Specimen storage in transport medium and detection of group B streptococci by culture

Recovery of group B streptococci (GBS) was assessed in 1,204 vaginorectal swabs stored in Amies transport medium at 4 or 21°C for 1 to 4 days either by direct inoculation onto Granada agar (GA) or by culture in blood agar (BA) and GA after a selective broth enrichment (SBE) step. Following storage at 4°C, GBS detection in GA was not affected after 72 h by either direct inoculation or SBE; however, GBS were not detected after SBE in the BA subculture in some samples after 48 h of storage and in GA after 96 h. After storage at 21°C, loss of GBS-positive results was significant after 48 h by direct inoculation in GA and after 96 h by SBE and BA subculture; some GBS-positive samples were not detected after 24 h of storage followed by SBE and BA subculture or after 48 h of storage followed by SBE and GA subculture. Storage of swabs in transport medium, even at 4°C, produced after 24 h an underestimation of the intensity of GBS colonization in most specimens. These data indicate that viability of GBS is not fully preserved by storage of vaginorectal swabs in Amies transport medium, mainly if they are not stored under refrigeration

Synthesis and transport of creatine in the CNS: importance for cerebral functions.

J. Neurochem. (2010) 10.1111/j.1471-4159.2010.06935.x Abstract Apart of its well known function of 'energetic buffer' through the creatine/phosphocreatine/creatine kinase system allowing the regeneration of ATP, creatine has been recently suggested as a potential neuromodulator of even true neurotransmitter. Moreover, the recent discovery of primary creatine deficiency syndromes, due to deficiencies in l-arginine : glycine amidinotransferase or guanidinoacetate methyltransferase (the two enzymes allowing creatine synthesis) or in the creatine transporter, has shed new light on creatine synthesis, metabolism and transport, in particular in CNS which appears as the main tissue affected by these creatine deficiencies. Recent data suggest that creatine can cross blood-brain barrier but only with a poor efficiency, and that the brain must ensure parts of its needs in creatine by its own endogenous synthesis. Finally, the recent years have demonstrated the interest to use creatine as a neuroprotective agent in a growing number of neurodegenerative diseases, including Parkinson's and Huntington's diseases. This article aims at reviewing the latest data on creatine metabolism and transport in the brain, in relation to creatine deficiencies and to the potential use of creatine as neuroprotective molecule. Emphasis is also given to the importance of creatine for cerebral function.

The gamma subunit is a specific component of the Na,K-ATPase and modulates its transport function.

The role of small, hydrophobic peptides that are associated with ion pumps or channels is still poorly understood. By using the Xenopus oocyte as an expression system, we have characterized the structural and functional properties of the gamma peptide which co-purifies with Na,K-ATPase. Immuno-radiolabeling of epitope-tagged gamma subunits in intact oocytes and protease protection assays show that the gamma peptide is a type I membrane protein lacking a signal sequence and exposing the N-terminus to the extracytoplasmic side. Co-expression of the rat or Xenopus gamma subunit with various proteins in the oocyte reveals that it specifically associates only with isozymes of Na,K-ATPase. The gamma peptide does not influence the formation and cell surface expression of functional Na,K-ATPase alpha-beta complexes. On the other hand, the gamma peptide itself needs association with Na,K-ATPase in order to be stably expressed in the oocyte and to be transported efficiently to the plasma membrane. Gamma subunits do not associate with individual alpha or beta subunits but only interact with assembled, transport-competent alpha-beta complexes. Finally, electrophysiological measurements indicate that the gamma peptide modulates the K+ activation of Na,K pumps. These data document for the first time the membrane topology, the specificity of association and a potential functional role for the gamma subunit of Na,K-ATPase.

Extremal principle of entropy production in the climate system

Paltridge found reasonable values for the most significant climatic variables through maximizing the material transport part of entropy production by using a simple box model. Here, we analyse Paltridge's box model to obtain the energy and the entropy balance equations separately. Derived expressions for global entropy production, which is a function of the radiation field, and even its material transport component, are shown to be different from those used by Paltridge. Plausible climatic states are found at extrema of these parameters. Feasible results are also obtained by minimizing the radiation part of entropy production, in agreement with one of Planck's results, Finally, globally averaged values of the entropy flux of radiation and material entropy production are obtained for two dynamical extreme cases: an earth with uniform temperature, and an earth in radiative equilibrium at each latitudinal point

Creatine, central nervous system and creatine deficiency syndromes

It was long thought that most of brain creatine was of peripheral origin. However, recentworks have demonstrated that creatine crosses blood-brain barrier only with poor efficiency, and thatCNS must ensure parts of its creatine needs by its own creatine synthesis pathway, thank to the brainexpression of AGAT and GAMT (creatine synthesis) and SLC6A8 (creatine transporter). This newunderstanding of creatine metabolism and transport in CNS allows a better comprehension of creatinedeficiency syndromes, which are due to deficiencies in AGAT, GAMT and SLC6A8 and mainly affectthe brain of patients who show severe neurodevelopmental delay and present neurological symptomsin early infancy.

Acoustic transport of charge and spins in GaAs-based heterostructures

Report for the scientific sojourn carried out at the Paul Drude Institut für Festkörperelektronik of the Stanford University, USA, from 2010 to 2012. The objective of this project is the transport and control of electronic charge and spin along GaAs-based semiconductor heterostructures. The electronic transport has been achieved by taking advantage of the piezolectric field induced by surface acoustic waves in non-centrosymmetric materials like GaAs. This piezolectric field separates photogenerated electrons and holes at different positions along the acoustic wave, where they acummulate and are transported at the same velocity as the wave. Two different kinds of structures have been studied: quantum wells grown along the (110) direction, both intrinsic and n-doped, as well as GaAs nanowires. The analysis of the charge acoustic transport was performed by micro-photoluminescence, whereas the detection of the spin transport was done either by analyzing the polarization state of the emitted photoluminescence or by Kerr reflectometry. Our results in GaAs quantum wells show that charge and spin transport is clearly observed at the non-doped structures,obtaining spin lifetimes of the order of several nanoseconds, whereas no acoutically induced spin transport was detected for the n-doped quantum wells. In the GaAs nanowires, we were able of transporting successfully both electrons and holes along the nanowire axis, but no conservation of the spin polarization has been observed until now. The photoluminescence emitted by these structures after acoustic transport, however, shows anti-bunching characteristics, making this system a very good candidate for its use as single photon emitters.

CAV2 vector gene transfer into central nervious System

Estudi realitzat a partir d’una estada al Institut de Génétique Moléculaire de Montpellier, França, entre 2010 i 2012. En aquest projecte s’ha avaluat les avantatges dels vectors adenovirals canins tipus 2 (CAV2) com a vectors de transferència gènica al sistema nerviós central (SNC) en un model primat no-humà i en un model caní del síndrome de Sly (mucopolisacaridosis tipus 7, MPS VII), malaltia monogènica que cursa amb neurodegeneració. En una primera part del projecte s’ha avaluat la biodistribució, l’eficàcia i la durada de l’expressió del transgen en un model primat no humà, (Microcebus murinus). Com ha vector s’ha utilitzat un CAV2 de primera generació que expressa la proteïna verda fluorescent (CAVGFP). Els resultats aportats en aquesta memòria demostren que en primats no humans, com en d’altres espècies testades anteriorment per l’equip de l’EJ Kremer, la injecció intracerebral de CAV2 resulta en una extensa transducció del SNC, siguent les neurones i els precursors neuronals les cèl•lules preferencialment transduïdes. Els vectors canins, servint-se de vesícules intracel•lulars són transportats, majoritàriament, des de les sinapsis cap al soma neuronal, aquest transport intracel•lular permet una extensa transducció del SNC a partir d’una única injecció intracerebral dels vectors virals. En una segona part d’aquest projecte s’ha avaluat l’ús terapèutic dels CAV2. S’ha injectat un vector helper-dependent que expressa el gen la b-glucuronidasa i el gen de la proteïna verda fluorescent (HD-RIGIE), en el SNC del model caní del síndrome de Sly (MPS VII). La biodistribució i la eficàcia terapèutica han estat avaluades. Els nivells d’activitat enzimàtica en animals malalts injectats amb el vector terapèutic va arribar a valors similars als dels animals no afectes. A més a més s’ha observat una reducció en la quantitat dels GAGs acumulats en les cèl•lules dels animals malalts tractats amb el vector terapèutic, demostrant la potencialitat terapèutica dels CAV2 per a malalties que afecten al SNC. Els resultats aportats en aquest treball ens permeten dir que els CAV2 són unes bones eines terapèutiques per al tractament de malalties que afecten al SNC.

Viral transport of DNA damage that mimics a stalled replication fork.

Adeno-associated virus type 2 (AAV2) infection incites cells to arrest with 4N DNA content or die if the p53 pathway is defective. This arrest depends on AAV2 DNA, which is single stranded with inverted terminal repeats that serve as primers during viral DNA replication. Here, we show that AAV2 DNA triggers damage signaling that resembles the response to an aberrant cellular DNA replication fork. UV treatment of AAV2 enhances the G2 arrest by generating intrastrand DNA cross-links which persist in infected cells, disrupting viral DNA replication and maintaining the viral DNA in the single-stranded form. In cells, such DNA accumulates into nuclear foci with a signaling apparatus that involves DNA polymerase delta, ATR, TopBP1, RPA, and the Rad9/Rad1/Hus1 complex but not ATM or NBS1. Focus formation and damage signaling strictly depend on ATR and Chk1 functions. Activation of the Chk1 effector kinase leads to the virus-induced G2 arrest. AAV2 provides a novel way to study the cellular response to abnormal DNA replication without damaging cellular DNA. By using the AAV2 system, we show that in human cells activation of phosphorylation of Chk1 depends on TopBP1 and that it is a prerequisite for the appearance of DNA damage foci.

Volatile transport during the crystallization of anatectic melts: Oxygen, boron and hydrogen stable isotope study on the metamorphic complex of Naxos, Greece

Crystallization of anatectic melts in high-temperature metamorphic terrains releases volatile-rich magmas that can be transported into adjacent lithologies. This study addresses the variations in the oxygen, boron and hydrogen isotopic composition of aplite-pegmatite dikes that formed during the crystallization of anatectic melts in regional high-temperature metamorphism on the island of Naxos, Greece, and propagated upward into the overlying sequences of metamorphic schist. The transport distance of these dikes was increased through a significant horizontal component of travel that was imposed by contemporaneous low-angle extensional shearing. Laser fluorination oxygen isotope analyses of quartz, tourmaline, garnet, and biotite mineral separates from the aplite-pegmatite dikes show a progressive rise in delta(18)O values with increasing distance from the core. Oxygen isotope fractionations among quartz, tourmaline, and garnet show temperature variations from > 700degreesC down to similar to400degreesC. This range is considered to reflect isotopic fractionation beginning with crystallization at high temperatures in water-undersaturated conditions and then evolving through lower temperature crystallization and retrograde sub-solidus exchange. Two processes are examined for the cause of the progressive increase in delta(18)O values: (1) heterogeneous delta(18)O sources and (2) fluid-rock exchange between the aplite/pegmatite magmas and their host rock. Although the former process cannot be ruled out, there is as yet no evidence in the exposed sequences on Naxos for the presence of a suitable high delta(18)O magma source. In contrast, a tendency for the delta(18)O of quartz in the aplite/pegmatite dikes to approach that of the quartz in the metamorphic rock suggests that fluid-rock exchange with the host rock may potentially be an important process. Advection of fluid into the magma is examined based on Darcian fluid flow into an initially water-undersaturated buoyantly propagating aplitic dike magma. It is shown that such advective flow could only account for part of the O-18-enrichment, unless it were amplified by repeated injection of magma pulses, fluid recycling, and deformation-assisted post-crystallization exchange. The mechanism is, however, adequate to account for hydrogen isotope equilibration between dike and host rock. In contrast, variations in the delta(11)B values of tourmalines suggest that B-11/B-10 fractionation during crystallization and/or magma degassing was the major control of boron geochemistry rather than fluid-rock interaction and that the boron isotopic system was decoupled from that of oxygen. Copyright (C) 2003 Elsevier Ltd.

The inositol Inpp5k 5-phosphatase affects osmoregulation through the vasopressin-aquaporin 2 pathway in the collecting system.

Inositol Inpp5k (or Pps, SKIP) is a member of the inositol polyphosphate 5-phosphatases family with a poorly characterized function in vivo. In this study, we explored the function of this inositol 5-phosphatase in mice and cells overexpressing the 42-kDa mouse Inpp5k protein. Inpp5k transgenic mice present defects in water metabolism characterized by a reduced plasma osmolality at baseline, a delayed urinary water excretion following a water load, and an increased acute response to vasopressin. These defects are associated with the expression of the Inpp5k transgene in renal collecting ducts and with alterations in the arginine vasopressin/aquaporin-2 signalling pathway in this tubular segment. Analysis in a mouse collecting duct mCCD cell line revealed that Inpp5k overexpression leads to increased expression of the arginine vasopressin receptor type 2 and increased cAMP response to arginine vasopressin, providing a basis for increased aquaporin-2 expression and plasma membrane localization with increased osmotically induced water transport. Altogether, our results indicate that Inpp5k 5-phosphatase is important for the control of the arginine vasopressin/aquaporin-2 signalling pathway and water transport in kidney collecting ducts.

Ammonium alters creatine transport and synthesis in a 3D culture of developing brain cells, resulting in secondary cerebral creatine deficiency.

Hyperammonemic disorders in pediatric patients lead to poorly understood irreversible effects on the developing brain that may be life-threatening. We showed previously that some of these NH4+-induced irreversible effects might be due to impairment of axonal growth that can be protected under ammonium exposure by creatine co-treatment. The aim of the present work was thus to analyse how the genes of arginine:glycine amidinotransferase (AGAT) and guanidinoacetate methyltransferase (GAMT), allowing creatine synthesis, as well as of the creatine transporter SLC6A8, allowing creatine uptake into cells, are regulated in rat brain cells under NH4+ exposure. Reaggregated brain cell three-dimensional cultures exposed to NH4Cl were used as an experimental model of hyperammonemia in the developing central nervous system (CNS). We show here that NH4+ exposure differentially alters AGAT, GAMT and SLC6A8 regulation, in terms of both gene expression and protein activity, in a cell type-specific manner. In particular, we demonstrate that NH4+ exposure decreases both creatine and its synthesis intermediate, guanidinoacetate, in brain cells, probably through the inhibition of AGAT enzymatic activity. Our work also suggests that oligodendrocytes are major actors in the brain in terms of creatine synthesis, trafficking and uptake, which might be affected by hyperammonemia. Finally, we show that NH4+ exposure induces SLC6A8 in astrocytes. This suggests that hyperammonemia increases blood-brain barrier permeability for creatine. This is normally limited due to the absence of SLC6A8 from the astrocyte feet lining microcapillary endothelial cells, and thus creatine supplementation may protect the developing CNS of hyperammonemic patients.

Identification of C(4)-dicarboxylate transport systems in Pseudomonas aeruginosa PAO1.

Pseudomonas aeruginosa utilizes preferentially C(4)-dicarboxylates such as malate, fumarate, and succinate as carbon and energy sources. We have identified and characterized two C(4)-dicarboxylate transport (Dct) systems in P. aeruginosa PAO1. Inactivation of the dctA(PA1183) gene caused a growth defect of the strain in minimal media supplemented with succinate, fumarate or malate, indicating that DctA has a major role in Dct. However, residual growth of the dctA mutant in these media suggested the presence of additional C(4)-dicarboxylate transporter(s). Tn5 insertion mutagenesis of the ΔdctA mutant led to the identification of a second Dct system, i.e., the DctPQM transporter belonging to the tripartite ATP-independent periplasmic (TRAP) family of carriers. The ΔdctA ΔdctPQM double mutant showed no growth on malate and fumarate and residual growth on succinate, suggesting that DctA and DctPQM are the only malate and fumarate transporters, whereas additional transporters for succinate are present. Using lacZ reporter fusions, we showed that the expression of the dctA gene and the dctPQM operon was enhanced in early exponential growth phase and induced by C(4)-dicarboxylates. Competition experiments demonstrated that the DctPQM carrier was more efficient than the DctA carrier for the utilization of succinate at micromolar concentrations, whereas DctA was the major transporter at millimolar concentrations. To conclude, this is the first time that the high- and low-affinity uptake systems for succinate DctA and DctPQM have been reported to function coordinately to transport C(4)-dicarboxylates and that the alternative sigma factor RpoN and a DctB/DctD two-component system regulates simultaneously the dctA gene and the dctPQM operon.

The second law of thermodynamics and the global climate system : A review of the maximum entropy production principle

The long-term mean properties of the global climate system and those of turbulent fluid systems are reviewed from a thermodynamic viewpoint. Two general expressions are derived for a rate of entropy production due to thermal and viscous dissipation (turbulent dissipation) in a fluid system. It is shown with these expressions that maximum entropy production in the Earth s climate system suggested by Paltridge, as well as maximum transport properties of heat or momentum in a turbulent system suggested by Malkus and Busse, correspond to a state in which the rate of entropy production due to the turbulent dissipation is at a maximum. Entropy production due to absorption of solar radiation in the climate system is found to be irrelevant to the maximized properties associated with turbulence. The hypothesis of maximum entropy production also seems to be applicable to the planetary atmospheres of Mars and Titan and perhaps to mantle convection. Lorenz s conjecture on maximum generation of available potential energy is shown to be akin to this hypothesis with a few minor approximations. A possible mechanism by which turbulent fluid systems adjust themselves to the states of maximum entropy production is presented as a selffeedback mechanism for the generation of available potential energy. These results tend to support the hypothesis of maximum entropy production that underlies a wide variety of nonlinear fluid systems, including our planet as well as other planets and stars

PDMP blocks brefeldin a-induced retrograde membrane transport from Golgi to ER: evidence for involvement of calcium homeostasis and dissociation from sphingolipid metabolism

In this study, we show that an inhibitor of sphingolipid biosynthesis, d,l-threo-1-phenyl-2- decanoylamino-3-morpholino-1-propanol (PDMP), inhibits brefeldin A (BFA)-induced retrograde membrane transport from Golgi to endoplasmic reticulum (ER). If BFA treatment was combined with or preceded by PDMP administration to cells, disappearance of discrete Golgi structures did not occur. However, when BFA was allowed to exert its effect before PDMP addition, PDMP could not ¿rescue¿ the Golgi compartment. Evidence is presented showing that this action of PDMP is indirect, which means that the direct target is not sphingolipid metabolism at the Golgi apparatus. A fluorescent analogue of PDMP, 6-(N-[7-nitro-2,1,3-benzoxadiazol-4-yl]amino)hexanoyl-PDMP (C6-NBD-PDMP), did not localize in the Golgi apparatus. Moreover, the effect of PDMP on membrane flow did not correlate with impaired C6-NBD-sphingomyelin biosynthesis and was not mimicked by exogenous C6-ceramide addition or counteracted by exogenous C6-glucosylceramide addition. On the other hand, the PDMP effect was mimicked by the multidrug resistance protein inhibitor MK571. The effect of PDMP on membrane transport correlated with modulation of calcium homeostasis, which occurred in a similar concentration range. PDMP released calcium from at least two independent calcium stores and blocked calcium influx induced by either extracellular ATP or thapsigargin. Thus, the biological effects of PDMP revealed a relation between three important physiological processes of multidrug resistance, calcium homeostasis, and membrane flow in the ER/ Golgi system.

The CbrA-CbrB two-component regulatory system controls the utilization of multiple carbon and nitrogen sources in Pseudomonas aeruginosa.

A novel two-component system, CbrA-CbrB, was discovered in Pseudomonas aeruginosa; cbrA and cbrB mutants of strain PAO were found to be unable to use several amino acids (such as arginine, histidine and proline), polyamines and agmatine as sole carbon and nitrogen sources. These mutants were also unable to use, or used poorly, many other carbon sources, including mannitol, glucose, pyruvate and citrate. A 7 kb EcoRI fragment carrying the cbrA and cbrB genes was cloned and sequenced. The cbrA and cbrB genes encode a sensor/histidine kinase (Mr 108 379, 983 residues) and a cognate response regulator (Mr 52 254, 478 residues) respectively. The amino-terminal half (490 residues) of CbrA appears to be a sensor membrane domain, as predicted by 12 possible transmembrane helices, whereas the carboxy-terminal part shares homology with the histidine kinases of the NtrB family. The CbrB response regulator shows similarity to the NtrC family members. Complementation and primer extension experiments indicated that cbrA and cbrB are transcribed from separate promoters. In cbrA or cbrB mutants, as well as in the allelic argR9901 and argR9902 mutants, the aot-argR operon was not induced by arginine, indicating an essential role for this two-component system in the expression of the ArgR-dependent catabolic pathways, including the aruCFGDB operon specifying the major aerobic arginine catabolic pathway. The histidine catabolic enzyme histidase was not expressed in cbrAB mutants, even in the presence of histidine. In contrast, proline dehydrogenase, responsible for proline utilization (Pru), was expressed in a cbrB mutant at a level comparable with that of the wild-type strain. When succinate or other C4-dicarboxylates were added to proline medium at 1 mM, the cbrB mutant was restored to a Pru+ phenotype. Such a succinate-dependent Pru+ property was almost abolished by 20 mM ammonia. In conclusion, the CbrA-CbrB system controls the expression of several catabolic pathways and, perhaps together with the NtrB-NtrC system, appears to ensure the intracellular carbon: nitrogen balance in P. aeruginosa.