Impact of mycelia on the accessibility of fluorene to PAH-degrading bacteria.

Mycelia have been recently shown to actively transport polycyclic aromatic hydrocarbons (PAH) in water-unsaturated soil over the range of centimeters, thereby efficiently mobilizing hydrophobic PAH beyond their purely diffusive transport in air and water. However, the question if mycelia-based PAH transport has an effect on PAH biodegradation was so far unsolved. To address this, we developed a laboratory model microcosm mimicking air-water interfaces in soil. Chemical analyses demonstrated transport of the PAH fluorene (FLU) by the mycelial oomycete Pythium ultimum that was grown along the air-water interfaces. Furthermore, degradation of mycelia-transported FLU by the bacterium Burkholderia sartisoli RP037-mChe was indicated. Since this organism expresses eGFP in response to a FLU flux to the cell, it was also as a bacterial reporter of FLU bioavailability in the vicinity of mycelia. Confocal laser scanning microscopy (CLSM) and image analyses revealed a significant increase of eGFP expression in the presence of P. ultimum compared to controls without mycelia or FLU. Hence, we could show that physically separated FLU becomes bioavailable to bacteria after transport by mycelia. Experiments with silicon coated glass fibers capturing mycelia-transported FLU guided us to propose a three-step mechanism of passive uptake, active transport and diffusion-driven release. These experiments were also used to evaluate the contributions of these individual steps to the overall mycelial FLU transport rate.

L'agrégation toxique des protéines: une forme de "delinquance moléculaire" activement combattue dans la cellule par les chaperones moléculaires et les protéases [The toxic aggregation of proteins: a kind of "molecular delinquency" actively fought in the cell by molecular chaperones and proteases]

Under various stresses, mutation-sensitised proteins may spontaneously convert into inactive, aggregation-prone structures, which may be cytotoxic and infectious. In the cell, this new kind of "molecular criminality" is actively fought against by a network of molecular chaperones that can specifically identify, isolate and unfold damaged (delinquent) proteins and favour their subsequent native refolding. Irreversibly damaged molecules unable to natively refold are preferentially "executed" and recycled by proteases. Failing that, they are "imprisoned" within compact amyloids, or "evicted" from the cell. Thus, striking parallels, although of questionable ethical value, exist between protein and human criminality, and between the cellular and social responses to these different types of criminality. Fundamental differences also exist. Whereas programmed death (apoptosis) is the preferred solution chosen by aged and aggregation-stressed cells, collective suicide is seldom an option chosen by lawless human societies. More significantly, there is no clear cellular equivalent for the role of the family and the education system, which are so essential to the proper shaping of functional individuals in the society, and give rise to humanism, that favours crime prevention, reeducation and reinsertion programs over capital punishment. To the cardiologist and transplantation surgeon, the interest of molecular chaperones, in particular of Hsp70, Hsp90 and Hsp27, lays in their ability to inhibit the signalling pathway of programmed cell death. Their induction before and during ischemia, by various treatments and drugs could significantly reduce damages from the post ischemic reperfusion of organs.

Enzyme activities and pectin breakdown of sapodilla submitted to 1-methylcyclopropene

The objective of this work was to investigate the influence of 1-methylcyclopropene (1-MCP) at 300 nL L-1 on activities of cell wall hidrolytic enzymes and pectin breakdown changes which Sapodilla (Manilkara zapota cv. Itapirema 31) cell wall undergoes during ripening. Sapodilla were treated with ethylene antagonist 1-MCP at 300 nL L-1 for 12 hours and then, stored under a modified atmosphere at 25º C for 23 days. Firmness, total and soluble pectin and cell wall enzymes were monitored during storage. 1-MCP at 300 nL L-1 for 12 hours delayed significantly softening of sapodilla for 11 days at 25º C. 1-MCP postharvest treatment affected the activities of cell wall degrading enzymes pectinmethylesterase and polygalacturonase and completely suppressed increases in beta-galactosidase for 8 days, resulting in less pectin solubilization. Beta-galactosidase seems relevant to softening of sapodilla and is probably responsible for modification of both pectin and xyloglucan-cellulose microfibril network.

Engaging students in an e-learning experience : how to manage information resources through wikis and web blogs

The purpose of this paper is to describe the collaboration between librarians and scholars, from a virtual university, in order to facilitate collaborative learning on how to manage information resources. The personal information behaviour of e-learning students when managing information resources for academic, professional and daily life purposes was studied from 24 semi-structured face-to-face interviews. The results of the content analysis of the interview' transcriptions, highlighted that in the workplace and daily life contexts, competent information behaviour is always linked to a proactive attitude, that is to say, that participants seek for information without some extrinsic reward or avoiding punishment. In the academic context, it was observed a low level of information literacy and it seems to be related with a prevalent uninvolved attitude.

Taking the fungal highway: mobilization of pollutant-degrading bacteria by fungi.

The capacity of fungi to serve as vectors for the dispersion of pollutant-degrading bacteria was analyzed in laboratory model systems mimicking water-saturated (agar surfaces) and unsaturated soil environments (glass-bead-filled columns). Two common soil fungi (Fusarium oxysporum and Rhexocercosporidium sp.) forming hydrophilic and hydrophobic mycelia, respectively, and three polycyclic aromatic hydrocarbon degrading bacteria (Achromobacter sp. SK1, Mycobacterium frederiksbergense LB501TG, and Sphingomonas sp. L138) were selected based on the absence of mutual antagonistic effects. It was shown that fungal hyphae act as vectors for bacterial transport with mobilization strongly depending on the specific microorganisms chosen: The motile strain Achromobacter sp. SK1 was most efficiently spread along hyphae of hydrophilic F. oxysporum in both model systems with transport velocities of up to 1 cm d(-1), whereas no dispersion of the two nonmotile strains was observed in the presence of F. oxysporum. By contrast, none of the bacteria was mobilized along the hydrophobic mycelia of Rhexocercosporidium sp. growing on agar surfaces. In column experiments however, strain SK1 was mobilized by Rhexocercosporidium sp. It is hypothesized that bacteria may move by their intrinsic motilitythrough continuous (physiological) liquid films forming around fungal hyphae. The results of this study suggest that the specific stimulation of indigenous fungi may be a strategy to mobilize pollutant-degrading bacteria leading to their homogenization in polluted soil thereby improving bioremediation.

Examining punishment at different explanatory levels.

Experimental studies on punishment have sometimes been over-interpreted not only for the reasons Guala lists, but also because of a frequent conflation of proximate and ultimate explanatory levels that Guala's review perpetuates. Moreover, for future analyses we may need a clearer classification of different kinds of punishment.

A comparative performance study of four-pole induction motors and synchronous reluctance motors in variable speed drives

Design aspects of the Transversally Laminated Anisotropic (TLA) Synchronous Reluctance Motor (SynRM) are studied and the machine performance analysis compared to the Induction Motor (IM) is done. The SynRM rotor structure is designed and manufactured for a30 kW, four-pole, three-phase squirrel cage induction motor stator. Both the IMand SynRM were supplied by a sensorless Direct Torque Controlled (DTC) variablespeed drive. Attention is also paid to the estimation of the power range where the SynRM may compete successfully with a same size induction motor. A technicalloss reduction comparison between the IM and SynRM in variable speed drives is done. The Finite Element Method (FEM) is used to analyse the number, location and width of flux barriers used in a multiple segment rotor. It is sought for a high saliency ratio and a high torque of the motor. It is given a comparison between different FEM calculations to analyse SynRM performance. The possibility to take into account the effect of iron losses with FEM is studied. Comparison between the calculated and measured values shows that the design methods are reliable. A new application of the IEEE 112 measurement method is developed and used especially for determination of stray load losses in laboratory measurements. The study shows that, with some special measures, the efficiency of the TLA SynRM is equivalent to that of a high efficiency IM. The power factor of the SynRM at rated load is smaller than that of the IM. However, at lower partial load this difference decreases and this, probably, brings that the SynRM gets a better power factor in comparison with the IM. The big rotor inductance ratio of the SynRM allows a good estimating of the rotor position. This appears to be very advantageous for the designing of the rotor position sensor-less motor drive. In using the FEM designed multi-layer transversally laminated rotor with damper windings it is possible to design a directly network driven motor without degrading the motorefficiency or power factor compared to the performance of the IM.

Evolutionary Games in Networked Populations: Models and Experiments

Cooperation and coordination are desirable behaviors that are fundamental for the harmonious development of society. People need to rely on cooperation with other individuals in many aspects of everyday life, such as teamwork and economic exchange in anonymous markets. However, cooperation may easily fall prey to exploitation by selfish individuals who only care about short- term gain. For cooperation to evolve, specific conditions and mechanisms are required, such as kinship, direct and indirect reciprocity through repeated interactions, or external interventions such as punishment. In this dissertation we investigate the effect of the network structure of the population on the evolution of cooperation and coordination. We consider several kinds of static and dynamical network topologies, such as Baraba´si-Albert, social network models and spatial networks. We perform numerical simulations and laboratory experiments using the Prisoner's Dilemma and co- ordination games in order to contrast human behavior with theoretical results. We show by numerical simulations that even a moderate amount of random noise on the Baraba´si-Albert scale-free network links causes a significant loss of cooperation, to the point that cooperation almost vanishes altogether in the Prisoner's Dilemma when the noise rate is high enough. Moreover, when we consider fixed social-like networks we find that current models of social networks may allow cooperation to emerge and to be robust at least as much as in scale-free networks. In the framework of spatial networks, we investigate whether cooperation can evolve and be stable when agents move randomly or performing Le´vy flights in a continuous space. We also consider discrete space adopting purposeful mobility and binary birth-death process to dis- cover emergent cooperative patterns. The fundamental result is that cooperation may be enhanced when this migration is opportunistic or even when agents follow very simple heuristics. In the experimental laboratory, we investigate the issue of social coordination between indi- viduals located on networks of contacts. In contrast to simulations, we find that human players dynamics do not converge to the efficient outcome more often in a social-like network than in a random network. In another experiment, we study the behavior of people who play a pure co- ordination game in a spatial environment in which they can move around and when changing convention is costly. We find that each convention forms homogeneous clusters and is adopted by approximately half of the individuals. When we provide them with global information, i.e., the number of subjects currently adopting one of the conventions, global consensus is reached in most, but not all, cases. Our results allow us to extract the heuristics used by the participants and to build a numerical simulation model that agrees very well with the experiments. Our findings have important implications for policymakers intending to promote specific, desired behaviors in a mobile population. Furthermore, we carry out an experiment with human subjects playing the Prisoner's Dilemma game in a diluted grid where people are able to move around. In contrast to previous results on purposeful rewiring in relational networks, we find no noticeable effect of mobility in space on the level of cooperation. Clusters of cooperators form momentarily but in a few rounds they dissolve as cooperators at the boundaries stop tolerating being cheated upon. Our results highlight the difficulties that mobile agents have to establish a cooperative environment in a spatial setting without a device such as reputation or the possibility of retaliation. i.e. punishment. Finally, we test experimentally the evolution of cooperation in social networks taking into ac- count a setting where we allow people to make or break links at their will. In this work we give particular attention to whether information on an individual's actions is freely available to poten- tial partners or not. Studying the role of information is relevant as information on other people's actions is often not available for free: a recruiting firm may need to call a job candidate's refer- ences, a bank may need to find out about the credit history of a new client, etc. We find that people cooperate almost fully when information on their actions is freely available to their potential part- ners. Cooperation is less likely, however, if people have to pay about half of what they gain from cooperating with a cooperator. Cooperation declines even further if people have to pay a cost that is almost equivalent to the gain from cooperating with a cooperator. Thus, costly information on potential neighbors' actions can undermine the incentive to cooperate in dynamical networks.

Generalist hydrocarbon-degrading bacterial communities in the oil-polluted water column of the North Sea.

The aim of this work was to determine the effect of light crude oil on bacterial communities during an experimental oil spill in the North Sea and in mesocosms (simulating a heavy, enclosed oil spill), and to isolate and characterize hydrocarbon-degrading bacteria from the water column. No oil-induced changes in bacterial community (3 m below the sea surface) were observed 32 h after the experimental spill at sea. In contrast, there was a decrease in the dominant SAR11 phylotype and an increase in Pseudoalteromonas spp. in the oiled mesocosms (investigated by 16S rRNA gene analysis using denaturing gradient gel electrophoresis), as a consequence of the longer incubation, closer proximity of the samples to oil, and the lack of replenishment with seawater. A total of 216 strains were isolated from hydrocarbon enrichment cultures, predominantly belonging to the genus Pseudoaltero monas; most strains grew on PAHs, branched and straight-chain alkanes, as well as many other carbon sources. No obligate hydrocarbonoclastic bacteria were isolated or detected, highlighting the potential importance of cosmopolitan marine generalists like Pseudoalteromonas spp. in degrading hydrocarbons in the water column beneath an oil slick, and revealing the susceptibility to oil pollution of SAR11, the most abundant bacterial clade in the surface ocean.

Action Dominates Valence in Anticipatory Representations in the Human Striatum and Dopaminergic Midbrain

The acquisition of reward and the avoidance of punishment could logically be contingent on either emitting or withholding particular actions. However,the separate pathways inthe striatumfor go and no-go appearto violatethis independence, instead coupling affect and effect. Respect for this interdependence has biased many studies of reward and punishment, so potential action- outcome valence interactions during anticipatory phases remain unexplored. In a functional magnetic resonance imaging study with healthy human volunteers, we manipulated subjects" requirement to emit or withhold an action independent from subsequent receipt of reward or avoidance of punishment. During anticipation, in the striatum and a lateral region within the substantia nigra/ventral tegmental area (SN/VTA), action representations dominated over valence representations. Moreover, we did not observe any representation associated with different state values through accumulation of outcomes, challenging a conventional and dominant association between these areas and state value representations. In contrast, a more medial sector of the SN/VTA responded preferentially to valence, with opposite signs depending on whether action was anticipatedto be emitted or withheld. This dominant influence of action requires an enriched notion of opponency between reward and punishment.

Characterization of an antennal carboxylesterase from the pest moth Spodoptera littoralis degrading a host plant odorant.

Background: Carboxyl/cholinesterases (CCEs) are highly diversified in insects. These enzymes have a broad range of proposed functions, in neuro/developmental processes, dietary detoxification, insecticide resistance or hormone/pheromone degradation. As few functional data are available on purified or recombinant CCEs, the physiological role of most of these enzymes is unknown. Concerning their role in olfaction, only two CCEs able to metabolize sex pheromones have been functionally characterized in insects. These enzymes are only expressed in the male antennae, and secreted into the lumen of the pheromone-sensitive sensilla. CCEs able to hydrolyze other odorants than sex pheromones, such as plant volatiles, have not been identified. Methodology: In Spodoptera littoralis, a major crop pest, a diversity of antennal CCEs has been previously identified. We have employed here a combination of molecular biology, biochemistry and electrophysiology approaches to functionally characterize an intracellular CCE, SlCXE10, whose predominant expression in the olfactory sensilla suggested a role in olfaction. A recombinant protein was produced using the baculovirus system and we tested its catabolic properties towards a plant volatile and the sex pheromone components. Conclusion: We showed that SlCXE10 could efficiently hydrolyze a green leaf volatile and to a lesser extent the sex pheromone components. The transcript level in male antennae was also strongly induced by exposure to this plant odorant. In antennae, SlCXE10 expression was associated with sensilla responding to the sex pheromones and to plant odours. These results suggest that a CCE-based intracellular metabolism of odorants could occur in insect antennae, in addition to the extracellular metabolism occurring within the sensillar lumen. This is the first functional characterization of an Odorant- Degrading Enzyme active towards a host plant volatile.

Integrated regulation of the type III secretion system and other virulence determinants in Ralstonia solanacearum

In many plant and animal bacterial pathogens, the Type III secretion system (TTSS) that directly translocates effector proteins into the eukaryotic host cells is essential for the development of disease. In all species studied, the transcription of the TTSS and most of its effector substrates is tightly regulated by a succession of consecutively activated regulators. However, the whole genetic programme driven by these regulatory cascades is still unknown, especially in bacterial plant pathogens. Here, we have characterised the programme triggered by HrpG, a host-responsive regulator of the TTSS activation cascade in the plant pathogen Ralstonia solanacearum. We show through genome-wide expression analysis that, in addition to the TTSS, HrpG controls the expression of a previously undescribed TTSS-independent pathway that includes a number of other virulence determinants and genes likely involved in adaptation to life in the host. Functional studies revealed that this second pathway co-ordinates the bacterial production of plant cell wall-degrading enzymes, exopolysaccharide, and the phytohormones ethylene and auxin. We provide experimental evidence that these activities contribute to pathogenicity. We also show that the ethylene produced by R. solanacearum is able to modulate the expression of host genes and can therefore interfere with the signalling of plant defence responses. These results provide a new, integrated view of plant bacterial pathogenicity, where a common regulator activates synchronously upon infection the TTSS, other virulence determinants and a number of adaptive functions, which act co-operatively to cause disease.

Environmental activity of bacteria degrading pollutants

Une des meilleures techniques pour décontaminer l'environnement d'éléments toxiques (comme par exemple le dibenzofuan, DBF et le 4-chlorophenol, 4CP) déposés par l'homme, à bas coûts et sans le perturber considérablement, est sans doute la biorémédiation, et particulièrement la bioaugmentation. Malheureusement, si plusieurs microorganismes ont démontré leur efficacité à dégrader les composés toxiques en conditions de laboratoire, plusieurs tentatives afin de les utiliser dans l'environnement n'ont pas abouti. Ces échecs sont probablement le résultat des pauvres connaissances des réactions de ces mêmes microorganismes dans l'environnement. L'objectif de mon travail a été de mieux comprendre les réponses de ces bactéries au niveau de leurs gènes lorsqu'elles sont introduites ou prospèrent dans des conditions plus proches de la réalité, mais encore suffisamment contrôlées pour pouvoir élucider leur comportement. Le fait de résister à des conditions de sécheresse a été considéré en tant que facteur clé dans la survie des bactéries amenées à être utilisées pour la biorémédiation; cela implique une série de mécanismes utilisés par la cellule pour faire face au stress hydrique. Le chapitre II, par une approche métagénomique, compare les réactions de trois souches prometteuses pour la biorémédiation (Arthrobacter chlorophenolicus A6, Sphingomonas wittichii RW1 and Pseudomonas veronii 1YdBTEX2) vis-à-vis du stress hydrique simulé en conditions de laboratoire. L'objectif ici est de découvrir et de décrire les stratégies de résistance au stress, communes ou spécifiques, employées par les bactéries. Mes résultats montrent que les trois souches ont des sensibilités différentes au stress hydrique. Entre les traits communs trouvés, il y a une diminution de l'expression des gènes flagellaires ainsi qu'une augmentation de l'expression de solutes compatibles, mais qui sont souche-spécifiques. J'ai étudié plus en détail la réponse génomique de RW1 par rapport aux inoculations ainsi que sa croissance dans le sable contaminé et non-stérile (chapitre III), et je les ai comparé à des cultures en milieu liquide. Mes résultats indiquent que RW1 peut résister efficacement et peut croître dans des conditions presque sèches et peut également dégrader le contaminant (DBF, dans le cas présent) si les pré-cultures sont réalisées dans le même type de contaminant. Par contre, notre hypothèse du chapitre II se révèle fausse car le comportement de RW1 est très diffèrent de celui observé dans des conditions avec stress hydrique induit par l'addition de sel ou de PEG. Plus intéressant, les réponses de RW1 en milieu liquide sont très différentes de celles observées dans le sable, révélant ainsi que cette souche peut reconnaître le milieu dans lequel elle se trouve. Les mêmes expériences en sable contaminé, cette fois-ci avec 4CP, ont été réalisées pour A6 (chapitre IV) dans l'espoir de compléter la comparaison entre le stress hydrique et l'adaptation dans le sol. Malheureusement, il n'a pas été possible d'obtenir d'échantillons de bonne qualité pour les hybridations des microarrays afin d'étudier la réponse transcriptionnelle dans les différentes phases de croissance dans le sable (contaminé ou non). Toutefois, j'ai appris qu'Arthrobacter ne peut pas croitre dans les sols hautement contaminés si les conditions du sol sont très sèches, elles ont en effet besoin de suffisamment d'eau pour dégrader des quantités importantes de 4CP. Ces observations dirigent l'attention sur le fait que les études sur l'efficacité de l'inoculation de bactéries doivent être testées dans des conditions le plus proche possible de l'environnement ciblé, tout comme les concentrations optimales pour l'inoculum. Finalement, nous avons étudié le comportement de A6 dans la phytosphère avec deux dégrés d'humidité (chapitre V). A6 ne montre pas de réaction particulière face aux changements d'humidité, et à nouveau, ces réponses ne peuvent être liées aux changements d'expression des gènes observées dans les conditions de stress hydrique simulées. Cette étude a permis d'identifier la présence de composés phénoliques dans les feuilles qui peuvent potentiellement améliorer les propriétés de dégradation ou qui permettent d'effectuer de façon plus rapide la réaction de dégradation des contaminants dans un processus de phytoremédiation par A. chlorophenolicus.

Sodium-dependent phosphate transporters in osteoclast differentiation and function.

Osteoclasts are multinucleated bone degrading cells. Phosphate is an important constituent of mineralized bone and released in significant quantities during bone resorption. Molecular contributors to phosphate transport during the resorptive activity of osteoclasts have been controversially discussed. This study aimed at deciphering the role of sodium-dependent phosphate transporters during osteoclast differentiation and bone resorption. Our studies reveal RANKL-induced differential expression of sodium-dependent phosphate transport protein IIa (NaPi-IIa) transcript and protein during osteoclast development, but no expression of the closely related NaPi-IIb and NaPi-IIc SLC34 family isoforms. In vitro studies employing NaPi-IIa-deficient osteoclast precursors and mature osteoclasts reveal that NaPi-IIa is dispensable for bone resorption and osteoclast differentiation. These results are supported by the analysis of structural bone parameters by high-resolution microcomputed tomography that yielded no differences between adult NaPi-IIa WT and KO mice. By contrast, both type III sodium-dependent phosphate transporters Pit-1 and Pit-2 were abundantly expressed throughout osteoclast differentiation, indicating that they are the relevant sodium-dependent phosphate transporters in osteoclasts and osteoclast precursors. We conclude that phosphate transporters of the SLC34 family have no role in osteoclast differentiation and function and propose that Pit-dependent phosphate transport could be pivotal for bone resorption and should be addressed in further studies.

Interleukin-6 and chondrocyte mineralisation act in tandem to promote experimental osteoarthritis.

OBJECTIVES: Basic calcium phosphate (BCP) crystal and interleukin 6 (IL-6) have been implicated in osteoarthritis (OA). We hypothesise that these two factors may be linked in a reciprocal amplification loop which leads to OA. METHODS: Primary murine chondrocytes and human cartilage explants were incubated with hydroxyapatite (HA) crystals, a form of BCP, and the modulation of cytokines and matrix-degrading enzymes assayed. The ability of IL-6 to stimulate chondrocyte calcification was assessed in vitro. The mechanisms underlying the effects of HA on chondrocytes were investigated using chemical inhibitors, and the pathways mediating IL-6-induced calcification characterised by quantifying the expression of genes involved in chondrocyte mineralisation. The role of calcification in vivo was studied in the meniscectomy model of murine OA (MNX), and the link between IL-6 and cartilage degradation investigated by histology. RESULTS: In chondrocytes, BCP crystals stimulated IL-6 secretion, further amplified in an autocrine loop, through signalling pathways involving Syk and PI3 kinases, Jak2 and Stat3 molecules. Exogenous IL-6 promoted calcium-containing crystal formation and upregulation of genes involved in calcification: the pyrophosphate channel Ank, the calcium channel Annexin5 and the sodium/phosphate cotransporter Pit-1. Treatment of chondrocytes with IL-6 inhibitors significantly inhibited IL-6-induced crystal formation. In meniscectomised mice, increasing deposits of BCP crystals were observed around the joint and correlated with cartilage degradation and IL-6 expression. Finally, BCP crystals induced proteoglycan loss and IL-6 expression in human cartilage explants, which were reduced by an IL-6 inhibitor. CONCLUSIONS: BCP crystals and IL-6 form a positive feedback loop leading to OA. Targeting calcium-containing crystal formation and/or IL-6 are promising therapeutic strategies in OA.