scholarly journals A Bacterial Microcompartment Is Used for Choline Fermentation byEscherichia coli536

2018 ◽  
Vol 200 (10) ◽  
Author(s):  
Taylor I. Herring ◽  
Tiffany N. Harris ◽  
Chiranjit Chowdhury ◽  
Sujit Kumar Mohanty ◽  
Thomas A. Bobik
Keyword(s):  
Electron Microscopy ◽  
Heart Disease ◽  
Gene Cluster ◽  
Transcriptional Regulator ◽  
Human Gut ◽  
Content Type ◽  
E Coli ◽  
Bacterial Microcompartment ◽  
New Type ◽  
Insight Into

ABSTRACTBacterial choline degradation in the human gut has been associated with cancer and heart disease. In addition, recent studies found that a bacterial microcompartment is involved in choline utilization byProteusandDesulfovibriospecies. However, many aspects of this process have not been fully defined. Here, we investigate choline degradation by the uropathogenEscherichia coli536. Growth studies indicatedE. coli536 degrades choline primarily by fermentation. Electron microscopy indicated that a bacterial microcompartment was used for this process. Bioinformatic analyses suggested that the choline utilization (cut) gene cluster ofE. coli536 includes two operons, one containing three genes and a main operon of 13 genes. Regulatory studies indicate that thecutXgene encodes a positive transcriptional regulator required for induction of the maincutoperon in response to choline supplementation. Each of the 16 genes in thecutcluster was individually deleted, and phenotypes were examined. ThecutX,cutY,cutF,cutO,cutC,cutD,cutU, andcutVgenes were required for choline degradation, but the remaining genes of thecutcluster were not essential under the conditions used. The reasons for these varied phenotypes are discussed.IMPORTANCEHere, we investigate choline degradation inE. coli536. These studies provide a basis for understanding a new type of bacterial microcompartment and may provide deeper insight into the link between choline degradation in the human gut and cancer and heart disease. These are also the first studies of choline degradation inE. coli536, an organism for which sophisticated genetic analysis methods are available. In addition, thecutgene cluster ofE. coli536 is located in pathogenicity island II (PAI-II536) and hence might contribute to pathogenesis.

scholarly journals A Novel CO-Responsive Transcriptional Regulator and Enhanced H2Production by an Engineered Thermococcus onnurineus NA1 Strain

2014 ◽  
Vol 81 (5) ◽  
pp. 1708-1714 ◽  
Author(s):  
Min-Sik Kim ◽  
Ae Ran Choi ◽  
Seong Hyuk Lee ◽  
Hae-Chang Jung ◽  
Seung Seob Bae ◽  
...  
Keyword(s):  
Production Rate ◽  
Gene Cluster ◽  
Type Strain ◽  
Wild Type Strain ◽  
Regulatory System ◽  
Transcriptional Regulator ◽  
Bioreactor Culture ◽  
Wild Type ◽  
Content Type ◽  
Transcriptional Regulatory

ABSTRACTGenome analysis revealed the existence of a putative transcriptional regulatory system governing CO metabolism inThermococcus onnurineusNA1, a carboxydotrophic hydrogenogenic archaeon. The regulatory system is composed of CorQ with a 4-vinyl reductase domain and CorR with a DNA-binding domain of the LysR-type transcriptional regulator family in close proximity to the CO dehydrogenase (CODH) gene cluster. Homologous genes of the CorQR pair were also found in the genomes ofThermococcusspecies and “CandidatusKorarchaeum cryptofilum” OPF8. In-frame deletion of eithercorQorcorRcaused a severe impairment in CO-dependent growth and H2production. WhencorQandcorRdeletion mutants were complemented by introducing thecorQRgenes under the control of a strong promoter, the mRNA and protein levels of the CODH gene were significantly increased in a ΔCorR strain complemented with integratedcorQR(ΔCorR/corQR↑) compared with those in the wild-type strain. In addition, the ΔCorR/corQR↑strain exhibited a much higher H2production rate (5.8-fold) than the wild-type strain in a bioreactor culture. The H2production rate (191.9 mmol liter−1h−1) and the specific H2production rate (249.6 mmol g−1h−1) of this strain were extremely high compared with those of CO-dependent H2-producing prokaryotes reported so far. These results suggest that thecorQRgenes encode a positive regulatory protein pair for the expression of a CODH gene cluster. The study also illustrates that manipulation of the transcriptional regulatory system can improve biological H2production.

scholarly journals Fur-Dam Regulatory Interplay at an Internal Promoter of the Enteroaggregative Escherichia coli Type VI Secretion sci1 Gene Cluster

2020 ◽  
Vol 202 (10) ◽  
Author(s):  
Yannick R. Brunet ◽  
Christophe S. Bernard ◽  
Eric Cascales
Keyword(s):  
Escherichia Coli ◽  
Gene Cluster ◽  
Gene Clusters ◽  
Secretion System ◽  
Target Cells ◽  
Type Vi Secretion System ◽  
Content Type ◽  
Internal Promoter ◽  
E Coli ◽  
Type Vi Secretion

ABSTRACT The type VI secretion system (T6SS) is a weapon for delivering effectors into target cells that is widespread in Gram-negative bacteria. The T6SS is a highly versatile machine, as it can target both eukaryotic and prokaryotic cells, and it has been proposed that T6SSs are adapted to the specific needs of each bacterium. The expression of T6SS gene clusters and the activation of the secretion apparatus are therefore tightly controlled. In enteroaggregative Escherichia coli (EAEC), the sci1 T6SS gene cluster is subject to a complex regulation involving both the ferric uptake regulator (Fur) and DNA adenine methylase (Dam)-dependent DNA methylation. In this study, an additional, internal, promoter was identified within the sci1 gene cluster using +1 transcriptional mapping. Further analyses demonstrated that this internal promoter is controlled by a mechanism strictly identical to that of the main promoter. The Fur binding box overlaps the −10 transcriptional element and a Dam methylation site, GATC-32. Hence, the expression of the distal sci1 genes is repressed and the GATC-32 site is protected from methylation in iron-rich conditions. The Fur-dependent protection of GATC-32 was confirmed by an in vitro methylation assay. In addition, the methylation of GATC-32 negatively impacted Fur binding. The expression of the sci1 internal promoter is therefore controlled by iron availability through Fur regulation, whereas Dam-dependent methylation maintains a stable ON expression in iron-limited conditions. IMPORTANCE Bacteria use weapons to deliver effectors into target cells. One of these weapons, the type VI secretion system (T6SS), assembles a contractile tail acting as a spring to propel a toxin-loaded needle. Its expression and activation therefore need to be tightly regulated. Here, we identified an internal promoter within the sci1 T6SS gene cluster in enteroaggregative E. coli. We show that this internal promoter is controlled by Fur and Dam-dependent methylation. We further demonstrate that Fur and Dam compete at the −10 transcriptional element to finely tune the expression of T6SS genes. We propose that this elegant regulatory mechanism allows the optimum production of the T6SS in conditions where enteroaggregative E. coli encounters competing species.

scholarly journals Antimicrobial Properties of a Chitosan Dextran-Based Hydrogel for Surgical Use

2011 ◽  
Vol 56 (1) ◽  
pp. 280-287 ◽  
Author(s):  
Manal A. Aziz ◽  
Jaydee D. Cabral ◽  
Heather J. L. Brooks ◽  
Stephen C. Moratti ◽  
Lyall R. Hanton
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Bacterial Species ◽  
Antimicrobial Properties ◽  
Sinus Surgery ◽  
Peptide Bonds ◽  
Content Type ◽  
E Coli ◽  
Transmission Electron

ABSTRACTA chitosan dextran-based (CD) hydrogel, developed for use in endoscopic sinus surgery, was tested for antimicrobial activityin vitroagainst a range of pathogenic microorganisms. The microdilution technique was used to determine minimum inhibitory, minimum bactericidal, and minimum fungicidal concentrations. In addition, the time-kill efficacy of CD hydrogel was determined for two bacterial species. Scanning and transmission electron microscopy were carried out to elucidate the antimicrobial mechanism of this compound. CD hydrogel was found to be effective againstStaphylococcus aureus,Streptococcus pyogenes,Escherichia coli, andClostridium perfringensat its surgical concentration of 50,000 mg/liter. Minimum bactericidal concentrations ranged from 2,000 to 50,000 mg/liter. Dextran aldehyde (DA) was found to be the antimicrobial component of the CD hydrogel with MBC ranging from 2,000 to 32,000 mg/liter.S. aureusappeared to be killed at a slightly faster rate thanE. coli. Candida albicansandPseudomonas aeruginosawere more resistant to CD hydrogel and DA. Scanning and transmission electron microscopy ofE. coliandS. aureusincubated with CD hydrogel and DA alone revealed morphological damage, disrupted cell walls, and loss of cytosolic contents, compatible with the proposed mode of action involving binding to cell wall proteins and disruption of peptide bonds. Motility and chemotaxis tests showedE. colito be inhibited when incubated with DA. The antibacterial activity of CD hydrogel may make it a useful postsurgical aid at other body sites, especially where there is a risk of Gram-positive infections.

scholarly journals AsnB is responsible for peptidoglycan precursor amidation in Clostridium difficile in the presence of vancomycin

Microbiology ◽  
2020 ◽  
Vol 166 (6) ◽  
pp. 567-578 ◽  
Author(s):  
Fariza Ammam ◽  
Delphine Patin ◽  
Héloise Coullon ◽  
Didier Blanot ◽  
Thierry Lambert ◽  
...  
Keyword(s):  
Clostridium Difficile ◽  
Gene Cluster ◽  
Enterococcus Faecalis ◽  
Type Species ◽  
Asparagine Synthetase ◽  
Structure Modification ◽  
Content Type ◽  
E Coli ◽  
Link Type ◽  
Peptidoglycan Structure

Clostridium difficile 630 possesses a cryptic but functional gene cluster vanG Cd homologous to the vanG operon of Enterococcus faecalis . Expression of vanG Cd in the presence of subinhibitory concentrations of vancomycin is accompanied by peptidoglycan amidation on the meso-DAP residue. In this paper, we report the presence of two potential asparagine synthetase genes named asnB and asnB2 in the C. difficile genome whose products were potentially involved in this peptidoglycan structure modification. We found that asnB expression was only induced when C. difficile was grown in the presence of vancomycin, yet independently from the vanG Cd resistance and regulation operons. In addition, peptidoglycan precursors were not amidated when asnB was inactivated. No change in vancomycin MIC was observed in the asnB mutant strain. In contrast, overexpression of asnB resulted in the amidation of most of the C. difficile peptidoglycan precursors and in a weak increase of vancomycin susceptibility. AsnB activity was confirmed in E. coli . In contrast, the expression of the second asparagine synthetase, AsnB2, was not induced in the presence of vancomycin. In summary, our results demonstrate that AsnB is responsible for peptidoglycan amidation of C. difficile in the presence of vancomycin.

scholarly journals Oleate Hydratase in Lactobacillus delbrueckii subsp. bulgaricus LBP UFSC 2230 Catalyzes the Reversible Conversion between Linoleic Acid and Ricinoleic Acid

2021 ◽  
Author(s):  
Gabriela Christina Kuhl ◽  
Ricardo Ruiz Mazzon ◽  
Brenda Lee Simas Porto ◽  
Tâmela Zamboni Madaloz ◽  
Guilherme Razzera ◽  
...  
Keyword(s):  
Linoleic Acid ◽  
Stress Tolerance ◽  
Lactobacillus Delbrueckii ◽  
Content Type ◽  
E Coli ◽  
Enzymatic Mechanism ◽  
The Impact ◽  
Insight Into ◽  
Oleate Hydratase ◽  
Lactobacillus Delbrueckii Subsp

This study provides insight into the enzymatic mechanism of CLA synthesis in L. delbrueckii subsp. bulgaricus and broadens our understanding of the bioconversion of LA and RA by OleH. The impact of OleH on the production of the c 9, t 11 CLA isomer and stress tolerance by E. coli has been assisted.

Localization, Assembly, and Activation of the Escherichia coli Cell Division Machinery

EcoSal Plus ◽  
2021 ◽  
Author(s):  
Petra Anne Levin ◽  
Anuradha Janakiraman
Keyword(s):  
Escherichia Coli ◽  
Cell Division ◽  
Bacterial Cell ◽  
Coli Cell ◽  
Bacterial Cell Division ◽  
Content Type ◽  
E Coli ◽  
Insight Into

Decades of research, much of it in Escherichia coli , have yielded a wealth of insight into bacterial cell division. Here, we provide an overview of the E. coli division machinery with an emphasis on recent findings.

scholarly journals Bactofencin A, a New Type of Cationic Bacteriocin with Unusual Immunity

mBio ◽  
2013 ◽  
Vol 4 (6) ◽  
Author(s):  
Eileen F. O'Shea ◽  
Paula M. O'Connor ◽  
Orla O'Sullivan ◽  
Paul D. Cotter ◽  
R. Paul Ross ◽  
...  
Keyword(s):  
Antimicrobial Peptides ◽  
Gene Cluster ◽  
Bacterial Resistance ◽  
Intestinal Bacteria ◽  
Cationic Antimicrobial Peptides ◽  
Content Type ◽  
Immunity Gene ◽  
New Type ◽  
Porcine Origin ◽  
Bacteriocin Gene

ABSTRACT Bacteriocin production is an important probiotic trait of intestinal bacteria. In this study, we identify a new type of bacteriocin, bactofencin A, produced by a porcine intestinal isolate Lactobacillus salivarius DPC6502, and assess its potency against pathogenic species including Staphylococcus aureus and Listeria monocytogenes. Genome sequencing of the bacteriocin producer revealed bfnA, which encodes the mature and highly basic (pI 10.59), 22-amino-acid defensin-like peptide. Matrix-assisted laser desorption ionization–time of flight (MALDI-TOF) mass spectral analysis determined that bactofencin A has a molecular mass of 2,782 Da and contains two cysteine residues that form an intramolecular disulfide bond. Although an ABC transporter and transport accessory protein were also present within the bacteriocin gene cluster, a classical bacteriocin immunity gene was not detected. Interestingly, a dltB homologue was identified downstream of bfnA. DltB is usually encoded within the dlt operon of many Gram-positive bacteria. It is responsible for d-alanylation of teichoic acids in the cell wall and has previously been associated with bacterial resistance to cationic antimicrobial peptides. Heterologous expression of this gene conferred bactofencin A-specific immunity on sensitive strains of L. salivarius and S. aureus (although not L. monocytogenes), establishing its role in bacteriocin immunity. An analysis of the distribution of bfnA revealed that it was present in four additional isolates derived from porcine origin and absent from five human isolates, suggesting that its distribution is host specific. Given its novelty, we anticipate that bactofencin A represents the prototype of a new class of bacteriocins characterized as being cationic, with a DltB homologue providing a cognate immunity function. IMPORTANCE This study describes the identification, purification, and characterization of bactofencin A, a novel type of bacteriocin produced by L. salivarius DPC6502. Interestingly, bactofencin A is not similar to any other known bacteriocin but instead shares similarity with eukaryotic cationic antimicrobial peptides, and here, we demonstrate that it inhibits two medically significant pathogens. Genome sequence analysis of the producing strain also revealed the presence of an atypical dltB homologue in the bacteriocin gene cluster, which was lacking a classical bacteriocin immunity gene. Furthermore, cloning this gene rendered sensitive strains resistant to the bacteriocin, thereby establishing its role in providing cognate bacteriocin immunity. Four additional L. salivarius isolates, also of porcine origin, were found to contain the bacteriocin biosynthesis genes and successfully produced bactofencin A, while these genes were absent from five human-derived strains investigated.

scholarly journals Shape Changes and Interaction Mechanism of Escherichia coli Cells Treated with Sericin and Use of a Sericin-Based Hydrogel for Wound Healing

2016 ◽  
Vol 82 (15) ◽  
pp. 4663-4672 ◽  
Author(s):  
Rui Xue ◽  
Yalong Liu ◽  
Qingsong Zhang ◽  
Congcong Liang ◽  
Huazhen Qin ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Escherichia Coli ◽  
Cell Membrane ◽  
Interaction Mechanism ◽  
Bacterial Cells ◽  
Content Type ◽  
E Coli ◽  
Membrane Blebbing ◽  
Sterile Gauze

ABSTRACTTo verify the interaction mechanism between sericin andEscherichia coli, especially the morphological and structural changes in the bacterial cells, the antimicrobial activity of sericin againstE. colias a model for Gram-negative bacteria was investigated. The antibacterial activity of sericin onE. coliand the interaction mechanism were investigated in this study by analyzing the growth, integrity, and morphology of the bacterial cells following treatment with sericin. The changes in morphology and cellular compositions of bacterial cells treated with sericin were observed by an inverted fluorescence microscope, scanning electron microscopy, and transmission electron microscopy. Changes in electrical conductivity, total sugar concentration of the broth for the bacteria, and protein expression of the bacteria were determined to investigate the permeability of the cell membrane. A sericin-based hydrogel was prepared for anin vivostudy of wound dressing. The results showed that the antibacterial activity of the hydrogel increased with the increase in the concentration of sericin from 10 g/liter to 40 g/liter. The introduction of sericin induces membrane blebbing ofE. colicells caused by antibiotic action on the cell membrane. The cytoplasm shrinkage phenomenon was accompanied by blurring of the membrane wall boundaries. WhenE. colicells were treated with sericin, release of intracellular components quickly increased. The electrical conductivity assay indicated that the charged ions are reduced after exposure to sericin so that the integrity of the cell membrane is weakened and metabolism is blocked. In addition, sodium dodecyl sulfate-polyacrylamide gel electrophoresis demonstrated that sericin hinders the expression of bacterial protein. Sericin may damage the integrity of the bacterial cell membrane, thereby eventually inhibiting the growth and reproduction ofE. coli. Compared to sterile gauze, the sericin-based hydrogel promoted fibroblast cell proliferation and accelerated the formation of granulation tissues and neovessels.IMPORTANCEThe specific relationship and interaction mechanism between sericin andE. colicells were investigated and elucidated. The results show that after 12 h of treatment, sericin molecules induce membrane blebbing ofE. colicells, and the bacteria show decreases in liquidity and permeability of biological membrane, resulting in alterations in the conductivity of the culture medium and the integrity of the outer membrane. The subsequentin vivoresults demonstrate that the sericin-poly(N-isopropylacrylamide-N,N′-methylene-bis-acrylamide [NIPAm-MBA]) hydrogel accelerated wound healing compared to that with sterile gauze, which is a beneficial result for future applications in clinical medicine and the textile, food, and coating industries.

scholarly journals Pseudomonas chlororaphis Produces Two Distinct R-Tailocins That Contribute to Bacterial Competition in Biofilms and on Roots

2017 ◽  
Vol 83 (15) ◽  
Author(s):  
Robert J. Dorosky ◽  
Jun Myoung Yu ◽  
Leland S. Pierson ◽  
Elizabeth A. Pierson
Keyword(s):  
Gene Cluster ◽  
Gene Clusters ◽  
Associated Bacteria ◽  
Content Type ◽  
Bacterial Competition ◽  
Transmission Electron ◽  
Lysis Cassette ◽  
First Time ◽  
Insight Into

ABSTRACT R-type tailocins are high-molecular-weight bacteriocins that resemble bacteriophage tails and are encoded within the genomes of many Pseudomonas species. In this study, analysis of the P. chlororaphis 30-84 R-tailocin gene cluster revealed that it contains the structural components to produce two R-tailocins of different ancestral origins. Two distinct R-tailocin populations differing in length were observed in UV-induced lysates of P. chlororaphis 30-84 via transmission electron microscopy. Mutants defective in the production of one or both R-tailocins demonstrated that the killing spectrum of each tailocin is limited to Pseudomonas species. The spectra of pseudomonads killed by the two R-tailocins differed, although a few Pseudomonas species were either killed by or insusceptible to both tailocins. Tailocin release was disrupted by deletion of the holin gene within the tailocin gene cluster, demonstrating that the lysis cassette is required for the release of both R-tailocins. The loss of functional tailocin production reduced the ability of P. chlororaphis 30-84 to compete with an R-tailocin-sensitive strain within biofilms and rhizosphere communities. Our study demonstrates that Pseudomonas species can produce more than one functional R-tailocin particle sharing the same lysis cassette but differing in their killing spectra. This study provides evidence for the role of R-tailocins as determinants of bacterial competition among plant-associated Pseudomonas in biofilms and the rhizosphere. IMPORTANCE Recent studies have identified R-tailocin gene clusters potentially encoding more than one R-tailocin within the genomes of plant-associated Pseudomonas but have not demonstrated that more than one particle is produced or the ecological significance of the production of multiple R-tailocins. This study demonstrates for the first time that Pseudomonas strains can produce two distinct R-tailocins with different killing spectra, both of which contribute to bacterial competition between rhizosphere-associated bacteria. These results provide new insight into the previously uncharacterized role of R-tailocin production by plant-associated Pseudomonas species in bacterial population dynamics within surface-attached biofilms and on roots.

scholarly journals Molecular Characterizations of Cytolethal Distending Toxin Produced by Providencia alcalifaciens Strains Isolated from Patients with Diarrhea

2012 ◽  
Vol 80 (4) ◽  
pp. 1323-1332 ◽  
Author(s):  
Ayaka Shima ◽  
Atsushi Hinenoya ◽  
Masahiro Asakura ◽  
Norihiko Sugimoto ◽  
Teizo Tsukamoto ◽  
...  
Keyword(s):  
Gene Cluster ◽  
Eukaryotic Cell ◽  
Nucleotide Sequence Analysis ◽  
M Cell ◽  
Transposase Gene ◽  
Content Type ◽  
Histone H2ax ◽  
E Coli ◽  
Shigella Boydii ◽  
Providencia Alcalifaciens

ABSTRACTCytolethal distending toxins (CDTs), which block eukaryotic cell proliferation by acting as inhibitory cyclomodulins, are produced by diverse groups of Gram-negative bacteria. Active CDT is composed of three polypeptides—CdtA, CdtB, and CdtC—encoded by the genescdtA,cdtB, andcdtC, respectively. We developed a PCR-restriction fragment length polymorphism assay for the detection and differentiation of five alleles ofcdtB(Cdt-I through Cdt-V) inEscherichia coliand used the assay to investigate the prevalence and characteristic of CDT-producingE. coliin children with diarrhea (A. Hinenoya et al., Microbiol. Immunol. 53:206–215, 2009). In these assays, two untypablecdtBgenes were detected and the organisms harboring thecdtBgene were identified asProvidencia alcalifaciens(strains AH-31 and AS-1). Nucleotide sequence analysis of thecdtgene cluster revealed that thecdtA,cdtB, andcdtCgenes ofP. alcalifaciensare of 750, 810, and 549 bp, respectively. To understand the possible horizontal transfer of thecdtgenes among closely related species, the presence ofcdtgenes was screened in variousProvidenciaspp. by colony hybridization assay, and thecdtgene cluster was found in only limited strains ofP. alcalifaciens. Genome walking revealed that thecdtgene cluster ofP. alcalifaciensis located adjacent to a putative transposase gene, suggesting the locus might be horizontally transferable. Interestingly, the CDT ofP. alcalifaciens(PaCDT) showed some homology with the CDT ofShigella boydii. Whereas filter-sterilized lysates of strains AH-31 and AS-1 showed distention of CHO but not of HeLa cells,E. coliCDT-I exhibited distention of both cells. This activity of PaCDT was confirmed by generating recombinant PaCDT protein, which could also be neutralized by rabbit anti-PaCdtB antibody. Furthermore, recombinant PaCDT was found to induce G2/M cell cycle arrest and phosphorylation of host histone H2AX, a sensitive marker of DNA double-strand breaks. To our knowledge, this is the first report showing that certain clinicalP. alcalifaciensstrains could produce variants of the CDTs compared.

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