• Journal of Microbiology and Biotechnology
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• v.19 no.9
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• pp.982-986
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• 2009

For the production of ghost bacteria vaccine to prevent the streptococcal disease in aquaculture fish species, a double cassettes vector was constructed and cloned in Escherichia coli DH5${\alpha}$. Ghost bacteria vaccine production from Escherichia coli DH5${\alpha}$/pHCE-InaN-GAPDH-Ghost 37 SDM (SIG) was maximized at a glucose concentration of 1 g/l, agitation of 300 rpm, and aeration of 1 vvm. The maximal efficiency of ghost bacteria formation was obtained at the mid-exponential phase ($OD_{600}=2.0$) with the concentration of 0.77 g/l for SIG. The molecular mass of GAPDH was detected at 67 kDa with the insoluble fraction, by SDS-PAGE and Western blot. The protective efficacy of ghost bacteria vaccine was evaluated by challenge test using olive flounder. The cumulative mortalities of the positive control, formalin-killed cell (FKC) vaccine, and SIG vaccine immunized groups were 91%, 74%, and 57%, respectively. These results suggest that SIG vaccine showed efficacy as a vaccine and had a higher potential to induce protective antibodies than did FKC vaccine.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.50 no.3
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• pp.302-310
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• 2017

Interferon regulatory factor 8 (IRF8) is essential for the development of B and T cells, as well as for the activity of dendritic cells and macrophages. We performed molecular characterization of IRF8 from rock fish, Sebastes schlegelii (Ss), and investigated the spatial and temporal profile of mRNA expression after challenge with lipopolysaccharide (LPS), polyinosinic:polycytidylic acid (poly I:C), or Streptococcus iniae. The full-length cDNA sequence of SsIRF8 was 1,657 bp, containing an ORF of 1,266 bp. The gene had a predicted molecular mass of 47.7 kDa and an isoelectric point of 5.99. The amino acid sequence coded by this gene showed the highest degree of identity (90.8%) and similarity (96.2%) with IRF8 from Oplegnathus fasciatus. The SsIRF8 mRNA was expressed ubiquitously, at varying levels, with the highest level of expression observed in the spleen. To confirm the role of SsIRF8 in mediating the immune response, we measured SsIRF8 mRNA expression in the splenic tissue at different time points after injection with LPS, poly I:C, or S. iniae. The qRT-PCR results showed that SsIRF8 mRNA expression in the poly I:C-injected group was highly upregulated 6 hr after exposure (P<0.05). Expression of SsIRF8 mRNA in the S. iniae-injected group peaked at 24 hr. These results suggest that SsIRF8 might be important in regulating the strength of the rockfish immune response to immunostimulatory agents.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.35 no.6
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• pp.578-582
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• 2002

Ethanol extract obtained from bark, roots, stem, leaves and seeds of 30 species of plants and mushroom, Dicyophora indusiata were examined antimicrobial effect on fish pathogenic bacteria, Listoneria anguillamm, Lactococcus garvieae, Streptococcus iniae by disk method. Ethanol extract of D. indusiata was found to be the most active against all fish pathogenic bacteria in test system. Ethyl acetate fraction from ethanol extract of D. indusiata showed the strongest compared with those from other solvent fractions such as dichloromethane, n-buthanol and water. The antimicrobial effect of ethyl acetate fraction was particularly evident against S. iniae.

• Food Science and Preservation
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• v.22 no.6
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• pp.886-892
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• 2015

The antimicrobial effects of ten isothiocyanates (ITCs) present in cruciferous vegetables and radish root hydrolysate were investigated against pathogenic bacteria from olive flounder. Minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were measured against two gram-positive bacterial strains (Streptococcus parauberis, S. iniae) and four gram-negative bacterial strains (Edwardsiella tarda, Vibrio ichthyoenteri, V. harveyi, Photobacterium damselae) by using a broth microdilution technique. The antibacterial activity of ITCs was in the order sulforaphane > sulforaphene > phenylethyl ITC > erucin > benzyl ITC > iberin > I3C > allyl ITC > phenyl ITC > hexyl ITC. The susceptibility of fish pathogens to ITCs was in the order of V. harveyi > E. tarda > P. damselae > S. parauberis > S. iniae > V. ichthyoenteri. Antimicrobial activity (MIC) of radish root hydrolysate was 0.250 mg/mL against S. iniae, 0.438 mg/mL against S. parauberis, and 0.500 mg/mL against both E. tarda and V. harveyi. The aliphatic ITCs were potent inhibitors of the growth of fish pathogens, followed by aromatic ITCs and indolyl ITC. The presence of a double bond in the chemical structure of ITCs decreased antibacterial activity, while ITCs with a thiol (-S-) group and a longer carbon chain increased antibacterial activity. These results suggest that ITCs have strong antibacterial activities and may be useful in the prevention of fish pathogens.

• Journal of Life Science
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• v.19 no.7
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• pp.845-851
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• 2009

A vector harboring double cassettes; a heterologous gene expression cassette of pHCE-InaN-antigen and a ghost formation cassette of pAPR-cI-E lysis 37 SDM was constructed and introduced to E. coli DH5a. For the production of a bacterial ghost vaccine, bacterial ghosts from E. coli / Streptococcus iniae with four different types of antigens - enolase, GAPDH, sagA and piaA - were produced by the optimization of fermentation parameters such as a glucose concentration of 1 g/l, agitation of 300 rpm and aeration of 1 vvm. Efficiency of ghost bacteria formation was evaluated with cultures of OD$_{600}$=1.0, 2.0 and 3.0. The efficiency of the ghost bacteria formation was 99.54, 99.67, 99.99 and 99.99% with inductions at OD$_{600}$=3.0, 1.0, 2.0 and 1.0 for E. coli/S. iniae antigens enolase, piaA, GAPDH and sagA, respectively. Ghost bacteria as a vaccine was harvested by centrifugation. The antigen protein expressions were analyzed by SDS-PAGE and western blot analysis, and the molecular weights of the enolase, piaA, GAPDH and sagA were 78, 26, 67 and 26 kDa, respectively. The molecular weights of the expressed antigens were consistent with theoretical sizes obtained from the amino acid sequences.

• Journal of fish pathology
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• v.6 no.2
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• pp.197-204
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• 1993

Flounder culture has been developed mainly in the western parts of japan, and, to date, following six bacterial diseases have been reported. Bacterial white enteritis occurs in 16 to 30-day-old flounder larvae and often causes mass mortality in seed production. Bacterium named Vibrio sp. INFL invades and multiplies in the mucosae of posterier part of intestine, and causes desquamative enteritis. Gliding bacterial disease occurs mostly in juvenile stage and in spring to summer. Diseased signs are partial discoloration and erosion of skin and fins. Histologically, epidermis are removed, and the causative bacterium, Flexibacter maritimus, multiplies on the surface of demis and invades into the muscular tissue. Vibriosis caused by Vibrio anguillarum and related organisum is one of the well-known diseases among marine fish. Outbreaks of the disease in flounder culture are relatively few, but mass mortalities in fingerlings due to the disease were reported. An outbreak of nocardiosis in the autumn of 1984 has been reported, but since then the disease scarcely occurred. The disease is characterized by formation of abscesses under the skin and white nodes in the gill, heart, spleen and kidney. Streptococcicosis occurs frequently in recent years. Beta-hemolytic streptococcus is the causative bacterium, which possesses the same biochemical and serological characteristics as $\beta$-streptococci isolated from some marine and freshwater fish, and is seemed to related to Streptococcus iniae. Edwardsiellosis is the disease that causes most damage in flounder culture in Japan. Characteristic symptoms are swelling of abdomen and intestinal protrusion from the anus due to accumulation of ascites. Edwardsiella tarda, a well-known pathogen of freshwater fish, is the causative bacterium of the disease.

• 한국생물공학회:학술대회논문집
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• 2005.10a
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• pp.899-903
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• 2005

Two infectious species of Streptococcosis pathogens were detected by multiplex PCR assay. Detection rates of Streptococcus iniae and S. parauberis could reach 44.9% and 55.1% respectively for one year during 2004 to 2005 in Jeju island. These findings showed that S. parauberis strains were important pathogen with streptococcosis of olive flounder in Jeju island. These findings showed that S. parauberis strains were important pathogen with streptococcosis of olive flounder in Jeiu island. In the present study we have investigated the interspecific relationship of all Jeju area of S. parauberis by RAPD analysis. Represent strains divided to four groups by RAPD fingerprints. The important differences observed between the olive flounder isolates suggest that they could constitute a well-differentiated group or a separate clonal line within this bacterial species. Though, serological research of S. parauberis strains in Jeju island not exist yet. These strains doing the serological evolution.

• Journal of fish pathology
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• v.22 no.3
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• pp.317-326
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• 2009

This study has shown the screening of anti-bacterial activity of three Indian medicinal plant choloroform : methanol (50:50) solvent leaf extracts (i.e. Azadirachta indica, Ocimum sanctum, and Curcuma longa) with different concentrations (10, 5, 2.5, 1.25, 0.625, 0.312, and 0.156 mg/ml) under in vitro conditions against fish pathogenic bacteria, Aeromonas hydrophila, Streptococcus iniae, Vibrio harveyi, V. anguillarum, and Edwardsiella tarda isolated from olive flounder farms, Jeju Island, South Korea. The anti-microbial activity of the A. indica and O. sanctum extracts yielded the zones of growth inhibition (ZI) was 3 and 1mm against A. hydrophila at concentration of 0.156 mg/ml when compared to that of tetracycline standard (3 mm). At highest concentration (10 mg/ml) of A. indica, O. sanctum, and C. longa, high inhibition was 9, 7, and 6 mm when compared to that of tetracycline (11 mm) against A. hydrophila. The minimum inhibitory concentration (MIC) of A. indica, O. sanctum, and C. longa at 0.156 mg/ml that yield 9, 10, and 13 CFU/ml for A. hydrophila, 16, 22, and 25 CFU/ml for S. iniae and 18, 22, and 23 CFU/ml for E. tarda compared to the tetracycline. At highest concentration (10 mg/ml) of the three extracts was better inhibiting the growth of A. hydrophila, S. iniae and E. tarda. A. indica, O. sanctum, and C. longa were determined to the potential antioxidant activityon the basis of their scavenging activity of the stable 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical. A. indica extract was 0.625 mg/ml which indicated that the strong anti-oxidant activity. However, O. sanctum and C. longa extracts showed weak anti-oxidant activity at this concentration. Hence, in vitro assay among the pathogens, A. hydropila is better inhibitory activity of the extracts. It is evident that the Indian medicinal plants extracts were subjected to its effectiveness against A. hydrophila, S. iniae, and E.tarda at low concentrations. The obtained results in the present study suggested that the Indian plant extracts is a prevention tools for Korean olive flounder aquaculture pathogens and its need further advance investigation.

• Journal of Food Hygiene and Safety
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• v.31 no.5
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• pp.380-385
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• 2016

The object of this study was to evaluate antibacterial activity of edible seaweed extracts against marine bacterial pathogens including Edwardsiella tarda, Streptococcus iniae, Streptococcus parauberis, Vibrio anguillarum, Vibiro harveyi and Vibrio scophthalm, which are associated with human or fish infectious disease. Ecklonia cava methanolic extract showed a strong and broad spectrum antibacterial activity against marine bacterial pathogens used in this study. Among solvent-soluble fractions of the E. cava extract, the ethyl acetate (EtOAc) soluble fraction showed the strongest antibacterial activity against marine bacterial pathogens tested in this study with MIC in the range of $128-256{\mu}g/mL$. Furthermore, HPLC analysis revealed that the soluble fraction contains abundant dieckol, a phlorotannin compound, compared to other solvent soluble fractions, suggesting that phlorotannins including dieckol would be a key antibacterial agent against marine bacterial pathogens.

• Proceedings of the Korean Society of Fisheries Technology Conference
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• 2002.05a
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• pp.515-516
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• 2002