Molecular interactions underlying the unusually high adenosine affinity of a novel Trypanosoma brucei nucleoside transporter

Trypanosoma brucei encodes a relatively high number of genes of the equilibrative nucleoside transporter (ENT) family. We report here the cloning and in-depth characterization of one T. brucei brucei ENT member, TbNT9/AT-D. This transporter was expressed in Saccharomyces cerevisiae and displayed a uniquely high affinity for adenosine (Km = 0.068 +/- 0.013 microM), as well as broader selectivity for other purine nucleosides in the low micromolar range, but was not inhibited by nucleobases or pyrimidines. This selectivity profile is consistent with the P1 transport activity observed previously in procyclic and long-slender bloodstream T. brucei, apart from the 40-fold higher affinity for adenosine than for inosine. We found that, like the previously investigated P1 activity of long/slender bloodstream trypanosomes, the 3'-hydroxy, 5'-hydroxy, N3, and N7 functional groups contribute to transporter binding. In addition, we show that the 6-position amine group of adenosine, but not the inosine 6-keto group, makes a major contribution to binding (DeltaG0 = 12 kJ/mol), explaining the different Km values of the purine nucleosides. We further found that P1 activity in procyclic and long-slender trypanosomes is pharmacologically distinct, and we identified the main gene encoding this activity in procyclic cells as NT10/AT-B. The presence of multiple P1-type nucleoside transport activities in T. brucei brucei facilitates the development of nucleoside-based treatments for African trypanosomiasis and would delay the onset of uptake-related drug resistance to such therapy. We show that both TbNT9/AT-D and NT10/AT-B transport a range of potentially therapeutic nucleoside analogs.

Loss of the high-affinity pentamidine transporter is responsible for high levels of cross-resistance between arsenical and diamidine drugs in African trypanosomes

Treatment of many infectious diseases is under threat from drug resistance. Understanding the mechanisms of resistance is as high a priority as the development of new drugs. We have investigated the basis for cross-resistance between the diamidine and melaminophenyl arsenical classes of drugs in African trypanosomes. We induced high levels of pentamidine resistance in a line without the tbat1 gene that encodes the P2 transporter previously implicated in drug uptake. We isolated independent clones that displayed very considerable cross-resistance with melarsen oxide but not phenylarsine oxide and reduced uptake of [(3)H]pentamidine. In particular, the high-affinity pentamidine transport (HAPT1) activity was absent in the pentamidine-adapted lines, whereas the low affinity pentamidine transport (LAPT1) activity was unchanged. The parental tbat1(-/-) line was sensitive to lysis by melarsen oxide, and this process was inhibited by low concentrations of pentamidine, indicating the involvement of HAPT1. This pentamidine-inhibitable lysis was absent in the adapted line KO-B48. Likewise, uptake of the fluorescent diamidine 4',6-diamidino-2-phenylindole dihydrochloride was much delayed in live KO-B48 cells and insensitive to competition with up to 10 muM pentamidine. No overexpression of the Trypanosoma brucei brucei ATP-binding cassette transporter TbMRPA could be detected in KO-B48. We also show that a laboratory line of Trypanosoma brucei gambiense, adapted to high levels of resistance for the melaminophenyl arsenical drug melarsamine hydrochloride (Cymelarsan), had similarly lost TbAT1 and HAPT1 activity while retaining LAPT1 activity. It seems therefore that selection for resistance to either pentamidine or arsenical drugs can result in a similar phenotype of reduced drug accumulation, explaining the occurrence of cross-resistance.

Arctic patterned-ground ecosystems: A synthesis of field studies and models along a North American Arctic Transect

Arctic landscapes have visually striking patterns of small polygons, circles, and hummocks. The linkages between the geophysical and biological components of these systems and their responses to climate changes are not well understood. The "Biocomplexity of Patterned Ground Ecosystems" project examined patterned-ground features (PGFs) in all five Arctic bioclimate subzones along an 1800-km trans-Arctic temperature gradient in northern Alaska and northwestern Canada. This paper provides an overview of the transect to illustrate the trends in climate, PGFs, vegetation, n-factors, soils, active-layer depth, and frost heave along the climate gradient. We emphasize the thermal effects of the vegetation and snow on the heat and water fluxes within patterned-ground systems. Four new modeling approaches build on the theme that vegetation controls microscale soil temperature differences between the centers and margins of the PGFs, and these in turn drive the movement of water, affect the formation of aggradation ice, promote differential soil heave, and regulate a host of system propel-ties that affect the ability of plants to colonize the centers of these features. We conclude with an examination of the possible effects of a climate wan-ning on patterned-ground ecosystems.

Detection, treatment, and prevention of carbapenemase-producing Enterobacteriaceae : Recommendations from an International Working Group

The prevalence of carbapenemase-producing Enterobacteriaceae (CPE) has increased during the past 10 years. Its detection is frequently difficult, because they do not always show a minimum inhibitory concentration (MIC) value for carbapenems in the resistance range. Both broth microdilution and agar dilution methods are more sensitive than disk diffusion method, Etest and automated systems. Studies on antimicrobial treatment are based on a limited number of patients; therefore, the optimal treatment is not well established. Combination therapy with two active drugs appears to be more effective than monotherapy. Combination of a carbapenem with another active agent — preferentially an aminoglycoside or colistin — could lower mortality provided that the MIC is #4 mg/l and probably #8 mg/l, and is administered in a higher-dose/prolonged-infusion regimen. An aggressive infection control and prevention strategy is recommended, including reinforcement of hand hygiene, using contact precautions and early detection of CPE through use of targeted surveillance.

Notch signaling in the brain: in good and bad times.

Notch signaling is an evolutionarily conserved pathway, which is fundamental for neuronal development and specification. In the last decade, increasing evidence has pointed out an important role of this pathway beyond embryonic development, indicating that Notch also displays a critical function in the mature brain of vertebrates and invertebrates. This pathway appears to be involved in neural progenitor regulation, neuronal connectivity, synaptic plasticity and learning/memory. In addition, Notch appears to be aberrantly regulated in neurodegenerative diseases, including Alzheimer's disease and ischemic injury. The molecular mechanisms by which Notch displays these functions in the mature brain are not fully understood, but are currently the subject of intense research. In this review, we will discuss old and novel Notch targets and molecular mediators that contribute to Notch function in the mature brain and will summarize recent findings that explore the two facets of Notch signaling in brain physiology and pathology.

Das offene Selbstgespräch in Dyaden: Dessen Zusammenhang mit Selbstdarstellungstendenzen und Einfluss auf den Partner

Ausgangslage und Fragestellung In der Forschung zum Selbstgespräch im Sport dominiert die Frage nach dessen Auswir-kungen auf die sprechende Person selber. Gemäss Diaz (1992) besitzt das offene Selbstgespräch aber neben der individuell-regulatorischen auch eine sozial-kommunikative Funktion. Diese sozial-kommunikative Funktion und der damit verbunde-ne Einfluss des Selbstgesprächs auf Beobachter wurden in der Sportwissenschaft bisher nur marginal untersucht (z.B. Gould & Weiss, 1981; Van Raalte, Brewer, Cornelius & Pe-titpas, 2006). Im angestrebten Forschungsvorhaben sollen zwei Fragen geklärt werden: Steht das of-fene, während dem Wettkampf geäusserte Selbstgespräch eines Athleten in Zusammen-hang mit dessen Selbstdarstellung? Beeinflusst das offene Selbstgespräch eines Athle-ten den Dyadenpartner und wie sie sich gegenseitig wahrnehmen? Methode Es werden Doppelpartien im Badminton per Videokamera aufgezeichnet. Mittels einer Korrelationsanalyse soll überprüft werden, ob die so erhobenen Selbstgesprächsaussa-gen mit dem Persönlichkeitsmerkmal „Selbstdarstellung“ der Probanden zusammenhän-gen. Nach Spielschluss werden die Probanden entsprechend eines video-stimulierten Fremdkonfrontationsinterviews mit Spielausschnitten konfrontiert, die offenes Selbstge-spräch ihres Dyadenpartners enthalten. Sie werden dabei nach ihren Kognitionen sowie Emotionen gefragt, die sie während der entsprechenden Spielsituation erlebten. Aktuelle Fragen Welcher Fragebogen zur Erhebung des Persönlichkeitsmerkmals „Selbstdarstellung“ soll Verwendung finden? Zur Auswahl steht ein noch zu übersetzender, validierter, engli-scher und sportspezifischer Fragebogen und die Verwendung eines deutschen Frage-bogens, der habituelle Selbstdarstellungstechniken im Allgemeinen erhebt. Wie soll beim video-stimulierten Fremdkonfrontationsinterview vorgegangen werden? Werden den Probanden z.B. das ganze Spiel oder nur ausgewählte Ausschnitte gezeigt, in denen offenes Selbstgespräch vorkommt? Wie zeitnah muss das Interview durchge-führt werden? Literatur Diaz, R. M. (1992). Methodological Concerns in the Study of Private Speech. In R. M. Diaz & L. E. Berk (Eds.), Private speech. From social interaction to self-regulation (pp. 55-81). Hillsdale, NJ: Lawrence Erlbaum. Gould, D. & Weiss, M. (1981). The effects of model similarity and model talk on self-efficacy and muscular endurance. Journal of Sport Psychology, 3, 17-29. Van Raalte, J. L., Brewer, B. W., Cornelius, A. E. & Petitpas, A. J. (2006). Self-presentational effects of self-talk on perceptions of tennis players. Hellenic Journal of Psychology, 3, 134-149.

A point mutation in cpsE renders Streptococcus pneumoniae nonencapsulated and enhances its growth, adherence and competence

BackgroundThe polysaccharide capsule is a major virulence factor of the important human pathogen Streptococcus pneumoniae. However, S. pneumoniae strains lacking capsule do occur.ResultsHere, we report a nasopharyngeal isolate of Streptococcus pneumoniae composed of a mixture of two phenotypes; one encapsulated (serotype 18C) and the other nonencapsulated, determined by serotyping, electron microscopy and fluorescence isothiocyanate dextran exclusion assay.By whole genome sequencing, we demonstrated that the phenotypes differ by a single nucleotide base pair in capsular gene cpsE (C to G change at gene position 1135) predicted to result in amino acid change from arginine to glycine at position 379, located in the cytoplasmic, enzymatically active, region of this transmembrane protein. This SNP is responsible for loss of capsule production as the phenotype is transferred with the capsule operon. The nonencapsulated variant is superior in growth in vitro and is also 117-fold more adherent to and more invasive into Detroit 562 human epithelial cells than the encapsulated variant.Expression of six competence pathway genes and one competence-associated gene was 11 to 34-fold higher in the nonencapsulated variant than the encapsulated and transformation frequency was 3.7-fold greater.ConclusionsWe identified a new single point mutation in capsule gene cpsE of a clinical S. pneumoniae serotype 18C isolate sufficient to cause loss of capsule expression resulting in the co-existence of the encapsulated and nonencapsulated phenotype. The mutation caused phenotypic changes in growth, adherence to epithelial cells and transformability. Mutation in capsule gene cpsE may be a way for S. pneumoniae to lose its capsule and increase its colonization potential.

Decrease in pneumococcal co-colonization following vaccination with the seven-valent pneumococcal conjugate vaccine.

Understanding the epidemiology of pneumococcal co-colonization is important for monitoring vaccine effectiveness and the occurrence of horizontal gene transfer between pneumococcal strains. In this study we aimed to evaluate the impact of the seven-valent pneumococcal conjugate vaccine (PCV7) on pneumococcal co-colonization among Portuguese children. Nasopharyngeal samples from children up to 6 years old yielding a pneumococcal culture were clustered into three groups: pre-vaccine era (n = 173), unvaccinated children of the vaccine era (n = 169), and fully vaccinated children (4 doses; n = 150). Co-colonization, serotype identification, and relative serotype abundance were detected by analysis of DNA of the total bacterial growth of the primary culture plate using the plyNCR-RFLP method and a molecular serotyping microarray-based strategy. The plyNCR-RFLP method detected an overall co-colonization rate of 20.1%. Microarray analysis confirmed the plyNCR-RFLP results. Vaccination status was the only factor found to be significantly associated with co-colonization: co-colonization rates were significantly lower (p = 0.004; Fisher's exact test) among fully vaccinated children (8.0%) than among children from the pre-PCV7 era (17.3%) or unvaccinated children of the PCV7 era (18.3%). In the PCV7 era there were significantly less non-vaccine type (NVT) co-colonization events than would be expected based on the NVT distribution observed in the pre-PCV7 era (p = 0.024). In conclusion, vaccination with PCV7 resulted in a lower co-colonization rate due to an asymmetric distribution between NVTs found in single and co-colonized samples. We propose that some NVTs prevalent in the PCV7 era are more competitive than others, hampering their co-existence in the same niche. This result may have important implications since a decrease in co-colonization events is expected to translate in decreased opportunities for horizontal gene transfer, hindering pneumococcal evolution events such as acquisition of antibiotic resistance determinants or capsular switch. This might represent a novel potential benefit of conjugate vaccines.

Interpersonale Konsequenzen von offenen Selbstgesprächen in Dyaden im Sport

In der sportpsychologischen Forschung werden sowohl offene als auch verdeckte Selbstgespräche mehrheitlich als rein intrapersonale Phänomene betrachtet und sowohl deren Antezedenzien als auch Konsequenzen nur auf die mit sich selbst sprechende Person bezogen untersucht. Erste laborexperimentelle Studien, die die Effekte von offenen Selbstgesprächen auf die Eindrucksbildung (Graf, Lier & Seiler, 2012; Van Raalte, Brewer, Cornelius & Petitpas, 2006) und die individuelle Leistung (Gould & Weiss, 1981) eines Beobachters nachweisen konnten und Aussagen verschiedener Tennisspieler (Rimmer, Greenlees, Graydon, Thelwell & Buscombe, 2008), dass sie die offene Selbstgespräche der Gegner bei der Bildung eines ersten Eindrucks verwenden würden, deuten jedoch auf die Bedeutsamkeit von interpersonalen Konsequenzen von offenen Selbstgesprächen hin. Um das Verständnis für die interpersonalen Konsequenzen von offenen Selbstgesprächen im Sport zu vertiefen, wurden zwei Studien mit unterschiedlicher Methodik durchgeführt. In einer qualitativen Feldstudie wurden mit 13 Badmintonspielern im Anschluss an ihr Badminton Herren-Doppel Selbstkonfrontationsinterviews durchgeführt, um die Wahrnehmungsergebnisse nach offenen, während eines realen Wettkampfes geäusserten Selbstgesprächen zu untersuchen. Die wahrgenommenen positiven, negativen und motivierenden Selbstgespräche führten zu Zuschreibungen von mentalen Zuständen und Kognitionen und vereinzelt zur Einschätzung des episodischen Leistungsvermögens. Weiter lösten die Selbstgespräche verschiedene affektive Reaktionen und Erwartungsreaktionen aus, deren mögliche Konsequenzen für den weiteren Spielverlauf diskutiert werden. Um einzelne Resultate aus dieser ersten Studie eingehender zu analysieren (zugeschriebene Kompetenz und stellvertretende Valenzeinschätzung; Birrer & Seiler, 2008), und zur Untersuchung der Effekte der offenen Selbstgespräche auf die individuelle Leistung eines Teampartners wurde eine zweite, laborexperimentelle Studie durchgeführt. Zur Herstellung der drei experimentellen Bedingungen kam ein gleichgeschlechtlicher Konfident zum Einsatz, der als vermeintlicher Teampartner fungierte und der während der Golf-Putt-Aufgabe als Reaktion auf die eigenen Putts positive, negative oder keine Selbstgespräche führte. Bezogen auf die zugeschriebene Kompetenz zeigte sich kein allgemeiner, sondern ein von der Valenz der Probanden bedingter Effekt der negativen Selbstgespräche: War den Probanden die Aufgabe wichtig, führten negative Selbstgespräche zur Zuschreibung einer tiefen Kompetenz. Weiter lösten sowohl positive als auch negative Selbstgespräche die Einschätzung aus, dass dem Partner die Aufgabe wichtig ist. Keine Effekte hatten die Selbstgespräche auf die individuelle Leistung der Probanden. Explorativ durchgeführte Nachanalysen deuten jedoch darauf hin, dass sich die Selbstgespräche des Partners womöglich nach einer gewissen Zeit auf die individuelle Leistung der wahrnehmenden Person auswirken könnten, weshalb methodische Anpassungen diskutiert werden, mit Hilfe derer der Effekt der Selbstgespräche auf die Leistung gesteigert werden könnte. Neben den hypothesenprüfenden Tests ermittelten verschiedene Post-Hoc-Analysen aufschlussreiche Resulate (z.B. zu affektiven Reaktionen). Die Ergebnisse der beiden Studien bringen klar zum Ausdruck, dass die interpersonale Perspektive bei der Erforschung der Effekte von offenen Selbstgesprächen bedeutsame Erkenntnisse für Theorie und Praxis liefern kann.

Thrombotic thrombocytopenic purpura during pregnancy versus imitator of preeclampsia.

BACKGROUND Thrombotic thrombocytopenic purpura (TTP) is a severe disorder affecting the microcirculation of multiple organs due to a systemic endothelial cell injury secondary to a deficiency in ADAMTS13 (a disintegrin and metalloprotease with thrombospondin type 1 motif, member 13) activity. TTP is a rare complication of pregnancy with a poor prognosis and high fetal mortality, especially when it occurs during the first trimester. Recent data have supported that effective treatment of TTP is plasma therapy. Unfortunately a major problem remains in the delay in diagnosis due to confounding factors between other "imitators of preeclampsia." Rapid and readily available laboratory testing to quickly diagnose TTP is desperately needed to improve care and to save mother and future child life. CASE REPORT We describe a rare case of successful pregnancy after TTP manifestations occurring in the first trimester; most importantly, our experience represents the first case of atypical manifestation due to neurologic and kidney manifestations preceding laboratory assay alterations. RESULTS We treated a patient with plasma replacement of 30 mL/kg/day and daily plasmapheresis in combination with continuous infusion of fresh-frozen plasma 10 mL/kg/day. The response of clinical manifestation immediately improved. At 30 weeks, the patient had multiple episodes of high blood pressure and concomitant decrease of hemoglobin and platelet count, so a cesarean section was immediately performed. She delivered a healthy female baby. CONCLUSION Early diagnosis by ADAMTS13 activity, occasionally occurring before clinical manifestations, aided us in promptly administering commended and life-saving treatments.