[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.

[Assessing the anthropogenic impact on lake Shira from antibiotic resistance of heterotrophic bacteria by neural networks methods].

A general approach to assessing the anthropogenic impact on lake ecosystems is proposed and exemplified for the case of Lake Shira (Republic of Khakasia, Russia). The impact strength is estimated by applying neural network-based methods to samples of data on interdependent marking features of autochthonous and allochthonous bacteria isolated from the lake in 1997-2001. The proposed combination of analysis methods makes it possible to determine the state of an ecosystem from both small- and large-size samples of data having complex interrelations.

[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.

[Influence of the salt concentration in culture on the copy number of plasmid pSH1 in cells of Micrococcus sp. 9].

The study of heterotrophic bacteria isolated from the brackish waters of Lake Shira has shown that some of them contain plasmid pSH1 of approximately 2.7 kb in size. The number of plasmid copies in plasmid-containing strains cultivated at a minimal concentration of sodium chloride is found to be low, whereas the subculturing of these strains at high salt concentrations increases the plasmid number. The role of natural pSH1 plasmid in the osmotolerance of host bacteria is discussed.

[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.

[Formation of structured communities by natural and transgenic naphthalene-consuming bacteria].

This study concerns the formation of structured communities by pure cultures and binary associations of Pseudomonasfluorescens transgenic strains and natural heterotrophic bacterial species in naphthalene-containing media with various osmotic pressures. It was shown that cells of P. fluorescens strain 5RL, harboring a recombinant construct in the chromosome, were more resistant to the combined action of the stress factors under study than P. fluorescens 82/pUTK21, harboring a recombinant construct within a plasmid. Natural P. fluorescens 1 strains, particularly Vibrio sp. 14, were more viable at high osmotic pressures and naphthalene concentrations. Experiments with the combined introduction of transgenic and natural bacterial strains at high osmotic pressures demonstrated the stable coexistence of bacterial associations in biofilms, independent of naphthalene concentration. Strains considered for introduction into the environment for bioremediation should be assessed with regard to their susceptibility to the combined effect of anthropogenic and natural stress factors. The design of bacterial associations for the same purpose should take into account the effect of factors important for their survival in polluted areas.

[Hygienic aspects of urinary system developing pathology in children (review)]. / Gigienicheskie aspekty formirovaniia patologii organov mochevoi sistemy u detei (obzor).

The paper deals with the impact of environmental factors of industrial cities and towns on children's health. Particular emphasis is laid on urinary tract pathology since the kidney is a xenobiotically exposed target organ. The main criteria for a risk of environment-induced diseases are ambient air pollution, low drinking water quality, soil heavy metal pollution, etc. The review covers the pathogenetic mechanisms and clinical features of the manifestation of renal diseases in children living in the regions with different heavy metal-loaded environment.

[Distribution of heterotrophic bacteria in Shira lake]. / Raspredelenie geterotrofnykh bakterii po akvatorii ozera Shira.

The study of the horizontal and vertical distribution of heterotrophic bacteria in brackish Lake Shira in summer periods showed that mesophilic bacteria dominated in all areas of the lake, whereas psychrotolerant bacteria dominated in the metalimnion and hypolimnion of its central part. Nonhalophilic bacteria were mostly mesophilic and dominated in coastal waters. Most psychrotolerant bacteria were able to grow in the presence of 5-10% NaCl. Heterotrophic bacteria isolated in different regions of the lake were identified to a generic level. The isolates were classified into autochthonous and allochthonous microorganisms on the bases of their distribution pattern in the lake water, halotolerance, and ability to grow at low temperatures.

[Conflict: induction-inhibition of transgene bacteria luminescence in studying expression of lux-genes]. / Konflikt: induktsiia-ingibirovanie liuminestsentsii transgennykh bakterii v izuchenii ékspressii lux-genov.

The relationship between the induction of the luminescent operon of lux-genes fused with the naphthalene and salicylate degradation genes and the inhibition of light emission caused by these compounds was studied. The quantitative correlations between these processes manifest themselves in the fact that light intensity linearly increased in a narrow concentration range of the inductor and then decreased due to the inhibition of the luminescence reaction itself, which is not related to the regulation of expression of lux-genes.

[The colony architectonics in Bacillus subtilis 2335]. / Osobennosti stroeniia kolonii Bacillus subtilis 2335.

Colonies grown from vegetative B. subtilis 2335 cells had a standard structure, with bacillar cells occupying the whole colony volume. At the same time, the colonies of this bacterium grown from germinated spores had an abnormal structure characterized by the location of cells in a surface layer 100-200 microns thick at the colony boundary with the air. The glycocalyx of the colonies grown from spores was characterized by a wetting angle theta e of 120 degrees-160 degrees, whereas that of the colonies grown from vegetative cells had an angle theta e as low as 5 degrees-30 degrees. It is suggested that spores and vegetative cells follow different strategies of substrate colonization and that the architectonics of bacterial colonies is determined by the physicochemical properties of the glycocalyx.

[Coexistence of transgenic Escherichia coli strains and natural microorganisms in experimental aquatic microcosms]. / Sosushchestvovanie transgennogo shtamma Escherichia coli s prirodnymi mikroorganizmami pri sovmestnoi introduktsii v vodnye mikrokosmy.

In experimental aquatic microcosms (AMCs), the population of the Escherichia coli strain Z905 harboring the recombinant plasmid pPHL7 (AprLux+) was found to gradually accumulate AMC-adapted cells, which retained the plasmid but differed from the original cells in some biochemical and physiological characteristics. Both the original and the AMC-adapted E. coli cells could coexist with the native AMC microflora for a year or longer. When introduced into AMCs together with native pseudomonads, the AMC-adapted E. coli Z905-33 (pPHL7) cells were more competitive than nonadapted cells.

[Mineral salt-dependent inhibition of light emission from the luminescent microorganism Escherichia coli Z9051]. / Zavisimoe ot mineral'nykh solei ingibirovanie svecheniia liuminestsentnogo mikroorganizma Escherichia coli Z9051.

The influence of some mineral salts on the recombinant strain Escherichia coli Z9051 was investigated. It was shown that the composition (NaCl, Na2SO4, MgCl2 and MgSO4) and concentration (5 and 10%) of the salts substantially affect the expression of genes for the luminescence system of light-emitting bacteria cloned in the plasmid under the control of the lac-promoter. In some cases, the luminescence level of the microorganism in the presence of salts was similar to the luminescence level under catabolite repression by glucose, the more strong influence of the salts exceeding the effect of catabolite repression. The possibility of adaptation of the genetically modified microorganism to the salinity factor is discussed.

[Stability of recombinant plasmids in transgenic microorganism cells under various environmental conditions]. / Stabil'nost' podderzhaniia rekombinantnykh plazmid v kletkakh transgennykh mikroorganizmov pri raznykh usloviiakh sushchestvovaniia.

The copy number of R plasmids weakly depends on the selective pressure of the respective antibiotic but does depend on the physiology of the host species and the type of plasmids and cloned genes, whose expression leads to a further load on the biosynthetic apparatus of cells. The last factor is critical in the maintenance of recombinant plasmids in transgenic microorganisms.

[Factor of salinity and adaptive capacity of recombinant strains of Escherichia coli and Bacillus subtilis]. / Faktor solenosti i adaptivnye vozmozhnosti rekombinantnykh mikroorganizmov Escherichia coli i Bacillus subtilis.

Effect of different concentrations of salts on natural and recombinant strains of Bacillus subtilis and Escherichia coli was studied. The recombinant strain of B. subtilis was found to be more osmotolerant than the wild-type strain of this bacterium, whereas the opposite situation was observed for the recombinant and wild-type strains of E. coli. Some salts exerted a bacteriostatic effect on E. coli and B. subtilis. The adaptive capacity of recombinant strains depended on the number of plasmid copies in the cells. The introduction of recombinant bacteria into model ecosystems resulted in the generation of their variants with increased osmotolerance.

[The study of heterogeneity of plasmid-bearing and plasmid-f ree populations of Bacillus subtilis under different environmental conditions]. / Izuchenie geterogennosti v populiatsiiakh plazmidosoderzhashchego i besplazmidnogo shtammov Bacillus subtilis v raznykh usloviiakh su shchestvovaniia.

The population heterogeneity of recombinant and plasmid-free Bacillus subtilis strains introduced into aquatic microcosms was studied. After introduction, the population of the plasmid-free strain B. subtilis 2335 in microcosms has long been represented by both vegetative cells and spores, whereas, already ten days after introduction, the population of the recombinant strain B. subtilis 2335/105 (Km[symbol: see text]nf+) was represented only by spores. The number of plasmid copies in the spore isolates of the recombinant strain was the same as before introduction, but the plasmid abundance in the vegetative isolates of this strain decreased. The isolates of B. subtilis 2335/105 obtained from microcosms and the variants of this strain obtained by ten successive subcultures on M9 and 0.1 x M9 media with and without kanamycin (Km) differed in the number of plasmid copies, Km resistance, and maximum biomass yield during batch cultivation. Irrespective of the presence of Km, more than 50% of the variants subcultured on M9 medium showed reduced plasmid abundance. At the same time, about 70% of the variants subcultured on 0.1 x M9 medium with Km and 90% of the variants subcultured on the same medium without Km retained the initial number of plasmid copies. The variants subcultured on media with Km retained the initial biomass level. In more than 70% of the variants isolated from media without Km, the biomass yield increased.

[The comparative growth characteristics of variants of a recombinant Escherichia coli strain and of the representatives of aboriginal microflora in aqueous microecosystems under selective and nonselective conditions]. / Sravnitel'naia kharakteristika rosta variantov rekombinantnogo shtamma Escherichia coli i prestavitelei aborigennoi mikroflory vodnykh mikroékosistem v selektivnykh i neselektivnykh usloviiakh.

We have studied the kinetic characteristics of several isolates of Escherichia coli Z905 recombinant strain after introduction of the strain into model aquatic ecosystems. Most E. coli Z905 isolates grown as batch cultures under selective conditions (0.5 microgram/ml ampicillin) showed better kinetic characteristics of growth than did related species of native microflora, which originally populated laboratory microcosms.

[Change in regulation of catabolite-dependent lux-operon, cloned in a recombinant plasmid, under the effect of environmental factors]. / Izmenenie reguliatsii katabolitzavisimogo lux-operona, klonirovannogo v rekominantnoi plazmide, pod vozdeistviem faktorov vneshnei sredy.

Expression of the lux-genes cloned on the recombinant plasmid pPHL7 (AprLux+) in Escherichia coli z905 cells was studied in various environments, including model aquatic ecosystems. Expression of the lux-genes strongly depended on the nutritional status of the medium. In particular, the cultivation of cells in nutrient-rich medium favored the maintenance of the initial level of expression of the lux-operon, whereas nutrient limitation induced recombinant cell variants with an impaired control of the catabolite-dependent lux-operon. On the other hand, long-term laboratory cultivation of recombinant strain in nutrient-deficient media or its long-term life in model aquatic ecosystems led to the accumulation of cells with a stringent control on the cloned lux-genes in the bacterial population. The presence of the selective factor (ampicillin) in the medium had no significant effect on the expression of the lux-operon.

[Introduction and long-term storage of recombinant luminescent Escherichia coli strain Z905 in laboratory water microecosystems]. / Introduktsiia i dlitel'noe sokhranenie rekombinantnogo liuminestsentnogo shtamma Escherichia coli Z905 v laboratornykh vodnykh mikroékosistemakh.

We studied preservation of recombinant Escherichia coli strain Z905 (AprLux+) in liquid microecosystems (LME) after the introduction. E. coli cells were shown to remain viable and preserve the ability to express the cloned lux genes for a long time (more than a year) in LME. The majority of the clones have reduced efficiency of the expression due to either changed regulation of the lux operon or decreased number of copies of the plasmid. These mechanisms could be realized either independently or simultaneously depending on LME conditions. We have exposed the major factors affecting the metabolic activity of the E. coli strain Z905 (AprLux+) introduced into model ecosystems and the level of expression of the cloned genes.