[Optimization of Streptococcus equi subsp. zooepidemicus cultivation process--producer of hyaluronic acid].

AIM: Selection of high-mucoid morphotype of Streptococcus equi subsp. zooepidemicus (Streptococcus zooepidemicus) and study of its morphological, physiological, biochemical and technological characteristics for providing increased secretion of hyaluronic acid (HA). MATERIALS AND METHODS: Submerged cultivation was performed in 100 ml glass flasks without baffles or in 1.5 or 10 1 laboratory bioreactors. LB and MRS media were used for cultivation. Mutagenesis was carried out by UV exposure with consequent selection of mucoid phenotype. HA was determined by carbazole method or after exhaustive acid hydrolysis by reaction of N-acetylglucosamine with Morgan-Elson reagent. Total hyaluronidase activity was evaluated by viscosimeter. Determination of cell and capsule size, ability to ferment carbohydrates and other microbiological, physiological and biochemical tests were performed by standard techniques. RESULTS: Instability of capsule phenotype of S. zooepidemicus B-8014 strain was revealed that is explained most probably by formation under certain conditions of bacterial hyaluronidase. This is confirmed by a reduction of HA concentration in cultural medium at pre- and stationary growth phases. Mucoid strain S. zooepidemicus KB-04 was obtained by mutagenesis with subsequent selection that is characterized by increased capsules. The strain was studied for HA formation. Optimization of growth medium composition, physical-chemical conditions and modes of cultivation allowed to significantly increase HA yield. CONCLUSION: The studies of morphologic, physiologic, biochemical and technological characteristics of the high-mucoid S. zooepidemicus KB-04 strain obtained by mutagenesis with consequent selection were performed, conditions of its cultivation and composition of growth mediu by carbon source and content of bivalent metal ions were optimized.

[Synthesis and degradation of hyaluronic acid by bacteria of Streptococcus genus].

Modern data on metabolism of hyaluronic acid by bacteria from Streptococcus genus are presented. Several species of bacteria forming capsule from hyaluronic acid, which is analogous to glycosaminoglycan of vertebrates, are considered. Different aspects of hyaluronic acid synthesis are described: biochemical synthesis pathway, genetic basis, regulation of expression of genes belonging to hyaluronic acid synthesis operon. Biological role and physiologic importance of hyaluronic acid for bacteria, including its role in overcoming immune barrier by pathogenic species, are discussed. Process of depolymerization of hyaluronic acid in presence of hyaluronatlyases secreted by certain streptococci is considered. Characteristic of streptococcal enzyme hyaluronatlyase, its mechanism of catalytic effect, and biological function are presented.

Functional activity of peritoneal macrophages from sensitive and resistant mouse strains during intravaginal infection with herpes simplex virus type 2 and mucosal vaccination.

In the early period after intravaginal infection with herpes simplex virus type 2 (2 h), macrophages from sensitive DBA/2 mice were characterized by higher capacity to engulf the antigen, decreased function of the lysosomal apparatus, lower activity of cathepsin D, and reduced oxygen metabolism compared to cells from resistant BALB/c mice. Mucosal vaccination with herpes vaccine and hyaluronic acid promoted the increase in functional activity of macrophages and improved survival of sensitive mice (by 60%).

[Mechanisms for the therapeutic effect of herpes polyvaccines in chronic ophthalmic herpes and genital herpes]. / Mekhanizm lechebnogo éffekta gerpeticheskoi polivaktsiny pri khronicheskom oftal'mogerpese i gerpes genitalii.

Commercial inactivated culture polyvaccine against herpes simplex viruses (types 1 and 2) developed at D. I. Ivanovsky Institute of Virology promoted cessation of viremia. During the first vaccination viremia coincided with appearance of a focal allergic test on the retina, which is proposed for the diagnosis of herpetic involvement of the posterior compartment of the eye. T-cellular immunity normalized after a course of vaccination. Experimental immunization of rats and vaccination of patients with chronic ophthalmic and genital herpes demonstrated the therapeutic activity of inactivated herpetic polyvaccine in suppositoria.

[Development of a suppository form of the interferon inducer poludan and potentiation of its action by hyaluronic acid]. / Razrobotka svechevoi formy induktora interferona poludan i potentsirovanie ego deistviia gialuronovoi kislotoi.

A promising dosage form of interferon inductor poludan has been developed for the treatment of patients with genital herpes and ophthalmic herpes: suppositoria. High interferon-producing and antiviral activity of poludan in suppositoria is potentiated by hyaluronic acid and antioxidants.

[Molecular mechanisms of autorepair of DNA double strand breaks]. / Molekuliarnye mekhanizmy samoreparatsii dvoinykh razryvov DNK.

The physical mechanism of interaction between nucleotides in intracellular liquid medium was studied. It was shown that during the contact between the thymine and guanine there exists a potential barrier the distance of about 10 A, which prevents the enzyme self-repairing in DNA after a double damage. All the remaining pairs of nucleotides have no such a barrier. Some processes of self-repairing of DNA for various types of double damages are analyzed. The geometry of an irreversible damage of DNA was determined. It was shown that with the increase in the number of hydrated electrons diminishing of the viscosity and increase in the temperature of intracellular medium the barrier vanishes and DNA undergoes a complete self-repairing.

[Mechanisms of the combined effect of SHF electromagnetic radiation and hydrogen peroxide on the viability of microorganisms]. / Issledovanie mekhanizmov kombinirovannogo deistviia élektromagnitnogo izlucheniia SVCh i perekisi vodoroda na zhiznesposobnost' mikroorganizmov.

The cellular (viability) and molecular (kinetics of changes in nucleoid viscosity) mechanisms of damage to various strains of microorganisms after single and combined action of SHF electromagnetic radiation and hydrogen peroxide are compared. The conditions are found in which marked synergetic interactions of the agents are realized. The role of repair systems in formation of final products of the interactions is discussed.

[Comparative analysis of mechanisms of the modification of microorganism viability under the effect of UHF heating and hyperthermia]. / Sravnitel'nyi analiz mekhanizmov modifikatsii zhiznesposobnosti mikroorganizmov pri vozdeistvii SVCh-nagreva i gipertermii.

In experiments with models of isogenic Escherichia coli strains, a comparative study was made of the effect of SHF of electromagnetic field and hyperthermia. The survival rate of bacteria was determined and, simultaneously, injuries to genetic supramolecular structures were registered through measuring the anomalous time dependence of cell lysate viscosity. The combined effects of the mixture of these factors with H2O2 microconcentrations were studied. The differences observed in their realization were attributed to the different mechanisms of action of these factors on the repair enzyme systems of the studied cells. It has been found that the effect of microwaves on microorganisms causes much severer damages to DNA that hyperthermia does.

[Physico-chemical and biological characteristics of Pseudomonas aeruginosa elastase]. / Nekotorye fiziko-khimicheskie i biologicheskie kharakteristiki élastazy Pseudomonas aeruginosa.

Elastase isolated from P. aeruginosa clinical strain hydrolyzes elastin, casein, hemoglobin, ovalbumin, gelatin, fibrin, collagen. The optimum pH ensuring the activity of the enzyme is 7.8-8.0. Elastase shows maximum stability at pH 6.6-9.0. Heating at 80 degrees C for 10 minutes results in its practically complete inactivation. Elastase is a highly radiosensitive enzyme. Chelating agents and zinc, cobalt, mercury ions suppress its activity. Sodium and ammonium chlorides selectively inhibit the elastolytic, but not proteolytic activity of the enzyme. Elastase shows pronounced dermonecrotic and keratolytic action.

[Effect of gamma radiation on the components of hepatitis B virus and the delta antigen]. / Vliianie gamma-izlucheniia na komponenty virusa gepatitia B i del'ta-antigen.

The influence of ionizing irradiation on the serological activity of purified HBsAg, receptors to polymerized albumin, DNA-polymerase activity, delta agent, and delta antigen was studied. Different radiosensitivity of hepatitis B virus components and delta-system was demonstrated. The possibility of sterilization by irradiation of purified HBsAg preparations suitable for construction of a vaccine against hepatitis B was shown.

[Mechanisms of the synergistic action of ionizing radiation and hydrogen peroxide on the viability of bacteria]. / Mekhanizmy sinergicheskogo deistviia ioniziruiushchego izlucheniia i perekisi vodoroda na zhizhnesposobnost' bakterii.

The combined effect of ionizing radiation and minor concentrations of hydrogen peroxide (0.1-0.01 M) has been studied on bacteria differing in the cell wall structure (Gy+, Gy-), radioresistance, and activity of DNA repair system. The H2O2 concentrations applied, which have a slight antibacterial action, enhance significantly the bactericidal effect of ionizing radiation producing a synergistic effect due to the decreased effectiveness of repair of DNA single-strand breaks.

[Mechanisms of bacterial inactivation as affected by cation surface-active substances]. / Issledovanie mekhanizmov inaktivatsii bakterii pri vozdeistvii kationnogo poverkhnostno-aktivnogo veshchestva.

The death kinetics and ultratructure of gram-positive and gram-negative bacteria exposed to alkyldimethylammonium chloride were studied. It was shown that at low concentrations (0.0001 per cent) the agent had a pronounced bactericidal effect, which was higher with respect to the gram-positive bacteria. Impairment of the intactness of the cytoplasmic membrane, a vitally important organoid of the bacterial cell, was the main factor in the mechanism of action of the cationic surface-active substance. Membrane structures of various configuration and localization not associated with the cell division were detected in the cytoplasm of the dead bacteria. These structures are probably the result of self-assembly of the membrane lipid components broken under the action of the detergent.

[Quantitative assessments of synergism]. / Kolichestvennye otsenki sinergizma.

The basic approaches to the assessment of the biological effects of combinations of different agents are briefly discussed. The situations of additivity, synergism and antagonism are defined with hetero- and isoaddition being taken into consideration. The practical application of the methods is illustrated by the analysis of the experimental data on the staphylococcus survival after the combined effect of gamma-irradiation and hydrogen peroxide or incubation at elevated temperature.

[Effect of gamma radiation on the immunobiological and immunochemical properties of cholera exotoxin. IV. The biological and immunochemical properties of purified irradiated choleragen]. / Vliianie gamma-izlucheniia na immunobiologicheskie i immunokhimi-cheskie svoistva kholernogo ékzotoksina. Soobshchenie IV. Biologicheskie i immunokhimicheskie svoistva ochishchennogo obluchennogo kholerogena.

The results of investigations carried out to study the effect of gamma radiation on the properties of the purified preparations of cholera exotoxin are presented. Irradiation has been shown to decrease the anterotoxicity of purified choleragen and the activity of its permeability factor, depending on the radiation dose. The investigations have revealed that in purified toxin enterotoxicity is completely inactivated with a lover radiation dose than in crude toxin filtrate (25 kGy). In immunochemical reactions the increase of the electrophoretic mobility of the choleragen components, correlated with the increase of the radiation dose, and the reduced number of protein zones have been observed. The irradiated preparations of purified choleragen have been found to retain their immunogenic properties and serological activity.

[Effect of gamma radiation on the immunobiological and immunochemical properties of cholera exotoxin. III. The serological activity and immunochemical properties of irradiated unpurified toxin]. / Vliianie gamma-izlucheniia na immunobiologicheskie i immunokhimicheskie svoistva kholernogo ékzotoksina. Soobshchenie III. Serologicheskaia aktivnost' i immunokhimicheskie svoistva obluchennogo neochischennogo toksina.

The immunochemical properties and serological activity of irradiated preparations of crude cholera exotoxin have been studied. This study has revealed that with the increase of the dose of ionizing radiation changes occur in the physico-chemical properties of the preparations of the toxin, which leads to an increase in the electrophoretic motility of the protein components of the toxin, to the aggregation and polymerization of individual fragments. The preparations of antigen exotoxins have been shown to retain their serological activity within the range of radiation doses under study (10-350 kGy).

[Effect of gamma radiation on the immunobiological and immunochemical properties of cholera exotoxin. I. Change in the biological activity of nonpurified cholera exotoxin as affected by ionizing radiation]. / Vliianie gamma-izlucheniia na immunobiologicheskie i immunokhimicheskie svoistva kholernogo ékzotoksina. Soobshchenie I. Izmenenie biologicheskoi aktivnosti neochishchennogo kholernogo ékzotoksina pod vozdeistviem ioniziruiushchei radiatsii.

Crude cholera exotoxin (filtrate toxin) was irradiated with increasing doses of gamma radiation. A significant drop in enterotoxicity, in the activity of the permeation factor and a decrease in toxicity were shown to occur as radiation doses increased. Radiation doses of 50-70 kGy were found to completely inactivate enterotoxicity in liquid toxic preparations. A higher radioresistance of dried preparations in comparison with liquid ones was registered: inactivation occurred at 150-200 kGy. Different batches of the initial filtrate toxin had varying radiosensitivity. The sterilizing effect of gamma radiation was achieved at doses of 20 kGy for liquid preparations and 30 kGy for dried preparations. During the prolonged storage of the irradiated preparations of crude toxin (the term of observation being 1.5 years) at different temperatures no reversion of toxicity was found to occur, while their immunogenic properties remained unchanged.