Characterization of the bound residues of the fungicide cyprodinil formed in plant cell suspension cultures of wheat

The non-extractable residues of the fungicide cyprodinil formed in heterotrophic cell suspension cultures of wheat were studied by application of [2-pyrimidyl-14C] or [2-pyrimidyl-13C]cyprodinil. The main objective was to examine whether solid-state and liquid 13C NMR spectroscopy can be used to examine plant bound residues of pesticides. For 14C experiments, wheat suspensions grown on glucose as carbon source were treated with 10 mg litre(-1) of 14C-cyprodinil. After incubation for 12 days, 20% of applied 14C was detected as non-extractable residues. The cell debris were treated with 0.1 M HCl (reflux), 1.0 M HCl (reflux), buffer, or 2 M NaOH (50 degrees C); Bj?rkman lignin and acidolysis lignin fractions were also prepared from the debris. Radioactivity liberated and solubilized by these procedures was examined by thin-layer chromatography and high-performance liquid chromatography. The results showed that cyprodinil and primary metabolites contributed to the fungicide's bound residues. Most of the residues (12% of applied 14C) remained associated with polar or polymeric/oligomeric endogenous cell materials in a stable manner. For the study with 13C-cyprodinil, wheat suspensions were cultivated on 13C-depleted glucose for four growth cycles, resulting in maximum 13C depletion of the natural cell components to about 0.10%. During the fourth cycle, 13C-labelled cyprodinil was applied, and cells were incubated (12 days). Cell debris was prepared and examined by solid-state 13C NMR spectroscopy. Debris was then treated as described above in the 14C experiment. Solubilized fractions were analyzed by liquid 13C NMR spectroscopy. However, none of the 13C NMR spectra recorded gave utilizable or unambiguous results, and all exhibited large inconsistencies, especially concerning the data from the conventional 14C experiment.     

Light-dependent oxidative stress determines physiological leaf spot formation in barley

ABSTRACT We reported previously that physiological leaf spot (PLS) formation in winter and spring barley is dependent on genotype-related oxidative stress under field conditions. In the present study, we searched for factors inducing PLS symptoms in the greenhouse similar to those observed in the field and investigated its relationship to reactive oxygen species (ROS) metabolism. We found that in the greenhouse, oxidative stress induced spring barley cv. Extract, which is sensitive to PLS, to express symptoms similar to those observed in the field. Leaves severely affected by PLS showed significantly lower activities of key enzymes in the Halliwell-Asada cycle such as ascorbate peroxidase, glutathione reductase, dehy-droascorbate reductase, and monodehydroascorbate reductase. The sensitive cultivar also showed lower levels of total superoxide dismutase (SOD) and Cu/Zn-SOD activity but a higher level of chloroplast-specific Fe-SOD activity than that of the insensitive cultivar. Thus, an unbalanced ROS metabolism in chloroplasts may trigger PLS incidence in sensitive cultivars, which is in agreement with the fact that light is essential for the induction of PLS expression under both field and greenhouse conditions. Accordingly, under greenhouse conditions, continuous light stress (7 days), but not light shock treatments, induced PLS similar to that of field conditions in sensitive cv. Extract, but not in resistant cv. Scarlett. Light with a high proportion of energy in the blue wavelength spectrum (350 to 560 nm) was significantly more PLS inductive than light with a pronounced red (photosynthetically active radiation) spectrum (580 to 650 nm). Exposure to ozone did not produce PLS-like symptoms. Furthermore, similar to earlier observations in the field, PLS symptom expression was closely correlated with the accumulation of superoxide (O(2) (-)) detected by both biochemical and histochemical assays. Taken together, these data suggest that PLS in barley is genotype-dependent but its expression appears to be induced by certain environmental stress factors, among which photosyn-thetically active radiation plays a major role.     

Endoscopic-assisted lumbosacral foraminotomy in the dog

OBJECTIVE: To determine if endoscopic-assisted foraminotomy significantly increased the area of the L7-S1 intervertebral foramen and if, over 12 weeks, stenosis would occur. STUDY DESIGN: Prospective, experimental study. ANIMAL POPULATION: Six clinically normal, 22-29 kg, adult dogs. METHODS: Using endoscopic assistance, unilateral L7-S1 foraminotomy was performed. Computed tomography of L7-S1 was performed preoperatively, immediately postoperatively, and at 12 weeks. Parasagittal foramen area (PFA) measurements were obtained at the entry, middle, and exit zones of the treated and control foramen for each period. Objective and subjective data were compared among dogs by time period and treatment status. RESULTS: Endoscopic-assisted foraminotomy resulted in a significant increase in the mean PFA of the entry and middle zones immediately postoperatively. The exit zone was not significantly larger at any time. The foramen remained significantly larger at 12 weeks only in the middle zone; however, some decrease in the surgically created foramen enlargement occurred at all 3 levels. The procedure was well tolerated but dogs did have transient, mild delay of functional return postoperatively. CONCLUSIONS: Endoscopic-assisted foraminotomy in dogs can be performed for certain foraminal regions, allowing enhanced visibility of the spinal canal. The foramen can be surgically enlarged at the entry and middle zones using this technique; however, some reduction of the foraminal enlargement occurs by 12 weeks. The clinical implications of this reduction cannot be determined from this study. CLINICAL RELEVANCE: Endoscopic-assisted foraminotomy could be used to improve intraoperative visualization in dogs with foraminal stenosis as a component of degenerative lumbosacral stenosis.     

Hypothyroid goitre in a ram: chemical analysis gives indirect evidence for a structurally altered type of ovine thyroglobulin

The thyroglobulin of a ram of the East Friesian milk sheep breed suffering from goitre was investigated by physico- and immunochemical methods. The respective ram was the only animal amongst the other sheep of the flock, that exhibited severe goitre, additionally showing depressed behaviour. Results of the thyroid-stimulating hormone response test were indicative of hypothyroidism. The dysfunction of the thyroid gland could be treated by additional iodine supplementation quite successfully, although all sheep had been given iodinated cattle salt throughout the course of the history. Without reducing conditions sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) of isolated thyroglobulin molecules of the ram and control sheep did not reveal different band patterns, but under reducing conditions different band patterns were evident for the respective animals: the ram's thyroglobulin displayed two main bands, those of healthy reference sheep only one. Both bands reacted equally with anti-thyroglobulin antibodies, even with those produced by immunizing rabbits with single bands. The reduced single thyroglobulin band of healthy sheep corresponded to a truncated form of that molecule, whereas the additional main band of the ram was a more resistant, intact thyroglobulin subunit, as was shown by mass spectrometry. In conclusion, results of physico- and immunochemical investigations gave evidence of a modification of thyroglobulin with suspected different iodine binding properties in the ram. The latter finding may have clinical relevance in similar cases in other species, as it is an example of the impact that a minor change in a protein molecule may have on a complete metabolic pathway. Additionally, it could be shown, that in the ovine species the generally found single main band of thyroglobulin after reduction is a truncated form and not an intact subunit. This truncation seems to be induced in vitro by the reductive sample pretreatment prior to SDS-PAGE.     

Bruchpilot promotes active zone assembly, Ca2+ channel clustering, and vesicle release

The molecular organization of presynaptic active zones during calcium influx-triggered neurotransmitter release is the focus of intense investigation. The Drosophila coiled-coil domain protein Bruchpilot (BRP) was observed in donut-shaped structures centered at active zones of neuromuscular synapses by using subdiffraction resolution STED (stimulated emission depletion) fluorescence microscopy. At brp mutant active zones, electron-dense projections (T-bars) were entirely lost, Ca2+ channels were reduced in density, evoked vesicle release was depressed, and short-term plasticity was altered. BRP-like proteins seem to establish proximity between Ca2+ channels and vesicles to allow efficient transmitter release and patterned synaptic plasticity.     

Spin coupling in engineered atomic structures

We used a scanning tunneling microscope to probe the interactions between spins in individual atomic-scale magnetic structures. Linear chains of 1 to 10 manganese atoms were assembled one atom at a time on a thin insulating layer, and the spin excitation spectra of these structures were measured with inelastic electron tunneling spectroscopy. We observed excitations of the coupled atomic spins that can change both the total spin and its orientation. Comparison with a model spin-interaction Hamiltonian yielded the collective spin configuration and the strength of the coupling between the atomic spins.     

The preparation and structures of hydrogen ordered phases of ice

Two hydrogen ordered phases of ice were prepared by cooling the hydrogen disordered ices V and XII under pressure. Previous attempts to unlock the geometrical frustration in hydrogen-bonded structures have focused on doping with potassium hydroxide and have had success in partially increasing the hydrogen ordering in hexagonal ice I (ice Ih). By doping ices V and XII with hydrochloric acid, we have prepared ice XIII and ice XIV, and we analyzed their structures by powder neutron diffraction. The use of hydrogen chloride to release geometrical frustration opens up the possibility of completing the phase diagram of ice.     

Toll-like receptor triggering of a vitamin D-mediated human antimicrobial response

In innate immune responses, activation of Toll-like receptors (TLRs) triggers direct antimicrobial activity against intracellular bacteria, which in murine, but not human, monocytes and macrophages is mediated principally by nitric oxide. We report here that TLR activation of human macrophages up-regulated expression of the vitamin D receptor and the vitamin D-1-hydroxylase genes, leading to induction of the antimicrobial peptide cathelicidin and killing of intracellular Mycobacterium tuberculosis. We also observed that sera from African-American individuals, known to have increased susceptibility to tuberculosis, had low 25-hydroxyvitamin D and were inefficient in supporting cathelicidin messenger RNA induction. These data support a link between TLRs and vitamin D-mediated innate immunity and suggest that differences in ability of human populations to produce vitamin D may contribute to susceptibility to microbial infection.