Search Results (325)

Lower respiratory tract infections caused by the respiratory syncytial virus (RSV) pose a major global public health challenge. The use of next-generation sequencing technologies enables detailed monitoring of viral genetic variability, which is crucial for evaluating the efficacy of immunoprophylactic measures. In this study, whole-genome sequencing of RSV was performed on 106 samples collected in the Russian Federation between September 2021 and April 2025. Three NGS platforms were employed: Illumina MiSeq, Oxford Nanopore Technologies MinION, and Qitan Tech QNome-3841. Using discrete phylogeographic methods, we estimated a minimum of 45 introduction events into Russia for RSV-A and 39 for RSV-B among the genomes included in the analysis. Most events were represented by a single Russian genome. These results indicate recurrent introductions of RSV into Russia from abroad. Given the limited genomic sampling available, most of these introductions were not associated with detectable transmission within the country. Full article

Wheat stem rust, caused by Puccinia graminis f. sp. tritici (Pgt), threatens food security owing to the pathogen’s high virulence and evolutionary variability, as well as the risk of selecting fungicide-resistant forms. For the first time in Russia, we performed a comparative analysis of virulence and genetic diversity in fungicide-target genes (SdhA–SdhD, Cyp51) in 35 monopustular Pgt isolates collected from regions with contrasting fungicide pressure (Saratov, n = 12; Chelyabinsk, n = 12; Moscow, n = 11). Phenotyping on an extended set of nearly isogenic lines carrying different Sr genes revealed regional specificity in effective resistance genes (Sr13, Sr26, Sr31, Sr32, Sr35), underscoring the vulnerability of genetic protection in Russian commercial cultivars. Targeted amplicon sequencing using the Oxford Nanopore Technologies platform produced 175 amplicons (~38,290 long reads, N50 = 1.06 kb), identifying 87% synonymous SNPs, 12% non-synonymous substitutions, and a single nonsense mutation in SdhA (position 580). The Cyp51 and SdhA genes were the most polymorphic. Recurrent non-synonymous mutations in these genes were detected in Pgt clones from regions with intensive agriculture and high use of chemical plant protection products. These results support the need for predictive breeding of resistant cultivars and adaptation of chemical control within integrated Pgt management in the European part of Russia. Full article

Adipogenic differentiation is essential for adipose tissue development, fat deposition, and metabolic regulation in livestock, but the role of RNA-binding proteins in sheep preadipocyte differentiation remains unclear. This study investigated whether Quaking (QKI) regulates sheep preadipocyte adipogenesis and explored its downstream regulatory mechanism. QKI expression was examined in sheep adipose tissues, and QKI knockdown was performed in sheep preadipocytes. Adipogenic marker expression, lipid accumulation, glucose consumption, Oxford Nanopore Technologies (ONT) full-length transcriptome sequencing, RNA immunoprecipitation coupled with quantitative real-time PCR (RIP-qPCR), actinomycin D assay, caveolae-associated protein 3 (Cavin3) knockdown, and simultaneous QKI and Cavin3 knockdown were used to evaluate the function and mechanism of QKI. QKI was expressed in different sheep adipose tissues, with relatively higher expression in tail fat. QKI knockdown reduced adipogenic marker expression, Oil Red O staining, and glucose consumption after adipogenic induction. ONT full-length transcriptome sequencing identified Cavin3 as a markedly upregulated candidate downstream target after QKI knockdown. RIP-qPCR showed enrichment of Cavin3 mRNA in QKI immunoprecipitates, and actinomycin D assays indicated that QKI knockdown delayed Cavin3 mRNA degradation. Cavin3 knockdown enhanced adipogenic marker expression, Oil Red O staining, and glucose consumption. Simultaneous QKI and Cavin3 knockdown increased adiponectin, fatty-acid-binding protein 4, and peroxisome proliferator-activated receptor gamma mRNA abundance and Oil Red O staining relative to the matched control, whereas glucose consumption was not significantly different. These findings suggest that QKI promotes sheep preadipocyte differentiation, at least in part, by reducing Cavin3 mRNA stability, providing insight into RNA-binding protein-mediated regulation of livestock adipogenesis. Full article

Background: Aztreonam–avibactam (ATM-AVI) is an important therapeutic option for infections caused by metallo-β-lactamase-producing Enterobacterales. However, the emergence of resistance may limit its clinical effectiveness. Here, we report two high-level ATM-AVI-resistant Escherichia coli isolates recovered from critically ill patients in a tertiary hospital in Guangzhou, both of whom had prior exposure to broad-spectrum antimicrobial agents. Methods: Antimicrobial susceptibility testing was performed by broth microdilution. Whole-genome sequencing (WGS) was conducted using the Illumina NovaSeq and Oxford Nanopore Technologies platforms for hybrid assembly. Resistome analysis, multilocus sequence typing (MLST), plasmid replicon typing, and phylogenomic analysis were performed. Comparative genomics with global ST410 isolates was used to investigate the evolutionary origin. Results: Antimicrobial susceptibility testing revealed high-level resistance to ATM-AVI. Hybrid whole-genome sequencing showed that both isolates belonged to the globally disseminated high-risk ST410 lineage and carried bla_NDM-5, CTX-M-type extended-spectrum β-lactamase genes, and an identical four-amino-acid insertion in penicillin-binding protein 3 (ftsI_I334IYRIK). Both isolates exhibited highly conserved chromosomal backbones, differed by only 29 core-genome single-nucleotide polymorphisms, and clustered within a China-associated ST410 clade. Notably, their key resistance determinants were located on distinct plasmid backgrounds: bla_NDM-5 was found on a predicted conjugative IncFIB/IncFIC plasmid in one isolate and on an IncX1 element with potential mobilization in the other. Conclusions: These findings provide important clinical evidence of high-level ATM-AVI resistance in Escherichia coli ST410 isolates in China and highlight the emergence of ATM-AVI resistance-associated determinants within a high-risk genomic background. Active phenotypic and genomic surveillance is warranted as ATM-AVI enters broader clinical use. Full article

Salivary microbiome profiling may represent a promising non-invasive approach for characterizing OSCC-associated microbial patterns and longitudinal microbiome dynamics during patient management. This exploratory pilot study aimed to longitudinally assess salivary microbiota profiles in patients with oral squamous cell carcinoma (OSCC) before and after tumor resection using Oxford Nanopore Technology. Unstimulated saliva samples were collected from 16 patients with OSCC at two time points (before and after tumor resection) and from 10 OSCC-free reference subjects. Microbial DNA was extracted using the QIAamp DNA Blood Kit (QIAGEN GmbH, Hilden, Germany) and subjected to long read metagenomic sequencing using the Oxford Nanopore MinION platform (v. 20.06.4, Oxford Nanopore Technologies, Oxford, UK). Taxonomic profiling was performed to longitudinally characterize salivary microbiota composition within patients and to provide descriptive comparisons with the OSCC-free reference cohort. Longitudinal analysis identified differences in salivary microbiota profiles between pre- and post-resection samples. Before surgery, an increased relative abundance of Neisseria subflava and Leptotrichia buccalis was observed. Post-surgical samples showed higher levels of Glaesserella parasuis, Streptomyces anulatus, and Lactobacillus species. Distinct microbial patterns were also descriptively observed between OSCC patients and OSCC-free controls, suggesting disease-associated dysbiosis. This exploratory longitudinal pilot study suggests differences in salivary microbiota profiles between samples collected before and after tumor resection in patients with OSCC, including changes in taxonomic composition and reduced alpha diversity. Given the limited sample size and the potential influence of unmeasured perioperative factors, these findings should be considered hypothesis-generating. Larger, well-controlled longitudinal studies incorporating standardized oral health assessment and detailed perioperative metadata are required to clarify the biological and clinical relevance of these observations. Full article

Accurate species identification is essential for biodiversity studies, ecological monitoring, and biosecurity, but current molecular approaches rely on PCR amplification, which requires universal primers. Here, we evaluated the possibility of primer-free species identification via direct RNA sequencing (dRNA-seq) by using Oxford Nanopore Technology (ONT). Total RNA isolated from three major eukaryotic kingdoms—fungal (Tuber magnatum Picco 1788), plant (Corylus avellana L. 1753), and animal (Eisenia fetida Savigny 1826) specimens—was sequenced without reverse transcription and PCR amplification. Taxonomic assignments based on similarity to ribosomal nuclear (fungi and animals) and chloroplast (plant) transcripts enabled reliable host species identification. Ribosomal RNA reads dominated the datasets and supported accurate identification of the target organisms, while additional sequences revealed associated microbiota and co-occurring taxa. Notably, dRNA-seq successfully detected symbiotic bacteria in E. fetida and latent fungal infection in C. avellana, highlighting the method sensitivity. However, the complex organization of nuclear ribosomal gene clusters may complicate taxonomic assignment in plants, underscoring the need for improved analytical pipelines. Overall, our results provide proof of concept that dRNA-seq enables primer-independent species identification while simultaneously providing insights into the host microbiomes. Direct sequencing of naturally present RNA molecules generated sufficient sequence information for species identification across different taxonomic kingdoms, indicating the technical feasibility of the developed technique. Further validation using larger biological datasets and direct comparisons with conventional sequencing methods will determine its wider applicability as an innovative and complementary species identification approach. Full article

The quail is a valuable model organism for avian developmental biology and genetic improvement; however, the full-length transcriptomic landscape and regulatory networks underlying critical embryonic developmental stages remain poorly characterized. In this study, we employed Oxford Nanopore Technologies full-length transcriptome sequencing to systematically analyze leg muscle tissues from quail embryos at embryonic day 8 (E8, pre-differentiation stage) and embryonic day 14 (E14, later differentiation stage), with four biological replicates per stage (eight libraries in total). Nanopore sequencing identified a total of 54,938 transcripts and 21,574 genes, including 49,678 known transcripts, 20,980 known genes, 5260 novel transcripts, and 594 novel genes. Analysis of transcriptome structure revealed that alternative first exon and exon skipping represented the predominant stage-specific alternative splicing events, and we identified 2692 lncRNAs. Differential expression analysis identified substantial transcriptome dynamics from E8 to E14, with 5934 differentially expressed transcripts and 3745 differentially expressed genes identified. Functional enrichment analyses, including KEGG, GSEA and PPI network analysis, revealed that the E8 transcriptome was predominantly enriched for pathways associated with cell proliferation, whereas the E14 transcriptome shifted toward the coordinated refinement of myofiber structural assembly and energy metabolism. Here, we present a high-resolution full-length transcriptional atlas of quail skeletal muscle development and identify candidate regulatory genes, offering a valuable theoretical framework and data resource for refining quail genome annotation. Full article

The reliable identification of cultivated and naturalised mulberries is complicated by morphological plasticity, the historical movement of the planting material, and partly discordant nuclear and plastid signals. We evaluated four field-identified Morus alba L. trees, one from each of four localities in Mangystau, Western Kazakhstan, using archived consensus sequences for ITS, matK, rbcL, and trnH-psbA, together with a low-coverage Oxford Nanopore Technologies (ONT) dataset. Because the design comprised one tree per locality (n = 4), analyses were restricted to accession-level descriptive comparisons, and no population-genetic, phylogeographic, or formal phylogenetic inference was attempted. Archived alignment summaries indicated mean pairwise distances of 0.13% for matK, 0.44% for ITS, 0.94% for rbcL, and 2.92% for trnH-psbA; these values are reported as retained dataset descriptors rather than estimates of population diversity. An additional product generated with Rosaceae-derived s6pdh primers was excluded because the target identity and orthology could not be verified. The ONT run yielded 19,958 pass reads (69.27 Mb; read N50 3574 bp). Reference-enriched assembly produced a 14,824 bp candidate plastid-associated contig, approximately 9.3% of a typical Morus plastome. Its short length, incomplete and non-collinear annotations, and the absence of retained depth, polishing, assembly graph, and join support diagnostics preclude its interpretation as a complete, circular, or structurally validated plastome. This study provides a transparent pilot baseline for Mangystau mulberries and establishes quality control criteria for replicated sampling, validated markers, and deeper organelle sequencing. Full article

HCT116 is a colorectal cancer cell line frequently used in anti-tumor drug development experiments as well as in studies of the molecular machinery of eukaryotic cells. It is well characterized by the presence of several single-nucleotide and short mutations in multiple oncogenes and tumor suppressor genes, including KRAS, PIK3CA, MLH1, CTNNB1, CDKN2A, TGFBR2, and BRCA2. However, its landscape of large genomic rearrangements (LGRs) and copy number variants (CNVs) is still far from being fully understood. Therefore, the aim of this study was to identify LGRs and CNVs in several HCT116 cell line samples using Oxford Nanopore sequencing technology, including three samples from the SRA NCBI database, and to compare common and unique variants across all samples. Using the recently developed eLaRodON tool, we identified 22,666 common LGRs, among which more than 70% of tandem duplications and deletions larger than 80 kb were confirmed by CNV analysis. Among LGRs affecting protein-coding sequences, two in-frame rearrangements were identified: a deletion of exons 4–6 and a duplication of exon 10 in the CCSER1 gene, which encodes a cell division regulator protein. Given its high rearrangement rate in various tumors and the clinical significance of its overexpression, this finding may be potentially useful in future research on this cell line. Regarding differences between samples, we found that LGRs in the laboratory sample and in one of the three SRA NCBI samples occurred more frequently via ALR/Alpha repeats than via Alu repeats, in contrast to common LGRs and those unique to the other samples, a finding that may indicate the presence of unique mechanisms of genomic instability. Thus, this study reveals a broad spectrum of large genomic rearrangements and copy number variants that can be identified in the HCT116 cell line using Oxford Nanopore sequencing, including rearrangements specific to distinct cell line samples. Full article

Oxford Nanopore technology enables cost-effective, portable, long-read analyses of pathogen genomes. Accurate detection and interpretation of small circular viral genomes, including Anelloviridae, remain challenging due to limited base-level error quantification in rolling-circle amplification (RCA)-derived datasets. Here, we characterized read-level sequencing error profiles using M13mp18, a 7.2 kb circular phage genome, subjected to 1X and 3X shearing during library preparation. M13mp18 DNA was serially diluted into pooled anellovirus-positive plasma DNA extracts. Using custom error-analysis pipelines, we quantified mismatch, insertion, and deletion rates and evaluated consensus reconstruction accuracy across simulated sequencing depths. Since metagenomic viromes contain mixtures of related genomes and uneven coverage across taxa, depth-normalized subsampling was used to assess the precision of read-level error estimates under heterogeneous coverage. Across four benchmarked datasets, per-base error rates ranged from 0.018 to 0.022 errors per aligned base. Complete M13mp18 reference reconstruction was achieved at input levels ≥ 4.6 log₁₀ copies, and consensus sequences reached 100% identity at depths ≥ 15X when sufficient reads were available. Below 4.6 log₁₀ input copies, recovery was inconsistent. These findings provide a controlled empirical characterization of read-level error behavior in RCA-derived nanopore sequencing and support the interpretation of circular DNA virome data generated in complex metagenomic backgrounds. Full article

The white rhinoceros (Ceratotherium simum) offers a unique model for investigating the genomic consequences of extreme demographic bottlenecks. However, the fragmented southern white rhinoceros genome assembly has limited chromosome-scale structural and evolutionary comparisons with the functionally extinct northern subspecies. Here, we report a chromosome-scale genome assembly for the southern white rhinoceros by integrating Oxford Nanopore Technology long-read sequencing, Illumina short-read polishing and high-throughput chromosome conformation capture (Hi-C) scaffolding. The final assembly spans 2.48 Gb and achieves a contig N50 of 42.06 Mb, representing a 452-fold improvement in contiguity over the previous assembly. In total, 2.46 Gb of sequence was anchored to 40 autosomes plus the X and Y chromosomes. Genome annotation identified 1.13 Gb of repetitive elements (45.7% of the assembly), 22,593 protein-coding genes, and 100.68 Mb of segmental duplications. Inspection of the major histocompatibility complex class II gene region further supported the local assembly and annotation reliability, revealing conserved gene composition and order between the southern and northern white rhinoceroses. Whole-genome comparison with the northern white rhinoceros assembly indicated extensive chromosome-scale synteny, along with localized structural variants between the two subspecies, including 111 inversions spanning 33.48 Mb and 497 translocations spanning 36.48 Mb. Furthermore, coalescent demographic reconstruction indicated asynchronous Pleistocene population dynamics for southern and northern white rhinoceroses, reflecting divergent responses to historical climate oscillations. Both subspecies also exhibit lower recent effective population sizes than estimated Pleistocene ancestral levels, underscoring persistent conservation concern. This assembly provides a useful resource for evaluating the genomic consequences of historical bottlenecks, informing future genomic-rescue plans, and strengthening the comparative framework for rhinoceros conservation and evolutionary genomics. Full article

Rapid and accurate diagnosis of infectious diseases in aquaculture is essential for preventing major economic and ecological losses. Traditional culture-based methods focus on isolation of individual pathogens, and often are burdened with extended processing times, particularly during investigations of polymicrobial infections. Application of Oxford Nanopore Technologies (ONT) sequencing offers a rapid, culture-independent workflow for the identification of bacterial and fungal pathogens directly from fish tissues. Swab and organ samples from four cases (1: Salmo spp.; 2: Cyprinus carpio; 3: Salvelinus fontinalis; 4: Heniochus acuminatus) were analyzed using ONT long-read sequencing for metagenomic screening and bioinformatic classification. The results revealed case-, species-, and tissue-specific microbial profiles, with external tissues showing higher microbial diversity and internal organs enriched in pathogenic taxa. Dominant pathogens included Streptococcus iniae, Aeromonas hydrophila, Pseudomonas spp., and Saprolegnia parasitica, alongside opportunistic zoonotic bacteria such as Escherichia coli and Acinetobacter baumannii. We demonstrate the potential for diagnostic application of ONT sequencing in investigations and detection of multi-pathogen infections, including assessments of microbial community structure changes during disease outbreaks in aquatic species. The presented workflow enables rapid, cost-effective, and comprehensive pathogen profiling, supporting early disease surveillance and improved management in aquatic veterinary practice. Full article

Long-read sequencing technologies provide portable and flexible service, making them attractive for small-scale sequencing studies. However, many existing RNA-sequencing analysis frameworks are designed for transcriptome-wide analyses and require substantial computational resources. Here we present a lightweight and reproducible computational pipeline for targeted long-read transcriptomic profiling using Oxford Nanopore Technologies (ONT) cDNA sequencing data. The pipeline was evaluated using targeted long-read transcriptomic datasets generated from formalin-fixed paraffin-embedded (FFPE) colorectal carcinoma samples previously classified as microsatellite instability—high (MSI-high) by PCR-based testing. Libraries were sequenced on the Oxford Nanopore MinION platform using R10.4.1 flow cells. Application of the workflow enabled rapid quantification of mismatch repair gene expression and detection of immune-related transcripts including CD8A, PDCD1, and HAVCR2 across multiplexed barcode samples. The pipeline performs targeted alignment of long-read sequencing data to a custom transcript reference panel using minimap2, followed by gene-level read counting and normalization using reads-per-million (RPM). Optional modules enable immune marker profiling, detection of reads aligning to multiple genes, exploratory variant analysis, and visualization of expression patterns. By combining simplicity, reproducibility, and minimal computational overhead, the present pipeline provides an accessible framework for targeted transcriptomic analysis of long-read sequencing data. It may facilitate adoption of ONT-based transcriptomic profiling in settings with restricted computational resources. Full article

Sea buckthorn (Hippophae rhamnoides L.) is a valuable plant whose fruits are rich in biologically active compounds. We sequenced the genome of variety Triumf of H. rhamnoides ssp. mongolica Rousi on the Oxford Nanopore Technologies (ONT) platform. With the Hifiasm algorithm optimized for ONT data, we assembled the 1.17-Gb genome into eleven complete chromosomes and one chromosome consisting of two contigs, which were scaffolded (Chr3). Eleven of twelve chromosomes had pronounced telomeric repeats at both ends and were assembled as telomere-to-telomere (T2T), and one chromosome (Chr12) had telomeric repeats only at one end. We also sequenced transcriptomes of thirteen Triumf organs/tissues and performed genome annotation using these and previously obtained RNA-Seq data for this variety. As a result, we predicted 25,915 genes and 30,527 transcripts. Repetitive elements comprised 66.9% of the genome size. The obtained near-T2T annotated genome assembly of H. rhamnoides ssp. mongolica variety Triumf enabled the identification of correct composition and sequences of important gene families in sea buckthorn. We demonstrated this with the FAT, SAD, and FAD gene families involved in fatty acid synthesis. Expression analysis revealed which FAT, SAD, and FAD genes are essential for specific organs/tissues. Thus, the Triumf genome assembly is a crucial tool for basic and applied studies of H. rhamnoides. Full article

Background: Human hospitals and veterinary centres are hotspots for resistant microbes and plasmids, and metagenomic sequencing offers an agnostic insight into microbiomes, resistomes, and mobilomes, informing strategies for reducing AMR spread. Methods: Environmental samples, including wastewater and surface swabs, were taken from a tertiary human hospital ward (36 samples) and a companion animal veterinary hospital (48 samples) in London. Whole DNA was extracted and metagenomic sequencing undertaken using Oxford Nanopore Technologies’ MinION. Data were analyzed for microbiomes, resistomes and mobilomes and compared. Results: Microbial diversity analyses highlight higher richness across human hospital (HH) environmental samples, but more evenness in veterinary hospital (VH) environmental samples. Diversity showed distinct microbial communities in the HH and VH samples. There were significantly more total antimicrobial resistance gene (ARG) types (p < 0.0001) in the environmental HH samples compared with the environmental VH samples. There was a significantly higher mean number of Enterobacteriales plasmid types (p ≤ 0.0001) in the HH samples. There were significantly more total Gram-Positive plasmid types (p ≤ 0.0001) in the VH samples. Discussion: This research highlights the presence of human and animal pathogens, ARGs and mobile genetic elements in clinical environments, underscoring the importance of multisectoral surveillance. Integrating taxonomic, resistome, and mobilome analyses provides a better understanding of the potential for AMR dissemination at the human–animal–environment interface. This provides insights relevant for the development of targeted surveillance and mitigation strategies within a OH framework. Full article