• Journal of Microbiology
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• v.43 no.5
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• pp.437-442
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• 2005

Sera from rabbits were infected with Vibrio vulnificus containing an antibody against major outer membrane protein (MOMP). MOMP of V. vulnificus ATCC 27562 were isolated and purified by Sarkosyl and TritonX-100 dual treatment. Molecular size of MOMP was identified as 36-kDa on $13\%$ SDS-PAGE. The sequence of the first 26 amino acid residues from the N-terminal end of the protein is AELYNQDGTSLDMGGRAEARLSMKDG, which is a perfect match with OmpU of V. vulnificus CMCP6 and YJ016. MOMP specific IgM and IgG were investigated in groups of mice. The group of mice immunized with MOMP and Alum showed higher levels of IgG2b than the group immunized with only MOMP. Vaccination with MOMP resulted in protective antibodies in the mouse infection experiment.

• Journal of Microbiology
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• v.41 no.4
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• pp.320-326
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• 2003

In order to identify Vibrio vulnificus in the Yellow Sea near Gunsan, Korea during the early and late summers, the efficiency of the real-time quantitative TaqMan PCR was compared to the efficiency of the conventional PCR and Biolog identification system^TM. Primers and a probe were designed from the hemolysin/cytolysin gene sequence of V. vulnificus strains. The number of positive detections by real-time quantitative TaqMan PCR, conventional PCR, and the Biolog identification system from seawater were 53 (36.8%), 36 (25%), and 10 strains (6.9%), respectively, among 144 samples collected from Yellow Sea near Gunsan, Korea. Thus, the detection method of the real-time quantitative TaqMan PCR assay was more effective in terms of accuracy than that of the conventional PCR and Biolog system. Therefore, our results showed that the real-time TaqMan probe and the primer set developed in this study can be applied successfully as a rapid screening tool for the detection of V. vulnificus.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.40 no.5
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• pp.293-299
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• 2007

The distribution of hemolytic Vibrio sp. from sea water of three main beaches located in Busan (Gwangan(G), Haeundae(H) and Songjeong(S) beaches) was investigated from June to September 2006 ; this is mid-summer and the main season for bathing. The monthly detection ratio from each beach was 29.2% (7 of 24 samples, G), 33.3% (8 of 24 samples, H), and 16.7% (4 of 24 samples, S). The most probable number(MPN) of strains detected ranged from 1.8-36(G), 1.8-180(H) and 1.8-18(S) MPN/100mL. Of the isolated strains, 24 strains showed definite hemolytic activity. These 24 strains were identified as Vibrio fluvialis, Vibrio vulnificus, Aeromonas hydrophila, Actinobacillus ureae and Eikenella corrodens. Vibrio fluvialis was detected from all three beaches investigated. Vibrio vulnificus was detected from Haeundae and Gwangan beaches. Gwangan beach had a higher detection ratio of Vibrio sp. than Haeundae and Songjeong beaches. These results suggest that seafood harvested from the vicinity of theses beaches may cause food poisoning and risk management to prevent Vibrio septicemia is required, especially for Haeundae and Gwangan beaches.

• Journal of fish pathology
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• v.35 no.1
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• pp.57-63
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• 2022

Vibrio parahaemolyticus and V. vulnificus are known to be infected to human via fisheries products. Therefore, food safety of fisheries products is important for public health and fish industry. This paper was conducted to know how well these human isolates can survive in olive flounder (Paralichthys olivaceus). The growth of V. parahaemolyticus and V. vulnificus showed about 50~60% reduced rates at 25℃ than at 37℃ and did not show any differences according to NaCl concentration of media except the increasing in the growth of V. vulnificus in medium containing 3% NaCl. Artificial infection of 1×10⁶ CFU/fish was carried out to confirm the sensitivity of olive flounder against V. parahaemolyticus and V. vulnificus. After 1 week from injection, no fish was dead. To evaluate nonspecific defense of olive flounder against V. parahaemolyticus and V. vulnificus, the antibacterial potency of serum and epidermal mucus were tested. The number of the vibrios exposed to serum obtained from olive flounder significantly decreased after 3 hours, and epidermal mucus showed decrease of the bacteria over than 90% until 12 hours from exposure. Phagocytosis of head kidney leucocytes of healthy olive flounder against V. parahaemolyticus and V. vulnificus showed in over 70% of leucocytes at the 2 hours. Therefore, cultured olive flounder only as vehicle for human pathogen in environmental water is well developed its antibacterial potency against human pathogens, so the viability of V. parahaemolyticus and V. vulnificus in cultured olive flounder was considered very low.

• Korean Journal of Microbiology
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• v.38 no.4
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• pp.281-286
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• 2002

Detection of Vibrio vulnificus in fish farm and searching for the bactericidal methods on this bacteria were studied. To detect this microorganism in sea water, mud, fish and mussels, selective isolation methods and detection of vvhA gene were used from January to October,2000. V. vulnificus was detected from May when the water temperature was over $17^{\circ}C$. From June to September, higher than $19^{\circ}C$, this bacteria could be isolated from most of the samples. Freezing and refrigerating did not inhibit the growth of V. vulnificus. Citric acid did not show the bactericidal effect, but more than 500 mg/l of EDTA did. With the aid of UV and photocatalyst, $TiO_{2}$ showed bactericidal effect after 15 minute treatment. Photocatalytic system consisted of glass bead coated with $TiO_{2}$ and UV illumination showed bactericidal effect on V. vulnificus at the turnover rate of 0.2/min.

• Journal of Life Science
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• v.14 no.6 s.67
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• pp.986-990
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• 2004

Five strains of Vibrio vulnificus, which cause serious septicemia in human worldwidely, was isolated from marine environments of Korea costal area from May to July of 2002. The isolated strains were identified by API 20E kit and partial 16S rDNA sequence analysis. 16S rDNA sequence of the isolates showed $99\%$ similarity with V. vulnificus ATCC 27562. The proteins of V. vulnificus isolates were examined by analyzing patterns of the cell lysates and outer membrane proteins (OMP). The OMP separated from cell lysates showed the common protein band. Therefore OMP profiles might be useful for the identification of V. vulnificus sp.

• Journal of fish pathology
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• v.12 no.2
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• pp.89-99
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• 1999

New sixteen heat shock proteins (Hsps) and ten ethanol shock proteins were appeared on the analysis with SDS-PAGE when cultivation temperature for the Vibrio vulnifrcus ATCC 27562 strain was shifted-up to $42^{\circ}C$ from $30^{\circ}C$ for 20 mins and treated with of 6% ethanol for 10 mins, respectively. Even the induction of thermotolerance in V. vulnificus was coincided with the induction of Hsps if the pre-shock was adjusted to thermal temperature. Outer membrane proteins (OMPs) that were purified from the membrane of cells after heat shock showed more immunodominant pattern to the immunized rabbit anti-V. vulnificus O serum in enzyme-linked immunosorbent assay (ELISA). On the western immunoblot analysis it was confirmed that both 62 kDa IMP and 69 kDa OMP in the Hsps and 48 kDa IMP a major OMP in the ethanol shock proteins were reacted with rabbit anti-V. vulnificus O sera. Agglutination titer of the heat shocked V. vulnificus with rabbit anti-V. vulnificus O serum was higher than that of the untreated bacteria.

• Journal of Microbiology and Biotechnology
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• v.14 no.2
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• pp.350-355
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• 2004

Sulforhodamine B (SRB) assay is a rapid, sensitive, and inexpensive method for measuring cell proliferation and chemosensitivity. However, the lactate dehydrogenase (LDH) release assay is generally used to measure cytototoxicity of infectious microorganisms against host cells. In this study, we investigated the possibility of applying the SRB assay to determine cytotoxicity for infectious microorganisms, and compared the results with those obtained by the LDH release assay. We used Vibrio vulnificus as a model of infectious microorganisms. The SRB assay showed that V vulnificus strongly induced cytotoxic activity against human intestinal cells, Caco-2 and INT-407 cells. The degree of cytotoxicity closely correlated with infection time and number ratios of V. vulnificus to intestinal cells (MOI, multiplicity of infection). Furthermore, cytotoxicity values obtained by SRB assay correlated well with results obtained by the LDH release assay, and both assays gave a linear response with respect to MOI Heat-inactivation of V. vulnificus for 35 min at $60^{\circ}C$ did not induce cytotoxic activity, indicating that viability of V. vulnificus is crucial for cytotoxic activity against intestinal cells. Although both assays are suitable as cytotoxicity endpoints, the SRB assay is recommended for measuring cytotoxicity of infectious microorganisms against host cells because of its significantly lower cost and more stable endpoint than the LDH release assay.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.36 no.3
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• pp.239-246
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• 2003

We have examined the 16S-23S rRNA intergenic spacer region (ISR) of Vibrio vulnificus KCTC 2959. ISRs were amplified by primers complementary to conserved regions of 16S and 23S rRNA genes. ISR amplicons were cloned and sequenced. Analysis of the ISR sequences showed that V. vulnificus KCTC 2959 contains five types of polymorphic ISRs. Size of ISRs ranged from 424 to 741 bp in length and the number of tRNA genes ranged from one to four. The ISRs were designated as ISR-E $(tRNA^{Glu}),\;ISR-IA\;(tRNA^{Ile}-tRNA^{Ala})$, ISR-EKV $(tRNA^{Glu}-tRNA^{Lys}-tRNA^{Val})$, ISR-IAV $(tRNA^{Ile}-tRNA^{Ala}-tRNA^{val})$ and ISR-EKAV $(tRNA^{Glu}-tRNA^{Lys}-tRNA^{Ala}-tRNA^{Val})$ based on their tRNA genes. Multiple alignment of representative sequences from different Vibrio species revealed several domains of high sequence variability. We used the sequences of variable domains to design species-specific primer for detection PCR. Specificity of the primers was examined using genomic DNA prepared from 18 different Vibrio species. The results showed that the PCR using primers designed in this study can be used to detect V. vulnificus from other Vibrio species.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.21 no.2
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• pp.80-84
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• 1988

The change of cell counts of Vibrio vulnificus in meat homogenates of fish and shellfish by the storage time and temperature was examined to get basic information for precautionary steps against septicemia from slices of raw fish (sashimi). Therefore, we inoculated raw and cooked meat homogenates of fish and shellfish with Vibrio vulificus M-8 (isolated from shellfish ) and stored them at $-20^{\circ}C,\;4^{\circ}C\;and\;30^{\circ}C$ for 72 hours. Vibrio vulnificus M-8 was not detected in 32 hours when it was frozen and stored at $-20^{\circ}C$ after inoculating them into phosphate buffer solution at concentration of $10^5\;cell/ml$, while the existance of Vibrio vulnificus was identified after 72 hours of storage at the same temperature in case of inoculation into the meat homogenate of yellow tail. The cell count of Vibrio vulnificus was decreased as about $20\%$ of initial count after 2 hours storage at $4^{\circ}C$ in phosphate buffer solution with fish and shellfish homgenates. From the experimental results it was recognized that Vibrio vulnificus was labile to the cold stress. In comparison to the growth of growth of Vibrio M-8 at $30^{\circ}C$ in the raw and cooked meat of the yellow tail(Seriola guingueradita), snapper(Chrysophrys major), ark shell(Anadra brouhgtonii), and oyster(Crassostrea gigas), the raw meat homogenates were more excellent than the cooked ones though all fish and shellfish meat homogenates were proves to be good for the growth of the microbe.