Vol 92, No 4 (2015)

In the article new line of research - infectious symbiology that studies interaction of microsymbionts with the host in case of infection under the conditions of their biocommunicative interrelations is discussed. The presence of interrelation of intestinal microbiota with human homeostasis disorders postulates thesis that: human organism is superorganism including a lot of prokaryote species where microbiota normoflora representatives are the main complex of extracorporeal physiological system acting as organism homeostasis regulator. When studying intermicrobial relations under microsymbiocenosis its system-forming factor has been identified that includes basic general functions of microorganisms - reproductive (growth/reproduction) and adaptive (biofilm formation and antilysozyme test). It has been shown that microbial interaction in dominant-associant vector providing microsymbiocenosis functioning is implemented via opposite (amplification/suppression) effect of reproductive and adaptive functions ofmicrosymbionts of dominant-associant pair contributing self-non-self discrimination. Intermicrobial discrimination of self-non-self microsymbionts goes before signaling of adaptive immunity of the host promoting microbe elimination or persistence in the body depending on the result of identification. It has been noted that microbial factor fulfilling the role oforganism homeostasis regulator performs synergistic functioning of dominant microflora with the host. When penetrating foreign associants (pathogens) into the organism host defense is implemented by means of induction of its adaptive immunity via cytokine reticulum. Key players of human intestinal microbiota are Bifidobacteria which along with induction of adaptive immunity and balance changes of pro- and anti-inflammatory cytokines of the host directly regulate microsymbiocenosis including system of hormones and neurotransmitters.

Aim. Study the prevalence and intensity of anti-hemoglobin and anti-lactoferrin activity in enterococci as representatives of symbiotic intestine microflora and causative agents of infection-inflammatory diseases in children. Materials and methods. 95 strains of enterococci were used in the study, that were isolated from intestines of children during examination for dysbiosis, and 34 clinical isolates. Strain identification was carried out by using multiplex PCR. Anti-lactoferrin and anti-hemoglobin activity were determined by O.V.Bukharin et al. (2005). Results. A higher level of prevalence and intensity of anti-lactoferrin, anti-hemoglobin activity, as well as a combination of those properties was established in clinical isolates of Enterococcus faecalis and E. faecium compared with strains, isolated from intestines of healthy individuals. Conclusion. The studied properties could be used for differentiation of pathogenic and non-pathogenic strains of enterococci and during selection of inhibitory preparations, suitable for therapy of enterococci infections.

Aim. Study the effect of exometabolites of associative microbiota on circadian dynamics of functional parameters, that reflect pathogenic and persistence properties of fungi and bacteria. Materials and methods. Clinical isolates of Candida albicans, isolated from intestine of healthy individuals and patients with candidosis, as well as clinical isolates and museum ATCC strains Staphylococcus aureus 25923, Escherichia coli 35218 and Pseudomonas aeruginosa 27853 were taken for study of proliferative, adhesive, catalase, protease, phospholipase, hemolytic, anti-lysozyme, biofilm-forming activity. The results were treated statistically. Results. C. albicans isolates, isolated from healthy individuals were revealed to be indifferent to the effect of bacterial metabolites. Chrono-infrastructure of biological properties of fungi altered under the effect of microbiota metabolites. Hospital isolates of S. aureus, E. coli and P. aeruginosa displayed a relative stability of physiological properties against the effect of bacterial-fungal metabolites as opposed to museum strains. Conclusion. The alterations of chrono-infrastructure of biological rhythms of microorganisms by bacterial-fungal metabolites of associants reflect the intensity of the biological system, that is inevitable during the process of formation of inter-microbial interactions.

Aim. Comparative phenotypical and genetical evaluation ofpathogenic potential of Staphylococcus aureus strains, isolated from resident bacterial carriers, residing on the territories with anthropogenic pollution of air environment of varying intensity. Materials and methods. S. aureus, isolated 3 times from mucous membrane of the anterior of nose from 210 children, were the object of the study. Anti-carnosine activity and biofilm formation was determined by a photometric method, antibiotics resistance - by a disc diffusion method. lukS, lukF, sec 3, clfA, clfB, agr and mecA gene detection, that are associated with S. aureus, was carried out by PCR. Results. S. aureus strains, isolated from children, residing on the territories with a high level of anthropogenic pollution of air environment, were characterized by antibiotics resistance, higher values of anti-carnosine activity, 2 times more frequently formed biofilms with higher values of the parameter. clfA and clfB genes, that determine colonization of mucous membranes, and agr gene were detected in all the studied S. aureus strains, lukF and sec 3 genes were detected in 20 - 40% of the strains, isolated from children, residing on both territories. mecA and lukS genetical determinants were not detected. Conclusion. S. aureus, isolated from children, residing on the territories with high levels of anthropogenic pollution of air environment, were characterized by higher values of the studied factors of persistence and stability against antibiotics. Genetical determinants of pathogenicity were not detected in S. aureus, isolated from individuals, residing on both territories.

Aim. Determination of modulating effect of acid-base state of medium for cultivation of enterobacteria on activity of C-type lysozyme. Materials and methods. Escherichia coli BL21(DE3) strain for protein expression, Escherichia coli K12 MG1655 model strain, Escherichia coli No. 242 strain, isolated from intestine biotope; 2 Klebsiella pneumoniae strains, one of those contained plasmid homologue of periplasmatic lysozyme inhibitor gene pliC; 1 typical Salmonella enterica ATCC 14028 strain and a Micrococcus luteus ATCC 15307 strain as a control - served as material for the study. The bacteria were cultivated for 24 hours in 2 ml of liquid medium LB at 37°C, 250 rpm. Determination of antilysozyme activity (ALA) was carried out by a photonepehlometri-cal method according to O.V Bukharin et al. (1999) with alterations. Results. All the studied microorganisms, including Micrococcus luteus, at the specified conditions 24 hours after cultivation were established to change the pH of the liquid nutrient medium LB from the initial value of 6.6±0.1 to 8.2±0.2 units. ALA determination in the cultivation medium without buffer correction was accompanied by a decline of lysozyme activity at an order of magnitude. The effect was absent during ALA measurement by a standard technique. Conclusion. The local shift of acid-base state of biotope under the conditions of buffer system insufficiency results in a reversible alteration of antimicrobial activity of muramidase, that among other non-specific factors of the environment determines the background of interactions on the level of associative symbiosis. This aspect should be taken into consideration during development of models, that are close to real conditions of microsymbiocenotical interactions.

A review of studies on pneumococcal biofilms as a form of persistence is presented. The following provisions are examined: formation of pneumococcal biofilm on abiotic and mucosal surfaces, pathogenetic significance of biofilm pneumococci, their immunogenicity, as well as resistance to antibiotics and unfavorable environmental factors. Differences between biofilm properties, that are formed in vivo and in vitro, are shown. The significance of pneumococcal biofilm formation as means of survival for a long time is underscored.

Aim. Evaluation of microbiological effectiveness of longidaze preparation use during therapy of active nonspecific endometritis in women with uterus myoma. Materials and methods. 2 groups of women were formed by a method of random selection. The first was composed of patients with active chronic endometritis, that had received standard etiotropic antibacterial therapy. The second included patients, that had received longidaze in addition to the standard therapy. Therapy effectiveness evaluation was carried out based on the analysis of microbial landscape of cervical canal and uterine cavity 2 months after the therapy during phase I of the menstrual cycle. Results. Nonplasmacoagulating staphylococci, enterococci and anaerobes were established to be present predominately in the examined women of both the first and the second group. Conclusion. The application of longidaze in combination with antibacterial therapy results in the most significant improvement of microbial landscape of cervical canal and uterine cavity.

Aim. Study the effect of RioFlora Immuno Neo and Bifiform probiotics on the colon microflora composition under the conditions of experimental dysbiosis in mice. Materials and methods. Medicinal dysbiosis was modelled in animals by gentamicin administration, probiotics were administered intragastrally by the end of dysbiosis formation. Quantitative and qualitative study of mucous microflora of mice colon was carried out by a bacteriological method. Results. Correction of dysbiosis by probiotics has resulted in correction of the intestine microflora composition, and more pronounced when RioFlora Immuno Neo was used. Conclusion. The results of the study allow to recommend the use of these preparations for selective correction of colon dysbiosis.

Aim. Evaluate immune regulatory properties ofbifidobacteria metabolites during eubiosis and dysbiosis of the human colon. Materials and methods. Anti-cytokine activity of metabolites of bifidobacteria clinical strains and their ability to influence the production of pro- and anti-inflammatory cytokines (IL-10) by peripheral blood mononuclear cells of healthy humans was studied, taking into account microecological state of the human intestine. Determination of final concentration of cytokines in experimental and control samples was carried out by EIA. Results. Sensitive parameters, that are suitable for evaluation of stability of human intestine microsymbiocenosis, were detected. The level of microbial seeding, concentration of TNF-а and anti-lysozyme activity turned out to be informative for bifidobacteria in eubiosis conditions. The ability of bifidoflora metabolites to influence the production of pro-inflammatory cytokines (INF-y, TNF-а, IL-8) by human mononuclears was a significant parameter during formation of 1 - 3 degree dysbiosis. Conclusion. The maintenance of physiological state of intestine homeostasis is determined by immune regulatory properties of bifidobacteria metabolites, that is realized via their interaction with both cytokines (anti-cytokine activity) and production of cytokines by host immune cells (peripheral blood mononuclears).

Aim. Study the frequency of detection of mycoplasma and ureaplasma in clinical material from urolithiasis patients. Materials and methods. Clinical material samples (blood sera, urine, uroliths) from 31 urolithiasis patients were obtained during operations of urolith removal. Cultural method, LAR and PCR were used in the study. Results. The study of clinical material from 31 patients by PCR has shown, that in 25 individuals (80.6%) DNA of mycoplasma and ureaplasma was detected, and mycoplasma DNA was more frequently detected in uroliths and less - in blood sera. Mycoplasma hominis DNA was detected in clinical material of a significantly larger number of patients. 23 cultures were isolated from 8 patients by a cultural method, that were identified by PCR as M. hominis. All the isolates have grown as «mini-colonies». Even after multiple passages in agar medium, reversion of «mini-colonies» into colonies with a classic morphology was not obtained. Conclusion. A high frequency of detection of mycoplasma and ureaplasma in clinical material of patients with urolithiasis was established. The isolated M. hominis cultures have only grown as «mini-colonies». The phenomenon discovered could give evidence on high variability of mycoplasma and a possibility of existence of previously unknown form of their persistence in human organism.

Aim. Selection of optimal dosage regimen, length of treatment course (frequency of administration), safety, tolerance and clinical effectiveness evaluation of the medical preparation fortepren in patients with chronical recurrent herpes virus infection of genital localization. Materials and methods. The medical product of antiviral and immune modulating effect - fortepren (sodium polyprenyl phosphate) as a 4 mg/ml solution for injections combined with the base course of acyclic nucleoside acyclovir, 400 mg tablets, held studies. 40 male and female patients participated in the study. After a 10-day acyclovir course (400 mg x 3 times a day) for removing the acute phase, 4 groups of 10 individuals were formed: 1 - 5 ml (20 mg) of fortepren i/m once at day 13+2 after the start of the study after the completion of the treatment of the acute phase of the disease; 2 - 5 ml (20 mg) fortepren i/m 3 times at an interval of 21 days; 3 - 2 ml (8 mg) fortepren i/m 3 times at an interval of 21 days; 4 (control) - 5 ml of placebo i/m at remission stage 3 times at an interval of 21 days. Increase of the duration of inter-recurrence period, decrease of the severity of the recurrences, state of skin and mucous damage elements, improvements of immunologic parameters were considered during effectiveness evaluation. Results. Significant differences in the frequency of recurrences of genital herpes were shown for 3 months of observation in experimental and control groups. A significant reduction of genital herpes recurrence frequency from 3.52+0.09 (before treatment) to 2.89+0.08 (after treatment) was noted in patients of group 3 (p<0,001). The frequency of recurrences in the control group was 3.84+0.10, that was higher than the parameters in all the experimental groups. A significant reduction of the rash area was noted in group 3, moreover, a reduction of frequency of detection of clinical manifestations of genital herpes in the form of vesicle elements after treatment in groups 2 (p=0.02) and 3 (p=0.005) was found. Evaluation of local symptoms has established that burning have caused minimal discomfort for patients of groups 3 and 4 and itch and soreness - of groups 1 and 3. The least pronounced exacerbations were noted in patients of group 3. Intramuscular administration of fortepren preparation was established to result in the increase of titers of leukocyte virus-induced interferon for the whole duration of treatment. Conclusion. An intramuscular dose of 2 ml (8 mg) at recurrence stage 3 times at an interval of 21 days after the completion of the 10-day base course of treatment of the acute phase of chronical recurrent herpes virus infection of genital localization using acyclovir was accepted as an optimal dosage regimen. Analysis of the obtained results has shown an acceptable safety profile and a good level of tolerance for fortepren preparation.