An Ustilago maydis Mutant Partially Blocked in P45014DM Activity Is Hypersensitive to Azole Fungicides

An Ustilago maydis ergosterol biosynthesis mutant (A14) which is partially blocked in sterol 14alpha-demethylase (P45014DM) activity is described. This mutant accumulated the abnormal 14alpha-methyl sterols, eburicol, 14alpha-methylfecosterol, and obtusifoliol, along with significant amounts of ergosterol. Although the A14 mutant grew nearly as well as the wild type, it was impaired in cell extension growth, which indicated a dysfunction in apical cell wall synthesis. The mutant was also found to be hypersensitive to the azole fungicides penconazole and tebuconazole.     

Hexazonium pararosaniline as a fixative for animal tissues

Hexazonium pararosaniline is a valuable reagent that has been used in enzyme activity histochemistry for 50 years. It is an aqueous solution containing the tris-diazonium ion derived from pararosaniline, an aminotriarylmethane dye, and it contains an excess of nitrous acid that was not consumed in the diazotization reaction. Other investigators have found that immersion for 2 min in an acidic (pH 3.5) 0.0015 M hexazonium pararosaniline solution can protect cryostat sections of unfixed animal tissues from the deleterious effects of aqueous reagents such as buffered solutions used in immunohistochemistry, while preserving specific affinities for antibodies. In the present investigation hexazonium pararosaniline protected lymphoid tissue and striated muscle against the damaging effects of water or saline. The same protection was conferred on unfixed sections treated with dilute nitrous or hydrochloric acid in concentrations similar to those in hexazonium pararosaniline solutions. Model tissues (solutions, gels or films containing gelatin and/or bovine albumin) responded predictably to well known cross-linking (formaldehyde) or coagulant (mercuric chloride) fixatives. Hexazonium pararosaniline solutions prevented the dissolution of protein gels in water only after 9 or more days of contact, during which time considerable swelling occurred. It is concluded that there is no evidence for a “fixative” action of hexazonium pararosaniline. The protective effect on frozen sections of unfixed tissue is attributable probably to the low pH of the solution.     

Impaired Excitatory Drive to Spinal Gabaergic Neurons of Neuropathic Mice

Adequate pain sensitivity requires a delicate balance between excitation and inhibition in the dorsal horn of the spinal cord. This balance is severely impaired in neuropathy leading to enhanced pain sensations (hyperalgesia). The underlying mechanisms remain elusive. Here we explored the hypothesis that the excitatory drive to spinal GABAergic neurons might be impaired in neuropathic animals. Transgenic adult mice expressing EGFP under the promoter for GAD67 underwent either chronic constriction injury of the sciatic nerve or sham surgery. In transverse slices from lumbar spinal cord we performed whole-cell patch-clamp recordings from identified GABAergic neurons in lamina II. In neuropathic animals rates of mEPSC were reduced indicating diminished global excitatory input. This downregulation of excitatory drive required a rise in postsynaptic Ca²⁺. Neither the density and morphology of dendritic spines on GABAergic neurons nor the number of excitatory synapses contacting GABAergic neurons were affected by neuropathy. In contrast, paired-pulse ratio of Aδ- or C-fiber-evoked monosynaptic EPSCs following dorsal root stimulation was increased in neuropathic animals suggesting reduced neurotransmitter release from primary afferents. Our data indicate that peripheral neuropathy triggers Ca²⁺-dependent signaling pathways in spinal GABAergic neurons. This leads to a global downregulation of the excitatory drive to GABAergic neurons. The downregulation involves a presynaptic mechanism and also applies to the excitation of GABAergic neurons by presumably nociceptive Aδ- and C-fibers. This then leads to an inadequately low recruitment of inhibitory interneurons during nociception. We suggest that this previously unrecognized mechanism of impaired spinal inhibition contributes to hyperalgesia in neuropathy.     

Seminal roots of wild and cultivated barley differentially respond to osmotic stress in gene expression,suberization, and hydraulic conductivity

Wild barley, Hordeum vulgare spp. spontaneum, has a wider genetic diversity than its cultivated progeny, Hordeum vulgare spp. vulgare. Osmotic stress leads to a series of different responses in wild barley seminal roots, ranging from no changes in suberization to enhanced endodermal suberization of certain zones and the formation of a suberized exodermis, which was not observed in the modern cultivars studied so far. Further, as a response to osmotic stress, the hydraulic conductivity of roots was not affected in wild barley, but it was 2.5-fold reduced in cultivated barley. In both subspecies, osmotic adjustment by increasing proline concentration and decreasing osmotic potential in roots was observed. RNA-sequencing indicated that the regulation of suberin biosynthesis and water transport via aquaporins were different between wild and cultivated barley. These results indicate that wild barley uses different strategies to cope with osmotic stress compared with cultivated barley. Thus, it seems that wild barley is better adapted to cope with osmotic stress by maintaining a significantly higher hydraulic conductivity of roots during water deficit.     

Characterization of novel mutants with an altered gibberellin spectrum in comparison to different wild-type strains of Fusarium fujikuroi

The rice pathogen Fusarium fujikuroi is known for producing a wide range of secondary metabolites such as pigments, mycotoxins, and a group of phytohormones, the gibberellic acids (GAs). Bioactive forms of these diterpenes are responsible for hyperelongation of rice stems, yellowish chlorotic leaves, and reduced grain formation during the bakanae disease leading to severely decreased crop yields. GAs are also successfully applied in agriculture and horticulture as plant growth regulators to enhance crop yields, fruit size, and to induce earlier flowering. In this study, six F. fujikuroi wild-type and mutant strains differing in GA yields and the spectrum of produced GAs were cultivated in high-quality lab fermenters for optimal temperature and pH control and compared regarding their growth, GA production, and GA gene expression levels. Comparative analysis of the six strains revealed that strain 6314/ΔDES/ΔPPT1, holding mutations in two GA biosynthetic genes and an additional deletion of the 4'-phosphopantetheinyl transferase gene PPT1, exhibits the highest total GA amount. Expression studies of two GA biosynthesis genes, CPS/KS and DES, showed a constantly high expression level for both genes under production conditions (nitrogen limitation) in all strains. By cultivating these genetically engineered mutant strains, we were able to produce not only mixtures of different bioactive GAs (GA₃, GA_4, and GA₇) but also pure GA₄ or GA₇. In addition, we show that the GA yields are not only determined by different production rates, but also by different decomposition rates of the end products GA₃, GA₄, and GA₇ explaining the varying GA levels of genetically almost identical mutant strains.     

Synthesis,docking, and biological studies of phenanthrene β-diketo acids as novel HIV-1 integrase inhibitors

In the present study we report the synthesis of halogen-substituted phenanthrene β-diketo acids as new HIV-1 integrase inhibitors. The target phenanthrenes were obtained using both standard thermal- and microwave-assisted synthesis. 4-(6-Chlorophenanthren-2-yl)-2,4-dioxobutanoic acid (18) was the most active compound of the series, inhibiting both 3′-end processing (3′-P) and strand transfer (ST) with IC₅₀ values of 5 and 1.3 μM, respectively. Docking studies revealed two predominant binding modes that were distinct from the binding modes of raltegravir and elvitegravir, and suggest a novel binding region in the IN active site. Moreover, these compounds are predicted not to interact significantly with some of the key amino acids (Q148 and N155) implicated in viral resistance. Therefore, this series of compounds can further be investigated for a possible chemotype to circumvent resistance to clinical HIV-1 IN inhibitors.     

Economic evaluation of water loss saving due to the biological control of water hyacinth at New Year’s Dam,Eastern Cape province,South Africa

Water hyacinth Eichhornia crassipes is considered the most damaging aquatic weed in the world. However, few studies have quantified the impact of this weed economically and ecologically, and even fewer studies have quantified the benefits of its control. This paper focuses on water loss saving as the benefit derived from biological control of this plant between 1990 and 2013 at New Year’s Dam, Alicedale, Eastern Cape, South Africa. Estimates of water loss due to evapotranspiration from water hyacinth vary significantly; therefore, the study used three different rates, high, medium and low. A conservative raw agriculture value of R 0.26 per m³ was used to calculate the benefits derived by the water saved. The present benefit and cost values were determined using 10% and 5% discount rates. The benefit/cost ratio at the low evapotranspiration rate was less than one, implying that biological control was not economically viable but, at the higher evapotranspiration rates, the return justified the costs of biological control. However, at the marginal value product of water, the inclusion of the costs of damage to infrastructure, or the adverse effects of water hyacinth on biodiversity, would justify the use of biological control, even at the low transpiration rate.     

Positioning Accuracy in Otosurgery Measured with Optical Tracking

Objectives

To assess positioning accuracy in otosurgery and to test the impact of the two-handed instrument holding technique and the instrument support technique on surgical precision. To test an otologic training model with optical tracking.

Study Design

In total, 14 ENT surgeons in the same department with different levels of surgical experience performed static and dynamic tasks with otologic microinstruments under simulated otosurgical conditions.

Methods

Tip motion of the microinstrument was registered in three dimensions by optical tracking during 10 different tasks simulating surgical steps such as prosthesis crimping and dissection of the middle ear using formalin-fixed temporal bone. Instrument marker trajectories were compared within groups of experienced and less experienced surgeons performing uncompensated or compensated exercises.

Results

Experienced surgeons have significantly better positioning accuracy than novice ear surgeons in terms of mean displacement values of marker trajectories. The instrument support and the two-handed instrument holding techniques significantly reduce surgeons’ tremor. The laboratory set-up presented in this study provides precise feedback for otosurgeons about their surgical skills and proved to be a useful device for otosurgical training.

Conclusions

Simple tremor compensation techniques may offer trainees the potential to improve their positioning accuracy to the level of more experienced surgeons. Training in an experimental otologic environment with optical tracking may aid acquisition of technical skills in middle ear surgery and potentially shorten the learning curve. Thus, simulated exercises of surgical steps should be integrated into the training of otosurgeons.     

Genes,symptoms, and the “asymptomatic ill”: toward a broader understanding of genetic discrimination

Since the early 1990s, the term “genetic discrimination” has been used to designate adverse treatment on the grounds of genetic makeup. However, the full spectrum of possible disadvantage associated with genetic information has not been addressed by either the international scientific debate or statutory arrangements on genetic discrimination. Informed by legal contexts, they almost all focus on one specific group: the “asymptomatic ill.” On the basis of the sociological study, “Genetic Discrimination in Germany,” this article proposes to revise the terms of the debate and discusses some limitations of the concept. Drawing on the experiences reported by affected individuals, it advocates a more expansive social understanding which does not require that a person has to be healthy to be at risk of genetic discrimination.