Exploration of surfactin production by newly isolated Bacillus and Lysinibacillus strains from food-related sources.

As a lipopeptide (LP), surfactin exhibits properties, such as emulsifying and dispersing ability, which are useful in food industry. Discovery of new LP-producing strains from food sources is an important step towards possible application of surfactin in foods. A total of 211 spore-forming, Gram-positive, and catalase-positive bacterial strains were isolated from fermented African locust beans (iru) and palm oil mill effluents in a screening process and examined for their ability to produce surfactin. This was achieved by a combination of methods, which included microbiological and molecular classification of strains, along with chemical analysis of surfactin production. Altogether, 29 isolates, positive for oil spreading and emulsification assays, were further identified with 16S rDNA analysis. The strains belonged to nine species including less commonly reported strains of Lysinibacillus, Bacillus flexus, B. tequilensis, and B. aryabhattai. The surfactin production was quantitatively and qualitatively analysed by high-performance thin-layer chromatography and liquid chromatography-mass spectrometry (LC-MS). Confirmation of surfactin by MS was achieved in all the 29 strains. Highest surfactin production capability was found in B. subtilis IRB2-A1 with a titre of 1444·1 mg L-1 .

Assessment of Bone and Muscle Measurements by Peripheral Quantitative Computed Tomography in Geriatric Patients.

The loss of bone and muscle mass increases the risk of osteoporotic fractures. Dual energy X-ray absorptiometry (DXA) loses sensitivity in older age. The purpose of this study was to evaluate bone and muscle measurements of peripheral quantitative computed tomography (pQCT) in a geriatric cohort with osteoporosis. Bone mineral density and muscle area of 168 patients aged 65 years and older (76.3 ± 6.5) were measured with pQCT at distal forearm additionally to an osteoporosis assessment consisting of anamnesis, blood test and DXA of lumbar spine and hip. Prior fractures were categorized in minor and major osteoporotic fractures. Logistic regression was used to show the association of bone mineral density and muscle area with major fractures. 54.8% of the participants had at least one major fracture. Bone mineral density measured with pQCT and muscle area were significantly associated with these fractures (total and trabecular bone mineral density OR 2.243 and 2.195, p < 0.01; muscle area OR 2.378, p < 0.05), whereas DXA bone mineral density showed no significant association. These associations remained after adjustment for age, sex, BMI, physical activity and other factors. In all models for patients >75 years only muscle area was significantly associated (OR 5.354, p < 0.05) with major fractures. Measurement of bone mineral density and muscle area with pQCT seems to have advantage over DXA in fracture association in geriatric patients. Measuring muscle area also adds useful information to estimate the presence of osteosarcopenia.

A new multi-channel Mach probe measuring the radial ion flow velocity profile in the boundary plasma of the W7-X stellarator.

Ion flow velocity measurement in the edge and scraper-off layer region is beneficial to understand the confinement related phenomenon in fusion devices such as impurity transport and plays an important role in impurity control. During the Wendelstein 7-X (W7-X) operation phase 1.2a, a multi-channel (MC) Mach probe mounted on the multi-purpose manipulator has been used to measure radial profiles of edge ion flow velocity. This MC-Mach probe consists of two polar and two radial arrays of directional Langmuir pins (28 pins in total) serving for different aims, of which the polar arrays could obtain a polar distribution of ion saturation current, while the radial arrays can be used to study the dynamic process of a radially propagated event. In this paper, we report the observation of the radially outward propagation of a low frequency mode with a speed of around 200 m/s. The first measurement of the radial ion flow velocity profile using the MC-Mach probe in the boundary plasma of the W7-X with an island divertor will also be presented.

Relatedness and heterogeneity at the near-terminal end of the genome of a parapoxvirus bovis 1 strain (B177) compared with parapoxvirus ovis (Orf virus).

The present study provides for the first time an extended investigation of individual genes located at the near-terminal right end of the genome of parapoxvirus bovis 1, Bovine papular stomatitis virus (BPSV) strain B177 and Orf virus (ORFV). Comparison of the respective DNA sequences of ORFV strain D1701 (9.9 kbp) and BPSV B177 (7.7 kbp) revealed a very similar organization of closely related genes transcribed in a rightward orientation. The most salient findings of this study were: (i) the absence of the ORFV-specific vascular endothelial growth factor (VEGF-E) gene in the BPSV isolate; (ii) the presence of an interleukin-10 (IL-10) orthologue; and (iii) the detection of three new genes encoding ankyrin-repeat-containing polypeptides. These results not only contribute to potential improvements of future molecular differentiation between the parapoxvirus species, but also shed new light on different pathobiologies among parapoxviruses.

Anisotropic scaling and generalized conformal invariance at Lifshitz points.

The behaviour of the 3D axial next-nearest-neighbor Ising model at the uniaxial Lifshitz point is studied using Monte Carlo techniques. A new variant of the Wolff cluster algorithm permits the analysis of systems far larger than in previous studies. The Lifshitz point critical exponents are alpha = 0.18(2), beta = 0.238(5), and gamma = 1.36(3). Data for the spin-spin correlation function are shown to be consistent with the explicit scaling function derived from the assumption of local scale invariance, which is a generalization of conformal invariance to the anisotropic scaling at the Lifshitz point.

Direct observation of membrane retrieval in chromaffin cells by capacitance measurements.

This study was focussed on the identification of the endocytic organelles in chromaffin cells which retrieve large, dense core vesicle (LDCV)-membrane components from the plasma membrane. For this purpose, 'on-cell' capacitance measurements and electron microscopy were employed. We found capacitance steps and capacitance flickers, corresponding to single exo- and endocytic events. The analysis revealed that the total membrane surface of completely fused LDCVs is recycled by large endocytic vesicles and smaller, most likely clathrin-coated vesicles, at approximately the same ratio. These results were confirmed by rapid-freeze immuno-electron microscopy, where an extracellular marker was rapidly internalized into endocytic vesicles that morphologically resembled LDCVs.

Critical properties of the reaction-diffusion model 2A-->3A, 2A-->0.

The steady-state phase diagram of the one-dimensional reaction-diffusion model 2A-->3A, 2A-->0 is studied through the non-Hermitian density matrix renormalization group. In the absence of single-particle diffusion the model reduces to the pair-contact process, which has a phase transition in the universality class of directed percolation (DP) and an infinite number of absorbing steady states. When single-particle diffusion is added, the number of absorbing steady states is reduced to 2 and the model no longer shows DP critical behavior. The exponents theta=nu(parallel)/nu(perpendicular) and beta/nu(perpendicular) are calculated numerically. The value of beta/nu(perpendicular) is close to the value of the parity conserving universality class, in spite of the absence of local conservation laws.

Aging, phase ordering, and conformal invariance.

In a variety of systems which exhibit aging, the two-time response function scales as R(t,s) approximately s(-1-a)f(t/s). We argue that dynamical scaling can be extended towards conformal invariance, thus obtaining the explicit form of the scaling function f. This quantitative prediction is confirmed in several spin systems, both for T

Generation of recombinant parapoxviruses: non-essential genes suitable for insertion and expression of foreign genes.

Orf virus (OV) is an epitheliotropic poxvirus and belongs to the genus Parapoxvirus (PPV). PPV, especially OV, is regarded as a promising candidate for an expression vector. Among available live vaccines only strain D1701 represents a highly attenuated OV strain with clearly reduced pathogenicity. Therefore, we started to identify potentially non-essential genes or regions of D1701, which might be suitable for insertion and expression of foreign genes. The present contribution reviews some of the progress using the vegf-e (homologue of the mammalian vascular endothelial growth factor) gene locus for the generation of recombinant D1701. The vegf-e gene of D1701 is dispensable for virus growth in vitro and in vivo, and represents a major virulence determinant of OV. It is shown that foreign genes can be inserted and functionally expressed in the vegf-e locus, also leading to the induction of a specific immune response in the non-permissive host. Furthermore, it is reported that adaptation to VERO cells led to the deletion of three further regions of the OV D1701 genome, which seems to be combined with additional virus attenuation in sheep. Molecular analysis of this OV D1701 variant allows the identification of new, potentially non-essential sites in the viral genome.

A gelatin sponge implantation model in swine for the determination of local immune reactions.

For in vivo determination of innate and memory immune reactions we have implanted sterile gelatin sponges subcutaneously in swine for the monitoring of the following immunological parameters: 1. Analysis of local cell population phenotypes after vascularization of the gelatin sponges using flow cytometry. 2. Comparative analysis of sponge-infiltrating cells after loading with viral antigen in primed as well as naive animals. 3. Performance of reverse transcription quantitative competitive PCR (RT-qcPCR) for the detection of porcine cytokine mRNA indicative for IFN-gamma, IL-2, IL-4, IL-8 and IL-10. The in vitro analysis, e.g. by re-exposure to viral antigens, allows the determination of cytokine reaction patterns of sponge derived cells, draining lymph node cells as well as PBMC of the same individual. Studies of innate reactions and modulation of cellular infiltration in transplanted gelatin sponges are possible. Functional assays, e.g. cell-mediated cytotoxicity, antigen specific cell proliferation, using sponge-derived cells will provide additional information about the suitability of the model for example in vaccine potency tests.

Parapoxviruses: potential alternative vectors for directing the immune response in permissive and non-permissive hosts.

Parapoxvirus (PPV) represents a genus of the poxviridae, and particularly PPV ovis (Orf virus, OV) seems to offer several potential advantages for the use of vector vaccine. Therefore, we started to investigate the genome of the highly attenuated OV strain D1701, which was only poorly characterised until now. Due to recombination of non-homologous sequences, part of the right hand end of the D1701 genome was duplicated and translocated to the opposite end of the genome. As a consequence gene deletion had occurred and the inverted terminal repeat region is increased. Results are described to identify viral genes, which are non-essential for virus replication and potentially influence viral pathogenesis, virulence, and host immunity. In more detail, we analysed the expression and functional activity of the OV-specific vascular endothelial growth factor (VEGF) gene homologue. Finally the construction and production of a D1701 mutant lacking the VEGF gene homologue is reported.

Endocytic delivery of intramolecularly quenched substrates and inhibitors to the intracellular yeast Kex2 protease1.

Kex2 in the yeast Saccharomyces cerevisiae is a transmembrane, Ca2+-dependent serine protease of the subtilisin-like pro-protein convertase (SPC) family with specificity for cleavage after paired basic amino acids. At steady state, Kex2 is predominantly localized in late Golgi compartments and initiates the proteolytic maturation of pro-protein precursors that transit the distal secretory pathway. However, Kex2 localization is not static, and its itinerary apparently involves transiting out of the late Golgi and cycling back from post-Golgi endosomal compartments during its lifetime. We tested whether the endocytic pathway could deliver small molecules to Kex2 from the extracellular medium. Here we report that intramolecularly quenched fluorogenic substrates taken up into intact yeast revealed fluorescence due to specific cleavage by Kex2 protease in endosomal compartments. Furthermore, the endocytic delivery of protease inhibitors interfered with Kex2 activity for precursor protein processing. These observations reveal that the endocytic pathway does intersect with the cycling itinerary of active Kex2 protease. This strategy of endocytic drug delivery has implications for modulating SPC protease activity needed for hormone, toxin and viral glycoprotein precursor processing in human cells.

Analysis of genomic rearrangement and subsequent gene deletion of the attenuated Orf virus strain D1701.

The orf virus (OV) strain D1701 belongs to the genetically heterogenous parapoxvirus (PPV) genus of the family Poxviridae. The attenuated OV D1701 has been licensed as a live vaccine against contagious ecthyma in sheep. Detailed knowledge on the genetic structure and organization of this PPV vaccine strain is an important prerequisite to reveal possible genetic mechanisms of PPV attenuation. The present study demonstrates a genomic map of the approximately 158 kbp DNA of OV D1701 established by hybridization studies of cloned restriction fragments covering the complete viral genome. The results show an enlargement of the inverted terminal repeats (ITR) to up to 18 kbp due to recombination between nonhomologous sequences during cell culture adaptation. DNA sequencing of the region adjacent to the ITR junction revealed the absence of one open reading frame designated E2L. In contrast to a transposition-deletion variant of the New Zealand OV strain NZ2 (Fleming et al., 1995) the two genes E3L (a homologue of dUTPase) and G1L neighbouring E2L are retained in OV D1701. DNA and RNA analyses proved the presence of E2L gene in wild-type OV isolated directly from scab material. The data presented indicate that the E2L gene is nonessential for virus replication in vitro and in vivo, and may represent one important viral gene in determining virulence and pathogenesis of OV.

[Surgical management of complications of open pelvic injuries]. / Chirurgisches Management der Komplikationen offener Beckenverletzungen.

Open pelvic fractures are rare fractures usually resulting from a severe trauma. Only 4 of 121 patients treated operatively in 1994 for a pelvic trauma showed an open injury. The high lethality is caused by two complications: in the early phase the patient is threatened by bleeding to death and in the following course sepsis determines the lethal outcome. Only an aggressive surgical management prevents or treats such complications. The presented concept comprises in the end the hemipelvectomy and an intensive care management for prophylaxis of septic complications. So the lethality was diminished.

[Correlation between pulmonary hypertension and portal hypertension--2 case reports of different forms of pre-sinusoidal portal hypertension]. / Zum Zusammenhang zwischen pulmonaler Hypertonie und portaler Hypertension--zwei Kasuistiken von unterschiedlichen Formen von präsinusoidaler portaler Hypertension.

Since its first description in 1951 by Mantz and Craig pulmonary hypertension in combination with portal hypertension has been observed more and more frequently. In a recent prospective study Hadengue et al. reported an incidence of 2%. Thus this simultaneous occurrence can no longer be considered to be coincidental. The etiology remains still unclear. It is most probable that the development is due to vasoactive substances which bypass the liver or which are produced in the lung itself, and which, due to a long-term vasoconstriction, causes irreparable damage to the arterioles and arteries in the lung. Such pulmonary hypertension can develop in the presence of a pre- as well as an intrahepatic block, even when the portal hypertension is partially or completely alleviated by a portosystemic anastomosis. This last circumstance can be illustrated by two cases which were observed by our group. Case A is of particular interest because it is the first documentation of a case of an intrahepatic block due to a (so-called) macronodular transformation of the liver in the absence of portal thrombosis (a so-called NRH: nodular regenerative hyperplasia) in combination with pulmonary hypertension. This type of non-cirrhotic portal hypertension can be associated with micronodular transformation of the liver as well. Post-hepatic blocks or the so-called BUDD-CHIARI Syndrome type appear to carry no risk of development of pulmonary hypertension. It remains unclear which particular etiologies increase susceptibility to later development of pulmonary hypertension.