RESEARCH AREAS AND PRACTICAL APPLICATION OF LUMINOUS BACTERIA

The research work in the collection done along the following lines:

·        Guaranteed preservation of viability and initial properties of the Collection gene pool;

·        Development of optimum methods of  gene pool storage;

·        Study of physiological, biochemical, and morphological properties of collection strains;

·        Screening of strains producing luciferases, oxidoreductases and other target products, including, using of E.coli as heterologic owner of such products;

·        Study of distribution of these microorganisms in waters of the World ocean and their taxonomy identification;

·        Study of population heterogeneity and dissociation under various cultivation conditions;

·        Research of structural organization and fatty acid composition of luminous bacteria.

·        Research of luminescent system localization;

·        Finding out of the mechanism of toxicant action on the cell and the population as a whole;

·        Research of activity and stability of enzymes of the luminescence reaction;

·        Culture collection information network, computer databases of microorganisms;

·        Development of high-sensitive toxicity biotest for monitoring of a environment;

·        Manufacturing of a complete set of reagents for the bioluminescent analysis on the basis of the isolated bienzyme system of luminous bacteria.

In the IBSO Collection, work is now underway toward creating the database within the framework of the international program Microbial Strain Data Network (MSDN) and the State Research - and - Development Project "Aids to Investigations in Physico-Chemical Biology and Biotechnology", "Microbial diversity" (the part "Culture collection of microorganisms"). The IBSO Culture Catalogue of Luminous Bacteria is available on disk in the MS-DOS format for personal computers.  The English version of the Catalogue is Accessible through the international network MSDN (News Letters, 1994; Kalakoutskii et al., 1996; Rodicheva et al., 1996). The IBSO Collection designs the database about features of natural and transgenic bacteria carring marker lux-gene (Medvedeva et al., 2001). Participation of the Collection in the MSDN makes its activity more effective and information exchange more expeditions.

Luminous bacteria are a convenient object to be used in solving general biological problems, genetic and genetic-engineering researchs. The collection of luminous bacteria can serve as a basis for researching the strain-superproducers of luciferases, NADH:FMN-oxidoreductase, decarboxylase, endonuclease of restriction, chitinase producers [Ruzgiene et al, 1990, 1991, 1992, Kolykhalov et al, 1991, Repin et al., 1993, 1995a,b, 1997a,b, Vydryakova et al., 1993]. They can be used successfully to determine toxicity of microquantity of different substances [Kratasyuk, Gitelson, 1987, Popova et al., 1993, Kudryasheva et al., 1996].

Strains from the luminous bacteria collection are used as a basis for preparation of lyophilized bacteria for measurements of the integral biological toxicity of different substances: heavy metals, phenols, narcotics, toxins, pesticides, medicines etc.  The method is based on quenching the bioluminescence of luminous bacteria by different toxicants.

The microbiosensor based on freeze-dried luminous bacteria has been used successfully to control the water quality in different water bodies, such as rivers, waste water of wood-pulp and paper production facilities and other industries [Stom et al., 1986, 1992, Popova et al., 1993, Geel et al., 1993, Kudryasheva et al, 1996, Kuznetsov et al., 1990, 1997-2000, Kratasyuk et al., 1991, 1997, 1998, 2001]. The preparation produced on the basis of freeze-dried luminous bacteria in comparable with the "Microtox" preparation produced by the Beckman but less expensive.

Unconventional techniques for cultivating of luminous bacteria [Rodicheva et al., 1973, Zavoruev et al., 1982, Gitelson et al., 1984] and deriving highly purified bacterial luciferases from four species (Photobacterium phosphoreum, Photobacterium leiognathi, Vibrio fischeri and Vibrio harveyi) have been developed [Mezhevikin et al, 1983, Bondar et al., 1988, Tyulkova, 1990, Sukovataya, Tyulkova, 2001].

At present, biotechnology department produces sets of analytical luminescence reagents based on the coupled enzyme system: luciferase - NAD(P)H:FMN-oxidoreductase isolated from luminous bacteria [Tyulkova, Illarionov, 1994, Kuznetsov et al., 1997]. A kit of reagents for analytical bioluminescence (KRAB) can be used in medicine, ecological monitoring, and for quality inspection of newly synthesized substances.

The diagnostic kit (KRAB) can be used for determination of NAD-dependent dehydrogenases, proteases and antiproteases in blood plasma and serum, platinoids etc [Petushkov et al., 1983, 1984, 1985, 1987, Kratasyuk et al., 1985, Gitelson et al., 1985, Kratasyuk, 1995]. Institute of Biophysics offers an immobilized reagent  for bioluminescent assay (IRBA) [Kratasyuk et al., 1994]. Using the kit of reagents for analytical bioluminescence (KRAB) and the hydrogenase of hydrogen bacteria, researchers of the Institute of Biophysics have developed methods for determining hydrogenase activity and the concentration of dissolved hydrogen [Petushkov V.N., Guseinov O.A., 1992].

Researchers of the Institute of Biophysics have developed the bioluminometers, a devices for measuring luminescence intensity.  Such devices are now produced by the Institute together with the "Nauka" Special Design and Development Office (Krasnoyarsk).

Bacterial bioluminescence can be used as a basis for making visual aids and creating tasks for laboratory work at schools and universities. Bioluminescence methods may be employed in investigating fine details of gene structure and functioning, conjugated chains of enzymatic reactions, determining the quantity of substrates, cofactors, etc. [Andrade, 1995; Kratasyuk et al., 1995, 1997, Kratasuyk, Kudinova, 1999]. Besides its evident educational advantages, the cold luminescence of life is aesthetically and emotionally attractive.

Remarkable opportunities offered by the use of luminous bacteria in the various areas of biology, their ecological and technological applications permit us to state that they deserve close attention of biologists and other experts of the adjacent areas of knowledge.