Background: Seasonal influenza remains a significant public health problem, and the constant antigenic drift of viruses requires regular vaccine updates. mRNA vaccines offer a promising platform for the development of new, effective influenza vaccines. Administration of the naked mRNA vaccine using a needle-free jet injection system further enhances its safety, reduces cost, and eliminates the need for lipid nanoparticles, which are traditionally used for mRNA delivery. Lyophilization of naked mRNA allows for long-term storage at +4 °C. Methods: We designed and produced an mRNA vaccine against seasonal influenza, designated mRNA-Vector-Flu, encoding the hemagglutinin (HA) of the A/Wisconsin/67/2022(H1N1)pdm09, A/Darwin/9/2021(H3N2), and B/Austria/1359417/2021 strains. The vaccine was lyophilized and stored for 1 month in a refrigerator (+4 °C). A comparative immunogenicity study was conducted between synthesized immediately before use prepared and lyophilized naked mRNA-Vector-Flu. The preparations were administered to BALB/c mice using a jet needleless injection twice, 3 weeks apart. Immunogenicity was assessed on day 35 of the study. Results: A comparative immunogenicity study of naked mRNA-Vector-Flu demonstrated that both the synthesized immediately before use prepared formulation and the lyophilized form, stored at +4 °C for a month, induced similar levels of virus-specific antibodies and generated a pronounced T-cell immune response. Conclusions: Delivery of the naked mRNA vaccine using a needle-free jet injection ensures a high-level immune response, which improves its safety, reduces its cost, and eliminates the need for lipid nanoparticles traditionally used for mRNA delivery. At the same time, lyophilization of the naked mRNA vaccine preserves its biological activity and ensures its storage for at least a month at +4 °C temperatures. Our results demonstrate that our proposed approach can be considered a promising direction for the development and improvement of the mRNA vaccine platform.

Background: Since the emergence of SARS-CoV-2 in the human population, the virus genome has undergone numerous mutations, enabling it to enhance transmissibility and evade acquired immunity. As a result of these mutations, most monoclonal neutralizing antibodies have lost their efficacy, as they are unable to neutralize new variants. Antibodies that neutralize a broad range of SARS-CoV-2 variants are of significant value in combating both current and potential future variants, making the identification and development of such antibodies an ongoing critical goal. This study discusses the strategy of using heterologous antigens in biopanning rounds. Methods: After four rounds of biopanning, nanobody variants were selected from a phage display library. Immunochemical methods were used to evaluate their specificity to the S protein of various SARS-CoV-2 variants, as well as to determine their competitive ability against ACE2. Viral neutralization activity was analyzed. A three-dimensional model of nanobody interaction with RBD was constructed. Results: Four nanobodies were obtained that specifically bind to the receptor-binding domain (RBD) of the SARS-CoV-2 spike glycoprotein and exhibit neutralizing activity against various SARS-CoV-2 strains. Conclusions: The study demonstrates that performing several rounds of biopanning with heterologous antigens allows the selection of nanobodies with a broad reactivity spectrum. However, the fourth round of biopanning does not lead to the identification of nanobodies with improved characteristics.

Previously only two families were known with progressive autosomal recessive deafness 103 (DFNB103, OMIM616042) caused by pathogenic variants of the CLIC5 gene. In this study we present the novel truncating variant c.644 G > A p.(Trp215*) of this gene which was found in homozygous state among 22 patients with hearing loss (HL) from 16 unrelated families living in the Sakha Republic of Russia (Eastern Siberia). Genotype-phenotype analysis in patients with DFNB103 showed that HL was sensorineural, symmetrical and variable by severity (from moderate to profound). Audiograms mostly have a down curve configuration, with pronounced loss of high and mid frequencies. In most cases this form of HL was detected in the post-lingual period (mean age 7.9 ± 1.2 years) and has a significant severity progression with age. In average the patients with DFNB103 lost 7.4 ± 13.65 dB on the speech frequency range in pure tone averages (PTA0.5,1.0,2.0,4.0 kHz) per year until reaching profound deafness in the second or third decade of the life. The high frequency of c.644 G > A p.(Trp215*) was found among Siberian GJB2-negative patients (9.9

Mitochondrial forms account approximately 1–2% of all nonsyndromic cases of hearing loss (HL). One of the most common causative variants of mtDNA is the m.1555A > G variant of the MT-RNR1 gene (OMIM 561000). Currently the detection of the m.1555A > G variant of the MT-RNR1 gene is not included in all research protocols. In this study this variant was screened among 165 patients with HL from the Republic of Buryatia, located in the Baikal Lake region of Russia. In our study, the total contribution of the m.1555A > G variant to the etiology of HL was 12.7% (21/165), while the update global prevalence of this variant is 1.8% (863/47,328). The m.1555A > G variant was notably more prevalent in Buryat (20.2%) than in Russian patients (1.3%). Mitogenome analysis in 14 unrelated Buryat families carrying the m.1555A > G variant revealed a predominant lineage: in 13 families, a cluster affiliated with sub-haplogroup A5b (92.9%) was identified, while one family had the D5a2a1 lineage (7.1%). In a Russian family with the m.1555A > G variant the lineage affiliated with sub-haplogroup F1a1d was found. Considering that more than 90% of Buryat families with the m.1555A > G variant belong to the single maternal lineage cluster we conclude that high prevalence of this variant in patients with HL in the Baikal Lake region can be attributed to a founder effect.

The annual number of reported human cases of flavivirus infections continues to increase. Measures taken by local healthcare systems and international organizations are not fully successful. In this regard, new approaches to treatment and prevention of flavivirus infections are relevant. One promising approach is to use monoclonal antibody preparations. The mouse mAb 10H10 is capable of interacting with viruses belonging to the genus Orthoflavivirus which are pathogenic to humans. ELISA and molecular modeling data can indicate that mAb 10H10 recognizes the fusion loop region of E protein. The KD of interaction between the mAb 10H10 and recombinant analogs of the E protein of the tick-borne encephalitis (TBEV), Zika (ZIKV) and dengue (DENV) viruses range from 1.5 to 4 nM. The aim of this study was to map the epitope of this antibody using phage display technology. After three rounds of biopanning, 60 individual phage clones were chosen. The amino acid sequences of the selected peptides were conveniently divided into five groups. Based on the selected peptides, bacteriophages were obtained carrying peptides on the surfaces of the pIII and pVIII proteins, which were tested for binding to the antibody in ELISA. Thus, the epitope of the mAb 10H10 is the highly conserved region 98-DRGWGNXXGLFGK-110 of the flavivirus E protein. The structures of the complexes of the identified peptides with the antibody paratope are proposed using the molecular docking and dynamics methods.

Among the many diseases that affect potato plants, viral infections are the most common and cause significant damage to farms, affecting both the yield and quality of potatoes. In this regard, an important condition for preserving the potato seed fund in Russia is systematic monitoring and early highly specific detection of potato viral infections. The purpose of the work is to study samples of potato varieties collected in the Novosibirsk region for the presence of viral infections using RT-PCR. 130 potato plants from three districts of the Novosibirsk region (NR) were studied. As a result of monitoring, the following viruses were identified: PVY (potato virus Y), PVS (potato virus S), PVM (potato virus M) and PVX (potato virus X). The quarantine pathogen potato spindle tuber viroid (PSTVd) was not detected in any of the samples analyzed. The maximum frequency of occurrence in the region was noted for three viruses: PVY, PVM and PVS. A significant proportion of the samples were mixed viral infections: the occurrence of the combination of infection PVY + PVM in plants was 25.0 %, and PVY + PVS, 22.6 %. To develop methods for determining the strain affiliation of the studied samples, the nucleotide sequences of the capsid protein genes of 10 Y-virus isolates were sequenced. Phylogenetic analysis of the studied sequences of NR isolates was carried out with a set of sequences of reference strains 261-4, Eu-N, N:O, NE-11, NTNa, NTNb, N-Wi, O, O5, SYR_I, SYR_II and SYR_III retrieved from GenBank. As a result of phylogenetic analysis, it was established that NR viral samples fell into two groups of strains: group 1, which also includes isolates of the reference strains 261-4/SYR_III, and group 2, NTNa. The obtained results of the strain affiliation of NR samples lay the basis for the development of DNA and immunodiagnostic systems for identifying PVY circulating in NR, as well as for elucidating the source and routes of entry of specific virus strains.

The removal of double-stranded RNA (dsRNA) contaminants during in vitro mRNA synthesis is one of the technological problems to be solved. Apparently, these contaminants are the result of the T7 RNA polymerase side activity. In this study, we used a modified method of mRNA purification based on the selective binding of dsRNA to cellulose in ethanol-containing buffer. It was shown both in vivo and in vitro that the cellulose-purified mRNA preparation leads neither to activation of the lymphocyte inflammatory marker CD69 nor to increased release of IFNα in mice, and does not contain impurities detectable by antibodies to dsRNA.

We represent a comparative analysis of GBSS1 gene fragment sequences for a number of species related to Elymus caninus: Elymus prokudinii, Elymus viridiglumis, Elymus goloskokovii, as well as a number of morphologically deviating biotypes, inhabiting Russia and Kazakhstan. Microevolutionary relationships between species were assessed from dendrograms derived from sequences of exons and introns. In all taxa, the St subgenome was represented by St(2) variants, rather typical of the North American ancestral line of Pseudoroegneria spicata than of the Asian line descending from Pseudoroegneria strigosa. All putative relatives of E. caninus had H-1 subgenome variants linked around the Asian diploid carrier of the H genome from Hordeum jubatum and were divided into two subclades. One of them (H1-1) contained most of the closely related E. caninus clones, including Elymus uralensis. Another subclade (H1-2) consisted of five variants phylogenetically related to Elymus mutabilis. We have also studied reproductive relationships between species E. goloskokovii, E. prokudinii, and E. viridiglumis and the degree of their integration into the E. caninus complex. Biotypes included in sexual hybridization formed a single recombination gene pool, within which slight differences in reproductive compatibility were observed. A comprehensive study of microevolutionary differentiation of taxa showed the expediency of taxonomic revision. The species mentioned should probably be relegated to the infraspecific rank within E. caninus s. l.

The analysis of DNA nucleotide sequences is an important source of information in the identification of plant taxa or plant components in the composition of medicines. In this work, we studied the possibility of using universal molecular markers for DNA barcoding of rare plant species distributed in the Altai Biosphere Reserve, located in the Altai Republic (South Siberia, Russia). In order to identify commercially-used endangered species, universal markers recommended for DNA barcoding were applied: ITS (Internal Transcribed Spacers) region of nuclear ribosomal DNA and four regions of chloroplast DNA: trnL-F, trnH-psbA rpl32-trnL, and rpl16. Among 61 species of rare flowering plants protected in the Altai Biosphere Reserve, ten species belonging to eight families were studied. The results of sequencing showed that ITS, rpl32-trnL and trnH-psbA core plant barcodes are the most effective markers for discriminating rare species from the Altai Biosphere Reserve.

The structure, mechanical characteristics, and high-temperature stability in vacuum and air of ZrB 2 and HfB 2 -based materials sintered at a high quasi-hydrostatic pressure (4.1 GPa) under hot pressing (at a pressure of 30 MPa) with and without SiC and Si 3 N 4 additives have been studied. It has been shown that short-term sintering (4 min) under high pressure conditions and at a comparatively low temperature (1800°C) essentially improves the mechanical properties of these materials as compared to the similar materials synthesized by the other method (hot pressing and spark-plasma sintering). In the case of sintering at a high pressure (4.1 GPa), the addition of 20 wt % SiC to ZrB 2 and 30 wt % SiC to HfB 2 leads to a decrease in the specific gravity of ZrB 2 and HfB 2 and increases their hardness by 17 and 46% and fracture toughness by 40 and 21%, respectively. When SiC is added, there occurs the formation of solid solutions through the mutual diffusion of C and Si into the ZrB 2 or HfB 2 matrix phases and the slight diffusion of Zr and Hf into SiC-enriched areas. The improvement of the mechanical properties of ZrB 2 and HfB 2 sintered at a high pressure without additives is explained by the formation of stronger bonds between the sintered material grains. The addition of SiC to ZrB 2 slightly decreases the Young modulus, but increases the damping ability of the synthesized materials. The simultaneous addition of SiC and Si 3 N 4 to ZrB 2 leads to an increase in the hardness to a smaller extent, but results in a further increase in fracture toughness. The melting temperature in vacuum of sintered ZrB 2 and HfB 2 has proven to be much higher as compared to the materials with SiC additives. The composite material synthesized from a HfB 2 –30 wt % SiC mixture has a density ρ = 6.21 g/cm 3 , a microhardness H V (9.8 N) = 38.1 ± 1.4 GPa, H V (49 N) = 27.7 ± 0.24 GPa, H V (98 N) = 26.3 ± 2.03 GPa, and a fracture toughness K І с (9.8 N) = 8.2 ± 0.2 MPa m 0.5 , K І с (49 N) = 6.8 ± 0.6 MPa m 0.5 , K І с (98 N) = 6.4 ± 0.11 MPa m 0.5 , which are much higher than the similar characteristics of HfB 2 sintered under the same conditions, but without the additives.

The GJB2 (Cx26) gene pathogenic variants are associated with autosomal recessive deafness type 1A (DFNB1A, OMIM #220290). Direct sequencing of the GJB2 gene among 165 hearing-impaired individuals living in the Baikal Lake region of Russia identified 14 allelic variants: pathogenic/likely pathogenic-nine variants, benign-three variants, unclassified-one variant, and one novel variant. The contribution of the GJB2 gene variants to the etiology of hearing impairment (HI) in the total sample of patients was 15.8% (26 out of 165) and significantly differed in patients of different ethnicity (5.1% in Buryat patients and 28.9% in Russian patients). In patients with DFNB1A (n = 26), HIs were congenital/early onset (92.3%), symmetric (88.5%), sensorineural (100.0%), and variable in severity (moderate-11.6%, severe-26.9% or profound-61.5%). The reconstruction of the SNP haplotypes with three frequent GJB2 pathogenic variants (c.-23+1G>A, c.35delG or c.235delC), in comparison with previously published data, supports a major role of the founder effect in the expansion of the c.-23+1G>A and c.35delG variants around the world. Comparative analysis of the haplotypes with c.235delC revealed one major haplotype G A C T (97.5%) in Eastern Asians (Chinese, Japanese and Korean patients) and two haplotypes, G A C T (71.4%) and G A C C (28.6%), in Northern Asians (Altaians, Buryats and Mongols). The variable structure of the c.235delC-haplotypes in Northern Asians requires more studies to expand our knowledge about the origin of this pathogenic variant.

Fragilaria and Ulnaria are two closely related diatom genera for which the delimitation and circumscription of several species is unclear. We studied strains isolated from Lake Baikal and compared them with the species from freshwater reservoirs in Europe and Asia using phylogenetic and species delimitation methods, microscopy and interclonal crossing experiments. The results of the phylogenetic analyses of the fragments of rbcL and 18S rRNA genes revealed that baikalian F. radians clade was independent from the representatives of the genus from other localities. Among Ulnaria we found the following 18S rRNA phylogenetic tree groups at species level: U. acus, U. ulna and U. danica. Genetic distance between genera varied between 3.9–10.2% substitutions in rbcL gene and 3.2–11.5% in 18S rRNA. The boundary between intraspecies and interspecies polymorphism for studied species of Ulnaria and Fragilaria in these marker genes was around 0.8% substitutions. Morphometric characters of individual strains showed their variability and division into F. radians, U. acus and U. ulna together with U. danica. Strains of U. acus and U. danica from different localities of Europe and Asia were sexually compatible inside the species. Sexual reproduction has never been observed in monoclonal cultures, either between this species or with strains of the Fragilaria.

The strain-induced martensitic transformation in a medical alloy from the ternary Ti–Nb–Mo system was studied. The low-doped Ti92.5Nb5Mo2.5 alloy was produced by arc remelting, followed by annealing, rolling at room temperature, reannealing, and water quenching. X-ray diffraction analysis showed that thermomechanical processing resulted in the alloy primarily consisting of orthorhombic martensite (αʺ) with a small amount of the β-titanium phase. Hysteresis loops were recorded in loading–unloading cycles with 1

The structure, mechanical characteristics, and high-temperature stability in vacuum and air of ZrB2 and HfB2-based materials sintered at a high quasi-hydrostatic pressure (4.1 GPa) under hot pressing (at a pressure of 30 MPa) with and without SiC and Si3N4 additives have been studied. It has been shown that short-term sintering (4 min) under high pressure conditions and at a comparatively low temperature (1800°C) essentially improves the mechanical properties of these materials as compared to the similar materials synthesized by the other method (hot pressing and spark-plasma sintering). In the case of sintering at a high pressure (4.1 GPa), the addition of 20 wt

Recent phylogenetic studies indicate taxonomic uncertainty of some species in the genera Arcyria (Arcyriaceae) and Trichia (Trichiaceae). For example, a phylogenetic position of Trichia brunnea still is not resolved. We revised a taxonomic position of this species based on extensively sampled 18S nrDNA sequences, as well as a morphological analysis of sporocarps and spores. The nomenclatural history of T. brunnea is briefly presented and acomprehensive morphological description of the species is provided. In result, we support the transfer of T.brunnea to Arcyria. Because of the name Arcyria brunnea exists already, we propose a new name, A. brunneo-iridescens (= Trichia brunnea).

Recent phylogenetic studies indicate taxonomic uncertainty of some species in the genera Arcyria (Arcyriaceae) and Trichia (Trichiaceae). For example, a phylogenetic position of Trichia brunnea still is not resolved. We revised a taxonomic position of this species based on extensively sampled 18S nrDNA sequences, as well as a morphological analysis of sporocarps and spores. The nomenclatural history of T. brunnea is briefly presented and acomprehensive morphological description of the species is provided. In result, we support the transfer of T.brunnea to Arcyria. Because of the name Arcyria brunnea exists already, we propose a new name, A. brunneo-iridescens (= Trichia brunnea).

Moose (Alces alces) recombinant chymosin with a milk-clotting activity of 86 AU/mL was synthesized in the Kluyveromyces lactis expression system. After precipitation with ammonium sulfate and chromatographic purification, a sample of genetically engineered moose chymosin with a specific milk-clotting activity of 15,768 AU/mg was obtained, which was used for extensive biochemical characterization of the enzyme. The threshold of the thermal stability of moose chymosin was 55 °C; its complete inactivation occurred after heating at 60 °C. The total proteolytic activity of moose chymosin was 0.332 A280 units. The ratio of milk-clotting and total proteolytic activities of the enzyme was 0.8. The Km, kcat and kcat/Km values of moose chymosin were 4.7 μM, 98.7 s−1, and 21.1 μM−1 s−1, respectively. The pattern of change in the coagulation activity as a function of pH and Ca2+ concentration was consistent with the requirements for milk coagulants for cheese making. The optimum temperature of the enzyme was 50–55 °C. The introduction of Mg2+, Zn2+, Co2+, Ba2+, Fe2+, Mn2+, Ca2+, and Cu2+ into milk activated the coagulation ability of moose chymosin, while Ni ions on the contrary inhibited its activity. Using previously published data, we compared the biochemical properties of recombinant moose chymosin produced in bacterial (Escherichia coli) and yeast (K. lactis) producers.

Recombinant chymosins (rСhns) of the cow and the camel are currently considered as standard milk coagulants for cheese-making. The search for a new type of milk-clotting enzymes that may exist in nature and can surpass the existing “cheese-making” standards is an urgent biotechnological task. Within this study, we for the first time constructed an expression vector allowing production of a recombinant analog of moose chymosin in the expression system of Escherichia coli (strain SHuffle express). We built a model of the spatial structure of moose chymosin and compared the topography of positive and negative surface charges with the correspondent structures of cow and camel chymosins. We found that the distribution of charges on the surface of moose chymosin has common features with that of cow and camel chymosins. However, the moose enzyme carries a unique positively charged patch, which is likely to affect its interaction with the substrate. Biochemical and technological properties of the moose rChn were studied. Commercial rСhns of cow and camel were used as comparison enzymes. In some technological parameters, the moose rChn proved to be superior to the reference enzymes. Сompared with the cow and camel rСhns, the moose chymosin specific activity is less dependent on the changes in CaCl2 concentration in the range of 1–5 mM and pH in the range of 6–7, which is an attractive technological property. The total proteolytic activity of the moose rСhn occupies an intermediate position between the rСhns of cow and camel. The combination of biochemical and technological properties of the moose rСhn argues for further study of this enzyme.

УкраїнаМетодою дугового перетопу одержано 8 стопів системи Ti-Nb-Mo.Показано, що вони мають типові для лиття дендритні мікроструктури.Визначено фазовий склад та встановлено періоди ґратниць утворених фаз.Використовуючи методу диференціальної термічної аналізи (ДТА) досліджено фазові перетворення у твердому стані, а також визначено температури початку топлення та кристалізації.Для стопів температура солідусу яких вище 2000°С разом з ДТА була також використана пірометрична метода Пірані-Альтермума.На основі експериментальних даних побудовані температурні залежності твердости стопів та розраховано енергії активації деформування матеріялу під індентором