Protection of yeast cells in micro-organized shells of natural polyelectrolytes during drying process

The encapsulation techniques are applied in various fields and for various applications. The layer-by-layer self-assembly (LbL), one of the encapsulation techniques, is built by the electrostatic attraction between oppositely charged polyelectrolytes, and the topmost layer determines essentially the sur-face properties of the edifice. This technique offers several advantages (low cost, simplicity of process and equipment, biocompatibility and biodegra-dation, etc.). In this present paper, results of the protection of Saccharomyces cerevisiae yeast cells in microorganized shell of natural polyelectrolytes during dehydration process are reported. To apply the LbL method to individually encaps…

La céramique médiévale et moderne (12e - 17e siècles) en Bourgogne : production, consommation, diffusion. Projet Collectif de Recherches 2018-2021. Bilan 2018-2021 - Programmation 2022-2024

Le programme collectif de recherches intitulé « La céramique médiévale et moderne (12e - 17e siècles) en Bourgogne : production, consommation, diffusion » a entamé ses travaux en 2018, pour une programmation sur quatre années, jusqu’en 2021. Il a pour objectifs de détecter les sites de fabrication potière médiévaux et moderne en Bourgogne, d’en caractériser la production et d’évaluer leur diffusion sur les sites d’habitat.Le nombre de chercheurs associés au programme, rattaché à l’UMR ARTEHIS (Dijon), varie légèrement d’année en année mais tourne en moyenne autour d’une douzaine (11 en 2018, 13 en 2019, 14 en 2020 et 2021). Ils sont issus de diverses institutions (Inrap, Etat, privé, UMR, b…

Influence d'humidité de l'air sur le séchage d'une goutte déposée sur une surface solide et sur la destruction microbienne.

International audience; This study was carried out in order to develop experimental methodology using a camera to monitor the evolution of the surface of a liquid droplet deposited on a solid surface composed of polypropylene. The droplet was exposed to various ambient relative humidity conditions (11.3%, 43.2%, 68.9% and 75.5%). Two types of liquid were investigated: distilled water and water containing nutritive substances (salmon “juice”). At 11.3% relative humidity, it takes 40% longer to evaporate a water droplet (initial weight 0.36 g, volume 360 μL, radius 6.5 × 10−3 m) than a salmon “juice” droplet (3.66 h for distilled water, 2.83 h for salmon “juice”). In the case of the distilled…

Drying parameters greatly affect the destruction of Cronobacter sakazakii and Salmonella Typhimurium in standard buffer and milk

International audience; Salmonella Typhimurium and Cronobacter sakazakii are two foodborne pathogens involved in neonatal infections from milk powder and infant formula. Their ability to survive in low-moisture food and during processing from the decontamination to the dried state is a major issue in food protection. In this work, we studied the effects of the drying process on Salmonella Typhimurium and Cronobacter sakazakii, with the aim of identifying the drying parameters that could promote greater inactivation of these two foodborne pathogens. These two bacteria were dried under different atmospheric relative humidities in milk and phosphate-buffered saline, and the delays in growth re…

Cardiolipin content controls mitochondrial coupling and energetic efficiency in muscle

Decreasing mitochondrial energy-production efficiency in skeletal muscle can confer protection against diet-induced obesity.

A Robust Generic Method for Grid Detection in White Light Microscopy Malassez Blade Images in the Context of Cell Counting

AbstractIn biology, cell counting is a primary measurement and it is usually performed manually using hemocytometers such as Malassez blades. This work is tedious and can be automated using image processing. An algorithm based on Fourier transform filtering and the Hough transform was developed for Malassez blade grid extraction. This facilitates cell segmentation and counting within the grid. For the present work, a set of 137 images with high variability was processed. Grids were accurately detected in 98% of these images.

La céramique médiévale et moderne (12e - 17e siècles) en Bourgogne :production, consommation, diffusion. Projet Collectif de Recherches 2018-2021. Bilan 2020-Programmation 2021

Axe Dynamique de Production matérielle; Le programme collectif de recherches intitulé « La céramique médiévale et moderne (12e - 17e siècles) en Bourgogne : production, consommation, diffusion » a entamé ses travaux en 2018, pour une programmationsur quatre années, jusqu’en 2021. Il a pour objectifs de détecter les sites de fabrication potière médiévaux et moderne en Bourgogne, d’en caractériser la production et d’évaluer leur diffusion sur les sites d’habitat.Le nombre de chercheurs associés au programme, rattaché à l’UMR ARTEHIS (Dijon), varie légèrement d’année en année mais tourne en moyenne autour d’une douzaine (11 en 2018, 13 en 2019, 14 en 2020 et 2021). Ils sont issus de diverses i…

Surviving the heat: heterogeneity of response inSaccharomyces cerevisiaeprovides insight into thermal damage to the membrane

Environmental heat stress impacts on the physiology and viability of microbial cells with concomitant implications for microbial activity and diversity. Previously, it has been demonstrated that gradual heating of Saccharomyces cerevisiae induces a degree of thermal resistance, whereas a heat shock results in a high level of cell death. Here, we show that the impact of exogenous nutrients on acquisition of thermal resistance differs between strains. Using single-cell methods, we demonstrate the extent of heterogeneity of the heat-stress response within populations of yeast cells and the presence of subpopulations that are reversibly damaged by heat stress. Such cells represent potential for…

Château de Saint-Germain (Ambérieu-en-Bugey, Ain). Fouille programmée triennale (2016-2018) – Campagne 2017

Management of Listeria monocytogenes on Surfaces via Relative Air Humidity: Key Role of Cell Envelope

International audience; Although relative air humidity (RH) strongly influences microbial survival, its use for fighting surface pathogens in the food industry has been inadequately considered. We asked whether RH control could destroy Listeria monocytogenes EGDe by envelope damage. The impact of dehydration in phosphate-buffered saline (PBS) at 75%, 68%, 43% and 11% RH on the bacterial envelope was investigated using flow cytometry and atomic force microscopy. Changes after rehydration in the protein secondary structure and peptidoglycan were investigated by infrared spectroscopy. Complementary cultivability measurements were performed by running dehydration–rehydration with combinations o…

Protection of living yeast cells by micro-organized shells of natural polyelectrolytes

International audience; Saccharomyces cerevisiae, a eukaryotic model organism, plays a key role in the oxidative stability of fermented products. In order to protect cells against environmental stresses, we report a method of modifying the cell surface architecture while maintaining the internal working properties of the system. The objective was to encapsulate living yeast cells in micro-organized polyelectrolyte shells using layer-by-layer (LbL) assembly. For the first time, the natural polyelectrolytes, β-lactoglobulin and sodium alginate, were alternately deposited on the surface of S. cerevisiae. Transmission electron microscopy coupled with immune-cytochemistry and scanning electron m…

8.2.3. La vaisselle médiévale et moderne

International audience

Recovery Estimation of Dried Foodborne Pathogens Is Directly Related to Rehydration Kinetics.

International audience; Drying is a common process which is used to preserve food products and technological microorganisms, but which is deleterious for the cells. The aim of this study is to differentiate the effects of drying alone from the effects of the successive and necessary rehydration. Rehydration of dried bacteria is a critical step already studied in starter culture but not for different kinetics and not for pathogens. In the present study, the influence of rehydration kinetics was investigated for three foodborne pathogens involved in neonatal diseases caused by the consumption of rehydrated milk powder: Salmonella enterica subsp. enterica serovar Typhimurium, Salmonella enteri…

Modeling the heat inactivation of foodborne pathogens in milk powder : High relevance of the substrate water activity

International audience; Due to the ability of foodborne pathogens to survive in low moisture foods, the decontamination of these products is an important issue in food hygiene. Up to now, such decontamination has mostly been achieved through empirical methods. The intention of this work is to establish a more rational use of heat treatment cycles. The effects of thermal treatment cycles on the inactivation of dried Salmonella Typhimurium, Salmonella Senftenberg, Cronobacter sakazakii and Escherichia coli were assessed. Bacteria were mixed with whole milk powder and dried down to different water activity levels (0.11, 0.25, 0.44 and 0.58). The rate of inactivated bacteria was determined afte…

Les Cruchades, Combronde (63)

International audience

Caco-2 Invasion by Cronobacter sakazakii and Salmonella enterica Exposed to Drying and Heat Treatments in Dried State in Milk Powder

International audience; Due to the ability of foodborne pathogens to survive in low moisture food, the decontamination of milk powder is an important issue in food protection. The safety of food products is, however, not always insured and the different steps in the processing of food involve physiological and metabolic changes in bacteria. Among these changes, virulence properties may also be affected. In this study, the effect of drying and successive thermal treatments on the invasion capacity of Salmonella Typhimurium, Salmonella Senftenberg, and Cronobacter sakazakii was assessed. Bacteria were dried on milk powder at three different water activity levels (0.25, 0.58, and 0.80) and hea…

Automatic Counting of Intra-Cellular Ribonucleo-Protein Aggregates in Saccharomyces cerevisiae Using a Textural Approach.

AbstractIn the context of microbiology, recent studies show the importance of ribonucleo-protein aggregates (RNPs) for the understanding of mechanisms involved in cell responses to specific environmental conditions. The assembly and disassembly of aggregates is a dynamic process, the characterization of the stage of their evolution can be performed by the evaluation of their number. The aim of this study is to propose a method to automatically determine the count of RNPs. We show that the determination of a precise count is an issue by itself and hence, we propose three textural approaches: a classical point of view using Haralick features, a frequency point of view with generalized Fourier…

Automatic Biological Cell Counting Using a Modified Gradient Hough Transform

AbstractWe present a computational method for pseudo-circular object detection and quantitative characterization in digital images, using the gradient accumulation matrix as a basic tool. This Gradient Accumulation Transform (GAT) was first introduced in 1992 by Kierkegaard and recently used by Kaytanli & Valentine. In the present article, we modify the approach by using the phase coding studied by Cicconet, and by adding a “local contributor list” (LCL) as well as a “used contributor matrix” (UCM), which allow for accurate peak detection and exploitation. These changes help make the GAT algorithm a robust and precise method to automatically detect pseudo-circular objects in a microscop…

La céramique médiévale et moderne (12e-17e siècle) en Bourgogne : production, consommation, diffusion. Projet Collectif de Recherches 2018-2021

Estimation of Microbial Viability Using Flow Cytometry.

For microorganisms in particular, viability is a term that is difficult to define and a state consequently difficult to measure. The traditional (and gold standard) usage equates viability and culturability (i.e., the ability to multiply) but the process of determining culturability is often too slow. Flow cytometry provides the opportunity to make rapid and quantitative measurements of dye uptake in large numbers of cells and we can therefore exploit the flow cytometric approach to evaluate so-called viability stains and to develop protocols for more routine assessments of microbial viability. This article provides a commentary and several protocols have been included to ensure that users …

An Off-On-Off Fluorescent Sensor for pH Windows Based on the 13aneN4-Zn 2+ System

International audience; The new ligand L was prepared and features a 13-membered tetraaza macrocyclic ring with a 1,8-naphthalimide fluorophore appended to a C atom of its backbone. The protonation constants of L as well as its complexation constants with Zn2+ ions were determined in 1:1 water/methanol solutions by potentiometric titrations. Fluorimetric pH titrations were performed with L alone and L in the presence of Zn2+ ions (1:1), and the species distributions (%) versus pH were compared. A window-shaped fluorescence trend was observed with pH for the L/Zn2+ system, which behaves as an off-on-off pH sensor. The on window is centred in the 6.5-7.5 pH range, in correspondence with the f…

Directed evolution of oenococcus oeni to improve acid-tolerance reveals fixed beneficial mutations in the citrate locus

Cellular Injuries in Cronobacter sakazakii CIP 103183T and Salmonella enterica Exposed to Drying and Subsequent Heat Treatment in Milk Powder

International audience; Because of the ability of foodborne pathogens to survive in low-moisture foods, their decontamination is an important issue in food protection. This study aimed to clarify some of the cellular mechanisms involved in inactivation of foodborne pathogens after drying and subsequent heating. Individual strains of Salmonella Typhimurium, Salmonella Senftenberg, and Cronobacter sakazakii were mixed into whole milk powder and dried to different water activity levels (0.25 and 0.58); the number of surviving cells was determined after drying and subsequent thermal treatments in closed vessels at 90 and 100 degrees C, for 30 and 120 s. For each condition, the percentage of unc…

The Impact of Drying on Foodborne Pathogens Salmonella enterica and Cronobacter sakazakii

International audience; [Technical Session 1 – Intervention Strategies] Introduction: Salmonella enterica and Cronobacter sakazakii are foodborne pathogens responsible for severe infant illness. Their ability to survive in harsh environmental conditions make these species a matter of concern for the low moisture food industry. Purpose: This study aimed to evaluate and understand the impact of drying conditions on survival, physiology, and invasion capacity of Salmonella Typhimurium and C. sakazakii.Methods: Salmonella Typhimurium and C. sakazakii were mixed into whole milk powder and dried at different water activity (aw) levels (0.25, 0.58 and 0.80). For each strain, the impact of each dry…

Hydrodynamic characterization of a new small-scale reactor mixed by a magnetic bar

International audience; The purpose of this study was to evaluate the mixing characteristics of a new milliliter-scale (mL-scale) reactor developed for studying enzymatic activity or physiological cell response. The mL-scale reactor was designed to enable the integration of several sensors to carry out dynamic measurements in a controlled environment. Rapid homogeneity of the entire system is essential to ensure reproducible and reliable results, consequently the reactor was stirred to optimize both mass and heat transfers. A comparative study using three different techniques was undertaken to study mixing performances in the system. Firstly, mixing time (4,) was estimated in the reactor us…

Exploration of lipid metabolism in relation with plasma membrane properties of Duchenne muscular dystrophy cells: influence of L-carnitine.

Duchenne muscular dystrophy (DMD) arises as a consequence of mutations in the dystrophin gene. Dystrophin is a membrane-spanning protein that connects the cytoskeleton and the basal lamina. The most distinctive features of DMD are a progressive muscular dystrophy, a myofiber degeneration with fibrosis and metabolic alterations such as fatty infiltration, however, little is known on lipid metabolism changes arising in Duchenne patient cells. Our goal was to identify metabolic changes occurring in Duchenne patient cells especially in terms of L-carnitine homeostasis, fatty acid metabolism both at the mitochondrial and peroxisomal level and the consequences on the membrane structure and functi…

HACD1, a regulator of membrane composition and fluidity, promotes myoblast fusion and skeletal muscle growth

International audience; The reduced diameter of skeletal myofibres is a hallmark of several congenital myopathies, yet the underlying cellular and molecular mechanisms remain elusive. In this study, we investigate the role of HACD1/PTPLA, which is involved in the elongation of the very long chain fatty acids, in muscle fibre formation. In humans and dogs, HACD1 deficiency leads to a congenital myopathy with fibre size disproportion associated with a generalized muscle weakness. Through analysis of HACD1-deficient Labradors, Hacd1-knockout mice, and Hacd1-deficient myoblasts, we provide evidence that HACD1 promotes myoblast fusion during muscle development and regeneration. We further demons…

High gas pressure survival/reactivation of soil microorganisms

International audience; Deep sea sediments constitute a considerable reserve of microorganisms belonging to different microbial communities. Our researches aimed to better understand cellular mechanisms related to cellular plasticity involved in resistance of such microbial communities to extreme conditions and more particularly to high level of pressure (> 50 MPa). Obviously, the first step is to isolate microorganisms present in deep sea sediments and then cultivate. The comparison of the cultivation of such microorganisms under atmospheric conditions and under pressure conditions will afford a possible reactivation of specific piezotolerants and/or piezophiles organisms from dormancy. Th…

Château de Saint-Germain (Ambérieu-en-Bugey, Ain). Rapport final d'opération 2016-2018

Improving acid tolerance in Oenococcus oeni by Adaptive Evolution

International audience; Oenococcus oeni is a lactic acid bacterium (LAB) mainly responsible for the malolactic fermentation (MLF) inwine. MLF plays an important role in determining the final quality of wines[1]. Even though this LAB is naturally present in musts, wines and oenological environment, spontaneous MLF are usually unpredictable because of the stressful conditions and especially due to acidity [2]. The consequence of the mismanagement ofthis step might lead to the depreciation of wine quality. To obtain a clone more tolerant to acidity, we undertaken to replicate O. oeni in a temporally varying environment (pH 5.3 to 3.2) to improve acid tolerance. To discriminate stress tolerance…

Adaptive evolution to improve acid tolerance in Oenococcus oeni

International audience; Oenococcus oeni is a lactic acid bacterium (LAB) mainly responsible for the malolactic fermentation (MLF) inwine. MLF plays an important role in determining the final quality of wines [1]. Even though this LAB is naturallypresent in musts, wines and oenological environment, spontaneous MLF are usually unpredictable because of thestressful conditions and especially due to acidity [2]. The consequence of the mismanagement of this step might leadto the depreciation of wine quality. To obtain a clone more tolerant to acidity, we undertook a replication of O. oeni until 450 generations in a temporally varying environment (pH 5.3 to 2.9) to improve acid tolerance. To discr…

Reliable Detection and Smart Deletion of Malassez Counting Chamber Grid in Microscopic White Light Images for Microbiological Applications

AbstractIn biology, hemocytometers such as Malassez slides are widely used and are effective tools for counting cells manually. In a previous work, a robust algorithm was developed for grid extraction in Malassez slide images. This algorithm was evaluated on a set of 135 images and grids were accurately detected in most cases, but there remained failures for the most difficult images. In this work, we present an optimization of this algorithm that allows for 100% grid detection and a 25% improvement in grid positioning accuracy. These improvements make the algorithm fully reliable for grid detection. This optimization also allows complete erasing of the grid without altering the cells, whic…

Phospholipidosis and down-regulation of the PI3-K/PDK-1/Akt signalling pathway are vitamin E inhibitable events associated with 7-ketocholesterol-induced apoptosis

International audience; Among the oxysterols accumulating in atherosclerotic plaque, 7-ketocholesterol (7KC) is a potent apoptotic inducer, which favours myelin figure formation and polar lipid accumulation. This investigation performed on U937 cells consisted in characterizing the myelin figure formation process; determining the effects of 7KC on the PI3-K/PDK-1/Akt signalling pathway; evaluating the activities of vitamin E (Vit-E) (α-tocopherol) on the formation of myelin figures and the PI3-K/PDK-1/Akt signalling pathway and assessing the effects of PI3-K inhibitors (LY-294002, 3-methyladenine) on the activity of Vit-E on cell death and polar lipid accumulation. The ultrastructural and b…

Survie et/ou réactivation de microorganismes du sol sous hautes pressions gazeuses

National audience; Le sol constitue une réserve considérable de microorganismes représentatifs de nombreuses communautés cellulaires. Les recherches reposent sur l’hypothèse qu’il existe, dans les échantillons de sol, des communautés microbiennes adaptées à des conditions physiques (pression, température) très différentes de celles rencontrées dans nos laboratoires : soit parce que les conditions de milieu ont changé (sol), soit parce que les conditions de prélèvement sont très différentes des conditions de culture (grands fonds marins). Dans cette optique, l’utilisation des hautes pressions gazeuses (200-1000 bars dans un premier temps) est envisagée afin d’essayer de réactiver des microor…

Extremely rapid acclimation of Escherichia coli to high temperature over a few generations of a fed-batch culture during slow warming

This study aimed to demonstrate that adequate slow heating rate allows two strains of Escherichia coli rapid acclimation to higher temperature than upper growth and survival limits known to be strain-dependent. A laboratory (K12-TG1) and an environmental (DPD3084) strain of E. coli were subjected to rapid (few seconds) or slow warming (1 degrees C 12 h(-1)) in order to (re) evaluate upper survival and growth limits. The slow warming was applied from the ancestral temperature 37 degrees C to total cell death 46-54 degrees C: about 30 generations were propagated. Upper survival and growth limits for rapid warming (46 degrees C) were lower than for slow warming (46-54 degrees C). The thermal l…

Design of new sensitive α,β-unsaturated carbonyl 1,8-naphtalimide fluorescent probes for thiol bioimaging

International audience; We report herein an efficient synthesis of news naphtalimide-based fluorescent derivatives functionalized with an a,(3-unsaturated carbonyl group. Probes were synthesized from reaction of an amino-naphtalimide precursor with maleic anhydride. Photophysical study of fluorescent probes and cells labelling were performed, and showed that the reactive derivatives exhibit rich turn on fluorescence properties in presence of different biological thiol (Glutathione and Cysteine) making these systems a very promising way for thiol bioimaging.