Cell envelope analysis of Coxiella Burneti phase I and phase II

1974 ◽  
Vol 20 (10) ◽  
pp. 1465-1470 ◽  
Author(s):  
T. R. Jerrells ◽  
David J. Hinrichs ◽  
L. P. Mallavia
Keyword(s):  
Electron Microscopy ◽  
Phase I ◽  
Phase Ii ◽  
Cell Envelope ◽  
Diaminopimelic Acid ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
Carbohydrate Concentration ◽  
Morphological Studies ◽  
Cell Envelopes

Cell envelopes of Coxiella burneti, Nine Mile, phase I and phase II were examined by electron microscopy and analyzed chemically in an attempt to characterize the phase phenomenon noted in this organism. Electron microscopy of intact organisms as well as cell envelope fractions revealed a morphology similar to many gram-negative bacteria and other rickettsiae. Morphological studies revealed no differences between C. burneti in phase I or II. Chemical analyses revealed a basic composition similar to that reported for members of the genus Rickettsia and many gram-negative bacteria. Cell envelopes of both phase types of this organism (I and II) contained similar amounts of extractable lipid, similar amino acid composition, and the amino sugars glucosamine and muramic acid. Diaminopimelic acid (DAP) was demonstrated in the cell envelope of both phase types. Coxiella burneti phase I differed from the phase II form in carbohydrate composition as well as protein and carbohydrate concentration. Glucuronic acid, glucose, and galactose were found in C. burneti phase I envelopes, whereas only glucose and galactose were present in phase II envelopes.

Fine structural observations of Desulfovibrio desulfuricans

1973 ◽  
Vol 19 (6) ◽  
pp. 753-756
Author(s):  
Terrence M. Hammill ◽  
Geno J. Germano
Keyword(s):  
Electron Microscopy ◽  
Cell Division ◽  
Cell Envelope ◽  
Desulfovibrio Desulfuricans ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
Ultrathin Sections ◽  
Basal Apparatus ◽  
Specialized Region ◽  
Whole Mounts

Glutaraldehyde-fixed, platinum-carbon-shadowed whole mounts, and ultrathin sections of glutaraldehyde-OsO4-fixed cells of Desulfovibrio desulfuricans were observed by electron microscopy. The preparations demonstrated a typical Vibrio form with a single polar flagellum. The cell envelope and the formation of external blebs were shown to be similar to other gram-negative bacteria. The protoplast, apparently devoid of mesosomes or other membranous structures, was densely packed with ribosomes and contained a fibrous nucleoid. A specialized region near the flagellar end of the cell was commonly observed and termed the basal apparatus. Cell division appeared to be by constriction.

scholarly journals The Peptidoglycan Sacculus of Myxococcus xanthus Has Unusual Structural Features and Is Degraded during Glycerol-Induced Myxospore Development

2008 ◽  
Vol 191 (2) ◽  
pp. 494-505 ◽  
Author(s):  
Nhat Khai Bui ◽  
Joe Gray ◽  
Heinz Schwarz ◽  
Peter Schumann ◽  
Didier Blanot ◽  
...  
Keyword(s):  
Fruiting Body ◽  
Bacterial Cell ◽  
Cell Envelope ◽  
Myxococcus Xanthus ◽  
Structural Features ◽  
Diaminopimelic Acid ◽  
Gram Negative Bacteria ◽  
Structural Elements ◽  
Vegetative Cells ◽  
Gram Negative

ABSTRACT Upon nutrient limitation cells of the swarming soil bacterium Myxococcus xanthus form a multicellular fruiting body in which a fraction of the cells develop into myxospores. Spore development includes the transition from a rod-shaped vegetative cell to a spherical myxospore and so is expected to be accompanied by changes in the bacterial cell envelope. Peptidoglycan is the shape-determining structure in the cell envelope of most bacteria, including myxobacteria. We analyzed the composition of peptidoglycan isolated from M. xanthus. While the basic structural elements of peptidoglycan in myxobacteria were identical to those in other gram-negative bacteria, the peptidoglycan of M. xanthus had unique structural features. meso- or ll-diaminopimelic acid was present in the stem peptides, and a new modification of N-acetylmuramic acid was detected in a fraction of the muropeptides. Peptidoglycan formed a continuous, bag-shaped sacculus in vegetative cells. The sacculus was degraded during the transition from vegetative cells to glycerol-induced myxospores. The spherical, bag-shaped coats isolated from glycerol-induced spores contained no detectable muropeptides, but they contained small amounts of N-acetylmuramic acid and meso-diaminopimelic acid.

scholarly journals Autotransporter-Based Antigen Display in Bacterial Ghosts

2014 ◽  
Vol 81 (2) ◽  
pp. 726-735 ◽  
Author(s):  
Anna Hjelm ◽  
Bill Söderström ◽  
David Vikström ◽  
Wouter S. P. Jong ◽  
Joen Luirink ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Flow Cytometry ◽  
Fluorescence Microscopy ◽  
Surface Display ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
Content Type ◽  
E Coli ◽  
Bacterial Ghosts ◽  
Cell Envelopes

ABSTRACTBacterial ghosts are empty cell envelopes of Gram-negative bacteria that can be used as vehicles for antigen delivery. Ghosts are generated by releasing the bacterial cytoplasmic contents through a channel in the cell envelope that is created by the controlled production of the bacteriophage ϕX174 lysis protein E. While ghosts possess all the immunostimulatory surface properties of the original host strain, they do not pose any of the infectious threats associated with live vaccines. Recently, we have engineered theEscherichia coliautotransporter hemoglobin protease (Hbp) into a platform for the efficient surface display of heterologous proteins in Gram-negative bacteria, HbpD. Using theMycobacterium tuberculosisvaccine target ESAT6 (early secreted antigenic target of 6 kDa), we have explored the application of HbpD to decorateE. coliandSalmonellaghosts with antigens. The use of different promoter systems enabled the concerted production of HbpD-ESAT6 and lysis protein E. Ghost formation was monitored by determining lysis efficiency based on CFU, the localization of a set of cellular markers, fluorescence microscopy, flow cytometry, and electron microscopy. Hbp-mediated surface display of ESAT6 was monitored using a combination of a protease accessibility assay, fluorescence microscopy, flow cytometry and (immuno-)electron microscopy. Here, we show that the concerted production of HbpD and lysis protein E inE. coliandSalmonellacan be used to produce ghosts that efficiently display antigens on their surface. This system holds promise for the development of safe and cost-effective vaccines with optimal intrinsic adjuvant activity and exposure of heterologous antigens to the immune system.

Neutrophil pseudoplatelets in subgingival plaque and crevicular fluid samples from periodontal diseased sites

1989 ◽  
Vol 47 ◽  
pp. 862-863 ◽  
Author(s):  
J Hanker ◽  
E.J. Burkes ◽  
G. Greco ◽  
R. Scruggs ◽  
B. Giammara
Keyword(s):  
Electron Microscopy ◽  
Bone Marrow ◽  
Peripheral Blood ◽  
Gingival Crevicular Fluid ◽  
Light And Electron Microscopy ◽  
Subgingival Plaque ◽  
Staining Reaction ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
Crevicular Fluid

The mature neutrophil with a segmented nucleus (usually having 3 or 4 lobes) is generally considered to be the end-stage cell of the neutrophil series. It is usually found as such in the bone marrow and peripheral blood where it normally is the most abundant leukocyte. Neutrophils, however, must frequently leave the peripheral blood and migrate into areas of infection to combat microorganisms. It is in such areas that neutrophils were first observed to fragment to form platelet-size particles some of which have a nuclear lobe. These neutrophil pseudoplatelets (NPP) can readily be distinguished from true platelets because they stain for neutrophil myeloperoxidase. True platelets are not positive in this staining reaction because their peroxidase Is inhibited by glutaraldehyde. Neutrophil pseudoplatelets, as well as neutrophils budding to form NPP, could frequently be observed in peripheral blood or bone marrow samples of leukemia patients. They are much more prominent, however, in smears of inflammatory exudates that contain gram-negative bacteria and in gingival crevicular fluid samples from periodontal disease sites. In some of these samples macrophages ingesting, or which contained, pseudoplatelets could be observed. The myeloperoxidase in the ingested pseudoplatelets was frequently active. Despite these earlier observations we did not expect to find many NPP in subgingival plaque smears from diseased sites. They were first seen by light microscopy (Figs. 1, 3-5) in smears on coverslips stained with the PATS reaction, a variation of the PAS reaction which deposits silver for light and electron microscopy. After drying replicate PATS-stained coverslips with hexamethyldisilazane, they were sputter coated with gold and then examined by the SEI and BEI modes of scanning electron microscopy (Fig. 2). Unstained replicate coverslips were fixed, and stained for the demonstration of myeloperoxidase in budding neutrophils and NPP. Neutrophils, activated macrophages and spirochetes as well as other gram-negative bacteria were also prominent in the PATS stained samples. In replicate subgingival plaque smears stained with our procedure for granulocyte peroxidases only neutrophils, budding neutrophils or NPP were readily observed (Fig. 6).

scholarly journals Long-distance electron transfer in a filamentous Gram-positive bacterium

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Yonggang Yang ◽  
Zegao Wang ◽  
Cuifen Gan ◽  
Lasse Hyldgaard Klausen ◽  
Robin Bonné ◽  
...  
Keyword(s):  
Electron Transfer ◽  
Microbial Fuel Cells ◽  
Extracellular Electron Transfer ◽  
Gram Negative Bacteria ◽  
Positive Bacterium ◽  
Long Distance ◽  
Gram Positive ◽  
Gram Negative ◽  
Force Microscopy ◽  
Cell Envelopes

AbstractLong-distance extracellular electron transfer has been observed in Gram-negative bacteria and plays roles in both natural and engineering processes. The electron transfer can be mediated by conductive protein appendages (in short unicellular bacteria such as Geobacter species) or by conductive cell envelopes (in filamentous multicellular cable bacteria). Here we show that Lysinibacillus varians GY32, a filamentous unicellular Gram-positive bacterium, is capable of bidirectional extracellular electron transfer. In microbial fuel cells, L. varians can form centimetre-range conductive cellular networks and, when grown on graphite electrodes, the cells can reach a remarkable length of 1.08 mm. Atomic force microscopy and microelectrode analyses suggest that the conductivity is linked to pili-like protein appendages. Our results show that long-distance electron transfer is not limited to Gram-negative bacteria.

scholarly journals Antibiotic transport kinetics in Gram-negative bacteria revealed via single-cell uptake analysis and mathematical modelling

2019 ◽  
Author(s):  
Jehangir Cama ◽  
Margaritis Voliotis ◽  
Jeremy Metz ◽  
Ashley Smith ◽  
Jari Iannucci ◽  
...  
Keyword(s):  
Rational Design ◽  
Cell Envelope ◽  
Time Lapse ◽  
Drug Accumulation ◽  
Cell Uptake ◽  
Gram Negative Bacteria ◽  
Label Free ◽  
Transport Pathways ◽  
Gram Negative ◽  
Accumulation Kinetics

AbstractThe double-membrane cell envelope of Gram-negative bacteria is a formidable barrier to intracellular antibiotic accumulation. A quantitative understanding of antibiotic transport in these cells is crucial for drug development, but this has proved elusive due to the complexity of the problem and a dearth of suitable investigative techniques. Here we combine microfluidics and time-lapse auto-fluorescence microscopy to quantify antibiotic uptake label-free in hundreds of individual Escherichia coli cells. By manipulating the microenvironment, we showed that drug (ofloxacin) accumulation is higher in growing versus non-growing cells. Using genetic knockouts, we provide the first direct evidence that growth phase is more important for drug accumulation than the presence or absence of individual transport pathways. We use our experimental results to inform a mathematical model that predicts drug accumulation kinetics in subcellular compartments. These novel experimental and theoretical results pave the way for the rational design of new Gram-negative antibiotics.

Diacylglycerol kinase A is essential for polymyxin resistance provided by EptA, MCR-1 and other lipid A phosphoethanolamine transferases.

2021 ◽  
Author(s):  
Alexandria B. Purcell ◽  
Bradley J. Voss ◽  
M. Stephen Trent
Keyword(s):  
Lipid A ◽  
Cell Envelope ◽  
Diacylglycerol Kinase ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
E Coli ◽  
Severe Reduction ◽  
Bacterial Fitness ◽  
Phosphoethanolamine Transferases ◽  
Kinase A

Gram-negative bacteria utilize glycerophospholipids (GPLs) as phospho-form donors to modify various surface structures. These modifications play important roles in bacterial fitness in diverse environments influencing cell motility, recognition by the host during infection, and antimicrobial resistance. A well-known example is the modification of the lipid A component of lipopolysaccharide by the phosphoethanolamine (pEtN) transferase EptA that utilizes phosphatidyethanoalmine (PE) as the phospho-form donor. Addition of pEtN to lipid A promotes resistance to cationic antimicrobial peptides (CAMPs), including the polymyxin antibiotics like colistin. A consequence of pEtN modification is the production of diacylglycerol (DAG) that must be recycled back into GPL synthesis via the diacylglycerol kinase A (DgkA). DgkA phosphorylates DAG forming phosphatidic acid, the precursor for GPL synthesis. Here we report that deletion of dgkA in polymyxin-resistant E. coli results in a severe reduction of pEtN modification and loss of antibiotic resistance. We demonstrate that inhibition of EptA is regulated post-transcriptionally and is not due to EptA degradation during DAG accumulation. We also show that the inhibition of lipid A modification by DAG is a conserved feature of different Gram-negative pEtN transferases. Altogether, our data suggests that inhibition of EptA activity during DAG accumulation likely prevents disruption of GPL synthesis helping to maintain cell envelope homeostasis.

scholarly journals How the assembly and protection of the bacterial cell envelope depend on cysteine residues

2020 ◽  
Vol 295 (34) ◽  
pp. 11984-11994 ◽  
Author(s):  
Jean-François Collet ◽  
Seung-Hyun Cho ◽  
Bogdan I. Iorga ◽  
Camille V. Goemans
Keyword(s):  
Amino Acid ◽  
Cell Envelope ◽  
Multilayered Structure ◽  
Cysteine Residues ◽  
Permeability Barrier ◽  
Gram Negative Bacteria ◽  
Amino Acid Residues ◽  
Gram Negative ◽  
Oxidative Inactivation ◽  
Peptidoglycan Cell Wall

The cell envelope of Gram-negative bacteria is a multilayered structure essential for bacterial viability; the peptidoglycan cell wall provides shape and osmotic protection to the cell, and the outer membrane serves as a permeability barrier against noxious compounds in the external environment. Assembling the envelope properly and maintaining its integrity are matters of life and death for bacteria. Our understanding of the mechanisms of envelope assembly and maintenance has increased tremendously over the past two decades. Here, we review the major achievements made during this time, giving central stage to the amino acid cysteine, one of the least abundant amino acid residues in proteins, whose unique chemical and physical properties often critically support biological processes. First, we review how cysteines contribute to envelope homeostasis by forming stabilizing disulfides in crucial bacterial assembly factors (LptD, BamA, and FtsN) and stress sensors (RcsF and NlpE). Second, we highlight the emerging role of enzymes that use cysteine residues to catalyze reactions that are necessary for proper envelope assembly, and we also explain how these enzymes are protected from oxidative inactivation. Finally, we suggest future areas of investigation, including a discussion of how cysteine residues could contribute to envelope homeostasis by functioning as redox switches. By highlighting the redox pathways that are active in the envelope of Escherichia coli, we provide a timely overview of the assembly of a cellular compartment that is the hallmark of Gram-negative bacteria.

Protein expression profiles in methicillin-resistant Staphylococcus aureus (MRSA) under effects of subminimal inhibitory concentrations of imipenem

2019 ◽  
Vol 366 (15) ◽  
Author(s):  
Jichun Wang ◽  
Junrui Wang ◽  
Yanyan Wang ◽  
Peng Sun ◽  
Xiaohui Zou ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Staphylococcus Aureus ◽  
Cell Wall ◽  
Cellular Proliferation ◽  
Methicillin Resistant Staphylococcus Aureus ◽  
Expression Profiles ◽  
Gram Negative Bacteria ◽  
Beta Lactam ◽  
Gram Negative ◽  
Methicillin Resistant

ABSTRACT Imipenem is a beta-lactam antibiotic mainly active against gram-negative bacterial pathogens and also could cause cell wall impairment in methicillin-resistant Staphylococcus aureus(MRSA). However, related antibacterial mechanisms of imipenem on MRSA and mixed infections of MRSA and gram-negative bacteria are relatively poorly revealed. This study was to identify proteins in the MRSA response to subminimal inhibitory concentrations (sub-MICs) of imipenem treatment. Our results showed that 240 and 58 different expression proteins (DEPs) in sub-MICs imipenem-treated S3 (a standard MRSA strain) and S23 (a clinical MRSA strain) strains were identified through the isobaric tag for relative and absolute quantitation method when compared with untreated S3 and S23 strains, respectively, which was further confirmed by multiple reactions monitoring. Our result also demonstrated that expressions of multiple DEPs involved in cellular proliferation, metabolism and virulence were significantly changed in S3 and S23 strains, which was proved by gene ontology annotations and qPCR analysis. Further, transmission electron microscopy and scanning electron microscopy analysis showed cell wall deficiency, cell lysis and abnormal nuclear mitosis on S23 strain. Our study provides important information for understanding the antibacterial mechanisms of imipenem on MRSA and for better usage of imipenem on patients co-infected with MRSA and other multidrug-resistant gram-negative bacteria.

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