Microflora of sputum and autopsy material of patients with COVID-19.

The rapid spread of a new coronavirus infection in the country actualizes the conduct of bacteriological studies of clinical material obtained from the respiratory tract of patients with COVID-19. During the experiments, 230 sputum samples and 260 autopsy lung samples from patients with COVID-19 were analyzed. 946 high-risk strains were isolated and identified by MALDI-ToF mass spectrometry on a Microflex LT instrument (Bruker®). According to the results of bacteriological cultures of sputum, a predominance of gram-positive ones was revealed, amounting to 50.5% (222 strains) of the total number of isolated pathogens. However, falling into this group is manifested by natural representatives of the microflora of the human mucous membranes from the genera Streptococcus, Rothia and Lactobacillus (109 strains in total), which can be manifested by the detection of improper sputum collection, causing contamination by the substance of intense salivation and nasopharyngeal discharge. In turn, the "classic" gram-positive causative agents of pneumonia were detected much less frequently: S. aureus in 5 cases, S. pneumoniae in 6 patients. The causative agents in the order Enterobacterales are represented by 42 strains, among which the most likely species are K.pneumoniae (27 strains). In the group of non-fermenting gram-negative bacteria, A. baumanii (29 strains) prevailed, and P. aeruginosa was also identified in 2 cases. When analyzing the results of a microbiological study of autopsy material (lungs) of patients with COVID-19, significant differences in the qualitative and quantitative composition of the microflora were revealed, compared with sputum. In the group of gram-positive bacteria, 15 strains of the natural microflora of the mucous membranes were identified, while sensitive species dominated among gram-negative pathogens: K. pneumoniae (102 strains), A. baumanii (75 strains), P. aeruginosa (11 strains). Regular microbiological monitoring is essential for antibiotic therapy and prevention of secondary bacterial infection. In the event of a fatal outcome, the results of microbiological analysis of autopsy material can determine the cause of death of the patient.

[Muscle post-effects and upright standing in healthy subjects and patients with sensory-motor integration disorders].

We compared the upright standing in 7 patients with sensory-motor disorders and 7 healthy subjects (control) before and after 30-s involuntary neck muscle contraction. A trajectory of the center of pressure was recorded during 30-s standing with the eyes open, eyes closed and standing on a foam-rubber with the eyes open. As compared to healthy subjects, patients exhibited an increased body sway area during standing with the eyes open on both the firm surface and foam-rubber and a backward shift of the center of pressure during standing with the eyes both open and closed. Closing the eyes affected the upright standing of patients to a lesser extent than standing of healthy subjects. Involuntary neck muscle contraction within 30 s elicited a backward shift of the center of pressure in healthy subjects, especially during standing with the eyes closed, and a decrease in the length of the center-of-pressure trajectory, especially of its frontal component during standing on the foam-rubber. In patients, a post-effect of the neck muscle contraction manifested itself as a decrease in the body sway area during standing on the foam-rubber and relative increase in the frontal component of the center-of-pressure trajectory during standing with the eyes closed. The results suggest that the upright standing of patients with sensory-motor disorders is more sensitive to somatosensory than visual input, and 30-s neck muscle contraction approach their postural stability to the age-matched control.

betaIV spectrin, a new spectrin localized at axon initial segments and nodes of ranvier in the central and peripheral nervous system.

We report the identification of betaIV spectrin, a novel spectrin isolated as an interactor of the receptor tyrosine phosphatase-like protein ICA512. The betaIV spectrin gene is located on human and mouse chromosomes 19q13.13 and 7b2, respectively. Alternative splicing of betaIV spectrin generates at least four distinct isoforms, numbered betaIVSigma1-betaIVSigma4 spectrin. The longest isoform (betaIVSigma1 spectrin) includes an actin-binding domain, followed by 17 spectrin repeats, a specific domain in which the amino acid sequence ERQES is repeated four times, several putative SH3-binding sites and a pleckstrin homology domain. betaIVSigma2 and betaIVSigma3 spectrin encompass the NH(2)- and COOH-terminal halves of betaIVSigma1 spectrin, respectively, while betaIVSigma4 spectrin lacks the ERQES and the pleckstrin homology domain. Northern blots revealed an abundant expression of betaIV spectrin transcripts in brain and pancreatic islets. By immunoblotting, betaIVSigma1 spectrin is recognized as a protein of 250 kD. Anti-betaIV spectrin antibodies also react with two additional isoforms of 160 and 140 kD. These isoforms differ from betaIVSigma1 spectrin in terms of their distribution on subcellular fractionation, detergent extractability, and phosphorylation. In islets, the immunoreactivity for betaIV spectrin is more prominent in alpha than in beta cells. In brain, betaIV spectrin is enriched in myelinated neurons, where it colocalizes with ankyrin(G) 480/270-kD at axon initial segments and nodes of Ranvier. Likewise, betaIV spectrin is concentrated at the nodes of Ranvier in the rat sciatic nerve. In the rat hippocampus, betaIVSigma1 spectrin is detectable from embryonic day 19, concomitantly with the appearance of immunoreactivity at the initial segments. Thus, we suggest that betaIVSigma1 spectrin interacts with ankyrin(G) 480/270-kD and participates in the clustering of voltage-gated Na(+) channels and cell-adhesion molecules at initial segments and nodes of Ranvier.

[Age-related structure of miners population in dependence on work conditions].

The authors analyzed age-related structure of miners population in major occupational groups with connection to special work conditions in one mine of Kouzbass. The data obtained prove certain influence of work conditions on age-related structure of occupational population.

[Microbial community of freshwater sponges in Lake Baikal].

The microbial community of Baikal sponges has been studied in five species belonging to the genera Swartschewskia, Baicalospongia, and Lubomirskia of the endemic family Lubomirskiidae. The results show that the total numbers of bacteria and bacterioplankton production have an effect on the growth of L. baicalensis body. Bacteria of the genera Pseudomonas, Bacillus, Micrococcus, Sarcina, Flavobacterium, Arthrobacter, and Acinetobacter living in the sponges are representatives of the Baikal bacterioplankton. Actinomycetes of the genera Streptomyces and Micromonospora are a permanent component of the cultivable sponge microbial community. The numbers and enzyme activities of heterotrophic, oligotrophic, and psychrophilic bacteria isolated from different sponge species and the surrounding water in autumn and in winter have been estimated.

The receptor tyrosine phosphatase-like protein ICA512 binds the PDZ domains of beta2-syntrophin and nNOS in pancreatic beta-cells.

Islet cell autoantigen (ICA) 512 of type I diabetes is a receptor tyrosine phosphatase-like protein associated with the secretory granules of neurons and endocrine cells including insulin-secreting beta-cells of the pancreas. Here we show that in a yeast two-hybrid assay its cytoplasmic domain binds beta2-syntrophin, a modular adapter which in muscle cells interacts with members of the dystrophin family including utrophin, as well as the signaling molecule neuronal nitric oxide synthase (nNOS). The cDNA isolated by two-hybrid screening corresponded to a novel beta2-syntrophin isoform with a predicted molecular mass of 28 kDa. This isoform included the PDZ domain, but not the C-terminal region, which in full-length beta2-syntrophin is responsible for binding dystrophin-related proteins. In vitro binding of the beta2-syntrophin PDZ domain to ICA512 required both ICA512's C-terminal region and an internal polypeptide preceding its tyrosine phosphatase-like domain. Immunomicroscopy and co-immunoprecipitations from insulinoma INS-1 cells confirmed the occurrence of ICA512-beta2-syntrophin complexes in vivo. ICA512 also interacted in vitro with the PDZ domain of nNOS and ICA512-nNOS complexes were co-immunoprecipitated from INS-1 cells. Finally, we show that INS-1 cells, like muscle cells, contain beta2-syntrophin-utrophin oligomers. Thus, we propose that ICA512, through beta2-syntrophin and nNOS, links secretory granules with the actin cytoskeleton and signaling pathways involving nitric oxide.

Expression of cloned genes of transgenic microorganisms introduced into man-made ecosystems.

Modeling of transgenic microorganism introduction into small man-made ecosystems can help forecast changes in expression of cloned genes under different conditions of existence. Introduction of the E. coli Z905/pPHL7 strain containing a plasmid with luminescent system genes of luminous bacteria led to changes in cell and colony morphology, reduction in metabolic activity of cells, and, as a result, a lower level of expression of cloned gene. A low concentration of nutrients has been shown to favor greatly the phenotypic change of cells of the recombinant strain. Expression of cloned genes changed due to: a lower concentration of plasmid DNA, a change in regulation of cloned genes, and a change in cells of biosynthesis of substrates needed for expression of luminescent genes. The conducted investigations can provide a basis for the use of marker transgenic microorganisms in closed ecosystems of different types. Grant numbers: 99-04-96017, 00-07-9011.

Effects of prolonged exposure of lettuce seeds to HZE particles on orbital stations.

In a study of the biological effects of cosmic HZE particles, lettuce (Lactuca sativa) seeds were flown on the orbital stations Salyut 6 and 7 for varying periods of time (from 40 to 457 days). The dependence of the biological damage on flight duration, physical parameters and the fact of passage of an HZE particle through the seed was estimated using the criterion of the frequency of aberrant cells. The arrangement of the flight biological container Biobloc made it possible to trace the location of tracks of individual HZE particles with Z > or = 6 and LET 200 keV/um. In seeds hit by HZE particles, for all exposure times, a statistically significant much higher yield of aberrant cells and also of cells containing multiple chromosome aberrations was observed than in the control material. The frequency of aberrant cells is markedly higher (by a factor of 1,5) in seeds hit than in non-hit ones. The changes of the yield of aberrant cells as a function of the absorbed dose (3.2-63.4 mGy) and the fluence (4.8-44.2 particles/cm2) are linear for the exposure duration ranging from 40 to 457 days.

Modelling of genetically engineered microorganisms introduction in closed artificial microcosms.

The possibility of introducing genetically engineered microorganisms (GEM) into simple biotic cycles of laboratory water microcosms was investigated. The survival of the recombinant strain Escherichia coli Z905 (Apr, Lux+) in microcosms depends on the type of model ecosystems. During the absence of algae blooming in the model ecosystem, the part of plasmid-containing cells E. coli decreased fast, and the structure of the plasmid was also modified. In conditions of algae blooming (Ankistrodesmus sp.) an almost total maintenance of plasmid-containing cells was observed in E. coli population. A mathematics model of GEM's behavior in water ecosystems with different level of complexity has been formulated. Mechanisms causing the difference in luminescent exhibition of different species are discussed, and attempts are made to forecast the GEM's behavior in water ecosystems.

Population dynamics of transgenic microorganisms in the different microecosystem conditions.

The role of key environmental factors in adaptation of spore-forming and non-spore-forming transgenic microorganisms (TM) have been studied in model ecosystems. Model TM Escherichia coli Z905 (bearing plasmid genes of bacterial luminescence Ap (r) Lux+) has been found to have a higher adaptation potential than TM Bacillus subtilis 2335/105 (bearing genes of human alpha 2-interferon Km (r) Inf+), planned for employment as a living vaccine under varying environmental conditions. Effects of abiotic factors on migration of natural and recombinant plasmids between microorganisms under model ecosystem conditions has been estimated. The transgenic microorganisms with low copy number survived better under introduction conditions in the microcosms studied. This trend has been shown to be independent of the microcosm type and its complexity. Grant numbers: 99-04-96017, 25, 00-07-9011.

[Synthesis of angiotensin converting enzyme inhibitors containing residues of substituted quinolines and a study of their biological activity]. / Sintez ingibitorov angiotenzinprevrashchaiushchego fermenta, soderzhashch ostatki zameshchennykh khinolinov, i issledovanie ikh biologicheskoi aktivnosti.

Inhibitors of the angiotensin-converting enzyme were synthesized by substituting N-and C-terminal amino acid residues of tripeptide Bz-Phe-Ala-Pro by the residues of 8-methoxy-5-sulphoquinoline and carboxy-1,2,3,4-tetrahydroquinoline, respectively, and their in vivo and in vitro biological activity was determined. The enzyme's S2' site proved to be non specific to the position of the carboxylic group in the C-terminal heterocyclic part of the inhibitor molecule. Introducing a modified quinoline residue into the N-terminal part of the inhibitor does not increase its specific interaction with the hydrophobic pocket of the angiotensin-converting enzyme.

[Silver staining of the nucleoli of pig embryonic kidney cells during the hyperactivation and inhibition of the synthesis of nucleolar RNA by UV microirradiation]. / Okrashivanie serebrom iadryshek kletok SPEV pri giperaktivatsii i ingibirovanii sinteza iadryshkovoi RNK u UF-mikrooblucheniem.

Silver staining of the nucleoli in pig embryo kidney cells (PK) was studied during the cell cycle and also upon mature nucleoli modifications induced by UV microirradiation. During anaphase only four silver-stained granules were revealed in each daughter set of chromosomes in the four nucleolus-organizing regions (NORs). In the following 1-2 hours, the number of granules in the NORs rapidly increased up to 25-30 per nucleus. During the next 20-25 hours of the cell cycle, the number of silver-stained granules was slowly doubling as the nucleoli grew in size. UV microirradiation of one nucleolus in the nucleus with two nucleoli induced a profound degradation of the injured nucleolus and a compensatory hypertrophy of the intact one. Such nucleolar modifications were accompanied by redistribution of the silver-stained granules between the injured and non-injured nucleoli and by alterations in the levels of nucleolar RNA synthesis in the NORs. These data support a hypothesis that silver-stained proteins may be involved in the regulation of the nucleolar activity.

[Laminar analysis of evoked potentials of the cat visual cortex during prenatal ontogenesis]. / Posloinyi analiz vyzvannykh potentsialov zritel'noi kory koshki v prenatal'nom ontogeneze

A laminar analysis of visual evoked potentials recorded from the cortical striate zone in response to stimulation of the optic nerve was made on the cats foeti in the second half of antenatal development and on kittens in the first few days of their life. In all the investigated terms of prenatal ontogenesis, two negativity foci are distinctly revealed in the specific zones of the visual cortex. The first appears with the least latency in the middle cortical layers after an afferent volley inflow to the cortex. The second, which reaches a maximum in 40 to 50 msec appears in the superficial cortical structures. Analysis of the data obtained shows that the first excitation focus is due to the summation of EPSP in the neural elements of the middle cortical layers, while the second is caused by development of depolarization in the apical dendrites of the pyramidal neurones.

[Functional maturation of the cat visual cortex during prenatal ontogenesis]. / Funktsional'noe sozrevanie zritel'noi kory koshki v prenatal'nom ontogeneze

Development of the perceiving function of the visual cortex was studied on the foeti of cats in the second stage of antenatal development with intact placental blood circulation. It has been found that functional maturing of the cortical end of the visual analyser starts at the beginning of the second half of antenatal life. At this period EPs to stimulation of the optic nerve are recorded throughout the dorsal cortex of the contralateral hemisphere. At first they appear as slow three-phase (positive-negative-positive) oscillations of a small amplitude. As the foetus develops, the EP amplitude increases, and the EP configuration in the striate zone of the cortex becomes complex. Two weaks before birth, a short-latency negative wave appears against the background of the primary positive oscillation. In the last week of antenatal development of the foeti and in the first few days of the kittens life, EPs are represented in the specific zone of the visual cortex (g. lateralis) as two negative oscillations, and in the so-called associative zone (the middle part of the suprasylvian gyrus) by one long-latency high-amplitude negative oscillation which corresponds by latency to the second negative EP component in the striate cortex.

[Role of several subcortical nuclei in the conduction of visual afferentation during early ontogeny]. / O roli nekotorykh podkorkovykh iader v provedenii zritel'noi afferentatsii v rannem ontogeneze.

Characteristics of EPs in different areas of the visual cortex elicited by stimulation of subcortical nuclei (LGB, posterior thalamic nuclei and superior colliculi) were studied in cat's fetuses during the last 10 days of their antenatal development. It was shown that both in the striate and in accessory visual areas the EPs are negative oscillations with a small initial positivity. Latencies of the EPs to stimulation of the thalamic nuclei make from 18 to 25 msec in different cortical areas and those to the stimulation of the superior colliculi--from 40 to 45 msec. Laminar analysis of EPs elicited in the occipital cortical area by the stimulation of LGB revealed two foci of depolarization; the one with shorter EP latency is localized in the depth of the cortex. The role of the extrageniculate ascending system and of different groups of fibers of the geniculocortical system in the formation of the main EP components in the visual cortex in early ontogenesis is discussed.

[Theoretical analysis of the continuous two-stage process of the biosynthesis of a metabolic product]. / Teoreticheskii analiz nepreryvnogo dvukhstadiinogo protsessa biosinteza produkta metabolizma.

A mathematical model for continuous biosynthesis of a metabolite in a battery of two apparatuses with ideal agitation is described and analysed. In the first apparatus of the battery it is advisable to maintain a high specific rate of culture growth for continuous accumulation of young active biomass while in the second apparatus a low growth rate is expedient which provides a change in the culture metabolism to biosynthesis of the required product. To make the continuous two-stage process efficient, it is necessary to add an extremely concentrated solution of the nutrients to the second apparatus of the battery. Influence of the oxygen transport velocity on the maximum attainable concentration of the biomass and the process capacity by the required product was studied.

[Introduction and long-term storage of recombinant luminescent Escherichia coli strain Z905 in laboratory water microecosystems]. / Introduktsiia i dlitel'noe sokhranenie rekombinantnogo liuminestsentnogo shtamma Escherichia coli Z905 v laboratornykh vodnykh mikroékosistemakh.

We studied preservation of recombinant Escherichia coli strain Z905 (AprLux+) in liquid microecosystems (LME) after the introduction. E. coli cells were shown to remain viable and preserve the ability to express the cloned lux genes for a long time (more than a year) in LME. The majority of the clones have reduced efficiency of the expression due to either changed regulation of the lux operon or decreased number of copies of the plasmid. These mechanisms could be realized either independently or simultaneously depending on LME conditions. We have exposed the major factors affecting the metabolic activity of the E. coli strain Z905 (AprLux+) introduced into model ecosystems and the level of expression of the cloned genes.

[Optimization of oxytetracycline biosynthesis]. / Optimizatsiia biosinteza oksitetratsiklina.

It was shown that rising of temperature up to 30 degrees C at the stage of the oxytetracycline-producing organism growth promoted acceleration of the culture growth rate and increasing of the antibiotic concentration by the 114th hour of the biosynthetic process. For the apparatus used in the study optimal aeration and agitation conditions were developed. To provide optimal parameters during biosynthesis of oxytetracycline, it was recommended to use the aeration rate of 1 v/v.min and the specific mechanical power for mixing of not less than 1 kW/m3.

[The comparative growth characteristics of variants of a recombinant Escherichia coli strain and of the representatives of aboriginal microflora in aqueous microecosystems under selective and nonselective conditions]. / Sravnitel'naia kharakteristika rosta variantov rekombinantnogo shtamma Escherichia coli i prestavitelei aborigennoi mikroflory vodnykh mikroékosistem v selektivnykh i neselektivnykh usloviiakh.

We have studied the kinetic characteristics of several isolates of Escherichia coli Z905 recombinant strain after introduction of the strain into model aquatic ecosystems. Most E. coli Z905 isolates grown as batch cultures under selective conditions (0.5 microgram/ml ampicillin) showed better kinetic characteristics of growth than did related species of native microflora, which originally populated laboratory microcosms.

[A synthetic fragment of insulin with insulin-like biological activity]. / Sinteticheskii fragment insulina s insulinopodobnoi biologicheskoi aktivnost'iu.

We have used computer modeling of insulin 3-D structure and experimental data about action of site point mutation on insulin activity to design functionally important domain with signaling activity and synthesized peptide than might be sufficient for the binding to insulin receptor. The designed and synthesized peptide consist of ten residues and may be obtained in two forms: oxidized and reduced (with or without disulfide bond). The synthesized decapeptide peptide represents functionally important site for binding to the insulin receptor. Amino acid residues at position 1-8 correlate with B-chain of insulin at position (B19-B26). Residues at position 9.10 correlate with A-chain at position A-10-A21. This peptide was tested with cell culture L-929 (glucose uptake) in comparison with bioactive commercial peptide (R-G-FF) and insulin. It was shown that synthesized peptide exhibit biological activity at molar concentration 0.01-1 mkM. Our results successfully demonstrate the synthetic insulin fragment have insulin-like biological activity.