[USE OF LASER RAMAN-LUMINESCENT TECHNOLOGIES FOR EVALUATION OF QUALITY OF MEAT PRODUCTS AND DETERMINATION OF THE DEGREE OF THEIR BACTERIAL CONTAMINATION].

AIM: Determination of the effect of microorganisms on spoilage of meat products during various temperature regimes of storage by integral indexes of luminescent lines in their spectra and development of an algorithm of microorganism indication by an express method using laser Raman-luminescent spectroscopy. MATERIALS AND METHODS: Minced meat from beef and pork was used. Determination of quantity of mesophilic aerobic and opportunistic-anaerobic microorganisms was carried out by serial 10-fold dilutions with subsequent parallel seeding into Rida count total 24 plates and Petri dishes with 5% blood agar. Sample study was carried out in luminescent software-hardware complex Enspectr L405 (a variant of Enspectr M software-hardware complexes). RESULTS: Meat spoilage was established to be caused to a large degree by Pseudomonas genus (P. fluorescens, P. putida, P. fragi et al.) bacterial growth. Raman-luminescent spectra of bacteria that compose microflora, characterizing and accompanying beef and pork spoilage, were measured and recorded into a database. CONCLUSION: The results obtained will allow to use this technique in the future for both express-indication and differentiation of microorganisms and express-evaluation of quality of meat products at all stages of their manufacturing, storage, transport and realization.

[Identification of microorganisms based on the effect of giant Raman scattering].

AIM: Development and evaluation of express diagnostics of microorganisms for reduction of time spent on study execution. MATERIALS AND METHODS: Giant Raman scattering effect and proposed substrate with metallic nano-balls of silver were used for identification and indication of microorganisms. Staphylococcus aureus, Staphylococcus haemolyticus, Staphylococcus saprophyticus, Candida albicans, Escherichia coli, Bacillus subtillis, Pseudomonas aeruginosa were used as objects of study. All the microorganisms were cultivated in dense nutrient media and non-fluorescent microbiological membrane filters. RESULTS: Giant Raman scattering method on the substrates used increases sensitivity of indication and specificity of identification of microbes compared with other existing express methods and allows to see the signal from a single bacterium when exposed to laser light. The methodic proposed was also used to determine sensitivity of bacteria to antimicrobial preparations. CONCLUSION: The proposed hardware and method may be used for express diagnostics of processes of microbial nature.

[Influence of siRNA complexes on the reproduction of influenza A virus (Orthomyxoviridae: Alphainfluenzavirus) in vivo].

INTRODUCTION: Influenza is one of the most pressing global health problems. Despite the wide range of available anti-influenza drugs, the viral drug resistance is an increasing concern and requires the search for new approaches to overcome it. A promising solution is the development of drugs with action that is based on the inhibition of the activity of cellular genes through RNA interference. AIM: Evaluation in vivo of the preventive potential of miRNAs directed to the cellular genes FLT4, Nup98 and Nup205 against influenza infection. MATERIALS AND METHODS: The A/California/7/09 strain of influenza virus (H1N1) and BALB/c mice were used in the study. The administration of siRNA and experimental infection of animals were performed intranasally. The results of the experiment were analyzed using molecular genetic and virological methods. RESULTS: The use of siRNA complexes Nup98.1 and Nup205.1 led to a significant decrease in viral reproduction and concentration of viral RNA on the 3rd day after infection. When two siRNA complexes (Nup98.1 and Nup205.1) were administered simultaneously, a significant decrease in viral titer and concentration of viral RNA was also noted compared with the control groups. CONCLUSIONS: The use of siRNAs in vivo can lead to an antiviral effect when the activity of single or several cellular genes is suppressed. The results indicate that the use of siRNAs targeting the cellular genes whose expression products are involved in viral reproduction is one of the promising methods for the prevention and treatment of not only influenza, but also other respiratory infections.

[The prospects for the use of drugs based on the phenomenon of RNA interference against HIV infection].

The human immunodeficiency virus (HIV) is currently one of the most pressing global health problems. Since its discovery in 1978, HIV has claimed the lives of more than 35 million people, and the number of people infected today reaches 37 million. In the absence of highly active antiretroviral therapy (HAART), HIV infection is characterized by a steady decrease in the number of CD4+ T-lymphocytes, but its manifestations can affect the central nervous, cardiovascular, digestive, endocrine and genitourinary systems. At the same time, complications induced by representatives of pathogenic and opportunistic microflora, which can lead to the development of bacterial, fungal and viral concomitant infections, are of particular danger. It should be borne in mind that an important problem is the emergence of viruses resistant to standard therapy, as well as the toxicity of the drugs themselves for the body. In the context of this review, of particular interest is the assessment of the prospects for the creation and clinical use of drugs based on small interfering RNAs aimed at suppressing the reproduction of HIV, taking into account the experience of similar studies conducted earlier. RNA interference is a cascade of regulatory reactions in eukaryotic cells, which results in the degradation of foreign messenger RNA. The development of drugs based on the mechanism of RNA interference will overcome the problem of viral resistance. Along with this, this technology makes it possible to quickly respond to outbreaks of new viral diseases.

[20-year experience with laserofluorescent diagnostics in clinical microbiology].

Results of rapid laser-assisted identification of microorganisms for diagnostics of microbial processes based on auto-fluorescence effect in bacteria-containing materials are summarized. It is proposed to use the auto-fluorescence technique for express diagnostics of pyoinflammatory diseases, evaluation of microflora conditions (eubiosis, dysbiosis) and sensitivity to antibiotics, monitoring and prognostication, assessment of the quality of antibiotic therapy. Priority in the development of this medical technology for laserofluorescent diagnostics and its practical application is protected by 15 patents.

Potential of application of the RNA interference phenomenon in the treatment of new coronavirus infection COVID-19.

COVID-19 has killed more than 4 million people to date and is the most significant global health problem. The first recorded case of COVID-19 had been noted in Wuhan, China in December 2019, and already on March 11, 2020, World Health Organization declared a pandemic due to the rapid spread of this infection. In addition to the damage to the respiratory system, SARS-CoV-2 is capable of causing severe complications that can affect almost all organ systems. Due to the insufficient effectiveness of the COVID-19 therapy, there is an urgent need to develop effective specific medicines. Among the known approaches to the creation of antiviral drugs, a very promising direction is the development of drugs whose action is mediated by the mechanism of RNA interference (RNAi). A small interfering RNA (siRNA) molecule suppresses the expression of a target gene in this regulatory pathway. The phenomenon of RNAi makes it possible to quickly create a whole series of highly effective antiviral drugs, if the matrix RNA (mRNA) sequence of the target viral protein is known. This review examines the possibility of clinical application of siRNAs aimed at suppressing reproduction of the SARS-CoV-2, taking into account the experience of similar studies using SARS-CoV and MERS-CoV infection models. It is important to remember that the effectiveness of siRNA molecules targeting viral genes may decrease due to the formation of viral resistance. In this regard, the design of siRNAs targeting the cellular factors necessary for the reproduction of SARS-CoV-2 deserves special attention.

[Rapid identification of mycobacteria using laser fluorescence].

AIM: To develop rapid method of identification of mycobacteria based on laser fluorescence. MATERIALS AND METHODS: Characteristics of laser-induced fluorescence of 19 bacteria species, including 17 species of mycobacteria, were studied. Identification of microorganisms was performed using measurement of spectral-fluorescent characteristics. RESULTS: Library of spectral-fluorescent characteristics of mycobacteria in different concentrations ratios and associations was created, which formed the basis of database for identification of mycobacteria by laser-fluorescent method. Principles of diagnostic algorithm of indication and differentiation of mycobacteria using this method were developed. Effect of myramistin for increasing the intensity of mycobacteria fluorescence, account of the diffracting characteristics of medium for adjustment of spectral characteristics of mycobacteria and processing of data by factor analysis are needed. Efficacy of the method was 80 - 90%. CONCLUSION: Principles of rapid identification of mycobacteria and their associations developed on the basis of laser-fluorescent method are experimentally founded and tested on unknown cultures of mycobacteria and objectively prove the possibility to apply this method for express identification of mycobacteria belonging to M. tuberculosis complex as well as non-tuberculous mycobacteria.

[Impact of liposomes and ceftriaxone-entrapped liposomes on skin wound healing in rats].

"Empty" liposomes and ceftriaxone (CT)-entrapped liposomes were developed. Lipids of natural origin, i. e. soybean phosphatidyl choline (PhCh) and cholesterol were used for the design. The conditions for production of the CT-entrapped liposomes were optimized to provide the maximum content of the antibiotic in the liposomes. The effect of the "empty" liposomes and the antibiotic-entrapped liposomes on healing of skin wounds was studied on rats. The wound on the skin surface of the rat back was purulent due to contamination with E. coli (10(9) cells/ml). The treatment with the antibiotic solution, "empty" liposomes or CT-entrapped liposomes was started in 48 hours. The use of the PhCh "empty" liposomes promoted more rapid healing of the wound vs. the control or the treatment with CT aqueous solution. The treatmen of the wound with the CT-entrapped PhCh liposomes provided 2 times more rapid healing vs. the control or the use of the CT aqueous solution.

[The role of regulatory cells CD4+CD25+ in the development of chronic infective diseases].

Regulatory T-cells CD4+CD25+Foxp3 (Treg) present a small subpopulation of CD4+ T lymphocytes, which develop in the thymus and are disseminated into peripheral lymphoid organs on the 3rd or the 4th day of the neonatal period. Treg play a decisive role in the pathogenesis of autoimmune diseases and the development of tolerance to transplantation antigens, regulate the immune response to allergens, and suppress antimicrobial immunity. Treg suppress proliferation as well as the cytotoxic effect and the secretion of Th1 and Th2 cytokines by effectory T lymphocytes, thus limiting the strength of the immune response of effectory T-cells, which makes them impossible to adequately control viral and bacterial infections. Recognition via antigen-presenting cells and the subsequent induction of the proliferation of antigen-reactive T and B lymphocytes, directed towards infectious agent elimination, is accompanied by the activation of regulatory T cells as well, which leads to immune response suppression; repeated microbial infections are not only able to strengthen T-cell immunity by generating memory T-cells, but can also strengthen the suppressive activity of endogenous T regulators CD4+CD25+. Moreover, T reg are capable of the direct recognition of a microbial product; these cells selectively express Toll-like receptors (TLR)-2, -4, -5, -7, and -8. Under normal conditions T reg are anergic, but are capable of direct proliferation in response to stimulation by TLR ligands, expressed on microbes and parasites. Treg removal enforces protective immune response to contagious microorganisms, including bacteria, viruses, and fungi, which leads to the elimination of pathogens from the host organism. The removal of Treg population will help to accomplish infectious pathogen elimination and diminish inflammation within a short period of time.

[Current status of laboratory diagnosis of infections caused by nonspore-forming anaerobes and ways of its improvement]. / Sovremennoe sostoianie laboratornoi diagnostiki infektsii, vyzyvaemykh nesporoobrazuiushchim anaerobami, i puti eë sovershenstvovaniia]

Nonspore-anaerobic bacteria are the most common pathogens of purulent opportunistic infection. However, this group of infections is not recorded in our country due to the lack of national culture media and special laboratory equipment for anaerobic bacteriology. The laboratory diagnosis of infections caused by nonspore-forming anaerobes are analyzed and urgent measures to solve this topical problem are defined.

[Microbiocenotic changes in the large intestine of patients with various diseases]. / Izmeneniia mikrobiotsenoza tolstoi kishki u bol'nykh razlichnymi zabolevaniiami.

The specific features of the components of human large bowel microbiocenosis were studied in healthy persons and patients with various pyo-inflammatory and other diseases in relation to the physiological status. The presence of a pathological process was demonstrated to be a powerful factor of intestinal microbiocenotic variations. No specific changes in the large bowel microflora were found in the abnormalities in question.

[Infections caused by non spore-forming anaerobic bacteria]. / Sostoianie problemy infektsii, vyzyvaemykh nesporoobrazuiushchimi anaérobnymi bakteriiami.

Pyo-inflammatory diseases continue to be one of the topical problems of modern medicine. Their etiological agents are opportunistic microorganisms among which non spore-forming anaerobes are prevalent in the normal microscopic flora. They cause a pathological process when the body's immunity is decreased. The authors consider the biological features of this group of bacteria, epidemiological and pathogenetic aspects, the problems of laboratory and clinical diagnosis of non clostridial anaerobic infection and outline immediate steps to be taken to solve the problems.

[Laser fluorescence in assessing the hygienic status of oral cavity]. / Primenenie lazernoi fliuorestsentsii dlia otsenki gigienicheskogo sostoianiia polost rta.

The issue related with an objective evaluation of a hygienic status of the oral cavity and a method to solve it by modern laser technologies are addressed in the paper. The efficiency of bacteriological and of laser-fluorescence express-diagnostics is comparatively analyzed. A hygienic index and its clinical value are substantiated.

[Microecology of non spore-forming anaerobes in health and in disease]. / Mikroékologiia nesporoobrazuiushchikh anaérobov v norme i pri patologii.

The authors studied the species-specific and quantitative composition of the large intestinal microbiocenosis on exposures to various factors of endo- and exogenous etiologies: the presence of a pathological process and its specific features, dietary trace element composition, unfavorable environmental (chemical) factors, as well as the impact of coexistence in the same family and the factors of family variability (the individual genotype of the macro-organism). The microenvironment of non spore-forming anaerobes that colonize the large bowel was found to be influenced by a number of various factors, both exo- and endogenous. The magnitude of these changes is associated with the intensity and specificity of an influencing factor.

[Problem in the diagnosis of anaerobic infections and dysbacteriosis in clinical dentistry]. / Problema diagnostiki anaérobnoi infektsii i disbakterioza v klinicheskoi stomatologii.

The fluorescence technique was proposed for rapid diagnosis of anaerobic infection. It determines the microbiocenosis of a wound, the gastrointestinal tract, maxillofacial region, and other pathologically changed organs and tissues. The technique allows one to follow the time course of changes in the microflora and in the pathological process during the treatment and follow-up of a patient. An attempt was made to establish a rapid diagnosis of intestinal dysbacteriosis and to evaluate its severity. A real-time feed-back unit was developed for the diagnosis of anaerobic infection. An anaerobic microflora model in dental caries was used to work out criteria for the informative value of the proposed technique and it was clinically tested.

[Human microecology studies at Department of Microbiology with Virology and Immunology, I.M. Sechenov Moscow Medical Academy]. / Issledovanie po mikroékologii cheloveka na kafedre mikrobiologii s virusologiei i immunologiei.

Studies of human microflora in health and disease and during exposure to professional and ecological factors is a traditional problem solved for many years by staff members of Department of Microbiology with Virology and Immunology, I. M. Setchenov Moscow Medical Academy. The purpose of research is to develop methods and means for diagnosis and prevention of human microbiocenosis disorders. Fundamental and applied research in cooperation with prophylactic and clinical institutions and departments yielded data contributing to solution of many pressing problems in prevention and diagnosis of infectious diseases.

[The role of nonsporulating anaerobic bacteria in the etiology of postabortion and postpartal endometritis]. / Rol' nesporoobrazuiushchikh anaérobnykh bakterii v étiologii posleabortnogo i poslerodovogo éndometritis.

As the result of the study carried out by the authors, a conclusion has been made on the leading role of polymicrobial aerobic-anaerobic complexes, capable of producing a synergic effect, in the etiology of endometritis, anaerobic microorganisms prevailing in their importance. In the etiology of endometritis the most important organisms are bacteroids and peptostreptococci among anaerobes, enterobacteria and group D streptococci among aerobes. Anaerobic bacteria causing endometritis are most sensitive to dalacin and lincomycin, least sensitive to benzylpenicillin, resistant to aminoglycosides. The amounts of aerobic and anaerobic microflora contaminating the uterine cavity correlate with the severity of endometritis.

[Morphologic characteristics of bacteria treated with human blood serum and immunoglobulin]. / Morfologicheskaia kharakteristika bakterii, obrabotannykh syvorokoi krovi i immunoglobulinom cheloveka.

The electron microscopic study of ultrathin sections of bacterial cells treated with blood serum has revealed the presence of the immunoglobulin cover located on the cell wall and consisting of blood serum components. Staphylococci (strain Smith), studied with the use of ruthenium red, have been found to possess the capsule and the immunoglobulin cover located at its base. The immunoglobulin cover has proved to be more pronounced in gram-positive bacterial (actinomycetes, peptococci, staphylococci) and faintly pronounced in gram-negative bacteria (Bacteroides, Proteus, Pseudomonas, Escherichia).

[Changes in the ultrastructure of staphylococci in kidney tissue in experimental infection]. / Izmenenie ul'trastruktury stafilokokkov v pochechnoi tkani pri eksperimental'noi infektsii

A study of ultrathin sections fo the kidney tissue of mice infected with staphylococci showed intracellular localization of the bacteria. In vivo there occurred a morphological reconstruction of external layers of the cell wall of staphylococci, and lysis of individual staphylococci. The middle part of the cell wall and the cytoplasmic membrane proved to be the structures the most stable to lysis. Specific changes of bacteria similar to the changes noted with the action of penicillin on staphylococci in vitro followed effective penicillin therapy of mice infected with staphylococci.

[The intestinal microflora of patients with inflammatory diseases of the maxillofacial area]. / Mikroflora kishechnika u bol'nykh vospalitel'nymi zabolevaniiami cheliustno-litsevoi oblasti.

The microflora of the large intestine in patients with odontogenic phlegmons of different localization were studied. 80.8% of such patients were found to have microecological disturbances, characterized by a decrease in the number of bacteria belonging to the genera Bifidobacterium, Lactobacillus, Enterococcus and Bacteroides and by an increase in the number of opportunistic microorganisms.