[The microbiocenosis of the palatal tonsils in the practically healthy people]. / Mikrobiotsenoz nebnykh mindalin u prakticheski zdorovykh lits.

The objective of the present study was the investigation of the quantitative and qualitative composition of the cultured microorganisms isolated from the proximal parts of the palatal tonsil lacunes of the practically healthy people. The data obtained made it possible to characterize the microbiocenosis of the palatal tonsil lacunes in the practically healthy subjects at the age from 18 to 30 years. A total of 153 strains were identified with the use of mass spectrometry; they represent six genera of two phylotypes that occur in the form of associations comprised of 4-5 microorganisms with the predominance of the species of the genus Streptococcus. The results of the study expand our knowledge about the oropharyngeal microbiota and create the prerequisites for the better uundderstanding of the contribution of the microbiotic communities to the development of pathology of the upper respiratory tracts.

Phagebiotics in treatment and prophylaxis of healthcare-associated infections.

We have developed a phagebiotic composition using 8 virulent bacteriophages (2 strains of each species) which are able to lyse Acinetobacter baumannii, Klebsiella pneumoniae, Pseudomonas aeruginosa and Staphylococcus aureus. The unique character of the developed composition is ensured by particular properties of each bacteriophage comprising the preparation, including their range of lytic activity toward specific bacterial pathogens, morphology of their plaques, cycle of their development, restriction profile of their DNAs, specificity of their genomes (based on complete genome sequencing), and other properties. The preparation did not produce any signs of acute or chronic intoxication in the experimental animals. Therapeutic and prophylactic efficiency of the phagebiotic composition was demonstrated in the prevention and treatment of the experimental acute K. pneumoniae infection in mice. The investigations have shown that the preparation possesses a high therapeutic efficiency and is highly competitive with ciprofloxacin which is very effective against the infective strain K. pneumoniae. Our small-scale clinical trial was aimed to evaluate therapeutic effectiveness of the phagebiotic composition in an epidemiological emergency situation in an intensive care unit, caused by multi-resistant strains of Acinetobacter baumannii, Klebsiella pneumoniae and Pseudomonas aeruginosa. Seventy nine per cent of the initial samples from 14 patients' endotracheal aspirate, blood and urine were contaminated. Twenty-four hours after the 3-day phage therapy (20 ml of cocktail at a titer for each phage 108 pfu/ml were introduced intragastrically through a tube once a day) contamination level dropped to 21%. Hence the obtained results enabled us to create a new phagebiotic composition that may be used as an alternative to antibiotics to treat these healthcare-associated infections.

A small-scale experiment of using phage-based probiotic dietary supplement for prevention of E. coli traveler's diarrhea.

Traveler's diarrhea (TD) is caused by Escherichia coli in 30% of cases. We have developed a phage cocktail for prophylaxis of TD caused by E.coli, Shigella flexneri, Shigella sonnei, Salmonella enterica, Listeria monocytogenes or Staphylococcus aureus, and investigated its effectiveness against infection caused by the non-pathogenic Lac (-) strain of E.coli K12 C600 in animal and human trials. On the 6th day of both animal and human trials E. coli K12 C600 strain was detected in titer of 104 CFU/g of mice feces and 106 CFU/g of human feces in the control (untreated) groups, while it was not detected in the samples of either of the study (phage-treated) groups. These results have great significance because the original coliphages included in the cocktail have a broad host-range including ETEC, EAEC and EHEC strains which cause severe cases of TD.

[Comparative analysis of informative-diagnostic properties of mucosa immune-reactivity parameters].

AIM: Study the features of immune-reactivity expression in mucosa depending on their topicity and etiopathogenesis of the pathological process. MATERIALS AND METHODS: Data from 30 clinically healthy children and 77 children with acute and recurrent diseases of respiratory tract: 51--with acute and 15--with chronic bronchitis; as well as 132 women: 41--with active stage of acute urogenital chlamydia infection, 29--with recurrent chronic process, 30--with non-recurrent form and 32 clinically healthy women were analyzed. Saline and urogenital tract mucosa discharge was analyzed for IgG, sIgA and secretory component, IL-1beta, 4, 6, 8, 9, 10, 12, IFNgamma, TNFalpha and GM-CSF, TLR-2, TLR-3, TLR-4, TLR-8 gene expression levels as well as content of lysozyme, total protein and leucocytes. RESULTS: Solidity, universality and practically single-stage triggering of mucosa immune reaction mechanisms to intervention by foreign agents regardless of their localization was confirmed. A dependence of immune-reactivity expression on the form of pathologic process, its localization and qualitative and quantitative characteristics of the infectious agents was clearly seen. The highest level of clinical-laboratory and immunological parameters is inherent for patients with acute processes in urogenital tract (cervical canal and urethra), especially cause by mixed infections. CONCLUSION: Immune diagnostic parameters of mucosa among which TLR system is especially notable have high information properties allowing not only diagnostics of inflammatory process but also differentiating its form and character our course.

[Molecular-genetic and phylogenetic characteristic of Streptococcus pneumoniae strains isolated from patients with meningitis and carriers].

AIM: Analyze genetic and phylogenetic interrelations between S. pneumoniae strains isolated from meningitis patients and carriers. MATERIALS AND METHODS: 23 S. pneumoniae isolates (9 from bacterial meningitis patients, 9 from nasopharynx of bacterial carriers, 5 strains from museum collection of Gabrichevsky Moscow Research Institute of Epidemiology and Microbiology) were studied. S. pneumoniae ATCC 49619, S. mitis ATCC 49456 served as control strains. S. pneumoniae serotype determination was carried out in latex-agglutination reaction and quelling reaction. Multiplex PCR according to WHO protocols was used for molecular-genetic study and pneumolysin (ply), autolysin (lytA), surface cellular adhesin A (psaA) and capsule polysaccharide (cpsA) gene determination. Multilocus sequence-typing was carried out according to WHO scheme for 7 "housekeeping" segments--aroE, gdh, gki, recP, spi, xpt and ddl. Computer programs from available internet resources were used for data processing and dendrogram building. RESULTS; The S. pneumoniae isolates analyzed were established to belong to 19 sequence types that may be combined into 4 subclusters. Results of molecular-genetic and serologic typing were completely comparable. CONCLUSION: Attribution of isolates from the same serotype and serogroup to different sequence-types gives evidence on the ongoing changes within serotype and as a result changes in allele profile of circulating S. pneumoniae isolates. Membership of isolates in separate subclusters gives evidence on close evolution relationship between isolates obtained from patients with bacterial meningitis and carriers. Certain isolates had not previously been registered in Russia and were probably imported from the territories of other countries.

[Development of polycomponent metabolite probiotic].

AIM: Construct composition of polycomponent probiotic based on lactobacilli cultural fluid (CF) supernatant. MATERIALS AND METHODS: Species membership of lactobacilli strains was specified by analysis of nucleotide sequences of 16S RNA genes. For comparison of probiotic effect of CF, supernatant and ultrafiltrate of a consortium of lactobacilli strains, studies of specter of peptides, amino- and other organic acids contained in them as well as profile of bactericidal and fungicidal activity was carried out. RESULTS; Technology of production of amicrobial probiotic agent based on CF of the consortium of lactobacilli strains was developed. During determination of concentration of metabolites of protein nature analogy of parameters was shown in CF supernatant and native lactobacilli cultures. In both the studied supernatants and CF the same set of organic acids was also reported to be present. During comparison of antimicrobial and anticandidosis activity of lactobacilli native culture and CF supernatant against Staphylococcus aureus 209, Enterococcus faecalis 1154 and Candida albicans 5 test-strains, an equivalent effect of both samples was shown. CONCLUSION: Polycomponent metabolite probiotic based on CF supernatant of Lactobacillus helveticus NKJC, Lactobacillus helveticus JCH and Lactobacillus casei KAA strains completely retains bactericidal and fungicidal activity of the native culture of the consortium of strains and at the same time lacks disadvantages of compositions containing live microorganisms.

[Comparative analysis of incidence and prevalence of cardiovascular diseases among military personnel and seafarers serving on the bank].

The study presents retrospective analysis of navy soldiers' cardiovascular diseases, which most often cause the growth in labor force, loss rates and navy soldiers' dismissal and mortality rates, and, therefore, are one of the most pressing problems of the military medical services. Comparative assessment conducted between emergence and pervasiveness of these diseases in navy with those of land-based soldiers. Analysis of correlation between morbidity rates and length of service in navy is also included.

[The phylogenetic position of Chlamydia strains isolated from monkeys and humans with Chlamydial pathology in the family Chiamydiaceae. Genotypic and phenotypic properties of this pathogen].

Based on the results of the comparative analysis concerning relatedness and evolutional difference of the 16S - 23S nucleotide sequences of the middle ribosomal cluster and 23S rRNA I domain, and based on identification of phylogenetic position for Chlamydophila pneumoniae and Chlamydia trichomatis strains released from monkeys, relatedness of the above stated isolates with similar strains released from humans and with strains having nucleotide sequences presented in the GenBank electronic database has been detected for the first time ever. Position of these isolates in the Chlamydiaceae family phylogenetic tree has been identified. The evolutional position of the investigated original Chlamydia and Chlamydophila strains close to analogous strains from the GenBank electronic database has been demonstrated. Differences in the 16S - 23S nucleotide sequence of the middle ribosomal cluster and 23S rRNA I domain of plasmid and non-plasmid Chlamydia trachomatis strains released from humans and monkeys relative to different genotype groups (group B- B, Ba, D, Da, E, L1, L2, L2a; intermediate group - F, G, Ga) have been revealed for the first time ever. Abnormality in incA chromosomal gene expression resulting in Chlamydia life and development cycle disorder and decrease of Chlamydia virulence can be related to probable changes in the nucleotide sequence of the gene under consideration.

[Colonization resistance and immunological reactivity of children's oropharyngeal mucosa in health and bronchopulmonary pathology].

The study group was comprised of 27 practically healthy children, 51 patients with acute bronchitis, 15 with chronic bronchitis and 11 with pneumonia. It was shown that changes of microbiocoenosis in back of the throat (BOT) were related to increased mucosal contamination with normal microflora and opportunistic microorganisms. The highest degree of contamination was observed in children with acute bronchitis. Normocoenosis was detected only in 13 practically healthy children. The disorders of microbiocoenosis took the form of disbiosis and acute inflammatory processes in patients with acute and chronic bronchitis and pneumonia. However, the large amount of normal flora together with the high Ig level ensured marked colonization resistance as evidenced by the values of natural colonization coefficient of nasopharyngeal epithelium (NCCNE) and balance coefficient (BC). These data suggested development of compensated secondary immunodeficiencies. In patients with acute bronchitis and pneumonia, local synthesis of Ig prevailed. It is shown that BC can be used to screen children for disorders of mucosal immunity. The presence of increased saliva IgE levels in patients with acute and chronic bronchitis supports the generally accepted concept of bronchi as a "shock organ" in allergic condition. It was demonstrated that IgE levels in saliva increase earlier than in serum and may be used as a prognostic criterion in patients with bronchopulmonary pathology.

[The role of lectins from probiotic microorganisms in sustaining the macroorganism].

The notion of lectins has been expanded with special reference to lectins of probiotic microorganisms (LPM). New data on LPM and their properties are considered in the context of a new concept of systemic organization of LPM being developed. Conditions for complete separation of poly (N-acetylgalactosamine-?)-binding lectins and polyMan-?/oligomannoside/mannan-binding lectins are developed on the assumption that they contain a single carbohydrate-binding site in their structure. It is shown that strain typing of probiotic microorganisms is possible based not only on protein but also on LPM composition. The observed peculiarities of LPM functioning substantially extend the concept of informational superiority of integral interatomic networks over their isolated components. This approach makes it possible to set functioning of LPM systems against that of lectin systems of other components of human microbiota as prevailing in the support of the normal status of the macrorganism. The results demonsatrate the unique character of LPM systems and suggest good prospects for their application in medicine, biotechnology and nanotechnology.

[Molecular mechanisms of induction of innate immunity].

Cellular and molecular mechanisms of congenital immunity at different levels are discussed including single cell expression patterns and intracellular localization of individual TLR, the use of adapter molecules for generation of activation signals in response to microbial and non-microbial pathogens, soluble trap receptors, and intracellular negative regulators.

[Classification of lectins as universal regulator molecules of biological systems].

Modern concepts of lectin classification reflecting tendencies in the development of functional classification of lectins (Ln) and Ln-like proteins are presented. Classification of bacterial Ln is proposed. Classification of Ln and Ln-like proteins of plants, animals, and bacteria is based on 8 major elements of protein secondary structure that allows for the evaluation of topographic diversity of their surfaces, potential compatibility of bacterial, vegetable, and mammalian Ln molecules and possible variants of their co-functioning.

[Opportunistic pathogenic microflora in intestine microbiocenosis of medical staff of multisectoral hospitals].

Microflora of colon in 81 medical staff was researched. Microecological disturbances mostly were represented by dysbacteriosis of second degree. The composition of intestinal microorganisms correlated with duration of the work in hospital. The study revealed decrease in medical staff with five years old work in hospital the detection rate of indigenous microflora and colonization of fecal microbiocenosis with Klebsiella, lactosonegative Escherichia coli, Staphylococcus aureus and Candida albicans. The biotype of medical staff working 6-10 years in hospital had mostly haemolytic microorganisms: E. coli, Staphylococci and Enterococci. Changes in intestinal microbiocenosis of medical staff are periodically repeated.

[Nanotechnologies and perspectives of their application in medicine and biotechnology].

The results of nanotechnologies application in biological and medical fields are summarized. The examples of nanotechnologies use in reactions of cell adhesion, cell recognizing, intracellular sorting, organ and tissue construction, nanobiosensing, drug delivering and cancer therapy are presented. The prospects of nanotechnologies use in biomedicine, biothechnology and cell biology are shown.

[General properties and functioning principles of lectins in biological systems].

The authors give the literature review with analysis of main conceptions of lectins - their structure and functions according to state-of-the-art science and production requirements. Up-to-date definition of lectins based on last achievements of science and practice represented in the article.

[The microflora of vaginal, gutter, and intestinal bioptats of women with a threat of early miscarriage].

A complex simultaneous assessment of the microbiota (the aerobic and anaerobic links as well as the parietal and lumen components) of the gutter, vagina, and intestines of women with a pathological pregnancy was performed. In 30% of women the study revealed system dysbiotic changes. Local immune reactivity was decreased, which may be considered a provoking factor. In conclusion, complex microbiological examination makes it possible to objectivize the picture of the pathological process and its outcome. A criterion for the administration of correcting measures is offered.

[Bifidobacterial plasmids and their application to genetic engineering].

Representatives of Bifidobacterium genus are considered to play many important roles in intestinal homeostasis. On the other hand, their molecular biology and genetics have been poorly studied. In order to broaden our understanding of their health-promoting mechanisms, it is extremely important to possess tools to manipulate them genetically. Another challenging task is to take advantage of genetic engineering technology for designing new probiotic bifidobacteria with unique therapeutic properties. An important step in such work is to isolate and characterize small bifidobacterial plasmids, which can be applied to the construction of cloning vectors. This article presents a review of several pioneering studied devoted to bifidobacterial plasmids and genetic engineering with bifidobacteria. Trends in and prospects of molecular genetics of bifidobacteria are discussed as well.

[Strategic aspects of constructing probiotics of the future].

The publication systematizes modern achievements in understanding the role of microbiocenoses in the existence of macroorganism, and possible ways of constructing of probiotics of the future.

[The role and biological significance of toll-like receptors in the anti-infectious resistance of the organism].

Modem conceptions of the role of toll-like receptors (TLR) in the innante immunity mechanisms realization and data on the interaction between TLR and pattern-associated molecular proteins of microbial or endogenic origin are presented in the review.

[Level of natural autoantibodies to interferon-alpha and characteristics of interferon status during chronic hepatitis and liver cirrhosis].

Level of autoantibodies to interferon-alpha and characteristics of interferon status were studied in 118 patients with chronic diffuse liver diseases (CDLD). During formation of liver cirrhosis in patients with CDLD level of natural autoantibodies to endogenous interferon-alpha increased. During progression of CDLD, decrease of interferon-alpha level and its functional activity were observed as well as change in the ratio between active fraction of interferon-alpha and its inactivated forms towards increase of the latter. One cause of observed decrease in biological functional activity of interferon-alpha in patients with chronic hepatitis and liver cirrhosis was increased titer of interferon-alpha neutralizing autoantibodies in blood. Level of natural autoantibodies to interferon-alpha, its antiviral activity and concentration during chronic hepatitis and liver cirrhosis can be used as markers of disease severity and predictors of its outcome.