Large Yellow Croaker Larimichthys Crocea Research Articles

Dried porcine soluble augments dietary fishmeal replacement by poultry by-product meal for large yellow croaker Larimichthys crocea

Two concurrent feeding trials was conducted to assess the effects of replacing fishmeal with poultry by-product meal (PBM) and supplementing dried porcine soluble (DPS) in large yellow croaker Larimichthys crocea diets at the optimal dietary protein level. In trial I, the reference diet (F40) contained fishmeal at 400 g/kg, and 40%, 60% and 80% of the fishmeal was substituted with PBM (the diets were termed as F24, F16 and F8). In trial II, the control diet (F16D0) contained fishmeal at 160 g/kg, and DPS was added at 4, 8, 12 and 16 g/kg in F16D0 (the diets were termed as F16D4, F16D8, F16D12 and F16D16). Fish initially weighing 20.0 ± 0.2 g were fed the test diets for 10 weeks. Fishmeal replacement level significantly influenced final body weight (FBW), weight gain, feed conversion ratio, nitrogen and energy retention efficiencies, ratio of fish-in to fish-out (FIFO), feed cost, activity of superoxide dismutase and catalase in plasma and liver, activity of glutathione peroxidase in plasma, and malondialdehyde content in liver. However, neither fishmeal replacement level nor DPS supplementation level resulted in significant change in condition factor, hepatosomatic index and body composition. There were no significant differences in weight gain and FBW either between F40, F24 and F16 or between F40 and F16D0, F16D4, F16D8, F16D12 and F16D16. The FBW of F16 declined by 3.2% relative to F40, while the FBW of F16D12 increased by 10.1% relative to F16D0. This study reveals that dietary fishmeal level for large yellow croaker could be reduced to 160 g/kg by using PBM as a sole substitute, and DPS supplementation could augment dietary fishmeal replacement by PBM.

Dietary cottonseed protein concentrate affected the flesh texture and myofiber characteristics of large yellow croaker Larimichthys crocea

The effects of replacing dietary fishmeal with cottonseed protein concentrate (CPC) on the muscle texture and water holding capacity of large yellow croaker Larimichthys crocea were investigated after a 90-day feeding trial. Results showed that the complete substitution of dietary fishmeal with CPC resulted in a significant reduction in muscle drip loss by decreasing collagen content and increasing heat soluble collagen content, without affecting the texture. Moreover, reduced drip loss is contingent upon diminished muscle salt-soluble protein content by enhancing skeletal muscle degradation (ubiquitin-proteasome system, calpain proteases and lysosomal cathepsins) and decreasing energy levels. In conclusion, the substitution of 30% dietary fishmeal with CPC (i.e., 20.94% of CPC in diet) did not show any significant alterations in muscle quality of large yellow croaker. The complete replacement of dietary fishmeal with CPC (i.e., 69.80% of CPC in diet) reduced the water holding capacity of muscle.

Structural and ultrastructural aspects of the skin of large yellow croaker Larimichthys crocea.

The skin color of the large yellow croaker (Larimichthys crocea) is a crucial indicator to determine its economic value. However, the location of pigment cells in the skin structure is uncertain. To determine the pigment cell type in the skin, the vertical order and ultrastructure of pigment cells were examined using light microscopy and transmission electron microscopy. Both dorsal and ventral skins comprise the epidermis, dermis, and hypodermis. Xanthophores, melanophores, and iridophores were observed in the dermis of the dorsal skin, whereas the latter two were in the dermis of the ventral skin. Interestingly, the size of xanthophores in the dorsal skin was significantly smaller than that of xanthophores in the ventral skin; however, the density of dorsal xanthophores was significantly higher than that of ventral xanthophores. The type L-iridophores with large crystalline structures were observed in the uppermost area of the upper pigment layer, which contributed to the strikingly metallic luster shown by the ventral skin. The melanophores were exclusively found in the dorsal skin, offering the purpose of camouflage. Taken together, our results indicated that the pigment cells display different arrangement patterns between dorsal and ventral skin, and the golden color in the ventral skin results from the coexistence of light-reflecting iridophores and light-absorbing xanthophores.

Effects of Dietary Fish Meal Replaced by Fish Steak Meal on Growth Performance, Antioxidant Capacity, Intestinal Health and Microflora, Inflammatory Response, and Protein Metabolism of Large Yellow Croaker Larimichthys crocea.

Although fish steak meal (FSM) is a potentially available protein source, its efficiency as a fish meal (FM) substitute remains unclear to date. To this end, this study was carried out to determine the effects of dietary FM replaced by FSM on growth performance, antioxidant capacity, intestinal health and microflora, inflammatory response, and protein metabolism of large yellow croaker. Five isolipidic and isonitrogenous diets were formulated by substituting FM with FSM at levels of 0% (FSM0, control diet), 25% (FSM25), 50% (FSM50), 75% (FSM75), and 100% (FSM100), and were fed to juvenile large yellow croaker for 8 weeks. Compared with the control diet, the replacement of 25% dietary FM with FSM did not markedly alter the weight gain (WG) and specific growth rate (SGR). When the FM substitution level was over 25%, WG and SGR markedly reduced. The intestinal structure observation found that the FSM75 and FSM100 diets markedly decreased villus height, villus width, and muscle thickness of the anterior intestine. The FSM75 and FSM100 diets significantly decreased enzyme activities of amylase (AMS), lipase (LPS), trypsin, catalase (CAT), and total superoxide dismutase (T-SOD) and the total antioxidant capacity (T-AOC), and increased the malondialdehyde (MDA) content in the liver of large yellow croaker. The mRNA expression levels of intestinal barrier and inflammatory response-related genes suggested that the FSM50, FSM75, and FSM100 diets significantly decreased the mRNA abundances of intestinal barrier-related genes and anti-inflammatory response-related genes, and increased the mRNA abundances of proinflammatory gene il-6 in the anterior intestine. The compositions of intestinal microflora displayed that the FSM50, FSM75, and FSM100 diets decreased relative abundances of Firmicutes phylum and increased relative abundances of Proteobacteria phylum. In addition, the results of protein expression levels showed that the phosphorylation level of mammalian target of rapamycin (mTOR) and 4E-binding protein 1 (4E-BP1) in FSM75 and FSM100 groups were markedly reduced. In conclusion, FSM can replace up to 25% dietary FM without compromising the growth performance, intestinal health, and protein metabolism of the large yellow croaker.

P38 MAPK involvement in the thermal stress response occurs via HSP27 and caspase3 in the large yellow croaker (Larimichthys crocea)

The p38 mitogen-activated protein kinase (p38 MAPK) is a multifunctional molecule that is involved in cellular response to various stressful stimuli. In the present study, the full-length cDNA sequence of p38 MAPK (Lcp38 MAPK) was identified from the large yellow croaker Larimichthys crocea, which encoded a polypeptide of 361 amino acid residues. The predicted Lcp38 MAPK protein contained a highly conserved Thr-Gly-Tyr (TGY) motif, a glutamate and aspartate (ED) site, a substrate binding site (Ala-Thr-Arg-Trp < ATRW>), and a serine/threonine kinase catalytic (S_TKc) domain characteristic of the MAPK family. The constitutive expression of Lcp38 MAPK was detected in most of the tissues examined with the strongest expression in intestine. Subcellular localization in LCK cells (kidney cell line from a L. crocea) revealed that Lcp38 MAPK existed in both the cytoplasm and cell nucleus. The expression of Lcp38 MAPK after temperature stress was tested in LCK cells. The results indicated that Lcp38 MAPK transcripts were significantly upregulated under both cold (10 °C) and heat stress (35 °C) (P < 0.05). Furthermore, the phosphorylation levels of p38 MAPK as well the transcriptional levels of heat shock protein 27 (HSP27) and caspase3 in LCK cells were significantly induced under thermal exposure (P < 0.05). However, the cold- and heat induced HSP27 and caspase3 expression was significantly suppressed by SB203580, a specific inhibitor of p38-MAPK (P < 0.05). These findings indicated that Lcp38 MAPK might be involved in the cellular stress response via HSP27 and caspase3 in large yellow croaker.

Environmental hypoxia induces apoptosis in large yellow croaker Larimichthys crocea via both intrinsic and extrinsic pathways

Environmental hypoxia induces apoptosis in large yellow croaker Larimichthys crocea via both intrinsic and extrinsic pathways

The optimal feeding regime for large yellow croaker Larimichthys crocea, with an emphasis on obviating raw fish diet in commercial farming

Commercial farming of large yellow croaker Larimichthys crocea still relies on raw fish diet. The present study assessed the possibility to replace raw fish diet with a formulated diet containing 490 g/kg crude protein by optimizing feed and feeding management. In experiment I, fish weighing 10.8 g were fed with the formulated diet at either five frequencies (FF13, 1/3 meal/d; FF12, 1/2 meal/d; FF11, 1 meal/d; FF21, 2 meal/d; FF31, 3 meal/d) or at 1 meal/d but different times (FF11, at 6:00; FF11A, at 18:00), respectively. In experiment II, fish were fed at 2 meal/d with the formulated diet (FF21) or a quality raw fish diet (RF21). The final body weight (FBW), weight gain, feed intake and body lipid content, fish in/out ratio (FIFO) and waste outputs (nitrogen, phosphorus and carbon) were higher, whereas the nutrient and energy retention efficiencies (NRE, nitrogen; PRE, phosphorus; CRE, carbon; ERE, energy) were lower, in FF21 and FF31 than in FF13, FF12 and FF11. No significant differences were detected in FBW and weight gain either between FF21 and FF31 or between FF13, FF12 and FF11. The feed conversion ratio (FCR) and feed cost were higher in FF31 than in FF13, FF12 and FF11. The activities of superoxide dismutase (SOD) and catalase (CAT) in plasm, activities of glutathione peroxidase (GSH-PX) and CAT in liver, and malondialdehyde (MDA) content in plasm and liver were higher in FF31 than in FF11. No significant differences were detected in FBW, weight gain, NRE, PRE, CRE, ERE, body composition, feed cost, FIFO-, waste outputs, activities of antioxidant enzymes and MDA content in plasm and liver between FF11 and FF11A despite FF11A had higher feed intake and FCR compared with FF11. The FBW, weight gain, feed intake, FCR, body lipid content, feed cost, FIFO, waste outputs, activities of SOD and GSH-PX in liver, plasm CAT activity, and MDA content in plasm and liver were higher, whereas the survival, NRE, PRE and ERE were lower, in RF21 than in FF21. Our study indicates that 2 meal/d is the optimal feeding frequency for large yellow croaker, and completely replacing raw fish diet with the formulated diet in commercial farming benefits to increase fish survival and yield and to reduce feed cost, FIFO, waste outputs and body lipid deposition.

Integration of transcriptomics and metabolomics reveals the effects of sea currents on overwintering of large yellow croaker Larimichthys crocea in cage culture

Large yellow croaker cultured in cages often face overwintering challenges, which have led to significant economic losses in aquaculture. In natural sea areas, apart from low temperatures, sea current is an important factor affecting the overwintering of large yellow croaker. Therefore, we have set up cages with different mesh sizes to investigate the impact of sea currents on the overwintering of large yellow croaker. Three cages with different mesh sizes (0.5 cm, 1.0 cm, and 2.0 cm) were constructed, and 500 fish were placed in each cage. The group with the 0.5-cm mesh served as the control, while the other two groups were experimental (T1 and T2). Water temperature was monitored daily, and water flow rate was measured at the highest tide point of each month (The first and fifteenth days of the lunar calendar). The experiment lasted for 78 days, during which the body weights and lengths of the fish were measured. Muscle samples were taken for body composition analysis, and liver tissues from control and T2 were collected for transcriptome and metabolome analyses. The water temperature ranged from 8.5 to 12.5 °C during the experimental period. The average sea current velocities in the three groups were 0.049 m/s, 0.167 m/s, and 0.286 m/s, respectively. The survival rates of fish in the control, T1 and T2 groups were 85.25%, 80.66%, and 70.32%, respectively. The weight and condition factor decreased significantly in all groups, with the T2 group experiencing the greatest decline. The lipid and protein contents of the fish in the T2 group were significantly lower than those in the control and T1 groups. Transcriptome analysis showed that the insulin and FoxO pathways were related to the regulation of glycolysis/gluconeogenesis. The PPAR signaling pathway was related to the regulation of lipid metabolism. Metabolome analysis revealed 663 significantly different metabolites in the two groups, of which 10 were significantly downregulated and 51 were significantly upregulated. The combined analysis of transcriptome and metabolome showed that both metabolites and genes involved in the ABC transporter pathway were significantly differentially expressed. The results of the study indicated that fish in high flow rate cages require more fat and protein to provide energy. The insulin and PPAR pathways played important roles in regulating glucose and lipid metabolism. The ABC transporter pathway may play important roles in the transport of small molecule substances and participate in energy metabolism during low-temperature activity in large yellow croaker.

Molecular Characterization of Galectin-3 in Large Yellow Croaker Larimichthys crocea Functioning in Antibacterial Activity.

Galectins are proteins that play a crucial role in the innate immune response against pathogenic microorganisms. Previous studies have suggested that Galectin-3 could be a candidate gene for antibacterial immunity in the large yellow croaker Larimichthys crocea. In this study, we cloned the Galectin-3 gene in the large yellow croaker, and named it LcGal-3. The deduced amino acid sequence of LcGal-3 contains a carbohydrate recognition domain with two conserved β-galactoside binding motifs. Quantitative reverse transcription PCR (qRT-PCR) analysis revealed that LcGal-3 was expressed in all the organs/tissues that were tested, with the highest expression level in the gill. In Larimichthys crocea kidney cell lines, LcGal-3 protein was distributed in both the cytoplasm and nucleus. Moreover, we found that the expression of LcGal-3 was significantly upregulated upon infection with Pseudomonas plecoglossicida, as demonstrated by qRT-PCR analyses. We also purified the LcGal-3 protein that was expressed in prokaryotes, and found that it has the ability to agglutinate large yellow croaker red blood cells in a Ca2+-independent manner. The agglutination activity of LcGal-3 was inhibited by lipopolysaccharides (LPS) in a concentration-dependent manner, as shown in the sugar inhibition test. Additionally, LcGal-3 exhibited agglutination and antibacterial activities against three Gram-negative bacteria, including P. plecoglossicida, Vibrio parahaemolyticus, and Vibrio harveyi. Furthermore, we studied the agglutination mechanism of the LcGal-3 protein using blood coagulation tests with LcGal-3 deletion and point mutation proteins. Our results indicate that LcGal-3 protein plays a critical role in the innate immunity of the large yellow croaker, providing a basis for further studies on the immune mechanism and disease-resistant breeding in L. crocea and other marine fish.

Effects of transportation on physiological indices and metabolomics of the large yellow croaker Larimichthys crocea.

The aim of this study was to investigate the survival rate, biochemical indices, and metabolome changes of the large yellow croaker after 48 h of live transportation. Two hundred and forty large yellow croakers (body weight: 23.4 ± 5.3 g, total length: 12.2 ± 0.7 cm) were used in this experiment. The transport buckets were filled with fresh seawater and the parameters of the water were a temperature of 16 ± 0.5 °C and a dissolved oxygen content of 6.0-7.2 mg/L. Large yellow crokers were first divided to 0, 10, 20, and 30 mg/L MS-222 groups to observe the 12 h survival rate. The survival rate of 10 mg/L MS-222 group (T1) was the 95%, highest of all, and was further analyzed. The results of liver biochemical indices indicated inhibition of gluconeogenesis and pentose phosphate pathway metabolism. In addition, metabolomics analysis identified significantly differentially expressed metabolites between T1 group and 0 mg/L MS-222 control (C) groups. Furthermore, Kyoto Encyclopedia of Genes and Genomes (KEGG) results revealed that the pathways of amino acid metabolism, especially the lysine, aspartate, and homoserine in the liver were significantly affected. In conclusion, the combination of metabolomics and liver biochemical assays provided a characterization of the response mechanism of L. crocea exposed to live transportation.

Spatial-temporal dynamics of bacterioplankton communities in the breeding area of large yellow croaker Larimichthys crocea in Sansha Bay, China

Spatial-temporal dynamics of bacterioplankton communities in the breeding area of large yellow croaker Larimichthys crocea in Sansha Bay, China

Immune response of large yellow croaker Larimichthys crocea towards a recombinant vaccine candidate targeting the parasitic ciliate Cryptocaryon irritans.

Cryptocaryon irritans, a parasitic ciliate, pose a major threat to marine teleost fish aquaculture. So far, no effective and safe control method is available. In this study, the protective efficacy of a recombinant truncated surface antigen of C. irritans (rCiSA32.6t) for large yellow croaker (Larimichthys crocea) against the parasite challenge with a sub-lethal dose of the infective theronts was evaluated by comparing the relative percent survivals (RPS), the specific antibody titers in sera, and the expression levels of the immune-related genes among the negative or adjuvant control fish, fish intraperitoneally immunized with rCiSA32.6t. The results showed that a RPS of 50.1% in rCiSA32.6t-immunized fish was achieved in comparison to negative control fish against C. irritans. A significant increase was noted in the antigen-specific immunoglobulin M (IgM) and immunoglobulin T (IgT) antibody levels in the sera of the rCiSA32.6t-vaccinated fish. Compared to the negative control fish, quantitative real-time PCR analysis indicated that the interleukin-1beta, IgT, and IgM heavy chain mRNA level in the fish head kidney, spleen, gill, and skin tissue were upregulated post-rCiSA32.6t immunization. This study indicates that the rCiSA32.6t can provide a high level of immune protection against C. irritans infection in grouper and is therefore pursued as a candidate C. irritans vaccine.

Renal metabolomic profiling of large yellow croaker Larimichthys crocea acclimated in low salinity waters

Cultivation of Larimichthys crocea in low salinity water has been regarded as an effective way to treat diseases induced by pathogens in seawater. The kidney of euryhaline teleost plays important roles in not only osmoregulation but also regulation of intermediary metabolism. However, the renal responses of metabolism and osmoregulation in L. crocea to low salinity waters are still rarely reported. In this work, renal metabolomic analysis based on MS technique was conducted on the L. crocea following cultivation in salinities of 24, 8, 6, 4, and 2 ppt for 40 days. A total of 485 metabolites covering organic acids and derivatives (34.17 %), lipids and lipid-like molecules (17.55 %), organoheterocyclic compounds (12.22 %), nucleosides, nucleotides, and analogues (11.91 %), and organic oxygen compounds (10.97 %), were identified in L. crocea kidney. Compared with control group (salinity 24), nearly all amino acids, nucleotides, and their derivatives were decreased in the kidney of L. crocea, whereas most of lipid-related metabolites including phospholipid, glycerophospholipids, and fatty acids were increased. The decrease in urea and inorganic ions as well as TMAO, betaine and taurine in L. crocea kidney suggested the less demand for maintaining osmotic homeostasis. Several intermediary metabolites covering amino acids, TCA cycle intermediates, and fatty acids were also significantly changed to match with the shift of energy allocation from osmoregulation to other biological processes. The reduced energy demand for osmoregulation might contribute to the promotion of L. crocea growth under low salinity environment. What is more, carbamoylphosphate and urea that showed linear salinity response curves and higher ED50 values were potential biomarkers to adaptation to low salinity water. Overall, the characterization of metabolomes of L. crocea kidney under low salinity provided a better understanding of the adaptive mechanisms to low salinity water and potentially contributed to a reference for optimal culture salinity and feed formula of L. crocea culture in low salinity water.

A new pathogenic isolate of Kocuria kristinae identified for the first time in the marine fish Larimichthys crocea.

In recent years, new emerging pathogenic microorganisms have frequently appeared in animals, including marine fish, possibly due to climate change, anthropogenic activities, and even cross-species transmission of pathogenic microorganisms among animals or between animals and humans, which poses a serious issue for preventive medicine. In this study, a bacterium was clearly characterized among 64 isolates from the gills of diseased large yellow croaker Larimichthys crocea that were raised in marine aquaculture. This strain was identified as K. kristinae by biochemical tests with a VITEK 2.0 analysis system and 16S rRNA sequencing and named K. kristinae_LC. The potential genes that might encode virulence-factors were widely screened through sequence analysis of the whole genome of K. kristinae_LC. Many genes involved in the two-component system and drug-resistance were also annotated. In addition, 104 unique genes in K. kristinae_LC were identified by pan genome analysis with the genomes of this strain from five different origins (woodpecker, medical resource, environment, and marine sponge reef) and the analysis results demonstrated that their predicted functions might be associated with adaptation to living conditions such as higher salinity, complex marine biomes, and low temperature. A significant difference in genomic organization was found among the K. kristinae strains that might be related to their hosts living in different environments. The animal regression test for this new bacterial isolate was carried out using L. crocea, and the results showed that this bacterium could cause the death of L. crocea and that the fish mortality was dose-dependent within 5 days post infection, indicating the pathogenicity of K. kristinae_LC to marine fish. Since K. kristinae has been reported as a pathogen for humans and bovines, in our study, we revealed a new isolate of K. kristinae_LC from marine fish for the first time, suggesting the potentiality of cross-species transmission among animals or from marine animals to humans, from which we would gain insight to help in future public prevention strategies for new emerging pathogens.

Antimicrobial activity of histone1-derived peptides from large yellow croaker Larimichthys crocea

The function of histones is well-known in the folding and packaging of DNA, gene regulation, tumor formation, and apoptosis and they also have bactericidal properties. In the present study, two histone1-derived antimicrobial peptides (LcH1–1 and LcH1–2) from Larimichthys crocea were described. The full-length cDNA of Lc-histione1 (LcH1) consisted of 1388 base pairs, including 588 bp open reading frame of, 5′ UTR of 249 bp and 3′ that of 551 bp. The molecular weight of LcH1 is 20,793.2 Da and the isoelectric point (pI) was 10.99. Two DNA-binding sites were predicted in LcH1. Tissue distribution analysis showed LcH1 presented a high expression pattern in gill, liver, and intestine. It also showed a time-dependent expression pattern after Cryptocaryon irritans infection. Maximum likelihood phylogenetic analysis showed that LcH1 had a relatively close relationship to homologous genes from Collichthys lucidus and Nibea albiflora. Both LcH1–1 and LcH1–2 formed an amphipathic α-helix structure and displayed a broad antimicrobial spectrum against a variety of microbial strains. In addition, scanning electronic microscope (SEM) showed that the LcH1–1 and LcH1–2 peptides were prone to damage the integrity of the bacteria membrane. These results suggested that LcH1 may be involved in the immunoreaction of L. crocea under C. irritans challenge and that LcH1-derived antimicrobial peptides may provide a potential strategy for the control of parasitic and bacterial diseases.

Growth-promoting effect of antimicrobial peptide Scy-hepc on mariculture large yellow croaker Larimichthys crocea and the underlying mechanism

With the antibiotics prohibition in feedstuffs worldwide, antimicrobial peptides (AMPs) are considered a more promising substitute for antibiotics to be used as feed additives, and positive results have been reported in livestock feeding studies. However, whether dietary supplementation of AMPs could promote the growth of mariculture animals such as fish and the underlying mechanism has not been elucidated yet. In the study, a recombinant AMP product of Scy-hepc was used as a dietary supplement (10 mg/kg) to feed mariculture juvenile large yellow croaker (Larimichthys crocea) with an average initial body weight (BW) of 52.9 g for 150 days. During the feeding trial, the fish fed with Scy-hepc showed a significant growth-promoting performance. Especially at 60 days after feeding, fish fed with Scy-hepc weighed approximately 23% more than the control group. It was further confirmed that the growth-related signaling pathways such as the GH-Jak2-STAT5-IGF1 growth axis, the PI3K-Akt and Erk/MAPK pathways were all activated in the liver after Scy-hepc feeding. Furthermore, a second repeated feeding trial was scheduled for 30 days using much smaller juvenile L. crocea with an average initial BW of 6.3 g, and similar positive results were observed. Further investigation revealed that the downstream effectors of the PI3K-Akt pathway, such as p70S6K and 4EBP1, were significantly phosphorylated, suggesting that Scy-hepc feeding might promote translation initiation and protein synthesis processes in the liver. Taken together, as an effector of innate immunity, AMP Scy-hepc played a role in promoting the growth of L. crocea and the underlying mechanism was associated with the activation of the GH-Jak2-STAT5-IGF1 axis, as well as the PI3K-Akt and Erk/MAPK signaling pathways.

Effects of rusted copper alloy in prevention and control of cryptocaryoniasis in large yellow croaker Larimichthys crocea

Copper alloy can be used to prevent and control cryptocaryoniasis in marine fish. However, it is not known whether the soaking corrosion of seawater can affect the parasiticidal effect of copper alloy. This study compared the killing effects of copper alloy mesh (CAM) and rust copper alloy mesh (RCAM) on the protomonts and tomonts of Cryptocaryon irritans and the control effects of cryptocaryoniasis in Larimichthys crocea. Results showed that with the increase of contact time with CAM and RCAM, the survival rate of the protomonts and tomonts of C. irritans were gradually decreased. The contents of Cu2+ and malondialdehyde (MDA) in the tomonts gradually increased, while the activities of superoxide dismutase (SOD), Na+/K+-ATPase and Ca2+/Mg2+-ATPase in the tomonts increased first and then decreased. Morphological observation showed that sufficient contact with CAM or RCAM damaged the cyst wall of the tomonts, and the cytoplasm have lysed and spilled out of the cell. In the treatment test of diseased fish, at the end of the assay (14 d), the survival rates of L. crocea in the CAM treatment group (group B) and the RCAM treatment group (group C) were 77.77% and 64.44%, respectively, which were significantly higher than those in the untreated group (group A, 0.00%) (P < 0.05). RCAM could still successfully prevent the outbreak of cryptocaryoniasis in a short time. Our study serves as a theoretical reference for the long–term application of metal materials in the prevention and control of cryptocaryoniasis in marine fish.

SARM suppresses TRIF, TRAF3, and IRF3/7 mediated antiviral signaling in large yellow croaker Larimichthys crocea.

As a TIR domain-containing molecular, sterile α-and armadillo motif-containing protein (SARM) acts as an adaptor in Toll-like receptor (TLR) signaling, and also plays important roles in mediating apoptosis and neuronal injury. In the present study, the ortholog of SARM, named as Lc-SARM, was cloned and identified in large yellow croaker (Larimichthys crocea). The full-length ORF of Lc-SARM consists of 2,154 bp, encoding a protein of 717 amino acids (aa), which is comprised of an N-terminal ARM domain, two SAM domains, and a C-terminal TIR domain. Confocal microscopy revealed that Lc-SARM was mainly distributed in the cytoplasm, and the mRNA expression level of Lc-SARM was broadly distributed in all the detected organs/tissues, with the highest expression level found in the brain. The expression patterns of Lc-SARM could be induced in response to poly I:C, LPS, PGN stimulations, and Pseudomonas plecoglossicida infection. Notably, although the overexpression of Lc-SARM could significantly induce NF-κB, IRF3, IRF7, and type I IFN promoter activation, whereas the co-expression of Lc-SARM with Lc-TRIF, Lc-TRAF3, Lc-IRF3, or Lc-IRF7 significantly down-regulated the induction of NF-κB, IRF3, IRF7, or type I IFN promoter activation, and suppressed the antiviral effects as well as the downstream antiviral-related genes expression compared to the only overexpression of Lc-TRIF, Lc-TRAF3, Lc-IRF3, or Lc-IRF7. Co-immunoprecipitation (Co-IP) assays also demonstrated that Lc-SARM interacts separately with Lc-TRIF, Lc-TRAF3, Lc-IRF3, and Lc-IRF7. It is thus collectively suggested that Lc-SARM functions as a negative regulator in Lc-TRIF, Lc-TRAF3, and Lc-IRF3/7 involved antiviral signaling.

Spatial Genetic Structure and Diversity of Large Yellow Croaker (Larimichthys crocea) from the Southern Yellow Sea and North-Central East China Sea: Implications for Conservation and Stock Enhancement

As a wild resource, the large yellow croaker Larimichthys crocea has been seriously threatened since the mid-1980s. Owing to the implementation of protection measures, such as the establishment of a protection zone, fishing prohibitions, restocking programs and successful mariculture, its resources have gradually recovered year by year. Limited by the low spatial resolution and incomplete spatial coverage of sampling stations, the spatial genetic structure and diversity of large yellow croakers from the southern Yellow Sea and north-central East China Sea remains unclear. In order to evaluate the genetic diversity status of this wild stock, 22 wild sites were collected from the southern Yellow Sea and north-central East China Sea and analyzed by investigating genetic variability and its population structure using mitochondrial COI sequence in this study. Among the 662 sequences, a total of 71 different haplotypes were defined. The haplotype diversity (h) and nucleotide diversity (π) values were 0.644~1.000 and 0.00220~0.00473 respectively. The highest h and π occurred in the southern Yellow Sea (YS). AMOVA analysis showed no genetic differentiation among those 22 sites after Bonferroni correction. By comparing with previous studies, the croaker has maintained relatively steady genetic diversity in recent years. Our result also suggested that the croakers in the South Yellow Sea and north-central East China Sea belonged to the same group. Thus, they can be released as a management unit without regard for heterogenicity among those in the sea area. The YS populations can serve as parents for released fish fries in the South Yellow Sea and north-central East China Sea.

Development and applications of chromosome-specific cytogenetic BAC-FISH probes in Larimichthys crocea

Large yellow croaker Larimichthys crocea (Richardson) is an important member in family Sciaenidae, and one of most productive mariculture fishes in China. Fluorescence in situ hybridization is a useful tool for cytogenetics and genomics research. Here, we demonstrated that bacterial artificial chromosome (BAC) clones could be used to identify individual chromosomes in large yellow croaker, and also to study chromosome evolution in the related species. By using BAC paired-end sequencing and sequence alignment, 435 BAC clones were anchored to 24 pseudochromosomes of large yellow croaker genome. Among them, 72 BAC clones with low repeat content were selected and passed PCR test, and then grouped by chromosome for FISH test. As a results, there were 67 BAC clones, 2 to 3 BAC clones per chromosome, generated specific and stable signal at expected position. Then, a dual-color FISH probe cocktail composed of 48 of these BAC clones was formulated and used to hybridize metaphase chromosome spreads, resulting in distinct signal patterns on each chromosome pair, which help to distinguish all chromosomes in the metaphase spreads of large yellow croaker. The chromosome-specific BAC-FISH probes were also applied to a close relative of large yellow croaker, Collichthys lucidus, demonstrating that its Y chromosome originated from the fusion of Chr.1 and Chr.7. Thus, our study provides the first set of chromosome-specific FISH probes in family Sciaenidae, which will play an important role in cytogenetics and genomics research in the family.