scholarly journals Regulation of the Salmonella enterica std Fimbrial Operon by DNA Adenine Methylation, SeqA, and HdfR

2008 ◽  
Vol 190 (22) ◽  
pp. 7406-7413 ◽  
Author(s):  
Marcello Jakomin ◽  
Daniela Chessa ◽  
Andreas J. Bäumler ◽  
Josep Casadesús
Keyword(s):  
Salmonella Enterica ◽  
Bacterial Population ◽  
Phase Variation ◽  
Oral Infection ◽  
Wild Type ◽  
Adenine Methylation ◽  
Dam Methylation ◽  
Serovar Typhimurium ◽  
Dna Adenine Methylase ◽  
In The Wild

ABSTRACT DNA adenine methylase (dam) mutants of Salmonella enterica serovar Typhimurium grown under laboratory conditions express the std fimbrial operon, which is tightly repressed in the wild type. Here, we show that uncontrolled production of Std fimbriae in S. enterica serovar Typhimurium dam mutants contributes to attenuation in mice, as indicated by the observation that an stdA dam strain is more competitive than a dam strain upon oral infection. Dam methylation appears to regulate std transcription, rather than std mRNA stability or turnover. A genetic screen for std regulators showed that the GATC-binding protein SeqA directly or indirectly represses std expression, while the poorly characterized yifA gene product serves as an std activator. YifA encodes a putative LysR-like protein and has been renamed HdfR, like its Escherichia coli homolog. Activation of std expression by HdfR is observed only in dam and seqA backgrounds. These data suggest that HdfR directly or indirectly activates std transcription. Since SeqA is unable to bind nonmethylated DNA, it is possible that std operon derepression in dam and seqA mutants may result from unconstrained HdfR-mediated activation of std transcription. Derepression of std in dam and seqA mutants of S. enterica occurs in only a fraction of the bacterial population, suggesting the occurrence of either bistable expression or phase variation.

scholarly journals Regulation of finP Transcription by DNA Adenine Methylation in the Virulence Plasmid of Salmonella enterica

2005 ◽  
Vol 187 (16) ◽  
pp. 5691-5699 ◽  
Author(s):  
Eva M. Camacho ◽  
Ana Serna ◽  
Cristina Madrid ◽  
Silvia Marqués ◽  
Raúl Fernández ◽  
...  
Keyword(s):  
Dna Sequences ◽  
Salmonella Enterica ◽  
Rna Stability ◽  
Virulence Plasmid ◽  
Methylation State ◽  
Adenine Methylation ◽  
Dam Methylation ◽  
Serovar Typhimurium ◽  
Dna Adenine Methylase ◽  
Gatc Site

ABSTRACT DNA adenine methylase (Dam−) mutants of Salmonella enterica serovar Typhimurium contain reduced levels of FinP RNA encoded on the virulence plasmid. Dam methylation appears to regulate finP transcription, rather than FinP RNA stability or turnover. The finP promoter includes canonical −10 and −35 modules and depends on the σ70 factor. Regulation of finP transcription by Dam methylation does not require DNA sequences upstream from the −35 module, indicating that Dam acts at the promoter itself or downstream. Unexpectedly, a GATC site overlapping with the −10 module is likewise dispensable for Dam-mediated regulation. These observations indicate that Dam methylation regulates finP transcription indirectly and suggest the involvement of a host factor(s) responsive to the Dam methylation state of the cell. We provide evidence that one such factor is the nucleoid protein H-NS, which acts as a repressor of finP transcription in a Dam− background. H-NS also restrains transcription of the overlapping traJ gene, albeit in a Dam-independent fashion. Hence, the decreased FinP RNA content found in Dam− hosts of S. enterica appears to result from H-NS-mediated repression of finP transcription.

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 DNA Adenine Methylation Regulates Virulence Gene Expression in Salmonella enterica Serovar Typhimurium

2006 ◽  
Vol 188 (23) ◽  
pp. 8160-8168 ◽  
Author(s):  
Roberto Balbontín ◽  
Gary Rowley ◽  
M. Graciela Pucciarelli ◽  
Javier López-Garrido ◽  
Yvette Wormstone ◽  
...  
Keyword(s):  
Gene Expression ◽  
Salmonella Enterica ◽  
Expression Patterns ◽  
Virulence Gene ◽  
Luria Bertani ◽  
Altered Expression ◽  
Virulence Gene Expression ◽  
Adenine Methylation ◽  
Dam Methylation ◽  
Serovar Typhimurium

ABSTRACT Transcriptomic analyses during growth in Luria-Bertani medium were performed in strain SL1344 of Salmonella enterica serovar Typhimurium and in two isogenic derivatives lacking Dam methylase. More genes were repressed than were activated by Dam methylation (139 versus 37). Key genes that were differentially regulated by Dam methylation were verified independently. The largest classes of Dam-repressed genes included genes belonging to the SOS regulon, as previously described in Escherichia coli, and genes of the SOS-inducible Salmonella prophages ST64B, Gifsy-1, and Fels-2. Dam-dependent virulence-related genes were also identified. Invasion genes in pathogenicity island SPI-1 were activated by Dam methylation, while the fimbrial operon std was repressed by Dam methylation. Certain flagellar genes were repressed by Dam methylation, and Dam− mutants of S. enterica showed reduced motility. Altered expression patterns in the absence of Dam methylation were also found for the chemotaxis genes cheR (repressed by Dam) and STM3216 (activated by Dam) and for the Braun lipoprotein gene, lppB (activated by Dam). The requirement for DNA adenine methylation in the regulation of specific virulence genes suggests that certain defects of Salmonella Dam− mutants in the mouse model may be caused by altered patterns of gene expression.

scholarly journals LuxS-Based Quorum Sensing Does Not Affect the Ability of Salmonella enterica Serovar Typhimurium To Express the SPI-1 Type 3 Secretion System, Induce Membrane Ruffles, or Invade Epithelial Cells

2009 ◽  
Vol 191 (23) ◽  
pp. 7253-7259 ◽  
Author(s):  
Charlotte A. Perrett ◽  
Michail H. Karavolos ◽  
Suzanne Humphrey ◽  
Pietro Mastroeni ◽  
Isabel Martinez-Argudo ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Quorum Sensing ◽  
Salmonella Enterica ◽  
Type Strain ◽  
Wild Type Strain ◽  
Wild Type ◽  
Protein Fusion ◽  
Serovar Typhimurium ◽  
In The Wild ◽  
Type 3

ABSTRACT Bacterial species can communicate by producing and sensing small autoinducer molecules by a process known as quorum sensing. Salmonella enterica produces autoinducer 2 (AI-2) via the luxS synthase gene, which is used by some bacterial pathogens to coordinate virulence gene expression with population density. We investigated whether the luxS gene might affect the ability of Salmonella enterica serovar Typhimurium to invade epithelial cells. No differences were found between the wild-type strain of S. Typhimurium, SL1344, and its isogenic luxS mutant with respect to the number and morphology of the membrane ruffles induced or their ability to invade epithelial cells. The dynamics of the ruffling process were also similar in the wild-type strain (SL1344) and the luxS mutant. Furthermore, comparing the Salmonella pathogenicity island 1 (SPI-1) type 3 secretion profiles of wild-type SL1344 and the luxS mutant by Western blotting and measuring the expression of a single-copy green fluorescent protein fusion to the prgH (an essential SPI-1 gene) promoter indicated that SPI-1 expression and activity are similar in the wild-type SL1344 and luxS mutant. Genetic deletion of luxS did not alter the virulence of S. Typhimurium in the mouse model, and therefore, it appears that luxS does not play a significant role in regulating invasion of Salmonella in vitro or in vivo.

scholarly journals Salmonella enterica Serovar Typhimurium Mutants Unable To Convert Malate to Pyruvate and Oxaloacetate Are Avirulent and Immunogenic in BALB/c Mice

2009 ◽  
Vol 77 (4) ◽  
pp. 1397-1405 ◽  
Author(s):  
Regino Mercado-Lubo ◽  
Mary P. Leatham ◽  
Tyrrell Conway ◽  
Paul S. Cohen
Keyword(s):  
Salmonella Enterica ◽  
Type Strain ◽  
Wild Type Strain ◽  
Salmonella Enterica Serovar Typhimurium ◽  
Tca Cycle ◽  
Tricarboxylic Acid ◽  
Oral Infection ◽  
Wild Type ◽  
The Tca Cycle ◽  
Serovar Typhimurium

ABSTRACT Previously, we showed that the Salmonella enterica serovar Typhimurium SR-11 tricarboxylic acid (TCA) cycle must operate as a complete cycle for full virulence after oral infection of BALB/c mice (M. Tchawa Yimga, M. P. Leatham, J. H. Allen, D. C. Laux, T. Conway, and P. S. Cohen, Infect. Immun. 74:1130-1140, 2006). In the same study, we showed that for full virulence, malate must be converted to both oxaloacetate and pyruvate. Moreover, it was recently demonstrated that blocking conversion of succinyl-coenzyme A to succinate attenuates serovar Typhimurium SR-11 but does not make it avirulent; however, blocking conversion of succinate to fumarate renders it completely avirulent and protective against subsequent oral infection with the virulent serovar Typhimurium SR-11 wild-type strain (R. Mercado-Lubo, E. J. Gauger, M. P. Leatham, T. Conway, and P. S. Cohen, Infect. Immun. 76:1128-1134, 2008). Furthermore, the ability to convert succinate to fumarate appeared to be required only after serovar Typhimurium SR-11 became systemic. In the present study, evidence is presented that serovar Typhimurium SR-11 mutants that cannot convert fumarate to malate or that cannot convert malate to both oxaloacetate and pyruvate are also avirulent and protective in BALB/c mice. These results suggest that in BALB/c mice, the malate that is removed from the TCA cycle in serovar Typhimurium SR-11 for conversion to pyruvate must be replenished by succinate or one of its precursors, e.g., arginine or ornithine, which might be available in mouse phagocytes.

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.

Oral immunization with a dam mutant of Yersinia pseudotuberculosis protects against plague

Microbiology ◽  
2005 ◽  
Vol 151 (6) ◽  
pp. 1919-1926 ◽  
Author(s):  
Victoria L. Taylor ◽  
Richard W. Titball ◽  
Petra C. F. Oyston
Keyword(s):  
Gene Expression ◽  
Bile Salts ◽  
Yersinia Pseudotuberculosis ◽  
Lethal Dose ◽  
Oral Immunization ◽  
Wild Type ◽  
Gene Encoding ◽  
Serovar Typhimurium ◽  
Dna Adenine Methylase ◽  
Methylase Gene

Inactivation of the gene encoding DNA adenine methylase (dam) has been shown to attenuate some pathogens such as Salmonella enterica serovar Typhimurium and is a lethal mutation in others such as Yersinia pseudotuberculosis strain YPIII. In this study the dam methylase gene in Yersinia pseudotuberculosis strain IP32953 was inactivated. Unlike the wild-type, DNA isolated from the mutant could be digested with MboI, which is consistent with an altered pattern of DNA methylation. The mutant was sensitive to bile salts but not to 2-aminopurine. The effect of dam inactivation on gene expression was examined using a DNA microarray. In BALB/c mice inoculated orally or intravenously with the dam mutant, the median lethal dose (MLD) was at least 106-fold higher than the MLD of the wild-type. BALB/c mice inoculated with the mutant were protected against a subcutaneous challenge with 100 MLDs of Yersinia pestis strain GB and an intravenous challenge with 300 MLDs of Y. pseudotuberculosis IP32953.

scholarly journals H-NS Represses Salmonella enterica Serovar Typhimurium dsbA Expression during Exponential Growth

2004 ◽  
Vol 186 (4) ◽  
pp. 910-918 ◽  
Author(s):  
C. V. Gallant ◽  
T. Ponnampalam ◽  
H. Spencer ◽  
J. C. D. Hinton ◽  
N. L. Martin
Keyword(s):  
Exponential Growth ◽  
Salmonella Enterica ◽  
Immunoblot Analysis ◽  
Exponential Phase ◽  
Exponential Growth Phase ◽  
Mobility Shift ◽  
Altered Expression ◽  
Serovar Typhimurium ◽  
In The Wild ◽  
Maximal Expression

ABSTRACT Disulfide bond formation catalyzed by disulfide oxidoreductases occurs in the periplasm and plays a major role in the proper folding and integrity of many proteins. In this study, we were interested in elucidating factors that influence the regulation of dsbA, a gene coding for the primary disulfide oxidoreductase found in Salmonella enterica serovar Typhimurium. Strains with mutations created by transposon mutagenesis were screened for strains with altered expression of dsbA. A mutant (NLM2173) was found where maximal expression of a dsbA::lacZ transcriptional fusion occurred in the exponential growth phase in contrast to that observed in the wild type where maximal expression occurs in stationary phase. Sequence analysis of NLM2173 demonstrated that the transposon had inserted upstream of the gene encoding H-NS. Western immunoblot analysis using H-NS and StpA antibodies showed decreased amounts of H-NS protein in NLM2173, and this reduction in H-NS correlated with an increase of StpA protein. Northern blot analysis with a dsbA-specific probe showed an increase in dsbA transcript during exponential phase of growth. Direct binding of H-NS to the dsbA promoter region was verified using purified H-NS in electrophoretic mobility shift assays. Thus, a reduction in H-NS protein is correlated with a derepression of dsbA in NLM2173, suggesting that H-NS normally plays a role in suppressing the expression of dsbA during exponential phase growth.

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.

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