High resolution microchemical analysis using soft X-ray lithographic techniques.

High resolution x-ray lithographic studies of cells from chick embryo hearts dried by the CO2 critical point method have been made with soft x-ray radiation of different wavelengths. A marked difference in the relief replica in polymethyl methacrylate (PMMA) resulting from the differential absorption by the dried cells of carbon K alpha radiation at 4.48 nm and broad band synchrotron radiation (SR) with lambda is greater than 1.5 nm demonstrates the potential usefulness of the technique in making high resolution (approximately or equal to 10 nm) chemical identification of the constitutents which make up the various parts of the cell.

Bacterial recognition of mineral surfaces: nanoscale interactions between Shewanella and alpha-FeOOH.

Force microscopy has been used to quantitatively measure the infinitesimal forces that characterize interactions between Shewanella oneidensis (a dissimilatory metal-reducing bacterium) and goethite (alpha-FeOOH), both commonly found in Earth near-surface environments. Force measurements with subnanonewton resolution were made in real time with living cells under aerobic and anaerobic solutions as a function of the distance, in nanometers, between a cell and the mineral surface. Energy values [in attojoules (10(-18) joules)] derived from these measurements show that the affinity between S. oneidensis and goethite rapidly increases by two to five times under anaerobic conditions in which electron transfer from bacterium to mineral is expected. Specific signatures in the force curves suggest that a 150-kilodalton putative iron reductase is mobilized within the outer membrane of S. oneidensis and specifically interacts with the goethite surface to facilitate the electron transfer process.

Distribution of norepinephrine transporters in the non-human primate brain.

Noradrenergic terminals in the central nervous system are widespread; as such this system plays a role in varying functions such as stress responses, sympathetic regulation, attention, and memory processing, and its dysregulation has been linked to several pathologies. In particular, the norepinephrine transporter is a target in the brain of many therapeutic and abused drugs. We used the selective ligand [(3)H]nisoxetine, therefore, to describe autoradiographically the normal regional distribution of the norepinephrine transporter in the non-human primate central nervous system, thereby providing a baseline to which alterations due to pathological conditions can be compared. The norepinephrine transporter in the monkey brain was distributed heterogeneously, with highest levels occurring in the locus coeruleus complex and raphe nuclei, and moderate binding density in the hypothalamus, midline thalamic nuclei, bed nucleus of the stria terminalis, central nucleus of the amygdala, and brainstem nuclei such as the dorsal motor nucleus of the vagus and nucleus of the solitary tract. Low levels of binding to the norepinephrine transporter were measured in basolateral amygdala and cortical, hippocampal, and striatal regions. The distribution of the norepinephrine transporter in the non-human primate brain was comparable overall to that described in other species, however disparities exist between the rodent and the monkey in brain regions that play a role in such critical processes as memory and learning. The differences in such areas point to the possibility of important functional differences in noradrenergic information processing across species, and suggest the use of caution in applying findings made in the rodent to the human condition.

Interactions between biofilms and the environment.

The surfaces of bacteria are highly interactive with their environment. Whether the bacterium is Gram-negative or Gram-positive, most surfaces are charged at neutral pH because of the ionization of the reactive chemical groups which stud them. Since prokaryotes have a high surface area-to-volume ratio, this can have surprising ramifications. For example, many bacteria can concentrate dilute environmental metals on their surfaces and initiate the development of fine-grained minerals. In natural environments, it is not unusual to find such bacteria closely associated with the minerals which they have helped develop. Bacteria can be free-living (planktonic), but in most natural ecosystems they prefer to grow on interfaces as biofilms; supposedly to take advantage of the nutrient concentrative effect of the interface, although there must also be gained some protective value against predators and toxic agents. Using a Pseudomonas aeruginosa model system, we have determined that lipopolysaccharide is important in the initial attachment of this Gram-negative bacterium to interfaces and that this surface moiety subtly changes during biofilm formation. Using this same model system, we have also discovered that there is a natural tendency for Gram-negative bacteria to concentrate and package periplasmic components into membrane vesicles which bleb-off the surface. Since some of these components (e.g., peptidoglycan hydrolases) can degrade other surrounding cells, the vesicles could be predatory; i.e., a natural system by which neighboring bacteria are targeted and lysed, thereby liberating additional nutrients to the microbial community. This obviously would be of benefit to vesicle-producing bacteria living in biofilms containing mixed microbial populations.

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.

Bacterial S-layers.

S-layers are produced by the self assembly of proteinaceous subunits on the surfaces of prokaryotes, so that planar, monomolecular-thick crystalline lattices are formed. Some archaeal and eubacterial S-layer proteins are glycosylated. These lattices typically have center-to-center spacings of less than 25 nm, which makes them attractive for biomimetic or nanotechnological applications.

Crystalline order to high resolution in the sheath of Methanospirillum hungatei: a cross-beta structure.

Electron microscopy and electron diffraction indicate that the outer sheath of the cell wall of the archaebacterium Methanospirillum hungatei contains a two-dimensional crystalline lattice having, at least to low resolution, p2 symmetry in projection with a = 5.66 nm, b = 2.81 nm and gamma = 85.6 degrees. At a resolution of 2 nm, the unit cell contains two lobes, whereas high-angle electron diffraction shows the presence of a substantial quantity of beta structure, with the 0.47 nm spacing (between polypeptide chains within a sheet) oriented circumferentially. The sheath is unusual when compared to other regular surface arrays found on bacteria in that it is a compact structure with small subunits. It may have a structural role analogous to barrel hoops since it tends to fragment perpendicular to its axis to give rings or hoops.

Evidence of dose saving in routine CT practice using iterative reconstruction derived from a national diagnostic reference level survey.

OBJECTIVE: To assess the influence and significance of the use of iterative reconstruction (IR) algorithms on patient dose in CT in Australia. METHODS: We examined survey data submitted to the Australian Radiation Protection and Nuclear Safety Agency (ARPANSA) National Diagnostic Reference Level Service (NDRLS) during 2013 and 2014. We compared median survey dose metrics with categorization by scan region and use of IR. RESULTS: The use of IR results in a reduction in volume CT dose index of between 17% and 44% and a reduction in dose-length product of between 14% and 34% depending on the specific scan region. The reduction was highly significant (p < 0.001, Wilcoxon rank-sum test) for all six scan regions included in the NDRLS. Overall, 69% (806/1167) of surveys included in the analysis used IR. CONCLUSION: The use of IR in CT is achieving dose savings of 20-30% in routine practice in Australia. IR appears to be widely used by participants in the ARPANSA NDRLS with approximately 70% of surveys submitted employing this technique. ADVANCES IN KNOWLEDGE: This study examines the impact of the use of IR on patient dose in CT on a national scale.

The brain gene expression profile of dopamine D2/D3 receptors and associated signaling proteins following amphetamine self-administration.

Persistent neuroadaptations following chronic psychostimulant exposure include reduced striatal dopamine D2 receptor (D2R) levels. The signaling of D2Rs is initiated by Gαi/o proteins and terminated by regulator of G protein signaling (RGS) proteins. The purpose of this study is to examine the association of the drug taking behavior and gene expression profile of D2/D3Rs, and their associated signaling proteins in the ventral tegmental area (VTA) and nucleus accumbens (NAc) using a rodent model of amphetamine (AMPH) self-administration. Rats were allowed to self-administer AMPH (0.187 mg/kg/infusion for a maximum of 40 injections in 6h daily sessions) for 5 days during which rats showed an escalated rate of AMPH intake across days. AMPH self-administration induced profound brain region-dependent alterations of the targeted genes. There was a positive correlation of the messenger ribonucleic acid (mRNA) levels of RGS10 between the VTA and the NAc in the control animals, which was abolished by AMPH self-administration. AMPH self-administration also produced a negative correlation of the mRNA levels of RGS7 and RGS19 between the two brain regions, which was not present in the control group. Furthermore, AMPH taking behavior was associated with changes in certain gene expression levels. The mRNA levels of RGS2 and RGS4 in both the VTA and NAc were positively correlated with the rate of AMPH intake. Additionally, the rate of AMPH intake was also positively correlated with RGS10 and negatively correlated with RGS17 and the short form of D2Rs mRNA level in the VTA. Although there were significant changes in the mRNA levels of RGS7 and RGS8 in the NAc, none of these measures were correlated with the rate of AMPH intake. The present study suggested that short-term AMPH self-administration produced pronounced changes in the VTA that were more associated with AMPH taking behavior than changes in the NAc.

Genomic, transcriptional and phenotypic analysis of ftsE and ftsX of Neisseria gonorrhoeae.

Although ftsE and ftsX are not universally present in bacteria, they are present in various Neisseria species as determined by Southern hybridization. The ftsE and ftsX genes of Neisseria gonorrhoeae (Ng) CH811 were cloned, sequenced and were shown to be co-transcribed from two promoters (P(E)1 and P(E)2) which were identified upstream of ftsE(Ng) by primer extension. Sequence analysis of FtsE(Ng) and alignment with other FtsE indicated that it contained the conserved motifs of ABC domains while sequence alignment of FtsX(Ng) with other published FtsX sequences predicted that they all contain four transmembrane segments and a conserved motif (Leu-hydrophobic aa-Gly-Ala/Gly) which may prove to be important for FtsX function. The viability of ftsE(Ng) and ftsX(Ng) mutants that were constructed by insertional inactivation indicated that these genes are not essential. The role of FtsE and FtsX is controversial. Analysis of ftsE(Ng) and ftsX(Ng) mutants by transmission electron microscopy showed that both exhibited morphological abnormalities indicative of defective division sites and in some cases aberrant condensation of DNA.

The efficacy and tolerability of cyclosporine G in human kidney transplant recipients.

Twelve consecutive first cadaveric kidney transplant recipients received cyclosporine G (CsG)(initial dose 12 mg/kg per day) as basic immunosuppressive treatment along with prednisone (initial dose 0.5 mg/kg per day) for the first three months after transplantation. Thereafter CsG was replaced by Sandimmun (cyclosporine, CsA). Evaluation of the immunosuppressive efficacy and assessment of possible side effects of CsG was made and compared with the results in 38 historical control patients starting with the same dose of CsA as part of the same immunosuppressive dosage schedule. Statistically, there was no difference in patient survival at three (91% in CsG group versus 95% in CsA group) and twelve months (91% in CsG group versus 92% in CsA group), or in graft survival at three (75% in CsG group versus 89% in CsA group) and twelve months (75% in CsG group versus 84% in the CsA group). At equivalent oral doses of CsG and CsA significantly higher blood levels of CsG were observed (2P less than 0.05). Nephrotoxicity assessed by graft biopsy could be demonstrated to a similar extent in both groups, whereas hepatotoxicity was more pronounced during CsG treatment. Sequential measurements of bilirubin revealed a significant increase in all patients but median values were significantly higher in the CsG patients. A pronounced and concordant elevation of liver enzymes occurred during CsG treatment in three out of 12 patients. Liver biopsies performed in these patients revealed histological alterations consistent with toxic liver injury. Thus, in human kidney transplant recipients CsG and CsA appeared to be equally immunosuppressive and nephrotoxic but more hepatotoxic. On the basis of this limited experience we conclude that in human kidney transplant recipients CsG has no advantage over CsA.

Pharmacokinetics of cyclosporine G in patients with renal failure.

The pharmacokinetics of the cyclosporine A (CsA, Sandimmune) analogue Nva2-cyclosporine, or cyclosporine G (CsG) was investigated in 6 patients with terminal renal failure after a 4-hr intravenous infusion (3.5 mg/kg) and after oral administration (600 mg) of the drug. Blood samples were collected up to 38 hr and CsG concentrations were measured by radioimmunoassay and high-performance liquid chromatography. The resulting pharmacokinetic parameters of CsG were similar to those described for CsA in the same patient population. Based on HPLC determinations, a mean terminal elimination half-life of 18.9 hr was calculated. The total body clearance was 0.55 L/hr/kg, the volume of the central compartment was 0.32 L/kg, and the steady-state volume of distribution was 5.97 L/kg. After oral administration maximum CsG concentrations in blood were reached between 2.5 and 3 hr, and the bioavailability was in the range of 24-55% (mean 36%). The ratios between the polyvalent RIA and HPLC determinations were considerably larger after oral dosing than after i.v. infusion. The blood-to-plasma ratio was 1.23, which is smaller than that observed for CsA. These results suggest that in patients undergoing renal transplantation the same dosing strategies can be applied for CsG as have been established for CsA.

Periplasm, periplasmic spaces, and their relation to bacterial wall structure: novel secretion of selected periplasmic proteins from Pseudomonas aeruginosa.

A brief overview of thin sections of cryopreserved walls from select eubacteria will be presented to suggest that all bacteria have functional periplasms, but that these are not necessarily confined to a periplasmic space such as found in typical gram-negative bacteria. Pseudomonas aeruginosa contains many components in its periplasmic space, some of which are required for infection. Throughout its growth cycle, P. aeruginosa blebs-off membrane vesicles that can possess DNA, endotoxin, phospholipase, protease, hemolysin, alkaline phosphatase, and autolysin, each of which must have a molecular phase that resides in the periplasm. These membrane packets make good delivery systems to convey these components to other bacteria and, possibly, tissue. Aminoglycoside antibiotics, such as gentamicin, produce a serious perturbation on the bacterium's surface (separate from the ribosomal effect), which contributes to the killing of the microorganism. Antibiotics such as this increase the size and number of the membrane blebs, which could contribute to septic shock of patients under drug therapy.

Structure and properties of novel inclusions in Shewanella putrefaciens.

Cytoplasmic inclusions surrounded by a bilayer membrane were seen in thin sections. negatively stained and freeze-fractured preparations of Shewanella putrefaciens. Cells harvested from the late exponential and early stationary phase showed a higher number of these vesicles than bacteria isolated from early exponential or late stationary phase. Chemical dyes for polyphosphate or poly-beta-hydroxybutyrate did not stain the material enclosed within these vesicles. Elemental analysis of the material indicated that the content was organic in nature and might be a protein. HPLC analysis of the material showed that it was probably not a carbon source, nor an electron acceptor used by S. putrefaciens.

Observations of the ultrastructure of infected kidney stones.

Struvite stones are formed as the result of urinary tract infection by urease-producing bacteria. Ultrastructural examination of calculi removed from a patient revealed bacteria incorporated throughout the stone matrix. Exopolysaccharide stained by ruthenium red was associated with most of the bacteria, but it represented only a small portion of the organic matrix in the stone. Localised deposits of calcium and phosphorus, components of carbonate-apatite, and magnesium, a struvite component, were detected in close proximity to the cells. Histochemical examinations revealed that several of the gram-negative bacteria within the stone matrix possessed high levels of urease activity. We propose that bacterial slime production, intimately involved in the initiation of stone matrix deposition, is less prominent in mature stones because of the increased incorporation of host-derived mucoproteins and mucopolysaccharides.

Whiting events: biogenic origin due to the photosynthetic activity of cyanobacterial picoplankton.

An annual whiting event occurs each year in late May to early June in Fayetteville Green Lake, New York. The initiation of this event correlates with exponential growth of the Synechococcus population within the lake. Synechococcus is the dominant (by approximately 4 orders of magnitude) autotrophic organism owing to the oligotrophic condition of the lake. The delta 13C values of the dissolved inorganic C range seasonally from -9.5% in winter to -6.2% in summer due to photosynthetic activity. Calcite precipitates principally in the microenvironment surrounding Synechococcus because of a photosynthetically driven alkalization process and the availability of the cells as nucleation sites. This calcite has a heavier delta 13C value (>4%) than does the dissolved inorganic C of the lake water owing to the cells' preferential uptake of 12C. A conceptual model suggests that photosynthetic activity and cell surface chemistry, together with the substantial surface area that arises from the great abundance of micron-sized cells, allow Synechococcus to dominate the annual whiting events in Fayetteville Green Lake.

Scanning probe microscopy in microbiology.

Scanning probe microscopy (SPM) is emerging as an important alternative to electron microscopy as a technique for analyzing submicron details on biological surfaces. Microbiological specimens such as viruses, bacteriophages, and ordered bacterial surface layers and membranes have played an important role in the development of scanning tunnelling microscopy (STM) and atomic force microscopy (AFM) in cellular and molecular biology. Early STM studies involving metal-coated bacteriophage T4 polyheads, Methanospirillum hungatei, and Deinococcus radiodurans HPI layer clearly demonstrated that resolution was comparable to TEM on similarly prepared specimens and only limited by metal graininess. However, except for thin films or layers, novel biological information has been difficult to obtain since imaging native surfaces of such biomaterials as proteins or nucleic acids by STM proved to be unreliable. With the development of atomic force microscopes, which allow imaging of similar native structures, SPM applications have widened to include straightforward surface structure analysis, analysis of surface elastic and inelastic properties, bonding force measurements between molecules, and micro-manipulations of such individual molecules as DNA. AFM images have progressed from relatively crude representations of specimen topography to nanometer scale representations of native hydrated surfaces. It appears from the study of microbiological specimens that direct visualization of dynamic molecular events or processes may soon become a reality.

Processing effects on the composition of sea buckthorn juice from Hippophae rhamnoides L. Cv. Indian Summer.

Sea buckthorn juice is one product that can be derived from the sea buckthorn berry, a new alternative crop for the Canadian western provinces. Fresh pressed juice separates into three phases when allowed to stand overnight in the refrigerator: an upper cream phase, juice in the middle portion, and a sediment at the bottom. Enzymatic hydrolysis with commercial, broad spectrum carbohydrate hydrolyzing enzyme preparations reduced the juice viscosity, assisted juice separation, and provided an opalescent juice. Soluble solids averaged 10.2 degrees Brix, pH averaged 3.13, ascorbic acid averaged 174.2 mg/100 mL, and titratable acidity averaged 1.97% as malic acid all determined on centrifuged (10 000 rpm, 15 min) juice. Soluble sugars included glucose, fructose, and an unidentified component that was not sucrose or other common soluble monomeric or dimeric sugar. Quinic acid was quantitatively most important, while malic was next, and oxalic, citric, and tartaric acids were minor components. Washing berries by dipping reduced soluble solids (degrees Brix) in juice suggesting uptake of wash water.

Sea buckthorn products: manufacture and composition.

Sea buckthorn (Hippophae rhamnoides) is a unique plant currently being domesticated. The fruit is the main component of value, although the leaves are occasionally made into sea buckthorn tea. The two main sources of valuable products are derived from the berries, juice from the fleshy tissue and seed as a single seed from each berry. The juice provides a nutritious beverage, high in suspended solids and very high in vitamin C and carotenes. The juice may contain an oil phase trapped within the suspended solids, or the oil may be removed as pulp oil and provided separately. The pulp remaining after juice removal provides for extraction of "sea buckthorn yellow", a pigment that has potential use as a food coloring material. The seed is a source of seed oil, which is very unsaturated and shows promise, because of its light absorption and emollient properties, as an ingredient in cosmetics, phytopharmaceuticals, or UV skin protectant preparations. It may be prepared by conventional extraction techniques or by supercritical carbon dioxide extraction. The manufacture of the main products derived from sea buckthorn is described, including several examples from the patent literature. The available compositional data for the main products are tabulated to form a comprehensive source of information on the manufacture and composition of sea buckthorn products.

Rheology of sea buckthorn (Hippophae rhamnoides L.) juice.

Juice extracted from sea buckthorn fruits was subjected to dynamic rheological measurements in a controlled stress rheometer. Sea buckthorn juice exhibited wide variations in flow behavior from pseudoplastic to dilitant with increasing temperature. The power law model suitably (r >/= 0.975, P