The diagnostic methods implemented at the Canadian Food Inspection Agency for the detection of Shiga toxin producing E. coli (STEC) are time consuming and tedious, taking up to 5 days before a positive sample can be confirmed. The goal of this project was to streamline the detection procedure for serogroup O157 and 6 important non-O157 serogroups of STEC. Following a short enrichment step (4-6 hrs), two approaches were considered: (1) the filtration of enrichment culture through a gradient of filtration membranes (decreasing pore sizes), followed by capture using specific monoclonal antibody (mAb)-coated Dynabeads, and detection via fluorescence microscopy, (2) the addition of enrichment culture into a flow through system consisting of a column packed with large polystyrene beads (≥ 100 μm) coated with specific mAbs for capture. The results indicate that the filtration approach can only be applied to simpler food matrices. However, at least 100 CFU of the target STEC could be recovered using the filtration system following 4 hrs of enrichment of these matrices spiked with ≤ 15CFU of the target STEC. Similar capture results were obtained in the second approach using specific mAbs immobilized on covalently coupled protein G polystyrene beads and diluted enrichment media. A combination of these strategies together with immunofluorescence microscopy (IMS) and polymerase chain reaction (PCR) could provide diagnostic laboratories with a means to confirm a positive sample within 2 days of testing.

Dans le cadre des procédés de clarification d'eau de surface, les flocs formés au cours des étapes de coagulation et de floculation peuvent être séparés de l'eau traitée par décantation ou par flottation. Dans ce dernier cas, le procédé actuellement en vigueur est la flottation à air dissous et présente un certain nombre d'inconvénients. Ainsi, dans ce contexte, la présente thèse consiste à développer un matériau innovant, à la fois flottant et floculant, pour une application dans un procédé de flottation sans air. Dans un premier temps, l'élaboration de billes de polystyrène expansibles utilisant l'eau ou l'éthanol comme agent gonflant a été réalisée. D'autres billes ont ensuite été préparées en présence de 4-(chlorométhyl)styrène comme co-monomère puis la polymérisation par transfert d'atome amorcée en surface (SI-ATRP) de l'acrylamide a été effectuée avec succès. L'impossibilité d'expanser ces matériaux a ensuite conduit à l'exploration de nouvelles voies de synthèse avec la fonctionnalisation de matériaux flottants existants par des techniques de « grafting from » ou « grafting onto ». Quelle que soit la voie envisagée, la première étape a consisté à réduire les fonctions nitrile en amine primaire en présence d'hydrure d'aluminium lithium. Après fonctionnalisation par du bromure de bromoisobutyryle ou du chlorure d'acryloyle, il a été possible de venir greffer de l'acrylamide par SI-ATRP ou de l'amidon via un amorceur redox, respectivement. Les matériaux flottants/floculants obtenus ont été testés lors de flottatests. Les meilleurs résultats ont été obtenus avec les microsphères fonctionnalisées par de l'amidon anionique. Ces dernières ont permis d'abattre la turbidité de l'eau, ont ensuite été régénérées avec succès, par des bains d'acide oxalique ou de dithionite de sodium, et employées durant plusieurs cycles flottatest/régénération avec des résultats reproductibles. / At the end of clarification process, after coagulation-flocculation steps, flocs can be removed from treated water by settling or flotation. In the latter case, Dissolved Air Flotation is the currently used process. However, this method showed important drawbacks, especially an important energetic cost due to the production of air saturated water. In that context, the goal of the reported work dealt with achieving air-free flotation using innovative floating materials. First of all, the synthesis of expandable polystyrene beads using water or ethanol as blowing agent was investigated. Other beads containing both styrene and 4-(chloromethyl)styrene were prepared. Then, surface initiated atom transfer radical polymerization of acrylamide (SI-ATRP) was achieved. Unfortunately, the expansion of such materials was not possible. Therefore, the second part focused on the functionalization of Expancel beads by “grafting from” or “grafting onto” techniques. The first step consisted in reducing some nitrile functions at the surface into primary amine ones. After functionalization with bromoisobutyryl bromide, the SI-ATRP of acrylamide was performed in water at room temperature. On the other hand, the acryloyl chloride was grafted onto amine functions, and grafting of starch was achieved using a redox initiator. All materials obtained have been used for flocculation/flotation tests and demonstrated satisfactory performances in terms of turbidity removal. Beads functionalized with starch have been successfully regenerated with oxalic acid and sodium dithionite and kept appropriate efficiency during several flotation/regeneration cycles.

Quantum dots (QDs) are potentially advantageous tools for both diagnostics and therapeutics due to their light emitting characteristics. The impact of QDs on biological systems however, is not fully understood. The aim of this project therefore, was to investigate the interaction of a series of different surface modifies QDs with macrophages and their subsequent toxicity. CdTe/CdSe (core), ZnS (shell) QDs with either an organic, COOH or NH2 polyethylene glycol (PEG) surface coatings were used. Fluorescent COOH polystyrene beads (PBs) at (Ø) 20nm and 200nm were also studied. J774.A1 murine ‘macrophage-like' cells were treated for two hours with QDs (40nM) of PBs ($50μg.ml^{-1}$) in the presence of 10% FCS prior to assessment of cellular uptake via confocal microscopy and flow cytometry. COOH and $NH_{2}$ (PEG) QDs, as well as 20nm and 200nm PBs entered macrophages within 30 minutes, and were found to locate within endosomes, lysosomes and the mitochondria. T.E.M. also illustrated particles, including organic QDs, to be present inside J774.A1 cells within membrane- bound vesicles at two hours. Organic QDs were unable to be visualised via fixed cell confocal microscopy. Live cell confocal microscopy (without 10% FCS) did suggest however, that organic QDs entered cells in low quantities up to 30 minutes, after which fluorescence declined. Particle toxicity was determined over 48 hours via the MTT, LDH and GSH assays, as well as via assessment of their potential to produce the pro-inflammatory cytokine (TNF-α) and effect cytosolic $Ca^{2+}$ signalling in the J774.A1 cells. Organic QDs were found to be highly toxic at all time points and concentrations used. Both COOH and $NH_{2 }$ (PEG) QDs induced significant (p<0.0001) cytotoxicity (MTT and LDH assays) at 80nM after 48 hours, as well as significant (p<0.01) GSH depletion over 24 hours at all doses, as well as increasing the level of cytosolic $Ca^{2+}$ at 40nM when assessed over 30 minutes. Organic and NH2 (PEG) QDs were found to significantly increase TNF-α production after 24 hours at 80nM. The findings of this study demonstrate that QDs differ in their uptake by macrophages according to their surface coating, with the organic surface coated QDs being the most toxic. At sub-lethal concentrations, in the presence of 10% FCS, the COOH and $NH_{2}$ (PEG) QDs are taken up resulting in GSH depletion and modulated $Ca^{2+}$ signalling, with $NH_{2}$ (PEG) QDs and organic QDs only eliciting limited TNF-α production. Interestingly however, despite these observations, QD surface coating does not affect the intracellular fate of these NPs, with all of the different surface coated QDs observed to be present in endosomes, lysosomes and the mitochondria within J774.A1 macrophage cells. Therefore, in conclusion, the surface coating of QDs plays a significant role in their interaction with macrophages, their uptake and their subsequent toxicity.