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Zinc finger Asp-His-His-Cys motif-containing (zDHHC) proteins, known for their palmitoyltransferase (PAT) activity, play crucial roles in diverse cellular processes, including immune regulation. However, their non-palmitoyltransferase immunomodulatory functions and involvement in teleost immune responses remain underexplored. In this study, we systematically characterized the zDHHC family in the large yellow croaker (Larimichthys crocea), identifying 22 members. Phylogenetic analysis unveiled that each of the 22 LczDHHCs formed distinct clusters with their orthologues from other teleost species. Furthermore, all LczDHHCs exhibited a highly conserved DHHC domain, as confirmed by tertiary structure prediction. Notably, LczDHHC23 exhibited the most pronounced upregulation following Pseudomonas plecoglossicida (P. plecoglossicida) infection of macrophage/monocyte cells (MO/MΦ). Silencing LczDHHC23 led to heightened pro-inflammatory cytokine expression and diminished anti-inflammatory cytokine levels in MO/MΦ during infection, indicating its anti-inflammatory role. Functionally, LczDHHC23 facilitated M2- type macrophage polarization, as evidenced by a significant skewing of MO/MΦ towards the pro-inflammatory M1 phenotype upon LczDHHC23 knockdown, along with the inhibition of MO/MΦ necroptosis induced by P. plecoglossicida infection. These findings highlight the non-PAT immunomodulatory function of LczDHHC23 in teleost immune regulation, broadening our understanding of zDHHC proteins in host-pathogen interactions, suggesting LczDHHC23 as a potential therapeutic target for immune modulation in aquatic species. [ABSTRACT FROM AUTHOR]

BACKGROUND: Large yellow croaker (Larimichthys crocea) is an economical marine fish consumed in China. Theaflavins have antibacterial and antioxidant properties. However, there is a lack of research into their application in large yellow croakers during refrigerated storage. This study investigated the effect of theaflavins on the quality of large yellow croaker (Larimichthys crocea) during 12 days of storage at 4 °C. RESULTS: The results showed that theaflavin treatment was able to inhibit microbial growth and reduce the production of total volatile basic nitrogen (TVB‐N). Meanwhile, theaflavins were beneficial in reducing the unfolding of myofibrillar proteins, decreasing the degree of protein aggregation, and improving the stability of protein structure. The degree of protein oxidation was lower in a theaflavin‐treated group compared with an untreated group. Theaflavin treatment effectively inhibited increases in acid value (AV), peroxide value (PV), and malonaldehyde (MDA) content. The effect of theaflavin was positively correlated with an increase in concentration under refrigeration conditions. This study therefore suggests that the use of theaflavins is a viable method for extending the period for which refrigerated large yellow croaker can be preserved. CONCLUSIONS: Adding theaflavins to large yellow croaker can be an effective method for preserving quality during refrigerated storage. © 2023 Society of Chemical Industry. [ABSTRACT FROM AUTHOR]

The large yellow croaker (Larimichthys crocea) is an economically important marine species with the highest annual production among the farmed marine fishes in China. However, the aquaculture industry of this species is suffering from severe problems that include weakened disease resistance, decreased growth rate, and reduced meat quality due to frequent inbreeding. Genome editing, which has a huge potential for solving those problems by introducing favorable genetic changes, is not yet available for the large yellow croaker. Here, we pioneered the techniques of embryo microinjection and genome editing using the CRISPR/Cas9 system in this species. Recombinant plasmids encoding green fluorescent protein (GFP) were introduced into the fertilized eggs of L. crocea by microinjection before the chorion had hardened. A high survival rate (40%) and GFP-positive larvae rate (81.8%) were achieved, indicating that the microinjection technique in L. crocea was successfully established. On this basis, Cas9 mRNA and sgRNA targeting the tyrosinase a gene in L. crocea (Lc-tyra) were coinjected into fertilized eggs of L. crocea. Mutant individuals with insertion and deletion mutations of Lc-tyra were detected. These results indicated that the CRISPR/Cas9-based genome editing technology established herein could efficiently introduce mutations at a specific site in the L. crocea genome. This method provides the potential for genetic improvement and functional genomic study in this species. This is the first report on effective CRISPR/Cas9-based genome editing in L. crocea. [ABSTRACT FROM AUTHOR]

Ultrasonic treatment (UT) was used to thaw large yellow croaker in this study, and the effect of various ultrasonic power levels on the quality of large yellow croaker was evaluated after thawing. The effects of ultrasonic on water holding capacity (WHC), moisture distribution, thiobarbituric acid‐reactive substance (TBARs), total volatile base nitrogen (TVB‐N), ATP degradation (related to K value), surface color change, free amino acid (FAA) content, total sulfhydryl group (SH) content, Fourier transform infrared absorption spectra (FT‐IR), fluorescence emission spectra, and microscopic observations of large yellow croaker myofibrillar proteins were investigated. The thawing times of the control sample, 200UT, 240UT, 280UT, and 320UT samples were 1750, 1190, 810, 580, and 570 s, respectively, which indicated that ultrasonic radiation could improve thawing efficiency. Additionally, ultrasonic thawing maintained better freshness and color and inhibited lipid oxidation. Compared with fresh samples, the TVB‐N of large yellow croaker thawed by ultrasonication increased by 12.68%, and the K value increased by 0.9%. The 240UT sample had tightly arranged myofibrils and fewer changes in the structures of myogenic fibrillar proteins than the fresh samples, and the SH content of 240UT was decreased by 8.17%. Use of excessive ultrasonic power (320 W) damaged the protein microstructure and the microstructure of large yellow croaker. In conclusion, sample 240UT maintained the quality of large yellow croaker better with minimal damage, which is recommended for rapid thawing. Practical Application: Ultrasonic waves improve the thawing efficiency of large yellow croaker and maintain the freshness and color of the fish. According to results, sample 240UT exhibited slight changes in the structure of the myofibril protein, but excessive ultrasonic power destroyed the microstructure and protein structure. Appropriate ultrasonic treatment to the thawing of fish has good prospects. [ABSTRACT FROM AUTHOR]

The mall heterodimer partner (SHP) plays an important regulatory role in mammal inflammation. The main objective of this study was to investigate the response of SHP to inflammatory stimulation and its underlying mechanism. The shp gene from large yellow croakers, was cloned, and this gene is mainly expressed in the liver and intestine. Lipopolysaccharide (LPS) stimulation induced the mRNA expression and protein level of SHP in macrophages of large yellow croakers. Overexpression of SHP significantly decreased mRNA expression of tnfα , il-1β , il-6 and cox2 induced by LPS treatment in macrophages. LPS stimulation increased the phosphorylation level of Adenosine 5'-monophosphate (AMP)-activated protein kinase (AMPK) in macrophages. AMPK inhibitor treatment significantly decreased the expression of SHP induced by LPS while AMPK activator significantly increased the expression of SHP. The nuclear factor-erythroid 2-related factor 2 (NRF2) increased the promoter activity of SHP in large yellow croakers and the level of nuclear NRF2 was increased by LPS stimulation and AMPK activation. NRF2 inhibitor treatment significantly decreased mRNA expression of shp induced by LPS and AMPK activator. In conclusion, LPS can induce SHP expression by activating the AMPK-NRF2 pathway while SHP could negatively regulate LPS-induced inflammation in large yellow croakers. This study may be benefit to the development of immunology of marine fish and provide new ideas for inflammation-related diseases. [ABSTRACT FROM AUTHOR]

Summary: Acid‐soluble collagens (ASCs) were extracted and characterised from different tissues of large yellow croaker (Larimichthys crocea). The yields of ASCs in bones, skins, scales and muscles were 3.22 ± 0.47%, 42.30 ± 1.15%, 2.82 ± 0.31% and 0.89 ± 0.12%, respectively. SDS‐PAGE and ATR‐FTIR analysis showed that all ASCs were type I collagen with intact triple helical structure and consisted of α1,3‐chain, α2‐chain, β‐chain and γ‐chain. Quantitative analysis of SDS‐PAGE revealed that the ratio of β‐chains in ASC from scales was lower than other tissues. The imino acid contents of ASC from bones, skins, scales and muscles were 169, 167, 162 and 173 residues/1000 residues, respectively. The maximum transition temperature (Tm) of ASC from scales was 30.15 °C, lower than the other three ASCs, indicated that the thermal stability of collagen was affected by both the content of imino acid and β‐chains. The results also implied that thermal stability of ASC might exist the tissue specificity among different fish species. [ABSTRACT FROM AUTHOR]

High levels of soybean oil (SO) in fish diets enriched with linoleic acid (LA, 18:2n-6) could induce strong inflammation. However, the molecular mechanism underlying LA-induced inflammation in the liver of large yellow croaker (Larimichthys crocea) has not been elucidated. Based on previous research, autophagy has been considered a new pathway to relieve inflammation. Therefore, the present study was performed to investigate the role of autophagy in regulating LA-induced inflammation in the liver of large yellow croaker in vivo and in vitro. The results of the present study showed that activation of autophagy in liver or hepatocytes could significantly reduce the gene expression of proinflammatory factors, such as tumor necrosis factor α (TNFα) and interleukin 1β (IL1β). The results of the present study also showed that inhibition of autophagy could upregulate the gene expression of proinflammatory factors and downregulate the gene expression of anti-inflammatory factors in vivo and in vitro. Furthermore, autophagy could alleviate LA-induced inflammatory cytokine gene expression in vivo and in vitro , while inhibition of autophagy obtained the opposite results. In conclusion, our study shows that autophagy could regulate inflammation and alleviate LA-induced inflammation in the liver of large yellow croaker in vivo and in vitro for the first time, which may offer considerable benefits to the aquaculture industry and human health. [ABSTRACT FROM AUTHOR]

Free fatty acid receptor 4 (FFAR4) plays a key role in regulating the inflammatory response in mammals. The present study aimed to investigate the function of large yellow croaker FFAR4 on inflammation. In the present study, ffar4 was widely expressed in 10 tissues of large yellow croaker including gill, head kidney and spleen. Further studies showed that treatment of head kidney macrophages with agonists (TUG891 or GSK137647A) or overexpression of ffar4 reduced the mRNA expression of pro-inflammatory genes induced by LPS, and increased the expression of pparγ. Treatment of macrophages with antagonist AH7614 increased the mRNA expression of pro-inflammatory genes induced by LPS, and decreased the mRNA expression of pparγ. In order to verify the immunomodulatory effect of PPARγ, PPARγ was overexpressed in macrophages which significantly reduced the mRNA expression of pro-inflammatory genes il6 , il1β , il8 , tnfα and cox2. Moreover, results of dual-luciferase assays showed that PPARγ downregulated the transcriptional activity of il6 and il1β promoters. In conclusion, FFAR4 showed anti-inflammatory effects on LPS-induced inflammation in large yellow croaker. [ABSTRACT FROM AUTHOR]

High-density single-nucleotide polymorphism (SNP) genotyping array is an essential tool for genetic analyses of animals and plants. Large yellow croaker (Larimichthys crocea) is one of the most commercially important marine fish species in China. Although plenty of SNPs have been identified in large yellow croaker, no high-throughput genotyping array is available. In this study, a high-throughput SNP array named NingXin-I with 600K SNPs was developed and evaluated. A set of 82 large yellow croakers were collected from different locations of China and re-sequenced. A total of 9.34M SNPs were identified by mapping sequence reads to the large yellow croaker reference genome. About 1.98M candidate SNPs were selected for further analyses by using criteria such as SNP quality score and conversion performance in the final array. Finally, 579.5K SNPs evenly distributed across the large yellow croaker genome with an average spacing of 1.19 kb were proceeded to array production. The performance of NingXin-I array was evaluated in 96 large yellow croaker individuals from five populations, and 83.38% SNPs on the array were polymorphic sites. A further test of the NingXin-I array in five closely related species in Sciaenidae identified 26.68–56.23% polymorphic SNP rate across species. A phylogenetic tree inferred by using the genotype data generated by NingXin-I confirmed the phylogenetic distance of the species in Sciaenidae. The performance of NingXin-I in large yellow croaker and the other species in Sciaenidae suggested high accuracy and broad application. The NingXin-I array should be valuable for quantitative genetic studies, such as genome-wide association studies (GWASs), high-density linkage map construction, haplotype analysis, and genome-based selection.

A proteomics analysis was conducted to investigate the effects of dietary carotenoids on the skin coloration of large yellow croaker (Larimichthys crocea) fed diets with different carotenoids for 9 weeks. The diet without carotenoids supplementation was used as the control group (CD). Based on the CD, the other four experimental diets were supplemented with 80 mg/kg of astaxanthin (AST80), 80 mg/kg of xanthophyll (X80), 40 mg/kg of astaxanthin and 40 mg/kg of xanthophyll (AST40 + X40), and 80 mg/kg of astaxanthin and 80 mg/kg of xanthophyll (AST80 + X80), respectively. Results showed that the growth performance, feed utilization, pH, water‐holding capacity, and texture in the muscle of large yellow croaker were not significantly affected. No significant differences were observed in redness (a*) of dorsal skin among all the groups. Nevertheless, the redness values of ventral skin in groups with dietary carotenoids supplementation were higher than that in the control group. The yellowness (b*) of dorsal and ventral skin was significantly enhanced in dietary carotenoid‐supplemented groups compared with CD group except for dorsal skin in the AST80 group. After astaxanthin or xanthophyll administration, the carotenoid contents in dorsal and ventral skin were significantly increased. After the proteomics analysis of the skin, 444 differentially expressed proteins (DEPs) and 352 DEPs have been found between AST80 and CD, and X80 and CD, respectively. The proteomics results showed that dietary carotenoids influenced the expression of myosin heavy chain, ribosomal proteins, metal iron binding related proteins, oxygen transporter activity associated proteins, lipid transport proteins and immune‐related proteins. The skin coloration of large yellow croaker might be directly regulated by myosin proteins and ribosomal proteins, such as 40S ribosomal protein S21 (RPS21), 60S ribosomal protein L23a (RPL23a), RPL24 and RPL27, which were induced by astaxanthin and xanthophyll. [ABSTRACT FROM AUTHOR]

The p53 activation is induced by stressors, such as DNA damage, oxidative stress, and activated oncogenes, and can promote cell cycle arrest, cellular senescence, and apoptosis. The large yellow croaker (Larimichthys crocea) is an important warm temperate marine fish in the Chinese aquiculture industry. However, few studies have investigated the role of p53 in the response of L. crocea to environmental stressors. Therefore, the aim of the present study was to assess the spatiotemporal mRNA expression levels of genes involved in the p53 signaling pathway of the large yellow croaker in response to cold stress. The results showed significant changes in the expression levels of p53, p21, MDM2, IGF-1, Gadd45, Fas, and Akt in various tissues of the large yellow croaker in response to cold stress for different times. As compared to the control group, p53 mRNA expression was upregulated in most of the examined tissues at 24 h with the exception of the gill. In the liver, the expression levels of p53 and Fas were significantly decreased at 12 h, while those of p21, MDM2, IGF-1, Gadd45 were dramatically increased. Akt expression was notably changed in response to cold in several tissues. These results suggested that p53 was potentially a key gene in the large yellow croaker response to cold and possibly other environmental stressors. [ABSTRACT FROM AUTHOR]

Visceral white nodules disease (VWND) caused by Pseudomonas plecoglossicida is a common disease in cage‐farmed large yellow croaker (Larimichthys crocea) in China. VWND usually occurred at water temperature of 16–19℃, resulting in high mortality in farmed large yellow croaker. Now, P. plecoglossicida as its pathogen has been considered nonpathogenic at 7–12℃. During February 2019, an infectious disease outbreak was observed in cage‐farmed large yellow croaker at a water temperature of 12℃ in Ningde, China. This disease is characterized by white granulomatous lesions in internal organs of the diseased fish, which was similar with the symptoms of the VWND in large yellow croaker. Then, we isolated a bacterial strain named PQLYC4 from visceral lesions of the diseased fish. The experimental infection studies demonstrated that the strain PQLYC4 was the pathogen of the disease, which was further identified as P. plecoglossicida by the analysis of morphology, 16s rRNA gene homology and average nucleotide identity based on the whole genome sequence. Our results revealed that P. plecoglossicida strain PQLYC4 could cause the outbreak of the VWND at 12℃, a water temperature lower than that reported previously, thus providing new knowledges of prevalence and prevention of the VWND in large yellow croaker. [ABSTRACT FROM AUTHOR]

High-density single-nucleotide polymorphism (SNP) genotyping array is an essential tool for genetic analyses of animals and plants. Large yellow croaker (Larimichthys crocea) is one of the most commercially important marine fish species in China. Although plenty of SNPs have been identified in large yellow croaker, no high-throughput genotyping array is available. In this study, a high-throughput SNP array named NingXin-I with 600K SNPs was developed and evaluated. A set of 82 large yellow croakers were collected from different locations of China and re-sequenced. A total of 9.34M SNPs were identified by mapping sequence reads to the large yellow croaker reference genome. About 1.98M candidate SNPs were selected for further analyses by using criteria such as SNP quality score and conversion performance in the final array. Finally, 579.5K SNPs evenly distributed across the large yellow croaker genome with an average spacing of 1.19 kb were proceeded to array production. The performance of NingXin-I array was evaluated in 96 large yellow croaker individuals from five populations, and 83.38% SNPs on the array were polymorphic sites. A further test of the NingXin-I array in five closely related species in Sciaenidae identified 26.68–56.23% polymorphic SNP rate across species. A phylogenetic tree inferred by using the genotype data generated by NingXin-I confirmed the phylogenetic distance of the species in Sciaenidae. The performance of NingXin-I in large yellow croaker and the other species in Sciaenidae suggested high accuracy and broad application. The NingXin-I array should be valuable for quantitative genetic studies, such as genome-wide association studies (GWASs), high-density linkage map construction, haplotype analysis, and genome-based selection. [ABSTRACT FROM AUTHOR]

To evaluate the quality changes in large yellow croaker (Pseudosciaena crocea) with ε‐polylysine and rosemary extract stored on ice at 4 ± 1°C. About 0.1% ε‐polylysine (PL) and 0.2% rosemary extract (RE) were individually or in combination with each other were treated with samples. Samples treated with deionized water were regarded as control check (CK) group. Physicochemical (texture profile analysis (TPA), pH, total volatile basis nitrogen (TVB‐N), thiobarbituric acid (TBA)), endogenous enzyme activity (cathepsin B and D), proteolytic degradation (Trichloroacetic Acid (TCA)‐soluble peptides and Sodium Salt‐polyacrylamide gel electrophoresis (SDS‐PAGE)), microbiological (Total viable count (TVC), Shewanella bacteria count, Pseudomonas bacteria count, Psychrophilic bacteria count) and sensory evaluation were conducted during the whole storage. As a result, PL + RE could delay the increase in pH, TVB‐N, TBA value, and improve the texture attributes compared with the CK group. In addition, PL + RE could inhibit cathepsin B and D activities, protein degradation, and microbial growth effectively. Moreover, the shelf life of samples could be prolonged at least 4 days when compared with the CK group according to the quality index method (QIM) and physicochemical assay, indicating that the PL + RE treatment could maintain the quality of large yellow croaker more effectively. Practical applications: The preservation of fish is becoming increasingly important in aquatic products. According to the fence theory, a combination of biopreservatives with different functions could be used to maintain the freshness synergistically. Furthermore, this research indicates that the combination of ε‐polylysine and rosemary extract, a promising method for the preservation of aquatic products, could slow down the deterioration of large yellow croaker and prolong its shelf life. [ABSTRACT FROM AUTHOR]

Fish tend to rely more on their innate immunity to executing defense against viral infection by inducing antiviral gene production. However, the expression pattern and underlying mechanism of fish antiviral responses have yet to be fully defined. In the present study, an in vitro viral infection model was established by exposing head kidney-derived macrophages of large yellow croaker to virus analog, poly(I:C). Transcriptome analysis indicated that poly(I:C) appeared to induce potent antiviral activity featuring dominant interferon a3 (IFNa3) expression through activation of toll-like receptors (TLRs)/TIR-domain-containing adapter-inducing interferon-β (TRIF) and retinoic acid-inducible gene I-like receptors (RLRs)/mitochondrial antiviral signaling protein (MAVS) pathways. Inhibition of nuclear factor κB (NF-κB) and stimulator of interferon genes (STING)/interferon regulatory factor 3 (IRF3) pathways diminished the expression of IFNa3. Mechanistically, transcription factors including p65 and IRF3 could promote expression of IRF3 , and activated IRF3 alone further increased the transcriptional activity of IFNa3. We also characterized the promoter of IFNa3 with direct IRF3 binding site which was sufficient to render the transcription of IFNa3. This effect was attenuated after deletion or mutation of the IRF3 binding sites. Taken together, our findings illustrate the distinct transcriptional profiling of fish macrophages triggered by poly(I:C). Also, this work provides new insights into the molecular mechanism underpinning coordinated activation of pathogen recognition and signaling transduction in the antiviral responses of non-model fish species. • Combined transcriptome and in vitro analyses reveal the antiviral mechanism of fish. • Poly(I:C) is potent to trigger a rapid induction of antiviral genes in fish macrophages. • Expression of antiviral genes primarily depends on NF-κB and IRFs signaling. • IRF3 alone could induce the transcription of itself and IFNa3. • IRF3 binding to specific promoter region is a prerequisite for IFNa3 expression. [ABSTRACT FROM AUTHOR]

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A 65‐day feeding trial was conducted to investigate the effects of dietary antimicrobial peptide APSH‐07 on the growth performance, anti‐oxidation responses, stress resistance and intestine microbiota of large yellow croaker Larimichthys crocea. Four isonitrogenous and isolipidic diets were formulated with 0, 30, 60 and 90 mg/kg of APSH‐07, respectively. The results showed that the fish fed with 90 mg/kg of dietary APSH‐07 had the significantly highest specific growth rate (p <.05). The activities of catalase, superoxide dismutase, the total anti‐oxidative capacity and lysozyme had the significantly highest values in liver of fish fed with 90 mg/kg of dietary APSH‐07 (p <.05). Serum glucose increased significantly in fish fed diets without APSH‐07 supplementation after temperature stress, while serum cortisol increased significantly in those after trawl stress (p <.05). Fish fed with 90 mg/kg of dietary APSH‐07 showed significantly higher operational taxonomic units, ACE estimator and phylogenetic diversity whole tree in intestine microbiota compared to fish fed without APSH‐07 (p <.05). In conclusion, under the present experimental condition, 90 mg/kg of antimicrobial peptide APSH‐07 supplementation in diet had the better growth performance, higher anti‐oxidation and stress resistance capacity, and a potentially more beneficial intestine microbial community of large yellow croaker. [ABSTRACT FROM AUTHOR]

Interleukin (IL)-11 is a multifunctional cytokine that exerts a series of important immunomodulatory effects and exists in many tissues and cells. A 1106-bp nucleotide sequence representing the complete cDNA of IL-11 was obtained from large yellow croaker (Larimichthys crocea), containing an open reading frame (ORF) of 603 bp encoding for 200 amino acids (aa). The predicted Lc IL-11 protein included a 12aa signal peptide and a conserved IL-11 domain. The polypeptide sequence identities between Lc IL-11 and its counterparts in mammals and other fish are from 84% to 92% with known fish IL-11a and 22%–27% with fish IL-11b. Lc IL-11 mRNA existed in most tissues with the most predominant expression in the gill. After immune challenge, the expression levels of Lc IL-11 were induced largely in vivo and in vitro , with the peak-value of 32 times as much as the control in the liver at 24 h after Vibrio parahaemolyticus injection (p < 0.05) and the greatest value of 13.9 times as much as the control in LCK cells at 12 h after poly I:C stimulation (p < 0.05). Furthermore, the overexpression vector pcDNA3.1- Lc IL-11 was constructed and transfected to LCK cells. Our results showed that the transcriptional expression levels of tumor necrosis factor (TNF)-α and myxovirus resistant protein (Mx) significantly up-regulated in LCK cells after Lc IL-11 overexpression (p < 0.05). However, no significant changes of IL-1β, janus kinase (JAK)2 and signal transducers and activators of transcription (STAT)5 was detected. Our finding indicated that Lc IL-11 might enhance TNF-α and antiviral protein Mx expression in large yellow croaker. • A 1106-bp nucleotide sequence representing the complete cDNA of IL-11 was obtained from large yellow croaker. • LcIL-11 shared 84%–92% with known fish IL-11a and 22%–27% with fish IL-11b. • LcIL-11 mRNA existed in most tissues with the most predominant expression in the gill. • LcIL-11 could be induced by bacterial and viral challenge. • Overexpression LcIL-11 could triggered the expression of TNF-α and Mx significantly. [ABSTRACT FROM AUTHOR]

Large yellow croaker (Larimichthys crocea, LYC) aquaculture is being threatened by intensive infectious diseases. Relevant studies have focused on LYC immune responses to infection. By contrast, little is known how and to what extent the gut microbiota responds to infection. Here, we explored the interactions between LYC immune responses and gut bacterial communities during Pseudomonas plecoglossicida infection. P. plecoglossicida successfully colonized into LYC gut microbiota, resulting in an increasing mortality rate. Relative gene expressions of pro-inflammatory cytokines (TNF-α1, TNF-α2 and IL-1β) and anti-inflammatory cytokine (IL-10) were consistently and significantly induced by P. plecoglossicida infection, whereas non-specific immune enzymes activities were only enhanced at the early infection stages. P. plecoglossicida infection caused an irreversible disruption in the gut microbiota, of which infection and hours post infection constrained 16.2% and 5.6% variations, respectively. In addition, top 18 discriminatory taxa that were responsible for the difference between treatments were identified, whose abundances were significantly associated with the immune activities of LYC. Using a structural equation modeling (SEM), we found that gut bacterial communities were primarily governed by the conjointly direct (−0.33) and indirect (0) effects of infection, which subsequently affect host immune responses. Our results suggest that an irreversible dysbiosis in gut microbiota could be the causality of increasing mortality. To our knowledge, this is the first study to provide an integrated overview among pathogen infection, immune response and gut microbiota of LYC. • The successful colonization of P. plecoglossicida is confirmed by sequencing. • P. plecoglossicida infection caused an irreversible disruption of gut microbiota, resulting in increasing mortality. • Abundances of the top 18 discriminatory taxa were significantly associated with the immune activities of LYC. • Interrelationship among pathogen infection, immune response and host gut microbiota was established. [ABSTRACT FROM AUTHOR]

Apart from mitigating endoplasmic reticulum (ER) stress, vast studies have demonstrated the crucial role of inositol-requiring transmembrane kinase and endonuclease 1α (IRE1α) - spliced X-box binding protein 1 (XBP1s) signaling pathway in inflammatory response in mammals. In addition, palmitic acid (PA)-induced inflammation has been verified in large yellow croaker (Larimichthys crocea). However, whether the IRE1α-XBP1s signaling pathway is involved in inflammatory response caused by PA remains poorly studied in fish. The present study was aimed at elucidating the role of the IRE1α-XBP1s signaling pathway in inflammatory response induced by PA in primary hepatocytes from large yellow croaker. In the present study, the full-length cDNA of ire1α and xbp1s were cloned and comprised 3793 bp and 1789 bp with an open reading frame of 3279 bp and 1170 bp, encoding 1093 and 390 amino acids, respectively. IRE1α protein possessed a protein kinase and endoribonuclease domain and XBP1s protein possessed a basic-leucine zipper domain. The IRE1α protein and XBP1s protein located to the ER membrane and nucleus respectively. The ire1α and xbp1s were widely transcribed in various tissues with the higher level in intestine, liver, adipose and head kidney. The ER stress-inducing agent tunicamycin (Tm) and PA treatment significantly activated the IRE1α-XBP1s signaling pathway and increased the pro-inflammatory genes expression including tumor necrosis factor α (tnfα), interleukin 6 (il-6) and interleukin 1β (il-1β) (P < 0.05). When KIRA6, the IRE1α kinase inhibitor, was used to block the IRE1α-XBP1s signaling pathway, the Tm and PA-induced pro-inflammatory genes expression was significantly suppressed (P < 0.05). These data indicated that the IRE1α-XBP1s signaling pathway was involved in the PA-induced inflammatory response in large yellow croaker. • IRE1α and XBP1s were cloned and characterized from the liver of large yellow croaker. • IRE1α-GFP fusion protein located to the ER membrane and XBP1s-GFP fusion protein located to the nucleus of HEK293t. • The IRE1α-XBP1s signaling pathway was involved in the PA-induced inflammatory response in large yellow croaker. [ABSTRACT FROM AUTHOR]

As a member of tumor necrosis factor receptor (TNFR)-associated factor (TRAF) family, TRAF3 is an important regulator of NF-κB and type I interferon (IFN) activation, especially in Toll-like receptors (TLRs)- and retinoic acid inducible gene I (RIG-I)-like receptors (RLRs)-mediated signaling pathway. In the present study, a TRAF3 homologue named Lc -TRAF3 was characterized in large yellow croaker (Larimichthys crocea). The open reading frame (ORF) of Lc -TRAF3 contains 1788 bp encoding a protein of 595 amino acids (aa). Sequence analysis indicated that Lc -TRAF3 is conserved in vertebrates, constituted with a N-terminal RING finger, two TRAF-type zinc fingers, and a C-terminal TRAF-MATH domain. The genome organization of Lc -TRAF3 is conserved in fish, with 13 exons and 12 introns, but different from that in birds or mammals, which contains 10 exons and 9 introns. Lc -TRAF3 was identified as cytosolic protein base on fluorescence microscopy analysis. Expression analysis revealed that Lc -TRAF3 was broadly distributed in examined organs/tissues, with the highest expression level in gill and weakest in brain, and could be up-regulated under poly I:C, LPS, PGN, and Pseudomonas plecoglossicida stimulation in vivo. Interestingly, overexpression Lc -TRAF3 could induce the activation of NF-κB, and Lc -TRAF3 co-transfected with Lc -TRIF induced a significantly higher level of NF-κB and IRF3 promoter activity, implying that Lc -TRAF3 may function as an enhancer in Lc -TRIF-mediated signaling pathway. • A TRAF3 homologue was characterized in large yellow croaker Larimichthys crocea. • Lc -TRAF3 can be induced by viral and bacterial PAMPs stimulation. • Lc -TRAF3 can induce NF-κB activation. • Lc -TRAF3 can positively regulate TRIF-mediated NF-κB and IRF3 activation. [ABSTRACT FROM AUTHOR]

Bacterial diversity in the foregut, midgut and hindgut of large yellow croaker Larimichthys crocea and black sea bream Sparus macrocephalus reared in an inshore net pen, formulated feed and seawater were analyzed with Illumina MiSeq high-throughput sequencing platforms. A total of 270 operational taxonomic units (OTUs) were identified from fish guts, formulated feed and seawater, which belonged to 17 phyla and 20 genera. Firmicutes and Proteobacteria dominated at the phylum level, while Bacillus, Lactococcus and Oceanobacillus dominated at the genus level, in fish gut. The similarity in bacterial community between the guts of two fish species was higher than that either between fish gut and formulated feed or between fish gut and seawater. This result indicates that bacterial communities in guts of large yellow croaker and black sea bream were independent on those in the formulated feed and seawater. The common OTUs between fish gut and formulated feed were more than that between fish gut and seawater, suggesting that the impact of the formulated feed on intestinal bacteria of large yellow croaker and black sea bream was greater relative to that of seawater in the net pen. Spatial heterogeneity in bacterial composition along the gut of large yellow croaker and black sea bream was also evaluated. [ABSTRACT FROM AUTHOR]

Cultured large yellow croaker (Larimichthys crocea) is frequently infected by bacteria, thus developing small white granulomas in the spleen and kidneys with high mortality in a few days. We performed a genome-wide association study (GWAS) on the trait of resistance to Pseudomonas plecoglossicida infection in a large yellow croaker population in 222 extreme-phenotype samplings using the whole-genome resequencing technology. With the 254,110 detected SNPs for the trait, three candidate regions associated with disease resistance were found on chromosomes 3, 5 and 20. By combing our GWAS results with the biological functional analysis of immunity genes, we found that IL10, THBS1, IGSF21, IGR and IGH as well as IRF8, TRAIL, CC, TNF and LINGO1 are candidate genes of the disease resistance trait in the large yellow croaker. Moreover, after comparison with transcriptome sequencing data, we found that the candidate genes from GWAS were actually up-regulated or down-regulated. This is the first time that a GWAS combined with a transcriptome-wide study was performed on disease resistance to detect candidate loci and find potential functionally relevant genes in the large yellow croaker. [ABSTRACT FROM AUTHOR]

The present study was conducted to investigate the effects of dietary lipid, carbohydrate and their interactions on growth performance, feed utilization, body composition and non‐specific immunity of large yellow croaker (Larimichthys crocea). The experimental fish were fed the diets with graded levels of dietary crude lipid (80, 110 and 140 g/kg diet) and dietary carbohydrate (60, 90 and 120 g/kg diet) for 7 weeks. Results showed that the specific growth rate decreased significantly as dietary carbohydrate increased, which partly resulted from the progressively reduced feed intake and protein efficiency ratio. Meanwhile, the result of postprandial glucose content confirmed glucose intolerance of the croaker. However, the specific growth rate was not significantly altered by dietary lipid, but the feed intake was remarkably reduced. The different growth performance and feed utilization of croaker in response to dietary lipid and carbohydrate resulted in the variation of body composition. In addition, the activity of lysozyme and classical complement pathway was significantly improved in croaker fed the diet with 110 g/kg lipid. In above, dietary carbohydrate and lipid of large yellow croaker should be maintained at 60 and 110 g/kg diet, respectively, based on the above indices. [ABSTRACT FROM AUTHOR]

As an adaptor in Toll-like receptor (TLR) signaling pathway, Toll/interleukin-1 receptor (TIR) domain containing adaptor inducing interferon-β (TRIF) mediates downstream signaling cascades and plays important roles in host innate immune responses. In the present study, a TRIF ortholog named Lc -TRIF was identified in large yellow croaker (Larimichthys crocea). Sequence comparison analysis revealed that Lc -TRIF has a conserved TIR domain but without TRAF6 binding motif. The genome structure of Lc -TRIF is conserved, with two exons and one intron. Syntenic comparison showed that the loci of fish TRIF was different from that in mammals or birds, and TRAM was absent in the genomes of fish, amphibians, and birds, but present in mammals and reptiles. Expression analysis revealed that Lc -TRIF was broadly expressed in examined organs/tissues, with the highest expression level in gill and weakest in brain, and could be up-regulated under poly I:C, LPS, PGN, and Pseudomonas plecoglossicida stimulation. Fluorescence microscopy results showed that Lc -TRIF exhibited a global localization throughout the entire cell including the nucleus in HEK 293T cells. Additionally, luciferase assays demonstrated that Lc -TRIF expression could significantly induce NF-κB, type I IFN, IRF3 as well as IRF7 promoter activation. These results collectively indicated that Lc -TRIF was function in host antiviral and antibacterial responses via NF-κB and IRF3/7 related signaling pathway. • A TRIF ortholog was characterized from large yellow croaker. • TRIF can be induced by viral and bacterial PAMPs stimulation. • TRAM was absent in bony fish. • TRIF can induce NF-κB, type I IFN, IRF3, and IRF7 activation. [ABSTRACT FROM AUTHOR]

The large yellow croaker (Larimichthys crocea) is one of the most valuable economic mariculture fish in China, and its yellowness in skin is considered as the most important traits that determine the market value. The fish skin colors present naturally silvery white during the daytime and golden yellow during the night-time. In the present study, we found that the xanthosomes of xanthophores aggregated during the daytime and dispersed during the night-time, while the melanosomes of melanophores always dispersed all day long in vivo. We further cloned three proopiomelanocortin (POMC) genes of the large yellow croaker. All these POMC genes were expressed in the pituitary, but only POMC-C mRNA was expressed in the skin. The results of single-cell RT-PCR showed that the POMC-C mRNA was expressed in isolated xanthophores but not in melanophores. The xanthophores, but not melanophores, were found in the ventral skin, where only one subtype of melanocortin receptor (MCR), i. e. MC5R, was expressed. MC1R, MC4R and MC5R mRNAs were detected in the dorsal skin, where there existed both the xanthophores and melanophores. The results of single-cell RT-PCR showed that only MC5R mRNA was expressed in the xanthophores, while only MC1R mRNA was detected in the melanophores. Thirty min after in vivo and in vitro treatment with melanocyte-stimulating hormone (MSH) peptides (α-MSH, Des-Ac-α-MSH, α-MSH-C and β-MSH-C), dispersion degrees of the xanthosomes and melanosomes significantly increased, even under light exposure. Interestingly, when we immediately observed the dorsal scales that have been maintained under in vitro complete dark condition without MSH peptides treatment for at least 30 min, the melanosomes aggregated as expected, but the xanthosomes dispersed. Both α-MSH and α-MSH-C exhibited similar, but stronger effects than Des-Ac-α-MSH on dispersion of both the xanthosomes and melanosomes. Although the two MSH peptides (α-MSH-C and β-MSH-C) are generated from POMC-C, the former showed stronger effect on xanthosome dispersion than the latter. Forskolin, an Adenylyl cyclase activator, dispersed pigment granules in both the xanthophores and melanophores. H 89 2HCl, a specific inhibitor of protein kinase A blocked α-MSH-induced pigment granule dispersion of xanthophores and melanophores. Taken together, our results indicated that the dispersion of melanosomes is stimulated by MSH, possibly secreted from pituitary, through endocrine pathway. The yellowness of the large yellow croaker skin was mainly due to the dispersion of xanthosomes. The dispersion of xanthosomes was regulated by MSH that is possibly released both from the pituitary through endocrine pathway and from the xanthophores through autocrine pathway. • The yellowness of the large yellow croaker skin was mainly due to the dispersion of xanthosomes. • Melanocyte-stimulating hormone dispersed the xanthosomes through both endocrine and autocrine pathway. • The light could specifically make the xanthosomes aggregated. • MSH activated its receptor (MC5R) in the xanthophores and further triggered AC-cAMP-PKA signal pathway. [ABSTRACT FROM AUTHOR]

Abstract The present study was conducted to investigate the metabolic responses of glucose and lipid in large yellow croaker Larimichthys crocea (initial weight, 36.80 ± 0.39 g) to high level of dietary soybean oil. Three isonitrogenous (46% crude protein) and isolipidic (13% crude lipid) experimental diets were designed, with 100% fish oil (FO), 50% fish oil and 50% soybean oil (FS) and 100% soybean oil (SO), respectively. After a 12-week growth trial, the results showed that compared with FO group, contents n-6 PUFAs increased while the n-3 PUFAs decreased significantly both in liver and muscle in FS and SO groups. Concentrations of blood glucose, leptin, free fatty acid and total triglyceride reached the highest values in SO group, while blood insulin showed no significant difference among all groups. The gene expressions of insulin receptor substrate-2, glucose-6-phosphatase, phosphoenolpyruvate carboxykinase, fatty acid synthetase, and lipoprotein lipase increased, and the insulin receptor substrate-1, phosphotidylinsositol-3-kinase (PI3K), hexokinase, glycogen synthetase and glucose transporter 2 in liver decreased significantly in SO group. Meanwhile, the phosphorylation of protein kinase B (AKT) also decreased significantly in this group. These results suggested that high level of dietary soybean oil depressed PI3K/AKT signaling pathway, and then affected glucose and lipid metabolism by glycolysis, gluconeogenesis, glucose transportation, glycogenesis and lipogenesis. Highlights • Large yellow croaker were fed with diet with high level of dietary soybean oil (6.5%). • The gene expression of the insulin receptor substrate-1 and phosphotidylinsositol-3-kinase (PI3K) were decreased. • The phosphorylation of the protein kinase B (AKT) was also decreased. • The glycolysis, gluconeogenesis, glucose transportation, glycogenesis and lipogenesis were significantly influenced. • High level of dietary soybean oil depressed PI3K/AKT signaling pathway, and then affected glucose and lipid metabolism. [ABSTRACT FROM AUTHOR]

Abstract Carnitine palmitoyltransferase 1 (CPT1) plays an essential role in maintaining energy supply via fatty acid oxidation, especially under fasting. In this study, the complete cDNA sequence of cpt1a was cloned from liver of large yellow croaker (Larimichthys crocea), with an open reading frame of 2319 bp encoding a protein of 772 amino acids. Bioinformatics analysis predicted the presence of conserved functional motifs and amino acid residues. The highest mRNA expression of cpt1a was observed in the liver. Phylogenetic tree clearly shows that CPT1A protein is a homologue of mammalian CPT1A. Recombinant protein rCPT1A showed catalytic activity, with Michaelis constant (km) (≈1.38 mM) and maximal reaction rates (Vmax) for carnitine (≈12.66 nmols/min/mg protein). The cpt1a mRNA expression dramatically increased and CPT1 activity remained unchanged after fasting. Fasting did not significantly change Vmax and free carnitine (FC) content in liver. Interestingly, catalytic efficiency (Vmax/Km) and FC/Km increased in fish fasted for 4 days, implying FC contents might be enough to ensure the optimal fatty acid oxidation. Contrarily, both indicators declined when fish fasted for 12 days. The present results demonstrated cpt1a has a biological function and showed that the transcriptional and kinetic regulation of CPT1 during fasting, emphasizing that fasting-induced fatty acid oxidation depends on changes in kinetic properties instead of CPT1 activity and transcription. Graphical abstract Unlabelled Image Highlights • A function of CPT1A was firstly demonstrated in fish by a yeast expression system. • Fasting affected CPT1A expression, Km for carnitine, and catalytic efficiency. • Short- and long-term fasting differently affected catalytic efficiency by changing Km. • Duration of fasting should be carefully considered when using it to reduce lipid content. [ABSTRACT FROM AUTHOR]

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Abstract In this study, we sequenced and characterized two homologues of interferon-stimulated gene ISG15, termed as LcISG15-1 and LcISG15-2, from the large yellow croaker (Larimichthys crocea). The LcISG15-1 encodes 159 amino acids and the LcISG15-2 encodes 155 amino acids, both of which contain two tandem ubiquitin-like domains and the conserved C-terminal LRGG conjugation motif. The sequence analyses showed that both the LcISG15-1 and LcISG15-2 exhibit high similarity with ISG15 from other fishes. A putative IFN-stimulatory response element (ISRE) was detected in promoter regions of both the LcISG15-1 and LcISG15-2. Phylogenetic analyses revealed a close evolutionary relationship of both the LcISG15-1 and LcISG15-2 with other teleostean ISG15. Molecular evolutionary analyses suggested a gene duplication event of ISG15 in the ancestor of the Sciaenidae, with a signature of positive selection was found in the ISG15-2 gene copy of sciaenid fishes. The Real-time PCR analyses showed that the LcISG15-1 and LcISG15-2 were both found to be ubiquitously expressed in ten examined organs in large yellow croaker, with predominant expressions both in peripheral blood. Expression analyses showed that both the LcISG15-1 and LcISG15-2 were rapidly and significantly upregulated in vivo after poly (I:C) challenge in liver and spleen organs. However, the LcISG15-1 and LcISG15-2 were both significantly induced after pathogen Vibrio parahemolyticus infection only in the liver but not in the spleen. These results indicated that there are two ISG15 homologues in the large yellow croaker, both of which are likely to be involved in host immune defense against viral and bacterial infection. Highlights • LcISG15-1 and LcISG15-2 were sequenced from Larimichthys crocea. • LcISG15-1 and LcISG15-2 both express ubiquitously with predominant expressions in peripheral blood. • LcISG15-1 and LcISG15-2 were significantly upregulated after poly(I:C) challenge in liver and spleen. • LcISG15-1 and LcISG15-2 were significantly upregulated after Vibrio parahemolyticus infection only in liver but not in spleen. • LcISG15-1 and LcISG15-2 both play important roles in host immune defense against viral and bacterial infection. [ABSTRACT FROM AUTHOR]

Abstract The shortage of fish oil (FO) leads to the extensive use of vegetable oil (VO) in marine fish diets. High replacement percentage of dietary FO by VO induced pro-inflammatory response of adipose tissue (AT) and liver tissue (LT) in large yellow croaker (Larimichthys crocea). Mammalian studies showed that the secretion of cytokines by AT affected the immune response of LT. To investigate whether or not the inflammation response of LT is related to AT in large yellow croaker, LT and AT cells from fish fed FO diet (FOL and FOA) and VO diet (VOL and VOA) were co-cultured in a trans -well system, which resulted in an assembly of the two cells types sharing the culture medium but being separated by the membrane of the insert. Co-culture of FOL and FOA was selected as the control group (FOL-FOA). Results indicated that, when compared with the control group, the expression of pro-inflammatory genes (toll like receptors [ TLRs ] , tumour necrosis factor α [ TNFα ], interleukin 1β [ IL1β ] , suppressor of cytokine signalling 3 [ SOCS3 ] and cyclooxygenase 2 [ COX2 ]) in FOL was significantly increased in the co-culture group of FOL and VOA (FOL-VOA), while the expression of anti-inflammatory genes (arginase I [ ArgI ] and transforming growth factor β1 [ TGFβ1 ]) in FOL was significantly depressed. On the contrary, a significantly depressed expression of pro-inflammatory genes (TLRs , TNFα , IL1β and COX2) and increased expression of anti-inflammatory genes (interleukin 10 [ IL10 ]) in VOL was observed in the co-culture group of VOL and FOA (VOL-FOA) when compared with the co-culture group of VOL and VOA (VOL-VOA). The change of immune-related gene expressions in LT cells was attributed to nuclear factor κB (NF-κB) signalling since the expression of the p65 protein was observed to show a similar trend to the expression of pro-inflammatory genes. It is speculated that dietary VO increased the secretion of cytokines, which induced pro-inflammatory response in LT cells. These ex vivo results indicate that AT plays a vital role in LT pro-inflammatory response in fish fed VO diet. Highlights • Adipose tissue cells from fish fed vegetable oil diets increased the expression of pro-inflammatory genes but decreased the expression of anti-inflammatory genes of liver tissue cells. • The change of immune-related gene expressions in liver tissue cells was attributed to NF-κB signalling. • Adipose tissue plays a vital role in liver tissue pro-inflammatory response of fish fed vegetable oil diet. [ABSTRACT FROM AUTHOR]

• Ultrasound increased free amino acids and nucleotides contents in the semi-dried fish. • Heat treatment would promote the production of volatile substances. • Na+ and K+ played an important role in flavor characteristics of semi-dried fish. • Partial substitution of KCl for NaCl could improve the flavor. • Coordination treatment was beneficial to improve the flavor. This study investigated the flavor characteristics of semi-dried yellow croaker made by KCl instead of partial NaCl combined with ultrasound treatment before and after low temperature vacuum heating. The electronic tongue, electronic nose, free amino acids, 5′-nucleotides, and gas chromatography-ion mobility spectrometry were employed. Electronic nose and electronic tongue results showed that different treatment groups had different sensitive signals to smell and taste. The odor and taste of each group were mainly affected by Na+ and K+. The difference between the groups becomes larger after thermal treatment. Ultrasound and thermal treatment both changed the content of taste components. In addition, each group contained 54 volatile flavor compounds. Among them, the combined treatment method gave semi-dried large yellow croaker pleasant flavor characteristics. Besides, it also improved the content of flavor substances. In conclusion, the semi-dried yellow croaker under sodium-reduced conditions showed better performance in flavor characteristics. [ABSTRACT FROM AUTHOR]

Abstract The aim of this study was to evaluate the preservation effect of chitosan (CS) and lysozyme (Lys) coating on the quality of large yellow croaker (Larimichthys crocea) via physicochemical analyses, microbiological and sensorial assessments during refrigerated storage. Results showed that CS films incorporated with 0.6 mg·mL−1 of lysozyme exhibited the strongest inhibition effect against S. aureus , whose inhibition zone was 16.10 ± 0.71 mm. The utilization of CS singly, or CS-lysozyme (Lys) coating significantly reduced lipid oxidation (thiobarbituric acid value, TBA) and improved the sensorial scores of fishes. Control and treated groups exceeded the maximum permissible level (30 mg N/100 g) of total volatile basic nitrogen (TVB-N) in fishery products on days 6 (control) and 9 (CS or CS-Lys treated), respectively. Fish samples reached the value of 7.0 Log CFU/g (Total Viable Count, TVC) on days 15 (control), whereas both CS and CS-Lys treated flesh samples never exceed this limit value during the refrigerated storage. Results of our study indicate CS-Lys have high efficiency in inhibiting microorganism growth and lipid oxidation and in maintaining sensorial quality, which is a promising method to maintain the quality and extend the shelf life of refrigerated fishes. Highlights • CS-Lys film was prepared and characterized. • CS-Lys film showed the improved efficiency against four bacteria compared to CS film. • CS or CS-Lys film could improve the quality of fish muscle. [ABSTRACT FROM AUTHOR]

Mammalian interleukin-4 (IL-4) and −13 (IL-13), two anti-inflammatory T helper cell type 2 (Th2) cytokines, play the central roles in mediating the alternative activation of monocytes/macrophages (MO/Mφs). However, exact functions in MO/Mφs polarization of IL-4/13 homologues in teleost fish remain largely unknown. In this study, we identified two IL-4/13 homologues from large yellow croaker Larimichthys crocea , Lc IL-4/13A and Lc IL-4/13B, which share low amino acid sequence identities to the known fish IL-4/13 molecules. Phylogenetic analysis showed that Lc IL-4/13A is evolutionarily closely related to Dicentrarchus labrax IL-4/13A, and Lc IL-4/13B to Takifugu rubripes IL-4/13B. The two Lc IL-4/13 genes were constitutively expressed in all examined tissues, but with different expression levels. Both Lc IL-4/13A and Lc IL-4/13B were up-regulated by inactivated trivalent bacterial vaccine in the head kidney, and Lc IL-4/13B appeared more responsive to bacterial vaccine than Lc IL-4/13A. Recombinant Lc IL-4/13A and Lc IL-4/13B proteins (rLcIL-4/13A and rLcIL-4/13B) produced in Escherichia coli could significantly decrease production of reactive oxygen species (ROS) and nitrogen oxide (NO) in the head kidney MO/Mφs from large yellow croaker. Furthermore, rLcIL-4/13A and rLcIL-4/13B obviously down-regulated expression of pro-inflammatory cytokine (IL-1β and TNF-α) and inducible NO synthase (iNOS) genes in MO/Mφs, while they increased mRNA expression of anti-inflammatory cytokines (TGF-β and VEGF) and arginase-2. Additionally, the phagocytic activity of MO/Mφs was also inhibited by rLcIL-4/13A or rLcIL-4/13B. All these results therefore indicated that both Lc IL-4/13A and Lc IL-4/13B, although exhibiting a lower degree of sequence identity of 15.6% and differential expression pattern, have the similar roles in promoting alternative activation of head kidney MO/Mφs. [ABSTRACT FROM AUTHOR]

Complement is a complex innate immune surveillance system, playing a key role in host homeostasis, inflammation, and in the defense against pathogens. Complement regulators are crucial to prevent the injudicious production of these mediators and potential injury to self tissues. Here, we identified the complement factor H (CFH) and its related gene 2 (CFHR2) homologs from large yellow croaker ( Larimichthys crocea ), named LcCfh and LcCfhr2 , respectively. The deduced LcCfh and LcCfhr2 proteins shared significant structural similarities and identified codes for a polypeptide consisting of various numbers of highly conserved SCR domains. LcCfh , LcCfhr1 and LcCfhr2 genes were detected in all examined tissues with predominantly expressions in liver, spleen and kidney, and their expressions all increased upon Vibrio alginolyticus challenge. In vitro assays showed that recombinant LcCfh was likely to act as a cofactor of CFI and played a negative regulation role in complement system, when recombinant LcCfhr2 seemed to play mechanisms independent of the activity of CFH. Both recombinant LcCfh and LcCfhr2 took participate in inflammatory reaction despite of the inequal ability to mediate pro-inflammation response. These data provide a new insight into the functional activities of teleost complement system. [ABSTRACT FROM AUTHOR]

Increasing demand, uncertain availability and high price of fish oil with the expansion of aquaculture made it essential to search alternative lipid sources. Vegetable oil has been proved to be the best candidate for the replacement of fish oil in aquafeeds. However, this replacement especially in high level potentially has some negative effects on fish. The present study was conducted to investigate the growth performance, anti-oxidative and inflammatory responses of large yellow croaker to replacement of dietary fish oil by soybean oil. Three isonitrogenous (46% crude protein) and isolipidic (13% crude lipids) diets were formulated to feed fish (initial body weight: 36.80 ± 0.39 g) for 12 weeks. The control diet was designed to contain 6.5% of fish oil, and named as FO. On the basis of the control diet, the fish oil was 50% and 100% replaced by soybean oil, and these two diets were named as FS and SO, respectively. Results showed that the specific growth rate significantly decreased in the SO group. Crude lipid contents in muscle and liver of fish fed SO diet were significantly higher than those in the FO group. The ratio of n-3 poly-unsaturated fatty acids (PUFAs) to identified fatty acids in liver decreased significantly, while n-6 PUFAs increased significantly with increasing dietary soybean oil inclusion. The levels of triacylglycerol, non-esterified fatty acid and tumour necrosis factor α, and the activity of aspartate aminotransferase in serum significantly increased in SO group. The total anti-oxidative capacity and expressions of the anti-oxidation-related genes (superoxide dismutase 1 and 2, catalase, glutathion peroxidase and nuclear factor erythroid 2-related factor 2) were significantly decreased by dietary soybean inclusion. Dietary soybean oil significantly decreased the gene expressions of the anti-inflammatory cytokines (arginase I and interleukin 10), and increased the pro-inflammatory cytokines (tumour necrosis factor α and interleukin 1β). The replacement of dietary fish oil by soybean oil also induced an over-expression of toll-like receptor 22 and myeloid differentiation factor 88 in liver. In conclusion, dietary soybean oil could suppress growth performance and liver anti-oxidative capacity, and induce inflammatory responses of large yellow croaker. [ABSTRACT FROM AUTHOR]

Abstract: The study was conducted to investigate the effects of dietary tea polyphenols (TP) on growth performance, biochemical and antioxidants responses, fatty acid composition, and lipid metabolism‐related gene expressions of large yellow croaker (Larimichthys crocea). Four diets were formulated with different levels of TP (0.00%, 0.01%, 0.02% and 0.05%). Results showed that growth performance of L. crocea were not different among dietary treatments. Compared with the control group, fish in 0.02% TP group had lower body and hepatic lipid content and lower total cholesterol content. The minimum content of triglycerides and low‐density lipoprotein‐cholesterol were found in 0.05% TP group. Hepatic n‐6 PUFA and n‐3 PUFA were significantly higher in TP supplementation groups. Malondialdehyde content was lower in TP supplementation groups, and superoxide dismutase activity was higher in 0.01% TP group than the control group. The mRNA expressions of carnitine palmitoyltransferase1, acyl‐CoA oxidase and peroxisome proliferators‐activated receptor α were up‐regulated in 0.01% and 0.02% TP groups, while lipoprotein lipase expression was down‐regulated in TP supplementation groups than the control group. Results suggested that 0.01%–0.02% TP supplementation could reduce the deposition of liver lipid of L. crocea caused by high‐lipid diet, which might be due to the increase in lipid oxidation related gene expressions. [ABSTRACT FROM AUTHOR]

In this study, we sequenced and characterized an interferon-stimulated gene Viperin homologue, LcViperin, from large yellow croaker ( Larimichthys crocea ). The LcViperin encodes 354 amino acids and contains an N-terminal amphipathic α-helix domain, a radical S-adenosyl- l -methionine (SAM) domain and a highly conserved C-terminal domain. The analyses of LcViperin promoter region revealed nine kinds of putative transcriptional factor binding sites, including five putative ICSBP (IRF-8) binding sites and one putative IRF-1 binding site, indicating that the expression of LcViperin might be induced by the type I IFN response. Phylogenetic analyses based on amino acid sequences showed that the Viperin of large yellow croaker is clustered together with its counterparts from other teleost fishes. The Real-time PCR analyses showed that the LcViperin was found to be ubiquitously expressed in ten examined tissues in large yellow croaker, with predominant expression in peripheral blood, followed by heart and gill. Expression analyses showed that the LcViperin was rapidly and significantly upregulated in vivo after poly (I:C) challenge in peripheral blood, head kidney, spleen and liver tissues. The results indicate that the LcViperin might play a pivotal role in antiviral immune responses. [ABSTRACT FROM AUTHOR]

Large yellow croaker ( Pseudosciaena crocea ) is a cultivated fish of great economic importance and abundant nutritional value. However, due to its high protein and water contents, it is susceptible to decomposition, leading to considerable economic loss and adverse effects on consumer health. Here, we assessed the function of the bacterial strain Bacillus coagulans L1208 (Bcoa) in preserving large yellow croaker during storage at 4 °C and found that Bcoa elongates the shelf-life significantly. Further investigations showed that Bcoa prolongs the storage time mainly by suppressing the growth of spoilage bacteria. Moreover, a novel bacteriocin, designated as Coagulin L1208 and produced by Bcoa, was purified and identified by N-terminal sequencing. Finally, the activity of Coagulin L1208 for suppressing spoilage bacteria during the preservation of large yellow croaker was assessed. Our results reveal the mechanism by which Bcoa aids the preservation of large yellow croaker and identify Coagulin L1208 as a potential novel antiseptic. [ABSTRACT FROM AUTHOR]

The large yellow croaker ( Larimichthys crocea ) has a well-developed innate immune system. We studied a component of this system, chemokine receptor CXCR family. In this study, we report the full-length open reading frames, as well as the identification and characterization of the chemokine receptor genes CXCR2 (Lyc CXCR2 ), CXCR3 (Lyc CXCR3 ), and CXCR4 (Lyc CXCR4 ) of large yellow croaker. We report that Lyc CXCR3 and Lyc CXCR4 are evolving neutrally according to PAML analyses. Quantitative real-time PCR analysis revealed that CXCR transcripts were expressed in all examined tissues. The expression of chemokine receptors Lyc CXCR2 , Lyc CXCR3 , and Lyc CXCR4 was elevated in the kidney, spleen, and particularly the liver of the large yellow croaker after challenge with Vibrio anguillarum and polyinosinic:polycytidylic acid (poly I:C). These results suggest that Lyc CXCR2 , Lyc CXCR3 , and Lyc CXCR4 may be important immune-related genes, playing crucial roles in immune defence against bacterial infection. [ABSTRACT FROM AUTHOR]

Understanding sex determination mechanisms is the first and most paramount step in the development of sex control breeding biotechnologies for fish. The large yellow croaker ( Larimichthys crocea ) is an economically vital fish in China and East Asia, which exhibits a sex-related growth dimorphism where females grow significantly faster than males. However, the lack of a rapid and accurate sex specific marker for the large yellow croaker has hampered the application of sex control in the breeding process, as well as sex-determination investigations of this species. In this study, a male-specific 15 bp deletion in the third intron of the Dmrt1 gene was identified by comparative analysis of genomic re-sequencing data from male and female samples. Two pairs of primers (MS and MFS) were carefully designed; thus, the male-specific marker could be genotyped by PCR amplification and agarose gel electrophoresis. More importantly, this male-specific marker could also be applied to precisely identify the genetic sex of multiple fish. This male specific DNA marker will have potential applications in the elucidation of the sex determination mechanism and will also accelerate the development of sex control breeding in the large yellow croaker. Moreover, our current data further validated an significant genetic evidence that an XY-male determination mechanism is utilized in the large yellow croaker. [ABSTRACT FROM AUTHOR]

The article focuses on the impact of low dietary fish meal (FM) content on volatile oils in muscle of Larimichthys crocea (LYC). Topics discussed include analysis of volatile indicators through statistically interpreted by principle component analysis (PCA), feeding with control diet, and presence of higher content of total esters.

To investigate the effect of dietary lipid levels on lipid metabolism in muscle and adipose tissue of large yellow croaker ( Larimichthys crocea), fish (average body weight 150.0 g) were fed to one of three diets with a low (6%), moderate (12%, the control diet) or high (18%) crude lipid level for 10 weeks. The expression of genes related to lipoprotein clearance ( lpl, ldlr, lrp1 and srbi), fatty acid transport ( cd36, fatp1, fabp3 and fabp11), lipogenesis ( fas and dgat2) and lipolysis ( atgl, cpt1 and aco) in muscle and adipose tissue was analysed. In muscle of fish fed the low-lipid diet, the expression of lpl, lrp1, fas, dgat2 and atgl was significantly downregulated, whereas fabp3 and fabp11 were significantly upregulated compared with those fed the control diet. In muscle of fish fed the high-lipid diet, the expression of lpl, ldlr, lrp1, cd36, fabp3, fas and dgat2 was significantly decreased, while fabp11 expression was significantly increased compared with those fed the control diet. Compared with the control group, only a significant downregulation of fabp3 expression in adipose tissue was observed in the low-lipid group. In the high-lipid group compared with the control group, the expression of ldlr, fabp11, dgat2, atgl and aco in adipose tissue was significantly upregulated, whereas cpt1 expression in adipose tissue was significantly downregulated. These results indicated that the expression of the selected key genes related to lipid metabolism was tissue-specific regulation by dietary lipid levels in large yellow croaker. [ABSTRACT FROM AUTHOR]

Large yellow croaker is an important marine aquaculture species in China. The aim was to determine an appropriate protocol of artificial fertilization for family construction in the breeding programme based on two trials. In trial 1, luteinizing hormone-releasing hormone A (LHRHA) was injected once, with a dosage of 2 μg/kg for females and 1 μg/kg for males. The latency time was in the range of 30-34 h. The maturation stage was checked by extracting a few eggs with a Pasteur pipette. The fertilization rate and hatching rate were 27 and 52%, respectively. The percentage of females with spawning difficulties was 30%. In trial 2, the females were injected LHRHA twice: with a first dose of 0.8 μg/kg and a second dose of 2 μg/kg, at an interval of 10 h, whereas the males were still injected once. The latency time was in the range of 29.5-35 h, determined by only observing courtship behaviour of males. The females with spawning difficulties decreased to 10%, and the fertilization rate and hatching rate also improved to 41 and 62%, respectively. [ABSTRACT FROM AUTHOR]

Thermal tolerance to high temperature was evaluated in the large yellow croaker Larimichthys crocea. The survival thermal maximum for L. crocea was 33.0°C, the 50% critical thermal maximum (50% CTMax) was 35.5°C, and the critical thermal maximum (CTMax) was 36.0°C. Three microsatellite markers (LYC0148, LYC0200 and LYC0435), associated with thermal tolerance were screened and identified using a Bulked Segregation Analysis (BSA) method. These markers have six amplified fragments in which four are related to thermal tolerance. These fragments were cloned and sequenced, and the results showed the core motif were all 'AC' repeats. For LYC0148 and LYC0200, the lengths of fragments are 181 bp and 197 bp, respectively. For LYC0435, which has two fragments, the fragment lengths are 112 bp and 100 bp. The results provide useful molecular markers for thermal-tolerance breeding of large yellow croaker in the near future. [ABSTRACT FROM AUTHOR]

A 9-week feeding experiment was conducted to determine the effect of dietary biotin levels on growth performance and non-specific immune response of large yellow croaker. Fish (6.16 ± 0.09 g) were fed twice daily to apparent satiation with diets containing 0.00 (as the basal diet), 0.01, 0.05, 0.25, 1.24 and 6.22 mg biotin kg−1 diet. Results showed that fish fed the basal diet had the lowest survival rate, and fish fed 0.05 mg kg−1 dietary biotin achieved significantly higher final weight and weight gain. Dietary biotin levels had no significant influence on carcass crude lipid, moisture and ash content, but significantly influenced the carcass crude protein. Liver biotin concentration significantly increased with the supplementation of biotin, but no tissue saturation was found within the supplementation scope of biotin. Broken-line regression analysis of weight gain showed that juvenile large yellow croaker requires a minimum dietary biotin of 0.039 mg kg−1 for maximal growth. The analyses of serum parameters showed that the moderate- (0.05 mg kg−1) and high-dose (6.22 mg kg−1) dietary biotin significantly improved both lysozyme and alternative complement pathway activities, indicating dietary biotin within a certain range could improve the non-specific immune response of large yellow croaker. [ABSTRACT FROM AUTHOR]

The unfolded protein response (UPR) is a mechanism to cope with perturbed endoplasmic reticulum (ER) functions or accumulation of unfolded protein in the ER in eukaryotic cells. Furthermore, the UPR also participates in a number of physiological and pathological processes, such as nutrient sensing, lipid synthesis, and inflammatory response. In this study, four UPR-related genes ( GRP78/BiP , ATF6α , XBP1 and CHO ) were isolated characterized from large yellow croaker ( Larimichthys crocea ), and their expression in response to dietary lipid sources (various fatty acids) such as fish oil (FO), palmic acid (PA), olive oil (OO), sunflower oil (SO), and perilla oil (PO), were examined following feeding. The results showed that the four UPR-related proteins contained highly conserved functional domains and had the closest phylogenetic relationships with other fishes. Additionally, these genes were ubiquitously expressed in large yellow croaker, as in zebrafish and medaka. Moreover, GRP78 , ATF6α and spliced XBP1 ( XBP1s ) mRNA levels in the liver, not in adipose tissue, were significantly increased in the SO group compared to the other groups ( P < 0.05). These results indicated that dietary SO activated UPR, and the activation of UPR might provide a mechanism to improve ER function, but probably stimulated lipid synthesis and caused inflammatory response in the liver of large yellow croaker. [ABSTRACT FROM AUTHOR]

• The integration of ecosystem model and management strategy evaluation for fisheries management. • An evaluation of effects of fisheries management procedures on ecosystem dynamics. • An assessment of ecological carrying capacity of Larimichthys crocea. Management Strategy Evaluation (MSE) evaluates the performance of alternative management strategies. However, the potential ecosystem effects of the management procedures (MPs) are less evaluated. As one of the four top commercial marine fisheries of China, the fishery of the large yellow croaker Larimichthys crocea (LYC) collapsed in the late 1980s due to the exceeded fishing pressure on spawning and over-wintering aggregations. Long-term and large scale restocking programs for the recovery of LYC fishery have been conducted widely along the eastern coast of China, but its ecosystem impacts and ecological carrying capacity (ECC) were not well understood and assessed. The species-specific assessment and management for LYC fishery were lack due to limited data availability. In this study, an Ecopath model was built as the operating model in MSE to evaluate the effects of MPs for the LYC fishery and estimate the LYC ECC in the Min River Estuary to explore the optimal management measure. A total of 12 candidate MPs were identified and tested, including 5 input controls (2 fishing-effort-based and 3 length-based) and 7 catch-based output controls. A 20-year simulation showed that only 4 MPs fully achieved the performance criteria in maintaining biomass and avoiding further overfishing, including curE75 (75 % of current fishing effort levels), minlenLopt1 (the minimum length of retention for maximizing biomass), matlenLim (fishing retention-at-length related to the maturity curve: 100 %) and matlenLim2 (fishing retention-at-length related to the maturity curve: 110 %). The Min River Estuary ecosystem could support up to 17 kg/km2 of LYC. The Ecosim model projects that the LYC biomass increased over 20 % after the application of curE75, and over 37 % with the combination of curE75 and national fishing moratorium policy after the 20-year simulation. The combination of releasing, fishing effort reduction and fishing moratorium is the optimal measure within current management systems for the recovery of LYC fishery in the Min River Estuary. This study provides an ecosystem-based fishery management approach to plan and evaluate restocking programs using data-limited methods and MSE for the LYC and other fisheries. [ABSTRACT FROM AUTHOR]

The physiological responses to different environmental salinities were assessed in juveniles of large yellow croaker Pseudosciaena crocea. This species shows a good capacity to adapt to comparatively low environmental salinities by evaluating some physiological responses, i.e. superoxide dismutase ( SOD), acid phosphatase ( ACP), alkaline phosphatase ( AKP) and lysozyme ( LZM) in the liver, spleen, gill and kidney respectively. Growth and survival at salinity 5‰, 10‰ and 25‰ were better than those at salinity 15‰ and 20‰. No significant differences in ACP, AKP, SOD and LZM in the liver were observed among different salinity treatments; SOD, AKP and LZM in the spleen among different treatments only showed significant differences at the beginning or the 2nd week; in the gill, no significant difference of AKP and LZM were observed during the whole experiment, SOD among different treatments showed significant difference at the beginning and the 8th week, and ACP only showed significant difference at the end of the experiment; in the kidney, significant differences in ACP, AKP and SOD among different salinities were merely observed at the end of experiment, and LZM showed significant difference among different treatments at the 2nd week. Overall, some slight stress responses were observed, but few significant differences were observed between low salinity and normal salinity, especially the growth and physiological functions were not influenced by low salinity, i.e. 5‰ and 10‰. We conclude that juvenile large yellow croaker is tolerant to low salinity and shows a potential for low salinity culture. [ABSTRACT FROM AUTHOR]

In this study, we sequenced and characterized melanoma differentiation-associated antigen 5 (LcMDA5), laboratory of genetics and physiology 2 (LcLGP2) and mitochondrial antiviral signaling protein (LcMAVS) from large yellow croaker ( Larimichthys crocea ). The LcMDA5 encodes 969 amino acids and contains two caspase-associated and recruitment domains (CARDs), a DExDc (DExD/H box-containing domain), a HELICc (helicase superfamily C-terminal domain) and a C-terminal regulatory domain (RD). The LcLGP2 encodes 679 amino acids and contains a DExDc, a HELICc and a RD. The LcMAVS encodes 512 amino acids and contains a CARD, a proline-rich domain, a transmembrane helix domain and a putative TRAF2-binding motif ( 269 PVQDT 273 ). Phylogenetic analyses showed that all the three genes of large yellow croaker are clustered together with their counterparts from other teleost fishes. The Real-time PCR analyses showed that all the three genes were found to be constitutively expressed in all examined tissues in large yellow croaker, but all with relatively low expression levels. Expression analyses showed that the three genes were all rapidly and significantly upregulated in vivo after poly (I:C) challenge in peripheral blood, liver, spleen and head kidney tissues. The results indicate that the LcMDA5, LcLGP2 and LcMAVS might play important roles in antiviral immune responses. [ABSTRACT FROM AUTHOR]