Effects of Clearcutting on Forage Production, Quality and Decomposition in the Caatinga Woodland of Northeast Brazil: Implications to Goat and Sheep Nutrition

Clearing of trees from the so-called caatinga woodland that characterizes the vegetation of the semi-arid region of northeast Brazil offers possibilities for increasing forage production. This research analyzed the first-year effects of clearing caatinga on dry season forage for goats and sheep. In addition, factors affecting litter decomposition on cleared and uncleared caatinga were assessed to evaluate the viability of deferring grazing of forages during the wet season for use later in the dry season. Removing the trees resulted in a sixfold increase in production of herbaceous vegetation, however, 88 percent of the increased yield on the cleared areas was in the form of stems from herbaceous vegetation. Seventy-two percent of the stems were unpalatable to goats and sheep because of the massive size of those stems. Leaf litter from trees was an important component of the diets of goats and sheep during the dry season and clearing reduced production of this forage threefold. Clearing resulted in increased decomposition of leaf litter. Changes in microclimate played only a minor role in this difference. The reduction in the amount of leaf litter from trees relative to litter from herbs had the greatest effect on decomposition rates of dry season forage because tree litter decomposed less rapidly than did herbaceous litter. The slow decomposition of leaf litter during the dry season suggests that deferment. of cleared or uncleared caatinga for use as forage in the latter part of the dry season is feasible. An analysis of the diets of esophageally fistulated goats and sheep indicated that clearing may be a viable alternative for improving the amount and the in vitro dry matter digestibility of the forage consumed during the dry season the first-year post-treatment. These increases were attributed to an absolute greater abundance of preferred herbaceous forages (i.e., foliage and leaf litter) and to the persistent green foliage on coppicing woody plants. Dietary nitrogen appeared to limit intake, and clearing did not improve availability of this nutrient to sheep and goats at the higher levels of grazing pressure applied in this study. Other ecosystem considerations such as watershed protection and long-term community stability must also be considered in decisions to remove the tree canopy of the caatinga

Studium der Wachstumskinetik von Intermediärfilamenten mit Hilfe von Vimentin

In dieser Arbeit wurde die Wachstumskinetik der in vitro Filamentebildung des Intermediärfilamentproteins Vimentin untersucht, und versucht diese quantitativ zu beschreiben. Als Grundlage dienten bestehende qualitative Beschreibungen der Filamentbildung. Das Protein Vimentin wurde rekombinant in E. coli hergestellt und über Säulenaustauschchromatographie gereinigt und in 8 M Harnstoff- Puffer gelagert. Die Renaturierung von Vimentin vor den Versuchen erfolgte durch ein Dialyseverfahren gegen Pi-Puffer (2 mM Na-Phosphat, pH 7.5). Schon während der Dialyse bilden sich zuerst Vimentin-Dimere gefolgt von Tetrameren. Nach der Dialyse liegt Vimentin als Tetramer im Pi-Puffer vor, was mit Hilfe der analytischen Ultrazentrifuge kontrolliert wurde. Um die Wachstumskinetik zu messen wurde die Salzkonzentration im Pi-Puffer auf 100 mM KCl erhöht. Dadurch beginnen die Vimentin-Tetramere lateral zu assoziieren und es entstehen ”Unit-Length- Filamente” (ULFs) aus acht Tetrameren. Diese ULFs können dann longitudinal zu langen Filamenten zusammenwachsen. ImVerlauf des Längenwachstums findet gleichzeitig eine Kompaktierung des Filaments statt. Diese radiale Kompaktierung konnte in dieser Arbeit nochmals bestätigt werden. Das Längenwachstum per Zeiteinheit wurde zuerst mit Hilfe des Elektronen- und Rasterkraftmikroskops (EM & SFM) untersucht. Im EM wurde das Wachstum nach definierten Zeitschritten durch Zugabe von Glutaraldehyd gestoppt. Im SFM dagegen wurde die Reaktion durch Verdünnen stark verlangsamt und dasWachstum durch Adsorption an eine Oberfläche endgültig beendet. Durch Bildanalyse wurde die Längenverteilung der Filamente auf den resultierenden Bilderserien bestimmt. Zudem wurde der Einfluss der eingesetzten Oberflächen (kohlebeschichtete Kupfernetze im EM sowie Glimmer und Glas im SFM) charakterisiert und die Ergebnisse miteinander mit Hilfe statistischer Methoden verglichen. Dadurch konnte bestätigt werden, dass eine kinetische Messung des Filamentwachstums auf diese Weise möglich ist und dass die Resultate aus den EM und SFM Messungen miteinander verglichen werden dürfen. Da mit Hilfe der Mikroskope insbesondere das lateraleWachstum vom Tetramer hin zum ULF innerhalb der ersten Sekunden nicht aufgelöst werden konnte, wurde diese Phase des Filamentwachstums mit einem ”Stopped-Flow”-Gerät untersucht. Hierbei wurde das Vermischen der Reaktionslösungen mit Hilfe eines Computers gesteuert, wodurch eine Beobachtung der Reaktion nach wenigen Millisekunden möglich war. Der Verlauf der Reaktion wurde mit Hilfe von Laserlichtstreuung gemessen. Zusätzlich wurden verschiedene kinetische Modelle auf Grundlage der bestehenden qualitativen Betrachtungen zum Filamentwachstum entwickelt. Aus diesen Modellen wurde ein Modell ermittelt, das die Messdaten am besten beschreibt. Durch mathematische Anpassung dieses Modells konnte dann eine erste qualitative Beschreibung derWachstumskinetik von Vimentin vorgeschlagen werden, die quantitativen Daten optimal wiedergibt. So konnte an dieser Stelle bestätigt werden, dass ULFs tatsächlich zum Filamentwachstum beitragen müssen und längere Filamente durch End-zu-End Fusion zusammenwachsen. Damit konnte im Rahmen dieser Arbeit eine, im Gegensatz zur Polymerisation von Aktin und Tubulin, neue Form des Filamentwachstums beschrieben werden, bei dem das Wachstum zuerst durch eine laterale Assoziation mit anschließender longitudinalen Verlängerung gekennzeichnet ist

Refractory metal-based ohmic contacts on β-Ga2O3 using TiW

The present work investigates the use of the refractory metal alloy TiW as a possible candidate for the realization of ohmic contacts to the ultrawide bandgap semiconductor β-Ga2O3. Ohmic contact properties were analyzed by transfer length measurements of TiW contacts annealed at temperatures between 400 and 900 °C. Optimum contact properties with a contact resistance down to 1.5 × 10-5 ω cm2 were achieved after annealing at 700 °C in nitrogen on highly doped β-Ga2O3. However, a significant contact resistance increase was observed at annealing temperatures above 700 °C. Cross-sectional analyses of the contacts using scanning transmission electron microscopy revealed the formation of a TiOx interfacial layer of 3-5 nm between TiW and β-Ga2O3. This interlayer features an amorphous structure and most probably possesses a high amount of vacancies and/or Ga impurities supporting charge carrier injection. Upon annealing at temperatures of 900 °C, the interlayer increases in thickness up to 15 nm, featuring crystalline-like properties, suggesting the formation of rutile TiO2. Although severe morphological changes at higher annealing temperatures were also verified by atomic force microscopy, the root cause for the contact resistance increase is attributed to the structural changes in thickness and crystallinity of the interfacial layer

CLEMSite, a software for automated phenotypic screens using light microscopy and FIB-SEM

This work was supported by EMBL funds and by by the Deutsche Forschungsgemeinschaft (DFG, German Research Foundation) – Project number 240245660 – SFB 1129 (project Z2).In recent years, Focused Ion Beam Scanning Electron Microscopy (FIB-SEM) has emerged as a flexible method that enables semi-automated volume ultrastructural imaging. We present a toolset for adherent cells that enables tracking and finding cells, previously identified in light microscopy (LM), in the FIB-SEM, along with the automatic acquisition of high-resolution volume datasets. We detect the underlying grid pattern in both modalities (LM and EM), to identify common reference points. A combination of computer vision techniques enables complete automation of the workflow. This includes setting the coincidence point of both ion and electron beams, automated evaluation of the image quality and constantly tracking the sample position with the microscope’s field of view reducing or even eliminating operator supervision. We show the ability to target the regions of interest in EM within 5 µm accuracy while iterating between different targets and implementing unattended data acquisition. Our results demonstrate that executing volume acquisition in multiple locations autonomously is possible in EM.Publisher PDFPeer reviewe

Photochemistry of Furyl- and Thienyldiazomethanes: Spectroscopic Characterization of Triplet 3-Thienylcarbene

Photolysis (λ \u3e 543 nm) of 3-thienyldiazomethane (1), matrix isolated in Ar or N2 at 10 K, yields triplet 3-thienylcarbene (13) and α-thial-methylenecyclopropene (9). Carbene 13 was characterized by IR, UV/vis, and EPR spectroscopy. The conformational isomers of 3-thienylcarbene (s-E and s-Z) exhibit an unusually large difference in zero-field splitting parameters in the triplet EPR spectrum (|D/hc| = 0.508 cm–1, |E/hc| = 0.0554 cm–1; |D/hc| = 0.579 cm–1, |E/hc| = 0.0315 cm–1). Natural Bond Orbital (NBO) calculations reveal substantially differing spin densities in the 3-thienyl ring at the positions adjacent to the carbene center, which is one factor contributing to the large difference in D values. NBO calculations also reveal a stabilizing interaction between the sp orbital of the carbene carbon in the s-Z rotamer of 13 and the antibonding σ orbital between sulfur and the neighboring carbon—an interaction that is not observed in the s-E rotamer of 13. In contrast to the EPR spectra, the electronic absorption spectra of the rotamers of triplet 3-thienylcarbene (13) are indistinguishable under our experimental conditions. The carbene exhibits a weak electronic absorption in the visible spectrum (λmax = 467 nm) that is characteristic of triplet arylcarbenes. Although studies of 2-thienyldiazomethane (2), 3-furyldiazomethane (3), or 2-furyldiazomethane (4) provided further insight into the photochemical interconversions among C5H4S or C5H4O isomers, these studies did not lead to the spectroscopic detection of the corresponding triplet carbenes (2-thienylcarbene (11), 3-furylcarbene (23), or 2-furylcarbene (22), respectively)

Filamentous Biopolymers on Surfaces: Atomic Force Microscopy Images Compared with Brownian Dynamics Simulation of Filament Deposition

Nanomechanical properties of filamentous biopolymers, such as the persistence length, may be determined from two-dimensional images of molecules immobilized on surfaces. For a single filament in solution, two principal adsorption scenarios are possible. Both scenarios depend primarly on the interaction strength between the filament and the support: i) For interactions in the range of the thermal energy, the filament can freely equilibrate on the surface during adsorption; ii) For interactions much stronger than the thermal energy, the filament will be captured by the surface without having equilibrated. Such a ‘trapping’ mechanism leads to more condensed filament images and hence to a smaller value for the apparent persistence length. To understand the capture mechanism in more detail we have performed Brownian dynamics simulations of relatively short filaments by taking the two extreme scenarios into account. We then compared these ‘ideal’ adsorption scenarios with observed images of immobilized vimentin intermediate filaments on different surfaces. We found a good agreement between the contours of the deposited vimentin filaments on mica (‘ideal’ trapping) and on glass (‘ideal’ equilibrated) with our simulations. Based on these data, we have developed a strategy to reliably extract the persistence length of short worm-like chain fragments or network forming filaments with unknown polymer-surface interactions

Bi-allelic ACBD6 variants lead to a neurodevelopmental syndrome with progressive and complex movement disorders

The acyl-CoA-binding domain-containing protein 6 (ACBD6) is ubiquitously expressed, plays a role in the acylation of lipids and proteins, and regulates the N-myristoylation of proteins via N-myristoyltransferase enzymes (NMTs). However, its precise function in cells is still unclear, as is the consequence of ACBD6 defects on human pathophysiology. Utilizing exome sequencing and extensive international data sharing efforts, we identified 45 affected individuals from 28 unrelated families (consanguinity 93%) with bi-allelic pathogenic, predominantly loss-of-function (18/20) variants in ACBD6. We generated zebrafish and Xenopus tropicalis acbd6 knockouts by CRISPR/Cas9 and characterized the role of ACBD6 on protein N-myristoylation with YnMyr chemical proteomics in the model organisms and human cells, with the latter also being subjected further to ACBD6 peroxisomal localization studies. The affected individuals (23 males and 22 females), with ages ranging from 1 to 50 years old, typically present with a complex and progressive disease involving moderate-to-severe global developmental delay/intellectual disability (100%) with significant expressive language impairment (98%), movement disorders (97%), facial dysmorphism (95%), and mild cerebellar ataxia (85%) associated with gait impairment (94%), limb spasticity/hypertonia (76%), oculomotor (71%) and behavioural abnormalities (65%), overweight (59%), microcephaly (39%) and epilepsy (33%). The most conspicuous and common movement disorder was dystonia (94%), frequently leading to early-onset progressive postural deformities (97%), limb dystonia (55%), and cervical dystonia (31%). A jerky tremor in the upper limbs (63%), a mild head tremor (59%), parkinsonism/hypokinesia developing with advancing age (32%), and simple motor and vocal tics were among other frequent movement disorders. Midline brain malformations including corpus callosum abnormalities (70%), hypoplasia/agenesis of the anterior commissure (66%), short midbrain and small inferior cerebellar vermis (38% each), as well as hypertrophy of the clava (24%) were common neuroimaging findings. acbd6-deficient zebrafish and Xenopus models effectively recapitulated many clinical phenotypes reported in patients including movement disorders, progressive neuromotor impairment, seizures, microcephaly, craniofacial dysmorphism, and midbrain defects accompanied by developmental delay with increased mortality over time. Unlike ACBD5, ACBD6 did not show a peroxisomal localisation and ACBD6-deficiency was not associated with altered peroxisomal parameters in patient fibroblasts. Significant differences in YnMyr-labelling were observed for 68 co- and 18 post-translationally N-myristoylated proteins in patient-derived fibroblasts. N-Myristoylation was similarly affected in acbd6-deficient zebrafish and Xenopus tropicalis models, including Fus, Marcks, and Chchd-related proteins implicated in neurological diseases. The present study provides evidence that bi-allelic pathogenic variants in ACBD6 lead to a distinct neurodevelopmental syndrome accompanied by complex and progressive cognitive and movement disorders