Efficient preparation of Arabidopsis pollen tubes for ultrastructural analysis using chemical and cryo-fixation

The pollen tube (PT) serves as a model system for investigating plant cell growth and morphogenesis. Ultrastructural studies are indispensable to complement data from physiological and genetic analyses, yet an effective method is lacking for PTs of the model plant Arabidopsis thaliana. Methods: Here, we present reliable approaches for ultrastructural studies of Arabidopsis PTs, as well as an efficient technique for immunogold detection of cell wall epitopes. Using different fixation and embedding strategies, we show the amount of PT ultrastructural details that can be obtained by the different methods. Results: Dozens of cross-sections can be obtained simultaneously by the approach, which facilitates and shortens the time for evaluation. In addition to in vitro-grown PTs, our study follows the route of PTs from germination, growth along the pistil, to the penetration of the dense stylar tissue, which requires considerable mechanical forces. To this end, PTs have different strategies from growing between cells but also between the protoplast and the cell wall and even within each other, where they share a partly common cell wall. The separation of PT cell walls in an outer and an inner layer reported for many plant species is less clear in Arabidopsis PTs, where these cell wall substructures are connected by a distinct transition zone. Conclusions: The major advancement of this method is the effective production of a large number of longitudinal and cross-sections that permits obtaining a detailed and representative picture of pollen tube structures in an unprecedented way. This is particularly important when comparing PTs of wild type and mutants to identify even subtle alterations in cytoarchitecture. Arabidopsis is an excellent plant for genetic manipulation, yet the PTs, several-times smaller compared to tobacco or lily, represent a technical challenge. This study reveals a method to overcome this problem and make Arabidopsis PTs more amenable to a combination of genetic and ultrastructural analyses

Two Distinct Coagulase-Dependent Barriers Protect Staphylococcus aureus from Neutrophils in a Three Dimensional in vitro Infection Model

Staphylococcus aureus is a pyogenic abscess-forming facultative pathogenic microorganism expressing a large set of virulence-associated factors. Among these, secreted proteins with binding capacity to plasma proteins (e.g. fibrinogen binding proteins Eap and Emp) and prothrombin activators such as Coagulase (Coa) and vWbp are involved in abscess formation. By using a three-dimensional collagen gel (3D-CoG) supplemented with fibrinogen (Fib) we studied the growth behavior of S. aureus strain Newman and a set of mutants as well as their interaction with mouse neutrophils by real-time confocal microscopy. In 3D-CoG/Fib, S. aureus forms microcolonies which are surrounded by an inner pseudocapsule and an extended outer dense microcolony-associated meshwork (MAM) containing fibrin. Coa is involved in formation of the pseudocapsule whereas MAM formation depends on vWbp. Moreover, agr-dependent dispersal of late stage microcolonies could be observed. Furthermore, we demonstrate that the pseudocapsule and the MAM act as mechanical barriers against neutrophils attracted to the microcolony. The thrombin inhibitor argatroban is able to prevent formation of both pseudocapsule and MAM and supports access of neutrophils to staphylococci. Taken together, this model can simulate specific stages of S. aureus abscess formation by temporal dissection of bacterial growth and recruitment of immune cells. It can complement established animal infection models in the development of new treatment options

En route to automated maintenance of industrial printing systems: digital quantification of print-quality factors based on induced printing failure

Tracking and tracing are a key technology for production process optimization and subsequent cost reduction. However, several industrial environments (e.g. high temperatures in metal processing) are challenging for most part-marking and identification approaches. A method for printing individual part markings on metal components (e.g. data matrix codes (DMCs) or similar identifiers) with high temperatures and chemical resistance has been developed based on drop-on-demand (DOD) print technology and special ink dispersions with submicrometer-sized ceramic and glass particles. Both ink and printer are required to work highly reliably without nozzle clogging or other failures to prevent interruptions of the production process in which the printing technology is used. This is especially challenging for the pigmented inks applied here. To perform long-term tests with different ink formulations and to assess print quality over time, we set up a test bench for inkjet printing systems. We present a novel approach for monitoring the printhead’s state as well as the print-quality degradation. This method does not require measuring and monitoring, e.g. electrical components or drop flight, as it is done in the state of the art and instead uses only the printed result. By digitally quantifying selected quality factors within the printed result and evaluating their progression over time, several non-stationary measurands were identified. Some of these measurands show a monotonic trend and, hence, can be used to measure print-quality degradation. These results are a promising basis for automated printing system maintenance

Hydrogel-based plasmonic sensor substrate for the detection of ethanol

The in-line monitoring of ethanol concentration in liquids is a crucial part of process monitoring in breweries and distilleries. Current methods are based on infrared spectroscopy, which is time-consuming and costly, making these methods unaffordable for small and middle-sized companies. To overcome these problems, we presented a small, compact, and cost-effective sensing method for the ethanol content, based on a nanostructured, plasmonically active sensor substrate. The sensor substrate is coated with an ethanol-sensitive hydrogel, based on polyacrylamide and bisacrylamide, which induces a change in the refractive index of the substrate surface. The swelling and shrinking of such hydrogels offer a means to measure the ethanol content in liquids, which can be determined in a simple transmittance setup. In our study, we demonstrated the capability of the sensor principle for the detection of ethanol content ranging from 0 to 30 vol% ethanol. Furthermore, we determined the response time of the sensor substrate to be 5.2 min, which shows an improvement by a factor of four compared to other hydrogel-based sensing methods. Finally, initial results for the sensor’s lifetime are presented

Fast response hydrogel-based plasmonic sensor substrate for the detection of ethanol

The inline monitoring of ethanol concentrations in liquids is a crucial part of process monitoring in breweries and distilleries. Current methods are based on infrared spectroscopy which are bulky and costly making them non-affordable for small and middle-sized companies. To overcome these problems, we present a small, compact and cost-effective sensing method, based on a nanostructured, plasmonically active sensor substrate. The sensor substrate is coated with a microstructured ethanol-sensitive acrylamide-bisacrylamide hydrogel which induces a change of the hydrogel’s refractive index in conjugation with the hydrogel swelling and shrinking. With such an approach, the ethanol concentration in liquids can be determined in a simple optical transmittance setup. In our study, we demonstrate the capability of the sensor principle for the detection of ethanol concentration ranging from 0 to 30 vol%. Furthermore, we determined the response time of the sensor substrate to be less than 10 seconds, which shows an enormous improvement compared to other hydrogel-based sensing methods. Finally, initial results for real sample measurements are presented

EU-Aktionsplan zur Unternehmensbesteuerung: Ein Weg zu größerer Fairness und Effizienz in der Steuerpolitik?

Die EU-Kommission hat einen Aktionsplan zur grundlegenden Reform der Unternehmensbesteuerung vorgelegt, der missbräuchlicher Steuergestaltung entgegenwirken und nachhaltige Einnahmen gewährleisten soll. Die Maßnahmen sollen die steuerlichen Rahmenbedingungen für Unternehmen in Europa gerechter, effizienter und wachstumsfreundlicher gestalten. Für Wolfgang Schön, Max-Planck-Institut für Steuerrecht und Öffentliche Finanzen, München, und Ludwig-Maximilians-Universität München, enthält der Aktionsplan wenige Positionen, die nicht bereits vorher diskutiert oder angekündigt waren. Der Gesamttext mache jedoch deutlich, wie stark sich die Kommission von ihren ursprünglichen Zielen europäischer Steuergesetzgebung entfernt habe. Dennis Klein, Leibniz-Fachhochschule Hannover, warnt vor zu hohen Erwartungen an den Aktionsplan. Er sei wenig mehr als bloßer Aktionismus der EU-Kommission. Für Markus Kerber, Bundesverband der Deutschen Industrie, ist es nachvollziehbar, dass die Kommission verstärkt gegen Steuervermeidung vorgehen möchte. Es dürfe jedoch nicht bei einer vordergründigen Diskussion bleiben: Im Zentrum des EU-Projekts stand zu Beginn das Ziel der steuerpolitischen Flankierung des europäischen Binnenmarktes. Eine Verschiebung des Brennpunktes auf die Vermeidung von Steuergestaltungen vermag, seiner Meinung nach, ein solch umfangreiches Projekt nicht zu tragen. Der Fokus sollte auf die Harmonisierung gerichtet sein, um die Attraktivität des europäischen Wirtschaftsstandorts zu erhöhen. Martin Ruf und Tanja Kroh, Universität Tübingen, sehen in dem EU-Aktionsplan nur punktuelle Lösungsansätze für das altbekannte Problem der »Steuerplanung unter Ausnutzung des internationalen Steuersatzgefälles«. Nach Ansicht von Katharina Finke, Universität Mannheim und Zentrum für Europäische Wirtschaftsforschung (ZEW), Mannheim, und Christoph Spengel, Universität Mannheim, gehen von dem Aktionsplan neue Impulse zur Weiterentwicklung der Unternehmensbesteuerung in Europa aus. Die EU-Ko

Geographic Atrophy and Foveal-Sparing Changes Related to Visual Acuity in Patients With Dry Age-Related Macular Degeneration Over Time

Purpose To correlate the area of geographic atrophy (GA) and residual foveal sparing (FS), and to identify the minimum FS and maximum GA area allowing sufficient visual acuity (VA) for daily tasks. Design Prospective cohort study. Methods Thirty-six eyes of 25 patients with GA and FS were followed for 18 months using spectral-domain optical coherence tomography and VA tests. Volume scans were imported into software enabling grading of areas in B-scans and computing of planimetric measurements in complete volume scans. Correlation of areas 1 (complete atrophy), 2 (FS in the central millimeter), and 3 (FS in the central 3 mm) with each other and with best-corrected VA (BCVA) were evaluated. Results Baseline means of areas 1, 2, and 3 were 6.15 mm, 0.49 mm, and 3.08 mm, respectively. At 1 year, area 1 increased by a mean of 1.33 mm, while areas 2 and 3 were decreased by 0.12 mm and 0.65 mm, respectively. From baseline to 18 months and from visit to visit, all areas and BCVA changed progressively (P < .001). Significant thresholds in GA size and FS for achieving a BCVA 70 ETDRS letters were detected (area 1: 6 mm; area 2: 0.48 mm; and area 3: 3.28 mm). Conclusion GA and FS changed inversely over time. In general, FS highly correlated with BCVA, while GA progression correlated with the central 3-mm FS regression, but not with BCVA. A threshold in GA and FS area could be determined for BCVA necessary for daily activity.(VLID)485261