Internal Transport Barrier Broadening through Subdominant Mode Stabilization in Reversed Field Pinch Plasmas.

The reversed field pinch (RFP) device RFX-mod features strong internal transport barriers when the plasma accesses states with a single dominant helicity. Such transport barriers enclose a hot helical region with high confinement whose amplitude may vary from a tiny one to an amplitude encompassing an appreciable fraction of the available volume. The transition from narrow to wide thermal structures has been ascribed so far to the transport reduction that occurs when the dominant mode separatrix, which is a preferred location for the onset of stochastic field lines, disappears. In this Letter we show instead that the contribution from the separatrix disappearance, by itself, is marginal and the main role is instead played by the progressive stabilization of secondary modes. The position and the width of the stochastic boundary encompassing the thermal structures have been estimated by applying the concept of a 3D quasiseparatrix layer, developed in solar physics to treat reconnection phenomena without true separatrices and novel to toroidal laboratory plasmas. Considering the favorable scaling of secondary modes with the Lundquist number, these results open promising scenarios for RFP plasmas at temperatures higher than the presently achieved ones, where lower secondary modes and, consequently, larger thermal structures are expected. Furthermore, this first application of the quasiseparatrix layer to a toroidal plasma indicates that such a concept is ubiquitous in magnetic reconnection, independent of the system geometry under investigation.

Persistence and degradation of new ß-lactam antibiotics in the soil and water environment.

The development of new antibiotics with low environmental persistence is of utmost importance in contrasting phenomena of antibiotic resistance. In this study, the persistence of two newly synthesized monocyclic ß-lactam antibiotics: (2R)-1-(methylthio)-4-oxoazetidin-2-yl acetate, P1, and (2R,3R)-3-((1R)-1-(tert-butyldimethylsilanyloxy)ethyl)-1-(methylthio)-4-oxoazetidin-2-yl acetate, P2, has been investigated in water in the pH range 3-9 and in two (calcareous and forest) soils, then compared to amoxicillin, a ß-lactam antibiotic used in human and veterinary medicine. P1 and P2 persistence in water was lower than that of amoxicillin with only a few exceptions. P1 hydrolysis was catalyzed at an acidic pH whereas P2 hydrolysis takes place at both acidic and alkaline pH values. P1 persistence in soils depended mainly on their water potential (t1/2: 35.0-70.7d at wilting point; <1d at field capacity) whereas for P2 it was shorter and unaffected by soil water content (t1/2 0.13-2.5d). Several degradation products were detected in soils at both water potentials, deriving partly from hydrolytic pathways and partly from microbial transformation. The higher LogKow value for P2 compared with P1 seemingly confers P2 with high permeability to microbial membranes regardless of soil water content. P1 and P2 persistence in soils at wilting point was shorter than that of amoxicillin, whereas it had the same extent at field capacity.

Migration kinetics of mineral oil hydrocarbons from recycled paperboard to dry food: monitoring of two real cases.

Mineral oil hydrocarbons present in printing inks and recycled paper migrate from paper-based food packaging to foods primarily through the gas phase. Migration from two commercial products packed in recycled paperboard, i.e. muesli and egg pasta, was monitored up to the end of their shelf life (1 year) to study the influence of time, storage conditions, food packaging structure and temperature. Mineral oil saturated and aromatic hydrocarbons (MOSH and MOAH, respectively), and diisopropyl naphthalenes (DIPN) were monitored using online HPLC-GC/FID. Storage conditions were: free standing, shelved, and packed in transport boxes of corrugated board, to represent domestic, supermarket and warehouse storage, respectively. Migration to food whose packs were kept in transport boxes was the highest, especially after prolonged storage, followed by shelved and free-standing packs. Tested temperatures were representative of refrigeration, room temperature, storage in summer months and accelerated migration testing. Migration was strongly influenced by temperature: for egg pasta directly packed in paperboard, around 30 mg kg⁻¹ of MOSH migrated in 8 months at 20°C, but in only 1 week at 40°C. Muesli was contained into an internal polyethylene bag, which firstly adsorbed hydrocarbons and later released them partly towards the food. Differently, the external polypropylene bag, containing pasta and recycled paper tray, strongly limited the migration towards the atmosphere and gave rise to the highest level of food contamination. Tests at increased temperatures not only accelerated migration, but also widened the migration of hydrocarbons to higher molecular masses, highlighting thus a difficult interpretation of data from accelerated simulation.

Bifurcation to 3D helical magnetic equilibrium in an axisymmetric toroidal device.

We report the first direct measurement of the internal magnetic field structure associated with a 3D helical equilibrium generated spontaneously in the core of an axisymmetric toroidal plasma containment device. Magnetohydrodynamic equilibrium bifurcation occurs in a reversed-field pinch when the innermost resonant magnetic perturbation grows to a large amplitude, reaching up to 8% of the mean field strength. Magnetic topology evolution is determined by measuring the Faraday effect, revealing that, as the perturbation grows, toroidal symmetry is broken and a helical equilibrium is established.

Saturated and aromatic mineral oil hydrocarbons from paperboard food packaging: estimation of long-term migration from contents in the paperboard and data on boxes from the market.

In the absence of a functional barrier, mineral oil hydrocarbons from printing inks and recycled fibres tend to migrate from paper-based food-packaging materials through the gas phase into dry food. Concentrations easily far exceed the limit derived from the acceptable daily intake (ADI) of the Joint FAO/WHO Expert Committee on Food Additives (JECFA). Since the estimation of long-term migration into the food by testing at 40°C for 10 days is difficult, it seems preferable (and easier) to use the mineral oil content in the paperboard. Evaporation experiments showed that hydrocarbons eluted up to about n-C24 are sufficiently volatile for relevant migration into dry food: in worst-case situations, about 80% migrate into the packed food. The extraction of the paperboard was optimised to give good recovery of the relevant hydrocarbons, but to discriminate against those of high molecular mass which tend to disturb gas chromatographic analysis in on-line coupled normal phase HPLC-GC-FID. Even though some of the relevant hydrocarbons had already evaporated, the average concentration of < C24 mineral oil saturated hydrocarbons (MOSH) in the paperboard boxes of 102 products from the Swiss and Italian market was 626 mg kg⁻¹. Nearly 15% of investigated boxes still contained more than 1000 mg kg⁻¹ < C24 MOSH up to over 3000 mg kg⁻¹ (maximum = 3500 mg kg⁻¹). This amount of MOSH in the board have the potential of contaminating the packed food at a level exceeding the limit, derived from the JECFA ADI, hundreds of times.

Single-helical-axis states in reversed-field-pinch plasmas.

The transition to a new magnetic topology, characterized by a quasi-single-helicity state with a single helical magnetic axis has been experimentally observed for the first time in a reversed-field-pinch plasma. The occurrence of the new state, which has been dubbed a single-helical-axis state, was found to provide magnetic chaos healing and enhanced thermal content of the plasma. The helical structure extends on both sides of the vessel geometric axis, and is related to exceeding a threshold in the ratio between the amplitude of the dominant MHD mode and the amplitude of the secondary ones.

[Specific immunotherapy and relocation in occupational allergic bakers]. / Recupero professionale di panificatori allergici mediante vaccino verso farina di frumento.

Occupational allergy to components of wheat flour is the main cause of rhinitis and asthma of workers in bakeries and similar activities. An immunological mechanism IgE-mediated is involved and the sensitising properties of some proteins of wheat where assessed. Nowadays it is possible to have an extract to be used for specific immunotherapy. The aim of this treatment should be a reduction of individual immunological reactivity and the possibility of going on the particular activity of allergic bakers, pastry makers or pizza makers. An observational crossectional retrospective study was performed on 41 sensitised workers that were diagnosed in the same occupational health unit. All underwent a subcutaneous specific immunotherapy (SCIT) with the same schedule and the same extract (Lofarma Allergeni, Milan) for 4 or more years, without avoiding their work activity. The outcome was investigated after five or ten years. Data were collected by a questionnaire. 34 subjects on 41 are still at work with an acceptable quality of life and a normal working efficiency, mainly in their small enterprises. In the "old" subgroup (19 cases), treated in the past, several bakers still at work stopped SCIT even from 4-10 years. In the "new" subgroup (15 cases), still in treatment, symptoms and drug use during the work activity resulted to be reduced or absent in the majority of cases. According to results of other immunotherapies by allergenic vaccines (pollens, mites) also for wheat flour occupational allergy a specific treatment seems to be possible and SCIT may be an useful tool to reduce and control the biological individual effects of allergy. By the occupational point of view wheat flour SCIT allows a relocation in many of cases and may be associated to other intervention of environmental prevention at workplaces, improving the relocation of occupational allergic subjects when requested.

Active-feedback control of the magnetic boundary for magnetohydrodynamic stabilization of a fusion plasma.

Stable operation with control on magnetohydrodynamic modes has been obtained in the modified reversed field experiment employing a set of 192 feedback controlled saddle coils. Improvements of plasma temperature, confinement (twofold), and pulse length (threefold) and, as a consequence of the magnetic fluctuation reduction, strong mitigation of plasma-wall interaction and mode locking are reported.

Species identification and confirmation of human and animal cell lines: a PCR-based method.

Misidentification and cross-contamination of cell lines are major problems of cell cultures that can make scientific results and their reproducibility unreliable. This paper describes a PCR-based method for easily identifying or confirming the species of origin of cell lines by using a panel of oligonucleotides specific for the nine animal species most common in cell culture laboratories. A panel of 35 human and animal cell lines, whose species of origin were previously confirmed by isoenzyme assay, was studied with nine species-specific primer pairs that specifically anneal to DNA sequences codifying for human, cat, dog, mouse, rat, horse, rabbit, African Green monkey cytochrome c oxidase subunit I (cox I), and one primer pair specific for the cytochrome b gene of Chinese hamster. The amplified fragments were analyzed by electrophoresis in ethidium bromide-stained 2% agarose gels. The method is simple, rapid, highly sensitive, and useful for routinely monitoring the species identity of cell cultures.

Specific localization of transcription factors in the chromatin of mouse mature spermatozoa.

We previously characterized a nuclease-hypersensitive fraction of mouse sperm chromatin, which is organized in a typical nucleosomal structure. A partial genomic library was constructed with the DNA from the nuclease-hypersensitive chromatin, which revealed a high content in retroposon/retroviral DNA sequences. Here we report that the cloned nuclease-hypersensitive DNA also contains clusters of potential sites for transcription factors: among those, binding sites for Oct-1, Oct-4, TBP, Ets-1, and C/EBP are most abundant. This observation prompted us to ask whether mature spermatozoa contain the corresponding protein factors. Indirect immunofluorescence experiments show that all analyzed factors are indeed present in the sperm heads. Moreover, transcription factors are associated with the nuclease-hypersensitive chromatin of spermatozoa, as endogenous nucleases that degrade the hypersensitive fraction also cause the concomitant release of transcription factors from sperm cells into the medium. Band-shift assays with proteins extracted from the supernatant, and immunofluorescence analysis of sperm pellets, indicate that transcription factors are largely recovered in the supernatant while being absent or poorly retained in spermatozoa. The possible involvement of these factors in early embryogenesis is discussed.

Accentuated response to phenylhydrazine and erythropoietin in mice genetically impaired for their GATA-1 expression (GATA-1(low) mice).

The response of mice genetically unable to up-regulate GATA-1 expression (GATA-1(low) mice) to acute (phenylhydrazine [PHZ]-induced anemia) and chronic (in vivo treatment for 5 days with 10 U erythropoietin [EPO] per mouse) erythroid stimuli was investigated. Adult GATA-1(low) mice are profoundly thrombocytopenic (platelet counts [x 10(9)/L] 82.0 +/- 28.0 vs 840 +/- 170.0 of their control littermates, P <.001) but have a normal hematocrit (Hct) (approximately.47 proportion of 1.0 [47%]). The spleens of these mutants are 2.5-fold larger than normal and contain 5-fold more megakaryocytic (4A5(+)), erythroid (TER-119(+)), and bipotent (erythroid/megakaryocytic, TER-119(+)/4A5(+)) precursor cells. Both the marrow and the spleen of these animals contain higher frequencies of burst-forming units-erythroid (BFU-E)- and colony-forming units-erythroid (CFU-E)-derived colonies (2-fold and 6-fold, respectively) than their normal littermates. The GATA-1(low) mice recover 2 days faster from the PHZ-induced anemia than their normal littermates (P <.01). In response to EPO, the Hct of the GATA-1(low) mice raised to.68 proportion of 1.0 (68%) vs the.55 proportion of 1.0 (55%) reached by the controls (P <.01). Both the GATA-1(low) and the normal mice respond to PHZ and EPO with similar (2- to 3-fold) increases in size and cellularity of the spleen (increases are limited mostly to cells, both progenitor and precursor, of the erythroid lineage). However, in spite of the similar relative cellular increases, the increases of all these cell populations are significantly higher, in absolute cell numbers, in the mutant than in the wild-type mice. In conclusion, the GATA-1(low) mutation increases the magnitude of the response to erythroid stimuli as a consequence of the expansion of the erythroid progenitor cells in their spleen.

Normal and cancer-prone human cells respond differently to extremely low frequency magnetic fields.

Human lymphoblastoid cells of normal origin and from genetic instability syndromes, i.e. Fanconi anemia (FA) group C and ataxia telangectasia, were continuously exposed to extremely low frequency magnetic field (ELF-MF). We report that ELF-MF, though not perturbing cell cycle progression, increases the rate of cell death in normal cell lines. In contrast, cell death is not affected in cells from genetic instability syndromes; this reflects a specific failure of the apoptotic response. Reintroduction of complementation group C in FA cells re-established the apoptotic response to ELF-MF. Thus, genes implicated in genetic instability syndromes are relevant in modulating the response of cells to ELF-MF.

Nucleosomal domains of mouse spermatozoa chromatin as potential sites for retroposition and foreign DNA integration.

Exogenous DNA molecules are spontaneously taken up by sperm cells, internalized in nuclei, and eventually integrated in the sperm genome. The actual occurrence of the integration suggests that the sperm chromosomal DNA is not uniformly and tightly packed with protamines, implying the existence of genomic sites where the chromosomal DNA is accessible to foreign molecules. We have characterized a hypersensitive, nucleosomal subfraction of mouse sperm chromatin that is highly enriched in unmethylated retroposon DNA from a variety of families. Here we propose that both the integration of exogenous DNA molecules, and the endogenous retroposition activity, occur in the same site(s) of sperm chromatin.

DNA dose and sequence dependence in sperm-mediated gene transfer.

We have tested three parameters in sperm-mediated gene transfer assays with mice and pigs: (i) the epididymal versus ejaculated origin of sperm cells, (ii) the primary structure, and (iii) the amount of the challenging foreign DNA. We have found that the pVLCNhGH construct, of retrotransposon origin, causes a massive embryo lethality and yet increases the yield of genetic transformation among born animals of both species compared to viral constructs. Arrest of embryonic development is a DNA dose-dependent effect, which is observed with high DNA doses, while lower doses are compatible with development. Finally, the overall efficiency of sperm-mediated gene transfer is higher when ejaculated, versus epididymal, spermatozoa are used. We suggest that this difference is related to the highly efficient apoptotic response in epididymal compared to ejaculated spermatozoa, triggered by the interaction of exogenous DNA molecules with the sperm membrane.

Reverse transcriptase activity in mature spermatozoa of mouse.

We show here that a reverse transcriptase (RT) activity is present in murine epididymal spermatozoa. Sperm cells incubated with human poliovirus RNA can take up exogenous RNA molecules and internalize them in nuclei. Direct PCR amplification of DNA extracted from RNA-incubated spermatozoa indicate that poliovirus RNA is reverse-transcribed in cDNA fragments. PCR analysis of two-cell embryos shows that poliovirus RNA-challenged spermatozoa transfer retrotranscribed cDNA molecules into eggs during in vitro fertilization. Finally, RT molecules can be visualized on sperm nuclear scaffolds by immunogold electron microscopy. These results, therefore, reveal a novel metabolic function in spermatozoa, which may play a role during early embryonic development.

Evaluation of pain and stress levels of animals used in experimental research.

This paper represents a contribution to researchers employing "in vivo" models in experimental surgery in order to obtain more reliable results in accordance with current legislation and Russel's "three R" statement: refinement, reduction, and replacement. After general consideration about the definition of pain and stress concerning laboratory animals, the authors suggest making an evaluation of the experimental protocol before approval by the local committee to allow assessment in terms of the costs/benefits of experimental research employing live animals.

A fraction of mouse sperm chromatin is organized in nucleosomal hypersensitive domains enriched in retroposon DNA.

We have characterized a nuclease hypersensitive chromatin fraction from murine spermatozoa. Endogenous nuclease activity can be induced in mouse epididymal spermatozoa by appropriate stimuli and cause the localized degradation of chromosomal DNA. Based on these observations, we have isolated nuclease hypersensitive chromatin regions released from spermatozoa in the supernatant of pelleted sperm cells, and have cloned and characterized the DNA. Gel electrophoresis of end-labelled released DNA fragments showed a typical nucleosomal distribution. Peripherally distributed nucleohistones were visualized by immunofluorescence in sperm nuclei, and histones were identified by western blot in sperm chromatin. Moreover, the released DNA is enriched in retroposon DNA from a variety of families. FISH and immunofluorescence analysis showed that retroposon DNA and nucleohistone chromatin co-localize and are both peripherically distributed in nuclei of spermatozoa. In contrast, a major satellite DNA probe, used for control, co-localizes with highly condensed chromatin in the central region of sperm nuclei. The nuclear Ran and RCC1 proteins were also visualized in the dorsal margin of sperm nuclei, and were abundantly released with the hypersensitive chromatin fraction. Together, these results indicate that nucleohistone chromatin fraction(s) with typical features of 'active' chromatin are present in murine spermatozoa, are hypersensitive to nuclease cleavage, enriched in retroposon DNA and organized in nucleosomal domains. These observations suggest that nucleohistone domains identify a fraction of the sperm genome which may be functional during early embryogenesis.

Matrix solid-phase dispersion microextraction and determination by high-performance liquid chromatography with UV detection of pesticide residues in citrus fruit.

A multiresidue method based on matrix solid-phase dispersion (MSPD) microextraction was studied to determine the carbamate, benfuracarb, and urea insecticides, diflubenzuron, flufenoxuron hexaflumuron and hexythiazox, used in control of citrus pests. Optimisation of different parameters, such as the type of solid support for matrix dispersion, elution solvents and the clean-up step were carried out. The method used 0.5 g of orange sample, C8 bonded silica as MSPD sorbent and dichloromethane as eluting solvent. Recoveries, at spiked concentrations below the maximum residue levels established by Spanish Government, were between 74 and 84% with relative standard deviations ranging from 2 to 4%. The limits of quantification were from 0.15 to 0.25 microgram/g using high-performance liquid chromatography with UV detection at 200 nm. The method may be useful as a screening protocol for the determination of these newly developed pesticides in citrus samples.

Increased production of mouse embryos in in vitro fertilization by preincubating sperm cells with the nuclease inhibitor aurintricarboxylic acid.

Exposure of spermatozoa to stress conditions causes a drastic reduction of their fertilizing ability. We report here that the decrease in fertilization can be effectively antagonized by preincubating sperm cells with the nuclease inhibitor drug aurintricarboxylic acid (ATA). Preincubation of mouse epididymal sperm cells with ATA increased the yield of 2-cell embryos produced by in vitro fertilization assays. The effect of ATA was selectively exerted via spermatozoa, since neither preincubation of eggs, nor the direct treatment of zygotes, modified the yield of 2-cell-stage embryos. Our results suggest that ATA does not directly improve the ability of sperm cells to penetrate the egg cytoplasm but instead acts by preserving sperm nuclei from induced or spontaneously occurring damage and/or favors events that trigger early embryogenesis.

Activation of endogenous nucleases in mature sperm cells upon interaction with exogenous DNA.

Mature sperm cells, either of epididymal origin or ejaculated and depleted of seminal fluid, are spontaneously able to bind exogenous DNA molecules which are subsequently internalized into sperm nuclei. Southern blot analysis showed that the internalized DNA was specifically cleaved by sperm endonucleases and showed typical fragmentation patterns of localized hypersensitivity. Nucleases were activated in response to the internalization of exogenous DNA by sperm cells and their activity increased with the DNA concentration. Nuclease activation was efficient in epididymal sperm cells, while being drastically reduced in ejaculated washed spermatozoa. Nucleases were Ca++ dependent, and were, respectively, inhibited and activated by preincubating sperm cells with Aurintricarboxylic Acid (ATA) and Ca++ Ionophore A23187, which are known to, respectively, inhibit and activate apoptosis in somatic cells. Moreover, nuclease activation also caused a partial degradation of the sperm endogenous chromosomal DNA; cleaved DNA fragments were released from the sperm cells to the medium. Taken together, these results suggest that a metabolically active process similar to apoptosis is triggered in the nuclei of mature sperm cells upon interaction with exogenous DNA.