Colonization by Staphylococcus aureus of Nano-Structured Fluorinated Surfaces, Formed by Different Methods of Ion-Plasma Technology.

Colonization of fluorinated surfaces produced by ion-plasma technology by Staphylococcus aureus was studied by scanning electron microscopy and surface energy analysis. It was shown that the intensity of colonization was determined by the surface relief and fluorine content. Formation of nanostructured surfaces accompanied by a sharp decrease in the surface energy prevented adhesion of Staphylococcus aureus cells to the fluorine-containing surface.

[ABILITY OF STAPHYLOCOCCUS OF VARIOUS STRAINS TO CREATE BIOFILMS AND THEIR EFFECT ON HUMAN BODY CELLS].

The urgency of the staphylococcus research is due to its ability to cause severe infections: softtissue infections, endocarditis, sepsis, toxic shock syndrome, and food poisoning. Coagulase-positive Staphylococcus aureus is the main infection agent of intrahospital infections. This agent has many factors of pathogenicity, which are well known. Among the coagulase-negative staphylococcus (CNS) strains, S. haemolyticus and S. epidermidis are clinically important, because they cause infections in patients with weak immune system. The mechanisms of the CNS pathogenicity are insufficiently understood. The goal of this work was to evaluate the potential pathogenicity of clinical strains of CNS from their capacity to create biofilms and the character of their interaction with human body cells by the example of the HT-29 cell culture. The research was carried out in laboratory strain S. aureus ATCC 29213 and clinical strains S. haemolyticus SH39, S. epidermidis SE36-1 isolated from the neonatal autopsy materials. The visual tests of biofilm formation by each strain and testing of the impact of the strains on the cell culture HT-29 was carried out in this work. The two species of CNS form biofilms at a higher rate than S. aureus. Upon incubation for 2 h of HT-29 cells with staphylococcus strains tested in this work, adhesion of bacteria on cell surface was observed. The adhesion was most pronounced in case of S. aureus ATCC 29213 and S. haemolyticus SH39. Upon 3 h of incubation with S. aureus ATCC 29213 and S. haemolyticus SH39, destruction of cell HT-29 monolayer was observed. The incubation for 24 h with the 3 strains tested in this work caused complete destruction of cell HT-29 monolayer. The maximal toxic effect on HT-29 cells was inherent in the strain S. haemolyticus SH39. The aggregate of the results obtained in this work indicates the presence of the pathogenicity factors in the strains S. haemolyticus SH39, which require additional research.

[A porous coral-like structure - a new insight into the morphology of the inner limiting membrane of the retina?] / Poristaya korallovidnaya struktura ­ novoe predstavlenie o morfologii vnutrennei pogranichnoi membrany setchatki?

To date, the internal limiting membrane (ILM), specifically, the side facing the retina, has never been studied by two parallel, mutually complementary methods. This is an attempt to explain favorable results of ILM peeling in various macular pathologies. AIM: By employing scanning (SEM) and transmission electron microscopy (TEM), to identify morphological features of epiretinal samples removed during vitrectomy in patients with lamellar macular hole (LMH) or epiretinal membrane (ERM). MATERIAL AND METHODS: We studied 23 eyes of 23 patients divided into two groups. The first group (13 samples, 11 eyes) consisted of patients with LMH; the second (12 samples, 12 eyes) - with ERM. The surgeries yielded a total of 21 epiretinal samples peeled simultaneously with the ILM and 4 epiretinal samples (2 eyes) peeled in two parts, the second part containing the ILM. One half of the samples was studied by SEM without prior dehydratation, the other - by TEM. RESULTS: The study revealed a high degree of ultrastructural similarity between the two groups of ILM samples. Judging from SEM findings, two sides of the membrane were clearly identified. Porous coral-like structures (PCS) were discovered on the side facing the retina. TEM in the area of PCS discovered parallel arrangement of multiple Muller cell (MC) bodies and processes separated by wide layers of the intercellular matrix. The vitreal side of all ILM samples was notable for numerous fibroblast-like cells. Many variously shaped petrified structures were found on both sides of the membrane. CONCLUSION: During the so called ILM peeling, the surgeon removes a layered structure that includes the basal membrane of MC, cells and fibers attached to its vitreal side, and one more layer comprised by PCS and rather readily torn off from the main massif. The functional significance of this previously unknown structure as well as the effect of its partial removal during surgical manipulations with neurosensory retina in the macular region is yet to be investigated.

[Efficacy of polyprenyl phosphates in the experimental genital herpes model].

An experimental model of the primary genital herpes (herpes simplex type 2, HSV-2) in the female guinea pigs was suggested to study the infectious process activity of polyprenyl phosphates (PPP) and PPP+acyclovir (AC) complex treatment. The morphofunctional features of the guinea pig ovaries were studied in the control and experimental groups (the latter were inoculated with PPP and/or AC as a primary infection treatment) at the stage of the recurrent genital herpes aggravation. It was shown that in the case of combined PPP +AC use significant changes in the disease symptoms were observed, as well as a decrease in the infectious process activity and duration, and positive remote effect on the ovarian morphophysiology.

[MICROBIAL COMMUNITIES ON KIDNEY STONES].

The clinical material obtained surgically in patients with kidney stone disease (KSD) was tested for content of the stone microflora using PCR and standard microbiological methods. It was demonstrated that about 50% of stones in patients with KSD were infected with various infection agents as observed using standard microbiological and molecular genetic methods. The percentage of detection of the Mycoplasma hominis using cultural method is lower than the percentage detected using PCR, which is due to difficult isolation and cultivation, as well as DNA fragments of mycoplasma observed after antibiotic therapy. Studies based on modern microscopy methods showed that microorganisms on the surface of the kidney stone formed multispecies biofilms.

[THE CHARACTERISTICS OF MORPHOLOGY OF BIOFILM OF PERIODONTIUM UNDER INFLAMMATORY DISEASES OF GUMS (CHRONIC CATARRHAL GINGIVITIS, CHRONIC PERIODONTITIS, CANDIDA-ASSOCIATED PERIODONTITIS) ACCORDING RESULTS OF ELECTRONIC MICROSCOPY].

The study was carried out to analyze morphology of biofilm of periodontium and to develop electronic microscopic criteria of differentiated diagnostic of inflammatory diseases of gums. The scanning electronic microscopy was applied to analyze samples of bioflm of periodont from 70 patients. Including ten patients with every nosologic form of groups with chronic catarrhal periodontitis. of light, mean and severe degree, chronic catarrhal gingivitis, Candida-associated paroperiodontitis and 20 healthy persons with intact periodontium. The analysis was implemented using dual-beam scanning electronic microscope Quanta 200 3D (FEI company, USA) and walk-through electronic micJEM 100B (JEOL, Japan). To detect marker DNA of periodont pathogenic bacteria in analyzed samples the kit of reagentsfor polymerase chain reaction "MultiDent-5" ("GenLab", Russia). The scanning electronic microscopy in combination with transmission electronic microscopy and polymerase chain reaction permits analyzing structure, composition and degree of development of biofilm of periodontium and to apply differentiated diagnostic of different nosologic forms of inflammatory diseases of periodontium, including light form of chronic periodontitis and gingivitis. The electronic microscopical indications of diseases ofperiodontium of inflammatory character are established: catarrhal gingivitis, (coccal morphological alternate), chronic periodontitis (bacillary morphological alternate), Candida-associated periodontitis (Candida morphological alternate of biofilm ofperiodontium).

[STUDY OF COLONIZATION PROCESSES AND PERSISTENCE OF MICROORGANISMS IN ARTIFICIAL MATERIALS FOR MEDICAL USE].

AIM: Study processes of microbial colonization and persistence of microorganisms in polymer materials for medical use. MATERIALS AND METHODS: Samples (1 x 1 cm plates) of polymer plastics for production of removable dental prosthesis based on polyurethane and acryl were used, that were incubated with clinical isolates of Pseudomonas aeuruginosa, Staphylococcus aureus in Luria-Bertani broth nutrient media for 24, 48 hours and 7, 14 days and for 1, 5 and 3 months at a temperature of 37 degrees C. Dynamics of interaction process of microorganisms with polymer materials were studied using scanning electron microscope Quanta 200 3D (FEI Company, USA). The samples were fixated after incubation with 10% of neutral formaldehyde, dehydration with alcohols or acetone, typical for SEM, was not carried out, that allowed to conserve the native structure of the samples, including exo-cell matrix of biofilms. RESULTS: Electron-microscopical data on stages of interaction of bacteria with the surface of medical plastics were obtained. Biofilms were shown to be formed on abiotic surfaces and biodestructive changes of plastics appeared. A question on the possibility of prolonged persistence of pathogenic for human microorganisms in artificial prosthesis is discussed. CONCLUSION: The developed experimental model of formation of biofilm on abiotic surfaces could be the basis for carrying out studies directed on the fight with biofilms, by using SEM.

[Structural changes of the adrenal cortex in experimental genital herpes virus infection].

The reactive changes in the adrenal gland cortex were studied in mature female guinea pigs (n=5) in an experimental model of acute genital herpes virus infection. The methods of light and transmission electron microscopy were used. To confirm the presence of viral antigen in the corticosterocytes (CSC), the methods of immunfluorescence and electron microscopic immunocytochemistry were used. It was shown that at day 7 of an acute process, focal CSC reactive changes appeared in the glomerular zone - at the light microscopic level, CSC had intact nuclei and optically empty cytoplasm, while at the electron microscopic level, these CSC demonstrated the damaged membranous organelles, and various membranous structures which were not found in the normal cells. The aggregates of hypertrophied CSC were found in the fasciculate zone. The changes described were reversible, as they practically disappeared by the onset of spontaneous recovery (day 21 after inoculation). The regeneration of CSC of glomerular and fasciculate zones of the adrenal cortex involves both intracellular and cellular mechanisms.

[Infectious genesis of nephroliths (electron-microscopic study)].

Scanning electron microscopy and x-ray microstructural analysis were employed in the study of nephroliths from patients suffering from nephrolithiasis. Bacterial biofilms, urease producing microorganisms, alkaline reaction of the urine are basic factors for local urine crystallization, formation of the base of the nephroliths and its rigid fixation to the pelvic mucosa. Mechanic trauma of the pelvic tissues by the concrement results in destruction of the pelvic mucosa epithelium at the site of the nephrolith. Subsequent inflammation in the underlying connective tissue contributes to formation of connective tissue commissures fixing the conrement in the kidney. It is shown that bacteria as a part of a biofilm are capable to persist in nephroliths for a long time. Destruction of the stones during operation or lithotripsy can trigger activation of growth of bacteria integrated in the biofilm and cause septic complications. Preservation of commissures with elements of the destroyed stone after lithotripsy or surgical removal is one of the leading causes of recurrent nephrolithiasis.

[Health conditions and physical development of soldiers during enrollment in the Armed Forces of Ukraine and military service in 2001-2010].

Fitness of soldiers in military reserve for military service at the stage in the Armed Forces of Ukraine has been studied in the article. It has been established that the growing number of soldiers in military reserve with changes in health and physical condition indicates insufficient level of their health which has a negative impact on their capability and gradually on their fitness for military service. Priorities of changes in organization of the process of completion by human resources of the soldiers' military reserve in the Armed Forces of Ukraine during their transition towards professional army have been defined, to include optimization of criteria of fitness for military service.

[Features of legionella biofilm formation in artificial and natural water systems].

AIM: Study the ability to form monospecies and associative biofilms as a characteristic oflegionella strains and features of organization of natural legionella biofilms in potentially dangerous water systems. MATERIALS AND METHODS: Comparative evaluation ofthe ability of 28 strains of Legionella spp. to form biofilms was determined in water according to previously developed procedure. Samples from biofilm of industrial enterprise coolers and systems of hot water supply of public buildings (hotels, trade centers, hospitals) were studied. Biofilms were studied by scanning and transmission electron microscopy methods. RESULTS: Legionella strains are divided into 3 groups by the ability to form biofilms. L. pneumophila BLR-05 strain that has the most pronounced ability to form monospecies biofilm and persistence in association with Pseudomonas aeruginosa was detected. Formation of massive legionella biofilm in association with bacteria of other taxonomic groups was detected on protective antibacterial filters in the system of hot water supply of a department of a therapeutic prophylaxis institution in the course of 2-3 weeks. Legionella biofilms on the surface of coolers resemble an aggregate of fungi, bacteria and blue-green algae enclosed into matrix. CONCLUSION: The ability to form artificial monospecies and associative biofilm may be a useful characteristic of legionella strains for evaluation of their adhesion and be used to evaluate epidemiological significance of the isolated strains. Prevention of formation of natural legionella biofilms in potentially dangerous water systems is necessary as an essential component of modern strategy of legionellosis prophylaxis.

[Renal calculus microflora in urolithiasis and search for agents of control of biofilms formed by uropathogenic bacteria].

AIM: Study bacterial biofilms in native material (renal calculus) by electron microscopy method and developmeit of biofilm model by isolates in vitro on sterile calculi of various chemical composition. MATERIALS AND METHODS: Bacterial spectra of microflora of renal calculus lavages were studied, isolated pure cultures were identified up to species. Comparisons of urine microflora obtained before operation in patients with urolithiasis with microflora of removed renal calculi were carried out. RESULTS: Urease activity and genes coding pathogenicity factors were detected, and the ability to form biofilms by isolates was studied. Model of formation of biofilms in vitro on sterile renal calculi was developed and candidate agents reducing the biofilm forming ability were tested. CONCLUSION: Uropathogenic microorganisms infecting renal calculi and forming biofilms on them not only support chronic infection by increased resistance to therapy but also facilitate novel lithogenesis.

Development of Chlamydial Type III Secretion System Inhibitors for Suppression of Acute and Chronic Forms of Chlamydial Infection.

The Type III secretion system (T3SS) is currently considered to be one of the main pathogenicity factors in Gram-negative bacteria, which exhibit different types of parasitizing activity. The presence of this structure is essential for the development of an acute infection; the chronicity of the infection is fundamentally dependent upon its functioning. In this regard, T3TS is one of the most promising targets for the development of broad-spectrum antimicrobial drugs that do not develop resistance and are efficacious for the acute and chronic forms of infection. The mechanism of action in drug development is based on the specific inhibition of T3SS, which should interrupt the infectious process, thereby enabling the immune system to eliminate the pathogen. As a result of pilot screening using specific cellular and bacterial tests, followed by chemical optimization and detailed characterization of the biological activity, a new class of chlamydial T3SS inhibitors was obtained. The selected compounds have obvious advantages over the currently available inhibitors of T3SS pathogens thanks to the high inhibitory activity of these compounds with minimal damaging effects on eukaryotic cells. Preclinical trials of the selected inhibitors are currently under way.

[Biofilms of pathogenic bacteria and their role in chronization of infectious process. The search for the means to control biofilms].

The role of biofilms in the environment and in the hosts of pathogenic bacteria has recently attracted much attention of researchers. Microorganisms form biofilms at any biotic and abiotic surfaces and thereby cause serious problems in medical practice and other areas of human activity. Biofilms have been shown to be pathogenetic factors responsible for chronization of infectious process. The data are presented illustrating ubiquitous nature of biofilms, their structural and functional characteristics, and modern methods for the study of microbial communities. The discussion is focused on the role of biofilms in chronization of infectious process, enhanced resistance of biofilm organisms to antibiotics and its underlying mechanisms. Approaches to the search for new means for biofilm control during chronic infections are considered.

[Modern peculiarities of dynamics and structure of servicemen of Armed Forces of Ukraine retiring in transitional period towards professional Army].

Operational readiness of personnel of Armed Forces of Ukraine was studied in the article based on peculiarities of dynamics and structure of retiring of all categories of servicemen. It was established that high rates of retiring related to specific classes of diseases indicate low health condition of servicemen which has a negative impact on their combat readiness and further fitness for military service. Priorities of changes in the process of completing of the Armed Forces of Ukraine with human resources in the conditions of transformation to professional Army, to include optimization of fitness for military service criteria, were defined.

[Characteristics of Armed Forces of Ukraine completing with conscripts and peculiaritis of life style and health of recruits].

Peculiarities of categories of servicemen of the Armed Forces of Ukraine along with health condition of recruits were studied in the article in order to define factors that have an influence on servicemen' health and play in favour of its worsening during the service time. It was identified that negative attitude of young people towards military service is connected with poor living conditions, unsatisfactory medical care, low level of life, bad quality of food, unwillingness to do physical training and sports. Priorities for the changes in organization of completing of the Army with human resources in the conditions of transformation to the completing of Armed Forces with contract servicemen were defined in the article. This is to include optimization of fitness for the military service criteria for young people.

[Intraovarian regulators of folliculogenesis].

The review summarizes information about the intraovarian modulators of folliculogenesis. Consistently described auto-and paracrine factors and mechanisms involved in the regulation of follicular development from the entry in the growth of primordial follicles before ovulation of the dominant follicle.

[Role of salmonella in biliary lithogenesis].

By the methods of light microscopy and immunocytochemistry studies of interaction between S. typhimurium and corpuscular biliary components was investigated in experimental model "bile-bacteria" It was shown that the results of this interaction was bacterial-biliary sludge formation. Bacterial extracellular mucopolysaccharides matrix and flagella's play crucial role in mechanism of sludge formation.

[Modern aspects of diagnostics of chronic chlamydiosis caused by persisting forms of Chlamydia].

AIM: To study the possible hematogenic route of dissemination of Chlamydia trachomatis and to analyze efficacy of methods of pathogen detection in clinical specimens (sera and scraping material). MATERIALS AND METHODS: Cultural method, electron microscopy, real-time PCR, immunofluorescent assay. RESULTS: C. trachomatis was detected in blood by using 2 tests (culture and PCR) in 95.2% of patients with confirmed Chlamydia infection. Chlamydia isolated from blood had infectious properties that could point to the presence of weakly studied hematogenic route of dissemination of C. trachomatis in host's organism. Study of diagnostic value of pathogen detection in serum showed that in case of chronic diseases of urogenital tract as well as extragenital diseases, rate of C. trachomatis detection in serum was significantly higher (61.1% of cases compared to 16.7% in scraping material). CONCLUSION: It is the first time when data about possible circulation of C. trachomatis in blood of patients was obtained. Detection of C. trachomatis in serum of patients with chronic and complicated forms of chlamydiosis provides essentially new approach for direct identification of the pathogen irrespectively from localization of infection's locus.

[Detection of coronavirus in experimental pneumocystic pneumonia].

Transmission electron microscopy (of ultrathin sections) was used to examine the biomass of lung tissue in the immunodeficiency minipigs experimentally infected with Pneumocystis carinii. The material was found to contain pneumocysts, bacteria, and coronaviruses. There was a clear coronavirus-pneumocyst structural relationship. The findings suggest the combined effect of microorganisms of different systematic groups on the development of a pathological process in the experimental infection etiologically determined by Pneumocystis carinii.