Search Results (2,147)

Background/Objectives: Targeted gene sequencing (TGS) for Comprehensive Genomic Profiling (CGP) use in sarcomas has recently increased in clinical practice. We report on TGS real-world data over a period of 3 years (2022–2025) at the IRCCS Istituto Ortopedico Rizzoli, with the aim of identifying potential actionable targets and providing therapeutic indications for advanced sarcoma patients. Methods: We analyzed 22 advanced sarcoma patients by using the VariantPlex Pan Solid Tumor kit panel, including 185 genes. In nine cases, saliva samples for germinal DNA analysis were available. Sequencing was performed on the NextSeq-500 Platform and analyzed with Archer Analysis software. The Cancer Genome Interpreter and OncoKB Database tools were used to find potential actionable targets. Results: We found the most frequent genetic variants, including missense, deletion, duplication, and delins, in the NOTCH4, AR, BARD1, MUC16, and ROS1 genes. Copy Number alterations affected the CDKN2A, CDKN2B, TP53, RHOA, MYC, CCND3, and DDR2 genes mainly in osteosarcoma samples. In four patients, longitudinal analyses of subsequent lesions showed the maintenance of most genomic alterations and enrichment in missense or splice variants in PMS2, SMARCA4, ARID1A, AKT1, BMPR1A, and PTEN, indicating the occurrence of tumor evolution. Germline variants subtraction identified the specific somatic tumor mutations. Advantages and disadvantages of our approach were considered in order to refine the analysis setting and better select possible actionable targets. Conclusions: Early access to genomic analyses, routine germline assessment, and broad gene panels would help in identifying possible targeted drugs with sufficient evidence of activity beneficial to each patient. In the clinical management of advanced sarcoma patients, when analyzing cost-effectiveness and sustainability, the role of the Molecular Tumor Board in the governance of the complexity introduced by mutational oncology should be considered. Full article

Side-channel attacks leveraging microarchitectural components such as caches and translation lookaside buffers (TLBs) pose increasing risks to cryptographic and machine-learning workloads. This paper presents a comparative study of performance and side-channel leakage under two page-size configurations—standard 4 KB pages and 2 MB huge pages—using paired attacker–victim experiments instrumented with both Performance Monitoring Unit (PMU) counters and precise per-access timing using rdtscp(). The victim executes repeated, key-dependent memory accesses across eight cryptographic modes (AES, ChaCha20, RSA, and ECC variants) while the attacker records eight PMU features per access (cpu-cycles, instructions, cache-references, cache-misses, etc.) and precise rdtscp() timing. The resulting traces are analyzed using a multilayer perceptron classifier to quantify key-dependent leakage. Results show that the 2 MB huge-page configuration achieves a comparable key-classification accuracy (mean 0.79 vs. 0.77 for 4 KB) while reducing average CPU cycles by approximately 11%. Page-index identification remains near random chance (3.6–3.7% for PMU side-channels and 1.5% for timing side-channel), indicating no increase in measurable leakage at the page level. These findings suggest that huge-page mappings can improve runtime efficiency without amplifying observable side-channel vulnerabilities, offering a practical configuration for balancing performance and security in user-space cryptographic workloads. Full article

Resource-constrained sensor nodes in Internet-of-Things (IoT) and embedded sensing applications frequently rely on low-cost microcontrollers, where even basic algorithmic choices directly impact latency, energy consumption, and memory footprint. This study evaluates six sorting algorithms—Bubble Sort, Insertion Sort, Selection Sort, Merge Sort, Quick Sort, and Heap Sort—in the restricted environment that microcontrollers provide. Three Arduino boards were used: Arduino Uno, Arduino Leonardo, and Arduino Mega 2560. Each algorithm was implemented in its unoptimized form and tested on datasets of increasing size, emulating buffered time-series sensor readings in random, ascending, and descending order. Execution time, number of write operations, and memory usage were measured. The tests show clear distinctions between the slower $O\left(n^2\right)$ algorithms and the more efficient $O(nlogn)$ algorithms. For random inputs of $n=1000$ elements, Bubble Sort required 1,958,193.75 $\mathsf{\mu }$son average, whereas Quick Sort completed it in 54,260.50 $\mathsf{\mu }$s and Heap Sort in 92,429.00 $\mathsf{\mu }$s, i.e., speedups of more than one order of magnitude compared to the quadratic baseline. These gains, however, come with very different memory footprints. Merge Sort kept the runtime below 100,000 $\mathsf{\mu }$s at $n=1000$ but required approximately 2023 bytes of additional static random-access memory (SRAM), effectively exhausting the 2 kB SRAM of the Arduino Uno. QuickSort used approximately 311 bytes of extra SRAM and failed to process larger ascending and descending datasets on the more constrained boards due to its recursive pattern and stack usage. Heap Sort offered the best overall trade-off: it successfully executed all tested sizes up to the SRAM limit of each board while using only about 12–13 bytes of additional SRAM and keeping the runtime below 100,000 $\mathsf{\mu }$s for $n=1000$. The results provide practical guidelines for selecting sorting algorithms on 8-bit AVR Arduino-class microcontrollers, which are widely used as simple sensing and prototyping nodes operating under strict RAM, program-memory, and energy constraints. Full article

The continuing spread of SARS-CoV-2 and the associated morbidity and mortality, especially in vulnerable populations, highlight the need for the development of antiviral therapeutics. Reverse genetics systems and reporter viruses are valuable for antiviral screening by simplifying methods to detect and quantify virus infections. This study aimed to generate wild-type and Nluc reporter full-length SARS-CoV-2 molecular clones and viruses as tools for high-throughput antiviral assays. The large SARS-CoV-2 genome (~30 kb) makes cDNA cloning and virus rescue technically challenging, so we opted to use cDNA chemical synthesis services to generate full-length wild-type and reporter Delta and Omicron clones. Clone-derived Delta and Omicron wild-type and reporter viruses were successfully rescued and showed replication kinetics comparable to patient-derived isolates. Nluc reporter viruses displayed stable luciferase expression that correlated with viral titres, supporting their reliability as replication substitutes. Antiviral assays measuring replication inhibition by Remdesivir, Molnupiravir, and Nirmatrelvir, based on Nluc expression, yielded IC₅₀ values and selectivity indices consistent with published ranges. Finally, Delta Nluc viruses replicated in primary human bronchial epithelial cells, demonstrating the application of clone-derived viruses in physiologically relevant models. The SARS-CoV-2 cDNA clones and Nluc reporter viruses derived from DNA synthesis services provide a rapid, scalable reverse genetics platform for generating new viruses and developing assays to rapidly assess antiviral compounds against current and emerging SARS-CoV-2 variants or coronaviruses that may emerge in the future. Full article

The aim of this study was to evaluate the effect of heat treatment, including solutionising and ageing in the temperature range of 20–250 °C, on the microstructural, mechanical, and tribological properties of the Al 7075 alloy. Microscopic analysis revealed that in the as-received condition and after natural ageing, the microstructure is characterised by the presence of elongated grains and a banded distribution of precipitates, whereas higher ageing temperatures lead to their coarsening and the initiation of recrystallisation processes. The highest hardness (189 HV) was obtained after ageing at 100 °C for 48 h, while further increases in temperature caused a systematic decrease in hardness—down to 85 HV at 250 °C for 4 h. Impact tests showed that in the as-received condition, the material reached a value of 7 J/cm², after natural ageing 15 J/cm², and the maximum (26 J/cm²) was achieved for samples aged at 250 °C for 4 h. Tribological tests conducted using the T-07 method confirmed the dependence of wear resistance on heat treatment parameters—the lowest relative abrasive wear resistance coefficient was observed after natural ageing (k_b = 0.860), and the highest after ageing at 250 °C for 4 h (k_b = 1.216). The results obtained indicate that moderate ageing conditions (100–150 °C) favour increased hardness, whereas higher temperatures (200–250 °C) lead to an improvement in impact strength and tribological resistance, which showed an inversely proportional relationship with hardness, contrary to Archard’s law. Full article

In this study we employed nanopore whole genome sequencing to analyze the resistance genes, genomic islands and prophage DNA in two multidrug resistant E. rhusiopathiae strains, i.e., 670 and 147, isolated from domestic geese. MLST profiles and core-genome phylogeny were determined to assess strain relatedness. In strain 670 (serotype 8, ST 113), a novel 53 kb prophage φEr670 carrying the lnuB and lsaE resistance genes was identified. Regions highly homologous to the φEr670 prophage were detected in 36 of 586 (6.14%) publicly available E. rhusiopathiae genomes, as well as in some other Gram-positive bacteria, and usually contained resistance genes. E. rhusiopathiae strain 147 (serotype 5, ST 243) was found to contain a composite 98 kb genomic island (GI_Er147) carrying the ant(6)-Ia and spw genes, as well as gene encoding a putative lincosamide nucleotidyltransferase designated lnu(J) and a vat family gene encoding a putative streptogramin A O-acetyltransferase. The lnu(J) gene exhibited 83.6% homology to the lnu(D) gene, and lnu(J)-positive E. rhusiopathiae strains displayed intermediate susceptibility to lincomycin. Vat-positive strain 147 and vat-negative E. rhusiopathiae strains showed similar susceptibility to quinupristin/dalfopristin. The presence of the Tn916 transposon carrying the tetM gene was confirmed in the genomes of both E. rhusiopathiae strains; in strain 147, however, Tn916 was located within ICEEr1012. Based on analyses of additional E. rhusiopathiae genomes, the integration sites of Tn916, ICEEr1012, and GI_Er147 were identified as genomic “hot spots,” contributing to the genome plasticity of E. rhusiopathiae. Prophage φEr670 and GI_Er147 as well as the Tn916 transposon and ICEEr1012 are most likely responsible for the dissemination of resistance genes in E. rhusiopathiae. Prophages highly homologous to φEr670 act as carriers of resistance genes in various Gram-positive bacteria. However, the transferability of the identified genetic elements and the functional role of the lnu(J) gene require further investigation. This study provides new insights into the diversity of MGEs in E. rhusiopathiae and advances understanding of the genomic mechanisms driving antimicrobial resistance in Gram-positive bacteria. Full article

Mandibular prognathism (Class III malocclusion) is a craniofacial anomaly characterized by an anteriorly positioned mandible, a concave facial profile and impaired mastication, and appears unusually frequently in Dolang sheep (Ovis aries). We combined clinical phenotyping and three-dimensional (3D) genome profiling to investigate this trait in a Dolang sheep flock. We examined 959 animals using standardized criteria, estimated a local prevalence of 10.3%, and assembled a 200 affected/200 unaffected case–control cohort for genomic analyses. As an exploratory pilot study of 3D genome architecture, we generated in situ Hi-C datasets from mandibular bone of two affected and two control sheep. At 40 kb resolution, global topologically associating domain (TAD) organization and boundary strength were broadly conserved between groups, but sliding-window analyses identified a small number of 1 Mb hotspots where affected animals showed increased TAD-boundary density and strengthened insulation. These UNDER-enriched windows lay near genes with plausible roles in craniofacial development, including ROBO2, COL27A1, VRK2 and a cytokine cluster (IL22/IL26/IFNG with MDM1). Together, our data indicate that mandibular prognathism in Dolang sheep is associated with localized remodeling of chromatin insulation at a restricted set of gene-proximal loci and highlight candidate regions and mechanisms for integration with whole-genome sequencing, association and transcriptomic data. Full article

Background: Gynostemma pentaphyllum (Thunb.) Makino is an important medicinal plant within the Cucurbitaceae family. Despite its economic and pharmacological importance, genomic resources for this species remain limited. Methods: We sequenced and assembled the complete mitochondrial genome of G. pentaphyllum. Comparative analyses were conducted to investigate the genomic structure, gene content, RNA editing events, and intracellular gene transfer (IGT) from chloroplasts. Additionally, phylogenomic relationships, synteny, and the selective pressure on mitochondrial genes were evaluated against related species within Cucurbitaceae. Results: The ~324 kb mitogenome has a multipartite architecture of six circular-mapping molecules. It encodes the typical complement of mitochondrial protein-coding genes, tRNAs, and rRNAs found in angiosperms. Extensive C-to-U RNA editing, including events that generate functional start and stop codons, points to substantial post-transcriptional regulation. We also detected multiple chloroplast-derived fragments, including several intact genes, indicating active intracellular gene transfer. Phylogenomic analyses of conserved mitochondrial genes place G. pentaphyllum firmly within Cucurbitaceae, clustering it with Thladiantha cordifolia and Momordica charantia, whereas synteny comparisons reveal pronounced structural rearrangements with respect to these close relatives. While most genes evolve under strong purifying selection, rps1, sdh3, and sdh4 show signatures of accelerated evolution; furthermore, haplotype networks based on conserved loci further corroborate the close affinity with T. cordifolia. Conclusions: This study provides the first high-resolution mitogenome resource for G. pentaphyllum and candidate mitochondrial markers for species authentication, evolutionary studies, and breeding in Gynostemma and related cucurbits. Full article

Background: Erysipelothrix rhusiopathiae is an important zoonotic pathogen in poultry, yet little is known about its antimicrobial resistance (AMR) dynamics in avian hosts. With growing concerns about subtherapeutic antimicrobial use in animal agriculture, poultry-origin isolates represent a potential but under-characterized reservoir of resistance genes. Methods: We phenotypically tested 38 E. rhusiopathiae strains isolated from geese, ducks, and turkeys in Hungary (2024) using broth microdilution against 18 antimicrobial agents, following Clinical Laboratory Standards Institute (CLSI) guidelines. Nineteen phenotypically resistant strains were selected for whole-genome sequencing (Illumina platform), followed by de novo hybrid assembly, gene annotation (Prokka, CARD, VFDB), mobile element detection (Mobile Element Finder), and phylogenetic inference (autoMLST). Results: All isolates were susceptible to β-lactams, including penicillin, amoxicillin, and third-generation cephalosporins. Resistance to tetracyclines (up to 10.5%) and florfenicol (5.3%) was most frequently detected. Genomic analysis revealed the presence of tetM (9/19), tetT (2/19), and erm(47) (2/19) genes, all associated with chromosomally integrated mobile elements, ICE Tn6009 and IS ISErh6. Phylogenomic analysis demonstrated tight clustering into four clades, suggesting clonal expansion. Notably, one strain harbored a 64.8 kb genomic island carrying ermC, the first such finding in poultry-derived E. rhusiopathiae. Conclusions: Our data highlights the early emergence of mobile AMR determinants in E. rhusiopathiae from poultry and suggests that horizontal gene transfer may drive resistance even in chromosomally encoded contexts. The genomic stability and phylogenetic homogeneity of avian isolates underscore the need for targeted AMR surveillance in poultry sectors to mitigate potential zoonotic transmission risks. Full article

Camellia sinensis ‘hainanensis’ (Hainan Sheng tea) is an endemic tea germplasm resource native to Hainan Island, China. Using complete chloroplast genome sequencing combined with comprehensive comparative analyses, we elucidated the genetic architecture of six C. sinensis accessions. The chloroplast genomes exhibited a typical quadripartite circular structure (~157 Kb) comprising 80 unique protein-coding genes, 30 tRNA genes, and 4 rRNA genes. Expansion and contraction of the inverted repeat (IR) regions led to boundary shifts affecting genes, while nucleotide diversity within the large single-copy (LSC) and small single-copy (SSC) regions (Pi > 0.0035) markedly exceeded that of the conserved IR regions. Phylogenetic reconstruction revealed that C. sinensis ‘hainanensis’ shared the closest evolutionary relationship with Yunnan large-leaf tea (Camellia grandibracteata), supporting its independent lineage within the genus. A polymorphic molecular marker derived from the hypervariable non-coding region (trnT–psbD) may serve as a useful preliminary marker for distinguishing C. sinensis ‘hainanensis’ from related taxa and hybrids. This study provides the first comprehensive comparison of complete chloroplast genomes of six C. sinensis ‘hainanensis’, identifies three distinct plastome types, and develops a molecular marker that can reliably distinguish these types, offering valuable genomic resources for future studies on tea evolution and germplasm identification. Full article

This article addresses the lack of low-cost, secure image-transmission solutions for IoT systems in remote environments. The design and implementation of a complete LoRa-based transmission system using ESP32 microcontrollers and Ebyte E220 modules, featuring AES-CBC encryption, HMAC integrity protection, and a custom retransmission protocol, are presented. The system achieves 100% packet delivery ratio (PDR) for 20 kB images over distances exceeding 2 km under line-of-sight conditions, with functional transmission up to 4.1 km. Image transmission time ranges from 35 s (0.1 m) to 110 s (600 m), while energy consumption increases from 4.95 mWh to 15.18 mWh. Critically, encryption imposes less than 1% overhead on total energy consumption. Unlike prior work focusing on isolated components, this article provides a complete, deployable architecture combining (i) low-cost hardware (<USD 50 total), (ii) long-range LoRa communication, (iii) custom reliability mechanisms for fragmenting 20 kB images into 100 packets, and (iv) end-to-end cryptographic protection, all evaluated experimentally across multi-kilometer distances. These findings demonstrate that secure long-range image transmission using commodity hardware is feasible and scalable for smart home and industrial monitoring applications. Full article

Background: Chemotherapy remains one of the main approaches for treating malignant tumors, but repeated exposure to cytostatics leads to multidrug resistance (MDR), increasing tumor aggressiveness and reducing therapeutic efficacy. Identifying adjuvant agents that restore tumor sensitivity to drugs while minimizing toxicity is a cornerstone challenge today. This study aimed to investigate the potential of mesyl phosphoramidate antisense oligonucleotides (µ-ASOs) targeting miR-17, miR-21, and miR-155 as agents for enhancing the efficacy of cisplatin (Cis) and doxorubicin (Dox) in MDR-positive human epidermoid carcinoma KB-8-5 cells. Methods: Optimal regimens for the simultaneous application of µ-ASOs and Dox or Cis in KB-8-5 cells, including a concentration-dependent analysis and the type of compound interaction in combinations (synergy/additivity/antagonism), were studied using the MTT assay. Antiproliferative effects of the combinations were assessed using the real-time cell monitoring xCELLigence system. The potential molecular mechanism underlying KB-8-5 cell sensitization to cytostatics was investigated using RT-PCR and Western blot hybridization, supported by bioinformatic reconstruction of the gene network. Results: The most effective combinations including µ-ASOs targeting miR-21 and miR-17 together with Cis or Dox demonstrated additive to moderately synergistic effects on KB-8-5 cell viability (HSA synergy score = 4.8–8.7). The co-application of µ-ASOs allowed a 5- to 20-fold reduction in the dose of cytostatics, while maintaining a strong antiproliferative effect of 70–95%. Sensitization of KB-8-5 cells to Cis or Dox following µ-ASO treatment was mediated by a 1.5- to 3-fold decrease in the levels of the well-known MDR marker ABCB1 as well as the newly identified MDR-associated targets ZYX, TUBA4A, and SEH1L. Conclusions: miRNA-targeted mesyl phosphoramidate oligonucleotides are effective tools for overcoming resistance to the clinically approved chemotherapeutics cisplatin and doxorubicin. The relationship between miR-21, miR-17, and miR-155 and the novel MDR markers such as SEH1L, TUBA4A, and ZYX was revealed, thereby expanding the current understanding of the molecular mechanisms underlying tumor cell resistance to chemotherapy. Full article

Duchenne muscular dystrophy (DMD) is a severe X-linked hereditary disorder caused by pathogenic variants in the DMD gene encoding the dystrophin protein. The absence of functional dystrophin leads to destabilization of the dystrophin-associated glycoprotein complex (DAPC), sarcolemmal damage, and progressive degeneration of muscle fibers. Current therapeutic strategies focus on restoring dystrophin expression using genome editing approaches. Adeno-associated virus (AAV) vectors represent the primary delivery platform due to their strong tropism for muscle tissue, low immunogenicity, and ability to achieve long-term transgene expression. However, the limited packaging capacity of AAV (~4.7 kb) necessitates the use of truncated mini- and micro-dystrophin transgenes as well as compact genome editing systems (SaCas9, NmeCas9, Cas12f, TIGR-Tas, and others). Major challenges include immune responses against the viral capsid and transgene products, as well as the inability to perform repeated administrations. Moreover, the duration of expression is limited by the episomal nature of AAV genomes and their loss during muscle fiber regeneration. Despite substantial progress, unresolved issues concerning safety, immunogenicity, and stability of genetic correction remain, defining the key directions for future research in DMD therapy. Full article

Sansevieria, a perennial herb known for its ornamental and medicinal value, has many varieties due to its leaf size and stripe color. However, it is very difficult to distinguish them during the seedling stage. In this study, we conducted chloroplast genome sequencing analysis on 10 cultivars of Sansevieria trifasciata. The chloroplast genomes exhibited a typical quadripartite circular structure (154.2–158.7 kb), encoding 113 functional genes with highly conserved gene order. Phylogenetic analysis supported the evolutionary linkage between Sansevieria and Dracaena. Dynamic inverted repeats (IR) boundary expansions/contractions, particularly species-specific patterns in ndhF and rps19 gene distributions across IR junctions, indicating its adaptive divergence. We also discovered the trnT-psbD marker, which is a deletion marker developed from hypervariable regions and can effectively distinguish closely related species. This work provides critical molecular tools and theoretical foundations for germplasm identification, phylogenetic reconstruction, and chloroplast genome evolution in Sansevieria, and also promotes taxonomic revisions in Asparagaceae. Full article

Helianthus annuus L. is one of the major oilseed crops worldwide, and its production is seriously affected by a highly destructive necrotrophic pathogen, Sclerotinia sclerotiorum (S. sclerotiorum). The use of resistant cultivars is the best control measure via molecular breeding; however, the gene action underlying resistance to this stress is not well-established. Here, we conducted QTL analysis for S. sclerotiorum resistance in a recombinant inbred line (RIL) population that were developed from parents with resistant (C6) and susceptible (B728) to the disease. A high-density genetic linkage map with 6059 single nucleotide polymorphism (SNP) markers and a total length of 2763 cM was developed. The lesion length (LL) and the lesion area (LA) in the field, under climate chamber conditions or greenhouse conditions, were assessed following standardized inoculation protocols. A total of 16 major QTL for LL and 12 for LA were detected across three experimental environments, explaining 1.58–32.86% of the phenotypic variation. Of these, a major-effect QTL, qSCL2.4 on chromosome 2, could explain 30.22% of phenotypic variance with alleles from parent C6 which had more increased resistance to S. sclerotiorum. Fine-mapping in the BC₁F₃ population narrowed the locus to a 226.7 kb interval. HaWRKY48, which encodes a WRKY transcription factor located in this region, was prioritized as the prime candidate gene. Polymorphism analysis of HaWRKY48 in 138 sunflower accessions revealed eight SNPs defining six haplotypes. Resistance was associated with Hap3 and susceptibility to Hap1/Hap6. These findings advance our understanding of the genetic mechanisms governing sunflower resistance to S. sclerotiorum and provide valuable genetic markers for molecular breeding of resistant cultivars. Full article