Genome sequencing and analysis of black flounder (Paralichthys orbignyanus) reveals new insights into Pleuronectiformes genomic size and structure.

Black flounder (Paralichthys orbignyanus, Pleuronectiformes) is a commercially significant marine fish with promising aquaculture potential in Argentina. Despite extensive studies on Black flounder aquaculture, its limited genetic information available hampers the crucial role genetics plays in the development of this activity. In this study, we first employed Illumina sequencing technology to sequence the entire genome of Black flounder. Utilizing two independent libraries-one from a female and another from a male-with 150 bp paired-end reads, a mean insert length of 350 bp, and over 35 X-fold coverage, we achieved assemblies resulting in a genome size of ~ 538 Mbp. Analysis of the assemblies revealed that more than 98% of the core genes were present, with more than 78% of them having more than 50% coverage. This indicates a somehow complete and accurate genome at the coding sequence level. This genome contains 25,231 protein-coding genes, 445 tRNAs, 3 rRNAs, and more than 1,500 non-coding RNAs of other types. Black flounder, along with pufferfishes, seahorses, pipefishes, and anabantid fish, displays a smaller genome compared to most other teleost groups. In vertebrates, the number of transposable elements (TEs) is often correlated with genome size. However, it remains unclear whether the sizes of introns and exons also play a role in determining genome size. Hence, to elucidate the potential factors contributing to this reduced genome size, we conducted a comparative genomic analysis between Black flounder and other teleost orders to determine if the small genomic size could be explained by repetitive elements or gene features, including the whole genome genes and introns sizes. We show that the smaller genome size of flounders can be attributed to several factors, including changes in the number of repetitive elements, and decreased gene size, particularly due to lower amount of very large and small introns. Thus, these components appear to be involved in the genome reduction in Black flounder. Despite these insights, the full implications and potential benefits of genome reduction in Black flounder for reproduction and aquaculture remain incompletely understood, necessitating further research.

Identification and functional characterization of annexin A2 in half-smooth tongue sole (Cynoglossus semilaevis).

Annexin A2 (AnxA2), belonging to the annexin family, plays a crucial role in immune responses. In this study, the cDNA of the AnxA2 gene was identified in half-smooth tongue sole, Cynoglossus semilaevis. The transcript of AnxA2 gene in C. semilaevis (CsAnxA2) showed broad tissue distribution, with the highest expression level observed in the gut. CsAnxA2 expression was significantly up-regulated in the intestine, spleen, and kidney tissues following exposure to Shewanella algae. Immunohistochemical staining revealed that CsAnxA2 was predominantly expressed in epithelial cells and significantly elevated after S. algae challenge. Subcellular localization showed that CsAnxA2 was primarily localized in the cytoplasmic compartment. Moreover, proinflammatory cytokines (IL-6, IL-8 and IL-1ß) exhibited significant upregulation after CsAnxA2 was overexpressed in vivo. One hundred and fifty-eight CsAnxA2-interacting proteins were captured in the intestinal tissue, showing the top two normalized abundance observed for actin beta (ACTB) and protein S100-A10 (p11). Fifty-four high abundance CsAnxA2-interacting proteins (HIPs) were primary enriched in ten pathways, with the top three significantly enriched pathways being Salmonella infection, glycolysis/gluconeogenesis, and peroxisome proliferator-activated receptor (PPAR) signaling pathway. These results provide valuable information for further investigation into the functional mechanism of AnxA2 in C. semilaevis.

Oral Administration of Antimicrobial Peptide NZ2114 Through the Microalgal Bait Tetraselmis subcordiformis (Wille) Butcher for Improving the Immunity and Gut Health in Turbot (Scophthalmus maximus L.).

Antibiotics are widely used in aquaculture to treat the bacterial diseases. However, the improper use of antibiotics could lead to environmental pollution and development of resistance. As a safe and eco-friendly alternative, antimicrobial peptides (AMPs) are commonly explored as therapeutic agents. In this study, a mutant strain of Tetraselmis subcordiformis containing AMP NZ2114 was developed and used as an oral drug delivery system to reduce the use of antibiotics in turbot (Scophthalmus maximus) aquaculture. The gut, kidney, and liver immune-related genes and their effects on gut digestion and bacterial communities in turbot fed with NZ2114 were evaluated in an 11-day feeding experiment. The results showed that compared with the group fed with wild-type T. subcordiformis, the group fed with T. subcordiformis transformants containing NZ2114 was revealed with decreased levels of both pro-inflammatory factors (TNF-α and IL-1ß), inhibitory effect on Staphylococcus aureus, Vibrio parahaemolyticus, and Vibrio splendidus demonstrated by the in vitro simulation experiments, and increased richness and diversity of the gut microbiota of turbot. In conclusion, our study provided a novel, beneficial, and low-cost method for controlling bacteria in turbot culture through the oral drug delivery systems.

Unraveling the transcriptomic landscape of eye migration and visual adaptations during flatfish metamorphosis.

Flatfish undergo a remarkable metamorphosis from symmetrical pelagic larvae to fully asymmetrical benthic juveniles. The most distinctive features of this transformation is the migration of one eye. The molecular role of thyroid hormone in the metamorphosis process in flatfishes is well established. However, the regulatory network that facilitates eye movement remains enigmatic. This paper presents a morphological investigation of the metamorphic process in turbot eyes, using advanced imaging techniques and a global view of gene expression. The study covers migrant and non-migrant eyes and aims to identify the genes that are active during ocular migration. Our transcriptomic analysis shows a significant up-regulation of immune-related genes. The analysis of eye-specific genes reveals distinct patterns during the metamorphic process. Myosin is highlighted in the non-migrant eye, while ependymin is highlighted in the migrant eye, possibly involved in optic nerve regeneration. Furthermore, a potential association between the alx3 gene and cranial restructuring has been identified. Additionally, it confirmed simultaneous adaptation to low light in both eyes, as described by changes in opsins expression during the metamorphic process. The study also revealed that ocular migration activates systems asynchronously in both eyes, providing insight into multifaceted reorganization processes during metamorphosis of flatfish.

Dietary methanotroph bacteria meal alleviates soybean meal-induced enteritis by improving immune tolerance and intestinal flora profile of juvenile turbot (Scophthalmus maximus L.).

An 8-week growth trial was performed to investigate the protective effects of methanotroph bacteria meal (MBM) produced from methane against soybean meal-induced enteritis (SBMIE) in juvenile turbot (Scophthalmus maximus L.). Five isonitrogenous and isolipidic diets were formulated: fishmeal-based diet (FM, the control group); FM with approximate 50% of fishmeal substituted by 399.4 g/kg soybean meal (SBM); SBM supplemented with 63.6, 127.2 and 190.8 g/kg MBM (named MBM1, MBM2 and MBM3), each diet was randomly assigned to triplicate fibreglass tanks. Results showed that fish fed with SBM exhibited enteritis, identified by reduced relative weight of intestine (RWI), as well as expanded lamina propria width and up-regulated gene expression of pro-inflammatory cytokines (tnf-α, il-6 and il-8) in intestine. While the above symptoms were reversed when diet SBM supplemented with MBM at the levels of 63.6 and 127.2 g/kg, as well as characterized by up-regulated gene expression of anti-inflammatory cytokines (tgf-ß and il-10) and tight junction protein (claudin3, claudin4 and claudin7) in intestine. Intestinal transcriptome analysis showed that the differentially expressed genes between groups FM and SBM predominantly enriched in the JAK-STAT signaling pathway, and the enrichment of differentially expressed genes between groups SBM and SBM supplemented with 63.6 g/kg MBM was in the inflammatory bowel disease (IBD) and JAK-STAT signaling pathway. To be specific, the expression of jak1, jak2b, stat1 and stat5a was significantly up-regulated when fish fed with SBM, suggested the activation of JAK-STAT signaling pathway, while the expression of these above genes was depressed by providing MBM to diet SBM, and the gene expression of toll-like receptors tlr2 and tlr5b showed a similar pattern. Moreover, intestinal flora analysis showed that community richness and abundance of beneficial bacteria (Cetobacterium and acillus_coagulans) were improved when fish fed with SBM supplemented with 63.6 g/kg MBM. Overall, methanotroph bacteria meal may alleviate SBMIE by regulating the expression of tight junction protein, toll-like receptors and JAK-STAT signaling pathway, as well as improving intestinal flora profile, which would be beneficial for enhancing the immune tolerance and utilization efficiency of turbot to dietary soybean meal.

First case of a reversed Parabothus taiwanensis Amaoka & Shen, 1993 from Taiwan (Pleuronectiformes: Bothidae), with first evidence of situs inversus viscerum in bothid.

Background: Reversed condition is rarely found in most flatfishes in natural environment, except for some certain species. The mechanism controlling the reversals in flatfishes has been studied in some cultivated species, whereas some have only few cases for the entire family and remain unclear. Here, we report the first record of a dextral (reversed) specimen of Parabothus taiwanensis Amaoka & Shen, 1993 collected off southwestern Taiwan recently. It represents the second reversed case ever recorded in Bothidae. We aim to provide a detailed description of this dextral specimen and compared to the sinistral (normal) specimens collected from the vicinity. Methods: Specimens were fixed in 4% formaldehyde and transferred to 70% ethanol for preservation. Meristic and morphometric characters were examined for both dextral and sinistral specimens. Dissections were made on specimens to confirm the position of internal organs. Lastly, X-radiographs were taken to elucidate the osteological features. Results: As a result, no differences of both meristic and morphometric characters were observed between the dextral and sinistral specimens. Nevertheless, situs inversus viscerum is discovered in the dextral specimen for the first time in Bothidae and the sixth record within flatfishes.

Optimal dietary lipid levels alleviated adverse effects of high temperature on growth, lipid metabolism, antioxidant and immune responses in juvenile turbot (Scophthalmus maximus L.).

Fish physiological health is often negatively impacted by high-temperature environments and there are few studies on how dietary lipids affect fish growth and physiology when exposed to heat stress. The main objective of this research was to examine the impact of dietary lipid levels on growth and physiological status of juvenile turbot (Scophthalmus maximus L.) and determine if dietary lipid concentration could alleviate the possible adverse effects of heat stress. Five diets containing 6.81%, 9.35%, 12.03%, 14.74%, and 17.08% lipid, respectively, were formulated and fed to turbot (initial weight 5.13 ± 0.02 g) under high-temperature conditions (24.0-25.0 °C). Meanwhile, the diet with 12.03% lipid (considered by prior work to be an optimal dietary lipid level) was fed to turbot of the same size at normal temperature. Results suggested that, among the different dietary lipid levels under high-temperature conditions, fish fed the optimal lipid (12.03%) exhibited better growth compared to non-optimal lipid groups, as evidenced by higher weight gain and specific growth rate. Simultaneously, the optimal lipid diet may better maintain lipid homeostasis, as attested by lower liver and serum lipid, along with higher liver mRNA levels of lipolysis-related genes (pgc1α, lipin1, pparα, lpl and hl) and lower levels of synthesis-related genes (lxr, fas, scd1, pparγ, dgat1 and dgat2). Also, the optimal lipid diet might mitigate oxidative damage by improving antioxidant enzyme activity, decreasing malondialdehyde levels, and up-regulating oxidation-related genes (sod1, sod2, cat, gpx and ho-1). Furthermore, the optimal lipid may enhance fish immunity, as suggested by the decrease in serum glutamic-oxalacetic/pyruvic transaminase activities, down-regulation of pro-inflammatory genes and up-regulation of anti-inflammation genes. Correspondingly, the optimal lipid level suppressed MAPK signaling pathway via decreased phosphorylation levels of p38, JNK and ERK proteins in liver. In summary, the optimal dietary lipid level facilitated better growth and physiological status in turbot under thermal stress.

Effect of dietary l-glutamine supplementation on the intestinal physiology and growth during Solea senegalensis larval development.

The maturation of the intestinal digestive and absorptive functions might limit the amount of absorbed nutrients to fulfil the high requirements of the fast-growing marine fish larva. Glutamine (Gln) has been described to improve intestinal epithelium functions, due to its involvement in energy metabolism and protein synthesis. The purpose of this study was to evaluate dietary 0.2% Gln supplementation on aspects of intestinal physiology, protein metabolism and growth-related genes expression in Senegalese sole larvae. Experiment was carried out between 12 and 33 days post hatching (DPH) and fish were divided into two experimental groups, one fed Artemia spp. (CTRL) and the other fed Artemia spp. supplemented with Gln (GLN). GLN diet had two times more Gln than the CTRL diet. Samples were collected at 15, 19, 26 and 33 DPH for biometry, histology, and digestive enzymes activity, and at 33 DPH for gene expression, protein metabolism and AA content determination. Growth was significantly higher for Senegalese sole fed GLN diet, supported by differences on protein metabolism and growth-related gene expression. Slight differences were observed between treatments regarding the intestinal physiology. Overall, GLN diet seems to be directed to enhance protein metabolism leading to higher larval growth.

Identification of mapk genes, and their expression profiles in response to low salinity stress, in cobia (Rachycentron canadum).

Mitogen-activated protein kinases (MAPKs) are a class of protein kinases that regulate various physiological processes, and play a crucial role in maintaining the osmotic equilibrium of fish. The objective of this study was to identify and characterize the mapk family genes in cobia (Rachycentron canadum) and examine their expression profiles under different low salinity stress regimes (acute: from 30‰ to 10‰ in 1 h, sub-chronic: from 30‰ to 10‰ over 4 d). A total of 12 cobia mapk genes (Rcmapks) were identified and cloned, including six erk subfamily genes (Rcmapk1/3/4/6/7/15), three jnk subfamily genes (Rcmapk8/9/10) and three p38 mapk subfamily genes (Rcmapk 11/13/14). Domain analysis indicated that the RcMAPKs possessed the typical domains including S_TKc and PKc_like domain. Phylogenetic analysis revealed that the Rcmapks were most closely related to those of the turbot (Scophthalmus maximus). The tissue distribution of mapk genes in adult cobia and the expression patterns of Rcmapks under different low salinity stress regimes were investigated using quantitative real-time PCR (qRT-PCR). The results revealed that Rcmapk3/9/10/11/13/14 exhibited a relatively broad expression distribution across 14 different tissues. For all these genes the highest expression level was in the brain, except for Rcmapk14 (highly expressed in the stomach, gill, and skin). The genes Rcmapk1/6/15 showed significantly higher expression in the testis. Under acute low salinity stress, expression of Rcmapk1/3/6/7/9/11/13/14 was significantly altered in the gill, intestine, and trunk kidney, however, the aforementioned genes exhibited very different expression patterns among the three tissues. In the gill, most of the genes from the erk (Rcmapk3/6/7) and p38 mapk subfamily (Rcmapk11/13/14) were significantly up-regulated at almost all the time points (P < 0.05); Similarly, the expression of Rcmapk3/9/11/13/14 genes were significantly increased in the trunk kidney; while in the intestine, most of the altered genes (Rcmapk6/7/9/11/13/14) were significantly down-regulated at 1 h. Following the sub-chronic low salinity stress, expression of Rcmapk1/3/6/7/9/11/13/14 genes were significantly altered in all three tissues. These findings provide important reference data for elucidating the roles of cobia mapk family genes in response to low salinity stress.

Transcriptomic analysis of turbot (Scophthalmus maximus) treated with zymosan a reveals that lncRNAs and inflammation-related genes mediate the protection conferred against Aeromonas salmonicida.

Aeromonas salmonicida is one of the most harmful pathogens in finfish aquaculture worldwide. Immunostimulants such as ß-glucans are used to enhance the immunity of cultured fish. However, their effects on fish physiology are not completely understood. In the present work, we evaluated the effect of a single intraperitoneal (ip) injection of zymosan A on fish survival against A. salmonicida infection. A single administration of this compound protected fish against A. salmonicida challenge and reduce the bacterial load in the head kidney one week after its administration. Transcriptome analyses of head kidney samples revealed several molecular mechanisms involved in the protection conferred by zymosan A and their regulation by long noncoding RNAs. The transcriptome profile of turbot exposed only to zymosan A was practically unaltered one week after ip injection. However, the administration of this immunostimulant induced significant transcriptomic changes once the fish were in contact with the bacteria and increased the survival of the infected turbot. Our results suggest that the restraint of the infection-induced inflammatory response, the management of apoptotic cell death, cell plasticity and cellular processes involving cytoskeleton dynamics support the protective effects of zymosan A. All this information provides insights on the cellular and molecular mechanisms involved in the protective effects of this widely used immunostimulant.

Circadian rhythm in turbot (Scophthalmus maximus): daily variation of blood metabolites in recirculating aquaculture systems.

INTRODUCTION: Animal welfare in aquaculture is becoming increasingly important, and detailed knowledge of the species concerned is essential for further optimization on farms. Every organism is controlled by an internal clock, the circadian rhythm, which is crucial for metabolic processes and is partially influenced by abiotic factors, making it important for aquaculture practices. OBJECTIVE: In order to determine the circadian rhythm of adult turbot (Scophthalmus maximus), blood samples were collected over a 24-h period and plasma metabolite profiles were analyzed by 1H-NMR spectroscopy. METHODS: The fish were habituated to feeding times at 9 am and 3 pm and with the NMR spectroscopy 46 metabolites could be identified, eight of which appeared to shift throughout the day. RESULTS: We noted exceptionally high values around 3 pm for the amino acids isoleucine, leucine, valine, phenylalanine, lysine, and the stress indicator lactate. These metabolic peaks were interpreted as either habituation to the usual feeding time or as natural peak levels in turbot in a 24-h circle because other indicators for stress (glucose, cortisol and lysozymes) showed a stable baseline, indicating that the animals had no or very little stress during the experimental period. CONCLUSION: This study provides initial insights into the diurnal variation of metabolites in adult turbot; however, further studies are needed to confirm present findings of possible fluctuations in amino acids and sugars. Implementing optimized feeding times (with high levels of sugars and low levels of stress metabolites) could lead to less stress, fewer disease outbreaks and overall improved fish welfare in aquaculture facilities.

pparß regulates lipid catabolism by mediating acox and cpt-1 genes in Scophthalmus maximus under heat stress.

Peroxisome proliferator-activated receptor ß (pparß) is a key gene-regulating lipid metabolism pathway, but its function in turbot remains unclear. In this study, the CDS of pparß was cloned from kidney for the first time. The CDS sequence length was 1533 bp encoding 510 amino acids. Structural analysis showed that the pparß protein contained a C4 zinc finger and HOLI domain, suggesting that the pparß gene of turbot has high homology with the PPAR gene of other species. The high expression patterns of pparß, acox, and cpt-1 at high temperatures, as shown through qPCR, indicated that high temperatures activated the transcriptional activity of pparß and increased the activity of the acox and cpt-1 genes. The expression of acox and cpt-1 was significantly inhibited when pparß was downregulated using RNAi technology and inhibitor treatments, suggesting that pparß positively regulated acox and cpt-1 expression at high temperatures and, thus, modulates lipid catabolism activity. These results demonstrate that pparß is involved in the regulation of lipid metabolism at high temperatures and expand a new perspective for studying the regulation of lipid metabolism in stress environments of teleost.

Tetraspanin CD53 regulates peripheral blood leucocytes vitality and pathogen infection in turbot (Scophthalmus maximus).

Cluster of differentiation 53 (CD53) also known as OX44 or tetraspanin 25 (TSPAN25) is a glycoprotein belonging to the tetraspanin family. Members of the tetraspanin family are characterized by four transmembrane domains, including intracellular N- and C-termini, and small and large extracellular domains. Currently, the function of CD53 in teleost is not well understood. In this study, we identified a CD53 (named SmCD53) from turbot (Scophthalmus maximus) and examined its expression and biological activity. SmCD53 contained 231 amino acid residues and was predicted to be a tetraspanin with small and large extracellular domains. SmCD53 expression was observed in different tissues, particularly in immune-related organs. Experimental infection with bacterial or viral pathogen significantly up-regulated SmCD53 expression in a time-dependent manner. Immunofluorescence microscopy analysis showed that SmCD53 was localized on the surface of PBL and was recognized by antibody against its large extracellular domain. Ligation of SmCD53 onto PBLs with antibodies suppressed the respiratory burst activity, inflammatory reaction, and enhanced cell viability. SmCD53 knockdown significantly enhanced bacterial dissemination and proliferation in turbot. Overall, these results underscore the importance of CD53 in the maintenance of the function and homeostasis of the immune system.

Assessing the role of ulvan as immunonutrient in Solea senegalensis.

Immunonutrition is a promising and viable strategy for the development of prophylactic measures in aquaculture. Ulvan, a sulphated marine polysaccharide from green seaweeds, has many biological activities including the immunomodulatory ones. The aim of this study was to assess the short and long-term effects of an ulvan-rich extract obtained from U. ohnoi as immunonutrient in Senegalese sole juveniles. In this work, an ulvan-rich extract from Ulva ohnoi has been obtained by the hot water method and isolated by ethanol precipitation. The FTIR analysis revealed that the ulvan-rich extact had very similar characteristics to previously published ulvan spectra. The total sulfate and protein content was 24.85 ± 3.98 and 0.91 ± 0.04 %, respectively. In vitro assays performed in Senegalese sole (Solea senegalensis) macrophages showed that the ulvan obtained in this study did not compromise the cell viability at concentrations up to 1 mg ml-1 and expression levels of lyg, irf1, il6, il10, c7, tf and txn were significantly upregulated in a concentration dependent-manner. Finally, S. senegalensis juveniles were fed basal diets and diets supplemented with the ulvan-rich extract at ratios 1 and 2 % for 30 days and then, challenged with Photobacterium damselae subsp. piscicida (Phdp). Thereafter, ulvan was withdrawn from the diet and all juveniles were fed the basal diet for 30 days. At 30 days post withdrawal (dpw), juveniles were challenged with Phdp. The expression profiles of a set of genes related to the immune system in spleen were evaluated as well as the lysozyme, peroxidase and bactericidal activity in plasma. Dietary effects of 1 % ulvan resulted in a boost of the immune response and increased disease resistance at short-term whereas juveniles fed diets supplemented with 2 % ulvan showed a significant decrease in the bactericidal activity and lack of protection against Phdp. At long-term (30 days after the withdrawal of ulvan), an improved response was observed in juveniles previously fed 1 % ulvan.

A comprehensive coding and microRNA transcriptome of vertebral bone in postlarvae and juveniles of Senegalese sole (Solea senegalensis).

Understanding vertebral bone development is essential to prevent skeletal malformations in farmed fish related to genetic and environmental factors. This is an important issue in Solea senegalensis, with special impact of spinal anomalies in postlarval and juvenile stages. Vertebral bone transcriptomics in farmed fish mainly comes from coding genes, and barely on miRNA expression. Here, we used RNA-seq of spinal samples to obtain the first comprehensive coding and miRNA transcriptomic repertoire for postlarval and juvenile vertebral bone, covering different vertebral phenotypes and egg-incubation temperatures related to skeleton health in S. senegalensis. Coding genes, miRNA and pathways regulating bone development and growth were identified. Differential transcriptomic profiles and suggestive mRNA-miRNA interactions were found between postlarvae and juveniles. Bone-related genes and functions were associated with the extracellular matrix, development and regulatory processes, calcium binding, retinol and lipid metabolism or response to stimulus, including those revealed by the miRNA targets related to signaling, cellular and metabolic processes, growth, cell proliferation and biological adhesion. Pathway enrichment associated with fish skeleton were identified when comparing postlarvae and juveniles: growth and bone development functions in postlarvae, while actin cytoskeleton, focal adhesion and proteasome related to bone remodeling in juveniles. The transcriptome data disclosed candidate coding and miRNA gene markers related to bone cell processes, references for functional studies of the anosteocytic bone of S. senegalensis. This study establishes a broad transcriptomic foundation to study healthy and anomalous spines under early thermal conditions across life-stages in S. senegalensis, and for comparative analysis of skeleton homeostasis and pathology in fish and vertebrates.

Chronic low salinity stress rescued masculinization effect in farmed Cynoglossus semilaevis population.

Salinity, being an indispensable abiotic factor crucial for the survival of marine organisms, has demonstrated diverse alterations globally in response to the current trend of global warming. In this study, the effect of chronic low salinity stress on teleosts' sex differentiation was investigated using Cynoglossus semilaevis, an economically important fish with both genetic and environmental sex determination system. The cultivation experiment was conducted employing artificially simulated seawater of 20 ppt and ambient sea water of 30 ppt to rear juveniles C. semilaevis. Throughout the experiment, the growth performance was assessed and the histology of gonadal development was examined, a significantly lower masculinization rate was observed in LS group. To gain further insights, transcriptome analysis was conducted using raw reads obtained from 53 libraries derived from gonads of 55 days post fertilization (dpf) and 100 dpf juveniles in both LS and CT groups. GO/KEGG enrichment were further proceeded, Terms and pathways involved in reproduction ability, germ cell proliferation, immune function, steroid metabolism etc., were illuminated and a possible crosstalk between HPI and HPG axis was proposed. WGCNA was conducted and two hub genes, hspb8-like and Histone H2A.V were exhibited to be of great significance in the changes of masculinization rate. Our findings provided solid reference for sex differentiation study of GSD + ESD species in a constantly changing ocean environment, as well as practice guiding significance for the environmental management for the culture of C. semilaevis.

Differential modulation of digestive enzymes and energy reserves at different times after feeding in juveniles of the marine estuarine-dependent flatfish Paralichthys orbignyanus (Valenciennes, 1839).

Integrative studies are lacking on the responses of digestive enzymes and energy reserves in conjunction with morphological traits at distinct postprandial times in marine estuarine-dependent flatfishes of ecological and economic importance, such as Paralichthys orbignyanus. We determined total weight (TW), hepato-somatic index (IH), activities of digestive enzymes in the intestine, and the concentration of energy reserves in the liver and the muscle at 0, 24, 72, and 360 h after feeding in juveniles of P. orbignyanus. Amylase activity decreased at 72 h (about 30%). Maltase, sucrose, and lipase activities reached peak at 24 h (67%, 600%, and 35%, respectively). Trypsin and aminopeptidase-N activities at 24 and 72 h, respectively, were lower than those at t = 0 (53% and 30%). A peak increase in the concentration of glycogen and triglycerides in the liver (24 h) (86% and 89%, respectively) occurred. In muscle, glycogen and triglyceride concentrations were unchanged at 24 h and higher at 72 and 360 h (100% and 60%). No changes were found in TW, IH, free glucose in the liver and muscle, and protein in the liver. The protein concentration in the muscle sharply increased at 24 and 360 h after feeding (60%). The results indicate a distinct and specific response of central components of carbohydrate, lipid, and protein metabolism that could be adjustments at the biochemical level upon periods of irregular feeding and even of long-term food deprivation inside coastal lagoons or estuaries. The distinct responses of digestive enzymes in the intestine and energy reserves in the liver and muscle suggest the differential modulation of tissue-specific anabolic and catabolic pathways that would allow the maintenance of physical conditions.

Comparative transcriptome analysis reveals growth and molecular pathway of body color regulation in turbot (Scophthalmus maximus) exposed to different light spectrum.

Fish body color changes play vital roles in adapting to ecological light environment and influencing market value. However, the initial mechanisms governing the changes remain unknown. Here, we scrutinized the impact of light spectrum on turbot (Scophthalmus maximus) body coloration, exposing them to red, blue, and full light spectra from embryo to 90 days post hatch. Transcriptome and quantitative real-time PCR (qRT-PCR) analyses were employed to elucidate underlying biological processes. The results showed that red light induced dimorphism in turbot juvenile skin pigmentation: some exhibited black coloration (Red_Black_Surface, R_B_S), while others displayed lighter skin (Red_White_Bottom, R_W_B), with red light leading to reduced skin lightness (L*) and body weight, particularly in R_B_S group. Transcriptomic and qRT-PCR analyses showcased upregulated gene expressions related to melanin synthesis in R_B_S individuals, notably tyrosinase (tyr), tyrosinase-related protein 1 (tyrp1), and dopachrome tautomerase (dct), alongside solute carrier family 24 member 5 (slc24a5) and oculocutaneous albinism type II (oca2) as pivotal regulators. Nervous system emerged as a critical mediator in spectral environment-driven color regulation. N-methyl d-aspartate (NMDA) glutamate receptor, and calcium signaling pathway emerged as pivotal links intertwining spectral conditions, neural signal transduction, and color regulation. The individual differences in NMDA glutamate receptor expression and subsequent neural excitability seemed responsible for dichromatic body coloration in red light-expose juveniles. This study provides new insights into the comprehending of fish adaptation to environment and methods for fish body color regulation and could potentially help enhance the economic benefit of fish farming industry.

Effects of miR-722 on gene expression and alternative splicing in the liver of half-smooth tongue sole after infection with Vibrio anguillarum.

MicroRNAs play crucial roles in various biological processes, including but not limited to differentiation, development, disease, and immunity. However, their immunoregulatory roles in half-smooth tongue sole are lacking. Our previous studies indicated that miR-722 could target C5aR1 to modulate the complement pathway to alleviate inflammatory response and even affect the mortality after the bacterial infection with Vibrio anguillarum. Driven by the purpose of revealing the underlying mechanisms, in this study, we investigated the effects of miR-722 on the gene expression and alternative splicing (AS) in the liver of half-smooth tongue sole after Vibrio anguillarum infection, with the approach of miR-722 overexpression/silencing and subsequent RNA-seq. Among the different comparisons, the I group (miR-722 inhibitor and V. anguillarum) versus blank control (PBS) exhibited the highest number of differentially expressed genes (DEGs), suggesting that the immune response was overactivated after inhibiting the miR-722. In addition, enrichment analyses were performed to reveal the functions of DEGs and differential AS (DAS) genes, reflecting the enrichment of RNA splicing and immune-related pathways including NF-κB and T cell receptor signaling pathway. Comparing the M group (miR-722 mimic and V. anguillarum) with the negative control (random sequence and V. anguillarum), two immune-related genes, cd48 and mapk8, were differentially expressed, of which mapk8 was also differentially spliced, indicating their importance in the immune response. Furthermore, representative gene analysis was performed, suggesting their corresponding functional changes due to AS. To verify the RNA-seq data, quantitative real-time PCR was employed with twenty pairs of primers for DEGs and DAS events. Overall, our results demonstrated that miR-722 could mediate the transcriptome-wide changes of gene expression and AS in half-smooth tongue sole, and provided insights into the regulatory role of miR-722 in immune responses, laying the foundation for further functional analyses and practical applications in aquaculture.

CsIL-20, a tongue sole interleukin-20, negatively mediates leucocyte activity and antibacterial defense.

Interleukin-20 (IL-20), as an essential member of IL-10 family, plays vital roles in mammalian immunological response such as antimicrobial, inflammation, hematopoiesis, and immune diseases. In teleost, the study about immune antimicrobial function of IL-20 is largely scarce. In this article, we revealed the expression profiles and the immunological functions of the IL-20 (CsIL-20) in tongue sole Cynoglossus semilaevis. CsIL-20 is composed of 183 amino acid residues, with seven cysteine residues and a typical IL-10 domain which comprises six α-helices and two ß-sheets, and shares 34.4-71.2 % identities with other teleost IL-20. CsIL-20 was constitutively expressed in a variety of tissues and regulated by bacterial invasion, and the recombinant CsIL-20 (rCsIL-20) could bind to different bacteria. In vitro rCsIL-20 could interact with the membrane of peripheral blood leukocytes (PBLs), leading to the attenuation of reactive oxygen species (ROS) production and acid phosphatase activity in PBLs. In line with In vitro results, In vivo rCsIL-20 could obviously suppressed the host immune against bacterial infection. Furthermore, knockdown of CsIL-20 in vivo could markedly enhance the host antibacterial immunity. Collectively, these observations offer new insights into the negative effect of CsIL-20 on antibacterial immunity.