[Show abstract][Hide abstract] ABSTRACT: A marine, magnetotactic bacterium, designated strain MMS-1(T), was isolated from mud and water from a salt marsh in Woods Hole, Massachusetts, USA, after enrichment in defined oxygen-concentration/redox-gradient medium. Strain MMS-1(T) is an obligate microaerophile capable of chemoorganoheterotrophic and chemolithoautotrophic growth. Optimal growth occurred at pH 7.0 and 24-26 °C. Chemolithoautotrophic growth occurred with thiosulfate as the electron donor and autotrophic carbon fixation was via the Calvin-Benson-Bassham cycle. The G+C content of the DNA of strain MMS-1(T) was 47.2 mol%. Cells were Gram-negative and morphologically variable, with shapes that ranged from that of a lima bean to fully helical. Cells were motile by means of a single flagellum at each end of the cell (amphitrichous). Regardless of whether grown in liquid or semi-solid cultures, strain MMS-1(T) displayed only polar magnetotaxis and possessed a single chain of magnetosomes containing elongated octahedral crystals of magnetite, positioned along the long axis of the cell. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain MMS-1(T) belongs to the family Rhodospirillaceae within the Alphaproteobacteria, and is distantly related to species of the genus Magnetospirillum. Strain MMS-1(T) is therefore considered to represent a novel species of a new genus, for which the name Magnetospira thiophila gen. nov., sp. nov. is proposed. The type strain of Magnetospira thiophila is MMS-1(T) ( = ATCC BAA-1438(T) = JCM 17960(T)).
INTERNATIONAL JOURNAL OF SYSTEMATIC AND EVOLUTIONARY MICROBIOLOGY 12/2011; 62(Pt 10):2443-50. · 2.11 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Scanning tunnelling microscopy (STM) images of Pt/Ir- and Pt/Ir/C-coated cell plugs of Methanospirillum hungatei showed paracrystalline structures with P6 symmetry and an 18-nm lattice constant, in agreement with electron microscopy studies. The three-dimensional STM images unambiguously distinguished the two morphologically different proteinaceous plug assemblies and led to an improved understanding of the natural internal organization of whole plugs. Tip convolution effects and the grain size of the metal coating complicated interpretation of finer structures. We discuss possible imaging mechanisms to explain observations in which part of the film was removed but the remaining part of the structure was still imaged reproducibly.
[Show abstract][Hide abstract] ABSTRACT: The frequency of intracytoplasmic membranes in several methanogens grown on H2–CO2 varied with the conditions of growth and varied from one strain to another. Methanobacterium thermoautotrophicum often generated large numbers of intracytoplasmic membranes, while Methanospirillum hungatei produced these membranes only rarely. Conditions allowing for rapid growth, including optimal temperature and high agitation rates, increased the production of intracytoplasmic membranes. These membranes consisted mainly of vesicles composed of one or several membrane layers, often positioned in the central region of the cytoplasm. Several mesophilic methanogens could be grown such that intracytoplasmic membranes were rarely or never observed in thin section or in replicas of cross-fractures from frozen cells. Since high rates of methane synthesis still occurred in these cultures, it follows that the intracytoplasmic membrane system is not a necessary organelle for methane formation in these strains. Negative staining for electron microscopy is not an accurate method to visualize intracytoplasmic membranes in these bacterial cells.
[Show abstract][Hide abstract] ABSTRACT: Adherent epilithic microorganisms recovered from rocks submerged 10 to 20 cm in two different rivers were examined by electron microscopy and enumerated after dispersion in M-9 salts by viable plate counts. Bacterial cells concentrated in microcolonies were often observed attached to the surface of algae, cyanobacteria, and organic detritus. This structured communal mode of growth was common among epilithic microbial communities of different rock types. However, counts of heterotrophic bacteria from limestone (106 to 107 cfu/cm2) were 10- to 100-fold greater than corresponding values from granite, gabbro, rhyolite, basalt, and quartz. Cyanobacteria and algae were an order of magnitude less abundant compared with their bacterial counterparts. These variations in population densities of epilithic microorganisms present on different rocks were inversely related to mineral substrate hardness.Key words: epilithic microorganisms, mineral hardness.
Canadian Journal of Microbiology 02/2011; 35(7):744-747. · 1.20 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Methanospirillum hungatei GP1 consists of chains of rod-shaped cells separated from one another by "cell spacers" comprised of two spacer plugs sandwiching a loose, amorphous material. The chain is encased within a highly ordered sheath to form a cylindrical, multicelled filament about 5 to 10 cells long under our growth conditions. Cells within the filament divide by septation in a manner similar to gram-positive eubacteria; the plasma membrane and wall grow inward to partition the cell in two. Yet, unlike gram-positive eubacteria, the wall is flexible, since cells round up when extruded from the sheath; the shape-maintaining structures are the sheath and spacer plugs. After septation and daughter cell separation, the cell spacer grows between the new cells. Initially, the growth of a spacer plug is detected by electron microscopy as the addition of electron-dense layers, exhibiting an 18.0-nm periodicity, at the surface of one new cell pole. Usually three layers develop at this pole before plug assembly is initiated at the opposite pole. As assembly proceeds, the two newly formed plugs separate from each other to form the loose, amorphous central zone of the spacer. Presumably, cell and cell spacer elongation requires sheath extension, since filament growth is observed. The amorphous cell spacer zone continues to expand as the spacer grows larger until lesions appear in the sheath near the zone's midpoint. Usually, the largest spacer zones are found towards each filament's centre and the lesions split the chain in two. Consequently, M. hungatei requires two separate events for filament division: cell replication which is a septation process and filament splitting which is a "cell spacer" breakage.
[Show abstract][Hide abstract] ABSTRACT: On Ellesmere Island, in the Canadian Arctic, dark-colored biofilms proliferate on moist surfaces, including exposed granodiorite outcrops. Transmission electron microscopy of these biofilms indicates that complex epilithic microbial communities developed, consisting of cyanobacteria and fungi symbiotically associated in a lichen, along with a consortium of free-living algae and gram-negative bacteria. The epilithic cyanobacteria and bacteria were shown to extensively precipitate phosphatic minerals, ranging from relatively large polyphosphate granules (approximately 250 nm in diameter) within their cytoplasmic membranes to smaller iron phosphate grains (generally less than 50 nm in diameter) associated with the periplasmic space and encompassing capsule. Complete encrustation of some bacterial cells by the iron phosphates was observed. Energy-dispersive X-ray spectroscopy suggested that these grains are compositionally similar to the mineral strengite (FePO4∙2H2O). This study clearly indicates that the Arctic supports a thriving microbial community that influences the biogeochemical cycling of PO4 in an environment of low nutrient availability. Nutritional requirements by the microorganisms were actively maintained through a relatively closed recycling mechanism, which restricted the immediate loss of phosphorus from the biofilm.
Canadian Journal of Earth Sciences 02/2011; 31(8):1320-1324. · 1.37 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: All biomass contains a significant quantity of metallic constituents, and mineralization in living and dead biodebris may contribute to element transport from the hydrosphere into sediments. The anionic cell walls of bacteria are remarkable in their ability to fix metals and provide sites for nucleation and growth of minerals. Results presented show the types of cell wall polymers that are responsible for metal binding in walls of Gram-positive and Gram-negative bacteria.
Canadian Journal of Earth Sciences 02/2011; 22(12):1893-1898. · 1.37 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Environmental growth conditions and cell physiology have the potential to influence bacterial surface-metal interactions in both planktonic and biofilm systems. Here, Pseudomonas aeruginosa was studied to determine the influence of these factors (pH, redox potential, and active respiration) on surface electrostatics and metal immobilization. Acid-base titrations revealed a decrease in ionizable ligands at pKa 5 (putative carboxyls) in cells grown below pH 6.2 and in cells grown anaerobically relative to cells grown under oxic and circumneutral pH conditions. This observation correlates with Western immunoblotting assays that revealed a reduction in carboxylated B-band lipopolysaccharide in these cells. Furthermore, spectrophotometric analysis revealed a decrease in zinc, copper, and iron immobilization in these cells, suggesting that lipopolysaccharide modification in response to environmental stimuli influences metal binding. The effect of active versus inactive metabolism on metal adsorption was also examined using respiration inhibitors carbonyl cyanide m-chlorophenylhydrazone and sodium azide. Cells treated with these compounds bound more zinc, copper, and iron than untreated controls, suggesting proton extrusion through respiration competes with metal cations for reactive groups on the cell surface. Accumulation of gold did not show the same trend, and transmission electron microscopy studies confirmed it was not a surface-mediated process. These results suggest that variations in growth environment and cell physiology influence metal accumulation by bacterial cell surfaces and may help to explain discontinuous accumulation of metal observed throughout microbial communities.
Canadian Journal of Microbiology 07/2010; 56(7):527-38. · 1.20 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: A thermophilic bacterium, designated strain CR11(T), was isolated from a filamentous sample collected from a terrestrial hot spring on the south-western foothills of the Rincón volcano in Costa Rica. The Gram-negative cells are approximately 2.4-3.9 microm long and 0.5-0.6 microm wide and are motile rods with polar flagella. Strain CR11(T) grows between 65 and 85 degrees C (optimum 75 degrees C, doubling time 4.5 h) and between pH 4.8 and 7.8 (optimum pH 5.9-6.5). The isolate grows chemolithotrophically with S(0), S(2)O(2)(3)(-) or H(2) as the electron donor and with O(2) (up to 16 %, v/v) as the sole electron acceptor. The isolate can grow on mannose, glucose, maltose, succinate, peptone, Casamino acids, starch, citrate and yeast extract in the presence of oxygen (4 %) and S(0). Growth occurs only at NaCl concentrations below 0.4 % (w/v). The G+C content of strain CR11(T) is 40.3 mol%. Phylogenetic analysis of the 16S rRNA gene sequence places the strain as a close relative of Thermocrinis ruber OC 1/4(T) (95.7 % sequence similarity). Based on phylogenetic and physiological characteristics, we propose the name Thermocrinis minervae sp. nov., with CR11(T) (=DSM 19557(T) =ATCC BAA-1533(T)) as the type strain.
International journal of systematic and evolutionary microbiology 09/2009; 60(Pt 2):338-43. · 2.11 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Bacterial biofilms are responsible for the majority of all microbial infections and have profound impact on industrial and geochemical processes. While many studies documented phenotypic differentiation and gene regulation of biofilms, the importance of their structural and mechanical properties is poorly understood. Here we investigate how changes in lipopolysaccharide (LPS) core capping in Pseudomonas aeruginosa affect biofilm structure through modification of adhesive, cohesive, and viscoelastic properties at an early stage of biofilm development. Microbead force spectroscopy and atomic force microscopy were used to characterize P. aeruginosa biofilm interactions with either glass substrata or bacterial lawns. Using isogenic migA, wapR, and rmlC mutants with defined LPS characteristics, we observed significant changes in cell mechanical properties among these strains compared to wild-type strain PAO1. Specifically, truncation of core oligosaccharides enhanced both adhesive and cohesive forces by up to 10-fold, whereas changes in instantaneous elasticity were correlated with the presence of O antigen. Using confocal laser scanning microscopy to quantify biofilm structural changes with respect to differences in LPS core capping, we observed that textural parameters varied with adhesion or the inverse of cohesion, while areal and volumetric parameters were linked to adhesion, cohesion, or the balance between them. In conclusion, this report demonstrated for the first time that changes in LPS expression resulted in quantifiable cellular mechanical changes that were correlated with structural changes in bacterial biofilms. Thus, the interplay between architectural and functional properties may be an important contributor to bacterial community survival.
Journal of bacteriology 09/2009; 191(21):6618-31. · 3.94 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Lipopolysaccharide (LPS) monolayers deposited on planar, hydrophobic substrates were used as a defined model of outer membranes of Pseudomonas aeruginosa strain dps 89. To investigate the influence of ions on the (out-of-plane) monolayer structure, we measured specular X-ray reflectivity at high energy (22 keV) to ensure transmission through water. Electron density profiles were reconstructed from the reflectivity curves, and they indicate that the presence of Ca(2+) ions induces a significant change in the conformation of the charged polysaccharide head groups (O-side chains). Monte Carlo simulations based on a minimal computer model of LPS molecules allow for the modelling of 100 or more molecules over 10(-3) s and theoretically explained the tendency found by experiments.
Journal of The Royal Society Interface 08/2009; 6 Suppl 5:S671-8. · 4.91 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The biofilm matrix contributes to the chemistry, structure, and function of biofilms. Biofilm-derived membrane vesicles (MVs) and DNA, both matrix components, demonstrated concentration-, pH-, and cation-dependent interactions. Furthermore, MV-DNA association influenced MV surface properties. This bears consequences for the reactivity and availability for interaction of matrix polymers and other constituents.
Journal of bacteriology 06/2009; 191(13):4097-102. · 3.94 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: There have been considerable strides made in the characterization of the dispensability of teichoic acid biosynthesis genes in recent years. A notable omission thus far has been an early gene in teichoic acid synthesis encoding the N-acetylmannosamine transferase (tagA in Bacillus subtilis; tarA in Staphylococcus aureus), which adds N-acetylmannosamine to complete the synthesis of undecaprenol pyrophosphate-linked disaccharide. Here, we show that the N-acetylmannosamine transferases are dispensable for growth in vitro, making this biosynthetic enzyme the last dispensable gene in the pathway, suggesting that tagA (or tarA) encodes the first committed step in wall teichoic acid synthesis.
Journal of bacteriology 05/2009; 191(12):4030-4. · 3.94 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Bacterial biofilms are the most prevalent mode of bacterial growth in nature. Adhesive and viscoelastic properties of bacteria play important roles at different stages of biofilm development. Following irreversible attachment of bacterial cells onto a surface, a biofilm can grow in which its matrix viscoelasticity helps to maintain structural integrity, determine stress resistance, and control ease of dispersion. In this study, a novel application of force spectroscopy was developed to characterize the surface adhesion and viscoelasticity of bacterial cells in biofilms. By performing microbead force spectroscopy with a closed-loop atomic force microscope, we accurately quantified these properties over a defined contact area. Using the model gram-negative bacterium Pseudomonas aeruginosa, we observed that the adhesive and viscoelastic properties of an isogenic lipopolysaccharide mutant wapR biofilm were significantly different from those measured for the wild-type strain PAO1 biofilm. Moreover, biofilm maturation in either strain also led to prominent changes in adhesion and viscoelasticity. To minimize variability in force measurements resulting from experimental parameter changes, we developed standardized conditions for microbead force spectroscopy to enable meaningful comparison of data obtained in different experiments. Force plots measured under standard conditions showed that the adhesive pressures of PAO1 and wapR early biofilms were 34 +/- 15 Pa and 332 +/- 47 Pa, respectively, whereas those of PAO1 and wapR mature biofilms were 19 +/- 7 Pa and 80 +/- 22 Pa, respectively. Fitting of creep data to a Voigt Standard Linear Solid viscoelasticity model revealed that the instantaneous and delayed elastic moduli in P. aeruginosa were drastically reduced by lipopolysaccharide deficiency and biofilm maturation, whereas viscosity was decreased only for biofilm maturation. In conclusion, we have introduced a direct biophysical method for simultaneously quantifying adhesion and viscoelasticity in bacterial biofilms under native conditions. This method could prove valuable for elucidating the contribution of genetic backgrounds, growth conditions, and environmental stresses to microbial community physiology.
[Show abstract][Hide abstract] ABSTRACT: Searles Lake occupies a closed basin harboring salt-saturated, alkaline brines that have exceptionally high concentrations of arsenic oxyanions. Strain SLAS-1(T) was previously isolated from Searles Lake (R. S. Oremland, T. R. Kulp, J. Switzer Blum, S. E. Hoeft, S. Baesman, L. G. Miller, and J. F. Stolz, Science 308:1305-1308, 2005). We now describe this extremophile with regard to its substrate affinities, its unusual mode of motility, sequenced arrABD gene cluster, cell envelope lipids, and its phylogenetic alignment within the order Halanaerobacteriales, assigning it the name "Halarsenatibacter silvermanii" strain SLAS-1(T). We also report on the substrate dynamics of an anaerobic enrichment culture obtained from Searles Lake that grows under conditions of salt saturation and whose members include a novel sulfate reducer of the order Desulfovibriales, the archaeon Halorhabdus utahensis, as well as a close homolog of strain SLAS-1(T).
Applied and Environmental Microbiology 03/2009; 75(7):1950-60. · 3.95 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Spirochaeta aurantia is a free-living saprophytic spirochete that grows easily in simple laboratory media, and thus can be used as a model for the investigation of surface carbohydrate structures in spirochetae, which are normally not available in sufficient amounts. Freeze-substitution electron microscopy indicated the presence of a capsule-like material projecting from the surface of S. aurantia. Extraction of cells gave two major glycolipids, the one with a higher molecular mass glycolipid was designated large glycolipid A (LGLA). LGLA contained small amount of branched and unsaturated O-linked fatty acids, L: -rhamnose, L: -fucose, D: -xylose, D: -mannose, D: -glucosamine, D: -glycero-D: -gluco-heptose (DDglcHep), D: -glycero-D: -manno-heptose (DDHep), and a novel branched tetradeoxydecose monosaccharide, which we proposed to call aurantose (Aur). The carbohydrate structure of LGLA was extremely complex and consisted of the repeating units built of 11 monosaccharides, arrangement of nine of them was determined as: - [- 3 - beta - DDglcHep - 3 - beta - D - GlcNAc - 2 - beta - D - Man - ] - which wasdeduced from the NMR and chemical data on the LGLA and its fragments, obtained by various degradations. Tentative position of two remaining sugars is proposed. LGLA was negative for gelation of Limulus amebocyte lysate, did not contain lipid A, and was unable to activate any known Toll-like receptors.
[Show abstract][Hide abstract] ABSTRACT: Slow-growing bacteria and biofilms are notoriously tolerant to antibiotics. Oritavancin is a lipoglycopeptide with multiple mechanisms of action that contribute to its bactericidal action against exponentially growing gram-positive pathogens, including the inhibition of cell wall synthesis and perturbation of membrane barrier function. We sought to determine whether oritavancin could eradicate cells known to be tolerant to many antimicrobial agents, that is, stationary-phase and biofilm cultures of Staphylococcus aureus in vitro. Oritavancin exhibited concentration-dependent bactericidal activity against stationary-phase inocula of methicillin-susceptible S. aureus (MSSA) ATCC 29213, methicillin-resistant S. aureus (MRSA) ATCC 33591, and vancomycin-resistant S. aureus (VRSA) VRS5 inoculated into nutrient-depleted cation-adjusted Mueller-Hinton broth. As has been described for exponential-phase cells, oritavancin induced membrane depolarization, increased membrane permeability, and caused ultrastructural defects including a loss of nascent septal cross walls in stationary-phase MSSA. Furthermore, oritavancin sterilized biofilms of MSSA, MRSA, and VRSA at minimal biofilm eradication concentrations (MBECs) of between 0.5 and 8 mug/ml. Importantly, MBECs for oritavancin were within 1 doubling dilution of their respective planktonic broth MICs, highlighting the potency of oritavancin against biofilms. These results demonstrate a significant activity of oritavancin against S. aureus in phases of growth that exhibit tolerance to other antimicrobial agents.
Antimicrobial Agents and Chemotherapy 01/2009; 53(3):918-25. · 4.57 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: An important feature of microbial communities is the spatial heterogeneity of extracellular chemistry. Scanning transmission X-ray microscopy (STXM) was used to map the spatial distribution of iron speciesthroughout Pseudomonas aeruginosa biofilms to assess the influence of chemical heterogeneity on biomineralization. P. aeruginosa biofilms were treated with Fe(III)-amended media. Speciation and quantitative mapping using STXM image sequences in the Fe 2p(3/2) (L3) absorption edge region revealed both Fe(II) and Fe(III) in localized microenvironments. Fe(III) was mainly associated with cell surfaces, while small amounts of Fe(II) was found in the extracellular space. Biofilms were also characterized using C 1s edge STXM image sequences. Anaerobic growth assays and confocal microscopy revealed the inability of P. aeruginosa to directly reduce Fe(III), implicating indirect iron reduction mechanisms in the formation of fine-grained, multivalent minerals. These studies suggest that geochemical microenvironments found throughout microbial communities are even more complex than previously believed.
Environmental Science and Technology 01/2009; 42(23):8766-72. · 5.26 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The major structural component of bacterial cell walls is the peptidoglycan sacculus, which is one of nature's strongest and largest macromolecules that maintains the large internal pressure within the cell while allowing the transport of molecules into and out of the cell and cell growth. The three-dimensional structure of this unique biopolymer is controversial, and two models have been proposed: the planar model, in which the glycan strands lie in the plane of the cell surface, and the scaffold model, in which the glycan strands lie perpendicular to the cell surface. We have used atomic force microscopy to investigate the high resolution structure of isolated, intact sacculi of Escherichia coli K12 bacteria. Atomic force microscopy-single molecule force spectroscopy was performed on single sacculi exposed to the tAmiB enzyme which cleaves the peptide-glycan bonds. Surprisingly, the measurements revealed individual strands of up to 250 nm in length. This finding combined with high resolution AFM images recorded on hydrated sacculi provide evidence for the validity of the planar model for the peptidoglycan structure in Gram-negative bacteria.
[Show abstract][Hide abstract] ABSTRACT: The ultrastructural effects of the lipoglycopeptide oritavancin on methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant enterococcus (VRE) were examined by transmission electron microscopy. Oritavancin but not vancomycin induced aberrant septum formation and loss of staining of nascent septal cross walls in MRSA. Septal distortions were also observed in VRE exposed to oritavancin.
Antimicrobial Agents and Chemotherapy 12/2008; 53(2):800-4. · 4.57 Impact Factor