acase@tulane.edu / The objective of this research is to study the surface/interfacial phenomena via video-microscopic observation and quantification inside a micro-channel or microcapillary, which can mimic the operating conditions of practical problems, such as ink-jet, lubricant oil neutralization and enhanced oil recovery. In the second chapter, a micropipette-in-microcapillary method is described for the surface tension measurement at high temperatures, which mimics the dimension and working environment of ink-jet print head. Temperature control within the confined space of a capillary was achieved by coating the outer surface of the housing microcapillary with an electrically conductive, transparent, tin-doped indium oxide (ITO) thin film as a heating jacket. The precision of this technique was discussed according to the comparisons of our results with published reference data for water, n-hexadecane, and n-decane at both room and elevated temperatures. Traditionally, the neutralization of sulfuric acid by engine oils has been the major focus, however, due to the introduce of biofuel or ethanol the acetic acid has become an important concern. In the third chapter, based on micropipette-in-square-channel video-microscopy setup, the neutralization reaction mechanism and reaction kinetics of acetic acid by fully formulated lubricant oil is discussed. It was found that the neutralization exists simultaneously on the oil-acid-interface and bulk-oil phase during the droplet shrinkage. Besides, FTIR and NMR analysis show the neutralization of acetic acid as an instantaneous process, and almost all of the dissolved acetic acid in the bulk is eventually neutralized. Due to the minor role of acetic acid dissolution compared to the interfacial reaction, an interface-reaction-rate-controlled kinetic mechanism is proposed as approximation to describe the neutralization process at different conditions. When glacial and diluted acetic acid droplets were neutralized in fully formulated lubricant oil, the experimentally measured shrinking radius agreed very well with the mathematical model. According to Arrhenius equation, the activation energy of neutralization reaction was determined to be constant and its range (Ea>21 kJ/mol) further validated the assumption of interface-controlled reaction kinetics. In the final chapter, an oil-soluble surfactant prepared by Eni S.p.A. was studied to enhance crude oil mobilization in cryolite-packed miniature bed, which provided a transparent porous media at the microscopic level. When the porous media was imbued with crude oil, the presence of the surfactant in the oil phase was able to improve the mobilization performance of crude oil by flushing. In order to deliver the oil-soluble surfactant and apply it to the removal of crude from porous media, an SDS solution was used to solubilize the surfactant, and the formation of SDS/Eni-Surfactant micellar solution was confirmed by Cryo-SEM images. Using the prepared micellar solutions in oil-removal tests on the packed bed, a very high effectiveness was demonstrated by image binarization, thus confirming the possibility to deliver liposoluble surfactants to the porous-media-trapped crude oil by means of hydrosoluble carriers. / 1 / Yufei Duan

Collecting microlymphatics play a vital role in promoting lymph flow from the initial lymphatics in the interstitial spaces to the large transport lymph ducts. In most tissues, the primary mechanism for producing this flow is the spontaneous contractions of the lymphatic wall. Individual units, known as lymphangion, are separated by valves that help prevent backflow when the vessel contracts, thus promoting flow through the lymphatic network. Lymphatic contractile activity is inhibited by flow in isolated lymphatics, however there are virtually no in situ measurements of lymph flow in these vessels. Initially, a high speed imaging system was set up to image in situ preparations at 500 fps. These images were then manually processed to extract information regarding lymphocyte velocity (-4 to 10 mm/sec), vessel diameter (25 to 165 um), and particle location. Fluid modeling was performed to obtain reasonable estimates of wall shear stress (-8 to 17 dynes/cm2). One of the difficulties encountered was the time consuming methods of manual particle tracking. Using previously captured images, an image correlation method was developed to automate lymphatic flow measurements and to track wall movements as the vessel contracts. Using this method the standard error of prediction for velocity measurements was 0.4 mm/sec and for diameter measurements it was 7.0 µm. It was found that the actual physical quantity being measured through this approach is somewhere between the spatially averaged velocity and the maximum velocity of a Poiseuille flow model.

Collecting microlymphatics play a vital role in promoting lymph flow from the initial lymphatics in the interstitial spaces to the large transport lymph ducts. In most tissues, the primary mechanism for producing this flow is the spontaneous contractions of the lymphatic wall. Individual units, known as lymphangion, are separated by valves that help prevent backflow when the vessel contracts, thus promoting flow through the lymphatic network. Lymphatic contractile activity is inhibited by flow in isolated lymphatics, however there are virtually no in situ measurements of lymph flow in these vessels. Initially, a high speed imaging system was set up to image in situ preparations at 500 fps. These images were then manually processed to extract information regarding lymphocyte velocity (-4 to 10 mm/sec), vessel diameter (25 to 165 um), and particle location. Fluid modeling was performed to obtain reasonable estimates of wall shear stress (-8 to 17 dynes/cm2). One of the difficulties encountered was the time consuming methods of manual particle tracking. Using previously captured images, an image correlation method was developed to automate lymphatic flow measurements and to track wall movements as the vessel contracts. Using this method the standard error of prediction for velocity measurements was 0.4 mm/sec and for diameter measurements it was 7.0 µm. It was found that the actual physical quantity being measured through this approach is somewhere between the spatially averaged velocity and the maximum velocity of a Poiseuille flow model.

Self-assembly of nanoparticles is a promising route to form complex, nanostructured materials with functional properties. Nanoparticle assemblies characterized by a crystallographic alignment of the nanoparticles on the atomic scale, i.e. mesocrystals, are commonly found in nature with outstanding functional and mechanical properties. This thesis aims to investigate and understand the formation mechanisms of mesocrystals formed by self-assembling iron oxide nanocubes. We have used the thermal decomposition method to synthesize monodisperse, oleate-capped iron oxide nanocubes with average edge lengths between 7 nm and 12 nm and studied the evaporation-induced self-assembly in dilute toluene-based nanocube dispersions. The influence of packing constraints on the alignment of the nanocubes in nanofluidic containers has been investigated with small and wide angle X-ray scattering (SAXS and WAXS, respectively). We found that the nanocubes preferentially orient one of their {100} faces with the confining channel wall and display mesocrystalline alignment irrespective of the channel widths.  We manipulated the solvent evaporation rate of drop-cast dispersions on fluorosilane-functionalized silica substrates in a custom-designed cell. The growth stages of the assembly process were investigated using light microscopy and quartz crystal microbalance with dissipation monitoring (QCM-D). We found that particle transport phenomena, e.g. the coffee ring effect and Marangoni flow, result in complex-shaped arrays near the three-phase contact line of a drying colloidal drop when the nitrogen flow rate is high. Diffusion-driven nanoparticle assembly into large mesocrystals with a well-defined morphology dominates at much lower nitrogen flow rates. Analysis of the time-resolved video microscopy data was used to quantify the mesocrystal growth and establish a particle diffusion-based, three-dimensional growth model. The dissipation obtained from the QCM-D signal reached its maximum value when the microscopy-observed lateral growth of the mesocrystals ceased, which we address to the fluid-like behavior of the mesocrystals and their weak binding to the substrate. Analysis of electron microscopy images and diffraction patterns showed that the formed arrays display significant nanoparticle ordering, regardless of the distinctive formation process.  We followed the two-stage formation mechanism of mesocrystals in levitating colloidal drops with real-time SAXS. Modelling of the SAXS data with the square-well potential together with calculations of van der Waals interactions suggests that the nanocubes initially form disordered clusters, which quickly transform into an ordered phase. / <p>At the time of the doctoral defense, the following papers were unpublished and had a status as follows: Paper 4: Manuscript. Paper 5: Manuscript.</p>

The experimental studies presented in this thesis deal with the photoluminescence intermittency of semiconductor quantum dots in different dielectric environments. Detailed analysis of intermittency statistics from single capped CdSe/ZnS, uncapped CdSe and water dispersed CdSe/ZnS QDs in different matrices provide experimental evidence for the model of photoionization with a charge ejected into the surrounding matrix as the source of PL intermittency phenomenon. The distribution of the dark state lifetimes can be described by a power law over a wide range while that of bright state can be described by a power law at shorter times followed by an exponential decay. The lifetimes of the bright and dark states are influenced by the dielectric properties of the surrounding environment. Our experimental results show that the lifetime of the dark state increases with the dielectric constant of the matrix. This is very clear from the linear correlation between αoff and f (ε). We propose a self-trapping model to explain the increase of dark state lifetimes with the dielectric constant of the matrix. A charge will be more stabilized in a medium with high dielectric constant. An energetically more favourable state for an electron in a high dielectric medium decreases the return probability which eventually increases the duration of the off-time. Moreover, the self-trapping model establishes a general model for distribution of states in a matrix. We like to mention, that in the case of bright states, a qualitative observation is the cross over of the on-time power law behavior to an exponential one. The power law part of the decay is nearly matrix independent while the exponential decay, which limits the maximum on-time, strongly depends on dielectric properties of the environment. The exponential part of the on-time probability decays much faster in a high dielectric medium and there exists a linear relation between the time constant of the exponential decay and f (ε). Theoretical background has been provided for the observed results using the recently published DCET model which correlates PL intermittency of QDs with properties of the environment. This supports our previous conjecture of a general model for matrix controlled blinking process. The disagreement between experimentally observed dependence of αoff and f (ε) for different matrices with that of the static tunnelling model proposed by Verberk is due to the fact that the tunneling model considers only an electron transfer between a QD and spatially distributed trap states in vacuum. These states are already stabilized states. It does not assume any medium in between. Therefore, matrix dependent blinking kinetics can not be explained quantitatively by tunneling model even though tunneling between a QD and spatially distributed trap states gives a power law distribution for the blinking kinetics. DCET is a more general (dynamic) model. The bright and dark state parabolas contain QD, charge and the matrix. Therefore, this model could in principle explain matrix dependent blinking kinetics in a better way, for example, the energy difference between the minima of the bright and dark state parabolas (-ΔG0) is defined by the stabilization energy of the system provided by the matrix. However, due to lack of the relevant intrinsic parameters we did not compare this relationship and dependence qualitatively. / Betrachtet man die Fluoreszenz einzelner Farbstoffmoleküle oder Halbleiternanokristalle bei kontinuierlicher Anregung, so stellt man fest, dass die im Zeitverlauf beobachtete Intensität einer stochastischen Variation unterliegt, d. h. dass das Chromophor zwischen emittierenden und nicht emittierenden Zuständen, auch Hell- und Dunkelzuständen genannt, hin- und herschaltet. Dieses als Blinken bekannte Phänomen ist physikalisch wie auch technologisch herausfordernd, lässt es doch einerseits die Realisierbarkeit einer Reihe von quantenoptischen Anwendungen, so z. B. auf dem Gebiet der Quantenkryptographie, dem Quantum Computing oder der optischen Schaltungstechnik auf Basis einzelner Quantenobjekte, in naher Zukunft möglich erscheinen. Andererseits setzt es gewissen Anwendungen, die auf die permanente Sichtbarkeit des Chromophors aufbauen, Grenzen, so zum Beispiel der Verwendung als Lumineszenzmarker in der medizinischen Diagnostik. Weiterhin ist festzustellen, dass das Blinken kritisch von den äußeren Bedingungen und von den Umgebungsparametern abhängt. Aus diesen und anderen Gründen ist ein fundamentales Verständnis der physikalischen Ursachen und der Wechselwirkungsprozesse unerlässlich. Die Forschung dazu steckt noch in den Kinderschuhen. Basierend auf umfangreiche Messungen der Fluoreszenzzeitreihen einzelner Nanokristalle aus CdSe und CdSe/ZnS in verschiedenen Umgebungen, zeigt diese Dissertation exemplarisch den Einfluss der Dielektrizitätsparameter auf das Blinken. Zur Erklärung des Sachverhalts wird ein so genanntes Self-Trapping-Modell zu Rate gezogen. Demnach kommt es zu einer Ionisation des Quantenobjekts und anschließender Ladungstrennung, woraufhin die abgetrennte Ladung für eine gewisse Zeit in der Umgebung lokalisiert bleibt. Die Dauer der Lokalisierung und damit der emittierenden und nicht emittierenden Perioden hängt von der dielektrischen Funktion des umgebenden Materials ab. Dies ist als direkter Nachweis für den photoinduzierten Ladungstransfer als Ursache des Fluoreszenzblinkens zu deuten. Die Arbeit demonstriert, dass die experimentellen Zeitreihen die charakteristischen Merkmale eines diffusionsgesteuerten Ladungstransferprozesses besitzen und nimmt dabei den gegenwärtigen wissenschaftlichen Diskurs über geeignete theoretische Modelle des Fluoreszenzblinkens auf.

Submitted by Luciana Sebin (lusebin@ufscar.br) on 2016-10-06T13:29:02Z No. of bitstreams: 1 DissBCGS.pdf: 7250050 bytes, checksum: df4b2203e5e586a2cba2f75ff4af7f2f (MD5) / Approved for entry into archive by Marina Freitas (marinapf@ufscar.br) on 2016-10-13T20:24:11Z (GMT) No. of bitstreams: 1 DissBCGS.pdf: 7250050 bytes, checksum: df4b2203e5e586a2cba2f75ff4af7f2f (MD5) / Approved for entry into archive by Marina Freitas (marinapf@ufscar.br) on 2016-10-13T20:24:21Z (GMT) No. of bitstreams: 1 DissBCGS.pdf: 7250050 bytes, checksum: df4b2203e5e586a2cba2f75ff4af7f2f (MD5) / Made available in DSpace on 2016-10-13T20:24:30Z (GMT). No. of bitstreams: 1 DissBCGS.pdf: 7250050 bytes, checksum: df4b2203e5e586a2cba2f75ff4af7f2f (MD5) Previous issue date: 2016-02-19 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / Over the last few years, a large number of researchers have directed their efforts and interests for the in vivo study of the cellular and molecular mechanisms of leukocyte-endothelial interactions in the microcirculation of many tissues under different inflammatory conditions. The main goal of these studies is to develop more effective therapeutic strategies for the treatment of inflammatory and autoimmune diseases. Nowadays, analysis of the leukocyte-endothelial interactions in small animals is performed by visual assessment from an intravital microscopy image sequences. Besides being time consuming, this procedure may cause visual fatigue of the observer and, therefore, generate false statistics. In this context, this work aims to study and develop computational techniques for the automatic detection and tracking of leukocytes in intravital video microscopy. For that, results from frame to frame processing (2D – spatial analysis) will be combined with those from the three-dimensional analysis (3D=2D+t – spatio-temporal analysis) of the volume formed by stacking the video frames. The main technique addressed for both processings is based on the analysis of the eigenvalues of the local Hessian matrix. While the 2D image processing aims the leukocyte detection without worrying about their tracking, 2D+t processing is intended to assist on the dynamic analysis of cell movement (tracking), being able to predict cell movements in cases of occlusion, for example. In this work we used intravital video microscopy obtained from a study of Multiple Sclerosis in mice. Noise reduction and registration techniques comprise the preprocessing step. In addition, techniques for the analysis and definition of cellular pathways comprise the post processing step. Results of 2D and 2D+t processing steps, compared with conventional visual analysis, have shown the effectiveness of the proposed approach. / Nos últimos anos, um grande número de pesquisadores tem voltado seus esforços e interesses para o estudo in vivo dos mecanismos celulares e moleculares da interação leucócitoendotélio na microcirculação de vários tecidos e em várias condições inflamatórias. O principal objetivo desses estudos é desenvolver estratégias terapêuticas mais eficazes para o tratamento de doenças inflamatórias e autoimunes. Atualmente, a análise de interações leucócito-endotélio em pequenos animais é realizada de maneira visual a partir de uma sequência de imagens de microscopia intravital. Além de ser demorado, esse procedimento pode levar à fadiga visual do observador e, portanto, gerar falsas estatísticas. Nesse contexto, este trabalho de pesquisa tem como objetivo estudar e desenvolver técnicas computacionais para a detecção e rastreamento automáticos de leucócitos em vídeos de microscopia intravital. Para isso, resultados do processamento quadro a quadro do vídeo (2D – análise espacial) serão combinados com os resultados da análise tridimensional (3D=2D+t – análise espaço-temporal) do volume formado pelo empilhamento dos quadros do vídeo. A principal técnica abordada para ambos os processamentos é baseada na análise local dos autovalores da matriz Hessiana. Enquanto o processamento de imagens 2D tem como objetivo a detecção de leucócitos sem se preocupar com seu rastreamento, o processamento 2D+t pretende auxiliar na análise dinâmica de movimentação das células (rastreamento), sendo capaz de prever movimentos celulares em casos de oclusão, por exemplo. Neste trabalho foram utilizados vídeos de microscopia intravital obtidos a partir de um estudo da Esclerose Múltipla realizado com camundongos. Técnicas de redução de ruído e estabilização do movimento das sequências de imagens compõem a etapa de pré-processamento, assim como técnicas para a definição e análise dos caminhos celulares compõem a etapa de pós-processamento. Resultados das etapas de processamento 2D e 2D+t, comparados com a convencional análise visual, demonstraram a eficácia da abordagem proposta. / FAPESP: 2013/26171-6

The experimental studies presented in this thesis deal with the photoluminescence intermittency of semiconductor quantum dots in different dielectric environments. Detailed analysis of intermittency statistics from single capped CdSe/ZnS, uncapped CdSe and water dispersed CdSe/ZnS QDs in different matrices provide experimental evidence for the model of photoionization with a charge ejected into the surrounding matrix as the source of PL intermittency phenomenon. The distribution of the dark state lifetimes can be described by a power law over a wide range while that of bright state can be described by a power law at shorter times followed by an exponential decay. The lifetimes of the bright and dark states are influenced by the dielectric properties of the surrounding environment. Our experimental results show that the lifetime of the dark state increases with the dielectric constant of the matrix. This is very clear from the linear correlation between αoff and f (ε). We propose a self-trapping model to explain the increase of dark state lifetimes with the dielectric constant of the matrix. A charge will be more stabilized in a medium with high dielectric constant. An energetically more favourable state for an electron in a high dielectric medium decreases the return probability which eventually increases the duration of the off-time. Moreover, the self-trapping model establishes a general model for distribution of states in a matrix. We like to mention, that in the case of bright states, a qualitative observation is the cross over of the on-time power law behavior to an exponential one. The power law part of the decay is nearly matrix independent while the exponential decay, which limits the maximum on-time, strongly depends on dielectric properties of the environment. The exponential part of the on-time probability decays much faster in a high dielectric medium and there exists a linear relation between the time constant of the exponential decay and f (ε). Theoretical background has been provided for the observed results using the recently published DCET model which correlates PL intermittency of QDs with properties of the environment. This supports our previous conjecture of a general model for matrix controlled blinking process. The disagreement between experimentally observed dependence of αoff and f (ε) for different matrices with that of the static tunnelling model proposed by Verberk is due to the fact that the tunneling model considers only an electron transfer between a QD and spatially distributed trap states in vacuum. These states are already stabilized states. It does not assume any medium in between. Therefore, matrix dependent blinking kinetics can not be explained quantitatively by tunneling model even though tunneling between a QD and spatially distributed trap states gives a power law distribution for the blinking kinetics. DCET is a more general (dynamic) model. The bright and dark state parabolas contain QD, charge and the matrix. Therefore, this model could in principle explain matrix dependent blinking kinetics in a better way, for example, the energy difference between the minima of the bright and dark state parabolas (-ΔG0) is defined by the stabilization energy of the system provided by the matrix. However, due to lack of the relevant intrinsic parameters we did not compare this relationship and dependence qualitatively. / Betrachtet man die Fluoreszenz einzelner Farbstoffmoleküle oder Halbleiternanokristalle bei kontinuierlicher Anregung, so stellt man fest, dass die im Zeitverlauf beobachtete Intensität einer stochastischen Variation unterliegt, d. h. dass das Chromophor zwischen emittierenden und nicht emittierenden Zuständen, auch Hell- und Dunkelzuständen genannt, hin- und herschaltet. Dieses als Blinken bekannte Phänomen ist physikalisch wie auch technologisch herausfordernd, lässt es doch einerseits die Realisierbarkeit einer Reihe von quantenoptischen Anwendungen, so z. B. auf dem Gebiet der Quantenkryptographie, dem Quantum Computing oder der optischen Schaltungstechnik auf Basis einzelner Quantenobjekte, in naher Zukunft möglich erscheinen. Andererseits setzt es gewissen Anwendungen, die auf die permanente Sichtbarkeit des Chromophors aufbauen, Grenzen, so zum Beispiel der Verwendung als Lumineszenzmarker in der medizinischen Diagnostik. Weiterhin ist festzustellen, dass das Blinken kritisch von den äußeren Bedingungen und von den Umgebungsparametern abhängt. Aus diesen und anderen Gründen ist ein fundamentales Verständnis der physikalischen Ursachen und der Wechselwirkungsprozesse unerlässlich. Die Forschung dazu steckt noch in den Kinderschuhen. Basierend auf umfangreiche Messungen der Fluoreszenzzeitreihen einzelner Nanokristalle aus CdSe und CdSe/ZnS in verschiedenen Umgebungen, zeigt diese Dissertation exemplarisch den Einfluss der Dielektrizitätsparameter auf das Blinken. Zur Erklärung des Sachverhalts wird ein so genanntes Self-Trapping-Modell zu Rate gezogen. Demnach kommt es zu einer Ionisation des Quantenobjekts und anschließender Ladungstrennung, woraufhin die abgetrennte Ladung für eine gewisse Zeit in der Umgebung lokalisiert bleibt. Die Dauer der Lokalisierung und damit der emittierenden und nicht emittierenden Perioden hängt von der dielektrischen Funktion des umgebenden Materials ab. Dies ist als direkter Nachweis für den photoinduzierten Ladungstransfer als Ursache des Fluoreszenzblinkens zu deuten. Die Arbeit demonstriert, dass die experimentellen Zeitreihen die charakteristischen Merkmale eines diffusionsgesteuerten Ladungstransferprozesses besitzen und nimmt dabei den gegenwärtigen wissenschaftlichen Diskurs über geeignete theoretische Modelle des Fluoreszenzblinkens auf.

Les virus bactériens (bactériophages), durant leur co-évolution avec les bactéries, ont su trouver de nombreuses voies pour détourner les machineries cellulaires dans le but de se multiplier efficacement. L’infection par le phage dès son entrée dans le cytoplasme est un bouleversement pour la bactérie en termes de ressources monopolisées à ses dépens et probablement de restructuration de l’espace cytoplasmique. Dans ce travail de thèse, l’impact de l’infection de la bactérie Gram-positive Bacillus subtilis par le bactériophage SPP1 a été étudié.La réplication de l’ADN est initiée par des protéines précoces virales. Elle mène au chargement de l’hélicase virale gp40 sur l’origine de réplication de SPP1 dont les brins d’ADN ont été ouverts par la protéine de liaison à l’origine, gp38. Le réplisome bactérien est ensuite recruté de manière massive au sein de l’usine de réplication formant un foyer défini dans le cytoplasme bactérien. L’interaction de gp40 avec les protéines cellulaires DnaX et DnaG assure fort probablement le recrutement du complexe cellulaire au foyer de réplication. La quantité d’ADN viral synthétisée représente presque 500 copies d’ADN viral par bactérie après 30 minutes d’infection, ce qui est équivalent à la taille de 5 génomes de B. subtilis. Des études de FRAP (Fluorescence Recovery After Photobleaching) montrent que l’usine de réplication est très dynamique. Ce comportement est inhibé par la présence de HPUra montrant qu’il dépend de la présence d’un réplisome actif.Les concatémères résultant de la réplication de l’ADN viral sont le substrat pour l’encapsidation du génome de SPP1 dans des procapsides préformées. La maturation de ces procapsides en particules virales infectieuses suit une voie d’assemblage spécifique. Deux protéines rapportrices de différentes étapes de cette voie ont été suivies : la protéine d’échafaudage gp11, présente à l’intérieur de la procapside avant encapsidation de l’ADN, et la protéine auxiliaire gp12, qui se fixe à la surface de la capside pendant l’encapsidation. Les procapsides colocalisent partiellement avec l’usine de réplication du génome viral. Après encapsidation de l’ADN, les capsides vont s’accumuler dans des foyers de stockage qui ont une localisation indépendante du foyer de réplication. Cette organisation est également observée dans des bactéries très allongées où deux régions de stockage sont retrouvées situées de part et d’autre de l’usine de réplication mais éloignées des pôles cellulaires. La microscopie électronique combinée à des immuno-marquages révèlent que cette compartimentation corrèle avec une réorganisation majeure de l’ultrastructure du cytoplasme bactérien.L’assemblage et la dynamique des foyers viraux dans la bactérie ont été suivis pendant toute la durée du cycle viral dans un système de microfluidique. Elle montre que les étapes de réplication de l’ADN viral et la formation de la particule du phage sont des processus compartimentés dans le cytoplasme de la bactérie tant spatialement que temporellement. Bien que la croissance cellulaire soit retardée, les bactéries continuent de s’allonger et de se diviser pendant l’infection par SPP1. Le virus exploite donc de manière efficace les machineries cellulaires et l’architecture de la bactérie pour une multiplication optimale. Ces stratégies sont probablement utilisées par de nombreux phages pour remodeler la cellule bactérienne à leur avantage. / During the co-evolution of viruses and cells, viruses exploited numerous ways to hijack cell machineries for their optimal multiplication and dissemination. Phage infection is a major challenge to bacteria, exploiting extensively cellular biosynthetic ressources and possibly re-organizing the cytoplasm space. The work in this thesis investigated the cellular impact of infection by SPP1, a well-characterized model tailed bacteriophage that infects the Gram-positive bacterium Bacillus subtilis.Viral DNA replication is initiated by early phage proteins whose activity culminates in loading of the SPP1 helicase gp40 at the melted phage origin of replication. The bacterial replisome is then massively recruited to the phage replication factory that is localized at a defined position of the cytoplasm. The interaction of gp40 with its two cellular partners DnaX and DnaG mediates most likely the hijacking of the B. subtilis replication machinery. More than 500 copies of the viral genome are synthesized within 30 minutes after initiation of infection, which is roughly the equivalent to five B. subtilis genomes. FRAP (Fluorescence Recovery After Photobleaching) experiments showed that the viral DNA factory is highly dynamic, a behavior that depends on active DNA replication.The concatemers resulting from DNA replication are the substrate for encapsidation of the SPP1 genome into preformed procapsids. Maturation of procapsids to infectious viral particles follows a defined pathway. The SPP1 scaffolding protein gp11, that occupies the interior of the procapsid before DNA packaging, and gp12, that binds to capsids during DNA packaging, were followed to dissect the steps of this process. Procapsids partially co-localize with DNA replication factories. After packaging the DNA-filled capsids fully segregate to spatially distinct warehouses where viral particles accumulate. Recruitment of SPP1 proteins to these compartments recapitulates the sequential order of their assembly to build the viral particle. The replication factory is most frequently flanked by two warehouses. Such pattern is also observed in very elongated cells where the viral compartments remain localized nearby each others and far from the bacterial poles. Immuno-electron microscopy of cryo-sections from infected cells highlights a complete remodelling of the bacterial cytoplasm dedicated to virus multiplication.The assembly and dynamics of the SPP1 replication factory and virions warehouses were visualized during the complete phage infection cycle in microfluidics experiments. The viral compartments are well individualized in the cytoplasm both in terms of space and time. Although bacterial growth is retarded, cells continue to elongate and to divide during SPP1 infection. Structuration of viral factories appears as a very efficient way for SPP1 to exploit bacterial resources and cytoplasmic space to optimize its multiplication. This strategy might be widely used by phages for remodelling the bacterial cell.

Systems of interacting colloidal particles are ideal tools for studies of pattern formation and collective non-equilibrium dynamics on the mesoscopic scale. These processes are governed by the interaction between the particles, which can be tuned by sophisticated fabrication. In this thesis, self-assembly of artificially designed magnetic spheres dispersed in water has been studied via video microscopy. The particles are based on silica microspheres with hemispherical ferromagnetic coating of [Co/Pd] multilayers with perpendicular magnetic anisotropy. These particles are exceptional in that they exhibit an off-centered net magnetic moment and yet obey rotational and mirror symmetry. It has been demonstrated how these magnetic properties provide innovative flexibility in pattern formation and collective dynamics based on magnetostatic interactions on the mesoscopic scale. The results are supported by analytical and numerical calculations of interacting spheres with radially shifted point dipoles (sd-particles). In two dimensions, the particles spontaneously self-assemble into branched structures as a result of a bistable assembly behavior where neighboring particles exhibit a non-collinear magnetic orientation. It has been shown that these features, which are atypical for homogeneous systems of magnetic particles, can be reproduced by simulation. It employs a theoretical model of a sphere that contains a distribution of three radially shifted point dipoles in analogy to the magnetization distribution in the coated particles. The stability of the assembly has been examined further by external manipulation using optical tweezers and homogeneous magnetic fields. A rich variety of stable structures with diverse spatial and magnetic ordering has been found. Particularly, the collective alignment of the specially designed particles in external fields opens completely new possibilities for the remote control over reversible pattern formation on the micrometer scale. In time-dependent fields, the collective dynamics of the anisotropic particles has revealed a novel approach for magnetically actuated translation. The variety of stable structures particularly enables control over this motion. / Kolloidale Suspensionen sind geeignete Systeme zur Untersuchung von Strukturbildung und kollektiver Nichtgleichgewichtsdynamik in mesoskopischen Größenskalen. Diese Vorgänge werden durch die Wechselwirkung zwischen den Teilchen bestimmt, welche durch geeignete Partikelherstellung angepasst werden kann. In der vorliegenden Arbeit wird ein System von künstlich hergestellten magnetischen Partikelsuspensionen mittels Videomikroskopie untersucht. Quarzglas-Mikrokugeln wurden halbseitig mit einer ferromagnetischen Dünnschicht aus [Co/Pd] Multilagen mit senkrechter Anisotropie beschichtet. Solche Partikel sind ausgezeichnet durch ein resultierendes magnetisches Moment mit Rotations- und Spiegelsymmterie, welches zusätzlich vom Mittelpunkt der Kugel verschoben ist. Die vorliegende Arbeit zeigt, dass diese Besonderheit zu einer bisher unbekannten Flexibilität bei der mesoskopischen Strukturbildung und der kollektiven Dynamik auf der Basis magnetostatischer Wechselwirkung führt. Die vorgestellten Ergebnisse werden durch analytische und numerische Berechnungen unterstützt, denen ein Modell einer idealen Kugel mit verschobenem Dipol zugrunde liegt. Die zweidimensionale Selbstanordnung der Partikel zeigt experimentell zwei stabile Formen der Verknüpfung, welche zu verzweigten Strukturen mit unterschiedlich magnetischer Ausrichtung benachbarter Partikel führen. Diese für ein homogenenes System magnetischer Partikel außergewöhnlichen Eigenschaften konnten in Simulationen durch ein Modellsystem aus Kugeln mit drei verschobenen Punktdipolen reproduziert werden. Darüber hinaus wurde die spontante Anordnung unter externer Manipulation mittels optischer Pinzette und magnetischen Feldern untersucht. Es konnte eine Vielfalt an stabilen Strukturen mit verschiedenen magnetischen und strukturellen Anordnungen gefunden werden. Insbesondere die kollektive Ausrichtung dieser Partikel in externen Feldern eröffnet neuartige Möglichkeiten, kontrolliert und reversibel Mikrostrukturen zu erzeugen. In zeitabhängigen Feldern zeigen die anisotropen Partikel zusätzlich eine kollektive Dynamik welche eine neue Möglichkeit zum magnetischen Antrieb von Partikelagglomeraten eröffnet. Die Vielfalt der möglichen stabilen Strukturen erlaubt es in besonderer Weise diese Bewegung zu steuern.

Avec près de 30 millions de morts depuis le début de la pandémie, l’infection par le VIH est un véritable fléau à l’échelle mondiale, surtout en Afrique sub-saharienne. Dans ce contexte, disposer d’une formulation microbicide efficace, facile à administrer par la voie vaginale, représenterait un outil de prévention idéal pour lutter contre cette pandémie. Ainsi, la conception rationnelle de telles formulations représente un enjeu majeur de santé publique.Ce travail décrit la recherche de formulations thermogélifiantes et mucoadhésives à base de pluronics et d’hydroxypropyl méthylcellulose (HPMC). Outre la caractérisation de leurs propriétés rhéologiques et d’adhésion, ainsi que des études en culture cellulaire suggérant une très faible toxicité de contact, ces hydrogels ont montré d’une part, leur capacité à véhiculer efficacement le peptide M48U1 (équipe L. Martin, CEA) destiné à bloquer l’entrée cellulaire du VIH et d’autre part à ralentir considérablement la diffusion de nanoparticules modèles mimant les particules virale matures du VIH-1, en comparaison d’hydrogel d'hydroxy éthylcellulose (HEC) fréquemment utilisés dans divers essais cliniques infructueux et aussi au mucus cervico-vaginal de macaque. L’ensemble des résultats suggère donc la capacité de ces formulations à constituer une double barrière, physique et pharmacologique, protectrice de la muqueuse vaginale vis-à-vis du VIH. / With nearly 30 million deaths since the start of the pandemic, HIV infection is a major problem globally, especially in sub-Saharan Africa. In this context, have an effective microbicide formulation, easily administered by the vaginal route, would be a great prevention tool to fight against this pandemic. Thus, the rational design of such formulations is a major issue of public health.This paper describes research of mucoadhesive and thermogelling formulations based on pluronics and hydroxypropyl methylcellulose (HPMC). Further characterization of their rheological properties and adhesion, and cell culture studies suggesting a very low contact toxicity, these hydrogels showed on one hand, their ability to effectively convey the peptide M48U1 (L. Martin team, CEA) for blocking the entry of HIV cellular and on the other hand to considerably slow down the diffusion of nanoparticles models mimicking viral particles mature HIV-1, compared to hydroxy ethylcellulose hydrogel (HEC) commonly used in various unsuccessful clinical trials and also in cervico-vaginal mucus of macaque. The overall results therefore suggest the ability of these formulations constitute a double barrier, physical and pharmacological, protective of the vaginal mucosa from the HIV.