Certaines microalgues peuvent excréter hors de la cellule des polysaccharides ayant des propriétés intéressant différents secteurs industriels (santé, cosmétique, agroalimentaire, etc.). Les exopolysaccharides (EPS) solubilisés dans le milieu de culture pourraient être concentrés et purifiés par filtration sur membrane après élimination des cellules. Dans ce contexte, la thèse a porté sur l’étude de la filtrabilité des solutions d’EPS de Porphyridium cruentum sur une membrane plane d’ultrafiltration en polyéthersulfone d’un seuil de coupure égal à 50 kDa muni d’un espaceur côté alimentation. Les solutions d’EPS ont été filtrées à petite échelle avec recyclage du rétentat et du perméat dans le bac d’alimentation. Des courbes flux de perméation-pression transmembranaire (PTM) ont été établies jusqu’au flux limite pour différentes conditions de vitesse tangentielle (de 0,3 à 1,2 m.s-1), de teneur en EPS (de 0,10 à 1,06 kg GlcEq.m-3) et de température (20 et 40 °C). Le colmatage de la membrane a été évalué par le biais du calcul des résistances hydrauliques de colmatage, du flux critique, et de cartographies FTIR-ATR et AFM du colmatage irréversible. Les résultats ont montré le caractère fortement colmatant des solutions d’EPS avec une prépondérance de la part du colmatage irréversible pour des pressions de filtration modérées et réverible pour des pressions proches de la PTM limite. Ainsi, la concentration des EPS d’un facteur 10 nécessiterait des vitesses tangentielles de l’ordre de 1 m.s-1, ce qui serait envisageable dans un module plan de type “filtre presse”, mais pas dans un module spiralé. / Some microalgae are able to excrete polysaccharides featuring properties which are of high interest and value to various fields (health, cosmetics, food, etc.). The exopolysaccharides (EPS) solubilized in the culture medium could be concentrated and purified by membrane filtration after removing the cells. Within this context, the thesis focused on the study of the filterability of Porphyridium cruentum EPS solutions on a flat polyethersulfone ultrafiltration membrane with a 50 kDa molecular weight cut-off. The EPS solutions were filtered at a lab-scale with a complete recirculation of both retentate and permeate to the feed tank. Permeate fluxtransmembrane pressure (TMP) curves were established up to the limiting flux for the filtration of solutions with various values of cross-flow velocity (0.3 to 1.2 m.s-1), EPS concentration (0.10 to 1.06 kg GlcEq.m-3) and temperature (20 and 40 °C). Membrane fouling was evaluated through the calculation of hydraulic fouling resistances, the critical flux and FTIRATR and AFM mapping of the irreversible fouling. The results showed the high fouling power of the EPS solutions as well as the preponderance of the irreversible fouling at moderate filtration pressures and the reversible one at pressures close to the limiting TMP. Thus, concentrating EPS by a factor of 10 would require cross-flow velocities of 1 m.s-1, which would be feasible in a flat module (plate and frame filter press) but not in a spiral-wound module.

No description available.

Thèse (M.Sc.)--Université Laval, 2009. / Bibliogr.: f. [110]-120. Publié aussi en version électronique dans la Collection Mémoires et thèses électroniques.

No description available.

Thesis (Ph. D.)--University of Hawaii at Manoa, 1995. / Includes bibliographical references (leaves 215-245). / Microfiche. / xii, 245 leaves, bound ill. (some col.) 29 cm

Thèse (Ph.D.)--Université Laval, 2008. / Bibliogr.: f. [150]-162. Publié aussi en version électronique dans la Collection Mémoires et thèses électroniques.

Lactobacilli are intestinal bacteria with known immunomodulatory competence. Numerous strains of this genus have been implicated in both the prevention and treatment of intestinal inflammation as well as in maintenance of immunological homeostasis. The frequent inclusion of lactobacilli in probiotic products attests to this ability. These lactic acid bacteria colonise the murine forestomach and burgeon in other environments similarly rich in carbohydrate-containing substrates. Accordingly, lactobacilli may utilise fermentable carbohydrates to synthesise exopolysaccharides (EPS). These polymers are secreted into the cellular milieu and, while the ecological function of EPS is yet to be defined, evidence points towards a protective role. This function may include bacterial protection from immunological attack, via EPS recognition by immune cells, resulting in modulation of immunological activity. Dendritic cells (DC) are potent antigen presenting cells, providing an essential link between the innate and adaptive arms of the intestinal immune system. DC efficiently sample intestinal antigens and present peptides to cognate naive CD4⁺ T cells in secondary lymphatic tissue. Under the influence of secreted cytokines, DC direct the differentiation of naive CD4⁺ T cells and therefore, instruct the resultant immune response. Anti-inflammatory Th2 and regulatory T cells can down-regulate the destructive Th1 pro-inflammatory effects associated with inflammatory bowel disease (IBD). As such, bioactives with the aptitude to direct DC activity and T cell differentiation have the potential to prevent or reduce intestinal inflammation. Therefore, this study aimed to determine whether heat-killed EPS-producing strains of lactobacilli, and their secreted EPS, exert an immunomodulatory effect on the murine gut which may down-regulate the immune reactions associated with IBD. Lactobacilli were screened for their ability to produce EPS when grown in the presence of glucose, sucrose or lactose. Heat-killed EPS-producing strains were then used to stimulate bone marrow-derived DC (BMDC) and the resultant cytokine profile was analysed. Nine Lactobacillus strains were found to produce EPS when grown in the presence of sucrose. Of these, L. reuteri 100-23 and L. johnsonii 100-33 exhibited potential anti-inflammatory effects. Therefore, these strains, as well as L. johnsonii 100-5 and L. johnsonii #21, with relatively weak BMDC stimulatory effect, were selected for further investigation. EPS of the potentially anti-inflammatory strain L. reuteri 100-23 was analysed. This sample contained approximately 85% carbohydrate and was composed of a (2[to]6)-β-fructofuranan (levan) and a mannan. The fructan, with an estimated molecular weight of 7 kDa, comprised at least 50% of the EPS, while the mannan made up at least 22%. The mannan component was likely linked to a protein and may have originated from the culture medium. The immunostimulatory capacity of heat-killed Lactobacillus bacterial cells and their EPS was determined in vitro. Firstly, the effect of lactobacilli and EPS on BMDC cytokine secretion, particularly levels of anti-inflammatory IL-10 and pro-inflammatory IL-12, as well as the expression of cell surface activation markers, was determined. L. reuteri 100-23 stimulated relatively high IL-10 secretion but low IL-12, while L. johnsonii 100-33-stimulated BMDC produced elevated levels of both IL-10 and IL-12. All bacterial cells up-regulated co-stimulatory molecules CD40 and CD80 on BMDC. The effect of these stimulated BMDC on T cell proliferation and cytokine production was then assessed, employing the ovalbuminDO11.10 T cell model. L. reuteri 100-23-stimulated BMDC down-regulated T cell production of the proliferation-stimulating cytokine, IL-2, up-regulated regulatory TGF-β secretion, but did not affect pro-inflammatory IFN-γ levels. The EPS of all strains did not stimulate significant BMDC cytokine production and failed to alter BMDC activation marker expression. However, BMDC stimulated with L. reuteri 100-23 and L. johnsonii 100-33 EPS significantly enhanced T cell IL-2 secretion, but did not alter TGF-β or IFN-γ levels. The effect of in vivo L. reuteri 100-23 and EPS intestinal stimulation on the reactivity of immune cells was subsequently investigated. Mesenteric lymph node (MLN) cells and splenic T cells from reconstituted Lactobacillus-free mice fed stimulant or PBS on two occasions were co-cultured with stimulated or unstimulated donor CD11c⁺ splenic DC ex vivo. Cellular proliferation as well as TGF-β and IFN-γ secretion was analysed, and IL-10 neutralisation assays were carried out to ascertain the involvement of this cytokine. Primary exposure of MLN cells to L. reuteri 100-23 resulted in suppressed cell proliferation in the presence of enhanced TGF-β levels, which may have also involved IL-10. Primed splenic T cells exhibited increased proliferation in the presence of elevated TGF-β levels following re-exposure to L. reuteri 100-23, and IL-10 may be involved in limiting this proliferation. L. reuteri 100-23 EPS did not alter MLN cell proliferation, possibly due to the suppressive activity of IL-10, but did enhance that of naive and primed splenic T cells. The effect of ingested L. reuteri 100-23 and EPS on intestinal sIgA concentration was assessed by quantifying IgA levels in the faecal supernatant of RLF mice previously ingesting L. reuteri 100-23 and EPS. L. reuteri 100-23 EPS-fed female mice exhibited significantly elevated levels of sIgA, while heat-killed bacteria did not affect antibody levels. The present study demonstrated that oral administration of heat-killed L. reuteri 100-23 and EPS exerts immunomodulatory effects on the murine intestine. These bioactives may promote a suppressive environment by conditioning DC to secrete a cytokine profile conducive to regulatory T cell induction and memory generation. Additionally, mucosal protection may be favoured by the stimulation of elevated sIgA levels. Therefore, a therapeutic composite is possibly obtained to preserve the intestinal barrier by defending against pathogen-induced injury and buffering inflammatory events. In these ways, L. reuteri 100-23 and EPS may confer long-lasting protection against, and down-regulate the immune reactions associated with, IBD.

The genes coding for the expression of a ropy exopolysaccharide responsible for commercially desirable textural and rhealogical traits in fermented milk products by a natural lactococcal ropy isolate were sought. Using a transposon mutagenesis vector, pGh9:ISS1, three mutants lacking expression of the ropy exopolysaccharide were isolated. One of the mutants was chosen for further characterization. Using a Southern hybridization analysis, the interrupted gene was localized to the chromosome. The non-ropy mutant was further characterized and shown to be unable to produce ropy exopolysaccharide in fermented milk. A 2006 by fragment of the interrupted gene was sequenced. The DNA sequence over a short region showed homology to sugar transfer enzymes found in exopolysaccharide biosynthesis pathways. The DNA sequence was translated into its predicted amino acid sequences and two partial open reading frames of 236 and 338 amino acid residues in length were identified. These open reading frames were found to exhibit identity to glycosyltransferases present in exopolysaccharide biosynthesis pathways in other bacteria. / Graduation date: 1999

L'objectif de cette thèse consistait à analyser la biodiversité des bactériophages infectant Lactococcus lactis. Les bactériophages sont l'une des principales causes d'inhibition de plusieurs procédés de fermentation. L'entrée continuelle de virions via le lait cru et l'absence d'une méthode efficace d'inactivation complète des phages dans l'industrie laitière empêche leur élimination. L'étude de la biodiversité des phages et de leur évolution permettra une action ciblée et une gestion plus intelligente des stratégies anti-phages actuellement disponibles. Depuis quelques années, la classification courante des phages de lactocoques fait l'objet de critiques. Ainsi, les bactériophages de référence représentant les espèces reconnues dans la littérature ainsi que des phages dont l'espèce n'avait pu être identifiée par diverses méthodes ont été étudiés. Une nouvelle proposition de classification contenant dix espèces de phages de L. lactis dont deux nouvelles jamais décrites auparavant a été soumise. De plus, des modifications ont été apportées à la méthode de détection par PCR multiplex des trois espèces prédominantes de phages de L. lactis. La seconde partie du projet porte sur la biodiversité des phages de L. lactis dans une usine québécoise fabriquant du fromage Cheddar. Ainsi, 25 bactériophages de L lactis ont été isolés, classés dans l'une des espèces connues de phages de L. lactis et comparés par leur profil RFLP et leur spectre lytique. Dans la troisième section, l'isolement de huit phages de l'espèce 936 infectant des souches de lactocoques productrices d'exopolysaccharides (EPS) a démontré que les EPS ne représentent pas une barrière efficace contre les phages de cette espèce. Des modifications ont également été proposées pour la méthode de classification des souches EPS+ de L. lactis par analyse du polymorphisme de la taille des fragments de restriction (RFLP). Finalement, bien que l'espèce 936 soit la plus fréquemment rencontrée, uniquement deux séquences génomiques complètes étaient disponibles au début de ce projet. L'analyse de la séquence nucléotidique de trois génomes additionnels apporte de l'information supplémentaire sur la biodiversité des phages à l'intérieur d'une même espèce.

Paenibacillus velaei, sp. nov. is a soil bacterium capable of producing an unusually large amount of exopolysaccharide (EPS). The EPS contains glucose, mannose, galactose and fucose in a molar ratio of 4:2:1:1. The molecular weight of the EPS is higher than 2x106. The viscosity of 1% EPS is 1300 cP when measured at a shear rate of 1 sec-1. Physiological parameters for optimal production of the EPS were studied and it was found that 1.4 g dry weight per 1 l of medium was produced when the bacteria were grown at 30EC and the pH adjusted at 7± 0.2 in a medium containing glucose as the carbon source. Growing the bacteria on different carbon sources did not alter the quantity or the composition of the EPS produced. No toxicity effects were observed in mice or rats when EPS was administered in amounts ranging from 20 to 200 mg per kg body weight. The data obtained from physical, chemical and biological properties suggest that the EPS may be employed in several industrial and environmental applications. It is an excellent emulsifier, it holds 100 times its own weight in water, it is not toxic, and it can be used to remove mercury, cadmium and lead from aqueous solutions.