Aquaculture

Selection for disease resistance forms one of the most highlighted areas of aquaculture breeding. A breeding program for Arctic charr has been operating in Sweden for over 40 years, making it the oldest of its kind worldwide for this species. However, the lack of available genomic resources prevented selection for any disease-resistance traits. A 72k Axiom SNP array was produced in this study and used to assess the potential to select for charr resistant to bacterial kidney disease (BKD), which is currently a major threat to the industry. Following a challenge experiment with Renibacterium salmoninarum, the causative agent of BKD, relevant phenotypic proxies were collected from approximately 2,000 charr. Thereafter, those animals were genotyped with the new 72k SNP array. The magnitude of the estimated variance components suggested potential for breeding for BKD resistance in charr, with relevant heritabilities ranging from 0.05 to 0.56 depending on the resistance proxy used. In addition, GWAS suggested that BKD resistance is a polygenic trait. Furthermore, genomic prediction approaches indicated that BKD-resistant animals can be identified using their SNP genotypes. Accuracies, expressed as Pearson correlation coefficients, when BKD resistance was analysed as a continuous trait, ranged from 0.42 to 0.52. In the scenario where BKD resistance was treated as a binary trait, the efficiency of genomic prediction was assessed using ROC curves, with an area under the curve of 0.72. Finally, no unfavourable correlations were found with growth traits. The developed 72k SNP array has the potential of being a pivotal tool for the Swedish Arctic charr breeding program. Moreover, our data support the use of genomic prediction in breeding BKD-resistant Arctic charr. As a critical next step, further validations in actual industry conditions would be required.

Viral haemorrhagic septicaemia (VHS) is a severe disease affecting rainbow trout (Oncorhynchus mykiss) and a wide range of wild freshwater and marine fish species. VHSV threatens rainbow trout aquaculture, as it may cause 100% mortality in fry. Previous studies identified a quantitative trait locus (QTL) on chromosome 3 associated with resistance to VHSV waterborne challenge and reduced viral replication in fin explants, although these findings were obtained using limited genetic diversity. The objective of this study was to validate and extend the identification of genomic regions associated with resistance to VHSV in the genetically diverse rainbow trout line designated "synthetic." A genome-wide association study (GWAS) was conducted using whole-genome sequences from parents of progeny classified as resistant or susceptible to a VHSV waterborne challenge. While the QTL on chromosome 3 was not validated in the synthetic line, four novel suggestive SNPs associated with survival following VHSV waterborne challenge were identified on chromosomes 6, 8, 17, and 32. Notably, one SNP on chromosome 17 was located within a gene potentially involved in antiviral defence, a paralog of lrp1 (low-density lipoprotein receptor-related protein 1). To further investigate its role, lrp1 function was analysed in vitro using CRISPR-Cas9 genome editing. Three independent lrp1-/- CHSE-EC cell lines were generated and challenged with VHSV. The results showed that lrp1 is not essential for viral entry but may modulate the inflammatory response during VHSV infection in epithelial cell lines.

Pseudomonas aeruginosa is a clinically significant bacterial pathogen that poses a serious threat to aquaculture. However, there are limited information on Massilia isolates against pathogenic P. aeruginosa in aquaculture. In the present study, a facultative predator, M. varians isolate P2-4, was isolated from aquaculture sediment using Chinese mitten crab Eriocheir sinensis-pathogenic P. aeruginosa as the prey bacterium, and its genomic feature, bacteriolysis-related genes, safety, bacteriolytic spectrum, and in vitro and in vivo antibacterial effects against pathogenic P. aeruginosa in E. sinensis were further characterized. Isolate P2-4 consisted of one chromosome and one plasmid (with a total of 75 tRNAs, 7 5S rRNAs, 7 16S rRNAs, 7 23S rRNAs, 34 sRNAs, 5,238 coding genes, 20 genomic islands, 1 prophage, 23 insertion sequences, and 102 repeat sequences), and harbored 19 bacteriolysis-related genes (pilA, pilB, pilC, pilD, pilF, pilG, pilH, pilM, pilO, pilP, pilQ, pilS, pilR, pilT, mltA, mltB, mltC, mltD, and dacB) associated with cellular motility and cell wall lysis. In addition, the isolate carried no virulence genes, was unable to produce haemolysin, hydrogen sulfide, nitrite and ammonia, and avirulent in E. sinensis with a 7-day acute intraperitoneal LD50 value of above 5.0 x 108 CFU/mL. Furthermore, the isolate possessed a wide bacteriolytic spectrum against pathogenic Shewanella algae, Aeromonas caviae, A. hydrophila, and Photobacterium damselae besides P. aeruginosa, exhibited bacteriolysis rates of 99.35% to 99.99% towards the pathogenic P. aeruginosa at 1.0x103 to 1.0x10{square} CFU/mL, and displayed relative percentage survivals of 42.31% to 73.08% against P. aeruginosa infection in E. sinensis at doses of 6.0 x 103 to 6.0 x 105 CFU/g diet. To our knowledge, this study for the first time demonstrates a M. varians strain as a potential biocontrol agent against pathogenic P. aeruginosa in aquaculture.

The slipper lobster (Thenus australiensis) is rapidly emerging as a high-potential species for commercial aquaculture. Because females exhibit superior growth characteristics due to less frequent moulting after sexual maturity, developing monosex breeding strategies is highly desirable for industry profitability. However, the lack of genomic resources and early sex-identification tools has hindered this development. Here, we report the first draft male genome assembly for T. australiensis, generated using a combination of whole-genome shotgun sequencing, DArT-seq, and multi-tissue transcriptomics. The curated assembly spans 0.913 Gbp with high functional completeness (93.0% BUSCO), providing a robust repertoire of 30,100 protein-coding genes. Through k-mer subtraction and population-level DArT-seq genotyping, we provide definitive evidence that T. australiensis utilizes an XX/XY sex-determination system. Crucially, by identifying male-specific structural variations within a neo-Y locus, we developed a diagnostic PCR assay targeting a male-exclusive sequence. This 171 bp marker achieved 100% accuracy in phenotypic sex identification across wild-caught populations. Ultimately, these foundational genomic resources, combined with a highly reliable molecular sexing tool, provide the critical framework necessary for early sex sorting, broodstock management, and the commercial advancement of monosex slipper lobster farming.

Currently, global conservation efforts for wildlife focus on a limited number of cell types and species. Although protocols from domestic and non-threatened related species have been applied to endangered species, cryopreservation techniques are species-specific and are constrained by a lack of understanding of reproductive biology in these species. Based on a review of 126 original studies from 27 countries, encompassing 160 species, we assess the current state of cryopreservation in wildlife, including gametes, embryos, somatic cells, and various tissues. Furthermore, we focused on the most homogeneous and frequently studied cell type in wildlife cryobanking: mammals sperm.. A meta-analysis of 27 studies was conducted to examine species-specific and protocol-dependent factors that affect post-thaw sperm quality. Our findings provide quantitative estimates of cryopreservation for various cell types and tissues in wildlife taxa. Furthermore, they serve as a crucial research roadmap, identifying major challenges in cryopreservation and proposing solutions.

Japanese chum salmon (Oncorhynchus keta) populations supported by one of the worlds largest hatchery programs have experienced severe declines in marine survival and abundance. To evaluate long-term changes in reproductive traits, we analyzed temporal and geographic variation in female fork length (FL) and egg size from 13 hatchery-enhanced rivers in Japan during 1999-2019. Females from northern rivers tended to have smaller FL but produced larger eggs, indicating persistent latitudinal differentiation in reproductive traits. Despite this geographic structure, both FL and egg size declined synchronously among rivers over time. Using nationwide annual means, exponential decay analyses showed that egg size declined by approximately 2% per generation, whereas marine survival and population abundance declined by approximately 22% per generation. Mean egg size was positively correlated with subsequent marine survival, explaining 45% of the variation in return rates. Mean female FL explained 64% of the variation in mean egg size. Exponential decay rates estimated from standardized variables were nearly identical among egg size, marine survival, and chum salmon abundance, suggesting common underlying drivers of long-term decline.

Charrs (Salvelinus spp.) are strong candidates for cyclical feed-restriction strategies due to their documented exceptional ability to withstand and recover from feed resources scarcity during winter (Drouin-Johnson et al. 2026; Le Francois et al. 2023; Savoie et al. 2017). Dietary nucleotides (NT) are supplements used to improve growth and health in fish, yet their integration into feed-restriction programs to enhance growth recovery remains largely unexplored. The one-carbon (1C) metabolism offers a mechanistic framework to investigate these effects, as it relies on vitamin B status and supports methylation reactions central to biosynthesis and growth. A large-scale trial involving S. alpinus x S. fontinalis was conducted, comprising a 90-day fasting period followed by refeeding on a commercial feed with or without NT. Growth and feeding metrics were evaluated alongside key metabolites of 1C metabolism to assess adaptations to fasting-induced vitamin B deficiency and the role of NT in recovery. Starved fish lost 5.2 % of body mass (p < 0.05) but displayed accelerated growth upon refeeding, further enhanced by NT supplementation by 27.8 % (p < 0.05), forecasting full compensation after 125 days. Starvation increased homocysteine and decreased methionine concentrations (p < 0.05), while S-adenosylmethionine, S-adenosylhomocysteine, and formate showed only minor fluctuations. These patterns indicate moderate but reversible fluctuations of 1C metabolism. NT supplementation may exert a methyl-unit sparing effect during refeeding, potentially supporting anabolic recovery. This study uniquely combines commercial-scale growth trials and molecular analyses, providing new insights into a species-adapted feeding strategy that enhances growth recovery performances and metabolic resilience.

Neoparameoba perurans causes Amoebic Gill Disease (AGD), a major parasitic disease of marine-phase Atlantic salmon and rainbow trout worldwide. Treatment options are limited to freshwater baths, which are costly at scale and exhibit only limited long-term efficacy. N. perurans contains an obligate eukaryotic symbiont, Perkinsela-like organism (PLO). PLO belongs to the class Kinetoplastida, which includes medically and veterinary important parasites such as Trypanosoma and Leishmania. As such, we hypothesised that trypanocidal drugs developed against other kinetoplastids might also affect N. perurans, potentially through disruption of its PLO symbiont, and used this hypothesis as a rationale for prioritising a focused panel of candidate compounds for screening. A holographic motility-based cytotoxicity assay was established to identify promising candidates in vitro, followed by controlled host tolerance testing and finally a field efficacy sea trial using naturally AGD-exposed site in the west of Ireland. Several compounds showed activity in vitro, especially miltefosine (EC50 1.84 uM, amoebicidal) and isometamidum (EC50 4.63 uM, amoebostatic). In vivo (two intramuscular injections, two weeks apart), miltefosine (Odds Ratio (OR) 0.62), isometamidum (OR 0.61) and benznidazole (OR 0.64) significantly improved gill score over four weeks, with miltefosine showing the largest effect size. Gill parasitaemia, measured via qPCR, was not reduced. Instead, two compounds increased apparent amoeba loads. This work support trypanocidal as potential AGD treatments in the field, although optimisation of dosing, delivery and mode of action requires further study.

Understanding the genetic structure of keystone species within river networks is essential for effective conservation and management. While population differentiation of anadromous species often occurs between river systems, less research has been conducted on differentiation within rivers with smaller catchment areas. In this study, we investigated the population genetic structure of wild Atlantic salmon (Salmo salar) across the small-scale river Ronne [a] system in southernmost Sweden using Restriction-site Associated DNA sequencing (RADseq). Although the Admixture analysis did not detect clearly defined genetic clusters, significant pairwise FST values and DAPC revealed emerging population differentiation among the Ronne [a] tributaries. The observed patterns are consistent with a system characterized by connectivity, where genetic flow is present but can be reduced by behavioral and ecological factors such as spawning homing behavior and selective movements. These findings suggest that, despite overall connectivity, Atlantic salmon populations in the Ronne [a] catchment area may function as partially independent sub-populations. This highlights the importance of conservation and management strategies in fragmented river systems to consider population genetic structure to support resilient salmon populations under ongoing anthropogenic pressures.

Congenital disorders of glycosylation and deglycosylation are rare, serious, and lethal disorders afflicting humans. CDGs and CDDGs result in loss of function enzymes which fail to build or break down oligosaccharides on proteins. This can produce protein aggregates and, in turn, reactive oxygen species that harm the cell eventually leading to autophagy and apoptosis. Because sperm contain high concentrations of polyunsaturated fatty acids, they are especially sensitive to these effects, which is understood as one of the leading factors in human male infertility. Sperm are developed in zebrafish similarly to humans and are useful models to examine human reproductive health, as well as genetic disorders. The combination of these advantages makes the analysis of sperm from zebrafish with heterozygous ALG1 or DPAGT1 CDGs or the NGLY1 CDDG suitable. Analysis of sperm concentration, motility, status, viability, and hypoosmotic swelling demonstrated the effects of these disorders on sperm quality. Results showed a significant decrease in sperm concentration, motility, and hypoosmotic swelling for all mutant zebrafish compared to the wild type. This suggests that CDGs and CDDGs influence the amount of sperm produced, the percentage of sperm cells that are mobile, and the integrity of the plasma membrane.

The age at which Atlantic salmon reaches sexual maturity shows a strong hereditary component associated with the vgll3a locus. The role of Vgll3 in maturation has remained unknown in vertebrates until recently, when it has been linked to pleiotropic roles in killifish, both delaying male maturation and affecting lifespan by protecting against cancer. As Atlantic salmon has two vgll3 paralogs, where only vgll3a has been associated with sexual maturation, it may provide a suitable model for studying the maturation-specific function of vgll3, as the other paralog may buffer for pleiotropic roles of vgll3. To address this, we used CRISPR/Cas9 to generate fish highly mutated in the vgll3a gene. We monitored their maturation and crossed highly mutated crispants to generate two year-classes of complete loss-of-function. All groups were reared under environmental conditions triggering early maturation in one-year-old males. We found a clear difference in the proportion of sexually maturing or mature fish between the different genotypes: in all experiments significantly fewer vgll3a-/- males entered puberty and reached final maturation compared to vgll3a+/- and vgll3a+/+ males. Furthermore, loss of vgll3a resulted in lower frequencies of maturation also in females. We conclude that Vgll3a stimulates maturation and that its complete removal significantly reduced maturation rates in both sexes in Atlantic salmon. Our findings also identify vgll3a as the causative gene in the locus associated with age at sexual maturity. Together, our findings support a new role for Vgll3 in initiating puberty in vertebrates and identifying salmon as a promising model for functional studies regarding the timing of sexual maturation.

The growing demand for sustainable aquafeeds has intensified interest in alternative protein ingredients capable of reducing reliance on fishmeal without compromising fish performance. Here, we evaluated microbial community-based single-cell protein (SCP) as a fishmeal substitute in juvenile Asian seabass (Lates calcarifer) diets in two independent feeding trials of juvenile fish conducted over 49 and 56 days, respectively and compared them to a previous study that lasted 24 days. SCP was produced from nutrient-rich soybean-processing side streams by microbial communities in fermenters and incorporated into experimental diets at inclusion levels ranging from 10% to 100% fishmeal replacement. In the 24-day trial, a diet containing 50% fishmeal replacement with lab-scale produced SCP achieved 100% survival and a feed conversion ratio (FCR), specific growth rate (SGR), and weight gain comparable to the fishmeal control diet. In the 49-day trial using pilot-scale produced SCP, a 50% fishmeal replacement also maintained an FCR and feed intake comparable to the control, whereas complete replacement reduced feed intake and growth performance. In a 56-day pilot-scale trial that used 500-L fish tanks, diets containing up to 50% fishmeal replacement maintained comparable survival, weight gain, and SGR, although moderately higher FCR values were observed at higher SCP inclusion levels. Proximate composition and essential amino acid profiles of fish fed control or SCP-containing diets were comparable. Genome-resolved metagenomic analyses revealed diverse microbial taxa associated with the SCP. Collectively, these findings support microbial community-based SCP as a scalable and reproducible alternative protein platform for aquaculture feeds across independent trials and production scales.

The sugar kelp Saccharina latissima is a promising candidate for sustainable aquaculture in the North Atlantic and North-East Pacific but genetic improvement has been hindered by limited understanding of the genetic basis of economically important traits. We conducted the first genome-wide association study (GWAS) for this species using 202 self-fertilised pseudo-F1 individuals derived from 12 populations spanning northern and southern European genetic clusters. Individuals were genotyped with ddRAD-seq-derived SNP markers and phenotyped in a common garden experiment for four morphological traits (blade length, blade width, blade area, stipe length) and six metabolic traits related to nitrogen metabolism. We identified 26 significant marker-trait associations, with phenotypic variance explained (PVE) ranging from 0.65% to 52.44%. Major-effect loci were detected for blade width (52.44% PVE) and blade area (45.22% PVE) and a locus on chromosome 17 influenced both blade length and blade area. Marker-based heritability estimates ranged from 0.75 to 0.99 for morphological traits and from 0.00 to 0.99 for metabolic traits, though with large standard errors. Cross-validation of genomic selection models yielded predictive abilities of 0.21-0.59 across traits. Our findings reveal a mixed genetic architecture with major-effect loci suitable for marker-assisted selection and polygenic traits amenable to genomic selection, providing a foundation for genomics-assisted breeding programs in kelp aquaculture.

BackgroundSkeletal muscle growth is a key determinant of body size and market value in salmonid aquaculture, yet the mechanisms linking genomic variation to muscle fiber hypertrophy remain poorly resolved. Myofiber cross-sectional area (CSA) provides a quantitative cellular proxy for fiber size and a direct link to macroscopic growth traits. MethodsWe performed histological phenotyping of white skeletal muscle from rainbow trout (Oncorhynchus mykiss) representing divergent fillet-yield selection lines (ARS-FY-H and ARS-FY-L), quantifying mean myofiber CSA and fiber number using high-throughput image analysis. Genome-wide association analysis (GWAS) was conducted using low-pass whole-genome sequencing ([~]1x) with genotype imputation and functional variant annotation. RNA sequencing was performed on fish representing high and low CSA extremes to identify differentially expressed genes and enriched biological pathways. ResultsMean myofiber CSA was significantly associated with body weight, muscle weight, visceral weight, and body length (p < 0.05), while fiber count showed no significant association with most growth traits, implicating hypertrophy as the primary driver of muscle mass variation. GWAS identified a significant QTL spanning [~]4.76 Mb on chromosome 2 (117 significant SNPs; Bonferroni-adjusted P [&le;] 0.05; {lambda} = 1.02). Associated variants were predominantly noncoding, enriched in intronic, intergenic, and enhancer-annotated regions. A high density of SNPs colocalized with the TGF-{beta}2 locus, overlapping strong and genic enhancer elements in white muscle. Transcriptomic comparisons revealed that high-CSA muscle showed elevated expression of genes related to contractile function, cytoskeletal organization, and translation, while low-CSA muscle exhibited upregulation of extracellular matrix and immune-related genes consistent with a tissue remodeling state. ConclusionsNoncoding regulatory variation within a significant QTL spanning the TGF-{beta}2 locus is associated with distinct transcriptional programs linked to muscle fiber hypertrophy in rainbow trout. By integrating genetic variation, chromatin-state annotation, and transcriptomic profiling, this study identifies candidate regulatory loci associated with variation in muscle cellularity and growth-related phenotypes in rainbow trout.

The aim of this study was to evaluate the impact of two sustainable dietary protein sources on the structure and composition of the gut microbiota in European sea bass (Dicentrarchus labrax) juveniles. These protein sources were incorporated to the aquafeeds containing (a) Lupinus albus meal, treated with either exogenous enzymes (Solid state hydrolysis-SSH) or fermented with Saccharomyces cerevisiae (Solid state fermentation, SSF) and (b) Lagocephalus sceleratus meal. In the first case (a), the control aquafeed simulated a standard commercial diet, containing soybean meal whereas in the rest of the diets soybean meal was partially or totally replaced by hydrolysed or fermented Lupin meal. In the second case (b) the fish were fed Lagocephalus sceleratus unprocessed fishmeal as well as treated at different temperatures to deactivate tetrodotoxin (TTX). A control diet with 30% commercial fish meal was also fed as a reference diet. Both diets in all inclusion levels did not cause any significant gut microbiota change, suggesting their neutral role in this aspect. However, the gut bacterial communities of the fish fed with 12.5% lupin meal inclusion, had increased amino acid biosynthetic pathways suggesting a beneficial effect.

Telomere length (TL) is increasingly used in ecology as a biomarker of individual quality and environmental stress, yet research on non-model species with complex life histories remains limited. Because TL varies among tissues and across ages in a species-specific manner, identifying non-lethal tissues that reliably reflect whole-organism telomere dynamics is essential for longitudinal telomere studies in the field. This study aimed to evaluate tissue-specific TL in Japanese eel (Anguilla japonica), an endangered catadromous fish. We first mapped the chromosomal distribution of telomeric sequences using fluorescent in situ hybridization (FISH), the first application of this method in this species. We then tested whether muscle and caudal fin, which can be sampled easily and non-lethally, can serve as suitable proxy tissues for TL measurements in wild individuals. Relative telomere length (RTL) was quantified by qPCR in blood, brain, caudal fin, gonads, heart, liver, and muscle. FISH analysis confirmed telomeric repeats at all chromosomal ends, with only weak interstitial signals on three chromosomal pairs unlikely to affect qPCR-based estimates. A generalized additive mixed model and Wilcoxons signed-rank tests revealed significant inter-tissue differences: RTL was shortest in the brain and muscle and longest in liver, blood and caudal fin. Muscle and caudal fin RTL were significantly correlated with RTL in many other tissues, supporting their use as proxy tissues for longitudinal TL monitoring, including responses to environmental variation. Both total length and age were tested as explanatory variables for RTL, and the model including total length showed a better fit than the age-based model. Non-linear relationships between RTL and total length observed in several tissues suggest physiological shifts associated with growth and sexual differentiation. Overall, these findings advance understanding of telomere dynamics in eels and establish muscle and caudal fin as suitable tissues for repeated, non-lethal TL assessment in ecological and conservation contexts.

Understanding population structure is critical for effective fisheries management in species with complex life histories and variable recruitment. Southern flounder (Paralichthys lethostigma) is a valuable flatfish species with declining populations in the Southeast United States. Improved management may depend on a better understanding of fine-scale and temporal population genetic structure in this region; however, such structure remains poorly characterized. To address our lack of understanding of the spatial and temporal population structure of this important species, we used double digest reduced-representation genome sequencing (ddRADSeq) on juveniles from estuaries in North Carolina and Texas between 2014 and 2023. We found significant genetic differentiation between the Gulf of Mexico and Atlantic populations, supporting the management of these regions as distinct stocks. By contrast, we detected significant variance in genetic structure within Texas and North Carolina populations that was not consistent across sampling years between estuaries in close proximity. The population genetic structure of southern flounder suggests significant, temporally variable genetic differences within estuarine locations that may result from variation in larval dispersal and recruitment patterns. Our findings highlight the value of integrating fine-scale, multi-year genetic data to capture temporal dynamics and avoid misleading conclusions based on single-year or broad-scale sampling.

Global climate change has increased the frequency and severity of stressful temperatures that freshwater fishes experience, necessitating rapid and sensitive methods to monitor wild populations. Tissues used to measure transcriptional responses traditionally involved invasive or lethal sampling, which may be undesirable for imperilled species. Epidermal mucus offers a non-lethal and minimally invasive alternative, but whether thermal thresholds can be detected in mucus to identify fish experiencing thermal stress is unclear. Bull trout (Salvelinus confluentus) are a legally protected salmonid and cold-water specialist, generally occupying waters 12 {degrees}C and below, with higher temperatures resulting in cellular stress. Therefore, we measured a suite of 56 genes using high-throughput qPCR to compare machine learning classifiers developed with transcriptional profiles of epidermal mucus, gill, liver, and muscle to classify laboratory reared juvenile bull trout as below (9 {degrees}C, 12 {degrees}C) or above (15 {degrees}C, 18 {degrees}C) cellular thermal thresholds. Mucus profiles most resembled gills but represented an intermediate transcriptional response to all tissues. A reduced biomarker panel of 10 genes in mucus assigned fish to stress categories with 94.1% (95% CI = 71.3-99.9%) accuracy, which was comparable to gill (100.0%, CI = 82.4- 100%), liver (95.0%, CI = 75.1-99.9%), and muscle (100.0%, CI = 80.5-100.0%). Sex-specific temperature effects were evident in all tissues, but less pronounced in mucus and gill than in liver and muscle. Our findings demonstrate that transcriptional profiling of mucus can reliably identify individuals experiencing thermal stress, highlighting the promise of this non-lethal approach for monitoring at-risk species.

In the multitude of suitable experimental systems used for functional studies in the field of developmental biology, the sea urchin plays a central role due to its amenability to various methods, including both transient and stable transgenesis. Among others, transposable elements represent powerful tools for generating stable transgenic specimens, and Minos transposon turned out to be an excellent genetic tool in marine organisms, despite its efficiency being host-dependent. This study provides new evidence for the activity of Minos transposable elements and their stable integration into the genome of the Mediterranean sea urchin Paracentrotus lividus. Using the Minos-based technology coupled with a fully-automated system used for the qPCR screening of the Minos transposon integration, we devised a new pipeline for performing transgenesis-based functional studies in P. lividus.

The moonfish Mene maculata (Bloch and Schneider, 1801), the sole extant species of the family Menidae, is widely distributed in the Indo-West Pacific. Although its spawning season has previously been inferred from seasonal variation in the gonadosomatic index (GSI), the histological basis for interpreting GSI as an indicator of spawning-capable status remains limited. Here we describe the gonadal structure of male and female M. maculata and evaluate how GSI and standard length relate to spawning-capable status based on germ cell development. The testis was lobular and exhibited an unrestricted spermatogonial distribution, a structure widely observed among neoteleosts, indicating that the phylogenetic distinctiveness of M. maculata was not associated with a distinctive testicular structure. In the spawning-capable female, oocytes at multiple developmental stages, from primary growth to oocyte maturation, co-occurred within the ovary, indicating asynchronous ovarian development. This finding suggests that M. maculata may be a batch spawner rather than a total spawner as previously inferred. The spawning-capable female had a GSI consistent with previously inferred spawning season estimates. By contrast, histologically examined males were spawning capable at GSI values lower than those previously associated with the inferred spawning season, suggesting that male spawning-capable status may persist beyond the period inferred from elevated GSI alone. This study provides the first histological description of reproductive biology in the phylogenetically distinctive M. maculata and establishes a histological basis for interpreting GSI as a reproductive indicator.