Molecular Characterization and Antibacterial Activity of a G-Type Lysozyme in Yellow Drum(Nibea albiflora)

Lysozyme(EC3.2.1.17)plays an important role in the immune response;as a nonspecific immune factor,it can resist causative agents.Lysozyme can be divided into c-type and g-type in fish.In a previous study,through genome-wide association analysis,the g-type lysozyme gene,which is named NaLyg in yellow drum(Nibea albiflora),was found to be a key candidate gene for disease resistance in response to Vibrio harveyi infection.The cDNA of NaLyg was 1025 bp,including four exons and three introns,and its open reading frame(ORF)had a full-length of 582 bp,encoding 193 amino acids.NaLyg was found to be conserved during evolution through bioinformatic analyses.The NaLyg protein possessed a sugar binding domain and three catalytic sites,including Glu71,Asp84 and Asp101.Quantitative qRT-PCR results confirmed that NaLyg gene mRNA was visibly increased after V.harveyi infection.The NaLyg protein purified by prokaryotic expression killed some gram-negative bacterial pathogens by inducing cell wall destruction,including V.harveyi,Aeromonas hydrophila and Edwardsiella tarda.Moreover,the NaLyg protein killed two gram-positive bacteria,Bacillus subtilis and Staphylococcus aureus.Taken together,the experimental results suggested that the NaLyg protein of N.albiflora played an important role in fighting bacterial infections.

Effects of Poria cocos polysaccharide on growth performance,physiological parameters,and lipid metabolism of spotted sea bass Lateolabrax maculatus

The aquaculture industry has developed significantly over the past few decades and has had a substantial impact on the global food supply and marine fisheries resources.However,some problems arise behind the scenes due to excessive intensive farming,such as slow animal growth,frequent disease,and lipid metabolism disorders.These problems have limited the sustainable development of the aquaculture industry,and a continuable solution is required.The use of fungal polysaccharide appears to provide a solution to these problems.Therefore,different supplemented levels of Poria cocos polysaccharide(PCP)(0,0.4,0.8,1.2,1.6,and 2.0 g/kg,respectively)were fed to spotted sea bass(Lateolabrax maculatus)in similar size(30.28±0.18 g)in current study.The effects of PCP on growth,physiological parameters,and lipid metabolism of spotted sea bass were investigated after a 4-week rearing period.Results showed,fish with PCP intake presented a significantly higher weight gain,specific growth rate,and a significantly lower feed conversion ratio.Significantly higher trypsin activity in liver and intestine were observed in fish with PCP intake.The superoxide dismutase activity in serum and liver of fish with PCP intake were significantly improved,while significantly higher serum total antioxidant capacity and hepatic catalase activity were also observed.However,no significant differences in lysozyme and alkaline phosphatase activity were evident among groups.Fish with PCP intake showed a significantly lower total cholesterol,but no noteworthy change in triglyceride and lipid-metabolismrelated genes expression were observed among groups.Results indicated that intake of PCP has a positive effect on growth and antioxidant capacity of spotted sea bass,but seems to have a limited effect on the non-specific immunity and lipid metabolism of spotted sea bass.Based on the regression analysis results,1.4 g/kg of PCP is the optimal dose for spotted sea bass in size(30.28±0.18 g).

Analytical and Experimental Research on Wave Scattering by an Open-Type Rectangular Breakwater with Horizontal Perforated Plates

This study proposes a novel open-type rectangular breakwater combined with horizontal perforated plates on both sides to enhance the sheltering effect of the rectangular box-type breakwaters against longer waves.The hydrodynamic characteristics of this breakwater are analyzed through analytical potential solutions and experimental tests.The quadratic pressure drop conditions are exerted on the horizontal perforated plates to facilitate assessing the effect of wave height on the dissipated wave energy of breakwater through the analytical solution.The hydrodynamic quantities of the breakwater,including the reflection,transmission,and energyloss coefficients,together with vertical and horizontal wave forces,are calculated using the velocity potential decomposition method as well as an iterative algorithm.Furthermore,the reflection and transmission coefficients of the breakwater are measured by conducting experimental tests at various wave periods,wave heights,and both porosities and widths of the horizontal perforated plates.The analytical predicted results demonstrate good agreement with the iterative boundary element method solution and measured data.The influences of variable incident waves and structure parameters on the hydrodynamic characteristics of the breakwater are investigated through further calculations based on analytical solutions.Results indicate that horizontal perforated plates placed on the water surface for both sides of the rectangular breakwater can enhance the wave dissipation ability of the breakwater while effectively decreasing the transmission and reflection coefficients.

Brain Transcriptome Analysis Reveals Metabolic Changes Adapting to Hyperhaline or Hypohaline Environments in Spotted Scat(Scatophagus argus)

The fish brain is crucial for adjusting to environmental changes.Metabolic changes play a vital role in the adaptation to salinity change in aquatic animals.However,few studies have evaluated the responses of the fish brain to salinity changes.To evaluate the response to various salinities,spotted scat(Scatophagus argus)was cultured in water with salinity levels of 5(low salinity:LS),25(control group:Ctrl),and 35(high salinity group:HS)for 22 days.The brain transcriptome was analyzed.In total,1698 differentially expressed genes(DEGs)were identified between the HS and Ctrl groups,and 841 DEGs were identified between the LS and Ctrl groups.KEGG analysis showed that the DEGs in the HS vs.Ctrl comparison were involved in steroid biosynthesis,terpenoid backbone biosynthesis,fatty acid biosynthesis,ascorbate and aldarate metabolism,other types of O-glycan biosynthesis,and fatty acid metabolism.Glyoxylate and dicarboxylate metabolism,one carbon pool by folate,steroid biosynthesis,and cysteine and methionine metabolism were significantly enriched in the LS vs.Ctrl comparison.Additionally,the genes related to metabolism(acc,fas,hmgcr,hmgcs1,mvd,soat1,nsdhl,sqle,cel,fdft1,dnmt3a and mtr)were significantly up-regulated in the HS vs.Ctrl comparison.The genes related to metabolism(lipa,sqle,acc,fas,bhmt,mpst,dnmt3a,mtr,hao2,LOC111225351 and hmgcs1)were significantly up-regulated,while hmgcr and soat1 were significantly down-regulated in the LS vs.Ctrl compparison.These results suggest that salinity stress affects signaling pathways and genes’expressions involved in metabolic processes in the brain,and the differences in metabolism play an important role in adaptation to hyperhaline or hypohaline environments in spotted scat.This research provides a comprehensive overview of transcriptional changes in the brain under hyperhaline or hypohaline conditions,which is helpful to understand the mechanisms underlying salinity adaptation in euryhaline fishes.

An Improved Chromosome-Level Genome Assembly and Annotation of Belted Beard Grunt(Hapalogenys analis)

Hapalogenys analis(order Lobotiformes)is an economically and ecologically significant fish species.It is a typical sedentary rocky reef fish and is primarily found in the northern Pacific Ocean.Here,we used Hi-C and PacBio sequencing technique to assemble a high-quality,chromosome-level genome for this species.The 539 Mb genome had a contig N50 with a size of 3.43 Mb,while 755 contigs clustered into 24 chromosomal groups with an anchoring rate of 99.02%.Of the total genomic sequence,132.74Mb(24.39%)were annotated as repeat elements.A total of 21360 protein-coding genes were identified,of which 20787 genes(97.32%)were successfully annotated to public databases.The BUSCO evaluation indicated that 96.90%of the total orthologous genes were matched.The phylogenetic tree representing H.analis and 14 other bony fish species indicated that the H.analis genome contained 364 expanded gene families related to olfactory receptor activity,compared with the common ancestor of H.analis and Sciaenidae.Comparative genomic analysis further identified 3584 contracted gene families.Branch-site modeling identified 277 genes experiencing positive selection,which may facilitate the adaptation to rocky reef environments.The genome reported here is helpful for ecological and evolutionary studies of H.analis.

Development of SNP parentage assignment techniques in the yellowfin seabream Acanthopagrus latus

Acanthopagrus latus is an essential aquaculture species on the south coast of China.However,there is a lack of systematic breeding of A.latus,which considerably limits the sustainable development of A.latus.As a result,genetic improvements are urgently needed to breed new strains of A.latus with rapid growth and strong resistance to disease.During selective breeding,it is necessary to estimate the genetic parameters of the target trait,which in turn depends on an accurate disentangled pedigree for the selective population.Therefore,it is necessary to establish the parentage assignment technique for A.latus.In this study,95 individuals selected from their parents and their 14 families were used as experimental material.SNPs were developed by genome resequencing,and highly polymorphic SNPs were screened on the basis of optimized filtering parameters.A total of 14392738 SNPs were discovered and 205 SNPs were selected for parentage assignment using the CERVUS software.In the model where the gender of the parents is known,the assignment success rate is 98.61%for the male parent,97.22%for the female parent,and 95.83%for the parent pair.In the model where the gender of the parents is unknown,the assignment success rate is 100%for a single parent and 90.28%for the parent pair.The results of this study were expected to serve as a reference for the breeding of new varieties of A.latus.

Construction of Molecular Biology and Experiment Case Library for Graduate Students

Molecular Biology and Experiment is considered fundamental for graduate students specializing in aquaculture at Guangdong Ocean University.This discipline focuses on the examination of the structure and function of macromolecules,including proteins and nucleic acids.Moreover,it elucidates biological phenomena and principles at the molecular level,making it an essential foundational course for students pursuing various biology majors.As a foundational course for the basic application of aquaculture,Molecular Biology and Experiment requires guidance through numerous examples and cases.However,there are several challenges to address in developing the case library.Consequently,a case library has been established to meet the course requirements of Molecular Biology and Experiment for modern graduate students,with the central goal of reforming the educational model of higher education institutions and enhancing the effectiveness and quality of talent development.This strategy is designed to nurture highly skilled professionals who can address the current needs of the industry.

Identification of Shell Color-Associated Genes Using Mantle Branch-Specific RNA Sequencing of Yellow-Colored Line of Pearl Oyster Pinctada fucata martensii

The yellow-colored line of pearl oyster Pinctada fucata martensii displays a yellow prismatic layer and a white nacreous layer that can be used as an ideal model for research on shell color formation.Micro-Raman spectroscopy and transcriptome analyses were performed to explore the potential molecular mechanism underlying the phenotype differentiation.The micro-Raman spectroscopy results indicate that the prismatic layer exhibits distinct characteristic peaks of carotenoids,while these peaks are not prominent in the nacreous layer.In the transcriptome comparison of the central zone of mantle and mantle edge tissue,which function in nacreous and prismatic layer formation,respectively,935 significantly differentially expressed genes(DEGs)were identified,with 385 genes upregulated and 227 genes downregulated(|log_(2)(Fold change)|>1 and false discovery rate<0.05)in the mantle edge tissue.Among these genes,some were associated with melanoma/melanogenesis,such as tyrosinase,zinc metalloprotease,glutathione S-transferase,and ATP-binding cassette sub-family;some were associated with the carotenoid-related pathway,including scavenger receptors,cytochrome P450 and lipoprotein receptor.Genes associated with porphyrin metabolism,including porphobilinogen deaminase,and copper/zinc superoxide dismutase,and genes associated with shell matrix protein,including amorphous calcium carbonate binding protein,shematrin,PIF,and collagen,also exhibited significantly different expressions.It is speculated that the different colours between prismatic layer and nacreous layer in the yellow-colored line of P.f.martensii might be resulted from melanin,carotenoids and porphyrin metabolism,while genes related to shell structure and biomineralization might also affect coloration.Our results provide new insights to understand the mechanism of shell color formation in mollusca.

Molecular Cloning and Bioinformatics Analysis of sucC Gene of Vibrio alginolyticus Strain HY9901

[Objectives]To clone the sucC gene of Vibrio alginolyticus strain HY9901 and conduct the bioinformatics analysis.[Methods]Based on the sucC gene of V.alginolyticus strain HY9901,specific primers were designed to amplify the full length sequence by PCR and make further analysis.[Results]The theoretical molecular weight of SucC protein was about 41528.45 Da,and the full length was 1167 bp,encoding 388 amino acids.It has no signal peptide and transmembrane region,and has a variety of functional sites.It is predicted that it is mainly located in the cytoplasm,and the ubiquitin and lactate modification sites overlap,and it has high gene homology with Vibrio parahaemolyticus.Theα-helix,random coil and extended strand are the main secondary structures.The similarity between the constructed three-level structure model and the template is high.[Conclusions]This study reveals the structural characteristics and functional potential of SucC protein,and provides a theoretical basis for the study of drug resistance mechanism and prevention strategies.

Cloning and Bioinformatics Analysis of hcp Gene in Aeromonas hydrophila

[Objectives]To explore the function of hcp gene in Aeromonas hydrophila.[Methods]A pair of specific primers was designed referring to the hcp gene sequence of A.hydrophila.The hcp gene was amplified by PCR,and performed bioinformatics analysis.[Results]The hcp gene had a total length of 1650 bp and encoded 549 amino acids.The theoretical molecular weight of the protein predicted was about 59476.44 kDa.After predicting the N-terminal signal peptide structure of the amino acid sequence,neither obvious signal peptide cleavage site nor signal peptide was found,and the protein had no transmembrane region.The amino acid sequence had a N-glycosylation site,4 protein kinase C phosphorylation sites,7 casein kinase II phosphorylation sites,9 N-myristoylation sites,4 isoprene binding sites,10 microbody C-terminal target signal sites,and an ATP/GTP binding site motif A(P-ring).The amino acid sequence of hcp gene of A.hydrophila was performed homology analysis with other Aeromonas strains,and it showed higher homology with A.veronii.In the secondary structure,theα-helix,β-sheet,random coil and extended strand accounted for 45.36%,6.01%,37.52%and 11.11%,respectively.The tertiary structure model consisted of 18α-helix and 22β-sheet.Analysis of protein-protein network interaction demonstrated that the proteins interacting with Hcp protein were AHA_3407,nrfA,nirB-1,nirB-2 and AHA_1112.[Conclusions]Through the bioinformatics prediction results,the basic information of hcp gene of A.hydrophila is preliminarily understood,and the possible function of this protein is predicted,in order to provide guidance for subsequent vaccine research.

Molecular Cloning of clpX Gene from Vibrio alginolyticus HY9901 and Its Bioinformatics Analysis

According to the clpX gene sequence of Vibrio alginolyticus HY9901,a pair of specific primers were designed,and the full length was cloned by PCR and subjected to bioinformatics analysis.The results showed that the clpX gene was 1281 bp in length and encoded 426 amino acids.Its molecular structure formula was C 3842 H 6405 N 1281 O 1598 S 260,with a theoretical protein molecular weight of approximately 1044473.4 kDa and a theoretical pI value of 5.04.The clpX gene was predominantly situated within the cytoplasm,exhibiting unstable and hydrophilic protein characteristics.It possessed a signal peptide cleavage site,lacked a transmembrane region,and was not associated with any KEGG metabolic pathway.Additionally,it possessed 2 glycine phosphorylation sites,a CAMP-dependent protein kinase phosphorylation site,a C-terminal amidation modification site,6 protein kinase C phosphorylation sites,7 microbody C-terminal target signal sites,and an ATP/GTP site.The clpX phylogenetic tree was constructed using the MEGA 5.0 software via the neighbor-joining method.The results demonstrated that the clpX of V.alginolyticus exhibited up to 100%affinity with the clpX of Vibrio spp.The single subunit 3D structure model of the ClpX protein was obtained using the SWISS-MODEL program.A structural and functional analysis of the protein revealed the presence of three distinct ClpX structural and functional domains.In the prediction of secondary structure,the proportions ofα-helix,random coil,β-sheet and extended strand were 40.38%,37.09%,5.40%and 17.14%,respectively.The analysis of the ClpX protein through the STRING database revealed that the proteins interacting with the ClpX protein were Tig,Atpd,Hflb,Msrb-2,Rpod,Clpp,Clpa,Lon-1,Hfq,and ANP63951.1.A computational analysis of the ClpX protein identified a number of post-translational modification sites,including phosphorylation,acetylation,ubiquitination,glycosylation,methylation,S-palmitoylation,and lactylation.The significance of this study is to analyze the function of the clpX gene and establish a robust foundation for subsequent investigations into the mechanism of the clpX gene in Vibrio alginolyticus.

Molecular Cloning and Bioinformatics Analysis of msrA Gene from Vibrio alginolyticus Strain HY9901

[Objectives]This study was conducted to understand the structure and function of MsrA protein.[Methods]With Vibrio alginolyticus HY9901 as the object of study,primers were designed to amplify the full-length gene of msrA,and its bioinformatics analysis was carried out.[Results]The full length of msrA gene was 639 bp,encoding 212 amino acids,and its theoretical molecular weight was about 23729.60 Da.The protein had a stable structure,and it was hydrophobic overall.The structure of signal peptides at the N terminal of the amino acid sequence was predicted,and it was found that there was no signal peptide cleavage site and no transmembrane region.The amino acid sequence of MsrA contained multiple signal binding sites.Protein subcellular localization showed that MsrA protein was most likely located in the cytoplasm.Homology analysis showed that MsrA of V.alginolyticus had high homology with other Vibrio species,and the highest homology with V.alginolyticus.In the prediction of functional domains,MsrA had the function of methionine sulfoxide reduction.In secondary structure prediction,MsrA contained random coils at a proportion of 46.70%,which was the highest.The similarity between the tertiary structure model of MsrA and template Q87SW6.1.A was 89.15%.PTM analysis showed that MsrA protein had many PTM modification sites such as phosphorylation and glycosylation sites.[Conclusions]This study provides some reference value for further study on the role of MsrA in bacterial antioxidant stress.

Gene Cloning and Bioinformatics Analysis of phoR Gene from Vibrio alginolyticus HY9901

PhoR is a histidine kinase in a two-component regulatory system that regulates phosphorus metabolic pathways and undertakes the key mission of information transmission in pathogenic bacteria.The full-length phoR gene was successfully cloned from the Vibrio alginolyticus HY9901 strain.A comprehensive analysis of the cloned gene was conducted using bioinformatics.Sequence analysis revealed that the total length of the phoR gene(GenBank accession No.:KJ958404.1)is 1299 bp,with the coding region containing a total of 432 amino acid residues.The phylogenetic tree of PhoR revealed that it belongs to the same subclade as V.diabolicus.The SMART program was employed for the purpose of functional domain prediction,which revealed that PhoR possesses three major functional domains:PAS(amino acids 98-166),HisKA(amino acids 205-272),and HATPase_c(amino acids 317-429).

Molecular Cloning and Bioinformatics Analysis of cyaA Gene of Vibrio alginolyticus Strain HY9901

[Objectives]This study was conducted to explore the biological functions of cyaA gene of Vibrio alginolyticus.[Methods]With DNA of V.alginolyticus HY 9901 as a template,primers were designed according to the sequence of cyaA gene,and the cyaA gene was amplified by PCR.Bioinformatics analysis was performed.[Results]The cyaA gene of V.alginolyticus HY9901 was 2529 bp in size,and encoded 842 amino acids.The molecular structure of CyaA protein was C_(4358)H_(6745)N_(1171)O_(1286)S_(35).Its theoretical molecular weight was 97.24167 kDa and the theoretical pI value was 5.56.It had no signal peptide and transmembrane domain.CyaA protein had three N-terminal glycosylation sites,one cAMP and cGMP-dependent protein kinase phosphorylation site,nine protein kinase C phosphorylation sites,nine casein kinase II phosphorylation sites,one tyrosine kinase phosphorylation site,seven N-terminal myristoylation sites,one pentenyl binding site and ten microbody C-terminal localization signal sites.Subcellular localization prediction showed that CyaA protein was mainly located in the nucleus and cytoplasm.Through multi-sequence alignment and phylogenetic tree construction,it was concluded that V.alginolyticus had high CyaA homology with other Vibrio species.cyaA of V.alginolyticus was clustered with Vibrio fluminensis and Vibrio marinisedimini,and they were closely related.The secondary structure of CyaA protein consisted ofα-helixes(43.11%),random coils(38.00%)and extended strands(14.49%).In protein network interaction,it was found that the proteins adjacent to CyaA protein were Crp-2,CpdA,Crr,PtsG-2,ANP67209.1,Crp-1,PykF,Pyk,RelA and Ndk.[Conclusions]This study provides a new idea for formulating strategies for the prevention and control of vibriosis.

Molecular Cloning of sodB Gene from Vibrio alginolyticus HY9901 and Its Bioinformatics Analysis

Vibrio alginolyticus is a zoonotic bacterium.A pair of specific primers was designed using the sodB gene sequence of Vibrio alginolyticus HY9901 in order to amplify the full length of the gene by PCR.The results indicated that the total length of the sodB gene was 585 bp and that it could encode 194 amino acids.The predicted amino acid sequence derivation indicated that the molecular weight of the protein was approximately 21.56 kDa,with an isoelectric point of 4.95.Upon prediction of the N-terminal signal peptide structure of the protein,no significant signal peptide cleavage site was observed,indicating that the protein lacked both a signal peptide and a transmembrane region.The amino acid sequence contained an N-glycosylation site,a casein kinase II phosphorylation site,a microsomal C-terminal target signal site,and a manganese and iron superoxide dismutase signal site.The probability of intracytoplasmic localization of the SodB protein was 56.5%,which was analyzed according to the subcellular localization of the protein.The amino acid sequence of the sodB gene of V.alginolyticus exhibited 98%-100%homology to other Vibrio species,clustering into the same subfamily with V.parahaem,indicating a relatively close relationship between them.In the prediction of protein structure,the proportions ofα-helix,random coil,β-sheet,and extended strand were 48.45%,30.41%,5.67%,and 15.46%,respectively.The similarity to template 1dt0.1.A reached 71.58%.A PTM site analysis revealed the presence of phosphorylation,glycosylation,ubiquitination,sumoylation,acetylation,and methylation modification sites,as well as the absence of lactylation modification sites.

Developing an intermediate-complexity projection model for China's fisheries:A case study of small yellow croaker(Larimichthys polyactis)in the Haizhou Bay,China

Projection models are commonly used to evaluate the impacts of fishing.However,previously developed projection tools were not suitable for China’s fisheries as they are either overly complex and data-demanding or too simple to reflect the realistic management measures.Herein,an intermediate-complexity projection model was developed that could adequately describe fish population dynamics and account for management measures including mesh size limits,summer closure,and spatial closure.A two-patch operating model was outlined for the projection model and applied to the heavily depleted but commercially important small yellow croaker(Larimichthys polyactis)fishery in the Haizhou Bay,China,as a case study.The model was calibrated to realistically capture the fisheries dynamics with hindcasting.Three simulation scenarios featuring different fishing intensities based on status quo and maximum sustainable yield(MSY)were proposed and evaluated with projections.Stochastic projections were additionally performed to investigate the influence of uncertainty associated with recruitment strengths and the implementation of control targets.It was found that fishing at FMSY level could effectively rebuild the depleted stock biomass,while the stock collapsed rapidly in the status quo scenario.Uncertainty in recruitment and implementation could result in variabilities in management effects;but they did not much alter the management effects of the FMSY scenario.These results indicate that the lack of science-based control targets in fishing mortality or catch limits has hindered the achievement of sustainable fisheries in China.Overall,the presented work highlights that the developed projection model can promote the understanding of the possible consequences of fishing under uncertainty and is applicable to other fisheries in China.

Marine Fishermen’s Perception about Consequences of Climate Change on Fisheries in Tamil Nadu State,India

Global climate change has become a critical barrier in the twenty-first century and it alarms the marine fishermen to respond and react until conditions become more adverse.Interaction with climate by humans occurs at all levels,but recent research has focused on industries,technological and demographic aspects.So far,less attention has been paid to investigate the factors which affect perception and decision-making at the individual level.The uncertain condition of climate on marine fisheries from the last two decades had created drastic change in ecosystems as well as in the fishers’livelihood.Fishers are the group who first experiences the bitter conditions of climate change on marine fisheries because their livelihood depends highly on the marine resources.Therefore,the fishers’perception about the consequences of climate change on fisheries may help to emerge the fisheries by implicating certain productive management measures.The research has been carried out in the two southern coastal districts of Ramanathapuram and Kanyakumari,Tamil Nadu state to find out the fishers’perception of the consequences of climate change on fisheries.A sample size consisting of 120 marine fishermen were randomly selected and interviewed.The results inferred that increase in cost of fishing,increase in fishing investments,decline in fishing days,reduction in fish diversity and fish catches were the major consequences faced by fishers due to the climate change.

Otolith microchemical evidence revealing multiple spawning site origination of the anadromous tapertail anchovy(Coilia nasus)in the Changjiang(Yangtze)River Estuary

The estuarine tapertail anchovy(Coilia nasus)is a high-value commercial fish.Estimating the spawning site or hatchery origin and habitat is essential for its conservation.This study aimed to determine the habitat use and life history characteristics of C.nasus from the Changjiang River Estuary.We investigated the environmental signatures of strontium(Sr)and calcium(Ca)in the otoliths of the collected specimens using electron probe microanalysis;additionally,we examined their gonadal maturity stage.Our results indicate that the 31 adult C.nasus specimens used in this study could be classified into two types based on their otolith Sr:Ca concentration ratios and their gonadal maturity stage.The long freshwater early life history type(Type LF)had wider central region in the otolith with low Sr:Ca concentration ratios ranging from 1.24±0.62 to 1.92±0.78 and a bluish pattern of low Sr content level.These fish are of riverine origin and had a relatively long early life history in freshwater and low gonadal maturation when captured.The short freshwater early life history type(Type SF)had a shorter central region in the otolith with low Sr:Ca concentration ratios ranging from 1.35±0.5 to 2.82±0.97 and a correspondingly bluish pattern.These fish also had a relatively short-term early life history in freshwater and high gonadal maturation when captured.The results of the otolith microchemical analysis indicated that Type LF and Type SF originated in spawning/hatching sites far from and close to the estuary,respectively.The mature gonads of Type SF fish indicated that they may breed in areas close to the estuary,whereas the immature gonads of Type LF fish indicated that they may breed in areas far from the estuary.This study is the first to reveal that the Changjiang River Estuary contains stocks of anadromous C.nasus originating in different spawning sites during the same season.The estuarine habitat plays a critical role in the connectivity between freshwater recruitment and the marine resources available to adult spawners of this commercially valued species.From a conservation perspective,this study provides important information for identifying anadromous C.nasus stocks originating in different spawning sites in the Changjiang River Basin.

DNA barcoding of fishes from Zhoushan coastal waters using mitochondrial COI and 12S rRNA genes

Accurate species identification is a key component of biodiversity research.DNA barcoding is an effective molecular method used for fish species identification.We aimed to study the DNA barcoding of fish in Zhoushan coastal waters,explore the differences and applicability of two gene fragments(12S rRNA and COI)of DNA barcoding in fish species identification,and established a comprehensive fish barcoding reference database.Two hundred and eighty-seven captured fish samples from Zhoushan coastal waters were identified using morphological characteristics and DNA barcoding.A total of 26412S rRNA sequences(belonging to eight orders,31 families,55 genera,and 66 species)and 188 COI sequences(belonging to seven orders,30 families,48 genera,and 58 species)were obtained.The lengths of the 12S rRNA sequences ranged from 165 to 178 bp,and the guanine-cytosine(GC)content was 45.37%.The average 12S rRNA interspecific and intraspecific genetic distances(K2P)were 0.10%and 26.66%,respectively.The length of the COI sequence ranged 574–655 bp,and the content of GC was 45.97%.The average 12S rRNA interspecific and intraspecific genetic distances(K2P)were 0.16%and 27.45%,respectively.The minimum interspecific genetic distances of 12S rRNA and COI(1.23%and 1.86%)were both greater than their maximum intraspecific genetic distances(2.42%and 8.66%).Three molecular analyses(NJ tree,ABGD,and GMYC)were performed to accurately identify and delineate species.Clustering errors occurred when the 12S rRNA sequences were delimited using the NJ tree method,and the delimitation results of ABGD and GMYC are consistent with the final species identification results.Our results demonstrate that DNA barcoding based on 12S rRNA and COI can be used as an effective tool for fish species identification,and 12S rRNA has good application prospects in the environmental DNA(eDNA)metabarcoding of marine fish.

A new species and a new record of free-living marine nematode of genus Diplolaimelloides(Monhysteridae)from Futian mangrove reservation in Shenzhen,China

A new species and a new record of free-living marine nematode of genus Diplolaimelloides’(Monhysteridae)inhabiting the sediment of mangrove reservation in Shenzhen,China are described and illustrated.Diplolaimelloides contortus sp.nov.is characterized by the combination of the following characters:ocelli present;four pairs of post-cloacal papillae arranged in(2+2)pattern,two midway on bursa and two near posterior extremities;spicules complicated in shape,twisted and curved at middle portion;gubernaculum wedge-shaped,with distal dorsal ridge well cuticularized,proximal dorsal ridge obscure sleeve-like,proximal end wrap the spicule;vulva located at 52.1%-53.6%of body length;male tail slender,10.0-11.4 times of anal body diameter,and female tail long filiform,13.1-13.6 times of anal body diameter.Diplolaimelloides delyi Andras sy,1958 is first recorded from China and redescribed in detail,it differs from the original description by Andrassy in 1958 for having larger body size of male,shorter tail,and posterior situation of vulva.However,it is much conform to the redescription to specimens from East Pakistan by Timm in 1966,except for shorter tail in female and absent pre-anal seta.Specimens are deposited in the Institute of Oceanology,Chinese Academy of Sciences.