[Low-Molecular Thiols as a Factor Improving the Sensitivity of Escherichia coli Mutants with Impaired ADP-Heptose Synthesis to Antibiotics].

Low molecular-weight thiols as glutathione and cysteine are an important part of the cell's redox regulation system. Previously, we have shown that inactivation of ADP-heptose synthesis in Escherichia coli with a gmhA deletion induces the oxidative stress. It is accompanied by rearrangement of thiol homeostasis and increased sensitivity to antibiotics. In our study, we found that restriction of cysteine metabolism (ΔcysB and ΔcysE) and inhibition of glutathione synthesis (ΔgshAB) lead to a decrease in the sensitivity of the ΔgmhA mutant to antibiotics but not to its expected increase. At the same time, blocking of the export of cysteine (ΔeamA) or increasing import (Ptet-tcyP) into cells of the oxidized form of cysteine-cystine leads to an even greater increase in the sensitivity of gmhA-deleted cells to antibiotics. In addition, there is no correlation between the cytotoxic effect of antibiotics and the level of reactive oxygen species (ROS), the total pool of thiols, or the viability of the initial cell population. However, a correlation between the sensitivity to antibiotics and the level of oxidized glutathione in cells was found in our study. Apparently, a decrease in the content of low-molecular-weight thiols saves NADPH equivalents and limits the processes of protein redox modification. This leads to increasing of resistance of the ΔgmhA strain to antibiotics. An increase in low-molecular-weight thiols levels requires a greater expenditure of cell resources, leads to an increase in oxidized glutathione and induces to greater increase in sensitivity of the ΔgmhA strain to antibiotics.

New Spin-Polarized Electron Source Based on Alkali Antimonide Photocathode.

New spin-dependent photoemission properties of alkali antimonide semiconductor cathodes are predicted based on the detected optical spin orientation effect and DFT band structure calculations. Using these results, the Na_{2}KSb/Cs_{3}Sb heterostructure is designed as a spin-polarized electron source in combination with the Al_{0.11}Ga_{0.89}As target as a spin detector with spatial resolution. In the Na_{2}KSb/Cs_{3}Sb photocathode, spin-dependent photoemission properties were established through detection of a high degree of photoluminescence polarization and high polarization of the photoemitted electrons. It was found that the multi-alkali photocathode can provide electron beams with emittance very close to the limits imposed by the electron thermal energy. The vacuum tablet-type sources of spin-polarized electrons have been proposed for accelerators, which can exclude the construction of the photocathode growth chambers for photoinjectors.

[Inactivation of Terminal Oxidase bd-I Leads to Supersensitivity of E. coli to Quinolone and Beta-Lactam Antibiotics].

In cells of Escherichia coli, terminal oxidase bd-I encoded by the cydAB gene catalyzes the reduction of O2 to water using hydroquinone as an electron donor. In addition to the cydAB operon, two other genes, cydC and cydD, encoding the heterodimeric ATP-binding cassette-type transporter are essential for the assembly of cytochrome bd-I. It was shown that inactivation of cytochrome bd-I by the introduction of cydB or cydD deletions into the E. coli chromosome leads to supersensitivity of the bacteria to antibiotics of the quinolone and beta-lactam classes. The sensitivity of these mutants to antibiotics is partially suppressed by introduction of a constitutively expressed gene katG under the control of the Ptet promoter into their genome. The increased level of hydrogen sulfide resulting from the introduction of the mstA gene, encoding 3-mercaptopyruvate sulfurtransferase, under the control of the Ptet promoter, leads to the same effect. These data demonstrate the important role of cytochrome bd-I in the defense of bacteria from oxidative stress and bactericidal antibiotics.

[Enhancement of the Bactericidal Effect of Antibiotics by Inhibition of Enzymes Involved in Production of Hydrogen Sulfide in Bacteria].

Counteraction of the origin and distribution of multidrug-resistant pathogens responsible for intra-hospital infections is a worldwide issue in medicine. In this brief review, we discuss the results of our recent investigations, which argue that many antibiotics, along with inactivation of their traditional biochemical targets, can induce oxidative stress (ROS production), thus resulting in increased bactericidal efficiency. As we previously showed, hydrogen sulfide, which is produced in the cells of different pathogens protects them not only against oxidative stress but also against bactericidal antibiotics. Next, we clarified the interplay of oxidative stress, cysteine metabolism, and hydrogen sulfide production. Finally, demonstrated that small molecules, which inhibit a bacterial enzyme involved in hydrogen sulfide production, potentiate bactericidal antibiotics including quinolones, beta-lactams, and aminoglycosides against bacterial pathogens in in vitro and in mouse models of infection. These inhibitors also suppress bacterial tolerance to antibiotics by disrupting the biofilm formation and substantially reducing the number of persister bacteria, which survive the antibiotic treatment. We hypothesise that agents which limit hydrogen sulfide biosynthesis are effective tools to counteract the origin and distribution of multidrug-resistant pathogens.

[RNA-Chromatin Interactome: What? Where? When?]

It was noticed in the early 1960s that a large amount of RNAs is associated with chromatin. What kind of RNAs are they? Where are they located on chromatin? When and in what processes do these RNAs perform their physiologically normal or pathogenic functions? The review describes the modern approaches that help, to some extent, to answer these questions. Consideration is given to the experimental methods that make it possible to obtain the complete RNA-chromatin interactome of a cell or the genome-wide interaction maps of individual RNAs with chromatin, as well as the methods to process the experimental data. Focus is placed on the noncoding RNAs that function in close contact with chromatin and chromatin-associated protein complexes. A variety of biological examples described using the methods makes it clear that RNAs interacting with chromatin play an important role in the functions of cell systems by finely tuning the chromatin architecture and thus changing the level of gene expression.

[Ultrastructure of neointima of native and artificial elements of the blood circulatory system]. / Ul'trastruktura neointimy nativnykh i iskusstvennykh elementov sistemy krovoobrashcheniya.

OBJECTIVE: To compare the neointima structure in conduits for coronary bypass grafting, bioprosthetic heart valves, tissue-engineered vascular grafts, and metal stents. MATERIAL AND METHODS: The objects of the study were the fragments of the human internal thoracic artery, experimental biodegradable vascular prostheses, leaflets of xenopericardial bioprostheses of heart valves, and fragments of stented vessels. Tissue samples were fixed in formalin and post-fixed in osmium tetroxide. After dehydration and epoxy resin embedding, the samples were ground and polished. Samples were counterstained with uranyl acetate and lead citrate and visualized by means of backscattered scanning electron microscopy. RESULTS: Neointimal pattern in all samples was similar. Neointima was comprised of endothelial cells, smooth muscle cells, fibroblasts, and the extracellular matrix. Endothelial cells showed significant diversity both between different elements of the circulatory system and within the same tissue, having either elongated or polygonal shape. Adhesion of leukocytes testified to the endothelial cell activation. In the absence of inflammation in the superficial layer of the neointima, the arrangement of smooth muscle cells and extracellular matrix fibers was parallel to the endothelium. Clusters of foam cells were frequently detected around the neointimal layers with solid inclusions (metal stents or calcium deposits). Thickening of the neointima was accompanied by the presence of capillaries and capillary-like structures. CONCLUSION: Neointima formation is a typical response to the damage inflicted to the elements of the circulatory system. Neointima underwent a constant remodeling characterized by an altered cellular composition, macrophage invasion, neovascularization, and calcification.

[Modern aspects of the treatment of submacular hemorrhages secondary to macular degeneration]. / Sovremennye aspekty terapii submakulyarnykh krovoizliyanii na fone makulyarnoi degeneratsii.

Age-related macular degeneration complicated by submacular hemorrhage if not treated timely leads to permanent loss of central vision. The basis of effective therapy is its early start, dislocation of blood from central parts of the retina and blockage of neovascularization. This review examines the current methods of treatment of submacular hemorrhages, anatomical and functional outcomes, risks of postoperative complications, as well as trends in the use of combined methods of surgical intervention.

Standardness of antimicrobial discs by various manufacturers in the light of import substitution.

In connection with the introduction of economic sanctions against the Russian Federation by the United States, the European Union, Japan, and a number of other countries, import substitution is becoming one of the strategic objectives of the Russian economy The comparative assessment of standardness and effectiveness of six antimicrobial commercial producer-varying disks for clinical and test strains of microorganisms has been conducted. It is based on analyzing of the variation coefficient (Cv) of inhibition zone diameters, mean values, and diameter values ranges, which are indicators of reproducibility, accuracy and efficiency, respectively. Discs from BD BBL result in insignificant variations of inhibition zone diameters, providing accurate and reproducible data. Imported discs available on the national market from a friendly country produced by HiMedia Company are showed more variation and inferior in quality to BD BBL ones. There are satisfactory results for some domestic discs, but discs with imipenem, meropenem, ciprofloxacin and gentamicin are not standard enough to assure reproducible results. To have reliable data on antibiotic susceptibility of infectious agents, incoming and routine quality control of antimicrobial disks is needed as a compulsory procedure for every bacteriological laboratory.

Analysis of the level of optopedical rehabilitation in studying the microbiome and evaluation of the degree of fixation of removable plated prosthesis.

The widespread use of traditional removable prosthetics is explained by the relative simplicity of the technological stages of manufacture and determines its availability. The development of prosthetic stomatitis of the oral cavity is facilitated by poor fixation and stabilization of removable orthopedic structures. Microbiome biofilms formed on the surface of dental orthopedic structures can help reduce their service life and cause an inflammatory process of the oral cavity of microbial etiology during dental prosthetics in the process of orthopedic rehabilitation. The purpose of the study: to assess the level of adaptation of patients during orthopedic rehabilitation based on the study of the microbiome and the assessment of the degree of fixation of removable lamellar dentures. Qualitative and quantitative characteristics of the microbiome of prostheses at the stages of orthopedic pealitation were assessed; facultative anaerobic species belonging to the genera Staphylococcus, Micrococcus, Enterococcus, Streptococcus, Klebsiella prevailed;noted the elimination of microorganisms of the genera Bifidobacterium and Lactobacterium, yeast-like fungi of the species Candida albicans were isolated. An analysis of the index of fixation of prostheses showed an increase depending on the duration of use; a good level of fixation of prostheses was established in groups of patients.

Effectiveness of using liquid transport media in bacteriological diagnostics of diphtheria infection.

The results of evaluating the effectiveness of the use of liquid transport media at the preanalytical stage of bacteriological diagnosis of diphtheria infection are presented. A typical toxigenic strain of C. diphtheriae biovar gravis № 665 was used. The experiments were carried out using a laboratory-prepared medium based on GRM-broth (State research center for applied biotechnology and microbiology, Obolensk), a transport system with a fleecy probe swab (DELTALAB) and a transport system ∑-Transwab ® with a polyurethane Sigma-swab (Medical Wire & Equipment Co. (Bath) Ltd.). The tampons were pooled with a 24-hour bacterial culture of C. diphtheriae, then immediately seeded on Tellurite-containing blood agar. Storage conditions were simulated for 6-24 hours: at room conditions +(20-25)° C, in the refrigerator +(4-8)° C, in a thermostat +(37±1)° C. Storage of C. diphtheriae was most optimal on two liquid transport systems in a refrigerator +(4-8)° C for 6 and 24 hours; in room conditions +(20-25)° C - there was a decrease in seeding after 6 hours and loss of pathological material after 24 hours, more pronounced on a fleecy probe swab; under thermostat conditions +(37±1)° C on both transport systems, a decrease in seeding was noted after 6 hours and a complete loss of pathological material after 24 hours. The results obtained demonstrated the efficiency of using the Amies liquid transport medium and justify the need to develop a domestic analogue of the transport system based on the Amies liquid medium for the bacteriological diagnosis of diphtheria infection.

Isolation and identification of listeria in clinical material.

Results from research on isolation, identification, and study of biological properties of L. monocytogenes clinical isolates and Listeria spp test strains are presented. Peculiarities of modern research methods for indicating and identifying pathogenic listeria to improve the quality of laboratory studies of clinical material are studied. The culture method provides reliable results of microbiological analyses upon detecting Listeria spp. The presented list and algorithm of the laboratory diagnostic methods can be used as a basis for elaborating regulatory documents for carrying out microbiological research on any biological material for the presence of bacteria of the genus Listeria spp. and L. monocytogenes species in it.

Analysis of micro-relief of biofilm of yeast Candida albicans of basic plastics by the method of laser modulation interference microscopy.

The development of mycotic colonization of the base surface with further biodegradation of acrylic plastics is currently of undoubted interest. The oral cavity is a favorable ecological niche for colonization by fungi and their subsequent possible invasion into the epithelium of the oral mucosa. The method of modulation interference laser microscopy is of considerable interest to researchers in medicine in the context of obtaining the necessary information about the morphological characteristics of microbial cells and the microbiome community as a whole during the colonization of a certain ecological niche in the human body. Purpose of the study: to analyze the microrelief of the biofilm of yeast-like fungi of the species Candida albicans of base plastics of the hot type of polymerization using the method of laser modulation interference microscopy. An experimental study was carried out in order to study biofilms of yeast-like fungi of the genus Candida on samples of basic plastics, an image of a biofilm of yeast-like fungi of the species Candida albicans was obtained on the surface of a plastic of a hot type of polymerization (polymethyl methacrylate) in the visualization of the phase portrait, a description of its horizontal and vertical bioprofile. As a result of the research, the heterogeneous structure of the biofilm was determined, due to the different density and accumulation of cells along the surface, the characteristics of the surface were established in accordance with the roughness criteria. The microrelief parameters on a separately arbitrarily selected section line allow one to determine the characteristics of the biofilm in the required area and make it possible to judge the nature of its formation in a certain biological niche.

Effect of different types of dry swabs and their storage conditions on the inoculation of Corynebacterium diphtheriae.

The results of evaluating the effectiveness of C. diphtheriae inoculation using different types of dry swabs in studies simulating various conditions of its storage at the preanalytical stage of a laboratory study for diphtheria are presented. A typical toxigenic strain of C. diphtheriae biovar gravis No. 665 was used. A commercial dry, sterile cotton swab probe (Ningbo Greetmed Medical Instruments Co., LTD, China), a commercial dry, sterile swab probe (plastic and viscose) (COPAN, Italy), tufters with a fluffy probe-tampon on a polystyrene applicator, standard (DELTALAB, SL, Spain). The tampons were pooled with a 24-hour bacterial culture of C. diphtheriae, then immediately seeded on Tellurite-containing blood agar and Corynebacagar. Storage conditions were simulated for 3 hours: at room conditions +(20-25)°C, in the refrigerator +(4-8)°C, in a thermostat +(37±1)°C. Optimal storage of C. diphtheriae on all three types of dry swabs at + (4-8) ° C; at +(20-25)° C - growth is observed when seeding from a cotton swab; in a swab with a fleecy probe-tampon, a decrease in the inoculation of C. diphtheriae was noted; when using a viscose swab - a significant loss of C. diphtheriae. At +(37±1)°C, a significant decrease in the inoculation of C. diphtheriae on all three types of tampons was noted, up to the absence of growth when using a viscose tampon. To exclude the loss of C. diphtheriae, it is necessary to observe the conditions for taking and storing biological material at the preanalytical stage of a laboratory study, which will improve the quality of laboratory microbiological studies for diphtheria infection.

Immune and microbiological disorders in women with infectious and inflammatory pathology of the cervix.

Penetration into the female genital tract of infectious agents is associated with the cause of the onset of cervicitis. In most cases, these are sexually transmitted diseases (STDs). A high level of viral and bacterial load in the study of clinical material from women with infectious and inflammatory pathology (IVP) of the cervix affects the activity of innate immunity reactions. The total number of examined patients was 76 patients of reproductive age, of which 36 women with cervical IVP and 40 women made up the comparison group. The imbalance of microbiocenosis can be accompanied by immune and microbiological disorders. The aim of the study was to study immune and microbiological disorders in women with infectious and inflammatory pathology of the cervix. To achieve the goal, the following tasks were set: to identify the relationship between the expression level of TLR2 and TLR4 genes in the epithelial cells of the cervical canal of women in which UPM is determined. To determine the species spectrum and etiological significance of the microbiota of bacterial biofilm of the cervical mucosa in the pathogenesis of cervicitis in women of reproductive age. It has been established that the determination of indicators of innate immunity, such as interferons, in blood serum and the expression of TLR2 and TLR4 receptors, with the study of their balance, taking into account the pro-inflammatory and anti-inflammatory properties in IVP of the cervix, has an important prognostic value. An increase in the level of the immune response in the form of hyperstimulation of antigens of IVP pathogens leads to the chronization of the inflammatory process in the urogenital tract, to scarring of tissues, which can play an important role in the development of infertility and termination of pregnancy in women of reproductive age.

Bloodstream infection in hospital therapeutic patients.

Bloodstream infection (BI) is the cause of high mortality. Hospital bloodstream infection (HBI) complicates hemodialysis, pneumonia, oncohematological diseases. Positive hemoculture obtaining depends on the volume of blood inoculation, the number of blood samples, the incubation time. To test the principles of microbiological culturomics in the diagnosis BI of hospital patients with a therapeutic profile. 848 hospital cardiac patients with suspected BI were included. 10 ml of blood were taken intravenously with a syringe, blood was inoculated into 200 ml of the heart-brain medium (HBM) in an anaerobic bottle. It was incubated for 7 or more days in a thermostat at +37º C. The hemocultures were obtained in 64.3% of cases with paired blood sampling with an interval of 30 minutes whereas an increase in the number of blood samples reduced the effectiveness of obtaining hemocultures to 9.1%. When incubating bottles for more than 7 days there were obtained 200 additional hemocultures containing 239 strains of microorganisms. Episodes of HBI were observed more often in the cases of the circulatory system (77.8%), including infectious endocarditis (IE) (47.0%), rheumatism (22.1%), myocarditis (14.6%). Episodes of HBI occurred more often in men with IE and coronary heart disease, in women - with rheumatism and myocarditis. Patients aged 45-75 were in the group of risk with a probability of complications of HBI up to 73.7%. When examining the blood of 848 hospital patients of cardiological profile HBI was detected in 38.3% of cases. Among clinical isolates gram-positive cocci with a great number S.epidermidis prevailed. Polymicrobial hemocultures (16.3%) were characterized by two and three associates in one blood sample. Among the hematological indicators in HBI there were: leukocytosis, increased ESR, lymphocytosis, decreased hemoglobin; increased values of fibrinogen, CRP, γ-globulin, α2-globulin, low levels of total protein and A/G coefficient. The techniques of microbiological culturomics were used. HBI was diagnosed in 38.3% of the therapeutic patients of cardiological profile. The etiology of HBI was characterized by polymicrobicity in 16.3% of cases. Hematological markers of HBI were identified.

The microbiome of oral cavity patients with periodontitis, adhesive and biofilm forming properties.

The work characterizes the intestinal microbiota of patients with ischemic stroke, including the spectrum, frequency and number of microorganisms, as well as the spectrum and amount of gas signaling molecules secreted by lactobacilli. It was found that in patients with ischemic stroke, the frequency of the main representatives of normal microflora, Bifidobacterium spp., Lactobacillus spp., Escherichia coli, decreased in 2-3 times, and the same time the prevalence of Clostridia spp., Bacillus spp., Peptostreptococcus spp., Klebsiella spp. increased in 2-3 times; yeast like fungi C. albicans was isolated in 25% of cases. Lactobacilli isolated from the intestinal microbiota of patients with ischemic stroke were represented by a wide variety of species: L. rhamnosus, L. fermentum, L. plantarum, L. brevis, L. pentosus, L. curvatus, L. salivarius. In most cases, they did not produce NO, they released CO 2 times less compared to healthy people. The most active NO producers - L. plantarum, CO - L. rhamnosus.

Development of a PCR assay for the detection of human herpes virus type 7.

A PCR assay has been developed to identify the DNA of the human herpes virus type 7. The search and selection of conserved regions was carried out by comparing the whole genome nucleotide sequences of HHV-7. A fragment duplicated in the HHV-7 genomes was chosen as a target for amplification. The performance of the assay was tested on a synthetic matrix and clinical samples. The developed assay has high sensitivity and specificity and showed good efficiency in detecting HHV-7 DNA in clinical samples.

Evaluation of the structure of the human microbiome in multiple sclerosis by the concentrations of microbial markers in the blood.

The relationship between multiple sclerosis and the state of the human microbiome was studied, namely, the change in the representation of microbiota phylotypes, the proportion of coccal flora, the proportion of anaerobic, gram-negative, proteolytically active microflora, as well as the concentration of markers of bacterial plasmalogen and endotoxin in the blood. Microbiome studies were carried out by gas chromatography - mass spectrometry of microbial markers in the blood. A statistically significant increase in blood concentrations of the total level of microbial markers of bacterial plasmalogen and endotoxin was determined in multiple sclerosis, which may be associated with an increase in the permeability of the intestinal wall. In multiple sclerosis, the proportion of coccal, gram-negative, anaerobic microflora with a proteolytic type of metabolic activity increases. The correlations of the representation of microbiota phylotypes change due to the switching of the direct relationship Proteobacteria-Bacteroides to Proteobacteria-Firmicutes. In multiple sclerosis, Actinobacteria and Proteobacteria increase and Firmicutes decrease. Conclusion. The multiple sclerosis disease may be associated with pathological changes in the structure of the microbiome and the growth of endotoxemia, which may be one of the factors in the pathogenesis of the disease. New laboratory markers for diagnosing and predicting the course of MS have been proposed.

Age changes of foot microbiome estimated by microscopy method.

Despite of great number of investigations in the area of tinea pedis, question is opened: to what extent dermatophyte fungi are spread among modern population and does their occurrence interrelated with host age? Investigated group included 99 volunteers from 14 to 73 years old. Skin scales were collected from heel area of foot, and signs of heel skin trouble were expressed in points. In contrast to usual laboratory microscope magnification x900 we worked at x1750, what allowed to estimate not only fungal, but bacterial forms too. Average abundances of microbial morphotypes were expressed in points. Heel skin trouble increased in the process of aging (Pirsons` coefficient r=0.954). Bacilli occurred in all persons independently from age, but their abundance increased with aging (0.821). On the contrary cocci were more common and abundant in young person`s feet (-0.620). Occurrence of dermatophytes increased with age (0.891), at that relatively high values took place in young persons (10.5% with mycelium and 73.7% with spores) and in group without any heel skin trouble symptoms (7.7% and 76.9%), what allow to refer these fungi to normal habitats of foot skin.

Laboratory diagnosis of community-acquired bloodstream infection in therapeutic pathology.

Community-acquired bloodstream infections (CBSIs) occur in the out-of-hospital setting (44%) and increase the overall mortality from bloodstream infections (BSIs) by 7.2% per year. The development of CBSIs depends on both comorbid and polymorbid diseases and the patients' age. The causes of CBSIs are: respiratory, hepatobiliary gastrointestinal and urogenital tracts and dental interventions. The etiology of CBSIs is characterized by the isolation of coagulase-negative staphylococci (CNS) (32%), E. coli (27%). To investigate community-acquired bloodstream infection in therapeutic patients. The study included out-of-hospital patients (n=382). 4.5 ml of blood were taken intravenously into a closed vacuum system in order to obtain a buffy coat of blood, which was put on glasses for microscopy and Petri dishes with blood agar for cultivating under aerobic and anaerobic conditions. Microorganisms were identified by mass spectrometry. Microscopy of blood smears was used for rapid diagnosis of infection in the bloodstream. BSI was diagnosed in 183 (48.0%) out of 382 out-of-hospital patients. The etiology of CBSIs was studied on 297 isolated strains of microorganisms. CBSIs rather often complicated the underlying disease in women and young people. The spectrum of CBSI pathogens included aerobic and anaerobic bacteria and fungi. Gram-positive cocci with the leadership of S.epidermidis (25.7%) were more often isolated among bacteria. 70% of all isolated pathogens grew under anaerobic conditions. CBSIs were characterized by polymicrobiality (33.5%) of two to four different microorganisms in one blood culture; the species of associates of polymicrobial blood cultures are shown. Microscopic examination of blood smears revealed microorganisms in 97.1% of cases, including associations of bacteria with fungi (66.9%). CBSIs occurred after contour plastic, in diseases of the respiratory system, genitourinary system, oral cavity, skin and subcutaneous tissue. Microbiological examination of the buffy coat is an alternative microbiological method of CBSIs diagnosis, which includes microscopy and blood cultivating and has a high diagnostic efficiency (97.1% and 48% respectively). It can become an option for replacing imported blood culture automated systems.