Modification de biopolymère pour l'étude des interactions bactéries - surface abiotique

Les travaux présentés dans ce mémoire ont pour objectif de concevoir un revêtement polymère favorisant l'adhésion et le développement d'un biofilm mature en vue de son utilisation dans des bioréacteurs afin d'en conserver l'efficacité. Ces revêtements ont été réalisés en greffant différentes fonctions chimiques par polymérisation de monomères vinyliques (HEMA, AAc, DMAEMA) à la surface de biopolyester comme le poly(hydroxybutyrate-cohydroxyvalérate) (PHBHV). Les perfonnances de différentes voies de greffage ont été analysées en tennes d'adhésion et de développement du biofilm bactérien. La modification chimique du PHBHV a été effectuée en greffant différents monomères hydrophiles par plusieurs voies de synthèses radicalaires. Les radicaux ont été générés par voie thennique ou par voie photochimique en présence d'un amorceur afin d'initier le greffage. Une nouvelle approche de modification de PHA a également été étudiée par le biais d'un amorceur immobilisé à la surface du film de PHBHV et la polymérisation a été initiée par transfert d'atome (SI-ATRP). La caractérisation de ces films de PHBHV greffés a montré une modification physicochimique des surfaces. Par ailleurs, la microscopie électronique à balayage (MEB) couplée à l'analyse X (EDX) a montré une distribution des greffons différente au sein du film en fonction de la technique de greffage étudiée. L'étude de l'adhésion et du développement du biofilm bactérien a ét réalisée sur les surfaces greffées ayant un taux de greffage relativement faible afin de conserver les propriétés biodégradables du PHBHV. Les observations par microscopie confocale montrent l'influence du greffage.The goal ofthis present work was to develop a polymer promoting the adhesion and the development ofa mature biofilm. The main application was intend to bioreactor and to maintain their efficiency. These surfaces were made by grafting various chemical functions by polymerizing different vinyl monomers (HEMA, AAC, DMAEMA) on the surface ofa biopolyester the poly(hydroxybutyrate-co-hydroxyvalerate) (PHBHV). Bacterial adhesion and biofilm development were tested on these different surfaces. Three routes of chemical radical grafting were carried out to immobilize the hydrophilic functions. Radical grafting were experimented by thennal initiation, photo initiation and a new approach by control radical polymerization the SI-A TRP. The grafted films were characterized by a large range of surface analysis such as scanning electon microscopy (SEM) associated with energy dispersive X-ray analysis (EDX), contact angle measurement, A TR-FTIR. These characterization techniques showed that depending on the grafting route the grafted chains were localized in different part of the PHBHV film. Evaluation of the bacterial adhesion and the biofilm fonnation were realized oflow graft yield to keep the biodegradation property of the PHBHV. The bacteria-surface study was followed by con focal laser scannin microscopy (CLSM).LORIENT-BU (561212106) / SudocSudocFranceF

Aptitude de Salmonella à contaminer l'oeuf coquille et élaboration d'un revêtement antibactérien

Salmonella enterica est une des espèces bactériennes les plus impliquées dans les toxi-infections alimentaires collectives (Tiac). Cette bactérie ubiquitaire, provoquant des gastroentérites, est capable de contaminer un large panel d aliments. Cependant, l apparition de Tiac est majoritairement provoquée par la consommation de volaille et d œufs. Dans ce contexte, l objectif de ce travail a été premièrement de caractériser quatre souches bactériennes du genre Salmonella, trois sérovar Enteritidis et un sérovar Typhimurium, et d évaluer leur aptitude à pénétrer dans l œuf coquille par contamination horizontale. Dans un second temps, l objectif s est porté sur la réalisation d un film alimentaire aux propriétés antibactériennes. Au cours de la caractérisation des quatre souches de Salmonella, leur morphologie et leur croissance à différentes températures ont été observées. La persistance de Salmonella dans l environnement a ensuite été analysée par l étude du morphotype rdar et l aptitude des quatre souches à former des biofilms. Des tests de mobilité bactérienne, swimming , twitching et swarming , ont également été effectués, ainsi que l extraction et la caractérisation des lipopolysaccharides. Enfin, l étude de la pénétration de ces quatre souches a été réalisée par la méthode agar-moulding afin de corréler les différents comportements et facteurs évoqués précédemment avec l aptitude de Salmonella à contaminer et à pénétrer dans l œuf coquille. La deuxième partie de ce travail concerne la réalisation d un film alimentaire aux propriétés antibactériennes vis-à-vis de Salmonella Enteritidis. Parmi trois biopolymères, le chitosan a été sélectionné et a été réticulé avec du tripolyphosphate pour améliorer sa stabilité. Une attention particulière s est ensuite portée sur l activité antibactérienne de la photocatalyse du dioxyde de titane afin d incorporer ce dernier au film de chitosan. Finalement, le film de chitosan seul, a été appliqué sur une population d œufs, qui a ensuite été contaminée dans le but d évaluer l impact du film sur la pénétration de Salmonella Enteritidis dans l œuf coquille.Salmonella enterica is one of the most common species responsible for foodborne disease throughout the world. This ubiquitous bacteria, inducing gastroenteritis, can contaminate a wide range of foods. Foodborne gastroenteritis is usually due to the consumption of poultry and eggs. In this context, the aim of this work was, first, to characterize four Salmonella strains and to evaluate the ability of the strains to penetrate eggs through the eggshell and secondly, to develop an antibacterial food coating. Salmonella characterization consists in studying bacterial morphology and growth at several temperatures. Next, Salmonella persistence in environment was evaluated by studying the rdar morphotype and the biofilm formation using confocal laser scanning microscopy. Bacterial motility was also observed: swimming , swarming , and twitching tests were performed. Finally, lipopolysaccharides were extracted and characterized by SDS-PAGE electrophoresis. All these experiments were carried out on four Salmonella enterica strains to determine if these behaviours and factors influenced the penetration of Salmonella in the egg content through the eggshell. Thus in conclusion, the ability of the four strains to penetrate through the eggshell was studied performing the agar-moulding method. The second part of the work related the elaboration of a food coating providing an antibacterial activity against Salmonella Enteritidis. Among three biopolymers, chitosan was chosen and was reticulated with tripolyphosphate to increase it stability. The antibacterial activity of the photocatalysis of titanium dioxide was also studied. The aim was to add titanium dioxide in the chitosan coating. It was then applied on eggs which were then contaminated with Salmonella Enteritidis to evaluate the impact of the chitosan coating on bacterial penetration in egg content.LORIENT-BU (561212106) / SudocSudocFranceF

Evaluation of anti-microfouling activity of marine paints by microscopical techniques

International audienc

Additives for Efficient Biodegradable Antifouling Paints

The evolution of regulations concerning biocidal products aims to increase protection of the environment (e.g., EU Regulation No 528/2012) and requires the development of new non-toxic anti-fouling (AF) systems. The development of these formulations implies the use of ingredients (polymers, active substances, additives) that are devoid of toxicity towards marine environments. In this context, the use of erodable antifouling paints based on biodegradable polymer and authorized biocides responds to this problem. However, the efficiency of paints could be improved by the use of specific additives. For this purpose, three additives acting as surface modifiers were studied (Tween 80, Span 85 and PEG-silane). Their effects on parameters involved in antifouling efficiency as hydrophobicity, hydration and copper release were studied. Results showed that the addition of 3% of additives modulated hydrophobicity and hydration without an increase of copper release and significantly reduced microfouling development. Efficient paints based on biodegradable polymer and with no organic biocide could be obtained by mixing copper thiocyanate and additives

Evaluation of anti-microfouling activity of marine paints by microscopical techniques

International audienc

Evaluation of antibacterial activity against Salmonella Enteritidis.

International audienceSalmonella enterica serovar Enteritidis is a well-known pathogenic bacterium responsible for human gastrointestinal enteritis mainly due to the consumption of eggs and egg-products. The first aim of this work was to study several virulence factors of a strain isolated from egg content: SEovo. First, bacterial growth was studied at several temperatures and cell morphology was observed by scanning electronic microscopy. These experiments showed Salmonella's ability to grow at low temperatures and to produce exoproducts. Next, Salmonella motility was observed performing swimming, twitching, and swarming tests. Results indicated a positive flagellar activity and the cell ability to differentiate and become hyperflagellated under specific conditions. Moreover, SEovo adherence and biofilm formation was carried out. All of these tests enabled us to conclude that SEovo is a potential pathogen, thus it can be used as a model to perform antibacterial experiments. The second part of the study was dedicated to the evaluation of the antibacterial activity of different molecules using several methods. The antibacterial effect of silver and copper aluminosilicates was tested by two different kinds of methods. On the one hand, the effect of these two antibacterial agents was determined using microbiological methods: viable cell count and agar-well diffusion. And on the other hand, the antibacterial activity was evaluated using CLSM and SYTO Red/SYTOX Green dyeing. CLSM allowed for the evaluation of the biocide on sessile cells, whereas the first methods did not. Results showed that adhered bacteria were more resistant than planktonic counterparts and that CLSM was a good alternative to evaluate antibacterial activity on fixed bacteria without having to carry out a removing step