Microorganisms associated with bacterial vaginosis: diversity and clinical and diagnostic significance

Bacterial vaginosis (BV) is the most common infectious non-inflammatory disease in women of reproductive age. The key feature of BV is that in the  absence of a specific pathogen, the disease develops against the background of vaginal biotope dysbiosis. According to the opinion of some authors, initially, the main role of BV in the pathogenesis was assigned to the species G. vaginalis. However, using of molecular methods made it possible to significantly expand the range of microorganisms found in women with BV, and to identify Gardnerella vaginalis, Atopobium vaginae, Mobiluncus spp., Prevotella spp. as the primary causative agents of BV. A number of studies have confirmed the sexual transmission of BV pathogens, with a new sexual partner being significant risk factors for episodic BV, and sexual contact with the same partner without using barrier methods of contraception for recurrent BV. At the same time, BV-associated bacterias  rarely exist as planktonic forms of one species, more often, they thrive in complex polymicrobial communities surrounded by an extracellular matrix, the so-called biofilms. In patients with BV, biofilms are detected in 90% of cases. G. vaginalis and Prevotella bivia are widespread in women with BV and, being the primary colonizers, create a bacterial biofilm, to which secondary colonizers can subsequently join, including A. vaginae, Sneathia spp. and potentially other BVABs. The resistance of bacterias in the biofilm to antimicrobial drugs is 1000 times higher than of planktonic forms, which can lead to chronicity of the infectious process and torpid course of the disease.The aim of this review was analyzing of modern studies on the prevalence, characteristics of opportunistic microorganisms associated with BV, to present their clinical and diagnostic significance and role in the pathogenesis of  diseases

Adaptive Redox Reactions Promote Naturalization of Rare Orchid Epipactis atrorubens on Serpentine Dumps Post Asbestos Mining

Epipactis atrorubens (Hoffm.) Besser. is a regionally rare orchid species with highly ornamental properties due to its very beautiful bright flowers, therefore it is of considerable interest as a horticultural plant for use in botanical gardens and greenhouses. The objective of the research was to assess metal accumulation and some pro-and antioxidant reactions in E. atrorubens, colonizing serpentine dumps post asbestos mining. Additionally, some physicochemical properties of substrates, microbiotic characteristics and water status were investigated in orchids growing on two serpentine dumps and in a natural forest habitat of the Middle Urals, Russia. The dump substrates were characterized by the strong stoniness and the high content of Mg, Ni, Cr and Co (by 1.8 times on average) compared to the natural habitat. In these sites, E. atrorubens was characterized by increased mycorrhization. In the rhizome and roots of E. atrorubens the concentrations of most metals studied were considerably higher (more than 4 times on average) than in the leaves. It was found that orchids colonizing serpentine dumps produced more lipid peroxidation products (by 1.4 times on average) in the leaves which was accompanied by the more active synthesis of such non-enzymatic antioxidants as ascorbate, free proline, soluble phenolic compounds (including flavonoids) and non-protein thiols. The study suggests that non-enzymatic antioxidants increased the adaptive potential of E. atrorubens and contributed to its naturalization on serpentine dumps post asbestos mining. © 2022 by the authors. Licensee MDPI, Basel, Switzerland.20-44-660011; Russian Foundation for Basic Research, РФФИ; Ministry of Education and Science of the Russian Federation, Minobrnauka; Ural Federal University, UrFU: FEUZ-2020-0057Funding: The reported study was partly funded by RFBR and the Government of Sverdlovsk region, project number 20-44-660011 and the Ministry of Science and Higher Education of the Russian Federation as part of state task of the Ural Federal University, FEUZ-2020-0057

Identification of Infectious Diseases Patterns in the Combined Use of Bacteriological Diagnostics and MALDI Biotyper

In multidisciplinary hospitals, there are conditions conducive to the emergence of healthcare-associated infections: high concentration of people with reduced immunity in a limited area, the presence of a significant number of sources of contagion (patients and carriers), a change in the biocenosis of the mucous membranes and skin of patients and medical personnel under the influence of widespread use of antibiotics and cytostatics. The aim of the research was in the intercomparison of the standardized bacteriological algorithms and the MALDI Biotyper system in the microbiological diagnosis of pathogens as illustrated by the healthcare-associated diseases.Materials and methods. Seventy-eight patients of a multidisciplinary hospital of a regional level (Irkutsk) in 2018–2019 were examined. The age of patients ranged from 1 to 15 years. The material for the study was blood, sputum, swabs from the tracheobronchial tree, throat, nose, wound, abdominal fluid, cerebrospinal fluid, and swabs from environmental objects. Identification of the isolated cultures (78 bacterial strains) was  carried out using generally accepted bacteriological methods, as well as using the MALDI Biotyper system.Results and discussions. In the structure of healthcare-associated infections, Pseudomonas aeruginosa occupied a leading position. Not all isolates of microorganisms were identified by standardized bacteriological methods. The identification of strains with characteristic manifestations of physiological and biochemical characteristics was more reliable. Identification difficulties arose in the presence of atypical properties of microorganisms when the use of MALDI Biotyper would be crucial.Conclusion. It is necessary to apply an integrated approach to conduct reliable diagnostics of pathogens. It includes standardized bacteriological methods and methods for identifying microorganisms using mass spectrometry in the subsequent stages

The effectiveness of biofilm formation of daily cultures of clinically significant strains of opportunistic bacteria

Background. The formation of biofilm structures by microorganisms living in the hospital environment is a serious medical problem. To conduct correct experimental studies, it is necessary to know the speed and efficiency of biofilm formation by clinically significant species of opportunistic bacteria.   Aim: to study the kinetics of plankton culture growth and the rate of biofilm formation by clinically significant pathogens of infections associated with medical care to substantiate the methodology of further research.   Materials and methods. The strains from the working collection of the Laboratory for Microbiome and Microecology of the Scientific Сentre for Family Health and Human Reproduction Problems were used. Experiments were carried out with conditionally pathogenic microorganisms of the Enterobacteriaceae family and non-fermenting gram-negative bacteria. The optical density was measured, the total microbial number of the cell suspension was determined, and the morphological structure of the biofilm was evaluated using a light microscope on sterile cover glasses for thespecies Pseudomonas aeruginosa, Klebsiella pneumoniae and Serratia marcescens.   Results. The duration of the lag phase of the kinetic curve of cell growth varied in isolates of S. marcescens, P. aeruginosa and K. pneumoniae from 1 to 4 and 6 hours of cultivation, respectively. Despite this, the exponential growth phase was the same for all tested isolates and amounted to 4 hours. Thus, isolates of clinically significant species entered the stationary growth phase after 5–10 hours of cultivation and were characterized as fast-growing. On abiotic surfaces, after 8 hours of incubation of the tested cultures, the initial stages of the formation of biofilm structures were observed, after 20 hours the formed multilayer biofilm was visualized, after 24 hours succession occurred, new single cells were attached to the place of the detached structures.   Conclusion. The data obtained on the duration of the main stages of growth kinetics compared with the visualization of the formation of biofilm structures on abiotic surfaces should be taken into account when studying the effects of disinfectants, antiseptics and antibacterial drugs on planktonic cells and biofilm associations of clinically significant opportunistic microorganisms

Frequency of strains with multiple antibiotic resistance in the structure of opportunitistic pathogens

Background. The problem of antibiotic resistance has remained significant for the medical community for more than half a century, since the first cases of resistance to penicillin were registered.   The aim. Analysis of the long-term dynamics of changes in the antibacterial resistance of microorganisms and the creation of a collection of multi-resistant strains of opportunistic microorganisms.   Materials and methods. The study included data from 3173 bacteriological samples of various loci of the human body for 2010 and 2020–2021. The sensitivity of isolated cultures was determined by the disk diffusion method to antimicrobial drugs of the following groups: penicillins, cephalosporins, fluoroquinolones, aminoglycosides, carbapenems, tetracyclines, macrolides, lincosamides, oxazolidinones, glycopeptides and others.   Results. In the general structure of conditionally pathogenic microorganisms, a significant increase in the frequency of isolation of multidrug-resistant representatives of the genus Staphylococcus by two or more times was observed in 2021 compared to 2010 and 2020. We also observed a significant increase in the proportion of multidrug-resistant Streptococcus spp. and non-fermenting gram-negative bacteria. These changes marked the beginning of the creation of a collection of conditionally pathogenic microorganisms with multiple antibacterial resistance. In the structure of multiresistant microorganisms included in the “Collection of human microbiota of the Irkutsk region”, the leading positions belong to Klebsiella pneumoniae (23.81 %), Escherichia coli (19.05 %) and Staphylococcus aureus (22.22 %).   Conclusion. Antibiotic resistance monitoring is an important measure to control the resistance of community-acquired and nosocomial (nosocomial) microorganisms both within a particular country and globally

Features of the Composition of the Colon Microbiota in Children of the First Year of Life with Functional Gastrointestinal Disorders

Background. The development of functional gastrointestinal disorders (FGID) in children, especially in the first year of life, is a key factor in the formation of an altered colon microbiota and its formation in older age. The article presents both clinical-detailed anamnesis, symptoms of FGID, and bacteriological characteristics – qualitative and quantitative composition of the colon microbiota in children of the first year of life.Aim: to determine the features of the colon microbiota in children of the first year of life with FGID.Materials and methods. The objects of the study were children of the first year of life (n = 28) with a diagnosis of FGID established by a gastroenterologist. The material was divided into two comparison groups, depending on the age of the subjects: the first group included stool sample cultures of children aged from birth to 6 months (n = 17), the second – from 6 to 12 months (n = 11). The bacteriological study of the quantitative and qualitative composition of the contents of the colon was performed using standard methods.Results. According to the anamnesis, colic and flatulence were the key symptoms of FGID (p < 0.05). The results show that the formation of FGID in children from birth to 6 months and from 6 to 12 months may be associated with altered colon microbiota. There was a decrease in the quantitative indicators of the indigenous microbiota: bifidobacteria, E. coli with normal sensitivity and Enterococcus spp., and an increase in the degree of contamination of Klebsiella spp., S. aureus, Clostridium spp.Conclusion. It is shown that the determining factor in reducing the risk of developing FGID in children of the first year of life is a high concentration and diversity of indigenous microbiota

Effects of аntimicrobials on <i>Pseudomonas aeruginosa</i> biofilm formation

Pseudomonas aeruginosa is one of the most problematic pathogens in medical institutions, which may be due to the ability of this microorganism to exist in a biofilm, which increases its resistance to antimicrobials, as well as its prevalence and survival ability in the external environment. This work aimed to evaluate the antimicrobial susceptibility of P. aeruginosa strains in planktonic and biofilm forms. We studied 20 strains of P. aeruginosa collected during 2018–2021 by specialists from the Laboratory of Microbiome and Microecology of the Scientific Centre for Family Health and Human Reproduction Problems. The identification of strains was carried out using test systems for differentiating gram-negative non-fermenting bacteria (NEFERMtest 24 Erba Lachema s.r.o., Czech Republic), and confirmed by mass spectrometric analysis and 16S rRNA gene sequencing. Antimicrobial activity was assessed by the degree of inhibition of cell growth in planktonic and biofilm forms (on a flat-bottomed 96-well plastic immunological plate). All clinical isolates of P. aeruginosa were biofilm formers, 47.6 % of the isolates were weak biofilm formers, and 52.4 % of the isolates were moderate biofilm formers. Planktonic cells and the forming biofilm of the tested P. aeruginosa strains were carbapenems-resistant. Biofilm formation was suppressed in more than 90 % of cases by the agents of the cephalosporin and aminoglycoside groups. Antimicrobial susceptibility of P. aeruginosa strains in the formed biofilm was significantly lower (p &lt; 0.05). Carbapenems and cephalosporins did not affect the mature biofilms of the tested P. aeruginosa strains in more than 60 % of cases. Only non-beta-lactam antibiotics (ciprofloxacin and amikacin) suppressed the growth of planktonic cells and destroyed the mature biofilm. The revealed differences in the effect of the tested antimicrobials on the P. aeruginosa strains biofilms correlate with resistance to a number of antibiotics. To prevent biofilm formation in the hospital strains of P. aeruginosa, the use of ceftazidime may be recommended, and antimicrobials such as ciprofloxacin and amikacin may be used to affect mature biofilms of P. aeruginosa

Probiotic consortiums: Structure and antagonistic activity against opportunistic bacteria and human normobiota (using the example of <i>Escherichia coli</i>) <i>in vitro</i>

Background. Using probiotic preparations based on consortia of microorganisms not only helps to restore the balance of the intestinal microbiota, but also increases the therapeutic effect of probiotics. Promising sources for obtaining probiotic consortia are milk products that have undergone natural fermentation with the help of spontaneously formed microbial consortia. The aim. To study the structure of five microbial consortia with probiotic properties from naturally fermented milk products and to assess in vitro their antagonistic activity against opportunistic bacteria and a representative of the human normobiota – Escherichia coli. Materials and methods. The structure of bacterial consortia was analyzed by sequencing methods. The antagonistic activity of the consortia was assessed by the disk diffusion method. Results. It has been established that the studied microbial consortiums are represented by Enterococcus spp. and Streptococcus spp. bacteria. In consortiums No. 1, No.  2, and No.  3, Enterococcus bacteria dominated, while in consortiums No.  4 and No. 5, Streptococcus dominated. Antagonistic activity was shown against four isolates of opportunistic bacteria: Klebsiella pneumoniae No.  493, Enterobacter hormaechei No. 372, Staphylococcus aureus No. 4 and Pseudomonas aeruginosa No. 25 IMB, as well as against one representative of the human normobiota – Escherichia coli No. 495. The highest growth delay zone is found in E. coli No. 495 isolate. Three test cultures (K. pneumoniae No. 509, E. coli ATCC25922 and P. aeruginosa No. 3 IMB) exhibited more dense growth around probiotic consortia. Conclusion. The results of the study showed that the effect of probiotic consortia differing in the composition of microorganisms can be neutral and bactericidal. The presence of antagonistic activity in the studied microbial consortia against multiresistant isolates of opportunistic bacteria is a prospect for creating probiotics with antibacterial properties

СЛУЧАИ ДИФФУЗНОЙ В-КРУПНОКЛЕТОЧНОЙ ЛИМФОМЫ С ФУНКЦИОНАЛЬНО ЗНАЧИМЫМИ ИНТРОННЫМИ МУТАЦИЯМИ В ГЕНЕ ТР53

Background. the presence of functionally significant intron mutations in the TP53 gene was demonstrated in experiments in vitro and on samples of patients with some variants of non-Hodgkin’s lymphomas. to date, the studies of TP53 in tumor tissue of patients with diffuse large B-cell lymphoma (dlBCl) have been focused on the mutations in 5–8 exons of the gene. For these reasons, further studies of the spectrum of changes in the intron sequences of the TP53 gene and the determination of their functional effect in dlBCl are required.Description. We present two case reports that are of interest as the cases for detection of functionally significant intron TP53 mutations in dlBCl tissue, first published in the Russian scientific literature. the first clinical case of dlBCl was clinically characterized by extranodal tumor at the onset of the disease, severe symptoms of tumor intoxication, high paraclinical tumor activity, and early hemoblastosis recurrence. the second case of dlBCl was characterized by initially massive tumor lesion, rapidly progressive course of the disease, severe symptoms of tumor intoxication, high paraclinical tumor activity and poor response to therapy with subsequent generalization of lymphoma. Conclusion. Mutations in the TP53 gene are the driver of the tumor process, serve not only as a marker of aggressive tumor progression, but also as an independent predictor of reduced sensitivity to treatment. the presented clinical cases show that an in-depth analysis of the results of the TP53 sequencing in tumors and functional assessment of all detected changes, including changes in the introns of the gene and involving in silico analysis techniques, are necessary.  Актуальность. наличие в гене ТР53 функционально значимых интронных изменений было продемонстрировано в экспериментах in vitro и на выборках больных отдельными вариантами неходжкинских лимфом. До настоящего времени при изучении ТР53 в опухолевой ткани больных диффузной В-крупноклеточной лимфомой (ДВккл) исследования были сосредоточены на поиске мутаций в 5–8 экзонах гена. По этим причинам требуются дальнейшие исследования спектра изменений в интронных последовательностях гена ТР53 и определение их функционального эффекта при ДВккл. Описание. В статье представлены два клинических наблюдения, которые представляют интерес как впервые  публикуемые в российской научной литературе случаи выявления функционально значимых интронных мутаций ТР53 в опухолевой ткани ДВккл. Первый клинический случай ДВккл характеризовался  экстранодальным опухолевым поражением в дебюте заболевания, выраженными симптомами опухолевой интоксикации, высокой параклинической активностью опухоли, развитием раннего рецидива гемобластоза с экстранодальным очагом поражения в забарьерной ткани, резистентным к лечению. Второй описанный клинический случай ДВккл характеризовался исходно массивным опухолевым поражением, быстропрогрессирующим течением заболевания, выраженными симптомами опухолевой интоксикации, высокой параклинической активностью опухоли и плохим ответом на терапию с последующей генерализацией лимфомы. Заключение. Мутации в гене ТР53 являются драйвером опухолевого процесса, служат не только маркером агрессивного течения опухоли, но и независимым предиктором снижения чувствительности к лечению. Представленные клинические случаи показывают, что необходимы углубленный анализ результатов  секвенирования ТР53 при опухолях и функциональная оценка всех выявляемых изменений, в том числе в интронах гена и с привлечением методов анализа in silico. 

Связь микро-РНК кластера miR-143/145 с онкогенезом: тканевой и клеточный контекст

The purpose of the study was to present up-to-date data on the regulation of expression, function in normal tissues and multidirectional activity in the oncogenesis of miR-143/145 microRNAs cluster, as well as to evaluate the possibilities and limitations of the therapeutic use of microRNAs of this cluster in malignant neoplasms. Material and methods. The search for available domestic and foreign literary sources published in PubMed and RSCI databases over the past 10 years has been carried out. 427 articles were found, of which 41 were included in this review. Results. The conservative cluster miR-143/145 is one of the most intensively studied in tumors. Based on the results of the analysis of differential miRNA expression, in vitro experiments in cancer cell lines and in vivo in mouse tumor models, a decrease in miR-143 and miR-145 levels was shown in malignant neoplasms of epithelial origin. Until recently, these miRNAs were considered classical oncosuppressors. The data presented in the review demonstrate that the results of a number of studies taking into account the cellular aspects of microRNA expression contradict this concept. miR-143 microRNA, for example, is known to participate in the metabolic restructuring of the tumor and the activation of neoangiogenesis. It has been shown that the oncosuppressive or pro-oncogenic activity of miR-143 and miR-145 depend on the tissue and cellular context and can be explained by the presence of several regulated targets that have opposite effects on oncogenesis. Taken together, the data obtained suggest the need to exercise caution when choosing the microRNAs of the described cluster for exogenous therapeutic delivery. Conclusion. Further detailed decoding of the mechanisms of miR-143 and miR-145 functioning in various types of tissues and cells, as well as identification of new MRNA targets are necessary for a better understanding of the involvement of these molecules in oncogenesis.Цель исследования - представить современные данные о регуляции экспрессии, функции в нормальных тканях и разнонаправленной активности в онкогенезе микроРНК кластера miR-143/145, а также оценить возможности и ограничения терапевтического использования микроРНК данного кластера при злокачественных новообразованиях. Материал и методы. Проведен поиск доступных отечественных и зарубежных литературных источников, опубликованных в базах данных PubMed и РИНЦ за последние 10 лет. Найдено 427 статей, из которых 41 была включена в данный обзор. Результаты. Консервативный кластер miR-143/145 является одним из наиболее интенсивно изучаемых при опухолях. На основании результатов анализа дифференциальной экспрессии микроРНК, экспериментов in vitro в раковых клеточных линиях и in vivo в мышиных моделях опухолей было показано снижение уровня miR-143 и miR-145 при злокачественных новообразованиях эпителиального происхождения. До недавнего времени данные микроРНК считались классическими онкосупрессорами. Приведенные в обзоре данные демонстрируют, что результаты целого ряда работ, учитывающих клеточные аспекты экспрессии микроРНК, противоречат этой концепции. Для микроРНК miR-143, например, известно участие в метаболической перестройке опухоли и активации неоангиогенеза. Показано, что онкосупрессорная или проонкогенная активность miR-143 и miR-145 зависят от тканевого и клеточного контекста и могут объясняться наличием у них нескольких регулируемых мишеней, оказывающих противоположные эффекты на онкогенез. В совокупности полученные данные говорят о необходимости проявлять осторожность при выборе микроРНК описываемого кластера для экзогенной терапевтической доставки. Заключение. Дальнейшая детальная расшифровка механизмов функционирования miR-143 и miR-145 в различных типах тканей и клеток, а также идентификация новых мРНК-мишеней необходимы для лучшего понимания участия данных молекул в онкогенезе