[Detection of uncultivable bacteria forms in probiotic lyophilized preparations].

AIM: Detection of viable but uncultivable forms among lyophilized cells of commercial probiotics. MATERIALS AND METHODS: 9 series of probiotic preparations (colibacterin, bificol, bifidumbacterin, bifiform) with expired (up to 30 years of storage) or valid shelf life were objects of the study. Total quantity of the bacteria was calculated under the microscope in Goryaev chamber, the number of viable cells was determined in luminescence microscope after staining by an array of fluorescent dyes, the quantity of CFU/ml was evaluated by method of seeding into the respective solid or semi-liquid cultivation medium. RESULTS: Juxtaposition of the specified parameter values allowed to establish that in lyophilized preparations of colibacterin with expired shelf life the amount of viable but not forming colonies (uncultivable forms) cells varied from 4.13 to 99.73% depending on date of production. For bifidumbacterin with unexpired shelf life live cells constituted 95.45 and 70.73%. The amount of viable bifidobacteria forming colonies was on the level of 100 and 50%, respectively. In bifidopreparations micro colonies that may possibly be formed spontaneously by awakened uncultivable forms were noted. A possibility of growth stimulation of these cells by 1 and 10% aminopeptide was shown. CONCLUSION: The presence of uncultivable forms in lyophilized preparations of probiotics was proven experimentally.

[Features of dynamics of growth and formation of uncultivable forms in Lactococcus lactis].

AIM: Study strain differences in dynamics of viability and formation of uncultivable forms of Lactococcus lactis. MATERIALS AND METHODS: 3 strains of L. lactis--MSU, 729 and F116 were used in the study. Uncultivable forms were obtained by prolonged incubation of cultures of lactococcus in synthetic medium under conditions of carbohydrate deprivation. Medium was inoculated by cultures grown in Elliker broth, washed by saline and without washing. The populations obtained were incubated at 30 degrees C without mixing for 4 months. Samples of cultures were studied periodically for viability and cultivability. RESULTS: In cultures obtained by using unwashed inoculate active growth of quantity of bacteria in the first days after seeding was noted. Speed of formation of uncultivable forms is the faster the higher the level of metabolic activity of cells in the population. A fact of phenotypic dissociation in lactococcus culture under stress was detected. CONCLUSION: The speed of formation of uncultivable forms of L. lactis, as well as stability of their existence may probably depend on the strain, cultivation conditions and metabolic activity of cells in the population.

[Changes of expression levels of innate immunity signaling receptors during Candida albicans infections in vitro and in vivo].

AIM: To study levels of expression of Toll-like receptors genes in response to Candida albicans antigens in vitro (using Vero cell line as well as mononuclear cells) and in vivo (using cervical canal cells of pregnant women). MATERIALS AND METHODS: Test-systems for measurement of expression levels of such genes as TLR 1, TLR2, TLR6 as well as system for quantitative measurement of tumor necrosis factor a level, all of which were developed earlier, were used. RESULTS: It was shown that antigens of C. albicans stimulated early increase of expression of innate immunity genes both in studied in vitro models and in cells of cervical canal of pregnant women with candidosis. CONCLUSION: Results of performed study allow to suggest that activation of innate immunity factors resulted from Candida albicans infection.

[The detection of bacteriogenic causes of opportunistic infections].

In order to select bacteriocin producing strains the screening of microorganisms from clinical material was perfomed. The delayed antagonism method was used. 154 strains were screened, 93 (60%) cultures produced antagonistic substances. Micrococcus luteus NCTC, 2665, Candida albicans ATCC 885-653, Escherichihia coli 168/59, Pseudomonas aeruginosa 27/99, Klebsiella peumoniae 1954 were used as test-cultures, Pseudomonas aenruginosa and Staphylococcys aureus were the most frequent species producing antibacterial substances and their products were of high potency, with a wide spectrum of antimicrobial activity.

[Study of genetic control of pediocins synthesis].

Plasmid profiles of 2 strains of Pediococcus acidilactici and 3 strains of P. pentosaceus were studied. Synthesis of bacteriocins in 2 strains of P. acidilactici was coded on the plasmid which was confirmed by experiment with elimination of plasmids and subsequent electrophoregram, which demonstrated synchronous disappearance of bacteriocins synthesis characteristic in plasmid-lacking variants. In 3 strains of P. pentosaceus plasmids were not found which points to chromosomal localization of genes for bacteriocins production.

[Biologic characteristics of new Bacillus subtilis isolates].

Characteristics of new Bacillus subtilis isolates from the environment and had high antagonistic activity were studied. Absence of virulence and toxin production allows to regard them as potential probiotic cultures. Growth media for cultivation of B.subtilis were chosen according to the principle of individual adequacy in system, which give the ability to optimize processes of cultivation, obtaining of inhibitory substances and antagonistic activity spectrum determination.

[Molecular basis of bacteriocins production and activity].

Bacteriocins--bacterial proteins or peptides--are envisaged as candidates for the next generation of effective antimicrobials. Analysis of characteristics of natural and genetically engineered bacteriocins with regard to the molecular basis of their production and activity has been performed. Most bacteriocins have narrow spectrum of the inhibitory activity. Some of the broad-spectrum bacteriocins have circular molecular structure (C- and N-terminals of the aminoacid chain are joined by a peptide sequence). Fixed position of molecules' ends possibly accounts for the ability of the proteins to bind with various receptors on the surface of the target cells. Genes encoding bacteriocins and functionally associated proteins can be expressed in heterologous cells including eukaryotic cells. Also there were reports of changing bacteriocin characteristics by the use of site-specific mutagenesis.

[Biotechnological conditions of the synthesis of bacteriocins].

In this review information on the cultivation of bacteriocin-producing microorganisms belonging to different taxonomic groups are presented. The data on the influence of the most important biotechnological parameters (nutrient medium, temperature, pH, phase of growth, etc.) on the synthesis of bacteriocins are given.

[Interaction of Staphylococcus aureus alpha-toxin with eukaryotic cells and their target].

In this review information on S. aureus alpha-toxin is expounded. The data on the targets of the action of the toxin on eukaryotic cells, its activity with respect to different membrane structures, effects established in the interaction of toxin and cell elements, as well as of its biological action on the macroorganism, are presented. Information on the factors inactivating the cytolytic action of the toxin is given.

[Bacillus subtilis protease: isolation, immobilization and properties].

During the cultivation of B. subtilis strain 3H under optimum conditions (adequate nutrient medium, seed culture, temperature, the level of dissolved oxygen) protease was produced. Protease could be obtained in the purified form by means of gel chromatography and ultrafiltration. The isolated protease was immobilized on polyglucin and stabilized by intramolecular cross-linking with the use of glutaraldehyde. The comparison of native protease modified with polyglucin and glutaraldehyde, as well as with polyglucin, revealed advantages in the stability of the latter.

[PCR system for the detection of the RNA of hepatitis A virus]. / Razrabotka i ispytanie PTSR test-sistemy dlia detektsii RNK virusa gepatita A.

The detection of the causative agent of hepatitis A in the patient's body is the necessary element of the diagnostics of this disease. In this work the PCR system for the analysis of the RNA of hepatitis A virus in the patient's blood is presented and characterized. The method of the detection of the RNA of hepatitis A virus is based on the "nest" principle and consists of two consecutive reactions. In the first reaction the reverse transcription and amplification with the external pair of primers are carried out. The product thus obtained is used as material for the second reaction with the internal pair of primers. This method was used for the study of 44 blood samples from hepatitis A patients and 23 blood samples from healthy donors. The detection rate of the RNA of hepatitis A virus in blood samples from the patients was 82%. Viral RNA could be detected in the serum in 72% of cases, both in the serum and in mononuclear blood cells in 20% of cases, in mononuclear blood cells only in 8% of cases.

[Properties of the isolated Bacillus subtilis strains and their influence on the intestinal microflora of experimental mice]. / Svoistva vydelennykh shtammov Bacillus subtilis i ikh vliianie na intestinal'nuiu mikrofloru éksperimental'nykh myshei.

The cultural, physiologo-biochemical adhesive and antagonistic properties of B. subtilis strains with good prospects for use as biotherapeutic preparations were studied. For further studies B. subtilis strain No. 1719 was chosen. In experiments on non-inbred white mice the animals were treated by the preparation Cifran used for their selective decontamination from opportunistic microflora and for the creation of the state of dysbiosis. The influence of the spore-forming microbe on the parietal microflora of the large intestine of the animals was shown. Reliable data on the changes in the number of microorganisms (CFU/ml) per 1 cm of the surface of the large intestine were established. As markers making it possible to evaluate the action of biotherapeutic and other medicinal remedies, easily determinable ratios of lac+/lac- of bacteria and Staphylococcus aureus/Staphylococcus spp. was proposed.

[Study of the effectiveness of the method of the nest polymerase chain reaction and the field of its use in hepatitis A diagnosis]. / Issledovanie éffektivnosti i oblasti primeneniia metoda gnezdovoi PTSR pri diagnostike gepatita A.

To determine the causative agent of hepatitis A in the blood of patients, we developed the method of the nest polymerase chain reaction (PCR) with reverse transcription. For the comparative evaluation of the diagnostic efficacy of the nest PCR and the enzyme immunoassay (EIA) serum samples and mononuclear blood cells obtained from 15 patients with diagnosed hepatitis A and from 33 patients having signs and carrying markers of other form of hepatitis were analyzed. On the whole, the method of PCR confirmed the results of EIA and in some cases exceeded it in sensitivity. In addition, hepatitis A virus RNA was detected in the blood of a clinically healthy person having had a contact with a hepatitis A patient. The PCR data were shown to correlate with the activity of liver amino transferases. The results obtained in this study indicate that the field of the use of PCR in the analysis of hepatitis A virus may include cases of mixed infection and virus carriership.

[Bacteriocins: criteria, classification, characteristics, methods of detection]. / Bakteriotsiny: kriterii, klassifikatsiia, svoistva, metody vyiavleniia.

The review on bacteriocins of Gram negative and Gram positive bacteria. Criteria making it possible to regard antagonistic substances as bateriocins or bacteriocin-like substances and on their classification are presented. Examples of bacteriocins naming depending on the taxonomic position of the producer culture are given. Information on the physico-chemical and biological properties of bacteriocins and their purification is presented as well as on detection tools of bacteriocins in microorganisms and evaluation of the producer activity of the bacteriological culture.

[Effectiveness of the oral administration of tomicide in experimental infection]. / Otsenka éffectivnosti énteral'nogo primeneniia "tomitsida" pri éksperimental'noi infektsii.

Experimental data on the oral administration of Tomicid using different schemes for the protection of white mice from staphylococcal infection are presented. The use of Tomicid, administered in the maximum dose admissible for mice, ensured the protection of up to 2/3 of the total number of mice. A single oral administration of the preparation immediately after infection protected 1/3 of the survived mice from local staphylococcal infection. Good prospects of using Tomicid for the prevention of catarrhal diseases in guinea pigs were established.

[Action of Spirulina platensis on bacterial viruses]. / Deistvie Spirulina platensis na virusy bakterii.

The impact of the biomass of the blue-green microalga (cyanobacterium) S. platensis on bacteriophage T4 (bacterial virus) has been evaluated. The study revealed that the addition of S. platensis biomass into the agar nutrient medium, followed by sterilization with 2% chloroform and thermal treatment, produced an inhibiting or stimulating effect on the reproduction of the bacteriophage in Escherichia coli B cells, depending on the concentration of S. platensis and the multiplicity of phage infection, as well as on the fact whether the microalgae were added during the first cycle of the development of the virus. The reproduction of the bacteriophage in E. coli B was influenced by the method and duration of the sterilization of the nutrient medium with S. platensis.

[Detection of biologically active substances with antagonistic and stimulating activity in Spirulina platensis]. / Vyiavlenie biologicheski aktivnykh veshchestv s antagonisticheskim stimuliruiushchim deistviem u Spirulina platensis.

The results of studies on the detection of biologically active substances (BAS) in biomass dilutions and culture fluid of Spirulina platensi and algae (Chlorella, Fucus, Laminaria) by the agar diffusion method are presented. After the sterilization of the solutions with chloroform (CF) a substance with lysozyme-like activity and 2 substances with antagonistic activity deep in agar and on its surface were detected with the use of the micrococcal indicator strain. After CF treatment, depending on the concentration of S. platensis strains, a compound stimulating the growth of bacteria and sensitive to heat treatment was detected. BAS were also detected with the use of other indicator cultures.

[Cultivation of Pseudomonas aeruginosa in media with chlorella fermentative hydrolysate as nutrient base]. / Kul'tivirovanie Pseudomonas aeruginosa na sredakh s ispol'zovaniem fermentativnogo gidrolizata khlorelly v kachestve pitatel'noi osnovy.

The specific features of the growth of P. aeruginosa in media prepared with the use of Chlorella fermentative hydrolysate as nutrient basis were studied. The accumulation of microbial biomass was shown to depend on increased content of amino nitrogen in the media, while the effectiveness of the pyocyanin formation was linked with decreased content of amino nitrogen and a rise in the amount of sodium chloride.

[Biological activity of Spirulina]. / Biologicheskaia aktivnost' spiruliny.

In this review information of Spirulina platensis (SP), a blue-green alga (photosynthesizing cyanobacterium) having diverse biological activity is presented. Due to high content of highly valuable proteins, indispensable amino acids, vitamins, beta-carotene and other pigments, mineral substances, indispensable fatty acids and polysaccharides, PS has been found suitable for use as bioactive additive. SP produces an immunostimulating effect by enhancing the resistance of humans, mammals, chickens and fish to infections, the capacity of influencing hemopoiesis, stimulating the production of antibodies and cytokines. Under the influence of SP macrophages, T and B cells are activated. SP sulfolipids have proved to be effective against HIV. Preparations obtained from SP biomass have also been found active against herpesvirus, cytomegalovirus, influenza virus, etc. SP extracts are capable in inhibiting cancerogenesis. SP preparations are regarded as functional products contributing to the preservation of the resident intestinal microflora, especially lactic acid bacilli and bifidobacteria, and to a decrease in the level of Candida albicans. The biological activity of SP with respect to microorganisms holds good promise for using these microalgae as components of culture media.

[Stimulating and inhibiting effect of Spirulina platensis on microorganisms]. / Stimuliruiushchee i ingibiruiushchee vozdeistvie Spirulina platensis na mikroorganizmy.

The blue-green microalga (cyanobacterium) S. platensis and the complex of its metabolites in the culture fluid being added into nutrient agar in doses of 0.01, 0.1 and 10 mg/ml, may produce a stimulating and inhibiting effect on microorganisms. In Spirulina biomass and in culture fluid substances which may be associated with the stimulation or inhibition of the growth of some microorganisms are supposedly contained. The manifestation of stimulating or inhibiting properties depended on the concentration of S. platensis and the complex of its metabolites, as well as on sterilization methods of nutrient media supplemented with these substances.