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Black porgy is an important marine aquaculture fish species whose production is at the fifth position in all kinds of marine-cultured fishes in China. In this study, Illumina high-throughput sequencing technology was used to sequence the total RNA of black porgy. Sixty-one candidate SNPs (Single Nucleotide Polymorphism) were screened out and genotyped through GATK4 (Genome Analysis ToolKit) software and MALDI-TOF MS (Matrix-Assisted Laser Desorption/ Ionization Time of Flight Mass Spectrometry). The experimental results showed that a total of sixty SNPs were successfully genotyped, with a success rate of 98.36%. The results of principal component analysis and correlation analysis of growth traits showed that body weight was the first principal component, with a cumulative contribution rate of 74%. There were significant correlations (p < 0.05) or extremely significant correlations (p < 0.01) between different growth traits. The results of genetic parameter analysis and association analysis showed that scaffold12-12716321, scaffold13-4787950, scaffold2-13687576 and scaffold290-11890 were four SNPs that met the requirement of polymorphic information content and conformed to the Hardy–Weinberg equilibrium. There were significant differences between their genotype and the phenotype of growth traits. The four SNP molecular markers developed in this research will lay a foundation for further exploration of molecular markers related to the growth traits of black porgy and will provide a scientific reference for the further study of its growth mechanisms. At the same time, these molecular markers can be applied to the production practices of black porgy, so as to realize selective breeding at the molecular level and speed up the breeding process.

Fatty acyl desaturase 2 (fads2) is a rate-limiting enzyme in long chain polyunsaturated fatty acids (LC-PUFAs) biosynthesis. In mammals, the lipid metabolism is modulated by a transcription factor, peroxisome proliferator-activated receptor alpha β (pparαβ); however, the detailed mechanism via pparαβ regulates fads2 remains unclear in fish. In the present study, we identified the sequence features of Trachinotus ovatus fatty acyl desaturase 2a (Tofads2a) and fatty acyl desaturase 2b (Tofads2b), which both encoded 442 amino acid polypeptides containing cytochrome-b5-like domains and three representative histidine-rich domains. The Phylogenetic and genome organization analyses revealed characteristic phylogeny: the majority of fads2s exhibited a highly conserved exon/intron architecture. Tissue expression patterns by quantitative real-time PCR (qRT-PCR) showed that the two Tofads2s were prominently expressed in the brain. A nutritional study indicated that the transcription of the two Tofads2s was significantly implicated by treatment with a 1: 1 ratio of fish oil: soybean oil (FO:SO) in the liver and brain. Furthermore, functional characterization in yeast demonstrated that both Tofads2a and Tofads2b possessed Δ4/Δ5/Δ8 desaturation activity. Furthermore, promoter activity assays showed that the expressions of the two Tofads2s were actively regulated by pparαβ. Moreover, mutation analyses showed that the M1 and M4/M5 binding sites of pparαβ were functionally vital for binding to Tofads2a and Tofads2b promoters, respectively. Transcriptional activities of the two Tofads2s promoters were significantly reduced after targeted mutation of M1 or M4/M5. Electrophoretic mobile shift assays (EMSAs) verified that pparαβ interacted with the M1 binding site in Tofads2a promoter to accommodate Tofads2a transcription. Briefly, pparαβ plays an important role in Tofads2 expression and may promote the LC-PUFAs biosynthesis by regulating the expression of two Tofads2s.

The whole mitochondrial genome ofLateolabrax maculatus(Cuvier, 1828) was used to investigate the reasons for the observed patterns of genetic differentiation among 12 populations in northern and southern China. The haplotype diversity and nucleotide diversity ofL. maculatuswere 0.998 and 0.00169, respectively. Pairwise FSTvalues between populations ranged from 0.001 to 0.429, correlating positively with geographic distance. Genetic structure analysis and haplotype network analysis indicated that these populations were split into two groups, in agreement with geographic segregation and environment. Tajima’s D values, Fu’s Fs tests and Bayesian skyline plot (BSP) indicated that a demographic expansion event may have occurred in the history ofL. maculatus. Through selection pressure analysis, we found evidence of significant negative selection at the ATP6, ND3, Cytb, COX3, COX2 and COX1 genes. In our hypotheses, this study implied that demographic events and selection of local environmental conditions, including temperature, are responsible for population divergence. These findings are a step forward toward the understanding of the genetic basis of differentiation and adaptation, as well as conservation ofL. maculatus.

Myoblast determination protein (MyoD), a muscle-specific basic helix-loop-helix (bHLH) transcription factor, plays a pivotal role in regulating skeletal muscle growth and development. However, the regulation mechanism of MyoD has not been determined in marine fishes. In the present study, we isolated the MyoD1 (AlMyoD1) and MyoD2 (AlMyoD2) genomic sequences and analyzed the expression patterns in different tissues of yellowfin seabream (Acanthopagrus latus). The open reading frame (ORF) sequences of AlMyoD1 and AlMyoD2 encoded 297 and 271 amino acids possessing three common characteristic domains, respectively, containing a myogenic basic domain, a bHLH domain, and a ser-rich region (helix III). Phylogenetic and genome structure analyses exhibited classic phylogeny and highly conserved exon/intron architecture. Furthermore, the AlMyoD1 and AlMyoD2 transcription levels were higher in white muscle than in the other tissues. In order to further study AlMyoD function in muscle, promoter sequence analysis found that several E-box binding sites were present. Additionally, binding sites of Almyomixer involved in mammal myoblast fusion, which expression was also the highest in white muscle, were found in the promoter of AlMyoD. Pomoter activity assays further confirmed that both AlMyoD1 and AlMyoD2 can dramatically activate Almyomixer expression, and the AlMyoD1 M2 and AlMyoD2 M5 E-box binding sites were functionally important for Almyomixer transcription based on mutation analysis and electrophoretic mobile shift assays (EMSA). In summary, two MyoDs play a core role in Almyomixer regulation and may promote myofibre formation during muscle development and growth by regulating Almyomixer expression.

Golden pompano Trachinotus ovatus (Linnaeus, 1758) is an important mariculture fish species with high commercial value in China. The present study thoroughly assessed the types and frequencies of skeletal deformities at the early developmental stages of golden pompano in an intensive aquaculture production system. Golden pompano (n = 500) were sampled 30 d posthatch (dph). The specimens were stained with Alcian blue and Alizarin red for the detection of deformities. The results of the study revealed that 77.2% of the specimens showed at least 1 spinal anomaly; most anomalies occurred in the prehemal region, and the most common deformity observed was vertebral fusion (37.4% incidence of deformities). The results of this study provide useful information for the early detection of skeletal deformities and for the optimization of fish fry breeding technologies.

The golden pompano (Trachinotus ovatus) is a marine fish species in the family Carangidae. We constructed a chromosome-level genome assembly of a male golden pompano. QTL-mapping and GWAS analysis showed that this species has a ZZ/ZW sex determination system and a sex defining SNP (Chr16:18219150:G/A), located on the splice donor site (GT-AG) of the first intron of Hsd17b1, was exclusively associated with the phenotypic sex. The W-linked coding sequences of Hsd17b1 were conserved across vertebrates, while Z-linked coding sequences introduced extra 64 bases and were malfunctional. The golden pompano and the greater amberjack (Seriola dumerili), divergent in 57 million years ago in the same family, share the same features of sex determination, including the same sex determining gene, malfunctional Z-linked haplotypes, undifferentiated sex chromosomes except that the sex defining SNPs are different. In simulation analysis, turnover of sex determining mutation, single mutation dominating sex determination and undifferentiated sex chromosomes were also observed. We proposed a hypothesis that W-linked haplotypes of the sex determining gene of Hsd17b1 were under purifying selection, Z-linked haplotypes may evolve near neutrally, recurrent and directional transformations from W-linked haplotypes to Z-linked haplotypes caused by inactivating mutations, relatively strong forces of drift and recombination comprehensively contributed to turnover of sex defining mutation and undifferentiation of sex chromosome. We also established zebrafish mutants and homozygous mutants were “all male”, which indirectly supported this hypothesis.

Heat shock protein 90 family, a class highly conserved protein and important molecular chaperone and its plays a role in enhancing animal adaptability to environmental stress and improving immunity. In this study, a full-length HSP90β cDNA (ToHSP90β) sequence was identified from Trachinotus ovatus. The cDNA of ToHSP90β was 2648 base-pairs (bp) long and contained a 5′-terminal untranslated region of 130 bp, a 3′-terminal untranslated region of 334 bp, and the open reading frame length 2184 bp, which can encode 728 amino acids. The ToHSP90β has a heat shock protein 90 family signature. The ToHSP90β amino acid sequence has high homology with other fish HSP90β amino acids (92.5-99.17%) and contains five conserved signal regions and a structural feature of the cytoplasmic HSP90 family. Phylogenetic tree analysis indicated that ToHSP90β was most closely related to Seriola dumerili HSP90β. Tissues expression analysis showed that the ToHSP90β was expressed in all nine tissues of T. ovatus, and its expression was the highest in the intestine. The stocking density significantly affected the expression of ToHSP90β in the kidney and brain tissues, and its expression levels increased significantly with increasing stocking densities, indicating that it plays an important regulatory role in the process of T. ovatus’ chronic stress response. After Cryptocaryon irritans infection, the mRNA expression of ToHSP90β was significantly increased in liver and kidney tissues, indicating that ToHSP90β would be involved in the immune regulatory response. This study was the first to analyze the expression pattern of HSP90β mRNA in T. ovatus tissues and their expression responses to stocking density stress and C. irritans infection. To provide a theoretical basis for further study of the function of fish HSP90 and its physiological mechanism in environmental stress and parasite infection.

This study aimed to investigate the intoxication mechanism of golden pompano (Trachinotus ovatus) exposed to high ammonia levels and the effects on the immune and antioxidant mechanisms of gills. Juvenile golden pompano was exposed to ammonia (total ammonia: 26.9 mg/L) to induce 96 h of ammonia stress, and a 96 h recovery experiment was performed after poisoning. Then, we evaluated hematological parameters, the histological structure and the expression of related genes. In this experiment, continuous exposure to high levels of ammonia led to a significant increase in plasma alkaline phosphatase (ALP), acid phosphatase (ACP) and lactate dehydrogenase (LDH) levels (P < 0.05), and the levels of triiodothyronine (T3) and tetraiodothyronine (T4) were significantly reduced (P < 0.05). Moreover, the expression of antioxidant genes such as superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPX) and inflammatory cytokines such as tumor necrosis factor α (TNF-α) and interleukin 1β (IL-1β) increased (P < 0.05). These results indicate that ammonia activates the active osmotic regulatory mechanism of fish gills and participates in defense and immune responses. However, with prolonged exposure to ammonia, the balance of the defense system is disrupted, leading to oxidative damage and inflammation of the gill tissue. This research not only helps elucidate the intoxication mechanism of golden pompano by ammonia at the molecular level but also provides a theoretical basis for further research on detoxification mechanisms.

Golden pompano (Trachinotus ovatus), a marine fish in the Carangidae family, has a wide geographical distribution and adapts to severe environmental rigours. It is also an economically valuable aquaculture fish. To understand the genetic mechanism of adaption to environmental rigours and improve the production in aquaculture, we assembled its genome. By combination of Illumina and Pacbio reads, the obtained genome sequence is 647.5 Mb with the contig N50 of 1.80 Mb and the scaffold N50 of 5.05 Mb. The assembly covers 98.9% of the estimated genome size (655 Mb). Based on Hi-C data, 99.4% of the assembled bases are anchored into 24 pseudo-chromosomes. The annotation includes 21,915 protein-coding genes, in which 95.7% of 2,586 BUSCO vertebrate conserved genes are complete. This genome is expected to contribute to the comparative analysis of the Carangidae family., Measurement(s)DNA • chromosome conformation capture assay • transcription profiling assayTechnology Type(s)DNA sequencing • Hi-C • RNA sequencingFactor Type(s)organism partSample Characteristic - OrganismTrachinotus ovatusSample Characteristic - Environmentocean biome Machine-accessible metadata file describing the reported data: 10.6084/m9.figshare.9933515

Golden pompano (Trachinotus ovatus) is a commercially important marine fish and is widely cultured in the coastal area of South China. Salinity is one of the most important environmental factors influencing the growth and survival of fish. The aims of this study are to investigate the growth, physiological, and molecular responses of juvenile golden pompano reared at different salinities. Juveniles reared at 15 and 25‰ salinity grew significantly faster than those reared at the other salinities. According to the final body weights, weight gain rate, and feed conversion ratio, the suitable culture salinity range was 15-25‰ salinity. The levels of branchial NKA activity showed a typical "U-shaped" pattern with the lowest level at 15‰ salinity, which suggested a lower energy expenditure on osmoregulation at this level of salinity. The results of this study showed that the alanine aminotransferase, aspartate aminotransferase, and cortisol of juveniles at 5‰ were higher than those of other salinity groups. Our results showed that glucose-6-phosphate dehydrogenase significantly increased at 5‰ and 35‰ salinity. Our study showed that osmolality had significant differences in each salinity group. GH, GHR1, and GHR2 had a wide range of tissue expression including the liver, intestine, kidneys, muscle, gills and brain. The expression levels of GH, GHR1 and GHR2 in the intestine, kidneys, and muscle at 15‰ salinity were significantly higher than those in other three salinity groups. Based on the growth parameters and physiological and molecular responses, the results of the present study indicated that the optimal salinity for rearing golden pompano was 21.36‰ salinity.

In this study, sequence analysis showed that ToGHR1 and ToGHR2 encoded polypeptides of 577 and 588 amino acids, respectively. Bioinformatics analysis showed that both ToGHR1 and ToGHR2 contain FN3 domains and transmembrane domains, which have glycosylation and phosphorylation sites. The exons of ToGHR1 and exons 4–10 of ToGHR2 are homologous to exons 2 and 4–9 in Homo sapiens genes, respectively. Only 3 SSR sites in ToGHR1 have SSR polymorphisms, and ToGHR2 has no SSR polymorphisms. ToGHR1 and ToGHR2 have high homology with GHR1 and GHR2 of many fish by BLAST. qRT-PCR was used to examine the expression profile of ToGHR mRNA in 12 normal liver and intestine tissue samples from 3 feed-type groups. The results showed that ToGHR is expressed in all 12 tissues, especially liver and muscle tissues, which showed higher ToGHR expression than that in other tissues (p

Longfin batfish (Platax teira) is a coastal marine fish life. The aims of this study are to investigate the compositions and content of amino acid and fatty acid in three parts of P. teira. The results showed that P. teira muscle protein content was as high as 21.00%. For amino acid composition, seventeen kinds of amino acids were detected in three parts. The essential amino acid score (EAAS) and essential amino acids (EAA) / total amino acids (TAA) ratio of P. teira were both above 70 and 0.4. EAAS in the upper part of the abdominal muscle showed higher than those two parts. The content of saturated fatty acids (SFA) and the polyunsaturated fatty acids (PUFA) in the upper part of the abdominal muscle was higher than that in other groups. No significant differences in monounsaturated fatty acids (MUFA) and n-3/n- 6 were detected among three parts. Overall, the upper part of the abdominal muscle was thought to have a higher nutritious value. These results showed P. teira is a high quality marine fish with high protein content and low fat, abundant amino acids with proper proportion, however, it was regarded as an excellent breeding varieties.

Golden pompano (Trachinotus ovatus) is a commercially important marine fish and is widely cultured in the coastal area of South China. We explored the distribution difference of intestinal microflora with the same growth condition and the difference of gut microflora with different growth rates fed with fermented soybean meal (FSM) and fish meal (FM) primary protein source. After 8 weeks of the breeding experiment, select significant growth fish (Zcd, 156.04 ± 1.04 g) average growth fish (Zc, 120.01 ± 1.05 g) stagnant growth fish (Zcx, 98.03 ± 1.08 g) in FSM and select average growth fish (Dc, 158.03 ± 1.02 g) in FM. Sequencing results show that a total of 86303 sequences had been obtained in the intestine of Trachinotus ovatus. The number of OTUs that can be annotated to the database was 2315 (98.97%). At the phylum level, Proteobacteria, Tenericutes, Spirochaetes and Firmicutes were the predominant phyla. Alpha diversity and beta diversity analyses show that gut microbes’ richness and diversity in the Dc group were the highest. The diversity of gut microbiota in the FSM: Zc > Zcd >Zcx. At the genus classification level, the differences between FM and FSM were mainly in Gueglia, Grimontia, Klebsiella, Alistipes, Staphylococcus, and Parasutterella (p

The ontogeny of the digestive tract in Platax teira was studied by means of optical microscopy from hatching to 30 days post-hatching. Collect fertilized eggs from P. teira broodstock, fertilize the eggs to hatch in hatching net bags, and rear the larvae in higher-place ponds. Observe the morphology and structure of the digestive tract of larvae 1-30 days post-hatching using tissue slicing technique. The results showed that the oral cavity of the larvae was closed and had not yet begun to differentiate at 1day post-hatching. Its digestive tract was close to the abdominal wall, and the yolk sac was completely dyed dark red. At 4 days post-hatching, the oropharyngeal cavity, esophagus, and stomach of the larvae had been communicated, and their oral cavity opened and food intake begins, entering the mixed period of endogenous nutrition and exogenous nutrition. Thereafter, the yolk sac gradually decreased to disappear, the esophagus, stomach, and intestines continued to differentiate, and their functions gradually improved. During 25-30 days post-hatching, the structure of the digestive tract of the larvae didn’t change much. Due to the increase in the size of the larvae, most changes in the digestive system are related to the complexity of the tissues and the size of the organs. Specifically, the number of mucosal folds and cells in the esophagus and intestine increases, and the gastric glands and pits in the stomach increase. The number of mucous membrane folds, mucosal epithelial cells and gastric glands in the digestive tract increase with the growth of the larvae, indicating that the development of the digestive system of P. teira is consistent with the growth of larvae, morphological development, feeding, metabolism and other functions consistently, its developmental stage is also adapted to the stages of the early development stage of P. teira.

The yellowfin seabream Acanthopagrus latus is the economically most important Sparidae fish species in the northern South China Sea. As euryhaline fish, they are perfect model for investigating osmoregulatory mechanisms in teleosts. Moreover, the reproductive biology of hermaphrodites has long been intriguing; however, very little is known about the molecular pathways underlying their sex change. To elucidate genetic mechanisms of osmoregulation and sex reversal in this fish, a high-quality reference genome of the yellowfin seabream was generated by a combination of Illumina and PacBio technologies. The draft genome of yellowfin seabream was 806 Mb, with 732 Mb scaffolds anchored on 24 chromosomes. The contig N50 and scaffold N50 were 2.6 Mb and 30.17 Mb, respectively. The assembly is of high integrity and includes 92.23% universal single-copy orthologues based on benchmarking universal single-copy orthologs (BUSCO) analysis. Moreover, among the 19,631 protein-coding genes, we found that the ARRDC3 and GSTA gene families related to osmoregulation underwent an extensive expansion in two euryhaline Sparidae fish genomes compared to other teleost genomes. Moreover, integrating sex-specific transcriptome analyses, several genes related to the transforming growth factor beta (TGF-β) signalling pathway involved in sex differentiation and development. This genomic resource will not only be valuable for studying the osmoregulatory mechanisms in estuarine fish and sex determination in hermaphrodite vertebrate species, but also provide useful genomic tools for facilitating breeding of the yellowfin seabream.

Thymosin β4 is a multifunctional protein in vertebrates that participates in physiological processes, such as wound healing, immune response, cell proliferation and migration. We assessed the multifarious roles of this small peptide in Pinctada fucata, an oyster commonly used in pearl culture in China. Our results showed that when P. fucata was challenged by bacterial pathogens or LPS, the relative expression level of Pfthymosin β4 mRNA was significantly up-regulated, suggesting its involvement in immune response of the animal. Recombinant Pfthymosin β4 (rPfthymosin β4) was produced and showed in vitro different antibacterial activities against different pathogenic bacteria; the inhibitory effect of rPfthymosin β4 on bacterial growth was relatively stronger in the broth culture than agar culture. The overexpression of Pfthymosin β4 in Escherichia coli BL21(DE3) cells could improve their resistance to Cu2+, Zn2+, Cd2+, and H2O2, suggesting that Pfthymosin β4 is likely involved with antioxidant. rPfthymosin β4 also significantly promoted the proliferation and migration of mouse aortic vascular smooth muscle cells as indicated by MTT assay and cell scratch assay, respectively. In addition, chemically synthesized or recombinant Pfthymosin β4 could transiently increase the circulating total hemocytes counts but down-regulated by RNAi in P. fucata. Taking together above results and previous studies suggested that Pfthymosin β4 is potentially able to promote wound healing through enhancing antibacterial activity and antioxidant capacity, promotion of cell proliferation and migration, and increase of circulating hemocytes in P. fucata due to nucleus implantation injury. Thus, the future of recombinant Pfthymosin β4 should be promising in the culture of pearls in P. fucata.

This study uses the CASA system and enzyme activity kit to detect movement parameters and enzyme activities of fresh and cryopreserved yellowfin seabream (Acanthopagrus latus) sperm, which could provide theoretical support for the protection of the germplasm resources of yellowfin seabream and the sustainable and healthy development of its aquacultural industry. The results showed that MOT, ALH, BCF and other indices were significantly reduced after cryopreservation (P 0.05). CAT, SOD, CK and other enzyme activities of fresh sperm decreased significantly after cryopreservation (P 0.05). After the second freezing treatment, the enzyme activities of SOD, T-GSH, ATP, CK and SDH of sperm were significantly reduced (P 0.05). When the cryopreservation time was extended from 5 days to 20 days, the enzyme activities of CAT, SOD, ATP and CK in sperm tended to increase first and then decrease, while the enzyme activities of GSH-Px and LDH tended to remain stable first and then decreased. This study indicates that cryopreservation has some effect on sperm motility parameters and that the activity compared with fresh sperm is slightly reduced. Cryopreservation causes damage to the antioxidant system of sperm to some extent. It also reduces the antioxidant capacity of antioxidant enzymes and energy metabolizing enzymes to generate energy.

Trachinotus ovatus is widely cultured in the ponds and gulf on the southeast coast of China. The dramatic salinity decrease caused by heavy rainfall could cause mass mortality of T. ovatus in aquaculture. It is very important to understand the osmoregulatory mechanism of T. ovatus. Na+/K+/2Cl− cotransporter 1a (NKCC1a) is involved in the osmoregulation of fish and plays a crucial role in cell volume homeostasis and maintenance of the electrolyte content. In this study, we characterized nkcc1a (designed as Tonkcc1a) from T. ovatus and investigated its expression responses to acute salinity changes. Tonkcc1a is approximately 70 kb in length and contains 26 exons and 25 introns. The phylogenetic analysis confirmed that ToNKCC1a belonged to the NKCC1a subclade. Quantitative real-time (qRT-PCR) analysis indicated that Tonkcc1a was ubiquitously expressed in all examined tissues, with the highest mRNA levels observed in gills, and the lowest level in liver. When T. ovatus were transferred from seawater (30‰) into fresh water, the expression levels of Tonkcc1a mRNA were significantly downregulated in gills and kidney, whereas its expression level was markedly upregulated in intestine. When transferred from seawater (30‰) to 10‰ sea water, the expression levels of Tonkcc1a mRNA were clearly increased in gills and kidney. When transferred from seawater (30‰) to 20‰ sea water, the expression of Tonkcc1a mRNA increased to some extent in gills, kidney, and intestine. When transferred from seawater (30‰) to 40‰ sea water, the expression levels of Tonkcc1a mRNA were dramatically upregulated in gills and intestine compared to that in the control. These results suggested that Tonkcc1a was involved in the response to acute salinity changes.

Golden pompano, Trachinotus ovatus, belongs to the family Carangidae. Within one decade, this species has rapidly become one of the most important cultured marine fish species in South China. However, the lack of a comprehensive genetic diversity assessment hinders the conservation of natural resources and management of cultured populations. Thus, we sampled one wild population and six cultured populations of golden pompano, which represented the whole cultured population, to assess the genetic diversity and population structure. The level of genetic diversity was low compared with those of other cultured fish, as the values of allelic richness, number of effective alleles and expected heterozygosity in the combined whole sample were 3.709, 2.592, and 0.591, respectively. These populations were little differentiated (Fst = 0.02091, p value = 0.00), and the whole sample did not show an explicit population structure at the individual level. The effective population size in each cultured population was small and in the whole sample was acceptable for a closed selection system. The pedigree reconstruction showed no evidence of artificial disturbance in mating. This general survey of cultured golden pompano populations showed the common characteristics of domestication with wild inputs. However, the low level of and potential decrease in genetic diversity should receive close attention in future breeding programs. In addition, we recommend wide surveys of natural resources and the establishment of closed breeding systems to increase genetic gain.

The interferon-g-inducible lysosomal thiol reductase (GILT) plays a significant character in the processing and presentation of MHC class II restricted antigen (Ag) by catalyzing disulfide bond reduction in mammals. To explore the function of GILT in the immune system of fish, we cloned a GILT gene homologue from Trachinotus ovatus, the full-length cDNA of GILT, which consisted of 2, 747 bp with a 771 bp open reading frame, encoding a protein of 256 amino acids. Moreover, similar to other species GILT gene, 7 exons and 6 introns were identified in T. ovatus, the deduced protein also possessed a representative characteristic of known GILT proteins. The result of real-time quantitative PCR showed that GILT mRNA was dramatically expressed in immune-associated tissues, such as spleen (p 0.01) and kidney (p 0.05). Bacterial challenge revealed that GILT mRNA level remarkably up-regulation in liver, spleen, kidney and intestine after induction with Photobacterium damsela. Furthermore, based on cloned sequences and genome BLAST, only one SNP site (ToGILT-S1-g.148CG) was identified, and the allele C was significantly associated with high-susceptibility (HS) group, nevertheless, the allele G was dramatically associated with high-resistance (HR) group, indicating potential application for disease resistant breeding selection in T. ovatus.

Trachinotus ovatus is an important economic species in China, with an annual output of over 100,000 tons (FAO). The major histocompatibility complex (MHC) genes, which are a highly polymorphic gene superfamily, play a vital role in the adaptive immune system. In our study, due to the threat caused by Photobacterium damselae, the associations between polymorphisms of exon 2 of MHC IIs and disease resistance/susceptibility were analysed in Trachinotus ovatus. The gene structure of MHC IIs was composed of six exons and five introns in T. ovatus, similar to Hippocampus erectus MHC IIs. Among the forty-three alleles, high levels of polymorphism were found (24.8% of nucleotide positions were variable). In the peptide-binding region (PBR) and non-PBR sites, the value of dN was 6.60 and 12.22 (> 1) times higher than the value of dS, respectively. Six alleles were chosen to calculate their distributions in the high-resistance (HR) and high-susceptibility (HS) groups. The analysis of the associations between alleles and resistance/susceptibility to P. damselae indicated that the TO-DAB-01 allele was associated with immunity to P. damselae and the TO-DAB-04, TO-DAB-05 and TO-DAB-10 alleles were associated with its sensitivity to P. damselae. The results suggest that the diversity of exon 2 of MHC IIs alleles could be used to explore the association between disease susceptibility/resistance and the multiformity of MHC IIs and to achieve the molecular-assisted selection of T. ovatus with enhanced disease resistance.

The golden pompano, Trachinotus ovatus, is a promising species in the mariculture. However, the sex identification in alive individual is a bottleneck step in selective breeding. The golden pompano and the greater amberjack, Seriola dumerili, belong to the same family, Carangidae. The genome assembly of golden pompano has been released and the sex determining mechanism in the greater amberjack has been deeply understood. The genome assembly of the golden pompano was improved with Hi-C sequencing and the presumed sex determining region was detected by aligning with the sex determining region of the greater amberjack. A SNP (Chr17:8325779) within the gene Hsd17b1 was successfully converted into a sex-specific marker that can be genotyped by kompetitive allele specific PCR and high resolution melting analysis with 100% the accuracy of sex identification. The results that only one presumed sex determining region was detected with gene Hsd17b1 lied in and the genotype of the developed sex-specific marker was heterozygous in female and homozygous in male supported the sex chromosome system was ZZ/ZW and the candidate sex determining gene was Hsd17b1. The results would benefit the selective breeding program of the golden pompano.

Continuous exposure to high levels of ammonia can cause oxidative damage to fish tissues and organs. To date, the mechanism by which juvenile golden pompano (Trachinotus ovatus) are poisoned by ammonia exposure has not been thoroughly elucidated. although the mechanisms of ammonia toxicity are not well described for the pompano, many other studies presented these effects to other fish species. So an overview would be given. First, an acute ammonia nitrogen toxicity experiment on juvenile golden pompano obtained a 96-h half-lethal concentration (96 h LC50) of 26.9 mg/L. In the ammonia exposure experiment, fish were sampled at 0 h, 6 h, 12 h, 24 h, 48 h, 72 h and 96 h after exposure to ammonia water (26.93 mg/L). The results showed that with the prolonged ammonia nitrogen exposure, plasma cortisol (COR), total cholesterol (TC), glutamic-pyruvic transaminase (ALT), glutamic oxalacetic transaminase (AST) and malonaldehyde (MDA) levels continued to rise, while glucose (GLU) levels first increased and later gradually decreased after 12 h. The activity of superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPX) in the liver and the mRNA expression levels of antioxidant genes (SOD, CAT, and GPX) first increased and subsequently decreased with increasing exposure time. Through microscopic observation, it was found that the degree of liver damage increased with increasing stress time and was most serious at 96 h. In the post-poison recovery experiment, the fish exposed to ammonia were transferred to clean water, and samples were taken at 24 h, 48 h, 72 h and 96 h after recovery. The results showed that with the increasing recovery time, each index recovered to the initial level to varying degrees, but the recovery time of 96 h was not enough for the fish to return to the normal level. We also examined the regulation of the Nrf2-Keap1 signaling pathway by the molecular mechanism of the antioxidant defense system. The results of this analysis showed that there was a positive correlation between Nrf2 and liver antioxidant gene expression levels, while there was a negative correlation between Keap1 and liver antioxidant gene expression levels, which may be observed because Nrf2 plays a key role in inducing antioxidant genes, and Keap1 may hinder the response to Nrf2. These results may provide a deeper and more comprehensive understanding of the impact of ammonia exposure on fish and help to provide a foundation for managing the healthy reproduction of juvenile fish.

Golden pompano (Trachinotus ovatus) is one of the most economically critical marine fishes in South China. Low oxygen stress, which is caused by a variety of factors, has resulted in substantial economic losses to the aquaculture of golden pompano. Genome-wide association study (GWAS) has been widely used to study the genetic basis of important economic traits in aquatic animals. In the present study, 100 individuals were used for genome-wide association study of hypoxia tolerance trait based on genome-wide resequencing technology to explore the SNPs and related candidate genes affecting hypoxia tolerance trait in golden pompano. Four significant SNPs were identified to be suggestively significantly associated with the hypoxia tolerance trait. In the 50 kb window surrounding the suggestive SNPs, 16 potential candidate genes affecting hypoxia tolerance trait were detected. Moreover, validation results of the screened significant loci in the large population were consistent with the results of the GWAS. Among these results, the survival times of individuals with different genotypes at SNP24101852 and SNP24194184 were significantly different (p

The marine species usually show high dispersal capabilities accompanied by high levels of gene flow. On the other hand, many physical barriers distribute along the continental marginal seas and may prevent dispersals and increase population divergence. These complexities along the continental margin generate serious challenges to population genetic studies of marine species. Chinese sea bass Lateolabrax maculatus distributes broad latitudinal gradient spanning from the tropical to the mid-temperate zones in the continental margin seas of the Northwest Pacific Ocean. Using the double digest restriction-site-associated DNA tag sequencing (ddRAD) approach, we genotyped 10,297 SNPs for 219 Chinese seabass individuals of 12 populations along the Chinese coast in the Northwest Pacific region. Genetic divergence among these populations was evaluated, and population structure was established. The results suggested that geographically distant populations in the Bohai Gulf and the Beibu Gulf retain significant genetic divergence, which are connected by a series of intermediate populations in between. The results also suggested that Leizhou Peninsula, Hainan Island, and Shandong Peninsula are major physical barriers and substantially block gene flow and genetic admixture of L. maculatus. We also investigated the potential genetic basis of local adaptation correlating with population differentiation of L. maculatus. The sea surface temperature is a significantly differentiated environmental factor for the distribution of L. maculatus. The correlation of water temperature and genetic variations in extensively distributed populations was investigated with Bayesian-based approaches. The candidate genes underlying the local selection in geographically divergent populations were identified and annotated, providing clues to understand the potential mechanisms of adaptive evolution. Overall, our genome scale population genetic analysis provided insight into population divergence and local adaptation of Chinese sea bass in the continental marginal seas along Chinese coast.

Myostatin (MSTN) was a conserved negative regulatory protein of muscle growth and development in numerous species. In this study, the MSTN gene was cloned and characterized from a noble scallop, Chlamys nobilis, and it was named CN-MSTN. To investigate the molecular characterization of MSTN and its gene expression profile of spatio-temporal, we isolated the MSTN cDNA sequence in C. nobilis and analysed expression patterns using quantitative real-time PCR. CN-MSTN cDNA contained a 1,374 bp open reading frame that encoded a 458 amino acids. Furthermore, the result of 3D model indicated that CN-MSTN mature peptide was similar to that of Chlamys farreri. Additionally, based on both the nucleotide and amino acid datasets, phylogenetic trees indicated that C. nobilis has the highest similarity with homologues of C. farreri. The expression patterns of different stages showed that MSTN expression was markedly higher in the gastrulae period. Additionally, remarkable expression of MSTN was observed in adductor muscle than with other tissues. Our data would provide expression and phylogenetic information of this economical important sea food, noble scallop.

In order to improve our understanding of the characterization of Cheilinus fasciatus mitogenome and the classification status in Labridae, the full-length mtDNA of C. fasciatus was sequenced, which...

In fish, interferon (IFN) regulatory factor 2 (IRF2) is a regulator of the type I IFN-dependent immune response, thereby playing a crucial role in innate immunity. However, the specific mechanism by which IRF2 regulates type II IFN in fish remains unclear. In the present study, first, to analyse the potential role of golden pompano (Trachinotus ovatus) IRF2 (ToIRF2) in the immune response, the mRNA level of ToIRF2 was detected by quantitative real-time polymerase chain reaction (qRT-PCR) after parasite infection. ToIRF2 was upregulated at early time points in both local infection sites (skin and gill) and system immune tissues (liver, spleen, and head-kidney) after stimulation with Cryptocaryon irritans. Second, to investigate the modulation effect of ToIRF2 on type II IFN (interferon gamma, IFNγ) expression, a promoter analysis was performed using progressive deletion mutations of ToIFNγ. The expression level of IFNγ-5 was highest among the five truncated mutants in response to ToIRF2, indicating that the core promoter region was located from −189 bp to +120 bp, which included the IRF2 binding sites. Mutation analyses showed that the activity of the ToIFNγ promoter dramatically decreased after the targeted mutation of the M1, M2 or M3 binding sites. Additionally, electrophoretic mobile shift assay (EMSA) confirmed that IRF2 interacted with the M1 binding site in the ToIFNγ promoter region to dominate ToIFNγ expression. Finally, overexpressing ToIRF2 in vitro notably increased ToIFNγ and the transcription of several type II IFN/IRF-based signalling pathway genes. These results suggested that ToIRF2 might be involved in the host defence against C. irritans infection and contribute to a better understanding of the transcriptional mechanisms by which ToIRF2 regulates type II IFN in fish.

Next‐generation sequencing has greatly promoted the investigation of single nucleotide polymorphisms, while studies of simple sequence repeats are sharply decreasing. However, simple sequence repeats still present some advantages in conservation genetics. In this study, an end‐to‐end pipeline referred to as MultiplexSSR was established to develop multiplex PCR assays in batches with highly polymorphic simple sequence repeats for capillary platforms from resequencing data. The distribution of single sequence repeats in the genome, the error profiles of genotypes and allelotypes, and the increase in the allele length range depending on the number of individuals were investigated. A total of 98% of single sequence repeats presented lengths of less than 100 bp. The error rate of the genotyping and allelotyping of dimeric patterns was ten times higher than those for other patterns. The error rate of allelotyping was less than that of genotyping. The allele length range reached approximate saturation with 10 individuals. This pipeline uses allele numbers to select highly polymorphic loci, masks loci with variation, and applies in silico PCR to improve primer specificity. The application of the developed multiplex SSR‐PCR assays validated the pipeline's robustness, showing higher polymorphism and stability for the developed simple sequence repeats and a lower cost for genotyping and providing low‐depth resequencing data from less than a dozen individuals for the development of markers. This pipeline fills the gap between next‐generation sequencing and multiplex SSR‐PCR., We established a pipeline referred to as MultiplexSSR to develop mutiplex SSR‐PCR from population resequencing data. The application of developed multiplex SSR‐PCRs validated this pipeline's robustness, showing higher polymorphism and stability for the developed simple sequence repeats and a lower cost for genotyping and providing low‐depth resequencing data from less than a dozen individuals for the development of markers.

The interferon regulatory factor 5 (IRF5) is a mediator of the type I IFN signalling pathways, thereby playing a key role in innate immunity. However, the detailed mechanism through which IRF5 regulates type I IFN in fish remains unclearly. In the present study, we first describe the identification of IRF5 (ToIRF5) from golden pompano (Trachinotus ovatus) and its features at the genomic sequence and expression level. The genomic DNA sequence consists of eight exons and seven introns. The full-length ToIRF5 cDNA is composed of 2, 059 bp and encodes for 499 amino acid polypeptides. The putative protein sequence shares 66.3%-82.9% identity to fish IRF5 and possesses three representative conserved domains (a DNA-binding domain (DBD) at the N-terminus, an IRF-associated domain (IAD), and a virus-activated domain (VAD) at the C-terminus) and one highly variable domain (middle region (MR)). Furthermore, the ToIRF5 transcript is constitutively expressed in all examined tissues, with higher levels observed in the immune relevant tissues. The mRNA levels of ToIRF5 are increased by polyinosinic: polycytidylic acid [poly (I: C)], lipopolysaccharide (LPS) and flagellin stimulation in the immune- and nonimmune-related tissues. The subcellular localization indicates that ToIRF5 is mainly localized in the cytoplasm with or without poly (I: C) induction. In addition, to explore whether ToIRF5 is a modulator of ToIFNa3, promoter analysis is performed. The region from -200 bp to +1 bp is identified as the core promoter by different truncated mutants of ToIFNa3. Mutation analyse declares that the activity of the ToIFNa3-5 promoter significantly decreases after targeted mutation of M2 binding sites. Moreover, overexpression of ToIRF5 in vitro memorably aggrandizes the expression of some IFN/IRF-based signalling pathway genes. These results provide new insights into the roles of teleost IRF5 in transcriptional mechanisms of type I IFN in the immunity process.

Interferon regulatory factor 8 (IRF8) increases type I IFN transcription levels by binding to IFN promoters, thereby playing a role in innate immunity. Nevertheless, the detailed mechanism through which IRF8 regulates type II IFN in fish remains ambiguous. In the present study, two genes from the golden pompano (Trachinotus ovatus), IRF8 (ToIRF8) and IFN gamma (ToIFNγ), were identified in the IFN/IRF-based signalling pathway. The full-length ToIRF8 cDNA was composed of 2,141 bp and encoded a 421 amino acid polypeptide; the genomic DNA was 2,917 bp in length and consisted of 8 exons and 7 introns. The putative protein showed the highest sequence identity (90-92%) with fish IRF8 and possessed a DNA-binding domain (DBD), an IRF-association domain (IAD) and a nuclear localization signal (NLS) motif consistent with those of IRF8 in other vertebrates. Furthermore, the ToIRF8 transcripts were expressed in all examined tissues of healthy fish, with higher levels observed in the central nervous and immune relevant tissues. They were upregulated by polyinosinic acid: polycytidylic acid [poly (I: C)], lipopolysaccharide (LPS) and flagellin treatments in the blood, liver, intestine and kidney. The results from assays of subcellular localization showed that ToIRF8 was localized to the cytoplasm. Moreover, to investigate whether ToIRF8 was a regulator of ToIFNγ, a promoter analysis was performed using progressive deletion mutations of ToIFNγ. The results indicated that the region from -601 bp to -468 bp includes the core promoter. Mutation analyses indicated that the activity of the ToIFNγ promoter significantly decreased after the targeted mutation of the M1-M3 binding sites. Additionally, overexpressed ToIRF8 in vitro notably increased the expression of several IFN/IRF-based signalling pathway genes. These results suggest that IRF8 is vital in the defence of T. ovatus against bacterial infection and contributes to a better understanding of the transcriptional mechanisms of ToIRF8 on type II IFN in fish.

Bone morphogenetic proteins (BMPs) play important roles in various physiological processes, especially during the formation and maintenance of various organs. In this study, we first obtained and characterized twenty BMP genes from the Trachinotus ovatus genome (designated as ToBMPs). Sequence alignment and phylogenetic analysis both indicated that the predicted amino acid sequences of ToBMP were highly conserved with corresponding homologs of other species. Moreover, a comparative analysis was performed with seven representative vertebrate genomes and found difference in number of BMP3 genes in different species, which three members, BMP3a, BMP3b-1, and BMP3b-2, existed in diploid T. ovatus, but there were four and two members in tetraploidized Cyprinus carpio (BMP3a-1, BMP3a-2, BMP3b-1, and BMP3b-2) and diploid Danio rerio (BMP3a and BMP3b), respectively. The amino acid alignment and genomic structure analysis of ToBMP3 also suggested that the BMP3 gene had expanded in T. ovatus. Furthermore, tissue expression patterns were assessed for the small intestine, liver, white muscle, brain, spleen, fin, gill, head kidney, stomach, blood, and gonads. It was discovered that BMP1, BMP2, BMP3a, BMP4, BMP6, BMP7b, BMP11, and BMP16 were ubiquitously expressed in all the tissues tested. To study the regulatory function of BMP in response to the intake of different types of food, the expression changes in BMP mRNAs were detected by qRT-PCR, and the results showed that the majority of the BMP genes had the highest mRNA levels in the small intestine and liver after ingesting pelleted feed. Our data provide a useful resource for further studies on how paralogous genes may have different expression profiles in T. ovatus.

Similar to mammals, fish possess interferon (IFN) regulatory factor 2 (IRF2)-dependent type I IFN responses. Nevertheless, the detailed mechanism through which IRF2 regulates type I IFNa3 remains largely unknown. In the present study, we first identified two genes from golden pompano (Trachinotus ovatus), IRF2 (ToIRF2) and IFNa3 (ToIFNa3), in the IFN/IRF-based signalling pathway. The open reading frame (ORF) sequence of ToIRF2 encoded 335 amino acids possessing four typical characteristic domains, including a conserved DNA-binding domain (DBD), an interferon association domain 2 (IAD2), a transcriptional activation domain (TAD), and a transcriptional repression domain (TRD). Furthermore, transcripts of ToIRF2 were significantly upregulated after stimulation by polyinosinic: polycytidylic acid [poly (I:C)], lipopolysaccharide (LPS) and flagellin in immune-related tissues (blood, liver, and head-kidney). Moreover, to investigate whether ToIRF2 was a regulator of ToIFNa3, promoter analysis was performed. The results showed that the region from -896 bp to -200 bp is defined as the core promoter using progressive deletion mutations of IFNa3. Additionally, ToIRF2 overexpression led to a clear time-dependent enhancement of ToIFNa3 promoter expression in HEK293T cells. Mutation analyses indicated that the activity of the ToIFNa3 promoter significantly decreased after targeted mutation of M4/5 binding sites. Electrophoretic mobile shift assays (EMSAs) verified that IRF2 interacted with the binding site of the ToIFNa3 promoter region to regulate ToIFNa3 transcription. Last, the promoter activity of ToIFNa3-2 was more responsive to treatment with poly (I:C) than LPS and flagellin. Furthermore, overexpression of ToIRF2 in vitro obviously increased the expression of several IFN/IRF-based signalling pathway genes after poly (I:C) abduction. In conclusion, the present study provides the first evidence of the positive regulation of ToIFNa3 transcription by ToIRF2 and contributes to a better understanding of the transcriptional mechanisms of ToIRF2 in fish.

Toll-like receptors (TLRs)are pattern recognition receptors (PRRs) that are important in invertebrate innate immunity for the recognition and elimination of pathogens. Although they were reported in many fishes, Toll-like receptors subfamily contain a large number of members with different functions that need to research in deep. In the present study, the full-length cDNA of TLR3 from the golden pompano, Trachinotus ovatus, was cloned and characterized. The full length of ToTLR3 cDNA was 3710 bp including an open reading frame of 2760 bp encoding a peptide of 919 amino acids. The derived amino acids sequence comprised of 14 leucine-rich repeats (LRR), capped with LRRCT followed by transmembrane domain and cytoplasmic Toll/IL-1R domain (TIR). Multiple sequence alignment and phylogenetic analysis revealed that ToTLR3 shared the highest similarity to the teleost fish and suggested ToTLR3 is fairly conservative in evolution process. Tissues distribution analysis indicated that ToTLR3 showed a tissue-specific variation with high expression in blood and liver. After the fish were stimulated by poly(I:C), flagellin and LPS, ToTLR3 expression in the liver, intestine, blood, kidney, skin and muscle was significantly upregulated in a time-depended manner, especially in immune related tissues such as liver, blood and kidney. Binding assay revealed the specificity of rToTLR3 for pathogen-associated molecular patterns (PAMPs) and bacteria that included Vibrio harveyi, V. vulnificus, V. anguillarum, Photobacterium damselae, Escherichia coli, Aeromonas hydrophila, Staphylococcus aureus and PolyI:C, LPS, Flagellin, and PGN. In addition, a luciferase reporter assay showed that overexpression ToTLR3 significantly increased NF-κB activity. Collectively, our results suggested that ToTLR3 might play an important role as a pattern recognition receptor (PRR) in the immune response towards pathogen infections, and transmiss the danger signal to downstream signaling pathways.

Taurine serves an important role in many species, including fish. The purpose of this study was to investigate the effects of varying dietary taurine levels on the growth, antioxidant capacity, and tissue deposition of taurine in golden pompano. In addition, the mechanism of taurine synthesis was studied via enzyme activity and gene expression of the following enzymes: cystathionine β-synthase (CBS), cystathionine γ-lyase (CSE), cysteine dioxygenase (CDO), cysteine sulfinic acid decarboxylase (CSAD), and 2-aminoethanethiol dioxygenase (ADO). Taurine concentrations of 1.3, 4.4, 7.4, 10.5 and 12.7 g kg−1 were added to the basic feed, hereafter labelled the T0, T1, T2, T3 and T4 diets, respectively. A control diet was supplemented with 1.3 g kg−1 taurine. Following 8 weeks of treatment, the results showed that the specific growth rate increased with the increase in dietary taurine content; the growth rate peaked at the 10.5 g kg−1 dietary taurine level (P 0.05). The gene expression of CBS, CSE, CDO, CSAD and ADO exhibited a downward trend with an increase of taurine (P

The elongases of very long-chain fatty acids (Elovls) are involved in the rate-limiting of the carbon chain elongation reaction in fatty acid (FA) biosynthesis in vertebrates. One member of the Elovls family, Elovl4, has been regarded as a critical enzyme involved in the biosynthesis pathway of polyunsaturated fatty acids (PUFAs). To explore the role of Elovl4 in PUFA synthesis in Trachinotus ovatus, the cDNA of the Elovl4b gene is cloned from T. ovatus (ToElovl4b). The ORF of ToElovl4b was 918 bp and encoded 305 amino acid (aa) protein sequences. Sequence alignment showed that the deduced amino acids contained significant structural features of the Elovl4 family, such as a histidine box motif (HXXHH), multiple transmembrane domains and an endoplasmic reticulum (ER) retention signal. Moreover, phylogenetic analysis revealed that ToElovl4b was highly conserved with that of Rachycentron canadum Elovl4b. Moreover, heterologous expression in yeast demonstrated that ToElovl4b could efficiently elongate 18:2n-6, 18:3n-6 and 20:5n-3 FAs up to 20:2n-6, 20:3n-6 and 22:5n-3, respectively. Furthermore, the tissue expression profile indicated that mRNA expression of ToElovl4b was higher in the gonads and brain than in other tissues. Additionally, nutritional regulation suggested the highest mRNA levels of ToElovl4b in liver and brain were under feeding with 1:1 FO-SO (fish oil, FO; soybean oil, SO) and 1:1 FO-CO (corn oil, CO)), respectively. These new insights were useful for understanding the molecular basis and regulation of LC-PUFA biosynthesis in fish.