• Plant Disease and Agriculture
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• v.5 no.1
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• pp.27-33
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• 1999

Frost injury of crops is closely related to the epiphytic population dynamics of ice nucleation-active (INA) bacteria, and the injury can be reduced by decreasing the INA bacterial population. In order to predict the epiphytic population of INA bacteria on crops, a rapid and accurate detection method has to be developed. In the previous report, we produced some antibodies against INA proteins purified from the outer membrane of INA bacteria. However it was difficult to produce the antibodies because the purification procedures of the INA proteins were complicated, and the final yield was too low. We designed a specific peptide from the N-terminal region of INA protein by computer analysis and synthesized the peptide in vitro in this experiment. The peptide sequence was Asp-Ser-Por-Leu-Ser-Leu-His-Ala-Asp, that is corresponding to the highly conserved region in several INA proteins, with predicted beta turn, coiling, and hydrophilic region. A polyclonal anti-INA peptide antiserum produced specifically recognized INA bacteria as few as 10 colony-forming units (CFU) in the ELISA reactions and did not respond to other non-INA bacteria. Serological specificity of the anti-INA peptide antiserum will facilitate the forecasting of the INA bacterial population dynamics on crops.

• Korean Journal of Food Science and Technology
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• v.29 no.2
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• pp.326-336
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• 1997

Four ice nucleation-active bacteria (INA-bacteria), Pseudomonas syringae, Xanthomonas campestris, Escherichia coli JM109/pEIN229 and Gluconobacter oxydans/pKIN230, were treated with heat, pressure and gamma-irradiation to compare viability and their ice nucleation activity (INA) after sterilization. Gamma-irradiated INA-bacteria showed the least decrease in T90 value (the temperature at which the 90% of drops are frozen). According to cumulative INA spectra, gamma-irradiated INA-bacteria showed little decrease in class A ice nuclei $(nucleate\;H_{2}O\;at\;higher\;than\;-5^{\circ}C)$, pressurized INA-bacteria showed more than 90% decrease in class A ice nuclei, and heat-treated INA-bacteria barely showed class A ice nuclei. Differential scanning calorimetry (DSC) was used to examine the effect of INA-bacteria on the thermophysical properties of water at freezing temperature. Freezing peaks were appeared at about $11{\sim}15^{\circ}C$ higher on thermograms and enthalpies of phase change were decreased for the water containing INA-bacteria compared with the pure water, while melting peaks were not shifted. INA measured by DSC method were significantly correlated with INA measured by drop freezing method $(R^{2}>0.993,\;p<0.0001)$, indicating that DSC can be used as a new, simple and precise method for measuring INA.

• Journal of Sericultural and Entomological Science
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• v.36 no.1
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• pp.62-68
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• 1994

Isolation, indentification, bioassay and distribution of ice nucleation active(INA) bacteria were done on branch rot of mulberry which was severely developed after harvest of mulberry branches in autumn. Twelve isolates and two isolates out of thirty-six isolates had ice nucleation activity from -5$^{\circ}C$ to -1$0^{\circ}C$, and over -5$^{\circ}C$, respercively. Isolates which formed ice nucleation from -5$^{\circ}C$ to -1$0^{\circ}C$ were not inclined to injure tomato and corn seedlings. However, Two isolates, SE9316 and SE9338 which formed ice nucleation over -5$^{\circ}C$ injured mulberry, tomato and corn seedlings. SE9316 and SE9338 were identified as Pseudomonas syringae based on the morphological, cultural and biochemical characteristics. Populations of ice nucleation active bacteria, fluorecent pseudomonads, were higer in February and April, but they decreased in May. Populations decreased as they become distant from the center of the symptom. The bacterial populations of all sampling times and sites were higer than 105 cfu/g which was enough to induce frost injury.

• 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.

• Journal of Microbiology and Biotechnology
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• v.24 no.3
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• pp.379-385
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• 2014

The purpose of this study was to compare the ability of an engineered Escherichia coli to degrade chlorpyrifos (Cp) using an organophosphorus hydrolase enzyme, encoded in both Flavobacterium sp. ATCC 27551 or Pseudomonas diminuta, by employing the Lpp-OmpA chimera and the N-terminal domain of the ice nucleation protein as anchoring motifs. Tracing of the expression location of the recombinant protein using SDS-PAGE showed the presentation of OPH by both anchors on the outer membrane. This is the first report on the presentation of OPH on the cell surface by Lpp-OmpA under the control of the T7 promoter. The results showed cell growth in the presence of Cp as the sole source of energy, without growth inhibition, and with higher whole-cell activity for both cells harboring plasmids pENVO and pELMO, at approximately 10,342.85 and 10,857.14 U/mg, respectively. Noticeably, the protein displayed by pELMO was lower than the protein displayed by pENVO. It can be concluded that Lpp-OmpA can display less protein, but more functional OPH protein. These results highlight the high potential, of both engineered bacteria, for use in the bioremediation of pesticide-contaminated sources in the environment.

• 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 the Korean Society of Food Science and Nutrition
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• v.23 no.2
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• pp.315-321
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• 1994

In order to prevent the postharvest decay and to promote the freshness retention of fruits and vegetables grapefruit seed extract(GFSE), natural microorganism control agent, was applied to the preservation of fresh fruits and vegetables. Freshfruits and vegetables treated with GFSE and stored in polyethylene film (0.1mm) at 1$0^{\circ}C$-15$^{\circ}C$ of natural low temperature low kept better qualities in color and texture than the GFSE -not- treated control. The treatment using GFSE ina 250ppm to 500ppm concentration seemed to be an effective one for the control of Botrytis cinerea isolated in red wine grapes. After 4 weeks of storage the firmness rate of cucumbers treated with the dilute GFSE was four times higher than that of non-treated ones. GFSE showed effective inhibitory action towards plant pathological bacteria and fungi which were involved in the decay of fruits and vegetables. Minimum inhibitory concentrations of GFSE towards them were in the range of 250ppm to 500ppm .Direct visualization of microbial cells and spores using electron microscopy showed microbial cells and fungal spores the function of which was destroyed by treating with the dilute solutions of GFSE. It was observed that GFSE would reduced disease damages and have bactericide & fungicide properties during the storage of such fruits and vegetables as egg plant, wild edible greens , kumquat, and kiwi fruit.

• Korean Journal Plant Pathology
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• v.12 no.1
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• pp.99-108
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• 1996

3종의 빙핵세균 Peudomonas syringae 8401, Pseudomonas fuorescens 8701, Erwinia herbicola 8701의 세포 외막으로부터 아무런 변성제도 사용치 않고 sucrose density gradient centrifugation, Sephacryl gel filtration chromatography, DEAE-cellulose ion exchange chromatography, non-denaturing buffer를 이용한 PAGE, electroelution, SDS-PAGE를 통해 빙핵활성 단백질을 고도로 정제할 수 있었다. P. suringae와 P. fluorescens에서는 각각 3종류(155 kD, 75 kD, 50 kD)의 빙핵활성 단백질이, E. herbicola에서는 155 kD를 제외한 2종류(75 kD, 50 kD)의 빙핵활성 단백질은 이 연구를 통해 처음 밝혀진 것으로 , 지금까지 보고된 빙핵활성 단백질(150 KD 이상)보다는 훨씬 작은 것이다. 이는 빙핵활성을 나타내는 단백질의 기본단위는 이 실험의 결과만에 의하면 최대 50 kD임을 시사한다. 이들 단백질은 그 유래된 세균의 종류나 또는 단백질 분자량의 크기에 관계없이 모두 -5.5~7.5$^{\circ}C$에서 물을 동결시키는 높은 빙핵활성을 갖고 있었다. 이는 지금까지 보고된 어느 정제단백질의 빙핵활성보다 높은 것이다. 정제된 단백질의 빙핵활성은 trypsin 처리에 의해 상실되었고, pH6~8범위에서는 안정하였으며, pH5이하, pH9이상에서는 활성을 상실하였다. 보존온도에 대한 영향은 3$0^{\circ}C$이상이 되면 점차 활성이 감소하는 경향을 보이다 37$^{\circ}C$이상에서는 활성이 완전히 상실되었다. 금속이온으로서 Hg\ulcorner이온과 SDS에 의해 활성이 상실되었으나 phosphatidylinositol의 첨가에 의해서는 활성이 약간 증가(-1$^{\circ}C$)하였다.

• Journal of Food Hygiene and Safety
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• v.3 no.4
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• pp.233-239
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• 1988

Vegetable mixed codiments, commercial products prepared using soybean,paste and natto, respectively as the ingredients, were used ina study to evaluate the efficacy of gamma irradiationas a means of decontamination and the emphasis was placed upon the determination of the effect of irradiation on the microbiological and some physicochemical properties of the samples. The number of microorganisms contaminated ranged from $10^{6}\;to\;10^{7}$ cells per gram in mesophilic total bacteria, which were composed of thermophilts and acid tolerant bacteria by over 90%. They were reduced by 3 to 4 log cycles with irradiation at 10 kGy. Gamma irradiation at 5 kGy could eliminate the microbial populations of yeasts and molds ($10^{2}\;to\;10^{3}$ cells per gram) and coliforms ($10^{6}\;to\;10^{6}$ cells per gram of natto condiments). However, total destruction of microorganisms in soybean-paste and natto condiments was shown to be possible at a dose-range more than 10 kGy. Irradiationup to 10 kGy was not detrimental to the physicochemical properties of the sample, such as pH. amino nitrogen, rancidity and color, even though some change was brought about in the content of sulfur-containing amino acids.

• Korean Journal of Microbiology
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• v.54 no.2
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• pp.152-154
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• 2018

Klebsiella pneumoniae is a Gram-negative, rod-shape bacterium causing disease in human and animal lungs. K. pneumoniae has been often found to gain antimicrobial resistance, thus it has been difficult to treat K. pneumoniae infection with antibiotics. For such infection, bacteriophage can provide an alternative approach for pathogenic bacterial infection with antimicrobial resistance, because of its sensitivity and specificity to the host bacteria. Bacteriophage KP1 was isolated in sewage and showed specific infectivity to K. pneumoniae. Here, we report the draft genome sequence of Klebsiella pneumoniae phage KP1. The draft genome of KP1 is 167,989 bp long, and the G + C content is 39.6%. The genome has 295 predicted ORFs and 14 tRNA genes. In addition, it encodes various enzymes which involve in lysis of the host cell such as lysozyme and holin.