An improved and extended dual-index multiplexed 16S rRNA sequencing for the Illumina HiSeq and MiSeq platform.

BACKGROUND: Recent advancements in next-generation sequencing (NGS) technology have ushered in significant improvements in sequencing speed and data throughput, thereby enabling the simultaneous analysis of a greater number of samples within a single sequencing run. This technology has proven particularly valuable in the context of microbial community profiling, offering a powerful tool for characterizing the microbial composition at the species level within a given sample. This profiling process typically involves the sequencing of 16S ribosomal RNA (rRNA) gene fragments. By scaling up the analysis to accommodate a substantial number of samples, sometimes as many as 2,000, it becomes possible to achieve cost-efficiency and minimize the introduction of potential batch effects. Our study was designed with the primary objective of devising an approach capable of facilitating the comprehensive analysis of 1,711 samples sourced from diverse origins, including oropharyngeal swabs, mouth cavity swabs, dental swabs, and human fecal samples. This analysis was based on data obtained from 16S rRNA metagenomic sequencing conducted on the Illumina MiSeq and HiSeq sequencing platforms. RESULTS: We have designed a custom set of 10-base pair indices specifically tailored for the preparation of libraries from amplicons derived from the V3-V4 region of the 16S rRNA gene. These indices are instrumental in the analysis of the microbial composition in clinical samples through sequencing on the Illumina MiSeq and HiSeq platforms. The utilization of our custom index set enables the consolidation of a significant number of libraries, enabling the efficient sequencing of these libraries in a single run. CONCLUSIONS: The unique array of 10-base pair indices that we have developed, in conjunction with our sequencing methodology, will prove highly valuable to laboratories engaged in sequencing on Illumina platforms or utilizing Illumina-compatible kits.

[Composition of oropharyngeal microbiota in patients with COVID-19 of different pneumonia severity].

AIM: To identify features of the taxonomic composition of the oropharyngeal microbiota of COVID-19 patients with different disease severity. MATERIALS AND METHODS: The study group included 156 patients hospitalized with confirmed diagnosis of COVID-19 in the clinical medical center of Yevdokimov Moscow State University of Medicine and Dentistry between April and June 2021. There were 77 patients with mild pneumonia according to CT (CT1) and 79 patients with moderate to severe pneumonia (CT2 and CT3). Oropharyngeal swabs were taken when the patient was admitted to the hospital. Total DNA was isolated from the samples, then V3V4 regions of the 16s rRNA gene were amplified, followed by sequencing using Illumina HiSeq 2500 platform. DADA2 algorithm was used to obtain amplicon sequence variants (ASV). RESULTS: When comparing the microbial composition of the oropharynx of the patients with different forms of pneumonia, we have identified ASVs associated with the development of both mild and severe pneumonia outside hospital treatment. Based on the results obtained, ASVs associated with a lower degree of lung damage belong predominantly to the class of Gram-negative Firmicutes (Negativicutes), to various classes of Proteobacteria, as well as to the order Fusobacteria. In turn, ASVs associated with a greater degree of lung damage belong predominantly to Gram-positive classes of Firmicutes Bacilli and Clostridia. While being hospitalized, patients with severe pneumonia demonstrated negative disease dynamics during treatment significantly more often. CONCLUSION: We have observed differences in the taxonomic composition of the oropharyngeal microbiota in patients with different forms of pneumonia developed outside hospital treatment against COVID-19. Such differences might be due to the presumed barrier function of the oropharyngeal microbiota, which reduces the risk of virus titer increase.

A hybrid interferometric system for velocity measurements in shock-wave experiments.

This paper presents a hybrid interferometric system designed to measure the surface velocity of tested specimens in shock-wave experiments. The system integrates the All-Fiber Velocity Interferometer System for Any Reflector (AFVISAR) and the Photonic Doppler Velocimeter (PDV) interferometric channels using a single probing system to measure the velocity of one surface point of specimens under study. This design allows the same optical signal containing the Doppler frequency shift to be processed by the AFVISAR and PDV independent interferometric devices. The interferometric system has been tested in dynamic experiments and provides the velocity measurement accuracy of at least 1.5 m/s with a nanosecond time resolution.

[Intestinal microbiome as a predictor of the anti-PD-1 therapy success: metagenomic data analysis]. / Mikrobiom kishechnika kak prediktor uspekha anti-PD1 terapii: analiz metagenomnykh dannykh.

Anti-PD-1 immunotherapy has a large impact on cancer treatment but the rate of positive treatment outcomes is 40-45% and depends on many factors. One of the factors affecting the outcome of immunotherapy is the gut microbiota composition. This effect has been demonstrated both in model objects and in clinical patients groups. However, in order to identify clear causal relationships between microbiota and anti-PD1 immunotherapy response, it is necessary to expand the number of patients and experimental samples. This work presents an analysis of metagenomic data obtained using whole-genome sequencing of stool samples from melanoma patients (n=45) with different responses to anti-PD1 therapy. The analysis of the differential representation of microbial species has shown a difference in the composition of the microbiota between the experimental groups. Results of this study indicate existence of a strong link between the composition of the gut microbiota and the outcome of anti-PD1 therapy. Expansion of similar research may help develop additional predictive tools for the outcome of anti-PD1 cancer immunotherapy, as well as increase its effectiveness.

Phase states, microstructure and dielectric characteristics of solid solutions (1 - x)NaNbO3 - xCa2Nb2O7 and (1 - x)NaNbO3 - xSr2Nb2O7.

Ceramics of binary systems solid solutions (1 - x)NaNbO3 - xCa2Nb2O7 and (1 - x)NaNbO3 - xSr2Nb2O7 with non-isostructural extreme components were prepared by the solid-phase reactions technique with the following sintering using conventional ceramic technology. It was found that ceramics with x ≤ 0.2 have a perovskite structure. Layered type of structure predominates in the concentration range 0.2 < x ≤ 1. Phase diagrams of both systems at room temperature have been determined in the perovskite area. It was shown that this area contains two concentration regions with the different crystal structures and the morphotropic phase boundary between them. Microstructure and dielectric characteristics of selected solid solutions were investigated. The influence of technological regulations, such as mechanical activation and variation of sintering temperatures, on the formation of the microstructure and dielectric characteristics was studied for the individually selected concentrations (x = 0.1 and x = 0.25). Dielectric characteristics of ceramics revealed the presence of the Maxwell-Wagner polarization and its corresponding relaxation in the solid solutions (1 - x)NaNbO3 - xCa2Nb2O7 at x > 0.20.

[Evaluation of metabolites levels in feces of patients with inflammatory bowel diseases]. / Otsenka urovnia metabolitov v fekaliiakh patsientov s vospalitel'nymi zabolevaniiami kishechnika.

Inflammatory bowel diseases (IBD), which include ulcerative colitis (UC) and Crohn's disease (CD), are chronic intestinal inflammatory disorders with an unknown etiology. They are characterized by chronic recurrent inflammation of the intestinal mucosa and lead to a significant decrease in the quality of life and death of patients. IBD are associated with suppression of normal intestinal microflora, including a decrease in bacteria, producers of short chain fatty acids (SCFAs), exhibiting anti-inflammatory and protective properties. Among the various methods of intestinal microflora correction, fecal microbiota transplantation (FMT), which engrafts the fecal microbiota from a healthy donor into a patient recipient, is of a particular interest. As a result, a positive therapeutic effect is observed, accompanied by the restoration of the normal intestinal microflora of the patient. A significant drawback of the method is the lack of standardization. Metabolites produced by intestinal microflora, namely SCFAs, allow objective assessment of the functional state of the intestinal microbiota and, consequently, the success of the FMT procedure. Using gas chromatography and nuclear magnetic resonance spectroscopy techniques, we have analyzed concentrations and molar ratios of SCFAs in fecal samples of 60 healthy donors. Results were in good accord when comparing two methods as well as with published data. Analysis of SCFAs in feces of patients with UC (19 patients) and CD (17 patients) revealed a general decrease in the concentration of fatty acids in the experimental groups with significant fluctuations in the values in experimental groups compared to control group of healthy donors. On the limited group of IBD patients (6 patients with UC and 5 patients with CD) concentration of SCFAs before and within 30 days of observation after FMT was determined. It was shown that FMT had a significant impact on the SCFAs levels within 1 month term; tendency to reach characteristics of healthy donors is unambiguously traced for both diseases.

[Intestinal microbiom modulates the response to antitumor immunotherapy]. / Vliianie mikrobioma kishechnika na éffektivnost' protivoopukholevoi immunoterapii.

Numerous studies confirm the high degree of involvement of the intestinal microbiota in most processes in the human body. There is evidence for the effect of intestinal microbiota on the success of chemo and immunotherapy of oncological diseases. It is assumed that the intestinal microbiota exhibits an indirect effect on the antitumor therapy through such mechanisms as general immunomodulation, an increase in cells that specifically respond to antigens of both microbial and tumor origin, metabolism, degradation (utilization) of drug compounds. The intestinal microbiota is currently considered as an additional, but important target for studying the effective use of antitumor therapy and reducing its toxicity, as well as a predictor of the success of immunotherapy. In this review, we highlight the results of studies published to date that confirm the relationship between gut microbiome and antitumor efficacy of immune checkpoint inhibitors. Despite the promising and theoretically substantiated conclusions, there are still some discrepancies among the existing data that will have to be addressed in order to facilitate the further development of this direction.

Markers of dysbiosis in patients with ulcerative colitis and Crohn's disease.

AIM: The aim of the study was to study the taxonomic and functional composition of the gut microbiota in ulcerative colitis (UC) and Crohn's disease (CD) patients to identify key markers of dysbiosis in IBD. MATERIALS AND METHODS: Fecal samples obtained from 95 IBD patients (78 UC and 17 CD) as well as 96 healthy volunteers were used for whole-genome sequencing carried out on the SOLiD 5500 W platform. Taxonomic profiling was performed by aligning the reeds, not maped on hg19, on MetaPhlAn2 reference database. Reeds were mapped using the HUNAnN2 algorithm to the ChocoPhlAn database to assess the representation of microbial metabolic pathways. Short-chain fatty acids (SCFA) level were measured in fecal samples by gas-liquid chromatographic analysis. RESULTS: Changes in IBD patients gut microbiota were characterized by an increase in the representation of Proteobacteria and Bacteroidetes phyla bacteria and decrease in the number of Firmicutes phylum bacteria and Euryarchaeota phylum archaea; a decrease in the alpha-diversity index, relative representation of butyrate-producing, hydrogen-utilizing bacteria, and Methanobrevibacter smithii; increase in the relative representation of Ruminococcus gnavus in UC and CD patients and Akkermansia muciniphila in CD patients. Reduction of Butyryl-CoA: acetate CoA transferase gene relative representation in CD patients, decrease of absolute content of SCFA total number as well as particular SCFAs and main SCFAs ratio in IBD patients may indicate inhibition of functional activity and number of anaerobic microflora and/or an change in SCFA utilization by colonocytes. CONCLUSION: the revealed changes can be considered as typical signs of dysbiosis in IBD patients and can be used as potential targets for IBD patients personalized treatment development.

Phenomenological theory of uniaxial relaxor ferroelectrics.

A phenomenological thermodynamic theory of uniaxial relaxor strontium barium niobate [Formula: see text] is developed using the Landau-Devonshire approach with two order parameters. The fourth-order thermodynamic potential allowed to explain the shape of the polarization hysteresis loops experimentally observed at different temperatures. We show that the broad maximum of the dielectric permittivity is not related to the phase transition and arise due to the coupling between polarization and true order parameter which has antiferroelectric nature. We found that the phase transition temperature is much higher than the maximum of the dielectric permittivity and very likely corresponds to so-called Burn's temperature. True order parameter has no simple relation with polar modes.

[Metagenomic analysis of taxonomic and functional changes in gut microbiota of patients with alcoholic dependence syndrome].

Here we present the first metagenomic study of gut microbiota in patients with alcohol dependence syndrome (ADS) performed in the whole-genome ("shotgun") format. Taxonomic analysis highlighted changes in community "drivers" abundance previously associated with inflammatory processes (including increase in Ruminococcus gnavus and torques, as well as decrease in Faecalibacterium and Akkermansia). Microbiota of alcoholics manifested presence of specific opportunistic pathogens rarely detected in healthy control subjects of the world. Differential analysis of metabolic potential basing on changes in KEGG Orthology groups abundance revealed increase in pathways associated with response to oxidative stress. Analysis of two specific gene groups--alcohol metabolism and virulence factors--also showed increase in comparison with the control groups. We suggest that gut microbiota distinct in alcoholics by both taxonomic and functional composition plays role in modulating the effect of alcohol on host organism.

[Specific features of the enteric microbiota composition in patients with alcoholic liver cirrhosis].

AIM: To establish the specific features of the taxonomic and functional composition of the enteric microbiota in patients with alcoholic liver cirrhosis (LC). SUBJECTS AND METHODS: Metagenomic analysis was used to study the taxonomic composition and functional potential of the enteric microbiota in 20 patients with alcoholic LC. Total DNA was isolated from the patients' fecal samples; thereafter full genome sequencing was carried out. The metagenomic analysis yielded the results of the relative taxonomic and functional abundance of microbial species in the test samples. These were comparatively analyzed with the previously published metagenomic datasets of healthy population cohorts in the Russian Federation, as well as in Denmark, China, and the USA. RESULTS: In the majority of patients, the dominant part of the intestinal community represented bacterial species constituting the normal human intestinal flora. At the same time, abnormal gut microbiota composition, which was suggestive of marked dysbacteriosis, was identified in a number of patients. In addition, pooled analysis of the data could identify a number of species with a statistically significantly increase and decrease in the relative abundance as compared to the control groups. Thus, the enteric microbiota of the patients with alcoholic LC showed a high proportion of bacteria characteristic of the oral cavity. Analysis of the pooled metabolic potential of the microbiota in these patients demonstrated the higher abundance of enzyme genes involved in alcohol metabolism. CONCLUSION: In the patients with alcoholic LC, the microbiota composition changes identified in individual bacterial species may be associated with gastrointestinal comorbidities, such as chronic erosive gastritis, chronic pancreatitis, and gastric ulcer. The alterations occurring in alcoholic cirrhosis promote the penetration and generation of oral cavity-specific microorganisms in the human intestine. This may a potential biomarker for the diagnosis of liver diseases. The bacterial enzyme genes involved in alcohol metabolism have an increased abundance in patients with alcoholic LC and healthy volunteers from the Russian Federation.

SOD2 gene polymorphism and muscle damage markers in elite athletes.

Exercise-induced oxidative stress is a state that primarily occurs in athletes involved in high-intensity sports when pro-oxidants overwhelm the antioxidant defense system to oxidize proteins, lipids, and nucleic acids. During exercise, oxidative stress is linked to muscle metabolism and muscle damage, because exercise increases free radical production. The T allele of the Ala16Val (rs4880 C/T) polymorphism in the mitochondrial superoxide dismutase 2 (SOD2) gene has been reported to reduce SOD2 efficiency against oxidative stress. In the present study we tested the hypothesis that the SOD2 TT genotype would be underrepresented in elite athletes involved in high-intensity sports and associated with increased values of muscle and liver damage biomarkers. The study involved 2664 Caucasian (2262 Russian and 402 Polish) athletes. SOD2 genotype and allele frequencies were compared to 917 controls. Muscle and liver damage markers [creatine kinase (CK), creatinine, alanine transaminase (ALT), aspartate transaminase (AST), alkaline phosphatase (ALP)] were examined in serum from 1444 Russian athletes. The frequency of the SOD2 TT genotype (18.6%) was significantly lower in power/strength athletes (n = 524) compared to controls (25.0%, p = 0.0076) or athletes involved in low-intensity sports (n = 180; 33.9%, p < 0.0001). Furthermore, the SOD2 T allele was significantly associated with increased activity of CK (females: p = 0.0144) and creatinine level (females: p = 0.0276; males: p = 0.0135) in athletes. Our data show that the SOD2 TT genotype might be unfavorable for high-intensity athletic events.

[Apparatus Aeltis-Synchro-02-"Yarilo" and vacuum laser therapeutic urologic massager AMVL 01-"Yarovit in the treatment of chronic prostatitis complicated with copulation disfunction]. / Primenenie apparatov Aéltis-Sinkhro-02-"Iarilo" i massazhera vakuumnogo lazernogo terapevticheskogo urologicheskogo AMVL 01-"Iarovit" v lechenii khronicheskogo prostatita i soputstvuiushchei kopuliativnoi disfunktsii.

There are three directions in pathogenetic treatment of chronic prostatitis (CP) which is conducted in parallel to etiotropic one (antimicrobial): general immunological; improving arterial inflow and venous outflow; creation of prostatic secretion outflow and that of seminal vesicles by means of contractions of the pelvic and perineal muscles, muscular fibers of the prostatic gland. The latter two directions can be managed with physiotherapy. It is proposed to use combination of drugs with physiotherapy conducted by means of the devices [see text].

[Genetic variants of haptoglobin in patients with destructive pulmonary tuberculosis]. / Geneticheskie varianty gaptoglobina u bol'nykh destruktivnym tuberkulezom legkikh.

Haptoglobin (Hb) phenotyping was carried out in 143 patients with tuberculosis and 221 practically healthy persons. Three phenotypes were distinguished: two homozygotic Hp. 1:1 and Hp. 2:2 and one heterozygotic Hp. 2:1. An analysis of the frequency of distribution of the above-mentioned phenotypes indicates that patients with tuberculosis as compared with the healthy showed a significant reduction of the number of Hp. 2:1 carriers while the number of Hp. 2:2 increased. There was a definite dependence between the haptoglobin phenotype and course of the disease. In carriers of the Hp. 2:1 abacillation of the sputum and closure of destructions occurred earlier than in carriers of Hp 1:1 and, in particular, Hp. 2:2.

[Changes in the biochemical indices of patients with a recurrence of pulmonary tuberculosis]. / Izmeneniia nekotorykh biokhimicheskikh pokazatelei u bol'nykh s retsidivom tuberkuleza legkikh.

It was found that patients with pulmonary tuberculosis recurrences revealed an imbalance between the elevated level of lipid peroxidation and possibilities of antioxidant protection. The proteinase inhibitor system balance was disturbed. The general level of haptoglobulin rises while the number of persons with homozygote of haptoglobulin 2:2 rose and the number of persons with the heterozygote phenotype 2:1 reduced. Normalization of the biochemical indices in patients with recurrences was delayed.

[The role of genetically determined haptoglobin phenotypes in patients with destructive pulmonary tuberculosis]. / Znachenie geneticheski determinirovannykh fenotipov gaptoglobina u bol'nykh s destruktivnym tuberkulezom legkikh.

Haptoglobin (Hp) was phenotyped in 567 apparently healthy individuals and 223 patients with destructive pulmonary tuberculosis, including 163 as newly-detected and 60 as recurrent cases. Three phenotypes of haptoglobin were identified, i.e. two homozygotic (Hp 1:1 and Hp 2:2) and one heterozygotic (Hp 2:1). It was revealed that among the pulmonary tuberculosis patients, both newly detected and recurrent cases, the number of Hp 2:2 carriers significantly increased, as compared to healthy individuals, while that of Ph 2:1 carriers decreased. A certain relation between the course of destructive pulmonary tuberculosis and genetically determined types of Hp was found. Sputum conversion and cavity closure in Hp 2:2 and Hp 1:1 phenotype carriers, as compared to Hp 2:1 ones, occurred more infrequently and in later periods. It is felt that the presence of the 2:2 phenotype is an adverse hereditary factor in relation to tuberculosis.

[Treatment of patients with destructive pulmonary tuberculosis with chemotherapeutic preparations combined with sodium nucleinate]. / Lechenie bol'nykh destruktivnym tuberkulezom legkikh khimiopreparatami v sochetanii s nukleinatom natriia.

The treatment efficacy was studied in 90 patients with destructive tuberculosis of the lungs. 50 of them were subjected to combined chemotherapy with inclusion of sodium nucleinate and 40 were treated with the tuberculostatics alone. It was shown that sodium nucleinate improved the host metabolism and in particular increased nucleonemia. As compared to the controls in the patients treated with sodium nucleinate there was a decrease in the periods of the host disintoxication, discontinuation of tubercle bacilli isolation and closing of destructive cavities. Sodium nucleinate was satisfactorily tolerated by the patients, it induced no adverse reactions and had a favourable effect on the patients immunity.