• The Journal of the Korean Society for Microbiology
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• v.22 no.1
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• pp.23-33
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• 1987

To detect lepotospiral strains which produce hemolysin and determine the optimal condition for assaying hemolysin, we screened reference strains and observed some property of hemolysin. Hemolysin activity was assayed with cell free culture liquids and erythrocyte suspension. The production of hemolysin by local strains isolated in Korea was assayed and compared with that of reference strains. The hemolysin was produced by 18 strains among 38 reference strains and 3 local strains isolated in Korea. The production of hemolysin began with growth of Leptospira cultured in EMJH medium and reached maximum at stationary phase. The optimum temperature for hemolytic activity was $37^{\circ}C$. At lower temperature the activity of hemolysin was decreased progressively. The hemolytic activity was completely inactivated after :30 minutes' exposure at $56^{\circ}C$. Hemolysis pattern was "hot-cold type" which showed increased hemolysis after cold incubation. The hemolysin was most active on sheep erythrocyte and less active on ox, goat, human and guinea pig erythrocyte with the decreasing order.

• Journal of Environmental Science International
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• v.6 no.3
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• pp.225-229
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• 1997

A halophilic V. vulnificus is an estuarine microorganism that has been associated with fatal wound Infection and life-threatening septicemia. Hemolysin is defined as toxic substance produced by various species of bacteria Including V. vulnificus. Hemolysin from marine V. vulnificus was purified and the effect of pH, temperature. metal ion on the activity of hemolysin, and thermostability of hemolysin were tested in this study. Hemolysin iysed the sheep red blood cell and the optimum pH was 8.0, the optimum temperature was 4$0^{\circ}C$, and $K^+$ increased but $Mn^{2+}$ decreased the hemolyic activity of hemolysin, but hemolysin was unstable to heat.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.30 no.4
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• pp.556-561
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• 1997

Vibrio cholerae non-O1 FM-3 was isolated from sea water, and it showed the same bacteriological characteristics as Vibrio cholerae non-O1 ATCC 25872. V. cholerae non-O1 FM-3 presented the highest hemolytic activity at stationary phase of its growth. The hemolytic activity was decreased in accordance with increasing of pretense activity of its cultural supernatant. The characteristics of the hemolysin produced by V. cholerae non-O1 FM-3 were investigated after partial purification with a Sephadex G-100 chromatography. The hemolytic activity of purified protein was stable at $4^{\circ}C$ while it was completely lost by heating at $60^{\circ}C$ for 30 min. The activity of hemolysin was increased by addition of divalent cations such as $Ca^{2+},\;Mg^{2+},\;and\;Mn^{2+}$ while it was inhibited by additions of $Zn^{2+}$. When the hemolysin was incubated with suspensions of erythrocytes at $4^{\circ}C$ and $37^{\circ}C$, respectively, hemolysis was not observed at $4^{\circ}C$ but at $37^{\circ}C$. It means that hemolysis by purified hemolysin was temperature-dependent while its binding step of hemolysin to cell membrane was temperature-independent.

• Microbiology and Biotechnology Letters
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• v.19 no.3
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• pp.303-307
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• 1991

The hemolyic polypeptide in delta-endotoxin from Bacillus thun'ngiensis subsp. darmstadiensis 73ElO-2 was purified by Sephadex G-IOO gel filtration and DEAE-cellulose ion exchange column chromatography. The purity of hemolysin was confirmed by ouchterlony test and SDS-PAGE. The molecular weight of the purified hemolysin was approximately 64 KDa by SDS-PAGE. The purified hemolysin has not mosquitocidal activity against larvae of Aedes agypti, but hemolytic activity on red blood cells of rat. There is no serological relationship between delta-endotoxin from B. thuringiensis subsp. israelensis and the purified hemolysin from the . strain 73ElO-2.

• Journal of Life Science
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• v.8 no.2
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• pp.145-157
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• 1998

We isolated 100 Vobrio sp. from marine products and sea from July to September, 1997. We attemped on purification of hemolysin produced by Vibrio sp. The growth, hemolysin production patterns by the 100 strains of Vibrio sp. showed identical, in general. V. unlnificus ys-1 produced hemolysis as the higtest titer. The optimal culture conditions for the hemolysin production by the V. vunificus ys-1 are followings; 1. Hemolysin production was optimal dering the late exponetial phage. 2. Maximal growth, hemolysin production were in heart infusion broth. 3. Maximal yields of hemolysin was obtained when the heart infusion broth had an intial pH of 8.0, 3$0^{\circ}C$, 3% NaCL. Hemolysin was purified from culture filtrate of the strain by ammonium sulfate recipitation, ion exchange and hydrophobic interaction chromatography. The results were as follows; 1. Hemogeneity of the purified hemolysin was demonstrated by revealing single band on SDS-PAGE. The molecular weight of purified hemolysin was 45KDa. 2. The absorbance rattern in ultraviolet wsa typical of those seen with most proteinb with 280nm. 3. Purified hemolysin was atable at 5$0^{\circ}C$ but 7$0^{\circ}C$ of the acivity was lost by heating for 30 min at 6$0^{\circ}C$/ Optimal temperature of purified hemolysin was 35$^{\circ}C$. 4. Purified hemolysin was stable at the pH range of 6~9, but in the less the pH5.0. above the pH 9.0, the hemolysin activity was lost completely.

• Journal of Life Science
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• v.12 no.4
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• pp.490-495
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• 2002

Hemolysin (VFH) of V. fluvialis, which is a pathogenic bacteria, causing watery diarrhea with vomiting, abdominal croup, was purified. V. fluvialis was cultivated in BHI medium and the culture supematant was precipitated by ammonium sulfate. The protein was purified by chromatographies on columns of DEAE-cellulose and Mono-Q. Molecular weight of the purified VFH was estimated as 79kDa by SDS-PAGE. The optimal temperature for a maximum hemolytic activity was at around 35$^{\circ}C$ and the activity was decreased at 4$0^{\circ}C$ Cytotoxicity of VFH was also investigated using RTG-2 cell line. LDH assay study showed that 50$\mu\textrm{g}$/m1 of VFH release 80% of total cellular LDH (lactate dehydrogenase) from RTG-2 cell and microscopic observation also showed the morphological change of cell.

• Microbiology and Biotechnology Letters
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• v.15 no.6
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• pp.425-429
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• 1987

The extra-cellular hemolysin from Bacillus thuringiensis subsp. israelensis was purified in the process of suiting with (NH$_4$)$_2$SO$_4$, Sephadex G-200 gel filtration, and DEAE-cellulose column chromatography. The purified hemolysin had molecular weight of approximately 47,000 dalton on SDS-polyacrylamide gel electrophoresis. The activity of purified hemolysin on human red blood cells was increased by thiol agents, but that was Inhibited by cholesterol, protease treatment, and metal salts such as CuSO$_4$, and FeSO$_4$, respectively.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.41 no.4
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• pp.243-247
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• 2008

The hemolysin of Vibrio cholerae non-O1 CT isolated from sea water was purified and characterized. The purified hemolysin displayed an optimum at $37^{\circ}C$ and exhibited more than 70% of residual hemolytic activity after incubation at $45^{\circ}C$ for 120 min. However, the activity dropped dramatically at temperature above $55^{\circ}C$. The purified protein showed the highest hemolytic activity at pH 7.0, while the activity was completely lost outside of the pH ranges of 5.0 and 10.0. The activity of hemolysin was inactivated by addition of divalent cations, such as $Cu^{2+},\;Fe^{2+},\;Hg^{2+},\;Mn^{2+},\;and\;Zn^{2+}$, however, the activity was not completely inhibited by additions of $Ca^{2+},\;Mg^{2+},\;K^+,\;Na^+,\;and\;Li^+$.

• Journal of Microbiology and Biotechnology
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• v.13 no.6
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• pp.1021-1026
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• 2003

Numerous secreted and cell-associated virulence factors have been proposed to account for the fulminating and destructive nature of Vibrio vulnificus infections. Among the putative virulence factors are an elastase, elastolytic protease, and a cytolytic hemolysin. Effects of the elastase on the hemolysin were assessed by evaluating changes of hemolytic activities either in the presence or absence of the protease. Although hemolytic activity in the culture supernatant was lowered by the purified elastase added in vitro, the cellular level of hemolytic activity was unaffected by the mutation of vvpE encoding the elastase. Growth kinetic studies revealed that hemolysin reached its maximum level in the exponential phase of growth, and the elastase appeared at the onset of the stationary phase. These results have provided insight into the regulation of virulence factors: temporally coordinate regulation of virulence factors is essential for the overall success of the pathogen during pathogenesis.

• Journal of Microbiology and Biotechnology
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• v.14 no.2
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• pp.330-336
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• 2004

Fifty-two pathogenic Vibrio parahaemolyticus strains were isolated from the environments of Busan and Yeosu, Korea. Forty-three of these strains showed protease activities, whereas 4 strains showed $\alpha / \beta$ hemolysin activities and 6 strains had urease activities. Their pathogenic factors were not overlapping except one strain, which had both protease and hemolysin activities. The 6 urease-positive strains (V. parahaemolyticus YKB4, YKB14, S25, YFB20, YFO21, and YFO22) showed the same biochemical characteristics as a reference strain [V. parahaemolyticus KCTC 2471 (urease-negative)], except for urease production. The 6 urease-positive strains showed different urease activities in their culture supernatant during the growth. The urease activity of S25 increased sharply at the late exponential phase, and was the highest at the initial stationary phase and was kept until the late stationary phase. The other 5 isolates, except C25, showed urease activities at the mid-stationary phase and increased steadily until the late stationary phase, when the urease activity was maximal. To compare the degree of virulence of V. parahaemolyticus with different pathogenic factors, hemolysin, protease, or urease-positive strains were injected into groups of 10 each of ICR mice (7- to l0-week-old males). The lethal rates of urease-positive V. parahaemolyticus, YKB14, YKB4, and S25, were significantly high, being 50, 70, and 80%, respectively. Protease-positive V. parahaemolyticus strains FM39 and FM50 showed 40% and 60% of lethal rate, respectively. Hemolysin-positive V. parahaemolyticus strains S34 and S72 had no mortality, similar to nonpathogenic V. parahaemolyticus FM12.