Growth of the extremophilic Deinococcus geothermalis DSM 11302 using co-substrate fed-batch culture

Deinococcus geothermalis metabolism has been scarcely studied to date, although new developments on its utilization for bioremediation have been carried out. So, largescale production of this strain and a better understanding of its physiology are required. A fed-batch experiment was conducted to achieve a high cell density non-limiting culture of D. geothermalis DSM 11302. A co-substrate nutritional strategy using glucose and yeast extract was carried out in a 20-L bioreactor in order to maintain a non-limited growth at a maximal growth rate of 1 h−1 at 45 °C. Substrate supplies were adjusted by monitoring online culture parameters and physiological data (dissolved oxygen, gas analyses, respiratory quotient, biomass concentration). The results showed that yeast extract could serve as both carbon and nitrogen sources, although glucose and ammonia were consumed too. Yeast extract carbon-specific uptake rate reached a value 4.5 times higher than glucose carbon-specific uptake rate. Cell concentration of 9.6 g L−1 dry cell weight corresponding to 99 g of biomass was obtained using glucose and yeast extract as carbon and nitrogen sources

Dynamique de la réponse physiologique d'Escherichia coli à des perturbations maîtrisées de son environnement (vers le développement de nouveaux outils de changement d'échelle)

Les bioréacteurs de grandes dimensions, en raison de phénomènes de transfert limitant, sont le siège d hétérogénéités se traduisant par des gradients locaux de concentration et température. Les microorganismes circulant au sein de ces bioréacteurs subissent donc des fluctuations environnementales qui peuvent affecter leur comportement aux niveaux métaboliques et/ou moléculaires. La réponse microbienne est fonction de la nature, de l intensité, de la fréquence et de la durée de la perturbation. L objectif de ce travail est l étude quantitative de l impact de l intensité, la fréquence et l amplitude d un stress nutritionnel sur le comportement dynamique d Escherichia coli, à savoir des ajouts pulsés de glucose lors de cultures continues en régime permanent. Un effort particulier est consacré au développement et à la validation des outils expérimentaux indispensables pour une caractérisation rigoureuse des dynamiques de réponses transitoires sur des échelles de temps allant de secondes à quelques minutes. Pour permettre le suivi in situ et en temps réel des changements métaboliques et moléculaires, une souche bioluminescente est mise en œuvre. Les réponses transitoires sont caractérisées par les vitesses spécifiques, les rendements, les profils d induction transcriptionnelle, les temps caractéristiques. Selon les différents scenarii réalisés, l ajustement du métabolisme face aux hétérogénéités de substrat est quantifié selon des échelles de temps aux niveaux macroscopiques et/ou moléculaires ; ces résultats originaux contribuent ainsi à l implémentation des connaissances sur les interactions dynamiques entre les phénomènes biologiques et les phénomènes physiques ; l enjeu réside à terme en l amélioration des processus d optimisation et d extrapolation des bioprocédés par l identification et la quantification des dynamiques des phénomènes limitantsIneffective mixing entailing heterogeneity issues within industrial bioreactors have been reported to affect microbial metabolisms at cellular and/or molecular levels. Substrate gradients inside large-scale bioreactors are common environmental fluctuations that microorganisms would have to encouter along with the bioprocess. Depending on intensity, frequency and duration of those fluctuations, microorganisms may respond in a different manner. The objective of this work is to study the impact of intensity, frequency and amplitude of glucose perturbations on the dynamics of Escherichia coli responses. An E. coli bioluminescent strain is used for in situ and real-time monitoring of both metabolic and transcriptional changes. For this purpose, short-term glucose excess was simulated, using pulse-based experiments into glucose-limited chemostat cultures. In addition, an important effort is devoted to the development and validation of technical and mathematical tools in order to acquire quantitative and kinetic data on time scales from seconds to minutes. The transient responses are characterized, using specific rates, yields, transcriptional induction profiles and characteristic response times, and are compared in the different defined perturbation scenarios. The results reflected the fact that short-term heterogeneities of substrate affect both cell metabolism and regulation at macroscopic and/or molecular levels. Quantitative understandings of the dynamics during transient responses to environmental perturbations can thus shed light on the bioprocess optimizationTOULOUSE-INSA-Bib. electronique (315559905) / SudocSudocFranceF

Quantitative Characterization of the Growth of Deinococcus geothermalis DSM-11302: Effect of Inoculum Size, Growth Medium and Culture Conditions

Due to their remarkable resistance to extreme conditions, Deinococcaceae strains are of great interest to biotechnological prospects. However, the physiology of the extremophile strain Deinococcus geothermalis has scarcely been studied and is not well understood. The physiological behaviour was then studied in well-controlled conditions in flask and bioreactor cultures. The growth of D. geothermalis type strains was compared. Among the strains tested, the strain from the German Collection of Microorganisms (Deutsche Sammlung von Mikroorganismen DSM) DSM-11302 was found to give the highest biomass concentration and growth rate: in a complex medium with glucose, the growth rate reached 0.75 h−1 at 45 °C. Yeast extract concentration in the medium had significant constitutive and catalytic effects. Furthermore, the results showed that the physiological descriptors were not affected by the inoculum preparation steps. A batch culture of D. geothermalis DSM-11302 on defined medium was carried out: cells grew exponentially with a maximal growth rate of 0.28 h−1 and D. geothermalis DSM-11302 biomass reached 1.4 g•L −1 in 20 h. Then, 1.4 gDryCellWeight of biomass (X) was obtained from 5.6 g glucose (Glc) consumed as carbon source, corresponding to a yield of 0.3 CmolX•CmolGlc−1 ; cell specific oxygen uptake and carbon dioxide production rates reached 216 and 226 mmol.CmolX −1•h −1 , respectively, and the respiratory quotient (QR) value varied from 1.1 to 1.7. This is the first time that kinetic parameters and yields are reported for D. geothermalis DSM-11302 grown on a mineral medium in well-controlled batch cultur

Changes in sensory and quality characteristics of S. Aethiopicum (Shum) and A. Lividus (Linn) leafy vegetables along the supply chain

Changes in sensory attributes of vegetables over time under different conditions have been reported, however, little has been done regarding profiling and assessing changes in sensory attributes of raw leafy vegetables particularly Solanum aethiopicum (S.) and Amaranthus lividus (L.). This study therefore fills an important knowledge gap of profiling sensory attributes and assessing changes in color, texture and appearance of S.aethiopicum and A.lividus leafy vegetables over time after harvest. A complete randomized design in a 3 ×3 factorial arrangement (each vegetable sample was subjected to three treatments (Time of the day) and three replicates) and data was collected by use of quantitative descriptive sensory analysis. Descriptive data was entered into Microsoft excel spread sheets, averages computed and graphs generated. The data was further subjected to ANOVA and a least significant difference test was used to compare means of samples for all attributes at 95% confidence interval. Correlation analysis using Statistical Package for Social Scientients’ (SPSS version 16.0) was also performed to assess relationship between sensory attributes. Descriptive sensory analysis results showed that all 9:00hrs samples were rated highly for each attribute compared to the 12:00hrs and 15:00hrs samples. ANOVA results for S. aethiopicum showed statistical significant (p<0.05) difference for all the attributes except for light green color of leaf stalk (p<0.05) whereas that for A. lividus showed significant differences for moist appearance, well spread appearance, smoothness and overall quality. Correlation results showed significant positive relationship (p<0.05) among attributes. This study observed that sensory attributes of leafy vegetables change with time after harvest andtraders are therefore encouraged to adopt local cooling systems to help preserve the sensory attributes of vegetables

A review of urinary angiotensin converting enzyme 2 in diabetes and diabetic nephropathy

Urinary angiotensin converting enzyme 2 (ACE2) is significantly increased in diabetes and diabetic nephropathy. While studies on its clinical significance are still underway, its urinary expression, association with metabolic and renal parameters has been in the recent past considerably studied. The recent studies have demystified urine ACE2 in many ways and suggested the roles it could play in the management of diabetic nephropathy. In all studies the expression of urinary ACE2 was determined by enzyme activity assay and/with the quantification of ACE2 protein and mRNA by methods whose reliability are yet to be evaluated. This review summarizes recent findings on expression of urinary ACE2, examines its relationship with clinical parameters and highlights possible applications in management of diabetic nephropathy

Experimental and statistical analysis of nutritional requirements for the growth of the extremophile Deinococcus geothermalis DSM 11300

Few studies concerning the nutritional requirements of Deinococcus geothermalis DSM 11300 have been conducted to date. Three defined media compositions have been published for the growth of this strain but they were found to be inadequate to achieve growth without limitation. Furthermore, growth curves, biomass concentration and growth rates were generally not available. Analysis in Principal Components was used in this work to compare and consequently to highlight the main compounds which differ between published chemically defined media. When available, biomass concentration, and/or growth rate were superimposed to the PCA analysis. The formulations of the media were collected from existing literature; media compositions designed for the growth of several strains of Deinococcaceae or Micrococcaceae were included. The results showed that a defined medium adapted from Holland et al. (Appl Microbiol Biotechnol 72:1074–1082, 2006) was the best basal medium and was chosen for further studies. A growth rate of 0.03 h-1 and a final OD600nm of 0.55 were obtained, but the growth was linear. Then, the effects of several medium components on oxygen uptake and biomass production by Deinococcus geothermalis DSM 11300 were studied using a respirometry-based method, to search for the nutritional limitation. The results revealed that the whole yeast extract in the medium with glucose is necessary to obtain a non-limiting growth of Deinococcus geothermalis DSM 11300 at a maximum growth rate of 0.64 h-1 at 45 °C

Development of a photon-counting near-fano-limited x-ray CMOS image sensor for THESEUS' SXI

THESEUS (Transient High Energy Sky & Early Universe Surveyor) is one of the three candidates for the M5 mission of the European Space Agency. The favoured mission will be announced in 2021 for an expected launch in 2032. THESEUS will be equipped with a Soft X-ray Imager (SXI) composed of a set of two telescopes using micro-pore optics offering an overall field of view of 0.5 sr (<2' accuracy) for X-ray energies between 300 eV and 5 keV. The focal plane of each SXI telescope has a 16 x 16 cm² cooled detector area. However, the limited radiator accommodation on the spacecraft prohibits the use of CCDs since cooling the focal planes to an optimal temperature for radiation hardness (<-100 °C) is not feasible. Therefore, the development of a suitable CMOS Image Sensor (CIS), capable of handling the expected levels of radiation at higher operating temperatures (approximately -30 °C) has been proposed. To demonstrate the performance required for the THESEUS SXI detector, a 2 x 2 cm² prototype is under development using Open University pixel designs in a Teledyne-e2v digital CMOS platform. The pixel design will allow full depletion over silicon thickness of 35 µm for optimal soft X-ray quantum efficiency and instrument background suppression, and will be capable of near-Fano-limited spectral resolution that will also be of prime interest for synchrotron and Free Electron Lasers (FEL) applications. In this paper, we will present the design considerations and simulations leading to the implemented structures complying with THESEUS' SXI requirements

Optical Methods and Their Limitation to Characterize the Morphology and Granulometry of Complex Shape Biological Materials

Background and aim: Particle size and morphology of biomass (microorganism, lignocellulosic substrates) stand out as the major determinants of the bioprocess efficiency. Through its impact on rheology, it affects momentum, heat and and mass transfers within the bioreactor. Various techniques are available to characterize in-situ and ex-situ size and shape of particles. The most common methods are classified into three groups: (i) analysis of microscopic images; (ii) laser light diffraction and (iii) settling kinetics. In present work, five techniques are compared and discussed with model particles, microorganisms and lignocellulosic substrates. Methods: The used techniques aim to characterize size and shape (0.1 to 2000µm). In-situ and ex-situ measurements were used: chord length measurement (FBRM), diffraction light scattering (DLS), morpho-granulometry (MG), cytometry (CYT) and settling velocity (TUL). A set of height polystyrene microspheres (1.0 to 15.0µm) and microbeads (40 and 80µm) were used as reference. Yarrowia lipolytica is strictly aerobic yeast, belonging to the family of hemiascomycetes. Cells are subjected to mycelial transition induced by pH changes. Its morphology evolves from ovoid shape (5-7µm) up to filament. It was used to appreciate the ability to qualify and quantify filamentous shape (width, length). Finally, two cellulosic matrices, microcrystalline cellulose and coniferous paper pulp were selected to investigate complex fiber morphologies. Results: Specifications and limits of instruments are scrutinized. Sampling methods and preparation should be carefully considered. Optical measurements provide raw data (light intensity, frequency, images) from which morphological parameters will be straightly extracted or calculated based on assumptions (optical properties, particles geometry, theory). Considering diameters and associated number and volume distribution functions, techniques are compared with model calibrated microspheres. The mean values appear consistent between techniques but the magnitude of standard deviation extensively varies. Few instruments (MG, CYT) provide access to additional morphological criteria (length, width, aspect ratio). Mycelial kinetics and magnitude is accurately described by fiber length (MG). However a poor reliability of width (time of flight, CYT) is noticeable. Considering more complex lignocellulosic particles, the relative diameter values usually indicate similar trends whatever the techniques is. However, absolute values should be carefully considered and may deviated in large extend (5-10 times)

Characterization of the Exopolysaccharide Produced by Salipiger mucosus A3T, a Halophilic Species Belonging to the Alphaproteobacteria, Isolated on the Spanish Mediterranean Seaboard

We have studied the exopolysaccharide produced by the type strain of Salipiger mucosus, a species of halophilic, EPS-producing (exopolysaccharide-producing) bacterium belonging to the Alphaproteobacteria. The strain, isolated on the Mediterranean seaboard, produced a polysaccharide, mainly during its exponential growth phase but also to a lesser extent during the stationary phase. Culture parameters influenced bacterial growth and EPS production. Yield was always directly related to the quantity of biomass in the culture. The polymer is a heteropolysaccharide with a molecular mass of 250 kDa and its components are glucose (19.7%, w/w), mannose (34%, w/w), galactose (32.9%, w/w) and fucose (13.4%, w/w). Fucose and fucose-rich oligosaccharides have applications in the fields of medicine and cosmetics. The chemical or enzymatic hydrolysis of fucose-rich polysaccharides offers a new efficient way to process fucose. The exopolysaccharide in question produces a solution of very low viscosity that shows pseudoplastic behavior and emulsifying activity on several hydrophobic substrates. It also has a high capacity for binding cations and incorporating considerable quantities of sulfates, this latter feature being very unusual in bacterial polysaccharides