scholarly journals Effects of Growth Phase and Temperature onσBActivity within aListeria monocytogenesPopulation: Evidence for RsbV-Independent Activation ofσBat Refrigeration Temperatures

2014 ◽  
Vol 2014 ◽  
pp. 1-11 ◽  
Author(s):  
Marta Utratna ◽  
Eoin Cosgrave ◽  
Claas Baustian ◽  
Rhodri H. Ceredig ◽  
Conor P. O’Byrne
Keyword(s):  
Growth Phase ◽  
Sigma Factor ◽  
Current Model ◽  
Exponential Phase ◽  
Alternative Sigma Factor ◽  
Alternative Route ◽  
Activation Pattern ◽  
Wild Type ◽  
Early Stationary Phase ◽  
Early Exponential Phase

The alternative sigma factorσBofListeria monocytogenesis responsible for regulating the transcription of many of the genes necessary for adaptation to both food-related stresses and to conditions found within the gastrointestinal tract of the host. The present study sought to investigate the influence of growth phase and temperature on the activation ofσBwithin populations ofL.monocytogenesEGD-e wild-type, ΔsigB,and ΔrsbVthroughout growth at both 4°C and 37°C, using a reporter fusion that couples expression of EGFP to the stronglyσB-dependent promoter oflmo2230. A similarσBactivation pattern within the population was observed in wt-egfpat both temperatures, with the highest induction ofσBoccurring in the early exponential phase of growth when the fluorescent population rapidly increased, eventually reaching the maximum in early stationary phase. Interestingly, induction ofσBactivity was heterogeneous, with only a proportion of the cells in the wt-egfppopulation being fluorescent above the background autofluorescence level. Moreover, significant RsbV-independent activation ofσBwas observed during growth at 4°C. This result suggests that an alternative route toσBactivation exists in the absence of RsbV, a finding that is not explained by the current model forσBregulation.

scholarly journals Roles of rpoN, fliA,and flgR in Expression of Flagella inCampylobacter jejuni

2001 ◽  
Vol 183 (9) ◽  
pp. 2937-2942 ◽  
Author(s):  
Aparna Jagannathan ◽  
Chrystala Constantinidou ◽  
Charles W. Penn
Keyword(s):  
Campylobacter Jejuni ◽  
Genome Sequence ◽  
Sigma Factor ◽  
Transcriptional Activator ◽  
Alternative Sigma Factor ◽  
Wild Type ◽  
Electron Microscopic ◽  
Global Regulation ◽  
Mutant Strains ◽  
Microscopic Studies

ABSTRACT Three potential regulators of flagellar expression present in the genome sequence of Campylobacter jejuni NCTC 11168, the genes rpoN, flgR, andfliA, which encode the alternative sigma factor ς54, the ς54-associated transcriptional activator FlgR, and the flagellar sigma factor ς28, respectively, were investigated for their role in global regulation of flagellar expression. The three genes were insertionally inactivated inC. jejuni strains NCTC 11168 and NCTC 11828. Electron microscopic studies of the wild-type and mutant strains showed that therpoN and flgR mutants were nonflagellate and that the fliA mutant had truncated flagella. Immunoblotting experiments with the three mutants confirmed the roles of rpoN, flgR, and fliA in the expression of flagellin.

scholarly journals Enhancing Isoprene Production by Genetic Modification of the 1-Deoxy-d-Xylulose-5-Phosphate Pathway inBacillus subtilis

2011 ◽  
Vol 77 (7) ◽  
pp. 2399-2405 ◽  
Author(s):  
Junfeng Xue ◽  
Birgitte K. Ahring
Keyword(s):  
Sigma Factor ◽  
Type Strain ◽  
Wild Type Strain ◽  
Stress Factors ◽  
Alternative Sigma Factor ◽  
Wild Type ◽  
Gram Positive ◽  
Gram Positive Bacteria ◽  
Overexpression Strain ◽  
As Stress

ABSTRACTTo enhance the production of isoprene, a volatile 5-carbon hydrocarbon, in the Gram-positive spore-forming rod-shaped bacteriumBacillus subtilis, 1-deoxy-d-xylulose-5-phosphate synthase (Dxs) and 1-deoxy-d-xylulose-5-phosphate reductoisomerase (Dxr) were overexpressed inB. subtilisDSM 10. For the strain that overexpresses Dxs, the yield of isoprene was increased 40% over that by the wild-type strain. In the Dxr overexpression strain, the level of isoprene production was unchanged. Overexpression of Dxr together with Dxs showed an isoprene production level similar to that of the Dxs overproduction strain. The effects of external factors, such as stress factors including heat (48°C), salt (0.3 M NaCl), ethanol (1%), and oxidative (0.005% H2O2) stress, on isoprene production were further examined. Heat, salt, and H2O2induced isoprene production; ethanol inhibited isoprene production. In addition, induction and repression effects are independent of SigB, which is the general stress-responsive alternative sigma factor of Gram-positive bacteria.

scholarly journals Comparative studies on Salmonella typhi grown in vivo and in vitro II. The effect of extracts from normal and infected organs on the bactericidal serum action on strains grown in vivo and in vitro

1963 ◽  
Vol 61 (1) ◽  
pp. 21-30 ◽  
Author(s):  
A. L. Olitzki ◽  
Ophira Kaplan
Keyword(s):  
Growth Phase ◽  
Complement System ◽  
Salmonella Typhi ◽  
Exponential Phase ◽  
Antigenic Structure ◽  
Bactericidal Action ◽  
Early Exponential Phase ◽  
Lower Extent

The sensitivity of in vitro- and in vivo-grown strains of S. typhi was determined not only by their antigenic structure, but also by their growth phase. An increased vulnerability to the antibody-complement system has been found in cells during the lag and the early exponential phase, while non-multiplying cells devoid of any nutrient were almost invulnerable to the antibody-complement system. Extracts of organs from normal and infected animals may promote the growth of S. typhi, and, therefore, increase its vulnerability to the antibody-complement system. The majority of extracts from organs of mice infected with strain Ty 2 inhibited markedly the bactericidal action of serum on this strain and, to a lower extent, on strain O 901.

scholarly journals Attenuation of Late-Stage Disease in Mice Infected bythe Mycobacterium tuberculosis Mutant Lacking theSigF Alternate Sigma Factor and Identification ofSigF-Dependent Genes by MicroarrayAnalysis

2004 ◽  
Vol 72 (3) ◽  
pp. 1733-1745 ◽  
Author(s):  
Deborah E. Geiman ◽  
Deepak Kaushal ◽  
Chiew Ko ◽  
Sandeep Tyagi ◽  
Yukari C. Manabe ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Stationary Phase ◽  
Mutant Strain ◽  
Microarray Analysis ◽  
Growth Phase ◽  
Sigma Factor ◽  
Granulomatous Inflammation ◽  
Exponential Growth Phase ◽  
Histopathological Analysis ◽  
Wild Type

ABSTRACT The Mycobacterium tuberculosis alternate sigma factor, SigF, is expressed during stationary growth phase and under stress conditions in vitro. To better understand the function of SigF we studied the phenotype of the M. tuberculosis ΔsigF mutant in vivo during mouse infection, tested the mutant as a vaccine in rabbits, and evaluated the mutant's microarray expression profile in comparison with the wild type. In mice the growth rates of theΔ sigF mutant and wild-type strains were nearly identical during the first 8 weeks after infection. At 8 weeks, theΔ sigF mutant persisted in the lung, while the wild type continued growing through 20 weeks. Histopathological analysis showed that both wild-type and mutant strains had similar degrees of interstitial and granulomatous inflammation during the first 12 weeks of infection. However, from 12 to 20 weeks the mutant strain showed smaller and fewer lesions and less inflammation in the lungs and spleen. Intradermal vaccination of rabbits with the M. tuberculosis ΔsigF strain, followed by aerosol challenge, resulted in fewer tubercles than did intradermal M. bovis BCG vaccination. Complete genomic microarray analysis revealed that 187 genes were relatively underexpressed in the absence of SigF in early stationary phase, 277 in late stationary phase, and only 38 genes in exponential growth phase. Numerous regulatory genes and those involved in cell envelope synthesis were down-regulated in the absence of SigF; moreover, the ΔsigF mutant strain lacked neutral red staining, suggesting a reduction in the expression of envelope-associated sulfolipids. Examination of 5′-untranslated sequences among the downregulated genes revealed multiple instances of a putative SigF consensus recognition sequence: GGTTTCX18GGGTAT. These results indicate that in the mouse the M. tuberculosis ΔsigF mutant strain persists in the lung but at lower bacterial burdens than wild type and is attenuated by histopathologic assessment. Microarray analysis has identified SigF-dependent genes and a putative SigF consensus recognition site.

scholarly journals RpoS-Regulated Genes of Escherichia coli Identified by Random lacZ Fusion Mutagenesis

2004 ◽  
Vol 186 (24) ◽  
pp. 8499-8507 ◽  
Author(s):  
Somalinga R. V. Vijayakumar ◽  
Mark G. Kirchhof ◽  
Cheryl L. Patten ◽  
Herb E. Schellhorn
Keyword(s):  
Escherichia Coli ◽  
Sigma Factor ◽  
Genetic Screen ◽  
The Other ◽  
Alternative Sigma Factor ◽  
Gene Fusions ◽  
Wild Type ◽  
Stress Response Genes ◽  
Independent Gene ◽  
Bacterial Genes

ABSTRACT RpoS is a conserved alternative sigma factor that regulates the expression of many stress response genes in Escherichia coli. The RpoS regulon is large but has not yet been completely characterized. In this study, we report the identification of over 100 RpoS-dependent fusions in a genetic screen based on the differential expression of an operon-lacZ fusion bank in rpoS mutant and wild-type backgrounds. Forty-eight independent gene fusions were identified, including several in well-characterized RpoS-regulated genes, such as osmY, katE, and otsA. Many of the other fusions mapped to genes of unknown function or to genes that were not previously known to be under RpoS control. Based on the homology to other known bacterial genes, some of the RpoS-regulated genes of unknown functions are likely important in nutrient scavenging.

Controlled induction of the RpoS regulon inEscherichia coli, using an RpoS-expressing plasmid

2003 ◽  
Vol 49 (12) ◽  
pp. 733-740 ◽  
Author(s):  
Guozhu Chen ◽  
Herb E Schellhorn
Keyword(s):  
Gene Expression ◽  
Sigma Factor ◽  
Exponential Phase ◽  
Nutrient Deprivation ◽  
Alternative Sigma Factor ◽  
Gram Negative Bacteria ◽  
Inescherichia Coli ◽  
Gram Negative ◽  
Glycogen Accumulation ◽  
Rpos Mrna

RpoS, an alternative sigma factor produced by many Gram-negative bacteria, primarily controls genes that are expressed in stationary phase in response to nutrient deprivation. To test the idea that induction of RpoS in the exponential phase, when RpoS is not normally expressed, increases RpoS-dependent gene expression, we constructed a plasmid carrying the rpoS gene under the control of an IPTG (isopropyl-β-D-thiogalactopyranoside)-inducible T7lac promoter. Northern and Western analyses revealed that levels of RpoS mRNA and protein, respectively, increased in response to the inducer IPTG. Assays of changes in RpoS-dependent functions (catalase activity and glycogen accumulation), confirmed that induced RpoS was functional in exponential phase and was sufficient for the expression of RpoS-dependent functions. Controlled expression of RpoS and RpoS-dependent genes by plasmid-encoded rpoS may thus offer a useful tool for the study of RpoS-dependent gene expression.Key words: RpoS, regulon, gene expression, Escherichia coli.

The role of catalase isozymes in the culturability of the root colonizer Pseudomonas putida after exposure to hydrogen peroxide and antibiotics

1994 ◽  
Vol 40 (5) ◽  
pp. 382-387 ◽  
Author(s):  
Martin G. Klotz ◽  
Anne J. Anderson
Keyword(s):  
Hydrogen Peroxide ◽  
Stationary Phase ◽  
Pseudomonas Putida ◽  
Growth Phase ◽  
Stationary Growth Phase ◽  
Exponential Phase ◽  
Wild Type ◽  
Early Stationary Growth Phase ◽  
Mutant Cells

The culturability of Pseudomonas putida cells after exposure to hydrogen peroxide and antibiotics was correlated with growth-dependent expression of catalase isozymes. Exponential phase wild-type cells, which contained catalase isozyme A, survived a 15-min treatment with less than 4 mM hydrogen peroxide, but were killed by higher concentrations. The culturability of P. putida mutant JIM, which lacked any functional catalase in exponential phase, was reduced by more than 75% after a 15-min exposure to ≥ 0.25 mM hydrogen peroxide. Because submillimolar concentrations of hydrogen peroxide are physiologically relevant in the bacterial cell, our results demonstrate that catalase isozyme A has essential housekeeping functions for growing cultures of P. putida. The accumulation of catalase isozymes B and C during growth into stationary phase coincided with a decrease in the sensitivity of wild-type and JIM cells of P. putida to hydrogen peroxide. Late stationary phase wild-type cells survived a 15-min exposure to even 50 mM hydrogen peroxide and mutant J1M cells survived exposure to 20 mM but not 50 mM hydrogen peroxide. The antibiotics tetracycline and kanamycin, which inhibit protein synthesis, were used to study the role of catalase induction in resistance to hydrogen peroxide. More than 40 and 80% of exponential phase cells of P. putida wild-type and J1M strains, respectively, were rendered nonculturable after a 20-min exposure to 45 μM tetracycline. Surprisingly, stationary phase cells of both P. putida strains were culturable after a 20-min exposure to tetracycline but remained sensitive to kanamycin. Exposure to tetracycline of stationary phase cells did not reduce the resistance of these cells to hydrogen peroxide. Tetracycline but not kanamycin increased the activity of catalase in lysates prepared from P. putida wild-type and mutant cells in early stationary growth phase. At this growth phase, only catalase isozyme B is operational in both strains, which suggests that tetracycline affects the activity of this enzyme.Key words: Pseudomonas putida, antibiotics, catalase, culturability, growth phase.

scholarly journals RpoN Modulates Carbapenem Tolerance in Pseudomonas aeruginosa through Pseudomonas Quinolone Signal and PqsE

2016 ◽  
Vol 60 (10) ◽  
pp. 5752-5764 ◽  
Author(s):  
Darija Viducic ◽  
Keiji Murakami ◽  
Takashi Amoh ◽  
Tsuneko Ono ◽  
Yoichiro Miyake
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Sigma Factor ◽  
Alternative Sigma Factor ◽  
Regulatory Effect ◽  
Rich Medium ◽  
Wild Type ◽  
Transcript Levels ◽  
Content Type ◽  
Pseudomonas Quinolone Signal ◽  
Nutrient Rich Medium

ABSTRACTThe ability ofPseudomonas aeruginosato rapidly modulate its response to antibiotic stress and persist in the presence of antibiotics is closely associated with the process of cell-to-cell signaling. The alternative sigma factor RpoN (σ54) is involved in the regulation of quorum sensing (QS) and plays an important role in the survival of stationary-phase cells in the presence of carbapenems. Here, we demonstrate that a ΔrpoNmutant grown in nutrient-rich medium has increased expression ofpqsA,pqsH, andpqsRthroughout growth, resulting in the increased production of thePseudomonasquinolone signal (PQS). The link betweenpqsAand its role in carbapenem tolerance was studied using a ΔrpoNΔpqsAmutant, in which the carbapenem-tolerant phenotype of the ΔrpoNmutant was abolished. In addition, we demonstrate that another mechanism leading to carbapenem tolerance in the ΔrpoNmutant is mediated throughpqsE. Exogenously supplied PQS abolished the biapenem-sensitive phenotype of the ΔrpoNΔpqsAmutant, and overexpression ofpqsEfailed to alter the susceptibility of the ΔrpoNΔpqsAmutant to biapenem. The mutations in the ΔrpoNΔrhlRmutant and the ΔrpoNΔpqsHmutant led to susceptibility to biapenem. Comparison of the changes in the expression of the genes involved in QS in wild-type PAO1 with their expression in the ΔrpoNmutant and the ΔrpoNmutant-derived strains demonstrated the regulatory effect of RpoN on the transcript levels ofrhlR,vqsR, andrpoS. The findings of this study demonstrate that RpoN negatively regulates the expression of PQS in nutrient-rich medium and provide evidence that RpoN interacts withpqsA,pqsE,pqsH, andrhlRin response to antibiotic stress.

scholarly journals The Staphylococcus aureus lrgAB Operon Modulates Murein Hydrolase Activity and Penicillin Tolerance

2000 ◽  
Vol 182 (7) ◽  
pp. 1794-1801 ◽  
Author(s):  
Kajetan H. Groicher ◽  
Brian A. Firek ◽  
David F. Fujimoto ◽  
Kenneth W. Bayles
Keyword(s):  
Staphylococcus Aureus ◽  
Regulatory System ◽  
Cell Lysis ◽  
Negative Control ◽  
Exponential Phase ◽  
Hydrolase Activity ◽  
Wild Type ◽  
Early Exponential Phase ◽  
Associated Proteins ◽  
Murein Hydrolase

ABSTRACT Previous studies in our laboratory have shown that theStaphylococcus aureus LytSR two-component regulatory system affects murein hydrolase activity and autolysis. A LytSR-regulated dicistronic operon has also been identified and shown to encode two potential membrane-associated proteins, designated LrgA and LrgB, hypothesized to be involved in the control of murein hydrolase activity. In the present study, a lrgAB mutant strain was generated and analyzed to test this hypothesis. Zymographic and quantitative analysis of murein hydrolase activity revealed that thelrgAB mutant produced increased extracellular murein hydrolase activity compared to that of the wild-type strain. Complementation of the lrgAB defect by providing thelrgAB genes in trans restored the wild-type phenotype, indicating that these genes confer negative control on extracellular murein hydrolase activity. In addition to these effects, the influence of the lrgAB mutation on penicillin-induced lysis and killing was examined. These studies demonstrated that thelrgAB mutation enhanced penicillin-induced killing of cells approaching the stationary phase of growth, the time at which thelrgAB operon was shown to be maximally expressed. This effect of the lrgAB mutation on penicillin-induced killing was shown to be independent of cell lysis. In contrast, thelrgAB mutation did not affect penicillin-induced killing of cells growing in early-exponential phase, a time in whichlrgAB expression was shown to be minimal. However, expression of the lrgAB operon in early-exponential-phase cells inhibited penicillin-induced killing, again independent of cell lysis. The data generated by this study suggest that penicillin-induced killing of S. aureus involves a novel regulator of murein hydrolase activity.

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