土壤中棉花黄萎病菌SYBR Green Ⅰ荧光RT-PCR定量检测技术研究

为实现田间土壤棉花黄萎病菌的早期检测,建立了土壤中棉花黄萎病菌的SYBR Green I荧光定量PCR检测方法。以含342bp PCR扩增产物的阳性质粒为参考,构建了标准曲线,并对该曲线的特异性、敏感性、可重复性进行了评价。结果表明,该方法具有快速、特异性强、敏感度高等特点。检测范围在3.8×103-3.8×108copies/μL之间有良好的线性关系,相关系数R2为0.996,扩增效率为101.5%,灵敏度比常规PCR方法高102倍。     

A New Basidiomycetous Yeast Species, Cryptococcus mycelialis, Related to Holtermannia Saccardo et Traverso

A new species of the genus Cryptococcus, Cr. mycelialis (the type strain VKM Y-2863), is described based on the taxonomic study of four strains isolated from soil and plant samples collected on the South Georgia and East Falkland islands. This species differs from the known Cryptococcus species in its ability to form a true monokaryotic mycelium with pseudoclamp connections and haustoria. The species can be distinguished from the phylogenetically related and phenotypically similar species Holtermannia corniformis and Cr. nyarrowii by some assimilatory reactions, maximum growth temperature, and sensitivity to mycocins.     

Purification, and biochemical and biophysical characterization of cellobiohydrolase I from Trichoderma harzianum IOC 3844

Because of its elevated cellulolytic activity, the filamentous fungus Trichoderma harzianum has a considerable potential in biomass hydrolysis applications. Trichoderma harzianum cellobiohydrolase I (ThCBHI), an exoglucanase, is an important enzyme in the process of cellulose degradation. Here, we report an easy single-step ion-exchange chromatographic method for purification of ThCBHI and its initial biophysical and biochemical characterization. The ThCBHI produced by induction with microcrystalline cellulose under submerged fermentation was purified on DEAE-Sephadex A-50 media and its identity was confirmed by mass spectrometry. The ThCBHI biochemical characterization showed that the protein has a molecular mass of 66 kDa and pI of 5.23. As confirmed by smallangle X-ray scattering (SAXS), both full-length ThCBHI and its catalytic core domain (CCD) obtained by digestion with papain are monomeric in solution. Secondary structure analysis of ThCBHI by circular dichroism revealed alpha- helices and beta-strands contents in the 28% and 38% range, respectively. The intrinsic fluorescence emission maximum of 337 nm was accounted for as different degrees of exposure of ThCBHI tryptophan residues to water. Moreover, ThCBHI displayed maximum activity at pH 5.0 and temperature of 50 degrees C with specific activities against Avicel and p-nitrophenyl-β-D-cellobioside of 1.25 U/mg and 1.53 U/mg, respectively.     

Changes in parameters of humoral immunity in patients with acute lung abscesses in the course of ultraviolet phototherapy

Ultraviolet fiber optic phototherapy of the blood and purulent cavities in patients with acute purulent lung abscesses results in a pronounced improvement of humoral immunity expressed in a tendency of all classes of immunoglobulins toward normalization. The bacterial flora of the purulent cavities and bronchial contents is markedly eradicated.     

Differences in the corticofugal facilitating effect on evoked potentials in the neostriatum of rats and monkeys

Identical facilitation of the primary response to peripheral stimuli in rats and monkeys has been induced by local superficial cooling of the cortex in the somatosensory area S1. Higher facilitation of the evoked potentials was observed in the neostriatum of rats. Correlation of the evoked potentials with primary responses was more significant with respect to temporal than amplitude parameters. In monkeys, "cool" facilitation of the cortical primary response did not result in facilitation of the evoked potential in the neostriatum. Temporal correlation between the primary response and the evoked potential was less significant, whereas correlation in the amplitude was absent. The data obtained indicate the existence of differences in functional organization of corticofugal influences upon the neostriatum in rodents and primates.     

Dominant form of cationic peroxidase from sorghum roots

A dominant form of cationic peroxidase (PO-2) was isolated from sorghum (Sorghum bicolor L. Moench) roots and purified to electrophoretically homogeneous state. The enzyme is a monomer with mol wt of 49.7 kD. The optimum pH and the main catalytic constants (K_M, V_max, k_cat) were determined for oxidation of the main substrates including Н₂О₂, 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulfonate) (ABTS), 2,7-diaminofluorene, syringaldazine, 2,6-dimethoxyphenol, and o-dianisidine. The K_M values increased in the sequence: H₂O₂ < 2,7-diaminofluorene < ABTS < o-dianisidine, whereas the maximum turnover number (93.9 s^–1) was found for 2,7-diaminofluorene. Based on the analysis of molecular and catalytic properties of the enzyme, it was proven that PO-2 is a typical cationic plant peroxidase. Polycyclic aromatic hydrocarbons (phenanthrene, anthracene, fluorene), 2,2'-diphenic acid, and Ni ions had no significant influence on the activity of PO-2. The enzyme was inhibited by p-aminobenzoic acid, NaN₃, 1-naphthol, 9,10-anthraquinone, and 9,10-phenanthrenequinone. In the presence of NaN₃, 1-naphthol, and 9,10-phenanthrenequinone, a mixed competitive/noncompetitive type of inhibition was noted. The peroxidase PO-2 was found to oxidize synthetic anthraquinone dyes, phenanthrene, and some oxygenated derivatives of polycyclic aromatic hydrocarbons (9-phenanthrol; 1-naphthol; and 1-hydroxy-2-naphthoic, salicylic, and 2,2'-diphenic acids), which indirectly confirms the coupled plant–microbial metabolism of these compounds in the root zone of sorghum. The results indicate that 9,10-phenanthrenequinone and 2,2'-diphenic acid are the products of peroxidase-catalyzed oxidation of 9-phenanthrol.     

Histochemical changes in neonatal liver caused by vaginal instillation of magnetic nanoparticles in pregnant mice

Drug delivery through the vagina is a novel and effective approach for treating embryonic diseases. Magnetic nanoparticles (MNPs) currently are used as drug delivery systems. The safety of MNPs for use with embryonic tissues remains unclear. We used pregnant mice to investigate the possible toxicity of MNPs toward neonatal liver at three embryonic ages using histochemical and immunohistochemical techniques. MNPs were instilled through the vaginas of pregnant mice at days 12 (E12), 15 (E15) and 17 (E17) after fertilization. We found MNPs in the neonatal liver parenchyma after delivery of the pups on day 20. We observed that MNPs caused mild apoptosis of hepatocytes, cytoplasmic vacuolation and lymphocytic infiltration in the neonatal liver after treatment at E15 compared to instillation at E12 and E17. We observed also that MNPs increased the production of caspase proteins and tumor necrosis factor receptor 2 proteins, which are indicators of apoptosis, in the neonatal liver after instillation of MNPs at E15 compared to instillation at E12 and E17. MNPs also increased the number of collagen fibers and the amounts of connective tissue growth factors in the neonatal liver parenchyma after instillation at E15 compared to instillation at E12 and E17. The general carbohydrates in the neonatal liver were decreased in a time-dependent manner after instillation at E17, E15 and E12 owing to the presence of MNPs in the parenchyma. Overall, we determined that MNPs were mildly toxic to neonatal liver.