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Recirculating aquaculture offers a prospect for successful fish farming, but this form of aquaculture presents a great potential for pathogenic microorganisms to become established in the system through the formation of biofilms. Biofilms are capable of forming on all aquaculture system components, incorporating the various microflora present in the water. Pathogenic microorganisms released from the biofilms are capable of causing recurring exposure to disease in both fish and humans. With the increased consumption of raw and rare fish, the presence of these bacteria in or on the fish could lead to ingestion of pathogens. There is also the possibility of cross-contamination during processing. The objectives of this study were to increase the understanding of pathogen incorporation into biofilms in recirculating aquaculture systems and to determine the effectiveness of various sanitizers in eliminating biofilms. Seven freshwater and two saltwater facilities were sampled, with eight different types of materials tested. Pathogenic bacteria were identified using Bacteriological Analytical Manual methods and rapid commercial test kits. Most of the pathogenic bacteria identified were opportunistic organisms ubiquitous in an aquatic environment. The most significant human pathogens were Bacillus cereus, the Shigella species and the Vibrio species. The major piscine pathogens of concern were Photobacterium damsela, the Vibrio species, and Aeromonas hydrophila. The most significant variation in biofilm pathogens was observed between facilities and not construction material. Buna-N rubber, polyvinyl chloride (PVC), chlorinated PVC, glass, fiberglass and stainless steel disks were suspended in 79.2 liter (20 gallon) aquariums stocked with Nile tilapia (Oreochromus niloticus). The tanks were inoculated with a known amount of green fluorescent protein (GFP) modified Escherichia coli and samples were removed on days 1,3, 7 and 15. The modified E. coli were isolated on Luria Broth Agar and plate counts were performed under ultraviolet light. There was no significant difference in the growth of the surrogate pathogen on the different materials. The GFP E. coli was isolated in the largest numbers 24 hours after inoculation of the tanks, with an approximate 1-log decrease after day 1. Days 3, 7, and 15 showed equivalent growth of the target organism. Two sets of disks were suspended in another six 79.2 liter (20 gallon) aquariums. The tanks were inoculated with a known amount of the surrogate pathogen, GFP E. coli, and after 24 hours one set of disks was removed from each tank. The second set of disks was removed and treated by spraying with water, alkaline cleanser, sodium hypochlorite, quaternary ammonium compound, or peracetic acid. Ozone was bubbled directly into one tank to treat another set of disks. The modified E. coli were isolated and counted. Total aerobic plate counts and Enterobacteriaceae counts were performed. Statistical analysis indicated that the type of material had no significant affect on the effectiveness of the sanitizers. It was determined that sodium hypochlorite (99.4591 overall reduction) and peracetic acid (98.8461 % overall reduction) were the most effective sanitizers overall, and ozone (32.9332 % overall reduction) was the least effective. / Ph. D.

The immune defense is geared to protect against a tremendous array of invaders. The ultimate goal of the immune system is to induce effective and balanced inflammatory responses that enable the efficient elimination of possible threats while avoiding both immunodeficiency and autoimmunity. The skewness towards inflammatory responses causing excessive collateral damage could lead to diverse autoimmune conditions. These conditions could be organ-specific or result from systemic immune dysregulations called systemic autoimmunity. The multifaceted nature and the intricate clinical heterogeneity of systemic autoimmune conditions indicate a strong influence of environmental factors on their immunopathogenesis, where environmental factors could either hinder or contribute to autoimmune development. We focused our research on deciphering the complex effects of environmental factors on the immunopathogenesis of systemic immune dysregulation, taking systemic lupus erythematosus (SLE or Lupus) as a model of systemic autoimmunity. SLE is one of the most mysterious autoimmune disorders with no known cure. In SLE, breaching of tolerance to self-antigens and the subsequent persistent inflammation and collateral tissue damage in multiple organs lead to very diverse clinical manifestations. These manifestations are a result from the interplay between multiple genetic susceptibilities and diverse environmental factors. To date, management plans for SLE are based on non-selective immunosuppressants that could impose significant side effects including increased risks of infection and infection-related mortalities. In parallel, environmental factors and the quality of life could significantly impact SLE management strategies. Therefore, delineating the immunomodulatory capacities of environmental factors would likely unravel more effective management strategies for SLE patients. The current research aims to investigate the central hypothesis that dietary and hygienic components modulate the immune dysregulations of SLE in a tissue- and disease stage-specific manner. We have focused on uncovering the complex effects of Vitamin A (VA) as an essential micronutrient with very diverse immunomodulatory capacities, and quaternary ammonium compound (QAC)-based disinfectants as ubiquitously used disinfectants that have been linked to immunotoxicity, on the immunopathogenesis of SLE. Due to the strong female bias of SLE where women especially of childbearing age are more prone to lupus, we have focused our research on delineating how these diverse factors shape the immunopathogenesis of SLE in female mice only. The first project dissected the immunomodulatory effects of VA, a potent immunomodulatory dietary component. Notably, VA exerts its function through a predominant metabolite known as all-trans-retinoic acid (tRA) that, as we have previously shown, has paradoxical and tissue-specific implications on lupus inflammation. Here, we utilized a pristane-induced model of lupus to investigate the disease stage-dependent effects of tRA. Oral supplementation of tRA was given either before pristane induction of lupus from weaning (3 weeks) to 3 months of age or after pristane induction of lupus from 3 to 9 months of age. We found that tRA treatment mediated disease stage-dependent effects and differentially affected the lupus-associated kidney inflammation (lupus nephritis) when given at the initiation vs. continuation phase of lupus. Unlike tRA treatment during active disease, pre-pristane treatment with tRA aggravated glomerulonephritis through potentiating leukocyte activation and trafficking to the kidney and augmenting renal pro-fibrotic signals. Post-pristane tRA treatment, on the other hand, exerted immunosuppressive functions of decreasing circulatory and renal deposition of autoantibodies as well as suppressing the renal expression of proinflammatory cytokines and chemokines. Interestingly, both pre- and post-pristane treatments with tRA reversed the pristane-induced leaky gut and similarly modulated the gut microbiota, suggesting a gut microbiota-independent mechanism by which tRA affects the initiation vs. continuation phase of lupus. As tRA could be protective against lupus nephritis especially during the active disease stage, and previous reports had shown hypovitaminosis A (reduced serum retinol levels) proceeding SLE, we expanded our investigation to decipher whether VA deficiency (VAD) was a contributing factor for severe SLE and to delineate how VAD affected the initiation and/or the progression of lupus nephritis in genetically-prone conditions. For that purpose, we utilized the classical murine lupus-prone model, MRL/lpr, and initiated VAD either during the gestation or after weaning to reveal potential time-dependent effects. VAD exacerbated lupus nephritis by provoking severe neutrophilic tubulointerstitial nephritis, and accelerated renal failure. This was concomitant with significantly higher mortality in all VAD mice. Mechanistically, VAD enhanced early activation of plasma cells and augmented their autoantibodies production. In addition, VAD led to an enhanced expansion of pathogenic T lymphocytes. In parallel, VAD increased renal infiltration of conventional and plasmacytoid dendritic cells. Our findings establish VAD as a driving factor for lupus nephritis progression in genetically predisposed conditions. These findings emphasize the importance of monitoring VA levels in SLE patients and urge for VA supplementations for patients at higher risk for hypovitaminosis A, especially during the maternal-neonatal interface. Additionally, this project warrants further investigations to delineate the molecular targets through which VA modulates cellular functions as well as immunopathogenesis of lupus nephritis. The information obtained from these studies may also benefit women with other autoimmune conditions and will pave the way for VA supplementations to be tested in clinical trials. The second project investigated the effects of ambient exposure to QAC-based disinfectants on the progression of murine SLE in genetically prone mice. We compared the disease progression in MRL/Lpr mice that have been exposed to QACs vs. those kept under a complete QAC-free condition. QAC-based disinfectants CP-64 or Labsan 256 were used under QAC-exposed conditions, while ethanol was used in the QAC-free environment. We found that compared to QAC-free mice, ambient exposure of lupus-prone mice to QACs led to smaller spleens with no change in circulating autoantibodies or the severity of glomerulonephritis. This suggests that QACs may have immunosuppressive effects on lupus. Using a microfluidic device, we showed that ambient exposure to QACs reduced directional migration of bone marrow-derived neutrophils toward an inflammatory chemoattractant ex vivo. Consistent with this, we found decreased infiltration of neutrophils into the spleen. While bone marrow-derived neutrophils appeared to exhibit a pro-inflammatory profile, upregulated expression of PD-L1 was observed on neutrophils that infiltrated the spleen, which in turn interacted with PD-1 on T cells and modulated their fate. Specifically, QAC exposure hindered activation of splenic T cells and increased apoptosis of effector T-cell populations. Collectively, these results suggest that ambient QAC exposure decreases lupus-associated splenomegaly likely through neutrophil-mediated toning of T-cell activation and/or apoptosis. However, our findings also indicate that even ambient exposure could alter immune cell phenotypes, functions, and their fate. Further investigations on how QACs affect immunity under steady-state conditions are warranted. Collectively, the findings of this doctoral research suggest temporal and spatial effects of diet and hygiene on systemic autoimmunity and emphasize the strong influence of environmental factors toning cellular immune responses and subsequently shaping autoimmune outcomes. Our findings could pave the way for more personalized healthcare plans for autoimmune patients that take into consideration tissue involvement, disease stages, and the patient's lifestyle. / Doctor of Philosophy / The immune system is efficiently toned to discriminate between friends and foes. It effectively protects against a wide array of pathogens while at the same time avoiding attacking self-tissues. The inability of immune defenses to achieve this optimal discrimination could lead to the breakdown of tolerance to self in a wide range of autoimmune conditions. Diverse genetic susceptibilities are implicated in the development of autoimmunity. In parallel, during the recent decades, the tremendous increase in the prevalence of autoimmune conditions coincides with evolving dietary and hygiene styles in Westernized societies. This suggests a strong influence of environmental factors such as dietary and hygienic components on the way that the immune system works. Therefore, the current research investigates whether diet and hygiene modulate the immune dysregulations of lupus disease as a model for systemic autoimmunity; and if so, whether such effects are tissue- and/or disease stage-specific. We utilized different mouse models to delineate the mechanisms by which essential nutrients such as vitamin A (VA) and widely used disinfectant compounds known as quaternary ammonium disinfectants (QACs) modulate the systemic autoimmunity in lupus disease. We found that these modulators influence various aspects of the cellular immune responses including (1) leukocyte activation and subsequent expansion of pathogenic (disease contributing) lymphocytes, production of antibodies directed against self-tissue molecules (i.e., autoantibodies), and production of inflammatory mediators (i.e., cytokines and chemokines); (2) cell trafficking and their infiltration into the tissues; (3) signal transduction pathways that modulate cell fate (e.g., PD-1: PD-L1 signaling). Importantly, environmental modulation of autoimmunity during different stages of autoimmune development could significantly impact the disease outcome. VA treatment, for example, differentially modulates the progression of kidney inflammation when given during the initiation vs. progressive disease stages. Similarly, VA deficiency has the most prominent effects on worsening kidney inflammation under genetically prone conditions when the deficiency is initiated early and at the prenatal stage. In parallel, the effects of environmental factors are also tissue-specific. For example, ambient exposure to QAC-based disinfectants exerted immunosuppressive effects on lupus-associated inflammation of lymphoid tissues with no change in circulating autoantibodies or the severity of kidney inflammation. Collectively, the findings of this doctoral research delineated the cellular mechanisms through which environmental factors could shape autoimmune responses. Further studies will dig into the underlying molecular pathways. Ultimately, our research emphasizes the strong influence of exogenous factors on immunity and will pave the way for more effective healthcare management plans and benefit vulnerable populations affected by autoimmune conditions such as lupus.

Compostos de amônio quaternário (QACs) têm sido amplamente utilizados como desinfetantes e antissépticos, sendo essenciais na prevenção e controle de infecções bacterianas na medicina humana e veterinária. Embora patógenos prioritários multirresistentes têm sido muito bem caracterizados quanto ao perfil de suscetibilidade e contexto genético da resistência aos antibióticos, dados de resistência aos QACs são limitados. Assim, o objetivo do presente estudo foi avaliar a atividade in vitro dos QACs de uso doméstico e hospitalar [cloreto de benzalcônio (BAC), cloreto de cetilpiridinio (CPC) e brometo de cetiltrimetilamônio (CTAB)], contra patógenos prioritários multirresistentes, identificando os principais genes de resistência associados. Foram estudadas 100 cepas multirresistentes previamente sequenciados usando as plataformas Illumina MiSeq e NextSeq representativas de diferentes hospedeiros (humanos e animais) e fontes (ambientes e alimentos). As cepas foram identificadas como Klebsiella pneumoniae (n= 24), Escherichia coli (n= 30); Pseudomonas aeruginosa (n= 10), Enterobacter spp, (n= 8), Acinetobacter baumannii (n= 11) e Salmonella spp. (n= 17). Genes de resistência aos QACs foram identificados in silico através do alinhamento dos contigs obtidos de cada cepa sequenciada com genes de referência obtidos do GenBank, utilizando o programa Geneious versão 8 (Biomatters Ltd). A identidade de cada gene foi analisada utilizando o programa BLASTx, no qual um critério baseado em ≥90% identidade resultou na identificação dos genes mdfA (77%), qacE (44%), qacEΔ1 (43%), sugE(c) (29%), emrE (21%), qacA (19%), sugE(p) (5%), qacF (7%), qacH (7%) e qacL (7%) em 85 cepas; enquanto que 15 cepas não possuíam nenhum gene de resistência aos QACs. A concentração inibitória mínima (CIM) dos QACs para as 100 cepas foi determinada pelo método de microdiluição em caldo. Os resultados sugeriram que a resistência em patógenos prioritários circulando na interface humano-ambiente-animal não é restrita aos antibióticos, uma vez que a elevada ocorrência de genes qacE, qacEΔ1 e mdfA poderia estar associada com uma redução da suscetibilidade para QACs. Consequentemente, a resistência aos QACs poderia também contribuir para a persistência e adaptação destes patógenos nos seres humanos e outros animais, assim como em ambientes impactados antropogenicamente. / Quaternary ammonium compounds (QACs) have been widely used as disinfectants and antiseptics, being applied as essential compounds in the prevention and control of bacterial infections in human-and veterinary hospital medicine. Although multiresistant priority pathogens have been well characterized with respect to their susceptibility profile and their genetic context of resistance for antibiotics, studies of resistance to QACs are limited. Thus, the objective of the present study was to evaluate the in vitro activity of QACs [(benzalkonium chloride (BAC), cetylpyridinium chloride (CPC) and cetyltrimethylammonium bromide (CTAB)] for household and hospital use against multiresistant priority pathogens, identifying the main resistance genes associated. A hundred multiresistant isolates (previously sequenced using the Illumina MiSeq and NextSeq platforms), representative of different hosts (humans and animals) and sources (environment and food) were studied. Isolates were identified as Klebsiella pneumoniae (n=24), Escherichia coli (n=30), Pseudomonas aeruginosa (n=10), Enterobacter spp. (n=8), Acinetobacter baumannii (n=11) and Salmonella spp. (n=17). In silico analysis for identification of genes conferring resistance to QACs were performed by aligning the contigs obtained from the strains with reference genes deposited in GenBank, using the Geneious version program (Biomatters Ltd). Similarities were analyzed using the BLASTx online program, considering the alignment criteria based on ≥ 90% identity. The result of these analysis revealed the presence of the following QAC genes: mdfA (77%), qacE (44%), qacEΔ1 (43%), sugE(c) (29%), emrE (21%), qacA (19%), sugE (p) (5%), qacF (7%), qacH (7%) e qacL (7%); while 15 strains showed no resistance genes for QACs. Determination of QACs minimum inhibitory concentration (MIC) for the 100 isolates, by the broth microdilution method. These results suggest that resistance to QACs in priority pathogens, circulating at the human-environment-animal interface, is not restricted to antibiotics, since the high occurrence of genes qacE, qacEΔ1 and mdfA were associated with a reduced susceptibility to QACs. Consequently, resistance to QACs could also contribute to the persistence and adaptation of these pathogens in humans and othes animals, as well as in anthropogenically impacted environments.

This thesis details investigations into the generation and synthetic utility of α,β-unsaturated acyl ammonium intermediates using isothioureas as Lewis base organocatalysts to generate a range of heterocyclic products. Initial investigations focussed on the development of a Michael addition-lactonisation protocol utilising α,β-unsaturated acyl ammonium intermediates (generated in situ from HBTM 2.1 and α,β-unsaturated homoanhydrides) and a range of 1,3-dicarbonyl nucleophiles. Products could be isolated as lactones or as ring-opened highly functionalised esters, giving good yields and excellent enantioselectivity. 1,3-Diketones were shown to generate a mixture of regioisomers and whereas 1,3-ketoesters afforded only a single regioisomer. A crystal structure of an α,β-unsaturated acyl ammonium intermediate was obtained that clearly demonstrated steric blocking of the Si- face of the alkene by the catalyst stereodirecting groups, therefore it can be postulated that enantiocontrol in the addition occurs by selective nucleophilic addition from the Re- face. α,β-Unsaturated acyl ammonium species were then shown to participate in asymmetric annulation processes with benzazole nucleophiles to afford highly functionalised heterocyclic products, with both lactone and lactam formation observed. The relationship between nucleophile structure and process regioselectivity was investigated and it was demonstrated that benzothiazole and benzimidazole nucleophiles afforded preferential N-cyclisation to give lactams whilst benzoxazoles exhibited O-cyclisation to form lactones. It was also possible to influence the regioselectivity by changing the electronic properties of the acyl group (R'). Due to the reactivity of this class of nucleophiles it was possible to access products with quaternary centres. Palladium-catalysed cross coupling reactions were also successful on 3-bromo substituted lactams, demonstrating the potential for further derivatising these interesting heterocyclic products. Finally, a cascade protocol was developed that employed Michael-Michael-lactonisation steps to give tricyclic products from enone malonate nucleophiles and α,β-unsaturated acyl ammonium intermediates (generated in situ by addition of HBTM 2.1 into acid chlorides). Interestingly, the reaction showed higher enantioselectivity at elevated temperatures (70 ˚C) and moderate regioselectivity (1,4- vs. 1,2-addition), which could not be improved after extensive screening. A range of lactones was isolated in moderate yields and enantioselectivity.

This study determines the differences in survival of eight Salmonella strains concentration of BAC in water, followed by regrowth in high and low nutrition conditions containing sublethal concentration of BAC. Our results show strain difference in survival, persistence of Salmonella in BAC at different concentrations. All eight strains of Salmonella were non-recoverable when exposed to 40 to 48 µg/ml BAC for 1h in water. When exposed to 24 µg/ml of BAC for 1 h in water, S. Typhimurium ATCC 14028 and S. Heidelberg ATCC 8326 cells were non-detectable from the initial 7 log CFU/ml. S. Blockley 7175, S. Virchow 7207, S. Poona 01A4242 and S. Poona 00A3208, were non-detectable after 1 h exposure at 24 µg/ml and continued to be non-detectable in 12 µg/ml BAC in both nutrient conditions after 24 h. 3/8 strains formed rugose morphotypes when exposed to BAC thus leading to understanding the role of strain differences.

A resina acrílica para base de próteses removíveis é capaz de acumular biofilme e assim favorecer o aparecimento de diversos problemas na cavidade bucal dos usuários de próteses. A imobilização de um agente antisséptico na matriz polimérica tem potencial preventivo frente a esse acúmulo, mas necessita de maior investigação. O objetivo deste estudo foi avaliar as propriedades biológicas do brometo de metacriloiloxiundecilpiridínio (MUPB), um composto antisséptico capaz de copolimerizar-se com as resinas acrílicas. Foram determinadas as concentrações inibitória e fungicida/bactericida mínimas (CIM, CFM/CBM) do MUPB frente às espécies Candida albicans, Candida dubliniensis, Candida glabrata, Lactobacillus casei, Staphylococcus aureus e Streptococcus mutans, em comparação ao cloreto de cetilpiridínio (CCP). A seguir, investigou-se a citotoxicidade do MUPB em fibroblastos, comparando-o com o metil metacrilato (MMA). A avaliação da atividade antimicrobiana de diferentes concentrações do MUPB em massa (0, 0,3% e 0,6%) incorporado em uma resina acrílica termopolimerizável para base de próteses foi realizada por meio de testes de difusão em disco e quantificação de unidades formadoras de colônia (UFC) aderidas à resina após contato com suspensões de cada micro-organismo. A adesão microbiana também foi avaliada por microscopia eletrônica de varredura. Comparações entre o MUPB e as demais substâncias, bem como entre as concentrações incorporadas na resina, foram realizadas com α=0,05. O MUPB apresentou CIM inferior ao CCP para C. dublinienses e S. mutans (P=0,046 e 0,043, respectivamente). Para as demais espécies, as diferenças não foram significantes. Para a CFM/CBM, só foi encontrada diferença significante para C. albicans (P=0,046). O MUPB não polimerizado mostrou-se em torno de 20 vezes mais citotóxico que o MMA. Independente da concentração incorporada e da espécie, não houve formação de halo de inibição em torno dos espécimes. A incorporação do MUPB só influenciou na adesão de C. albicans (P=0,003), com menores contagens de UFC para o grupo com 0,6%. Conclui-se que o MUPB não polimerizado tem capacidade antimicrobiana próxima à do CCP, e alta citotoxicidade, se comparado ao MMA. A atividade antimicrobiana, após incorporação em uma resina acrílica para base de prótese, não depende de sua eluição, porém apresentou-se restrita à C. albicans. / Acrylic resins for removable denture base are capable of accumulating biofilm and thus favor the appearance of various problems in the edentulous oral cavity. An antiseptic agent immobilized within the polymeric matrix has a preventive potential against this accumulation, but requires further investigation. The aim of this study was to evaluate the biological properties of methacryloyloxyundecylpyridinium bromide (MUPB), an antiseptic monomer capable of copolymerizing with acrylic resins. The minimum inhibitory and fungicidal/bactericidal concentrations (MIC, MFC/MBC) of MUPB were determined against the species Candida albicans, Candida dubliniensis, Candida glabrata, Lactobacillus casei, Staphylococcus aureus and Streptococcus mutans, in comparison with cetylpyridinium chloride (CCP). After this, the cytotoxicity of MUPB was investigated in fibroblasts, compared with methyl methacrylate (MMA). The antimicrobial activity of different concentrations of MUPB (0, 0.3% and 0.6% w/w) incorporated into a heat polymerized denture base acrylic resin was evaluated by means of disk diffusion tests, and the CFUs adhered to the resin after contact with suspensions of each microorganism were quantified. Microbial adhesion was also evaluated by scanning electron microscopy. Comparisons were made between MUPB and the other substances, as well as between the concentrations incorporated into the resin (α=.05). MUPB presented a lower MIC than CCP for C. dubliniensis and S. mutans (P=.046 and .043, respectively). For the other species, the differences were not significant. For MFC or MBC, significant difference was found only for C. albicans (P=.046). Non polymerized MUPB was shown to be 20 times more cytotoxic than MMA. Irrespective of the concentration incorporated and the species, there was no growth of inhibition halo around the specimens. The incorporation of MUPB only influenced the adhesion of C. albicans (P=.003), with lower CFU counts for the group with the concentration of 0.6%. It was concluded that non polymerized MUPB has an antimicrobial capacity close to that of CCP, and high cytotoxicity when compared with MMA. The antimicrobial activity after incorporation within a denture base acrylic resin did not depend on its elution, but was shown to be restricted to C. albicans.

A resina acrílica para base de próteses removíveis é capaz de acumular biofilme e assim favorecer o aparecimento de diversos problemas na cavidade bucal dos usuários de próteses. A imobilização de um agente antisséptico na matriz polimérica tem potencial preventivo frente a esse acúmulo, mas necessita de maior investigação. O objetivo deste estudo foi avaliar as propriedades biológicas do brometo de metacriloiloxiundecilpiridínio (MUPB), um composto antisséptico capaz de copolimerizar-se com as resinas acrílicas. Foram determinadas as concentrações inibitória e fungicida/bactericida mínimas (CIM, CFM/CBM) do MUPB frente às espécies Candida albicans, Candida dubliniensis, Candida glabrata, Lactobacillus casei, Staphylococcus aureus e Streptococcus mutans, em comparação ao cloreto de cetilpiridínio (CCP). A seguir, investigou-se a citotoxicidade do MUPB em fibroblastos, comparando-o com o metil metacrilato (MMA). A avaliação da atividade antimicrobiana de diferentes concentrações do MUPB em massa (0, 0,3% e 0,6%) incorporado em uma resina acrílica termopolimerizável para base de próteses foi realizada por meio de testes de difusão em disco e quantificação de unidades formadoras de colônia (UFC) aderidas à resina após contato com suspensões de cada micro-organismo. A adesão microbiana também foi avaliada por microscopia eletrônica de varredura. Comparações entre o MUPB e as demais substâncias, bem como entre as concentrações incorporadas na resina, foram realizadas com α=0,05. O MUPB apresentou CIM inferior ao CCP para C. dublinienses e S. mutans (P=0,046 e 0,043, respectivamente). Para as demais espécies, as diferenças não foram significantes. Para a CFM/CBM, só foi encontrada diferença significante para C. albicans (P=0,046). O MUPB não polimerizado mostrou-se em torno de 20 vezes mais citotóxico que o MMA. Independente da concentração incorporada e da espécie, não houve formação de halo de inibição em torno dos espécimes. A incorporação do MUPB só influenciou na adesão de C. albicans (P=0,003), com menores contagens de UFC para o grupo com 0,6%. Conclui-se que o MUPB não polimerizado tem capacidade antimicrobiana próxima à do CCP, e alta citotoxicidade, se comparado ao MMA. A atividade antimicrobiana, após incorporação em uma resina acrílica para base de prótese, não depende de sua eluição, porém apresentou-se restrita à C. albicans. / Acrylic resins for removable denture base are capable of accumulating biofilm and thus favor the appearance of various problems in the edentulous oral cavity. An antiseptic agent immobilized within the polymeric matrix has a preventive potential against this accumulation, but requires further investigation. The aim of this study was to evaluate the biological properties of methacryloyloxyundecylpyridinium bromide (MUPB), an antiseptic monomer capable of copolymerizing with acrylic resins. The minimum inhibitory and fungicidal/bactericidal concentrations (MIC, MFC/MBC) of MUPB were determined against the species Candida albicans, Candida dubliniensis, Candida glabrata, Lactobacillus casei, Staphylococcus aureus and Streptococcus mutans, in comparison with cetylpyridinium chloride (CCP). After this, the cytotoxicity of MUPB was investigated in fibroblasts, compared with methyl methacrylate (MMA). The antimicrobial activity of different concentrations of MUPB (0, 0.3% and 0.6% w/w) incorporated into a heat polymerized denture base acrylic resin was evaluated by means of disk diffusion tests, and the CFUs adhered to the resin after contact with suspensions of each microorganism were quantified. Microbial adhesion was also evaluated by scanning electron microscopy. Comparisons were made between MUPB and the other substances, as well as between the concentrations incorporated into the resin (α=.05). MUPB presented a lower MIC than CCP for C. dubliniensis and S. mutans (P=.046 and .043, respectively). For the other species, the differences were not significant. For MFC or MBC, significant difference was found only for C. albicans (P=.046). Non polymerized MUPB was shown to be 20 times more cytotoxic than MMA. Irrespective of the concentration incorporated and the species, there was no growth of inhibition halo around the specimens. The incorporation of MUPB only influenced the adhesion of C. albicans (P=.003), with lower CFU counts for the group with the concentration of 0.6%. It was concluded that non polymerized MUPB has an antimicrobial capacity close to that of CCP, and high cytotoxicity when compared with MMA. The antimicrobial activity after incorporation within a denture base acrylic resin did not depend on its elution, but was shown to be restricted to C. albicans.

Les zéolithes sont largement utilisées en catalyse. Un enjeu majeur est d'obtenir des cristaux nanométriques qui offrent des perspectives prometteuses dans la conception de catalyseurs acides plus actifs et plus sélectifs, notamment pour les procédés de conversion des coupes lourdes pétrolières. L'obtention de ces nano-cristaux peut résulter de l'utilisation d'inhibiteurs de croissance. Cette thèse s'est attachée à identifier deux familles de composés organiques limitant la croissance des cristaux. Pour la première, l'inhibition est envisagée par adsorption de composés organiques (polycations, acides aminés...) sur la surface des cristaux en formation. Cette étude a été réalisée en suivant une méthodologie d'expérimentation à haut-débit et a conduit à des cristaux de zéolithe Y (FAU) de 300 nm par l'ajout de L-lysine. La seconde famille est dérivée de l’approche de l’équipe de Ryoo et consiste en l’utilisation de composés bifonctionnels comportant une fonction structurante et une fonction inhibitrice de croissance. Cette étude a démarré par la synthèse de zéolithe MFI. La modélisation moléculaire a permis d'identifier un mono-ammonium alkylé favorisant la formation de nanofeuillets de zéolithe ZSM-5 d'épaisseur voisine de 2 nm. L'étude cinétique a révélé par ailleurs que cette zéolithe est synthétisée à partir d’un polysilicate lamellaire formé in situ. Cette stratégie d'identification, couplée à une méthodologie d'expérimentation à haut débit, a alors été appliquée à la synthèse des zéolithes EMC-1 (FAU) et EMC-2 (EMT), et a conduit à l'élaboration de nouveaux agents structurants et composés bi-fonctionnels. / Zeolites are widely used in catalysis. One of today’s major challenges is to obtain nanometer-sized crystals, offering promising prospects for the design of more active and more selective acid catalysts, in particular for heavy oil conversion processes. Zeolite nanocrystals can be obtained by using growth inhibitors. This thesis focused on the identification of two families of organic compounds limiting the crystals growth. For the first one, the growth inhibition is favored by the adsorption of organic compounds (polycations, amino acids…) on the surface of growing crystals. This study was conducted using a high-throughput experiment methodology and led to zeolite Y (FAU) crystals of 300 nm by the addition of L-lysine. The second family is derived from Ryoo’s team approach and consists of the use of bifunctional compounds including one structure-directing function and one growth-inhibiting function. This study started with the synthesis of MFI zeolite. The molecular modelling allowed identifying an alkyl mono-ammonium directing the formation of 2 nm-thick nanosheets of zeolite ZSM-5. The kinetic study revealed that this zeolite is synthesized from a lamellar polysilicate formed in situ. This identification strategy, coupled to a high-troughput experiment methodology, was applied to the synthesis of zeolites EMC-1 (FAU) and EMC-2 (EMT) and conducted to the elaboration of new structure-directing agents and their bifunctional counterparts.