• Journal of Microbiology and Biotechnology
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• 제22권2호
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• pp.234-238
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• 2012

Recombinant Escherichia coli displaying organophosphorus hydrolase (OPH) was used to overcome the diffusion barrier limitation of organophosphorus pesticides. A new anchor system derived from the N-terminal domain of ice-nucleation protein from Pseudomonas syringae InaV (InaV-N) was used to display OPH onto the surface. The designed sequence was cloned in the vector pET-28a(+) and then was expressed in E. coli. Tracing of the expression location of the recombinant protein using SDS-PAGE showed the presentation of OPH by InaV-N on the outer membrane, and the ability of recombinant E. coli to utilize diazinon as the sole source of energy, without growth inhibition, indicated its significant activity. The location of OPH was detected by comparing the activity of the outer membrane fraction with the inner membrane and cytoplasm fractions. Studies revealed that recombinant E. coli can degrade 50% of 2 mM chlorpyrifos in 2 min. It can be concluded that InaV-N can be used efficiently to display foreign functional protein, and these results highlight the high potential of an engineered bacterium to be used in bioremediation of pesticide-contaminated sources in the environment.

• 공업화학
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• 제25권4호
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• pp.386-391
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• 2014

본 연구에서는 상압 저온 코로나 방전 플라즈마를 화산암재(스코리아) 분말의 살균에 적용하였다. 스코리아 분말에 Escherichia coli (E. coli) 배양액을 살포하여 균일하게 혼합한 후, 코로나 방전 플라즈마 특성 인자인 방전전력, 방전시간, 주입기체, 전극간격 등의 조건을 변화시키며 E. coli 살균효율을 조사하였다. 실험 결과 상압 저온 코로나 방전 플라즈마는 분말상의 스코리아 살균에 아주 효과적인 것으로 나타났으며, 방전전력 15 W에서 5 min 동안 살균한 결과 E. coli가 99.9% 이상 사멸하였다. 방전전력, 방전시간, 인가전압이 증가할수록 사멸율이 향상되었다. 반응기에 주입되는 기체의 종류에 따른 살균력 실험 결과, 산소 > 모사공기(산소 20%) > 질소 순으로 나타났다. 코로나 방전 플라즈마에 의한 E. coli 살균은 자외선과 활성산화종(산소라디칼, OH라디칼, 오존 등)에 의한 세포막 침식 및 에칭, 그리고 플라즈마 방전 스트리머에 의한 대장균 세포막 파괴로 설명할 수 있다.

• Journal of Microbiology and Biotechnology
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• 제25권11호
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• pp.1835-1841
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• 2015

A cell surface display system for heterologous expression of the multifunctional cellulase, CelEx-BR12, in Escherichia coli was developed using truncated E. coli outer membrane protein C (OmpC) as an anchor motif. Cell surface expression of CelEx-BR12 cellulase in E. coli harboring OmpC-fused CelEx-BR12, designated MC4100 (pTOCBR12), was confirmed by fluorescence-activated cell sorting and analysis of outer membrane fractions by western blotting, which verified the expected molecular mass of OmpC-fused CelEx-BR12 (~72 kDa). Functional evidence for exocellulase activity was provided by enzymatic assays of whole cells and outer membrane protein fractions from E. coli MC4100 (pTOCBR12). The stability of E. coli MC4100 (pTOCBR12) cellulase activity was tested by carrying out repeated reaction cycles, which demonstrated the reusability of recombinant cells. Finally, we showed that recombinant E. coli cells displaying the CelEx-BR12 enzyme on the cell surface were capable of growth using carboxymethyl cellulose as the sole carbon source.

• Membrane and Water Treatment
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• 제2권1호
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• pp.25-37
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• 2011

PES UF membranes containing silver were prepared to impart antibacterial properties for waste water treatment. Asymmetric membranes for antibacterial application were prepared from polyethersulfone (PES) and silver nitrate ($AgNO_3$) (PES/$AgNO_3$=15/2 by weight) solution in N-Methyl-2-pyrrolidone (NMP) via simple wet phase inversion technique. These membranes were characterized by polyvinylpyrrolidone (PVP) and polyethylene glycol (PEG) of different molecular weights (1000 ppm in water) at room temperature and on operating pressure of 5 bars. It was observed that the water flux of PES-$AgNO_3$ membrane is slightly lower than virgin PES but still increased linearly with the increment of pressure applied. The morphology of the resulting membranes was examined using Field-Emission Scanning Electron Microscope (FESEM) coupled with Energy Dispersive Spectroscopy (EDS). Elemental analysis using EDS proved that silver is successfully loaded on the membrane surfaces. Due to the success of loading silver on membrane surfaces, antibacterial activities were evaluated via agar diffusion method against Escherichia coli (E.coli) and Staphylococcus aureus (S.aureus) culture. By incorporating 2 wt% of silver nitrate, PES-$AgNO_3$ showed significant inhibition ring on both E.coli and S.aureus. Filtration of E.coli solution (OD 0.31) showed satisfactory rejection data with ~100% inhibition growth after 24 hours incubation at $37^{\circ}C$. Resultant membranes also exhibit better tensile strength (compared to virgin PES) up to 71% may be due to the suggested interactions. The residual silver during fabrication was measured using ICP-MS and result showed that the residual silver content of PES-$AgNO_3$ membrane was only ~1% of the original silver added in the polymer solution. These studies have shown that PES-$AgNO_3$ UF membranes are potential in improving the filtration in water treatment.

• Journal of Microbiology and Biotechnology
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• 제28권11호
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• pp.1782-1790
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• 2018

Assessment of microorganism viability is useful in many industrial fields. A large number of methods associated with the use of fluorescent probes have been developed, including fluorimetry, fluorescence microscopy, and cytometry. In this study, a microvolume spectrofluorometer was used to measure the membrane potential variations of Escherichia coli. In order to estimate the sensitivity of the device, the membrane potential of E. coli was artificially disrupted using an ionophore agent: carbonyl cyanide 3-chlorophenylhydrazone. The membrane potential was evaluated using two ratiometric methods: a Rhodamine 123/4',6-diamidino-2-phenylindole combination and a JC-10 ratiometric probe. These methods were used to study the impact of freezing on E. coli, and were compared with the conventional enumeration method. The results showed that it was beneficial to use this compact, easy-to-use, and inexpensive spectrofluorometer to assess the viability of bacterial cells via their membrane potential.

• 한국수산과학회지
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• 제26권3호
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• pp.205-213
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• 1993

APHA-MPA(American Public Health Association-Most Probable Numbers)은 환경수중에 존재하는 위생지표세균 추정에 사용되는 미국 FDA의 유일한 공인방법이기는 하나 결과를 얻기까지 많은 시간, 인력 및 경비가 소요될 뿐 아니라 일반적으로 정밀도도 낮다는 결점이 있다. 이러한 문제를 극복하기 위하여 개발된 막여과 방법중 mTEC(membrane thermotolerant E. coli)은 분변계대장균 및 E. coli의 회수율이 좋을 뿐 아니라 경비도 절약할 수 있는 방법으로서 알려져 있어, 미국 Rhode Island주의 Greenwich Bay의 위생조사를 통하여 MPN과 비교하여 보았다. 그 결과 mTEC의 분변계 대장균 및 E. coli 회수율은 MPN보다 평균치로써 각각 1.08 및 1.27배 높게 나타나, MPN의 대체 방법으로서의 가능성이 확인되었으며, 따라서 이 방법이 앞으로 위생지표세균의 검출방법으로서 공인된다면 우리나라 수출용 패류생산 지정해역의 관리는 물론 패류양식장의 위생학적인 분류도 효율적으로 할 수 있으리라 기대된다.

• 한국식품저장유통학회지
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• 제7권1호
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• pp.124-131
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• 2000

Antibacterial activities of powdered spices(garlic , ginger, cinnamon and clove) against pathogenic Escherichia coli )157:H7 and Staphyloccus auresus were investigated. Spice powder was added in was exponetial phase of each bacterial culture . Growth inhibition was determined by the absorbance at 660nm and morphological changes of the cells were observed by transmission electron microscope (TEM). Ginger powder has the highest antibacterial activity, following cinnamon , clove and garlic has the least activity.Growth of Escherichia coli O157:H7 and Staphyloccus aureus were completely inhibited within 5 hours after addition of 1 % of garlic , 0.3% of ginger or cinnamon , 0.5% of clove powder on the exponential phase of the cells. Spice untreated cells of E. coli and S. aureus, the cytoplasm was entirely surrounded by rigid cell wall and cell walls formed a smooth layer well attached to the plasma membrane. In the cells of E. coli and S. aureus treated with spice powder, cell wall and plasma membrane were lysed and severely damaged. E.coli cells growth in the presence of spice powder showed plammolysis, the loss of electron dense material, the formation of extra cellular blebs and cytoplasm burst out from the cell. S .sureus cells grown in the presence of spice powder showed swell of cell wall, the loss of electron dense material , coagulation of cell cytoplasm and formation of extra cellular blebs. Severely damaged cells of S. aureus lost whole cytoplasm and left as ghost of the cell. Spice powder stimulated autolyssi and induced cell death.

• Journal of Microbiology and Biotechnology
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• 제30권10호
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• pp.1552-1558
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• 2020

With an increase in the consumption of non-heated fresh food, foodborne shiga toxin-producing Escherichia coli (STEC) has emerged as one of the most problematic pathogens worldwide. Endolysin, a bacteriophage-derived lysis protein, is able to lyse the target bacteria without any special resistance, and thus has been garnering interest as a powerful antimicrobial agent. In this study, rV5-like phage endolysin targeting E. coli O157:H7, named as LysECP26, was identified and purified. This endolysin had a lysozyme-like catalytic domain, but differed markedly from the sequence of lambda phage endolysin. LysECP26 exhibited strong activity with a broad lytic spectrum against various gram-negative strains (29/29) and was relatively stable at a broad temperature range (4℃-55℃). The optimum temperature and pH ranges of LysECP26 were identified at 37℃-42℃ and pH 7-8, respectively. NaCl supplementation did not affect the lytic activity. Although LysECP26 was limited in that it could not pass the outer membrane, E. coli O157: H7 could be effectively controlled by adding ethylenediaminetetraacetic acid (EDTA) and citric acid (1.44 and 1.14 log CFU/ml) within 30 min. Therefore, LysECP26 may serve as an effective biocontrol agent for gram-negative pathogens, including E. coli O157:H7.

• Journal of Biosystems Engineering
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• 제38권4호
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• pp.335-340
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• 2013

Purpose: This study was performed to develop a lateral flow strip sensor for the detection of pathogenic Escherichia coli O157:H7 in various samples. Also, feasibility of using an image analysis method to improve the interpretation of the strip sensor was evaluated. Methods: The lateral flow strip sensor has been fabricated based on nitrocellulose lateral-flow membrane. Colloidal gold and E. coli O157:H7 antibodies were used as a tag and a receptor, respectively. Manually spotted E. coli O157:H7 antibody and anti-mouse antibody on nitrocellulose membrane were used as test and control dots, respectively. Feasibility of the lateral flow strip sensor to detect E. coli O157:H7 were evaluated with serially diluted E. coli O157:H7 cells in PBS or food samples. Test results of the lateral flow strip sensor were measured with an image analysis method. Results: The intensity of the test dot started to increase with higher concentration of the cells were introduced. The sensitivities of the sensor were both $10^4$ CFU/mL Escherichia coli O157:H7 spiked in PBS and in chicken meat extract, respectively. Conclusions: The lateral flow strip sensor and image analysis method could detect E. coli O157:H7 in 20 min, which is significantly quicker than conventional plate counting method.

• Archives of Pharmacal Research
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• 제19권5호
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• pp.353-358
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• 1996

E. coli 11 and E. coli 101, clinical isolates of Escherichia coli were resistant to various quinolones, especially MICs to norfloxacin of both strains were higher than 100 mg/ml. In the presence of carbonyl cyanide m-chlorophenylhydrazone, a proton gradient uncoupler, norfloxacin uptake in both strains was increased, suggesting that an efflux system play an important role in the norfloxacin resistance. Outer membrane proteins of the susceptible and resistant strains which could affect the route of norfloxacin entry into cells were different. When quinolone resistance determining region(QRDR) of gyrA was amplified using PCR and cut with Hinf I, QRDR in the susceptible strain yielded two fragments while QRDRs in E. coli 11 and E. coli 101 yielded only one uncut fragment. When DNA sequence of QRDR was analyzed, there were two mutations as Ser-83 and Asp-87 in both resistant strains. these residues were changed to Leu-83 and Asn-87, respectively. These results showed that the norfloxacin resistance of E. coli 11 and E. coli 101 was resulted from multiple changes-an altered DNA gyrase A subunit, a change in route of drug entry, and reduction in quinolone concentration inside cells due to an efflux system.