- Email: [email protected]
Study on restoring the genetic diversity of the Japanese scallop (Mizuhopecten yessoensis) in China
Abstracts / Journal of Biotechnology 136S (2008) S548–S557
1.9 × 107 to 6.3 × 107 , 1.4 × 107 to 2.3 × 108 , 1.72 × 107 to 1.62 × 108 , and 2.2 × 107 to 6.4 × 108 CFU/mL, respectively, at Pyoseon area, 1.9 × 107 to 2.7 × 107 , 1.3 × 107 to 5.6 × 108 , 1.46 × 107 to 8.0 × 108 , and 2.4 × 107 to 5.1 × 108 CFU/mL, respectively, at Wimi area, and at Daejoung area, those were 1.2 × 107 to 5.1 × 107 , 1.4 × 107 to 4.5 × 107 , 3.4 × 107 to 8 × 108 , and 2.2 × 107 to 9.5 × 108 CFU/mL, respectively. It was shown that 12 strains of Bacillus sp. (2 strains), Staphylococcus sp. (2 strains), Streptomyces sp. (1 strain), Micrococcus sp. (1 strain), Roseovarius sp. (1 strain), Acinetobacteria sp. (1 strain), Enterobacter sp. (1 strain), Psudoalteromonas sp. (2 strains), Tenabaculum sp. (1 strain) were distributed in the sediment samples at seoungsan, 19 strains of Shwanella sp. (2 strains), Vibrio sp. (1 strain), Krokinobacter sp. (1 strain), Brevundimonas sp. (1 strain), Erythrobacter sp. (1 strain), Jannaschia sp. (1 strain), Micrococcus sp. (1 strain), Streptomyces sp. (3 strains), Paenibacillus sp. (1 strain), Bacillus sp. (7 strains) at Pyoseon area, 12 strains of Staphylococcus sp. (1 strain), Salinicoccus sp. (1 strain), Bacillus sp. (1 strain), Streptomyces sp. (1 strain), Dietzia sp. (1 strain), Micrococcus sp. (1 strain), Vibrio sp. (1 strain), Alterromonas sp. (1 strain), Shewanella sp. (1 strain), Roseovarius sp. (1 strain), Tenacibaculum sp. (1 strain), Muricauda sp. (1 strain) at Wimi area, and 12 strains of Bacillus sp. (4 strains), Salinicoccus sp. (1 strain), Streptomyces sp. (1 strain), Vibrio sp. (1 strain), Pseudoalteromonas sp. (4 strains), Tenacibaculum sp. (1 strain) at Daejoung area.
Acknowledgement Supported by grants from JETC doi:10.1016/j.jbiotec.2008.07.1304 VI3-P-019 Novel bacterial surface display systems based on outer membrane anchoring elements from marine bacterium Vibrio anguillarum Zhao Yang, Qin Liu, Qiyao Wang, Yuanxing Zhang ∗ State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, China E-mail address: [email protected] (Y. Zhang). Surface display of heterologous peptides and proteins such as receptors, antigens, and enzymes on live bacterial cells is of considerable value for various biotechnological and industrial applications. In this work, a series of novel cell surface display systems were examined by employing Vibrio anguillarum outer membrane protein and outer membrane lipoprotein as anchoring motifs. These display systems consist of: (1) the signal sequence and first eleven N-terminal amino acids of V. anguillarum outer membrane lipoprotein Wza, or the signal sequence and first nine N-terminal amino acids of the mature major Escherichia coli lipoprotein Lpp; (2) transmembrane domains of V. anguillarum outer membrane proteins Omporf1, OmpU or Omp26La. In order to assay the translocation efficiency of constructed display systems in bacteria, green fluorescent protein (GFP) was inserted to the systems and the results of GFP surface localization confirmed that four of the six surface display systems could successfully display GFP on Escherichia coli surface. For assaying its potential application in live bacteria carrier vaccines, an excellent display system Wza-Omporf1 was fused with the major capsid protein (MCP) of large yellow croaker iridovirus (LYCIV) and introduced into an attenuated V. anguillarum strain
S555
MVAV6203, and subsequent analysis of MCP surface localization proved that the novel display system Wza-Omporf1 could function as a strong tool in V. anguillarum carrier vaccines development. References Drummelsmith, J., Whitfield, C., 2000. Translocation of group 1 capsular polysaccharide to the surface of Escherichia coli requires a multimeric complex in the outer membrane. EMBO J. 19, 57–66. Francisco, J.A., Earhart, C.F., Georgiou, G., 1992. Transport and anchoring of betalactamase to the external surface of Escherichia coli. Proc. Natl. Acad. Sci. U.S.A. 89, 2713–2717. Georgiou, G., Stephens, D.L., Stathopoulos, C., Poetschke, H.L., Mendenhall, J., Earhart, C.F., 1996. Display of beta-lactamase on the Escherichia coli surface: outer membrane phenotypes conferred by Lpp -OmpA -beta-lactamase fusions. Protein Eng. 9, 239–247. Georgiou, G., Stathopoulos, C., Daugherty, P.S., Nayak, A.R., Iverson, B.L., Curtiss, R., 1997. Display of heterologous proteins on the surface of microorganisms: from the screening of combinatorial libraries to live recombinant vaccines. Nat. Biotechnol. 15, 29–34.
doi:10.1016/j.jbiotec.2008.07.1305 VI3-P-021 Study on restoring the genetic diversity of the Japanese scallop (Mizuhopecten yessoensis) in China Weidong Liu 1,2,∗ , Wenbo Wang 2 , Xiangbo Bao 2 , Wei Zhang 1 , Xingju Yu 1 1
Dalian Institute of Chemical Physics, CAS, Dalian 116023, China Liaoning Ocean and Fisheries Science Research Institute, Dalian 116023, China 2
E-mail address: [email protected] (W. Liu). The Japanese scallop Patinopecten yessoensis was introduced form Japan to China in 1982. Because of the limited number of foundation population, the genetic variability has been lost during hatchery production. For restoring the genetic diversity of Japanese scallop in China, some means and steps for seed yielding, such as re-introducing Japanese wild population from Japanese as parents or hybridizing Chinese hatchery population (♀) with Japanese wild population (♂), were adopted by hatching plant. In this study, seven microsatellite loci were used to investigate the genetic variations of two different populations, namely, one Japanese hatchery population which parents came from Japanese wild population, one hybrid population from Chinese hatchery population (♀) and Japanese wild population (♂), and to compare with one Chinese hatchery population and one natural seed population from Dalian in China. The average effective allele numbers per locus (Ae) of the above corresponding populations were 3.753, 3.523, 3.156 and 3.593, the average expected heterozygosities (He) were 0.701, 0.696, 0.669 and 0.701, respectively. These data explained that the seed breed from Japanese wild population as parents had obvious high level genetic diversity than the Chinese hatchery population, and except for the Chinese hatchery population, the genetic variations of the others had no obvious variance. References Appleyard, S.A., Renwick, J.M., Mather, P.B., 2001. Individual heterozygosity levels and relative growth performance in Oreochromis niloticus (L.) cultured under Fijian conditions. Aquacult. Res. 32, 287–296. Knibb, W., 2000. Genetic improvement of marine fish—which method for industry? Aquacult. Res. 31, 11–23. Li, Q., Xu, K.F., Yu, R.H., 2007. Genetic variation in Chinese hatchery populations of the Japanese scallop (Patinopecten yessoensis) inferred from microsatellite data. Aquaculture 269, 211–219.
S556
Abstracts / Journal of Biotechnology 136S (2008) S548–S557
Wang, L.L., Zhang, H., Song, L.S., Guo, X.M., 2007. Loss of allele diversity in introduced populations of the hermaphroditic bay scallop Argopecten irradians. Aquaculture 271, 252–259.
doi:10.1016/j.jbiotec.2008.07.1306 VI3-P-022 Abalone NF-B transcription factor requires a nuclear localization signal for nuclear import Yusheng Zhang 1
Jiang 2,∗ , Jinyan
Shang 1 , Xinzhong
Wu 1,2 , Chuanxi
1
Zhejiang University, Hangzhou 310029, PR China College of Life Sciences and Technology, Dalian Fisheries University, Dalian116023, PR China 2
E-mail address: [email protected] (Y. Jiang). Abalone, Haliotis diversicolor supertexta is economically important marine gastropod cultured in the south coast of China. However, this industry has often been devastated by disease outbreak. Up to date, few studies on the molecular mechanism of immunology were reported on this ancient invertebrate. A homologue of Rel\NFB transcription factor, named Ab-Rel, was identified from this species, and proved to play a possible role in the immune response (Jiang and Wu, 2007; Jiang et al., 2007). Since the DNA-binding capacity of NF-B in hemocytes can be specifically up-regulated by immune challenge, it is presumed that proper nuclear transport of Ab-Rel was occurred within this process (Birbach et al., 2002). To investigate the mechanisms of nuclear import, we analyzed the effect of truncated mutations on subcellular distribution of Ab-RelEGFP using the modified bac to bac insect cell expression system. It demonstrated that a consensus nuclear localization signal (NLS) is involved in the translocation event. Our data provide further evidence that Ab-Rel is functionally conserved as one of Rel\NF-B members, allowing us to study this important transcription factor in an evolutionary context. References Birbach, A., Gold, P., Binder, B.R., Hofer, E., Martin, R.D., Schmid, J.A., 2002. Signaling molecules of the NF-B pathway shuttle constitutively between cytoplasm and nucleu. J. Biol. Chem. 277, 10842–10851. Jiang, Y.S., Wu, X.Z., 2007. Characterization of a Rel\NF-B homologue in a gastropod abalone, Haliotis diversicolor supertexta. Dev. Comp. Immunol. 31 (2), 121–131. Jiang, Y.S., Wu, X.Z., Zhang, Y., 2007. Magnetocapture of abalone transcription factor NF-B: a new strategy for isolation and detection of NF-B both in vitro and in vivo. J. Biotechnol. 127 (3), 385–391.
doi:10.1016/j.jbiotec.2008.07.1307 VI3-P-023 Abalone NF-B transcription factor is composed of two different subunits Yusheng Jiang 2,∗ , Junfeng Li 2 , Xinzhong Wu 1,2 , Yue Ma 1,2 , Gang Liu 2 1
College of Animal Sciences, Zhejiang University, Hangzhou 310029, PR China 2 College of Life Sciences and Technology, Dalian Fisheries University, Dalian 116023, PR China E-mail address: [email protected] (Y. Jiang). Rel\NF-B signal transduction pathway is evolutionarily conserved and involved in numerous biological processes (Hayden and Ghosh, 2004). Previously, we cloned and functionally characterized a
homologue of Rel\NF-B transcription factor in a gastropod Haliotis diversicolor supertexta, named Ab-Rel (Jiang and Wu, 2007). Using an antibody raised against the Ab-Rel RHD (Rel homology domain) which is the signature motif of the Rel families, detected two specific immunoreactive bands on the Western blot, leading us to extrapolate that abalone NF-B is consisted of two subunits. To confirm this hypothesis, a plasmid containing a specific fragment corresponding to transactivation domain (TAD) of Ab-Rel was constructed to express recombinant protein in E. coli. The expressed products were purified to immunize rabbits, and the antibody was obtained with the titer determined by an indirect ELISA. Western blot analysis of total extracts of abalone haemocytes using the AbRel specific antibody detected one specific immunoreactive band. These results are consistent with northern blot analysis using the nucleotide probe to Ab-Rel RHD or TAD. Taken together, all results indicated that Ab-Rel is one of the two subunits of abalone NFB (Huxford et al., 1998). This is the first report on the molecular structure of NF-B protein in shellfish. References Hayden, M.S., Ghosh, S., 2004. Signaling to NF-B. Genes Dev. 18, 2195–2224. Huxford, T., Huang, De-B., Malek, S., Ghosh, G., 1998. The crystal structure of the IBa/NF-B complex reveals mechanisms of NF-B inactivation. Cell 95, 759–770. Jiang, Y.S., Wu, X.Z., 2007. Characterization of a Rel\NF-B homologue in a gastropod abalone, Haliotis diversicolor supertexta. Dev. Comp. Immunol. 31 (2), 121–131.
doi:10.1016/j.jbiotec.2008.07.1308 VI3-P-024 Effect of l-ascorbic acid as immunity enhancer for juvenile sea cucumber, Apostichopus japonicus Kailai Wang ∗ , Ziru Liu, Yongping Xu, Gang Liu, Tongju Ren, Liji Jin School of Environmental and Biological Science and Technology, Dalian University of Technology, Dalian, 116024, People’s Republic of China E-mail address: [email protected] (K. Wang). A trail was conducted to determine the efficiency of vitamin C (ascorbic acid, AsA) as an immunity enhancer to stimulate the non-specific immune response and to protect juvenile sea cucumbers from infectious pathogen. Five diets of different AsA levels were fed to five groups of juvenile sea cucumbers (G1—0, G2—487, G3—1372, G4—4836, G5—13,857 mg/kg diet), with an average initial weight 1.19 ± 0.46 g. During the 40 days feeding trial, higher but not significantly survival rates were observed in groups G3, G4, G5 than the AsA free group G1. The bactericidal activity of skin mucus and coelom fluid (CF) of sea cucumbers from group G3, G4 were found significantly (P < 0.05) enhanced. Moreover, a significantly higher lysozyme activity of CF was showed in G3. The coelom cytocrit demonstrated an increased trend along with the increased diet AsA level, while the coelom cells of G4, G5 showed significantly higher phagocytic activity and respiratory burst activity. A greater CF lectin titer was described significantly in G4, and higher mucus lectin titer activity was observed in animals from G3, G4 and G5. In G1, both alkaline phosphatase (ALP) and acid phosphatase (ACP) were found to possess a significantly lower activity than AsA containing groups. However, there was no significant difference observed in phenoloxidase activity in all the five groups. Bacterial challenged study showed that typical infection signs were first observed among the five groups, respectively, on day 4, 6, 8,
∗ Corresponding author. Tel.: +86 411 84706359; fax: +86 411 84706359.
1.9 × 107 to 6.3 × 107 , 1.4 × 107 to 2.3 × 108 , 1.72 × 107 to 1.62 × 108 , and 2.2 × 107 to 6.4 × 108 CFU/mL, respectively, at Pyoseon area, 1.9 × 107 to 2.7 × 107 , 1.3 × 107 to 5.6 × 108 , 1.46 × 107 to 8.0 × 108 , and 2.4 × 107 to 5.1 × 108 CFU/mL, respectively, at Wimi area, and at Daejoung area, those were 1.2 × 107 to 5.1 × 107 , 1.4 × 107 to 4.5 × 107 , 3.4 × 107 to 8 × 108 , and 2.2 × 107 to 9.5 × 108 CFU/mL, respectively. It was shown that 12 strains of Bacillus sp. (2 strains), Staphylococcus sp. (2 strains), Streptomyces sp. (1 strain), Micrococcus sp. (1 strain), Roseovarius sp. (1 strain), Acinetobacteria sp. (1 strain), Enterobacter sp. (1 strain), Psudoalteromonas sp. (2 strains), Tenabaculum sp. (1 strain) were distributed in the sediment samples at seoungsan, 19 strains of Shwanella sp. (2 strains), Vibrio sp. (1 strain), Krokinobacter sp. (1 strain), Brevundimonas sp. (1 strain), Erythrobacter sp. (1 strain), Jannaschia sp. (1 strain), Micrococcus sp. (1 strain), Streptomyces sp. (3 strains), Paenibacillus sp. (1 strain), Bacillus sp. (7 strains) at Pyoseon area, 12 strains of Staphylococcus sp. (1 strain), Salinicoccus sp. (1 strain), Bacillus sp. (1 strain), Streptomyces sp. (1 strain), Dietzia sp. (1 strain), Micrococcus sp. (1 strain), Vibrio sp. (1 strain), Alterromonas sp. (1 strain), Shewanella sp. (1 strain), Roseovarius sp. (1 strain), Tenacibaculum sp. (1 strain), Muricauda sp. (1 strain) at Wimi area, and 12 strains of Bacillus sp. (4 strains), Salinicoccus sp. (1 strain), Streptomyces sp. (1 strain), Vibrio sp. (1 strain), Pseudoalteromonas sp. (4 strains), Tenacibaculum sp. (1 strain) at Daejoung area.
Acknowledgement Supported by grants from JETC doi:10.1016/j.jbiotec.2008.07.1304 VI3-P-019 Novel bacterial surface display systems based on outer membrane anchoring elements from marine bacterium Vibrio anguillarum Zhao Yang, Qin Liu, Qiyao Wang, Yuanxing Zhang ∗ State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, China E-mail address: [email protected] (Y. Zhang). Surface display of heterologous peptides and proteins such as receptors, antigens, and enzymes on live bacterial cells is of considerable value for various biotechnological and industrial applications. In this work, a series of novel cell surface display systems were examined by employing Vibrio anguillarum outer membrane protein and outer membrane lipoprotein as anchoring motifs. These display systems consist of: (1) the signal sequence and first eleven N-terminal amino acids of V. anguillarum outer membrane lipoprotein Wza, or the signal sequence and first nine N-terminal amino acids of the mature major Escherichia coli lipoprotein Lpp; (2) transmembrane domains of V. anguillarum outer membrane proteins Omporf1, OmpU or Omp26La. In order to assay the translocation efficiency of constructed display systems in bacteria, green fluorescent protein (GFP) was inserted to the systems and the results of GFP surface localization confirmed that four of the six surface display systems could successfully display GFP on Escherichia coli surface. For assaying its potential application in live bacteria carrier vaccines, an excellent display system Wza-Omporf1 was fused with the major capsid protein (MCP) of large yellow croaker iridovirus (LYCIV) and introduced into an attenuated V. anguillarum strain
S555
MVAV6203, and subsequent analysis of MCP surface localization proved that the novel display system Wza-Omporf1 could function as a strong tool in V. anguillarum carrier vaccines development. References Drummelsmith, J., Whitfield, C., 2000. Translocation of group 1 capsular polysaccharide to the surface of Escherichia coli requires a multimeric complex in the outer membrane. EMBO J. 19, 57–66. Francisco, J.A., Earhart, C.F., Georgiou, G., 1992. Transport and anchoring of betalactamase to the external surface of Escherichia coli. Proc. Natl. Acad. Sci. U.S.A. 89, 2713–2717. Georgiou, G., Stephens, D.L., Stathopoulos, C., Poetschke, H.L., Mendenhall, J., Earhart, C.F., 1996. Display of beta-lactamase on the Escherichia coli surface: outer membrane phenotypes conferred by Lpp -OmpA -beta-lactamase fusions. Protein Eng. 9, 239–247. Georgiou, G., Stathopoulos, C., Daugherty, P.S., Nayak, A.R., Iverson, B.L., Curtiss, R., 1997. Display of heterologous proteins on the surface of microorganisms: from the screening of combinatorial libraries to live recombinant vaccines. Nat. Biotechnol. 15, 29–34.
doi:10.1016/j.jbiotec.2008.07.1305 VI3-P-021 Study on restoring the genetic diversity of the Japanese scallop (Mizuhopecten yessoensis) in China Weidong Liu 1,2,∗ , Wenbo Wang 2 , Xiangbo Bao 2 , Wei Zhang 1 , Xingju Yu 1 1
Dalian Institute of Chemical Physics, CAS, Dalian 116023, China Liaoning Ocean and Fisheries Science Research Institute, Dalian 116023, China 2
E-mail address: [email protected] (W. Liu). The Japanese scallop Patinopecten yessoensis was introduced form Japan to China in 1982. Because of the limited number of foundation population, the genetic variability has been lost during hatchery production. For restoring the genetic diversity of Japanese scallop in China, some means and steps for seed yielding, such as re-introducing Japanese wild population from Japanese as parents or hybridizing Chinese hatchery population (♀) with Japanese wild population (♂), were adopted by hatching plant. In this study, seven microsatellite loci were used to investigate the genetic variations of two different populations, namely, one Japanese hatchery population which parents came from Japanese wild population, one hybrid population from Chinese hatchery population (♀) and Japanese wild population (♂), and to compare with one Chinese hatchery population and one natural seed population from Dalian in China. The average effective allele numbers per locus (Ae) of the above corresponding populations were 3.753, 3.523, 3.156 and 3.593, the average expected heterozygosities (He) were 0.701, 0.696, 0.669 and 0.701, respectively. These data explained that the seed breed from Japanese wild population as parents had obvious high level genetic diversity than the Chinese hatchery population, and except for the Chinese hatchery population, the genetic variations of the others had no obvious variance. References Appleyard, S.A., Renwick, J.M., Mather, P.B., 2001. Individual heterozygosity levels and relative growth performance in Oreochromis niloticus (L.) cultured under Fijian conditions. Aquacult. Res. 32, 287–296. Knibb, W., 2000. Genetic improvement of marine fish—which method for industry? Aquacult. Res. 31, 11–23. Li, Q., Xu, K.F., Yu, R.H., 2007. Genetic variation in Chinese hatchery populations of the Japanese scallop (Patinopecten yessoensis) inferred from microsatellite data. Aquaculture 269, 211–219.
S556
Abstracts / Journal of Biotechnology 136S (2008) S548–S557
Wang, L.L., Zhang, H., Song, L.S., Guo, X.M., 2007. Loss of allele diversity in introduced populations of the hermaphroditic bay scallop Argopecten irradians. Aquaculture 271, 252–259.
doi:10.1016/j.jbiotec.2008.07.1306 VI3-P-022 Abalone NF-B transcription factor requires a nuclear localization signal for nuclear import Yusheng Zhang 1
Jiang 2,∗ , Jinyan
Shang 1 , Xinzhong
Wu 1,2 , Chuanxi
1
Zhejiang University, Hangzhou 310029, PR China College of Life Sciences and Technology, Dalian Fisheries University, Dalian116023, PR China 2
E-mail address: [email protected] (Y. Jiang). Abalone, Haliotis diversicolor supertexta is economically important marine gastropod cultured in the south coast of China. However, this industry has often been devastated by disease outbreak. Up to date, few studies on the molecular mechanism of immunology were reported on this ancient invertebrate. A homologue of Rel\NFB transcription factor, named Ab-Rel, was identified from this species, and proved to play a possible role in the immune response (Jiang and Wu, 2007; Jiang et al., 2007). Since the DNA-binding capacity of NF-B in hemocytes can be specifically up-regulated by immune challenge, it is presumed that proper nuclear transport of Ab-Rel was occurred within this process (Birbach et al., 2002). To investigate the mechanisms of nuclear import, we analyzed the effect of truncated mutations on subcellular distribution of Ab-RelEGFP using the modified bac to bac insect cell expression system. It demonstrated that a consensus nuclear localization signal (NLS) is involved in the translocation event. Our data provide further evidence that Ab-Rel is functionally conserved as one of Rel\NF-B members, allowing us to study this important transcription factor in an evolutionary context. References Birbach, A., Gold, P., Binder, B.R., Hofer, E., Martin, R.D., Schmid, J.A., 2002. Signaling molecules of the NF-B pathway shuttle constitutively between cytoplasm and nucleu. J. Biol. Chem. 277, 10842–10851. Jiang, Y.S., Wu, X.Z., 2007. Characterization of a Rel\NF-B homologue in a gastropod abalone, Haliotis diversicolor supertexta. Dev. Comp. Immunol. 31 (2), 121–131. Jiang, Y.S., Wu, X.Z., Zhang, Y., 2007. Magnetocapture of abalone transcription factor NF-B: a new strategy for isolation and detection of NF-B both in vitro and in vivo. J. Biotechnol. 127 (3), 385–391.
doi:10.1016/j.jbiotec.2008.07.1307 VI3-P-023 Abalone NF-B transcription factor is composed of two different subunits Yusheng Jiang 2,∗ , Junfeng Li 2 , Xinzhong Wu 1,2 , Yue Ma 1,2 , Gang Liu 2 1
College of Animal Sciences, Zhejiang University, Hangzhou 310029, PR China 2 College of Life Sciences and Technology, Dalian Fisheries University, Dalian 116023, PR China E-mail address: [email protected] (Y. Jiang). Rel\NF-B signal transduction pathway is evolutionarily conserved and involved in numerous biological processes (Hayden and Ghosh, 2004). Previously, we cloned and functionally characterized a
homologue of Rel\NF-B transcription factor in a gastropod Haliotis diversicolor supertexta, named Ab-Rel (Jiang and Wu, 2007). Using an antibody raised against the Ab-Rel RHD (Rel homology domain) which is the signature motif of the Rel families, detected two specific immunoreactive bands on the Western blot, leading us to extrapolate that abalone NF-B is consisted of two subunits. To confirm this hypothesis, a plasmid containing a specific fragment corresponding to transactivation domain (TAD) of Ab-Rel was constructed to express recombinant protein in E. coli. The expressed products were purified to immunize rabbits, and the antibody was obtained with the titer determined by an indirect ELISA. Western blot analysis of total extracts of abalone haemocytes using the AbRel specific antibody detected one specific immunoreactive band. These results are consistent with northern blot analysis using the nucleotide probe to Ab-Rel RHD or TAD. Taken together, all results indicated that Ab-Rel is one of the two subunits of abalone NFB (Huxford et al., 1998). This is the first report on the molecular structure of NF-B protein in shellfish. References Hayden, M.S., Ghosh, S., 2004. Signaling to NF-B. Genes Dev. 18, 2195–2224. Huxford, T., Huang, De-B., Malek, S., Ghosh, G., 1998. The crystal structure of the IBa/NF-B complex reveals mechanisms of NF-B inactivation. Cell 95, 759–770. Jiang, Y.S., Wu, X.Z., 2007. Characterization of a Rel\NF-B homologue in a gastropod abalone, Haliotis diversicolor supertexta. Dev. Comp. Immunol. 31 (2), 121–131.
doi:10.1016/j.jbiotec.2008.07.1308 VI3-P-024 Effect of l-ascorbic acid as immunity enhancer for juvenile sea cucumber, Apostichopus japonicus Kailai Wang ∗ , Ziru Liu, Yongping Xu, Gang Liu, Tongju Ren, Liji Jin School of Environmental and Biological Science and Technology, Dalian University of Technology, Dalian, 116024, People’s Republic of China E-mail address: [email protected] (K. Wang). A trail was conducted to determine the efficiency of vitamin C (ascorbic acid, AsA) as an immunity enhancer to stimulate the non-specific immune response and to protect juvenile sea cucumbers from infectious pathogen. Five diets of different AsA levels were fed to five groups of juvenile sea cucumbers (G1—0, G2—487, G3—1372, G4—4836, G5—13,857 mg/kg diet), with an average initial weight 1.19 ± 0.46 g. During the 40 days feeding trial, higher but not significantly survival rates were observed in groups G3, G4, G5 than the AsA free group G1. The bactericidal activity of skin mucus and coelom fluid (CF) of sea cucumbers from group G3, G4 were found significantly (P < 0.05) enhanced. Moreover, a significantly higher lysozyme activity of CF was showed in G3. The coelom cytocrit demonstrated an increased trend along with the increased diet AsA level, while the coelom cells of G4, G5 showed significantly higher phagocytic activity and respiratory burst activity. A greater CF lectin titer was described significantly in G4, and higher mucus lectin titer activity was observed in animals from G3, G4 and G5. In G1, both alkaline phosphatase (ALP) and acid phosphatase (ACP) were found to possess a significantly lower activity than AsA containing groups. However, there was no significant difference observed in phenoloxidase activity in all the five groups. Bacterial challenged study showed that typical infection signs were first observed among the five groups, respectively, on day 4, 6, 8,
∗ Corresponding author. Tel.: +86 411 84706359; fax: +86 411 84706359.