Ability of different restorative materials to prevent in situ secondary caries: analysis by polarized light-microscopy and energy-dispersive X-ray

Secondary caries is the main cause of direct restoration replacement. The purpose of this study was to analyze enamel adjacent to different restorative materials after in situ cariogenic challenge using polarized-light microscopy (PLM), scanning electron microscopy (SEM) and energy-dispersive X-ray analysis (EDS). Twelve volunteers, with a low level of dental plaque, a low level of mutans streptococci, and normal salivary flow, wore removable palatal acrylic appliances containing enamel specimens restored with Z250 composite, Freedom composite, Fuji IX glass-ionomer cement, or Vitremer resin-modified glass-ionomer for 14 days. Volunteers dripped one drop of 20% sucrose solution (n = 10) or distilled water (control group) onto each specimen 8 times per day. Specimens were removed from the appliances and submitted to PLM for examination of the lesion area (in mm(2)), followed by dehydration, gold-sputtering, and submission to SEM and EDS. The calcium (Ca) and phosphorus (P) contents were evaluated in weight per cent (%wt). Differences were found between Z250 and Vitremer, and between Z250 and FujiIX, when analyzed using PLM. Energy-dispersive X-ray analysis results showed differences between the studied materials regarding Ca %wt. In conclusion, enamel adjacent to glass-ionomer cement presented a higher Ca %wt, but this material did not completely prevent enamel secondary caries under in situ cariogenic challenge.

Measurement of the t(t)over-bar production cross section in p(p)over-bar collisions at root s=1.96 TeV using secondary vertex b tagging

We report a new measurement of the t (t) over bar production cross section in p (beta) over bar collisions at a center-of-mass energy of 1.96 TeV using events with one charged lepton (electron or muon), missing transverse energy, and jets. Using 425 pb(-1) of data collected using the D0 detector at the Fermilab Tevatron Collider, and enhancing the t (t) over bar content of the sample by tagging b jets with a secondary vertex tagging algorithm, the t (t) over bar production cross section is measured to be sigma(p (t) over bar -> t (t) over bar +X)=6.6 +/- 0.9(stat+syst) +/- 0.4(lum) pb. This cross section is the most precise D0 measurement to date for t (t) over bar production and is in good agreement with standard model expectations.

Analysis of secondary phases segregated and precipitated in SnO2-based varistors

Transmission and scanning electron microscopy techniques were used to study the heterogeneities found in the microstructure of (SnO2Co3O4Nb2O5Fe2O3)-Co-.-Nb-.-Fe-. and (SnO2ZnONb2O5FC2O3)-Zn-.-Nb-.-F-. varistors. Second phases encountered both inside the grains and ingrain boundary regions were identified using energy dispersive spectrometry and electron diffraction patterns. Through the electrical characterisation, the presence of iron oxide among the additives was determined to highlight the non-linear properties of the specimens. A discussion on the influence of second phases on the non-linear features of these systems is also addressed. (C) 2004 Elsevier Ltd. All rights reserved.

Organic complexation and translocation of ferric iron in podzols of the Negro River watershed. Separation of secondary Fe species from Al species

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Field-free and field-stimulated electron emission from solids

Electron irradiation of solids produces a backemission of secondary electrons (energies between 0 and 50 eV) and reflected primaries (energies between 50 eV and that of the incident beam). For insulators, it is shown that an externally applied positive electric field penetrating into the solid material, energizes electrons generated by the primary irradiation and enables them to travel back to the surface of incidence and be emitted (stimulated secondary emission).

The ultrastructural aspects of vitellogenesis or oocyte secondary growth in Serrasalmus spilopleura (Teleostei, Characiformes, Serrasalminae)

Oocyte secondary growth in S. spiloleura corresponds to the period in which different vesicular structures are formed, including the cortical alveoli and the yolk granules. The oocytes with cortical alveolus formation show vesicular structures with filamentous content in the cortical cytoplasmic region, which are the cortical alveolus precursors. In these oocytes, electron-dense vesicles of heterogenous content are dispersed in the inner cytoplasmic region and their nuclei are irregular, showing many nucleoli of different sizes. The oocytes in vitellogenesis are filled with many vesicles. The cortical alveolus precursors are in the peripheral region, and electron-dense granules are seen near to the nucleus. These fuse and form yolk granules. The oocytes in vitellogenesis show a very irregular nucleus that has nucleoli of different sizes. In the oocytes in final vitellogenesis, the yolk granules are scattered throughout the cytoplasm, displacing the cortical alveoli toward cell periphery. The nucleus is similar to the other stages.

Low energy elastic and electronically inelastic electron scattering from biomolecules.

Reactions initiated by collisions with low-energy secondary electrons has been found to be the prominent mechanism toward the radiation damage on living tissues through DNA strand breaks. Now it is widely accepted that during the interaction with these secondary species the selective breaking of chemical bonds is triggered by dissociative electron attachment (DEA), that is, the capture of the incident electron and the formation of temporary negative ion states [1,2,3]. One of the approaches largely used toward a deeper understanding of the radiation damage to DNA is through modeling of DEA with its basic constituents (nucleotide bases, sugar and other subunits). We have tried to simplify this approach and attempt to make it comprehensible at a more fundamental level by looking at even simple molecules. Studies involving organic systems such as carboxylic acids, alcohols and simple ¯ve-membered heterocyclic compounds are taken as starting points for these understanding. In the present study we investigate the role played by elastic scattering and electronic excitation of molecules on electron-driven chemical processes. Special attention is focused on the analysis of the in°uence of polarization and multichannel coupling e®ects on the magnitude of elastic and electronically inelastic cross-sections. Our aim is also to investigate the existence of resonances in the elastic and electronically inelastic channels as well as to characterize them with respect to its type (shape, core-excited or Feshbach), symmetry and position. The relevance of these issues is evaluated within the context of possible applications for the modeling of discharge environments and implications in the understanding of mutagenic rupture of DNA chains. The scattering calculations were carried out with the Schwinger multichannel method (SMC) [4] and its implementation with pseudopotentials (SMCPP) [5] at di®erent levels of approximation for impact energies ranging from 0.5 eV to 30 eV. References [1] B. Boudai®a, P. Cloutier, D. Hunting, M. A. Huels and L. Sanche, Science 287, 1658 (2000). [2] X. Pan, P. Cloutier, D. Hunting and L. Sanche, Phys. Rev. Lett. 90, 208102 (2003). [3] F. Martin, P. D. Burrow, Z. Cai, P. Cloutier, D. Hunting and L. Sanche, Phys. Rev. Lett. 93, 068101 (2004). [4] K. Takatsuka and V. McKoy, Phys. Rev. A 24, 2437 (1981); ibid. Phys. Rev. A 30, 1734 (1984). [5] M. H. F. Bettega, L. G. Ferreira and M. A. P. Lima, Phys. Rev. A 47, 1111 (1993).

Electron spectroscopy of novel charge transfer systems based on polycyclic aromatic hydrocarbons

Organische Ladungstransfersysteme weisen eine Vielfalt von konkurrierenden Wechselwirkungen zwischen Ladungs-, Spin- und Gitterfreiheitsgraden auf. Dies führt zu interessanten physikalischen Eigenschaften, wie metallische Leitfähigkeit, Supraleitung und Magnetismus. Diese Dissertation beschäftigt sich mit der elektronischen Struktur von organischen Ladungstransfersalzen aus drei Material-Familien. Dabei kamen unterschiedliche Photoemissions- und Röntgenspektroskopietechniken zum Einsatz. Die untersuchten Moleküle wurden z.T. im MPI für Polymerforschung synthetisiert. Sie stammen aus der Familie der Coronene (Donor Hexamethoxycoronen HMC und Akzeptor Coronen-hexaon COHON) und Pyrene (Donor Tetra- und Hexamethoxypyren TMP und HMP) im Komplex mit dem klassischen starken Akzeptor Tetracyanoquinodimethan (TCNQ). Als dritte Familie wurden Ladungstransfersalze der k-(BEDT-TTF)2X Familie (X ist ein monovalentes Anion) untersucht. Diese Materialien liegen nahe bei einem Bandbreite-kontrollierten Mottübergang im Phasendiagramm.rnFür Untersuchungen mittels Ultraviolett-Photoelektronenspektroskopie (UPS) wurden UHV-deponierte dünne Filme erzeugt. Dabei kam ein neuer Doppelverdampfer zum Einsatz, welcher speziell für Milligramm-Materialmengen entwickelt wurde. Diese Methode wies im Ladungstransferkomplex im Vergleich mit der reinen Donor- und Akzeptorspezies energetische Verschiebungen von Valenzzuständen im Bereich weniger 100meV nach. Ein wichtiger Aspekt der UPS-Messungen lag im direkten Vergleich mit ab-initio Rechnungen.rnDas Problem der unvermeidbaren Oberflächenverunreinigungen von lösungsgezüchteten 3D-Kristallen wurde durch die Methode Hard-X-ray Photoelectron Spectroscopy (HAXPES) bei Photonenenergien um 6 keV (am Elektronenspeicherring PETRA III in Hamburg) überwunden. Die große mittlere freie Weglänge der Photoelektronen im Bereich von 15 nm resultiert in echter Volumensensitivität. Die ersten HAXPES Experimente an Ladungstransferkomplexen weltweit zeigten große chemische Verschiebungen (mehrere eV). In der Verbindung HMPx-TCNQy ist die N1s-Linie ein Fingerabdruck der Cyanogruppe im TCNQ und zeigt eine Aufspaltung und einen Shift zu höheren Bindungsenergien von bis zu 6 eV mit zunehmendem HMP-Gehalt. Umgekehrt ist die O1s-Linie ein Fingerabdruck der Methoxygruppe in HMP und zeigt eine markante Aufspaltung und eine Verschiebung zu geringeren Bindungsenergien (bis zu etwa 2,5eV chemischer Verschiebung), d.h. eine Größenordnung größer als die im Valenzbereich.rnAls weitere synchrotronstrahlungsbasierte Technik wurde Near-Edge-X-ray-Absorption Fine Structure (NEXAFS) Spektroskopie am Speicherring ANKA Karlsruhe intensiv genutzt. Die mittlere freie Weglänge der niederenergetischen Sekundärelektronen (um 5 nm). Starke Intensitätsvariationen von bestimmten Vorkanten-Resonanzen (als Signatur der unbesetzte Zustandsdichte) zeigen unmittelbar die Änderung der Besetzungszahlen der beteiligten Orbitale in der unmittelbaren Umgebung des angeregten Atoms. Damit war es möglich, präzise die Beteiligung spezifischer Orbitale im Ladungstransfermechanismus nachzuweisen. Im genannten Komplex wird Ladung von den Methoxy-Orbitalen 2e(Pi*) und 6a1(σ*) zu den Cyano-Orbitalen b3g und au(Pi*) und – in geringerem Maße – zum b1g und b2u(σ*) der Cyanogruppe transferiert. Zusätzlich treten kleine energetische Shifts mit unterschiedlichem Vorzeichen für die Donor- und Akzeptor-Resonanzen auf, vergleichbar mit den in UPS beobachteten Shifts.rn

Identification and dissolution behavior of the secondary uranium minerals in the corrosion products of Depleted Uranium (DU) ammunition formed in soils

In der vorliegenden Studie wurden verschiedene Techniken eingesetzt um drei Proben (4, 7, and 8) die aus denrnKorrosionsprodukten von aus dem Kosovo Krieg stammenden Munitionskugeln, bestehend aus abgereichertem Uranrn(Depleted Uranium - DU), zu untersuchen. Als erstes Verfahren wurde die Raman-Spektroskopie eingesetzt. Hierbeirnzeigte sichin den Proben, charakterisiert durch einen Doppelpeak, die Anwesenheit von Schoepitrn(UO2)8O2(OH)12(H2O)12. Der erste und zweite Peakzeigte sich im Spektralbereich von 840,3-842,5 cm-1rnbeziehungsweise 853,6-855,8 cm-1. Diese Werte stimmen mit den Literaturwerten für Raman-Peaks für Schoepitrnüberein. Des Weiteren wurde bei dieser Untersuchungsmethode Becquerelite Ca(UO2)6O4(OH)6(H2O)8 mit einemrnPeak im Bereich zwischen 829 to 836 cm-1 gefunden. Aufgrund des Fehlens des Becquerelitespektrums in derrnSpektralbibliothek wurde eine in der Natur vorkommende Variante analysiert und deren Peak bei 829 cm-1 bestimmt,rnwas mit den Ergebnissen in den Proben korrespondiert. Mittels Röntgenbeugung (X-Ray Diffraction, XRD) zeigtenrnsich in allen Proben ähnliche Spektren. Das lässt darauf schließen, dass das pulverisierte Material in allen Probenrndas gleiche ist. Hierbei zeigte sich eine sehr gute Übereinstimmung mit Schoepit und/oder meta-rnSchoepit(UO2)8O2(OH)12(H2O)10, sowie Becquerelite. Weiterhin war weder Autunit, Sabugalit noch Uranylphosphatrnanwesend, was die Ergebnisse einer anderen Studie, durchgeführt an denselben Proben, wiederlegt. DiernAnwesenheit von P, C oder Ca im Probenmaterial konnte ausgeschlossen werden. Im Falle von Calciumkann diesrnmit der Anwesenheit von Uran erklärt werden, welches aufgrund seines Atomradius bevorzugt in Becquerelite (1:6)rneingebaut wird. Die beiden Hauptpeaks für Uran lagen im Falle von U 4f 7/2 bei 382.0 eV und im Falle von U 4f 5/2rnbei 392 eV. Diese Werte mit den Literaturwerten für Schoepit und meta-Schoepitüberein. Die Ergebnissernelektronenmikroskopischen Untersuchung zeigen U, O, Ca, Ti als dominante Komponenten in allen Messungen.rnElemente wie Si, Al, Fe, S, Na, und C wurden ebenfalls detektiert; allerdings kann nicht ausgeschlossen werden,rndass diese Elemente aus dem Boden in der unmittelbaren Umgebung der Munitionsgeschosse stammen. Gold wurdernebenfalls gemessen, was aber auf die Goldarmierung in den Probenaufbereitungsbehältern zurückgeführt werdenrnkann. Die Elektronenmikroskopie zeigte außerdem einige Stellen in denen elementares Uran und Bodenmineralernsowie sekundäre Uranminerale auftraten. Die Elementübersicht zeigt einen direkten Zusammenhang zwischen U andrnCa und gleichzeitig keine Korrelation zwischen U und Si, oder Mg. Auf der anderen Seite zeigte sich aber einrnZusammenhang zwischen Si und Al da beide Konstituenten von Bodenmineralen darstellen. Eine mit Hilfe derrnElektronenstrahlmikroanalyse durchgeführte quantitative Analyse zeigte den Massenanteil von Uran bei ca. 78 - 80%,rnwas mit den 78,2% and 79,47% für Becquerelite beziehungsweise Schoepit aufgrund ihrer Summenformelrnkorrespondiert. Zusätzlich zeigt sich für Calcium ein Massenanteil von 2% was mit dem Wert in Becquerelite (2.19%)rnrecht gut übereinstimmt. Der Massenanteil von Ti lag in einigen Fällen bei 0,77%, was auf eine noch nicht korrodierternDU-Legierung zurückzuführen ist. Ein Lösungsexperiment wurde weiterhin durchgeführt, wobei eine 0,01 M NaClO4-rnLösung zum Einsatz kam in der die verbliebene Probensubstanz der Korrosionsprodukte gelöst wurde;rnNatriumperchlorate wurde hierbei genutzt um die Ionenstärke bei 0,01 zu halten. Um Verunreinigungen durchrnatmosphärisches CO2 zu vermeiden wurden die im Versuch für die drei Hauptproben genutzten 15 Probenbehälterrnmit Stickstoffgas gespült. Eine Modelkalkulation für den beschriebenen Versuchsaufbau wurde mit Visual MINTEQrnv.3.0 für die mittels vorgenannten Analysemethoden beschriebenen Mineralphasen im pH-Bereich von 6 – 10 imrnFalle von Becquerelite, und Schoepit berechnet. Die modellierten Lösungskurven wurden unter An- und Abwesenheitrnvon atmosphärischem CO2 kalkuliert. Nach dem Ende des Lösungsexperiments (Dauer ca. 6 Monate) zeigten diernKonzentrationen des gelösten Urans, gemessen mittels ICP-OES, gute Übereinstimmung mit den modelliertenrnSchoepit und Becquerelite Kurven. Auf Grund des ähnlichen Löslichkeitverhaltens war es nicht möglich zwichen denrnbeiden Mineralen zu unterscheiden. Schoepit kontrolliert im sauren Bereich die Löslichkeit des Urans, währendrnbecquerelit im basichen am wenigsten gelöst wird. Des Weiteren bleibt festzuhalten, dass ein Anteil an CO2 in diernverschlossenen Probenbehälter eingedrungen ist, was sich mit der Vorhersage der Modeldaten deckt. Die Löslichkeitrnvon Uran in der Lösung als Funktion des pH-Wertes zeigte die niedrigsten Konzentrationen im Falle einer Zunahmerndes pH-Wertes von 5 auf 7 (ungefähr 5,1 x 10-6 mol/l) und einer Zunahme des pH-Wertes auf 8 (ungefähr 1,5 x 10-6rnmol/l bei). Oberhalb dieses Bereichs resultiert jeder weitere Anstieg des pH-Wertes in einer Zunahme gelösten Uransrnin der Lösung. Der ph-Wert der Lösung wie auch deren pCO2-Wert kontrollieren hier die Menge des gelösten Urans.rnAuf der anderen Seite zeigten im Falle von Becquerelite die Ca-Konzentrationen höhere Werte als erwartet, wobeirnwahrscheinlich auf eine Vermischung der Proben mit Bodensubstanz zurückgeführt werden kann. Abschließendrnwurde, unter Berücksichtigung der oben genannten Ergebnisse, eine Fallstudie aus Basrah (Irak) diskutiert, wo inrnzwei militärischen Konflikten Uranmunition in zwei Regionen unter verschiedenen Umweltbedingungen eingesetztrnwurden.

Improvement of photon transport model by including coupled photon-electron transport and kernel refinement

The first part of this work deals with the inverse problem solution in the X-ray spectroscopy field. An original strategy to solve the inverse problem by using the maximum entropy principle is illustrated. It is built the code UMESTRAT, to apply the described strategy in a semiautomatic way. The application of UMESTRAT is shown with a computational example. The second part of this work deals with the improvement of the X-ray Boltzmann model, by studying two radiative interactions neglected in the current photon models. Firstly it is studied the characteristic line emission due to Compton ionization. It is developed a strategy that allows the evaluation of this contribution for the shells K, L and M of all elements with Z from 11 to 92. It is evaluated the single shell Compton/photoelectric ratio as a function of the primary photon energy. It is derived the energy values at which the Compton interaction becomes the prevailing process to produce ionization for the considered shells. Finally it is introduced a new kernel for the XRF from Compton ionization. In a second place it is characterized the bremsstrahlung radiative contribution due the secondary electrons. The bremsstrahlung radiation is characterized in terms of space, angle and energy, for all elements whit Z=1-92 in the energy range 1–150 keV by using the Monte Carlo code PENELOPE. It is demonstrated that bremsstrahlung radiative contribution can be well approximated with an isotropic point photon source. It is created a data library comprising the energetic distributions of bremsstrahlung. It is developed a new bremsstrahlung kernel which allows the introduction of this contribution in the modified Boltzmann equation. An example of application to the simulation of a synchrotron experiment is shown.

Generalized eMC implementation for Monte Carlo dose calculation of electron beams from different machine types

The electron Monte Carlo (eMC) dose calculation algorithm available in the Eclipse treatment planning system (Varian Medical Systems) is based on the macro MC method and uses a beam model applicable to Varian linear accelerators. This leads to limitations in accuracy if eMC is applied to non-Varian machines. In this work eMC is generalized to also allow accurate dose calculations for electron beams from Elekta and Siemens accelerators. First, changes made in the previous study to use eMC for low electron beam energies of Varian accelerators are applied. Then, a generalized beam model is developed using a main electron source and a main photon source representing electrons and photons from the scattering foil, respectively, an edge source of electrons, a transmission source of photons and a line source of electrons and photons representing the particles from the scrapers or inserts and head scatter radiation. Regarding the macro MC dose calculation algorithm, the transport code of the secondary particles is improved. The macro MC dose calculations are validated with corresponding dose calculations using EGSnrc in homogeneous and inhomogeneous phantoms. The validation of the generalized eMC is carried out by comparing calculated and measured dose distributions in water for Varian, Elekta and Siemens machines for a variety of beam energies, applicator sizes and SSDs. The comparisons are performed in units of cGy per MU. Overall, a general agreement between calculated and measured dose distributions for all machine types and all combinations of parameters investigated is found to be within 2% or 2 mm. The results of the dose comparisons suggest that the generalized eMC is now suitable to calculate dose distributions for Varian, Elekta and Siemens linear accelerators with sufficient accuracy in the range of the investigated combinations of beam energies, applicator sizes and SSDs.

Observation and applications of single-electron charge signals in the XENON100 experiment

The XENON100 dark matter experiment uses liquid xenon in a time projection chamber (TPC) to measure xenon nuclear recoils resulting from the scattering of dark matter weakly interacting massive particles (WIMPs). In this paper, we report the observation of single-electron charge signals which are not related to WIMP interactions. These signals, which show the excellent sensitivity of the detector to small charge signals, are explained as being due to the photoionization of impurities in the liquid xenon and of the metal components inside the TPC. They are used as a unique calibration source to characterize the detector. We explain how we can infer crucial parameters for the XENON100 experiment: the secondary-scintillation gain, the extraction yield from the liquid to the gas phase and the electron drift velocity.

Modeling Techniques for the High-Resolution Interpretation of Cryo-Electron Microscopy Reconstructions

Essential biological processes are governed by organized, dynamic interactions between multiple biomolecular systems. Complexes are thus formed to enable the biological function and get dissembled as the process is completed. Examples of such processes include the translation of the messenger RNA into protein by the ribosome, the folding of proteins by chaperonins or the entry of viruses in host cells. Understanding these fundamental processes by characterizing the molecular mechanisms that enable then, would allow the (better) design of therapies and drugs. Such molecular mechanisms may be revealed trough the structural elucidation of the biomolecular assemblies at the core of these processes. Various experimental techniques may be applied to investigate the molecular architecture of biomolecular assemblies. High-resolution techniques, such as X-ray crystallography, may solve the atomic structure of the system, but are typically constrained to biomolecules of reduced flexibility and dimensions. In particular, X-ray crystallography requires the sample to form a three dimensional (3D) crystal lattice which is technically di‑cult, if not impossible, to obtain, especially for large, dynamic systems. Often these techniques solve the structure of the different constituent components within the assembly, but encounter difficulties when investigating the entire system. On the other hand, imaging techniques, such as cryo-electron microscopy (cryo-EM), are able to depict large systems in near-native environment, without requiring the formation of crystals. The structures solved by cryo-EM cover a wide range of resolutions, from very low level of detail where only the overall shape of the system is visible, to high-resolution that approach, but not yet reach, atomic level of detail. In this dissertation, several modeling methods are introduced to either integrate cryo-EM datasets with structural data from X-ray crystallography, or to directly interpret the cryo-EM reconstruction. Such computational techniques were developed with the goal of creating an atomic model for the cryo-EM data. The low-resolution reconstructions lack the level of detail to permit a direct atomic interpretation, i.e. one cannot reliably locate the atoms or amino-acid residues within the structure obtained by cryo-EM. Thereby one needs to consider additional information, for example, structural data from other sources such as X-ray crystallography, in order to enable such a high-resolution interpretation. Modeling techniques are thus developed to integrate the structural data from the different biophysical sources, examples including the work described in the manuscript I and II of this dissertation. At intermediate and high-resolution, cryo-EM reconstructions depict consistent 3D folds such as tubular features which in general correspond to alpha-helices. Such features can be annotated and later on used to build the atomic model of the system, see manuscript III as alternative. Three manuscripts are presented as part of the PhD dissertation, each introducing a computational technique that facilitates the interpretation of cryo-EM reconstructions. The first manuscript is an application paper that describes a heuristics to generate the atomic model for the protein envelope of the Rift Valley fever virus. The second manuscript introduces the evolutionary tabu search strategies to enable the integration of multiple component atomic structures with the cryo-EM map of their assembly. Finally, the third manuscript develops further the latter technique and apply it to annotate consistent 3D patterns in intermediate-resolution cryo-EM reconstructions. The first manuscript, titled An assembly model for Rift Valley fever virus, was submitted for publication in the Journal of Molecular Biology. The cryo-EM structure of the Rift Valley fever virus was previously solved at 27Å-resolution by Dr. Freiberg and collaborators. Such reconstruction shows the overall shape of the virus envelope, yet the reduced level of detail prevents the direct atomic interpretation. High-resolution structures are not yet available for the entire virus nor for the two different component glycoproteins that form its envelope. However, homology models may be generated for these glycoproteins based on similar structures that are available at atomic resolutions. The manuscript presents the steps required to identify an atomic model of the entire virus envelope, based on the low-resolution cryo-EM map of the envelope and the homology models of the two glycoproteins. Starting with the results of the exhaustive search to place the two glycoproteins, the model is built iterative by running multiple multi-body refinements to hierarchically generate models for the different regions of the envelope. The generated atomic model is supported by prior knowledge regarding virus biology and contains valuable information about the molecular architecture of the system. It provides the basis for further investigations seeking to reveal different processes in which the virus is involved such as assembly or fusion. The second manuscript was recently published in the of Journal of Structural Biology (doi:10.1016/j.jsb.2009.12.028) under the title Evolutionary tabu search strategies for the simultaneous registration of multiple atomic structures in cryo-EM reconstructions. This manuscript introduces the evolutionary tabu search strategies applied to enable a multi-body registration. This technique is a hybrid approach that combines a genetic algorithm with a tabu search strategy to promote the proper exploration of the high-dimensional search space. Similar to the Rift Valley fever virus, it is common that the structure of a large multi-component assembly is available at low-resolution from cryo-EM, while high-resolution structures are solved for the different components but lack for the entire system. Evolutionary tabu search strategies enable the building of an atomic model for the entire system by considering simultaneously the different components. Such registration indirectly introduces spatial constrains as all components need to be placed within the assembly, enabling the proper docked in the low-resolution map of the entire assembly. Along with the method description, the manuscript covers the validation, presenting the benefit of the technique in both synthetic and experimental test cases. Such approach successfully docked multiple components up to resolutions of 40Å. The third manuscript is entitled Evolutionary Bidirectional Expansion for the Annotation of Alpha Helices in Electron Cryo-Microscopy Reconstructions and was submitted for publication in the Journal of Structural Biology. The modeling approach described in this manuscript applies the evolutionary tabu search strategies in combination with the bidirectional expansion to annotate secondary structure elements in intermediate resolution cryo-EM reconstructions. In particular, secondary structure elements such as alpha helices show consistent patterns in cryo-EM data, and are visible as rod-like patterns of high density. The evolutionary tabu search strategy is applied to identify the placement of the different alpha helices, while the bidirectional expansion characterizes their length and curvature. The manuscript presents the validation of the approach at resolutions ranging between 6 and 14Å, a level of detail where alpha helices are visible. Up to resolution of 12 Å, the method measures sensitivities between 70-100% as estimated in experimental test cases, i.e. 70-100% of the alpha-helices were correctly predicted in an automatic manner in the experimental data. The three manuscripts presented in this PhD dissertation cover different computation methods for the integration and interpretation of cryo-EM reconstructions. The methods were developed in the molecular modeling software Sculptor (http://sculptor.biomachina.org) and are available for the scientific community interested in the multi-resolution modeling of cryo-EM data. The work spans a wide range of resolution covering multi-body refinement and registration at low-resolution along with annotation of consistent patterns at high-resolution. Such methods are essential for the modeling of cryo-EM data, and may be applied in other fields where similar spatial problems are encountered, such as medical imaging.

Structural conservations and diversities of dsRNA viruses: Structural studies of the cytoplasmic polyhedrosis virus by electron cryomicroscopy and 3D reconstruction

The Reoviridae virus family is a group of economically and pathologically important viruses that have either single-, double-, or triple-shelled protein layers enclosing a segmented double stranded RNA genome. Each virus particle in this family has its own viral RNA dependent RNA polymerase and the enzymatic activities necessary for the mature RNA synthesis. Based on the structure of the inner most cores of the viruses, the Reoviridae viruses can be divided into two major groups. One group of viruses has a smooth surfaced inner core, surrounded by complete outer shells of one or two protein layers. The other group has an inner core decorated with turrets on the five-fold vertices, and could either completely lack or have incomplete outer protein layers. The structural difference is one of the determinant factors for their biological differences during the infection. ^ Cytoplasmic polyhedrosis virus (CPV) is a single-shelled, turreted virus and the structurally simplest member in Reoviridae. It causes specific chronic infections in the insect gut epithelial cells. Due to its wide range of insect hosts, CPV has been engineered as a potential insecticide for use in fruit and vegetable farming. Its unique structural simplicity, unparalleled capsid stability and ease of purification make CPV an ideal model system for studying the structural basis of dsRNA virus assembly at the highest possible resolution by electron cryomicroscopy (cryoEM) and three-dimensional (3D) reconstruction. ^ In this thesis work, I determined the first 3D structure of CPV capsids using 100 kV cryoEM. At an effective resolution of 17 Å, the full capsid reveals a 600-Å diameter, T = 1 icosahedral shell decorated with A and B spikes at the 5-fold vertices. The internal space of the empty CPV is unoccupied except for 12 mushroom-shaped densities that are attributed to the transcriptional enzyme complexes. The inside of the full capsid is packed with icosahedrally-ordered viral genomic RNA. The interactions of viral RNA with the transcriptional enzyme complexes and other capsid proteins suggest a mechanism for RNA transcription and subsequent release. ^ Second, the interactions between the turret proteins (TPs) and the major capsid shell protein (CSPs) have been identified through 3D structural comparisons of the intact CPV capsids with the spikeless CPV capsids, which were generated by chemical treatments. The differential effects of these chemical treatment experiments also indicated that CPV has a significantly stronger structural integrity than other dsRNA viruses, such as the orthoreovirus subcores, which are normally enclosed within outer protein shells. ^ Finally, we have reconstructed the intact CPV to an unprecendented 8 Å resolution from several thousand of 400kV cryoEM images. The 8 Å structure reveals interactions among the 120 molecules of each of the capsid shell protein (CSP), the large protrusion protein (LPP), and 60 molecules of the turret protein (TP). A total of 1980 α-helices and 720 β-sheets have been identified in these capsid proteins. The CSP structure is largely conserved, with the majority of the secondary structures homologous to those observed in the x-ray structures of corresponding proteins of other reoviruses, such as orthoreovirus and bluetongue virus. The three domains of TP are well positioned to play multifunctional roles during viral transcription. The completely non-equivalent interactions between LPP and CSP and those between the anchoring domain of TP and CSP account for the unparalleled stability of this structurally simplest member of the Reoviridae. ^

Geochemical analyses of secondary minerals from ODP Leg 115 basalts (Tables 1+2)

Basalts recovered along the Reunion hotspot track on Ocean Drilling Program (ODP) Leg 115 range in age from 34 Ma at Site 706 to 64 Ma at Site 707. They have undergone various degrees of secondary alteration. Within single holes the amount of alteration can vary from a few percent to near complete replacement of phenocrysts and groundmass by secondary minerals. Olivine appears to be the most susceptible to alteration and in some sections it is the only mineral altered. In other sections, olivine, pyroxene and plagioclase phenocrysts, and groundmass have been completely replaced by secondary minerals. Clays are the predominant form of secondary mineralization. In addition to replacing olivine, pyroxene, glass, and groundmass, clays have filled veins, vesicles, and voids. Minor amounts of calcite, zeolites, and K-feldspar were also detected. The clays that filled vesicles and veins often show color zonations of dark, opaque bands near the edges that grade into tan or green transparent regions in the centers of the veins. The electron microprobe was used to obtain chemical analyses of these veins as well as to characterize isolated clays that replaced specific minerals and filled voids and vesicles.