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Two new paramagnetic defects ( and ) have been revealed in neutron-irradiated natural -quartz by using electron paramagnetic resonance (EPR) spectroscopy. The EPR spectra of the center as well as a previously reported but incompletely characterized center demonstrate that their super-hyperfine structures arise from interaction with 27 Al, the first-ever examples of Al-associated centers in crystalline quartz. The matrices g and , of , g and of the center, have been determined.

The crystal structures of two elbaites from the Paraiba deposit with copper contents of 3.51 and 1.61 wt% CuO [ a = 15.881(1), 15.840(3); c = 7.112(1), 7.1028(9), respectively], as well as a synthetic Cu-bearing olenite with a CuO content of 8.39 wt% [ a = 15.840(4), c = 7.091(1)] have been refined to R-indices of 2.2, 3.1, and 4.1% using X-ray single-crystal diffraction. On the basis of original and published data (for six structures) the crystal-chemical relationships of copper-bearing tourmalines has been analyzed. It is shown that copper cations and cations of other 3d elements (Mn, Zn, Fe) occupy almost exclusively the Y site. Such an ordered distribution results in a change of the size of the Y-octahedra. There is an inverse correlation between the content of 3d elements (mainly Cu2+ cations) and Al3+ cations in the Y-octahedron and also between the value of and the content of Al3+ cations in the Y site. The direct correlation between values of -distance in the structures of Cu-bearing elbaites and the parameters ( a , c ) of the unit cell have been found. Distortions of polyhedra in the structures of Cu-bearing elbaites are similar, and significantly higher than those present in the structure of the synthetic Cu-bearing olenite.

Marine Bacteroidetes have pronounced capabilities of degrading high molecular weight organic matter such as proteins and polysaccharides. Previously we reported on 76 Bacteroidetes-affiliated fosmids from the North Atlantic Ocean's boreal polar and oligotrophic subtropical provinces. Here, we report on the analysis of further 174 fosmids from the same libraries. The combined, re-assembled dataset (226 contigs; 8.8 Mbp) suggests that planktonic Bacteroidetes at the oligotrophic southern station use more peptides and bacterial and animal polysaccharides, whereas Bacteroidetes at the polar station (East-Greenland Current) use more algal and plant polysaccharides. The latter agrees with higher abundances of algae and terrigenous organic matter, including plant material, at the polar station. Results were corroborated by in-depth bioinformatic analysis of 14 polysaccharide utilisation loci from both stations, suggesting laminarin-specificity for four and specificity for sulfated xylans for two loci. In addition, one locus from the polar station supported use of non-sulfated xylans and mannans, possibly of plant origin. While peptides likely represent a prime source of carbon for Bacteroidetes in open oceans, our data suggest that as yet unstudied clades of these Bacteroidetes have a surprisingly broad capacity for polysaccharide degradation. In particular, laminarin-specific PULs seem widespread and thus must be regarded as globally important.

The stable paramagnetic center Cd+(2 S 1/2) in natural sedimentary calcium phosphates containing a cadmium impurity has been studied using electron paramagnetic resonance. The parameters of the spectrum have been determined, and the influence of different radiation types on the paramagnetic center has been investigated.

Synthetic stishovite before and after electron- and γ-ray irradiation has been investigated by single-crystal electron paramagnetic resonance (EPR) spectroscopy for the first time. Room-temperature single-crystal EPR spectra of as-is stishovite reveal two high-spin (S = 3/2) Cr 3+ centers: one with D 2 h symmetry and another of triclinic symmetry. Quantitatively determined spin Hamiltonian parameters, including matrices g , D , and A ( 53 Cr) and high-spin Zeeman term BS 3 , suggest that the D 2 h center represents a substitutional Cr 3+ ion at the Si site without an immediate charge compensator. The triclinic center, which is characterized by a well-resolved 1 H superhyperfine structure, also arises from a substitutional Cr 3+ ion at the Si site but has an H + charge compensator bonded to one of the four equatorial oxygen atoms. The magnitude and orientation of the 1 H superhyperfine structure yield the location of the H atom at (0.46, 0.12, 0). These structural models for Cr and H in stishovite also have been corroborated by periodic density functional theory (DFT) calculations using the Vienna ab initio simulation package (VASP), with 2 × 2 × 4 supercells, plane-wave basis sets and the projector augmented wave (PAW) potentials. In addition, 85 K EPR spectra of irradiated stishovite show that the two Cr 3+ centers are both converted to an S = 1/2 Cr 5+ center characterized by two 29 Si superhyperfine structures arising from interactions with two nearest and eight second-nearest Si atoms, respectively. The spin Hamiltonian parameters of this Cr 5+ center provide further support for the location of the two Cr 3+ centers at the Si site.

A natural quartz, annealed first at 800°C and then irradiated with a 3.5 MeV electron beam, has been investigated by single-crystal electron paramagnetic resonance (EPR) spectroscopy at X- and W-band frequencies from 110 K to 298 K. The W-band EPR spectra allow better separation of two previously reported radiation-induced defects (D and E) and improved determinations of their spin Hamiltonian parameters. These defects have similar g tensors with the gmax axes approximately along an O—O pair and gmin axes perpendicular to the short Si—O bonds, but different 27Al hyperfine structures. Centre E is also characterized by a 29Si hyperfine structure (A/geβe = ~0.4 mT). These spin Hamiltonian parameters, together with results from density functional theory (DFT) calculations, suggest centre E to be a new variant of peroxy radicals in quartz, whereas a peroxy radical model for centre D remains tentative. Thermal stabilities and decay kinetics of centres D and E have been investigated by use of isochronal and isothermal annealing experiments on a neutron-irradiated quartz and six smoky quartz crystals in druses from a U deposit.

Gammaproteobacterial Reinekea spp. were detected during North Sea spring algae blooms in the years 2009-2012, with relative abundances of up to 16% in the bacterioplankton. Here, we explore the ecophysiology of 'R. forsetii' strain Hel1_31_D35 that was isolated during the 2010 spring bloom using (i) its manually annotated, high-quality closed genome, (ii) re-analysis of in situ data from the 2009-2012 blooms and (iii) physiological tests. High resolution analysis of 16S rRNA gene sequences suggested that 'R. forsetii' dominated Reinekea populations during these blooms. This was corroborated by retrieval of almost complete Hel1_31_D35 genomes from 2009 and 2010 bacterioplankton metagenomes. Strain Hel1_31_D35 can use numerous low-molecular weight substrates including diverse sugar monomers, and few but relevant algal polysaccharides such as mannan, α-glucans, and likely bacterial peptidoglycan. It oxidizes thiosulfate to sulfate, and ferments under anoxic conditions. The strain can attach to algae and thrives at low phosphate concentrations as they occur during blooms. Its genome encodes RTX toxin and secretion proteins, and in cultivation experiments Hel1_31_D35 crude cell extracts inhibited growth of a North Sea Polaribacter strain. Our data suggest that the combination of these traits make strain Hel1_31_D35 a versatile opportunist that is particularly competitive during spring phytoplankton blooms.

The mechanism of induction of apoptosis by the novel anti-cancer drug 1-O-octadecyl-2-methyl-rac-glycero-3-phosphocholine (ET-18-OCH3) was investigated in p53-defective SV40 immortalized rat hepatocytes (CWSV1). Exposure to 12 microM ET-18-OCH3 for 36 h induced apoptosis as determined using classical morphological features and agarose gel electrophoresis of genomic DNA. Increased levels of reactive oxygen species (ROS) were detected spectrophotometrically using a nitroblue tetrazolium (NBT) assay in cells treated with ET-18-OCH3. Both the increased generation of ROS and the induction of apoptosis were inhibited when cells were treated concurrently with ET-18-OCH3 in the presence of the antioxidant alpha-tocopherol. Similar results were achieved when cells were switched acutely to choline-deficient (CD) medium in the presence of the antioxidant. The possible role of mitochondria in the generation of ROS was investigated. Both ET-18-OCH3 and CD decreased the phosphatidylcholine (PC) content of mitochondrial and associated membranes, which correlated with depolarization of the mitochondrial membrane as analyzed using 5,5',6,6'-tetramethylbenzimidazolcarbocyanine iodide (JC-1), a sensitive probe of mitochondrial membrane potential. Rotenone, an inhibitor of the mitochondrial electron transport chain, significantly reduced the intracellular level of ROS and prevented mitochondrial membrane depolarization, correlating with a reduction of apoptosis in response to either ET-18-OCH3 or CD. Taken together, these results suggest that the form of p53-independent apoptosis induced by ET-18-OCH3 is mediated by alterations in mitochondrial membrane PC, a loss of mitochondrial membrane potential, and the release of ROS, resulting in completion of apoptosis.

Continuous-wave electron paramagnetic resonance (EPR) spectroscopy has been the technique of choice for the studies of radiation-induced defects in silica (SiO2) for 60 years, and has recently been expanded to include more sophisticated techniques such as high-frequency EPR, pulse electron nuclear double resonance (ENDOR), and pulse electron spin echo envelope modulation (ESEEM) spectroscopy. Structural models of radiation-induced defects obtained from single-crystal EPR analyses of crystalline SiO2 (alfa-quartz) are often applicable to their respective analogues in amorphous silica (a-SiO2), although significant differences are common.

The globally abundant, uncultured unicellular cyanobacterium UCYN-A was recently discovered living in association with a eukaryotic cell closely related to a prymnesiophyte. Here, we established a double CAtalysed Reporter Deposition-Fluorescence In Situ Hybridization (CARD-FISH) approach to identify both partners and provided quantitative information on their distribution and abundance across distinct water masses along a transect in the North Atlantic Ocean. The N2 fixation activity coincided with the detection of UCYN-A cells and was only observed in oligotrophic ( 0.067 NO3(-) μM and 0.04 PO4(3-) μM) and warm ( 18°C) surface waters. Parallel 16S ribosomal RNA gene analyses among unicellular diazotrophs indicated that only UCYN-A cells were present. UCYN-A cells were associated with an algal partner or non-associated using the double CARD-FISH approach. We demonstrated that UCYN-A cells living in association with Haptophyta were the dominant form (87.0 ± 6.1%), whereas non-associated UCYN-A cells represented only a minor fraction (5.2 ± 3.9%). Interestingly, UCYN-A cells were also detected living in association with unknown single-celled eukaryotes in small amounts (7.8 ± 5.2%), presumably Alveolata. The proposed ecological niche of UCYN-A as an oligotrophic, mesophilic and obligate symbiotic nitrogen-fixing microorganism is evident for the North Atlantic Ocean.

E' centers as fundamental radiation-induced defects in amorphous and crystalline SiO2 have been thought to be related to oxygen vacancies for50years. However, direct proof for oxygen vacancies from experimental (17)O hyperfine data has never been provided. In this contribution, we report on the most complete set of spin Hamiltonian parameter matrices g, A((73)Ge), P((73)Ge), A((17)O), and A((29)Si) for the Ge analog of the classic E1(') center (denoted GeE1(')), determined from single-crystal electron paramagnetic resonance (EPR) analyses of two fast-electron-irradiated, Ge-doped α-quartz samples, including one grown from (17)O-enriched water. These experimental data, particularly the three distinct matrices A((17)O), not only confirm the GeE1(') center to represent spin trapping on a substitutional Ge atom coordinated to three nonequivalent nearest-neighbor O atoms (i.e., the GeO moiety involving an oxygen vacancy) and two next-nearest-neighbor Si atoms but also permit determination of the spin population on all of these atoms. These EPR data also have been evaluated by periodic density functional theory (DFT) calculations to demonstrate that the widely popular single-oxygen-vacancy V(O)(+) model for the GeE1(') center cannot account for A((17)O) arising from three nonequivalent nearest-neighbor oxygen atoms. A new tri-vacancy with an Al impurity model V(SiO2)(Al), on the other hand, reproduces all the EPR experimental hyperfine matrices A((73)Ge), A((17)O), and A((29)Si) and explains the common associations of the E' centers with both Al related defects and superoxide/peroxy radicals in quartz and amorphous silica.

Immortalized CWSV-1 rat hepatocytes, in which p53 protein is inactivated by SV40 large T antigen, had increased numbers of cells with strand breaks in genomic DNA (terminal dUTP end labeling) when grown in 0 Micron choline (67-73% of cells) than when grown in 70 Micron choline (2-3% of cells). Internucleosomal fragmentation of DNA (DNA ladders) was detected in cells grown with 5 Micron and 0 Micron choline for 72h. Cells treated with 0 or 5 Micron choline for 72h detached from the substrate in high numbers (58% of choline deficient cells vs. 1.4% of choline sufficient cells detached) exhibited a high incidence of apoptosis (apoptotic bodies were seen in 55-75% of cells; 67-73% had DNA strand breaks), and an absence of mitosis and proliferating cell nuclear antigen (PCNA) expression. Cells undergoing DNA fragmentation had functioning mitochondria. At 24h, cells grown in 0 or 5 Micron choline synthesize DNA more rapidly than those grown in 70 Micron choline. By 72h, the cells grown in 0 or 5 Micron choline ...

The use of ribosomal RNA (rRNA) probe technology for the characterization of complex microbial communities is reviewed and illustrated by discussing the results of a long-term study of four anaerobic fixed-bed biofilm reactors. Two distinct approaches were used to characterize the microbial community structure in these biofilm reactors. The first used a collection of phylogenetically defined oligonucleotide rRNA probes for methanogens and sulfate-reducing bacteria (SRB) to quantify their populations. Population abundance was linked to the functional behavior of the biofilm reactor community by determining the effluent concentrations of the substrates, intermediates, and final products of microbial metabolism. This analysis indicated that the presence of SRB (especially Desulfovibrio-species) was not dependent upon the presence of sulfate. Methanobacteriales-species were the major competitors for hydrogen with these SRB in the absence of sulfate. The second approach involved selective amplification, cloning, sequencing, and whole cell hybridization to identify, visualize, and isolate a biofilm community member (strain PT-2). Subsequently, it was determined that the growth rate of strain PT-2 was significantly higher in young biofilms than in established biofilms.

Nitrogen (N₂) fixation is a globally important process often mediated by diazotrophic cyanobacteria in the open ocean. In 2010, seawater was collected near Cape Verde to identify and measure N₂ and carbon (C) fixation by unicellular diazotrophic cyanobacteria. The nifH gene abundance (10⁴-10⁶ nifH L⁻¹) and nifH gene transcript abundance (10²-10⁴ cDNA nifHL⁻¹) for two unicellular groups, UCYN-A and UCYN-B, were detected. UCYN-A was also identified and quantified (10⁴-10⁵cells L⁻¹) by new probes (UCYN-A732 and UCYN-A159) using Catalyzed Reporter Deposition-Fluorescence In Situ Hybridization (CARD-FISH) assays. The UCYN-A were observed as free cells or attached to a larger unidentified eukaryotic cell. A Halogen In Situ Hybridization-Secondary Ion Mass Spectrometry (HISH-SIMS) assay using the UCYN-A732 probe was applied on samples previously incubated with ¹³C-bicarbonate and ¹⁵N₂. Free UCYN-A cells were enriched in both ¹³C and ¹⁵N and estimated C and N₂ fixation rates for UCYN-A were lower compared to co-occurring unicellular cyanobacteria cells similar in size (3.1-5.6 μm) and pigmentation to diazotroph Crocosphaera watsonii. Here, we identify and quantify two common co-occurring unicellular groups and measure their cellular activities by nanoSIMS.

Summary Six oligonucleotide probes complementary to the 16S rRNAs of members of the genus Fibrobacter were designed for identification at the levels of genus, species and subspecies. The temperature of dissociation (T d ) of probe-target hybrid was determined for each probe and used to control the stringency of hybridization. Specificity was demonstrated by comparing hybridization to rRNA from target and nontarget species immobilized on nylon membranes. Hybridization to nucleic acid isolated from cattle and goat gut contents suggested greater genetic diversity among this assemblage of fiber-digesting bacteria than now represented in pure culture.

Structural alterations in mitochondrial DNAs (mtDNAs) from a plant of a sterile sugar beet line, callus derived from it, suspension-cultured cells and plants regenerated from the callus were studied. BamHI restriction analysis revealed that structural alterations between the mtDNAs of the callus and the control plant had occurred. Multiple rearrangements were also demonstrated in the mtDNA from the suspension culture, of which some were similar to those appearing in the callus, and others had arisen de novo. Rearrangements were also identified by means of blot hybridization of BamHI-digested mtDNA from suspension-cultured cells with the genes encoding subunit II of cytochrome oxidase (cox II) and subunit 1 of NADH-dehydrogenase (Nd1). No alterations were observed in the mitochondrial genome of the callus and regenerants. The location of the genes for the α-subunit of F1-ATPase (atpA) and apocytochrome b (cob) in the mtDNA remained unchanged. Our salient finding was of a plant with an altered mitochondrial genome as judged by EcoRI and BamHI restriction analysis. This exceptional plant had retained the sterile phenotype like all of the other regenerants and the parent. The set of plasmid-like molecules of mtDNA remained the same as that in the control plant and in all of the regenerants, callus and suspension-cultured cells. The only type of plasmid-like molecule found in all of the DNAs was the 1.6-kbp minicircle, which is a feature of sterile cytoplasms. These structural changes in mtDNA were obviously a consequence of somaclonal variation during the in vitro cultivation of the sugar beet cells.

The probability that damage occurs in closely opposed sites on complementary DNA strands increases when DNA is heavily modified with mutagenic agents. Enzymatic excision of the opposite lesions produces DNA double-strand breaks which give rise to genomic rearrangements (deletions, insertions, etc.). Plasmid systems were developed for studying chemical lesions leading to double-strand breaks and the fate of broken plasmid molecules within bacterial cells. Deletions result from the base-pairing of fortuitously located direct repeats flanking the DNA broken ends; as a consequence, the latter are joined, while the DNA fragment between the direct repeats is deleted. Genomic rearrangements arise during the repair of the DNA double-strand breaks, and both events are due to similar repair enzymes which maintain the integrity of the DNA primary structure when conditions are not stressful. A number of genomic rearrangements and point mutations seem to be predetermined by the DNA primary structure.

Mitochondrial (mt) DNA from eight cytoplasmic male-sterile (cms) lines of sugar beet from different breeding stations was investigated by restriction fragment analysis and Southern hybridization. All cms lines showed similar but not identical restriction and hybridization signal patterns, readily distinguishable from those of fertile (N) cytoplasm. Digestion of the mtDNA with BamHI, EcoRI, SalI, and XhoI revealed distinct differences between the sterile lines, and six subtypes of the S cytoplasm could be distinguished. Differences between the sterile lines were confirmed by hybridization with a \(\overline{\overline {COX}} II\) gene probe revealing minor, line-specific hybridization signals. The data presented provide evidence for the existence of considerable variation within the only commercially used source of cms in the sugar beet, the Owen's type of cytoplasm.