scholarly journals Release of Compact Nucleoids with Characteristic Shapes from Escherichia coli

2001 ◽  
Vol 183 (17) ◽  
pp. 5041-5049 ◽  
Author(s):  
Steven B. Zimmerman ◽  
Lizabeth D. Murphy
Keyword(s):  
Escherichia Coli ◽  
Genomic Dna ◽  
Phase Contrast ◽  
Cell Envelope ◽  
Simple Procedure ◽  
Sodium Dodecyl ◽  
Cell Lysis ◽  
Exponential Phase ◽  
Stable Form ◽  
General Shape

ABSTRACT The genomic DNA of bacteria is contained in one or a few compact bodies known as nucleoids. We describe a simple procedure that retains the general shape and compaction of nucleoids from Escherichia coli upon cell lysis and nucleoid release from the cell envelope. The procedure is a modification of that used for the preparation of spermidine nucleoids (nucleoids released in the presence of spermidine) (T. Kornberg, A. Lockwood, and A. Worcel, Proc. Natl. Acad. Sci. USA 71:3189–3193, 1974). Polylysine is added to prevent the normal decompaction of nucleoids which occurs upon cell lysis. Nucleoids retained their characteristic shapes in lysates of exponential-phase cells or in lysates of cells treated with chloramphenicol or nalidixate to alter nucleoid morphology. The notably unstable nucleoids of rifampin-treated cells were obtained in compact, stable form in such lysates. Nucleoids released in the presence of polylysine were easily processed and provided well-defined DNA fluorescence and phase-contrast images. Uniform populations of nucleoids retaining characteristic shapes could be isolated after formaldehyde fixation and heating with sodium dodecyl sulfate.

scholarly journals Thermococcus coalescens sp. nov., a cell-fusing hyperthermophilic archaeon from Suiyo Seamount

2005 ◽  
Vol 55 (6) ◽  
pp. 2507-2514 ◽  
Author(s):  
Tomohiko Kuwabara ◽  
Masaomi Minaba ◽  
Yukihiro Iwayama ◽  
Isao Inouye ◽  
Miwako Nakashima ◽  
...  
Keyword(s):  
Cell Fusion ◽  
Genomic Dna ◽  
Cell Envelope ◽  
16S Rrna Gene Sequencing ◽  
Exponential Phase ◽  
Rrna Gene ◽  
Hyperthermophilic Archaeon ◽  
Intercalating Dye ◽  
A Cell ◽  
Rrna Gene Sequencing

A cell-fusing hyperthermophilic archaeon was isolated from hydrothermal fluid obtained from Suiyo Seamount of the Izu-Bonin Arc. The isolate, TS1T, is an irregular coccus, usually 0·5–2 μm in diameter and motile with a polar tuft of flagella. Cells in the exponential phase of growth fused at room temperature in the presence of DNA-intercalating dye to become as large as 5 μm in diameter. Fused cells showed dark spots that moved along in the cytoplasm. Large cells with a similar appearance were also observed upon culture at 87 °C, suggesting the occurrence of similar cell fusions during growth. Transmission electron microscopy revealed that cells in the exponential phase possessed a thin and electron-lucent cell envelope that could be lost subsequently during culture. The fragile cell envelope must be related to cell fusion. The cells grew at 57–90 °C, pH 5·2–8·7 and at NaCl concentrations of 1·5–4·5 %, with the optima being 87 °C, pH 6·5 and 2·5 % NaCl. The isolate was an anaerobic chemo-organotroph that grew on either yeast extract or tryptone as the sole growth substrate. The genomic DNA G+C content was 53·9 mol%. Phylogenetic analysis based on 16S rRNA gene sequencing indicated that the isolate was closely related to Thermococcus species. However, no significant DNA–DNA hybridization was observed between genomic DNA of strain TS1T and phylogenetically related Thermococcus species. We propose that isolate TS1T represents a novel species, Thermococcus coalescens sp. nov., with the name reflecting the cell fusion activity observed in the strain. The type strain is TS1T (=JCM 12540T=DSM 16538T).

Cell lysis directed by σ E in early stationary phase and effect of induction of the rpoE gene on global gene expression in Escherichia coli

Microbiology ◽  
2005 ◽  
Vol 151 (8) ◽  
pp. 2721-2735 ◽  
Author(s):  
Md. Shahinur Kabir ◽  
Daisuke Yamashita ◽  
Satoshi Koyama ◽  
Taku Oshima ◽  
Ken Kurokawa ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Stationary Phase ◽  
Cell Envelope ◽  
Expression System ◽  
Global Gene Expression ◽  
Cell Lysis ◽  
Specific Cell ◽  
Early Stationary Phase ◽  
Gene Encoding ◽  
Number Of Genes

It has been shown that Escherichia coli cells with increased expression of the rpoE gene encoding σ E exhibit enhanced cell lysis in early stationary phase. Further analysis of the lysis phenomenon was performed using a transient expression system of the rpoE gene and by DNA microarray. The former analysis revealed a σ E-directed cell lysis, specific for early stationary phase but not for the exponential phase. The microarray analysis with RNAs from exponential and early stationary phase cells revealed that a large number of genes were up- or down-regulated when the rpoE gene was induced, and that several genes were induced in a phase-specific manner. The upregulated genes include many previously identified σ E regulon genes, suggesting that a large number of genes are under the control of σ E in this organism. These genes are involved in various cellular activities, including the cell envelope, cellular processes, regulatory functions, transport and translation. Genes that are presumably related to phase-specific cell lysis in E. coli are discussed.

scholarly journals An Active Type IV Secretion System Encoded by the F Plasmid Sensitizes Escherichia coli to Bile Salts

2004 ◽  
Vol 186 (16) ◽  
pp. 5202-5209 ◽  
Author(s):  
James E. Bidlack ◽  
Philip M. Silverman
Keyword(s):  
Escherichia Coli ◽  
Bile Salt ◽  
Bile Salts ◽  
Cell Envelope ◽  
Sodium Dodecyl ◽  
Salt Resistance ◽  
Salt Sensitivity ◽  
Plating Efficiency ◽  
Active System ◽  
Type Iv

ABSTRACT F+ strains of Escherichia coli infected with donor-specific bacteriophage such as M13 are sensitive to bile salts. We show here that this sensitivity has two components. The first derives from secretion of bacteriophage particles through the cell envelope, but the second can be attributed to expression of the F genes required for the formation of conjugative (F) pili. The latter component was manifested as reduced or no growth of an F+ strain in liquid medium containing bile salts at concentrations that had little or no effect on the isogenic F− strain or as a reduced plating efficiency of the F+ strain on solid media; at 2% bile salts, plating efficiency was reduced 104-fold. Strains with F or F-like R factors were consistently more sensitive to bile salts than isogenic, plasmid-free strains, but the quantitative effect of bile salts depended on both the plasmid and the strain. Sensitivity also depended on the bile salt, with conjugated bile salts (glycocholate and taurocholate) being less active than unconjugated bile salts (deoxycholate and cholate). F+ cells were also more sensitive to sodium dodecyl sulfate than otherwise isogenic F− cells, suggesting a selectivity for amphipathic anions. A mutation in any but one F tra gene required for the assembly of F pili, including the traA gene encoding F pilin, substantially restored bile salt resistance, suggesting that bile salt sensitivity requires an active system for F pilin secretion. The exception was traW. A traW mutant was 100-fold more sensitive to cholate than the tra+ strain but only marginally more sensitive to taurocholate or glycocholate. Bile salt sensitivity could not be attributed to a generalized change in the surface permeability of F+ cells, as judged by the effects of hydrophilic and hydrophobic antibiotics and by leakage of periplasmic β-lactamase into the medium.

scholarly journals Exopolysaccharide defects cause hyper-thymineless death in Escherichia coli via massive loss of chromosomal DNA and cell lysis

2020 ◽  
Vol 117 (52) ◽  
pp. 33549-33560
Author(s):  
T. V. Pritha Rao ◽  
Andrei Kuzminov
Keyword(s):  
Escherichia Coli ◽  
Cell Envelope ◽  
Glucose Production ◽  
Cell Lysis ◽  
Chromosomal Dna ◽  
Chromosomal Replication ◽  
Enterobacterial Common Antigen ◽  
Acute Starvation ◽  
Thymineless Death ◽  
Dna Loss

Thymineless death in Escherichia coli thyA mutants growing in the absence of thymidine (dT) is preceded by a substantial resistance phase, during which the culture titer remains static, as if the chromosome has to accumulate damage before ultimately failing. Significant chromosomal replication and fragmentation during the resistance phase could provide appropriate sources of this damage. Alternatively, the initial chromosomal replication in thymine (T)-starved cells could reflect a considerable endogenous dT source, making the resistance phase a delay of acute starvation, rather than an integral part of thymineless death. Here we identify such a low-molecular-weight (LMW)-dT source as mostly dTDP-glucose and its derivatives, used to synthesize enterobacterial common antigen (ECA). The thyA mutant, in which dTDP-glucose production is blocked by the rfbA rffH mutations, lacks a LMW-dT pool, the initial DNA synthesis during T-starvation and the resistance phase. Remarkably, the thyA mutant that makes dTDP-glucose and initiates ECA synthesis normally yet cannot complete it due to the rffC defect, maintains a regular LMW-dT pool, but cannot recover dTTP from it, and thus suffers T-hyperstarvation, dying precipitously, completely losing chromosomal DNA and eventually lysing, even without chromosomal replication. At the same time, its ECA+thyA parent does not lyse during T-starvation, while both the dramatic killing and chromosomal DNA loss in the ECA-deficient thyA mutants precede cell lysis. We conclude that: 1) the significant pool of dTDP-hexoses delays acute T-starvation; 2) T-starvation destabilizes even nonreplicating chromosomes, while T-hyperstarvation destroys them; and 3) beyond the chromosome, T-hyperstarvation also destabilizes the cell envelope.

Survival of Low-pH Stress by Escherichia coli O157:H7: Correlation between Alterations in the Cell Envelope and Increased Acid Tolerance

1999 ◽  
Vol 65 (7) ◽  
pp. 3048-3055 ◽  
Author(s):  
Kieran N. Jordan ◽  
Lynn Oxford ◽  
Conor P. O’Byrne
Keyword(s):  
Escherichia Coli ◽  
Stationary Phase ◽  
Cell Envelope ◽  
Polyacrylamide Gel Electrophoresis ◽  
Protein Composition ◽  
Acid Tolerance ◽  
Low Ph ◽  
Exponential Phase ◽  
Viability Loss ◽  
High Level

ABSTRACT Survival of a nontoxigenic isolate of Escherichia coliO157:H7 at low pH (pH 3.0) was examined over prolonged time periods for each of three population types: exponential-phase cells, stationary-phase cells, and acid-adapted exponential-phase cells. In each population, approximately 5 × 104 CFU ml−1 were detected after a 24-h incubation at pH 3.0. Even after 3 days at pH 3.0, significant numbers of survivors from each of the three populations could be detected. The high level of acid tolerance exhibited by these survivors was found to be quickly lost once they were transferred to conditions which permitted growth to resume, indicating that they were not mutants. Proton flux measurements on the three populations of cells revealed that the initial rates of viability loss at pH 3.0 correlated well with net proton accumulation. Cells showing a high initial rate of viability loss (exponential-phase cells) accumulated protons at the highest rate, whereas resistant populations (adapted or stationary-phase cells) accumulated protons only slowly. Differences in the protein composition of the cell envelope between the three populations were studied by two-dimensional polyacrylamide gel electrophoresis. Complex differences in the pattern of proteins expressed by each population were uncovered. The implications of these findings are discussed in the context of a possible model accounting for acid tolerance in this important food-borne pathogen.

scholarly journals Thermococcus celericrescens sp. nov., a fast-growing and cell-fusing hyperthermophilic archaeon from a deep-sea hydrothermal vent

2007 ◽  
Vol 57 (3) ◽  
pp. 437-443 ◽  
Author(s):  
Tomohiko Kuwabara ◽  
Masaomi Minaba ◽  
Noriko Ogi ◽  
Masahiro Kamekura
Keyword(s):  
Surface Layer ◽  
Hydrothermal Vent ◽  
Genomic Dna ◽  
Cell Envelope ◽  
Optimal Growth ◽  
Exponential Phase ◽  
Cell Multiplication ◽  
Rrna Gene ◽  
Hyperthermophilic Archaeon ◽  
Fast Growing

A fast-growing and cell-fusing hyperthermophilic archaeon was isolated from a hydrothermal vent at Suiyo Seamount, Izu-Bonin Arc, Western Pacific Ocean. Strain TS2T is an irregular, motile coccus that is generally 0.7–1.5 μm in diameter and possesses a polar tuft of flagella. In the mid-exponential phase of growth, cells that appeared black under phase-contrast microscopy fused at room temperature in the presence of a DNA-intercalating dye, as previously observed in Thermococcus coalescens. Cell fusion was not observed in later growth phases. Transmission electron microscopy revealed that the cells in the mid-exponential phase had a 5 nm-thick, electron-dense cell envelope that appeared to associate loosely with the cytoplasmic membrane. As the growth stage progressed, a surface layer developed on the membrane under the envelope and the envelope eventually peeled off. These observations suggest that the surface layer prevents the fusion of cells. Cells of strain TS2T grew at 50–85 °C, pH 5.6–8.3 and at NaCl concentrations of 1.0 to 4.5 %, with optimal growth occurring at 80 °C, pH 7.0 and 3.0 % NaCl. Under optimal growth conditions, strain TS2T grew very fast with an apparent doubling time of 20 min. It is suggested that the biosynthesis of the surface layer cannot catch up with cell multiplication in the mid-exponential phase and thus cells without the surface layer are generated. Strain TS2T was an anaerobic chemo-organotroph that grew on either yeast extract or tryptone as the sole growth substrate. The genomic DNA G+C content was 54.6 mol%. Phylogenetic analysis based on 16S rRNA gene sequencing indicated that the isolate belongs to the genus Thermococcus. However, no significant DNA–DNA hybridization was observed between the genomic DNA of strain TS2T and phylogenetically related Thermococcus species. On the basis of this evidence, strain TS2T is proposed to represent a novel species, Thermococcus celericrescens sp. nov., a name chosen to reflect the fast growth of the strain. The type strain is TS2T (=NBRC 101555T=JCM 13640T=DSM 17994T).

scholarly journals Inhibition of Escherichia coli O157:H7 and Salmonella enterica Isolates on Spinach Leaf Surfaces Using Eugenol-Loaded Surfactant Micelles

Foods ◽  
2019 ◽  
Vol 8 (11) ◽  
pp. 575
Author(s):  
Songsirin Ruengvisesh ◽  
Chris R. Kerth ◽  
T. Matthew Taylor
Keyword(s):  
Escherichia Coli ◽  
Salmonella Enterica ◽  
Limit Of Detection ◽  
Sodium Dodecyl ◽  
Microbial Pathogens ◽  
Escherichia Coli O157 ◽  
Human Pathogens ◽  
Surfactant Micelles ◽  
Leaf Surfaces ◽  
Spinach Leaf

Spinach and other leafy green vegetables have been linked to foodborne disease outbreaks of Escherichia coli O157:H7 and Salmonella enterica around the globe. In this study, the antimicrobial activities of surfactant micelles formed from the anionic surfactant sodium dodecyl sulfate (SDS), SDS micelle-loaded eugenol (1.0% eugenol), 1.0% free eugenol, 200 ppm free chlorine, and sterile water were tested against the human pathogens E. coli O157:H7 and Salmonella Saintpaul, and naturally occurring microorganisms, on spinach leaf surfaces during storage at 5 °C over 10 days. Spinach samples were immersed in antimicrobial treatment solution for 2.0 min at 25 °C, after which treatment solutions were drained off and samples were either subjected to analysis or prepared for refrigerated storage. Whereas empty SDS micelles produced moderate reductions in counts of both pathogens (2.1–3.2 log10 CFU/cm2), free and micelle-entrapped eugenol treatments reduced pathogens by >5.0 log10 CFU/cm2 to below the limit of detection (<0.5 log10 CFU/cm2). Micelle-loaded eugenol produced the greatest numerical reductions in naturally contaminating aerobic bacteria, Enterobacteriaceae, and fungi, though these reductions did not differ statistically from reductions achieved by un-encapsulated eugenol and 200 ppm chlorine. Micelles-loaded eugenol could be used as a novel antimicrobial technology to decontaminate fresh spinach from microbial pathogens.

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