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PubMed1993

Antigenic properties of Campylobacter rectus (Wolinella recta) major S-layer proteins.

The antigenic properties of the surface layer (S-layer) proteins of various Campylobacter rectus strains including 24 clinical isolates and the type strain ATCC 33238 were examined. S-layer proteins were extracted from whole cells by acid treatment according to the method of McCoy et al. (Infect. Immun. 11, 517-525, 1975). The acid extracts from 23 of the isolates and ATCC 33238 contained two major proteins with molecular masses of 130 kDa and 150 kDa, both of which were identified as subunits of the S-layer after comparison with the protein profiles of acid-treated (S-layer-deficient) cells. An S-layer protein from one isolate (CI-808) demonstrated a different molecular mass (160 kDa). Both the 150-kDa proteins of ATCC 33238 and isolate CI-306 and the 160-kDa protein of CI-808 were purified by ion-exchange chromatography in the presence of urea. In Ouchterlony immunodiffusion experiments with these purified proteins and rabbit antiserum raised to each purified protein, both common and strain-specific antigenic determinants were identified in the C. rectus S-layer proteins.

Antigens, Bacterial↗
PubMed1997

Functions of S-layers.

Although S-layers are being increasingly identified on Bacteria and Archaea, it is enigmatic that in most cases S-layer function continues to elude us. In a few instances, S-layers have been shown to be virulence factors on pathogens (e.g. Campylobacter fetus ssp. fetus and Aeromonas salmonicida), protective against Bdellovibrio, a depository for surface-exposed enzymes (e.g. Bacillus stearothermophilus), shape-determining agents (e.g. Thermoproteus tenax) and nucleation factors for fine-grain mineral development (e.g. Synechococcus GL 24). Yet, for the vast majority of S-layered bacteria, the natural function of these crystalline arrays continues to be evasive. The following review up-dates the functional basis of S-layers and describes such diverse topics as the effect of S-layers on the Gram stain, bacteriophage adsorption in lactobacilli, phagocytosis by human polymorphonuclear leukocytes, the adhesion of a high-molecular-mass amylase, outer membrane porosity, and the secretion of extracellular enzymes of Thermoanaerobacterium. In addition, the functional aspect of calcium on the Caulobacter S-layer is explained.

Abortion, Veterinary↗
PubMed1989

Ultrastructural relationship of cell envelope layers in Wolinella recta.

The cell envelope of Wolinella recta was studied by electron microscopy. In thin sections the surface layer (S-layer) seemed to be in close contact to the outer membrane (OM). A hexagonally arranged pattern of the S-layer was revealed by freeze-etching. Unlike in other oral bacteria with an S-layer, the cleavage occurred along the S-layer by freeze-fracture, suggesting an exceptionally tight relationship between the two outermost layers. The convex surface of the S-layer was usually revealed by this technique. When cleavage occurred through the outer membrane, no periodicity was seen in the arrangement of the intramembranous particles.

Bacteroidaceae↗
PubMed2005

Fatigue resistance of dentin/composite interfaces with an additional intermediate elastic layer.

According to the "elastic bonding" concept, a thick intermediate layer of flexible resin has been suggested to absorb part of the polymerization shrinkage stress and to absorb shocks during function. In this study, the effect of an additional intermediate layer of a low-viscosity resin on the microrotary fatigue resistance (microRFR) of a hybrid composite bonded to dentin was evaluated. The hypotheses tested were that an intermediate layer of a low-viscosity resin (i) increases the microRFR to dentin, but (ii) has no effect on the static bond strength. Microtensile bond strength (microTBS) samples were loaded until failure or inserted in a microrotary fatigue testing device. Specimens were tested at 4 Hz until failure or until 10(5) cycles were reached. An additional intermediate elastic layer had no effect on the static microTBS, but significantly lowered the median microRFR from 28.4 MPa to 21.6 MPa. However, the application of an intermediate flexible layer had, no effect on the static microTBS. In conclusion, an additional elastic intermediate layer did decrease significantly the microRFR (rejection of hypothesis i), but did not alter the microTBS (acceptance of hypothesis ii). The decrease in microRFR most likely may be explained by the lower mechanical properties of the intermediary layer.

Composite Resins↗
PubMedDate not supplied

High-permeability layers for remediation of ground water; go wide, not deep.

A nitrate-reactive porous media layer comprising wood particles with very high hydraulic conductivity (K approximately 1 cm/s) was used to successfully treat nitrate in a shallow sand-and-gravel aquifer in southern Ontario. Nitrate concentrations of 1.3 to 14 mg/L as N in the aquifer were attenuated to <0.5 mg/L as N in the reactive layer. Borehole dilution testing indicated that ground water velocities in the reactive layer, although variable, averaged five times higher than in the surrounding aquifer, suggesting that the layer was capturing ground water flow from deeper in the aquifer. The use of high-K reactive media opens up the possibility of installing permeable reactive barriers as horizontal layers in the shallow water table zone that do not necessarily have to penetrate the full depth of a contaminant plume to be effective. Model simulations show that the depth of capture of a high-K layer increases as the layer width in the direction of flow increases. Shallower emplacement could decrease barrier costs at some sites.

Computer Simulation↗
PubMed1982

Spontaneous contractility of isolated muscle layers of the pig oviductal isthmus.

A new dissection technique for the mechanical separation of the smooth muscle layers of the pig oviductal isthmus is described. The spontaneous contractility of the isolated muscle layers was recorded using transducers designed to reflect the proper force development of the tubal muscle tissue specimens under in vitro conditions. A series of well defined stages of the entire estrual cycle was surveyed, and the isolated tubal muscle contractility was compared with that of the intact isthmus, according to previous work. The circular and longitudinal layers are both active during the estrual cycle, showing definite patterns in each stage explored. The circular layer revealed a changing pattern similar to, but of lower activity than, the intact isthmus, specially during the heat, postovulation and luteal phases. The periods are considered relevant because, during heat, the spermatozoa might be actively transported through the isthmus by the concerted contractions and relaxations of the circular muscles. The same would happen during the luteal phase, specially during the time the ova are descending into the uterus. As important would be the postovulation time, during which the sustained contraction of the circular layer could "lock" the ithmic lumen, thus preventing improper timing of ova descent. The longitudinal layer follows the intensity of the contraction patterns of the circular layers, being specially active during the periovulatory period.

Animals↗
PubMed1991

Fatty layer of the precorneal film in the 'office eye syndrome'.

The thickness of the fatty layer of the precorneal film was estimated by the semiquantitative inteference method in 70 office workers from two town halls in Copenhagen County and compared with that in 110 controls from the general population. There were significant correlations between use of eye make-up and a thin fatty layer (less than or equal to 50 nm, P = 0.023) and between investigation time before 12 h and a thick fatty layer (greater than or equal to 150 nm, P = 0.046). After correction for these confounders, the fatty layer was significantly reduced in the office population compared with the controls (P = 0.0032). Furthermore, absence of biomicroscopically dry eyes (expressed as a combination of premature break-up of the tear film and lissamine green-stained epithelial damage of the bulbar conjunctiva) was significantly correlated to a thick fatty layer (greater than or equal to 150 nm, P = 0.015), indicating that a thick fatty layer may protect the eyes against development of dry eyes. The results indicate that deficiency in available Meibomina oil is involved in the dry eye syndrome we earlier found to be associated with eye irritation in the office environment and termed the 'office eye syndrome'. Together with observation of foam in the eye canthus, estimation of break-up time, and examination for clusters of epithelial damage on the bulbar conjunctiva, measurement of the fatty layer appears to be an easy tool for diagnosing the 'office eye syndrome' or for differentiating this disorder from other conditions with complaints of dryness or irritation.

Adult↗
PubMed1978

Ocular dominance in layer IV of the cat's visual cortex and the effects of monocular deprivation.

1. The relation between the physiological pattern of ocular dominance and the anatomical distribution of geniculocortical afferents serving each eye was studied in layer IV of the primary visual cortex of normal and monocularly deprived cats. 2. One eye was injected with radioactive label. After allowing sufficient time for transeuronal transport, micro-electrode recordings were made, and the geniculocoritcal afferents serving the injected eye were located autoradiographically. 3. In layer IV of normal cats, cell were clustered according to eye preference, and fewer cells were binocularly driven than in other layers. Points of transition between groups of cells dominated by one eye and those dominated by the other were marked with electrolytic lesions. A good correspondence was found between the location of cells dominated by the injected eye and the patches of radioactively labelled geniculocortical afferents. 4. Following prolonged early monocular deprivation, the patches of geniculocortical afferents in layer IV serving the deprived eye were smaller, and those serving the non-deprived eye larger, than normal. Again there was a coincidence between the patches of radioactively labelled afferents and the location of cells dominated by the injected eye. 5. The deprived eye was found to dominate a substantial fraction (22%) of cortical cells in the fourth layer. In other cortical layers, only 7% of the cells were dominated by the deprived eye. 6. These findings suggest that the thalamocortical projection is physically rearranged as a consequence of monocular deprivation, as has been demonstrated for layer IVc of the monkey's visual cortex (Hubel, Wiesel & Le Vay, 1977).

Animals↗
PubMed1987

Differences in the amplitude of X-cell responses as a function of depth in layer A of lateral geniculate nucleus in cat.

1. Extracellular recordings were made from 131 X-like cells (hereafter called X cells) at different depths in layer A of the lateral geniculate nucleus (l.g.n.) in the cat. The depths of the cells were measured accurately by orienting the electrodes parallel to the borders of the layer and reconstructing the electrode tracks from histological sections. 2. Each cell was stimulated with a standardized stimulus; a 500 ms, low-contrast (10%) spot of light, matched in size to the centres of the receptive fields. The responses were later compared with the depths of the cells in the layer. 3. The amplitudes of the responses, which ranged between peak values of 20 and 400 spikes/s in different cells, were related to the depths of the cells in the layer. On average, the response amplitudes of the X-on cells near the borders of the layer were small compared to those in the middle. The responses of the X-off cells were reciprocal; large near the borders of the layer and small in the middle. 4. The results reveal a sublaminar organization in layer A based on the signs and amplitudes of X-cell responses. It is suggested that the pattern reflects differences in signalling within the X-on and X-off channels that are preserved in the relay to the visual cortex.

Action Potentials↗
PubMed1990

Layered structure of bacterial aggregates produced in an upflow anaerobic sludge bed and filter reactor.

The ultrastructure of bacterial granules that were maintained in an upflow anaerobic sludge bed and filter reactor was examined. The reactor was fed a sucrose medium, and it was operated at 35 degrees C. Scanning and transmission electron microscopy revealed that the granular aggregates were three-layered structures. The exterior layer of the granule contained a very heterogeneous population that included rods, cocci, and filaments of various sizes. The middle layer consisted of a slightly less heterogeneous population than the exterior layer. A more ordered arrangement, made up predominantly of bacterial rods, was evident in this second layer. The third layer formed the internal core of the granules. It consisted of large numbers of Methanothrix-like cells. Large cavities, indicative of vigorous gas production, were evident in the third layer. On the basis of these ultrastructural results, a model that presents a possible explanation of granule development is offered.

Bacteria↗
PubMed2004

Construction of a functional S-layer fusion protein comprising an immunoglobulin G-binding domain for development of specific adsorbents for extracorporeal blood purification.

The chimeric gene encoding a C-terminally-truncated form of the S-layer protein SbpA from Bacillus sphaericus CCM 2177 and two copies of the Fc-binding Z-domain was constructed, cloned, and heterologously expressed in Escherichia coli HMS174(DE3). The Z-domain is a synthetic analogue of the B-domain of protein A, capable of binding the Fc part of immunoglobulin G (IgG). The S-layer fusion protein rSbpA(31-1068)/ZZ retained the specific properties of the S-layer protein moiety to self-assemble in suspension and to recrystallize on supports precoated with secondary cell wall polymer (SCWP), which is the natural anchoring molecule for the S-layer protein in the bacterial cell wall. Due to the construction principle of the S-layer fusion protein, the ZZ-domains remained exposed on the outermost surface of the protein lattice. The binding capacity of the native or cross-linked monolayer for human IgG was determined by surface plasmon resonance measurements. For batch adsorption experiments, 3-microm-diameter, biocompatible cellulose-based, SCWP-coated microbeads were used for recrystallization of the S-layer fusion protein. In the case of the native monolayer, the binding capacity for human IgG was 5.1 ng/mm(2), whereas after cross-linking with dimethyl pimelimidate, 4.4 ng of IgG/mm(2) was bound. This corresponded to 78 and 65% of the theoretical saturation capacity of a planar surface for IgGs aligned in the upright position, respectively. Compared to commercial particles used as immunoadsorbents to remove autoantibodies from sera of patients suffering from an autoimmune disease, the IgG binding capacity of the S-layer fusion protein-coated microbeads was at least 20 times higher. For that reason, this novel type of microbeads should find application in the microsphere-based detoxification system.

Autoimmune Diseases↗
PubMed1998

Surface structure, hydrophobicity, phagocytosis, and adherence to matrix proteins of Bacillus cereus cells with and without the crystalline surface protein layer.

Nonopsonic phagocytosis of Bacillus cereus by human polymorphonuclear leukocytes (PMNs) with particular attention to bacterial surface properties and structure was studied. Two reference strains (ATCC 14579(T) and ATCC 4342) and two clinical isolates (OH599 and OH600) from periodontal and endodontic infections were assessed for adherence to matrix proteins, such as type I collagen, fibronectin, laminin, and fibrinogen. One-day-old cultures of strains OH599 and OH600 were readily ingested by PMNs in the absence of opsonins, while cells from 6-day-old cultures were resistant. Both young and old cultures of the reference strains of B. cereus were resistant to PMN ingestion. Preincubation of PMNs with the phagocytosis-resistant strains of B. cereus did not affect the phagocytosis of the sensitive strain. Negatively stained cells of OH599 and OH600 studied by electron microscopy had a crystalline protein layer on the cell surface. In thin-sectioned cells of older cultures (3 to 6 days old), the S-layer was observed to peel off from the cells. No S-layer was detected on the reference strains. Extraction of cells with detergent followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed a major 97-kDa protein from the strains OH599 and OH600 but only a weak 97-kDa band from the reference strain ATCC 4342. One-day-old cultures of the clinical strains (hydrophobicity, 5.9 to 6.0%) showed strong binding to type I collagen, laminin, and fibronectin. In contrast, reference strains (hydrophobicity, -1.0 to 4.2%) as well as 6-day-old cultures of clinical strains (hydrophobicity, 19.0 to 53.0%) bound in only low numbers to the proteins. Gold-labelled biotinylated fibronectin was localized on the S-layer on the cell surface as well as on fragments of S-layer peeling off the cells of a 6-day-old culture of B. cereus OH599. Lactose, fibronectin, laminin, and antibodies against the S-protein reduced binding to laminin but not to fibronectin. Heating the cells at 84 degreesC totally abolished binding to both proteins. Benzamidine, a noncompetitive serine protease inhibitor, strongly inhibited binding to fibronectin whereas binding to laminin was increased. Overall, the results indicate that changes in the surface structure, evidently involving the S-layer, during growth of the clinical strains of B. cereus cause a shift from susceptibility to PMN ingestion and strong binding to matrix and basement membrane proteins. Furthermore, it seems that binding to laminin is mediated by the S-protein while binding to fibronectin is dependent on active protease evidently attached to the S-layer.

Bacillus cereus↗
PubMed1973

Freeze-etching and x-ray diffraction of the isolated double-track layer from the cell wall of a gram-negative marine pseudomonad.

The isolated double-track layer of the cell wall of the gram-negative marine pseudomonad studied here contains a cleavage plane. This finding localizes the single cleavage plane of the cell wall and shows that the molecular architecture of this layer provides the lipid-enriched layer which cleaves preferentially in the frozen cell. The observation that the isolated double-track layer of the cell wall is sufficiently ordered at the molecular level to yield a well-defined X-ray diffraction pattern with a d-spacing of 0.44 nm shows that its molecular architecture is very similar to that of true membranes. This specific d-spacing is produced by the highly ordered packing of the hydrophobic portions of phospholipid molecules. Therefore, the double-track layer of the cell wall has been shown, by these two biophysical means, to have a molecular architecture which would allow it to function as the membrane-like "molecular sieve" layer, whose presence has been deduced from physiological data. This layer is important in the retention of cell wall-associated enzymes and in the control of the movement of large molecules through the cell wall.

Bacteria↗
PubMed1975

Electron microscopy studies of the limiting layers of the rickettsia Coxiella burneti.

Surface layers of Coxiella burneti studied at a high resoulution reveal a plasma membrane and an outer surface membrane 6 to 7 nm thick, and a thin, moderately electron-dense intermediate layer associated with the inner surface of the outer membrane of many cells. This layer appears to be unaffected by lysozyme treatment. Ruthenium red staining was used to delineate a layer of filamentous material external to the outer membrane; this fuzzy layer has a mean thickness of 20 nm and is not often seen on the surface of cells prepared by conventional means. Both antigenic phase I and II cells show a ruthenium red-binding surface layer. It is suggested that this fuzzy layer may be, among other possibilities, a highly branched mucopolysaccharide.

Animals↗
PubMed1976

Isolation and characterization of the rodlet layer of Trichophyton mentagrophytes microconidial wall.

The rodlet layer of the microconidial wall of Trichophyton mentagrophytes was isolated and partially characterized. The purified microconidial walls were first extracted with urea (8M), mercaptoethanol (1%), and sodium dodecyl sulfate (1%) followed by enzymatic digestion with glusulase (snail intestinal enzymes) and purified (1 leads to 3)-beta-D-glucanase and chitinase. The purified rodlet layer was 15 to 30 nm thick and accounted for approximately 10% of the original wall weight. The pattern of rodlet patches, as revealed by electron microscopy of freeze-etched preparations of the isolated layer, was essentially the same as that observed on the intact microconidial wall. The rodlet layer was found to be resistant to most of the common organic solvents, cell wall lytic enzymes, mild acid treatments, and surface-active agents, but was solubilized in boiling 1 N NaOH with concomitant disorientation of the rodlet patterns. A melanin or melanin-like pigment appeared to be intimately associated with this rodlet layer and was solubilized during a hot-alkali treatment. Protein (80 to 85%) and glucomannan (7 to 10%) were the major components of the rodlet layer. The rodlet layer did not contain any appreciable amounts of lipid or phosphorus.

Cell Fractionation↗
PubMed1979

Isolation, characterization, and in vitro assembly of the tetragonally arrayed layer of Bacillus sphaericus.

The tetragonally arranged cell wall layer (T-layer) of Bacillus sphaericus NTCC 9602 was isolated and characterized. Parallel studies were made on a spontaneous variant of the wild-type strain which had a T-layer subunit of altered molecular weight. A purification method for the T-layers was devised which involved separation of the cell walls from the cytoplasmic contents, urea dissociation of the T-layer from the cell walls, removal of soluble contaminants by differential centrifugation, and finally selective adsorption of uncleaved subunits to sacculi. The purified subunits retained the capacity to form an assembly in vitro with the same lattice parameters as that observed on whole cells or cell walls and could readsorb to the cell walls from which they had been extracted. Both the wild-type and the variant subunits behaved as single, homogeneous polypeptide chains. Carbohydrate assay and isoelectric point determinations revealed that both subunit types were acidic glycoproteins. Values obtained for thebuoyant density, isoelectric point, and extinction coefficient differed minimally; major differences were observed in the molecular weight and the characterisitc width of cylinders formed by in vitro-assembled T-layer of the wild-type and variant. Assembled T-layer was subject to alkaline or acid dissociation and in acid titration dissociated at its isoelectric point.

Bacillus↗
PubMed1985

Structures of two different surface layers found in six Bacteroides strains.

The structures of crystalline layers from six Bacteroides strains were studied by electron microscopy. Two different hexagonal crystalline surface layers were found, one with a unit cell spacing of 21.5 nm and another with a spacing of 7.7 nm. A three-dimensional structure of the 21.5-nm layer and a two-dimensional projection of the 7.7-nm layer were determined to 3.0- and 3.8-nm resolution, respectively, by computerized image processing of electron micrographs. Both of these two crystalline layers were found in all six strains studied: B. pentosaceus NP333T and WPH61, B. capillus ATCC 33690T and ATCC 33691, and B. buccae ATCC 33574T and ES57. This further supports the identity of B. pentosaceus, B. capillus, and B. buccae as suggested by M. Haapasalo, K. Lounatmaa, H. Ranta, H. Shah, and K. Ranta (Int. J. Syst. Bacteriol. 35:65-72, 1985). The surface layer with 21.5-nm spacing is an intricate network with two classes of pores through the layer.

Bacteroides↗
PubMed1986

Two patterns in the Aeromonas salmonicida A-layer may reflect a structural transformation that alters permeability.

Electron micrographs of negatively stained regular surface layers (A-layers) of Aeromonas salmonicida showed two square patterns having p4 symmetry. Computer image processing demonstrated that, at a resolution of 2.3 nm, both square arrays were composed of two different morphological units arranged alternatively to give a face-centered lattice in which the four nearest neighbors of each unit were the other type of unit. The lattice constant was slightly but significantly different in the two patterns, and the orientation of one of the two morphological units changed by about 20 degrees between patterns. These patterns were probably not derived from different strains present in the preparation, since both were seen in material that appeared to come from a single layer. This and the difference in lattice constant made it unlikely that they represented different sides of the A-layer. However, it is possible that the two patterns may reflect a structural transformation of the layer. In this respect, it is interesting that the rotation of one morphological subunit changed the size of the gaps between units in the layer. This raised the possibility that the transformation could be related to a change in permeability of the A-layer, possibly analogous to that proposed for gap junctions in eucaryotic cells.

Aeromonas↗