Salmonella enterica Serovar Typhimurium Braun Lipoprotein (Lpp) and Acetyltransferase (MsbB) Mutants.

    PubMed

    Erova, Tatiana E; Kirtley, Michelle L; Fitts, Eric C; Ponnusamy, Duraisamy; Baze, Wallace B; Andersson, Jourdan A; Cong, Yingzi; Tiner, Bethany L; Sha, Jian; Chopra, Ashok K

    2016-01-01

    We evaluated the extent of attenuation and immunogenicity of the ΔlppAB and ΔlppAB ΔmsbB mutants of Salmonella enterica serovar Typhimurium when delivered to mice by the oral route. These mutants were deleted either for the Braun lipoprotein genes (lppA and lppB) or in combination with the msbB gene, which encodes an acetyltransferase required for lipid A modification of lipopolysaccharide. Both the mutants were attenuated (100% animal survival) and triggered robust innate and adaptive immune responses. Comparable levels of IgG and its isotypes were produced in mice infected with wild-type (WT) S. typhimurium or its aforementioned mutant strains. The ΔlppAB ΔmsbB mutant-immunized animals resulted in the production of higher levels of fecal IgA and serum cytokines during later stages of vaccination (adaptive response). A significant production of interleukin-6 from T-cells was also noted in the ΔlppAB ΔmsbB mutant-immunized mice when compared to that of the ΔlppAB mutant. On the other hand, IL-17A production was significantly more in the serum of ΔlppAB mutant-immunized mice (innate response) with a stronger splenic T-cell proliferative and tumor-necrosis factor-α production. Based on 2-dimensional gel analysis, alterations in the levels of several proteins were observed in both the mutant strains when compared to that in WT S. typhimurium and could be associated with the higher immunogenicity of the mutants. Finally, both ΔlppAB and ΔlppAB ΔmsbB mutants provided complete protection to immunized mice against a lethal oral challenge dose of WT S. typhimurium. Thus, these mutants may serve as excellent vaccine candidates and also provide a platform for delivering heterologous antigens.

  20. Zinc Supplementation, via GPR39, Upregulates PKCζ to Protect Intestinal Barrier Integrity in Caco-2 Cells Challenged by Salmonella enterica Serovar Typhimurium.

    PubMed

    Shao, Yu-Xin; Lei, Zhao; Wolf, Patricia G; Gao, Yan; Guo, Yu-Ming; Zhang, Bing-Kun

    2017-07-01

    Background: Zinc has been shown to improve intestinal barrier function against Salmonella enterica serovar Typhimurium (S. typhimurium) infection, but the mechanisms involved in this process remain undefined.Objective: We aimed to explore the roles of G protein-coupled receptor (GPR)39 and protein kinase Cζ (PKCζ) in the regulation by zinc of intestinal barrier function.Methods: A Transwell Caco-2 monolayer was pretreated with 0, 50, or 100 μM Zn and then incubated with S. typhimurium for 0-6 h. Afterward, cells silenced by the small interfering RNA for GPR39 or PKCζ were pretreated with 100 μM Zn and incubated with S. typhimurium for 3 h. Finally, transepithelial electrical resistance (TEER), permeability, tight junction (TJ) proteins, and signaling molecules GPR39 and PKCζ were measured.Results: Compared with controls, S. typhimurium decreased TEER by 62.3-96.2% at 4-6 h (P < 0.001), increased (P < 0.001) permeability at 6 h, and downregulated (P < 0.05) TJ protein zonula occludens (ZO)-1 and occludin by 104-123%, as well as Toll-like receptor 2 and PKCζ by 35.1% and 75.2%, respectively. Compared with S. typhimurium-challenged cells, 50 and 100 μM Zn improved TEER by 26.3-60.9% at 4-6 h (P < 0.001) and decreased (P < 0.001) permeability and bacterial invasion at 6 h. A total of 100 μM Zn increased ZO-1, occludin, GPR39, and PKCζ 0.72- to 1.34-fold (P < 0.05); however, 50 μM Zn did not affect ZO-1 or occludin (P > 0.1). Silencing GPR39 decreased (P < 0.05) zinc-activated PKCζ and blocked (P < 0.05) the promotion of zinc on epithelial integrity. Furthermore, silencing PKCζ counteracted the protective effect of zinc on epithelial integrity but did not inhibit GPR39 (P = 0.138).Conclusion: We demonstrated that zinc upregulates PKCζ by activating GPR39 to enhance the abundance of ZO-1, thereby improving epithelial integrity in S. typhimurium-infected Caco-2 cells. © 2017 American Society for Nutrition.

  1. SadA, a Trimeric Autotransporter from Salmonella enterica Serovar Typhimurium, Can Promote Biofilm Formation and Provides Limited Protection against Infection ▿ †

    PubMed Central

    Raghunathan, Dhaarini; Wells, Timothy J.; Morris, Faye C.; Shaw, Robert K.; Bobat, Saeeda; Peters, Sarah E.; Paterson, Gavin K.; Jensen, Karina Tveen; Leyton, Denisse L.; Blair, Jessica M. A.; Browning, Douglas F.; Pravin, John; Flores-Langarica, Adriana; Hitchcock, Jessica R.; Moraes, Claudia T. P.; Piazza, Roxane M. F.; Maskell, Duncan J.; Webber, Mark A.; May, Robin C.; MacLennan, Calman A.; Piddock, Laura J.; Cunningham, Adam F.; Henderson, Ian R.

    2011-01-01

    Salmonella enterica is a major cause of morbidity worldwide and mortality in children and immunocompromised individuals in sub-Saharan Africa. Outer membrane proteins of Salmonella are of significance because they are at the interface between the pathogen and the host, they can contribute to adherence, colonization, and virulence, and they are frequently targets of antibody-mediated immunity. In this study, the properties of SadA, a purported trimeric autotransporter adhesin of Salmonella enterica serovar Typhimurium, were examined. We demonstrated that SadA is exposed on the Salmonella cell surface in vitro and in vivo during infection of mice. Expression of SadA resulted in cell aggregation, biofilm formation, and increased adhesion to human intestinal Caco-2 epithelial cells. Immunization of mice with folded, full-length, purified SadA elicited an IgG response which provided limited protection against bacterial challenge. When anti-SadA IgG titers were enhanced by administering alum-precipitated protein, a modest additional protection was afforded. Therefore, despite SadA having pleiotropic functions, it is not a dominant, protective antigen for antibody-mediated protection against Salmonella. PMID:21859856

  2. Metabolic Engineering of a Novel Propionate-Independent Pathway for the Production of Poly(3-Hydroxybutyrate-co-3-Hydroxyvalerate) in Recombinant Salmonella enterica Serovar Typhimurium

    PubMed Central

    Aldor, Ilana S.; Kim, Seon-Won; Jones Prather, Kristala L.; Keasling, Jay D.

    2002-01-01

    A pathway was metabolically engineered to produce poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV), a biodegradable thermoplastic with proven commercial applications, from a single, unrelated carbon source. An expression system was developed in which a prpC strain of Salmonella enterica serovar Typhimurium, with a mutation in the ability to metabolize propionyl coenzyme A (propionyl-CoA), served as the host for a plasmid harboring the Acinetobacter polyhydroxyalkanoate synthesis operon (phaBCA) and a second plasmid with the Escherichia coli sbm and ygfG genes under an independent promoter. The sbm and ygfG genes encode a novel (2R)-methylmalonyl-CoA mutase and a (2R)-methylmalonyl-CoA decarboxylase, respectively, which convert succinyl-CoA, derived from the tricarboxylic acid cycle, to propionyl-CoA, an essential precursor of 3-hydroxyvalerate (HV). The S. enterica system accumulated PHBV with significant HV incorporation when the organism was grown aerobically with glycerol as the sole carbon source. It was possible to vary the average HV fraction in the copolymer by adjusting the arabinose or cyanocobalamin (precursor of coenzyme B12) concentration in the medium. PMID:12147480

  3. Functional insight from the tetratricopeptide repeat-like motifs of the type III secretion chaperone SicA in Salmonella enterica serovar Typhimurium.

    PubMed

    Kim, Jin Seok; Kim, Bae-Hoon; Jang, Jung Im; Eom, Jeong Seon; Kim, Hyeon Guk; Bang, Iel Soo; Park, Yong Keun

    2014-01-01

    SicA functions both as a class II chaperone for SipB and SipC of the type III secretion system (T3SS)-1 and as a transcriptional cofactor for the AraC-type transcription factor InvF in Salmonella enterica subsp. enterica serovar Typhimurium. Bioinformatic analysis has predicted that SicA possesses three tetratricopeptide repeat (TPR)-like motifs, which are important for protein-protein interactions and serve as multiprotein complex mediators. To investigate whether the TPR-like motifs in SicA are critical for its transcriptional cofactor function, the canonical residues in these motifs were mutated to glutamate (SicAA44E , SicAA78E , and SicAG112E ). None of these mutants except SicAA44E were able to activate the expression of the sipB and sigD genes. SicAA44E still has a capacity to interact with InvF in vitro, and despite its instability in cell, it could activate the sigDE operon. This suggests that TPR motifs are important for the transcriptional cofactor function of the SicA chaperone.

  4. Enhancement of Th1-biased protective immunity against avian influenza H9N2 virus via oral co-administration of attenuated Salmonella enterica serovar Typhimurium expressing chicken interferon-α and interleukin-18 along with an inactivated vaccine

    PubMed Central

    2012-01-01

    Background Control of currently circulating re-assorted low-pathogenicity avian influenza (LPAI) H9N2 is a major concern for both animal and human health. Thus, an improved LPAI H9N2 vaccination strategy is needed to induce complete immunity in chickens against LPAI H9N2 virus strains. Cytokines play a crucial role in mounting both the type and extent of an immune response generated following infection with a pathogen or after vaccination. To improve the efficacy of inactivated LPAI H9N2 vaccine, attenuated Salmonella enterica serovar Typhimurium was used for oral co-administration of chicken interferon-α (chIFN-α) and chicken interleukin-18 (chIL-18) as natural immunomodulators. Results Oral co-administration of S. enterica serovar Typhimurium expressing chIFN-α and chIL-18, prior to vaccination with inactivated AI H9N2 vaccine, modulated the immune response of chickens against the vaccine antigen through enhanced humoral and Th1-biased cell-mediated immunity, compared to chickens that received single administration of S. enterica serovar Typhimurium expressing either chIFN-α or chIL-18. To further test the protective efficacy of this improved vaccination regimen, immunized chickens were intra-tracheally challenged with a high dose of LPAI H9N2 virus. Combined administration of S. enterica serovar Typhimurium expressing chIFN-α and chIL-18 showed markedly enhanced protection compared to single administration of the construct, as determined by mortality, clinical severity, and feed and water intake. This enhancement of protective immunity was further confirmed by reduced rectal shedding and replication of AIV H9N2 in different tissues of challenged chickens. Conclusions Our results indicate the value of combined administration of chIFN-α and chIL-18 using a Salmonella vaccine strain to generate an effective immunization strategy in chickens against LPAI H9N2. PMID:22776696

  5. Oral administration of live attenuated Salmonella enterica serovar Typhimurium expressing chicken interferon-α alleviates clinical signs caused by respiratory infection with avian influenza virus H9N2.

    PubMed

    Rahman, Md Masudur; Uyangaa, Erdenebileg; Han, Young Woo; Kim, Seong Bum; Kim, Jin Hyoung; Choi, Jin Young; Yoo, Dong Jin; Hong, Jin Tae; Han, Sang-Bae; Kim, Bumseok; Kim, Koanhoi; Eo, Seong Kug

    2011-12-29

    Low pathogenic avian influenza (LPAI) H9N2 has attracted considerable attention due to severe commercial losses in the poultry industry. Furthermore, avian influenza virus (AIV) H9N2-infected chickens can be a reservoir for viral transmission to mammals including pigs and humans, complicating control of viral mutants. Chicken interferon-alpha (chIFN-α) may be useful as an exogenous antiviral agent to control AIV H9N2 infection. However, a superior vehicle for administration of chIFN-α is needed because of challenges of protein stability, production cost, and labor associated with mass administration. Presently, oral administration of single dose of attenuated Salmonella enterica serovar Typhimurium expressing chIFN-α alleviated clinical signs and histopathological changes caused by respiratory infection with AIV H9N2 and reduced the excretion of virus in cloacal swab samples. Similarly, chickens administered S. enterica serovar Typhimurium expressing chIFN-α showed inhibited replication of AIV H9N2 in several different tissues including trachea, lung, cecal tonsil, and brain. Furthermore, immune responses specific for challenged AIV H9N2 were enhanced in chickens administered S. enterica serovar Typhimurium expressing chIFN-α, as determined by hemagglutination inhibition assay of sera, proliferation and IFN-γ and interleukin-4 expression by AIV H9N2 antigen-stimulated peripheral blood mononuclear cells and splenocytes. Therefore, oral administration of S. enterica serovar Typhimurium expressing chIFN-α can successfully control clinical signs caused by respiratory infection with AIV H9N2, which provides valuable insight into the use of attenuated Salmonella vaccine as an oral delivery system of chIFN-α to prevent the replication of AIV H9N2 in respiratory tract. Copyright © 2011 Elsevier B.V. All rights reserved.

  6. Evaluation of near-infrared pasteurization in controlling Escherichia coli O157:H7, Salmonella enterica serovar typhimurium, and Listeria monocytogenes in ready-to-eat sliced ham.

    PubMed

    Ha, Jae-Won; Ryu, Sang-Ryeol; Kang, Dong-Hyun

    2012-09-01

    This study was conducted to investigate the efficacy of near-infrared (NIR) heating to reduce Salmonella enterica serovar Typhimurium, Escherichia coli O157:H7, and Listeria monocytogenes in ready-to-eat (RTE) sliced ham compared to conventional convective heating, and the effect of NIR heating on quality was determined by measuring the color and texture change. A cocktail of three pathogens was inoculated on the exposed or protected surfaces of ham slices, followed by NIR or conventional heating at 1.8 kW. NIR heating for 50 s achieved 4.1-, 4.19-, and 3.38-log reductions in surface-inoculated S. Typhimurium, E. coli O157:H7, and L. monocytogenes, respectively, whereas convective heating needed 180 s to attain comparable reductions for each pathogen. There were no statistically significant (P > 0.05) differences in reduction between surface- and internally inoculated pathogens at the end of NIR treatment (50 s). However, when treated with conventional convective heating, significant (P < 0.05) differences were observed at the final stages of the treatment (150 and 180 s). Color values and texture parameters of NIR-treated (50-s treatment) ham slices were not significantly (P > 0.05) different from those of nontreated samples. These results suggest that NIR heating can be applied to control internalized pathogens as well as surface-adhering pathogens in RTE sliced meats without affecting product quality.

  7. Evaluation of Near-Infrared Pasteurization in Controlling Escherichia coli O157:H7, Salmonella enterica Serovar Typhimurium, and Listeria monocytogenes in Ready-To-Eat Sliced Ham

    PubMed Central

    Ha, Jae-Won; Ryu, Sang-Ryeol

    2012-01-01

    This study was conducted to investigate the efficacy of near-infrared (NIR) heating to reduce Salmonella enterica serovar Typhimurium, Escherichia coli O157:H7, and Listeria monocytogenes in ready-to-eat (RTE) sliced ham compared to conventional convective heating, and the effect of NIR heating on quality was determined by measuring the color and texture change. A cocktail of three pathogens was inoculated on the exposed or protected surfaces of ham slices, followed by NIR or conventional heating at 1.8 kW. NIR heating for 50 s achieved 4.1-, 4.19-, and 3.38-log reductions in surface-inoculated S. Typhimurium, E. coli O157:H7, and L. monocytogenes, respectively, whereas convective heating needed 180 s to attain comparable reductions for each pathogen. There were no statistically significant (P > 0.05) differences in reduction between surface- and internally inoculated pathogens at the end of NIR treatment (50 s). However, when treated with conventional convective heating, significant (P < 0.05) differences were observed at the final stages of the treatment (150 and 180 s). Color values and texture parameters of NIR-treated (50-s treatment) ham slices were not significantly (P > 0.05) different from those of nontreated samples. These results suggest that NIR heating can be applied to control internalized pathogens as well as surface-adhering pathogens in RTE sliced meats without affecting product quality. PMID:22773635

  8. Overexpression of the Multidrug Efflux Operon acrEF by Insertional Activation with IS1 or IS10 Elements in Salmonella enterica Serovar Typhimurium DT204 acrB Mutants Selected with Fluoroquinolones

    PubMed Central

    Olliver, Anne; Vallé, Michel; Chaslus-Dancla, Elisabeth; Cloeckaert, Axel

    2005-01-01

    High-level fluoroquinolone (FQ) resistance in Salmonella enterica serovar Typhimurium phage type DT204 has been previously shown to be essentially due to both multiple target gene mutations and active efflux by the AcrAB-TolC efflux system. In this study we show that in intermediatly resistant acrB-inactivated serovar Typhimurium DT204 mutants, high-level resistance to FQs can be restored on in vitro selection with FQs. In each FQ- resistant mutant selected from serovar Typhimurium DT204 acrB mutant strains, an insertion sequence (IS1 or IS10) was found integrated upstream of the acrEF operon, coding for AcrEF, an efflux pump highly homologous to AcrAB. In one of the strains, transposition of IS1 caused partial deletion of acrS, the putative local repressor gene of the acrEF operon. Sequence analysis showed that both IS1 and IS10 elements contain putative promoter sequences that might alter the expression of adjacent acrEF genes. Indeed, reverse transcription-PCR experiments showed an 8- to 10-fold increase in expression of acrF in these insertional mutants, relative to their respective parental strain, which correlated well with the resistance levels observed to FQs and other unrelated drugs. It is noteworthy that AcrEF did not contribute to the intrinsic drug resistance of serovar Typhimurium, since acrF deletion in wild-type strains did not result in any increase in drug susceptibility. Moreover, deletion of acrS did not cause any acrF overexpression or any decrease in drug susceptibility, suggesting that acrEF overexpression is mediated solely by the IS1 and IS10 promoter sequences and not by inactivity of AcrS. Southern blot experiments showed that the number of chromosomal IS1 and IS10 elements in the serovar Typhimurium DT204 genome was about 5 and 15 respectively. None were detected in epidemic serovar Typhimurium DT104 strains or in the serovar Typhimurium reference strain LT2. Carrying IS1 and/or IS10 elements in their chromosome may thus be a selective

  9. Proteomic Analysis of Salmonella enterica Serovar Typhimurium Isolated from RAW 264.7 Macrophages: identification of a novel protein that contributes to the replication of serovar Typhimurium inside macrophages

    SciTech Connect

    Shi, Liang; Adkins, Joshua N.; Coleman, James R.; Schepmoes, Athena A.; Dohnalkova, Alice; Mottaz, Heather M.; Norbeck, Angela D.; Purvine, Samuel O.; Manes, Nathan P.; Smallwood, Heather S.; Wang, Haixing H.; Forbes, John; Gros, Philippe; Uzzau, Sergio; Rodland, Karin D.; Heffron, Fred; Smith, Richard D.; Squier, Thomas C.

    2006-09-01

    ABSTRACT: To evade host resistance mechanisms, Salmonella enterica serovar Typhimurium (STM), a facultative intracellular pathogen, must alter its proteome following macrophage infection. To identify new colonization and virulence factors that mediate STM pathogenesis, we have isolated STM cells from RAW 264.7 macrophages at various time-points following infection and used a liquid chromatography-mass spectrometry (LC-MS)-based proteomic approach to detect the changes in STM protein abundances. Because host resistance to STM infection is strongly modulated by the expression of a functional host resistant regulator, i.e., natural resistance associated macrophage protein 1 (Nramp1, also called Slc11a1), we have also examined the effects of Nramp1 activity on the changes of STM protein abundances. A total of 315 STM proteins have been identified from isolated STM cells, which are largely house-keeping proteins whose abundances remain relatively constant during the time-course of infection. However, 39 STM proteins are strongly induced after infection, suggesting their involvement in modulating colonization and infection. Of the 39 induced proteins, 6 proteins are specifically modulated by Nramp1 activity, including STM3117, as well as STM3118-3119 whose time-dependent abundance changes were confirmed using Western blot analysis. Deletion of the gene encoding STM3117 resulted in a dramatic reduction in the ability of STM to colonize wild-type RAW 264.7 macrophages, demonstrating a critical involvement of STM3117 in promoting the replication of STM inside macrophages. The predicted function common for STM3117-3119 is biosynthesis and modification of the peptidoglycan layer of STM cell wall, emphasizing their important roles in the colonization of macrophages by Salmonella.

  10. Percolation and Survival of Escherichia coli O157:H7 and Salmonella enterica Serovar Typhimurium in Soil Amended with Contaminated Dairy Manure or Slurry▿

    PubMed Central

    Semenov, Alexander V.; van Overbeek, Leo; van Bruggen, Ariena H. C.

    2009-01-01

    The effect of cattle manure and slurry application on percolation and survival of Escherichia coli O157:H7 and Salmonella enterica serovar Typhimurium was investigated for different soil depths after the addition of water. Four treatments were chosen for the first set of experiments: (i) addition of inoculated farmyard manure on the soil surface, (ii) mixing of inoculated farmyard manure with the top 10 cm of soil, (iii) addition of inoculated slurry on the soil surface, and (iv) injection of inoculated slurry into the top 10 cm of the soil. Homogeneity of water distribution in the soil profile was confirmed by a nondestructive nuclear magnetic resonance method. Survival data were fitted to a modified logistic model, and estimated survival times were compared. In the second set of experiments, pathogen-inoculated farmyard manure or slurry was applied to soil columns with 1-month-old lettuce plants. More pathogen cells percolated to greater depths after slurry than after manure application. Survival of E. coli O157:H7 was significantly longer in soil with slurry than in that with manure, while survival of Salmonella serovar Typhimurium was equally high with manure and slurry. The densities of the pathogens were not different in the rhizosphere compared to the bulk soil with manure, while the densities were higher by 0.88 ± 0.11 and 0.71 ± 0.23 log CFU per g (dry weight), respectively, in the rhizosphere than in bulk soil after slurry application. Our results suggest that surface application of manure may decrease the risk of contamination of groundwater and lettuce roots compared to injection of slurry. PMID:19270130

  11. Percolation and survival of Escherichia coli O157:H7 and Salmonella enterica serovar Typhimurium in soil amended with contaminated dairy manure or slurry.

    PubMed

    Semenov, Alexander V; van Overbeek, Leo; van Bruggen, Ariena H C

    2009-05-01

    The effect of cattle manure and slurry application on percolation and survival of Escherichia coli O157:H7 and Salmonella enterica serovar Typhimurium was investigated for different soil depths after the addition of water. Four treatments were chosen for the first set of experiments: (i) addition of inoculated farmyard manure on the soil surface, (ii) mixing of inoculated farmyard manure with the top 10 cm of soil, (iii) addition of inoculated slurry on the soil surface, and (iv) injection of inoculated slurry into the top 10 cm of the soil. Homogeneity of water distribution in the soil profile was confirmed by a nondestructive nuclear magnetic resonance method. Survival data were fitted to a modified logistic model, and estimated survival times were compared. In the second set of experiments, pathogen-inoculated farmyard manure or slurry was applied to soil columns with 1-month-old lettuce plants. More pathogen cells percolated to greater depths after slurry than after manure application. Survival of E. coli O157:H7 was significantly longer in soil with slurry than in that with manure, while survival of Salmonella serovar Typhimurium was equally high with manure and slurry. The densities of the pathogens were not different in the rhizosphere compared to the bulk soil with manure, while the densities were higher by 0.88 +/- 0.11 and 0.71 +/- 0.23 log CFU per g (dry weight), respectively, in the rhizosphere than in bulk soil after slurry application. Our results suggest that surface application of manure may decrease the risk of contamination of groundwater and lettuce roots compared to injection of slurry.

  12. Chloramphenicol and tetracycline decrease motility and increase invasion and attachment gene expression in specific isolates of multidrug-resistant Salmonella enterica serovar Typhimurium

    PubMed Central

    Brunelle, Brian W.; Bearson, Bradley L.; Bearson, Shawn M. D.

    2015-01-01

    Salmonella enterica serovar Typhimurium is one of the most common serovars isolated from humans and livestock, and over 35% of these isolates are resistant to three or more antibiotics. Multidrug-resistant (MDR) Salmonella is a public health concern as it is associated with increased morbidity in patients compared to antibiotic sensitive strains, though it is unknown how the antibiotic resistant isolates lead to a more severe infection. Cellular invasion is temporally regulated in Salmonella and normally occurs during late-log and stationary growth. However, our previous work determined that a 30 min exposure to a sub-inhibitory concentration of tetracycline can induce the full invasion phenotype during early-log growth in certain MDR S. Typhimurium isolates. The current study examined whether sub-inhibitory concentrations of other antibiotics could also induce the invasiveness in the same set of isolates. Ampicillin and streptomycin had no effect on invasion, but certain concentrations of chloramphenicol were found to induce invasion in a subset of isolates. Two of the isolates induced by chloramphenicol were also inducible by tetracycline. RNA-seq analyses demonstrated that chloramphenicol and tetracycline both down-regulated motility gene expression, while up-regulating genes associated with attachment, invasion, and intracellular survival. Eleven fimbrial operons were up-regulated, which is notable as only three fimbrial operons were thought to be inducible in culture; six of these up-regulated operons have been reported to play a role in Salmonella persistence in mice. Overall, these data show that the normal progression of the genetic pathways that regulate invasion can be expedited to occur within 30 min due to antibiotic exposure. This altered invasion process due to antibiotics may play a role in the increased intensity and duration of infection observed in patients with MDR Salmonella. PMID:25688233

  13. Pediatric Epidemic of Salmonella enterica Serovar Typhimurium in the Area of L’Aquila, Italy, Four Years after a Catastrophic Earthquake

    PubMed Central

    Nigro, Giovanni; Bottone, Gabriella; Maiorani, Daniela; Trombatore, Fabiana; Falasca, Silvana; Bruno, Gianfranco

    2016-01-01

    Background: A Salmonella enterica epidemic occurred in children of the area of L’Aquila (Central Italy, Abruzzo region) between June 2013 and October 2014, four years after the catastrophic earthquake of 6 April 2009. Methods: Clinical and laboratory data were collected from hospitalized and ambulatory children. Routine investigations for Salmonella infection were carried out on numerous alimentary matrices of animal origin and sampling sources for drinking water of the L’Aquila district, including pickup points of the two main aqueducts. Results: Salmonella infection occurred in 155 children (83 females: 53%), aged 1 to 15 years (mean 2.10). Of these, 44 children (28.4%) were hospitalized because of severe dehydration, electrolyte abnormalities, and fever resistant to oral antipyretic and antibiotic drugs. Three children (1.9%) were reinfected within four months after primary infection by the same Salmonella strain. Four children (2.6%), aged one to two years, were coinfected by rotavirus. A seven-year old child had a concomitant right hip joint arthritis. The isolated strains, as confirmed in about the half of cases or probable/possible in the remaining ones, were identified as S. enterica serovar Typhimurium [4,5:i:-], monophasic variant. Aterno river, bordering the L’Aquila district, was recognized as the main responsible source for the contamination of local crops and vegetables derived from polluted crops. Conclusions: The high rate of hospitalized children underlines the emergence of a highly pathogenic S. enterica strain probably subsequent to the contamination of the spring water sources after geological changes occurred during the catastrophic earthquake. PMID:27164121

  14. The In Vitro Redundant Enzymes PurN and PurT Are Both Essential for Systemic Infection of Mice in Salmonella enterica Serovar Typhimurium

    PubMed Central

    Jelsbak, Lotte; Mortensen, Mie I. B.; Kilstrup, Mogens

    2016-01-01

    Metabolic enzymes show a high degree of redundancy, and for that reason they are generally ignored in searches for novel targets for anti-infective substances. The enzymes PurN and PurT are redundant in vitro in Salmonella enterica serovar Typhimurium, in which they perform the third step of purine synthesis. Surprisingly, the results of the current study demonstrated that single-gene deletions of each of the genes encoding these enzymes caused attenuation (competitive infection indexes [CI] of <0.03) in mouse infections. While the ΔpurT mutant multiplied as fast as the wild-type strain in cultured J774A.1 macrophages, net multiplication of the ΔpurN mutant was reduced approximately 50% in 20 h. The attenuation of the ΔpurT mutant was abolished by simultaneous removal of the enzyme PurU, responsible for the formation of formate, indicating that the attenuation was related to formate accumulation or wasteful consumption of formyl tetrahydrofolate by PurU. In the process of further characterization, we disclosed that the glycine cleavage system (GCV) was the most important for formation of C1 units in vivo (CI = 0.03 ± 0.03). In contrast, GlyA was the only important enzyme for the formation of C1 units in vitro. The results with the ΔgcvT mutant further revealed that formation of serine by SerA and further conversion of serine into C1 units and glycine by GlyA were not sufficient to ensure C1 formation in S. Typhimurium in vivo. The results of the present study call for reinvestigations of the concept of metabolic redundancy in S. Typhimurium in vivo. PMID:27113361

  15. The In Vitro Redundant Enzymes PurN and PurT Are Both Essential for Systemic Infection of Mice in Salmonella enterica Serovar Typhimurium.

    PubMed

    Jelsbak, Lotte; Mortensen, Mie I B; Kilstrup, Mogens; Olsen, John E

    2016-07-01

    Metabolic enzymes show a high degree of redundancy, and for that reason they are generally ignored in searches for novel targets for anti-infective substances. The enzymes PurN and PurT are redundant in vitro in Salmonella enterica serovar Typhimurium, in which they perform the third step of purine synthesis. Surprisingly, the results of the current study demonstrated that single-gene deletions of each of the genes encoding these enzymes caused attenuation (competitive infection indexes [CI] of <0.03) in mouse infections. While the ΔpurT mutant multiplied as fast as the wild-type strain in cultured J774A.1 macrophages, net multiplication of the ΔpurN mutant was reduced approximately 50% in 20 h. The attenuation of the ΔpurT mutant was abolished by simultaneous removal of the enzyme PurU, responsible for the formation of formate, indicating that the attenuation was related to formate accumulation or wasteful consumption of formyl tetrahydrofolate by PurU. In the process of further characterization, we disclosed that the glycine cleavage system (GCV) was the most important for formation of C1 units in vivo (CI = 0.03 ± 0.03). In contrast, GlyA was the only important enzyme for the formation of C1 units in vitro The results with the ΔgcvT mutant further revealed that formation of serine by SerA and further conversion of serine into C1 units and glycine by GlyA were not sufficient to ensure C1 formation in S Typhimurium in vivo The results of the present study call for reinvestigations of the concept of metabolic redundancy in S Typhimurium in vivo.

  16. Compensatory role of PspA, a member of the phage shock protein operon, in rpoE mutant Salmonella enterica serovar Typhimurium.

    PubMed

    Becker, Lynne A; Bang, Iel-Soo; Crouch, Marie-Laure; Fang, Ferric C

    2005-05-01

    Sigma(E) is an alternative sigma factor that responds to and ameliorates extracytoplasmic stress. In Salmonella enterica serovar Typhimurium (S. Typhimurium), sigma(E) is required for oxidative stress resistance, stationary-phase survival and virulence in mice. Microarray analysis of stationary-phase gene expression in rpoE mutant bacteria revealed a dramatic increase in expression of pspA, a member of the phage shock protein (psp) operon. The psp operon can be induced by filamentous bacteriophages or by perturbations of protein secretion, and is believed to facilitate the maintenance of proton motive force (PMF). We hypothesized that increased pspA expression may represent a compensatory response to the loss of sigma(E) function. Increased pspA expression was confirmed in rpoE mutant Salmonella and also observed in a mutant lacking the F(1)F(0) ATPase. Alternatively, expression of pspA could be induced by exposure to CCCP, a protonophore that disrupts PMF. An rpoE pspA double mutant strain was found to have a stationary-phase survival defect more pronounced than that of isogenic strains harbouring single mutations. The double mutant strains were also more susceptible to killing by CCCP or by a bactericidal/permeability-increasing protein (BPI)-derived anti-microbial peptide. Using fluorescence ratio imaging, differences were observed in the Deltapsi of wild-type and rpoE or pspA mutant bacteria. These findings suggest that pspA expression in S. Typhimurium is induced by alterations in PMF and a functional sigma(E) regulon is essential for the maintenance of PMF.

  17. Molecular epidemiology of Salmonella enterica serovar typhimurium isolates from cattle in hokkaido, Japan: evidence of clonal replacement and characterization of the disseminated clone.

    PubMed

    Tamamura, Yukino; Uchida, Ikuo; Tanaka, Kiyoshi; Okazaki, Hizuru; Tezuka, Satoru; Hanyu, Hideki; Kataoka, Natsumi; Makino, Sou-Ichi; Kishima, Masato; Kubota, Takayuki; Kanno, Toru; Hatama, Shinichi; Ishihara, Ryoko; Hata, Eiji; Yamada, Hironari; Nakaoka, Yuuji; Akiba, Masato

    2011-03-01

    The molecular epidemiology of 545 Salmonella enterica serovar Typhimurium isolates collected between 1977 and 2009 from cattle in Hokkaido, Japan, was investigated using pulsed-field gel electrophoresis (PFGE). Nine main clusters were identified from 116 PFGE patterns. Cluster I comprised 248 isolates, 243 of which possessed a sequence specific to definitive phage type 104 (DT104) or U302. The cluster I isolates were dominant in 1993 to 2003, but their numbers declined beginning in 2004. Beginning in 2002, an increase was observed in the number of cluster VII isolates, consisting of 21 PFGE patterns comprising 165 isolates. A total of 116 isolates representative of the 116 PFGE profiles were analyzed by multilocus variable-number tandem-repeat analysis (MLVA). Other than two drug-sensitive isolates, 19 isolates within cluster VII were classified in the same cluster by MLVA. Among the cluster VII isolates, an antibiotic resistance type showing resistance to ampicillin, chloramphenicol, streptomycin, sulfonamides, tetracycline, kanamycin, cefazolin, and sulfamethoxazole-trimethoprim and a resistance type showing resistance to ampicillin, streptomycin, sulfonamides, tetracycline, and kanamycin were found in 23 and 125 isolates, respectively. In the 19 isolates representative of cluster VII, the bla(TEM-1) gene was found on a Salmonella serotype Typhimurium virulence plasmid, which was transferred to Escherichia coli by electroporation along with resistance to two to four other antimicrobials. Genomic analysis by subtractive hybridization and plasmid analysis suggested that the bla(TEM-1)-carrying virulence plasmid has a mosaic structure composed of elements of different origin. These results indicate an emerging multidrug-resistant S. Typhimurium clone carrying a virulence-resistance plasmid among cattle in Hokkaido, Japan.

  18. Oral immunization with attenuated Salmonella enterica serovar Typhimurium encoding Cryptosporidium parvum Cp23 and Cp40 antigens induces a specific immune response in mice.

    PubMed

    Benitez, Alvaro J; McNair, Nina; Mead, Jan R

    2009-09-01

    Attenuated Salmonella enterica serovar Typhimurium vaccine strain SL3261 was used as an antigen delivery system for the oral immunization of mice against two Cryptosporidium parvum antigens, Cp23 and Cp40. Each antigen was subcloned into the pTECH1 vector system, which allows them to be expressed as fusion proteins with highly immunogenic fragment C of tetanus toxin under the control of the anaerobically inducible nirB promoter. The recombinant vector was introduced into Salmonella Typhimurium vaccine strain SL3261, and the stable soluble expression of the chimeric protein was evaluated and confirmed by Western blotting with polyclonal C. parvum antisera. Mice were inoculated orally with a single dose of SL3261/pTECH-Cp23 or Cp40, respectively, and plasmid stability was demonstrated both in vitro and in vivo. Specific serum immunoglobulin G (IgG) antibodies against the Cp23 or Cp40 antigen were detected by enzyme-linked immunosorbent assay 35 days after immunization. Also, serum IgA and mucosal (feces) IgA antibodies were detected in 30% of the mice immunized with Cp23. In addition, prime-boosting with Cp23 and Cp40 DNA vaccine vectors followed by Salmonella immunization significantly increased antibody responses to both antigens. Our data show that a single oral inoculation with recombinant S. Typhimurium SL3261 can induce specific antibody responses to the Cp23 or Cp40 antigen from C. parvum in mice, suggesting that recombinant Salmonella is a feasible delivery system for a vaccine against C. parvum infection.

  19. Mixed biofilm formation by Shiga toxin-producing Escherichia coli and Salmonella enterica serovar Typhimurium enhanced bacterial resistance to sanitization due to extracellular polymeric substances.

    PubMed

    Wang, Rong; Kalchayanand, Norasak; Schmidt, John W; Harhay, Dayna M

    2013-09-01

    Shiga toxin-producing Escherichia coli O157:H7 and Salmonella enterica serovar Typhimurium are important foodborne pathogens capable of forming single-species biofilms or coexisting in multispecies biofilm communities. Bacterial biofilm cells are usually more resistant to sanitization than their planktonic counterparts, so these foodborne pathogens in biofilms pose a serious food safety concern. We investigated how the coexistence of E. coli O157:H7 and Salmonella Typhimurium strains would affect bacterial planktonic growth competition and mixed biofilm composition. Furthermore, we also investigated how mixed biofilm formation would affect bacterial resistance to common sanitizers. Salmonella Typhimurium strains were able to outcompete E. coli strains in the planktonic growth phase; however, mixed biofilm development was highly dependent upon companion strain properties in terms of the expression of bacterial extracellular polymeric substances (EPS), including curli fimbriae and exopolysaccharide cellulose. The EPS-producing strains with higher biofilm-forming abilities were able to establish themselves in mixed biofilms more efficiently. In comparison to single-strain biofilms, Salmonella or E. coli strains with negative EPS expression obtained significantly enhanced resistance to sanitization by forming mixed biofilms with an EPS-producing companion strain of the other species. These observations indicate that the bacterial EPS components not only enhance the sanitizer resistance of the EPS-producing strains but also render protections to their companion strains, regardless of species, in mixed biofilms. Our study highlights the potential risk of cross-contamination by multispecies biofilms in food safety and the need for increased attention to proper sanitization practices in food processing facilities.

  20. Antibody-based Detection of Escherichia coli O157:H7 and Salmonella enterica Serovar Typhimurium Grown in Low-shear Modeled Microgravity

    NASA Astrophysics Data System (ADS)

    Nyquist-Battie, Cynthia; Freeman, Laura; Leckband, Kristen; Martinez, Stephanie; Ansley, Ariel; Lund, Deanna; Lim, Daniel V.

    2008-06-01

    With the advent of prolonged spaceflights, it is important to determine if antibody-based assays can be used to monitor food and water for bacterial contaminants. In the present work, a ground-based high aspect ratio vessel (HARV) was used to determine if low shear modeled microgravity (LSMMG) alters antibody-binding to E. coli O157:H7 and Salmonella enterica serovar Typhimurium. Antibody-bacteria binding was similar under LSMMG and normal gravity because there was no difference in amount of captured bacteria measured by colony forming units (CFU) between assays conducted in the HARV and a conventional roller flask. The ability of E. coli O157:H7 and Salmonella Typhimurium grown in LSMMG to bind specific antibodies was also studied. After incubations of 4, 18 or 36 h in the HARV or a shaking incubator, bacteria were harvested for enzyme-linked immunosorbent assays (ELISA). In the E. coli O157:H7 ELISA using a goat polyclonal primary antibody, LSMMG did not alter the linear range of detection (105-107 cells/ml) nor the signal to noise ratio at any bacterial concentration. Although insignificant changes in signal to noise ratios were evident, LSMMG did not alter the range of detection (105-107 cells/ml) for Salmonella Typhimurium in ELISAs using either a polyclonal or a monoclonal antibody. These results suggest that immunoassays may be used in spacecrafts because LSMMG does not have significant deleterious effects on antibody-binding to bacteria nor does it significantly alter surface antigens necessary for antibody-based methods.

  1. Porins from Salmonella enterica Serovar Typhimurium Activate the Transcription Factors Activating Protein 1 and NF-κB through the Raf-1-Mitogen-Activated Protein Kinase Cascade

    PubMed Central

    Galdiero, Massimiliano; Vitiello, Mariateresa; Sanzari, Emma; D’Isanto, Marina; Tortora, Annalisa; Longanella, Anna; Galdiero, Stefania

    2002-01-01

    In this study we examined the ability of Salmonella enterica serovar Typhimurium porins to activate activating protein 1 (AP-1) and nuclear factor κB (NF-κB) through the mitogen-activated protein kinase (MAPK) cascade, and we identified the AP-1-induced protein subunits. Our results demonstrate that these enzymes may participate in cell signaling pathways leading to AP-1 and NF-κB activation following porin stimulation of cells. Raf-1 was phosphorylated in response to the treatment of U937 cells with porins; moreover, the porin-mediated increase in Raf-1 phosphorylation is accompanied by the phosphorylation of MAPK kinase 1/2 (MEK1/2), p38, extracellular-signal-regulated kinase 1/2, and c-Jun N-terminal kinase. We used three different inhibitors of phosphorylation pathways: 2′-amino-3′-methoxyflavone (PD-098059), a selective inhibitor of MEK1 activator and the MAPK cascade; 4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)1H-imidazole (SB203580), a specific inhibitor of the p38 pathway; and 7β-acetoxy-1α,6β,9α-trihydroxy-8,13-epoxy-labd-14-en-11-one (forskolin), an inhibitor at the level of Raf-1 kinase. PD-098059 pretreatment of cells decreases AP-1 and NF-κB activation by lipopolysaccharide (LPS) but not by porins, and SB203580 pretreatment of cells decreases mainly AP-1 and NF-κB activation by porins; in contrast, forskolin pretreatment of cells does not affect AP-1 and NF-κB activation following either porin or LPS stimulation. Our data suggest that the p38 signaling pathway mainly regulates AP-1 and NF-κB activation in cells treated with S. enterica serovar Typhimurium porins. Antibody electrophoretic mobility shift assays showed that JunD and c-Fos binding is found in cells treated with porins, in cells treated with LPS, and in unstimulated cells. However, by 30 to 60 min of stimulation, a different complex including c-Jun appears in cells treated with porins or LPS, while the Fra-2 subunit is present only after porin stimulation

  2. Porins from Salmonella enterica serovar Typhimurium activate the transcription factors activating protein 1 and NF-kappaB through the Raf-1-mitogen-activated protein kinase cascade.

    PubMed

    Galdiero, Massimiliano; Vitiello, Mariateresa; Sanzari, Emma; D'Isanto, Marina; Tortora, Annalisa; Longanella, Anna; Galdiero, Stefania

    2002-02-01

    In this study we examined the ability of Salmonella enterica serovar Typhimurium porins to activate activating protein 1 (AP-1) and nuclear factor kappaB (NF-kappaB) through the mitogen-activated protein kinase (MAPK) cascade, and we identified the AP-1-induced protein subunits. Our results demonstrate that these enzymes may participate in cell signaling pathways leading to AP-1 and NF-kappaB activation following porin stimulation of cells. Raf-1 was phosphorylated in response to the treatment of U937 cells with porins; moreover, the porin-mediated increase in Raf-1 phosphorylation is accompanied by the phosphorylation of MAPK kinase 1/2 (MEK1/2), p38, extracellular-signal-regulated kinase 1/2, and c-Jun N-terminal kinase. We used three different inhibitors of phosphorylation pathways: 2'-amino-3'-methoxyflavone (PD-098059), a selective inhibitor of MEK1 activator and the MAPK cascade; 4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)1H-imidazole (SB203580), a specific inhibitor of the p38 pathway; and 7beta-acetoxy-1alpha,6beta,9alpha-trihydroxy-8,13-epoxy-labd-14-en-11-one (forskolin), an inhibitor at the level of Raf-1 kinase. PD-098059 pretreatment of cells decreases AP-1 and NF-kappaB activation by lipopolysaccharide (LPS) but not by porins, and SB203580 pretreatment of cells decreases mainly AP-1 and NF-kappaB activation by porins; in contrast, forskolin pretreatment of cells does not affect AP-1 and NF-kappaB activation following either porin or LPS stimulation. Our data suggest that the p38 signaling pathway mainly regulates AP-1 and NF-kappaB activation in cells treated with S. enterica serovar Typhimurium porins. Antibody electrophoretic mobility shift assays showed that JunD and c-Fos binding is found in cells treated with porins, in cells treated with LPS, and in unstimulated cells. However, by 30 to 60 min of stimulation, a different complex including c-Jun appears in cells treated with porins or LPS, while the Fra-2 subunit is present only

  3. Antimicrobial Effects of Mustard Flour and Acetic Acid against Escherichia coli O157:H7, Listeria monocytogenes, and Salmonella enterica Serovar Typhimurium

    PubMed Central

    Rhee, Min-Suk; Lee, Sun-Young; Dougherty, Richard H.; Kang, Dong-Hyun

    2003-01-01

    This study was designed to investigate the individual and combined effects of mustard flour and acetic acid in the inactivation of food-borne pathogenic bacteria stored at 5 and 22°C. Samples were prepared to achieve various concentrations by the addition of acetic acid (0, 0.5, or 1%) along with mustard flour (0, 10, or 20%) and 2% sodium chloride (fixed amount). Acid-adapted three-strain mixtures of Escherichia coli O157:H7, Listeria monocytogenes, and Salmonella enterica serovar Typhimurium strains (106 to 107 CFU/ml) were inoculated separately into prepared mustard samples stored at 5 and 22°C, and samples were assayed periodically. The order of bacterial resistance, assessed by the time required for the nominated populations to be reduced to undetectable levels against prepared mustards at 5°C, was S. enterica serovar Typhimurium (1 day) < E. coli O157:H7 (3 days) < L. monocytogenes (9 days). The food-borne pathogens tested were reduced much more rapidly at 22°C than at 5°C. There was no synergistic effect with regard to the killing of the pathogens tested with the addition of 0.5% acetic acid to the mustard flour (10 or 20%). Mustard in combination with 0.5% acetic acid had less bactericidal activity against the pathogens tested than did mustard alone. The reduction of E. coli O157:H7 and L. monocytogenes among the combined treatments on the same storage day was generally differentiated as follows: control < mustard in combination with 0.5% acetic acid < mustard alone < mustard in combination with 1% acetic acid < acetic acid alone. Our study indicates that acidic products may limit microbial growth or survival and that the addition of small amounts of acetic acid (0.5%) to mustard can retard the reduction of E. coli O157:H7 and L. monocytogenes. These antagonistic effects may be changed if mustard is used alone or in combination with >1% acetic acid. PMID:12732572

  4. Association of Salmonella enterica Serovar Enteritidis YafD with Resistance to Chicken Egg Albumen

    PubMed Central

    Lu, Sangwei; Killoran, Patrick B.; Riley, Lee W.

    2003-01-01

    Salmonella enterica serovar Enteritidis is a major cause of food-borne diseases in industrialized countries. The incidence of S. enterica serovar Enteritidis infections has increased substantially in recent decades, and S. enterica serovar Enteritidis is now one of the leading serovars of Salmonella in the United States. A unique epidemiological characteristic of S. enterica serovar Enteritidis is its association with chicken shell eggs, since approximately 80% of all human gastrointestinal diseases can be traced to contaminated egg products. Eggs are contaminated when bacteria from reproductive tissues of infected hens are packaged into the eggs and persist inside the hostile egg albumen environment. Therefore, resistance to egg albumen is an important aspect in the transmission of S. enterica serovar Enteritidis. We identified a gene, yafD from S. enterica serovar Enteritidis, whose overexpression conferred upon S. enterica serovar Typhimurium enhanced resistance to egg albumen, while disruption of this gene in S. enterica serovar Enteritidis rendered the organism more susceptible to egg albumen. YafD is homologous to members of an exonuclease-endonuclease-phosphatase family, including some enzymes involved in DNA repair. Furthermore, we discovered that egg albumen has nuclease activities and uses both circular and linear DNA as substrates. We propose that YafD provides a survival advantage to S. enterica serovar Enteritidis in eggs by repairing DNA damage caused by egg albumen and that it may be one of the biologic determinants that contribute to the epidemiological association of S. enterica serovar Enteritidis with egg products. PMID:14638758

  5. Association of Salmonella enterica serovar enteritidis yafD with resistance to chicken egg albumen.

    PubMed

    Lu, Sangwei; Killoran, Patrick B; Riley, Lee W

    2003-12-01

    Salmonella enterica serovar Enteritidis is a major cause of food-borne diseases in industrialized countries. The incidence of S. enterica serovar Enteritidis infections has increased substantially in recent decades, and S. enterica serovar Enteritidis is now one of the leading serovars of Salmonella in the United States. A unique epidemiological characteristic of S. enterica serovar Enteritidis is its association with chicken shell eggs, since approximately 80% of all human gastrointestinal diseases can be traced to contaminated egg products. Eggs are contaminated when bacteria from reproductive tissues of infected hens are packaged into the eggs and persist inside the hostile egg albumen environment. Therefore, resistance to egg albumen is an important aspect in the transmission of S. enterica serovar Enteritidis. We identified a gene, yafD from S. enterica serovar Enteritidis, whose overexpression conferred upon S. enterica serovar Typhimurium enhanced resistance to egg albumen, while disruption of this gene in S. enterica serovar Enteritidis rendered the organism more susceptible to egg albumen. YafD is homologous to members of an exonuclease-endonuclease-phosphatase family, including some enzymes involved in DNA repair. Furthermore, we discovered that egg albumen has nuclease activities and uses both circular and linear DNA as substrates. We propose that YafD provides a survival advantage to S. enterica serovar Enteritidis in eggs by repairing DNA damage caused by egg albumen and that it may be one of the biologic determinants that contribute to the epidemiological association of S. enterica serovar Enteritidis with egg products.

  6. Diversity of Genome Structure in Salmonella enterica Serovar Typhi Populations†

    PubMed Central

    Kothapalli, Sushma; Nair, Satheesh; Alokam, Suneetha; Pang, Tikki; Khakhria, Rasik; Woodward, David; Johnson, Wendy; Stocker, Bruce A. D.; Sanderson, Kenneth E.; Liu, Shu-Lin

    2005-01-01

    The genomes of most strains of Salmonella and Escherichia coli are highly conserved. In contrast, all 136 wild-type strains of Salmonella enterica serovar Typhi analyzed by partial digestion with I-CeuI (an endonuclease which cuts within the rrn operons) and pulsed-field gel electrophoresis and by PCR have rearrangements due to homologous recombination between the rrn operons leading to inversions and translocations. Recombination between rrn operons in culture is known to be equally frequent in S. enterica serovar Typhi and S. enterica serovar Typhimurium; thus, the recombinants in S. enterica serovar Typhi, but not those in S. enterica serovar Typhimurium, are able to survive in nature. However, even in S. enterica serovar Typhi the need for genome balance and the need for gene dosage impose limits on rearrangements. Of 100 strains of genome types 1 to 6, 72 were only 25.5 kb off genome balance (the relative lengths of the replichores during bidirectional replication from oriC to the termination of replication [Ter]), while 28 strains were less balanced (41 kb off balance), indicating that the survival of the best-balanced strains was greater. In addition, the need for appropriate gene dosage apparently selected against rearrangements which moved genes from their accustomed distance from oriC. Although rearrangements involving the seven rrn operons are very common in S. enterica serovar Typhi, other duplicated regions, such as the 25 IS200 elements, are very rarely involved in rearrangements. Large deletions and insertions in the genome are uncommon, except for deletions of Salmonella pathogenicity island 7 (usually 134 kb) from fragment I-CeuI-G and 40-kb insertions, possibly a prophage, in fragment I-CeuI-E. The phage types were determined, and the origins of the phage types appeared to be independent of the origins of the genome types. PMID:15805510

  7. Mobilome differences between Salmonella enterica serovars Anatum and Typhimurium isolated from cattle and humans and potential impact on virulence

    USDA-ARS?s Scientific Manuscript database

    Salmonella enterica subsp. enterica is an important group of pathogens capable of inhabiting a range of niches and hosts with varying degrees of impact, from commensal colonization to invasive infection. Recent outbreaks of multi-drug resistant S. enterica, attributed to consumption of contaminated ...

  8. Effects of nitrate or nitro supplementation, with or without added chlorate, on Salmonella enterica serovar Typhimurium and Escherichia coli in swine feces.

    PubMed

    Anderson, Robin C; Jung, Yong S; Oliver, Christy E; Horrocks, Shane M; Genovese, Kenneth J; Harvey, Roger B; Callaway, Todd R; Edrington, Thomas S; Nisbet, David J

    2007-02-01

    The effects of coincubating the active agent of an experimental chlorate product with nitrate or select nitro compounds, possible inducers and competing substrates for the targeted respiratory nitrate reductase, on concentrations of experimentally inoculated Salmonella enterica serovar Typhimurium and indigenous Escherichia coli were determined. Studies were completed in swine fecal suspensions as a prelude to the administration of these inhibitors to pigs. Results confirmed the bactericidal effect of chlorate (5 to 10 mM) against these fecal enterobacteria, reducing (P < 0.05) concentrations by > 2 log CFU ml(-1) after 3 to 6 h of incubation. An effect (P < 0.05) of pH was observed, with considerable regrowth of Salmonella and E. coli occurring after 24 h of incubation in suspensions buffered to pH 7.1 but not in suspensions buffered to pH 6.5 or 5.6. A 24-h coincubation of fecal suspensions with 5 to 10 mM chlorate and as little as 2.5 mM nitrate or 10 to 20 mM 2-nitro-1-propanol, 2-nitroethanol, and, sometimes, nitroethane decreased (P < 0.05) Salmonella but not necessarily E. coli concentrations. 2-Nitro-1-propanol and 2-nitroethanol exhibited inhibitory activity against Salmonella and E. coli by an undetermined mechanism, even in the absence of added chlorate.

  9. Leaching of Cryptosporidium parvum Oocysts, Escherichia coli, and a Salmonella enterica Serovar Typhimurium Bacteriophage through Intact Soil Cores following Surface Application and Injection of Slurry▿

    PubMed Central

    Forslund, Anita; Markussen, Bo; Toenner-Klank, Lise; Bech, Tina B.; Jacobsen, Ole Stig; Dalsgaard, Anders

    2011-01-01

    Increasing amounts of livestock manure are being applied to agricultural soil, but it is unknown to what extent this may be associated with contamination of aquatic recipients and groundwater if microorganisms are transported through the soil under natural weather conditions. The objective of this study was therefore to evaluate how injection and surface application of pig slurry on intact sandy clay loam soil cores influenced the leaching of Salmonella enterica serovar Typhimurium bacteriophage 28B, Escherichia coli, and Cryptosporidium parvum oocysts. All three microbial tracers were detected in the leachate on day 1, and the highest relative concentration was detected on the fourth day (0.1 pore volume). Although the concentration of the phage 28B declined over time, the phage was still found in leachate at day 148. C. parvum oocysts and chloride had an additional rise in the relative concentration at a 0.5 pore volume, corresponding to the exchange of the total pore volume. The leaching of E. coli was delayed compared with that of the added microbial tracers, indicating a stronger attachment to slurry particles, but E. coli could be detected up to 3 months. Significantly enhanced leaching of phage 28B and oocysts by the injection method was seen, whereas leaching of the indigenous E. coli was not affected by the application method. Preferential flow was the primary transport vehicle, and the diameter of the fractures in the intact soil cores facilitated transport of all sizes of microbial tracers under natural weather conditions. PMID:21948848

  10. Fur negatively regulates hns and is required for the expression of HilA and virulence in Salmonella enterica serovar Typhimurium.

    PubMed

    Troxell, Bryan; Sikes, Michael L; Fink, Ryan C; Vazquez-Torres, Andres; Jones-Carson, Jessica; Hassan, Hosni M

    2011-01-01

    Iron is an essential element for the survival of living cells. However, excess iron is toxic, and its uptake is exquisitely regulated by the ferric uptake regulator, Fur. In Salmonella, the Salmonella pathogenicity island 1 (SPI-1) encodes a type three secretion system, which is required for invasion of host epithelial cells in the small intestine. A major activator of SPI-1 is HilA, which is encoded within SPI-1. One known regulator of hilA is Fur. The mechanism of hilA regulation by Fur is unknown. We report here that Fur is required for virulence in Salmonella enterica serovar Typhimurium and that Fur is required for the activation of hilA, as well as of other HilA-dependent genes, invF and sipC. The Fur-dependent regulation of hilA was independent of PhoP, a known repressor of hilA. Instead, the expression of the gene coding for the histone-like protein, hns, was significantly derepressed in the fur mutant. Indeed, the activation of hilA by Fur was dependent on 28 nucleotides located upstream of hns. Moreover, we used chromatin immunoprecipitation to show that Fur bound, in vivo, to the upstream region of hns in a metal-dependent fashion. Finally, deletion of fur in an hns mutant resulted in Fur-independent activation of hilA. In conclusion, Fur activates hilA by repressing the expression of hns.

  11. Expression of the Fis protein is sustained in late-exponential- and stationary-phase cultures of Salmonella enterica serovar Typhimurium grown in the absence of aeration.

    PubMed

    O Cróinín, Tadhg; Dorman, Charles J

    2007-10-01

    The classic expression pattern of the Fis global regulatory protein during batch culture consists of a high peak in the early logarithmic phase of growth, followed by a sharp decrease through mid-exponential growth phase until Fis is almost undetectable at the end of the exponential phase. We discovered that this pattern is contingent on the growth regime. In Salmonella enterica serovar Typhimurium cultures grown in non-aerated SPI1-inducing conditions, Fis can be detected readily in stationary phase. On the other hand, cultures grown with standard aeration showed the classic Fis expression pattern. Sustained Fis expression in non-aerated cultures was also detected in some Escherichia coli strains, but not in others. This novel pattern of Fis expression was independent of sequence differences in the fis promoter regions of Salmonella and E. coli. Instead, a clear negative correlation between the expression of the Fis protein and of the stress-and-stationary-phase sigma factor RpoS was observed in a variety of strains. An rpoS mutant displayed elevated levels of Fis and had a higher frequency of epithelial cell invasion under these growth conditions. We discuss a model whereby Fis and RpoS levels vary in response to environmental signals allowing the expression and repression of SPI1 invasion genes.

  12. Effect of frequency and waveform on inactivation of Escherichia coli O157:H7 and Salmonella enterica Serovar Typhimurium in salsa by ohmic heating.

    PubMed

    Lee, Su-Yeon; Ryu, Sangryeol; Kang, Dong-Hyun

    2013-01-01

    The effect of frequency of alternating current during ohmic heating on electrode corrosion, heating rate, inactivation of food-borne pathogens, and quality of salsa was investigated. The impact of waveform on heating rate was also investigated. Salsa was treated with various frequencies (60 Hz to 20 kHz) and waveforms (sine, square, and sawtooth) at a constant electric field strength of 12.5 V/cm. Electrode corrosion did not occur when the frequency exceeded 1 kHz. The heating rate of the sample was dependent on frequency up to 500 Hz, but there was no significant difference (P > 0.05) in the heating rate when the frequency was increased above 1 kHz. The electrical conductivity of the sample increased with a rise in the frequency. At a frequency of 60 Hz, the square wave produced a lower heating rate than that of sine and sawtooth waves. The heating rate between waveforms was not significantly (P > 0.05) different when the frequency was >500 Hz. As the frequency increased, the treatment time required to reduce Escherichia coli O157:H7 and Salmonella enterica serovar Typhimurium to below the detection limit (1 log CFU/g) decreased without affecting product quality. These results suggest that ohmic heating can be effectively used to pasteurize salsa and that the effect of inactivation is dependent on frequency and electrical conductivity rather than waveform.

  13. Long-Term Dissemination of CTX-M-5-Producing Hypermutable Salmonella enterica Serovar Typhimurium Sequence Type 328 Strains in Russia, Belarus, and Kazakhstan

    PubMed Central

    Ilina, Elena N.; Malakhova, Maja V.; Carattoli, Alessandra; Azizov, Ilya S.; Tapalski, Dmitry V.; Kozlov, Roman S.; Edelstein, Mikhail V.

    2014-01-01

    In this paper, we present evidence of long-term circulation of cefotaxime-resistant clonally related Salmonella enterica serovar Typhimurium strains over a broad geographic area. The genetic relatedness of 88 isolates collected from multiple outbreaks and sporadic cases of nosocomial salmonellosis in various parts of Russia, Belarus, and Kazakhstan from 1996 to 2009 was established by multilocus tandem-repeat analysis (MLVA) and multilocus sequence typing (MLST). The isolates belong to sequence type 328 (ST328) and produce CTX-M-5 β-lactamase, whose gene is carried by highly related non-self-conjugative but mobilizable plasmids. Resistance to nalidixic acid and low-level resistance to ciprofloxacin is present in 37 (42%) of the isolates and in all cases is determined by various single point mutations in the gyrA gene quinolone resistance-determining region (QRDR). Isolates of the described clonal group exhibit a hypermutable phenotype that probably facilitates independent acquisition of quinolone resistance mutations. PMID:24957829

  14. Protection of neonatal mice from lethal enterovirus 71 infection by maternal immunization with attenuated Salmonella enterica serovar Typhimurium expressing VP1 of enterovirus 71.

    PubMed

    Chiu, Cheng-Hsun; Chu, Chishih; He, Chao-Che; Lin, Tzou-Yien

    2006-06-01

    This study describes the potential use of attenuated Salmonella enterica serovar Typhimurium strains to express and deliver VP1 of enterovirus 71 (EV71) as a vaccination strategy to prevent EV71 infection in mice. When orally administered to BALB/c mice, both attenuated carrier strains, CNP101 and SL7207, were able to efficiently invade livers and spleens, while only the virulence plasmid-carrying strain SL7207 persisted for more than 30 days in these organs. A recombinant in vivo-regulated promoter expression plasmid expressing VP1 antigen of EV71 was constructed. The expression of the VP1, directed by the pagC promoter, in attenuated Salmonella was confirmed by Western blot hybridization. Both humoral and cellular immune responses were elicited in mice by oral immunization with such Salmonella-based VP1 vaccines. We evaluated the protective efficacy of the vaccines in mice using a maternal immunization protocol. With a lethal challenge, ICR newborn mice born to dams immunized with Salmonella-based VP1 vaccine showed a 50-60% survival; in contrast, none of the mice in the control group survived the challenge. Our data indicated that Salmonella-based VP1 subunit vaccines are a promising vaccine strategy in the prevention of EV71 infection.

  15. Effect of Salmonella enterica serovar Typhimurium VNP20009 and VNP20009 with restored chemotaxis on 4T1 mouse mammary carcinoma progression.

    PubMed

    Coutermarsh-Ott, Sheryl L; Broadway, Katherine M; Scharf, Birgit E; Allen, Irving C

    2017-05-16

    A variety of bacterial strains have been evaluated as bio-therapeutic and immunomodulatory agents to treat cancer. One such strain, Salmonella enterica serovar Typhimurium VNP20009, which is attenuated by a purine auxotrophic mutation and modified lipid A, is characterized in previous models as a safely administered, tumor colonizing agent. However, earlier work tended to use less aggressive cancer cell lines and immunocompromised animal models. Here, we investigated the safety and efficacy of VNP20009 in a highly malignant murine model of human breast cancer. Additionally, as VNP20009 has recently been found to have a defective chemotaxis system, we tested whether restoring chemotaxis would improve anti-cancer properties in this model system. Exposure to VNP20009 had no significant effect on primary mammary tumor size or pulmonary metastasis, and the tumor colonizing process appeared chemotaxis independent. Moreover, tumor-bearing mice exposed to Salmonella exhibited increased morbidity that was associated with significant liver disease. Our results suggest that VNP20009 may not be safe or efficacious when used in aggressive, metastatic breast cancer models utilizing immunocompetent animals.

  16. Effect of Salmonella enterica serovar Typhimurium VNP20009 and VNP20009 with restored chemotaxis on 4T1 mouse mammary carcinoma progression

    PubMed Central

    Scharf, Birgit E.; Allen, Irving C.

    2017-01-01

    A variety of bacterial strains have been evaluated as bio-therapeutic and immunomodulatory agents to treat cancer. One such strain, Salmonella enterica serovar Typhimurium VNP20009, which is attenuated by a purine auxotrophic mutation and modified lipid A, is characterized in previous models as a safely administered, tumor colonizing agent. However, earlier work tended to use less aggressive cancer cell lines and immunocompromised animal models. Here, we investigated the safety and efficacy of VNP20009 in a highly malignant murine model of human breast cancer. Additionally, as VNP20009 has recently been found to have a defective chemotaxis system, we tested whether restoring chemotaxis would improve anti-cancer properties in this model system. Exposure to VNP20009 had no significant effect on primary mammary tumor size or pulmonary metastasis, and the tumor colonizing process appeared chemotaxis independent. Moreover, tumor-bearing mice exposed to Salmonella exhibited increased morbidity that was associated with significant liver disease. Our results suggest that VNP20009 may not be safe or efficacious when used in aggressive, metastatic breast cancer models utilizing immunocompetent animals. PMID:28431394

  17. Effect of Frequency and Waveform on Inactivation of Escherichia coli O157:H7 and Salmonella enterica Serovar Typhimurium in Salsa by Ohmic Heating

    PubMed Central

    Lee, Su-Yeon; Ryu, Sangryeol

    2013-01-01

    The effect of frequency of alternating current during ohmic heating on electrode corrosion, heating rate, inactivation of food-borne pathogens, and quality of salsa was investigated. The impact of waveform on heating rate was also investigated. Salsa was treated with various frequencies (60 Hz to 20 kHz) and waveforms (sine, square, and sawtooth) at a constant electric field strength of 12.5 V/cm. Electrode corrosion did not occur when the frequency exceeded 1 kHz. The heating rate of the sample was dependent on frequency up to 500 Hz, but there was no significant difference (P > 0.05) in the heating rate when the frequency was increased above 1 kHz. The electrical conductivity of the sample increased with a rise in the frequency. At a frequency of 60 Hz, the square wave produced a lower heating rate than that of sine and sawtooth waves. The heating rate between waveforms was not significantly (P > 0.05) different when the frequency was >500 Hz. As the frequency increased, the treatment time required to reduce Escherichia coli O157:H7 and Salmonella enterica serovar Typhimurium to below the detection limit (1 log CFU/g) decreased without affecting product quality. These results suggest that ohmic heating can be effectively used to pasteurize salsa and that the effect of inactivation is dependent on frequency and electrical conductivity rather than waveform. PMID:23023752

  18. Leaching of Cryptosporidium parvum oocysts, Escherichia coli, and a Salmonella enterica serovar Typhimurium bacteriophage through intact soil cores following surface application and injection of slurry.

    PubMed

    Forslund, Anita; Markussen, Bo; Toenner-Klank, Lise; Bech, Tina B; Jacobsen, Ole Stig; Dalsgaard, Anders

    2011-11-01

    Increasing amounts of livestock manure are being applied to agricultural soil, but it is unknown to what extent this may be associated with contamination of aquatic recipients and groundwater if microorganisms are transported through the soil under natural weather conditions. The objective of this study was therefore to evaluate how injection and surface application of pig slurry on intact sandy clay loam soil cores influenced the leaching of Salmonella enterica serovar Typhimurium bacteriophage 28B, Escherichia coli, and Cryptosporidium parvum oocysts. All three microbial tracers were detected in the leachate on day 1, and the highest relative concentration was detected on the fourth day (0.1 pore volume). Although the concentration of the phage 28B declined over time, the phage was still found in leachate at day 148. C. parvum oocysts and chloride had an additional rise in the relative concentration at a 0.5 pore volume, corresponding to the exchange of the total pore volume. The leaching of E. coli was delayed compared with that of the added microbial tracers, indicating a stronger attachment to slurry particles, but E. coli could be detected up to 3 months. Significantly enhanced leaching of phage 28B and oocysts by the injection method was seen, whereas leaching of the indigenous E. coli was not affected by the application method. Preferential flow was the primary transport vehicle, and the diameter of the fractures in the intact soil cores facilitated transport of all sizes of microbial tracers under natural weather conditions.

  19. ubiJ, a New Gene Required for Aerobic Growth and Proliferation in Macrophage, Is Involved in Coenzyme Q Biosynthesis in Escherichia coli and Salmonella enterica Serovar Typhimurium

    PubMed Central

    Aussel, Laurent; Loiseau, Laurent; Hajj Chehade, Mahmoud; Pocachard, Bérengère; Fontecave, Marc; Pierrel, Fabien

    2014-01-01

    Ubiquinone (coenzyme Q or Q8) is a redox active lipid which functions in the respiratory electron transport chain and plays a crucial role in energy-generating processes. In both Escherichia coli and Salmonella enterica serovar Typhimurium, the yigP gene is located between ubiE and ubiB, all three being likely to constitute an operon. In this work, we showed that the uncharacterized yigP gene was involved in Q8 biosynthesis in both strains, and we have renamed it ubiJ. Under aerobic conditions, an ubiJ mutant was found to be impaired for Q8 biosynthesis and for growth in rich medium but did not present any defect anaerobically. Surprisingly, the C-terminal 50 amino acids, predicted to interact with lipids, were sufficient to restore Q8 biosynthesis and growth of the ubiJ mutant. Salmonella ubiE and ubiB mutants were impaired in Q8 biosynthesis and in respiration using different electron acceptors. Moreover, ubiE, ubiJ, and ubiB mutants were all impaired for Salmonella intracellular proliferation in macrophages. Taken together, our data establish an important role for UbiJ in Q8 biosynthesis and reveal an unexpected link between Q8 and virulence. They also emphasize that Salmonella organisms in an intracellular lifestyle rely on aerobic respiration to survive and proliferate within macrophages. PMID:24142253

  20. Outer membrane vesicles derived from Salmonella Typhimurium mutants with truncated LPS induce cross-protective immune responses against infection of Salmonella enterica serovars in the mouse model.

    PubMed

    Liu, Qiong; Liu, Qing; Yi, Jie; Liang, Kang; Liu, Tian; Roland, Kenneth L; Jiang, Yanlong; Kong, Qingke

    2016-12-01

    Salmonella enterica cause diarrheal and systemic diseases and are of considerable concern worldwide. Vaccines that are cross-protective against multiple serovars could provide effective control of Salmonella-mediated diseases. Bacteria-derived outer membrane vesicles (OMVs) are highly immunogenic and are capable of eliciting protective immune responses. Alterations in lipopolysaccharide (LPS) length can result in outer membrane remodeling and composition of outer membrane proteins (OMPs) changing. In this study, we investigated the impact of truncated LPS on both the production and immunogenicity of Salmonella OMVs, including the ability of OMVs to elicit cross-protection against challenge by heterologous Salmonella strains. We found that mutations in waaJ and rfbP enhanced vesiculation, while mutations in waaC, waaF and waaG inhibited this process. Animal experiments indicated that OMVs from waaC, rfaH and rfbP mutants induced stronger serum immune responses compared to OMVs from the parent strain, while all elicited protective responses against the wild-type S. Typhimurium challenge. Furthermore, intranasal or intraperitoneal immunization with OMVs derived from the waaC and rfbP mutants elicited significantly higher cross-reactive IgG responses and provided enhanced cross-protection against S. Choleraesuis and S. Enteritidis challenge than the wild-type OMVs. These results indicate that truncated-LPS OMVs are capable of conferring cross protection against multiple serotypes of Salmonella infection.

  1. Identification of cptA, a PmrA-Regulated Locus Required for Phosphoethanolamine Modification of the Salmonella enterica Serovar Typhimurium Lipopolysaccharide Core

    PubMed Central

    Tamayo, R.; Choudhury, B.; Septer, A.; Merighi, M.; Carlson, R.; Gunn, J. S.

    2005-01-01

    In response to the in vivo environment, the Salmonella enterica serovar Typhimurium lipopolysaccharide (LPS) is modified. These modifications are controlled in part by the two-component regulatory system PmrA-PmrB, with the addition of 4-aminoarabinose (Ara4N) to the lipid A and phosphoethanolamine (pEtN) to the lipid A and core. Here we demonstrate that the PmrA-regulated STM4118 (cptA) gene is necessary for the addition of pEtN to the LPS core. pmrC, a PmrA-regulated gene necessary for the addition of pEtN to lipid A, did not affect core pEtN addition. Although imparting a similar surface charge modification as Ara4N, which greatly affects polymyxin B resistance and murine virulence, neither pmrC nor cptA plays a dramatic role in antimicrobial peptide resistance in vitro or virulence in the mouse model. Therefore, factors other than surface charge/electrostatic interaction contribute to resistance to antimicrobial peptides such as polymyxin B. PMID:15866924

  2. Regulation of marA, soxS, rob, acrAB and micF in Salmonella enterica serovar Typhimurium.

    PubMed

    Hartog, Efrat; Ben-Shalom, Lior; Shachar, Dina; Matthews, Karl R; Yaron, Sima

    2008-12-01

    Importance of the overexpression of AcrAB efflux pumps in the low-level resistance of pathogens to antimicrobials requires a better understanding of the AcrAB regulation. The goal of the present research was to study the transcription of acrAB, as well as the genes that play a role in its regulation in Salmonella enterica serovar Typhimurium. We monitored the transcription of these genes during growth at 30 degrees C and 37 degrees C, and thoroughly studied the effect of salicylate, paraquat and decanoate. The strengths of the promoters' activities were ordered from strong to weak as micF > rob > acrAB > soxS, marRAB. At both temperatures, marRAB was mainly upregulated by salicylate and decanoate, soxS by paraquat and acrAB and micF by all three compounds. rob was always downregulated. Transcription rates of all promoters were at the greatest level at the beginning of the stationary phase and, except for soxS, levels of transcription and induction were greater at 37 degrees C. Results show that despite the promoters' similar activity and the sequence homology between Escherichia coli and S. typhimurium, regulation of the investigated genes of both strains differed in the response to temperature. This difference was found to be dependent on the promoters' sequence, as the marRAB and acrAB promoters maintained their original dependence on temperature when they were analyzed in the other strain. Hence, it is most likely that the nutrient-limited environment at 37 degrees C in the human body will lead to increased transcription of marA, acrAB and micF in Salmonella, enhancing the resistance properties of the bacteria.

  3. No protective effects of high-dosage dietary zinc oxide on weaned pigs infected with Salmonella enterica serovar typhimurium DT104.

    PubMed

    Janczyk, Pawel; Kreuzer, Susanne; Assmus, Jens; Nöckler, Karsten; Brockmann, Gudrun A

    2013-05-01

    Twenty-eight-day-old weaned pigs were fed diets with a low (LZn), medium (MZn), or high (MZn) Zn concentration (50 to 80, 150, or 2,500 mg Zn/kg of diet, respectively) provided as zinc oxide (ZnO)(24 pigs per group). They were infected orally with Salmonella enterica serovar Typhimurium DT104 on day 32. Salmonellae were cultivated from feces (up to 42 days postinfection [dpi]) and organs (2 and 42 dpi). Activation of the adaptive systemic and mucosal immune systems was investigated by recording anti-Salmonella IgG levels and levels of B and T lymphocyte subpopulations in blood and gut-associated lymphatic tissue. Growth performance was recorded as well. Salmonellae were shed at higher levels and for longer periods in the HZn group (P < 0.05), with no differences in the tissues. At 2 dpi, the relative percentages of CD4(+) T helper cells (P < 0.01) and of CD2(+) T and NK cells (P < 0.01) in blood were reduced from the relative cell counts obtained at 0 dpi, irrespective of the Zn group. The lowest percentage of cytotoxic T cells was found 14 dpi in the HZn group relative to the MZn (P < 0.05) and LZn (P < 0.01) groups. Supplementation of the feed with 2,500 mg Zn/kg of diet immediately after weaning could positively affect the immune responses of piglets infected with Salmonella Typhimurium, but for a short period only. After 2 weeks, all positive effects disappeared, and rather negative effects, such as higher shedding of salmonellae, lower T cell frequencies, and worse performance, occurred. Thus, supplementation with ZnO at high levels in the pig industry should be limited to 2 to 3 weeks.

  4. Development of a novel in-water vaccination protocol for DNA adenine methylase deficient Salmonella enterica serovar Typhimurium vaccine in adult sheep.

    PubMed

    Mohler, V L; Heithoff, D M; Mahan, M J; Hornitzky, M A; Thomson, P C; House, J K

    2012-02-14

    Intensive livestock production is associated with an increased incidence of salmonellosis. The risk of infection and the subsequent public health concern is attributed to increased pathogen exposure and disease susceptibility due to multiple stressors experienced by livestock from farm to feedlot. Traditional parenteral vaccine methods can further stress susceptible populations and cause carcass damage, adverse reactions, and resultant increased production costs. As a potential means to address these issues, in-water delivery of live attenuated vaccines affords a low cost, low-stress method for immunization of livestock populations that is not associated with the adverse handling stressors and injection reactions associated with parenteral administration. We have previously established that in-water administration of a Salmonella enterica serovar Typhimurium dam vaccine conferred significant protection in livestock. While these experimental trials hold significant promise, the ultimate measure of the vaccine will not be established until it has undergone clinical testing in the field wherein environmental and sanitary conditions are variable. Here we show that in-water administration of a S. Typhimurium dam attenuated vaccine was safe, stable, and well-tolerated in adult sheep. The dam vaccine did not alter water consumption or vaccine dosing; remained viable under a wide range of temperatures (21-37°C); did not proliferate within fecal-contaminated trough water; and was associated with minimal fecal shedding and clinical disease as a consequence of vaccination. The capacity of Salmonella dam attenuated vaccines to be delivered in drinking water to protect livestock from virulent Salmonella challenge offers an effective, economical, stressor-free Salmonella prophylaxis for intensive livestock production systems.

  5. The Impact of 18 Ancestral and Horizontally-Acquired Regulatory Proteins upon the Transcriptome and sRNA Landscape of Salmonella enterica serovar Typhimurium.

    PubMed

    Colgan, Aoife M; Kröger, Carsten; Diard, Médéric; Hardt, Wolf-Dietrich; Puente, José L; Sivasankaran, Sathesh K; Hokamp, Karsten; Hinton, Jay C D

    2016-08-01

    We know a great deal about the genes used by the model pathogen Salmonella enterica serovar Typhimurium to cause disease, but less about global gene regulation. New tools for studying transcripts at the single nucleotide level now offer an unparalleled opportunity to understand the bacterial transcriptome, and expression of the small RNAs (sRNA) and coding genes responsible for the establishment of infection. Here, we define the transcriptomes of 18 mutants lacking virulence-related global regulatory systems that modulate the expression of the SPI1 and SPI2 Type 3 secretion systems of S. Typhimurium strain 4/74. Using infection-relevant growth conditions, we identified a total of 1257 coding genes that are controlled by one or more regulatory system, including a sub-class of genes that reflect a new level of cross-talk between SPI1 and SPI2. We directly compared the roles played by the major transcriptional regulators in the expression of sRNAs, and discovered that the RpoS (σ38) sigma factor modulates the expression of 23% of sRNAs, many more than other regulatory systems. The impact of the RNA chaperone Hfq upon the steady state levels of 280 sRNA transcripts is described, and we found 13 sRNAs that are co-regulated with SPI1 and SPI2 virulence genes. We report the first example of an sRNA, STnc1480, that is subject to silencing by H-NS and subsequent counter-silencing by PhoP and SlyA. The data for these 18 regulatory systems is now available to the bacterial research community in a user-friendly online resource, SalComRegulon.

  6. The Impact of 18 Ancestral and Horizontally-Acquired Regulatory Proteins upon the Transcriptome and sRNA Landscape of Salmonella enterica serovar Typhimurium

    PubMed Central

    Colgan, Aoife M.; Diard, Médéric; Hardt, Wolf-Dietrich; Puente, José L.; Sivasankaran, Sathesh K.; Hokamp, Karsten; Hinton, Jay C. D.

    2016-01-01

    We know a great deal about the genes used by the model pathogen Salmonella enterica serovar Typhimurium to cause disease, but less about global gene regulation. New tools for studying transcripts at the single nucleotide level now offer an unparalleled opportunity to understand the bacterial transcriptome, and expression of the small RNAs (sRNA) and coding genes responsible for the establishment of infection. Here, we define the transcriptomes of 18 mutants lacking virulence-related global regulatory systems that modulate the expression of the SPI1 and SPI2 Type 3 secretion systems of S. Typhimurium strain 4/74. Using infection-relevant growth conditions, we identified a total of 1257 coding genes that are controlled by one or more regulatory system, including a sub-class of genes that reflect a new level of cross-talk between SPI1 and SPI2. We directly compared the roles played by the major transcriptional regulators in the expression of sRNAs, and discovered that the RpoS (σ38) sigma factor modulates the expression of 23% of sRNAs, many more than other regulatory systems. The impact of the RNA chaperone Hfq upon the steady state levels of 280 sRNA transcripts is described, and we found 13 sRNAs that are co-regulated with SPI1 and SPI2 virulence genes. We report the first example of an sRNA, STnc1480, that is subject to silencing by H-NS and subsequent counter-silencing by PhoP and SlyA. The data for these 18 regulatory systems is now available to the bacterial research community in a user-friendly online resource, SalComRegulon. PMID:27564394

  7. Genome-wide analysis of the PreA/PreB (QseB/QseC) regulon of Salmonella enterica serovar Typhimurium

    PubMed Central

    2009-01-01

    Background The Salmonella PreA/PreB two-component system (TCS) is an ortholog of the QseBC TCS of Escherichia coli. In both Salmonella and E. coli, this system has been shown to affect motility and virulence in response to quorum-sensing and hormonal signals, and to affect the transcription of the Salmonella enterica serovar Typhimurium (S. Typhimurium) pmrAB operon, which encodes an important virulence-associated TCS. Results To determine the PreA/PreB regulon in S. Typhimurium, we performed DNA microarrays comparing the wild type strain and various preA and/or preB mutants in the presence of ectopically expressed preA (qseB). These data confirmed our previous findings of the negative effect of PreB on PreA gene regulation and identified candidate PreA-regulated genes. A proportion of the activated loci were previously identified as PmrA-activated genes (yibD, pmrAB, cptA, etc.) or were genes located in the local region around preA, including the preAB operon. The transcriptional units were defined in this local region by RT-PCR, suggesting three PreA activated operons composed of preA-preB, mdaB-ygiN, and ygiW-STM3175. Several putative virulence-related phenotypes were examined for preAB mutants, resulting in the observation of a host cell invasion and slight virulence defect of a preAB mutant. Contrary to previous reports on this TCS, we were unable to show a PreA/PreB-dependent effect of the quorum-sensing signal AI-2 or of epinephrine on S. Typhimurium with regard to bacterial motility. Conclusion This work further characterizes this unorthadox OmpR/EnvZ class TCS and provides novel candidate regulated genes for further study. This first in-depth study of the PreA/PreB regulatory system phenotypes and regulation suggests significant comparative differences to the reported function of the orthologous QseB/QseC in E. coli. PMID:19236707

  8. Two draft genome sequences of a new serovar of Salmonella enterica, serovar Lubbock

    USDA-ARS?s Scientific Manuscript database

    Salmonella enterica is principally a foodborne pathogen that shows considerable serovar diversity. In this report, we present two draft genome sequences of Salmonella enterica subsp. enterica serovar Lubbock, a novel serovar....

  9. Two Draft Genome Sequences of a New Serovar of Salmonella enterica, Serovar Lubbock

    PubMed Central

    den Bakker, Henk C.; Nightingale, Kendra K.; Brichta-Harhay, Dayna M.; Edrington, Thomas S.; Loneragan, Guy H.

    2015-01-01

    Salmonella enterica is principally a foodborne pathogen that shows considerable serovar diversity. In this report, we present two draft genome sequences of Salmonella enterica subsp. enterica serovar Lubbock, a novel serovar. PMID:25883279

  10. Use of multiple-locus variable-number of tandem repeats analysis (MLVA) to investigate genetic diversity of Salmonella enterica subsp. enterica serovar Typhimurium isolates from human, food, and veterinary sources.

    PubMed

    Mateva, Gergana; Pedersen, Karl; Sørensen, Gitte; Asseva, Galina; Daskalov, Hristo; Petrov, Petar; Kantardjiev, Todor; Alexandar, Irina; Löfström, Charlotta

    2017-08-23

    Salmonella enterica subspecies enterica serovar Typhimurium is the most common zoonotic pathogen in Bulgaria. To allow efficient outbreak investigations and surveillance in the food chain, accurate and discriminatory methods for typing are needed. This study evaluated the use of multiple-locus variable-number of tandem repeats analysis (MLVA) and compared results with antimicrobial resistance (AMR) determinations for 100 S. Typhimurium strains isolated in Bulgaria during 2008-2012 (50 veterinary/food and 50 human isolates). Results showed that isolates were divided into 80 and 34 groups using MLVA and AMR, respectively. Simpson's index of diversity was determined to 0.994 ± 0.003 and 0.945 ± 0.012. The most frequently encountered MLVA profiles were 3-11-9-NA-211 (n = 5); 3-12-9-NA-211 (n = 3); 3-12-11-21-311 (n = 3); 3-17-10-NA-311 (n = 3); 2-20-9-7-212 (n = 3); and 2-23-NA-NA-111 (n = 3). No clustering of isolates related to susceptibility/resistance to antimicrobials, source of isolation, or year of isolation was observed. Some MLVA types were found in both human and veterinary/food isolates, indicating a possible route of transmission. A majority (83%) of the isolates were found to be resistant against at least one antimicrobial and 44% against ≥4 antimicrobials. Further studies are needed to verify MLVA usefulness over a longer period of time and with more isolates, including outbreak strains. © 2017 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.

  11. Optimization of inactivated H5N9 highly pathogenic avian influenza vaccine and inactivated Salmonella enterica serovar Typhimurium vaccine with antigen dose and prime-boost regimen in domestic ducks.

    PubMed

    Yuk, Seong-Su; To, Eredene-Ochir; Kwon, Jung-Hoon; Noh, Jin-Yong; Hong, Woo-Tack; Jeong, Jei-Hyun; Gwon, Gyeong-Bin; Song, Chang-Seon

    2017-09-01

    Owing to the increase in the number of diseases affecting ducks and the demand for food safety by consumers, vaccination has become one of the factors that influence duck meat productivity. The highly pathogenic avian influenza (HPAI) virus is one of the most prevalent and causes one of the most lethal diseases in domestic ducks, and Salmonella enterica serovar Typhimurium is a food-borne pathogen persistent in the domestic duck population. To better understand the optimal usage of HPAI and S. enterica serovar Typhimurium vaccines, we aimed to determine antigen dose, oil and gel adjuvant usage with prime-boost regimen, and vaccination age, inducing the best immune response in ducks, without an effect on body weight gain. In the case of the inactivated H5N9 vaccine, a single dose of vaccine was inadequate to induce proper antibody titer when administered to day-old ducks, which necessitates boost vaccination. Administration of the oil-adjuvanted H5N9 vaccine administration in day-old and 2-week-old ducks resulted in a lower body weight at the time of slaughtering, compared to that of gel-adjuvanted H5N9 vaccine. However, gel-adjuvanted H5N9 vaccine failed to induce proper immune response to an extent recommend by OIE-World Organization for Animal Health. In the case of the Salmonella enterica serovar Typhimurium vaccine, a moderate or low dose of vaccine was appropriate for day-old ducks receiving the gel prime-oil boost vaccination. Single vaccination with oil adjuvants affects the mean body weight of 7-week-old ducks, suggesting that the gel adjuvant is more suitable for meat production. We expect that the use of adjuvants in a prime-boost regimen and at antigen doses set in this study will be helpful to maximize body weight in the case of domestic duck production at the actual farm site. © 2017 Poultry Science Association Inc.

  12. Survival of Salmonella enterica serovar Typhimurium within late endosomal-lysosomal compartments of B lymphocytes is associated with the inability to use the vacuolar alternative major histocompatibility complex class I antigen-processing pathway.

    PubMed

    Rosales-Reyes, Roberto; Alpuche-Aranda, Celia; Ramírez-Aguilar, María de la Luz; Castro-Eguiluz, Angel Denisse; Ortiz-Navarrete, Vianney

    2005-07-01

    Gamma interferon (IFN-gamma)-activated macrophages use an alternative processing mechanism to present Salmonella antigens to CD8(+) T lymphocytes. This pathway involves processing of antigen in a vacuolar compartment followed by secretion and loading of antigenic peptides to major histocompatibility complex class I (MHC-I) molecules on macrophage cell surface and bystander cells. In this study, we have shown that B lymphocytes are not able to process Salmonella antigens using this alternative pathway. This is due to differences in Salmonella enterica serovar Typhimurium-containing vacuoles (SCV) when comparing late endosomal-lysosomal processing compartments in B lymphocytes to those in macrophages. The IFN-gamma-activated IC21 macrophage cell line and A-20 B-cell line were infected with live or dead Salmonella enterica serovar Typhimurium. The SCV in B cells were in a late endosomal-lysosomal compartment, whereas SCV in macrophages were remodeled to a non-characteristic late endosomal-lysosomal compartment over time. Despite the difference in SCV within macrophages and B lymphocytes, S. enterica serovar Typhimurium survives more efficiently within the IFN-gamma-activated B cells than in activated macrophage cell lines. Similar results were found during in vivo acute infection. We determined that a lack of remodeling of late endosomal-lysosomal compartments by live Salmonella infection in B lymphocytes is associated with the inability to use the alternative MHC-I antigen-processing pathway, providing a survival advantage to the bacterium. Our data also suggest that the B lymphocyte late endosome-lysosome environment allows the expression of Salmonella virulence mechanisms favoring B lymphocytes in addition to macrophages and dendritic cells as a reservoir during in vivo infection.

  13. Inactivation of Bacillus cereus and Salmonella enterica serovar Typhimurium by aqueous ozone (O3): Modeling and Uv-Vis spectroscopic analysis

    USDA-ARS?s Scientific Manuscript database

    Ozone (O3) is a natural antimicrobial agent with potential applications in food industry. In this study, inactivation of Bacillus cereus and Salmonella enterica Typhimurium by aqueous ozone was evaluated. Ozone gas was generated using a domestic ozone generator with an output of 200 mg/hr (approx. 0...

  14. Immunological responses induced by asd and wzy/asd mutant strains of Salmonella enterica serovar Typhimurium in BALB/c mice.

    PubMed

    Piao, Hong Hua; Tam, Vo Thi Minh; Na, Hee Sam; Kim, Hyun Ju; Ryu, Phil Youl; Kim, Soo Young; Rhee, Joon Haeng; Choy, Hyon E; Kim, Suhng Wook; Hong, Yeongjin

    2010-08-01

    Attenuated bacteria have long been developed as vaccine candidates but can have some disadvantages, such as the potential for damage to immune organs due to insufficient clearance. To minimize these disadvantages, we generated Salmonella enterica serovar Typhimurium mutants SHJ2104 (asd::cm) and HTSaYA (wzy::km, asd::cm). The wzy gene codes for the O-antigen polymerase, which is involved in lipopolysaccharide (LPS) biosynthesis, and asd codes for aspartate beta-semialdehyde dehydrogenase, which participates in cell wall formation. The strains synthesized LPS with a short-chain length, and showed lower cytotoxicity and reduced intracellular proliferation in animal cells compared to wild-type bacteria. After oral infection, the mutants were cleared in immune tissues, including the Peyer's patch, mesenteric lymph node, and spleen, within 5 days. The LD50 of the mutants in Balb/c mice was estimated to be 10(6) higher than wild-type bacteria when administered either via an oral or i.p. route, indicating that the two strains are highly attenuated. To compare the immune response to and protective effects of the mutants against wild-type bacterial infection, we inoculated the mutants into mice via an oral (1x10(10)CFU) or i.p. (1x10(7) CFU) route once or twice at a two week interval. All immune responses, such as serum IgG and secretory IgA levels, cytokine production, and delayed hypersensitivity, were highly induced by two rounds of immunization. HTSaYA and SHJ2104 induced similar immune responses, and mice immunized with HTSaYA or SHJ2104 via an i.p. route were protected against wild-type Salmonella infection even at 100-fold of the LD(50) (5x10(6) CFU). Taken together, these data indicate that HTSaYA and SHJ2104 could be developed as live attenuated Salmonella vaccine candidates.

  15. Subspecies IIIa and IIIb Salmonellae are defective for colonization of murine models of salmonellosis compared to Salmonella enterica subsp. I serovar typhimurium.

    PubMed

    Katribe, Erin; Bogomolnaya, Lydia M; Wingert, Heather; Andrews-Polymenis, Helene

    2009-04-01

    Non-subspecies I salmonellae are commensals of cold-blooded vertebrates and cause sporadic disease in mammals. The reasons why non-subspecies I salmonellae do not circulate in populations of warm-blooded vertebrates, but instead only cause occasional disease in this niche, are unknown. We examined the ability of Salmonella enterica subsp. IIIa (subsp. arizonae) and subsp. IIIb (subsp. diarizonae) isolates to grow competitively with subspecies I (serovar Typhimurium) ATCC 14028 in vitro, to colonize Salmonella-sensitive BALB/c mice, and to persist in the intestine of Salmonella-resistant CBA/J mice in competitive infections. Subspecies IIIa had severely reduced intestinal colonization, intestinal persistence, and systemic spread in mice. Subspecies IIIa is nonmotile on swarming agar and thus may also have reduced motility under viscous conditions in vivo. Surprisingly, subspecies IIIb colonizes the intestinal tract of BALB/c mice normally yet does not spread systemically. Subspecies IIIb colonization of the intestine of CBA/J mice is reduced late in infection. In order to understand why these isolates do not colonize systemic sites, we determined that subspecies IIIa and IIIb are not internalized well and do not replicate in J774-A.1 murine macrophages, despite normal adherence to these cells. We further show that selected effectors of both type III secretion systems 1 and 2 are secreted by subspecies IIIa and IIIb in vitro but that each of these isolates secretes a different combination of effectors. We outline the phenotypic differences between these subspecies and subspecies I and provide a possible explanation for the inability of these strains to spread systemically in murine models.

  16. β-Galactomannan and Saccharomyces cerevisiae var. boulardii modulate the immune response against Salmonella enterica serovar Typhimurium in porcine intestinal epithelial and dendritic cells.

    PubMed

    Badia, Roger; Brufau, M Teresa; Guerrero-Zamora, Ana Maria; Lizardo, Rosil; Dobrescu, Irina; Martin-Venegas, Raquel; Ferrer, Ruth; Salmon, Henri; Martínez, Paz; Brufau, Joaquim

    2012-03-01

    Salmonella enterica serovar Typhimurium is a facultative intracellular pathogen that causes inflammation, necrosis, and diarrhea in pigs, as well as being an important source of food-borne diseases in humans. Probiotics and prebiotics are promising alternatives to antibiotics to control and prevent intestinal infections. The present work investigated a recently developed β-galactomannan (βGM) prebiotic compared to the proven probiotic Saccharomyces cerevisiae var. boulardii on porcine ileum intestinal epithelial cells (IECs) of the IPI-2I line and monocyte-derived dendritic cells (DCs) cocultured in vitro with Salmonella. We observed that both S. cerevisiae var. boulardii and βGM inhibited the association of Salmonella with IECs in vitro. Our data indicated that βGM has a higher ability than S. cerevisiae var. boulardii to inhibit Salmonella-induced proinflammatory mRNA (cytokines tumor necrosis factor alpha [TNF-α], interleukin-1α [IL-1α], IL-6, and granulocyte-macrophage colony-stimulating factor [GM-CSF] and chemokines CCL2, CCL20, and CXCL8) and at protein levels (IL-6 and CXCL8). Additionally, βGM and S. cerevisiae var. boulardii induced some effects on DCs that were not observed on IECs: βGM and S. cerevisiae var. boulardii showed slight upregulation of mRNA for TNF-α, GM-CSF, and CCR7 receptor on porcine monocyte-derived dendritic cells (DCs). Indeed, the addition of βGM or S. cerevisiae var. boulardii on DCs cocultured with Salmonella showed higher gene expression (mRNA) for TNF-α, GM-CSF, and CXCL8 compared to that of the control with Salmonella. In conclusion, the addition of βGM inhibits Salmonella-induced proinflammatory profiles in IECs but may promote DC activation, although associated molecular mechanisms remain to be elucidated.

  17. Peroral Ciprofloxacin Therapy Impairs the Generation of a Protective Immune Response in a Mouse Model for Salmonella enterica Serovar Typhimurium Diarrhea, while Parenteral Ceftriaxone Therapy Does Not

    PubMed Central

    Endt, Kathrin; Maier, Lisa; Käppeli, Rina; Barthel, Manja; Misselwitz, Benjamin; Kremer, Marcus

    2012-01-01

    Nontyphoidal Salmonella (NTS) species cause self-limiting diarrhea and sometimes severe disease. Antibiotic treatment is considered only in severe cases and immune-compromised patients. The beneficial effects of antibiotic therapy and the consequences for adaptive immune responses are not well understood. We used a mouse model for Salmonella diarrhea to assess the effects of per os treatment with ciprofloxacin (15 mg/kg of body weight intragastrically 2 times/day, 5 days) or parenteral ceftriaxone (50 mg/kg intraperitoneally, 5 days), two common drugs used in human patients. The therapeutic and adverse effects were assessed with respect to generation of a protective adaptive immune response, fecal pathogen excretion, and the emergence of nonsymptomatic excreters. In the mouse model, both therapies reduced disease severity and reduced the level of fecal shedding. In line with clinical data, in most animals, a rebound of pathogen gut colonization/fecal shedding was observed 2 to 12 days after the end of the treatment. Yet, levels of pathogen shedding and frequency of appearance of nonsymptomatic excreters did not differ from those for untreated controls. Moreover, mice treated intraperitoneally with ceftriaxone developed an adaptive immunity protecting the mice from enteropathy in wild-type Salmonella enterica serovar Typhimurium challenge infections. In contrast, the mice treated intragastrically with ciprofloxacin were not protected. Thus, antibiotic treatment regimens can disrupt the adaptive immune response, but treatment regimens may be optimized in order to preserve the generation of protective immunity. It might be of interest to determine whether this also pertains to human patients. In this case, the mouse model might be a tool for further mechanistic studies. PMID:22354292

  18. β-Galactomannan and Saccharomyces cerevisiae var. boulardii Modulate the Immune Response against Salmonella enterica Serovar Typhimurium in Porcine Intestinal Epithelial and Dendritic Cells

    PubMed Central

    Brufau, M. Teresa; Guerrero-Zamora, Ana Maria; Lizardo, Rosil; Dobrescu, Irina; Martin-Venegas, Raquel; Ferrer, Ruth; Salmon, Henri; Martínez, Paz

    2012-01-01

    Salmonella enterica serovar Typhimurium is a facultative intracellular pathogen that causes inflammation, necrosis, and diarrhea in pigs, as well as being an important source of food-borne diseases in humans. Probiotics and prebiotics are promising alternatives to antibiotics to control and prevent intestinal infections. The present work investigated a recently developed β-galactomannan (βGM) prebiotic compared to the proven probiotic Saccharomyces cerevisiae var. boulardii on porcine ileum intestinal epithelial cells (IECs) of the IPI-2I line and monocyte-derived dendritic cells (DCs) cocultured in vitro with Salmonella. We observed that both S. cerevisiae var. boulardii and βGM inhibited the association of Salmonella with IECs in vitro. Our data indicated that βGM has a higher ability than S. cerevisiae var. boulardii to inhibit Salmonella-induced proinflammatory mRNA (cytokines tumor necrosis factor alpha [TNF-α], interleukin-1α [IL-1α], IL-6, and granulocyte-macrophage colony-stimulating factor [GM-CSF] and chemokines CCL2, CCL20, and CXCL8) and at protein levels (IL-6 and CXCL8). Additionally, βGM and S. cerevisiae var. boulardii induced some effects on DCs that were not observed on IECs: βGM and S. cerevisiae var. boulardii showed slight upregulation of mRNA for TNF-α, GM-CSF, and CCR7 receptor on porcine monocyte-derived dendritic cells (DCs). Indeed, the addition of βGM or S. cerevisiae var. boulardii on DCs cocultured with Salmonella showed higher gene expression (mRNA) for TNF-α, GM-CSF, and CXCL8 compared to that of the control with Salmonella. In conclusion, the addition of βGM inhibits Salmonella-induced proinflammatory profiles in IECs but may promote DC activation, although associated molecular mechanisms remain to be elucidated. PMID:22301691

  19. Pronounced susceptibility to infection by Salmonella enterica serovar Typhimurium in mice chronically exposed to lead correlates with a shift to Th2-type immune responses

    SciTech Connect

    Fernandez-Cabezudo, Maria J.; Ali, Sumaya A.E.; Ullah, Azim; Hasan, Mohammed Y.; Kosanovic, Melita; Fahim, Mohamed A.; Adem, Abdu; Al-Ramadi, Basel K. . E-mail: ramadi.b@uaeu.ac.ae

    2007-02-01

    Persistent exposure to inorganic lead (Pb) is known to adversely affect the immune system. In the present study, we assessed the effect of chronic Pb exposure on susceptibility to infection by the facultative intracellular pathogen Salmonella enterica serovar Typhimurium. Mice were exposed to 10 mM Pb-acetate in drinking water for {approx} 16 weeks, resulting in a significant level of Pb in the blood (106.2 {+-} 8.9 {mu}g/dl). Pb exposure rendered mice susceptible to Salmonella infection, manifested by increased bacterial burden in target organs and heightened mortality. Flow cytometric analysis of the splenic cellular composition in normal and Pb-exposed mice revealed no gross alteration in the ratios of B and T lymphocytes or myeloid cells. Similarly, the capacity of B and T cells to upregulate the expression of activation antigens in response to mitogenic or inflammatory stimuli was not hindered by Pb exposure. Analysis of the ability of ex vivo-cultured splenocytes to secrete cytokines demonstrated a marked reduction in IFN-{gamma} and IL-12p40 production associated with Pb exposure. In contrast, secretion of IL-4 by splenocytes of Pb-treated mice was 3- to 3.6-fold higher than in normal mice. The increased capacity to produce IL-4 correlated with a shift in the in vivo anti-Salmonella antibody response from the protective IgG2a isotype to the Th2-induced IgG1 isotype. We conclude that chronic exposure to high levels of Pb results in a state of immunodeficiency which is not due to an overt cytotoxic or immunosuppressive mechanism, but rather is largely caused by a shift in immune responsiveness to Th2-type reactions.

  20. Structural Alterations of the Cysteine Desulfurase IscS of Salmonella enterica Serovar Typhimurium Reveal Substrate Specificity of IscS in tRNA Thiolation

    PubMed Central

    Lundgren, Hans K.; Björk, Glenn R.

    2006-01-01

    The cysteine desulfurase IscS in Salmonella enterica serovar Typhimurium is required for the formation of all four thiolated nucleosides in tRNA, which is thought to occur via two principally different biosynthetic pathways. The synthesis of 4-thiouridine (s4U) and 5-methylaminomethyl-2-thiouridine (mnm5s2U) occurs by a transfer of sulfur from IscS via various proteins to the target nucleoside in the tRNA, and no iron-sulfur cluster protein participates, whereas the synthesis of 2-thiocytidine (s2C) and N6-(4-hydroxyisopentenyl)-2-methylthioadenosine (ms2io6A) is dependent on iron-sulfur cluster proteins, whose formation and maintenance depend on IscS. Accordingly, inactivation of IscS should result in decreased synthesis of all thiolated nucleosides. We selected mutants defective either in the synthesis of a thiolated nucleoside (mnm5s2U) specific for the iron-sulfur protein-independent pathway or in the synthesis of a thiolated nucleoside (ms2io6A) specific for the iron-sulfur protein-dependent pathway. Although we found altered forms of IscS that influenced the synthesis of all thiolated nucleosides, consistent with the model, we also found mutants defective in subsets of thiolated nucleosides. Alterations in the C-terminal region of IscS reduced the level of only ms2io6A, suggesting that the synthesis of this nucleoside is especially sensitive to minor aberrations in iron-sulfur cluster transfer activity. Our results suggest that IscS has an intrinsic substrate specificity in how it mediates sulfur mobilization and/or iron-sulfur cluster formation and maintenance required for thiolation of tRNA. PMID:16585765

  1. CorA affects tolerance of Escherichia coli and Salmonella enterica serovar Typhimurium to the lactoperoxidase enzyme system but not to other forms of oxidative stress.

    PubMed

    Sermon, Jan; Wevers, Eva M-R P; Jansen, Leentje; De Spiegeleer, Philipp; Vanoirbeek, Kristof; Aertsen, Abram; Michiels, Chris W

    2005-11-01

    The enzyme lactoperoxidase is part of the innate immune system in vertebrates and owes its antimicrobial activity to the formation of oxidative reaction products from various substrates. In a previous study, we have reported that, with thiocyanate as a substrate, the lactoperoxidase system elicits a distinct stress response in Escherichia coli MG1655. This response is different from but partly overlapping with the stress responses to hydrogen peroxide and to superoxide. In the current work, we constructed knockouts in 10 lactoperoxidase system-inducible genes to investigate their role in the tolerance of E. coli MG1655 to this antimicrobial system. Five mutations resulted in a slightly increased sensitivity, but one mutation (corA) caused hypersensitivity to the lactoperoxidase system. This hypersensitive phenotype was specific to the lactoperoxidase system, since neither the sensitivity to hydrogen peroxide nor to the superoxide generator plumbagin was affected in the corA mutant. Salmonella enterica serovar Typhimurium corA had a similar phenotype. Although corA encodes an Mg2+ transporter and at least three other inducible open reading frames belonged to the Mg2+ regulon, repression of the Mg stimulon by Mg2+ did not change the lactoperoxidase sensitivity of either the wild-type or corA mutant. Prior exposure to 0.3 mM Ni2+, which is also transported by CorA, strongly sensitized MG1655 but not the corA mutant to the lactoperoxidase system. Furthermore, this Ni2+-dependent sensitization was suppressed by the CorA-specific inhibitor Co(III) hexaammine. These results indicate that CorA affects the lactoperoxidase sensitivity of E. coli by modulating the cytoplasmic concentrations of transition metals that enhance the toxicity of the lactoperoxidase system.

  2. Peroral ciprofloxacin therapy impairs the generation of a protective immune response in a mouse model for Salmonella enterica serovar Typhimurium diarrhea, while parenteral ceftriaxone therapy does not.

    PubMed

    Endt, Kathrin; Maier, Lisa; Käppeli, Rina; Barthel, Manja; Misselwitz, Benjamin; Kremer, Marcus; Hardt, Wolf-Dietrich

    2012-05-01

    Nontyphoidal Salmonella (NTS) species cause self-limiting diarrhea and sometimes severe disease. Antibiotic treatment is considered only in severe cases and immune-compromised patients. The beneficial effects of antibiotic therapy and the consequences for adaptive immune responses are not well understood. We used a mouse model for Salmonella diarrhea to assess the effects of per os treatment with ciprofloxacin (15 mg/kg of body weight intragastrically 2 times/day, 5 days) or parenteral ceftriaxone (50 mg/kg intraperitoneally, 5 days), two common drugs used in human patients. The therapeutic and adverse effects were assessed with respect to generation of a protective adaptive immune response, fecal pathogen excretion, and the emergence of nonsymptomatic excreters. In the mouse model, both therapies reduced disease severity and reduced the level of fecal shedding. In line with clinical data, in most animals, a rebound of pathogen gut colonization/fecal shedding was observed 2 to 12 days after the end of the treatment. Yet, levels of pathogen shedding and frequency of appearance of nonsymptomatic excreters did not differ from those for untreated controls. Moreover, mice treated intraperitoneally with ceftriaxone developed an adaptive immunity protecting the mice from enteropathy in wild-type Salmonella enterica serovar Typhimurium challenge infections. In contrast, the mice treated intragastrically with ciprofloxacin were not protected. Thus, antibiotic treatment regimens can disrupt the adaptive immune response, but treatment regimens may be optimized in order to preserve the generation of protective immunity. It might be of interest to determine whether this also pertains to human patients. In this case, the mouse model might be a tool for further mechanistic studies.

  3. Role of Salmonella enterica Serovar Typhimurium Two-Component System PreA/PreB in Modulating PmrA-Regulated Gene Transcription

    PubMed Central

    Merighi, Massimo; Carroll-Portillo, Amanda; Septer, Alecia N.; Bhatiya, Aditi; Gunn, John S.

    2006-01-01

    The PmrA/PmrB two-component system encoded by the pmrCAB operon regulates the modification of Salmonella enterica serovar Typhimurium lipopolysaccharide leading to polymyxin B resistance. PmrA and PhoP are the only known activators of pmrCAB. A transposon mutagenesis screen for additional regulators of a pmrC::MudJ fusion led to the identification of a two-component system, termed PreA/PreB (pmrCAB regulators A and B), that controls the transcription of the pmrCAB operon in response to unknown signals. The initial observations indicated that insertions in, or a deletion of, the preB sensor, but not the preA response regulator, caused upregulation of pmrCAB. Interestingly, the expression of pmrCAB was not upregulated in a preAB mutant grown in LB broth, implicating PreA in the increased expression of pmrCAB in the preB strain. This was confirmed by overexpression of preA+ in preAB or preB backgrounds, which resulted in significant upregulation or further upregulation of pmrCAB. No such effect was observed in any tested preB+ backgrounds. Additionally, an ectopic construct expressing a preA[D51A] allele also failed to upregulate pmrC in any of the pre backgrounds tested, which implies that there is a need for phosphorylation in the activation of the target genes. The observed upregulation of pmrCAB occurred independently of the response regulators PmrA and PhoP. Although a preB mutation led to increased transcription of pmrCAB, this did not result in a measurable effect on polymyxin B resistance. Our genetic data support a model of regulation whereby, in response to unknown signals, the PreB sensor activates PreA, which in turn indirectly upregulates pmrCAB transcription. PMID:16352830

  4. Transcriptome and proteome analysis of Salmonella enterica serovar Typhimurium systemic infection of wild type and immune-deficient mice

    PubMed Central

    Oshota, Olusegun; Fookes, Maria; Schreiber, Fernanda; Chaudhuri, Roy R.; Yu, Lu; Clare, Simon; Choudhary, Jyoti; Thomson, Nicholas R.; Lio, Pietro

    2017-01-01

    Salmonella enterica are a threat to public health. Current vaccines are not fully effective. The ability to grow in infected tissues within phagocytes is required for S. enterica virulence in systemic disease. As the infection progresses the bacteria are exposed to a complex host immune response. Consequently, in order to continue growing in the tissues, S. enterica requires the coordinated regulation of fitness genes. Bacterial gene regulation has so far been investigated largely using exposure to artificial environmental conditions or to in vitro cultured cells, and little information is available on how S. enterica adapts in vivo to sustain cell division and survival. We have studied the transcriptome, proteome and metabolic flux of Salmonella, and the transcriptome of the host during infection of wild type C57BL/6 and immune-deficient gp91-/-phox mice. Our analyses advance the understanding of how S. enterica and the host behaves during infection to a more sophisticated level than has previously been reported. PMID:28796780

  5. Effects of Salmonella enterica serovars Typhimurium (ST) and Choleraesuis (SC) on chemokine and cytokine expression in swine ileum and jejunal epithelial cells.

    PubMed

    Skjolaas, K A; Burkey, T E; Dritz, S S; Minton, J E

    2006-06-15

    The gastrointestinal epithelium represents a barrier to potentially invasive enteric pathogens, maintains a role in innate immune surveillance, and is a source of both chemokine and cytokine chemotactic mediators in response to bacterial invasion. In the current study, we evaluated cytokine and chemokine mediators known to regulate movement of macrophages (macrophage migration inhibitory factor; MIF), neutrophils (IL8), dendritic cells (CCL20), and epithelial remodeling (osteopontin; OPN) in response to invasive swine enteropathogens Salmonella enterica serovar Typhimurium (ST) or Choleraesuis (SC). For the in vivo experiment, weaned pigs served as uninfected controls (0 h) or were given 3 x 10(9) CFU ST orally. Pigs were sacrificed at 8, 24, 48, and 144 h after inoculation and total RNA was extracted from defined segments of proximal (PI) and distal (DI) ileum. Relative expression of MIF and OPN were not affected by ST. IL8 expression was increased numerically (P = 0.17 for the interaction term) at 24 and 144 h in the PI and these increases accounted for greater expression in the PI relative to the DI (P < 0.05). Relative expression of CCL20 was increased at 24 h after ST (P < 0.05). Next, we evaluated the time course of MIF, IL8, CCL20, and OPN mRNA expression induced by application of lipopolysaccharide (LPS), ST or SC in vitro using pig jejunal epithelial cells (IPEC-J2). Cells were grown to confluency on permeable membranes, and treated apically with LPS (10 ng/mL), ST or SC (10(8)/well). After 1 h, cells were washed to remove LPS or extracellular bacteria, and media containing gentamicin was added to kill remaining extracellular bacteria. Media and RNA were collected at 1.5, 3, and 6 h after treatment. MIF mRNA was not affected by LPS or bacterial treatment. Similarly, IL8 expression was not affected by LPS, but was increased by ST and SC relative to controls at 1.5 and 3 h post exposure (P < 0.05 for all comparisons). Treatment with SC increased CCL20 m

  6. Genomic Analysis of Salmonella enterica Serovar Typhimurium Characterizes Strain Diversity for Recent U.S. Salmonellosis Cases and Identifies Mutations Linked to Loss of Fitness under Nitrosative and Oxidative Stress

    PubMed Central

    Hayden, Hillary S.; Matamouros, Susana; Hager, Kyle R.; Brittnacher, Mitchell J.; Rohmer, Laurence; Radey, Matthew C.; Weiss, Eli J.; Kim, Katie B.; Jacobs, Michael A.; Sims-Day, Elizabeth H.; Yue, Min; Zaidi, Mussaret B.; Schifferli, Dieter M.; Manning, Shannon D.; Walson, Judd L.

    2016-01-01

    ABSTRACT Salmonella enterica serovar Typhimurium is one of the most common S. enterica serovars associated with U.S. foodborne outbreaks. S. Typhimurium bacteria isolated from humans exhibit wide-ranging virulence phenotypes in inbred mice, leading to speculation that some strains are more virulent in nature. However, it is unclear whether increased virulence in humans is related to organism characteristics or initial treatment failure due to antibiotic resistance. Strain diversity and genetic factors contributing to differential human pathogenicity remain poorly understood. We reconstructed phylogeny, resolved genetic population structure, determined gene content and nucleotide variants, and conducted targeted phenotyping assays for S. Typhimurium strains collected between 1946 and 2012 from humans and animals in the United States and abroad. Strains from recent U.S. salmonellosis cases were associated with five S. Typhimurium lineages distributed within three phylogenetic clades, which are not restricted by geography, year of acquisition, or host. Notably, two U.S. strains and four Mexican strains are more closely related to strains associated with human immunodeficiency virus (HIV)-infected individuals in sub-Saharan Africa than to other North American strains. Phenotyping studies linked variants specific to these strains in hmpA and katE to loss of fitness under nitrosative and oxidative stress, respectively. These results suggest that U.S. salmonellosis is caused by diverse S. Typhimurium strains circulating worldwide. One lineage has mutations in genes affecting fitness related to innate immune system strategies for fighting pathogens and may be adapting to immunocompromised humans by a reduction in virulence capability, possibly due to a lack of selection for its maintenance as a result of the worldwide HIV epidemic. PMID:26956590

  7. Small outer-membrane lipoprotein, SmpA, is regulated by sigmaE and has a role in cell envelope integrity and virulence of Salmonella enterica serovar Typhimurium.

    PubMed

    Lewis, Claire; Skovierova, Henrieta; Rowley, Gary; Rezuchova, Bronislava; Homerova, Dagmar; Stevenson, Andrew; Sherry, Aileen; Kormanec, Jan; Roberts, Mark

    2008-03-01

    SmpA is a small outer-membrane lipoprotein that is a component of the essential YaeT outer-membrane protein assembly complex. In Salmonella enterica serovar Typhimurium (S. Typhimurium), expression of the smpA gene was shown to be directed by two promoters, smpAp1 and smpAp2. The more distal promoter, smpAp1, is dependent upon the extracytoplasmic stress response sigma factor sigma(E). An smpA null mutant was constructed in S. Typhimurium SL1344 and was shown to be more sensitive than its wild-type parent to growth at high temperature and in the presence of sodium cholate, SDS plus EDTA, and the hydrophobic antibiotic rifampicin. The lack of SmpA in S. Typhimurium elicits a sigma(E)-dependent stress response. These findings are indicative of altered outer-membrane integrity in the smpA mutant, probably due to a defect in outer-membrane protein biogenesis. SmpA was not important for entry or survival within murine macrophages; however, the S. Typhimurium smpA mutant was attenuated in mice by both the oral and parenteral routes of infection, and SmpA appeared to be most important for the growth of S. Typhimurium at systemic sites.

  8. A multi-pronged search for a common structural motif in the secretion signal of Salmonella enterica serovar Typhimurium type III effector proteins

    SciTech Connect

    Buchko, Garry W.; Niemann, George; Baker, Erin Shammel; Belov, Mikhail E.; Smith, Richard D.; Heffron, Fred; Adkins, Joshua N.; McDermott, Jason E.

    2010-11-08

    Many pathogenic Gram-negative bacteria use a type III secretion system (T3SS) to deliver effector proteins into the host cell where they reprogram host defenses and facilitate pathogenesis. While it has been determined that the first 20 - 30 N-terminal residues usually contain the ‘secretion signal’ that targets effector proteins for translocation, the molecular basis for recognition of this signal is not understood. Recent machine-learning approaches, such as SVM-based Identification and Evaluation of Virulence Effectors (SIEVE), have improved the ability to identify effector proteins from genomics sequence information. While these methods all suggest that the T3SS secretion signal has a characteristic amino acid composition bias, it is still unclear if the amino acid pattern is important and if there are any unifying structural properties that direct recognition. To address these issues a peptide corresponding to the secretion signal for Salmonella enterica serovar Typhimurium effector SseJ was synthesized (residues 1-30, SseJ) along with scrambled peptides of the same amino acid composition that produced high (SseJ-H) and low (SseJ-L) SIEVE scores. The secretion properties of these three peptides were tested using a secretion signal-CyaA fusion assay and their structures systematically probed using circular dichroism, nuclear magnetic resonance, and ion mobility spectrometry-mass spectrometry. The signal-CyaA fusion assay showed that the native and SseJ-H fusion constructs were secreted into J774 macrophage at similar levels via the SPI-2 secretion pathway while secretion of the SseJ-L fusion construct was substantially retarded, suggesting that the SseJ secretion signal was sequence order dependent. The structural studies showed that the SseJ, SseJ-H, and SseJ-L peptides were intrinsically disordered in aqueous solution with only a small predisposition to adopt nascent helical structure in the presence of the powerful structure stabilizing agent, 1

  9. Effect of chlorate, molybdate, and shikimic acid on Salmonella enterica serovar Typhimurium in aerobic and anaerobic cultures

    USDA-ARS?s Scientific Manuscript database

    Chlorate is a bactericide that has potential as a pre-slaughter feed additive to improve food safety of meat products. The aims of the present study were to examine the effects of chlorate (5mM), molybdate (1 mM), and shikimate (0.34 mM) on the growth and chlorate-resistance of Salmonella enterica ...

  10. Dose-dependent effects on survival of Salmonella enterica serovar Typhimurium in house flies (Musca domestica L.)

    USDA-ARS?s Scientific Manuscript database

    Adult house flies ingest variable numbers of bacteria when they encounter microbe-rich substrates. Bacterial abundance may affect survival within the fly gut, which subsequently impacts vector potential. This study investigated the dose-dependent survival of GFP-expressing Salmonella enterica serova...

  11. Shifts from glucose to certain secondary carbon-sources result in activation of the extracytoplasmic function sigma factor sigmaE in Salmonella enterica serovar Typhimurium.

    PubMed

    Kenyon, William J; Thomas, Sheena M; Johnson, Erin; Pallen, Mark J; Spector, Michael P

    2005-07-01

    Salmonella enterica serovar Typhimurium (S. Typhimurium) elicits the starvation-stress response (SSR) due to starvation for an essential nutrient, e.g. a carbon/energy source (C-source). As part of the SSR, the alternative sigma factor sigma(E) is activated and induced. The authors suspect that this activation is, in part, triggered by changes in the S. Typhimurium cell envelope occurring during the adaptation from growth to carbon/energy starvation (C-starvation), and resulting in an increased need for sigma(E)-regulated factors involved in the proper folding and assembly of newly synthesized proteins destined for this extracytoplasmic compartment. This led to the hypothesis that a sigma(E) activation signal might arise during C-source shifts that cause the induction of proteins localized to the extracytoplasmic compartment, i.e. the outer membrane or periplasm, of the cell. To test this hypothesis, cultures were grown in minimal medium containing enough glucose to reach mid-exponential-phase, plus a non-limiting amount of a secondary 'less-preferred' but utilizable carbon/energy source. The sigma(E) activity was then monitored using plasmids carrying rpoEP1- and rpoEP2-lacZ transcriptional fusions, which exhibit sigma(E)-independent and -dependent lacZ expression, respectively. The secondary C-sources maltose, succinate and citrate, which have extracytoplasmic components involved in their utilization (e.g. LamB), resulted in a discernible diauxic lag period and a sustained increase in sigma(E) activity. Growth transition from glucose to other utilizable phosphotransferase (PTS) and non-PTS C-sources, such as trehalose, mannose, mannitol, fructose, glycerol, d-galactose or l-arabinose, did not cause a discernible diauxic lag period or a sustained increase in sigma(E) activity. Interestingly, a shift from glucose to melibiose, which does not use an extracytoplasmic-localized protein for uptake, did cause an observable diauxic lag period but did not result in a

  12. The agricultural antibiotic carbadox induces generalized transducing phage in multidrug-resistant Salmonella enterica serovar Typhimurium DT104

    USDA-ARS?s Scientific Manuscript database

    Non-typhoidal Salmonella, a leading cause of U.S. foodborne disease and food-related deaths, often asymptomatically colonizes food-producing animals. In fact, >50% of U.S. swine production facilities test positive for Salmonella. The multidrug-resistant (MDR) Salmonella Typhimurium DT104 NCTC13348 c...

  13. Effect of zinc on growth performance, gut morphometry, and cecal microbial community in broilers challenged with Salmonella enterica serovar typhimurium.

    PubMed

    Shao, Yuxin; Lei, Zhao; Yuan, Jianmin; Yang, Ying; Guo, Yuming; Zhang, Bingkun

    2014-12-01

    To evaluate the effects of supplemental zinc on growth performance, gut morphometry, and the cecal microbial community in broilers challenged with Salmonella typhimurium, 180, 1-day-old male Cobb 500 broiler chicks were randomly assigned to 3 treatments with ten replicates for a 42 day experiment. The 3 treatments were: unchallenged, S. typhimurium-challenged, and S. typhimurium-challenged with 120 mg/kg of zinc supplementation in the diet. Salmonella infection caused a reduction in body-weight gain and feed intake, disrupted the intestinal structure by decreasing the villus-height/crypt-depth ratio of the ileum and increasing the apoptotic index of ileal epithelial cells. Moreover, the cecal microbial community was altered by Salmonella infection, as demonstrated by a reduced number of Lactobacillus and total bacteria. Dietary zinc supplementation improved growth performance by increasing the body-weight gain and feed intake in the challenged broilers. In addition, zinc repaired intestinal injury by reducing the apoptotic index of ileal epithelial cells, enhancing villus height and the villus-height/crypt-depth ratio of the ileum, and the proliferation index of ileal epithelial cells. Finally, zinc regulated the cecal microbial community by increasing the number of total bacteria and beneficial Lactobacillus bacteria, and reducing the number of Salmonella. The results indicated that dietary zinc supplementation improved growth performance, intestinal morphology, and intestinal microbiota in S. typhimurium-challenged broilers.

  14. Minimal effects of high-pressure treatment on Salmonella enterica serovar Typhimurium inoculated into peanut butter and peanut products.

    PubMed

    Grasso, Elizabeth M; Somerville, Jeremy A; Balasubramaniam, V M; Lee, Ken

    2010-10-01

    About 1.2 billion pounds of peanut butter are consumed annually in the United States. In 2008 to 2009, an outbreak involving Salmonella Typhimurium in peanut butter led to a recall of over 3900 products by over 200 companies. More than 700 people became sick, 100 were hospitalized, and 9 people died from this outbreak. This study examines the efficacy of high-pressure processing (HPP) to decrease S. Typhimurium American Type Culture Collection (ATCC) 53647 inoculated into peanut butter and model systems. The viability of S. Typhimurium in peanut butter stored at room temperature was investigated. A culture of S. Typhimurium (6.88 log CFU/g) was inoculated into peanut butter. Following 28 d at 20 °C there was a 1.23-log reduction. Approximately 10(6) to 10(7) CFU/g S. Typhimurium were inoculated into 4 brands of peanut butter, 3 natural peanut butters and peanut flour slurries at 2, 5, and 10% peanut flour protein in peanut oil and in distilled water. All were treated at 600 MPa for 5 min at 45 °C. While significant differences were found between natural peanut butter and peanut protein mixtures, the reduction was <1.0 log. The peanut flour/oil mixtures had a 1.7, 1.6, and 1.0-log reduction from HPP (2, 5, and 10% protein, respectively) whereas peanut flour/water mixtures had a 6.7-log reduction for all protein levels. Oil had a protective effect indicating HPP may not help the microbial safety of water-in-oil food emulsions including peanut butter. Practical Application: There have been multiple outbreaks of foodborne illness involving peanut butter products. This study looks at the potential use of high-pressure processing to reduce the bacteria that may be in peanut butter.

  15. Intestinal invasion of Salmonella enterica serovar Typhimurium in the avian host is dose dependent and does not depend on motility and chemotaxis.

    PubMed

    Olsen, John Elmerdahl; Hoegh-Andersen, Kirsten Hobolt; Rosenkrantz, Jesper Tjørnholt; Schroll, Casper; Casadesús, Josep; Aabo, Søren; Christensen, Jens Peter

    2013-08-30

    Salmonella enterica serotype Typhimurium (S. Typhimurium) can invade in the intestine of the avian host, and knowledge on the mechanisms that govern this is potentially important for prevention of disease. This study investigated the invasion of S. Typhimurium in the avian host and to which extent it depended on motility and chemotaxis. Wild type and previously well-characterized transposon mutants in flagella genes fliC and fljB and in chemotaxis genes cheA, cheB and cheR were used as challenge strains in intestinal loop experiments. Invasion was shown to be dose dependent, but did not require functional flagella or chemotaxis genes. In support of the results from intestinal loop experiments, flagella and chemotaxis genes were not significantly important to the outcome of an oral infection. The results showed that S. Typhimurium invasion in the avian host was dose dependent and was not affected by the loss of flagella and chemotaxis genes.

  16. RNA-seq-based analysis of drug-resistant Salmonella enterica serovar Typhimurium selected in vivo and in vitro.

    PubMed

    Li, Lin; Dai, Xingyang; Wang, Ying; Yang, Yanfei; Zhao, Xia; Wang, Lei; Zeng, Minghua

    2017-01-01

    The aim of this study was to characterize the mechanism of fluoroquinolone (FQ) resistance in Salmonella Typhimurium. We established the Caenorhabditis elegans-Salmonella Typhimurium model to select for ciprofloxacin resistance in Salmonella Typhimurium colonizing C. elegans, generating the resistant strains TN4. Gradient doses of ciprofloxacin were used to generate the resistant strain TW4 in vitro. RNA sequencing was used to establish the whole-transcriptome profile of three strains of Salmonella Typhimurium. The gene expression patterns of resistant strains TN4 and TW4 differed from those of the parental strain. In TN4, 2,277 genes were differentially expressed (1,833 upregulated and 444 downregulated) relative to the parental strain, and in TW4, 3,464 genes were differentially expressed (3,433 upregulated and 31 downregulated). Among these differentially expressed genes, 28 were associated with drug resistance and 26 were associated with the two-component systems in the two resistant strains. Seven different pathways were significantly sffected in two strains. Efflux pump overexpression was identified as one of the main mechanisms underlying FQ resistance in the two resistant strains. TW4 differentially expressed more efflux pump genes than TN4 and most of these genes were more strongly expressed than in TN4. However, expression of the efflux pump repressor gene and the mar operon was downregulated in TN4 but not in TW4. Two-component systems are also important in drug resistance. Our findings provide an important basis for further studies of the complex network that regulate FQ resistance in Salmonella.

  17. Dam methylation regulates the expression of SPI-5-encoded sopB gene in Salmonella enterica serovar Typhimurium.

    PubMed

    Giacomodonato, Mónica N; Llana, Mariángeles Noto; Castañeda, María del Rosario Aya; Buzzola, Fernanda; García, Mauro D; Calderón, Marina Gallo; Sarnacki, Sebastián H; Cerquetti, María C

    2014-08-01

    DNA adenine methylation is an essential factor in Salmonella virulence. Here, we investigate the involvement of DNA adenine methylase (Dam) in the expression and translocation of a SPI-5-encoded effector of S. Typhimurium. SopB expression and secretion were determined using SopB-FLAG-tagged wild type and dam strains of S. Typhimurium. Western blot and quantitative reverse transcriptase PCR analysis showed that the dam mutant expresses lower levels of SopB protein and sopB mRNA than the wild type strain under SPI-1 and SPI-2 inducing conditions in vitro. SopB secretion was also considerably impaired in the absence of dam. In agreement with in vitro experiments, SopB synthesis in dam mutants recovered from infected epithelial cells and from murine mesenteric lymph nodes was reduced by 40% respect to the wild type strain (p < 0.05). SopB translocation was neither detected in the cytosol of epithelial cells nor in the cytosol of cells isolated from mesenteric lymph nodes infected with the dam mutant. Taken together, our results demonstrate that, in S. Typhimurium, Dam methylation modulates the expression and translocation of SPI-5-encoded SopB effector.

  18. Improvement of bacterial clearance and relief of clinical signs of Salmonella enterica serovar Typhimurium infection in pigs through upregulation of Th 1-specific responses by administration of a combination of two silicate minerals, biotite and bentonite

    PubMed Central

    LEE, Jin-A; JUNG, Bock-Gie; KIM, Tae-Hoon; KIM, Yun-Mi; KOH, Hong-Bum; LEE, Bong-Joo

    2015-01-01

    Biotite and bentonite are phyllosilicate minerals that were originally used in industrial applications. Several beneficial activities of them have recently been reported, especially regulation of the immune system and antimicrobial effects. Therefore, we investigated the immune-enhancing and bacterial clearance effects of a biotite and bentonite mixture (BBM) on experimental infection of Salmonella enterica serovar Typhimurium (S. Typhimurium) to determine whether the BBM could be used as an alternative antibiotic. We administered 1% or 2% BBM as a feed supplement. We then evaluated the bacterial clearance effects of the BBM against S. Typhimurium. We also evaluated the immune-enhancing effect of the BBM through several immunological experiments that included examination of the lysozyme activity, CD4+/CD8+ T lymphocyte ratio and the T-helper type 1 (Th 1) cytokine profile. The clinical signs of S. Typhimurium and the number of viable bacteria in feces and tissues were significantly decreased in both BBM groups, especially in the 2% BBM group. The BBM also markedly enhanced the lysozyme activity, CD4+/CD8+ T lymphocyte ratio and expression levels of IFN-γ and IL-12 in S. Typhimurium-challenged pigs. Therefore, the BBM could be a good candidate as an alternative antibiotic that improves Th 1-specific immune responses and the bacterial clearance effect. PMID:25947887

  19. Improvement of bacterial clearance and relief of clinical signs of Salmonella enterica serovar Typhimurium infection in pigs through upregulation of Th 1-specific responses by administration of a combination of two silicate minerals, biotite and bentonite.

    PubMed

    Lee, Jin-A; Jung, Bock-Gie; Kim, Tae-Hoon; Kim, Yun-Mi; Koh, Hong-Bum; Lee, Bong-Joo

    2015-09-01

    Biotite and bentonite are phyllosilicate minerals that were originally used in industrial applications. Several beneficial activities of them have recently been reported, especially regulation of the immune system and antimicrobial effects. Therefore, we investigated the immune-enhancing and bacterial clearance effects of a biotite and bentonite mixture (BBM) on experimental infection of Salmonella enterica serovar Typhimurium (S. Typhimurium) to determine whether the BBM could be used as an alternative antibiotic. We administered 1% or 2% BBM as a feed supplement. We then evaluated the bacterial clearance effects of the BBM against S. Typhimurium. We also evaluated the immune-enhancing effect of the BBM through several immunological experiments that included examination of the lysozyme activity, CD4(+)/CD8(+) T lymphocyte ratio and the T-helper type 1 (Th 1) cytokine profile. The clinical signs of S. Typhimurium and the number of viable bacteria in feces and tissues were significantly decreased in both BBM groups, especially in the 2% BBM group. The BBM also markedly enhanced the lysozyme activity, CD4(+)/CD8(+) T lymphocyte ratio and expression levels of IFN-γ and IL-12 in S. Typhimurium-challenged pigs. Therefore, the BBM could be a good candidate as an alternative antibiotic that improves Th 1-specific immune responses and the bacterial clearance effect.

  20. Influence of Salmonella enterica serovar Typhimurium infection on intestinal goblet cells and villous morphology in broiler chicks.

    PubMed

    Fasina, Y O; Hoerr, F J; McKee, S R; Conner, D E

    2010-06-01

    Live broiler chickens are important in the transmission of Salmonella to humans. Reducing Salmonella levels in the intestine of broiler chickens, in part, requires understanding of the interactions between Salmonella and the intestinal barriers that represent the first line of defense. Such barriers include the mucus layer (composed of mucins secreted by goblet cells) and the underlying epithelium. Two experiments were conducted to evaluate the effect of Salmonella Typhimurium infection on intestinal goblet cell dynamics (density and size) and villous morphology in broiler chicks. In Experiment 1, broiler chicks were either challenged with sterile media (control treatment) or orally given 7.4 x 10(7) colony-forming units (CFU) at 3 days of age (termed the CST treatment). Treatments were similar in Experiment 2, except that chicks in the CST treatment were challenged with 7.8 x 10(6) CFU at 4 days of age. Duration of each experiment was 14 days. At 7 days postchallenge (PC) in Experiment 1, jejunal tissue sections were collected, formalin-fixed, and routinely processed for histologic measurement of villous morphometric indices. In Experiment 2, at 10 days PC, jejunal tissue sections were collected and processed for histologic determination of goblet cell numbers and size, in addition to villous measurements. Results showed that Salmonella Typhimurium infection increased goblet cell density, reduced villous surface area, increased the incidence of epithelial exfoliation, and increased the incidence of heterophil influx into the lamina propria (P < 0.05). It was concluded that Salmonella Typhimurium infection impacts goblet cell biology and exerts morphopathologic changes in the jejunum of broiler chicks.

  1. Novel synergistic approach to exploit the bactericidal efficacy of commercial disinfectants on the biofilms of Salmonella enterica serovar Typhimurium.

    PubMed

    Singla, Richu; Goel, Honey; Ganguli, Abhijit

    2014-07-01

    Combined effect of malic acid and ozone as sanitizer to inhibit the biofilm formation by Salmonella typhimurium on different food contact surfaces was investigated in this study. Different surfaces used in food industry including PVC pipes, polyethylene bags, plastic surfaces and fresh produce were analyzed for the biofilm formation by S. typhimurium ST1 and ST2. Malic acid alone was not able to inhibit biofilm formation in all the samples. However, combination of malic acid with ozone reduced the biofilm formation on plastic bags as well as on PVC pipes suggesting as an effective disinfectant for food contact surfaces. Five- and six-fold reduction in biofilm formation was observed in microtitre plates after 20 h and 40 h. Scanning electron micrographs of carrot and turnip showed control over the biofilms. Malic acid as sanitizer in food industry was effective for the complete inhibition of biofilm in carrot and other food contact surfaces, besides this, combined sanitizer (malic acid and ozone) was effective in turnip. Biofilms in food-processing industries can survive even after the sanitizer treatment and may represent reservoirs of product contamination leading to subsequent spoilage and/or food safety risks.

  2. Complex Function for SicA, a Salmonella enterica Serovar Typhimurium Type III Secretion-Associated Chaperone

    PubMed Central

    Tucker, Stephanie C.; Galán, Jorge E.

    2000-01-01

    Salmonella enterica encodes a type III secretion system within a pathogenicity island located at centisome 63 that is essential for virulence. All type III secretion systems require the function of a family of low-molecular-weight proteins that aid the secretion process by acting as partitioning factors and/or secretion pilots. One such protein is SicA, which is encoded immediately upstream of the type III secreted proteins SipB and SipC. We found that the absence of SicA results in the degradation of both SipB and SipC. Interestingly, in the absence of SipC, SipB was not only stable but also secreted at wild-type levels in a sicA mutant background, indicating that SicA is not required for SipB secretion. We also found that SicA is capable of binding both SipB and SipC. These results are consistent with a SicA role as a partitioning factor for SipB and SipC, thereby preventing their premature association and degradation. We also found that introduction of a sicA null mutation results in the lack of expression of SopE, another type III-secreted protein. Such an effect was shown to be transcriptional. Introduction of a loss-of-function sipC mutation into the sicA mutant background rescued sopE expression. These results indicate that the effect of sicA on sopE expression is indirect and most likely exerted through a regulatory factor(s) partitioned by SicA from SipC. These studies therefore describe a surprisingly complex function for the Salmonella enterica type III secretion-associated chaperone SicA. PMID:10735870

  3. Improving resolution of public health surveillance for human Salmonella enterica serovar Typhimurium infection: 3 years of prospective multiple-locus variable-number tandem-repeat analysis (MLVA)

    PubMed Central

    2012-01-01

    Background Prospective typing of Salmonella enterica serovar Typhimurium (STM) by multiple-locus variable-number tandem-repeat analysis (MLVA) can assist in identifying clusters of STM cases that might otherwise have gone unrecognised, as well as sources of sporadic and outbreak cases. This paper describes the dynamics of human STM infection in a prospective study of STM MLVA typing for public health surveillance. Methods During a three-year period between August 2007 and September 2010 all confirmed STM isolates were fingerprinted using MLVA as part of the New South Wales (NSW) state public health surveillance program. Results A total of 4,920 STM isolates were typed and a subset of 4,377 human isolates was included in the analysis. The STM spectrum was dominated by a small number of phage types, including DT170 (44.6% of all isolates), DT135 (13.9%), DT9 (10.8%), DT44 (4.5%) and DT126 (4.5%). There was a difference in the discriminatory power of MLVA types within endemic phage types: Simpson's index of diversity ranged from 0.109 and 0.113 for DTs 9 and 135 to 0.172 and 0.269 for DTs 170 and 44, respectively. 66 distinct STM clusters were observed ranging in size from 5 to 180 cases and in duration from 4 weeks to 25 weeks. 43 clusters had novel MLVA types and 23 represented recurrences of previously recorded MLVA types. The diversity of the STM population remained relatively constant over time. The gradual increase in the number of STM cases during the study was not related to significant changes in the number of clusters or their size. 667 different MLVA types or patterns were observed. Conclusions Prospective MLVA typing of STM allows the detection of community outbreaks and demonstrates the sustained level of STM diversity that accompanies the increasing incidence of human STM infections. The monitoring of novel and persistent MLVA types offers a new benchmark for STM surveillance. A part of this study was presented at the MEEGID × (Molecular Epidemiology

  4. Effects of dietary clays on performance and intestinal mucus barrier of broiler chicks challenged with Salmonella enterica serovar Typhimurium and on goblet cell function in vitro.

    PubMed

    Almeida, J A S; Ponnuraj, N P; Lee, J J; Utterback, P; Gaskins, H R; Dilger, R N; Pettigrew, J E

    2014-04-01

    In vivo and in vitro experiments were conducted to test for beneficial effects of dietary clays on broiler chicks challenged with Salmonella enterica serovar Typhimurium and to explore potential mechanisms. First, two hundred forty 1-d-old male broilers (initial BW: 41.6 ± 0.4 g) were allotted in a 2 × 4 factorial arrangement in a randomized complete block design. There were 2 infection treatments (with or without Salmonella) and 4 diets: basal (BAS), 0.3% smectite A (SMA), 0.3% smectite B, and 0.3% zeolite. The Salmonella reduced (P < 0.05) the growth rate of chicks fed the BAS, and feeding clay largely restored it (challenge × diet interaction, P < 0.05). Goblet cell number and size were increased (P < 0.05) by Salmonella in chicks fed the BAS and were reduced (P < 0.05) in Salmonella-challenged chicks by feeding SMA. Villus height was reduced by the Salmonella challenge in the chicks fed dietary clays (P < 0.01) but not in chicks fed the BAS (interaction P < 0.05). A human adenocarcinoma cell line (LS174T) was cultured in vitro in 3 separate experiments in the absence or presence of 3 concentrations (0.05, 0.10, and 0.50%) of SMA. Expression of mucin 2 (MUC2), resistin-like molecule β (RELMß), and trefoil factor 3 (TFF3) were determined by real-time reverse-transcription PCR. The expression of RELMβ was increased and expression of MUC2 was reduced (P < 0.05) by 0.10% SMA. Also, LS174T cells were cultured without or with SMA (0.05 and 0.10%) and the medium and cell lysate were analyzed for RELMβ using an immunoblot assay. Protein expression of RELMß in the cell lysate was reduced (P < 0.05) by SMA addition but increased in the medium, indicating that SMA increased secretion of RELMß, thus depleting the cell and concentrating this protein in the medium. In conclusion, the dietary clays restored the growth depression caused by Salmonella, and changes in goblet cell function may contribute to the benefits of one of the clays, specifically SMA.

  5. Curcumin Reduces the Motility of Salmonella enterica Serovar Typhimurium by Binding to the Flagella, Thereby Leading to Flagellar Fragility and Shedding

    PubMed Central

    Balakrishnan, Arjun; Negi, Vidya Devi; Sakorey, Deepika; Chandra, Nagasuma

    2016-01-01

    ABSTRACT One of the important virulence properties of the pathogen is its ability to travel to a favorable environment, cross the viscous mucus barrier (intestinal barrier for enteric pathogens), and reach the epithelia to initiate pathogenesis with the help of an appendage, like flagella. Nonetheless, flagella can act as an “Achilles heel,” revealing the pathogen's presence to the host through the stimulation of innate and adaptive immune responses. We assessed whether curcumin, a dietary polyphenol, could alter the motility of Salmonella, a foodborne pathogen. It reduced the motility of Salmonella enterica serovar Typhimurium by shortening the length of the flagellar filament (from ∼8 μm to ∼5 μm) and decreasing its density (4 or 5 flagella/bacterium instead of 8 or 9 flagella/bacterium). Upon curcumin treatment, the percentage of flagellated bacteria declined from ∼84% to 59%. However, no change was detected in the expression of the flagellin gene and protein. A fluorescence binding assay demonstrated binding of curcumin to the flagellar filament. This might make the filament fragile, breaking it into smaller fragments. Computational analysis predicted the binding of curcumin, its analogues, and its degraded products to a flagellin molecule at an interface between domains D1 and D2. Site-directed mutagenesis and a fluorescence binding assay confirmed the binding of curcumin to flagellin at residues ASN120, ASP123, ASN163, SER164, ASN173, and GLN175. IMPORTANCE This work, to our knowledge the first report of its kind, examines how curcumin targets flagellar density and affects the pathogenesis of bacteria. We found that curcumin does not affect any of the flagellar synthesis genes. Instead, it binds to the flagellum and makes it fragile. It increases the torsional stress on the flagellar filament that then breaks, leaving fewer flagella around the bacteria. Flagella, which are crucial ligands for Toll-like receptor 5, are some of the most important

  6. Reduction of Salmonella enterica serovar typhimurium DT104 infection in experimentally challenged weaned pigs fed a lactobacillus-fermented feed.

    PubMed

    Yin, Fugui; Farzan, Abdolvahab; Wang, Qi Chuck; Yu, Hai; Yin, Yulong; Hou, Yongqing; Friendship, Robert; Gong, Joshua

    2014-08-01

    Salmonella Typhimurium is a foodborne pathogen and commonly present on pig farms. Probiotics have shown potential as a means of reducing Salmonella shedding in pigs. Three experimental challenge trials were conducted to investigate the potential application of newly isolated Lactobacillus isolates for controlling Salmonella infection in pigs. In each trial, 16 Yorkshire piglets (28-d old) were randomly allocated to 1 of 4 dietary treatments: (1) basal diet (BD), (2) naturally fermented (NF) feed, (3) Lactobacillus zeae-fermented (LZ-F) feed, and 4) Lactobacillus casei-fermented (LC-F) feed. All pigs consumed their assigned diets for 3 d prior to the challenge of Salmonella Typhimurium DT104 (approximately 6 log colony-forming units/pig) through gavage. Pediococcus pentosaceus, L. zeae, and L. casei were most abundant in NF, LZ-F, and LC-F feed, respectively. After the challenge, pigs on fermented feed had lower rectal temperature, diarrhea scores, serum haptoglobin concentrations, and intestinal Salmonella counts than the control group (BD) (p ≤ 0.01). Salmonella spp. were detected in both ileocecal lymph nodes (ICLN) and spleens from all pigs on BD, NF, and LC-F, but only 50% of spleens from pigs on LZ-F. Pigs had a dynamic spatial and temporal immune response to Salmonella infection and dietary treatments, as indicated by up- and downregulation in gene expression of inflammatory cytokines (interleukin (IL)-1β, IL-6, IL-10, interferon-γ, and tumor necrosis factor) in the ileum, ICLN, and spleen. The alternation in cytokine expression by fermented feed, particularly LZ-F, appeared to benefit pigs in combating Salmonella infection.

  7. An enteric pathogen Salmonella enterica serovar Typhimurium suppresses tumor growth by downregulating CD44high and CD4T regulatory (Treg) cell expression in mice: the critical role of lipopolysaccharide and Braun lipoprotein in modulating tumor growth.

    PubMed

    Liu, T; Chopra, A K

    2010-02-01

    An antitumor activity associated with several bacterial pathogens, including Salmonella enterica serovar Typhimurium, has been reported; however, the underlying immunological mechanism(s) that lead to an antitumor effect are currently unclear. Furthermore, such pathogens cannot be used to suppress tumor growth because of their potential for causing sepsis. Recently, we reported the characterization of S. Typhimurium isogenic mutants from which Braun lipoprotein genes (lppA and B) and the multicopy repressor of high temperature requirement (msbB) gene were deleted. In a mouse infection model, two mutants, namely, lppB/msbB and lppAB/msbB, minimally induced proinflammatory cytokine production at high doses and were nonlethal to animals. We showed that immunization of mice with these mutants, followed by challenge with the wild-type S. Typhimurium, could significantly suppress tumor growth, as evidenced by an 88% regression in tumor size in lppB/msbB mutant-immunized animals over a 24-day period. However, the lppAB/msbB mutant alone was not effective in modulating tumor growth in mice, although the lppB/msbB mutant alone caused marginal regression in tumor size. Importantly, we showed that CD44(+) cells grew much faster than CD44(-) cells from human liver tumors in mice, leading us to examine the possibility that S. Typhimurium might downregulate CD44 in tumors and splenocytes of mice. Consequently, we found in S. Typhimurium-infected mice that tumor size regression could indeed be related to the downregulation of CD44(high) and CD4(+)CD25(+) T(reg) cells. Importantly, the role of lipopolysaccharide and Braun lipoprotein was critical in S. Typhimurium-induced antitumor immune responses. Taken together, we have defined new immune mechanisms leading to tumor suppression in mice by S. Typhimurium.

  8. Evidence of Metabolic Switching and Implications for Food Safety from the Phenome(s) of Salmonella enterica Serovar Typhimurium DT104 Cultured at Selected Points across the Pork Production Food Chain

    PubMed Central

    Martins, Marta; McCusker, Matthew P.; McCabe, Evonne M.; O'Leary, Denis; Duffy, Geraldine

    2013-01-01

    Salmonella enterica serovar Typhimurium DT104 is a recognized food-borne pathogen that displays a multidrug-resistant phenotype and that is associated with systemic infections. At one extreme of the food chain, this bacterium can infect humans, limiting the treatment options available and thereby contributing to increased morbidity and mortality. Although the antibiotic resistance profile is well defined, little is known about other phenotypes that may be expressed by this pathogen at key points across the pork production food chain. In this study, 172 Salmonella enterica serovar Typhimurium DT104/DT104b isolated from an extensive “farm-to-fork” surveillance study, focusing on the pork food chain, were characterized in detail. Isolates were cultured from environmental, processing, retail, and clinical sources, and the study focused on phenotypes that may have contributed to persistence/survival in these different niches. Molecular subtypes, along with antibiotic resistance profiles, tolerance to biocides, motility, and biofilm formation, were determined. As a basis for human infection, acid survival and the ability to utilize a range of energy sources and to adhere to and/or invade Caco-2 cells were also studied. Comparative alterations to biocide tolerance were observed in isolates from retail. l-Tartaric acid and d-mannose-1-phosphate induced the formation of biofilms in a preselected subset of strains, independent of their origin. All clinical isolates were motile and demonstrated an enhanced ability to survive in acidic conditions. Our data report on a diverse phenotype, expressed by S. Typhimurium isolates cultured from the pork production food chain. Extending our understanding of the means by which this pathogen adapts to environmental niches along the “farm-to-fork” continuum will facilitate the protection of vulnerable consumers through targeted improvements in food safety measures. PMID:23770904

  9. Evidence of metabolic switching and implications for food safety from the phenome(s) of Salmonella enterica serovar Typhimurium DT104 cultured at selected points across the pork production food chain.

    PubMed

    Martins, Marta; McCusker, Matthew P; McCabe, Evonne M; O'Leary, Denis; Duffy, Geraldine; Fanning, Séamus

    2013-09-01

    Salmonella enterica serovar Typhimurium DT104 is a recognized food-borne pathogen that displays a multidrug-resistant phenotype and that is associated with systemic infections. At one extreme of the food chain, this bacterium can infect humans, limiting the treatment options available and thereby contributing to increased morbidity and mortality. Although the antibiotic resistance profile is well defined, little is known about other phenotypes that may be expressed by this pathogen at key points across the pork production food chain. In this study, 172 Salmonella enterica serovar Typhimurium DT104/DT104b isolated from an extensive "farm-to-fork" surveillance study, focusing on the pork food chain, were characterized in detail. Isolates were cultured from environmental, processing, retail, and clinical sources, and the study focused on phenotypes that may have contributed to persistence/survival in these different niches. Molecular subtypes, along with antibiotic resistance profiles, tolerance to biocides, motility, and biofilm formation, were determined. As a basis for human infection, acid survival and the ability to utilize a range of energy sources and to adhere to and/or invade Caco-2 cells were also studied. Comparative alterations to biocide tolerance were observed in isolates from retail. l-Tartaric acid and d-mannose-1-phosphate induced the formation of biofilms in a preselected subset of strains, independent of their origin. All clinical isolates were motile and demonstrated an enhanced ability to survive in acidic conditions. Our data report on a diverse phenotype, expressed by S. Typhimurium isolates cultured from the pork production food chain. Extending our understanding of the means by which this pathogen adapts to environmental niches along the "farm-to-fork" continuum will facilitate the protection of vulnerable consumers through targeted improvements in food safety measures.

  10. The effect of electron beam irradiation on the survival of Salmonella enterica serovar typhimurium and psychrotrophic bacteria on raw chicken breasts stored at four degrees celsius for fourteen days.

    PubMed

    Sarjeant, K C; Williams, S K; Hinton, A

    2005-06-01

    The effect of high-energy electron beam irradiation on the survival of Salmonella enterica serovar Typhimurium and psychrotrophic bacteria on commercial chicken breast meat was evaluated. Fresh chicken breast meat was purchased from a local poultry processor, inoculated with 8 log10 cfu/mL Salmonella, packaged in Styrofoam trays and over wrapped with a polyvinyl chloride film, and subjected to 0, 1, 2, or 3 kGy of irradiation. The packaged samples were stored at 4 degrees C and analyzed for Salmonella Typhimurium and psychrotrophic organisms at 0, 2, 4, 6, 8, 10, 12, and 14 d of storage. Direct plating and enrichment methods were used for S. Typhimurium analyses. The direct plating method revealed a 4 log reduction in Salmonella for chicken breasts inoculated and treated with 1, 2, or 3 kGy of irradiation. Psychrotrophic counts were conducted at 7 degrees C for 10 d and 25 degrees C for 5 d to determine the effect of incubation methods on the recovery of psychrotrophic organisms. The enrichment method resulted in the repair of injured Salmonella cells and an elevated Salmonella Typhimurium count for all irradiation dosages when compared with data reported for the direct plating method. In general, psychrotrophic counts increased as storage time increased. However, psychrotrophic counts decreased (P < 0.05) as the irradiation dosage increased.

  11. Transcriptional response of selected genes of Salmonella enterica serovar Typhimurium biofilm cells during inactivation by superheated steam.

    PubMed

    Ban, Ga-Hee; Kang, Dong-Hyun; Yoon, Hyunjin

    2015-01-02

    Superheated steam (SHS), produced by the addition of heat to saturated steam (SS) at the same pressure, has great advantages over conventional heat sterilization due to its high temperature and accelerated drying rate. We previously demonstrated that treatment with SHS at 200°C for 10 sec inactivated Escherichia coli O157:H7, Salmonella Typhimurium, and Listeria monocytogenes biofilm cells on the surface of stainless steel to below the detection limit. However, bacteria withstanding heat stress become more resistant to other stress conditions, and may be more virulent when consumed by a host. Herein, we studied the transcriptional regulation of genes important for stress resistance and virulence in Salmonella biofilms after SHS treatments. Genes encoding heat shock proteins and general stress resistance proteins showed transcriptional surges after 1 sec of SHS treatment at 200°C, with parallel induction of stress-related regulator genes including rpoE, rpoS, and rpoH. Interestingly, Salmonella biofilm cells exposed to SHS showed decreased transcription of flagella and Salmonella pathogenicity island-1 (SPI-1) genes required for motility and invasion of host cells, respectively, whereas increased transcription of SPI-2 genes, important for bacterial survival and replication inside host cells, was detected. When the transcriptional response was compared between cells treated with SHS (200°C) and SS (100°C), SHS caused immediate changes in gene expression by shorter treatments. Understanding the status of Salmonella virulence and stress resistance induced by SHS treatments is important for wider application of SHS in controlling Salmonella biofilm formation during food production.

  12. Cold plasma inactivation of internalised bacteria and biofilms for Salmonella enterica serovar Typhimurium, Listeria monocytogenes and Escherichia coli.

    PubMed

    Ziuzina, Dana; Han, Lu; Cullen, Patrick J; Bourke, Paula

    2015-10-01

    Microbial biofilms and bacteria internalised in produce tissue may reduce the effectiveness of decontamination methods. In this study, the inactivation efficacy of in-package atmospheric cold plasma (ACP) afterglow was investigated against Salmonella Typhimurium, Listeria monocytogenes and Escherichia coli in the forms of planktonic cultures, biofilms formed on lettuce and associated bacteria internalised in lettuce tissue. Prepared lettuce broth (3%) was inoculated with bacteria resulting in a final concentration of ~7.0 log10 CFU/ml. For biofilm formation and internalisation, lettuce pieces (5 × 5 cm) were dip-inoculated in bacterial suspension of ~7.0 log10 CFU/ml for 2 h and further incubated for 0, 24 and 48 h at either 4 °C or room temperature (~22 °C) in combination with light/dark photoperiod or at 4 °C under dark conditions. Inoculated samples were sealed inside a rigid polypropylene container and indirectly exposed (i.e. placed outside plasma discharge) to a high voltage (80 kVRMS) air ACP with subsequent storage for 24 h at 4 °C. ACP treatment for 30s reduced planktonic populations of Salmonella, L. monocytogenes and E. coli suspended in lettuce broth to undetectable levels. Depending on storage conditions, bacterial type and age of biofilm, 300 s of treatment resulted in reduction of biofilm populations on lettuce by a maximum of 5 log10 CFU/sample. Scanning electron and confocal laser microscopy pointed to the incidence of bacterial internalisation and biofilm formation, which influenced the inactivation efficacy of ACP. Measured intracellular reactive oxygen species (ROS) revealed that the presence of organic matter in the bacterial suspension might present a protective effect against the action of ROS on bacterial cells. This study demonstrated that high voltage in-package ACP could be a potential technology to overcome bacterial challenges associated with food produce. However, the existence of biofilms and internalised bacteria should be

  13. Effects of Cattle Feeding Regimen and Soil Management Type on the Fate of Escherichia coli O157:H7 and Salmonella enterica Serovar Typhimurium in Manure, Manure-Amended Soil, and Lettuce

    PubMed Central

    Franz, Eelco; van Diepeningen, Anne D.; de Vos, Oscar J.; van Bruggen, Ariena H. C.

    2005-01-01

    Survival of the green fluorescent protein-transformed human pathogens Escherichia coli O157:H7 and Salmonella enterica serovar Typhimurium was studied in a laboratory-simulated lettuce production chain. Dairy cows were fed three different roughage types: high-digestible grass silage plus maize silage (6:4), low-digestible grass silage, and straw. Each was adjusted with supplemental concentrates to high and low crude protein levels. The pathogens were added to manure, which was subsequently mixed (after 56 and 28 days for E. coli O157:H7 and Salmonella serovar Typhimurium, respectively) with two pairs of organically and conventionally managed loamy and sandy soil. After another 14 days, iceberg lettuce seedlings were planted and then checked for pathogens after 21 days of growth. Survival data were fitted to a logistic decline function (exponential for E. coli O157:H7 in soil). Roughage type significantly influenced the rate of decline of E. coli O157:H7 in manure, with the fastest decline in manure from the pure straw diet and the slowest in manure from the diet of grass silage plus maize silage. Roughage type showed no effect on the rate of decline of Salmonella serovar Typhimurium, although decline was significantly faster in the manure derived from straw than in the manure from the diet of grass silage plus maize silage. The pH and fiber content of the manure were significant explanatory factors and were positively correlated with the rate of decline. With E. coli O157:H7 there was a trend of faster decline in organic than in conventional soils. No pathogens were detected in the edible lettuce parts. The results indicate that cattle diet and soil management are important factors with respect to the survival of human pathogens in the environment. PMID:16204535

  14. Effects of cattle feeding regimen and soil management type on the fate of Escherichia coli O157:H7 and salmonella enterica serovar typhimurium in manure, manure-amended soil, and lettuce.

    PubMed

    Franz, Eelco; van Diepeningen, Anne D; de Vos, Oscar J; van Bruggen, Ariena H C

    2005-10-01

    Survival of the green fluorescent protein-transformed human pathogens Escherichia coli O157:H7 and Salmonella enterica serovar Typhimurium was studied in a laboratory-simulated lettuce production chain. Dairy cows were fed three different roughage types: high-digestible grass silage plus maize silage (6:4), low-digestible grass silage, and straw. Each was adjusted with supplemental concentrates to high and low crude protein levels. The pathogens were added to manure, which was subsequently mixed (after 56 and 28 days for E. coli O157:H7 and Salmonella serovar Typhimurium, respectively) with two pairs of organically and conventionally managed loamy and sandy soil. After another 14 days, iceberg lettuce seedlings were planted and then checked for pathogens after 21 days of growth. Survival data were fitted to a logistic decline function (exponential for E. coli O157:H7 in soil). Roughage type significantly influenced the rate of decline of E. coli O157:H7 in manure, with the fastest decline in manure from the pure straw diet and the slowest in manure from the diet of grass silage plus maize silage. Roughage type showed no effect on the rate of decline of Salmonella serovar Typhimurium, although decline was significantly faster in the manure derived from straw than in the manure from the diet of grass silage plus maize silage. The pH and fiber content of the manure were significant explanatory factors and were positively correlated with the rate of decline. With E. coli O157:H7 there was a trend of faster decline in organic than in conventional soils. No pathogens were detected in the edible lettuce parts. The results indicate that cattle diet and soil management are important factors with respect to the survival of human pathogens in the environment.

  15. Transcriptional Profiling of a Cross-Protective Salmonella enterica serovar Typhimurium UK-1 dam Mutant Identifies a Set of Genes More Transcriptionally Active Compared to Wild-Type, and Stably Transcribed across Biologically Relevant Microenvironments

    PubMed Central

    Miller, Claire B.; Pierlé, Sebastian Aguilar; Brayton, Kelly A.; Ochoa, Jennine N.; Shah, Devendra H.; Lahmers, Kevin K.

    2014-01-01

    Vaccination with Salmonella enterica serovar Typhimurium lacking DNA adenine methyltransferase confers cross-protective immunity against multiple Salmonella serotypes. The mechanistic basis is thought to be associated with the de-repression of genes that are tightly regulated when transiting from one microenvironment to another. This de-repression provides a potential means for the production of a more highly expressed and stable antigenic repertoire capable of inducing cross-protective immune responses. To identify genes encoding proteins that may contribute to cross-protective immunity, we used a Salmonella Typhimurium DNA adenine methyltransferase mutant strain (UK-1 dam mutant) derived from the parental UK-1 strain, and assessed the transcriptional profile of the UK-1 dam mutant and UK-1 strain grown under conditions that simulate the intestinal or endosomal microenvironments encountered during the infective process. As expected, the transcriptional profile of the UK-1 dam mutant identified a set of genes more transcriptionally active when compared directly to UK-1, and stably transcribed in biologically relevant culture conditions. Further, 22% of these genes were more highly transcribed in comparison to two other clinically-relevant Salmonella serovars. The strategy employed here helps to identify potentially conserved proteins produced by the UK-1 dam mutant that stimulate and/or modulate the development of cross-protective immune responses toward multiple Salmonella serotypes. PMID:25364573

  16. Specificity of Salmonella Typhimurium strain (ATCC 14028) growth responses to Salmonella serovar-generated spent media.

    PubMed

    Calo, Juliany Rivera; Park, Si Hong; Baker, Christopher A; Ricke, Steven C

    2015-01-01

    Salmonella enterica is one of the most prevalent pathogens responsible for foodborne illness worldwide. Numerous Salmonella serovars have been associated with the consumption of a variety of products, and limiting food-borne illness due to Salmonella serovars is a continuing problem for food producers and public health. The emergence and prevalence of Salmonella serovars has been studied but the predominant serovars have varied somewhat over the years. The aims of this research were to compare the aerobic growth responses of selected predominant foodborne Salmonella serovars, and evaluate how the spent media from different serovars affects the growth of a well-characterized Salmonella Typhimurium strain. Growth responses were similar for most strains in spent media except for S. Typhimurium (ATCC 14028), which exhibited a decrease in growth in the presence of Salmonella Heidelberg (ARI-14) spent media. This research will provide a better understanding of the growth differences among several Salmonella serovars in nutrient limited spent media.

  17. Complete Genome and Methylome Sequences of Salmonella enterica subsp. enterica Serovar Panama (ATCC 7378) and Salmonella enterica subsp. enterica Serovar Sloterdijk (ATCC 15791)

    PubMed Central

    Yao, Kuan; Muruvanda, Tim; Roberts, Richard J.; Payne, Justin; Allard, Marc W.

    2016-01-01

    Salmonella enterica spp. are pathogenic bacteria commonly associated with food-borne outbreaks in human and animals. Salmonella enterica spp. are characterized into more than 2,500 different serotypes, which makes epidemiological surveillance and outbreak control more difficult. In this report, we announce the first complete genome and methylome sequences from two Salmonella type strains associated with food-borne outbreaks, Salmonella enterica subsp. enterica serovar Panama (ATCC 7378) and Salmonella enterica subsp. enterica serovar Sloterdijk (ATCC 15791). PMID:26988049

  18. Complete Genome and Methylome Sequences of Salmonella enterica subsp. enterica Serovar Panama (ATCC 7378) and Salmonella enterica subsp. enterica Serovar Sloterdijk (ATCC 15791).

    PubMed

    Yao, Kuan; Muruvanda, Tim; Roberts, Richard J; Payne, Justin; Allard, Marc W; Hoffmann, Maria

    2016-03-17

    Salmonella enterica spp. are pathogenic bacteria commonly associated with food-borne outbreaks in human and animals. Salmonella enterica spp. are characterized into more than 2,500 different serotypes, which makes epidemiological surveillance and outbreak control more difficult. In this report, we announce the first complete genome and methylome sequences from two Salmonella type strains associated with food-borne outbreaks, Salmonella enterica subsp. enterica serovar Panama (ATCC 7378) and Salmonella enterica subsp. enterica serovar Sloterdijk (ATCC 15791).

  19. Effect of feed particle size and feed processing on morphological characteristics in the small and large intestine of pigs and on adhesion of Salmonella enterica serovar Typhimurium DT12 in the ileum in vitro.

    PubMed

    Hedemann, M S; Mikkelsen, L L; Naughton, P J; Jensen, B B

    2005-07-01

    A 2 x 2 factorial experiment with pigs was undertaken to investigate the effect of particle size (fine and coarse) and feed processing (pelleted and nonpelleted) on morphological characteristics in the small intestine, cecum, and colon of pigs and on the adhesion of Salmonella enterica serovar Typhimurium DT12 to the ileum in vitro. Ninety-six pigs (average BW = 33 +/- 7 kg) were fed the experimental diets. After 4 wk, 24 pigs were selected (six pigs per diet) and euthanized, and tissue samples were taken from the mid and distal small intestine, cecum, and distal colon. The effects of particle size and feed processing on villus height and crypt depth in the small intestine were minor. Feeding coarse diets increased (P = 0.05) the crypt depth in the colon. The crypt depth was 420 +/- 12 and 449 +/- 12 microm in pigs fed finely and coarsely ground feed, respectively. Pigs fed pelleted diets had a larger (P = 0.01) staining area for neutral mucins, as well as for acidic and sulfomucins on the villi of the distal small intestine than pigs fed nonpelleted diets. The area was 41, 46, and 33% larger for neutral, acidic, and sulfomucins, respectively. The mucin-staining areas of the crypts in the cecum and the colon were not affected by the experimental diets. Examination of lectin binding characteristics of the distal small intestine and the cecum did not reveal any differences between the experimental diets. Using a pig intestine organ culture model, Salmonella adhered less (P < 0.05) to the ileal tissue of pigs fed the nonpelleted diets than to those fed pelleted diets; the adherence was 60% less in these pigs. Results of this study suggest that pigs fed pelleted diets secrete mucins that are capable of binding Salmonella enterica serovar Typhimurium DT12 and thereby allowing for colonization. Therefore, pigs fed a nonpelleted diet are better protected against Salmonella infections than pigs fed a pelleted diet.

  20. Simultaneous near-infrared radiant heating and UV radiation for inactivating Escherichia coli O157:H7 and Salmonella enterica serovar Typhimurium in powdered red pepper (Capsicum annuum L.).

    PubMed

    Ha, Jae-Won; Kang, Dong-Hyun

    2013-11-01

    This study was conducted to investigate the efficacy of the simultaneous application of near-infrared (NIR) heating and UV irradiation for reducing populations of food-borne pathogens, including Salmonella enterica serovar Typhimurium and Escherichia coli O157:H7 in red pepper powder and to clarify the mechanisms of the lethal effect of the NIR-UV combined treatment. Also, the effect of the combination treatment on quality was determined by measuring changes in color and pungency constituents. Simultaneous NIR-UV combined treatment for 5 min achieved 3.34- and 2.78-log CFU reductions in S. Typhimurium and E. coli O157:H7, respectively, which involved 1.86- and 1.31-log CFU reductions, respectively, which were attributed to the synergistic effect. Through qualitative and quantitative analyses, damage to the cell envelope was identified as the main factor contributing to the synergistic lethal effect of NIR-UV combined treatment. Color values and capsaicin and dihydrocapsaicin content of NIR-UV simultaneously treated red pepper powder were not significantly (P > 0.05) different from those of untreated samples. These results suggest that simultaneous application of NIR and UV treatment can be effectively used to control food-borne pathogens in powdered red pepper without affecting quality.

  1. Emergence of a multidrug-resistant (ASSuTTm) strain of Salmonella enterica serovar Typhimurium DT120 in England in 2011 and the use of multiple-locus variable-number tandem-repeat analysis in supporting outbreak investigations.

    PubMed

    Paranthaman, Karthikeyan; Haroon, Sophie; Latif, Samia; Vinnyey, Natalie; de Souza, Valerie; Welfare, William; Tahir, Mamoona; Cooke, Edward; Stone, Kirsten; Lane, Chris; Peters, Tansy; Puleston, Richard

    2013-10-01

    In summer 2011, two outbreaks of a unique, multidrug-