An AFLP Genetic Linkage Map of Pacific Abalone （Haliotis discus hannai）

A genetic linkage map of Pacific abalone （Haliotis discus hannai） was constructed using AFLP markers based on a two-way pseudo-testcross strategy in a full-sib family. With 33 primer combinations, a total of 455 markers （225 from the female parent and 230 from the male parent） segregated in a 1 ： 1 ratio, corresponding to DNA polymorphism： heterozygous in one parent and null in the other. The female framework map consisted of 174 markers distributed in 18 linkage groups, equivalent to the H. discus hannai haploid chromosome number, and spanning a total length of 2031.4 cM, with an average interval of 13.0 cM between adjacent markers. The male framework map consisted of 195 markers mapped on 19 linkage groups, spanning a total length of 2273.4cM, with an average spacing of 12.9cM between adjacent markers. The estimated coverage for the framework linkage maps was 81.2% for the female and 82.1% for the male, on the basis of two estimates of genome length. Fifty-two markers （11.4%） remained unlinked. The level of segregation distortion observed in this cross was 20.4%. These linkage maps will serve as a starting point for linkage studies in the Pacific abalone with potential application for marker-assisted selection in breeding programs.     

Isolation, cloning and sequencing of AFLP markers related to disease-resistance traits in Fenneropenaeus chinensis

Amplified fragment length polymorphisms （AFLP） technique was used to analyze the fingerprinting of four successive generations of Fenneropenaeus chinensis to reveal their disease-resistance traits. Some loci showed quite different genetic frequencies due to artificial selection, which implied that these fragments were putative markers related to the disease-resistance trait. We developed a simple and effective method to further characterize these AFLP fragments. Specific AFLP bands were cut directly from polyacrylamide gels, re-amplified, cloned and sequenced. Eight putative genetic markers were sequenced and their sizes ranged from 63 to 209 bp. The sequences were submitted to dbGSS （database of Genome Sequence Survey）; and the BLAST analysis showed low similarity to the function genes, indicating these markers were tightly linked to a disease-resistance trait but were not functional genes.     

Hematological changes in white spot syndrome virus-infected shrimp, Fenneropenaeus chinensis (Osbeck)

The pathological changes of hemocytes in the haemolymph and hepatopancreas were examined in experimentally and naturally WSSV (white spot syndrome virus) infected Fenneropenaeus chinensis. The results showed that the pathological manifestations of hemocytes were similar among moribund shrimps infected via injection, feeding and by nature. Firstly, the total hemocyte counts (THCs) in WSSV-infected shrimp were significantly lower than those in healthy shrimp. Secondly, necrotic, broken and disintegrated cells were often observed, and a typical hematolysis was present in the haemolymph smear of WSSV-infected shrimp. Thirdly, necrosis and typical apoptosis of hemocytes were detected with TEM in the peripheral haemolymph of WSSV-infected shrimp. Hyalinocytes and semi-granulocytes with masses of WSSVs in their nuclei often appeared, whereas no granular hemocytes with WSSV were found in the hepatopancreas of moribund infected shrimps. All our results supported that hemocytes were the main target cells of WSSV, and hyalinocytes and semigranular hemocytes seemed to be more favorable for WSSV infection in F. chinensis.     

A microsatellite genetic linkage map of black rockfish (Sebastes schlegeli)

Ovoviviparous black rockfish (Sebastes schlegeli) is an important marine fish species for aquaculture and fisheries in China. Genetic information of this species is scarce because of the lack of microsatellite markers. In this study, a large number of microsatellite markers of black rockfish were isolated by constructing microsatellite-enriched libraries. Female- and male-specific genetic linkage maps were constructed using 435 microsatellite markers genotyped in a full-sib family of the fish species. The female linkage map contained 140 microsatellite markers, in which 23 linkage groups had a total genetic length of 1334.1 cM and average inter-marker space of 13.3 cM. The male linkage map contained 156 microsatellite markers, in which 25 linkage groups had a total genetic length of 1359.6 cM and average inter-marker distance of 12.4 cM. The genome coverage of the female and male linkage maps was 68.6% and 69.3%, respectively. The female-to-male ratio of the recombination rate was approximately 1.07:1 in adjacent microsatellite markers. This paper presents the first genetic linkage map of microsatellites in black rockfish. The collection of polymorphic markers and sex-specific linkage maps of black rockfish could be useful for further investigations on parental assignment, population genetics, quantitative trait loci mapping, and marker-assisted selection in related breeding programs.     

Peptidomic analysis of Chinese shrimp (Fenneropenaeus chinensis) hemolymph by magnetic bead-based MALDI-TOF MS

Peptides in shrimp hemolymph play an important role in the innate immune response.Analysis of hemolymph will help to detect and identify potential novel biomarkers of microbial infection.We used magnetic bead-based purification(ClinProt system) and matrix-assisted laser desorption/ionization time of flight mass spectrometry(MALDI-TOF MS) to characterize shrimp hemolymph peptides.Shrimp serum and plasma were used as the source of samples for comparative analysis,and it was found that serum was more suitable for shrimp hemolymph peptidomic analysis.To screen potential specific biomarkers in serum of immune-challenged shrimps,we applied magnetic bead-based MALDI-TOF MS to serum samples from 10 immune-challenged and 10 healthy shrimps.The spectra were analyzed using FlexAnalysis 3.0 and ClinProTools 2.1 software.Thirteen peptide peaks significantly different between the two groups were selected as candidate biomarkers of lipopolysaccharide(LPS)-infection.The diagnostic model established by genetic algorithm using five of these peaks was able to discriminate LPS-challenged shrimps from healthy control shrimps with a sensitivity of 90% and a specificity of 100%.Our approach in MALDITOF MS-based peptidomics is a powerful tool for screening bioactive peptides or biomarkers derived from hemolymph,and will help to enable a better understanding of the innate immune response of shrimps.     

Purification and identification of a clotting protein from the hemolymph of Chinese shrimp (Fenneropenaeus chinensis)

The clotting protein (CP) plays important and diverse roles in crustaceans, such as coagulation and lipid transportation. A clotting protein was purified from the hemolymph of Chinese shrimp Fenneropenaeus chinensis (named as Fc-CP) with Q sepharose HP anion-exchange chromatography and phenyl sepharose HP hydrophobic interaction chromatography. Fc-CP was able to form stable clots in vitro in the presence of hemocyte lysate and Ca²⁺, suggesting that the clotting reaction is catalyzed by a Ca²⁺-dependent transglutaminase in shrimp hemocytes. The molecular mass of Fc-CP was 380 kDa under non-reducing conditions and 190 kDa under reducing conditions as was determined with SDS-PAGE. CP exists as disulfide-linked homodimers and oligomers. The N-terminal amino acid sequence of Fc-CP was identical to that of shrimps including Penaeus monodon, Farfantepenaeus paulensis and Litopenaeus vannamei; and similar to that of other decapods. The purified Fc-CP was digested with trypsin and verified on an ABI 4700 matrix-assisted laser desorption/ionization tandem time-of-flight (MALDI-TOF/TOF) mass spectrometry. Our results will aid to better understanding the coagulation mechanism of shrimp hemolymph.     

Requirements of shrimp,Penaeus chinensis O'sbeck for potassium,sodium, magnesium and iodine

Potassium, sodium, magnesium and iodine requirements of shrimp,Penaeus chinensis were studied. Orthogonal design was employed in this experiment. The composition of the basal diet consisted of fish meal, peanut cake, corn meal, soybean cake, wheat bran, vitamin mix and mineral mix, and supplementations of potassium, sodium, magnesium and iodine in the basal diet were made according to the L₉(3⁴) orthogonal table. The results indicated that iodine supplementation improved growth of the shrimp significantly and raised survival very significantly, iodine requirement of the shrimp was 0.003%; sodium requirement was 0.87% or less; and that 1.1–1.3% potassium and 0.18–0.38% magnesium in the diet were proper nutrition supplements for the shrimp. Contribution No. 2342 from the Institute of Oceanology, Chinese Academy of Sciences.     

Genetic improvement on Chinese shrimp （Fenneropenaeus chinensis）： growth and viability performance in F1 hybrids of different populations

Fenneropenaeus chinensis distributed in the Yellow Sea and Bohai Sea of China and the west coast of the Korean Peninsula. Different geographical populations represent potentially different genetic resources. To learn further the characteristics of different geographical population, crosses among two wild and three farmed populations were produced. The two wild populations were from the Yellow Sea and Bohai Sea （WYP）, and the west coast of the Korean Peninsula and coast （WKN）. The three farmed populations included the offspring of first generation of wild shrimp from coast in Korea （FKN）, the Huang Hai （the Yellow Sea in Chinese） No.1 （HH1）, and JK98. The phenotypes growth and survival rates of these populations were compared to confirm the feasibility for crossbreeding. The body length （BL）, carapace length （CL）. carapace width （CW）, height of the second and third abdominal segment （HST）, width of the second and third abdominal segment （WST）, length of the first abdominal segment （LF）, length of the last abdominal segment （LL）, live body weight （BW）, and survival rate were measured, Different combinations were statistically performed with ANOVA and Duncan＇s Multiple Range Test. The results show that the survival rate of JK98（♀）×WKN（♂） was the highest, followed by WYP（♀）×WKN（♂）, FKN（♀）×WYP（♂）, FKN（♀）×HHI（♂） and WYP（♀）×FKN（♂）; the body weight of FKN（♀）×HHI（♂） was the highest, followed by FKN（♀）×WYP（♂）, WYP（♀）×WKN（♂）, WYP（♀）×FKN（♂） and JK98（♀）×WKN（♂）; the total length had the same ranking as the body weight. All growth traits in hybrids JK98（♀）×WKN（♂） were the lowest among all combinations. F1 hybrids had significant difference （P〈0.05） in BL, CL, HST, LL, and BW; and insignificant difference （P〉0.05） in other growth traits and survival rate. The results of Duncan＇s Multiple Range Test are that BL and CL of J     

Cloning,characterization,and expression analysis of the DEAD-box family genes,Fc-vasa and Fc-PL10a,in Chinese shrimp (Fenneropenaeus chinensis)

RNA helicases of the DEAD-box and related families are involved in various cellular processes including DNA replication,DNA repair,and RNA processing.However,the function of DEAD-box proteins in aquaculture species is poorly understood at molecular level.We obtained the full-length cDNA sequences of two genes encoding helicase-related proteins,Fc-vasa and Fc-PL10a,from the testes of Chinese shrimp,Fenneropenaeus chinensis.The two predicted amino acid sequences contain all the conserved motifs characterized ...     

Chromosomal localization of 5S rDNA in Chinese shrimp (Fenneropenaeus chinensis): a chromosome-specific marker for chromosome identification

Chinese shrimp(Fenneropenaeus chinensis)is an economically important aquaculture species in China.However,cytogenetic and genomic data is limited in the organism partly because the chromosomes are difficult to isolate and analyze.In this study,fluorescence in-situ hybridization(FISH) was used to identify the chromosomes of F.chinensis.The 5S ribosomal RNA gene(rDNA)of F. chinensis was isolated,cloned and then used as a hybridization probe.The results show that the 5S rDNA was located on one pair of homologous chromosomes in F.chinensis.In addition,triploid shrimp were used to evaluate the feasibility of chromosome identification using FISH and to validate the method.It was confirmed that 5S rDNA can be used as a chromosome-specific probe for chromosome identification in F.chinensis.The successful application of FISH in F.chinensis shows that chromosome-specific probes can be developed and this finding will facilitate further research on the chromosomes of penaeid shrimps.     

Nitrogen and Phosphorus Budget of a Polyculture System of Sea Cucumber （Apostichopus japonicus）, Jellyfish （Rhopilema esculenta） and Shrimp （Fenneropenaeus chinensis）

The nitrogen(N) and phosphorus(P) budget and the ecological efficiency of a polyculture system of sea cucumber(Apostichopus japonicus), jellyfish(Rhopilema esculenta) and shrimp(Fenneropenaeus chinensis) were studied in a cofferdam, 120.2 ha in size. The nutrients were supplied by spring tide inflow. In total, 139600 kg N yr-1 and 9730 kg P yr-1 input to the system; while 118900 kg N yr-1 and 2840 kg P yr-1 outflowed from the system concurrently, thus the outflow was 85.7%(N) and 29.2%(P) of inflow. The production of N and P was 889.5 kg yr-1 and 49.28 kg yr-1(sea cucumber) and 204 kg yr-1 and 18.03 kg yr-1(jellyfish and shrimp), respectively. The utilization rate of N and P by polycultured animals was 7.8‰ and 6.9‰, respectively, 21.9% and 38% higher than that of monocultured sea cucumber. Our results indicated that the polyculture system was an efficient culture system of animals and a remediation system of coastal environment as well; it scavenged 14.3% and 70.8% of N and P, respectively. Such an ecological efficiency may be improved further by increasing either the stocking density or the size of sea cucumber or both.     

Chromosomal localization of 5S rDNA in Chinese shrimp (Fenneropenaeus chinensis): a chromosome-specific marker for chromosome identification

Chinese shrimp (Fenneropenaeus chinensis) is an economically important aquaculture species in China. However, cytogenetic and genomic data is limited in the organism partly because the chromosomes are difficult to isolate and analyze. In this study, fluorescence in-situ hybridization (FISH) was used to identify the chromosomes of F. chinensis. The 5S ribosomal RNA gene (rDNA) of F. chinensis was isolated, cloned and then used as a hybridization probe. The results show that the 5S rDNA was located on one pair of homologous chromosomes in F. chinensis. In addition, triploid shrimp were used to evaluate the feasibility of chromosome identification using FISH and to validate the method. It was confirmed that 5S rDNA can be used as a chromosome-specific probe for chromosome identification in F. chinensis. The successful application of FISH in F. chinensis shows that chromosome-specific probes can be developed and this finding will facilitate further research on the chromosomes of penaeid shrimps.     

Erratum to: Cloning, characterization, and expression analysis of the DEAD-box family genes, Fc-vasa and Fc-PL10a, in Chinese shrimp (Fenneropenaeus chinensis)

<正>In the original article, the name of the fourth author is incorrect. The correct one is Kyoung Ho Kang, in which "Kang" is the family name. The publishing companies apologize to the author.     

Effects of salinity fluctuation frequency on the growth, molting rate and hemolymph 20-hydroxyecdysone concentration in Juvenile Chinese shrimp, Fenneropenaeus chinensis

This experiment was conducted to examine the effects of salinity fluctuation frequency on the growth, molting rate and hemolymph 20-hydroxyecdysone (20-HE) concentration for juvenile Fenneropenaeus chinensis with an initial wet body weight of 1.115 g±0.012 g. The salinity of the control treatment (represented by D0) was 28 throughout the experiment, whereas treatments D2, D4, D6 and D8 were subjected to different salinity fluctuation frequencies of 2, 4, 6 and 8 d, respectively. The range of salinity fluctuation was 4. The salinity in treatments D2, D4, D6 and D8 was kept at 28 for 2, 4, 6 and 8 d, respectively; each original salinity was decreased abruptly to salinity 24, which first lasted for another 2 d and was then raised to its initial value 28. This constituted a salinity fluctuation cycle and afterwards the cycle repeated. It was found that: 1) The weight gain of shrimp in treatment D4 was 202.9% and significantly higher than that in treatment D0 (P<0.05). The molting rate in treatment D2 was the lowest, while that in treatment D6 was the highest and significantly higher than those in treatments D0 and D2 (P<0.05). 2) The hemolymph 20-HE concentration of shrimps in all treatments was at low levels and increased first gradually during the inter-molt period and then increased sharply. It reached a peak value during the pre-molt stage, and then decreased abruptly during the post-molt stage when the lowest level occurred. 3) During the post-molt stage, the hemolymph 20-HE concentration in treatment D2 was 7.47 pg μL⁻¹ and significantly higher than that in treatment D0 (P<0.05). There were, however, no significant differences in hemolymph 20-HE concentration among all five treatments during the inter-molt stage (P>0.05). During the pre-molt stage, the hemolymph 20-HE concentration in treatment D6 was significantly higher than that in treatment D0 (P<0.05), whereas the hemolymph 20-HE concentration in treatment D2 was significantly lower than that in other treatments (P<0.05). Adequate salinity fluctuation promoted the molting rate through increasing hemolymph 20-HE concentration at the pre-molt stage.     

Effects of Salinity Fluctuation Frequency on the Growth, Molting Rate and Hemolymph 20-Hydroxyecdysone Concentration in Juvenile Chinese shrimp, Fenneropenaeus chinensis

This experiment was conducted to examine the effects of salinity fluctuation frequency on the growth, molting rate and hemolymph 20-hydroxyecdysone (20-HE) concentration for juvenile Fenneropenaeus chinensis with an initial wet body weight of 1.115 g±0.012 g. The salinity of the control treatment (represented by D0) was 28 throughout the experiment, whereas treatments D2,D4, D6 and D8 were subjected to different salinity fluctuation frequencies of 2, 4, 6 and 8d, respectively. The range of salinity fluctuation was 4. The salinity in treatments D2, D4, D6 and D8 was kept at 28 for 2, 4, 6 and 8d, respectively; each original salinity was decreased abruptly to salinity 24, which first lasted for another 2d and was then raised to its initial value 28. This constituted a salinity fluctuation cycle and afterwards the cycle repeated. It was found that: 1) The weight gain of shrimp in treatment D4 was 202.9% and significantly higher than that in treatment D0 (P<0.05). The molting rate in treatment D2 was the lowest, while that in treatment D6 was the highest and significantly higher than those in treatments D0 and D2 (P<0.05). 2) The hemolymph 20-HE concentration of shrimps in all treatments was at low levels and increased first gradually during the inter-molt period and then increased sharply. It reached a peak value during the pre-molt stage, and then decreased abruptly during the post-molt stage when the lowest level occurred. 3) During the post-molt stage, the hemolymph 20-HE concentration in treatment D2 was 7.47pgμL-1 and significantly higher than that in treatment D0(P<0.05). There were, however, no significant differences in hemolymph 20-HE concentration among all five treatments during the inter-molt stage (P>0.05). During the pre-molt stage, the hemolymph 20-HE concentration in treatment D6 was significantly higher than that in treatment D0(P<0.05), whereas the hemolymph 20-HE concentration in treatment D2 was significantly lower than that in other treatments (P<0.05). Adequate salinity fluctuation promoted the molting rate through increasing hemolymph 20-HE concentration at the pre-molt stage.     

Effect of Co-Culture of Chinese Shrimp(Fenneropenaeus chinensis) and Sea Cucumber(Apostichopus japonicus Selenka) on Pond Environment

Monoculture of sea cucumber(pond S) and polyculture of shrimp with sea cucumber(pond SS) were established to evaluate the effect of shrimp on the environmental conditions of sea cucumber farming pond. Contributions of sediment organic matter(SOM2) resuspended from benthic sediment and the suspended particulate organic matter(SPOM) deposited from the water column to the precipitated organic matter(SOM1) collected with sediment traps were estimated with carbon stable isotope analysis. The results showed that the levels of SPOM and SOM2 in pond SS significantly decreased in comparison with those in pond S at the end of experiment(P 0.05), indicating that co-culturing shrimp in sea cucumber farming pond could purify the farming water. Carbon stable isotope analysis showed that the proportion of SOM2 in SOM1 in pond SS(84.97% ± 0.38%) was significantly lower than that in pond S(95.20% ± 0.30%)(P 0.05), suggesting that the resuspension of organic matter from benthic sediment into overlying water was reduced in polyculture pond. In contrast, the proportion of SPOM in SOM1 in pond SS(15.03% ± 0.38%) was significantly higher than that in pond S(4.80% ± 0.30%)(P 0.05), indicating that the sedimentation of SPOM from water column was enhanced in pond SS owing to the biodeposition effect of shrimp.