Trend reversal of nitrate in Danish groundwater--a reflection of agricultural practices and nitrogen surpluses since 1950

This paper assesses the long-term development in the oxic groundwater nitrate concentration and nitrogen (N) loss due to intensive farming in Denmark. First, up to 20-year time-series from the national groundwater monitoring network enable a statistically systematic analysis of distribution, trends, and trend reversals in the groundwater nitrate concentration. Second, knowledge about the N surplus in Danish agriculture since 1950 is used as an indicator of the potential loss of N. Third, groundwater recharge CFC (chlorofluorocarbon) age determination allows linking of the first two data sets. The development in the nitrate concentration of oxic groundwater clearly mirrors the development in the national agricultural N surplus, and a corresponding trend reversal is found in groundwater. Regulation and technical improvements in the intensive farming in Denmark have succeeded in decreasing the N surplus by 40% since the mid 1980s, while at the same time maintaining crop yields and increasing the animal production of especially pigs. Trend analyses prove that the youngest (0-15 years old) oxic groundwater shows more pronounced significant downward nitrate trends (44%) than the oldest (25-50 years old) oxic groundwater (9%). This amounts to clear evidence of the effect of reduced nitrate leaching on groundwater nitrate concentrations in Denmark.     

Development and application of new nucleic acid-based technologies for microbial community analyses in foods

Several challenges still persist in the analysis of microorganisms in foods, particularly in studies of complex communities. Nucleic acid-based methods are promising tools in addressing new questions concerning microbial communities. We have developed several new methods in the field of nucleic acid-based microbial community analyses. These methods cover both sample preparation and detection approaches. The sample preparation method involves simplified DNA purification using paramagnetic beads. As an extension of this method, the same paramagnetic beads are used for both cell separation and DNA purification. This enables full automation. The separate detection of viable and dead bacteria is a major issue in nucleic acid-based diagnostics. We have applied a living/dead dye that binds covalently to DNA and inhibits the PCR from dead cells. In addition, a DNA array-based detection assay has been developed. The assay combines the specificity obtained by enzymatic labeling of DNA probes with the possibility of detecting several targets simultaneously by DNA array hybridization. In combination with 16S rDNA amplification, this is a promising tool for community analyses. Also, we have developed a novel approach for multiplex quantitative PCR. The multiplex PCR has been combined with our DNA array-based detection method. Finally, we are now in the process of adapting a system for monitoring microbial growth and death in real-time through the tagging of bacteria with green fluorescent protein (GFP) combined with fluorescence detection using a high-resolution confocal laser scanner.     

An ester bond linking a fragment of a serine proteinase to its serpin inhibitor

Most known members of the serpin superfamily are serine proteinase inhibitors. Serpins are therefore important regulators of blood coagulation, complement activation, fibrinolysis, and turnover of extracellular matrix. Serpins form SDS-resistant complexes of 1:1 stoichiometry with their target proteinases by reaction of their P1-P1' peptide bond with the active site of the proteinases. The nature of the interactions responsible for the high stability of the complexes is a controversial issue. We subjected the complex between the serine proteinase urokinase-type plasminogen activator (uPA) and the serpin plasminogen activator inhibitor-1 (PAI-1) to proteolytic digestion under nondenaturing conditions. The complex could be degraded to a fragment containing two disulfide-linked peptides from uPA, one of which included the active site Ser, while PAI-1 was left undegraded. By further proteolytic digestion after denaturation and reduction, it was also possible to degrade the PAI-1 moiety, and we isolated a fragment containing 10 amino acids from uPA, encompassing the active site Ser, and 6 amino acids from PAI-1, including the P1 Arg. Characterization of the fragment gave results fully in agreement with the hypothesis that it contained an ester bond between the hydroxyl group of the active site Ser and the carboxyl group of the P1 Arg. These data for the first time provide direct evidence that serine proteinases are entrapped at an acyl intermediate stage in serine proteinase-serpin complexes.     

Autometallographic demonstration of zinc ions in rat sperm cells

An in-vitro technique for autometallographic (AMG) demonstration of chelatable zinc in electroejaculated sperm cells and spermatozoa from the epididymis is presented and the localization of zinc ions in rat spermatozoa is described. Sperm cells from caput epididymis showed zinc staining in all parts of the tail and a sparse, dispersed staining in the acrosome. Spermatozoa from cauda epididymis showed heavy staining in the acrosome but no staining in the tail, or post-acrosomal part of the sperm head. This distinct acrosomal AMG staining was also found in ejaculated spermatozoa, but additionally a segmentation of the tail was seen based on differences in staining intensity. The membrane penetrating chelator diethyldithiocarbamate (DEDTC) was found to block the AMG staining whereas calcium-EDTA, known not to pass through cell membranes, did not influence the staining, proving that the detected zinc ions are intracellularly located. Two different approaches for demonstrating the presence of a chelatable zinc pool at electron microscope levels are presented, and the ultrastructural presence of AMG grains located in the acrosome and in the mitochondria of the midpiece is demonstrated. It is postulated that an exchange of zinc ions takes place between the epididymal epithelium and the sperm cells as they pass along the epididymal duct.     

The neuropsychology of the prefrontal cortex

Single photon emission computed tomography with the xenon inhalation technique is used to compare activation of regional cerebral blood flow in frontal brain regions during the performance of four widely used neuropsychological tests: the Continuous Performance Test, the Wisconsin Card Sorting Test, the Tower of London, and Porteus Mazes. Healthy normal volunteers performing these tasks show significant increases in frontal regions during the Continuous Performance Test, the Wisconsin Card Sorting Test, and the Tower of London, but not the Porteus Mazes. Activation produced by the Continuous Performance Test and the Tower of London are mesial and bilateral and may reflect stimulation of midline attentional circuits. The Wisconsin Card Sorting Test produces a left dorsolateral area of prefrontal activation. These findings indicate that regional activation of the frontal lobes occurs in response to cognitive challenges produced through performance of standard neuropsychological tests.     

Lipid peroxidation, arachidonic acid and products of the lipoxygenase pathway in ischaemic preconditioning of rat heart

Progenitor and pluripotent stem cells reside within connective tissue compartments. They are also present in granulation tissue. This study examined the effects of treating these two cell populations with eight bioactive factors. Cells were assayed for DNA content as a measure of proliferation and for tissue-specific phenotypic markers as measures of lineage progression and lineage commitment. Platelet-derived endothelial growth factor and insulin-like growth factor-II did not induce proliferation in either population. However, dexamethasone, insulin, insulin-like growth factor-I, muscle morphogenetic protein, platelet-derived growth factor-AA, and platelet-derived growth factor-BB stimulated proliferation in one or both cell populations. Platelet-derived growth factor-BB was the most potent stimulator of proliferation in either population. Phenotypic expression markers were induced in the progenitor cells by insulin, insulin-like growth factor-I, insulin-like growth factor-II, dexamethasone, and muscle morphogenetic protein. However, only dexamethasone and muscle morphogenetic protein induced phenotypic expression markers in the pluripotent cells. Platelet-derived endothelial cell growth factor, platelet-derived growth factor-AA, and platelet-derived growth factor-BB did not induce phenotypic expression markers in progenitor or pluripotent cells. This study suggests the potential for using progenitor and pluripotent cells as an in vitro model to ascertain the effects of various bioactive factors on stem cells potentially involved in tissue maintenance and repair.     

Symptomless hypoglycaemia

We report an unusual case of hypoglycaemia, in which a pregnant woman with a Rh-negative problem and random blood glucose levels of less than 10 mg/100 ml was found to have no clinical symptoms of hypoglycaemia. The patient, who had a blood glucose concentration of 7 mg/100 ml, behaved as normally as a person with the expected average physiological value. This finding did not seem to be connected in any way with the patient's serious Rh-ve problem. We have been unable to find reports of a similar situation in the literature available to us.     

Partial purification and characterization of free and immobilized lipases fromMucor miehei

An extracellular lipase, a glycoprotein, produced by fermentation with a selected strain ofMucor miehei has been partially purified in two forms, A and B. The two forms have a high degree of antigenic identity and have similar pH-activity profiles with tributyroylglycerol as a substrate with optima at pH 7. They differ as follows: A, in contrast to B, requires activation at alkaline pH before analysis; A binds with concanavalin-A more completely than B; the net charges are slightly different at pH 8; and the isoelectric points are different. Our results indicate that the B lipase is formed by partial deglycosylation of the A lipase and that this influences the activity toward emulsions. In addition, the two enzymes have been immobilized by adsorption. These preparations and the soluble forms were highly specific for primary esters of triacylglycerols (TG); they usually hydrolyzed TG of 12∶0, 14∶0, 16∶0, and 18∶1 more rapidly than those of 4∶0, 6∶0, and 8∶0 and 10∶0 in mixtures of monoacid TG (4∶0 to 18∶1); and they were not stereospecific for TG. Immobilization altered the specificity of the preparations somewhat, in that slightly more 14∶0 and 16∶0 were released. Scientific Contribution No. 1192, Storrs Agricultural Experiment Station, University of Connecticut, Storrs, Connecticut.     

Scattering from bodies of revolution

The problem of scattering of a plane electromagnetic wave from an arbitrary metallic body of revolution is solved by a theoretical method for arbitrary incidence and polarization. The method permits numerical computations by high-speed digital computers, and examples are given. The incident wave is expanded in cylindrical modes, and an integral equation is solved for the induced current distribution of each mode. The scattering cross section, including the back-scattering or radar cross section, is found by summation of the mode scattered fields. The method is limited to a maximum perimeter length of twenty wavelengths. While the cases discussed in the paper pertain to perfectly conducting bodies, other surface boundary conditions, an arbitrary surface impedance or coatings by lossy dielectrics, can also be treated with equal precision.