Search Results (40)

Missense mutations in the BCR::ABL1 kinase domain are found in approximately 12–80% of patients with chronic myeloid leukemia (CML). Clinically significant mutations include T315I, M244V, Y253H/F, E255K/V, V299L, and F359V. The aim of this study was to create a diagnostic system for rapid and inexpensive detection of the above mutations. We used genomic DNA and RNA from peripheral blood and bone marrow cells of 57 patients with a Ph-positive CML diagnosis established in the chronic phase. We have developed a method to detect mutations in the BCR::ABL1 gene based on allele-specific real-time polymerase chain reaction (AS-PCR). In parallel, we analyzed the RNA sequence of the protein kinase domain of the same samples by next-generation sequencing (NGS) covering the points of putative mutations. In this work, we compared the results obtained by both methods for mutation detection and variant allele frequency (VAF) estimation of mutated vs. normal alleles. The sensitivity and specificity of our diagnostic system were also evaluated. It was found that AS-PCR gives reliable results at VAF up to 0.01%. AS-PCR has high sensitivity and may serve as an alternative for the more time-consuming NGS in some cases, as well as for monitoring CML treatment and for analyzing archival material. Full article

Background/Objectives: Humoral immunity directed against neuraminidase (NA) of the influenza virus may soften the severity of infection caused by new antigenic variants of the influenza viruses. Evaluation of NA-inhibiting (NI) antibodies in combination with antibodies to hemagglutinin (HA) may enhance research on the antibody response to influenza vaccines. Methods: The study examined 64 pairs of serum samples from patients vaccinated with seasonal inactivated trivalent influenza vaccines (IIVs) in 2018 according to the formula recommended by the World Health Organization (WHO) for the 2018–2019 flu season. Antibodies against drift influenza viruses A/Guangdong-Maonan/SWL1536/2019(H1N1)pdm09 and A/Brisbane/34/2018(H3N2) were studied before vaccination and 21 days after vaccination. To assess NI antibodies, we used an enzyme-linked lectin assay (ELLA) with pairs of reassortant viruses A/H6N1 and A/H6N2. Anti-HA antibodies were detected using a hemagglutination inhibition (HI) test. The microneutralization (MN) test was performed in the MDCK cell line using viruses A/H6N1 and A/H6N2. Results: Seasonal IIVs induce a significant immune response of NI antibodies against influenza A/H1N1pdm09 and A/H3N2 viruses. A significantly reduced ‘herd’ immunity to drift influenza A/H1N1pdm09 and A/H3N2 viruses was shown, compared with previously circulating strains. This reduction was most pronounced in strains possessing neuraminidase N2. Seasonal IIVs caused an increase in antibodies against homologous and drifted viruses; however, an increase in antibodies to drifting viruses was observed more often among older patients. The level of NI antibodies for later A/H1N1pdm09 virus in response to IIVs was statistically significantly lower among younger people. After IIV vaccination, the percentage of individuals with HI antibody levels ≥ 1:40 and NI antibody levels ≥ 1:20 was 32.8% for drift A/H1N1pdm09 virus and 17.2% for drift A/H3N2 virus. Antisera containing HI and NI antibodies exhibited neutralizing properties in vitro against viruses with unrelated HA of the H6 subtype. Conclusions: Drift A/H1N1pdm09 and A/H3N2 viruses demonstrated significantly lower reactivity to HI and NI antibodies against early influenza viruses. In response to seasonal IIVs, the level of seroprotection has increased, including against drift influenza A viruses, but protective antibody levels against A/H1N1pdm09 have risen to a greater extent. A reduced immune response to the N1 protein of the A/H1N1pdm09 drift virus was obtained in individuals under 60 years of age. Based on our findings, it is hypothesized that in the cases of a HA mismatch, vaccination against N1-containing influenza viruses may be necessary for individuals under 60, while broader population-level vaccination against N2-containing viruses may be required. Full article

Background/Objectives: Influenza viruses and SARS-CoV-2 are currently cocirculating with similar seasonality, and both pathogens are characterized by a high mutational rate which results in reduced vaccine effectiveness and thus requires regular updating of vaccine compositions. Vaccine formulations combining seasonal influenza and SARS-CoV-2 strains can be considered promising and cost-effective tools for protection against both infections. Methods: We used a licensed seasonal trivalent live attenuated influenza vaccine (3×LAIV) as a basis for the development of a modified 3×LAIV/CoV-2 vaccine, where H1N1 and H3N2 LAIV strains encoded an immunogenic cassette enriched with conserved T-cell epitopes of SARS-CoV-2, whereas a B/Victoria lineage LAIV strain was unmodified. The trivalent LAIV/CoV-2 composition was compared to the classical 3×LAIV in the golden Syrian hamster model. Animals were intranasally immunized with the mixtures of the vaccine viruses, twice, with a 3-week interval. Immunogenicity was assessed on day 42 of the study, and the protective effect was established by infecting vaccinated hamsters with either influenza H1N1, H3N2 or B viruses or with SARS-CoV-2 strains of the Wuhan, Delta and Omicron lineages. Results: Both the classical 3×LAIV and 3×LAIV/CoV-2 vaccine compositions induced similar levels of serum antibodies specific to all three influenza strains, which resulted in comparable levels of protection against challenge from either influenza strain. Protection against SARS-CoV-2 challenge was more pronounced in the 3×LAIV/CoV-2-immunized hamsters compared to the classical 3×LAIV group. These data were accompanied by the higher magnitude of virus-specific cellular responses detected by ELISPOT in the modified trivalent LAIV group. Conclusions: The modified trivalent live attenuated influenza vaccine encoding the T-cell epitopes of SARS-CoV-2 can be considered a promising tool for combined protection against seasonal influenza and COVID-19. Full article

Background. Influenza and SARS-CoV-2 viruses are two highly variable pathogens. We have developed a candidate bivalent live vaccine based on the strain of licensed A/Leningrad/17-based cold-adapted live attenuated influenza vaccine (LAIV) of H3N2 subtype, which expressed SARS-CoV-2 immunogenic T-cell epitopes. A cassette encoding fragments of S and N proteins of SARS-CoV-2 was inserted into the influenza NA gene using the P2A autocleavage site. In this study, we present the results of preclinical evaluation of the developed bivalent vaccine in a non-human primate model. Methods. Rhesus macaques (Macaca mulatta) (n = 3 per group) were immunized intranasally with 7.5 lg EID₅₀ of the LAIV/CoV-2 bivalent vaccine, a control non-modified H3N2 LAIV or a placebo (chorioallantoic fluid) using a sprayer device, twice, with a 28-day interval. The blood samples were collected at days 0, 3, 28 and 35 for hematological and biochemical assessment. Safety was also assessed by monitoring body weight, body temperature and clinical signs of the disease. Immune responses to influenza virus were assessed both by determining serum antibody titers in hemagglutination inhibition assay, microneutralization assay and IgG ELISA. T-cell responses were measured both to influenza and SARS-CoV-2 antigens using ELISPOT and flow cytometry. Three weeks after the second immunization, animals were challenged with 10⁵ PFU of Delta SARS-CoV-2. The body temperature, weight and challenge virus shedding were monitored for 5 days post-challenge. In addition, virus titers in various organs and histopathology were evaluated on day 6 after SARS-CoV-2 infection. Results. There was no toxic effect of the immunizations on the hematological and coagulation hemostasis of animals. No difference in the dynamics of the average weight and thermometry results were found between the groups of animals. Both LAIV and LAIV/CoV-2 variants poorly replicated in the upper respiratory tract of rhesus macaques. Nevertheless, despite this low level of virus shedding, influenza-specific serum IgG responses were detected in the group of monkeys immunized with the LAIV/CoV-2 bivalent but not in the LAIV group. Furthermore, T-cell responses to both influenza and SARS-CoV-2 viruses were detected in the LAIV/CoV-2 vaccine group only. The animals were generally resistant to SARS-CoV-2 challenge, with minimal virus shedding in the placebo and LAIV groups. Histopathological changes in vaccinated animals were decreased compared to the PBS group, suggesting a protective effect of the chimeric vaccine candidate. Conclusions. The candidate bivalent vaccine was safe and immunogenic for non-human primates and warrants its further evaluation in clinical trials. Full article

Duchenne muscular dystrophy is secondarily accompanied by Ca²⁺ excess in muscle fibers. Part of the Ca²⁺ accumulates in the mitochondria, contributing to the development of mitochondrial dysfunction and degeneration of muscles. In this work, we assessed the effect of intraperitoneal administration of rhodacyanine MKT077 (5 mg/kg/day), which is able to suppress glucose-regulated protein 75 (GRP75)-mediated Ca²⁺ transfer from the sarcoplasmic reticulum (SR) to mitochondria, on the Ca²⁺ overload of skeletal muscle mitochondria in dystrophin-deficient mdx mice and the concomitant mitochondrial dysfunction contributing to muscle pathology. MKT077 prevented Ca²⁺ overload of quadriceps mitochondria in mdx mice, reduced the intensity of oxidative stress, and improved mitochondrial ultrastructure, but had no effect on impaired oxidative phosphorylation. MKT077 eliminated quadriceps calcification and reduced the intensity of muscle fiber degeneration, fibrosis level, and normalized grip strength in mdx mice. However, we noted a negative effect of MKT077 on wild-type mice, expressed as a decrease in the efficiency of mitochondrial oxidative phosphorylation, SR stress development, ultrastructural disturbances in the quadriceps, and a reduction in animal endurance in the wire-hanging test. This paper discusses the impact of MKT077 modulation of mitochondrial dysfunction on the development of skeletal muscle pathology in mdx mice. Full article

Zr/Nb nanoscale multilayer coatings (NMCs) were studied after hydrogenation in a gaseous environment at 400 °C. The hydrogen distribution and content were determined by pressure and hydrogenation time. Increasing the pressure from 0.2 to 2 MPa resulted in different hydrogen distribution within the Zr/Nb NMCs, while the concentration remained constant at 0.0150 ± 0.0015 wt. %. The hydrogen concentration increased from 0.0165 ± 0.001 to 0.0370 ± 0.0015 wt. % when the hydrogenation time was extended from 1 to 7 h. The δ-ZrH hydride phase was formed in the Zr layers with Zr crystals reorienting towards the [100] direction. The Nb(110) diffraction reflex shifted towards smaller angles and the interplanar distance in the niobium layers increased, indicating significant lateral compressive stresses. Despite an increase in pressure, the nanohardness and Young’s modulus of the Zr/Nb NMCs remained stable. Increasing the hydrogen concentration to 0.0370 ± 0.0015 wt. % resulted in a 40% increase in nanohardness. At this concentration, the relative values of the Doppler broadening variable energy positron annihilation spectroscopy (S/S₀) increased above the initial level, indicating an increase in excess free volume due to hydrogen-induced defects and changes. However, the predominant positron capture center remained intact. The Zr/Nb NMCs with hydrogen content ranging from 0.0150 ± 0.0015 to 0.0180 ± 0.001 wt. % exhibited a decrease in the free volume probed by positrons, as demonstrated by the Doppler broadening variable energy positron annihilation spectroscopy. This was evidenced by opposite changes in S and W (S↓W↑). The microstructural changes are attributed to defect annihilation during hydrogen accumulation near interfaces with the formation of hydrogen–vacancy clusters and hydrides. Full article

Background: Influenza viruses continue to cause a significant social and economic burden globally. Vaccination is recognized as the most effective measure to control influenza. Live attenuated influenza vaccines (LAIVs) are an effective means of preventing influenza, especially among children. A reverse genetics (RG) system is required to rapidly update the antigenic composition of vaccines, as well as to design LAIVs with a broader spectrum of protection. Such a system has been developed for the Russian LAIVs only for type A strains, but not for influenza B viruses (IBV). Methods: All genes of the B/USSR/60/69 master donor virus (B60) were cloned into RG plasmids, and the engineered B60, as well as a panel of IBV LAIV reassortants were rescued from plasmid DNAs encoding all viral genes. The engineered viruses were evaluated in vitro and in a mouse model. Results: The B60 RG system was successfully developed, which made it possible to rescue LAIV reassortants with the desired antigenic composition, including hybrid strains with hemagglutinin and neuraminidase genes belonging to the viruses from different IBV lineages. The LAIV candidate carrying the HA of the B/Victoria-lineage virus and NA from the B/Yamagata-lineage virus demonstrated optimal characteristics in terms of safety, immunogenicity and cross-protection, prompting its further assessment as a broadly protective component of trivalent LAIV. Conclusions: The new RG system for B60 MDV allowed the rapid generation of type B LAIV reassortants with desired genome compositions. The generation of hybrid LAIV reassortants with HA and NA genes belonging to the opposite IBV lineages is a promising approach for the development of IBV vaccines with broad cross-protection. Full article

Reconstructing interglacial marine environments helps us understand the climate change mechanisms of the past. To contribute to this body of knowledge, we studied a high-resolution 455 cm-thick sediment sequence of the Boreal (Eemian) marine beds directly overlying Moscovian (Saalian) moraine in the Bychye-2 section on the Pyoza River. We analyzed lithological and microfossil (foraminifers, ostracods, pollen, aquatic palynomorphs) variations at the studied site. Stratigraphical zonation is based on the local and well-established regional pollen zones, correlated with the western European pollen zones. The studied marine beds accumulated from the end of the Moscovian glacial (>131 ka) until ca. 119.5 ka. We distinguished three successive phases: a seasonally sea-ice-covered, relatively deep, freshened basin in the initial rapid flooding stage (>131–130.5 ka); a deep basin in the maximum flooding phase with less extensive sea ice cover (130.5–130.25 ka); and a shallow basin with reduced sea ice cover (130.25–119.5 ka). According to a pollen zone comparison with other sites, the regional glacioisostatic rebound started ca. 130 ka. The diverse warm-water assemblages of benthic foraminifers and ostracods containing typical Baltic Sea species occurred during the regression, mainly 128–124 ka, thus giving evidence for a relatively long-lasting connection between the White and Baltic Seas. Full article

Influenza virus strain A/South Africa/3626/2013 (H1N1)pdm09 (SA-WT) is a non-mouse-adapted model strain that has naturally high pathogenic properties in mice. It has been suggested that the high pathogenicity of this strain for mice could be due to the three strain-specific substitutions in the polymerase complex (Q687R in PB1, N102T in PB2, and E358E/K heterogeneity in PB2). To evaluate the role of these replacements, SA-WT was passaged five times in mouse lungs, and the genome of the mouse-adapted version of the SA-WT strain (SA-M5) was sequenced. SA-M5 lost E358E/K heterogeneity and retained E358, which is the prevalent amino acid at this position among H1N1pdm09 strains. In addition, in the hemagglutinin of SA-M5, two heterogeneous substitutions (G155G/E and S190S/R) were identified. Both viruses, SA-M5 and SA-WT, were compared for their toxicity, ability to replicate, pathogenicity, and immunogenicity in mice. In mice infected with SA-M5 or SA-WT strains, toxicity, virus titer in pulmonary homogenates, and mouse survival did not differ significantly. In contrast, an increase in the immunogenicity of SA-M5 compared to SA-WT was observed. This increase could be due to the substitutions G155G/E and S190S/R in the HA of SA-M5. The prospects for using SA-M5 in studying the immunogenicity mechanisms were also discussed. Full article

Cryopreservation is one way to preserve rare, endangered species. However, during the cryopreservation process, plant cells undergo considerable stress, which may lead to cell death. In our work, orthodox Stipa seeds of six rare species were cryopreserved: S. sareptana, S. ucrainica, S. tirsa, S. dasyphylla, S. adoxa, and S. pulcherríma. Short-term cryopreservation (14 days) stimulated germination of all Stipa species studied. Prolonged cryopreservation (70 days and more) decreased the germination of all Stipa seeds except S. sareptana. The decrease in germination progressed over time as a result of the cumulative stress of cryopreservation rather than the initial stress. To stimulate germination, seeds were stratified and treated with GA₃, KNO₃, NaOH, and H₂O₂. After four years of seed cryopreservation, it was possible to obtain seedlings of all the Stipa species studied with 30 days of stratification and 180 days of germination. After five years of cryopreservation and seed treatment with 30% NaOH for one hour, the best germination was obtained in S. adoxa and S. pulcherrima. After treatment with 5% H₂O₂ for 20 min, the best germination was obtained in S. sareptana, S. ucrainica, and S. dasyphylla. S. sareptana seeds germinated in all the aforementioned experiments. S. sareptana has a non-deep physiological dormancy and is the most widespread and drought-tolerant Stipa species studied. The best habitat adaptation and stress tolerance correlated with this species’cryotolerance. S. sareptana was recommended for further cryopreservation, while storage protocols for the other Stipa species studied need further improvements. Full article

Pathogenic CFTR variants cause cystic fibrosis (CF), and CF-related disorders (CF-RD), including bilateral aplasia of the vas deferens (CBAVD). The spectrum of clinical manifestations depends on the CFTR genotype. The frequency and spectrum of the CFTR variants vary between populations and clinical groups. CFTR variants and genotypes were analyzed in Russian men with CF (n = 546) and CBAVD syndrome (n = 125). Pathogenic variants were detected in 93.95% and 39.2% of the CF and CBAVD alleles, respectively. The most frequent c.1521_1523del (F508del; p.Phe508del) variant was found in 541 (49.5%) CF alleles. A total of 162 CFTR genotypes were revealed in CF patients, including 152 homozygous and 394 compound-heterozygous. The most common CF-genotype was F508del/F508del (24.9%). Other frequent CF-genotypes were F508del/3849+10kbC>T, F508del/CFTRdele2,3, and F508del/E92K. CF-causing variants and/or 5T allele were found in 88% of CBAVD patients: 5T/CFTRmut (48.0%), CFTRmut/N (17.6%), CFTRmut/CFTRmut (6.4%), 5T/5T (10.4%), 5T/N (5.6%) and N/N (12.0%), with the most common CBAVD-genotype being F508del/5T (29.6%). The allele frequencies of F508del, CFTRdele2,3 394delTT, and 3849+10kbC>T were significantly higher in CF patients. L138ins/L138ins, 2184insA/E92K, and L138ins/N genotypes were found in CBAVD, but not in CF patients. The results indicate certain differences in the frequency of some CFTR variants and genotypes in Russian CF and CBAVD patients. Full article

Inherited retinal diseases (IRDs) constitute a prevalent group of inherited ocular disorders characterized by marked genetic diversity alongside moderate clinical variability. Among these, ABCA4-related eye pathology stands as a prominent form affecting the retina. In this study, we conducted an in-depth analysis of 96 patients harboring ABCA4 variants in the European part of Russia. Notably, the complex allele c.[1622T>C;3113C>T] (p.Leu541Pro;Ala1038Val, or L541P;A1038V) and the variant c.5882G>A (p.Gly1961Glu or G1961E) emerged as primary contributors to this ocular pathology within this population. Additionally, we elucidated distinct disease progression characteristics associated with the G1961E variant. Furthermore, our investigation revealed that patients with loss-of-function variants in ABCA4 were more inclined to develop phenotypes distinct from Stargardt disease. These findings provide crucial insights into the genetic and clinical landscape of ABCA4-related retinal dystrophies in this specific population. Full article

Currently, more than 500,000 cases of various helminthes in humans are reported annually in the Russian Federation. This figure may not reflect the true incidence of helminthes, as only nine separate nosological forms are compulsory notifiable. The rest of the species of detected helminthes are included in a separate category of “other helminthes” or “rare helminthes”. The bulk of the latter is represented by the helminthes with a rate of incidence that does not exceed one case per 100,000 people. This review is based on data derived from publications in the Russian language, both from the Russian Federation and international, as well as data available from various health treatment facilities in Russia. These data largely cover the period of the 1990s–2010s. A total of 15 species of “rare helminthes” are described in this review: anisakiosis, capillariosis, clonorchosis, dioctophymosis, dipylidiosis, echinochasmosis, fasciolosis, gastrodiscoidosis (amphistomiosis), metagonimosis, metorchiosis, nanophyetosis, pseudamphistomosis, sparganosis (spirometrosis), strongyloidosis and trichostrongylosis. Details of their geographical distribution, clinical and epidemiological peculiarities, and the difficulties they pose in diagnosis are provided. The public health importance of “rare helminthes” in Russia at present and in the forthcoming years is stressed. Full article

(1) Introduction: Pathogenic variants in the CFTR (Cystic Fibrosis Transmembrane conductance Regulator, OMIM: 602421) gene cause Cystic Fibrosis (CF, OMIM: 219700) and CF-related disorders (CF-RD), often accompanied by obstructive azoospermia due to congenital bilateral aplasia of vas deferens (CBAVD, OMIM: 277180) in male patients. The L138ins (c.413_415dup; p. (Leu138dup)) is a mild variant in the CFTR gene that is relatively common among CF-patients in Slavic populations. The frequency of this variant in Russian infertile men has not been sufficiently studied; (2) Materials and Methods: The sample consisted of 6033 Russian infertile men. The patients were tested for 22 common in Russian populations pathogenic variants of the CFTR gene and the IVS9Tn-polymorphic locus of the intron 9. Molecular-genetic studies were performed using amplified fragment length polymorphism (AFLP-PCR), multiplex ligation-dependent probe amplification (MLPA), and nested PCR (for analysis of the IVS9Tn-polymorphic locus); (3) Results: Pathogenic variants in the CFTR were detected in 3.9% of patients. The most frequent variants were F508del and CFTRdele2.3(21kb), accounted for 61.0% and 7.1% of detected variants, respectively. The L138ins variant was detected in 17 (0.28%) individuals: one of them was homozygous, 10 patients were heterozygous, and 6 patients were compound-heterozygous (F508del/L138ins, n = 4; L138ins/N1303K, n = 1; L138ins/5T, n = 1). Two pathogenic CF-causing variants in the CFTR gene were detected in 8 patients, including 7 compound heterozygous (F508del/L138ins, n = 4; F508del/N1303K, n = 1; 2184insA/E92K, n = 1; 3849+10kbC>T/E92K, n = 1) and one homozygous (L138ins/L138ins). The L138ins variant was found in 7 out of 16 (43.75%) chromosomes in six of these patients. The most common pathogenic variant, F508del, was identified in five out of them, in 5 of 16 (31.25%) chromosomes. The allele frequency (AF) of the L138ins variant in the sample has been found to be 0.0014.; (4) Conclusions: The L138ins variant of the CFTR gene is the third most common variant after F508del and CFTRdele2.3(kb) among Russian infertile men. Full article

The effect of proton irradiation on the structure, phase composition, defect state and nanohardness of Zr/Nb nanoscale multilayer coatings was investigated. Preservation of the Zr/Nb layered structure with 50 and 100 nm thick layers, was observed after irradiation with protons at 1720 keV energy and 3.4 × 10¹⁵, 8.6 × 10¹⁵ and 3.4 × 10¹⁶ ions/cm² fluences, and the interfaces remained incoherent. In the Zr/Nb nanoscale multilayer coatings with individual layer thicknesses of 10 and 25 nm, there were insignificant fluctuations in interplanar distance, which were influenced by changes in irradiation fluence, and the interfaces were partially destroyed and became semicoherent. Changing irradiation fluence in the investigated ranges led to a decrease in the nanohardness of the Zr/Nb nanoscale multilayer coatings with individual layer thicknesses of 10–50 nm. Variable-energy positron Doppler broadening analysis revealed that these changes are primarily caused by peculiarities of the localization and accumulation of the embedded ions and do not cause a significant increase in the S-parameters of Zr/Nb nanoscale multilayer coatings with a layer thickness less than 100 nm. Full article