Detection of Hispidin by a Luminescent System from Basidiomycete Armillaria borealis.

In in vitro experiments, the possibility of using a luminescent system extracted from the luminous fungus Armillaria borealis has been shown to detect and determine the concentration of hispidin. A linear dependence of the luminescent response on the content of hispidin in solutions in the concentration range of 5.4 × 10-5-1.4 × 10-2 µM was detected. The stability of the enzyme system and the high sensitivity of the bioluminescent reaction allows carrying out multiple measurements with the analyte detection limit of 1.3 × 10-11 g. The obtained results show the prospects of creating a rapid bioluminescent method for the analysis of medical substances or extracts from various biological objects for the presence of hispidin.

[Total Peroxidase and Catalase Activity of Luminous Basidiomycetes Armillaria borealis and Neonothopanus nambi in Comparison with the Level of Light Emission].

The peroxidase and catalase activities in the mycelium of luminous basidiomycetes Armillaria borealis and Neonothopanus nambi in normal conditions and under stress were compared. An increase in the luminescence level was observed under stress, as well as an increase in peroxidase and catalase activities. Moreover, the peroxidase activity in extracts of A. borealis mycelium was found to be almost one and a half orders of magnitude higher, and the catalase activity more than two orders of magnitude higher in comparison with the N. nambi mycelium. It can be suggested that the difference between the brightly luminescent and dimly luminescent mycelium of N. nambi is due to the content of H2O2 or other peroxide compounds.

[Effect of organic and inorganic toxic compounds on luminescence of luminous fungi].

The possibility of the development of the solid phase bioluminescent biotest using aerial mycelium of the luminous fungi was investigated. Effect of organic and inorganic toxic compounds (TC) at concentrations from 10(-6) to 1 mg/ml on luminescence of aerial mycelia of four species of luminous fungi-Armillaria borealis (Culture Collection of the Institute of Forest, Siberian Branch, Russian Academy of Sciences), A. mellea, A. gallica, and Lampteromyces japonicus (Fungi Collection of the Botanical Institute, Russian Academy of Sciences)--has been studied. Culture of A. mellea was shown to be most sensitive to solutions of the model TC. It was demonstrated that the sensitivity of the luminous fungi is comparable with the sensitivity of the bacteria that are used for environmental monitoring. Use of the aerial mycelium of the luminous fungi on the solid support as a test object is a promising approach in biotesting for the development of continuous biosensors for air monitoring.

[Quantitative criteria for the estimation of the effectiveness of bioluminescence expression in natural and transgenic luminescent bacteria].

Computation coefficients for estimating the effectiveness of bioluminescence expression in natural luminescent bacteria P. leiognathi 54 and transgenic strain E. coli Z905/pPHL7 bearing lux-operon in multicopy plasmid are suggested, and their use on the molecular, cell, and population levels was considered. It was shown that, on the population level, all transgenic variants got the better of natural variants of P. leiognathi 54 irrespective of the type of lux-operon regulation. On the cell level, in the bright and dim variants of the transgenic strain, the effectiveness of bioluminescence expression increases by several orders. On the level of one lux-operon, the effectiveness of expression of the bright variant of transgenic strain is substantially higher than in the natural bright variant; in dim variants, the efficiency values are similar, and the effectiveness of bioluminescence expression in the dark variant of E. coli Z905-2 /pPHL7 is by two orders lower than that in the dark variant of P. leiognathi 54.

[Heterogeneity of the populations of marine luminescent bacteria Photobacterium leiognathi under different conditions of cultivation].

Manifestation of pleiotropic effects in the isogenic variants of luminescent bacteria Photobacterium leiognathi 54 was investigated. The decrease or increase of the expression level of bioluminescence was caused by changes in lux operon regulation. The dynamics of the bioluminescence of dark and dim variants did not differ from the dynamics of the initial luminescent variant, but dependence of the level of luminescence intensity on the exogenous autoinductor of the lux operon was revealed. The investigated variants of P. leiognathi 54 inherited fairly stable morphological characteristics, colony architectonics, level of luminescence, and activity of some enzymes; variants with reduced bioluminescence formed colonies of the S type. Stable bright variants with S- and R-type colonies appeared both in the initial strain population and in the dark variant population, but with smaller frequency. Populations of the bright variant with R-type colonies were most heterogeneous; this can be determined by the lack of glucose repression of the bioluminescence in contrast to other investigated variants of P. leiognathi.

[Database on natural and transgenic luminous microorganisms: "BiolumBase"].

The database "BiolumBase" is designed for the selection and systematization of available world information on microorganisms containing bioluminescent systems; it includes two sections: "natural" and "transgenic" luminous microorganisms. At present, logic schemes of divisions, classification of the objects, presentation of characteristics, and the inputs of relative information, as well as the necessary program modules including links to the database, are developed. The database is constructed on the basis of published data and our own experimental results; the subsequent linkage of the database to the Internet is envisaged. Users will be able to obtain not only the catalogues of strains but also information concerning the properties and functions of the known species of luminous bacteria, the structure, regulatory mechanisms, and application of bioluminescent systems and genetically engineered constructions with lux genes, as well as to find references and to search strains by using any set of attributes. The database will provide information that is of interest for the development of microbial ecology and biotechnology, in particular, for the prediction of biological hazard from the application of transgenic strains.

Transfer of xenobiotics through cell membranes of luminous bacteria.

The influence of some chemical substances on luminous bacteria was studied to elucidate the interrelation between the xenobiotics action on bacterial luminescence and cell ultrastructure. Such substances as quinones, phenols, chlorides of heavy metals (in concentrations of substances inhibiting luminescence by 50%) resulted in damaging effects upon bacteria: a lot of cells had damage of membranes due to changes in their permeability. It was found that the high concentration of EDTA and toluene decreased the luminescence and caused the condensation of DNA-fibrils and the cell damage after long-term and short-term action. The low concentration of EDTA and toluene did not decrease the bacterial luminescence; the noticeable damage of cell membranes did not take place during short-term treatment. However, the long action of these substances changed the membrane permeability resulting in increased sensitivity of bacterial luminescence to some toxic substances.

Analysis of river water by bioluminescent biotests.

The bacterial bioluminescence has high sensitivity to the action of various inhibitors of biological activity. The lyophilized luminous bacteria Photobacterium phosphoreum (Microbiosensor B17 677F) and luminous strain Escherichia coli (Microbiosensor EC) from the Culture Collection IBSO were used to create bioluminescent biotests. They have been applied in ecological monitoring to determine the overall toxicity of the Yenisei and Angara Rivers and some water sources of Altai Territory. As a rule the heaviest pollution of water in studied rivers was registered near cities and settlements. The luminous bacteria biotests are simple and convenient in work, standardized and quantitative, have rapid response to actions of different substances and high sensitivity to environmental pollutants. It takes less than 30 min to do the biotest (the other biotests take 48--96 h).

Growth and luminescence of luminous bacteria promoted by agents of microbial origin.

The examination of four species of luminous bacteria Photobacterium leiognathi, Photobacterium phosphoreum, Vibrio fischeri and Vibrio harveyi has enabled us to reveal some nutrient medium components effecting growth, luminescence intensity and luciferase synthesis. These agents are nucleic components (nucleotides, nucleotides and amine bases), amino acids and vitamins, which are part of hydrolysates from the biomass of various lithotrophic microorganisms, hydrogen-oxidizing, iron-oxidizing and carboxydobacteria. The effect of promoting agents essentially alters the physiological state and ultrastructure of the cells of luminous bacteria and increases luciferase biosynthesis two- to three-fold compared to a control.

Effect of carbon monoxide on metabolism and ultrastructure of carboxydobacteria.

Growth of Seliberia carboxydohydrogena was inhibited by CO at 10 to 40% (v/v), resulting in increased substrate utilization and enhanced synthesis of cytochromes and cyclopropane and saturated fatty acids. The bacteria showed increased formation of new membrane structures, with pronounced folding of their cell walls.

Bioluminescent method in studying the complex effect of sewage components.

The inhibition of bacterial luminescence has been used in testing industrial enterprises sewage. The toxicity of the sewage is less than the total toxicity of separate components due to neutralization of quinone products of polyphenol oxidation in the reactions with the other phenol components of sewage. Toxicity increase is due to their influence on the cell membrane. Studies of cell ultrastructure confirm this fact. The studied mechanism of the complex effect allowed a more accurate forecast of the ecological situation during the discharge of phenol compounds and metals. It also showed the necessity of taking into account the complex effect of sewage components on contaminant discharge into water reservoirs.

Immunoelectronmicroscopic study of the nucleoid structure of hydrogen bacteria.

Electron microscopical studies of the nucleoid structure of hydrogen bacteria using ultrahin sections and spread DNA from bacterial cell lysates revealed a different DNA packaging in the cell. A compact state of the major part of DNA at all growth stages and stability of nucleosome-like structures were shown. The use of antibodies to HU protein of E. coli labelled by protein A-colloidal gold demonstrated the immunological relationship between HU protein of E. coli and histone-like proteins of Alcaligenes eutrophus and their possible role in the nucleosome-like DNA packaging in procariotic genome.

[Deposition of potassium and water in the brown adipose tissue of cold-adapted rats]. / Deponirovanie kaliia i vody v buroi zhirovoi tkani u adaptirovannykh k kholodu krys.

The rats adapted to 0-4 degrees C increased water content in the BAT. The Na and K intracellular contents was unaltered during first month of the adaptation. K content in the BAT was considerably increased by the 10th and 18th weeks of the adaptation. A positive correlation between Na and K contents was revealed in the BAT of both adapted and unadapted rats. The increase of K concentration and the decrease of Na/K ratio suggest that, in prolonged adaptation, the Na-pump power grows more than is necessary for compensation of the high fluxes of cations through the cell membranes in the BAT.

[Ultrastructural organization of marine luminescent bacteria and their mutants]. / Osobennosti ul'trastrukturnoi organizatsii morskikh svetiashchikhsia bakterii i ikh mutantov.

The aim of this work was to study the submicroscopic organization of luminescent bacteria belonging to the genera Photobacterium and Lucibacterium as well as that of their "dark" mutants incapable of luminescence. The ultrastructural organization of all studied bacteria is typical of gram-negative species. The luminescent bacteria are characterized by the presence, in their cytoplasm, of osmophilic formations 22--110 nm in size. The cells of "dark" mutants accumulate volutin and contain complex membrane systems which are related to decelerated growth of the cultures.