Search Results (906)

Gametogenesis is a fundamental biological process that ensures both genetic recombination and the continuity of successive generations. Interspecific hybrids can reproduce through modified mechanisms, such as hybridogenesis, by transmitting clonal, unrecombined genomes of only one of the parental species via their gametes. Pelophylax grafi (RP) is a natural hybrid frog composed of mixed genomes (subgenomes) of two related species, Pelophylax perezi (P) and Pelophylax ridibundus (R), and coexists in populations with P. perezi. This study tested the involvement of programmed genome elimination in gamete production of P. grafi, providing new insight into reproductive mechanisms of hybrid vertebrates. Using comparative genomic hybridization (CGH) and fluorescent in situ hybridization (FISH), we examined the genomic constitution of germline cells in tadpoles and adult male and female P. grafi. Controlled crosses between P. perezi and P. grafi produced F1 hybrid tadpoles, whose genotypes confirmed that P. grafi parents transmitted the R subgenome through their gametes. In the early germline cells (gonocytes) of these tadpoles, P chromosomes were selectively eliminated via micronuclei formation during interphase. The occasional presence of the R genome and mixed R/P genome micronuclei suggests variability and imperfect fidelity in the elimination process. In adult hybrids, the majority of diplotene oocytes, spermatogonial stem cells (SSC) and spermatocytes carried R subgenomes. We demonstrated that programmed genome rearrangement in Pelophylax hybrids is an evolutionarily conserved mechanism underlying this unique reproductive strategy. Full article

Periodontal disease (PD) is one of the most widespread oral inflammatory diseases in dogs, with the potential to cause systemic consequences. The purpose of this study was to evaluate the inhibitory potential of yeast-derived postbiotics and probiotic bacterial strains by using a previously developed in vitro biofilm model mimicking canine PD-associated biofilm including five bacterial species: Neisseria zoodegmatis, Corynebacterium canis, Porphyromonas cangingivalis, Peptostreptococcus canis, and Enterococcus faecalis. After we confirmed the presence of these five bacterial species by employing Fluorescence In Situ Hybridization, the biofilm inhibitory and eradication activity of 11 yeast-derived postbiotics and probiotic bacterial strains, as well as selected dual biotic combinations, against the polymicrobial biofilm were determined using a modified version of the Calgary Biofilm Pin Lid Device and the crystal violet method; additionally, hemolytic activity was evaluated using canine red blood cells. The results show that the inhibitory activity against the polymicrobial PD biofilm ranged from 0% to 22.55%, and eradication ability varied between 0% and 17.28%; however, when combined, the biotics achieved a maximum inhibition rate of 71%. Probiotic strain BC-05 exhibited the lowest in vitro hemolytic activity. Overall, based on the results, four yeast-derived postbiotics and one probiotic bacterial strain were selected as promising candidates for further evaluation, aiming at in vivo application. Full article

We report a unique pediatric acute myeloid leukemia (AML) case characterized by a CBFB::MYH11 fusion located on a supernumerary ring chromosome 16. Following diagnosis through comprehensive blood and bone marrow assays, the patient was enrolled in the Children’s Oncology Group (COG) study AAML1831 and randomized to the experimental treatment arm (Arm B). She received induction chemotherapy with CPX-351 (liposomal daunorubicin and cytarabine), gemtuzumab and ozogamicin (GO), and the cardioprotectant dexrazoxane and achieved complete remission (CR). The patient completed the treatment with sustained CR for 18 months. This case represents a rare cytogenetic phenomenon that is not well-documented in the current literature. Through a review of relevant publications, we contextualize this case within the spectrum of core binding factor AML (CBF-AML), highlighting diagnostic approaches, treatment strategies, and prognostic implications, particularly in cases involving atypical CBFB::MYH11 fusions. The durable remission observed in this patient, despite the unusual cytogenetic presentation, provides valuable insights into therapeutic management. This report underscores the cytogenetic and molecular heterogeneity of CBFB::MYH11 AML and emphasizes the importance of comprehensive genetic profiling using advanced techniques such as chromosomal microarray and next-generation sequencing. Full article

Periodontitis is a biofilm-driven inflammatory disease in which conventional indices (probing depth, clinical attachment level, and radiographs) quantify tissue destruction without capturing the biology of infection. In this review, we synthesized microbiological diagnostics, from chairside tools to omics. We outline sampling strategies and emphasize the quantitative monitoring of bacterial load. Enzymatic assays (e.g., N-benzoyl-DL-arginine-2-naphthylamide hydrolysis assay test) measure functional activity at the point of care. Immunological methods include rapid immunochromatography for Porphyromonas gingivalis and enzyme-linked immunosorbent assay for the high-throughput measurement of bacterial antigens. Molecular platforms encompass quantitative polymerase chain reaction (qPCR) (TaqMan, SYBR, multiplex panels; propidium monoazide quantitative-qPCR for viable cells), checkerboard DNA–DNA hybridization for semi-quantitative community profiling, loop-mediated isothermal amplification (LAMP)/molecular beacon-LAMP for portable isothermal detection, and microarrays. Complementary modalities such as fluorescent in situ hybridization, next-generation sequencing, and Fourier transform infrared spectroscopy provide spatial, ecological, and biochemical resolutions. We discuss the limitations of current approaches, including sampling bias, presence–activity discordance, semi-quantitation, method biases, limited strain/function resolution, low-biomass artifacts, and lack of validated cutoffs. To address these challenges, we propose a pragmatic hybrid strategy: site-specific quantitative panels combined with activity and host-response markers interpreted alongside clinical metrics under standardized quality assurance/quality control. Priorities include outcome-linked thresholds, strain-aware/functional panels, robust point-of-care chemistry, and harmonized protocols to enable personalized periodontal care. Full article

In this study, SweAMI FISH fluorescence in situ hybridization and immunofluorescence were used to analyze the spatiotemporal expression characteristics of the TBXT gene and Brachyury protein in 16-day-old Hulunbuir sheep embryos and 19-day-old Hu sheep embryos and to explore their regulatory effects on the development of sheep tails and related organs. The study showed the following: At 16 days of embryonic age, the TBXT gene was concentratedly expressed in the heart, somites, neural tube, and mesonephros of both sheep breeds; at 19 days of embryonic age, it was concentratedly expressed in the limb ectoderm and tail bud of Hulunbuir sheep, and in the midgut and tail bud of Hu sheep. At 16 days of embryonic age, the BRACHYURY protein was concentratedly expressed in the neural tube, somites, brain vesicles, and mesonephros of both sheep breeds; at 19 days of embryonic age, it was concentratedly expressed in the heart and hindgut of Hulunbuir sheep, and in the tail bud and hindgut of Hu sheep. In summary, this shows that there are differences in the temporal and spatial expressions of the TBXT gene and BRACHYURY protein between the two sheep breeds. There are also site-specific and time-specific differences in the regulation of the above genes and proteins during tail and related organ development between the two breeds, which confirms that the molecular regulation pathways of tail and related organ development are different between the two breeds. This study provides an experimental basis for screening molecular markers related to goat tail development and breed improvement. Full article

Structural variation (SV) serves as a fundamental driver of phenotypic diversity and environmental adaptation in plants and animals, significantly influencing key agronomic traits in crops. Common wheat (Triticum aestivum L.), an allohexaploid species, harbors extensive chromosomal SVs and distant hybridization-induced recombination events that provide critical resources for genetic improvement. This study utilizes non-denaturing fluorescence in situ hybridization (ND-FISH) and oligonucleotide multiplex probe-based FISH (ONPM-FISH) to analyze the karyotypes of 153 BC₁F₄–BC₁F₆ lines derived from the hybrid line Xiaoyan 7430 and common wheat Yannong 1212. The results revealed that Xiaoyan 7430 carries 8 alien chromosome pairs and 20 wheat chromosome pairs (lacking 6B), and Yannong 1212 contains 21 pairs of wheat chromosomes. The parental lines exhibited presence/absence variations (PAVs) on chromosomes 2A, 6A, 5B, 1D, and 2D. Chromosomal variations, including numerical chromosomal variation (NCV), structural chromosomal variation (SCV), and complex chromosomal variation (CCV), were detected in the progeny lines through ONPM-FISH analysis. The tracking of alien chromosomes over three consecutive generations revealed a significant decrease in transmission frequency, declining from 61.82% in BC₁F₄ to 26.83% in BC₁F₆. Telosomes were also lost during transmission, declining from 21.82% in BC₁F₄ to 9.76% in BC₁F₆. Alien chromosome 1J^S, 4J, and 6J exhibited the highest transmission stability and were detected across all three generations. Association analysis showed that YN-PAV.2A significantly affected the length/width ratio (LWR) and grain diameter (GD); YN-PAV.6A, XY-PAV.6A, and PAV.5B increased six grain traits (+2.25%~15.36%); YN-PAV.1D negatively affected grain length (GL) and grain circumference (GC); and XY-PAV.2D exerted positive effects on thousand-grain weight (TGW). Alien chromosomes differentially modulated grain characteristics: 1J^S and 6J both reduced grain length and grain circumference; 1J^S increased LWR; and 4J negatively impacted TGW, grain width (GW), GD, and grain area (GA). Meanwhile, increasing alien chromosome numbers correlated with progressively stronger negative effects on grain traits. These findings elucidate the genetic mechanisms underlying wheat chromosomal variations induced by distant hybridization and their impact on wheat grain traits, and provide critical intermediate materials for genome design breeding and marker-assisted selection in wheat improvement. Full article

Karyotype analysis and the investigation of chromosomal variations in Populus are challenging due to its small and morphologically similar chromosomes. Despite its utility in chromosome identification and karyotype evolutionary research, satellite DNA (satDNA) remains underutilized in Populus. In the present study, 12 satDNAs were identified from P. trichocarpa, and the copy numbers and chromosomal distributions of each satDNA were analyzed bioinformatically in the reference genomes of P. trichocarpa, P. simonii, and P. nigra. Ten satDNA probes for fluorescence in situ hybridization (FISH) were successfully developed and validated on chromosomes of P. xiaohei (poplar hybrid P. simonii × P. nigra). By integrating bioinformatic genomic satDNA distribution patterns with experimental FISH signals, we constructed a molecular karyotype of P. xiaohei. Comparative analysis revealed errors in current poplar genome assemblies. Comparative karyotype analysis of P. xiaohei and its doubled haploid (DH) lines revealed chromosomal variations in the DH lines relative to the donor tree. The results demonstrate that the newly developed satDNA probes constitute robust cytogenetic tools for detecting structural variations in Populus, while molecular karyotyping provides new insights into the genetic mechanisms underlying chromosome variations in P. xiaohei and the DH plants derived. Full article

Circulating tumor cells (CTCs) are crucial biomarkers for lung cancer metastasis and recurrence, garnering significant clinical attention. Despite this, efficient and cost-effective detection methods remain scarce. Consequently, there is an urgent demand for the development of highly sensitive CTC detection technologies to enhance lung cancer diagnosis and treatment. This study utilized microspheres and A549 cells to model CTCs, assessing the impact of acoustic field forces on cell viability and proliferation and confirming capture efficiency. Subsequently, CTCs from the peripheral blood of patients with lung cancer were captured and identified using fluorescence in situ hybridization, and the results were compared to the immunomagnetic bead method to evaluate the differences between the techniques. Finally, epidermal growth factor receptor (EGFR) mutation analysis was conducted on CTC-positive samples. The findings showed that acoustic microfluidic technology effectively captures microspheres, A549 cells, and CTCs without compromising cell viability or proliferation. Moreover, EGFR mutation analysis successfully identified mutation types in four samples, establishing a basis for personalized targeted therapy. In conclusion, acoustic microfluidic technology preserves cell viability while efficiently capturing CTCs. When integrated with EGFR mutation analysis, it provides robust support for the precise diagnosis and treatment of lung cancer as well as personalized drug therapy. Full article

Background: Although telomere length maintenance is a common characteristic of hematological malignancies, the role of telomere length as a prognostic factor to stratify acute lymphoblastic leukemia (ALL) patients depending on their risk of relapse remains elusive. Methods: This knowledge gap motivated us to examine telomere length values in children with ALL at the time of diagnosis and after treatment using quantitative polymerase chain reaction (qPCR) (n = 35). To achieve high-resolution precision and cell specificity, a quantitative fluorescence in situ hybridization (qFISH) technique was developed (n = 5). Results: The results demonstrated statistically significant evidence of telomere shortening in the lymphoblasts of children with ALL but not in the lymphocytes of children after remission following treatment. Our findings also suggested a significant association between telomere shortening and a high risk of relapse disease. Last but not least, our preliminary results showed a trend that telomere shortening was more pronounced in children with B-ALL compared to those with T-ALL in a non-significant manner. Conclusions: Consequently, the current study provides preliminary insights into the potentially substantial prognostic value of telomere length in the progression of pediatric ALL, with the possibility of predicting treatment response. To clarify the application of telomere length as a possible biomarker for disease progression and treatment response in children with ALL, the telomere length values of additional participants need to be examined in further studies. Full article

The PML::RARA fusion resulting from t(15;17) is the genetic hallmark of acute promyelocytic leukemia (APL), typically detected by cytogenetics and/or fluorescence in situ hybridization (FISH) studies. Rarely, APL patients present with normal cytogenetics and FISH findings, complicating diagnosis and delaying life-saving therapy. We report a 23-year-old male with clinical, morphologic and immunophenotypic features consistent with APL but negative for FISH studies. Despite prompt initiation of all-trans retinoic acid (ATRA) based on clinical suspicion, the patient succumbed to intracranial hemorrhage. Quantitative reverse transcriptase PCR (qRT-PCR) confirmed a long isoform PML::RARA fusion. A review of 34 published cytogenetics- and FISH-negative cases since 1995 demonstrates that RT-PCR-based methods reliably detect cryptic fusions. While advanced genomic approaches may identify these fusions at higher resolution, their accessibility, complexity, cost, and turnaround time often limit diagnostic utility in the urgent setting of APL. Given the extreme rarity of this subset, cytogenetics and FISH remain the standard frontline tests; however, these cases underscore the critical need to incorporate molecular testing into routine workflows. Early recognition and timely therapy are essential to reducing mortality in cryptic APL, and these cases also provide insight into mechanisms of atypical leukemia biology. Full article

Background/Objectives: Synovial sarcoma (SS) is an aggressive soft-tissue tumor characterized by the chromosomal translocation t(X;18) (p11.2;q11.2), most commonly involving the fusion of the SYT gene on chromosome 18 with the SSX1 or SSX2 genes on chromosome X. This study aims to explore the clinicopathological and molecular characteristics of synovial sarcoma in a cohort of Moroccan patients. Methods: We analyzed 48 cases of synovial sarcoma using formalin-fixed, paraffin-embedded (FFPE) tissue samples. Histological grading was performed according to the FNCLCC system. Immunohistochemical staining was employed to detect cytokeratin (CK) and epithelial membrane antigen (EMA). Molecular analysis included fluorescence in situ hybridization (FISH) to identify SS18 gene rearrangements and reverse transcription–polymerase chain reaction (RT-PCR) to detect SYT-SSX fusion transcripts. Results: Among the cohort, 56% of cases showed SS18 gene rearrangements via FISH, while RT-PCR confirmed the presence of SS18-SSX1 and SS18-SSX2 transcripts in 60% and 32% of cases, respectively. The remainder was classified as undifferentiated sarcoma. Notably, no significant associations were observed between SYT-SSX fusion type and clinicopathological features. Conclusions: These findings underscore the importance of integrating molecular techniques for precise diagnosis in synovial sarcoma. The results align with global patterns, emphasizing the necessity for molecular testing to enhance diagnostic accuracy and informing potential therapeutic advancements. Full article

Background: Leukemia, a hematologic malignancy, is the major fluid tumor. However, there is a paucity in laboratory characterization in South Africa due to limited diagnostic infrastructure. Chromosomal aberrations play a crucial role in leukemia pathogenesis, influencing classification, prognosis, and treatment. Aim: This study aimed to characterize chromosomal aberrations in leukemia patients using the fluorescence in situ hybridization (FISH) method, with the goal of improving diagnostic precision and guiding tailored treatment in resource-limited settings. Methodology: This study was a retrospective analysis of 349 leukemia patient records from the NHLS Corporate Data Warehouse, covering cases diagnosed between January 2019 and January 2024. Chromosomal aberrations were assessed using FISH, including cases of CML, AML, CLL, and ALL. Results: CML was the most prevalent leukemia subtype (40%), followed by AML (31%). Age-specific distributions were significant across subtypes (p < 0.0001). FISH detected subtype-specific aberrations: t(1;19) and t(12;21) in 25% of ALL cases; t(8;21) and t(15;17) in 22–33% of AML cases; and t(9;22) in 100% of CML cases. In CLL, 13q deletions were most common (53% complex, 33% simple). Conclusions: This study reveals distinct chromosomal aberration patterns in leukemia patients in Gauteng, with CML as the most prevalent subtype. Distinct patterns were observed across ALL, AML, and CLL, with age and gender-specific trends. Findings highlight regional genetic influences, diagnostic gaps, and healthcare challenges, emphasizing the urgent need to expand cytogenetic and molecular testing to enable targeted diagnostics, risk stratification, and personalized therapies in sub-Saharan Africa. Full article

Insights into the state of individual cells within a living organism are essential for identifying diseases and abnormalities. The internal state of a cell is reflected in its morphological features and changes in the localization of intracellular molecules. Using this information, it is possible to infer the state of the cells with high precision. In recent years, technological advancements and improvements in instrument specifications have made large-scale analyses, such as single-cell analysis, more widely accessible. Among these technologies, imaging flow cytometry (IFC) is a high-throughput imaging platform that can simultaneously acquire information from flow cytometry (FCM) and cellular images. While conventional FCM can only obtain fluorescence intensity information corresponding to each detector, IFC can acquire multidimensional information, including cellular morphology and the spatial arrangement of proteins, nucleic acids, and organelles for each imaging channel. This enables the discrimination of cell types and states based on the localization of proteins and organelles, which is difficult to assess accurately using conventional FCM. Because IFC can acquire a large number of single-cell morphological images in a short time, it is well suited for automated classification using machine learning. Furthermore, commercial instruments that combine integrated imaging and cell sorting capabilities have recently become available, enabling the sorting of cells based on their image information. In this review, we specifically highlight practical applications of IFC in four representative areas: cell cycle analysis, protein localization analysis, immunological synapse formation, and the detection of leukemic cells. In addition, particular emphasis is placed on applications that directly contribute to elucidating molecular mechanisms, thereby distinguishing this review from previous general overviews of IFC. IFC enables the estimation of cell cycle phases from large numbers of acquired cellular images using machine learning, thereby allowing more precise cell cycle analysis. Moreover, IFC has been applied to investigate intracellular survival and differentiation signals triggered by external stimuli, to monitor DNA damage responses such as γH2AX foci formation, and more recently, to detect immune synapse formation among interacting cells within large populations and to analyze these interactions at the molecular level. In hematological malignancies, IFC combined with fluorescence in situ hybridization (FISH) enables high-throughput detection of chromosomal abnormalities, such as BCR-ABL1 translocations. These advances demonstrate that IFC provides not only morphological and functional insights but also clinically relevant genomic information at the single-cell level. By summarizing these unique applications, this review aims to complement existing publications and provide researchers with practical insights into how IFC can be implemented in both basic and translational research. Full article

Allograft inflammatory factor-1 (AIF1) is a conserved calcium-binding protein involved in inflammatory and neuro-immune responses and expressed in Pomacea canaliculata (Pc-AIF1) during cephalic tentacle regeneration. Here, we investigated the expression and distribution of Pc-AIF1 in control conditions and during cephalic tentacle regeneration. A transcriptomic analysis of 315 RNA-seq datasets revealed maximal Pc-AIF1 expression in circulating hemocytes and hemocyte-rich tissues. Pc-AIF1 was also highly expressed in neural ganglia. Fluorescence in situ hybridization (FISH) evidenced Pc-AIF1 in circulating hemocytes and in the phagocytic hemocyte aggregates in the posterior kidney. qPCR showed the constitutive expression of Pc-AIF1 in cerebral ganglia. FISH experiments showed Pc-AIF1-positive cells within the cephalic tentacle blastema at 24 h post-amputation (hpa). Even if the amputation left them untouched, both the ipsilateral and contralateral cerebral ganglia increased Pc-AIF1 expression until 48 hpa. Immunocytochemical experiments evidenced positive cells to RCA120 (a microglial marker in mammals) among circulating hemocytes, in the connective tissue surrounding the cerebral ganglia, and within the regenerating tentacles. These findings suggest that Pc-AIF1 is a neuro-immune marker constitutively expressed in hemocyte populations and neural tissues; it is associated with the immediate hemocyte response to wounding and the neuro-immune interplay during the regeneration of sensory organs. 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