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Dans le domaine du diagnostic in vitro, l'étape d'extraction de micro-organismes à partir d'un échantillon complexe est une étape clé pour permettre l'identification du pathogène responsable d'une infection. Pour les septicémies, cette étape d'extraction est généralement précédée d'une étape de culture, ce qui conduit à une obtention des résultats au bout de plusieurs jours. Un résultat plus rapide (typiquement inférieur à 24h) permettrait d'augmenter le taux de survie des patients, et aurait ainsi une forte valeur ajoutée pour le corps médical. Le but de ces travaux est donc de développer une nouvelle méthode d'extraction et de concentration de pathogènes directement à partir d'un échantillon sanguin, sans étape de culture. Une stratégie en deux modules microfluidiques associés en série est proposée : elle repose sur la modification de la conductivité et de l'osmolarité de l'échantillon dans un premier module, puis sur la capture des micro-organismes par diélectrophorèse dans un second module. L'étude du premier module a permis de déterminer l'impact de la conductivité et de l'osmolarité du milieu sur les propriétés diélectriques des cellules. Deux voies ont ainsi été abordées, afin de diriger les cellules du sang et les micro-organismes vers un milieu de conductivité et d'osmolarité contrôlées : la dilution, et l'utilisation de forces acoustiques. L'étude du deuxième module a ensuite permis de démontrer la possibilité de capturer et concentrer des micro-organismes à partir d'un échantillon hypotonique et faiblement conducteur dans un écoulement microfluidique par diélectrophorèse. L'architecture d'un microsystème dédié a été définie grâce à un modèle numérique, puis validé expérimentalement avec des échantillons sanguins et différents micro-organismes (E. coli, S. epidermidis et C. albicans). La capture générique des micro-organismes est démontrée, et un taux de capture de 97% a été obtenu pour la séparation de \EC, avec une vitesse moyenne de l'échantillon dans le microsystème de 100 à 200 µm.s-1. Enfin, des perspectives d'amélioration sont présentées pour permettre d'effectuer cette étape de séparation sur un gros volume d'échantillon (1 à 10mL) en quelques heures, afin de répondre aux exigences imposées par l'urgence des tests de diagnostic des septicémies. / Extraction of pathogens from a biological sample is a key step for efficient diagnostic tests of infectious diseases. For bloodstream infections, current diagnostic methods are usually based on bacterial growth and take several days to provide valuable information. An accelerated result would have a high medical value to adjust therapeutic strategies. The aim of this study is to design a new approach for separation and concentration of microorganisms directly from a blood sample, to avoid time-consuming growth stages. We report a method based on two different microsystems connected in series: it combines modification of conductivity and osmolarity of the sample with generic capture of microorganisms by dielectrophoresis. First we explore the impact of conductivity and osmolarity on the dielectric properties of blood cells and microorganisms. Dilution and acoustic forces are both analyzed to transfer blood cells and microorganisms to the optimized buffer. Then we demonstrate the feasibility of achieving the dielectrophoretic separation of microorganisms from blood cells in a low conductivity and low osmolarity medium inside a fluidic device. The structure of the device is optimized with numerical simulations and experiments performed on blood samples and various microorganisms (E. coli, S. epidermidis and C. albicans).The generic capture of microorganisms is validated, and we achieved a separation of 97% efficiency with E. coli, with an optimal inlet velocity around 100-200 µm.s-1. Finally, we propose an improved microsystem to perform the sample preparation step on a larger volume (1-10mL) in a few hours, in order to fit the medical need.

Le football moderne est caractérisé par des efforts intermittents de très haute intensité. Pendant un match, les joueurs réalisent des performances, qu'elles soient physiques ou techniques, en lien direct avec la spécificité de leur poste de jeu, leur rôle tactique et leur positionnement sur le terrain. Un match de football de haut niveau induit des variations de fréquence cardiaque, une baisse de réserves énergétiques, une augmentation des dommages musculaires, du stress oxydatif et une affectation du statut immunitaire. Incidences physiologiques auxquelles se rajoutent des modifications de perception de la fatigue, des douleurs musculaires, du bien-être, de la qualité du sommeil, du stress psychologique et de l'humeur. Toutes ces incidences se mesurent, se quantifient et s'analysent en lien direct avec des facteurs contextuels comme le lieu du match, le moment de la journée, le système de jeu, …, et les périodes d'enchainement de match (e.g. deux à trois matchs par semaine) qui peuvent avoir une influence significative. La présente thèse a pour objectif principal l'étude de l'influence de l'enchainement de matchs sur les performances physiques et sur les cinétiques de récupération mesurées sur des marqueurs sanguins, salivaires et des questionnaires de perception, sur des joueurs de football de haut-niveau / Modern football is characterized by very high-intensity intermittent efforts. During a match, players perform technical and physical tasks in relation to their specific positions on the field. A high-level football match induces heart-rate variations, energetic storage lowering, muscular damage and oxidative stress increase and immune status alteration. These physiological variations are accompanied by modifications of fatigue perceived, muscle soreness, wellness, sleep quality, psychological stress and overall mood. All these incidences can be measured, quantified and analyzed, in direct relation to contextual factors like game location, time of the day, playing system, …, and congested period of matches (e.g. two to three matches per week). The present thesis aims to report all the ways to monitor match load and fatigue and aims to analyze the influence of playing matches during congested periods on physical activity and on physiological post-match kinetics, over high-level football players

Inom klinisk verksamhet är det av stor betydelse att känna till hållbarheten på olika analyter för att säkerställa att analysen inte genomförs på för gamla prover. Vid litteratursökning framgick att bland de publicerade studier som undersökt hållbarheten på blodprover har de flesta undersökt hållbarheten på venösa men inte kapillära blodprover. En vanligt förekommande analys med både venösa och kapillära prover är blodstatus, som inkluderar bland annat bestämning av hemoglobinkoncentrationen (Hb-), leukocyt- samt trombocytpartikelkoncentrationen (LPK och TPK). Analys av blodets celler kan ge allmän information om hälsotillståndet hos patienter och är därav viktiga och vanligt förekommande analyser. Syftet med examensarbetet var att undersöka hållbarheten för analyserna Hb, LPK och TPK på kapillära blodprover efter fyra och sex timmars förvaring i rumstemperatur. Provmaterialet utgjordes av 100 kapillära prover som analyserades på hematologiinstrumentet Sysmex XN-10. Analysmetoden för Hb var fotometri med natriumlaurylsulfat- (SLS-) metoden. LPK bestämdes med flödescytometri medan TPK bestämdes med impedansmetoden alternativt flödescytometri. Resultatet visade att det för Hb inte förekom någon signifikant statistisk skillnad mellan den initiala analysen och efter fyra respektive sex timmar. Analysresultatet för Hb var därmed stabilt under förhållandena i denna studie. För LPK och TPK förekom statistiskt signifikanta skillnader mellan resultatet vid den initiala analysen jämfört med efter fyra respektive sex timmar. Analysresultaten för LPK och TPK var därmed inte lika stabila som Hb är under förhållandena i studien. Den statistiska skillnaden bedömdes däremot inte ha någon klinisk betydelse, vilket ledde till slutsatsen är att kapillära prover för analys av Hb, LPK och TPK är hållbara i sex timmar vid förvaring i rumstemperatur. / One important factor in clinical practice is understanding the stability of analytes and for how long blood samples can be stored before analysis. Most published studies are based on venous blood samples rather than capillary. The knowledge about storage time for cells in capillary blood is therefore limited. Plenty of information can be obtained by analysing the blood cells and its components, including haemoglobin, white blood cells and platelets. These analyses are therefore some of the most common in clinical practice. The aim of this study was to evaluate the effect of the analysis result for haemoglobin, white blood cells and platelets in capillary blood samples after storage at room temperature for four and six hours, respectively. In the study, 100 capillary samples from anonymous patients were analysed with the haematology analyser Sysmex XN-10. The method for analysing haemoglobin was the sodium lauryl sulphate (SLS) detection method with spectrophotometry. White blood cells were analysed with fluorescens flow cytometry. Platelets were analysed with either impedance or fluorescens flow cytometry. The result of the study showed no statistically significant difference between the initial analysis results in haemoglobin and at four and six hours, concluding that within the conditions of this study, the analysis result for haemoglobin was stable. Significant statical differences were identified for both white blood cells and platelet analysis results at four and six hours. However, the identified statistical significance was not esteemed to have any clinical relevance. In conclusion haemoglobin, white blood cells and platelets in capillary blood can be stored for at least six hours at room temperature.

Diabetes mellitus is a frequent endocrinopathy in dogs. Exogenous insulin and nutritional management are the mainstays of therapy. High carbohydrate and fibre diets have been traditionally recommended for canine diabetes mellitus. However, recent studies in healthy and diabetic dogs have challenged the use of these traditional diabetic diets. In addition, dietary carbohydrate content was found to be the main determinant of postprandial blood glucose concentrations over 1.5 hours in healthy dogs. Increasing carbohydrate content increased blood glucose concentration. In a search of the literature, no studies in dogs were found comparing the effects on postprandial plasma glucose, insulin, triglyceride and free fatty acid concentrations of a traditionally recommended high carbohydrate and fibre diabetes diet with a moderate carbohydrate and fibre diet, or a commercially-available maintenance diet. Although fasting plasma triglyceride concentrations are commonly used for diagnosis and management of canine hyperlipidemia, a review of the literature found that in human beings, it may not to be predictive of the highest postprandial concentration. Importantly, in overweight and obese dogs with postprandial plasma triglyceride concentrations >5 mmol/L (>445 mg/dL), it was found there was a 6 fold increase in risk of laboratory evidence of exocrine pancreatic disease. However, there are limited studies on the effect of maintenance diets on postprandial plasma triglyceride concentrations in dogs, and no reference intervals determined for a triglyceride meal challenge test in healthy dogs. For the dietary studies in this thesis, animal welfare was of paramount importance, and minimally invasive peripheral venous catheterization and blood collection techniques were needed to collect 4 mL blood samples over 13 hours in conscious dogs. A search of the literature found no studies evaluating the reliability of using peripheral catheterization via the cephalic vein for the collection of larger blood volumes (for example, 4 mL). The aims of this thesis were firstly, to investigate the influence of three diets with varying carbohydrate and fibre content on postprandial plasma glucose, insulin, triglyceride and free fatty acid concentrations in healthy dogs over 12 hours; secondly, to develop a triglyceride meal challenge test and associated reference intervals; and finally, to evaluate a minimally invasive catheterization and blood collection technique for reliability in collecting the blood volumes required for these studies. For the studies in this thesis, twelve healthy dogs were fed each of three diets for three weeks in a three period cross-over design. For the meal response test at the end of each three-week period, 10 blood samples were collected over 13 hours. Reference intervals for fasting and postprandial triglyceride concentrations at single and multiple time points after eating a standard meal were determined in healthy dogs. Associations between fasting and postprandial triglyceride concentrations, and time to measure highest postprandial triglyceride concentration were also evaluated. Blood volume obtained, resistance on aspirating the blood sample, and methods used to improve blood flow during collection were recorded for the 358 samples collected. The results of the studies in this thesis demonstrated that a moderate carbohydrate diet resulted in lower peak and postprandial glucose concentrations compared with a traditional diabetic diet (CHO 55 %ME) and a maintenance diet (CHO 45 %ME). Fasting triglyceride concentrations were found not to be predictive of peak postprandial triglyceride concentrations. The highest triglyceride concentration measured at 2, 5, and 6 hours after eating had the closest agreement with peak postprandial triglyceride concentration. Glucose, insulin and triglyceride concentrations in some dogs were significantly above baseline concentrations at 12 hours after eating each of the diets. Of 358 blood samples collected, 93 % achieved the required 4 mL volume, while the remaining samples were greater than 1.5 mL, and 87 % were obtained with minimal resistance. It was concluded that the moderate carbohydrate and fibre test diet warrants evaluation in diabetic dogs. It was found that when following the same protocol, blood samples should be collected 2, 5, and 6 hours after eating to determine the highest postprandial plasma triglyceride concentration. For future nutritional studies, dogs may need to be fasted for at least 12 hours, and analytes measured over at least 12 hours after feeding to evaluate fasting and postprandial plasma analyte concentrations. Finally it was demonstrated that percutaneous catheterization of the cephalic vein using a 20-gauge catheter allows for successful collection of multiple 4 mL blood samples over 13 hours in conscious dogs. This thesis presents new information for future nutritional studies in healthy and diabetic dogs by suggesting that there may be no glycemic benefit feeding the high carbohydrate and fibre diet compared with a moderate carbohydrate and fibre diet. A triglyceride meal challenge test was developed to assist the diagnosis of canine hyperlipidemia. Finally, a minimally invasive method for obtaining serial blood samples was evaluated and described. These findings are expected to help in designing future studies in the nutritional management in healthy dogs, and dogs with diabetes mellitus and hyperlipidemia.