scholarly journals A Salmonella enterica Serovar Typhimurium Succinate Dehydrogenase/Fumarate Reductase Double Mutant Is Avirulent and Immunogenic in BALB/c Mice

2007 ◽  
Vol 76 (3) ◽  
pp. 1128-1134 ◽  
Author(s):  
Regino Mercado-Lubo ◽  
Eric J. Gauger ◽  
Mary P. Leatham ◽  
Tyrrell Conway ◽  
Paul S. Cohen
Keyword(s):  
Succinate Dehydrogenase ◽  
Salmonella Enterica ◽  
Double Mutant ◽  
Wild Type Strain ◽  
Salmonella Enterica Serovar Typhimurium ◽  
Tca Cycle ◽  
Fumarate Reductase ◽  
Subsequent Infection ◽  
The Tca Cycle ◽  
Serovar Typhimurium

ABSTRACT Previously we showed that the tricarboxylic acid (TCA) cycle operates as a full cycle during Salmonella enterica serovar Typhimurium SR-11 peroral infection of BALB/c mice (M. Tchawa Yimga et al., Infect. Immun. 74:1130-1140, 2006). The evidence was that a ΔsucCD mutant (succinyl coenzyme A [succinyl-CoA] synthetase), which prevents the conversion of succinyl-CoA to succinate, and a ΔsdhCDA mutant (succinate dehydrogenase), which blocks the conversion of succinate to fumarate, were both attenuated, whereas an SR-11 ΔaspA mutant (aspartase) and an SR-11 ΔfrdABCD mutant (fumarate reductase), deficient in the ability to run the reductive branch of the TCA cycle, were fully virulent. In the present study, evidence is presented that a serovar Typhimurium SR-11 ΔfrdABCD ΔsdhCDA double mutant is avirulent in BALB/c mice and protective against subsequent infection with the virulent serovar Typhimurium SR-11 wild-type strain via the peroral route and is highly attenuated via the intraperitoneal route. These results suggest that fumarate reductase, which normally runs in the reductive pathway in the opposite direction of succinate dehydrogenase, can replace it during infection by running in the same direction as succinate dehydrogenase in order to run a full TCA cycle in an SR-11 ΔsdhCDA mutant. The data also suggest that the conversion of succinate to fumarate plays a key role in serovar Typhimurium virulence. Moreover, the data raise the possibility that S. enterica ΔfrdABCD ΔsdhCDA double mutants and ΔfrdABCD ΔsdhCDA double mutants of other intracellular bacterial pathogens with complete TCA cycles may prove to be effective live vaccine strains for animals and humans.

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.

Double deletion of cpxR and tolC significantly increases the susceptibility of Salmonella enterica serovar Typhimurium to colistin

2021 ◽  
Author(s):  
Meng-Ke Zhang ◽  
Meng-Yao Zhang ◽  
Shuo-Bo Liu ◽  
Ying-Ying Yang ◽  
Ya-Jun Zhai ◽  
...  
Keyword(s):  
Salmonella Enterica ◽  
Molecular Mechanisms ◽  
Salmonella Enterica Serovar Typhimurium ◽  
Growth Curves ◽  
Tca Cycle ◽  
Lower Growth ◽  
Double Deletion ◽  
Broth Microdilution Method ◽  
Serovar Typhimurium ◽  
Time Kill

Abstract Background The increasing use of colistin causes a serious breach in our last line of defence against MDR Gram-negative pathogens. Our previous study showed that CpxR overexpression increases the susceptibility of acrB and cpxR double-deleted Salmonella enterica serovar Typhimurium to colistin. Objectives To identify the mechanism of CpxAR and efflux pumps that synergistically enhance the susceptibility of S. Typhimurium to colistin. Methods A series of cpxR- and tolC-deleted mutants and a cpxR-complemented strain from a multidrug-susceptible standard strain of S. Typhimurium (JS) were generated in our previous study. Herein, we investigated the susceptibility of these strains to colistin through the broth microdilution method, time–kill curves and survival assays. Growth curves were measured by OD600 in LB broth, tryptone-soy broth (TSB) and M9-glucose (0.2%) minimal media. Finally, molecular mechanisms underlying the mode of action were elucidated by transcriptomic analysis. Results We found that in contrast to JS (0.8 mg/L), the MIC of colistin for JSΔtolC::kan showed a 16-fold decrease (0.05 mg/L). Notably, JSΔcpxRΔtolC and JSΔcpxRΔtolC/pcpxR were associated with a 256-fold decrease (0.0031 mg/L) compared with JS. Growth curves identified that JSΔcpxRΔtolC and JSΔcpxRΔtolC/pcpxR displayed a markedly lower growth rate and poorer adaptability. In addition, time–kill curves and survival assays showed that JSΔcpxRΔtolC and JSΔcpxRΔtolC/pcpxR were more susceptible to colistin. Lastly, double deletion of cpxR and tolC enhanced oxidative damage through promoting oxidative phosphorylation, the tricarboxylic acid (TCA) cycle and trimethylamine N-oxide (TMAO) respiration. Conclusions Our findings revealed that double deletion of cpxR and tolC significantly increases the susceptibility of S. Typhimurium to colistin.

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 Altering the Length of the Lipopolysaccharide O Antigen Has an Impact on the Interaction of Salmonella enterica Serovar Typhimurium with Macrophages and Complement

2006 ◽  
Vol 188 (7) ◽  
pp. 2735-2739 ◽  
Author(s):  
Gerald L. Murray ◽  
Stephen R. Attridge ◽  
Renato Morona
Keyword(s):  
Vibrio Cholerae ◽  
Salmonella Enterica ◽  
Double Mutant ◽  
Salmonella Enterica Serovar Typhimurium ◽  
Shigella Flexneri ◽  
O Antigen ◽  
Repeat Units ◽  
Complement Resistance ◽  
Serovar Typhimurium ◽  
First Time

ABSTRACT A panel of isogenic Salmonella enterica serovar Typhimurium strains that vary only in the length of the O antigen was constructed through complementation of a wzz double mutant (displaying unregulated O-antigen length) with one of two homologous (wzz ST and wzz fepE) or three heterologous (wzz O139 of Vibrio cholerae and wzz SF and wzz pHS-2 of Shigella flexneri) wzz genes. Each gene was functional in the S. enterica serovar Typhimurium host and specified production of O-antigen polymers with lengths typical of those synthesized by the donor bacteria (ranging from 2 to >100 O-antigen repeat units). By use of this panel of strains, it was found that O-antigen length influences invasion/uptake by macrophage cells; this is the first time this has been shown with Salmonella. O-antigen length was confirmed to be related to complement resistance, with a minimum protective length of >4 and <15 repeat units. O antigen of 16 to 35 repeat units was found to activate complement more efficiently than other lengths, but this was unrelated to complement resistance. No evidence was found to suggest that modifying the length of the O-antigen polymer affected expression of the O1, O4, or O5 antigenic factors.

scholarly journals sciS, an icmF Homolog in Salmonella enterica Serovar Typhimurium, Limits Intracellular Replication and Decreases Virulence

2005 ◽  
Vol 73 (7) ◽  
pp. 4338-4345 ◽  
Author(s):  
Duncan A. Parsons ◽  
Fred Heffron
Keyword(s):  
Legionella Pneumophila ◽  
Salmonella Enterica ◽  
Wild Type Strain ◽  
Salmonella Enterica Serovar Typhimurium ◽  
The Body ◽  
Wild Type ◽  
Serovar Typhimurium ◽  
Intracellular Multiplication ◽  
Late Stages ◽  
Made In

ABSTRACT Salmonella enterica serovar Typhimurium utilizes macrophages to disseminate from the intestine to deeper tissues within the body. While S. enterica serovar Typhimurium has been shown to kill its host macrophage, it can persist intracellularly beyond 18 h postinfection. To identify factors involved in late stages of infection, we screened a transposon library made in S. enterica serovar Typhimurium for the ability to persist in J774 macrophages at 24 h postinfection. Through this screen, we identified a gene, sciS, found to be homologous to icmF in Legionella pneumophila. icmF, which is required for intracellular multiplication, is conserved in several gram-negative pathogens, and its homolog appears to have been acquired horizontally in S. enterica serovar Typhimurium. We found that an sciS mutant displayed increased intracellular numbers in J774 macrophages when compared to the wild-type strain at 24 h postinfection. sciS was maximally transcribed at 27 h postinfection and is repressed by SsrB, an activator of genes required for promoting intracellular survival. Finally, we demonstrate that an sciS mutant is hypervirulent in mice when administered intragastrically. Taken together, these data indicate a role for SciS in controlling intracellular bacterial levels at later stages of infection and attenuating virulence in a murine host

scholarly journals Role of Gluconeogenesis and the Tricarboxylic Acid Cycle in the Virulence of Salmonella enterica Serovar Typhimurium in BALB/c Mice

2006 ◽  
Vol 74 (2) ◽  
pp. 1130-1140 ◽  
Author(s):  
Merlin Tchawa Yimga ◽  
Mary P. Leatham ◽  
James H. Allen ◽  
David C. Laux ◽  
Tyrrell Conway ◽  
...  
Keyword(s):  
Salmonella Enterica ◽  
Tricarboxylic Acid Cycle ◽  
Carbon Sources ◽  
Tca Cycle ◽  
Tricarboxylic Acid ◽  
Glyoxylate Bypass ◽  
The Tca Cycle ◽  
Mouse Tissues ◽  
Serovar Typhimurium ◽  
Genes Encoding

ABSTRACT In Salmonella enterica serovar Typhimurium, the Cra protein (catabolite repressor/activator) regulates utilization of gluconeogenic carbon sources by activating transcription of genes in the gluconeogenic pathway, the glyoxylate bypass, the tricarboxylic acid (TCA) cycle, and electron transport and repressing genes encoding glycolytic enzymes. A serovar Typhimurium SR-11 Δcra mutant was recently reported to be avirulent in BALB/c mice via the peroral route, suggesting that gluconeogenesis may be required for virulence. In the present study, specific SR-11 genes in the gluconeogenic pathway were deleted (fbp, glpX, ppsA, and pckA), and the mutants were tested for virulence in BALB/c mice. The data show that SR-11 does not require gluconeogenesis to retain full virulence and suggest that as yet unidentified sugars are utilized by SR-11 for growth during infection of BALB/c mice. The data also suggest that the TCA cycle operates as a full cycle, i.e., a sucCD mutant, which prevents the conversion of succinyl coenzyme A to succinate, and an ΔsdhCDA mutant, which blocks the conversion of succinate to fumarate, were both attenuated, whereas both an SR-11 ΔaspA mutant and an SR-11 ΔfrdABC mutant, deficient in the ability to run the reductive branch of the TCA cycle, were fully virulent. Moreover, although it appears that SR-11 replenishes TCA cycle intermediates from substrates present in mouse tissues, fatty acid degradation and the glyoxylate bypass are not required, since an SR-11 ΔfadD mutant and an SR-11 ΔaceA mutant were both fully virulent.

scholarly journals Nucleoid-associated protein HU controls three regulons that coordinate virulence, response to stress and general physiology in Salmonella enterica serovar Typhimurium

Microbiology ◽  
2011 ◽  
Vol 157 (4) ◽  
pp. 1075-1087 ◽  
Author(s):  
Michael W. Mangan ◽  
Sacha Lucchini ◽  
Tadhg Ó Cróinín ◽  
Stephen Fitzgerald ◽  
Jay C. D. Hinton ◽  
...  
Keyword(s):  
Salmonella Enterica ◽  
Double Mutant ◽  
Salmonella Enterica Serovar Typhimurium ◽  
Expression Of Genes ◽  
Hu Proteins ◽  
Epithelial Cell Lines ◽  
Serovar Typhimurium ◽  
Associated Proteins ◽  
Response To Stress

The role of the HU nucleoid-associated proteins in gene regulation was examined in Salmonella enterica serovar Typhimurium. The dimeric HU protein consists of different combinations of its α and β subunits. Transcriptomic analysis was performed with cultures growing at 37 °C at 1, 4 and 6 h after inoculation with mutants that lack combinations of HU α and HU β. Distinct but overlapping patterns of gene expression were detected at each time point for each of the three mutants, revealing not one but three regulons of genes controlled by the HU proteins. Mutations in the hup genes altered the expression of regulatory and structural genes in both the SPI1 and SPI2 pathogenicity islands. The hupA hupB double mutant was defective in invasion of epithelial cell lines and in its ability to survive in macrophages. The double mutant also had defective swarming activity and a competitive fitness disadvantage compared with the wild-type. In contrast, inactivation of just the hupB gene resulted in increased fitness and correlated with the upregulation of members of the RpoS regulon in exponential-phase cultures. Our data show that HU coordinates the expression of genes involved in central metabolism and virulence and contributes to the success of S. enterica as a pathogen.

scholarly journals Effect of Inactivation of the HtrA-Like Serine Protease DegQ on the Virulence of Salmonella enterica Serovar Typhimurium in Mice

2004 ◽  
Vol 72 (12) ◽  
pp. 7357-7359 ◽  
Author(s):  
Jacinta Farn ◽  
Mark Roberts
Keyword(s):  
Serine Protease ◽  
Salmonella Enterica ◽  
Type Strain ◽  
Wild Type Strain ◽  
Salmonella Enterica Serovar Typhimurium ◽  
Wild Type ◽  
Virulence Determinant ◽  
Serovar Typhimurium

ABSTRACT DegQ is a serine protease that is highly homologous to HtrA, an important virulence determinant of Salmonella enterica serovar Typhimurium. We examined if DegQ is involved in serovar Typhimurium pathogenesis. A serovar Typhimurium degQ mutant was as virulent as the wild-type strain in mice. However, a serovar Typhimurium htrA degQ mutant survived less well in murine organs, particularly in the liver, than a serovar Typhimurium htrA mutant. DegQ is not essential for serovar Typhimurium pathogenesis but may play a small role during salmonella growth at systemic sites.

scholarly journals The Conserved Cys-X1-X2-Cys Motif Present in the TtcA Protein Is Required for the Thiolation of Cytidine in Position 32 of tRNA from Salmonella enterica serovar Typhimurium

2004 ◽  
Vol 186 (3) ◽  
pp. 750-757 ◽  
Author(s):  
Gunilla Jäger ◽  
Ramune Leipuviene ◽  
Michael G. Pollard ◽  
Qiang Qian ◽  
Glenn R. Björk
Keyword(s):  
Site Selection ◽  
Salmonella Enterica ◽  
Type Strain ◽  
Wild Type Strain ◽  
Salmonella Enterica Serovar Typhimurium ◽  
Mixed Population ◽  
Wild Type ◽  
Serovar Typhimurium ◽  
Trna Species ◽  
A Site

ABSTRACT The modified nucleoside 2-thiocytidine (s2C) has so far been found in tRNA from organisms belonging to the phylogenetic domains Archaea and Bacteria. In the bacteria Escherichia coli and Salmonella enterica serovar Typhimurium, s2C is present in position 32 of only four tRNA species— \batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \(tRNA_{ICG}^{Arg}\) \end{document} , \batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \(tRNA_{CCG}^{Arg}\) \end{document} , \batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \(tRNA_{mnm^{5}UCU}^{Arg}\) \end{document} , and \batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \(tRNA_{GCU}^{Ser}\) \end{document} . An in-frame deletion of an S. enterica gene (designated ttcA, for “two-thio-cytidine”) was constructed, and such a mutant has no detectable s2C in its tRNA. The TtcA protein family is characterized by the existence of both a PP-loop and a Cys-X1-X2-Cys motif in the central region of the protein but can be divided into two distinct groups based on the presence and location of additional Cys-X1-X2-Cys motifs in terminal regions of the sequence. Mutant analysis showed that both cysteines in this central conserved Cys-X1-X2-Cys motif are required for the formation of s2C. The ΔttcA1 mutant grows at the same rate as the congenic wild-type strain, and no growth disadvantage caused by the lack of s2C was observed in a mixed-population experiment. Lack of s2C32 did not reduce the selection rate at the ribosomal aminoacyl-tRNA site (A-site) for \batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \(Arg-tRNA_{ICG}^{Arg}\) \end{document} at any of its cognate CGN codons, whereas A-site selection at AGG by \batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \(Arg-tRNA_{mnm^{5}UCU}^{Arg}\) \end{document} was dependent on the presence of s2C32. The presence of s2C32 in peptidyl- \batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \(tRNA_{CCU}^{Arg}\) \end{document} or in peptidyl- \batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \(tRNA_{mnm^{5}UCU}^{Arg}\) \end{document} interfered with decoding in the A-site. The presence of s2C32 in \batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \(tRNA_{ICG}^{Arg}\) \end{document} decreased the rate of translation of the CGA codon but not that of the CGU codon.

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