Search Results (234)

Long-term monitoring of tree crown condition is essential for assessing forest resilience under increasing climatic variability. This study presents a comprehensive evaluation of oak (Quercus spp.) defoliation trends in Poland from 2015 to 2024, based on national forest health monitoring data. Mean defoliation remained relatively stable until 2018, followed by a significant increase in 2019 (+5.1 percentage points; p < 0.001), coinciding with a major drought event across Central Europe. In subsequent years, defoliation gradually decreased and stabilised, indicating partial canopy recovery. Segmented regression and spline models revealed a consistent breakpoint in 2019 across all age classes, with the most severe crown damage recorded in stands older than 100 years. Younger stands showed lower defoliation levels and higher regenerative capacity. A nonlinear relationship between defoliation and growing-season precipitation was also identified, showing that when rainfall fell below 40 mm, canopy loss exceeded 30%. The results confirm that oak defoliation reflects both short-term climatic stress and long-term structural changes. Integrating monitoring data with climatic analyses and statistical modelling improves the detection of stress-related drivers and the assessment of recovery processes. The combined use of these approaches supports adaptive forest management strategies, including the promotion of mixed-species and multi-aged stands, improvement of soil nutrient conditions, and targeted monitoring of drought-sensitive age classes, thereby enhancing the resilience of oak ecosystems to climate change. Full article

Watermelon chlorotic stunt virus (WmCSV), a bipartite begomovirus, poses a severe and expanding threat to global cucurbit and watermelon production, driving an urgent need to unravel its genetic diversity indices and evolutionary complexities. To elucidate its evolutionary history, this study investigated the genetic diversity, evolutionary dynamics, and global dispersal of its genomic components, DNA-A (WmA) and DNA-B (WmB). The analyses uncovered striking contrasts between the components. WmB exhibited markedly greater genetic diversity (π = 0.0508 vs. 0.0119 for WmA), a slightly faster evolutionary rate (1.26 × 10⁻⁴ vs. 1.44 × 10⁻³ substitutions/site/year), and a far more complex recombination profile, with 34 events detected compared to only one in WmA. The abundance of recombination breakpoints in WmB underscores its central role in shaping genomic variability and adaptive potential. Phylogenetic analyses of both components unveil eight robustly supported clades per segment, predominantly shaped by geographical boundaries, hinting at localized evolutionary trajectories with constrained long-distance gene flow, with the exception of Oman. Bayesian time-scaled phylogenies and phylogeographic reconstructions further illuminate distinct dissemination pathways, suggesting an intriguing origin, with WmA likely emerging from the United States and WmB tracing back to Saudi Arabia, while the Middle East emerges as a dynamic epicenter for regional spread and subsequent incursions into the Americas. Together, these findings reveal contrasting evolutionary forces driving WmCSV diversification and provide critical insights into its origins and ongoing global emergence. Full article

Background/Objectives: Salmonella enterica is a major cause of foodborne infection globally, with poultry acting as an important reservoir. However, data from Central Asia remain limited. This study provides preliminary phenotypic and genomic characterization of S. enterica isolates recovered from poultry farms in southern Kazakhstan, focusing on antimicrobial resistance (AMR), serotypes/sequence types and phylogenetic relationships. Methods: In October 2024, 335 poultry and environmental samples were collected from three regions of southern Kazakhstan using a cross-sectional, detection-focused sampling strategy. Isolation of Salmonella enterica followed enrichment and selective culturing, with confirmation by biochemical assays, slide agglutination serology and real-time PCR. Antimicrobial susceptibility testing was performed using the Kirby-Bauer disk diffusion method and interpreted according to CLSI veterinary breakpoints (VET01/VET08) and CLSI M100 where veterinary criteria were unavailable. Whole-genome sequencing (Illumina) was used for in silico serotyping, MLST, AMR gene detection, plasmid replicon typing and SNP-based phylogenetic reconstruction. Results: Nine S. enterica isolates were confirmed (overall yield 2.7%; 9/335), comprising S. Enteritidis (ST11; n = 4), S. Infantis (ST32; n = 3) and ST68 (n = 2; Choleraesuis/Paratyphi C lineage). All isolates were resistant to ciprofloxacin, and most displayed resistance to ampicillin, gentamicin and trimethoprim-sulfamethoxazole. Plasmid-associated AMR determinants, including blaTEM-116, tet(A), sul1 and dfrA14, were frequently identified on IncF-type replicons. Phylogenetic analysis revealed that the isolates clustered with previously described Eurasian poultry-associated lineages. Conclusions: In this small, exploratory sample from poultry farms in southern Kazakhstan, all recovered S. enterica isolates were multidrug-resistant, with universal fluoroquinolone resistance and frequent plasmid-borne AMR genes. These preliminary findings provide baseline genomic evidence and highlight the need for broader, harmonized AMR surveillance in the regional poultry sector. Full article

Accurate detection of oncogenic gene fusions is becoming increasingly important given the availability of highly effective targeted therapies. However, their identification in clinical practice remains challenging due to the rarity of individual events, diversity of partner genes, and variability of breakpoint locations. Conventional approaches such as immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) lack multiplexing capacity and demonstrate variable sensitivity and specificity, while direct identification of fusion transcripts in whole-transcriptome sequencing (RNA-seq) profiles provides broader applicability but limited sensitivity, as fusion junctions are frequently supported by a minimal number of reads or even no reads at all. In this study, a novel approach was employed to accurately detect clinically actionable RET (REarranged during Transfection) fusions. This approach entailed the measurement of the imbalance in RNA-seq read coverage of potential fusion oncogenes at their 3′ and 5′ exons. A total of 1327 experimental solid tumor RNA-seq profiles were screened, including 154 non-small cell lung cancer and 221 thyroid cancer samples. The RET status was validated in 78 selected cases by targeted NGS and Sanger sequencing. An analysis of the coverage imbalance was conducted, which enabled the accurate discrimination between true and false positive RET fusions. This approach outperformed other methods and yielded 100% sensitivity and specificity with optimized thresholds. The findings were validated using an independent cohort of 79 thyroid cancer cases, confirming the reliability of the results. Among the 18 RET fusion-positive samples, one was identified as an extremely rare case (RUFY3::RET), and two were determined to be novel fusions (FN1::RET, PPP1R21::RET). The findings of this study demonstrate that exon coverage imbalance analysis serves as a robust complement to computational RNA-seq analysis pipelines for the detection of clinically relevant RET fusions. Full article

Background/Objectives: Short-read-sequencing (SRS) is currently the standard for genetic testing in inherited human diseases. Intrinsic limitations include PCR dependency, restricted read length, and challenges in identifying structural variants (SVs), copy number variations (CNVs), and intronic small variants (SNVs/indels). Long-read-sequencing (LRS) enables the sequencing of long DNA molecules, detection of deep intronic variants, rapid testing of few samples, and improved resolution of SVs, CNVs, and SNVs/indels. We therefore aimed to validate Oxford Nanopore Technologies (ONT) LRS for potential clinical application. Methods: We evaluated the ONT’s ability to detect pathogenic/likely pathogenic (P/LP) variants previously identified by SRS and confirmed via Sanger sequencing, Multiplex-Ligation-dependent-Probe-Amplification (MLPA), or quantitative-PCR (qPCR). In total, 509 samples were analyzed, including 393 with P/LP variants and 116 negative controls. We used CE-IVD panels HEVA pro, CARDIO pro, BRaCA panel, and ClinEX pro (4Bases-CH). Sequencing was performed on MinION, GridION, and PromethION-2 platforms. Data were analyzed using the 4eVAR pipeline. Results: ONT successfully identified all P/LP variants across the panels (sensitivity 100%); identified a previously missed CNV in ENG gene; precisely defined the breakpoints of a del(13q) (unsuspected and diagnosed as BRCA2 del ex2–14); improved the coverage profiles in difficult-to-map regions (e.g., ex1 TGFBR1, PSM2CL); expanded the coverage of out-of-target deep intronic regions; and allowed for the set-up of fast-track tests (<24 h) for urgent clinical needs. Conclusions: Our findings demonstrate that ONT LRS provides diagnostic performance comparable to SRS, with significant advantages in resolving complex and previously undetectable variants. Ongoing developments are further increasing read length, expanding detectable targets, and potential clinical applications. Full article

This study proposes a hybrid analytical framework for detecting chondromalacia using vibroacoustic (VAG) signals from patients with knee osteoarthritis (OA) and healthy controls (HCs). The methodology combines nonlinear signal decomposition, feature extraction, and deep learning classification. Raw VAG signals, recorded with a custom multi-sensor system during open (OKC) and closed (CKC) kinetic chain knee flexion–extension, underwent preprocessing (denoising, segmentation, normalization). Ensemble Empirical Mode Decomposition (EEMD) was used to isolate Intrinsic Mode Functions (IMFs), and Detrended Fluctuation Analysis (DFA) computed local (α₁) and global (α₂) scaling exponents as well as breakpoint location. Frequency–energy features of IMFs were statistically assessed and selected via Neighborhood Component Analysis (NCA) for support vector machine (SVM) classification. Additionally, reconstructed α₁/α₂-based signals and raw signals were converted into continuous wavelet transform (CWT) scalograms, classified with convolutional neural networks (CNNs) at two resolutions. The SVM approach achieved the best performance in CKC conditions (accuracy 0.87, AUC 0.91). CNN classification on CWT scalograms also demonstrated robust OA/HC discrimination with acceptable computational times at higher resolutions. Results suggest that combining multiscale decomposition, nonlinear fluctuation analysis, and deep learning enables accurate, non-invasive detection of cartilage degeneration, with potential for early knee pathology diagnosis. Full article

Inland lakes across the United States are increasingly impacted by nutrient pollution, sedimentation, and algal blooms, with significant ecological and economic consequences. While satellite-based monitoring has advanced our ability to assess water quality at scale, many lakes remain analytically underserved due to their spatial heterogeneity and the multivariate nature of pollution dynamics. This study presents an integrated framework for detecting spatiotemporal pollution patterns using satellite remote sensing, trend segmentation, hierarchical clustering and dimensionality reduction. Taking Horseshoe Lake (Illinois), a shallow eutrophic–turbid system, as a case study, we analyzed Sentinel-2 imagery from 2020–2024 to derive chlorophyll-a (NDCI), turbidity (NDTI), and total phosphorus (TP) across five hydrologically distinct zones. Breakpoint detection and modified Mann–Kendall tests revealed both abrupt and seasonal trend shifts, while correlation and hierarchical clustering uncovered inter-zone relationships. To identify lake-wide pollution windows, we applied Kernel PCA to generate a composite pollution index, aligned with the count of increasing trend segments. Two peak pollution periods, late 2022 and late 2023, were identified, with Regions 1 and 5 consistently showing high values across all indicators. Spatial maps linked these hotspots to urban runoff and legacy impacts. The framework captures both acute and chronic stress zones and enables targeted seasonal diagnostics. The approach demonstrates a scalable and transferable method for pollution monitoring in morphologically complex lakes and supports more targeted, region-specific water management strategies. Full article

Virtual Machine Introspection (VMI) is a powerful technology used to detect and analyze malicious software inside Virtual Machines (VMs) from the outside. Asynchronous access to the VM’s memory can be insufficient for efficient monitoring of what is happening inside of a VM. Active VMI introduces breakpoints to intercept VM execution at relevant points. Especially for frequently visited breakpoints, and even more so for production systems, it is crucial to keep performance overhead as low as possible. In this paper, we present an empirical study that compares the performance of four VMI breakpoint-implementation variants—EPT switching (SLAT view switching) with and without fast single-stepping acceleration, instruction repair, and instruction emulation—from two VMI applications (DRAKVUF, SmartVMI) with the XEN hypervisor on 20 Intel Core i processors ranging from the fourth to the thirteenth generation. Instruction emulation was the fastest method across all 20 tested platforms. Modern processors such as the Intel Core i7 12700H and Intel Core i9 13900HX achieved median breakpoint-processing times as low as 15 µs for the emulation mechanism. The slowest method was instruction repair, followed by EPT switching and EPT switching with FSS. The order was the same for all measurements, indicating that this is a strong and generalizable result. Full article

Duchenne muscular dystrophy (DMD) is typically described in boys with a pathogenic variant in the DMD. However, in certain cases, females may also exhibit symptoms of this X-linked disorder. In the present study, the cause of Duchenne muscular dystrophy in three girls was reciprocal translocations t(X;2), t(X;12), and t(X;16), with breakpoints located within the DMD gene sequence. All patients had global development delay, predominantly proximal muscle weakness, calf muscle hypertrophy, and elevated creatine kinase levels up to 100 times the normal range (16,000–26,694 U/L). All underwent cardiac ultrasound and electromyography, and two of the girls also had muscle MRI data. After receiving negative results of MLPA aimed at the detection of DMD deletions and duplications, as well as the limb-girdle muscular dystrophy gene panel sequencing, the patients were referred to whole genome sequencing, which allowed to detect a translocation involving the short arm of the X chromosome and with breakpoints in the DMD. Karyotyping confirmed reciprocal translocations in all patients, with de novo status established in all three cases. The results of this study contribute to the understanding of clinical polymorphism and genetic heterogeneity of the disease, highlighting the importance of a comprehensive approach to genetic diagnostics in atypical cases. Full article

In Brazil, Nyssorhynchus darlingi stands out as the primary malaria vector. Chromosome inversions have long been recognized as critical evolutionary mechanisms in diverse organisms. In this study, we used biallelic SNPs to show that it is possible to detect chromosome inversions reliably with low coverage sequence data. We estimated chromosome inversions in an Amazon Basin sample of Ny. darlingi and compared them with Anopheles gambiae and Anopheles albimanus genomes in synteny analysis. The An. gambiae dataset benchmarked the inversion detection pipeline with known inversions. Genotyping by sequencing was performed using the LCSeqTools workflow for the lcWGS dataset with an average sequencing depth of 2x. A synteny analysis was performed for Ny. darlingi inversions regions with An. gambiae and An. albimanus genomes. The sliding window analysis of PCA components revealed 10 high-confidence candidate regions for chromosome inversions in Ny. darlingi genome and two known inversions for An. gambiae with possible identification of breakpoints and adjacent regions at lower resolution. We demonstrate that lcWGS is a cost-effective and accurate method for detecting chromosome inversions. We reliably detected chromosome inversions in Ny. darlingi from the Brazilian Amazon that does not share similar inversion arrangements in An. gambiae or An. albimanus genomes. Full article

The expanded newborn screening (NBS) program in the Russian Federation, launched in 2023, includes the detection of severe forms of T- and B-cell immunodeficiencies via TREC/KREC quantification. We report a rare case of a male infant having multiple congenital anomalies and lymphopenia identified through this program. Genetic testing revealed a 25.8 Mb terminal deletion spanning 13q31.2–qter, consistent with 13q deletion syndrome. Initial NBS revealed reduced TREC levels, prompting further evaluation. The patient exhibited a complex phenotype, including central nervous system malformation (alobar holoprosencephaly), severe congenital heart disease, renal hypoplasia, limb and genitourinary anomalies, and facial dysmorphism. Postnatal complications included pneumonia, pleuritis, and chylothorax. Flow cytometry demonstrated mild T- and B-cell lymphopenia. The genomic defect was characterized using long-read third-generation sequencing, enabling precise breakpoint identification and accurate mapping of deleted genes. The deletion was confirmed via subtelomeric FISH analysis. The patient died at 7 months of age due to the progression of underlying congenital anomalies and associated complications. Our findings broaden the clinical characterization of distal 13q deletion syndrome and demonstrate the value of long-read sequencing in structural chromosomal analysis. They further highlight the difficulties of caring for neonates having complex malformations and immune dysfunction. Given the potential for both primary and secondary immune disturbances, comprehensive immunological evaluation should be considered in patients having 13q deletion syndrome to improve diagnostic accuracy and inform appropriate clinical management. Full article

Cutibacterium acnes subspecies/phylotypes can cause infections requiring antibiotic therapy. Phylotyping of 73 (55 acneic and 18 non-acneic) C. acnes strains was performed, and antibiotic susceptibility was tested by E tests, breakpoint susceptibility test, or disk diffusion method. The dominant phylotype in both acneic and non-acneic strains was IA1 (56.2%). Phylotype II was >3-fold more frequent in non-acneic than acneic isolates. Resistance in acneic strains was >41% for clindamycin, 36.4% for tetracycline and 15.9% for levofloxacin, and that in non-acneic strains was >38% for clindamycin, 22.2% for tetracycline and 5.6% for levofloxacin. No strain was piperacillin/tazobactam or vancomycin resistant. Amoxicillin resistance was found in both acneic (5.4%) and non-acneic strains (11.1%), and was rare (1.8%) in phylotype I but higher (23.5%) in other strains. Double resistance was found in 32.6% of acneic and 22.2% of the non-acneic strains, and 9.3% of acneic strains displayed multidrug resistance. In conclusion, IA1 phylotype was dominant in both acneic and non-acneic strains, and type II was more frequent in non-acneic isolates. The detection (at >6%) of amoxicillin resistance represents a rare yet important finding. The presence of double/multidrug resistance strongly implies the need of susceptibility-guided therapy of the associated infections. Full article

Background/Objectives: Fluorescence in situ hybridization (FISH) is a standard diagnostic tool for detecting gene fusions and rearrangements in lymphomas but is limited by incomplete genomic coverage, dependence on predefined probes, and difficulty identifying atypical or noncanonical fusion partners. These constraints often result in inconclusive diagnoses in complex lymphoma cases. This study evaluates a novel Hi-C-based sequencing assay from formalin-fixed paraffin-embedded (FFPE) samples to detect clinically significant gene fusions and rearrangements in cases where conventional FISH was inconclusive or expected biomarkers were not detected. Methods: Five diffuse large B-cell lymphoma cases with previously atypical gene fusions or rearrangements by FISH were analyzed using both standard FISH and a Hi-C-based lymphoma assay. Standard FISH was performed using break-apart probes targeting MYC, BCL2, and BCL6, and dual-fusion probes targeting IGH::MYC and IGH::BCL2. The Hi-C assay utilized high-resolution sequencing of FFPE tissue to map chromatin interactions and identify structural variations across the genome and assessment of their clinical relevance. Results: In this series of five lymphoma cases, Hi-C detected additional structural variants beyond those identified by FISH. It identified typical and atypical translocation partners of key oncogenes (MYC, BCL2, BCL6), cryptic breakpoints, and novel genomic events, including TP53 loss, KMT2A amplification, and complex rearrangements, which were undetectable by FISH. The Hi-C assay’s whole-genome coverage enabled comprehensive profiling. Conclusions: The Hi-C-based lymphoma assay offers a transformative diagnostic tool, overcoming FISH limitations by providing unbiased, high-resolution detection of structural variations. This approach enhances diagnostic accuracy and supports personalized therapeutic strategies in lymphoma management, warranting further validation for clinical adoption. Full article

Complex chromosomal rearrangements (CCRs) are rare structural abnormalities involving at least three chromosomal breakpoints and often two or more chromosomes. Owing to their inherent genomic complexity, CCRs are frequently associated with abnormal phenotypes, including developmental delay, congenital anomalies, and infertility. In this study, we report four male patients, three of them with de novo rare structural chromosomal rearrangement detected through a combination of Giemsa-Trypsin (GTG) banding, fluorescence in situ hybridization (FISH), and high-resolution microarray techniques (SNP array and array CGH). Each of the four cases turned out to be of a different type: in addition to two exceptional CCRs, an inv dup del 18q and a cluster rearrangement involving the long arm of chromosome 4 were identified. Despite the limitations of the testing methods, we performed a detailed analysis of the relationship between the most detailed genotype data and the associated phenotype. Our study provides further valuable evidence that the use of molecular cytogenetic methods is of paramount importance even in cases with abnormal karyotypes detected by light microscopy, as high-resolution data may reveal unsuspected genomic complexity, which is essential for genetic counseling in these patients. Full article

Chromosomal BCR-ABL1 fusions are the defining molecular lesions of chronic myeloid leukemia (CML) and Philadelphia-positive acute lymphoblastic leukemia, and are rarely observed in acute myeloid leukemia. Their detection have transformed treatment paradigms by enabling effective use of specific tyrosine kinase inhibitors (TKIs). Although many BCR-ABL1 rearrangements are identified by standard cytogenetics, a clinically relevant subset is cryptic and can escape detection. High-depth RNA sequencing assays have improved our capacity to detect expressed fusion transcripts. Here, we introduce two myeloid cases in which cryptic BCR-ABL1 rearrangements and precise breakpoints detection required an integrated molecular approach: we describe the initial diagnostic pitfalls, detail the downstream therapeutic and prognostic implications and offer practical recommendations for integrating targeted sequencing and cytogenetics into routine practice. In the first case, a patient initially diagnosed with a myelodysplastic/myeloproliferative neoplasm was reclassified as CML following the discovery of a cryptic e13a2 BCR-ABL1 rearrangement, enabling effective TKI treatment. In the second case, a previously undetected BCR-ABL1 fusion was identified in a relapsed AML patient, along with additional molecular lesions, underscoring the aggressive nature of the disease. Our findings support a systematic, multimodal screening strategy in patients with atypical presentations to ensure the timely detection of clinically actionable fusion events. Full article