Establishment of <i>Rhodiola quadrifida</i> Hairy Roots and Callus Culture to Produce Bioactive Compounds

Rhodiola quadrifida is a rare mountain medicinal plant whose root extracts are used in traditional Chinese medicine as a hemostatic, antitussive, and tonic in the treatment of gynecological diseases. The aim of the study was to obtain R. quadrifida cultures at different degrees of differentiation in vitro and compare their growth characteristics and the content of salidroside and rosavin. Hairy roots were obtained by incubating cotyledons and hypocotyls in a suspension of Agrobacterium rhizogenes strain A4. The presence of the rolB and rolC genes was proven by polymerase chain reaction. The obtained roots were cultivated in Murashige-Skoog medium (MS). Calluses were obtained from the hairy roots in MS medium with the addition of hormones: 3 mg/L 2,4 D and 0.5 mg/L BAP. The presence of the main secondary metabolites of R. quadrifida, salidroside and rosavin, in calluses and salidroside in hairy roots by HPLC/MS was confirmed. The content of salidroside in callus culture was significantly higher than in hairy roots, 0.158 and 0.047%, respectively. The content of rosavin in callus culture was 0.07%. The content of rosavin and salidroside in callus culture was close to the level of these substances in the rhizomes of R. quadrifida plants growing in vivo, making this culture promising for its possible biotechnological use.     

Effect of bacterial satellites on Chlamydomonas reinhardtii growth in an algo-bacterial community

The growth characteristics of an algo-bacterial community (Chlamydomonas reinhardtii and bacterial satellites) were studied, as well as the mechanism and patterns of bacterial effect on algae. Four strains of predominant bacteria were isolated and partially characterized. They were assigned to the following taxa: Rhodococcus terrea, Micrococcus roseus, and Bacillus spp. A pure culture of the alga under study was obtained by plating serial dilutions on agarized media. Within the algo-bacterial association, the alga had a higher growth rate (0.76 day^?1) and yield (60 μg chlorophyll/ml culture) than in pure cultures (0.4 day^?1 and 10 μg chlorophyll/ml culture, respectively). The viability of the algal cells within the association was retained longer than in pure culture. Among the isolated bacterial satellites, strains B1 and Y1, assigned to the species Rhodococcus terrae, had the highest stimulatory effect on algal growth. The culture liquid of bacteria incubated under the conditions not permitting growth stimulated algal growth; the culture liquid of actively growing bacteria had an opposite effect.     

A huge,undescribed soil ciliate (Protozoa: Ciliophora) diversity in natural forest stands of Central Europe

We investigated 12 natural forest stands in eastern Austria for soil ciliate diversity, viz., eight beech forests and two lowland and Pinus nigra forests each. The stands span a wide range of climatic (e.g., 543–1759 mm precipitation, 160–1035 m above sea-level) and abiotic (e.g., pH 4–7.4) factors. Samples were taken twice in autumn and late spring and analysed with the non-flooded Petri dish method. Species were identified in vivo, in silver preparations, and in the scanning electron microscope. A total of 233 species were found, of which 30 were undescribed, a surprising number showing our ignorance of soil ciliate diversity, even in Central Europe. Species number varied highly from 45 (acidic beech on silicate) to 120 (floodplain forest) and was strongly correlated with pH and overall habitat quality, as measured by climate, the C/P quotient (ratio of r-selected colpodean and k-selected polyhymenophorean ciliates), and the proportion of mycophagous ciliate species; multivariate analysis showed further important variables, viz., the general nutrient status (glucose, nitrogen, C/ N ratio) and microbial (urease) activity. The highest species number occurred in one of the two floodplain soils, supporting the intermediate disturbance hypothesis. The three main forest types could be clearly distinguished by their ciliate communities, using similarity indices and multidimensional scaling. Individual numbers varied highly from 135^–1 (lowland forest) to 10,925 ml^–1 (beech on silicate) soil percolate and showed, interestingly, a weak correlation with soil protozoan phospholipid fatty acids. Eight of the 30 new species found and a forgotten species, Arcuospathidium coemeterii (Kahl 1943) nov. comb., are described in detail, as examples of how species were recognized and soil protozoan diversity should be analyzed: Latispathidium truncatum bimicronucleatum, Protospathidium fusioplites, Erimophrya sylvatica, E. quadrinucleata, Paragonostomum simplex, Periholosticha paucicirrata, P. sylvatica, and Australocirrus zechmeisterae.     

Morphological characteristics of natural strains of certain species of basidiomycetes and biological analysis of antimicrobial activity under submerged cultural conditions

Twenty-one strains belonging to 18 species of basidiomycetes from different ecological groups of fungi were isolated from natural sources. Light and electron microscopy was used to determine the morphological properties of the cultures, which confirmed their classification as basidiomycetes and facilitated their identification in monocultures. The capacity of the fungal strains for biosynthesis of antibiotics was determined by one- or two-stage cultivation on seven nutrient media. It was established that, under submerged cultivation, antimicrobial substances were formed by 13 strains (81.25%) of 12 fungal species (Armillaria sp., Coprinus comatus, Flammulina velutipes, Hypsizygus ulmarius, Lentinus tigrinus, Lycoperdon pyriforme, Macrolepiota procera, Panellus serotinus, Pholiota aurivella, Pholiota lenta, Rhodocollybia maculate, and Sparassis crispa). The antibiotics formed were efficacious against bacterial test strains, including the methicillin-resistant strain Staphylococcus aureus (MRSA) and the strain Leuconostoc mesenteroides VKPM B-4177 that is resistant to the glycopeptide antibiotics. No antibiotic activity was revealed against fungal test cultures (Aspergillus niger INA 00760 and Saccharomyces cerevisiae RIA 259).     

Production of singlet oxygen in a non-self-sustained discharge

The production of O₂(a¹Δ_g) singlet oxygen in non-self-sustained discharges in pure oxygen and mixtures of oxygen with noble gases (Ar or He) was studied experimentally. It is shown that the energy efficiency of O₂(a¹Δ_g production can be optimized with respect to the reduced electric field E/N. It is shown that the optimal E/N values correspond to electron temperatures of 1.2–1.4 eV. At these E/N values, a decrease in the oxygen percentage in the mixture leads to an increase in the excitation rate of singlet oxygen because of the increase in the specific energy deposition per O₂ molecule. The onset of discharge instabilities not only greatly reduces the energy efficiency of singlet oxygen production but also makes it impossible to achieve high energy deposition in a non-self-sustained discharge. A model of a non-self-sustained discharge in pure oxygen is developed. It is shown that good agreement between the experimental and computed results for a discharge in oxygen over a wide range of reduced electric fields can be achieved only by taking into account the ion component of the discharge current. The cross section for the electron-impact excitation of O₂(a¹Δ_g and the kinetic scheme of the discharge processes with the participation of singlet oxygen are verified by comparing the experimental and computed data on the energy efficiency of the production of O₂(a¹Δ_g and the dynamics of its concentration. It is shown that, in the dynamics of O₂(a¹Δ_g molecules in the discharge afterglow, an important role is played by their deexcitation in a three-body reaction with the participation of O(³P) atoms. At high energy depositions in a non-self-sustained discharge, this reaction can reduce the maximal attainable concentration of singlet oxygen. The effect of a hydrogen additive to an Ar: O₂ mixture is analyzed based on the results obtained using the model developed. It is shown that, for actual electron beam current densities, a significant energy deposition in a non-self-sustained discharge in the mixtures under study can be achieved due to the high rate of electron detachment from negative ions. In this case, however, significant heating of the mixture can lead to a rapid quenching of O₂(a¹Δ_g molecules by atomic hydrogen.     

Optimized Fractioning and Structure Analysis of the Reactivating Factor from Luteococcus japonicus subsp. casei

Microbiology - The chemical structure of the extracellular reactivating factor (RF) from Luteococcus japonicus subsp. casei was determined; this factor promotes survival of a small subpopulation of...     

Changes in the carbohydrate and hormonal status in <Emphasis Type="Italic">Tulipa bifloriformis</Emphasis> bulbs forced into bloom in a greenhouse and in the open ground

Time-course of ABA, cytokinins, monosaccharides (MS), and water-soluble polysaccharides (WSP) contents were followed during cold dormancy period in the bulbs of tulip (Tulipa bifloriformis Vved.) that stayed over winter in the open ground or were cold-forced into bloom in a greenhouse. In both cases, the level of monosaccharides and water-soluble polysaccharides in the storage scale tissues increased, whereas the MS/WSP ratio in the bulbs planted in the open remained essentially the same and in case of forcing treatment decreased almost fivefold. In the apical bud tissues, the level of monosaccharides also rose in both cases, but the MS/WSP ratio in the open was greater throughout the whole experiment. The level of cytokinins in the apical bud tissues in the open was higher than in the forcing case, although the changes in their total content were identical following both treatments. Following the forcing treatment, the contents of free and bound ABA in the apical bud tissues increased reaching their peaks by the end of cold period. In the open, there were two peaks of free ABA: in October (when early frosts occurred) and in March (at the end of the wintering period). The winter forcing treatment resulted in rapid depletion of energy and plastic resources in T. bifloriformis and the death of 20% of embryonic flower buds (in the open, all flower buds survived). Nevertheless, plant adaptation potential ensured the development of generative shoots with 4–5 normal flowers, which makes it possible to use this species of multiflorous tulip for winter forcing in a greenhouse.     

Patterns of pyrite oxidation by different microorganisms

The bacterial-chemical oxidation of natural pyrites with different physical, chemical, and electrophysical characteristics by bacteria Acidithiobacillus ferrooxidans, Sulfobacillus thermotolerans, and the archaeon Ferroplasma acidiphilum were studied. The electrophysical characteristics of three natural pyrites differed in the K _thermoEMF value (pyrites 3, 4, hole conduction (p-type conductivity); pyrite 5, mixed type conductivity (n-p)) and in the logarithm of electric resistance. Chemical oxidation of pyrites 3 and 5 resulted in no changes of K _thermoEMF. When pyrite 4 was oxidized chemically, the K _thermoEMF values remained in the same range as in the initial sample, but the ratio of grains with different K _thermoEMF values in the sample was changed: the number of grains with a higher K _thermoEMF value increased. The same changes were also observed in the course of bacterio-chemical oxidation of pyrite 4. Of the three pyrites studied, an increase in the logarithm of resistance was observed only for chemical oxidation of pyrite 4 at 28°C. At higher experimental temperatures, the logarithm of resistance increased accordingly; more active bacterial-chemical oxidation resulted in a more pronounced increase in the logarithm of resistance than chemical oxidation. On bacterial-chemical oxidation of pyrites 3 and 5 by A. ferrooxidans and S. thermotolerans strains, iron was leached more actively than sulfur. Preferred bacterial-chemical oxidation of certain fractions from the pyrite samples was shown, namely of the pyrite 3 fraction with higher K _thermoEMF values by the F. acidiphilum strain and of a fraction from the pyrite 5 sample with medium K _thermoEMF values by the A. ferrooxidans and S. thermotolerans strains. The comparative assessment of bacterial-chemical pyrite oxidation by three types of microorganisms showed the direction of changes in the K _thermoEMF values to be the same in the case of bacteria Acidithiobacillus ferrooxidans and Sulfobacillus thermotolerans and different in the case of the archaeon Ferroplasma acidiphilum.