A single nucleotide exchange in the wzy gene is responsible for the semirough O6 lipopolysaccharide phenotype and serum sensitivity of Escherichia coli strain Nissle 1917

Structural analysis of lipopolysaccharide (LPS) isolated from semirough, serum-sensitive Escherichia coli strain Nissle 1917 (DSM 6601, serotype O6:K5:H1) revealed that this strain's LPS contains a bisphosphorylated hexaacyl lipid A and a tetradecasaccharide consisting of one E. coli O6 antigen repeating unit attached to the R1-type core. Configuration of the GlcNAc glycosidic linkage between O-antigen oligosaccharide and core (beta) differs from that interlinking the repeating units in the E. coli O6 antigen polysaccharide (alpha). The wa(*) and wb(*) gene clusters of strain Nissle 1917, required for LPS core and O6 repeating unit biosyntheses, were subcloned and sequenced. The DNA sequence of the wa(*) determinant (11.8 kb) shows 97% identity to other R1 core type-specific wa(*) gene clusters. The DNA sequence of the wb(*) gene cluster (11 kb) exhibits no homology to known DNA sequences except manC and manB. Comparison of the genetic structures of the wb(*)(O6) (wb(*) from serotype O6) determinants of strain Nissle 1917 and of smooth and serum-resistant uropathogenic E. coli O6 strain 536 demonstrated that the putative open reading frame encoding the O-antigen polymerase Wzy of strain Nissle 1917 was truncated due to a point mutation. Complementation with a functional wzy copy of E. coli strain 536 confirmed that the semirough phenotype of strain Nissle 1917 is due to the nonfunctional wzy gene. Expression of a functional wzy gene in E. coli strain Nissle 1917 increased its ability to withstand antibacterial defense mechanisms of blood serum. These results underline the importance of LPS for serum resistance or sensitivity of E. coli.     

PAF-mediated pulmonary edema: a new role for acid sphingomyelinase and ceramide

Platelet-activating factor (PAF) induces pulmonary edema and has a key role in acute lung injury (ALI). Here we show that PAF induces pulmonary edema through two mechanisms: acid sphingomyelinase (ASM)-dependent production of ceramide, and activation of the cyclooxygenase pathway. Agents that interfere with PAF-induced ceramide synthesis, such as steroids or the xanthogenate D609, attenuate pulmonary edema formation induced by PAF, endotoxin or acid instillation. Our results identify acid sphingomyelinase and ceramide as possible therapeutic targets in acute lung injury.     

In vitro models for metabolic studies of small peptide hormones in sport drug testing

Peptide hormones represent an emerging class of potential doping agents. Detection of their misuse is difficult due to their short half‐life in plasma and rapid elimination. Therefore, investigating their metabolism can improve detectability. Unfortunately, pharmacokinetic studies with human volunteers are often not allowed because of ethical constraints, and therefore alternative models are needed. This study was performed in order to evaluate in vitro models (human liver microsomes and S9 fraction) for the prediction of the metabolism of peptidic doping agents and to compare them with the established models. The peptides that were investigated include desmopressin, TB‐500, GHRP‐2, GHRP‐6, hexarelin, LHRH and leuprolide. Several metabolites were detected for each peptide after incubation with human liver microsomes, S9 fraction, and serum, which all showed endopeptidase and exopeptidase activity. In vitro models from different organs (liver vs. kidney) were compared, but no significant differences were recorded. Deamidation was not observed in any of the models and was therefore evaluated by incubation with α‐chymotrypsin. In conclusion, in vitro models are useful tools for forensic and clinical analysts to detect peptidic metabolic markers in biological fluids. Copyright © 2014 European Peptide Society and John Wiley & Sons, Ltd.     

Energy potential for combustion and anaerobic digestion of biomass from low-input high-diversity systems in conservation areas

In this study, we assessed the potential for bioenergy production of Low-Input High-Diversity (LIHD) systems in temperate West-European conservation areas. A wide range of seminatural ecosystems (wet and dry grasslands, marshes, tall-herb vegetation and heathlands) was sampled. Because LIHD biomass is often scattered and discontinuously available, we only considered the potential for anaerobic digestion and combustion. Both technologies are suitable for decentralized biomass utilization. The gross energy yield showed a promising range between 46–277  GJ per hectare per mowing cycle (MC). The energy efficiency of the anaerobic digestion process was rather low (10–30%) with a methane energy yield of 5.5–35.5 GJ ha⁻¹ MC⁻¹, experimentally determined by batch digestion tests. The water content, functional group composition and biochemical composition (hemicellulose, cellulose, lignin and Kjeldahl nitrogen) of the biomass were analyzed to assess the suitability of different valorization pathways. On the basis of the results, we were able to propose recommendations regarding the appropriate conversion techniques. Biomass from plant communities with ‘late’ harvest dates (August–October) or a high fraction of woody species like heathland and dune slacks, is best valorized through combustion, while herbaceous biomass of ‘early’ harvested grasslands (June–July) and tall-herb vegetation can better be digested. The main advantages of the production of bioenergy from LIHD biomass originating from conservation management are the minimization of the competition with food production and its potential to reconcile renewable energy policies and biodiversity goals.     

Das Verhalten von Extremitätenregeneraten des weissen und pigmentierten Axolotl bei heteroplastischer,heterotoper und orthotoper Transplantation und sukzessiver Regeneration

Ohne Zusammenfassung     

Atkinesin-13A Modulates Cell-Wall Synthesis and Cell Expansion in Arabidopsis thaliana via the THESEUS1 Pathway

Growth of plant organs relies on cell proliferation and expansion. While an increasingly detailed picture about the control of cell proliferation is emerging, our knowledge about the control of cell expansion remains more limited. We demonstrate here that the internal-motor kinesin AtKINESIN-13A (AtKIN13A) limits cell expansion and cell size in Arabidopsis thaliana, with loss-of-function atkin13a mutants forming larger petals with larger cells. The homolog, AtKINESIN-13B, also affects cell expansion and double mutants display growth, gametophytic and early embryonic defects, indicating a redundant role of the two genes. AtKIN13A is known to depolymerize microtubules and influence Golgi motility and distribution. Consistent with this function, AtKIN13A interacts genetically with ANGUSTIFOLIA, encoding a regulator of Golgi dynamics. Reduced AtKIN13A activity alters cell wall structure as assessed by Fourier-transformed infrared-spectroscopy and triggers signalling via the THESEUS1-dependent cell-wall integrity pathway, which in turn promotes the excess cell expansion in the atkin13a mutant. Thus, our results indicate that the intracellular activity of AtKIN13A regulates cell expansion and wall architecture via THESEUS1, providing a compelling case of interplay between cell wall integrity sensing and expansion.     

Der Einfluss mechanisch veränderter Augenstellungen auf die Richtungslokalisation bei Fischen

Zusammenfassung Wenn man Fischen die Augen um eine der Körperlängsachse parallele Achse mechanisch verstellt, so lokalisieren sie die Richtung des Lichteinfalls um den gleichen Winkelbetrag falsch; der äußere Einfluß auf das Auge wird also bei der Richtungslokalisation nicht einberechnet. Das gilt unabhängig davon, wie die Spannungsverhältnisse der Augenmuskeln sich bei dieser aufgezwungenen Augenstellung ändern.Anzeichen für eine allmähliche Kompensation dieser Fehllokalisation wurden nicht beobachtet. Dagegen wird ein Kompensationsvorgang, den die Ausschaltung eines Auges in Gang setzt, von der Lage, die man dem anderen Auge aufzwingt, stark beeinflußt.Es wird gezeigt, daß die Befunde durch das Reafferenzprinzip (v. Holst und Mittelstaedt) physiologisch verständlich werden. Otto Hahn zum 75. Geburtstag.     

Phenotyping of Partial Physiological Resistance to Rice Sheath Blight

Sheath blight, caused by Rhizoctonia solani, is one of the most important rice diseases worldwide especially under irrigated agro‐ecosystems. To date, no rice accession with complete resistance to sheath blight has been reported. However, a number of genotypes with varying levels of resistance have been reported. Twelve genotypes (including mega varieties) viz. Tetep, Jasmine 85, Te‐Qing, Duduruchi, Betichikon, Khatochalani, D‐6766, D‐256, Swarna, Sarju‐52, MTU‐1010 and Samba Mashuri were evaluated for quantitative measurement of partial physiological resistance to sheath blight under controlled conditions using detached tiller method. Three independent experiments, each involving three replications, were conducted. Seven days after inoculation, the following disease variables were measured: number of lesions, lesion length, vertical sheath colonization (VSC) on the tiller, disease severity, relative vertical sheath colonization (RVSC) and survival of the leaf blade. Variation between rice genotypes was observed for all the disease variables. Disease severity and VSC were the two most correlated variables, whereas the number of lesions and mean lesion length were the least correlated variables. The ranking of varieties often differed depending on the disease variable considered. Amongst the genotypes tested, D‐256, Tetep and Jasmin‐85 had the lowest number of lesions and disease severity. Similarly, Tetep and D‐256 showed the lowest levels of RVSC, whilst Jasmine‐85 was found to be intermediate. D‐6766, Samba Mashuri and Betichikon showed the highest levels of disease variables. The fraction of dead leaves ranged from 0.00 to 0.38. No dead leaves were observed in Te‐Qing, Swarna and MTU‐1010. The highest fraction of dead leaves was observed for Betichikon (0.38) followed by Duduruchi and D‐6766 (0.33). Our results suggest that this method in combination with other phenotyping methods could be used to quantify partial resistance to rice sheath blight.     

The In ovo CAM-assay as a Xenograft Model for Sarcoma

Sarcoma is a very rare disease that is heterogeneous in nature, all hampering the development of new therapies. Sarcoma patients are ideal candidates for personalized medicine after stratification, explaining the current interest in developing a reproducible and low-cost xenotransplant model for this disease. The chick chorioallantoic membrane is a natural immunodeficient host capable of sustaining grafted tissues and cells without species-specific restrictions. In addition, it is easily accessed, manipulated and imaged using optical and fluorescence stereomicroscopy. Histology further allows detailed analysis of heterotypic cellular interactions.This protocol describes in detail the in ovo grafting of the chorioallantoic membrane with fresh sarcoma-derived tumor tissues, their single cell suspensions, and permanent and transient fluorescently labeled established sarcoma cell lines (Saos-2 and SW1353). The chick survival rates are up to 75%. The model is used to study graft- (viability, Ki67 proliferation index, necrosis, infiltration) and host (fibroblast infiltration, vascular ingrowth) behavior. For localized grafting of single cell suspensions, ECM gel provides significant advantages over inert containment materials. The Ki67 proliferation index is related to the distance of the cells from the surface of the CAM and the duration of application on the CAM, the latter determining a time frame for the addition of therapeutic products.