scholarly journals Characterization of grvA, an Antivirulence Gene on the Gifsy-2 Phage in Salmonella enterica Serovar Typhimurium

2001 ◽  
Vol 183 (2) ◽  
pp. 611-620 ◽  
Author(s):  
Theresa D. Ho ◽  
James M. Slauch
Keyword(s):  
Virulence Factors ◽  
Salmonella Enterica ◽  
Single Copy ◽  
Null Mutation ◽  
Multicopy Plasmid ◽  
Wild Type ◽  
Reading Frame ◽  
Serovar Typhimurium ◽  
Phage Production ◽  
Superoxide Dismutase Gene

ABSTRACT The lambdoid phage Gifsy-2 contributes significantly toSalmonella enterica serovar Typhimurium virulence. The phage carries the periplasmic superoxide dismutase gene,sodCI, and other unidentified virulence factors. We have characterized the gene grvA, a single open reading frame inserted in the opposite orientation in the tail operon of the Gifsy-2 phage. Contrary to what is observed with classic virulence genes,grvA null mutants were more virulent than wild type as measured by intraperitoneal competition assays in mice. We have termed this effect antivirulence. Wild-type grvA in single copy complemented this phenotype. However, grvA +on a multicopy plasmid also conferred the antivirulence phenotype. Neither a grvA null mutation nor thegrvA + plasmid conferred a growth advantage or disadvantage in laboratory media. The antivirulence phenotype conferred by the grvA null mutation and thegrvA + plasmid required wild-typesodCI but was independent of other virulence factors encoded on Gifsy-2. These results suggest that in a wild-type situation, GrvA decreases the pathogenicity of serovar Typhimurium in the host, most likely by affecting resistance to toxic oxygen species. These virulence phenotypes were independent of functional Gifsy-2 phage production. Our data suggest that the contribution of Gifsy-2 is a complicated sum of both positive virulence factors such assodCI and antivirulence factors such asgrvA.

scholarly journals Fis Regulates Transcriptional Induction of RpoS in Salmonella enterica

2005 ◽  
Vol 187 (5) ◽  
pp. 1568-1580 ◽  
Author(s):  
Matthew Hirsch ◽  
Thomas Elliott
Keyword(s):  
Exponential Growth ◽  
Salmonella Enterica ◽  
Single Copy ◽  
Regulation Of Transcription ◽  
Wild Type ◽  
Protein Activity ◽  
Serovar Typhimurium ◽  
Fold Induction ◽  
Transcriptional Induction

ABSTRACT The sigma factor RpoS is known to regulate at least 60 genes in response to environmental sources of stress or during growth to stationary phase (SP). Accumulation of RpoS relies on integration of multiple genetic controls, including regulation at the levels of transcription, translation, protein stability, and protein activity. Growth to SP in rich medium results in a 30-fold induction of RpoS, although the mechanism of this regulation is not understood. We characterized the activity of promoters serving rpoS in Salmonella enterica serovar Typhimurium and report that regulation of transcription during growth into SP depends on Fis, a DNA-binding protein whose abundance is high during exponential growth and very low in SP. A fis mutant of S. enterica serovar Typhimurium showed a ninefold increase in expression from the major rpoS promoter (P rpoS ) during exponential growth, whereas expression during SP was unaffected. Increased transcription from P rpoS in the absence of Fis eliminated the transcriptional induction as cells enter SP. The mutant phenotype can be complemented by wild-type fis carried on a single-copy plasmid. Fis regulation of rpoS requires the presence of a Fis site positioned at −50 with respect to P rpoS , and this site is bound by Fis in vitro. A model is presented in which Fis binding to this site allows repression of rpoS specifically during exponential growth, thus mediating transcriptional regulation of rpoS.

scholarly journals Tissue-Specific Gene Expression Identifies a Gene in the Lysogenic Phage Gifsy-1 That Affects Salmonella enterica Serovar Typhimurium Survival in Peyer's Patches

2000 ◽  
Vol 182 (16) ◽  
pp. 4406-4413 ◽  
Author(s):  
Theresa L. Stanley ◽  
Craig D. Ellermeier ◽  
James M. Slauch
Keyword(s):  
Small Intestine ◽  
Salmonella Enterica ◽  
Specific Gene ◽  
Peyer’S Patch ◽  
Wild Type ◽  
Peyer's Patch ◽  
Reading Frame ◽  
Serovar Typhimurium ◽  
Sense And Respond

ABSTRACT In vivo expression technology was used to identify Salmonella enterica serovar Typhimurium genes that are transcriptionally induced when the bacteria colonize the small intestines of mice. These genes were subsequently screened for those that are transcriptionally inactive during the systemic stages of disease. This procedure identified gipA, a gene that is specifically induced in the small intestine of the animal. The gipA gene is carried on the lambdoid phage Gifsy-1. Consistent with the expression profile, the sole defect conferred by a gipA null mutation is in growth or survival in a Peyer's patch. The gipA strain is wild type in its ability to initially colonize the small intestine and invade the intestinal epithelium. The mutant also survives and propagates at wild-type levels during the systemic stages of disease. The gipA open reading frame is homologous to a family of putative insertion sequence elements, although our evidence shows that transposition is not required for gipA function in the Peyer's patch. These results suggest that the bacteria sense and respond to the particular environment of the Peyer's patch, a critical site for the replication of Salmonella serovar Typhimurium.

scholarly journals A network of Rab GTPases controls phagosome maturation and is modulated by Salmonella enterica serovar Typhimurium

2007 ◽  
Vol 176 (3) ◽  
pp. 263-268 ◽  
Author(s):  
Adam C. Smith ◽  
Won Do Heo ◽  
Virginie Braun ◽  
Xiuju Jiang ◽  
Chloe Macrae ◽  
...  
Keyword(s):  
Salmonella Enterica ◽  
Salmonella Enterica Serovar Typhimurium ◽  
Dominant Negative ◽  
Rab Gtpases ◽  
Wild Type ◽  
Intracellular Growth ◽  
Non Invasive ◽  
Serovar Typhimurium ◽  
Guanosine Triphosphatase ◽  
Kinetics Of

Members of the Rab guanosine triphosphatase (GTPase) family are key regulators of membrane traffic. Here we examined the association of 48 Rabs with model phagosomes containing a non-invasive mutant of Salmonella enterica serovar Typhimurium (S. Typhimurium). This mutant traffics to lysosomes and allowed us to determine which Rabs localize to a maturing phagosome. In total, 18 Rabs associated with maturing phagosomes, each with its own kinetics of association. Dominant-negative mutants of Rab23 and 35 inhibited phagosome–lysosome fusion. A large number of Rab GTPases localized to wild-type Salmonella-containing vacuoles (SCVs), which do not fuse with lysosomes. However, some Rabs (8B, 13, 23, 32, and 35) were excluded from wild-type SCVs whereas others (5A, 5B, 5C, 7A, 11A, and 11B) were enriched on this compartment. Our studies demonstrate that a complex network of Rab GTPases controls endocytic progression to lysosomes and that this is modulated by S. Typhimurium to allow its intracellular growth.

scholarly journals Salmonella enterica Serovar Typhimurium Uses PbgA/YejM To Regulate Lipopolysaccharide Assembly during Bacteremia

2019 ◽  
Vol 88 (1) ◽  
Author(s):  
Melina B. Cian ◽  
Nicole P. Giordano ◽  
Revathi Masilamani ◽  
Keaton E. Minor ◽  
Zachary D. Dalebroux
Keyword(s):  
Salmonella Enterica ◽  
Lipid A ◽  
Salmonella Enterica Serovar Typhimurium ◽  
Transmembrane Domain ◽  
Wild Type ◽  
Content Type ◽  
Serovar Typhimurium ◽  
Periplasmic Domain ◽  
Inner Membrane Protein ◽  
Substitution Mutations

ABSTRACT Salmonella enterica serovar Typhimurium (S. Typhimurium) relies upon the inner membrane protein PbgA to enhance outer membrane (OM) integrity and promote virulence in mice. The PbgA transmembrane domain (residues 1 to 190) is essential for viability, while the periplasmic domain (residues 191 to 586) is dispensable. Residues within the basic region (residues 191 to 245) bind acidic phosphates on polar phospholipids, like for cardiolipins, and are necessary for salmonella OM integrity. S. Typhimurium bacteria increase their OM cardiolipin concentrations during activation of the PhoPQ regulators. The mechanism involves PbgA’s periplasmic globular region (residues 245 to 586), but the biological role of increasing cardiolipins on the surface is not understood. Nonsynonymous polymorphisms in three essential lipopolysaccharide (LPS) synthesis regulators, lapB (also known as yciM), ftsH, and lpxC, variably suppressed the defects in OM integrity, rifampin resistance, survival in macrophages, and systemic colonization of mice in the pbgAΔ191–586 mutant (in which the PbgA periplasmic domain from residues 191 to 586 is deleted). Compared to the OMs of the wild-type salmonellae, the OMs of the pbgA mutants had increased levels of lipid A-core molecules, cardiolipins, and phosphatidylethanolamines and decreased levels of specific phospholipids with cyclopropanated fatty acids. Complementation and substitution mutations in LapB and LpxC generally restored the phospholipid and LPS assembly defects for the pbgA mutants. During bacteremia, mice infected with the pbgA mutants survived and cleared the bacteria, while animals infected with wild-type salmonellae succumbed within 1 week. Remarkably, wild-type mice survived asymptomatically with pbgA-lpxC salmonellae in their livers and spleens for months, but Toll-like receptor 4-deficient animals succumbed to these infections within roughly 1 week. In summary, S. Typhimurium uses PbgA to influence LPS assembly during stress in order to survive, adapt, and proliferate within the host environment.

scholarly journals Novel Salmonella enterica Serovar Typhimurium Protein That Is Indispensable for Virulence and Intracellular Replication

2001 ◽  
Vol 69 (12) ◽  
pp. 7413-7418 ◽  
Author(s):  
Tahar van der Straaten ◽  
Angela van Diepen ◽  
Kitty Kwappenberg ◽  
Sjaak van Voorden ◽  
Kees Franken ◽  
...  
Keyword(s):  
Amino Acid ◽  
Protein Function ◽  
Salmonella Enterica ◽  
Host Cells ◽  
Wild Type ◽  
Intracellular Replication ◽  
Oxygen Intermediates ◽  
Serovar Typhimurium

ABSTRACT Upon contact with host cells, the intracellular pathogenSalmonella enterica serovar Typhimurium promotes its uptake, targeting, and survival in intracellular niches. In this process, the bacterium evades the microbicidal effector mechanisms of the macrophage, including oxygen intermediates. This study reports the phenotypic and genotypic characterization of an S. enterica serovar Typhimurium mutant that is hypersusceptible to superoxide. The susceptible phenotype is due to a MudJ insertion-inactivation of a previously undescribedSalmonella gene designated sspJ that is located between 54.4 and 64 min of the Salmonellachromosome and encodes a 392-amino-acid protein. In vivo, upon intraperitoneal injection of 104 to 107bacteria in C3H/HeN and 101 to 104 bacteria in BALB/c mice, the mutant strain was less virulent than the wild type. Consistent with this finding, during the first hour after ingestion by macrophage-like J774 and RAW264.7 cells in vitro, the intracellular killing of the strain carrying sspJ::MudJ is enhanced fivefold over that of wild-type microorganisms. Wild-type salmonellae displayed significant intracellular replication during the first 24 h after uptake, but sspJ::MudJ mutants failed to do so. This phenotype could be restored to that of the wild type by sspJ complementation. The SspJ protein is found in the cytoplasmic membrane and periplasmic space. Amino acid sequence homology analysis did reveal a leader sequence and putative pyrroloquinoline quinone-binding domains, but no putative protein function. We excluded the possibility that SspJ is a scavenger of superoxide or has superoxide dismutase activity.

scholarly journals A Plasmid-Encoded FetMP-Fls Iron Uptake System Confers Selective Advantages to Salmonella enterica Serovar Typhimurium in Growth under Iron-Restricted Conditions and for Infection of Mammalian Host Cells

2020 ◽  
Vol 8 (5) ◽  
pp. 630
Author(s):  
Vanesa García ◽  
Ana Herrero-Fresno ◽  
Rosaura Rodicio ◽  
Alfonso Felipe-López ◽  
Ignacio Montero ◽  
...  
Keyword(s):  
Iron Content ◽  
Salmonella Enterica ◽  
Iron Uptake ◽  
Type Strain ◽  
Wild Type Strain ◽  
Salmonella Enterica Serovar Typhimurium ◽  
Iron Acquisition ◽  
Clinical Strain ◽  
Wild Type ◽  
Serovar Typhimurium

The resistance plasmid pUO-StVR2, derived from virulence plasmid pSLT, is widespread in clinical isolates of Salmonella enterica serovar Typhimurium recovered in Spain and other European countries. pUO-StVR2 carries several genes encoding a FetMP-Fls system, which could be involved in iron uptake. We therefore analyzed S. Typhimurium LSP 146/02, a clinical strain selected as representative of the isolates carrying the plasmid, and an otherwise isogenic mutant lacking four genes (fetMP-flsDA) of the fetMP-fls region. Growth curves and determination of the intracellular iron content under iron-restricted conditions demonstrated that deletion of these genes impairs iron acquisition. Thus, under these conditions, the mutant grew significantly worse than the wild-type strain, its iron content was significantly lower, and it was outcompeted by the wild-type strain in competition assays. Importantly, the strain lacking the fetMP-flsDA genes was less invasive in cultured epithelial HeLa cells and replicated poorly upon infection of RAW264.7 macrophages. The genes were introduced into S. Typhimurium ATCC 14028, which lacks the FetMP-Fls system, and this resulted in increased growth under iron limitation as well as an increased ability to multiply inside macrophages. These findings indicate that the FetMP-Fls iron acquisition system exceeds the benefits conferred by the other high-affinity iron uptake systems carried by ATCC 14028 and LSP 146/02. We proposed that effective iron acquisition by this system in conjunction with antimicrobial resistance encoded from the same plasmid have greatly contributed to the epidemic success of S. Typhimurium isolates harboring pUO-StVR2.

scholarly journals An Attenuated Salmonella enterica Serovar Typhimurium Strain and Galacto-Oligosaccharides Accelerate Clearance of Salmonella Infections in Poultry through Modifications to the Gut Microbiome

2017 ◽  
Vol 84 (5) ◽  
Author(s):  
M. Andrea Azcarate-Peril ◽  
Natasha Butz ◽  
Maria Belen Cadenas ◽  
Matthew Koci ◽  
Anne Ballou ◽  
...  
Keyword(s):  
United States ◽  
Gut Microbiota ◽  
Gut Microbiome ◽  
Salmonella Enterica ◽  
The United States ◽  
Wild Type ◽  
Content Type ◽  
Salmonella Infections ◽  
Serovar Typhimurium ◽  
The Impact

ABSTRACT Salmonella is estimated to cause one million foodborne illnesses in the United States every year. Salmonella -contaminated poultry products are one of the major sources of salmonellosis. Given the critical role of the gut microbiota in Salmonella transmission, a manipulation of the chicken intestinal microenvironment could prevent animal colonization by the pathogen. In Salmonella , the global regulator gene fnr ( f umarate n itrate r eduction) regulates anaerobic metabolism and is essential for adapting to the gut environment. This study tested the hypothesis that an attenuated Fnr mutant of Salmonella enterica serovar Typhimurium (attST) or prebiotic galacto-oligosaccharides (GOS) could improve resistance to wild-type Salmonella via modifications to the structure of the chicken gut microbiome. Intestinal samples from a total of 273 animals were collected weekly for 9 weeks to evaluate the impact of attST or prebiotic supplementation on microbial species of the cecum, duodenum, jejunum, and ileum. We next analyzed changes to the gut microbiome induced by challenging the animals with a wild-type Salmonella serovar 4,[5],12:r:− (Nal r ) strain and determined the clearance rate of the virulent strain in the treated and control groups. Both GOS and the attenuated Salmonella strain modified the gut microbiome but elicited alterations of different taxonomic groups. The attST produced significant increases of Alistipes and undefined Lactobacillus , while GOS increased Christensenellaceae and Lactobacillus reuteri . The microbiome structural changes induced by both treatments resulted in a faster clearance after a Salmonella challenge. IMPORTANCE With an average annual incidence of 13.1 cases/100,000 individuals, salmonellosis has been deemed a nationally notifiable condition in the United States by the Centers for Disease Control and Prevention (CDC). Earlier studies demonstrated that Salmonella is transmitted by a subset of animals (supershedders). The supershedder phenotype can be induced by antibiotics, ascertaining an essential role for the gut microbiota in Salmonella transmission. Consequently, modulation of the gut microbiota and modification of the intestinal microenvironment could assist in preventing animal colonization by the pathogen. Our study demonstrated that a manipulation of the chicken gut microbiota by the administration of an attenuated Salmonella strain or prebiotic galacto-oligosaccharides (GOS) can promote resistance to Salmonella colonization via increases of beneficial microorganisms that translate into a less hospitable gut microenvironment.

scholarly journals Flagellar Phase Variation of Salmonella enterica Serovar Typhimurium Contributes to Virulence in the Murine Typhoid Infection Model but Does Not InfluenceSalmonella-Induced Enteropathogenesis

2001 ◽  
Vol 69 (5) ◽  
pp. 3021-3030 ◽  
Author(s):  
Jack S. Ikeda ◽  
Clare K. Schmitt ◽  
Stephen C. Darnell ◽  
Patricia R. Watson ◽  
Jennifer Bispham ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Salmonella Enterica ◽  
Phase Variation ◽  
Selective Advantage ◽  
Peritoneal Macrophages ◽  
Infection Model ◽  
Wild Type ◽  
Two Phase ◽  
Serovar Typhimurium

ABSTRACT Although Salmonella enterica serovar Typhimurium can undergo phase variation to alternately express two different types of flagellin subunit proteins, FljB or FliC, no biological function for this phenomenon has been described. In this investigation, we constructed phase-locked derivatives of S. enterica serovar Typhimurium that expressed only FljB (termed locked-ON) or FliC (termed locked-OFF). The role of phase variation in models of enteric and systemic pathogenesis was then evaluated. There were no differences between the wild-type parent strain and the two phase-locked derivatives in adherence and invasion of mouse epithelial cells in vitro, survival in mouse peritoneal macrophages, or in a bovine model of gastroenteritis. By contrast, the locked-OFF mutant was virulent in mice following oral or intravenous (i.v.) inoculation but the locked-ON mutant was attenuated. When these phase-locked mutants were compared in studies of i.v. kinetics in mice, similar numbers of the two strains were isolated from the blood and spleens of infected animals at 6 and 24 h. However, the locked-OFF mutant was recovered from the blood and spleens in significantly greater numbers than the locked-ON strain by day 2 of infection. By 5 days postinfection, a majority of the mice infected with the locked-OFF mutant had died compared with none of the mice infected with the locked-ON mutant. These results suggest that phase variation is not involved in the intestinal stage of infection but that once S. enterica serovar Typhimurium reaches the spleens of susceptible mice those organisms in the FliC phase can grow and/or survive better than those in the FljB phase. Additional experiments with wild-type S. enterica serovar Typhimurium, fully capable of switching flagellin type, supported this hypothesis. We conclude that organisms that have switched to the FliC+phase have a selective advantage in the mouse model of typhoid fever but have no such advantage in invasion of epithelial cells or the induction of enteropathogenesis.

scholarly journals aro Mutations in Salmonella enterica Cause Defects in Cell Wall and Outer Membrane Integrity

2008 ◽  
Vol 190 (9) ◽  
pp. 3155-3160 ◽  
Author(s):  
Alena Sebkova ◽  
Daniela Karasova ◽  
Magdalena Crhanova ◽  
Eva Budinska ◽  
Ivan Rychlik
Keyword(s):  
Gene Expression ◽  
Cell Wall ◽  
Outer Membrane ◽  
Salmonella Enterica ◽  
Membrane Integrity ◽  
Innate Immune ◽  
Multicopy Plasmid ◽  
Oral Vaccines ◽  
Serovar Typhimurium ◽  
Serum Albumen

ABSTRACT In this study we characterized aro mutants of Salmonella enterica serovars Enteritidis and Typhimurium, which are frequently used as live oral vaccines. We found that the aroA, aroD, and aroC mutants were sensitive to blood serum, albumen, EDTA, and ovotransferrin, and this defect could be complemented by an appropriate aro gene cloned in a plasmid. Subsequent microarray analysis of gene expression in the aroD mutant in serovar Typhimurium indicated that the reason for this sensitivity might be the upregulation of murA. To confirm this, we artificially overexpressed murA from a multicopy plasmid, and this overexpression caused sensitivity of the strain to albumen and EDTA but not to serum and ovotransferrin. We concluded that attenuation of aro mutants is caused not only by their inability to synthesize aromatic metabolites but also by their defect in cell wall and outer membrane functions associated with decreased resistance to components of innate immune response.

scholarly journals Modulation of Virulence by Two Acidified Nitrite-Responsive Loci of Salmonella enterica Serovar Typhimurium

2003 ◽  
Vol 71 (6) ◽  
pp. 3196-3205 ◽  
Author(s):  
Charles C. Kim ◽  
Denise Monack ◽  
Stanley Falkow
Keyword(s):  
Salmonella Enterica ◽  
Fluorescent Protein ◽  
Dependent Manner ◽  
Wild Type ◽  
Independent Manner ◽  
Inducible Promoters ◽  
Serovar Typhimurium ◽  
Bacterial Genes ◽  
Green Fluorescent

ABSTRACT Two acidified nitrite-inducible genes of Salmonella enterica serovar Typhimurium were identified with a green fluorescent protein-based promoter-trap screen. The nitrite-inducible promoters were located upstream of loci that we designated nipAB and nipC, which correspond to hcp-hcr (hybrid cluster protein) of Escherichia coli and norA of Alcaligenes eutrophus, respectively. Maximal induction of the promoters by nitrite was dependent on pH. The nipAB promoter was regulated by oxygen in an Fnr-dependent manner. The nipC promoter was also regulated by oxygen but in an Fnr-independent manner. The promoters were upregulated in activated RAW264.7 macrophage-like cells, which produce NO via the inducible nitric oxide synthase (iNOS), and the induction was inhibited by aminoguanidine, an inhibitor of iNOS. Although the nipAB and nipC mutants displayed no defects under a variety of in vitro conditions or in tissue culture infections, they exhibited lower oral 50% lethal doses (LD50s) than did the wild type in C57BL/6J mouse infections. The lower LD50s reflected an unexpected increased ability of small inoculating doses of the mutant bacteria to cause lethal infection 2 to 3 weeks after challenge, compared to a similar challenge dose of wild-type bacteria. We conclude that these genes are regulated by physiological nitrogen oxides and that the absence of these bacterial genes in some way diminishes the ability of mice to clear a low dose infection.

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