Abstract:Samples of Ostracoda were collected from the Taiwan Strait in August 1984.Through the analysis of all the samples, a new species of Euphilomedes(Ostracoda:Philomedidae)is found, Euphilomedes multiangular sp.nov.The type specimen is deposited in the Third Institute of Oceanography, SOA.The specific diagnoses of new species are as follows:Euphilomedes multiangular sp.nov.(♂)The new species is distinguished by the truncate posterior with three clusters of spines along the edge;the frontal organ has five joints with three spines on the tip of the teminal segment;The third segment of the endopodite of the second antenna is elongate with uneven margins, and the long bristles near middle with marginal spines, the tip of this segment is provided with five ranged ridges.

Abstract:Stable isotope and laboratory(feeding selectivity & detritus size-fraction)experiments were used to investigate feeding behavior and ecological function of the gastropod, Turbo cornutus Solander(T.cornutus), living on the seaweed bed during the summer, 2013.Stable isotope analysis demonstrated that T.cornutus is herbivore and primary consumers(trophic level is 2.12)in seaweed bed, and prefers seaweed(>94.5%)to other potential food resources(phytoplankton, POM and SOM).Laboratory experiments revealed that T.cornutus could feed(>0.1 g individual^-1day^-1, wet weight)11 different seaweeds(including the dominant seaweed species of Gouqi Island: Sargassum horneri and Grateloupia livida).Detritus size-fraction experiments provided evidence that T.cornutus can digest seaweed, then excrete detritus(0.016-0.094)g/d into small size(dominant detritus size is 1Φ:>250 μm), these change of matter size-fractionate caused by T.cornutus grazing behavior could make contribution to detritus in seaweed bed.Stable isotope experiment, combined with laboratory(feeding selectivity & detritus size-fraction)experiments suggest T.cornutus plays an important role in connecting grazing food chain and detritus food chain in food web of seaweed bed, Gouqi Island.

Abstract:In order to collect more data of intensive aquaculture of Cynoglossus semilaevis, setting up two aquaculture modes of open flow through and recirculating aquaculture system, adult fish[(0.640±0.063)kg]in rapid growth period were observed, the index of growth and quality in muscles were detected.The experiment continued 7 months and the results were as follows:1)The survival rate of C.semilaevis in recirculating aquaculture and open flow through mode was 79.89% and 90.38%, weight gain rate was 0.154 7 and 0.110 9 kg/month, fatness was 0.920 and 0.838, growth hormone was 2.812 and 2.706 μg/L, respectively.There were respectively 13.13%, 39.5%, 9.8% and 3.92% higher in survival, weight gain, fatness and growth hormone of C.semilaevis in recirculating aquaculture mode than those in open flow through mode.2)Comparing two kinds of aquaculture models, open flow through mode has obvious superiority in crude protein, amino acid and PUFA, while recirculating mode has obvious superiority in crude fat and MUFA, these were respectively 41.732% and 16.912% higher than those in open flow through mode, which means recirculating aquaculture mode was more suitable for C.semilaevis's fattening.This study showed that C.semilaevis has a good adaptability in recirculating aquaculture mode, the growth and quality in muscles were all in a better condition, also showed that recirculating aquaculture mode can greatly develop the culture capacity of C.semilaevis.This was a preponderant culture mode for intensive culture of C.semilaevis.

Abstract:The present experiment was conducted to investigate gastric evacuation rates, change of digesta moisture content, pH values and digestive enzyme specific activity along the gastrointestinal tract of channel catfish[body weight(115.55±7.54)g]fed plant protein based diet.Three kinds of models used to fit the gastric evacuation were compared.The results showed that:all of these three kinds of models could fit the experimental data quite well, and the square root model was the best, followed by the exponential model and the linear model.The asymptotic level of 50% and 98% gastrointestinal evacuation calculated by the square root model(dry weight basis)apparently reached at 12.8 h and 36 h, respectively.Digesta moisture in the stomach showed a slow increasing trend, the upper intestine and middle intestine showed an increasing trend after the fall, while no changes were found in the lower intestine.The pH values in each part of the gastrointestinal tract decreased significantly within 3 h after feeding(P<0.05), no remarkable changes were found(P>0.05)from 3 to 30 h after feeding, while a noteworthy increasing level, recovering or drawing near the initial level was observed in intestine, within 30-36 h after feeding(P<0.05).The digestive enzyme activity had a significant increase within 3 h(P<0.05)and kept at a stable level within 3-30 h, while had a significant decrease(P<0.05)30 h after feeding.The digestive enzyme specific activity varied in different gastrointestinal sections, and ranged as upper intestine>middle intestine>lower intestine>stomach.In summary:In this study the digesta moisture, pH value and digestive enzyme along the gastrointestinal tract of channel catfish showed a regular change after feeding, and gastric evacuation pattern was affected by the interaction of these factors.

Abstract:GIFT tilapia[initial weight(64.4±1.5)g]were fed purified diets containing six levels(0.08, 0.57, 1.13, 2.09, 4.11 and 8.09 mg/kg diet, respectively)of thiamin for 12 weeks.The experiment was used to determine the dietary thiamin requirement of GIFT tilapia on the basis of growth parameters, hepatic thiamin saturation and a thiamin-dependent biochemical function.The results showed that, with the increasing dietary thiamin level, weight gain rate of tilapia first increased linearly then remained nearly unchanged.Hepatic thiamin contents presented the same tendency with weight gain rate.Fish fed diets supplemented with VB₁ showed significantly higher hepatic transketolase expression, serum high density lipoprotein cholesterol and lower serum pyruvate than fish fed diets without supplementation of VB₁(P<0.05).Fish fed the control diet without supplementation of thiamin showed significantly higher serum pyruvate content than fish fed diets supplemented with thiamin(P<0.05), while protein, moisture, ash, lipid ontent in whole body had no significant differences among various treatments(P>0.05).Based on broken method of the relationships between weight gain rate, hepatic thiamin contents and dietary VB₁ concentrations, it can be concluded that the dietary VB₁ requirement for GIFT tilapia was considered to be 1.16 and 2.06 mg/kg diet respectively.

Abstract:The experiment was conducted to investigate the effects of dosage and treatments of Enteromorpha prolifera on growth, digestibility, digestive enzymes and non-specific immunity enzymes of juvenile sea cucumber(Apostichopus japonicus Selenka).Juvenile sea cucumbers with the initial weight of(1.44±0.01)g were fed with 6 practical diets according to a single factor design:a basal diet contained 0, 5%, 10%, 15%, 20% and 25% E.prolifera for 49 days firstly, and then juvenile sea cucumber with the initial weight of(4.58±0.23)g were fed with 3 practical diets according to a single factor design:the diet contained 20% three treatments[dry crushing(DC), dry crushing after cellulase treatment(DCC)and dry crushing after protease treatment(DCP)]disposal(Wrong word)of E.proliferat for 60 days.There were 3 replicates per group with 35 sea cucumbers.The experiment results showed as follows:1)E.prolifera had significant influence on enhancing specific growth rate(SGR), the amylase(AMS)and the superoxide dismutase(SOD)in gut of sea cucumber and reducing feed conversion ratio(FCR)(P<0.05), and 20% E.prolifera were the best.The apparent digestibility coefficient of dry matter(ADCd)and apparent digestibility coefficient of crude protein(ADCp)of diets were significantly declined with the increasing level of E.prolifera(P<0.05).The different contents of E.prolifera had no significance on feed intake(FI), survival rate(SR), the pepsin(PP)and acid phosphatase(ACP)in gut of sea cucumber(P>0.05).2)E.prolifera with DCC and DCP had significant influence on enhancing SGR, ADCd, ADCp, AMS, PP, SOD and ACP in gut of sea cucumber and reducing FCR of sea cucumbers(P<0.05), and E.prolifera with DCC were the best.Three treatment of E.prolifera had no significant influence on FI and SR(P>0.05).Under the conditions of this experiment, the optimal content of E.prolifera in the diets of juvenile sea cucumber is 20%.And E.prolifera with drying crush after cellulase treatment is an ideal way to deal with E.prolifera.

Abstract:To detect Vibrio mimicus' dynamic distribution in infected C.idella, the gene encoding enhanced green fluorescent protein(EGFP)was cloned into the expression plasmid pBAD24 and transformed into the host cells V.mimicus strain 04-14 by electroporation to generate recombinant V.mimicus.The recombinant EGFP was highly expressed under induction with arabinose.The EGFP on the cell surface was also confirmed by fluorescence microscopy and flow cytometric analysis.Under nonselective condition, the stability of recombinant plasmid was 100% in EGFP-labeled V.mimicus after 30-generation time, and there was no significant difference between EGFP-labeled strain and wild strain in morphology, growth characteristics and adhesion ability to EPC cells.C.idella were immersed in water containing EGFP-labeled V.mimicus at the concentration of 5×10⁷ CFU/mL.At least three fish per group were sacrificed at different time post infection, and in different tissues and organs, such as gill, intestine, muscle, head kidney, spleen and liver were sampled separately.EGFP-labeled V.mimicus in different tissues and organs was examined by the signal of fluorescence and the method of plate culture count within 4 days after infection.The green fluorescent signals appeared firstly in the intestine and gill at 4 h after infection, and the number of bacteria reached up to 3.60×10⁸ and 2.36×10⁶ CFU/g, respectively.The number of bacteria in the intestine and gill had no obvious changes from 4 to 10 h post infection, then declined gradually after 12 h, but continued to exist until fish death.Dynamics of bacteria in muscle, head kidney, spleen and liver were similar, the green fluorescent signals in these tissues and organs were found 24 h later after infection, and the number of bacteria increased at first but later decreased from 24 to 85 h.The number of bacteria in muscle, head kidney, spleen and liver reached their peaks 48 h post infection, being 9.58×10⁴, 8.75×10⁴, 1.50×10⁴ and 4.50×10⁴ CFU/g, respectively, but the number of bacteria which was detected in them was lower than that in the intestine.These results suggest that the intestine is the primary target organ for the adhesion, colonization and reproduction of V.mimicus.

Abstract:In order to increase the understanding of the genetic resources of main economic loach species in China, seven microsatellite markers and mitochondrial D-Loop partial sequences were used to analyse three stocks of loach species, consisting of pond loach(Misgurnus anguillicaudatus, MA), largescale loach(Paramisgurnus dabryanus, PD), and Taiwan large loach(Uncategorized, TW).The results showed that, six microsatellite markers were commonly amplified in three stocks, whereas, one microsatellite marker(Mac239)specifically amplified in MA but failed to obtain effective amplified bands in PD and TW.Thirty-two haplotypes of D-Loop partial sequence detected in ninety individuals from three stocks were analyzed in this study, and there was only one which shared between PD and TW.A total of sixty-five variable sites were detected, twenty-nine of which were specific sites between MA and the others.It would be recently decreased of effective population size in TW, according the bottleneck and neutral test results.Based on the genetic variation analyses using microsatellite markers and D-Loop partial sequence, the closest Nei's genetic distance and K2P distance were found between TW and PD(0.297, and 0.006, respectively), obviously less than the genetic distance between MA and others(1.011-1.899, and 0.095-0.099, respectively);and the analysis of molecular variance(AMOVA)showed that extremely significant genetic differentiation among three stocks were detected in the present study(P<0.01).The results of genetic structure and haplotype network analysis suggested that relative genetic independence was showed among three stocks, whereas, similar genetic resources were only found in TW and PD.In conclusion, the distinct genetic differences in loach species would be effectively identified using molecular markers;and this paper speculated that Taiwan large loach might be the ecology stock or genetic improved population of P.dabryanus, but not effective taxonomy species.

Abstract:The purpose of this study is to explore the rational use of antimicrobial drugs, and prevent bacterial resistance to antibiotics through determination minimal inhibitory concentration(MIC), mutant prevention concentration(MPC)and mutant selection window(MSW)of monotherapy and double medicine antibiotics on Aeromonas punctata.Resistances of 10 antibiotics were tested via drugs sensitivity assays, 6 sensitive antimicrobial drugs with different mechanisms were screened and their MIC, MPC, selection index(SI)and MSW to Aeromonas punctata were determined respectively via single-drug and double-drug's combination by agar plate dilution method.The MIC of chloramphenicol, oxytetracycline, rifampicin, erythromycin, tetracycline hydrochloride, cephalosporins were 0.512, 0.416, 6.272, 2.048, 0.386, 0.512 μg/mL, respectively.The MPC of 6 antibiotics were 524.288, 1 277.952, 802.816, 1 024.000, 1 185.792, 10.880 μg/mL.The SI were 1 024.000, 3 072.000, 128.000, 500.000, 3 072.000, 21.250.The combination index of oxytetracycline and erythromycin, tetracycline hydrochloride and erythromycin, tetracycline hydrochloride and cephalosporins, oxytetracycline and cephalosporins, tetracycline hydrochloride and oxytetracycline, chloramphenicol and oxytetracycline, chloramphenicol and tetracycline hydrochloride, rifampicin and chloramphenicol, rifampicin and cephalosporins, rifampicin and tetracycline hydrochloride, rifampicin and oxytetracycline, erythromycin and cephalosporins were 1.000, 1.000, 1.000, 1.000, 0.834, 1.500, 1.500, 0.834, 2.000, 2.000, 1.000, 2.000, respectively.It was confirmed that the mutant selection window of A.punctata was narrowed effectively via double drug combination.The SIs declined from 128.000 and 3 072.000 to 1.000 and 1.000 after combination of rifampicin and tetracycline, so this demonstrated that combination of rifampicin and tetracycline closed the resistant mutant selection window, and inhibited effectively the resistant growth of A.punctata.

Abstract:Aeromonas is widely distributed in freshwater environments.Since the late 1980s, Aeromonas has caused serious economic losses to the aquaculture industry in our country.The published species names are increasing in the genus Aeromonas.In order to determine the disease-causing bacterium at one fish farm of Nanjing, some tissue samples from diseased fish were collected and the pathogenic bacteria were identified by means of bacterial culture, biochemical characterizations and gyrB gene sequencing.One strain of bacterium was isolated and named LK-25.The results of biochemical tests showed that the bacterial strain has many similarities compared with A.hydrophila.Then the strain was subjected to sequencing of the gyrB gene for species identification, and then it demonstrated that the strain shared 99% homology with the A.aquariorum strain MDC 47.The corresponding gyrB phylogenetic analysis including representative strains from some known Aeromonas species showed that A.aquariorum LK-25 was very closely related to the A.hydrophila ssp.dhakensis.The detection result of virulence factors indicated that A.aquariorum LK-25 strain carried five virulent genes, including act, ahp, epr, hcp and alt genes, which encoded the cytotoxin, serine proteases, extracellular protease, hemolysin co-mediate protein and cytotonic enterotoxins, respectively.A.aquariorum LK-25 also showed strong hemolytic and proteolytic activities.The median lethal dose(LD50)with 1.02×10³ CFU/ml in zebrafish indicated that A.aquariorum LK-25 is a highly virulent strain.A.aquariorum was a new species originally isolated from ornamental fish in 2008, and widely distributed in clinical and environmental specimens.Recent reports highlighted the importance of this species in cases of gastroenteritis, soft tissue infections, bacteremia and peritoneal dialysates in human, suggesting its public health significance.To our knowledge, this was the first report of A.aquariorum in China.The study will lay a good foundation for a better understanding of the occurrence and spread of A.aquariorum infection.

Abstract:To study the mechanisms of immune system formation at the early developmental stages of the Pacific cod, Gadus macrocephalus, transcriptional levels of RAG1 and IgM were measured to characterize the immune system development.According to the sequences of RAG1 and IgM in GenBank, pairs of specific primers were designed separately to amplify both genes from the head kidney of Pacific cod.Then the fragments of RAG1 and IgM genes were inserted into pMD18-T cloning vector to construct the standard plasmids of RAG1 and IgM genes.Both absolute(A-qPCR)and relative quantification PCR(R-qPCR)were established and optimized.The results showed that a linear regression equation of RAG1 is y=-3.266 2x+33.772, and the regression coefficient(R²)of the standard curve is 0.996 1;a linear regression equation of IgM is y=-3.119 7x+27.612, and the regression coefficient(R²)of the standard curve is 0.998 1.The results of A-qPCR and R-qPCR revealed a consistent trend in gene transcription.The RAG1 was only detected in the thymus and head kidney, and the expression level in thymus was higher than that in the head kidney.The IgM was detected in the thymus, kidney, liver and spleen, mainly in the spleen and head-kidney.The expression level of RAG1 did not increase until 61 to 95 days post-hatching(dph), while the expression level of IgM started to increase from 33 to 65 dph and significantly increased on 95 dph.Further analysis showed that the method of A-qPCR is reliable and specific to analyze the expression of target genes.

Abstract:In order to establish a simple, rapid, sensitive and specific method for detection of the SB strain of Ostreid herpesvirus-1(OsHV-1), a cross priming amplification(CPA)assay was developed based on primers developed according to the conserved regions selected across the whole genome sequences of the SB strain of OsHV-1.Then the reaction temperature, time and concentrations of dNTPs and Mg²⁺ were optimized.Our results showed the optimum temperature and time of the assay were 63 ℃ and 60 min, and the optimum concentration of dNTP and Mg²⁺were 1.4 mmol/L and 6 mmol/L respectively.The CPA assay was highly sensitive with detection limit approximately 30 copies recombinant plasmid DNA per μL.The assay was also highly specific for OsHV-1-SB detection, with no cross-reactions was found when acute viral necrosis virus, abalone herpes viruses, Perkinsus sp., Bonamia sp., Martelia sp., white spot syndrome virus and Vibrio parahaemolyticus were used as controls.The prevalence of OsHV-1-SB in 22 blood clams, Scapharca broughtonii, collected from Gyeongsangnam-do in Korea, Shandong and Liaoning Province in China were detected with the CPA assay established in this study.The results showed that the CPA assay is a simple, rapid, sensitive, specific and reliable technique.Additionally, since the results of CPA assay could be read directly through centrifugation of the reaction mix or adding GeneFinder^TM to the mix, this assay can be used in shellfish farms and local laboratories with poor conditions.

Abstract:Trehalose-6-phosphate synthase(TPS), the key enzyme involved in trehalose biosynthesis, plays particularly important roles in a variety of stress responses.In this study, based on unigene sequences which were obtained from whole transcriptome sequencing of Pyropia haitanensis, three full-length PhTPS genes were obtained by rapid amplification of cDNA ends(RACE), and named PhTPS1, PhTPS2-1 and PhTPS2-2.The full-length cDNA of the PhTPS gene comprised 3 557 nucleotides and contained an open reading frame of 3 462 bp(GenBank accession:KM580358), encoding a protein of 1 153 amino acid residues with the predicted molecular weight of 124.2 ku and theoretical isoelectric point of 6.73;the full-length cDNA of the PhTPS2-1 gene comprised 4 264 nucleotides and contained an open reading frame of 4 044 bp(GenBank accession:KM519457), encoding a protein of 1 374 amino acid residues with the predicted molecular weight of 145.2 ku and theoretical isoelectric point of 5.96;and the full-length cDNA of the PhTPS2-2 gene comprised 3 733 nucleotides and contained an open reading frame of 3 324 bp(GenBank accession:KM519458), encoding a protein of 1 107 amino acid residues with the predicted molecular weight of 120.2 ku and theoretical isoelectric point of 5.42.On the basis of conserved motifs and phylogenetic tree analysis, PhTPS1 belongs to the Ⅰ subfamily of TPS and PhTPS2-1 and PhTPS2-2 belong to the Ⅱ subfamily of TPS.The expressions of the three PhTPS genes, as measured by real-time quantitative PCR, were significantly induced by high-temperature stress and desiccation stress, but had different expression patterns.During high-temperature stress, the expression levels of the three PhTPS genes were all significantly upregulated firstly and then decreased, but as the high-temperature stress continues, the expression levels were upregulated again.However, during desiccation, the expression levels of PhTPS1 and PhTPS2-2 were significantly upregulateded only when the water loss was >60% or 75%, but the expression level of PhTPS2-1 was significantly upregulat when the water loss was >15%.These results suggested that the three PhTPS genes may play important roles in the response to higher desiccation stress.

Abstract:To inquire the correlation of cathepsins activities and texture and myofibrillar protein degradation of tilapia fillets during iced storage, the changes of cathepsins B, H, L and D activities over a period of iced storage were measured.And these variation trends of texture of fillets as well as myofibrillar protein with iced time and correlation among them were also determined.The results showed that cathepsins B and D activities presented a fluctuatingly upgraded tendency on the whole, and cathepsins L, sum of B and L activities presented a first descending and then upgraded tendency.A highly significant positive correlation was observed between cathepsins B and sum of B and L activities and iced storage period.Through the duration of storage, changing scale of cathepsin H activity was small.Myofibrillar protein had a maximum content on the iced 2^nd day, and its overall variation trend showed a significant negative correlation with iced time.The SDS-PAGE pattern showed that myosin heavy chain degraded obviously, while actin and tropomyosin had no remarkable changes, and α-actinin degraded gradually in the first iced four days.Texture indices including hardness, gumminess and chewiness on the whole presented a fluctuating tendency, and cohesiveness as well as springiness did not change remarkably through the iced storage, whereas adhesiveness and adhesive force increased gradually two days after iced storage.The correlation analysis of cathepsins activities and texture as well as myofibrillar protein content indicated that cathepsin B activity was largely related to springiness and myofibrillar protein content, and cathepsins L and D presented a significant correlation with hardness, gumminess and chewiness.Cathepsins L and D were the most likely to be directly involved in the postmortem softening of tilapia fillets muscle during iced storage.Lysosomal cathepsins may have participated jointly in autolysis of fish.

Abstract:Bangia atropurpurea has been inadequately studied in China.To understand the basic biological characteristics of B.atropurpurea in China, the morphology, reproduction, chromosome characteristics and pigment contents of B.atropurpurea were studied in this research.Three B.atropurpurea samples were collected from Niangziguan(Shanxi Province), Wuquanshan(Gansu Province)and Xinglongshan(Gansu Province).The growth environment conditions of the Bangia samples were studied in the fields.The morphological and reproduction characteristics of these three samples were examined under the light microscope.The nuclear division processes of samples were observed under a photomicroscope with an aceto-iron-haematoxylin-chloral hydrate stain.The freshwater Bangia populations in these three different locations were found adhered to rocks, waterfall and fast flowing streams, and the Bangia populations were often found to coexist with Cladophora insignis.These Bangia samples were unbranched filaments and the length ranged from 1.1 cm to 3.5 cm.Phycobiliprotein contents of three samples were ranged from 7.26 to 9.46 mg/g, in which the sample from Niangziguan contained the highest concentration, followed by the sample from Wuquanshan.The Xinlongshan sample contained the lowest phycobiliprotein contents concentration in comparison with other samples.These Bangia samples reproduced asexually and the released archeospores germinated into gametophytes continously.The nuclear division of three Bangia samples were mitosis and the sister chromosomes always assembled as a "8" or a "X" shape in the early metaphase.All freshwater samples had the consistent somatic chromosome numbers of n=4.The length of chromosomes were ranged from 0.42 to 1.67 μm and one of the chromosomes was usually found to be shorter than the other three.This is the first time that the biology characteristics of Bangia samples from Wuquanshan and Xinglongshan have been systematically studied in China.That provides useful information for the B.atropurpurea research.

Abstract:There is a growing interest for the progress and application of transcriptomics technology in recent years.Two types of techniques, including hybridization-or sequence-based method, have been developed for transcriptome investigations.With the emergence of next-generation sequencing techniques, those technologies have been widely used in the genetic studies of economic traits and molecular-aided selections for aquatic animals.This review has briefly summarized the progress of transcriptomic researches in aquaculture, involving immune response, growth, development, evolution and toxicology.Meanwhile, we have discussed challenges in both transcriptomic technologies and elaborate applications in aquaculture and analyzed prospects of further development of transcriptome researches on aquatic animals.