Teleost fishes display remarkable diversity in sex determination mechanisms, yet this aspect remains uninvestigated in the family Hexagrammidae. In this study, we performed whole-genome resequencing of Hexagrammos otakii to characterize its sex determination system. Genome-wide association studies and population differentiation index analysis identified chromosome 9 as the sex chromosome, supporting a male heterogametic (XY) system. A 3.11 Mb sex-determining region (SDR) was delineated, containing the potential candidate gene gdf6a. Comparative genomic analysis revealed that the H. otakii sex chromosome is non-homologous to those reported in other Cottioidei species, suggesting an independent origin and a dynamic evolutionary trajectory. In addition, we identified a 99 bp male-specific deletion within the SDR and developed a PCR-based marker that successfully differentiates sex with 100

Sex-controlled breeding holds significant value in aquaculture. By controlling the sex of fish, faster-growing sexes can be preferentially produced, shortening the rearing time to market size. Sebastes schlegelii exhibits a significant female growth advantage, and thus all-female aquaculture can substantially improve economic benefits. The objective of this study was to induce XX pseudo-males in S. schlegelii using exogenous 17 alpha-methyltestosterone (MT) and letrozole (LZ) for all-female breeding, and to clarify the underlying hormonal and transcriptomic mechanisms. S. schlegelii juveniles before sex differentiation were treated with MT at concentrations of 20, 40, 60 mg/kg and LZ at concentrations of 300, 600, 900 mg/kg for 30 and 40 days, respectively. Phenotypic and histological analysis showed that all treatment groups achieved 100 % sex reversal efficiency, with pseudo-males successfully induced. Growth performance results indicated that the 30-day treatment with MT (20 mg/kg) or LZ (300 mg/kg) had no significant difference from the untreated control group, while higher concentrations and longer treatment durations inhibited growth and testicular development. Endocrinology showed that the serum steroid levels (elevated testosterone and 11-ketotestosterone, reduced estradiol) of pseudo-males were closer to those of normal males. Transcriptome analysis showed that the downregulation of genes related to ovarian development (cyp19a1a, foxl2) and the upregulation of genes related to testicular development (amh, sox9, dmrt1) jointly regulated sex reversal, and the pseudo-males induced by both hormones had enriched steroid hormone biosynthesis and TGF-beta signaling pathways. This study establishes a feasible sex reversal strategy in S. schlegelii and provides molecular insights into gonadal differentiation, offering a technical basis for all-female breeding.

Innate immunity in teleost fish relies heavily on granulocytes for host defense. However, how these cells functionally adapt to distinct tissue microenvironments remains poorly understood. To address this, we constructed a multi-tissue immune atlas for Cynoglossus semilaevis by integrating single-cell RNA sequencing (scRNA-seq) data from blood, liver, spleen, and head kidney. We identified five transcriptionally distinct granulocyte subsets (Gr1-Gr5), each with divergent roles in inflammation, immunoregulation, and complement activity; moreover, these subsets exhibited tissue-specific distributions. Pseudotime analysis revealed a bifurcated developmental trajectory from a shared progenitor pool residing in the head kidney and spleen, characterized by the dynamic expression of the cxcr2/cxcr4 axis and branch-specific transcription factors. While sharing core metabolic pathways, granulocyte subsets displayed tissue-specific adaptations and an inferred communication strategy involving a conserved cd44b-mediated module. Our findings systematically characterize the patterns of granulocyte adaptation in a teleost model, providing insights into fish immunology and aquaculture disease control.

Sexual size dimorphism (SSD) refers to the differences in body size between males and females. Epigenetic mechanisms, such as DNA methylation, can shape phenotypes; however, their role in SSD remains unclear. Here, we used Cynoglossus semilaevis, a species with larger females (female-biased SSD), as a model to investigate the role of methylation in the emergence and regulation of SSD. Methylomes and transcriptomes were constructed in four tissues (brain, liver, muscle, and gonad) from three key life stages: juveniles (3 months post-fertilization), male mature stage (1 year post-fertilization; ypf), and female mature stage (3 ypf). In 1 ypf and 3 ypf females showed lower methylation levels, while the number of differentially methylated regions between sexes increased over the lifetime. Genes at the top of the growth hormone/insulin-like growth factor (GH/IGF) axis and hypothalamus-pituitary-gonadal axis (gh1, igf1, gnrh3, errγ, etc.) showed differential methylation between sexes, consistent with sex-related differences in growth. The female-specific W chromosome showed higher methylation than the autosomes or the Z chromosome. Genes of the IGF signaling network that negatively regulate growth and are located on the W chromosome were more highly methylated than their Z homologs. Furthermore, genes with a faster evolutionary rate in C. semilaevis exhibited lower methylation levels than the background genes, suggesting an important evolutionary role for DNA methylation in shaping SSD. Our results provide a comprehensive depiction of methylation regulation of SSD in vertebrates and improve our understanding of how methylation regulation can help organisms to respond to natural selection and allocate resources between the sexes.

Developing effective salinity acclimation strategies can improve the welfare and farming efficiency of juvenile rainbow trout in mariculture. This study employed three acclimation groups and a freshwater control group: gradient salinity treatment (SG), acute salinity treatment (SA), gradient salinity with hydrocortisone supplementation treatment (SG_H), no salinity treatment (SC). The aim is to evaluate shared and specific adaptive mechanisms among different strategies through analysis of production performance, physiological and biochemical indicators, and gene expression. Results showed that survival rates and feed conversion efficiency were lower in all salinity-treated groups compared to the control group. At 7d, Mg2 + concentrations were significantly lower in all salinity-treated groups compared to the SC (P < 0.05). while the urea nitrogen and Na+ concentrations were higher than those in the SC group. Transcriptome analysis indicates that ion transport and nitrogen metabolism as shared regulatory mechanisms across salinity treatments (SG_vs_SC, SA_vs_SC), driven by upregulation of genes including trpm7, cftr, cps1, atp1a1. The SG group exhibited higher survival rates, feed conversion efficiency, ATP content than the SA group. KEGG analysis revealed significant enrichment in pathways related to energy metabolism, amino acid biosynthesis, driven by the significant upregulation of gstp, idh3a, and gulb genes. The SG_H group showed feed conversion efficiency was lower than the SG and SA groups. KEGG analysis indicated significant enrichment of the intestinal immune network for IgA production and Cytokine-cytokine receptor interaction pathways. This study provides molecular insights for developing salinity acclimation strategies in the saltwater farming of rainbow trout juveniles.

The rainbow trout (Oncorhynchus mykiss) exhibits extensive karyotypic diversity (2n = 58-64) driven by Robertsonian translocations, yet widely used reference genomes are derived from North American lineages, leaving Chinese aquaculture populations underrepresented. Here, we present a near telomere-to-telomere (T2T) genome assembly of a farmed rainbow trout from China. Integrating PacBio HiFi, ONT ultra-long reads, and Hi-C data, we assembled a 2.29 Gb genome with 99.04% anchored to 30 chromosomes. Notably, the genome contains only 18 gaps, with 16 gap-free chromosomes and 13 achieving T2T status. Comparative synteny analysis revealed a third chromosomal fission/fusion iteration in which Swanson Omy14 splits into Arlee Omy14 and Omy32. Annotation identified 43,137 protein-coding genes, with a BUSCO completeness of 98.9%. This dataset provides a valuable resource for resolving lineage-specific structural variation, supporting pangenome construction and facilitating molecular breeding in rainbow trout.

Atlantic salmon (Salmo salar) is a major aquaculture species and an important model for studying salmonid evolution. However, the highly repetitive genome and extensive duplicated regions retained from the salmonid-specific fourth whole-genome duplication (Ss4R) have impeded complete genome assembly, leaving unresolved gaps and structurally ambiguous regions in previous references. Here, we present a gap-free genome assembly of salmon generated using PacBio HiFi, Oxford Nanopore (ONT) ultra-long reads, and Hi-C data. The 2.81 Gb assembly is anchored to 29 chromosomes with contig N50 of 110.45 Mb. Notably, 8 chromosomes are resolved to a complete T2T level. Quality assessments confirm the high accuracy of the assembly, yielding a quality value of 51.06 and the genome BUSCO completeness of 99.67%. Gene annotation identified 45,163 protein-coding genes with BUSCO completeness of 98.80%. By improving continuity and completeness in repetitive and duplicated regions, this resource provides an enhanced reference for structural variant discovery, comparative genomics, and breeding applications in Atlantic salmon.

Aquatic germplasm resources refer to genetic materials of aquatic animals and plants that possess actual or potential value, including species, subspecies, varieties, strains, etc. These resources hold significant economic, ecological, and scientific importance in the fields of fisheries and agriculture, serving as the foundation for aquaculture, genetic improvement, and biodiversity conservation. Aquatic germplasm resources encompass aquatic organisms such as fish, shrimp, shellfish, algae, and echinoderms, along with their genetic material. They are not only a core element supporting the sustainable development and international competitiveness of the aquaculture industry but also a strategic resource for safeguarding national food security, ecological security, and biodiversity. Building strengthened protection of aquatic germplasm resources, how to enhance the scientific utilization and innovative development of superior aquatic germplasm is both an urgent national need for biological and genetic diversity security and a pressing demand for high-quality, healthy aquatic products driven by growing public expectations. This review conducts a safety assessment from three perspectives: fundamental research on the utilization of aquatic germplasm resources, relevant technologies and platforms, and artificially cultivated and genetically improved germplasm. Based on a review of research progress in these areas, it analyzes existing problems within these three aspects and proposes recommendations for improving the utilization of aquatic germplasm resources. Future recommendations for enhancing the safety of aquatic germplasm resource utilization in China mainly include: strengthening the construction of a shared genomic resource platform for aquatic species based on China's independent intellectual property; increasing efforts to decipher the genetic mechanisms underlying economically important traits; advancing the development of low-cost, high-throughput, and intelligent technologies for precise phenotyping and genotyping, along with platforms for precise functional gene manipulation; and enhancing science popularization regarding transgenic and gene editing technologies while strengthening the protection of intellectual property rights for new varieties.

Salinity difference is a significant characteristic that distinguishes marine and terrestrial aquatic ecosystems. The carbonic anhydrase (CA) plays an important role in responding to the salinity changes in organisms, including regulating ion transport, osmotic balance, and internal environmental homeostasis. However, there is little research on the multiple ca gene family members under salinity stress. Therefore, we analyzed and researched the ca gene family members under different salinity conditions (0 %o and 30 %o) in a euryhaline fish of marine medaka (Oryzias melastigma). In this study, we have screened and identified 18 members of the family in its genome, which are scattered across 10 chromosomes. Although some members had multiple copies, for example, ca16 with 3 copies, there was no homologous pair among the ca genes. These proteins were localized in different cellular compartments (cytoplasm, membrane, mitochondrion, extracellular space, and nucleus) and exhibited alpha helices, beta sheets, and loops, with all containing at least one CA catalytic domain. According to the results of transcriptome and qPCR, we concluded that CA1, CA5A, CA12, CAR12, and CAHZA may respond to low salinity stress. Furthermore, RPF2 may be an important interacting protein with these CA proteins. Our results provide a molecular basis for the salinity domestication of fishes.

The whitespotted conger (Conger myriaster) is an ecologically and economically significant benthic marine species widely distributed across East Asia’s coastal waters. Despite this importance, the genomic resources for this species remain limited, hindering evolutionary and aquaculture research. Here, we present the first high-quality chromosome-level genome assembly of C. myriaster using PacBio CLR, WGS, 10X Genomics and Hi-C data. The resulting 1.09 Gb genome assembly exhibits excellent contiguity, with 97.49% of sequences anchored onto 19 chromosomes. The assembled genome achieved a BUSCO completeness stands at 98.00%, containing 34.80% repetitive sequences and 24,063 predicted protein-coding genes. This foundational genomic resource overcomes a major limitation, providing the essential framework for future investigations into the evolutionary adaptations and for the genetic improvement of C. myriaster in aquaculture.

Sexual size dimorphism (SSD) refers to the differences in body size between males and females. Epigenetics such as methylation contribute to shaping phenotypes, nevertheless, their role in SSD is still unclear. Here, we used Cynoglossus semilaevis , a species with larger females known as female-biased SSD, as a model to investigate the role of methylation in the emergence and regulation of SSD. Methylomes and transcriptomes were constructed in four tissues (brain, liver, muscle, and gonad) from three key life stages: juveniles (3-months post fertilization), male mature stage (1-year post fertilization; ypf), and female mature stage (3 ypf). In 1 ypf and 3 ypf females showed lower methylation, while the number of differentially methylated regions between sexes increased during the lifetime. Genes at the top of the growth hormone/insulin-like growth factor (GH/IGF) axis and hypothalamus-pituitary-gonadal axis ( gh1 , igf1 , gnrhr2r-like , err γ, etc .) showed differential methylation between sexes, consistent with sex-related differences in growth. The female-specific W chromosome showed higher methylation than the autosomes or the Z chromosome. Genes of the IGF signaling network that negatively regulate growth and located on W chromosome were hyper-methylated than their Z homologs. Furthermore, genes with a faster evolutionary rate in C. semilaevis exhibited lower methylation levels than the background genes, suggesting an important evolutionary role for DNA methylation in shaping SSD. Our results provide a comprehensive depiction of methylation regulation of SSD in vertebrates, and improve our understanding of how methylation regulation can help organisms to respond to natural selection, and allocate resources between the sexes. ### Competing Interest Statement The authors have declared no competing interest.

The tiger puffer fish (Takifugu rubripes) is highly valued both economically and for its culinary appeal. However, it currently faces a significant risk of disease. This study aimed to establish a spleen cell line to aid in disease prevention and control for this species. We established a new stable cell line (TRSC) from the spleen tissue of the tiger puffer fish. The TRSC cells exhibited stable growth over more than 90 generations in L-15 medium supplemented with 4 ng/mL recombinant human basic fibroblast growth factor (bFGF) and 2 ng/mL recombinant human leukemia inhibitory factor (LIF) at 24 degrees C. Chromosomal analysis revealed a diploid number of 2n = 44. The TRSC cells were susceptibly infected by both viral hemorrhagic septicemia virus (VHSV) and infectious hematopoietic necrosis virus (IHNV), exhibiting a clear cytopathic effect (CPE). After 48 h of infection, alterations in cell morphology and disintegration were observed. Quantitative reverse-transcription polymerase chain reaction (qRT-PCR) analysis revealed differential replication of the two viruses in TRSC cells. Transcriptome sequencing identified 1308 differentially expressed genes (DEGs) following IHNV infection, with 668 genes upregulated and 640 downregulated. These DEGs were significantly enriched in MAPK signaling pathway, p53 signaling pathway, and DNA replication and so on. The establishment of the TRSC cell line provides valuable insights into viral infection mechanisms, aids in the identification of key immune genes, and offers a more effective approach to disease control in aquaculture, supporting the sustainable development of the industry.

Flow velocity is a critical environmental factor influencing the growth, energy metabolism, and physiological health of aquaculture species. This study investigated the physiological and molecular responses of spotted sea bass (Lateolabrax maculatus) under experimental conditions simulating flow velocities typical of land-based recirculating aquaculture systems (RAS) and deep-sea cage systems. High flow velocities (HFV, 0.35-0.65 body lengths per second [BL/s]) enhanced growth performance compared to low flow velocity (LFV, 2.28-2.85 BL/s) conditions. Histological analysis revealed reduced hepatic lipid accumulation under HFV, while LFV promoted lipid storage. Serum analyses showed elevated antioxidant enzyme activity in the LFV group but higher oxidative stress markers in the HFV group. Transcriptomic profiling identified foxo3 as a key regulatory hub orchestrating metabolic and oxidative stress adaptations. Genes associated with oxidative damage repair, lipid catabolism, and glucose metabolism were significantly enriched under hydrodynamic stress. Enrichment of the FoxO signaling pathway highlighted its central role in mediating oxidative stress mitigation and energy mobilization. These findings demonstrate the dual effects of flow velocity, where higher velocities promote growth and metabolic activity at the cost of oxidative stress, and lower velocities conserve energy while maintaining oxidative stability. Tailored flow velocity conditions can optimize fish welfare and productivity across aquaculture systems. Future studies should investigate the systemic effects of hydrodynamic stress using multi-omics approaches to advance sustainable aquaculture practices.

The Yadong trout (Salmo trutta), a species endemic to the Yatung River in Tibet, China, was classified as a second-class protected species in the 20th century. Now, it is considered one of the most important fishery resources in China. In this study, we assembled a near-complete genome of the S. trutta, integrating PacBio HiFi, Hi-C, and ONT sequencing technologies. The genome assembly spans 2.49 Gb, with 96.87% of the sequence anchored onto 40 chromosomes. In this assembly, a total of 12 chromosomes were assembled to a gap-free level, with 8 of them reaching the telomere-to-telomere level. The completeness of this assembly was assessed at 99.50% by BUSCO, containing approximately 63.24% repetitive sequences, and predicted to encode 41,782 protein-coding genes. This is the first near-complete genome assembly of the S. trutta, providing an essential resource for molecular breeding and germplasm conservation of this important species.

Pufferfish exhibit the smallest vertebrate genomes, making them ideal models for investigating evolutionary patterns and processes that affect genome size. While the Takifugu rubripes genome was fully sequenced two decades ago, key evolutionary drivers remain elusive. We sequenced 10 pufferfish genomes and generated 35 transcriptomes and 13 methylomes to understand genomic evolutionary mechanisms. Comparative genomics revealed that transposable element suppression-rather than lineage-specific conserved element loss-primarily underlies genome compaction. This is mediated by reductions in transposon-associated enzymes that limit transposable element propagation and modify DNA repair mechanisms that promote genomic streamlining. Based on resolved phylogeny among nine Takifugu, it is found that introgression drives speciation of T. niphobles and T. oblongus, while long-term linked selection dominates divergence in other species. Positive selection analyses highlighted mechanotransduction pathway genes (integrins, ion channel transport) that are functionally convergent with mammalian lung cell mechanisms, potentially supporting inflation-based anti-predatory strategies. Additionally, positive selection variants in genes that control lineage-specific skin patterning and coloration, which are either selected for or are rewired in regulatory processes, might suggest a role for pigmentation during the rapid speciation of this lineage. This findings shed light on mechanisms enabling extreme vertebrate genome compaction and provide insights for genome engineering applications.

Black rockfish (Sebastes schlegelii) is a marine ovoviviparous teleost that exhibits significant sexual dimorphism, with females growing faster and reaching larger sizes than males. Establishing stable oogonial stem cells (OSCs) is critical for understanding germline stem cell dynamics and facilitating all-female breeding. In this study, we successfully isolated and cultured OSCs from S. schlegelii for 12 passages. These cells exhibited alkaline phosphatase activity, expressed germline marker genes (ddx4, cdh1, klf4), and maintained a diploid karyotype (2n = 48). Transcriptomic comparisons between early (P3) and late (P12) passages revealed significant metabolic dysfunction and cell cycle arrest in the late-passage cells. Specifically, the down-regulation of glutathione-related and glycolysis-related genes (gstm3, gstt1, mgst3, gsta1, gsta4, gsto1, gapdh) and key mitotic regulators (cdk1, chk1, cdk4, e2f3, ccne2, ccnb1) suggested that metabolic imbalance contributes to oxidative stress, resulting in cell cycle inhibition and eventual senescence. This study provides a marine fish model for investigating metabolism-cell cycle interactions in germline stem cells and lays the foundation for future applications in germ cell transplantation and all-female breeding.

Circular RNAs (circRNAs) are a large class of widely expressed RNAs with covalently closed continuous structures. However, it is currently unknown if circRNAs shows allele-specific expression, as are the consequences of genetic variation on their circularization efficiency and subsequent biological function. Here, we propose a novel pipeline, ASE-circRNA, to accurately quantify both circRNA and their related linear RNA for each allele, and then assess the allele-specificity of the expression of a circular RNA. We identified and analyzed allele-specific circRNAs from human tissue, as well as brains from reciprocal crosses between pairs of highly divergent strains of both mice and pigs by next generation sequencing. Droplet digital PCR (ddPCR) was used to confirm the circularization efficiency measured by next generation sequencing. We found that variation in intron sequences affect the circularization efficiency of circRNAs. Furthermore, we demonstrate that a circRNA, circHK1, regulates the expression of POLR2A to influence the rate of cell proliferation. Our study provides new insight into the molecular mechanisms impacted by variation in genome sequence in the origin of human disease and phenotype.

The identification of sex chromosomes is fundamental for exploring the mechanism and evolution of sex determination. Platichthys stellatus, a species exhibiting clear sexual dimorphism and homomorphic chromosome pairs, has received limited research concerning its sex determination mechanisms. Clarifying the sex chromosome of P. stellatus will enhance our understanding of sex chromosome evolution in Pleuronectiformes. This study employed whole-genome resequencing to investigate the sex chromosome and sex determination system in P. stellatus. Notably, Chr23 was identified as the sex chromosome in P. stellatus, with the sex-determining region (SDR) occupying 48.1% of the chromosome and featuring an XX/XY system. Sex chromosome turnover was observed within Pleuronectiformes, with P. stellatus, Verasper variegatus, and Hippoglossus hippoglossus sharing a common ancestral karyotype. No inversions were detected within the SDR of P. stellatus, although chromosomal rearrangements between sex chromosomes and autosomes were identified. Additionally, a sex-specific marker for P. stellatus was ascertained, enabling genetic sex identification, with significant implications for improving breeding programs and aquaculture practices.

DNA methylation is a key epigenetic mechanism orchestrating gene expression networks in many biological processes. Nonetheless, studying the role of specific gene methylation events in fish faces challenges. In this study, we validate the regulation of DNA methylation on empty spiracles homeobox 2 (emx2) expression with decitabine treatment in Chinese tongue sole testis cells. We used the emx2 gene as the target gene and developed a new DNA methylation editing system by fusing dnmt3a with catalytic dead Cas9 (dCas9) and demonstrated its ability for sequence-specific DNA methylation editing. Results revealed that utilizing dCas9-dnmt3a to target emx2 promoter region led to increased DNA methylation levels and decreased emx2 expression in Chinese tongue sole testis cells. More importantly, the DNA methylation editing significantly suppressed the expression of MYC proto-oncogene, bHLH transcription factor (myc), one target gene of emx2. Furthermore, we assessed the off-target effects of dCas9-dnmt3a and confirmed no significant impact on the predicted off-target gene expression. Taken together, we developed the first DNA methylation editing system in marine species and demonstrated its effective editing ability in Chinese tongue sole cells. This provides a new strategy for both epigenetic research and molecular breeding of marine species.