Etablierung und Optimierung einer PCR-basierten Methode zur Detektion von genetischen Veränderungen im PDPK1-Gen bei Kindern mit intrauterinem und postnatalem Kleinwuchs

Untersuchungen zeigen, dass Kinder mit einem Geburtsgewicht und/oder einer Geburtslänge unterhalb der 3. Perzentile der populationsspezifischen Norm (SGA) u. a. ein erhöhtes Risiko für kardiovaskuläre und metabolische Krankheitsbilder haben. Mausembryonen mit einer Funktionseinschränkung der PDPK1 sind überlebensfähig, haben aber ein um 40-50 Prozent vermindertes Längenwachstum als Wurfgeschwister. Im Rahmen dieser Arbeit wurden bei 111 SGA-Kindern mit nachgewiesenem proportioniertem Kleinwuchs ohne Aufholwachstum, dem Phänotyp einer IGF1 Resistenz und einem im Vorfeld ausgeschlossenem IGF1 Rezeptordefekt die Nukleotidabfolge der kodierenden Bereiche der PDPK1 untersucht. Dabei konnte für die Exons 1, 3 und 4, sowie 7 bis 14 eine PCR-Reaktion etabliert werden, wobei das PCR Produkt mittels dHPLC und/oder Sequenzierung untersucht wurde. Methodische Probleme ergaben sich aus der Existenz eines Pseudogen, sodass für die Exons 2-10 eine Vor-PCR etabliert werden musste. Aufgrund der Länge des homologen Bereichs und der damit verbundenen relativ weiten Entfernung zu spezifischen Primerbindungsstellen konnten nicht alle Exons mit ausreichender Spezifität amplifiziert werden. Es wurden insgesamt 6 verschiedene intronische Varianten bei 11 Patienten nachgewiesen, welche bisher in der Literatur noch nicht beschrieben worden sind. Ob oder in wie weit diese Sequenzvarianten eine Rolle bei der Entwicklung des SGA-Phänotyps der Kinder gespielt haben, ist aufgrund der Lokalisation außerhalb der Exons bzw. wegen ihres seltenen Auftretens auch in der Gesundpopulation unklar. Möglich wären z. B. Wechselwirkungen mit Promoterregionen und eine daraufhin eingeschränkte Funktion der Aktivität der PDPK1. Weiterführende methodische Optimierungen oder alternative Analysetechniken sowie die Untersuchung größerer Patientenkollektive mit geeigneten Kontrollgruppen werden helfen, die Relevanz der PDPK1 bei der Ausprägung eines isolierten Minderwuchses bei Patienten mit IGF1-Resistenz zu etablieren

Structurally rich dry grasslands – Potential stepping stones for bats in open farmland

Agricultural intensification has caused decrease and fragmentation of European semi-natural dry grasslands. While a high biodiversity value of dry grasslands is acknowledged for plants and insects, locally and on landscape level, their relevance for mobile species, such as bats, is unknown. Here we investigate the use of dry grassland fragments by bats in an agriculturally intensified region in Germany and evaluate local and landscape factors influencing bat activity and assemblages. Specifically, we predicted that a combination of local dry grassland structural richness and landscape features as well as their interactions affect bat activity and foraging above dry grasslands. We also expected that these features influence compositions of local bat assemblages. We repeatedly sampled at 12 dry grassland plots with acoustic monitoring and assessed activity and foraging of bat species/sonotypes, which we grouped into guilds known for foraging in open land, at vegetation edges and in narrow spaces. We determined structural richness of the dry grassland plots in field and derived landscape features from digital landscape data. A relatively high proportion of bat species/sonotypes used dry grasslands regularly. The edge space foragers responded positively to higher local structural richness. Their dry grassland use increased when surrounding forests and woody features were less available, but they foraged more on dry grasslands closer to water bodies. Narrow space bat activity on dry grasslands decreased with less landscape connectivity. Open and narrow space foragers responded to local structural richness only in landscape context. For all bat guilds we found increased use of structurally richer dry grasslands when there was more open farmland in the surroundings. This was also the case for edge space foragers, when landscapes were more homogeneous. Lastly, with increasing structural richness, bat assemblages were more dominated by edge space foragers. We show the importance of European dry grassland fragments for the highly mobile group of bats under certain local structural and landscape compositional conditions. Our results underline the value of heterogeneous dry grassland fragments as potential stepping stones in intensively used farmland areas and contribute to evidence based decision making in dry grassland management and bat conservation

Methods for the automatic recording of bird calls and songs in field ornithology

Der gegenwärtige Kenntnisstand über automatisierte Methoden zur akustischen Erfassung von Rufen und Gesängen von Vögeln wird dargelegt. Die Grundlage für eine automatisierte Erfassung bilden Langzeitaufzeichnungen. Es wird der Frage nachgegangen, inwiefern Tonaufzeichnungen für eine qualitative und auch quantitative Analyse von Vogelbeständen geeignet sind. Spezielles Augenmerk wird autonomen Aufzeichnungsmethoden und der Auswertung von Langzeitaufzeichnungen unter Nutzung von Algorithmen der akustischen Mustererkennung gewidmet. Sinnvolle Einsatzszenarien für automatisierte Methoden im Rahmen avifaunistischer Feldforschung sind die Erfassung des nächtlichen Vogelzuges, die Erfassung nachtaktiver Brutvogelarten und die Datenerhebung in Kernzonen von Schutzgebieten.This review presents our current knowledge on automated methods for acoustic recording of calls and songs of birds. Acoustic long-term recordings can serve as a basis for an automated bird census. We stress the question of whether sound recordings are suitable for qualitative and quantitative analysis of bird populations. Special attention is devoted to autonomous recording methods and the evaluation of long-term recordings by use of acoustic pattern recognition algorithms. Realistic scenarios for the use of automated methods in field ornithology we see in the investigation of nocturnal bird migration, the census of nocturnal bird species, and data collection in core areas of nature reserves

The genome of Romanomermis culicivorax:revealing fundamental changes in the core developmental genetic toolkit in Nematoda

Background: The genetics of development in the nematode Caenorhabditis elegans has been described in exquisite detail. The phylum Nematoda has two classes: Chromadorea (which includes C. elegans) and the Enoplea. While the development of many chromadorean species resembles closely that of C. elegans, enoplean nematodes show markedly different patterns of early cell division and cell fate assignment. Embryogenesis of the enoplean Romanomermis culicivorax has been studied in detail, but the genetic circuitry underpinning development in this species has not been explored. Results: We generated a draft genome for R. culicivorax and compared its gene content with that of C. elegans, a second enoplean, the vertebrate parasite Trichinella spiralis, and a representative arthropod, Tribolium castaneum. This comparison revealed that R. culicivorax has retained components of the conserved ecdysozoan developmental gene toolkit lost in C. elegans. T. spiralis has independently lost even more of this toolkit than has C. elegans. However, the C. elegans toolkit is not simply depauperate, as many novel genes essential for embryogenesis in C. elegans are not found in, or have only extremely divergent homologues in R. culicivorax and T. spiralis. Our data imply fundamental differences in the genetic programmes not only for early cell specification but also others such as vulva formation and sex determination. Conclusions: Despite the apparent morphological conservatism, major differences in the molecular logic of development have evolved within the phylum Nematoda. R. culicivorax serves as a tractable system to contrast C. elegans and understand how divergent genomic and thus regulatory backgrounds nevertheless generate a conserved phenotype. The R. culicivorax draft genome will promote use of this species as a research model

Salmonella Typhimurium impairs glycolysismediated acidification of phagosomes to evade macrophage defense

Regulation of cellular metabolism is now recognized as a crucial mechanism for the activation of innate and adaptive immune cells upon diverse extracellular stimuli. Macrophages, for instance, increase glycolysis upon stimulation with pathogen-associated molecular patterns (PAMPs). Conceivably, pathogens also counteract these metabolic changes for their own survival in the host. Despite this dynamic interplay in host-pathogen interactions, the role of immunometabolism in the context of intracellular bacterial infections is still unclear. Here, employing unbiased metabolomic and transcriptomic approaches, we investigated the role of metabolic adaptations of macrophages upon Salmonella enterica serovar Typhimurium (S. Typhimurium) infections. Importantly, our results suggest that S. Typhimurium abrogates glycolysis and its modulators such as insulin-signaling to impair macrophage defense. Mechanistically, glycolysis facilitates glycolytic enzyme aldolase A mediated v- ATPase assembly and the acidification of phagosomes which is critical for lysosomal degradation. Thus, impairment in the glycolytic machinery eventually leads to decreased bacterial clearance and antigen presentation in murine macrophages (BMDM). Collectively, our results highlight a vital molecular link between metabolic adaptation and phagosome maturation in macrophages, which is targeted by S. Typhimurium to evade cell-autonomous defense. Copyright

Standardized high-throughput evaluation of cell-based compound screens

<p>Abstract</p> <p>Background</p> <p>High-throughput screening of pharmaceutical compound activity in tissue culture experiments requires time-consuming repeated analysis of the large amounts of data generated. Automation of the evaluation procedure and assessment of measurement accuracy can save time and improve the comparability of results.</p> <p>Results</p> <p>We present a tool for simultaneous evaluation of an arbitrary number of compound screens including a standardized statistical validation. It is provided as a novel R package with a Tcl/Tk-based GUI for convenient use in the lab and runs on usual platforms like Linux, Windows and Mac OS. In a compound screen of lung cancer cells, the tool was successfully and efficiently applied for data analysis.</p> <p>Conclusion</p> <p>The package provides an efficient and intuitive platform for automatic evaluation of compound screens, improving the performance and standardization of data analysis.</p

Mutations in NNT encoding nicotinamide nucleotide transhydrogenase cause familial glucocorticoid deficiency

This work has been supported by the Medical Research Council UK (New Investigator Research Grant G0801265 to L.A.M., Clinical Research Training Fellowship Grant G0901980 to C.R.H. and Project Grant G0700767 to P.J.K.)

Control of mitogenic and motogenic pathways by miR-198, diminishing hepatoma cell growth and migration

Abstract Hepatocellular carcinoma (HCC) is one of the leading causes of cancer deaths, worldwide. MicroRNAs, inhibiting gene expression by targeting various transcripts, are involved in genomic dysregulation during hepatocellular tumorigenesis. In previous studies, microRNA-198 (miR-198) was shown to be significantly downregulated in HCV-positive hepatocellular carcinoma (HCC). Herein, the function of miR-198 in hepatocellular carcinoma cell growth and gene expression was studied. In hepatoma cell-types with low levels of liver-specific transcription factor HNF1α indicating a low differentiation grade, miR-198 expression was most downregulated. However, miR-198 treatment did not restore the expression of the liver-specific transcription factors HNF1α or HNF4α. Importantly, overexpression of miR-198 in Pop10 hepatoma cells markedly reduced cell growth. In agreement, comprehensive gene expression profiling by microarray hybridisation and real-time quantification revealed that central signal transducers of proliferation pathways were downregulated by miR-198. In contrast, genes mediating cellular adherence were highly upregulated by miR-198. Thus, the low expression of E-cadherin and claudin-1, involved in cell adhesion and cell-cell contacts, was abolished in hepatoma cells after miR-198 overexpression. This definite induction of both proteins by miR-198 was shown to be accompanied by a significantly impaired migration activity of hepatoma Pop10 cells. In conclusion, miR-198 acts as a tumor suppressor by repression of mitogenic and motogenic pathways diminishing cell growth and migration