scholarly journals DNA Microarray Analysis of the Expression Profile of Escherichia coli in Response to Treatment with 4,5-Dihydroxy-2-Cyclopenten-1-One

2002 ◽  
Vol 184 (23) ◽  
pp. 6725-6729 ◽  
Author(s):  
Sangita Phadtare ◽  
Ikunoshin Kato ◽  
Masayori Inouye
Keyword(s):  
Oxidative Stress ◽  
Escherichia Coli ◽  
Antibacterial Activity ◽  
Microarray Analysis ◽  
Dna Microarray ◽  
Cell Metabolism ◽  
Response To Treatment ◽  
E Coli ◽  
Number Of Genes ◽  
Genes Encoding

ABSTRACT We carried out DNA microarray-based global transcript profiling of Escherichia coli in response to 4,5-dihydroxy-2-cyclopenten-1-one to explore the manifestation of its antibacterial activity. We show that it has widespread effects in E. coli affecting genes encoding proteins involved in cell metabolism and membrane synthesis and functions. Genes belonging to the regulon involved in synthesis of Cys are upregulated. In addition, rpoS and RpoS-regulated genes responding to various stresses and a number of genes responding to oxidative stress are upregulated.

DNA Microarray-Based Genomic Characterization of the Pathotypes of Escherichia coli O26, O45, O103, O111, and O145 Isolated from Feces of Feedlot Cattle†

2019 ◽  
Vol 82 (3) ◽  
pp. 395-404 ◽  
Author(s):  
PRAGATHI B. SHRIDHAR ◽  
ISHA R. PATEL ◽  
JAYANTHI GANGIREDLA ◽  
LANCE W. NOLL ◽  
XIAORONG SHI ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Dna Microarray ◽  
Virulence Genes ◽  
Feedlot Cattle ◽  
Clinical Strains ◽  
E Coli ◽  
Type Iii ◽  
Virulence Potential ◽  
Genes Encoding ◽  
Secretory System

ABSTRACT Shiga toxin–producing Escherichia coli (STEC) serogroups O26, O45, O103, O111, O121, and O145, referred to as the top six non-O157 serogroups, are responsible for more than 70% of human non-O157 STEC infections in North America. Cattle harbor non-O157 strains in the hindgut and shed them in the feces. The objective of this study was to use the U.S. Food and Drug Administration (FDA) E. coli identification (ECID) DNA microarray to identify the serotype, assess the virulence potential of each, and determine the phylogenetic relationships among five of the six non-O157 E. coli serogroups isolated from feedlot cattle feces. Forty-four strains of STEC, enterohemorrhagic E. coli (EHEC), enteropathogenic E. coli (EPEC), or putative nonpathotype E. coli (NPEC) of cattle origin and five human clinical strains of EHEC were assayed with the FDA-ECID DNA microarray. The cattle strains harbored diverse flagellar genes. The bovine and human strains belonging to serogroups O26, O45, and O103 carried stx1 only, O111 carried both stx1 and stx2, and O145 carried either stx1 or stx2. The strains were also positive for various subtypes of intimin and other adhesins (IrgA homologue adhesin, long polar fimbriae, mannose-specific adhesin, and curli). Both human and cattle strains were positive for LEE-encoded type III secretory system genes and non–LEE-encoded effector genes. SplitsTree4, a program used to determine the phylogenetic relationship among the strains, revealed that the strains within each serogroup clustered according to their pathotype. In addition to genes encoding Shiga toxins, bovine non-O157 E. coli strains possessed other major virulence genes, including those for adhesins, type III secretory system proteins, and plasmid-borne virulence genes, similar to human clinical strains. Because virulence factors encoded by these genes are involved in the pathogenesis of various pathotypes of E. coli, the bovine non-O157 strains could cause human illness. The FDA-ECID DNA microarray assay rapidly provided a profile of the virulence genes for assessment of the virulence potential of each strain.

scholarly journals The soxRS response of Escherichia coli can be induced in the absence of oxidative stress and oxygen by modulation of NADPH content

Microbiology ◽  
2011 ◽  
Vol 157 (4) ◽  
pp. 957-965 ◽  
Author(s):  
Adriana R. Krapp ◽  
María Victoria Humbert ◽  
Néstor Carrillo
Keyword(s):  
Oxidative Stress ◽  
Escherichia Coli ◽  
Methyl Viologen ◽  
Transcriptional Regulator ◽  
E Coli ◽  
Enhanced Sensitivity ◽  
Genes Encoding ◽  
Cellular Signal ◽  
Redox Shuttle

The soxRS regulon protects Escherichia coli cells against superoxide and nitric oxide. Oxidation of the SoxR sensor, a [2Fe–2S]-containing transcriptional regulator, triggers the response, but the nature of the cellular signal sensed by SoxR is still a matter of debate. In vivo, the sensor is maintained in a reduced, inactive state by the activities of SoxR reductases, which employ NADPH as an electron donor. The hypothesis that NADPH levels affect deployment of the soxRS response was tested by transforming E. coli cells with genes encoding enzymes and proteins that lead to either build-up or depletion of the cellular NADPH pool. Introduction of NADP+-reducing enzymes, such as wheat non-phosphorylating glyceraldehyde-3-phosphate dehydrogenase or E. coli malic enzyme, led to NADPH accumulation, inhibition of the soxRS regulon and enhanced sensitivity to the superoxide propagator methyl viologen (MV). Conversely, expression of pea ferredoxin (Fd), a redox shuttle that can oxidize NADPH via ferredoxin-NADP(H) reductase, resulted in execution of the soxRS response in the absence of oxidative stress, and in higher tolerance to MV. Processes that caused NADPH decline, including oxidative stress and Fd activity, correlated with an increase in total (NADP++NADPH) stocks. SoxS expression can be induced by Fd expression or by MV in anaerobiosis, under conditions in which NADPH is oxidized but no superoxide can be formed. The results indicate that activation of the soxRS regulon in E. coli cells exposed to superoxide-propagating compounds can be triggered by depletion of the NADPH stock rather than accumulation of superoxide itself. They also suggest that bacteria need to finely regulate homeostasis of the NADP(H) pool to enable proper deployment of this defensive response.

scholarly journals Genes of Escherichia coli O157:H7 That Are Involved in High-Pressure Resistance

2006 ◽  
Vol 72 (4) ◽  
pp. 2661-2671 ◽  
Author(s):  
Aaron S. Malone ◽  
Yoon-Kyung Chung ◽  
Ahmed E. Yousef
Keyword(s):  
Escherichia Coli ◽  
High Pressure ◽  
Microarray Analysis ◽  
Dna Microarray ◽  
Spontaneous Mutation ◽  
Cluster Assembly ◽  
E Coli ◽  
Dna Microarray Analysis ◽  
Pressure Resistance ◽  
K 12

ABSTRACT Seventeen Escherichia coli O157:H7 strains were treated with ultrahigh pressure at 500 MPa and 23 ± 2°C for 1 min. This treatment inactivated 0.6 to 3.4 log CFU/ml, depending on the strain. The diversity of these strains was confirmed by pulsed-field gel electrophoresis (PFGE) analysis, and there was no apparent association between PFGE banding patterns and pressure resistance. The pressure-resistant strain E. coli O157:H7 EC-88 (0.6-log decrease) and the pressure-sensitive strain ATCC 35150 (3.4-log decrease) were treated with a sublethal pressure (100 MPa for 15 min at 23 ± 2°C) and subjected to DNA microarray analysis using an E. coli K-12 antisense gene chip. High pressure affected the transcription of many genes involved in a variety of intracellular mechanisms of EC-88, including the stress response, the thiol-disulfide redox system, Fe-S cluster assembly, and spontaneous mutation. Twenty-four E. coli isogenic pairs with mutations in the genes regulated by the pressure treatment were treated with lethal pressures at 400 MPa and 23 ± 2°C for 5 min. The barotolerance of the mutants relative to that of the wild-type strains helped to explain the results obtained by DNA microarray analysis. This study is the first report to demonstrate that the expression of Fe-S cluster assembly proteins and the fumarate nitrate reductase regulator decreases the resistance to pressure, while sigma factor (RpoE), lipoprotein (NlpI), thioredoxin (TrxA), thioredoxin reductase (TrxB), a trehalose synthesis protein (OtsA), and a DNA-binding protein (Dps) promote barotolerance.

Synthesis of Chalcones and Nucleosides Incorporating [1, 3, 4]Oxadiazolenone Core and Evaluation of their Antifungal and Antibacterial Activities

2020 ◽  
Vol 15 (6) ◽  
pp. 665-679
Author(s):  
Alok K. Srivastava ◽  
Lokesh K. Pandey
Keyword(s):  
Escherichia Coli ◽  
Staphylococcus Aureus ◽  
Antibacterial Activity ◽  
Bacillus Subtilis ◽  
Antifungal Activity ◽  
Aspergillus Niger ◽  
Gram Negative ◽  
E Coli ◽  
Gram Positive Bacteria ◽  
Good Antibacterial Activity

Background: [1, 3, 4]oxadiazolenone core containing chalcones and nucleosides were synthesized by Claisen-Schmidt condensation of a variety of benzaldehyde derivatives, obtained from oxidation of substituted 5-(3/6 substituted-4-Methylphenyl)-1, 3, 4-oxadiazole-2(3H)-one and various substituted acetophenone. The resultant chalcones were coupled with penta-O-acetylglucopyranose followed by deacetylation to get [1, 3, 4] oxadiazolenone core containing chalcones and nucleosides. Various analytical techniques viz IR, NMR, LC-MS and elemental analysis were used to confirm the structure of the synthesised compounds.The compounds were targeted against Bacillus subtilis, Staphylococcus aureus and Escherichia coli for antibacterial activity and Aspergillus flavus, Aspergillus niger and Fusarium oxysporum for antifungal activity. Methods: A mixture of Acid hydrazides (3.0 mmol) and N, Nʹ- carbonyl diimidazole (3.3 mmol) in 15 mL of dioxane was refluxed to afford substituted [1, 3, 4]-oxadiazole-2(3H)-one. The resulted [1, 3, 4]- oxadiazole-2(3H)-one (1.42 mmol) was oxidized with Chromyl chloride (1.5 mL) in 20 mL of carbon tetra chloride and condensed with acetophenones (1.42 mmol) to get chalcones 4. The equimolar ratio of obtained chalcones 4 and β -D-1,2,3,4,6- penta-O-acetylglucopyranose in presence of iodine was refluxed to get nucleosides 5. The [1, 3, 4] oxadiazolenone core containing chalcones 4 and nucleosides 5 were tested to determined minimum inhibitory concentration (MIC) value with the experimental procedure of Benson using disc-diffusion method. All compounds were tested at concentration of 5 mg/mL, 2.5 mg/mL, 1.25 mg/mL, 0.62 mg/mL, 0.31 mg/mL and 0.15 mg/mL for antifungal activity against three strains of pathogenic fungi Aspergillus flavus (A. flavus), Aspergillus niger (A. niger) and Fusarium oxysporum (F. oxysporum) and for antibacterial activity against Gram-negative bacterium: Escherichia coli (E. coli), and two Gram-positive bacteria: Staphylococcus aureus (S. aureus) and Bacillus subtilis(B. subtilis). Result: The chalcones 4 and nucleosides 5 were screened for antibacterial activity against E. coli, S. aureus and B. subtilis whereas antifungal activity against A. flavus, A. niger and F. oxysporum. Compounds 4a-t showed good antibacterial activity whereas compounds 5a-t containing glucose moiety showed better activity against fungi. The glucose moiety of compounds 5 helps to enter into the cell wall of fungi and control the cell growth. Conclusion: Chalcones 4 and nucleosides 5 incorporating [1, 3, 4] oxadiazolenone core were synthesized and characterized by various spectral techniques and elemental analysis. These compounds were evaluated for their antifungal activity against three fungi; viz. A. flavus, A. niger and F. oxysporum. In addition to this, synthesized compounds were evaluated for their antibacterial activity against gram negative bacteria E. Coli and gram positive bacteria S. aureus, B. subtilis. Compounds 4a-t showed good antibacterial activity whereas 5a-t showed better activity against fungi.

scholarly journals Proteomic response of Escherichia coli to a membrane lytic and iron chelating truncated Amaranthus tricolor defensin

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Tessa B. Moyer ◽  
Ashleigh L. Purvis ◽  
Andrew J. Wommack ◽  
Leslie M. Hicks
Keyword(s):  
Escherichia Coli ◽  
Antibacterial Activity ◽  
Antimicrobial Peptides ◽  
Mechanism Of Action ◽  
Gram Negative Bacteria ◽  
Amaranthus Tricolor ◽  
Core Motif ◽  
Gram Negative ◽  
E Coli ◽  
Membrane Lysis

Abstract Background Plant defensins are a broadly distributed family of antimicrobial peptides which have been primarily studied for agriculturally relevant antifungal activity. Recent studies have probed defensins against Gram-negative bacteria revealing evidence for multiple mechanisms of action including membrane lysis and ribosomal inhibition. Herein, a truncated synthetic analog containing the γ-core motif of Amaranthus tricolor DEF2 (Atr-DEF2) reveals Gram-negative antibacterial activity and its mechanism of action is probed via proteomics, outer membrane permeability studies, and iron reduction/chelation assays. Results Atr-DEF2(G39-C54) demonstrated activity against two Gram-negative human bacterial pathogens, Escherichia coli and Klebsiella pneumoniae. Quantitative proteomics revealed changes in the E. coli proteome in response to treatment of sub-lethal concentrations of the truncated defensin, including bacterial outer membrane (OM) and iron acquisition/processing related proteins. Modification of OM charge is a common response of Gram-negative bacteria to membrane lytic antimicrobial peptides (AMPs) to reduce electrostatic interactions, and this mechanism of action was confirmed for Atr-DEF2(G39-C54) via an N-phenylnaphthalen-1-amine uptake assay. Additionally, in vitro assays confirmed the capacity of Atr-DEF2(G39-C54) to reduce Fe3+ and chelate Fe2+ at cell culture relevant concentrations, thus limiting the availability of essential enzymatic cofactors. Conclusions This study highlights the utility of plant defensin γ-core motif synthetic analogs for characterization of novel defensin activity. Proteomic changes in E. coli after treatment with Atr-DEF2(G39-C54) supported the hypothesis that membrane lysis is an important component of γ-core motif mediated antibacterial activity but also emphasized that other properties, such as metal sequestration, may contribute to a multifaceted mechanism of action.

scholarly journals Ferric uptake regulator (Fur) reversibly binds a [2Fe-2S] cluster to sense intracellular iron homeostasis in Escherichia coli

2020 ◽  
Vol 295 (46) ◽  
pp. 15454-15463 ◽  
Author(s):  
Chelsey R. Fontenot ◽  
Homyra Tasnim ◽  
Kathryn A. Valdes ◽  
Codrina V. Popescu ◽  
Huangen Ding
Keyword(s):  
Escherichia Coli ◽  
Iron Content ◽  
Iron Homeostasis ◽  
Ferric Uptake Regulator ◽  
Intracellular Iron ◽  
Free Iron ◽  
E Coli ◽  
Genes Encoding ◽  
Fur Protein ◽  
Mutant Cells

The ferric uptake regulator (Fur) is a global transcription factor that regulates intracellular iron homeostasis in bacteria. The current hypothesis states that when the intracellular “free” iron concentration is elevated, Fur binds ferrous iron, and the iron-bound Fur represses the genes encoding for iron uptake systems and stimulates the genes encoding for iron storage proteins. However, the “iron-bound” Fur has never been isolated from any bacteria. Here we report that the Escherichia coli Fur has a bright red color when expressed in E. coli mutant cells containing an elevated intracellular free iron content because of deletion of the iron–sulfur cluster assembly proteins IscA and SufA. The acid-labile iron and sulfide content analyses in conjunction with the EPR and Mössbauer spectroscopy measurements and the site-directed mutagenesis studies show that the red Fur protein binds a [2Fe-2S] cluster via conserved cysteine residues. The occupancy of the [2Fe-2S] cluster in Fur protein is ∼31% in the E. coli iscA/sufA mutant cells and is decreased to ∼4% in WT E. coli cells. Depletion of the intracellular free iron content using the membrane-permeable iron chelator 2,2´-dipyridyl effectively removes the [2Fe-2S] cluster from Fur in E. coli cells, suggesting that Fur senses the intracellular free iron content via reversible binding of a [2Fe-2S] cluster. The binding of the [2Fe-2S] cluster in Fur appears to be highly conserved, because the Fur homolog from Hemophilus influenzae expressed in E. coli cells also reversibly binds a [2Fe-2S] cluster to sense intracellular iron homeostasis.

scholarly journals Detection and Quantification of Superoxide Formed within the Periplasm of Escherichia coli

2006 ◽  
Vol 188 (17) ◽  
pp. 6326-6334 ◽  
Author(s):  
Sergei Korshunov ◽  
James A. Imlay
Keyword(s):  
Escherichia Coli ◽  
Pathogenic Bacteria ◽  
Phagocytic Cells ◽  
Free Living ◽  
E Coli ◽  
Superoxide Formation ◽  
Genes Encoding ◽  
Copper Zinc ◽  
Primary Substrate

ABSTRACT Many gram-negative bacteria harbor a copper/zinc-containing superoxide dismutase (CuZnSOD) in their periplasms. In pathogenic bacteria, one role of this enzyme may be to protect periplasmic biomolecules from superoxide that is released by host phagocytic cells. However, the enzyme is also present in many nonpathogens and/or free-living bacteria, including Escherichia coli. In this study we were able to detect superoxide being released into the medium from growing cultures of E. coli. Exponential-phase cells do not normally synthesize CuZnSOD, which is specifically induced in stationary phase. However, the engineered expression of CuZnSOD in growing cells eliminated superoxide release, confirming that this superoxide was formed within the periplasm. The rate of periplasmic superoxide production was surprisingly high and approximated the estimated rate of cytoplasmic superoxide formation when both were normalized to the volume of the compartment. The rate increased in proportion to oxygen concentration, suggesting that the superoxide is generated by the adventitious oxidation of an electron carrier. Mutations that eliminated menaquinone synthesis eradicated the superoxide formation, while mutations in genes encoding respiratory complexes affected it only insofar as they are likely to affect the redox state of menaquinone. We infer that the adventitious autoxidation of dihydromenaquinone in the cytoplasmic membrane releases a steady flux of superoxide into the periplasm of E. coli. This endogenous superoxide may create oxidative stress in that compartment and be a primary substrate of CuZnSOD.

scholarly journals Escherichia coli Mutants Lacking All Possible Combinations of Eight Penicillin Binding Proteins: Viability, Characteristics, and Implications for Peptidoglycan Synthesis

1999 ◽  
Vol 181 (13) ◽  
pp. 3981-3993 ◽  
Author(s):  
Sylvia A. Denome ◽  
Pamela K. Elf ◽  
Thomas A. Henderson ◽  
David E. Nelson ◽  
Kevin D. Young
Keyword(s):  
Escherichia Coli ◽  
Cell Wall ◽  
Binding Proteins ◽  
Kanamycin Resistance ◽  
Open Reading Frames ◽  
Penicillin Binding Proteins ◽  
E Coli ◽  
Peptidoglycan Biosynthesis ◽  
Kanamycin Resistance Cassette ◽  
Genes Encoding

ABSTRACT The penicillin binding proteins (PBPs) synthesize and remodel peptidoglycan, the structural component of the bacterial cell wall. Much is known about the biochemistry of these proteins, but little is known about their biological roles. To better understand the contributions these proteins make to the physiology ofEscherichia coli, we constructed 192 mutants from which eight PBP genes were deleted in every possible combination. The genes encoding PBPs 1a, 1b, 4, 5, 6, and 7, AmpC, and AmpH were cloned, and from each gene an internal coding sequence was removed and replaced with a kanamycin resistance cassette flanked by two ressites from plasmid RP4. Deletion of individual genes was accomplished by transferring each interrupted gene onto the chromosome of E. coli via λ phage transduction and selecting for kanamycin-resistant recombinants. Afterwards, the kanamycin resistance cassette was removed from each mutant strain by supplying ParA resolvase in trans, yielding a strain in which a long segment of the original PBP gene was deleted and replaced by an 8-bpres site. These kanamycin-sensitive mutants were used as recipients in further rounds of replacement mutagenesis, resulting in a set of strains lacking from one to seven PBPs. In addition, thedacD gene was deleted from two septuple mutants, creating strains lacking eight genes. The only deletion combinations not produced were those lacking both PBPs 1a and 1b because such a combination is lethal. Surprisingly, all other deletion mutants were viable even though, at the extreme, 8 of the 12 known PBPs had been eliminated. Furthermore, when both PBPs 2 and 3 were inactivated by the β-lactams mecillinam and aztreonam, respectively, several mutants did not lyse but continued to grow as enlarged spheres, so that one mutant synthesized osmotically resistant peptidoglycan when only 2 of 12 PBPs (PBPs 1b and 1c) remained active. These results have important implications for current models of peptidoglycan biosynthesis, for understanding the evolution of the bacterial sacculus, and for interpreting results derived by mutating unknown open reading frames in genome projects. In addition, members of the set of PBP mutants will provide excellent starting points for answering fundamental questions about other aspects of cell wall metabolism.

scholarly journals Antibacterial activity of Escherichia coli from squid ink (Loligo sp.) n-Hexane extracts

2021 ◽  
Vol 869 (1) ◽  
pp. 012033
Author(s):  
D R Utami ◽  
I Irwan ◽  
S Agustina ◽  
S Karina ◽  
S Afriani
Keyword(s):  
Escherichia Coli ◽  
Antibacterial Activity ◽  
Secondary Metabolite ◽  
Bioactive Compound ◽  
Inhibition Zone ◽  
Hexane Extract ◽  
Inhibition Zone Diameter ◽  
E Coli ◽  
Hexane Extracts ◽  
Intermediate Concentration

Abstract Squid is one of the export commodities in Indonesia. In general, the use of squid meat, while the ink is only as waste. In fact, Squid ink contain bioactive compound that potential as anti-inflammatory, antihypertensive, anti-diabetic,anti-microbial and anti-malaria agents. The purpose of the study is to determine the types of secondary metabolite compounds contained in n-hexane extract of Loligo sp. ink using maceration method to determine its antibacterial activity against Escherichia coli. The results of secondary metabolite compounds obtained from the n-hexane extract of Loligo sp. ink are alkaloid, saponins, glycosides and phenol. The results of antibacterial test against E. coli using the disc method obtained the average of inhibition zone diameter at the concentration of 4% is 6.3 mm (intermediate), concentration of 8% is 7.83 mm (intermediate), concentration of 16% is 14.5 mm (susceptible) and concentration of 32% is 10.83 mm (intermediate). The antibacterial activity in n-hexane extract of Loligo sp. ink is optimal at the concentration of 16% against E. coli bacteria.

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