Spinyhead Croaker Research Articles

Comparative transcriptome analysis reveals sex bias in expression patterns of genes related to sex steroids and immunity in the skin of spinyhead croaker Collichthys lucidus.

Fish skin is the first barrier against external invasion, and also an important interface for communication between males and females during reproduction. Nonetheless, sexual dimorphism in the physiology of fish skins is still poorly understood. Herein, transcriptomes of skin were comparatively analysed between males and females in spinyhead croaker, Collichthys lucidus. Totally, 170 differentially expressed genes (DEG) were detected, including 79 female-biased genes and 91 male-biased genes. Gene ontology (GO) annotation items of the DEGs were mainly enriched in biological process items (86.2%), including regulation of biological processes, responses to chemical and biological stimuli, transport and secretion, movement, immune response, tissue development, etc. In KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway enrichment analysis, the male-biased genes were enriched in pathways including those related to immunity such as the TNF signalling pathway and IL-17 signalling pathway, whereas the female-biased genes were enriched in pathways including those related to female steroids such as ovarian steroidogenesis and oestrogen signalling pathway. In addition, odf3 was found to be a male-specific expression gene, being a candidate marker for phenotypic sex. Thus, the sexual difference in gene expression in fish skin in spawning season was uncovered by transcriptome analysis for the first time, providing new insights into sexual dimorphism in the physiology and functions of fish skin.

Spinyhead Croaker Germ Cells Gene dnd Visualizes Primordial Germ Cells in Medaka.

HighlightsCldnd was specifically expressed in adult gonads.Cldnd RNA was enriched in spermatogonia and early oocytes and decreased with gametogenesis.Gfp-Cldnd 3′ UTR mRNA could identify primordial germ cells in medaka.Spinyhead croaker (Collichthys lucidus) is an economically important fish suffering from population decline caused by overfishing and habitat destruction. Researches on the development of primordial germ cell (PGC) and reproduction biology were an emergency for the long-term conservation of the involved species. Dead end (dnd) gene plays an indispensable role in PGC specification, maintenance, and development. In the current study, we report the cloning and expression patterns of dnd in C. lucidus (Cldnd). RT-PCR analysis revealed that Cldnd was specifically expressed in both sexual gonads. In the ovary, Cldnd RNA was uniformly distributed in the oocytes and abundant in oogonia, and gradually decreased with oogenesis. A similar expression pattern was also detected in testis. Dual fluorescent in situ hybridization of Cldnd and Clvasa demonstrated that they almost had the same distribution except in oocytes at stage I, in which the vasa RNA aggregated into some particles. Furthermore, Cldnd 3′ UTR was sufficient to guide the Green Fluorescent Protein (GFP) specifically and stably expressed in the PGCs of medaka. These findings offer insight into that Cldnd is an evolutionarily conserved germline-specific gene and even a potential candidate for PGC manipulation in C. lucidus.

Life history of spinyhead croaker Collichthys lucidus (Sciaenidae) differentiated among populations from Chinese coastal waters

Otolith microchemistry provides valuable information about the environmental history of individual fish, but few studies have considered the influence of population or stock on life history traits. This study used the Sr/Ca ratio as an index of habitat use of spinyhead croakerCollichthys lucidusfrom 2 different populations (northern and southern China populations), including lifetime otolith microchemistry profiles (n = 63 fish) and determinations of natal habitat selection (n = 352 fish). The otolith data revealed 3 life history patterns inC. lucidus: Pattern 1 individuals spent most of their lifetime (>95%) in mesohaline waters (Sr/Ca range: 3-7 mmol mol-1), particularly during early growth; Pattern 2 individuals migrated from hyperhaline waters (Sr/Ca ≥ 7 mmol mol-1) to mesohaline waters; and Pattern 3 individuals migrated from oligohaline waters (Sr/Ca < 3 mmol mol-1) to mesohaline waters. Pattern 2 and Pattern 3 were specific to the northern and southern China populations, respectively, and Pattern 1 was shared by the 2 populations. The otolith core Sr/Ca ratios showed that mostC. lucidusselected mesohaline waters as their natal habitat (i.e. 90.6% of 352 total individuals); 12.5% (19 of 152) selected hyperhaline waters and fell within the northern China population, and 7% (14 of 200) selected oligohaline waters and fell within the southern China population. These results indicate that life history diversity exists in this species, as the pattern of habitat use largely differed between the northern and southern China populations. The overall findings highlight that the behavior of different populations should be considered when determining fish migration histories.

Genome‐wide identification and transcriptome‐based expression profiling of the Sox gene family in the spinyhead croaker (Collichthys lucidus)

Sox genes encode transcription factors with a high-mobility group (HMG) box, playing critical roles in the initiation and maintenance of a variety of developmental processes, such as sex determination and differentiation. In the present study, we identified 26 Sox genes in the genome of spinyhead croaker Collichthys lucidus (Richardson, 1844) with homology-based analysis of the HMG box. The transcriptome-based expression profiles revealed that the expression of the Sox gene in gonads began to differ between sexes when the body length was 2.74 ± 0.24 cm. At that time, three Sox genes (Sox11b, Sox8a and Sox19) were significantly upregulated, accompanied by the downregulation of 12 Sox genes in the ovary, and six Sox genes were temporarily significantly upregulated in the testis. Afterwards, the expression profile of Sox genes changed only with a small amplitude in both the ovary and testis. For adult tissues, huge differences were observed in the expression profiles of Sox genes between ovaries and testes, as well as small differences in somatic tissues between sexes. These results provide clues to further decipher the role of Sox genes in the processes of sex determination and differentiation in spinyhead croaker and other teleosts.

Whole-Genome Sequencing and Genome-Wide Studies of Spiny Head Croaker (Collichthys lucidus) Reveals Potential Insights for Well-Developed Otoliths in the Family Sciaenidae.

Spiny head croaker (Collichthys lucidus), belonging to the family Sciaenidae, is a small economic fish with a main distribution in the coastal waters of Northwestern Pacific. Here, we constructed a nonredundant chromosome-level genome assembly of spiny head croaker and also made genome-wide investigations on genome evolution and gene families related to otolith development. A primary genome assembly of 811.23 Mb, with a contig N50 of 74.92 kb, was generated by a combination of 49.12-Gb Illumina clean reads and 35.24 Gb of PacBio long reads. Contigs of this draft assembly were further anchored into chromosomes by integration with additional 185.33-Gb Hi-C data, resulting in a high-quality chromosome-level genome assembly of 817.24 Mb, with an improved scaffold N50 of 26.58 Mb. Based on our phylogenetic analysis, we observed that C. lucidus is much closer to Larimichthys crocea than Miichthys miiuy. We also predicted that many gene families were significantly expanded (p-value <0.05) in spiny head croaker; among them, some are associated with “calcium signaling pathway” and potential “inner ear functions.” In addition, we identified some otolith-related genes (such as otol1a that encodes Otolin-1a) with critical deletions or mutations, suggesting possible molecular mechanisms for well-developed otoliths in the family Sciaenidae.

An Integrated Approach to Determine the Stock Structure of Spinyhead Croaker Collichthys lucidus (Sciaenidae) in Chinese Coastal Waters

An integrated approach including analyses of different biological traits is a proven and powerful tool used to assess the population structures of fish species, which is vital for fishery stock conservation and management of wild resources. This study evaluates the use of three natural tags (i.e., microsatellites, parasites, and otolith nucleus chemistry) in order to describe the population structure of the spinyhead croaker, Collichthys lucidus, in the coastal waters of China, on evolutionary and ecological time scales. Spinyhead croaker was assigned with 86% accuracy to its regional origin (northern China vs. southern China) using all three natural tags. Accuracy decreased when incorporating only one type of natural tag (genetics: 19–69%; parasites: 30–60%; otolith chemistry: 51–86%) or when assigning the fish to a fine scale (sea areas: 44–64%; sampled estuaries: 19–66%). However, the overall accuracy assignment improved slightly compared with otolith chemistry (estuaries: 55 vs. 51%; sea areas: 66 vs. 64%; regions: 86 vs. 86%). Three natural tags and integrated results show that C. lucidus in Chinese coastal waters can be separated into distinct northern and southern Chinese stocks. Finally, this information should promote the development of effective conservation strategies and integrated fisheries management plans for this commercially important species.

Development of sex-specific markers in Spinyhead croaker, Collichthys lucidus

Spinyhead croaker (Collichthys lucidus) is a commercially important marine sciaenid fish, being found diverging into northern and southern lineages. A sex diagnostic marker has been developed in the croaker, but its application scope was limited in the populations belonging to southern lineage. In present study, novel diagnostic markers were developed to break the limitation. Resequencing was performed with samples from Zhousan (ZS) population and Ningde (ND), representing northern and southern lineages respectively. Based on the resulted data, the candidate sexually dimorphic SNPs and InDels were screened out, and their genomic distributions were visualized. For ZS population, 52,621 SNPs and 8594 InDels were screened out, mainly distributing on Chr.1 and Chr.8 along almost entire chromosome. For ND population, 2094 SNPs and 350 InDels were screened out, mainly clustering within a 0.55-Mb segment from 4.31–4.86 Mb on Chr.1 and a 1.58-Mb segment from 27.12–28.70 Mb on Chr.7. Based on a common male-specific delete on Chr.1, PCR primer pair CMS-1 was designed and demonstrated applicable to all four tested populations covering both lineages; while based on a male-specific delete of Chr.8 for ZS population, PCR primer pair ZS-8 was designed and demonstrated applicable only to the northern populations but not the southern populations. The novel sex-specific markers developed in the present study extend the toolkit for both sex control breeding and basic study on sex determination in the croaker. Besides, different number and location of sexually dimorphic SNPs/InDels between two lineages provide new insights in sex chromosome evolution in spinyhead croaker.

De novo transcriptome assembly of four organs of Collichthys lucidus and identification of genes involved in sex determination and reproduction.

The spinyhead croaker (Collichthys lucidus) is a commercially important fish species, which is mainly distributed in the coastal regions of China. However, little is known about the molecular regulatory mechanism underlying reproduction in C. lucidus. A de novo transcriptome assembly in brain, liver, ovary and testis tissues of C. lucidus was performed. Illumina sequencing generated 60,322,004, 57,044,284, 60,867,978 and 57,087,688 clean reads from brain, liver, ovary and testis tissues of C. lucidus, respectively. Totally, 131,168 unigenes with an average length of 644 bp and an N50 value of 1033 bp were assembled. In addition, 1288 genes were differentially expressed between ovary and testis, including 442 up-regulated and 846 down-regulated in ovary. Functional analysis revealed that the differentially expressed genes between ovary and testis were mainly involved in the function of sexual reproduction, sex differentiation, development of primary male sexual characteristics, female gamete generation, and male sex differentiation. A number of genes which might be involved in the regulation of reproduction and sex determination were found, including HYAL and SYCP3 and BMP15. Furthermore, 35,476 simple sequence repeats (SSRs) were identified in this transcriptome dataset, which would contribute to further genetic and mechanism researches. De novo transcriptome sequencing analysis of four organs of C. lucidus provides rich resources for understanding the mechanism of reproductive development of C. lucidus and further investigation of the molecular regulation of sex determination and reproduction of C. lucidus.

Development of a PCR-based genetic sex identification method in spinyhead croaker (Collichthys lucidus)

Spinyhead croaker (Collichthys lucidus) is a commercially important marine sciaenid fish, exhibiting sexual dimorphism in growth, and being of a special multiple sex system X1X1X2X2/X1X2Y. In the present study, we developed the first PCR-based method for identifying the genetic sex in spinyhead croaker with a next generation sequencing (NGS)-based approach. After whole genome resequencing was performed on 10 individuals and 2 pools for each sex, approximately 404 G clean data and 2,857,406 high quality SNPs were obtained. With genome-wide association analysis (GWAS), 420 SNPs on 8 chromosomes were identified as sex significantly associated loci. Most of the sex associated loci aggregated on Chr. 1 and Chr. 7, suggesting that they might be the X1 and X2 chromosome. On the segment of Chr. 7 that aggregated sex associated SNPs, three candidate male specific deletions were detected by comparing the sequencing depth between sexes. Based on one of the putative male specific deletions (68 bp), a pair of primers for sex identification were designed and validated with PCR. The primers were applicable to ND and JR populations belonging to the southern group, but not to ZS population belonging the northern group. The methods for sex identification developed in the present study along with its limited versatility provided a necessary tool and important clues for deciphering the sex determination mechanism in spinyhead croaker.

Study of Spinyhead Croaker (Collichthys lucidus) Fat Content Forecasting Model Based on Electronic Nose and Non-linear Data Resolution Model

In this paper, spinyhead croaker (Collichthys lucidus) fat content forecasting model using electronic nose (e-nose) and non-linear stochastic resonance (SR) has been studied. Spinyhead croaker samples are stored at 4 °C temperature. Physical/chemical indexes (firmness, total volatile basic nitrogen (TVB-N), pH, and fat content) are examined to provide quality references for e-nose. E-nose responses are treated by principal component analysis (PCA), bistable SR, and double-layered cascaded serial SR (DCSSR). SR and DCSSR SNR maximal (SNR-Max) values discriminate croakers clearly. Multi-variables regressions (MVR) are conducted between physical/chemical indexes and SR/DCSSR SNR-Max values. MVR results demonstrate that DCSSR feature values have more significant linearity relation with physical/chemical indexes. Spinyhead croaker fat content forecasting model is developed via linear fitting regression on SR SNR-Max values. Validating experimental results demonstrate that the developed model has good accuracy.

Evolutionarily conserved vasa identifies embryonic and gonadal germ cells in spinyhead croaker Collichthys lucidus.

In this study, a 2198 bp full-length cDNA of spinyhead croaker Collichthys lucidus vasa gene encoding 616 amino-acid residues was obtained. Multiple alignment revealed that C. lucidus vasa has eight conserved characteristic motifs of the DEAD box protein family and has the highest identity to large yellow croaker Larimichthys croceas. Reverse-transcription (RT)-PCR and Western blot analyses indicated that the vasa messenger (m)RNA and Vasa protein are specifically expressed in the gonads in both sexes. In situ hybridisation (ISH) demonstrated that vasa RNA is exclusively detected in the germ cells in C. lucidus gonads and its temporospatial expression reveals a dynamic pattern during oogenesis. Surprisingly, C. lucidus vasa 3'UTR can direct stable and specific GFP expression in the primordial germ cells (PGC) of medaka Oryzias latipes embryos. Taken together, these results suggest that because C. lucidus vasa expression delineates critical stages of oogenesis, it may be a useful molecular marker for the identification of gonadal germ cells, facilitating the isolation and utilization of germ cells in future study.

Fine‐scale population structure of Collichtys lucidus populations inferred from microsatellite markers

The spinyhead croaker Collichthys lucidus (Richardson) is a small sciaenid species distributed along the inshore waters of northwestern Pacific Ocean, and now has been listed as Key Protected Commercial Sources of Aquatic Animals and Plants in China. To delineate stock boundaries and inform conservation policy for its management, samples were collected from eight locations across the Chinese coastal waters and analyzed at nine microsatellite loci. C. lucidus populations showed low genetic diversity (expected heterozygosity = 0.445–0.542; observed heterozygosity = 0.392–0.539; Polymorphism Information Content = 0.268–0.684). Strong genetic fdifferentiation (Fst = 0.065–0.510, all significant after Bonferroni correction) among all populations and high levels of self-recruitment (89.2%–91.5%) were observed, which suggested limited genetic exchange for this species. Clustering results of discriminant analysis of principal components and STRUCTURE found strong support for obvious genetic clusters (populations FZ, XM and SZ vs. populations SH, YRE, ZS, WZ and ND). The results of the present study not only supported the phylogeographic pattern of north-south differentiation, but also suggested that C. lucidus populations may be predominantly sustained by self-replenishment rather than by recruitment from distant populations.

Cytogenetic characterization and description of an X1X1X2X2/X1X2Y sex chromosome system in Collichthys lucidus (Richardson, 1844)

The chromosomes of spinyhead croaker Collichthys lucidus (Richardson, 1844) were characterized for the first time by fluorescence staining, self genomic in situ hybridization (self-GISH), and multicolor fluorescence in situ hybridization (FISH) with 18S rDNA, 5S rDNA and telomeric sequence probes. The female karyotype has exclusively 24 pairs of acrocentric chromosomes (2n=48a, NF=48), while the male one consists of 22 pairs of acrocentric chromosomes, 2 monosomic acrocentric chromosomes and a metacentric chromosome (2n=1m+46a, NF=48). The difference between female and male karyotypes indicates the presence of a sex chromosome of X1X1X2X2/X1X2Y type, where Y is the unique metacentric chromosome in the male karyotype. As revealed by FISH, 5S rDNA and 18S rDNA sites were mapped at syntenic position of the largest acrocentric chromosome (X1), and the short arms of the Y chromosome as well. An X1-chromosome specific interstitial telomeric signal (ITS) was detected overlapping the 5S rDNA sites. In addition, self-GISH revealed that the repetitive DNAs accumulated on all the putative sex chromosome. Chromosome fusion accompanied by a partial deletion in the ancestral karyotype (2n=48a) is hypothesized for the origin of such multiple sex chromosome system. The present study, as the first description of differentiated sex chromosome in family Sciaenidae, will give clues to the studies on the sex chromosome of other Sciaenids.

Development and characterization of SNP derived from spinyhead croaker (Collichthys lucidus) by RNA-seq

A novel set of SNP markers were detected from RNA-seq data of spinyhead croaker (Collichthys lucidus). Totally 49,617,970 clean reads and 78,671 putative SNPs were obtained. Among them, 40 SNPs were randomly selected to perform validation in 30 wild individuals of C. lucidus captured from 3 geographic stocks in the coast of China. Eventually, 29 SNP loci were successfully validated and showed bi-allelic polymorphism in genotyping by Sequenom MassARRAY. These novel SNPs provided useful genomic resources for C. lucidus, especially in population genetics, molecular assisted selection and conservation studies.

Spinyhead croaker (Collichthys lucidus) quality determination using multi-walled carbon nanotubes gas-ionization sensor array

In this paper, a novel spinyhead croaker (Collichthys lucidus) quality analysis method using multi-walled carbon nanotubes (MWCNT) gas-ionization sensor array and stochastic resonance (SR) was proposed. The sensor array consisted of two MWCNT gas sensors. One was used as anodic sensor, and the other as cathodal sensor. Sensor array response to croakers under different storage time was measured. Responses of two MWCNT sensors were processed by SR. Signal-to-noise ratio (SNR) was utilized to characterize croaker quality. Meanwhile, total volatile basic nitrogen index was examined to provide quality reference to cold stored croaker samples in each day. Croaker quality was determined by SNR maximal values and its located noise intensities.

RESEARCH ARTICLE Molecular cloning and expression analysis of a matrix Gla protein gene in the spinyhead croaker, Collichthys lucidus.

The matrix Gla (gamma-carboxyglutamic acid-rich) protein (MGP), a vitamin K-dependent and Gla-containing protein, is a calcification inhibitor that mainly functions in tissue calcification and mineralization. In this study, we obtained the complete cDNA sequence of MGP from the spinyhead croaker (Collichthys lucidus), which we named Cl-MGP. Cl-MGP was 923 bp long with a 384-bp open reading fragment that encoded 127 amino acids. The predicted MGP protein sequence contained a 19-residue hydrophobic signal peptide, suggesting that it possesses secretory characteristics. The Gla domain and the invariant unit ErraEtCedyspC, which has been identified in all known vitamin K-dependent vertebrate proteins, were highly conserved in Cl-MGP, suggesting that it uses the same mechanism to function as the known proteins. An alignment analysis revealed that Cl-MGP had the highest identity with Larimichthys crocea (93%), which had lost five amino acid residues in the C-terminal. A quantitative real-time polymerase chain reaction revealed that Cl-MGP expression was highest in the gill, followed by the cholecyst and spleen, with almost no expression in the blood, muscle, or testes. The high Cl-MGP expression in the gill is similar to that observed in other fish species, but the relatively high expression found in the cholecyst and spleen is not seen in all species. Future studies should investigate the tissue distributions of both mRNA and proteins in different species, in order to understand the function and evolution of MGP in different species.

Characterization, molecular cloning, and expression analysis of Ecsit in the spinyhead croaker, Collichthys lucidus.

Evolutionarily conserved signaling intermediate in Toll pathways (Ecsit) is reported to play an essential role in innate immunity, embryogenesis, and assembly or stability of the mitochondrial complex I. In this study, the full-length cDNA of Ecsit was cloned from the spinyhead croaker Collichthys lucidus based on the expressed sequence tags from our cDNA library constructed using the SMART technique. The cDNA was 1669 bp long, including a 5'-terminal untranslated region (UTR) of 121 bp, a 3'-terminal UTR of 183 bp, and an open reading frame of 1365 bp encoding a 454-amino acid polypeptide. The estimated molecular weight of C. lucidus Ecsit (ClEcsit) was 52.50 kDa with an isoelectric point of 6.14, and contained a typical Ecsit domain that is conserved in other Ecsits. Multiple alignment of ClEcsit with other selected Ecsits suggested that some amino acid residues were highly conserved. Phylogenetic analysis indicated that ClEcsit was more similar to its identities in Sciaenidae and grouped with Ecsits from other Perciformes. Quantitative real-time reverse transcription PCR analysis revealed broad expression of ClEcsit and the transcript was strongly expressed in the gill and weakly expressed in other tissues.

Molecular cloning and gene expression analysis of cystatin C-like proteins in spinyhead croaker Collichthys lucidus.

Cystatins are natural tight-binding reversible inhibitors of cysteine proteases. In this study, a cDNA library was constructed from Collichthys lucidus using the SMART technique. A complete cDNA sequence with high identity to the conserved sequence of the cystatin C gene was cloned from the library using EST analysis and rapid amplification of cDNA ends (RACE), then subjected to further investigation. The full-length cDNA of cystatin C from C. lucidus (Clcys) was 699 bp long, including a 5'-terminal untranslated region (5'-UTR) of 52 bp, a 3'-UTR of 290 bp, and an open-reading frame of 357 bp. The gene encoded a polypeptide of 118 amino acids, constituting a predicted molecular weight of 12.875 kDa and a theoretical isoelectric point of 8.81. The amino acid sequence of Clcys possessed typical features of type II cystatins and had the highest identity with cystatin C of Pseudosciaena crocea (89%); therefore, it clustered with the cystatin C group in the UPGMA phylogenetic tree. Quantitative real-time reverse transcription analysis revealed that the highest expression was found in the kidney, followed by the liver, heart, and testis, with the lowest expression in muscle. Interestingly, Clcys had relatively low identity with cystatin C genes from other fish and mammals, and its expression pattern did not possess features of a housekeeping gene. Based on these findings, we suspect that the classification of cystatins in fish is somewhat confusing, and the identification of more cystatin gene sequences is needed before a definite conclusion can be drawn.

Molecular cloning and mRNA expression of a hepcidin gene from the spinyhead croaker, Collichthys lucidus.

Antimicrobial peptides are important components that participate in host innate immune activities and play crucial roles in host defense against microbial invasion. Hepcidin is an antimicrobial peptide and iron-regulatory molecule that primarily functions in the liver. In the present study, we first obtained a full-length cDNA sequence of hepcidin and its corresponding genomic DNA sequence from Collichthys lucidus using RT-PCR and rapid amplification of cDNA ends (RACE), and then analyzed these sequences using bioinformatics software. The results showed that C. lucidus hepcidin (CL-hepc) possesses two introns and three exons in the genomic DNA, with a length of 816 bp. The open reading frame was 264 bp, encoding an 87 amino acid peptide, and with high similarity (88.89%) to 83416593 Larimichthys crocea (ABC18307) and relatively low similarity (47.73%) to 158358729 L. crocea (ABY84845.1). The pre-peptide contained a signal peptide (28 amino acids), a prodomain (34 amino acids), and a mature peptide (25 amino acids). The predicted 25 amino acid hepcidin mature peptide included 8 conserved cysteine residues. Quantitative real-time reverse transcription-PCR analysis revealed specific expression patterns of CL-hepc, with the highest expression observed in the liver, relatively low expression observed in the gill and spleen, and almost no expression detected in other tissues analyzed. In conclusion, we identified a hepcidin from C. lucidus that has common expression patterns with other hepcidins. However, as this hepcidin is inconsistent with two other hepcidins from L. crocea in terms of the phylogenetic tree, the presence of another hepcidin gene warrants further investigation.

Estuarine dependency in Collichthys lucidus of the Yangtze River Estuary as revealed by the environmental signature of otolith strontium and calcium

This paper represents the first effort to study the environmental Sr and Ca signatures in otolith of spinyhead croaker Collichthys lucidus, an economically important sciaenid fish in China, by means of Electron Probe Microanalysis. Results from life-history transect analyses and X-ray intensity maps suggested that all croakers in this study collected from the Yangtze River Estuary were estuarine-resident individuals, in contrast to previous reports that C. lucidus is a marine fish. However, the high Sr core suggested that the stimulation of relatively high salinity water seemed to be required for spawning, embryogenesis and early larval growth of C. lucidus.