Visnyk of Dnipropetrovsk University. Biology, medicine

47-51

Authors: I.I. Mykaylo, M.V. Kryvtsova, V.I. Nikolaichuk

The aim of our research was to conduct a monitoring study of nitrate content in plant products of Uzhgorod district and to accomplish comparative analysis of the survey results in different periods of crop ripening. Selection of vegetable samples was carried out in Uzhgorod district in the early spring and summer periods. Determination of the nitrate content was performed using an ion-selective method at the Chemical and Toxicological Department of the Regional State Veterinary Medicine Laboratory in the Transcarpathian region of Ukraine. Vegetables were tested for nitrate content using the ion-selective method with the laboratory ion meter AI-123. Core investigation samples were crushed and homogenized. A 10.0 g weight of the investigated product, which was prepared according to MIR № 5048-89, was placed in a flat-bottomed or a conical flask, which was then filled with 50 cm3 potassium alumens solution and shaken in a shaking-machine for 5 minutes and then transferred into a measuring glass. The nitrate weight fraction in milligrams per kilogram was obtained together with the weight concentration value of nitrate ions in solution. For our study we selected vegetables grown in both public and private gardens of Uzhgorod district, namely: common onions, radishes, garden parsley, cucumbers, tomatoes, bell peppers, white cabbages, carrots and table beets. 25 samples were selected for each type of vegetable. Nitrate content was determined in the early spring growing period (from February 9 to May 27, 2011) and in the summer growing period (from June 3 to September 28, 2011), because in these particular periods we recorded the most frequent cases of food poisoning from nitrates among the population of the region. A clear trend has been traced towards increasing the nitrate content in food plant production, at levels which exceed the maximum permissible concentration (MPC). The results of our research demonstrate that the nitrate content exceeded the maximum permissible concentration in 9 kinds of vegetables out of the 11 selected for the investigation, which composes 82% of total production. In particular, among the selected vegetables an excess of nitrate content, above MPC was recorded in 100% of cucumbers, 92% of carrots, 40% samples of green onions, 40% of radishes, 40% of tomatoes, 28% bell peppers, and 16% of early white cabbages. However, the most significant nitrate excess, which was more than double the MPC, was observed in 56% of cucumbers, 20% of tomatoes, 8% of radishes in the early spring period and 4% of tomatoes in the summer period. Consequently, it has been established that the consumption of early vegetable production contributes to the ingestion by humans of significant amounts of nitrates. The application of an agrochemical system based upon sound measurement of the nitrate content would allow us to solve the task of increasing soil fertility and to form a deficit-free and positive balance of biogenic elements and humus in the "soil – plant – fertilizer" system and develop a system of crop production which is balanced in its chemical composition and nutritional value. To sum up, detailed determination of the factors that lead to the accumulation of nitrates in vegetable crops and the development of methods to reduce nitrate concentrations in crop production require further investigation.

57-62

Authors: O.V. Melnyk, O.P. Kornijchuk, O.I. Pershyn, Z.D. Vorobets

A significant role in the development and course of reactive arthritis (ReA) is played by T-lymphocytes as their development and systemic manifestations are based on immunological mechanisms. Additionally, the pathogenesis of many diseases is linked to changes in the structure and function of ion-transporting systems. Therefore, the aim of the study was to find out the kinetic properties of ATP-hydrolysis reaction involving Na+,K+-ATPase of peripheral blood lymphocytes of healthy individuals and patients with ReA. We used the current methodological approaches to the study of ATPase activity in saponin permeabilized cells. We conducted an analysis of the kinetic properties of ouabainsensitive Na+,K+-ATPase activity of saponin-perforated peripheral blood lymphocytes of healthy donors and patients with rheumatoid arthritis (RеA). We found out that in peripheral blood lymphocytes of patients with RеA primary active transport of Na+, K+ ions is slower and less intensive, though characterised by the same capacity, as in healthy donors. The affinity constant for ATP in peripheral blood lymphocytes in patients with RеA is greater by 2.9 times than its value in comparison with healthy donors. We established that in conditions of rheumatic pathology in immunocompetent cells, inhibition of Na+,K+-ATPase activity is not caused by reduction of speed of enzyme work, but by increase of affinity of ouabainsensitive Na+,K+-ATPase to ATP. At the same time, the Mg2+-binding center of Na+,K+-ATPase in patients with RеA is endogenous. We also found that affinity Na+,K+-ATPase to the ions K+ in peripheral blood lymphocytes of healthy donors is 2.4 times higher than in patients with RеA. We observed that Na+,K+-ATPase of peripheral blood lymphocytes of patients with RеA retains its endogenous receptor properties – sensitivity to ouabain does not change. It is assumed that under conditions of rheumatic pathology the impact on the Na+,K+-ATPase structure occurs both externally and on the cytoplasmic membrane surface. The above experimental data can be used for further clarification of the membrane mechanisms of ion exchange in immunocompetent cells of patients suffering from autoimmune diseases.

71-75

Authors: N.P. Kilochytska

From the epidemiological viewpoint the greatest threat to human health is represented by female mosquitoes in the hematophagous stage. Contact with these bloodsucking insects occurs most frequently in their places of diurnal concentration. The main daytime habitats of mosquitoes in urban areas are recreational areas, especially near water – forests, parks, botanical gardens, cemeteries, green zones in residential areas, residential buildings and buildings with livestock, basements and cellars. In the context of global warming and extreme high summer temperatures a tendency has been observed for mosquitoes to make increasing use of residential premises and outbuildings as a daytime habitat. In the Solomianskyi district of Kiev a six-year monitoring programme of mosquitoes’ diurnal habitat distribution was conducted. It showed that the majority of female mosquitoes which attacked humans (60%) were concentrated in areas of recreation and outdoors in residential areas. Simultaneously a significant number of mosquitoes was found to use as their daytime habitat places where they were in close proximity to humans – in domestic outbuildings (sheds under various uses), basements and cellars. The lion’s share of these mosquitoes was represented by Anopheles maculipennis (25%) and Culex pipiens (7,6%). At the same time a significant number of representatives of the genera Ochlerotatus (O. cantans, O. sticticus, O. cataphylla) and Aedes (Ae. v. vexans) were also recorded using residential premises as their daytime habitat on a scale which had not been previously observed for these species of mosquito. It is possible that female mosquitoes fly into livestock buildings at twilight because they are attracted by electric light and ammonia vapors. However, these factors are almost absent in basements and cellars. It remains to be noted that mosquitoes use basements and cellars as a daytime habitat at periods of hightemperatures and low humidity in the city. To test this assumption the distribution of mosquitoes was analysed during the warm season in 2010 in Solomyanskyi district with measurement of temperature at the locations of collection. It turned out that the air temperature in the sheds was 2–4 °C lower than outside in the shade, in the basements lower by 4–11 °C, and in the cellars lower by 4–12 °C. Comparison of the temperature in the daytime habitats and number of mosquitoes found there showed a direct relation between the outdoor temperature and the number of mosquitoes in the daytime habitat on the premises. The data indicate that there is a tendency for the number of synanthropic blood-sucking mosquito species to increase owing to the occupation of domestic premises as a daytime habitat by those species of mosquitoes for which this phenomenon was not typical earlier. If global warming and the current trend to increase in summer temperatures persist, this can cause a deterioration of the epidemiological situation in the megalopolis.

10-13

Authors: O. Rivis, M. Kryvtsova, V. Nikolajchuk

It is admitted that the normal human microflora plays an important role in supporting homeostasis, forming immune mechanisms and metabolism. Nowadays, there is a constant growth of different diseases due to microbiological imbalance in a human organism. Preparations containing “good bacteria” have been used for therapeutic purposes since ancient times. The mechanism of probiotics influence comprises their ability to compete for adhesion sites with the pathogens, to exhibit antagonistic activity and stimulate the immune system of a host. Most of probiotics commonly used are the spores of Bacillus. Initially the main focus of their use was the prevention of gastrointestinal disorders. So, the use of probiotics in dental practice is a poorly studied area. In recent years, probiotics have been investigated to provide the oral health. Therefore the study of using probiotics for correction of the oral microflora in people with inflammatory diseases of the periodontal tissues is promising. Our previous studies have shown changes in microbial community of an oral cavity in patients with periodontitis. In particular, the reducing number of obligate microorganisms and increasing number of pathogens was demonstrated. The paper describes the current data on the potential benefits and basic properties of the Bacillus spore probiotics, which demonstrate the viability and relevance in dental practice. The study tested antagonistic activity of commercial Bacillus probiotics "Biosporin" ("Biopharma", Ukraine), "Subalinum" ("Biopharma", Ukraine), "Normaflore" ("Sanofi-Aventis Zrt.", Hungary) and "Enterogermina" ("Sanofi-Synthelabo SpA", Italy) against clinical strains of microorganisms isolated from the oral cavity of patients with periodontitis. Thus, further studies on the role of spore probiotics in correction of the oral cavity microflora as a part of complex treatment of periodontitis should be carried out.

23-29

Authors: N. N. Donets, I. E. Sokolova, A. A. Moskalenko, A. L. Drobina, A. I. Vinnikov

The paper presents monitoring results of the spread of opportunistic and pathogenic microorganisms in patients of surgical departments of the Dniprodzerzhynsk city hospital No 7. 1464 strains of bacteria isolated from biological material of the patients from January to December 2012 were studied. Relevant standard methods of research and data interpretation in accordance with the regulatory guidelines were used. The microorganisms’ sensitivity to antibiotics was determined by the disk diffusion method. Assessment of the resistance of isolated microorganisms to antibiotics was made with the software Whonet 5.1. At the first stage of investigation sampling biological material and inoculation in the culture medium were made. The discharges of wounds, throat, nose, ears, vagina and urethra, and also urine from patients of surgical departments were sampled for bacteriological analysis. The main substratum was 5% blood agar. There may additionally be used the selective growth media (yolk-salt agar, Endo, and Saburo). At the second stage we identify microorganisms with bacterioscopic, bacteriological and biochemical methods. Identifying microorganisms of the genus Staphylococcus was made by the reaction of lecithinase presence, plasma-coagulation reaction and the mannitol oxidation reaction. For the identification of bacteria of the family Streptococcaceae the growth pattern in 0.5% sugar medium was used. It was differentiated from bacteria of the genus Enterococcus by plating onto egg yolk agar base and milk with 0.1% methylene blue. Identification of bacteria of the Enterobacteriaceae family was made by studying their colonies on dense differential diagnostic media. Suspicious colonies were transferred on a combined medium for primary identification (Olkenitsky's medium). Then the biochemical signs of enterobacteria were studied in the minimum number of tests. The third phase of the study included the determination of the sensitivity of microorganisms to antibiotics. To do that, we use the disc diffusion method. The antibiogram tests of isolated bacterial strains used Mueller-Hinton agar. According to the data obtained we assign of microorganisms to certain category of sensitivity: sensitive, moderately resistant or resistant. Monitoring of prevalent microorganisms showed that 61% (893 of 1464) isolates were gram-positive bacteria, 696 strains of them are staphylococci. 477 of these are identified as S. aureus, representing 68.5% of all Staphylococcus. We found coagulase negative staphylococci in 192 patients, and the strains of S. haemolyticus are dominated. Strains of the family Streptococcaceae isolated from 197 patients. Among them the E. faecalis accounted for 66% of the total number of Streptococcus. Gram negative bacteria were presented by the families Enterobacteriaceae and Pseudomonaceae. Enterobacteriaceae accounted for 90.7% (518 of 571). E. coli plays the leading role and accounted for 42% of all Enterobacteriaceae. The strains of P. aeruginosa were identified in 53 patients. S. haemolyticus has played an important role as a pathogen as well as S. aureus. Its resistance to antibiotics is much higher than that of S. aureus. Although S. haemolyticus is opportunistic coagulase negative, it can be isolated from patients not only with chronic, but with acute infection. Thus nowadays the strains of S. haemolyticus gained high pathogenic and virulent properties.

35-41

Authors: O. V. Goncharenko, O. O. Konovalova

The aim of this work was to study the mechanisms of action of sub-toxic dose (LD50) of the manganese chloride on cell metabolism in rats’ organs and tissues under model conditions. A model of assessment of integral response of an organism to exogenous manganese at exposure levels close to the threshold that was realized as the repeated impact of sub-toxic doses. White Wistar rats aged 3 months were injected intramuscularly with MnCl2 with the metal concentration of 50 mg/kg for 8 days. Effect of exogenous manganese load on the content of macro- and microelements, such as Ca, Mg, Mn, Cu, Zn, and Pb, was studied in liver, heart, spleen, kidneys, muscles and bones of experimental animals the use of C-115-M1 atomic absorption spectrophotometer. Study of enzymes activity and lipid peroxidation products in serum and liver homogenates of experimental animals was conducted on a spectrophotometer Spekord UV/VIS. Hemolysis of red blood cells was determined by the Yacher method. Sorption capacity of the erythrocytes glycocalyx to alcian blue was determined by the Artsishevsky method. A study of the influence of Mn-load on the functional activity of energy metabolism was performed on a model system of rat liver mitochondria by the polarography. Intake of MnCl2 in a dose of LD50 generates weight loss of the rats by 44%, hemoglobin level decrease by 25%, and reduction of the transaminases activity by 40%. That indicates serious disorders in ions homeostasis and metabolic processes. MnCl2 load leads to a significant increase in the content of manganese in all tissues tested: the largest amount was found in the spleen (600% increase), liver (300%) and muscles (240%) as compared with the initial state. Apparently, these organs are depots of manganese. It was established that the introduction of MnCl2 in a dose of LD50 causes a redistribution of elements in the body, resulting in a significant reduction in the concentration of magnesium ions in heart, bones, muscles, liver and spleen. It was accompanied by increasing calcium content in liver, heart, muscle, kidneys and bones as well as by disorders of Ca/Mg ratios. MnCl2causes significant redistribution of the microelements in the rats’ organs. It is characterized by a decrease of copper, zinc and nickel contents in almost all studied tissues. The most antagonistic effect of manganese manifested in relation to nickel and copper in heart and spleen. A reduction of zinc content was most pronounced in spleen, while its contents in bones and kidneys almost don’t change. The study of the impact of manganese on biochemical parameters of membranes proved for the first time the malfunction of erythrocytes’ membranes. It results in increasing sorption capacity of the red blood cells glycocalyx to alcian blue. Using the erythrocyte model we established that manganese cations cause a significant increase in sorption capacity of the red blood cells (53.4 ± 1.8%) and their osmotic fragility, as evidenced by an increase of spontaneous hemolysis to 42%. The other evidence is the change of surface properties (glycocalyx), which indicated by an increase in the sialic acid content by 60% as compared with the control. The obtained data of the model study of the dynamics of the sorption capacity of erythrocytes glycocalyx to alcian blue, osmotic resistance of erythrocytes, activation of lipid peroxidation and increased level of sialic acid may be a signal that the primary mechanism of manganese intoxication is a damage of cell (plasma) membranes. The data obtained on a mitochondrial model suggests that MnCl2, acting as an antagonist of magnesium, has the ability to disturb respiration and oxidative phosphorylation that inhibits the energy metabolism of a cell. Mitochondrial oxidation of malate+glutamate was affected by MnCl2 in narrow range concentrations 3–4.5 mM that cause disengagement (3 mM) and complete inhibition (4.5 mM). The effectiveness of manganese intoxicated rats treatment with the poly-elemental phyto-complex “Valeoton” and zinc-pectin (drug-antagonist of manganese) was experimentally proved. Thus, the overall effect of manganese is its accumulation in organs and antagonistic relations to all divalent metals. Experimental results with manganese chloride loading point not only to changes in the concentrations of elements and their redistribution, but also to serious disturbances in regulation of ions homeostasis.