The growth hormone gene (GH) affects animal growth and is a potential target for genetic studies of variation related to growth traits. In this study, we analyzed single nucleotide polymorphisms (SNPs) in GH intron re...The growth hormone gene (GH) affects animal growth and is a potential target for genetic studies of variation related to growth traits. In this study, we analyzed single nucleotide polymorphisms (SNPs) in GH intron regions and their associations with growth traits in large yellow croaker, Larimichthys crocea, from Zhejiang and Fujian stocks. The results of PCR-single strand conformation polymorphism showed two haplotypes of intron 1, named AA and AB genotypes, in Zhejiang stock. AB exhibited an SNP at position 196 (G A) that was negatively correlated with body height and positively correlated with standard length/body height (P 0.05). Two different genotypes, CC and CD, were identified in intron 2 in Fujian stock, with CD showing an SNP at position 692 (T C). The CD genotype had a significantly positive correlation with both weight and total length (P 0.01). These basic data highlight the potential for using GH as a genetic marker of fish growth in marker assisted selection.展开更多
We estimated the reproductive success of adult brood fish and the heritability of growthrelated traits for large yellow croaker （Larimichthys crocea）. We created two independent full-factorial cross groups （1 and 2...We estimated the reproductive success of adult brood fish and the heritability of growthrelated traits for large yellow croaker （Larimichthys crocea）. We created two independent full-factorial cross groups （1 and 2） by crossing 4 malesx4 females and 4 males^3 females, respectively. We measured the body weight （BW）, body length （BL）, and body height （BH） of 281 individuals from group 1 and 318 individuals from group 2 at 20 months post hatch （harvest age）. We also collected a tissue sample from each individual. The parents and offspring were genotyped using six polymorphic mierosatellites. Of the 599 offspring, 99.2% were assigned to a single pair of parents. In both groups, some parent pairs produced a large number of offsprings while other pairs did not produce any offspring. The genetic diversity and putative No were lower in the offsprings than in the parents in both groups. The heritability estimates at 20-month age were 0.13±0.10 for BW, 0.19±0.13 for BL, and 0.09±0.06 for BH. The genetic and phenotype correlation between BW, BL, and BH was close to 1. The results provide basic information for selective breeding and further genetic characterization of large yellow croaker.展开更多
Host-pathogen interactions are highly complex,involving large dynamic changes in gene expression during infection. These interactions are fundamental to understanding anti-infection immunity of hosts, as well as the p...Host-pathogen interactions are highly complex,involving large dynamic changes in gene expression during infection. These interactions are fundamental to understanding anti-infection immunity of hosts, as well as the pathogenesis of pathogens. For bacterial pathogens interacting with animal hosts, timeresolved dual RNA-seq of infected tissue is difficult to perform due to low pathogen load in infected tissue. In this study, an acute infection model of Larimichthys crocea infected by Pseudomonas plecoglossicida was established. The spleens of infected fish exhibited typical symptoms, with a maximum bacterial load at two days post-injection(dpi). Time-resolved dual RNA-seq of infected spleens was successfully applied to study hostpathogen interactions between L. crocea and P.plecoglossicida. The spleens of infected L. crocea were subjected to dual RNA-seq, and transcriptome data were compared with those of noninfected spleens or in vitro cultured bacteria. Results showed that pathogen-host interactions were highly dynamically regulated, with corresponding fluctuations in host and pathogen transcriptomes during infection. The expression levels of many immunogenes involved in cytokine-cytokine receptor,Toll-like receptor signaling, and other immunerelated pathways were significantly up-regulated during the infection period. Furthermore, metabolic processes and the use of oxygen in L. crocea were strongly affected by P. plecoglossicida infection. The WGCNA results showed that the metabolic process was strongly related to the entire immune process.For P. plecoglossicida, the expression levels of motility-related genes and flagellum assemblyrelated genes were significantly up-regulated. The results of this study may help to elucidate the interactions between L. crocea and P.plecoglossicida.展开更多
Oxygen is an essential molecule for animal respiration,growth,and survival.Unlike in terrestrial environments,contamination and climate change have led to the frequent occurrence of hypoxia in aquatic environments,thu...Oxygen is an essential molecule for animal respiration,growth,and survival.Unlike in terrestrial environments,contamination and climate change have led to the frequent occurrence of hypoxia in aquatic environments,thus impacting aquatic animal survival.However,the adaptative mechanisms underlying fish responses to environmental hypoxia remain largely unknown.Here,we used large yellow croaker(Larimichthys crocea)and large yellow croaker fry(LYCF)cells to investigate the roles of the Hif-1α/Hsf1/Hsp70 signaling pathway in the regulation of cellular redox homeostasis,and apoptosis.We confirmed that hypoxia induced the expression of Hif-1α,Hsf1,and Hsp70 in vivo and in vitro.Genetic Hsp70 knockdown/overexpression indicated that Hsp70 was required for maintaining redox homeostasis and resisting oxidative stress in LYCF cells under hypoxic stress.Hsp70 inhibited caspase-dependent intrinsic apoptosis by maintaining normal mitochondrial membrane potential,enhancing Bcl-2 mRNA and protein expression,inhibiting Bax and caspase3 mRNA expression,and suppressing caspase-3 and caspase-9 activation.Hsp70 suppressed caspaseindependent intrinsic apoptosis by inhibiting nuclear translocation of apoptosis-inducing factor(AIF)and disturbed extrinsic apoptosis by inactivating caspase-8.Genetic knockdown/overexpression of Hif-1αand dual-luciferase reporter assay indicated that Hif-1αactivated the Hsf1 DNA promoter and enhanced Hsf1 mRNA transcription.Hsf1 enhanced Hsp70 mRNA transcription in a similar manner.In summary,the Hif-1α/Hsf1/Hsp70 signaling pathway plays an important role in regulating redox homeostasis and anti-apoptosis in L.crocea under hypoxic stress.展开更多
An （AC）n-microsatellite-enriched library for Larimichthys crocea was constructed in this study. Primers for fifty simple sequence repeat （SSR） loci were synthesized and genotyped on 30 L. crocea individuals from G...An （AC）n-microsatellite-enriched library for Larimichthys crocea was constructed in this study. Primers for fifty simple sequence repeat （SSR） loci were synthesized and genotyped on 30 L. crocea individuals from Guanjingyang wild population （WP） in Fujian Province and 38 individuals from Ningbo cultured population （CP） in Zhejiang Province. Only 21 loci were successfully amplified and polymorphic in two populations. In WP, the observed heterozygosity （Ho） ranged from 0.233 to 0.900 and the expected heterozygosity （HE） ranged from 0.326 to 0.893, with an average of 7.8 alleles/locus, the polymorphism information content （PIC） ranged from 0.283 to 0.866 （mean 0.731）. In CP, the Ho ranged from 0.189 to 0.892 and the HE ranged from 0.333 to 0.800, with an average of 4.4 alleles/locus. The probability test showed significant departures from HWE in 9 and 2 of the 21 loci in WP and in CP, respectively. Deficiency of heterozygotes at four loci showed the presence of null alleles （P 〈0.01）. The PIC of 20 microsatellite loci in WP were greater than 0.50. Overall, these novel markers are potentially useful for the study of population genetics, construction of genetic linkage and quantitative trait loci maps in large yellow croaker.展开更多
The large yellow croaker(Larimichthys crocea)is an important mariculture fish in China.Farmed large yellow croaker undergo periods of fasting to adapt to the environment or to improve meat quality.To better understand...The large yellow croaker(Larimichthys crocea)is an important mariculture fish in China.Farmed large yellow croaker undergo periods of fasting to adapt to the environment or to improve meat quality.To better understand the physiological responses of their muscle tissues to fasting stresses,we analyzed the transcriptomes and proteome s of both normally-fed and fasting fish groups and identified 7578 differentially expressed genes(DEGs)and 297 differentially expressed proteins(DEPs)among them.Gene ontology and KEGG analysis showed that the enriched biological pathways were mainly involved in various synthetic and catabolic pathways,especially the protein metabolism.Based on the omics data,nine DEGs related to muscle composition(CAN3,MYL3,and TNNC2),growth(MSTN and MYF5),autophagy(TSC2 and ULK1),and the ubiquitin proteasome pathway(PRS6 B and UCHL3)were examined using qPCR.In response to fasting stress,MYL3 and TNNC2 were significantly downregulated,while genes associated with autophagy and the ubiquitin proteasome pathway were significantly upregulated.In re sponse to fasting stres s,MYL3,TNNC2,and MYF5 positively correlated with muscle growth were significantly downregulated,while inhibiting growth MSTN and genes associated with autophagy and the ubiquitin proteasome pathways were significantly upregulated.These results clarify the effects of fasting on metabolic changes in their muscle components and growth at the molecular level.展开更多
The large yellow croaker, Larimichthys crocea is an important marine fish in China with a high economic value. In the last decade, the stock conservation and aquaculture industry of this species have been facing sever...The large yellow croaker, Larimichthys crocea is an important marine fish in China with a high economic value. In the last decade, the stock conservation and aquaculture industry of this species have been facing severe challenges because of wild population collapse and degeneration of important economic traits. However, genes contributing to growth and immunity in L. crocea have not been thoroughly analyzed, and available molecular markers are still not sufficient for genetic resource management and molecular selection. In this work, we sequenced the transcriptome in L. eroeea liver tissue with a Roche 454 sequencing platform and assembled the transcriptome into 93 801 transcripts. Of them, 38 856 transcripts were successfully annotated in nt, nr, Swiss-Prot, InterPro, COG, GO and KEGG databases. Based on the annotation information, 3 165 unigenes related to growth and immunity were identified. Additionally, a total of 6 391 simple sequence repeats （SSRs） were identified from the transcriptome, among which 4 498 SSRs had enough flanking regions to design primers for polymerase chain reactions （PCR）. To access the polymorphism of these markers, 30 primer pairs were randomly selected for PCR amplification and validation in 30 individuals, and 12 primer pairs （40.0%） exhibited obvious length polymorphisms. This work applied RNA-Seq to assemble and analyze a live transeriptome in L. crocea. With gene annotation and sequence information, genes related to growth and immunity were identified and massive SSR markers were developed, providing valuable genetic resources for future gene functional analysis and selective breeding ofL. eroeea.展开更多
As an economically important marine fish,the large yellow croaker Larimichthys crocea suffered from marine white spot disease caused by the ectoparasite Cryptocaryon irritans in recent years.This disease not only coul...As an economically important marine fish,the large yellow croaker Larimichthys crocea suffered from marine white spot disease caused by the ectoparasite Cryptocaryon irritans in recent years.This disease not only could result in physiological damage,but also lead to secondary bacterial invasion.Reports indicated some AMPs(antimicrobial peptides)were of antiparasitic activity to C.irritans.Hepcidin-like(Lc-HepL)was one of the significant differential expression genes excavated from the transcriptome following a challenge with C.irritans.In this study,we characterized this AMP’s bioactivity based on the levels of mRNA and protein.After challenged by C.irritans,qRT-PCR showed Lc-HepL was significantly upregulated in six tissues,including gill,muscle,liver,head kidney and spleen during theront infection,trophont falling off,and secondary bacterial invasion stages,which implicated a role Lc-HepL played in the immune defense against C.irritans and secondary bacterial infection.Recombinant Lc-HepL(rLc-HepL)was induced and purified successfully.rLc-HepL exhibited antibacterial activity to certain bacteria in a dose-and time-dependent manners.Anti-C.irritans activity was explored for the first time and found it could cause the theronts membrane rupture and contents leakage.These results provided the first evidence that Lc-HepL had strong antiparasitic activity against marine fish ectoparasites C.irritans theronts.Together,data indicated that Lc-HepL might be an important component in the innate immune system against C.irritans and has the potential to be employed in future drug development.展开更多
Larimichthys crocea is a marine fish species cultured in China.Short-term starvation is often applied to improve the quality of cultured L.crocea,and the expression of ghrelin in tissues of stomach,muscle,brain,intest...Larimichthys crocea is a marine fish species cultured in China.Short-term starvation is often applied to improve the quality of cultured L.crocea,and the expression of ghrelin in tissues of stomach,muscle,brain,intestines,liver,and kidney,involved in starvation response,under starvation conditions were studied to understand the effect of starvation on the expression of ghrelin in L.crocea juveniles.The ghrelin expression was tissue-specific,and expression was significantly higher in the stomach compared to other tissues(P<0.01).Additionally,ghrelin expression in different tissues changed along with prolongation of fasting.In the stomach,ghrelin expression levels increased gradually at the beginning of the fast,and then declined after eight days of fasting.Gene expression in the brain and intestines increased at the beginning of the fast,and then decreased with longer fasting time.Interestingly,ghrelin expression declined at the beginning of the fast,then increased with longer fasting in the kidneys and muscles.These results suggest that ghrelin is involved in starvation response in L.crocea juveniles.This study provids insights into ghrelin function and an important reference for the development of reasonable feeding strategies for L.crocea juveniles.展开更多
Antitumor activity is one characteristic function of some certain antimicrobial peptides(AMPs)found in recent years.In the present study,we attempted to detect potential anticancer activity of a recombinant piscidin 5...Antitumor activity is one characteristic function of some certain antimicrobial peptides(AMPs)found in recent years.In the present study,we attempted to detect potential anticancer activity of a recombinant piscidin 5-like from Larimichthys crocea(r Lc-P5L)which owned widely antibacterial and strong antiparasitic activity in vitro.The light microscope observation indicated r Lc-P5L was of antitumor activity to He La cells,293 T cells and L929 cells.MTT assay showed the toxic sensitivity of r Lc-P5L to three tumor cell strains was 293 T>L929>He La.Scanning electron microscope(SEM)results showed r Lc-P5L behaved like a lytic peptide to cause damage on cells membrane of L929 cells by forming globular clusters,even pores at 60μmol/L,or degrading membrane to make it completely lose cytoskeleton structure at 80μmol/L;r Lc-P5L treatment also resulted in DNA degradation.Fluorescence observation results indicated r Lc-P5L could cause L929 cells at least two obvious changes:one is nucleus,nuclear chromatin condensed in the margin,nuclear volume became smaller and shrank to be out of shape,or lysed to be debris;the other is cytoskeleton,they became disordered and polarized to make cells atrophic shapes,or even lysed to be debris.In summary,r Lc-P5L owned potential anticancer activity causing membrane structure damage and genome DNA degradation.Interestingly,treatment with different concentration of r Lc-P5L seemingly caused the similar but different changes,whether it indeed gave rise to cancer cells diverse death way,the further studies should be performed,and the detailed mechanisms were still need further explored.展开更多
Elovl4 is a fatty acyl elongase which participates in the biosynthesis of long-chain polyunsaturated fatty acids(LCPUFA).Herein we isolated and functionally characterized the elovl4a gene in the large yellow croaker L...Elovl4 is a fatty acyl elongase which participates in the biosynthesis of long-chain polyunsaturated fatty acids(LCPUFA).Herein we isolated and functionally characterized the elovl4a gene in the large yellow croaker Larimichthys crocea,and investigated the regulatory effects of transcription factors Hnf4α,Lxrα,Pparαon the activity of elovl4a promoter.Tissue expression pattern revealed that elovl4a was widely expressed in several tissues and predominantly in brain.Heterologous expression in yeast showed that L.crocea Elovl4a could effectively elongate both C18 and C20 PUFA substrates to C22 fatty acid.In addition,the affinity of large yellow croaker Elovl4a to n-6 series fatty acids is weak and cannot prolong 18:2n-6 and 18:3n-6.Moreover,L.crocea elovl4a reporter activities were elevated by 1.31-,1.39-and 1.48-fold via over-expression of Lxrα,Pparαand Hnf4α,respectively.The above findings can enrich the knowledge of biosynthesis pathway of LC-PUFA in the large yellow croaker and contribute to elucidate the LC-PUFA anabolism mechanism in fish.展开更多
To ascertain the nutrient components and nutritive quality of the flesh of big yellow croaker in three culture conditions （ traditional cage, offshore cage and cage-free）, basic nutritional components,amino acid,fat...To ascertain the nutrient components and nutritive quality of the flesh of big yellow croaker in three culture conditions （ traditional cage, offshore cage and cage-free）, basic nutritional components,amino acid,fatty acid and mineral elements were determined. The results indicated that crude protein in flesh of the big yellow croaker in cage-free culture was higher than that in offshore cage and much higher than that in traditional cage （ P 〈0.05）. Crude fat of the croaker cul- tured in the traditional cage was twice as high as that in cage-free culture, while that in the offshore cage was in the middle. Proline content in the cage-free culture was much higher than that in the offshore cage, and also than that in the traditional cage （P 〈 0.05 ）. There was no significant difference in the content of alanine, methionine and tryptophan （ P 〈 0.05）. Contents of other amino acids had no significant difference between the cage-free culture and offshore cage, but were much lower in the traditional cage （P 〈 0.05 ）. Top six fatty acids were 9-Hexadecenoic acid, palmitic acid,9-Octadecenoic acid, Octadecauoic acid, DHA and EPA. The palmitic acid content was the highest in cage-free culture and in traditional cage, 9-Octadeeenoic acid content was the highest in offshore cage. Content of unsaturat- ed fatty acids in cage-free culture, offshore cage and traditional cage was 63.60,66.32,57.67, respectively, and polyunsaturated fatty acid was 29.10, 28.57, and 24.40. Content of DHA in cage-free culture was significantly higher than that in the offshore and traditional cage. Content of zinc had no significant difference in three culture models. Content of phosphorus had no significant difference between that in cage-free culture and offshore cage, was lower in the traditional cage. The cage-free cultured croakers had the highest content of calcium and phosphorus. Content of selenium was about the same between the offshore cage and the traditional cage stocking, higher in the cage-free culture. This research has considerable application value for identifying quality and sources of the big yellow croakers.展开更多
The aim of the present study was to determine the preventive effects of the polysaccharide of Larimichthys crocea swim bladder(PLCSB) on CCl4-induced hepatic damage in ICR mice.The in vitro preventive effects of PLCSB...The aim of the present study was to determine the preventive effects of the polysaccharide of Larimichthys crocea swim bladder(PLCSB) on CCl4-induced hepatic damage in ICR mice.The in vitro preventive effects of PLCSB on CCl4-induced liver cytotoxic effect were evaluated in BRL 3A rat liver cells using the MTT assay.The serum levels of AST,ALT,and LDH in mice were determined using commercially available kits.The levels of IL-6,IL-12,TNF-α,and IFN-γ were determined using ELISA kits.The pathological analysis of hepatic tissues was performed with H and E staining,and the gene and protein expressions were determined by RT-PCR and Western blotting,respectively.PLCSB(20 μg·m L-1) could increase the growth of BRL 3A rat liver cells treated with CCl4.The serum levels of AST,ALT,and LDH were significantly decreased when the mice were treated with two doses of PLCSB,compared with the control mice(P < 0.05).PLCSB-treated groups also showed reduced levels of the serum pro-inflammatory cytokines IL-6,IL-12,TNF-α,and IFN-γ.PLCSB could decrease the liver weight,compared to the CCl4-treated control mice.The histopathology sections of liver tissues in the 100 mg·kg-1 PLCSB group indicated that the animals were recovered well from CCl4 damage,but the 50 mg·kg-1 PLCSB group showed necrosis to a more serious extent.The 100 mg·kg-1 PLCSB group showed significantly decreased mR NA and protein expression levels of NF-κB,i NOS,and COX-2,and increased expression of IκB-α compared with the CCl4-treated control group.In conclusion,PLCSB prevented from CCl4-induced hepatic damage in vivo.展开更多
The large yellow croaker Larimichthys crocea is one of the most important fish species in China. To estimate the re- productive success of breeders, three independent full-factorial crosses were created and the fins o...The large yellow croaker Larimichthys crocea is one of the most important fish species in China. To estimate the re- productive success of breeders, three independent full-factorial crosses were created and the fins of breeders and progenies were sampled for microsatellite analysis. Out of 959 offspring from three sets, 99.6% were assigned to their parents using 6-7 mi- crosatellite markers. In all crosses, some parent pairs produced a large number of offspring and some parent pairs did not produce any offspring. The contributions of male or female parents were unequal, ranging from 1.0-89.3% across the three sets. The loss of putative Ne was 69.6% in set 1, 31.2% in set 2 and 57.6% in set 3. These results suggest that the unequal contribution of parents is universal in artificial breeding ofL. crocea, especially in a small population, and this should be taken into account in hatcheries or when releasing animals for resource enhancement展开更多
In mammals,mitofusin 2(MFN2)is involved in mitochondrial fusion,and suppresses the virus-induced RIG-I-like receptor(RLR)signaling pathway.However,little is known about the function of MFN2 in non-mammalian species.In...In mammals,mitofusin 2(MFN2)is involved in mitochondrial fusion,and suppresses the virus-induced RIG-I-like receptor(RLR)signaling pathway.However,little is known about the function of MFN2 in non-mammalian species.In the present study,we cloned an MFN2 ortholog(LcMFN2)in large yellow croaker(Larimichthys crocea).Phylogenetic analysis showed that MFN2 emerged after the divergence of amphioxus and vertebrates.The protein sequences of MFN2 were well conserved from fsh to mammals.LcMFN2 was expressed in all the tissues/organs examined at diferent levels,and its expression was upregulated in response to poly(I:C)stimulation.Overexpression of LcMFN2 inhibited MAVS-induced type I interferon(IFN)promoter activation and antiviral gene expression.In contrast,knockdown of endogenous LcMFN2 enhanced poly(I:C)induced production of type I IFNs.Additionally,LcMFN2 enhanced K48-linked polyubiquitination of MAVS,promoting its degradation.Also,overexpression of LcMFN2 impaired the cellular antiviral response,as evidenced by the increased expression of viral genes and more severe cytopathic efects(CPE)in cells infected with spring viremia of carp virus(SVCV).These results indicated that LcMFN2 inhibited type I IFN response by degrading MAVS,suggesting its negative regulatory role in cellular antiviral response.Therefore,our study sheds a new light on the regulatory mechanisms of the cellular antiviral response in teleosts.展开更多
基金Supported by the National Basic Research Program of China(973Program)(No.2010CB126304)the Special Fund for Agro-scientific Research in the Public Interest(No.200903046)the Project from the Ministry of Science and Technology of China(No.2006DKA30470017)
文摘The growth hormone gene (GH) affects animal growth and is a potential target for genetic studies of variation related to growth traits. In this study, we analyzed single nucleotide polymorphisms (SNPs) in GH intron regions and their associations with growth traits in large yellow croaker, Larimichthys crocea, from Zhejiang and Fujian stocks. The results of PCR-single strand conformation polymorphism showed two haplotypes of intron 1, named AA and AB genotypes, in Zhejiang stock. AB exhibited an SNP at position 196 (G A) that was negatively correlated with body height and positively correlated with standard length/body height (P 0.05). Two different genotypes, CC and CD, were identified in intron 2 in Fujian stock, with CD showing an SNP at position 692 (T C). The CD genotype had a significantly positive correlation with both weight and total length (P 0.01). These basic data highlight the potential for using GH as a genetic marker of fish growth in marker assisted selection.
基金Supported by the Special Fund for Agro-Scientific Research in the Public Interest (No. 200903029-4)Key Project of Science and Technology Department of Fujian Province (No. 2010N0021)the Foundation for Young Professors of Jimei University (No. 2009C002)
文摘We estimated the reproductive success of adult brood fish and the heritability of growthrelated traits for large yellow croaker （Larimichthys crocea）. We created two independent full-factorial cross groups （1 and 2） by crossing 4 malesx4 females and 4 males^3 females, respectively. We measured the body weight （BW）, body length （BL）, and body height （BH） of 281 individuals from group 1 and 318 individuals from group 2 at 20 months post hatch （harvest age）. We also collected a tissue sample from each individual. The parents and offspring were genotyped using six polymorphic mierosatellites. Of the 599 offspring, 99.2% were assigned to a single pair of parents. In both groups, some parent pairs produced a large number of offsprings while other pairs did not produce any offspring. The genetic diversity and putative No were lower in the offsprings than in the parents in both groups. The heritability estimates at 20-month age were 0.13±0.10 for BW, 0.19±0.13 for BL, and 0.09±0.06 for BH. The genetic and phenotype correlation between BW, BL, and BH was close to 1. The results provide basic information for selective breeding and further genetic characterization of large yellow croaker.
基金the National Natural Science Foundation of China(31672694,31972836)Fujian Provincial Special Fund for Marine and Fishery Protection and Development(MCZ[2019]062)。
文摘Host-pathogen interactions are highly complex,involving large dynamic changes in gene expression during infection. These interactions are fundamental to understanding anti-infection immunity of hosts, as well as the pathogenesis of pathogens. For bacterial pathogens interacting with animal hosts, timeresolved dual RNA-seq of infected tissue is difficult to perform due to low pathogen load in infected tissue. In this study, an acute infection model of Larimichthys crocea infected by Pseudomonas plecoglossicida was established. The spleens of infected fish exhibited typical symptoms, with a maximum bacterial load at two days post-injection(dpi). Time-resolved dual RNA-seq of infected spleens was successfully applied to study hostpathogen interactions between L. crocea and P.plecoglossicida. The spleens of infected L. crocea were subjected to dual RNA-seq, and transcriptome data were compared with those of noninfected spleens or in vitro cultured bacteria. Results showed that pathogen-host interactions were highly dynamically regulated, with corresponding fluctuations in host and pathogen transcriptomes during infection. The expression levels of many immunogenes involved in cytokine-cytokine receptor,Toll-like receptor signaling, and other immunerelated pathways were significantly up-regulated during the infection period. Furthermore, metabolic processes and the use of oxygen in L. crocea were strongly affected by P. plecoglossicida infection. The WGCNA results showed that the metabolic process was strongly related to the entire immune process.For P. plecoglossicida, the expression levels of motility-related genes and flagellum assemblyrelated genes were significantly up-regulated. The results of this study may help to elucidate the interactions between L. crocea and P.plecoglossicida.
基金This work was supported by the National Key Research and Development Program of China(2018YFC1406300)NSFC-Zhejiang Joint Fund for the Integration of Industrialization and Informatization(U1809212)+4 种基金Scientific and Technical Project of Zhejiang Province(2021C02069-1,2016C02055-7)Scientific and Technical Project of Ningbo City(2021Z002,2015C110005)Ningbo Science and Technology Plan Projects(2018A610228)Teaching and Research Project of Ningbo University(XYL19023)Collaborative Innovation Center for Zhejiang Marine High-Efficiency and Healthy Aquaculture,K.C.Wong Magna Fund in Ningbo University。
文摘Oxygen is an essential molecule for animal respiration,growth,and survival.Unlike in terrestrial environments,contamination and climate change have led to the frequent occurrence of hypoxia in aquatic environments,thus impacting aquatic animal survival.However,the adaptative mechanisms underlying fish responses to environmental hypoxia remain largely unknown.Here,we used large yellow croaker(Larimichthys crocea)and large yellow croaker fry(LYCF)cells to investigate the roles of the Hif-1α/Hsf1/Hsp70 signaling pathway in the regulation of cellular redox homeostasis,and apoptosis.We confirmed that hypoxia induced the expression of Hif-1α,Hsf1,and Hsp70 in vivo and in vitro.Genetic Hsp70 knockdown/overexpression indicated that Hsp70 was required for maintaining redox homeostasis and resisting oxidative stress in LYCF cells under hypoxic stress.Hsp70 inhibited caspase-dependent intrinsic apoptosis by maintaining normal mitochondrial membrane potential,enhancing Bcl-2 mRNA and protein expression,inhibiting Bax and caspase3 mRNA expression,and suppressing caspase-3 and caspase-9 activation.Hsp70 suppressed caspaseindependent intrinsic apoptosis by inhibiting nuclear translocation of apoptosis-inducing factor(AIF)and disturbed extrinsic apoptosis by inactivating caspase-8.Genetic knockdown/overexpression of Hif-1αand dual-luciferase reporter assay indicated that Hif-1αactivated the Hsf1 DNA promoter and enhanced Hsf1 mRNA transcription.Hsf1 enhanced Hsp70 mRNA transcription in a similar manner.In summary,the Hif-1α/Hsf1/Hsp70 signaling pathway plays an important role in regulating redox homeostasis and anti-apoptosis in L.crocea under hypoxic stress.
基金The Special Fund for Agro-Scientific Research in the Public Interest under contract No.200903046-5the National High Technology Research and Development Program of China(863program)under contract No.2006AA10A405+1 种基金the National Natural Science Foundation of China under contract No.30771663the Foundation for Innovation Research Team of Jimei University,China under contract No.2006A001
文摘An （AC）n-microsatellite-enriched library for Larimichthys crocea was constructed in this study. Primers for fifty simple sequence repeat （SSR） loci were synthesized and genotyped on 30 L. crocea individuals from Guanjingyang wild population （WP） in Fujian Province and 38 individuals from Ningbo cultured population （CP） in Zhejiang Province. Only 21 loci were successfully amplified and polymorphic in two populations. In WP, the observed heterozygosity （Ho） ranged from 0.233 to 0.900 and the expected heterozygosity （HE） ranged from 0.326 to 0.893, with an average of 7.8 alleles/locus, the polymorphism information content （PIC） ranged from 0.283 to 0.866 （mean 0.731）. In CP, the Ho ranged from 0.189 to 0.892 and the HE ranged from 0.333 to 0.800, with an average of 4.4 alleles/locus. The probability test showed significant departures from HWE in 9 and 2 of the 21 loci in WP and in CP, respectively. Deficiency of heterozygotes at four loci showed the presence of null alleles （P 〈0.01）. The PIC of 20 microsatellite loci in WP were greater than 0.50. Overall, these novel markers are potentially useful for the study of population genetics, construction of genetic linkage and quantitative trait loci maps in large yellow croaker.
基金the Agricultural Major Project of Ningbo Municipality(No.2015 C110005)the K.C.Wong Magna Fund in Ningbo University。
文摘The large yellow croaker(Larimichthys crocea)is an important mariculture fish in China.Farmed large yellow croaker undergo periods of fasting to adapt to the environment or to improve meat quality.To better understand the physiological responses of their muscle tissues to fasting stresses,we analyzed the transcriptomes and proteome s of both normally-fed and fasting fish groups and identified 7578 differentially expressed genes(DEGs)and 297 differentially expressed proteins(DEPs)among them.Gene ontology and KEGG analysis showed that the enriched biological pathways were mainly involved in various synthetic and catabolic pathways,especially the protein metabolism.Based on the omics data,nine DEGs related to muscle composition(CAN3,MYL3,and TNNC2),growth(MSTN and MYF5),autophagy(TSC2 and ULK1),and the ubiquitin proteasome pathway(PRS6 B and UCHL3)were examined using qPCR.In response to fasting stress,MYL3 and TNNC2 were significantly downregulated,while genes associated with autophagy and the ubiquitin proteasome pathway were significantly upregulated.In re sponse to fasting stres s,MYL3,TNNC2,and MYF5 positively correlated with muscle growth were significantly downregulated,while inhibiting growth MSTN and genes associated with autophagy and the ubiquitin proteasome pathways were significantly upregulated.These results clarify the effects of fasting on metabolic changes in their muscle components and growth at the molecular level.
基金Supported by the National Natural Science Foundation of China(Nos.U1205122,31172397)the Key Project of Agricultural Science and Technology of Fujian Province(No.2011N5010)the Foundation for Innovation Research Team of Jimei University(No.2010A02)
文摘The large yellow croaker, Larimichthys crocea is an important marine fish in China with a high economic value. In the last decade, the stock conservation and aquaculture industry of this species have been facing severe challenges because of wild population collapse and degeneration of important economic traits. However, genes contributing to growth and immunity in L. crocea have not been thoroughly analyzed, and available molecular markers are still not sufficient for genetic resource management and molecular selection. In this work, we sequenced the transcriptome in L. eroeea liver tissue with a Roche 454 sequencing platform and assembled the transcriptome into 93 801 transcripts. Of them, 38 856 transcripts were successfully annotated in nt, nr, Swiss-Prot, InterPro, COG, GO and KEGG databases. Based on the annotation information, 3 165 unigenes related to growth and immunity were identified. Additionally, a total of 6 391 simple sequence repeats （SSRs） were identified from the transcriptome, among which 4 498 SSRs had enough flanking regions to design primers for polymerase chain reactions （PCR）. To access the polymorphism of these markers, 30 primer pairs were randomly selected for PCR amplification and validation in 30 individuals, and 12 primer pairs （40.0%） exhibited obvious length polymorphisms. This work applied RNA-Seq to assemble and analyze a live transeriptome in L. crocea. With gene annotation and sequence information, genes related to growth and immunity were identified and massive SSR markers were developed, providing valuable genetic resources for future gene functional analysis and selective breeding ofL. eroeea.
基金Foundation item:The National Key Research and Development Program of China under contract No.2018YFC1406302the Local Science and Technology Development Project Guide by the Central Government under contract No.2017L3019+2 种基金the Development Project Guide by the Central Government under contract No.2017L3019the Technical Innovation Platform for Large Yellow Croaker under contract No.XDHT2018143Athe Major Special Projects of Fujian Province under contract No.2016NZ0001.
文摘As an economically important marine fish,the large yellow croaker Larimichthys crocea suffered from marine white spot disease caused by the ectoparasite Cryptocaryon irritans in recent years.This disease not only could result in physiological damage,but also lead to secondary bacterial invasion.Reports indicated some AMPs(antimicrobial peptides)were of antiparasitic activity to C.irritans.Hepcidin-like(Lc-HepL)was one of the significant differential expression genes excavated from the transcriptome following a challenge with C.irritans.In this study,we characterized this AMP’s bioactivity based on the levels of mRNA and protein.After challenged by C.irritans,qRT-PCR showed Lc-HepL was significantly upregulated in six tissues,including gill,muscle,liver,head kidney and spleen during theront infection,trophont falling off,and secondary bacterial invasion stages,which implicated a role Lc-HepL played in the immune defense against C.irritans and secondary bacterial infection.Recombinant Lc-HepL(rLc-HepL)was induced and purified successfully.rLc-HepL exhibited antibacterial activity to certain bacteria in a dose-and time-dependent manners.Anti-C.irritans activity was explored for the first time and found it could cause the theronts membrane rupture and contents leakage.These results provided the first evidence that Lc-HepL had strong antiparasitic activity against marine fish ectoparasites C.irritans theronts.Together,data indicated that Lc-HepL might be an important component in the innate immune system against C.irritans and has the potential to be employed in future drug development.
基金The Key Research and Development Project of Zhejiang Province under contract Nos 2016C02055-7,2017C02013。
文摘Larimichthys crocea is a marine fish species cultured in China.Short-term starvation is often applied to improve the quality of cultured L.crocea,and the expression of ghrelin in tissues of stomach,muscle,brain,intestines,liver,and kidney,involved in starvation response,under starvation conditions were studied to understand the effect of starvation on the expression of ghrelin in L.crocea juveniles.The ghrelin expression was tissue-specific,and expression was significantly higher in the stomach compared to other tissues(P<0.01).Additionally,ghrelin expression in different tissues changed along with prolongation of fasting.In the stomach,ghrelin expression levels increased gradually at the beginning of the fast,and then declined after eight days of fasting.Gene expression in the brain and intestines increased at the beginning of the fast,and then decreased with longer fasting time.Interestingly,ghrelin expression declined at the beginning of the fast,then increased with longer fasting in the kidneys and muscles.These results suggest that ghrelin is involved in starvation response in L.crocea juveniles.This study provids insights into ghrelin function and an important reference for the development of reasonable feeding strategies for L.crocea juveniles.
基金The National Key R&D Program of China under contract No.2018YFC1406302the Zhoushan Science and Technology Special Project under contract No.2020C21005+1 种基金the Zhejiang Education Department General Project under contract No.Y201942430the National Natural Science Foundation of China under contract No.41606418。
文摘Antitumor activity is one characteristic function of some certain antimicrobial peptides(AMPs)found in recent years.In the present study,we attempted to detect potential anticancer activity of a recombinant piscidin 5-like from Larimichthys crocea(r Lc-P5L)which owned widely antibacterial and strong antiparasitic activity in vitro.The light microscope observation indicated r Lc-P5L was of antitumor activity to He La cells,293 T cells and L929 cells.MTT assay showed the toxic sensitivity of r Lc-P5L to three tumor cell strains was 293 T>L929>He La.Scanning electron microscope(SEM)results showed r Lc-P5L behaved like a lytic peptide to cause damage on cells membrane of L929 cells by forming globular clusters,even pores at 60μmol/L,or degrading membrane to make it completely lose cytoskeleton structure at 80μmol/L;r Lc-P5L treatment also resulted in DNA degradation.Fluorescence observation results indicated r Lc-P5L could cause L929 cells at least two obvious changes:one is nucleus,nuclear chromatin condensed in the margin,nuclear volume became smaller and shrank to be out of shape,or lysed to be debris;the other is cytoskeleton,they became disordered and polarized to make cells atrophic shapes,or even lysed to be debris.In summary,r Lc-P5L owned potential anticancer activity causing membrane structure damage and genome DNA degradation.Interestingly,treatment with different concentration of r Lc-P5L seemingly caused the similar but different changes,whether it indeed gave rise to cancer cells diverse death way,the further studies should be performed,and the detailed mechanisms were still need further explored.
基金the Natural Science Foun-dation of Fujian Province(No.2018J01450)the National Natural Science Foundation of China(No.U1705231)+2 种基金the National Marine Fisheries Industrial Technology System Post Scientist Project(No.CARS-47-G04)the Scientific Research Foundation of Jimei University(No.ZQ2019046)the Central Public-Interest Scientific Institution Basal Re-search&Key Laboratory of Sustainable Development of Marine Fisheries,Ministry of Agriculture,P.R.China,CAFS(No.2017HY-XKQ01).
文摘Elovl4 is a fatty acyl elongase which participates in the biosynthesis of long-chain polyunsaturated fatty acids(LCPUFA).Herein we isolated and functionally characterized the elovl4a gene in the large yellow croaker Larimichthys crocea,and investigated the regulatory effects of transcription factors Hnf4α,Lxrα,Pparαon the activity of elovl4a promoter.Tissue expression pattern revealed that elovl4a was widely expressed in several tissues and predominantly in brain.Heterologous expression in yeast showed that L.crocea Elovl4a could effectively elongate both C18 and C20 PUFA substrates to C22 fatty acid.In addition,the affinity of large yellow croaker Elovl4a to n-6 series fatty acids is weak and cannot prolong 18:2n-6 and 18:3n-6.Moreover,L.crocea elovl4a reporter activities were elevated by 1.31-,1.39-and 1.48-fold via over-expression of Lxrα,Pparαand Hnf4α,respectively.The above findings can enrich the knowledge of biosynthesis pathway of LC-PUFA in the large yellow croaker and contribute to elucidate the LC-PUFA anabolism mechanism in fish.
基金Sponsored by Research Program of Zhejiang Provincial Department of Education(Y201328411)Program of Zhejiang Provincial Department of Science and Technology(2009C22007)
文摘To ascertain the nutrient components and nutritive quality of the flesh of big yellow croaker in three culture conditions （ traditional cage, offshore cage and cage-free）, basic nutritional components,amino acid,fatty acid and mineral elements were determined. The results indicated that crude protein in flesh of the big yellow croaker in cage-free culture was higher than that in offshore cage and much higher than that in traditional cage （ P 〈0.05）. Crude fat of the croaker cul- tured in the traditional cage was twice as high as that in cage-free culture, while that in the offshore cage was in the middle. Proline content in the cage-free culture was much higher than that in the offshore cage, and also than that in the traditional cage （P 〈 0.05 ）. There was no significant difference in the content of alanine, methionine and tryptophan （ P 〈 0.05）. Contents of other amino acids had no significant difference between the cage-free culture and offshore cage, but were much lower in the traditional cage （P 〈 0.05 ）. Top six fatty acids were 9-Hexadecenoic acid, palmitic acid,9-Octadecenoic acid, Octadecauoic acid, DHA and EPA. The palmitic acid content was the highest in cage-free culture and in traditional cage, 9-Octadeeenoic acid content was the highest in offshore cage. Content of unsaturat- ed fatty acids in cage-free culture, offshore cage and traditional cage was 63.60,66.32,57.67, respectively, and polyunsaturated fatty acid was 29.10, 28.57, and 24.40. Content of DHA in cage-free culture was significantly higher than that in the offshore and traditional cage. Content of zinc had no significant difference in three culture models. Content of phosphorus had no significant difference between that in cage-free culture and offshore cage, was lower in the traditional cage. The cage-free cultured croakers had the highest content of calcium and phosphorus. Content of selenium was about the same between the offshore cage and the traditional cage stocking, higher in the cage-free culture. This research has considerable application value for identifying quality and sources of the big yellow croakers.
基金supported by Program for Innovation Team Building at Institutions of Higher Education in Chongqing(KJTD201325)the Program for Innovative Research Team in Chongqing University of Education(No.KYC-cxtd03-20141002)
文摘The aim of the present study was to determine the preventive effects of the polysaccharide of Larimichthys crocea swim bladder(PLCSB) on CCl4-induced hepatic damage in ICR mice.The in vitro preventive effects of PLCSB on CCl4-induced liver cytotoxic effect were evaluated in BRL 3A rat liver cells using the MTT assay.The serum levels of AST,ALT,and LDH in mice were determined using commercially available kits.The levels of IL-6,IL-12,TNF-α,and IFN-γ were determined using ELISA kits.The pathological analysis of hepatic tissues was performed with H and E staining,and the gene and protein expressions were determined by RT-PCR and Western blotting,respectively.PLCSB(20 μg·m L-1) could increase the growth of BRL 3A rat liver cells treated with CCl4.The serum levels of AST,ALT,and LDH were significantly decreased when the mice were treated with two doses of PLCSB,compared with the control mice(P < 0.05).PLCSB-treated groups also showed reduced levels of the serum pro-inflammatory cytokines IL-6,IL-12,TNF-α,and IFN-γ.PLCSB could decrease the liver weight,compared to the CCl4-treated control mice.The histopathology sections of liver tissues in the 100 mg·kg-1 PLCSB group indicated that the animals were recovered well from CCl4 damage,but the 50 mg·kg-1 PLCSB group showed necrosis to a more serious extent.The 100 mg·kg-1 PLCSB group showed significantly decreased mR NA and protein expression levels of NF-κB,i NOS,and COX-2,and increased expression of IκB-α compared with the CCl4-treated control group.In conclusion,PLCSB prevented from CCl4-induced hepatic damage in vivo.
基金Aeknowleflgements Thanks to Chen Qing-kai for rearing fish families. This study was supported by Special Fund for Agro-scientific Research in the Public Interest （200903029-4）, the National Natural Science Foundation of China （30771663）, Key Project of Fujian Provincial Department of Science ＆ Technology （2010N0021）, the Foundation for Innovation Research Team of Jimei University, China（2010A002） and the Open Foundation from Ocean Fishery Science and Technology in the Most Important Subjects of Zhejiang （20100203）.
文摘The large yellow croaker Larimichthys crocea is one of the most important fish species in China. To estimate the re- productive success of breeders, three independent full-factorial crosses were created and the fins of breeders and progenies were sampled for microsatellite analysis. Out of 959 offspring from three sets, 99.6% were assigned to their parents using 6-7 mi- crosatellite markers. In all crosses, some parent pairs produced a large number of offspring and some parent pairs did not produce any offspring. The contributions of male or female parents were unequal, ranging from 1.0-89.3% across the three sets. The loss of putative Ne was 69.6% in set 1, 31.2% in set 2 and 57.6% in set 3. These results suggest that the unequal contribution of parents is universal in artificial breeding ofL. crocea, especially in a small population, and this should be taken into account in hatcheries or when releasing animals for resource enhancement
基金This work was supported by National Key Research and Development Program of China under Grant No.2022YFD2401001National Natural Science Foundation of China under Grant No.U1905204+2 种基金China Agriculture Research System of MOF and MARA under Grant No.CARS-47Fujian Science and Technology Department under Grant No.2021N5008Institute of Oceanology of Fuzhou(2021F02).
文摘In mammals,mitofusin 2(MFN2)is involved in mitochondrial fusion,and suppresses the virus-induced RIG-I-like receptor(RLR)signaling pathway.However,little is known about the function of MFN2 in non-mammalian species.In the present study,we cloned an MFN2 ortholog(LcMFN2)in large yellow croaker(Larimichthys crocea).Phylogenetic analysis showed that MFN2 emerged after the divergence of amphioxus and vertebrates.The protein sequences of MFN2 were well conserved from fsh to mammals.LcMFN2 was expressed in all the tissues/organs examined at diferent levels,and its expression was upregulated in response to poly(I:C)stimulation.Overexpression of LcMFN2 inhibited MAVS-induced type I interferon(IFN)promoter activation and antiviral gene expression.In contrast,knockdown of endogenous LcMFN2 enhanced poly(I:C)induced production of type I IFNs.Additionally,LcMFN2 enhanced K48-linked polyubiquitination of MAVS,promoting its degradation.Also,overexpression of LcMFN2 impaired the cellular antiviral response,as evidenced by the increased expression of viral genes and more severe cytopathic efects(CPE)in cells infected with spring viremia of carp virus(SVCV).These results indicated that LcMFN2 inhibited type I IFN response by degrading MAVS,suggesting its negative regulatory role in cellular antiviral response.Therefore,our study sheds a new light on the regulatory mechanisms of the cellular antiviral response in teleosts.
