No description available.

The formation of bacterial biofilms on surfaces and their subsequent biofouling pose extensive safe and healthy concerns to a variety of industries. Biofilms are ubiquitous, and the biofilm state is considered the default mode of growth for the majority of the world's bacteria population. Once mature, biofilms are difficult to remove completely and have improved resistance against antibacterial agents. Given this, there has been significant interest to mitigate or at least manage biofilm formation on surfaces. One such method has been through the material design of surfaces, and to the interest of this study, through the development of antimicrobial stainless steels. Stainless steel is not an inherently antimicrobial material. Stainless steels alloyed with small amounts of either copper (Cu) or silver (Ag), both well-known natural antimicrobial agents, have been investigated since their initial development in the late 1990's onward. This class of materials have been proven to show significant antimicrobial effect over their traditional counterparts without compromising the characteristic mechanical properties of the stainless steels. However, most of the antimicrobial assessments for these materials documented within literature are conducted over a 24-hour timeframe and do not adequately account for the biofilm mode of growth. As so, this study aimed to assess how biofilms grow on this class of antimicrobial steels over a longer duration of growth and under growth conditions which more adequately modeled the biofilm mode of life. The same strain of Escherichia coli commonly used in antimicrobial surface testing, ATCC 8739, was grown on submicron-polished coupons of a ferritic Cu-alloyed stainless steel (1.50 wt. % Cu), an austenitic Ag-alloyed stainless steel (0.042wt. % Ag), and a standard 304 series stainless steel, used as a baseline. Following ASTM-E2647-13, the E. coli/SS coupons were grown using a drip flow bioreactor under low shear conditions at either ambient temperature or 37 ± 3 degrees C with a batch phase of 6 hours and a continuous phase of 48 hours up to 96 hours. Directly after harvesting, the coupons were analyzed with an Environmental Scanning Electron Microscope (E-SEM) under low vacuum with a water vapor environment. The effect of surface chemistry and alloy microstructure, surface roughness, rinsing the surfaces prior to inoculation and after harvesting, temperature, and growth duration on the resulting E. coli biofilms were all investigated in some capacity. Growth on the submicron finished surfaces indicated there were no significant differences between the biofilms grown on the three different steel compositions. Bacterial attachment appeared non-preferential to surface chemistry or alloy microstructure, suggesting that E. coli interacted with the surfaces effectively the same under the given growth conditions. To account for apparent randomness in bacterial attachment, it is hypothesized that the surface features of interest were on a size scale irrelevant to the size of single bacterial cells. To account for the lack of an observed biocidal effect from the Cu- and Ag-alloyed stainless steels, it is hypothesized that an organic conditioning film which developed on the surfaces from the fluid environment may have effectively inhibited the release of Cu and Ag ions from the steel surfaces. / MS / Bacteria frequently self-organize into what are commonly called bacterial biofilms, or an aggregation of bacterial cells that attach to a surface and which are embedded within a self-generated matrix of polymeric substances, such as proteins and polysaccharides. The biofilm state offers a lot of survival advantages to bacteria, and once biofilms form on a surface they are very difficult to remove. The formation of bacterial biofilms on surfaces and their subsequent biofouling pose extensive safe and healthy concerns to a variety of industries. There has been significant interest to stop or at least manage biofilm formation on surfaces. One such method has been through the design of surfaces, and to the interest of this study, through the development of antimicrobial stainless steels. Stainless steel is not an inherently antimicrobial material. Stainless steels which include small amounts of either copper or silver have been proven to show a significant antimicrobial effect over their traditional stainless steel counterparts without compromising the other desirable properties of the steels. However, most of the documented antimicrobial assessments for these materials have been conducted over a 24-hour timeframe and do not adequately account for the biofilm mode of growth. This study aimed to assess how biofilms grow on this class of steels over a longer duration of growth and under growth conditions which more adequately modeled the biofilm mode of life. This was done by growing a single strain of E. coli bacteria onto coupons of these stainless steel materials for either a 48-hour or a 96-hour timeframe within a low-flow, continuously-fed bioreactor. The coupons were visualized with an environmental scanning electron microscope to assess the effect of the material properties on the observed biofilms grown during this study. Overall there were little differences observed between the E. coli biofilms grown on the copper-containing stainless steel, the silver-containing stainless steel, and the standard stainless steel used within this study. Mirror finish smooth surfaces were needed in order to adequately visualize the steel coupons. The bacteria appeared to attach randomly without any preference for steel surface chemistry or other surface features. This suggested that under the given growth conditions the bacteria interacted with the smooth steel surfaces the same. To account for this randomness, it is hypothesized that the relevant surface features were significantly smaller than the size of single bacterial cells. E. coli cells are between 1 – 2 micrometers long and 0.5 – 1 micrometers in diameter. There was also no antimicrobial effect observed on the copper-containing and silver-containing stainless steels. To account for the lack of an observed antimicrobial effect, it is hypothesized that a conditioning film of carbon-based molecules formed on the surface of the steels from the liquid growth medium environment, preventing bacterial cells from being damaged by the copper and silver within the steel surfaces.

Lithium-ion batteries have become a vital part of our modern life and play an essential role in electric vehicle development. One of the most feasible strategies to enhance the energy density of Li-ion batteries is to use layered, Ni-rich cathode materials. However, higher nickel content causes several problems and therefore, several methods, including doping and coating, have been utilized to stabilize their structure and boost their performance. This thesis aims to understand the microstructure of such engineered complex cathodes and provide valuable contributions by comprehensively understanding and establishing a link between the composition, structure, performance, and properties of these complex materials. In this regard, the most advanced electron- and photon-based techniques have been used to uncover the fundamental underlying reasons for the enhanced performance or degradation in these complex cathode structures. This study shows that introducing W cation inside the LiNiO2 results in new W-variants with a heterogeneous concentration on the top surface and through grain boundaries of the host secondary particles. These W-rich regions play a reinforcing role in grain boundaries and protect the outer surface of LiNiO2 particles. However, synthesis defects, such as porosities, could reduce these benefits by increasing the electrolyte infiltration inside the cathode particles. It is also demonstrated that the degradation process can be studied through the changes in electron energy loss near-edge structure spectra. The investigation of a coating approach on LiNi0.8Co0.15Al0.05O2 materials through the mechanofusion process illustrates more microscopic-scale details regarding the thickness unevenness of the coating and some degree of physical intermixing between the core (LiNi0.8Co0.15Al0.05O2) and coating (LiFePO4 and alumina) precursors. In addition to good physical contact between the core and coating materials, further analysis at higher resolution reveals some nanoscale grains and defective areas near the top surface of the secondary particles following the mechanofusion coating process. / Thesis / Doctor of Philosophy (PhD)

The pursuit of high quality, large area graphene has been a major research focus of contemporary materials science research, in the wake of the discovery of the multitude of exceptional properties exhibited by the material. The DPhil project was undertaken with the objective of developing an understanding of the growth of large graphene sheets by chemical vapour deposition (CVD), and also in the subsequent characterisation of their material properties. By conducting atmospheric pressure CVD growth at high methane flow rates, it was found that few-layered graphene (FLG) could be deposited on a copper catalyst. It is demonstrated that the self-limiting property of a copper catalyst is not universal to all deposition conditions, and shown that FLG grows in a terrace-like configuration. In depth transmission electron microscopy (TEM) studies were carried out on FLG. By selective image reconstruction from the inverse power spectrum of the TEM images, it was possible to elucidate the inter-grain connectivity of few-layer graphenes. It was determined that there were two possible inter-grain configurations possible; specifically an overlap of graphene layers or a discrete atomic bonding edge. The perturbation of the few-layer structure when subject to an out of plane distortion was found to incur a shift in the conventional AB-Bernal stacking of FLG. By utilising the aberration corrected TEM (AC-TEM) at Oxford it was possible to resolve atomic detail in CVD synthesised monolayer films, including atomic bond rotations and vacancies. The use of a high current density at low accelerating voltage (80 kV) was demonstrated to allow for the controlled defect creation in graphene sheets. This permitted the creation of monovacancies and iron doped vacancy complexes suitable for further study. The behaviour of these two defect types under electron beam irradiation was subsequently studied.

Formaldehyde-based adhesives have been reported to be detrimental to health. Petrochemical-based adhesives are non-renewable, limited and costly. Therefore, the improvement of environmental-friendly adhesive from natural agricultural products has awakened noteworthy attention. A novel adhesive for wood application was successfully prepared with enhanced shear strength and water resistance. The Fourier transmform infrared spectra showed the surface functionalities of the functionalized carbon nanotubes (FCNTs) and soy protein isolate nanocomposite adhesive. The attachment of carboxylic functional group on the surface of the carbon nanotubes (CNTs) after purification contributed to the effective dispersion of the CNTs in the nanocomposite adhesive. Hence, enhanced properties of FCNTs were successfully transferred into the SPI/CNTs nanocomposite adhesive. These unique functionalities on FCNTs however, improved the mechanical properties of the adhesive. The shear strength and water resistance of SPI/FCNTs was higher than that of the SPI/CNTs. SEM images showed the homogenous dispersion of CNTs in the SPI/CNTs nanocomposite adhesive. The carbon nanotubes were distributed uniformly in the soy protein adhesive with no noticeable clusters at relatively reduced fractions of CNTs as shown in the SEM images, which resulted into better adhesion on wood surface. Mechanical (shear) mixing and ultrasonication with 30 minutes of shear mixing both showed an improved dispersion of CNTs in the soy protein matrix. However, ultrasonication method of dispersion showed higher tensile shear strength and water resistance than in mechanical (shear) mixing method. Thermogravimetric analysis of the samples also showed that the CNTs incorporated increases the thermal stability of the nanocomposite adhesive at higher loading fraction. Incorporation of CNTs into soy protein isolate adhesive improved both the shear strength and water resistance of the adhesive prepared at a relatively reduced concentration of 0.3%.The result showed that tensile shear strength of SPI/FCNTs adhesive was 0.8 MPa and 7.25MPa at dry and wet state respectively, while SPI/CNTs adhesive had 6.91 MPa and 5.48MPa at dry and wet state respectively. There was over 100% increase in shear strength both at dry and wet state compared to the pure SPI adhesive. The 19% decrease in value of the new adhesive developed compared to the minimum value of ≥10MPa of European standard for interior wood application may be attributed to the presence of metallic particles remaining after purification of CNTs. The presence of metallic particles will prevent the proper penetration of the adhesive into the wood substrate. The type of wood used in this study as well as the processing parameters could also result into lower value compared to the value of European standard. Therefore, optimization of the processing parameter as well as the conversion of carboxylic acid group on the surface of the CNTs into acyl chloride group may be employed in future investigation. However, the preparation of new nanocomposite adhesive from soy protein isolate will replace the formaldehyde and petrochemical adhesive in the market and be of useful application in the wood industry. / Civil and Chemical Engineering / M. Tech. (Chemical Engineering)

La grande majorité des essais cliniques de transfert de gènes in vivo utilise des vecteurs viraux. Si ces derniers sont efficaces, ils présentent des risques immunogènes, toxiques, voire mutagènes avérés. Les vecteurs synthétiques (non viraux), par leur grande modularité et leur faible toxicité représentent une alternative très prometteuse. Le principal frein à leur utilisation est leur manque d’efficacité. L’objectif majeur de ce travail de thèse a été de comprendre le mécanisme de transfert de gènes associé à différents complexes vecteurs synthétiques/ADN plasmidique, ce qui est indispensable pour une conception rationnelle de nouveaux vecteurs. Nous avons étudié, sur cellules en culture, le mécanisme de transfert de gènes associé à deux lipides cationiques ; le BGTC (bis(guanidinium)-tren-cholesterol) et la DOSP (DiOleylamine A-Succinyl-Paromomycine) qui sont connus pour être des vecteurs efficaces in vitro. Nous avons ainsi pu visualiser par microscopie électronique leurs voies d’entrée, leurs remaniements structuraux ainsi que leur échappement endosomal qui représente une étape clé du processus de transfert de gènes. L’identification non ambigüe des lipoplexes tout au long de leur trafic intracellulaire a été rendue possible grâce au marquage de l’ADN par des nanoparticules de silice dotées d’un cœur de maghémite (Fe2O3) dense aux électrons. Cette stratégie de marquage a également été appliquée à l’étude du mécanisme d’action d’un autre vecteur synthétique de type polymère, le copolymère à blocs non ionique P188 ou Lutrol. Contrairement à la plupart des vecteurs synthétiques, celui-ci présente une efficacité de transfection in vivo chez la souris par injection in situ pour le tissu musculaire ou en intra trachéale dans le poumon. En revanche, il est totalement inefficace in vitro. Nous avons montré que le Lutrol permet une augmentation de l’internalisation d’ADN par les cellules mais n’induit pas son échappement endosomal, ce qui expliquerait son absence d’efficacité in vitro. D’autres voies d’entrée sont alors à envisager in vivo pour comprendre son mécanisme d’action. / The vast majority of clinical trials of gene transfer in vivo use viral vectors. Although they are effective, they induce immunogenic, toxic or mutagenic risks. Due to their high modularity and low toxicity, synthetic vectors (non viral), represent a promising alternative despite their lack of effectiveness. The major objective of this work was to understand the mechanism of gene transfer using two prototypic synthetic vectors, in the context of a rational design of new vectors. We studied on cultured cells, the mechanism of action of two cationic lipids; BGTC (bis(guanidinium)-tren-cholesterol) and DOSP (DiOleylamine A-Succinyl-Paromomycine) formulated with plasmid DNA (lipoplexes) which are in vitro efficient vectors. We have been able to visualize by electron microscopy, their intracellular pathways, their structural alterations and their endosomal escape, the latter being a key step in the process of gene transfer. The unambiguous identification of lipoplexes throughout their intracellular trafficking has been made possible thanks to the labelling of DNA by core-shell silica nanoparticles with an electron dense maghemite core (Fe2O3). The labeling strategy has also been applied to study the mechanism of action of a nonionic block copolymer (P188 or Lutrol). Interestingly, these synthetic vectors have an in vivo transfection efficiency in mice lung and muscle tissue while they are totally inefficient in vitro. We have shown that Lutrol induces an increase of DNA internalization into cells and fails to trigger endosomal escape, which would explain the lack of in vitro efficacy. These findings suggest that the in vivo mechanism of action of Lutrol would involve other internalization pathways.

A terapia de fotobiomodulação (PBMT do inglês photobiomodulation therapy) exerce efeitos benéficos em processos relevantes para a regeneração tecidual. A técnica de membranas celulares (CSs; cell sheets) pode gerar grande quantidade de células organizadas em uma matriz extracelular (MEC) produzida por essas células. A constituição de MEC das CSs pode ser de importância para a regeneração de tecidos. O colágeno tipo I, a fibronectina e a tenascina são proteínas da MEC já detectadas em CSs de células-tronco da polpa dentária humana. O objetivo deste estudo foi investigar os efeitos de diferentes parâmetros de PBMT sobre a arquitetura (histologia), composição proteica (Western blotting e imunoistoquímica) e ultraestrutura (MEV e MET) da MEC de CSs de células-tronco da polpa dental humana. As células-tronco foram descongeladas e recaracterizadas através da análise de seu perfil imunofenotípico avaliado pela expressão de moléculas de superfície utilizando citometria de fluxo para marcadores associados a células-tronco mesenquimais (MSC do inglês mesenchymal stem cells; CD105, CD146 e CD44) e não associados (CD45, CD34 e CD14). As CSs foram formadas em placas de cultivo celular após 15 dias em cultura em meio clonogênico suplementado com vitamina C (20 ?g/ml). As culturas celulares foram alocadas em 3 grupos experimentais diferentes, como segue: Controle: nenhum tratamento adicional; PBMT1 e PBMT2. A PBMT foi realizada com um laser diodo vermelho contínuo, aplicando-se os seguintes parâmetros gerais: 660nm, 20mW, 0,028cm2 e 0,71W/cm2. Os parâmetros do PBMT1 foram: 4s, 3J/cm2 e 0,08J por ponto, e o PBMT2: 7s, 5J/cm2 e 0,14J por ponto. As irradiações foram realizadas em dias alternados durante todo o período do experimento, em modo pontual (5 pontos / poço ou 13 por placas de 100 mm de diâmetro) em contato com a base da placa. Após a formação das CSs, 15 dias após o plaqueamento, estas foram submetidas a análises histológica, imunoistoquímica, Western blotting, microscopia eletrônica de transmissão e de varredura. Comparações estatísticas foram realizadas (p <0,05). As células apresentaram perfil imunofenotípico clássico de MSCs, mostrando a expressão de marcadores associados a MSCs, enquanto a expressão dos marcadores não associados a MSCs estavam ausentes. O colágeno tipo I, colágeno tipo III e fibronectina estavam presentes no MEC das CSs. Western blotting revelou maior síntese de fibronectina nas CSs submetidas ao PBMT1. A ultraestrutura geral dos CSs foi diversa nos 3 grupos experimentais. As CSs do grupo PBMT1 apresentaram aspecto epitelióide, enquanto no grupo PBMT2 as CSs apresentaram células isoladas e fusiformes dispostas em feixes unidirecionais. MET identificou uma MEC mais madura e sinais de apoptose nos grupos submetidos à PBMT. A PBMT influenciou a composição e ultraestrutura da MEC de CSs de células-tronco da polpa dentária. Assim, a PBMT pode ser importante na determinação da qualidade mecânica das CSs, o que pode favorecer a terapia celular, facilitando o transplante das células-tronco. / Photobiomodulation therapy (PBMT) improves processes relevant to tissue regeneration. The technique of cell sheets (CSs) can generate large amount of cells organized in an extracellular matrix (ECM) produced by these cells. The constitution of the ECM of CSs could be of importance for tissue regeneration. Type I collagen, fibronectin and tenascin are ECM proteins already detected in CSs of human dental pulp stem cells. The aim of this study was to investigate the effects of different PBMT parameters on the architecture (histology), protein composition (Western blotting and immunohistochemistry) and ultrastructure (SEM and MET) of the MEC of CSs of human dental pulp stem cells. Dental pulp stem cells were thawed and recharacterized by the expression profile of the surface molecules using flow cytometry for mesenchymal stem cells (MSC) -associated (CD105, CD146 and CD44,) and non associated (CD45, CD34 and CD14) markers. The CSs were formed in cell culture plates after 15 days in culture in clonogenic medium supplemented with vitamin C (20 ?g/ml). The cell cultures were allocated in 3 different experimental groups, as follows: Control: no further treatment; PBMT1 and PBMT2. PBMT was performed with CW red diode laser applying the following general parameters: 660nm, 20mW, 0.028cm2 spot size and 0.71W/cm2. The PBMT1 parameters were: 4s, 3J/cm2 and 0.08J per point, and the PBMT2: 7s, 5J/cm2, and 0.14J per point. The irradiations were done on alternate days throughout the experimental time, in a punctual mode (5 points/well or 13 points/100mm-diameter dishes) in contact at the base of the plate. After the CSs formation, 15 days after plating, they were submitted to histology, immunohistochemistry, Western blotting, transmission electron microscopy and scanning electron microscopy analyses. Statistical comparisons were performed (p<0.05). The cells presented the classical immunoprofile of MSCs showing the expression of MSCs-associated markers, whereas the expression of the MSCs non-associated markers were absent. The type I collagen, type III collagen and fibronectin were present in the MEC of the CSs. Western blotting revealed a higher amount of fibronectin in the CSs submitted to PBMT1. The overall ultrastructure of the CSs was diverse in the 3 experimental groups. PBMT1 leads to epithelial-like CS, whereas the PBMT2 leads to a CSs with isolated fusiform cells arranged in unidirectional bundles. MET identified a more mature ECM and signs of apoptosis in the PBMT groups. PBMT may influence the composition and ultrastructure of the ECM of CSs of dental pulp stem cells. Thus, PBMT could be of importance in the determination of the mechanical quality of CSs, which may favor cell therapy by improving the CS transplantation approach.

Les maladies ostéoarticulaires représentent environ 10% de l’ensemble des pathologiessurvenant en France chaque année. Les difficultés pour identifier les causes de ces maladiesproviennent pour une part d’un manque de compréhension du mécanisme de lubrificationd’une articulation synoviale saine. Dans ce contexte, un premier objectif de ce travail a été d’analyser la structurediscontinue du liquide synovial à partir de prélèvements animaux sains et de la reproduire àpartir de composants biomoléculaires commerciaux afin de comprendre le mécanisme delubrification dans le cas sain. Le deuxième objectif de cette thèse a été d’analyser l’évolution de la structure et despropriétés lubrifiantes du liquide synovial dans le stade précoce de pathologies noninflammatoiresou inflammatoires à partir de prélèvements pathologiques humains. Afind’étudier plus finement l’évolution de la lubrification pathologique cette thèse a visé àdévelopper aussi des modèles de lubrifiants obtenus à partir de cultures cellulaires desynoviocytes humains en combinant l’action de facteurs inflammatoire (cytokines).L’ensemble des résultats montre l’importance de la structure supramoléculaire duliquide synovial dans l’obtention de bonnes propriétés lubrifiantes ; cette relation devraitconstituer d’une part un paramètre clé dans le diagnostic précoce des pathologies articulaireset d’autre part une voie de développement de liquides thérapeutiques à base de lubrifiantsnanostructurés / Osteoarticular diseases account for approximately 10% of all diseases occurring inFrance each year. The difficulties in identifying their causes are also due to a deficiency inunderstanding the lubrication mechanism of healthy synovial joint.In this context, the first objective of the present study was to analyze the discontinuousstructure of synovial fluid samples from healthy animals and to reproduce it using commercialbiomolecular components in order to understand the lubrication mechanism in the healthycase. The second objective of this thesis was to analyze the evolution of the structure andthe lubricating properties of synovial fluid in the early stage of non-inflammatory andinflammatory diseases using pathological human fluid samples. In order to study moreprecisely the evolution of pathological lubrication this work aimed to develop lubricantmodels obtained from cell cultures of human synoviocytes adding the action of pathologicalinflammatory factors (cytokines). All together the results show the importance of the supramolecular structure of thesynovial fluid in obtaining good lubricating properties what may be a key parameter in theearly diagnosis of joint diseases and even more a chance to develop therapeutic fluids basedon nanostructured lubricants

Avanços na tecnologia introduziram o laser como um grande aliado na remoção seletiva do tecido cariado. Entretanto, o efeito dos lasers e seus diferentes parâmetros nos tecidos duros dentais e fibrilas colágenas não foi completamente estudado. O objetivo deste estudo foi avaliar os efeitos da irradiação com os lasers de Er:YAG e Er,Cr:YSGG na morfologia e na resistência de união da dentina irradiada à resina composta. Sessenta e quatro hemi-discos de dentina obtidos de terceiros molares humanos foram aleatoriamente divididos em 9 grupos (n=7): G1 Controle (não irradiado); G2 Laser de Er:YAG 250 mJ, 4 Hz; G3 Laser de Er:YAG 200 mJ, 4 Hz; G4 Laser de Er:YAG 180 mJ, 10 Hz; G5 Laser de Er:YAG 160 mJ, 10 Hz; G6 - Laser de Er,Cr:YSGG 2W, 20 Hz; G7 - Laser de Er,Cr:YSGG 2,5W, 20 Hz; G8 - Laser de Er,Cr:YSGG 3W, 20 Hz; G9 - Laser de Er,Cr:YSGG 4W, 20 Hz. Destes, vinte e sete amostras (n=3) foram processadas e destinadas à avaliação em Microscopia Eletrônica de Varredura (MEV) para análise morfológica de superfície e em corte transversal da dentina irradiada. Nove fragmentos (n=1) foram submetidos à Microscopia Eletrônica de Transmissão (MET) e vinte e sete (n=3) foram restaurados para análise da interface adesiva. Para o teste de resistência de união, 45 superfícies dentinárias planificadas foram restauradas e, após 24 horas de armazenamento em água, foram submetidas ao teste de microtração. O teste de ANOVA (=5%) foi realizado e verificou-se que o G1 apresentou os maiores valores de resistência de união quando comparado aos demais grupos experimentais. Não houve diferença estatisticamente significativa entre os grupos irradiados, e a análise morfológica revelou para estes grupos uma superfície irregular, com aparência escamosa, sem smear layer e com os túbulos dentinários abertos. Nos cortes transversais foi possível observar que esses efeitos se estendem à subsuperfície dentinária e resultam na formação de um padrão modificado dos tags durante o processo de hibridização. Os resultados deste estudo in vitro sugerem que a irradiação com os lasers de Er:YAG e Er,Cr:YSGG promovem um padrão morfológico específico da superfície dentinária, os quais podem interferir negativamente nos valores de resistência de união à resina composta. / Improvements on technology have introduced the laser as a great ally in the selected removal of dental caries. However, the different laser parameters and their effects on dental hard tissues and collagen fibrils have not yet been completely studied. The aim of the present study was to evaluate the effect of Er:YAG and Er,Cr:YSGG laser irradiation on dentin morphology and on microtensile bond strength of resin composite to the irradiated dentin. Sixty-four dentin hemi-disks obtained from sound human third molars were randomly divided into 9 groups (n=7): G1 Control, G2 Er:YAG Laser 250 mJ, 4 Hz; G3 Er:YAG Laser 200 mJ, 4 Hz; G4 Er:YAG Laser 180 mJ, 10 Hz; G5 Er:YAG Laser 160 mJ, 10 Hz; G6 - Er,Cr:YSGG Laser 2W, 20 Hz; G7 - Er,Cr:YSGG Laser 2,5W, 20 Hz; G8 - Er,Cr:YSGG Laser 3W, 20 Hz; G9 - Er,Cr:YSGG Laser 4W, 20 Hz. Twenty seven specimens (n=3) were processed for Scanning Electron Microscopy (SEM) for surface and cross-sectional morphological analysis. Nine specimens (n=1) were processed for Transmission Electron Microscopy (TEM) and twenty seven (n=3) were restored to evaluate the adhesive interface. For the microtensile bond strength (^TBS) test, 45 flat occlusal dentin surfaces were restored and after storage in water for 24h, were submitted to ^TBS test. ANOVA (=5%) showed that G1 (control group) presented the highest ^TBS values when compared with the irradiated groups, which did not differ from each other. SEM analysis showed an irregular scaly surface, free of smear layer and with open dentinal tubules. The cross-sectional dentin micrographs showed that the effects of laser extended to the dentin subsurface resulting in a pattern of tags modified in the hibridization process. The findings of this in vitro study showed that both Er:YAG and Er,Cr:YSGG laser irradiation resulted in a specific morphological pattern of dentin that negatively affects the bond strength to resin composite to such dentin.

X. fastidiosa é uma bactéria fitopatogênica limitada ao xilema, que tem afetado um grande número de plantas no Brasil e no mundo. Muitos trabalhos já foram realizados sobre esta bactéria, mas pouco se conhece a respeito da adesão, colonização e expressão dos sintomas. Os objetivos deste trabalho foram: a) através do uso da microscopia eletrônica e de luz, determinar e relacionar o número de vasos colonizados de citros, cafeeiro e ameixeira com a sintomatologia em folhas; b) estudar a adesão, migração radial e colonização dos vasos do xilema do pecíolo de folhas de citros pela bactéria; c) estudar algumas variáveis experimentais que afetam a expressão dos sintomas em fumo; d) verificar os sítios de ligação da bactéria em cigarrinhas vetores; e) estudar a adesão e a formação do biofilme por X. fastidiosa em superfície de poliestireno, como uma nova metodologia. Os resultados mostraram em ameixeira e cafeeiro uma relação entre o número de vasos colonizados e a expressão de sintomas necróticos, relação esta que não pode ser observada para citros, o qual apresentava um número de vasos colonizados do pecíolo bem menor que o das outras duas espécies. No estudo da bactéria nos vaso do xilema de citros foi possível verificar as diversas fases do processo de colonização do xilema, bem como a capacidade da bactéria em degradar a parede celular primária da pontuação e migrar para os vasos adjacentes. Neste estudo foi também possível verificar respostas da planta à bactérias caracterizadas pela produção de cristais no lúmen dos vasos do xilema e o acúmulo de goma e hiperplasia de células nas folhas. No estudo com variedades de fumo verificou-se que a cultivar Havana apresentou expressão de sintomas mais intensa que as variedades TNN e RP1 e que o aparecimento dos mesmos não foi influenciado pelo volume de inóculo e pelo local de inoculação, mas sim pela adubação com sulfato de amônio, a qual retardou o aparecimento dos sintomas e reverteu os sintomas inicias em folhas após a aplicação. Em cigarrinhas, células bacterianas com morfologia similar as de X. fastidiosa, foram visualizadas aderidas pela parte lateral na câmara do cibário (sulco longitudinal, parede lateral e membrana do diafragma) de Acrogonia citrina, e Oncometopia facialis, no canal do apodeme de Dilobopterus costalimai e pela parte polar no precibário de O. facialis. Finalmente, no estudo da adesão de X. fastidiosa a película de poliestireno, os resultados revelaram as várias fases da formação do biofilme, aspectos da sua arquitetura, e indicaram que a técnica é uma ferramenta adequada para o estudo da formação do biofilme e também da morfologia das bactérias. Os resultados são discutidos em termos de modelos de adesão e colonização, da bactéria e importância para o conhecimento dos mecanismos de patogenicidade da bactéria em plantas e transmissão pelos vetores. / X. fastidiosa is a xylem-limited bacterium that has been affecting a high number of plants in Brazil and in the world. A lot of researches were already accomplished on this bacterium, but little is known regarding the adhesion, colonization and expression of the symptoms in plants. The objectives of this work were: a) through the use of electron microscopy and of light microscopy determine and to correlate the number of xylem colonized vessels of petiole of sweet orange, coffee and plum with chlorosis and leaf scorching in leaves; b) study the adhesion, radial migration and colonization of the vessels of the petiole xylem of sweet orange by the bacterium; c) study some experimental variables that affect the expression of symptoms in tobacco; d) verify the retention sites of the bacterium in sharpshooters; d) study the adhesion and biofilm formation by X. fastidiosa on polystyrene surface. The results showed a relationship between the number of colonized vessels in plum and coffee and the expression of necrotic symptoms. However, that relationship was not observed for sweet orange, which presented a number of colonized vessels smaller than the other two species. In the study of the bacterium in the xylem vessels of sweet orange it was possible to verify the several phases of the colonization process of the xylem as well as the ability of the bacterium to degrade the primary cell wall of the pit and migrate to adjacent vessels. It was also possible to verify responses of the plant to the bacterium characterized by the production of crystals in the lumen of the xylem vessels and gum accumulation and hyperplasia in the leaf cells. Regarding the tobacco varieties it was verified that the expression of symptoms is more intense in the cultivar Havana than in the cultivars TNN and RP1. It was also seen that symptoms expression was not influenced by the inoculum volume or the inoculation place, but it was altered by fertilization with ammonium sulfate, which delayed the beginning of the symptoms and reverted the symptoms in leaves after the application. In sharpshooters, bacterial cells exhibiting morphology similar to X. fastidiosa were visualized attached to the lateral side in the cibarium camera (longitudinal, lateral wall and membrane of the diaphragm) of Acrogonia citrina, and Oncometopia facialis, in the apodemal channel of Dilobopterus costalimai, and in the polar part in the pre-cibarium of O. facialis. Finally, in the study of the adhesion of X. fastidiosa on polystyrene surface, the results revealed the several phases of biofilm formation; aspects of its architecture, and it also indicated that the technique is an appropriate tool to study of the formation of biofilms and also of the bacterial morphology. The results are discussed regarding adhesion models, colonization, and distribution of the bacterium in the plant and the importance of knowing the pathogenicity mechanisms of X. fastidiosa and its transmission by the insect vectors.