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The high specificity of human antibodies to blood group A and B antigens is impressive, especially when considering the structural difference between these antigens (tetrasaccharides) is a NHAc versus a hydroxyl group on the terminal monosaccharide residue. It is well established that in addition to anti-A and anti-B there is a third antibody, anti-A,B capable of recognizing both A and B antigens. To analyze this AB specificity, we synthesized a tetrasaccharide, where the NHAc of the A antigen was replaced with an NH2. This NH2 group was then used to attach the glycan to an affinity resin, creating an AB epitope (ABep) adsorbent where the critical site for recognition by A and B antibodies was not accessible, while the rest of the (conformationally compact) tetrasaccharide remained accessible. Anti-ABep antibodies were then isolated from blood group O donors and found to have expected A,B specificity against immobilized and red cell bound synthetic antigens, including ABep, and were able to agglutinate both A and B red cells. The amount of these anti-ABep (anti-A,B) antibodies found in the blood of group O donors was comparable to levels of anti-A and anti-B found in group B and A individuals. Using STD-NMR the location for the AB epitope on the tetrasaccharide was found.
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The North Caucasus played a key role during the ancient colonization of Eurasia and the formation of its cultural and genetic ancestry. Previous archeogenetic studies described a relative genetic and cultural continuity of ancient Caucasus societies, since the Eneolithic period. The Koban culture, which formed in the Late Bronze Age on the North Caucasian highlands, is considered as a cultural "bridge" between the ancient and modern autochthonous peoples of the Caucasus. Here, we discuss the place of this archeological culture and its representatives in the genetic orbit of Caucasian cultures using genome-wide SNP data from five individuals of the Koban culture and one individual of the early Alanic culture as well as previously published genomic data of ancient and modern North Caucasus individuals. Ancient DNA analysis shows that an ancient individual from Klin-Yar III, who was previously described as male, was in fact a female. Additional studies on well-preserved ancient human specimens are necessary to determine the level of local mobility and kinship between individuals in ancient societies of North Caucasus. Further studies with a larger sample size will allow us gain a deeper understanding of this topic.
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Virus diseases affect the yield and fruit quality and shorten the productive life of stone fruits (Prunus spp. in the family Rosaceae). Of over fifty known viruses infecting these crops, cherry virus A (CVA) is among the most common, and little cherry virus 1 (LChV1) is one of the most economically important. Using high-throughput sequencing, full-length genomes of CVA and LChV1 isolates, found on interspecies hybrids in the Prunus collection of the Nikita Botanical Gardens, Russia, were sequenced, assembled, and characterized. CVA was found in the P. cerasifera × P. armeniaca hybrid and in phylogenetic analysis clustered with non-cherry virus isolates. The LChV1 isolate Stepnoe was detected in ((P. cerasifera Ehrh. × P. armeniaca L.) × P. brigantiaca Vill.) trihybrid suggesting that both P. cerasifera and P. brigantiaca potentially can be the LChV1 hosts. The isolate Stepnoe was most closely related to the Greece isolate G15_3 from sweet cherry, sharing 77.3% identity at the nucleotide level. Possibly, the highly divergent Russian isolate represents one more phylogroup of this virus. This is the first report of CVA and LChV1 from Russia, expanding the information on their geographical distribution and genetic diversity.
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In different countries, interest in the commercial cultivation of the olive has recently greatly increased, which has led to the expansion of its range. The Crimean Peninsula is the northern limit of the common olive (Olea europaea L.) range. A unique collection of common olive's cultivars and hybrids has been collected in the Nikitsky Botanical Gardens (NBG). The aim of this study was to assess the genetic diversity of 151 samples (total of several biological replicates of 46 olive cultivars including 29 introduced and 11 indigenous genotypes) using the ddRAD sequencing method. Structural analysis showed that the studied samples are divided into ten groups, each of which mainly includes cultivars of the same origin. Cultivars introduced to the Crimean Peninsula from different regions formed separate groups, while local cultivars joined different groups depending on their origin. Cultivars of Crimean origin contain admixtures of mainly Italian and Caucasian cultivars' genotypes. Our study showed that the significant number of Crimean cultivars contains an admixture of the Italian cultivar "Coreggiolo". Genetic analysis confirmed the synonymy for the cv. "Otur" and "Nikitskaya 2", but not for the other four putative synonyms. Our results revealed the genetic diversity of the olive collection of NBG and provided references for future research studies, especially in selection studies for breeding programs.
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Olea , Olea/genética , Repeticiones de Microsatélite/genética , Fitomejoramiento , Genotipo , Variación GenéticaRESUMEN
Paleoclimatic changes during the Pleistocene-Holocene transition is suggested as a main factor that led to species extinction, including the woolly mammoth (Mammuthus primigenius), Steller's sea cow (Hydrodamalis gigas) and the Don-hare (Lepus tanaiticus). These species inhabited the territory of Eurasia during the Holocene, but eventually went extinct. The Don-hare is an extinct species of the genus Lepus (Leporidae, Lagomorpha), which lived in the Late Pleistocene-Early Holocene in Eastern Europe and Northern Asia. For a long time, the Don-hare was considered a separate species, but at the same time, its species status was disputed, taking into account both morphological data and mitochondrial DNA. In this study, mitochondrial genomes of five Don-hares, whose remains were found on the territory of Northeastern Eurasia were reconstructed. Firstly, we confirm the phylogenetic proximity of the "young" specimens of Don-hare and mountain or white hare, and secondly, that samples older than 39 Kya form a completely distinct mitochondrial clade.
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Liebres , Lagomorpha , Animales , Femenino , Bovinos , Liebres/genética , Filogenia , ADN Antiguo , Lagomorpha/genética , AsiaRESUMEN
Significant palaeoecological and paleoclimatic changes that took place during Late Pleistocene-Early Holocene transition are considered important factors that led to megafauna extinctions. Unlike many other species, the brown bear (Ursus arctos) has survived this geological time. Despite the fact that several mitochondrial DNA clades of brown bears became extinct at the end of the Pleistocene, this species is still widely distributed in Northeast Eurasia. Here, using the ancient DNA analysis of a brown bear individual that inhabited Northeast Asia in the Middle Holocene (3460 ± 40 years BP) and comparative phylogenetic analysis, we show a significant mitochondrial DNA similarity of the studied specimen with modern brown bears inhabiting Yakutia and Chukotka. In this study, we clearly demonstrate the maternal philopatry of the Northeastern Eurasian U. arctos population during the several thousand years of the Holocene.
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Ursidae , Animales , Ursidae/genética , ADN Antiguo , Filogenia , ADN Mitocondrial/genética , Mitocondrias/genéticaRESUMEN
Fig mosaic disease is spread worldwide and is believed to have a viral etiology. Divergent isolates of grapevine badnavirus 1 (GBV1), named fGBV1, were discovered on Ficus carica, F. palmata, F. virgata, and F. afghanistanica in the fig germplasm collection of the Nikita Botanical Gardens, Russia, expanding the list of viruses infecting this crop. The complete genomes of five fGBV1 isolates from F. carica and F. palmata trees were determined using high-throughput and Sanger sequencing. The genomes comprised 7283 base pairs, contained four overlapping open reading frames, were 99.7 to 99.9% identical to each other, and related to GBV1 (83.2% identity). The reverse transcriptase RNase H genome regions of fGBV1 and GBV1 share 84.6% identity, indicating that fGBV1 is a divergent isolate of GBV1, which was found on the new natural hosts from a different family (Moraceae). Further, fGBV1-specific primers were developed to detect the virus using RT-PCR. Survey of 47 trees, belonging to four fig species and 14 local and introduced F. carica cultivars, showed the high fGBV1 prevalence in the collection (93.6%), including trees with no obvious symptoms of fig mosaic disease.
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Interspecific hybridization has occurred relatively frequently during the evolution of vertebrates. This process usually abolishes reproductive isolation between the parental species. Moreover, it results in the exchange of genetic material and can lead to hybridogenic speciation. Hybridization between species has predominately been observed at the interspecific level, whereas intergeneric hybridization is rarer. Here, using whole-genome sequencing analysis, we describe clear and reliable signals of intergeneric introgression between the three-spined stickleback (Gasterosteus aculeatus) and its distant mostly freshwater relative the nine-spined stickleback (Pungitius pungitius) that inhabit northwestern Russia. Through comparative analysis, we demonstrate that such introgression phenomena apparently take place in the moderate-salinity White Sea basin, although it is not detected in Japanese sea stickleback populations. Bioinformatical analysis of the sites influenced by introgression showed that they are located near transposable elements, whereas those in protein-coding sequences are mostly found in membrane-associated and alternative splicing-related genes.
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Prunus persica is one of the main stone fruit crops in Crimea and southern Russia. The P. persica genome has recently been sequenced and annotated in good quality. However, for a deeper assessment of the peach genome, it is necessary to include in the research other cultivars that are in the collection of the Nikitsky Botanical Garden. The cultivars of the Nikitsky Botanical Garden are unique and differ from Western European and American ones, as they are derived from cultivars and forms originating from Central Asian, North Caucasian, Transcaucasian and Eastern European countries. In this paper, we present the assembly of the P. persica cv. 'Sovetskiy' genome obtained using Oxford Nanopore long reads and Illumina short reads by hybrid assembly methods. The assembled genome of P. persica cv. 'Sovetskiy' is 206.26 MB in 226 scaffolds, with N50 24 Mb, including 8 chromosomes. It contains 27140 coding genes, 26973 (99.38%) of which are annotated in at least one functional database. More than 36.05% of the genome regions were identified as repeating elements.
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Nanoporos , Prunus persica , Secuencia de Bases , Genoma , Secuenciación de Nucleótidos de Alto Rendimiento , Prunus persica/genéticaRESUMEN
Diffuse large B-cell lymphoma (DLBCL) is the most common lymphoid tumor among other non-Hodgkin lymphomas (30-40% of all cases). This type of lymphoma is characterized by significant differences in treatment response and the heterogeneity of clinical traits. Approximately 60% of patients are cured using standard chemotherapy (CT), while in 10-15% of cases, the tumor is characterized by an extremely aggressive course and resistance to even the most high-dose programs with autologous stem cell transplantation (auto-SCT). The activated B-cell (ABC) subtype of DLBCL is characterized by poor prognosis. Here, we describe a clinical case of diffuse ABC-DLBCL with an atypical disease course. Complete remission was achieved after four courses of CT, followed by autologous hematopoietic stem cell transplantation (auto-HSCT). However, early relapse occurred 2 months after the completion of treatment. According to the results of cytogenetic studies, significant chromosome breakdowns were observed. Exome sequencing allowed for the detection of several novel mutations that affect components of the NOTCH2 and NF-κB signaling pathways, a number of epigenetic regulators (KMT2D, CREBBP, EP300, ARID1A, MEF2B), as well as members of the immunoglobulin superfamily (CD58 and CD70). Whether these mutations were the result of therapy or were originally present in the lymphoid tumor remains unclear. Nevertheless, the introduction of genomic technologies into clinical practice is important for making a diagnosis and developing a DLBCL treatment regimen with the use of targeted drugs.
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The peach (Prunus persica L. Batsch) is one of the important stone fruit crops in the Crimea Peninsula and the southern part of Russia. The complete chloroplast genome of the peach cultivar 'Sovetskiy' is published in this paper. The chloroplast genome size is 157,756 bp. It contains 126 genes, including 81 protein-coding genes (PCGs), eight ribosomal RNA (rRNA) genes, and 37 transfer RNA (tRNA) genes. The chloroplast genome also contains a large single-copy region of 85,960 bp, a small single-copy (SSC) region of 19,045 bp, and two inverted repeats regions of 26,375 bp and 26,372 bp. The overall base composition of the genome in descending order is 31.2% - A, 32.1% - T, 18.7% - C, and 18.0% - G. The total GC content of the chloroplast genome is 36.7%. Maximum-likelihood phylogenetic analysis involving nine chloroplast genomes of the Prunus genus revealed a separate cluster for P. persica and its possible landrace - P. ferganensis.
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Anthropogenic activity is the top factor directly related to the extinction of several animal species. The last Steller's sea cow (Hydrodamalis gigas) population on the Commander Islands (Russia) was wiped out in the second half of the 18th century due to sailors and fur traders hunting it for the meat and fat. However, new data suggests that the extinction process of this species began much earlier. Here, we present a nuclear de novo assembled genome of H. gigas with a 25.4× depth coverage. Our results demonstrate that the heterozygosity of the last population of this animal is low and comparable to the last woolly mammoth population that inhabited Wrangel Island 4000 years ago. Besides, as a matter of consideration, our findings also demonstrate that the extinction of this marine mammal starts along the North Pacific coastal line much earlier than the first Paleolithic humans arrived in the Bering sea region.
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Dugong/genética , Genoma , Animales , ADN Mitocondrial/genética , Dugong/clasificación , Extinción Biológica , Humanos , Mutación , Filogenia , Polimorfismo de Nucleótido Simple , Federación de Rusia , Análisis de Secuencia de ADNRESUMEN
Interspecies hybridization is driven by a complex interplay of factors where introgression plays an important role. In the present study, the transfer of genetic material, between two quite distant fish species from different genera, through spontaneous hybridization was documented with dedicated molecular and bioinformatics tools. We investigate the genomic landscape of putative stickleback-relative introgression by carefully analyzing the tractable transposable elements (TE) on the admixed genome of some individuals of two sympatric stickleback species inhabiting northwestern Russia, namely the three-spined (Gasterosteus aculeatus) and the nine-spined (Pungitius pungitius) sticklebacks. Our data revealed that unique TE amplification types exist, supporting our proposed hypothesis that infers on the interspecific introgression. By running a restriction site-associated DNA sequencing (RAD-Seq) with eight samples of G. aculeatus and P. pungitius and subjecting further the results to a contrasting analysis by variated bioinformatic tools, we identified the related introgression-linked markers. The admixture nature observed in a single sample of the nine-spined stickleback demonstrated the possible traces of remote introgression between these two species. Our work reveals the potential that introgression has on providing particular variants at a high-frequency speed while linking blocks of sequence with multiple functional mutations. However, even though our results are of significant interest, an increased number of samples displaying the introgression are required to further ascertain our conclusions.
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Fig mosaic virus (FMV) (genus Emaravirus in the family Fimoviridae) is considered the etiological agent of fig mosaic disease (FMD) that is recorded in most of the fig growing areas with an average global infection rate of 33%. The multipartite FMV genome is comprised of six negative monocistronic ssRNAs, each of which is separately encapsidated (Preising et al. 2020). Although FMD-like symptoms, which include mosaic, chlorotic ringspots, and oak leaf patterns, were observed in approximately a third of 400 fig accessions in the Nikita Botanical Gardens, Yalta, Russia (Mitrofanova et al. 2016), FMV has not been identified as the causal agent of the disease. In June of 2020, total RNA was isolated from symptomatic leaves of 59 thirty two-year-old trees representing 31 local and 27 introduced Ficus carica L. cultivars and a single F. pseudocarica Miq. tree using RNeasy Plant Mini kit (Qiagen, USA). FMV was tested by RT-PCR using primer sets E5 (Elbeaino et al. 2009) and EMARAVGP (Walia et al. 2009), which amplify a 302-bp fragment of RNA1 and a 468-bp fragment of RNA2, respectively. PCR products of the expected sizes were generated in all samples, indicating a high FMV incidence in the plantings. The genome sequences of FMV isolates from F. carica cvs. Bleuet, Kraps di Hersh, Smena, Temri, and F. pseudocarica (Fig. S1) were determined by high-throughput sequencing on MiSec Illumina platform. Double-stranded RNA was isolated from FMV-positive leaves using Viral Gene-spin™ Viral DNA/RNA Extraction Kit (iNtRON, Korea), followed by cDNA library preparation with the NEBNext® Ultra™ II RNA Library Prep Kit (New England Biolabs, USA). In average, 695,000 quality-filtered 150 bp pair-ended reads per a library were produced and used in a de novo assembly using metaSpades program version 3.14 (Nurk et al. 2017). In each of five samples, BLASTn analysis found six FMV-related contigs. The contigs spanned 99 to 100% of corresponding genomic segments of the most closely related isolates. In addition to FMV, fig cryptic virus-related contigs were also detected in some samples. The FMV contigs covering RNA1 to RNA6 had the highest identity to corresponding genomic segments of isolates AM941711 (96.5 to 96.6%), FM864225 (94.4 to 94.6%), FM991954 (97.9 to 98.2%), AB697863 (96.4 to 96.6%), AB697879 (93.3 to 93.4%), and AB697895 (95.4 to 97.0%), respectively. Five Russian isolates shared 99.2 to 100% nucleotide sequence identity, depending on the genomic segment. Their sequences were deposited in GenBank under accession numbers MW201216 to MW201230 and MW208662 to MW208676. Phylogenetic analysis of six ORFs showed that ORF1 to ORF3 and ORF6 of the Russian isolates clustered with FMV isolates from Italy while ORF4 grouped with the isolate JTT-Pa (AB697863) from Japan (Fig. S2). ORF5 of the Russian isolates formed a separate cluster with the isolates SB1 and SB2 from Serbia and JTT-Vi from Japan (AB697879 to AB697884). Incongruency of phylogenetic relationship among the genomic segments suggests reassortment among ancestors of the Russian FMV isolates. In addition, similar to the SB1, SB2 and JTT-Vi, ORF5 of the Russian isolates encodes a protein of 486 amino acid (aa) residues in contrast to the corresponding protein of Italian isolates consisting of 502 aa. To the best of our knowledge, this is the first report of FMV in Russia. This finding not only expands the information on the geographical distribution of FMV, but also extends knowledge on F. pseudocarica as a natural host of the virus.
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BACKGROUND: The α1,3-galactosyltransferase gene-knockout (GalT KO) mice are able to produce natural anti-αGal antibodies apparently without any specific immunization. GalT KO mice are commonly used as a model immunological system for studying anti-αGal responses to Gal-positive xenografts in human. In this study, we compared the specificity of mouse and human αGal antibodies to realize the adequacy of the murine model. METHODS: Using hapten-specific affinity chromatography antibodies against Galα1-3Galß1-4GlcNAcß epitope were isolated from both human and GalT KO mice blood sera. Specificity of isolated antibodies was determined using a printed glycan array (PGA) containing 400 mammalian glycans and 200 bacterial polysaccharides. RESULTS: The quantity of isolated specific anti-Galα antibodies corresponds to a content of <0.2% of total Ig, which is an order of magnitude lower than that generally assumed for both human and murine peripheral blood immunoglobulin, with a high predominance of IgM over IgG (95% vs 5%). Analysis using a printed glycan array has demonstrated that (a) antibodies from both species bind not only the Galα1-3Galß1-4GlcNAcß epitope, but also unrelated glycans; (b) particularly, for human (but not mouse) antibodies the best binders appear to be bacterial polysaccharides; (c) the profile of mouse antibodies is broader, it is noteworthy that they recognize a variety of human blood group B epitopes and even glycans without the α-galactosyl residue. CONCLUSIONS: We believe that the mouse model should be used cautiously in xenotransplantation experiments when the fine epitope specificity of antibodies is critical.
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Anticuerpos , Galactosiltransferasas , Animales , Galactosiltransferasas/genética , Humanos , Ratones , Ratones Noqueados , Polisacáridos , Trasplante HeterólogoRESUMEN
Common carp (Cyprinus carpio) has an outstanding economic importance in freshwater aquaculture due to its high adaptive capacity to both food and environment. In fact, it is the third most farmed fish species worldwide according to the Food and Agriculture Organization. More than four million tons of common carp are produced annually in aquaculture, and more than a hundred thousand tons are caught from the wild. Historically, the common carp was also the first fish species to be domesticated in ancient China, and now, there is a huge variety of domestic carp strains worldwide. In the present study, we used double digestion restriction site-associated DNA sequencing to genotype several European common carp strains and showed that they are divided into two distinct groups. One of them includes central European common carp strains as well as Ponto-Caspian wild common carp populations, whereas the other group contains several common carp strains that originated in the Soviet Union, mostly as cold-resistant strains. We believe that breeding with wild Amur carp and subsequent selection of the hybrids for resistance to adverse environmental conditions was the attribute of the second group. We assessed the contribution of wild Amur carp inheritance to the common carp strains and discovered discriminating genes, which differed in allele frequencies between groups. Taken together, our results improve our current understanding of the genetic variability of common carp, namely the structure of natural and artificial carp populations, and the contribution of wild carp traits to domestic strains.
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Strong discrepancies in published data on the levels and epitope specificities of antibodies against the xenogenic N-glycolyl forms of sialoglycans (Hanganutziu-Deicher Neu5GcÉ2-3Galß1-4Glc and related antigens) in healthy donors prompted us to carry out a systematic study in this area using the printed glycan array and other methods. This article summarizes and discusses our published and previously unpublished data, as well as publicly available data from the Consortium for Functional Glycomics. As a result, we conclude that (1) the level of antibodies referred to as anti-Neu5Gc in healthy individuals is low; (2) there are antibodies that seem to interact with Neu5Gc-containing epitopes, but in fact they recognize internal fragments of Neu5Gc-containing glycans (without sialic acids), which served as antigens in the assays used and; (3) a population capable of interacting specifically with Neu5Gc (it does not bind the corresponding NAc analogs) does exist, but it binds the monosaccharide Neu5Gc better than the entire glycans containing it. In other words, in healthy donors, there are populations of antibodies capable of binding the Neu5Gc monosaccharide or the inner core -Galß1-4Glc, but very few true anti-Neu5GcÉ2-3Galß1-4Glc antibodies, i.e., antibodies capable of specifically recognizing the entire trisaccharide.
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Anticuerpos/inmunología , Epítopos/inmunología , Ácidos Neuramínicos/inmunología , Anticuerpos/sangre , Epítopos/sangre , Epítopos/química , Voluntarios Sanos , Humanos , Ácidos Neuramínicos/sangre , Ácidos Neuramínicos/químicaRESUMEN
The enigmatic and poorly studied sturgeon genus Pseudoscaphirhynchus (Scaphirhynchinae: Acipenseridae) comprises three species: the Amu Darya shovelnose sturgeon (Pseudoscaphirhynchus kaufmanni (Bogdanow)), dwarf Amu Darya shovelnose sturgeon P. hermanni (Kessler), and Syr Darya shovelnose sturgeon (P. fedtschenkoi (Bogdanow). Two species - P. hermanni and P. kaufmanni - are critically endangered due to the Aral Sea area ecological disaster, caused by massive water use for irrigation to support cotton agriculture, subsequent pesticide pollution and habitat degradation. For another species - P. fedtschenkoi - no sightings have been reported since 1960-s and it is believed to be extinct, both in nature and in captivity. In this study, complete mitochondrial (mt) genomes of these three species of Pseudoscaphirhynchus were characterized using Illumina and Sanger sequencing platforms. Phylogenetic analyses showed the significant divergence between Amu Darya and Syr Darya freshwater sturgeons and supported the monophyletic origin of the Pseudoscaphirhynchus species. We confirmed that two sympatric Amu Darya species P. kaufmanni and P. hermanni form a single genetic cluster, which may require further morphological and genetic study to assess possible hybridization, intraspecific variation and taxonomic status and to develop conservation measures to protect these unique fishes.
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Especies en Peligro de Extinción , Peces/genética , Genoma Mitocondrial , Filogenia , Animales , Asia Central , Ecosistema , Extinción Biológica , Peces/clasificación , Hidrobiología , Especificidad de la Especie , Contaminación Química del AguaRESUMEN
Modification of vaccine carriers by decoration with glycans can enhance binding to and even targeting of dendritic cells (DCs), thus augmenting vaccine efficacy. To find a specific glycan-"vector" it is necessary to know glycan-binding profile of DCs. This task is not trivial; the small number of circulating blood DCs available for isolation hinders screening and therefore advancement of the profiling. It would be more convenient to employ long-term cell cultures or even primary DCs from murine blood. We therefore examined whether THP-1 (human monocyte cell line) and DC2.4 (immature murine DC-like cell line) could serve as a model for human DCs. These cells were probed with a set of glycans previously identified as binding to circulating human CD14low/-CD16+CD83+ DCs. In addition, we tested a subpopulation of murine CD14low/-CD80+СD11c+CD16+ cells reported as relating to the human CD14low/-CD16+CD83+ cells. Manα1-3(Manα1-6)Manß1-4GlcNAcß1-4GlcNAcß bound to both the cell lines and the murine CD14low/-CD80+СD11c+CD16+ cells. Primary cells, but not the cell cultures, were capable of binding GalNAcα1-3Galß (Adi), the most potent ligand for binding to human circulating DCs. In conclusion, not one of the studied cell lines proved an adequate model for DCs processes involving lectin binding. Although the glycan-binding profile of BYRB-Rb (8.17)1Iem mouse DCs could prove useful for assessing human DCs, important glycan interactions were missing, a situation which was aggravated when employing cells from the BALB/c strain. Accordingly, one must treat results from murine work with caution when seeking vaccine targeting of human DCs, and certainly should avoid cell lines such as THP-1 and DC2.4 cells.