Hybridogenesis is a unique type of reproduction found in hybrids, producing offspring that are partial (hemiclonal) genetic replicas of one parent. Along the southern coast of Hokkaido, Japan, two types of interspecies hybrids have been identified among three Hexagrammos species: H. octogrammus (Hoc), H. otakii (Hot), and H. agrammus (Hag). These hybrids are characterized by exclusively fertile females reproducing via hybridogenesis. During oogenesis of hybrids, the paternal haploid genome (Hag or Hot) is excluded, while only the maternal genome (Hoc) retains and transmits to eggs. This study investigates the mechanism of genome elimination in Hexagrammos hybrids through a comparative analysis of gonadal development in 1-year-old juveniles and adult individuals of hybrids and their parental species. Comparative genomic hybridization on whole-mount gonads from Hoc/Hot and (Hoc/Hag) × Hag hybrids revealed gonocytes containing genomes of both parental species and gonocytes with only Hoc genome, suggesting that genome elimination occurs during early gametogenesis. In Hoc but not in Hot or Hag species, we observed heterochromatin foci enriched with H3K9me3 epigenetic histone modification. Similar foci were found in hybrids, confirming the presence of Hoc genome in their gonocytes. Furthermore, we detected micronuclei in the cytoplasm of gonocytes in parental species and hybrids. Although micronuclei were rare, their frequency was significantly higher in hybrids compared to parental species. Within micronuclei in Hoc/Hot and (Hoc/Hag) × Hag hybrids, we identified Hot and Hag chromosomes or their fragments, respectively. Thus, in Hexagrammos hybrids, genome elimination likely occurs during early gametogenic stages and is accompanied by micronuclei formation.

• Keywords
• comparative genomic hybridization, fertility, gametogenesis, genome, heterochromatin, hybridization, reproduction,
• MeSH
• DNA * genetics   metabolism   MeSH
• Gametogenesis * genetics   physiology   MeSH
• Hybridization, Genetic * MeSH
• Oogenesis MeSH
• Triatominae * genetics   physiology   MeSH
• Animals MeSH
• Check Tag
• Male MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• DNA * MeSH

BACKGROUND AND AIMS: There is an urgent need for effective prognostic biomarkers to better stratify patients with advanced hepatocellular carcinoma (HCC) undergoing systemic therapy. Shallow whole-genome sequencing (sWGS) of cell-free DNA (cfDNA) is a cost-effective approach for assessing circulating tumour DNA (ctDNA), genomic alterations, and fragmentomic patterns. This study aimed to evaluate sWGS-derived biomarkers as predictors of outcomes in advanced HCC patients receiving systemic treatment. METHODS: Pretreatment plasma samples from 134 patients treated with either tyrosine kinase inhibitors (TKIs; n = 83) or immune checkpoint inhibitors (ICIs; n = 51) were analysed using sWGS. Tumour fraction (TF) and copy number alterations (CNAs) were derived using ichorCNA, while DNA fragmentation was assessed using the DELFI approach. Associations with progression-free survival (PFS) and overall survival (OS) were evaluated. RESULTS: High TF was significantly associated with shorter median PFS (3.0 vs. 10.6 months; p<0.001) and OS (7.8 vs. 21.6 months; p=0.02). High fragmentation similarly correlated with reduced PFS (4.7 vs. 10.7 months; p<0.001) and OS (13.1 vs. 22.3 months; p<0.001). In multivariate analysis within the ICI subgroup, high fragmentation and BCLC stage independently predicted shorter PFS and OS. High AFP levels, advanced BCLC stage, and larger tumours correlated with elevated TF. Frequent CNA gains were observed in chromosomal arms 1q and 8q. CONCLUSION: High TF and high fragmentation levels are associated with worse clinical outcomes in advanced HCC patients treated with systemic therapy, making them promising prognostic biomarkers to consider for guiding treatment decisions.

• Keywords
• Circulating cell-free DNA, Circulating tumour DNA, Hepatocellular carcinoma, Immunotherapy, Shallow whole-genome sequencing, Tumour fraction, Tyrosine kinase inhibitors,
• MeSH
• Circulating Tumor DNA * genetics   blood   MeSH
• Adult MeSH
• Carcinoma, Hepatocellular * genetics   drug therapy   pathology   blood   mortality   MeSH
• Immune Checkpoint Inhibitors therapeutic use   MeSH
• Protein Kinase Inhibitors therapeutic use   MeSH
• Middle Aged MeSH
• Humans MeSH
• Biomarkers, Tumor * genetics   blood   MeSH
• Liver Neoplasms * genetics   drug therapy   pathology   blood   mortality   MeSH
• Prognosis MeSH
• Whole Genome Sequencing * methods   MeSH
• Aged, 80 and over MeSH
• Aged MeSH
• DNA Copy Number Variations MeSH
• Treatment Outcome MeSH
• Check Tag
• Adult MeSH
• Middle Aged MeSH
• Humans MeSH
• Male MeSH
• Aged, 80 and over MeSH
• Aged MeSH
• Female MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Circulating Tumor DNA * MeSH
• Immune Checkpoint Inhibitors MeSH
• Protein Kinase Inhibitors MeSH
• Biomarkers, Tumor * MeSH

Male infertility is a multifactorial condition influenced by hormonal, anatomical, genetic, and lifestyle factors. Although the male factor contributes to up to 40% of infertility cases, the exact etiology often remains unknown (so-called idiopathic infertility). Current diagnostic approaches include detailed clinical evaluation, semen analysis, hormonal assessment, genetic testing, and imaging techniques. Significant progress has been achieved in sperm DNA fragmentation testing and oxidative stress evaluation, offering a broader insight into sperm quality. Treatment strategies vary depending on the underlying cause, ranging from pharmacological therapy and surgical interventions to assisted reproductive technologies. Rapid advances in regenerative medicine and gene therapy show promise for novel therapeutic options. A healthy lifestyle and risk factor management are integral to both prevention and treatment.

• Keywords
• Male infertility, spermatogenesis, diagnostics, assisted reproduction, sperm DNA fragmentation, oxidative stress,
• MeSH
• Semen Analysis MeSH
• Humans MeSH
• Infertility, Male * diagnosis   therapy   etiology   MeSH
• Check Tag
• Humans MeSH
• Male MeSH
• Publication type
• Journal Article MeSH
• Review MeSH

BACKGROUND: The quality and yield of DNA specimens are essential for downstream analyses. Complex biological or environmental samples contain interfering compounds requiring advanced protocols for removing salts, small organic molecules, and/or proteins without compromising the DNA fragments distribution in the original sample. High-molecular-weight DNA fragments are often discriminated by standard extraction methods in attempts to remove the highly concentrated inorganic sample components. RESULTS: Here, we introduce a simple device for the simplification, purification, desalting, and buffer exchange of complex biological samples based on electrodialysis in discontinuous electrolyte systems. The device combines a 3D-printed body with two electrolyte reservoirs and commercially available cassettes with dialysis membranes as inserts. Upon applying an electric current via electrodes inside the electrolyte reservoirs, the sample ions, injected into the cassette or one of the electrolyte reservoirs, electromigrate across the dialysis membranes. During this process, small sample ions pass through the membrane and are substituted by the ions of the selected buffers loaded into the electrolyte reservoirs. The larger ions, such as DNA fragments or proteins, stay in the sample reservoir. The performance of the device was evaluated using DNA samples in blood plasma. Several milliliters of a plasma sample were purified in 30 min with 90 % DNA recovery. SIGNIFICANCE: The device enables fast and effective purification of highly complicated samples. It can be easily scaled for the available sample amount (ranging from microliters to milliliters) and the analyte size of interest by selecting appropriate membrane pore sizes. The device's applicability was optimized on DNA samples with high salt content and tested on plasma samples. But it also has the potential for purification/desalting of a wide range of biological compounds, including proteins, peptides, or charged oligosaccharides, as well as organelles, cells, viruses, or bacteria.

• Keywords
• 3D printed device, Buffer replacement, Desalting, Electromigration, Sample purification,
• MeSH
• Printing, Three-Dimensional * MeSH
• Dialysis instrumentation   MeSH
• DNA * blood   isolation & purification   MeSH
• Electrochemical Techniques * instrumentation   MeSH
• Electrolytes * chemistry   MeSH
• Humans MeSH
• Buffers MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• DNA * MeSH
• Electrolytes * MeSH
• Buffers MeSH

Even after decades of research on diversification in the Neotropics, our understanding of the evolutionary processes that shaped Neotropical clades is still incomplete. In the present study, we used different divergence times and likelihood-based methods to investigate the influence of biogeography and host plant associations on the diversification of the most species-rich psyllid genus Melanastera (Liviidae) from the Neotropics as a model group of herbivorous insects. We used molecular phylogenetic data from seven gene fragments (four mitochondrial and three nuclear). The putatively monophyletic group of Neotropical Melanastera species has an estimated crown node age of 20.2 Ma (ML, CI 20.2-30.6) or 23.2 Ma (BI, 95 % HPD 16.6-32.6), with diversification occurring mainly in the Upper Miocene, although some species groups diversified in the Pliocene or Pleistocene. Biogeographic analysis suggests that the Neotropical Melanastera originated from the Pacific region of South and Central America. We detected a shift in diversification rates that likely occurred either at the time of origin of Melanastera or during the main colonisation of the Atlantic and Amazon Forests, followed by a subsequent slowdown in speciation rates. State-dependent speciation and extinction models revealed a significant relationship between this diversification shift and the shift of Melanastera to the plant families Melastomataceae and Annonaceae, reflecting the impact of host switching on speciation rates in this group. This period also coincides with several independent dispersal events from the Atlantic and Amazon Forests to other parts of the Neotropics. Taken together, the results of the current study suggest that diversification of Melanastera was facilitated by shifts to new host families, which may have promoted the dispersal of Melanastera into new adaptive zones with subsequent processes of local speciation.

• Keywords
• Adaptive radiation, Biogeography, Brazil, Dispersal, Diversification, Herbivorous insects, Host shifts, Jumping plant lice,
• MeSH
• Biological Evolution MeSH
• Phylogeny * MeSH
• Phylogeography MeSH
• Hemiptera * genetics   classification   MeSH
• DNA, Mitochondrial genetics   MeSH
• Models, Genetic MeSH
• Evolution, Molecular MeSH
• Likelihood Functions MeSH
• Sequence Analysis, DNA MeSH
• Genetic Speciation * MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Geographicals
• South America MeSH
• Central America MeSH
• Names of Substances
• DNA, Mitochondrial MeSH

The use of microfluidic sperm sorting (MFSS) systems in infertility treatment is increasing due to their practicality and ease of use. While often presented as highly effective, their efficacy in patients with varying sperm analysis results remains uncertain. In this study, we evaluated the effectiveness of MFSS compared with the swim-up (SU) technique in terms of oxygen radical levels and spermiogram parameters. Samples from each patient were processed using both methods, followed by assessments of sperm concentration, motility, morphology, DNA integrity, acrosomal status, and mitochondrial membrane potential. Participants were selected based on sperm analysis and categorized as normozoospermic (n = 40) or non-normozoospermic (n = 28). An analysis of separation techniques revealed no significant differences, except for a lower percentage of DNA-fragmented sperm in the MFSS group compared with SU within the non-normozoospermic cohort (SU: 10.0% vs. MFSS: 5.69%, p = 0.027). No differences were observed between SU and MFSS in normozoospermic men. The MFSS method is a simple technique, frequently used in laboratories, that yields good results but does not offer a substantial advantage over SU. The primary benefit of MFSS appears to be a significant reduction in the proportion of sperm with DNA fragmentation compared with SU in patients with abnormal sperm analysis results.

• Keywords
• IVF/ICSI, infertility, sperm analysis, sperm separation, spermatozoa,
• MeSH
• Semen Analysis methods   MeSH
• Adult MeSH
• DNA Fragmentation MeSH
• Sperm Injections, Intracytoplasmic * methods   MeSH
• Humans MeSH
• Membrane Potential, Mitochondrial MeSH
• Microfluidics * methods   MeSH
• Sperm Motility * MeSH
• Infertility, Male therapy   MeSH
• Cell Separation * methods   MeSH
• Spermatozoa * cytology   metabolism   MeSH
• Check Tag
• Adult MeSH
• Humans MeSH
• Male MeSH
• Publication type
• Journal Article MeSH
• Comparative Study MeSH

DNA double-strand breaks (DSBs) are formed in meiosis, so their repair in the homologous recombination (HR) pathway will lead to crossover formation, which is essential for successful chromosome segregation. HR contains 2 subpathways: synthesis-dependent strand annealing (SDSA) that creates noncrossover and double Holliday junction (dHJ) that generates crossovers. RAD-51 is a protein essential to the formation of all products of HR, as it assembles on the processed DSB, allowing the invasion of the single-stranded DNA into a region of homology. RAD-51 is removed by RAD-54.L after invasion to allow for repair to occur. Here, we investigate a separation of function allele of rad-51, rad-51::FLAG, as compared to 2 other RAD-51 alleles: rad-51::degron and GFP::rad-51. rad-51::FLAG displays slowed repair kinetics, resulting in an accumulation of RAD-51 foci. rad-51::FLAG worms also activate the DSB checkpoint, but to a less extant than that of rad-51 null mutants. In a proximity ligation assay, RAD-54.L and RAD-51 show enriched colocalization in rad-51::FLAG germlines (but not in rad-51::degron), consistent with stalling at the strand invasion step in HR. The defects in RAD-51 disassembly in rad-51::FLAG mutants lead to formation of chromosomal fragments, similar in their magnitude to ones observed in rad-51 or rad-54.L null mutants. However, rad-51::FLAG mutants (unlike a rad-51 null, GFP::rad-51 or rad-54.L null mutants) displayed no defects in the formation of crossover-designated sites (via GFP::COSA-1 localization). Given that rad-51::FLAG worms show checkpoint activation and chromosomal fragments, these results suggest that crossover repair concludes normally, while the noncrossover pathway is perturbed. This is strikingly different from rad-51::degron and GFP::rad-51 strains, which are proficient or deficient in both pathways, respectively. These results suggest that noncrossovers vs crossovers have distinct recombination intermediates and diverge prior to RAD-51 disassembly.

• Keywords
• C. elegans, RAD51, crossover, meiosis,
• MeSH
• Alleles MeSH
• Crossing Over, Genetic MeSH
• DNA Breaks, Double-Stranded MeSH
• Homologous Recombination MeSH
• DNA, Cruciform * genetics   metabolism   MeSH
• Meiosis MeSH
• DNA Repair MeSH
• Caenorhabditis elegans Proteins MeSH
• Rad51 Recombinase * genetics   metabolism   MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• DNA, Cruciform * MeSH
• Caenorhabditis elegans Proteins MeSH
• rad-51 protein, C elegans MeSH Browser
• Rad51 Recombinase * MeSH

High-throughput sequencing technologies have advanced RNA virus genomics, but recovering viral genomes from mammalian tissues remains challenging due to the predominance of host RNA. We evaluated two metatranscriptomic workflows to address these challenges. Our results demonstrate that the methods differed significantly in performance, with Method B achieving a 5-fold increase in RNA yield and improved RNA integrity over Method A. These differences resulted in the recovery of 4 complete hepacivirus genomes with Method B compared to fragmented or incomplete genomes with Method A. Additionally, Method B's library preparation workflow, incorporating rRNA depletion, enhanced viral genome recovery by reducing host RNA background. Our novel approach integrates an optimized RNA purification protocol with a customized bioinformatics strategy for improved viral genome recovery. Overall, our findings highlight the critical role of optimized homogenization, RNA purification, and library preparation in metatranscriptomic workflows, facilitating the more effective RNA virus genome recovery from complex mammalian tissues.

• MeSH
• Genome, Viral * genetics   MeSH
• Gene Library MeSH
• Humans MeSH
• RNA, Viral * genetics   isolation & purification   MeSH
• RNA Viruses * genetics   MeSH
• Mammals virology   MeSH
• Transcriptome * MeSH
• High-Throughput Nucleotide Sequencing * methods   MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• RNA, Viral * MeSH

Direct detection of DNA in complex biological samples bears several challenges regarding the selectivity and sensitivity of analyses. Therefore, DNA pre-extraction from bio-fluids is an emerging tool in biologically related fields. Specifically, a newly developing family of liquid biopsy techniques using PCR detection of circulating tumor DNA from blood serum or blood plasma could be significantly improved by harnessing fast and high-throughput DNA sample preparation. To address these needs, a 3D-printed device and a method based on gel electrophoresis combined with electrodialysis for the time-, cost- and labor-efficient preparative separation of DNA fragments from blood was developed. The proposed system also successfully eliminated large DNA fragments from the samples. Recovery for short DNA fragments was reaching up to 80 %. The method was tested on human genomic DNA and blood and blood serum spiked with DNA standards, and it significantly alleviated the signal of matrix DNA.

• Keywords
• Cancer, Circulating tumor DNA (ctDNA), Liquid biopsy, PCR, Preparative gel electrophoresis,
• MeSH
• Printing, Three-Dimensional * economics   MeSH
• Time Factors MeSH
• DNA * blood   isolation & purification   MeSH
• Humans MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• DNA * MeSH

The aim of this study is to examine the influence of food restriction on rabbit ovarian functions. A total of eight females were fed ad libitum (NF), while eight females were subjected to 50% food restriction (RF). One month later, all females were euthanized. Weights and lengths of ovaries and uterine horns were measured. Representative parts of the ovaries were subjected to histomorphometry analysis of folliculogenesis. Granulosa cells were isolated and cell viability, proliferation (accumulation of PCNA, cyclin B1, and BrdU-positive cells), apoptosis (accumulation of bax, caspase 3, and DNA fragmentation) were evaluated. Granulosa cells were subjected to proteomic analysis by using the nano HPLC-Chip-MS/MS method. Estradiol and progesterone release by ovarian and granulosa cells was assessed by ELISA. Ovarian and uterine horn weights were lower in RF than NF. The diameter of follicles and oocytes and the thickness of the theca and granulosa cells were higher in RF than NF. RF showed a lower percentage of cells containing bax and caspase 3, occurrence of DNA fragmented cells, and estradiol and progesterone. RF had higher incorporation of BrdU, a higher proportion of cells containing PCNA and cyclin B1, and a lower percentage of viable cells. RF produced more specific proteins than NF, including peptides involved in cell differentiation, proliferation/division, mitotic cell cycle, and GTP-ase activity. In conclusion, food restriction can activate reproduction by (1) selection of the growing primordial follicles, (2) better transformation of secondary to preovulatory follicles, (3) increasing growth of oocytes, (4) increasing proliferation and decreasing apoptosis in granulosa cells, (5) changes in ovarian secretory activity, and (6) changes in the number of peptides.

• Keywords
• apoptosis, food restriction, hormone, ovarian follicle, rabbit,
• Publication type
• Journal Article MeSH