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1.
Cultured Apostichopusjaponicus in China suffers from a kind of skin ulceration disease that has caused severe economic loss in recent years. The disease, pathogens of which are supposed to be bacteria by most researchers, is highly infectious and can often cause all individuals in the same culture pool to die in a very short time. The 16S rRNA gene phylogenesis of the culturable bacteria from the lesions of diseased individuals was conducted to study the biodiversity of the bacterial communities in the lesions and to identify probable pathogen(s) associated with this kind of disease. S. japonica samples were selected from a hatchery located in the eastern part of Qingdao, China. Bacterial universal primers GM5F and DS907R were used to amplify the 16S rRNA gene of bacteria colonies, and touchdown PCR was performed to amplify the target sequences. The results suggest that γ- proteobacteria (Alteromonadales and Vibrionales) of CFB group, many strains of which have been also determined as pathogens in other marine species, are the predominant bacterial genera of the diseased Apostichopusjaponicus individuals.  相似文献   

2.
The purpose of this study was to investigate the antibacterial activity of extracts from Zostera marina against the pathogens of Apostichopus japonicus skin ulceration disease. When 95% ethanol (v/v) solvent was used to extract Zostera marina at 50°C, aqueous extract (ZA) showed obvious bacteriostatic effects on the tested bacterial strains (inhibition halo diameters between 8.23 mm and 13.62 mm), whereas the ethyl acetate extract (ZE) was almost inactive. The minimal inhibitory concentration (MIC) of ZA against four pathogens were homogeneous at 12.8 g L−1. ZA components were analyzed by thin layer chromatography (TLC) assay and six fractions were obtained. In another study, the six fractions showed inhibitory effects against the tested bacteria while their functions seemed to counteract the ZA activity.  相似文献   

3.
Studies were conducted to determine the cause of the acute mortality of half-smooth tongue sole Cynoglossus semilaevis Günther juveniles in a fish farm in Jimo, Shandong Province, China, in June 2006. Gross signs of the diseased tongue sole included several petechiae and ecchymoses on the body and fin necrosis and hemorrhagic lesion at the base of the fin. Bacteria were isolated from kidney, liver and hemorrhagic lesions of the diseased tongue sole. Among14 strains, SJ060621 was proved to be highly virulent to juvenile tongue sole with LD50 value of <1.0×105 colony forming units (CFU) mL−1, while the remaining 13 were avirulent. Among the 16 antibiotics tested, SJ060621 was sensitive to gentamicin and nitrofurantoin. It was identified as Listonella anguillarum with conventional plate and tube tests in combination with API 20E analysis. 16S rRNA gene and partial HSP60 gene sequenceing analysis revealed that the strain was highly homologous with L. anguillarum. Examination of the infected musculature by electron microscopy indicated numerous bacteria and lots of macrophages containing phagocytosed bacteria. Histopathological investigations revealed severe necrotic degenerative changes in the infected organs. Indirect immunofluorescence assay (IFA) was employed to detect the location of occurrence of bacteria, and bacteria were found in aggregations in the inflammatory areas in musculature.  相似文献   

4.
This study investigated the bacterial diversity of gut content of sea cucumber(Apostichopus japonicus) and its habitat surface sediment in a bottom enhancement area using PCR-based denaturing gradient gel electrophoresis(DGGE) technique. Bacterial diversity evaluation showed that the value of the Shannon-Wiener index of gut content in different intestinal segments of A. japonicus varied between 2.88 and 3.00, lower than that of the surrounding sediment(3.23). Phylogenetic analysis showed that bacterial phylotypes in gut content and the surrounding sediment of A. japonicus were closely related to Proteobacteria includingγ-, α-, δ-and ε-proteobacteria, Bacteroidetes, Firmicute, and Actinobacteria, of which γ-proteobacteria were predominant. These results suggested that the sea cucumber A. japonicus was capable of feeding selectively, and PCR-DGGE was applicable for characterizing the bacterial community composition in gut content and the surrounding sediment of sea cucumber. Further investigation targeting longer 16S rDNA gene fragments and/or functional genes was recommended for obtaining more information of the diversity and function of bacterial community in the gut content of sea cucumber.  相似文献   

5.
The relationship between microsatellite polymorphism and body weight of captive bred Chinese sea cucumber Apostichopus japonicus was investigated in two local populations in Dalian. Among ten loci discovered, nine show changes except for AJ07 loci. Seven loci were found highly polymorphic in both populations. For each locus in two populations, the average number of alleles is 6.428 6 and 6.285 7, the average observed heterozygosity at 0.225 7 and 0.245 9, the expected heterozygosity at 0.776 8 and 0.748 8, the polymorphism information content (PIC) at 0.709 2 and 0.674 6, respectively. Further analysis show significant correlation between A. japonicus body weight and occurrence markers AJ02 and AJ04. The findings of the relation may be helpful for molecular breeding, as well as the marker-assisted selection of sea cucumbers. Supported by the National High Technology Research and Development Program of China (863 Program, No. 2006AA10A411), the Natural Science Foundation of Liaoning Province (No.20072139), the Grant of Dalian Fisheries University, the Key Laboratory Foundation of the Educational Department of Liaoning Province (No.2009S024)  相似文献   

6.
The sea cucumber Apostichopus japonicus is a commercially and ecologically important species in China. A total of 3056 potential unigenes were generated after assembling 7597 A. japonicus expressed sequence tags (ESTs) downloaded from Gen-Bank. Two hundred and fifty microsatellite-containing ESTs (8.18%) and 299 simple sequence repeats (SSRs) were detected. The average density of SSRs was 1 per 7.403 kb of EST after redundancy elimination. Di-nucleotide repeat motifs appeared to be the most abundant type with a percentage of 69.90%. Of the 126 primer pairs designed, 90 amplified the expected products and 43 showed polymorphism in 30 individuals tested. The number of alleles per locus ranged from 2 to 26 with an average of 7.0 alleles, and the observed and expected heterozygosities varied from 0.067 to 1.000 and from 0.066 to 0.959, respectively. These new EST-derived microsatellite markers would provide sufficient polymorphism for population genetic studies and genome mapping of this sea cucumber species.  相似文献   

7.
The aquaculture of sea cucumber Apostichopus japonicus (Echinodermata, Holothuroidea) has grown rapidly during recent years and has become an important sector of the marine industry in Northern China. However, with the rapid growth of the industry and the use of non-standard culture techniques, epidemic diseases of A. japonicus now pose increasing problems to the industry. To screen the genes with stress response to bacterial infection in sea cucumber at a genome wide level, we constructed a cDNA library from A. japonicus Selenka (Aspidochirotida: Stichopodidae) after infecting them with Vibrio sp. for 48 h. Total RNA was extracted from the intestine, mesentery and coelomocyte of infected sea cucumber using Trizol and mRNA was isolated by Oligotex mRNA Kits. The ligated cDNAs were transformed into DH5α, and a library of 3.24×105 clones (3.24×105 cfu mL-1) was obtained with the sizes of inserted fragments ranging from 0.8 to 2.5 kb. Sequencing the cDNA clones resulted in a total of 1106 ESTs that passed the quality control. BlastX and BlastN searches have identified 168 (31.5%) ESTs sharing significant homology with known sequences in NCBI protein or nucleotide databases. Among a panel of 25 putative immunity-related genes, serum lectin isoform, complement component 3, complement component 3-like genes were further studied by real-time PCR and they all increased more than 5 fold in response to Vibrio sp. challenge. Our library provides a valuable molecular tool for future study of invertebrate immunity against bacterial infection and our gene expression data indicates the importance of the immune system in the evolution and development of sea cucumber.  相似文献   

8.
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9.
Fatty acids （FAs） provide energy and also can be used to trace trophic relationships among organisms. Sea cucumber Apostichopus japonicus goes into a state of aestivation during warm summer months. We examined fatty acid profiles in aestivated and non-aestivated A. japonicus using multivariate analyses （PERMANOVA, MDS, ANOSIM, and SIMPER）. The results indicate that the fatty acid profiles of aestivated and non-aestivated sea cucumbers differed significantly. The FAs that were produced by bacteria and brown kelp contributed the most to the differences in the fatty acid composition of aestivated and non- aestivated sea cucumbers. Aestivated sea cucumbers may synthesize FAs from heterotrophic bacteria during early aestivation, and long chain FAs such as eicosapentaenoic （EPA） and docosahexaenoic acid （DHA） that produced from intestinal degradation, are digested during deep aestivation. Specific changes in the fatty acid composition of A. japonicus during aestivation needs more detailed study in the future.  相似文献   

10.
Immunostimulants may improve disease resistance of aquaculture animals by promoting the nonspecific immunity response of the organisms. Five types of saccharides, including chitosan, yeast polysaccharide, burdock oligosaccharide, seaweed polysaccharide and lentinus edodes polysaccharide, were screened for potential use as immunostimulants by using spectrophotometry. The saccharides were injected into Apostichopus japonicus, a sea cucumber, and the lysozyme and superoxide dismutase (SOD) activities of the coelomic fluid and epidermal slime were monitored in six consecutive days. The results show that the lysozyme activity of the animal’s coelomic fluid was significantly stimulated on day 2, day 4 and day 6 after the injection of the saccharides (P<0.05). The effects of chitosan and yeast polysaccharide were the most notable. The lysozyme activity of the epidermal slime was significantly increased by chitosana, yeast polysaccharide, seaweed polysaccharide, and burdock oligosaccharide on day 1 and day 2 after the injection (P<0.05). The SOD activity of the coelomic fluid was significantly promoted by the saccharides on day 2 and day 4 post-injection (P<0.05), while the SOD activity of the epidermal slime increased on day 2. These findings indicate that chitosan and yeast polysaccharide are the most effective immunostimulants and potential healthy anti-disease feedstuff for A. japonicus. Supported by the National Key Technology Research and Development Program (863 Program, No. 2006BAD09A06) and the Special Fund of Chinese Central Government for Basic Scientific Research Operations in Commonweal Research Institutes (No. 02-2007B03)  相似文献   

11.
Studies were conducted to determine the cause of the acute mortality of half-smooth tongue sole Cynoglossus semilaevis Günther juveniles in a fish farm in Jimo, Shandong Province, China, in June 2006. Gross signs of the diseased tongue sole included several petechiae and ecchymoses on the body and fin necrosis and hemorrhagic lesion at the base of the fin. Bacteria were isolated from kidney, liver and hemorrhagic lesions of the diseased tongue sole. Among14 strains, SJ060621 was proved to be highly virulent to juvenile tongue sole with LD50 value of 1.0×105 colony forming units (CFU)mL-1, while the remaining 13 were avirulent. Among the 16 antibiotics tested, SJ060621 was sensitive to gentamicin and nitrofurantoin. It was identified as Listonella anguillarum with conventional plate and tube tests in combination with API 20E analysis. 16S rRNA gene and partial HSP60 gene sequenceing analysis revealed that the strain was highly homologous with L. anguillarum. Examination of the infected musculature by electron microscopy indicated numerous bacteria and lots of macrophages containing phagocytosed bacteria. Histopathological investigations revealed severe necrotic degenerative changes in the infected organs. Indirect immunofluorescence assay (IFA) was employed to detect the location of occurrence of bacteria, and bacteria were found in aggregations in the inflammatory areas in musculature.  相似文献   

12.
From May to August 2008, a large "green tide", consisting of the alga Ulva (Enteromorpha) prolifera, occurred in the Yellow Sea, China, affecting the local marine ecosystem and human activities. We investigated the influence of the green tide on the microbial community in the surface seawater, at four sites from July to August 2008, using bacterial 16S rRNA gene clone libraries. We sequenced 228 clones of unique patterns identified by restriction fragment length polymorphism (RFLP) techniques. The results show that 228 sequenced clones fell into six bacterial phyla:Proteobacteria, Bacteroidetes, Cyanobacteria, Verrucomicrobia, Actinobacteria, and Planctomycetes. Alphaproteobacteria (33%), Gammaproteobacteria (25%), Bacteroidetes (23%) and Cyanobacteria (9%) dominated the assemblage. Comparison between samples collected in July (during the tide) and those collected in August (after the tide) showed that, in the microbial community, diversities of Alphaproteobacteria and Cyanobacteria increased after the tide, while those of Gammaproteobacteria and Bacteroidetes decreased. These results indicate that the green tide influenced the growth of some bacteria, and provide information for further studies on the interactions and relationships between U. prolifera and the bacterial community. This study suggests that microbial community analysis is a good approach to monitoring green tides.  相似文献   

13.
Optimal stocking densities were investigated for the sea cucumber Apostichopus japonicus Selenka under feed-supplement and non-feed-supplement regimes in net enclosures for 333 d. Substantial weight loss occurred during the aestivation phase (AE). Decreased growth rates were also observed during the winter phase (WT). In contrast, sea cucumbers showed rapid growth during the spring (SP) and autumn (AU) phases. Feeding regimes considerably influenced the growth performance, i.e., sea cucumbers grew faster under feed-supplement regime than under non-feed-supplement regime (P < 0.05). The average survival rates of sea cucumbers under feed-supplement regime were higher than those under non-feed-supplement regime for both the autumn phase and spring phase, but the differences were only significant for the latter phase (P < 0.05). The fitted B-N curves showed that the optimal stocking densities, in terms of net production, were 22.3 ind. m-2 for feed-supplement regime and 14.1 ind. m-2 for non-feed-supplement regime.  相似文献   

14.
In this study, the intestinal microbiota of kuruma shrimp (Marsupenaeus japonicus) was examined by molecular analysis of the 16S rDNA to identify the dominant intestinal bacteria and to investigate the effects of Bacillus spp. on intestinal microbial diversity. Samples of the intestines of kuruma shrimp fed normal feed and Bacillus spp. amended feed. PCR and denaturing gradient gel electrophoresis (DGGE) analyses were then performed on DNA extracted directly from the guts. Population fingerprints of the predominant organisms were generated by DGGE analysis of the universal V3 16S rDNA amplicons, and distinct bands in the gels were sequenced. The results suggested that the gut of kuruma shrimp was dominated by Vibrio sp. and uncultured gamma proteobacterium. Overall, the results of this study suggest that PCR-DGGE is a possible method of studying the intestinal microbial diversity of shrimp.  相似文献   

15.
Leaves of terrestrial and aquatic plants are home to a wide diversity of bacterial species. However, the diversity and variability of epiphytic bacteria on their submerged plant hosts remains poorly understood. We investigated the diversity and composition of epiphytic bacteria from two common submerged macrophytes: Vallisneria natans and Hydrilla verticillata in Taihu Lake, Jiangsu, China, using methods of terminal restriction fragment length polymorphisms (T-RFLP) and clone library analyses targeted at bacterial 16S rRNA genes. The results show that: (1) the libraries of the two waterweeds contain wide phylogenetic distribution of bacteria, and that the sequences of the two libraries can be separated into 93 OTUs (at 97% similar value); (2) Betaproteobacteria, including Burkholderiales, was the most abundant bacterial group on both plants. Cyanobacteria and Gammaproteobacteria were the second largest groups on V. natans and H. verticillata, respectively. Both clone libraries included some sequences related to those of methanotrophs and nitrogen-fixing bacteria; (3) Cluster analysis of the T-RFLP profiles showed two distinct clusters corresponding to the two plant populations. Both ANOSIM of the T-RFLP data and Libshuff analysis of the two clone libraries indicated a significant difference in epiphytic bacterial communities between the two plants. Therefore, the epiphytic bacterial communities on submerged macrophytes appear to be diverse and host-specific, which may aid in understanding the ecological functions of submerged macrophytes in general.  相似文献   

16.
Sea cucumber Apostichopus japonicus samples were collected in Changdao, Penglai (PL), 27 individuals, and Lingshandao, Qingdao (QD), 30 individuals, in the Shandong Peninsula, China. Ten SSR primers were used to assess the genetic variation and relationship between and within the two stocks. Respectively, for each stock, the percentage of polymorphic bands was 85.2% and 86.9%; the gene diver- sity was 0.360 5 and 0.342 8; and the Shannon’s information index was 0.515 0 and 0.499 0. At species level, the percentage of polymorphic bands was 92.2%, the total gene diversity was 0.378 9 and the Shannon’s information index was 0.550 4. The coefficient of overall genetic differentiation and the ge- netic distances between the stocks were also calculated to be 0.073 0 and 0.079 6 using the POPGENE program. Results show that the genetic diversity of the two stocks is still large but the genetic distance between the two stocks is close. A dendrogram was constructed for the 57 individuals from the two stocks, showing that the genetic structure was unitary for PL stock but complex for QD stock.  相似文献   

17.
Lytic Characteristics and Identification of Two Alga-lysing Bacterial StrainsBeaulieu, S. E., M. R. Sengco, and D. M. Anderson, 2005. Using clay to control harmful algal blooms: deposition and resuspension of clay/algal flocs. Harmful Algae . (4): 123-138…  相似文献   

18.
The complete mitochondrial cytochrome oxidase subunit Ⅱ （COⅡ） gene of Penaeinae shrimp Fenneropenaeus chinensis was cloned and sequenced. The gene is 688 bp in length and codes for 229 amino acids. It shows 83.2%, 87.0% and 83.8% sequence similarity to Marsupenaeus Japonicus, Penaeus monodon and Farfantepenaeus notialis, respectively. The A＋T content of the whole gene and that at the third position of codons are 64.7% and 78.2%, respectively. The phylogenetic relationship between F. chinensis and three other species representing genera Farfanatepenaeus, Marsupenaeus and Penaeus was analyzed. Results showed that the genetic distances among the four taxa ranged from 0.144 0 to 0.200 5, exceeding those estimated with COⅠ and partial 16S rRNA gene sequences among Marsupenaeus, Litopenaeus and Melicertus, and being therefore larger than the value among subgenera. It has been suggested that the COⅡ gene has a faster evolutionary rate than that of the COⅠ gene and partial 16S rRNA gene and could be used for phylogenetic analysis at genus or species level. The results of the present study indicated that Farfantepenaeus, Fenneropenaeus, Marsupenaeus and Penaeus are at a higher phylogenetic level than subgenus, which supports the opinion of the elevation of phylogenetic status of the four subgenera to genus level.  相似文献   

19.
Marine sponge Tedania anhelans distributes throughout the intertidal zone of Fujian, southeastern China, and is a potential source of natural bioactive products. The sponge harbors a large number of bacterial groups that have been identified using various techniques, including fluorescent in situ hybridization (FISH). Fractionation of dissociated sponge allowed isolation of 25 bacterial species. Based on 16S rRNA gene sequencing, phylogenetic analysis attributed most of these eubacteria to α-Proteobacteria, γ-Proteobacteria, Cytophaga/Flavobacterium/Bacteroidetes (CFB group), and the family Bacillaceae of Gram-positive bacteria. In sequence similarity, five putatively novel species were identified with less than 98% similarity to other strains in the NCBI database. Tests for antimicrobial activities were performed against Gram-positive bacteria, Gram-negative bacteria, fungi, antitumor indicators Escherichia coli 343/591 (with DNA repair deficiency), regular E. coli 343/636 (with different DNA repair capacity), and 10 bacterial isolates exhibited inhibitory bioactivities. Among these strains, three isolates were detected involving function gene NRPS-A domains, which were most closely related to the amino acid sequences of linear gramicidin synthetase and pyoverdine synthetase. These results contribute to our knowledge of the microbes associated with marine sponges and further reveal novel bacterial resources for the screening of bioactive marine natural products.  相似文献   

20.
Gut microorganisms play an important role in the digestion of their host animals. The purpose of this research was to isolate and assess the enzyme-producing microbes from the Apostichopus japonicus gut. Thirty-nine strains that can produce at least one of the three digestive enzymes (protease, amylase, and cellulase) were qualitatively screened based on their extracellular enzyme-producing abilities. The enzyme-producing strains clustered into eight groups at the genetic similarity level of 100% by analyzing the restriction patterns of 16S rDNA amplified with Mbo I. Phylogenetic analysis revealed that 37 strains belonged to the genus Bacillus and two were members of the genus Virgibacillus. Enzyme-producing capability results indicate that the main enzyme-producing microflora in the A. japonicus gut was Bacillus, which can produce protease, amylase, and cellulase. Virgibacillus, however, can only produce protease. The high enzyme-producing capability of the isolates suggests that the gut microbiota play an important role in the sea cucumber digestive process.  相似文献   

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