Universal Method Based on Layer-by-Layer Assembly for Aptamer-Based Sensors for Small-Molecule Detection.

Development of a fast and accurate pesticide analysis system is a challenging task, as a large amount of commonly used pesticide has negative effects on humans' health. Detection of pesticide residues is crucial for food safety management and environmental protection. Aptamers─short single-stranded oligonucleotides (RNA or DNA) selected by aptamer selection method SELEX─can selectively bind to their target pesticide molecules with high affinity. Thus, in the present study, we developed an electrochemical biosensor based on aptamers to detect the commonly used pesticide, glyphosate. Carbon fibers were used as the platform to assemble polyelectrolyte layers with the incorporated aptamers selectively binding with glyphosate molecules for electrochemical detection. The best limit of detection of 0.3 µM was achieved at open-circuit potential measurements, which is comparable to the current need in detection of glyphosate. The developed method can be implemented into existing systems for the determination of pesticides on farms to control residual concentrations of glyphosate in soil and water.

The Succession of the Cellulolytic Microbial Community from the Soil during Oat Straw Decomposition.

The process of straw decomposition is dynamic and is accompanied by the succession of the microbial decomposing community, which is driven by poorly understood interactions between microorganisms. Soil is a complex ecological niche, and the soil microbiome can serve as a source of potentially active cellulolytic microorganisms. Here, we performed an experiment on the de novo colonization of oat straw by the soil microbial community by placing nylon bags with sterilized oat straw in the pots filled with chernozem soil and incubating them for 6 months. The aim was to investigate the changes in decomposer microbiota during this process using conventional sequencing techniques. The bacterial succession during straw decomposition occurred in three phases: the early phase (first month) was characterized by high microbial activity and low diversity, the middle phase (second to third month) was characterized by low activity and low diversity, and the late phase (fourth to sixth months) was characterized by low activity and high diversity. Analysis of amplicon sequencing data revealed three groups of co-changing phylotypes corresponding to these phases. The early active phase was abundant in the cellulolytic members from Pseudomonadota, Bacteroidota, Bacillota, and Actinobacteriota for bacteria and Ascomycota for fungi, and most of the primary phylotypes were gone by the end of the phase. The second intermediate phase was marked by the set of phylotypes from the same phyla persisting in the community. In the mature community of the late phase, apart from the core phylotypes, non-cellulolytic members from Bdellovibrionota, Myxococcota, Chloroflexota, and Thermoproteota appeared. Full metagenome sequencing of the microbial community from the end of the middle phase confirmed that major bacterial and fungal members of this consortium had genes of glycoside hydrolases (GH) connected to cellulose and chitin degradation. The real-time analysis of the selection of these genes showed that their representation varied between phases, and this occurred under the influence of the host, and not the GH family factor. Our findings demonstrate that soil microbial community may act as an efficient source of cellulolytic microorganisms and that colonization of the cellulolytic substrate occurs in several phases, each characterized by its own taxonomic and functional profile.

The Structure of Stable Cellulolytic Consortia Isolated from Natural Lignocellulosic Substrates.

Recycling plant matter is one of the challenges facing humanity today and depends on efficient lignocellulose degradation. Although many bacterial strains from natural substrates demonstrate cellulolytic activities, the CAZymes (Carbohydrate-Active enZYmes) responsible for these activities are very diverse and usually distributed among different bacteria in one habitat. Thus, using microbial consortia can be a solution to rapid and effective decomposition of plant biomass. Four cellulolytic consortia were isolated from enrichment cultures from composting natural lignocellulosic substrates-oat straw, pine sawdust, and birch leaf litter. Enrichment cultures facilitated growth of similar, but not identical cellulose-decomposing bacteria from different substrates. Major components in all consortia were from Proteobacteria, Actinobacteriota and Bacteroidota, but some were specific for different substrates-Verrucomicrobiota and Myxococcota from straw, Planctomycetota from sawdust and Firmicutes from leaf litter. While most members of the consortia were involved in the lignocellulose degradation, some demonstrated additional metabolic activities. Consortia did not differ in the composition of CAZymes genes, but rather in axillary functions, such as ABC-transporters and two-component systems, usually taxon-specific and associated with CAZymes. Our findings show that enrichment cultures can provide reproducible cellulolytic consortia from various lignocellulosic substrates, the stability of which is ensured by tight microbial relations between its components.

Consensus guidelines for botulinum toxin therapy: general algorithms and dosing tables for dystonia and spasticity.

Botulinum toxin (BT) therapy is a complex and highly individualised therapy defined by treatment algorithms and injection schemes describing its target muscles and their dosing. Various consensus guidelines have tried to standardise and to improve BT therapy. We wanted to update and improve consensus guidelines by: (1) Acknowledging recent advances of treatment algorithms. (2) Basing dosing tables on statistical analyses of real-life treatment data of 1831 BT injections in 36 different target muscles in 420 dystonia patients and 1593 BT injections in 31 different target muscles in 240 spasticity patients. (3) Providing more detailed dosing data including typical doses, dose variabilities, and dosing limits. (4) Including total doses and target muscle selections for typical clinical entities thus adapting dosing to different aetiologies and pathophysiologies. (5) In addition, providing a brief and concise review of the clinical entity treated together with general principles of its BT therapy. For this, we collaborated with IAB-Interdisciplinary Working Group for Movement Disorders which invited an international panel of experts for the support.

Investigation of Bemethyl Biotransformation Pathways by Combination of LC-MS/HRMS and In Silico Methods.

Bemethyl is an actoprotector, an antihypoxant, and a moderate psychostimulant. Even though the therapeutic effectiveness of bemethyl is well documented, there is a gap in knowledge regarding its metabolic products and their quantitative and qualitative characteristics. Since 2018, bemethyl is included to the Monitoring Program of the World Anti-Doping Agency, which highlights the challenge of identifying its urinary metabolites. The objective of the study was to investigate the biotransformation pathways of bemethyl using a combination of liquid chromatography-high-resolution mass spectrometry and in silico studies. Metabolites were analyzed in a 24 h rat urine collected after oral administration of bemethyl at a single dose of 330 mg/kg. The urine samples were prepared for analysis by a procedure developed in the present work and analyzed by high performance liquid chromatography-tandem mass spectrometry. For the first time, nine metabolites of bemethyl with six molecular formulas were identified in rat urine. The most abundant metabolite was a benzimidazole-acetylcysteine conjugate; this biotransformation pathway is associated with the detoxification of xenobiotics. The BioTransformer and GLORY computational tools were used to predict bemethyl metabolites in silico. The molecular docking of bemethyl and its derivatives to the binding site of glutathione S-transferase has revealed the mechanism of bemethyl conjugation with glutathione. The findings will help to understand the pharmacokinetics and pharmacodynamics of actoprotectors and to improve antihypoxant and adaptogenic therapy.

Proteomics of the 26S proteasome in Spodoptera frugiperda cells infected with the nucleopolyhedrovirus, AcMNPV.

Baculoviruses are large DNA viruses that infect insect species such as Lepidoptera and are used in biotechnology for protein production and in agriculture as insecticides against crop pests. Baculoviruses require activity of host proteasomes for efficient reproduction, but how they control the cellular proteome and interact with the ubiquitin proteasome system (UPS) of infected cells remains unknown. In this report, we analyzed possible changes in the subunit composition of 26S proteasomes of the fall armyworm, Spodoptera frugiperda (Sf9), cells in the course of infection with the Autographa californica multiple nucleopolyhedrovirus (AcMNPV). 26S proteasomes were purified from Sf9 cells by an immune affinity method and subjected to 2D gel electrophoresis followed by MALDI-TOF mass spectrometry and Mascot search in bioinformatics databases. A total of 34 homologues of 26S proteasome subunits of eukaryotic species were identified including 14 subunits of the 20S core particle (7 α and 7 ß subunits) and 20 subunits of the 19S regulatory particle (RP). The RP contained homologues of 11 of RPN-type and 6 of RPT-type subunits, 2 deubiquitinating enzymes (UCH-14/UBP6 and UCH-L5/UCH37), and thioredoxin. Similar 2D-gel maps of 26S proteasomes purified from uninfected and AcMNPV-infected cells at 48hpi confirmed the structural integrity of the 26S proteasome in insect cells during baculovirus infection. However, subtle changes in minor forms of some proteasome subunits were detected. A portion of the α5(zeta) cellular pool that presumably was not associated with the proteasome underwent partial proteolysis at a late stage in infection.

Strategies for treatment of dystonia.

Treatment of dystonias is generally symptomatic. To produce sufficient therapy effects, therefore, frequently a multimodal and interdisciplinary therapeutic approach becomes necessary, combining botulinum toxin therapy, deep brain stimulation, oral antidystonic drugs, adjuvant drugs and rehabilitation therapy including physiotherapy, occupational therapy, re-training, speech therapy, psychotherapy and sociotherapy. This review presents the recommendations of the IAB-Interdisciplinary Working Group for Movement Disorders Special Task Force on Interdisciplinary Treatment of Dystonia. It reviews the different therapeutic modalities and outlines a strategy to adapt them to the dystonia localisation and severity of the individual patient. Hints to emerging and future therapies will be given.

Inhibitory effects of Lactobacillus fermentum on microbial growth and biofilm formation.

Beneficial effects of Lactobacilli have been reported, and lactic bacteria are employed for conservation of foods. Therefore, the effects of a Lactobacillus fermentum strain were analyzed regarding inhibitory effects on staphylococci, Candida albicans and enterotoxigenic enterobacteria by transmission electron microscopy (TEM). TEM of bacterial biofilms was performed using cocultures of bacteriocin-producing L. fermentum 97 with different enterotoxigenic strains: Staphylococcus epidermidis expressing the ica gene responsible for biofilm formation, Staphylococcus aureus producing enterotoxin type A, Citrobacter freundii, Enterobacter cloaceae, Klebsiella oxytoca, Proteus mirabilis producing thermolabile and thermostable enterotoxins determined by elt or est genes, and Candida albicans. L. fermentum 97 changed morphological features and suppressed biofilm formation of staphylococci, enterotoxigenic enterobacteria and Candida albicans; a marked transition to resting states, a degradation of the cell walls and cytoplasm, and a disruption of mature bacterial biofilms were observed, the latter indicating efficiency even in the phase of higher cell density.

Validation of two automatic sphygmomanometers according to the modified International Standardization Organization 81060-2:2018 protocol in adults with a mid-upper arm circumference of 22 centimeters or less.

OBJECTIVE: A recent International Organization for Standardization (ISO) Task Group report calls for research to investingate potential special populations in validation studies of automated blood pressure (BP) devices. Accordingly, we aimed to determine the accuracy of two previously validated BP monitors passed in a general population when measured in adults with a mid-upper arm circumference (MUAC) ≤ 22 cm. METHODS: Test device A was the OMRON HEM 7121 equipped with the HEM CS24 cuff designed for an arm circumference of 17-22 cm. Test device B was the YuWell YE660Е with the YuWell '360°' cuff (18-36 cm). Data from 37 participants aged 20.14 ± 2.23 (18-28) years were analyzed according to criterion 1 of ISO Standard 81060-2 : 2018. RESULTS: According to criterion 1, the mean ± SD of the BP differences for the device A was -7.81 ± 5.20/-10.66 ± 5.48 mmHg (systolic/diastolic) and for the device B was -8.00 ± 6.30/-16.11 ± 5.15 mmHg (systolic/diastolic), respectively. This means that neither device met the requirements. CONCLUSION: Since devices A and B, which had passed in a general population study, failed in adults aged 18-28 years with an MUAC ≤ 22 cm, such individuals might be considered as new special population for validation studies.

Comparative Evaluation of the Activity of Various Lentiviral Vectors Containing Three Anti-HIV Genes.

A promising direction in the treatment of HIV infection is a gene therapy approach based on the insertion of antiviral genes aimed at inhibiting HIV replication into the genome of host cells. We obtained six constructs of lentiviral vectors with different arrangements of three antiviral genes: microRNAs against the CCR5 gene, the gene encoding the C-peptide, and the gene encoding the modified human TRIM5a protein. We found that despite containing the same genes, these vectors were produced at different titers and had different effects on cell viability, transduction efficiency, and expression stability. Comparative evaluation of the antiviral activity of three of the six developed vectors that showed stable expression was carried out using the continuous SupT1 lymphocytic cell line. All of the vectors protected cells from HIV infection: the viral load was several orders of magnitude lower than in control cells, and with one vector, complete cessation of virus growth in modified cells was achieved.

Development of MVA-d34 Tetravalent Dengue Vaccine: Design and Immunogenicity.

Dengue fever, an infectious disease that affects more than 100 million people every year, is a global health problem. Vaccination may be the most effective prevention strategy for the disease. However, the development of vaccines against dengue fever is complicated by the high risk of developing an antibody-dependent increase in infection. This article describes the development of an MVA-d34 vaccine against the dengue virus based on a safe and effective MVA viral vector. The DIII domains of the envelope protein (E) of the dengue virus are used as vaccine antigens, as antibodies against these domains do not cause an enhancement of infection. The use of the DIII domains of each of the four dengue virus serotypes made it possible to generate a humoral response against all four dengue virus serotypes in immunized mice. We also showed that the sera of vaccinated mice present virus-neutralizing activity against dengue serotype 2. Thus, the developed MVA-d34 vaccine is a promising candidate vaccine against dengue fever.

A Moderate Decrease in ADAMTS13 Activity Correlates with the Severity of STEC-HUS.

Atypical hemolytic uremic syndrome (HUS) develops as a result of damage to the endothelium of microvasculature vessels by Shiga toxin produced by enterohemorrhagic Escherichia coli (STEC-HUS). STEC-HUS remains the leading cause of acute kidney injury (AKI) in children aged 6 months to 5 years. The pathomorphological essence of the disease is the development of thrombotic microangiopathy (TMA). One of the key causes of TMA is an imbalance in the ADAMTS13-von Willebrand factor (vWF)-platelet system. The goal of the work was to clarify the role of a moderate decrease in ADAMTS13 activity in the pathogenesis of STEC-HUS. The activity of ADAMTS13 was determined in 138 children (4 months-14.7 years) in the acute period of STEC-HUS and the features of the course of the disease in these patients were analyzed. The study revealed a decrease in the activity and concentration of ADAMTS13 in 79.8% and 90.6% of patients, respectively. Measurements of von Willebrand factor antigen content and the activity of von Willebrand factor in the blood plasma of part of these patients were carried out. In 48.6% and 34.4% of cases, there was an increase in the antigen concentration and the activity of the Willebrand factor, respectively. Thrombocytopenia was diagnosed in 97.8% of children. We have demonstrated that moderately reduced ADAMTS13 activity correlates with the risk of severe manifestations of STEC-HUS in children; the rate of developing multiple organ failure, cerebral disorders, pulmonary edema, and acute kidney injury with the need for dialysis increases. It is assumed that reduction in ADAMTS13 activity may serve as a predictor of disease severity.

Dynamic of the Soil Microbiota in Short-Term Crop Rotation.

Crop rotation is one of the oldest and most effective methods of restoring soil fertility, which declines when the same plant is grown repeatedly. One of the reasons for a reduction in fertility is the accumulation of pathogenic and unfavorable microbiota. The modern crop rotation schemes (a set of plant species and their order in the crop rotation) are highly effective but are designed without considering soil microbiota dynamics. The main goal of this study was to perform a short-term experiment with multiple plant combinations to access the microbiological effects of crop rotation. It could be useful for the design of long-term crop rotation schemes that take the microbiological effects of the crop rotation into account. For the analysis, five plants (legumes: vetch, clover, and cereals: oats, wheat, and barley) were used. These five plants were separately grown in pots with soil. After the first phase of vegetation, the plants were removed from the soil and a new crop was planted. Soil samples from all 25 possible combinations of primary and secondary crops were investigated using v4-16S rDNA gene sequencing. It was shown that the short-term experiments (up to 40 days of growing) are effective enough to find microbial shifts in bulk soil from different plants. Both primary and secondary cultures are significant factors for the microbial composition of microbial soil communities. Changes are the most significant in the microbial communities of vetch soils, especially in the case of vetch monoculture. Growing clover also leads to changes in microbiota, especially according to beta-diversity. Data obtained can be used to develop new crop rotation schemes that take into account the microbiological effects of various crops.

Electrochemical Sensor to Detect Antibiotics in Milk Based on Machine Learning Algorithms.

The present study is dedicated to the problem of electrochemical analysis of multicomponent mixtures, such as milk. A combination of cyclic voltammetry facilities and machine learning techniques made it possible to create a pattern recognition system for the detection of antibiotic residues in skimmed milk. A multielectrode sensor including copper, nickel, and carbon fiber was fabricated for the collection of electrochemical data. Processes occurring at the electrode surface were discussed and simulated with the help of molecular docking and density functional theory modeling. It was assumed that the antibiotic fingerprint reveals a potential drift of electrodes, owing to complexation with metal ions present in milk. The gradient boosting algorithm showed the best efficiency in training the machine learning model. High accuracy was achieved for the recognition of antibiotics in milk. The elaborated method may be incorporated into existing milking systems at dairy farms for monitoring the residue concentrations of antibiotics.

Development of a Universal Epitope-Based Influenza Vaccine and Evaluation of Its Effectiveness in Mice.

Vaccination is an effective and economically viable means of protection against the influenza virus, but due to rapid viral evolution, modern seasonal vaccines are not effective enough. Next-generation vaccines are designed to provide protection against a wide range of influenza virus strains, including pandemic variants. In our work, we made an epitope-based universal vaccine, rMVA-k1-k2, against the influenza virus based on the modified vaccinia Ankara (MVA) vector and using our own algorithms to select epitopes from conserved fragments of the NP, M1 and HA proteins of influenza A and B. We show that double immunization protects mice with a 67% or greater efficiency against viral influenza pneumonia when infected with various strains of the H1N1, H2N2, H3N2 and H5N1 subtypes of influenza A. In animals, the level of protection provided by the rMVA-k1-k2 vaccine was comparable to that provided by the universal M001 and MVA-NP+M1 (Invictus) vaccines, which have shown success in clinical trials, against strains of the H1N1 and H3N2 subtypes.

Development of Modified Vaccinia Virus Ankara-Based Vaccines: Advantages and Applications.

Modified vaccinia virus Ankara (MVA) is a promising viral vector for vaccine development. MVA is well studied and has been widely used for vaccination against smallpox in Germany. This review describes the history of the origin of the virus and its properties as a vaccine, including a high safety profile. In recent years, MVA has found its place as a vector for the creation of vaccines against various diseases. To date, a large number of vaccine candidates based on the MVA vector have already been developed, many of which have been tested in preclinical and clinical studies. We discuss data on the immunogenicity and efficacy of some of these vaccines.

Iatrogenic Glossopharyngeal Neuropathy in Aesthetic Practice: A Case Report.

Iatrogenic glossopharyngeal nerve injuries are commonly associated with procedures such as a tonsillectomy, carotid endarterectomy, and endotracheal intubation. We present a previously unreported complication of a thread lift procedure in the jowl region, causing damage to the glossopharyngeal nerve through compression. The glossopharyngeal nerve belongs to the bulbar group of the cranial nerves. It is connected with the vagus and, therefore, is closely associated with the latter functionally and anatomically. Damage to the former may present with cardiovascular complications associated with the vagus nerve. The presented case demonstrates the diagnostic and treatment aspects of iatrogenic injury to the glossopharyngeal nerve.

Soft Hydrogel Actuator for Fast Machine-Learning-Assisted Bacteria Detection.

We demonstrate that our bio-electrochemical platform facilitates the reduction of detection time from the 3-day period of the existing tests to 15 min. Machine learning and robotized bioanalytical platforms require the principles such as hydrogel-based actuators for fast and easy analysis of bioactive analytes. Bacteria are fragile and environmentally sensitive microorganisms that require a special environment to support their lifecycles during analytical tests. Here, we develop a bio-electrochemical platform based on the soft hydrogel/eutectic gallium-indium alloy interface for the detection of Streptococcus thermophilus and Bacillus coagulans bacteria in various mediums. The soft hydrogel-based device is capable to support bacteria' viability during detection time. Current-voltage data are used for multilayer perceptron algorithm training. The multilayer perceptron model is capable of detecting bacterial concentrations in the 104 to 108 cfu/mL range of the culture medium or in the dairy products with high accuracy (94%). Such a fast and easy biodetection is extremely important for food and agriculture industries and biomedical and environmental science.

Sputnik V Effectiveness against Hospitalization with COVID-19 during Omicron Dominance.

Mass vaccination campaigns against COVID-19 affected more than 90% of the population in most developed countries. The new epidemiologic wave of COVID-19 has been ongoing since the end of 2021. It is caused by a virus variant B.1.1.529, also known as "Omicron" and its descendants. The effectiveness of major vaccines against Omicron is not known. The purpose of this study is to evaluate the efficacy of the Sputnik V vaccine. The main goal is to assess its protection against hospitalization in the period of Omicron dominance. We conducted our study based on a large clinical center in Moscow (Russia) where 1112 patients were included. We used the case-population method to perform the calculations. The data we obtained indicate that the Omicron variant causes at least 90% of infections in the studied cohort. The effectiveness of protection against hospitalization with COVID-19 in our study was 85.9% (95% CI 83.0-88.0%) for those who received more than one dose. It was 87.6% (95% CI 85.4-89.5%) and 97.0% (95% CI 95.9-97.8%) for those who received more than two or three doses. The effectiveness in cases of more severe forms was higher than for less severe ones. Thus, present study indicates the high protective efficacy of vaccination against hospitalization with COVID-19 in case of Omicron lineage.