Svetlana E. Medvedeva

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.

The Chemical Basis of Fungal Bioluminescence

Many species of fungi naturally produce light, a phenomenon known as bioluminescence, however, the fungal substrates used in the chemical reactions that produce light have not been reported. We identified the fungal compound luciferin 3-hydroxyhispidin, which is biosynthesized by oxidation of the precursor hispidin, a known fungal and plant secondary metabolite. The fungal luciferin does not share structural similarity with the other eight known luciferins. Furthermore, it was shown that 3-hydroxyhispidin leads to bioluminescence in extracts from four diverse genera of luminous fungi, thus suggesting a common biochemical mechanism for fungal bioluminescence.

New insights in to the treatment of myocardial infarction.

This study investigated the effects of the L‐17 compound of the group of substituted 5R1, 6H2‐1,3,4‐thiadiazine‐2‐amines on the inflammatory cellular infiltration and myocardial remodelling which occurs after acute myocardial infarction (MI) in rats. The study is based upon recent clinical and experimental work which demonstrated the role of local and systemic inflammatory reactions in postinfarction remodelling. Acute MI in rats was induced by left coronary artery coagulation. Animals were sacrificed on day one, five and seven after MI induction. The myocardiumal samples were taken from all parts of the heart and examined by histology. This included areas of infarction, infraction and areas that were peri‐infarctiom and left ventricular areas distant from the damaged tissues. Serum activity of creatine phosphokinase (CPK), aspartate aminotransferase (AST), isoenzymes 1 and 2 and lactate dehydrogenase (LDH1‐2) were investigated on the same three days, before and in the process of MI development was investigated (at days 1, 5 and 7). The L‐17 compound to not only decreased the area of initial infarction but also changed the pattern of inflammatory reaction in the affected myocardium fundamentally. Laboratory studies of effects of L‐17 compound on the development and course of experimental MI showed that administration decreased blood AST and CPK levels significantly and provided useful the data about the correlation between the activity of these enzymes and the dimensions of the significantly necrotic area. In this model of experimental MI the use of the L‐17 compound induced led to the replacement of the exudative destructive inflammation that is seen under standard conditions with a more cellular “productive” pattern of inflammation, with associated reduction in initial necrosis area and the, decrease in myocardial ischaemia and reperfusion injury may account for the accelerated repair process.

Isolation of luminescence system from the luminescent fungus Neonothopanus nambi

56 This study is devoted to the problem of isolation of the lighttemitting system able to luminesce in vitro from the fungus Neonothopanus nimbi. The study was performed with the mycelium of the luminous higher fungus N. nambi, inhabiting the tropp ical forests of South Vietnam. [1] The fungus culture was kindly provided for experiments by Vietnamese researcher Dao Thi Van (private collection of strains To obtain fungal biomass, the mycelium was cultured in Petri plates in a liquid nutrient medium by the techh nology described earlier [2]. The grown mycelium was taken from the Petri dish and washed with deionized (DI) water (MilliiQ system, Millipore, United States) to remove the residual components of the nutrient medium and exometaboliltes. After washing, the remaining water was removed from the mycelial biom ass with filter paper. The isolation of the luminescent system of N. nambi mycelium included the following steps, which were carried out at 0–4°C. Mycelium washed with DI water was ground in the cold with scissors and transferred to a beaker placed in an ice bath. The bioo mass was poured with cold 0.1 M phosphate buffer (pH 7.0) supplemented with 0.1–1.0% BSA (Serva, Germany) in the ratio 1 : 5 (wet biomass weight : buffer volume). The biomass was destroyed with a Volna ultrasonic disintegrator (Russia). Sonication was perr formed at a power of 200 W three times for 5 s at 11min intervals. The homogenate was transferred into chilled tubes and centrifuged at 48 000 g for 30–60 min in an Avanti ® JE centrifuge (BeckmannCoulter, United States). The pellet was discarded, and the supernatant was either used immediately for study (in this case, it was stored at 4°C) or immediately frozen at –20°C and stored at this temperature. The luminescence of the supernatants was meaa sured using a BLM 8801 luminometer (Nauka Special Engineering and Design Department, Krasnoyarsk, Russia) calibrated using the Hastings–Weber radioacc tive standard [3] (one luminescent unit was 10 8 phoo tons per 1 s). The signals were recorded using an LKB 2210 recorder (LKB, Sweden). It was found that supernatants isolated from the mycelium of the luminous fungus N. nambi by the method described above emitted long luminescence (Fig. 1). This fact allowed us to conclude that a selff sufficient luminescent system that ensures luminess cence in vitro was isolated from this fungal species. After filtering the supernatant through a membrane with an …

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.

Chemiluminescent emission of tissues of fruit bodies of higher fungi

105 Among the dozens of thousands of species of higher fungi known to date, more than 80 species possess bioluminescence—the ability to emit light that is visii ble with the naked eye. This ability was found in the to note that the luminescent species phylogenetically coexist with the nonluminescent ones. Sometimes even one taxonomically defined species includes both luminescent and nonluminescent forms [2, 3]. Such a mosaic distribution of bioluminescence suggests that the ability to luminesce has occurred in the kingdom of fungi repeatedly and independently and that its evolutionary basis is a fundamental bioo chemical process, a small deviation in which (even in one or two stages of the metabolic chain) gives rise to luminescence. However, such a process remains unree vealed as yet. The absence of intermediate forms makes it highly difficult to elucidate the evolutionary pathway of the emergence of bioluminescence in the kingdom of fungi. Although the notions on the mechanism of light emission by fungi are far from complete [3, 4], it is obviously distinct from the understood emission mechanisms in animals and bacteria. Weak chemiluu minescence is characteristic of animal tissues [5]. It is known that the major source of luminescence in animals is lipid peroxidation [6] and that chemilumii nescence in plants is related to the photosynthesis system [7]. We studied the emission of nonbioluminescent higher fungi. Studies were performed with different species of higher fungi growing in forests of the Eastt ern Siberian region of Russia (Krasnoyarsk Krai). The objects of the study were 150 samples of fungi collected in forests in vicinities of Krasnoyarsk in summer 2011. The collections were representatives of five orders, 15 families, 21 genera, and 13 species. As many as 136 samples of the collected material were identified to the genus level and 35 samples to the species level (table). We measured luminescence of fungal fragments taken from different parts of the fruit body. Measuree ments were performed with the Glomax 20/20 lumii nometer (Promega, United States), which was calii brated using the Hastings–Weber radioactive standard [8] (2.7 × 10 3 quanta/s was taken as one unit of lumii nescence). The emission of each sample was recorded for 10 s. Signals exceeding the background level by at least 5 times were taken as reliable. Then, each sample was air dried to a constant weight to calculate the spee cific luminosity per unit mass. The table shows the …

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).

Heterogeneity of the Populations of Marine Luminescent Bacteria Photobacterium leiognathi under Different Conditions of Cultivation

Manifestation of pleiotropic effects in the isogenic variants of the luminescent bacterium 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 autoinducer 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 inherited variants of P. leiognathi.

Biotesting of effluent and river water by lyophilized luminous bacteria biotest

Waters of the Yenisei River, certain rivers and lakes of the Altai Territory, and effluents of some industrial factories in Krasnoyarsk were studied by luminous bacteria biotest Microbiosensor B17 677F. The lyophilized luminous bacteria Photobacterium phosphoreum from the IBSO collection were used to design this biotest. The bioluminescent test is based on bioluminescence quenching resulting from the action of water samples on luminous bacteria. The test results indicated locations and zones of impaired water quality. The heaviest pollution of water in the Yenisei River was recorded in the zones 0–116 km downstream from Krasnoyarsk (Krasnoyarsk and satellite towns). The effluents of most factories were found to be toxic. Underground and surface waters of some areas of the Altai Territory had different toxicity levels; there were deviations from the norm in most water samples taken from the different lakes and rivers. The data from this study show that the luminous bacteria biotest is simple and convenient, and that the results obtained are within acceptable levels of accuracy for the evaluation of the toxicity of effluent and river water. It takes no more than 30 min to do the biotest. It can be used in ecological monitoring like the Microtox toxicity test.

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 lower, 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.